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+{"text": "P=0.015), T stage (P=0.006), lymph node metastasis (P=0.028) and the worst prognosis of PC patients (P=0.004). Meanwhile, a positive relationship between FAM172A and E-cadherin (E-cad)  was observed in clinical samples, which contributed to the better prognosis of PC patients (P=0.014). FAM172A silencing induced EMT in both AsPC-1 and BxPC-3 cells, including inducing the increase of Vimentin, MMP9 and pERK and the decrease of E-cad and \u03b2-catenin expression, stimulating EMT-like cell morphology and enhancing cell invasion and migration in PC cells. However, MEK1 inhibitor PD98059 reversed FAM172A silencing-enhanced EMT in PC cells. We conclude that FAM172A inhibits EMT of PC cells via ERK-MAPK signaling.FAM172A, as a newly discovered gene, is little known in cancer development, especially in pancreatic cancer (PC). We investigated the potential role and molecular mechanism of FAM172A in epithelial to mesenchymal transition (EMT) in both human clinical samples and PC cells. FAM172A was downregulated in human PC tissues compared with that in non-cancerous pancreas cells by immunohistochemistry and qRT-PCR. FAM172A expression was negatively associated with tumor size ( Summary: FAM172A inhibits EMT in pancreatic cancer via specifically regulating ERK-MAPK signaling. Pancreatic cancer (PC), as a deadly malignancy, ranks as the fourth leading cancer-related cause of death in Europe . It is aFamily with sequence similarity 172 member A (FAM172A), cloned from human aortic tissues, is subsequently observed in human endothelial and vascular smooth muscle cells, and macrophages . HoweverThe ERK-MAPK signaling plays a significant role in regulating cell proliferation, differentiation, survival, migration senescence and apoptosis via transmitting signals from cell surface receptors . ERK-MAPP<0.01) (P=0.002)  A. In mosP<0.01) B, and viP<0.01) C. SpearmP=0.002) .Fig. 1.P=0.015), T stage (P=0.006), lymph node metastasis (P=0.028), but had no relationship with age, gender, tumor location, differentiation, vascular invasion and preoperative CA199 level in PC patients (P>0.05) (P=0.003) (P=0.159) (P=0.014) (P=0.020) (Chi-squared test showed that FAM172A was negatively associated with tumor size ((P>0.05) . PatientP=0.003) D. ThoughP=0.159) E, patienP=0.014) F. In mulP=0.020) .Table\u00a02.P=0.002)  A. Both FP=0.002) B,C. The in vitro.AsPC-1 and BxPC-3 cells with high FAM172A expression were used for FAM172A silencing experiments. FAM172A protein expression in the FAM172AsiRNA group was much lower than that in the siRNAcontrol group in both cell lines . FAM172APD98059 was identified as a highly selective inhibitor of MEK1 activation and the MAP kinase cascade . PD98059In addition, FAM172AsiRNA transfected AsPC-1 and BxPC-3 cells presented EMT-like cell morphology: most cells lost their epithelial properties of tight junction, and presented a spindle-shaped and fibroblast-like morphology . Howeverin vitro  was significantly reversed by MEK1 inhibitor PD98059. PD98059 also inhibited FAM172A silencing-induced EMT like cell morphology and cell invasiveness. Previous studies have shown that FAM172A promotes the proliferation of papillary thyroid carcinoma cells via p38/MAPK signaling , which were cultured in the recommended growth media with 10% FBS .IHC was performed as described previously ,b. BriefAs described previously , whole-cell lysates of PC cells were put into 12% SDS-polyacrylamide gels, transferred to PVDF membrane  and incubated with primary FAM172A , E-cadherin , \u03b2-catenin , N-cadherin , Vimentin , MMP9 , and \u03b1-Smooth muscle actin , pERK , ERK  and GAPDH  antibodies overnight. Then membranes were incubated with secondary antibodies (Proteintech) at room temperature and were visualized with the ECL machine . PC cells were pretreated with 20\u2005\u00b5M of MEK1 inhibitor PD98059  for 2\u2005h before western blotting. The experiment was repeated three times.As described previously , qRT-PCRThe FAM172AsiRNA sequences were: sense: 5\u2032-GCCACTGAGAGTGAACCAAAG -3\u2032, which were synthesized from GenePharma company . siRNA transfections (20\u2005\u00b5M) were mixed with Oligofectamine 3000  for transient transfection following the manufacturer\u2019s instructions.As described previously , FAM172ABriefly, FAM172AsiRNA and siRNAcontrol transfected AsPC-1 and BxPC-3 cells (pretreated with 20\u2005\u00b5M of PD98059 for 2\u2005h only once) was seeded onto 8.0-\u00b5M pore size membrane inserts  coated with matrigel  in 24-well plates with FBS-free growth media. Growth media plus 10% FBS was added to the bottom wells as a chemoattractant. 24\u2005h later, cells that moved to the underside of the inserts were stained with Crystal Violet Hydrate . The migratory cells were counted in five random fields per well. Results were expressed as cells migrated per field and repeated three times. The transwell assay was performed in a similar way without matrigel.in vitro were described as means\u00b1s.e. (standard deviation) and were compared via t-test. P-value is presented as follow: *P<0.05; **P<0.01.Under SPSS software 20.0 , paired nonparametric test, chi-squared test and spearman correlation test were used for IHC assays, respectively. The log-rank test and Cox's regression was used to evaluate the postoperative survival time of PC patients. Western blotting, qRT-PCR and transwell assays"}
+{"text": "The present study evaluates the protective effects of myricitrin and its solid lipid nanoparticle (SLN) on diabetic nephropathy (DN) induced by streptozotocin-nicotinamide (STZ-NA) in mice. In this experimental study, 108 adult male NMRI mice were divided into 9 groups: control, vehicle, diabetes, diabetes + myricitrin 1, 3, and 10 mg/kg and, diabetes + SLN containing myricitrin 1, 3, and 10 mg/kg. After the experimental period, the plasma and tissue samples were collected for experimental, histopathological, real-time PCR and apoptosis assessments. Total antioxidant capacity, catalase, glomerular filtration rate, plasma level of albumin, urine (BUN) and, creatinine (Cr) levels decreased, and the kidney weight, intake/output, malondialdehyde, plasma level of BUN and Cr, urine level of sodium, potassium, albumin and glucose, fractional excretions of sodium and potassium, transforming growth factor-\u03b2 (TGF-\u03b2) and nuclear factor kappa B (NF-\u03baB) gene expression, red blood cell accumulation and infiltration of inflammatory cells, and kidney apoptosis increased in untreated diabetic mice compared to the control group, and administration of myricitrin and its SLN recovered all of these changes. Ultimately, myricitrin and its SLN administration improved DN changes by reducing oxidative stress and increasing antioxidant enzymes level, and these effects were more prominent in the SLN-administered mice. Myrica cerifera (Diabetic nephropathy (DN), known as a kidney progressive disease is characterized by persistent albuminuria, progressive decrease in glomerular filtration rate (GFR) and increases in plasma level of creatinine (Cr). This disease occurs in 45 % of diabetic patients . The rencerifera . Oral bicerifera . Solid lcerifera . Streptocerifera . Howevercerifera . So, basChemicals and experimental kitsUnited Kingdom) assay kits, blood urea nitrogen (BUN), Cr, albumin, glucose assay kits , RNeasy Mini Kit , cDNA Synthesis Kit, Sybergreen , TGF-\u03b2 and NF-\u03bab, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) primers , TUNEL assay kit , Proteinase K .Myricitrin (Purity 98 %) , Citrate buffer, Tween 80 , Streptozotocin , Nicotinamide, Hematoxylin, Eosin, , Ketamine/Xylazine , phosphate-buffered saline (PBS) , Iran), malondiadehyde (MDA), total oxidant capacity (TAC), catalase (CAT) , superoxide dismutase (SOD)  and encapsulation efficiency (EE %) were assessed in that study .Animalsad libitum. After one-week acclimatization of mice, T2DM induced by intraperitoneal injection of STZ (65 mg/kg) and NA (120 mg/kg) (dissolved in a citrate buffer (pH: 4.5) and normal saline, respectively) with 15 min interval. Mice with fasting blood glucose above 200 mg/dl were considered as diabetic and entered the study at 3 days after STZ-NA administration  mice (25-30 g) were obtained from the Ahvaz Jundishapur University of Medical Sciences (AJUMS) animal facility and, treated in accordance with the principles and guidelines on animal care of AJUMS as reviewed by an ethics committee (IR.AJUMS.REC.1395.136) and kept at a 20 \u00b0C \u00b1 4 \u00b0C temperature with a 12 hr / 12 hr light and dark cycle. They received tap water and commercial chow Group I: ControlGroup II: Vehicle  solvents)Group III: DiabetesGroups IV, V and, VI: Diabetes in addition to administration of Myricitrin 1, 3, and 10 mg/kg, respectively.Groups VII, VIII and, IX: Diabetes in addition to administration of SLN containing myricitrin 1, 3, and 10 mg/kg, respectively  injected anesthetized mice. Then, plasma samples were prepared by blood samples centrifuging at 3500 rpm for 20 min. Following that, the kidney of all animals was separated for histological, gene expression, and apoptosis assessment. The plasma and kidney samples were maintained at \u221280 \u00b0C until the laboratory measurements were performed . Experimental measurementsThe kidney homogenate was prepared using Sharma and Singh method. In brief, the separated kidney slab on the dry ice pack, homogenized in 1/5 (w/v) PBS (pH: 7.4) with a Teflon homogenizer, and centrifuged at 2000 rpm for 10 min. Finally, the supernatant was used to measure the levels of lipid peroxidation (MDA) and antioxidant defense variables  by specific commercial kits .Plasma and urine level of the BUN, Cr, albumin, sodium (Na), potassium (K) and, urine level of glucose were measured by using an Autoanalyzer  electrolyte analyzer devices and biochemical assay kits . Also, GFR, and fractional excretion of sodium and potassium  were assessed by the following formula: GFR= Urine Cr \u00d7 Urine volume / Plasma Cr; FE Na= (Urine Na \u00d7 Plasma Cr / Plasma Na \u00d7 Urine Cr) \u00d7 100; FE K= (Urine K \u00d7 Plasma Cr / Plasma K \u00d7 Urine Cr) \u00d7 100 .Gene expression assessment-\u0394\u0394CT). The sequences of forward and reverse primer for TGF-\u03b2, NF-\u03bab, and GAPDH genes are presented in Real-time PCR method was used for gene expression measurement in the present study. In brief, total RNA and cDNA were purified and synthesized from the kidney by the RNeasy mini kit and Reverse Transcriptase kit, respectively according to their instructions . Real-tiHistopathological and apoptosis assessmentsIn Situ Cell Death Detection Kit, POD, and the dark brown stained nucleus was included as dead cells. The index of apoptosis was measured in 3 randomly slides/animal and ten fields for each slide as a percentage of TUNEL-positive cells  and hematoxylin and eosin (H&E) staining method. Then, seven microscopic slides of tissue sections (5 to 7 \u00b5m) were assessed and read using blind method. Apoptosis assessment was carried out by TUNEL staining according to the labeling of DNA strand breaks by administration of ve cells .Statisticspost hoc least significant difference (LSD) tests. Moreover, the data were represented as the mean\u00b1standard error of the mean (SEM), and the statistically significant differences were considered at P<0.05.The obtained results were statistically analyzed by SPSS software (version 16) with one-way analysis of variance (ANOVA) and The role of myricitrin and its SLN on kidney weight and intake/outputP<0.01) compared to the control and decreased in the groups II (P<0.01), IX (P<0.05), and others (P<0.01) versus diabetes group. The water and food intake showed a significant increase in diabetic mice when compared to control . The water intake decreased in groups IV, V (P<0.05), and others (P<0.01) compared to the diabetes group. Food intake assessment showed a similar effect in the groups II, IV, V, VI (P<0.01), VII, VIII, and IX (P<0.001) compared to diabetes group. The urinary volume of untreated diabetic mice increased significantly (P<0.01). Further, the urine output decreased in groups IX (P<0.001) and others (P<0.01) versus diabetes group.As shown in Lipid peroxidation and antioxidant enzymes level in the kidneyP<0.01). This factor of lipid peroxidation reduced in the groups II (P<0.05), IV, V (P<0.01), VI, and VII (P<0.01) versus diabetes group (P<0.05) and group VI (P<0.01), and increased in groups V (P<0.01), VIII (P<0.05), and IX (P<0.001) in comparison with control. Further, this variable increased in groups II (P<0.05), IV (P<0.01), V (P<0.001), VII (P<0.01), VIII and IX (P<0.001) compared to untreated diabetic mice (P<0.001), groups IV (P<0.01), VI (P<0.001), VIII and IX (P<0.05) when compared to control, but this variable increased in groups II (P<0.001), IV (P<0.05), V (P<0.001), VII, VIII (P<0.01), and IX (P<0.01) versus diabetes group . This variable increased in the groups II (P<0.01), VIII (P<0.001), and others (P<0.05) versus diabetes group. Plasma level of BUN increased in diabetes (P<0.001) and groups IV (P<0.001), V (P<0.01), VI (P<0.05), and VII (P<0.01) compared to the control. Also, this factor decreased in the groups II (P<0.001), IV (P<0.01), and others (P<0.001) compared to untreated diabetic mice. Plasma Cr level showed a significant increase in diabetes and groups IV, V, VI, and IX in comparison with the control group (P<0.001). This variable decreased in the groups II (P<0.001), IV, V (P<0.05), VIII, and IX (P<0.001) versus diabetes group. Plasma level of albumin reduced in all experimental groups except group II versus the control (P<0.001). Moreover, this factor increased in the groups II (P<0.001), V, and VIII (P<0.05) when compared to the diabetes group (P<0.001). Further, this variable increased in the groups IV, VII (P<0.01), and others (P<0.001) versus diabetes group (P<0.01) and groups VI and IX (P<0.001) compared to the control group. Also, this factor increased in the groups II, IV, V, VII, and VIII (P<0.01) when compared to diabetes group (P<0.001) and groups IV, V (P<0.01), VI (P<0.001), VII (P<0.01), VIII (P<0.01), and IX (P<0.001) versus the control group. Moreover, urine albumin level decreased in the groups II (P<0.001), V, and VIII (P<0.01) when compared to untreated diabetic mice  compared to the control. Also, this variable decreased in all groups (P<0.01) except IV and VII versus the diabetes group. The urine K level increased in diabetes compared to control (P<0.01). Also, this factor decreased in the groups II (P<0.01), IV, V (P<0.05), VI (P<0.001), VII (P<0.05), VIII (P<0.01), and IX (P<0.001) when compared to diabetes group. FE Na showed a remarkable increase in diabetes (P<0.001) and groups IV (P<0.05), VI (P<0.001) and, IX (P<0.01) versus to the control group. Further, this variable reduced in the groups II (P<0.001), IV (P<0.01), V (P<0.001), VII, VIII (P < 0.001) and, IX (P<0.05) compared to the diabetes. The results of FE K indicated a significant increase in diabetes (P<0.001) and groups IV, V (P<0.05), VI (P<0.01), and IX (P<0.01) in comparison with the control group. Moreover, FE K decreased in the groups VI (P<0.01) and others (P<0.001) when compared to the diabetes group  and groups V (P<0.01), VI (P<0.001), VIII (P<0.05), and IX (P<0.001) compared to the control group. Also, TGF-\u03b2 gene expression decreased in the groups II (P<0.001), IV (P<0.001), V (P<0.01), VII, and VIII (P<0.001), and increased in the group IX (P<0.001) when compared to the diabetes group . NF-\u03bab gene expression showed a remarkable increase in diabetes (P<0.001) and groups VIII (P<0.01) and IX (P<0.001) versus the control group. This variable decreased in the groups II (P<0.001), IV (P<0.001), V (P<0.01), VII (P<0.001), and VIII (P<0.001) compared to the diabetes group .The gene expression of TGF-\u03b2 increased in diabetes (Effect of myricitrin and its SLN on the kidney histopathology and apoptosisP<0.001) and groups IV (P<0.01), V (P<0.05), VI (P<0.001), VII (P<0.05), and IX (P<0.01) when compared to the control. Furthermore, this variable decreased in the groups II (P<0.001), IV, V (P<0.001), VI (P<0.05), VII, VIII, and IX (P<0.001) compared to the diabetes group  accumulation and infiltration of inflammatory cells in diabetes group compared to the control group. These alterations decreased in the groups II and others treated groups versus diabetes group, and this decreasing effect was more obvious in the groups VII and VIII . The reses group .et al. demonstrated an increase in kidney weight of type 2 diabetic rats due to renal enlargement, hypertrophy, and hyperfunctioning, and treatment with silymarin as a plant-derived antioxidant improved kidney weight (The present study showed that polyphagia, polydipsia, and polyuria occurred in untreated diabetic mice, and myricitrin and its SLN administration improved these disorders. The present results indicated that the kidney weight increased in diabetic mice and administration of myricitrin and its SLN recovered this alteration. Consists with this result Sheela y weight . When thy weight . TherefoT2DM increases ROS production and attenuates free radical scavenging molecules such as antioxidant enzymes. Free radicals induce basement membrane damage and lead to altering the membrane fluidity, ion transports and increased urinary albumin excretion . It seemMurraya paniculata ameliorated kidney dysfunctions of DN in diabetic rats through reducing oxidative stress and increasing antioxidant enzymes level (Urinary albumin excretion, hypoalbuminemia, elevated BUN, serum Cr, urine glucose, and decreased GFR levels occurred in DN, which may be related to T2DM and formation of free radicals . The enhes level . MoreoveGlomerular damage may accelerate tubulointerstitial injury by multiple pathways such as tubular chemokine expression that results in inflammatory cell infiltration in DN . AbnormaThe obtained results indicated that T2DM induced by STZ-NA causes to DN, kidney apoptosis and, inflammation via the increasing lipid peroxidation and decreasing antioxidant defense. Moreover, administration of myricitrin and its SLN in low and moderated doses improved all of the DN changes through reducing oxidative stress and increasing antioxidant enzymes level, and these effects were more potent in the SLN-administered diabetic mice."}
+{"text": "Brachycephalus sulfuratus was recently described from southeastern and southern Brazil. In its description, the authors overlooked previous records of flea toads that had been identified as \u201cBrachycephalus sp. nov.\u201d and B. hermogenesi occurring in the same regions, which could suggest the possibility of up to three flea toads coexisting in southern Brazil. In addition, B. sulfuratus is characterized by substantial phenotypic variability, to an extent that compromises its current diagnosis with respect to its congener B. hermogenesi. Therefore, the current state-of-affairs regarding the geographical distribution of these two species and the identification of previously known populations is hitherto uncertain. Our goals are to reassess previous records of flea toads attributable to B. hermogenesi, B. sulfuratus and \u201cBrachycephalus sp. nov.\u201d, considering the description of B. sulfuratus, and to review the diagnosis of B. sulfuratus.The flea toad B. hermogenesi, B. sulfuratus, or to a potentially undescribed species from southeastern and southern Brazil was based either on the analysis of morphology or on their advertisement calls. These analyses include our independent examinations of specimens and, when not possible, examinations of published descriptions. To allow for a consistent comparison of advertisement calls between B. hermogenesi and B. sulfuratus, we made recordings of both species, including in the type locality of the former.A critical analysis of the species identity of flea toad specimens attributable to B. sulfuratus in relation to B. hermogenesi vary intraspecifically. Live individuals with ventral yellow spots correspond to B. sulfuratus; individuals without yellow spots can be either B. sulfuratus or B. hermogenesi. In preservative, they are indistinguishable. Previous records of Brachycephalus sp. nov. correspond to B. sulfuratus. We propose that the reduced number of notes per call and the presence of only isolated notes in the call of B. sulfuratus, as opposed to a high number of notes per call with isolated notes and note groups in the call of B. hermogenesi, as the only diagnostic characters between them. Regarding their distributions and based in our assessment, only B. sulfuratus occurs in southern Brazil, without any overlap with B. hermogenesi. There is a narrow gap between the distributions of these species around the southeast of the city of S\u00e3o Paulo. Our revision also revealed that some records previously attributed to B. hermogenesi in Rio de Janeiro and north S\u00e3o Paulo represent a distinct, unidentified flea toad that is not B. sulfuratus. Both species occur side by side in Corcovado, S\u00e3o Paulo, a locality from where five paratypes of B. hermogenesi were obtained. Biogeographic events that might have led to vicariance between B. hermogenesi and B. sulfuratus are discussed.We found that morphological and call characters originally proposed as diagnostic for  Brachycephalus Fitzinger, 1826 includes 36 small diurnal anuran species that live in the leaf litter across the Brazilian Atlantic Rainforest , municipality of Guaraque\u00e7aba, in the northern coast of Paran\u00e1 . Third, if the unidentified species of B. sulfuratus, there could be a single species of flea toad in southern Brazil (B. sulfuratus).The absence of a nomenclatural review of records of flea toads in southern Brazil can be evidenced by the fact that a single location in Santa Catarina, called Castelo dos Bugres, was recorded as harboring specimens identified as \u201c nov. 1\u201d , or Bracs sp. 1.  and B. smogenesi , Brachycalus sp.  and B. slfuratus . Second, records  were assp. nov.\u201d , leadingBrachycephalus sp. nov. 1\u201d , Curitiba, Paran\u00e1, Brazil, Cole\u00e7\u00e3o Herpetol\u00f3gica do Departamento de Zoologia (DZUP), Universidade Federal do Paran\u00e1, Curitiba, Paran\u00e1, Brazil, and Museu de Hist\u00f3ria Natural (ZUEC), Universidade Estadual de Campinas, Campinas, S\u00e3o Paulo, Brazil. The sound collection analyzed include MHCNI, Xeno-Canto sound collection (www.xeno-canto.org), and Fonoteca Neotropical Jacques Vielliard .The critical analysis of the species identity of specimens attributable to B. sulfuratus .The analyses began by the assessment of the original diagnosis of lfuratus . We lookB. sulfuratus and B. hermogenesi, we noticed that the calls of B. hermogenesi described by B. hermogenesi  and digital  devices, with Sennheiser ME 66 and ME 67 microphones. Analogical recordings were digitized at 44.1 kHz and 16 bit using Raven Pro 1.4 . Digital recordings were made equally with sampling frequency rate of 44.1 kHz and 16-bit resolution. We analyzed calls under note-centered approach , as BornB. sulfuratus and B. hermogenesi, vouchered with specimens collected and deposited in the MHNCI. Collection permits were issued by ICMBIO . Geographical coordinates are based on the WGS84 datum. Elevations for literature records and author\u2019s records were obtained from Google Earth, after plotting the location point  It \u201cdiffers from\u2026 B. hermogenesi\u2026 by having (in life) yellow blotches on the ventral surfaces of the throat, chest, arms, and forearms\u201d . The inverted v-shaped mark can be absent in individuals of B. sulfuratus \u201d .\u201dorearms\u201d : 43, 50;genesi\u2026\u201d : 43, 50;ogenesi\u201d : 50. SpeCatarina  have revCatarina . Moreove compare . Additio pulex)\u201d : 50. Als pulex)\u201d : 46. Fin pulex)\u201d : 50: \u201cThB. sulfuratus from B. hermogenesi. However, for identification purposes, we considered individuals with yellow spots on their ventral side as B. sulfuratus, whereas individuals without yellow spots could be either B. sulfuratus or B. hermogenesi. It is important to note that specimens with yellow spots of B. sulfuratus must be observed in life because in the preservative the change in color prevents separate them in relation to specimens of B. hermogenesi.Currently, there are no unique morphological character that could differentiate either live or preserved specimens  for B. sB. sulfuratus the following parameters of the advertisement call: \u201cadvertisement call long, composed of a set of 4\u20137 high-frequency notes (6.2\u20137.2 kHz) repeated regularly.\u201d In the section \u201cComparisons with other species\u201d, B. hermogenesi is the most similar to the new species (B. sulfuratus), being quite similar in frequency (dominant frequency = 6.8 kHz), which are the highest recorded for the genus. However, the advertisement call of B. hermogenesi can be simple or composed of 2\u20137 shorter notes with 1\u20133 pulses  is within the range of B. sulfuratus (6.2\u20137.2).In addition to morphological characters, 3 pulses .\u201d In sumB. hermogenesi call can be simple or composed  and the presence of \u201cattenuated notes\u201d  with only isolated notes of B. sulfuratus, while in B. hermogenesi the advertisement call has a high number of notes (\u226524) with the presence of isolated notes and note groups . One of these records involves \u201cB. hermogenesi\u201d from the municipality of Piedade, state of S\u00e3o Paulo, of HQ435682.1 and HQ435709.1; B. sulfuratus, which was placed on the tree together with a specimen from the Municipality of Barra do Turvo, in an early-diverging branch of the B. sulfuratus clade on the tree , also examined by us, collected in the state of Paran\u00e1, that was first identified as \u201c nov. 1\u201d , \u201cBrachys sp. 1\u201d , and, fifuratus\u201d ). There part.), two localities previously considered as occurrence of B. hermogenesi . The phylogenetic analysis revealed that the specimen from Municipality of Paraibuna is indeed B. hermogenesi  with distinct durations (= transliteration size) related to the number of notes in the call. This diagnosis between B. sulfuratus and B. hermogenesi is only feasible under the note-centered approach. Considering their calls under the call-centered approach, there would be no diagnosis to be proposed between them at this moment, because each note would represent a call , consolidate the benefit of the note-centered approach over the call-centered approach in describing calls of species of this genus , for example, from the B. ephippium group, which includes most species from the state of S\u00e3o Paulo to the north up to Esp\u00edrito Santo and Minas Gerais.The advertisement calls of ix group , 2018a, B. hermogenesi in southern Brazil and the presence of B. sulfuratus as far north as the east of S\u00e3o Paulo city, only 25 km in straight line from the southernmost site of a confirmed record of B. hermogenesi  since 2009, focusing on their distribution, ecology and conservation , on the border between the municipalities of Jaragu\u00e1 do Sul and Massaranduba, where we described B. albolineatus . In the studies . In the  studies  and snak studies .Brachycephalus toads and Scytalopus birds of S\u00e3o Paulo, Paran\u00e1, and Santa Catarina originated less than 2\u20135 million years ago  and Mesozoic, respectively  and Eleoscytalopus indigoticus , and calls of B. hermogenesi in N\u00facleo Santa Virg\u00ednea, Parque Estadual da Serra do Mar, municipality of S\u00e3o Luiz do Paraitinga, S\u00e3o Paulo, can be heard in a recording of E. indigoticus  at Corcovado, S\u00e3o Paulo, is the only confirmed case of sympatry between species of Brachycephalus in the same group. Other cases of sympatry include Brachycephalus from distinct groups . RIO DE JANEIRO: Trilha do Corisco, municipality of Paraty MHNCI 206\u201312. S\u00c3O PAULO: Corcovado, municipality of Ubatuba MHNCI 193\u2013205.10.7717/peerj.10983/supp-1Supplemental Information 1Abbreviation: NA = not available.Click here for additional data file.10.7717/peerj.10983/supp-2Supplemental Information 2MHNCI 124 Brachycephalus sulfuratus near Jurupara dam voucher MHNCI 10791 or MHNCI 10792 29 set 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-3Supplemental Information 3MHNCI 124 Brachycephalus sulfuratus near Jurupara dam voucher MHNCI 10791 or MHNCI 10792 29 set 2016 MRBornschein cutted.Click here for additional data file.10.7717/peerj.10983/supp-4Supplemental Information 4MHNCI 125 Brachycephalus sulfuratus near Jurupara dam not collected 29 set 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-5Supplemental Information 5MHNCI 126 Brachycephalus sulfuratus Nucleo Itutinga Piloes ind 1 not collected 08 out 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-6Supplemental Information 6MHNCI 127 Brachycephalus sulfuratus Nucleo Itutinga Piloes ind 2 not collected 08 out 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-7Supplemental Information 7MHNCI 128 Brachycephalus sulfuratus Biquinha not collected 18 set 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-8Supplemental Information 8MHNCI 129 Brachycephalus sulfuratus Serra Agua Limpa MHNCI 11583 26 out 2011 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-9Supplemental Information 9MHNCI 130 Brachycephalus sulfuratus Serra do Guarau not collected 22 out 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-10Supplemental Information 10MHNCI 131 Brachycephalus sulfuratus Caratuval near Parque Estadual Lauraceas MHNCI 11571 L Correa.Click here for additional data file.10.7717/peerj.10983/supp-11Supplemental Information 11MHNCI 132 Brachycephalus sulfuratus Caratuval Parque Estadual Lauraceas not collected 18 dez 2010 L Correa.Click here for additional data file.10.7717/peerj.10983/supp-12Supplemental Information 12MHNCI 133 Brachycephalus sulfuratus Reserva Particular do Patrimonio Natural Salto Morato not collected 10 marco 2016 MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-13Supplemental Information 13MHNCI 134 Brachycephalus sulfuratus Fazenda Thalia not collected MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-14Supplemental Information 14MHNCI 135 Brachycephalus sulfuratus Truticultura not collected MRBornschein.Click here for additional data file.10.7717/peerj.10983/supp-15Supplemental Information 15MHNCI 136 Brachycephalus sulfuratus Morro Garuva not collected 02 nov 2016 Andre Confetti.Click here for additional data file.10.7717/peerj.10983/supp-16Supplemental Information 16MHNCI 137 Brachycephalus sulfuratus Morro do Garraf\u00e3o not collected LFRibeiro.Click here for additional data file.10.7717/peerj.10983/supp-17Supplemental Information 17MHNCI 165 000107 0122S4 Brachy hermogenesi Corcovado specimen one two starting notes lost without interruption.Click here for additional data file.10.7717/peerj.10983/supp-18Supplemental Information 18MHNCI 168 190330 05 Brachy hermogenesi ind 03 Boraceia 30 marco 2019 MRB.Click here for additional data file.10.7717/peerj.10983/supp-19Supplemental Information 19MHNCI 169 190330 06 Brachy hermogenesi ind 04 Boraceia 30 marco 2019 MRB.Click here for additional data file.10.7717/peerj.10983/supp-20Supplemental Information 20MHNCI 172 BLR00061 Brachy hemogenesi Picinguaba ind 01 ex 01 sem detalhes 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-21Supplemental Information 21MHNCI 173 BLR00062 B hermogenesi Picinguaba ind2 ex1 4 notas perdidas 13dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-22Supplemental Information 22MHNCI 174 BLR00064 B hermogenesi Picinguaba ind3 6 notas perdidas 13dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-23Supplemental Information 23MHNCI 175 BLR00065 B hermogenesi Picinguaba ind1ex2 tudo 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-24Supplemental Information 24MHNCI 176 BLR00067 B hermogenesi Picinguaba ind1ex3 tudo ind4ex1 3 perdidas 13dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-25Supplemental Information 25MHNCI 177 BLR00068 B hermogenesi_Picinguaba ind2 ex2 tudo 05cm 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-26Supplemental Information 26MHNCI 178 BLR00069 B hermogenesi Picinguaba ind3 ex2 5 notas perdidas 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-27Supplemental Information 27MHNCI 179 BLR00070 B hermogenesi Picinguaba ind 04 ex 02 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-28Supplemental Information 28MHNCI 180 BLR00071 B hermogenesi Picinguaba ind 03 ex 03 sem detalhes 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-29Supplemental Information 29MHNCI 181 BLR00072 B hermogenesi Picinguaba ind3 ex4 perdeu uma nota 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-30Supplemental Information 30MHNCI 182 BLR00073 B hermogenesi Picinguaba ind2 ex3 soh o final do canto 13 dez 2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-31Supplemental Information 31MHNCI 183 BLR00076 B hermogenesi Picinguaba ind4ex3 tudo 5-20cm distancia 13dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-32Supplemental Information 32MHNCI 184 000106 0115S4 Brachy hermogenesi Picinguaba specimen one several starting notes lost without interruption.Click here for additional data file.10.7717/peerj.10983/supp-33Supplemental Information 33MHNCI 185 000106 0117S4 Brachy hermogenesi Picinguaba specimen two two starting notes lost without interruption.Click here for additional data file.10.7717/peerj.10983/supp-34Supplemental Information 34MHNCI 186 000106 0119S4 Brachy hermogenesi Picinguaba specimen three one starting notes lost without interruption.Click here for additional data file.10.7717/peerj.10983/supp-35Supplemental Information 35MHNCI 187 000106 0120S4 Brachy hermogenesi Picinguaba specimen four several starting notes lost without interruption.Click here for additional data file.10.7717/peerj.10983/supp-36Supplemental Information 36MHNCI 213 080409 02 B hermogenesi ind1 muito perdido ruim Paranapiacaba MRB.Click here for additional data file.10.7717/peerj.10983/supp-37Supplemental Information 37MHNCI_188_000103_0135S4_Brachy hermogenesi Trilha do Ipiranga ind 1 e unknown starting notes with interruption.Click here for additional data file.10.7717/peerj.10983/supp-38Supplemental Information 38MHNCI 189 000103 0137S4 Brachy hermogenesi Trilha do Ipiranga ind 2 three starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-39Supplemental Information 39MHNCI 190 000103 0140S4 Brachy hermogenesi Trilha do Ipiranga ind 3 three starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-40Supplemental Information 40MHNCI 191 000103 0141S4 Brachy hermogenesi Trilha do Ipiranga ind 4 four starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-41Supplemental Information 41MHNCI 193 BLR00084 Brachy sp ind1 parte final Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-42Supplemental Information 42MHNCI 194 BLR00085 Brachy sp ind2 2m Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-43Supplemental Information 43MHNC 195 BLR00086 Brachy sp ind3 perdeu notas 20cm Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-44Supplemental Information 44MHNCI 196 BLR00087 Brachy sp ind4ex1 perdeu 3 notas 30-100cm Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-45Supplemental Information 45MHNCI 197 BLR00089 Brachy sp ind5 soh final Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-46Supplemental Information 46MHNCI 198 BLR00091 Brachy sp ind6ex1 4 notas perdidas 10-50cm Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-47Supplemental Information 47MHNCI 199 BLR00092 Brachy sp ind6ex2 soh final 50-80cm Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-48Supplemental Information 48MHNCI 200 BLR00093 Brachy sp ind4ex2 perdeu 7 notas 20cm Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-49Supplemental Information 49MHNCI 201 BLR00094 Brachy sp ind7 perdeu 4 notas 20-100cm Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-50Supplemental Information 50MHNCI 202 BLR00095 Brachy sp ind8 perdeu 6 notas Corcovado 18dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-51Supplemental Information 51MHNCI 203 000102 0129S4 Brachycephalus sp ind1 Corcovado two starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-52Supplemental Information 52MHNCI 204 000102 0130S4 Brachycephalus sp ind2 Corcovado two starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-53Supplemental Information 53MHNCI 205 000102 0132S4 Brachycephalus sp ind4 ind5 Corcovado two starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-54Supplemental Information 54MHNCI 211 000104 0113S4 Brachycephalus sp ind1 Trilha do Corisco several starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-55Supplemental Information 55MHNCI 212 000104 0114S4 Brachycephalus sp ind2 Trilha do Corisco several starting notes lost.Click here for additional data file.10.7717/peerj.10983/supp-56Supplemental Information 56MHNCI 206 BLR00053 Brachycephalus sp Trilha do Corisco ind1 perdeu inicio 40 cm 12dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-57Supplemental Information 57MHNCI 207 BLR00054 Brachycephalus sp Trilha do Corisco ind2 perdeu inicio 12dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-58Supplemental Information 58MHNCI 208 BLR00055 Brachycephalus sp Trilha do Corisco ind3 microfone flutuou 12dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-59Supplemental Information 59MHNCI 209 BLR00056 Brachycephalus sp Trilha do Corisco ind4 20-60 cm 12dez2017 MRB.Click here for additional data file.10.7717/peerj.10983/supp-60Supplemental Information 60MHNCI 217 190206 05 Brachycephalus sp mais B sulfuratus Serra do Pico MRB 6fev2019.Click here for additional data file.10.7717/peerj.10983/supp-61Supplemental Information 61MHNC 218 BLR00165 Brachy sp mais Brachy sulfuratus Torre Embratel MRB n\u00e3o coletado.Click here for additional data file.10.7717/peerj.10983/supp-62Supplemental Information 62MHNCI 219 200123 0448S4 Brachycephalus sulfuratus Entroncamento Teba 20 cm from microphone LFR JAN-2020.Click here for additional data file.10.7717/peerj.10983/supp-63Supplemental Information 63MHNCI 220 Brachycephalus sulfuratus Morro do Canal 150 cm from microphone LFRibeiro JAN-2017.Click here for additional data file.10.7717/peerj.10983/supp-64Supplemental Information 64MHNCI 221 161115 02 Brachycephalus sulfuratus Monte Crista 100 cm from microphone Luiz Fernando Ribeiro NOV-2016.Click here for additional data file.10.7717/peerj.10983/supp-65Supplemental Information 65Brachycephalus hermogenesi ind1 parte do canto Morro do Cantagalo-Caraguatatuba LC.MHNCI 222 180802 08 Click here for additional data file.10.7717/peerj.10983/supp-66Supplemental Information 66Brachycephalus hermogenesi ind2 parte do canto Morro do Cantagalo-Caraguatatuba LC.MHNCI 223 180802 09 Click here for additional data file."}
+{"text": "Pancreatic ductal adenocarcinoma (PDAC) is a fatal malignant neoplasm. It is necessary to improve the understanding of the underlying molecular mechanisms and identify the key genes and signaling pathways involved in PDAC.GSE28735, GSE62165, and GSE91035 were downloaded from the Gene Expression Omnibus. Differentially expressed genes (DEGs) were identified by integrated bioinformatics analysis, including protein\u2013protein interaction (PPI) network, Gene Ontology (GO) enrichment, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. The PPI network was established using the Search Tool for the Retrieval of Interacting Genes (STRING) and Cytoscape software. GO functional annotation and KEGG pathway analyses were performed using the Database for Annotation, Visualization, and Integrated Discovery. Hub genes were validated via the Gene Expression Profiling Interactive Analysis tool (GEPIA) and the Human Protein Atlas (HPA) website.The microarray datasets FN1, COL1A1, COL3A1, BGN, POSTN, FBN1, COL5A2, COL12A1, THBS2, COL6A3, VCAN, CDH11, MMP14, LTBP1, IGFBP5, ALB, CXCL12, FAP, MATN3, and COL8A1. These genes were validated using The Cancer Genome Atlas (TCGA) and Genotype\u2013Tissue Expression (GTEx) databases, and the encoded proteins were subsequently validated using the HPA website. The GO analysis results showed that the most significantly enriched biological process, cellular component, and molecular function terms among the 20 hub genes were cell adhesion, proteinaceous extracellular matrix, and calcium ion binding, respectively. The KEGG pathway analysis showed that the 20 hub genes were mainly enriched in ECM\u2013receptor interaction, focal adhesion, PI3K-Akt signaling pathway, and protein digestion and absorption. These findings indicated that FBN1 and COL8A1 appear to be involved in the progression of PDAC. Moreover, patient survival analysis performed via the GEPIA using TCGA and GTEx databases demonstrated that the expression levels of COL12A1 and MMP14 were correlated with a poor prognosis in PDAC patients (p\u00a0<\u00a00.05).A total of 263 DEGs (167 upregulated and 96 downregulated) were common to the three datasets. We used STRING and Cytoscape software to establish the PPI network and then identified key modules. From the PPI network, 225 nodes and 803 edges were selected. The most significant module, which comprised 11 DEGs, was identified using the Molecular Complex Detection plugin. The top 20 hub genes, which were filtered by the CytoHubba plugin, comprised MMP14 and COL12A1 is associated with poor overall survival, and these might be a combination of prognostic biomarkers in PDAC.The results demonstrated that upregulation of  Pancreatic ductal adenocarcinoma (PDAC) is the most common malignant tumor of the pancreas and is a lethal malignancy with poor prognosis, which is in part due to its rapid progression and the lack of diagnostic and therapeutic targets. In 2018, pancreatic cancer (PC) ranked 11th among the most common cancers, with 458,918 new cases and 432,242 deaths due to PC worldwide . Recent KRAS  and biological pathways linked to various malignant tumors. Therefore, microarray techniques are promising and efficient ways to identify candidate biomarkers involved in the pathogenesis of PDAC. The purpose of our study was to determine significant DEGs and pathways implicated in PDAC by integrated bioinformatics analysis and to provide novel insights into the progression, diagnosis, and therapeutic targets of PDAC.https://www.ncbi.nlm.nih.gov/geo/) is a public repository of high-throughput gene expression genomics datasets  is an online analysis tool that is based on the R programming language and can be used to identify DEGs that differentiate between cancer and normal samples in a GEO series  is an online application that can be used to assess DEG-encoded proteins and protein\u2013protein interaction (PPI) networks  is a public online bioinformatics database (p\u00a0<\u00a00.05. We performed enrichment of the GO terms and KEGG pathways for the candidate DEGs using DAVID.The Gene Ontology (GO) is used to perform enrichment analysis, which covers the cellular component (CC), biological process (BP), and molecular function (MF), of the selected genes . The Kyodatabase  that conhttp://gepia.cancer-pku.cn/) is used to perform functions such as survival analysis, the detection of similar genes, and correlation analysis to clarify the relationships between diseases and DEGs  database and the Genotype\u2013Tissue Expression (GTEx) database, the Gene Expression Profiling Interactive Analysis tool  website on the basis of spatial proteomics data and quantitative transcriptomics data (RNA-Seq) obtained from immunohistochemical analysis of tissue microarrays.The expression of proteins encoded by the PDAC candidate genes was validated using the Human Protein Atlas Functional and pathway enrichment analyses were accomplished using DAVID. GO analysis showed that the most significant module was mainly enriched in cell adhesion, extracellular matrix structural constituent, and proteinaceous extracellular matrix  Table 2Table 2. COL12A1 and MMP14 were correlated with an unfavorable prognosis in PDAC patients (p\u00a0<\u00a00.05) (p\u00a0>\u00a00.05).Patient survival analysis performed via the GEPIA using TCGA and GTEx databases demonstrated that the high expression levels of \u00a0<\u00a00.05) . The oveFN1, COL1A1, COL3A1, BGN, POSTN, FBN1, COL5A2, COL12A1, THBS2, COL6A3, VCAN, CDH11, MMP14, LTBP1, IGFBP5, FAP, MATN3, and COL8A1 were significantly overexpressed in PDAC tissues in comparison with normal pancreatic tissues, whereas ALB was underexpressed in PDAC tissues (p\u00a0<\u00a00.05) .To ensure the reliability of the identification of the top 20 hub genes, we validated these via the GEPIA using TCGA and GTEx databases. Boxplots of the hub genes associated with PDAC were downloaded from the GEPIA. The results demonstrated that \u00a0<\u00a00.05) . CXCL12 COL5A2, IGFBP5, and MATN3 are reported on the HPA website, and expression profiles of the other 17 genes in PDAC clinical specimens are shown in We obtained the expression levels of proteins encoded by the 20 hub genes associated with PDAC from the HPA website. No data for proteins encoded by FN1, MMP14, COL12A1, COL3A1, COL1A1, POSTN, VCAN, LTBP1, FBN1, and FAP were upregulated in PDAC tissues in comparison with normal tissues, with only ALB being downregulated in PDAC tissues. COL6A3, COL8A1, CDH11, and CXCL12 were not expressed in either PDAC tissues or normal tissues, and BGN and THBS2 were overexpressed in both cancer and normal tissues.The protein expressions of GSE28735, GSE62165, and GSE91035. The main findings deduced from the studies used to compile GSE28735 were that dipeptidase 1 and a unique set of free fatty acids played roles in the development, progression, and prognosis of PC and might be potential targets in PDAC (GSE62165 found that hepatocyte nuclear factor (HNF)-1 \u03b1 and HNF-1 \u03b2 seem to be good candidates as tumor suppressors in PDAC . Ca2+ is a ubiquitous and versatile second messenger involved in the regulation of numerous cellular functions, including gene transcription, vesicular trafficking, and cytoskeletal rearrangements  and enhances the production of type I collagen by increasing transforming growth factor- \u03b2 signaling  are a family of calcium- and zinc-dependent membrane-anchored or secreted endopeptidases that are overexpressed in various diseases, including breast cancer . MMP14 i of PDAC . Moreoveasive PC , and MMPasive PC . Type I al cells , and COLignaling . Ottaviac tissue . These fCOL1A1, COL3A1, COL5A2, COL6A3, FN1, and THBS2, were significantly associated with ECM\u2013receptor interactions, focal adhesion, and the phosphatidylinositol-3-kinase\u2013protein kinase B (PI3K-Akt) signaling pathway. In addition, collagen-encoding genes, including COL1A1, COL3A1, and COL5A2, were also enriched in protein digestion and absorption and platelet activation.The KEGG pathway analysis showed that six hub genes, namely, COL1A1 and COL3A1 were significantly downregulated in PC (p <0.0001) after treatment with gemcitabine in combination with EC359   . As we hFBN1 and COL8A1 appear to be involved in the progression of PDAC. FBN1 encodes a structural component of the microfibrils of the ECM that have diameters of 10\u201312 nm, which impart both regulatory and structural properties to load-bearing connective tissues  and the related enriched functions or pathways, which regulate the progression and metastatic invasion of PDAC, as well as overall survival. The results demonstrate that the upregulation of MMP14 and COL12A1 in PDAC is closely associated with poor overall survival, that these might be a potential combination of prognostic biomarkers in patients with PDAC, and that FBN1 and COL8A1 might be biomarkers of PDAC. In brief, our study increases the understanding of the potential critical genes and related pathways that participate in the pathogenesis of PDAC.In conclusion, we screened the top 20 hub genes (10.7717/peerj.10419/supp-1File S1Click here for additional data file.10.7717/peerj.10419/supp-2Table S1Click here for additional data file.10.7717/peerj.10419/supp-3Supplemental Information 3Click here for additional data file.10.7717/peerj.10419/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.10419/supp-5Supplemental Information 5Click here for additional data file.10.7717/peerj.10419/supp-6Supplemental Information 6Click here for additional data file.10.7717/peerj.10419/supp-7Supplemental Information 7Click here for additional data file.10.7717/peerj.10419/supp-8Supplemental Information 8Click here for additional data file.10.7717/peerj.10419/supp-9Supplemental Information 9Click here for additional data file.10.7717/peerj.10419/supp-10Supplemental Information 10Click here for additional data file.10.7717/peerj.10419/supp-11Supplemental Information 11Click here for additional data file.10.7717/peerj.10419/supp-12Supplemental Information 12Click here for additional data file.10.7717/peerj.10419/supp-13Supplemental Information 13Click here for additional data file.10.7717/peerj.10419/supp-14Supplemental Information 14Click here for additional data file.10.7717/peerj.10419/supp-15Supplemental Information 15Click here for additional data file.10.7717/peerj.10419/supp-16Supplemental Information 16Click here for additional data file.10.7717/peerj.10419/supp-17Supplemental Information 17Click here for additional data file.10.7717/peerj.10419/supp-18Supplemental Information 18Click here for additional data file.10.7717/peerj.10419/supp-19Supplemental Information 19Click here for additional data file.10.7717/peerj.10419/supp-20Supplemental Information 20Click here for additional data file.10.7717/peerj.10419/supp-21Supplemental Information 21Click here for additional data file.10.7717/peerj.10419/supp-22Supplemental Information 22Click here for additional data file.10.7717/peerj.10419/supp-23Supplemental Information 23Click here for additional data file.10.7717/peerj.10419/supp-24Supplemental Information 24Click here for additional data file.10.7717/peerj.10419/supp-25Supplemental Information 25Click here for additional data file.10.7717/peerj.10419/supp-26Supplemental Information 26Click here for additional data file.10.7717/peerj.10419/supp-27Supplemental Information 27Click here for additional data file.10.7717/peerj.10419/supp-28Supplemental Information 28Click here for additional data file.10.7717/peerj.10419/supp-29Supplemental Information 29Click here for additional data file.10.7717/peerj.10419/supp-30Supplemental Information 30Click here for additional data file.10.7717/peerj.10419/supp-31Supplemental Information 31Click here for additional data file.10.7717/peerj.10419/supp-32Supplemental Information 32Click here for additional data file.10.7717/peerj.10419/supp-33Supplemental Information 33Click here for additional data file.10.7717/peerj.10419/supp-34Supplemental Information 34Click here for additional data file.10.7717/peerj.10419/supp-35Supplemental Information 35Click here for additional data file.10.7717/peerj.10419/supp-36Supplemental Information 36Click here for additional data file.10.7717/peerj.10419/supp-37Supplemental Information 37Click here for additional data file.10.7717/peerj.10419/supp-38Supplemental Information 38Click here for additional data file.10.7717/peerj.10419/supp-39Supplemental Information 39Click here for additional data file.10.7717/peerj.10419/supp-40Supplemental Information 40Click here for additional data file.10.7717/peerj.10419/supp-41Supplemental Information 41Click here for additional data file.10.7717/peerj.10419/supp-42Supplemental Information 42Click here for additional data file.10.7717/peerj.10419/supp-43Supplemental Information 43Click here for additional data file.10.7717/peerj.10419/supp-44Supplemental Information 44Click here for additional data file.10.7717/peerj.10419/supp-45Supplemental Information 45Click here for additional data file.10.7717/peerj.10419/supp-46Supplemental Information 46Click here for additional data file.10.7717/peerj.10419/supp-47Supplemental Information 47Click here for additional data file.10.7717/peerj.10419/supp-48Supplemental Information 48Click here for additional data file.10.7717/peerj.10419/supp-49Supplemental Information 49Click here for additional data file.10.7717/peerj.10419/supp-50Supplemental Information 50Click here for additional data file.10.7717/peerj.10419/supp-51Supplemental Information 51Click here for additional data file.10.7717/peerj.10419/supp-52Supplemental Information 52Click here for additional data file.10.7717/peerj.10419/supp-53Supplemental Information 53Click here for additional data file.10.7717/peerj.10419/supp-54Supplemental Information 54Click here for additional data file.10.7717/peerj.10419/supp-55Supplemental Information 55Click here for additional data file.10.7717/peerj.10419/supp-56Supplemental Information 56Click here for additional data file.10.7717/peerj.10419/supp-57Supplemental Information 57Click here for additional data file.10.7717/peerj.10419/supp-58Supplemental Information 58Click here for additional data file.10.7717/peerj.10419/supp-59Supplemental Information 59Click here for additional data file.10.7717/peerj.10419/supp-60Supplemental Information 60Click here for additional data file.10.7717/peerj.10419/supp-61Supplemental Information 61Click here for additional data file.10.7717/peerj.10419/supp-62Supplemental Information 62Click here for additional data file.10.7717/peerj.10419/supp-63Supplemental Information 63Click here for additional data file.10.7717/peerj.10419/supp-64Supplemental Information 64Click here for additional data file.10.7717/peerj.10419/supp-65Supplemental Information 65Click here for additional data file.10.7717/peerj.10419/supp-66Supplemental Information 66Click here for additional data file.10.7717/peerj.10419/supp-67Supplemental Information 67Click here for additional data file.10.7717/peerj.10419/supp-68Supplemental Information 68Click here for additional data file.10.7717/peerj.10419/supp-69Supplemental Information 69Click here for additional data file.10.7717/peerj.10419/supp-70Supplemental Information 70Click here for additional data file.10.7717/peerj.10419/supp-71Supplemental Information 71Click here for additional data file.10.7717/peerj.10419/supp-72Supplemental Information 72Click here for additional data file.10.7717/peerj.10419/supp-73Supplemental Information 73Click here for additional data file.10.7717/peerj.10419/supp-74Supplemental Information 74Click here for additional data file.10.7717/peerj.10419/supp-75Supplemental Information 75Click here for additional data file.10.7717/peerj.10419/supp-76Supplemental Information 76Click here for additional data file.10.7717/peerj.10419/supp-77Supplemental Information 77Click here for additional data file.10.7717/peerj.10419/supp-78Supplemental Information 78Click here for additional data file.10.7717/peerj.10419/supp-79Supplemental Information 79Click here for additional data file.10.7717/peerj.10419/supp-80Supplemental Information 80Click here for additional data file.10.7717/peerj.10419/supp-81Supplemental Information 81Click here for additional data file.10.7717/peerj.10419/supp-82Supplemental Information 82Click here for additional data file."}
+{"text": "Fear of Falling (FOF) is common among community-dwelling older adults and is associated with increased fall-risk. In this cross-sectional study we examined the relationships between FOF and factors associated with fall-risk such as gait quality, cognition, and walking-confidence. Using baseline data from older adult participants in a randomized exercise trial , we quantified the following outcome measure for (1) gait quality: harmonic ratio (smoothness) and time-frequency spatiotemporal variables from triaxial accelerometry during 6 minute walk; gait speed, step-time CoV (variability) and walk-ratio (step-length/cadence) on an instrumented walkway; (2) cognition: Trails A and B (3) walking-confidence: Gait efficacy Scale. Mann Whitney U-tests indicated individuals without FOF had better gait quality (p<0.05): greater smoothness (2.38\u00b1.58 vs 1.14\u00b1.73), speed (1.10\u00b1.15 vs 1.04\u00b1.17 m/s) and walk-ratio (.56\u00b1.07 vs .53\u00b1.08 cm/steps/min), lower step-time CoV (3.72\u00b11.24 vs 4.17\u00b11.66), and greater walking-confidence (89\u00b111 vs 79\u00b113). A random forest classifier predicted FOF with 64% (gait only) and 70%  accuracy; Gini-index based ranking indicated gait quality  direction, walking speed) were consistently important variables. Linear Support Vector Machine learning yielded accuracies of 60% (only gait) and 68% : smoothness V, mediolateral frequency bandwidth, gait speed among top 4 ranked variables in both models, and walking-confidence in the additional measures model; smoothness-V the highest weighted coefficient (-0.52). Based on these findings, interventions targeted for gait quality and walking-confidence may be important to overcome FOF and reduce fall risks."}
+{"text": "Staphylococcus aureus (MRSA), there have been few studies focused on the molecular characterization of methicillin-susceptible Staphylococcus aureus (MSSA). In this cross-sectional study, 85 MSSA isolates were characterized by antimicrobial susceptibility testing, virulence genes analysis, accessory gene regulator (agr) typing, and S. aureus protein A locus (spa) typing.Compared to methicillin-resistant agr types detected in tested strains were mainly type I (76.5%), II (12.9%), and III (10.6%). Of 85 MSSA examined isolates, 48 (56.5%) isolates were toxinogenic with 27 producing pvl (31.8%) and 21 tst (24.7%). The findings of the study show a high genetic diversity in MSSA strains warranting continued surveillance to provide critical insights into control and treatment of MSSA infections.In present study, 9 different clonal complexes namely CC8-MSSA-t037 (22.4%), CC8-MSSA-t008 (11.8%), CC7-MSSA-t091 and CC30-MSSA-t021 (each 9.4%), CC8-MSSA-t037 (8.3%), CC398-MSSA-t034 (7.1%), CC22-MSSA-t005 (5.9%), CC5-MSSA-t002 and CC15-MSSA-t084 (each 4.7%), CC22-MSSA-t790 and CC59-MSSA-t437 (each 3.5%), CC22-MSSA-t1869, CC5-MSSA-t045, and CC45-MSSA-t015 (each 2.3%), CC30-MSSA-t318 and CC15-MSSA-t491 (each 1.2%) were found.  Staphylococcus aureus is a common hospital- and community-acquired pathogen [S. aureus strain diversity appears to differ by geographic region, there has been a dramatic increase in the prevalence of S. aureus strains associated with human infections around the world and this appears to be especially true for methicillin-susceptible Staphylococcus aureus (MSSA) [S. aureus infections [pathogen . It is rpathogen . Althougs (MSSA) \u20135. MSSA s (MSSA) , 6. Comps (MSSA) . Antimicfections , 8. Knowfections , 10. AltS. aureus isolates phenotypically by using standard microbiological techniques. Polymerase chain reaction (PCR) assay targeting the S. aureus-specific nuc gene was applied to verify the isolates [S. aureus isolates susceptible to cefoxitin disc  in a disc diffusion assay using established methods (CLSI 2018) and negative for the presence of mecA gene by PCR were considered as MSSA strains [Eighty-five MSSA isolates were obtained from hospitalized patients at four hospitals affiliated to Shahid Beheshti University of Medical Sciences during an 9-month collection period from March 2019 to November 2019. This study protocol was approved by the Ethics Committee of the Shahid Beheshti University of Medical Sciences in Tehran, Iran (IR. SBMU. MSP.REC. 1398. 774). Furthermore, we confirmed isolates , 12. TheS. aureus ATCC 25923, ATCC 43300, and ATCC 29213 strains.In present study, Kirby\u2013Bauer disk diffusion method was applied to determine the antimicrobial susceptibility of isolates based on the clinical and laboratory standards institute (CLSI) criteria (CLSI 2018). The antimicrobial agents included penicillin, teicoplanin, gentamicin, kanamycin, amikacin, tobramycin, clindamycin, erythromycin, tetracycline, linezolid, rifampicin, mupirocin, ciprofloxacin, quinupristin\u2013dalfopristin, and trimethoprim\u2013sulfamethoxazole . The minimal inhibitory concentrations (MIC) values of vancomycin was evaluated by broth microdilution method. Susceptibility test was quality controlled by using eta, and etb), Panton-Valentine leukotoxin gene (pvl), and toxic shock syndrome toxin (tsst-1) by PCR assay [S. aureus ATCC49775 and toxin positive S. aureus\u00a0strains obtained from our previous were used as reference strains [S. aureus strain ATCC 25923 were also used as negative control.Genomic DNA was isolated using the phenol\u2013chloroform extraction method. All of the isolates were screened for virulence encoding genes namely: exfoliative toxin genes (CR assay \u201314. The agr type detection using primer set comprising a common forward primer (Pan) and reverse primers  specific to each agr group [agr types were identified by comparing the banding patterns of isolates to RN6390 (agr type I), RN6607 (agr type II), RN8465 (agr type III), RN4550 (agr type IV), and RN6911 (negative control), as reference strains. PCR amplification was used for spa typing as described previously [spa gene amplified by PCR with forward (5\u2032-AGACGATCCTTCGGTGAGC-3\u2032) and reverse (5\u2032-GCTTTTGCAATGTCATTTACTG-3\u2032) primers. The purified PCR products were sequenced and then edited. The Ridom SpaServer database (http://www.spaserver.ridom.de) was applied to determine the spa type of each isolate. Each set of PCR reactions include a spa-type t008 isolate from our previous study as positive control sample (14), and a reaction mixture containing no template DNA as a control for possible false positive results.Multiplex PCR was performed for gr group . agr typeviously . In thispvl (31.8%) and 21 tst (24.7%).The sources of isolates included: skin and soft tissue wounds (44.7%), purulent exudates from wounds or abscesses 17.7%), urine (14.1%), blood (11.8%), sputum (8.2%), and other body fluids (3.5%). Out of 85 MSSA isolates, 29 isolates were obtained from hospital H1 (34.1%), 25 isolates from hospital H2 (29.4%), 20 isolates from hospital H3 (23.5%), and 11 isolates from hospital H4 (13%). Among 85 MSSA isolates tested, the highest rate of resistance was detected for penicillin (74.1%), and gentamicin (54.1%). , II (12.9%), and III (10.6%). spa results showed 16 types corresponding to nine clonal complexes (CCs), namely CC8 (42.3%), CC22 (11.8%), CC30 (10.6%), CC7 (9.4%), CC5 (7.1%), CC398 (7.1%), CC15 (5.9%), CC59 (3.5%), and CC45 (2.3%). spa type t037 was the most common spa type identified among 85 MSSA isolates, with a frequency of 22.3%, followed by t008 (11.8%), t021 and t091 (each 9.4%), t030 (8.2%), t034 (7%), t005 (5.9%), t084 and t002 (each 4.7%), t790 and t437 (each 3.5%), t1869, t045, and t015 (each 2.4%), t318 and t491 (each 1.2%) (Table\u00a05%), II 1.9%, and Inducible clindamycin resistance was observed in CC8-MSSA-t037 (n\u2009=\u20095), CC30-MSSA-t021 (n\u2009=\u20094) CC8-MSSA-t008 (n\u2009=\u20093) isolates; while constitutive resistance phenotype was observed in CC8-MSSA-t037 (n\u2009=\u20098), CC8-MSSA-t030 (n\u2009=\u20093), CC-MSSA-t034 (n\u2009=\u20094), CC-MSSA-t091 (n\u2009=\u20095), CC-MSSA-t045 (n\u2009=\u20094), CC-MSSA-t005 (n\u2009=\u20094), and CC-MSSA-t790 (n\u2009=\u20091) isolates. Detailed results of the genotype distribution are presented in Table\u00a0spa types among MSSA isolates. We detected CC8 in 42.3% of isolates. Although multi-resistant CC8 was previously reported as one of the main international CCs of MRSA, the predominance of the CC8-MSSA clone has previously been reported in Europe [Infection with MSSA as the most common pathogen in hospitalized patients is becoming increasingly problematic globally and requires special attention . As presn Europe  and Afrin Europe , 19. We n Europe , Irelandn Europe , and Nign Europe . We confn Europe , 21. As n Europe .We noted a relatively low prevalence of CC7 (9.4%), CC5 (7.1%), CC398 (7.1%), CC15 (5.9%), CC59 (3.5%), and CC45 (2.3%) in our study. A study conducted in China during the 4-year period indicated that CC22-t309 26.0%), CC188-t189 (5.1%), CC796-t796 (4.8%), CC121-t435 (4.8%), and CC398-t571 (3.6%) were the most dominant clones among MSSA isolates . They de.0%, CC18pvl encoding gene. Prevalence PVL positive MSSA strains has varied among studies from different geographic regions including China (34.4%) [Present results demonstrated that out of the 85 tested MSSA, 27 isolates 31.8%) carried 1.8% carr 34.4%) , Ireland (34.4%) , Colombi.4% , Ire (34.4%) , Russia  (34.4%) . In a re (34.4%) . However (34.4%) , 19. In  (34.4%) , 26, 28, (34.4%) .tst positive. This reported rate was different from the earlier studies in Africa (7%) [tst gene were associated with CC8-MSSA-t037 (11.8%), CC8-MSSA-t0307 (5.9%), CC22-MSSA-t790 (3.5%), and CC59-MSSA-t437 (3.5%) clones. A multicenter study\u00a0from china reported that 4.0% of MSSA isolates carried tst\u00a0gene which belonged to CC5 clone [tst-positive MSSA was distributed into four STs and the majority of them belonged to CC30 clone [tst encoding gene among S. aureus clinical isolates (21.3%) [tst were significantly higher in MSSA isolates in comparison to MRSA isolates (18.1% vs. 11.6%) [tst-positive MSSA clones seems geographically different and CC8 may represent a newly emerging clone in Iran.In our strain collection, 24.7% of isolates were ica (7%) , China (ica (7%)  and Turkica (7%) . Contrarica (7%) , 31 whicC5 clone . A recenSSA-t037 .8%, CC8- (21.3%) . In a st. 11.6%) . Overallagr genotypes are strictly associated with the clonal lineages [agr type I, as the most predominant type (76.4%), was associated with CC8, CC22, CC7, CC45, CC398, and CC59 isolates. In line with our results, Croes et al. demonstrated that CC7, CC8, CC22, CC25 and CC45 clonal lineages harbored agr I [agr type I (68.8%), followed by agr III (18.7%), and agr IV (12.5%) among MSSA isolates [According to the evidence, the lineages , 35. In ant type 6.4%, wasOur research had some limitations. Firstly, present work lacks detailed clinical information about the patients, Secondly, our samples were not collected consecutively. Thirdly, whole genome sequencing technique was not applied in the present work due to some of technical limitations."}
+{"text": "The results showed that a HSD, salt gene overexpression and dFOXO knockdown significantly reduced climbing endurance, climbing index, survival, dFOXO expression and SOD activity level, and increased malondialdehyde level in aging flies. Inversely, in a HSD aging flies, endurance exercise and dFOXO overexpression significantly increased their climbing ability, lifespan and antioxidant capacity, but they did not significantly change the salt gene expression. Overall, current results indicated that a HSD accelerated the age-related decline of climbing capacity and mortality via upregulating salt expression and inhibiting the dFOXO/SOD pathway. Increased dFOXO/SOD pathway activity played a key role in mediating endurance exercise resistance to the low salt tolerance-induced impairment of climbing capacity and longevity in aging Drosophila.A high-salt diet (HSD) is a major cause of many chronic and age-related defects such as myocardial hypertrophy, locomotor impairment and mortality. Exercise training can efficiently prevent and treat many chronic and age-related diseases. However, it remains unclear whether endurance exercise can resist HSD-induced impairment of climbing capacity and longevity in aging individuals. In our study, flies were given exercise training and fed a HSD from 1-week old to 5-weeks old. Overexpression or knockdown of This article has an associated First Person interview with the first author of the paper. Summary: Increased dFOXO/SOD pathway activity played a key role in mediating endurance exercise resistance to the salt tolerance-induced impairment of climbing capacity and longevity in aging Drosophila. Sodium chloride from dietary salt supplies essential electrolytes to the human body, and it plays a vital role in maintaining the stability of the intracellular and extracellular environments . DespiteExercise training is an efficient strategy for the prevention and treatment of many chronic diseases caused by diet or aging such as myocardial hypertrophy, hypertension and obesity . For exaDrosophila melanogaster provides significant practical advantages over other model systems to study the molecular mechanisms of exercise training, nutrition and aging, which include developed exercise and diet programs, a short lifespan (2\u20133\u2005months), low genetic redundancy compared with mammals, and a plethora of tools available to easily manipulate gene expression      , but a 2%-SD had no remarkable influence on CI (P>0.05)   , a 4%-SD and an 8%-SD significantly reduced the climbing fatigue time (CFT) (ctively) B. In 3-wctively) C. In 5-wctively) D. In addctively) E\u2013H. Nextctively) I. In 1-w(P>0.05) J. In 3-wctively) J. In 5-wctively) J. These Drosophila. In this experiment, the results showed that a 2%-SD significantly reduced the lifespan of Drosophila (P<0.05), and a 4%-SD and an 8%-SD acutely reduced lifespan of flies   K. These Drosophila  (P>0.05)  (P>0.05)  (P>0.05) , and senility significantly decreased their CI (P<0.01) (P<0.05), and senility significantly decreased their CI (P<0.01) (P<0.05), but the CI of 4%-SD and 5-week-old flies did not change significantly after exercise training (P>0.05), and senility significantly decreased their CI (P<0.01) (P>0.05), and senility significantly decreased their CI (P<0.01) (P<0.05) (P>0.05)  A\u2013C, but (P>0.05) D. Furthectively) E\u2013G, but (P>0.05) H. Moreovctively) I,J, and (P>0.05) K,L. The (P<0.01) M. In 2%-(P<0.01) N. In 2%-(P<0.01) O. The CI(P<0.01) P. What i(P<0.05) Q,U, but (P>0.05) V,W. ThesD. melanogaster , and the expression of Salt gene of 2%-SD flies, 4%-SD flies and 8%-SD flies did not change significantly after exercise training (P>0.05) (dFOXO gene (P<0.01), and exercise training significantly increased the dFOXO gene expression in the 0%-SD flies, 2%-SD flies and 4%-SD flies , but dFOXO gene expression of the 8%-SD flies did not change significantly after exercise training (P>0.05) (P<0.01), and exercise training significantly increased the SOD level of the 0%-SD flies, 2%-SD flies and 4%-SD flies , but the SOD level of the 8%-SD flies did not change significantly after exercise training (P>0.05) , and exercise training significantly decreased the MDA level of the 0%-SD flies, 2%-SD flies and 4%-SD flies (P<0.05), but the MDA level of the 8%-SD flies did not change significantly after exercise training (P>0.05)  A. In add(P>0.05) B. Moreov(P>0.05) C. Finall(P>0.05) D. These FOXO is associated with an increase in the expression of SOD and SOD activity , exercise training significantly increased the CFT (P<0.05) , exercise training significantly increased the CFT (P<0.05) , exercise training could not be said to significantly increase the CFT (P>0.05)  , but exercise training significantly increased the CI (P<0.05) , but exercise training significantly increased the CI (P<0.01) , but exercise training significantly increased the CI (P<0.01) , but exercise training significantly increased lifespan (P<0.05)  A. In 3-w(P<0.05) B. Howeve(P>0.05) C. Next, (P<0.01) D. In 1-w(P<0.05) E. In 3-w(P<0.01) E. In 5-w(P<0.01) E. Moreov(P<0.05) F,G. ThesSalt gene (P<0.01), but exercise training did not significantly decrease the expression of the Salt gene (P>0.05) , but exercise training significantly increased the expression of dFOXO gene (P<0.05) , but exercise training significantly increased the SOD level (P<0.01) , but exercise training significantly increased the MDA level (P<0.01)  H. In add(P<0.05) I. Moreov(P<0.01) J. Finall(P<0.01) K. These salt gene knockdown can improve salt tolerance, and it can increase the survival of a HSD fly , and a HSD also did not significantly reduce the CFT (P>0.05)  , and a HSD also did not significantly reduce the CI (P>0.05) , and a HSD also did not significantly reduce the lifespan (P>0.05) , but a HSD did not significantly increased the salt gene expression (P>0.05)   A\u2013C. In s(P<0.01) D. Howeve(P>0.05) E. What i(P>0.05) F,G, whic(P>0.05) . Next, t(P>0.05) H. Finall(P>0.05) I\u2013K. ThesSalt gene function takes part in regulating salt tolerance, and downstream it regulates antioxidant function, climbing capacity and aging.Two principal classes of manipulation are usually employed to study gene function. Loss-of-function (LOF) approaches attempt to eliminate gene function partially or completely. Gain-of-function (GOF) approaches attempt to obtain functional information by creating conditions where the gene is excessively or ectopically expressed or its function exaggerated . In thisdFOXO gene expression was also changed by UAS/Gal4 system in flies, and their CFT, CI and lifespan were measured.In animals, it has been identified that a HSD increases the oxidative stress and causes cell damage . In fliedFOXO gene overexpression significantly increased the CFT (P<0.05), and it significantly increased the CFT in HSD 1-week-old flies (P<0.001) (dFOXO gene overexpression significantly increased the CFT (P<0.05), and it also significantly increased the CFT in HSD 3-week-old flies (P<0.001) (dFOXO gene overexpression significantly increased the CFT (P<0.05), and it also significantly increased the CFT in HSD 5-week-old flies (P<0.01)  (dFOXO gene significantly increased the CI (P<0.05), and it also significantly increased the CI in HSD 1-week-old flies (P<0.05) (dFOXO gene significantly increased the CI (P<0.05), and it also significantly increased the CI in HSD 3-week-old flies (P<0.01) (dFOXO gene significantly increased the CI (P<0.01), and it also significantly increased the CI in HSD 5-week-old flies (P<0.01) (dFOXO gene significantly increased the lifespan of flies (P<0.05), and it also significantly increased the lifespan of HSD flies (P<0.01)  A. In 3-wP<0.001) B. In 5-w(P<0.01) C. Moreov(P<0.01) D. In 1-w(P<0.05) E. In 3-w(P<0.01) E. In 5-w(P<0.01) E. Furthe(P<0.01) F,G. ThesdFOXO gene (P<0.01), it also significantly increase the dFOXO gene expression in HSD flies (P<0.01) , and it also significantly did not change the salt gene expression in HSD flies (P>0.05) (dFOXO gene significantly increased the SOD level (P<0.01), and it also significantly increased the SOD level in HSD flies (P<0.01) (dFOXO gene significantly decreased the MDA level (P<0.01), and it also significantly increased the MDA level in HSD flies (P<0.01)  I. In add(P>0.05) H. Howeve(P<0.01) J. Finall(P<0.01) K. These dFOXO gene knockdown significantly decreased the CFT and CI in flies   , and a HSD aggravated the lifespan reduction in dFOXO gene knockdown flies (P<0.01) , but a HSD significantly increased the salt gene expression in dFOXO gene knockdown flies (P<0.01) (dFOXO gene RNAi significantly decreased the dFOXO gene expression (P<0.01), and a HSD aggravated dFOXO gene expression reduction in dFOXO gene knockdown flies (P<0.01) (dFOXO gene knockdown significantly decreased the SOD activity level (P<0.01), and a HSD aggravated SOD activity level reduction in dFOXO gene knockdown flies (P<0.01) (dFOXO gene knockdown significantly increased the MDA level (P<0.01), and a HSD aggravated MDA level increase in dFOXO gene knockdown flies (P<0.01)  A\u2013E, and P<0.001) A\u2013E. More(P<0.01) F,G. Howe(P<0.01) H. Next, (P<0.01) I. Simila(P<0.01) J. Finall(P<0.01) K. TherefdFOXO gene function is closely to lifespan in flies. For instance, the dFOXO overexpression decreased mortality and increased lifespan of flies via reducing oxidative stress, and the dFOXO is required both for transcriptional changes that mark the fly's dietary history and for nutritional programming of lifespan by excess dietary sugar . The UAS-salt-overexpression flies , the UAS-dFOXO-overexpression flies , and the arm-gal4  flies were obtained from the Bloomington Stock Center. The UAS-salt-knockdown flies  and the UAS-dFOXO-knockdown flies were obtained from the Vienna Drosophila Resource Center .The KO saltmir-1014-KO\u2019 and \u2018arm-gal4>w*; TI /total dead numbers. Sample sizes were 200\u2013210 flies per group  method. Ten fresh-frozen flies were converted to homogenates in a homogenizer filled with 1\u2005ml PBS (pH 7.2\u20137.4). The homogenates were centrifuged at 4\u00b0C for 15\u2005min with a speed of 2000\u2005r/min. The supernatant was mixed with the reagents supplied in an MDA Assay Kit  and incubated at 95\u00b0C for 40\u2005min. After cooling at room temperature, the mixture was centrifuged at 4000\u2005The total SOD activity in flies was determined using the hydroxylamine method. Ten fresh-frozen flies were converted to homogenates in a homogenizer filled with 1\u2005ml PBS (PH7.2\u20137.4). The homogenates were centrifuged at 4\u00b0C for 15\u2005min with a speed of 2000\u2005r/min. The supernatant was mixed with the reagents supplied in a SOD Assay Kit . The mixture was incubated at room temperature for 10\u2005min, and the absorbance of the compound was then measured at 550\u2005nm. All operations were according to the manufacturer's instructions. Each preparation was tested in triplicate. SOD activity was expressed as U/ml. All assays were repeated three times.salt were as follows: F: 5\u2032-TTAATCGCAGGCGCGTCAGTG-3\u2032; R: 5\u2032-GGACGAGACCACCGTGTTAATCAG-3\u2032. Primer sequences of dFOXO were as follows: F: 5\u2032-AACAACAGCAGCATCAGCAG-3\u2032; R: 5\u2032-CTGAACCCGAGCATTCAGAT-3\u2032. Primer sequences of Rp49 were as follows: F:5\u2032-CTAAGCTGTCGCACAAATGG-3\u2032; R:5\u2032-AACTTCTTGAATCCGGTGGG-3\u2032.About ten flies were homogenized in Trizol. First, 10\u2005\u03bcg of the total RNA was purified by organic solvent extraction from the Trizol . The purified RNA was treated with DNase I  and used to produce oligo dT-primed cDNAs , which were then used as templates for quantitative real-time PCR. The rp49 gene was used as an internal reference for normalizing the quantity of total RNAs. Real-time PCR was performed with SYBR green using an ABI7300 Real time PCR Instrument (Applied Biosystems), with three biological replicates. Expression of the various genes was determined by the comparative CT method . Primer sequences of dFOXO overexpression on flies. Using a non-parametric followed by a log-rank test for analyzing survival and time to fatigue. Analyses were performed using the Statistical Package for the Social Sciences (SPSS) version 16.0 for Windows , with statistical significance set at P<0.05. Data are represented as means\u00b1s.e.m.The one-way ANOVA with LSD tests were used to identify differences among the relevant groups. A two-way ANOVA was used to analyze the effects of a HSD and"}
+{"text": "Correction to: Glob Health Res Policy (2020) 5:54https://doi.org/10.1186/s41256-020-00183-yAfter publication of this article , it is nThe correct reference 7 is below:Wang X, Yu Y, Hu Y, Yu C. COVID-19 analysis and forecast based on exponential smoothing model in Hubei Province. J Public Health Prev Med. 2020;31(1):1\u20134. (in Chinese)The original article has been updated."}
+{"text": "Correction to: Mol Cancer (2019) 18:126https://doi.org/10.1186/s12943-019-1054-7After publication of the article , the autThe correct figures are updated below.Fig.3i and jFig.4hAdditional file 3: Figure S2d"}
+{"text": "Candidatus Dehalogenimonas etheniformans\u201d strain GP couples growth with the reductive dechlorination of vinyl chloride and several polychlorinated ethenes. The genome sequence comprises a circular 2.07-Mb chromosome with a G+C content of 51.9% and harbors 50 putative reductive dehalogenase genes.\u201c Candidatus Dehalogenimonas etheniformans\u201d strain GP couples growth with the reductive dechlorination of vinyl chloride and several polychlorinated ethenes. The genome sequence comprises a circular 2.07-Mb chromosome with a G+C content of 51.9% and harbors 50 putative reductive dehalogenase genes.\u201c Dehalogenimonas are obligate organohalide-respiring bacteria implicated in the turnover of naturally occurring and anthropogenic chlorinated compounds in anoxic environments . Paired-end sequencing (2\u2009\u00d7\u2009150\u2009bp) was performed using a NovaSeq PE150 flow cell . The sequence reads were trimmed and filtered (https://github.com/lh3/readfq), resulting in 4,073,240 high-quality reads. Assembly using NanoPore raw long reads  and Illumina short reads  was performed using Unicycler version 0.4.7 (Genomic DNA from strain GP biomass was extracted using the cetyltrimethylammonium bromide method . For Nanon 0.4.7 . Unicyclon 0.4.7 .rdh) subunit A genes, including cerA (locus tag number HX448_10020), which encodes a VC rdh  of D. formicexedens strain NSZ-14T (D. formicexedens strain NSZ-14T (GenBank accession number CP018258.1), D. alkenigignens strain IP3-3T (shotgun sequencing project accession number LFDV00000000.1), Dehalogenimonas sp. strain WBC-2 (GenBank accession number CP011392.1), and D. lykanthroporepellens strain BL-DC-9T (GenBank accession number CP002084.1), respectively. The new data expand the Dehalogenimonas pangenome and rdh sequence diversity.The genome comprises one circular 2,068,322-bp chromosome with a G+C content of 51.9%. The genome contains 2,029 coding sequences, 47 tRNA genes, and single copies of the 5S, 16S, and 23S rRNA genes. The GP genome harbors 50 nonidentical reductive dehalogenase (CP058566.1). The BioSample and BioProject accession numbers are SAMN15398252 and PRJNA258024, respectively. The raw reads were deposited in the Sequence Read Archive under accession numbers SRR12774736 (Nanopore) and SRR12774735 (Illumina).The genome has been deposited at the DNA Data Bank of Japan, the European Nucleotide Archive, and GenBank (accession number"}
+{"text": "Solanum tuberosum L., is the No. 1 vegetable crop and a critical food security crop. The genome sequence of DM1\u20133 516 R44, a doubled monoploid clone of S. tuberosum Group Phureja, was published in 2011 using a whole-genome shotgun sequencing approach with short-read sequence data. Current advanced sequencing technologies now permit generation of near-complete, high-quality chromosome-scale genome assemblies at minimal cost.Worldwide, the cultivated potato, Here, we present an updated version of the DM1\u20133 516 R44 genome sequence (v6.1) using Oxford Nanopore Technologies long reads coupled with proximity-by-ligation scaffolding (Hi-C), yielding a chromosome-scale assembly. The new (v6.1) assembly represents 741.6 Mb of sequence (87.8%) of the estimated 844 Mb genome, of which 741.5 Mb is non-gapped with 731.2 Mb anchored to the 12 chromosomes. Use of Oxford Nanopore Technologies full-length complementary DNA sequencing enabled annotation of 32,917 high-confidence protein-coding genes encoding 44,851 gene models that had a significantly improved representation of conserved orthologs compared with the previous annotation. The new assembly has improved contiguity with a 595-fold increase in N50 contig size, 99% reduction in the number of contigs, a 44-fold increase in N50 scaffold size, and an LTR Assembly Index score of 13.56, placing it in the category of reference genome quality. The improved assembly also permitted annotation of the centromeres via alignment to sequencing reads derived from CENH3 nucleosomes.Access to advanced sequencing technologies and improved software permitted generation of a high-quality, long-read, chromosome-scale assembly and improved annotation dataset for the reference genotype of potato that will facilitate research aimed at improving agronomic traits and understanding genome evolution. Solanum tuberosum L., NCBI:txid4113) was published in 2011 by the Potato Genome Sequencing Consortium (PGSC) using a whole-genome shotgun sequencing approach [The genome of the vegetable crop potato . Howeveg; a total of 6.2 g of shoot tissue was split across 6 separate nuclei isolations. Modifications to the protocol include squeezing the homogenate through 5 layers of Miracloth instead of gravity filtering alone and 2 washes with nuclear isolation buffer. DNA was isolated from nuclei using the Nanobind Plant Nuclei Big DNA\u2014Alpha Version kit  following the Nanobind Plant Nuclei Big DNA Kit Handbook v0.17 (May 2018). DNA libraries were prepared using the ONT SQK-LSK109 Ligation Sequencing kit . Six libraries were prepared and sequenced on 6 separate R9 ONT flow cells . DNA repair and end preparation  were performed with an input of 1 \u03bcg of DNA. The repair and end preparation reaction were incubated for 5\u201345 minutes at 20\u00b0C and 5\u201345 minutes at 60\u00b0C. The reaction was cleaned using Agencourt AMPure XP beads  with an incubation time of 5\u201310 minutes on a rotator mixer and eluted for 2\u20135 minutes. Ligation of adapters to the prepared DNA was performed at room temperature for 10\u201360 minutes. The ligation reaction was cleaned using Agencourt AMPure XP beads on a rotator mixer with an incubation time of 5\u201310 minutes with an elution time of 10 minutes. Sequencing was performed on an ONT MinION   with the current release of MinKNOW (version 1.15.0). Sequencing was run for 48\u201392 hours (RRID:SCR_016383) in paired-end mode, generating 150\u00a0nt reads (DM plants were grown in Murashige and Skoog (MS) medium , shoots harvested, and flash frozen in liquid nitrogen. Nuclei were isolated following the Workman et al.  protocol92 hours . DNA wasnt reads . Hi-C lint reads .RRID:SCR_018927) [RRID:SCR_017016) [RRID:SCR_017642) [RRID:SCR_018550) [RRID:SCR_016157) [RRID:SCR_002105) [RRID:SCR_014731) [RRID:SCR_011841) [RRID:SCR_010910) [RRID:SCR_006525) [RRID:SCR_006525), all using default parameters. Pilon was run using the \u201c\u2013fix bases\u201d option. The polished contigs are composed of 1,382 contigs with a total size of 745.6 Mb with an N50 contig size of 17.3 Mb and a maximum contig length of 42.1 Mb  on an A_018927)  to remov_017016)  with the_017642) . For eac_018550)  with the_016157) . Reads w_016157)  with the_002105) . An upda_014731) . The Ill_011841)  to remov_006525) , and theRRID:SCR_017226) [RRID:SCR_017227) [RRID:SCR_016665) [S. tuberosum mitochondrion genome  using blastn v2.9.0  [To construct chromosome-scale pseudomolecules, the Hi-C library was first processed using the juicer.sh pipeline from the Juicer package (git commit 6403a27)  . The pse_017227)  and the _017227)  (Supplem_017227)  database_016665)  with the_001598) . FifteenRRID:SCR_011841) [RRID:SCR_010910) [RRID:SCR_015008) [k-mers (k = 21) from the cleaned Illumina whole-genome shotgun library (PEP_AA_01) using Jellyfish2 v2.2.10  [k-mer count histogram was analyzed by the online version of GenomeScope  [To assess completeness and accuracy of the v6.1 assembly, \u223c458 million paired-end reads from a whole-genome Illumina sequencing library   was used_010910) . Alignme_015008)  software_015008) . Of 1,61_005491) . The k-m_017014)  and the RRID:SCR_018970) [RRID:SCR_018969) [RRID:SCR_017623) [Arabidopsis thaliana TAIR10 (LAI = 14.9), Fragaria vesca v4.1 (LAI = 16.9), and Solanum pennellii (LAI = 14.8) [The Long Terminal Repeat (LTR) Assembly Index (LAI)  metric w_018970) , LTR_FIN_018969) , and LTR_017623) . LTR seq = 14.8) . The LAIin situ hybridization (oligo-FISH) probes, which mark 26 regions on the 12 chromosomes, have been used to characterize potato karyotypic variation [Solanum species. We aligned the oligo-FISH probes to v6.1 using BWA-MEM v0.7.12-r1039  [Two \u201cbarcode\u201d oligonucleotide fluorescent ariation , as well_010910)  to confiRRID:SCR_018968) [RRID:SCR_010910) [RRID:SCR_006646) [Cen12 is properly positioned on the short arm of v6.1 chromosome 12   with 200_018968) . ChIP-se_010910)  using de 12 Fig.\u00a0. The imp6.1 Fig.\u00a0. In v4.0nce Fig.\u00a0. The siznce Fig.\u00a0. These cRRID:SCR_018967) [To better depict the improved contiguity and accuracy of v6.1 relative to v4.04, D-GENIES   was usedRRID:SCR_015027) [RRID:SCR_001653) [RRID:SCR_012954) [A custom repeat library (CRL) was generated using RepeatModeler2 v2.0.1   with the_015027)  using bl_001653)  with an Cen4, Cen6, Cen10, Cen11, and Cen12) lack typical centromere-specific satellite repeats and, instead, are composed of single- or low-copy sequences resembling neocentromeres [Cen1, Cen2, Cen3, Cen5, Cen7, and Cen8) that contain megabase arrays of satellite repeats. Interestingly, the satellite repeats for these 6 centromeres are unique to individual chromosomes, some of which are derived from retrotransposons. Centromeric repeat sequences from Gong et al. [RRID:SCR_004870) [Cen2, Cen5, and Cen7 in v.6.1 but not in v4.04   with aliRRID:SCR_018966) [RRID:SCR_018927) [RRID:SCR_018550) with the parameters -a -x splice -uf -G 5000; 5,783,924 (99.98%) of the 5,784,833 filtered reads aligned to the DM assembly. The cDNA alignments were assembled using Stringtie2 v2.1.2  [RRID:SCR_018965) [RRID:SCR_011841) [RRID:SCR_015530) [RRID:SCR_016323) [RRID:SCR_018965) [To facilitate annotation of gene models, ONT complementary DNA (cDNA) sequences were generated from DM. DM was grown under a 16-hour day length in tissue culture and RNA was isolated from whole tissue-culture plants using a modified hot borate method . DNA con_018966)  to ident_018927)  to remov_016323)  (-L -m 5_018965) . Illumin_011841)  with the_015530)  with the_016323)  with the_018965) . Both thRRID:SCR_018964) [RRID:SCR_008417) [RRID:SCR_011930) [ab initio gene predictions. The BRAKER2 pipeline was run using the command line: braker.pl \u2013species = DM_v6_1 \u2013gff3 \u2013softmasking \u2013UTR = off \u2013bam {RNA-seq.alns.bam}. Ab initio gene predictions were refined using PASA2 v2.4.1  [RRID:SCR_004726) [RRID:SCR_005305) [RRID:SCR_016582) [The BRAKER2 (git commit 6219573)   gene pre_011930)  and the _005305)  with a c_016582) .Arabidopsis proteome  [RRID:SCR_004726) [RRID:SCR_002380). Search results were processed in the same order, and the function of the first hit encountered was assigned to the gene model. The quality of the annotation was evaluated using BUSCO [High-confidence gene models were defined as having a TPM value >\u00a00 in \u22651 RNA-Seq library and/or having a PFAM domain match. Gene models that were partial or had matches to transposable element\u2013related PFAM domains were excluded from the high-confidence model set. A total of 32,917 loci encoding 44,851 gene models are contained within the high-confidence set . To assi_004618) , the PFA_004726) , and theng BUSCO , and botUsing improved sequencing technologies, the genome sequence of the reference potato genotype DM was vastly improved in contiguity relative to the previous release, DM v4.04. Version 6.1 of the DM genome assembly represents 87.8% of the estimated genome, with 595-fold increase in N50 contig size, 99% reduction in number of contigs, and a 44-fold increase in N50 scaffold size. Importantly, 731.2 Mb of the 741.6-Mb assembly is non-gapped and anchored to the 12 chromosomes, indicating a high degree of contiguity that was reflected in a \u201creference quality\u201d LAI score, demonstrating the ability of advanced sequencing methods to assemble large contiguous regions of a medium-sized plant genome. With access to full-length cDNA sequences, 32,917 high-confidence protein-coding genes encoding 44,851 gene models were annotated, which provided a substantial improvement in representation of conserved orthologs compared with the previous annotation that will facilitate future studies in potato biology, genetics, and genomics.GigaScience GigaDB [The clone, DM1\u20133516 R44, is available through the United States Department of Agriculture Potato Genebank via PI GS 233 . The rawe GigaDB , Dryad De GigaDB , and on e GigaDB , 73 via Supplementary Figure S1. Hi-C contact map showing the inter- and intra-chromosomal chromatin interactions in DM v6.1. Inter-chromosomal chromatin interactions are off the diagonal axis and intra-chromosomal chromatin interactions are within the blue boxes. Each pixel represents the degree of interaction between each 1-Mb locus, with a dark red color indicating a greater number of reads involved in the interaction. The blue boxes represent the boundaries of each pseudomolecule, and individual scaffold boundaries are represented by the green boxes.k-mer of 21.Supplementary Figure S2. Estimation of heterozygosity of the DM genome as determined by GenomeScope. The DM genome has an estimated heterozygosity rate of 0.0383% using a RRID:SCR_018968) [Supplementary Figure S3. Mapping of the DM \u00d7 RH F1 population markers to the (a) DM v4.04 and the (b) DM v6.1 assembly. Flanking sequence (200\u00a0nt) of the markers was used for sequence alignments to the assembly using Vmatch  . The y-aSupplementary Table S1. Sequence datasets used in this study. Total reads for Oxford Nanopore Technologies sequencing are passed reads after base-calling.Supplementary Table S2. Oxford Nanopore Technologies whole-genome shotgun sequence reads used in the DM v6.1 assembly.Supplementary Table S3. Illumina whole-genome shotgun sequence read mapping statistics.Supplementary Table S4. BUSCO  results Supplementary Table S5. Centromere positions in the DM v6.1 assembly.Supplementary Table S6. Repetitive sequence content in v4.04 and v6.1 DM 1\u20133516 R44 genome assemblies.Supplementary Table S7. DM v6.1 gene annotation summary.giaa100_GIGA-D-20-00167_Original_SubmissionClick here for additional data file.giaa100_GIGA-D-20-00167_Revision_1Click here for additional data file.giaa100_Response_to_Reviewer_Comments_Original_SubmissionClick here for additional data file.giaa100_Reviewer_1_Report_Original_SubmissionJian-Feng Mao, Ph.D. -- 7/7/2020 ReviewedClick here for additional data file.giaa100_Reviewer_2_Report_Original_SubmissionAlexis Sullivan -- 8/10/2020 ReviewedClick here for additional data file.giaa100_Supplemental_FilesClick here for additional data file.in situ hybridization; ONT: Oxford Nanopore Technologies; PASA: Program to Assemble Spliced Alignments; PGSC: Potato Genome Sequencing Consortium; RNA-Seq: RNA-sequencing; SRA: Sequence Read Archive; TPM: transcripts per million.BLAST: Basic Local Alignment Search Tool; bp: base pairs; BUSCO: Benchmarking Universal Single-Copy Orthologs; cDNA: complementary DNA; ChIP-Seq: chromatin immunoprecipation sequencing; CRL: custom repeat library; Gb: gigabase pairs; kb: kilobase pairs; LAI: LTR Assembly Index; LTR: long terminal repeat; Mb: megabase pairs; mRNA: messenger RNA; NCBI: National Center for Biotechnology Information; nt: nucleotide; oligo-FISH: oligonucleotide fluorescent The authors declare that they have no competing interests.C.R.B. conceived the study. G.M.P., J.P.H., B.V., and J.C.W. performed the experiments. J.T.B., J.P.H., J.J., G.M.P., S.O., B.V., J.C.W., and H.Z. analyzed data. C.R.B., J.P.H., J.J., G.M.P., B.V., J.C.W., and H.Z. wrote the manuscript. All authors approved the final manuscript."}
+{"text": "Ephemerella sp. Yunnan-2018, Serratella zapekinae, Serratella sp. Yunnan-2018, Serratella sp. Liaoning-2019, Torleya grandipennis and T. tumiforceps. These mitogenomes were employed to reveal controversial phylogenetic relationships among the Ephemeroptera, with emphasis on the phylogenetic relationships among Ephemerellidae. The lengths of the six mayfly mitogenomes ranged from 15,134 bp to 15,703 bp. Four mitogenomes of Ephemerella sp. Yunnan-2018, Serratella zapekinae, Serratella sp. Yunnan-2018 and Serratella sp. Liaoning-2019 had 22 tRNAs including an inversion and translocation of trnI. By contrast, the mitogenomes of T. tumiforceps and T. grandipennis had 24 tRNAs due to an extra two copies of inversion and translocation of trnI. Within the family Ephemerellidae, disparate gene rearrangement occurred in the mitogenomes of different genera: one copy of inversion and translocation trnI in the genera Ephemerella and Serratella, and three repeat copies of inversion and translocation of trnI in the genus Torleya. A large non-coding region (\u2265200 bp) between trnS1 (AGN) and trnE was detected in T. grandipennis and T. tumiforceps. Among the phylogenetic relationship of the Ephemeroptera, the monophyly of almost all families except Siphlonuridae was supported by BI and ML analyses. The phylogenetic results indicated that Ephemerellidae was the sister clade to Vietnamellidae whereas Teloganellidae was not a sister clade of Ephemerellidae and Vietnamellidae.As a small order of Pterygota (Insecta), Ephemeroptera has almost 3,500 species around the world. Ephemerellidae is a widely distributed common group of Ephemeroptera. However, the relationship among Ephemerellidae, Vietnamellidae and Teloganellidae is still in dispute. In this study, we sequenced six complete mitogenomes of three genera from Ephemerellidae (Insecta: Ephemeroptera):  BecauseSiphluriscus chinensis (Siphluriscidae) had one extra trnK-like gene (trnK 2 (AAA)  had an inversion of trnI (Alainites yixiani (GU479735) (Baetidae) showed a gene arrangement of trnI-trnW-trnQ-trnY-trnM. Therefore, these different sets of data suggested that specific gene rearrangements may occur in the mitogenomes of different families. The present study is important and original in that it explores the relationships between mitogenome rearrangements and taxonomic categories of Ephemeroptera.Gene rearrangements can be frequently observed in insect mitogenomes , whereas 2 (AAA) . Within  of trnI . AdditiotrnI inversion in the Family Ephemerellidae and explore the phylogenetic relationships of Ephemerellidae, we sequenced six complete mitogenomes belonging to three genera of Ephemerellidae. The compositional and structural features of these six mitogenomes are described and gene rearrangements were analyzed to explain the mechanism of mitochondrial gene rearrangements.In order to discuss the characteristics of Ephemerella sp. Yunnan-2018, S.\u00a0zapekinae, Serratella sp. Yunnan-2018, Serratella sp. Liaoning-2019, T.\u00a0grandipennis and T.\u00a0tumiforceps were separately captured from Ning\u2019er Yunnan province, Xiuyan Liaoning province, Ning\u2019er Yunnan province, Kuandian Liaoning province, Longquan Zhejiang province and Tonglu Zhejiang province, China, respectively. After morphological identification, the specimens were deposited at \u221240\u00a0\u00b0C in the Animal Specimen Museum, College of Life Sciences and Chemistry, Zhejiang Normal University, China. Total genomic DNA was extracted from tissues of each complete specimen by Ezup Column Animal Genomic DNA Purification Kit . The Animal Research Ethics Committees of Zhejiang Normal University approved the experimental design.The voucher specimens of Takara Taq and Takara LA Taq DNA polymerase in a reaction volume of 50 \u03bcL. The amplifications for both normal PCR and Long-PCR were performed under the conditions as described in Several partial fragments were amplified using common primers , as deschttp://mitos.bioinf.uni-leipzig.de/index.py) (http://rna.tbi.univie.ac.at/forna) (12S and 16S rRNA). The trnI sequences of T.\u00a0grandipennis and T.\u00a0tumiforceps were aligned using the Clustal W program implemented in Mega 7.0 (MT274127\u2013MT274132. The mitogenome maps of the six mayfly species were drawn using GenomeVx (http://wolfe.ucd.ie/GenomeVx) (http://tandem.bu.edu/trf/trf.submit.options.html)   using tht/forna) . After ut/forna)  to compaMega 7.0 . Then, wMega 7.0 . The sixenomeVx) . Codon uenomeVx) . Using tenomeVx) , we calcns.html) . Additions.html) .Siphluriscus chinensis acted as the outgroup  to \u22120.017 (Serratella sp. Yunnan-2018) whereas the GC-skew for the majority strand ranged from \u22120.234 (Ephemerella sp. Yunnan-2018) to \u22120.159 (S. zapekinae). In addition, we analyzed the sizes and nucleotide compositions of the previously published mayfly mitogenomes. Differences in mitochondrial sequences of Ephemeroptera were mainly determined by different sizes of the CR. Among all sequenced Ephemeroptera mitogenomes  was the longest because of the longest CR  (Baetis sp. PC-2010 (GU936204)  was the shortest CR (340 bp). The AT-skews of mitogenomes on the majority strand were negative ranging from Baetis sp. PC-2010 (GU936204) (\u22120.093) to Caenis sp. YJ-2009 (GQ502451) (\u22120.001) except for positive skews in Ephemera orientalis (EU591678) (0.03), S. chinensis (0.019) (Ameletus sp. MT-2014 (0.01) (Habrophlebiodes zijinensis (GU936203) (\u22120.296) to S. chinensis (\u22120.139) (Baetis sp. PC-2010 (GU936204) (0.006), A. yixiani (GU479735) (0.14) and Ameletus sp. MT-2014 (0.202)  , whereas (0.019)  and Amel4 (0.01) . The GC-(\u22120.139)  except f (0.202) .ND3 (Ephemerella sp. Yunnan-2018) and ND6  along with GTG assigned to ND1 (Serratella sp. Yunnan-2018), ND3 (Serratella sp. Yunnan-2018), ND4 (Serratella sp. Liaoning-2019), and ND5 . Typical stop codons (TAA/TAG) were found in most PCGs, except for incomplete terminal codons, such as T used for COX2 , COX3 , Cyt b , ND4 (Serratella sp. Liaoning-2019), ND5  and ND6 (Serratella sp. Liaoning-2019), as well as TA used for ND4 . Incomplete terminal codons perform a significant function in polycistronic transcription cleavage and polyadenylation processes  of the six mayfly mitogenomes . The anaEphemerella sp. Yunnan-2018, S. zapekinae, Serratella sp. Yunnan-2018 and Serratella sp. Liaoning-2019 had 22 tRNA genes  in S. zapekinae (trnQ and U-G in the T \u03c8C (T) arm of trnN and trnS1 (AGN). In general, these mismatches are fundamental units of tRNA secondary structure and are nearly isomorphic to normal base pairs, as commonly observed in mayflies or other insects  and Epeorus sp. MT-2014 (A. yixiani ( GU479735)]. It has also been observed in other insect orders such as Mantodea , such as Anthonomus rectirostris, A. rubi, A. eugenii and A.\u00a0pomorum  and trnV. Unexpectedly, the 12S rRNA was located between trnV and trnI due to the translocation of trnI. The sizes of the 16S rRNA genes in these six mayfly species varied from 1,212 bp (T. grandipennis) to 1,230 bp (Serratella sp. Yunnan-2018), and the sizes of 12S rRNA ranged from 769 bp (T. grandipennis and T. tumiforceps) to 778 bp (Serratella sp. Yunnan-2018). Their length fit within the lengths detected in other Ephemeroptera mitogenomes (Serratella sp. Yunnan-2018 had the highest A+T content of the rRNA genes (68.8%) whereas the mitogenome of T. grandipennis had the lowest (63.1%). In the six mayfly mitogenomes, the AT bias of both 16S rRNA and 12S rRNA was slightly positive (\u22640.075), whereas the GC bias was strongly positive (\u22650.22), which revealed that the contents of A and G outnumbered T and C nucleotides, respectively  to 68.5% (Serratella sp. Liaoning-2019)  . The CRsng-2019) . We obseapekinae . It has apekinae . The secapekinae , are shoslippage . These sslippage .Ephemerella sp. Yunnan-2018, S. zapekinae and Serratella sp. Yunnan-2018 were obviously lower than the A+T content of corresponding whole mitogenomes, which was also observed in other mayflies  (Mecopoda niponensis) and Hemiptera  , Caenis sp. JYZ-2018  and Epeorus sp. MT-2014 (Siphlonurus immanis (FJ606783)] and Isonychiidae [Isonychia ignota (HM143892)].Interestingly, the A+T content of CRs in us youi) . This phrostris) . The abnrostris) . In the of 43.6% . Also, tsequence . Moreove insects . HoweverJYZ-2018  and CaenJYZ-2020 ], Heptag MT-2014 ], SiphloS. zapekinae, Serratella sp. Yunnan-2018 and Serratella sp. Liaoning-2019 contained 4, 6, and 6 non-coding intergenic spacer sequences, respectively, with total lengths of 22 bp, 32 bp and 29 bp, whereas Ephemerella sp. Yunnan-2018 had 7 non-coding intergenic spacer sequences of 92 bp in total length due to an intergenic spacer of 68 bp between ND4L and trnT. An intergenic spacer between trnS2 (UCN) and ND1 ranging from 17 bp to 19 bp was also detected in these four mitogenomes and is commonly found in Ephemeroptera. In terms of the intergenic spacer between ND4L and trnT in Ephemerella sp. Yunnan-2018, we also detected a similar sequence of 62 bp with a similarity of 72.58% in Ephemerella sp. MT-2014 , we observed a large intergenic spacer between trnS1 (AGN) and trnE within the mitogenomes of T. tumiforceps and T. grandipennis, with lengths of 531 bp and 200 bp, respectively. The large intergenic region in T. tumiforceps and T. grandipennis had 52.17% and 52% A+T content, respectively, with a strongly positive GC-skew (0.31 and 0.13). The large intergenic region in T. tumiforceps and T. grandipennis had a similar starting sequence of 20 bp (GGGSCKAAAGGSCCTACCTA) and a conserved sequence of 19 bp (TTTTTAGCGAACTTAGGGG), respectively, with 4 tandem repeats of 87 bp and 3 tandem repeats of 42 bp along with 4 partial sequences, respectively. The repeat unit of T. tumiforceps could be folded into three stem-loop secondary structures, whereas T. grandipennis folded into two stem-loop secondary structures  and trnE could be synapomorphic for the genus Torleya owing to the fact that sequences located between trnS1 (AGN) and trnE within mitogenomes of other Ephemeroptera were short (<13 bp) and the large intergenic spacer has only been found in Torleya. This is the first finding of a large intergenic region in the mitogenomes of mayflies. As a consequence of the non-coding particularity of the intergenic spacer sequence, these species may well have gone through further sequence divergence swiftly ], whereas an intergenic spacer between trnQ and trnM ranging from 26 bp to 53 bp was found in Caenidae  and Ephemeridae [E. orientalis (EU591678)]. In terms of the mitogenomes of Heptageniidae, we detected an intergenic spacer between trnA and trnR, ranging from 33 bp to 47 bp ))), as reported by P. youi + (P. cupulatus + (Epeorus sp. JZ-2014 + (Epeorus sp. MT-2014 + E. herklotsi)))), as reported by P. youi, P. cupulatus and (Epeorus sp. JZ-2014 + (Epeorus sp. MT-2014 + E. herklotsi)), with low support. On the whole, the monophyly of all Ephemeroptera families was supported except the family Siphluriscidae.Derived from both BI and ML analyses, the phylogenetic trees showed an identical topology, excluding the internal relationship among Heptageniidae  and 5. T12S, 16S, 18S, 28S and H3 genes, Ephemerellidae was supported as a monophyletic group, and their relationship with Teloganellidae and Vietnamellidae remains problematic  + Serratella sp. Yunnan-2018) + (T. tumiforceps + T. grandipennis)) + (Ephemerella sp. Yunnan-2018 + Ephemerella sp. MT-2014)). The inversion and translocation of one copy of trnI was found in Ephemerella and Serratella, whereas the inversion and translocation of three copies of trnI was found in Torleya (Serratella + Torleya) + Ephemerella), we can deduce that the inversion and translocation of one copy of trnI is characteristic of the ancestral Ephemerellidae. Unfortunately, the presence of only one mitogenome of Teloganellidae restricted a discussion of its monophyly, which requires more species to be added.Our findings indicated that Ephemerellidae was the sister clade to Vietnamellidae, but Teloganellidae was the sister clade to Baetidae , which mblematic . In addi Torleya . AccordiEphemerella sp. Yunnan-2018, S. zapekinae, Serratella sp. Yunnan-2018, Serratella sp. Liaoning-2019, T. grandipennis and T. tumiforceps. The mitogenomes of the genera Ephemerella and Serratella showed similar gene features to Ephemerella sp. MT-2014 (KM244691) and had the same inversion and translocation of trnI, whereas two extra copies of trnI were found in the genus Torleya. The translocation and extra copies of trnI could be explained by the tandem duplication/random loss model, whereas the inversion of trnI could be interpreted by the mitogenome recombination model. The evidence from this study suggests that these specific mitogenome rearrangements may be molecular markers of the family Ephemerellidae, especially the three copies of trnI as a synapomorphy for the genus Torleya. The occurrences and mechanisms of the large intergenic region between trnS1 (AGN) and trnE, detected in T. tumiforceps and T. grandipennis, require more mitogenomes of Torleya species to be investigated. Phylogenetic analyses based on BI and ML trees both supported the monophyly of Ephemerellidae and Vietnamellidae. Moreover, Ephemerellidae acted as the sister clade to Vietnamellidae whereas Teloganellidae existed in the other clade. Considerable work may be needed to increase the number of mitogenomes to resolve the phylogenetic relationships of Ephemeroptera.We successfully determined the complete mitogenomes of 10.7717/peerj.9740/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.9740/supp-2Supplemental Information 2Click here for additional data file.10.7717/peerj.9740/supp-3Supplemental Information 3Click here for additional data file.10.7717/peerj.9740/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.9740/supp-5Supplemental Information 5Click here for additional data file.10.7717/peerj.9740/supp-6Supplemental Information 6Click here for additional data file.10.7717/peerj.9740/supp-7Supplemental Information 7 Ephemerella sp. Yunnan-2018, Serratella zapekinae, Serratella sp. Yunnan-2018. and Serratella sp. Liaoning-2019; (B) Torleya grandipennis and Torleya tumiforceps. The figure is written permission from Yu-Rou Cao to publish. She helped drawing under our CC BY 4.0 license.External genes on the circle are encoded by the positive strand (5\u2019 \u21923\u2019) and internal genes are encoded by the negative strand (3\u2019 \u21925\u2019). (A)Click here for additional data file.10.7717/peerj.9740/supp-8Supplemental Information 8trnI; (B) trnQ; (C) trnM; (D) trnW; (E) trnC; (F) trnY; (G) trnL (CUN); (H) trnK; (I) trnD; (J) trnG; (K) trnA; (L) trnR; (M) trnN; (N) trnS (AGN); (O) trnE; (P) trnF; (Q) trnH; (R) trnT; (S) trnP; (T) trnS (UCN); (U) trnL (UUR); (V) trnV.A) Click here for additional data file.10.7717/peerj.9740/supp-9Supplemental Information 9trnI; (B) trnQ; (C) trnM; (D) trnW; (E) trnC; (F) trnY; (G) trnL (CUN); (H) trnK; (I) trnD; (J) trnG; (K) trnA; (L) trnR; (M) trnN; (N) trnS (AGN); (O) trnE; (P) trnF; (Q) trnH; (R) trnT; (S) trnP; (T) trnS (UCN); (U) trnL (UUR); (V) trnV.(A) Click here for additional data file.10.7717/peerj.9740/supp-10Supplemental Information 10trnI; (B) trnQ; (C) trnM; (D) trnW; (E) trnC; (F) trnY; (G) trnL (CUN); (H) trnK; (I) trnD; (J) trnG; (K) trnA; (L) trnR; (M) trnN; (N) trnS (AGN); (O) trnE; (P) trnF; (Q) trnH; (R) trnT; (S) trnP; (T) trnS (UCN); (U) trnL (UUR); (V) trnV.(A) Click here for additional data file.10.7717/peerj.9740/supp-11Supplemental Information 11trnI; (B) trnQ; (C) trnM; (D) trnW; (E) trnC; (F) trnY; (G) trnL (CUN); (H) trnK; (I) trnD; (J) trnG; (K) trnA; (L) trnR; (M) trnN; (N) trnS (AGN); (O) trnE; (P) trnF; (Q) trnH; (R) trnT; (S) trnP; (T) trnS (UCN); (U) trnL (UUR); (V) trnV.(A) Click here for additional data file.10.7717/peerj.9740/supp-12Supplemental Information 12trnI; (B) trnI; (C) trnI; (D)trnQ; (E) trnM; (F) trnW; (G) trnC; (H) trnY; (I) trnL (CUN); (K) trnK; (K) trnD; (L) trnG; (M) trnA; (N)trnR; (O) trnN; (P) trnS (AGN); (Q) trnE; (R) trnF; (S) trnH; (T) trnT; (U) trnP; (V) trnS (UCN); (W) trnL (UUR); (X) trnV.(A) Click here for additional data file.10.7717/peerj.9740/supp-13Supplemental Information 13trnI; (B) trnI; (C) trnI; (D)trnQ; (E) trnM; (F) trnW; (G) trnC; (H) trnY; (I) trnL (CUN); (K) trnK; (K) trnD; (L) trnG; (M) trnA; (N)trnR; (O) trnN; (P) trnS (AGN); (Q) trnE; (R) trnF; (S) trnH; (T) trnT; (U) trnP; (V) trnS (UCN); (W) trnL (UUR); (X) trnV.(A) Click here for additional data file.10.7717/peerj.9740/supp-14Supplemental Information 14Click here for additional data file.10.7717/peerj.9740/supp-15Supplemental Information 15Serratella zapekinae;(B) the repeat unit (100 bp) in Serratella zapekinae;(C) the repeat unit (109 bp) in Serratella sp. Liaoning-2019; (D) the repeat unit (72 bp) in Serratella sp. Liaoning-2019.(A) the repeat unit (90 bp) in Click here for additional data file.10.7717/peerj.9740/supp-16Supplemental Information 16T. tumiforceps; (B) the repeat unit (42 bp) in T. grandipennis.(A) the repeat unit (87 bp) in Click here for additional data file.10.7717/peerj.9740/supp-17Supplemental Information 17trnS1 (AGN) and trnE. The CS indicates the 19 bp conserved sequence TTTTTAGCGAACTTAGGGG. Gene sizes are not drawn to scale. Genes located on the majority strand are shown along the top of the boxes whereas genes located on the minority strand are shown on the bottom. White boxes represent genes with the same relative position as in the ancestral insect arrangement pattern. Grey boxes represent non-coding regions. The remaining genes and gene orders that were identical to the ancestral insect are left out.The duplication/random loss model can explain the NCR between Click here for additional data file.10.7717/peerj.9740/supp-18Supplemental Information 18Click here for additional data file.10.7717/peerj.9740/supp-19Supplemental Information 19Click here for additional data file.10.7717/peerj.9740/supp-20Supplemental Information 20Ephemerella sp. Yunnan-2018. Blue shading represents Serratella zapekinae. Yellow shading represents Serratella sp. Yunnan-2018. Green shading represents Serratella sp. Liaoning-2019.Orange shading represents Click here for additional data file.10.7717/peerj.9740/supp-21Supplemental Information 21Click here for additional data file.10.7717/peerj.9740/supp-22Supplemental Information 22Click here for additional data file.10.7717/peerj.9740/supp-23Supplemental Information 23Click here for additional data file."}
+{"text": "Macadamia integrifolia is a representative of the large basal eudicot family Proteaceae and the main progenitor species of the Australian native nut crop macadamia. Since its commercialisation in Hawaii fewer than 100 years ago, global production has expanded rapidly. However, genomic resources are limited in comparison to other horticultural crops. The first draft assembly of M. integrifolia had good coverage of the functional gene space but its high fragmentation has restricted its use in comparative genomics and association studies. Here we have generated an improved assembly of cultivar HAES 741  using a combination of Illumina paired and PacBio long read sequences. Scaffolds were anchored to 14 pseudo-chromosomes using seven genetic linkage maps. This assembly has improved contiguity and coverage, with >120 Gb of additional sequence. Following annotation, 34,274 protein-coding genes were predicted, representing 90% of the expected gene content. Our results indicate that the macadamia genome is repetitive and heterozygous. The total repeat content was 55% and genome-wide heterozygosity, estimated by read mapping, was 0.98% or an average of one SNP per 102 bp. This is the first chromosome-scale genome assembly for macadamia and the Proteaceae. It is expected to be a valuable resource for breeding, gene discovery, conservation and evolutionary genomics. As a member of the Gondwanan family Proteaceae  (Macadamia (F.Muell.) is a basal eudicot and is phylogenetically divergent from other tree crops  , Macadamtivation , is a mitivation . MacadamMacadamia  were then anchored and oriented to 14 pseudo-chromosomes using methods to maximize colinearity across seven genetic linkage maps . An herbarium specimen was submitted to the Southern Cross Plant Science Herbarium [accession PHARM-13-0813]. On collection, samples were snap frozen in liquid nitrogen or placed on dry ice and stored at -80\u00b0 prior to extraction. Previously described methods for and DNA/RNA extraction for Illumina , new libraries with 200, 350 and 550 bp insert sizes were prepared with TruSeq DNA PCR-free kits and sequenced with Illumina HiSeq 2500 producing 57.0, 29.8 and 29.6 Gb paired-end sequence data. A PacBio library (20-Kb) was prepared and sequenced across 10 SMRT cells on a PacBio RSII system (P6-C4 chemistry) generating 6.44 Gb data. Transcriptome sequencing of leaf, shoot and flower tissue followed previously described methods .de novo assembly, scaffolding and gap closing was performed using MaSuRCA , Eucalpytus grandis and Coffea canephora (coffee) genomes. Pairwise sequence similarities were determined applying a BLASTP E-value cut-off of 1E-05 and an inflation value of 1.5 for OrthoMCL Markov clustering. Macadamia specific gene clusters were tested for GO enrichment using OrthoVenn2.Orthologous gene clusters were identified and compared using OrthoVenn2   with 180 \u00d7 coverage of the genome and a scaffold N50 of 413 kb  and 93.5% of corrected PacBio reads tagged as primary alignments by Minimap2 aligned reliably to the assembly. No Pacbio reads aligned to two different scaffolds at a cutoff of 1/2 read length. At 1/3 read length, 130 reads (mean length 7833 bp representing < 0.14% of the assembly) aligned to two scaffolds and no reads aligned to more than two scaffolds.M. integrifolia of 652 Mb (600-700 Mb) that was based on 51.6 Gb Illumina short read data  of the assembly. As reported for many other plant genomes, long terminal repeat (LTR) retrotransposons were the most abundant repeat type (30.5%). Small RNAs including pseudogenes and active small RNA genes including tRNA and snRNA accounted for 0.54% of the assembly .N. nucifera, A. thaliana, P. persica, E. grandis and C. canephora identified 8,961 orthologous clusters of which 2,051 contained a single gene from each of the six eudicot species. M. integrifolia and N. nucifera both belong to the basal eudicot order Proteales. These species and contained similar numbers of clusters with 13,491 and 13,321 respectively and shared the largest number of clusters (503) of any species pair (P = 4.8E-05) and fruit ripening .Annotation of the final assembly, identified 34,274 high-confidence gene models. Of these, 25,779 (75.2%) were anchored to pseudo-chromosomes and 8,495 were located on unanchored scaffolds . Comparaies pair . Tests fHere we present the first chromosome-scale genome assembly for the nut crop macadamia and the large Gondwanan plant family Proteaceae. This provides a platform for unraveling the genetics of macadamia and is expected to underpin future breeding, and comparative and horticultural genomics research.Versions and parameters of the tools implemented in data analysis are provided below:BBMap version 36.62: ktrim = r k = 23 mink = 11 hdist = 1 tpe tbo maxns = 1 minlen = 50 maq = 8 qtrim = rl trimq = 20LoRDEC version 0.4: lordec-correct -2 input_for_read_correction.fastq -k 19 -S out.stat.txt -s 3 -T 12 -i PacBio_filtered_reads.fasta -o out.pacbio.corrected.fastaNUM_THREADS = 16, JF_SIZE = 20000000000 (jellyfish hash size)MaSuRCA version 3.2.1. Default parameters except: vs.macaAssembly.psl -noHead -out = pslL_RNA_scaffolder: blat -t = dna -q = dna scaffolds_gapClosed_min1000.fa Trinity.fasta transcript.ALLMAPS version 0.7.7: Default parametersJellyfish version 2.0: jellyfish count -t 14 -C -m 27 -s 8G -o 27mer_maca_illumOnly_out <all Illumina-only WGS fastqs > jellyfish histo -t14 27mer_maca_illumOnly_out> 27mer_maca_illumOnly_out.histofindGSE: Default parametersGenomeScope version 2.2.6:kmer length 27, read length 125bp, Max kmer coverage 1000Trinity, version 2.0.3: Trinity\u2013seqType fq\u2013max_memory 100G \u2013left reads_S_R1_clean.fq,reads_F2_R1_clean.fq,reads_YL_R1_clean.fq\u2013right reads_S_R2_clean.fq,reads_F2_R2_clean.fq,reads_YL_R2_clean.fq\u2013CPU 8RepeatModeler version 2.0.1. Default parametersRepeatMasker version 4.0.9. Default parametersMAKER, version 2.31.10. Default parameters except: Gene prediction methods Augustus and SNAP (trained with previously generated macadamia gene models); AED score = 0.40; Minimum protein length: = 50 amino acidsBUSCO version 3.0.2: busco -i proteins.fasta -l viridiplantae_odb10 -m proteins -o output_nameMinimap2 version 2.17. Default parameters (paftool.js bundled with Minimap2): minimap2 -ax sr ref_assemblyfasta fastq1stReadPair fastq2ndReadPair > ref_readPairs_aln.paf; k8 paftools.js stat ref_readPairs_aln.paf > alignment_mapstatscf.gz -ERC GVCF -heterozygosity 0.01GATK HaplotypeCaller version 4.1.4.1: gatk\u2013java-options \u201c-Xmx8g\u201d HaplotypeCaller -R refGenome.fasta -I input.bam -O output.g.vcf.gz -O output.vcf.gz\u2013heterozygosity 0.01GATK GenotypeGVCFs version 4.1.4.1: gatk\u2013java-options \u201c-Xmx4g\u201d GenotypeGVCFs -R refGenome.fasta -V input.g.vcf.gz -F CHROM -F POS -F TYPE -F REF -F ALT -F HET -GF GT -GF AD -O output_genoGVCF.table.txt.GATK VariantsToTable version 4.1.4.1: gatk-4.1.4.1/gatk\u2013java-options \u201c-Xmx8g\u201d VariantsToTable -V output.v"}
+{"text": "RAS wild-type metastatic colorectal cancer (mCRC) treated with anti-EGFR  based first-line treatment is still debated. Methods: This multicenter, real-world, retrospective study is aimed at evaluating the effectiveness of second-line Bevacizumab- and Aflibercept-based treatments after an anti-EGFR based first-line regimen. Clinical outcomes measured were: objective response rate (ORR), progression free survival (PFS), overall survival (OS) and adverse events (AEs) profiles. Results: From February 2011 to October 2019, 277 consecutive mCRC patients received Bevacizumab-based  or Aflibercept-based  regimen. No significant difference was found regarding ORR. The median follow-up was 27.7 months (95%CI: 24.7\u201334.4). Aflibercept-treated group had a significantly shorter PFS compared to Bevacizumab-treated group  (HR = 1.34 (95%CI: 0.95\u20131.89); p = 0.0932). The median OS of the Bevacizumab-treated group and Aflibercept-treated group was 16.2 (95%CI: 15.3\u201318.1) and 12.7 (95%CI: 8.8\u201317.5) months, respectively (HR= 1.31 (95%CI: 0.89\u20131.93) p = 0.16). After adjusting for the key covariates  Bevacizumab-based regimens revealed to be significantly related with a prolonged PFS (HR = 1.44 (95%CI: 1.02\u20132.03); p = 0.0399) compared to Aflibercept-based regimens, but not with a prolonged OS (HR = 1.47 (95%CI: 0.99\u20132.17); p = 0.0503). The incidence of G3/G4 VEGF inhibitors class-specific AEs was 7.5% and 26.5% in the Bevacizumab-treated group and the Aflibercept-treated group, respectively (p = 0.0001). Conclusion: Our analysis seems to reveal that Bevacizumab-based regimens have a slightly better PFS and class-specific AEs profile compared to Aflibercept-based regimen as second-line treatment of RAS wild-type mCRC patients previously treated with anti-EGFR based treatments. These results have to be taken with caution and no conclusive considerations are allowed.Background: The optimal anti-angiogenic strategy as second-line treatment in  RAS genotype  1\u20132 and 3\u20134). Toxicities were summarized and compared among subgroups according to three key subgroups: VEGF inhibitors class-specific AEs , hematologic AEs , and non-hematologic AEs . Only AEs which occurred in more than 5% of patients were included in the safety analysis.KRAS, NRAS and BRAF mutational status was assessed with Sanger sequencing, real-time PCR techniques and next-generation sequencing (NGS) ; Pyromark Q96 ID System, Qiagen ; EasyPGX and Myriapod Colon Status, Diatech Pharmacogenetics ). MSI (microsatellite instability) status and/or MMR (mismatch repair) proteins expression were assessed with molecular sequencing  and Immunohistochemistry (IHC) ; Ultraview Universal Detection Kit and Ventana platform, Roche Tissue Diagnostics and Ventana Medical Systems ).All the molecular analyses were performed according to the local clinical practice of the participating centers. p < 0.05. Hazard Ratios (HRs) with 95% confidence intervals (CIs) were calculated using the logistic regression model. All statistical analyses were performed using MedCalc Statistical Software version 18.11.3 .Baseline patients\u2019 characteristics were reported with descriptive statistics and compared among subgroups with the Chi-square test. Chi-square test was also used to compare ORR and the incidence of AEs across subgroups. Logistic regression was used for the multivariate analysis of ORR. Median PFS and median OS were evaluated using the Kaplan\u2013Meier method. Median period of follow-up was calculated according to the reverse Kaplan-Meier method. Cox proportional hazards regression was used for the univariate and multivariate analysis of PFS and OS. The alpha level for all analyses was set to RAS wild-type mCRC patients were treated with Bevacizumab-based  or Aflibercept-based  second-line regimens. The median age was 64.5 years (range: 29\u201384). Patients features  are summarized in p < 0.0001). According to the clinical indication of Aflibercept, also the previously received first-line regimens (p < 0.0001) and second-line chemotherapy backbone (p = 0.0148) were significantly different. A total of 277 consecutive The activity profile for the overall population and according to subgroups is summarized in p = 0.0932) (p = 0.1600) (p = 0.0399) compared to Aflibercept-based regimens, but not with a prolonged OS (HR = 1.47 (95%CI: 0.99\u20132.17); p = 0.0503).The second-line median follow-up for the study population was 27.7 months (95%CI: 24.7\u201334.4); median PFS and median OS were 7.1 months  and 15.7 months . Median PFS of the Bevacizumab-treated group was 7.1 months , while median PFS of the Aflibercept-treated group was 5.6 months , without statistically significant difference at the univariate analysis (HR = 1.34 (95%CI: 0.95\u20131.89);  0.0932) A. Median 0.1600) B. Table p = 0.1908). The incidence of G3/G4 VEGF inhibitors class-specific AEs was 7.5% and 26.5% in the Bevacizumab-treated group and in the Aflibercept-treated group, respectively (p = 0.0001) (p = 0.0033), while the incidence of G3/G4 non hematologic AEs was 4.4% and 10.2%, respectively (p = 0.1032). The incidence of G1/G2 hematologic AEs was 24.6% and 22.4% in the Bevacizumab-treated group and in the Aflibercept-treated group, respectively (p = 0.7545), while the incidence of G3/G4 hematologic AEs was 3.1% and 18.4%, respectively (p < 0.0001).The toxicity profile for the overall study population and according to subgroups is summarized in  0.0001) . The incp = 0.2236). A total of 136 patients (69.4%) and 24 patients (60%) were treated with a third-line systemic therapy, among those who discontinued second-line treatment in the Bevacizumab-treated group and Aflibercept-treated group, respectively (p = 0.5930). A total of 67 patients (29.4%) and nine patients (18.4%) underwent a maintenance therapy after an induction phase, in the Bevacizumab-treated group and in the Aflibercept-treated group, respectively , and this might have affected the clinical outcomes [BRAF mutant patients [BRAF mutational status , identifying a study population with good prognosis overall [Despite the unbalanced grouping of the study population according to the received regimens (82.3% Bevacizumab-based vs. 17.7% Aflibercept-based), most of the patients characteristics were balanced between the subgroups, such as elderly patients, number of metastatic sites and primary tumor location see . On the outcomes . The clipatients , are ali overall .RAS wild-type patients were eligible) might also partially explain these discrepancies. Interestingly, genotype based post-hoc analyses reported an OS of 15.4 months for KRAS wild-type patients of the experimental arm of the ML18147 [RAS wild-type patients of the experimental arm of the VELOUR trial [Even though studies results comparisons are not methodologically correct, some speculations are allowed. The median PFS of the Bevacizumab-treated group (7.1 months) was comparable to the PFS reported in the E3200 and ML18147 trials  ,13, wher ML18147 , and an UR trial . Moreovep = 0.0399), whereas a not significant trend towards a shorter OS was reported (HR = 1.47 (95%CI: 0.99\u20132.17); p = 0.0503). Concerning safety data, we found a significant higher incidence of G3/G4 VEGF inhibitors class-specific AEs among Aflibercept-treated patients, compared to the Bevacizumab-treated patients . This aspect might be also related to the different pharmacodynamic mechanisms of action of Bevacizumab  and Aflibercept ) [p = 0.0033) and G3/G4 hematologic AEs (3.1% vs. 18.4%) to the detriment of the Aflibercept-treated patients, was found. The latter aspect could be related to the different chemotherapy backbone .Intriguingly, the multivariate analysis revealed that the Aflibercept-treated group had a statistically significant shorter PFS compared to the Bevacizumab-treated group (HR = 1.44 (95%CI: 1.02\u20132.03);  (PIGF)) . FurtherOur results suggest a slightly better clinical performance for second-line Bevacizumab-based regimens compared to Aflibercept-based regimens. In our opinion, the different safety profile might had affected the effectiveness of Aflibercept-based regimens compared to Bevacizumab-based regimens, leading to a higher discontinuation rate  and a worse PFS.According to the RAISE trial results , it woulRAS wild-type patients, are awaited. The STRATEGIC-S1 trial (NCT01910610) [RAS wild-type mCRC patients: an oxaliplatin-based second-line regimen with Bevacizumab after fist line FOLFIRI-Cetuximab vs. an irinotecan-based second-line regimen with Bevacizumab after a first-line OPTIMOX Bevacizumab, followed by an anti-EGFR based third-line treatment. The DISTINCTIVE trial (NCT04252456) [RAS wild-type mCRC patients after an oxaliplatin/fluoropyrimidines-based first-line regimen combined with either Panitumumab or Cetuximab. Results from important prospective phase II-III studies, comparing different sequencing strategies of available biological agents for 1910610)  is an in4252456)  is a proRAS wild-type mCRC patients.There are some obvious limitations in this study, including its retrospective design, which expose to selection bias, therefore the results must be taken with caution. Further analysis with a larger sample size and a prospective translational design are certainly needed to better define and personalize the anti-angiogenic strategy as a second-line treatment in RAS wild-type mCRC patients who received first-line anti-EGFR based treatments. These results have to be taken with caution and no conclusive consideration are allowed.Our analysis seems to reveal that Bevacizumab-based regimens have a slightly better efficacy and safety profile compared to Aflibercept-based regimens as second-line treatment of"}
+{"text": "Kernel weight and morphology are important traits affecting cereal yields and quality. Dissecting the genetic basis of thousand kernel weight (TKW) and its related traits is an effective method to improve wheat yield.QTkw.cas-1A.2 and QTkw.cas-4A explained the largest portion of the phenotypic variance for TKW and kernel length (KL), and the other two stable QTLs QTkw.cas-6A.1 and QTkw.cas-7D.2 contributed more effects on kernel width (KW). Conditional QTL analysis revealed that the stable QTLs for TKW were mainly affected by KW. The QTLs QTkw.cas-7D.2 and QKw.cas-7D.1 associated with TKW and KW were delimited to the physical interval of approximately 3.82\u2009Mb harboring 47 candidate genes. Among them, the candidate gene TaFT-D1 had a 1\u2009bp insertions/deletion (InDel) within the third exon, which might be the reason for diversity in TKW and KW between the two parents. A Kompetitive Allele-Specific PCR (KASP) marker of TaFT-D1 allele was developed and verified by PG-RIL and a natural population consisted of 141 cultivar/lines. It was found that the favorable TaFT-D1 (G)-allele has been positively selected during Chinese wheat breeding. Thus, these results can be used for further positional cloning and marker-assisted selection in wheat breeding programs.In this study, we performed quantitative trait loci (QTL) analysis using recombinant inbred lines derived from the cross \u2018PuBing3228\u2009\u00d7\u2009Gao8901\u2019 (PG-RIL) to dissect the genetic basis of kernel traits. A total of 17 stable QTLs related to kernel traits were identified, notably, two stable QTLs TaFT-D1 could be the candidate gene for QTLs QTkw.cas-7D.2 and QKw.cas-7D.1.Seventeen stable QTLs related to kernel traits were identified. The stable QTLs for thousand kernel weight were mainly affected by kernel width.  Triticum aestivum L.) is one of the most important cereal crops for feeds 40% of population in the world (http://www.fao.org/). Wheat yield is determined by thousand kernel weight (TKW), kernel number per spike, and effective tiller number [Common wheat  in tetraploid wheat [Recently, high-density single nucleotide polymorphism (SNP) arrays technology provides a superior approach to identify QTLs for wheat kernel-related traits \u201331. To did wheat , 42. HowUsing a RIL population derived from \u2018PuBing 3228 (P3228)\u2009\u00d7\u2009Gao8901 (G8901)\u2019, the objectives of this study were to (i) identify stable and major QTLs for TKW, KL, KW and KL/W under different field conditions; (ii) reveal the contribution of the other kernel traits to TKW using conditional QTL analysis; (iii) predict candidate gene(s) for targeted QTLs interval based on reference genome annotation information; (iv) develop KASP markers of the candidate gene(s) and verified by PG-RIL and a natural population consisted of 141 cultivar/lines for marker-assisted selection in high-TKW wheat breeding.H) of four traits were higher than 0.60  are listed in Table\u00a0A total of 47 putative QTLs were detected for TKW, KL, KW and KW/L Figs.\u00a0.Fig. 3GQTkw.cas-1A.2, QTkw.cas-4A, QTkw.cas-5D, QTkw.cs-6A.1 and QTkw.cas-7D.2  ranged from 8.31 to 11.84%  were significant in at least two environments . The favorable alleles of the five QTLs  were derived from the parent G8901 , 1B, 4B, 6A, 7A (two) and 7D, respectively  were found in at least two environments .A total of 10 QTLs for KL/W were identified on chromosomes 1A, 1B, 2A, 5A (two), 5D, 7A (two) and 7D (two), with PVE of individual QTL ranging from 1.79 to 22.41% \u2009=\u200921.93%), indicating that the major QTLs QTkw.cas-4A and QKl.cas-4A for TKW and KL, respectively, were significantly affected by the environment  interactions were detected at 43 loci for TKW, KL, KW and KW/L  were identified to be associated with KW, but independent of KL and KL/W , a homolog of Arabidopsis FLOWERING LOCUS T, was considered as the candidate gene for QTkw.cas-7D.2 and QKw.cas-7D.1  closely linked to the two stable QTLs (QTkw.cas-7D.2 and QKw.cas-7D.1) and 1\u2009bp InDel of TaFT-D1 were further converted to KASP markers -allele) and P3228-allele (TaFT-D1(\u2212)-allele) plants -allele was significantly higher than those of the TaFT-D1(\u2212)-allele .Two SNPs markers -allele in modern cultivars was higher in the seven agro-ecological wheat production regions , suggesting that TaFT-D1(G)-allele have undergone positive selection during wheat breeding process -allele can be used in different wheat production regions.To determine whether the two g season , 52. ComQTkw.cas-1A.2, QTkw.cas-5D, QTkw.cas-6A.1, QTkw.cas-7D.2) account for TKW, while two (QTkw.cas-4A and QTkw.cas-5D) on KL -allele associated with TKW and KW has been positively selected during Chinese wheat breeding. In addition, the current study provided new options for dissecting the genetic basis of yield and molecular-assisted breeding.In this study, we performed QTL analysis using the PG-RIL population in four environments for kernel-related traits , which were mainly distributed on chromosomes 1A, 1B, 4A, 5D, 6A, 7A and 7D Fig. . Notably6\u20139 RILs derived from \u2018PuBing3228\u2009\u00d7\u2009Gao 8901 \u2018was developed by single seed descent method. The wheat germplasm P3228 was developed by Dr. Lihui Li . G8901 is a commercial cultivar released by Gaocheng institute of agricultural science, Hebei, China. The P3228 has higher kernel number per spike and the G8901 has higher thousand kernel weight . The H was calculated using the QGAStation 2.0 (http://ibi.zju.edu.cn/software/qga/v2.0/index c.htm) and the following formula H\u2009=\u2009VG/VP; where VG and VP are the genetic variance and phenotypic variance, respectively.For the four environments, 10 representative plants were sampled from each plot to investigate kernel-related traits. At seed maturity, the TKW and kernel morphometric traits  of at least 500 kernels were measured three times using the rapid SC-G grain appearance quality image analysis system . Analysis of variance (ANOVA), mean values of traits, standard deviations and variation coefficients (CV) were performed with SPSS Statistics v20.0 software . Effects among genotypes, environments, and GE interaction were estimated by ANOVA. BLUP for all four traits across four environments was calculated using R software ) of TKW in wheat were obtained by the mixed-model approach. The conditional phenotypic value can be partitioned as.Conditional QTL analysis was performed to analyze the genetic contributions of kernel-related traits to TKW, by the procedure of inclusive composite interval mapping . The cony(TKW|KL)=\u03bc(TKW|KL)\u2009+\u2009G(TKW|KL)\u2009+\u2009E(TKW|KL)\u2009+\u2009e(TKW|KL).y(TKW|KL) is the conditional phenotypic value of TKW on KL; \u03bc(TKW|KL) is the conditional population mean, G(TKW|KL) is the conditional general genotypic effect; E(TKW|KL) is the conditional effect for the environment and e(TKW|KL) is the conditional residual error.where (TKW|KL) denote TKW conditional on KL; y(TKW|KL), y(TKW|KW) and y(TKW|KL/W)) are the conditional phenotypic value of TKW on KL, KW or KL/KW in the corresponding environment, which were estimated using QGAStation2.0 (http://ibi.zju.edu.cn/software/qga/). MapChart\u00a02.2 (http://www.biometris.nl/uk/Software/MapChart/) was used to draw the genetic map.The conditional phenotypic values  and physical (right) positions for SNPs mapped on the chromosome 7DS in PG-RIL genetic map. Fig. S2 A 1\u2009bp InDel in TaFT-D1 caused a frameshift mutation of the protein. (a) Sequence alignment of TaFT-D1 showing 1\u2009bp InDel between P3228 and G8901. (b) Protein alignment of TaFT-D1 showing frameshift mutation in P3228. Fig. S3 Heatmap showing the expression profile of DEGs at 15 development stages. Fig. S4 Allelic segregation of KASP markers AX-111061288 (a) and AX-111184541 (b) for QTkw.cas-7D.2 and QKw.cas-7D.1."}
+{"text": "Chronic isoleucine supplementation prevents diet-induced weight gain in rodents. Acute-isoleucine administration improves glucose tolerance in rodents and reduces postprandial glucose levels in humans. However, the effect of chronic-isoleucine supplementation on body weight and glucose tolerance in obesity is unknown. This study aimed to investigate the impact of chronic isoleucine on body weight gain and glucose tolerance in lean and high-fat-diet (HFD) induced-obese mice. Male C57BL/6-mice, fed a standard-laboratory-diet (SLD) or HFD for 12 weeks, were randomly allocated to: (1) Control: Drinking water; (2) Acute: Drinking water with a gavage of isoleucine (300 mg/kg) prior to the oral-glucose-tolerance-test (OGTT) or gastric-emptying-breath-test (GEBT); (3) Chronic: Drinking water with 1.5% isoleucine, for a further six weeks. At 16 weeks, an OGTT and GEBT was performed and at 17 weeks metabolic monitoring. In SLD- and HFD-mice, there was no difference in body weight, fat mass, and plasma lipid profiles between isoleucine treatment groups. Acute-isoleucine did not improve glucose tolerance in SLD- or HFD-mice. Chronic-isoleucine impaired glucose tolerance in SLD-mice. There was no difference in gastric emptying between any groups. Chronic-isoleucine did not alter energy intake, energy expenditure, or respiratory quotient in SLD- or HFD-mice. In conclusion, chronic isoleucine supplementation may not be an effective treatment for obesity or glucose intolerance. The branched-chain amino acids (BCAAs), isoleucine, leucine and valine, are essential amino acids accounting for ~35% of the essential amino acids comprising muscle proteins in humans and ~40% of the pre-formed amino acids required by all mammals . In popudb/db) mice, a model of morbid obesity and hyperglycaemia [Acute administration of isoleucine and leucine in rats improved glucose tolerance, with isoleucine showing greater effectiveness than leucine . This walycaemia . This evIt is known that postprandial blood glucose levels are influenced by the rate of gastric emptying . In partThe current study aimed to determine whether chronic dietary supplementation with the BCAA isoleucine, alters body weight gain, adiposity, glucose tolerance, and energy metabolism in mice with HFD-induced obesity.This study was approved  by the South Australian Health and Medical Research Institute Animal Ethics Committee. All experimental protocols were performed in alignment with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes.n = 54) were group-housed in a 12:12 h light-dark cycle within a temperature (24 \u00b1 1 \u00b0C) controlled facility. Mice were provided ad libitum access to either a standard laboratory diet  or HFD . Consistent with the previous literature, the HFD-induced obese mouse model was chosen as both a model of obesity [n = 10; SLD-Chronic (Ch)) and HFD-mice  received ad libitum isoleucine  supplemented in the drinking water. The remaining SLD (n = 20) and HFD-mice (n = 16) continued with ad libitum access to normal drinking water. At 16 weeks, all mice were singly housed and underwent an OGTT (at 1400 h) and gastric emptying breath test (at 0900 h) in random order with a three-day recovery between tests. In each diet group, the mice provided normal drinking water were subdivided into two groups, receiving either an oral gavage of water ; n = 8, HFD-C) or isoleucine ; n = 8, HFD-A) 30 min before the OGTT or GEBT. The SLD and HFD-Ch mice received an oral gavage of drinking water similar to the control groups. The doses for acute and chronic isoleucine treatments were chosen based on previous studies [Eight-week-old male C57BL/6 mice  to consume voluntarily within 1 min. Breath samples were collected before isoleucine administration  and again at regular intervals  after egg consumption. Breath samples were analysed for the 13CO2 content using an isotope ratio mass spectrometer . The 13CO2 excretion data were analysed by non-linear regression analysis for curve fitting and for calculation of gastric half emptying time (t \u00bd) [Gastric emptying of a solid meal was determined using a non-invasive breath test as previously described ,22. Miceme (t \u00bd) . Gastric2/VO2), and total activity  were measured and analysed using the ExpeData data analysis software .Mice were individually housed in Promethium metabolic cages  for 72 h of continuous metabolic monitoring. Energy intake (kJ), energy expenditure (kJ/lean mass), respiratory quotient  then anaesthetised with isoflurane . The nose-to-tail length and abdominal circumference of mice were measured. Blood was collected from the abdominal aorta and transferred to ethylenediaminetetraacetic acid (EDTA) tubes . Plasma was extracted by centrifugation at 1000 g and 4 \u00b0C for 15 min, and snap-frozen in liquid nitrogen prior to storage at \u221280 \u00b0C until further analysis. Liver, gonadal fat pads, and inter-scapula brown fat pads were collected and weighed. Lean mass was determined by the final body weight minus the weight of collected fat pads. A section of the liver was fixed in 4% paraformaldehyde for 4 h, cryoprotected overnight in 30% sucrose in a phosphate buffer, frozen in Tissue-Tek O.C.T. compound  and stored at \u221280 \u00b0C before processing for histology. Plasma total triglycerides, total cholesterol, and high-density lipoprotein (HDL)-cholesterol concentrations were measured using commercial enzymatic kits ) on a Beckman AU480 clinical analyser . Plasma low-density lipoprotein (LDL)-cholesterol concentrations were estimated using the Friedewald equation : LDL-cho2 area using the ImageJ-win64 software.The histological lysochrome lipid stain Oil Red O was performed on liver sections using a standard protocol . Slides 0\u2013120 min blood glucose area under the curve (AUC) was generated using the IBM SPSS Statistics 26 software . A correlation between gastric half emptying time (t \u00bd) and OGTT0\u2013120 min blood glucose AUC for SLD groups was performed in the GraphPad Prism v8 software. The coefficient of determination value (r2) was considered significant at p < 0.05.Results are expressed as the mean \u00b1 SEM. A two-way ANOVA was performed to assess diet and isoleucine treatment effects with a Tukey\u2019s post hoc test for multiple comparisons, using the GraphPad Prism v8 software . The OGTTp < 0.001, unpaired t-test; p < 0.0001, F  = 28.17, diet effect; p = 0.1262, F  = 2.418, isoleucine effect; SLD-C/A; 34.8 \u00b1 0.7 g (n = 20), SLD-Ch; 37.2 \u00b1 1.3 g (n = 10), HFD-C/A; 42.3 \u00b1 1.6 g (n = 16), and HFD-Ch; 44.1 \u00b1 1.7 g (n = 8)).At the beginning of week 12, prior to isoleucine supplementation, HFD-mice gained significantly more weight than SLD-mice  = 19.21, diet effect; In weeks 12\u201315, HFD-mice continued to gain more weight compared to SLD-mice  = 24.78, diet effect; SLD-C, 9.3 \u00b1 0.2 cm, SLD-Ch, 9.9 \u00b1 0.5 cm, HFD-C, 11.4 \u00b1 0.3 cm, and HFD-Ch, 11.2 \u00b1 0.3 cm). In addition, HFD-mice had heavier gonadal fat pads and brown fat pads than SLD-mice  = 11.13 and F  = 11.44, respectively, diet effect; At week 18, abdominal circumference was greater in HFD-mice than SLD-mice, but was not affected by chronic isoleucine treatment  = 18.34, diet effect; p < 0.0001, F  = 37.31, diet effect; HFD-mice had heavier livers than SLD-mice  = 4.157, diet effect; p < 0.05, F  = 5.715, diet effect; HFD-mice consumed more energy across 24 h  = 9.151, p < 0.05, F  = 6.499, p < 0.01, F  = 11.25, respectively, diet effect; p < 0.05, F  = 5.416, interaction; p < 0.05, F  = 6.468, interaction; p < 0.05, Sidak\u2019s post hoc test), but not in HFD-mice.HFD-mice had a significantly lower energy expenditure  compared to SLD-mice across 24 h, during the light phase or dark phase  = 19.66, diet effect; p < 0.05, F  = 4.082, diet effect; p < 0.001, F  = 38.4, diet effect; HFD-mice had lower RQ values compared to SLD-mice across 24 h  = 31.28, diet effect; p < 0.001, F  = 40.02, diet effect; p < 0.001, F  = 40.16, diet effect; p < 0.0001, F  = 25.36, diet effect; HFD-mice had elevated plasma total triglycerides  = 19.34, diet effect; HFD-mice had higher fasting blood glucose levels than SLD-mice  = 34.44, diet effect; p < 0.05, one-way ANOVA; HFD-mice had a greater glucose AUC than SLD-mice (n = 8); SLD-A, 144.4 \u00b1 15.3 min (n = 8) and SLD-Ch, 126.7 \u00b1 5.1 min (n = 7)).There was no significant difference in the gastric half emptying time (t \u00bd) between different isoleucine groups in SLD-mice ; SLD-C, r2 = 0.0197, SLD-A, r2 = 0.2602, and SLD-Ch, r2 = 0.004292).There was no correlation between gastric half emptying time and blood glucose AUC in SLD-mice  mice, significantly reduced blood glucose levels in response to an OGTT, albeit at a higher dose of 0.5 g/kg [In the current study, acute isoleucine orally administered at a dose of 0.3 g/kg, had no effect on postprandial blood glucose levels in SLD- or HFD-mice. Previously, acute administration of 0.3 g/kg isoleucine in lean  and diet0.5 g/kg . ConsideConsistent with a previous report , the chr13CO2 content in the breath following a gastric emptying breath test, but did not significantly affect the 13CO2 AUC, Cmax, or Tmax values (peak concentration and the time at which it occurred) compared to controls [Slowing gastric emptying allows for efficient digestion and absorption of nutrients, including glucose ,36,37. Icontrols .The acute and chronic isoleucine supplementation had no beneficial effect to limit HFD-induced body weight gain, adiposity, fasting blood glucose levels, and glucose tolerance. In contrast, the chronic isoleucine treatment impaired glucose tolerance in SLD-mice. Therefore, the chronic isoleucine supplementation is unlikely to be an effective dietary intervention for the treatment of obesity and type 2 diabetes."}
+{"text": "Campylobacter (Campylobacter jejuni and Campylobacter coli) isolates (n = 158) recovered from broiler neck skin and caecal contents in Ireland over a one-year period, resistant to at least one of three clinically relevant antimicrobial classes, were screened for resistance determinants. All ciprofloxacin-resistant isolates (n = 99) harboured the C257T nucleotide mutation (conferring the Thr-86-Ile substitution) in conjunction with other synonymous and nonsynonymous mutations, which may have epidemiological value. The A2075G nucleotide mutation and amino acid substitutions in L4 and L22 were detected in all erythromycin-resistant isolates (n = 5). The tetO gene was detected in 100% (n = 119) of tetracycline-resistant isolates and three of which were found to harbour the mosaic tetracycline resistance gene tetO/32/O. Two streptomycin-resistant C. jejuni isolates (isolated from the same flock) harboured ant(6)-Ib, located in a multidrug resistance genomic island, containing aminoglycoside, streptothricin (satA) and tetracycline resistance genes (truncated tetO and mosaic tetO/32/O). The ant(6)-Ie gene was identified in two streptomycin-resistant C. coli isolates. This study highlights the widespread acquisition of antimicrobial resistance determinants among chicken-associated Campylobacter isolates, through horizontal gene transfer or clonal expansion of resistant lineages. The stability of such resistance determinants is compounded by the fluidity of mobile genetic element.Campylobacteriosis is the leading cause of human bacterial gastroenteritis, very often associated with poultry consumption. Thermophilic  Campylobacter is the most commonly reported foodborne bacterial pathogen causing human gastroenteritis in the European Union (EU) and Ireland, most often associated with the broiler reservoir [Campylobacter spp. of food animal origin continues to limit the spectrum of clinically useful antimicrobials and is internationally recognised as a major societal challenge. Veterinary antimicrobials used therapeutically and prophylactically are often the same as, or belong to the same class as those used clinically [eservoir . Irelandeservoir . Frequeninically .Campylobacter spp. isolates, while resistance to erythromycin is typically low to moderate across Europe [Macrolides, fluoroquinolones and aminoglycosides are classified as critically important antimicrobials, while tetracycline is considered a highly important antimicrobial . Resistas Europe ,5,6. Macs Europe ,8,9. Syss Europe  and resigyrA and gyrB, respectively), catalysing ATP-dependent negative supercoiling of DNA to regulate replication, repair and gene expression [Campylobacter spp. is largely mediated by chromosomal mutations in the quinolone resistance-determining region (QRDR) of gyrA, typically conferred by the C257T nucleotide mutation (Thr-86-Ile) [In Gram negative bacteria, DNA gyrase is the primary target of fluoroquinolones . DNA gyrpression ,13,14. R-86-Ile) . The QRD-86-Ile) . rplD and rplV, respectively) or the presence of the emerging ermB gene contribute to macrolide resistance [ermB was reported recently for the first time in thermophilic Campylobacter spp., located on a chromosomal multidrug resistance genomic island (MDRGI) (likely originating from a Gram positive species) in a high-level erythromycin-resistant Campylobacter coli (C. coli) isolate (ZTC113) of swine origin in China [Macrolides act by binding to the 50S bacterial ribosomal subunit and inhibit translational elongation, and interfere with protein synthesis and subsequent ribosomal subunit assembly ,17. Polysistance ,18. \u0392etasistance ,20. Mutasistance . The ribin China . ErmB diin China . Campylobacter spp. contributes to baseline resistance against structurally diverse antimicrobials [cmeABC operon encodes a tripartite multidrug efflux pump that consists of an outer membrane channel protein (cmeC), an inner membrane efflux transporter (cmeB) and a periplasmic fusion protein (cmeA) [cmeR) binding to an inverted repeat (IR) (TGTAATAAATATTACA) in the intergenic region between cmeR and cmeA transcriptionally represses the cmeABC operon [The intrinsic, chromosomally encoded resistance-nodulation-division (RND) CmeABC (Campylobacter multidrug efflux) efflux pump in crobials ,24,25,26n (cmeA) . RepressC operon ,29. ConsC operon ,30.Campylobacter spp. is largely conferred by a ribosomal protection protein (RPP), TetO, capable of displacing tetracycline from its primary binding site on the 30S ribosomal subunit [Campylobacter tetO can be located chromosomally but is often plasmid-mediated [tetO gene exists in at least eleven bacterial genera, including four Gram negative and seven Gram positive genera [tetO between members of different bacterial genera indicates that conjugative plasmids, transposons, or recombination events contribute to the dissemination and maintenance of the tetO gene [Tetracycline resistance in  subunit ,31. Bact subunit ,33,34. Cmediated ,37,38,39tetO acquisition is the most prevalent genetic event conferring tetracycline resistance among Campylobacter spp., mosaic tet genes  have also been reported within the genus [tetO, W, 32 [tetM, S [tet genes are widespread among Gram positive and Gram negative genera in human and animal isolates [Although he genus . Mosaic O, W, 32 ,40,41,42[tetM, S ) to form[tetM, S . Mosaic Campylobacter spp. resistance to aminoglycosides is mediated by reduced antimicrobial binding affinity for target sites due to enzymatic modification, via acetylation, phosphorylation or adenylation of amino or hydroxyl groups of the aminocyclitol nucleus or sugar moieties [ant(6)-I-sat4-aphA3), first isolated from Staphylococci [Campylobacter spp. isolates [Aminoglycosides are broad spectrum antimicrobials and inhibit protein synthesis by binding to 16S rRNA of the 30S ribosome ,45. Campmoieties ,45,46. Amoieties ,48,49, wmoieties ,51. The ylococci ,53. ANT and C. coli isolates recovered from Irish broiler neck skin and caecal samples over a one-year period (2017\u20132018).Despite the high prevalence of  broiler ,57,58,59 broiler ,59,60,61 broiler  or rumin broiler ,63 isolagyrA gene and 100% of isolates (n = 99) harboured a C257T point mutation, which is the dominant mutation conferring resistance among campylobacters. Resistant isolates were grouped into C. jejuni gyrA and C. coli gyrA  sequence types based on the presence of synonymous and nonsynonymous mutations present in the portion of the gyrA gene sequenced (C. jejuni isolates (n = 85) were grouped into three GTJs . A large proportion (47.1%) carried the Thr-86-Ile substitution exclusively (GTJ1). Synonymous mutations T72C, C243T, T357C, C360T, C471T, T483C, and C622T were exclusively associated with C. jejuni and were present in both GTJ-II and GTJ-III. Nonsynonymous Ser-22-Gly (A64G) and Ala-206-Thr (G616A) mutations were present in 35.3% and 17.7% of isolates of ciprofloxacin-resistant C. jejuni isolates, respectively and were the basis of defining GTJ-II and GTJ-III, respectively. Both GTJ-II and GTJ-III were associated with the Asn-203-Ser (A608G) substitution. All CIP-resistant C. coli isolates tested (n = 14) harboured one nonsynonymous mutation only (Thr-86-Ile), but were grouped into seven GTCs based on the presence of various synonymous mutations quinolone-resistant isolates tested and minimum inhibitory concentrations (MICs) ranged from 0.5\u20138 mg/L.rplD gene was detected in the five isolates, and the partial sequences shared 100% homology with the rplD gene of erythromycin-sensitive C. coli isolates (GenBank accession numbers: MH084640.1 and MH084639.1) [rplV sequence in all erythromycin-resistant isolates: double point mutation at positions 308 and 309 , A325G (Thr-109-Ala), a double point mutation at positions 332 and 333 (Ala-111-Glu), G340A (Ala-114-Thr) and C358A (Pro-120-Thr). Nonsynonymous mutations  in rplV were also identified in all erythromycin-resistant isolates. Partial rplV sequences in this study were homologous to a high-level erythromycin-resistant clinical C. coli isolate rplV gene (GenBank accession number: GU384982.1) [cmeR-cmeA intergenic region, upstream of the IR (positions 66-80), homologous to sequences derived from erythromycin-resistant (GenBank accession number: FJ797673.1) and erythromycin-sensitive (GenBank accession number: FJ797671.1) strains [ermB gene was not detected among the erythromycin-resistant isolates tested.Five erythromycin-resistant isolates were screened for mutations contributing to erythromycin resistance. All five erythromycin-resistant isolates harboured the A2075G mutation in the 23S rRNA gene. A T82C mutation (Ser-28-Pro substitution) in the  strains . The ermtetO gene was detected in 100% of tetracycline-resistant isolates (n = 119). Three isolates , accounting for 2.5% of the tetracycline-resistant isolates, harboured the mosaic tetO/32/O type II gene, confirmed by PCR/partial sequencing and genomic sequencing (Streptococcus suis (S. suis) (GenBank accession number: KY994102.1) tetO/32/O gene (Campylobacter spp. (GenBank accession numbers: WP_052855148.1) and Clostridiales (WP_117823345.1) TetO/32/O sequences. A portion of the quencing B. Isolat2/O gene . EquallyC. jejuni isolates CITCj625-18 and CITCj727-18 were found to harbour multiple resistance genes, including a truncated tetO , mosaic tetO/32/O type II (1920 bp), aminoglycoside-6-nucleotidyltransferase (ant(6)-Ib)), and streptothricin acetyltransferase (satA). These antimicrobial resistance genes were located circumjacent to proteins involved in replication and recombination (C. jejuni isolates CITCj625-18 (first thin)  and CITCj727-18   were nearly identical with an average nucleotide identity (ANI) of 99.99% and 30.5% GC content, indicating that the isolates had been circulating within and had been maintained by the flock. The pair belonged to sequence type ST-45 clonal complex (ST-137) and harboured LOS locus class C.bination A. The MDant(6)-Ie gene (900 bp), with almost identical sequences (99.89% identity). ANT(6)-Ie in this study shared 99.66% amino acid identity with each other, where CITCc3448-18 harboured a nonsynonymous G820A mutation  and was identical to a C. coli ANT(6)-Ie protein (GenBank: WP_052786298.1). CITCc1631-18 and CITCc3448-18 ANT(6)-Ie shared between 29.7-36.1% identity to ANT(6)-Ia, ANT(6)-Ib, ANT(6)-Ic, and ANT(6)-Id amino acid sequences  [CITCc1631-18 and CITCc3448-18 belonged to ST-828 clonal complex  and harboured the ctively) ,51,65. N-acetyltransferase (AAC(3)) (261 amino acids) was also detected in both CITCj625-18 and CITCj727-18, 44.3% identical to a aminoglycoside 3-N-acetyltransferase (AAC(3)) variant (239 amino acids) in both CITCc1631-18 and CITCc3448-18. However, these isolates were gentamicin-susceptible, although this protein may confer resistance to other aminoglycoside antibiotics [C. coli variant was identical to AAC(3) genes reported in Campylobacter spp., 37.5% identical to AAC(3)-Ia described in Serratia marcescens (GenBank accession number: Q7B9H0), and 19\u201332.25% similar to AAC(3) orthologous, including types AAC(3)-Ib/Ic/Id/Ie/IIa/IIb/IIc/IId/IIe/IIf/IIIa/IIIb/IIIc/IVa/VIa/VIIIa/Ixa/Xa/XIa. The AAC(3) variant in CITCj625-18 and CITCj727-18 was identical to Campylobacter AAC(3) variants and was 19.3\u201325.84% similar to orthologues reported in other bacterial genera.Aminoglycoside 3-ibiotics . The C. Campylobacter spp. isolates  have been detailed previously [The antimicrobial resistance rates of this pool of broiler-associated thermophilic eviously , and areCampylobacter isolates, collected throughout the Republic of Ireland, from the three largest poultry processors, over a one-year period (2017\u20132018). This study reports the antimicrobial resistance determinants circulating among Irish broiler-associated Campylobacter spp. [n = 99) tested in the current study, similar to reports published worldwide [Campylobacter spp. isolates [The Thr-86-Ile mutation is the predominant genetic alteration conferring (fluoro)quinolone resistance among ter spp. , and wasorldwide ,70,71. Sisolates ,62,72, iCampylobacter spp. are ecologically competitive and persistent even in the absence of antimicrobial selective pressure [Campylobacter isolates (3.1% to 28.9%) in Ireland between 1998 and 2000 [Fluoroquinolone-resistant pressure ,74,75. Dand 2000 ,76, in 2and 2000 . Clonal and 2000 ,78,79.C. jejuni isolates harbouring multiple amino acid substitutions in the gyrA QRDR (GTJ-II and GTJ-III)  were necessary to produce high level moxifloxacin resistance [gyrA [gyrA QRDR have a negligible effect on (fluoro)quinolone resistance.GTJ-III)  had ciprsistance ,85. MoxigyrA alleles within a population of Campylobacter isolates have been identified as epidemiological markers and may serve as a supplementary approach to classical epidemiological typing methods [C. coli gyrA type in 0.23% (n = 1) of 430 C. jejuni isolates tested, and 1.4% (n = 4) C. coli isolates harboured a typical C. jejuni gyrA type. The amino acid substitutions present in each of the three GTJ , while MICs ranged from 128 mg/L to \u2265 128 mg/L. Three of these isolates were collected from the same flock in north-central Ireland while one isolate was collected from a farm approximately 10 km away, the following week. The fifth erythromycin-resistant isolate was recovered from a farm in the mid-south-west of Ireland, two months previously, but all birds from these farms were processed in the same processing plant. Identical mutations in the 23S rRNA, rplV gene, encoding the 50S ribosomal protein L22 were detected in the C-terminal region (amino acids 109\u2013142) [cmeR regulatory IR. The effects of mutations detected in this study in rplD, rplV, and the intergenic region of cmeR-cmeA are unknown, but may contribute to erythromycin resistance.The A2075G point mutation in the 23S rRNA gene remains the most prevalent genetic event conferring macrolide resistance ,92 and w109\u2013142) , includi109\u2013142) . Mutatio109\u2013142) ,93) are ermB gene was not detected among the erythromycin-resistant isolates tested. Resistance mediated by ermB in Campylobacter spp. is largely confined to China, which may reflect the extensive use of antimicrobials in food producing animals in China [ermB-positive C. coli isolates recovered from poultry exist in Europe [ermB-positive isolate was detected for the first time in the United States in a C. jejuni isolate of clinical origin [ermB gene was recently detected in 18.3% of 240 thermophilic Campylobacter spp. retail meat associated-isolates in South Africa [C. jejuni 23S rRNA has been associated with a fitness cost and reduced doubling times [The in China . Three rn Europe ,95,96 anl origin , while th Africa . Mutationg times ,100,101,tetO gene, while three isolates harboured the mosaic tetO/32/O type II gene were detected. Mosaic tetracycline resistance genes in Campylobacter spp. are typically derived from tetO and tet32 sequences in the type II conformation, with a shorter central tet32 segment [Campylobacter spp. The first mosaic tetracycline RPP gene was detected in Megasphera elsdenii, composed of a central tetW region flanked by two tetO regions [tetO primers [tetO gene and enable the detection of mosaic tet genes with a central portion flanked by an initial tetO region until position 228. The tetO/32/O type II gene reported here was associated exclusively with an MIC of 64 \u00b5g/L. However, 27 of 119 (22.7%) tetracycline-resistant Campylobacter spp. isolates tested had MICs of 64 \u00b5g/L or \u2265 64 \u00b5g/L, indicating that other factors contribute to enhanced tetracycline resistance. It should be noted that in this study, all three isolates harbouring mosaic tetracycline genes were also co-resistant to streptomycin, enabling co-selection and persistence of these antimicrobials. The burden of mosaic tetracycline resistance genes within the genus should be considered as part of the approach to elucidate developing and newly acquired antimicrobial resistance determinants within the genus. All tetracycline-resistant isolates harboured a portion of the  segment , althoug regions . However regions ,43. The  primers  used in n = 4) isolates had MICs of \u2265 16 mg/L. Streptomycin-resistant C. jejuni isolates CITCj625-18 and CITCj727-18 (ST-137) harboured sialylated LOS locus class C, which has been identified as a risk factor for post-infectious Guillain-Barr\u00e9 syndrome and increased severity of enteric disease [C. jejuni ST-45 clonal complex [All streptomycin resistant  revealed identical genes in a multidrug resistance genomic island  and Campylobacter spp. (GenBank accession number: WP_002823161.1). The presence of multiple tet genes (coding for similar or different mechanisms) in Gram negative isolates has also been documented [tetO genes have also been reported in C. coli MDGRI containing aadE (ant(6)-Ib) and ermB [ant(6)-Ib (867 bp)) and streptothricin resistance (satA) genes were also located within the multidrug resistance island (satA) is frequently reported in Gram positive bacilli [sat4) reported in Campylobacter [Genomic sequencing of tetracycline-/streptomycin-resistant c island A. Both icumented . Howeverand ermB . Aminogle island A. Strept bacilli  and sharlobacter ,111,113.C. coli of swine origin in Spain, America, and Grenada [C. coli ST-6543 on the PubMLST Campylobacter database at the time of writing [CITCc3448-18 belonged to ST-828 clonal complex (ST-1096). ST-1096 has been isolated from a case of gastroenteritis in the United Kingdom (UK) in 2016  and has  Grenada ,115,116. Grenada . There a writing , includiC. coli isolates (CITCc1631-18 and CITCc3448-18) harboured the emerging ant(6)-Ie gene (900 bp), found widely disseminated among clinical and animal C. coli isolates [Campylobacter ant(6) genes, ant(6)-Ie appears to be inherent to C. coli and does not have a Gram positive ancestor [ant(6)-Ie gene was originally detected in a hypervariable genomic region, unaccompanied by other resistance genes [ant(6)-Ie genes were located chromosomally, and were not located near other resistance determinants. Both streptomycin-resistant C. coli isolates were co-resistant to ciprofloxacin/nalidixic acid (GTC-V) and tetracycline. C. coli isolate 1631-18 harboured tetO (1920 bp), while C. coli isolate CITCc3448-18 also harboured the mosaic tetracycline resistance gene, tetO/32/O (1290 bp), highly homologous with that detected in CITCj625-18 and CITCj727-18. Both streptomycin-resistant isolates ,65. UnliCampylobacter isolates (296 C. jejuni and 54 C. coli) were recovered from free range and intensively-reared broiler carcasses  using ISO 10272-2:2017 [C. jejuni and 18 C. coli) isolates tested were resistant to at least one antimicrobial and were subsequently tested for resistance determinants.A total of 350 thermophilic 2-2:2017  for thei2-2:2017 ,119. Ove2, 10% CO2, 85% N2) and subcultured. DNA was extracted using the PureLink Genomic DNA Mini Kit , according to manufacturer\u2019s instructions and DNA was standardised to 50\u2013100 ng/\u03bcL.Briefly, isolates were recovered from \u221280 \u00b0C on Columbia blood agar  and incubated for 24 h at 42 \u00b0C, under microaerobic conditions (5% OtetO/32/O and tetO/W/O) were designed on SnapGene2.3.2 software  and regions of primer complementarity were assessed on Primer-BLAST [Primer sets, target genes and annealing temperatures are listed in er-BLAST .PCRs were performed with 2.5 U of Amplitaq polymerase , 1 \u00d7 PCR buffer I and 2.5 mM magnesium chloride (Applied Biosystems), 0.2 mM of each deoxyribonucleotide , 0.2 pmol/\u03bcL of each primer, and 1 \u00b5L (1\u20132 ng/\u03bcL) of DNA. Reaction conditions were denaturation at 94 \u00b0C for 2 min, 35 cycles of denaturation at 94 \u00b0C for 30 s, annealing  for 30 sn = 99) were screened for mutations in the QRDR of the gyrA gene [gyrA of the reference C. jejuni (GenBank accession number: L04566.1 and AL111168.1) and C. coli sequences (GenBank accession number: AF092101.1 and NZ_UIGM01000003.1) on SnapGene 2.3.2. Ciprofloxacin-resistant isolates  and the regulatory site of the cmeABC operon in five erythromycin-resistant C. coli isolates are listed in C. coli type strain NCTC 11366 (ATCC 33559) 23S rRNA (GenBank accession number: GQ167698.1), rplD (GenBank accession number: DQ639752.1 and UIGM01000003.1), rplV (GenBank accession number: UIGM01000003.1) and cmeABC operon (GenBank accession number: FJ797670.1) sequences was performed on SnapGene 2.3.2. Isolates were also screened for the presence of ermB, according to Wang et al. (2014). The primers used for the amplification and sequencing of domain V of 23S rRNA, n = 119) were screened for the presence of tetO, according to Aminov et al. (2001) and products were visualised on 2% agarose gel electrophoresis. The tetO amplicon of a tetracycline-resistant C. jejuni isolate (CITCj382-18) from this study was purified and sequenced (as described above) as a positive control. Consensus sequences were aligned to the C. jejuni tetO gene (GenBank accession number: M18896.2).Tetracycline-resistant isolates (tetO/W/O (tetO (GenBank accession number: M18896.2) and mosaic tetO/WO genes (GenBank accession numbers: EF065524.1 and AY196921.1). Two tetO/32/O-targeting primers (tetO (GenBank accession number: M18896.2) and tetO/32/O type I genes with a longer central tet32 region (GenBank accession numbers: AJ295238.3 and JQ740052.1) and tetO/32/O type II genes with a shorter central tet32 segment  [Primers were designed to target tetO/W/O  based on primers  were des00033.1) .n = 4) were screened for sialylated LOS locus class A/B and C (C. jejuni 81-176 (ATCC BAA2151) and C. jejuni NCTC 11168 (DSM 27585), respectively, as positive controls. Streptomycin-resistant isolates  were tested for moxifloxacin susceptibility. Briefly, isolates were recovered from \u221280 \u00b0C on CBA (Fannin Ltd) for 24 h at 42 \u00b0C under microaerobic conditions, and subcultured to CBA for 20 \u00b1 2 h at 42 \u00b0C under microaerobic conditions. In microtiter plates, 100 \u00b5L serial dilutions of moxifloxacin (Sigma Aldrich) were prepared in of Mueller Hinton broth with lysed horse blood  ranging from 0.125\u201316 mg/L. A 0.5 McFarland inoculum was prepared in 5 mL demineralised water (Thermo Fisher Scientific) and 100 \u00b5L was transferred to 11 mL of Mueller Hinton broth with lysed horse blood (Thermo Fisher Scientific). Moxifloxacin serial dilutions were inoculated with 100 \u00b5L of cell suspension and incubated for 20 \u00b1 2 hat 42 \u00b0C under microaerobic conditions.All (fluoro)quinolone-resistant isolates (C. jejuni isolates CITCj625-18 and CITCj727-18 and C. coli isolates CITCc1631-18 and CITCc3448-18) were sequenced. DNA was quantified in triplicate with the Quant-iT dsDNA Assay Kit (Thermo Fisher Scientific). Genomic DNA libraries were prepared using the Nextera-XT protocol , with changes including 2 ng of input DNA and a minute PCR elongation time. DNA quantification and library preparation were performed on a Hamilton Microlab STAR system. Pooled libraries were quantified using the Kapa Biosystems library quantification kit on a Roche light cycler 96 qPCR machine. Libraries were sequenced on the Illumina HiSeq using a 250 bp paired-end protocol. Reads were adapter trimmed using Trimmomatic 0.30 with a sliding window quality cut-off of Q15 [The genomes of four streptomycin-resistant isolates (f of Q15 . De novof of Q15  and assef of Q15 .Genomes were annotated using Prokka 1.14.3 . SimilargyrA GT was submitted for each type. This whole-genome project has been deposited at DDBJ/ENA/GenBank under the accession number PRJNA612628. CITCj625-18, CITCj727-18, CITCc1631-18, and CITCc3448-18 genomes can be accessed using SAMN14379027, SAMN14379028, SAMN14379029, and SAMN14379030. The partial and complete gene sequences deposited in GenBank are listed in Campylobacter spp. hosts. The broiler industry serves as a reservoir for the dissemination of resistant campylobacters. The enrichment and stability of Campylobacter spp. resistance determinants is noteworthy but the natural competence and potential of recombination or acquisition of mobile genetic elements contributes to the Campylobacter. Taken together, the data collected in this study point to the diversity of resistance determinants circulating among Irish broilers, contributing to the development of resistance to clinically relevant antimicrobials. Although non-poultry sources contribute to campylobacteriosis incidence, poultry are natural thermophilic"}
+{"text": "Splenocyte proliferation was tested using thymidine incorporation. Th1 and Th17 cells were analyzed by flow cytometry. Results: Oral KLHTT treatment (50 and 100 mg/kg) ameliorated mouse CIA by decreasing the levels of interleukin (IL)-1\u03b2, IL-6, IL-17A, and tumour necrosis factor-\u03b1 in the paw homogenates and serum. KLHTT also suppressed anti-CII antibody formation, splenocyte proliferation, and splenic Th1 and Th17 cell numbers. Additionally, KLHTT showed antioxidant activity by reducing the concentrations of MDA and H2O2 in paw tissues. Conclusions: The therapeutic effects of KLHTT in CIA mice were through regulating oxidative stress and inflammatory responses. Our results suggest that KLHTT has potential to treat RA.Background: Kan-Lu-Hsiao-Tu-Tan (KLHTT) exhibits anti-psoriatic effects through anti-inflammatory activity in mice. However, the therapeutic effects of KLHTT on rheumatoid arthritis (RA), another significant autoimmune inflammatory disorder, have not been elucidated. Herein, we explored the anti-arthritic effects of KLHTT on collagen-induced arthritis (CIA) in mice. Methods: KLHTT was extracted by boiling water and subjected to spectroscopic analysis. Chicken collagen type II (CII) with complete Freund\u2019s adjuvant was intradermally injected to induce CIA in DBA/1J mice. Anti-CII antibody, cytokines, malondialdehyde (MDA), and hydrogen peroxide (H Rheumatoid arthritis (RA) is an autoimmune disease affecting approximately 1% of the global population, which is characterised by synovitis, cartilage damage, and bone resorption in the joint , cervicaRA is characterised by synovitis accompanied by the infiltration of immune cells , includiKan-Lu-Hsiao-Tu-Tan (KLHTT), a Chinese medicine (CM), has been used to treat inflammatory conditions such as RA, systemic lupus erythematosus, dermatomyositis , sinusit3H labelled thymidine was purchased from Amersham Pharmacia Biotech, Arlington Heights, IL, USA. Brefeldin A and Freund\u2019s adjuvant were bought from Sigma-Aldrich, St. Louis, MO, USA. Phycoerythrin (PE)-conjugated anti-mouse CD4 (clone GK1.5), FITC-conjugated anti-mouse IL-17A (clone TC11-18H10.1), and FITC-conjugated anti-mouse interferon (IFN)-\u03b3 (clone XMG1.2) antibodies were ordered from Biolegend, San Diego, CA, USA. Formalin was bought from AVANTOR, Center Valley, PA. Bovine serum albumin (BSA) was purchased from EMD Millipore, Billerica, MA, USA. Tween 20 was obtained from EMD Millipore, Alsace, France.KLHTT (batch number: 0503-2-403-01) was supplied by Sun Ten Pharmaceutical Corporation, New Taipei City, Taiwan. Chicken collagen type II (CII) was purchased from Chondrex, Inc., Woodinville, WA, USA. Mycobacterium tuberculosis H37RA was bought from Difco Laboratories Inc., Detroit, MI, USA. Methotrexate (MTX) and EDTA were ordered from Bio Basic Inc., Toronto, ON, Canada. Enzyme-linked immunosorbent assay (ELISA) kits  were obtained from eBioscience, San Diego, CA, USA. An IL-17A ELISA kit was purchased from R&D Systems Inc., Minneapolis, MN, USA. A hydrogen peroxide assay kit was purchased from Cell Biolabs, Inc., San Diego, CA, USA. Goat anti-mouse IgG1 and goat anti-mouse IgG2a secondary antibodies were purchased from Jackson ImmunoResearch Laboratories, West Grove, PA, USA. 2,2\u2019-Azinobis [3-ethylbenzothiazoline-6-sulfonic acid]-diammonium salt (ABTS) substrate solution was ordered form Roche Diagnostic Systems, South San Francisco, CA, USA. A Artemisia capillaris Thunb. (seedling), 4.17 g Scutellaria baicalensis Georgi (root), 2.50 g Acorus gramineus Soland. (rhizome), 2.08 g Clematis armandii Franch. (rattan and stem), 2.08 g Fritillaria cirrhosa D. Don (bulb), 1.67 g Pogostemon cablin (Blanco) Benth. (plant shoot), 1.67 g Forsythia suspensa (Thunb.) Vahl (fruits), 1.67 g Amomum kravanh Pierre ex Gagnep. (fruits), 1.67 g Mentha haplocalyx Briq. (stem and leaf plot), and 1.67 g Belamcanda chinensis (L.) DC (rhizome)) was extracted by boiling water (12 times the weight of the herbs) for 1 h, and then concentrated to a voucher specimen (CGU_KLHTT-01) by the freeze dryer  [The herbs of KLHTT were purchased and identified by Sun Ten Pharmaceutical Corporation, New Taipei City, Taiwan. A total of 27.93 g of herbs  comprising an LC-30AD pump, SIL-30AC auto-sampler, CTO-20AC column, and SPD-M20A photodiode array detector . Prior to being loaded onto the UPLC column, 1 mg of KLHTT extract was first dissolved in 1 mL of methanol and filtered through a 0.45 \u03bcm membrane. Sample injections of 1 \u03bcL were then performed automatically. Liquid chromatography was performed using a CORTECS UPLC C18 column  . The mobile phase was a mixture of MeCN (A) and water . A gradient sequence was executed as follows: 0\u201310 min, 10\u201320% A; 10\u201314 min, 20\u201325% A; 14\u201324 min, 25\u201330% A; 24\u201328 min, 30\u201340% A; 28\u201333 min, 40\u201350% A; 33\u201338 min, 50\u201375% A; 38\u201340 min, 75\u2013100% A; and 40\u201343 min, 100% A. The column temperature was set at 35 \u00b0C. The flow rate was at 0.4 mL/min. The range of detection wavelengths was fixed in the 190\u2013500 nm.Multiple reaction monitoring (MRM) experiments (in negative) were carried out using Shimazu LCMS-8045 triple quadrupole mass spectrometry to identify the constituents of KLHTT extract. The precursor ion settings of the corresponding profiling peaks were determined using the full scan experiment (50\u20131000 amu). The product ions were settled according to previously reported data. The dwell time was fixed at 100 ms and the collision energy was set at 25\u201345 eV. All MS data were acquired and processed using LCMS LabSolutions software Version 5.93 .DBA/1J mice  were purchased from Jackson Laboratories  and maintained at 20\u201325 \u00b0C with half day light/dark cycle under a specific pathogen-free condition. All mice were treated according to the guidelines of the Institutional Animal Care and Use Committee of National Chung Hsing University (NCHU). The study protocol was approved by NCHU ethics committee .2O, KLHTT, or MTX was administered orally once a day from day 21 to 42. Mice were divided into four groups (n = 6/group) randomly as follows: Group I, Normal; Group II, Vehicle (ddH2O) + CII; Group III, KLHTT (50 mg/kg) + CII; Group IV, KLHTT (100 mg/kg) + CII. MTX (0.5 mg/kg) was used as a positive control. Mice were euthanized with CO2 exposure (100% CO2 for 5 min) by experienced experimenters humanely on day 42. The arthritis severity score was recorded every 3 days after the treatment of drugs. The biochemical and histological assays were performed on day 42.CIA was induced by active immunisation with chicken CII . BrieflyThe body weight and arthritis severity score were obtained . The artAfter the mice were humanely sacrificed, the hind limbs were fixed in 10% buffered formalin, decalcified in 15% EDTA, and embedded in paraffin. Serial paraffin sections (5 \u03bcm) were stained with haematoxylin and eosin. The severity of histopathological lesions was scored  as folloHind paw was dissected and homogenised in ice-cold saline using a tissue homogeniser. After being centrifuged at 3000 rpm , the hind paw homogenates were harvested. Blood was collected from the heart. The levels of cytokines in hind paw homogenates and serum were measured by ELISA .2O2) concentration was measured using a colorimetric OxiSelect\u2122 hydrogen peroxide assay kit at 560 nm [Malondialdehyde (MDA) concentration was determined by thiobarbituric acid reactive substances assay at 532 nm. The standard curve was obtained using 1,1,3,3-tetramethoxypropane. Hydrogen peroxide  at 4 \u00b0C overnight. The plates were washed and incubated with goat anti-mouse secondary antibodies IgG1 (1:500 dilution) or IgG2a (1:500 dilution) at 25\u201327 \u00b0C for 1 h. After being washed, ABTS substrate was added and the reactions were stopped by adding H2SO4. The level of IgG1 and IgG2a was measured at 450 nm by an ELISA reader  [Serum samples were diluted 1:250 for IgG1 or 1:125 for IgG2a in Tris-buffered saline , and then transferred to CII (10\u2009\u03bcg/mL) pre-coated 96-well plates (Microtiterzerland) .5 cells/well) were cultured with chicken CII (50 \u03bcg/mL) at 37 \u00b0C for 40 h, and then incubated with 3H for 8 h. Cell proliferation was evaluated by radioactive thymidine incorporation [Splenocytes (4 \u00d7 10poration .6 cells/well) were cultured with chicken CII (50 \u03bcg/mL) at 37 \u00b0C for 48 h, and then brefeldin A (5 \u03bcg/mL) was added for 6 h. Cells were harvested and extracellularly stained with PE-conjugated anti-mouse CD4 antibodies. After being fixed and permeabilised with Cytofix/Cytoperm solution (BD Pharmingen), cells were then intracellularly labelled with FITC-conjugated anti-mouse IL-17A and anti-mouse IFN-\u03b3 antibodies. Splenocytes were detected by an Accuri C5 flow cytometer  and analysed by BD Accuri\u2122 C6 Plus software [Splenocytes , chrysin 6-C-glucoside-8-C-arabinoside (2), baicalin (3), norwogonin-7-O-\u03b2-d-glucuronide (4), chrysin 7-O-\u03b2-d-glucuronide (5), oroxylin A 7-O-\u03b2-d-glucuronide (6), wogonoside (7), and baicalein (8)  B (Table lein (8) .The CIA mouse model is a well-established and commonly used model mimicking the clinical symptoms and immunopathogenesis of human RA . ImmunisHistopathological analysis of the CIA mice revealed inflammatory cell infiltration into articular tissues, exudates within the synovial space, synovial hyperplasia, and cartilage erosion. KLHTT-treated mice demonstrated well-preserved joint spaces with minimal inflammatory exudates, normal cartilage structure, and clear synovial spaces, along with improved histological arthritis severity scores . MTX 0..The pathogenesis of RA involves activated T cells promoting macrophages to release pro-inflammatory cytokines . Therefo2O2 were noted in the hind paw homogenates of CIA mice. KLHTT (50 and 100 mg/kg) significantly reduced the levels of MDA  and MDA (a lipid peroxidation marker) in joint homogenates from CIA mice. MDA-related reactions are highly immunogenic. MDA levels correlate with RA severity and can be used to predict RA severity [Oxidative stress is involved in the pathogenesis of RA ,43. In tseverity . The newseverity . RA patiseverity ,47. Inflseverity . Adalimuseverity . In thisC-arabinoside-8-C-glucoside, chrysin 6-C-glucoside-8-C-arabinoside, baicalin, norwogonin-7-O-\u03b2-d-glucuronide, chrysin 7-O-\u03b2-d-glucuronide, oroxylin A 7-O-\u03b2-d-glucuronide, wogonoside, and baicalein. Previous studies have reported that baicalin ameliorated CIA in mice by down-regulating Janus kinase 1/signal transducer and activator of transcription 3 signalling and inhibiting IL-17-mediated joint inflammation [We identified eight flavonoids from KLHTT, including chrysin 6-ammation ,51. Baicammation . Intrapeammation . Chrysinammation ,55. The ammation . ObservaIn summary, our results indicate that KLHTT, a CM formulation, shows significant anti-inflammatory effects, antioxidant activities, and immunomodulatory functions in CIA mice. The present study also demonstrates that KLHTT has potential to treat RA."}
+{"text": "Helicobacter pylori (H. pylori) is one of the most common human pathogens, affecting half of the world\u2019s population. Approximately 20% of the infected patients develop gastric ulcers or neoplastic changes in the gastric stroma. An infection also leads to the progression of epithelial\u2013mesenchymal transition within gastric tissue, increasing the probability of gastric cancer development. This paper aims to review the role of H. pylori and its virulence factors in epithelial\u2013mesenchymal transition associated with malignant transformation within the gastric stroma. The reviewed factors included: CagA (cytotoxin-associated gene A) along with induction of cancer stem-cell properties and interaction with YAP (Yes-associated protein pathway), tumor necrosis factor \u03b1-inducing protein, Lpp20 lipoprotein, Afadin protein, penicillin-binding protein 1A, microRNA-29a-3p, programmed cell death protein 4, lysosomal-associated protein transmembrane 4\u03b2, cancer-associated fibroblasts, heparin-binding epidermal growth factor (HB-EGF), matrix metalloproteinase-7 (MMP-7), and cancer stem cells (CSCs). The review summarizes the most recent findings, providing insight into potential molecular targets and new treatment strategies for gastric cancer. Helicobacter pylori (H. pylori) is a helix-shaped, Gram-negative, microaerophilic, flagellated bacterium that is capable of biofilm formation and converting from spiral to coccoid form ..51].As a physiological process, EMT is observed during organogenesis, tissue development, remodeling, and wound healing ,53,54; cc-MYC [During this process, epithelial cells undergo a series of biochemical changes, which lead to the loss of polarity and migratory capacity of cells, resulting in cell shape changes (cell elongation). EMT promotes the transformation of immobile epithelial cells into motile mesenchymal cells, enhancing the metastatic properties ,70. Furtc-MYC ,77,78,79c-MYC . Moreovec-MYC ,82,83.H. pylori infection significantly affects the gastric microenvironment by induction of several inflammatory responses via infiltrating macrophages, neutrophils, regulatory T-cells, and natural killer cells [H. pylori virulence factors are considered being associated with the promotion of EMT in gastric cells, which consequently causes neoplasia and malignant transformation. This review summarizes several mechanisms associated with epithelial\u2013mesenchymal transition, gastric tumor microenvironment, and the influence of H. pylori infection, although some described mechanisms are not only H. pylori-specific. Even though H. pylori-induced carcinogenesis is not fully understood, several mechanisms have already been deciphered.er cells ,85. Infler cells ,87,88. Ter cells ,90,91. AH. pylori, cag pathogenicity island (cagPAI) probably plays a key role in carcinogenesis [CagA oncoprotein [CagA into a cell, transforming its shape into the so-called \u201chummingbird phenotype\u201d characterized by an elongated cell shape commonly observed in EMT [CagA into the cell via the T4SS induces signal transduction, with one of the most relevant mecahnisms being the nuclear factor \u03baB (NF-\u03baB) signaling pathway involving extracellular regulated kinases 1/2 (ERK-1/2) [H. pylori [CagA in host cells is tyrosine phosphorylated and interacts with protein tyrosine phosphatase 2 (SHP-2), also inducing the progression of the \u201chummingbird phenotype\u201d [CagA enhances EMT via the stabilization of Snail protein, which is essential in carcinogenesis, mainly by the reduction of glycogen synthase kinase-3 (GSK-3) activity [CagA-positive H. pylori strains with CagA containing phosphorylation-functional EPIYA motifs present significantly higher expression of mesenchymal markers such as vimentin, Snail, and ZEB-1 and the stem cell marker CD44 [CagA-positive H. pylori strains induce a higher probability of gastric carcinogenesis and induction of EMT process [CagA-positive H. pylori infection present poor clinical outcome, and higher invasion and metastatic characteristics [Among many virulence factors of ogenesis ,93,94. Ioprotein ,96. The d in EMT ,98. InjeERK-1/2) ,100,101.ERK-1/2) . The inh. pylori . CagA inenotype\u201d ,105. Cagactivity . CagA-poker CD44 ,109,110. process ,112,113.eristics . Besideseristics ,116.CagA-positive H. pylori strain infection [Recent research suggests that cells that undergo EMT obtain the ability to acquire cancer stem cell (CSC) properties ,122,123.nfection ,129. Hignfection ,135,136.H. pylori in the generation of cells with CSC properties, including several gastric epithelial cell lines  [CagA in the induction of CSC properties was studied on H. pylori 7.13 wild-type (WT) CagA-positive strain and its knock-out mutants\u20147.13CagA-negative and 7.13DCagE-negative H. pylori strains. Only the wild-type (WT) (7.13CagA-positive) strain-induced mesenchymal changes and EMT in cells; cells infected by CagA-positive strains, presented higher expression of mesenchymal markers\u2014CD44, Snail1, vimentin, or zinc finger E-box binding homeobox 1 (ZEB1), while an expression of epithelial markers\u2014cytokeratin 7 (CK7) or osteopontin (SSP-1) was lowered [CagA-positive H. pylori strains. The relevance of the infection by CagA-positive H. pylori strain on EMT in gastric cancer was confirmed by significantly higher expression of CD44 and mesenchymal markers in tumor samples.Bess\u00e8de et al. (2014) studied the role of  MKN-74) . The rol lowered . The migCagA-positive H. pylori EMT and the induction of CSC properties [CagA-positive H. pylori infection with N-nitrosoguanidine (MNNG) stimulation causes CSC features with dysplastic lesions and mesenchymal phenotype. Amieva and Peek (2016) observed that CD44+ cells with CSC features displayed increased CagA half-life [CagA was confirmed in CD44+ gastric CSCs [H. pylori-infected (CagA-positive) gastric cancer cells exhibit CSC properties via increased expression of surface markers\u2014CD44 and Lgr5 with Nanog, Oct4, and c-myc upregulation [CagA impairs transcription factor CDX1 expression, promoting EMT, and CSC features in gastric epithelial cells [The Wnt/\u03b2-catenin signaling pathway is involved in operties . Likewisalf-life . Accumulric CSCs . H. pylogulation ,140. Cagal cells .MYC oncogene [Yes-Associated-Protein (YAP) is an element of the Hippo tumor suppressor signaling pathway, which plays a crucial role in the maintenance of the proper size of organs, tissue homeostasis, cell proliferation, and stem cell maintenance ,143,144.oncogene ,149. Besoncogene .CagA-positive H. pylori strain induces higher YAP expression and decreases E-cadherin, N-cadherin, and Slug levels, promoting EMT.Li et al. (2018) showed a significantly higher expression of YAP and TAZ  in cancerous gastric tissues with a positive correlation between YAP, TAZ levels, and tumor size . During CagA, in the enhancement of the YAP pathway [CagA-positive H. pylori upregulates YAP1 and large tumor suppressor 2 (LATS2) expression in gastric epithelial cells [YAP1 is associated with aggressiveness and poor prognosis [YAP1/SLC35B4 axis; another mechanism involves altered IL-1\u03b2 levels [Other studies have shown a potential role of GSK3/\u03b2-catenin or AMP-activated protein kinase (AMPK) pathways, which are regulated by  pathway ,153. Molal cells . Overexprognosis . Gastric\u03b2 levels ,157. TheH. pylori strains produce a high quantity of tumor necrosis factor-\u03b1 (TNF-\u03b1)-inducing protein (Tip\u03b1 protein), which binds to the cell surface nucleolin and induces carcinogenic alterations [H. pylori virulence [\u03b1 induces mesenchymal changes of cells via EMT progression [\u03b1, as a carcinogenic factor, activates NF-\u03baB, promoting gastric carcinogenesis by inhibiting miR-3178 expression [\u03b1 release, Tip\u03b1 also induces the expression of several chemokines, including chemokine (C-C motif) ligand 2 (Ccl2), chemokine (C-C motif) ligand 7(Ccl7), chemokine (C-C motif) ligand 20 (Ccl20), C-X-C motif chemokine 11 (Cxc11), chemokine (C-X-C motif) ligand 2 (Cxcl2), C-X-C motif chemokine 5 (Cxcl5), and C-X-C motif chemokine 10 (Cxcl10) [\u03b1 (TNF-\u03b1), or hypoxia-inducible factor 1\u03b1 (HIF1\u03b1) [\u03b1, along with CagA and VacA, plays an underlying role primarily in mucosal damage [erations ,162,163.irulence ,165. By gression ,167,168.pression ,170. Bes(Cxcl10) ,173,174. (HIF1\u03b1) ,176. In l damage ,178.H. pylori infection increases the quantity of Tip\u03b1 protein in gastric cancer tissue, inducing tumor progression in H. pylori carcinogenesis [\u03b1 reduces cell stiffness and phosphorylates various oncoproteins; it also enhances filopodia formation, morphological, and conformational changes within cells, and expression of vimentin via MEK-ERK phosphorylation, confirming its role in EMT progression [Suganuma et al. showed that ogenesis . Tip\u03b1 regression ,180.\u03b1 and nucleolin receptor (88 kDa) on the migration properties of gastric cancer cells [\u03b1 could be inhibited via small interfering RNAs targeted for the nucleolin receptor, showing its role in EMT progression [H. pylori infection by inhibiting Tip\u03b1 and nucleolin interaction [Researchers have suggested the role of Tiper cells ,181. Fujgression . Thus, peraction ,184.\u03b1 induces the formation of filopodia and a reduction of cell stiffness, predisposing cells to higher motility [\u03b1 induces higher levels of IL-1\u03b2, TNF-\u03b1, and IL-8 in SGC7901 cells [H. pylori infection promotes the Il-6/STAT3 signaling pathway in AGS cells, which is one factor promoting EMT [\u03b1 did successfully eradicate H. pylori infection [Tipmotility . It has 01 cells . Furtherting EMT . Inoue enfection .H. pylori virulence factor, promoting its colonization and survival properties [H. pylori, which enhances the stimulation of cell proliferation. Lpp20 expression is increased in the acidic pH of stomach juice and during iron depletion [The Lpp20 protein plays an important role as an operties . Lpp20 iepletion ,190.\u03b1), is one of the carcinogenic factors released by H. pylori via NF-\u03baB pathway activation [\u03b1 and Lpp20 induce EMT mainly by stimulating cell proliferation, migratory properties of cells, and filopodia formation [Lpp20 , these proteins show similar effects on gastric cancer cells by stimulating EMT and gastric cancer progression.Even though Lpp20 and TipH. pylori lowered the expression of Afadin protein independently of CagA and VacA in MKN74 and NCI-N87 cell lines [H. pylori.Afadin is a cytoplasmic actin-filament-binding protein responsible for the formation and stabilization of tight and adherens junctions ,194,195.ll lines . DownregH. pylori with an affinity for penicillin binding; any modifications within PBP might induce resistance to \u03b2-lactam antibiotics  [PBP2X, PBP2B, and PBP1A [H. pylori PBPs might present unique characteristics because of several putative genes encoding PBPs.Penicillin-binding protein (PBP) is a specialized acyl serine transferase released by icillin) . These mnd PBP1A . It was PBP1A mutation-positive H. pylori (H. pylori CagA+/P+) strain and CagA+/P\u2212 in terms of clinicopathological characteristics of gastric cancer and EMT induction [H. pylori CagA+/P+ infection resulted in progressive and clinically significant EMT and gastric cancer severity. Researchers have observed decreased levels of E-cadherin and increased \u03b1-smooth muscle actin (\u03b1-SMA) levels; likewise, it was observed that PBP1A mutation is associated with decreased miR-134 levels. MiR-134 suppresses gastric carcinogenesis by targeting the KRAS gene and Golgi phosphoprotein 3 (GOLPH3) downregulation; another essential target gene of miR-134 is FoxM1 [H. pylori activity decline [Huang et al. 2019) showed significant differences between nduction . H. pylois FoxM1 ,201. The9 showed  decline . Marcus  decline .Micro-RNAs constitute a family of small noncoding RNAs, which are regulators of posttranscriptional gene expression by binding to the 3\u2032-untranslated region of mRNA, promoting an inappropriate translation of targeted mRNA. There is an increasing interest in micro-RNA involvement in many diseases and malignancies ,206,207.H. pylori [H. pylori infection induced the overexpression of microRNA-29a-3p in gastric cancer tissue samples, which resulted in an enhanced ability of the migration of gastric epithelial cells. Furthermore, the silencing of A20 increased the EMT markers Snail, vimentin, and N-cadherin, and decreased E-cadherin levels. The authors have suggested that the overexpression of micro-RNA-29a-3p might be associated with a higher probability of EMT induction in gastric cells.Sun et al. presented the relevance of upregulation of microRNA-29a-3p and its role in the decreased expression of A20 in patients infected by . pylori . A20 is . pylori . H. pyloTWIST1 gene [Programmed cell death protein 4 (PDCD4) is a tumor suppressor responsible for the inhibition of cell growth, tumor invasion, metastasis, or induction of apoptosis . It is lST1 gene .TWIST1, and vimentin levels in gastric cancerous tissues [CagA-positive H. pylori strain infection. PDCD4 overexpression reduced EMT. MicroRNA-21 overexpression in gastric cancer modulates the expression of the tumor suppressors phosphatase and tensin homolog (PTEN) and PDCD4, altering molecular pathways associated with cell growth, invasion, migration, and apoptosis [PDCD4 is one factor responsible for the induction of carcinogenesis . Yu et a tissues . PDCD4 dpoptosis .LAPTM4B) is a proto-oncogene relevant in the progression of tumorigenesis, regulating molecular mechanisms, and cellular functioning, including proliferation, migration, and invasion [LAPTM4B upregulation has been observed in various cancers, including hepatocellular carcinoma, breast cancer, and lung adenocarcinoma [Lysosomal-associated protein transmembrane 4\u03b2  while lowering E-cadherin levels and dysregulating cell\u2013cell junctions. Additionally, there was a noticeable increase of ZEB1, \u03b2-catenin, Snail, and Slug markers, whereas tight junction protein-1  level was decreased. LAPTM4B seems to be a factor strictly associated with EMT induction since it is responsible for regulating cells\u2019 filopodia, increasing motility of gastric cancer cells, and a higher invasion potential [LAPTM4B results in poorer prognosis in various cancers [It has been reported that ormation . Researcotential . Overexp cancers .\u03b1 in fibroblasts, which is common during CagA+VacA+ H. pylori infection [Physiologically, natural gastric stroma contains only a few fibroblasts, and those are mainly myofibroblasts. The number of fibroblasts is significantly increased in tissues affected by inflammation or neoplasm ,230,231.nfection .H. pylori infection results in the transformation of fibroblasts and myofibroblasts into CAFs [CagA+VacA+ H. pylori infection results in the overexpression of fibroblast activation protein (FAP), fibroblast surface protein (FSP) mRNA, and increased levels of the proinflammatory factors IL-6, IL-8, COX-2, and SDF-1 [\u03b1-SMA, N-cadherin, vimentin, Snail, Twist) and lowered levels of E-cadherin, antiapoptotic B-cell lymphoma 2 (Bcl-2), and proliferative marker Ki-67 protein within fibroblasts [CagA+VacA+ H. pylori infection resulted in the aberrant apoptosis process, an increased level of collagen production, and impaired fibroblast and myofibroblast differentiation into CAFs. This induced desmoplastic reactions and EMT within gastric cells, promoting carcinogenesis. The increased levels of \u03b21-integrin and COX-2, which are crucial in invasion, metastasis, and angiogenesis, enhanced the migration properties of the infected cells. CagA+Vac+ H. pylori infection results in the upregulation of HIF-1\u03b1 and tenascin-C (TNC), enhancing EMT and tumor progression, the release of proangiogenic factors, and further CAFs activation [CagA+Vac+ H. pylori infection activates gastric fibroblasts and induces the release of TGF-\u03b2 [nto CAFs . CagA+Vand SDF-1 . Overexpnd SDF-1 ,248,249.roblasts . CagA+Vativation ,251,252.of TGF-\u03b2 .The tumor-promoting ability of CAFs is enhanced by microRNA dysregulation . CAFs stH. pylori infection results in the upregulation of HB-EGF gene expression, which is a potential serological biomarker for gastric cancer [H. pylori g-glutamyltranspeptidase, activating PI3K, and p38 kinase-dependent signal transduction pathways [Heparin-binding epidermal growth factor (HB-EGF) is an epidermal growth factor receptor (EGFR) cognate ligand with mitogenic and chemotactic properties for fibroblasts and smooth muscle cells . H. pyloc cancer ,266,267.c cancer . HB-EGF pathways .H. pylori infection and is associated with EMT [H. pylori infection [Matrix metalloproteinase-7 (MMP-7) is an enzyme responsible for the degradation of adherence between cells in the extracellular matrix, and overexpression of MMP-7 induces a more aggressive course of gastric cancer ,272,273.with EMT ,275. Liknfection ,278,279.nfection ,280.H. pylori infection results in the increased release of MMP-7 and soluble HB-EGF, promoting EMT [H. pylori-induced inflammation and damage via M1 macrophage polarization suppression [Yin et al. showed that excessive gastrin secretion due to ting EMT . HB-EGF ting EMT . Besidesting EMT ,284,285.ting EMT . Gastricting EMT ,288. Howpression .H. pylori infection, neoplasia, or cancer, being involved in the migration properties of gastric cancer cells [Mesenchymal stem cells (MSCs) present a multipotent potential to differentiate into various cell types, while keeping a capacity for self-renewal ,291. MSCer cells ,293,294.er cells ,296. Naner cells .H. pylori gained a fibroblastic phenotype, enhancing EMT progression in gastric cancer cells [H. pylori showed a significant decrease in the levels of E-cadherin and overexpression of mesenchymal markers  and inflammatory cytokines , VEGF, epidermal growth protein (EGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and monocyte chemoattractant protein 1 (MCP-1). High concentrations of IL-15 secreted by gastric cancer MSCs contribute to tumor cell metastasis; increased IL-17 levels promote gastric cancer invasiveness [Zhang et al. showed that human umbilical cord MSCs (hucMSCs) infected by er cells . Furthersiveness ,300.SOX4 induces EMT via a release of TGF-\u03b2 and contributes to the development of stem cell characteristics of gastric cancer cells [CagA-positive H. pylori infection induces EMT, and CSCs feature in gastric mucosal cells via overexpression of the Wnt/\u03b2-catenin signaling pathway [KRAS, STAT3, Rac1, Wnt, Notch, PTEN, ERK, NF-\u03baB signaling pathways, or hypoxic microenvironment [Increased expression of er cells . Furtherer cells . CagA-po pathway . The pro pathway ,306,307. pathway . Besidesironment ,317,318.ironment .H. pylori infection and gastric carcinogenesis remains controversial. In fact, some mechanisms are yet not fully understood. H. pylori infection seems to play an important role in terms of EMT induction within gastric mucosa (The association between c mucosa .H. pylori virulence factors. Likewise, cellular components that are associated with EMT progression can be influenced by H. pylori infection (H. pylori along with its properties and virulence factors (H. pylori during the early stages of infection, which could significantly decrease the number of gastric cancer incidents.The severity and progression of gastric cancer depends on the presence of specific nfection . EMT and factors . Specifi"}
+{"text": "Escherichia coli isolates, four (13.79%) Klebsiella pneumoniae isolates, and one (3.45%) Enterobacter cloacae isolate. The analysis based on the nucleotide sequences of the ESBL resistance genes showed that all cefotaximase-Munichs (CTX-Ms) were CTX-M-15 and that all sulfhydryl variables (SHVs) were SHV-11: 41.67% CTX-M-15-producing E. coli, 16.67% CTX-M-15+SHV-11-producing E. coli, 8.33% CTX-M-15-producing K. pneumoniae, 25% CTX-M-15+SHV-11-producing K. pneumoniae, and 8.33% CTX-M-15-produced E. cloacae. This study shows for the first time the presence of multiresistant ESBL-producing enterobacteria in fruit bats in Makokou.In Gabon, terrestrial mammals of protected areas have been identified as a possible source of antibiotic-resistant bacteria. Some studies on antibiotic resistance in bats have already been carried out. The main goal of our study was to detect extended-spectrum beta-lactamases (ESBLs) that are produced by enterobacteria from bats in the Makokou region in Gabon. Sixty-eight fecal samples were obtained from 68 bats caught in the forests located 1 km from the little town of Makokou. After culture and isolation, 66 Gram-negative bacterial colonies were obtained. The double-disk diffusion test confirmed the presence of ESBLs in six (20.69%)  Bats are an important, widespread, and abundant taxon of mammals with over 1300 described species . Bats haThe existence of antibiotic multi-resistant (AMR) enterobacteria that have been recurrently documented in wildlife in recent years provide important insights into the potential role of wildlife as reservoirs of resistant bacteria. AMR enterobacteria are also pertinent markers to understand the dynamics of zoonosis and the complex transmission routes among wildlife and humans . Many waPteropus poliocephalus), McDougall et al. detected the presence of resistance to several antibiotic families in 5.3% of wild flying foxes from different Australian colonies , CH17 (2) CTX-M-15 [MK559065], CH18 (3) CTX-M-15 [MK559057], CH38 (2) CTX-M-15 [MK559063], CH41 (1) CTX-M-15 [MK559058], CH41 (2) CTX-M-15 [MK559064], CH42 (1) CTX-M-15 [MK559059], CH42 (2) CTX-M-15 [MK559060], CH43 (1) CTX-M-15 [MK559066], CH71 (1) CTX-M-15 [MK559061], CH82 (1) CTX-M-15 [MK559062], CH38 (2) SHV-11 [MK590053], CH42 (2) SHV-11 [MK590054] and CH43 (1) SHV-11 [MK590055].Epomops franqueti and 22 Megaloglossus woermanni were caught; a total of 68 bats of the Pteropodidae family were collected. Of the 68 bacterial strains identified, 66 were Gram-negative bacteria (GNB). Among the GNB, 29 (42.65%) were enterobacteria, which consisted of 11 (37.93%) Escherichia coli, five (17.24%) Klepsiella pneumoniae, three (10.34%) Enterobacter aerogenes, two (5.88%) Enterobacter cloacae, two (6.89%) Serratia plymuthica, one Citrobacter freundii (3.45%), one (3.45%) Enterobacter hormaechei, one (3.45%) Ewingella americana, one (3.45%) Morganella morganii, one (3.45%) Pantoea sp., and one (3.45%) Proteus vulgaris  K. pneumoniae isolates, and one (3.45%) E. cloacae isolate  ESBL-producing enterobacteria. Among beta-lactams, resistance was observed by using amoxicillin (100%), ticarcillin (100%), cefotaxime (100%), ceftazidime (100%), cefpodoxime (100%), aztreonam (100%), ticarcillin/clavulanic acid (90.90%), piperacillin (90.90%), cephalexin (100%), cefepime (81.81%), amoxicillin/clavulanic acid (54.54%), cefoxitin (54.54%), ertapenem (36.36%), piperacillin/tazobactam (54.54%), and imipenem (0.00%) .As for other antibiotics tested on these positive ESBL strains, resistance was observed as follows: Erythromycin (100%), streptomycin (100%), ciprofloxacin (90.90%), kanamycin (81.81%), tetracycline (81.81%), trimethoprim/sulfamethoxazole (72.72%), gentamycin (63.63%), nalidixic acid (63.63%), tobramycin (63.63%), colistin (54.54%), ofloxacin (54.54%), levofloxacin (45.45%), netilmicin (36.36%), amikacin (27.27%), nitrofurantoin (18.18%), and chloramphenicol (0%) .E. franqueti bats species and four ESBL-producing Enterobacteriae from M. woermanni bats species were identified. These bats belong to the family of Pteropodidae -producing E. coli, 9.09% ESBL (CTX-M-15)-producing K. pneumoniae, 27.27% ESBL -producing K. pneumoniae and 9.09% ESBL (CTX-M-15)-producing E. cloacae _SHV-11, CH43 (2)_SHV-11, CH42 (2)_SHV-11 and CH38 (2)_SHV-11) clustered with human bacterial strains from Tunisia  and E. cloacae and K. pneumoniae (9.09%) or was associated with SHV-11 in K. pneumoniae (27.27%). The prevalence of ESBL-producing enterobacteria (37.93%) was higher than that obtained in E. coli sampled in bats and other wild animals in Nigeria [K. pneumoniae was found in a gorilla in its natural habitat in the Dzanga Ndoki National Park [Tadarida teniotis (bats) in Portugal [E. coli, followed by K. pneumoniae [blaCTX-M beta-lactam resistance genes are the most common in the human and veterinary strains [blaCTX-M-15 is the most prevalent ESBL gene in human samples worldwide [blaCTX-M-15 and blaSHV-11 genes are recognized as plasmid-mediated resistance genes [The beta-lactam resistance phenotypes obtained in our study are also comparable to those obtained in a study on bats in Algeria , which s Nigeria ,15 and t Nigeria , in whiceumoniae .27%. Thenal Park ,46 and iPortugal . Enterobeumoniae . However strains ,49,50. borldwide  and is porldwide . In addice genes . All of ce genes ,52,53. blaCTX-M-15 genes had already been identified in ESBL-producing Enterobacteriaceae from patients in the Albert Schweitzer Hospital in Lambar\u00e9n\u00e9 and in poultry [blaCTX-M-15 and blaSHV-11 genes is probably linked to antibiotics used by humans, since our phylogenetic analyses show that all the sequences obtained in bats that carry CTX-M-15 or SHV-11 clustered with one human bacterial strain from Turkey and Tunisia, respectively. This suggests that the prevalence of antibiotic resistance in wild animals depends on the antibiotics consumed by human populations at each site and the density of the human population in contact with fauna [In Gabon, the  poultry ,32. The th fauna .E. coli isolated from wildlife may be acquired from the food [E. coli has already been described as an important source of infection in mammals [E. coli infections can result from food contamination [Tetrapleura tetraptera, the fruit of which is widely consumed in Gabon) and lemon. Hence, it is possible that some fruits partly eaten by humans and contaminated have been ingested by bats. Fruit bats are potential vectors and reservoirs of pathogenic bacteria such as E. coli, which carry acquired resistance [In our samples, the highest prevalence of resistance was obtained in resistance to aminoglycosides , fluoroquinolones  and tetracycline, which are the most common antibiotics consumed by the Gabonese people ,54. The the food ,57,58,59the food  and watethe food ; resistathe food ,52. Furt mammals . Foodbormination ,63. Secomination ,64. Mostmination ,66,67,68mination ,70. Thesmination . Severalmination ,72,73. Tsistance ,59. The sistance ,75. Batssistance . In thisThe phylogeny of CTX-M-15 was constructed using v. 1.8.1 in BioEdit v. 7.0.9.0 software. These analyses were performed with a multiple alignment matrix of obtained partial CTX-M-15 sequences and the GenBank reference sequences of human, poultry and swine from Africa (Nigeria), Asia , Europe , the Middle East (Iran), and South America (Brazil). Enterobacteriaceae carrying CTX-M-15 isolated from fruit bats of Makokou in Gabon (in pink color).K. pneumoniae from bats. Enterobacteriaceae carrying SHV-11 isolated from fruit bats of Makokou in Gabon (in pink color).The phylogeny of SHV-11 was constructed using v. 1.8.1 in BioEdit v. 7.0.9.0 software. These analyses were performed with a multiple alignment matrix of obtained partial SHV-11 sequences and the GenBank reference sequences of human, water and cattle from Europe , Asia , the Middle East (Iran), and Africa (Tunisia and Egypt). All sequences of SHV-11 were from This study showed for the first time the presence of multiresistant ESBL-producing enterobacteria in Makokou fruit bats in Gabon . The source of the contamination has not been clearly determined, but the presence of this type of resistance in bats suggests that these wild mammals could spread ESBL-producing Enterobacteriacea over long distances or across the urban landscape.Our results reinforce the need to monitor antimicrobial resistance in wild animals, in protected or unprotected areas, in order to assess environmental responses to anthropogenic pressures."}
+{"text": "Scientific Reports 10.1038/s41598-019-41524-3, published online 01 April 2019Correction to: The Acknowledgements sectionin this Article is incomplete.\u201cWe are grateful to Dr. D.W. Stamer and his collaborators from Duke University Eye Center  for their gift of human primary TM cells. The present work was supported by Ministerio de Economia y Competitividad and Instituto de Salud Carlos III of Spain FIS PI14/00141 (X.G.), FIS P17/00296 (X.G.), RETICs Oftared RD12/0034/0003 (X.G.), RD16/0008/0014 (X.G.), RD12/0034/0015 (M.C.), RD16/0008/0023 (M.C.), Generalitat de Catalunya 2014SGR1165 (X.G.), and National Eye Institute - NIH, EY-11906 (T.B.).\u201dshould read:\u201cWe are grateful to Dr. D.W. Stamer and his collaborators from Duke University Eye Center  for their gift of human primary TM cells. The present work was supported by European Union, Fondo Europeo de Desarrollo Regional (FEDER), Ministerio de Economia y Competitividad and Instituto de Salud Carlos III of Spain FIS PI14/00141 (X.G.), FIS P17/00296 (X.G.), RETICs Oftared RD12/0034/0003 (X.G.), RD16/0008/0014 (X.G.), RD12/0034/0015 (M.C.), RD16/0008/0023 (M.C.), Generalitat de Catalunya 2014SGR1165 (X.G.), and National Eye Institute - NIH, EY-11906 (T.B.).\u201d"}
+{"text": "In: Imsanguan W., Bupachat S., Wanchaithanawong V., Luangjina S., Thawtheong S., Nedsuwan S., et al. Contact tracing for tuberculosis, Thailand. Bull World Health Organ. 2020 Mar 1;98(3):212\u201318 On page 215, Table 2, should read as follows:"}
+{"text": "M-SAL yielded 29.7% ND and average concentrations of 2.26 \u00b1 1.15 (SD) log10 PFU/L (somatic) and 0.59 \u00b1 0.82 (SD) log10 PFU/L (F+ ). DMF performance was inferior to D-HFUF-SAL and M-SAL methods , indicating this procedure is unsuitable for 1 L surface water sample volumes. This study represents an important step toward the use of a coliphage method for recreational water quality criteria purposes.Coliphages are alternative fecal indicators that may be suitable surrogates for viral pathogens, but majority of standard detection methods utilize insufficient volumes for routine detection in environmental waters. We compared three somatic and F+ coliphage methods based on a paired measurement from 1 L samples collected from the Great Lakes (n = 74). Methods include: 1) dead-end hollow fiber ultrafilter with single agar layer (D-HFUF-SAL); 2) modified SAL (M-SAL); and 3) direct membrane filtration (DMF) technique. Overall, D-HFUF-SAL outperformed other methods as it yielded the lowest frequency of non-detects  and the highest average concentrations of recovered coliphage for positive samples (2.51 \u00b1 1.02  log Enteric viruses are the leading cause of recreational waterborne disease outbreaks , but detE. coli or enterococci) are present at 1\u20132 orders of magnitude higher concentrations (Current coliphage standard culture-based methods include single-agar layer (SAL) plaque assay , double-trations . A simplIn this study, we evaluate the performance of three somatic and F+ coliphage methods with 1 L sample volumes, including dead-end hollow fiber ultrafiltration  with SALSurface waters originated from Lake Michigan  (n = 37) and nearby Trail Creek (n = 37). Samples were collected during the 2015 Great Lakes beach season at a frequency of five samples per week. The sample collection procedures are detailed elsewhere . The D-HAll data were log10 transformed and expressed as plaque forming unit (PFU) per liter (L) for positive samples only as ND samples were not included in the concentration data. Statistical analyses were performed using SigmaPlot version 13.0 . One-way analysis of variance (ANOVA) followed by Holm-Sidak multiple comparisons or Kruskal-Wallis ANOVA on ranks (followed by Tukey tests) were applied to somatic and F+ coliphage datasets (from both sites), while Wilcoxon signed rank tests were used for overall comparisons between the coliphage types or sites.10 PFU/L) were comparable to M-SAL , but concentrations obtained by DMF (0.30 \u00b1 0.44 [SD] log10 PFU/L) (p < 0.001) lower than either D-HFUF-SAL or M-SAL methods. The DMF method did not yield any F+ coliphage results from Lake Michigan samples, but they were detected using the other two methods (10 PFU/L) (p < 0.001) for positive samples than either M-SAL or DMF methods, but there was no statistically significant difference in F+ coliphage concentrations obtained by the M-SAL and DMF methods (p = 0.899).Method performance metrics are summarized in 0 PFU/L)  were sig methods  with D-H0 PFU/L) . D-HFUF-10 PFU/L and 3.25 \u00b1 0.44 (SD) log10 PFU/L, respectively (p = 0.252) (p < 0.001) of somatic coliphage for positive samples were observed with the DMF method (2.75 \u00b1 0.49 [SD] log10 PFU/L) (p = 0.659) average concentrations of F+ coliphage in positive samples were found in Trail Creek waters using the D-HFUF-SAL method (1.25 \u00b1 0.68 [SD] log10 PFU/L) and M-SAL (1.22 \u00b1 0.85 [SD] log10 PFU/L), but concentrations obtained by the DMF method were significantly lower (p < 0.001) (p < 0.001), and concentrations of both coliphage types were typically higher in the Trail Creek samples compared to the Lake Michigan samples (p < 0.001).Somatic coliphage were consistently detected in Trail Creek water samples irrespective of the method . D-HFUF-= 0.252) . Signifi0 PFU/L) . Compara< 0.001) . OverallRegarding frequency of NDs, D-HFUF-SAL exhibited the lowest ND range overall, with 0%\u20132.7% (somatic) and 5.4%\u201335% (F+) . M-SAL r2, CaCl2, MgSO4, tryptone, glucose, Tween 80, Antifoam Y-30, sodium hexametaphosphate) but we\u2019ve excluded common laboratory disposables such as serological pipettes, pipette tips, gloves and similar. Regarding the processing time, we have assumed that sample is being processed by a single analyst familiar with the routine water quality assessment (e.g. membrane filtration and /or defined substrate technology for enumeration of E. coli and enterococci) and possessing basic knowledge of microbiological culture techniques. The cost per sample for D-HFUF-SAL and DMF was comparable, while M-SAL was approximately ten times higher, reflecting the increase in supplies and reagents needed to process 1 L sample volumes  to 25% (F+) . M-SAL mSupp Data"}
+{"text": "Correction to: BMC Public Health (2020) 20:1616https://doi.org/10.1186/s12889-020-09721-2It was highlighted that in the original article  the lettCorrect Table 1footnotesa Mann-Whitney testb \u03c72 testc Mortality rate per 1000 person-yearCOPD Chronic Obstructive Pulmonary Disease, CKD Chronic Kidney Disease, ICU Intensive Care Unit, IMSS Mexican Institute of Social Services, ISSSTE Institute of Social Security and Services for State Workers, SS Mexican Ministry of HealthCorrect equation in Results section2 = 31.83, df = 12, p < 0.001)(\u03c7The original article has been updated."}
+{"text": "O.decamerasp. nov. and O.joshiisp. nov., of the ant genus Ooceraea are described from India. These species differ from other known congeners on the basis of number of antennal segments. An illustrated key to the known species based on the worker caste is also provided.Two new species,  Ooceraea Roger, 1862 has been challenging, since its inception based on the type species O.fragosa. The taxonomic ambiguity has led to its uncertain placements in different subfamilies: in Myrmicinae  and no constrictions between abdominal segments IV, V and VI. Ooceraea can be distinguished from the closely allied Syscia Roger, 1861 on the basis of abdominal segment III relatively narrow in dorsal view and similar in size to the preceding abdominal segment II (petiole); in lateral view, abdominal tergite IV not folding over sternite and the anterior portion of the sternite visible; hind basitarsi not dilating distally, circular in cross-section and metabasitarsal glands absent  is probably native to the Asian continent, and has been introduced to Southeast Asia, the Pacific islands, Madagascar and the Caribbean islands  and Ooceraeabesucheti  with nine- and eleven-segmented antennae respectively ;SL Scape length: maximum length of antennal scape excluding basal condylar bulb;MW Mesosomal width: maximum width of promesonotum in dorsal view;ML Mesosomal length: maximum diagonal length of mesosoma in lateral view, measured from posterodorsal border of pronotal flange to posterior basal angle of metapleuron;PL Petiolar length: maximum length of petiole in lateral view;PH Petiolar height: maximum height of petiole in lateral view (including subpetiolar process);PW Petiolar width: maximum width of petiole in dorsal view;PPL Postpetiolar length: maximum length of postpetiole in lateral view;PPH Postpetiolar height: maximum height of postpetiole in lateral view;PPW Postpetiolar width: maximum width of postpetiole in dorsal view;CI Cephalic index: HW/HL \u00d7 100;SI Scape index: SL/HW \u00d7 100;PI1 Petiolar index 1: PL/PH \u00d7 100;PI2 Petiolar index 2: PW/PL \u00d7 100;PPI1 Postpetiolar index 1: PPL/PPH \u00d7 100;PPI2 Postpetiolar index 2: PPW/PPL \u00d7 100;WI Waist index: PPW/PW \u00d7 100.DepositoriesPUACPunjabi University Patiala Ant Collection at Department of Zoology and Environmental Sciences, Punjabi University, Patiala, Punjab, India;MCZCMuseum of Comparative Zoology, Harvard University, Cambridge, Massachusetts, United States.Taxon classificationAnimaliaHymenopteraFormicidae9DCFFE46-DF69-52AC-8558-AA33512A3640http://zoobank.org/182F8A89-653B-4604-8337-F7A5F258080B9.5627\u00b0N, 77.2348\u00b0E, 780 m.India, Kerala, Periyar Tiger Reserve Holotype worker and one paratype worker, both India, Kerala, Periyar Tiger Reserve 9.5627\u00b0N, 77.2348\u00b0E, 780 m, leaf litter, Winkler, 21 January 2017, Tarun Dhadwal leg. [PUAC].HL 0.57; HW 0.56; SL 0.34; MW 0.39; ML 0.68; PL 0.29; PH 0.34; PW 0.27; PPL 0.32; PPH 0.34; PPW 0.30; CI 98; SI, 61; PI1 85; PI2 93; PPI1 94; PPI2 88; WI 111. Paratype: HL 0.57; HW 0.56; SL 0.33; MW 0.39; ML 0.68; PL 0.29; PH 0.33; PW 0.26; PPL 0.32; PPH 0.34; PPW 0.30; CI 98; SI, 59; PI1 88; PI2 89; PPI1 94; PPI2 88; WI 111.Holotype: Head in full-face view, almost as long as broad, with lateral margin weakly convex and converging anteriorly, with posterior margin concave medially and posterior lateral corners rounded. Anterior clypeal margin reduced and slightly concave in the middle. Eyes present, small in size, with two ommatidia, parafrontal ridge prominently produced. Mandibles edentate, sub-triangular. Antenna 10-segmented; scape short and clavate, reaching almost mid-length of the head; apical funicular segment fusiform. Frontal lobes reduced. Antennal sockets fully exposed from above.Mesosoma in lateral view weakly convex; promesonotal suture and metanotal groove absent. Pronotum in dorsal view anteriorly marginate. Propodeum in dorsal view with posterior margin concave; propodeal declivity in lateral view slightly concave, with lateral margin slightly marginate; propodeal lobe reduced. Petiolar node in dorsal view as long as broad, rounded anteriorly, in lateral view hemiglobular; subpetiolar process well-developed, with sickle-shaped anteroventral apex. Postpetiole in dorsal view subtrapezoidal, with anterior margin transverse and posterior margin convex, in lateral view with anteroventral corner angulate. Gastral segment I  large, occupying the most part of gaster, in lateral view with dorsal margin weakly and roundly convex.Sculpture. Head foveolate-reticulate; mesosoma, petiole and postpetiole foveolate-reticulate; gaster foveolate, with foveae smaller than those of head and mesosoma.Pilosity and Pubescence. Body covered with erect or sub-erect hairs; sides of head and legs covered with shorter hairs; scape and funicular segments covered with short decumbent or subdecumbent hairs.Body coloration. Head and gaster light brown; mesosoma, petiole and postpetiole darker than the head; legs yellowish brown.Queen. Unknown.Male. Unknown.Ooceraeajoshii sp. nov. and O.decamera sp. nov. (described below) are distinctly separated from the other valid congeners by having 10-segmented antennae. Furthermore, the two new species are well distinguished from each other by a combination of the following characters: head shape ; presence of ommatidia (present in O.joshii sp. nov. and absent in O.decamera sp. nov.); propodeal lobes (reduced versus roundly produced); petiolar node in lateral view (hemiglobular versus rectangular); subpetiolar process ; pilosity (head and body comparatively more pilose in O.joshii sp. nov.); and sculpturation .The type series was found in leaf litter samples collected from the Medaganam region of the Periyar Tiger Reserve situated at an elevation of 780 meters. The region is composed of an undisturbed tropical moist evergreen forest with low light penetration, with a mean average daytime temperature of 30 \u00b0C.Known only from the type locality.The species has been named in honor of Professor Amitabh Joshi, a distinguished evolutionary biologist based at Jawaharlal Nehru Centre for Advanced Scientific Research (JNCASR), Bengaluru, India.Taxon classificationAnimaliaHymenopteraFormicidaeEFC710C7-7193-5101-BA40-0E0AD282BD0Chttp://zoobank.org/D8C9E609-7416-4081-A54A-83CAA01CEAD610.02308\u00b0N, 77.833333\u00b0E, 250\u2013350 m alt.India: Madras, Alagarkovil, 21 km. N Madurai, Holotype worker, India, Madras, Alagarkovil, 21 km N Madurai, 10.02308\u00b0N, 77.833333\u00b0E, 250\u2013350 m alt.; 2 November 1972; Besuchet Lobt Mussard leg. (Specimen number/barcode: MCZ-ENT00649398) [MCZC].HL 0.62; HW 0.46; SL 0.26; MW 0.38; ML 0.78; PL 0.26; PH 0.42; PW 0.30; PPL 0.34; PPH 0.41; PPW 0.40; CI 74; SI 57; PI1 62; PI2 93; PPI1 81; PPI2 118; WI 133.CI 74), with lateral sides weakly convex, with posterior margin concave medially, with occipital lobes/corners angulate. Anterior clypeal margin slightly projecting forward. Eyes absent. Parafrontal ridge prominent and elevated. Mandibles edentate but weakly serrate. Antennae with 10 segments; scape short, clavate, slightly surpassing the mid-length of head. Frontal lobes reduced. Antennal sockets fully exposed from above.Head in full-face view rectangular, distinctly longer than broad  large occupying the most part of gaster, in lateral view with dorsal margin almost straight, base of cinctus of first gastral tergite cross-ribbed.Sculpture. Head, mesosoma, petiole and postpetiole shallowly foveolate-reticulate; mandibles and dorsal surface of gaster sparsely foveolate, foveae somewhat smaller as compared to those present on head, mesosoma, petiole, and postpetiole.Pilosity and pubescence. Whole body covered with pale yellow erect and sub-erect hairs; appressed pubescence abundant on antennae and legs.Body coloration. Mandibles, antennae, legs, subpetiolar process and gaster light brown; head, mesosoma and gaster dark brown.Queen. Unknown.Male. Unknown.The two species significantly differ from each other on the basis of dimensions of head capsule and shape of subpetiolar process.Unknown.Known only from the type locality. The place has been transformed into agricultural land and is prone to anthropogenic activities. Thus, this reinforces the concept that important biodiversity components, which are already rare, are imperiled due to local extinctions.decamera refers to the ten-segmented antennal count.The species epithet"}
+{"text": "Plakobranchus cf. ocellatus (Heterobranchia: Sacoglossa), a so-called \u2018solar-powered\u2019 sea slug with long-term retention of chloroplasts. The mitochondrial genome was 14,177\u2009bp in length containing the standard set of 13 protein-coding genes, 2 rRNAs, and 22 tRNAs. The base composition of 27.3% A, 15.6% C, 18.6% G, and 38.5% T showed a strong A\u2009+\u2009T bias. The genome organization of P. cf. ocellatus is identical to the other sacoglossan mitogenomes sequenced so far, except for Ascobulla fragilis.We present the complete mitochondrial genome sequence of  Plakobranchus cf. ocellatus van Hasselt, 1824 belongs to a small group of marine, heterobranch slugs, called Sacoglossa, of which certain species have the ability to sequester chloroplasts from its food algae  in December 2012. The heads of about 60 slugs were dissected to remove the post-pharyngeal nerve ring, which were directly deep frozen in liquid nitrogen and then stored (\u221280\u2009\u00b0C). From these, genomic DNA was extracted using the QIAGEN DNeasy\u00ae Blood & Tissue Kit . Voucher material is stored in absolute ethanol at the Zoological Research Museum Alexander Koenig (voucher no. ZFMK-TIS-29490).Specimens of \u00ae DNA PCR-Free Library Preparation Kit  according to the manufacturer\u2019s protocol and 125\u2009bp paired-end reads were sequenced on an Illumina HiSeq 2500 platform . All read pairs were used for mitochondrial genome assembly with MITObim 1.8  as seed. After verification of correct circularity, genome annotation was done with MITOS revision 656  were prepared using the TruSeqElysia chlorotica, E. ornata, Thuridilla gracilis, and Placida sp. 1 NY-2013, except for Ascobulla fragilis.The complete mitochondrial genome (GenBank accession number: KX853083) had a length of 14,177\u2009bp encoding for 13 protein-coding genes, 2 rRNAs, and 22 tRNAs. The base composition is 27.3% A, 15.6% C, 18.6% G, and 38.5% T. A total of 183\u2009bp nucleotides were observed in multiple small intergenic regions, ranging from 1 to 54\u2009bp (found between the genes coding for COX3 and trnI(gat)). The gene order was as follows: trnK(ttt), cox1, trnV(tac), rrnL, trnL1(tag), trnA(tgc), trnP(tgg), nad6, nad5, nad1, trnW(tca), trnY(gta), nad4l, cob, trnD(gtc), trnF(gaa), cox2, trnG(tcc), trnH(gtg), -trnQ(ttg), -trnL2(taa), -atp8, -trnN(gtt), trnC(gca), -atp6, -trnR(tcg), -trnE(ttc), -rrnS, -trnM(cat), -nad3, -trnS2(tga), trnS1(gct), nad4, -trnT(tgt), -cox3, trnI(gat), nad2 and is therefore identical to the other sacoglossan mitogenomes sequenced so far, i.e. P. cf. ocellatus and all other sacoglossan species with fully sequenced mitogenome were extracted, aligned with MAFFT v7.271 (G-INS-i mode) (Katoh & Standley The amino acid sequence of the 13 protein-coding genes of"}
+{"text": "Chromatographic separation of androgen derivatives was optimized, carefully separating isobaric interferents and acceptable outputs for precision and trueness achieved following injection of 0.4 pg on column (approximately 34 pmol/L). HMP derivatives of all androgens tested could be detected in low plasma volumes: male (100 \u00b5L) and post-menopausal female (200 \u00b5L), and derivatives were stable over 30 days at -20\u00b0C. In conclusion, HMP derivatization, in conjunction with LC-MS/MS, is suitable for quantitative analysis of DHT, testosterone and androstenedione in low plasma volumes, offering advantages in sensitivity over current methodologies.Liquid Chromatography tandem mass spectrometry (LC-MS/MS) is the gold-standard approach for androgen analysis in biological fluids, superseding immunoassays in selectivity, particularly at low concentrations. While LC-MS/MS is established for analysis of testosterone and androstenedione, 5\u03b1-dihydrotestosterone (DHT) presents greater analytical challenges. DHT circulates at low nanomolar concentrations in men and lower in women, ionizing inefficiently and suffering from isobaric interference from other androgens. Even using current LC-MS/MS technology, large plasma volumes (>0.5 mL) are required for detection, undesirable clinically and unsuitable for animals. This study investigated derivatization approaches using hydrazine-based reagents to enhance ionization efficiency and sensitivity of analysis of DHT by LC-MS/MS. Derivatization of DHT using 2-hydrazino-1-methylpyridine (HMP) and 2-hydrazino-4-(trifluoromethyl)-pyrimidine (HTP) were compared. A method was validated using an UHPLC interfaced by electrospray with a triple quadruple mass spectrometer , analyzing human plasma  following solid-phase extraction. HMP derivatives were selected for validation affording greater sensitivity than those formed with HTP. HMP derivatives were detected by selected reaction monitoring (DHT-HMP  Twydrazone , specifiution MS .3.1.1.2m/z 449 , m/z 447 (androstenedione) and m/z 451 (DHT) (m/z 257 (C11H12N4F3) and m/z 269 (C12H12N4F3) for testosterone, androstenedione and EpiT resulting from the cleavage of the steroid A and B ring (m/z 288 (C19H30ON) and m/z 164 (C5H5N3F3) whereas, m/z 286 (C19H28ON) and m/z 164 (C5H5N3F3) were selected for DHA and DHEA respectively. Heterolytic cleavage of hydrazine N-N bond of derivative may form ions at m/z 288 and m/z 286, and m/z 164 characterizes the protonated trifluoromethyl-pyrimidine moiety. Ions obtained following collision induced dissociation of derivatives of stable-isotopically labeled testosterone, androstenedione and DHT corroborated potential fragmentation   14] andd B ring  14,46]. andd B r. andd B entation .3.1.23.1.2.1E and Z isomers which elute as double peaks for each androgen HMP derivatization forms two 18 columns with modified stationary phases, particle sizes and dimensions. SunFire\u00ae C18 , ACE\u00ae Excel PFP-C18 , ACE\u00ae UltraCore 2.5 SuperC18 , ACE\u00ae Excel super C18 , Acquity UPLC\u00ae BEH C18  and Acquity UPLC\u00ae BEH C18  were evaluated aiming to achieve baseline resolution of isomers and isotopologues within reasonable analytical times. Ultimately rapid chromatographic separation was achieved between isomeric HMP derivatives of each androgen and their endogenous isomers using a UPLC\u00ae BEH C18  column operated at 50\u00b0C, with run times of 10 min  and gradient method  and the estimated LOQs were 0.4, 0.8 and 0.4 pg  on column for DHT, testosterone and androstenedione respectively. Thus, estimated LOQs were improved ~21-fold and ~ 12.5-fold for HMP-DHT and HMP-androstenedione respectively in comparison to our LC-MS/MS method for underivatized androgens 3.23.2.113C3-T 97\u00b19%, 13C3-A4 99\u00b11% and 13C3-DHT 98\u00b15%). Extraction from plasma was assessed using stable-isotope labeled standards as surrogates, showing similar results . Ion suppression of internal standards was also within acceptable limits . Establishing conditions which limited ion suppression reduced the potential for variability in quantitation brought about by impact of subtle variations in matrix composition, particularly for low abundance analytes such as DHT. Minimal differences (<0.05 mins) in retention time between 13C3 IS and endogenous steroid due to the presence of heavy isotope were recorded. The assessment of the whole process also demonstrated that the presence of matrix did not adversely affected derivatization efficiency.Recovery of analytes by extraction was maximized and matrix effect minimized to ensure the cleanest possible sample to be prepared for analysis, and thus maximising column life-span and limiting source contamination. Analytes were well retained on lipophilic polymeric adsorbent Oasis\u00ae HLB ,50. and 3.2.213C3-T 1.7, 13C3-A4 1.8, and 13C3-DHT 2.7), were comparable to those in plasma, showing differences less than 20% .Interferents close to the retention times of the peaks of interest were not observed visually. The ratios of peak areas of quantifier to qualifier mass transitions ratios in standards , but acceptable within the reference range , intra and inter assay deemed acceptable (<20% RSD for precision and trueness at the LOQ  and <15%3.2.5The HMP derivatives demonstrated acceptable stability upon storage in an auto-sampler (10\u00b0C) over 24 h, with limited degradation measured for derivatized testosterone , androstenedione  and DHT  in extracts of plasma from male and post-menopausal female subjects, respectively. Derivatives were stable upon short-term storage in the freezer  for 7 days; reduction from original response testosterone , androstenedione  and DHT  in extracts of samples from males and testosterone , androstenedione  and DHT  from post-menopausal female. Moreover, derivatives were stable upon long-term storage in the freezer  for 30 days; reduction from original response testosterone , androstenedione  and DHT  in extracts of samples from males and testosterone , androstenedione  and DHT  from post-menopausal females.3.3The method was applied to samples from post-menopausal females and males and compared to typical reference ranges  1,33]. ,33. 1,334. While multi-dimensional detection in conjunction with chromatography brings anticipated benefits in selectivity In summary derivatization to form HMP derivatives of androgens, in conjunction with LC-MS/MS, is suitable for quantitative analysis of androgens in low abundance in biological fluids, offering advantages in sensitivity over analysis of underivatized DHT. The approach allows concomitant analysis of testosterone, androstenedione and DHT and this method may be extended to include further steroids with ketone functions e.g. 11-oxygenated androgens. Robust measurement was achieved across typical physiological ranges found in post-menopausal women and men offering an attractive alternative to immunoassays, which lack selectivity in the lower physiological range of concentrations and typically report only one androgenAbdullah MM Faqehi: Methodology, Formal analysis, Project administration, Resources, Validation, Writing - original draft. Scott G Denham: Methodology, Formal analysis. Gregorio Naredo: Methodology, Formal analysis, Writing - review & editing. Diego F Cobice: Methodology, Formal analysis, Writing - review & editing. Shazia Khan: Methodology, Formal analysis, Writing - review & editing. Joanna P Simpson: Methodology, Formal analysis, Writing - review & editing. Ghazali Sabil: Methodology, Formal analysis, Writing - review & editing. Rita Upreti: Investigation, Writing - review & editing. Fraser Gibb: Investigation, Writing - review & editing. Natalie ZM Homer: Methodology, Validation, Project administration, Supervision, Writing - review & editing. Ruth Andrew: Conceptualization, Methodology, Validation, Funding acquisition, Project administration, Supervision, Writing - review & editing.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper."}
+{"text": "Lactococcus lactis (L. lactis)/pNZ8150-phosphatidylglycerophosphate synthetase A (pgsA)-HAsd, in which pgsA was used as an anchor protein, and investigated the immunogenicity of HAsd in a mouse model by oral administration without the use of a mucosal adjuvant. Compared with L. lactis/pNZ8150-pgsA, mice were orally vaccinated with L. lactis/pNZ8150-pgsA-HAsd and then produced strong humoral and mucosal immune responses. Importantly, L. lactis/pNZ8150-pgsA-HAsd provided cross-protection against H5N1, H3N2 and H1N1 virus infections. Our data support the hypothesis that HAsd presented on the surface of L. lactis can provide cross-protective immunity against divergent influenza A viruses. Taken together, these findings suggest that L. lactis/pNZ8150-pgsA-HAsd can be considered an alternative approach to developing a novel universal vaccine during an influenza A pandemic.Most of the current approaches to influenza vaccine design focus on antibodies against influenza (HA). However, these influenza vaccines typically provide strain-specific protection against mostly homologous subtypes. There is an urgent need to develop a universal vaccine that confers cross-protection against influenza viruses. Of note, the HA stalk domain (HAsd) is a promising target for such an influenza vaccine. In this study, we generated recombinant Abbreviations: HA, HAsd, HA stalk domain; L. lactis, Lactococcus lactis; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; IFA, immunofluorescence assay; PBS, phosphate-buffered saline; pgsA, phosphatidylglycerophosphate synthetase A; SPF, specific pathogen-free; CFU, colony-forming unit; BSL-3, biosafety level-3 laboratory; TCID50, 50% tissue culture infective dose; ELISA, enzyme-linked immunosorbent assay; OD, optical density; LTB, liable enterotoxin B subunit; CTB, cholera toxin B subunit. Influenza A viruses cause a highly infectious respiratory disease that remains a public health problem worldwide . Humans Lactococcus lactis (L. lactis) has been engineered to express heterologous proteins [L. lactis expressing HA, HA1, neuraminidase or nucleoprotein of avian influenza H5N1 virus is a safe and effective vaccine candidate against H5N1 virus infection in mouse and chicken models with the use of a mucosal adjuvant or enteric capsule [L. lactis in the absence of a mucosal adjuvant can provide cross-protective immunity against divergent influenza A viruses.proteins . Our pre capsule . HoweverL. lactis, recombinant L. lactis/pNZ8150-pgsA-HAsd was constructed, in which pgsA derived from the chromosomal DNA of Bacillus subtilis [L. lactis/pNZ8150-pgsA-HAsd, which elicited robust humoral and mucosal immune responses against influenza A viruses. Most importantly, unadjuvanted L. lactis/pNZ8150-pgsA-HAsd could provide cross-protective immunity against lethal challenge with homologous and heterologous influenza A viruses. These data suggest that recombinant L. lactis/pNZ8150-pgsA-HAsd is expected to serve as an alternative approach for a truly protective universal influenza vaccine against divergent influenza A viruses.Thus, to investigate the immunogenicity of the HAsd presented on the surface of subtilis , and wasL. lactis was determined by western blot analysis. A highly specific band (approximately 100 kDa) was detected for the pgsA-HAsd fusion protein in the L. lactis/pNZ8150-pgsA-HAsd cells . In contrast, no band was detected in the L. lactis/pNZ8150-pgsA cells .The HAsd gene  of codon-optimized A/Vietnam/1203/2004(H5N1)  was clonsd cells , lane 3.L. lactis, L. lactis/pNZ8150-pgsA and L. lactis/pNZ8150-pgsA-HAsd cells were examined by immunofluorescence assay (IFA) and flow cytometric analysis. There were no positive signals in the L. lactis/pNZ8150-pgsA cells . By contrast, L. lactis/pNZ8150-pgsA-HAsd consistently exhibited specific fluorescent signals, indicating that HAsd was stably expressed on the surface of L. lactis (right side).To confirm that HAsd protein was expressed on the surface of L. lactis/pNZ8150-pgsA or L. lactis/pNZ8150-pgsA-HAsd group at the prime immunization. A highly significant increase (IgG titer \u2265 25) was observed in the L. lactis/pNZ8150-pgsA-HAsd group at the boost immunization, IgG titer induced by L. lactis/pNZ8150-pgsA-HAsd reach 28.4\u00b10.894, whereas there were still no significant (IgG titer <25) changes in the PBS or L. lactis/pNZ8150-pgsA group.HA-specific antibody responses were measured by ELISA. As shown in L. lactis/pNZ8150-pgsA or L. lactis/pNZ8150-pgsA-HAsd group at the prime immunization. Only L. lactis/pNZ8150-pgsA- HAsd could induce an elevated level of IgA antibodies after the boost immunization, IgA titers in the intestine and upper respiratory washes were 27.8\u00b11.30 and 27.4\u00b11.14, respectively.Mucosal IgA antibodies were also detected in the intestine and upper respiratory washes  and c. TL. lactis/pNZ8150-pgsA-HAsd orally after the prime-boost immunization could produce an elevated level of HA-specific IgG and IgA antibody responses.Collectively, these results demonstrated that mice administered L. lactis/pNZ8150-pgsA-HAsd, all vaccinated mice were challenged with 20\u00a0\u00b5L of 104 EID50 of A/Vietnam/1203/2004(H5N1), A/Beijing/47/1992(H3N2) or A/California/04/2009(H1N1) and monitored for 14\u00a0days. As shown in L. lactis/pNZ8150-pgsA showed significant body weight loss and increased virus shedding in the lung and died within 8\u00a0days after challenge with the lethal dose of virus . In contrast, mice vaccinated orally with L. lactis/pNZ8150-pgsA-HAsd experienced only mild weight loss and recovered by 14\u00a0days . Importantly, L. lactis/pNZ8150-pgsA-HAsd could confer 100% (5/5), 80% (4/5) and 80% (4/5) protection against A/Vietnam/1203/2004(H5N1), A/Beijing/47/1992(H3N2) and A/California/04/2009(H1N1), respectively .Last, to assess the cross-protective efficacy of of virus ,b and c. 14\u00a0days , b and c 14\u00a0days , E and Fectively ,\u00a0h and iNde I or EcoR I sites underlined . The resulting Nde I/EcoR I fragment was cloned into pNZ8150-pgsA at the C-terminal end, in which pgsA was used as an anchor protein [L. lactis NZ9000. The positive clone of L. lactis/pNZ8150-pgsA-HAsd was screened, and the plasmid was expressed as described previously [L. lactis/pNZ8150-pgsA was used as a negative control for subsequent analysis.The HAsd (876 bp) gene of A/Vietnam/1203/2004(H5N1) that included part of HA1 (from 831 bp to 1041 bp) and complete HA2 gene (from 1041 bp to 1707 bp)  was PCR  protein , and theeviously . L. lactL. lactis surface was determined by western blot analysis as described previously [6-cell L. lactis/pNZ8150-pgsA-HAsd pellets were mixed with 60\u00a0\u00b5L of 6 \u00d7\u00a0loading buffer, boiled for 10\u00a0minutes, subjected to sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a nitrocellulose membrane . The membrane was incubated with a 1:500-diluted monoclonal mouse anti-HA of A/Vietnam/1203/2004(H5N1) antibody  after blocking with 5% skim milk at room temperature for 2\u00a0h. After incubation overnight at 4\u00b0C, the membrane was incubated with affinity-purified horseradish peroxidase (HRP)-conjugated anti-mouse IgG . The membrane was reacted with West Pico Chemiluminescent Substrate  and imaged using a Molecular Imager ChemiDoc XRS System . Precision Plus Protein\u2122 WesternC\u2122  was used as a standard protein marker.The expression level of HAsd displayed on the recombinant eviously . BrieflyL. lactis surface was by an IFA . Briefly, 106L. lactis/pNZ8150-pgsA-HAsd cells were washed three times with sterile PBS, incubated with monoclonal mouse anti-HA antibody  at 4\u00b0C for 1\u00a0h, followed by FITC-conjugated goat anti-mouse IgG at 4\u00b0C for 30\u00a0minutes, and resuspended in 100\u00a0\u00b5L of sterile PBS. Finally, L. lactis/pNZ8150-pgsA-HAsd cells were used for the IFA. L. lactis/pNZ8150-pgsA cells were used as negative controls.The HAsd protein displayed on the L. lactis/pNZ8150-pgsA-HAsd was analyzed by flow cytometric analysis .Flow cytometry was performed very similarly to IFA, as described above. Five hundred microliters of Specific-pathogen-free (SPF) six-week-old female BALB/c mice were used  in this study and housed in cages ventilated under negative pressure with high efficiency particulate air filter.L. lactis/pNZ8150-pgsA-HAsd cells was adjusted to 1012 colony forming unit (CFU)/ml with sterile PBS. The mice (n\u00a0=\u00a039 per group) were vaccinated orally with 1012 CFU of L. lactis/pNZ8150-pgsA-HAsd at days 0, 1, 2, and 3 for prime immunization and boosted at day 17, 18, 19, and 20. PBS and L. lactis/pNZ8150-pgsA cells were used as controls.The concentration of recombinant At days 15 and 34 after the initial immunization, blood samples  and intestine and upper respiratory washes  were collected.4 EID50 of lethal dose of A/Vietnam/1203/2004(H5N1) (n\u00a0=\u00a08 per group), A/Beijing/47/1992(H3N2) (n\u00a0=\u00a08 per group)or A/California/04/2009(H1N1) virus (n\u00a0=\u00a08 per group). The mice were monitored for 14\u00a0days, and body weight loss and survival rate post challenge were calculated.Two weeks after the last immunization, all vaccinated mice (n\u00a0=\u00a024 per group) were transferred into an enhanced biosafety level-3 laboratory (BSL-3) containment facility and challenged intranasally with 20\u00a0\u00b5L containing 1050).Additionally, lungs from vaccinated mice were harvested for the detection of virus shedding at day 3 post infection (n\u00a0=\u00a03 per group). Tissue samples were homogenized and processed as described previously . The virAntibody responses of serum IgG and IgA in the intestinal washes and upper respiratory washes were determined by ELISA using recombinant HA protein (2\u00a0\u00b5g/mL) of A/Vietnam/1203/2004(H5N1)  as a coating antigen, as described previously . A 1:5,0p-value of less than 0.05 was considered statistically significant.A two-tailed Student\u2019s t-test and one-way ANOVA were used for all statistical analyses. A Due to continuous antigen gene occurring mutation, the influenza A virus is considered to have the potential to cause the next pandemic . Most ofL. lactis may provide cross-protective immunity against divergent influenza A viruses. Generally, the HAsd, including HA2 and part of HA1, was displayed on the surface of L. lactis, and L. lactis/pNZ8150-pgsA-HAsd showed a specific binding profile that could be directly with anti-HA antibody and FITC-conjugated goat anti-mouse IgG. It was then tested by IFA and flow cytometric analysis. The binding profiles observed positively correlated with the display efficacy of HAsd, demonstrating that HAsd presented on the surface of L. lactis has reactive activity in vitro, which may contribute to immunogenicity in vivo.It has been hypothesized that the HAsd present on the surface of E. coli heat-liable enterotoxin B subunit (LTB) and cholera toxin B subunit (CTB) are potent mucosal adjuvants and promote the induction of protective immunity. However, the adverse effects of these adjuvants in clinical trials, including diarrhea, low-grade fever, and vomiting, limit their further application [L. lactis has an adjuvant function because of its inherent safety profile [L. lactis/pNZ8150-pgsA-HAsd could induce an adequate humoral response and mucosal response in the absence of a mucosal adjuvant. Moreover, the contributions of HA-specific antibodies for broad-spectrum protection against divergent influenza A viruses were emphasized.It is generally recognized that lication ,24. Furt profile , and sti profile ,27. ThusL. lactis/pNZ8150-pgsA-HAsd were protected completely (100%) (5/5) from homologous A/Vietnam/1203/2004(H5N1) virus challenge, as well as 80% (4/5) against heterologous H3N2 or H1N1 virus infection, and showed a reduced degree of virus shedding in the lung. These findings provide reliable evidence that L. lactis/pNZ8150-pgsA-HAsd is an effective vaccine candidate to induce cross-protective immunity against divergent influenza A viruses.Antigen epitope is crucial in designing vaccine. The HAsd designed in this study was codon-optimized for completely matching the antigen epitope with homologous H5N1 and 60% HAsd protein sequence similarity with heterologous H3N2 or H1N1, which provides the basis of cross-protective immunity. Furthermore, virus challenge is regarded as the gold standard for assessing the efficacy of an influenza vaccine. Therefore, mice administered orally with L. lactis/pNZ8110-pgsA-HA1, in which pgsA has been used as an anchor protein, provided immune protection against homologous H5N1 virus in the mouse model [L. lactis surface. The HAsd represents a promising research target for a novel universal influenza vaccine. Notably, the HAsd presented on the surface of L. lactis has the potential to induce cross-protective immunity against divergent influenza A viruses. Thus, this study strongly demonstrates that the discovery of L. lactis/pNZ8150-pgsA-HAsd may allow for an alternative design of influenza vaccines that would afford broad coverage.It has been demonstrated previously that adjuvanted se model . In this"}
+{"text": "Marine sponges are well known as rich sources of biologically natural products. Growing evidence indicates that sponges harbor a wealth of microorganisms in their bodies, which are likely to be the true producers of bioactive secondary metabolites. In order to promote the study of natural product chemistry and explore the relationship between microorganisms and their sponge hosts, in this review, we give a comprehensive overview of the structures, sources, and activities of the 774 new marine natural products from sponge-derived microorganisms described over the last two decades from 1998 to 2017. The term \u201csymbiosis\u201d was first defined by the German mycologist Heinrich Anton de Bary in 1879 as \u201cthe living together of unlike organisms\u201d . Fr  [14]. F124 16)  were iso JBIR-59  and new   were is [18\u201319)  [18] werTrichoderma sp. derived from the Caribbean sponge Agelas dispar, four novel sorbicillinoid polyketide derivatives (20\u201323) (20\u201323 showed no bioactivities . A . A 60\u201361 (86\u201388) . Compoun4 \u00b5g/mL) .Arthrinium arundinis ZSDS1-F3 cultured from a Phakellia fusca marine sponge , four new cytochalasins, arthriniumnins A\u2013D (89\u201392), a new natural product, ketocytochalasin (93), three new 4-hydroxy-2-pyridone alkaloids, arthpyrones A\u2013C (94\u201396), and a new natural product, phenethyl 5-hydroxy-4-oxohexanoate (97)  inhibitory activity with an IC50 value of 0.81 \u03bcM ..107\u2013109)126\u2013127)  showed superior cytotoxicity at nanomolar concentrations toward 36 human tumor cell lines . Fr. Fr166) ue (177)  was puri  was yieAspergillus insuetus (OY-207) originated from the Mediterranean sponge Psammocinia sp. , and it afforded three novel meroterpenoids insuetolides A\u2013C (178\u2013180) and one new drimane sesquiterpene (181)  (Arthrinium sp. associated with the marine sponge Sarcotragus muscarum (Turkey) . Th. ThAsperne (192)  72]. A . A Asper2) (193) , was proA-MB-231 . Examina194\u2013201) . Their a202\u2013207), and a new polyketide, scedogiine G (208)   , were pr209\u2013210)  were isoe sponge ,76.211) and a monocyclic butoxyl derivative (212) , and the new phthalide herbaric acid (215) . However, their bioassays were disappointing . Th. ThDichone (262) . Compoun\u00b1 0.3 \u00b5M .263)  and ent-gloeosteretriol (266)  led to the isolation of two new natural products, evariquinone and isoemericellin (267\u2013268) (267 showed antiproliferative activity toward KB and NCI-H460 cells at a concentration of 3.16 \u03bcg/mL . F. FPetros364\u2013365) , were isth Korea . New \u03b1-p368\u2013369)  were iso Island) . Two new370\u2013371) , were deectively . A study F (373) . Both co 33.1 \u03bcM ,120. Two374\u2013375)  were isoectively . Futher ve (376) , which e\u201396.4 \u00b5M . A new d\u00b5M . A. AS)-2,43) (434)  was fromioassays . A new dys anthracene derivative called mayamycin (497), and three new streptophenazines I\u2013K (498\u2013500)  afforded two unusual compounds, Chlorohydroaspyrones A\u2013B (501\u2013502)  was found to produce two new indole derivatives (503\u2013504) (505\u2013508) . No bioactivities were detected . F. FStrept520\u2013521) . Lobophof 7.5 \u03bcM . A marinin 524) . Compounectively . From th9  172].  . Compou 526\u2013527  were ideectively . Chemicaid (531) , which wne (533)  were detn, China . Chemicane (534) . Compouncavenger .l-seryl-l-prolyl-l-glutamyl) and cyclo-(glycyl-l-prolyl-l-glutamyl) (535\u2013536)  was found to produce seven new drimane sesquiterpenoids (537\u2013543), five new ophiobolin-type sesterterpenoids (544\u2013548), and the two new pyrrolidine alkaloids (549\u2013550)   and exhibited cytotoxicity against the L929 (mouse fibroblast) cell line with an IC50 value approximated to 30 \u03bcM, while compound 553 was the most effective inhibitor of NF-\u03baB nuclear translocation with an IC50 value of 71 \u03bcM  (654) (Aspergillus ostianus strain TUF 01F313 isolated from a marine sponge collected at Pohnpei. Three new chlorinated compounds (647\u2013649) showed antibacterial activity against the Ruegeria atlantica strain. Compounds 650\u2013652 showed cytotoxic activity against mouse lymphocytic leukemia cells (L1210) with LD50 values of 2.1, 71.0, and 2.0 \u03bcg/mL, respectively  (654)  were isoectively ,219,220.655\u2013656) . Both co657\u2013658) . Both coectively .659)  (Aspergillus sp. (strain CNK-371) that was isolated from an unidentified sponge collected at Manele Bay, Hawaii. Compounds 660\u2013661 showed moderate cytotoxicity against human colon carcinoma (HCT-116) with IC50 values of 13.2, 10.9, and 13.9 \u03bcM, respectively . C. CAspergin (690) . Compoun25 \u03bcg/mL . One newmL [691)  was isol0.07) mm .Penicillium sp. MWZ14-4, which was isolated from an unidentified sponge (collected from the South China Sea), yielded 10 new fungal metabolites, including three hydroisocoumarins, penicimarins A\u2013C (692\u2013694), three isocoumarins, penicimarins D\u2013F (695\u2013697), and four benzofurans, penicifurans A\u2013D (698\u2013701) (698) showed moderate inhibitory activity against Staphylococcus albus with an MIC value of 3.13 \u03bcM and weak activity against B. cereus [702\u2013703) . Compounds 702\u2013703 may show antiphotoaging activity in UVB-irradiated models [Chemical examination of the sponge-derived fungus 698\u2013701) . Penicif. cereus . Two new702\u2013703) , were isd models .704) , amycocyclopiazonic acid (706), and amycolactam (707) (Amycolatopsis sp. isolated from a sponge sample gathered from Micronesia. Amycolactam (707) displayed significant to moderate cytotoxicity against the gastric cancer cell line SNU638, the colon cancer cell line HCT116, A546, K562, and SK-HEP1 with IC50 values of 0.8, 2.0, 13.7, 9.6, and 8.3 \u03bcM, respectively [Penicillium adametzioides AS-53 from an unidentified marine sponge  afforded a new spiroquinazoline derivative, N-Formyllapatin A (708), two new bisthiodiketopiperazine derivatives, adametizines A\u2013B (709\u2013710), two new acorane sesquiterpenes, adametacorenols A\u2013B (711\u2013712), a new dithiodiketopiperazine derivative, peniciadametizine A (713), and a highly oxygenated new analogue, peniciadametizine B (714)  with an LD50 value of 4.8 \u03bcM and moderate antimicrobial potency against several microbes with MIC values ranging from 8 to 32 \u03bcg/ mL, respectively, whereas compound 710 only showed weak activity against S. aureus . Compound 712 showed significant selective activity against the NCI-H446 cell line (IC50 = 5.0 \u03bcM). Compounds 713 and 714 showed inhibitory activity against the pathogenic fungus Alternaria brassicae with MIC values of 4.0 and 32.0 \u03bcg/mL, respectively [Aspergillus terreus MXH-23, isolated from an unidentified sponge , yielded a new butyrolactone derivative, namely butyrolactone VIII (715)  , was isoendently . Three nam (707)  were isoectively . Chemica B (714) . Compounectively ,245,246.II (715) . Derivatactivity .716) sesquiterpenoid and two new (717\u2013718) . Compounds 717\u2013718 exhibited significant inhibitory activity against cyclooxygenase (COX-2) with IC50 values of 10.6 and 8.9 \u03bcM, respectively. Besides, compound 718 displayed activities against intestinal virus EV71 with IC50 values of 30.1 \u03bcM [Micromonospora sp. NPS2077 isolated from an unidentified marine sponge  led to the isolation of a novel \u03b2-hydroxyl-\u03b4-lactone compound, neomacquarimicin (719) (Bacillus subtilis and Escherichia coli [720) . No bioactivities were detected [721)  [Verrucosispora sp. FIM06054 from a marine sponge sample (the East China Sea) afforded a new compound, FW054-1 (722)   xanthone 30.1 \u03bcM . The exain (719) , which ehia coli . One newli [720) , was isodetected . One newed [721)  was from, China) . Chemica-1 (722) . Compoun 6.88 \u03bcM .Aspergillus sp. OUCMDZ-1583 from an unidentified sponge XD10410  yielded 18 new compounds named aspergones A\u2013Q (723\u2013739) and 6-O-demethylmonocerin (740)  [Penicillium chrysogenum SYP-F-2720 from an unidentified sponge  afforded a novel benzoic acid (741)  afforded four new sesquiterpene lactones (742\u2013745) , a new cyclohexapeptide, similanamide (747), and a new pyripyropene derivative, named pyripyropene T (748) (747 exhibited weak activity against the MCF-7 (breast adenocarcinoma), NCI-H460 , and A373 (melanoma) cell lines, and neither of them showed antibacterial activity [Fractionation of the marine fungus in (740) . Compoun37.3 \u03bcM) . Chemicaid (741) . When adc effect . Study o742\u2013745) . Compoun193.3 \u03bcM . The inv T (748) . Only coactivity .Corynespora cassiicola XS-200900I7, isolated from an unidentified sponge XS-2009001 , yielded 12 new chromone derivatives, corynechromones A\u2013L (749\u2013760), two new naphthalenones, corynenones A\u2013B (761\u2013762), and one new depsidone, corynesidone E (763) (764\u2013765)  [766\u2013767)  led to the isolation of three new meroterpenoids, named austalides S\u2013U (768\u2013770)  . The bio764\u2013765) , were me20.0 \u03bcM) . Two new766\u2013767) , were meectively . Chemica768\u2013770) . Compounof 90 \u00b5M .Streptomyces albus PVA94-07 strain derived from an unidentified sponge afforded two new deferoxamine analogues, compounds 771\u2013772  afforded two new polyketides, aspergchromones A\u2013B (773\u2013774)  . Compounrculosis .As what we described above, temperate and tropical sea areas have become the main regions with sponge-derived microorganisms related to natural product chemistry. The statistical data shows that all the sponge hosts associated with microbes were distributed in the classes of Calcarea, Demospongiae, and unidentified sponges. The class Demospongiae sponges accounts for absolute majority at 77%, the class Calcarea and others sponges account for 1% and 22%, respectively. In the class Demospongiae, the order Haplosclerida provides the most sponge species accounting for 26.75% of the totality, followed by the orders Suberitida, Axinellida, Dictyoceratida, Tetractinellida, and Poecilosclerida etc. see .Aspergillus and Penicillium, followed by Trichoderma, Acremonium, Arthrinium, and Talaromyces. The genera Aspergillus and Penicillium obtained from many different sponge species take a percentage of 25% of the total microbes reported, the majority of which displayed wonderful chemistry diversity. Actinomycetes and bacteria account for 16% and 11% of sponge-derived microbes, respectively, which offered many novel and unique compounds. The actinomycetes studied were mainly derived from the genera Streptomyces, and the bacteria were mainly derived from the genera Pseudomonas, Pseudoalteromonas, and Bacillus  [50 value of 8.8 \u03bcM against H1N1 virus  [687) showed potent cell growth inhibitory activity against the murine P388 cell line with an ED50 value of 57 ng/mL  [Natural products isolated from sponge-derived microorganisms have interesting pharmaceutical activities such as cytotoxicity, antioxidant, antifungal, antiviral, and antibacterial activities. Some novel compounds showed significant cytotoxic activities in the nM levels, such as two new indolocarbazole alkaloids  and line14.8 \u03bcM) . Truncat46.5 \u03bcM) . Dankast8 ng/mL) . As we r8 ng/mL) . Another8 ng/mL) , antipla8 ng/mL) ,134, neu8 ng/mL) ,178, andA total of 774 new compounds from sponge-derived microorganisms, covering the last two decades from 1998 to 2017, were reviewed. These new compounds presented abundant chemical diversity except for the well-known types such as terpenes, alkaloids, peptides, aromatics, meroterpenoids, macrolides, polyketides, steroids, and so on.The total amount of new compounds from sponge-derived microorganisms has increased rapidly and has not yet reached a climax, especially in the last five years see . Among tWe should make more efforts to study the pharmacodynamic relationships and pharmacological effects of promising compounds, and conduct clinical trials to complete the drug-like evaluation of the compound. The study on the chemical constituents of sponges and their co-existing microorganisms will promote the study of the relationship between sponges and their co-existing microorganisms, develop and utilize medicinal resources, and systematically study the diversity of biodiversity and ecosystems. Given the structural differences and biodiversity of compounds derived from sponge symbiotic microorganisms, we believe that there are more resources waiting to be mined."}
+{"text": "Dear Editor,Several results have been reported on the efficacy of tranexamic acid (TXA) in patients with acute traumatic brain injury (TBI). As such, Lawati et al. (2020) conductRowell et al. (2020) conductIn contrast, an RCT report presented that the adjusted RRs (95% CIs) of TXA for mild-to-moderate and severe TBI death were 0.78 (0.64-0.95) and 0.99 (0.91-1.07), respectively , and a The author declares no conflict of interest."}
+{"text": "AbstractFanjingagen. nov., with type species F.digitata Yu & Yang, sp. nov., is described from the Guizhou Province of Southwest China. Habitus photos and illustrations of male genitalia of the new species are provided.A new leafhopper genus,  Alebroides group of Empoascini leafhoppers includes 25 genera and 156 species, widely distributed in the Palaearctic Region and Old World tropics . For morphological terminology we mostly follow Taxon classificationAnimaliaHemipteraCicadellidaeYu & Yanggen. nov.04EA95A4-B242-55E4-AD37-69DA953F1F0Fhttp://zoobank.org/F1EB5835-F39D-4C1A-862C-3F55C8618661Fanjingadigitata Yu & Yang, sp. nov., here designated.Alebroides group of Empoascini, the new genus runs to Nulliata Lu, Xu & Qin in the key to genera by Nulliata in having the abdominal apodemes long and from Nulliata and other genera in having a finger-like process bearing micro-setae on the lower posterior margin of the male pygofer.Within the Relatively robust Figs . Head slMale basal abdominal sternal apodemes well developed Fig. . Male pyThe genus name is an arbitrary combination of letters.China (Guizhou).Taxon classificationAnimaliaHemipteraCicadellidaeYu & Yangsp. nov.FE13424B-E8A8-5677-B624-2DA047EAD319http://zoobank.org/F427C9A4-AEFC-472A-AFAB-B158C408A80ESize. Male 3.94\u20134.00mm.Whole body yellowish, anterior margin of vertex marked with two brownish patches Figs . PronotuBasal sternal abdominal apodemes extending to end of segment VI Fig. . Male py\u2642, China, Guizhou, Fanjing Mountain, 21-August-2017, coll. Yarong Gao (GUGC). Paratypes: 3\u2642, same collecting data as holotype .The new species is named from the Latin \u201cdigitalis\u201d, meaning finger, for the finger-like process of the male pygofer.Euonymus (Celastraceae).China (Guizhou)."}
+{"text": "Correction to: Genome Biol (2019) 20:134https://doi.org/10.1186/s13059-019-1732-1Following publication of the original paper , the autIn the Additional file Additional file 1: Additional text 1. Supplementary methods. Additional text 2. Mapping SMURF-seq reads. Additional text 3. Short molecule sequencing with long-read sequencers. Additional table 3. Summary of sequencing runs. Figure S1. Distribution of length between restriction sites computed by measuring the distance between the recognition sites on the human reference genome. Figure S2. Schematic of SMURF-seq protocol. Figure S3. Sequencing of restriction enzyme digested normal diploid genome without SMURF-seq. Figure S4. Sequencing normal diploid genome using SMURF-seq. Figure S5. Sequencing normal diploid genome using SMURF-seq with 1D Rapid kit. Figure S6. Sequencing SK-BR-3 cancer genome using SMURF-seq. Figure S7. Replicate sequencing run of normal diploid genome using SMURF-seq. Figure S8. Replicate sequencing run of SK-BR-3 cancer genome using SMURF-seq. Figure S9. High-resolution CNV profile generated using SMURF-seq is highly concordant with the profile generated with Illumina WGS. Figure S10. SMURF-seq generates fragments at a faster rate than sequencing short molecules directly. Figure S11. CNV profile with reads obtained in first few minutes of sequencing. Figure S12. Multiplexed sequencing of normal diploid (barcode01) and SK-BR-3 cancer genome (barcode02) in a single sequencing run. Figure S13. Speed of nanopore sequencing as a function of read length. Figure S14. Biases correlated with GC content are reduced with LOWESS smoothing."}
+{"text": "This letter summarizes recommendations from the interdisciplinary working group of renal tumors (IAGN) of the German Cancer Society for the systemic treatment of advanced/metastatic renal cell carcinoma in the context of the current SARS-CoV-2 pandemic During the SARS-CoV-2 pandemic, oncologists faced several new challenges  of the German cancer society towards treatment of mRCC.During the process of treatment evaluation, the assessment of treatment indication must be made on an individual basis. However, once selecting treatment, patient, and tumor-specific parameters, patient\u00b4s comorbidity and the availability of local caregiving facilities should be taken into account. In our opinion, two major scenarios according to systemic treatment have to be considered: initiation or change medication of a systemic treatment upon progressing diseases and measures taken during ongoing systemic treatment.For systemic treatment, the appropriate treatment should be selected based on patient-specific factors that take the overall picture into account when searching for treatment , as well as a surprising high rate of SARS-CoV-2 diseases among cancer patients receiving surgery  benefit was gained by any of the immune combinations for patients with a favorable risk profile (HR for OS: ipilimumab/nivolumab vs. sunitinib: 1.19 (95% CI 0.77\u20131.85), axitinib/pembrolizumab vs. sunitinib: 0.94 (95% CI 0.43\u20132.07), axitinib/avelumab vs. sunitinib: 0.812 (95% CI 0.336\u20131.960) ; axitinib/pembrolizumab intermediate prognosis: 0.53 (95% CI 0.35\u20130.82), poor prognosis: 0.43 (95% CI 0.23\u20130.81); axitinib/avelumab: intermediate prognosis: 0.86 (95% CI 0.615\u20131.202), poor prognosis: 0.57 (95% CI 0.363\u20130.895); Cabozantinib vs. sunitinib: 0.80 (0.53\u20131.21)] (Motzer et al. Although precise guidelines according to mRCC treatment reflects best efficacy and QoL data, within the SARS-CoV-2 pandemic a patient-centered treatment choice, which is adapted to the local pandemic situation is warranted (s. Table"}
+{"text": "Dear Editor,9/L in the platelet count for mortality was 0.60 (0.43-0.84). They monitored changes in the platelet count during treatments and whether maintaining a normal platelet count was related to good prognosis. A meta-analysis conducted by Lippi et al.  have presented with thrombocytopenia. Figliozzi et al. (20209 have pr) reporte 0.60 0.4-0.84. ThHowever, some reports present an inverse association. Iba\u00f1ez et al. (2020) describAlthough two meta-analyses presented that thrombocytopenia reflected poor prognosis in patients with COVID-19 , the poThe author declares no conflict of interest."}
+{"text": "Microcerotermes eugnathus Silvestri  showed structural damage in Bir al-Shaghala cemeteries located in the oasis of Dakhla, Egypt. The mud tubes of this termite spread inside and over the mural painted floors of the tombs. Extracts from Lavandula latifolia, Origanum vulgare, and Syzygium aromaticum were tested for their anti-termitic activity and compared with the bio-insecticide, Bacillus thuringiensis var. kurstaki (Protecto 9.4% WP) and Dursban (Chlorpyrifos 48%). The bioassay experimental showed that the extracts have low activity against M. eugnathus compared to Protecto and Dursban, but the extract from O. vulgare showed promising natural termiticides.The termite Lavandula latifolia (Spike lavender), Origanum vulgare (Marjorum), and Syzygium aromaticum (Clove) against the termite Microcerotermes eugnathus. Plant extract results were compared to two commercially used termite pesticides, the bio-insecticide, Bacillus thuringiensis var. kurstaki (Protecto 9.4% WP) and Dursban (Chlorpyrifos 48%). Gas chromatography\u2013mass spectrometry (GC-MS) analysis was used to identify the main compounds in the three plant extracts. The main compounds in Lavandula Latifolia were linalool (21.49%), lavandulol (12.77%), \u03b2-terpinyl acetate (10.49%), and camphor (9.30%). Origanum vulgare extract contained thymol (14.64%), m-cymene (10.63%), linalool (6.75%), and terpinen-4-ol (6.92%) as main compounds. Syzygium aromaticum contained eugenol (99.16%) as the most abundant identified compound. The extract of O. vulgare caused the highest termite death rate, with an LC50 of 770.67 mg/L. Exposure to lavender extract showed a high death rate with an LC50 of 1086.39 mg/L. Clove extract did not show significant insecticidal activity with an LC50 > 2000 mg/L. Significant termiticide effects were found, with LC50 values of 84.09 and 269.98 mg/L for soldiers and workers under the application of Dursban and Protecto, respectively. The LC50 values reported for nymphs were <120, <164.5, and 627.87 mg/L after exposure to Dursban, Protecto, and O. vulgare extract, respectively. The results of the study show that some of the extracts have low toxicity compared to the bioagent and Dursban, and may show promise as natural termiticides, particularly as extracts from O. vulgare.A trend towards environmentally friendly chemicals for use in termite management has been occurring globally. This study examined three naturally occurring plant extracts from  Dakhla Oasis, Egypt, are the site of heavy infestation by the termite Microcerotermes eugnathus Silvestri  [Termites, although important in ecosystems, are economically important structural pests to wood and cellulosic material in service ,3,4,5,6.mitidae) ,8,9,10. M. eugnathus [In Egypt, this termite was reported along the northwestern coast  and in tugnathus .Treatments used to control this termite include bioagents, chemical pesticides, and natural products. Natural products investigated for termite management include plant extracts. In many parts of the world, chemical pesticides are considered poor for the environment and new research is examining the use of naturally occurring, less environmentally toxic, compounds ,12,13.Bacillus thuringiensis var. kurstaki and Stenotrophomonas maltophilia Palleroni & Bradbury have potential application as biological control agents of termites [B. thuringiensis is an example of this [S. maltophilia has shown potential application as a biocontrol agent and could degrade chitin in the termites, Coptotermes heimi (Wasmann) (Blattodea: Rhinotermitidae) and Heterotermes indicola (Wasmann) (Blattodea: Rhinotermitidae) [Bioagents such as termites ,15,16. T of this ,18. ChitDursban (Chlorpyrifos 48%) is an organophosphate pesticide that acts on the insect nervous system by inhibiting acetylcholinesterase (AChE) ,20,21. DC. intermedius Silvestri (Blattodea: Rhinotermitidae) died within 30, 40, and 110 min when exposed to 70% ethanol, aqueous, and acetone extracts from seeds of Parkia biglobosa (Jacq) Benth, respectively, at the concentration of 4 gm/L [Khaya senegalensis (Desr.) A. Juss., K. ivorensis A. Chev. and Swietenia mahagoni (L.) Jacq. showed potential insecticidal efficacy towards the dry wood termite Kalotermes flavicollis   [Larix leptolepis (Lamb.) Carr. Wood, with its large quantities of flavonoids, showed strong feeding deterrent activity against the subterranean termite Coptotermes formosanus Shiraki (Insecta: Blattodea: Rhinotermitidae) [Jatropha curcas L. oil showed maximum wood protection against Microcerotermes beesoni Snyder (Termitidae: Amitermitinae) at a concentration of 20% [Natural products are being studied as potential termiticides. The termite f 4 gm/L . Seed oimitidae) . Water emitidae) . Jatrophn of 20% .Tectona grandis L.f., Dalbergia sissoo Roxb., Cedrus deodara (Roxb.) G.Don and Pinus roxburghii Sarg. rapidly lowered numbers of protozoans in the hindgut of Reticulitermes flavipes (Kollar) (Insecta: Blattodea: Rhinotermitidae) and H. indicola workers, which were closely correlated with the mortality of worker [Morus alba L. heartwood resulted in 100% mortality in R. flavipes after feeding [Microcerotermes beesoni Snyder (Termitidae: Amitermitinae) [D. sissoo showed termite mortality at 10 mg/mL against H. indicola and R. flavipes with LC50s at 5.54 and 3.89 mg/mL, respectively [H. indicola mortality reached more than 75% after feeding on Populus deltoides W.Bartram ex Marshall wood treated with extract from M. alba [Some heartwoods show resistance to termite attack, which is attributed to the presence of toxic compounds within the heartwood ,30. For  feeding . Methanomitinae) . Heartwoectively . H. indi M. alba . Cymbopogon citratus (DC.) Stapf, Eucalyptus globulus Labill., Syzygium aromaticum (L.) Merr. and L. M. Perry, Origanum vulgare L., Rosmarinus officinalis L., Cinnamomum verum J.Presl, and Thymus vulgaris L. showed strong termiticidal activity against Odontotermes assamensis Holmgren (Blattodea: Termitidae) [Cananga odorata (Lam.) Hook.f. and Thomson flower extract of 2 mg/filter paper showed cumulative mortalities of 18% and 94%, after 2 and 7 days of exposure, respectively, against the termite Reticulitermes speratus (Kolbe) (Rhinotermitidae: Blattodea) [K. flavicollis pseudergates fed Casuarina sp. wood wafers, previously treated separately with Taxodium distichum (L.) Rich., E. citriodora Hook. and Cupressus sempervirens L. extracts, showed reduced numbers and vigour in spirochaete and flagellate populations [Tagetes erecta L. leaf extract, caused 100% mortality in Odontotermes obesus (Rambur) (Blattodea: Termitidae) after 24 h of exposure [Microcerotermes eugnathus.Many plants contain bioactive compounds such as phenols, terpenoids , aldehydes, ketones, and other compounds that have a strong effect against microorganisms and termites ,39,40,41mitidae) . Insectiattodea) . K. flavulations . At a doexposure . TherefoLavandula latifolia Medik.) belongs to the family Lamiaceae and is native to the Mediterranean. It is also cultivated in different regions of the world. The bioactivity of L. angustifolia Mill. and L. latifolia extracts from Spain were found to be associated with the presence of linalool, camphor, p-cymene, and limonene [\u03b1-pinene, thujene, \u03b2-pinene, sabinene, myrcene, p-cymene, limonene, camphene, camphor, 1,8-cineole, (Z)- and (E) ocimene, terpinene, terpinene-4-ol, lavandulol, lavandulylacetate, \u03b2-caryophyllene, and (Z)- and (E) farnesene [Lavender  species are characterized by the presence of two main chemotypes, thymol and carvacrol, while others have \u03b3-terpinene, p-cymene, cis-sabinene hydrate, terpinen-4-ol, \u03b1-terpineol, \u03b1-terpinene, \u03b3-terpinene, sabinene, and linalool as their main compounds [O. vulgare L. ssp. vulgare extract showed the presence of caryophyllene, spathulenol, germacrene-D, and \u03b1-terpineol and had antimicrobial properties [\u03b3-terpinene, \u03b1-terpineol, thymol, carvacrol, cis-sabinene-hydrate, linalool, p-cimene, 1,8-cineole were identified as the main compounds in the extract of O. vulgare L. [O. vulgare extract and its major compounds carvacrol, p-cymene, and \u03b3-terpinene showed potent larvicidal activity against the cotton bollworm, Helicoverpa armigera (H\u00fcbner) (Lepidoptera: Noctuidae: Heliothinae) [Several ompounds ,50,51,52operties . Terpinelgare L. . O. vulgothinae) . Syzygium aromaticum (L.) Merr. and L.M.Perry, Myrtaceae family) bud extract has shown the presence of eugenol, eugenol acetate and \u03b2-caryophyllene as main compounds with percentages of 88.61, 8.89 and 1.89%, respectively. It exhibited strong contact toxicity against Cacopsylla chinensis (Yang and Li) (Hemiptera: Psyllidae) with LD50 values of 0.730 \u00b5g/adult and 1.795 \u00b5g/nymph, respectively [S. aromaticum extract gave LC50 values of 13.871 and 15.551 \u03bcL against larvae and adult Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) 48 h after fumigation [50 values of 9.45 and 10.15 \u03bcL/g, against Acanthoscelides obtectus (Say) (Coleoptera: Chrysomelidae: Bruchinae) and Sitophilus zeamais Motschulsky (Coleoptera: Curculionidae), respectively [Clove  and the chemical compound Dursban (chlorpyrifos 48%). In addition, the chemical composition of the three extracts were analyzed with using GC-MS apparatus.This research aimed to (1) determine the chemical composition of three plant extracts  with potential anti-termitic activities; (2) evaluate the anti-termitic activity of the three plant extracts against Microcerotermes eugnathus foraging tubes. Termite tunnels could be observed on the floors and tomb substructure, causing cracks and separation of mortar. This termite species produces winged alates during the months of April and January, the remnants of which (wings) could often be observed around the tombs [Termite infestation in the cemeteries of Bir al-Shaghala located at the oases of Dakhla, Egypt was noted by the presence of he tombs . Figure Lavandula latifolia Medik., Lamiaceae) dry flower, Marjoram  dry leaves and Clove (Syzygium aromaticum (L.) Merrill and Perry, Mytaceae) dry buds were selected for their potential effect on termites. The extraction process was carried out at the National Research Center in Cairo, Egypt. Plant material was shade air-dried in the laboratory at room temperature for one week, then ground into a fine powder in an electrical blender. About 100 g of the fine powdered material was put into a 2-L flask containing 1000 mL of distilled water (DW) and extracted by the hydrodistillation method using a Clevenger-type apparatus for 3 h [Three plant extracts from Spike Lavender  with a TG\u20135MS direct capillary column (30 m \u00d7 0.25 mm \u00d7 0.25 \u00b5m film thickness). Extracts were diluted in n-hexane solvent in the ratio of 3:1 (3 n-heaxane: 1 extract sample) before being injected to the GC-MS. The column oven temperature was initially held at 50 \u00b0C and then increased by 5 \u00b0C/min to 250 \u00b0C and held for 2 min, then increased to a final temperature of 310 \u00b0C by 25 \u00b0C/min and held for 2 min. The injector and MS transfer line temperatures were kept at 270 and 260 \u00b0C, respectively; helium was used as a carrier gas at a constant flow rate of 1 mL/min. The solvent delay was 3 min, and diluted samples of 2 \u00b5L were injected automatically using an Autosampler AS1310 coupled with GC in the splitless mode. EI mass spectra were collected at 70 eV ionization voltages over the m/z range of 50\u2013650 in full scan mode. The ion source temperature was set at 250 \u00b0C. The components were identified by comparison of their retention times and mass spectra with those in the WILEY 09 and NIST 11 mass spectral databases [The chemical composition of extracts from atabases . The matatabases ,40,62,63Bacillus thuringiensis var. kurstaki\u201d (Protecto 9.4% WP) was diluted in distilled water to obtain the following concentrations: 164.5, 258.5, 376, 756, and 1410 mg/L. Dursban (Chlorpyrifos 48%) was diluted in distilled water and prepared in concentrations of 120, 240, 360, 480, and 960 mg/L.Extract concentrations for laboratory testing were prepared by mixing with 0.01% of Tween 80 and diluting with distilled water to obtain the following concentrations: 125, 250, 500, 1000, and 2000 mg/L. The bioagent \u201cMicrocerotermes eugnathus castes were collected from soil around cemeteries and kept in a container with a portion of termite nesting material. Briefly, from the experimental location, termites were placed inside a special box (4-L in capacity) for preservation and the box was covered with wool textile to maintain the temperature, because the temperatures in the interior oases are as high as 50 \u00b0C. During the transfer to the laboratory in Cairo, the death of the termites occurs due to the different environmental conditions, so it was necessary to warm the termites. Termites\u2019 death was also observed during transportation to the laboratory or even in the laboratory during the preparation for the laboratory experiment. Therefore, 10% glucose sugar solution [solution ,65 was pA no-choice bioassay method ,67 was e50) expressed as mg/L was calculated from log-concentration mortality regression lines [Untreated control replicates (five replicates) contained 10 workers, five soldiers and five nymphs, who were supplied with pieces of untreated cardboard paper only. Each concentration tested was replicated five times. All the treatments were kept in darkness at 48 \u00b1 3 \u00b0C. After seven days, the test was halted, the number of surviving termites was counted, and percent mortality was calculated. The lethal concentration , lavandulol (12.77%), \u03b2-terpinyl acetate (10.49%), camphor (9.30%), linalyl acetate (7.87%), (\u2212)-\u03b2-fenchol (5.12%), isobornyl acetate (5.00%), \u03b1-pinene (3.81%), terpineol (3.64%), and L-\u03b1-terpineol (2.62%).O. vulgare dry leaves. The main components were thymol (14.64%), m-cymene (10.63%), terpinen-4-ol (6.92%), linalool (6.75%), estragole (5.42%), \u03b3-terpinene (5.39%), anethole (5.00%), eucalyptol (3.61%), borneol (3.29%), \u03b2-caryophyllene (3.29%), \u03b1-terpinene (2.89%), carvacrol (2.58%), camphor (2.36%) and carvacryl methyl ether (2.34%). S. aromaticum extract. The main component was eugenol (99.16%).Microcerotermes eugnathus. After a seven-day exposure to this extract, the LC50 was 770.67 mg/L against soldiers and workers and 627.87 mg/L against nymphs. Lavender extract had less of toxic effect against M. eugnathus than marjoram, with an LC50 of 1086.39 mg/L for the group of soldiers and workers and 1140.74 mg/L for nymphs. Clove extract showed the lowest insecticidal activity against M. eugnathus for both nymphs and soldier/workers, with an LC50 > 2000 mg/L  extract from Macedonia [p-cymen-8-ol and bornyl acetate with values of 32.3%, 12.4%, 11.7%, 8.7%, 7.7%, and 4.2%, respectively, were the main compounds in L. latifolia extract from the Northern Tunisia [L. latifolia extracts from Spain, characterized by 1,8-cineole with high amounts (36.3\u201333.65%) [L. latifolia from Iran showed the presence of linalool (31.9\u201330.6%) as the main compound [Lavandula, 1,8-cineole, 2-\u03b2-pinene, \u03b1-thujone, camphor and cis-\u03b1-bisabolene were the main compounds isolated from L. angustifolia extract [Linalool, borneol and terpinene-4-ol, 1,8-cineole, camphor, and linalyl acetate were reported in lavandin  ,82, whilcompound . In othe extract .\u03b1-terpinene, \u03b3-terpinene, cis-sabinene-hydrate, linalool, terpinen-4-ol, terpineol, thymol, and carvacrol as the main compounds in O. vulgare extract with percentages of 5.58, 12.32, 3.35, 3.47, 21.43, 3.17, 9.45, and 11.67%, respectively [O. vulgare extract contained p-cymene, linalool, thymol, caryophyllene, and carvacrol. These authors showed insecticidal activity against Schistocerca gregaria (Forsk\u00e5l) (Orthoptera: Acrididae) [O. vulgare extract. Other studies reported this same compound at 9.45% [Coleus amboinicus Lour. extract caused 100% mortality to the termite Odontotermes obesus at a dose of 2.5 \u00d7 10\u22122 mg/cm3. This study found that thymol was the main compound, with 94.3% [Pittosporum undulatum Vent., Lippia sidoides Cham. and Lippia gracilis Schauer showed 100% mortality to the termite Heterotermes sulcatus Mathews (Blattodea: Rhinotermitidae) after 14 days of exposure, where the activity was related to the major compounds, limonene (80.8%), carvacrol (45.6%) and thymol (78.55%), that were identified in each extract, respectively [Previous work identified ectively . Other wrididae) . In the at 9.45% , 44.55% at 9.45% , 26.75% at 9.45% , and 58.at 9.45% . After 5th 94.3% . Extractectively .O. vulgare extract [O. vulgare extract with its main compounds of thymol (40.35%), p-cymene (17.32%), \u03b3-terpinene (15.66%), and carvacrol (12.15%), observed 72% and 88% mortality at 48 and 72 h, respectively, against Sitophilus oryzae (L.) (Coleoptera: Curculionidae) [O. vulgare extract showed toxicity in higher doses against Plutella xylustella L.  [O. vulgare extract at 40% exhibited 100% repellency against Cimex lectularius Linnaeus (Hemiptera: Cimicidae) [Terpineol (22.85%), \u03b1\u2013terpinene (20.60%), caryophyllene (6.75%) and thymol (4.53%) were the main compounds isolated from  extract . O. vulgionidae) . O. vulgralidae) . After amicidae) .Origanum were shown to have insecticidal activity against larvae of Culex pipiens L. (Diptera: Culicidae) [Lippia sidoides contained 44.55% thymol and showed high insecticidal activity against the termite Nasutitermes corniger (Blattodea: Termitidae) with 48 h exposure at a dose of 0.27 \u03bcg/mg [Oxygenated monoterpenes such as thymol, eucalyptol, linalool and carvacrol found in the extracts from the present study are reported to be more toxic against termite workers . For exalicidae) . The ext27 \u03bcg/mg .Microcerotermes eugnathus with an LC50 value of >2000 mg/L. Eugenol has been shown to be an effective fumigant and feeding deterrent against the termite C. formosanus [\u03b3-aminobutyric acid, causing inhibitory symptoms in the nervous system [Odontotermes assamensis Holmgren (Blattodea: Termitidae) at 2.5 mg/g after 8 days [Reticulitermes messperatus Kolbe (Blattodea: Rhinotermitidae) nymphs [S. aromaticum extract at 0.5 mL/L after 10 min of exposure showed 100% mortality against Coptotermes formosanus Shiraki (Insecta: Blattodea: Rhinotermitidae) [R. speratus [In this study, clove extract showed the lowest activity against rmosanus . Carvacrrmosanus . It causs system . Carvacrr 8 days . At 1.5 ) nymphs ,97. S. amitidae) . Clove bsperatus .M. eugnathus against termiticides than those of other termites. In addition, our results suggest that O. vulgare extract caused moderate activity against the termite M. eugnathus compared to commercial termiticides tested. Further research may show that O. vulgare extract or its derivatives could potentially be used to control and manage some termite infestations and lessen or limit the amount of more toxic pesticides currently in use. Our present results show a higher resistance of M. eugnathus. However, due to the highly toxic nature of Dursban and the fact that it has been withdrawn for use in some countries, the search for less toxic and natural termiticides from plants is of interest. Extract from the plant Origanum vulgare showed some promise as a plant-based toxicant for this termite. Chemical pesticides had the greatest impact on the mortality of the termite"}
+{"text": "Bacillus sp. strains EKM417B and EKM420B (from Citrullus lanata [watermelon]) and EKM501B (from Cucurbita moschata [butternut squash]) and Paenibacillus sp. strain EKM301P (from Cucurbita pepo L. var. pepo L. [pumpkin]). These strains previously demonstrated biostimulant and biocontrol activities.Here, we announce the draft genome sequences of four endophytic bacilli isolated from surface-sterilized seeds of three cucurbit species,  Bacillus sp. strains EKM417B and EKM420B (from Citrullus lanata [watermelon]) and EKM501B (from Cucurbita moschata [butternut squash]) and Paenibacillus sp. strain EKM301P (from Cucurbita pepo L. var. pepo L. [pumpkin]). These strains previously demonstrated biostimulant and biocontrol activities.Here, we announce the draft genome sequences of four endophytic bacilli isolated from surface-sterilized seeds of three cucurbit species,  Bacillus/Paenibacillus genera) (Bacillus sp. strains EKM417B and EKM420B (from Citrullus lanata [watermelon]), Bacillus sp. strain EKM501B (from Cucurbita moschata [butternut squash]), and Paenibacillus sp. strain EKM301P (Cucurbita pepo L. var. pepo L. [pumpkin]) using 16S rRNA gene primer pair 799F and 1492R and then submitted to GenBank   in planta; EKM501B grew on N2-free medium, produced indole-3-acetic acid (IAA/auxin) and ribonucleases, and suppressed Rhizoctonia solani; and EKM301P secreted cellulase and antagonized Fusarium graminearum and Rhizoctonia solani (Plant microbiomes have evolved to perform defensive/growth-promoting functions \u20133. 16S I\u2013 genera) , consist genera) . We isolde novo assembled using the EvoCAT pipeline (Evogene Clustering and Assembly Toolbox) and identified using KmerFinder 3.2 (Bacillus velezensis strain KD1 (GenBank accession number NZ_CP014990.2) (EKM417B and EKM420B), Bacillus cereus strain FORC087 (NZ_CP029454.1) (EKM501B), and Paenibacillus polymyxa strain SQR-21 (NZ_CP006872.1) . Genomic DNA was extracted using DNeasy UltraClean microbial kits  and adjusted to 50\u2009ng/\u03bcl. Libraries were constructed using TruSeq DNA Nano library prep kits  and then sequenced using the Illumina NovaSeq 6000 system to produce 1,461,384 (EKM417B), 1,659,509 (EKM420B), 1,823,824 (EKM501B), and 2,124,086 (EKM301P) raw reads in 150-bp paired-end format. Quality-trimmed reads  were 06872.1) . The pro06872.1) . Peptide06872.1) . DefaultGenome mining identified candidate genes involved in biofertilizer/biocontrol metabolic pathways, including those discussed above. These genes encode proteins involved in nitrogen fixation, phytase, alkaline phosphatase, carbon-nitrogen hydrolase, trehalose-6-phophate hydrolase, and tryptophan synthase (IAA/auxin production) \u201314. Bioc\u201314\u2013This whole-genome shotgun project and the Illumina raw reads have been deposited in DDBJ/EMBL/GenBank and the SRA, respectively, under the accession numbers provided in"}
+{"text": "Y.Tsiang 7712) when he described Tsugalongibracteata W.C.Cheng. Later, researchers suggested that the type is either in NAS or in PE. However, we found more than one duplicate of the type collection in both NAS and PE. Following the Shenzhen Code, we lectotypify the name T.longibracteata with Y.Tsiang 7712 (PE00003223) that bears a handwritten identification of W.C.Cheng.W.C.Cheng did not clearly indicate the herbarium repository of the type specimen ( Tsuga (Endl.) Carri\u00e8re: Tsugalongibracteata W.C.Cheng. This species differs from all known species of Tsuga in both vegetative and reproductive characters , two in NAS (NAS00070064 and NAS00070063), two in HUH (A00052508 and A00052510), one each in E (E00215871), IBSC (IBSC0012857), K (K000288277), NY (NY00001279), and S (S-C-4796) respectively. Shenzhen Code .1A55C046-BD56-5079-A68C-4A134EA9F396Nothotsugalongibracteata (W.C.Cheng) Hu ex C.N.Page, Notes Roy. Bot. Gard. Edinburgh 45(2): 390 .  \u2261Y.Tsiang (\u848b\u82f1) 7712 .China. Guizhou (\u8d35\u5dde): Yinjiang Tujiazu Miaozu Zizhixian , Fanjing Shan , in densely shaded ravine, alt. 400\u2013500 m, 19 December 1930,"}
+{"text": "Correction to: Arthritis Res Ther 22, 98 (2020)https://doi.org/10.1186/s13075-020-02185-0Following publication of the original article , the aut1. In Fig.\u00a02. Supplementary Figure\u00a0The original article has been corrected.Additional file 1:Figure S1. Distribution of FOI PVM enhancement in joints with increasing degree of osteoarthritis."}
+{"text": "Food and Drug Administration in 2019 for the treatment of community-acquired bacterial pneumonia (CABP). In this study we evaluated the The test strains were isolated from patients across China during the period from 2017 to 2019, including 634 strains of respiratory pathogens. The minimum inhibitory concentrations (MICs) of lefamulin and comparators were determined by broth microdilution method.Streptococcus pneumoniae and Staphylococcus evidenced by 100% inhibition at 0.25 mg/L, and favorable MIC50/90 (0.125/0.125 mg/L) against S. pneumoniae (penicillin MIC \u2265 2 mg/L), MIC50/90 (\u22640.015/0.125 mg/L) against methicillin-resistant S. aureus, and MIC50/90 (\u22640.015/0.06 mg/L) against methicillin-resistant S. epidermidis. Lefamulin also had good activity against Streptococcus pyogenes and Streptococcus agalactia (MIC50/90: \u22640.015/\u22640.015 mg/L), \u03b2-lactamase-producing Haemophilus influenzae (MIC50/90: 0.5/1 mg/L), \u03b2-lactamase-negative H. influenzae (MIC50/90: 1/1 mg/L), Moraxella catarrhalis (MIC50/90: 0.25/0.25 mg/L), and Mycoplasma pneumoniae (MIC50/90: 0.03/0.03 mg/L) regardless of resistance to azithromycin. Lefamulin was generally more active than the comparators against the test strains.Lefamulin showed potent activity against Staphylococcus, S. pneumoniae, \u03b2-hemolytic Streptococcus, H. influenzae, M. catarrhalis and M. pneumoniae). In vitro activity supports the use of lefamulin in the treatment of CABP in China.In summary, lefamulin has good and broad-spectrum coverage of respiratory pathogens (methicillin-sensitive and -resistant  Clitopilus scyphoides, Clitopilus passeckerianus, or other Clitopilus species in basidiomycota. Lefamulin is the first-in-class semi-synthetic pleuromutilin antibiotic for systemic use. Its molecular formula is C28H45NO5S (molecular weight 567.79 g). Lefamulin inhibits bacterial protein synthesis by binding to \u201cA\u201d and \u201cP\u201d sites of the peptidyl transferase center (PTC) of the 23s rRNA of the 50S ribosomal subunit of bacterial cell. The binding is through the mutilin core and C-14 side chain in the forms of hydrogen bonds, hydrophobic interactions, and conformational change to prevent correct orientation of tRNA\u2019s 3\u2019-CCA ends for peptide transfer  coded by the transposon Tn5406 and vga(A) carried by plasmids (encoding ABC transporter)  . So far,Streptococcus pneumoniae (PRSP), macrolide-resistant Mycoplasma pneumoniae, and methicillin-resistant S. aureus (MRSA)  . In Auguin vitro activity of lefamulin against a broad range of respiratory pathogens.The antibacterial spectrum and activity of lefamulin have been studied in the United States and Europe , 2019, bMycoplasma pneumoniae. These strains were isolated from 29 hospitals across China, representing 23 provinces and municipalities, during the period from October 2017 to July 2019. Specifically, the test strains included S. aureus (n = 121), S. epidermidis (n = 30), \u03b2-lactamase-producing Haemophilus influenzae (n = 48), \u03b2-lactamase-negative H. influenzae (n = 48), Haemophilus parainfluenzae (n = 10), Moraxella catarrhalis (n = 54), S. pneumoniae (n = 172), Streptococcus pyogenes (n = 30), and Streptococcus agalactiae (n = 13). All the strains were re-identified before susceptibility testing. Species identification was confirmed by MALDI-TOF/MS system , and antimicrobial susceptibility testing were controlled with reference strains S. aureus ATCC29213, S. pneumoniae ATCC49619, H. influenzae ATCC49247, and M. pneumoniae ATCC 29342.A total of 634 non-duplicate strains of respiratory pathogens were tested, including 580 strains of bacteria and 54 strains of M. pneumoniae were measured according to the methods for antimicrobial susceptibility testing for human mycoplasmas described in CLSI document M43-A (2011) (The minimum inhibitory concentrations (MICs) of lefamulin and the comparators were determined by broth microdilution method according to the Clinical and Laboratory Standards Institute  2018) M M2018) MA (2011) . The antth Edition .Staphylococcus strains , 0.06 mg/L against methicillin-sensitive S. aureus (MSSA), and 0.03 mg/L against methicillin-sensitive S. epidermidis (MSSE). Lefamulin displayed MIC values ranging from \u22640.015 mg/L to 0.25 mg/L (MIC90: \u22640.25 mg/L) against 172 strains of S. pneumoniae, including penicillin-susceptible (PSSP) strains (penicillin MIC \u22640.06 mg/L), penicillin-intermediate (PISP) strains (penicillin MIC: 0.125 mg/L\u20131 mg/L), and penicillin-resistant (PRSP) strains (penicillin MIC \u2265 2 mg/L). Lefamulin inhibited the growth of all PSSP strains at \u22640.015 mg/L and all PISP and PRSP strains at 0.25 mg/L. The MIC50/90 values of lefamulin were \u22640.015/\u22640.015 mg/L against S. pyogenes and \u22640.015/0.06 mg/L against S. agalactiae. Lefamulin inhibited the growth of all the S. pyogenes and S. agalactiae strains at 0.06 mg/L . Its activity was comparable to moxifloxacin and significantly superior to erythromycin and azithromycin  in this respect.In the present study, lefamulin displayed excellent antimicrobial activity against all the respiratory pathogens, including MRSA, MSSA, MRSE, MSSE, 015\u20132016 , 2019. THaemophilus and M. catarrhalis. Lefamulin was comparable to ceftriaxone in activity against S. pneumoniae strains  and \u03b2-hemolytic Streptococcus, but better than ceftriaxone against PRSP, better than penicillin against PISP and PRSP, and similar to penicillin against \u03b2-hemolytic Streptococcus. Lefamulin had similar activity as moxifloxacin, vancomycin, and linezolid against Streptococcus. It inhibited the growth of all Streptococcus species at 0.125 mg/L, which was lower than the above mentioned three agents. Lefamulin was significantly better than erythromycin and azithromycin in the activity against S. pneumoniae and \u03b2-hemolytic Streptococcus.Lefamulin also displayed high antimicrobial activity against H. influenzae and M. catarrhalis. As for the \u03b2-lactamase-negative strains, lefamulin provided significantly better activity than azithromycin. Lefamulin was comparable to tigecycline, ceftriaxone, and levofloxacin, and significantly superior to azithromycin and trimethoprim-sulfamethoxazole in the activity against M. catarrhalis. These results are consistent with those reports from other countries  (It has been reported that the 02 mg/L) .90 value of lefamulin was 0.5 \u03bcg/mL against the 50 strains of S. pneumoniae isolated from the patients in phase III clinical trial LEAP 1  was 0.12\u20130.25 \u03bcg/mL. The post-treatment bacterial clearance rate was up to 100%. Research results at home and abroad have shown that lefamulin had similar antimicrobial activity against S. epidermidis and S. aureus  over the MIC (24 h AUC/MIC). Lefamulin achieves rapid and predictable penetration into human tissues, with a mean 5.7-fold higher concentration in the pulmonary epithelial lining fluid compared with plasma. Percent probabilities of attaining the median AUCELF/MIC ratio targets associated with a 1-log10 CFU reduction from baseline by MIC were 97.0% at a MIC of 0.5 \u03bcg/mL for S. pneumoniae and 99.4% at a MIC of 0.25 \u03bcg/mL for S. aureus .DZ and FH designed the study. SW, YZ, YG, and DY performed the experimental work. SW and YZ collected the data. FH analyzed the data. All authors read and approved the final manuscript, contributed to the article, and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "T(temperature), Moisture content (MC), sediment total phosphorus (STP), ion-exchangeable form of ammonia (IEF-NH4+-N), weak-acid extractable form of ammonia (WAEF-NH4+-N), weak-acid extractable form of nitrate (WAEF-NO3\u2212-N), and strong-alkali extractable form of ammonia (SAEF-NH4+-N) were the dominant environmental factors and explained 11.1%, 8.2%, 10.7%, 6.9%, 9.3%, 8.1%, 10.5%, 7.5%, and 7% variation, respectively, of the total variation in the microbial community. Furthermore, the network analysis showed that symbiotic relationships accounted for a major percentage of the microbial networks. The keystone aerobic denitrifying bacteria belonged to Comamonas, Rhodobacter, Achromobacter, Aeromonas, Azoarcus, Leptothrix_Burkholderiales, Pseudomonas, Thauera, unclassified_Burkholderiales, and unclassified_bacteria. The composition of the keystone aerobic denitrifying microbial community also exhibited significant differences  on the basis of the adonis analysis. T, STP, IEF-NH4+-N, ion-exchangeable form of nitrate (IEF-NO3\u2212-N), WAEF-NO3\u2212-N, SAEF-NH4+-N, and TN were the dominant environmental factors that explained 8.4%, 6.2%, 4.6%, 5.9%, 5.9%, 4.5%, and 9.4% variation, respectively, of the total variation in the keystone aerobic denitrifying microbial community. The systematic investigation could provide a theoretical foundation for the evolution mechanism of the aerobic denitrifying microbial community in Baiyangdian Lake.Here, the ion-exchangeable form of nitrogen (IEF-N), weak-acid extractable form of nitrogen (WAEF-N), strong-alkali extractable form of nitrogen (SAEF-N), strong-oxidant extractable form of nitrogen (SOEF-N), residue nitrogen (Res-N), and total nitrogen (TN) showed spatial differences, and most of the sediment nitrogen fractions exhibited positive correlations in Baiyangdian Lake. High-throughput sequencing analysis revealed that the aerobic denitrification microbial community was composed of proteobacteria (42.04%\u201399.08%) and unclassified_bacteria (0.92%\u201357.92%). Moreover, the microbial community exhibited significant differences (R Thiosphaera pantotropha) [Excessive nitrogen concentrations present problems for water quality and cause eutrophication . An incrotropha)  has beenotropha) , Wadden otropha) , wetlandotropha) , coastalotropha)  and reseotropha) ,10. Therotropha) ,12.napA is the biomarker of aerobic denitrifying bacteria [napA) in Baiyangdian Lake.Currently, some full-scale experiments on bioaugmentation with aerobic denitrification bacteria have been conducted successfully for the remediation of wastewater, urban river, and river sediment. Such as, Duan et al. 2015) explored the nitrogen removal performance of halophilic heterotrophic nitrification-aerobic denitrification SF-16 in saline wastewater ; Du et a5 explorebacteria ,21. TherIn this study, our objectives were to (1) investigate the characteristics of environmental factors; (2) examine the composition of the aerobic denitrifying microbial community and keystone operational taxonomic unit (OTU); (3) evaluate the differences in abundance, diversity, and community structure; (4) analyze the relationship between the microbial community structure and environmental driving factors; and (5) estimate the relative contributions of the environmental driving factors.Baiyangdian Lake, located in Xiong\u2019an New Area, is the largest freshwater lake in the North China Plain a. Baiyan3\u2212-N, NO2\u2212-N, and NH4+-N were measured using DR6000  [T, DO, pH, ORP, and EC of the water samples were determined using Hydrolab DS5 . According to the standard methods, TN, NOny, USA) . The difny, USA) ,24.napA [https://submit.ncbi.nlm.nih.gov/subs/sra/) database with the accession number PRJNA623955.The whole DNA of the sediment was extracted using the Soil DNA Kit . After DNA purification, the extracted DNA was stored at \u221280 \u00b0C for PCR amplification. The extracted DNA was amplified using primer V66F, 5\u2032-TAYTTYYTNHSNAARATHATGTAYGG-3\u2032, and V67R, 5\u2032- DATNGGRTGCATYTCNGCCATRTT-3\u2032, for aerobic denitrifying bacterial napA ,25. PCR napA . High-thOn the basis of the OTU data, the richness index, Shannon index, Simpson index, Pielou index, Chao1 index, ACE index, and Good\u2019s Coverage were calculated to evaluate the alpha diversity. The differences in the microbial communities were evaluated using principal co-ordinate analysis (PCoA) and permutational MANOVA (adonis) in the vegan package (R.3.5.3). Redundancy analysis (RDA) was used to determine the correlation between the microbial community and predominant environmental factors with variance inflation factor (VIF) < 10 in the vegan package (R.3.5.3). Indicator species analysis was conducted using labdsv package (R.3.5.3). Hierarchical partitioning (HP) analysis was performed to quantitatively evaluate the relative influences of the environmental factors on the aerobic denitrifying bacterial community by using the rdacca.hp package (R.3.5.3) .Network analysis was performed to investigate the biotic interactions between the microbial populations, and the results were visualized with Gephi software (0.9.2). In this study, a positive correlation implied a mutualistic interaction, whereas a negative correlation may indicate competition . The \u201ckeNatural area. The IEF-N, WAEF-N, SAEF-N, SOEF-N, and Res-N values were 100.61, 2831.38, 1487.38, 995.42, and 1093.82 mg/kg, respectively , whereas it was negatively correlated with IEF-NO3\u2212-N , SAEF-NH4+-N , IEF- N , and IEF-NH4+-N  , IEF-N , SAEF-N , IEF-NH4+-N , SAEF-NO3\u2212-N , and TAN   and IEF-NO3\u2212-N . IEF-NO3\u2212-N was positively correlated with IEF-N  , SAEF-NO3\u2212-N , and TAN  (p < 0.05), IEF-NH4+-N , SAEF-NO3\u2212-N , TAN , SOEF-NO3\u2212-N , SOEF-N , and SOEF-NH4+-N  , SAEF-NO3\u2212-N , and TAN  , TAN , SOEF-NO3\u2212-N , SOEF-N , and SOEF-NH4+-N  , SOEF-NO3\u2212-N , SOEF-N , and SOEF-NH4+-N  , SOEF-N , and SOEF-NH4+-N   and SOEF-NH4+-N  .WAEF-NO < 0.05) . Res-N w< 0.001) . WAEF-NH < 0.01) . SAEF-NH < 0.01) . IEF-N w < 0.05) b. SAEF-N< 0.001) . IEF-NH4 < 0.05) . SAEF-NO < 0.05) . TAN was < 0.01) b. SOEF-N< 0.001) . SOEF-N After quality trimming, a total of 223754 sequences with an average length of 373 bp were obtained for the 14 sediment samples. MiSeq revealed a total of 1886 OTUs with 97% similarity . The Sha\u03b1-diversity was influenced by PCoA1 , MC (R = 0.70), STP (R = 1.00), IEF-NH4+-N (R = 0.97), and WAEF-NH4+-N (R = 0.80) were the dominant environmental parameters  , unclassified_bacteria (0.92%\u201357.92%), 0\u20130.25%) . ProteobAt the class level , BetaproComamonas (0\u201383.37%), unclassified_bacteria (1.44\u201358.39%), Rhodobacter (0.01\u201350.84%), unclassified_Burkholderiales (0.46\u201328.84%), Aeromonas (0.08\u201323.42%), Azoarcus (1.19\u201322.78%), Thauera (0.31\u201318.17%), Achromobacter (0.02\u201311.86%), Pseudomonas (0.11\u201310.78%), Sulfuritalea (0.09\u20139.75%), unclassified_betaproteobacteria (0.28\u20139.28%), Cedecea (0\u20138.16%), Dinoroseobacter (0\u20137.61%), Sulfurospirillum (0\u20137.54%), Leptothrix_Burkholderiales (0.29\u20135.61%), Magnetospirillum (0.19\u20134.59%), Azospirillum (0.03\u20133.52%), Azospira (0\u20133.30%), Rhodopseudomonas (0.16\u20133.09%), and Rhizobium (0\u20132.10%) (Comamonas (BH), unclassified_Betaproteobacteria (BGYH), Achromobacter (BGYH), Sulfurospirillum (PH), Azospira (BGYH), and Azospirillum (PH); for the breeding area, unclassified_bacteria (FYD) and Thauera (SHD); for the tourist area, Sulfuritalea (YYD) and Dinoroseobacter (YYD); for the living area, unclassified_Burkholderiales (BTZXD), Aeromonas (ZLZ), Magnetospirillum (ZLZ), Cedecea (PYD), Leptothrix_Burkholderiales (BTZXD), and Rhizobium (BTZXD); and for the natural area, Azoarcus (ZZD), Rhodobacter (ZZD), Pseudomonas (ZZD), and Rhodopseudomonas (ZZD). Previous studies have shown that these dominant genera are aerobic denitrifying bacteria. For example, Comamonas is capable of heterotrophic nitrification-aerobic denitrification nitrogen removal, and it has been successfully used for bioaugmentation of municipal wastewater [Rhodobacter is a typical denitrifying bacterium in alkaline copper mine drainage [Aeromonas sp. HN-02 [Aeromonas, Pseudomonas, and Acinetobacter) could be successfully enriched and conducted denitrification to remove 90% of TN in a wetland test system [Azoarcus was an important genus in a denitrifying quinoline-removal bioreactor [Thauera was identified as the key potential heterotrophic nitrification and aerobic denitrification genus for principal nitrogen removal in granular reactors [Achromobacter xylosoxidans CF-S36, a typical heterotrophic nitrification-aerobic denitrification bacterium, were investigated through molecular analysis [Pseudomonas indoloxydans YY-1, through physiological and transcriptomic analyses [Pseudomonas putida strain NP5 [Azospira was the main dentrifier in a tidal flow constructed wetland [Halomonas was the dominant heterotrophic nitrifying/aerobic denitrifying genus under hypersaline conditions [Rhodopseudomonas is a potential autotrophic denitrifying genus that can use electrons and reducing power from cathodes [At the genus level, the top 20 abundant genera are as follows: 0\u20132.10%) . Moreovestewater  and enristewater . Liu et nas 0\u201383.%, unclasdrainage . Chen et 1.44\u201358.%, Rhodobt system . Azoarcuoreactor , and it  0.08\u201323.%, Azoarcreactors . The dynanalyses  and Yangriales 0.\u201328.84%,  wetland , reservo1.44\u201358.3%, Rhodob wetland . Halomonnditions . Rhodopster 0.01\u2013.84%, uncDifferences in the composition of the aerobic denitrifying microbial community in the five areas were investigated with PCoA and NMDS analysis. RDA was used to demonstrate the links between the environmental parameters and microbial community. HP analysis was performed to investigate the relative influence of the environmental driving factors on microbial community composition.For the whole aerobic denitrifying microbial community. The PCoA results showed that PCoA1 and PCoA2 accounted for 42.91% and 17.42%, respectively, of the variability in the microbial community composition  on the basis of the adonis analysis. The stress result (stress = 0.06 < 0.1) of NMDS showed that the NMDS analysis exhibited good representation  on the basis of the adonis analysis. The stress result (stress = 0.04 < 0.05) of NMDS showed that NMDS analysis exhibited excellent representation , module 4 , module 6 , module 7 , and module 8 ; STP and WAEF-NO3\u2212-N exhibited a significantly negative correlation with modules 1\u20138, and the correlation reached \u22120.56 to \u22120.58 (p < 0.05) and \u22120.64 (p < 0.05); WAEF-NH4+-N, Res-N, and TN were positively correlated with modules 1\u20138, and the correlation reached 0.54\u20130.64 (p < 0.05), 0.65\u20130.70 (p < 0.05), and 0.67\u20130.71 (p < 0.01). WAEF-N was positively correlated with module 1 , module 2 , module 5 , and other modules . Especially, the keystone aerobic denitrifying microbial community exhibited a significantly positive correlation with T , WAEF-NH4+-N , Res-N , and TN  and a significantly negative correlation with STP . On the basis of all the results, T, STP, WAEF-NH4+-N, WAEF-NO3\u2212-N, and TN were important environment factors, which is consistent with the RDA and HP analysis results.The correlations between the modules and environmental factors were investigated in Baiyangdian Lake c. The ennapA). In this study, the environmental parameters  exhibited significantly spatial differences among the different functional areas, and most of the sediment nitrogen fractions exhibited positive correlations in Baiyangdian Lake. MiSeq revealed a total of 1886 OTUs identified as proteobacteria (42.04\u201399.08%), unclassified_bacteria (0.92\u201357.92%), and Deinococcus-Thermus (0.00\u20130.25%). The unclassified genus accounted for an important part in Baiyangdian Lake. The RDA and VIF results showed that T, MC, STP, IEF-NH4+-N, and WAEF- NH4+- N were the dominant environment parameters and explained 11%, 13.6%, 27.6%, 16.3%, and 2.8% variation in \u03b1-diversity. Moreover, the microbial community exhibited significantly spatial differences . T, MC, STP, IEF-NO3\u2212-N, WAEF-NH4+-N, WAEF-NO3\u2212-N, and SAEF-NH4+-N were the dominant environmental factors and explained the 11.1%, 8.2%, 10.7%, 9.3%, 8.1%, 10.5%, and 7.5% variation of the total variation in the whole aerobic denitrifying microbial community, respectively. The network analysis showed that symbiotic relationships accounted for a major percentage of the microbial networks, and 40.18% nodes belonged to betaproteobacteria. The keystone OTUs belonged to Comamonas, Rhodobacter, Achromobacter, Aeromonas, Azoarcus, Leptothrix_Burkholderiales, Magnetospirillum, Pseudomonas, Sulfuritalea, and Thauera. Furthermore, the composition of the keystone aerobic denitrifying microbial community also exhibited significantly spatial differences , and T, STP, IEF-NH4+-N, IEF-NO3\u2212-N, WAEF-NO3\u2212-N, SAEF-NH4+-N, and TN were the dominant environmental factors. From all the results, this study supplied a new view on investigating the distribution characteristics and driving factors of the aerobic denitrifying microbial community in Baiyangdian Lake.This is the first report of the characteristics and driving factors of the aerobic denitrifying microbial community, especially, the \u201ckeystone species\u201d and the dominant environment factor in natural water ecosystem through the aerobic denitrification functional gene ("}
+{"text": "Correction to: Isr J Health Policy Res 10, 12 (2021)https://doi.org/10.1186/s13584-021-00449-xFollowing publication of the original article , the autBelow the incorrect and correct table description:Incorrect: Table\u00a01 Cumulative COVID-19 vaccination doses administered per 100 people by Feb. 1, 2021, or latest date availableCorrect: Table\u00a01: Cumulative COVID-19 vaccination doses administered per 100 people by Feb. 11, 2021, or latest date availableThe original article has been updated."}
+{"text": "Stem rust in recent years has acquired an epiphytotic character, causing significant economic damagefor wheat production in some parts of Western Siberia. On the basis of a race composition study of the stem rustpopulations collected in 2016\u20132017 in Omsk region and Altai Krai, 13 pathotypes in Omsk population and 10 inAltai population were identified. The race differentiation of stem rust using a tester set of 20 North AmericanSr genes differentiator lines was carried out. The genes of stem rust pathotypes of the Omsk population are avirulentonly to the resistance gene Sr31, Altai isolates are avirulent not only to Sr31, but also to Sr24, and Sr30. A lowfrequency of virulence (10\u201325 %) of the Omsk population pathotypes was found for Sr11, Sr24, Sr30, and for Altaipopulation \u2013 Sr7b, Sr9b, Sr11, SrTmp, which are ineffective in Omsk region. Field evaluations of resistance to stemrust were made in 2016\u20132018 in Omsk region in the varieties and spring wheat lines from three different sources.The first set included 58 lines and spring bread wheat varieties with identified Sr genes \u2013 the so-called trap nursery. The second set included spring wheat lines from the Arsenal collection,that were previously selected according to a complex of economically valuable traits, with genes for resistanceto stem rust, including genes introgressed into the common wheat genome from wild cereal species. The thirdset included spring bread wheat varieties created in the Omsk State Agrarian University within the framework ofa shuttle breeding program, with a synthetic wheat with the Ae. tauschii genome in their pedigrees. It was establishedthat the resistance genes Sr31, Sr40, Sr2 complex are effective against stem rust in the conditions of WesternSiberia. The following sources with effective Sr genes were selected: (Benno)/6*LMPG-6 DK42, Seri 82, Cham 10,Bacanora (Sr31), RL 6087 Dyck (Sr40), Amigo , Siouxland , Roughrider , Sisson, and Fleming , Pavon 76 (Sr2 complex) from the ISRTN nursery; No. 1 BC1F2(96 \u00d7 113) \u00d7 145 \u00d7 113 , No. 14\u0430 F3 (96 \u00d7 113) \u00d7 145 , No. 19 BC2F3 (96 \u00d7 113) \u00d7 113 , and No. 20 F3 (96 \u00d7 113) \u00d7 145  from the Arsenal collection; and the Omsk State AgrarianUniversity varieties Element 22 , Lutescens 27-12, Lutescens 87-12 , Lutescens 70-13, andLutescens87-13 . These sources are recommended for inclusion in the breeding process for developingstem rust resistant varieties in the region. Stem rust of wheat caused by Puccinia graminis f. sp. triticiErikss. for a long time had a weak manifestation in the territoryof Western Siberia and only in the recent years acquiredan epiphytotic nature, causing significant economic damagefor wheat production in the region. First of all, this is due tothe deterioration of the phytosanitary situation in the region,the general trend of climate warming and cultivation of susceptiblewheat varieties on large area . The threat of stem rust race Ug99 appearance and theemergence of new pathotypes of this race, affecting varietieswith genes Sr24 and Sr36 present a serious threat for wheatproduction in West Siberian region. Genetic diversity of cultivatedwheat varieties for resistance to Ug99 and stem rustin general is very limited .Enhancement of genetic resistance to pathogens can besolved germplasm exchange, and also cultivation of varietieswith different level of resistance to diseases and to differentraces. Crop protection is necessary to restrain the evolutionof pathogens and the emergence of new virulent races.Such programs are widely used in Europe and America. Theduration of the variety cultivation in advanced countries is3\u20134 years, while in Russia \u2013 7\u201310 years . Inthis regard, the breeding of spring wheat varieties, whichhave a diverse genetic basis of resistance to stem rust, is veryrelevant.Amigo .Since the 1950s, many resistance genes introduced intobread wheat have lost their effectiveness .The most significant genes for breeding practice are Sr2,Sr23, Sr24, Sr25, Sr31, Sr33, Sr36, Sr38, Sr45, Sr50, SrTmp,Sr1RSIntrogression of resistance genes of wild and cultivatedwheat relatives allows to expand the genetic diversity ofvarieties and contributes to their long-term protection . To date, about 86 Sr genes have beenidentified,of which 26 stem rust resistance genes havebeen transferred into bread wheat from other cereal species. For example, T. turgidum was thesource of the stem rust resistance genes Sr2, Sr9d, Sr9e, Sr9g,Sr11, Sr12, Sr13, Sr14, and Sr17, of which the Sr2, Sr13, andSr14 genes are effective against Ug99 race; T. monococcumwas the source of Sr21, Sr22, and Sr35 genes .Genes that caused the resistance to stem rust have beenintroduced into wheat gene pool from the genome of variousAegilops L. species: Ae. speltoides \u2013 Sr32, Sr39, Sr47;Ae. comosa \u2013 Sr34; Ae. ventricosa \u2013 Sr38 . Ae. tauschii contributed genes Sr33, Sr45, Sr46 . Direct hybridization of T. aestivum withAe. tauschii and following backcrosses allowed introductionof new resistance genes SrTA1662, SrTA1017, and SrTA10187effective against Ug99 race . The searchof new resistance genes in wild wheat relatives continues, forexample, G. Yu et al. (2017) identified two new Sr genes inAe. sharonesis.One of the objectives of Kazakh-Siberian Spring WheatImprovement Network (KASIB) is expanding of the geneticpolymorphism of new varieties, including resistance to harmfuldiseases . This is based onshuttle breeding with CIMMYT (Mexico). Varieties andbreeding lines developed through shuttle breeding with participationof Ae. tauschii and T. dicoccum, as well as linesof the \u201cArsenal\u201d collection, which have wild species in theirpedigree are of interest for breeding for resistance to stemrust in the region.The aim of the research was analysis of the racial compositionof the Western-Siberian stem rust population, resistanceassessment of spring bread wheat lines and varieties with identified resistance genes and identification of the sourceswith effective Sr genes for breeding under Western Siberianconditions.http://agro.au.dk/forskning/internationaleplatforme/wheatrust).The racial composition of Puccinia graminis f. sp. tritici populationscollected in 2016\u20132017 in Omsk region ) and Altai region  were analyzed in the Global Rust Reference Center was carriedout. Monopustule isolates were reproducted to identifyrace Ug99 with usage of the test PCR-Stage 1. A total of19 single pustule isolates were selected from Omsk populationand 20 \u2013 from Altai population (Table 1).Selection of single pustule isolates according to requirementsof GRRC protocols (Differentiation of stem rust races was performed with useof the set of 20 North American differentiator lines containingSr genes: Sr5 (ISr5-Ra), Sr21 (CnS_Triticum monoc.Deriv.), Sr9e (Vernstein), Sr7b (ISr7b-Ra), Sr11 (ISr11-Ra), Sr6 (ISr6a-Ra), Sr8a (ISr8a-Ra), Sr9g (CnSr9g), Sr36(W2691SrTt-1), Sr9b (W2691Sr9b), Sr30 (BtSr30Wst),Sr17+13 (Combination VII), Sr9a (ISr9a-Ra), Sr9d (ISr9d-Ra), Sr10 (W2691Sr10), SrTmp (CnsSrTmp), Sr24 (LcSr24Ag),Sr31 (Benno Sr31/6*LMPG), Sr38 (VPM-1), SrMcN(McNair 701). Infected plants were evaluated in 14\u201316 daysafter inoculation according to modified E.C. Stakman scale. Virulence phenotypes were classifiedaccording to North American system .The varieties and lines of bread wheat from three germplasmsets were evaluated in Omsk at least 4\u20135 times for reaction tostem rust on scales recommended by Koyshibaev et al. (2014).The type of reaction on E.B. Mains and H.S. Jacksonscale(1926) and severity \u2013 on modified Peterson scale  were considered: 0 \u2013 immunity, uredopustulesnot formed; R (Resistance \u2013 high resistance), 1 score, severity 5\u201310 %; MR (Moderately resistant \u2013 average resistance),2 score, severity 10\u201325 %; M (heterogeneous type), pustulesof different sizes, surrounded by chlorotic and necrotic spotsor without them; MS (Moderately susceptible \u2013 averagesusceptibility), 3 score, severity 40\u201350 %; S (Susceptible \u2013susceptibility), 4 score, severity more than 60 %.In 2016\u20132018, International Stem Rust Trap Nursery with58 genotypes with identified Sr genes was evaluated to Omskstem rust population (Table 2). Varieties and lines of nurserytrapwere sown manually in 100 cm-long rows with stem rustresistant (Element 22) and susceptible checks (Chernyava 13)alternating every entries.1F2 (96 \u00d7 113) \u00d7 145 \u00d7 113]; No. 13, 14\u0430 [F3 (96 \u00d7 113) \u00d7 145];No. 16, 17, 17\u0430 [BC1F4 (96 \u00d7 113) \u00d7 113]; No. 19 [BC2F3(96 \u00d7 113) \u00d7 113]; No. 20, 22\u0430 [F3 (96 \u00d7 113) \u00d7 145]. The lineswere studied in 2016\u20132018 in un-replicated trial with the plotsize of 2 m2.In 2015, 9 spring wheat lines originating from wide crosses\u201cArsenal\u201d collection were kindly provided by I.F. Lapochkinafor evaluation in Omsk. These lines carry a pyramid ofstem rust resistance genes  \u2013 No. 1[BCNine spring wheat varieties and breeding lines from advancedyield trial at Omsk SAU developed through utilizationof synthetic wheat with the Ae. tauschii genome (Lutescens24-12 (Kasibovskaya), Lutescens 27-12, Lutescens87-12, Lutescens 70-13, Lutescens 87-13, Lutescens 88-13(Silantiy), Lutescens 124-13, Lutescens 53-15, Lutescens 128-15) were evaluated for stem rust resistance and other traits in2016\u20132018. The plot size was 25 m2 with four replications.The checks were Pamyati Azieva (early maturing), Duet (mediummaturing), and Element 22 (late maturing).http://maswheat.ucdavis.edu/protocols/StemRust/index.htm). The resistance genes of spring bread wheat lines and varietiesfrom nursery-trap and from collection \u201cArsenal\u201d were identifiedearlier .Sr genes of Omsk SAU varieties were identified usingmolecular markers: Xsts638 \u2013 Sr15, Xcfa2123 \u2013 Sr22,Xgwm210 \u2013 Sr23, Xscs73 \u2013 Sr24, Xwmc221 \u2013 Sr25,BE518379 \u2013 Sr26, Xscm09 \u2013 Sr31, SCS421 \u2013 Sr34,Xcfa2170 \u2013 Sr35, Xstm773-2 \u2013 Sr36, Ventriup-LN2 \u2013 Sr38,Lr34plus \u2013 Sr57, according to established protocol . In all studiedWestern-Siberian populations of P. graminis Ug99 and Sicilianraces were not identified. Genes of stem rust pathotypesof Omsk population were avirulent only to Sr31 gene, whileAltai pathotypes were avirulent to Sr31, Sr24, and Sr30.The race composition analysis of stem rust populationsidentified a significant number of pathotypes: in the Omskpopulation \u2013 13 and in Altai population \u2013 10 (see Table 1).Unlike many regions of the world where stem rust is a harmfuldisease for decades, for example in Krasnodar region of Russia, for Western Siberia this is surprisingresult considering a short period of time since its appearance.Most of the identified pathotypes of stem rust population inOmsk and Altai regions were not identical in virulence to thepathotypes, which were found in recent years in Asia andAfrica (Low frequency of virulence (10\u201325 %) of Omsk populationpathotypes was established for Sr11, Sr24, Sr30 genes, forAltai population \u2013 for Sr7b, Sr9b, Sr11, SrTmp genes, whichwere ineffective in Omsk region. The results of laboratoryevaluation of virulence of P. graminis pathotypes collectedin Omsk region were confirmed by field of trap nursery withidentified Sr genes (see Table 2).Genotypes with Sr31: Sr31(Benno)/6*LMPG-6 DK42,Seri 82, PBW343=Attila with Sr31, Cham 10=Kauz//Kauz/star, Bacanora=Kauz\u2019s\u2019 showed high level of resistance toOmsk stem rust population in all years of study (2016\u20132018).Line 28 LcSr24Ag + BTSr24Ag with Sr24 gene was characterizedby moderate resistance. For some Sr genes, resistanttype of reaction under epiphytotic conditions was observedon the stage of adult plants, and susceptible type \u2013 on theseedling stage in the laboratory conditions.For example, variety Trident (entries 46 and 47) withSr38 gene had high resistance (R\u20135MR) in the field; varietyEinkorn (entry 25) with Sr21 gene, and line W2691SrTt-1CI 17385 (entry 44) with Sr36 gene had moderate resistance(10M) in the field conditions. In the laboratory conditionsthe seedlings plants with above mentioned genes were classifiedas susceptible. Genotypes of ISRTN nursery with agene pyramid had high resistance to stem rust in all years ofresearch: entry 50 Amigo (Sr24 + 1RS-Am), entry 51 Siouxland(Sr24 + Sr31), entry 52 Roughrider (Sr6 + Sr36), entry 53Sisson (Sr6 + Sr31 + Sr36), entry 55 Fleming (Sr6 + Sr24 +Sr36 + 1RS-Am). The results of stem rust resistance evaluationof \u201cArsenal\u201d collection and Omsk SAU germplasm arepresented in Table 3.Lines from \u201cArsenal\u201d collection are of great interest assources of resistance to pathogen since they possess thegene pyramid: Sr2 (T. turgidum), Sr36, Sr40 (T. timopheevii),Sr44 (Th. intermediate). The pedigree of selected linescontains spring wheat line 13/00/i-4 with 7 resistance genes: Sr2, Sr36, Sr39, Sr40, Sr44, Sr47, Sr15, and winter lineGT 96/90 with genes Sr15, Sr24, Sr31, Sr36, Sr40, Sr47.In Omsk SAU varieties 3 resistance genes were identified:Sr23, Sr31, Sr36. Variety Element 22, which has winter wheatAurora in its pedigree also possesses wheat-rye translocation1BL.1RS with Sr31 gene . Thecombination of effective resistance genes Sr31 and Sr35 inthis variety results a high level of resistance to stem rust. Element22 is one of the few varieties with combined resistanceto stem and leaf rust. It was included into State register ofbreeding achievements in Western Siberian region. This varietyis the check of the late maturity group at the State VarietyTrials in Omsk region.Stem rust resistant breeding lines Lutescens 27-12, Lutescens70-13, Lutescens 87-13, Lutescens 88-13 were selectedfrom a cross Lutescens 30-94*2/3/T. dicoccon PI 94625/Ae. squarrosa (372)//3*Pastor involving Kazakhstan springwheat line Lutescens 30-94 and CIMMYT line developedby hybridization of synthetic wheat with variety Pastor. Theline Lutescens 87-12 originated from a cross Kazakhstanskaya25/2*Attila/3/T. dicoccon PI 94625/Ae. squarrosa alsoinvolving synthetic wheat. Omsk SAU germplasm possesseddifferent combinations of genes Sr23, Sr31, and Sr36.In modern conditions, stem rust is the most dangerous diseasefor grain production in Western Siberia. In the epiphytoticyears the grain losses of wheat in the region were about 2 milliontons. Unfortunately, stem rust resistant varieties includedinto the State register occupy about 10\u201315 % of the totalwheat sowing area in the region. In 2015\u20132016, evaluationof spring wheat varieties at Moskalenskiy State Variety Trialof Omsk region (southern forest-steppe zone) demonstratedthat out of 57 varieties tested only Element 22 (Sr31 + Sr35),Omskaya 37, Sigma, Uralosibirskaya (Sr31), and Sigma 2(Sr31 + Sr25) were resistant to stem rust (5\u201315MR). The othervarieties were affected by pathogen in medium and high degreerequiring the use of chemical protection . Previously, Shamanin et al. (2016b) identified thestem rust resistance genes in the germplasm developed bybreeding institutions of Western Siberia. High frequency ofgenes Sr25, Sr31, and their combination was observed. Highvariability of the race composition of the pathogen population,as shown in our studies, and the uniformity of resistance genesto stem rust in cultivated varieties, threaten grain productionstability in Western Siberia.The breeding strategy should focus on limiting diseasedevelopment in the region. The study of the populations ofP. graminis, formed on wheat in the different regions, is veryessential to guide the breeding efforts. There were no clonesavirulent to Sr24 gene in Omsk population of P. graminiswhile in Altai region there were no clones virulent to Sr24,which remains its effectiveness in Novosibirsk region . The results of the population compositioncomparison suggest that Omsk and Altai subpopulations haverelatively independent sources of genetic diversity and the contact zone. Western Siberian population of P. graminis hasquite complex structure. Two subpopulations are assumed toexist: Omsk and Altai \u2013 with independent sources of geneticdiversity, and zone of genotypic exchange on wheat crop inNovosibirsk region .Omsk stem rust population analysis showed that the spectrumof effective resistance genes has narrowed due to lossesof some genes to the local population of P. graminis.Highly resistant varieties and lines of ISRTN nurserywere identified: Sr31 (Benno)/6*LMPG-6 DK42, Seri 82,Cham 10, Bacanora (Sr31), RL 6087 Dyck (Sr40), Amigo, Siouxland , Roughrider , Sisson , Fleming , Pavon 76 (Sr2 complex). Selected varieties andlines are recommended for using as sources of resistance inbreeding programs to create resistant wheat varieties to stemrust. Effective resistance genes Sr31, Sr40, Sr2 complex,and their combinations with ineffective genes are recommendedfor use in breeding, taking into account the constantrotation, combination of genes of nonspecific resistance, aswell as the possibility of infection threat from neighboringterritory.The resistance gene Sr2, widely used in breeding for resistanceto virulent stem rust races, is common in commercial varietiesin a number of countries around the world, particularlyin the United States, Australia, India, and Mexico. This geneis practically absent in the commercial varieties of RussianFederation, however, for effective protection against stem rust,its pyramiding with other resistance genes is recommended.For the development of varieties with long-term resistance,the strategy of combining genes responsible for differenttypes of resistance in one genotype is used. Pyramiding ofspecific resistance genes  withAPR gene Sr2, which causes the slow development of thedisease (slow rusting), will provide longer protection ofwheat crops from stem rust in Western Siberia in the presentphytosanitary situation.1F2 (96 \u00d7 113) \u00d7 145 \u00d7 113 ; No. 14\u0430 F3(96 \u00d7 113) \u00d7 145 ; No. 19 BC2F3 (96 \u00d7 113) \u00d7 113; No. 20 F3 (96 \u00d7 113) \u00d7 145  represent a promising starting material for breedingand creation of varieties with long-term resistance.In this regard, the lines from \u201cArsenal\u201d collection \u2013 No. 1BCIt is justified to include resistance sources to stem rust withminimum number of negative traits that reduce their breedingvalue. In this regard, stem rust resistant germplasm fromOmsk SAU with identified effective genes Element 22 , Lutescence 27-12, Lutescence 87-12 ,Lutescence 70-13, Lutescence 87-13 ,Lutescence 88-13 (Sr23) are valuable starting material forbreeding in the region.1F2 (96 \u00d7 113) \u00d7 145 \u00d7 113 , No. 14\u0430 F3(96 \u00d7 113) \u00d7 145 , No. 19 BC2F3 (96 \u00d7 113) \u00d7 113, No. 20 F3 (96 \u00d7 113) \u00d7 145 , varieties of Omsk Agrarian University \u2013 Element 22, Lutescens 27-12, Lutescens 87-12 ,Lutescens 70-13, Lutescens 87-13  arerecommendedfor inclusion into breeding process of thecreation of resistant to stem rust varieties in the region. Furthermonitoring of the virulence of stem rust pathogen andcoordination strategy of breeding programs in Western Siberia,and neighboring regions of the Kazakhstan Republic isrecommended. Incorporation of effective resistance genes, inparticular Sr2 and Sr40, will improve the phytosanitary situationand expand the segment of resistant varieties in the region.Thus, the genetic similarity of spring wheat varieties on stemrust resistance genes cultivated over large areas in WesternSiberia, and the predominance of varieties with race specific resistance genes contribute to spreading and high variabilityof the pathogen. The lines from collection \u201cArsenal\u201d \u2013 No. 1BCThe authors declare no conflict of interest.Ablova I.B., Bespalova L.A., Kolesnikov F.A., Nabokov G.D.,Kovtunenko V.Ya., Filobok V.A., Davoyan R.O., KhudokormovaZh.N., Mokhova L.M., Levchenko Yu.G., Tarkhov A.S. Principlesand methods of wheat breeding on tolerance to diseasesin KRIA named after P.P. Lukiyanenko. Zernovoe KhozjaistvoRossii = Grain Economy of Russia. 2016;5:32-36. (in Russian)Baranova O.A., Lapochkina I.F., Anisimova A.V., Gajnullin N.R.,Iordanskaya I.V., Makarova I.Yu. Identification of Sr genes innew common wheat sources of resistance to stem rust race Ug99using molecular markers. Russ. J. Genet.: Appl. Res. 2016;6(3):344-350.Gomez-Becerra H., Morgounov A., Abugalieva A. Evaluation ofyield grain stability, reliability and cultivar recommendation inspring wheat (Triticum aestivum) from Kazakhstan and Siberia.Central Eur. J. Agriculture. 2006;6:649-660.Jin Y., Szabo L.J., Pretorius Z.A., Singh R.P., Ward R., Fetch T.,Jr. Detection of virulence to resistance gene Sr24 within raceTTKS of Puccinia graminis f. sp. tritici. Plant Dis. 2008;92:923-926.Kerber E.R., Dyck P.L. Resistance to stem and leaf rust in Aegilopssquarrosa and transfer of a gene for stem rust resistanceto hexaploid wheat. In: Ramanujam S. (Ed.). Proceeding of the5th International Wheat Genetics Symposium. New Delhi: Ind.Soc. of Genetics and Plant Breeding, Ind. Agric. Res. Institute,1979;358-364.Koyshybaev M., Shamanin V.P., Morgunov A.I. Screening of Wheatfor Resistance to Major Diseases. Ankara: FAO-SEK, 2014. (inRussian)Lapochkina I.F., Baranova O.A., Shamanin V.P., Volkova G.V.,Gainullin N.R., Anisimova A.V., Galinger D.N., Lazareva E.N.,Gladkova E.V., Vaganova O.F. The development of the initialmaterial of spring common wheat for breeding for resistance tostem rust (Puccinia graminis Pers. f. sp. tritici), including theUg99 race, in Russia. Russ. J. Genet.: Appl. Res. 2017;7(3):308-317. DOI 10.1134/S207905971703008X.Leonova I.N., Orlovskaya O.A., R\u00f6der M.S., Nesterov M.A., BudashkinaE.B. Molecular diversity of common wheat introgressionlines (T. aestivum/T. timopheevii). Russ. J. Genet.: Appl.Res. 2015; 5(3):191-197. DOI 10.1134/S2079059715030090.Mains E.B., Jackson H.S. Physiologic specialization in the leaf rustof wheat, Puccinia triticina Erikss. Phytopathology. 1926;16(2):89-120.McIntosh R.A., Dubcovsky J., Rogers W.J., Morris C., Xia X.C.Catalogue of gene symbols for wheat: 2017 Supplement. Availableat: http://shigen.nig.ac.jp/wheat/komugi/genes/macgene/supplement 2017.pdfMcIntosh R.A., Yamayaki Y., Dubcovsky J., Rogers W.J., MorrisC., Appels R., Xia X. Catalogue of Gene Symbols for Wheat:2013\u20132014 Supplement. Available at: http://www.shigen.nig.ac.jp/wheat/komugi/genes/macgene/supplement2013.pdfOlson E.L., Rous M.N., Pumphrey M.O., Bowden R.L., Gill B.S.,Poland J.A. Introgression of stem rust resistance genesSrTA10187 and SrTA10171 from Aegilops tauschii to wheat.Theor. Appl. Genet. 2013;126:2477-2484. DOI 10.1007/s00122-013-2148-z.Peterson R.F., Campbell A.B., Hannah A.E. A diagrammatic scalefor estimating rust intensity on leaves and stems of cereals. Can.J. Res. (Sect. C). 1948;26:496-500.Roelfs A.P., Martens J.W. An international system of nomenclaturefor Puccinia graminis f. sp. tritici. Phytopathology. 1988;78:526-533.Sanin S.S. Problems of phytosanitary of grain production. In: Protectionof Cereal Crops Against Diseases, Pests, and Weeds:Progress and Problems: Proc. of the Int. sci. and pract. conf.Bolshye Vyazemy, 5\u20139 Dec. 2016. Moscow, 2016;4-15. (in Russian)Schneider A., Moln\u00e1r I., Moln\u00e1r-L\u00e1ng M. Utilisation of Aegilops(goatgrass) species to widen the genetic diversity of cultivatedwheat. Euphytica. 2008;163:1-19. DOI 10.1007/s10681-007-9624-y.Shamanin V.P., Morgunov A.I., Petukhovskiy S.L., Likhenko I.E.,Levshunov M.A., Salina E.A., Pototskaya I.V., Trushchenko A.Ju. Breeding of spring bread wheat for resistance to stem rustin West Siberia. Omsk: FGOU VPO OmGAU Publ., 2015. (inRussian)Shamanin V.P., Pototskaya I.V., Klevakina M.V. Assessment of theSiberian collection of spring wheat on resistance to stem rustin the southern forest-steppe of Western Siberia. Vestnik KazanskogoGAU = The Herald of Kazan State Agrarian University.2016a;2(40):55-59. (in Russian)Shamanin V., Salina \u0415., Wanyera R., Zelenskiy Yu., Morgounov A.Genetic diversity of spring wheat from Kazakhstan and Russiafor resistance to stem rust Ug99. Euphytica. 2016b;12:287-296.DOI 10.1007/s10681-016-1769-0.Singh R.P., Hodson D.P., Huerta-Espino J., Jin Y., Njau P., WanyeraR., Herrera-Foessel S.A., Ward R.W. Will stem rust destroythe world\u2019s wheat crop? Adv. Agron. 2008;98:271-309. DOI10.1016/S0065-2113(08)00205-8.Singh R.P., Hodson D.P., Jin Y., Lagudah E.S., Ayliffe M.A., BhavaniS., Rouse M.N., Pretorius Z.A., Szabo L.J., Huerta-EspinoJ., Basnet B.R., Lan C., Hovm\u00f8ller M.S. Emergence andspread of new races of wheat stem rust fungus: Continued threatto food security and prospects of genetic control. Phytopathology.2015;105:872-884.Singh R.P., Huerta-Espino J., Bhavani S., Herrera-Foessel S.A.,Singh D., Singh P.K., Velu G., Mason R.E., Jin Y., Njau P., CrossaJ. Race non-specific resistance to rust diseases in CIMMYTspring wheats: breeding and advances. Euphytica. 2011;179:175-186. DOI 10.1007/s10681-010-0322-9.Skolotneva E.S., Bukatich E.Ju., Bojko N.I., Piskarev V.V., SalinaE.A. Screening of an international stem rust nursery trapfor Ug99 in the Priobie forest-steppe in 2017. In: Gene Pooland Plant Breeding: Proc. IV Int. sci. and pract. conf., 4\u20136 April2018. Novosibirsk, 2018;313-318. (in Russian)Skolotneva E.S., Kel\u2019bin V.N., Morgunov A.I., Bojko N.I., ShamaninV.P., Salina E.A. Races composition of the Novosibirskpopulation of Puccinia graminis f. sp. tritici. Mikologiya iFitopatologiya= Mycology and Phytopathology. 2020;54(1):49-58. (in Russian)Yu G., Champouret N., Steuernagel B., Olivera P.D., Simmons J.,Williams C., Johnson R., Moscou M.J., Hern\u00e1ndez-Pinz\u00f3n I.,Green P., Sela H., Millet E., Jones J.D.G., Ward E.R., SteffensonB.J., Wulff B.B.H. Discovery and characterization of twonew stem rust resistance genes in Aegilops sharonensis. Theor.Appl. Genet. 2017;130:1207-1222."}
+{"text": "The F2 populations derived from these crosses were genotyped using InDel marker specific to crtRB1. Severe marker segregation distortion was observed. Seventeen crtRB1 inbreds developed through marker-assisted pedigree breeding and seven inbreds generated using marker-assisted backcross breeding were characterized using 77 SSRs. Wide variation in gene diversity (0.08 to 0.79) and dissimilarity coefficient (0.28 to 0.84) was observed. The inbreds were grouped into three major clusters depicting the existing genetic diversity. The crtRB1-based inbreds possessed high \u03b2-carotene (BC: 8.72\u03bcg/g), \u03b2-cryptoxanthin (BCX: 4.58\u03bcg/g) and proA (11.01\u03bcg/g), while it was 2.35\u03bcg/g, 1.24\u03bcg/g and 2.97\u03bcg/g in checks, respectively. Based on their genetic relationships, 15 newly developed crtRB1-based inbreds were crossed with five testers (having crtRB1 gene) using line \u00d7 tester mating design. 75 experimental hybrids with crtRB1 gene were evaluated over three locations. These experimental hybrids possessed higher BC (8.02\u03bcg/g), BCX (4.69\u03bcg/g), proA (10.37\u03bcg/g) compared to traditional hybrids used as check . Environment and genotypes \u00d7 environment interaction had minor effects on proA content. Both additive and dominance gene action were significant for proA. The mean proportion of proA to total carotenoids (TC) was 44% among crtRB1-based hybrids, while 11% in traditional hybrids. BC was found to be positively correlated with BCX (r = 0.68) and proA (r = 0.98). However, no correlation was observed between proA and grain yield. Several hybrids with >10.0 t/ha grain yield with proA content >10.0 \u03bcg/g were identified. This is the first comprehensive study on development of diverse proA rich maize hybrids through marker-assisted pedigree breeding approach. The findings provides sustainable and cost-effective solution to alleviate vitamin-A deficiency.Malnutrition has emerged as one of the major health problems worldwide. Traditional yellow maize has low provitamin-A (proA) content and its genetic base in proA biofortification breeding program of subtropics is extremely narrow. To diversify the proA rich germplasm, 10 elite low proA inbreds were crossed with a proA rich donor (HP702-22) having mutant  DeficieVarious avenues namely, food-fortification, medical-supplementation and food-diversification are implemented to alleviate VAD ,7. HowevZea mays L.) is an important cereal crop grown in almost all parts of the world and cultivated across diverse climatic spectrum [CrtRB1 gene that codes for \u03b2-carotene hydroxylase is associated with higher accumulation of proA especially BC and BCX in maize. Rare natural variation in crtRB1 gene limits the hydroxylation of BC and BCX [crtRB1, while the TE is absent in mutant version [Maize  characterize the newly developed crtRB1-based inbreds using microsatellite markers, (iii) study combining ability of the crtRB1-based inbreds for different carotenoid fractions, and (iv) identify promising hybrids with high proA and grain yield.Diverse proA rich inbreds have been developed in the tropics \u201319. Howeermplasm ,20. Therviz., UMI-1200, UMI-1230, BML-6Q, BML-7Q, LM-11Q, LM-12Q, LM-13Q, LM-14Q, PDM-4341 and PDM-4251, possessing good general combining ability for greater yield but low in proA were selected as recipient parents. To incorporate favourable allele of crtRB1 into these inbreds, a CIMMYT-HarvestPlus bred inbred i.e., HP704-22 was used as donor. All these recipient inbreds represent a great extent of adaptation range as parental lines of released/promising hybrids while the donor HP704-22 has poor adaptation in Indian conditions (1s were raised and selfed during winter season (December-April) of 2015\u201316 at the Winter Nursery Centre (WNC), ICAR-Indian Institute of Maize Research (ICAR-IIMR), Hyderabad (17\u00b021\u00b450.39\u02baN and 78\u00b029\u00b442.31\u02baE). Ten F2 populations consisting of 99\u2013111 plants of each cross were then grown at ICAR-IARI, New Delhi during rainy season (2016)  .1s were tested for hybridity using crtRB1-based InDel marker present in 3\u2019UTR. The true F1s were selfed to derive F2 populations. Genomic DNA was extracted from three weeks old seedlings of F2 progenies using standard procedure of CTAB with minor modification [3\u2019TE-InDel-based marker for crtRB1 [viz., Forward (F): 5\u2019ACACCACATGGACAAGTTCG3\u2019, Reverse1 (R1):5\u2019ACACTCTGGCCCATGAACAC3\u2019 and Reverse2 (R2): 5\u2019ACAGCAATACAGGGGACCAG3\u2019. The primers were custom synthesized from Macrogen Inc., Seoul, South Korea. Polymerase chain reaction (PCR) protocol for crtRB1 [In-vitro amplification using ready-to-use master mix OnePCRTM  including Taq buffer, MgCl2, dNTPs and Taq Polymerase was used to perform PCR reaction in 96 well microtiter plate (M/s Genaxy) using GenePro thermal cycler (M/s Hangzhou Bioer Technology Co. Ltd.). The amplified product was resolved on 1.5% Seakem LE agarose gel .The Ffication . The quar crtRB1 , viz., Fr crtRB1  standard2 segregants with homozygous crtRB1 were selfed and derived F3 progenies were raised at ICAR-IARI, New Delhi during rainy season (2017). The F4s were raised during winter season (2017\u201318) at WNC, ICAR-IIMR, Hyderabad, and F5 progenies during rainy season (2018) at ICAR-IARI, New Delhi. To avoid chances of out-crossing, the presence of favourable allele of crtRB1 in each generation was validated using crtRB1-based InDel marker. Desirable segregants were advanced to further generation based on plant-, ear- and grain- characteristics.The FcrtRB1-based inbreds (MGU-PVMAS-1 to MGU-PVMAS-15) from 10 F2 populations were selected for molecular characterization  earlier developed by marker-assisted backcross breeding (MABB) at ICAR-IARI were also included. PMI-PV-1 and PMI-PV-2 are parental inbreds of India\u2019s first proA rich maize hybrid (Pusa Vivek QPM9 Improved). PMI-PV-5, PMI-PV-6, PMI-PV-7, PMI-PV-8 and PMI-PV-9 are the parents of MABB-derived proA rich hybrids . Further, HP465-41, a CIMMYT-derived proA rich inbred derived through marker-assisted pedigree breeding was also included for characterization. Two low proA elite inbreds  were also included as control for quality analysis. These 26 inbreds were planted in randomized complete block design (RCBD) at ICAR-IARI, New Delhi in rainy season of 2018. Each inbred was planted with two replications, in rows of 3 m with a plant-to-plant distance of 20 cm. The rows were spaced 75 cm apart. Recommended cultural practices were followed to raise a good experimental crop. To avoid contamination by foreign pollens, 2\u20133 plants in each row were selfed for estimation of carotenoids.Fifteen rization . The doncrtRB1 were used for molecular analysis using simple sequence repeats (SSRs) markers. DNA was isolated from seeds using standard sodium dodecyl sulphate (SDS) extraction protocol [http://www.maizegdb.org). PCR was carried out as per Choudhary et al. (2016) [Selfed seeds of 24 inbreds with favourable allele of protocol . A totalviz., gene diversity, major allele frequency, total number of alleles detected, heterozygosity and polymorphism information content (PIC) were estimated using PowerMarker v3.0 [For each allele, presence of a band in a genotype was indicated by 1 and absence of the band as 0. Five parameters, ker v3.0 . An alleker v3.0 . Principker v3.0 .crtRB1 and two inbreds with unfavourable allele of crtRB1) were extracted from maize endosperm through protocol of Kurilich and Juvik (1999) [30 column  and detected with a diode array detector-3000 (RS). The mobile phase comprised of methanol: tert-butyl methyl ether  with flow rate at 1 ml/min and peaks were detected at 450 nm. For each carotenoid component, viz., BC, BCX, LUT and ZEA, six dilutions of standards were used to construct the regression curve. To estimate proA concentration, amount of BC was added to one-half of BCX amount, while sum of LUT and ZEA gave the non-proA fractions [Carotenoids from the selfed seeds of 26 inbreds   with modractions . Total cractions .crtRB1-based inbreds developed under the current study were crossed with five crtRB1-based tester inbreds  as per line \u00d7 tester mating design [crtRB1-based promising inbred. These five testers belonged to two different groups viz., -A (PMI-PV-5 and HP465-41), -B . The 75 hybrid combinations and five commercial check hybrids  were evaluated using RCBD at three locations, viz. ICAR-IARI, New Delhi; CCS-HAU Regional Station, Uchani ; and ICAR-IARI Regional Research Centre, Dharwad  in rainy season of 2018. Each entry was evaluated in two replications, and was grown in a single row of 3 m length, with a row-to-row distance of 75 cm and plant-to-plant distance of 20 cm. Combining ability of the inbreds for carotenoids was calculated as per Singh and Choudhary (1985) [A set of 15 g design  at WNC, y (1985) .In each of the 75 experimental hybrid combinations generated from crosses, 2\u20133 plants were selfed to avoid contamination by the foreign pollen. The selfed seeds were used for estimation of carotenoids using UHPLC as per the procedure depicted for the inbreds.Grain yield (YLD) per plot was converted to t/ha as per the standard procedure. Magnitude of heterosis in hybrids over five commercial checks was estimated following Singh and Choudhary (1985) .The statistical analyses on ANOVA, correlation coefficients and combining ability were computed using Windostat 8.0.1s had both 296 and 543 bp amplicons. The 10 F2 populations were genotyped using crtRB1-specific 3\u2019TE-InDel-based marker  were 407 and 435, respectively. Thus, crtRB1 showed severe marker segregation distortion both cumulatively as well as in individual populations. Out of 201 favourable homozygotes, 75 segregants were selected based on ear- and grain- characteristics, and advanced to generate F3 progenies. Finally, 15 locally adapted F4 progenies  to 270 bp (bnlg1537). The average major allele frequency was 0.62 with a range from 0.29 (phi126) to 0.96 (umc2167 and umc1068). A set of 24 SSRs showed major allele frequency of \u22640.5. The average gene diversity was 0.49, ranging from 0.08 (umc2167 and umc1068) to 0.79 (bnlg1740) (umc2167 and umc1068) to 0.75 (bnlg1740) with an average of 0.43. Of the 77 SSRs, 29 loci had PIC \u22650.5. The current study also detected 14 unique alleles and 23 rare alleles. The heterozygosity existing among the inbreds varied from 0.00 to 0.25 with a mean of 0.04.Some loci such as umc1446 (0.25) and bnlg1346 (0.25) showed high heterozygosity  to 0.84 (MGU-PVMAS-13 and MGU-PVMAS-8) with mean of 0.67  . The PICzygosity . Genetic of 0.67 . Cluster of 0.67 . PCoA di of 0.67 .crtRB1-based inbreds and two check inbreds . Among 15 inbreds developed under the study, MGU-PVMAS-11 (14.63 \u03bcg/g), MGU-PVMAS-5 (13.09 \u03bcg/g), MGU-PVMAS-4 (12.53 \u03bcg/g), MGU-PVMAS-12 (12.52 \u03bcg/g) and MGU-PVMAS-2 (12.28 \u03bcg/g) were the most promising ones (crtRB1-based inbreds had significantly low mean LUT (12.16 \u03bcg/g), ZEA (5.86 \u03bcg/g) and non-proA (18.02 \u03bcg/g) compared to check inbreds . However,TC was nearly comparable among the crtRB1-based (31.31 \u03bcg/g) and check inbreds (36.34 \u03bcg/g).ANOVA revealed significant variation for BC, BCX, proA, LUT, ZEA, non-proA and total carotenoid (TC) among 24  inbreds . The meaectively , Fig 4. ing ones . Six inbcrtRB1-based inbreds, BC and BCX contributed 28% and 15% of TC, while LUT and ZEA contributed 39% and 18%, respectively. The contribution of BC and BCX to TC was only 6% and 3% in check inbreds, while the same for LUT and ZEA was 58% and 33%, respectively.Among crtRB1-based hybrids had significant variation for BC, BCX, proA, LUT, ZEA, non-proA and TC (\u00d7 environment (G \u00d7 E) interaction was also low with 8% (non-proA) to 22% (BCX).Pooled ANOVA revealed that A and TC . The proThe mean BC and BCX among experimental hybrids were 8.02 \u03bcg/g and 4.69 \u03bcg/g, as compared to 2.36 \u03bcg/g (BC) and 1.53 \u03bcg/g (BCX) among low-proA checks . High-prThe experimental crosses possessed low LUT  and ZEA  relative to low-proA checks . High-proA checks also possessed low LUT (5.22 \u03bcg/g) and ZEA (10.56 \u03bcg/g). The non-proA fraction among experimental hybrids varied from 10.78 to 23.50 \u03bcg/g, with an average of 16.47 \u03bcg/g. The non-proA fraction in low-proA checks was higher (31.88 \u03bcg/g) than that of high-proA checks (15.78 \u03bcg/g). TC in experimental hybrids varied from 21.92 to 37.86 \u03bcg/g, with a mean of 29.18 \u03bcg/g. Low-proA checks showed a mean of 35.77 \u03bcg/g, while high-proA commercial checks showed a mean of 29.80 \u03bcg/g .In case of experimental hybrids, the contribution of BC and BCX towards TC was 27% and 17%, while LUT and ZEA contributed 38% and 18%, respectively . In caseCorrelation analysis revealed that BC was positively correlated with BCX (r = 0.68**). Whereas, BC and BCX were negatively correlated with LUT  and ZEA , respectively. LUT and ZEA, however, were positively correlated (r = 0.70**). ProA and non-proA carotenoids were also negatively correlated (r = -0.57**). BC, BCX and proA were not correlated with TC, while LUT (r = 0.72**), ZEA (r = 0.72**) and non-proA (r = 0.78**) showed strong positive association with TC. The present study revealed that kernel carotenoid components and grain yield exhibited non-significant relationships (r = -0.18 to 0.15) except for ZEA (r = 0.29**).Pooled ANOVA revealed that environment was highly significant for all the carotenoids except for ZEA . Line efThe proportion of additive and dominance variance, and the contribution of lines, testers and line \u00d7 testers for pooled dataset are presented in GCA effect for proA varied from -1.52 to 1.27. Nine lines and one tester showed significant positive GCA effects for proA . Among tat par with CoMH-08-292. Among high proA checks, \u2018Pusa HQPM-5 Improved\u2019 emerged as the best check with 8.6 t/ha. A total of 26 experimental hybrids were significantly better than \u2018Pusa HQPM-5 Improved\u2019 for grain yield. These hybrids showed heterosis of 8.04% to 31.63% over the best high proA check.Heterosis of the 75 experimental hybrids was estimated over commercial low-proA and high proA checks. CMH08-292 and DHM-121 were medium maturing hybrids with low-proA. \u2018Pusa Vivek QPM-9 Improved\u2019 is an early maturing proA rich hybrid and was released as country\u2019s first proA rich maize hybrid during 2017. \u2018Pusa HQPM-5 Improved\u2019 and \u2018Pusa HQPM-7 Improved\u2019 are the medium maturing proA rich hybrids released in 2020. All the experimental hybrids matured in 92\u2013100 days, thus had medium maturity. The mean grain yield among experimental hybrids was 8.9 t/ha, with the highest yield of 11.4 t/ha  Tables. crtRB1 significantly enhances proA in maize kernel [crtRB1 in the Indian maize germplasm is quite low (3.38%) [crtRB1 inbreds have been developed in India. Thus, targeted breeding approach for selection of crtRB1 is essential for broadening the genetic base of proA rich maize germplasm [Plant based-food is the major source of nutrition especially in developing world. Traditional yellow maize lacks the required level of proA . The mute kernel . Diversee kernel ,19. Howe (3.38%) . So far,ermplasm .2 populations indicated that crtRB1 did not segregate as per the expected 1:2:1. This observation drives strength from earlier results of Lu et al. (2002) [crtRB1 gene could be precisely selected due to the reliable gene-based marker. In case of linked marker, there is always chance of selection of false positive individuals due to crossing over between the gene and marker [crtRB1-based inbreds using marker-assisted pedigree breeding. Earlier, Muthusamy et al (2014) [crtRB1 into the elite inbreds using MABB approach. In case of MABB, the improved lines are genetically similar to the recurrent parents except for gene under introgression [crtRB1-based inbreds were developed from F2 populations, the genetic makeup remains novel leading to development of new and diverse crtRB1-based inbreds. The MAS-based selection of crtRB1 is quite cost-effective, as selection of genotypes for high proA using UHPLC involves US$30\u201335 per sample. On the contrary, PCR marker-based selection of crtRB1 employed here costs only US$0.5\u20131.0 per sample. Molecular breeding is now a preferred choice among the maize breeders to develop proA rich germplasm [Genotyping of F. 2002)  and Babu. (2002) . The seg. (2002) . Since, d marker . The prel (2014) , Liu et l (2014) , Zunjarel (2014)  and Goswl (2014)  have intgression . In the 002  and Knowledge of the genetic relationship among inbreds is essential for efficient exploitation in breeding programme. Molecular markers have been successfully employed to derive the genetic distance in maize . High dicrtRB1-based inbreds and hybrids recorded nearly 3\u20134 folds more proA (8\u201314 \u03bcg/g) over the traditional checks (2\u20133 \u03bcg/g). Muthusamy et al., (2014) [crtRB1-based maize genotypes, respectively. However, traditional yellow maize possesses low concentration of proA (<2.5 ppm) [crtRB1, while low proA genotypes harbour the wild-type allele [The , 2014)  and Zunj, (2014)  also rep14  and Z2.5 ppm) . The acce allele ,17.phytoene synthase 1 (psy1) or Yellow 1 (Y1) gene condenses two geranyl-geranyl pyrophosphate molecules into one molecule of phytoene [y1 allele is unable to catalyze the reactions and white grains are formed due to no synthesis of carotenoids. However, when the Y1 is functional, crtRB1 causes hydroxylation of BC and BCX into ZEA. CrtRB1 located on chromosome 10 codes for \u03b2-carotene hydroxylase. The mutant version of crtRB1 drastically slows down the conversion, leading to more accumulation of proA carotenoids [crtRB1 allele, a large variation in proA was observed. This could be due to variation in the activity of other key genes such as psy1, lycopene \u03b2-cyclase (lcyB), lycopene \u03b5-cyclase (lcyE), phytoene desaturase (pds) and \u03b6-carotene desaturase (zds) catalyzing the carotenoid biosynthesis [lcyE present on chromosome 8 has been observed [lcyE along with crtRB1 is beneficial in enhancing proA in maize. Besides, several modifier loci/QTLs alone or in combination with other pathway genes could also influence the accumulation of proA in maize [In the carotenoid biosynthesis pathway, phytoene . The mutotenoids . Though ynthesis . Allelicobserved . Zunjareobserved  reportedin maize ,48.crtRB1-based and check- genotypes, thereby, suggesting greater flux of lycopene towards \u03b1-branch than the \u03b2-branch of pathway [crtRB1-based genotypes, the conversion of BC to ZEA is partially blocked, leading to higher proportion of BC next to LUT. ProA content among the crtRB1-based genotypes constituted 43\u201344% of TC as against 9\u201311% in the check genotypes. The non-proA component was less (56\u201357%) among crtRB1-based genotypes compared to 89\u201391% in checks [Among carotenoids, proportion of LUT was higher in both  pathway . LUT ser pathway . Howevern checks ,49. In sn checks .BC, BCX and proA showed strong positive correlations, as BCX is produced from BC, while both contribute to proA . Since, crtRB1 possesses much higher proA compared to heterozygote [crtRB1 in all hybrids. Since, all the hybrids were homozygous for favourable allele of crtRB1, other modifier loci could be the reason for dominance effects for further influencing the proA. Several lines including testers were identified as the best general combiners for proA. MGUH-57 (14.90 \u03bcg/g), MGUH-52 (14.60 \u03bcg/g), MGUH-27 (14.36 \u03bcg/g) and MGUH-32 (14.26 \u03bcg/g) possessed high proA, and had both the parents being high in GCA effects as well. Besides, MGUH-1, MGUH-14, MGUH-15, MGUH-18, MGUH-19, MGUH-28, MGUH-31, MGUH-48, MGUH-50 and MGUH-75 had one of the parents (used as line) having high GCA effects for proA. The inbreds with high GCA for proA thus serve as a promising inbreds in the future breeding programme [The current study revealed that environments had minor effect on BC, BCX and proA. Muthusamy et al. (2015b)  also reprozygote ,17,22. Crogramme .at par with CoMH08-292  and significantly better than DHM-121  for grain yield potential.Several promising experimental hybrids with >10.0 t/ha grain yield and >10.0 \u03bcg/g proA were identified. MGUH-15 , MGUH-50 , MGUH-72 , MGUH-54  and MGUH-55  were the most promising combinations. Most of these promising hybrids belongs to different clusters. These hybrids are much higher yielding than the \u2018Pusa Vivek QPM-9 Improved\u2019 , the first proA rich hybrid released in India. This is primarily due to extra-early maturity of \u2018Pusa Vivek QPM-9 Improved\u2019. Besides, two of the proA hybrids, \u2018Pusa HQPM5 Improved\u2019  and \u2018Pusa HQPM7 Improved\u2019  in the medium maturity groups have been recently released. Thus, these identified hybrids are higher yielding than the proA check hybrids as well. These selected hybrids also possessed higher proA than the normal best check (CoMH08-292 and DHM-121), but were crtRB1-donor was only 1.4t/ha, thereby suggesting its poor adaptability in subtropical conditions. The high grain yield of new crtRB1-based inbreds depicts better adaptability. The high yielding proA rich hybrids identified here would thus provide more productivity and profit to the farmers, and offer higher vitamin-A to the consumers [Further, parents of these proA rich hybrids also produce high yield from seed production perspective. The average grain yield among parental inbreds (used as lines) developed under this programme was 3.1t/ha with a range of 2.6\u20133.5t/ha. The same in onsumers . The procrtRB1-based inbreds have been developed in the study through marker-assisted pedigree breeding. These new inbreds possessed significantly higher proA than the checks. Molecular characterization of the inbreds depicted their diverse genetic nature. Genetic analysis revealed that both additive and non-additive variances were important. The crtRB1-based inbreds were successfully used in development of proA rich hybrids. Promising high yielding hybrids with very high concentration of proA have been identified. These proA rich hybrids are higher yielding than the exiting proA checks and at par with the normal checks in grain yield but with higher proA. The present study demonstrated the successful application of markers-assisted pedigree breeding in broadening the genetic base, and developing promising hybrids with higher grain yield as well as improved nutritional quality.Diverse S1 Tableopaque2 versions, TNAU: Tamil Nadu Agricultural University; ANGRAU: Acharya N. G. Ranga Agricultural University; PAU: Punjab Agricultural University.Q: Represents the (DOC)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table1. MGU-PVMAS-13, 2. PMI-PV-8, 3. PMI-PV-7, 4. MGU-PVMAS-6, 5. MGU-PVMAS-5, 6. MGU-PVMAS-11, 7. MGU-PVMAS-9, 8. MGU-PVMAS-4, 9. MGU-PVMAS-8, 10. MGU-PVMAS-15, 11. MGU-PVMAS-14, 12. MGU-PVMAS-7, 13. HP465-41, 14. PMI-PV-1, 15. MGU-PVMAS-3, 16. MGU-PVMAS-2, 17. MGU-PVMAS-12, 18. MGU-PVMAS-10, 19. HP704-22, 20. PMI-PV-6, 21. PMI-PV-9, 22. PMI-PV-5, 23. PMI-PV-2, 24. MGU-PVMAS-1, Inb. = Inbred.(DOCX)Click here for additional data file.S4 TableBC: \u03b2- Carotene, BCX: \u03b2-Cryptoxanthin, ProA: Provitamin A, LUT: Lutein, ZEA: Zeaxanthin, Non-ProA: Non-Provitamin A, TC: Total carotenoids, YLD: Yield.(DOCX)Click here for additional data file.S5 Table*Significant at p = 0.05; **Significant at p = 0.01, BC: \u03b2- Carotene, BCX: \u03b2-Cryptoxanthin, ProA: Provitamin-A, LUT: Lutein, ZEA: Zeaxanthin, Non-ProA: Non-Provitamin-A, TC: Total carotenoids, SE: Standard Error.(DOCX)Click here for additional data file.S6 Table*Significant at p = 0.05; **Significant at p = 0.01.(DOCX)Click here for additional data file.S1 File(DOCX)Click here for additional data file."}
+{"text": "Coronavirus disease-2019 (COVID-19) is now an unprecedented worldwide pandemic. However, there are no specific antiviral drugs available for its treatment. A handful of studies have summarized convalescent plasma (CP) transfusion in severe or critical cases ,4,5,6, w9/L with 78% neutrophils, 9.2% lymphocytes, 2.3% basophils, 0.4% eosinophils, and 10.1% monocytes; hemoglobin of 51 g/L; platelet count of 79x109/L; high-sensitivity C-reactive protein of 57.43 mg/L; and interleukin-6 level of 59.25 pg/mL. The real-time polymerase chain reaction (RT-PCR) assay of the throat swab was positive for severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection. A chest CT obtained on February 21 revealed bilateral ground-glass opacities primarily distributed along the pleura  starting in November 2017, but a drug-related hematologic adverse event occurred quickly. As a result, dasatinib (150 mg/day) was given instead. She had not achieved complete hematological remission at the diagnosis of COVID-19 due to poor responses to these therapies. At admission , the most relevant clinical findings included white blood cell count of 4.93x10e pleura  and 1d. e pleura  that expe pleura . RT-PCR e pleura  accordin2) oscillating between 90% and 93%, after receiving conventional antiviral therapy including arbidol (200 mg three times daily), oseltamivir (75 mg twice daily), ribavirin (500 mg every 12 hours), and interferon-alpha-2b inhalation (5 million units twice daily). A follow-up chest CT scan showed increased consolidation and extended opacities . Three repetitive RT-PCR tests were negative from the 6th to 8th day after CP transfusion. Chest images showed absorption of opacities within 10 days (th\u00a0day of admission.The patient developed worsening hypoxemia, with oxyhemoglobin saturation (SaOpacities  and 1e.  10 days  and 1f. A recent study showed a 10% case rate of COVID-19 among 128 patients with hematological cancers in Wuhan . The tre"}
+{"text": "Tepidicella baoligensis strain B18-50T, isolated from a high-temperature oil well in Baolige Oilfield, China. The estimated genome is 2.87\u2009Mb, with 2,653 protein-coding sequences.We report the draft genome sequence of  Tepidicella baoligensis strain B18-50T, isolated from a high-temperature oil well in Baolige Oilfield, China. The estimated genome is 2.87\u2009Mb, with 2,653 protein-coding sequences.We report the draft genome sequence of  Tepidicella species are slightly thermophilic Gram-negative rods that are distributed in geothermal areas. These organisms do not assimilate carbohydrates or polyols but use organic acids as carbon and energy sources and oxidize thiosulfate to sulfate in the presence of an assimilable carbon source  in China. The liquid sample was diluted with sterilized enrichment medium (EN), plated onto EN agar, and incubated at 50\u00b0C , and 1.0 g liter\u22121 tryptone (Oxoid). Genomic DNA was prepared from an overnight culture by using the phenol-chloroform extraction method (Strain B18-50 at 50\u00b0C . A pure 2 medium  supplemehttps://github.com/aquaskyline/SOAPdenovo2)  libraries were prepared from the sheared fragments using the NEXTflex rapid DNA-Seq kit  and sequenced in 150-bp paired-end mode using the Illumina HiSeq X Ten platform at Majorbio Bio-Pharm Technology, Inc. . The sequencing generated 3,449,944 pairs of raw reads totaling 1,041,883,088\u2009bp, giving approximately 362\u00d7 coverage. The reads were quality trimmed with Trimmomatic v0.36 (http://ccb.jhu.edu/software/glimmer/index.shtml) (http://trna.ucsc.edu/software/) (https://github.com/tseemann/barrnap) was used for rRNA prediction. The predicted CDSs were annotated from the nonredundant (NR), Swiss-Prot, Pfam, GO, COG, and KEGG databases using the sequence alignment tools BLAST+ v2.3.0   was usedftware/)  was used+ v2.3.0 , Diamond+ v2.3.0 , and HMM+ v2.3.0 . A totalJACCDB000000000; the raw reads are available under SRA accession number SRR12179937. This announcement represents the first version of the genome.This whole-genome shotgun sequencing project has been deposited in DDBJ/ENA/GenBank under the accession number"}
+{"text": "Orthobunyavirus, family Peribunyaviridae) is an arthropod-borne virus that causes congenital abnormalities in ruminants. Here, we report the complete genome sequences of two AKAV strains causing nonsuppurative encephalomyelitis in cattle by postnatal infection in Japan.Akabane virus (AKAV)  is an arthropod-borne virus that causes congenital abnormalities in ruminants. Here, we report the complete genome sequences of two AKAV strains causing nonsuppurative encephalomyelitis in cattle by postnatal infection in Japan.Akabane virus (AKAV) (genus  Akabane orthobunyavirus . Since it was first isolated from mosquitos in Gunma, Japan, in 1959, it has been widely detected in Australia, Asia, and Africa (Culicoides spp.) and causes spontaneous abortions, stillbirths, and congenital malformations in pregnant ruminants (Akabane virus (AKAV) is an enveloped, tripartite, negative-sense RNA virus of the species AKAV strains are clustered into four genogroups based on phylogenetic analysis of their medium (M) segments , 2. Genohttps://jorgensen.biology.utah.edu/wayned/ape), with the OBE-1 strain (the genogroup II strain that is the progenitor of the vaccine strain used in Japan) as the reference .Both virus strains were propagated in HmLu-1 cells. Their genomic RNA was extracted using the QIAamp virus RNA minikit . Reverse transcription-PCR was performed using SuperScript IV reverse transcriptase  and PrimeSTAR GXL DNA polymerase . Primer sequences are available on request. The 5\u02b9- and 3\u02b9-terminal sequences of all of the genomic RNA segments were determined using 5\u2032 and 3\u2032 rapid amplification of cDNA ends (RACE) . The seqThe full-length S, M, and L segments of KM-2/Br/06 are 856, 4,309, and 6,869 nucleotides (nt), respectively, whereas those of FI-1/Br/06 are 867, 4,308, and 6,867\u2009nt, respectively. The GC contents of the segments were also determined . The nucThe sequences determined in this study will be useful in identifying the virulence factors implicated in bovine postnatal encephalomyelitis.LC552047.1 (KM-2/Br/06 S segment), LC552048.1 (KM-2/Br/06 M segment), LC552049.1 (KM-2/Br/06 L segment), LC552050.1 (FI-1/Br/08 S segment), LC552051.1 (FI-1/Br/08 M segment), and LC552052.1 (FI-1/Br/08 L segment).The sequences are available in GenBank under the accession numbers"}
+{"text": "Correction to: BMC Rheumatol 4, 4 (2020)https://doi.org/10.1186/s41927-019-0105-4Following publication of the original article , the autThe original article has been updated.AbstractResults: In those without DM GC use had a 4.4-fold increased all-cause mortality RR : 3.77 to 5.07) compared to non-use, whilst those with DM had a lower RR for GC use ). However, those with DM had a higher RD associated with GC use because of their higher baseline risk. In those with DM, GC use was associated with an additional 44.9 deaths/1000 person-years (pyrs) (95% CI: 32.9 to 56.8) compared to non-use, while in those without DM GC use was associated with an additional 34.4 deaths/1000 pyrs (95% CI: 30.1 to 38.7). A similar pattern was seen for CV mortality. The adjusted Cox proportional hazards model showed no evidence of multiplicative interaction, but additive interaction indicated a non-significant increased risk. For CV mortality there was no interaction on either scale.In the \u201cAll-cause mortality\u201d sub-section it now correctly reads \u201cDuring follow-up there were 1005 deaths\u201d rather than 1002 deaths."}
+{"text": "Background: Cancer treatments induce symptoms/signs superimposing on individual patient's clinical status, determining heterogenous toxicity syndromes (TS). We reviewed intensive first line triplet chemotherapy-based regimens in metastatic gastro-intestinal cancers (mGI), based on FIr/FOx schedule, fluorouracil and weekly alternating irinotecan/oxaliplatin, to point out limiting TS (LTS) relevance.Methods: Metastatic colo-rectal (mCRC), pancreatic ductal adenocarcinoma (mPDAC), gastric carcinoma (mGC) patients were enrolled by careful decision-making including age, performance status (PS), and comorbidity status in real life phase II studies: FIr-B/FOx adding bevacizumab (B) overall, FIr-C/FOx-C adding cetuximab (C) in KRAS/NRAS wild-type mCRC; FIr/FOx in mPDAC; FD/FOx adding docetaxel (D) in mGC. Toxicity, individual LTS, LT alone  or associated to other limiting/G2 toxicities  were evaluated, compared by chi-square test. In FIr-C/FOx-C, 5-fluorouracil/irinotecan pharmacogenomic biomarkers, 5-fluorouracil degradation rate (5-FUDR), SNPs ABCB1, CYP3A4, DYPD, UGT1A1 were evaluated, related with LTS.Results: FIr-B/FOx, FIr-C/FOx-C in mCRC, FIr/FOx in mPDAC, FD/FOx in mGC, showed activity, efficacy, toxicities similar to reported triplet regimens. LTS: mCRC FIr-B/FOx 44%, LTS-ms 24%, LTS-ss 20%, in young-elderly 46%, LTS-ms significantly increased vs. LTS-ss; FIr-C/FOx-C 65.5%, significantly increased LTS-ms vs. LTS-ss, in young-elderly 83%; mPDAC FIr/FOx 27.5%, mostly LTS-ms, in young-elderly 38.4% all LTS-ms; mGC FD/FOx 30%, all LTS-ms, in young-elderly 25%. Reduced FUDR, SNPs CYP3A4, UGT1A1, >1 positive pharmacogenomic biomarkers were prevalent in patients with gastrointestinal LTS.Conclusions: LTS is an innovative clinical parameter of toxicity burden, differential treatment-related TS in individual patient. LTS can evaluate pharmacogenomic biomarkers predictive relevance to select mGI patients fit for intensive treatments, at risk of limiting gastrointestinal toxicity.Trial Registrations: The trials were registered at Osservatorio Nazionale sulla Sperimentazione Clinica dei Medicinali (OsSC) Agenzia Italiana del Farmaco (AIFA) Numero EudraCT 2007-004946-34, and Osservatorio Nazionale sulla Sperimentazione Clinica dei Medicinali (OsSC) Agenzia Italiana del Farmaco (AIFA) Numero EudraCT 2009- 016793-32. Addition of more drugs in a chemotherapy combination requires the design of proper schedule and doses, to provide the balance between projected/received (>80%) dose intensity (DI) and treatment-related toxicity . This clThe conventional evaluation of toxicity of cancer treatments depends upon grading of the clinical relevance of each toxicity symptom and sign, by evaluation of the type, prevalence and intensity of toxicity, more specifically of limiting G3-4) toxicity, related to the administered treatment regimen, and determined by National Cancer Institute Common Toxicity Criteria . Thus, safety profile and toxicities induced by cancer treatments are conventionally defined, according to the number of administered cycles and treated patients, as severe (G3-4), moderate G2), mild (G1), or absent. This evaluation does not individually describe the toxicity burden experienced in a single patient, defining a spectrum of toxicity syndromes (TS). To better evaluate individual toxicity of intensive treatments, we introduced the concept of Limiting Toxicity Syndromes (LTS), consisting of at least a limiting toxicity alone  or associated to other limiting or G2-3 non-limiting toxicities   66.7%, progression-free survival (PFS) 12 months, and overall survival (OS) 20 months as first line mCRC treatment. Thus, we developed intensive first line regimens, based on FIr/FOx triplet chemotherapy schedule: FIr-B/FOx (We previously developed doublet chemotherapy schedule of 12 h (10 p.m. to 10 a.m.) timed-flat infusion (TFI) 5-FU , associaIr-B/FOx , respectIr-B/FOx ; FD/FOx Ir-B/FOx .Patients treated with triplet chemotherapy-based regimens were enrolled by decision-making process including age, performance status (PS), and comorbidity status evaluated by Cumulative Illness Rating Scale (CIRS) . CIRS stin label for treatment in Italian public hospitals, and published in Gazzetta Ufficiale Repubblica Italiana . MCRC clinical trials were approved by Local Ethical Committee , and conducted in accordance with Declaration of Helsinki. PDAC study was approved by the Regional Review Board . All patients provided written, informed consent concerning the proposed treatment, and biological evaluations.Proposed regimens were approved by Agenzia Italiana del Farmaco for administration 2, days 1\u20132, 8\u20139, 15\u201316, 22\u201323; CPT-11 160 mg/m2 days 1,15; OXP 80 mg/m2, days 8, 22; every 4 weeks; B 5 mg/kg, days 1,15. In KRAS/NRAS wild-type mCRC, FIr-C/FOx-C . In mGC, FD/FOx  week week2, dschedule  11): TF: TF2, da, FD/FOx  14): TF: TF2, da6 cells/min, and Single Nucleotide Polimorphisms (SNPs) ABCB1 , CYP3A4 , DYPD1 , UGT1A1 (28) were preliminarily related with the occurrence of LTS . To better evaluate toxicity of intensive treatments in the individual patient, LTS, consisting of at least a limiting toxicity alone (LTS-ss) or associated to other limiting or G2-3 non-limiting toxicities (LTS-ms) were evaluated , 14, 20.e of LTS , 25.2, respectively  per cycle for all the drugs at the recommended doses were \u226580% (2/w); CPT-11 76% (45.5 mg/m2/w); OXP 66% (26.5 mg/m2/w); C 71% (178.5 mg/m2/w); yE patients showed significantly worse OS compared to non-elderly (P 0.045) . . 2, respP = 0.022). In mPDAC patients treated with FIr/FOx, median rDI per cycle were: 5-FU 70.4% (1268.5 mg/m2/w); CPT-11 70% (56 mg/m2/w); OXP 72.5% (29 mg/m2/w). In yE, 5-FU 83.3%, CPT-11 80%, OXP 85%. Overall, 17% discontinued FIr/FOx treatment due to LT. Limiting cumulative G3-5 toxicities were: diarrhea 17%, asthenia 14%, neutropenia 17%, mucositis 6%, hypokaliemia 7%, hypertransaminasemia 7%, nausea/vomiting, hypoalbuminemia, anemia, thrombocytopenia 3%, respectively. One case of toxic death (3%) was observed.In mPDAC patients, FIr/FOx showed ORR 53%, PFS 4 months, OS 11 months; among yE/old-elderly patients, median PFS 4 months, and median OS 5 months ; D 91% (22.75 mg/m2/week); OXP 83.8% (33.5 mg/m2/week). Cumulative G3-4 toxicities were represented by: asthenia 20%, neutropenia 50%, leucopenia 20%, mucositis and hypoalbuminemia 10%.In mGC patients, FD/FOx at recommended 5-FU 1000 mg/m7 months . Median KRAS/NRAS wild-type mCRC patients treated with FIr-B/FOx, overall LTS were observed in 22 patients (44%) (P 0.05). LTS were prevalently characterized by G2-3 diarrhea, 9 patients (69.2%), 8 LTS-ms and 1 LTS-ss. The 2 LTS-ms with double LT were observed in yE patients. The 10 LTS-ms, defined by LT added to other, at least G2, non-limiting toxicities, were prevalently determined by G2-3 diarrhea (90%), plus G2-3 nausea/vomiting (70%). The 10 LTS-ss were prevalently characterized by G3 diarrhea (50%), G3 asthenia, hypertension, hypertransaminasemy, neutropenia, thrombocytopenia 10%, respectively.In ts (44%) : LTS-ms ts (44%) . LTS-ms KRAS/NRAS wild-type mCRC patients treated with FIr-C/FOx-C, LTS were observed in 19 patients (65.5%), 5 out of 6 yE (83%): LTS-ms 17 (59%), LTS-ss 2 (7%) (P 0.006). LTS-ms characterized by \u22652 LT 7 (24%); LT associated to other toxicities 10 (34%). LTS were prevalently characterized by G3-4 diarrhea and G3 asthenia associated to other toxicities.In s 2 (7%) . LTS werIn mPDAC patients treated with FIr/FOx, overall LTS were 8 (27.5%); 5 out of 13 yE/old-elderly (38.4%); LTS-ms 7 (24.1%), LTS-ss 1 (3.4%) . LTS-ms CYP3A4, UGT1A1, and >1 positive pharmacogenomic biomarkers. Specifically, the exploratory analysis of 5-FUDR reduction and ABCB1, CYP3A4, DYPD, UGT1A1 SNPs evaluated in 14 KRAS/NRAS wild-type mCRC patients (48.3%) treated with FIr-C/FOx-C .Furthermore, the analysis of SNPs represented an exploratory analysis in colorectal cancer patients treated with intensive triplet chemotherapy plus cetuximab, according to FIr-C/FOx-C schedule, to better evaluate the safety profile, particularly gastrointestinal. Patients who reported gastrointestinal LTS, prevalently showed reduced fluorouracil degradation rate (FUDR), Single Nucleotide Polimorphisms (SNP) -C/FOx-C , 25, andOver the past 10 years, we developed intensive triplet chemotherapy-based regimens in fit mGI cancer patients , 11, 14,KRAS/NRAS wild-type mCRC , to obtain the description of cumulative and prevalent limiting (G3-G4), moderate (G2), mild (G1), or absent toxicities directly determined by the cancer treatment, according to the number of administered cycles and treated patients. This way does not really represent the clinical burden of toxicity in the individual patient, nor define prevalent individual TS affecting a patient population equivalently treated and their variability. Thus, in reported intensive first line triplet chemotherapy-based regimens developed in mGI cancer patients, we added the description of individual TS, and specifically of LTS, to describe cumulative and individual toxicity, that include differential spectrum and intensities of TS, depending from medical treatment and patients' individual clinical conditions.The integration of the evaluation of LTS to the conventional treatment-related toxicity, contributed a patient-related clinical indicator of toxicity burden, providing a global evaluation of patient-related limiting toxicity , 14, 20:In yE patients, LTS evaluation showed the differential tolerability of intensive triplet chemotherapy based regimens: in mCRC, FIr-B/FOx LTS 46%, but significantly prevalent LTS-ms , 27; FIrKRAS/NRAS wild-type MCRC patients treated with FIr-C/FOx-C showed that reduced 5-FUDR, and CYP3A4 and UGT1A1 SNPs may predict individual LTS occurrence, particularly at recommended doses, specifically gastrointestinal LTS  and in 9 who showed LTS (47.4%), exploratory data of pharmacogenomic biomarkers compared with LTS occurrence in Thus, LTS meets the need of an innovative clinical parameter of patient-related toxicity burden, to measure personalized safety of intensive first line triplet chemotherapy-based regimens proposed in mGI, also associated to targeted agents (B or C). Its clinical relevance should be prospectively evaluated as a model in clinical practice. The integration of LTS and conventional toxicity evaluations may help proper selection of patients fit for intensive first line medical treatments in mGI cancers, to more properly weigh toxicity analysis with activity and clinical outcome and its contribution to address selection of patients suitable for intensive triplet chemotherapy-based regimens. The equivalent, integrated evaluation and monitoring of individual safety by LTS, can help properly select first line intensive medical treatment, and safely administer and manage an intensive first line regimen, that could guarantee increased clinical outcomes and good safety profile in real life.In the era of precision oncology, integrating molecular characterization of cancer affecting the individual patient to specifically address targeted treatments, such as in mCRC \u201332, the Analysis of TS could be integrated in the therapeutic pathway of cancer patients in clinical practice and in clinical studies to globally evaluate tolerability of cancer treatments. It could be, also, particularly useful for a more proper evaluation of tolerability in the therapeutic pathway of individual cancer patients unfit for standard treatments, due to elderly status and/or their clinical status, or unfit for intensive regimens , 34, to More, in intensive regimens such as FIr-C/FOx-C, characterized by high discontinuation rate of treatment, highly prevalent (>50%) LTS and heterogeneity of LT, LTS may help directly relate individual patient- and drugs-related toxicity with pharmacogenomics biomarkers referred to the individual genetic identity of the cancer patient.TS, specifically LTS, represents an innovative clinical parameter of cumulative and individual patient-related toxicity burden, defining differential spectrum and intensities of treatment-related TS, depending from the clinical status of the individual cancer patient, particularly according to elderly, and PS.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation, to any qualified researcher.Proposed regimens were approved by Agenzia Italiana del Farmaco for administration in label for treatment in Italian public hospitals, and published in Gazzetta Ufficiale Repubblica Italiana . MCRC clinical trials were approved by Local Ethical Committee , and conducted in accordance with Declaration of Helsinki. PDAC study was approved by the Regional Review Board . All patients provided written, informed consent concerning the proposed treatment, and biological evaluations.GB contributed in conceptualization, data curation, formal analysis, investigation, methodology, project administration, validation, and writing original draft. ER contributed in conceptualization, data curation, formal analysis, investigation, methodology, project administration, supervision, validation, and writing\u2014review original draft.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Populus trichocarpa) upon infection with the generalist root pathogen Phytophthora cactorum (Oomycetes). P. cactorum infection led to an induced accumulation of terpenes, aromatic compounds, and fatty acids in poplar roots. Transcriptome analysis of uninfected and P. cactorum-infected roots revealed a terpene synthase gene PtTPS5 that was significantly induced upon pathogen infection. PtTPS5 had been previously reported as a sesquiterpene synthase producing two unidentified sesquiterpene alcohols as major products and hedycaryol as a minor product. Using heterologous expression in Escherichia coli, enzyme assays with deuterium-labeled substrates, and NMR analysis of reaction products, we could identify the major PtTPS5 products as -guaia-4(15)-en-11-ol and -guaia-4-en-11-ol, with the former being a novel compound. The transcript accumulation of PtTPS5 in uninfected and P. cactorum-infected poplar roots matched the accumulation of -guaia-4(15)-en-11-ol, -guaia-4-en-11-ol, and hedycaryol in this tissue, suggesting that PtTPS5 likely contributes to the pathogen-induced formation of these compounds in planta.Pathogen infection often leads to the enhanced formation of specialized plant metabolites that act as defensive barriers against microbial attackers. In this study, we investigated the formation of potential defense compounds in roots of the Western balsam poplar ( Plants are constantly under attack from a multitude of pests, including pathogens and herbivores. Such biotic stresses often induce the formation of specialized plant metabolites that play major roles in plant defense. Terpenoids represent the largest class of natural compounds, and to date, more than 200,000 terpenoids are known, of which ~40,000 can be produced by plants . Beside 10), -farnesyl diphosphate , and -geranylgeranyl diphosphate . The prenyl diphosphates are substrates for terpene synthases (TPS), which catalyze the formation of the basic mono-(C10), sesqui-(C15), and diterpene (C20) skeletons D20 = +47.8 . HRMS (EI): m/z = 222.1978 . GC (HP5-MS): I = 1660. MS : m/z (%) = 222 (0.3), 204 (29), 189 (24), 175 (2), 161 (24), 149 (22), 133 (11), 121 (19), 107 (31), 91 (41), 81 (54), 67 (25), 59 (100), 53(13), 41 (35). IR (diamond ATR): /cm\u20131 = 2953 (m), 2923 (s), 2854 (m), 1714 (w), 1650 (w), 1456 (m), 1376 (m), 1260 (m), 1094 (s), 1020 (s), 873 (m), 800 (s).-Guaia-4-en-11-ol (2). Yield: 0.6 mg . TLC (pentane/ether = 4:1): Rf = 0.21. Optical rotation: [\u03b1]D20 = +21.7 . HRMS (EI): m/z = 222.1975 . GC (HP5-MS): I = 1661. MS : m/z (%) = 222 (4), 204 (81), 189 (69), 175 (7), 161 (75), 147 (27), 133 (29), 119 (37), 105 (61), 91 (81), 79 (68), 67 (30), 59 (100), 51(5), 41 (55). IR (diamond ATR): /cm\u20131 = 2954 (s), 2923 (s), 2854 (s), 1723 (w), 1670 (w), 1459 (m), 1376 (m), 1260 (w), 1096 (w), 1025 (w), 800 (w).S,5S,7R,10R)-guaia-4(15)-en-11-ol and -guaia-4-en-11-ol. For the reactions with DMAPP (1 mg in 1 mL water) and (Z)-IPP or (E)-IPP (1 mg in 1 mL water), protein preparations of PtTPS5 (1 mL) and FPPS  and incubation buffer (6 mL) were added. For the reactions with (R)-IPP or (S)-IPP (1 mg in 1 mL water), protein preparations of PtTPS5 (1 mL), FPPS  and incubation buffer (6 mL) were added. After incubation with shaking at 28 \u00b0C overnight, the reaction mixtures were extracted with C6D6, the extracts were dried with MgSO4 and analyzed by NMR and GC-MS.Isotopic labelling experiments were performed to determine the absolute configurations of , MgCl2 S , IDI , MgCl2 Throughout the manuscript, data are presented as means \u00b1 SE. Statistical analysis was performed with SigmaPlot 11.0 for Windows  and is described in the figure and table legends for the respective experiments. Whenever necessary, the data were log transformed to meet statistical assumptions such as normality and homogeneity of variances.Raw reads of the RNAseq experiment were deposited in the NCBI Sequence Read Archive (SRA) under the BioProject accession PRJNA660564 \u2018Oomycete-induced changes in the root transcriptome of poplar\u2019."}
+{"text": "T (Collection de Souches de l\u2019Unit\u00e9 des Rickettsie CSUR P6417\u2009=\u2009Colecci\u00f3n Espa\u00f1ola de Cultivos Tipo CECT 9771) is a new Pedobacter species isolated from the planarian Schmidtea mediterranea. Schmidtea mediterranea are flatworms living in freshwater and exhibiting an unusual ability to regenerate amputated parts. To date, the gut microbiota of Schmidtea mediterranea remains poorly studied. Here, via the culturomics strategy that consists in using diversified culture conditions, we isolated a new bacterium, strain EG, that we characterized using the taxono-genomics approach that combines phenotypic assays and genome sequencing and analysis. Strain EG exhibits a 16S rRNA sequence similarity of 98.29% with Pedobacter nyackensis strain NWG-II14T, its closest neighbour with standing in nomenclature. It is an aerobic bacterium belonging to the family Sphingobacteriaceae. Colonies are small, round, smooth and transparent. Bacterial cells are Gram-negative, rod-shaped, motile and non-spore-forming bacilli with positive catalase and oxidase activities. The genome sequence is 6,198,518 bp\u2013long with a G\u2009+\u2009C content of 41.13%, and the Ortho-ANI and dDDH values when compared to P. nyackensis are 77.34% and 21.50%, respectively. Strain EGT exhibits unique characteristics that classify it as the type strain of new bacterial species for which we propose the name Pedobacter schmidteae sp. nov.Pedobacter schmidteae sp. nov. strain EG Schmidtea mediterranea is an invertebrate living in environmental water. This flatworm is used as a model of development, because of its extraordinary abilty to regenerate after amputation, because of his high contents in stem cells known as neoblasts1. It has been shown that S. mediterranea is an excellent model to investigate host-pathogen relationships3, notably in the context of human pathogens. To date, the gut microbiota of S. mediterranea remains poorly studied5. Using the microbial culturomics approach6, we investigated the S. mediterranea microbiota. Culturomics is a concept in which diversified culture conditions are used to enable isolation of a maximum of bacterial species from the human microbiota10. During this analysis, we isolated a bacterial strain from S. mediterranea that could not be identified using Matrix Assisted Laser Desorption-Ionisation Time of Flight-Mass Spectrometry (MALDI-TOF-MS). We used the taxono-genomics strategy that combines phenotypic assays and genome sequencing to further characterize this bacterium14. This enabled us to describe a new bacterial strain that exhibited enough genetic and phenotypic differences with closely related bacteria. We propose it as a new species named Pedobacter schmidteae sp. nov.Schmidtea mediterranea asexual clonal line ClW42 is a laboratory planarian strain that has been preserved in our laboratory for the past 10 years, by cutting animals in tree fragments each month. S. mediterranea were kept in the dark, in filtered tap water, at 19\u2009\u00b0C without antibiotics. The animals were fed once per week with homogenized calf liver and were starved for at least two weeks prior to studying them. Filtered water was obtained using a device consisting of two 0.2\u2009\u00b5m filters, one containing charcoal and ceramics , and the other being a 0.20\u2009\u00b5m membrane . Filtered water was checked for sterility prior to be used for planarian culture.S. mediterranea were washed in filter-sterilized water and then one ground worm was inoculated in Buffered Charcoal Yeast Extract (BCYE) , Luria Bertani (LB) and 5% sheep blood-enriched Columbia agar . All inoculated media were incubated at 19, 28 and 37\u2009\u00b0C. Each individual bacterial colony was harvested and identified by MALDI-TOF-MS 15. The obtained spectra were imported into the MALDI Biotyper 3.0 software  and analysed against the reference spectra of bacteria included in the database (Bruker database constantly updated with the Mediterranee-Infection database (http://www.mediterranee-infection.com/article.php?larub=280&titre=urms-database). The MALDI Biotyper RTC software was used to interpret the results according to the obtained score values: a colony was likely identified at the species level for a score > 2.0, probably identified for a score between 1.99 and 1.7, but not identified for a score <1.7.Following two weeks of starvation, https://www.codoncode.com/). For taxonomic assignation, a BLASTn search was performed against the nr database16. A sequence similarity threshold of 98.65% by comparison with the phylogenetically closest species with standing in nomenclature was used to delineate a putative new species17. Phylogenetic relationships were inferred from the comparison of 16S rRNA sequences using the MEGA7 software18.Bacterial colonies that were not identified at the species level using MALDI-TOF MS were further tested using 16S rRNA sequencing. Genomic DNA was extracted using an EZ1 automate and the DNA tissue kit . The complete 16S rRNA gene amplification and sequencing was performed using eight primers on an ABI Prism 3130xl Genetic Analyzer capillary sequencer . The primers used were Fd1 (5\u2032-AGAGTTTGATCCTGGCTCAG-3\u2032), Rp2 (5\u2032-ACGGCTACCTTGTTACGACTT-3\u2032), F536 (5\u2032-CAGCAGCCGCGGTAATAC-3\u2032), R536 (5\u2032-GTATTACCGCGGCTGCTG-3\u2032), F800 (5\u2032-ATTAGATACCCTGGTAG-3\u2032), R800 (5\u2032-CTACCAGGGTATCTAAT-3\u2032), F1050 (5\u2032-TGTCGTCAGCTCGTG-3\u2032) and R1050 (5\u2032-CACGAGCTGACGACA-3\u2032) . The CodonCode Aligner software was used for sequence alignment, assembly and correction (strain EG and P. nyackensis strain NWG-II14T (DSM19625) was attempted at various growth temperatures  in 5% sheep blood-enriched Columbia agar (bioM\u00e9rieux) under anaerobic, aerobic and microaerophilic atmospheres using GasPak\u2122 EZ generators . Sporulation was tested by thermal shock, which consists in exposing bacteria to a temperature of 80\u2009\u00b0C for 30\u2009minutes and then monitoring their growth for 4 days. Various salinity  and pH  conditions were tested. Gram staining and motility from fresh colonies were observed using a DM1000 photonic microscope  with a 40 \u00d7 objective lens. Catalase and oxidase activities were tested by using a BBL DrySlide according to the manufacturer\u2019s instructions . The size of bacterial cells was measured using transmission electron microscopy. API strips  were used to study the biochemical characteristics of the strains. Bacterial susceptibility to benzylpenicillin, amoxicillin, ampicillin, ceftriaxone, imipenem, ciprofloxacin, amikacin, gentamicin, streptomycin, daptomycin, doxycycline, metronidazole, rifampicin, and vancomycin was assessed using E-tests and a 0.5 McFarland concentration of strains EG and NWG-II14T. Cellular fatty acid methyl ester (FAME) analysis was performed by GC/MS. Two samples were prepared with approximately 120\u2009mg of bacterial biomass per tube harvested from several culture plates. Fatty acid methyl esters were prepared as described by Sasser30 and GC/MS analysis was carried out as previously described31. Briefly, FAMES were separated using an Elite 5-MS column and monitored by mass spectrometry . Spectral database search was performed using MS Search 2.0 operated with the following Standard Reference Database 1\u2009A  and FAMEs mass spectral database .Culture of strain 32. Briefly, sequencing was performed using the Mate-Pair strategy and a Miseq sequencer . A concentration of 1.5\u2009\u03bcg of gDNA prepared following the Nextera Mate-Pair Illumina guide was used to prepare the Mate-Pair library. The gDNA was simultaneously fragmented and tagged using a Mate-Pair junction adapter. Then the fragmentation pattern was validated using a DNA 7500 labchip on a BioAnalyzer . The size of the DNA fragments ranged from 1.5\u2009kb to 11\u2009kb. No size selection was performed and 662\u2009ng of labelled fragments were circularized. Next, circularized DNA was mechanically sheared using a Covaris device S2  into small fragments with an optimal size at 1200\u2009bp. The library profile was analysed on a High Sensitivity Bioanalyzer LabChip (Agilent Technologies) and the concentration library was measured at 61.4 nmol/l. Then, the library was loaded on the sequencer after a denaturation step and a dilution at 15 pM. Automated cluster generation and sequencing were performed in a single 39-h run in a 2\u2009\u00d7\u2009251-bp and sequencing reads were assembled using the A5 pipeline. Genomic annotation was obtained using the Prokka software. A search for virulence factors was performed by comparaison with the VFDB database (http://www.mgc.ac.cn/VFs/) using BLASTn34.The bacterial genomic DNA (gDNA) of strain EG was extracted using an EZ1 automate and the DNA tissue kit  and then was quantified using a Qubit assay  at 82.6\u2009ng/\u03bcL. The bacterial gDNA was prepared and sequenced as previously describedPedobacter africanus strain DSM 12126\u2009T (NZ_FWXT00000000.1), P. antarcticus strain 4BYT (NZ_JNFF00000000.1), P. ginsengisoli strain T01R-27 (NZ_CP024091.1), P. heparinus strain DSM 2366\u2009T (NC_013061.1), P. nutrimenti strain DSM 27372 (NZ_QKLU00000000.1), P. nyackensis strain DSM 19625 (NZ_FWYB00000000.1), P. panaciterrae strain 048 (NZ_LGEL00000000.1) and P. steynii strain DX4 (NZ_CP017141.1). Degrees of genomic similarity between strain EG and compared genome were evaluated using the GGDC (http://ggdc.dsmz.de/ggdc.php#)16 and Orthologous Average Nucleotide Identity (Ortho-ANI) (https://www.ezbiocloud.net/tools/orthoani)35 softwares.The genome from strain EG was compared to those of Pedobacter nyackensis strain NWG-II14T with which it exhibited a sequence similarity of 98.29%. When compared to other Pedobacter species, strain EG exhibited 16S rRNA similarity values of 97.84%, 97.69%, 97.63%, 97.93%, 97.43%, 97.28%, 97.33%, 97.62%, 97.93%, 97.32%, 97.32% and 97.0% with P. heparinus strain DSM 2366\u2009T, P. steynii strain WB2.3\u201345\u2009T, P. metabolipauper strain WB2.3\u201371\u2009T, P. nutrimenti strain J22T, P. duraquae strain WB2.1\u201325\u2009T, P. africanus strain NBRC 100065\u2009T, P. panaciterrae strain Gsoil 042\u2009T, P. ginsengisoli strain Gsoil 104\u2009T, P. seoulensis strain THG-G12T, P. trunci strain THG-DN3.19\u2009T, P. humi strain THG S15\u20132T and P. antarcticus strain 4BYT\u200940, respectively .Strain EG was isolated on 5% sheep blood-enriched Columbia agar (bioM\u00e9rieux) after 2 days at 28\u2009\u00b0C in aerobic atmosphere at pH 7. The 16S rRNA-based phylogenetic tree demonstrated that strain EG was most closely related to S. mediterranea revealed the presence of strain EG in 11 S. mediterranea sp. nov. Strain EG is a member of the family Sphingobacteriaceae within the phylum Bacteroidetes(Table\u00a01).The analysis of 13 , rod-shaped, motile, and non-spore-forming bacilli. Using the Image J software, their mean length and width were 1.98\u2009\u00b5m and 0.69\u2009\u00b5m, respectively , without any flagellum. For the two strains EG and P. nyackensis NWG-II14T, no growth was obtained in anaerobic or microaerophilic conditions. Both strains grew at temperatures ranging from 6 to 30\u2009\u00b0C in aerobic atmosphere at pH values ranging from 5.5 to 9; the strains also grew at salinity concentrations lower than 25\u2009g/L. Catalase and oxidase activities were positive for both strains.After 4 days of culture on blood-enriched Columbia agar, colonies from strain EG were small (0.4\u2009mm of diameter), transparent, round with a convex shape and smooth. Bacterial cells were Gram-negative Fig.\u00a0, rod-shaely Fig.\u00a0, withoutP. nyackensis strain NWG-II14T were susceptible to benzylpenicillin, ampicillin, ceftriaxone, imipenem, gentamicin, doxycycline and rifampicin (Table\u00a0). Both strains were resistant to amoxicillin, amikacin, daptomycin, metronidazole and vancomycin. Pedobacter nyackensis, but not P. schmidteae, was susceptible to ciprofloxacin and streptomycin.Strain EG and in Table\u00a0. Both stP. nyackensis NWG-II14T, strain EG differed by exhibing of Esterase (C4), Esterase lipase (C8), Valine arylamidase, \u03b2-galactosidase and \u03b2-glucosidase activities (Table\u00a0). By comparaison with all other tested species, strain EG differed in production of esterase (C4) and \u03b2-glucosidase.Positive and negative reactions obtained using API 50CH, API 20NE, API Zym, and API 20E strips are presented in Table\u00a0es Table\u00a0. By comp(Table\u00a0). 15-methyl-Hexadecenoic acid, 14-Methylpentadec-9-enoic acid is detected only in strain EGT. In contrast to P. nyackensis NWG-II14T, the strain EG have in its membrane composition the 3-hydroxy-Hexadecanoic acid, 13-methyl-Tetradecanoic acid, 3-hydroxy-13-methyl-Tetradecanoic acid, 3-hydroxy-15-methyl-Hexadecanoic acid, 15-methyl-Hexadecenoic acid and 14-Methylpentadec-9-enoic acid; and for the absence of Dodecanoic acid, 3-hydroxy-8-methyl-nonanoic acid, 9-Heptadecenoic acid, 9-Octadecenoic acid.The analysis of the composition in fatty acid of the strain EG revealed that the major fatty acids were 13-methyl-tetradecanoic acid 49.8%), 9-Hexadecenoic acid (27.7%) and 3-hydroxy-15-methyl-Hexadecanoic acid (7.5%). Other fatty acids included 15-methyl-Hexadecenoic acid (3.6%), 14-Methylpentadec-9-enoic acid (3.4%), Hexadecanoic acid (3.3%), 3-hydroxy-13-methyl-Tetradecanoic acid (1.9%), Tetradecanoic acid (1.4%) and 3-hydroxy-Hexadecanoic acid (1.0%). Minor amounts (<1%) of the fatty acids were also detected as Pentadecanoic acid and 9,12-Octadecadienoic acid 9.8%, 9-H\u22123) and a score of 52 with the GacS [GacS/GacA two-component system]42 sensor histidine kinase/response regulator in Pseudomonas putida strain KT2440 according to the VFDB database. It has been reported that the two-component system signaling pathways are the major signaling mechanisms in bacteria and as well in Archaea43 to monitor external and internal stimuli 44. This pathway is also found in simple eukaryota and higher plants45. In opportunistic bacterial pathogens, the use of two-components systems is required to regulate the expression of genes necessary for the transition from the environmental reservoir to the host46. Digital DNA-DNA hybridization values (dDDH) obtained using the GGDC software are reported in Table\u00a0P. nutrimenti to 21.50% with P. nyackensis. Such values were lower than the 70% threshold recognized to delineate distinct species. Similarly, Ortho-ANI values  ranged from 70.98% with P. nutrimenti to 74.65% with P. nyackensis, which is lower than the 95% threshold used to discriminate bacterial species. Thus, we could confirm that strain EG belonged to a separate Pedobacter species for which we propose the name Pedobacter schmidteae sp. nov.The genome sequence from strain EG was assembled into one contig of 6,198,518\u2009bp with a G\u2009+\u2009C content of 41.13%. We identified a total of 5,012 predicted protein-coding genes, in addition to 3 complete rRNA operons, 52 tRNAs and 1 tmRNA. Comparison of these genomic data with those from of closely related species is presented in Table\u00a0P. schmidteae sp. nov. Interestingly, P. schmidteae sp. nov. is present in 84.6% of S. mediterranea worms tested. To best of our knowledge, P. schmidteae sp. nov has never been identified anywhere else than in S. mediterranea. Indeed, no nucleotide sequence linked to this new strain has been found in the nr database (BLASTn https://blast.ncbi.nlm.nih.gov). To date, we assume that P. schmidteae sp. nov is unique to S. mediterranea, but we cannot exclude formally that it might be present in other living organisms of the environment.Using the taxono-genomic approach, we concluded that strain EG is the representative strain of the new species 47. Pedobacter schmidteae . The bacterium belongs to the family Sphingobacteriaceae within the phylum Bacteroidetes. The type strain EGT (CSUR P6417\u2009=\u2009CECT9771) was isolated on 5% sheep blood-enriched Columbia agar after 2 days at 28\u2009\u00b0C in aerobic atmosphere at pH 7 from the microbiota of the planarian S. mediterranea. Colonies are small, round, smooth, transparent and convex. Bacterial cells are Gram-negative, rod-shaped, motile and non-spore-forming bacilli with positive catalase and oxidase activities. The main cellular fatty acids detected are 13-methyl-tetradecanoic acid (49.8%), 9-Hexadecenoic acid (27.7%), 3-hydroxy-15-methyl-Hexadecanoic acid (7.5%), 15-methyl-Hexadecenoic acid (3.6%), 14-Methylpentadec-9-enoic acid (3.4%), Hexadecanoic acid (3.3%), 3-hydroxy-13-methyl-Tetradecanoic acid (1.9%), Tetradecanoic acid (1.4%) and 3-hydroxy-Hexadecanoic acid (1.0%). We found traces (<1%) of Pentadecanoic acid and 9,12-Octadecadienoic acid.The protologue is to standardize the format of descriptions of new taxa, supported by the Judicial Commission of the International Committee on Systematic BacteriologyT exhibits positive reactions for catalase, oxidase, alkaline phosphatase, esterase (C4), esterase lipase (C8), leucine arylamidase, valine arylamidase, acid phosphatase, Naphtol-AS-BI-phosphohydrolase, \u03b2-galactosidase, \u03b1-glucosidase, \u03b2-glucosidase, N-acetyl-\u03b2-glucosaminidase, esculin ferric citrate, glucose, D-mannose, N-Acetyl-glucosamine and D-maltose, but negative reaction for lipase (C14), cystine arylamidase, trypsin, \u03b1-chymotrypsin, \u03b1-galactosidase, \u03b2-glucuronidase, \u03b1-mannosidase, \u03b1-fucosidase.Using an APIZYM strip, strain EGT is unable to metabolize glycerol, starch, amygdalin, arbutin, D-adonitol, D-arabinose, D-arabitol, D-cellobiose, D-fructose, D-fucose, D-galactose, D-glucose, D-lactose, D-melezitose, D-melibiose, D-raffinose, D-ribose, D-saccharose, D-sorbitol, D-tagalose, D-trehalose, D-turanose, dulcitol, D-xylose, erythritol, gentiobiose, glycogen, inositol, inulin, L-arabinose, L-arabitol, L-fucose, L-rhamnose, L-sorbose, L-xylose, methyl-\u03b1D-Glucopyranoside, methyl-\u03b1D-Mannopyranoside, methyl-\u03b2D-xylopyranoside, potassium 2-CetoGluconate, potassium 5-Cetogluconate, potassium gluconate, salicin, xylitol.Using a API 50CH strip, strain EGT use potassium nitrate, L-tryptophane, L-arginine, urea, gelatin, D-mannitol, malic acid, phenylacetic acid, L-lysin, trinatriumcitrate, L-ormithin, Esculin ferric citrate, Glucose, D-mannose, N-Acetyl-glucosamine and D-maltose.In addition, strain EGT is 6,198,518 bp\u2013long with a G\u2009+\u2009C content of 41.13%.The genome of strain EGThe 16S rRNA gene and genome sequence are deposited in GenBank under accession numbers LS453293 and LS999839, respectively.Pedobacter schmidteae strain EGT is available at http://www.mediterranee-infection.com/article.php?laref=936The MALDI-TOF MS spectrum of T is TA00631 in the http://imedea.uib-csic.es/dprotologue/ website.The 16S rRNA gene sequence and genome sequence were deposited in GenBank under accession numbers LS453293 and LS999839, respectively. The Digital Protologue database Taxonumber for strain EGT  was deposited in the CSUR (Collection de Souches de l\u2019Unit\u00e9 des Rickettsies WDCM 875) and CSCT (Colecci\u00f3n Espa\u00f1ola de Cultivos Tipo) strain collections under numbers CSUR P6417 and CECT9771, respectively.Strain EG"}
+{"text": "Escherichia coli strain Tj, isolated from the Varzob River in Tajikistan, is presented. This strain possesses four prophage elements related to Shigella phage SfV, E. coli O157:H7-specific phage \u03d5V10, lambdoid phage HK225, and coliphage Ayreon. It contains a gene encoding a hemolysin E toxin.The 4.6-Mbp draft genome sequence of  Escherichia coli strain Tj, isolated from the Varzob River in Tajikistan, is presented. This strain possesses four prophage elements related to Shigella phage SfV, E. coli O157:H7-specific phage \u03d5V10, lambdoid phage HK225, and coliphage Ayreon. It contains a gene encoding a hemolysin E toxin.The 4.6-Mbp draft genome sequence of  Escherichia coli (Migula 1895) Castellani and Chalmers 1919 is a Gram-negative, facultatively anaerobic, enteric bacterium that inhabits the intestines of warm-blooded animals and humans and frequently contaminates environmental waters and food. E. coli strain Tj was isolated from the Varzob River in Tajikistan, during a laboratory course at the Center of Biotechnology of the Tajik National University, on Endo LES agar plates at 44\u00b0C as a dark-red colony and confirmed as E. coli by its API 20E profile (5-0-4-4-5-5-2). The 16S rRNA gene sequence, obtained by PCR amplification and Sanger sequencing as described elsewhere . The genome was sequenced at Eurofins Genomics using a NEBNext Ultra II DNA library preparation kit and Illumina HiSeq 2500 paired-end sequencing technology with a read length of 2 \u00d7 150\u2009bp, yielding 6,416,190 reads and 1,924,857,000 sequenced bases. Reads with a maximum of 7 bases with a Phred score below 28 were initially discarded, and additional quality control procedures were performed using the Trim Reads tool in the CLC Genomics Workbench v. 8.5.1. Unless otherwise stated, all software was used with default values. Assembly was performed using the CLC de novo assembly tool, resulting in 4,627,784\u2009bp of unique sequence data distributed into 96 contigs with an N50 value of 111,402\u2009bp, coverage of 417\u00d7, and GC content of 50.8%. The draft genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) (https://www.ncbi.nlm.nih.gov/genome/annotation_prok). Genome completeness was estimated as 99.96% by CheckM v. 1.0.18  values of\u2009\u226597.0% (http://www.mgc.ac.cn/VFs) (E. coli phage Ayreon (P-1) (Shigella flexneri (P-2) (NC_019717), and phage \u03d5V10, which specifically infects E. coli O157:H7 (P-4)   using IsE. coli strain Tj have been deposited in DDBJ/ENA/GenBank under accession numbers MT920313 and GCA_007049865.1, respectively. The associated BioProject, SRA, and BioSample accession numbers are PRJNA506592, SRR12336902, and SAMN10464432, respectively.The partial 16S rRNA gene and whole-genome shotgun sequences of"}
+{"text": "In Russia, cancer is the second leading cause of death following cardiovascular diseases. Adoptive transfer of NK cells is a promising approach to fight cancer; however, for their successful use in cancer treatment, it is necessary to ensure their robust accumulation at tumor foci, provide resistance to the immunosuppressive tumor microenvironment, and to engineer them with higher cytotoxic activity. NK lymphocytes are known to kill cancer cells expressing a number of stress ligands; and the balance of signals from inhibitory and activating receptors on the surface of the NK cell determines whether a cytotoxic reaction is triggered. We hypothesized that stronger cytotoxicity of NK cells could be achieved via gene editing aimed at enhancing the activating signaling cascades and/or weakening the inhibitory ones, thereby shifting the balance of signals towards NK cell activation and target cell lysis. Here, we took advantage of the CRISPR/Cas9 system to introduce mutations in the coding sequence of the shp-2 (PTPN11) gene encoding the signaling molecule of inhibitory pathways in NK cells. These shp-2 knock-outNK cells were additionally transduced to express a chimeric antigen receptor (CAR) that selectively recognized the antigen of interest on the target cell surface and generated an activating signal. We demonstrate that the combination of shp-2 gene knockout and CAR expression increases the cytotoxicity of effector NK-like YT cells against human prostate cancer cell line Du-145 with ectopic expression of PSMA protein, which is specifically targeted by the CAR. Natural killer cells are cells whose primary function is toeliminate infected and transformed cells in the body. Cytotoxicactivity of NK cells is regulated by the balance of signalstriggered by the interaction of activating and inhibitory receptorswith appropriate ligands on the surface of target cells.Inhibitory receptors play an central role in NK cell biologysince most of their ligands are represented by Major HistocompatibilityComplex I (MHC-I) molecules, that are presenton the surface of nearly all healthy cells and frequently absenton cancer or infected cells . This ensures that normal cells are spared by NK cells,whereas the cells that have lost MHC-I expression becomeeliminated . Nevertheless,absence of MHC-I expression does not universallyguarantee that an NK cell would kill a target cell, as this activityis also heavily dependent on the presence of ligands toactivating receptors of NK cells . Inhibitory receptor signal is known to be mainlytransduced via SH2-containing inositol 5\u2032 polyphosphatase 1(SHIP-1) and/or tyrosine phosphatases Shp-1 and Shp-2. Thelatter two proteins dephosphorylate tyrosine residues withinthe ITAM motifs of activating receptors thereby aborting thesignaling cascade .NK cells are successfully used in therapy of oncologicaldiseases . Nevertheless, because tumorcells actively withstand elimination by establishing an immunosuppressivetumor microenvironment, the cytotoxicityof NK cells can be strongly affected . There are several waysof enhancing the cytotoxicity of NK cells. These includemodification of signaling pathways, altering the levels andrepertoire of NK cell receptors, or boosting the activity ofNK cells with appropriate cytokines . In this study, we explored whethermodification of the inhibitory signaling pathway through theshp-2 gene knockout in NK cells would result in the increaseof the cytotoxic activity of model NK cells.Cell lines and cell culture. YT, Du-145-PSMA, and HEK293Tcell lines were maintained in Iscove\u2019s Modified Dulbecco\u2019sMedium (IMDM) supplemented with 10 % fetal bovine serum(FBS), 100 U/ml penicillin, and 100 \u03bcg/ml streptomycin(Sigma) in a humidified atmosphere of 5 % CO2 at 37 \u00b0C.Plasmid construction. Two shp-2-specific sgRNAs wereidentified using publicly available bioinformatics tools . The following target sequences were selected: gtgcagatcctacctctgaaagg andacagtactacaactcaagcagg (PAM site underlined).The lentiCRISPRv2 vector provided by Prof. Feng Zhang  was usedfor co-delivery of sgRNAs and Cas9. Oligonucleotides correspondingto the selected targets were denatured in a boilingwater bath followed by slow renaturation and cloned into thelentiCRISPRv2 vector between BsmBI restriction sites.The plasmid DNA of clones lentiCRISPRv2-Shp2g1 andlentiCRISPRv2-Shp2g2 encoding shp2-specific sgRNA1 andsgRNA2, was mixed with the DNA of packaging plasmidspsPAX2 and pMD2.G (provided by Prof. D. Trono) in thefollowing weight ratio: 10:10:7.5:2.5, in the total amountof 3 \u03bcg. Using calcium phosphate transfection , the resulting plasmid DNA mix was delivered toHEK293T cells. Supernatants containing VSV-G pseudotypedlentiviral particles were collected 48 hours after transfection,filtered through 0.45 \u03bcm PES filters and used either fresh orstored at \u201370 \u00b0C.YT cell transduction. To improve transduction rate ofYT cells, a spinoculation protocol was used . YT cells were seeded in 24-well plates (1 \u00d7 105 cells) inthe presence of polybrene (8 \u03bcg/ml) followed by the additionof supernatants containing pseudotyped lentiviral particles.Cells were centrifuged at 500 g for 40 minutes at 32 \u00b0C andincubated in a CO2 incubator for 16 hours. The next day,the supernatant was replaced with a fresh culture medium.After 3 days, selection of transduced cells was performed byadding puromycin  to a final concentrationof 5 mg/ ml (1 week), with non-transduced cells serving ascontrols.Western blot analysis. For western blot analysis, transducedYT cells and control non-transduced cell lines YT-wtand HEK293T were lysed in lysis buffer . Cell lysateswere centrifuged, boiled in a water bath for 5 minutes,separated by SDS-PAGE in a 10 % polyacrylamide gel andtransferred onto a nitrocellulose membrane . After blocking in 3 % milk in PBST, the membrane wasincubated with rabbit monoclonal antibodies against Shp-2, followedby an HRP-labeled secondary goat-anti-rabbit antibody. Equal loading was controlled byusing hybridization with control antibodies against \u03b2-actin. Signal was detected usingan ECL-prime substrate in accordance with the manufacturer\u2019srecommendations  and AmershamImager 600 with an exposure time of 1 min.Flow cytometry. For phenotyping CAR-YTshp-2\u2013/\u2013 celllines, 105 cells were washed in PBS and incubated for 30 minuteswith the biotinylated protein L (3.3 \u03bcg/ml)  at a temperature of 4 \u00b0C. Then the cells werewashed and incubated with streptavidin-APC  in accordance with the manufacturer\u2019s recommendations.Vital dye 7AAD  was used to excludedead cells from the analysis. The samples were analyzed usingBD FACSCanto\u00ae II (Becton Dickinson and Company) andBD FACSDiva software.Real Time Cytotoxicity Assay (RTCA). Adherent targetcells (5 \u00d7 104 cells per well) were seeded in 8-well E-plates and left to grow for 16\u201318 hours.The next day, the medium was removed and replaced witha fresh one containing 105 effector cells. Cell growth wasmonitored for 24 hours using the RTCA iCELLigence system.Cytotoxicity was calculated using the formula: [CI (targetcells without effector cells) \u2013 CI (target cells with effectorcells) \u00d7 100/CI (target cells without effector cells)], where CIis the normalized impedance value in the wells .Flow cytometry-based cytotoxicity assay. Cells were labeledwith Cell Proliferation Dye eFluor 670 . 50,000 target cells per well were seeded into a 96-wellplate. Next, an equal number of effectors was added .Cells were incubated for 4 hours in a humidified atmosphereof 5 % CO2 at 37 \u00b0C. Then the vital dye 7AAD was added toeach well and the percentage of living target cells was measuredon a BD FACSCantoII flow cytometer.Statistical analysis  was performed using Prism software(GraphPad version 8.0). One-way analysis of variance (onewayANOVA) was used to detect the differences between theactivity of control cell lines that did not carry the shp-2 geneknockout and that of knockout cells. All data are presentedas mean \u00b1 standard error of the mean.In this study, we have chosen human immortalized cell lineYT that has an NK-like phenotype. These cells are known todisplay perforin-mediated target cell lysis, lack Fc receptor expressionand do not require conditioning of growth media withIL2 . To knock-out shp-2, CRISPR/Cas9 system was used.Two sgRNAs specific to the 3rd and 5th exons of the genewere cloned into a lentiviral vector lentiCRISPRv2 . The two resulting constructs lentiCRISPRv2-Shp2g1 and lentiCRISPRv2-Shp2g2 were used for producingVSV-G-pseudotyped lentiviral particles, which then wereused for transduction of YT-cells. Given that lentiCRISPRv2vector carries a puromycin resistance gene, transduced cellswere subjected to antibiotic selection and single-cell cloning. A panel of monoclonal derivatives of YT cells was analyzedby Sanger-sequencing the genomic fragments centered atsgRNA-binding sites in the shp-2 gene, in order to identifythe clones carrying biallelic mutations in this gene .Four clones  were obtained that displayedtruncated shp-2 sequence . Knock-outswere verified by western-blot analysis, which confirmed thatthese clones did not express a full-length Shp-2 protein .One way to re-target NK cell cytotoxicity is via expressionof chimeric antigen receptors (CARs). Even if the target celllacks typical stress markers recognized by endogenous NK cellreceptors, binding of a CAR to its cognate target may inducepro-activation signaling in a CAR-NK cell. To test whetherCAR-mediated cytotoxicity is enhanced upon shp2-knock out,we used our previously published second-generation CAR(scFv(J591)-CD8hinge-CD28TM-CD28-CD3z)  that is specific for PSMA, a common surfacemarker of prostate cancer cells . Wild-type and shp2-mutant YT cell lines weretransduced to express this CAR, which was verified by flowcytometry .shp-2\u2013/\u2013 cells exerted significantlyhigher cytotoxicity than CAR-YT cells with a functionalshp- 2 gene; no difference between the knock-out subcloneswas observed .Next, we compared the cytotoxic activity of the B1-CAR,C1-CAR, and C4-CAR cells vs CAR-YT cells against Du-145 prostate cancer cells engineered to ectopically expressPSMA. Du-145 cells have been reported to be resistant toNK cell killing , which establishes theman appropriate model for assaying possible enhancement ofNK cell-mediated cytotoxicity. We used iCELLigence platform for the cytotoxicity analysis.It was found that CAR-YTshp-2\u2013/\u2013 cells may displayunwanted cytotoxicity against healthy cells, cytotoxicity assaywas conducted with PBMCs isolated from a healthy donor. Weobserved that the percentage of dead normal cells did not differsignificantly between the wells where CAR-YTshp-2\u2013/\u2013 cells orno effector cells (control) were added . This was alsotrue for cultures, where normal cells were co-incubated witheither CAR-YT or CAR-YTshp-2\u2013/\u2013 cells.To understand whether CAR-YTCytotoxic activity of NK cells is known to be regulated bythe balance of activating and inhibitory receptor signals . Cytotoxic reaction occurswhen two conditions are met. Namely, if an activating signalis present, (i. e. when the target cell expresses the ligand(s) tothe activating receptors) and if the negative signaling is sufficientlyreduced or absent  . Whenever positive signalingis absent or negative signaling outweighing the positive signaling,cytotoxic reaction will not be launched. In the contextof NK cell therapy, this translates to the failure to eliminatethe tumor cells .Absence of activating signal can be effectively solved byengineering CAR expression in NK cells, an approach alreadyactively used in practice . Nonetheless,tumor cells overexpressing the ligands of inhibitoryNK cell receptors may still remain protected from CAR-NKcell mediated lysis .One way to address this problem is to create NK cells withknock-outs of inhibitory receptors. Although technicallyfeasible, this may not be as straightforward, considering thelarge number of these receptors. In our opinion, knockingout Shp-2, a single \u201chub\u201d of NK cell signaling appears as aviable alternative . Our experiments show that YT cell line derivativeslacking SHP2 expression display stronger CAR-mediatedcytotoxicity towards NK-resistant cell line Du-145-PSMA, unlike CAR-YT cells . Thisis likely attributableto the fact that Du-145-PSMA cellshave a high density of inhibitory ligands, which results in thesuppression of activating signaling in NK cells, even uponCAR engagement. Indeed, it was reported that Du-154 cellsexpress low levels of ligands to the activating NK receptorsNKp30 and NKp46, while MHC-I expression is very high.shp-2\u2013/\u2013 cells are safe, testing ona wide panel of different healthy cell types obtained frommultiple donors may be required, which is a challenging taskfrom a technical and ethical points of view. In the currentstudy, we have measured cytotoxicity of CAR-YTshp-2\u2013/\u2013 cellstowards the lymphocytes of a healthy donor, and no significantdifference with negative control was observed. This suggeststhat reducing the inhibitory signaling in NK cells likely posesno threat provided that no strong activating signals are available.However, to address this important safety concern a morecomprehensive study is indeed required. We envisage that arepresentative and diverse panel iPS-derived normal humancell types can be obtained to test for the possible off-targetactivity of shp2-mutant YT cells.Safety of such combination is yet to be found, as it is possiblethat this may lead to increased cytotoxicity towardshealthy tissues, as a result of weakening of inhibitory signaling.To ensure that the YTThus, the best option for enhancing cytotoxicity of NK cellsand the YT cell line in particular is to induce activating signalsusing CARs in combination with the suppression of inhibitorysignaling via knocking out its key mediator, the SHP-2protein. In the future it is necessary to comprehensivelystudy the safety of modified lymphocytes, in particular, tostudy their cytotoxic activity against a wider range of healthytissues.The authors declare no conflict of interest.Becker P.S.A., Suck G., Nowakowska P., Ullrich E., Seifried E.,Bader P., Tonn T., Seidl C. 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DOI 10.4049/jimmunol.170.9.4539."}
+{"text": "Egglestonichthysfulmensp. nov. (Teleostei: Gobiidae) is described on the basis of a single specimen (21.7 mm in standard length) collected from 250 m depth off Okinawa Island, Ryukyu Islands, Japan. The new species is characterized by the following combination of characters: anal-fin rays I, 9; pectoral-fin rays 17, lower rays not free from membrane; longitudinal scale series 25; transverse scales 8; pre-dorsal-fin scale rows 8; cheek and opercle naked; pelvic frenum absent; caudal fin lanceolate, its length 32.2% of SL; interorbital width very narrow, 1.2% of HL (much narrower than pupil diameter); no spicules or odontoid processes on outer surface of gill arches; and body whitish, upper half with broken zigzag pattern of bright yellow patches and associated scattered black melanophores in fresh specimens (melanophores retained in preserved specimens). Several characters, including pectoral-fin ray count, interorbital width, and coloration uniquely distinguish the new species from congeners. Egglestonichthys Miller & Wongrat, 1979. Although a generic reassessment of the genus Egglestonichthys is needed , Egglestonichthyspatriciae Miller & Wongrat, 1979, Egglestonichthysrubidus Allen, Erdmann & Brooks, 2020, and Egglestonichthysulbubunitj Larson, 2013]   Fig. ; no spicer) Fig. .SL (% of HL in parentheses): head length 30.3; head depth 16.7 (55.1); head width 18.3 (60.6); body depth 14.9; body width 12.9; caudal-peduncle length 19.1, depth 9.4; snout length 5.9 (19.3); eye diameter 7.3 (24.0); interorbital width 0.4 (1.2); upper-jaw length 11.8 (39.0); pectoral-fin length 31.0; pelvic-fin length 28.1; caudal-fin length 32.2; and longest dorsal-fin spine (2nd) length 12.8.Dorsal-fin rays VI + I, 10; anal-fin rays I, 9; pectoral-fin rays 17; segmented caudal-fin rays 16; caudal-fin ray pattern 9/7; branched caudal-fin rays 7/5; unsegmented (procurrent) caudal-fin rays 6/6; longitudinal series scales 25; transverse scales 8; pre-dorsal-fin scale rows 8 ; circumpeduncular scales 12; gill rakers on outer face of first arch 3 + 11 (counted on right side); vertebrae 10 + 16. The following morphometrics are expressed as percentage of Body slender, compressed, width much less than depth Fig. . Anus siHead lacking sensory canal pores Fig. . Head seBody covered with relatively large cycloid scales. Pre-dorsal- and pelvic-fin regions covered with cycloid scales; anterior margin of pre-dorsal-fin scales reaching to vertical through preopercle margin; pre-pelvic-fin scales reaching to just behind anteroventral point of gill opening. Pectoral-fin base with 2 relatively large cycloid scales. Side of head  naked , last rays well separated from caudal-fin base. Pectoral fin long, pointed, middle rays longest, tips reaching to below origin of 2nd dorsal-fin soft ray; all rays connected by membrane, uppermost and lower 3 rays unbranched. Pelvic fins completely connected by membrane, without frenum; posterior tip reaching below 2nd dorsal-fin origin when appressed; pelvic-fin origin located just below ventral end of pectoral-fin base. Caudal fin lanceolate, its length much greater than head length.First dorsal fin triangular, 2Fresh coloration . Scattered black melanophores throughout bright yellow patches on head and body. Dorsal-fin rays whitish with yellow interspaces (possibly barred in undamaged specimens). Other fins whitish; upper end of pectoral-fin base with yellow spot; caudal-fin base with faint yellow vertical bar and broad rectangular patch of melanophores.ion Fig. . Body whColoration when preserved , neither could be confirmed in the present specimen. A detailed investigation of species currently assigned to Egglestonichthys must precede any redefinition of the genus .The holotype and only known example of the new species lacks head sensory pores, but has a transverse sensory papillae pattern on the cheek and a wide gill opening , thereby matching the diagnostic characters of aracters . Althougenus see : 153. NeEgglestonichthysfulmen is unlikely to be misidentified as one of its congeners, having the following characters: 17 pectoral-fin rays ; interorbit very narrow (width 1.2% of HL), much less than pupil diameter [vs. variously broad: 38.5\u201347.6% HL in E.bombylios (described as 4.9\u20135.9 in HL in Larson and Hoese 1996); 17.0\u201320.4% HL in E.melanoptera (4.9\u20135.9 in HL in Larson and Hoese 1996); equal to pupil diameter in E.patriciae .E.fulmen and E.melanoptera both lack a pelvic frenum, the former has much lower scale counts than the latter . E.ulbubunitj; all are connected by membrane in E.fulmen. In addition, E.fulmen has fewer anal-fin rays . Egglestonichthysfulmen also has fewer longitudinal series scales than E.bombylios and E.patriciae , a lanceolate caudal fin (length 32.2% of SL), and scales absent on the cheek and opercle .Although"}
+{"text": "Mycobacterium smegmatis mc2155. While the Awesomesauce genome is 57,054 bp with 94 protein-coding genes, the LastJedi genome is 55,149 bp with 94 protein-coding genes. Nucleotide sequence comparison in Phamerator detected synteny between Awesomesauce gp49 to gp61 and singleton LilSpotty. Whole-genome BLASTn alignments revealed that LastJedi strongly resembles Clifton (99.41% identity).Bacteriophages Awesomesauce and LastJedi infect  Mycobacterium smegmatis mc2155. While the Awesomesauce genome is 57,054 bp with 94 protein-coding genes, the LastJedi genome is 55,149 bp with 94 protein-coding genes. Nucleotide sequence comparison in Phamerator detected synteny between Awesomesauce gp49 to gp61 and singleton LilSpotty. Whole-genome BLASTn alignments revealed that LastJedi strongly resembles Clifton (99.41% identity).Bacteriophages Awesomesauce and LastJedi infect  Mycobacterium smegmatis mc2155, a prominent acid-fast Gram-positive soil bacterium. Awesomesauce inhabited damp, shaded soil at Brown University , while LastJedi was from mud near shrubs at the State University of New York College at Old Westbury . Both were isolated (with enrichment at 23 to 24\u00b0C), purified, and amplified in M. smegmatis mc2155, as described in the SEA-PHAGES Discovery Guide (https://seaphagesphagediscoveryguide.helpdocsonline.com/home). The bacteriophages form clear plaques (\u223c1 mm), exhibit Siphoviridae morphology (images are available at https://phagesdb.org/phages/Awesomesauce and https://phagesdb.org/phages/LastJedi), and are F cluster/F1 subcluster members (Bacteriophage Awesomesauce and LastJedi genomes were examined in collaboration with the Science Education Alliance-Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) program in efforts to characterize viral diversity. Both bacteriophages infect https://phagesdb.org/DNAMaster). PECAAN resources applied to identify genes included Glimmer v3.0 (https://seaphages.org/software), ARAGORN v1.2.38 (http://www.cbs.dtu.dk/services/TMHMM), TOPCONS v2 (Following DNA extraction (Promega Wizard DNA clean-up system), sequencing libraries were produced (NEBNext Ultra II DNA library prep kit) and sequencing on an Illumina MiSeq system (MiSeq reagent kit v3)  was perfmer v3.0 , GeneMarmer v3.0 , Phameramer v3.0 , Starter v1.2.38 , and tRN v1.2.38 . Functio v1.2.38 , HHpred  v1.2.38 , TMHMM2 PCONS v2 , and thePCONS v2 . All proIn both genomes, key structural and assembly genes are on the left arm and nonstructural coding sequences are on the right arm. On the left arm, both genomes encode small and large terminase subunits, portal protein, capsid maturation protease, scaffolding protein, major capsid protein, head-to-tail adaptor, head-to-tail stopper, tail terminator, major tail protein, tail assembly chaperones, tape measure protein, minor tail proteins, lysin A/lysin B, and DNA polymerase III subunit. On the right arm, both code for genes regulating lysogeny , as well as WhiB family transcription factor, mobile element MPME1, and two glycosyltransferases.MH020243) , Misha28 (MH020242) , and Piper2020 (MK494120) . The Awesomesauce genome also includes Cas4-like exonuclease (gp66), two HNH endonucleases (gp1 and gp74), and two DNA methylases (gp73 and gp76). A rare synteny block (gp49 to gp61) that is highly conserved in 4 of the other 169 F1 mycobacteriophages  was identified. BLASTn comparison revealed remarkable colinearity and sequence similarity (98% identity) between the Awesomesauce synteny block and gp49 to gp61 in the singleton LilSpotty (MK977707), suggesting a common origin for the two. Excise (gp48) and MPME1 (gp62) border the synteny block and may facilitate its transposition. It was also noted that homologues to Awesomesauce gp82 (function unknown) occurred in 13 subcluster A1 bacteriophages but only 2 F1 mycobacteriophages (TootsiePop and Misha28) in addition to LilSpotty, suggesting shared origins for A1 and F1 bacteriophages.Whole-genome BLASTn alignments  revealedMH371115; 99.41% identity, with 95% coverage) and Brocalys . Notable features include a toxin/antitoxin system (gp35 and gp36), two helix-turn-helix DNA binding domains (gp53 and gp54), Ku-like double-stranded DNA break-binding protein (gp57), AAA-ATPase (gp58), HNH endonuclease (gp59), DNA helicase (gp60), and two GIY-YIG endonucleases (gp65 and gp82).LastJedi is highly homologous to F1 bacteriophages Clifton  (The Awesomesauce and LastJedi genomes are available in DDBJ/ENA/GenBank under accession numbers sdb.org) , MargotM"}
+{"text": "The 2010 Patient Protection and Affordable Care Act includes the Annual Wellness Visit (AWV) for older adult (OA) patients. Medicare pays for an initial AWV per beneficiary and subsequent visits annually. Many Medicare beneficiaries have not taken advantage of the AWV preventive health benefit. The Saint Louis University Geriatrics Workforce Enhancement Program (GWEP) developed an AWV for OA, NH residents. This project describes the NH AWV and reports results. Data include age, gender, comorbidities, medications, hospitalizations, depression, frailty, pain, sarcopenia, sensory impairment, cognition, nutrition, smoking, falls, and advance directives. Two suburban academic for-profit NHs are included in this study (2016-17). OA NH residents (N=247) completed an AWV and 36.8% (n=91) had a 1-year follow-up AWV. OA NH residents were female  and a majority ages 75+ . Most (96.3%) had a documented advance directive. Comorbidities (7.8\u00b12), polypharmacy (92.3%), vision impairment (52.8%), hearing impairment (52.8%), depression (65.2%), frailty (75.7%), sarcopenia (84.4%), risk of weight loss (53.9%), MCI (11.7%), and dementia (75.8%) were prevalent. Among OA NH residents (n=91) with an AWV follow-up, there was modest worsening in total comorbidities and medications as well as frailty, sarcopenia, and cognition scores (ps\u22640.05). Pain, depression, and nutrition did not change. To our knowledge, no one has specifically analyzed the Medicare AWV in NHs. Data from the traditional AWV is an extension of the routine clinical care of OAs and therefore could also be useful for healthcare professionals focused on providing care to OA patients in the NH setting."}
+{"text": "Culex (Melanoconion) (Diptera: Culicidae) as vectors of several arboviruses that cause diseases in humans and other animals, there are few taxonomic studies focusing on species of the subgenus, especially providing morphological keys for species identification.Despite the importance of some species of Culex (Melanoconion) were reviewed, five new species are described, and two taxonomic changes are proposed: Cx. (Mel.) exedrus Root, 1927 and Cx. (Mel.) loturus Dyar, 1925 are resurrected from synonymy with Cx. (Mel.) dunni Dyar, 1918 and Cx. (Mel.) zeteki Dyar, 1918, respectively. The Atratus Group now includes fourteen species: Cx. (Mel.) atratus Theobald, 1901; Cx. (Mel.) caribeanus Galindo & Blanton, 1954; Cx. (Mel.) columnaris S\u00e1 & Hutchings n. sp.; Cx. (Mel.) commevynensis Bonne-Wepster & Bonne, 1919; Cx. (Mel.) comptus S\u00e1 & Sallum n. sp.; Cx. (Mel.) dunni; Cx. (Mel.) ensiformis Bonne-Wepster & Bonne, 1919; Cx. (Mel.) exedrus; Cx. (Mel.) longisetosus S\u00e1 & Sallum n. sp.; Cx. (Mel.) longistylus S\u00e1 & Sallum n. sp.; Cx. (Mel.) loturus; Cx. (Mel.) spinifer S\u00e1 & Sallum n. sp.; Cx. (Mel.) trigeminatus Clastrier, 1970; and Cx. (Mel.) zeteki. Keys, descriptions and illustrations for the identification of the male, female, pupal and fourth-instar larval stages of each species are provided. The treatment of each species includes a complete synonymy, descriptions of available life stages, a taxonomic discussion, updated bionomics and geographical distribution, and a list of material examined.Thirteen species of the Atratus Group of Culex (Melanoconion) is updated, including descriptions of five new species. The number of valid species is greater than the number recognized in the previous taxonomic study of the group, increasing from seven to 14 species. Distributional and bionomical data are updated. Morphology-based identification keys for females, males, fourth-instar larvae and pupae provided in this study will facilitate species identification.The taxonomy of the Atratus Group of  Melanoconion Theobald, 1903 of Culex Linnaeus, 1758 are considered to be of public health importance because they are vectors of several arboviruses, such as the West Nile virus, viruses of the Venezuelan equine encephalitis complex, and eastern equine encephalomyelitis virus [Species of the subgenus is virus \u20139. Althois virus .Melanoconion includes 160 valid species and 79 synonyms for several species from both the Spissipes and the Melanoconion Sections [Melanoconion was proposed by Sirivanakarn [The subgenus Sections , 10\u201312. vanakarn , with soc oxidase subunit 1 (cox1) and two nuclear genes: hunchback (hb) and carbamoyl-phosphate synthetase 2, aspartate transcarbamylase and dihydroorotase (CAD) of 43 species. The authors demonstrated the monophyly of the Spissipes and Melanoconion Sections, and that most of the morphology-based groups of the Spissipes Section are also monophyletic, corroborating the morphological classification previously proposed by Sirivanakarn [The Spissipes Section comprises 23 species separated into eight groups and three subgroups . The Melvanakarn . In contCx. atratus Theobald, 1901 ; Cx. caribeanus Galindo & Blanton, 1954; Cx. commevynensis Bonne-Wepster & Bonne, 1919; Cx. dunni Dyar ; Cx. ensiformis Bonne-Wepster & Bonne, 1919; Cx. trigeminatus Clastrier, 1970; and Cx. zeteki Dyar, 1918 . The geographical distribution of the Atratus Group ranges from southern South America to northern Central America with Cx. atratus dispersed on some Caribbean islands, and Cx. dunni as the only member of the group recorded in Mexico [The Atratus Group includes seven valid species  and fiven Mexico \u201316.Culex dunni has epidemiological importance as a potential vector of arboviruses that can infect and cause encephalitis in humans, as it has been found naturally infected with Pacora (PCA) virus [Cx. dunni has been reported to be vector of Venezuelan equine encephalitis (VEE) virus in Panama [A) virus . In addin Panama .Culex atratus, Cx. zeteki, Cx. dunni, Cx. commevynensis, Cx. ruffinis and Cx. loturus in the Melanoconion Section. Edwards [Cx. commevynensis in Group B, and Cx. atratus, Cx. zeteki, Cx. dunni (syn. Cx. ensiformis), Cx. ruffinis (syn. Cx. exedrus) and Cx. loturus in Group C. Komp [Cx. ruffinis as a synonym of Cx. dunni and Cx. loturus as a synonym of Cx. zeteci. Rozeboom & Komp [Cx. zeteci to Cx. zeteki, in accordance with provisions of the International Code of Zoological Nomenclature. Galindo & Blanton [Cx. caribeanus and Clastrier [Cx. trigeminatus, both based on unique features of the male genitalia. Sirivanakarn [Several taxonomic changes have been made related to the species of the Atratus Group before the classification proposed by Sirivanakarn . Dyar 1 placed C Edwards , based o C. Komp  considerm & Komp  correcte Blanton  describelastrier  describevanakarn  classifiAccurate species identification is necessary for studies focusing on biology, ecology, vectorial capacity and vector competence. This study aimed to review the taxonomy of the Atratus Group and update the data on species bionomics and distributions. Additionally, five new species are formally named and described, two species are elevated from synonymy and illustrated identification keys to the species level are provided for females, males, fourth-instar larvae and pupae.Cx. commevynensis Bonne-Wepter & Bonne, 1919 and Cx. trigeminatus Clastrier, 1970. Female and male genitalia along with immature specimens from the same locality and habitat were examined when available. When available, male genitalia, larval and pupal exuviae associated to the pinned adult were mounted on the same slide. Character measurements, of 2\u20135 specimens when available, were obtained in the same manner as Sallum & Hutchings [Culex classification adopted is that proposed by Harbach [The specimens examined during this study are from the Cole\u00e7\u00e3o Entomol\u00f3gica de Refer\u00eancia, Faculdade de Sa\u00fade P\u00fablica, Universidade de S\u00e3o Paulo (FSP-USP), S\u00e3o Paulo, Brazil and from the Cole\u00e7\u00e3o de Invertebrados, Instituto Nacional de Pesquisas da Amaz\u00f4nia (INPA), Manaus, Brazil. All specimens come from field collections made in several localities in the Brazilian states of Acre, Amazonas, Mato Grosso do Sul, Minas Gerais, Par\u00e1, Rond\u00f4nia and S\u00e3o Paulo. Type specimens of the nominal species, deposited in the Diptera Collection in the National Museum of Natural History (USNM), Washington, D.C., USA and in the Natural History Museum (NHM), London, UK, were also examined, except for the types of utchings . Illustrutchings , 26, witutchings . Only th Harbach . The Ano Harbach .Geographical distributions are based on both literatute records and material examined, including field collections and museum specimens examined. Distribution records of the material examined are listed in the following format: country, state, municipality and/or locality name, latitude and longitude.Culex commevynensis is not included in this revision because the type specimen could not be examined.International Code of Zoological Nomenclature (ICZN) [To comply with the regulations set out in Article 8.5 of the amended 2012 version of the e (ICZN) , detailsThe abbreviations used are: L, larva; Le, larval exuviae; P, pupa; Pe, pupal exuviae; \u2642, male; \u2640, female; \u2642G, male genitalia; Syn., synonym; distr., distribution; tax., taxonomy; info., information; desig., designation; emend., emendation; FSP-USP, Faculdade de Sa\u00fade P\u00fablica of Universidade de S\u00e3o Paulo, S\u00e3o Paulo, Brazil; INPA, Instituto Nacional de Pesquisas da Amaz\u00f4nia, Manaus, Amazonas, Brazil; NHM, Natural History Museum, London, UK; USNM, National Museum of Natural History, Washington DC, USA; MNHN, Museum National d\u02bcHistoire Naturelle, Paris, France.Atratus GroupAccording to Sirivanakarn , the folTaxonomic treatmentCulex (Melanoconion) atratusTheobald, 19011901Culex atratus Theobald, 1901: 55 [1901: 55   l1909Culex falsificator Dyar & Knab, 1909: 258 [909: 258  (\u2642) lect1938Culex advieri Senevet, 1938: 185 [938: 185   Melanoconion atratus of Theobald (1903: 238) [Melanoconion).03: 238)     [15: 388)   atratus of Dyar (1923: 187) [23: 187)  (\u2642G); Bo23: 187)    (\u2642G); Fo23: 187)  (type in23: 187)  (type in23: 187)      (tax.); 23: 187)   [909: 50)  (distr.)909: 50)  (tax.); 909: 50)   falsificator of Edwards (1932: 214) [Cx. atratus); Stone & Knight (1957: 49) [32: 214)  (synonym957: 49)  (desig. 957: 49) .Culex advieri of Rozeboom & Komp (1950: 87) [Cx. atratus)950: 87)  (synonymCulex (Melanoconion) advieri of Floch & Abonnenc (1945: 29) [945: 29)    (type in945: 29) .Type material: Lectotype, pinned adult male (NHM 010630134) in poor condition associated with male genitalia, and paralectotype pinned adult female (NHM 0106300135) in good condition in the Diptera Collection, Natural History Museum (NHM), London, UK; topotypic male and female in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA. Lectotype male and paralectotype female of Cx. atratus examined as photographs provided by the Natural History Museum, London, UK, for comparisons.Material examined: 36 specimens: 25 \u2642G, 5 Le, 3 Pe, 7\u2642. USNM, Jamaica: JA798-90 (\u2642); JA 798-30 ; JA798-32 (\u2640); JA6- 101 (\u2642G); JA25-102 (\u2642G); JA2/3-84 (\u2642G); JA871-102 (\u2642G); JA744-90 ; JA798-96 ; JA414-103 (\u2642G); JA701-67/208-2 (\u2642G); JA727-102 ; JA719-6/208-3 (\u2642G); JA871-103 (\u2642G); JA899-10 (\u2642G); JA701-67/213-15 (\u2642G); JA719-67/213-16 (\u2642G); JA759-91 (\u2642G); JA798-40 (\u2642G); JA744-95 (\u2642G); JA862-67/213-17 (\u2642G); JA862-67/208-4 (\u2642G); JA744-1 (Le); JA798-3 (Le); JA899 (Le); JA862-2 (Le). Dominican Republic: RD1 ; RD2 (Pe); RD3 (Pe). French West Indies: FWI201-14 (\u2642G); FWI198-101 (\u2642G). Haiti: HAT11-103 (\u2642G); HAT8-680923-8 (\u2642G). Synonym Culex falsificator: Lectotype, pinned adult male (USNM no. 12108), with associated dissected genitalia (USNM no. 408) in good condition in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Distribution:Culex atratus has been found in Cuba [ in Cuba ; Jamaica in Cuba ; Dominic in Cuba ; Dominic in Cuba ; Brazil  in Cuba \u201350; Caym in Cuba ; Florida in Cuba ; Venezue in Cuba  and Pana in Cuba . The occDescriptionMale. [Figs.\u00a0Head: antenna dark, verticillate; length 1.17\u20121.25 (1.20) (n\u2009=\u20093); proboscis entirely dark-scaled, length 1.56\u20131.74 (1.67) (n\u2009=\u20093); maxillary palpus dark-scaled, length 2.08\u20132.21 (2.13) (n\u2009=\u20093); palpomere III with few long, strong setae at apex and inconspicuous basal white ring; palpomeres IV-V entirely covered with long, strong setae; clypeus and antennal pedicel dark. Vertex with narrow, white decumbent falcate scales on central area and erect, dark forked scales; large lateral patch of broad, decumbent white scales extending dorsally; ocular line with narrow, white falcate scales extending dorsally; occiput with dark erect forked scales. Thorax: integument brown; scutum covered with narrow, dark brown falcate scales with lightly golden-coppery reflections. Scutal setae large, dark brown with golden-coppery reflections. Scutellar scales similar to prescutellar scales on median and lateral lobes; median lobe with 4\u20136 large, dark setae; lateral lobes each with 3 or 4 setae. Pleural integument with pattern of pale and dark brown areas as follows: dark brown on postspiracular area, upper mesokatepisternum, upper and lower mesepimeron; pale on lower mesokatepisternum, mesomeron. Pleural setae with 2 types of colouring: setae dark brown with bronzy reflections: 7 or 8 antepronotal, 4\u20136 prealar; and pleural setae pale golden, hyaline: 4 or 5 upper mesokatepisternal, 3\u20135 lower mesokatepisternal, 3 or 4 upper mesepimeral and 1 large lower mesepimeral. Pleura with patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few scales not forming patch. Wing: dark-scaled, length 2.26\u20132.31 (2.29) (n\u2009=\u20093). Dorsal scales broad, dark distally on veins R1, R2, R3, R4+5, M1+2 and M3+4; appressed, dark spatulate scales on veins C, Sc, R, proximally on M, Cu1, Cu2 and A1; linear scales on Rs, R2+3; remigium with appressed spatulate scales and 2 setae. Halter: scabellum yellowish; pedicel yellowish, narrow, with brown dorsal strip; capitellum brown, with few scales with golden reflections. Legs: coxae pale; ventral surface of fore- and midfemur with longitudinal stripe of white scales; tibiae dark-scaled; joints of fermur-tibia and tibia-tarsomere I with ring of pale scales; tarsi entirely dark-scaled. Abdomen: tergum I with dark scales, terga III-VIII dark-scaled with basolateral patches of white scales; sterna II-VII with broad basal white bands. Genitalia: ninth tergal lobes pear-shaped, each with 12\u201314 slender, aciculate setae inserted at 0.67 from base, apex glabrous; distance between lobes 0.6 of width of one lobe at base. Gonocoxite small, narrow, oblong; subapical lobe divided into 2 columnar divisions; proximal division with 2 parallel, apically pointed setae (a and b); seta a shorter, slender, inserted basal to seta b; seta b spatulate and stronger than seta a. Distal division with short columnar process, with 5 setae: 3 filiform, narrow, pointed, apically inserted, subequal in size (setae f), 1 long seta, with hook-like apex (seta h), 1 large, broad, asymmetrical, ribbed seta arising subapically (seta l); 1 saber-like, ribbed seta (seta s) arising apically. Gonocoxite with 4 or 5 broad, hyaline, flattened, apically curved setae borne ventromesally between proximal and distal divisions. Gonostylus slender, slightly curved, tapering towards apex; apex moderately blunt, ventral surface with 2 apical hyaline setae; one short leaf-like gonostylar claw arising apically. Aedeagal sclerite and lateral plate equal in length; lateral process of lateral plate sclerotized, slightly pointed, directed dorsolaterally; ventral process almost straight. Aedeagal sclerite curved in lateral view. Proctiger with tergum X somewhat triangular, slender in outline, inner process pointed. Basal plate with concave inner margin. Paraproct elongate, crown with 9 or 10 simple blades. Cercal sclerite with 1 or 2 setae.e. Figs.\u00a0, 2a SmalFemale. Not examined.Pupa. [Figs.\u00a0Cephalothorax: setae 1,2-CT 4-branched (n\u2009=\u20092); setae 3,4-CT 2-branched; seta 5-CT 4-branched; seta 6-CT 2-branched; seta 7-CT 3-branched; seta 8-CT 5- or 6-branched; seta 9-CT 2- or 3-branched; seta 10-CT 5- or 6-branched; seta 11-CT single; seta 12-CT 2- or 3-branched. Trumpet moderately tanned. Pinna small, V-shaped in lateral view; tracheoid area, darker, extending almost 0.45 from base; trumpet index c.7. Abdomen: lightly tanned; seta 9-VIII with 4 aciculate branches. Paddle weakly tanned; setae 1,2-Pa single, 1-Pa longer than 2-Pa.a. Figs.\u00a0a, 4a IntLarva. [Figs.\u00a0Head: wider than long; capsule moderately tanned; lateralia and collar darker; length and width not measured; dorsomentum with 1 large median tooth and 4 small teeth on either side. Antenna lightly tanned with dark rings at base and level of seta 1-A; setae 2,3-C absent (n\u2009=\u20092); seta 4-C single; seta 5-C with 6 long branches reaching 6-C insertion; seta 6-C single, long, reaching anterior margin of head, with sparse minute spicules on basal 0.5; seta 7-C with 12 aciculate branches; seta 8-C 5- or 6 branched; seta 9-C 4-branched; seta 10-C 3-branched; seta 11-C double; seta 12-C 4- or 5-branched; seta 13-C single; seta 14-C 3-branched; seta 15-C with multiple hyaline branches. Thorax: integument hyaline; pleura without darker patches under integument. Abdomen: integument hyaline; comb of segment VIII with 19\u201328 sub-equal scales arranged in 3 rows. Segment X with complete saddle, apico-lateral margin dark with spicules; seta 1-X with 4 hyaline branches; seta 2-X with 1 long branch, 3 shorter; seta 3-X single; ventral brush (seta 4-X) with 5 pairs of 5-branched setae. Anal papillae slender, gradually tapering to apex. Siphon: long, at least 3 times longer than saddle, darker in mid-length; pecten with 18 spines on basal 0.3. Seta 1-S usually in 4 ventral pairs and 4 dorsal pairs; seta 2-S hook-shaped with small, curved secondary branch.a. Figs.\u00a0a, 6a HeaBionomics. Immature stages of Cx. atratus were collected in permanent and semi-permanent partially shaded habitats, such as ponds, stream margins, swamps and ditches, in association with herbaceous vegetation such as reeds, grass and algae, in fresh, clear or dark water. Larvae and pupae were found in association with Nyssorhynchus albimanus Wiedemann, 1820 and Anopheles grabhamii Theobald, 1901 and less frequently with Cx. nigripalpus Theobald, 1901 Uranotaenia socialis Theobald, 1901 and Ur. cooki Root, 1937 [Cx. atratus were found in artificial containers with Aedes albopictus  in Florida Key, Florida, USA [ot, 1937 . Larvae ida, USA . In the ida, USA  found laida, USA .RemarksCulex atratus was described as a species of the genus Culex by Theobald [Cx. atratus to the newly created genus Melanoconion, based on the arrangement of wing scales. Dyar [Cx. atratus as the type of genus Melanoconion, and Howard et al. [Cx. falsificator from adults collected in Cuba, and Bonne & Bonne-Wepster [Cx. falsificator with Cx. atratus, which was accepted by Edwards [Culex advieri was described by Senevet [Cx. advieri from Guadeloupe. Rozeboom & Komp [Cx. advieri with Cx. atratus; the synonymy was also recognized later by Belkin et al. [Cx. atratus can be identified by the following combination of characters: wings dark-scaled; small patch of pale scales on upper mesokatepisternum; scutum with very narrow, bronzy scales; mesepimeron entirely dark, without median pale area; terga II-VIII with basolateral patches of white scales. Males can be readily distinguished from the other species of the Atratus Group by the presence of 4 or 5 broad, hyaline, flattened, apically curved setae arising ventromesally between the proximal and distal divisions of the gonocoxite. In addition, other characteristics of the male genitalia can be employed to identify Cx. atratus: lateral plate without apical process and lateral process directed dorsolaterally, and ninth tergal lobe pear-shaped with aciculate setae arising at basal 0.6. Fourth-instar larvae can be distinguished by having the scales of the comb of segment VIII of equal size and arranged in four irregular rows; seta 5-C reaching the insertion of seta 6-C; siphonal pecten spines with large coarse marginal denticles. Culex atratus pupae can be distinguished by having a V-shaped pinna and seta 9-VIII with 4 or 5 aciculate branches.Theobald  based onTheobald  transferes. Dyar  selectedd et al.  described et al.  describe-Wepster  synonymi Edwards  and Belk Edwards . Culex a Senevet  from mal Senevet  associatm & Komp  synonymin et al.  and Belkn et al. . In spitCulex(Melanoconion)caribeanusGalindo & Blanton, 19541954Culex (Melanoconion) caribeanus Galindo & Blanton, 1954: 244 [954: 244  (\u2642) holoCulex (Melanoconion) caribeanus of Pecor et al.  [16, 124)      (distr.)Type material: Holotype, adult male mounted on slide with dissected male genitalia (USNM 01347) and paratype male mounted on slide with dissected male genitalia (USNM 01160) deposited in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Material examined: 4 specimens: 4 \u2642G, 2 \u2642: INPA, Brazil: Amazonas State, Ipixuna Municipality, Lago Grande, Seringal Recreio, Greg\u00f3rio River , coll. Hutchings et al. 2011, 20\u201321.v.2011, det. Hutchings & Sallum, 3.vii.2012 ; coll. Hutchings et al. 2011, 18\u201319.v.2011, det. Hutchings & Sallum, 3.vii.2012 . Amazonas State, Barcelos Municipality, Ararinha, Padauari River , coll. Hutchings et al. 2010, 6\u20137.vi.2010, det. Hutchings & S\u00e1, 01.iii.2017 . Amazonas State, Mau\u00e9s Municipality, Picada Pirarara, Abacaxis River , coll. Hutchings et al. 2008, 28\u201329.v.2008, det. S\u00e1, 01.iii.2017 .Distribution:Culex caribeanus has been found in Mojinga Swamp, Canal Zone, Panama [Cx. trigeminatus) [, Panama  and in tminatus) , Barcelominatus)  and IpixDescriptionMale. [Figs.\u00a0Head: antenna dark, verticillate, length 1.04\u20131.38 (1.21) (n\u2009=\u20092); proboscis dark-scaled, with conspicuous, median, dorsal patch of whitish scales, length 1.30\u20131.51 (1.40) (n\u2009=\u20092); maxillary palpus length 1.69\u20131.90 (1.79) (n\u2009=\u20092); palpomere I entirely whitish-scaled; palpomere II with basal patch of whitish scales; palpomere III with conspicuous patch of whitish scales on median portion. Thorax: scutum covered with bronzed scales, except whitish scales on anterior promontory, scutal fossa, dorsocentral and supraalar areas forming a pattern. Scutellar scales whitish; median lobe with 4 or 5 setae; lateral lobes with 3 or 4 setae each. Pleural setae with 2 types of colouring: dark brown with bronzy reflections: 7 or 8 antepronotal, 5 or 6 prealar; and pleural setae golden, hyaline: 4 or 5 upper mesokatepisternal, 3 or 4 lower mesokatepisternal, 5 or 6 upper mesepimeral, and 1 large lower mesepimeral. Pleura with patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few scales not forming patch. Wing: dark-scaled as in Cx. atratus; length 2.31\u20132.47 (2.39) mm (n\u2009=\u20092). Halter: scabellum and pedicel whitish; capitellum brown with few scales with golden reflections. Legs: fore- and midfemora with preapical ring of white scales. Abdomen: tergum I dark-scaled; terga III-VIII dark-scaled with basal bands of white scales. Genitalia: tergum IX lobes with concave inner margin, pointed and apex glabrous, median portion each with 14\u201316 slender, simple and aciculate setae; distance between lobes smaller than half basal width of 1 lobe. Gonocoxite narrow, oblong; proximal division of subapical lobe with 4 parallel, apically pointed setae : seta a more basal, spoon-shaped; seta b longer than others, spatulate, sinuous subapically; seta c thin, slender, filiform, inserted between setae b and d; seta d implanted on tubercle apical to seta b, filiform, long. Distal division with elongated columnar process, with 5 setae: 3 filiform, narrow, pointed, apically inserted, subequal in size (setae f), 1 filiform, with hook-like apex (seta h) and 1 large, broad, asymmetrical, ribbed seta arising subapically (l seta); 1 saber-like, ribbed seta (seta s) arising apically. Gonocoxite with 4 or 5 slender, hyaline, short, inconspicuous setae borne ventromesally between proximal and distal divisions. Gonostylus as in Cx. atratus, except on dorsal surface of apex with 2 or 3 inconspicuous folds. Ventral process of lateral plate with large convexity on upper border and conspicuous pointed projection directed ventrobasally. Proctiger with tergum X somewhat triangular in outline, inner process pointed and long.e. Figs.\u00a0b, 7 HeadFemale, pupa and larva. Unknown.Bionomics. Adult males were collected using CDC light traps with UV lamps in upland (terra firme) Amazon Forest [n Forest .RemarksCulex caribeanus was described by Galindo & Blanton [Cx. caribeanus can be misidentified as Cx. trigeminatus if the male genitalia are not properly dissected and mounted in lateral view. Based on characteristics of the female, Cx. caribeanus is similar to Cx. trigeminatus in possessing preapical rings of white scales on the fore- and midfemora and proboscis with patch of whitish scales on the median portion of ventral surface. Culex caribeanus differs from Cx. trigeminatus in having palpomere I entirely white-scaled, palpomere II with basal patch of white scales, palpomere III with conspicuous patch of white scales on median portion close to pale patch of proboscis, and wings entirely dark-scaled on ventral and dorsal surfaces. In Cx. trigeminatus, the palpomeres I and II are dark-scaled, palpomere III has a small basal patch of pale scales, and palpomere IV has an inconspicuous proximal patch of whitish scales, and the wings have veins C and R with basal patches of white scales. The male genitalia of Cx. caribeanus have simple and aciculate setae on the median portion of the ninth tergal lobes, whereas in Cx. trigeminatus the setae are simple. In addition, Cx. caribeanus differs from Cx. trigeminatus in possessing a pronounced convexity on the apical margin of the lateral plate, and a conspicuous pointed projection directed ventrobasally in the ventral process. In Cx. trigeminatus, the apical margin of the ventral process of the lateral plate is straight and bears a short projection. Blanton  from mal Blanton  found thCulex(Melanoconion)columnarisS\u00e1 & Hutchings n. sp.Type locality: Senador Guiomard Municipality in Fazenda Experimental Catuaba, UFAC, BR-364 Km 23 , Acre State, Brazil. Adults were collected using a CDC trap with UV light in terra firme forests at an elevation of 205 m.Type material: Holotype, pinned adult male with associated dissected genitalia on slide , with following collection data: Brazil: Acre State, Senador Guiomard Municipality, Fazenda Experimental Catuaba, UFAC, BR-364 Km 23 , coll. Hutchings & Carmo, 23-24.viii.2016, det. S\u00e1, 2017, deposited in the Cole\u00e7\u00e3o de Invertebrados, Instituto Nacional de Pesquisas da Amaz\u00f4nia (INPA), Manaus, Amazonas State, Brazil. Paratypes: 2 pinned adult males with dissected genitalia on separate slides  from same collection as holotype and deposited in INPA; and 2 pinned adult males with dissected genitalia on separate slides , from same collection as holotype and deposited in the Cole\u00e7\u00e3o Entomol\u00f3gica de Refer\u00eancia, Faculdade de Sa\u00fade P\u00fablica, Universidade de S\u00e3o Paulo (FSP-USP), S\u00e3o Paulo State, S\u00e3o Paulo municipality, Brazil.ZooBank registration: The Life Science Identifier (LSID) for Culex (Melanoconion) columnaris n. sp. is urn:lsid:zoobank.org:act: 139D4046-50EC-4D2A-AE1B-51502EC47A44.Etymology: From the Latin adjective columnaris, meaning rising in the form of column, in reference to the long columnar process of the proximal division of the subapical lobe.DescriptionMale. [Figs.\u00a0Head: antennal length 1.06\u20131.60 (1.40) (n\u2009=\u20095); proboscis entirely dark-scaled, length 1.44\u20131.70 (1.55) (n\u2009=\u20095); maxillary palpus dark-scaled, length 1.82\u20132.21 (1.96) (n\u2009=\u20095); occiput with dark brown erect forked scales. Thorax: scutum with narrow, brown, falcate scales with golden reflection, prescutellar area with whitish scales. Scutellar scales whitish, median lobe with 6 setae; lateral lobes each with 3 or 4 large setae. Pleural setae with 2 types of colouring: dark brown: 3\u20135 antepronotal, 4\u20136 prealar; and pleural pale golden, slender setae: 4 or 5 upper mesokatepisternal, 3 or 4 lower mesokatepisternal, 4 or 5 upper mesepimeral; lower mesepimeron with one strong, long, pale golden seta. Mesepimeral integument dark, with indistinct pale spot on median area, not dividing upper and lower areas. Pleura with less evident patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few white scales. Wing: dark-scaled; length 2.19\u20132.54 (2.35) (n\u2009=\u20095). Halter: scabellum, pedicel and capitellum whitish. Legs: coxae pale; ventral surface of fore- and midfemur with a longitudinal stripe of white scales; tibiae dark-scaled; joints of femur-tibia and tibia-tarsomere I with ring of pale scales; tarsi entirely dark-scaled. Abdomen: tergum I with dark scales; terga III-VII dark-scaled with proximal white bands; tergum VIII with dark scales. Genitalia: tergum IX lobes elongate, each with 14 slender, apically bifid, simple setae in median portion; apex glabrous. Distance between lobes about 1/3 of basal width of 1 lobe. Gonocoxite oblong, narrow; proximal division with long, apically divided, columnar process bearing 2 parallel setae (a and b): seta a basal, slender with pointed apex; seta b slightly sinuous with apex curved. Distal division with long columnar process with 5 setae: 3 narrow, filiform, apically pointed setae of different sizes (seta f), 1 long seta hook-like at apex (seta h), and 1 large, long, asymmetrical seta arising subapically (seta l); 1 saber-like seta with broad apex and without peduncle on base (seta s) arising apically; gonocoxite with 2 short, hyaline setae on ventromesal surface. Gonostylus slender, slightly curved, with moderately pointed apex, ventral surface with 2 apical hyaline setae; 1 short gonostylar claw arising apically. Aedeagus with apical process slightly curved dorsally and ventral process with rounded prominence. Proctiger with tergum X asymmetrical, with outer process rounded.e. Figs.\u00a0c, 8 InteRemarksCx. columnaris n. sp. bear more morphological similarities to Cx. zeteki than to other species of the Atratus Group. However, adult specimens of the new species differ from Cx. zeteki in having the mesepimeron with a slightly light stain on the median area, not divided into upper and lower areas. The male genitalia differ from those of Cx. zeteki in having the seta s without peduncle on base and with a broad apex, IX tergal lobes with bifid setae in the ventromedial region, and a lateral plate without undulations on the apical process. Furthermore, Cx. columnaris n. sp. has the proximal division of subapical lobe with an apically divided long columnar process which bears only two setae (a and b).The male genitalia and adults of Culex(Melanoconion)comptusS\u00e1 & Sallum n. sp.Type locality: Presidente Epit\u00e1cio Municipality near Horto Florestal , S\u00e3o Paulo State, Brazil. Larvae were collected in partially shaded, permanent habitats, with turbid water, associated with Pistia sp., in remnants of the Atlantic Forest and in transition areas between the Cerrado and the Atlantic Forest biomes, cohabiting with Cx. dunni.Other localities: Bolivia, Brazil, Panama and Suriname. In Brazil, the species occurs in the municipalities of Presidente Epit\u00e1cio and Dourado, S\u00e3o Paulo State, in Santo Ant\u00f4nio do I\u00e7\u00e1, Manacapuru and Juta\u00ed, Amazonas State, and in the municipality of Juruti, Par\u00e1 State (present study).Type material: Holotype, pinned adult male with dissected genitalia, larval and pupal exuviae on the same slide , with following collection data: Brazil, S\u00e3o Paulo State, Presidente Epit\u00e1cio Municipality, near Horto Florestal , coll. S\u00e1 & Chaves, 15.iii.2016, det. S\u00e1, 2016, deposited in the Cole\u00e7\u00e3o Entomol\u00f3gica de Refer\u00eancia, Faculdade de Sa\u00fade P\u00fablica, Universidade de S\u00e3o Paulo (FSP-USP), S\u00e3o Paulo Municipality, S\u00e3o Paulo State, Brazil. Paratypes: 2 pinned adult males with dissected genitalia, larval and pupal exuviae on separate slides ; 2 pinned adult females associated with larval and pupal exuviae on separate slides ; 2 pinned adult males with dissected genitalia and pupal exuviae on separate slides ; 1 pinned adult female with associated pupal exuviae on slide , from same collection as holotype and deposited in FSP-USP; 2 pinned adult males with dissected genitalia on separate slides , with following collection data: Brazil, Amazonas State, Santo Ant\u00f4nio de I\u00e7\u00e1 Municipality, Parana do Canini, Solim\u00f5es River , coll. Hutchings et al. 2003; 1 pinned adult male with dissected genitalia on slide , with following collection data: Brazil, Amazonas State, Jutai Municipality, S\u00e3o Raimundo, Parana do Cervalho, Solim\u00f5es River , coll. Hutchings et al. 2003; 1 pinned adult male with dissected genitalia on slide , with following collection data: Brazil, Amazonas State, Manacapuru Municipality, Parana do Cururu, Solim\u00f5es River , coll. Hutchings et al. 2003; and 1 pinned adult male associated with dissected male genitalia , with following collection data: Brazil, Par\u00e1 State, Juruti Municipality, Recreio, Parana de Dona Rosa, Amazon River , coll. Hutchings et al. 2003, deposited in the Cole\u00e7\u00e3o de Invertebrados, Instituto Nacional de Pesquisas da Amaz\u00f4nia (INPA), Manaus, Amazonas State, Brazil.Material examined: 21 specimens: 10 \u2642G, 16 Pe, 14 Le, 4 \u2642, 10 \u2640. FSP-USP, Brazil, S\u00e3o Paulo State, Presidente Epit\u00e1cio Municipality , coll. S\u00e1 & Chaves, 15.iii.2016, det. S\u00e1, 2016: SP172-06 ; SP172-17 ; SP172-28 ; SP172-32 ; SP172-34 ; SP172-36 ; SP172-37 ; SP172-39 ; SP172-43 ; SP 172-45 ; SP172-46 ; SP172-47 ; SP172-49 ; SP172-110 . S\u00e3o Paulo State, Dourado Municipality, SP255Km, Obelisco  coll. Sallum et al., 7.v.2009, det Sallum 2012: E-15439 (Pe). Dourado, SP255 km, Santa Leonor Farm , coll. Sallum et al. 7.v.2009, det. Sallum 2012: E-15440 . USNM, Panama: PA37-115 (\u2642G); PA2-101 (\u2642G). Suriname: (USNM) S.S det., 1978: MEP-AC634-20 (\u2642G); MEP-AC634-21 (\u2642G). Bolivia: Catalog no. 82164 (\u2642G).ZooBank registration: The Life Science Identifier (LSID) for Culex (Melanoconion) comptus n. sp. is urn:lsid:zoobank.org:act: 260B62E3-BD5B-4384-980A-AD7000EC8E17.Etymology: From the Latin adjective comptus, meaning ornate, adorned, in reference to the dark brown to black and pale golden scales that form a pattern on the scutum.DescriptionFemale. Integument dark brown, with pale areas on thoracic pleura. Head: antenna dark, flagellum normal, whorls with 5 setae, length 1.00\u20131.38 (1.21) (n\u2009=\u20095); proboscis dark-scaled, length 1.35\u20131.44 (1.42) (n\u2009=\u20095); maxillary palpus dark-scaled, length 0.21\u20130.26 (0.24) (n\u2009=\u20095). Occiput with dark brown erect forked scales. Thorax: integument dark brown; scutum covered with narrow, dark brown to black falcate scales; may have pale golden scales forming a pattern on anterior promontory, scutal fossa, dorsocentral, prescutelar and supraalar areas. Scutellar scales whitish; median lobe with 5 or 6 setae; lateral lobes each with 3 or 4 setae. Pleural setae with 2 types of colouring: brown, large: 3\u20137 antepronotal, 3\u20135 prealar; and pleural setae golden, hyaline: 4 or 5 upper mesokatepisternal, 3 or 4 lower mesokatepisternal, 4 or 5 upper mesepimeral and 1 large lower mesepimeral. Pleura with patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few white scales. Wing: dark-scaled; vein C with small proximal patch of white scales, vein Sc occasionally with inconspicuous proximal patch of white scales, vein R with proximal patch of white scales separated by median patch of dark scales; wing length 2.44\u20133.05 (2.79) (n\u2009=\u20095). Halter: scabellum and pedicel whitish; pedicel with narrow, brown, dorsal strip; capitellum brownish. Legs: coxae pale; ventral surface of fore- and midfemur with longitudinal stripe of white scales; tibiae dark-scaled; joints of femur-tibia and tibia-tarsomere I with ring of pale scales; tarsi entirely dark-scaled. Abdomen: tergum I dark-scaled; terga II-VIII dark-scaled with basal bands of white scales.Male. [Figs.\u00a0Head: antenna verticillate, length 0.92\u20131.37 (1.10) (n\u2009=\u20095); proboscis length 1.48\u20131.59 (1.55) (n\u2009=\u20095); maxillary palpus length 1.54\u20132.16 (1.89) (n\u2009=\u20095). Wing: length 2.37\u20132.63 (2.51) (n\u2009=\u20095). Genitalia: tergum IX lobes somewhat conical, elongate, narrow with glabrous apex, median portion each with 11\u201315 slender, simple setae; distance between lobes larger than basal width of 1 lobe. Gonocoxite oblong; proximal division of subapical lobe with 2 parallel, pointed setae (a and b): seta a basal, short, narrower than seta b, with pointed apex; seta b long, spatulate, with pointed apex and implanted on salient tubercle. Distal division with long columnar process, with 5 setae: 3 filiform, narrow, pointed, apically inserted, subequal in size (seta f); 1 filiform, with hooked apex (seta h); 1 large, long, broad, asymmetrical, ribbed seta arising subapically (seta l); and 1 saber-like, ribbed seta (seta s) arising apically. Gonocoxite with 3 or 4 hyaline, filiform, median, inconspicuous setae on ventromesal surface. Gonostylus slender, slightly curved, with moderately blunt apex, ventral surface with 2 apical hyaline setae, gonostylar claw extremely short. Aedeagal sclerite with few, inconspicuous spicules on ventral surface; lateral process pointed and directed dorsolaterally. Proctiger with tergum X somewhat triangular in outline, inner process pointed.e. Figs.\u00a0d, 9 EssePupa. [Figs.\u00a0Cephalothorax: trumpet cylindrical, pinna small, irregular in shape, pinna length 0.07\u20130.12 (0.10) (n\u2009=\u200910), distal margin opposite meatal cleft, with small emargination; tracheoid area darker, extending 0.20\u20130.25 (0.23) (n\u2009=\u200910) from base; trumpet index 13.8\u201316.2 (15.4) (n\u2009=\u200910). Abdomen: seta 9-VIII with 3 simple branches; paddle index 1.32\u20131.57 (1.47) (n\u2009=\u200910).a. Figs.\u00a0b, 4b CepLarva. [Figs.\u00a0Head: length 0.62\u20130.71 (0.67) (n\u2009=\u200910), width 1.04\u20131.10 (1.06) (n\u2009=\u20095). Antennal length 0.49\u20130.60 (0.55) (n\u2009=\u200910). Seta 1-A inserted 0.38\u20130.40 (0.38) (n\u2009=\u200910) from antennal base; seta 4-C double; seta 5-C with 3 or 4 long branches reaching 6-C insertion; seta 13-C double. Abdomen: comb of segment VIII with 25\u201328 sub-equal scales arranged in 3 rows. Segment X length 0.34\u20130.38 (0.36) (n\u2009=\u200910), saddle complete, apicolateral margin dark with spicules; siphon/saddle index 4.17\u20135.18 (4.67) (n\u2009=\u200910). Siphon: long, slender, index 8.7\u201311.0 (9.7) (n\u2009=\u200910); pecten with 12 marginal spines on basal 0.30 of siphon. Seta 1-S usually with 4 ventral pairs and 4 dorsal pairs.a. Figs.\u00a0b, 6b HeaRemarksCx. comptus n. sp. are more morphologically similar to Cx. commevynensis than to any other species of the Atratus Group. These species have in common characteristics such as the shape and size of seta l of the distal division, the length of the columnar process of the distal division and the number and form of setae in the proximal division. However, the male genitalia of Cx. comptus n. sp.\u00a0differ from those of Cx. commevynensis in possessing slightly conical, narrow and elongate tergum IX lobes. On the other hand, in the original description of the Cx. commevynensis adult, Bonne-Wepster & Bonne [Cx. comptus n. sp., the vertex and occiput possess whitish, curved, narrow scales and the scutum is covered with narrow and dark brown to black falcate scales and some pale golden scales forming conspicuous pattern on the anterior promontory, scutal fossa, dorsocentral, prescutelar and supraalar areas. The fourth-instar larva of Cx. comptus n. sp. differs from the larva of Cx. zeteki in having three rows of comb scales and slender pecten spines with serrate edges. The pupa of Cx. comptus n. sp. differs from the pupa of Cx. zeteki in having the pinna of the trumpet slightly smaller and differs from the pupa of Cx. trigeminatus in having the trumpet clearer and with the distal margin bearing a more conspicuous notch opposite the meatal cleft.Based on the original description of Bonne-Wepster & Bonne , and ill & Bonne  mention Culex(Melanoconion)dunniDyar, 19181918Culex (Melanoconion) dunni Dyar, 1918: 123 [918: 123   l1924Culex (Melanoconion) ruffinis Dyar & Shannon, 1924: 144 [924: 144   Culex (Melanoconion) dunni of Dyar (1923: 188) [Cx. ensiformis); Dyar (1928: 340) [23: 188)  (synonym of Dyar 23: 188 [28: 340)  (\u2642G); Fo28: 340)  ; 28: 340)  (desig. 28: 340)  (distr.)28: 340)  (Guatema28: 340)  (Venezue28: 340)  (distr.)Culex (Melanoconion) ruffinis of Dyar (1928: 341) [Cx. dunni). of Dyar 28: 341 [28: 341)  , with dissected genitalia on slide (USNM no. 901).Material examined: 56 specimens: 45 \u2642G, 15 Le, 18 Pe, 18 \u2642, 16 \u2640. FSP-USP, Brazil: S\u00e3o Paulo State, Canan\u00e9ia Municipality, Vilarinho , coll. S\u00e1 et al. 14.vii.2015, det. S\u00e1 2015: SP166-03 ; SP166-04 (Pe); SP166-21 . S\u00e3o Paulo State, Presidente Epit\u00e1cio Municipality , coll. S\u00e1 & Chaves 2016, det. S\u00e1 2016: SP172-35 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Itapitangui , coll. Forattini et al. 1985, det. Sallum 1985: EP014-1 ; EP058-1 ; EP058-3 ; EP070-1 . S\u00e3o Paulo State, Dourado , coll. Forattini et al. 1980, det. Sallum 1980: 02 (\u2642G); 28 (\u2642G); 148 . S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, Pariquera-Mirim , coll. Forattini et al. 24.vii.1984, det. Sallum 1984: HEP414-7 ; HEP414-8 ; HEP414-9 ; HEP414-18 ; HEP414-20 ; HEP429-3 ; HEP429-7 ; HEP440-7 . Minas Gerais State, Goian\u00e1 Municipality , coll. Bergo et al. 30.xi.2008, det. Sallum 2008: MG24-102 , Minas Gerais State, Carmo da Mata Municipality, Rural das Pedras Farm , coll. Bergo et al. 13.iv.2010, det. Sallum 2014: MG46-02 . INPA, Brazil: Par\u00e1 State, Almeirim Municipality, Arumanduba, Amazon River , coll. Hutchings et al. 19\u201320.viii.2003, det. Hutchings: ProV-047641 (\u2642G); ProV-047649 (\u2642G); ProV-047741 (\u2642G). Par\u00e1 State, Almeirim Municipality, Paraiso, Paranacuara, Amazon River , coll. Hutchings et al. 21\u201322.viii.2003, det. Hutchings: ProV-055512 (\u2642G). Par\u00e1 State, Prainha Municipality, Fazenda JK, Amazon River , coll. Hutchings et al. 22\u201323.x.2003, det. Hutchings & S\u00e1: ProV-049099 (\u2642G). Par\u00e1 State, Prainha Municipality, Curuauna River , coll. Hutchings et al. 24\u201325.x.2003, det. Hutchings & S\u00e1: ProV-049615 (\u2642G). Par\u00e1 State, Juruti Municipality, Recreio, Parana de Dona Rosa, Amazon River , coll. Hutchings et al. 30\u201331.x.2003, det. Sallum, Hutchings & S\u00e1: ProV-053664 (\u2642G); ProV-053667 (G); ProV-053670 (G); ProV-053674 (G); ProV-053678 (\u2642G); ProV-053679 (\u2642G); ProV-053688 (\u2642G). Amazonas State, Iranduba Municipality, Ramal do Lago Grande , coll. Hutchings et al. 8\u201310.ix.2008, det. Hutchings: IRam-000751 (\u2642G); IRam-000752 (\u2642G); IRam-000759 (\u2642G); IRam-000760 (\u2642G); IRam-000945 (\u2642G); IRam-002062 (\u2642G); IRam-002061 (\u2642G); IRam-002060 (\u2642G); IRam-001933 (\u2642G); IRam-001932 (\u2642G); IRam-001931 (\u2642G); IRam-001212 (\u2642G). Amazonas State, Juru\u00e1 Municipality, Igarap\u00e9 de Tamaniqua, Solim\u00f5es River , coll. Hutchings et al. 19.ix.2003, det. Sallum, Hutchings & S\u00e1: ProV-015891 (\u2642G); ProV-015900 (\u2642G). Amazonas State, Juta\u00ed Municipality, S\u00e3o Raimundo , coll. Hutchings et al. 16\u201317.ix.2003, det. S\u00e1 1.iii.2017: ProV-006542 (\u2642G). Amazonas State, Urucara Municipality, L\u00edrio do Vale , coll. Hutchings et al. 3.xi.2003, det. Sallum, Hutchings & S\u00e1: ProV-056752 (\u2642G). Synonym species Culex ruffinis: lectotype, pinned adult male in good conditions and male genitalia (USNM no. 1928), deposited in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Distribution:Culex dunni has been found in Central and South America, including Belize [Cx. dunni was collected in the municipalities of Iranduba, Juru\u00e1, Juta\u00ed, Manaus and Urucar\u00e1, Amazonas State; in Bataguassu Municipality, Mato Grosso do Sul State; Carmo da Mata and Goian\u00e1 Municipalities, Minas Gerais State; Almeirim, Juruti and Prainha Municipalities, Par\u00e1 State; Canan\u00e9ia, Dourado, Pariquera-A\u00e7u, and Presidente Epit\u00e1cio Municipalities in S\u00e3o Paulo State (present study).g Belize , Brazil g Belize , Colombig Belize , 65, Cosg Belize , French g Belize , 67, Guag Belize , Mexico g Belize , 16, Nicg Belize , Panama g Belize , Surinamg Belize  and Veneg Belize , 62, 68.DescriptionFemale.Head: antenna dark, flagellum normal, whorls with 4 or 5 setae, length 1.21\u20131.35 (1.26) (n\u2009=\u20095); proboscis dark-scaled, length 0.58\u20131.44 (1.19) (n\u2009=\u20095); maxillary palpus dark-scaled, length 0.18\u20130.19 (0.19) (n\u2009=\u20095). Occiput with dark brown erect forked scales. Thorax: integument brown; scutum covered with narrow, bronze falcate scales; possibly with whitish scales on anterior promontory, scutal fossa, dorsocentral and supraalar areas, but not forming a pattern. Scutellar scales withish; median lobe with 5 or 6 setae; lateral lobes each with 3 or 4 setae. Pleural setae with 2 types of colouring: brown with bronzy reflections: 5\u20137 antepronotal, 3\u20135 prealar; and pleural setae golden, hyaline: 5 or 6 upper mesokatepisternal, 4 or 5 lower mesokatepisternal, 4 or 5 upper mesepimeral, and 1 large lower mesepimeral. Pleura with patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few white scales. Wing: dark-scaled, vein R with 2 proximal patches of white scales separated by large patch of dark scales; occasionally vein C with small proximal patch of white scales; length 2.67\u20133.00 (2.86) (n\u2009=\u20095). Halter: scabellum, pedicel and capitellum brownish. Legs: as in Cx. atratus. Abdomen: tergum I dark-scaled; terga III-VIII dark-scaled with basal bands of white scales.Male. [Figs.\u00a0Head: antenna verticillate, length 1.03\u20131.19 (1.13) (n\u2009=\u20095); proboscis length 1.13\u20131.68 (1.45) (n\u2009=\u20095); maxillary palpus length 1.25\u20131.77 (1.51) (n\u2009=\u20095); palpomere III with inconspicuous whitish basal ring. Wing: length 2.49\u20132.76 (2.61) (n\u2009=\u20095). Genitalia: tergum IX lobes slightly globose, apex glabrous, median portion each with 15\u201320 slender, simple setae; distance between lobes smaller than half basal width of 1 lobe. Gonocoxite oblong; proximal division of subapical lobe with 4 parallel setae : seta a more basal, narrow, with pointed apex; seta b long, spatulate, rounded apex, implanted on salient tubercle; seta c thin, slender, filiform, slightly curved, inserted between setae b and d; seta d filiform, long, spatulate, implanted on tubercle, with blunt apex. Distal division with short columnar process, with 5 setae: 3 filiform, narrow, pointed, apically inserted, subequal in size setae (seta f), 1 filiform seta with hooked apex (seta h), and 1 large, long, broad, asymmetrical, ribbed seta arising subapically (seta l); 1 saber-like, ribbed seta (seta s) arising apically. Gonocoxite with 5 slender, hyaline, short, inconspicuous setae on ventromesal surface; sternomesal surface with long, strong evenly dispersed setae. Gonostylus as in Cx. atratus, with large gonostylar claw, with slightly rounded apex. Aedeagus with ventral process of lateral plate with numerous spicules; lateral process pointed and directed dorsolaterally. Proctiger with tergum X long, sinuous, somewhat elongated in outline, inner process pointed, narrow and long.e. Figs.\u00a0e, 10 EssPupa. [Figs.\u00a0Cx. atratus except for followings characters. Cephalothorax: setae 1,2-CT 4- or 5-branched; seta 4-CT 3-branched; seta 8-CT 4-branched; seta 11-CT single or double; seta 12-CT double. Trumpet with pinna of median size, irregular in shape, length 0.16\u20130.23 (0.20) (n\u2009=\u200910), distal margin opposite to meatal cleft, which has a large and conspicuous emargination; tracheoid area, darker, extending 0.15\u20130.23 (0.20) (n\u2009=\u200910) from base; trumpet index 12.4\u201320.0 (14.5) (n\u2009=\u200910). Abdomen: seta 9-VIII with 2 simple branches; paddle index 1.56\u20131.95 (1.71) (n\u2009=\u200910).a. Figs.\u00a0c, 4c SimLarva. [Figs.\u00a0Cx. atratus except for followings characters. Head: length 0.64\u20130.79 (0.74) (n\u2009=\u200910), width 1.02\u20131.16 (1.11) (n\u2009=\u20095). Antennal length 0.48\u20130.57 (0.54) (n\u2009=\u200910). Seta 1-A inserted 0.34\u20130.38 (0.37) (n\u2009=\u200910) from antennal base; seta 14-C double, strong. Abdomen: comb of segment VIII with 25\u201335 scales of similar size arranged in 3 or 4 rows. Segment X length 0.31\u20130.35 (0.34) (n\u2009=\u200910), siphon/saddle index 3.36\u20134.22 (3.87) (n\u2009=\u200910). Siphon: long, slender, index 6.2\u20138.9 (7.4) (n\u2009=\u200910); pecten with 12 marginal spines as from on basal 0.30. Seta 1-S with 4 ventral pairs and 6 dorsal pairs.a. Figs.\u00a0c, 6c In Bionomics. Immature specimens of Cx. dunni were collected in permanent and semi-permanent partially shaded ground habitats, with slightly turbid water, associated with herbaceous vegetation such as Pistia sp. The larvae were collected in remnants of the Atlantic Forest in southeastern Brazil and in transition areas between the Cerrado and the Atlantic Forest biomes, in association with Cx. ensiformis and Cx. comptus\u00a0n. sp.RemarksCulex dunni was described by Dyar [Cx. dunni to be identical to Cx. ensiformis of Bonne-Wepster & Bonne based on characteristics of the male genitalia. Bonne & Bonne-Wepster [Cx. dunni with Cx. ensiformis, and that Cx. ensiformis can be distinguished from Cx. dunni by the crescent-shaped plate at base of gonocoxite and the pattern of scales on the scutum. Dyar & Shannon [Cx. ruffinis from an adult male from Barro Colorado Island, Canal Zone, Panama. Komp [Cx. ruffinis with Cx. dunni, considering a possible misinterpretation of some features of the male genitalia in the original description. In addition, the author hypothesized that the male genitalia of Cx. commevynensis were similar to those of Cx. dunni, and that the differences noted by Bonne-Wepster & Bonne [Cx. dunni to a probable confusion with Cx. ensiformis, considered by them as a morphologically close species. Rozeboom & Komp [Cx. commevynensis possesses \u201ca hair-like\u201d seta on the proximal division of the subapical lobe, while Cx. dunni has several \u201cspines\u201d in that position, distinguishing the species. Later, Foote [Cx. commevynensis and suspected that this species was not valid; this author considered Cx. dunni, Cx. commevynensis and Cx. zeteki to be closely related species. Although there has been intense discussion about the taxonomy of Cx. dunni, this species bears characteristics that clearly differ from the other species of the Atratus Group, especially with regard to features of the male genitalia and of the immature forms. The male genitalia of Cx. dunni differ from those of Cx. ensiformis and Cx. commevynensis in having four parallel setae on the proximal division of the subapical lobe while the other species have only two setae. Culex dunni has a long seta l in the distal division of the subapical lobe and Cx. ensiformis has a short seta. Culex dunni also differs in possessing several spicules on the ventral process of the lateral plate and in having tergum X appearing long, sinuous and elongated in outline while Cx. ensiformis and Cx. commevynensis have a lateral plate without spicules and a shorter tergum X. Based on larval characteristics, Cx. dunni differs from Cx. ensiformis in having subequal comb scales; double and strong seta 14-C, and short pecten spines with a conspicuously serrate border. Additionally, Cx. dunni differs from Cx. ensiformis and the other species in having a pinna of median size and a conspicuous emargination on the distal margin opposite the meatal cleft. With respect to adult specimens, both Cx. dunni and Cx. ensiformis bear a patch of pale scales separated by dark scales on the base of vein R and occasionally a small pale patch on the base of vein C. However, Cx. dunni has bronze scales on the scutum, not forming a pattern and in the male, an inconspicuous whitish basal ring on palpomere III, different to what is observed in Cx. ensiformis, which possesses scutal scales with different colour that form a pattern and palpomere III of the male dark-scaled. by Dyar  from spe by Dyar , conside-Wepster  mentione Shannon  describema. Komp  synonymi & Bonne  were the & Bonne  attributm & Komp  verifiedr, Foote  describeCulex(Melanoconion)ensiformisBonne-Wepster & Bonne, 19191919Culex (Melanoconion) ensiformis Bonne-Wepster & Bonne, 1919: 176 [919: 176   ensiformis of Dyar (1923: 188) [Cx. dunni); Bonne & Wepster-Bonne (1925: 272) [Cx. zeteki); Foote (1954: 97) [Cx. zeteki, lectotype desig.); Pecor et al. (1992: 25) [23: 188)  (synonym25: 272)  (resurre25: 272)  (\u2642G); Ro25: 272)  (synonym954: 97)  (resurre992: 25)  (distr.)992: 25)    (Brazil)992: 25)  (distr.)Type material: Paratypes, pinned adult male with associated larval and pupal exuviae on slide, in poor condition (USNM no. 22709-BB638) and pinned adult female with associated larval and pupal exuviae on slide (USNM no. 22709-BB625) deposited in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Material examined: 155 specimens: 77\u2642G, 98Le, 143Pe, 7\u2642, 5\u2640. FSP-USP, Brazil: S\u00e3o Paulo State, Canan\u00e9ia Municipality, Vilarinho , coll. S\u00e1 et al. 14.vii.2015, det. S\u00e1 2015: SP166-02 ; SP166-07 ; SP166-08 ; SP166-09 ; SP166-11 ; SP166-12 ; SP166-14 ; SP166-15 ; SP166-16 ; SP166-17 ; SP166-18 ; SP166-19 ; SP166-20 ; SP166-22 ; SP166-24 ; SP166-25 ; SP166-26 ; SP166-27 ; SP166-28 ; SP166-33 ; SP166-34 ; SP166-35 ; SP166-37 ; SP166-38 ; SP166-39 ; SP166-40 ; SP166-41 ; SP166-42 ; SP166-44 ; SP166-46 ; SP166-47 ; SP166-48 ; SP166-49 ; SP166-52 ; SP166-53 ; SP166-55 ; SP166-56 ; SP166-57 ; SP166-58 ; SP166-59 ; SP166-60 ; SP166-62 ; SP166-65 ; SP166-66 ; SP166-68 ; SP166-69 ; SP166-70 ; SP166-73 ; SP166-75 ; SP166-76 ; SP166-77 ; SP166-78 ; SP166-79 ; SP166-80 ; SP166-81 ; SP166-82 ; SP166-88 ; SP166-89 ; SP166-91 ; SP166-93 ; SP166-94 ; SP166-96 ; SP166-97 ; SP166-99 ; SP166-100 (Pe); SP166-101 (Pe); SP166-102 ; SP166-103 (Pe); SP166-105 ; SP166-111 (Pe); SP166-112 (Pe); SP166-113 ; SP166-115A (Pe); SP166-115B (Pe); SP166-117 ; SP166-118 (Pe); SP166-119 ; SP166-120 (Pe); SP166-121 ; SP166-301 ; SP166-304 ; SP166-305 ; SP166-311 ; SP166-313 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Folha Larga Farm , coll. S\u00e1 et al. 14.vii.2015, det. S\u00e1 2015: SP167-07 ; SP167-09 ; SP167-10 ; SP167-13 ; SP167-16 ; SP167-17 ; SP167-18 ; SP167-20 ; SP167-21 ; SP167-24 ; SP167-25 ; SP167-32 ; SP167-33 ; SP167-39 ; SP167-124 ; SP167-127 ; SP167-129 ; SP167-133 ; SP167-134 (Pe). S\u00e3o Paulo State, Canan\u00e9ia Municipality, Vilarinho , coll. S\u00e1 et al. 12.ii.2016, det. S\u00e1 2016: SP171-07 ; SP171-08 ; SP171-09 ; SP171-10 ; SP171-11 ; SP171-12B ; SP171-111 ; SP171-118 (Pe); SP171-119 ; SP171-143 ; SP171-144 (Pe); SP171-146 (Pe); SP171-147 (Pe). B.M.M-9 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Itapitangui , coll. Forattini et al. 1983, det. Sallum 1983: HEP81-04 ; HEP81-05 (Pe); HEP 81-06 ; HEP81-07 (Pe); HEP81-08 ; HEP81-11 ; HEP81-12 (Pe); HEP81-13 ; HEP81-14 ; HEP81-15 ; HEP81-16 ; HEP81-18 ; HEP81-19 ; HEP81-20 ; HEP81-21 ; HEP81-23 ; HEP81-24 ; HEP81-25 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Itapitangui , coll. Forattini et al. 1983, det. Sallum 1983: HEP81-10 ; HEP81-31 ; HEP81-33 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, estrada de Canan\u00e9ia , coll. Forattini et al. 12.ix.1984, det. Sallum 1984: HEP434-04 . Amazonas State, Humait\u00e1 Municipality, Realidade, , coll. Chaves et al. 22.vii.2016, det. S\u00e1 2016: Coleta07-Humait\u00e1-01 ; Coleta07-Humait\u00e1-02 . INPA, Brazil: Amazonas State, Manaus Municipality, Acampamento Colosso, Fazenda Esteio , coll. Hutchings et al. 2002, det. Sallum & Hutchings: Fam-000631 ; Fam-000632 (\u2642G); Fam-002314 (\u2642G); Fam-002925 (\u2642G); Fam-002927 (\u2642G); Fam-002932 (\u2642G). Amazonas State, Manaus Municipality, Fazenda Esteio ; coll. Hutchings et al. 2003, det. Sallum & Hutchings: Fam-004667 (\u2642G). Amazonas, Manaus, Fazenda Esteio , coll. Hutchings et al. 2002, det. Sallum & Hutchings: Fam-003224 (\u2642G). Amazonas State, Manaus Municipality, Fazenda Porto Alegre, BR-174 , coll. Hutchings et al. 2002, det. Sallum & Hutchings: Fam-003557 (\u2642G). Amazonas State, Manaus Municipality, Fazenda Porto Alegre, BR-174 , coll. Hutchings et al. 2002, det. Sallum & Hutchings: Fam-002343 (\u2642G). USNM, Brazil: Par\u00e1 State, Altamira Municipality, km 158, coll. Reinert et al. 9.xi.1974: 446Coll- 111-109 (Pe \u2642); 111-133 (Pe \u2640); 111-107 (Pe \u2640); 111-129 (Pe \u2642).Distribution:Culex ensiformis has been collected in Belize [n Belize , Bolivian Belize , Brazil n Belize , French n Belize  and SuriDescriptionFemale.Head: antenna dark, flagellum normal, whorls with 5 setae, length 1.25\u20131.74 (1.49) (n\u2009=\u20095); proboscis dark-scaled, length 1.16\u20131.52 (1.40) (n\u2009=\u20095); maxillary palpus with dark scales, length 0.24\u20130.31 (0.27) (n\u2009=\u20095). Occiput with dark brown erect forked scales. Thorax: scutum covered with narrow, bronze, and golden falcate scales on acrostichal and dorsocentral areas; occasionally whitish scales on anterior promontory, supraalar, and prescutellar areas forming a pattern. Scutellar scales whitish; median lobe with 5 or 6 dark setae; lateral lobes each with 3 or 4 setae. Pleural setae with 2 types of colouring: brown with bronzy reflections: 8 or 9 antepronotal, 4 or 5 prealar; and pleural setae golden, hyaline: 3\u20135 upper mesokatepisternal, 3 or 4 lower mesokatepisternal, 4 or 5 upper mesepimeral and 1 large lower mesepimeral. Pleura with large patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few white scales. Wing: dark-scaled, vein R with 2 proximal patches of white scales separated by small patch of dark scales; vein C with proximal patch of white scales; wing length 2.58\u20133.38 (3.06) (n\u2009=\u20095). Halter: scabellum, pedicel and capitellum brownish. Legs: as in Cx. atratus. Abdomen: tergum I with dark scales; terga III-VII dark-scaled with basal bands of white scales; tergum VIII dark-scaled.Male. [Figs.\u00a0Head: antenna verticillate, length 1.05\u20131.63 (1.36) (n\u2009=\u20095); proboscis length 1.39\u20131.93 (1.65) (n\u2009=\u20095); maxillary palpus length 1.71\u20132.15 (1.99) (n\u2009=\u20095). Wing: length 2.69\u20133.16 (2.88) (n\u2009=\u20095). Genitalia: tergum IX lobes conical, elongate, slender, with apex glabrous, median portion each with 14\u201320 slender, simple setae; distance between lobes equivalent to basal width of 1 lobe. Gonocoxite oblong; proximal division of subapical lobe with 2 parallel setae (a and b): seta a more basal, narrow, slender, with pointed apex; seta b long, spatulate, with pointed apex, implanted on salient tubercle. Distal division with elongate columnar process, with 5 setae: 3 filiform, narrow, pointed, inserted apically, subequal in size (seta f), 1 filiform seta with hooked apex (seta h), and 1 short, broad, asymmetrical, ribbed seta arising subapically (seta l); 1 saber-like, ribbed seta (seta s) arising apically. Additionally, 1 small, hyaline, inconspicuous seta basally on columnar process. Gonocoxite with 3 or 4 slender, hyaline, short, inconspicuous setae on ventromesal surface. Gonostylus as Cx. atratus, with large gonostylar claw, with slightly pointed apex. Aedeagus with ventral process of lateral plate with small convexity; aedeagal sclerite with spicules on ventral surface. Proctiger with tergum X narrow, somewhat triangular in outline, with slightly pointed inner process.e. Figs.\u00a0f, 11 SimPupa. [Figs.\u00a0Cephalothorax: setae 1,2-CT 4- or 5-branched; seta 3-CT double; seta 4-CT 5-branched; seta 5-CT 3-branched; seta 6,7-CT 3-branched; seta 8-CT 7-branched; seta 9-CT 3-branched; seta 10-CT 6-branched; seta 11-CT double; seta 12-CT 4-branched. Trumpet long, slender, with dilated apex; pinna small, opening circular, pinna length 0.07\u20130.13 (0.11) (n\u2009=\u200910), distal margin opposite meatal cleft with shallow depression; tracheoid area extending 0.20\u20130.34 (0.29) (n\u2009=\u200910) from base; trumpet index 16.2\u201329.7 (24.2) (n\u2009=\u200910). Abdomen: seta 9-VIII with 2 simple branches; paddle index 1.49\u20131.95 (1.68) (n\u2009=\u200910).a. Figs.\u00a0d, 4d CepLarva. [Figs.\u00a0Head: length 0.70\u20130.81 (0.79) (n\u2009=\u200910), width 1.08\u20131.20 (1.13) (n\u2009=\u200910). Antennal length 0.62\u20130.70 (0.67) (n\u2009=\u200910); seta 1-A inserted 0.42\u20130.46 (0.45) (n\u2009=\u200910) from antennal base. Seta 5-C with 8 long branches; seta 11-C 3-branched; seta 13-C double. Abdomen: comb of segment VIII with 16\u201322 scales of different sizes arranged in 2 or 3 rows: upper rows with small, pointed scales; lower row with 5\u20139 large, pointed scales. Segment X length 0.32\u20130.39 (0.36) (n\u2009=\u200910), siphon/saddle index 4.26\u20134.95 (4.51) (n\u2009=\u200910). Siphon: long, slender, index 7.6\u201310.4 (9.2) (n\u2009=\u200910); pecten with 10 spines on basal 0.30 of siphon. Seta 1-S usually with 4 ventral pairs and 4 dorsal pairs.a. Figs.\u00a0d, 6d HeaBionomics. Immature specimens of Cx. ensiformis were collected in semipermanent partially shaded groundwater habitats with herbaceous vegetation such as Pistia sp. in remnants of the Atlantic Forest in association with Cx. dunni. Adults were collected in the Amazon Forest.RemarksCulex ensiformis was described by Bonne-Wepster & Bonne [Cx. dunni. Bonne & Wepster-Bonne [Cx. dunni. According to Bonne & Wepster-Bonne [Cx. dunni and Cx. ensiformis in the same material, because Cx. ensiformis possesses morphological differences, such as the crescent-shaped lateral plate and the scale pattern of the scutum, that can distinguish it from Cx. dunni. Senevet & Abonnec [Cx. ensiformis to be close to but distinct from Cx. dunni, and resurrected it from synonymy again. Rozeboom & Komp [Cx. ensiformis with Cx. zeteki and considered the former to be a synonym of Cx. zeteki based on features of the male genitalia and the color pattern of the scales on the scutum. Likewise, Foote [Cx. ensiformis in synonymy with Cx. zeteki based on the presence of two types of comb scales in the larva. Belkin [Cx. ensiformis as a distinct species close to Cx. commevynensis but not conspecific with Cx. zeteki, and designated a lectotype male associated with larval and pupal exuviae while resurrecting Cx. ensiformis . Pecor et al. [Cx. ensiformis and Cx. commevynensis based on the morphology of the pupal stage. According to these authors, Cx. ensiformis is most readily distinguished from Cx. commevynensis and the other species belonging to Atratus Group in the pupal stage, because it bears morphological characteristics markedly unique to this species, as follows: trumpet distinctly flared at the apex and a trumpet index greater than 10. Culex zeteki has a trumpet with a smaller pinna than in Cx. ensiformis. Culex dunni has a trumpet with a larger pinna than in Cx. ensiformis and with a conspicuous emargination. Culex commevynensis has a straight pinna which is not flared apically as in Cx. ensiformis. Regarding the larval stage, Cx. ensiformis can be distinguished from the other species belonging to the group in having seta 5-C with 5 or more long branches , and comb scales of two different sizes in 2 or 3 rows . Comb scales of uneven sizes can also be found in Cx. trigeminatus; however, Cx. ensiformis differs from Cx. trigeminatus in having a few large scales and in having seta 5-C with long branches reaching the base of seta 6-C. Adults of Cx. ensiformis can be distinguished from the other species in possessing vein R with two proximal patches of white scales separated by small patch of dark scales, vein C with a proximal patch of white scales and the scutum with a pattern of whitish, bronze and golden scales, whereas Cx. zeteki has entirely dark-scaled wings, Cx. trigeminatus has a single large patch of whitish scales on vein R, Cx. dunni has two short patches of whitish scales on vein R and Cx. comptus n. sp.\u00a0has dark and whitish scales on the scutum. Regarding male genitalia, Cx. ensiformis can be readily distinguished from Cx. dunni, Cx. zeteki and Cx. trigeminatus in having two parallel spatulate setae on the proximal division of the subapical lobe and conspicuous spicules on the ventral surface of the aedeagal sclerite. Culex ensiformis differs from Cx. comptus n. sp.\u00a0in having a short columnar process and a short seta l on the distal division of the subapical lobe. & Bonne  from adu & Bonne  synonymier-Bonne  resurrecer-Bonne , Dyar ma Abonnec  considerm & Komp  comparede, Foote  maintain. Belkin  considerr et al.  providedCulex(Melanoconion)exedrusRoot, 19271927Culex (Melanoconion) exedrus Root, 1927: 580 [927: 580   exedrus of Dyar (1928: 341) [Cx. ruffinis); Rozeboom & Komp (1950: 89) [Cx. dunni), Stone & Knight (1957: 49) [ of Dyar 28: 341 [950: 89)  (synonym957: 49)  , in poor condition, with dissected genitalia on slide (USNM no. 30-1) deposited in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Distribution:Culex exedrus has been collected in the Porto das Caixas and Paracambi municipalities, Rio de Janeiro State, Brazil [, Brazil .Description Male. [Fig.\u00a0Cx. dunni, except as follows: Genitalia: gonocoxite with long, strong setae, aligned from base to apex on sternomesal surface. Proximal division of subapical lobe with 3 parallel setae : seta a inserted basally, narrow; seta b long, spatulate, borne on salient tubercle; seta c filiform, long, spatulate, borne on small tubercle, apex blunt; and 1 saber-like, ribbed seta (seta s) with broad apex, arising apically. Proctiger with tergum X long, sinuous, somewhat elongate in outline, inner process pointed, long and wide.le. Fig.\u00a0 EssentiaFemale. Not examined.Pupa and larva. Unknown.Bionomics. Immatures of Cx. exedrus were collected in ground water sites such as river margins, lagoons and ponds, associated with thick aquatic vegetation [getation .RemarksCulex exedrus was described by Root [Cx. exedrus with Cx. ruffinis Dyar & Shannon [Cx. exedrus as a junior synonym of Cx. dunni and maintained this taxonomic status until Cx. exedrus was resurrected from synonymy. Culex exedrus can be distinguished from Cx. dunni by features of the male genitalia, mainly a large number of long setae visibly lined up on the sternomesal surface of the gonocoxite; additionally, Cx. exedrus has seta s of the gonocoxite with a wider apex and a proctiger with the inner process of tergum X wider than in Cx. dunni. by Root  based on by Root  synonymi Shannon . Rozeboo Shannon  considerCulex(Melanoconion)longisetosusS\u00e1 & Sallum n. sp.Type locality: Pariquera-A\u00e7u Municipality , S\u00e3o Paulo State, Brazil. Adults were collected in the Atlantic Forest and in seasonally flooded v\u00e1rzea forests along the Amazonas and Solim\u00f5es Rivers.Other localities: Pariquera-A\u00e7u Municipality, S\u00e3o Paulo State; Santo Ant\u00f4nio do I\u00e7\u00e1, Juta\u00ed, Coari and Itacoatiara municipalities, Amazonas State; and, in Juruti and Almeirim municipalities, Par\u00e1 State, Brazil.Type material: Holotype, pinned adult male with associated dissected genitalia on slide , with following collection data: Brazil, S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality , coll. Forattini et al., 11.i.1979, with Shannon trap, deposited in the Cole\u00e7\u00e3o Entomol\u00f3gica de Refer\u00eancia, Faculdade de Sa\u00fade P\u00fablica, Universidade de S\u00e3o Paulo (FSP-USP), S\u00e3o Paulo, S\u00e3o Paulo State, Brazil. Paratypes: 7 pinned adult males with associated dissected genitalia on slide from same collection as holotype and deposited in FSP-USP: specimen field no. 2661, accession no. FSP-USP E-15900 (coll. 10.vi.1980), specimen field no. 2695, accession no. FSP-USP E-15892 (coll. 17.vi.1980), specimen field no. 2756, accession no. FSP-USP E-15893 (coll. 11.xii.1980), specimen field no. 02, accession no. FSP-USP E-15894 (coll. 9.ii.1981), specimen field no. 2173, accession no. FSP-USP E-15895 (coll. 10.ii.1981), specimen field no. 01, accession no. FSP-USP E-15896 (coll. 29.i.1981), and specimen field no. 2967, accession no. FSP-USP E-15897 (coll. 12.iii.1981); 1 pinned adult male with associated dissected genitalia on slide , with following collection data: Brazil, S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, Experimental Farm, 7.v.1984; 1 pinned adult male with associated dissected genitalia on slide , with following collection data: Brazil, S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, Pariquera-Mirim district  on 2-II-1985, both deposited in the FSP-USP; and 5 pinned adult males with associated dissected genitalia on separate slides, from different locations: specimen field no. ProV-053607, accession no. INPA-DIP 004574, with following collection data: Brazil, Par\u00e1 State, Juruti, Recreio, Parana de Dona Rosa, Amazon River , coll. Hutchings et al. 30-31.x.2003, det. Sallum & Hutchings 2016; specimen field no. ProV-047936, accession no. INPA-DIP 004575, with following collection data: Brazil, Par\u00e1 State, Almeirim Municipality, Arumanduba, Amazon River , coll. Hutchings et al. 19-20.x.2003, det Sallum, Hutchings & S\u00e1 2017; specimen field no. ProV-005165, accession no. INPA-DIP 004576, with following collection data: Brazil, Amazonas State, Santo Ant\u00f4nio do I\u00e7\u00e1 Municipality, Parana do Canini, Solim\u00f5es River , coll. Hutchings et al. 15-16.ix.2003, det. Hutchings & S\u00e1 2017; specimen field no. ProV-044118, accession no. INPA-DIP 004577, with following collection data: Brazil, Amazonas State, Coari Municipality, Ilha do Botija, Trocaris, Solim\u00f5es River , coll. Hutchings et al. 25-26.ix.2003, det. Hutchings & S\u00e1 2017; specimen field no. ProV-057487, accession no. INPA-DIP 004573) with following collection data: Brazil, Amazonas State, Itacoatiara Municipality, S\u00e3o Jorge, Parana da Eva, Amazon River , coll. Hutchings et al. 7-8.ix.2003, det. Hutchings & S\u00e1 2017, all deposited in the Cole\u00e7\u00e3o de Invertebrados, Instituto Nacional de Pesquisas da Amaz\u00f4nia (INPA), Manaus, Amazonas State, Brazil.Material examined: 3 G\u2642, 3 \u2642. INPA, Brazil: Par\u00e1 State, Juruti Municipality, Recreio, Parana de Dona Rosa, Amazon River , coll. Hutchings et al. 30\u201331.x.2003, det. Sallum & Hutchings 2016: ProV-053597 (\u2642G). Par\u00e1 State, Almeirim Municipality, Arumanduba, Amazon River , coll. Hutchings et al. 19\u201320.x.2003, det. Sallum, Hutchings & S\u00e1 2017: ProV-047940 (\u2642G). Amazonas State, Juta\u00ed Municipality, S\u00e3o Raimundo, Parana do Cervalho, Solim\u00f5es River , coll. Hutchings et al. 16-17.ix.2003, det. Hutchings & S\u00e1 2017: ProV-007278 (\u2642G).ZooBank registration: The Life Science Identifier (LSID) for Culex (Melanoconion) longisetosus n. sp. is urn:lsid:zoobank.org:act: 2F0C0B21-08FE-458E-94D3-EB8D2550B5C8.Etymology: The name longisetosus is derived from a combination of the Latin noun saeta, meaning \u201cseta, bristle\u201d and with the Latin adjective l\u0101tus, meaning \u201cextensive, broad\u201d. Culex longisetosus is named in reference to the four long and spatulate setae borne ventromesally between the proximal and distal divisions of the subapical lobe of the male genitalia.Description Male. [Figs.\u00a0Head: antennal length 1.02\u20131.71 (1.25) (n\u2009=\u20095); proboscis entirely dark-scaled, length 1.04\u20131.65 (1.43) (n\u2009=\u20095); maxillary palpus dark-scaled, length 1.32\u20131.98 (1.71) (n\u2009=\u20094); palpomere II with small, basal patch of whitish scales; palpomere III with inconspicuous proximal patch of whitish scales; palpomeres IV and V dark-scaled, with long, strong setae. Occiput with dark brown, erect forked scales. Thorax: scutum covered with narrow, dark brown falcate scales, except anterior promontory and prescutellar area with whitish scales. Median scutellar lobe with 6 dark large setae; lateral lobes each with 4 setae. Pleural setae with 2 types of colouring: dark brown: 3\u20135 antepronotal, 4 or 5 prealar; and pleural pale golden, slender setae: 4 upper mesokatepisternal, 5 lower mesokatepisternal, 4 or 5 upper mesepimeral; lower mesepimeron with 1 long, strong seta. Pleura with distinct patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few white scales. Wing: dark-scaled; length 2.37\u20132.49 (2.45) (n\u2009=\u20095). Halter: scabellum and pedicel whitish, capitellum whitish with few brown scales. Legs: coxae pale; ventral surface of fore- and midfemur with longitudinal stripe of white scales; tibiae dark-scaled; joints of femur-tibia and tibia-tarsomere I with ring of pale scales; tarsi entirely dark-scaled. Abdomen: tergum I with dark scales; terga III-VII dark-scaled, with white basal bands. Genitalia: tergum IX lobes elongate, each with 12\u201314 slender, apically bifid, and simple setae in median portion; apex glabrous; distance between lobes less than basal width of 1 lobe. Gonocoxite oblong, narrow, small; subapical lobe divided into 2 columnar divisions; proximal division with 2 pointed setae (a and b); seta a shorter, slender, inserted basal to seta b; seta b spatulate, robust; gonocoxite with 4 long, spatulate setae on ventromesal surface; distal division with long columnar process, with 5 setae: 3 filiform, narrow, pointed, different in size (seta f), 1 long seta with hooked apex (seta h), and 1 large, broad, asymmetrical seta arising subapically (seta l); 1 saberlike seta (seta s) arising apically. Gonostylus with apex moderately rounded, short leaf-like gonostylar claw borne apically. Aedeagus with ventral process with small convexity.e. Figs.\u00a0g, 13 HeaRemarksCx. longisetosus n. sp. differ from Cx. atratus in possessing an inconspicuous basal patch of whitish scales on palpomere III and a small basal patch of whitish scales on palpomere II. The male genitalia of Cx. longisetosus n. sp.\u00a0can be distinguished from those of other species of the Atratus Group in having a long columnar process in the distal division, elongate and slightly widened from the base to the apex of the ninth tergal lobe, and 4 long, spatulate setae on the ventromesal surface of the gonocoxite.Adults of Culex(Melanoconion)longistylusS\u00e1 & Sallum n. sp.Type locality: Dourado Municipality , S\u00e3o Paulo State, Brazil. Adults were collected in transitional vegetation areas between the Atlantic Forest and Cerrado biomes, and in seasonally flooded v\u00e1rzea forest areas along the Amazon River.Other localities: Dourado and Presidente Epit\u00e1cio municipalities, S\u00e3o Paulo State; Bataguassu Municipality, Mato Grosso do Sul State; Senador Guiomard Municipality, Acre State; Itacoatiara Municipality, Amazonas State; Almeirim, Prainha, Obidos, Santar\u00e9m and Juruti municipalities, Par\u00e1 State.Type material: Holotype male pinned with associated dissected genitalia on slide , with following collection data: Brazil, S\u00e3o Paulo State, Dourado Municipality , coll. Forattini et al. 2.ix.1980, with CDC light trap at the edge of the forest, deposited in the Cole\u00e7\u00e3o Entomol\u00f3gica de Refer\u00eancia, Faculdade de Sa\u00fade P\u00fablica, Universidade de S\u00e3o Paulo (FSP-USP), S\u00e3o Paulo, S\u00e3o Paulo State, Brazil. Paratypes: 1 pinned adult male with dissected genitalia on slide , from the same collection as the holotype and deposited in the same institution (FSP-USP); 1 pinned adult male with dissected genitalia on slide , with following collection data: Brazil, Par\u00e1 State, Santar\u00e9m Municipality, Parana de Ituqui, Amazon River , coll. Hutchings et al. 25\u201326.x.2003, det. Hutchings 2015; 2 pinned adult males with dissected genitalia on separate slides , with following collection data: Brazil, Acre State, Senador Guiomard Municipality, Fazenda Experimental Catuaba, UFAC, BR-364 km 23 , coll. Hutchings & Carmo 23\u201324.viii.2016, S\u00e1 3.iii.2017; 2 pinned adult males with dissected genitalia on separate slides , with following collection data: Brazil, Par\u00e1 State, Almeirim Municipality, Para\u00edso, Paranaquara, Amazon River , coll. Hutchings et al. 21\u201322.x.2003, det. Hutchings 2015, deposited in the Cole\u00e7\u00e3o de Invertebrados, Instituto Nacional de Pesquisas da Amaz\u00f4nia (INPA), Manaus, Amazonas State, Brazil.Material examined: 50 G\u2642, 2 \u2642. FSP-USP, Brazil: S\u00e3o Paulo State, Presidente Epit\u00e1cio Municipality, Peixe River , coll. Gomes et al. 10.xii.1997, det. S\u00e1 2015: LAM. no. 01 (\u2642G); LAM. no. 02 (\u2642G). S\u00e3o Paulo State, Presidente Epit\u00e1cio Municipality, Jo\u00e3o Baiano Farm , coll. Gomes et al. 2.v.1998, det. S\u00e1 2015: LAM. no.05 (\u2642G); LAM. no. 06 (\u2642G); LAM. no. 07 (\u2642G). S\u00e3o Paulo State, Presidente Epit\u00e1cio Municipality, Campinal , coll. Gomes et al. 4.i.1999, det. S\u00e1 2015: LAM. no. 03 (\u2642G). Mato Grosso do Sul State, Bataguassu Municipality, Romualdo Farm , coll. Gomes et al. 2.viii.1997, det. S\u00e1 2015: LAM. no. 08 (\u2642G). INPA, Brazil: Par\u00e1 State, Almeirim Municipality, Arumanduba, Amazon River , coll. Hutchings et al. 19\u201320.x.2003, Hutchings and Sallum det.: ProV-047634 (\u2642G); ProV-047673 (\u2642G); ProV-047692 (\u2642G); ProV-047764 (\u2642G); ProV-047962 (\u2642G). Par\u00e1 State, Prainha Municipality, Fazenda JK, Parana do Mouratuba, Amazon River , coll. Hutchings et al. 22\u201323.x.2003, det. Hutchings 2015: ProV-048997 ; ProV-049054 (\u2642G); ProV-048945 (\u2642G). Par\u00e1 State, Prainha Municipality, Boca do Rio Curuauna, Amazon River , coll. Hutchings et al. 24-25.x.2003, det. Hutchings 2015: ProV-049608 . Par\u00e1 State, Almeirim Municipality, Para\u00edso, Paranaquara, Amazon River , coll. Hutchings et al. 2003, det. Hutchings 2015: ProV-055464 ; ProV-055376 . Par\u00e1 State, Obidos Municipality, Ilha do Amador \u201cIlha Grande\u201d, Parana do Capivara, Amazon River , coll. Hutchings et al. 29\u201330.x.2003, det. Hutchings 2015: ProV-050860 (\u2642G); ProV-050878 (\u2642G); ProV-050909 (\u2642G); ProV-050910 (\u2642G); ProV-050943 (\u2642G); ProV-050964 (\u2642G); ProV-050974 (\u2642G); ProV-050978 (\u2642G); ProV-050982 (\u2642G); ProV-051027 (\u2642G); ProV-051033 (\u2642G); ProV-051049 (\u2642G); ProV-051053 (\u2642G); ProV-051055 (\u2642G); ProV-051079 (\u2642G); ProV-051092 (\u2642G); ProV-051104 (\u2642G); ProV-051106 (\u2642G); ProV-051181 (\u2642G). Par\u00e1 State, Juruti Municipality, Recreio, Parana de Dona Rosa, Amazon River , coll. Hutchings et al. 30\u201331.x.2003, det. Hutchings 2015: ProV-053577 (\u2642G); ProV-053593 (\u2642G); ProV-053599 (\u2642G); ProV-053600 (\u2642G); ProV-053618 (\u2642G); ProV-053657 (\u2642G); ProV-053668 (\u2642G); ProV-053683 (\u2642G). Amazonas State, Itacoatiara Municipality, S\u00e3o Jorge, Parana da Eva, Amazon River , coll. Hutchings et al. 7\u20138.xi.2003, det. Hutchings 2015: ProV-057494 (\u2642G); ProV-057497 (\u2642G). USNM, Ecuador: (as Cx. ensiformis), coll. 3.xii.1981: EC8-1263(n 866) (\u2642G). Brazil: S\u00e3o Paulo State, Iguape Municipality (as Cx. ensiformis), coll. unknown, det. S.S. 1987: no. 050977-14 (\u2642G); S\u00e3o Paulo State, Canan\u00e9ia Municipality (as Cx. ensiformis) coll. unknown, det. S.S. 1987: no. 050977-15 (\u2642G).ZooBank registration: The Life Science Identifier (LSID) for Culex (Melanoconion) longistylus n. sp. is urn:lsid:zoobank.org:act:C0FD06D0-B7E2-4775-A8BB-DC776858132F.Etymology: The specific epithet longistylus is a combination of the Latin adjective longus (long) and the Latin noun stylus , in reference to the long columnar process in the distal division of the subapical lobe of the male genitalia.DescriptionMale. [Figs.\u00a0Head: antennal length 0.92\u20131.47 (1.23) (n\u2009=\u20096); proboscis entirely dark-scaled, length 1.23\u20131.64 (1.48) (n\u2009=\u20096); maxillary palpus dark-scaled, length 1.40\u20132.23 (1.73) (n\u2009=\u20096). Occiput with dark brown erect forked scales. Thorax: scutum covered with narrow, dark brown falcate scales, except prescutellar area with whitish scales. Median scutellar lobe with 6 large, dark setae; lateral lobes each with 4 setae. Pleural setae with 2 types of colouring: dark brown with bronzy reflections: 3\u20136 antepronotal; 3\u20135 prealar; and pleural setae golden, hyaline: 4 or 5 upper mesokatepisternal, 4 or 5 lower mesokatepisternal; 5 upper mesepimeral; lower mesepimeron with 1 long, strong seta. Pleura with patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few scales, extending dorsally on posterior margin. Wing: mostly dark-scaled, sometimes with minute patch of white scales at proximal end of vein C; length 2.08\u20132.45 (2.27) (n\u2009=\u20096). Halter: scabellum and pedicel whitish; capitellum pale brown with few golden scales. Legs: coxae pale; ventral surface of fore- and midfemur with longitudinal stripe of white scales; tibiae dark-scaled; joints of fermur-tibia and tibia-tarsomere I with ring of pale scales; tarsi entirely dark-scaled. Abdomen: tergum I with dark scales, terga III\u2013VII dark-scaled with white basal bands. Genitalia: tergum IX as illustrated ; seta a short, slender, inserted basal to seta b; seta b spatulate; gonocoxite with 3 short filiform setae with pointed apices on ventromesal surface; distal division with long columnar process, with 5 setae: 3 narrow, filiform, apically pointed setae, subequal in size (seta f), 1 long seta with hook-like apex (seta h), and 1 large, broad, ribbed asymmetrical seta arising subapically (seta l); 1 saber-like seta (seta s) arising apically. Gonostylus slender, slightly curved, tapering towards apex, apex moderately blunt, ventral surface with 2 apical hyaline setae; 1 short leaf-like gonostylar claw. Aedeagus with sclerotized, slightly pointed, dorsolaterally directed lateral process; ventral process straight; apical process convex. Proctiger with tergum X somewhat triangular in outline, inner process pointed. Paraproct elongate, crown with 9 or 10 simple blades. Cercal sclerite with 1 seta.e. Figs.\u00a0h, 14 HeaRemarksCulex longistylus n. sp. differ from the adults of Cx. atratus in having dark-scaled wings, occasionally with an inconspicuous patch of white scales on the base of vein C, and dark-scaled terga III-VII with white basal bands. Based on male genitalia, Cx. longistylus n. sp.\u00a0can be distinguished from the other species of the Atratus Group in possessing fine, subapically bifid setae interspersed with simple setae on tergum IX lobes, a long columnar process of the distal division with a large, broad and ribbed seta, lateral plate of the phallosome with a straight ventral process, and 3 short filiform setae on the ventromesal surface of the gonocoxite.Adults of Culex(Melanoconion)loturusDyar, 19251925Culex (Melanoconion) loturus Dyar, 1925: 241 [925: 241  (\u2642) holoCulex (Melanoconion) loturus of Dyar (1928: 342) [Cx. zeteki). of Dyar 28: 342 [28: 342)  , in poor condition, with associated genitalia on slide (USNM no. 30-1) deposited in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Distribution:Culex loturus was collected in Venezuela at the margin of the Catatumbo River [bo River .DescriptionMale. [Fig.\u00a0Cx. zeteki, except as follows: Genitalia: distal division of subapical lobe with median columnar process; seta l large, long, asymmetrical, ribbed; aedeagus with apical process of lateral plate pointed, without ripples.le. Fig.\u00a0 EssentiaFemale, pupa and larva. Unknown.RemarksCulex loturus was described by Dyar [Cx. loturus and in Cx. zeteki he mentioned the presence of two appendages. Komp [Cx. loturus with Cx. zeteki Dyar [Cx. loturus which are identical to those of Cx. zeteki. Although Cx. loturus bears three setae on the proximal division, this species can be distinguished from Cx. zeteki in possessing a large subapical seta l on the distal division, and a slender apical process of the lateral plate that lacks ripples. by Dyar  based on by Dyar  mentionees. Komp  synonymieki Dyar  based onCulex(Melanoconion)spiniferS\u00e1 & Sallum n. sp.Type locality: Pariquera-A\u00e7u Municipality , S\u00e3o Paulo State, Brazil. Adults were collected in the southeastern Atlantic Forest.Type material: Holotype, pinned adult male with dissected genitalia on slide (accession no. FSP-USP E-15901), with following collection data: Brazil, S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality , coll. Forattini et al. 6.iii.1980, det. Sallum 1980, deposited in the Cole\u00e7\u00e3o Entomol\u00f3gica de Refer\u00eancia, Faculdade de Sa\u00fade P\u00fablica, Universidade de S\u00e3o Paulo (FSP-USP), S\u00e3o Paulo, S\u00e3o Paulo State, Brazil. Paratypes: 1 pinned adult male with dissected genitalia on slide (accession no. FSP-USP E-15902) from the same collection as the holotype; 1 pinned adult male with dissected genitalia on slide (accession no. FSP-USP E-15903), coll. Forattini et al. 19.iv.1979, det. Sallum 1980; 1 pinned adult male with dissected genitalia on slide (accession no. FSP-USP E-15904), coll. Forattini et al. 6.ix.1980, det. Sallum 1980, all deposited in FSP-USP.ZooBank registration: The Life Science Identifier (LSID) for Culex (Melanoconion) spinifer n. sp. is urn:lsid:zoobank.org:act:4C1F6F37-F488-4EBF-9F69-3523902A29F1.Etymology: From the Latin adjective spinifer meaning spiny. Culex spinifer is named in reference to the spicules present on the ventral process of the lateral plate of the aedeagus.DescriptionMale. [Figs.\u00a0Head: antennal length 1.41\u20131.71 (1.59) (n\u2009=\u20094); proboscis dark-scaled, with inconspicuous median, dorsal patch of whitish scales; proboscis length 1.41\u20131.66 (1.52) (n\u2009=\u20094); maxillary palpus dark-scaled, length 2.28\u20132.05 (2.15) (n\u2009=\u20094); palpomere II with inconspicuous basal patch of whitish scales; palpomere III with small basal patch of whitish scales; palpomere IV with inconspicuous basal patch of whitish scales; palpomere V dark-scaled, with long, strong setae. Occiput with dark brown forked erect scales. Thorax: integument dark brown; scutum with narrow, dark brown forked scales, mainly on median prescutellar area and median scutal fossa; with whitish scales on anterior promontory and other prescutellar areas. Median scutellar lobe with 6 large, dark setae; lateral lobes each with 3 or 4 setae. Pleural setae with 2 types of colouring: dark brown: 3\u20136 antepronotal, 3 or 4 prealar; and pleural pale golden, slender setae: 4 or 5 upper mesokatepisternal, 4 or 5 lower mesokatepisternal, 4 or 5 upper mesepimeral; lower mesepimeron with 1 strong, long seta. Pleura with distinct patch of broad, white scales on upper mesokatepisternum; lower mesokatepisternum with few white scales. Wing: dark-scaled, with inconspicuous basal patch of whitish scales on vein C; large basal patch of whitish scales on vein R; wing length 2.93\u20132.65 (2.79) (n\u2009=\u20094). Halter: scabellum, pedicel and capitellum whitish. Legs: coxae pale; ventral surface of fore- and midfemur with a longitudinal stripe of white scales; tibiae dark-scaled; joints of femur-tibia and tibia-tarsomere I with ring of pale scales; tarsi entirely dark-scaled. Abdomen: tergum I with dark scales; terga III-VII dark-scaled, with basal bands of white scales. Genitalia: tergum IX lobes elongate, each with 7\u201310 slender simple setae and few apically bifid setae in median portion, apex glabrous. Distance between lobes equivalent to basal width of 1 lobe. Gonocoxite oblong, small; subapical lobe divided into 2 columnar divisions; proximal division with 2 pointed setae (a and b); seta a shorter, slender, inserted basal to seta b; seta b spatulate, robust and stronger than seta a; gonocoxite with 2 or 3 short, pointed, hyaline setae on ventromesal surface; distal division with short columnar process, with 5 setae: 3 narrow filiform, apically pointed setae of different in sizes (seta f), 1 longer hook-like seta (seta h), and 1 short, broad, asymmetrical seta arising subapically (seta l); 1 saber-like seta (seta s) arising apically. Gonostylus with broad leaf-like gonostylar claw with pointed apex, arising apically. Aedeagus with ventral process slightly convex and with spicules. Proctiger with tergum X with slightly pointed inner process.e. Figs.\u00a0i, 16 HeaRemarksCulex spinifer n. sp. has spicules on the ventral process of the lateral plate similar to Cx. dunni. However, Cx. spinifer n. sp.\u00a0differs from Cx. dunni in having elongate and slender ninth tergal lobes. Moreover, it has only two filiform setae on the proximal division of the subapical lobe and a somewhat triangular-shaped tergum X. Culex spinifer n. sp.\u00a0differs from Cx. comptus\u00a0n. sp., Cx. longisetosus n. sp.\u00a0and Cx. longistylus n. sp.\u00a0by having a short columnar process on the proximal division of the subapical lobe, a broad seta l, however shorter than filaments of seta f, and a large and broad gonostylar claw. Additionally, adults of Cx. spinifer n. sp.\u00a0differ from those of Cx. caribeanus and Cx. trigeminatus in having the femora without pre-apical whitish rings and palpomere II with an inconspicuous proximal patch of whitish scales.Culex(Melanoconion)trigeminatusClastrier, 19701970Culex (Melanoconion) trigeminatus, Clastrier 1970: 473 [970: 473  (\u2642) holoCulex (Melanoconion) trigeminatus of Pecor et al. (1992: 27) [992: 27)  (distr.)992: 27)  (distr.)Type material: Holotype, pinned adult male from For\u00eat du Gallion, French Guiana, collected on 19\u201320.iv.1968 , deposited in the Museum National d\u02bcHistoire Naturelle (MNHN), Paris, France.Material examined: 56 specimens: 40 G\u2642, 24 Le, 37 Pe. FSP-USP, Brazil: S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, Bra\u00e7o Magro, coll. S\u00e1 et al. 2014, 20.viii.2014, det. S\u00e1 2014: SP152-01 ; SP152-02 ; SP152-03 ; SP152-04 ; SP152-05 ; SP152-09 ; SP152-12 ; SP152-14 ; SP152-15 ; SP152-19 ; SP152-100 ; SP152-101 ; SP152-102 ; SP152-104 ; SP152-105 ; SP152-106 ; SP152-107 ; SP152-108 . S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, road to Bra\u00e7o Magro farm, Lagoon in forest environment, coll. S\u00e1 et al. 2014, 16.ix.2014, det. S\u00e1 2014: SP157-03 ; SP157-08 ; SP157-10 ; SP157-12 ; SP157-16 ; SP157-18 ; SP157-19 ; SP157-101 ; SP157-104 ; SP157-107 . S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, road to Bra\u00e7o Magro farm, stream on forest A, coll. S\u00e1 et al. 16.ix.2014, det. S\u00e1 2014: SP158A-01 ; SP158A-08 ; SP158A-09 ; SP158A-16 . S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, Bra\u00e7o Magro, Lagoon, coll. S\u00e1 et al. 2016, det. S\u00e1 2016: SP184-22 (\u2642G). S\u00e3o Paulo State, Canan\u00e9ia Municipality, Taquari , coll. Forattini et al. 25.iii.1980, det. Sallum 1980: no. 241 ; no. 225 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Itapu\u00e3 Farm , coll. Forattini et al. 6.iv.1981, det. Sallum 1981: no. 136 . S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality , coll. Forattini et al. 8.i.1981, det. Sallum 1981: no. 2242 ; no. 2491 . S\u00e3o Paulo State, Iguape Municipality , coll. Forattini et al. 6.x.1982, det. Sallum 1982: no. 3251 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Folha Larga Farm \u2212\u200924.89273, \u2212\u200947.919048), coll. Forattini et al. 19.iv.1983, det. Sallum 1983: no. 3470 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Vilarinho farm , coll. Forattini et al. 7.ii.1984, det. Sallum 1984: no. 3515 . S\u00e3o Paulo State, Canan\u00e9ia Municipality, Itapitangui , coll. Forattini et al. 11.iv.1985, det. Sallum 1985: EP035-01 , EP035-08 .Distribution:Culex trigeminatus has been collected in Brazil and French Guiana [h Guiana . In Brazh Guiana , Canan\u00e9ih Guiana , Par\u00e1 StDescriptionFemale.Head: antennal length 1.28\u20131.70 (1.46) (n\u2009=\u20095); proboscis dark-scaled, with median, dorsal patch of whitish scales, length 1.23\u20131.33 (1.28) (n\u2009=\u20095); maxillary palpus dark-scaled, length 0.21\u20130.25 (0.23) (n\u2009=\u20095). Occiput with erect, forked, pale brown scales. Thorax: scutum with narrow, dark brown to black falcate scales and narrow, whitish falcate scales on scutal fossa, dorsocentral, anterior promontory and supraalar areas forming a pattern. Scutellar scales whitish, median lobe with 5 or 6 setae, lateral lobes each with 3 setae. Pleural setae with 2 types of colouring: dark brown: 4\u20136 antepronotal, 4 or 5 prealar; and pleural setae golden: 4 upper mesokatepisternal, 3 or 4 lower mesokatepisternal, 4 upper mesepimeral, 1 large lower mesepimeral. Pleura with distinct patch of broad, whitish scales. Wing: dark-scaled, vein C with small proximal patch of whitish scales, vein R with large proximal patch of whitish scales; wing length 2.63\u20132.84 (2.67) (n\u2009=\u20095). Halter: scabellum, pedicel and capitellum pale brown. Legs: fore- and midfemur with conspicuous preapical ring of white scales. Abdomen: terga II-VII with basal bands of white scales, tergum VIII dark-scaled.Male. [Figs.\u00a0Head: antenna verticillate, length 0.96\u20131.12 (1.11) (n\u2009=\u20095); proboscis dark-scaled, with median patch of whitish scales, proboscis length 1.47\u20131.63 (1.53) (n\u2009=\u20095); maxillary palpus length 1.71\u20132.28 (1.96) (n\u2009=\u20095), palpomere III with basal patch of whitish scales; palpomeres IV and V with small basal patch of whitish scales. Wing: length 2.32\u20132.81 (2.51) (n\u2009=\u20095). Genitalia: tergum IX lobes with convex outer edge, apex glabrous, median portion each with 20\u201322 slender, simple setae; distance between lobes shorter than half basal width of 1 lobe. Gonocoxite narrow, oblong; proximal division with 4 parallel setae : seta a more basal, spoon-shaped; seta b robust, spatulate, inserted on tubercle; seta c thin, slender, filiform, inserted between setae b and d; seta d borne on tubercle apical to seta b, filiform, long, with slightly narrowed apex. Distal division with medium-sized, elongate columnar process, with 5 setae: 3 filiform, narrow, pointed, apically inserted, subequal sized (seta f), 1 filiform, hook-like apex (seta h), and 1 large, broad, asymmetrical ribbed seta with apex slightly pointed on median portion, arising subapically (seta l); and 1 saber-like, ribbed seta (seta s) arising apically. Gonocoxite with slender, hyaline, short, inconspicuous setae on ventromesal surface. Gonostylus as in Cx. atratus, except for dorsal surface of the apex which may bear 2 or 3 superficial, inconspicuous emarginations. Lateral plate of aedeagus with rounded apical process, ventral process with short pointed projection directed ventrobasally. Proctiger with tergum X somewhat triangular in outline, inner process pointed and short.e. Figs.\u00a0j, 17 EssPupa. [Figs.\u00a0Cephalothorax: seta 4-CT 3-branched; seta 5-CT 4-branched; seta 8-CT 6-branched; seta 12-CT 3- or 4- branched. Trumpet long, slender; pinna small, opening circular, pinna length 0.05\u20130.08 (0.06) (n\u2009=\u20099), distal margin opposite meatal cleft with small notch; tracheoid area extending 0.15\u20130.25 (0.22) (n\u2009=\u20099) from base; trumpet index 14.8\u201330.5 (20.9) (n\u2009=\u20099). Abdomen: seta 9-VIII with 2 simple branches; paddle index 1.40\u20131.71 (1.50) (n\u2009=\u20099).a. Figs.\u00a0e, 4e CepLarva. [Figs.\u00a0Head: length 0.65\u20130.73 (0.69) (n\u2009=\u20099), width 1.05\u20131.12 (1.09) (n\u2009=\u20099). Antennal length 0.52\u20130.63 (0.56) (n\u2009=\u20099); seta 1-A inserted 0.39\u20130.46 (0.41) (n\u2009=\u20099) from antennal base. Seta 5-C with 5 short branches not reaching 6-C insertion; seta 10-C 4-branched; seta 13-C 3-branched. Abdomen: comb of segment VIII with 18\u201322 scales of different sizes arranged in 2 or 3 irregular rows: upper rows with small, pointed scales; lower row with 7\u20139 large, pointed scales. Segment X length 0.30\u20130.36 (0.33) (n\u2009=\u20099), siphon/saddle index 4.62\u20135.28 (4.88) (n\u2009=\u20099). Siphon: long, slender, index 7.7\u201313.0 (10.8) (n\u2009=\u20099); pecten with 13 spines on basal 0.30 of siphon. Seta 1-S usually with 4 ventral pairs and 2 dorsal pairs.a. Figs.\u00a0e, 6e HeaBionomics. Immatures of Cx. trigeminatus were collected in large, shaded lagoons with aquatic vegetation and, in small, flooded, shaded depressions and floodplain terraces of streams in Atlantic Forest, associated with Cx. albinensis and Cx. zeteki.RemarksCulex trigeminatus was described by Clastrier [Culex trigeminatus is more closely related to Cx. caribeanus within the Atratus Group, especially regarding adult specimens. However, Cx. trigeminatus differs from Cx. caribeanus in having palpomeres I and II dark-scaled, palpomere III with small basal whitish patch, palpomeres IV and V with inconspicuous whitish basal patches, and wings with proximal patches of white scales on veins C and R. The male genitalia of Cx. trigeminatus differ from those of Cx. caribeanus by having the median portion of the apex of seta l  slightly pointed, robust and with blunt apex and proximal division with seta b strong, the lateral plate of the aedeagus having a rounded apical process and a ventral process with short, pointed projection directed ventrobasally. Fourth-instar larvae of Cx. trigeminatus differ from those of the other species of the Atratus Group by having seta 5-C with short branches that do not reach the insertion of seta 6-C and a siphon with only two pairs of dorsal setae. Furthermore, Cx. trigeminatus differs from Cx. ensiformis in possessing strongly serrated comb scales and short pecten spines. With respect to pupae, Cx. trigeminatus can be distinguished from the other species by having a slender trumpet and with the pinna small and appearing heart-shaped in dorsal view.lastrier  based onCulex(Melanoconion)zetekiDyar, 19181918Culex (Melanoconion) zeteki Dyar, 1918: 122 [Cx. zeteci) deposited in the USNM. Type locality: Gat\u00fan, Canal Zone, Panama.918: 122  holotypeCulex (Melanoconion) zeteki of Dyar (1928: 339) [Cx. zeteci); Komp (1935: 9) [Cx. zeteci); Rozeboom & Komp (1950: 98) [Cx. zeteki); Pecor et al. (1992: 56) [ of Dyar 28: 339 [1935: 9)  (\u2642 as Cx950: 98)    (distr.)992: 56)   with dissected genitalia (USNM no. 953) on slide, in poor condition, deposited in the Diptera Collection, National Museum of Natural History (USNM), Washington, DC, USA.Material examined: 37 specimens: 15 Le, 20 Pe, 10 \u2640, 10 \u2642. (FSP-USP): Brazil: S\u00e3o Paulo State, Pariquera-A\u00e7u Municipality, Pariquera-Mirim , coll. S\u00e1 et al. 2014, 21.viii.2014, det. S\u00e1 2014: SP155-29 . S\u00e3o Paulo, Pariquera-A\u00e7u, coll. S\u00e1 et al. 2014, 16.ix.2014, det. S\u00e1 2014: SP157-105 ; SP158A-04 ; SP158A-05 ; SP158A-06 ; SP158A-07 . Minas Gerais State, Cl\u00e1udio Municipality, Marcelo Farm, V\u00e1rzea da Rocinha , coll. Bergo et al. 2010, 13.iv.2010, det. Sallum 2014: MG50-10 . S\u00e3o Paulo State, Dourado Municipality , coll. Forattini et al. 7.i.1981, det. Sallum 1981: no.147 (\u2642G); no.150 ; no.151 (\u2642G); no.152 ; no.156 ; no.157 (\u2642G). S\u00e3o Paulo, Canan\u00e9ia Municipality, Iririaia-A\u00e7u , coll. Forattini et al. 18.i.1984, det. Sallum 1984: HEP352-07 . S\u00e3o Paulo, Canan\u00e9ia, Itapitangui , coll. Forattini et al. 11.iv.1984, det. Sallum 1984: HEP387-01 ; HEP387-03 ; HEP387-04 ; HEP394-05 . S\u00e3o Paulo, Canan\u00e9ia, Folha larga , coll. Forattini et al. 26.vii.1985, det. Sallum 1985: EP0005-01 ; EP0005-02 ; EP0005-04 (Pe); EP0005-05 ; EP0005-06 ; EP0005-07 ; EP0005-09 . S\u00e3o Paulo, Canan\u00e9ia, Itapitangui , coll. Forattini et al. 25.x.1988, det. Sallum 1988: EP0003-06 . Amazonas State, Humait\u00e1 Municipality, Realidade , coll. Chaves et al. 22.vii.2016, det. S\u00e1 2017: Coleta07-Humait\u00e1-03 (\u2640); Coleta07-Humait\u00e1-04 (\u2640); Coleta07-Humait\u00e1-05 (\u2640); Coleta07-Humait\u00e1-06 (\u2640); Coleta07-Humait\u00e1-74 (\u2640). Amazonas State, L\u00e1brea Municipality, Umari  coll. Sallum et al. viii.2015, det. S\u00e1 2017: AM47-06 (\u2642). (INPA): Amazonas State, Manaus Municipality, Acampamento Colosso, Fazenda Esteio , coll. Hutchings & Aquino 2002, det. Sallum & Hutchings: Fam-000630 (\u2642G); Fam-000939 (\u2642G); Fam-002679 (\u2642G). Amazonas State, Jutai Municipality, S\u00e3o Raimundo, Parana do Cervalho, Solim\u00f5es River , coll. Hutchings et al. 16\u201317.ix.2003, det. S\u00e1 2017: ProV-007250 (\u2642G); ProV-007254 (\u2642G).Distribution:Culex zeteki has been found in Belize [n Belize , Brazil n Belize , Colombin Belize , French n Belize , Honduran Belize , Nicaragn Belize , Paraguan Belize , Panama n Belize , Surinamn Belize , 15, 69 n Belize , 70, 76,n Belize , Minas Gn Belize  and S\u00e3o n Belize , 81.DescriptionFemale.Head: antennal length 0.81\u20131.31 (1.08) (n\u2009=\u20095); proboscis dark-scaled, length 1.07\u20131.33 (1.20) (n\u2009=\u20095); maxillary palpus dark-scaled, length 0.16\u20130.26 (0.22) (n\u2009=\u20095). Occiput with brown, erect, forked scales. Thorax: scutum with narrow, bronzy falcate scales. Median lobe of scutellum with 6 setae, lateral lobes each with 4 setae. Pleural setae with 2 types of colouring: dark brown: 4\u20136 antepronotal, 4 or 5 prealar; and pleural setae golden: 3 or 4 upper mesokatepisternal, 3 or 4 lower mesokatepisternal, 4 upper mesepimeral, 1 large lower mesepimeral. Pleura with indistinct broad patch of whitish scales. Mesepimeral integument dark, with distinct median whitish area completely separating darker upper and lower areas. Wing: dark-scaled, length 2.23\u20132.88 (2.55) (n\u2009=\u20095). Halter: scabellum, pedicel and capitellum whitish. Legs: as in Cx. atratus. Abdomen: tergum II-VII with basal bands of white scales, tergum VIII dark-scaled.Male. [Figs.\u00a0Head: antennal length 0.96\u20131.32 (1.15) (n\u2009=\u20095); proboscis entirely dark-scaled, length 1.24\u20131.81 (1.63) (n\u2009=\u20095); maxillary palpus dark-scaled, length 1.68\u20132.19 (1.85) (n\u2009=\u20095). Wing: length 2.28\u20132.74 (2.64) (n\u2009=\u20095). Genitalia: tergum IX lobes elongate, each with 20\u201322 slender, simple setae on median portion, apex glabrous, slightly pointed. Distance between lobes shorter than basal width of 1 lobe. Gonocoxite oblong, narrow; proximal division with 3 long, parallel setae : seta a long, slender with \u201copened\u201d apex; seta b, long with rounded apex; seta c slender, filiform, with curved apex. Distal division with median columnar process with 5 setae: 3 filiform, narrow, apically pointed and differently sized setae (setae f), 1 long seta hook-like at apex (seta h), and 1 large, long, asymmetrical seta arising subapically (seta l); 1 saber-like seta (seta s) arising apically; gonocoxite with short, inconspicuous, hyaline setae on ventromesal surface. Gonostylus as in Cx. atratus, except for dorsal surface of apex with 3 or 4 conspicuous folds and large gonostylar claw. Aedeagus with apical process with rounded ripples; ventral process slightly straight. Proctiger with tergum X asymmetrical, with rounded outer and inner processes.e. Figs.\u00a0k, 18 EssPupa. [Figs.\u00a0Cephalothorax: seta 5-CT 4-branched; seta 7-CT double. Trumpet slender; pinna small, asymmetrical, length 0.09\u20130.15 (0.12) (n\u2009=\u200910), distal margin opposite the meatal cleft with small, inconspicuous emargination; tracheoid area extending 0.15\u20130.21 (0.18) (n\u2009=\u200910) from base; trumpet index 12.0\u201320.0 (15.9) (n\u2009=\u200910). Abdomen: seta 9-VIII with 3 simple branches; paddle index 1.57\u20132.03 (1.69) (n\u2009=\u200910).a. Figs.\u00a0f, 4f CepLarva. [Figs.\u00a0Head: length 0.66\u20130.77 (0.71) (n\u2009=\u200910), width 1.04\u20131.13 (1.09) (n\u2009=\u200910). Antennal length 0.49\u20130.57 (0.53) (n\u2009=\u200910); seta 1-A inserted 0.35\u20130.39 (0.37) (n\u2009=\u200910) from antennal base. Seta 5-C with 4 long branches; seta 13-C double; seta 14-C with 2 strong branches. Abdomen: comb of segment VIII with 28\u201334 scales equal in size arranged in 3 or 4 irregular rows. Segment X length 0.30\u20130.37 (0.33) (n\u2009=\u200910), siphon/saddle index 3.90\u20134.88 (4.43) (n\u2009=\u200910). Siphon: long, slender, index 6.5\u20138.9 (8.1) (n\u2009=\u200910); pecten with 12 spines on basal 0.30 of siphon. Seta 1-S usually with 4 ventral pairs and 4 dorsal pairs.a. Figs.\u00a0f, 6f HeaBionomics. Immature specimens of Cx. zeteki were collected in shaded, stagnant lagoons with abundant aquatic vegetation.RemarksCulex zeteki was described by Dyar [Cx. zeteci. Rozeboom & Komp [Cx. zeteki. Culex zeteki differs from Cx. columnaris n. sp.\u00a0in having the gonostylus with folds on the dorsal surface. Fourth-instar larvae of Cx. zeteki can be distinguished by having comb scales with lateral fringes on the middle of the lateral margins and seta 5-C with 4 or 5 long branches which may reach seta 7-C insertion. Pupae of Cx. zeteki can be distinguished from the other species of the group by possessing a slender trumpet with a small pinna and having the distal margin opposite the meatal cleft with an inconspicuous rounded emargination. by Dyar  as Cx. zm & Komp  correcteThe primary diagnostic characters of the larval and pupal forms are summarized in Tables\u00a0Keys for identification of the Atratus Group and the species of the group(i)Keys based on adult morphology1Vertex with narrow, curved or linear decumbent scales; broad decumbent scales restricted to small lateral patches\u20262\u2013Vertex with broad decumbent scales\u2026Melanoconion Section (in part)2(1)Vertex with few narrow decumbent scales restricted to median area; lateral patch of broad decumbent scales large, evident in dorsal view\u20263\u2013Vertex with numerous narrow decumbent scales; lateral patch of broad decumbent scales small, almost indistinct in dorsal view\u2026Spissipes Section (in part)3(2)Thoracic pleural integument yellowish or lighter, contrasting with brown scutal integument\u2026Spissipes Section\u2013Thoracic pleural integument similar in color or lighter, not contrast sharply with scutal integument\u202644(3)Thoracic pleural integument lighter than scutal integument, with pattern of dark and pale areas on mesepimeron and mesokatepisternum; upper mesokatepisternum with a patch of white scales; legs with conspicuous or inconspicuous ring of white scales at all femur-tibia joints\u2026Atratus Group (Melanoconion Section)\u2013Thoracic pleural integument without pattern of dark and pale areas; upper mesokatepisternum without or with a small patch or a few white scales; legs with or without ring or patch of white scales at femur-tibia joints\u2026Other Groups (Melanoconion Section)Atratus GroupProboscis with distinct dorsal and median patches of whitish scales; fore- and midfemur with preapical ring of whitish scales Fig.\u00a0a\u20262Proboscis dark-scaled or with indistinct patch of pale scales; fore- and midfemur without preapical ring of whitish scales Fig.\u00a0b\u20263Fig.\u00a01Cx. caribeanusCosta (C) dark-scaled; vein R entirely dark-scaled. Male: palpomere I pale-scaled; palpomere III with long, distinct median patch of whitish scales on dorsal surface Fig.\u00a0a\u2026Cx. carCx. trigeminatusCosta (C) with proximal patch of whitish scales; vein R with a long line of whitish scales proximally. Male: palpomere I entirely dark-scaled; palpomere III with small, basal patch of whitish scales Fig.\u00a0b\u2026Cx. triCx. spinifer n. sp.Proboscis with indistinct dorsomedian patches of whitish scales; costa (C) with inconspicuous proximal patch of whitish scales; vein R with 1 long basal patch of whitish scales. Male: palpomere III with inconspicuous proximal patch of whitish scales Fig.\u00a0a\u2026Cx. spiProboscis without dorsal patch of whitish scales; vein R dark-scaled or with 2 basal patches of whitish scales separated by a dark scale-patch Fig.\u00a0b\u20264Fig.\u00a02Vein R with 2 basal whitish patches separated by dark-scaled patch Fig.\u00a0a\u20265Vein R entirely dark-scaled Fig.\u00a0b\u20267Fig.\u00a02Scutal integument dark brown/black with whitish and golden scales  with patch of broad whitish scales as long as the mesothoracic spiracle (MS) Fig.\u00a0a\u20268Upper corner of mesokatepisternum (Mkm) with patch of broad whitish scales shorter than the mesothoracic spiracle (MS) Fig.\u00a0b\u20269Fig.\u00a02Cx. longisetosus n. sp.Costa entirely dark-scaled Fig.\u00a0a. Male: Cx. longistylus n. sp.Costa with proximal whitish scale-patch Fig.\u00a0c. Male: Cx. zeteki / Cx. loturusMesepimeral integument (Mm) dark with distinct median whitish area completely separating darker upper and lower areas Fig.\u00a0a\u2026Cx. zetMesepimeral integument (Mm) entirely dark or with indistinct median whitish area not completely separating the dark areas Fig.\u00a0b\u202610Fig.\u00a0Cx. atratusMesepimeral integument (Mm) dark, without median pale area; terga II-VIII (II-VIII-Te) with basolateral patches of whitish scales, without basal pale bands Fig.\u00a0a\u2026Cx. atrCx. columnaris n. sp.Mesepimeral integument (Mm) dark, with indistinct median pale area that does not completely separate the dark area; terga II-VIII (II-VIII-Te) with pale basal bands Fig.\u00a0b\u2026Cx. col Keys based on the morphology of male genitaliaAedeagal sclerite (AeS) broad and curved in lateral view, largely connected to dorsal process of lateral plate (LP)\u2026Spissipes SectionAedeagal sclerite (AeS) narrow and curved in lateral view, lightly connected to dorsal process of lateral plate (LP)\u20262Gonocoxite (Gc) small, narrow, oblong; gonostylus (Gs) narrow, dorsal area without setae; lateral plate (LP) with concave ventral process and pointed apical process\u2026Atratus Group (Melanoconion Section)Gonocoxite (Gc) conical, ovoid or globose; gonostylus variously modified; lateral plate (LP) with ventral and lateral processes otherwise modified\u2026Other Groups (Melanoconion Section)Atratus GroupProximal division of subapical lobe (pSL) without columnar process, setae only inserted on tubercles or directly inserted on surface of the gonocoxite Fig.\u00a0a\u20262Cx. columnaris n. sp.Proximal division of subapical lobe (pSL) with long columnar process divided apically Fig.\u00a0b\u2026Cx. colProximal division of subapical lobe (pSL) with 2 setae Fig.\u00a0a\u20263Proximal division of subapical lobe (pSL) with 3 or more setae Fig.\u00a0b\u20268Fig.\u00a03b of proximal division) columnar process  with a short  columnar process  with a long   with seta th) Fig.\u00a0a; tergumth) Fig.\u00a0b; gonocoth) Fig.\u00a0c\u20265l long   with seta th) Fig.\u00a0d; tergumth) Fig.\u00a0e; gonocoth) Fig.\u00a0f\u2026Cx. atrCx. ensiformisTergum IX lobe (IX-TL) with simple setae Fig.\u00a0a; aedeagCx. spinifer n. sp.Tergum IX lobe (IX-TL) with simple and apically bifid setae Fig.\u00a0d; aedeagTergum IX lobe (IX-TL) elongate, longer than wide, with only simple setae or simple and few apically bifid setae Fig.\u00a0a\u20267Cx. longistylus n. sp.Tergum IX lobe (IX-TL) somewhat rounded, wider than long, with few simple and several apically bifid setae Fig.\u00a0b\u2026Cx. lonCx. comptus n. sp.Tergum IX lobe (IX-TL) slender, with simple setae Fig.\u00a0a; aedeagCx. longisetosus n. sp.Tergum IX lobe (IX-TL) slightly broad, with simple, apically bifid setae Fig.\u00a0d; aedeagLateral plate (LP) of aedeagus with spicules on ventral process Fig.\u00a0a; tergumLateral plate (LP) of aedeagus without spicules on ventral process Fig.\u00a0c; tergumCx. dunniGonocoxite (Gc) with long and dispersed setae on sternomesal surface Fig.\u00a0a; tergumCx. exedrusGonocoxite (Gc) with long and aligned setae on sternomesal surface Fig.\u00a0c; tergumProximal division of subapical lobe (pSL) with 3 setae Fig.\u00a0a; apicalProximal division of subapical lobe (pSL) with 4 setae Fig.\u00a0c; lateral of distal division of subapical lobe (dSL) with rounded and laterally directed apex  of aedeagus with distinct ripples on apical process Fig.\u00a0a; seta lpex Fig.\u00a0b; gonostpex Fig.\u00a0c\u2026Cx. zetl of distal division of subapical lobe (dSL) with pointed and laterally directed apex  of aedeagus with indistinct ripples on apical process Fig.\u00a0d; seta lpex Fig.\u00a0e; gonostpex Fig.\u00a0f\u2026Cx. lotl of distal division of subapical lobe (dSL) with rounded apex; seta b of proximal division of subapical lobe (pSL) long, slender, apically sinuous  with pointed apex; seta b of proximal division of subapical lobe (pSL) long and stout Keys based on pupal morphology1Seta 9-VIII inserted at or near to caudolateral margin; caudolateral angle of segment VIII blunt\u2026Spissipes Section\u2013Seta 9-VIII inserted above of caudolateral margin; caudolateral angle of segment VIII slightly pointed\u202622(1)Trumpet narrow, long, length/width ratio 7\u00a0to\u00a030\u2026Atratus Group (Melanoconion Section)\u2013Trumpet thick, of shorter length, length/width ratio 5\u00a0to\u00a08\u2026Other Groups (Melanoconion Section)Atratus GroupTrumpet (T) with distal margin of pinna (Pi) without emargination; if present, emargination indistinct Fig.\u00a0a\u20262Trumpet (T) with distal margin of pinna with conspicuous emargination Fig.\u00a0b\u20264Fig.\u00a04Cx. atratusTrumpet (T) with a V-shaped pinna (Pi); trumpet large, wider at apex than at base Fig.\u00a0a; trumpePinna (Pi) small, not V-shaped; trumpet narrow from base to apex Fig.\u00a0c; trumpeCx. trigeminatusTrumpet index 20; pinna (Pi) short, somewhat rounded, meatal cleft (MC) short; distal margin opposite meatal cleft with small notch Fig.\u00a0a\u2026Cx. triCx. zetekiTrumpet index 17; pinna (Pi) heart-shaped in lateral view, becoming slender at base, meatal cleft (MC) long; distal margin opposite meatal cleft with small, rounded emargination Fig.\u00a0b\u2026Cx. zetCx. ensiformisTrumpet (T) long; pinna (Pi) cup-shaped in lateral view; trumpet index > 20; distal margin opposite meatal cleft (MC) with indistinct, shallow transverse depression Fig.\u00a0a\u2026Cx. ensTrumpet (T) moderately long; pinna (Pi) narrow; trumpet index < 16; distal margin opposite meatal cleft with indistinct longitudinal notch Fig.\u00a0b\u20265Fig.\u00a04c.14\u2026Cx. dunniTrumpet (T) widened distally; distal margin opposite meatal cleft with large emargination Fig.\u00a0a; trumpec.15\u2026Cx. comptus n. sp.Trumpet (T) narrow; distal margin opposite meatal cleft with indistinct longitudinal fissure Fig.\u00a0b; trumpe(iv)Keys based on fourth-instar larvae morphology1Seta 2-C present; seta 14-C inserted anteriorly to 15-C\u2026Spissipes Section \u201cpartim\u201d\u2013Seta 2-C absent; seta 14-C and 15-C inserted at same level or 14-C slightly anterior to 15-C\u202622(1)Siphon slender and long, index 7\u201310; seta 1-S with 2\u20134 short pairs of dorsolateral setae; thoracic and abdominal integument without spicules; segment X with few spicules on posterior margin\u2026Atratus Group (Melanoconion Section)\u2013Siphon thick and short, index lower than 7; seta 1-S with 2 short or long pairs of dorsolateral setae; thoracic and abdominal integument with distinct or indistinct spicules; segment X with several spicules on posterior margin\u2026Other Groups (Melanoconion Section)Atratus GroupComb scales (CS) of segment VIII different in size and shape: long, pointed and laterally fringed, and shorter apicolaterally fringed scales Fig.\u00a0a\u20262Comb scales (CS) of segment VIII similar in size and shape Fig.\u00a0b\u20263Fig.\u00a04Cx. trigeminatusSeta 5-C with 5 short branches not reaching 6-C insertion; seta 13-C triple Fig.\u00a0a; siphonCx. ensiformisSeta 5-C with 8 long branches extending beyond 6-C insertion; seta 13-C double Fig.\u00a0d; siphonSegment VIII with comb scales in 3 rows Fig.\u00a0a; seta 1Segment VIII with comb scales in 4 rows Fig.\u00a0c; seta 1Cx. atratusSeta 13-C simple Fig.\u00a0a; pectenCx. comptus n. sp.Seta 13-C double Fig.\u00a0c; pectenCx. zetekiSeta 2-VIII with 2 or 3 branches; seta 5-VIII with 3 branches Fig.\u00a0a; comb sCx. dunniSeta 2-VIII with 1 or 2 branches; seta 5-VIII with 3 or 4 branches Fig.\u00a0c; comb sCx. atratus, Cx. caribeanus, Cx. commevynensis, Cx. dunni, Cx. ensiformis, Cx. trigeminatus, and Cx. zeteki. Adults of the group can be readily identified by having the vertex with narrow, decumbent scales restricted to the central area, a pleural integument with a striking pattern of dark and pale areas, and a patch of white scales on upper mesokatepisternum. In the male genitalia, the aedeagal sclerite is slender and curved in lateral view, the gonocoxite is oblong and narrow, and the gonostylus is narrow, simple and tapering to the apex. The trumpet of pupae is thin, long with a length/width ratio of 10 or higher. The fourth-instar larvae have a long, slender and tapering to apex siphon, with 3 or 4 pairs of small dorsolateral setae 1-S.The Atratus Group proposed by Sirivanakarn  includedCx. atratus is 7.0 and Cx. trigeminatus larvae possess only 2 pairs of dorsolateral setae on the siphon. These characteristics disagree with the diagnosis for the group provided by Sirivanakarn [Cx. atratus and Cx. trigeminatus from the other species of the Group, being diagnostic of these two species.Thus, according to Sirivanakarn , immaturvanakarn  but theyCx. theobaldi (Lutz).In addition, there are several characters that are useful for identification of species of the Atratus Group. Our comparative observations of adults of all examined species clearly indicate that the presence of a ring of white scales on all of the femoro-tibial joints, represents a character that helps recognize species of the group. However, other species of the Melanoconion Section also possess white scales on the knees, such as Culex exedrus can be distinguished from Cx. dunni by having several long setae visibly lined up on the sternomesal surface of the gonocoxite, seta s of gonocoxite with a large apex, and a proctiger with a large inner process of tergum X. Therefore, these features justify the resurrection of Cx. exedrus from synonymy with Cx. dunni. Similarly, Cx. loturus was resurrected from synonymy with Cx. zeteki based on the possession of a large seta l borne subapically on the distal division of the subapical lobe and a slender apical process without ripples of the lateral plate of the aedeagus.Furthermore, examination of the available material resulted in the discovery of five new species of the Atratus Group, based on morphological characters of adults, male genitalia and, where possible, of immature stages.Culex (Melanoconion) currently comprises 14 species and it has been markedly updated with respect to the number of known species, their bionomics and distribution, providing tools to facilitate the identification of the adult and immature stages of the species in the group. As a result, the current knowledge leads us to suggest the following composition of the Atratus Group: Cx. atratus , Cx. caribeanus, Cx. commevynensis, Cx. columnaris n. sp., Cx. comptus n. sp., Cx. dunni (syn. Cx. ruffinis), Cx. ensiformis, Cx. exedrus, Cx. longisetosus n. sp., Cx. longistylus n. sp., Culex loturus, Culex spinifer n. sp., Culex trigeminatus and Culex zeteki. Additional studies utilizing molecular methods, particularly to investigate phylogenetic relationships within the Atratus Group, and to determine the placement of the group within the genus Melanoconion, are necessary.The Atratus Group of"}
+{"text": "Gryllus bimaculatus, is registered as an edible insect by the Korean food and drug administration. The aim of this study was to investigate the effect of Gui-A-Gra on testicular damage induced by experimental left varicocele in male Sprague Dawley rats. A total of 72 rats were randomly divided into the following six groups (12 rats in each group): a normal control group (CTR), a group administrated with Gui-A-Gra 1.63 gm/kg (G1.63), a group administrated with Gui-A-Gra 6.5 gm/kg (G6.5), a varicocele (VC)-induced control group (VC), a VC-induced group administrated with Gui-A-Gra 1.63 gm/kg (VC + G1.63), and a VC-induced group administrated with Gui-A-Gra 6.5 gm/kg (VC + G6.5). Rats were administrated 1.63 or 6.5 gm/kg Gui-A-Gra once daily for 42 days. Indicators of sperm parameters, histopathology, reproductive hormones, oxidative stress, endoplasmic reticulum (ER) stress, inflammation, and mitochondrial apoptosis were analyzed to evaluate effects of Gui-A-Gra on VC-induced testicular dysfunction. Gui-A-Gra administration to VC-induced rats significantly (p < 0.05) increased sperm count and sperm motility, Johnsen score, spermatogenic cell density, serum testosterone, testicular superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase, GPx4, and steroidogenic acute regulatory protein (StAR) level. Moreover, pretreatment with Gui-A-Gra significantly (p < 0.05) decreased terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) positive cells/tubules, serum luteinizing hormone (LH), serum follicle-stimulating hormone (FSH), testicular tumor necrosis factor-\u03b1 (TNF-\u03b1), interleukin-6 (IL-6), malondialdehyde (MDA), reactive oxygen species (ROS)/reactive nitrogen species (RNS) level, glucose-regulated protein-78 (Grp-78), phosphorylated c-Jun-N-terminal kinase (p-JNK), phosphorylated inositol-requiring transmembrane kinase/endoribonuclease 1\u03b1 (p-IRE1\u03b1), cleaved caspase-3, and BCL2 associated X protein: B-cell lymphoma 2 (Bax: Bcl2) ratio in VC rats. These results suggest that protective effects of Gui-A-Gra on VC-induced testicular injury might be due to its antioxidant, anti-inflammatory, and androgenic activities that might be mediated via crosstalk of oxidative stress, ER stress, and mitochondrial apoptosis pathway.Gui-A-Gra, a commercial insect powder from  Varicocele (VC) is characterized by dilation and tortuosity of internal spermatic vein (pampiniform plexus), a network of veins that drains testicular tissues . VC is cOxidative stress and testicular apoptosis are well documented causes of testicular dysfunction ,3. PlasmGryllus bimarculatus powder that has recently been registered as an edible insect in Korea [Gryllus bimarculatus has pharmacological activity such as antioxidant and anti-inflammatory effects [Gryllus bimarculatus have beneficial antioxidant, hypocholesterolemic, and antimicrobial properties [G. bimaculatus include glycosaminoglycan which exhibits anti-inflammatory effects [Gryllus bimarculatus extract can prevent diabetes, liver disease, and arthritis [Gryllus bimarculatus on male reproductive functions has not been reported yet. Therefore, the objective of this study was to determine the protective effect of Gryllus bimaculatus powder for the recovery from testicular dysfunction and its associated male fertility condition in a VC-induced rat model. The roles of inflammation, oxidative stress, endoplasmic reticulum (ER) stress, and mitochondrial (intrinsic) pathway in VC testicular tissues were also examined to determine molecular mechanisms involved in the protective effect of Gryllus bimaculatus.It has been reported that antioxidant supplements can alleviate male infertility by scavenging ROS and modulating inflammatory responses and the antioxidant system . Gui-A-Gin Korea . Gryllus effects ,18. A reoperties . The con effects . Recent rthritis ,22,23. Hp < 0.05). Pretreatment with G 6.5 significantly increased sperm count in both vas deference and epididymis of VC rats (p < 0.05). Sperm motility in vas deference and epididymis in VC rats showed remarkable increases after pretreatment with G1.63 or G6.5 (p < 0.05)  . The fer < 0.05) .p < 0.05). Johnsen\u2019s score and spermatogenic cell density in the VC group were significantly (p < 0.05) decreased compared to those in the CTR group  as compared to the CTR group. However, serum LH and FSH levels were significantly decreased in VC + G1.63 and VC + G6.5 groups. Levels of IL-6 and TNF-\u03b1 were remarkably (p < 0.05) upregulated in the VC group as compared to the CTR group. Pretreatment with Gui-A-Gra significantly (p < 0.05) downregulated these parameters in the VC + G1.63 and VC + G6.5 groups (Serum testosterone level was significantly decreased in the VC group as compared to the CTR group . It was 5 groups .p < 0.05) enhanced MDA level and ROS/RNS level compared to rats in the CTR group. Treatment with G1.63 and G6.5 significantly (p < 0.05) restored these parameters in the VC group. VC-induced rats showed significant decreases of testicular levels of SOD, GPx, and catalase levels as compared to rats in the CTR group. These levels were remarkably (p < 0.05) increased after pre-treatment with G1.63 or G6.5. No significant difference in these parameters was observed between VC + G1.63 and VC + G6.5 groups.Results of lipid peroxidation parameters , ROS/RNS) and antioxidant enzymes  in testicular tissues are presented in p < 0.05) increases of GRP-78, p-IRE1\u03b1/ IRE1\u03b1, and p-JNK/JNK as compared to rats in the CTR group. In contrast, levels of GRP-78, p-IRE1\u03b1/IRE1\u03b1, and p-JNK/JNK levels in VC + G6.5 group were remarkably (p < 0.05) decreased as compared to those in the VC group. Levels of GRP 78 and p-JNK/JNK in VC + G1.63 group were lower than in the VC group, although differences between the two groups were not statistically significant. However, the p-IRE1\u03b1/IRE1\u03b1 level in VC + G1.63 group was significant decreased as compared to the VC group. Levels of pro-caspase-3 were similar among all groups. VC-induced rats showed prominently increased expression of cleaved caspase-3 and Bax:Bcl2 ratio compared to rats in the CTR group, although the increase was not statistically significant compared with the CTR group. In contrast, the cleaved caspase 3 and Bax:Bcl2 ratio in the VC + G6.5 group was significantly (p < 0.05) decreased as compared to the VC group : B-cell lymphoma 2 (Bcl2) ratio, StAR, and Gpx4 were analyzed in testis tissues by western blotting. Results are shown in VC group E. The clVC-induced rats showed significantly decreased expression levels of StAR as compared to rats in the CTR group. However, treatment with Gui-A-Gra restored levels of StAR in varicocelized rats. Cell-specific expression of StAR, Grp 78, and cleaved caspase 3 proteins in testis tissues was further analyzed by immunohistochemistry staining A\u2013C. A daThe present work used an experimental VC model to determine treatment effect of Gui-A-Gra on VC. This work extended studies on pathophysiological mechanisms associated with VC and suggested an ameliorative effect of Gui-A-Gra on VC. This study is first to report a protective effect of Gui-A-Gra on testicular dysfunction and male infertility using a VC-induced rat model. Pretreatment with Gui-A-Gra significantly improved sperm parameters, regulated levels of reproductive hormones, and promoted spermatogenesis in VC rats. Results of testicular morphological analysis, TUNEL assay, immunohistochemistry, and western blot analyses indicated that Gui-A-Gra could also decrease germ cell apoptosis.Previous studies have shown testicular hypotrophy in patients with clinical VC, but not in patients with subclinical VC . In the In our study, VC-induced rats showed degenerative changes in mammalian testis such as testicular atrophy, apoptosis of germ cell, aberration of STs, and impaired spermatogenesis. Similar findings have been reported in previous experimental VC studies ,2,3. TesLeydig cells are located in the interstitial tissue of the testis where testosterone is synthesized. They are essential for spermatogenesis . The StAIn the present study, higher concentrations of testicular MDA were detected in VC rats. The results of our study were similar to the MDA results of human testicular biopsy samples from infertile patients with VC . FurtherApoptosis is another detrimental factor involved in the impairment of sperm quality in patients with VC . IncreasGryllus bimarculatus powder material under the name of Gui-A-Gra used for the experiments was supplied from 239bio Inc. , a facility accredited for hazard analysis and critical control point (HACCP) and good manufacturing practice (GMP). Cultivated adult G. bimarculatus were subjected to 1-day of defecation before sacrifice. The G. bimarculatus were then cleansed with tap water three times, sterilized at 100 \u00b0C for 3\u20135 min, and dried for 6 h to have moisture less than 5%. Finally, dried G. bimaculatus were ground and homogenized. The powder was then stored at \u221220 \u00b0C for further use.A commercial Seventy-two 7\u20138 weeks old male Sprague Dawley rats were purchased from Koatech  and randomly divided into six groups (12 rats/group): (1) normal control (CTR) group; (2) Gui-A-Gra 1.63 gm/kg p.o. group (G1.63); (3) Gui-A-Gra 6.5 gm/kg p.o. group (G6.5); (4) varicocele (VC) group; (5) VC + Gui-A-Gra 1.63 gm/kg p.o. group (VC + G1.63); and (6) VC + Gui-A-Gra 6.5 gm/kg p.o. group (VC + G6.5). Rats were maintained in plastic cases (47 \u00d7 18 \u00d7 40 cm) (four rats per case) in controlled environment conditions . Rats had free access to water ad libitum and standard rat chow diet. All experiments were performed according to the Institutional Animal Care and Use Committee (IACUC) of Jeonbuk National University Hospital Laboratory Animal Center (cuh-IACUC-2017-13). After one week of acclimatization, left VC was induced for VC group, VC + G1.63 group, and VC + VC + G6.5 group of rats according to standard protocols . CTR and6 sperms/mL. Motile spermatozoa were analyzed with the formula of  \u00d7 100). Results for sperm motility are expressed as percentages.Epididymal and vas deference sperm count and sperm motility were evaluated as described previously . In brieTestis tissues were fixed in Bouin\u2019s solution, dehydrated, embedded in paraffin wax, and sectioned into 5 \u03bcm slices. Testis sections were subjected to hematoxylin and eosin (H&E) staining and TUNEL staining  as described previously . At leasSerum levels of testosterone , luteinizing hormone (LH) , and follicle-stimulating hormone (FSH)  were measured using enzyme-linked immunosorbent assay (ELISA) per each manufacturer\u2019s protocols.Whole tissue supernatants were prepared as described previously . TesticuMalondialdehyde (MDA) levels in testis tissues were assessed using reagents provided by Northwest Life Science Specialties LLC . Briefly, MDA reaction of a colored complex was measured at an absorbance of 532 nm using a kinetic spectrophotometer . MDA levels are expressed as \u03bcmole of MDA/mg of wet testis tissue. Testicular ROS/RNS levels were quantified using an OxiSelect in Vitro ROS/RNS Assay Kit  according to the manufacturer\u2019s protocol. Fluorescence of dichlorofluorescein (DFC) was measured with a SpectraMax Gemini XS fluorimeter  at excitation/emission wavelengths of 480/530 nm as described previously [Paraffin-embedded consecutive testis tissue sections (5 \u03bcm in thickness) were deparaffinized with xylene, dehydrated with alcohol, and subjected to heat-mediated 1\u00d7 target retrieval solution at pH 6.0 . Endogenous peroxide activity was blocked with peroxidase-blocking solution (DAKO) for 15 min and washed with 1\u00d7 PBS buffer twice (5 min each). Non-specific bindings of testis section were further blocked with serum block solution for 10 min (DAKO) and incubated with primary antibodies cleaved caspase 3 , StAR , and Grp 78  at 4 \u00b0C overnight. Sections were then washed with 1\u00d7 PBS twice (5 min each) and incubated with a horseradish peroxidase (HRP)-labeled micropolymer conjugated secondary antibody  at room temperature for 1 h. Prior to incubation, sections were rinsed with AEC substrate chromogens . Slides were then rinsed with deionized water for 5 min, counter-stained with hematoxylin, rinsed with distilled water, and mounted with mounting medium .Extraction of protein from testis tissue was conducted as described previously . Proteinp < 0.05. GraphPad PRISM 6.0 was used for graphical analysis .All quantitative data are expressed as mean \u00b1 standard error of the mean (SEM). Statistical analyses were performed using SPSS version 22 . One-way analysis of variance (ANOVA) was used to compare groups followed by Tukey\u2019s test for post-hoc analysis. Statistical significance was considered at In summary , our res"}
+{"text": "Enterobacterales and its association with plasmid mediated quinolone resistance (PMQR), aminoglycoside modifying enzymes (AME) and Trimethoprim/Sulfamethoxazole (TMP-SMX) resistance is little known from North India. Therefore, the current study was aimed to investigate the frequency of Non-\u03b2-Lactam antibiotic resistance associated genes in extended spectrum \u03b2-Lactamase producing Enterobacterales. For this study, Non-Duplicate phenotypically confirmed ESBL producing Enterobacterales isolates (N = 186) were analyzed for ESBLs, PMQRs, AMEs and TMP-SMX resistance genes using polymerase chain reaction (PCR). PCR detected presence of PMQR genes in 81.29% (N = 139) of ESBL isolates (N = 171), AME genes in 60.82% and TMP-SMX resistance genes in 63.74% of the isolates. Molecular characterization of ESBL producing Enterobacterales showed 84.79% TEMbla followed by 73.68% CTX-Mbla, 43.86% SHVbla, 19.88% PER andbla 9.94% VEBbla, respectively. Analysis of PMQR genes revealed 77.7% aac(6\u2032)-lb-cr the most commonly detected gene followed by 67.63% oqxB, 62.59% oqxA, 43.17% qnrB, 19.42% qnrD, 18.7% qnrS, 9.35% qnrA, 3.6% qepA and 2.88% qnrC, respectively. Analysis of AMEs gene profile demonstrated 81.73% aac(6\u2032)-Ib, the most frequently encountered gene followed by 46.15% aph(3\u2032)-Ia, 44.23% ant(3\u201d)-Ia, respectively. A 100% prevalence of sul1, followed by dfrA (54.63%) and sul2 (15.74%) was observed. In summary, prevalence of ESBL-Producing genes (particularly TEMbla and CTX-Mbla) along with PMQR, AMEs, and TMP-SMX resistant genes may potentially aid in the transfer of antimicrobial resistance among these strains.Genetic context of extended spectrum \u03b2-Lactamase (ESBL) producing  Enterobacterales [Enterobacterales and other clinically significant Gram-negative bacteria [Enterobacterales [Enterobacterales [qnr) gene mediated (ii) quinolones modifying aminoglycoside acetyltransferase encoding genes (aac(6\u2032)-Ib-c), (iii) plasmid-mediated quinolone efflux pumps qepA, oqxA, and oqxB [\u03b2-Lactam antibiotics are used for treating most of the human infections that are caused by Gram-negative bacteria belonging to the family cterales . Howevercterales . ESBLs abacteria ,2. Genesbacteria . Furthercterales . Chromoscterales ,5. Genercterales ,6,7,8. PEnterobacterales demonstrate resistance to TMP-SMX as well. However, little has been reported on the genetic context of various species of ESBL producing Enterobacterales and its association with, PMQR, AMEs TMP-SMX resistance exclusively from North India. Therefore, the current study was designed to analyze the distribution of non-\u03b2-lactam antibiotic resistance associated genes prevailing in ESBL producing Enterobacterales. To our knowledge, this is the first study that investigated the distribution of AME, PMQR and TMP-SMX resistance genes among ESBL producing Enterobacterales that are isolated from North India. Furthermore, this study included clinical isolates of different members of Enterobacterales , while the earlier studies limited their molecular characterization in two species (E. coli and K. pneumoniae) [Enterobacterales, an observation that has been made only a few times earlier.Literature indicates that synthesis of aminoglycoside modifying enzymes (AMEs) is an important AMR mechanism that produce high level of aminoglycoside resistance among Gram- negative bacteria ,10. Thesumoniae) ,10,13. IK. pneumoniae 16.13% (30/186), E. coli 9.67% (18/186), K. oxytoca 2.15% (4/186), P. mirabilis 1.61% (3/186) and P. vulgaris 0.53% (1/186) were resistant to all the antibiotics tested. Enterobacterales. The highest antibiotic resistance rate was noted for ampicillin 95.89% (70/73) followed by TMP-SMX 90.86% (169/186). The resistance rate of \u03b2-lactams were: cefazolin 94.08% (175/186), amoxicillin-clavulanic acid 77% (124/161), cefotaxime 82.8% (154/186), cefepime 66.67% (124/186), ceftazidime 89.78% (164/186), ceftriaxone 88.17% (164/186), cefoxitin 76.83% (126/164), cefpodoxime 86.11% (155/180), cefuroxime 86.02% (160/186), and ceftizoxime 83.3% (155/186), respectively. The resistance rate of fluoroquinolones were 75.27% (140/186), 69.89% (130/186), 76.88% (143/186), 75.27% (140/186), and 71.51% (133/186) for ciprofloxacin, levofloxacin, nalidixic acid, gatifloxacin, and moxifloxacin, respectively. Similarly, the resistance rate of aminoglycosides were 64% (119/186), 61% (114/186), 49% (91/186), and 59% (109/186) for gentamycin, tobramycin, amikacin, and kanamycin, respectively. The lowest resistance rates were associated with imipenem 43.01% (80/186) and meropenem 46.77% (87/186), respectively  obtained, ectively .Enterobacterales. Among the total phenotypically confirmed ESBL isolates (N = 186), PCR confirmed presence of ESBL genes  in 91.94% (171/186) of Enterobacterales isolates. However, genes coding for ESBL production were not detected from K. pneumoniae 2.69% (5/186), E. coli 2.69% (5/186), K. oxytoca 1.07% (2/186), P. mirabilis 1.07% (2/186), C. freundii 0.54% (1/186) and therefore, these isolates were not included for further analysis. ESBL genes were detected from pus (N = 97), respiratory tract specimens (N = 44), blood and body fluids (N = 30), respectively , E. coli 30.99% (53/171), P. mirabilis 7.6% (13/171), C. freundii 7.02% (12/171), K. oxytoca 5.2% (9/171), E. cloacae 5.2% (9/171), P. vulgaris 1.75% (3/171), and M. morganii 1.75% (3/171), respectively. Of the 91.94% (N = 171) genotypically confirmed isolates, TEMbla 84.79% (145/171) was the most common ESBL gene followed by CTX-Mbla 73.68% (126/171), SHVbla 43.86% (75/171), PERbla 19.88% (34/171) and VEBbla 9.94% (17/171), respectively.ectively . The detEnterobacterales (N = 139) as follows; K. pneumoniae 44.6% (62/139), E. coli 31.65% (44/139), P. mirabilis 8.63% (12/139), C. freundii 5.03% (7/139), K. oxytoca 5.03% (7/139), E. cloacae 2.16% (3/139), P. vulgaris 2.16% (3/139), and M. morganii 0.72% (1/139), respectively (aac(6\u2032)-lb-cr 77.7% (108/139) followed by oqxB 67.63% (94/139), oqxA 62.59% (87/139), qnrB 43.17% (60/139), qnrD 19.42% (27/139), qnrS 18.71% (26/139), qnrA 9.35% (13/139), qepA 3.6% (5/139), and qnrC 2.88% (4/139), respectively , majority of the isolates to had co-existence with TEMbla (13/13) and CTX-Mbla (8/13) gene (qnrB gene (N = 60) demonstrated coexistence with TEMbla (46/60), CTX-Mbla (43/60), and SHVbla (27/60). In contrast, strains carrying Aac-ib-cr gene (N = 108), most of the isolates showed coexistence with TEMbla (98/108), CTX-Mbla (84/108), SHVbla (47/108), PERbla (28/108), and VEBbla (13/108) (oqxA and oqxB as well. Among the isolates that carrying oqxA (N = 87) gene, majority of the isolates were possessing TEMbla (72/87), CTX-Mbla (66/87) and SHVbla (39/87), PERbla (17/87) and VEBbla (12/87), respectively (oqxB genes was carrying TEMbla (80/94), CTX-Mbla (71/94) and SHVbla (46/94), PEblaR (15/94) and VEBbla (11/94), respectively. It was also noted that Enterobacterales isolates that having higher minimum inhibitory concentration (MIC) values for ciprofloxacin (\u226516 \u03bcg/mL), nalidixic acid (\u226564 \u03bcg/mL), levofloxacin (\u226532 \u03bcg/mL),gatifoxacin (\u226532 \u03bcg/mL) and moxifloxacin (\u226532 \u03bcg/mL) mainly harboured Aac-ib-cr, qnrB, oqxA and oqxB genes. Further, the frequency of PMQR genes were high in isolates with higher quinolone MIC than their low MIC counterparts , 81.72% (152/186), 70.43% (131/186), and 90.32% (168/186), were resistant to fluoroquinolones, aminoglycosides and TMP-SMX, respectively. Out of the 91.94% (171/186) of the genotypically confirmed ESBL strains, PCR could detect 81.29% (PMQR), 60.82% (AME), 63.74% (TMP-SMX) of genes in these strains. Of the total PMQR genes (81.29%) detected, the distribution of genes in different members of ectively . PMQR geectively . Of the ectively . Among t13) gene . Similar(13/108) . This coectively . Similarterparts .Enterobacterales (N = 104) is as follows; K. pneumoniae 44.23% (46/104), E. coli 38.46% (40/104), P. mirabilis 2.89% (3/104), C. freundii 5.77% (6/104), K. oxytoca 3.84% (4/104), E. cloacae 2.89% (3/104), P. vulgaris 0.96% (1/104), and M. morganii 0.96% (1/104), respectively (aac(6\u2032)-Ib 81.73% (85/104), followed by aph(3\u2032)-Ia 46.15% (48/104), ant(3\u201d)-Ia 44.23% (46/104), aac(3)-IIa 45.19% (47/104), aph(3\u201d)-Ib 35.58%, (37/104), armA 14.42% (15/104), ant(4\u201d)-IIa 12.5% (13/104), aac(3)-Ib 10.58%(11/104), ant(2\u201d)-Ia 8.65% (9/104), aac(3)-Ia 6.73% (7/104), respectively (aac(2\u2032)-Ia, aac(6\u2032)-Ia, aac(6\u2032)-Ic and aph(3\u201d)-Ia were not detected in any of the strains analyzed (Out of the 60.82% (104/171) of AME genes detected, the distribution of these genes in the ectively . The disectively . Howeveranalyzed .TEMbla (43/47) and CTX-Mbla (38/47) genes. Similarly, strains carrying aac(6\u2032)-Ib gene (N = 85) demonstrated coexistence with TEMbla (75/85), CTX-Mbla (72/85), and SHVbla (43/85). However, strains carrying ant (3\u201d)-Ia gene (N = 46), majority of isolates showed co-existence with TEMbla (41/46), CTX-Mbla (35/46) and SHVbla (23/46). This coexistence of genes were also observed among the isolates that carrying aph(3\u201d)-Ia and aph(3\u201d)-Ib wherein majority of the strains were equally harbouring both TEMbla and CTX-Mbla genes (aac(6\u2032)-Ib, aph(3\u201d)-Ia, aac(3)-IIa, ant(3\u201d)-Ia genes, respectively  that carried aac(3)-IIa genes (N = 47), majority of the isolate had co-existence with -M genes . Furtherectively . The ratectively .Enterobacterales isolates, the distribution of TMP-SMX resistance genes are as follows; K. pneumoniae 42.59% (46/108), E. coli 34.26% (37/108), P. mirabilis 5.56% (6/108), C. freundii 6.5% (7/108), K. oxytoca 5.56% (6/108), E. cloacae 4.63% (5/108), P. vulgaris 1.86% (2/108), and M. morganii 0.92% (1/108), respectively (sul1 100% (108/108), followed by dfrA 54.63% (59/108) and sul2 15.74% (17/108), respectively (Of the 63.74% (108/171) of TMP-SMX resistant ESBL producing ectively . These Tectively .TEMbla (95/108), CTX-Mbla (81/108) and SHVbla (51/108) genes, respectively. Similarly, strains carrying dfrA1 gene (N = 59) demonstrated coexistence with TEMbla (53/59) and CTX-Mbla (48/59)  for TMP-SMX mainly harbored sul1 and dfrA1 genes. It was also observed that the frequency of TMP-SMX genes was higher in strains with high TMP-SMX MIC values , majority of the isolates were having co-existence with  (48/59) . FurtherC values .p < 0.05) different from that found in the respiratory tract specimens  were comparable with earlier studies [TEMbla (84.79%) followed by CTX-Mbla (73.68%), SHVbla (43.86%), PEblaR (18.71%) and SHVbla (9.94%), respectively. These findings were in accordance with an earlier study wherein most prevalent ESBL gene found was TEMbla (73%) followed by CTX-Mbla (25\u2013100%) and SHVbla (23%) [PERbla and VEBbla in the present study were comparable with the data reported by Khurana et al. [The current study investigated the frequency of non-\u03b2-lactam antibiotic resistance associated genes among ESBL producing  studies ,22. The K. pneumoniae (44.6%) was having higher number of PMQR genes detected followed by E. coli (31.65%). This observation is in accordance with an earlier study wherein, PMQR genes were more frequently encountered among E. coli, Klebsiella species, and Enterobacter species [Enterobacter cloacae in this study. This is probably due to the difference in the geographical location as the earlier study was conducted in the southern part of India (where the usage of antibiotics is different). Further, the widespread antibiotic resistance prevalent in India may be attributed to readily availability of antibiotics across the pharmacy counters. This could play a major role in increased distribution of antibiotic-resistance genes throughout the population. This study included isolates that obtained from wound, respiratory tract, body fluid and blood specimens while other studies were performed mostly using isolates that are collected from urine [aac(6\u2032)-lb-cr (77.7%) which is in agreement with an earlier report wherein the prevalence rate was found to be 64.5% [qnrB (43.17%) was observed, which was in accordance with Yang et.al observation  [qnrD, qnrS, qnrA, and qnrC [Enterobacterales isolates were found to be oqxB (67.63%), oqxA (62.59%) and qepA (3.6%), respectively. However, this efflux pump mediated drug resistance mechanism of bacteria can be subdued using various efflux inhibitory molecules [Enterobacterales in North India. However, the oqxA and oqxB prevalence rates were comparable with earlier reports wherein the prevalence rates were found to be 88% and 30% for oqxA and oqxB genes, respectively [oqxA and oqxB genes may be attributed to the geographical distribution and type of isolates studied, as most of the studies were conducted on E. coli and K. pneumonia isolates [qepA observed (3.6%) was similar with previous data (2%) in the literature [qepA in the current study may also indicate low incidence of qepA gene among different strains of Enterobacterales across the world [TEMbla and CTX-Mbla) were more likely to carry aac(6\u2032)-Ib-cr and qnrB genes. The genes that code for both ESBL and PMQR proteins are usually located on same plasmids and consequently that may have higher chances of transfer among the members of Enterobacterales. Therefore, it is very pertinent to comprehend the drug resistance mechanisms prevalent among the members of medically important bacteria as it may be a major concern for patient safety and in determination of therapeutic strategies.In the present study, PCRs detected presence of PMQR genes in 81.29% N = 139) of genotypically confirmed ESBL isolates (N = 171), indicating the presence of high frequency of PMQR genes among ESBL strains. Interestingly, 9 of genoeumoniae 4.6% was om urine . Of the coli 31.6%. This oand qnrC ,23. It wolecules , for insolecules . Furtherolecules . Furtherectively . Furthercoli 31.6%. This oisolates . The lowterature . This locoli 31.6%. This oEnterobacterales. This relatively higher prevalence rate of AMEs in ESBL producing strains may be due to co-existence of genes encoding ESBLs and AMEs in Gram-negative bacteria [K. pneumoniae (44.23%) and E. coli (38.46%). This was in accordance with an earlier study conducted by Haidar et al., wherein a higher prevalence of AME genes were reported among K. pneumoniae [aac(6\u2032)-Ib (81.73%) which is in agreement with the earlier studies  [aac(6\u2032)-Ib with other AME genes observed may be attributed to the fact that the gene coding for aac(6\u2032)-Ib enzyme is frequently located within class I integrons. Further, it is known that the gene cassettes that carry other genes coding for AMEs can be easily incorporated into class I integrons resulting in the development of resistance to currently used aminoglycosides [aph(3\u2032)-Ia (46.15%), followed by ant(3\u201d)-Ia (44.23%), aac(3)-IIa (45.19%), and aph(3\u201d)-Ib (35.58%). The higher prevalence of aph(3\u2032)-Ia and aph(3\u201d)-Ib is alarming as this type of resistance may usually produce high level of aminoglycoside resistance. The prevalence of other AME genes were found to be relatively low, which is comparable with earlier studies [Genes encoding AMEs are prevalent in various groups of bacteria ,30,31,32bacteria . In thiseumoniae . In the was 73%) . The preycosides . The oth-IIa 45.1%, and aponiae 44.% and E. sul1, sul2, or dfrA genes are likely to be present either on chromosome or on plasmids [Enterobacterales. Further, among the genotypically confirmed TMP-SMX resistant strains (N = 108), 42.59% of K. pneumonia isolates were carrying TMP-SMX resistance genes, followed by E. coli (34.26%). However, due to the paucity of literatures, the comparison with earlier studies could not performed. To our knowledge, this new study report, the prevalence of TMP-SMX resistance genes among various clinical strains of ESBL producing Enterobacterales. A higher prevalence of sul1 (100%) followed by dfrA (54.63%) and sul2 (15.74%), genes were noted in this study. This higher prevalence of sul1 gene may be attributed to the fact that the sul1 genes are usually located within class I integrons and this particular characteristic  might have further helped in its wide distribution [dfrA1 (33.91%) gene in the present study was in agreement with an earlier study wherein in the prevalence rate was also found to be high [Enterobacterales isolates that showed elevated MICs for TMP-SMX mainly harbored sul1 and dfrA1 genes indicating the likelihood of these genes in imparting resistance to TMP-SMX. However, no genes were detected from number of isolates that were having higher MIC values, suggesting the existence of alternative pathways of resistance in TMP-SMX resistant isolates.The TMP-SMX resistance genes such as plasmids ,34. In tplasmids . Howeverribution . The com be high . EnterobEnterobacterales. This hospital laboratory receives samples from two civil hospitals and three primary health care centers that are attached to it. Among the total samples analyzed (N = 2134), a total of 186 non-repetitive phenotypically confirmed ESBL producing Enterobacterales isolates (one organism per patient was included to avoid duplication) were obtained. All the isolates were identified by manual API\u00ae system  and the results interpreted as recommended by the manufacturer. The strains included were K. pneumoniae (N = 74), E. coli (N = 58), P. mirabilis (N = 15), C. freundii (N = 13), K. oxytoca (N = 11), E. cloacae (N = 9), P. vulgaris (N = 3) and M. morganii (N = 3). All the clinical isolates were identified by standard laboratory procedure [Between July 2018 and June 2019, a total of 2134 clinical samples  received in the Microbiology laboratory, Government Medical College and Hospital, Badaun, India were analyzed for ESBL producing strains of toca N = , E. cloaK. pneumoniae, C. freundii, K. oxytoca, E. cloacae, P. vulgaris, and M. morganii against Ampicillin (b) C. freundii, E. cloacae, and M. morganii against amoxicillin- clavulanic acid (c) C. freundii, and E. cloacae against cefoxitin (d) P. vulgaris, and M. morganii against cefpodoxime, respectively [E. coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27853).The antibiotic susceptibility testing was conducted by modified Kirby Bauer disc diffusion method as recommended by the Clinical Laboratory Standards Institute (CLSI) . The antectively . QualityThe MIC of fluoroquinolones , aminoglycosides  and TMP-SMX were determined by broth micro dilution method and the results were interpreted in accordance with the CLSI guidelines . The MICE. coli, K. pneumoniae, and K oxytoca). However, zone size with \u226422 mm cefpodoxime (10 \u03bcg), \u226422 mm with ceftazidime (30 \u03bcg), \u226427 mm with cefotaxime, indicated ESBL production for P. mirabilis [ESBL activity was initially screened using disk diffusion method as recommended by CLSI . Isolateirabilis .The double-disk synergy diffusion test (phenotypic confirmatory test for ESBL) was carried out as per CLSI recommendations. Briefly, ceftazidime (30 \u03bcg) and cefotaxime (30 \u03bcg) alone and in combination with clavulanic acid  were used. The ESBL production is confirmed when there is an increase of zone diameter (\u22655 mm) around disk with antibiotic-clavulanic acid combination .E. cloacae, C. freundii, P. vulgaris, and M. morganii strains. This test was performed as disc diffusion assay on Mueller-Hinton agar (MHA). Briefly, antibiotic discs containing aztreonam (30 \u03bcg), ceftazidime (30 \u03bcg), ceftriaxone (30 \u03bcg), and cefotaxime (30 \u03bcg) were kept 30 mm apart (center to center) around amoxicillin-clavulanic acid (20 \u03bcg + 10 \u03bcg) disc on MHA plate inoculated with the organism to be tested. The MHA plates were incubated at 37 \u00b0C for 24 h. An increased zone of inhibition of any of the test antibiotic towards amoxicillin-clavulanic acid was considered as ESBL production [E. coli ATCC 25922 (non-beta-lactamases producer) and K. pneumoniae ATCC 700603 (ESBL producer), respectively.The disc approximation test was used to confirm the ESBL production in Enterobacterales using QIAamp DNA Mini Kit . The DNA samples obtained by this procedure were segregated into two aliquots; first aliquot was used as template for the subsequent PCR reactions and the second aliquot was stored at \u221280 \u00b0C , Ghaziabad, India) for future use.Bacterial DNA was obtained from the phenotypically confirmed ESBL strains of Enterobacterales isolates were subjected to molecular characterization of the relevant encoding genes such as SHV, blaCTX-M, blaTEM, blaPERbla, and VEBbla by PCR using primers and PCR conditions shown in All the phenotypically confirmed ESBL producing Enterobacterales isolates that resistant to fluoroquinolones were subjected to PCRs for detection of (i) qnr proteins coding genes such as qnrA, qnrB, qnrC, qnrD, and qnrS (ii) quinolones modifying aminoglycoside acetyltransferase coding genes (acc(6\u2032)-Ib-cr) and (iii) plasmid-mediated quinolone efflux pump protein encoding genes  using primers and PCR conditions depicted in ESBL producing Enterobacterales were screened for AMEs coding genes aac(2\u2032)-Ia, aac(3)-Ia, aac(3)-Ib, aac(3)-IIa, aac(6\u2032)-Ia, aac(6\u2032)-Ib,aac(6\u2032)-Ic,ant(2\u201d)-Ia, ant(3\u201d)-Ia, ant(4\u201d)-IIa, aph(3\u2032)-Ia, aph(3\u201d)-Ia, aph(3\u201d)-Ib, armA using primers described in ESBL producing aminoglycosides resistant strains of sul1, sul2 and dfrA genes using primers and PCR conditions shown in 4)2SO4, 3 mM MgCl2, 0.2% Tween 20), 200 mM of each dNTPs, 0.5 \u00b5M of each primer, and 2.0 units of AURA Taq DNA polymerase. Amplification was carried out as follows: initial denaturation at 95 \u00b0C for 5 min; 30 cycles of denaturation at 95 \u00b0C for 30 s, annealing at 50\u201362 \u00b0C for 30 s, extension/elongation at 72 \u00b0C for 45 s; and a final elongation step at 72 \u00b0C for 5 min. PCR-generated products were detected by electrophoresis of 7 \u03bcL of each amplification mixture in 2% agarose gels in 1% Tris Borate-EDTA buffer and 0.5 \u00b5g/mL ethidium bromide.All isolates phenotypically resistant to TMP-SMX were subjected to PCR for detection of To identify the ESBL, PMQR, AME, and TMP-SMX resistance genes detected in the PCR assays, automated DNA sequencing of the amplicons were conducted. More specifically, multiplex PCR-generated products were separated in 2% low melting agarose gel with 1% Tris-acetate-EDTA buffer. The PCR products were excised from the agarose gel and purified using the QIAquick PCR purification kit  as recommended by the manufacturer. The nucleotide sequencing of amplicons was conducted using an ABI 3730xl DNA Analyzer . Basic Local Alignment Search Tool (BLAST) program was used to compare each ESBL, PMQR, AME, and TMP-SMX resistance gene sequences against those available in gene bank at the National Center of Biotechnology Information database.p < 0.05 were considered as significant.Chi-squared test was used to compare the association between the origin of strain and type of resistance genes detected. The null hypothesis will be accepted if the presence of genes in all the groups  were similar. Dunn\u2019s multiple comparisons test was performed to compare the differences in the number of resistance genes obtained between two categories of samples. All statistical analyses were performed using Graph pad Prism . The statistical difference values showing Enterobacterales and the association of these genetic determinants with PMQR, AME, and TMP-SMX resistance genes in the north India. The current study demonstrated widespread occurrence of PMQR, AME, and TMP-SMX drug resistant genetic determinants in the ESBL producing Enterobacterales strains. Screening of PMQR genetic elements in ESBL producing Enterobacterales strains revealed high prevalence of both aac(6\u2032)-Ib-cr and qnrB. However, molecular analysis of AMEs producing Enterobacterales strains showed high prevalence of aac(6\u2032)-Ib followed by aph (3\u2032)-Ia. On examination of TMP-SMX resistant strains, sul1 was found to be the most frequently encountered gene followed by dfrA. The association of ESBL-producing genes with the PMQR, AMEs, and TMP-SMX resistance genes may potentially aid in transfer of drug resistance determinants among these strains. Therefore, a complete understanding of PMQRs, AMEs, and other drug resistance mechanisms will help in determining rationale of treatment and infection control measures in hospital settings.In summary, this is the first study that investigated the occurrence of genetic determinants prevailing in ESBL producing"}
+{"text": "Introduction: Intensive oncological treatment integrated with resection of metastases raised the clinical outcome of metastatic colorectal cancer (MCRC). In clinical practice, complex evaluation of clinical , and biological  parameters addresses tailored, personalized multidisciplinary treatment strategies. Patients with MCRC unsuitable for first-line intensive medical treatments are prevalent and showed worse clinical outcome. After progression to oxaliplatin-based chemotherapy, aflibercept/FOLFIRI significantly improved clinical outcome, even if no survival benefit was reported in adjuvant fast relapsers by aflibercept addition. The case reported a young-elderly (yE) patient with KRAS mutant colorectal cancer rapidly progressing to adjuvant chemotherapy, unfit owing to comorbidities, with multiple pharmacogenomic alterations, who gained long-term survival in clinical practice by multidisciplinary treatment strategy consisting of first-line and re-introduction of aflibercept-containing chemotherapy and two-stage lung metastasectomies.Case presentation: A 71-years-old yE patient, unfit for intensive oncological treatments owing to Cumulative Illness Rating Scale (CIRS) stage secondary, affected by KRAS c.35 G>T mutant colorectal cancer, rapidly progressing with lung metastases after adjuvant XelOx chemotherapy, reached long-term survival 66 months with no evidence of disease after first-line and re-introduction of tailored, modulated aflibercept (4 mg/kg) d1,15-irinotecan (120 mg/m2) d1,15-5-fluorouracil (750 mg/m2/day) dd1\u20134, 15\u201318; and secondary radical bilateral two-stage lung metastasectomies. Safety profile was characterized by limiting toxicity syndrome at multiple sites (LTS-ms), requiring 5-fluorouracil discontinuation and aflibercept reduction (2 mg/kg), because of G2 hand-foot syndrome (HFS) for >2 weeks, and G3 hypertension. Pharmacogenomic analyses revealed multiple alterations of fluoropyrimidine and irinotecan metabolism: severe deficiency of fluorouracil degradation rate (FUDR), single nucleotide polymorphisms of UGT1A1*28 variable number of tandem repeats (VNTR) 7R/7R homozygote, ABCB1 c.C3435T, c.C1236T, MTHFR c.C667T homozygote, DPYD c.A166G, TSER 28bp VNTR 2R/3R heterozygote.Conclusions: In clinical practice, a complex management evaluating clinical parameters and RAS/BRAF genotype characterizing an individual patient with MCRC, particularly elderly and/or unfit owing to comorbidities, is required to properly address tailored, multidisciplinary medical and surgical treatment strategies, integrated with careful monitoring of superimposing toxicity syndromes, also related to pharmacogenomic alterations, to gain optimal activity, and long-term efficacy. KRAS/NRAS/BRAF genotype . We previously demonstrated that first-line intensive FIr-B/FOx triplet chemotherapy plus bevacizumab reached an objective response rate (ORR) of 82%, median progression-free survival (PFS) of 12 months, and overall survival (OS) of 28 months . Integragenotype . In non-tastases . In patitastases . Patienttastases , 8.KRAS/NRAS/BRAF genotype) parameters addresses tailored, multidisciplinary treatment strategies  and biological  d1, capecitabine (825 mg/m2 bid) d1\u201314, cycles repeated every 21 days, for six cycles. Safety profile was characterized by LTS-ms, specifically G2 HFS associated with G2 anemia  d1,15-irinotecan (120 mg/m2) d1,15-5-fluorouracil (750 mg/m2/day) dd1\u20134, 15\u201318, cycles repeated every 28 days  of secondary lung metastases with intra-lesion necrosis  mutant MCRC rapidly progressing to adjuvant chemotherapy with bilateral lung metastases, unfit for intensive medical treatment owing to comorbidities, who was treated by first-line and re-introduction of aflibercept-containing chemotherapy followed by two-stage lung metastasectomies and gained in clinical practice by multidisciplinary treatment strategy long-term OS 66 months of metastatic disease with no evidence of disease at PFI 40 months.The case reported a yE patient with KRAS c.35 G>T (G12V) mutation  mutation  and shownd death , with sind death ; the poond death , 19. KRA cancers .KRAS c.35 G>A mutant status was significantly associated with worse clinical outcomes of patients with MCRC treated with FIr-B/FOx compared with KRAS/BRAF wild-type and other KRAS mutant patients  , consistidedness . Adjuvanidedness , 8; in Sidedness , 30 and with PFS , 32. Aflwith PFS . Afliberwith PFS . Afliberwith PFS . At progS mutant . TripletS mutant .Then, because of long-term control of bilateral lung metastastes during first-line aflibercept/irinotecan and re-challenge, bilateral resections of lung metastases were performed. The diagnosis of mucinous lung metastases may justify such a long OS 66 months of metastatic disease without evidence of disease at PFS 50 months and PFI 40 months from second stage lung metastasectomies, thus realizing the effectiveness of the integrated lung metastasectomies. In a retrospective Spanish real-life analysis of 32 patients who underwent surgical resection after aflibercept/FOLFIRI , PR was Reported yE, unfit patient underwent first-line MCRC treatment with reduced doses of aflibercept, irinotecan, and fluorouracil. Nevertheless, safety profile was characterized by LTS-ms, particularly G2 HFS lasting >2 weeks with different other G2\u2013G1 toxicities. Because of limiting HFS and multiple pharmacogenomic alterations, 5-fluorouracil was discontinued; then, LTS-ms was observed, characterized by G3 hypertension, and re-introduction was planned with further aflibercept dose reduction. Reported prevalent toxicities with FOLFIRI/aflibercept were diarrhea (19.3%), mucositis 13.7%), asthenia (16.9%), HFS 2.8%), hypertension (2.9%), arterial (1.8%) and venous (7.9%) thromboembolic events, neutropenia (36.7%), and thrombocytopenia (3.3%) %, hypert. In Span.7%, asthUGT1A1*28 homozygote 7 repeats, ABCB1 c.C3435T and C1236T heterozygote, MTHFR\u2014c.C667T homozygote, DPYD c.A166G heterozygote, and TSER 28bp heterozygote VNTR 2R/3R. Dihydropyrimidine dehydrogenase gene (DPYD) and UGT1A1 SNPs variably influence fluoropyrimidines and irinotecan tolerability (DPYD c.496A > G (P = 0.022) and c.1896 T > C (P = 0.027), trendly with UGT1A1*28 SNPs (P = 0.054) , ABCB1 (71%), CYP3A4 (14%), and DYPD (15%), in the reported range , thus predicting occurrence of LTS-ms in patients at risk of gastrointestinal LT  . UGT1A1*tropenia \u201349. 5-FUtropenia , 51. To ed range , 49. MosRAS/BRAF genotype of patients with MCRC, particularly the elderly and/or those who are unfit because of comorbidities, is required to properly tailor multidisciplinary medical and surgical treatment strategies to gain optimal activity and long-term efficacy, integrated with careful monitoring of toxicity syndromes, potentially related to pharmacogenomic alterations.In clinical practice, a complex management evaluating patient-related clinical parameters and Written informed consent was obtained from the patient for the publication of any potentially identifiable images or data included in this article.GB and ER were the medical oncologists responsible for clinical and bioclinical management of patient. AD'A, SG, and EAR were abdominal and thoracic surgeons responsible for surgical management of patient. MS performed pharmacogenomic analyses. All authors read and approved the final manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer AB declared a past co-authorship with one of the authors MS to the handling editor."}
+{"text": "Chrysomallon squamiferum) is highly adapted to deep-sea hydrothermal vents and has drawn much interest since its discovery. However, the limited information on its genome has impeded further related research and understanding of its adaptation to deep-sea hydrothermal vents.The scaly-foot snail (Gigantopelta aegis), which inhabits similar environments. Using Oxford Nanopore Technology, 10X Genomics, and Hi-C technologies, we obtained a chromosome-level genome of C. squamiferum with an N50 size of 20.71 Mb. By constructing a phylogenetic tree, we found that these 2 deep-sea snails evolved independently of other snails. Their divergence from each other occurred \u223c66.3 million years ago. Comparative genomic analysis showed that different snails have diverse genome sizes and repeat contents. Deep-sea snails have more DNA transposons and long terminal repeats but fewer long interspersed nuclear elements than other snails. Gene family analysis revealed that deep-sea snails experienced stronger selective pressures than freshwater snails, and gene families related to the nervous system, immune system, metabolism, DNA stability, antioxidation, and biomineralization were significantly expanded in scaly-foot snails. We also found 251 H-2 Class II histocompatibility antigen, A-U \u03b1 chain-like  genes, which exist uniquely in the Gigantopelta aegis genome. This finding is important for investigating the evolution of major histocompatibility complex (MHC) genes.Here, we report the whole-genome sequencing and assembly of the scaly-foot snail and another snail (Our study provides new insights into deep-sea snail genomes and valuable resources for further studies. Bathymodiolus platifrons) showed that, while most of the genes present in a related shallow-water mussel (Modiolus philippinarum) have been retained, many gene families have expanded in the\u00a0B. platifrons genome. These families include those that are associated with stabilizing protein structures, removing toxic substances from cells, and the immune response to symbionts , Octopus bimaculoides [GCF_0\u00a0011\u00a094135.1], Biomphalaria glabrata [GCF_000457365.1], Crassostrea gigas [GCF_000297895.1], Lottia gigantea [GCF_000\u00a0327385.1], Pomacea canaliculata [GCF_003073045.1], Pinctada fucata [GCA_0\u00a0022\u00a016045.1], and Helobdella robusta [GCF_000\u00a0326865.1] from the NCBI database and C. squamiferum and G. aegis from this research) were compared using blastp with the E-value threshold set as 1e\u22127. Then, alignment segments of each protein pair were concatenated using the in-house software Solar v0.9.6 [The TreeFam tool   was usedr v0.9.6 . H-scorer v0.9.6 .RRID:SCR_011812) [A. californica\u2014C. gigas (\u223c516.3\u2013558.3 MYA), A. californica\u2014P. canaliculata (\u223c310\u2013496 MYA), A. californica\u2014O. bimaculoides (\u223c551\u2013628 MYA), C. gigas\u2014H. robusta (\u223c585\u2013790 MYA), and C. gigas\u2014P. fucata (394\u2009 MYA) [We obtained 406 one-to-one single-copy orthology gene families based on gene family classification. Then, these gene families were extracted and aligned using guidance from amino acid alignments created using the default parameters of the MUSCLE   programm_011812) , 74 was 94\u2009 MYA)  were useC. squamiferum and G. aegis using BWA-MEM (v0.7.12-r1039)  [RRID:SCR_002105) [RRID:SCR_001876) [Approximately 50\u00d7 clean WGS reads were mapped to genomes of _010910)  with def_002105)  and \u201cSor_001876)  with def_001876) .We used CAFE v2.1  to analyCNP0000854. The raw sequencing reads were also uploaded to the SRA database under accession No. CNP0000854. All supporting data are available in the GigaScience GigaDB database [The genome assemblies of these 2 genomes have been deposited in GenBank under accession No. database .Supplementary Figure S1.17-mer frequency distribution for C. squamiferum and G. aegis genomes.Supplementary Figure S2.Construction of Phylogenetic trees for ten representative molluscs using coding sequences of 407 single-copy orthologs.Supplementary Figure S3.Box plot of Ka and Ks values of 1,324 single copy orthologous genes from two deep-sea snails, one shallow-water snail, and two fresh-water snails.Supplementary Figure S4.Expansion pattern of HTR4 genes in two deep-sea snails.Supplementary Figure S5.KEGG enrichment analysis of unique gene families of G. aegis.Supplementary Table S1.Statistics of raw sequencing data of Chrysomallon squamiferum.Supplementary Table S2.Statistics of raw sequencing data of Gigantopelta aegis.Supplementary Table S3.Summary from the genome assembly of Chrysomallon squamiferum without using Hi-C data.Supplementary Table S4.Lengths of the 16 chromosomes assembled for Chrysomallon squamiferum.Supplementary Table S5.BUSCO assessment of the assembled genome of Chrysomallon squamiferum using metazoa_odb9 database.Supplementary Table S6.Summary of the genome assembly for Gigantopelta aegis.Supplementary Table S7.BUSCO assessment of the assembled genome for Gigantopelta aegis using the metazoa_odb9 database.Supplementary Table S8.General statistics of predicted protein-coding genes of Chrysomallon squamiferum.Supplementary Table S9.General Statistics of Predicted Protein-coding Genes of Gigantopelta aegis.Supplementary Table S10.Summary of predicted gene functions of the Chrysomallon squamiferum gene set.Supplementary Table S11.Summary of predicted gene functions of the Gigantopelta aegis gene set.Supplementary Table S12.Summary of repeat contents in four selected species.Supplementary Table S13.Gene family clusters in selected species.Supplementary Table S14.Estimation of mutation rates of two deep-sea snails.Supplementary Table S15.Ka and Ks values of 1,324 single copy orthologous genes from five snails.Supplementary Table S16.KEGG enrichment of expanded gene families of C. squamiferum.Supplementary Table S17.KEGG enrichment of unique gene families in G. aegis genome.bp: base pairs; BTBD6: BTB/POZ domain-containing protein 6; BUSCO: Benchmarking Universal Single-Copy Orthologs; BWA: Burrows-Wheeler Aligner; CAFE: Computational Analysis of gene Family Evolution; CAT: catalase; CR1: chicken repeat 1; DLD: dihydrolipoyl dehydrogenase; DLST: dihydrolipoyl succinyltransferase; DMBT1: deleted in malignant brain tumours 1; DNB: DNA nanoball; GAFT: glutamine-fructose-6-phosphate transaminase; GATK: Genome Analysis Toolkit; Gb: gigabase pairs; GP340: glycoprotein-340; Hi-C: high-throughput chromosome conformation capture; HIF-1\u03b1: hypoxia-inducible factor-1\u03b1; HIV: human immunodeficiency virus; Hsp90: heat shock protein 90; HTR4: 5-hydroxytryptamine receptor 4; IDH: isocitrate dehydrogenase; IUCN: International Union for Conservation of Nature; kb: kilobase pairs; KEGG: Kyoto Encyclopedia of Genes and Genomes; LINE: long interspersed nuclear element; LTR: long terminal repeat; Mb: megabase pairs; MHC: major histocompatibility complex; ML: maximum likelihood; MYA: million years ago; NCBI: National Center for Biotechnology Information; OGDC: \u03b1-ketoglutarate dehydrogenase complex; OGDH: oxoglutarate dehydrogenase; ONT: Oxford Nanopore Technologies; ORF: open reading frame; PAML: Phylogenetic Analysis by Maximum Likelihood; PSMC: pairwise sequential Markovian coalescent; SAG: salivary agglutinin; SNP: single-nucleotide polymorphism; SRA: Sequence Read Archive; SRCR: scavenger receptor cysteine-rich; TCA: tricarboxylic acid; TE: transposable element; TLR13: Toll-like receptor 13; TNF: tumour necrosis factor; Txn1: Thioredoxin 1; WGS: whole-genome sequencing.The authors declare that they have no competing interests.This work was supported by the National Key R&D Programme of China (No. 2018YFC0310702).Zongze Shao, S.L., G.F., and X.L. conceived and managed this project and amended the manuscript. X.Z., Y.Z., L.M., and I.S. performed the evolutionary analysis and wrote the manuscript. L.M., J.C., and Y.S. performed genome assembly and annotation. J.B., S.L., X.F., C.W., Zenghua Shao, H.L., N.L., and L.W. were responsible for sample collection, DNA extraction, and library construction.giaa139_GIGA-D-20-00187_Original_SubmissionClick here for additional data file.giaa139_GIGA-D-20-00187_Revision_1Click here for additional data file.giaa139_Response_to_Reviewer_Comments_Original_SubmissionClick here for additional data file.giaa139_Reviewer_1_Report_Original_SubmissionTakeshi Takeuchi, Ph.D -- 7/31/2020 ReviewedClick here for additional data file.giaa139_Reviewer_1_Report_Revision_1Takeshi Takeuchi, Ph.D -- 10/9/2020 ReviewedClick here for additional data file.giaa139_Reviewer_2_Report_Original_SubmissionMathew J. Jenny -- 8/5/2020 ReviewedClick here for additional data file.giaa139_Reviewer_3_Report_Original_SubmissionReuben William Nowell, Ph.D. -- 8/10/2020 ReviewedClick here for additional data file.giaa139_Supplemental_Tables_and_FiguresClick here for additional data file."}
+{"text": "MAPK mutations favor HNSCC survival, revealing the broad clinical utility of MAPK pathway mutations in prognosis and potentially in precision immunotherapy. TP53 mutations (median \u223c14 yr). We explored underlying outcome-favoring mechanisms with omics followed by preclinical models. Strikingly, multiple hotspot and non-hotspot MAPK mutations  all abrogated ErbB3 activation, a well-established HNSCC progression signal. Inhibitor studies functionally defined ERK activity negatively regulating phospho-ErbB3 in MAPK-mutants. Furthermore, pan-pathway immunoprofiling investigations identified MAPK-mutant tumors as the only \u201cCD8+ T-cell\u2013inflamed\u201d tumors inherently bearing high-immunoreactive, constitutive cytolytic tumor microenvironments. Immunocompetent MAPK-mutant HNSCC models displayed active cell death and massive CD8+ T-cell recruitment in situ. Consistent with CD8+ T-inflamed phenotypes, MAPK-mutant HNSCC patients, independent of tumor-mutational burden, survived 3.3\u20134 times longer than WT patients with anti-PD1/PD-L1 immunotherapies. Similar prognosticity was noted in pan-cancers. We uncovered clinical, signaling, and immunological uniqueness of MAPK-mutant HNSCC with potential biomarker utilities predicting favorable patient survival.MAPK pathway mutations affect one-fifth of head and neck squamous cell carcinoma (HNSCC). Unexpectedly, MAPK pathway aberrations are associated with remarkably long patient survival, even among patients with  The incidence of head and neck squamous cell carcinoma (HNSCC) is approaching 0.7 million/year globally . DespiteTP53 wild-type (WT) status which allows proper execution of drug-induced apoptosis by functional p53  responses in HNSCC, with proven biology demonstrated in PI3K-mutant, PI3K-activated preclinical models of HNSCC and retrospective patient cohorts  positivity in patients\u2019 tumors can be potentially used as a favorable prognostic biomarker (mainly for cancer from the oropharynx subsite), which can clinically stratify patients for treatment de-intensification purposes . The bioonal p53 . Similar cohorts . These sTP53 mutations (median \u223c14 yr), much longer than that of HPV-positive HNSCC (median \u223c5.5 yr). The favorable prognosticity of MAPK pathway mutations in HNSCC was found to be independent of HPV. Subsequent molecular dissections revealed two plausible underlying mechanisms operative by MAPK mutations in patient tumors, followed by preclinical HNSCC models. First, multiple hotspot and non-hotspot MAPK mutations  all abrogated ErbB3 activation, an established signal for HNSCC progression. Inhibitor studies functionally defined ERK activity negatively regulating p-ErbB3 in MAPK-mutants. Second, comprehensive pan-pathway immune landscape investigations identified MAPK-mutant tumors as the only \u201cCD8+ T-cell\u2013inflamed\u201d tumors with inherently immunoreactive tumor microenvironments with constitutive cytolysis, subsequently validated in immunocompetent HNSCC models. As low tumoral phospho-ErbB3 levels and elevated CD8+ T-cell infiltrations are recently established events driving good patient survivals in HNSCC , were able to identify subsets of HNSCC patients benefiting significantly from anti-PD1/PD-L1 immunotherapy clinically. Similar prognosticity of MAPK pathway mutations were noted in pan-cancer immunotherapy settings. Our study uncovers novel clinical, biological, and immunological uniqueness of MAPK-mutant HNSCC and may indicate important clinical utilities of MAPK pathway mutations in this cancer.Here, we first reported that MAPK pathway mutations in HNSCC predict remarkably long patient survival, even among patients bearing in HNSCC , our stuAs of today, the clinical significance of somatic genomic aberrations in HNSCC remains underexplored. Here, using a comprehensive pan-pathway approach, we examined the prognostic impact of mutational events of seven major cancer-related signaling pathways on HNSCC patient survivals with The Cancer Genome Atlas (TCGA) dataset . These include three major mitogenic pathways, the PI3K, MAPK(ERK), and JAK/STAT pathways; two differentiation-related pathways, the Notch and TGF-\u03b2/Smad pathways; the inflammation-related NF-\u03baB pathway; and the stem cell\u2013related WNT pathway, representing a total of 81% of cases . PathwayNOTCH1 mutations predict recurrences with poor outcomes .Among all seven pathways examined, only mutations in the MAPK pathway  and the Notch pathway  are associated with patient outcomes . Mutatiooutcomes . Unexpec p.E322K ; Fig S2)TP53-mutated patients  (TP53 double mutant\u201d patients (N = 363) have an extreme long median OS of 169.25 mo (\u223c14 yr), which is 4.77 times longer than that of the \u201cMAPK-WT/TP53-mutated\u201d counterparts . The double mutant patients also have a 55.26% reduction in chances of death (OR = 0.4474) versus MAPK-WT counterparts .Most strikingly, MAPK pathway mutations are also prognostic for markedly improved survival even among iveness) . In factP = 0.01003, 0.03372, respectively, Table S1). Importantly, unlike HPV-positive HNSCC with favorable outcomes, MAPK pathway mutations span multiple head and neck anatomic subsites, including the oral cavity sites, larynx, oropharynx, and others (Table S2). More than 87% (83/95 cases) of MAPK pathway-mutated tumors are HPV-negative. Upon HPV stratification, MAPK pathway mutations are still found to be prognostic for HPV-negative HNSCC , but potentially associated with lower alcohol intake per TCGA alcohol history, and a higher occurrence in females .Table S1 Subsite distribution of MAPK pathway mutations in HNSCC tumors (upper panel) and subsite distributions of HPV positivity in TCGA HNSCC cohort (lower panel).Table S2 Pan-cancer analyses results for associations between MAPK proteome components  in patient tumors and overall survival in 33 cancer types (TCPA database).Table S3 H/N/K-RAS, A/B-RAF, RAF-1, MAP2K1/2, and MAPK1) were also prognostic of favorable HNSCC patient outcome  expression versus control  expressions in HNSCC cell models  and the hotspot ARAF p.S214F mutations inhibited p-ErbB3(Y1289) expressions. In addition to these mutants, overexpression of either MAP2K1-WT or BRAF-WT was also sufficient to inhibit p-ErbB3(Y1289) expressions by \u223c60% and 50%, respectively.ErbB3 activation contributes to HNSCC proliferation  and is ag/tcpa/, , 2017a) in HNSCC , we thusl models  and S4A.Top 19 proteins significantly associated with patient OS in the HNSCC TCPA dataset.Table S4 MAPK1 p.R135K and HRAS p.G12S mutations and the HSC-6 cell line harboring MAPK1 p.E322K, >100\u2013200% increases of p-ErbB3 expression were observed, whereas such an increase was not observed in MAPK-WT cells (HSC-4). The data suggested MAPK (ERK) activity negatively regulates p-ErbB3 in a mutant-specific manner in HNSCC. Such a mutant-specific phenomenon was further supported by proteomic findings in MAPK-mutant HNSCC cell lines  and 0.4333  levels. Therefore, we further tested this non-canonical MAPK/ErbB3 relationship using the pharmacological GDC-0994 MAPK1/2-RSK inhibitor in HNSCC cells harboring endogenous MAPK pathway mutations. As shown in a [CCLE] ) and in Response to therapy is an important factor determining cancer patient survival. Yet, there was no clear association between MAPK mutations and responses to cisplatin, docetaxel, 5-flurouracil, methotrexate, and cetuximab in CCLE HNSCC drug-sensitivity database . SimilarSupplemental Data 1, there were 1,793 significant differentially expressed genes (DEGs) in MAPK-mutated versus WT HNSCC tumors with P-values < 0.05 and false discovery rate (FDR) < 0.05, among which 130 protein-coding DEGs showed a >1.41-fold difference (log2 fold-change > 0.5) in gene expression with FDR < 0.05 (Perforin (PRF1), and four granzymes, GZMA (Cytotoxic T-Lymphocyte-Associated Serine Esterase-3), GZMB (Cytotoxic T-Lymphocyte-Associated Serine Esterase-1), GZMH (Cytotoxic Serine Protease C), and GZMK (Tryptase II) were all significantly up-regulated in MAPK-mutated tumors  differentially expressed protein-coding genes (DEGs) in MAPK-mutated tumors (versus WT) with all immune-related genes highlighted in red.A list of 130 significantly up-regulated .A complete list of 1,793 significant differentially expressed genes (DEGs) in MAPK-mutated tumors versus WT tumors in TCGA HNSCC provisional cohort . Most importantly, MAPK-mutant tumors have the most remarkable \u201cCD8+ T-cell\u2013inflamed status\u201d along with most significant and concurrent increases in all three major immunoreactive signature scores, including high cytolytic process (CYT)  and neutrophil infiltration levels in MAPK-mutant tumors  and neutrophils, which were discrete from signatures for B cell, CD4+ T-cell, and macrophage infiltrations  analysis , 2017 ress (CYT) , T-effecss (CYT) , and antss (CYT) , all eviss (CYT) . Whereas in situ . We alsotrations . Overall as well  and S8B.+ T-cells (accompanied by dendritic cells and neutrophil infiltrations) by immunohistochemistry, consistent with the TIMER-predicted CD8+ T-cell\u2013inflamed, immunoactive microenvironments borne by MAPK-mutated HNSCC tumors in TCGA , whereas such infiltrates were not detected in WT tumors .Increases of CD8+ T-cell infiltration in HNSCC tumor microenvironment alone are known to independently predict favorable patient survival  Hartman. Our in- in TCGA  and S10.+ T-cell infiltration in HNSCC dictate patient outcomes , and their abilities to induce intratumoral CD8+ T-cell infiltration in vivo were compared. As shown in HRAS p.G12V (mHRAS p.G12V), mMAPK1 p.D319N , and mMAPK1 p.E320K mutations , were all potent recruiters for CD8+ T-cell infiltration in situ. Furthermore, consistent with our findings that MAPK-mutated human HNSCC tumors have inherently high active cytolytic signatures  were all survival-favoring features among HNSCC patients . Importantly, MAPK pathway-mutated HNSCC patients bearing high IFN-\u03b3 scores, high CYT scores, and high T-eff scores have long-term survivals . At the most relaxed median cutoff, both the IFN-\u03b3 and T-eff scores could still separate patient survival (P < 0.05), but not CYT score (P = 0.2) (In addition to high CD8P = 0.2) \u2013S13.Notably, among MAPK-mutated HNSCC patients, we found that those with high IFN-\u03b3, CYT, or T-eff scores (both at 20% and 50% cutoffs) do not significantly overlap with patients of low p-ErbB3  and oropharyngeal cancer (N = 32) were two major cancers with sufficient case numbers for survival analysis. Strikingly, among advanced oral cancer patients treated with PD1/PD-L1 inhibitors, MAPK pathway mutations appeared to predict a 3.3 times longer median OS versus WT   . This wact test) .Notably, as high as 49% of HNSCC tumors (53/110) sequenced by + T-cell\u2013inflamed/cytolytically active HNSCC patients who may potentially benefit from ICIs. Our study uncovers novel clinical, biological, and immunological uniqueness of MAPK-mutant HNSCC and may indicate wide clinical utilities of MAPK pathway mutations in this cancer.Overall, MAPK pathway mutations may represent novel and important HNSCC biomarkers for prognosis in HNSCC. Based on our transcriptome findings, validated in immunocompetent models, MAPK mutations likely identify highly CD8TP53-mutated HNSCC patients (median OS >14 yr). Thus, MAPK mutations may represent novel prognostic or even de-intensification biomarkers for HNSCC with broad applicability in terms of HNSCC subsites.We reported for the first time that MAPK pathway mutations have significant survival-favoring clinical, signaling, and immunological impacts in HNSCC. Among seven major cancer-signaling pathways examined, MAPK (ERK) pathway mutations, predominated by many activating mutations, were prognostic for remarkably long patient survivals. This is contradictory to the known role of MAPK-mitogenic signaling in HNSCC tumorigenesis and progression . This fiHRAS and MAPK1 hotspot mutations and potentially others, have remarkable tumoral ErbB3-suppression . Mechanistically, our findings identified a previously undescribed mechanism of p-ErbB3 regulation by MAPK pathway-mutants. Such a negative regulation of p-ErbB3 by ERK activity is uniquely found in MAPK-mutant, but not in MAPK-WT HNSCC. Most importantly, MAPK-mutant HNSCC tumors are the only tumors having significant \u201cCD8+ T-cell\u2013inflamed\u201d and inherently immunoactive tumor microenvironment (versus six other pathway-mutant tumors), with constitutive cytolysis. The ability of MAPK mutations to drive a CD8+ T-cell\u2013inflamed status in vivo with marked apoptosis is proven in immunocompetent HNSCC models. As low tumoral phospho-ErbB3 levels and elevated CD8+ T-cell infiltrations are recently established events indicative of good patient survivals in HNSCC , MAPK1/3(ERK2/1), RPS6KA1, SHC1/2/3/4, GRB2, and Erk1/2-specific DUSP3/5/6/7/9. The PI3K pathway was defined as AKT1/2/3, PIK3CA/B/D/G/2A/2B/2G, PIK3AP1, PIK3IP1, PDK1, MTOR, TSC1, TSC2, PTEN, RICTOR, RPTOR, RHEB, and PIK3R1/2/3/4/5/6. The NF-\u03baB pathway was defined as TAB1/2/3, MAP3K7/14, CHUK, IKBKB, IKBKG, NFKBIA, NFKBIE, REL, RELA/B, NFKB1/2, LTBR, TNF, TNFAIP3, TNFSF11/13B, TNFRSF1A/8/11A/13C, BTRC, CYLD, NLRC5, TRADD, CD40, CD40LG, LTA, TRAF2/3/5/6, IL1B, and IL1R1. The JAK/STAT pathway was defined as JAK1/2/3, STAT1/2/3/4/5A/5B/6, PTPN11, IL6, IL6R, IL6ST, and SOCS3. The Notch pathway was defined as DLL1/3/4, JAG1/2, NOTCH1/2/3/4, NUMB, DTX1/3L, NEDD4, MAML1, RBPJ, POFUT1, HES1/5, and HEY1/2/L. The WNT pathway was defined as WNT1/3A/5A/5B/7A, CTNNB1, HNF1A, FZD1/2/3/7/8/9/10, AXIN1, LEF1, LOXL2, DVL2/3, NKD1/2, TAB1/2, GSK3B, CSNK1A1, NLK, and LRP5/6. The TGF-\u03b2/Smad pathway was defined as SMAD1/2/3/4/5/6/7/9, TGFB1/2/3, TGFBR1/2, INHBA/B/C/E, NODAL, ACVR1/1B/1C/2A/2B, BMP2/4/7, BMPR1A/B/2, AMHR2, LTBP1, BAMBI, ZFYVE9, SMURF1/2, and LIMK1. All TCGA whole-exome sequencing data and clinical data of TCGA are downloaded from the www.cbioportal.org  generated from Kaplan\u2013Meier curves as in P-value < 0.05 (significant) would we consider them having favorable/unfavorable OS. CCLE-proteomic database was downloaded from DepMap portal (http://www.depmap.org) as published by the CCLE study . The Platinum-A (PLAT-A) retrovirus packaging cell line was purchased from Cell Biolabs. The HSC-6 cell line was a generous gift from Dr J Inazawa , and SCC VII mouse HNSCC cell line was kindly provided by Dr Sven Branduau . Pt-25 primary cultures were prepared from a female recurrent HNSCC patient. For GDC-0994 treatment, the cells were plated at \u223c30% confluency for overnight and then subjected to either vehicle or 0.5 \u03bcM of GDC-0994 for 30 min. The cells were then washed with 1\u00d7 PBS, and protein lysates were prepared for Western blot analyses.2. Retroviruses were removed from the infection medium at postinfection, and cells were then cultured in their respective culture media. Gene expression were then validated by Western blotting. For mouse HNSCC tumor cell inoculation  was used for ectopic expression of genes and mutants into HNSCC cells. In brief, the desired genes/mutants cloned into the pMXs-puro retroviral expression vector backbone were transfected into PLAT-A cells using Lipofectamine 3000 (Thermo Fisher Scientific) for the generation of retroviruses. Retroviruses were filtered through a 0.45-\u03bcm mixed cellulose ester membrane filter to remove cell debris and subsequently used for infection of FaDu cells or SCC VII cells for 48\u201372 h at 37\u00b0C, 5% COckground ), 2\u20133 miCells were washed with cold 1\u00d7 PBS and lysed with the NP40 lysis buffer . Cell lysates were centrifuged, and the supernatant was quantified with Protein Assay Dye Reagent (Bio-Rad). 50 \u00b5g of protein lysates were mixed with a 4\u00d7 protein loading dye and then separated by SDS\u2013PAGE. Separated proteins were transferred to nitrocellulose membrane, which was then blocked with 5% nonfat dry milk  and probed with primary antibody at 4\u00b0C overnight. Primary antibodies include AKT (#9272), pi-AKT (#9271), ARAF(#4432), pi-ARAF(S299) (#4431), BRAF (#2696), pi-BRAF(S445) (#2696), pi-ErbB3 (#2842), ErbB3 (#12708), MAPK (#9102), pi-MAPK (#9101), pi-MEK1/2 (#9154), and RSK1 (#8408), all from Cell Signaling Technology, USA. Anti-\u03b2-actin (sc-69879) antibody was from Santa Cruz. Anti-MEK1/2 (YT2714) and anti-N/H/K-RAS (YT2960) antibodies were from ImmunoWay. The probed membrane was then washed three times with 1\u00d7 TBST, followed by a 1\u20132-h incubation with the respective secondary antibody , followed by 3\u00d7 washings with 1\u00d7 TBST. ECL detection solution was then applied onto the membrane for the development of chemiluminescence, which was captured by autoradiography.+ T-cell, CD4+ T-cell, macrophage, neutrophil, and dendritic cell) in 512 TCGA-HNSCC head and neck squamous carcinoma tumor samples are extracted from the TIMER Web site (https://cistrome.shinyapps.io/timer/_w_20aca96c/immuneEstimation.txt) and grouped according to the mutational information downloaded from cBioPortal . The Wilcoxon rank sum test was performed to calculate the statistical significance between two groups in the TIMER plots.We adopted the methodology of TIMER analysis for immune infiltration level estimation , 2017b. https://portal.gdc.cancer.gov/repository?facetTab=files&filters=%7B%22op%22%3A%22and%22%2C%22content%22%3A%5B%7B%22op%22%3A%22in%22%2C%22content%22%3A%7B%22field%22%3A%22cases.project.project_id%22%2C%22value%22%3A%5B%22TCGA-HNSC%22%5D%7D%7D%2C%7B%22op%22%3A%22in%22%2C%22content%22%3A%7B%22field%22%3A%22files.data_category%22%2C%22value%22%3A%5B%22Transcriptome%20Profiling%22%5D%7D%7D%5D%7D&searchTableTab=files)   using GDGZMA and PRF1 as previous described by GZMA, GZMB, PRF1, IFN-\u03b3, EOMES, and CD8A (offset 0.01)  was calculated by the geometric mean of the TPM of et 0.01) . AntitumMAP2K1 and MAPK1. It also captured selection regions of ARAF, BRAF, HRAS, KRAS, and MAP2K2 to cover all well-known hotspot sites. The sequenced locations for these partially sequenced genes are listed in the following table, and variants were called by the Ion Reporter Software (Thermo Fisher Scientific).Tumor tissues and blood samples were collected from patients under written informed consents according to clinical research approvals by the Institutional Review Board of the University of Hong Kong/Hospital Authority Hong Kong East Cluster Research Ethics Committee , the Joint Chinese University of Hong Kong\u2013New Territories East Cluster Clinical Research Ethics Committee , Hong Kong SAR, and the Research Ethics Committee, Kowloon West Cluster , Hong Kong SAR. Genomic DNA from samples were extracted with the DNeasy Blood & Tissue Kit (QIAGEN), followed by quantification and targeted sequencing by next-generation sequencing using the IonS5 platform (Thermo Fisher Scientific). All samples were sequenced with a mean depth of >500\u00d7 using a custom-designed gene panel on major MAPK pathway genes. The custom gene panel consisted of amplicons that covered all exons of Patient tumors were freshly fixed with 10% formalin and dehydrated in a serial manner in ethanol. The formalin fixed and paraffin embedded tumor samples were then sectioned and dewaxed and rehydrated in xylene, 100% ethanol, 70% ethanol, and running tap water. Antigen retrieval was performed at 95\u00b0C for 20 min in citrate buffer . The VECTASTAIN Elite ABC Universal PLUS Kit Peroxidase (Horse Anti-Mouse/Rabbit IgG) (Cat. no. PK-8200) was used for immunohistochemical staining. Endogenous peroxidase activity was quenched by BLOXALL Blocking Solution, followed by blocking in 2.5% Normal Horse Serum for 20 min at room temperature. CD8 mouse antihuman antibody , CD11c rabbit antihuman antibody , and neutrophil elastase rabbit antihuman antibody  and phospho-HER3/ErbB3(Tyr1289) (21D3) rabbit antihuman antibody  were used as primary antibodies to stain patient tumors for overnight incubation at 4\u00b0C. CD8 rabbit antimouse antibody  and cytokeratin mouse antibody  were used. The secondary antibody (prediluted biotinylated horse antimouse/rabbit IgG secondary antibody) was added for 1-h incubation at room temperature. For signal amplification and detection, the slides were incubated with VECTASTAIN Elite ABC reagent for 30 min and ImmPACT DAB EqV solutions for 30 s. After counterstaining with hematoxylin for 1 min, clearing, and mounting, pictures were taken under a light microscope. The Roche In Situ Cell Death Detection Kit, Fluorescein (Cat. no. 11684795910) was applied for TUNEL assay. Proteinase K solution  was used to pretreat the rehydrated sections. Then the TUNEL reaction mixture was incubated on the section in dark at 37\u00b0C for 1 h in dark. The section was mounted with VECTASHIELD vibrance antifade mounting medium (Cat. no. H-1800)."}
+{"text": "The structures were determined based on spectroscopic evidence including 1D and 2D nuclear magnetic resonance (NMR) spectroscopy and high resolution mass spectrometry (HRESIMS) data, along with comparisons to reported data. The leaves were used to extract compounds with different solvents. The extracts were tested for antioxidant activity with a variety of in vitro tests including 2,2-diphenyl-1-picrylhydrazyl (DPPH\u2022), 2,2\u2032-azino-bis (3-ethylbenzothiazoline-6-sulfonate (ABTS\u2022+), ferric reducing antioxidant power assay (FRAP), superoxide, and nitric oxide radical scavenging assays. The dichloromethane fraction was most active, displaying significant free radical scavenging activity. The n-butanol fraction also showed notable activity in all assays. Therefore, these findings support the potential use of A. vacillans leaves as an antioxidant medication due to the presence of polyphenolic compounds.A new dihydroisocoumarin glucoside, vacillanoside ( Aloe spp. are members of the bitter or Asphodelaceae family (previously known as Liliaceae). This family is represented by more than 600 species endemic to tropical and southern Africa, Madagascar, Jordan, the Arabian Peninsula, East Asian countries, and various islands in the Indian Ocean + , m/z 529.1320 [M+Na]+ , and m/z 545.1069 [M+K]+  with 162 amu more than that of 2, suggesting the presence of a monosaccharide moiety. A significant fragment using the high-resolution electron impact mode (HR-EIMS) appeared at 327.0868 corresponding to C18H15O6 (M+-glucose). A positive Molisch\u2019s test reflected the glycosidic nature \u2212 at m/z 579.1509  with 17 degrees of unsaturation. Thirty carbon resonances were clear in the 13C-NMR and were sorted in a DEPT experiment into two oxymethylenes , 16 methines, and 12 quaternary carbons. The HR-EI-MS for both compounds showed strong peaks at 256.0735 with a relative abundance of 100% calculated for C15H12O4 and assigned to the aloe-emodin anthrone aglycone  in 10 and  in 11. These data matched data reported for caffeic acid + , m/z 507.1500 [M+H]+ , m/z 529.1320 [M+Na]+ , m/z 545.1069 [M+K]+ .White amorphous powder (1 mg); [\u03b1]23D + 16.8\u00b0 ; UV \u03bbmax MeOH (log \u03b5) nm: 209 (4.63), 244 (4.42), 300 (3.92), 330 (3.61); IR (KBr) vmax 3400, 1720, 1640, 1606, 1511, 1240 cm\u22121; 1H and 13C NMR .Red amorphous powder (6 mg); [\u03b1] NMR see ; HR-ESI-23D \u2212 4.7\u00b0 ; UV \u03bbmax MeOH (log \u03b5) nm: 209 (4.63), 244 (4.42), 300 (3.92), 330 (3.61); IR (KBr) vmax 3400, 1720, 1640, 1606, 1511, 1240 cm\u22121; 1H and 13C NMR , m/z 581.1651 [M+H]+ , m/z 603.1469 [M+Na]+ .Red amorphous powder (8 mg); [\u03b1] NMR see ; HR-ESI-t-test. Data were expressed as mean \u00b1 SD, and the difference was considered significant at p < 0.05 compared to the control. All statistical calculations used OriginLab software  and Microsoft Excel.Analysis of variance (ANOVA) was used to evaluate significance differences, followed by the Student\u2019s 3), 3,4 dihydro-6 glucopyranozyl-8-hydroxy-3--methyl-1H-[6), and two new anthraquinone derivatives, vacillantins A and B (10 and 11) were isolated from the leaves of A. vacillans (Asphodelaceae) together with nine known compounds . The structures of these compounds were elucidated through extensive spectroscopic analyses. The total alcohol extract and different fractions were tested for their antioxidant activities in five spectrophotometric assays. The dichloromethane fraction exhibited promising free radical scavenging activity in most of the assays. Our findings add new information to the literature on the structural diversity and pharmacological activities of Aloe species. Our results suggest A. vacillans as a potential source of secondary metabolites with pharmacological and industrial importance. Moreover, these results advocate further investigation of the remaining fractions with the aim of isolating bioactive compounds exhibiting interesting biological capacities.In summary, a new dihydroisocoumarin derivative, vacillanoside ("}
+{"text": "P < 0.001 for both comparisons). Positive resistin expression was significantly associated with tumor size, grade, stage, estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) status, and molecular classification; strongly positive resistin expression was associated with tumor grade, ER, PR, HER2 status, and molecular classification. Significantly positive correlations were observed between positive and strongly positive resistin expression and corresponding levels of EGFR expression. Relapse-free and overall survival was worse for patients with high levels of both proteins than for those with high levels of only one protein or normal levels of both proteins. Our evidence suggests that combined high levels of resistin and EGFR expression correlate with survival in patients with breast cancer.Elevated levels of resistin and epidermal growth factor receptor (EGFR) facilitate the development of breast cancer, although there are no reports of any correlation between these proteins. This study analyzed 392 human breast cancer tissue specimens and 42 samples of adjacent normal tissue. Rates of positive and strongly positive resistin expression were significantly higher in breast cancer tissue than in the adjacent nontumor tissue (83.2% vs. 23.8% and 20.9% vs. 0.0%, respectively;  Elevated levels of circulating resistin have been linked to a higher risk of breast cancer , 2, and Epidermal growth factor receptor (EGFR), a member of the HER family of receptor tyrosine kinases, is abnormally expressed and activated in many epithelial tumors and plays a critical role in the initiation and development of cancer via modulating downstream signaling pathways , 10. OurThis paper describes our insights derived from immunohistochemistry (IHC) analyses of resistin and EGFR expression in breast cancer and healthy normal breast tissue specimens obtained from 392 Chinese Han women. We detail the clinical prognostic significance of a positive association between resistin and EGFR expression in breast cancer.+/PR+[\u226520%]/HER2\u2013, Ki\u201067 < 14%); luminal B, containing hormone receptor-positive cases that did not meet the conditions of luminal A; HER2-enriched ; or triple-negative breast cancer (TNBC) . Follow-up information was available for 239 breast cancer patients with a median follow-up time of 60 months . The Ethics Committee of the Affiliated Dongyang Hospital of Wenzhou Medical University approved this study. All study methods satisfied the relevant guidelines and regulations issued by the Affiliated Dongyang Hospital of Wenzhou Medical University.Breast cancer tissue samples were obtained from 392 untreated Chinese Han women aged 24\u201390 years (median 50 years) who underwent surgery in the Affiliated Dongyang Hospital of Wenzhou Medical University  between 2007and 2019. Forty-two samples of adjacent normal breast tissue were also obtained following surgical resection. Pathohistological diagnoses followed the World Health Organization breast tumor classification criteria . HistoloWe followed the methods described by Wang et al. .We followed the methods of Wang et al. . The priThe entire section was scanned and scored independently by 2 pathologists. Resistin and EGFR expression staining intensity was scored on a 4-point scale from 0 (negative) to 1 (weak), 2 (moderate), or 3 (strong) . A case We followed the methods of Wang et al. .We followed the methods of Wang et al. . MultivaP < 0.001 and P = 0.001, respectively) (P = 0.012), grade (P < 0.001), stage (P = 0.042), ER (P < 0.001) and PR (P < 0.001) status, HER2 status (P = 0.021), and molecular classification (P < 0.001). Strongly positive resistin expression was associated with higher tumor grade (P < 0.001), ER (P < 0.001) and PR (P < 0.001) status, HER2 status (P = 0.001), and molecular classification (P < 0.001).The rate of positive resistin expression in breast cancer tissue specimens was 83.2% (326/392), which included 82 (20.9%) strongly positive cases; corresponding rates in normal breast tissue specimens were 23.8% (10/42) and 0.0% (0/42), respectively. Rates of positive and strongly positive resistin expression were significantly higher in breast cancer tissue than in normal breast tissue (ctively) . As showP < 0.001) and 15.2% [10/66] (P = 0.001), respectively, r = 0.250 and P < 0.001 and r = 0.166 and P = 0.001, respectively). Similarly, as shown in P < 0.001) and 27.4% [85/310] (P < 0.001), respectively). Spearman correlation analysis revealed significantly positive correlations between strongly positive levels of resistin expression and EGFR-positive or strongly positive expression in breast cancer tissue specimens . Similarly, a positive correlation was observed between staining intensity scores of resistin and EGFR in breast cancer tissues  (Rates of positive and strongly positive EGFR expression in breast cancer tissues were 52.0% (204/392) and 32.4% (127/392), respectively. When we analyzed the relationship between resistin and EGFR expression in breast cancer tissue specimens, we found significantly higher levels of positive or strongly positive EGFR expression among resistin-positive cases  compared with resistin-negative cases (24.2% [16/66] (< 0.001) .P = 0.237, P = 0.128, P = 0.171, and P = 0.105, respectively)   had a mean RFS of 51.5 months (an estimated 5-year RFS rate of 68.9%); patients whose tumors were either resistin-positive or strongly EGFR-positive (n = 129) had a mean RFS of 54.3 months (an estimated 5-year RFS rate of 83.7%), while patients whose tumors were resistin-negative and not strongly EGFR-positive (n = 36) had a mean RFS of 54.1 months . Patients whose tumors were both resistin-positive and strongly EGFR-positive experienced worse OS compared with patients whose tumors were either resistin-positive or strongly EGFR-positive and patients whose tumors were resistin-negative and not strongly EGFR-positive (P = 0.099).As shown in Figures n = 29) had a mean RFS of 49.1 months (an estimated 5-year RFS rate of 62.1%); patients whose tumors were either strongly resistin-positive or strongly EGFR-positive (n = 73) had a mean RFS of 54.2 months (an estimated 5-year RFS rate of 79.5%), and patients whose tumors were not strongly resistin-positive and not strongly EGFR-positive (n = 137) had a mean RFS of 53.9 months . Patients whose tumors were both strongly resistin-positive and strongly EGFR-positive experienced worse OS compared with patients whose tumors were either strongly resistin-positive or strongly EGFR-positive and patients whose tumors were not strongly positive for either protein (P = 0.453). Cox proportional hazards regression analysis did not reveal any significant associations between resistin-positive and strongly EGFR-positive tumor tissue (hazard ratio (HR) = 0.736, 95%CI = 0.198\u20132.734, P = 0.648; 0.765, 0.110\u20135.323, P = 0.786), strongly resistin-positive and strongly EGFR-positive tumor tissue , and RFS or OS.As shown in Figures n = 61) had a mean RFS of 51.3 months (an estimated 5-year RFS rate of 67.2%); patients whose tumors were either resistin-positive or strongly EGFR-positive (n = 111) had a mean RFS of 55.2 months (an estimated 5-year RFS rate of 86.5%), while patients whose tumors were resistin-negative and not strongly EGFR-positive (n = 36) had a mean RFS of 54.1 months . Non-TNBC patients whose tumors were both resistin-positive and strongly EGFR-positive experienced worse OS compared with patients whose tumors were either resistin-positive or strongly EGFR-positive and also patients whose tumors were resistin-negative and not strongly EGFR-positive (P = 0.065). As shown in Figures n = 23) had a mean RFS of 48.3 months (an estimated 5-year RFS rate of 60.9%); patients whose tumors were either strongly resistin-positive or strongly EGFR-positive (n = 59) had a mean RFS of 54.5 months (an estimated 5-year RFS rate of 78.0%), and patients whose tumors were not strongly resistin-positive and not strongly EGFR-positive (n = 126) had a mean RFS of 54.6 months . Non-TNBC patients whose tumors were both strongly resistin-positive and strongly EGFR-positive experienced worse OS compared with patients whose tumors were either strongly resistin-positive or strongly EGFR-positive and patients whose tumors were not strongly positive for either protein (P = 0.135).We analyzed the effect of combined high expression of resistin and EGFR on the prognosis of non-TNBC or TNBC, ER-negative or ER-positive, HER-negative or HER2-positive tumors. As shown in Figures As shown in Figures The cytokine resistin participates in several physiological and pathological processes, including metabolism, inflammation, autoimmunity, and various cancers, including breast cancer \u20136, 21\u201324\u03baB)/signal transducer and activator of transcription 3 (STAT3) signaling pathway-mediated induction of mesenchymal phenotypes and stemness properties [Our previous study have shown that EGFR promotes breast cancer invasion through downstream p44/p42 MAPK (ERK1/2) signaling , and resoperties . These rIn this study, both resistin and EGFR impacted adversely upon survival in breast cancer, but neither protein alone had a significant impact. Breast cancer patients whose tumors were both resistin-positive and strongly EGFR-positive, or were both strongly resistin-positive and strongly EGFR-positive, had worse RFS than all other breast cancer patients. In the non-TNBC and TNBC subgroup analyses, similar results were observed with non-TNBC, but not with TNBC. This may be due to the fact that there were too few cases for analysis. Further study should examine the effects of combined high resistin and EGFR expression on the prognosis of patients with TNBC. These results suggest that resistin and EGFR are potential clinical biomarkers of disease progression and prognosis in breast cancer and that simultaneously targeting these proteins is a potentially useful therapeutic strategy in this disease."}
+{"text": "Candida infections  C. glabrata, in cases of azole-resistant isolates , the general and long-term therapeutic use of FLZ (or other triazoles) can result in acquisition of molecular mechanisms that enable Candida isolates to exhibit antifungal resistance  , fluconafections , 3. Howeisolates . Except sistance . It is ksistance . Fenticoisolates . Thus, aisolates  but alsosistance .C. albicans (20 isolates) and C. glabrata (30 isolates) species, respectively. In each isolate\u2019s pair, the FLZ-resistant (derivative) isolate originated from the FLZ-susceptible or susceptible-dose-dependent  isolate following resistance development during patient infection  isolates exhibit known molecular resistance mechanisms, which consisted of upregulation of drug efflux pump-encoding genes  and/or point mutations of 14-\u03b1-lanosterol demethylase-encoding ERG11 gene . MIC values to FEZ and FLZ\u2014both obtained as standard powders from Sigma-Aldrich \u2014were determined using the protocol specified in the Clinical and Laboratory Standards Institute (CLSI) M27-A3 document without modifications -specific clinical breakpoints reported in the CLSI M27-S4 document (P\u2009<\u20090.05) differences between GM MIC values obtained for isolate groups from each species (see below), using repeated-measures analysis of variance (ANOVA) on log2 MICs followed by Bonferroni-Dunn\u2019s multiple-comparison test  from nfection . All excications . Only foson test .in vitro, 15 C. glabrata and 1 C. albicans isolate overexpressed drug efflux pumps alone, whereas 9 C. albicans isolates combined overexpression of drug efflux pumps and ERG11 amino acid substitution(s) (Table S1). For C. albicans isolates, FEZ MIC ranges were 0.25 to 2\u2009mg/liter among FLZ-nonresistant isolates  and 1 to 8\u2009mg/liter among FLZ-resistant isolates . For C. glabrata isolates, FEZ MIC ranges were 0.5 to 2\u2009mg/liter among FLZ-nonresistant isolates  and 0.5 to 4\u2009mg/liter among FLZ-resistant isolates . C. albicans and C. glabrata, respectively. Interestingly, the GM MICs \u00b1 CIs of C. albicans FEZ MICs in FLZ-nonresistant isolates differed significantly from that in FLZ-resistant isolates . Conversely, no significant difference was seen between the GM \u00b1 CIs of C. glabrata FEZ MICs in FLZ-nonresistant isolates and that in FLZ-resistant isolates .Of 25 isolates with molecular mechanisms contributing to the FLZ resistance phenotype observed Candida species, including C. albicans and C. glabrata. Remarkably, differences were more prominent in FLZ-resistant isolates than their nonresistant counterparts but were statistically significant only for C. albicans. Our data demonstrate that FEZ exhibits higher activity than FLZ. FEZ activity was less dependent on drug efflux pump-mediated FLZ resistance in Candida species such as C. glabrata. Based on these findings, FEZ should be evaluated as a candidiasis treatment, particularly in patients with recurrent or antifungal-recalcitrant Candida infections.In conclusion, we showed that FEZ MIC values were lower than FLZ MIC values in 50 well-characterized isolates from two clinically relevant"}
+{"text": "In contrast, after the subchronic administration of therapeutic-relevant concentration of valproate (VPA), acute administration of therapeutic-relevant concentration of CLZ drastically increased Cx43-associated astroglial releases. VPA increased Cx43 expression in cytosol fraction without affecting plasma membrane fraction, whereas CLZ increased Cx43 expression in both fractions. Acute administration of therapeutic-relevant concentration of CLZ drastically increased Cx43 expression in the plasma membrane fraction of astrocytes subchronically treated with VPA. The present findings suggest that CLZ-induced the activation of Cx43-associated channel activity and transported Cx43 to plasma membrane, probably contribute to the double-edged sword clinical action of CLZ, such as improvement of cognitive dysfunction and CLZ-induced myocarditis.Clozapine (CLZ) is a gold-standard antipsychotic against treatment-refractory schizophrenia, but is one of the most toxic antipsychotic agents. Pharmacological mechanisms of the double-edged sword clinical action of CLZ remain to be clarified. To explore the mechanisms of CLZ, the present study determined the astroglial transmission associated with connexin43 (Cx43), which is the most principal expression in astrocytes and myocardial cells, and expression of Cx43 in primary cultured astrocytes. Both acute and subchronic administrations of CLZ concentration-dependently increased Cx43-associated astroglial release of  N-methyl-d-aspartate (NMDA)/glutamate receptor (NMDAR) contributes to the pathophysiology of schizophrenia [It has been established that dysfunctions of both dopaminergic and glutamatergic transmission play important roles in the pathophysiology of schizophrenia, with various antipsychotics improving dysfunctions of mesolimbic and mesocortical dopaminergic transmissions with thalamocortical glutamatergic transmission ,4,5,6,7;ophrenia ,10,11,12ophrenia ,14 and eophrenia . Moreoveophrenia . Based ol-glutamate and d-serine induced by CLZ are one of the major mechanisms of agonistic action of CLZ against impaired NMDAR [Clozapine (CLZ) is considered the double-edged sword antipsychotic drug, since CLZ is one of the most effective but toxic antipsychotics against antipsychotics-resistant schizophrenia . In facted NMDAR .In terms of management of CLZ-induced seizure, possible strategies include reduction of CLZ dose and addition of antiepileptic drugs, including valproate (VPA), which is the most commonly used for management of CLZ-induced seizures ,21 due tBoth clinical and preclinical studies have emphasized that thalamocortical disturbance is particularly relevant for cognitive dysfunction in several neuropsychiatric disorders, including schizophrenia, Attention-deficit hyperactivity disorder (ADHD), intellectual disability, autism, and epileptic psychosis ,9,26,27.Accumulating evidences indicate connexin (Cx) composed transmembrane channels (Cxs) are crucial to the coordination and maintenance of physiologic activity including neuronal excitability, synaptic plasticity, tripartite synaptic transmission, and homeostasis maintenance in the central nervous system ,29. Cx il-glutamate and d-serine associated with Cxs and the astroglial expression of Cx43 using primary cultured astrocytes.Our recent preclinical study demonstrated that CLZ acutely activated astroglial Cxs activity, in a concentration-dependent manner. Clinically, survival analysis has suggested that the lower limit of the therapeutic range of CLZ serum concentration is 0.6 \u03bcM , whereasClozapine (CLZ) was purchased from Sigma . Sodium valproate (VPA) was obtained from Wako Chemicals . The hemichannel/gap junction (Cxs) blocker, carbenoxolone (CBX) , and selAll animal care and experimental procedures described in this report complied with the Ethical Guidelines established by the Institutional Animal Care and Use Committee at Mie University, Japan (No. 2019-3-R1) and are reported in accordance with the Animal Research: Reporting of In Vivo Experiments (ARRIVE) guidelines . AstrocyN = 48) sacrificed by decapitation at 0\u201324 h of age. The cerebral hemispheres were removed under dissecting microscope. Tissue was chopped into fine pieces using scissors and then triturated briefly with micropipette. Suspension was filtered using 70 \u00b5m nylon mesh  and centrifuged. Pellets were then resuspended in 10 mL Dulbecco\u2019s modified Eagle\u2019s medium containing 10% fetal calf serum (fDMEM), which was repeated three times. After culture for 14 days (DIV14), contaminating cells were removed by shaking in standard incubator for 16 h at 200 rpm. On DIV21, astrocytes were removed from flasks by trypsinization and seeded directly onto translucent poly ethylene terephthalate (PET) membrane (1.0 \u03bcm) with 24-well plates (BD) at a density of 1 \u00d7 105 cells/cm2 for experiments From DIV21 to DIV28, the culture medium (fDMEM) was changed twice a week, and CLZ  or VPA  was added for subchronic administrations (7 days). On DIV28, cultured astrocytes were washed out using ACSF, and this was repeated three times. The ACSF comprised NaCl 150.0 mM, KCl 3.0 mM, CaCl2 1.4 mM, MgCl2 0.8 mM, and glucose 5.5 mM, buffered to pH 7.3 with 20 mM HEPES buffer. The remaining adherent cells contained 95% GFAP-positive and A2B5-negative cells, detected using immunohistochemical staining [2 incubator (pretreatment incubation). After the pretreatment, astrocytes were then incubated in ACSF, 50 mM K+  or 100 mM K+  containing the same agents of pretreatment (20 min) and collection of the ACSF, MK-ACSF, or HK-ACSF for analysis. Each 100 \u03bcL of collected ACSF, MK-ACSF, or HK-ACSF was filtered by Vivaspin 500-3K  and freeze-dried for storage at \u221280 \u00b0C until needed for analyses.Pregnant Sprague-Dawley rats , which were housed individually in cages, were kept in air-conditioned rooms  set at 12 h light/dark cycle, with free access to food and water. Cultured astrocytes were prepared from cortical astrocyte cultures of neonatal Sprague-Dawley rats . Both plasma membrane and cytosol fractions were solubilized by radio immunoprecipitation assay buffer  containing protease inhibitor cocktail .+ containing ACSF), or HK-ACSF (100 mM K+ containing ACSF) containing the same agent of pretreatment incubation for 20 min. Especially, to determine the astroglial releases associated with Cxs, after pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent of pretreatment incubation with Cx43 selective inhibitor, CAP19 (20 \u03bcM), or nonselective Cxs inhibitor, CBX (100 \u03bcM), for 20 min.During DIV21 to DIV28, astrocytes were incubated in fDMEM, not containing any target agents. On DIV28 during pretreatment incubation, astrocytes were incubated in ACSF containing CLZ  for 60 min. After pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF . On DIV28 during pretreatment incubation, astrocytes were incubated in ACSF containing the same agent for 60 min. After pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent for 20 min. Especially, to determine the astroglial releases associated with Cxs, after pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent with CAP19 (20 \u03bcM) or CBX (100 \u03bcM) for 20 min.l-glutamate and d-serine, during DIV21 to DIV28, astrocytes were incubated in fDMEM, not containing any target agents. On DIV28 during pretreatment incubation, astrocytes were incubated in ACSF containing VPA  for 60 min. After pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent of pretreatment incubation for 20 min.To determine the effects of acute administration of VPA on astroglial releases of l-glutamate and d-serine, during DIV21 to DIV28, astrocytes were incubated in fDMEM containing VPA . On DIV28 during pretreatment incubation, astrocytes were incubated in ACSF containing the same agent for 60 min. After pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent of pretreatment incubation for 20 min.To determine the effects of subchronic administration of VPA on astroglial releases of During DIV21 to DIV28, astrocytes were incubated in fDMEM containing CLZ . On DIV28 during pretreatment incubation, astrocytes were incubated in ACSF containing the same agent (CLZ) with therapeutic-relevant concentration of VPA (1000 \u03bcM) for 60 min. After pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent of pretreatment incubation for 20 min.During DIV21 to DIV28, astrocytes were incubated in fDMEM containing the therapeutic-relevant concentration of VPA (1000 \u03bcM). On DIV28 during pretreatment incubation, astrocytes were incubated in ACSF containing the same agent (1000 \u03bcM VPA) with CLZ  for 60 min. After pretreatment incubation, astrocytes were incubated in ACSF, MK-ACSF, or HK-ACSF containing the same agent for 20 min.l-cysteine and o-phthalaldehyde [l-cysteine (2 mg) and o-phthalaldehyde (1 mg) in 0.1 mL ethanol, followed by the addition of 0.9 mL sodium borate buffer . Automated pre-column derivatives were obtained by drawing up a 5 \u03bcL aliquot of the standard or blank solution and 5 \u03bcL of the derivative reagent, and holding them in vials for 5 min before injection. The derivatized samples (5 \u03bcL) were injected by autosampler . Analytical column  was maintained at 45 \u00b0C and a flow rate of 500 \u03bcL/min. Linear gradient elution was performed over 10 min in mobile phases A  and B . The excitation/emission wavelengths of the fluorescence detector were set at 280 and 455 nm, respectively [Levels of L-glutamate and GABA were determined by the fluorescence resonance energy transfer method ,47,48 usaldehyde ,26,27. Dectively ,47,48.Simple Western analyses were performed using Wes  according to the ProteinSimple user manual . Lysate N = 6) that were predetermined based on our previous studies [p value less than 0.05 was considered statistically significant. Statistical analyses were performed in BellCurve for Excel Version 3.2. . Astroglial transmitter concentrations were analyzed by Mauchly\u2019s sphericity test, followed by multivariate analysis of variance (MANOVA) using BellCurve for Excel. When the data did not violate the assumption of sphericity (p > 0.05), the F-value of MANOVA was analyzed using sphericity-assumed degrees of freedom. However, if the assumption of sphericity was violated (p < 0.05), the F-value was analyzed using Chi-Muller\u2019s-corrected degrees of freedom. When the F-value for the agent/concentration factors of MANOVA was significant, the data was analyzed by Tukey\u2019s post hoc test. Protein expression of Cx43 in cytosol and plasma membrane fractions was analyzed by two-way analysis of variance (ANOVA) with Tukey\u2019s post hoc test using BellCurve for Excel.All experiments were designed with equal-sized groups  = 39.4 (p < 0.01), FCLZ = 210.0 (p < 0.01), Fstimulation*CLZ = 114.4 (P < 0.01)) and D-serine  = 55.4 (p < 0.01), FCLZ = 179.5 (p < 0.01), Fstimulation*CLZ = 86.7 (p < 0.01)) (+-evoked stimulation (50 and 100 mM) concentration-dependently increased astroglial releases of L-glutamate and D-seine  concentration-dependently increased basal and K< 0.01)) A,B. The  D-seine A,B. The +-evoked astroglial releases of L-glutamate  = 8.5 (p < 0.01), FCLZ = 259.1 (p < 0.01), FGAP19*CLZ  = 22.6 (p < 0.01)) and D-serine  = 11.8 (p < 0.01), FCLZ = 220.8 (p < 0.01), FGAP19*CLZ = 13.8 (p < 0.01))  = 20.6 (p < 0.01), FCLZ = 226.1 (p < 0.01), FCBX*CLZ = 66.3 (p < 0.01)) and D-serine  = 26.5 (p < 0.01), FCLZ = 178.1 (p < 0.01), FCBX*CLZ = 48.1 (p < 0.01)) , decreased the CLZ-induced 100 mM K< 0.01)) A,B. The < 0.01)) A,B. Ther+-evoked astroglial releases of L-glutamate  = 67.1 (p < 0.01), FCLZ = 268.1 (p < 0.01), Fstimulation*CLZ = 128.9 (p < 0.01)) and D-serine  = 80.6 (p < 0.01), FCLZ = 287.8 (p < 0.01), Fstimulation*CLZ = 125.2 (p < 0.01)) (+-evoked stimulation (50 and 100 mM) concentration-dependently increased astroglial releases of L-glutamate and D-seine  concentration-dependently increased basal and K< 0.01)) A,B. The  D-seine A,B. The +-evoked astroglial releases of L-glutamate  = 19.7 (p < 0.01), FCLZ = 276.5 (p < 0.01), FGAP19*CLZ = 27.0 (p < 0.01)) and D-serine  = 22.7 (p < 0.01), FCLZ = 336.1 (p < 0.01), FGAP19*CLZ = 18.6 (p < 0.01))  = 35.6 (p < 0.01), FCLZ = 279.7 (p < 0.01), FCBX*CLZ = 72.0 (p < 0.01)) and D-serine  = 38.3 (p < 0.01), FCLZ = 296.3 (p < 0.01), FCCBX*CLZ = 52.0 (p < 0.01)) , decreased the CLZ-induced 100 mM K< 0.01)) A,B. The < 0.01)) A,B. Ther+-evoked astroglial releases of L-glutamate or D-serine  = 13.6 (p < 0.01), FVPA = 97.9 (p < 0.01), Fstimulation*VPA = 16.4 (p < 0.01)) and D-serine  = 16.5 (p < 0.01), FVPA = 66.0 (p < 0.01), Fstimulation*VPA = 13.1 (p < 0.01)) (+-evoked releases of L-glutamate and D-serine were increased by supratherapeutic concentration of VPA (3000 \u03bcM), but not affected by therapeutic-relevant concentration of VPA (300\u20131000 \u03bcM)  did not affect basal or KD-serine A,B. Cont< 0.01)) C,D. Basa1000 \u03bcM) C,D.+-evoked astroglial releases of L-glutamate and D-serine    A,B. Acut\u2013100 \u03bcM) A,B.+-evoked astroglial releases of L-glutamate and D-serine  = 37.9 (p < 0.01), FCLZ = 439.4 (p < 0.01), FVPA*CLZ = 265.6 (p < 0.01)) and D-serine  = 48.3 (p < 0.01), FCLZ = 270.2 (p < 0.01), FVPA*CLZ = 184.9 (p < 0.01)) (+-evoked releases of L-glutamate (from 100 \u03bcM to 10 \u03bcM) and D-serine (from 30 \u03bcM to 10 \u03bcM).Acute administration of CLZ concentration-dependently increased KD-serine A,B. Subc< 0.01)) A,B. Subc+-evoked releases of L-glutamate  = 30.1 (p < 0.01), FCLZ = 291.8 (p < 0.01), FVPA*CLZ = 40.3 (p < 0.01)) and D-serine  = 16.3 (p < 0.01), FCLZ = 94.7 (p < 0.01), FVPA*CLZ = 11.5 (p < 0.01)) (+-evoked releases of L-glutamate (from 30 \u03bcM to 3 \u03bcM) and D-serine (from 10 \u03bcM to 3 \u03bcM).Subchronic administration of therapeutic-relevant concentration of VPA (1000 \u03bcM) increased astroglial CLZ-induced 100 mM K< 0.01)) A,B. SubcFragment = 7.4 (p < 0.01), FLevel = 15.0 (p < 0.01), FFragment*Level = 3.8 (p < 0.05))  = 5.0 (p < 0.05), FLevel = 22.1 (p < 0.01), FFragment*Level = 6.2 (p < 0.01)) (p < 0.05) (Subchronic administrations of VPA (1000 and 3000 \u03bcM) concentration-dependently increased Cx43 expression in cytosol fraction without affecting that in plasma membrane fraction (F< 0.05)) A. Contra< 0.01)) B. The th < 0.05) B.Fragment = 33.4 (p < 0.01), FLevel = 15.6 (p < 0.01), FFragment*Level = 17.0 (p < 0.01)) (p < 0.01)  concentration-dependently increased Cx43 expression in plasma membrane but not cytosol fraction of astrocytes subchronically treated with therapeutic-relevant concentration of VPA (1000 \u03bcM) (F< 0.01)) C. The th < 0.01) C.+-evoked astroglial releases of L-glutamate and D-serine were 100 \u03bcM, 30\u2013100 \u03bcM and 10\u201330 \u03bcM, respectively. The K+-evoked astroglial release was composed of astroglial exocytosis [2+, K+, and plasma membrane voltage [+-evoked astroglial releases of L-glutamate and D-serine. These results suggest that the K+-evoked astroglial releases of L-glutamate and D-serine, at least partially, comprise the output through Cxs. Therefore, acute administration of CLZ concentration-dependently enhances astroglial releases of L-glutamate and D-serine associated with Cxs, but the stimulatory effects of CLZ on astroglial releases were observed in the supratherapeutic range. The therapeutic concentration of VPA was considered, ranged 350\u2013700 \u03bcM [+-evoked astroglial releases of L-glutamate and D-serine. Therefore, acute administration of VPA did not affect Cxs activities.The present study demonstrated that CLZ enhanced astroglial releases of L-glutamate and D-serine, in a concentration-dependent manner. Survival analysis has suggested that effective relapse prevention requires the maintenance of patients at CLZ serum concentrations above 0.6 \u03bcM ; howeverocytosis ,24,44, oocytosis ,41,42, a voltage . During  voltage . In the 0\u2013700 \u03bcM . Neither+-evoked astroglial releases of L-glutamate and D-serine. The stimulatory effects of subchronic administration of CLZ on basal and K+-evoked astroglial releases were predominant compared with those of acute CLZ administration, since the threshold concentrations of subchronic administration of CLZ on basal, 50 mM and 100 mM K+-evoked astroglial releases of L-glutamate and D-serine were reduced to 30 \u03bcM (acute: 100 \u03bcM), 30 \u03bcM (acute: 30\u2013100 \u03bcM), and 10 \u03bcM (acute: 10\u201330 \u03bcM), respectively. Similar to CLZ, subchronic administration of supratherapeutic concentration of VPA (3000 \u03bcM) increased K+-evoked astroglial releases of L-glutamate and D-serine without affecting basal release. These discrepancies between acute and subchronic administrations of CLZ and VPA on basal and K+-evoked astroglial releases of L-glutamate and D-serine suggest that subchronic administration of CLZ and VPA possibly increases expression of Cxs in plasma membrane.Contrary to acute administrations, chronic administration of CLZ also increased basal and KIndeed, simple western analysis detected the subchronic administration of CLZ and VPA increased astroglial expression of Cx43, which is the principal Cx isoform in astrocytes ; howeverThe Cx gene regulation system, transcriptional factors , and epigenetic processes  have been identified ,52. HistContrary to VPA, CLZ activates the inhibitory and stimulatory expression regulating system on Cx43 synthesis, such as HDAC2, Wnt pathway, transcription factor specificity protein 1, and activator protein 1 complex ,60,61,62Cx43 is the principal Cxs in astrocytes, but Cx26 and Cx30 are also expressed in astrocytes [The incidence of seizures with CLZ is ranged from 2% to 7.5%, and the risk of CLZ-induced seizures increased with higher doses . The risIn the present study, neither acute nor subchronic administrations of therapeutic-relevant concentration of CLZ (1\u20133 \u03bcM) affected astroglial transmission; however, acute administration of therapeutic-relevant concentration of CLZ drastically increased astroglial releases of L-glutamate and D-serine from astrocytes which were chronically administrated with therapeutic-relevant concentration of VPA (1000 \u03bcM). Therapeutic-relevant concentration of CLZ (3 \u03bcM) also drastically increased Cx43 expression in the plasma membrane fraction without affecting that in cytosol fraction of therapeutic-relevant concentration of VPA-administrated astrocytes. These results suggest that increased Cx43 in the cytosol fraction induced by subchronic administration of therapeutic-relevant concentration of VPA (1000 \u03bcM) is transported to plasma membrane by CLZ. The rapid titration of CLZ dose and VPA administration at the commencing CLZ are significantly associated with increasing risks of CLZ-induced myocarditis/cardiomyopathies . It has The present study determined the concentration- and time-dependent effects of CLZ and VPA on astroglial transmission of L-glutamate and D-serine associated with Cx43, to explore the mechanisms of the multimodal action of CLZ. Both acute and subchronic administrations of CLZ and VPA increased astroglial releases of L-glutamate and D-serine concentration-dependently, but the therapeutic-relevant concentration of neither CLZ nor VPA affected these releases. The stimulatory effects of subchronic administrations of CLZ and VPA on astroglial releases were more predominant compared with those of acute administrations. Subchronic administrations of both VPA and CLZ concentration-dependently increased Cx43 expression in astrocytes, but the therapeutic-relevant concentration of neither CLZ nor VPA affected Cx43 expression. Especially, VPA increased Cx43 expression in cytosol fraction of astrocytes, whereas CLZ increased Cx43 expression in both cytosol and plasma membrane fractions. After subchronic administration of CLZ, acute administration of therapeutic-relevant concentration of VPA did not affect CLZ-induced astroglial transmitter releases; however, after subchronic administration of therapeutic-relevant concentration of VPA, acute administration of CLZ drastically increased astroglial transmitter releases. Therapeutic-relevant concentration of CLZ alone could not affect astroglial release, whereas after the subchronic administration of therapeutic-relevant concentration of VPA, acute administration of therapeutic-relevant concentration of CLZ could increase astroglial transmitter releases. During subchronic administration of therapeutic-relevant concentration of VPA, acute administration of therapeutic-relevant concentration of CLZ enhanced the transport of cytosol Cx43 to plasma membrane. Therefore, CLZ enhances astroglial functional Cxs via activation of transport of cytosol Cx43 to plasma membrane. These results suggest that the hyperactivation of astroglial Cxs activities induced by supratherapeutic concentration or rapid titration of CLZ, at least partially, contributes to CLZ-induced seizure, but inhibitory effects of VPA on CLZ-induced seizure are not modulated with astroglial transmission associated with Cxs. Interestingly, VPA intake at the commencing CLZ increases risk of CLZ-induced myocarditis/cardiomyopathies, probably via reciprocal activation of Cxs between VPA and CLZ."}
+{"text": "Kocuria varians 80 was sequenced and assembled into a single 2.82-Mb chromosome. Genome sequence comparison of K. varians strain 80 and previously sequenced strains revealed predicted proteomic differences that can impact its technological properties.The genome of the meat starter culture strain  Kocuria varians 80 was sequenced and assembled into a single 2.82-Mb chromosome. Genome sequence comparison of K. varians strain 80 and previously sequenced strains revealed predicted proteomic differences that can impact its technological properties.The genome of the meat starter culture strain  Kocuria varians is used as a starter culture for processed meat and dairy products. In combination with other starter cultures or by itself, it can participate in proteolysis, denitrification, and biogenic amine degradation (\u2013Staphylococcus carnosus TM300 (Staphylococcus xylosus (Lactobacillus sakei (Kocuria varians strain 80 (SSCA-2165).radation \u20133, whichus TM300 , Staphyl xylosus , Lactobaus sakei , and othKocuria varians strain 80 was received from State Scientific Center for Antibiotics Culture  and was originally isolated from dried fermented sausages in 1978 . We used the NEBNext Ultra DNA library prep kit  for Illumina HiSeq 2500 sequencing. For PacBio RS II single-molecule real-time (SMRT) sequencing, we used the SMRTbell template prep kit v1.0  following the 20-kb template preparation protocol using the BluePippin size selection system. In total, there were 2.1 billion (2\u2009\u00d7\u2009250-bp) Illumina reads and 158,304 PacBio reads .K. varians was assembled with SPAdes v3.11 , and K. varians G6 (GCA_900014985.1). After circularization using Simple Circularise v1.0 (https://github.com/Kzra/Simple-Circularise), the genome assembly consisted of a single contig with a 2,823,038-bp circular chromosome . The genome has a 70.55% GC content. We found 2,465 protein-coding sequences, 48 tRNA genes, and 9 rRNA genes.After read and adapter trimming with Trimmomatic v0.36  with thees v3.11 , with eres v3.11 ; every pes v3.11  search. es v3.11  for K. vK. varians C6, both K. varians 80 and K. salsicia G1 contain the xylB gene, according to KEGG pathway analysis. Also unlike K. varians G6 and K. salsicia G1, strain 80 has a complete metabolic pathway from d-glucose-1P to l-rhamnose, which is potentially important to its properties as a meat-fermenting starter culture.Unlike SRR13177364 and SRR13177365, respectively. The genome sequence of K. varians strain 80 (SSCA-2165) has been deposited at GenBank under accession number GCF_002250745.2.The PacBio and Illumina reads were submitted to the SRA under accession numbers"}
+{"text": "J Clin Endocrinol Metab. 2020;105(8):2515\u20132531; doi: 10.1210/clinem/dgaa238), the following transcriptional error occurred in the published paper: \u201cThe y-axis title for Figure 3 (page 9) incorrectly describes the plasma testosterone (T) concentration in units of ng/mL instead of ng/dL.\u201dIn the above-named article by Swerdloff RS, Wang C, White WB, Kaminetsky J, Gittelman MC, Longstreth JA, Dudley RE, and Danoff TM (The authors have provided an updated version of Figure 3.10.1210/clinem/dgaa238doi:"}
+{"text": "Common clinical features of patients with Coronavirus disease-2019 (COVID-19) vary from fever, to acute severe respiratory distress syndrome. Several laboratory parameters are reported as indicators of COVID-19 severity. We hereby describe the possible novel severity biomarkers for COVID-19, CD11b+CD33+HLA-DR-CD14+ cells and CD11b+CD33+HLA-DR-CD66b+ cells. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiologic agent of COVID-19. COVID-19 has already led to massive infection and deaths worldwide and was declared a pandemic by the World Health Organization.Common clinical features of patients with COVID-19 vary from fever, cough, dyspnea (among others) to acute severe respiratory distress syndrome (ARDS), and shock. Reports of fatal outcomes in hospitalized patients vary from 6.2 to 21.5%  in patients recently infected with SARS-CoV-2.n = 16, 7 males, 9 females, median age 49.7 years).We investigated the presence of CD33+CD11b+HLA-DR-CD14+ and CD33+CD11b+HLA-DR-CD66b+ cells in the peripheral blood of 104 patients infected with COVID-19, 62 males and 42 females. The diagnosis of COVID-19 was confirmed by the detection of SARS-CoV-2 RNA by reverse-transcriptase polymerase chain reaction (RT-PCR), and patients tested negative for respiratory syncytial virus (RSV) and influenza. The study was approved by the Ethics Committee of Hospital das Cl\u00ednicas da Faculdade de Medicina da Universidade de S\u00e3o Paulo\u2014HCFMUSP (no. 30800520.7.0000.0068-2020) and was carried out in conformity with the 2013 revision of the Declaration of Helsinki. Healthy controls (HC), without SARS-CoV-2, RSV or influenza infection, were recruited for flow cytometry assays .During hospitalization, COVID-19 patients received systemic treatment. All patients received antibiotics and anticoagulants, 45/104 systemic corticosteroids, 39/104 antivirals. EDTA plasma samples were obtained from a single venipuncture. Laboratory analysis was performed at the Central Laboratory of Hospital das Clinicas, Faculdade de Medicina da Universidade de S\u00e3o Paulo , and included: complete blood counts (CBC), coagulogram, liver enzymes , alkaline phosphatase (AP), bilirubin, urea, creatinine, glucose, sodium, potassium, lactate dehydrogenase, magnesium, phosphorus, total proteins  and fractions immunoglobulins, CRP, ferritin, pH, pO2, pCO2, D-dimer, and erythrocyte sedimentation rate in arterial blood collected in K2EDTA tubes.P < 0.05).Flow cytometry analysis was performed using 0.1 mL of whole blood in K2EDTA collection tubes . Samplesp = 0.0382), pCO2 (p = 0.0167), mean corpuscular volume (p = 0.0226), enhanced leukocytes (p = 0.0001), neutrophils (p = 0.0001), neutrophil-to-leukocyte ratio (p = 0.0001), and augmented levels of alkaline phosphatase (p = 0.0150), total bilirubin (p = 0.0363), direct bilirubin (p = 0.0113), glutamyl transferase gamma (p = 0.0388), creatinine (p = 0.0121), lactate dehydrogenase (p = 0.0121), CRP (p = 0.0408), urea (p = 0.0016) and D-dimer (p = 0.0012)  .Such laboratory parameters reinforce the need for biomarkers such as neutrophil-to-lymphocyte ratio, urea and d-dimer as a predictive factor of disease outcome , 4. MorePrevious reports verified an immune dysregulation in circulating immune cells in the blood of COVID-19 patients, with a significant increase in neutrophil-to-lymphocyte ratio in COVID-19, and a further increase in severe COVID-19 . Here, wp = 0.0471). We also verified an increase in CD33+CD11b+HLA-DR-CD14+ cells in the blood of ICU patients  in comparison with HC individuals (p < 0.0001) and GW patients (p = 0.0388)  .p = 0.0178). Again, we also verified an increase in CD33+CD11b+HLA-DR-CD66b+ cells in the blood of ICU patients  in comparison with HC individuals (p < 0.0001) and GW patients (p = 0.0175)  patients in comparison with HC individuals  ( 0.0175) .Overall, these findings suggest a correlation between the quantification of CD33+CD11b+HLA-DR-CD14+ cells and CD33+CD11b+HLA-DR-CD66b+ cells and the severity of COVID-19. These cells have been previously characterized as myeloid derived suppressor (MDSC) cells by this cytometry panel , but furEnhanced CD33+CD11b+HLA-DR\u2013CD14\u2013CD66b+ and CD33+CD11b+HLA-DR\u2013CD14+CD66b\u2013 cells in the blood of ICU patients, classified as severe COVID patients, could represent not only a predictor of prognosis for COVID-19, but also specific therapy targets for treating this virus infection in the near future.All datasets generated for this study are included in the article/supplementary material.This studies involving human participants were reviewed and approved by Ethics Committee of Hospital das Cl\u00ednicas da Faculdade de Medicina da Universidade de S\u00e3o Paulo\u2014HCFMUSP (no. 30800520.7.0000.0068-2020). The patients/participants provided their written informed consent to participate in this study.RA and MS: conception, performed experiments, analysis, write, and review. MAn, AB, AP, NP, IF, LO, FT, DB, EO, SG-S, YR, and CdB: performed experiments and review. MAr, RO, VA, and AD: patient care and review. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Technology in Cancer Research & Treatment. 2020;19: 1-8. doi: 10.1177/1533033819892251Xuefang Z, Ruinian Z, Liji J, et al. miR-331-3p Inhibits Proliferation and Promotes Apoptosis of Nasopharyngeal Carcinoma Cells by Targeting elf4B-PI3K-AKT Pathway. The published version of Figure\u00a02 is incorrect. The correct version of the figure is as given below:"}
+{"text": "Sir,Escherichia coli have identified a globally disseminated high-risk clone named ST131, with strains belonging to three clades  and three different subclades . While C2 is associated with CTX-M-15, clade C1-M27 has been associated with CTX-M-27.blaCTX-M-27 nor E. coli ST131 C1-M27 have been reported so far.Surveillance studies of ESBL-producing Perna perna) and oysters (Crassostrea spp.) were collected from 14 near-shore sites located at different distances from the port of Santos (the largest port of Latin America). Mussel (n\u2009=\u200910) and oyster (n\u2009=\u200910) samples, collected from each site, were placed into sterile plastic bags. The samples were kept refrigerated and processed within 3\u2009h after collection. Following standard methods for the examination, 25\u2009g of bivalves were distributed in sterile plastic bags containing 225\u2009mL of Brain Heart Infusion broth and incubated at 37\u00b0C for 24\u2009h. Subsequently, the samples were streaked onto MacConkey agar plates supplemented with ceftriaxone (2\u2009mg/L), meropenem (2\u2009mg/L) or colistin (2\u2009mg/L), following incubation at 37\u00b0C for 24\u2009h.During a local surveillance study conducted to monitor the presence of WHO critical priority pathogens in impacted marine ecosystems, brown mussels (E. coli isolates were recovered from mussel (E. coli 6M) and oyster (E. coli MO) samples collected from two different sites  close to the port. Antimicrobial susceptibility testing, performed by disc diffusion and/or Etest methods,,E. coli MO was resistant to nalidixic acid (>32\u2009mg/L) and ciprofloxacin (>4\u2009mg/L). PCR screening and Sanger sequencing revealed that these isolates were positive for the blaCTX-M-27 ESBL gene.Two ceftriaxone-resistant E. coli strains were subjected to WGS using the Illumina NextSeq (2\u205f\u00d7\u205f150\u2009bp) platform . De novo assemblies were performed using Spades v. 3.11. WGS data were analysed using bioinformatics tools available from the Center for Genomic Epidemiology (www.cge.dtu.dk).E. coli 6M (accession number: NCWA00000000.1) belonged to serotype O86:H18 and sequence type ST38/CC38, whereas E. coli MO (accession number: NCVZ00000000.1) belonged to serotype O25b:H4 and ST131/CC131. These STs have been globally disseminated among humans, animals and aquatic environments, being commonly associated with CTX-M variants.,,E. coli 6M revealed the presence of iss , astA (EAST-1 toxin), eatA (enterotoxigenic autotransporter A), capU (hexosyltransferase homologue), nfaE (diffuse adherence fibrillar adhesin) and eilA  genes, whereas iha (adherence protein), sat (secreted autotransporter toxin), gad (glutamate decarboxylase), senB (enterotoxin) and iss genes were found in E. coli MO. Moreover, E. coli MO carried fimH30 (associated with ST131) and the C1 subclade-specific prophage-like region (M27PP1).iha, sat, gad, iss and senB genes) of E. coli MO was identical to that of other E. coli strains of ST131 and C1-M27 clade.,strA, strB and aadA5), \u03b2-lactams (blaCTX-M-27), sulphonamides (sul1 and sul2), trimethoprim (dfrA17) and tetracycline [tet(A)], as previously observed in E. coli of ST131 and C1-M27;2,3 whereas mutations in the quinolone resistance-determining regions of gyrA , parC  and parE (Ile529Leu) genes were only identified in the E. coli MO strain. FIB and FII, and Col156, FIA, FIB and FII replicon types were identified in E. coli 6M and MO strains, respectively.blaCTX-M-27 genes, was achieved by bacterial transformation using E. coli TOP10. FIB and FII replicons were identified in p6M (FAB formula F2:A\u2212:B10), whereas FIA, FIB and FII replicon types were confirmed in pMO (FAB formula F1:A2:B20). The complete sequence of the pMO plasmid (GenBank accession no. MG886288) was obtained using de novo assembly, followed by gap closure by PCR and Sanger sequencing.Mobilization of plasmids \u223c130\u2009kb in size (named pMO and p6M), bearing blaCTX-M-27, the pMO plasmid harboured aadA5, sul1, dfrA17, tet(A) and mphA resistance genes, similarly to F1:A2:B20 plasmids harboured by the C1-M27 clade , of which 129 CDS encoded proteins with known functions . Besides blaCTX-M-27 genes, carried by both E. coli strains, revealed the presence of IS26 and IS903 mobile elements, E. coli MO presented a truncated IS903 upstream of the blaCTX-M-27 gene, whereas E. coli 6M presented a truncated ISEcp1 downstream of the blaCTX-M-27 gene, and tonB and ORF genes (Figure\u2009Although analysis of the genetic environment of E. coli strains, of ST131 and clade C1-M27, in Brazil. In this regard, since CTX-M-27-positive E. coli strains were recovered from areas impacted by intensive maritime traffic and transoceanic shipping activities, a possible introduction of international clones via commercial shipping routes could be speculated.E. coli in South American countries remains necessary to elucidate the local epidemiology and dynamics of the transmission of high-risk clades with pandemic potential.In summary, to our knowledge, we report the first identification of CTX-M-27-producing"}
+{"text": "T isolated from the roots of Acacia mangium Willd. showed potential plant growth promoting (PGP) activity. Phylogenetic analysis, based on 16S rRNA gene, indicated that strain GKU 173T was the most closely related to Fodinicola feengrottensis HKI 0501T\u2014the only species in the genus Fodinicola. Morphology and chemotaxonomy of strain GKU 173T indicated that it belongs to the genus Fodinicola by having meso-diaminopimelic acid in the cell wall and xylose as the characteristic cell-wall sugars. The cellular fatty acid profile mainly comprised iso-C16:0, anteiso-C17:0, iso-C18:0, and iso-C17:0. The major menaquinones were MK-9(H4), MK-9(H6), and MK-9(H8). The main polar phospholipids contained diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). Genome analysis signified DNA G+C content of 67.81 mol%. The level of digital DNA-DNA relatedness between strain GKU 173T and the type strain was 21.30%. On the basis of polyphasic characteristics, strain GKU 173T clearly represents a novel species of the genus Fodinicola, for which the name Fodinicolaacaciae sp. nov. (= TBRC 10620T = NBRC 114213T) is proposed. Furthermore, genome analysis of both strains suggested that members of the genus Fodinicola are promising sources of beneficial PGP-actinomycetes and novel secondary metabolites.A novel endophytic actinomycete strain GKU 173 Endophytic actinomycetes colonize the internal plant tissues and usually have a beneficial effect to the host plant by promoting growth and protecting the plant from biotic and abiotic stresses without any damage or morphological changes to the plant ,2. IsolaFodinicola was first proposed by Carlsohn et al.  was determined on ISP 3 agar for three weeks. Catalase and oxidase activities were observed with 3% (v/v) hydrogen peroxide solution and 1% (v/v) tetramethyl-p-phenylenediamine solution [Growth of strain GKU 173solution , respectsolution . The utisolution . Degradasolution  after insolution ,38. Hydrsolution . Milk coT in Bennett\u2019s broth on a rotary sharker at 28 \u00b0C for 14 days. Cells were harvested by centrifugation, washed with distilled water, and freeze-dried. Cell wall peptidoglycan was prepared using the modified method described by Kawamoto et al. [N\u03b1--D-leucinamide (FDLA) derivatization according to the method of Fujii et al. [N-acyl group of muramic acid in the peptidoglycan was examined using the previously described method [For chemotaxonomic analysis, biomass was prepared by growing strain GKU 173o et al.  to analyo et al.  after N\u03b1i et al. . The LC/i et al.  and by Li et al. . The isoi et al. . N-acyl d method . Mycolicd method . Phosphod method . Cellulad method  by the Sd method , and anaT and F. feengrottensis HKI 0501T were extracted from the culture grown in Bennett\u2019s broth at 28 \u00b0C for seven days according to standard procedure [T and F. feengrottensis HKI 0501T were sequenced using Illumina Hiseq PE 150 serviced by Novogene  and PacBio Sequel systems at Chulabhorn Royal Academy, Thailand. Prior to assembly, the reads were trimmed for adapter sequences and filtered for sequence quality and checked by Fast QC tool [Genomic DNA of strain GKU 173rocedure . Whole g QC tool . Sequenc QC tool  and dete QC tool . G+C conAverage Nucleotide Identity (ANI) and Ortho ANI  were comT was isolated from surface-sterilized roots of the black wattle tree (Acacia mangium Willd.) on starch-casein agar. It is an aerobic Gram-stain-positive non-sporulating actinobacterium. Morphological observation on ON agar indicated that strain GKU 173T formed a branched substrate mycelium and abundant white aerial hyphae that fragmented into irregular rod-like elements  tricalcium phosphate . The strain also showed a remote relationship (<95% similarity) with other species including Cryptosporangium eucalypti EURKPP3H10T (94.64%), Cryptosporangium phraense A-T 5661T (94.57%), Cryptosporangium aurantiacium DSM 46144T (94.50%), Jiangella anatolica GTF31T (94.36%), Cryptosporangium minutisporangium IFO 15962T (94.36%), and Frankia coriariae BMG5.1T (94.34%). Neighbor-joining phylogenetic analysis indicated that strain GKU 173T grouped tightly with the only member of the genus Fodinicola  indicated that strain GKU 173dinicola . The posp values . Genus Fn et al.  is monopngiaceae  within t0501T 97.%. The st in 2008 . Based oT and its closely related species, F. feengrottensis HKI 0501T, were phenotypically characterized. Strain GKU 173T showed better growth than F. feengrottensis HKI 0501T in most tested media  with good growth at concentration up to 3% (w/v). Strain GKU 173T showed oxidase-negative and catalase-positive. The difference of acidic production, and carbon/nitrogen utilization of strain GKU 173T and F. feengrottensis HKI 0501T are summarized in T can be distinguished from F. feengrottensis HKI 0501T.Strain GKU 173ed media . Its sub pigment . Strain T contained meso-diaminopimelic acid, D-alanin, D-glutamic acid, and glycine. Cell wall sugars were xylose and mannose. The N-acyl type of muramic acid in the peptidoglycan was N-acetyl. Mycolic acids were absent. The polar phospholipids comprised of diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), phosphatidylinositol (PI), several unknown phospholipids, and unknown ninhydrin-positive compounds (16:0 (58.22%), anteiso-C17:0 (9.94%), iso-C18:0 (9.35%), and iso-C17:0 (6.58%); and a small proportion of 10-methyl C17:0 (3.87%), 10-methyl C18:0 (1.9%), iso-C16:1 (1.63%), sum in feature 8 , and summed feature 8 . The menaquinones were MK-9(H4) (25%), MK-9(H6) (36%) and MK-9(H8) (39%). All chemotaxonomic results revealed that strain GKU 173T shared apparent characteristics with the only member of the genus Fodinicola particularly, the presence of xylose as the diagnostic cell-wall sugar [T as a novel species within the genus Fodinicola.The analysis of the cell wall peptidoglycan by TLC  and LC/Mompounds . The fatll sugar . AltogetT and strain HKI 0501T were sequenced and compared. The assembled draft genomes of strain GKU 173T and strain HKI 0501T contained 4 and 107 contigs with a total length of 8.86 Mbp (GenBank accession number WOTN00000000) and 8.71 Mbp (GenBank accession number WOTO00000000), respectively. The N50 value of genomes of strains GKU 173T and HKI 0501T was 2.35 Mbps (L50 = 2) and 1.15 Mbps (L50 = 26), respectively. The ANI value between strain GKU 173T and strain HKI 0501T was 77.92% which was much lower than the ANI threshold range (95%\u201396%) for species delineation [T and F. feengrottensis HKI 0501T were 67.81% and 66.61%, respectively. The difference in G+C value was higher than 1% indicating distinct species according to Meier-Kolthoff et al. [T was distinguished from the closely related type strain and represents a novel species of the genus Fodinicola, for which the name Fodinicola acaciae sp. nov. (= TBRC 10620T = NBRC 114213T) is proposed.Genomes of strain GKU 173ineation . The dDDineation . The calf et al. . On the Fodinicola acaciae myo-inositol, lactose, mannitol, D-raffinose, D-sorbitol, sucrose, D-tretalose, and D-xylose, but not from rhamnose. L-tyrosine, Tween 20, and Tween 80 are degraded, but adenine, hypoxanthine, starch, urea, xanthine, and xylan are not. Adonitol, L-arabinose, D-fructose, D-glucose, glycerol, myo-inositol, lactose (weakly), mannitol, D-raffinose, rhamnose, D-sorbitol, sucrose, D-trehalose, and D-xylose are used as carbon sources. L-arginine, L-cysteine, L-histidine, L-isoleucine, L-phenylalanine, potassium nitrate, L-tryptophan, and L-valine are used as nitrogen sources. By API ZYM system, production of acid phosphatase, alkaline phosphatase, \u03b1-chymotrypsin (weakly), cysteine arylamidase (weakly), esterase (C4), \u03b1-galactosidase, \u03b2-galactosidase, N-acetyl-\u03b2-glucosaminidase, \u03b1-glucosidase, \u03b2-glucosidase, leucine arylamidase, lipase (C8), lipase (C14), \u03b1-mannosidase, naphthol-AS-BI-phosphohydrolase, and trypsine are positive, but not \u03b1-fucosidase and \u03b2-glucuronidase. The cell wall peptidoglycan contains meso-A2pm, D-alanine, D-glutamic acid, and glycine. The muramic acid in peptidoglycan is N-acetylated. The cell-wall sugars are xylose and mannose. The predominant menaquinones are MK-9(H4), MK-9(H6), and MK-9(H8). The polar phospholipids comprise diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), phosphatidylinositol (PI), several unknown phospholipids, and unknown ninhydrine-positive compounds. Mycolic acids are absent. The predominant cellular fatty acid profiles contain iso-C16:0, anteiso-C17:0, iso-C18:0, and iso-C17:0. The G+C content is 67.81% (determined from the genome sequence).Gram-stain-positive, aerobic, non-motile, catalase-positive, oxidase-negative actinomycete that produces branched substrate mycelium and abundant aerial mycelium. Aerial hyphae break up into irregular rod-like elements. Colonies are wrinkled, beige to orange in color. Red-series diffusible pigment is produced on Bennett, ISP 2, ISP 3, organic medium 79, and MS media. Growth occurs between 14 and 42 \u00b0C, good growth at 28 \u00b0C, and no growth below 14 \u00b0C or above 42 \u00b0C. Good growth occurs at pH 7.0 and pH 8.0, but no growth occurs at pH 3 nor pH 12. NaCl tolerance is up to 6% (w/v). Nitrate is not reduced. Gelatin liquefaction, milk coagulation and peptonization are positive. Acid production from adonitol, L-arabinose, D-fructose, glycerol, D-glucose, T (= TBRC 10620T = NBRC 114213T) was isolated from the roots of black wattle tree (Acacia mangium Willd.) collected at Kasetsart University, Bangkok, Thailand. Type strain GKU 173T showed 8705 coding sequences and 55 RNAs, while F. feengrottensis HKI 0501T showed 11,978 coding sequences and 57 RNAs. The genome annotation revealed genes related to the PGP traits according to those in vitro activities of strain GKU 173T and HKI 0501T  of lucA/lucC family protein [RAST annotation of genome sequences of strain GKU 173KI 0501T . Indole-lubility . The gen protein .T and F. feengrottensis HKI 0501T were investigated by antiSMASH. The results showed that strain GKU 173T carried 16 BGCs including 5 known BGCs and 11 uncharacterized BGCs  constitutes a novel species within the genus Fodinicola of which Fodinicola acaciae sp. nov. was proposed under the type strain GKU 173T (= TBRC 10620T = NBRC 114213T). Genome analysis of the novel strain and the closely related strain, F. feengrottensis HKI 0501T, revealed potential PGP-traits including genes involved in phosphate solubilization, IAA and siderophore production; and a variety of uncharacterized BGCs comprising NRPs, RiPPs, Type I PKs, Type II PKs, Type I PK-NRP hybrid, and terpenes. The results suggested that members in the genus Fodinicola have potential activity as beneficial PGP-bacteria and specialized secondary metabolite producers. Based on the polyphasic taxonomic study demonstrated that strain GKU 173"}
+{"text": "Their absorption and emission behaviours in dilute solutions were investigated in order to explain structure\u2013property associations and demonstrate the impact of different aryl substituents on the terpyridine scaffold as well as the role of the metal on the complexes. Photo-luminescence analysis of the complexes in acetonitrile solution revealed a transition from hypsochromic to bathochromic shift. All the compounds displayed remarkable photo-luminescent properties and various maximum emission peaks owing to the different nature of the functional groups. Furthermore, the anti-microbial potential of ligands and complexes was evaluated with docking analyses carried out to investigate the binding affinity of terpyridine-based ligands along with corresponding proteins (shikimate dehydrogenase and penicillin-binding protein) binding sites. To obtain further insight into molecular orbital distributions and spectroscopic properties, density functional theory calculations were performed for representative complexes. The photophysical activity and interactions between chromophore structure and properties were both investigated experimentally as well as theoretically.A series of different substituted terpyridine (tpy)-based ligands have been synthesized by Kr\u00f6hnke method. Their binding behaviour was evaluated by complexing them with Co(II), Fe(II) and Zn(II) ions, which resulted in interesting coordination compounds with formulae, [Zn(tpy) Thorough photophysical and computational studies of the new complexes provide an insight into their electronic structure, which provides a significant extension of newly published terpyridine-transition metal complexes. In addition, we have established a detailed structure\u2013activity relationship (SAR) between these architectures and their biological and spectrophotometric properties.2.3) and acetonitrile (CH3CN) solutions, respectively, on a Jasco UV-Vis V-660 instrument using a QUARTZ cell. The luminescence spectra were obtained using a Shimadzu 8101AFT-IR instrument. Thin-layer chromatography (TLC) was conducted on silica gel TLC plates purchased from Merck and chromatograms were viewed with a UV-lamp at 254 and 365 nm.All chemicals were bought from Merck and Sigma-Aldrich and used as received. Melting points have been recorded with an electrothermal device and are uncorrected. The FTIR spectra have been obtained on a Bio-Rad spectrophotometer and the infrared values are listed in \u1fe1 units. The UV-Vis spectra were recorded in chloroform  in methanol (20 ml), a substituted aryl aldehyde (10.0 mmol) was added, followed by potassium hydroxide (KOH) pellets  and 35% aqueous ammonia solution (40.0 ml). The reaction mixture was refluxed for 4\u20136 h. After the reaction was completed (determined by TLC monitoring), the solvent was removed under vacuum and the precipitates were filtered, washed with plenty of distilled H2.1.2.2Cl2 solution (20.0 ml) of the substituted terpyridine ligand  under continuous stirring. The colour of the reaction mixture instantly changed and the reaction mixture was stirred for approximately 2 h at room temperature. Upon addition of an excess of NH4PF6 precipitation occurred. The precipitates were filtered, washed with ice-cold MeOH (5.0 ml) and (C2H5)2O (15.0 ml) to obtain a pure complex. Recrystallization from acetonitrile or methanol or a mixture of both yielded the analytically purified complex.A hot methanolic solution (20.0 ml) of metal salt (0.5 mmol) was introduced dropwise to a CH2.2.1\u2013L9L) and 520 nm for complexes (1\u2013C27C).RF-6000 Spectrofluorophotometer was used to produce an emission spectrum for ligands and their complexes. The emission spectra of the synthesized ligand in chloroform and of complexes in acetonitrile were obtained by using a fluorescence spectrophotometer. The emission spectra were checked by keeping the excitation value of the synthesized compounds at 325 nm ,67\u201372. T2.3.2.3.1.http://www.rcsb.org/pdb/home/home.do] [Chemical structures of the terpyridines were developed with ChemBio Draw which also produced their MOL format files. The shikimate dehydrogenase (PDB ID: 3DON) and penicillin-binding protein (PDB ID: 1VQQ) three-dimensional crystal structures have been accessed and downloaded from the protein data bank database (PDB) [home.do] .2.3.2.The active sites of the target protein were examined using the molecular operating environment (MOE) software. An active site was distinct from the ligand coordinates in the original protein sites ,72\u201375.2.3.3.A theoretical ligand-target docking method has been used to classify structural complexes of the shikimate dehydrogenase crystal structure and penicillin-target protein with ligand molecules to recognize the molecular basis of the specificity of these protein targets. Eventually, the MOE program performs docking. The energy of these derivatives to interact with the protein targets is given \u2018grid level\u2019.2.4.13C were optimized using the B3LYP* hybrid functional and double zeta (DZ) basis sets.Computational calculations were performed by using DFT-B3LYP*(DZ) in the Amsterdam Density Functional (ADF) modelling suite . The gro2.5.2.5.1.in vitro anti-bacterial activity against Gram-negative Escherichia coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 9721) and Gram-positive Staphylococcus aureus (ATCC 6538) bacterial strains. Cefixime was used as a positive standard and dimethyl sulfoxide (DMSO) as a negative control. Various sample solutions were prepared by dissolving 4.0 mg of each sample in 1.0 ml of DMSO. The lawn was developed on nutrient agar plates using bacterial strains of equal turbidity that is accomplished using 0.5% of McFarland's solution. The well depth was 8 mm and they were all made at the correct distances from each other. The sample and standard are poured into their respective tubes. The sample quantity used was 80 \u00b5l in a well together with the two controls. The prepared dishes were incubated for 24 h at 37\u00b0C, and the findings were reported as the average diameter of zone of inhibition (ZOI) of bacterial development around the discs in millimetres for each compound +\u00b7 : 387.0713.Light-purple amorphous solid; Yield: 85%; m.p. above 300\u00b0C; UV 3.12.\u03bbmax (CH3CN) = 314 nm; \u03bbem (CH3CN): 379, 776 nm; FTIR (cm\u22121): 1616 (C=N), 1471 (C=C), 1245 (C-N), 1110 (C-H), 623 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 705.2176.Brick-red amorphous solid; Yield: 85%; m.p. above 300\u00b0C; UV 3.13.\u03bbmax (CH3CN) = 327 nm; \u03bbem (CH3CN): 394, 742 nm; FTIR (cm\u22121): 1611 (C=N), 1483 (C=C), 1286 (C-N), 1058 (C-H), 627 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 702.2194.Dark-purple amorphous solid; Yield: 79%; m.p. above 300\u00b0C; UV 3.14.\u03bbmax (CH3CN) = 243 nm; \u03bbem (CH3CN): 370, 521, 732 nm; FTIR (cm\u22121): 1595 (C=N), 1476 (C=C), 1250 (C-N), 1107 (C-H), 584 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 416.0979.Brown amorphous solid; Yield: 72%; m.p. above 300\u00b0C; UV 3.15.\u03bbmax (CH3CN) = 290 nm; \u03bbem (CH3CN): 547, 683 nm; FTIR (cm\u22121): 1595 (C=N), 1473 (C=C), 1248 (C-N), 1125 (C-H), 625 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 763.2707.Candy-orange amorphous solid; Yield: 76%; m.p. above 300\u00b0C; UV 3.16.\u03bbmax (CH3CN) = 324 nm; \u03bbem (CH3CN): 365, 638, 751 nm; FTIR (cm\u22121): 1587 (C=N), 1468 (C=C), 1247 (C-N), 1090 (C-H), 629 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 760.2725.Dark-purple amorphous solid; Yield: 77%; m.p. above 300\u00b0C; UV 3.17.\u03bbmax (CH3CN) = 428 nm; \u03bbem (CH3CN): 294, 359, 451, 717 nm; FTIR (cm\u22121): 1660 (C=N), 1487 (C=C), 1286 (C-N), 1105 (C-H), 589 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 540.1292.Brown amorphous solid; Yield: 80%; m.p. above 300\u00b0C; UV 3.18.\u03bbmax (CH3CN) = 308 nm; \u03bbem (CH3CN): 273, 453, 722 nm; FTIR (cm\u22121): 1584 (C=N), 1506 (C=C), 1284 (C-N), 1050 (C-H), 621 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 1011.3310.Marmalade (dark-yellow) amorphous solid; Yield: 70%; m.p. above 300\u00b0C; UV 3.19.\u03bbmax (CH3CN) = 437 nm; \u03bbem (CH3CN): 341, 747 nm; FTIR (cm\u22121): 1582 (C=N), 1505 (C=C), 1285 (C-N), 1150 (C-H), 627 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 1008.3351.Maroon amorphous solid; Yield: 66%; m.p. above 300\u00b0C; UV 3.20.\u03bbmax (CH3CN) = 286 nm; \u03bbem (CH3CN): 454, 856 nm; FTIR (cm\u22121): 1600 (C=N), 1477 (C=C), 1295 (C-N), 1110 (C-H), 581 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 403.0663.Greyish-purple amorphous solid; Yield: 78%; m.p. above 300\u00b0C; UV 3.21.\u03bbmax (CH3CN) = 286 nm; \u03bbem (CH3CN): 426, 704, 856 nm; FTIR (cm\u22121): 1610 (C=N), 1473 (C=C), 1301 (C-N), 1098 (C-H), 625 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 737.2075.Clay-brown amorphous solid; Yield: 66%; m.p. above 300\u00b0C; UV 3.22.\u22121; Yield: 58%; m.p. above 300\u00b0C; UV \u03bbmax (CH3CN) = 336 nm; \u03bbem (CH3CN): 409, 503, 759, 854 nm; FTIR (cm\u22121): 3345 (C=N), 1607 (C=C), 1432 (C-N), 1057 (C-H), 628 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 734.2092.Dark-purple amorphous solid; Mol. Wt.: 1024.55 g mol3.23.\u03bbmax (CH3CN) = 287 nm; \u03bbem (CH3CN): 853 nm; FTIR (cm\u22121): 1616 (C=N), 1443 (C=C), 1326 (C-N), 1298 (C-H), 583 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 441.0431.Red-violet amorphous solid; Yield: 73%; m.p. above 300\u00b0C; UV 3.24.\u03bbmax (CH3CN) = 285 nm; \u03bbem (CH3CN): 405, 737, 850 nm; FTIR (cm\u22121): 1617 (C=N), 1473 (C=C), 1327 (C-N), 1250 (C-H) 627 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 813.1611.Brick-red amorphous solid; Yield: 55%; m.p. above 300\u00b0C; UV 3.25.\u03bbmax (CH3CN) = 286 nm; \u03bbem (CH3CN): 403, 685, 850 nm; FTIR (cm\u22121): 1612 (C=N), 1448 (C=C), 1327 (C-N), 1128 (C-H), 630 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 810.1629.Dark-purple amorphous solid; Yield: 70%; m.p. above 300\u00b0C; UV 3.26.\u03bbmax (CH3CN) = 283 nm; \u03bbem (CH3CN): 401, 685, 851 nm; FTIR (cm\u22121): 1617 (C=N), 1447 (C=C), 1327 (C-N), 1284 (C-H), 590 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 441.0431.Lilac (light purple) amorphous solid; Yield: 65%; m.p. above 300\u00b0C; UV 3.27.\u03bbmax (CH3CN) = 285 nm; \u03bbem (CH3CN): 361, 735, 859 nm; FTIR (cm\u22121): 1615 (C=N), 1476 (C=C), 1328 (C-N), 1197 (C-H), 627 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 813.1611.Brick-red amorphous solid; Yield: 78%; m.p. above 300\u00b0C; UV 3.28.\u03bbmax (CH3CN) = 286 nm; \u03bbem (CH3CN): 361, 580, 737, 865 nm; FTIR (cm\u22121): 1612 (C=N), 1450 (C=C), 1328 (C-N), 1265 (C-H), 629 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 810.1629.Dark-purple amorphous solid; Yield: 71%; m.p. above 300\u00b0C; UV 3.29.\u03bbmax (CH3CN) = 286 nm; \u03bbem (CH3CN): 409, 856 nm; FTIR (cm\u22121): 1619 (C=N), 1448 (C=C), 1329 (C-N), 1198 (C-H), 592 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 441.0431.Light-purple amorphous solid; Yield: 70%; m.p. above 300\u00b0C; UV 3.30.\u03bbmax (CH3CN) = 289 nm; \u03bbem (CH3CN): 360, 730, 867 nm; FTIR (cm\u22121): 1617 (C=N), 1480 (C=C), 1332 (C-N), 1267 (C-H), 623 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 813.1611.Chocolate-brown amorphous solid; Yield: 65%; m.p. above 300\u00b0C; UV 3.31.\u03bbmax (CH3CN) = 286 nm; \u03bbem (CH3CN): 358, 578, 732, 869 nm; FTIR (cm\u22121): 1615 (C=N), 1457 (C=C), 1334 (C-N), 1210 (C-H), 626 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 810.1629.Dark-purple amorphous solid; Yield: 66%; m.p. above 300\u00b0C; UV 3.32.\u03bbmax (CH3CN) = 347 nm; \u03bbem (CH3CN): 365, 680, 743, 855; FTIR (cm\u22121): 1603 (C=N), 1467 (C=C), 1348 (C-N), 1073 (C-H), 585 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 310.040.Light-purple amorphous solid; Yield: 74%; m.p. above 300\u00b0C; UV 3.33.\u03bbmax (CH3CN) = 334 nm; \u03bbem (CH3CN): 391, 699 nm; FTIR (cm\u22121): 1614 (C = N), 1530 (C=C), 1246 (C-N), 1151 (C-H), 627 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 767.1565.Chocolate-brown amorphous solid; Yield: 65%; m.p. above 300\u00b0C; UV 3.34.\u03bbmax (CH3CN) = 328 nm; \u03bbem (CH3CN): 402, 684, 856 nm; FTIR (cm\u22121): 1614 (C=N), 1474 (C=C), 1351 (C-N), 1025 (C-H), 629 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 764.1584.Dark-purple amorphous solid; Yield: 70%; m.p. above 300\u00b0C; UV 3.35.\u03bbmax (CH3CN) = 287 nm; \u03bbem (CH3CN): 474, 850 nm; FTIR (cm\u22121): 1619 (C=N), 1476 (C=C), 1359 (C-N), 1167 (C-H), 587 (Zn-N); MS (MALDI-ToF) of [M]+\u00b7 : 310.040.Caramel (brown) amorphous solid; Yield: 77%; m.p. above 300\u00b0C; UV 3.36.\u03bbmax (CH3CN) = 289 nm; \u03bbem (CH3CN): 428, 563, 850 nm; FTIR (cm\u22121): 1616 (C=N), 1471 (C=C), 1245 (C-N), 1245 (C-H), 628 (Co-N); MS (MALDI-ToF) of [M]+\u00b7 : 767.1565.Chocolate-brown amorphous solid; Yield: 76%; m.p. above 300\u00b0C; UV 3.37.\u03bbmax (CH3CN) = 287 nm; \u03bbem (CH3CN): 565, 848 nm; FTIR (cm\u22121): 1618 (C=N), 1472 (C=C), 1350 (C-N), 1287 (C-H), 630 (Fe-N); MS (MALDI-ToF) of [M]+\u00b7 : 764.1598.Dark-purple amorphous solid; Yield: 83%; m.p. above 300\u00b0C; UV 3.38.1\u2013L9L), strong, broad absorption bands appear in the range 234\u2013325 nm, while for the complexes (1\u2013C27C), the absorption bands were seen in the range 240\u2013429 nm. All these absorptions are attributed to \u03c0\u2013\u03c0* electronic transitions in the aromatic system of the terpyridines. The absorption maxima for 1\u2013C27C are somewhat shifted from those of the free ligands by affixing either electron-releasing or -withdrawing groups at the central pyridine. The longest absorption wavelength in the 1L UV-Vis spectrum (\u03bbmax = 325 nm) is broad and comparable to that of the un-substituted parent terpyridine  at \u03bbmax = 587 nm and the bands at 284 nm and 426 nm are due to ligand-centred (LC) transitions. A slight redshift when related to the reference iron (II) terpyridine complex has been detected. The spin allowed the MLCT band in the visible region to undergo a rise in intensity and a redshift, regardless of electron-donor or -acceptor nature of the substituents. The intense absorption bands of the Zn(II) complexes  were seen in the range 215\u2013586 nm. These absorption peaks are mainly due to the metal\u2013ligand charge transfer of \u03c0\u2013\u03c0* and n\u2013\u03c0* transitions having \u03bbmax = 586 nm. This redshift could be credited to a decrease in the LUMO energy values of the complexes. The absorption bands of the Co(II) complexes  were seen in the range 209\u2013571 nm and their MLCT transitions have a maximum value at 530 nm.With a broad class of terpyridine compounds in hand, we examined their initial photophysical properties. The UV-Vis spectra of terpyridines and their metal complexes are recorded in chloroform and acetonitrile solutions, respectively, and the related data are given in pyridine . The UV-p-methoxyphenyl at the 4\u2032-position of the terpyridine ring in Zn(II) and Co(II) complexes shifts \u03bbmax towards longer wavelengths, resulting in a 30 nm bathochromic shift. This result supports the concept that the MLCT state can be higher in energy by nitration of the terpyridine ligand-based metal complexes, the 4-methoxyphenyl group at the 4\u2032-position of the terpyridine ring in the complexes have a shorter wavelength, and a significant maximum enhancement in MLCT band energy was observed. The absorption bands are influenced significantly by the aryl substituents on the terminal phenyl moiety. Generally, the absorption spectra of all complexes are typically well correlated with those found for the respective uncomplexed ligands.Surprisingly, the electron-donating group 3.39.p-position may not only open several paths towards the development of advanced supramolecular structures but may also influence the fluorescence properties of the compounds. Due to their potential applications in chemical sensors, electroluminescent displays and fluorescent materials, inorganic-organic hybrid complexes, particularly those with d10 transition metal centres, are now of great interest. From the structural analysis, the ligand can be seen to have a large p-conjugated aromatic system that can possess strong fluorescence and efficient transfer of energy. The photo-luminescence spectra of the Zn, Co and Fe (1\u2013C27C) complexes were recorded in acetonitrile solutions at ambient temperature with an excitation \u03bb of 320 nm for the Zn(II) complexes, 286 nm for the Co complexes and 295 nm for the Fe complexes. All 1\u2013C27C complexes display emission bands in the visible region of the electromagnetic spectrum. The maximum emission peaks for the Zn, Co and Fe complexes were observed at 857, 859 and 865 nm, respectively, as described in 1\u2013L9L was observed at about 340 nm irrespective of the location of the substitution and the number of phenyl groups. The excitation spectrum observed between 330 and 400 nm at any stage was similar to the corresponding absorption spectra, indicating that the excitation of the lowest energy absorption of 1\u2013L9L led to the emitting state. In the p-position of the 4-phenyl complex, several substituents with different electron-donating or -withdrawing properties were added to further analyse and modify the fluorescence properties of 4-phenyl terpyridine. The p-substituted terpyridine complexes 1\u2013C27C absorption and fluorescence maxima are listed in Attaching various functional groups to terpyridine at the 1\u2013C4C in solution and at ambient temperature are characterized by the fluorescence spectra presented in figures\u00a01C has two bands in its emission spectrum, a lower intensity one at ca 405 nm and another with higher intensity at ca 704 nm. Similarly, compound 2C has two bands in its emission spectra, a low-intensity one at ca 379 nm and another with higher intensity at ca 776 nm. Compared with the previously reported 2+ complexes  have revealed that introduction of either electron-donating 2, -N(Ph)2, -OCH3) or electron-withdrawing  groups onto the aryl-terpyridine scaffold alters absorbance as well as fluorescence properties and thus resulted in bathochromic shifted and structurally varied absorption and emission spectra. In addition, we have observed that the introduction of d-block transition metals  into the terpyridine derivatives modified their absorption and emission properties as compared with terpyridine ligands (1\u2013L9L), and most complexes (1\u2013C27C) showed red-shifted absorption and emission spectra of varied appearance in contrast with the 2,2\u2032:6\u2032\u2032,2\u2032-terpyridines. Many of the new compounds have interesting photo-luminescent properties with high emissions. Importantly, almost all the functionalized terpyridine complexes display longer emission \u03bb and they are stronger emissive in contrast with the unsubstituted 2,2\u2032:6\u2032,2\u2033-terpyridine.We have presented the spectroscopic, photophysical, and anti-microbial studies of Zn(II), Co(II) and Fe(II) complexes of the variously substituted 2,2\u2032:6\u2032,2\u2032\u2032-terpyridine motif. New substrates of symmetrically substituted in vitro). The analysis revealed that the complexes (1\u2013C27C) exhibited more potent activity than free ligands (1\u2013L9L). Overall, the entire compounds exhibited moderate to excellent anti-microbial activities. These results provide new possibilities for therapeutic purposes. The experimental results complemented with in silico studies provided insights into structure\u2013property relationships. We are quite confident that the reported structures may find applications in organic electronics as well as medicines as potential materials and this will be the target of our future investigations. The investigation permitted the selection of materials with the most promising properties with special emphasis on the nature of the substituents.The ligands and complexes were also evaluated for their anti-microbial potential ("}
+{"text": "Bothrops brazili, is a poorlystudied pit viper distributed in lowlands of the equatorial rainforests ofsouthern Colombia, northeastern Peru, eastern Ecuador, southern andsoutheastern Venezuela, Guyana, Suriname, French Guiana, Brazil, andnorthern Bolivia. Few studies have been reported on toxins isolated fromvenom of Ecuadorian and Brazilian B. brazili. The aim ofthe present study was to elucidate the qualitative and quantitative proteincomposition of B. brazili venom from Par\u00e1 (Brazil), and tocarry out a comparative antivenomics assessment of the immunoreactivity ofthe Brazilian antibothropic pentavalent antivenom [soroantibotr\u00f3pico (SAB) in Portuguese] against the venoms ofB. brazili and reference species, B.jararaca.The Brazil\u2019s lancehead, B.brazili venom from Par\u00e1 (Brazil). Using third-generationantivenomics, the specific and paraspecific immunoreactivity of theBrazilian SAB against homologous (B. jararaca) andheterologous (B. brazili) venoms was investigated. We have applied a quantitative snake venomics approach, includingreverse-phase and two-dimensional electrophoretic decomplexation of thevenom toxin arsenal, LC-ESI-MS mass profiling and peptide-centric MS/MSproteomic analysis, to unveil the overall protein composition of 2 molecules; 7-11 snake venommetalloproteinases of classes PI (21%) and PIII (6%); 10-12 serineproteinases (14%), and 1-2 L-amino acid oxidases (6%). Other toxins,including two cysteine-rich secretory proteins, one C-type lectin-likemolecule, one nerve growth factor, one 5'-nucleotidase, onephosphodiesterase, one phospholipase B, and one glutaminyl cyclase molecule,represent together less than 2.7% of the venom proteome. Third generationantivenomics profile of the Brazilian pentabothropic antivenom showedparaspecific immunoreactivity against all the toxin classes of B.brazili venom, with maximal binding capacity of132.2 mg venom/g antivenom. This figure indicates that 19% of antivenom'sF(ab')2 antibodies bind B. brazili venomtoxins. The venom proteome of the Brazil\u2019s lancehead (Par\u00e1) is predominantly composedof two major and three minor acidic (19%) and two major and five minor basic(14%) phospholipase AB.brazili and B. jararaca (the reference venomfor assessing the bothropic antivenom's potency in Brazil), provides cluesabout the proper use of the Brazilian antibothropic polyvalent antivenom inthe treatment of bites by the Brazil\u2019s lancehead. The proteomics outcome contribute to a deeper insight into the spectrum oftoxins present in the venom of the Brazil\u2019s lancehead, and rationalize thepathophysiology underlying this snake bite envenomings. The comparativequalitative and quantitative immunorecognition profile of the Brazilianpentabothropic antivenom toward the venom toxins of  Bothrops includes at least 50 species of pit vipers that are widely distributed throughout the Americas, fromMexico to southern Argentina, in different ecoregions, from tropical and subtropicalforests to arid and semiarid regions, and from sea level to altitudes of more than3000 m  neutralizes 50 mg of B.jararaca venom (the reference venom for assessing the bothropicantivenom potency in Brazil).Pooled venom from olecules , 33. A vg for 10 min at room temperature, and the proteinscontained in 40\u00b5L (600 \u00b5g) were separated by RP-HPLC using a Agilent LC 1100High Pressure Gradient System equipped with a Teknokroma Europa C18  column and a DAD detector. The columnwas developed at a flow rate of 1.0 mL/min with a linear gradient of 0.1% TFA inMilliQ\u00ae water (solution A) and 0.1% TFA in acetonitrile (solutionB), isocratic (5% B) for 5 min, followed by 5-25% B for 10 min, 25-45% B for 60min, and 45-70% B for 10 min. Protein detection was carried out at 215 nm with areference wavelength of 400 nm. Fractions were collected manually across theentire elution range, dried in a vacuum centrifuge , and redissolved in MilliQ\u00ae water. Molecular masses ofthe purified proteins were estimated by non-reduced and reduced Tris-TricineSDS-PAGE (on 15% polyacrylamide gels) , with orwithout 40 mM DTT. IPG strips were then placed on top of SDS-15% polyacrylamidegels and run in a Protean II (Bio-Rad) electrophoresis unit at room temperature.Protein spots were visualized by Coomassie Brilliant Blue G250 staining. \u00ae,ThermoSavant), redissolved in 14\u00b5L of 5% ACN containing 0.1% formic acid, and7\u00b5L submitted to LC-MS/MS. Tryptic peptides were separated by nano-AcquityUltraPerformance LC\u00ae (UPLC\u00ae) as above. Protein bands of interest were excised from Coomassie Brilliant Blue-stainedSDS-PAGE and 2-DE gels and subject to in-gel disulfide bond reduction  and cysteine alkylation , followed by overnight digestion withsequencing-grade trypsin , using a Genomics Solution ProGest\u2122 Protein Digestion Workstation.Tryptic digests were dried in a vacuum centrifuge (SPD SpeedVacde novo sequencing), ii) usingthe on-line form of the MASCOT Server (version 2.6) at http://www.matrixscience.com against the last update  of NCBI non-redundant database, and iii) processed inWaters Corporation\u2019s ProteinLynx Global SERVER 2013 version 2.5.2. (withExpression version 2.0). The following search parameters were used: Taxonomy:bony vertebrates; Enzyme: trypsin ; MS/MS masstolerance was set to \u00b1 0.6 Da; carbamidomethyl cysteine and oxidation ofmethionine were selected as fixed and variable modifications, respectively. Allmatched MS/MS data were manually checked. Peptide sequences assigned byde novo MS/MS were matched to homologous proteins availablein the NCBI non-redundant protein sequences database using the online BLASTPprogram was determined spectrophotometrically using an extinction coefficient for a 1mg/mL concentration (\u03b50.1%) at 280 nm of 1.36 (mg/mL)-1cm-1  by the [total amount of antibody(F(ab')2) (in mg) per antivenom vial] [The percentage of antivenom anti-toxin F(ab')om vial] , 45, 46.B. brazili (Par\u00e1)was decomplexed and quantified by reverse-phase HPLC and downstream SDS-PAGEanalysis of the chromatographic peaks (B. brazili (Par\u00e1) venom investigated inthis work, RP-HPLC fractions 2-46 were submitted to molecular mass determinationby LC-ESI-MS mass profiling. The venom proteome of 600 \u00b5g of crude venom of ic peaks . Twenty ic peaks , inset. 50 in the range of 0.15-0.95 mM) of the fibrinogenolyticactivity of multiple snake venom Zn2+-metalloproteinases (SVMP) of 27,623.1, 27,455.2 and 13,930.5; 27,636.5 and 13,803.2; and30,360.4, 29,663.4 and 13,781.9, respectively. These molecular masses maycorrespond to the minor (<0 .1% TVC) PLA13.1 Da) ; MTx-II 13.1 Da) ; Brazili, Brazil  or SerpeB.atrox , 48, andave: 29,899.2 Da, 30,130.2 Da and 30,421.9 Da), 19, 20 , 38 and 42/43  were ten2s, which together account for approximately 30% (w/w) of itsproteome, one minor (1.6%) CRISP molecule, one major  and at leastten minor (< 2.3%) SVSPs, and 2-3 abundant  and amajor  PI-SVMPs. In addition, SDS-PAGE analysis displayed inAs a whole, the above data suggested that the Brazil\u2019s lancehead venom comprisednine minor  and five major (> 4.4%)PLAdenovo gathered peptide ion sequences  was fractionated by RP-HPLC/SDS-PAGE and 2-DEB. brazili (Par\u00e1), comprisedby at least 40-47 components  and PIII , 10-12 SVSPs (14%) and 1-2 LAOs (6%). Other toxinclasses are: two CRISPs, one C-type lectin-like (CTL), one nerve growth factor(NGF), one 5'-nucleotidase (5'NT), one phosphodiesterase (PDE), onephospholipase B (PLB), and one glutaminyl cyclase (GC) represent together lessthan 2.7% of the venom proteome (50) and hemorrhagic, dermonecrotic and defibrinogenatingeffects reported for Peruvian B. brazili venom in the murinemodel  andMTx-II   , 54, andliase-II .ClearlyTwo-dimensional electrophoretic (2-DE) analysis provides a rapid way to visualizethe overall venom protein complexity of a snake's venom in a single image. 2-DEand RP-HPLC/SDS-PAGE are complementary approaches that combined provide a morecomprehensive view of a venom proteome than each approach separately. Inaddition, each of these approaches serves, by itself, a specific purpose. Thus,the presence and subunit composition of covalent complexes in a venom proteomecan be conveniently addressed by comparing the 2-DE protein maps resolved undernon-reducing (NRed) conditions in both directions (IEF and SDS-PAGE) versusnon-reducing/reducing (Red) conditions . 2 subproteome, which is made of two major acidic , two strongly basic (pI 9.5-9.8) (spots 13 and 14), and onemildly basic (pI 7.8) (spot 12), and five low abundant PLA2 molecules(spots 3-7) within the pI range 5.3-7.3. The latter spots also yielded CTLpeptide ions, and molecules belonging to this toxin family were identified inspots 6, 9-11  andsemi-quantitative (ELISA) evidence that these antivenoms exhibited variableparaspecific immunoreactivity towards nineteen venoms of bothropic snakes,including B. brazili in addition to B.alternatus, B. atrox, B.bilineatus, B. castelnaudi, B.cotiara, B. erythromelas, B.fonsecai, B. hyoprorus, B.insularis, B. itapetiningae, B.jararaca, B. jararacussu, B.leucurus, B. marajoensis, B.moojeni, B. neuwiedi, B. pirajai,and B. pradoi. In Brazil, envenomings by bothropic species are clinically treated with equinepolyspecific pentabothropic (SAB) or antibothropic-lachetic F(ab')z et al.  haverepB. brazili (Par\u00e1) and B. jararaca(reference venom). Analysis of the concentration-dependent immunocapturingprofile of the SAB antivenom affinity columns showed paraspecificimmunoreactivity against all the toxin classes of B. brazilivenom 2 antigen-recognition sites were occupied, the antivenomicsresults suggest that 19% of the SAB antibodies recognized toxins from B.brazili venom. This figure fall within the range of percentages(6-28%) of antitoxin antibodies determined for a number of commercial antivenoms[Here, we have applied third-generation antivenomics , 41 to clivenom , Table 1B. jararaca(SE)  venom proteinsper vial. Assuming full occupancy of the two F(ab')2antigen-recognition sites, the antivenomics results indicate that 23.7% of SABF(ab')2 are toxin-binding antibodies. Moreover, theneutralization potency of the SAB antivenom specified by Butantan Institute, 50mg of Bothrops jararaca reference venom/vial (10 mL), mirrorsits maximal binding capacity, indicating that virtually all (50/50.6 = 98.8%)toxin-binding F(ab')2 antibodies may contribute to the capability ofthe SAB antivenom to neutralize the lethality of the homologous venom. On theother hand, the paraspecificity of SAB toward toxins of the heterologousB. brazili venom is due to the remarkable conservation ofantigenic determinants already present in the venom of the last common ancestorof the \u201cjararaca\u201d and \u201cjararacussu\u201d clades, an event that has been dated closeto the base of the radiation of genus Bothrops in the middleMiocene 14.07 Mya (CI95% 16.37-11.75 Mya) [For comparison, analysis of the concentration-dependent antivenomics profile ofthe SAB antivenom against the reference venom of aca(SE) , Table 2.75 Mya) , 57.in vitro preclinical data to an invivo scenario is not straightforward. Thus, although the similartotal binding capacity of SAB antivenom towards B. jararaca andB. brazili venoms could be interpreted as indicative forits equivalent therapeutic potential against human envenomings by eitherspecies, the devil is in the details. In this regard, it is worth noting thatalthough the major toxin classes PLA2, PIII-SVMP, PI-SVMP, and SVSPrepresent, respectively, 30.6%, 24.6%, 15.5%, and 13.5% of the total venomarsenal of B. brazili, the SAB antivenom\u2019s antibodiescontributing to its paraspecific recognition of B. brazilitoxins are biasedly distributed against PI-SVMP (41%), PIII-SVMP (32%), SVSP(9.3%), and PLA2 (8.8%). This suggests that the ability of SAB toneutralize the toxic activities of Brazil\u2019s lancehead venom associated withPIII- and PI-SVMPs, and SVSPs is equivalent to, or greater than, the B.jararaca reference venom. On the other hand, counteracting thetoxic activities of the major B. brazili venom PLA2molecules may require several times the amount of antivenom. Translating B. brazili is about 270 mg dry weight. For comparison, theaverage yield reported for B. jararaca .These figures suggest that the same therapeutic potency of SAB against bothvenoms. However, the treatment of a Brazil's lancehead bite injecting an averageamount of venom would require a 5-13 higher SAB dose than for a similarenvenoming by B. jararaca. The average venom yield of y weight ;40-70 mB. brazili geographically restricted to regions north(Guiana Shield clade) and south  of the AmazonRiver [B. brazili clades has been dated back to the Miocene-Plioceneborder, 4.65 Mya, and the best\u2010fit scenario includes colonization of the AtlanticForest from an ancestor from the Guiana Shield region through a northern bridgeduring the Pleistocene about 0.36 Mya, pointing to former rain forest expansions innorth\u2010eastern South America [The Brazil\u2019s lancehead is a wide-ranging species endemic to lowlands of equatorialrainforests of northern South America. Phylogenetic analyses recovered two majorlineages of onRiver . The div America . B. brazili are consistent withthe idea that the establishment of the Amazon River has favored divergence bypromoting vicariant separation between lineages [Historical demographic analyses of lineages . The orilineages , 61. Thelineages , 63. TheB. brazili venom provide a reference map for future comparativestudies of the intraspecific intra- and inter-population variations of the venomproteome of this wide geographic distributed, yet poorly studied, rainforest snakespecies. This work represents the first comprehensive characterization of the venom proteomeof the Brazil\u2019s lancehead. The venom was sourced from Par\u00e1, a Brazilian state southof the Amazon River. The complementary RP-HPLC/SDS-PAGE and 2-DE protein profiles ofB.jararaca and B. brazili  withpotencies of 5.5 and 1.6 mg venom/mL, respectively. The antivenom showed potenciesof 6.2 and 1.4 mg/mL, respectively, in the neutralization of the PLA2activity of B. jararaca and B. brazili venoms. Thevolume of SAB antivenom that neutralized one minimal hemorrhagic dose (MHD) [B. jararaca andB. brazili venoms was 5 mL and 7.8 mL, respectively.Understanding the basis of the different effectivity of SAB antivenom againsthomologous (B. jararaca) and heterologous (B.brazili) venoms demands the quantitative assessment of itstoxin-resolved immunorecognition profile. The ability of SAB antivenom to recognize a broad spectrum of medically importantbothropic venoms has been documented in previous works spanning the last threedecades , 64-67. se (MHD)  of B. jaB. brazili is mostly due to large conservation ofimmunoreactive epitope on hemorrhagic PI- and PIII-SVMPs across much of the naturalhistory of Bothrops. On the contrary, SAB paraspecificity againstPLA2s, which comprise the major toxin class of the Brazil\u2019s lanceheadvenom arsenal, is disproportionately diminished. Our antivenomics data allow therationalization, in molecular terms, of the conclusions of the invivo neutralization assays of Muniz et al. [Herein, we have applied third-generation antivenomics to compare the specific andparaspecific immunoreactivity of the SAB antivenom against these venoms. Theremarkable paraspecificity exhibited by the Brazilian SAB antivenom against thevenom of z et al. , and pro50: median lethal dose; MCD-F:minimum coagulant dose against fibrinogen; MCD-P: minimum coagulant dose againstplasma; MDD: minimum defibrinogenating dose; MHD: minimum hemorrhagic dose; MND:minimum dermonecrotic dose; NGF: nerve growth factor; NR: non-retained; P-BAV:Peruvian bothropic antivenom; PDE: phosphodiesterase; PLA2:phospholipase A2; PLB: phospholipases B; R: retained; SAB:soro antibotr\u00f3pico (Portuguese); SDS-PAGE:SDS-polyacrylamide gel electrophoresis; SVMP: snake venom metalloproteinase;SVSP: snake venom serine proteinase; TFA: trifluoroacetic acid; TVC: total venomcomponents. 2-DE: two-dimensional gel electrophoresis; 5'-NT: 5\u2032-nucleotidase; ACN:acetonitrile; CTL: C-type lectin-like; GC: glutaminyl cyclase; IEF: isoelectricfocusing; LAO: L-amino acid oxidase; LD"}
+{"text": "Objective: Associate the impact of oral health with quality of life and subjective well-being in the community-dwelling older adults in Mexico. Methods: Non-random sample; 326 subjects: age collected (60-69 / \u2265 70); gender ; marital status (couple / no partner); schooling (0-6 years / \u22657); income for basic needs (yes / no); no depression (GDS-15), no cognitive impairment (MMSE) and comorbidity (no disease / \u2265 1 disease) to control biases. Oral conditions; Caries index (ICPOD) WHO criteria: Very low-Low; Moderate and High. Need for dental prostheses : No prostheses needed ; moderate to severe xerostomia > 17 points. Dependent variables: Quality of Life Related to Oral Health (GOHAI); 57-60 points: High perception. Subjective well-being: Moral Scale of the Geriatric Center of Philadelphia (PGCMS): Low score (0-11). Results: Age: 71.84 \u00b1 7,278; female / male (70.9 / 29.1%). Controlling confounding factors, multiple logistic regression showed that the need for multi-unit or total prostheses; high CPOD index; severe xerostomia; and low perception of well-being subjective, were associated with low GOHAI scores: P = 0.000; P = 0.004; P = 0.003; P = 0.02 respectively. Subjective well-being only was associated with severe xerostomia and low CVRSO perception: P = 0.0 1; P = 0.02 respectively. Conclusion: Taking into account various confounding factors, the Quality of Life related to Oral Health was the most affected by the deterioration of oral health."}
+{"text": "Glaesserella parasuis (G. parasuis) is a respiratory pathogen of swine and the etiological agent of Gl\u00e4sser\u2019s disease. The structural organization of genetic information, antibiotic resistance genes, potential pathogenicity, and evolutionary relationships among global G. parasuis strains remain unclear. The aim of this study was to better understand patterns of genetic variation, antibiotic resistance factors, and virulence mechanisms of this pathogen.The whole-genome sequence of a ST328 isolate from diseased swine in China was determined using Pacbio RS II and Illumina MiSeq platforms and compared with 54 isolates from China sequenced in this study and 39 strains from China and eigtht other countries sequenced by previously. Patterns of genetic variation, antibiotic resistance, and virulence mechanisms were investigated in relation to the phylogeny of the isolates. Electrotransformation experiments were performed to confirm the ability of pYL1\u2014a plasmid observed in ST328\u2014to confer antibiotic resistance.6678 transposon harbouring a unique resistance determinant. It also contained a small broad-host-range plasmid pYL1 carrying aac(6\u2019)-Ie-aph(2\u201d)-Ia and blaROB-1; when transferred to Staphylococcus aureus RN4220 by electroporation, this plasmid was highly stable under kanamycin selection. Most (85.13\u201391.74%) of the genetic variation between G. parasuis isolates was observed in the accessory genomes. Phylogenetic analysis revealed two major subgroups distinguished by country of origin, serotype, and multilocus sequence type (MLST). Novel virulence factors  and drug resistance genes  in G. parasuis were identified. Resistance determinants -Ib, norA, bacA, ksgA, and bcr) were widespread across isolates, regardless of serovar, isolation source, or geographical location.The ST328 genome contained a novel TnG. parasuis isolates provides valuable insight into the emergence and transmission of G. parasuis in the swine industry. The result suggests the importance of transposon-related and/or plasmid-related gene variations in the evolution of G. parasuis.Our comparative genomic analysis of worldwide  Glaesserella parasuis, a gram-negative bacterium in the family Pasteurellaceae  and multiple target gene mutations. To date, two \u03b2-lactam resistance genes (blaROB\u22121 and blaTEM), an aminoglycoside-resistance gene (aac (6\u2032)-Ib-cr) and a mutation in the six copies of the 23S rRNA gene, associated with macrolide resistance, have been reported in G.\u00a0parasuis -Ib, aph(6)-Id, floR, sul1, and sul2 (hhdBA), iron acquisition genes , the restriction modification system hsdS, and genes involved in sialic acid utilization   . RecentlG.\u00a0parasuis strains, the structural organization of genetic information, ARGs, potential pathogenicity determinants, and evolutionary relationships among global G.\u00a0parasuis strains remain unclear. In this study, we sequenced a multidrug-resistant isolate from diseased swine in Dongguan, China, then compared this genome sequence with those of 54 isolates from China sequenced by us and 39 strains from China and eight other countries sequenced by other researchers in order to improve our understanding of genomic diversity in G.\u00a0parasuis and provide information for gaining better control to treat these infections.Though significant effort has been focused on exploring ARGs, virulence factors and other genetic characteristics of various G.\u00a0parasuis isolate HPS-1 examined in this study belongs to serotype 4 and was originally isolated from the lungs of a pig suffering from Gl\u00e4sser\u2019s disease in a commercial pig farm in Dongguan city, Guangdong province, China, in 2017. Susceptibility to 19 antimicrobial agents was determined by the disc agar diffusion method and the broth microdilution method .Isolates were cultured on tryptic soy agar or in tryptic soy broth  supplemented with 10 mg/mL nicotinamide adenine dinucleotide and 5% bovine serum at 37\u00a0\u00b0C in 5% COCP040243. The plasmid pYL1 and transposon Tn6678 of HPS-1 were submitted to GenBank under accession number MK182379 and and MK994978, respectively. Genomic libraries of the other 53 genomes were generated and sequenced using the Illumina HiSeq 4000 system  as previously described  against 6 databases: Kyoto Encyclopedia of Genes and Genomes (KEGG), Clusters of Orthologous Groups (COG), NCBI non-redundant protein database (NR), Swiss-Prot, Gene Ontology (GO), and TrEMBL.Among the 55 isolates, one multidrug-resistant isolate (HPS-1) and one sensitive isolate (HPS-2) from diseased swine in Guangdong were randomly selected for WGS using the PacBio RSII  and Illumina MiSeq  platforms as previously described . The genescribed . WGS datescribed . Gene prescribed , and a wescribed  searchesG.\u00a0parasuis strains and 39 previously published genome sequences using single-copy core orthologs and single nucleotide polymorphisms (SNPs)  software to generate non-paralogous gene clusters .Two phylogenetic trees were constructed to assess the relatedness of the 55 s (SNPs) . Phyloges (SNPs) . The WAGE-value \u2264 1e\u22125, minimal alignment length percentage \u2265 80%) was performed against four databases for pathogenicity and drug resistance analysis: Pathogen Host Interactions (PHI), Virulence Factors of Pathogenic Bacteria (VFDB), Carbohydrate-Active enZYmes Database (CAZy), and Integrated Antibiotic Resistance Genes Database (IARDB).A whole-genome BLAST search -Ie-aph(2\u2033)-Ia, which confer to \u03b2-lactams and aminoglycosides resistance. To determine the contributions of pYL1 to penicillin and aminoglycoside antibiotic resistance, electrotransformation experiments were performed using Staphylococcus aureus RN4220 as the recipient as previously described (aac(6\u2032)-Ie-aph(2\u2033)-Ia and blaROB\u22121 genes in transformants was confirmed by PCR amplification followed by DNA sequence analysis. The primers for blaROB\u22121 (494 bp) were 5\u2032-CGCTTTGCTTATGCGTCCAC-3\u2032 (forward) and 5\u2032-ACTTTCCACGATGTTGGCGT-3\u2032. The primers for aac(6\u2032)-Ie-aph(2\u2033)-Ia (412 bp) were 5\u2032-AGAGCCTTGGGAAGATGAAGTT-3\u2032 (forward) and 5\u2032-TGCCTTAACATTTGTGGCATT-3\u2032 (reverse). The primers were designed using NCBI Primer-BLAST. The PCR conditions were as follows: initial denaturation at 95\u00a0\u00b0C for 5 min, 30 cycles of amplification , followed by extension at 72\u00a0\u00b0C for 10 min. The PCR products were purified and sequenced by Majorbio Company . The MICs of S.\u00a0aureus RN4220 and five transformants were determined by Etest (Liofilchems.r.l.) according to the manufacturer\u2019s instructions.Plasmid pYL1 harboring two antimicrobial resistance genes, escribed . Transfoaac(6\u2032)-Ie-aph(2\u2033)-Ia and blaROB\u22121 was determined by serial passages for 15 consecutive days at 1:1000 dilutions into fresh BHI, with or without antibiotic (kanamycin) pressure. Serially diluted cultures were spread on BHI agar plates with or without kanamycin (8 \u00b5g/mL), and the resistance retention rate was determined by randomly picking at least 50 colonies from the BHI plates, spotting them onto new BHI plates with kanamycin (8 \u00b5g/mL), and calculating the ratio of resistant to total colonies. Both the resistant and susceptible colonies from the plates were randomly picked and subjected to PCR for detection of blaROB\u22121 and aac(6\u2032)-Ie-aph(2\u2033)-Ia.The stability of plasmids carrying G. parasuis gene content compared with the core genomes and the number of unique genes ranged from 0 to 103 indicating that 0\u20134.6% of the genome consists of strain-specific accessory genes  core biosynthesis, and galU and galE, resulting in impaired biofilm formation, were universally present in the G. parasuis isolates.Variation in virulence and stress resistance genes was observed among ubgroups . All 94 G. parasuis, including the \u03b2-lactam-resistant gene blaROB\u22121, tetracycline resistance genes tetB, aminoglycoside resistance genes aph(3\u2032)-Ib and aac(6\u2032)-Ie-aph(2\u2033)-Ia, fluoroquinolone resistance gene norA, chloramphenicol resistance genes catIII and floR, sulfonamide resistance gene sul2 were discovered (sul2 and aph(3\u2032)-Ib, and \u03b2-lactam-resistant genes pbp1a and pbp3a were universally present in the G. parasuis isolates , aminoglycosides (aph(3\u2032)-Ib, aac(6\u2032)-Ie-aph(2\u2033)-Ia), and \u03b2-lactam (blaROB\u22121) (bcr) and multidrug resistance (acrB).Following sequencing and assembly, a 2,326,414-bp chromosome with an average G+C content of 40.03%, and a 7,777-bp small plasmid sequence (pYL1) with an average G+C content of 33.32% were identified in strain HPS-1 . HPS-1 elaROB\u22121) . Furtherhttps://transposon.lstmed.ac.uk/). This transposon harbours two 966-bp IS110 family transposases at both ends, two toxin genes pilT and phd, two genes associated with the two-component signal transduction system cpxA and cpxR, one efflux pump-associated gene bcr, and four genes encoding hypothetical proteins with unknown function (34)-methyltransferase MnmD/FAD-dependent 5-carboxymethylaminomethyl-2-thiouridine (34) oxidoreductase MnmC flanked the transposon to the right and left, respectively.We also identified a novel transposon in the ST328 isolate, designated Tn6678 in the Tn Number Registry , SH0165 , ZJ0906 , and str. Nagasaki ]. The only differences in these five chromosomes were in the transposases, but transposon Tn6678 had two complete inverted repeats of IS110 transposases flanked by 32-bp inverted repeats of ISNme5 at both ends  were identified in four oth ends , suggestbcr gene returned a large set of divergently related sequences using default parameters. These sequences were annotated as bicyclomycin/multidrug efflux system, Bcr/CflA family drug resistance efflux transporter, Bcr/CflA family multidrug efflux major facilitator superfamily (MFS) transporter or drug resistance transporter, and Bcr/CflA subfamily. Phylograms revealed that the bcr gene in HPS-1 was most closely related to homologs identified in other members of the Pasteurellaceae, particularly G. parasuis, Actinobacillus indolicus, Bibersteinia trehalosi, Actinobacillus , and Mannheimia (M. haemolytica and M. varigena), all of which are known causative agents of upper respiratory tract infections -Ie-aph(2\u2033)-Ia. Four ORFs were identified to encode a 3\u2032-truncated transposase protein ISApl1 (30 amino acids), a Rep-like protein (444 amino acids) involved in plasmid replication, and two Mob proteins, MobC (144 amino acids) and MobA (541 amino acids), associated with plasmid mobilization . This finding indicated that plasmid pYL1 carrying blaROB\u22121 and aac(6\u2032)-Ie-aph(2\u2033)-Ia contributed to the penicillin resistance and aminoglycoside antibiotic resistance in S. aureus RN4220 transformants. Furthermore, the plasmid showed low stability in S. aureus without antibiotic pressure, as only 52.5%, 30.48%, and 2.68% of transformants maintained the kanamycin resistance after five, six, and seven subcultures, respectively. However, the plasmid can be conserved in S. aureus cultured with kanamycin, as 100% of the colonies remained resistant to kanamycin after 10 subcultures, as confirmed by PCR mapping.Transformation of pYL1 into G. parasuis pan-genome is vast, and unique genes can be continuously be identified upon sequencing more G. parasuis genomes. However, the isolates in this study with \u223c3.34% core genes, primarily isolated from China, displayed further diversity and higher variability than isolates with only \u223c14% core genes, primarily obtained from the UK , in line with potentially virulent strains isolated from the nasal cavities of healthy pigs -Ib, tetB, tetD, aac(6\u2032)-Ie-aph(2\u2033)-Ia, catIII, and floR genes have previously been identified in G. parasuis  family, has been analysed to date. Efflux pump AcrB may play a role in multidrug resistance, and the acrAB gene cluster could affect the efflux of macrolides in G. parasuis  have been reported in G. parasuis -Ie-aph(2\u2033)-Ia. The ROB-1 of plasmid pYL1 had a typical size of 305 bp, in line with functionally active members of the ROB-1 family from different plasmids in Pasteurellaceae species. AAC(6\u2032)-Ie-APH(2\u2032)-Ia, the most important aminoglycoside-resistance enzyme in gram-positive bacteria conferring resistance to almost all known aminoglycoside antibiotics in clinical use, also had a typical size of 479 amino acids in this family (aac(6\u2032)-Ib-cr is considered the most prominent aminoglycoside-resistance gene in G. parasuis  and drug resistance genes  in G. parasuis. Resistance determinants -Ib, norA, bacA, ksgA, and bcr) were widespread across isolates, regardless of serovar, isolation source, or geographical location. Future research focused on a larger sample of G. parasuis isolates worldwide will further increase understanding of the rapid development of antibiotic resistance associated with mobile genetic elements in this important animal pathogen.In summary, our results shed new light on the importance of genomic variations, especially transposon-related and/or plasmid-related gene variations, in the evolution of 10.7717/peerj.9293/supp-1Table S1P. aeruginosa .MIC: minimum inhibitory concentration; R: resistant; S: susceptible. a Interpreted according to Clinical and Laboratory Standards Institution (CLSI) guidelines for Click here for additional data file.10.7717/peerj.9293/supp-2Table S2N.A. represents unknown information; ST: sequence type; UT: untypeable.Click here for additional data file.10.7717/peerj.9293/supp-3Table S3Click here for additional data file.10.7717/peerj.9293/supp-4Table S4Click here for additional data file.10.7717/peerj.9293/supp-5Table S5Click here for additional data file.10.7717/peerj.9293/supp-6Figure S1The tree was constructed with MEGA 7 using a maximum-likelihood optimality criterion as implemented in PhyML v3.0 with 1, 000 bootstrap replicates. The annotation rings surrounding the tree, from inside to outside, depict (1) serotype, (2) host, (3) geographic region and (4) year of sample collection. The branch colors denote two major lineages, lineage I (pink) and lineage II (green).Click here for additional data file.10.7717/peerj.9293/supp-7Figure S2Glaesserella sp. 15\u2013184 was chosen as an outgroup. The branch colors denote two major lineages, lineage I (pink) and lineage II (green).Click here for additional data file.10.7717/peerj.9293/supp-8Figure S3From inside to outside, the first circle represents the genome size of HPS-1; the second circle represents the GC skew; the third circle represents the GC content; the fourth circle and the seventh circle represent the COG (cluster of orthologous groups) designation of each coding sequence (CDS); the fifth and sixth circles represent the position of CDS, tRNA, and rRNA on the genome.Click here for additional data file.10.7717/peerj.9293/supp-9Supplemental Information 1Click here for additional data file.10.7717/peerj.9293/supp-10Supplemental Information 2Click here for additional data file.10.7717/peerj.9293/supp-11Supplemental Information 3Click here for additional data file.10.7717/peerj.9293/supp-12Supplemental Information 4Click here for additional data file.10.7717/peerj.9293/supp-13Supplemental Information 5Click here for 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+{"text": "These fungi have worldwide distribution, but with the exception of Articulosporatetracladia, no aquatic hyphomycete taxon was previously recorded on Madeira Island. Aquatic hyphomycetes were sampled from 40 sites, distributed by 27 permanent streams in 2015, to provide the distribution of aquatic hyphomycetes in Madeira Island streams.Aquatic hyphomycetes were recorded belonging to three classes of Ascomycota. All taxa are new records for Madeira Archipelago, except Articulosporatetracladia and four are reported for the first time in Macaronesian biogeographic region.In this study, a total of 21 species of aquatic  Aquatice scarce . In factal zones . Despiteems e.g. .fungi started almost two centuries ago with the work of th century, many mycological studies increased the number of records for Madeira Archipelago (North Atlantic), including numerous descriptions of species new to science . Lying in the subtropical region, Madeira\u2019s climate is influenced by winds from NE and the Canary Islands current. The Island has a mild oceanic climate, both in winter and summer with mild temperatures ranging from 15.9\u00b0C in February up to 22.3\u00b0C in August , relative humidity between 55 and 75% and annual rainfall between 500 and 1,000 mm  distributed by 27 permanent streams : 384 (1942)3690A495-A84F-50C3-A2E6-2BEDD6529505Cosmopolitan .Madeira distribution: Streams in agricultural and natural areas at low to moderate altitude: Ribeira de S\u00e3o Vicente (MAD04); Ribeira do Juncal (MAD15); Ribeira do Faial (MAD16); Ribeira Primeira (MAD18); Ribeira de S\u00e3o Jorge (MAD37).Habitat: Submerged leaf litter .(H.J. Huds.) L. Lombard & Crous, in Lombard, van der Merwe, Groenewald & Crous, Stud. Mycol. 80: 207 (2015)672BA860-8D1E-5A9B-9963-0C35130DCB8ECosmopolitan .Madeira distribution: Stream in agricultural and natural areas at low to moderate altitude: Ribeira de S\u00e3o Vicente (MAD04); Ribeira Brava (MAD07); Ribeira do Juncal ; Ribeira do Faial (MAD16); Ribeira da Janela (MAD21); Ribeira de S\u00e3o Roque do Faial (MAD33); Ribeira dos Arcos (MAD36); Ribeira de S\u00e3o Jorge (MAD17); Ribeira da Fonte do Bugio (MAD39).Habitat: Submerged leaf litter .(R.H. Petersen) Dyko, Trans. Br. mycol. Soc. 70 (3): 412 (1978)8A859C84-48D6-5382-BE6E-FC0C03AD9EDDCosmopolitan .Madeira distribution: Streams in natural areas at low to high altitude: Ribeira Brava (MAD07); Corgo da Ribeira de An\u00e9is (MAD11); Ribeira do C\u00f3rrego do Arrochete (MAD30); Ribeira de S\u00e3o Jorge (MAD37).Habitat: Submerged leaf litter [e.g. Ilexperado (xperado ].(Ingold) Sv. Nilsson ex Marvanov\u00e1 & Sv. Nilsson, Trans. Br. mycol. Soc. 57 (3): 532 (1971)15B4E8EC-42DF-52E6-B357-42CBE90D2558Cosmopolitan .Madeira distribution: Streams in natural areas at high altitude: Ribeira da Janela ; Ribeira do C\u00f3rrego do Arrochete (MAD30).Habitat: Submerged leaf litter .(Sacc. & Therry) L. Lombard & Crous, in Lombard, van der Merwe, Groenewald & Crous, Phytopath. Mediterr. 53(3): 528 (2014)9B3C3176-C801-5961-B75C-48C31DFB8EEDCosmopolitan .Madeira distribution: Streams in urban, agricultural and natural areas at low to high altitude: Ribeira Brava (MAD02); Ribeira da Vargem (MAD03); Ribeira Brava ; Ribeira dos Socorridos (MAD09); Ribeira do Juncal ; Ribeira do Machico (MAD17); Ribeira Primeira (MAD18); Ribeira da Janela ; Ribeira do Alecrim (MAD28); Ribeira do C\u00f3rrego do Arrochete (MAD30).Habitat: Submerged leaf litter .(Grove) Ingold, Trans. Br. mycol. Soc. 25(4): 371(1942)29EA0E24-82EE-5239-85A5-1555AED4BAD6Cosmopolitan .Madeira distribution: Streams in agricultural areas at low to moderate altitude: Ribeira dos Socorridos (MAD01); Ribeira Brava (MAD02); Ribeira do Juncal (MAD15).Habitat: Submerged leaf litter [e.g. Clethraarborea, Ilexperado : 624 (1974)68D79614-DC15-5AF0-991B-F21A84FD1786Cosmopolitan .Madeira distribution: Streams in urban, agricultural and natural areas at low to high altitude: Ribeira dos Socorridos (MAD01); Ribeira de S\u00e3o Vicente (MAD04); Ribeira Grande (MAD05); Ribeira Brava ; Ribeira do Cidr\u00e3o (MAD12); Ribeira do Juncal ; Ribeira do Faial (MAD16); Ribeira do Machico (MAD17); Ribeira Primeira ; Ribeira da Janela ; Ribeiro Frio (MAD29); Ribeira do C\u00f3rrego do Arrochete (MAD30); Ribeira da Metade (MAD31); Ribeira das Lajes (MAD32); Ribeira de S\u00e3o Jorge (MAD35); Ribeira dos Arcos (MAD36).Habitat: Submerged leaf litter [e.g. Acaciamelanoxylon, Acerrubrum, Alnusglutinosa, Clethraarborea, Cryptomeriajaponica, Ilexperado, Pittosporumundulatum, Quercusrobur, Rhododendronmaximum : 346 (1975)F45FE042-099D-5651-8E96-FC87B0CB74C9Cosmopolitan .Madeira distribution: Streams in agricultural and natural areas at low to moderate altitude: Ribeira dos Socorridos (MAD01); Ribeira Brava (MAD08); Ribeira do Juncal (MAD15); Ribeira do Faial (MAD16); Ribeira do Machico (MAD17); Ribeira Primeira (MAD18); Ribeira da Janela (MAD21); Ribeiro Frio (MAD29); Ribeira do C\u00f3rrego do Arrochete (MAD30); Ribeira das Lajes (MAD32); Ribeira Seca (MAD34); Ribeira de S\u00e3o Jorge (MAD35); Ribeira dos Arcos (MAD36); Ribeira de Santa Luzia (MAD38).Habitat: Submerged leaf litter [e.g. Acaciamelanoxylon, Alnusglutinosa, Clethraarborea, Ilexperado, Pittosporumundulatum, Quercusrobur : 152 (1943)E9A99FEC-221E-5C74-98E4-1A77CEA78B80Cosmopolitan .Madeira distribution: Streams in urban, agricultural and natural areas at low to high altitude: Ribeira dos Socorridos (MAD01); Ribeira Brava ; Ribeira de S\u00e3o Vicente (MAD04); Ribeira Grande ; Ribeira da Gomeira (MAD10); Ribeira do Cidr\u00e3o (MAD12); Ribeira do Juncal ; Ribeira do Faial (MAD16); Ribeira do Machico (MAD17); Ribeira Primeira (MAD18); Ribeira da Janela (MAD21); Ribeira dos Cedros (MAD25); Ribeiro Frio (MAD29); Ribeira de S\u00e3o Roque do Faial (MAD33); Ribeira Seca (MAD34); Ribeira de S\u00e3o Jorge ; Ribeira dos Arcos (MAD36); Ribeira de Santa Luzia (MAD38).Habitat: Submerged leaf litter [e.g. Acaciamelanoxylon, Acerrubrum L., Alnusglutinosa, Clethraarborea, Eucalyptusglobulus, Ilexperado, Pittosporumundulatum, Quercusrobur, Rhododendronmaximum , Leotiomycetes encompassed 43% of the taxa recorded. At the order level, the 21 ascomycetes were distributed by Helotiales (12 spp.), Hypocreales (2 spp.), Leotiales (1 sp.), Microascales (1 sp.), Sordariales (1 sp.) and Incertae sedis (4 spp.), according to In the present study, we found a total of 21 aquatic Articulosporatetracladia, Clavariopsisaquatica, Lemonnieraaquatica, Lunulosporacurvula, Tetrachaetumelegans, Tetracladiummarchalianum, Tricladiumchaetocladium and Triscelophorusmonosporus, which were the most ubiquitous aquatic hyphomycetes in Madeira streams. Two taxa, Lemonniera sp. and Tetracladiumfurcatum, had a sporadic occurrence, being found at only one or two sampling sites. A maximum of 14 species was recorded in MAD15 and a minimum of one species in MAD13 and MAD20, with a mean richness of eight species per stream site. Higher altitude stream sites (> 800 m a.s.l.) in natural areas, such as MAD07, MAD08, MAD24 and MAD26, displayed higher taxa richness , compared with coastal (< 25 m a.s.l.) and urban stream sites, such as MAD13, MAD20, MAD32, MAD33, MAD37, MAD38, MAD39 and MAD40 (5.8 \u00b1 1.2). All species, with the exception of Articulosporatetracladia reported by From the 21 species identified, none occurred in all 40 studied stream sites and only 8 taxa occurred in more than 50% of the stream sites: hyphomycetes in insular streams from Madeira Island. Twenty-one taxa are recorded for Madeira Island, which is lower than what is reported for the Azores archipelago  of the Archipelago. In this context, it is essential to increase the sampling effort for both archipelagos, as well as to survey multiple matrices  in order to find a greater diversity of aquatic hyphomycetes. To the best of our knowledge, no data of recorded species exist for the other Macaronesia archipelagos, such as Canary Islands and Cabo Verde Archipelago.Here we present the first study that explored the distribution of aquatic hyphomycetes species have a cosmopolitan distribution Aquatic Data typeData recordsBrief descriptionMetafileFile: oo_404009.xmlhttps://binary.pensoft.net/file/404009Raposeiro, P.M.; Faustino, H.; Ferreira, V.; Gon\u00e7alves, V.23207FAB-3574-536A-BFDF-036E130E693C10.3897/BDJ.8.e53690.suppl4Supplementary material 4Occurrence of the aquatic hyphomycete species found in this study in 15 geographic regions defined based on the geographic location and climatic influence in the western and eastern hemispheres Data typeTableBrief descriptionTableFile: oo_420217.csvhttps://binary.pensoft.net/file/420217Raposeiro, P.M.; Faustino, H.; Ferreira, V.; Gon\u00e7alves, V."}
+{"text": "This is the first report of plasmid-mediated colistin resistance mcr-8 gene from a clinical Klebsiella pneumoniae K9 isolate recovered from Lebanon. The isolate was characterized phenotypically and genotypically through both short and long read whole-genome sequencing, plasmid typing and conjugation assays. k9 belonged to sequence type 15 and harbored 31 antimicrobial resistance genes. The mcr-8.1 variant was carried on a novel ~\u2009300\u2009kb multireplicon plasmid having IncFIA, IncR and IncHI1B. The plasmid was conjugative and carried a plethora of antimicrobial resistance determinants. The introduction of novel mcr variants in Lebanon poses an alarming health concern. Surveillance and screening for colistin resistant Enterobacteriaceae and mcr in livestock, animal farms, imported meat and poultry is highly recommended along with monitoring antibiotic use.Colistin is considered as a last resort treatment for infections caused by multidrug-resistant Enterobacteriaceae. Plasmid-mediated mobile colistin resistance ( Klebsiella pneumoniae (CRKP) [mcr-1, was identified in 2015 in Enterobacteriaceae, mainly Escherichia coli and K. pneumoniae, collected from animals and humans in China [mcr variants have been described, designated as mcr-1 -9, isolated from humans, animals and the environment [Colistin is the last-line treatment option for infections caused by carbapenem resistant Enterobacteriaceae such as carbapenem-resistant e (CRKP) . The firironment .mcr-8 located on an IncFII-type conjugative plasmid was first described in 2018 in China in K. pneumoniae collected from both animals and humans [d humans . Shortlyd humans  and Alged humans .K. pneumoniae in Lebanon was in 2017 [mcr-1-positive E. coli from a human clinical isolate in Lebanon has been recently reported [mcr-1 was widely detected in Gram-negative bacilli isolated from poultry [mcr-8.1 gene variant in an ST-15\u2009K. pneumoniae isolate recovered from Lebanon.The first report of a colistin resistant reported . On the  poultry , swine f poultry  and wate poultry . This isKlebsiella pneumoniae and designated as k9.The isolate was recovered from the urine sample of a 50-years old female patient on the 28th of August 2018 at the Clinical Microbiology Laboratory of the American University of Beirut Medical Centre (AUBMC), Lebanon. The identification of the isolate was performed using Matrix-Assisted Laser Desorption/Ionization Time of Flight (MALDI-TOF) system . The isolate was identified as E. coli transconjugant was further determined by using the broth microdilution method according to the recommendations from the European Committee on Antimicrobial Susceptibility Testing (EUCAST) [Antimicrobial susceptibility testing was performed using the disk diffusion assay on Mueller-Hinton agar and included a panel of 24 antibiotics belonging to 16 different classes (Table S(EUCAST) .DNA extraction was performed from fresh bacterial colonies using the Nucleospin\u00ae Tissue kit  according to the manufacturer\u2019s instructions.mcr-8 was confirmed through PCR using MCR-8F 5\u2032-AACCGCCAGAGCACAGAATT-3\u2032 and MCR-8R 5\u2032-TTCCCCCAGCGATTCTCCAT-3\u2019primer pair as previously described [The presence of escribed .Plasmid characterization was performed using the DIATHEVA plasmid based replicon typing (PBRT) kit  through a polymerase chain reaction (PCR) based replicon typing method consisting of eight multiplex PCR assays for the amplification of 25 replicons: A/C, B/O, FIA, FIB, FIB-M, FIC, FII, FIIK, FIIS, HI1, HI2, HIB-M, I1, I2, K, L/M, N, P, R, T, U, W, X1, X2, and Y found in the family Enterobacteriaceae. Positive controls were included for all reactions. All PCR reactions were performed according to the manufacturer\u2019s instructions.E. coli J53 strain. Selection of transconjuguants was done on Uriselect agar  supplemented with 100\u2009mg/L sodium azide and 4\u2009\u03bcg/ml colistin. E. coli transconjugants were typed by PBRT to detect plasmid replicons.Conjugation was performed with an azide-resistant Genomic libraries were constructed using the Nextera XT DNA library preparation kit with dual indexing (Illumina). The libraries were sequenced on an Illumina MiSeq with 250\u2009bp\u2009\u00d7\u20092 read length. Genome assembly was performed de novo using Spades Genome Assembler Version 3.6.0 . Qualitymcr8-carrying plasmid was sequenced using PacBio long-read sequencing technology on the Sequel platform . Library preparation was performed according to the manufacturer\u2019s instructions for microbial multiplexing. G-tubes  were used for DNA shearing, and no size selection was performed.The http://rast.nmpdr.org) [www.genomicepidemiology.org) were used to determine the ST, presence of resistance genes and plasmid content, respectively. The sequences encoding for plasmids were then extracted and aligned to references obtained from NCBI as previously described [https://www-is.biotoul.fr) [wzi gene was performed using Kaptive (www.kaptive.holtlab.net) [http://bigsdb.pasteur.fr. PlasmidsSpades was used to assemble the plasmid sequences [The assembled genomes were annotated using the RAST online server (pdr.org) . MLST 2.pdr.org) , ResFindpdr.org) , PointFipdr.org)  and Plaspdr.org)  availablescribed . The ISftoul.fr)  was usedlab.net) . The preequences . Plasmidequences .pmrA, pmrB, pmrC, pmrD, phoP, phoQ, mgrB, crrA and crrB genes were investigated in silico using both nucleic and amino acid alignments compared to the wild-type sequence of K. pneumoniae MGH 78578 (GenBank accession no. CP000647). SNAP [http://www.hiv.lanl.gov) was used to calculate synonymous and non-synonymous substitution rates based on the codon-aligned nucleotide sequences of pmrA and pmrB against the K. pneumoniae MGH 78578 references.The presence of known mutations conferring colistin resistance on the 7). SNAP   and fimbriae .K9 genome size was 5,656,813\u2009bp with a GC content of 56.9%. The best match for the capsular locus was KL107 (Blastn identity: 89.60%) with idFinder  includinmcr-8.1 positive isolates recovered so far (Table\u00a0R. ornithinolytica QDRO2 (Accession no. MK097469.1) [n\u2009=\u200931) than other mcr-8.1 positive isolates with 16, 18 and 23 antimicrobial resistance genes (AMRs) detected in isolates KP95, KP91 and QDRO2, respectively. The latter isolates represented different STs and were recovered from both humans and animal sources [k9 was compared to all other 97469.1)  KP91, KP97469.1) , respect sources .Table 1k9 had a colistin MIC of 10\u2009\u03bcg/mL , aminoglycosides (aac(6\u2032)-Ib-cr, aadA1, aadA16, aadA2b and aph(3\u2032)-Ia), quinolones (aac(6\u2032)-Ib-cr, oqxA, oqxB, qnrB2, qnrB4, and qnrB52), sulphonamides (sul1 and sul3), fosfomycin (fosA), macrolides (mph(A),mph(E),msr(E)), tetracycline (tetD), trimethoprim (dfrA27), phenicol , rifampicin (arr-3) and colistin (mcr-8).Studying antibiotic resistance mechanisms revealed the presence of 31 different resistance genes on the plasmid including resistance to: \u03b2-lactams  revealed a dN/dS ratio of 1.131 for pmrA and 1.181 for pmrB indicating a positive selection for non-synonymous mutations , ompK36 sequence (Accession no. Z33506.1) and ompK37 (Accession no. AJ011502.1). We detected a premature stop codon in ompK35 (Y76*), and 22 different amino acids in ompK36 and two in ompK37. A premature stop codon was also detected in ramR (K194*); a positive regulator of the AcrAB efflux system, compared to an intact ramR reference genes (Accession no. KY465996.1).Mutations in mcr variant was mcr-8.1 showing 100% identity to mcr-8.1 carried by K. pneumoniae plasmid pKP91 (Accession no. MG736312), a\u2009~\u200990-kb IncFII-type plasmid obtained from swine fecal material in China [Raoutella ornithinolytica QDRO2 (Accession no. MK097469.1) [Blast analysis revealed that the in China  and in R97469.1) mcr-8.1 recovered from k9 was further compared with other mcr-8 variants. The amino acid sequences and the genetic environment of mcr-8.1 was most similar to mcr-8.3 and mcr-8.4 then to mcr-8.2, the latter being recovered from K. quasipneumoniae in China [mcr-8.2 also harboured more mutations compared to mcr-8.1 than the other variants  and pKPN-065  isolated from China and USA [rep genes also found in pKP91 and IncHI1B rep gene also found in pKPN-065. IncFIA was bracketed by two IS1 insertion sequences while IS6 was present downstream of IncR. Parts of pk9 plasmid carrying various AMR determinants also aligned to pR50\u201374 (Accession no. CP040362.1) collected from a rabbit fecal sample in China in 2017, pYDC676 collected from patients in the USA in 2014 (Accession no. KT225462.1) [ and USA , respectsul1, aadA16, dfrA27, arr-3 and aac(6\u2032)-Ib-cr5. It closest identity  to Enterobacter hormaechei plasmid pM206-NDM1 collected in Japan (Accession no. AP018830.1) [sul1, qnr, sul1, aadA16 and aac(6\u2032)-Ib-cr5 showing 100% identity and 98% coverage to K. pneumoniae Kp_29407 (Accession no. LR736030.1). Other resistance determinants present on the pk9 included tetA, qnrB4, blaDHA-1, apha1b, sul3, aadA1\u20133, cmlA, floR and ampC. Various sulfonamide resistance genes were identified including four copies of sul1 in addition to sul3 and bcr (bicyclomycin resistance protein).A plethora of resistance genes were detected on pk9. An AMR cassette was located between 14,200\u2009bp and 18,300\u2009bp encoding 18830.1) . Anothermcr gene variants represents a major health concern worldwide. In this study, we identified a multireplicon plasmid carrying the mcr-8.1 gene variant in a K. pneumoniae clinical isolate recovered from a urine sample in Lebanon. This represents the first report of mcr-8 from Lebanon. mcr-8.1 showed 100% gene sequence similarity to mcr-8.1 encoded by pKP91 plasmid originating from a K. pneumoniae KP91 collected in 2018 from swine fecal material in China [mcr variants have been described (mcr-1 to mcr-9) from human, animal and environmental sources [mcr gene in the wildlife and particularly in surface water samples [The plasmid-mediated spread of in China . The detin China , 32. Sin sources , showing samples .mcr-8.1 was located on a 324,283\u2009bp multireplicon plasmid showing highest nucleotide similarity (99\u2013100%) to plasmids pF10AN_1 (Accession no. CP026154.1) and pKPN-065 (Accession no. CP015026.1) isolated from China and USA [rep genes also found in pKP91 [rep gene also found in pKPN-065. Recently, mcr-8.2 was detected on a large, hybrid plasmid containing IncQ, IncR, and IncFII replicons [mcr gene spread [ and USA , respecteplicons  confirmie spread .mcr gene family could be transmitted via the food chain, and so its prudent use in both human and veterinary medicine is of paramount importance [mcr-8 in livestock and animal farms in Lebanon as well as in imported meat and poultry is recommended to track the environmental influences contributing to the development and dissemination of resistance determinants propagating on large mobile genetic vehicles.Colistin resistant bacteria and the portance . Polymyxportance . FurtherAdditional file 1: Table S1. Antimicrobial susceptibility and MIC results of K. pneumoniae k9. Antimicrobial susceptibility was performed using the disk diffusion technique; MICs of ertapenem, imipenem and meropenem were determined using the E-test methodology; MIC of colistin was determined using the broth microdilution method. Antibiotics were divided by their drug categories. GM: gentamycin 10\u03bcg; IP: imipenem; MEM: meropenem 10\u03bcg; IPM: imipenem 10\u03bcg; PM: cefepime; CTX: cefotaxime 30\u03bcg; CXM: cefuroxime 30\u03bcg; FEP: cefepime 30\u03bcg; CZD: ceftazidime 10\u03bcg; CZN: cefazolin 30\u03bcg; SCF: cefoperazone/sulbactam 75/30\u03bcg; FOX: cefoxitin 30\u03bcg; NOR: norfloxacin 5\u03bcg; CIP: ciprofloxacin 5\u03bcg; FAD: fusidic acid 10\u03bcg; TGC: tigecycline 15\u03bcg; CMN: clindamycin 2\u03bcg; ERY: erythromycin 15\u03bcg; ATM: aztreonam 30\u03bcg; LZD: linezolid 30\u03bcg; AMX: amoxicilin 25\u03bcg; PIR: pipercacillin 100\u03bcg; AMP: amipicillin 10\u03bcg; TZP: pipercacillin/tazobactam 100/10\u03bcg; FOS: fosfomycin 200\u03bcg; TE: tetracycline 30\u03bcg; TS: trimethoprim/sulfamethoxazole; CO: colistin; Dark blue: resistant, light blue: intermediate resistance, white: susceptible; MICs are in \u03bcg/mL; NA: not available.Additional file 2: Figure S1. SNAP plots with potential synonymous and non-synonymous substitutions in pmrA (A) and pmrB (B). Alignment was performed against the query sequence of K. pneumoniae MGH 78578 wild-type chromosomal genes."}
+{"text": "Since electronic cigarettes (e-cigarettes) entered the U.S. marketplace in 2007, the landscape has evolved to include different product types  and flavored e-liquids , which have contributed to increases in youth use , from 7.7 million to 17.1 million units per 4-week interval. ; however, within the context of this general increase, sales fluctuated . During Among total e-cigarette unit sales during September 2014\u2013August 2019, the proportion that were prefilled cartridges increased from 47.5% to 89.4% (AIPC\u00a0=\u00a01.0) (p<0.05) . The proAmong total e-cigarette unit sales during September 2014\u2013August 2019, the proportion accounted for by mint products increased from 0.01% to 43.4% (AIPC\u00a0=\u00a010.5) (p<0.05) . During Among prefilled cartridge sales during September 2014\u2013August 2019, the percentage that were mint increased from <0.1% to 47.6% (AIPC\u00a0=\u00a014.1) (p<0.05) . During Among disposable e-cigarette sales during September 2014\u2013May 2020, the percentage of sales of tobacco-flavored and menthol-flavored products decreased; sales of tobacco-flavored e-cigarettes accounted for 17.2% and menthol-flavored accounted for 10.2% of all disposable e-cigarette sales in May 2020, (p<0.05). . During 1,2).During November 2016\u2013August 2019, total e-cigarette unit sales in the U.S. increased nearly 300%. Although prefilled cartridges remained the leading product type sold, disposable sales increased beginning in August 2019, reaching 19.8% of total sales by May 2020. Among prefilled cartridge sales, the proportion of mint-flavored products declined beginning in August 2019; by May 2020, menthol (61.8%) and tobacco (37.1%) flavors dominated the market. Among disposable e-cigarette sales, tobacco-flavored and menthol-flavored sales decreased during September 2014\u2013May 2020; during the same period, the proportion of sales that were mint and all other flavors increased, with mint reaching 10.5% and all other flavors reaching 62.1% of total sales by May 2020. Continued monitoring of e-cigarette sales could inform strategies to reduce use among U.S. youths, including strategies that address youth-appealing product innovations and flavors  products. Flavored e-cigarette sales patterns by product type are likely influenced by multiple factors. For example, JUUL voluntarily removed mango, creme, fruit, and cucumber flavored cartridges from retail stores (November 2018) and online (October 2019)The findings in this report are subject to at least three limitations. First, sales data did not include purchases from the Internet or \u201cvape shops,\u201d which accounted for approximately one half of U.S. e-cigarette sales in 2019;1,2). In the U.S., e-cigarette use is markedly higher among youths than adults; in 2018, current use of e-cigarettes was 20.8% (past 30-day use) among high school students, 7.6% (everyday/someday use) among adults aged 18\u201324 years, and 3.2% (everyday/someday use) among adults aged \u226518 years  entered the U.S. marketplace in 2007, the landscape has evolved to include disposable e-cigarettes and rechargeable e-cigarettes with prefilled cartridges and flavored e-liquids .During September 2014\u2013May 2020, e-cigarette sales increased by 122.2%. Sales of prefilled cartridges increased during September 2014\u2013August 2019; since then, sales of disposable products have increased. Prefilled mint cartridge e-cigarette sales increased from September 2014 to August 2019, then decreased, as menthol sales increased during August 2019\u2013May 2020.Continued monitoring of e-cigarette sales and use is critical to inform strategies to minimize risks. As part of a comprehensive approach, such strategies could include those that address youth-appealing product innovations and flavors."}
+{"text": "This study was aimed at exploring the effect of long noncoding RNA LINC00324 (LINC00324) on gastric cancer (GC) and the potential molecular mechanisms.  The expression of LINC00324 and miR-3200-5p in GC tissues and cells was detected by qRT-PCR. LINC00324 was silenced in GC cells by transfection of si-LINC00324. Then, the proliferation, migration, and invasion of GC cells were analyzed by MTT, wound healing, and transwell assays, respectively. The interactions between LINC00324 and miR-3200-5p and between miR-3200-5p and BCAT1 were determined by a dual-luciferase reporter and/or RNA pull-down assay.  The expression of LINC00324 was upregulated in GC cells and tissues, but miR-3200-5p was downregulated. Silencing of LINC00324 inhibited the proliferation, migration, and invasion of GC cells. LINC00324 directly targeted miR-3200-5p, and miR-3200-5p directly targeted BCAT1. si-LINC00324 negatively regulated BCAT1 expression via binding to miR-3200-5p. Furthermore, silencing of LINC00324 reversed the promoting effects of BCAT1 on the proliferation, migration, and invasion of GC cells.  Silencing of LINC00324 inhibited the proliferation, migration, and invasion of GC cells through regulating the miR-3200-5p/BCAT1 axis. Gastric cancer (GC), also known as stomach cancer, is the third most frequent malignancy worldwide . Less thLong noncoding RNAs (lncRNAs) are noncoding RNA transcripts, taking part in the occurrence and progression of diverse digestive organ cancers including esophageal cancer , pancreaMicroRNAs (miRNAs) are small noncoding RNAs which mediate target genes by interacting with the 3\u2032 untranslated regions (3\u2032 UTRs) . Some miBranched-chain amino acid transaminase 1 (BCAT1) is a cytosolic BCAT isozyme expressed in specific tissues . BCAT1 tHere, the effects of LINC00324 on the proliferation, migration, and invasion of GC cells were evaluated. Then, we investigated whether miR-3200-5p is a downstream target of LINC00324 and the association between miR-3200-5p and BCAT1. This study may reveal a hopeful therapeutic target for GC and the underlying mechanisms for the treatment of GC.In total, 60 GC patients  were collected from our hospital between April 2016 and May 2018. Fresh GC tissues and adjacent normal tissues were obtained from GC patients who underwent surgery at our hospital. The tissues were frozen in liquid nitrogen immediately after resection and stored at -80\u00b0C for future use. This study was permitted by the ethics committee of Taian City Central Hospital, and written informed consent was obtained from all the patients.2.Three human GC cell lines, AGS, MGC803, and MKN-45, and the human normal gastric epithelial cell line GES-1 were obtained from the American Type Culture Collection. Cells were cultured in Dulbecco's Modified Eagle's Medium  containing 10% fetal bovine serum  at 37\u00b0C with 5% COThe siRNA-negative control (si-NC), si-LINC00324-1, si-LINC00324-2, mimics NC, miR-3200-5p mimics, miR-3200-5p inhibitor, inhibitor NC, pcDNA3.1-NC (pcDNA-NC), and pcDNA3.1 BCAT1 (pcDNA-BCAT1) were purchased from Shanghai GenePharma . MGC803 and MKN-45 cells grown to 80% confluence were transfected with these above agents using the Lipofectamine 3000 reagent . MGC803 and MKN-45 cells were divided into si-NC, si-LINC00324-1, si-LINC00324-2, mimics NC, miR-3200-5p mimics, inhibitor NC, miR-3200-5p inhibitor, si-NC+pcDNA-NC, si-NC+pcDNA-BCAT1, and si-LINC00324+pcDNA-BCAT1 groups. Cells without transfection were considered the control or blank groups.\u03b2-actin were used for the normalization of LINC00324, miR-3200-5p, and BCAT1, respectively. The primer sequences are shown in \u0394\u0394Ct- method.Total RNA was extracted from tissues and cells using the TRIzol reagent . RNA was quantified with a NanoDrop ND-1000 spectrophotometer . A total of 300\u2009ng RNA was reverse-transcribed into cDNA using a PrimeScript RT reagent kit . PCR reaction was performed on the ABI 7500HT Fast Real-Time PCR System  with the following conditions: 95\u00b0C for 3\u2009min and 40 cycles of 95\u00b0C for 15\u2009s and 60\u00b0C for 30\u2009s. GAPDH, U6, and 3 cells/well). After 24, 48, 72, and 96\u2009h of culturing, 20\u2009\u03bcL MTT  was added into each well, and the cells were incubated for 4\u2009h at 37\u00b0C with 5% CO2. After the supernatant was discarded, 150\u2009\u03bcL dimethyl sulfoxide  was added to dissolve the formazan. The absorbance at 450\u2009nm was measured using a microplate reader .MGC803 and MKN-45 cells were seeded into 96-well plates . The wound healing rate was calculated by the fraction of cell coverage across the line.When MGC803 and MKN-45 cells were cultured at 80% confluence, an artificial scratch was created using a 10\u2009\u03bcm pore size) . MGC803 and MKN-45 cells (2 \u00d7 105) in serum-free medium were added to the upper chamber precoated with Matrigel . Roswell Park Memorial Institute- (RPMI-) 1640  medium with 10% FBS (Invitrogen) was added to the lower chamber. After 48\u2009h of incubation at 37\u00b0C, cells were removed from the upper chamber with a cotton swab, and cells in the lower chamber were fixed in methanol and stained with 0.5% crystal violet for 2\u2009min. Five random fields were photographed.The transwell assay was used to determine the cell invasion using a transwell chamber . The vectors were then cotransfected with mimics NC, miR-3200-5p mimics, and miR-3200-5p mimics+LINC00324 into MGC803 and MKN-45 cells using Lipofectamine 3000 (Invitrogen) for 48\u2009h. The luciferase activity was detected by a dual-luciferase reporter gene assay system (Promega).Bio-LINC00324-Wt, Bio-LINC00324-Mut, and Bio-NC (GenePharma) were transfected into GC cells (MGC803 and MKN-45) using Lipofectamine 3000 (Invitrogen). After culturing for 48\u2009h, cells were incubated with Dynabeads M-280 streptavidin beads  for 1\u2009h. The qRT-PCR assay was employed to assess the enrichment of BCAT1.t-test. The correlation was determined by the Pearson correlation analysis. A P value < 0.05 was considered statistically significant.Statistical analysis was performed using SPSS 23.0 . Data were presented as mean \u00b1 standard\u2009deviation (SD). The differences among multiple groups were analyzed by one-way ANOVA followed by Tukey's multiple comparison test. The differences between two groups were assessed using Student's P < 0.001) (P < 0.01) (P < 0.05), lymphoma metastasis (P < 0.01), and TNM stage in GC patients (P < 0.01). However, LINC00324 expression had no connection with age, gender, and histological grade in GC patients (P > 0.05) (P < 0.01)  . Additio < 0.01) . The cor > 0.05) . qRT-PCR < 0.01) . MGC803 P < 0.01)  (P < 0.01) (P < 0.01)  . si-LINC < 0.05) . The wou < 0.01) . The tra < 0.01) .P < 0.01) (P < 0.01) (P < 0.01) (P < 0.001)  (P < 0.01)  . The dua < 0.01) . In addi < 0.01) . The exp< 0.001) . There w < 0.01) . qRT-PCR < 0.01) .P < 0.01), and overexpression of LINC00324 reversed the reducing effect of miR-3200-5p mimics on the relative luciferase activity (P < 0.05) (P < 0.01) (P < 0.01) (P < 0.001)   (The downstream target of miR-3200-5p was predicted by TargetScan. A binding site of miR-3200-5p on the 3\u2032 UTR of BCAT1 was predicted . Subsequ < 0.05) . The RNA < 0.01) . In addi < 0.01) . These r< 0.001) . There w < 0.01)  and a po < 0.05) .P < 0.01). The expression of BCAT1 in the si-LINC00324+pcDNA-BCAT1 group was markedly decreased compared with that in the si-NC+pcDNA-BCAT1 group (P < 0.01)  and 72\u2009h (P < 0.01) postculturing was markedly increased compared with that in the si-NC+pcDNA-NC group. Silencing of LINC00324 markedly reversed the promoting effect of BCAT1 on the proliferation of MGC803 cells at 72\u2009h postculturing (P < 0.01) (P < 0.01). Silencing of LINC00324 markedly reversed the promoting effects of BCAT1 on the migration and invasion of MGC803 cells (P < 0.01)  . Rescue  < 0.01) . Wound h Figures .Abnormal expression of lncRNAs is related to the tumorigenesis of GC , 21. SomPrevious researches have confirmed that LINC00324 takes part in cell proliferation, invasion, and migration in cancer. Wu et al. have proven that LINC00324 increases the proliferation and migration of osteosarcoma cells via targeting WDR66 . Pan et lncRNAs can serve as molecular sponges to competitively regulate miRNAs in GC. For instance, lncRNA-RMRP promotes cell proliferation by acting as a sponge of miR-206 in GC . lncRNA The expression of BCAT1 is usually upregulated in diverse cancers, such as ovarian cancer , breast In conclusion, the expression of LINC00324 was upregulated in GC tissues and cells. Silencing of LINC00324 inhibited the proliferation, migration, and invasion of GC cells through regulating the miR-3200-5p/BCAT1 axis. Our research indicated that LINC00324 might act as a potential therapeutic target for GC. However, the detailed action mechanisms of LINC00324 on GC remain limited, and further researches are still needed."}
+{"text": "Orthodontic management of a non-syndromic patient with concomitant bimaxillary hypohyperdontia: a case report. Dental Press J. Orthod. [Internet]. 2020 Jan [cited 2020 Apr 15] ; 25( 1 ): 36-46.How to cite this article\u201d: Now the article should have the following \u201cEi Hsu Hlaing Ei, Ishihara Yoshihito, Fujisawa Atsuro, Yamashiro Takashi, Kamioka Hiroshi. Orthodontic management of a non-syndromic patient with concomitant bimaxillary hypohyperdontia: a case report. Dental Press J. Orthod. [Internet]. 2020 Jan [cited 2020 Apr 15] ; 25( 1 ): 36-46."}
+{"text": "Azadirachta indica, Azadirachta excelsa, Azadirachta siamens, Melia azedarach, Melia toosendan, and Melia volkensii, have been subject to botanical pesticide evaluation. This review focuses on Melia volkensii, which has not been intensively studied. M. volkensii, a dryland tree species native to East Africa, has shown activity towards a broad range of insect orders, including dipterans, lepidopterans and coleopterans. Its extracts have been reported to have growth inhibiting and antifeedant properties against Schistocerca gregaria, Trichoplusia ni, Pseudaletia unipuncta, Epilachna varivestis, Nezara viridula, several Spodoptera species and other insect pests. Mortality in mosquitoes has also been reported. Several limonoids with a wide range of biological activities have been isolated from the plant, including volkensin, salannin, toosendanin, trichilin-class limonoids, volkendousin, kulactone among others. This paper presents a concise review of published information on the phytochemical composition and potential of M. volkensii for application in insect pest management.Due to potential health and environmental risks of synthetic pesticides, coupled with their non-selectivity and pest resistance, there has been increasing demand for safer and biodegradable alternatives for insect pest management. Botanical pesticides have emerged as a promising alternative due to their non-persistence, high selectivity, and low mammalian toxicity. Six Meliaceae plant species,  The continuous and indiscriminate use of synthetic pesticides in crop protection has led to an increase in pest resistance, health and environmental concerns . This haMeliaceae plant family has been reported to produce a wide range of compounds, including flavonoids, chromones, coumarins, benzofurans, mono-, sesqui-, di-, and triterpenoids, but tetranortriterpenoids with a \u03b2-substituted furanyl ring at C17\u03b1 are the best known for the production of limonoids [Azadirachta indica A. Juss (Indian neem tree) is reported as a potential plant for the control of vector mosquitoes [Melia azedarach Linnaeus (chinaberry) has been reported outlining its use in folk medicine having antifertility, antiviral, cytotoxic, antibacterial, immunomodulatory, repellent, antifeedant, antilithic and anthelmintic activity from various parts of the plant [A. indica has reported its use in agriculture for application as manure, fertilizer, soil conditioner, fumigant, and as botanical pesticide [Melia volkensii (Gurke) has also been identified as one of the pesticidal plants in Africa [The imonoids . Limonoiimonoids . Alkaloiimonoids . Reviewssquitoes . Reviewssquitoes ,7. A phyhe plant ,9. A revesticide . Melia vn Africa . Anothern Africa . Detailen Africa .A. indica (Indian neem tree), Azadirachta excelsa Jack (Philippine neem tree), Azadirachta siamens Valeton (Siamese neem tree), M. azedarach (chinaberry), Melia toosendan Siebold and Zucc., and M. volkensii [A. indica (neem tree) and M. azedarach (chinaberry) [A. indica, have been effective growth regulators and feeding deterrents for a wide range of insect species [M. azedarach have also shown antifeedant activity against the juvenile and adult Xanthogaleruca luteola Muller (elm leaf beetles) and mortality against its larvae [M. azedarach are also effective against Napomyza lateralis Fallen (agromyzid leafminers) and Trialeurodes vaporariorum Westwood (whiteflies) [M. azedarach which has been commercialized in China, is an effective growth inhibitor against Ostrinia nubilalis H\u00fcbner (European corn borer), effective repellent against Pieris brassicae Linnaeus (cabbage moth) and an oviposition deterrent against Trichoplusia ni H\u00fcbner (cabbage looper) [Several plant species of the Meliaceae have shown promising bioactivity against a variety of insects . Their iolkensii . Howevernaberry) . Azadira species . Azadira species . Extracts larvae . Fruit eteflies) . Toosend looper) . Toosend looper) .M. volkensii, a dryland tree species native to East Africa has, however, not been intensively studied [M. volkensii. These include: 1-O-cinnamoyltrichilin, meliavolkinin, 1,3-diacetylvilasinin, meliavolkin, volkensin, volkensinin, 12\u03b2- and 6\u03b2-hydroxykulactone, meliavolkenin, meliavolin, meliavolen, melianinone, meliavolkensin A and B, melianin C, (E)- and (Z)-volkendousin, meliavosin, 2-9-epoxymeliavosin [M. volkensii seed kernel extracts have more insect growth inhibitory and acute lethal toxicity than azadirachtin-containing fractions from neem seed kernel extracts [M. volkensii dried fruit powder and residual fruit cake obtained after extraction with ethanol are used as goat feed, their growth and performance are not negatively affected, indicating that both fruit powder and its cake could be used as safe ruminant feed supplement [M. volkensii have also traditionally been used to control ticks and fleas in goats [M. volkensii offers a key indigenous tree species that can be used to mitigate against desertification in arid and semi-arid lands [M. volkensii in insect pest management. studied . It is a studied . The tre studied . It rema studied . Severalextracts . It has pplement . Its usepplement , and trapplement . Aqueousin goats . M. volkid lands , while aid lands . This paM. volkensii have been reported to pose activity towards a broad range of insect orders including Diptera, Lepidoptera, Coleoptera among others [Schistocerca gregaria Forsk. (desert locusts) [S. gregaria when seed extract was applied to their preferred host plants mainly Schouwia thebaica Webb, Fagonia olivieri DC (fagonbush plant) and Hyoscyamus muticus Linnaeus (Egyptian henbane) in a field trial experiment [M. volkensii extracts could affect hormone levels in S. gregaria larvae [S. gregaria, 80% mortality was recorded 48 hours after injection with crude ethanolic and methanolic extracts at a concentration of 30 \u03bcg/g of the insect [S. gregaria, M. volkensii and neem oil have been reported to cause mortality of up to 91% and 92%, respectively, after 14 days in a comparative study [Crude fruit extracts from g others  as shownlocusts) . Repelleperiment . Althouga larvae . In fifte insect . When spve study . In contve study . Trichoplusia ni H\u00fcbner (cabbage looper) and Pseudaletia unipuncta Haworth (true armyworm) [T. ni (dietary EC50 = 7.6 ppm) and feeding deterrent (DC50 = 0.9 \u03bcg/cm2) [M. volkensii extracts has been observed to lead to a decrease in antifeedant response when tested against T. ni implying that the insect could develop tolerance to the extracts [Plutella xylostella Linnaeus (diamondback moth) and P. unipuncta, there was no significant decrease in feeding deterrent response to the extracts following continuous exposure [M. volkensii are responsible for the insecticidal activity in T. ni [M. volkensii extracts, other Meliaceae plant extracts  and commercial botanical insecticides  when tested against T. ni and P. unipuncta [Antifeedant and larval growth inhibitory effects of fruit extracts have been observed in rmyworm) ,28. Crud \u03bcg/cm2) . Prolongextracts . Howeverexposure . It has in T. ni . Comparanipuncta . M. volkensii fruit extracts when tested at concentrations ranging from 1 to 50 \u03bcg/\u03bcL showed feeding deterrence, growth disruption and mortality against Nezara viridula Linnaeus (stink bug), a polyphagous pest which attacks a variety of crops, including nuts, corn, cotton, grains and tomatoes [N. viridula [N. viridula when exposed to fruit extracts, with 10 \u03bcg/\u03bcL causing malformation in up to 85.70% of surviving adults [Coranus arenaceus Walker even though there were no deformities in resultant adults after topical application of the M. volkensii extracts at 1, 5, and 10 \u03bcg/\u03bcL [tomatoes . The disviridula . Furtherg adults . A delay10 \u03bcg/\u03bcL . M. volkensii fruit extract showed potent antifeedant properties against Epilachna varivestis Mulsant (Mexican bean beetle) [P. unipuncta (dietary EC50 = 12.5 ppm) with refined seed extracts to the leaf discs in a choice bioassay [P. unipuncta and P. xylostella, and adults of E. varivestis  [M. volkensii seed extracts have been recorded to have stronger antifeedant activity compared to pure allelochemicals: digitoxin, cymarin, xanthotoxin, toosendanin, thymol and trans-anethole against P. unipuncta, P. xylostella and E. varivestis [Spodoptera litura Fabricius, neem, rotenone, M. volkensii extract, toosendanin, Annona squamosal L. extract and pyrethrum at 1% concentration recorded larval growth (% relative to control) of 4.1, 97.5, 26.2, 48.3, 61.4, and 56.6%, respectively after 96 h in a comparative study [When applied to cabbage leaf disks in a choice bioassay,  beetle) . Growth bioassay . The seectively) . In factrivestis . When apM. volkensii fruit extracts have shown growth-inhibiting activity against Aedes aegypti Linnaeus (yellow fever mosquito) larvae at 2 \u03bcg/mL in water, whilst recording high mortality during the molting and melanization process with LC50 of 50 \u03bcg/mL in 48 h [A. aegypti could be a result of synergistic effects of a plethora of limonoid compounds or a single active compound exerting these effects [Dried  in 48 h . At a hi in 48 h . Growth  effects . M. volkensii fruit kernel extract showed growth-inhibiting activity against Anopheles arabiensis Giles with an LC50 of 5.4 \u03bcg/mL in 48 h [50 of 34.72 \u03bcg/mL in 48 h) and oviposition deterrence was observed in second-instar larvae of Culex quinquefasciatus Say (Southern house mosquito) when treated with refined methanolic fruit extracts [M. volkensii fruit acetone extract showed S- and U-shaped postures and frequent stretching in C. quinquefasciatus; such postures and stretching are a characteristic of mosquito larvae reared in M. volkensii fruit extract [C. quinquefasciatus within 36 h [M. volkensii seeds have recorded growth inhibitory effects and equal toxicity (LD50 of 30 \u03bcg/mL) for larvae and pupae of C. pipiens f. molestus Forsk\u00e5l (London underground mosquito) [M. azedarach seed extracts recorded lower toxicity (LD50 of 40 \u03bcg/mL) while pure azadirachtin A recorded higher toxicity (LD50 of 1\u20135 \u03bcg/mL) against C. pipiens when compared with M. volkensii extracts [M. volkensii may indicate the presence of saponins as toxic principles thus making it an interesting candidate for application against aquatic insects such as mosquitoes and other vectors of diseases [A column chromatography-purified fraction of  in 48 h . Mortaliextracts . The graosquito) . M. azedextracts . The watdiseases .M. volkensii, as shown in M. volkensii. This indicates that insect control bioactivity is, therefore, based on other compounds than azadirachtin [M. volkensii fruit is more lipophilic than azadirachtin [Insect antifeedants have been found in major classes of secondary metabolites\u2014alkaloids, phenolics, and terpenoids . Howeverirachtin . It is pirachtin . Botanicirachtin .1) and salannin (2) have been isolated from seed extracts of M. volkensii [1) and salannin (2) were isolated from the whole fruits of M. volkensii [1) has shown antifeedant activity against Spodoptera frugiperda Smith  larvae with an ED50 of 3.5 \u03bcg/cm2 [2) has also shown antifeedant activity against insect pests such as Acalymma vittata Fabricius (striped cucumber beetle), Musca domestica Linnaeus (housefly), Epilachna varivestis Mulsant (Mexican bean beetle), Heliothis virescens Fabricius (tobacco budworm), S. frugiperda and Spodoptera littoralis Boisduval (cotton leafworm) [2) has also been reported to cause feeding suppression against larvae of Earias insulana Boisduval (Egyptian stemborer), weight reduction (59%\u201389%) in Cnaphalocrocis medinalis Guenee (rice leafroller) and reduction in activities of acid phosphatases (ACP), alkaline phosphatases (ALP) and adenosine triphosphatases (ATPase), implying that gut enzyme activities were affected. 2\u2019,3\u2019-Dihydrosalannin (3), 1-detigloyl-1-isobutylsalannin (4) and 1\u03b1,3\u03b1-diacetylvilasinin (5) have also been isolated from the plant [The insect antifeedants volkensin  . Salannihe plant .M. volkensii seed extracts, extracted in cold water, have been reported to contain unsaturated fatty acids (oleic acid (6), linoleic acid (7) and gadoleic acid (8)) and saturated fatty acids , stearic acid (10) and arachidic acid (11)) as shown in 6), linoleic acid (7), linolenic acid, and ricinoleic acid have enhanced insecticidal activity of organophosphates and carbamates when applied against sucking insects and defoliating insects [ insects . M. volkensii are shown in 12), which has been isolated from the root bark of M. volkensii [O. nubilalis, an effective repellent against P. brassicae and an oviposition deterrent against T. ni [13) has shown antifeedant activity towards S. littoralis having minimum antifeedant concentration (MAC) value of 1000 mg/L and a significant antibacterial activity against Porphyromonas gingivalis ATCC 33277 with minimum inhibitory concentration (MIC) value of 15.6 \u03bcg/mL [14) has been reported to have antifeedant activity against several Spodoptera species  [13, 1-acetyltrichilinin 15, 1-tigloyltrichilinin 16) were tested against Spodoptera eridania Stoll (Southern armyworm) where the 12\u03b1-OH function was the most potent, followed by 12\u03b2-OH, 12-desoxy, and 12\u03b1-acetoxy groups in order of decreasing potency [13, 15, 16) [17, which has weak activity with marginally significant selectivity for breast cancer cell line (MCF-7), has also been isolated from the root bark of M. volkensii [18), isolated from methanolic extract of M. volkensii fruits [19), both insect antifeedants have also been reported [Other chemical compounds that have been isolated from various parts of olkensii , has beest T. ni . 1-Cinnaolkensii . Ohchinii fruits , and melreported . M. volkensii with different biological activities. These include volkensinin, as isolated from ethanolic extracts of M. volkensii root bark [50 = 57 \u03bcg/mL) and weak cytotoxicity against six human tumor cell lines with ED50 values of 27.90, 28.35, 33.56, 29.55, 8.43, and 28.51 \u03bcg/mL in A-498 (human kidney carcinoma), PC-3 (prostate adenocarcinoma), PACA-2 (pancreatic carcinoma), A-549 (human lung carcinoma), MCF-7 (human breast carcinoma), and HT-29 (human colon adenocarcinoma), respectively [Escherichia coli Migula and Aspergillus niger Tiegh. with respective minimum inhibitory concentration (MIC) values of 12.5 and 6.25 \u03bcg/mL [M. volkensii, showed cytotoxicity against six human solid tumor cell lines: A-549, MCF-7, HT-29, A-498, PACA-2, and PC-3 [M. volkensii root bark led to the isolation of meliavolkenin which showed moderate cytotoxicity against three human tumor cell lines with a respective ED50 value of 10.33 \u03bcg/mL, 4.30 \u03bcg/mL, and 0.67 \u03bcg/mL in A-549, MCF-7, and HT-29 cells [M. volkensii [50 values of 0.49 \u03bcg/mL and 0.25 \u03bcg/mL, respectively [E)-volkendousin, isolated from M. volkensii root bark, also showed activity against six human tumor cell lines  [50 of 11.25 \u03bcg/mL, 0.57 \u03bcg/mL and 6.65 \u03bcg/mL in A-549, MCF-7 and HT-29 cells, respectively [M. volkensii root bark following activity-directed fractionation with brine shrimp test [M. volkensii and exhibited significant activity against E. coli and A. niger with a respective minimum inhibitory concentration (MIC) value of 12.5 and 6.25 \u03bcg/mL [Mycobacterium tuberculosis Zopf resulted in the isolation of 12\u03b2-hydroxykulactone, 6\u03b2-hydroxykulactone and kulonate from M. volkensii seeds with MIC values of 16 \u03bcg/mL, 4 \u03bcg/mL, and 16 \u03bcg/mL, respectively [50 = 0.57 \u03bcg/mL), MCF-7 (ED50 = 0.26 \u03bcg/mL), and HT-29 (ED50 = 0.12 \u03bcg/mL) [M. volkensii include 3-episapelin, meliavolen, melianinone [M. volkensii include scopoletin [Other compounds have also been isolated from oot bark , which sectively . Toosend25 \u03bcg/mL . Melianiand PC-3 . Bioacti29 cells . The bioolkensii , have shectively . (E)-volnd PC-3) . Meliavo25 \u03bcg/mL . Bioactiopoletin , melianiM. volkensii have proved to be effective insect antifeedants and growth inhibitors. Extensive research has been done on mosquito control using M. volkensii; however, more research needs to be done on insect pests of agricultural importance. M. volkensii has no reported adverse effect on the environment or mammals, making it a potential botanical pesticide for the biosafe application in integrated pest management. The availability of renewable resources from the tree, such as fruits, stem bark, and leaves makes this plant a potential candidate for insect control with minimal interference on the plant. In this regard, M. volkensii could be further exploited as a source of natural insecticide. Extracts and pure compounds isolated from"}
+{"text": "Escherichia coli isolate INF13/18/A, which was isolated from Universiti Sains Malaysia (USM) Hospital. This isolate was identified as an extended-spectrum \u03b2-lactamase-producing Escherichia coli strain harboring the antimicrobial resistance genes TEM, CTX-M-1, and CTX-M-9.We describe here the draft genome sequence and basic characteristics of  Escherichia coli isolate INF13/18/A, which was isolated from Universiti Sains Malaysia (USM) Hospital. This isolate was identified as an extended-spectrum \u03b2-lactamase-producing Escherichia coli strain harboring the antimicrobial resistance genes TEM, CTX-M-1, and CTX-M-9.We describe here the draft genome sequence and basic characteristics of  Escherichia coli is one of the most versatile and widely studied species in the world  Hospital.Gram-negative he world . The emehe world . The precrobials . In thisThe clinical sample was collected aseptically and incubated into the Bactec 9240 blood culture system . Gram staining and biochemical testing . The isoE. coli , with read-trimming and filtered read length threshold values of 30 and 50\u2009bp, respectively. The filtered and trimmed reads  were assembled de novo using SPAdes v3.9.0  (E. coli isolates. The complete genome sequences of E. coli O157 strain AR-0427 (GenBank accession no. CP044148.1) and E. coli strain AR216.2b (GenBank accession no. CP043942), which were the top two hits, were selected to scaffold the draft genome using a graph-based approach with Medusa v1.6  v4.10  (usa v1.6 . The assP) v4.10 , which pEscherichia coli INF/13/18/A has been deposited in GenBank under the accession no. WRPL01000000 (BioProject accession no. PRJNA593306 and BioSample accession no. SAMN13474918). The raw reads are available under the SRA accession no. SRR10587526.The whole-genome project for the ESBL-producing strain"}
+{"text": "Human immunodeficiency virus (HIV) is associated with co-morbid affective and stress-sensitive neuropsychiatric disorders that may be related to dysfunction of the hypothalamic-pituitary-adrenal (HPA) stress axis. The HPA axis is perturbed in up to 46% of HIV patients, but the mechanisms are not known. The neurotoxic HIV-1 regulatory protein, trans-activator of transcription (Tat), may contribute. We hypothesized that HPA dysregulation may contribute to Tat-mediated interactions with oxycodone, a clinically-used opioid often prescribed to HIV patients. In transgenic male mice, Tat expression produced significantly higher basal corticosterone levels with adrenal insufficiency in response to a natural stressor or pharmacological blockade of HPA feedback, recapitulating the clinical phenotype. On acute exposure, HIV-1 Tat interacted with oxycodone to potentiate psychomotor and anxiety like-behavior in an open field and light-dark transition tasks, whereas repeated exposure sensitized stress-related psychomotor behavior and the HPA stress response. Pharmacological blockade of glucocorticoid receptors (GR) partially-restored the stress response and decreased oxycodone-mediated psychomotor behavior in Tat-expressing mice, implicating GR in these effects. Blocking corticotrophin-releasing factor (CRF) receptors reduced anxiety-like behavior in mice that were exposed to oxycodone. Together, these effects support the notion that Tat exposure can dysregulate the HPA axis, potentially raising vulnerability to stress-related substance use and affective disorders. Human immunodeficiency virus type 1 (HIV-1) continues to be a serious health concern with over 1 million infected individuals in the U.S. . HoweverA common, but under-investigated neurological complication of HIV involves disruption of the hypothalamic-pituitary-adrenal (HPA) stress axis. Despite treatment with cART, 14\u201346% of HIV+ patients demonstrate a dysregulated HPA axis characterized by elevated basal glucocorticoids with seemingly paradoxical adrenal insufficiency in response to a stressor ,8,9. ComThe mechanisms of HIV-mediated HPA dysregulation are not known but may involve neurotoxic HIV-1 proteins. One important therapeutic target that may contribute to these effects is the regulatory protein, trans-activator of transcription (Tat). HIV-1 Tat exerts direct effects on neurons to promote excitotoxic injury and activates monocyte-derived cells (and astrocytes to a lesser extent) to promote neuroinflammation via cytokine production . We haveIn the present study, we hypothesized that HIV-1 Tat and/or oxycodone would interact to promote HPA axis dysregulation, potentiating oxycodone-mediated effects including psychomotor stimulation and affective dysfunction in a conditionally-inducible Tat transgenic mouse model. We further expected pharmacological blockade of GR and/or CRF receptors to restore HPA function and to ameliorate behavioral deficits.In Experiment 1, Tat-tg male mice had HIV-1 Tat expression induced (or not) via doxycycline administration for five days. After two days of doxycycline washout (to limit non-specific anti-inflammatory effects), mice were (or were not) exposed to a 15-min swim stress. Following stress, mice were acutely administered an injection of saline  or oxycodone  and psychomotor and anxiety-like behavior were assessed 15 min later A.F = 5.00, p < 0.05]  = 5.00, p < 0.05]  = 5.04, p < 0.05]  = 27.16, p < 0.05]  = 27.34, p < 0.05] of travel and decreased the frequency  and time spent rearing , compared to saline administration  = 8.69, p < 0.05] and made fewer transitions between compartments  compared to Tat(\u2212) controls  = 5.15, p < 0.05]  controls, Tat(+) mice traveled a significantly greater distance [ < 0.05] B; see *  < 0.05]  in an op < 0.05] . Irrespe < 0.05] A; see \u2020 stration . In a licontrols . No signcontrols . When ci < 0.05] C. Tat(+)F = 13.83, p < 0.05] and speed  of travel; however, no differences were observed between Tat(\u2212) and Tat(+) mice  = 13.88, p < 0.05]  mice D, nor we(+) mice . Similar(+) mice . As expe < 0.05] E; see *.While the initial opioid response is indicative of later abuse liability, many HIV+ patients are prescribed oxycodone and are exposed repeatedly. How repeated oxycodone exposure modifies the HPA stress response in Tat-exposed mice was of interest. In Experiment 2, Tat-tg male mice had HIV-1 Tat expression induced (or not) via doxycycline administration for five days (with two days of washout). During this time, mice received daily injections of saline  or oxycodone . Mice were (or were not) exposed to a 15-min swim stress prior to testing A.F = 46.98, p < 0.05]  = 46.68, p < 0.05]  = 6.08, p < 0.05]  = 11.18, p < 0.05]  = 4.05, p = 0.05]; no such effect was observed on Tat(\u2212) controls  = 3.99, p = 0.05]  = 4.93, p = 0.05]  = 14.79, p < 0.05]  mice spent significantly less time in the brightly-lit compartment, compared to Tat(\u2212) mice [ = 0.05] . Irrespe = 0.05] . Tat(+)  < 0.05] C; see *.F = 7.01, p < 0.05]  = 7.10, p < 0.05]  = 3.86, p = 0.05]  = 9.60, p < 0.05]  mice. As seen in Experiment 1, overall motor behavior was reduced following swim stress compared to that observed in non-stressed mice. Repeated oxycodone and genotype interacted such that oxycodone\u2013administered Tat(+) mice traveled a significantly greater distance [ < 0.05] D; see \u2021  < 0.05]  than did = 0.05] . No diff = 0.05] . As obse < 0.05] E; see \u00a7.In order to begin identifying the important aspects of the HPA axis that are involved in the psychomotor, anxiety-like, and glucocorticoid response to oxycodone and Tat, mice were administered glucocorticoid receptor (GR) and/or corticotrophin-releasing factor-receptor (CRF-R) inhibitors concurrent with doxycycline administration A. To bloF = 2.86, p < 0.05] and speed  traveled. As before, oxycodone significantly increased the distance . All Tat(+) mice administered a GR and/or CRF-R inhibitor demonstrated a modest, but significant, reduction in the distance  mice  = 11.54, p < 0.05]. Pretreatment with RU-486  produced a significant and large increase in circulating corticosterone among Tat(\u2212) mice  = 6.11, p < 0.05]. Irrespective of genotype, at t5 oxycodone-administered mice exhibited significantly lower circulating corticosterone compared to saline-administered mice  and promoting adrenal insufficiency in response to HPA activation to a natural or pharmacological challenge. HPA dysregulation occurred concurrent with anxiety-like behavior and combined Tat and oxycodone-mediated psychostimulation. Repeated oxycodone exposure was able to sensitize the Tat-suppressed HPA response, suggesting potential for pharmacological therapeutic intervention. Blocking GR attenuated combined Tat- and oxycodone-mediated psychostimulation and produced a proportional increase in corticosterone, indicating its importance in these effects. Thus, therapeutics with the capacity to restore the HPA axis may be important mediators of HIV-1 Tat/opioid interactions.Procedures were preapproved by the Institutional Animal Care and Use Committee (IACUC) at the University of Mississippi . All experiments were carried in accordance with ethical guidelines defined by the National Institutes of Health (NIH Publication No. 85-23).n = 324) were bred in the vivarium at the University of Mississippi . Mice were housed 2\u20135 per cage and be kept in a temperature-and humidity-controlled environment on a 12:12 h light:dark cycle (lights off at 09:00 h) with ad libitum access to food and water. HIV-1 Tat1\u201386 is conditionally expressed by administration of doxycycline as described previously  via the administration of doxycycline QD for 5 days i.p. Given that doxycycline can attenuate neuroinflammation and could therefore mask potential effects of Tat, 2 days of doxycycline washout was carried out . All behn = 8\u20139), Tat(+)/saline (n = 7\u20138), Tat(\u2212)/oxycodone (n = 12), Tat(+)/oxycodone (n = 8\u20139), and the following swim-stressed groups were yielded: Tat(\u2212)/saline (n = 8), Tat(+)/saline (n = 9), Tat(\u2212)/oxycodone (n = 8), Tat(+) oxycodone (n = 9).To begin to determine the HPA-axis interactions involved in exposure to HIV-1 Tat and acute oxycodone, mice were randomly-assigned to undergo a 15-min swim stress (or not) followed by administration of vehicle  or oxycodone  only once prior to behavioral testing. Fifteen minutes after drug administration, mice were assessed in an open field to determine their psychomotor response followed immediately by assessment in a light-dark transition test to determine anxiety-like behavior A. Acrossn = 7\u20138), Tat(+)/saline (n = 10), Tat(\u2212)/oxycodone (n = 8), Tat(+)/oxycodone (n = 10), and the following swim-stressed groups were yielded: Tat(\u2212)/saline (n = 8), Tat(+)/saline (n = 8), Tat(\u2212)/oxycodone (n = 8), Tat(+) oxycodone (n = 9).Given that most patients are exposed to opioids on a repeated dosing schedule, some mice were administered sterile saline (0.9%) or oxycodone (3 mg/kg) daily throughout the 7-day doxycycline-induction/washout schedule A. As befn = 8), Tat(+)/saline/vehicle (n = 7\u20138), Tat(\u2212)/oxycodone/vehicle (n = 8\u20139), Tat(+)/oxycodone/vehicle (n = 9), Tat(\u2212)/saline/antalarmin (n = 8), Tat(+)/saline/antalarmin (n = 8\u20139), Tat(\u2212)/oxycodone/antalarmin (n = 8\u20139), Tat(+)/oxycodone/antalarmin (n = 10), Tat(\u2212)/saline/RU-486 (n = 8\u20139), Tat(+)/saline/RU-486 (n = 7\u20138), Tat(\u2212)/oxycodone/RU-486 (n = 8\u20139), Tat(+)/oxycodone/RU-486 (n = 7\u20138), Tat(\u2212)/saline/antalarmin+RU-486 (n = 9), Tat(+)/saline/antalarmin+RU-486 (n = 8), Tat(\u2212)/oxycodone/antalarmin+RU-486 (n = 9\u201310), Tat(+)/oxycodone/antalarmin+RU-486 (n = 9\u201310).To begin to identify the important receptor sites involved in HIV-1 Tat- or oxycodone-mediated disruption of the HPA axis, some mice were pretreated with the GR antagonist, RU-486, and/or the CRF-R antagonist, antalarmin, prior to testing. RU-486 was administered daily throughout the 7-day doxycycline-induction/washout schedule and 30 min prior to behavioral testing A. Antalan = 5/group) in order to assess the time-course for HPA activation. Prior work revealed peak HPA circulating corticosterone levels in HIV-1 Tat females within 30 min of testing  as the between-subjects factors. For Experiment 3, behavioral and steroid analyses were assessed via separate three-way ANOVAs with pretreatment , drug condition , and genotype  as the between-subjects. For Experiment 4, corticosterone steroid analyses were assessed via repeated measures ANOVA with drug condition  and genotype  as the between-subjects factors and time  as the within-subjects factor. Main effects were followed by Fisher\u2019s Protected Least Significant Difference"}
+{"text": "Neritimorpha, six Caenogastropoda, and 28 Heterobranchia). Illustrations and brief taxonomic notes/remarks are provided for every species. We also described Georrisacarinata Sutcharit & Jirapatrasilp, sp. nov. based on some distinct shell morphological characters. Since the first descriptions during the colonial period in the nineteenth century, some land snail species  have not been reported subsequently. This probably reflects a lack of knowledge concerning land snail biodiversity in this country. To our knowledge, this is the first comprehensive survey of land snails in southern Cambodia. A need for more field research and systematic revision of the land snails in this interesting region is also highlighted and demonstrated.Prior to this study, few collections and records were made of the land snails in Cambodia and the historical taxa had never been reviewed. Herein a report on the land snail diversity based on specimens collected recently from karstic and non-karstic areas in southern Cambodia is provided. This checklist presents 36 species of land snails (two  Cambodia forms a part of the Indo-Chinese sub-region of the Indo-Burma biodiversity hotspot . Its terHemiptera  and The Natural History Museum . The placement of each genus within higher order classification followsIFReDI) of the Fisheries Administration, Phnom Penh; Chulalongkorn University Museum of Zoology (CUMZ), Bangkok; Zoological Reference Collection of the Lee Kong Chian Natural History Museum, National University of Singapore (ZRC); The Natural History Museum, London (NHMUK).All the specimens were identified to genus or species level based on shell characteristics by referring to the historical literature including original descriptions, recent catalogues of land snails from Laos by Field surveys were conducted in karstic areas in southwestern Cambodia, Kampot Province. In addition, caves and cave-like chambers which provide appropriate microhabitats for karst-dwelling snails were also surveyed. This area has a monsoonal climate with wet season (May to November) and dry season (December to April). The karst landscape in Kampot is a small, isolated hill rising precipitously from the flat lowlands Fig. . The forLowland habitats of the eastern areas of Kirirom National Park are a conglomerate of hills and a plateau reaching 900 m in elevation, straddling the Kampong Speu and Koh Kong Provinces. The bulk of the plateau is covered with a mosaic of grassland and a reticulated network of pine forest plantations Fig. . The sloThe Preah Monivong Bokor National Park, Kampot Province is in the southeastern portion of the Cardamom Mountain Ranges within a range known as the Elephant Mountains. The plateau reaches an elevation of 1,100 m, and the floral composition of this range is greatly affected by continuous, monsoonal winds arising from the Gulf of Thailand. The climate promotes a mixture of grassland and tropical moist forest that shrouds the upper elevations of the Bokor Plateau in thick fog for much of the year , the conNeritimorpha . The distribution data given under each species was retrieved from the past records.A total of 180 voucher specimen lots was collected over the survey. The total of 36 species . The aperture is round to slightly ovate, with a closed umbilicus. Although the specimens from Perak have denser spiral striation , withoutG.decora M\u00f6llendorff, 1900 and G.chrysacme M\u00f6llendorff, 1900 both of which were described from \u201cTouranne\u201d , by having a conic shell with ca. ten strong spiral ribs on the last whorl. However, G.decora has an ovate conic shape with fine radial ribs on the last whorl, and G.chrysacme has an elongate conic shape with a deep and narrow suture. In addition, the shell shape of G.monterosatiana approaches the shape of G.insulaeThis species differs from Taxon classificationAnimaliaCycloneritidaHydrocenidaeSutcharit & Jirapatrasilpsp. nov.5B5771A6-B6EC-59E9-BA7E-E23BB4736AE5http://zoobank.org/CA891381-B719-4A88-B4A9-B0E36BAB121ECUMZ-CM094/1  from locality no. 11; CUMZ-CM042 (21 shells), IFReDI (10 shells), ZRC (10 shells) and NHMUK (10 shells) from locality no. 9.Holotype 4/1 Fig.  from loc10\u00b034'1.77\"N, 104\u00b028'6.13\"E).Phnom Kampong Trach Cave Temple, Kampong Trach District, Kampot Province, Cambodia, Locality no. 11 (CUMZ-CM086 (23 shells). Locality no. 17: CUMZ-CM102 (18 shells).Locality no. 12: Shell minute , conic, solid, translucent, yellowish to pale orange with darker colour on apex. Whorls 4\u00bc, last whorl large ca. two-thirds of shell height. Protoconch ca. one whorl; sculpture nearly smooth and discontinuous spiral appearing immediately after protoconch. Following whorls slightly keeled, sculptured with thin and uneven growth lines; upper periphery with irregular and strong sculpture; below periphery with discontinuous spiral ribs. Sutures angular and impressed. Aperture round to slightly ovate. Umbilicus closed. Operculum unknown.carinata represents its keeled whorls of this new species.The Latin specific name Hypselostoma spp.This new species is found from Kampot and Kep Provinces. The snails were found to live on limestone wall syntopically with other G.bocourti  described from \u201cEaux douces de Preck-Scholl. Haut M\u00e9kong\u201d , by having a conic shell with 4\u00bc whorls, which are slightly keeled and sculptured with thin and uneven growth lines without conspicuous spiral ribs. However, G.bocourti has a turriform shell with 6\u20137 whorls and sculptured with conspicuous spiral ribs (Rochebrune 1881a). Georissacarinata sp. nov. differs from G.poirieri Mabille, 1887 and G.conspicua Mabille, 1887 described from \u201cTonkin\u201d [Vietnam] in that the latter two species are larger  and has a turriform shell. In addition, G.poirieri has very thin, tight, wavy spiral ribs, while G.conspicua has uneven spiral ribs with additional protuberances unequally arranged along the longitudinal rows 262B8A2B-3FD7-5E65-B8DF-40373B15A5F2Cyclostoma (Cyclophorus) amoenum Pfeiffer, 1854[1852]: 62. Type locality: unknown.Cyclophorusamoenus : CUMZ-CM053 (21 shells), CUMZ-CM054 . Locality no. 11: CUMZ-CM071 (2 shells). The empty shells were collected from the ground among leaf litter.Locality no. 10: Cambodia and Thailand . The disCyclophorus is the genus encompassing highly variable shell morphology of both inter- and intraspecific entities. The demarcation among different species is poorly understood. Thus, both intensive and thorough revision and redescription require more effective taxonomic characters, e.g., morphometric analysis of large series of specimens and perhaps molecular phylogeny to clarify the exact species boundaries  [The mountains of Dey-Crahom (red earth)], sur la rive droite du grand fleuve [on the right bank of the great river (Mekong River)]. Cyclophorus (Eucyclophorus) paviei : CUMZ-CM119 (1 shell). Locality no. 6: CUMZ-CM176 (1 shell). Locality no. 9: CUMZ-CM036 (2 shells), CUMZ-C037 . Locality no. 12: CUMZ-CM096 (1 shell). Locality no. 16: CUMZ-CM168 (2 shells), CUMZ-CM169 (1 specimen in ethanol), CUMZ-CM170 (1 specimen in ethanol). The snails were found to live on the ground among leaf litter.Locality no. 13: Cambodia . The disCyclophoruspaviei was described from \u201cLes montagnes de Dey-Crahom\u201d, from Cambodia. It differs from C.cambodgensis Morlet, 1885, which was described from the same area in having a smaller (shell width 32 mm) conical shell, with a white-yellowish apertural lip, while C.cambodgensis has a larger (shell width 42 mm) and turbinate conic shell, with an orange to reddish apertural lip.Taxon classificationAnimaliaArchitaenioglossaCyclophoridaeB24D7F7F-272C-5CAE-BE1A-4ADB522E60F7Rhiostomabernardii Pfeiffer, 1862: 45, 46, pl. 6, fig. 5. Type locality: Siam [Thailand]. Cyclotusbernardii : Opisthoporusbernardii : CUMZ-CM043 (8 shells). Locality no. 13: CUMZ-CM127 (2 shells), CUMZ-CM118 (2 shells). Locality no. 10: CUMZ-CM062 . The snails were found to live on the ground among leaf litter.Locality no. 9: Cambodia, Laos and Thailand .Opisthoporusbernardii was originally described from \u201cSiam\u201d [Thailand], and it has been reported from Cambodia A8BA2B42-822F-538E-889B-28DC9E7AA101Cyclophorusklobukowskii Morlet, 1885[1884]: 391, 392, pl. 12, fig. 1. Type locality: Near the Kamchay rapids, around the K\u00e9bal-R\u00e9m\u00e9as cave (Kampot-Hatien road); commonly found on mountains, in forests, up to Compong-Som, and on the banks of Tap-Ch\u00e9ang. Lagocheilusklobukowskii : CUMZ-CM044 (7 shells), CUMZ-CM045 (12 specimens in ethanol). Locality no. 10: CUMZ-CM068 (3 specimens in ethanol). Locality no. 11: CUMZ-CM079 (2 shells). Locality no. 13: CUMZ-CM128 (3 shells), CM129  \u2026\u201d. We collected topotypic specimens that tend to have a variable shell colour from yellowish Fig. . This liLagocheilusklobukowskii was originally placed in the genus Cyclophorus and later was transferred to the genus Lagocheilus , and a thick calcareous, multispiral and plate-like operculum, whereas Cyclophorus has a turbinate shell, a thick and expanded lip, and a corneous multispiral operculum.ilus see . The disTaxon classificationAnimaliaArchitaenioglossaCyclophoridae614B5E69-1CA9-527D-9220-C0A030BE666ECyclophoruslandesi Morlet, 1885[1884]: 392, 393, pl. 11, fig. 5, 5a. Type locality: extr\u00e9mit\u00e9 de la cha\u00eene de \u013e\u00c9l\u00e9phant, non loin de la mer .Cyclophoruslaudesi [sic]: Lagocheiluslandesi : CUMZ-CM080 . Locality no. 12: CUMZ-CM104 (4 shells). Locality no. 14: CUMZ-CM152 , and the thick calcareous, multi-spiral and plate-like operculum characteristic of Pupina Vignard, 1829Taxon classificationAnimaliaArchitaenioglossaPupinidaeMorlet, 1883283DD709-264C-5D8D-A991-BD4F39799F14Pupinacrosseana Morlet, 1883: 108, 109, pl. 4, fig. 5. Type locality: Cambodge [Cambodia]. CUMZ-CM029 (1 shell). Locality no. 9: CUMZ-CM039 (10 shells). Locality no. 10: CUMZ-CM066 (1 shell), CUMZ-CM067 . Locality no. 13: CUMZ-CM133 (1 specimen in ethanol). Locality no. 17: CUMZ-CM142 (1 specimen in ethanol). The snails were found to live on the ground among leaf litter.Locality no. 7: Cambodia .This species was originally described from \u201cCambodge\u201d [Cambodia]. The diagnostic characters of this porcelain shell are a pupoid shell with varying shell colour from brownish to whitish, having a large, ovate last whorl ca. two-thirds of shell height. The shell has a thickened parietal callus, with a small posterior plica that is located some distance from an angular corner of aperture, which possesses a wide posterior canal. The anterior canal is a narrowly transverse slit overhung by a square and thickened columella plica. The aperture is circular with a white, thickened and slightly expanded lip.Valiguna Grimpe & Hoffmann, 1925Taxon classificationAnimaliaSystellommatophoraVeronicellidae59FA60F6-E456-561B-8B22-19E795F8A8D9Vaginulussiamensis Martens, 1867: 68, pl. 5, fig. 3. Type locality: Petshaburi .Valigunasiamensis : CUMZ-CM116  is thickened, with dark colour and scattered with brownish spots, and without median stripe. The ventral side (hyponotum) is with much lighter, pale creamy colouration, with tiny greyish spots distributed across hyponotum, and a narrow foot sole located in the middle. The foot sole is as long as and slightly narrower than the hyponotum, with pale yellowish brown colour. This slug is different from Succinea Draparnaud, 1801Taxon classificationAnimaliaStylommatophoraSuccineidaeMorelet, 1865EA06A16E-E337-58B7-B934-EF322D27ECB2Succineatenuis Morelet, 1865: 225, 226. Type locality: Cochinchina [South Vietnam]. Succineatenella Morelet, 1875: 244, pl. 12, fig. 5 [unjustified emendation].CUMZ-CM106 (2 shells), CUMZ-CM107 . Locality no. 9: CUMZ-CM087 .Hypselostomacambodjense tends to be abundant and widely distributed in several karstic hills in southern Cambodia and Vietnam B2EDA1FE-87E6-5097-B863-9DCB9ACC6E2FBulimusgracilis (?) Hutton, 1834: 84, 85, 93. Type locality: Mirzapoor; Futtehpoor Sikra; between Agra and Neemuch . (?) Allopeasgracilis [sic]: CUMZ-CM105 . The snails were found to live on the ground among leaf litter.Locality no. 12: Pantropical and subtropical .A.gracile in Cambodia. This species could be found in both natural and transformed anthropogenic habitats. This widespread and pantropical species has been introduced into many countries, including in greenhouses in temperate regions, and occurs throughout Laos, Thailand and Vietnam 3E6BACC4-C635-59E3-824E-AE4521A0DFD0Achatinafulica Bowdich, 1822: pl. 13, fig. 3. Type locality: unknown.Lissachatinafulica : CUMZ-CM065 . The snails were found to live on tree trunks and on the ground among leaf litter.Locality no. 10: Pantropical and subtropical .Angiostrongyluscantonensis . Locality no. 11: CUMZ-CM074 , CUMZ-CM076 , CUMZ-CM099 (1 shell), CUMZ-CM100 (9 specimens in ethanol). Locality no. 13: CUMZ-CM121 (21 shells), CUMZ-CM122 (1 specimen in ethanol). The snails were found to live on the ground among leaf litter in the limestone area.Locality no. 6: H.michaui , but the latter is more ovate and less oblique in shell shape. In addition, this species can be distinguished from H.pellucens , H.porrectus  and H.perlissus Vermeulen et al., 2019 by having strong and prominent radial ridges. For comparison, the latter three species have a smooth to nearly smooth shell surface, H.pellucens has an oblique-ovate shell shape, H.porrectus and H.perlissus have an oblique heliciform shell shape , CUMZ-CM156 (4 shells), CUMZ-CM157 . The snails were found to live on tree trunks and leaves.Locality no. 15: Bertiacambojiensis  by having a brownish shell, with spirally undulated surfaces, while B.cambojiensis has a smooth surface  and D.retrorsa  by having a dark brown shell, with wide angle of peripheral keels. In contrast, D.retrorsa tends to have sharp peripheral keel, D.salangana has round periphery and usually with brownish peripheral band, and both species are pale brownish in shell colour  located on the plateau of Preah Monivong Bokor National Park may be young individuals, as their shell size is relatively small compared to those of other congeners recorded from peninsular Thailand. It differs from l colour .Taxon classificationAnimaliaStylommatophoraDyakiidae9963D788-2D1B-5D3C-883F-9E6D421B1761Helixweinkauffiana Crosse & Fischer, 1863: 350, 351. Type locality: Cochinchine [Southern Vietnam].Quantulaweinkauffiana : CUMZ-CM002 (8 shells). Locality no. 2: CUMZ-CM006 (10 shells). Locality no. 5: CUMZ-CM011 (2 shells). Locality no. 7: CUMZ-CM013 (82 shells), CUMZ-CM014 (1 shell), CUMZ-CM015 , CUMZ-CM035 (3 shells). Locality no. 10 CUMZ-CM052 (1 shell). Locality no. 12: CUMZ-CM093 (5 shells). Locality no. 13: CUMZ-CM120 (5 shells). Locality no. 17: CUMZ-CM135 (3 shells). Locality no. 18: CUMZ-CM143 (3 shells), CUMZ-CM144 (1 shell). Locality no. 16: CUMZ-CM166 (4 shells), CUMZ-CM177 (1 shell). Locality no. 6: CUMZ-CM174 (9 shells), CUMZ-CM175 EDC62306-0411-5BDB-93D2-C99EEEB8A0C5Helixpaviei Morlet, 1885[1884]: 386, 387, pl. 11, fig. 1, 1a. Type locality: dans les for\u00eats, entre Kampot et Phnom-Penh, particuli\u00e8rement pr\u00e8s des rapides de Kamchay (rivi\u00e8re de Kampot), sur les bois pourris et les petite plantes .Trochomorphapaviei : CUMZ-CM153 (3 specimens in ethanol). Locality no. 15: CMZ-CM162 (2 shells), CUMZ-CM163 . The snails were found to live on tree trunks and on the ground among leaf litter.Locality no. 14: Cambodia, Laos and Vietnam .T.paviei closely resembles T.saigonensis  that was described from \u201cPoulo-Condor and Saigon, Cochinchine\u201d. The latter species is slightly smaller (shell width 11 mm), having the last whorl with a wide angled peripheral keel and being slightly convex below the periphery. The type specimens of both species were recently figured in This species was originally described from \u201cDans les for\u00eats, entre Kampot et Phnom-Penh\u201d. The unique characters are a depressed conic shell (shell width 12 mm) with a very strong and sharp peripheral keel, and a widely opened and deep umbilicus. The shell surface has thin and regular radial ridges, and very thin spiral ridges. Based on shell morphology, Taxon classificationAnimaliaStylommatophoraTrochomorphidaesp.4907C676-3C20-5516-A416-2FE97F57D66CCUMZ-CM057 , CUMZ-CM059 (1 specimen in ethanol). Locality no. 13: CUMZ-CM134 (3 shells). The snail was found to live on a tree trunk.Locality no. 10: T.paviei and T.saigonensis in having a larger shell (shell width 14 mm), an elevated domed spire, more whorls, and being nearly flat below the periphery. However, the identification is provisional, and further evidence from examination of genitalia or DNA will be necessary to elucidate their status.The specimens from Prasat Phnom Totong  have a conic shell with a very strong and sharp peripheral keel, widely opened and deep umbilicus, and slightly convex below the periphery. The shell surface has irregular growth lines and very thin spiral ridges. These specimens tend to differ from Cryptozona M\u00f6rch, 1872Taxon classificationAnimaliaStylommatophoraAriophantidaeC459181B-410C-5886-80E0-0BA6909196F3Helixsiamensis Pfeiffer, 1856: 32. Type locality: Siam [Thailand].Hemiplectadichromatica Morlet, 1889: 124, 175, 176, pl. 6, fig. 2. Type locality: de Srak\u00e9o \u00e0 Ang-Son (Siam) .Cryptozonasiamensis : CUMZ-CM147 . The snails were found to live on the ground among leaf litter.Locality no. 4: Cambodia, Laos, Malaysia, Singapore and Thailand .C.siamensis is found throughout the country and the allozyme analysis by C.siamensis probably occupies almost all habitat types through accidental introduction or horticultural trade activities, and this species is especially abundant in human-modified landscapes.This widespread species has recently been recorded from Singapore and Peninsular Malaysia , Laos I and YunnHemiplectadichromatica Morlet, 1889\u201d which was subsequently considered to be conspecific with this species , in which its habitats are associated with human activities. In contrast, Q.weinkauffiana could be found commonly in both natural forest and anthropogenic habitats.The historical record of this species from Cambodia was probably under the name \u201c species . In thisTaxon classificationAnimaliaStylommatophoraAriophantidae4A498BFE-3341-5F92-8207-B3B3E08EF2DAHelixdistincta Pfeiffer, 1850: 69, 70. Type locality: insulis Moluccis [Molucca Islands].Hemiplectadistincta : CUMZ-CM021 (6 shells), CUMZ-CM022 (1 specimen in ethanol), CUMZ-CM023 (1 specimen in ethanol). Locality no. 11: CUMZ-CM069 (2 shells), CUMZ-CM070 . Locality no. 3: CUMZ-CM123 (1 specimen in ethanol). Locality no. 17: CUMZ-CM136 (4 shells). Locality no. 18: CUMZ-CM145 (1 shell). Locality no. 14: CUMZ-CM149 (1 shell). Locality no. 16: CUMZ-CM171 . Locality no. 6: CUMZ-CM180 (3 specimens in ethanol). The small juveniles were found on tree trunks and leaves, while the adults were found to live on the ground among leaf litter.Locality no. 7: Cambodia, Laos, Thailand and Vietnam .Hemiplectadistincta has a wide distribution from Southern Vietnam, throughout Cambodia, northeastern Thailand, and throughout Laos , CUMZ-CM033 (3 shells). Locality no. 10: CUMZ-CM050 (24 shells), CUMZ-CM051  by having a reddish-brown shell with a wide whitish or creamy area surrounding the umbilicus. Sarikabocourti, which is described from \u201cBattambang, Cambodje\u201d, has a uniform brownish shell . Locality no. 13: CUMZ-CM117 . These specimens differ from M.psyche in having a nearly flattened to slightly elevated spire, with a slightly shouldered last whorl and milky shell colour, while M.psyche has a slightly sunken spire, with a well-rounded last whorl and whitish shell colour , CUMZ-CM092  have a small shell (diameter ca. 10 mm), which is depressed, slightly thick, translucent, shiny, and pale reddish-brown. The shell surface is smooth with obvious irregular growth lines. The shell has 5 to 6 whorls, with wide and shallow suture. The spire is convex, with an elevated apex. The last whorl has a well-rounded periphery, with an ovate-lunate aperture and a simple lip. An umbilicus is widely open and deep.Macrochlamyspsyche, Sarika sp. 1 and sp. 2 by having a small size and slightly elevated spire. In contrast, M.psyche and Sarika sp. 1 have a large, whitish shell and a flatten to slightly shrunken spire, while Sarika sp. 2 has a larger, reddish-brown shell with whitish area surrounding the umbilicus. Live specimens are required so that the anatomical characters can be used to discriminate among the species.These specimens can be distinguished from Cambodiparmarion Kuznetsov & Kuzminykh, 1999Taxon classificationAnimaliaStylommatophoraAriophantidaeKuznetsov & Kuzminykh, 19997F1D24E2-D479-5F7E-96A0-A98EDACC1FA6Cambodiparmariondoroshenkoi Kuznetsov & Kuzminykh, 1999: 113\u2013116, figs 1, 2. Type locality: In tropical forest between Motel Lomherkay and Hotel Koh Pos, SW end of Kompong Som [= Sihanoukville], Kompong Som district, Kampot province, Cambodia.CUMZ-CM108 (4 shells). Locality no. 13: CUMZ-CM130 (2 shells), CUMZ-CM131 . The semi-slug was found to live on tree trunks and leaves in the limestone area.Locality no. 12: Known only from the type locality .Microparmarion Simroth,C.doroshenkoi was described, the authors did not mention the characters used to discriminate this species from Parmarionmartensi Simroth,C.doroshenkoi has a solid, ear-shape shell with ca. 2 whorls, a blackish body and mantle, and a long flagellum, while P.martensi has a thin nail-shape shell with a trace of shell coiling, a greyish to blackish body and a short flagellum , CUMZ-CM151 . The semi-slugs were found to live on tree trunks and leaves.Locality no. 14: Cambodia, Laos, Malaysia and Singapore .Parmarionmartensi has also been reported as an introduced species to Samoa and Hawaii , in which the shell is usually entirely covered with movable mantle lobes. d Hawaii .Taxon classificationAnimaliaStylommatophoraHelicarionidaeD96F0748-53E1-5B2B-845F-79B4D0954B75Vitrinarusseola Morelet, 1865: 225. Type locality: Cochinchina. Megausteniarusseola : CUMZ-CM030  was figured in Durgella. Further additional anatomical examination is necessary since the shell morphology is insufficient for species identification.A syntype of Taxon classificationAnimaliaStylommatophoraAriophantidaesp.456EF9A1-F892-5A09-BB2F-C813B4D198A7CUMZ-CM031 . It is distinguished from  lamella .Amphidromus Albers, 1850Taxon classificationAnimaliaStylommatophoraCamaenidae4392F96C-9762-5E88-9AAE-9603DB4DEBB0Bulimusleucoxanthus Martens, 1864: 526. Type locality: unknown.Amphidromus (Amphidromus) atricallosusleucoxanthus : CUMZ-CM018 41CDD142-CBE9-5E02-A065-AB263656D157Bulimus (Amphidromus) semitessellatus Morlet, 1885[1884]: 387, 388, pl. 11, fig. 2, 2a. Type locality: les montagnes qui bordent le grand fleuve au del\u00e0 de Stung-Treng. Les for\u00eats et les montagnes de Kampot \u00e0 Compong-Som .Amphidromus (Syndromus) semitessellatus : Amphidromussemitessellatus : CUMZ-CM040 (2 shells). Locality no. 10: CUMZ-CM055 (1 shell). Locality no. 12: CUMZ-CM101 . The empty shells were collected from the ground, and the living snails probably live on tree trunks and leaves.Locality no. 9: Cambodia, Laos, Thailand and probably in Vietnam , 2019.A.semitessellatus due to the similarity of both brown supra-peripheral and sub-peripheral bands on the penultimate whorls and the geographical proximity. The subgenus Syndromus typically has a small shell which exhibits high variation on shell size, colour, and pattern F49A8378-19A8-5EED-A5E4-A1D43D30DF08Helixnorodomiana Morlet, 1883: 106, 107, pl. 4, fig. 3, 3a, b. Type locality: Khamchay [Cambodia].Chloritisnorodomiana : CUMZ-CM024 (2 shells). Locality no. 9: CUMZ-CM041 (35 shells). Locality no. 10: CUMZ-CM056 (1 shell). Locality no. 11: CUMZ-CM077 . Locality no. 12: CUMZ-CM103 (1 shell). Locality no. 13: CUMZ-CM125 (4 shells), CUMZ-CM126 (1 specimen in ethanol). Locality no. 16: CUMZ-CM173 CC0F820D-5140-5F03-8B7B-20ADBBD3E1F4Helix (Geotrochus) perakensis Crosse, 1879: 199, 200, pl. 8, fig. 4. Type locality: Perak .Ganesellaperakensis : CUMZ-CM159 , CUMZ-CM161  species complex which is composed of 11 nominal species and widely distributed from Western Ghats of India to Indochina and the Greater Sunda Islands  from \u201cTonquin\u201d and G.vatheleti  from \u201cVan Bu, Tonkin\u201d by having a strong peripheral keel. For comparison, G.subperakensis is convex below periphery, with less strong peripheral keel and without brownish spiral band , which was described from Ha-Giang (Northern Vietnam) and Siam [Thailand]. The description itself was based mainly on the Ha-Giang specimen  in having a smaller shell (shell height up to 13 mm), shallow suture with 6 to 7 convex whorls, while G.lantenoisi performs an elongate trochoid, larger shell (shell height up to 18 mm), suture flattened and smooth 9 to 10 whorls. However, further investigations with both genitalia and DNA analysis will be necessary to elucidate the exact relationship between them.This species is very similar to Taxon classificationAnimaliaStylommatophoraCamaenidaeS. Tumpeesuwan & C. Tumpeesuwan, 20209E87B72C-4CCB-5F6D-86FE-0B4A4924EFC2Ganesella (Giardia)Mollusca: Eupulmonata: Camaenidae). Preoccupied by K\u00fcnstler, 1882: (Metamonada: Diplomonadida: Hexamitidae). Ancey, 1907: 195, 203 (Pseudobuliminus (Giardia) : Pseudobuliminus (Girardius) Richardson, 1983: 94. [incorrect subsequent spelling]Giardia : Anceyoconcha Tumpeesuwan & Tumpeesuwan in The distinguished shell character of this genus is sinistral, elongate cylindrical to more or less conical, with 6\u201310 convex whorls. The last whorl is rounded (not keeled), with the aperture ovate to slightly trapezoid and the apertural lip expanded. The columella is vertical, with the umbilicus narrowly opened.Giardia as the subgenus of Ganesella Blanford, 1863 to include two Indochinese sinistral species: Bulimussiamensis Redfield, 1853 and Bulimusrhombostomus Pfeiffer, 1861. Subsequently, this nominal name was used as a subgeneric level of Buliminopsis Heude, 1890 (family Fruticicolidae) by Bradybaenidae as the subgenus of Pseudobuliminus Gredler, 1886, and also designated Bulimussiamensis Redfield, 1853 as the type species. Zilch\u2019s classification was subsequently accepted and used by later authors . authors . Recentlmaenidae . HoweverBradybaenidae, Girardius\u201d, accompanied by diagnostic characters and attributed Bulimussiamensis Redfield, 1853 as the type species. However, this name is considered incorrect subsequent spelling  comb. nov., A.maestratii  comb. nov., A.mantongensis  comb. nov., A.obesa  comb. nov., A.ovoideus  comb. nov., A.pharangensis  comb. nov., A.rhombostomapupoidea  comb. nov., A.rhombostomarhombostoma, A.siamensismaxima  comb. nov., A.siamensisnobilis  comb. nov., A.siamensisobesula  comb. nov., A.siamensispervariabilis  comb. nov., A.siamensissiamensis, A.siamensiszonifera  comb. nov. and A.vignei  comb. nov.Anceyoconcha is probably within the Indochinese region of Cambodia, Laos, Thailand, and Vietnam 773E6D49-5BCA-5F83-8110-6C8F386BBA31Bulimusrhombostomus Pfeiffer, 1861: 194, 195. Type locality: Camboja [Cambodia].Ganesellarhombostoma : Anceyoconcharhombostoma : CUMZ-CM047 (66 shells), CUMZ-CM048 . Locality no. 11: CUMZ-CM085 . Locality no. 13: CUMZ-CM132 (3 shells). Locality no. 17: CUMZ-CM139 (5 shells), CUMZ-CM140 ADF2BB29-F75B-52E1-8FCE-0F006E8D1E31Buliminussiamensis var. obesulaAncey, 1888: 352. Type locality: Saigon, dans le jardin du gouverneur. CUMZ-CM027 (14 shells), CUMZ-CM028 , ovate shell and lower number of whorls. For comparison, A.siamensissiamensis 345FACC2-57F6-5EBD-A592-13F4FB14F3B7Petraeuschaudoensis Rochebrune, 1881a: 70. Type locality: Montagnes de Chaudoe Cambodge .Enachaudocensis [sic]: CUMZ-CM003 . Locality no. 3: CUMZ-CM010 (1 specimen in ethanol). The snails were found to live on tree trunks and leaves.Locality no. 1: Cambodia .This species was originally described from \u201cMontagnes de Chaudoe Cambodge\u201d  probably in the area bordering Cambodia and Vietnam. The original description of this species was brief and without illustration. This species has a sinistral elongate conic to slightly ovate conic shell, with 7 to 9 whorls, which increase regularly; cylindrical, having convex whorl and wide and impressed suture. The shell surface possesses fine growth lines, and the periostracum is thin and brownish. The last whorl is well rounded and without keel near aperture and has a similar diameter to the penultimate whorl. The shell colour is light brownish and translucent (becoming whitish when worn). The aperture is semi-ovate, with expanded and whitish lip and thin or thickened with whitish parietal callus. The columella is straight and dilated, with a rimate umbilicus.A.chaudoensis can be distinguished from A.siamensisobesula in having an elongate cylindrical shell and higher number of whorls, while the latter species has an ovate conic shell and a smaller number of whorls.Based on the original description, Alycaeidae, Clausiliidae, and Plectopylidae. It is possible that the geography of the area without high mountains and other structured habitat types result in comparatively fewer species. In comparison, most of the species Sesarapolita that does not exceed 12 mm in width. However, most land snails collected in our study are large (more than 10 mm) and cover most taxonomic groups, with the exception of the families Assimineidae and Diplommatinidae, both of which have been reported by Cambodia has received the least attention from malacologists for inventorying the land snail fauna, compared to other adjacent countries within the Indo-Chinese region, e.g. Thailand , Laos I and Viet"}
+{"text": "Methanobrevibacter smithii and Methanobrevibacter oralis DNA sequences, were detected in 21/527 (3.9%) sputum samples, 2/188 (1.06%) bronchoalveolar lavages, and none of 193 tracheo-bronchial aspirations. Further, fluorescence in situ hybridization detected methanogens in three sputum investigated specimens with stick morphology suggesting M. oralis and in another one bronchoalveolar lavage sample investigated, diplococal morphology suggesting M. smithii. These observations extend the known territory of methanogens to the respiratory tract and lay the foundations for further interpretation of their detection as pathogens in any future cases of isolation from bronchoalveolar lavages and the lungs.Methanogens, the sole microbes producing methane, are archaea commonly found in human anaerobic microbiota. Methanogens are emerging as opportunistic pathogens associated with dysbiosis and are also detected and cultured in anaerobic abscesses. Their presence in the respiratory tract is yet unknown. As a preliminary answer, prospective investigation of 908 respiratory tract samples using polyphasic approach combining PCR-sequencing, real-time PCR, fluorescent in situ hybridization (FISH), and methanogens culture was carried out.  Methanobrevibacter oralis, Methanosarcina mazei, Methanobacterium congolense, Methanobrevibacter smithii, and Methanobrevibacter massiliense) [M. smithii [M. smithii, M. oralis, Methanosphaera stadtmanae, Methanomassilicoccus luminyensis, and Methanobrevibacter arboriphilicus) [M. smithii and M. oralis have been associated with dysbiosis affecting the oral cavity, the gut [Methanogenic archaea (referred to in this study as methanogens), the sole organisms known to produce methane, have been characterized in the oral cavity microbiota (iliense) ,2,3, in  smithii , in digehilicus) ,6 and rehilicus)   and blast programme of NCBI (https://blast.ncbi.nlm.nih.gov/Blast.cgi) were used for sequence analyses. A total of 5\u03bcL of distilled water were used as a negative control for sputum samples and bronchial aspirates, and 5\u03bcL of 0.9% NaCl saline for bronchoalveolar lavage.All samples were examined by standard polymerase chain reaction (PCR) assays targeting the 16S rRNA gene in an automated 5A PTC-200 thermal cycler . The 16S gene was amplified using broad-range rRNA primers SDArch0333aS15 (5\u2032-TCCAGGCCCTACGGG-3\u2032) and SDArch0958aA19 (5\u2032-YCCGGCGTTGTTGAMTCCAATT-3\u2032)  . The PCRM. oralis cnp602F primer 5-GCTGGTGTAATCGAACCTAAACG-3 (Eurogentec); cnp602R primer 5-CACCCATACCCGGATCCATA-3 (Eurogentec) and the probe cnp602-FAM 5-AGCAGTGCACCTGCTGATATGGAAGG-3 (Eurogentec) [M. smithii ; Smit.16S-862R, 5-CTCCCAGGGTAGAGGTGAAA-3 (Eurogentec) and the probe Smit.16S FAM, 5-CCGTCAGAATCGTTCCAGTCAG-3 (Eurogentec) [The PCR products were sequenced using the same primers as used for PCRs following this program: a 1-min denaturation step at 96 \u00b0C, followed by 25 cycles of denaturation of 10 s at 96 \u00b0C, a 20-s annealing at 50 \u00b0C and a 4-min extension at 60 \u00b0C. The MultiScreen 96-well plates Millipore  containing 5% of Sephadex G-50  were used to purify the sequencing products. The sequences were analyzed and edited following the protocol described previously . Furtherogentec) ) speciesBacteroides thetaiotaomicron (104 cells/mL) for hydrogen production [B. thetaiotaomicron in SAB broth were incubated at 37 \u00b0C for 7 days. Subculture was seeded on a Petri dish containing SAB medium supplemented with 15 g/L agar (Sigma-Aldrich) and deposited in the upper chamber of a double-chamber box. Methanogen colonies appeared after 9\u201312 days of incubation [A volume of 250 \u03bcL of each sample diluted in PBS was seeded under anaerobic chamber in a sterile Hungate tube  ,21. Eachoduction  and the cubation .Only PCR positive samples were analyzed by FISH using the method previously described ,4. In brWe used a negative methanogen PCR sputum sample and a negative methanogen bronchoalveolar lavage sample as negative controls.M. oralis-positive samples exhibiting 99% sequence similarity with M. oralis strain ZR (GenBank: NR_104878.1) and two M. smithii-positive samples exhibiting 99% sequence similarity with M. smithii ATCC 35,061 (GenBank NR: 074235.1); in 2/188 (1.06%) of bronchoalveolar lavage samples  and one M. smithii-positive sample exhibiting 99.97% similarity with M. smithii strain C2 CSUR P5816 (GenBank: LR590664.1). None of the bronchial aspirate samples were PCR-positive.In the presence of negative control which remained negative, PCR-sequencing disclosed methanogens in 21/527 (3.98%) of the sputum samples including 19 M. smithii 16S rRNA gene yielded a median Ct of 35.7 \u00b1 0.47, indicating a M. smithii load of 1.32 \u00d7 104 \u00b1 0.56 \u00d7 104/mL in the case of two sputum positive samples and Ct of 36.2 indicative of an M. smithii load of 0.8 \u00d7 104/mL in case of one bronchoalveolar lavage positive sample.In the presence of negative control which remained negative, all the positive samples by PCR-sequencing were positive by real-time PCR. Real-time PCR analyses targeting the M. oralis cnp-60 gene yielded a median Ct of 33.2 \u00b1 1.07 indicative of an M. oralis load of 2.61 \u00d7 105 \u00b1 1.23 \u00d7 105/mL in the case of 19 sputum positive samples and Ct of 35.3 indicative of an M. oralis load of 1.41 \u00d7 104 in case of one bronchoalveolar lavage positive sample  in the saliva ,32. Inte"}
+{"text": "Beilschmiedia erythrophloia, B. robusta, B. yunnanensis, Cryptocarya concinna, C. impressa, C. infectoria, Litsea viridis, Machilus balansa, M. grandifolia, Neolitsea ellipsoidea, and Phoebe angustifolia) have been obtained by hydrodistillation and the chemical compositions analyzed by gas chromatography \u2013 mass spectrometry (GC-MS) and gas chromatography with flame ionization detection (GC-FID). The essential oils were screened for larvicidal activity against Aedes aegypti, Ae. albopictus, and Culex quinquefasciatus, and for antimicrobial activity against Enterococcus faecalis, Staphylococcus aureus, Bacillus cereus, Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, and Candida albicans. The leaf essential oil of N. ellipsoidea, rich in (E)-\u03b2-ocimene (87.6%), showed excellent larvicidal activity against Ae. aegypti with a 24 h LC50 of 6.59 \u03bcg/mL. The leaf essential oil of C. infectoria, dominated by germacrene D (55.5%) and bicyclogermacrene (11.4%), exhibited remarkable larvicidal activity against Cx. quinquefasciatus (48 h LC50 = 0.40 \u03bcg/mL). N. ellipsoidea leaf essential oil also demonstrated notable antibacterial activity against E. faecalis and B. cereus with minimum inhibitory concentration (MIC) values of 16 \u03bcg/mL, while the leaf essential oil of C. impressa showed excellent anticandidal with an MIC of 16 \u03bcg/mL. Leaf essential oils from the Lauraceae should be considered for utilization as alternative agents for controlling mosquito populations and as antimicrobial agents.The Lauraceae is a family rich in aromatic and medicinal plants. Likewise, essential oils derived from members of this family have demonstrated a myriad of biological activities. It is hypothesized that members of the Lauraceae from Vietnam will yield essential oils that may be useful in controlling mosquito populations and treating microbial infections. In this work, the leaf essential oils of eleven species of Lauraceae ( Persea americana Mill.) for its fruit, bay leaf (Laurus nobilis L.) used in cooking, and the spice cinnamon (Cinnamomum verum J. Presl) [Sassafras albidum (Nutt.) Nees) [Lindera benzoin (L.) Blume) [Aniba rosaeodora Ducke) [Cinnamomum camphora (L.) J. Presl.) [Litsea cubeba (Lour.) Pers.) [The Lauraceae is made up of around 55 genera and 3000 species of tropical and warm temperate trees and shrubs, with Southeast Asia and Brazil serving as species-rich hot spots . Several. Presl) . Several) Blume) . Many spa Ducke) , camphor Presl.) , and aro) Pers.) .Based on the utility and properties of Lauraceae essential oils, it is hypothesized that members of the Lauraceae found in Vietnam have biologically active essential oils that may be useful in controlling mosquito populations or as antimicrobial agents. Eleven species of Lauraceae from north-central Vietnam have been collected, the essential oils obtained by hydrodistillation, chemical compositions analyzed, and the oils screened for mosquito larvicidal activity and for antimicrobial activity.Beilschmiedia Nees is comprised of around 250 species of trees and shrubs [Beilschmiedia has been reviewed [The genus d shrubs  and are d shrubs . The phyreviewed .Beilschmiedia erythrophloia Hayata is a tree found in Taiwan, southern China, Hainan Island, and Ryukyu Islands (Japan) [B. erythrophloia have revealed endiandric acid derivatives from the roots [E)-caryophyllene and \u03b1-humulene [ (Japan) ,13. In V (Japan) . Previouhe roots ,16, the humulene .Beilschmiedia robusta C.K. Allen is a tree, 10\u201315 m tall that is recorded from Guangzi, southwestern Guizhou, Xizang, and Yunnan provinces in China [B. robusta.in China ,18. In Vin China . A perusBeilschmiedia yunnanensis H.H. Hu is a tree, up to 18 m tall and is found in Guangdong, southern Guangxi, and southern Yunnan provinces in China [B. yunnanensis.in China . In Vietin China . A literCryptocarya R. Br. is a pantropical genus of around 300 species [Cryptocarya concinna Hance  is a tree up to 18 m tall, and ranges from southern China  to northern Vietnam [C. concinna have shown the roots to contain cytotoxic cryptocaryone [C. concinna. species . Cryptoc Vietnam ,12. In V Vietnam . Previouocaryone , the leaocaryone , and theocaryone . There hCryptocarya impressa Miq. is native to Vietnam, Laos, the Malay Peninsula, Borneo and Sumatra [C. impressa. Sumatra . In Viet Sumatra . To our Cryptocarya infectoria (Blume) Miq. (syn. Cylicodaphne infectoria Blume) is a tree up to 33 m tall that is native to Indo-China and Malesia [C. infectoria [N-methylisococlaurine, and N-methyllaurotetanine have also been isolated from the bark of C. infectoria [ Malesia ,25,26. I Malesia . The cytfectoria ,28. The fectoria . There hLitsea Lam. consists of around 300 species distributed in tropical and warm subtropical regions of Asia, Australia, and the Americas [Litsea viridis H. Liu is a small tree, 3-6 m tall, found in south-eastern Yunnan province (China) and Cao B\u1eb1ng, Ngh\u1ec7 An, \u0110\u00e0 N\u1eb5ng, and \u0110\u1eafk L\u1eafk provinces (Vietnam) [The genus Americas . Litsea Vietnam) ,14. TherMachilus Rumph. ex Nees is comprised of around 100 species distributed in southern and south-eastern Asia [Machilus balansae (Airy Shaw) F.N. Wei & S.C. Tang  is endemic to Vietnam and is generally found at low elevations in north Vietnam [Machilus grandifolia S.K. Lee & F.N. Wei is now regarded as a new synonym of M. balansae [M. balansae or M. grandifolia.The genus ern Asia ,14. Mach Vietnam . Machilubalansae . To our Neolitsea (Benth.) Merr. Contains around 85 species distributed from Indo-Malaysia to East Asia [Neolitsea ellipsoidea K.C. Allen is a tree up to 30 m in height [The genus ast Asia ,14. Neoln height . The spePhoebe Nees [There are around 100 species in the genus ebe Nees , which rebe Nees .Phoebe angustifolia Meisn. is a small shrub found in southeastern Yunnan (China), Myanmar, India, and Vietnam [P. angustifolia from Vietnam has been reported, which showed the major components to be spathulenol (17.0%), palmitic acid (13.0%), sabinene (6.4%), bicyclogermacrene (5.9%), and artemisia triene (5.1%) [ Vietnam . In Viet Vietnam . The leae (5.1%) .The essential oil collection details and yields are summarized in B. erythrophloia, B. robusta, and B. yunnanensis are compiled in Beilschmiedia leaf essential oils were dominated by sesquiterpene hydrocarbons. A preponderance of sesquiterpene hydrocarbons has been previously seen in Beilschmiedia leaf essential oils from Malaysia [The essential oil compositions of Malaysia  and fromMalaysia .B. erythrophloia essential oil were bicyclogermacrene (30.5%), (Z)-\u03b2-ocimene (26.1%), and (E)-caryophyllene (18.3%). While qualitatively similar, there are notable differences between the essential oil from Vietnam in this work and that reported by Su and Ho from Taiwan [The major components in m Taiwan ; the samB. robusta and B. yunnanensis leaf oils were rich in (E)-caryophyllene , \u03b1-humulene (13.4% and 9.9%), and bicyclogermacrene (8.6% and 8.4%). The leaf oil of B. robusta had a high concentration of germacrene D (20.3%), while B. yunnanensis oil was rich in 9-epi-(E)-caryophyllene (21.2%).Both C. concinna (from two locations), C. impressa, and C. infectoria are listed in C. impressa and C. infectoria leaf essential oils, while oxygenated sesquiterpenoids were abundant in C. concinna essential oil from Nam Dong and monoterpene hydrocarbons dominated the leaf oil of C. concinna from Pu Hoat.The leaf essential compositions of C. concinna from two different collection sites were qualitatively similar, but quantitatively different. That is, the abundant components in the Nam Dong sample were also observed in the Pu Hoat sample, and vice versa. Thus, for example, \u03b1-pinene, \u03b2-pinene, and myrcene were abundant in the Pu Hoat sample  but were found in lower concentrations in the sample from Nam Dong . Conversely, the sesquiterpenoids, (E)-caryophyllene, spathulenol, and caryophyllene oxide were abundant in the sample from Nam Dong , but less concentrated in the Pu Hoat sample .The leaf essential oils of C. impressa were bicyclogermacrene (18.7%), (E)-caryophyllene (10.8%), dodecanal (10.8%), -\u03b1-farnesene (7.9%), and \u03b1-humulene (6.3%). Germacrene D (55.5%) dominated the essential oil composition of C. infectoria, which was also composed of bicyclogermacrene (11.4%) and \u03b4-elemene (5.1%) as major components. The major components of the leaf essential oil of L. viridis, M. balansae, M. grandifolia, N. ellisoidea, and P. angustifolia are compiled in The chemical compositions of the leaf essential oils of L. viridis leaf essential oil were bicyclogermacrene (25.5%), decanal (14.4%), \u03b1-pinene (11.1%), and \u03b2-pinene (8.3%). This is the first report on the essential oil from this plant.The major components in M. balansae and M. grandifolia are considered conspecific, the essential oil compositions showed pronounced differences. The leaf oil of M. balansae was dominated by bicyclogermacrene (41.5%), which was not detected in the essential oil of M. grandifolia. Likewise, the sesquiterpene alcohols (E)-nerolidol and globulol were abundant constituents in M. grandifolia , but (E)-nerolidol was much lower in M. balansae (8.7%) and globulol was not detected. The taxonomy of these two plants deserves closer scrutiny.Although N. ellipsoidea was dominated by (E)-\u03b2-ocimene (87.6%). (E)-\u03b2-Ocimene was also found to be the dominant compound (85.6%) in the leaf essential oil of N. polycarpa from Vietnam [N. sericea from Korea (13.3%) [N. variabillima from Taiwan (13.4%) [N. aciculata from Korea (9.7%) [E)-\u03b2-ocimene was only a minor component in the leaf oils of N. australiensis, N. brassii, or N. dealbata from Australia [N. pallens from India [N. foliosa leaf essential oil from India [The leaf essential oil of  Vietnam , and one (13.3%) , N. vari (13.4%) , and N. a (9.7%) . In contustralia , and N. om India , and wasom India .P. angustifolia from P\u00f9 Ho\u1ea1t Nature Reserve (northern Vietnam) in this study was rich in \u03b1-pinene (26.9%), \u03b2-pinene (20.8%), spathulenol (5.4%), (E)-caryophyllene (5.3%), and p-cymene (5.0%), which differs markedly from a previous study on the leaf essential oil from Sao La Nature Reserve . The previous work reported spathulenol (17.0%), palmitic acid (13.0%), sabinene (6.4%), bicyclogermacrene (5.9%), and artemisia triene (5.1%) to be the major components [The leaf essential oil of mponents . There iThe 24-h and 48-h larvicidal activities of Lauraceae leaf essential oils from Vietnam are summarized in N. ellipsoidea showed the greatest activity against Ae. aegypti with 24-h and 48-h LC50 values of 6.59 and 4.04 \u03bcg/mL, respectively. Similar larvicidal activities were observed against Cx. quinquefasciatus (24-h and 48-h LC50 = 7.47 and 4.65 \u03bcg/mL) for this essential oil. Interestingly, although the larvicidal activities of C. infectoria leaf essential oil were not as impressive against Ae. aegypti or Ae. albopictus, the essential oil did show much better activity against Cx. quinquefasciatus (24-h LC50 = 10.8 \u03bcg/mL), particularly after 48 h of exposure (48-h LC50 = 0.402 \u03bcg/mL). Unfortunately, the limited quantities available for several of the essential oils precluded larvicidal screening. However, the larvicidal activity of the untested essential oils will be investigated in future studies.Of the Lauraceae essential oils screened for larvicidal activity, N. ellipsoidea leaf essential oil, (E)-\u03b2-ocimene (87.6%), is not likely responsible for the observed larvicidal activity. The (E)-\u03b2-ocimene-rich (94.8%) essential oil of Porophyllum ruderale showed an LC50 of 173.7 \u03bcg/mL against Ae. aegypti [Syzygium jambolana, with (Z)-\u03b2-ocimene (27.2%) and (E)-\u03b2-ocimene (12.2%), was inactive against Ae. aegypti (LC50 = 433 \u03bcg/mL) [N. ellipsoidea essential oil can likely be attributed to synergistic effects involving minor components.The major component of  aegypti . Likewis3 \u03bcg/mL) . The excC. infectoria was rich in the germacrene sesquiterpenes germacrene D (55.5%) and bicyclogermacrene (11.4%), and these compounds may be responsible for the larvicidal activity. Germacrene D has demonstrated notable larvicidal activity against Ae. aegypti and Cx. quinquefasciatus  [Ae. albopictus and Cx. tritaeniorhynchus  [The leaf essential oil of ctively) , and bicctively) .C. concinna from Nam Dong is consistent with the marginal activities observed for the major components. (E)-Caryophyllene, caryophyllene oxide, and \u03b1-pinene have shown modest mosquito larvicidal activities [50 = 65 \u03bcg/mL against Cx. quinquefasciatus) [50 = 15.4 \u03bcg/mL against Ae. aegypti) [50 = 22.4 \u03bcg/mL against Ae. aegypti) [Tephrosia toxicaria (42.3% spathulenol) had an LC50 of 63.1 \u03bcg/mL against Ae. aegypti [Guarea sylvatica essential oil from branches (14.3% spathulenol) showed LC50 against Ae. aegypti of 274 \u03bcg/mL [The marginal larvicidal activity of tivities . \u03b2-Pinensciatus) , (LC50 =aegypti) . Spathul aegypti , while G74 \u03bcg/mL .L. viridis and N. ellipsoidea leaf essential oils demonstrated particularly notable activities against E. faecalis and B. cereus with minimum inhibitory concentration (MIC) values of 16 \u03bcg/mL. The leaf essential oil of C. impressa also showed excellent anticandidal activity against C. albicans with an MIC of 16 \u03bcg/mL.Several of the leaf essential oils of the Lauraceae were screened for antimicrobial activity . All of L. viridis leaf oil, bicyclogermacrene, has shown antibacterial activity against B. cereus [E. faecalis [L. viridis leaf essential oil, bicyclogermacrene, decanal, \u03b1-pinene, and \u03b2-pinene, can account for the observed antibacterial activity.The major component of . cereus . Likewisfaecalis  as well faecalis . Similarfaecalis ,56. Decafaecalis ,58. ThusE)-\u03b2-Ocimene dominated the leaf essential oil of N. ellipsoidea, but this compound has demonstrated relatively marginal antibacterial activity [E)-\u03b2-ocimene with minor essential oil components may play a role in the antibacterial activity of N. ellipsoidea leaf oil.-caryophyllene nor \u03b1-humulene have shown strong anti-Candida albicans activity [E)-caryophyllene do not exhibit notable activity against Candida spp. [C. albicans with an MIC of 125 \u03bcg/mL [The components responsible for the anticandidal activity of activity ,56. The ida spp. ,60. Dode25 \u03bcg/mL .Leaves were collected from wild-growing trees in north-central Vietnam. Plants were identified by Do Ngoc Dai and voucher specimens  have bee2 (1 mL/min), injector temperature (PTV: programmable temperature vaporization) 250 \u00b0C, detector temperature 260 \u00b0C, column temperature programmed from 60 \u00b0C (2 min hold) to 220 \u00b0C (10 min hold) at 4 \u00b0C/min. Samples were injected using a split mode with a split ratio of 10:1. The volume injected was 1.0 \u03bcL. Inlet pressure was 6.1 kPa.Gas chromatographic (GC) analysis was performed on an Agilent Technologies HP 7890A Plus Gas chromatograph equipped with a FID and fitted with HP-5ms column . The analytical conditions were: carrier gas HAn Agilent Technologies  HP 7890A Plus Chromatograph fitted with a fused silica capillary HP-5ms column  and interfaced with a mass spectrometer HP 5973 MSD was used for the GC/MS analysis, under the same conditions as those used for GC analysis. The conditions were the same as described above with He (1 mL/min) as carrier gas. The MS conditions were as follows: ionization voltage 70 eV; emission current 40 mA; acquisitions scan mass range of 35\u2013350 amu at a sampling rate of 1.0 scan/s. Compound identification was carried out by comparison of the MS fragmentation patterns and calculated retention indices with those available in the databases ,37 and, Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus were carried out as previously described [50 values, LC90 values, and 95% confidence limits were determined by log-probit analysis using Minitab\u00ae 19 .Larvicidal activities against escribed ; LC50 vaEnterococcus faecalis (ATCC299212), Staphylococcus aureus (ATCC25923), Bacillus cereus (ATCC14579), three strains of Gram-negative test bacteria, Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC27853), Salmonella enterica (ATCC13076) and one strain of yeast, Candida albicans (ATCC 10231). Minimum inhibitory concentration (MIC) and median inhibitory concentration (IC50) values were measured by the microdilution broth susceptibility assay as previously described [The bacterial growth inhibition of the essential oils was evaluated using three strains of Gram-positive test bacteria, escribed . Neolitsea ellipsoidea, dominated by (E)-\u03b2-ocimene, showed excellent larvicidal activity against Aedes aegypti and antibacterial activity against Enterococcus faecalis and Bacillus cereus; Cryptocarya infectoria leaf essential oil, rich in germacrene D and bicyclogermacrene, showed excellent larvicidal activity on Culex quinquefasciatus and anticandidal activity against Candida albicans. The leaf essential oil of Litsea viridis, which was rich in bicyclogermacrene, also showed good antibacterial properties. The biological properties of these Lauraceae essential oils suggest that they may serve as potential \u201cgreen\u201d alternatives, as also described for Lamiaceae family plants [Of the eleven species of Lauraceae examined in this work, the leaf essential oil of y plants , for use"}
+{"text": "Ribosomal protein S6 (RPS6) is the only phosphorylatable protein of the eukaryotic 40S ribosomal subunit.Ribosomes with phosphorylated RPS6 can selectively translate 5\u2019TOP--containing mRNAsthat encode most proteins of the translation apparatus. The study of translational control of 5\u2019TOP-mRNAs, which arepreferentially translated when RPS6 is phosphorylated and cease to be translated when RPS6 is de-phosphorylated,is particularly important. In Arabidopsis thaliana, AtRPS6 is phosphorylated by kinase AtRPS6K2, which should in turnbe phosphorylated by upper level kinases (AtPDK1 \u2013 at serine (S) 296, AtTOR \u2013 at threonine (T) 455 and S437) for fullactivation. We have cloned AtRPS6K2 cDNA gene and carried out in vitro mutagenesis replacing codons encodingS296, S437 and T455 by triplets of phosphomimetic glutamic acid (E). After the expression of both natural and mutatedcDNAs in Escherichia coli cells, two recombinant proteins were isolated: native AtRPS6K2 and presumably constitutivelyactive AtRPS6K2. The activity of these variants was tested in vitro. Both kinases couldphosphorylate wheat (Triticum aestivum L.) TaRPS6 as part of 40S ribosomal subunits isolated from wheat embryos,though the non-mutated variant had less activity than phosphomimetic one. The ability of recombinant non-mutatedkinase to phosphorylate TaRPS6 can be explained by its phosphorylation by bacterial kinases during the expressionand isolation steps. The phosphomimetically mutated AtRPS6K2 can serve as a tool to investigatepreferential translation of 5\u2019TOP-mRNAs in wheat germ cell-free system, in which most of 40S ribosomal subunits havephosphorylated TaRPS6. Besides, such an approach has a biotechnological application in producing genetically modifiedplants with increased biomass and productivity through stimulation of cell growth and division. Growth and division of cells depending on the availability ofnutrients, energy resources, as well as responding to internaland external stimuli are coordinated by signaling system basedon a multilevel cascade of serine-threonine protein kinases.These kinases transmit signals from internal and externalevents to the protein synthesis apparatus, causing inhibitionor enhancement of protein synthesis . The target of rapamycin (TOR) kinase \u2013is the master signaling integrator, central hub synchronizingcell growth according to the nutrient and energy status as wellas environmental influences . In mammals,TOR forms two functionally distinct protein complexes:mTORC1 containing RAPTOR (regulatory-associated proteinof mTOR), and mTORC2 containing RICTOR (rapamycininsensitivecompanion of mTOR) .In favorable conditions mTORC1 phosphorylates RPS6K. Complete activation of mammalian RPS6K byphosphorylationis dependent on another upper level PDK1kinase . The fully activated RPS6K inturn phosphorylates the S6 ribosomal protein (RPS6) .At transcriptional level, phosphorylation of pRPS6 in nucleolusleads to activation of rRNA gene promoter and ribosomogenesis. In cytosol,RPS6 phosphorylation promotes the selective translationof special group of cellular mRNAs, containing 5\u2032-terminaloligo-pyrimidine tract (5\u2032TOP) in their 5\u2032-untranslated regions(5\u2032UTRs) . The number of these5\u2032TOP-containing mRNAs, according to various estimates,ranges from one hundred to two hundred and forty . They encode almost all theproteins of the translation apparatus -binding proteins, etc.) , as well asother protein families associated with lysosome functions, metabolismand proliferation .As in yeast and animals, TOR kinase is involved in controllingplant growth and cell division . But inplants, only orthologs of genes encoding mTORC1 were found. No clear orthologs of theRICTOR have yet been found in plants . TOR proteins are highly conserved ineukaryotes. For example, in A. thaliana and Homo sapiensthey share 73 % amino acid sequence identity in the kinasedomains .Although functioning of this main regulator of cell processeshas been well studied in other eukaryotes, knowledgeof the regulation of translation and gene expression in plantsis very limited. Most studies of the regulation of cellular processby plant RPS6-kinase were performed on a model objectA. thaliana containing two very similar forms \u2013 At RPS6K1and At RPS6K2. It was shown that only At RPS6K2 is able tophosphorylate RPS6 and stimulate an increase in cell size . Forthe complete activation of At RPS6K2, it is necessary that it bephosphorylated by pPDK1 kinase (at Ser296), pTOR kinase (atThr455), as well as by one more, unknown, kinase (at Ser437).Although pTOR\u2192S6K signaling plays multiple roles intranslational control , mechanisms used byTOR kinase to impact global protein synthesis in plants are notwell understood . New data are currently appearing on the involvementof pRPS6K1 in the promotion of translation reinitiationof upstream open reading frame (uORF)-containing viral andcellular mRNAs via phosphorylation of eIF3h  and in regulation of translation initiation underenergy-deficient conditions via formation of the functionaleIF4F complex . Nevertheless, the role ofplant pRPS6K2 and pRPS6 phosphorylation in translationregulation in the cytosol remains unclear .It is practically impossible to control the multiple and simultaneousphosphorylation of At RPS6K2 kinase by the kinases of theupper regulatory level for experimental purposes. Therefore, wedecided to use a different approach to achieve the phosphorylationof plant RPS6 using the mutated form of At RPS6K2,which should be stably active. We have cloned the AtRPS6K2cDNA gene and performed in vitro mutagenesis of this cDNAby replacing codons encoding serines at positions 296 and 437,as well as threonine at position 455 with triplets encoding thephosphomimetic amino acid \u2013 glutamic acid. After expressionof non-mutated and mutated cDNA gene in E. coli cells the nativeAt RPS6K2 and the phosphomimetic At RPS6K2 recombinant protein was obtained. The secondone is expected to have stable kinase activity, regardless of theupper-level kinases, that could be used as a unique tool for theartificial phosphorylation of TaRPS6 in a wheat germ cell-freetranslation system. Mutated version of cDNA gene encodingthe constantly active form of AtRPS6K2 may also be used toobtain genetically modified plants with increased productivity,earlier ripening and a higher rate of biomass accumulation.Cloning of AtRPS6K2 cDNA gene. The total RNA was isolatedfrom A. thaliana (Col-0 ecotype) leaves using Tri-reagent(Sigma). The reverse transcription reaction was performedusing Maxima Reverse Transcriptase (Thermo) and \u2018AtS6K2-rev-3UTR\u2019 primer (5\u2032-GAATTCGAGAAATAGGTTTCTTCAAACAACCGTTGATTTTG), which allowed to differentiateAt RPS6K2 from At RPS6K1 mRNAs. RT-PCR was performed in 25 \u03bcl reaction using Phusion High-fidelity DNApolymerase(Thermo), 0.2 pM primers \u2018Nde-AtS6K2-for\u2019(5\u2032- GGGCGAATTGGGTCATATGGTTTCTTCTCAGTG)and \u2018AtS6K2-Xho-rev\u2019 (5\u2032-AAACTCGAGCTACAAGTTGGATGTGGTCCGATGA) and 2.5 \u03bcl of RT-reaction mixture.Temperature regime: stage 1\u20135 min at 94 \u00b0C, 1 cycle; stage2\u201310 s at 98 \u00b0C, 20 s at 49 \u00b0C, 45 s at 72 \u00b0\u0421, 4 cycles; stage3\u201310 s at 98 \u00b0C, 20 s at 52 \u00b0C, 45 s at 72 \u00b0\u0421, 30 cycles; stage4\u20135 min at 72 \u00b0C, 1 cycle. The PCR product (~1425 bp)was digested with NdeI/XhoI and cloned into pET19b vectordigested with the same enzymes resulting \u2018Pet19b-His-At RPS6K2\u2019 plasmid.Mutagenesis. In vitro mutagenesis was performed in threesteps using QuikChange II Site-Directed Mutagenesis Kit(Agilent technologies) according to the manufacturer\u2019s protocol.At the first step \u2018Pet19b-His-AtRPS6K2\u2019 plasmid wasamplified entirely using Pfu Ultra High-Fidelity DNA polymerase(Thermo) and complementary primers: \u2018S296-Glu-dir\u2019(5\u2032-AAACACAAGATCAAACGAAATGTGTGGGACTACGGA) and \u2018S296-Glu-rev\u2019 (5\u2032-TCCGTAGTCCCACACATTTCGTTTGATCTTGTGTTT) containing correspondingnucleotide substitutions. Temperature regime: stage 1\u201330 s at95 \u00b0C, 1 cycle; stage 2\u201330 s at 95 \u00b0C, 1 min at 55 \u00b0C, 7 min30 s at 68 \u00b0C, 18 cycles. The reaction mixture was furthertreated with restriction enzyme DpnI, which cleaves methylatedDNA into fragments at 5\u2032-Gm6ATC-3\u2032 sequences. Sincethe original plasmid was methylated (dam+ E. coli strain DH5was used for plasmid enrichment), the restriction enzymeDpnI had cleaved the original non-mutated plasmid, whereas\u2018Pet19b-His-AtRPS6K2(S296E)\u2019 plasmid synthesized duringPCR-step remained intact. Subsequently, the competent E. colicells (XL1-Blue strain) were transformed with the reactionmixture. Another two mutagenesis steps for the production of\u2018Pet19b-His-At RPS6K2\u2019 and \u2018Pet19b-His-At RPS6K2\u2019 plasmids were done inthe same manner using \u2018S437-Glu-dir\u2019 (5\u2032-ACATGTCTGTTTTGGATGAACCAGCAAGTAGTCCCA)/\u2018S437-Glu-rev\u2019(5\u2032-TGGGACTACTTGCTGGTTCATCCAAAACAGACATGT) or \u2018T455-Glu-dir\u2019 (5\u2032-ACCCTTTTACAAACTTCGAATACGTCAGGCCTCCTCA)/\u2018T455-Glu-rev\u2019 (5\u2032-TGAGGAGGCCTGACGTATTCGAAGTTTGTAAAAGGGT)primers respectively. Resulting DNA-constructs were usedas templates for the next in vitro mutagenesis step. The insertscloned into the recombinant plasmids were sequencedfrom both ends by the dideoxy chain termination methodusing Big Dye Terminator v.3.1 sequencing kit (Thermo) onthe 310 genetic analyzer (Applied Biosystems) according tomanufacturer\u2019s recommendations.Expression and purification of recombinant proteins.E. coli strain BL21(DE3) cells transformed with recombinant\u2018Pet19b-His-AtRPS6K2\u2019 or \u2018Pet19b-His-At RPS6K2\u2019 plasmid were grown in 100 ml of LB mediumcontaining ampicillin (100 \u03bcg/ml) at 30 \u00b0C to A600of 0.5 unit. The expression of recombinant proteins wasinduced by 0.8 mM isopropyl \u03b2-D-1-thiogalactopyranoside(IPTG) at 30 \u00b0C for 4 h. Cells were collected by centrifugation,resuspended in His-buffer  containing 10 mM imidazole, and then lysedby addition of lysozyme (1 mg/ml) and sonication. The celldebris was removed by centrifugation at 10000 g for 20 min at 4 \u00b0C. Supernatant was combined with PerfectPro Ni\u2013NTAresin suspension (5-Prime), shaken at 4 \u00b0C for 1 h followedby flow throw in column. The resin was washed twice byHis-buffer containing 20 mM imidazole at 4 \u00b0C. His-taggedproteins bound to the resin were eluted with His-buffer containing250 mM imidazole and dialyzed against dialysis buffer2, pH 7.6) at4 \u00b0C for 12 h. Dialyzed proteins were concentrated by centrifugationin 10,000 MWCO HY columns (Sartorius) accordingto the manufacturer\u2019s instructions. Protein concentration wasestimated by the Bradford protein assay .Gel-electrophoresis. Proteins were separated by standardSDS-PAGE in Tris-Glycine gel system  according toU.K. Laemmli (1970). After electrophoresis, the gels werefixed and stained in PageBlue Protein Staining Solution(Thermo) or subjected to semi-dry blotting in transfer buffer ethanol) for1 h at 0.8 mA/cm2 and 20 V using 0.22 \u03bcm pore NitroBindnitrocellulose membranes (GVS).Western blotting. For immunodetection of His-At RPS6K2and His-At RPS6K2 proteins, the blotswere first \u2018blocked\u2019 by submerging them in blocking solution Tween 20, pH 7.5) containing 5 % skim milk) for 1 h at25 \u00b0C with gentle shaking. The blots were then incubated withPenta-His mouse antibodies (5 Prime) diluted  in theblocking solution for 1 h at 25 \u00b0C, thoroughly washed threetimes with TBST buffer, and incubated for 1 h at 25 \u00b0C withhorseradish peroxidase-conjugated goat anti-mouse antibodies(Santa Cruz) diluted  in blocking solution. Afterdouble washes in TBST and double washes in TBS, the blotswere chemiluminescence developed using ChemiluminescentPeroxidase Substrate-3 detection reagents (Sigma). Animage of the membrane was then produced on X-ray film.Monoclonal Anti-Phosphoserine Mouse Antibodies (Sigma)and Monoclonal Anti-Phosphothreonine Mouse Antibodies(Sigma) were used as 1st antibodies (at 1:300 dilution in TBSTcontaining 5 % BSA) for the detection of phosphorylationstatus of proteins.260 unit corresponds to 50 pmolof 40S subunits.40S ribosomal subunits isolation. 40S ribosomal subunitswere isolated from wheat  embryos, purified from endosperm, as described previouslyfor ribosomal subunits isolation from human placenta with the ratio of buffer to embryos of6:1. It was considered that 1 AKinase assay. The reaction mixture in 20 \u03bcl contained20 mM TrisAc (pH 7.6), 90 mM KAc, 2.5 mM Mg(OAc)2,1 mM DTT, 10 pmol of 40S ribosomal subunits, 0.1 mM ATP.Purified His-At RPS6K2 or His-At RPS6K2 were added in amount of 2.5 ng/\u03bcl. The mixtures wereincubated for 20 min at 26 \u00b0C.Cloning and mutagenesis of AtRPS6K2 cDNA gene. A totalRNA preparation was isolated from A. thaliana, and reversetranscription was performed using \u2018AtS6K2-rev-3UTR\u2019primer, complementary to 3\u00b4UTR of AtRPS6K2 mRNA, butnot AtRPS6K1 mRNA, allowing to discriminate between them. Then, AtRPS6K2 cDNA was amplified by RT-PCR andcloned into pET19b vector. According to sequencing analysis,AtRPS6K2 cDNA corresponded to #AT3G08720 (GeneBank)sequence.Thus obtained \u2018Pet19b-His-AtRPS6K2\u2019 plasmid was mutatedin vitro in three steps to introduce three phosphomimeticmutations into AtRPS6K2 cDNA. At the first step, theTCC triplet encoding serine at position 296 was replaced bythe GAA triplet, which encodes glutamic acid that imitatesphosphorylated serine. In a second step, the TCT tripletof obtained mutated AtRPS6K2(S296E) cDNA encodingserine at position 437 was mutated to GAA triplet to formAtRPS6K2 cDNA. In the third step, the ACAtriplet of AtRPS6K2 cDNA encoding threonineat position 455 was replaced by the GAA triplet to form themutated AtRPS6K2 cDNA.Expression and purification of recombinant kinases.AtRPS6K2 and AtRPS6K2 cDNAgenes were expressed in E. coli cells, then recombinant Histaggedproteins  respectively) were isolated using immobilizedmetal ion affinity chromatography (IMAC) followed by immunoblottinganalysis .Isolated proteins were purified by dialysis and concentrated.Preparations isolated under native conditions contained acertain amount of impurity polypeptides. Content of recombinantproteins in preparations was corrected according todensitometric analysis data (by ImageJ 1.42). The yield ofpurified and concentrated full-length recombinant proteins His-AtRPS6K2 and His-At RPS6K2was 5.22 mg and 4.52 mg per L of media respectively.Testing the activity of recombinant kinases. Both forms ofkinase (the intact one and that carrying three phosphomimeticsubstitutions) were tested for their ability to phosphorylateTaRPS6 in the composition of 40S ribosomal subunits isolatedfrom wheat embryos. The phosphorylation state of proteinswas tested using monoclonal antibodies against phosphoserine.As can be seen from the data presented in Fig. 2, bothkinases are able to phosphorylate the plant ribosomal proteinS6 (TaRPS6) in composition of 40S ribosomal subunits,although activity of His-At RPS6K2is obviously higher than that of non-mutated His-At RPS6K2. In wheat germ, there are at least two forms of the S6ribosomal protein (A and B); therefore, two bands are observed.Initially, we expected that non-mutated kinase should haveno activity since for its activation in plant cells phosphorylationat three sites is required by upper-level kinases. Thephosphorylation state of purified recombinant kinases waschecked using monoclonal antibodies against phosphoserineand phosphothreonine .As can be seen from the data presented in Fig. 3, thenon-mutated recombinant His-At RPS6K2 kinase producedin E. coli cells was phosphorylated both at serine residues and threonine residues . Thus, some bacterial kinases were able to phosphorylate His-At RPS6K2 protein resulting in its activation.It should be noted that certain non-mutated serine residuesof mutated His-At RPS6K2 recombinantkinase were also phosphorylated ,althoughthis kinase was not phosphorylated at threonineresidues .The interest in studying the mechanisms of TOR-mediatedregulation of mRNA translation in plants is high because othermechanisms of regulation of protein biosynthesis, which arewell described for mammals and yeast, either do not work orfunction within very narrow limits in plants. Indeed, in plantcells eIF4E binding proteins (eIF4E-BPs) were not found, andthere are no genes for these proteins in plant genome . The mechanism of translation suppressionby phosphorylation of peEF2 is not realized in plants. Then,out of four protein-kinases  thatphosphorylate \u03b1-subunit of meIF2 in mammalian cells, onlypGCN2-kinase was detected in plants, that can be activatedunder several but not all stresses. Moreover, it was shown,that factor eIF2B is not necessary for cyclic functioning ofplant peIF2 , and neither its biochemicalactivity nor peIF2B-like factor orthologs were detected inplants till now . These circumstancesmake the TOR system one of the few currently known effectiveregulators of protein biosynthesis in plants.Having obtained the constitutively active protein kinaseAt RPS6K2 with phosphomimeticsubstitutions of key amino acids, we acquire a convenienttool that allows to considerably increase phosphorylationof TaRPS6 in the composition of 40S ribosomal subunits in wheat germ cell-free system. This allows studying importantmechanisms of preferential translation of a specific groupof cellular 5\u2032TOP-containing mRNAs, which is preferablytranslated when pRPS6 is phosphorylated and ceases to betranslated when RPS6 is de-phosphorylated . In addition to fundamental interest the use of cDNA encodingconstitutively active RPS6-protein kinase would opennovel routes for increasing crop yield through stimulation ofribosomogenesis and subsequent growth and division of plantcells. It is known that augmented expression of the At TORgene results in a dose-dependent decrease or increase, in organand cell size, seed production . In addition to regulating theprotein synthesis process, TOR acts as a master regulator ofthe cell cycle, coordinator of rRNA transcription, activation ofribosomal protein genes, ribosome assembly and may also regulate long non-coding RNAs (lncRNAs)expression . Therefore, artificial increasingof TOR gene expression in plant cells can lead to seriousundesirable consequences while using of AtRPS6K2 cDNA may help to avoid these complications.We have cloned the AtRPS6K2 cDNA gene encoding kinase 2of ribosomal protein S6 from A. thaliana and performed itsmutagenesis to obtain the AtRPS6K2kinase containing phosphomimetic substitutions. Such mutatedenzyme with constant RPS6-kinase activity may be usedto study specific molecular mechanisms mediating efficienttranslation of 5\u2032TOP-mRNAs depending on phosphorylationof RPS6 in plant cells. 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+{"text": "New cultivars adapted to major durum wheat growing environments are essential for the cultivationof this crop. The development of new cultivars has required the availability of diverse genetic material and theirextensive field trials. In this work, a collection of tetraploid wheat consisting of 85 accessions was tested in thefield conditions of Almaty region during 2018 and 2019. The accessions were ranged according to nine agronomictraits studied, and accessions with the highest yield performance for Almaty region of Kazakhstan were revealed.The ANOVA suggested that the performance of agronomic traits were influenced both by Environment and Genotype.Also, the collection was analyzed using seven SSR (simple sequence repeats) markers. From 3 to 6 alleles perlocus were revealed, with an average of 4.6, while the effective number of alleles was 2.8. Nei\u2019s genetic diversitywas in the range of 0.45\u20130.69. The results showed high values of polymorphism index content (PIC) in the rangeof 0.46\u20130.70, with an average of 0.62, suggesting that 6 out of 7 SSRs were highly informative (PIC > 0.5). Phylogeneticanalysis of the collection has allowed the separation of accessions into six clusters. The local accessions werepresented in all six clusters with the majority of them grouped in the first three clusters designated as A, B, and C,respectively. The relations between SSR markers and agronomic traits in the collection were studied. The resultscan be efficiently used for the enhancement of local breeding projects for the improvement of yield productivityin durum wheat. Othertetraploid wheat species Triticum turgidum L. ssp. turanicum(Jakubz.) \u00c1. L\u00f6ve & D. L\u00f6ve, Triticum turgidum L. ssp. polonicum(L.) Thell., Triticum turgidum L. ssp. carthlicum(Nevski) \u00c1. L\u00f6ve & D. L\u00f6ve, Triticum turgidum ssp. dicoc-cum(Shrank ex Sch\u00fcbler) Thell. are used as food and feedcrops in different world regions. Wild species Triticum turgidumssp. dicoccoides (Korn. ex Asch. & Graebn.) Thell. isalso often included in crossing schemes as a source for resistanceto abiotic and biotic stresses .Durum wheat (Triticum turgidum L. ssp. turgidum convar. durum(Desf.) MacKey) is a tetraploid species of wheat and is themain crop to producers of pasta and cereals. The growing areaunder durum wheat is about 17 million hectares in the worldand production is 37 million tons . In 2019, durum wheat production in Kazakhstanamounted to 560 thousand tons . Hence, the comprehensive study ofthe diverse germplasm is a very important prerequisite forthe successful conservation and rational use of plant geneticresources, including both wild and cultivated tetraploid wheatspecies . Theappropriate assessment of the genetic diversity in these collectionsdepends on the application of informative and efficienttypes of DNA markers. In many centers of the world,research is underway to find and use different types of DNAmarkers with the aim of using them to study genetic diversity,inventory, genotyping, mapping, and identifying genesassociated with useful traits of cultivated plant varieties andlines . Various types of DNA markershave been developed and are successfully used to study thegenetic diversity of accessions of the genus Triticum L. . PCR-basedmarkers,such as RAPD, AFLP, and SSR, are widely used toolsfor studying genetic diversity and discrimination both durumand common wheat .The wheat genome contains a class of specific nucleotide sequencescalled microsatellites, also known as SSRs or simplesequences repeats . SSR markers havemany advantages, being highly polymorphic, codominant,informative, reliable, and the availability of information onchromosomal localization . Microsatellites are hypervariable, they often havedozensof alleles at one locus, differing from each other inthe number of repeats. They are widely used to study geneticdiversity, as well as for the analysis of paternity and mappingof quantitative trait loci (QTLs), kinship, belonging to aspecific population, for studying hybridization, evolutionaryprocesses, and for searching for paralogs .Durum wheat polymorphism studies are currently underwayworldwide. The survey of reports demonstrated the successfuluse of SSR markers for assessment of the genetic diversityin different collections of Europe , Africa , China , Russia, Turkey ,Syria , etc. Microsatellites are also highlyeffective in tagging specific genes that play an important rolein variation for yield components and biotic stress resistance.A number of studies reported relations between SSR lociand wheat traits, such as yield, etc. For instance, Zhang etal. (2013) showed that the Xgwm11-1B locus is significant( p < 0.001) for plant height. In the study reported by Li etal. (2015) it was shown that the marker Xgwm148-2B is associatedwith the manifestations of the traits \u201cthousand grainweight\u201d, \u201cspike yield index\u201d and \u201cweight of kernels perspike\u201d. Xgwm251 was associated with lipoxygenase (LOX)activity, which is an important factor determining the colorof flour and end-use products of wheat .Vinod et al. (2014) have identified the significant associationbetween Xgwm234 and the resistance of T. turgidum to leafrust. Golabadi et al. (2011) showed that the Xcfa2114-6Amarker was responsible for 20 % of the phenotypic variationin the yield index and thousand grain weights (TGW) underdifferent environmental conditions. SSR marker Xgwm219was also shown to be associated with TGW . These examples suggest that the assessment of thegenetic diversity of the varietal gene pool of durum wheatmay provide not only proper genetic documentation of theaccessions but also hinting the identification of a valuablesource of genes associated with agronomic traits.The purpose of this work was the study the genetic diversityusing seven SSR markers and phenotypic variation in yieldcomponents in the collection of tetraploid species harvestedin the conditions of South-East Kazakhstan.1. Seeds were provided bythe Research Center for Grain and Industrial Crops , University of Bologna , Aktobe andKarabalyk Agricultural Experimental Stations (Kazakhstan).The collection included 21 cultivars and 15 promising linesof durum wheat from Kazakhstan (see Suppl. Table 1).Plant material and experimental site conditions. The plantmaterial consisted of 85 accessions of tetraploid wheat (2 Triticumturgidum ssp. dicoccoides (Korn. ex Asch. & Graebn.)Thell., 2 Triticum turgidum ssp. dicoccum (Shrank ex Sch\u00fcbler)Thell., 65 Triticum turgidum L. ssp. turgidum convar. durum(Desf.) MacKey, 10 Triticum turgidum L. ssp. turanicum(Jakubz.) \u00c1. L\u00f6ve & D. L\u00f6ve, 4 Triticum turgidum L. ssp. polonicum(L.) Thell., and 2 Triticum turgidum L. ssp. carthlicum(Nevski) \u00c1. L\u00f6ve & D. L\u00f6ve from different geographicalorigins (Supplementary Table 1)1 Supplementary Tables 1 & 2 are available in the online version of the paper:http://www.bionet.nsc.ru/vogis/download/pict-2020-24/appx9.pdfThe studied collection of tetraploid wheat was evaluated intwo randomized replicates in the field conditions of Almatyregion (Table 1).Each accession was planted in two rows with a row spacingof 15 cm, 25 seeds per row. In total, nine agronomic traits connected with the vegetation period, plant morphology, and yieldcomponents were studied. The list of traits included the headingtime , flowering time , seed maturationtime , plant height , spike length , number of fertile spikes , number of kernelsper spike , thousand kernel weight , andyield per plant  .DNA extraction and SSR genotyping. Genomic DNAwas isolated from individual 4-day-old wheat seedlings, accordingto Dellaporta et al. (1983). The quality and quantityof isolated DNA were evaluated using a NanoDrop 2000 and agarose electrophoresisin 1 % gel. The list of markers used for SSR analysis wasthe following: Xgwm11, Xgwm148, Xgwm251, Xgwm234,Xcfa2114, Xgwm169, and Xgwm219 (Supplementary Table 2).Polymerase chain reaction (PCR) was conducted in a Veriti\u2122Thermal Cycler . The PCRreaction mixture (10 \u03bcl) contained from 2.5 mM of 10\u00d7 Taqbuffer; 0.2 mM of each dNTP; 1.5 mM MgCl2; 250 \u03bcM ofeach primer; 1 unit Taq polymerase  and50 ng of genomic DNA.The amplification program included the following cycles:94 \u00b0C \u2013 3 min; 40 cycles: 94 \u00b0C \u2013 1 min; annealing temperature(55 or 60 \u00b0C depending on the primer) \u2013 1 min; 72 \u00b0C \u20132 min; and 72 \u00b0C \u2013 10 min. PCR products were separated on6 % polyacrylamide gels  run in 0.5\u00d7TBE buffer pH 8.0 at 250 V for 1.5 h. Gels were stained withethidium bromide, and the images were recorded with a Bio-Rad Image System . Allele sizes wereestimated in comparison with 100 bp DNA ladder .http://software.dell.com/products/statistica).Statistical analyses of field data were estimated usingSPSS 22.0 and STATISTIKA 13.2 software . Thevalues of the PIC index (polymorphism information content)suggested the effectiveness of the markers used, giventhat markers with a value of PIC > 0.5 considered as highlyinformative; 0.5 > PIC > 0.25 as informative; and PIC \u2264 0.25as marginally informative . Variationamong populations was studied using Principal CoordinateAnalysis (PCoA) in the software GenAlex, ver.6.5 . The resulting similarity matrix was furtheranalyzed using the neighbor-joining clustering algorithm forthe construction of the dendrogram. The phylogenetic treewas constructed using PAST v.3.25 software . Analyses of marker-trait associations were conductedusing a simple t-test .Phenotypic variation in the studied collectionField trials for two years revealed a sharp difference inthe vegetationperiod between species of tetraploid wheat(Table 2).All accessions reached the ripening stage, with an exceptfor the wild accession PI346783 .The shortest HT was observed in genotypes of T. dicoccoides(56.5 \u00b1 3.5 days), the longest \u2013 in T. polonicum(60.7 \u00b1 3.9 days) (see Table 2).Plant height is one of the important morphological traitsof the crops. According to the species, the highest ones werethe samples from T. carthlicum (117.9 \u00b1 5.4 cm), while theaccessions from T. dicoccum were the lowest (97.4 \u00b1 7.4 cm).On the other hand for T. durum genotypes the PH rangedfrom 58.0 \u00b1 3.7 cm  to 137.6 \u00b1 3.0 cm for cultivarKargala 66 (see Suppl. Table 1). As for the SL, the lowestvalue (5.0 \u00b1 0.2 cm) had the cultivar PI 184526 , while the highest value (17.5 \u00b1 1.7 cm) wasin accession PI 210845 (T. polonicum from Iran).The value of a cultivar is determined by its productivity,which consists of several components, including TKW whichis significantly affected by weather conditions, violation ofmoisture supply, and mineral nutrition of plants during theformation and maturation of grain. The highest averagedTKW values were revealed for three T. turanicum accessions and T. polonicumfrom Iraq (PI 208911 \u2013 61.8 \u00b1 4.5 g). The lowestTKW value was in accessions of T. carthlicum (29.9 \u00b1 1.1 g).The NFS ranged from 3.9 \u00b1 0.6 pcs/plant in the accessionPI 343446 (T. dicoccoides) to 2.0 \u00b1 0.5 pcs/plant in genotypesPI 210845 and PI 266846 of T. polonicum.As for NKS and YPP the highest value were on accessionsof T. durum and the lowest to T. dicoccoides (see Table 2). Themin value of NKS (24.8 \u00b1 3.8 pcs) under both conditions wasobtained in PI 343446 , the max \u2013 inKazakh cultivar Gordeiforme 254 (67.7 \u00b1 7.1 pcs) and Canadiancultivar Strongfield (62.2 \u00b1 1.2). Overall 31 T. durumaccessions prevailed the local check cultivar Gordeiforme 254(4.4 \u00b1 1.6 g/plant) by YPP. Top twenty accessions by yieldcontained cultivars from Canada (Strongfield \u2013 7.6 \u00b1 1.9 g/plant), Spain (Granizo \u2013 7.0 \u00b1 1.9 g/plant), Italy (Capeiti-8and Ancomarzio), Syria (Sharm5), Russia , Ukraine (Har\u2019kovskaya 90 andHar\u2019kovskaya 9), USA (LO92), as well as 5 cultivars and4 breeding lines (e. g. G 2607 \u2013 7.2 \u00b1 1.4 g/plant), from Kazakhstan(see Suppl. Table 1).The Pearson index analysis revealed a significant positivecorrelation ( p < 0.01) between yield components and phenotypictraits. The ANOVA test based on two-years field trialssuggested that Genotype significantly influenced the SMT,NFS, SL, and all yield components with p < 0.001 (Table 3).Microsatellite analysis of the tetraploid wheat collectionThe lines and cultivars of the studied tetraploid wheat collectionwere analyzed using 7 polymorphic microsatellite markers(see Suppl. Table 2) localized on 6 wheat chromosomes \u2013 1B,2B, 4B, 5B, 6A, 6B. The results based on using 7 SSR markershave allowed identifying a total of 32 alleles, with average4.57 alleles per marker (Table 4).The effective number of alleles ranged from 1.82 to 3.27,with a mean value of 2.77. Nei\u2019s genetic diversity index averaged0.62 (see Table 4). The average value of polymorphisminformation content (PIC) was 0.62, ranging from 0.46 forXgwm219 to 0.7 for Xgwm148, Xgwm251, and Xgwm11,respectively.The PCoA was conducted based on SSR genotyping of85 tetraploid wheat accessions using 7 SSR markers. Accessionsof the studied collection were divided into groupsdepending on their attribution to species and place of origin,respectively .The first principal component in the PCoA (46.31 %) clearlyseparated T. polonicum and T. turanicum from other species. The most genetically distant from other specieswas T. carthlicum. PCoA using origin data revealed that localgenotypes were genetically closer to the North American accessions. The accessions from Russia and NorthAfrica were genetically distant from other groups of origin.Based on the genetic diversity results using 7 polymorphicSSR markers, a phylogenetic tree of 85 accessions of tetraploidwheat was constructed .The analysis revealed a division into two large clusters. Thefirst cluster consisted mostly of cultivars of tetraploid wheatfrom Kazakhstan and North America. The second cluster wasdivided into three sub-clusters. Although the European accessionswere dominated in all three subclusters of cluster 2, allthree sub-clusters included cultivars and lines of Kazakhstan.The t-test was performed to confirm the significance of theSSR markers for the studied traits. The results identified themost informative SSR markers related to major agronomictraits (Table 5). Xgwm251 showed a significant relationshipto HT and FT. Four markers were related to variance in PH.DiscussionInitially, the studied collection was separated according totheir species classification and origin (see Suppl. Table 1).The average yield analysis in the collection of tetraploid accessionsover two years (2018 and 2019) suggested that it ishighly correlated with all studied phenotypic traits ( p < 0.01), confirming the importance of selected characters in the trials.The two-way ANOVA showed that Environment greatly influencedHT and SMT. In addition, it was found that SMT isalso influenced by Genotype, showing the prospects of possibilityto adjust maturation time in the breeding process, asearly seed maturation is vital to avoid abiotic stresses duringthe important stages of plant growth. Particularly, it was shownthat in T. polonicum the seeds are ripening nearly five daysearlier than in T. durum (see Table 2). The field trials haveallowed the identification of accessions with outstanding fieldperformances. For instance, the cultivar Strongfield (Canada)showed 7.6 \u00b1 1.9 g/plant, which was the highest yield valueamong 31 T. durum accessions that prevailed local standardGordeiforme 254 (4.4 \u00b1 1.6 g/plant). In general, two-wayANOVA indicated the great influence of the environmentalfactors, as they were affected both adaptation-related traits,such as HT and SMT, and yield components, such as SL andNKS (see Table 3).The entire collection was studied using seven SSR markersthat were located on six different chromosomes (see Suppl.Table 2). According to the previous works, a list of markers inthis study was most useful to evaluation of genetic diversityand associations with agronomic traits of durum wheat . The average PIC value was higher than 0.6,suggesting that the level of polymorphism was very high.The high level of variation in the collection has effectivelyallowed the separation of accessions according to their speciesclassification . Notably, the PC1 (46.3 %)separated T. polonicum and T. turanicum from the remainingspecies, and the PC1 (34.1 %) distinguished T. carthlicum andT. durum from T. dicoccum and T. dicoccoides. Interestingly,the accessions originated in Kazakhstan were geneticallyclose to North American samples , and it is tosome extent confirm the phylogeny of hexaploid bread wheatstudies using SNP (single nucleotide polymorphism) markers. The PC plot is suggesting that sixaccessions of durum wheat from the Russian Federation aredistinctly different from accessions with other origins . The Neighbor-joining phylogenetic tree suggestedthat all accessions can be divided into two clusters, wherecluster 1 was mostly populated by accessions from Kazakhstan.The significance of each SSR marker for studied traits wasassessed using a two-tailed t-test . The results of the test suggested that five out ofseven SSRs were significant at least for one studied trait (seeTable 5). The PH was the trait where four SSR markers, twowith negative and two with positive values, were significantlycorrelated. In addition, the test showed that Xgwm234 is significantlycorrelated with TKW and Xgwm219 and Xgwm169with YPP (see Table 5). Thus, the application of SSR markersin the analysis of tetraploid wheat collection consistingof 85 accessions was used for (1) genetic documentation ofsamples, (2) for phylogenetic clusterization based on the speciesclassification and geographic origin, and (3) associationsbetween DNA markers and studied phylogenetic traits. Hence,the results can be efficiently used for the enhancement of localbreeding projects for the improvement of yield productivityin durum wheat.The phenotypic analysis of the tetraploid wheat collectionconsistingof 85 accessions showed a high correlation ofYPP with all 8 phenotypic traits in conditions of South-EastKazakhstan. The ANOVA suggested that the environmentalconditionssignificantly affected the variation in HT and SMT,while Genotype has contributed significantly to main yieldcomponents, including TKW. Overall, 31 accessions of T. durumshowed higher average yield values in comparison withlocal check cultivar Gordeiforme 254 (4.4 \u00b1 1.6 g/plant), andCanadian cultivar Strongfield was with the highest yield value(7.6 \u00b1 1.9 g/plant). The application of seven SSR markerssuggested that local accessions were distinctly different fromdurum accession from other parts of the world. Particularly,the Principal Coordinate plot showed that local durum sampleswere most close to North American samples. The Neighborjoiningphylogenetic tree separated 85 samples to two mainclusters, where the cluster 1 was mainly represented by Kazakhaccessions and cluster 2 mostly by European accessions. Theapplication of the t-test indicated that five out of seven SSRswere significant at least with one agronomic trait. 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DOI 10.5958/j.0975-6906.74.1.002.Wang H., Wang X., Chen P., Liu D. Assessment of genetic diversityof Yunnan, Tibetan, and Xinjiang wheat using SSR markers.J. Genet. Genom. 2007;34(7):623-633. DOI 10.1016/S1673-8527(07)60071-X.Yildirim A., S\u00f6nmezo\u011flu \u00d6z., G\u00f6kmen S., Kandemir N., Aydin N. Determinationof genetic diversity among Turkish durum wheat landracesby microsatellites. Afr. J. Biotechnol. 2011;10(19):3915-3920.DOI 10.5897/AJB10.2240.Za\u00efm M., El Hassouni Kh., Gamba F., Filali-Maltouf A., Belkadi B.,Sourour A., Amri A., Nachit M., Taghouti M., Bassi F.M. Widecrosses of durum wheat (Triticum durum Desf.) reveal good diseaseresistance, yield stability, and industrial quality across Mediterraneansites. Field Crops Res. 2017;214:219-227. DOI 10.1016/j.fcr.2017.09.007.Zhang B., Shi W., Li W., Chang Xi, Jing R. Efficacy of pyramidingelite alleles for dynamic development of plant height in commonwheat. Mol. Breed. 2013;32:327-338. DOI 10.1007/s11032-013-9873-5."}
+{"text": "Caseinolytic protease P (ClpP), which is located on the inner mitochondrial membrane, degrades mitochondrial proteins damaged by oxidative stress. The role of ClpP varies among tumor types. However, the expression pattern and biological functions of ClpP in breast cancer (BC) have not yet been investigated.The Cancer Genome Atlas (TCGA) and Kaplan Meier-plotter database were used to analyze the expression level of ClpP in BC tissues, relationships with clinicopathological characteristics, and the influence on the prognosis of BC. Protein and mRNA expression levels of ClpP in BC cell lines and tissues were detected by quantitative real-time PCR, western blot and immunohistochemical (IHC) analyses. The colony formation assay, transwell assay and flow cytometric analysis were performed to assess various functions of ClpP. Western blot analysis was also conducted to determine the mechanism of ClpP.ClpP expression was markedly increased in BC cells and tissues. High expression of ClpP was significantly correlated with the T stage, estrogen receptor (ER) expression, and poor recurrence-free survival (RFS) in TCGA and Kaplan Meier-plotter database. ClpP silencing significantly inhibited proliferation, migration, invasion, and promoted apoptosis of BC cells, which resulted in suppression of the Src/PI3K/Akt signaling pathway. The gain-of-function assay confirmed partial these results. Breast cancer (BC) is the most common malignant tumor among women worldwide . At presThe mitochondria have been a focus of cancer research since the 1950s, when it was discovered that cancer cells produce adenosine triphosphate (ATP) and employ different mechanisms to support cell growth than the normal surrounding tissues. Therefore, it was suggested that a defect in the mitochondrial mechanism will not only lead to increased glycolysis, but also to the transformation of normal cells into cancer cells . The maiin vivo, and correlated with shortened survival. However, the ClpP and ClpX subunits may not have completely overlapping function(s) in the tumor mitochondria (The ClpXP protease complex is composed of two proteins: hexamers of a AAA+ ATPase (ClpX) and the tetradecameric peptidase caseinolytic protease P (ClpP) , which cchondria .ClpP is encoded by nuclear genes in mammalian cells and plays a central role in the quality control of mitochondrial proteins via the degradation of misfolded proteins. ClpP was first identified in bacteria and has since aroused interest as a potential anti-microbial therapeutic target . StudiesThe expression level of ClpP is greater in acute myeloid leukemia (AML) cells than in normal hematopoietic cells . FurtherHuman BC tissues and corresponding adjacent normal tissues were collected from patients who underwent surgery at the First Affiliated Hospital of Chongqing Medical University from 2014 to 2018, snap-frozen in liquid nitrogen, and then stored at \u221280\u00a0\u00b0C. This study was approved by the Institutional Ethics Committees of the First Affiliated Hospital of Chongqing Medical University  and conducted in accordance with the tenets of the Declaration of Helsinki. Each participant signed an informed consent form prior to study inclusion.2/95% air.Seven human BC cell lines  and two normal mammary epithelial cell lines (MCF-10A and HBL-100) were obtained from the American Type Culture Collection . MCF-10A cells were cultured as described previously  and all ClpP-specific and the negative control siRNAs were synthesized by OriGene . The following siRNAs sequences were generated: SR305388A-rGrCrUrCrArArGrArArGrCrArGrCrUrCrUrArUrArArCrATC; SR305388B-rGrUrUrUrGrGrCrArUrCrUrUrArGrArCrArArGrGrUrUrCTG; and SR305388C-rGrGrCrCrArUrCrUrArCrGrArCrArCrGrArUrGrCrArGrUAC. The expression vector pCMV6-Myc-DDK-ClpP was produced by OriGene ; the empty pCMV6-Myc-DDK vector was used as a control. Lipofectamine 2000  was used for siRNAs and plasmids transfection, in accordance with the manufacturer\u2019s protocols.Total RNA was extracted from cells and tissues with TRIzol reagent (Invitrogen) in accordance with the manufacturer\u2019s instructions. RT-qPCR was performed using an ABI 7500 Real-Time PCR System  with the SYBR Green kit (Invitrogen). \u03b2-actin was used as an internal control. Each sample was tested in triplicate. The followig primers were used for RT-qPCR analysis: ClpP, forward primer: 5\u2032-GCC AAG CAC ACC AAA CAG A-3\u2032, reverse primer: 5\u2032-GGA CCA GAA CCT TGT CTA AG-3\u2032; \u03b2-actin, forward primer: 5\u2032-CCT GTG GCA TCC ACG AAA CT-3\u2032, reverse primer: 5\u2032-GAA GCA TTT GCG GTG GAC GAT- 3\u2032.Total proteins were extracted with radioimmunoprecipitation assay lysis buffer . Protein concentrations were determined using the bicinchoninic acid protein assay kit . Western blot analysis was conducted as previously described  with theAll specimens were formalin-fixed, paraffin-embedded and cut into 4 \u00b5m-thick sections, which were mounted onto glass slides. IHC analysis was conducted as described previously . BrieflyCell apoptosis was detected with the Annexin V-FITC Apoptosis Detection Kit  in accordance with the manufacturer\u2019s protocol and a FACSCalibur flow cytometer (BD Biosciences).MDA-MB-231 and ZR-75-1 cells were seeded into triplicate wells of 6-well plates at 1,000 cells/well, and cultured for 7 days. The cells were then fixed with 4% paraformaldehyde and stained with 0.1% crystal violet solution .4 cells/well) were added into the upper transwell chamber, and 800 \u00b5L of medium containing 10% FBS were added to the lower chamber. After incubation at 37\u00a0\u00b0C under an atmosphere of 5% CO2/95% air for 24 h (MDA-MB-231 and ZR-75-1 cells), or 72\u00a0h (MCF-7 and T47D cells), cells that migrated through the membrane pores were fixed in 4% paraformaldehyde for 30 min and stained with 0.1% crystal violet  for 15 min at room temperature. Matrigel\u2122-coated transwell filters  were used to evaluate the invasion capability of the cells. The subsequent procedures were the same as those for the cell migration assay. Cells from six random fields were counted under a microscope. All experiments were repeated three times.Transwell chambers  were used to detect the migratory and invasive capabilities of BC cells. For the transwell migration assays, 200-\u00b5lL aliquots of transfected MDA-MB-231, ZR-75-1, MCF-7 and T47D cells (4 * 10https://tcga-data.nci.nih.gov/tcga/). The study cohort consisted of a total of 1010 BC patients. ClpP expression data of 112 normal breast samples were included to compare differences in ClpP expression levels in BC tissues. Overall survival (OS) and complete clinicopathological data of 990 BC patients, and RFS data of 792 BC patients were screened. ClpP expression levels were ranked from low to high based on median values. The first 50% of patients were considered as the low-expression group and the second 50% as the high-expression group.Gene expression data of BC tissues were downloaded from the TCGA database .Prognosis based on ClpP levels in BC patients was analyzed using the Kaplan Meier-plotter database  and GraphPad Prism 7.0 software . The two-tailed Student\u2019s p\u00a0<\u00a00.001) (p\u00a0<\u00a00.0001) (p\u00a0<\u00a00.01) (p = 0.029) (p\u00a0<\u00a00.001) (p = 0.0154) and ER expression (p = 0.0164). Kaplan\u2013Meier survival curves from TCGA indicated that ClpP was not associated with RFS (p = 0.506) or OS (p = 0.619) (p = 0.00071)  . Receive\u00a00.0001) . To veri\u00a0<\u00a00.01) . The AUC= 0.029) . ClpP pr<\u00a00.001) \u20131G. Thes<\u00a00.001) . High Cl= 0.619)  and 2B. 0.00071)  and 2D. ClpP mRNA and protein expression levels in a panel of BC and normal breast epithelial cell lines were determined. The results showed that ClpP was increased in most malignant cell lines, as compared with normal cells  and 3B. p\u00a0<\u00a00.01 and <0.001, respectively) (The suppressive effect of ClpP silencing on cancer cell growth was confirmed by the colony formation assay. The colony formation capabilities of the BC cell lines MDA-MB-231 and ZR-75-1 were markedly inhibited by si-ClpP-B (ctively) \u20134E.The transwell migration and invasion assay was performed to quantitatively assess cell metastasis and invasiveness. The results showed that cell migration and invasion were significantly reduced in cells transfected with si-ClpP-B, as compared with control cells, due to the down-regulation of MMP7 and vimentin, and the up-regulation of E-cadherin \u20135T. Convp\u00a0<\u00a00.01 and <0.001, respectively), accompanied by increased levels of cleaved caspase-9, cleaved caspase-8 and cleaved poly (ADP-ribose) polymerase (PARP) (Flow cytometry was carried out to detect cell apoptosis. The percentages of apoptotic MDA-MB-231 and ZR-75-1 cells were increased by si-ClpP-B (e (PARP) \u20135N.To confirm the specificity of ClpP siRNAs, the same functional experiments were performed with a second siRNA (si-ClpP-A). The results of the loss-of-function studies were consistent .These data revealed the anti-proliferative, anti-migration, anti-invasion and pro-apoptotic roles of silencing ClpP in BC cells.in vivo by increasing phosphorylation of the key cellular kinases Akt and Src  of 14 analyzed datasets. Importantly, high ClpP expression was correlated with shorter metastasis-free survival in BC patients and reduced RFS in those with lung adenocarcinoma . ConsistClpP is a subunit of the ClpXP complex. The Akt signaling pathway is important in the regulation of various cellular functions, including metabolism, growth, proliferation, survival, transcription, protein synthesis and tumorigenesis . MutatioIn summary, the present study provides the first evidence that ClpP is frequently up-regulated in BC and that ClpP has high diagnostic value.ClpP expression is markedly increased in BC and significantly correlated with the T stage, ER expression, and poor RFS. Silencing of ClpP significantly inhibited proliferation, migration and invasion, and promoted apoptosis of BC cells, resulting in suppression of the Src/PI3K/Akt signaling pathway. These data indicate that ClpP is an oncogene, and may be a promising diagnostic biomarker and therapeutic target in BC.10.7717/peerj.8754/supp-1Figure S1p < 0.01, ***p < 0.001.(A) The overexpressing efficiency in ClpP (MCF-7 and T47D cells) was evaluated by RT-qPCR.  The effects of overexpressed-ClpP on BC cell migration were evaluated using transwell assays.  The effects of overexpressed-ClpP on BC cell invasion were evaluated using transwell assays. The data are presented as mean \u00b1 SD, **Click here for additional data file.10.7717/peerj.8754/supp-2Figure S2p < 0.01, ***p < 0.001. The effects of si-ClpP-A on BC cell proliferation were analyzed using the colony formation assay.  The effects of si-ClpP-A on BC cell migration were evaluated using the transwell assay.  The effects of si-ClpP-A on BC cell invasion were evaluated using the transwell assay.  The effect of si-ClpP-A on MDA-MB-231 cells apoptosis was analyzed using flow cytometry. The data are presented as the mean \u00b1 SD, **Click here for additional data file.10.7717/peerj.8754/supp-3Supplemental Information 3Click here for additional data file.10.7717/peerj.8754/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.8754/supp-5Supplemental Information 5Click here for additional data file.10.7717/peerj.8754/supp-6Supplemental Information 6Click here for additional data file.10.7717/peerj.8754/supp-7Supplemental Information 7Click here for additional data file.10.7717/peerj.8754/supp-8Supplemental Information 8Click here for additional data file.10.7717/peerj.8754/supp-9Supplemental Information 9Click here for additional data file.10.7717/peerj.8754/supp-10Supplemental Information 10Click here for additional data file.10.7717/peerj.8754/supp-11Supplemental Information 11Click here for additional data file.10.7717/peerj.8754/supp-12Supplemental Information 12Click here for additional data file.10.7717/peerj.8754/supp-13Supplemental Information 13Click here for additional data file.10.7717/peerj.8754/supp-14Supplemental Information 14Click here for additional data file.10.7717/peerj.8754/supp-15Supplemental Information 15Click here for additional data file.10.7717/peerj.8754/supp-16Supplemental Information 16Click here for additional data file.10.7717/peerj.8754/supp-17Supplemental Information 17Click here for additional data file.10.7717/peerj.8754/supp-18Supplemental Information 18Click here for additional data file.10.7717/peerj.8754/supp-19Supplemental Information 19Click here for additional data file.10.7717/peerj.8754/supp-20Supplemental Information 20Click here for additional data file.10.7717/peerj.8754/supp-21Supplemental Information 21Click here for additional data file.10.7717/peerj.8754/supp-22Supplemental Information 22Click here for additional data file.10.7717/peerj.8754/supp-23Supplemental Information 23Click here for additional data file.10.7717/peerj.8754/supp-24Supplemental Information 24Click here for additional data file.10.7717/peerj.8754/supp-25Supplemental Information 25Click here for additional data file.10.7717/peerj.8754/supp-26Supplemental Information 26Click here for additional data file.10.7717/peerj.8754/supp-27Supplemental Information 27Click here for additional data file.10.7717/peerj.8754/supp-28Supplemental Information 28Click here for additional data file.10.7717/peerj.8754/supp-29Supplemental Information 29Click here for additional data file.10.7717/peerj.8754/supp-30Supplemental Information 30Click here for additional data file.10.7717/peerj.8754/supp-31Supplemental Information 31Click here for additional data file.10.7717/peerj.8754/supp-32Supplemental Information 32Click here for additional data file.10.7717/peerj.8754/supp-33Supplemental Information 33Click here for additional data file.10.7717/peerj.8754/supp-34Supplemental Information 34Click here for additional data file.10.7717/peerj.8754/supp-35Supplemental Information 35Click here for additional data file.10.7717/peerj.8754/supp-36Supplemental Information 36Click here for additional data file.10.7717/peerj.8754/supp-37Supplemental Information 37Click here for additional data file.10.7717/peerj.8754/supp-38Supplemental Information 38Click here for additional data file.10.7717/peerj.8754/supp-39Supplemental Information 39Click here for additional data file.10.7717/peerj.8754/supp-40Supplemental Information 40Click here for additional data file.10.7717/peerj.8754/supp-41Supplemental Information 41Click here for additional data file.10.7717/peerj.8754/supp-42Supplemental Information 42Click here for additional data file.10.7717/peerj.8754/supp-43Supplemental Information 43Click here for additional data file.10.7717/peerj.8754/supp-44Supplemental Information 44Click here for additional data file.10.7717/peerj.8754/supp-45Supplemental Information 45Click here for additional data file.10.7717/peerj.8754/supp-46Supplemental Information 46Click here for additional data file.10.7717/peerj.8754/supp-47Supplemental Information 47Click here for additional data file.10.7717/peerj.8754/supp-48Supplemental Information 48Click here for additional data file."}
+{"text": "Acanthosaura have been recorded so far, including Acanthosauraarmata from the southern region, A.cardamomensis from the eastern region, A.crucigera from the western region, A.lepidogaster from the northern region and A.phuketensis from the Phuket Island and south-western region. However, comprehensive studies of diversity patterns and distribution of Acanthosaura are still lacking in some areas and need further information for designating areas of special conservation importance and nature protection planning in Thailand.In Thailand, five species of Acanthosauraaurantiacrista is a new species of long-horned lizard of the genus Acanthosaura from northern Thailand. It is distinguished from all other species of Acanthosaura by a dagger-like nuchal spine with yellowish-orange colouration in females, bright yellow colouration in males and a combination of other morphological characters: a greater tail length to snout-vent length ratio; a larger postorbital spine, nuchal spine, dorsal spine and occipital spine compared to its head length; a smaller diastema to snout-vent length ratio; a greater number of subdigital lamellae on the fourth finger and fourth toe; and a larger gular pouch than other Acanthosaura species. Analysis of mitochondrial ND2 gene sequences revealed a sister clade between the A.aurantiacrista lineage and the A.crucigera lineage with a 100% probability of divergence, according to Bayesian analysis and strong support value for Maximum Likelihood analysis. The pairwise distance ranged from 13.8-15.0% between A.aurantiacrista and A.cardamomensis, 10.9-14.5% between A.aurantiacrista and A.crucigera and 0-1.2% amongst A.aurantiacrista populations. The discovery of this lizard increases the known endemic herpetological diversity and underscores the importance of conservation in the mountain rainforest region of northern Thailand. Acanthosaura Gray, 1831 are recognised, ranging from Myanmar, east through Thailand, Cambodia, Laos, Vietnam and southern China and southwards through the Malaysian Peninsula and archipelagoes , A.capra , A.cardamomensis Wood, Grismer, Grismer, Neang, Chav & Holden, 2010, A.murphyi Nguyen, Do, Hoang, Nguyen, McCormack, Nguyen, Orlov, Nguyen & Nguyen, 2018, A.nataliae Orlov, Truong, & Sang, 2006 and A.phuketensis Pauwels, Sumontha, Kunya, Nitikul, Samphanthamit, Wood & Grismer, 2015 are classified as a long-horned group exhibiting the largest nuchal and dorsal spines with lengths greater than 10 mm; A.bintangensis Wood, Grismer, Grismer, Northayati, Chan & Bauer, 2009, A.brachypoda Ananjeva, Orlov, Nguyen & Ryabov, 2011, A.crucigera Boulenger, 1885, A.lepidogaster , A.phongdienensis Nguyen, Jin, Vo, Nguyen, Zhou, Che, Murphy, & Zhang, 2019, A.titiwangsaensis Wood, Grismer, Grismer, Northayati, Chan & Bauer, 2009 and A.tongbiguanensis Liu & Rao, 2019 belong to a short-horned group exhibiting nuchal and dorsal spines that are shorter than 10 mm; and the nuchal crest is absent in A.coronata G\u00fcnther, 1861  or bright yellow  nuchal crest on the dorsum. To clarify the taxonomic status of this Acanthosaura population, we describe the new species on the basis of its distinctive morphological and colouration characteristics and a genetic analysis. Its name is provided to be included in conservation planning for agamid species in wildlife sanctuaries and national parks in the Thanon Thong Chai Mountain Range.A recent field sampling of Specimens were collected by hand from two locations in Thanon Thong Chai Mountain Range, Thailand: one specimen from Mae Sariang District, Mae Hong Son Province on 27 April 2018; seven specimens from Sop Khong Subdistrict, Omkoi District, Chiang Mai Province on 14 November 2018; and one specimen from Nang Lae Subdistrict, Mueang District, Chiang Rai Province on 24 January 2019. For all specimens, photographs were taken to document their colour patterns prior to euthanisation. Liver samples were taken and stored in absolute ethanol. Specimens were fixed in 10% formalin before being transferred to 70% ethanol for permanent storage.Acanthosaura in Thailand were examined from the Natural History Museum, National Science Museum, Technopolis, Pathum Thani Province (THNHM) and the Queen Saovabha Memorial Institute, Thai Red Cross Society, Bangkok Province, Thailand (QSMI): Acanthosauraarmata from Hala-Bala, Narathiwat Province (THNHM15209), Sungai Kolok District, Narathiwat Province (THNHM18884); Acanthosauracardamomensis from Koh Kut, Trat Province , Khao Yai National Park, Nakhon Ratchasrima Province ; Acanthosauracrucigera from Na Yong District, Trang Province , Taksin Maharat National Park, Muang District, Tak Province , Thong Pha Phum District, Kanchanaburi Province (THNHM22658), Huai Kha Khaeng, Lan Sak District, Uthai Thani Province (THNHM18594); Acanthosauralepidogaster from Phu Luang District, Loei Province , Phu Kieo District, Chaiyaphum Province (THNHM19619), Ban Sun Phae Kae, Chiang Dao District, Chiang Mai Province (THNHM20537), Roi Praputabath, Umphang District, Tak Province (THNHM20647), Huai Na Tee, Pua District, Nan Province (THNHM10080), Doi Khun Tan National Park, Lam Phun Province ; Acanthosauraphuketensis from Ton Sai Waterfall, Thalang District, Phuket Province (THNHM08865), Khao Sok, Ban Ta Khun, Surat Thani Province (THNHM22663); and Acanthosauranataliae from Xe Sap, Samoy, Lao .Comparative materials of all currently recognised species of Acanthosaura were also taken from original descriptions and subsequent studies  obtained from GenBank  was prepared using an EP0402 TAQ DNA POLYMERASE. Two primers, METF6  and ACANTHND2.833. R1 (5\u2019-AGGGAGGTTATTGTTGCTAG-3\u2019), were used to amplify a 698 bp fragment of the NADH dehydrogenase subunit 2 (ND2) gene . PCR proThe ND2 sequences were aligned using ClustalW v.1.83  integratBayesian Interference was performed in Mr.Bayes v.3.1.2 , based o212B9A56-ACDD-536A-958C-C3115F384365urn:lsid:zoobank.org:act:E0D80F05-8884-4426-B2D6-6C939310D2A4Type status:Holotype. Location: country: Thailand (northern region); stateProvince: Mae Hong Son Province; county: Mae Sariang District; verbatimElevation: 728 m; verbatimLatitude: 18\u00b009'02.8\"N; verbatimLongitude: 97\u00b058'50.2\"E; verbatimCoordinateSystem: degrees minutes seconds; verbatimSRS: WGS84; Event: year: 2018; month: April; day: 27; habitat: evergreen forests on hills up to at least 600 m elevation; fieldNotes: collector = Poramad Trivalaira; Record Level: institutionCode: THNHM; collectionCode: 28064; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = adult femaleType status:Paratype. Location: country: Thailand (northern region); stateProvince: Chiang Mai Province; county: Omkoi District; locality: Sop Khong Subdistrict; verbatimElevation: 935; verbatimLatitude: 17\u00b039'45.4\"N; verbatimLongitude: 98\u00b011'53.6\"E; verbatimCoordinateSystem: degrees minutes seconds; verbatimSRS: WGS84; Event: year: 2018; month: November; day: 14; habitat: evergreen forests on hills up to at least 600 m elevation; fieldNotes: collector = Kirati Kunya; Record Level: institutionCode: THNHM; collectionCode: 28521; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = adult femaleType status:Paratype. Record Level: institutionCode: THNHM; collectionCode: 28522; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = adult female, same collection date, collector and location as paratype THNHM28521Type status:Paratype. Record Level: institutionCode: THNHM; collectionCode: 28523; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = subadult male, same collection date, collector and location as the THNHM28521Type status:Paratype. Record Level: institutionCode: THNHM; collectionCode: 28524; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = subadult male, same collection date, collector and location as the THNHM28521Type status:Paratype. Record Level: institutionCode: QSMI; collectionCode: 1446; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = adult female, same collection date, collector and location as the THNHM28521Type status:Paratype. Record Level: institutionCode: QSMI; collectionCode: 1447; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = adult female, same collection date, collector and location as the THNHM28521Type status:Paratype. Record Level: institutionCode: QSMI; collectionCode: 1448; basisOfRecord: PreservedSpecimen; dynamicProperties: sex = adult male, same collection date, collector and location as the THNHM28521Acanthosauraaurantiacrista sp. n.  ; differences of 13.8-15.0% compared to two specimens of A.cardamomensis (GenBank GU817397 and GU817400); differences of 16.2-16.3% compared to two specimens of A.armata (GenBank AB266452 and NC014175); differences of 18.0-19.6% compared to two specimens of A.lepidogaster (GenBank AF128499 and KR092427); and differences of 19.1-19.8% compared to a specimen of A.capra (GenBank AF128498) (Table 4). The phylogenetic relationships within the genus Acanthosaura revealed through Maximum-Likelihood trees and Bayesian Inference tree of the ND2 gene showed high posterior probabilities and high bootstrap support values , more FI (17-23 vs. 13-17), more TO (25-29 vs. 19-26), fewer NS (5-6 vs. 6-10), fewer NCS (11-13 vs. 10-17) and the presence of a BEP and more GP (1-4 vs. 1) , greater PS/HL ratio (0.24-0.84 vs. 0.07-0.19), greater DS/HL ratio (0.15-0.38 vs. 0.08-0.09), fewer DIASN (8-9 vs. 11-15), more VENT (63-66 vs. 51-55), fewer NSSOS (5 vs. 6-7), fewer NS (5-6 vs. 8) and the presence of a LKP.Acanthosauraaurantiacrista sp. n. differs from A.brachypoda in presenting greater PS/HL ratio (0.24-0.84 vs. 0.11), greater DS/HL ratio (0.15-0.38 vs. 0.06), fewer RS (4-6 vs. 5-9), fewer NS (5-6 vs. 9) and more GP (1-4 vs. 0).Acanthosauraaurantiacrista sp. n. differs from A.capra in presenting fewer INFRAL (9-11 vs. 12-13), more FI (17-23 vs. 16-17), more TO (25-29 vs. 22-24), fewer NS (5-6 vs. 9) and the presence of an occipital spine and scales surrounding the occipital spine.Acanthosauraaurantiacrista sp. n. differs from A.cardamomensis in presenting fewer RS (4-6 vs. 7-9), fewer NS (5-6 vs. 7-10) and fewer NSSLC (9-13 vs. 10-19).Acanthosauraaurantiacrista sp. n. differs from A.coronata in presenting greater TaL/SVL ratio (1.40-1.70 vs. 0.60-1.00), more FI (17-23 vs. 13-14), more TO (25-29 vs. 17-19), fewer RS (4-6 vs. 9), fewer NS (5-6 vs. 7-9), fewer NR (1-2 vs. 3-4) and the presence of a postorbital spine, nuchal spine, dorsal spine, diastema, occipital spine, YAS, ND, BEP and more GP (1-4 vs. 0).Acanthosauraaurantiacrista sp. n. differs from A.crucigera in presenting fewer DIASN (8-9 vs. 9-25), more VENT (63-66 vs. 55-63), more FI (17-23 vs. 16-18), more TO (25-29 vs. 21-26), fewer RS (4-6 vs. 7-8), fewer NS (5-6 vs. 7-9) and more GP (1-4 vs. 1-2).Acanthosauraaurantiacrista sp. n. differs from A.lepidogaster in presenting greater TaL/SVL ratio (1.40-1.70 vs. 1.00-1.50), greater PS/HL ratio (0.24-0.84 vs. 0.06-0.17), greater NSL/HL ratio (0.35-0.95 vs. 0.12-0.15), greater DS/HL ratio (0.15-0.38 vs. 0.06-0.15), fewer DIASN (8-9 vs. 10-14), more VENT (63-66 vs. 52-61), more TO (25-29 vs. 22-23), greater OS/HL ratio (0.19-0.44 vs. 0.14-0.15), fewer NS (5-6 vs. 7-8), fewer PM (4 vs. 5), absence of scale on tympanum and more GP (1-4 vs. 0-1).Acanthosauraaurantiacrista sp. n. differs from A.murphyi in presenting greater PS/HL ratio (0.24-0.84 vs. 0.16-0.34), greater NSL/HL ratio (0.35-0.95 vs. 0.24-0.43), fewer INFRAL (9-11 vs. 12-14), more FI (17-23 vs. 15-18), more TO (25-29 vs. 21-23), fewer RS (4-6 vs. 8-9), fewer NS (5-6 vs. 7-8), fewer NR (1-2 vs. 3-4) and the absence of a tympanum scale.Acanthosauraaurantiacrista sp. n. differs from A.nataliae in presenting fewer VENT (63-66 vs. 64-71), fewer RS (4-6 vs. 7), fewer NSSLC (9-13 vs. 13-16) and the presence of an occipital spine, scales surrounding the occipital spine, ND and LKP.Acanthosauraaurantiacrista sp. n. differs from A.phongdienensis in presenting greater PS/HL ratio (0.24-0.84 vs. 0.06-0.09), greater NSL/HL ratio (0.35-0.95 vs. 0.07-0.18), larger DS/HL ratio (0.15-0.38 vs. 0.03-0.07), more FI (17-23 vs. 14-17), more TO (25-29 vs. 19-23) and the presence of a diastema.Acanthosauraaurantiacrista sp. n. differs from A.phuketensis in presenting greater NSL/HL ratio (0.35-0.95 vs. 0.21-0.39), fewer DIASN (8-9 vs. 12-17), more FI (17-23 vs. 15-17), more TO (25-29 vs. 21-24), fewer RS (4-6 vs. 5-9), fewer NS (5-6 vs. 7-8) and more GP (1-4 vs. 1-2).Acanthosauraaurantiacrista sp. n. differs from A.titiwangsaensis in presenting greater PS/HL ratio (0.24-0.84 vs. 0.14-0.18), greater NSL/HL ratio (0.35-0.95 vs. 0.11-0.18), greater DS/HL ratio (0.15-0.38 vs. 0.07-0.09), fewer DIASN (8-9 vs. 10-13), more VENT (63-66 vs. 47-57), fewer NS (5-6 vs. 8) and the presence of a LKP.Acanthosauraaurantiacrista sp. n. differs from A.tongbiguanensis in presenting greater PS/HL ratio (0.24-0.84 vs. 0.13-0.19), greater NSL/HL ratio (0.35-0.95 vs. 0.15-0.21), greater OS/HL ratio (0.19-0.44 vs. 0.16-0.23), fewer RS (4-6 vs. 6-9), fewer NS (5-6 vs. 8-9) and more GP (1-4 vs. 1-2).Acanthosauraaurantiacristasp. n. is differentiated from all other congeners by this combination of characters: A large size  and a single long conical spine above the posterior margin of the eye; a large spine on the occiput between the tympanum and nuchal crest; tympanum naked, large, roundish; large developed gular pouch; scales on flanks randomly intermixed with small keeled and small tubercle scales; large nuchal crest with 8 large dagger-like and pointed spines; narrow diastema with 8-9 scales between the nuchal and vertebral crests; vertebral crest composed of large dagger-like, pointed spines beginning at the shoulder region and decreasing in size until the base of the tail; nuchal and dorsal crests are orange in females and yellow in males; tail 1.40-1.70 times the SVL; and black collar and black eye patch present, extending posteriorly until reaching the nuchal crest.Description of the holotype: Adult female. SVL 105.7 mm; TaL 151.8 mm (1.44 times SVL), tail complete; HL (23.3 mm) slightly longer than HW (18.9 mm); HL one-fifth SVL (0.22 times SVL), HW narrow (0.179 times SVL) and HD tall (0.64 times HL); head triangular in dorsal and lateral views; SL moderately long (0.46 times HL); RW wide (2.31 times RH); steeply sloping anteriorly; CS prominent, forming a large projecting shelf extending above eye, composed of 14/13 large scales; shelf terminates with a notch anterior to postorbital spine; rostrum moderate in size, rectangular, bordered laterally by first SUPRALs and posteriorly by five smaller scales; nostrils roundish, surrounded by one prenasal anteriorly, four postnasals posteriorly and two subnasals; six NS; oval supranasals; large scales above orbit weakly keeled; three rows of moderately-keeled scales below orbit extending from the anterior margin of the eye to posterior; large EYE (0.28 times HL) and ORBIT (0.44 times HL); interorbital, prefrontal and frontal scales slightly keeled and smaller than scales below orbit; seven large, keeled, azygous prefrontal scales arranged in a Y-shaped pattern; parietal eyespot surrounded by a larger row of scales; large conical PS above posterior margin of the eye surrounded by five small lanceolate scales; single row of seven large keeled scales extending from suborbital below posterior margin of eye to above tympanic margin, increasing in size posteriorly; elongated conical OS on lateral margin of nape surrounded by a rosette of five small lanceolate NSSOS; tympanum exposed, roundish, with a size two-thirds that of EYE (0.69 times EYE), surrounded by tiny conical scales; thirteen rectangular SUPRALs similar in size; mental pentagonal, larger than adjacent INFRALs; two postmentals similar in size, four scales contacting PM; chin shields large, extending posteriorly to angle of jaw, separated from infralabials by one scale row anteriorly and three at angle of jaw; eleven rectangular INFRALs of similar size; gular scales sharply keeled and spinose with a larger midventral row; extensible dewlap present; nuchal crest composed of eight elongated, dagger-like scales, bordered on each side by two rows of large, flat, keeled, triangular scales; nuchal crest followed by a diastema of nine DIASN at base of nape; dorsal body crest extending from posterior margin of diastema to base of tail; dorsal crest composed of small laterally compressed, triangular epidermal scales, bordered by a row of smaller paravertebral triangular scales; DS slightly decreasing to sacrum, then fading progressively; and nuchal and dorsal crests present as orange in live specimen  and OS (5.3-7.5 vs. 4.5) and greater NSL (9.6-12.7 vs. 8.3), wider WNC (0.9-1.6 vs. 0.6), more SUPRAL (10-11/10 vs. 13/13) and fewer number of FI (18-20/17-20 vs. 23/21), higher number of NSSLC (10 vs. 13) in QSMI1446, QSMI1447, THNHM28521 and THNHM28522, and larger GP (3 vs. 1) in QSMI1446 and THNHM28522. The adult male paratype (QSMI1448) differs from the adult female holotype in presenting a longer SVL (130.1 vs. 105.7), greater TaL (202.2 vs. 151.8) and SL (22.3 vs. 10.7), wider TBW (19.2 vs. 10.3), greater HD (21.7 vs. 14.9), larger ORBIT (8.5 vs. 6.4), longer PS (19.1 vs. 5.7), OS (10 vs. 4.5), greater NSL (21.6 vs. 8.3), FOREL (54.2 vs. 49.8), HINDL (71.4 vs. 59.7) and GP (4 vs. 1) size, wider WNC (2.9 vs. 0.6), narrower DIAS (3.5 vs. 5.4) and fewer number of SUPRAL (10 vs. 13) and FI (19/18 vs. 23/21). Two subadult male paratypes are smaller and present fewer differences in morphological characters compared with the holotype, except for a longer PS (5.5-7.5 vs. 5.7) and OS (5.9 vs. 4.5) in THNHM28523, a greater WNC (0.7-0.8 vs. 0.6) in THNHM28523 and THNHM28524 and higher number of TO (29/29 vs. 27/26)  and crista (crest). The name refers to a distinctive characteristic of the first discovered female specimen, which exhibited nuchal and dorsal crests with an orange colour. We suggest the following common names: kingkakhaownaam seesom (Thai), orange crested horned lizard (English), orange-verzierter geh\u00f6rnter Nackenstachler (German) and Acanthosaurus \u00e0 cr\u00eate orange (French).The specific epithet Acanthosauraaurantiacrista sp. n. occurs in the Thanon Thong Chai Mountain Range in northern Thailand: Mae Sariang District, Mae Hong Son Province  at 728 m a.s.l.; Sop Khong Subdistrict, Omkoi District, Chiang Mai Province  at 935 m a.s.l.; and Nang Lae Subdistrict, Mueang District, Chiang Rai Province  at 636 m a.s.l. This species usually lives in rainforests on mountains at elevations over 600 m a.s.l. . This herpetological discovery yields the sixth member of the genus Acanthosaura from Thailand and the taxa in this genus that have not yet been identified, will be described in the near future.The comparisons revealed morphological characters of a new long-horned lizard species, Thailand . In addiAcanthosauraaurantiacrista sp. n. is expected to distribute throughout the Thanon Thong Chai Mountain Range, which includes several important protected areas where the biological fauna is preserved in northern regions of Thailand, such as Doi Inthanon National Park; Doi Suthep-Pui National Park; Khun Khan National Park; Mae Ngao National Park; Mae Ping National Park; Mae Tho National Park; Mae Wang National Park; Namtok Mae Surin National Park; Op Luang National Park; Op Khan National Park; Chiang Dao Wildlife Sanctuary; Lum Nam Pai Wildlife Sanctuary; Mae Lao-Mae Sae Wildlife Sanctuary; Mae Tuen Wildlife Sanctuary; and Om Koi Wildlife Sanctuary. Moreover, our research team recommends this new long-horned species, whose nuchal and dorsal crests have an appearance similar to fire, as a representative animal for drawing the attention of the local people, scientific community and government towards addressing the current problem of forest burning in northern regions of Thailand.Many forest areas in northern regions of Thailand are currently faced with deforestation due to timber logging and forest fire for human use, which negatively influences the conservation of the herpetological fauna . However6CF57014-D7C4-5583-91CF-B24FDE50E47610.3897/BDJ.8.e48587.suppl1Supplementary material 1Acanthosaura and Acanthosauraaurantiacrista sp. n., \u201c?\u201d = data not available.Comparison of morphometric (in mm) and meristic data for all currently recognised species of Data typemorphologicalBrief descriptionAcanthosaura lizardThe comparison table of morphometric for File: oo_395107.docxhttps://binary.pensoft.net/file/395107Poramad Trivalairat"}
+{"text": "Callitris and Widdringtonia, the diterpene acids from Widdringtonia have never been described and no comparison to the Australian clade sister genus Callitris has been made. The critically endangered South African Clanwilliam cedar, Widdringtonia wallichii (syn. W. cedarbergensis), of the Cederberg Mountains was once prized for its enduring fragrant timbers and an essential oil that gives an aroma comparable to better known Mediterranean cedars, predominantly comprised by widdrol, cedrol, and thujopsene. In South Africa, two other \u2018cedars\u2019 are known, which are called W. nodiflora and W. schwarzii, but, until now, their chemical similarity to W. wallichii has not been investigated. Much like Widdringtonia, Callitris was once prized for its termite resistant timbers and an \u2018earthy\u2019 essential oil, but predominantly guaiol. The current study demonstrates that the essential oils were similar across all three species of Widdringtonia and two known non-volatile diterpene acids were identified in leaves: the pimaradiene sandaracopimaric acid (1) and the labdane Z-communic acid (2) with a lower yield of the E-isomer (3). Additionally, in the leaves of the three species, the structures of five new antimicrobial labdanes were assigned: 12-hydroxy-8R,17-epoxy-isocommunic acid (4), 8S-formyl-isocommunic acid (5), 8R,17-epoxy-isocommunic acid (6), 8R-17R-epoxy-E-communic acid (7), and 8R-17-epoxy-E-communic acid (8). Australian Callitris columellaris (syn. C. glaucophylla) also produced 1 and its isomer isopimaric acid, pisiferal (9), and pisiferic acid (10) from its leaves. Callitris endlicheri (Parl.) F.M.Bailey yielded isoozic acid (11) as the only major diterpene. Diterpenes 4\u20136, pisiferic acid (10), spathulenol, and guaiol (12) demonstrated antimicrobial and acaricidal activity.In spite of the evidence for antimicrobial and acaricidal effects in ethnobotanical reports of  Widdringtonia were once prized for their timbers and have been heavily exploited over the course of more than a century. The most pronounced impact has been on the now critically endangered W. wallichii Endl. ex Carri\u00e8re (=W. cedarbergensis J.A.Marsh) + ; 8S-formyl-isocommunic acid (5), HRESIMS m/z 341.2094 [M + Na]+ , 8R,17-epoxy-isocommunic acid (6), HRESIMS m/z 319.2258 [M + H]+ , 8R-17-epoxy-E-communic acid (7), HRESIMS m/z 341.2081 [M + Na]+ , 8R-17-epoxy-Z-communic acid (8), HRESIMS m/z 341.2081 [M + Na]+ .Five of the compounds are undescribed. These are structures 4\u20138, which were all isolated as amorphous translucent white solids: 12-hydroxy-8Staphylococcus epidermidis (ATCC 12228), Staphylococcus aureus (ATCC 29213 & a methicillin resistant strain), Pseudomonas aeruginosa (ATCC 27703), Bacillus subtilis (University of New England strain), and Escherichia coli (ATCC 25922). The positive control used was tetracycline.The minimum inhibitory concentration (MIC) method described by Eloff , which im/z 30\u2013400.Relative abundances of essential oil components and esterified diterpene acids were studied using gas chromatography with mass spectrometric detection (GC-MS). GC-MS analyses were performed using an Agilent Technologies 7890A GC-System coupled with an Agilent 5975C mass selective detector . An autosampler unit (Agilent Technologies 7693-100 positions) held samples. Separation of 1-\u03bcL injections used an HP-5MS Agilent column (30 m \u00d7 250 \u03bcm \u00d7 0.25 \u03bcm). Operating conditions were as follows: injector split ratio 25:1, temperature 250 \u00b0C, carrier gas helium, 1.0 mL/min, and constant flow. Column temperature was 50 \u00b0C (no hold) and 5 \u00b0C per minute. Then, at 280 \u00b0C, it was held at 5 min. Mass fragmentation patterns were acquired at \u221270 eV using a mass scan range of n-alkanes, when compared with values published in Adams [Primary identifications were performed by comparison of mass spectra with an electronic library database  and confin Adams  by visuain Adams . Semi-quHRESIMS spectra were recorded using an AB Sciex 5600 TripleTOF mass spectrometer in positive mode."}
+{"text": "Acinetobacter baumannii (XDRAB), isolated in Thailand. These results revealed multiple antimicrobial-resistant genes, each involving two sequence type 16 (ST16) isolates, ST2, and a novel sequence type isolate, ST1479.Here, we report the complete genome sequences of four clinical isolates of extensively drug-resistant  Acinetobacter baumannii (XDRAB), isolated in Thailand. These results revealed multiple antimicrobial-resistant genes, each involving two sequence type 16 (ST16) isolates, ST2, and a novel sequence type isolate, ST1479.Here, we report the complete genome sequences of four clinical isolates of extensively drug-resistant  Acinetobacter baumannii is one of the multidrug-resistant bacteria listed as a priority by the World Health Organization, as it exhibits resistance to most commercially available antibiotics and causes hospital-acquired infections (\u2013Acinetobacter baumannii (CRAB) infections present limited therapeutic options and are associated with high morbidity and mortality as well as longer hospitalization (A. baumannii (XDRAB) infections have been reported worldwide, including in Thailand (n = 3) and bile (n = 1) samples by a tertiary hospital in northeastern Thailand between 2017 and 2018 (gyrB multiplex PCR (n = 2) and no clonal group (n = 2) based on a multiplex PCR assay . Quality control of ONT reads was undertaken using NanoPlot v1.28.1 (https://github.com/wdecoster/NanoPlot). For the Illumina platform, the sequencing library was generated using the NEBNext Ultra II DNA library prep kit for Illumina  following the manufacturer\u2019s recommendations. We applied Fastp v0.19.5 (https://github.com/relipmoc/skewer). Quality checking of the Illumina reads was performed using FastQC v0.11.8 (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/). Hybrid assemblies with the ONT and Illumina data were performed using Unicycler v0.4.8 (https://github.com/rrwick/Unicycler/releases), and the genome sequences were checked for quality using QUAST v5.0.2 (http://quast.sourceforge.net/). Complete circular DNA structures of the bacterial chromosomes and plasmids were automatically produced using Unicycler software. The genome sequences were submitted to the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v4.12 for annotation. Default parameters were used for all software unless otherwise specified.Bacterial genomic DNA samples extracted using ZymoBIOMICS DNA kits  were sequenced using the Oxford Nanopore Technologies (ONT) and Illumina platforms. Library preparation for ONT sequencing followed the rapid barcoding DNA sequencing protocol with the SQK-RBK004 kit without DNA size selection, to preserve the plasmid DNA, and the libraries were sequenced using a single R9.4.1/FLO-MIN106 flow cell on a MinION Mk1B sequencer. We base called and demultiplexed the raw data using Guppy v3.4.5 (ONT), specifying the high-accuracy model (-c dna_r9.4.1_450bps_hac.cfg). The ONT adapters were trimmed using Porechop v0.2.4 ( v0.19.5  (https:/r v0.4.8  (https:/T v5.0.2  (http://https://cge.cbs.dtu.dk/services/ResFinder/) and CARD (https://card.mcmaster.ca/): blaNDM-1, blaOXA-23, blaOXA-66, blaTEM1D, blaVEB-1, blaIMP-14, blaADC-25, sul1, sul2, cmlA1, mphE, msrE, ARR2, tetB, tet(39), aac(3)-IId, aadA1, ant(2\u2033)-Ia, aph(3\u2032)-Ia, aph(3\u2033)-Ib, aph(3\u2032)-VI, aph(6)-Id, and armA. The multilocus sequence type (MLST) findings using the PubMLST database indicated that two isolates were ST16 (or ST355), and the other isolates were ST2 (or ST195) and ST1479 (new sequence type), according to the Pasteur MLST scheme (with the Oxford MLST scheme in parentheses) (https://pubmlst.org/abaumannii/).All isolates contained plasmids, and we detected seven plasmids in one isolate . From thand CARD  (https:/ntheses)  (https:/PRJNA647677. The accession numbers and assembly statistics are provided in These sequences have been submitted to GenBank under BioProject accession number"}
+{"text": "Doppler echocardiographic (Echo) measurements of the mitral and pulmonary venous flow have been used to assess diastolic dysfunction (DD). Velocity-encoded cardiac magnetic resonance (Ve-CMR) can measure similar parameters and may potentially be used for the assessment of DD.To assess the correlation of Ve-CMR with Echo in assessing mitral inflow E/A ratio, left atrial (LA) size, and pulmonary vein (PV) flow.We prospectively enrolled 22 outpatients with a mitral inflow pattern by Echo that was normal (8), impaired relaxation (7), pseudo-normal (2), or restrictive (5). The mean left ventricular ejection fraction (LVEF) was 60.8 and two patients had left ventricular hypertrophy on echo. Ve-CMR was performed within 2 hours of the Echo. The following indices were measured: LA size, mitral inflow E-velocity (E), A-velocity (A), E/A ratio, PV waves .The mean age of the patients was 51 years. The average Echo mitral inflow absolute velocities were close to double the Ve-CMR {Echo E-velocity = 85.8 cm/sec vs. Ve-CMR E velocity = 41.4 cm/sec, Echo A-velocity 70 cm/sec vs. Ve-CMR A-velocity 32.5 cm/sec}. The average E/A ratio was 1.41 by echocardiogram and 1.48 by Ve-CMR. There was a significant correlation in LA size and Mitral inflow E/A ratio between Ve-CMR and Echo . There were no significant correlations in PV waves . Figure 1Ve-CMR has a very high correlation with Echo in the assessment of mitral inflow E/A ratio and LA size. Ve-CMR maybe useful for the assessment of diastolic dysfunction."}
+{"text": "EGFR mutation-positive NSCLC at a major cancer center. This study showed the efficacy of 1G or 2G EGFR-TKIs as the 1L treatment, and subsequent therapy including 3G EGFR-TKIs in the real-world setting.The present study showed the comprehensive analysis of disease characteristics and treatment patterns in uncommon EGFR) mutations in non-small-cell lung cancer (NSCLC) are uncommon EGFR mutations. Although the efficacy of second (2G) or third generation (3G) EGFR tyrosine kinase inhibitors (EGFR-TKIs) in the patients with uncommon EGFR mutation has been proven, further studies are warranted to define the optimal treatment approach for uncommon EGFR mutation-positive NSCLC. This study retrospectively investigated the treatment patterns and outcomes of patients with uncommon EGFR mutation-positive NSCLC from January 2011 to December 2019 at the Samsung Medical Center, Seoul, Korea. During the study, 2121 patients with EGFR mutation-positive NSCLC received first-generation  or 2G EGFR-TKI (afatinib) as the first-line (1L) systemic therapy. Of this, 135 (6.4%) patients harbored uncommon EGFR mutations. Of 135, 54  patients had overlapping mutations with major EGFR mutations. The objective response rate (ORR) for the 1L EGFR-TKI was 63.3%. The median progression-free survivals (PFSs) were 8.6 months (95% CI: 3.8\u201313.5), 11.7 months (95% CI: 6.6\u201316.7), 7.7 months (95% CI: 4.9\u201317.4), and 5.0 months (95% CI: 3.7\u20136.1) for major uncommon EGFR mutation , compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), other compound mutation, and other uncommon mutations, respectively. The median overall survivals (OSs) were 25.6 months (16.9\u201334.2), 28.8 (95% CI: 24.4\u201333.4), 13.5 months (95% CI: 7.4\u201327.8), and 9.4 months (95% CI: 3.4\u201310.5) for major uncommon EGFR mutation (G719X), compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), other compound mutation, and other uncommon mutations, respectively. The response rate, median PFS, and OS were 63.3%, 16.3 months (95% CI: 15.6\u201316.9), and 37.5 months (95% CI: 35.4\u201339.6) for common EGFR mutation-positive NSCLC. After failing 1L EGFR-TKI, repeated tissue or liquid biopsy were carried out on 44.9% (35/78) of patients with T790M detected in 10/35 (28.6%) patients. With subsequent 3G EGFR-TKI after failing the first-line EGFR-TKI, the ORR and PFS for 3G EGFR-TKI were 80% and 8.9 months (95% CI: 8.0\u20139.8). These patients showed a median OS of 34.6 months (95% CI: 29.8\u201339.4). The ORR, PFS and OS were poorer in patients with uncommon  than those with common EGFR mutations. T790M was detected in 28.6% of the uncommon EGFR mutation-positive patients for whom prior 1G/2G EGFR-TKIs failed and underwent repeat biopsy at the time of progression.Approximately 10% of the epidermal growth factor receptor ( Uncommon EGFR mutations account for 7\u201323% of EGFR mutation-positive NSCLCs. Uncommon EGFR mutations can be categorized as follows: (i) de novo T790M; (ii) exon 20 insertions; (iii) \u201cmajor\u201d uncommon mutations Gly719Xaa (G719X), Leu861Gln (L861Q), and Ser768Ile (S768I); (iv) compound mutations; (v) other uncommon mutations [Exon 19 deletions (Del19) and epidermal growth factor receptor EGFR mutation-positive NSCLC has been revolutionized with the development of next-generation EGFR tyrosine kinase inhibitors (EGFR-TKIs). Currently, five EGFR-TKIs are available as the first-line (1L) therapy for advanced EGFR mutation-positive NSCLC\u2014first-generation (1G) reversible EGFR-TKIs, gefitinib, and erlotinib; second-generation (2G) irreversible ErbB family blockers, afatinib, and dacomitinib; third-generation (3G) irreversible EGFR-TKIs, osimertinib. Of the EGFR-TKI prospective randomized trials undertaken to date, only Iressa Pan-Asia Study (IPASS) [EGFR mutations. However, these studies included a small number of cases with uncommon EGFR mutations. Thus, it is unclear whether it is the best practice to use 2G or 3G EGFR-TKIs as the treatment of choice.The treatment of neration G irreverneration G irrever (IPASS) ,5 includEGFR-mutation positive NSCLC, recently, the highly anticipated findings of the phase III FLAURA trial showed 38.6 months of overall survival (OS) with frontline osimertinib, a 3G EGFR-TKI, versus 31.8 months with erlotinib or gefitinib [EGFR mutation-positive NSCLC who received sequential afatinib/osimertinib. The median OS was 41.3 months : 36.8\u201346.3) in the total population and 45.7 months (90% CI: 45.3\u201351.5) in patients with Del 19 [As to which treatment strategy is the best for the patients with major EGFR-TKIs and best sequential treatment, the bulk of these trials were limited to patients whose tumors harbored common EGFR mutations.While robust data from clinical trials have demonstrated the efficacy, tolerability, and benefits of EGFR-TKIs, followed by sequential treatment including 3G EGFR-TKI and cytotoxic chemotherapy in patients with uncommon EGFR mutation-positive NSCLC. In this study, we describe the real-world data of practice pattern and treatment outcomes in patients with uncommon EGFR mutation-positive NSCLC, including objective response rate (ORR), progression-free survival (PFS), and OS.To the best of our knowledge, there have been no prospective studies on the use of 1G or 2G EGFR mutation-positive NSCLC received gefitinib, erlotinib, or afatinib as 1L chemotherapy at the Samsung Medical Center. A total of 135 (6.4%) patients harbored uncommon EGFR mutations such as G719A/G719C/G719X, L861Q, S781I, H351I, E709K, and de novo T790M. The types of uncommon EGFR mutations are described in EGFR insertion 20 who received EGFR-TKIs as a 1L treatment. We classified patients with uncommon EGFR mutation into the major uncommon EGFR mutation , L861Q alone (n = 18), compound mutation with major EGFR mutation , other compound mutations (n = 13), other uncommon mutations (n = 2) and T790M (n = 15). There were 47.4% (64/135) male, and ex- or current smokers were 44.4% (60/135). About 25.9% of patients had symptomatic or asymptomatic brain metastasis at initial diagnosis.From January 2011 to December 2019, 2121 patients with EGFR-TKIs was 63.3% (76/120). The median PFS was 11.1 months (95% CI: 7.2\u201315.0) for uncommon EGFR mutation-positive NSCLC , compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), compound mutation , and other uncommon mutations , respectively. The median OS were 25.6 months (16.9\u201334.2), 28.8 (95% CI: 24.4\u201333.4), 13.5 months (95% CI: 7.4\u201327.8), and 9.4 months (95% CI: 3.4\u201310.5) for major uncommon EGFR mutation (G719X), compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), compound mutation, and other uncommon mutations, respectively. From October 2014 to December 2019, in the uncommon EGFR mutation-positive NSCLC group, the median PFS was 15.1 months (95% CI: 12.5\u201317.7) for afatinib and 7.7 months (95% CI: 1.3\u201314.1) for gefitinib or erlotinib (p = 0.165). The median OS was 25.6 months (95% CI: 18.2\u201333.0) for uncommon EGFR mutation-positive NSCLC  for afatinib and 15.5 months (95% CI: 9.0\u201322.0) for gefitinib or erlotinib (p = 0.03). There was no significant difference in sequential treatment after failing 1L treatment among the three EGFR-TKIs.The ORR  for the 1L 1G or 2G ve NSCLC . The medrlotinib A  patients (EGFR-TKIs (8/10 patients had overlapping EGFR mutations with Del 19 or EGFR exon 21 p.L858R), seven patients who did not have T790M received other EGFR-TKIs , 41 patients received cytotoxic chemotherapy, and 20 patients did not receive any sequential treatment.Among 120 patients (except de novo T790M), 78 experienced disease progression until January 2020. Among them, patients . T790M wEGFR-TKIs and cytotoxic chemotherapy, respectively (p = 0.05). In 58 patients who experienced disease progression and received sequential treatment, the median OS2 was 15.1 months (95% CI: 9.0\u201321.2) and 11.0 months (95% CI: 5.1\u201316.9) for 3G EGFR-TKIs and cytotoxic chemotherapy, respectively (p = 0.214). The median OS was 34.6 months (95% CI: 29.8\u201339.4), 24.4 months (95% CI: 17.4\u201331.4), and 5.3 months (95% CI: 2.9\u20137.7) for 3G EGFR-TKIs, cytotoxic chemotherapy, and no sequential treatment, respectively (p < 0.001).In 58 patients who experienced disease progression and received sequential treatment, the median PFS2 was 8.9 months (95% CI: 8.0\u20139.8) and 4.2 months (95% CI: 2.3\u20136.2) for third-generation ectively A (p = 0.ectively B (p = 0.EGFR-TKIs after failing the 1L EGFR-TKI treatment. The median PFS was 4.9 months (95% CI: 3.8\u20136.0), and the median OS was 24.0 months (95% CI: 0.7\u20134.8). Patients who received 3G EGFR-TKIs showed a median OS of 38.0 months (95% CI: 10.5\u201365.4).Among the 15 patients with de novo T790M mutation, 11 patients had an EGFR exon 21 p.L858R mutation and four patients had a Deletion 19 as a coexisting mutation. As a 1L treatment, seven patients received gefitinib, one patient received erlotinib, and seven patients received afatinib. Among them, eight patients received 3G EGFR mutation group, there were relatively more male patients and current/ex-smokers than in the common EGFR mutation group (EGFR mutation group was 66.6% (75% between October 2014 and December 2019), and in the uncommon EGFR mutation group it was relatively low at 44.9%. Although there was a difference between the two groups in the re-biopsy rate, the T790M detection rate in the patients who underwent re-biopsy was 63.5% in the common EGFR mutation group but was 28.6% in the uncommon EGFR mutation group. The response rate was 63.3% and 86.6% for the uncommon and common EGFR mutation groups, respectively. The median PFS was 16.3 months (95% CI: 15.6\u201316.9) and 11.1 months (95% CI: 7.2\u201315.0) for common EGFR mutation-positive NSCLC and uncommon EGFR mutation-positive NSCLC, respectively (p < 0.001).In the uncommon on group . During ectively  . In the patients who received 3G EGFR-TKIs after failing 1L EGFR-TKIs, the median OS was 44.4 months (95% CI: 38.9\u201349.9) and 34.6 months (95% CI: 29.8\u201339.4) for common EGFR mutation-positive NSCLC and uncommon EGFR mutation-positive NSCLC, respectively.In the overall population, the median OS was 37.5 months (95% CI: 35.4\u201339.6) and 25.6 months (95% CI: 18.2\u201333.0) for common ectively  , compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), afatinib showed superior OS results. In our study, less than one-third of the patients with uncommon EGFR mutation-positive NSCLC had a T790M mutation in the re-biopsy after failing 1L EGFR-TKIs. Most T790M mutation positive cases had overlapping mutations with major (Del 19 or EGFR exon 21 p.L858R) and uncommon EGFR mutations. The rate of T790M mutations after failing 1L EGFR TKI treatment in our study was only 28.6%, relatively lower than that of common EGFR mutation [EGFR mutation-positive NSCLCs than in common EGFR mutation-positive NSCLCs (27.1 % vs. 45.2%) [EGFR mutations after failing EGFR-TKI is lower than in patients with common EGFR mutations.The median ORR, PFS and OS of patients with uncommon ve NSCLC . The PFS G719X, L61Q, compmutation ,11. In a. 45.2%) . It needEGFR mutations are heterogeneous and have various sensitivities to EGFR-TKIs. In addition, the mechanism of acquired resistance to 1L EGFR-TKIs has not yet been well defined. Several retrospective studies and case reports of 1G EGFR-TKIs showed inconsistent responses in patients with uncommon EGFR mutation-positive NSCLC. A post-hoc analysis of prospectively collected data from the participants of the LUX-Lung 2, LUX-Lung 3, and LUX-Lung 6 trials showed the clinical activity of afatinib in patients with advanced uncommon EGFR mutation-positive NSCLC, especially G719X, L861Q, and S768I. However, it reported a low activity against T790M and exon 20 insertion mutations. Afatinib showed an ORR of 71% and a median PFS of 10.7 months (95% CI: 5.6\u201314.7), except for those with T790M or exon 20 insertion mutations, for whom ORR was 9\u201314%, and PFS was less than 3 months. The median OS was 19.4 months (95% CI: 16.4\u201326.9) [n = 315), it showed activity against major uncommon mutations , compound mutations , other uncommon mutations , and some exon 20 insertions  in EGFR-TKI na\u00efve patients [Uncommon .4\u201326.9) . In a reEGFR-TKI na\u00efve patients with uncommon EGFR mutation-positive NSCLC (KCSG-LU15-09), 22 of the 36 patients received osimertinib as the 1L treatment [EGFR-TKIs. Osimertinib conferred an ORR of 50%. The median PFS was 8.2 months (95% CI: 5.9\u201310.5), and the median OS was not reached. Furthermore, the response rate and median duration of response (11.2 months) were lower than those observed with osimertinib in patients with common mutations . In our study, the median PFS of 1G or 2G generation EGFR-TKI was 11.1 months (95% CI: 7.2\u201315.0), and the median OS was 25.6 months (95% CI: 18.2\u201333.0) in patients with uncommon EGFR mutant-positive NSCLC.In the phase II study of osimertinib for reatment . A totalEGFR mutations because of low sensitivity compared to the NGS [EGFR mutation detection accuracy [P53) in common EGFR mutation-positive NSCLC. However, in our study, few patients had an NGS result; therefore, further studies are needed to determine the prognostic effect of co-mutations in uncommon EGFR mutation-positive NSCLC.This study has several limitations due to its retrospective nature and single center experience. We had a small number of patients with brain metastasis at initial diagnosis. This study could not evaluate for CNS-specific outcomes or subgroup analysis. Among other things, only 44.9% of the patients were evaluated by repeated tissue or liquid biopsy at the time of progression because there was no accessible site for biopsy . Furthermore, the polymerase chain reaction-based or direct sequencing methods might have had limitations in detecting compound  the NGS ,16. Succaccuracy . It is aEGFR mutation-positive NSCLC, who started 1L gefitinib, erlotinib, or afatinib treatment for recurrent or metastatic NSCLC at the Samsung Medical Center between January 2011 and December 2019. This non-interventional observational study through big data analysis retrospectively collected de-identified patient data from a clinical data warehouse (CDW) using a unique algorithm with Standard Query Language (SQL) called the ROOT project. Patient demographic characteristics, such as age, sex, smoking history, performance status, and EGFR mutation type, were reviewed. We classified the uncommon EGFR mutation-positive NSCLC into the group with major uncommon EGFR mutations , compound mutations with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), other compound mutations, other uncommon mutations  and T790M. Demographic information was obtained when the 1L EGFR-TKI treatment was initiated. EGFR mutations were identified using a peptide nucleic acid (PNA)-clamp kit and real-time polymerase chain reaction, COBAS (cobas\u00aeEGFR Mutation Test v2 [TM).This study included patients with  Test v2 , or nextThis study was reviewed and approved by the Institutional Review Board (IRB) at the Samsung Medical Center (IRB No. 2018-05-130). The trial was conducted following the Declaration of Helsinki (as revised in 2013).EGFR-TKIs until progression or death resulting from any cause. PFS2 was defined as the first date of sequential treatment after failing 1L EGFR-TKIs until progression or death resulting from any cause. OS was defined as the first date of EGFR-TKIs until death resulting from any cause. OS2 was defined as the first date of sequential treatment after failing 1L EGFR-TKIs until death resulting from any cause. All p-values were two-sided, and p-value < 0.05 was considered statistically significant. The Korea Food and Drug Administration approved afatinib in October 2014. Among the three 1G or 2G EGFR-TKIs, PFS and OS were analyzed in patients who received EGFR-TKIs between October 2014 and December 2019.The all-data cut-off date for the analyses was February 2020. PFS and OS were calculated using a Kaplan\u2013Meier estimator and compared using the log-rank test. PFS and OS were presented as median values, with two-sided 95% CIs. PFS was defined as the first date of EGFR-mutant NSCLC showed lower ORR, PFS, and OS than the patients with common EGFR-mutant NSCLC. In particular, among the uncommon EGFR mutation, compared to the group with compounding mutation with major EGFR mutation and the group with major uncommon EGFR mutation, the group with other compounding mutation or other uncommon EGFR mutation showed inferior outcomes. Compared to common EGFR mutation-positive NSCLC, a lower percentage of patients underwent repeated biopsy and showed a lower detection rate of T790M. Less than one-third of the patients received 3G EGFR-TKI after failing the 1L EGFR-TKIs. However, like the common EGFR group, patients with T790M mutation after failing the 1L EGFR-TKI who received 3G TKI showed a favorable OS compared to other sequential treatments in the uncommon EGFR group. With the availability of more EGFR TKIs and a better understanding of tumor biology, further prospective studies are warranted to define the optimal treatment approach for uncommon EGFR mutant-positive NSCLC.The patients with uncommon"}
+{"text": "Older adults self-administer prescribed medication regimens to treat chronic diseases which can lead to mismanagement, medication related harm and hospitalizations. We examined the extent to which source of purchased medications influenced the occurrence of self-reported medication mistakes and hospitalizations in community-dwelling participants who managed medications independently (N= 3899). The majority (65%) picked-up medications, 18% had medications delivered, and 17% used both (picked-up and delivery). Compared to those picking up their medications, those using delivery only were less likely to have a hospital stay (OR=0.691 [95% CI 0.507-0.943]) and no difference in odds of medication mistakes (OR=1.051 [95% CI 0.764-1.445]), while those using both methods were more likely to report hospital stays (OR=1.429 [95% CI 1.106-1.846]) and medication mistakes (OR = 1.576[95% CI 1.078-2.304]). Older adults who picked-up medications from a local pharmacy and had medications delivered were more likely to report medication mistakes and hospitalizations."}
+{"text": "Scientific Reports 10.1038/s41598-019-39851-6, published online 27 February 2019Correction to: This Article contains errors in the legend of Figure 7.http://imaging.org.au/AMBMC/Model.\u201d\u201ca) Association matrix for calculating modules. The modules in (b) non-fasted and (c) fasted groups. Background image from should read:http://imaging.org.au/AMBMC/Model.\u201d\u201cAssociation matrix for calculating modules in (a) non-fasted and (b) fasted groups. The modules in (c) non-fasted and (d) fasted groups. Background image from"}
+{"text": "Mycobacterium abscessus strain Taiwan-45 that lytically infects M. abscessus strain BWH-C; phiT45-1 also infects M. abscessus ATCC 1997 but not Mycobacterium smegmatis. Phage phiT45-1 has a 43,407-bp genome and carries a polymorphic toxin-immunity cassette associated with type VII secretion systems.Mycobacteriophage phiT45-1 is a newly isolated bacteriophage spontaneously released from  Mycobacterium abscessus strain Taiwan-45 that lytically infects M. abscessus strain BWH-C; phiT45-1 also infects M. abscessus ATCC 19977 but not Mycobacterium smegmatis. Phage phiT45-1 has a 43,407-bp genome and carries a polymorphic toxin-immunity cassette associated with type VII secretion systems.Mycobacteriophage phiT45-1 is a newly isolated bacteriophage spontaneously released from  Mycobacterium abscessus is often antibiotic resistant and refractory to treatment. M. abscessus infections are frequent among cystic fibrosis patients and those with bronchiectasis and can disseminate in immunosuppressed patients  strains  are mycobacterial species that do not cause tuberculosis or leprosy . Among tpatients , 3. The  strains . The risernative .M. abscessus to contain prophages (M. abscessus Taiwan-45 onto a lawn of M. abscessus strain BWH-C (both provided by Chidiebere Akusobi and Eric Rubin) on solid medium at 37\u00b0C using standard methods (http://cobamide2.bio.pitt.edu)  . Putativitt.edu)  and HHpritt.edu) , 15. No itt.edu) . All tooM. smegmatis  (48 and 49) and several predicted HNH endonucleases  and immunity repressor (35) is consistent with phiT45-1 being temperate. Interestingly, phiT45-1 codes for a polymorphic toxin (PT) cassette, including an immunity protein (30), a polymorphic toxin (31) with RipA-like and WXG-100 domains (32)  to phages isolated on megmatis , althougviridae) . Early l domains , 21, andins (32) , situateMW570842 and BioProject accession no. PRJNA488469. The sequencing reads are available in the SRA under accession no. SRX10050651.Phage phiT45-1 is available at GenBank under accession no."}
+{"text": "T is an aerobic, non-motile and non-spore-forming Gram-positive coccus isolated from the stools of a Burkinabe woman. In this report, we present its phenotypic description including MALDI-TOF mass spectrometry analysis and genome sequencing. Strain Marseille-Q0835T; 2.9768-Mb genome exhibited a 41.9 mol% G+C content and 2699 predicted genes. Considering phenotypic features and comparative genome studies, we propose the strain Marseille-Q0835T as the type strain of Enterococcus burkinafasonensis sp. nov., a new species within the family Enterococcaceae.Strain Marseille-Q0835 Enterococcus species purposely named Enterococcus burkinafasonensis.Culturomics strategy is a high-throughput culturing method . This stBiotyper 3.0 software  and analysed against the main spectra of the bacteria included in the database.In September 2018, a fresh stool sample was collected from an apparently healthy 28-year-old Burkinabe woman who was admitted for diagnosis check-up in the Regional Tuberculosis Control Centre, Bobo-Dioulasso, Burkina Faso. A stool sample was sent to the collaborative laboratory at IHU in Marseille, France for culturomics analysis, which isolated an unidentified bacterial strain from the stool. The study was validated by the Science and Health Research Ethics Committee of Bobo-Dioulasso, under number (N/Ref.002-2018-CEIRS). The bacterium here referred to as strain Marseille-Q0835 was isolated on Columbia sheep blood agar after a 24-hour incubation under aerobic atmosphere at 37\u00b0C and pH 7.5. Purified colonies could not be identified by MALDI-TOF MS. The screening was performed on a Microflex LT spectrometer , as previously described [CodonCode Aligner software (http://www.codoncode.com). BLASTn research was conducted using nucleotide databases for cross-species comparison (https://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastn&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome). The search was limited to records that include sequences from type material, and exclude uncultured/environmental sample sequences. The result showed that strain Marseille-Q0835 exhibited a 97.80% sequence identity with Enterococcus gallinarum strain LMG 13129 (GenBank accession number NR_104559.2), the phylogenetically closest species with standing in nomenclature  and sequencing using the Big Dye\u00ae Terminator v1.1 Cycle Sequencing Kit and ABI Prism 3130xl Genetic Analyzer capillary sequencer , as previously described . The 16Snclature . We consEnterococcus sp. Marseille- Q0835T showed negative catalase and oxidase activities. API 50CH and API ZYM tests  were performed at 37\u00b0C under aerobic conditions and the results are summarized in Enterococcus sp. nov. strain Marseille-Q0835T with other bacterial species  with the EZ1 DNA tissue kit and then sequenced on MiSeq technology  with the Nextera XT Paired end (Illumina), as previously described . The assVelvet , Spades  [Spades  and Soapp Denovo ) on trimmmomatic ) or raw  removed . The besEnterococcus sp. Marseille-Q0835T with closely related species was estimated using the OrthoANI software version 0.93.1 (https://www.ezbiocloud.net/tools/orthoani) [Enterococcus malodoratus strain DSM 20681 and Enterococcus gallinarum strain LMG 13129 to 91.81% for Enterococcus devriesei strain DSM 22802 and Enterococcus viikkiensis strain LMG 26075. When the isolate was compared with these closely related species, values ranged from 70.07% with E.\u00a0gallinarum strain LMG 13129 to 73.41% with Enterococcus pseudoavium strain CBA7133. These values are lower than the 95% threshold used to discriminate bacterial species [.The degree of genomic similarity of rthoani) . Values  species .In silico DNA\u2013DNA hybridization values obtained using the GGDC version 2.0 online tool (http://ggdc.dsmz.de/ggdc.php) are reported in T, these values ranged from 20% with E.\u00a0devriesei strain DSM 22802 to 26.8% with Enterococcus faecium strain ISMMS VRE 1. Such values were lower than the 70% threshold recognized as delineating distinct species [. species ,17.TableT exhibited a 16S rRNA gene sequence divergence <98.65%, DNA\u2013DNA hybridization values\u00a0<\u00a070% and an OrthoANI value\u00a0<\u00a095% with its phylogenetically closest species with standing in nomenclature, together with unique phenotypic features. We formally propose strain Marseille-Q0835T as the type strain of the new species named Enterococcus burkinafasonensis.Strain Marseille-Q0835Enterococcaceae within the phylum Firmicutes. The type strain Marseille-Q0835T (CSUR P0835) was isolated after a 24-hour incubation at 37\u00b0C and pH 7.5 in an anaerobic atmosphere of a fresh stool sample collected from a 28-year-old Burkinabe woman. Colonies were smooth, white with entire edges with an average diameter of 1 mm. Bacterial cells were Gram-positive, coccus-shaped, non-motile and non-spore-forming with negative catalase and oxidase activities . The bacterium belongs to the family ties see .Fig.\u00a04HeT exhibits positive reaction for esterase lipase (C8), lipase (C14), naphthol-AS-BI-phosphohydrolase, \u03b1-galactosidase, \u03b2-galactosidase and N-acetyl-\u03b2-glucosaminidase, but negative reaction for alkaline phosphatase, esterase (C4), leucine arylamidase, valine arylamidase, cystine arylamidase, trypsin, \u03b1-chymotrypsin, acid phosphatase, \u03b2-glucuronidase, \u03b1-glucosidase, \u03b2-glucosidase, \u03b1-mannosidase and \u03b1-fucosidase. Using an API 50CH strip, positive reactions were obtained for d-galactose, d-glucose, d-fructose, l-arabinose, d-ribose, d-mannose, l-rhammose, d-mannitol, N-acetylglucosamine, arbutin, esculin, salicin, d-cellobiose, d-maltose, d-trehalose and gentibiose.Using an APIZYM strip, strain Marseille-Q0835T genome is 2.9768 Mb long, with a G-C content of 41.9%.The strain Marseille-Q0835The 16S rRNA gene and genome sequences were deposited in GenBank under accession number LR742708 and NZ_CACSLH000000000, respectively.T has been deposited in the French culture collection centre, Collection de Souches de l\u2019Unit\u00e9 des Rickettsies (CSUR), under the number Q0835.The strain Marseille-Q0835None to declare.10.13039/100007356IHU M\u00e9diterran\u00e9e Infection  and by the French Government under the Investissements d'avenir (Investments for the Future) programme managed by the 10.13039/501100001665Agence Nationale de la Recherche , (reference: M\u00e9diterran\u00e9e Infection 10-IAHU-03).This work was funded by the"}
+{"text": "E. coli. Terpene synthase activity was detected for 15 of these enzymes, and included mono-, sesqui- and diterpene synthase activities. A number of confirmed sesquiterpene synthases also exhibited promiscuous monoterpene synthase activity, suggesting that bacteria are potentially a richer source of monoterpene synthase activity then previously assumed. Several terpenoid products not previously detected in bacteria were identified, including aromandendrene, acora-3,7(14)-diene and longiborneol. Overall, we have identified promiscuous terpene synthases in bacteria and demonstrated that terpene synthases with substrate promiscuity are widely distributed in nature, forming a rich resource for engineering terpene biosynthetic pathways for biotechnology.Terpenes are the largest class of natural products with extensive structural diversity and are widely used as pharmaceuticals, herbicides, flavourings, fragrances, and biofuels. While they have mostly been isolated from plants and fungi, the availability and analysis of bacterial genome sequence data indicates that bacteria also possess many putative terpene synthase genes. In this study, we further explore this potential for terpene synthase activity in bacteria. Twenty two potential class I terpene synthase genes (TSs) were selected to represent the full sequence diversity of bacterial synthase candidates and recombinantly expressed in  The vast majority of terpenoids have been isolated from plants and fungi; however, bacteria are also known producers of volatile odoriferous metabolites. All terpenoids are synthesised from the universal C5 isoprenoid precursors isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), which are joined by isoprenyl transferases to form isoprenyl diphosphate substrates of varying lengths, such as geranyl diphosphate , farnesyl diphosphate  and geranylgeranyl diphosphate . Terpene synthases (TSs) convert the linear isoprenyl diphosphate substrates into structurally diverse mono- (C10), sesqui- (C15) and diterpene (C20) scaffolds. Due to their structural diversity, terpenoids have a wide range of industrial applications as pharmaceuticals, flavourings, fragrances, antimicrobials, pesticides and alternative fuels . Al. AlSoranous work . When Scous work . These rubstrate .Streptomyces clavuligerus are well studied [S. clavuligerus accepts GPP and FPP [Kitasatospora setae with GPP yielded linalool as a major product and small amounts of several acyclic and cyclic monoterpenes [Until now, only two bacterial monoterpene synthases linalool synthase (bLinS) and 1,8-cineole synthases (bCinS) from  studied  and stru studied . Broad s studied , a bLinS and FPP , and incterpenes .CFM47198.1, ELQ82238.1, ACY98649.1, EXG82599.1, ABU58787.1, EJL71407.1, KFE96946.1 were tested for compound production with FPP and GPP but no terpenoids were detected. in vivo product formation, all identified TSs were cloned into the pBbB2a-GPPS  backbone plasmid and co-expressed with pMVA plasmid in E. coli [in vivo conditions ,4,11-germacratriene as seen in the in vitro assays 11-diene and (+)-\u03b3-gurjunene when expressed in E. coli  from  E. coli . WhereasSM 43183  and EXG8SM 44712  producednditions .Streptomyces coelicolor A3(2) when expressed in E. coli produced various monoterpenes: \u03b2-myrcene, \u03b2-ocimene, linalool and geraniol  from E. coli with two mutations D2G, C155G (ispAM22), which can function as geranylgeranyl diphosphate synthase [in vivo. GGPP synthase activity using the variant prenyltransferase was confirmed by overexpression of spata-13, 17-diene synthase from Streptomyces xinghaiensis [ispAM22). This yielded the diterpene spata-13,17-diene, where the produced compound was extracted using a nonane layer  from Roseiflexus castenholzii DSM 13941  and Arctic Express (DE3) cells for recombinant protein expression.(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 TableKnown TSs annotated in the tree in (DOCX)Click here for additional data file.S1 FigNeighbour-joining tree constructed from the amino acid sequences of characterized terpene synthases in this study (underlined) and in previous studies. Geosmin synthases (dark red), 2-methylisoborneol synthases (orange), monoterpene (light blue), diterpene (dark blue) and sesquiterpene synthases (purple) are shown and uncharacterized bacterial terpene synthases in this study (in red).(DOCX)Click here for additional data file.S2 Fig-1) on a HP5 column and its retention time at 7.5 minutes. B. Mass spectra of geosmin.GC-MS traces showing the separation of geosmin (0.1 mg mL(DOCX)Click here for additional data file.S3 FigGC-MS chromatogram of geosmin synthase with 75 \u03bcM of FPP. Peak 1: geosmin (rt: 7.5), peak 2: Germacrene D (rt: 7.86), peak 3: germacradienol (rt: 8.84). B. Mass spectra of compounds observed in the extracts.(DOCX)Click here for additional data file.S4 Fig-1) produced in this study on a HP5 column. The internal standard, sec-butyl benzene , has a retention time of 9.5 minutes. Peak 1: R- Limonene (rt: 9.41). B: Mass spectra of limonene.A. GC-MS traces showing the separation of limonene (0.1 mg mL(DOCX)Click here for additional data file.S5 FigA. GC chromatogram of 10 \u03bcg of limonene synthase (pJBEI-6410) crude extracts with 75 \u03bcM of GPP. Peak 1: limonene (rt: 9.41). B. Mass spectra of limonene.(DOCX)Click here for additional data file.S6 FigA. GC-MS traces showing the separation of standard nerolidol mix (0.1 mg mL-1) on a HP5 column. B. MS spectra of -cis and -trans Nerolidol. C. GC-MS chromatogram of trans-nerolidol produced by AHY47823 with FPP. D. Mass spectra for trans-nerolidol produced by AHY47823.(DOCX)Click here for additional data file.S7 Fig-1) on a CP-Chirasil-DEX-CB column. B. GC-MS chromatogram of linalool isomers produced by AHY47823 with FPP. Samples were analyzed by GC on an Agilent Technologies 7890A GC system equipped with an FID detector, a 7693 auto sampler, and a CP-Chirasil-DEX-CB column . For linalool, the program initiated at a temperature of 70\u00b0C which was then increased to 90\u00b0C at 8\u00b0C/min. This was followed by an increase in temperature to 150\u00b0C at a rate of 2\u00b0C/min and then to 190\u00b0C at 40\u00b0C/min (1 min hold). The FID detector was maintained at a temperature of 200\u00b0C with a flow of hydrogen at 30 mL/min.A. GC-MS traces showing the separation of standard linalool mix (0.1 mg mL(DOCX)Click here for additional data file.S8 Fig-1) on a CP-Chirasil-DEX-CB column. B. GC-MS chromatogram of linalool isomers produced by AHY45426 with FPP. Samples were analyzed by gas chromatography on an Agilent Technologies 7890A GC system equipped with an FID detector, a 7693 auto sampler, and a CP-Chirasil-DEX-CB column . For linalool, the program initiated at a temperature of 70\u00b0C which was then increased to 90\u00b0C at 8\u00b0C/min. This was followed by an increase in temperature to 150\u00b0C at a rate of 2\u00b0C/min and then to 190\u00b0C at 40\u00b0C/min (1 min hold). The FID detector was maintained at a temperature of 200\u00b0C with a flow of hydrogen at 30 mL/min.A. GC-MS traces showing the separation of standard linalool mix (0.1 mg mL(DOCX)Click here for additional data file.S9 FigTotal ion chromatograms of products obtained from an incubation RrNerS (left) and RrBerS (right) with FPP for 3 hours at variable temperatures. Samples were analyzed by GC-QToF on HP5 column.(DOCX)Click here for additional data file.S10 FigA. \u03b1\u2013bergamotene produced by RrBerS, B. Aromandendrene produced by ScAroS, C and D. cadinene and longiborneol produced by BpLonS, E. germacrene D produced byKaGerS, F and G. copaene and acora-3,7(14)\u2013diene produced byAbAcoS, and H and I. \u03b2-elemene and 1(10),4,7(11)-germacra-triene produced by MiGerS.The reference spectra from the NIST library shown in blue and the compound spectra shown in red.(DOCX)Click here for additional data file.S11 FigA. GC-MS traces showing the separation of monoterpenoids (0.1 mg mL-1) standards used in this study on a HP5 column. The internal standard used, sec-butyl benzene , has a retention time of 9 minutes. Peak 1: cis-ocimene (rt: 9.56), peak 2: trans-ocimene (rt: 9.72), 3: linalool (rt: 10.43). B. Comparison of obtained mass spectra with NIST Library spectra and standard as shown in A. of products yielded by CAN96536 with GPP in in vitro assay.(DOCX)Click here for additional data file.S12 FigA. BpTPS; B. ScTPS1; C. CvTPS; D. SsTPS; E. ScTPS2; F. SniTPS; G.SnaTPS. The peaks in the chromatogram do not correspond to any terpenoids.GC-MS chromatograms of extracts profile with FPP are shown for (DOCX)Click here for additional data file.S13 FigA. GC-MS chromatogram for geosmin synthase products, B. mass spectra for Germacradienol, C\u2014G. Comparison of obtained mass spectra with NIST Library spectra of products yielded by GeoS.(DOCX)Click here for additional data file.S14 FigA\u2013B. SrGuaS C. TcCubS D. CaCubS.(DOCX)Click here for additional data file.S15 FigA. spata-13,17-diene produced by overexpression of ispAM22 and WP_095757924 in E. coli. B. Mass spectra of produced spata-13,17-diene.(DOCX)Click here for additional data file.S16 Fig(DOCX)Click here for additional data file.S17 Fig(DOCX)Click here for additional data file.S18 Fig(DOCX)Click here for additional data file."}
+{"text": "Vitamin D (VitD) deficiency during pregnancy has been associated with adverse neonatal outcomes and increased risk of late pregnancy complications. We planned to correlate serum VitD biomarkers; 25-hydroxyvitamin D (25-OH-VitD) and 1,25-dihydroxyvitamin D  levels; and their ratio with the frequency of feto-maternal pregnancy complications. A prospective cross-sectional case-control study was conducted at Aljouf Maternity and Children Hospital, Sakaka, Saudi Arabia, during the period of September 1, 2017 to September 30, 2019. 322 pregnant women were stratified into 2 groups: controls (110 cases) and complicated group (212 cases). The later comprised severe preeclamptic toxemia associated with intrauterine growth restriction (58 cases), gestational diabetes mellitus , abortion (26 cases), undisturbed ectopic pregnancy (16 cases), premature rupture of membranes , and, inevitable preterm labour (16 cases). After clinical assessment, peripheral blood samples were collected. Serum biomarkers were measured using specific immunoassays. The direct 1,25-diOH-VitD/25-OH-VitD ratio was calculated. Serum 25-OH-VitD indicated widely spreading VitD deficiency among participants with significantly higher levels in controls vs. GDM subgroup only. 1,25-diOH-VitD levels and the ratio were markedly reduced in the six complicated subgroups vs. controls, with non-significant differences amongst the complicated subgroups. ROC analysis showed very high sensitivity and specificity, to differentiate patients from controls, only for 1,25-diOH-VitD  followed by the ratio but not 25-OH-VitD. In conclusions, 25-OH-VitD did not show significant changes except for GDM. 1,25-diOH-VitD levels and the ratio showed strong associations with pregnancy complications. Serum 1,25-di-OH-VitD and its ratio to 25-OH-VitD are more reliable and physiologically relevant biomarkers for VitD status in pregnancy. Vitamin D (VitD) deficiency became a global epidemic, particularly for women of reproductive age. Nuclear receptor-mediated, the dihydoxy active form of VitD, 1,25-diOH-VitD, controls cellular proliferation, differentiation and apoptosis through targeting \u22651000 genes. Nevertheless, pathogenetic role of hypovitaminosis D in a number of comorbidities is still hotly debated Linked to hypovitaminosis D, adverse fetal outcomes included abortion, intrauterine growth restriction, fetal death and congenital malformations. Maternal adverse effects involved preeclampsia, gestational diabetes mellitus (GDM) and increased risk of preterm labor We planned to assess the relationship between hypovitaminosis D and feto-maternal pregnancy complications and morbidities, using the more patho-physiologically relevant biomarkers. We measured serum 25-OH-VitD, as the established VitD biomarker, and, 1,25-diOH-VitD as a functional VitD biomarker, and their ratio, and, correlated them with the existing pregnancy feto-maternal status.This prospective cross-sectional case control study was carried out at Aljouf Maternity and Children Hospital, Sakaka, Saudi Arabia, during the period between September 1, 2017 and September 30, 2019. It was approved by the local bioethical committees of Jouf University and Ministry of Health (#6-16-4/40). Written informed consents were obtained. The exclusion criteria were uncertain diagnosis, obesity, anemia, endocrinal disorders , immobilization for any reason, on anti-convulsion drug, acute infections, malabsorption syndromes, comorbid chronic medical disorder , multiple pregnancy, antepartum hemorrhage and superimposed preeclampsia.Among 450 pregnant women attending the hospital, 322 voluntary participants fulfilling the inclusion criteria were recruited. The 322 participants were classified into 2 groups; control group of 110 women who had normal feto-maternal pregnancy, and, complicated pregnancy group comprising 212 participants. The latter group was further subdivided into: severe preeclamptic toxemia (PET) associated with intrauterine growth restriction , GDM (82 cases), abortion (26 cases), undisturbed ectopic pregnancy (16 cases), premature rupture of membranes , and, inevitable preterm labor (16 cases).After history taking  and general examination , the head, face, neck, chest, heart, back, lower limbs and abdomen were systematically examined. The pregnancy was assessed ultra-sonographically for viability and fetal biometry to determine pregnancy duration and the expected fetal weight, condition of amniotic fluid and for diagnosing fetal anomalies, if present. Women with routine laboratory findings that contradict the inclusion criteria were excluded.oC. Specific quantitative ELISA immunoassay kits  were used to measure total 25-OH-VitD  and 1,25-DiOH-VitD . The direct 1,25-DiOH-VitD/25-OH-VitD ratio was calculated for each patient. Participants were stratified using the clinically relevant 25-OH-VitD cutoff levels of high/toxic as \u226550/>80 ng/mL, normal as \u226530-50 ng/mL, insufficient as \u226520 - <30 ng/mL, deficient as \u226510 - <20 ng/mL and severely deficient as <10 ng/mL Serum was recovered from peripheral blood samples by centrifugation and was aliquotted and stored till used at -80 Data was analyzed using SPSS . We expressed descriptive qualitative data as frequency and percentage, and, quantitative data as range and mean \u00b1 standard deviation (SD). Normal distribution was checked by Kolmogorov-Sminov test. One-way ANOVA with Tukey's post-test was employed for multiple comparisons. Correlation among variables was assessed by Spearman correlation coefficient test. Receiver Operating Characteristic (ROC) curve analysis was used to determine the area under curve (AUC) for 25-OH-VitD, 1,25-DiOH-VitD and their ratio to check their sensitivity and specificity to differentiate the cases from controls. P value of <0.05 at a confidence level of 95% was considered significant.Age was nonsignificantly different among all groups except for preeclamptics vs. each of controls (p = 0.011) and PROMs (p = 0.014), and, PROMs vs. GDM (p = 0.035). BMI showed nonsignificant difference among the investigated groups except comparing GDM vs. each of controls (p = 0.002), abortion (p = 0.02), ectopic pregnancy (p <0.001) and PROMs (p <0.001); and preeclamptics vs. each of ectopic pregnancy (p <0.028) and PROM (p = 0.045). Gravidity showed nonsignificant difference among the investigated groups except comparing GDM vs. each of controls (p <0.001), preeclamptics (p <0.001), abortion (p <0.036), ectopic pregnancy (p = 0.002) and PROMs (p = 0.002). Parity showed nonsignificant difference among the investigated groups except comparing GDM vs. each of preeclamptics (p <0.001), abortion (p = 0.035), ectopic pregnancy (p = 0.002) and PROMs (p <0.035). Duration of pregnancy showed very strong significant difference among the investigated groups with a few non-significant exceptions; it was significantly different comparing controls vs. all complication subgroups (p <0.01 - 0.001) except GDM, comparing preeclamptics vs. all other subgroups (p <0.05 - <0.001) except preterm labour, comparing GDM vs. the others (p <0.001), comparing abortion vs. the others (p <0.001) except ectopic pregnancy, and, comparing ectopic pregnancy vs. the others (p <0.001) except PROMs.Serum 25-OH-VitD levels showed nonsignificant difference among the investigated groups except comparing GDM vs. each of controls (p <0.001) and ectopic pregnancy (p = 0.01). Serum 1,25-diOH-VitD levels showed nonsignificant difference among the investigated groups of patients. However, controls had markedly higher levels vs. each of the six complicated groups (p <0.001). Similarly, the direct 1,25-diOH-VitD/25-OH-VitD ratio showed very marked significant reduction comparing controls vs. each of the six patients' groups (p <0.001), albeit, with nonsignificant difference among the complicated groups.Using serum 25-OH-VitD as the biomarker for VitD status, 43.636% of controls were deficient (\u226510 ng/mL), 38.181% were insufficient (\u226520 ng/mL), and, equal proportions (9.09%) were either normal (\u226530 ng/mL) or having toxic levels (>80 ng/mL). 82.759% of preeclamptic cases were insufficient, 13.793% were deficient and 3.448% had normal levels. 82.927% of GDM cases were deficient, 12.195% were insufficient and 4.878 had normal levels. 84.615% of abortion cases deficient, and equal proportions (7.692%) had either insufficient or toxic levels. 50.0% of ectopic pregnancy cases were deficient, 37.5% were insufficient and the rest (12.5%/2) had toxic levels. 71.429%/ of PROMs cases were deficient and 28.571% were insufficient. 75% of preterm labor cases were insufficient and 25.0% were deficient. Fortunately, severe deficiency (<10 ng/mL) was not observed in our cases. Therefore, VitD insufficiency/deficiency is widely spreading among the study participants . Controls presented a better picture only considering insufficiency/deficiency proportion, and, % of normal/super-normal levels.ROC curve analysis showed that 1,25-DiOH-VitD level is the most sensitive and most specific biomarker for differentiating between cases and controls; with an AUC of 0.965 . It is followed by 1,25-DiOH-VitD25-OH-VitD ratio; with AUC of 0.843 . 25-OH-VitD levels fell near the diagonal line with no effect, i.e., showing least sensitivity and least specificity  , parity (0.468/<0.001) and BMI (0.837/<0.001), and negative relationship with the ratio (-0.155/<0.05). Gravidity showed positive relationship vs. parity (0.958/<0.001) and BMI (0.480/<0.001). Parity correlated positively with BMI (0.448/<0.001). BMI correlated negatively vs. the ratio (-0.184/= 0.027). Pregnancy duration correlated negatively with 1,25-DiOH-VitD (-0.241/<0.006) and ratio (-0.192/= 0.022). 25-OH-VitD correlated positively vs. 1,25-DiOH-VitD (0.157/<0.05) and negatively vs. the ratio (-0.647/<0.001). 1,25-DiOH-VitD correlated positively with the ratio (0.562/<0.001).For preeclamptic cases, age had positive correlation vs. gravidity (0.682/<0.001), parity (0.608/<0.001) and BMI (0.690/<0.001), and, negative correlation vs. 1,25-DiOH-VitD (-0.267/= 0.021). Gravidity related had positively with parity (0.962/<0.001) and BMI (0.497/<0.001), but, negatively vs. 1,25-DiOG-VitD (-0.282/= 0.016). Parity correlated positively vs. BMI (0.424/<0.001), and, negatively with 1,25-DiOH-VitD (-0.218/<0.05). BMI correlated positively vs. duration of pregnancy (0.333/= 0.005) and negatively vs. 1,25-DiOH-VitD (-0.275/= 0.017). 25-OH-VitD correlated negatively with the ratio (-0.490/<0.001). 1,25-DiOH-VitD correlated positively vs. the ratio (0.853/<0.001).For GDM cases, age correlated positively vs. each of gravidity (0.680/<0.001), parity (0.732/<0.001), BMI (0.657/<0.001) and 25-OH-VitD (0.239/= 0.015), but, negatively vs. each of 1,25-DiOH-VitD (-0.179/<0.05) and the ratio (-0.320/<0.002). Gravidity correlated positively with parity (0.865/<0.001), BMI (0.671/<0.001) and 25-OH-VitD (0.216/<0.026), and, negatively vs. duration of pregnancy (-0.294/<0.004) and the ratio (-0.183/<0.05). Parity correlated positively vs. BMI (0.725/<0.001) and 25-OH-VitD (0.187/0.047), and, negatively vs. duration of pregnancy (-0.179/<0.05). BMI correlated positively vs. 25-OH-VitD (0.412/<0.001) and negatively vs. the ratio (-0.225/<0.021). 25-OH-VitD correlated negatively with the ratio (-0.757/<0.001). 1,25-DiOH-VitD had positive correlation vs. the ratio (0.592/<0.001).For abortion cases, age correlated positively with gravidity (0.406/<0.02), parity (0.535/= 0.0024), BMI (0.757/<0.001), and pregnancy duration (0.337/= 0.046), and, negatively vs. 1,25-DiOH-VitD (-0.669/<0.001) and the ratio (-0.380/<0.028). Gravidity correlated positively with parity (0.815/<0.001) and duration of pregnancy (0.565/= 0.0013), and, negatively vs. each of 1,25-DiOH-VitD (-0.333/= 0.048) and the ratio (-0.381/ = 0.027). Parity related positively with duration of pregnancy (0.358/= 0.036). BMI associated negatively with 1,25-DiOH-VitD (-0.385/= 0.026). Pregnancy duration correlated negatively vs. 1.25-DiOH-VitD (-0.371/<0.031) and the ratio (-0.490/<0.006). 25-OH-VitD correlated negatively vs. the ratio (-0.773/<0.001). 1,25-DiOH-VitD correlated positively vs. the ratio (0.665/<0.001).For the ectopic pregnancy cases, age correlated positively vs. gravidity (0.884/<0.001), parity (0.927/<0.001), and BMI (0.922/<0.001), but, negatively vs. 1,25-DiOH-VitD (-0.639/<0.005) and the ratio (-0.527/<0.02). Gravidity correlated positively vs. parity (0.906/<0.001) and BMI (0.854/<0.001), and, negatively vs. 1,25-DiOH-VitD (-0.565/<0.013). Parity correlated positively with BMI (0.933/<0.001), and, negatively vs. 1,25-DiOH-VitD (-0.624/<0.006) and the ratio (-0.437/<0.05). BMI correlated positively vs. duration of pregnancy (0.433/<0.05), and, negatively vs. 1,25-DiOH-VitD (-0.443/= 0.044). Pregnancy duration related negatively vs. 1,25-DiOH-VitD (-0.504/<0.027). 25-OH-VitD correlated negatively vs. the ratio (-0.690/= 0.002). 1,25-DiOH-VitD correlated positively with the ratio (0.647/= 0.004).Among PROMs cases, age correlated positively vs. gravidity (0.578/<0.018), parity (0.578/<0.018), BMI (0.727/= 0.002), 1,25-DiOH-VitD (0.500/<0.038) and the ratio (0.474/<0.05). Gravidity correlated positively vs. parity (1.0/<0.001), BMI (0.517/= 0.03) and duration of pregnancy (0.459/<0.05), but, negatively vs. 25-OH-VitD (-0.543/<0.026). Parity associated positively vs. BMI (0.517/= 0.03) and pregnancy duration (0.467/<0.05), but, negatively vs. 25-OH-VitD (-0.543/<0.026). BMI positively correlated with 1,25-DiOH-VitD (0.800/<0.001) and the ratio (0.655/= 0.007). 25-OH-VitD correlated positively with 1,25-DiOH-VitD (0.468/<0.048). 1,25-DiOH-VitD correlated positively vs. the ratio (0.786/<0.001).Within the preterm labor cases, age correlated positively vs. gravidity (0.898/<0.001), parity (0.908/<0.001), BMI (0.952/<0.001), and the ratio (0.452/<0.042), but, negatively vs. 25-OH-VitD (-0.439/<0.05). Gravidity correlated positively vs. parity (0.933/<0.001), BMI (0.802/<0.001) and the ratio (0.503/= 0.025). Parity correlated positively vs. BMI (0.835/<0.001) and the ratio (0.454/= 0.04), but, negatively with 25-OH-VitD (-0.454/= 0.04). BMI associated negatively with 25-OH-VitD (-0.500/<0.027), but, positively vs. the ratio (0.548/= 0.016). 1,25-DiOH-VitD correlated positively vs. the ratio (0.786/<0.001).st trimester of pregnancy, PROMs occurs after 37th weeks and before onset of labor and GDM is common at beginning of the last trimester), it showed significant differences comparing all groups Age was insignificantly different among our complicated groups. It had significant difference only when controls are compared to preeclamptics. Preeclampsia usually attacks extremely aged mothers; mainly primigravida. Gravidity showed a significant difference between the controls and GDM groups. Repeated pregnancy is a definite risk factor for GDM development. BMI was significantly different only comparing GDM and controls. GDM usually associates with higher BMI. As the gestational age is unique for the occurring complication  the traditionally low VitD foods, 2) a scalding local summer sun and ice-cold winter that give very narrow room for sun exposure, 3) low supplement intake, and, 4) absence of a national mandatory VitD food fortification program.The difficulties faced us carrying out this study included: 1) Absence of a reasonable number of other pregnancy complications such as congenital anomalies, stillbirth, isolated IUGR, 2) we could not correlate findings with comorbid rate of infection and inflammation, length of hospital stays, and long-term outcomes, & 3) Longitudinal analysis was not accessible.VitD deficiency/insufficiency is widely spreading in the studied population of pregnant women. Levels of 25-OH-VitD were not different among controls and complicated pregnancies except for a significantly lower level in GDM. 25-OH-VitD levels did not correlate with most of the characteristics of participants and pregnancy complications. 1,25-diOH-VitD and its ratio to 25-OH-VitD revealed very highly significant reduction in complicated pregnancy and were highly sensitive and specific in differentiating controls from complicated pregnancies. They also correlate with most of the characteristics of the participating women. 1,25-diOH-VitD and its ratio to 25-OH-VitD are potentially more functionally relevant biomarkers for VitD status particularly during pregnancy."}
+{"text": "Noroviruses  are a common cause of acute gastroenteritis in all age groups. These small non-envelope viruses with a single-stranded (+)RNA genome are characterized by high genetic variability. Continuous changes in the genetic diversity of co-circulating noroviruses and the emergence of new recombinant variants are observed worldwide. Recently, new recombinant noroviruses with a novel GII.P16 polymerase associated with different capsid proteins VP1 were reported. As a part of the surveillance study of sporadic cases of acute gastroenteritis in Novosibirsk, a total of 46 clinical samples from children with diarrhea were screened in 2016. Norovirus was detected in six samples from hospitalized children by RT-PCR. The identified noroviruses were classified as recombinant variants GII.P21/GII.3, GII. Pe/GII.4_Sydney_2012, and GII.P16/GII.4_Sydney_2012 by sequencing of the ORF1/ORF2 junction. In Novosibirsk, the first appearance of the new recombinant genotypeGII.P16/ GII.4_Sydney_2012 was recorded in spring 2016. Before this study, only four complete genome sequences of the Russian GII.P16/GII.3 norovirus strains were available in the GenBank database. In this work, the complete genome sequence of the Russian strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 (GenBank KY210980) was determined. A comparison of the nucleotide and the deduced amino acid sequences showed a high homology of the Russian strain with GII.P16/GII.4_Sydney_2012 strains from other parts of the world. A comparative analysis showed that several unique substitutions occurred in the GII.P16 polymerase, N-terminal p48 protein, and minor capsid protein VP2 genes, while no unique changes in the capsid VP1 gene were observed. A functional significance of these changes suggests that a wide distribution of the strains with the novel GII.P16 polymerase may be associated both with several amino acid substitutions in the polymerase active center and with the insertion of glutamic acid or glycine in an N-terminal p48 protein that blocks the secretory immunity of intestinal epithelial cells. Further monitoring of genotypes will allow determining the distribution of norovirus recombinants with the polymerase GII.P16 in Russia. Noroviruses  areconsidered to be one of the common causes of outbreaks andsporadic cases of acute gastroenteritis (AGE) in humans ofall ages . Norovirus infection can causesevere outcomes of the disease in very young and elderlyindividuals, as well as chronic diarrhea, lasting from severalmonths to several years, in immunocompromised and cancerpatients, and humans after organ transplantation . Dueto the low infectious dose  and highresistance in the environment, noroviruses are rapidly transmittedperson-to-person, by food and water . A meta-analysis of epidemiological datafrom many countries showed that the incidence of norovirusinfection among patients with AGE regardless of their age was17\u201320 % in 2008\u20132014 . The prevalenceof asymptomatic norovirus infection is estimated from 4 to18 % in different regions .The polyadenylated single-stranded (+)RNA genome ofnorovirus (~7.5 kb) contains three overlapping open readingframes (ORF1\u2013ORF3) . ORF1 encodes alarge polyprotein that is post-translationally cleaved by viralprotease into six nonstructural proteins, including RNAdependentRNA polymerase (RdRp); ORF2 and ORF3 encodemajor (VP1) and minor (VP2) capsid proteins, respectively.Two mechanisms of norovirus genetic variability have beenidentified: point mutations and recombination . Due to recombination events occurring in the norovirusgenome near the overlapping region of the 3\u02b9-end of ORF1(RdRp) and 5\u02b9-end of ORF2 (VP1), a dual nomenclature ofnoroviruses defining the RdRp/VP1 genotypes was recentlydeveloped .https://www.rivm.nl/mpf/typingtool/norovirus/). The average duration ofgenotype-specific immunity after norovirus infection can befrom 4 to 8 years , however, due to theexistence of a wide range of genetic variants, subsequent norovirusinfection with other antigenic variants or \u201cimmunotypes\u201dcan occur in a shorter time .Noroviruses exhibit significant genetic and antigenic diversity.Based on the VP1 amino acid sequence, noroviruses arecurrently classified into at least seven genogroups (GI\u2013GVII),which are further sub-divided into more than forty genotypes. It has been established that GI, GIIand GIV noroviruses can cause disease in humans . In the most common genogroup GII,at least 31 RdRp genotypes and 23 VP1 genotypes are distinguished,and their combination is designated as GII.Px/GII.x. In 2012, a newrecombinant GII.4 norovirus classified as GII.Pe/GII.4_Sydney_2012 appeared and later became the dominant strainworldwide . However, changes in themolecular epidemiology of norovirus have been observed inrecent years. In the winter season 2014/2015, a new GII.P17/GII.17 strain, which was first registered in China, quicklyreplaced the GII.Pe/GII.4_Sydney_2012 variant and initiallyspread to Asia, and later, to other regions . Recently, the prevalence of new recombinant norovirusstrains with the GII.P16 polymerase associated with multipleVP1 genotypes, including GII.4_Sydney_2012, has beenreported in different regions .In Novosibirsk, long-term monitoring of the genetic diversityof enteric viruses showed that noroviruses GII.P4/GII.4were a common cause of sporadic AGE cases in 2003\u20132012,while noroviruses with the GII.P16 polymerase were rarelydetected . In the spring of2016, we recorded the emergence of a new recombinant variantGII.P16/GII.4_Sydney_2012 in Novosibirsk. Before thisstudy, only four complete genome sequences of recombinantGII.P16/GII.3 noroviruses from Russia were available in theGenBank database . Theaim of this study was complete genome sequencing of the new Russian GII.P16/GII.4_Sydney_2012 strain and comparativeanalysis with similar strains from other regions and with Russian2005\u20132012 strains in which the GII.P16 polymerase wasin association with various other VP1 genotypes.Origin of virus strains. As a part of the surveillance studygenetic diversity of enteric viruses, clinical samples werecollected from children with diarrhea who were hospitalizedat Children\u2019s City Clinical Hospital No. 3 and were onoutpatient treatment in 2016. Written informed consent wasobtained from each parent/guardian of the child to participatein the study, in compliance with voluntariness in accordancewith the Federal Law \u201cOn the Principles of the Protection ofCitizens\u2019 Health in the Russian Federation\u201d. Detection anddifferentiation of viral RNA were performed by RT-PCR usinga verified laboratory primer panel, as previously described.Sequencing. The detected noroviruses were characterizedby sequencing of the genome region (~1400 nt), includingthe ORF1/ORF2 junction (20 nt). The nucleotide sequenceswere determined by the Sanger method using the BigDye\u2122Terminator v.3.1 Cycle Sequencing Kit and 3500 GeneticAnalyzer . The completegenome sequencing of the strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 was performed by the \u201cprimerwalking\u201dmethod using a panel of newly designed primers.The obtained data were analyzed by FinchTV . Partial fragments were assembled into a full-length genomesequence using SeqMan from the Lasergene EvolutionSuite software package .Norovirus genotype was determined using the Norovirus TypingTool v. 2.0  .Phylogenetic analysis. Reference norovirus sequenceswere obtained by search on BLAST 2.9.0+ (https://www.ncbi.nlm.nih.gov/). ClustalW alignment and phylogenetic analysisof nucleotide sequences were performed using MEGA 7(https://www.megasoftware.net/). Phylogenetic trees wereconstructed using the Neighbor-Joining method with theKimura 2-parameter model. The analysis was performed with abootstrap of 1000 replicas; only values >80 % were indicated.The identity of the nucleotide and amino acid sequences was calculated using BioEdit v7.2.6 software (http://www.mbio.ncsu.edu/bioedit/bioedit.html). The variability of the deducedamino acid sequences of the polyprotein comprising GII.P16RdRp, as well as the capsid proteins VP1 and VP2 of variantGII.4_Sydney_2012 was determined using the Sequence DataExplorer from MEGA 7.The nucleotide sequences obtained in this study were annotatedand deposited in the GenBank database with accessnumbers KY210919, KY210976\u2013KY210980, KY210983,MG892912 \u0438 MG892914.https://www.ncbi.nlm.nih.gov/) and RIVM (https://www.rivm.nl/mpf/typingtool/norovirus/) showed that the identified noroviruses belongedto three recombinant variants (Table 1).A total of 46 fecal samples from children aged 1 monthto 8 years were tested by RT-PCR. Enteric viruses weredetected in 15 (32.6 %) samples. Norovirus infection wasidentified in six hospitalized children aged 1 to 9 months.Analysis of nucleotide sequences (~1400 nt), including theORF1/ORF2 junction, by BLAST (The nucleotide sequences of the norovirus strains that weremost homologous to the sequences obtained in this studywere determined by search on BLAST. Isolate NS16-C32related to GII.P21/GII.3 genotype had high homology withstrains circulating in Novosibirsk (97.6\u201398.9 %) in 2010\u20132012 and in Europe (96.5\u201397.2 %) in2014\u20132016 . Two isolates NS16-C12and NS16-C13 genotyped as GII.Pe/GII.4_Sydney_2012had a high similarity (99.3\u201399.5 %) with GII.Pe/GII.4 strains(2014\u20132016) from Southeast Asia and Great Britain  and 96.9\u201397.1 % identity with strains that previouslycirculated in Novosibirsk .Nucleotide sequences of three remaining isolates NS16-C36,NS16-C37, and NS16-C38 related to the new genotype GII.P16/GII.4_Sydney_2012 had 100 % identity to each otherand 96.2 % homology with isolates NS16-C12 and NS16-C13. Genetic similarity of isolates NS16-C36, NS16-C37 andNS16-C38 with strains GII.P16/GII.4_Sydney_2012 fromother regions was 97.4\u201398.9 %.Based on complete norovirus sequences available in theGenBank database, several sets of original primers were designedfor complete sequencing of ORF1, RdRp, ORF2, and ORF3 of different genotypes. For two isolates, NS16-C13and NS16-C32, nucleotide sequences (~4300 nt), includingRdRp, ORF2 and ORF3, were identified and depositedin the GenBank database as strains Hu/GII.Pe-GII.4/RUS/Novosibirsk/NS16-C13/2016 (GenBank KY210977) andHu/GII.P21-GII.3/RUS/Novosibirsk/NS16-C23/2016 (Gen-Bank KY210919).For isolate NS16-C38, complete genome sequence(7560 nt) including the 3\u02b9-untranslated region (47 nt) wasdetermined and deposited in the GenBank database as thestrain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016(GenBank KY210980). Analysis of the deduced amino acidsequences showed that ORF1 (5100 nt) encoded a polyproteinof 1700 amino acid (aa) residues of length; ORF2 (1623 nt) andORF3 (807 nt) encoded the capsid proteins VP1 (541 aa) andVP2 (269 aa), respectively. The complete nucleotide sequenceof the Russian strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 had 98\u201399 % homology with recombinantGII.P16/GII.4_Sydney_2012 strains that appeared in theUSA and the UK in the winter season 2015/2016. Since thestrain studied was recombinant, a comparative analysis wasperformed separately for each ORF.Comparative analysis of the ORF1Phylogenetic analysis of complete ORF1 nucleotide sequenceswith GII.P16 RdRp available in GenBank showed that theanalyzed strains were divided into three clusters ; separate clades (supported on >85 %) within them wereformed by strains with the same VP1 genotype .Cluster III contained contemporary recombinant strains withGII.P16 RdRp associated with VP1 of four genotypes, GII.1,GII.2, GII.3, and GII.4_Sydney_2012. The ORF1 homology ofthe Russian strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 with other strains of cluster III was 97.9\u201399 %.Comparative analysis showed that the complete ORF1sequences of the GII.P16 RdRp strains varied from 5094 nt(reference strain AY772730_Hu/GII.P16-GII.16/DEU/Neustrelitz/2000) to 5100 nt (cluster III). When aligned, theGAA insert in the region encoding the N-terminal protein p48was found in recombinant GII.P16/GII.17 strains (cluster I)and in all strains from cluster II. Additional insertion of GAAor GGA was detected in contemporary recombinant strainsfrom cluster III in the same region of the p48 gene. For ORF1,1317 (25.8 %) variable sites were determined, of which906 (17.7 %) were informative, i. e. were in two or more strains.For polyprotein, 182 (10.7 %) variable sites were found,104 (6.1 %) of which were informative (Table 2). Comparativeanalysis showed that 14 variable sites were unique to thenovel GII.P16 RdRp lineage (cluster III), and the substitutionsin seven positions 52, 53, 644, 845, 853, 1546, and 1549resulted in a change in amino acid chemistry. An assessmentof the functional significance of the changes revealed thatthree non-synonymous substitutions  and77E/G insert were found in p48, which plays a role in virusentry through the host cell membrane . Four of the five non-synonymous substitutions  within GII.P16 RdRp occurredin the active center (see Table 2), and this may have affectedthe norovirus transmission.Comparative analysis of the ORF2In addition to the sequences determined in this study, phylogeneticanalysis of ORF2 included strains of different capsidgenotypes, which were identified in combination with theGII. P16 RdRp. On phylogenetic tree of the partial ORF2sequences,the analyzed strains formed separate clusters inaccordance with the capsid genotype and were further subdividedinto separate clades depending on the RdRp genotype.The GII.4 nucleotide sequences divided into two majorclades. The most polymorphic clade includes strains withthe GII.P4 RdRp, which previously circulated in Novosibirskand belonged to six GII.4 epidemic variants: Farmington_Hills_2002, Hunter_2004, Yerseke_2006a, Den_Haag_2006b,Apeldoorn_2007 and New-Orleans_2009 . The second clade was formed by GII.4_Sydney_2012 strains, which were further divided into two separateclusters, depending on the RdRp genotype \u2013 GII.Pe or GII. P16(supported on 99\u2013100 %).The ORF2 similarity of the Russian strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 to the GII.P16/GII.4_Sydney_2012 strains from other regions was 98.8\u201399 %.Comparison of complete ORF2 sequences of the GII. P16/GII.4_Sydney_2012 strains with the GII.Pe/GII.4_Sydney_2012 and GII.P4/GII.4_New_Orleans_2009 strains revealed236 (14.5 %) variable sites, of which 132 (8.1 %) were informative.In deduced amino acid sequences of VP1, eightvariable sites were unique to the GII.4_Sydney_2012 variant, which distinguishedthem from the GII.4_New_Orleans_2009 variant,and only two of these changes (at 368 and 373) were locatedin the hypervariable epitope A (Table 3). Notably, only onevariable site at position 540 was unique to the new lineage ofGII.P16/GII.4_Sydney_2012, however, it was not located inantigenic regions of the major capsid protein VP1.Comparative analysis of the ORF3Phylogenetic analysis of complete ORF3 sequences ofstrains with the GII.P16 RdRp showed that the analyzedsequences were divided into separate clusters depending onthe VP1 genotype . Within each VP1 genotype, strainswith different RdRp genotypes were grouped into separateclades. Comparative analysis of complete ORF3 sequencesof GII.P16/GII.4_Sydney_2012 strains with those of GII.Pe/GII.4_Sydney_2012 and GII.P4/GII.4_New_Orleans_2009strains revealed 109 (13.5 %) variable sites, and 55 (6.8 %)of them were informative.In the deduced amino acid sequences of the minor capsidprotein VP2 of GII.4_Sydney_2012 strains, eight uniquevariable sites at positions 81, 108, 148, 149, 158, 164, 205,and 241 were identified (Table 4), which differed them fromGII.4_New_Orleans_2009 strains. In addition, two other variablesites (at 155 and 157) were unique to the new lineage ofGII.P16/GII.4_Sydney_2012.This work is a part of long-term monitoring of the geneticdiversity of noroviruses associated with sporadic AGE cases inNovosibirsk, Russia. In March 2016, a new variant of GII. P16/GII.4_Sydney_2012 norovirus was first isolated in feces fromhospitalized children. In samples from hospitalized adults,this variant was first identified in autumn 2016 . GenBank searchshowed that in the European part of Russia, similar GII.4_Sydney_2012 noroviruses (GenBank MK033810\u2013MK033811)were found in samples of children from Nizhny Novgorod alsoat the end of 2016, unfortunately, the RdRp genotype was notdetermined for those isolates.Until recently, noroviruses with the GII.P16 RdRp wereconsidered uncommon, although local outbreaks associatedwith GII.P16/GII.2 (2009/2010 and 2012/2014) were reportedin Japan , and thosecaused by GII.P16/GII.13 (2009/2010) in Nepal . During the 10-year (2003\u20132012) monitoring ofnorovirus genotypes in Novosibirsk, Russia , GII.P16 RdRp was identified in five samples:GII.P16/GII.16 , GII.P16/GII.3  and GII.P16/GII.5(GenBank HM596590). Until 2016 in the Russian Federation,except Novosibirsk, GII.P16/GII.3 noroviruses were rarelydetected in Omsk  andSmolensk (GenBank KF895841), and GII.P16/GII.16 in Moscowand St. Petersburg . InNovosibirsk, recombinant noroviruses with the novel GII.P16RdRp, which differed from RdRp of variant 2010\u20132012 andwas in combination with multiple capsid genotypes , were often found in samplesfrom adult AGE patients since 2016 (data not published).Our results confirmed the hypothesis of the spread of newlyemerged recombinant norovirus strains with the novel GII. P16RdRp in different regions of the world .Before this study, only four complete genome sequencesof recombinant GII.P16/GII.3 norovirus strains from Russianwere available in the GenBank database . In this work, complete genome sequenceof the Russian strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 related to the newly emerged recombinantgenotype GII.P16/GII.4_Sydney_2012 was determined. Thecomparative analysis showed that unique changes occurred inthe amino acid sequences of two non-structural proteins \u2013 theN-terminal protein p48 and GII.P16 RdRp, as well as in the minor capsid protein VP2; at the same time, no significantchanges were detected in the major capsid protein VP1 ofGII.4_Sydney_2012.RNA-dependent RNA polymerase plays a crucial role inthe replication of the norovirus genome. Recent studies haveshown that the RdRp coding region is changing quickly;however, variable mutation rates were observed in differentRdRp genotypes . Our findings are consistentwith the hypothesis of Ruiz et al. (2017) that unusualworldwide distribution of the novel GII.P16 lineage is mainlydue to changes in the polymerase active center, which couldincrease the norovirus transmission. However, we assumethat changes in the N-terminal protein p48 have also playedsome role in the wide distribution of these new recombinantstrains. It was previously shown that p48 can bind the hostrestriction factors in an infected cell, allowing norovirus toavoid the host immune response, and its coding region hasa higher evolution rate than the complete norovirus genome. In addition, it is known that p48 is ableto block the local secretory immunity of intestinal epithelialcells, induce the disintegration of the Golgi apparatus anddisrupt the intracellular traffic of proteins . We assume that the insertion ofglutamic acid residue into the region, which already containsfour consecutive glutamic acid residues, increases the negativecharge at the N-terminus of p48, and this may affect boththe norovirus entry into the intestinal epithelial cells and theGolgi disintegration rate.The minor capsid protein VP2, playing an important role invirus replication  and viral particlestability , is also involved in modulation ofthe host immune response . The mutationrate of VP2 is higher than that for the major capsid protein VP1. Identified amino acid substitutionscould affect the ability of VP2 to suppress the presentationof antigens on cell membranes and the induction of humanprotective immunity.As the result of long-term monitoring of noroviruses RdRp/VP1 genotypes, the emergence of the novel GII.P16/GII.4_Sydney_2012 recombinant was recorded in Russia. Theanalysis showed that the distribution of the newly emergedrecombinant GII.P16/GII.4_Sydney_2012 is not associatedwith changes in the antigenic profile of the major capsid proteinVP1, which usually led to the emergence of new epidemicGII.4 variants. In GII.P16/GII.4_Sydney_2012 strains, a certainrole was probably played by changes in the minor proteinVP2 that might affect the antigenic composition of the viralparticle and help to avoid the cellular immune response. Inaddition, the multiple mutations in two non-structural proteins,the N-terminal protein of p48 and RdRp, probably increasedthe transmission of noroviruses with the novel GII.P16 RdRp.Further monitoring of genotypes will allow estimation of thespread of emerged recombinant noroviruses with the novelGII.P16 RdRp lineage in the Russian Federation and predictionof their epidemic potential.The authors declare no conflict of interest.Ahmed S.M., Hall A.J., Robinson A.E., Verhoef L., Premkumar P.,Parashar U.D., Koopmans M., Lopman B.A. Global prevalenceof norovirus in cases of gastroenteritis: a systematic review andmeta-analysis. Lancet Infect. Dis. 2014;14(8):725-730. DOI10.1016/S1473-3099(14)70767-4.Barreira D.M.P.G., Fumian T.M., Tonini M.A.L., Volpini L.P.B.,Santos R.P., Ribeiro A.L.C., Leite J.P.G., Souza M.T.B.M.,Brasil P., da Cunha D.C., Miagostovich M.P., Spano L.C. 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DOI 10.1007/s00705-013-1708-5.Lin Y., Fengling L., Lianzhu W., Yuxiu Z., Yanhua J. Functionof VP2 protein in the stability of the secondary structure ofvirus-like particles of genogroup II norovirus at different pHlevels: function of VP2 protein in the stability of NoV VLPs.J. Microbiol. 2014;52(11):970-975. DOI 10.1007/s12275-014-4323-6.Lun J.H., Hewitt J., Yan G.J.H., Enosi T.D., Rawlinson W.D.,White P.A. Recombinant GII.P16/GII.4 Sydney 2012 was thedominant norovirus identified in Australia and New Zealand in2017. Viruses. 2018;10(10):548. DOI 10.3390/v10100548.Mallory M.L., Lindesmith L.C., Graham R.L., Baric R.S. GII.4 humannorovirus: surveying the antigenic landscape. Viruses.2019;11(2):177. DOI 10.3390/v11020177.Motomura K., Boonchan M., Noda M., Tanaka T., Takeda N.Norovirus epidemics caused by new GII.2 chimera viruses in2012\u20132014 in Japan. Infect. Genet. Evol. 2016;42:49-52. 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DOI 10.1098/rsos.170602.van Beek J., Ambert-Balay K., Botteldoorn N., Eden J.S., FonagerJ., Hewitt J., Iritani N., Kroneman A., Vennema H., Vinje J.,White P.A., Koopmans M., on behalf of NoroNet. Indicationsfor worldwide increased norovirus activity associated with emergenceof a new variant of genotype II.4, late 2012. Eurosurveillance.2013;18(1):pii=20345. Available online: https://www.eurosurveillance.org/content/10.2807/ese.18.01.20345-en.Vinje J. Advances in laboratory methods for detection and typingof norovirus. J. Clin. Microbiol. 2015;53(2):373-381. DOI10.1128/JCM.01535-14.Vongpunsawad S., Venkataram Prasad B.V., Estes M.K. Norwalkvirus minor capsid protein VP2 associates within the VP1shell domain. J. Virol. 2013;87(9):4818-4825. DOI 10.1128/JVI.03508-12.Woodward J., Gkrania-Klotsas E., Kumararatne D. Chronic norovirusinfection and common variable immunodeficiency. Clin.Exp. Immunol. 2017;188(3):363-370. 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+{"text": "Cnidium monnieri (L.) Cuss (C. monnieri) is one of the most widely used traditional herbal medicines, exhibiting a wide range of pharmacological functions for treating asynodia, trichomonas vaginitis, and osphyalgia. Its important medicinal value comes from its abundance of coumarins. To identify genes involved in coumarin biosynthesis and accumulation, we analyzed transcriptome data from flower, leaf, root and stem tissues of C. monnieri. A total of 173,938 unigenes with a mean length of 1,272 bp, GC content of 38.79%, and N50 length of 2,121 bp were assembled using the Trinity program. Of these, 119,177 unigenes were annotated in public databases. We identified differentially expressed genes (DEGs) based on expression profile analysis. These DEGs exhibited higher expression levels in flower tissue than in leaf, stem or root tissues. We identified and analyzed numerous genes encoding enzymes involved in coumarin biosynthesis, and verified genes encoding key enzymes using quantitative real-time PCR. Our transcriptome data will make great contributions to research on C. monnieri and provide clues for identifying candidate genes involved in coumarin metabolic pathways. Cnidium monnieri (L.) Cuss, an annual plant of the Umbelliferae family native to China and Vietnam, exhibits a diverse set of pharmacological activities including anti-osteoporotic  has been widely applied as a powerful method for discovering and predicting functional genes, and revealing gene expression patterns, especially in non-model species for which reference genome sequences are insufficient . To dateus albus , Artemisia argyi , Arisaemophyllum , Clinopochinense  and Polyyrtonema . HoweverC.\u00a0monnieri using BGI-500 sequencing technology. Numerous potential genes and differentially expressed genes (DEGs) involved in coumarin biosynthesis were screened using de novo transcriptome sequencing. Our findings will increase our understanding of the molecular mechanisms of coumarin biosynthesis in C.\u00a0monnieri and provide clues for exploring functional genes in other plants having close relationships with C.\u00a0monnieri.The purpose of this study was to uncover functional genes associated with or playing regulatory roles in coumarin biosynthesis. We conducted transcriptome sequencing and gene expression profiling in C.\u00a0monnieri plants were collected from Tongcheng city in Anhui Province, China in May 2018, with verbal permission of the Manager Zhongquan Fang , and authenticated by Professor Dequn Wang (Anhui University of Chinese Medicine). Prior to the experiment, the plants were grown at a temperature of 22\u221226\u00a0\u00b0C/14\u221218\u00a0\u00b0C (day/night) and relative humidity of 65\u201380%. All samples were rinsed in ultrapure water, and leaves, roots, flowers and stems, which were harvested from three individual C.\u00a0monnieri plants, were placed in centrifuge tubes, frozen in liquid nitrogen immediately and stored at \u221280\u00a0\u00b0C.Whole Total RNA was extracted from each tissue using RNA plant Plus Reagent  according to the manufacturer\u2019s instructions. The extracted RNA was checked using a NanoDrop 2000 , and the concentration of the isolated RNA, 28S/18S and integrity (RIN) were estimated using an RNA Nano 6000 Assay Kit and the Agilent Bioanalyzer 2100 system  .C.\u00a0monnieri samples from flowers, roots, leaves and stems were used for isolation of osthole as previously reported  to remove DNA residues and then mixed with magnetic beads containing oligo (dT) to purify mRNA. After purification, the mRNA was fragmented into small pieces under elevated temperature. The cleaved RNA fragments were copied into first-strand cDNA using reverse transcriptase and random primers. Second-strand cDNA was synthesized using DNA Polymerase I and RNase H. These cDNA fragments had the addition of a single adenine at the 3\u2032\u00a0end for subsequent ligation of adapters. The products were then purified and enriched using PCR amplification. The quality of each sample library was evaluated using an Agilent 2100 Bioanalyzer system . Each cDNA library was sequenced using the BGISEQ-500 system at Beijing Genomics Institute (BGI)  with paired-end (PE) sequencing length of 100 bp.i\u00a0\u2212\u00a0A 0.25). In the absence of a reference genome, clean reads were subjected to transcriptome assembly using Trinity software (version 2.5.1) applying the following parameters: \u2013min_contig_length 150 \u2013CPU 8 \u2013min_kmer_cov 3 \u2013min_glue 3 \u2013bfly_opts \u2019-V 5 \u2013edge-thr =0.1 \u2013stderr\u2019  , NCBI non-redundant protein sequences (NR) (update date: 23 June 2019) , clusters of euKaryotic Orthologous Groups (KOG) (http://www.ncbi.nlm.nih.gov/KOG), Kyoto Encyclopedia of Genes and Genomes (KEGG) (update date: 1 July 2018) (http://www.genome.jp/kegg/) (http://www.uniprot.org/). Assembled unigenes were annotated against NR database using the software BLAST . And the results of NR annotation were used for Gene Ontology (GO) (update date: 1 January 2019) annotation using Blast2GO (version v2.5.0) program (Assembled unigenes were annotated against the NCBI nucleotide sequences (NT) (p/kegg/) , and a m program . Pfam  using Power SYBR\u00ae Green PCR Master Mix . The actin gene (CL3748.Contig7) of C.\u00a0monnieri was used as a reference. Primers for all selected genes were designed using Premier 6.0 and are listed in \u00ae Green Master Mix. Reactions were conducted under the following conditions: denaturation at 95\u00a0\u00b0C for 1 min, followed by 40 cycles of 95\u00a0\u00b0C for 15 s and 63\u00a0\u00b0C for 25 s. Each qRT-PCR was performed using three biological replicates. The 2Ct\u2212\u0394\u0394 method was used to calculate the relative expression level of genes . qRT-PCR analysis of each gene was performed on a QuantStudioof genes .After assembly, clean reads were mapped to unigenes using Bowtie2 (version 2.2.5) , and theThe open reading frames (ORFs) of unigenes were initially detected using getorf (version EMBOSS:6.5.7.0)  and furtP value <0.05 was considered to be statistically significant.The contents of osthole and all data from qRT-PCR were presented as mean\u00a0\u00b1\u00a0standard deviation. Statistics were done by GraphPad Prism 6.0. Heatmap and boxplot were performed using R software (version 3.4.4). The one-way analysis of variance (ANOVA) was used to compare the data between groups with Duncan t test. C.\u00a0monnieri. Osthole content was highest in flowers (1.372%) and lowest in stems (0.283%)  . OstholeC.\u00a0monnieri using BGISEQ-500 sequencing technology. After eliminating low-quality reads, we obtained a total of 10.6 Gb, 10.47 Gb, 10.81 Gb and 10.38 Gb of clean reads from 83.42 Mb, 81.65 Mb, 84.27 Mb and 82.14 Mb sets of raw data from leaf, stem, flower and root tissue, respectively, with all Q30 values greater than 85%  against seven public databases , and a total of 110,992 (63.81%), 95,603 (54.96%), 80,216 (46.12%), 86,568 (49.77%), 87,523 (50.32%), 79,335 (45.61%) and 56,429 (32.44%) unigenes were aligned, respectively (Daucus carota subsp. sativus (82.2%) and others (17.8%)  (e <10ectively . We annoectively . We focuectively . Additio (17.8%) .We determined expression values of transcripts with fragments per kilobase of transcript per million mapped reads (FPKM) >1 for each tissue and found 55,421, 52,787, 48,287 and 44,717 expressed unigenes in flower, leaf, stem and root tissues, respectively . The oveC. monnieri, 86,568 (49.77%) unigenes were annotated and assigned to 135 KEGG pathways (19 subcategories) , cinnamate 4-hydroxylase , 4-coumarate\u2013CoA ligase , shikimate hydroxycinnamoyl transferase , 5-O-(4-coumaroyl)-D-quinate 3\u2032-monooxygenase , caffeoyl-CoA O-methyltransferase  and feruloyl-CoA ortho-hydroxylase   . Fourteeegories) . Phenylpnigenes) . These dnigenes) .C. monnieri (C4H), CL16061.Contig1 (4CL) and CL8379.Contig6 (F6H1) genes was highest in root tissue, whereas relative expression of the CL11351.Contig3 (PAL) and CL2358.Contig9 (BGA) genes was highest in flower tissue. Unigene 20867 (BGA) had a high expression level in leaf tissue. The expression levels determined by qRT-PCR were consistent with the FPKM values.We selected six unigenes encoding five key enzymes and used quantitative real-time PCR (qRT-PCR) to determine their relative expression levels in the leaf, stem, flower and root tissues of monnieri . RelativWe identified 12,635 unigenes uniquely expressed in flowers and 77,762 shared unigenes exhibiting expression in all four tissues . Total DAll DEGs were annotated in the KEGG database to further describe and evaluate their biological functions. This analysis revealed 32,060 DEGs in flower versus leaf associated with 137 pathways; these DEGs were mainly enriched in the \u201cplant hormone signal transduction,\u201d \u201cplant-pathogen interaction,\u201d \u201ccircadian rhythm - plant\u201d and \u201cphotosynthesis - antenna proteins\u201d pathways . In flowA total of 18,881 up-regulated DEGs showed flower-specific expression with |log2 (fold changes) | >2. We inferred the nature of these DEGs via GOSlim functional analysis. Sequence homology revealed that each of the 18,881 flower-specific DEGs mapped to at least one ontology term, including 2,814 for biological processes, 3,232 for cellular components and 4,508 for molecular function; many genes were enriched in \u201ccellular process\u201d and \u201ccatalytic activity\u201d subcategories within the biological processes and molecular function categories, respectively .To evaluate the biological functions of these DEGs, the 18,881 flower-specific DEGs were annotated in the KEGG database. KEGG enrichment analysis showed that these DEGs were significantly enriched in the \u201cribosome,\u201d \u201cstarch and sucrose metabolism,\u201d \u201cpentose and glucuronate interconversions\u201d and \u201cphenylpropanoid biosynthesis\u201d categories . AdditioC. monnieri transcriptome database, including 1,169, 1,032 and 920 unigenes up-regulated in flowers compared with leaves, stems and roots, respectively (Transcription factors (TFs) play an important role in regulating secondary metabolites by modulating the expression of genes related to biosynthetic pathways. We identified 3,799 unigenes encoding putative TFs in the ectively . The majectively . Of thesC. monnieri. Their structures and physicochemical properties have been well studied  genes were mapped in at least one public database, while 54,761 (31.48%) genes remained unannotated because of the lack of available genomic information.Coumarins are the most important bioactive components of  studied . HoweverPAL), CL2358.Contig9 (BGA), Unigene 20867 (BGA), CL3766.Contig1 (C4H), CL16061.Contig1 (4CL) and CL8379.Contig6 (F6H1) examined using qRT-PCR were consistent with the FPKM values determined by RNA-Seq. Coumarins are a major group of natural plant products derived from the phenylpropanoid pathway  in Angelica gigas , 4CL (2 DEGs), HCT (13 DEGs), COMT (7 DEGs) and F6\u2032H1 (8 DEGs) were related to coumarin biosynthesis . These kArabidopsis results in increased biosynthesis and accumulation of scopoletin, which belongs to the coumarins  Standard curve of osthole at 322 nm. (B) Osthole content from flowers, leaves, roots and stems of Click here for additional data file.10.7717/peerj.10157/supp-2Figure S2Click here for additional data file.10.7717/peerj.10157/supp-3Figure S3Click here for additional data file.10.7717/peerj.10157/supp-4Figure S4Click here for additional data file.10.7717/peerj.10157/supp-5Figure S5Click here for additional data file.10.7717/peerj.10157/supp-6Figure S6Click here for additional data file.10.7717/peerj.10157/supp-7Figure S7Red boxes represent enzymes annotated in the KEGG database. Photo credit: Kanehisa Laboratories.Click here for additional data file.10.7717/peerj.10157/supp-8Figure S8actin gene (CL3748.Contig7) as a reference gene for normalization. FPKM values of these genes are shown as red lines, qRT-PCR results of these genes are shown as blue bars.Relative expression of (A) CL11351.Contig3 (PAL), (B) CL2358.Contig9 (BGA), (C) Unigene 20867 (BGA), (D) CL3766.Contig1 (C4H), (E) CL16061.Contig1 (4CL) and (F) CL8379.Contig6 (F6H1) was analyzed by qRT-PCR using the Click here for additional data file.10.7717/peerj.10157/supp-9Figure S9Click here for additional data file.10.7717/peerj.10157/supp-10Supplemental Information 10Click here for additional data file.10.7717/peerj.10157/supp-11Supplemental Information 11Click here for additional data file.10.7717/peerj.10157/supp-12Supplemental Information 12Click here for additional data file.10.7717/peerj.10157/supp-13Supplemental Information 13Click here for additional data file.10.7717/peerj.10157/supp-14Supplemental Information 14Click here for additional data file.10.7717/peerj.10157/supp-15Supplemental Information 15Click here for additional data file.10.7717/peerj.10157/supp-16Supplemental Information 16Click here for additional data file.10.7717/peerj.10157/supp-17Supplemental Information 17Click here for additional data file.10.7717/peerj.10157/supp-18Supplemental Information S1Click here for additional data file."}
+{"text": "This study examined the diagnostic accuracy of 18F-DCFPyL (PSMA) PET/CT for lymph-node staging in primary PCa.The detection of lymph-node metastases (N1) with conventional imaging such as magnetic resonance imaging (MRI) and computed tomography (CT) is inadequate for primarily diagnosed prostate cancer (PCa). Prostate-specific membrane antigen (PSMA) PET/CT is successfully introduced for the staging of  recurrent PCa. Besides the frequently used 18F-DCFPyL PET/CT prior to robot-assisted radical prostatectomy (RARP) with extended pelvic lymph-node dissection (ePLND). Patients were included between October 2017 and January 2020. A Memorial Sloan Kettering Cancer Centre (MSKCC) nomogram risk probability of \u2265\u20098% of lymph-node metastases was set to perform ePLND. All images were reviewed by two experienced nuclear physicians, and were compared with post-operative histopathologic results.This was a prospective, multicentre cohort study. Patients with primary PCa underwent 18F-DCFPyL PET/CT detection of pelvic lymph-node metastases on a patient level were 41.2% : 19.4\u201366.5%), 94.0% (CI 86.9\u201397.5%), 53.8% (CI 26.1\u201379.6%) and 90.4% (CI 82.6\u201395.0%), respectively.A total of 117 patients was analysed. Lymph-node metastases (N1) were histologically diagnosed in 17/117 patients (14.5%). The sensitivity, specificity, positive predictive value and negative predictive value for the 18F-DCFPyL PET/CT showed a high specificity (94.4%), yet a limited sensitivity (41.2%) for the detection of pelvic lymph-node metastases in primary PCa. This implies that current PSMA PET/CT imaging cannot replace diagnostic ePLND. Further research is necessary to define the exact place of PSMA PET/CT imaging in the primary staging of PCa. Prostate cancer (PCa) is the most frequently diagnosed cancer in men in the Western world , 2. Init68gallium-labelled PSMA tracers. High detection rates for metastases were demonstrated even at low prostate-specific antigen (PSA) values (i.e. 45% for PSA <\u20090.5\u00a0ng/mL and over 95% for PSA \u2265\u20092.0\u00a0ng/mL) fluoro-pyridine-3-carbonyl)-amino]-pentyl)-ureido)-pentanedioic acid)  112\u2013123\u00a0min) of a median dose of 311\u00a0MBq 18F-DCFPyL (IQR 297\u2013324\u00a0MBq) within a median of 4.1\u00a0weeks (IQR 2.1\u20136.6) after prostate biopsy and within a median of 5.9\u00a0weeks (IQR 3.6\u201312.0) prior to surgery. Image acquisitions were performed using a Philips Ingenuity TF  and a Siemens Biograph-16 TruePoint  PET/CT system. The scan trajectory included mid-thigh to skull base, with 4\u00a0min (Philips) and 5\u00a0min (Siemens) per bed position. All PET scans were combined with a low-dose CT (33/117 patients) or contrast-enhanced CT scan (84/117 patients) . Images were corrected for decay, scatter, random coincidences and photon attenuation.Patients were staged with Images were reconstructed with a BLOB-based Ordered-Subsets Expectations Maximization algorithm   and the All scans were clinically and prospectively interpreted in the participating centres by one of two nuclear medicine physicians  with ample experience in PSMA PET interpretation (>\u2009300 scans). Upon completion of the study, all scans were reviewed by a second independent reader who was blinded to initial scan interpretation, surgery and histopathology results . A joint re-evaluation was performed in case of incongruent scan interpretation (consensus read), and used for final analysis. Lymph-node metastases were defined as increased PSMA expression, higher than the background, incompatible with physiological uptake, and in a typical site of PCa. A significant CT substrate was not an absolute prerequisite. The following parameters were recorded: detection of the primary tumour, tumour stage and presence of pelvic lymph-node metastases (N1). Pelvic lymph-node metastases were further classified in accordance with the four sections of the ePLND .To assess inter-observer variability, the proportional agreement was calculated, based on the two individual scan interpretations . Proportpeak), with correction for background uptake [peak is defined as the highest local intensity of uptake with a 6-mm-radius sphere [peak of the prostate tumour for patients with and without lymph-node metastases, the Mann-Whitney U test was used (significance set at p\u2009<\u20090.05). To compare the median SUVpeak of the prostate tumour with the corresponding Gleason score of the lesion, the Kruskall-Wallis test was used (significance set at p\u2009<\u20090.05). A linear regression was run to predict PSA from SUVpeak of the prostate index lesion (significance set at p\u2009<\u20090.05).PET/CT scans with PSMA-avid lesions in the prostate were delineated according to the reports of the nuclear medicine physicians. Semi-automatic delineation on the PET scans from both participating centres was performed with the in-house developed ACCURATE tool\u00a9 . The ACCd uptake . SUVpeaks sphere . To compThe ePLND surgical template includes removal of fatty lymphoid tissue overlying the common and external iliac vessels and within the obturator fossa . The medProstate specimens and resected lymph-node templates were fixated in formaldehyde (10%) directly after surgery and processed according to routine clinical standards . Individ18F-DCFPyL PET/CT to detect pelvic lymph-node metastases. The diagnostic accuracy of 18F-DCFPyL PET/CT was calculated with histopathologic evaluation of ePLND as a reference. The sensitivity, specificity and positive and negative predictive values  of 18F-DCFPyL PET/CT for the detection of pelvic lymph-node metastases (pN1) were calculated both on a patient level and on a surgical template level. The surgical template analysis was based on the abovementioned 4 surgical templates of the ePLND, and was applied to approximate lesion based-detection using 18F-DCFPyL PET/CT. To compare the median diameter of PET/CT-detected lymph-node metastases vs. PET/CT-undetected lymph-node metastases, the Mann-Whitney U test was used (significance set at p\u2009<\u20090.05).The primary outcome of this study was the patient-based sensitivity of 18F-DCFPyL PET/CT to differentiate local advancement  from prostate-confined disease (T2). This study did not investigate the exact location of local PCa advancement, only the presence of extracapsular or seminal vesicle invasion was noted. Local advancement was defined as PSMA expression outside the borders of the prostate gland, not suspect for overprojection or bladder/urethral physiological activity. Numerical variables were summarized with median values and interquartile ranges (IQR), categorical variables with proportions (%). Statistical analysis was done with IBM\u00ae SPSS\u00ae Statistics for Windows\u00ae, version 26.For the assessment for the local tumour stage (pT), we measured the accuracy of 18F-DCFPyL PET/CT, as presented in Fig.\u00a0A total of 120 patients were included in this study, and scheduled for ePLND with RALP after 18F-DCFPyL PET/CT suspicious for lymph-node metastases. Hence, the patient-based sensitivity to detect lymph-node metastases using 18F-DCFPyL PET/CT was 41.2%  19.4\u201366.5), with a specificity of 94.0% (95%CI 86.9\u201397.5), a PPV of 53.8% (95%CI 26.1\u201379.6) and a NPV of 90.4% (95%CI 82.6\u201395.0), as shown in Table Pathological features after RARP and ePLND are listed in Table 18F-DCFPyL PET/CT preoperatively identified 38 PSMA-avid regions suspect for lymph-node metastases in 18 surgical ePLND templates. Therefore, the template-based sensitivity for the detection of lymph-node metastases using 18F-DCFPyL PET/CT was 34.7% (95%CI 17.1\u201357.1), with a specificity of 97.7% (95%CI 95.7\u201398.9), a PPV of 44.4 (95%CI 22.4\u201368.6) and a NPV of 96.6% (95%CI 94.4\u201398.0) as seen in Table 18F-DCFPyL PET/CT, please see Fig.\u00a0In the 17 patients with lymph-node metastases, 31 lymph-node metastases were histologically identified in 23 surgical ePLND templates. n\u2009=\u200912) had a median tumour size of 5.5\u00a0mm (IQR 2.4\u20136.6), whereas the PET/CT-undetected lymph-node metastases (n\u2009=\u200919) had a significantly smaller median tumour size of 1.5\u00a0mm (IQR 1.0\u20134.5) (p\u2009=\u20090.03). A clinical example of a patient with both missed and detected lymph-node metastases is shown in Fig.\u00a0The median diameter of resected lymph-node metastases was 2.5\u00a0mm (IQR 1.0\u20136.0). The PET/CT-detected lymph-node metastases  p\u2009=\u20090.0. A clinimm IQR 2.\u20136.6, whe18F-DCFPyL PET/CT to detect locally advanced tumour growth (pT3-4) were 45.2% (95%CI 32.7\u201358.2%), 94.4% (95%CI 83.7\u201398.6%), 90.3% (95%CI 73.1\u201397.5%) and 60.0% (95%CI 48.8\u201370.7%), respectively, as seen in Table 18F-DCFPyL PET/CT were 18.1% (95%CI 8.7\u201333.2), 97.2% (95%CI 89.4\u201399.5), 80.0% (95%CI 44.2\u201396.5) and 66.0% (95%CI 56.1\u201374.8), respectively. For the detection of pT3b sub-stage, the sensitivity, specificity, PPV and NPV of 18F-DCFPyL PET/CT were 52.9% (95%CI 28.5\u201376.1), 89.9% (95%CI 81.8\u201394.8), 47.4% (95%CI 25.2\u201370.5) and 91.8% (95%CI 83.9\u201396.1), respectively. A clinical example of seminal vesicle (pT3b) PCa involvement on PET/CT is presented in Fig.\u00a0A total of 116/117 patients (99.1%) showed PSMA expression in the prostate at PET/CT. The sensitivity, specificity, PPV and NPV of owth pT3- were 45.peak of the PSMA-avid prostate lesions was found between patients with and without lymph-node metastases  (p\u2009=\u20090.46). A Gleason score of 4\u2009+\u20095\u2009=\u20099 or higher was associated with a higher median SUVpeak when compared with the lower Gleason scores . A Gleason score of 4\u2009+\u20094\u2009=\u20099 or higher was not associated with a higher median SUVpeak when compared with the lower scores . No correlation was found between SUVpeak of the PSMA-avid prostate lesions and PSA (R2\u2009=\u20090.12).No significant difference in the median SUVProportional agreement for the detection of lymph-node metastases using PET/CT was present in 94.7% (95%CI 89.8\u201397.7) overall, with a positive agreement of 78.6% (95%CI 53.4\u201393.9) and a negative agreement of 97.0% (95%CI 92.4\u201399.2). Proportional agreement for locally advanced tumours was observed in 79.1% (95%CI 71.7\u201385.3), with a positive agreement of 55.6% (95%CI 38.2\u201372.0), and a negative agreement of 86.2% (95%CI 77.9\u201391.5).18F-DCFPyL PET/CT for primary staging of PCa, assessing the diagnostic accuracy for the detection of pelvic lymph-node metastases. The results of a total of 117 patients with intermediate and high-risk PCa that underwent 18F-DCFPyL PET/CT and ePLND were analysed.PSMA PET/CT imaging is currently the imaging technique of choice for patients with biochemically recurrent disease after initial curative local treatment (EAU guidelines) . Its val18F-DCFPyL PET/CT imaging demonstrated a limited sensitivity for pelvic lymph-node metastases of 41.2%, at 94.0% specificity. The limited sensitivity indicates that 18F-DCFPyL PET/CT does not detect all lymph-node metastases. Therefore, albeit invasive, ePLND remains the gold standard for nodal staging. The 90% NPV might suggest that a negative test is reliable in the majority of times, though we should mind the low prevalence of lymph-node metastases in this cohort (14.5%). This prevalence was congruent with the median MSKCC risk for lymph-node metastases of 14.4% (IQR 10.1\u201330.2). Although specificity for lymph-node metastases was favourable in this study, the PPV was moderate at 53.8% . In this clinical setting, not all positive 18F-DFCPyL PET/CT results for pelvic lymph-node metastases represent actual metastatic disease. Good intra-observer agreement for the detection of pelvic lymph-node metastases using 18F-DFCPyL PET/CT was observed (95%).In this study, 68Ga-PSMA study from van Kalmthout et al. [n\u2009=\u2009103 patients), applying the same standardized ePLND techniques, histopathology analyses and PET positivity criteria. It revealed a patient-based sensitivity for lymph-node metastases of 41.5% (95%CI 26.7\u201357.8) and a specificity of 90.9% (95%CI 79.3\u201396.6) [18F-DCFPyL PET/CT is at least comparable with 68Ga-PSMA imaging.Our results appear to be in line with a recent prospective t et al. . This st.3\u201396.6) . Althoug68Ga PSMA ligands, which reported a specificity of 90% and higher [18F-DCFPyL PET/CT as an imaging tool for initial staging of PCa [The high specificity presented in the current study confirms results from previous retrospective studies with d higher , 29, 30.g of PCa . In 25 pg of PCa . PotentiU test: p\u2009=\u20090.03). This may explain the imperfect imaging sensitivity reported in this study. The 5-mm spatial resolution offered by PET is still an improvement compared with the detection limits of CT and MRI (i.e. >\u200910\u00a0mm [68Ga-PSMA PET/CT vs. MRI (65% vs. 41%) [PET/CT-detected lymph-node metastases were larger than lymph-node metastases that were not detected by PET/CT . This d >\u200910\u00a0mm . This st18F-DCFPyL PET/CT should still be followed by curative treatment in the form of a RALP with an ePLND. However, the threshold to perform ePLND with certain amounts of detected number of lymph-node metastases remains unclear.The therapeutic consequence of PSMA-detected pelvic lymph-node metastases remains a matter of debate. Previous research showed that patients with lymph-node metastases detected intra-operatively (with frozen sections) or preoperatively (with CT) still benefit from radical prostatectomy and complete lymph-node dissection \u201334. This18F-DCFPyL PET/CT, yet the sensitivity was limited at 45.2% (95%CI 32.7\u201358.2). Moreover, the promising specificity of the detection of pT3a-b using 18F-DCFPyL PET/CT of 94.4% (95%CI 83.7\u201398.6) is in line with previous reports on 68Ga-PSMA (specificity >\u200990% for T3b) [18F-DCFPyL PET/CT is comparable with that of mpMRI, for which a meta-analysis revealed a specificity of 88% (95%CI 85\u201397%) compared with our 94.4% (95%CI 83.7\u201398.6) [A total of 116/117 patients (99.1%) showed PSMA expression in the prostate at PET/CT. A promising PPV for the detection of pT3a-b of 90.3% (95%CI 73.1\u201397.5) was observed using for T3b) , 35, 36..7\u201398.6) . The sen.7\u201398.6) . Altoget18F-DCFPyL PET/CT for detecting distant metastases. Only patients undergoing RARP and ePLND were considered for analysis, which naturally excludes patients with distant metastases in which radical surgery is forgone. As such, our results should be interpreted as the accuracy of 18F-DCFPyL PET/CT for N-staging in patients (expected to be) free from distant metastases . Therefore, we decided to focus our study on determining the accuracy of 18F-DCFPyL PET/CT for N-staging, as hereto a solid reference standard is available (ePLND). Determining the accuracy of M-staging is certainly of interest, yet any such analysis is limited to providing a PPV, as the true prevalence of distant metastases cannot be known.Our study has inherent limitations. Firstly, this study did not assess the accuracy of 18F-DCFPyL PET images were obtained, again detecting positive lymph nodes in the surgical template. Metastasis-directed radiotherapy to these lesions was followed by a PSA-response. The ePLND may possibly have missed these lesions initially that were rightfully detected on the first PET/CT scan.Since the PET/CT resolution is confined at 5\u00a0mm, limited diagnostic accuracy for micro metastases is to be expected. Secondly, this study might not have been adequately powered, since the expected prevalence was higher than the actual prevalence (30% vs. 14.5%). Moreover, the sensitivity used for the power analysis was higher than actually realized (90% vs. 41.2%) due to the high expectations for the sensitivity of PSMA PET/CT. Lastly, we should consider that the golden standard (ePLND) is not always flawless: in two patients with PET-positive lymph nodes, the ePLND was reported to be technically challenging. Histopathological analysis did not reveal any lymph-node metastases, yet these patients soon developed a biochemical recurrence. Repeated 18F-DCFPyL PET/CT scan could be used to withhold PCa patients from an ePLND. Future studies are therefore needed to assess whether the diagnostic accuracy of 18F-DCFPyL PET/CT, its high specificity in particular, could assist in proper treatment planning of patients with intermediate and high-risk stages of disease.The follow-up data of this cohort is necessary to investigate whether a specific risk profile in combination with a negative 18F-DCFPyL PET/CT imaging for the detection of lymph-node metastatic disease in men with intermediate and high-risk prostate cancer, undergoing radical surgery. We found a limited sensitivity of 41.2% (95%CI 19.4\u201366.5) at excellent specificity (94.0%). Based on current results, 18F-DCFPyL PET/CT imaging should not replace ePLND.In this prospective cohort study, we evaluated the accuracy of"}
+{"text": "Ulmus parvifolia cvs. Allee and Drake in Lake County, Florida. Nematode species were identified using both molecular analysis and morphology of perineal patterns. Meloidogyne enterolobii and M. javanica were identified from U. parvifolia cv. Allee. Meloidogyne arenaria and M. javanica were identified from U. parvifolia cv. Drake. This is a first report of these nematode species infecting Chinese Elm in Florida.Samples of galled roots, resembling those induced by root-knot nematodes, and rhizosphere soil were collected from potted plants of  Ulmus parvifolia Jacq.: Ulmaceae: Rosales) are valued for their tough lumber, resistance to some elm pests and pathogens  were collected directly from the rhizosphere of U. parvifolia. These samples were designated with internal FDACS-DPI sample identifiers N20-110, N20-113 (both from U. parvifolia cv. Allee), and N20-115 (from U. parvifolia cv. Drake). Round galls, resembling those commonly induced by M.\u00a0enterolobii, were observed on secondary and tertiary roots of U. parvifolia cv. Allee in one of the samples  and used for qPCR, conventional PCR, and sequencing. The cytochrome c oxidase subunit I (COI) and intergenic spacer 2 (IGS2) qPCR assays were repeated with DNA from J2 extracted directly from the roots of each cultivar. Standard PCRs targeted NADH-ubiquinone oxidoreductase chain 5 (NADH5) and cytochrome c oxidase subunit II (COXII) using the primers NAD5F2/NADH5R1 and COX2F/COX2R, respectively, and thermocycle conditions described by R assays , isozymeR assays . DNA wasMeloidogyne isolates with both NADH5 and COXII sequences were further analyzed. The alignments of NADH5 (448\u2009bp) and COXII (323\u2009bp) were trimmed until data were 100% complete for each terminal taxon. Alignments were then concatenated  and analyzed simultaneously. K2P . Newly generated sequences  were aligned in MEGA7  using thsly. K2P  distancesly. K2P .Meloidogyne spp. found infecting U. parvifolia is provided (n = 6) and were positively identified as M. enterolobii from both qPCR assays. All other samples had undetermined Ct values for the qPCR assays. COXII sequences from N20-110 samples were 100% BLASTn matches to previously published M. enterolobii data. With the concatenated matrix, isolates N20-115-1A, N20-115-6A, N20-115-10A, N20-115-16B, N20-113-1B, N20-113-14B, N20-113-18B were 100% matches to M. javanica  Chitwood, 1949  Chitwood, 1949 (M.\u00a0sp. n. 1 (n = 26 for each sample) and morphology of perineal patterns were consistent with those reported for M. enterolobii  isolated singly from U.\u00a0parvifolia cv. Allee (N20-110) and M. javanica  also found singly on this same cultivar (N20-113), and both M.\u00a0arenaria  and M. javanica  identified as mixed species infecting U. parvifolia cv. Drake (N20-115). To our knowledge this is the first report of Ulmus parvifolia as a host of M. arenaria, M.\u00a0enterolobii, and M. javanica in Florida.A summary of identified provided . Samples"}
+{"text": "Escherichia coli (STEC) O26:H11 strain, MBT-5 , and two draft genome sequences of Listeria monocytogenes strains MBT-6 and MBT-7 belonging to the virulent sequence types 1  and 59 , respectively, were determined. The strains were isolated in 2015 from ready-to-eat mixed greens in Germany.The complete genome sequence of a Shiga toxin-producing  Escherichia coli (STEC) O26:H11 strain, MBT-5 , and two draft genome sequences of Listeria monocytogenes strains MBT-6 and MBT-7 belonging to the virulent sequence types 1  and 59 , respectively, were determined. The strains were isolated in 2015 from ready-to-eat mixed greens in Germany.The complete genome sequence of a Shiga toxin-producing  Escherichia coli (STEC) serotype O26 can lead to life-threatening infections and is the second most common serotype (after O157) found in clinical and food samples in Europe  for E. coli. The AX Bacteria+ kit is suitable for getting high-molecular-weight DNA. DNA quantification and paired-end  and long-read  sequencing were performed as previously described (https://github.com/s-andrews/FastQC). Hybrid assembly of short and long sequence reads of STEC strain MBT-5, obtained from Illumina and MinION sequencing, respectively, was performed with Unicycler v0.4.8 (L. monocytogenes strains MBT-6 and MBT-7 was done with SPAdes v3.10.0 (Cultures were grown overnight at 37\u00b0C in brain heart infusion (BHI) broth, and genomic DNA was extracted using the ZR fungal/bacterial DNA miniprep kit  for escribed . For sho v3.10.0 .dnaA as the origin with Geneious v9.2. Plasmids were not rotated. Strain MBT-5 was identified as serotype O26:H11, sequence type 21 (ST21), and adhesin fimH type 440 using SerotypeFinder v2.0, MLST v2.0, and FimTyper v1.0, respectively (\u2013ehxA (enterohemolysin), espA (extracellular serine protease), toxB (toxin B), and katP , while all other virulence genes were located on the chromosome.The hybrid assembly of STEC strain MBT-5 generated a single circular chromosome of 5,705,580\u2009bp, with 6 circular plasmids . The circular chromosome of MBT-5 was rotated to ectively \u201312. Viructively \u2013 13). In. IndnaA L. monocytogenes strains MBT-6 and MBT-7 generated 21 and 24 contigs, respectively. No plasmid sequences could be determined. Genome characteristics are shown in L. monocytogenes strains showed genetic characteristics similar to those of previously reported clinical, food, and outbreak-associated populations   , indicatPRJNA643697. The sequences of STEC strain MBT-5 and L. monocytogenes strains MBT-6 and MBT-7 were deposited at DDBJ/ENA/GenBank under the accession numbers CP058682 (chromosome), CP058683 (plasmid 5.1), CP058684 (plasmid 5.2), CP058685 (plasmid 5.3), CP058686 (plasmid 5.4), CP058687 (plasmid 5.5), CP058688 (plasmid 5.6), JACBGM000000000, and JACBGL000000000. The raw sequencing data were also deposited at the SRA with the accession numbers shown in The genome sequences have been deposited in GenBank under the BioProject number"}
+{"text": "Correction to: Pilot and Feasibility Studies 6, 69 (2020)https://doi.org/10.1186/s40814-020-00613-1Following publication of the original article , the autStudies into TFT for non-refugee traumatized populations show higher effects (d\u2009=\u20091.08\u20141.40) [7], than for refugee populations (g\u2009=\u2009.25\u20141.01) [8].Which will be replaced by:There is evidence that non-refugee traumatized populations benefit more from TFT than traumatized groups with a refugee background .This also involves changes in reference [8].Epidemiology and psychiatric sciences, 28(4), 376\u2013388.doi.org/10.1017/S2045796019000027\u201d.\u201cTurrini, G., Purgato, M., Acarturk, C., Anttila, M., Au, T., Ballette, F., ... & Hall, J. (2019). Efficacy and acceptability of psychosocial interventions in asylum seekers and refugees: systematic review and meta-analysis. should replace:\u201cLambert JE, Alhassoon OM. Trauma-focused therapy for refugees: Meta-analytic findings. Journal of counseling psychology. 2015;62(1):28. DOI: 10.1037/cou0000048\u201d.The publishers apologise for this error."}
+{"text": "E. coli pathogenic lineage ST131 suggesting the important role of flies in disseminating highly virulent pathogens in clinical settings and beyond.Multidrug-resistant gram-negative (MRGN) bacteria are a serious threat to global health. We used genomics to study MRGN obtained from houseflies in a tertiary Rwandan hospital. Our analysis revealed a high abundance of different MRGN including  Escherichia (E.) coli, Klebsiella spp., Enterobacter (E.) cloacae, Acinetobacter spp., and Pseudomonas (P.) aeruginosa, and others, and cause a variety of severe infections like diarrhea, pneumonia, sepsis, endocarditis and urinary tract infection (UTI). Studies estimate 700.000 fatalities caused by antibiotic-resistant pathogens each year with increasing numbers [E. coli from flies captured in a livestock facility was thus unsurprising [Multidrug-resistant gram-negative (MRGN) bacteria include  numbers . In addi numbers . The detrprising . Anotherrprising . We inverprising , flies mE. coli, Klebsiella spp., Enterobacter spp., Acinetobacter spp., P. aeruginosa, Citrobacter spp., and Raoultella spp., we confirmed the bacterial species using MALDI-TOF . Additional phenotypic resistance screening was performed on the VITEK 2 system  and for colistin resistance on 96-well microtiter plates investigating minimal inhibitory concentrations in triplicates. Randomly selected strains . After de-novo assembly using shovill/SPAdes  and Velvet, draft genomes were polished by mapping all trimmed reads back to the contigs with bwa (https://github.com/lh3/bwa/releases/download/v0.7.17/bwa-0.7.17.tar.bz2) and calling variants with Pilon (https://github.com/broadinstitute/pilon/releases/download/v1.23/pilon-1.23.jar). E. coli plasmid sequences of PBIO711 and PBIO1939 were manually extracted using similarity searches (BLASTn Megablast) against the NCBI nucleotide collection for visualization in BRIG (Blast Ring Image Generator) (https://sourceforge.net/projects/brig/files/dev/BRIG-0.95-dev.0004.zip/download). Sequence type (ST), antibiotic resistance/virulence gene and single-nucleotide polymorphism (SNP) detection was carried out using mlst, abricate, and snippy . We inferred a core SNP phylogeny for ST5474. Alignments were filtered for recombinations using Gubbins (https://github.com/sanger-pathogens/gubbins/archive/v2.3.4.tar.gz) and core SNPs extracted using snp-sites . A maximum likelihood tree was inferred with RAxML-NG (https://github.com/amkozlov/raxml-ng/releases/download/0.9.0/raxml-ng_v0.9.0_linux_x86_64.zip) using GTR\u2009+\u2009G. The best-scoring maximum likelihood tree was midpoint-rooted and visualized in FigTree .We examined 42 flies randomly captured in fly traps within 4 weeks in a tertiary hospital in Rwanda in 2014 . SamplinE. coli, 19% (8/42) E. cloacae, 9% (4/42) K. oxytoca, 7% (3/42) C. freundii, 4% (2/42) R. ornithinolytica, 4% (2/42) P. aeruginosa, and 2% (1/42) A. baumannii. Twelve flies (29%) carried more than one antibiotic-resistant bacterial genus of which three  carried three different pathogens  of flies carried antibiotic-resistant bacteria. Thirty-six percent 15/42) carried ESBL-producing 5/42 carrblaCTX-M-15,aac [-IIa, and tet(A)/(B)  Table\u00a0. Eight dd to UTI   causing diarrhea [In addition, we observed five diarrhea . This miE. coli strains , which did not only originate from individual flies captured in different wards but belonged to two different clonal lineages (ST410 and ST617), carried similar resistance genes  (Table The two C) Table . OverallC) Table b.E. coli lineage ST131. High pre-admission and even higher discharge rates at this facility [Our results demonstrate that half of the flies in this tertiary hospital in Rwanda carried virulent MRGN pathogens including the pathogenic clonal facility  may suggfacility  at that facility .Additional file 1: Table S1. Results based on whole genome sequence analysis. Abbreviations: BLA: beta-lactams ; GEN: gentamicin; CIP: ciprofloxacin/moxifloxacin; SXT: sulfamethoxazole-trimethoprim; TET: tetracycline; ST: sequence type; resistance, virulence and plasmid genes are based on the abricate (https://github.com/tseemann/abricate) abbreviations using the databases Resfinder, ARG-ANNOT, CARD, NCBI Bacterial Antimicrobial Resistance Reference Gene Database, PlasmidFinder, VFDB, and Ecoli_VF.Additional file 2: Figure S2. Phylogenomic tree of five E. coli sequence type (ST) 5474 fly isolates (strain names colored according to Fig."}
+{"text": "Drosophila; CRB1-3 in mammals) is a transmembrane determinant of epithelial cell polarity and a regulator of Hippo signalling. Crb is normally localized to apical cell-cell contacts, just above adherens junctions, but how apical trafficking of Crb is regulated in epithelial cells remains unclear. We use the Drosophila follicular epithelium to demonstrate that polarized trafficking of Crb is mediated by transport along microtubules by the motor protein Dynein and along actin filaments by the motor protein Myosin-V (MyoV). Blocking transport of Crb-containing vesicles by Dynein or MyoV leads to accumulation of Crb within Rab11 endosomes, rather than apical delivery. The final steps of Crb delivery and stabilisation at the plasma membrane requires the exocyst complex and three apical FERM domain proteins \u2013 Merlin, Moesin and Expanded \u2013 whose simultaneous loss disrupts apical localization of Crb. Accordingly, a knock-in deletion of the Crb FERM-binding motif (FBM) also impairs apical localization. Finally, overexpression of Crb challenges this system, creating a sensitized background to identify components involved in cytoskeletal polarization, apical membrane trafficking and stabilisation of Crb at the apical domain.Crumbs (Crb in   \u2022Crumbs is transported apically along microtubules and actin filaments.\u2022Dynein is the key microtubule motor protein transporting Crumbs.\u2022Myosin V is the key F-actin motor protein transporting Crumbs.\u2022Crumbs polarises the cytoskeleton, which then directs further Crumbs delivery. How epithelial cells polarize is a fascinating unsolved problem in biology. A key determinant of epithelial polarity is the transmembrane protein Crumbs, which localises specifically to the apical domain of epithelial cells and then helps direct the apical-basal polarization of the entire epithelial cell. How Crumbs itself becomes localized apically is still poorly understood. Here we define a key role for two motor proteins which can transport Crumbs apically along microtubules and actin filaments, respectively. Thus, a polarised cytoskeleton directs Crumbs delivery, which then helps define the apical-basal axis, which feeds back to polarize the cytoskeleton - a positive feedback loop. In fertS.\u00a0pombe . The actS.\u00a0pombe . HoweverS.\u00a0pombe . In Drossymmetry . In epitDrosophila epithelial cells exhibit a more complex polarization than oocytes, with the plasma membrane divided into distinct apical and basolateral domains, separated by a ring of adherens junctions  and Patronin  for acentrosomal nucleation apically . The actcrb and sdt mRNA (Here we show that the actin motor protein Myosin-V (MyoV), previously implicated in Rab11-mediated apical secretion and found to co-localise with Crb , as wellsdt mRNA , both fusdt mRNA  to the asdt mRNA . Crb mus2Drosophila ovarian follicle cell epithelium. Previously, loss of Dynein was reported to cause decreased levels of Crb protein in follicle cells due to failure of crb or sdt mRNA transport  driver  driver . We findle cells C. Co-standosomes D. These dynein-RNAi expressing follicle cells is disrupted by simultaneous expression of kinesin-RNAi, leading to an abnormal depolarised localization of Crb , which has been previously shown to form a complex with Crb and to promote apical secretion from Rab11 endosomes in the receptor . Class Vilaments , 2004, ailaments , suggestilaments , fails tilaments A,B. Use l domain C\u2013E. The extruded F\u2013H, or bextruded . Co-staiextruded F,G. ThesGiven our findings with MyoV, we sought to confirm that the F-actin cytoskeleton is also necessary for apical localization of Crb. As expected, disruption of F-actin by acute treatment with Latrunculin A (Lat A) caused a strong loss of MyoV-GFP and Crb-GFP from the apical domain, with Crb-GFP accumulating in endosomes A\u2013F. TreaDrosophila , Expanded (Ex) and Moesin (Moe). FERM domains link the actin cytoskeleton to the plasma membrane  and bindosophila . Mer andr domain . Moe hasr domain . We find mutants A,B. Doubt on Crb C. Howevelization D. Accordlization  also redlization E,F. ThesS.\u00a0cerevisiae   . Importacomplex) . Thus, tDrosophila neuroblasts, which are polarized along the apical-basal axis by Baz, there is no role for either Crb or membrane trafficking . Notably, CRB3 lacks the homophillic extracellular domain of CRB1 and CRB2, but is still able to localise to tight junctions, suggesting that other tight junction proteins such as JAMs, Occludins or Claudins may mediate extracellular domain clustering, with the entire complex clustered by intracellular multi-PDZ domain proteins such as ZO-1 and MUPP1/PATJ  driver, the actin \u2018flip-out\u2019 and MARCM systems. The tj.Gal4 line is weakly expressed from the beginning of follicular development and strongly from stage 7 onward. For \u2018flip-out\u2019 clones, third instar larvae were heat-shocked at 37\u00a0\u200b\u00b0C for 20\u00a0\u200bmin, and dissected 3 days after eclosion. Fly crosses were kept at a temperature of 25\u00b0.Drosophila stocks were obtained from the Bloomington Drosophila Stock Centre and are described in FlyBase. Mitotic clones were generated using the FLP/FRT system and were either marked positively  or negatively (absence of GFP). Third instar larvae were heat-shocked once at 37\u00a0\u200b\u00b0C for 1\u00a0\u200bh and dissected 3 days after eclosion. Expression of UAS-driven transgenic lines was achieved with 4.1Ovaries were dissected in PBS, fixed for 20\u00a0\u200bmin in 4% paraformaldehyde in PBS, washed for 30\u00a0\u200bmin in PBS/0.1% Triton X-100 (PBT) and blocked for 15\u00a0\u200bmin in 5% normal goat serum/PBT (PBT/NGS). Primary antibodies were diluted in PBT/NGS and samples were incubated overnight at 4\u00a0\u200b\u00b0C. Secondary antibodies were used for 2\u00a0\u200bh at room temperature and then mounted on slides in Vectashield (Vector Labs). Images were taken with a Leica SP5 confocal using 40x oil immersion objective and processed with Adobe Photoshop and ImageJ.Primary antibodies used were: rat anti-Crumbs , mouse anti-Crumbs (Cq4) , rat anti-Crb intra (1:500\u00a0\u200bM.Bhat), rabbit anti-Lgl , mouse anti-Dlg  and FITC-conjugated anti-GFP .Secondary antibodies used were goat Alexa fluor 488, 546 or 647 , Phalloidin  to stain F-actin and DAPI  to visualize nuclei.4.2Crb:GFP was performed by isolating egg chambers and culturing them as described Fig 1A: w; tj.Gal4/+; crb-GFP/UAS.dynein-IR(28054 VDRC)Fig 1B: w hs.flp; actin.FRT.STOP.FRT.Gal4 UAS.GFP/UAS.dynein-IR(28054 VDRC)Fig 1C: w hs.flp; actin.FRT.STOP.FRT.Gal4 UAS.GFP/UAS.dynein-IR(28054 VDRC)Fig 1D: w;crb-GFP/+Fig 2A: w; tj.Gal4/+; crb-GFP/UAS.dynein-IR(28054 VDRC)Fig 2B: w; tj.Gal4/UAS.kinesin-IR; crb-GFP/Fig 2C: w; tj.Gal4/UAS.kinesin-IR; crb-GFP/UAS.dynein-IR(28054 VDRC)Fig 2D: w; tj.Gal4/+; UAS.MyoV-GFP/+Fig 3A: w; tj.Gal4/+; UAS.GFP-MyoV-GT/+Fig 3B: wFig 3C: w; tj.Gal4/+; UAS.GFP-MyoV-GT/+Fig.\u00a03D\u2013E&G: w; tj.Gal4/+; UAS.GFP-MyoV-GT/UAS.dynein-IR(28054 VDRC)Fig.\u00a03F&H: wFig.\u00a04A\u2013B: w; tj.Gal4/+; UAS.MyoV-GFP/+Fig.\u00a04C\u2013D: w;crb-GFP/+Fig.\u00a04E\u2013G: w hs.flp FRT19A moePL106/FRT19A ubi.RFPFig 5A: w hs.flp FRT19A mer4/FRT19A ubi.RFP; FRT40Aexe1/FRT40A GFPFig 5B: w hs.flp FRT19A mer4moePL106/FRT19A ubi.RFPFig 5C: w hs.flp FRT19A mer4moePL106/FRT19A ubi.RFP; FRT40Aexe1/FRT40A GFPFig 5D: w hs.flp; FRT82B crb\u0394FBM/FRT82B ubi.nlsGFP Fig 5E: w hs.flp; FRT82B crb\u0394FBM/FRT82B ubi.nlsGFP Fig 5F: w hs.flp; FRT82Bsec151/FRT82B ubi.nlsGFPFig 5G: yw hs.flp tub.Gal4 UAS.GFPnls/+; FRT40Asec5e10/FRT40A tub.Gal80Fig 5H: wFig 6A: (27654 VDRC)w; tj.Gal4/UAS.patronin-IR(41858 BLOOMINGTON)/+w; tj.Gal4/+; UAS.shot-IRw; tj.Gal4/+; UAS.myoV-IR/+w; tj.Gal4/+; UAS.GFP-MyoV-GT/+w; tj.Gal4/+; UAS.\u03b1-Cat-IR/+1/FRT82B tub.Gal80w hs.flp UAS.GFPnls tub.Gal4/+;FRT82B sec15w; tj.Gal4/UAS.Crb-FLFig 6B: (27654 VDRC)w; tj.Gal4/UAS.Crb-FL UAS.patronin-IR(41858 BLOOMINGTON)/+w; tj.Gal4/UAS.Crb-FL; UAS.shot-IRw; tj.Gal4/UAS.Crb-FL; UAS.myoV-IR/+w; tj.Gal4/UAS.Crb-FL; UAS.GFP-MyoV-GT/+w; tj.Gal4/UAS.Crb-FL; UAS.\u03b1-Cat-IR/+1/FRT82B tub.Gal80w hs.flp UAS.GFPnls tub.Gal4/+; UAS.Crb-FL/+; FRT82B sec1532/FRT82B tub.Gal80yw hs.flp UAS.GFPnls tub.Gal4/+; UAS.Crb-FL/+; FRT82B kibyw hs.flp tub.Gal4 UAS.GFPnls/+; fosCrbICD/+; FRT82B crb11A22/FRT82B tub.Gal80Fig.\u00a06C\u2013E:"}
+{"text": "Enterobacteriaceae (CPE) carriage, we studied 21 CPE carriers for \u00bb1 year. Mean carriage duration was 86 days; probability of decolonization in 1 year was 98.5%, suggesting that CPE-carriers\u2019 status can be reviewed yearly. Prolonged carriage was associated with use of antimicrobial drugs.To determine the duration of carbapenemase-producing  Enterobacteriaceae (CPE) poses a public health threat , and species identification was performed with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry . Antimicrobial susceptibility testing was performed by using VITEK-2 (bioM\u00e9rieux). All Enterobacteriaceae isolates with a MIC of >2 mg/L for meropenem or >1.0 mg/L for ertapenem underwent PCR to detect blaNDM-1, blaKPC, blaOXA-48, blaIMI-1, and blaIMP genes (https://www.illumina.com). We used the Shannon diversity index to measure \u03b1-diversity for fecal microbial communities (https://cran.r-project.org/web/packages/vegan/vegan.pdf).The fecal samples were inoculated onto selective chromogenic agar  colonized, CP\u2013Klebsiella pneumoniae (CP-KP) colonized, and CP-EC/KP co-colonized (https://www.R-project.org) and RStan (http://mc-stan.org).We analyzed data by using Bayesian multistate Markov models to account for interval censoring . First, olonized  Figure 22 of difference in proportions = 14.8, simulated p = 0.0005) (K. pneumoniae (18 [85.7%]) and E. coli (16 [76.2%]). The most frequently observed carbapenemase genes were blaOXA-48 (11 [52.4%]) and blaKPC (8 [38.1%]). We obtained 76 CP-KP isolates from the samples; the most common sequence type (25 [32.9%]) was 307. Among the 83 CP-EC isolates, the most common sequence type (22 [26.5%]) was 131. Sample positivity was continuous until clearance for most (17 [81.0%]) of the 21 participants. For 4 participants, negative samples were followed by positive samples; the longest period was 3 negative samples over 3 consecutive weeks (We enrolled 21 patients . Mean (\u00b1 0.0005)  Table 1.ve weeks  Figure 5The estimated mean duration of CPE carriage was 86  days. The probability of decolonization in 1 year was 98.5% (95% CrI 95.0%\u201399.8%), assuming a constant decolonization rate within the time interval. The longest observed carriage duration was 387 days. We performed a sensitivity analysis that included 16 participants who became decolonized during follow-up . This analysis gave a mean carriage time of 77 (95% CrI 53\u2013108) days and a 98.8% (95% CrI 96.5%\u201399.9%) probability of decolonization within 1 year.As time-fixed covariates, we analyzed age, co-colonization with other multidrug-resistant organisms, presence of a urinary catheter, antimicrobial drug use during follow-up, Charlson Comorbidity Index score, and readmission; as a time-varying covariate, we used the Shannon Diversity Index score to explore the covariates\u2019 association with decolonization  Figure 3CPE infections are typically preceded by asymptomatic carriage, especially in vulnerable patients such as those who are immunocompromised and critically ill  patients had intervening negative samples, 1\u20133 weeks apart. This finding suggests that a patient should have Enterobacteriaceae carriage in hospital patients.Supplementary methods and results for study of duration of carbapenemase-producing"}
+{"text": "Enterobacter hormaechei 2B-MC1, isolated from a shrimp sample collected from a farmer\u2019s market in Atlanta, Georgia. The assembled genome sequence observed was 4,661,561 bp long with a G+C content of 55.3%. The isolate harbored sul1, sul2, qnrA1, oqxB, dfrA23, blaACT, floR, fosA, tet(A), aph(6)-Id, and aph(3\u2033)-Ib antibiotic resistance genes.Here, we announce the draft genome sequence of  Enterobacter hormaechei 2B-MC1, isolated from a shrimp sample collected from a farmer\u2019s market in Atlanta, Georgia. The assembled genome sequence observed was 4,661,561 bp long with a G+C content of 55.3%. The isolate harbored sul1, sul2, qnrA1, oqxB, dfrA23, blaACT, floR, fosA, tet(A), aph(6)-Id, and aph(3\u2033)-Ib antibiotic resistance genes.Here, we announce the draft genome sequence of  Enterobacter hormaechei 2B-MC1 strain from raw farm-raised shrimp from Ecuador which was collected from a farmer\u2019s market in Atlanta, Georgia, on 30 March 2019. Thirty-one shrimp samples were screened for the presence of extended-spectrum \u03b2-lactam strains as previously described . The quality and quantity of the DNA were measured using a NanoDrop One spectrophotometer (Thermo Fisher). Libraries from the DNA sample were prepared and sequenced at a Florida State University  molecular cloning facility. Libraries were sequenced using a MiSeq 2 microkit (Illumina) at 9 pM with 5% PhiX. Default parameters for all software were used during analyses unless otherwise specified. The raw Illumina sequences were quality filtered using Trimmomatic (v0.36) (de novo assembled with Velvet (1.2.10)   v0.36) . The hig . The hiine tool  with theine tool  annotatilvet 1.2. . A total of 1,567 genes were associated with KEGG pathways. The closest relatives of E. hormaechei 2B-MC1 found by MiGA in the database were Enterobacter sp. strain CRENT-193 (GenBank accession number NZ_CP024812.1) and E. hormaechei (NZ_CP030076) with an average nucleotide identity of 99.19%. AMRFinder identified genes and mutations that confer resistance to sulfonamides (sul1 and sul2), quinolones (qnrA1), phenicol/quinolones (oqxB), trimethoprim (dfrA23), beta-lactamases (blaACT), phenicols (floR), fosfomycin (fosA), tetracycline [tet(A)], and aminoglycosides [aph(6)-Id and aph(3\u2033)-Ib]. The following six putative plasmids were present in the draft genome: pESBL176 (MT230180.1), pESBL31 , pESBL87 (MT230381.1), pESBL96 (MT230424.1 and MT230426.1), unnamed4 plasmid (NZ_CP020513.1), and pKP2442_7c331 (NZ_KX434882.1).A total of 4,397,799 paired-end reads were sequenced. These raw sequences were trimmed and then quality filtered (about 8% of the reads were eliminated), resulting in approximately 363,705 reads. These high-quality reads were assembled into contigs. CheckM estimated the completeness of this genome as 99.8%. This assembly produced 160 contigs and an JACVEK000000000. The version described in this paper is the first version, JACVEK010000000. Raw Illumina data have been deposited in the NCBI Sequence Read Archive with the accession number SRR12586817 under the BioProject accession number PRJNA661375.This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession number"}
+{"text": "In this paper, we present the first optimized implementation of ARIA block cipher on low-end 8-bit Alf and Vegard\u2019s RISC processor (AVR) microcontrollers. To achieve high-speed implementation, primitive operations, including rotation operation, a substitute layer, and a diffusion layer, are carefully optimized for the target low-end embedded processor. The proposed ARIA implementation supports the electronic codebook (ECB) and the counter (CTR) modes of operation. In particular, the CTR mode of operation is further optimized with the pre-computed table of two add-round-key, one substitute layer, and one diffusion layer operations. Finally, the proposed ARIA-CTR implementations on 8-bit AVR microcontrollers achieved 187.1, 216.8, and 246.6 clock cycles per byte for 128-bit, 192-bit, and 256-bit security levels, respectively. Compared with previous reference implementations, the execution timing is improved by 69.8%, 69.6%, and 69.5% for 128-bit, 192-bit, and 256-bit security levels, respectively. Data encryption is a fundamental technology for secure network communication in the Internet of Things (IoT). However, the data encryption operation imposes high overheads for low-end microcontrollers. For this reason, the efficient implementation of data encryption is important to achieve the high availability of IoT services. Many block cipher algorithms have been suggested by cryptography researchers to achieve this goal.The international block cipher standard suggested by the National Institute of Standards and Technology (NIST) is the Advanced Encryption Standard (AES) was first introduced in 1998\u00a0. AES hasThe ARIA block cipher was first introduced in 2004\u00a0. This blIn this work, we first optimized the ARIA block cipher on low-end embedded processors. Two modes of operation, including the electronic codebook (ECB) and the counter (CTR) operation, are efficiently implemented with optimized rotation operation, a substitute layer, a diffusion layer, and a pre-computed table for repeated data of the initialization vector (IV) in the CTR.Primitive operations for the ARIA block cipher, including a substitute layer, a diffusion layer, and rotation operation, are efficiently implemented on target 8-bit AVR microcontrollers. The proposed method reduces the number of memory accesses and the number of instructions required for primitive operations. Compared with previous implementations, the proposed implementations for key scheduling and encryption optimized the execution timing by 89.1% and 68.0%, respectively.The ARIA-CTR mode of operation is further optimized with repeated data of IV. Two add-round-key, one substitute layer, and one diffusion layer are pre-computed in the form of a look-up table (LUT). By accessing the pre-computed table, these expensive operations are efficiently optimized away. ARIA-CTR implementations on 8-bit AVR microcontrollers require 187.1, 216.8, and 246.6 clock cycles per byte for 128-bit, 192-bit, and 256-bit key lengths, respectively.The remainder of this paper is organized as follows. A round of the ARIA block cipher consists of three steps, including add-round-key, a substitution layer, and a diffusion layer. The add-round-key performs XOR operation with a 128-bit round key and plaintext. The substitution layer is defined as four types of substitution operations; S-BOX, which are an affine transformation of the inversion function over A overview of the ARIA encryption and decryption processes is presented in The electronic codebook (ECB) mode is the simplest of the encryption modes. The long message is divided into blocks. Each block is encrypted separately.An alternative mode of operation is the counter (CTR) mode. The counter mode turns a block cipher into a stream cipher. The CTR mode generates the next keystream block by encrypting successive values of a counter value.FLASH memory, 8 MHz working frequency, two-stage pipeline design, and 4 KB RAM. The number of available registers is 32. Among them, six registers (i.e R26 \u223c R31) are reserved for address pointers, and the remaining registers are used for general purpose registers. The basic arithmetic instruction takes one clock cycle, while the memory access takes two clock cycles per byte. A detailed instruction set summary for implementation is presented in AVR is a modified Havard architecture 8-bit RISC single-chip microcontroller\u00a0. AVR micA number of implementation studies have been conducted to improve the performance of block ciphers on 8-bit AVR microcontrollers. Block cipher structures are largely divided into two categories. First, Addition, Rotation, and eXclusive- or (ARX)-based block ciphers have been efficiently implemented on low-end microcontrollers\u00a0,14,15,16In WISA\u201913, the LEA block cipher was introduced by an institute attached to Electronics and Telecommunications Research Institute (ETRI)\u00a0. The worIn CHES\u201906, the HIGHT block cipher was introduced\u00a0. 64-bit The US National Security Agency (NSA) presented two lightweight block ciphers, namely, SIMON and SPECK . The SIMZ address pointer for fast memory access. The mix-column computation was efficiently handled with the conditional branch skip. However, previous implementations have mainly focused on ECB mode of operation. However, the CTR mode of operation is most widely used in practice  [Second, Substitution Permutation Network (SPN)-based block ciphers have also been actively investigated. Among them, AES implementations have received considerable attention because the block cipher is an international standard. In , the S-bTLS/SSL) . In CHESTLS/SSL) . The FACTLS/SSL) . With a In this work, we first implemented the ARIA block cipher on low-end 8-bit AVR microcontrollers. Then, the CTR mode of operation for the ARIA block cipher were optimized. By utilizing the repeated IV data and the inner architecture of ARIA, two add-round-key, one substitute layer, and one diffusion layer are replaced with one LUT access.The ARIA block cipher consists of key scheduling, encryption, and decryption functions. As encryption and decryption operations can be performed in one architecture, only the implementation of encryption operation is required. First, the ARIA-ECB mode of operation is optimized. This is the most basic mode of operation for block ciphers, in which 128-bit plaintext is encrypted with the ARIA encryption in specific security keys . The encryption operation outputs 128-bit ciphertext.Key scheduling generates round keys based on the master key. First, the master key is transformed to 128-bit variables :Instead of registers, these variables are stored in a Z address pointer, the target address of the STACK memory is accessible.After the address setting by adjustment of the 0x00 value. Only higher 8-bit of address includes the S-BOX starting address. As the offset of the table is 8-bit long, only the lower address must be updated for memory access.The substitute layer of ARIA consists of sixteen 8-bit-wise S-BOX layers, including four S-BOX1 (i.e. SBOX1_tbl) is set to the higher address of the Z pointer. In Steps 2\u20139, four S-BOX1 accesses are performed with input intermediate results  by assigning them to the lower address of the Z pointer (R30). Afterward, results are loaded  from the FLASH memory to input registers .Algorithm 1 Optimized four S-BOX1 accesses in a source code level.Input: Higher address of S-BOX1 SBOX1_tbl, intermediate results .4: MOV R30, reg2Output: Output results 5: LPM reg2, Z1: LDI R31, hi8(SBOX1_tbl)6: MOV R30, reg2: MOV R30, reg18: MOV R30, reg43: LPM reg1, 9: LPM reg4, ZThe memory access is performed in a grouped way. In each group, four S-BOX layers are grouped as shown in The diffusion layer requires several XOR operations with input variables. Some of these XOR operation duplicate each other. The diffusion layer is optimized in  by re-orAlgorithm 2 8-bit optimized diffusion layer [on layer .Input: Intermediate results , and the remaining XOR operations for these registers are also performed. Similarly, the STACK memory  rather than the registers. In Steps 49, 54, 59, and 64, intermediate results are pushed to the STACK memory. Similarly, in the STACK memory in Steps 73, 78, 83, and 88. In Steps 89 to 96, the pushed results are restored from the STACK memory to the registers. In Steps 97 to 104, intermediate results are moved to the output registers for result alignment.Algorithm 3 Proposed implementation of 8-bit optimized diffusion layer in a source code level.Input: Intermediate results (Y0\u223cY15), temporal register .Output: diffusion layer intermediate results (Y0\u223cY15).// 2:EOR TMP1, Y43:EOR TMP1, Y94:EOR TMP1, Y145:MOV Z0, TMP16:EOR Z0, Y67:EOR Z0, Y88:EOR Z0, Y139:MOV Z5, TMP110:EOR Z5, Y111:EOR Z5, Y1012:EOR Z5, Y1513:MOV Z11, TMP114:EOR Z11, Y215:EOR Z11, Y716:EOR Z11, Y1217:MOV Z14, TMP118:EOR Z14, Y019:EOR Z14, Y520:EOR Z14, Y11// 21:MOV TMP1, Y222:EOR TMP1, Y523:EOR TMP1, Y824:EOR TMP1, Y1525:MOV Z1, TMP126:EOR Z1, Y727:EOR Z1, Y928: EOR Z1, Y1229: MOV Z4, TMP130:EOR Z4, Y031:EOR Z4, Y1132:EOR Z4, Y1433:MOV Z10, TMP134:EOR Z10, Y335:EOR Z10, Y636:EOR Z10, Y1337:MOV Z15, TMP138:EOR Z15, Y139: EOR Z15, Y440:EOR Z15, Y10// 41:MOV TMP1, Y142:EOR TMP1, Y643:EOR TMP1, Y1144:EOR TMP1, Y1245:EOR TMP1, Y1246:EOR TMP2, Y447:EOR TMP2, Y1048:EOR TMP2, Y1549:PUSH TMP250:MOV TMP2, TMP151:EOR TMP2, Y352:EOR TMP2, Y853:EOR TMP2, Y1354:PUSH TMP255:MOV TMP2, TMP156:EOR TMP2, Y057:EOR TMP2, Y558:EOR TMP2, Y1459:PUSH TMP260:MOV TMP2, TMP161:EOR TMP2, Y262:EOR TMP2, Y763:EOR TMP2, Y964:PUSH TMP2// 65:MOV TMP1, Y066: EOR TMP1, Y767:EOR TMP1, Y1068:EOR TMP1, Y1369:MOV TMP2, TMP170: EOR TMP2, Y571:EOR TMP2, Y1172:EOR TMP2, Y1473:PUSH TMP274:MOV TMP2, TMP175:EOR TMP2, Y276:EOR TMP2, Y977:EOR TMP2, Y1278:PUSH TMP279:MOV TMP2, TMP180:EOR TMP2, Y181:EOR TMP2, Y482:EOR TMP2, Y1583:PUSH TMP284:MOV TMP2, TMP185:OR TMP2, Y386:EOR TMP2, Y687:EOR TMP2, Y888: PUSH TMP2//Finalization89:POP Y1390:POP Y891:POP Y692:POP Y393:POP Y1294:POP Y995:POP Y796:POP Y297:POP Y298:MOV Y5, Z599:MOV Y11, Z11100:MOV Y14, Z14101:MOV Y1, Z1102:MOV Y4, Z4103:MOV Y10, Z10104:MOV Y15, Z15The 8-bit optimized diffusion layer approach is efficiently implemented on 8-bit AVR microcontrollers. Detailed descriptions are given in Algorithm 3. The process of x) is efficiently implemented on 8-bit AVR microcontrollers. First, the offset for multiple of 8-bit is performed byte-wise rather than bit-wise. Then, the remaining offset is performed bit-wise. The ARIA block cipher requires five different rotation operations. The 8-bit optimized rotation operation is as follows.The ARIA block cipher requires 128-bit wise rotation operation. Multi-precision rotation on 128-bit wise data (reg16.Algorithm 4ROR_1: 1-bit right rotation for 128-bit data.Input: Intermediate results (reg1\u223creg16)Output: 1-bit right rotated intermediate results (reg1\u223creg16)1:BST reg16, 02:LSR reg13:ROR reg24:ROR reg35:ROR reg46:ROR reg57:ROR reg68:ROR reg79:ROR reg810:ROR reg911:ROR reg1012:ROR reg1113:ROR reg1214:ROR reg1315:ROR reg1416:ROR reg1517:ROR reg1618:BLD reg1, 7Taking an example of 19-bit right rotation, 2-byte is right rotated first and then 3-bit is right rotated. Efficient 1-bit right rotation for 128-bit data is given in Algorithm 4. In Step 1, the most significant bit is cached. Afterward, 1-bit is shifted from the least significant byte to the most significant byte. In Step 18, the least significant bit is replaced with the cached bit from MOVW instruction, which ensures 2-byte-wise register copying. In Steps 10 to 12, the remaining 3-bit right rotation is performed with Algorithm 4  by calling 3 times.Algorithm 5ROR_19: 19-bit right rotation for 128-bit data.Input: Intermediate results (reg1\u223creg16), temporal registers (tmp_reg1)Output: 19-bit right rotated intermediate results (reg1\u223creg16).1:MOVW tmp_reg1, reg152:MOVW reg15, reg133:MOVW reg13, reg114:MOVW reg11, reg95:MOVW reg9, reg76:MOVW reg7, reg57:MOVW reg5, reg38:MOVW reg3, reg19:MOVW reg1, tmp_reg110:ROR_1 reg1, \u2026, reg1611:ROR_1 reg1, \u2026, reg1612:ROR_1 reg1, \u2026, reg16The process of 19-bit right rotation for 128-bit data is given in Algorithm 5. First, 16-bit wise  right rotation is performed with the Algorithm 6ROL_1: 1-bit left rotation for 128-bit data.Input: Intermediate results (reg1\u223creg16), temporal register (tmp_reg).Output: 1-bit left rotated intermediate results (reg1\u223creg16).1:CLR tmp_reg2:LSL reg163:ROL reg154:ROL reg145:ROL reg136:ROL reg127:ROL reg118:ROL reg109:ROL reg910:ROL reg811:ROL reg712:ROL reg613:ROL reg514:ROL reg415:ROL reg316:ROL reg217:ROL reg118:ADC reg16, tmp_regEfficient 1-bit left rotation for 128-bit data is given in Algorithm 6. In Step 1, one register is initialized. Then, 1-bit is shifted to the left from the most significant byte to the least significant byte. In Step 18, the most significant bit is replaced by the carry bit generated from Step 17.MOVW instruction. In Step 11, the remaining 1-bit right rotation is performed with Algorithm 6.Algorithm 7ROR_31: 31-bit right rotation for 128-bit data.Input: Intermediate results (reg1\u223creg16), temporal registers (tmp_reg1).Output: 31-bit right rotated intermediate results (reg1\u223creg16).1:MOVW tmp_reg3, reg152:MOVW tmp_reg1, reg133:MOVW reg15, reg114:MOVW reg13, reg95:MOVW reg11, reg76:MOVW reg9, reg57:MOVW reg7, reg38:MOVW reg5, reg19:MOVW reg3, tmp_reg310:MOVW reg1, tmp_reg111:ROL_1 reg1, \u2026, reg16, tmp_reg1The process of 31-bit right rotation for 128-bit data is given in Algorithm 7. First, 32-bit wise  right rotation is performed with the As shown in In this section, efficient implementations of ARIA-CTR encryption for low-end processors are proposed. The main idea is caching the primitive operations of the ARIA block cipher; this approach skips the operations by the add-round-key of round 2.The first operation of the ARIA block cipher is add-round-key. This is a byte-wise XOR operation with plaintext and round keys. In particular, the CTR mode of operation assigns a (non-constant) 32-bit counter and a (constant) 96-bit IV. Between the first and second blocks, only counter 1 is different in the 32-bit counter section. After the add round key operation, this difference is maintained because it only performs XOR operations. By exploiting this condition, the output of the add-round-key operation can be cached except the counter parts. Detailed descriptions are given in The cache table is further extended to the add-round-key operation of round 2. The substitution layer only updates the data byte-wise. The other (constant) bytes are maintained and can be cached. This is presented in detail in However, for the diffusion layer, one byte updates other bytes. Taking an example of After the 256-th block, 2 bytes , RAM (byte), and execution time (clock cycles per byte). The software was implemented over Atmel Studio 7, and the code was compiled in -O2 option. All ARIA implementations are written in assembly language. The function call and variable assignment are written in C language.The proposed ARIA implementations were evaluated on a low-end 8-bit ATmega128 microcontoller. The microcontroller supports a 128KB In Previous works saved four S-BOX tables in RAM. Each S-BOX table requires 256-byte\u00a0[FLASH memory, which reduces the expensive RAM consumption\u00a0[The execution timings of the proposed ARIA-128-ECB for key scheduling, encryption, and decryption are 214.9, 198.3, and 198.3 clock cycles per byte, respectively. Compared with previous reference implementations, the proposed implementations for key scheduling and encryption optimized the execution timing by 89.1% and 68.0%, respectively [The execution timing of the proposed ARIA-CTR-128 requires 187.1 clock cycles per byte. This result is 5.6% faster than the speed-optimized ECB implementation. Similarly, the implementations of ARIA-CTR-192 and ARIA-CTR-256 require 216.8 and 246.6 clock cycles per byte, respectively. These are faster than ECB implementations by 4.9% and 4.3%, respectively. The code sizes of the speed-optimized ARIA-CTR implementation for key scheduling and encryption are 5938 bytes and 3602 bytes, respectively. Compared with the ECB implementation, the CTR implementation requires 1 KB more for the pre-computed substitute layer and diffusion layer.In this paper, we presented the optimized implementation of the ARIA block cipher on AVR microcontrollers. Optimization techniques are generally divided into AVR specific optimization and generic optimization. In this section, we describe these optimizations in detail.First, memory access is efficiently performed in a grouped way. The memory address is aligned 8-bit wise, which ensures multiple memory accesses with simple offset modifications. This is described in detail in Algorithm 1.Second, the 8-bit optimized diffusion layer is presented. The target microcontroller has a limited number of registers. The proposed approach reduces the number of memory accesses by utilizing available registers. This is described in detail in Algorithm 3.Finally, 5 different rotation operations are optimized for the 8-bit microcontroller. This reduces the offset only below 8-bit wise. This is described in detail in Algorithms 4 and 5.Although the ARIA-CTR encryption (ACE) method is optimized for low-end microcontrollers, the proposed method is a generic algorithm. For this reason, the ACE method can optimize the implementation of ARIA-CTR encryption on other platforms, such as 32-bit ARM and Intel processors, without difficulties. The main idea of the proposed method is pre-computation of the ARIA round function. Because the 96-bit nonce value is constant, a large portion of the round function can be re-used. The pre-computed table skips two add-round-key, one substitute layer, and one diffusion layer operations.In this paper, we proposed optimized implementations of ARIA\u2013ECB and ARIA-CTR on low-end 8-bit AVR microcontrollers. The implementation of ARIA\u2013ECB is improved with optimized rotation, substitute layer, and diffusion layer operations. Then, ARIA\u2013CTR implementation is further optimized with two cache tables. This novel approach skips ARIA\u2013CTR computations by the add-round-key operation of Round 2. With these efficient implementation methods, ARIA-CTR implementations on 8-bit AVR microcontrollers require 187.1, 216.8, and 246.6 clock cycles per byte for 128-bit, 192-bit, and 256-bit, respectively.In future work, the proposed method will be applied to other lightweight block ciphers, such as SIMON and SPECK. Furthermore, we will investigate other microcontrollers to achieve high-speed implementation of the ARIA block cipher."}
+{"text": "Few longitudinal studies link objectively assessed sleep and cognitive performance in ecologically-valid environments. Participants enrolled in the community-based Einstein Aging Study cohort . Wake after sleep onset (WASO) was associated with worse cognitive performance with and without MCI . Models include age, gender, ethnicity, education, learning effects, sleep duration, WASO*MCI interaction . Associations were stronger among those with MCI, thirty minutes more nightly WASO predicted a 500ms longer Symbol Match response time (p<0.0001), 5.05% higher Color Dot error proportion (p=0.002), 0.184 points lower Color Shape accuracy (p<0.0001). In those without MCI, WASO was associated with worse cognition: thirty minutes more nightly WASO predicted +166.7ms Symbol Match response time (p=0.03) and -0.06 points Color Shape accuracy (p=0.013). Actigraphic sleep quality associated with ambulatory cognitive performance (and worse with MCI status) suggests targets for prevention/mitigation of cognitive decline. Part of a symposium sponsored by the Sleep, Circadian Rhythms and Aging Interest Group."}
+{"text": "Correction to: BMC Cell Biol 17, 21 (2016)https://doi.org/10.1186/s12860-016-0101-0Following publication of the original article , an erroc Morphological changes in HKGECs induced by TGF-\u03b21 (20\u2009ng/ml) were examined using phase contrast microscopy. Cells were counterstained with DAPI to visualize nuclei (blue).Fig. 1"}
+{"text": "Correction to: World Journal of Emergency Surgery 15, 88 (2020)https://doi.org/10.1186/s13019-020-01143-wThe age in Biological AVR group should instead be 65.6 +/\u2212\u20094.3The age in Mechanical AVR group should instead be 64.8 +/\u2212\u20095.1The original article  contains"}
+{"text": "Correction to: BMC Biol 18, 163 (2020)https://doi.org/10.1186/s12915-020-00893-2The original publication of this article  containeIn this correction article the correct and incorrect version of figure 4 are published along with the updated Additional file The original article has been updated.Incorrect figure 4.Correct Figure Additional file 1: Fig. S1. Excision of the primary tumor elicits gradual regression of early-stage metastases. Table S1. Cytokines pointed out by the cytokine array. Fig. S2. ELISA validation of in-vitro tumor secretion of the chosen cytokines. Fig. S3. Elevated levels of Serpin E1, IL-8, MIF and PDGF-AA are correlated to poor survival in lung cancer patients. Fig. S4. Associations between levels of DKK1, IL-6, M-CSF and LIF and survival in breast cancer patients."}
+{"text": "Scientific Reports 10.1038/s41598-019-48799-6, published online, 23 August 2019Correction to: This Article containedan error in the Author Information section where:\u201cE. Rasmark Roepke, V. Bruno, M. Rub\u00e9r and J. Ernerudh contributed equally.\u201dnow reads:\u201cE. Rasmark Roepke and V. Bruno contributed equally. M. Rub\u00e9r and J. Ernerudh jointly supervised this work.\u201dThis error has now been corrected in the PDF and HTML versions of the Article."}
+{"text": "Correction to: BMC Evolutionary Biology (2020) 20:44https://doi.org/10.1186/s12862-020-01605-8Following publication of the original article , the autFig. 1 STRUCTURE bar plot showing the individual membership coefficients assigned to each K number of groups (K\u2009=\u20092) defined as populations. The individuals are classified according to their sampling site: 1) Carrizal de Bravo, Guerrero. (2) San Mateo R\u00edo Hondo, Oaxaca. (3) Motozintla, Chiapas. (4) Tacan\u00e1 volcano, Chiapas. (5) San Crist\u00f3bal de las Casas, Chiapas. (6) Acatenango volcano, Chimaltenango. (7) Chimaltenango Quetzaltenango. (8) Totonicap\u00e1n. (9) Sierra de los Cuchumatanes, Huehuetenango."}
+{"text": "The article presents a variant of maturity onset diabetes of the young type 2, caused by a rare mutationin the GCK gene. Maturity onset diabetes of the young (MODY) is a hereditary form of diabetes with an autosomaldominant type of inheritance, an onset at a young age, and a primary defect in pancreatic \u03b2-cell function. Thistype of diabetes is different from classical types of diabetes mellitus (DM1 and DM2) in its clinical course, treatmentstrategies, and prognosis. Clinical manifestations of MODY are heterogeneous and may vary even amongmembers of the same family, i. e., carriers of identical mutations. This phenotypic variation is due to the interactionof mutations with different genetic backgrounds and the influence of environmental factors . Usingnext-generation sequencing technology, the c.580\u20131G>A substitution  located in anacceptor splice site of intron 5 of the GCK gene was found in a proband. The identified variant cosegregated witha pathological phenotype in the examined family members. The GCK gene encodes glucokinase (hexokinase 4),which catalyzes the first step in a large number of glucose metabolic pathways such as glycolysis. Mutations in thisgene are the cause of MODY2. The illness is characterized by an insignificant increase in the fasting glucose level, isa well-controlled disease without medication, and has a low prevalence of micro- and macrovascular complicationsof diabetes. The presented case of MODY2 reveals the clinical significance of a mutation in the splice site of theGCK gene. When nonclassical diabetes mellitus is being diagnosed in young people and pregnant women, genetictesting is needed to verify the diagnosis and to select the optimal treatment method.Key words: human; maturity onset diabetes of the young; MODY2; glucokinase gene; next-generation sequencing;genetic analysis; bioinformatics. Maturity onset diabetes of the young (MODY) is a hereditaryform of diabetes with autosomal dominant inheritance and ischaracterized by onset at a young age and by the presenceof an initial defect in pancreatic \u03b2-cell function. This type ofdiabetes differs from classic types of diabetes mellitus-type 1(DM1) and type 2 (DM2) in disease progression, in treatmentstrategies, and prognosis . Up to 80 % ofMODY cases are not detected or are misdiagnosed as DM1 orDM2; therefore, patients with an incorrectly diagnosed typeof diabetes are often prescribed inadequate therapy . On average, MODY is detected in 2\u20135 % of casesof diabetes (the rest being mostly DM1 and DM2) . To reliably diagnose MODY in a patient, moleculargenetic analysis should be carried out. To date, 14 types ofMODY (MODY1 through MODY14) have been identified,each associated with mutations in a specific gene: HNF4A,GCK, HNF1A, PDX1, HNF1B, NEUROD1, KLF11, CEL,PAX4, INS, BLK, KCNJ11, ABCC8 and APPL1 . FourteenMODY-associated genes explain 70\u201385 % of the disease casesand are involved in various stages of glucose metabolismregulation . According to various researchers, 11to 30 % of MODY cases are caused by mutation in other genes. These formsof MODY are commonly referred to as MODY-X. Becausethe vast majority of pathogenic mutations are found in exonsand adjacent splicing sites of genes , it isreasonable to perform whole-exome sequencing on genomicDNA from individuals with MODY, with subsequent genetictesting of their relatives for the identified mutation. MODYverification allows for successful patient management andensures healthy pregnancy and provision of genetic counselingto families . Examination of relatives ofMODY probands makes it possible to diagnose hyperglycemiain the preclinical phase.In this report, we describe a clinical case of a family withMODY2 associated with a rare splice site mutation in theglucokinase (GCK ) gene identified by the next-generationsequencing technology.The study protocol was approved by the local Ethics Committeeof the Institute of Internal and Preventive Medicine. Written informedconsent to be examined and to participate in the study wasobtained from each patient. For individuals younger than18 years, the informed consent form was signed by a parentor legal guardian.Blood samples were collected from the ulnar vein forbiochemical analysis in the morning on an empty stomach.Lipid levels  andglucose concentration were determined on a KoneLab 300ibiochemical analyzer  with Thermo Fisher Scientific reagents.https://broadinstitute.github.io/picard/).Genomic DNA was isolated from leukocytes of venousblood by phenol-chloroform extraction . Quality of the extracted DNA was assessed on acapillary electrophoresis system, Agilent 2100 Bioanalyzer. Sequencing of patients\u2019DNA was carried out on an Illumina HiSeq1500 instrument. The enrichment andlibrary preparation were performed with the SureSelectXTHuman All Exon V5 + UTRs Kit . Reads were mapped to the reference human genome(GRCh37) by means of the Burrow\u2013Wheeler Alignment tool(BWA v.0.7.12) . Polymerase chain reaction(PCR)-generated duplicates were removed in the PICARDsoftware (A search for single-nucleotide variants (SNVs) was conductedusing the Genome Analysis Toolkit v.3.3 package bythe procedure for local remapping of short insertions/deletionsand recalibration of read quality . Thedepth of coverage was 34\u00d7 to 53\u00d7. SNVs with genotype qualityscores < 20 and coverage depth < 10\u00d7 were filtered out andexcluded from further analysis. Annotation of the SNVs wasperformed in the ANNOVAR software using the 1000 Genomes Project  and The Genome Aggregation Database(gnomAD)  databases. We selectedthe spectrum of rare and novel sequence variants in MODYgenes .Rare variants were selected if their minor allele frequency(MAF) was \u2264 0.5 in the 1000 Genomes Project and gnomAD.Heterozygous substitution c.580\u20131G>A (IVS5 \u20131G>A) at anacceptor splice site of intron 5 of the GCK gene was found in the proband and her sister. To predict the possible effectof the SNV on splicing regulation, we employed the SPANRsoftware .The substitution was corroborated by Sanger sequencingof the DNA fragment containing exons 5 and 6, intron 5,and parts of introns 4 and 6 using the following forward andreverse primers: 5\u2032-CAGGGAGCCTCAGCAGTCTGGA-3\u2032and 5\u2032-GCCACGAGGCCTATCTCTCCCC-3\u2032. The oligonucleotideswere designed in the Primer-Blast software (https://www.ncbi.nlm.nih.gov/tools/primer-blast/) and were synthesizedby the Biosset company . Thesequencing reactions were carried out on an automated ABI3500 DNA sequencer  withthe BigDye Terminator v3.1 Cycle Sequencing Kit . PCR was set up using BioMasterLR HS-PCR (2\u00d7) , 1 \u03bcL of each primer,and 1 \u03bcL of DNA, with a total final volume of 25 \u03bcL. Thethermocycling program consisted of initial denaturation at94 \u00b0C for 3 min and then 35 cycles at 94 \u00b0C for 30 s, 66 \u00b0Cfor 30 s, and 72 \u00b0C for 50 s. The PCR products were evaluatedby electrophoresis in a 5 % polyacrylamide gel after visualizationwith an ethidium bromide solution. A 100 bp DNALadder (BioLabMix) was simultaneously run on the gel asmolecular size markers. The amplicons were purified usingAgencourt AMPure Xp beads . Thesequencing reactions were conducted on an automated ABI3500 DNA sequencer  viathe BigDye Terminator v3.1 Cycle Sequencing Kit (ThermoFisher Scientific). The sequences were analyzed in the VectorNTI\u00ae Advance software (Thermo Fisher Scientific). The hg19version of the human genome served as a reference sequencefor the alignment.The white European 44-year-old female proband was undermedical observation. When she underwent routine screening in2012 (at age 40), hyperglycemia at 7.2 mmol/L was revealed.No complaints were registered. During subsequent glycemiacontrol, maximal fasting glucose was 7.2 mmol/L, and thepostprandial one was 8.9 mmol/L. C-peptide was 1.83 ng/mL(reference range 0.5\u20133.2 ng/mL), immunoreactive insulin was7.9 \u03bcU/mL (reference range 2.0\u201325.0 \u03bcU/ mL), and glycatedhemoglobin (HbA1c) was 7.1 %. Antibodies to insulin, topancreatic islet cells, and to glutamic acid decarboxylasewere absent. Blood biochemical analysis and determinationof thyroid status did not reveal any abnormalities. Ultrasonographyof internal organs, echocardiography, and a studyof brachiocephalic vessels did not uncover any pathology.The body mass index (BMI) was 20.2 kg/m2. DM2 was diagnosedin the patient, and sitagliptin was prescribed. At theage of 26, the patient spontaneously delivered a healthy girlat 39 weeks of gestation; hyperglycemia was not detectedduring the pregnancy.The sister of the proband is a white European 35-yearoldwoman. At age 23, during tests before mastectomy formastopathy, she got a diagnosis of fasting hyperglycemia(6.3 mmol/L). The patient did not have any complains, anda proper diet was recommended. At the age of 29, duringadditional examination before cholecystectomy for cholelithiasis,she received a diagnosis of DM2, and vildagliptin was prescribed at the dose of 50 mg twice a day. At age 31, the proband\u2019ssister visited an endocrinologist at the outpatient clinicof the Institute of Internal and Preventive Medicine  withcomplaints of a failure to get pregnant within a year. On examination,BMI was 20.6 kg/m2, and the objective status was unremarkable.Blood biochemical analysis revealed hypercalcemia(3.25 mmol/L), increased levels of high-density lipoproteincholesterol (85 mg/dL), hypercholesterolemia (220 mg/ dL),and hyperglycemia (6.8 mmol/L), but other analyzed parameterswere within reference ranges. The HbA1c level was7.1 %. Antibodies to insulin, to pancreaticislet cells, and toglutamic acid decarboxylase were absent. Thyroid-stimulatinghormone concentration was 0.759 mU/ mL (referencerange0.4\u20134.0), whereas the prolactin level was 216 ng/ mL (referencerange 1.2\u201319.5). Echocardiography, Doppler sonography ofextracranial parts of cerebral vessels, and abdominal and renalultrasonographic examination revealed no pathology. Cystswere found in both thyroid lobes during the ultrasonography.Given the existence of the proband\u2019s relatives with impairedglucose metabolism, persistence of normal C-peptide levels,the absenceof diabetes-associated autoantibodies, normalBMIs of the proband and her sister, and stable mild hyperglycemia,MODY was assumed.Exons and adjacent splice sites of MODY-associated geneswere analyzed by whole-exome sequencing in the proband andher sister. As a result, heterozygous substitution c.580\u20131G>A(IVS5 \u20131G>A) at an acceptor splice site of intron 5 of theGCK gene was found in the proband and her sister. The IVS5(\u20131G>A) polymorphism of GCK was submitted in ClinVarwith an accession number of rs1554335421 , but was absent in the 1000 Genomes Project , in gnomAD projectdatabases at the moment of publication. Subsequent geneticanalysis by Sanger sequencing of the family members  uncovered segregationof the substitution with DM as an autosomal dominanttrait .Our results, literature data, and databasessuggest that this splice site mutation is likely pathogenic.After confirmation of GCK-MODY in the proband and hersister, their relatives were screened for carbohydrate metabolism disorders. The proband\u2019s mother and daughter did nothave any abnormalities. The proband\u2019s father showed impairedfasting glucose. No complaints were registered, venous plasmafasting glucose was 6.3 mmol/L, and 2 h after the oral glucosetolerance test, it was 7.5 mmol/L. At present, the man doesnot take any medication. The same heterozygous substitutionrs1554335421 (IVS5 \u20131G>A) in the proband\u2019s father\u2019s GCKgene was detected by genetic testing.The proband\u2019s sister had her first pregnancy in 2014 (atage 31). In 2015, a boy weighing 3640 g was born by a caesareansection at 39 weeks of gestation. The pregnancy wascomplicated: premature rupture of membranes, weakness oflabor, and fetal hypoxia. After delivery, due to stable glycemicindexes, it was decided that insulin therapy should bediscontinued. In January 2018, during treatment with diet,the patient\u2019s HbA1c was 6.4 %.The neonatal period of the proband\u2019s nephew was unremarkable.In 2017, his blood biochemical analysis resultedin a diagnosis of hyperglycemia (6.9 mmol/L). HbA1c was6.3 %, and the C-peptide level was 0.54 ng/mL. Antibodiesto insulin, pancreatic \u03b2-cells, and glutamic acid decarboxylasewere undetectable. The same heterozygous substitutionrs1554335421  in the GCK gene was identifiedby genetic testing. At present, the child is under medical observationat Almazov Federal Medical Research Centre ; because of GCK-MODY, a balanceddietwas recommended.DiscussionIt is known that in young patients with impaired carbohydratemetabolism, DM1, DM2, or rarer monogenic forms of diabetesmay be diagnosed. At the onset of the disease, the probandand her sister had no symptoms characteristic for the commontypes of diabetes, fasting hyperglycemia was not progressing,and carbohydrate metabolism disorders were detected duringroutine screening. The presence of DM in the proband\u2019s sister,persistence of normal C-peptide levels, a lack of autoantibodies,and a normal BMI in the proband and her sister pointedto MODY .Heterozygous splice site mutation c.580\u20131G>A(rs1554335421) in intron 5 of their GCK gene was identifiedby genetic testing. Mutations in this gene are associated withDM2, MODY, and neonatal DM . More than 600 variants of the GCK gene associated with MODY have been described, and the list ofthe mutations is constantly growing. The vast majority of themutations are missense substitutions, but splice site mutations,deletions, and insertions are reported too .The GCK gene is located in chromosomal region 7p15.3-p15.1 and consists of 12 exons that encode a 465-amino-acidprotein, glucokinase , which is one of fourmembers of the hexokinase family of enzymes. In 1992, GCKwas the first gene to be linked to MODY. It plays an importantregulatory role in glucose metabolism. Glucokinase catalyzesphosphorylation of glucose to produce glucose-6-phosphate asthe first step of glycolysis in pancreatic \u03b2-cells . Most individuals with heterozygousGCK mutations show fasting plasma glucose levelsbetween 5.5 and 8.0 mmol/L and a small increase in plasmaglucose (< 3 mmol/L in 70 % of the patients) 2 h after the oralglucose test . This feature also explainsasymptomatic fasting hyperglycemia (HbA1c range 5.8\u20137.6 %(40\u201360 mmol/mol)) and rare microvascular and macrovascularcomplications in patients with GCK-MODY . Most patients have an aberrantfasting glucose level or impaired glucose tolerance, and lessthan 50 % of the affected individuals have diabetes, whichis diagnosed during childhood, adolescence, or pregnancy. In a study on Italian patients under 18years of age with incidental hyperglycemia, it was estimatedthat 15 % of these cases are caused by GCK mutations .It was found here that the proband and her sister carrya heterozygous substitution, c.580\u20131G>A , at an acceptor splice site of GCK intron 5.This allelic variant is of interest because consensus donor(GT dinucleotide) and acceptor (AG dinucleotide) splicesites are highly conserved. Point mutations at these loci canlead to cryptic splice site activation and synthesis of aberrantprotein isoforms.In silico analysis of the functional significance of this substitutionsuggested that the inclusion of exons 5, 6, and 7 ingene transcripts will be reduced in case of the detected variant(see the Table).DM and a family history of gestational diabetes. Experimentson lymphoblastoid cells indicate that rs1554335421(IVS5 \u20131G>A) of GCK can activate a cryptic splice site inintron 5 and cause retention of 27 bp of the intron . Information about rs1554335421 (IVS5 \u20131G>A)of GCK is absent in the 1000 Genomes Project, The ExomeAggregation Consortium, and GNOMAD (https://gnomad.broadinstitute.org/) databases; however, taking into accountthe previous study and the data we obtained here, carriage ofthe A allele at position \u20131 of intron 5 is most likely a causative dominant variant of the GCK gene in MODY-affected people.Cryptic splice site activation and formation of several alternativetranscripts with intron 7 fragments\u2019 retention weredemonstrated in a system of model GCK minigenes withacceptor site mutation IVS7 (\u20131G>C) .Model mice with the homozygous mutation in the splicesite of \u03b2-cell-speci\ufb01c exon 1 IVS1A (\u20131G>T) show hyperglycemia,glucosuria, and growth retardation and die within the\ufb01rst week after birth. This phenotype can be explained by exonskipping or intron retention . Splicing sitesaffected by mutations have been described for many pathologicalphenotypes: neurofibromatosis type 1 ,familial hypercholesterolemia ,Wiskott\u2013Aldrich syndrome and chronic colitis , hypophosphatemic rickets , andothers. Mutations affecting splicing have been found not onlyin canonical splicing sites but also in introns and exons andmay have a tissue-specific effect, as in familial dysautonomia. Thatanalysis indicated that the donor splice site mutations weremore prevalent than the acceptor splice site variants (ratio1.5:1.0) . Because the mutationsin the GCK gene can cause a mild clinical phenotype, whichcan vary under the influence of many genetic and lifestylefactors, research on the carriers of these mutations is essentialfor identifying additional risk factors.GCK-MODY is inherited as an autosomal dominant traitmanifested throughout the lifespan as stable, mild fastinghyperglycemia usually reaching 6.7 mmol/L and higher onlyin middle age . A similar patternwas observed here in the proband and her sister. Nonetheless,the metabolic disturbances in the carriers of GCK mutationsare present from birth and can be identified already in the firstyears of life, almost all of them after puberty . The proband\u2019s nephew, who is a heterozygous mutationcarrier, developed carbohydrate metabolism disorders attwo years of age.It has been reported that carriers of GCK gene mutationswith a long history of hyperglycemia (48.6 years on average)usually have micro- and macrovascular complications of diabetesand are at a risk of cardiovascular diseases that is identicalto that in the general population .Patients with GCK-MODY in childhood and adolescencecan be treated only with diet in most cases, and glucoseloweringtherapy should be considered during pregnancy. The present subtype of MODY2 (in theproband and her sister) currently is treated with diet resultingin sufficient glycemic control.The presence or absence of a GCK mutation in the fetusaffects its sensitivity to maternal hyperglycemia . If the fetus does not have the mutation, then itwill secrete insulin excessively and as a result have a risk ofmacrosomia . In that case, low doses ofinsulin should be prescribed during pregnancy . During her pregnancy, the proband\u2019s sister wasgiven insulin injections in small doses. After delivery, insulintherapy was discontinued. Subsequently, the child was foundto carry the same substitution.The presented subtype of MODY2 reveals the clinical significanceof the mutation in a splice site of the GCK gene. Whennonclassical diabetes mellitus is being diagnosed in youngpeople and pregnant women, genetic testing is needed to verifythe diagnosis and to select the optimal treatment method.The authors declare no conflict of interest.Abramowicz A., Gos M. Splicing mutations in human genetic disorders:examples, detection, and confirmation. J. Appl. Genet. 2018;59:253-268. DOI 10.1007/s13353-018-0444-7.An\u0131k A., \u00c7atl\u0131 G., Abac\u0131 A., B\u00f6ber E. Maturity-onset diabetes of theyoung (MODY): an update. J. Pediatr. Endocrinol. Metab. 2015;28:251-263. DOI 10.1515/jpem-2014-0384.Bonnefond A., Philippe J., Durand E., Dechaume A., Huyvaert M.,Montagne L., Marre M., Balkau B., Fajardy I., Vambergue A., VatinV., Delplanque J., Le Guilcher D., De Graeve F., Lecoeur C.,Sand O., Vaxillaire M., Froguel P. Whole-exome sequencing andhigh throughput genotyping identified KCNJ11 as the thirteenthMODY gene. PLoS One. 2012;7(6):e37423. 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+{"text": "Micronutrients are minerals and vitamins and they are essential for normal physiological activities. The objectives of the study were to describe the progress and determinants of micronutrient levels and to assess the effects of micronutrients in the treatment outcome of kalazar.A prospective cohort study design was used. The data were collected using patient interviews, measuring anthropometric indicators, and collecting laboratory samples. The blood samples were collected at five different periods during the leishmaniasis treatments: before starting anti-leishmaniasis treatments, in the first week, in the second week, in the third week, and in the 4th week of anti-leishmaniasis treatments. Descriptive statistics were used to describe the profile of patients and to compare the treatment success rate. The generalized estimating equation was used to identify the determinants of serum micronutrients.Hookworm (\u2212\u20094.48 [\u2212\u20096.82 - -2.14]), chronic diseases (B -7.44 [95% CI: \u2212\u20099.75 - -5.13]), and HIV (B -5.51 [95% CI: \u2212\u20098.23 - -2.78]). The serum selenium level of visceral leishmaniasis patient was affected by HIV (B -18.1 [95% CI: \u2212\u200920.63 - -15.58]) and family size (B -11.36 [95% CI: \u2212\u200913.02 - -9.7]). The iodine level of visceral leishmaniasis patient was affected by HIV (B -38.02 [95% CI: \u2212\u200941.98 - -34.06]), DDS (B 25 .84 [95% CI: 22.57\u201329.1]), smoking (B -12.34 [95% CI: \u2212\u200915.98 - -8.7]), chronic illness (B -5.14 [95% CI: \u2212\u20097.82 - -2.46]), and regular physical exercise (B 5.82 [95% CI: 0.39\u201311.26]). The serum vitamin D level of visceral leishmaniasis patient was affected by HIV (B -9.43 [95% CI: \u2212\u200910.92 - -7.94]), DDS (B 16.24 [95% CI: 14.89\u201317.58]), malaria (B -0.61 [95% CI: \u2212\u20093.37 - -3.37]), and family size (B -1.15 [95% CI: \u2212\u20092.03 - -0.28]). The serum vitamin A level of visceral leishmaniasis patient was affected by residence (B 0.81 [95% CI: 0.08\u20131.54]), BMI (B 1.52 [95% CI: 0.42\u20132.6]), DDS (B 1.62 [95% CI: 0.36\u20132.88]), family size (B -5.03 [95% CI: \u2212\u20095.83 - -4.22]), HIV (B -2.89 [95% CI: \u2212\u20094.44 - -1.34]),MUAC (B 0.86 [95% CI: 0.52\u20131.21]), and age (B 0.09 [95% CI: 0.07\u20130.12]).The mean age of the patients were 32.88\u2009years [SD (standard deviation) \u00b115.95]. Male constitute 62.3% of the patients and problematic alcohol use was present in 11.5% of the patients. The serum zinc level of visceral leishmaniasis patients was affected by alcohol (B\u2009\u2212\u20092.7 [95% CI: \u2212\u20094.01 - -1.5]), DDS (B 9.75 [95% CI: 7.71\u201311.79]), family size (B -1.63 [95% CI: \u2212\u20092.68 - -0.58]), HIV (B -2.95 [95% CI: \u2212\u20094.97 - -0.92]), and sex (B\u2009\u2212\u20091.28 [95% CI: \u2212\u20092.5 - -0.07]). The serum iron level of visceral leishmaniasis patients was affected by alcohol (B 7.6 [95% CI: 5.86\u20139.35]), family size (B -5.14 [95% CI: \u2212\u20097.01 - -3.28]), malaria (B -12.69 [95% CI: \u2212\u200914.53 - -10.87]), The micronutrient levels of visceral leishmaniasis patients were significantly lower. The anti-leishmaniasis treatment did not increase the serum micronutrient level of the patients. The three forms of leishmaniasis are cutaneous leishmaniasis, mucocutaneous leishmaniasis, and visceral leishmaniasis . Male constitute 62.3% of study participants and problematic alcohol use was observed in 11.5% of the patients (Table\u00a0mcg/dl). Female kalazar patients had 1.28 mcg/dl less zinc level than males. High dietary diversification increases the serum zinc level by 9.75 mcg/dl. High family size decreases the serum zinc level by 1.63 mcg/dl. The serum zinc level of HIV positive visceral leishmaniasis patients was 2.95 mcg/dl less than HIV negative visceral leishmaniasis patients. The anti-leishmaniasis treatment did not increase the serum zinc level of the patients.Problematic alcohol use decreases the serum zinc level by 2.7 micrograms per deciliter (mcg/dl. Chronic illness decreases the serum iron level of the patients by 7.44 mcg/dl. Malaria co-infection decreases the serum iron level of the patients by 12.69 mcg/dl. Hookworm infection decreases the serum iron level of visceral leishmaniasis patients by 4.48 mcg/dl. High family size decreases the serum iron level of the patients by 5.14 mcg/dl. HIV infection decreases the serum iron level of the patients by 5.54 mcg/dl. The serum iron level of patient increase by 0.11mcq/dl per a year increase in the patient age. The serum iron level of the patient increase by 0.75 mcq/dl per a centimeter increase in the MUAC of the patient. The anti-leishmaniasis treatment increases the serum iron level of the patient by 0.67 mcg/dl.Problematic alcohol use increases the serum iron level of the patients by 7.6 ng/ml less serum selenium level than HIV negative visceral leishmaniasis patients. High family size decreases the serum selenium level of the patients by 11.36\u2009ng/ml. The anti-leishmaniasis treatment increases the serum selenium level by 3.04\u2009ng/ml.HIV positive visceral leishmaniasis patient had 18.1\u2009mcg/l. High DDS increases the iodine level of the patients by 25.84 mcg/l. Smoking decreases the iodine level of the patients by 12.34 mcg/l. HIV decreases the iodine level of visceral leishmaniasis patients by 38.02 mcg/l. Chronic illness decreases the iodine level of visceral leishmaniasis patients by 5.14 mcg/l. The anti-leishmaniasis treatment increases the iodine level of patients by 13.67 mcg/l.Malaria decreases the iodine level of visceral leishmaniasis patients by 3.78 mcg/dl. Chronic illness decreases the serum vitamin A level of the patients by 2.56 mcg/dl. Urban residence increases the serum vitamin A level of the patient by 0.81 mcq/dl. High DDS increases the serum vitamin A level of the patient by 1.62 mcg/dl. Malaria co-infection decreases the serum vitamin A level of the patient by 4.8 mcg/dl. High family size decreases the serum vitamin A level of visceral leishmaniasis patients by 5.03 mcg/dl. HIV infection decreases the serum vitamin A level of the patients by 2.89 mcg/dl. A centimeter increase in the MUAC of visceral leishmaniasis patients increases the serum vitamin A level by 0.86 mcg/dl. The anti-leishmaniasis treatment did not increase the serum vitamin A level of the patient.Problematic alcohol use decreases the serum vitamin A level of visceral leishmaniasis patients by 1.09 ng/ml. High DDS increases the serum vitamin D level of the patients by 16.24\u2009ng/ml. Malaria decreases the serum vitamin D level of the patients by 0.61\u2009ng/ml. In the presence of hookworm infection, the serum vitamin D level of visceral leishmaniasis patients decreased by 3.94\u2009ng/ml. High family size decreases the serum vitamin D level of the patients by 1.15\u2009ng/ml. HIV co-infection decreases the serum vitamin D level of the patient by 9.43\u2009ng/ml. The anti-leishmaniasis treatment did not increase the serum vitamin D level of visceral leishmaniasis patients (Table A unit increase in the BMI of visceral leishmaniasis patients increases the serum vitamin D level by 1.52\u2009The micronutrient level directly affects treatment outcome of visceral leishmaniasis; especially the treatment outcome was not successful if the serum zinc, iron, vitamin A and vitamin D levels were lower than the first quartile. The overall treatment success rate of visceral leishmaniasis treatment was 84.7% [95% CI: 82.77 - 86.67%] Tables\u00a0, 4.Tablmcg/dl, and the serum vitamin A level by 1.09 mcg/dl. This finding was in line with previous research outputs . A systematic review and meta-analysis estimate also supports this finding .Low serum zinc level decreases the treatment outcome of visceral leishmaniasis. This finding was in line with finding from India . This isHigher patient iron level increases the treatment success rate of visceral leishmaniasis. This finding supports the results of previously published work . This isHigher serum vitamin A and Vitamin D level favors good treatment outcome in visceral leishmaniasis. This finding agrees with previous researchers outputs \u201379. ThisPossible limitation of this study was a failure to address all the vitamins and minerals status of visceral leishmaniasis patients, but since practically it is very difficult to address all of them this study gives the baseline evidence on main vitamins and mineral levels.The serum micronutrient levels of visceral leishmaniasis patients were low. Problematic alcohol use affects the serum zinc, iron, vitamin A levels. DDS affects the serum zinc, iodine, vitamin A, and vitamin D level. Family size affects the serum zinc, iron, selenium, vitamin A, and vitamin D levels. HIV infection affects the serum zinc, iron, selenium, iodine, vitamin A, and vitamin D levels. Anti-leishmaniasis drug slightly increases the serum iodine and selenium levels, but it doesn\u2019t increase the serum iron, zinc, vitamin A, and vitamin D levels. The serum levels of zinc, iron, vitamin A, and vitamin D significantly affect the treatment outcomes of visceral leishmaniasis.The visceral leishmaniasis treatment guideline should incorporate supplementing the micronutrients as part of anti-leishmaniasis intervention."}
+{"text": "Klebsiella pneumoniae (K. pneumoniae) and Escherichia coli (E. coli) were the most common (52.7% and 44.2%). K. pneumoniae was most susceptible to colistin, amikacin, ertapenem, and imipenem . E. coli was most susceptible to colistin (100%), amikacin (94.1%), imipenem (90.4%), and ertapenem (83.6%). ESBL was detected in 96.2% and ESBL genotypes included blaCTX-M-15 (70.1%), blaTEM-OSBL (48.5%), blaSHV-OSBL (27.9%), and blaCTX-M-14 (10.7%). AmpC resistance genes were identified in 9.7% of the isolates, dominated by blaCMY-2 (5.7%). Carbapenem resistance genes were detected in 45.3% of the isolates. In K. pneumoniae, blaOXA-48 dominated (40.6%), followed by blaNDM-1 (23.7%) and blaOXA-232 (4.5%). In E. coli, the most frequent genes were blaNDM-5 (9.6%), blaOXA-181 (5.5%), blaOXA-244 (3.7%), and blaNDM-1 (3.7%). blaKPC-2 was identified in 0.4% of isolates. Notably, 32.3% of isolates carried more than one resistance gene. Our findings emphasize the continued need for molecular surveillance of MDR pathogens, implementation of strict infection control measures, and antimicrobial stewardship policies in our hospitals.High rates of antimicrobial resistance (AMR) among Gram-negative pathogens (GNP) have been reported in Egypt. Antimicrobial surveillance and identifying the genetic basis of AMR provide important information to optimize patient care. In this study, we aimed to identify the beta-lactam resistance phenotypes and genotypes of multidrug-resistant (MDR) non-repetitive GNP from 3 tertiary hospitals in Egypt. WZe studied 495 non-repetitive MDR Gram-negative isolates from patients with complicated intra-abdominal infections (cIAI), complicated urinary tract infection (cUTI), and lower respiratory tract infection (LRTI), collected as part of the \u201cStudy for Monitoring Antimicrobial Resistance Trends\u201d (SMART) conducted in 3 tertiary hospitals in Cairo, Egypt, from 2015 to 2016. Identification and susceptibility testing of GNP to antimicrobials were tested in each hospital laboratory and confirmed in a reference laboratory , Inc., Schaumburg, IL, USA). Molecular identification of extended-spectrum beta-lactamases (ES\u0392Ls), AmpC, and carbapenem resistance genes was conducted in IHMA. Among the 495 MDR isolates,  A high rate of AMR has been reported in Egypt since more than 20\u00a0years, among GNP causing nosocomial infections and outbreaks \u20133. AMR rin Egypt \u20138. As thEnterobacteriaceae, because no CLSI breakpoints exist [This study was conducted in 3 major tertiary care Egyptian hospitals participating in the \u201cStudy for Monitoring Antimicrobial Resistance Trends\u201d (SMART) from 2015 to 2016. The hospitals were Ain Shams University Hospital, Ain Shams Specialized University Hospital, and Dar Al-Fouad Hospital. Isolates were collected according to SMART protocol as previously reported \u201311. Briets exist .The suscblaTEM, blaSHV, blaCTX-M, blaVEB, blaPER, and blaGES; blaAmpC \u03b2-lactamase genes (class C)\u2014blaACC, blaACT, blaCMY, blaDHA, blaFOX, blaMIR, and blaMOX; and carbapenemases (class A)\u2014blaKPC and blaGES; (class B)\u2014blaNDM, blaIMP, blaVIM, blaGIM, and blaSPM; and (class D)\u2014blaOXA-48-like. Then the genes encoding ES\u0392L, carbapenemases, and AmpC were sequenced in their entirety in IHMA [The molecular characterization of ESBL and carbapenemases was done using the Check-Points MDR CT103  microarray kit, which detects most carbapenemase, ESBL, and AmpC genes: ES\u0392Ls (class A)\u2014 in IHMA .Klebsiella pneumoniae and Escherichia coli were the most common (52.7% and 44.2%) and were also the predominant organisms in cIAI , cUTI , and LRTI  . E. coli remained most susceptible to colistin (100%), amikacin (94.1%), imipenem (90.4%), and ertapenem (83.6%).The 495 MDR isolates were derived from cIAI 181, 36%), LRTI , and cUTI . Overall 81, 36%, blaCTX-M-15 (70.1%), blaTEM-OSBL (48.5%), blaSHV-OSBL (27.9%), and blaCTX-M-14 (10.7%). The predominant ESBL gene in both E. coli and K. pneumoniae was blaCTX-M-15 , followed by blaNDM-1 (23.7%) and blaOXA-232 (4.5%). In E. coli, the most frequent genes were blaNDM-5 (9.6%), blaOXA-181 (5.5%), blaOXA-244 (3.7%), and blaNDM-1 (3.7%). blaKPC-2 and blaVIM-2 were less frequently identified (Table Carbapenem resistance genes were detected in 45.3% of the MDR isolates. In blaCMY-2 was the most predominant one (Table blaCTX-M-15 with blaNDM-5 (2.6%); the commonest combination of 3 genes was blaSHV-OSBL, blaCTX-M-15, and blaNDM-1 (9.2%); the commonest combination of 4 genes was blaSHV-OSBL, blaTEM-OSBL, blaCTX-M-15, and blaNDM-1 (12.4%); and the commonest combination of more than 4 genes was blaSHV-OSBL, blaTEM-OSBL, blaCTX-M-15, blaCTX-M-14, blaNDM-1, and blaOXA-48 (5.9%) . We identified 43 resistance phenotypes distributed among isolates from the 3 hospitals. Typing of nosocomial GNP in each hospital based on the phenotypic resistance patterns showed no clonal spread except for few isolates in each hospital (data not shown).K. pneumoniae was a common pathogen in all 3 types of infection, and most isolates were MDR. Due to its large accessory genome including plasmids and chromosomal loci, K. pneumoniae isolates may act as opportunistic pathogens. Such strains infect critically ill and immunocompromised patients mostly, whereas other strains of K. pneumoniae (hyper-virulent) may even infect healthy people in community settings. Many of the virulent strains encode carbapenemases [E. coli was a frequently identified pathogen from cIAI and cUTI, which is consistent with previous reports, and was also a frequent pathogen in LRTI. E. coli pneumonia is uncommon and may result from micro-aspiration of colonized upper airway secretions in severely ill patients; hence, it is a well-known cause of nosocomial pneumonia [E. coli pneumonia may also be community-acquired in patients who have underlying diseases such as diabetes mellitus, alcoholism, chronic obstructive pulmonary disease, and E. coli UTI [E. coli isolates compared with 42.5% and 48.1% in K. pneumoniae isolates, respectively.enemases . E. colineumonia . Howevercoli UTI . Our rescoli UTI  that shocoli UTI , ESBL anblaCTX-M-15 and blaCTX-M-14 are the most commonly identified worldwide as the genes encoding CTX-M enzymes (blaCTX-M) can be horizontally mobilized by various genetic elements [blaCMY-42, blaDHA, and blaACT-like. This is in keeping with recent reports on acquisition of plasmid-mediated cephalosporinase producing Enterobacteriaceae after a travel to the tropics and North Africa including Egypt [E. coli isolates in Egypt [Among the many types of ES\u0392Ls reported, elements . This reng Egypt , 21. Theng Egypt . Moreovein Egypt . This isin Egypt , 25.blaOXA followed by blaNDM genes dominated, while only 2 isolates of K. pneumoniae harbored blaKPC-2 genes. These results confirm previous small-scale reports that the blaNDM and blaOXA genes are the predominant in Egypt and Middle East [The most important mechanism of carbapenem resistance is the production of carbapenemases; therefore, all isolates were investigated to identify the carbapenemase genes. We identified them in 45.4% of tested isolates; dle East .The present study detected massive coexistence of different resistance genes among tested isolates. This coexistence could have contributed to the observed elevated variability in resistance phenotypes and genotypes among GNP in Egypt .In conclusion, our study detected alarming rates of resistance and identified many resistance mechanisms in clinical GNP from Egyptian tertiary care hospitals. These high resistance rates highlight the importance of continuous monitoring of the resistance trends, adherence to infection control policies, and underscore urgently implementing a national antimicrobial stewardship plan in Egypt."}
+{"text": "Cryptophyceae sp. CCMP2293. The circular genome is 139,208\u2009bp in length and contains 142 protein-coding genes (PCGs), 30 transfer RNA (tRNA) genes, 6 ribosome RNA (rRNA) genes, and 1 transfer-messenger RNA (tmRNA) gene. The overall nucleotide composition is: 33.6% A, 32.5% T, 16.8% C, and 17.1% G with a total A\u2009+\u2009T content of 66.1%. The phylogenetic tree was constructed to explore the taxonomic status of Cryptophyceae sp. CCMP2293, which is closely related to G. theta and R. salina.In this study, we present the complete plastid genome of  Cryptophyte algae are an evolutionarily significant group which inhabits marine, brackish, and freshwater environments . The single specimen was provided by the Culture Collection of Marine at the Ocean University of China in Qingdao (OUC-2013060210). Illumina paired-end DNA library was prepared and sequenced using HiSeq 2500 Sequencing System (Gene Denovo Laboratory). The pre-processed sequences were assembled using NOVOPlasty  and aligned via BLASTX and BLASTN searches at NCBI (http://blast.ncbi.nlm.nih.gov/). The tRNAs were identified using the tRNAscan-SE 1.21 web server (http://lowelab.ucsc.edu/tRNAscan-SE/), and the rRNAs were identified using the RNAmmer 1.2 server (http://www.cbs.dtu.dk/services/RNAmmer/). The complete plastid genome of Cryptophyceae sp. CCMP2293 is 139,208\u2009bp in length and contains 142 protein-coding genes (PCGs), 30 transfer RNA (tRNA) genes, 6 ribosome RNA (rRNA) genes and 1 transfer-messenger RNA (tmRNA). There are 4 function-unknown open reading frames. The overall nucleotide composition is: 33.6% A, 32.5% T, 16.8% C, and 17.1% G, with a total A\u2009+\u2009T content of 66.1%.Here, we reported and characterized the complete Cryptophyceae sp. CCMP2293, the phylogenetic tree was constructed with 6 published complete plastid genomes obtained from the Genbank, where Costaria costata served as outgroup. 56 concatenated protein-coding amino acid sequences were aligned using the program MAFFT (Katoh et\u00a0al. Cryptophyceae sp. CCMP2293 clustered with G. theta and R. salina (To elucidate the phylogenetic position of . salina . The det"}
+{"text": "Desulfuromonas sp. strain AOP6, with iron(III)-reducing activity, was isolated from subseafloor sediment in Nankai Trough. We report the complete genome of this strain determined by Illumina MiSeq sequencing and PCR/Sanger sequencing-based gap closing. The genome includes the genes encoding c-type cytochromes, type IV pili, and fatty acid degradation enzymes. Desulfuromonas sp. strain AOP6, with iron(III)-reducing activity, was isolated from subseafloor sediment in Nankai Trough. We report the complete genome of this strain determined by Illumina MiSeq sequencing and PCR/Sanger sequencing-based gap closing. The genome includes the genes encoding c-type cytochromes, type IV pili, and fatty acid degradation enzymes. Geobacter spp., can reduce not only soluble ferric iron but also poorly crystalline Fe(III) oxides (Desulfuromonas sp. strain AOP6 (formerly described as Pelobacter sp. AOP6) was isolated from subseafloor sediment in Nankai Trough using acetate and goethite (\u03b1-FeOOH) as the electron donor and acceptor, respectively  reduction is an important anaerobic respiration process \u20134. Some \u2013) oxides , 5. Howelmitatis , followeylenicus  and Geoaerraneus . Here, whttps://github.com/najoshi/sickle/releases/tag/v1.33) with default settings, a two-step combination of the genome assembly was performed. Briefly, the obtained sequence was preassembled using Unicycler v0.4.8 -NTA] as the electron donor and acceptor, respectively . These tr v0.4.8  with defr v0.4.8  with ther v0.4.8 . This rerrn operons, 2 CRISPR features, 50 tRNA loci, and 3,000 protein-coding sequences (CDSs) were predicted using the DDBJ Fast Annotation and Submission Tool (DFAST) v1.2.2  oxides . Two ) v1.2.2  with def) v1.2.2 . Functio) oxides , 16. Mor) oxides . AdditioDesulfuromonas sp. strain AOP6 has been deposited in DDBJ/ENA/GenBank under accession number AP022810. The raw sequence reads for the paired-end and mate pair libraries are available at accession numbers DRR194005 and DRR194004, respectively.The complete genome sequence of"}
+{"text": "AbstractFothergilla is a small genus of deciduous shrubs native to the southeastern United States that depending on circumscription comprises two to four species. Recent treatments recognized only two species in the genus: F.gardenii (tetraploid) and F.major (hexaploid). Until recently, no diploid taxon of Fothergilla was known. However, recent investigations identified a number of diploid populations in Alabama, Florida, Georgia, and South Carolina. A subsequent phylogenomic analysis showed that the diploids segregated into two, well-supported lineages, corresponding to largely allopatric populations. A re-examination of the morphology of diploid plants, in combination with the genetic evidence, has led us to the recognition of two species of diploids in the genus \u2013 a resurrected F.parvifolia and a new species  \u2013 bringing the total number of recognized species in Fothergilla to four. A revised taxonomic treatment of the genus is provided. Fothergilla L.  is a small genus of deciduous shrubs native to the southeastern United States that depending on circumscription comprises two to four species . F.major based on \u201chaving a spike of flowers, three inches or more in length; [...] later flowering, and [...] leaves [...] very broad, and much more toothed\u201d , leaf base (cordate in F.parvifolia vs. cuneate to rounded in F.gardenii), and leaf margin (toothed from below the middle to the apex in F.parvifolia vs. toothed only near the apex in F.gardenii). This circumscription was followed by F.parvifolia as a synonym of F.gardenii (F.gardenii (incl. F.parvifolia) and F.major (incl. F.monticola) (Kearney in  segregatgardenii . In factnticola) .Fothergillagardenii is found in wet savannas and pocosins in the coastal plains of North Carolina, South Carolina, Georgia, Florida, and Alabama, whereas F.major occurs primarily in woodlands, bluffs, and riverbanks of the upper Piedmont and mountains of North Carolina, South Carolina, Georgia, Alabama, Tennessee, and Arkansas , smaller leaves (<5.2 cm wide vs. > 5.2 cm wide in F.major), leaf dentations , base symmetry (symmetric vs. asymmetric in F.major), hypanthium length (3\u20134.5 mm vs. 4\u20139.2 mm in F.major), number of stamens per flower (12\u201324 vs. 22\u201332 in F.major), and seed size (4.8\u20136.3 mm long vs. 6.2\u20137.8 mm long in F.major)  . Weaver F.major and F.gardenii sensu n = 5x = 60) and named the nothospecies F.\u00d7intermedia Ranney & Fantz, a finding that cleared up previous controversy as to whether common cultivars (such as \u2018Mount Airy\u2019) represented F.major or F.gardenii. No pentaploids have been identified in nature.Although Fothergilla was known. However, recent sampling and cytometric analysis identified a number of diploid populations in Alabama, Florida, Georgia, and South Carolina (F.gardenii and F.major and their relationship to the diploid populations (F.gardenii and F.major). Furthermore, the diploid OTUs segregated into two, well-supported lineages, corresponding to largely allopatric populations. A re-examination of the morphology of diploid plants, in combination with the genetic evidence, has led us here to the recognition of two species of diploids in the genus: a resurrected F.parvifolia and a new species (F.milleri) as described below. A revised taxonomic treatment of the genus is provided.Until recently, no diploid taxon of Carolina . This woulations . These aFothergilla from throughout the southeastern United States, planted and grown in a common garden at the Mountain Horticultural Crops Research and Extension Center in Mills River, North Carolina (Table PSC) sensu IW), and (2) the length of the midvein interval between the junction of the midvein and lowermost secondary vein and the junction of the midvein and the next-most distal secondary vein on the same side of the leaf . We also searched the SERNEC portal  to identify any additional specimens of the diploid taxa we recognized. This search resulted in six additional specimens, which we added to the list of exsiccatae in the taxonomic treatment below . The combined sets of specimens of known ploidy (Table Specimens studied in the course of preparing this revision included: 1) 34 accessions of  34 accesPopulation Aggregation Analysis revealed four distinct aggregate profiles, each corresponding to one of the major lineages identified by F.parvifolia Kearney , the shape is actually a combination of a short cuneate section adjoining the petiole that broadens out laterally into the more general cordate or rounded shape. We here use the terms V-cordate and V-rounded to describe this type of base (the \u201cV\u201d representing the cuneate section) and consider it structurally distinct from the neatly cordate leaf bases of F.parvifolia , F.major (6x), F.milleri (2x), and F.parvifolia (2x). An updated taxonomic treatment follows.Based on these results, Taxon classificationPlantaeSaxifragalesHamamelidaceaeL., Syst. Veg. ed. 13. 418. 1774F9AB60B8-9179-5DF2-813D-AEFB92A8716DFothergillagardenii L., Syst. Veg. ed. 13. 418. 1774 [as F.Gardeni]  \u00d7 0.9\u20132.7(\u20134.3) mm; petioles 3.9\u201310.5 mm long, usually \u00be the length of the IL or longer, brown-yellow pubescent; blades mostly spreading, green, narrowly ovate to ovate, (1.8\u2013)2.8\u20138.7 \u00d7 (1.0\u2013)2.5\u20135.3 cm, pinnately 8\u201311-veined, bases asymmetrical or symmetrical, usually rounded to cuneate, sometimes shallowly cordate, margins crenate to serrate above middle, teeth 3\u201310, apices acute to obtuse, both surfaces stellate-pubescent, rarely glabrous, abaxial surface sometimes glaucous, IW:IL < or = 0.49 (x\u0304 = 0.33). Inflorescences appearing before leaves, spikes terminal, appearing lateral on short lower branches, sessile or on short peduncles. Flowers: stamens 10\u201324, filaments 3.6\u201313.8 mm long. Capsules 6.6\u20139.0 \u00d7 5.5\u20136.6 mm. Seeds white to mottled brown or red-brown, ellipsoid to slightly ovoid, 5.1\u20135.8 \u00d7 2.5\u20133.5 mm, apices mostly obtuse. Genome size and ploidy 3.33\u20133.76 pg, tetraploid (2n = 4x = 48).Flowering beginning late Mar; fruiting by late Apr through Jul.Acerrubrum L., Amelanchierobovalis (Michx.) Ashe, Aroniaarbutifolia (L.) Pers., Clethraalnifolia L., Cyrillaracemiflora L., Gaylussaciadumosa (Andrews) Torr. & A. Gray, G.tomentosa (A. Gray) Pursh ex Small, Ilexcoriacea (Pursh) Chapm., I.glabra (L.) A. Gray, I.vomitoria Aiton, Kalmiacuneata Michx., K.latifolia L., Liquidambarstyraciflua L., Lyonia Nutt. spp., Magnoliavirginiana L., Perseapalustris (Raf.) Sarg., Pinusserotina Michx., Pteridiumaquilinum (L.) Kuhn, Quercuslaevis Walter, Q.virginiana Mill., Rhododendronviscosum (L.) Torr., Vacciniumcrassifolium Andrews, V.myrsinites Lam., and Zenobiapulverulenta (W. Bartram ex Willd.) Pollard (fide colectoris).This species can be found along the Atlantic coastal plains of North Carolina, South Carolina, and Georgia Fig. . It occuFothergillagardenii was apparently cultivated in England as early as 1765, grown at Kew Gardens by 1789, and English and French plant nurseries were offering seeds for sale by the early 1800s  (mblebees . The autrphidae) .Fothergillagardenii s.l. is considered Vulnerable, currently ranked by NatureServe as follows: G3G4; Alabama (S1), Florida (S1), Georgia (S2), Mississippi (SNR), North Carolina (S3S4), South Carolina (SNR) . In light of the removal of the diploid taxa F.milleri and F.parvifolia from the broader historical concept of F.gardenii, the conservation status of this species, as well as that of the diploid taxa, warrants re-evaluation; such re-evaluation is likely to result in a more imperiled ranking than the G3G4 current assessment, following removal of a significant number of populations and a narrower distribution following its narrower taxonomic circumscription. Due to its sensitive status, we provide only skeletal collections data below..Lynch 43 (NCSC); 2012-078, 13 Jun 2014 (V), Phillips 49 (NCSCm); 12 Jul 1997 (V), Sorrie 9349 (NCUm). McDuffie: Jun 1911 (V), Bartlett 2636 .Georgia. Effingham: 2012-078, 15 Apr 2014 (FL), Coker s.n. (NCU); 14 May 1966 (FL), Blair 415 (NCSC); 21 Jun 1965 (FR), Sawyer 2475 (NCUm); 5 Nov 1956 (V), Ahles & Leisner 21471 (NCUm); 20 Apr 1957 (FL), Ahles & Ramseur 23593 (NCUm); 31 May 2003 (FR), Horn 4437 (BRITm); 25 Jan 1937 (FL), Melvin 3613 (BRITm); 7 Jul 1994 (V), Nifong 400 (NCUm). Brunswick: 8 Jun 1951 (FR), Boyce & Wells 1656 (NCSC); 13 May 1950 (FL), Godfrey & Wiebe 50341 ; 18 Apr 1999 (FL), Hill 31327 (BRIT); 6 Mar 1974 (V), Kologiski 54 (NCSCm); 7 May 1974 (FL), Kologiski 125 (NCSC); 23 Jul 1974 (V), Kologiski 311 (NCSC); 6 Jun 1975 (FR), Kologiski 423 (NCSC); 27 Apr 1976 (FL), Kologiski 551 (NCSC); 18 Apr 1965 (FL), Mullen s.n. (NCSC). Carteret: 2011-096, 26 Mar 2012 (FL), Lynch 9 (NCSC); 2011-096, 2 Jul 2012 (V), Lynch 76 (NCSCm); 2011-096, 13 Jun 2014 (FR), Phillips 50 (NCSC); 2011-103, 21 Mar 2012 (FL), Lynch 3 (NCSC); 2011-103, 2 Jul 2012 (V), Lynch 75 (NCSCm); 22 May 1976 (V), Wilson 1792 (NCUm); 9 Apr 1977 (FL), Wilson 3067 (NCU). Columbus: 25 Apr 1958 (FL), Bell 11417 (NCU). Craven: 19 Apr 1958 (Fl), Radford 31924 (NCU). Cumberland: 11 Oct 1957 (V), Ahles 36621 (NCUm); 28 Apr 1933 (FL), Tallin & Harbison s.n. (NCU). Duplin: 27 Apr 1957 (V), Ahles & Ramseur 23990 (VDBm); 27 Apr 1957 (FR), Radford & Ramseur 23990 (NCUm). Harnett: 10 Apr 1957 (FL), Laing 843 (NCU); 30 Jun 2005 (FR), Sorrie 11634 (NCUm). Hoke: 2011-097, 21 Mar 2012 (FL), Lynch 4 (NCSC); 2011-097, 2 Jul 2012, (V), Lynch 72 (NCSCm); 2011-097, 13 Jun 2014 (FR), Phillips 71 (NCSC); 16 May 1976 (V), Kral 58099 (VDB m); 26 Jun 1975 (FR), Ahles & Haesloop 29627 (NCUm). Lee: 19 Apr 1958 (FL), Stewart 149 (NCU); 7 Jun 1958 (FR), Stewart 451 (NCUm). Montgomery: 9 Oct 1956 (V), Radford 19633 (NCUm). Moore: 17 Jul 1942 (FR), Wicker s.n. (NCUm); 8 Apr 1973 (FL), Carter 449 (NCU); 24 Apr 1949 (FL), Woods 2256 (NCSC). New Hanover: 6 Jun 1929 (FR), Wells s.n. (NCSCm); 29 Jun 1963 (V), McCrary 607 (NCU m); 30 Mar 1991 (FL), Pyne & Seneca 91-013 (NCSC). Onslow: 11 May 1948 (FR), Boyce & Moreland 647 (NCSC); 28 Apr 1951 (FL), Beaman s.n. (NCSC); 2 June 1948 (FR), Boyce & Moreland 700 (NCSCm); 24 Jun 1965 (FR), Wilbur 8398 (BRITm). Pamlico: 12 Oct 1957 (V), Radford 42285 (NCUm). Pender: 17 May 1925 (FR), A.C.W. s.n. (NCSC); 1 Jun 1945 (V), Wells s.n. (NCSC); 19 Apr 1957 (FL), Ahles & Ramseur 23440 (NCU); 25 Apr 1947 (FL), Fox & Wells 162 (NCSC); 12 May 1951 (FR), Fox 4621 (NCSC); 26 Aug 1983 (V), Leonard 8199 (UWFPm); 14 Apr 1925 (FL), Wells s.n. (NCSC). Richmond: 2011-123, 11 Apr 2012 (FL), Lynch 13 (NCSC); 2011-123, 21 Aug 2012 (V), Lynch 74 (NCSCm); 2011-085, 21 Mar 2012 (FL), Lynch 5 (NCSC); 2011-085, 2 Jul 2012 (V), Lynch 73 (NCSC); 2011-085, 13 Jun 2014 (V), Phillips 55 (NCSCm); 19 May 2007 (FL), Boyle 1 (NCU). Robeson: 18 Apr 1956 (FL), Terrell 3019 (NCU). Sampson: 10 Apr 1938 (FL), Godfrey 3394 (NCSC). Scotland: 20 Apr 1999 (FL), Hill 31345 (BRITm); 20 Jun 1957 (FR), Ahles & Haesloop 28637 (NCU); 20 Jun 1957 (FR), Ahles & Haesloop 28601 (NCUm); 4 Jun 2004 (FR), Sorrie 11264 (NCUm).North Carolina. Beaufort: 27 Mar 1949 (FL), Duncan 5923 (NCSC). Charleston: 2012-075, 15 Apr 2014 (FL), Lynch 42 (NCSC); 2012-075, 13 Jun 2014 , Phillips 48 (NCSC m); 2012-77, 15 Apr 2014 (FL), Lynch 37 (NCSC); 2012-077, 21 Aug 2012 (V), Lynch 30 (NCSCm); 2012-77, 13 Jun 2014 (FR), Phillips 53 (NCSC); 2 Apr 1944 (FL), Duncan 5885 (NCSC). Chesterfield: 25 Apr 1968 (FR), Ewel 666 (NCSC); 5 Jun 1956 (V), Radford 12431 (NCUm); 5 Apr 1968 (FL), Leonard & Radford 1219 (NCU). Clarendon: 20 Apr 1957 (FL), Radford 21097 (NCU). Colleton: 5 Apr 1956 (FL), Bell 1862 (NCU); 17 Apr 1974 (FL), Hardin 13420 . Darlington: 25 Mar 1935 (Fl), Matthews s.n. (NCU); 26 Mar 1935 (FL), Matthews & Smith s.n. (NCU); 10 Apr 1940 (FL), Smith 1378 (NCU). Dillon: 7 Apr 1940 (FL), Radford & Stewart 56 (NCU). Dorchester: 20 Jul 1957 (V), Ahles & Leisner 31966 (NCUm). Georgetown: 18 Apr 1987 (FL), Taggart 78 (NCU); Near Georgetown, 28 Jul 1928 (FR), Ashe s.n. (NCUm). Horry: 2012-076, 15 Apr 2014 (FL), Lynch 41 (NCSC); 2012-076, 13 Jun 2014 , Phillips 51 (NCSC m). Lee: 26 Jul 1957 (V), Radford 27396 (NCUm). Marlboro: 4 May 1968 (FL), Leonard & Radford 1218 (NCU).South Carolina. Berkeley: 9 Apr 1944 (FR), Taxon classificationPlantaeSaxifragalesHamamelidaceae2.Lodd., Bot. Cab. 16: Pl. 1520. 18298DC644F4-B01B-567E-B9EB-DF10E90AE717F.monticolaW.W. Ashe 1509  Ashe, Bot. Gaz. 24: 374. 1897. Type. North Carolina, mountains, C. Loddiges Illustration Pl. 1520. Figs Shrub, erect, robust, frequently >1 m tall (to 8 m); stems in clumps of 3 or more, branching. Leaves: stipules ovate to lanceolate, 3.5\u201311.8 \u00d7 1.9\u20134.0 mm; petioles 8.3\u201317.1(\u201318.9) mm, \u00bd as long or longer than the IL, brown-yellow pubescent; blades mostly spreading, green, broadly ovate or elliptic to suborbiculate, rarely obovate, 3.6\u201313.7 \u00d7 3.2\u201313.0 cm, most within the upper end of those ranges, pinnately 7\u201312-veined, bases oblique, occasionally symmetrical, usually V-cordate, rarely nearly rounded, margins crenate or serrate to nearly entire, toothed from at or below middle to the apex, teeth 11\u201324, apices acute to obtuse, often pubescent, both surfaces glabrous to sparsely stellate-pubescent, abaxial surface sometimes glaucous, IW:IL > or = 0.51 (x\u0304 = 0.96). Inflorescences usually appearing with leaves, spikes sessile or on short peduncles. Flowers: stamens 14\u201332, filaments 7.3\u201316.2 mm long. Capsules 6.8\u201312.7 \u00d7 6.5\u20138.6 mm. Seeds nearly white to completely brown or red-brown, usually ellipsoid, 5.3\u20137.4 \u00d7 2.7\u20133.6 mm, apices rounded to obtuse. Genome size and ploidy 5.21\u20135.25 pg, hexaploid (2n = 6x = 72).Flowering beginning late Mar; fruiting late May through mid-Jul.F.major can be found growing in seepage community with Coastal Plain-affinity species. In this habitat, where burning is frequent, this species may be <1 m in height. However, height variance is apparently not simply a factor of fire frequency. Fothergillamajor varies considerably in habit, ranging from colonial shrubs less than 2 m tall, typically found growing on sandy dampish sites in the southwestern part of the range and lower elevations  to larger tree forms growing to 6 m, with offsets but no rhizomes, typically found in more upland mountain dry forest sites with igneous geology found to the North and East . Commonly associated upland species include Acerrubrum, Aesculussylvatica W. Bartram, Alnusserrulata, Aroniaarbutifolia (Aiton) Willd., Calycanthusfloridus L., Carpinuscaroliniana Walter, Cornusflorida L., Gaylussaciafrondosa (L.) Torr. & A. Gray, Hamamalisvirginiana L., Hepaticaamericana (DC.) Ker Gawl, Hypericumnudiflorum Michx. ex Willd., Ilexopaca Aiton, Kalmialatifolia, Liquidambarstyraciflua, Liriodendrontulipifera L., Magnoliaacuminata (L.) L., Polystichumacrostichoides (Michx.) Schott, Quercusalba L., Q.rubra L., Rhododendroncatawbiense Michx., and Stewartiaovata (Cav.) Weath. (fide colectoris).This species can be found in the mountains and Piedmont of Alabama, Arkansas, Georgia, North Carolina, South Carolina, and Tennessee Fig. . HabitatFothergillamajor is considered Vulnerable rangewide, ranked by NatureServe as follows: G3; Alabama (S2), Arkansas (S1), Georgia (S1), North Carolina (S3), South Carolina (S2), Tennessee (S2); http://explorer.natureserve.org/, accessed 11 Dec 2019). Due to its sensitive status, we provide only skeletal collections data below..Lynch 91 (NCSCm). Cherokee: 12 Apr 1969 (FL), Kral 34269 ; 10 May 1970 (FL), Kral 39052 (BRITm); 14 Jul 1966 (FR), Clark & Landers 5122 . Cullman: 17 Apr 1931 (FL), Ashe s.n. (NCU); 16 Apr 1924 (FR), Wolf s.n. (AUAm); 4 Jul 1911 (FR), Wolf 813 (AUAm); 28 May 1931 (V), Wolf s.n. (VDB). DeKalb: 2008-009, 21 Mar 2012 (FL), Lynch 7 (NCSC); 2008-009, 2 Jul 2012 (V), Lynch 81 (NCSCm); 14 Jul 1966 (FR), Clark & Landers 5023 ; 22 May 1972 (FL), Whetstone 1935 (NCUm). Marshall: 2011-092, s.d. (FL), Lynch 79 (NCSC); 2011-092, 2 Jul 2012 (V), Lynch 17 (NCSCm); 2011-147, 21 Aug 2012 (V), Lynch 24 (NCSCm); 15 Apr 1973 (FL), Kral 49636 . St. Clair: 31 May 1947 , McVaugh 8594 (BRIT m): Unspecified: n.d. (FL), Buckley s.n. (FLAS).Alabama. Blount: 2011-093, 2 Jul 2012 (V), Lynch 10 (NCSC); 2011-082, 2 Jul 2012 (FR), Lynch 80 (NCSCm).Arkansas. Searcy: 2011-082, 26 Mar 2012 (FL), Duncan & McDowell 12200 (NCU); 30 Mar 1951 (FL), Duncan & McDowell 12200 ; 6 May 1951 (FL), Duncan & Venard 12339 (BRIT m). Fulton: 2011-169, 21 Aug 2012 (V), Lynch 28 (NCSC). Lumpkin: 2011-164, 21 Aug 2012 (V), Lynch 27 (NCSCm). Walker: 2011-146, 21 Aug 2012 (V), Lynch 25 (NCSC); 15 Apr 1986 (FL), Coile 4547 (NCSC).Georgia. Bartow: 30 Mar 1951 (FL), Gavalak 3306V90 (BRIT m).Illinois. DuPage: Cultivated at Morton Arboretum, 9 May 1990 (FL), Dewolf & Bruns 2235 (BRIT).Massachusetts. Suffolk: Cultivated at Arnold Arboretum, Harvard University, 13 May 1968 (FL), Gillis 15041 (BRIT m).Michigan. Ingham: Cultivated at Michigan State University, 9 May 1979 (FL), Lynch 81 (NCSCm); 7 Oct 1966 (V), Downs 408 (NCSC); 2 Aug 1977 (FR), Kral 60712 (VDBm); 25 Aug 1989 (FR), Lance & Wood s.n. (NCUm); 1 Sep 1952 (FR), Radford 6676 (NCUm); 9 Sep 1976 (V), Smith 182 (NCSC); 5 Jul 1940 (FR), Stewart 1554 (NCU); 27 May 1964 (FR), Wilbur 7012 (VDBm). Chatham: 25 Apr 1988 (FL), Swab 75 (NCSC); 4 May 1988 (FL), Swab 97 (NCSC); 28 May 1988 (FR), Swab 236 (NCSC). Gaston: 19 May 1919 (FR), Coker s.n. (NCUm). Harnett: 18 Apr 2006 (V), Sorrie & Hart 11770 (NCUm). Montgomery: 2011-122, 21 Aug 2012 (V), Lynch 23 (NCSCm); 11 Oct 2002 (FR), Diamond 1606 (NCUm); 14 Jul 2004 (V), Schwartzman 30 (NCUm); 20 Jul 2004 (V), Weakley s.n. (NCUm); 15 Oct 2004 (V), Weakley s.n. (NCU). Orange: 2011-124, 19 Apr 2012 (FL), Lynch 14 (NCSC); 2011-124, 2 Jul 2012, Lynch 78 (NCSCm); 2011-124, 24 Jun 2014 (FR), Phillips 62 (NCSC); Apr 1899 (FL), Ashe s.n. (NCU); Jun 1899 (FR), Ashe s.n. (NCU); 31 Apr 200 (FL), Wally & Wichmann 125 (NCUm); 4 May 1910 (FR), Clerces s.n. (NCU). Person: 7 Jul 2005 (FR), LeGrand s.n. (NCUm). Polk: 30 May 1930 (FL), Ashe s.n. (NCSC); 20 Apr 1897 (FL), Biltmore 6565 (BRITm); 20 Apr 1897 (FL), Biltmore 708 (NCU). Randolph: 22 Apr 1958 (FL), Melvin s.n. (NCU). Rutherford: 2011-163, 21 Aug 2012 (V), Lynch 26 (NCSCm); 2011-163, 15 Apr 2013 (FL), Lynch 39 (NCSC); 2011-163, 24 Jun 2014 (FR), Phillips 64 (NCSC); 21 Apr 1956 (FL), Bell 2118 (NCU). Stokes: 23 Apr 1950 (FL), Fox et al. 3565 (NCSC); s.d. (FR), Harbison s.n. (NCU); 5 May 1936 (FL), Hunt 13474 ; 7 Jul 1969 (FR), Leonard & Russ 2553 (NCUm); 21 Apr 1974 (FL), Massey 3900 (VDB); 26 Apr 1958 (FL), Matthews 54 (BRIT); 15 Jun 1967 (FR), Radford 45392 ; 4 Jun 1958 (FR), Radford 34675 (VDB m); 9 Apr 1933 (FL), Schallert 3613 (BRIT). Surry: 19 May 1935 (V), Harbison s.n. (NCU). Transylvania: 2011-112, 2 Jul 2012 (V), Lynch 16 (NCSC); 2011-112, 24 Apr 2014 (FL), Lynch 47 (NCSC); 2011-112, 24 Jun 2014 (FR), Phillips 63 (NCSC); 2011-131, 21 Aug 2012 (FR), Lynch 22 (NCSCm); 19 Jun 1965 (FR), Eggers 1262 (VDB m); 26 Jul 1962 (FR), Rodgers 62064a (NCUm); 24 May 2006 (FR), Schwartzman 29 (NCUm). Wake: 28 Oct 2005 (V), Bruhn s.n. (NCU).North Carolina. Burke: 2011-105, 2 Jul 2012 (V), Fogg 1 (UWFP).Pennsylvania. Marion: Cultivated at the Arboretum of the Barnes Foundation, 27 Apr 1968 (FL), Kral 58130 (VDB m); 15 Oct 1988 (V), Hill 20066 (BRIT m); 23 May 1996 (V), Townsend 874 (VDB m). Oconee: 2011-091, 28 Mar 2012 (FL), Lynch 12 (NCSC); 2011-091, 2 Jul 2012 (V), Lynch 77 (NCSCm); 26 Apr 1965 (FL), Radford 44707 (NCU); 20 Apr 1973 (FL), Rogers & Green 73078 (FLAS-2). Pickens: 15 June 1992 (FT), Hill 23387 (BRIT m);South Carolina. Greenville: 17 May 1976 (FR), Sharp 556 ; 30 Apr 1931 (FL), Jennison & Sharp s.n. (NCU); May 3 1936 (Fl), Sharp & Underwood 4205 . Greene: 17 May 1970 (FL), Sharp et al. 45204 . Scott: 2012-065, 21 Aug 2012 (V), Lynch 29 (NCSC m); 2012-065, 15 Apr 2014 (FL), Lynch 44 (NCSC); 23 Apr 1972 (FL), Clebsch s.n. (VDB); 19 Apr 1979 (FL), Whitten & Noss s.n. (FLAS-2); 23 Jun 1978 (FR), Wofford et al. 78-112 (VDB m). Sevier: 1 May 1960 (Fl), Sharp 26818 ; 25 Apr 1970 (FL), Williams 82 (AUA m); 25 Apr 1970 (FL), Pippin 83 (VDB); 2 Apr 1938 (FL), Jennison 53 (VDB m); 26 Apr 1957 (FL), Sharp 21575 (BRIT m).Tennessee. Grainger: 20 Apr 1931 (FL), Meyer 37144 (FLAS).Washington D.C. U.S. National Arboretum, 16 Oct 1990 (V), Taxon classificationPlantaeSaxifragalesHamamelidaceae3.W.D.Phillips & J.E.Haynessp. nov.DC5E3CB6-8D0A-57AA-96CC-7D6838D9DC9Furn:lsid:ipni.org:names:77208304-1Nathan Lynch 70 . Figs Florida. Walton Co.: Voucher specimen from containerized plant at Mountain Horticultural Crops Research and Extension Center, Mills River, NC (leaves and stem collected 19/7/2012), , 2011-088, 19 Jul 2012 (V), Fothergillamilleri is morphologically most similar to F.gardenii, but differs from the latter by leaves held erect (vs. spreading in F.gardenii), blades blue-green or gray-green (vs. green in F.gardenii), petioles 1/3\u20131/2 the length of the IL (vs. \u00be the length of the IL or longer in F.gardenii), and seed apices acute to acuminate (vs. rounded or obtuse in F.gardenii).Shrub, rhizomatous, erect, to 1.5 m tall, clump-forming, multi-stemmed, branching. Leaves: stipules ovate to lanceolate, 2.5\u20137.8 \u00d7 1.0\u20133.0 mm; petioles 2.6\u20138.0 mm long, usually 1/3\u20131/2 the length of the IL, brown-yellow pubescent; blades erect, blue-green to gray-green, obovate, 3.2\u20138.0 \u00d7 3.0\u20134.8 cm, pinnately 8\u201310-veined, bases rounded to truncate, margins crenate to serrate above middle and mostly near the apex, sometimes appearing crenate to entire, apices mostly obtuse to acute, both surfaces conspicuously stellate-pubescent, sometimes sparsely so, abaxial surface not glaucous, IW:IL < or = 0.48 (x\u0304 = 0.29). Inflorescences appearing before leaves, spikes on short peduncles or sessile. Flowers: stamens 15\u201322, filaments 4.3\u201310 mm long. Capsules 7.0\u20139.0 \u00d7 5.0 \u20137.0 cm. Seeds usually completely brown to red-brown, ovoid, 4.5\u20136.2 \u00d7 2.4\u20133.2 mm, apices acuminate, often recurved. Genome size and ploidy: 1.70\u20131.78 pg, diploid (2n = 2x = 24).Flowering beginning late Mar; fruiting late May through the end of Jul.Cyrillaracemiflora or Taxodiumascendens swamp forests. Plant associates include Acerrubrum, Arundinariatecta  Muhl., Clethraalnifolia, Hibiscusaculeatus Walter, Hypericumcistifolium Lam., Juncustrigonocarpus Steud., Dichantheliumscoparium (Lam.) Gould., Osmundastrumcinnamomeum (L.) C. Presl, Rhexiavirginica L., Rhynchosporachalarocephala Fernald & Gale, and Xyrisfimbriata Elliott (fide colectoris).This species can be found in the panhandle of Florida, in Alabama in the Gulf coastal plain, and is also known from one county in Georgia Fig. . This spThe species is named in honor of Dr Ron Miller, Pensacola, Florida, who championed this project, originally suggested that diploid cytotypes might still exist, and ultimately found them. Dr Miller\u2019s extensive effort and field work (with colleagues) provided the foundation for this research and establishment of ex situ living collections, including accessions in the U.S. National Plant Germplasm System.F.major. Consequently, only skeletal collections data are provided below.The conservation status of this species must be reassessed. It is presently known from only seven counties, and would appear to have an imperilment status at least as severe as that of .Lynch 1 (NCSC); 2011-087, 2 Jul 2012 (V), Lynch 68 (NCSCm); 2011-087, 13 Jun 2014 (FR), Phillips 58 (NCSC); 16 Jun 1984 (FR), Hedges 156 (UWFPm); 19 Apr 2001 (FR), Schotz 1830 (UNA [online!]). Covington: 25 Jun 1906 (FR), Harper 108 (NY [online!]). Escambia: 11 Apr 1964 (FL), Kral 19698 (AUAm). Geneva: 25 Jul 1968 (V), Kral 32029 (VDBm); 26 Apr 1969 (FL), Kral 34524 (VDBm); 7 Sep 1994 (V), Simmers s.n. (HTTU [online!]). Monroe: 15 Jun 1959 (V), Grelen s.n. (FLASm).Alabama. Baldwin: 2011-087, 21 Mar 2012 (FL), Lynch 2 (NCSC); 2011-083, 2 Jul 2012 (FL), Lynch 67 (NCSC); 2011-083, 13 Jun 2014 (FR), Phillips 57 (NCSCm); 20 Apr 1991 (FL), Burkhalter 12638 ; 5 Aug 1990 (V), Burkhalter 12211 (UWFPm); 13 Jul 1996 (V), Burkhalter 15064 (UWFP); 18 May 1993 (FR), Naczi 3028 (KNK [online!]). Walton: 8 Apr 1899 (FL), Biltmore Herbarium 7609b (FLASm); 2011-088, 28 Mar 2012 (FL), Lynch 11 (NCSU); 2012-060, 19 Mar 2013 (FL), Lynch 34 (NCSC); 2012-060, 2 Jul 2012 (V), Lynch 15 (NCSC); 2012-060, 13 Jun 2014 (FR) Phillips 59 (NCSC); 18 Aug 1990 (V), Orzell and Bridges 14757 .Florida. Okaloosa: 2011-083, 21 Mar 2012 (FL), Lynch 8 (NCSC); 2011-178, 2 Jul 2012 (V), Lynch 69 (NCSCm).Georgia. Taylor: 2011-178, 26 Mar 2012 (FL), Taxon classificationPlantaeSaxifragalesHamamelidaceae4.Kearney in Small, Fl. S.E. U.S. 509, 1331. 1903988E10E7-FD07-532B-8B40-80EC41C51904Kearney s.n. . Figs Georgia. Wayne Co., Jesup, dry soil, 4 June 1893 (FR), Shrub, rhizomatous, erect, to 1 m tall, clump-forming, multi-stemmed, branching. Leaves: stipules ovate to lanceolate, 3.5\u20136.0 \u00d7 8.0\u201313.0 mm, curved downward; petioles 4.4\u201317.8 mm long, usually nearly as long to longer than the IL, brown-yellow pubescent; blades drooping, green, mostly ovate, 3.2\u201312.1 \u00d7 3.0\u20136.2 cm, pinnately 8\u201310-veined, bases cordate, margins crenate to serrate from middle, apices acute, adaxial and abaxial surfaces not glaucous, stellate-pubescent, sometimes sparsely so, IW:IL > or = 0.62 (x\u0304 = 0.96). Inflorescences appearing before leaves, spikes on short peduncles or sessile. Flowers: stamens 15\u201320 per flower, filaments 6.6\u20139.3 mm long. Capsules 7.5\u201310.0 \u00d7 5.0 \u20137.6 mm. Seeds usually completely brown to red-brown, ovoid, apex obtuse, 4.5\u20136.2 \u00d7 2.4\u20133.2 mm, apices obtuse to acute. Genome size and ploidy 1.73\u20131.82 pg, diploid (2n = 2x = 24).Flowering Mar\u2013May; fruiting May\u2013Sep.This species range is restricted to Georgia, South Carolina, and Alabama Fig. . BecauseF.major. Consequently, only skeletal collections data are provided below.The conservation status of this species needs to be assessed. It is presently known from only eight counties, and would appear to have an imperilment status at least as severe as that of .Harbison 1033 (NCUm).Alabama. Montgomery: 12 Sep 1899 (FR), Cuthbert s.n. (FLAS). Brantley: 23 Aug 1947 (V), Thorne & Norris 6284 (GEO [online!]). Emanuel: 2011-170, 2 Jul 2012 (V), Lynch 19 (NCSCm); 2011-170, 15 Mar 2013 (FL), Lynch 33 (NCSC); 2011-170, 14 Jun 2014 (FR), Phillips 60 (NCSC). Long: 2011-171, 2 Jul 2012 (V), Lynch 20 (NCSCm); 2011-171, 14 Mar 2013 (FL), Lynch 32 (NCSC); 2011-171, 13 Jun 2014 (FR), Phillips 61 (NCSC); Tattnall: 2011-168, 19 Mar 2013 (FL), Lynch 35 (NCSC); 2011-168, 2 Jul 2012 (V), Lynch 18 (NCSCm). Wayne: 31 Aug 1904 (V), Biltmore Herbarium 14940 (NY [online!]).Georgia. Augusta-Richmond: 2 Apr 1898 (FL), Lynch 40 (NCSC); 2012-084, 13 Jun 2014 (V), Phillips 56 (NCSCm). Lexington: 13 Sept 1988 (V), Pittman 9139817 (BRITm); 27 May 1957 (FR), Radford 23378 (NCUm).South Carolina. Aiken: 2012-084, 15 Apr 2014 (FL),"}
+{"text": "Numerous individual studies have investigated the diagnostic value of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG detection for nasopharyngeal carcinoma (NPC), but the conclusions remain controversial. This meta-analysis aimed to determine the value of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG detection in the diagnosis of NPC.PROSPERO registration number: CRD42019145532. PubMed, EMBASE, Cochrane Library, and Chinese data libraries  were searched up to January 2019. The pooled sensitivity, specificity, and positive likelihood, negative likelihood, and diagnostic odds ratios were conducted in this meta-analysis. Summary receiver operating characteristic curves evaluated the test-performance global summary. Publication bias was examined by Deek\u2019s funnel plot asymmetry test.EBV-DNA in diagnosis of NPC were: 0.76 (95% CI 0.73\u20130.77), 0.96 (95% CI 0.95\u20130.97), 14.66 (95% CI 9.97\u201321.55), 0.19 (95% CI 0.13\u20130.28), 84 (95% CI 50.45\u2013139.88), 0.96 (SE: 0.001), and 0.55 (95% CI 0.54\u20130.57), 0.96 (95% CI 0.96\u20130.97), 12.91 (95% CI 9.55\u201317.45), 0.35 (95% CI 0.29\u20130.43), 39.57 (95% CI 26.44\u201359.23), 0.94 (SE: 0.002) for the EA-IgA, and 0.85 (95% CI 0.84\u20130.85), 0.89 (95% CI 0.88\u20130.89), 6.73 (95% CI5.38\u20138.43), 0.17 (95% CI 0.12\u20130.23), 43.03 (95% CI 31.51\u201358.76), 0.93 (SE: 0.007) for the VCA-IgA, and 0.86 (95% CI 0.85\u20130.88), 0.87 (95% CI 0.88\u20130.90), 7.55 (95% CI 5.79\u20139.87), 0.16 (95% CI 0.13\u20130.19), 50.95 (95% CI 34.35\u201375.57), 0.94 (SE: 0.008) for the EBNA1-IgA, and 0.70 (95% CI 0.69\u20130.71), 0.94 (95% CI 0.94\u20130.95), 9.84 (95% CI 8.40\u201311.54), 0.25 (95% CI 0.21\u20130.31), 40.59 (95% CI 32.09\u201351.35), 0.95 (SE: 0.005) for the Rta-IgG. The EBV-DNA had larger AUC compared with other EBV-based antibodies (P\u2009<\u20090.05), while the difference between EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG was not statistically significant (P\u2009>\u20090.05).Forty-seven studies with 8382 NPC patients (NPC group) and 15,089 individuals without NPC (Control group) were included in this meta-analysis. The sensitivity, specificity, positive likelihood (+\u2009LR), negative likelihood (-LR), DOR and AUC of EBV-DNA, VCA-IgA, EBNA1-IgA and Rta-IgG detection have high accuracy in early diagnosis NPC. In addition, EBV-DNA detection has the higher diagnosis accuracy in NPC. On the other hand, EA-IgA is suitable for the diagnosis but not NPC screening.The online version contains supplementary material available at 10.1186/s12935-021-01862-7. Nasopharyngeal carcinoma (NPC) is the most common malignant tumor in head and neck surgery and it is highly prevalent in southern China and Southeast Asia . UnfortuPROSPERO registration number: CRD42019145532.Literature review was separately conducted by two investigators that queried online databases, including PubMed, EMBASE, Cochrane Library, and Chinese data libraries , and the search concluded in January 2019, using the following keywords: nasopharyngeal carcinoma, Epstein-Barr virus, capsid antigen-IgA, early antigen antibody, nuclear antigen antibody, BRLF1 transcription activator IgG, EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG.Studies that assessed the performance of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG detection for untreated NPC identification;All patients included in the study were diagnosed using a reference test ;Studies that used a pre-specified threshold;Studies that clearly stated the number of true positive, false positive, false negative, and true negative results in the diagnosis of NPC or these values could be calculated from the data;Studies that provided a clear definition of the control sources ;In cases of multiple reports describing the same population, the most recent or most complete report was selected.Reported results were insufficient for construction of the 2\u2009\u00d7\u20092 table;Studies that failed to clearly define the control types;The NPC group contained other tumors;Basic research, review articles, comments, letters, case reports, abstracts in conference, responding letters and experimental animal studies.Study quality assessment was conducted using the diagnostic accuracy (QUADAS) II checklist . StudiesQ test and inconsistency index (I2) were used to estimate the heterogeneity within studies [P\u2009<\u20090.05 or I2\u2009>\u200950%. If statistically significant heterogeneity existed, meta-analysis was performed using the random effects model, otherwise, a fixed effect model was used.Standard methods recommended for meta-analyses of diagnostic test evaluations were used to perform this meta-analysis . Review  studies . HeterogThe accuracy indexes of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG was pooled by meta-analysis, such as sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), diagnostic odds ratio (DOR) and AUC. The likelihood ratios (PLR and NLR) are clinically meaningful for the measurement of diagnostic accuracy; PLR\u2009>\u200910 and NLR\u2009<\u20090.1 are considered high . The DORP values were two sides and P\u2009<\u20090.05 was regarded as statistically significant.Sensitivity analyses were performed to explore the sources of heterogeneity of the included studies by removing each included study consecutively. The heterogeneity was investigated by meta-regression according to different covariates, including publication year (Year\u2009\u2265\u20092011 or\u2009<\u20092011), NPC size (NPC\u2009\u2265\u2009100 or NPC\u2009<\u2009100), control sources , detection method, and article language (English or Chinese). Publication bias was examined by Deek\u2019s funnel plot asymmetry test. All In this meta-analysis, 47 publications on the role of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG concentrations in the diagnosis of NPC that met the criteria for inclusion were included in the analysis \u201358. FiguI2\u2009=\u200977.5%, P\u2009<\u20090.001; EA-IgA:77.3%, P\u2009<\u20090.001; VCA-IgA: 87.0%, P\u2009<\u20090.001; EBNA1-IgA:78.9%, P\u2009<\u20090.001; Rta-IgG: 60.5%, P\u2009<\u20090.001) indicated significant heterogeneity among the studies. The result showed that there was no threshold effect in the pooled analysis of EBV-DNA (P\u2009<\u20090.001), EA-IgA (P\u2009<\u20090.001), VCA-IgA (P\u2009<\u20090.001), EBNA1-IgA (P\u2009<\u20090.001) and Rta-IgG (P\u2009<\u20090.001).The inconsistency index (EBV-DNA: P\u2009<\u20090.05), while EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG were no statistically different from each other (P\u2009>\u20090.05) in Table The pooled sensitivity, specificity, PLR, NLR, DOR and AUC for the value of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG in the diagnosis of NPC are displayed in Table P\u2009<\u20090.0095).The sensitivity analysis showed that the results were not affected by the exclusion of any individual trial. As meta-regression result indicated that publication year, NPC or control size, control sources, detection method, cutoff value, and article language are not the DOR heterogeneity of EBV-DNA, VCA-IgA, EBNA1-IgA and Rta-IgG, whereas detection method was possible DOR heterogeneity sources for the EA-IgA (P\u2009=\u20090.14), EA-IgA (P\u2009=\u20090.26), EBNA1-IgA (P\u2009=\u20090.56) and Rta-IgG (P\u2009=\u20090.16), other than VCA-IgA (P\u2009=\u20090.03)  and 0.89 (95% CI 0.88\u20130.90) [CI 0.90\u20130.92) and specificity 0.92 (95% CI 0.92\u20130.93), with SROC 0.98. But the Li\u2019s study existed language bias and publication bias. For the Rta-IgG, Cui et al.pooled 17 studies involving 2658 NPC patients, and the\u00a0results pointed out that the sensitivity of Rta-IgG for detecting NPC was 90.83 (95% CI 0.78\u20130.87), the specificity was 0.92 (95% CI 0.90\u20130.93) [Previous meta-analyses have been published on the value of some EBV-based markers in the detection of NPC. For the EBV-DNA, Han et al.conducted a meta-analysis based on 18 studies involving 1492 NPC cases and 2461 health controls in Asians, in which the pooled sensitivity and specificity of EBV-DNA detection for NPC were 0.73 (95% 88\u20130.90) . Further88\u20130.90) . The Han88\u20130.90) , which wP\u2009<\u20090.05). Additionally, the pooled DOR of EBV-DNA (84.00) that differed 33.05\u201344.43 was higher than other EBV-based antibodies. These results indicated that EBV-DNA detection had higher accuracy in diagnosis of NPC. In addition, a meta-analysis included 8128 NPC cases showed that pre-EBV-DNA levels can also be a prognostic indictor for patients with NPC [To our knowledge, this is the first meta-analysis to determine the usefulness of EA-IgA and EBNA1-IgA and compare the accuracy EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgA in diagnosis of NPC. In this study,\u00a0the highest sensitivity was EBNA1-IgA (0.86), and the specificity of EBV-DNA (0.96) and EA-IgA (0.96) were highest. Besides, the sensitivity of EA-IgA (0.55) was lowest, and screening for NPC using only EA-IgA may lead to misdiagnosis, but the specificity was high, which indicated that EA-IgA was suitable for the diagnosis but not screening of NPC. EA-IgA or EBV-DNA detection combined with other indicators may also improve the sensitivity and specificity for the serological diagnosis of NPC , 51. Thewith NPC . A recenwith NPC , seventywith NPC . In addiwith NPC , 71. In with NPC , 72, it Several strengths of present meta-analysis should be highlighted. This study compered five EBV-related diagnostic markers for NPC with comprehensive calculations of their diagnostic performance, shedding light on the value of these tests in clinical settings. In the context of current availability of studies on EBV-DNA and immunoglobin antibody tests in the literature, this meta-analysis covers a large sample size pooled from rigorously included studies, and the results were stable. However, this meta-analysis should be interpreted with caution due to certain limitations. First, there was large heterogeneity among the included studies with differences in characteristics of the study and participants. The inability to obtain raw data on patient age and gender may have led to the heterogeneity and hindered a more detailed analysis. Second, most of the included populations were Chinese, which could lead to population selection bias and should not allow for generalization to other ethnicity groups, rendering further research needed. Third, technical methods for testing of EBV-related markers vary across different studies, including the inconsistent cut-off values and different antigen sets used despite of a same generic name. Use of enzyme-linked immunosorbent assay or immunofluorescence assay may have contributed to the different results . FinallyNotwithstanding heterogeneity of currently available data, the studies included in our analysis are of a large sample size and high-quality, thus providing a considerable power. EBV-DNA, VCA-IgA, EBNA1-IgA and Rta-IgG detection have high accuracy in early diagnosis NPC and can improve the effectiveness of screening. In addition, EBV-DNA detection has the higher diagnosis accuracy in NPC. On the other hand, EA-IgA is suitable for the diagnosis but not NPC screening. Further well-designed clinical trials need to be carried out in order to improve early diagnosis rate.Additional file 1: Forest plots of sensitivity, specificity, PLR, NLR, DOR for acoustic analysis of EBV-DNA, EA-IgA, VCA-IgA, EBNA1-IgA and Rta-IgG, and Funnel plots for publication bias test."}
+{"text": "AbstractPhyllodiaptomus (Phyllodiaptomus) roietensissp. nov. was collected from temporary water bodies in Roi Et and Nakhon Ratchasima provinces in northeastern Thailand and Kampong Thom Province in central Cambodia. The new species is closely related to Phyllodiaptomus (P.) surinensis Sanoamuang & Yindee, 2001 in that it shares common morphological characters in the males: urosomites 2\u20133, P5 intercoxal sclerite, right P5 Exp-2, and left P5 Exp. Minor differences on the right antennule, right caudal ramus, P5 basis and Enp exist. The females differ in their Pdg 5, genital double-somite, and P5. An updated key to the species of the genus Phyllodiaptomus Kiefer, 1936 is provided. Phyllodiaptomus Kiefer, 1936, is among the most common freshwater copepods in Southeast Asia (Phyllodiaptomus (Phyllodiaptomus) blanci  from Uzbekistan; P. (Ctenodiaptomus) annae  from Sri Lanka; P. (P.) tunguidus Shen & Tai, 1964 from China; P. (P.) longipes Kiefer, 1965 from Indonesia; P. (C.) sasikumari Ranga Reddy & Venkateswarlu, 1989 and P. (C.) wellekensae Dumont & Ranga Reddy, 1993 from India; P. (C.) praedictus Dumont & Ranga Reddy, 1994, P. (P.) christineae Dumont, Ranga Reddy & Sanoamuang, 1996, P. (P.) surinensis Sanoamuang & Yindee, 2001, and P. (P.) thailandicus Sanoamuang & Teeramaethee, 2006 from Thailand; and P. (P.) irakiensis Khalaf, 2008 from Iraq. In addition, P. (C.) praedictussulawesensis as a subspecies of P. (C) praedictus from Indonesia; this subspecies was later found in the Philippines  roietensis sp. nov. from two localities in Roi Et and Nakhon Ratchasima provinces, northeast Thailand, and two localities in Kampong Thom Province in central Cambodia . After 10 min the animals were transferred to pure glycerol. The animals were dissected and prepared in a glycerin-mounted slide under a stereomicroscope at 40\u2013100-x magnifications. The dissected specimens were mounted in pure glycerin on a glass slide and sealed under a cover glass with transparent nail varnish. All un-dissected specimens were stored in 70% ethanol in 1.5 mL microtubes.All appendages and body ornamentation were examined at 1000-x magnification under an Olympus CX31 compound microscope. The drawings were made using an Olympus U-DA drawing tube mounted on a compound microscope. The final versions of the drawings were made using the CORELDRAW 12.0 graphic program.SEM) were dehydrated in an ethanol series  for 15 min at each concentration. Specimens were dried in a critical-point dryer and were mounted on stubs using adhesive tape under a stereomicroscope. Dried specimens were coated with gold in a sputter-coater. The SEM photographs were taken using a scanning electron microscope (FEI Helios NanoLab G3 CX).Specimens for scanning electron microscopy  and at the Applied Taxonomic Research Center, Khon Kaen University (Thailand) (KKU).Specimens were deposited at the Abbreviations used in this paper are as follows:ae aesthetasc;Enp endopod;Exp exopod;Exp/Enp-n exopodal segment n/endopodal segment n;Pdg pediger;Pdg 1\u20135 pedigers 1\u20135;P1\u2013P5 legs 1\u20135;sp spine.The descriptive terminology follows Diaptominae Kiefer, 1932Sub-family Phyllodiaptomus Kiefer, 1936Genus Phyllodiaptomus Dumont, Ranga Reddy & Sanoamuang, 1996Subgenus Taxon classificationAnimaliaCalanoidaDiaptomidae0B9F97DA-0E9E-5345-88D4-29E2AF212FBEhttp://zoobank.org/59131C6D-A0DE-4BE0-9383-20C0DA8A709D-1.A pool in the rice field at Ban Nakae, Khilek Subdistrict, Pathum Rat District, Roi Et Province, northeastern Thailand; pH of water 8.6, water conductivity 126 \u00b5S cmHolotype: one adult male completely dissected , Ban Nakae , Khilek Subdistrict, Pathum Rat District, Roi Et Province, northeastern Thailand; collected on 12 June 1999 by L. Sanoamuang. Allotype: one adult female completely dissected ; same data as for holotype. Paratypes: two adult females and three adult males undissected preserved in 70% ethanol (NHMUK 2019.9\u201313), one adult female completely dissected (KKU-COP-2019-S-01); one adult female with eggs and three adult males undissected preserved in 70% ethanol (KKU-COP-2019-T-01); same data as for holotype.15\u00b010'55\"N, 102\u00b023'46\"E), Than Lalot Subdistrict, Phimai District, Nakhon Ratchasima Province, northeastern Thailand; collected on 17 October 2017 by N. Plangklang; (2) a roadside canal, Tropeang Chouk village , Baray District, Kampong Thom Province, central Cambodia; collected on 14 June 2007 by R. Chaicharoen; (3) a temporary pond, Kropeu village , Steung Sen District, Kampong Thom Province, central Cambodia; collected on 14 June 2007 by R. Chaicharoen.(1) a temporary pond, Ban Non Lakki , 3+ae (II), 1+ae (III), 1(IV), 1+ae (V), 1 (VI), 1+ae (VII), 1+sp (VIII), 2+ae (IX), 1 (X), 1 (XI), 1+ae+sp (XII), 1 (XIII), 1+ae (XIV), 1 (XV), 1+ae (XVI), 1 (XVII), 1 (XVIII), 1+ae (XIX), 1 (XX), 1 (XXI), 2 (XXII), 2 (XXIII), 2 (XXIV), 4+ae (XXV).Antennule Fig.  symmetriExp-1\u20137 with 1, 3, 1, 1, 1, 1, and 1 inner seta, respectively; Exp-7 with three additional apical setae. Enp-1 with two inner median setae. Enp-2 with eight inner and seven apical setae.Antenna Fig.  biramousEnp-1 with four inner distal setae; Enp-2 with nine apical setae plus tiny spinules along posterior surface. Exp-1\u20134 with 1, 1, 1, 3 setae, respectively.Mandible Fig.  with sixEnp with seven apical setae. Exp with six setae.Maxillule Fig.  with 13 Enp-1 and 2 with three setae each.Maxilla Fig.  with twoEnp-1\u20136 with 2, 3, 2, 2, 2, 4 setae, respectively.Maxilliped Fig.  with fouExp longer than Enp, Exp and Enp three-segmented except P1 Enp bi-segmented. Armature formula of P1\u2013P4 as follows :P1\u2013P4 Fig.  with rouExp-1. Exp-1 sub-rectangular, more than twice as long as wide, longer than Enp. Exp-2 triangular, right side stout and shorter than left one; with row of strong spinules along margins and proximolateral spine at basal Exp-3; with two longitudinal grooves on anterior view , 3+ae (II), 1+ae (III), 1 (IV), 1+ae (V), 1 (VI), 1+ae (VII), 1+sp (VIII), 2+ae (IX), 1+sp (X), 1+sp (XI), 1+ae+sp (XII), 1+ae+sp (XIII), 2+ae+sp (XIV), 2+ae+sp (XV), 2+ae+sp (XVI), 1+sp (XVII), 1+sp (XVIII), 2+ae+sp (XIX), 3+sp (XX), 2 (XXI), 4+ae (XXII); geniculated between segments 18 and 19; segment 20 (antepenultimate) with serrated process distally (3\u20134 teeth), and with longitudinal hyaline membrane along outer margin.Antennule Figs , 7G, H aLeft antennule, antenna, mouthparts, and P1\u2013P4 as in female.Exp-1. Enp  roietensis sp. nov. with the male P5 Exp-2 displays an affinity to the subgenus Phyllodiaptomus sensu Exp-2, medially concave in posterior view, principal lateral spine inserted on distal to mid-outer margin and three accessary spines arranged from proximal, middle, and distal, respectively; the left Exp-2 with patch of strong spinules along medial margin.Exp-1 (4) Exp-2 oval and concave, with strong, flat, curved principal spine and three accessary spines, and (5) bi-lobed Enp. The left P5 with long and narrow hyaline lamella along inner margin, Exp-2 with patch of strong spinules along medial margin, and bi-segmented Enp.The male of the new species has serrated outgrowth on the antepenultimate segment of the right antennule. The right caudal ramus with small chitinous spine near distal margin on ventral side and triangular prominence along proximal one-third length of outer margin. The P5 intercoxal sclerite produced, with convex distal margin. The right P5 with (1) short, strong spine on posterior lobe of coxa, (2) triangular hyaline lamella on proximal inner margin and large chitinous outgrowth on posterior surface of basis, (3) acute distal outer corner of Pdg 5 wings, left wing more elongated in posterio-lateral direction; posterior and dorsal spines short and strong. Genital double-somite with posterolateral directed process on right side. One pair of genital spines on lateral side slightly symmetrical and strong. P5 Exp-2 with conveyor canal on anterior surface. P5 with bi-segmented Enp.Female with asymmetrical roietensis is taken after the type locality, Roi Et Province. The name with the Latin suffix \u201c-ensis\u201d is the adjective for a location.The specific name Dentodiaptomusjavanus , Eodiaptomussanoamuangae Ranga Reddy & Dumont, 1998, Mongolodiaptomuscalcarus , M.malaindosinensis , Neodiaptomuslaii Kiefer, 1974, and Phyllodiaptomus (Phyllodiaptomus) christineae Dumont, Ranga Reddy & Sanoamuang, 1996.Known only from four temporary water bodies from Roi Et and Nakhon Ratchasima provinces, Thailand and Kampong Thom Province, Cambodia Fig. . PresencPhyllodiaptomus has been recorded in Asia, including south China, Turkey, Israel, Uzbekistan, Iran, Iraq, India, Sri Lanka, Nepal, Indonesia, Thailand, Laos, Philippines and Cambodia (P. (C.) annae, P. (C.) wellekensae, and P. (C.) sasikumari) are endemic to India; two species (P. (P.) thailandicus and P. (P.) surinensis) are endemic to Thailand; P. (P.) tunguidus, P. (P.) irakiensis, and P. (P.) longipes are endemic to China, Iraq, and Indonesia, respectively. Only P. (P.) blanci is widely distributed, extending across many countries. Five species have been recorded in Thailand, namely P. (C.) praedictus, P. (P.) christineae, P. (P.) thailandicus, P. (P.) surinensis, and P. (P.) roietensis sp. nov. (P. (P.) surinensis and P. (P.) roietensis sp. nov. are rare. In 3,000 samples collected within Thailand, each has been recorded in only two localities in the northeast. This is in contrast to another endemic Thai species, P. (P.) thailandicus, which is widely distributed in both temporary and permanent water bodies in the east and south of Thailand  roietensis sp. nov., Mongolodiaptomusloeiensis Watiroyram & Sanoamuang, 2017, and Mongolodiaptomusmekongensis Sanoamuang & Watiroyram, 2018 differ from P. (P.) surinensis and other diaptomids by having additional spines on segments 17\u201319  thailandicus. However, the characteristic that differentiates the new diaptomid from P. (P.) thailandicus is the presence of a single process on each side of the genital double-somite; P. (P.) thailandicus has two processes only on the right side  surinensis have unique females which can be easily differentiated. The female P5 Exp-2 of the new species is obviously asymmetrical compared with that of P. (P.) surinensis which has a slightly asymmetrical P5 Exp-2. Exp-2 in females is species-specific and unique to the genus Phyllodiaptomus: the new species has two longitudinal ridges on the anterior surface versus multi-longitudinal ridges in P. (P.) surinensis  surinensis. The new species has substantial left genital double-somite proximal bulging versus only slight asymmetry in P. (P.) surinensis. The genital double-somite of P. (P.) surinensis has a bi-lobed hyaline outgrowth ventrally, which is absent in the new species. The genital spines in the female of the new species are oriented to a posterolateral direction in dorsal view, whereas they are pointed to the lateral direction in P. (P.) surinensis. The new species has tiny spinules on Pdg 4\u20135 laterally; these are present dorsally in P. (P.) surinensis  roietensis sp. nov. has a number of morphological differences from other members of the blanci-species group as follows:The male of P. (P.) longipes.a) Antepenultimate segment with a serrated process versus smooth in P. (P.) thailandicus, P. (P.) christineae, and P. (P.) blanci.b) Urosomite(s) with a long hair or hair-like setae versus bare in P. (P.) surinensis and P. (P.) tunguidus. However, a ventral prominence is also present on the left ramus of P. (P.) tunguidus (but is absent in the left ramus of the new species) and there are only two prominences in the new species but five in P. (P.) surinensis.c) Right caudal ramus with ventral prominences as in P. (P.) irakiensis and P. (P.) thailandicus. The new species has a round or semi-circular distal margin versus triangular in P. (P.) blanci, P. (P.) christineae, P. (P.) longipes, and P. (P.) tunguidus.d) Intercoxal sclerite is modified distally into single lobe versus two lobes in P. (P.) thailandicus and slender in P. (P.) christineae.e) Right P5 coxal spine is strong and acute versus rectangular in P. (P.) irakiensis and P. (P.) blanci. In addition, three species, P. (P.) longipes, P. (P.) christineae, and P. (P.) tunguidus, have a longitudinal ridge on the posterior surface, which is absent in the new species (the first one has two minute prominences on the ridge). The right P5 basis has a triangular hyaline lamella at inner margin versus elongated in P. (P.) christineae, P. (P.) longipes, and P. (P.) tunguidus, and round in P. (P.) blanci. The left P5 basis has inner lamella versus bare in P. (P.) irakiensis and P. (P.) longipes, digitiform in P. (P.) tunguidus, and small in P. (P.) blanci. The new species lacks any ornamentation on the anterior surface but P. (P.) surinensis has two minute lateral spines (see f) Right P5 basis with a three-lobed chitinous prominence on posterior surface versus bare in ines see : fig. 54Exp-2 with three accessary lateral spines versus bare in P. (P.) tunguidus and P. (P.) blanci, and one in P. (P.) irakiensis, P. (P.) christineae, and P. (P.) longipes.g) Right P5 Enp with a bi-lobed shape versus conical in the rest of the species except P. (P.) surinensis.h) Right P5 Enp versus one-segmented in P. (P.) thailandicus, P. (P.) surinensis, P. (P.) blanci, P. (P.) christineae, and P. (P.) longipes.i) Left P5 with bi-segmented P. (P.) surinensis. However, there are three major differences among the males, i.e. the right caudal ramus, left P5 basis, and left P5 Enp as described above. The fine detail on its inner hyaline lamella on the right P5 basis is also different: triangular in the new species versus oval bi-lobed in P. (P.) surinensis.With regard to the comparative morphology above, the male of the new species is most similar to those of Phyllodiaptomus (P. (P.) tunguidus, P. (P.) blanci, P. (P.) longipes, P. (C.) annae, P. (C.) wellekensae, and P. (C.) sasikumari); he also gave morphological descriptions of these six species. In this study, the key is updated as follows:Males:Females:"}
+{"text": "Adiantum philippense revealed bioactive Nigrospora sphaerica from the leaf segment. Chemical and biological profiling through TLC\u2013bioautography and hyphenated spectroscopic techniques confirmed the presence of phomalactone as an antimicrobial metabolite.Endophyte bestows beneficial aspects to its inhabiting host, along with a contribution to diverse structural attributes with biological potential. In this regard, antimicrobial profiling of fungal endophytes from medicinal plant Escherichia coli by disc diffusion assay. The MIC was found to be significant against both Escherichia coli and Xanthomonas campestris in the case of bacteria and dermatophyte Candida albicans at 150\u2009\u03bcg/ml, respectively.The chemical investigation of the broth extract by bioassay-guided fractionation confirmed phomalactone as a bioactive antimicrobial secondary metabolite. The antimicrobial activity of phomalactone was found to be highest against Nigrospora sphaerica exhibiting a broad spectrum of antimicrobial activity against human and phytopathogenic bacteria and fungi. This work is the first report regarding the antibacterial activity of phomalactone.Overall, the results highlighted the antimicrobial potential of phomalactone from the endophyte  Taxomyces andreanae inhabiting Taxus brevifolia -5, 6-dihydroxy-2\u2009h-pyran-2-one].HPLC: The purified compound eluted at RT-4.89 with 86% purity  : 2.05 , 2.80 , 4.53 , 4.76  5.67\u20135.69 , 6.03\u20136.04  : 17.6 (C-9), 63.8 (C-5), 122.3 (C-3), 132.7 (C-8), 144.8 (C-4), 162.1 , 83.3 (C-6), 123.5 (C-7). MS : 154.06, (100%), 155.07 (M + 1) , 1750\u20131720 (ether), 1680\u20131600 , 1448 (CH3-bond), 1050\u20131150 (> C=O (1160) stretch), 1380 (\u2013CH-bond) . The results obtained indicated that Gram-negative bacteria were found to be more susceptible when compared to Gram-positive.The MIC was found to be lowest against Nigrospora sphaerica has also been reported from a vast range of host viz., Ginkgo biloba [Moringa oleifera [Phoenix dactylifera [Saccharum arundinaceum [N. sphaerica, extracted with ethyl acetate showed a significant antimicrobial activity against bacteria and fungus viz., S. aureus, B. subtilis, S. epidermidis, E. coli, K. pneumonia, P. aeruginosa, S. typhi, S. flexneri, X. campestris, V. parahaemolyticus, and C. albicans, respectively. Several bioactive compounds have been reported from N. sphaerica which suggests the potential for discovery of pharmaceutically relevant substances. Previously, it was reported N. sphaerica to produce new antimicrobial bioactive secondary metabolites, such as nigrosporolides [o biloba , Moringaoleifera , Phoenixtylifera  and Saccdinaceum . In thisorolides , nigrosporolides , lactoneorolides , epoxydoorolides , and nigorolides .Nigrospora [Xylaria [Aspergillus [Verticillium [Drechslera [Hirsutella (entomopathogenic) [Paecilomyces (entomopathogenic) [Phoma [Aspergillus fumigatus [Phomalactone is quite insecticidal\u00a0 and has grospora , 33, Xyl[Xylaria , Aspergiergillus , Verticiicillium , Drechslechslera , Hirsutehogenic) , Paecilohogenic) , and Pho) [Phoma . Phomalaumigatus , nematocumigatus , immunomumigatus , insectiumigatus , and phyumigatus .Xylaria sp. exhibited to possess weak activity at 13\u2009\u03bcg/ml against protozoan Plasmodium falciparum was reported by Carlos et al. 2008. A study conducted by Kumudini et al. 2015 reported phomalactone from N. sphaerica had larvicidal, adulticide activity against Aedes aegypti and Anopheles quadrimaculatus mosquitoes with LD50 value 0.64\u2009\u03bcg/org and 0.20\u2009\u03bcg/org, respectively. The investigation of biopotential endophytes inhabiting Zoysia japonica Steud by Kim et al. 2001 revealed phomalactone isolated from N. sphaerica exhibited potent fungitoxic activity against Phytophthora infestans and Phytophthora capsici. Its ecological role(s) in each of these species has not been rigorously studied, but its phytotoxic, fungitoxic, and insecticidal activities may be important to the various fungi that produce phomalactone.Phomalactone from endophyte N. sphaerica from medicinal plant A. philippense. Chemical investigation through bioautography N. sphaerica displayed a broad spectrum of antimicrobial activity, which indicated that endophytic N. sphaerica as a potent producer of bioactive phomalactone derivative with great potential as a natural product drug molecule.In conclusion, this is the first report on the isolation of Additional file 1: Supplementary Figure 1. HPLC analysis of purified bioactive compound phomalactone.Additional file 2: Supplementary Figure 2. LCMS spectra of purified bioactive compound phomalactone.Additional file 3: Supplementary Figure 3. Proton (1H) NMR Spectra of purified bioactive compound phomalactone.Additional file 4: Supplementary Figure 4.13C NMR Spectra of purified bioactive compound phomalactone.Additional file 5: Supplementary Figure 5. FT-IR Spectra of purified bioactive compound phomalactone."}
+{"text": "Escherichia coli is the causative agent of diarrhea in infants and animals worldwide. Many isolated strains recovered from pigs with postweaning diarrhea are multidrug resistance (MDR), and hybrids of E. coli are potentially more virulent, as enterotoxigenic E. coli (ETEC)/Shiga-toxigenic E. coli (STEC) hybrids. Here, we used whole-genome sequencing to analyze clinical isolates of the five colistin-resistant E. coli. The E. coli CAU15104, CAU15134, and CAU16060 belonged to ETEC/STEC hybrids, displaying the same serotype O3:H45 and sequence type ST4214. The E. coli CAU16175 and CAU16177 belonged to atypical enteropathogenic E. coli (aEPEC), display O4:H11 and O103:H2, ST29, and ST20, respectively. The E. coli CAU16175 carries six plasmids. An IncHI2-type plasmid, pCAU16175_1, harbors an IS26-enriched MDR region, which includes 16 antimicrobial-resistant genes. An IncFII-type plasmid, pCAU16175_3, harbors mcr-1.1, tet(M), and blaTEM\u22121B, whereas mcr-1.1 is located within a Tn2 derivative. Our findings indicate that the ETEC/STEC strains of the O3:H45 serotype as well as the aEPEC strains of the O4:H11 and O103:H2 serotypes are associated with postweaning diarrhea in swine and that some of diarrheagenic E. coli contains IS26-enriched MDR region and the mcr-1 gene located within a Tn2 derivative on IncFII plasmid.Diarrheagenic  Escherichia coli (DEC) is a leading cause of infectious diarrhea in humans and animals around the world (E. coli has six well-described pathotypes: enteropathogenic E. coli (EPEC), which is subdivided into typical EPEC (tEPEC) and atypical EPEC (aEPEC), enterotoxigenic E. coli (ETEC), enteroinvasive E. coli, enteroaggregative E. coli, diffusely adherent E. coli, and enterohemorrhagic E. coli, which is subgroup of Shiga-toxigenic E. coli (STEC)  encoded by ltc, and heat-stable enterotoxin (ST) encoded by st (stx2e (E. coli strains are defined as forming the attaching and effacing (A/E) lesions mediated by genes located on the locus of enterocyte effacement (LEE) pathogenicity island (including eae) in the intestinal epithelium but not produce Shiga-like toxin (Stx)  are present during PWD  , 4. All i (STEC) . Enteroted by st . In pigst (stx2e . Enteropin (Stx) . Moreoveand stx\u2212 . Three pring PWD , 10. Thering PWD . Atypicaring PWD .E. coli isolates, 94.15% of the strains were MDR, with antimicrobial resistance rates ranging from 2.34% for meropenem to 90.05% for nalidixic acid  Gram-negative Enterobacteriaceae has been increasing worldwide between humans and animals . In the xic acid . Among tmcr-1), which has received widespread attention in different species of Enterobacteriaceae found in animals and humans around the world since it was first reported (mcr-1 (mcr-2 to mcr-10) have been detected (mcr-1 (mcr-1.1 to mcr-1.22), mcr-2 (mcr-2.1 to mcr-2.3), mcr-3 (mcr-3.1 to mcr-3.30), mcr-4 (mcr-4.1 to mcr-4.6), mcr-5 (mcr-5.1 to mcr-5.4), and mcr-8 (mcr-8.1 and mcr-8.2) (E. coli isolates (mcr-1 were higher than the rates of mcr-1\u2013positive E. coli (64.6 vs. 49.2%)  .26 and Tn2, which is a member of the IS6 family and Tn3 family, respectively, play a key role in the dissemination of antimicrobial-resistant genes in Gram-negative bacteria  and transposons (Tns), arguably most numerous autonomous transposable elements, are crucial to shape their host genomes, particularly important in the bacterial antimicrobial resistance . IS26 anbacteria , 26. IS2 of IS26 . Tn2 is E. coli. . ISSwi1-with Tn2 . ISApl1,of mcr-1 , 30.E. coli isolates recovered from pigs with PWD and to characterize the complete sequence of a Tn2 derivative carrying mcr-1.1 on IncFII plasmid, pCAU16175_3, from a swine aEPEC isolate.In the present study, we used whole-genome sequencing (WGS) to analyze virulence and resistant genes of the five clinical colistin-resistant E. coli strains were obtained from feces samples or small intestinal content from pigs with diarrhea in China between 2014 and 2016 and identified with polymerase chain reaction (PCR) amplification of the uspA gene  based on the multilocus sequence typing and caused severe damage to IPEC-J2 cells , and two MDR aEPEC isolates (E. coli CAU16175 and CAU16177) were randomly chosen for WGS.A total of 455 Escherichia coli ATCC 25922 was used as the quality control. According to the MIC determined for each antimicrobial, the isolates were defined as \u201csusceptible,\u201d \u201cintermediate,\u201d or \u201cresistant.\u201dThe susceptibility of the five isolates to 18 antimicrobials were tested by determining the minimum inhibitory concentration (MIC) using the US Clinical and Laboratory Standards Institute (CLSI) broth micro method . The resE. coli isolates and further used WGS by the Oxford Nanopore Technologies MinION platform to get complete genome of the E. coli CAU16175.Bacterial isolates were recultured from stock; DNA was extracted using a TIANamp Bacteria DNA kit . We used WGS by the Illumina Hiseq platform to get draft genome of the five https://cge.cbs.dtu.dk). Insertion sequence typing was carried out using the ISFinder database (https://www-is.biotoul.fr/). The complete genome sequences were initially annotated with Rapid Annotation using the Subsystem Technology server (http://rast.nmpdr.org) and curated manually using the BLAST server (https://blast.ncbi.nlm.nih.gov/Blast). The obtained plasmid sequences were aligned with homologous plasmid sequences from NCBI using the BRIG tool . The serotype, multilocus sequence type, plasmid type, antimicrobial-resistant gene, and virulence gene detection were performed using the Center for Genomic Epidemiology server (RIG tool .E. coli CAU16175 as the donor and the E. coli J53 (resistant to sodium azide) as the recipient. The transconjugant was screened on BHI agar plates containing sodium azide (150 mg/L) and colistin (4 mg/L). The presence of mcr-1 in the transconjugant was confirmed by PCR, and the MICs of antimicrobial agents for the transconjugant were determined using the agar dilution method. The conjugation experiment was repeated three times, and the conjugation frequencies were calculated as the number of transconjugants per recipient. The transconjugant strains were distinguished from donor occurring natural mutants of sodium azide\u2013resistant E. coli by verifying eae, which is marker gene for E. coli CAU16175 donor.The conjugation experiment was carried out using the E. coliCAU15104, CAU15134, CAU16060, and CAU16177 isolates have been deposited at GenBank under SRA accession no. SRR10828049, SRR10828048, SRR10828047, and SRR10828046, respectively. The complete genome sequences of the E. coli CAU16175 have been deposited at GenBank under SRA accession no. SRR10813965.The draft genome sequences of the Antimicrobial susceptibility testing of the five isolates showed that they were MDR exhibiting resistant to at least three different classes of antimicrobials . All theE. coli CAU15104, CAU15134, and CAU16060 belonged to ETEC/STEC  hybrid strains, display the same serotype O3:H45 and sequence type ST4214, which are identical sequence type with the strains swine19 (LVMV00000000), swine22 (LVMY00000000), swine54 (LVMV00000000), and swine67 (LVOR00000000) from the NCBI database, all carrying fedA, fedF, iha, stb, ltcA, sepA, stx2A, and stx2B virulence genes and antimicrobial-resistant genes conferring resistant to KAN (aph(3\u2032)-Ia), STR , TE [tet(A) and tet(M)], SXT  and PB (mcr-1.1)  and aac(6\u2032)Ib-cr (associated to quinolone and aminoglycoside antibiotic). The E. coli CAU15134 and CAU16060 carry dfrA1 (associated to SXT), mcr-3.1 (associated to PB), oqxA, and oqxB, which are efflux pump conferring antibiotic resistance, and exhibit resistant to AMP associated to blaTEM\u22121B.The clinical isolates of the mcr-1.1) , 2. Besitibiotic . The E. E. coli CAU16175 and CAU16177, belonged to aEPEC , display different serotype and sequence type, O4:H11 and O103:H2, ST29 and ST20, respectively. They both carry cif, eae, espA, espB, espJ, gad, iss, nleB, and tir virulence genes and antimicrobial-resistant genes conferring resistance to beta-lactam antibiotic (blaOXA\u22121), KAN (aph(3\u2032)-Ia), GEN (aac(3)-IVa), STR (aadA1 or aadA2 or strA or strB), TE [tet(A) and/or tet(M)], SXT , and PB (mcr-1.1 and/or mcr-3.1) (E. coli CAU16175 and CAU16177 both carry aac(6\u2032)Ib-cr (associated to quinolone and aminoglycoside antibiotic), aph(4)-Ia (associated to aminoglycoside antibiotic), ARR-3 (associated to rifamycin antibiotic), catB3, cmlA1, and floR (associated to phenicol antibiotic) .The clinical isolates of the mcr-3.1) , 2. In aibiotic) . InteresE. coli CAU16175 has a total GC content of 50.47% with a single chromosome and six plasmids. The 5.62 Mb chromosome has a GC content of 50.66%. The six plasmids sequences of E. coli CAU16175 range in length from 6.99 to 190.22 kb with a GC content from 47.10 to 51.35% (mdf (A) antimicrobial-resistant gene. The pCAU16175_1 (IncHI2 type) harbors an IS26-enriched MDR region, which includes blaOXA\u22121 and 15 additional antimicrobial-resistant genes. The pCAU16175_2 (IncFIB type) harbors katP, sepA, and perA virulence genes. The pCAU16175_3 (IncFII type) harbors mcr-1.1, tet(M), and blaTEM\u22121B antimicrobial-resistant genes. The pCAU16175_5 harbors celb virulence gene (The 6.07-Mb complete genome of o 51.35% . The chrnce gene .mcr-1) increased to 8 and 16 mg/L, respectively  database. An overall nucleotide sequence identity (99.66\u201399.86%) with query coverages of 90\u201399% to pSH16G4498 (MH522423), pSH16G2457 (MH522421), pHNYJC8 (KY019259), pHNLDF400 (KY019258), pHXY0908 (KM877269), and other 25 IncHI2-type plasmids were observed (6 family insert sequences (IS26/IS15DI/IS15DIV/IS1006/ISEc59) . The structure of the MDR region comprises twelve IS6 family insert sequence  that flank containing different antimicrobial-resistant genes. The MDR region contains floR, sul2, aph(4)-Ia, aac(3)-IVa, aac(6')-Ib-cr, blaOXA\u22121, catB3, ARR-3, sul1 (two copies), dfrA12, aph(3')-Ia, sul3, aadA1, cmlA1, aadA2b, and tet(A). The MDR region is also interspersed with a number of different mobile elements including \u0394TnAs3, \u0394ISVsa3, ISVsa3, ISAba1, \u0394Tn5393, \u0394Tn2, \u0394IS15, and \u0394TnAs1.The 190.22-kb plasmid observed . We chos/ISEc59) . The pCAin China . In the pCAU16175_3 was blasted against the GenBank Nucleotide collection (nr/nt) database. An overall nucleotide sequence identity (94.46\u201397.88%) with query coverages of 80\u201389% to pEC1515-3 (CP021847), pEC974-3 (CP021843), pH1038-142 (KJ484634), pFORC_081_1 (CP029058), plasmid R1 (KY749247), and other 27 IncFII-type plasmids were observed (mcr-1pKP81-BE (KU994859) for comparison analysis , which are close to the mcr-1.1 gene.The 76.63-kb plasmid observed . We chosanalysis . Althoug2 derivative (mcmM (encoded microcin M). A five-nucleotide direct duplication (TATTT) was identified flanking the structure. Unlike the Tn2, this structure harbors mcr-1\u2013pap2-\u0394ISApl1 and IS1X2-hp-tet(M)-IS15DI, deletes the resolvase (tnpR), and truncates the transposase (tnpA). Besides, the striking features of the insertion sites of ISApl1 were found, which includes high AT content and 2-bp direct repeats (DRs) (AA)  (AA) . From thRs) (AA) . The mcrE. coli isolates recovered from feces samples or small intestinal content from pigs with diarrhea in China between 2014 and 2016 to know the E. coli pathotype and the antimicrobial susceptibility of the isolates -Ia), phenicol-resistant genes (cmlA1 and floR), tetracycline-resistant genes [tet(A) and tet(M)], trimethoprim-resistant gene (dfrA12), sulfonamide-resistant genes , streptomycin-resistant genes (strA and strB), and colistin-resistant genes (mcr-1.1). The ST4214 ETEC/STEC also harbored fimbrial adhesin genes (fedA and fedF), adherence gene (iha), heat-labile enterotoxin (LT) gene (ltcA), heat-stabile enterotoxin (ST) gene (stb), Shigella extracellular protein gene (sepA), and Shiga toxin 2 variant e genes (stx2A and stx2B). We observed the ST4214 strains were recovered from different provinces  and years  in China, which indicated this clone strains have spread in swine and should be brought to our attention.The clinical isolates of the ype ST10 . In the E. coli CAU16175 and CAU16177 belonged to aEPEC, displaying different serotype and sequence type, O4:H11 and O103:H2, ST29 and ST20, respectively. For aEPEC, serotypes O45 and O123 are frequently occurring in diarrheagenic pigs , aminoglycoside-resistant genes (aac(3)-IVa, aac(6\u2032)Ib-cr, aph(3\u2032)-Ia and aph(4)-Ia), phenicol-resistant genes , tetracycline-resistant genes [tet(A)], trimethoprim-resistant gene (dfrA12), sulfonamide-resistant genes , rifampicin-resistant gene (ARR-3), and colistin-resistant genes (mcr-1.1). Interestingly, E. coli CAU16177 also harbored mcr-3.1. Four mcr-1\u2013 and mcr-3-positive E. coli have been reported, and three E. coli isolates were recovered from pigs , non\u2013LEE-encoded effector gene (nleB), glutamate decarboxylase gene (gad), increased serum survival gene (iss), a marker gene for EPEC, intimin gene (eae), and translocated intimin receptor gene (tir).In addition, the clinical isolates of the nic pigs , 42. TheE. coli CAU16175 carries IS26-enriched MDR region in pCAU16175_1 (IncHI2 type). The IncHI2-type plasmids have been discovered as genetic elements mediating the transmission of MDR genes . The emergence of the mcr-1 gene can be traced back to the E. coli isolated in the 1980s, and the outbreak of chicken-derived mcr-1\u2013containing E. coli started in 2009 , we noticed the mcr-5 gene and the mcr-3.6 gene could be mobilized by Tn6452 and Tn6518 (belonged to Tn2 family). For the Tn6452, the mcr-5 gene was embedded within a Tn3-family Tn with 38-bp inverted repeats and flanked by 5-bp DRs, which were usually generated during the insertion  can be found below: The animal study was reviewed and approved by the Animal Ethics Committee of the China Agricultural University under the protocol CAU20140616-1.LG and YZ conceived and designed the experiments. LG, JW, SW, JS, and XW performed the experiments. LG analyzed the sequencing data and wrote the manuscript. YZ revised the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Hibiscus schizopetalus (Dyer) Hook.f.  is an ornamental plant. The aim was to investigate its antimicrobial and antioxidant activities. In vitro antiviral, antibacterial, and antioxidant activities of the 70% ethanolic extract (Et-E) of the aerial parts of the plant were determined. The Dichloromethane Fraction (DCM-F) and the n-Butanol Fraction (Bu-F) were assessed using Liquid chromatography\u2013mass spectrometry (LC-MS). The DCM-F showed higher antiviral activities against Coxsackie B4 (CoxB4) viruses (IC50 = 64.13 \u00b5g/mL) and adenoviruses (IC50 = 54.88 \u00b5g/mL) than acyclovir . The DCM-F showed higher anti-helicobacter pylori activity (MIC = 3.9 \u00b5g/mL) than clarithromycin (MIC = 1.95 \u00b5g/mL). The DCM-F inhibited Herpes Simplex Virus (HSV) Type I (IC50 = 29.85 \u00b5g/mL) and HSV Type II (IC50 = 74.17 \u00b5g/mL). The Bu-F showed higher anti-mycobacterial activity (MIC = 7.81 \u00b5g/mL) than isoniazid (MIC = 0.24 \u00b5g/mL) and higher antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA)(MIC = 7.81 \u00b5g/mL) than vancomycin (MIC = 3.9 \u00b5g/mL). Antioxidant assays included total antioxidant capacity (TAC), 2,2\u2032-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS), 2,2-diphenyl-1-picryl-hydrazyl (DPPH), and iron reducing power. The Bu-F showed the highest antioxidant activity. Chemical profiles were analyzed using HPLC-HR\u2013ESI\u2013MS to identify the metabolites responsible for these biological activities. We identified more than 60 metabolites that belong to anthocyanins, flavonoids, phenolics, terpenes, sterols, and fatty acids. In conclusion, Hibiscusschizopetalus is endowed with metabolites that could be used against viruses and antibiotic-resistant bacteria. They can also be potent antioxidants. Antimicrobial resistance to classical antibiotics is a major problem that negatively impacts treatments of common infectious diseases caused by fungi, bacteria, viruses, and parasites. In order to overcome this major problem, natural products offer a viable alternative .Hibiscus is a member of Malvaceae family and is cultivated globally as an ornamental and medicinal plant -2,5-diphenyl tetrazolium bromide (MTT) was performed spectrophotometrically using a Perkin-Elmer ELISA reader (HTS 7000 plus) at 540 nm . The minimum concentration required for the inhibition of bacterial growth (MIC) and the concentration required for inhibition of 90% of the bacterial growth (MIC90) were determined from the corresponding dose-response curve. The MTT assay was done using an automatic BioTek plate reader at 620 nm. Activity against n method . H. pyloM. tuberculosis (M. Tub ATCC 27294) was detected using the Microplate Alamar Blue Assay (MABA) [M. tuberculosis inoculum (105 CFU/mL) was added. Plates were incubated at 37 \u00b0C for 4 days, then 20 \u03bcL of Alamar Blue solution and 12.5 \u03bcL of 20% Tween 80 were added. The plates were re-incubated at 37 \u00b0C for 24 h. The color intensity was measured using an ELISA microplate reader at 590 nm. The experiment was done in triplicates.The anti-mycobacterial activity against y (MABA) . Isoniaz(MRSA ATCC 27294) was assessed using the microdilution method in the presence of tetrazolium salts [6 CFU/mL), and 50 \u03bcL of the sample dilutions (0.24\u2013125 \u03bcg/mL) were added. The plates were incubated at 37 \u00b0C for 24 h. To each well, 40 \u03bcL of XTT were added and incubated for 1 h in the dark at 37 \u00b0C. For each well, the color intensity was measured at 492 nm.The anti-MRSA activity um salts . VancomyA.Total antioxidant capacity assay (TAC).The antioxidant activity of the samples was assessed using the phosphomolybdenum method ,49. AbsoB.2,2\u2032-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical scavenging assay.Hibiscus extract and its fractions [The ABTS radical scavenging assay was used to assess the free radical scavenging activity of the ractions ,49. MeasC.2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging assay.The anti-oxidant activity was measured as previously described  AbsorbanD.Reducing power assay.The reducing power was assessed by the evaluating the ability of the samples to change potassium ferricyanide to potassium ferrocyanide. Absorbance was measured at 700 nm ,49. The Experiments for antioxidants were done in triplicates and means were calculated.m/z 150\u20131500. The crude methanolic extract and fractions for mass spectrometry were prepared at 1 mg/mL. Metabolomic analysis of the samples was done using LC-HR-ESI-MS ,51. We uThe data were imported and analyzed using MZmine 2.20 . The masp < 0.05.Data analysis was done using analysis of variance (ANOVA). The significance of differences between means was done using Duncan\u2019s multiple range tests at H. schizopetalus is endowed with a lot of chemical compounds (more than 60 metabolites) that could be clinically used in fighting viruses, such as adenoviruses, HSVI, and CoxB4. They can also be used as effective therapies against antibiotic resistant pathogens such as M. tuberculosis, MRSA, and H. pylori. Finally, they can be potent antioxidants that can protect cells from free radical damage in cancer, heart diseases, and other serious health conditions."}
+{"text": "Tumor metastasis accounts for the majority of cancer-related deaths; it is therefore important to develop preclinical models that faithfully recapitulate disease progression. Here, we generated paired organoids derived from primary tumors and matched liver metastases in the same colorectal cancer (CRC) patients. Despite the fact that paired organoids exhibit comparable gene expression and cell morphology, organoids from metastatic lesions demonstrate more aggressive phenotypes, tumorigenesis, and metastatic capacity than those from primary lesions. Transcriptional analyses of the paired organoids reveal signature genes and pathways altered during the progression of CRC, including SOX2. Further study shows that inducible knockdown of SOX2\u00a0attenuated invasion, proliferation, and liver metastasis outgrowth. Taken together, we use patient-derived paired primary and metastatic cancer organoids to model CRC metastasis and illustrate that SOX2 is associated with CRC progression and may serve as a potential prognostic biomarker and therapeutic target of CRC. To the Editor,Tumor heterogeneity plays a key role in cancer progression and therapy resistance . HoweverInvasion is a fundamental step in tumor progression toward metastasis. To study collective invasion, we cultured paired organoids in a 3D invasion matrix Fig. a. AlthouP < 0.05; fold change > 2.5) that were significantly upregulated in metastatic organoids  inducible expression of shRNA targeting SOX2 was established in metastatic organoids Fig. SC and D. nes Fig. E-G. Furtnts Fig. H. We thents Fig. . Taken tIn summary, the present study highlights the potential of patient-derived paired primary and metastatic cancer organoids as an experimental model for investigating CRC progression. We identified a significantly dysregulated gene between paired organoids, SOX2, which could be a prognostic biomarker, and perhaps a potent therapeutic target in the treatment of CRC.Additional file 1: Supplementary\u00a0Table S1. Summary of patient-derived CRC organoid lines and corresponding clinical data.Additional file 2: Supplementary Figures. Figure S1. Paired organoids derived from primary and metastatic CRC recapitulate the histopathological structure of parental tumor. A and B, Organoids architecture resembles parental tumor epithelium. Representative bright-field images of organoids together with H&E staining of parental tumors and patient-derived organoids. The scale bar represents 100 \u03bcm. C and D, Representative IHC sections for the intestinal epithelial marker CDX2. The scale bar represents 100 \u03bcm. Figure S2. Organoids derived from liver metastatic lesions exhibited the most aggressive phenotypes with significant high tumorigenic and metastatic potential. A, Representative micrographs of organoids in 3D invasion assay. Tumor organoids showed the smooth and protrusive leading fronts, respectively. B, Micrographs of the paired organoids stained with phalloidin\u2013F-actin (right). The scale bar represents 100 \u03bcm. C, qRT-PCR analysis of MMP-2 in paired organoid lines. Error bars indicate SEMs. *P=0.0207, **P =0.0083, ***P=0.0008 , ***P=0.0006 . D, Western blot analysis of the MMP-2 protein expression in paired organoid lines. \u03b1-tubulin was used as a loading control. E, Representative IHC sections for Ki67 in human colorectal tumor tissues and paired organoid lines. The scale bar represents 100 \u03bcm. F, Representative IHC sections for Ki67 in organoid xenografts and organoids derived from xenografts. The scale bar represents 100 \u03bcm. G, Representative gross image, histopathology and Ki67 staining of whole liver from primary organoid xenografts. The scale bar of the whole liver represents 1cm. The black scale bar represents 100 \u03bcm. Figure S3. A, Representation of the up-regulated genes in 13L organoids. Error bars indicate SEMs. *P = 0.0283, ***P = 0.0004, ****P < 0.0001 (one-way ANOVA). B, qRT-PCR analysis of the up-regulated genes in 13L organoids. Values were normalized to mean levels in 13a organoids. Error bars indicate SEMs. *P=0.02, ***P=0.0008 ****P < 0.0001 (one-way ANOVA). C, RNA sequencing analysis of SOX2 in human colorectal cancer tissues and normal colon tissues. D, qRT-PCR analysis of SOX2 in tumor tissues and paired normal colon tissues. Values were normalized to mean levels in normal colon tissues. Error bars indicate SEMs. **P = 0.0018, ****P < 0.0001 (one-way ANOVA). E, Western blot analysis of the SOX2 protein expression in tumor tissues and paired normal colon tissues. \u03b1-tubulin was used as a loading control. F, Representative IHC sections for SOX2 in human colorectal tumor tissues and paired normal colon tissues. The scale bar represents 100 \u03bcm. Figure S4. Silencing SOX2 in metastatic organoids attenuated cell invasion and proliferation and suppressed liver metastasis in vivo. A, qRT-PCR analysis of SOX2 in paired organoid lines (left). Values were normalized to mean levels in 21a organoids. Error bars indicate SEMs. ****P < 0.0001(Unpaired t test). Western blot analysis of the SOX2 protein expression in paired organoid lines. \u03b1-tubulin was used as a loading control (right). B, Representative IHC sections for SOX2 in human colorectal tumor tissues and organoids. The scale bar represents 100 \u03bcm. C, qRT-PCR analysis of SOX2 in the 13L-shRNA-1/2 (left) and 21L-shRNA-1/2 (right) organoids after 3 days of Dox-induction. The level of SOX2 was compared to that in the untreated sample. Error bars indicate SEMs. ***P=0.0001, ****P < 0.0001 (two-way ANOVA). D, Western blot analysis of the expression of SOX2 in the 13L-shRNA-1/2 and 21L-shRNA-1/2 organoids after 3 days of Dox-induction. \u03b1-tubulin was used as a loading control. E, Representative micrographs of Dox-transduced the 21L-shRNA-1/2 organoids after 5 days and stained with phalloidin\u2013F-actin. The scale bar represents 100 \u03bcm. F, Proliferation of the 21L-shRNA-1/2 organoids were examined by CTG cell viability assays following 3- or 5-days growth in the presence or absence of Dox. Error bars indicate SEMs. **p=0.0019, ****P < 0.0001 (two-way ANOVA). G, Representative micrographs of colonies arising from the 21L-shRNA-1/2 organoids (top), with magnified insets showing colonies (bottom). The scale bar represents 200 \u03bcm. Colony forming efficiency in G was calculated and compared (right). Error bars indicate SEMs. **P=0.0015 (two-way ANOVA). H, Representative Ki67 immunostaining of liver metastases of the 13L-shRNA-1/2 organoids (left) and ratio of Ki67-positive tumor cells in liver metastases (right). Scale bars, 200 \u03bcm (top) and 100 \u03bcm (bottom). Each dot indicates individual mice. Error bars indicate SEMs. *p=0.0197, **P=0.0044 (two-way ANOVA). Figure S5. SOX2 is overexpressed in primary organoids. A, qRT-PCR analysis of SOX2 in primary organoid lines. Values were normalized to mean levels in control organoids. Error bars indicate SEMs. ****P < 0.0001 (one-way ANOVA). B, Western blot analysis of the SOX2 protein expression in primary organoid lines. \u03b1-tubulin was used as a loading control. C, Representative micrographs of colonies arising from the control, LV-Vector and LV-SOX2 primary organoid lines. The scale bar represents 200 \u03bcm. D-F, Colony forming efficiency in C was calculated and compared. Error bars indicate SEMs. ***P=0.0008, ****P < 0.0001 (one-way ANOVA). G, Proliferation of the control, LV-Vector and LV-SOX2 primary organoid lines were examined by CTG cell viability assays. Error bars indicate SEMs. ***P=0.0008, ****P < 0.0001 (one-way ANOVA). H, Representative micrographs of organoids in 3D invasion assay. Arrowheads, protrusive leading fronts. Micrographs of the paired organoids stained with phalloidin\u2013F-actin. The scale bar represents 100 \u03bcm.Additional file 3. Supplementary Materials and Methods."}
+{"text": "Correction to: Mol Neurodegeneration 15, 34 (2020)https://doi.org/10.1186/s13024-020-00383-7The original article [incorrectly displays 6th author, Bj\u00f6rn Oskarsson\u2019s name with middle initial. The correct presentation can be viewed in this Correction article. article  incorrec"}
+{"text": "In this study, we investigated the presence of carbapenemase genes, mcr-1, and tet(X) in 298 Escherichia coli strains obtained from a teaching hospital in China. In total, eight (2.68%), six (2.01%), and one (0.34%) E. coli isolates carried blaNDM, mcr-1, and tet(X4), respectively. The blaNDM gene was located on IncX3 (n = 4), F2:A-:B- (n = 3), and F2:A1:B1 (n = 1) plasmids, with high similarity to multiple plasmids belonging to the same incompatibility type from Enterobacteriaceae. Six MCR-producing strains contained mcr-1-carrying IncI2 plasmids, organized similarly to other mcr-1-bearing IncI2 plasmids from animals in China. The blaCTX\u2212M\u221255/64/132/199 gene located within a typical transposition unit (ISEcp1-blaCTX\u2212M-orf477\u0394) was inserted near dnaJ to generate 5-bp direct repeats in four mcr-1-positive plasmids. The tet(X) and another four resistance genes  were co-located on an IncX1 plasmid, highly similar to other tet(X4)-carrying IncX1 plasmids from Escherichia and Klebsiella of animal or food origin, except that the conjugative transfer region of IncX1 plasmids was absent in our plasmid. Although a low prevalence of blaNDM, mcr-1, and tet(X) was observed in E. coli from patients in this study, their dissemination associated with some successful pandemic plasmids is of great concern. The continued surveillance of these crucial resistance genes in patients should be strengthened.The emergence and spread of carbapenemase genes, colistin resistance genes  Escherichia coli as the leading pathogen  bacteria are rapidly increasing, which poses a serious threat to clinical therapy and public health WHO, . In 2019Escherichia coli is one of the predominant carriers of blaNDM; some lineages, such as ST167, ST410, and ST617, and popular plasmids  are associated with blaNDM global dissemination in E. coli  in E. coli from pigs in China can reduce the clinical efficacy of these agents   gene has recently been increasingly reported in E. coli from different sources , mainly in China  in E. coli , and carbapenemases in human or animal gut microbiomes , and carbapenemases is worrisome, as options for therapy are limited. Therefore, there is an urgent need for evaluating and monitoring the prevalence of pathogenic bacteria carrying mcr, tet(X), and carbapenemases. In this study, we investigated the prevalence and characterization of mobilized resistance genes tet(X4), mcr-1, and blaNDM in E. coli strains isolated from a teaching hospital in Jiangsu Province, China.A previous study reported the discovery of E. coli strains were isolated from patients in a teaching hospital in Jiangsu Province, China. The presence of tet(X), mcr-1, and carbapenemase genes  was detected by PCR and sequencing according to previously described protocols -carrying E. coli is described in From November 2019 to November 2021, 298 non-duplicate blaNDM/mcr-1/tet(X)-carrying E. coli isolates were tested for susceptibility to 14 antimicrobial agents, namely, ampicillin, cefotaxime, meropenem, gentamycin, amikacin, streptomycin, tetracycline, tigecycline, chloramphenicol, nalidixic acid, ciprofloxacin, colistin, fosfomycin, and sulfamethoxazole/trimethoprim, by using the agar dilution method or broth microdilution method (limited to colistin and tigecycline). E. coli ATCC 25922 was used as the quality control strain. The results were interpreted according to Clinical and Laboratory Standards Institute (CLSI) M100, 30th edition .The on CLSI, . StreptoblaNDM/mcr-1/tet(X)-carrying E. coli isolates as donor bacteria and E. coli C600 (high-level streptomycin resistance) as the recipient strain following a previously described protocol , colistin (for mcr-1), or tigecycline [for tet(X)]. They were further confirmed by detecting the presence of blaNDM, mcr-1, and tet(X) using PCR. Transfer frequencies were calculated as the number of transconjugants per recipient. Conjugation experiments were performed at least in triplicate.Conjugation experiments were performed using blaNDM/mcr-1/tet(X)-carrying E. coli isolates using a TIAN amp Bacteria DNA kit  and sequenced by the Illumina Hiseq platform. The 150 bp pair-end raw reads were trimmed with the NGSQC toolkit v.2.3.3 and assembled into contigs using SPAdes v.3.8.2 , XYEH3783 (blaNDM \u2212 5), and XYEH3934 [tet(X4)], were sequenced using Nanopore MinION and assembled with Unicycler version 0.4.9. The remaining blaNDM/mcr-1-carrying plasmid contigs were assembled into a complete plasmid sequence using PCR and Sanger sequencing (http://blast.ncbi.nlm.nih.gov/Blast.cgi). The genome sequences were analyzed for multi-locus sequence typing (MLST) and resistance genes using MLST and ResFinder, respectively (https://cge.food.dtu.dk//services). Analysis and annotation of all blaNDM/mcr-1/tet(X4)-carrying plasmids were performed using the RAST server , ResFinder, PlasmidFinder (https://cge.food.dtu.dk/services/PlasmidFinder/), BLAST, and the Gene Construction Kit 4.5 . Blast Ring Image Generator (BRIG) version 0.95 was used to visualize the blaNDM-carrying plasmid comparisons -positive E. coli strains and blaNDM-positive E. coli ST167 in this study and the NCBI database was constructed based on core genome single nucleotide polymorphisms (SNPs) using ParSNP v. 1.2 (https://github.com/marbl/parsnp), respectively. ChiPlot (http://chiplot.online/#) was used to visualize the phylogenetic tree.The phylogenetic tree of all blaNDM/mcr-1/tet(X4)-carrying plasmids have been deposited in GenBank under accession number PRJNA901528.The genome sequences and blaNDM gene has been globally distributed as the third most common carbapenemase-encoding gene  or blaNDM \u2212 6 (n = 1) (E. coli were resistant to meropenem (MIC \u2265 8 mg/L), ampicillin, cefotaxime, gentamicin, tetracycline, chloramphenicol, nalidixic acid, ciprofloxacin, and sulfamethoxazole/trimethoprim , ST167 (n = 2), ST361 (n = 1), and ST1193 (n = 1), respectively  . None ofethoprim . Resistaectively . E. coliecipient .blaNDM gene was located on IncX3 (n = 4), F2:A-:B- (n = 3), and F2:A1:B1 (n = 1) plasmids  were obtained from four ST156 NDM-5-producing E. coli strains. These IncX3 plasmids were identical, except that two copies of IS1294 were inserted into the accessory replication gene bis and DNA relaxase gene taxC, respectively, in pYUXEEH1271-NDM and pYUXEEH1278-NDM  of animal or human origin  and F2:A1:B1 plasmid pYUXYEH3783-NDM . These IncFII plasmids showed high similarity to multiple plasmids belonging to the same incompatibility type from Enterobacteriaceae  from food-producing animals or humans  have been identified in Enterobacteriaceae from animals, humans, and the environment , ampicillin, cefotaxime, nalidixic acid, ciprofloxacin, and some other agents , tet(A) (n = 3), fosA3 (n = 3), and rmtB (n = 1) (All r agents . In addi (n = 1) . The pre (n = 1) .mcr-1-positive E. coli strains could transfer mcr-1 to E. coli C600 by conjugation with frequencies of 10\u22126 to 10\u22124 transconjugants/recipient  from E. coli and pSCS23  from Salmonella  , was inserted near dnaJ to generate 5-bp direct repeats (5\u2032-GAAAA-3\u2032)  . The blaAAAA-3\u2032) . The co-tet(X4) in E. coli from pigs in China in 2019  has been rarely reported in E. coli from patients. It was detected in 4.5%  fecal samples of hospital patients in Guangdong province, and only 0.07%  E. coli isolates from patients in China were identified to carry tet(X4) -positive E. coli isolate, XYEH3934. This strain exhibited high-level tigecycline resistance (MIC = 32 mg/L) and was also resistant to ampicillin, cefotaxime, streptomycin, tetracycline, chloramphenicol, nalidixic acid, fosfomycin, and sulfamethoxazole/trimethoprim (tet(X4) to E. coli C600 by conjugation.Since the discovery of the plasmid-mediated tigecycline resistance gene ethoprim . Howevertet(X) and another four resistance genes  were co-located on an IncX1 plasmid pYUXYEH3934-2 with a size of 31,816 bp. It was highly similar to other tet(X4)-carrying IncX1 plasmids from porcine E. coli in China, such as plasmids pYUYZMP62 (MW439254) and pDW28-tet(X4) (ON390803), as well as tet(X4)-positive plasmids from Escherichia and Klebsiella (tet(X4) was also associated with ISCR2 in pYUXYEH3934-2 (tet(X4) is mainly mediated by the horizontal transfer of insertion sequences  and plasmids such as IncX1 in the present study -positive E. coli strains has been previously described, such as E. coli ST877, ST10, and ST48 between animals and humans  and four plasmids . Among tebsiella . Howeverebsiella , which mebsiella , tet(X4)EH3934-2 . The rap Aminov, . HoweverblaNDM/mcr-1/tet(X4)-positive E. coli isolates in the present study. The results showed that the 15 E. coli strains were divided into four clades  and Italy (n = 1) were clustered in one clade with a close relationship  was observed in E. coli isolates from patients in one hospital in this study, their dissemination in E. coli associated with some successful pandemic plasmids  is of great concern. Conversely, E. coli strains from food or food-producing animals in China show a higher prevalence of blaNDM, mcr-1, and tet(X)  can be found below: LS and JW conceived the study. LS, G-ZS, YJ, C-YM, H-YW, and G-MK carried out the experiments. LS, YJ, Z-YW, and JW analyzed the data. JW wrote the manuscript. XJ revised the manuscript. All authors have read and approved the final version of the manuscript."}
+{"text": "Aspergillus is a filamentous fungus renowned for its extraordinary ability to produce active natural products of high therapeutic value and economic importance. This review is the first to focus on Aspergillus-derived alkaloids. Through an extensive literature review and data analysis, 263 alkaloids are categorized according to their structural features into those containing cytochalasans, diketopiperazine alkaloids, quinazoline alkaloids, quinoline alkaloids, indole alkaloids, pyrrolidine alkaloids, and others. These metabolites exhibited diverse biological activities, such as antibacterial activity, cytotoxicity, anti-inflammatory activity, and \u03b1-glucosidase, ACE, and DPPH inhibitory activities. The bioactivity, structural diversity, and occurrence of these alkaloids are reviewed in detail.Alkaloids represent a large family of natural products with diverse structures and bioactivities. These compounds and their derivatives have been widely used in clinics to treat various diseases. The endophytic  Aspergillus is one of the most widely studied filamentous fungi and renowned for its extraordinary productivity when it comes to active natural products with therapeutic values, making it of economic importance quinazoline-3,6-dione (163) and protuboxepin K (164). Compounds 153, 160\u2013164 exhibited mosaic virus TMV inhibitory activity, with IC50 values of 34.8, 37.9, 32.2, 42.4, 39.5, and 35.2 \u03bcM, respectively -L-tryptophan methyl ester (236), N-[(2S)-2hydroxy-1-oxo-3-phenylpropyl]-L-tryptophan (237), acudioxomorpholine (238), and emindole SB (239) benzodiazepine-5,11-dione (241)  [The fungus ne 241) , Table 6 , Table de (242) .A. fumigatus was studied and yielded the known metabolites 14-norpseurotin (243) and pseurotin A (244). Compound 243 promoted the neurite outgrowth of PC12 cells at 10.0 \u00b5M [244 was also produced from Aspergillus sp. EJC08 associated with Bauhinia guianensis [A. fumigatus associated with Erythrophloeum fordii Oliv [A. fumigatus associated with Heteroscyphus tener (Steph.) Schiffn [A. fumigatus D associated with Edgeworthia chrysantha Lindl [Aspergillus sp. 87 [244 exhibited inhibition of S. aureus, B. subtilis, Pseudomonas aeruginosa, and E. coli, with MICs of 15.62, 31.25, 31.25, and 15.62 \u03bcg/mL, respectively [50 value of 5.20 \u00b5M [The endophyte  10.0 \u00b5M . Compounianensis , A. fumidii Oliv , A. fumi Schiffn , A. fumiha Lindl , and Asps sp. 87 . Compounectively , and inh 5.20 \u00b5M .A. fumigatus LN-4 obtained from Melia azedarach led to the discovery of 244 and pseurotin A1 (245), which demonstrated nontoxicity toward brine shrimps [A chemical study of  shrimps .246), and the known compound 11-O-methylpseurotin A (247) were collected from A. fumigatus Y0107 of Crocus sativus Linn (saffron). These compounds were inactive against Pantoea agglomerans, Agrobacterium tumefaciens, Erwinia sp, and Ralstonia solanacearum [A new alkaloid, 11-acetyl-pseurotin A2  and adenine (250). Compound 248 was also isolated from A. fumigatus LN-4 obtained from Melia azedarach. Compounds 248 and 250 suppressed melanin production in B16 melanoma cells with IC50 values of 144.7 \u00b1 2.35 and 100.4 \u00b1 3.05 \u00b5M, respectively [A chemical study on unds 248 , Table 7ectively ,38.251), obtained from A. fumigatus collected from Erythrophloeum fordii Oliv. (Leguminosae), did not inhibit NO production [The known alkaloid lumichrome (oduction .Aspergillus sp. TJ23 collected from leaves of Hypericum perforatum (St John\u2019 Wort) yielded a new pyridone alkaloid, asperpyridone A (252), which improved glucose uptake in HepG2 cells at 50 \u03bcM [The endophyte at 50 \u03bcM .253) and 4-amino-2-hydroxymethylpyridin-5-ol (254) and the known compound 5-hydroxy-2-hydroxymethylpyridine-4(1H)-one (255) were obtained from A. flavus GZWMJZ-288 on Garcinia multiflora. None of them demonstrated any inhibitory activity against gram-positive S. aureus ATCC6538, S. aureus ATCC25923, MRSA, gram-negative Pseudomonas aeruginosa ATCC10145, or E. coli ATCC11775, nor against the pathogenic fungi C. albicans ATCC10231 or C. glabrata ATCC2001 at 100 \u03bcg/mL [New alkaloids 2-Hydroxymethyl-5-(3-oxobutan-2-yl)aminopyran-4(4H)-one , cyclopeptine (257), and trans-3-(3\u2032-hydroxybenzylidene)-3,4-dihydro-4-methyl-1H-1,4-benzodiazepin2,5-dione (258). None of them showed any AChE inhibitory activity [A study on activity .259), together with known compounds penipanoid A (260), and fuscoatramide (261) were obtained from A. flavipes DZ-3 of Eucommia ulmoides Oliver. These compounds were inactive against antioxidant and \u03b1-glucosidase capacities [A new alkaloid, asperflaloid B (pacities .262), obtained from the fungus Aspergillus sp. 87, was devoid of antibacterial activity [A new alkaloid, aspergilamide A -1-(4-hydroxy-phenyl)-4-(4-methoxyphenyl)buta-1,3-diene-2,3-diyl)diformamide (263) was isolated from the A. fumigatus HQD24, but it did not display inhibition of splenic lymphocyte growth [N,N\u2019-, followed by A. versicolor , A. flavipes , A. terreus , A. nidulans , other species  including A. aculeatus (1), A. amstelodami (1), A. cristatus (1), A. creber (1), A. micronesiensis (1), and A. ochraceus (1), and Aspergillus unknown species . Detailed analysis revealed that the discovery probability of known alkaloids is high (61.98%)  . The micrsicolor , 10.87%,82) displayed potent inhibitory activity against the human FLT3-ITD mutant AML cell line MV4-11, with an IC50 value of 0.22 \u03bcM [252) improved glucose uptake and is a potential hypoglycemic agent [The alkaloids summarized in this literature exhibited antibacterial activity; cytotoxicity; anti-inflammatory activity; and \u03b1-glucosidase, ACE, and DPPH inhibitory activities. Many of these metabolites demonstrated potent biological activity. For example, gartryprostatin C ( 0.22 \u03bcM . Asperpyic agent . HoweverAspergillus is a dominant community in natural products exploration. In this literature, the bioactivity, structural diversity, and biosources of alkaloids derived from plant endophytic Aspergillus species during January 2004 to May 2023 were described. Approximately 263 alkaloids isolated from 46 strains of Aspergillus species were reviewed according to their structural features, including cytochalasans, diketopiperazine alkaloids, quinazoline alkaloids, quinoline alkaloids, indole alkaloids, pyrrolidine alkaloids, and others. Among them, 149 alkaloids have significant physiological activities, such as antibacterial activity, cytotoxicity, anti-inflammatory activity, and \u03b1-glucosidase, ACE, and DPPH inhibitory activities. Therefore, these active alkaloids have tremendous potential as lead compounds for the exploitation of new drugs. The interdisciplinary research of chemistry, biology, and pharmacology for alkaloids derived from plant endophytic Aspergillus sp. has attributed to driving the application of alkaloids in the drug discovery and development.Plant endophytic fungi have provided abundant resources of natural products with unique structural features and diverse biological activities, which play a critical role for drug development. The plant endophytic"}
+{"text": "Limbic-predominant age-related TDP-43 encephalopathy neuropathologic change (LATE-NC)  frequentThe amygdala is also preferentially affected by argyrophilic grains (AGs) and age-related tau astrogliopathy (ARTAG). AGs are age-related lesions in which four-repeat tau is selectively accumulated . AGs areAt present, the pathogenic relationship between LATE-NC, AGs, and GFAs in the amygdala remains to be elucidated. For example, although a few previous studies supported the potential relationship between LATE-NC and AGs , 4, a reTo address these issues, first, we examined whether amygdala GFAs and AGs are independent risk factors of LATE-NC in a Japanese series with a low to moderate Braak neurofibrillary tangle (NFT) stage. Then, independent pathological risk factors of the formation of AGs in this series were also explored. Finally, whether LATE-NC, AGs, and amygdala GFAs have independent effects on severe loss of neurons in the amygdala was examined.From 1,180 autopsy cases who were registered in the database at the Department of Neuropsychiatry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences as of the end of June 2023, we first selected 501 cases in which all of the following pathological data were available: Braak NFT stage, Thal phase, CERAD neuritic plaque score, LBD subtypes, Braak Parkinson\u2019s disease stage, Saito AG stage , LATE-NCFrom 501 cases, we excluded any cases having following pathologies: NFTs with Braak stages V-VI, Lewy bodies in any regions including the olfactory bulb, NFTs which quantity fit the NINDS-PSP criteria, tufted astrocytes and astrocytic plaques in any region, amyotrophic lateral sclerosis or frontotemporal lobar degeneration with TDP-43-positive or FUS-positive inclusions, Pick\u2019s disease, globular glial tauopathy, multiple system atrophy, trinucleotide repeat diseases, inflammatory diseases, leukodystrophies, and lysosomal storage diseases. Finally, 72 cases that were Braak NFT stages I-IV and Thal phases 0\u20134 but lacked any other neurodegenerative changes except for AGs, GFAs, LATE-NC, or HS which is closely associated with LATE-NC, were extracted .The mean age at death in 72 cases was 71.6\u2009\u00b1\u200911.6\u00a0years. Of 72 cases, 52 cases had GFAs in the amygdala (72.2%), and 26 had AGs (36.1%) Table : Fig. S2Four of 72 cases (5.6%) had HS Figs.\u00a0E. All HSp < 0.001, Fig. Among 72 cases, five (6.9%) had severe neuronal loss in the amygdala (Table p\u2009=\u20090.0264) and amygdala GFA stage 4  were significant predictors of LATE-NC  demonstrated that the age at death  and amygdala GFA stage 4  were independent risk factors of AGs  revealed that the age at death  was an independent risk factor of severe neuronal loss with stage 3.In the multivariate binomial logistic regression regarding tissue degeneration in the amygdala , only Saito AG stages II-III  independently contribute to amygdala degeneration. Although LATE-NC was not a statistically significant factor in the present study, it might be explained by the small number of LATE-NC-positive cases in our series. Indeed, it was reported that LATE-NC was associated with the amygdala volume assessed using postmortem MRI . RegardiAdditional file 1: Fig. S1. Representative figures of neuronal loss stage in the amygdala. A\u2013C Stage 0. Neuronal loss and glial proliferation is absent. D\u2013F Stage 1. Mild neuronal loss with minimal gliosis is noted. G\u2013I Stage 2. Moderate neuronal loss with gliosis is present, but tissue rarefaction is not evident. J\u2013L Stage 3. Severe neuronal loss with remarkable glial proliferation is noted. Tissue rarefaction is also seen. Scale bars: A, D, G, J 200 \u03bcm, B, E, H, K 100 \u03bcm, C, F, I, L 50 \u03bcm.Additional file 2: Table S1. Demographic data in cases with Braak stages I-IV by LATE-NC status.Additional file 3: File S1. Supplementary file 1: Supplementary materials and methodsAdditional file 4: Fig. S2. TDP-43 pathology, granular fuzzy astrocytes (GFAs), argyrophilic grains, hippocampal sclerosis, and tissue degeneration in the amygdala in representative cases. A\u2013F Pathological findings in a case with Braak NFT stage II, Thal phase 0, amygdala GFA stage 2, Saito AG stage III, and LATE-NC stage 2. A, B Phosphorylated TDP-43 accumulation in the amygdala A and dentate gyrus in the hippocampus B. pS409/410 immunohistochemistry. Scale bar: 25 \u03bcm. C A GFA in the amygdala. AT8 immunohistochemistry. Scale bar: 25 \u03bcm. D AGs in the amygdala. Gallyas method. Scale bar: 25 \u03bcm. E Hippocampal sclerosis. Hematoxylin-eosin stain. Scale bar: 100 \u03bcm. F Severe loss of neurons with gliosis in the amygdala. Hematoxylin-eosin stain. Scale bar: 25 \u03bcm. G\u2013L Pathological findings in a case with Braak stage II, Thal phase 0, amygdala GFA stage 4, Saito AG stage III, and LATE-NC stage 0. G This case lacked phosphorylated TDP-43-positive lesion in any region. The amygdala. pS409/410 immunohistochemistry. Scale bar: 25 \u03bcm. H, I GFAs in the amygdala. AT8 immunohistochemistry. Scale bar: 25 \u03bcm. J AGs in the amygdala. Gallyas method. Scale bar: 25 \u03bcm. K Neither loss of pyramidal neurons nor gliosis is noted in the hippocampal CA1. Hematoxylin-eosin stain. Scale bar: 100 \u03bcm. L Severe neuronal loss with gliosis in the amygdala. Hematoxylin-eosin stain. Scale bar: 25 \u03bcm."}
+{"text": "S-adenosylmethionine (SAM) and total cysteine (tCys) concentrations. Serum 5-MTHF concentrations demonstrated a stronger negative correlation with tHcy/tCys than with tHcy alone. The negative correlation between betaine and tHcy concentrations was stronger in the low 5-MTHF group than in the high 5-MTHF group. The 5-MTHF status could be linked to Hcy flux into the transsulfuration pathway via SAM. Therefore, the tHcy/tCys ratio may be a more sensitive indicator of the 5-MTHF status than tHcy alone. Furthermore, a low 5-MTHF status can enhance Hcy metabolism via betaine.Maintaining optimal one-carbon metabolism (OCM) is essential for health and pregnancy. In this cross-sectional study, folate status was assessed based on 5-methyltetrahydrofolate (5-MTHF) levels, and the association between 5-MTHF and OCM-related metabolites was investigated in 227 female Japanese university students aged 18\u201325 years. The participants were divided into high and low 5-MTHF groups based on their folate status. Serum samples of the participants were collected while they were fasting, and 18 OCM-related metabolites were measured using stable-isotope dilution liquid chromatography\u2013electrospray tandem mass spectrometry. The association between serum 5-MTHF and OCM-related metabolite concentrations was assessed using Spearman\u2019s rank correlation coefficient. Serum 5-MTHF concentrations were negatively correlated with total homocysteine (tHcy) concentrations and positively correlated with  S-adenosylmethionine (SAM)-dependent methyl transfer reactions [One-carbon metabolism (OCM) comprises a folate cycle and choline metabolic pathway linked to a methionine cycle; homocysteine (Hcy) in the methionine cycle is connected to the transsulfuration pathway . OCM is eactions , nucleiceactions ,2, and aeactions .2-dependent MTHF reductase  [S-methyltransferase , which is expressed primarily in the liver and kidneys and remethylates Hcy to produce methionine and dimethylglycine (DMG) [S-adenosylhomocysteine (SAH) [6-dependent enzyme cystathionine-\u03b2-synthase , which condenses serine to Hcy thiol [6-dependent enzyme cystathionine \u03b3-lyase  to Cys [Folate mediates the transfer of one-carbon units in OCM . Folic a.5.1.20) . The maj.5.1.20) ,8. Methi.5.1.20) ,7,8. Cho.5.1.20) . Betainene (DMG) ,11. Methne (DMG) . SAM is ne (SAH) . Methylane (SAH) ,15. SAH ne (SAH) . Both thne (SAH) . Hcy is ne (SAH) ,19. Hcy cy thiol ,20. Then) to Cys ,21. Cys ) to Cys ,19.OCM metabolic markers have been linked to various diseases. Intracellular Hcy concentrations increase when OCM fails to function adequately ,22. The Previous studies have assessed folate concentrations in circulation using a microbiological assay ; howeverThe characteristics of the study population are summarized in 6, folate, and vitamin C intakes, but significantly lower sodium intake than the low 5-MTHF group.p = 0.059). However, tHcy and cystathionine concentrations and tCys/tHcy ratios were significantly lower in the high 5-MTHF group than in the low 5-MTHF group. Serum homocysteic acid, pyridoxamine, and pyridoxine concentrations were below the limit of quantification in all samples.The serum OCM-related metabolite concentrations in the high and low 5-MTHF groups and in the overall study sample are summarized in p < 0.001) than in the high 5-MTHF group . The positive correlation between DMG and tHcy concentrations was weaker in the low 5-MTHF group  than in the high 5-MTHF group . The correlation matrix between serum OCM-related metabolite concentrations and enzyme activity indices is summarized in p < 0.001) than in the high 5-MTHF group . The 5-MTHF concentrations and betaine/DMG ratios showed a significant positive correlation. The significant negative correlation between 5-MTHF concentrations and tHcy/tCys ratios  was stronger than the correlation between 5-MTHF and tHcy concentrations alone  than the low 5-MTHF group. On the other hand, the high 5-MTHF group exhibited higher energy and micronutrient intake than the low 5-MTHF group, suggesting a potential dietary influence on OCM. Because food nutrients are complex, it is likely that the intake of folate-rich foods, such as vegetables, altered the intake of other micronutrients. The intake of methionine, cystine, glycine, serine, zinc, riboflavin, and vitamin BHerein, the median serum 5-MTHF concentration in young Japanese women was 19.2 nmol/L, which was comparable to or slightly higher than that reported in previous studies involving healthy adults in countries where FA fortification of grains is not mandated, such as Japan ,64. HoweConcentrations of 5-MTHF exhibited a significant positive correlation with SAM and tCys concentrations and a significant negative correlation with tHcy concentrations. Similarly, a previous study involving older adults found a positive correlation between 5-MTHF and SAM concentrations, and FA intervention significantly increased plasma SAM concentrations in the intervention group compared with the control group . In addiThe negative correlation between serum 5-MTHF concentrations and tHcy/tCys ratios was stronger than that between 5-MTHF and tHcy concentrations alone . The tHcHerein, 5-MTHF concentrations exhibited significant positive correlations with betaine concentrations or betaine/DMG ratios . SimilarHerein, a negative association between serum 5-MTHF and cystathionine concentrations was observed . A negatSerum FA concentrations were not correlated with OCM-related metabolite concentrations associated with 5-MTHF concentrations. This suggests that the concentration of 5-MTHF, a bioactive folate molecule species, as a biomarker may be more useful than that of total folate, which was considered in previous studies .This study has several strengths. Early morning fasting blood samples were collected, and standardized blood collection protocols were used to minimize the following effects on OCM-related metabolites: changes in the blood concentration of the measured substance due to eating immediately before the blood sample is collected; errors related to human activities at the time of blood collection; and variation in the deterioration of the substance being measured. Furthermore, the serum concentrations of OCM-related metabolites were measured using the stable-isotope dilution mass spectrometry method with internal standards for all measured components, yielding high-quality quantitative results .12-dependent reaction, and vitamin B12 is an important determinant of Hcy [6 intake may have promoted transsulfuration pathway flux. Finally, single-nucleotide polymorphisms may have an impact on OCM [However, this study has several limitations. First, given the cross-sectional survey design, it is impossible to establish a causal association between OCM metabolic dynamics. Further intervention studies on FA or 5-MTHF are required. Second, serum OCM-related metabolites do not always directly reflect OCM dynamics in organ cells. Third, 5-MTHF-dependent Hcy remethylation is a vitamin Bt of Hcy ,107; howt of Hcy ; howevert of Hcy ,109. In t on OCM ; howeverThis study was conducted at Kagawa Nutrition University, Saitama, Japan, between October and December 2018. This was a cross-sectional study designed to investigate the association between diet and blood components such as biochemical test values, OCM-related metabolites, fatty acids, and antioxidants; we reported associations between serum OCM-related metabolites.Healthy female university students aged 18\u201325 years majoring in nutrition were included. The following students were excluded: (1) those with health conditions that may affect the biomarker concentration; (2) those with a history of, or who have serious hepatic, renal, cardiac, pulmonary, gastrointestinal (including gastrectomy), or organ disorders; diabetes; food allergies; or other serious diseases; (3) those receiving medicines that can affect lipid metabolism or FA metabolism or anti-inflammatory and antioxidant drugs that may affect the measured values in this study; (4) those pregnant and lactating or planning for pregnancy and lactation; (5) those with systolic blood pressure <90 mmHg; (6) those who had previously donated large amounts of blood; (7) those participating in other clinical trials or studies or within 4 weeks of the end of those studies; (8) those with BMI >25 because BMI may affect blood SAM and SAH levels ,111; andn = 9), lost contact (n = 6), health problems (n = 1), busyness (n = 4), and withdrawals (n = 11), leaving 227 participants. The participants completed a lifestyle questionnaire and underwent physical measurements, and their fasting blood samples were collected after at least 10 h of fasting.A total of 258 eligible participants consented to the study, including proxy consent for minors. After the study began, 31 participants dropped out owing to consent withdrawals (The study protocol was approved by the Ethics Committee of Kagawa Nutrition University (protocol code no. 204) and was conducted in accordance with the Helsinki Declaration. All participants provided written informed consent before participating in the study.g, and the serum was then separated within an hour and frozen at \u221280 \u00b0C. Under these storage conditions, the concentrations of OCM-related metabolites measured in this study have been reported to be generally stable [To minimize dietary effects ,67,112, y stable ,113,114.m/z 460.2\u2192313.2 (5-MTHF), m/z 465.2\u2192313.2 (5-MTHF\u221213C5), m/z 442.2\u2192295.2 (FA), m/z 447.2\u2192295.2 (FA\u221213C5), m/z 104.0\u219260.1 (choline), m/z 113.0\u219269.1 (choline-d9), m/z 118.1\u219258.0 (betaine), m/z 129.0\u219266.1 (betaine-d11), m/z 104.1\u219258.1 (DMG), m/z 110.1\u219264.0 (DMG-d6), m/z 150.1\u2192104.0 (methionine), m/z 153.0\u2192107.0 (methionine-d3), m/z 399.1\u2192250.3 (SAM), m/z 402.2\u2192250.2 (SAM-d3), m/z 385.2\u2192134 (SAH), m/z 389.2\u2192136.2 (SAH-d4), m/z 136.0\u219290.0 (Hcy), m/z 140.0\u219293.9 (Hcy-d4), m/z 184.1\u2192138.3 (homocysteic acid), m/z 188.1\u2192142.0 (homocysteic acid-d4), m/z 223.2\u2192134.1 (cystathionine), m/z 227.1\u2192138.1 (cystathionine-d4), m/z 122.1\u219259.1 (Cys), m/z 124.2\u219261.0 (Cys-d2), m/z 126.1\u2192107.8 (taurine), m/z 128.1\u2192110.2 (taurine\u221213C2), m/z 106.1\u219260.1 (serine), m/z 109.1\u219263.1 (serine-d3), m/z 76.0\u219230.0 (glycine), m/z 78.0\u219231.9 (glycine-d2), m/z 377.1\u2192243.1 (riboflavin), m/z 383.2\u2192249.1 (riboflavin\u221213C415N2), m/z 169.2\u2192152.2 (pyridoxamine), m/z 172.1\u2192155.1 (pyridoxamine-d3), m/z 170.1\u2192134.1 (pyridoxine), and m/z 172.1\u2192136.0 (pyridoxine-d2)) [We previously described the method for measuring OCM-related metabolites . After cine-d2)) . Hcy andine-d2)) . AnalystData regarding the participants\u2019 ages were collected using a self-administered questionnaire.Dietary data were collected for 7 days before blood collection using a continuous, non-weighted dietary record in conjunction with digital images captured using a digital camera or smartphone according to a validated dietary survey method ,118,119.Blood pressure was measured using a P2000 Electronic Blood Pressure Monitor . The mean systolic and diastolic blood pressure was measured twice in the resting state according to the Japanese guidelines for managing hypertension .The participants\u2019 anthropometric measurements were collected using standardized methods during laboratory visits for blood sampling. Body weight and body fat percentage were measured using TBF-110 .U test was used to compare these groups. The enzyme activity indices included the betaine/DMG ratio [p-value of <0.05. IBM Statistical Package for the Social Sciences Statistics, Version 28  was used for statistical analysis.The distribution of subject characteristics data, nutrient intake, and serum concentrations of OCM-related metabolites used in the analysis were mostly non-normal and continuous variables were expressed as medians . Since there are no thresholds established for serum 5-MTHF concentrations such as deficiency, insufficiency, sufficiency, or excess, based on a previous study , a mediaMG ratio  for BHMTMG ratio  for methMG ratio  for enzyPrevious studies assessed folate status without considering folate molecular species. Thus, we extended the findings of previous studies by comprehensively measuring OCM-related metabolites, including 5-MTHF, a major reduced folate. FA showed no association with important OCM-related metabolites, indicating that blood 5-MTHF levels could be useful for evaluating folate status. The 5-MTHF state could be linked to the Hcy flux into the transsulfuration pathway via SAM. Therefore, the tHcy/tCys ratio may be a more sensitive indicator of the 5-MTHF status than tHcy concentrations alone. A low 5-MTHF status may affect the choline metabolic pathway, particularly BHMT or CBS activity via betaine, implying that the folate cycle and choline metabolic pathway should be evaluated simultaneously in Hcy studies. Further intervention studies on FA or 5-MTHF are warranted to determine the causal association between the abovementioned findings."}
+{"text": "Correction: Journal of Translational Medicine (2013) 11:29 10.1186/1479-5876-11-29Since the publication of our article , it has https://jeccr.biomedcentral.com/articles/10.1186/1756-9966-27-15. Copyright \u00a9 2008, Liao et al.; licensee BioMed Central Ltd.(A) The levels of CSE1L expression in B16-EV, B16-CSE1L, COLO-EV, and COLO-CSE1L cells were assayed by immunoblotting with anti-CSE1L antibody. The \u03b2-actin levels were assayed as a control. The invasive ability of the cells was analyzed by in vitro invasion assays using chemotaxis chambers, as described in \u201cMaterials and Methods\u201d. The left panel is adapted with permission from Fig.\u00a05C in Liao CF, Luo SF, Li LT, Lin CY, Chen YC, Jiang MC. CSE1L/CAS, the cellular apoptosis susceptibility protein, enhances invasion and metastasis but not proliferation of cancer cells. J Exp Clin Cancer Res. 2008; 27:15."}
+{"text": "Infections after LT are common, but standard of care testing (SOCT) often fails to identify a cause. We prospectively compared longitudinal mcfDNA sequencing vs SOCT to determine the utility of mcfDNA sequencing for infection surveillance after LT.Single center prospective observational study of new LT patients (pts). We collected plasma on days 0, 1, 2, 3, 7, 14, and 30 post LT for mcfDNA sequencing; results were not available clinically. We reviewed outcomes and SOCT through 6 months (mo).Candida, oral or gastrointestinal commensals, Enterococci, or obligate pathogens ; most had polymicrobial mcfDNA sequencing results. In 36.67% (11/30), mcfDNA sequencing detected pathogens of unclear significance . In 8 pts with positive SOCT, mcfDNA sequencing did not detect at least 1 pathogen found on SOCT. In 13% (4/30), mcfDNA sequencing detected a pathogen before it was detected by SOCT (n=1 each): VRE surgical site infection (SSI), invasive candidiasis, Enterobacter pneumonia, and M. hominis SSI (a known donor-derived syndrome) with HHV-8/Kaposi Sarcoma .We enrolled 30 LT pts . 93.3% (28/30) received broad-spectrum antibiotics, although only 40.0% (12/30) had SOCT yielding pathogenic bacteria. 43.33% (13/30) had unexplained leukocytosis. Table 2 shows serial mcfDNA sequencing results, clinical course, and adjudication of mcfDNA sequencing. 96% (29/30) had at least 1 positive mcfDNA sequencing test, including 68.75% (11/16) with presumed infection or unexplained leukocytosis and negative SOCT, but diversity of organisms on mcfDNA sequencing, e.g., Table 1.Table 2 legend.Table 2.Plasma mcfDNA sequencing revealed potential occult explanations for unexplained leukocytosis and infectious syndromes with negative SOCT early post LT. mcfDNA sequencing also detected infection earlier than SOCT in 4 pts but did not detect known pathogens from 8 pts; > 1/3 had presumed FP mcfDNA sequencing results. Larger studies with longer follow-up are needed to determine the utility of mcfDNA sequencing in diagnosing infection, optimizing antibiotic use, and monitoring donor-derived infection post LT.Ghady Haidar, MD, Allovir: Grant/Research Support|AstraZeneca: Advisor/Consultant|AstraZeneca: Grant/Research Support|Karius: Advisor/Consultant|Karius: Grant/Research Support|NIH: Grant/Research Support Timothy A. Blauwkamp, PhD, Karius: Board Member|Karius: Ownership Interest Georgios Kitsios, MD, PhD, Karius, Inc: Grant/Research Support"}
+{"text": "This study reports on the E. coli and 92 S. saprophyticus isolates were collected during 2022 from 45 medical centers located within the United States. Most isolates (77%) tested were cultured from urine specimens collected from patients seen in ambulatory, emergency, family practice, and outpatient medical services. Bacterial identifications were confirmed by MALDI-TOF MS. Isolates were tested for susceptibility by CLSI methods at a central laboratory (JMI Laboratories). MIC results for oral antibiotics licensed for the treatment of uUTI and drug-resistant subsets were interpreted per CLSI guidelines.A total of 1,001 50/90, 2/4 \u00b5g/mL) displayed good activity against 1,001 E. coli isolates as 98.8% of all observed gepotidacin MICs were \u22644 \u00b5g/mL (Table). Other oral agents tested against these isolates demonstrated the following rates of susceptibility (S): amoxicillin-clavulanate (85.2% S), ampicillin (55.6% S), ciprofloxacin (78.8%S), fosfomycin (99.3% S), mecillinam (93.5%S), nitrofurantoin (97.5% S), and trimethoprim-sulfamethoxazole (73.3% S). Gepotidacin maintained similar MIC50 values (ranging from 1 \u2013 2 \u00b5g/mL) and MIC90 values (ranging from 2 \u2013 4 \u00b5g/mL) against these drug-resistant subsets. Against S. saprophyticus isolates, gepotidacin  inhibited all isolates at \u22640.12 \u00b5g/mL. Most oral agents showed >90% S against S. saprophyticus isolates, except for penicillin (7.6% S).Gepotidacin (MICin vitro activity against contemporary E. coli and S. saprophyticus from US urine isolates. This activity remained unaffected by resistance to other oral standard-of-care antibiotics.Gepotidacin demonstrated S J Ryan Arends, PhD, Cipla: Grant/Research Support|GSK: Grant/Research Support Renuka Kapoor, PhD, GSK: Grant/Research Support Didem Torumkuney, PhD, GSK: Grant/Research Support Nicole E. Scangarella-Oman, MS, GSK: Employee and shareholder Rodrigo E. Mendes, PhD, AbbVie: Grant/Research Support|Basilea: Grant/Research Support|Cipla: Grant/Research Support|Entasis: Grant/Research Support|GSK: Grant/Research Support|Paratek: Grant/Research Support|Pfizer: Grant/Research Support|Shionogi: Grant/Research Support"}
+{"text": "Acinetobacter spp., including carbapenem-resistant Acinetobacter baumannii-calcoaceticus complex (ABC) organisms. In this study, the susceptibility testing of zosurabalpin was carried out against a panel of 150 randomly selected Acinetobacter spp. isolates (100 A. baumannii & 50 non-A. baumannii) representing 11 sites in China and a broad susceptibility profile (65% of which were multi-drug resistant [MDR]) collected in 2021.Zosurabalpin (RG6006) is the first representative of a novel class of tethered macrocyclic peptide antibiotics active against MICs were performed using the Clinical Laboratory Standards Institute (CLSI) broth dilution method cation-adjusted Mueller Hinton broth (CA-MHB) supplemented with either 20% of human serum (HS) or 20% of horse serum (HoS). For a fraction of isolates (10-25%), MIC determination for zosurabalpin is affected by aberrant readings  in CA-MHB. This effect complicates routine susceptibility testing. Supplementation of CAMHB with serum allows accurate MIC determinations without any major effects on the MIC distribution.Summary data from the study are shown in Table 1.Acinetobacter spp., with an MIC50/90 of 0.12/0.5 \u03bcg/mL and 0.25/1 \u03bcg/mL in CA-MHB supplemented with HoS and HS, respectively (MIC range of 0.015/0.03 to 8 \u03bcg/mL). Against ABC isolates (n=133), the MIC50/90 for zosurabalpin was 0.12/0.25 \u03bcg/mL and 0.25/0.5 \u03bcg/mL, in HoS and HS, respectively (MIC range of 0.015/0.03 to 1 \u03bcg/mL). A similar activity was observed against carbapenem-resistant ABC isolates.Zosurabalpin was active against all in vitro antibacterial activity against Acinetobacter clinical isolates circulating in China. These data support the continued clinical development of zosurabalpin for infections caused by ABC isolates, including difficult to treat carbapenem-resistant isolates.In addition to the potent activity observed against isolates from USA and Europe, zosurabalpin exhibited potent Stephen Hawser, PhD, Allecra: study funding|Innoviva Specialty Therapeutics, Inc.: Honoraria|Roche: Honoraria|Roche: This project has been funded by BARDA (HHSO100201600038C). Nimmi Kothari, PhD, Allecra: Allecra (study funding)|Innoviva Specialty Therapeutics, Inc.: Honoraria|Roche: Honoraria|Roche: This project has been funded by BARDA (HHSO100201600038C). Thomas Valmont, BS, Roche: Honoraria|Roche: This project has been funded by BARDA (HHSO100201600038C). S\u00e9verine Louvel, PhD, F. Hoffmann-La Roche Ltd: employee of the company Claudia Zampaloni, n/a, F. Hoffmann-La Roche Ltd.: Full time employee of Roche"}
+{"text": "Detection of SARS-CoV-2 in hospitalized patients is an important measure to prevent transmission to vulnerable individuals. Admission screening using nucleic acid amplification test such as PCR testing or rapid antigen testing (RAT) is proven to be beneficial in certain situations. However, the usefulness of SARS-CoV-2 antibody testing for COVID-19 screening in real-world setting is unclear.In this study, we aimed to investigate the effect of antibody testing on the results of SARS-CoV-2 screening tests. Anti-SARS-CoV-2 spike protein antibody (S-Ab) and anti-SARS-CoV-2 nucleocapsid protein antibody (N-Ab) were measured using an electrochemiluminescence immunoassay  in all patients who presented to the emergency department and subsequently administered to Nagasaki University Hospital. PCR testing and RAT were also performed for the detection of SARS-CoV-2. The diagnostic performance of each antibody testing to PCR and RAT was retrospectively evaluated by Fisher's exact test and odds ratio (OR).A total of 2,692 patients were tested for S-Ab, N-Ab, RT-PCR, and RAT from Jan 2022 to Feb 2023. Among them, S-Ab were positive ( >0.7 U/mL) for 2,195 patients (81.5%), N-Ab ( >1.0 U/mL) 307 patients (11.4%), PCR 215 patients (7.9%), RAT 174 patients (6.5%).P=0.35, OR 0.8397) and RAT 1.8% . For those with N-Ab positive (n=307), PCR positivity was 6.8%  and RAT 2.6% . Comparing the group of both S-Ab > 10,000 U/mL and N-Ab positive (n=128) with that of both negative (n=2564), PCR positivity was 4.7% vs. 8.1% , and RAT positivity was 1.6% vs. 6.7% .For those with S-Ab > 10,000 U/mL (n=625), PCR positivity was 7.0% (Antibody testing of S-Ab, N-Ab, or a combination of both showed performance in reducing the probability of positive PCR or RAT tests, indicating vaccine and infection-induced protective effect. However, the need for PCR and RAT tests for virus detection was suggested.Kosuke Kosai, M.D., Ph.D., FUJIFILM Toyama Chemical Co., Ltd.: Commissioned research|KYORIN Pharmaceutical Co.,Ltd.: Commissioned research Koichi Izumikawa, M.D., Ph.D., Asahi Kasei Pharma Corporation: Grant/Research Support|Asahi Kasei Pharma Corporation: Honoraria|Astellas Pharma Inc.: Honoraria|DAIICHI SANKYO COMPANY, LIMITED: Grant/Research Support|DAIICHI SANKYO COMPANY, LIMITED: Honoraria|KYORIN Pharmaceutical Co.,Ltd.: Honoraria|Merck & Co., Inc.: Honoraria|Pfizer Japan Inc.: Honoraria|Shionogi & Co., Ltd.: Grant/Research Support|Shionogi & Co., Ltd.: Honoraria|Sumitomo Pharma Co., Ltd.: Grant/Research Support|Sumitomo Pharma Co., Ltd.: Honoraria|TAIHO PHARMACEUTICAL CO., LTD.: Grant/Research Support Katsunori Yanagihara, MD, PhD, FUJIFILM Toyama Chemical Co., Ltd.: Commissioned research|KYORIN Pharmaceutical Co.,Ltd.: Commissioned research"}
+{"text": "Artemisia heptapotamica Poljak led to the isolation of ten known compounds, including four alkyl p-coumarates: octadecyl trans-p-coumarate (1), icosy trans-p-coumarate (2), docosyl trans-p-coumarate (3), and tetracosyl trans-p-coumarate (4), one sesquiterpene lactone: santonin (5), four flavonoids; axillarin (6), quercetin 3-O-methyl ether (7), luteolin (8), and quercetin (9), and one phenolic acid derivative: p-coumaric acid (10). The structures of the isolated compounds were identified by various spectroscopic analyses. Additionally, the antimicrobial activity of the total extract and different fractions was screened, and they exhibited no inhibition of the growth of Candida albicans, C. neoformans, Aspergillus fumigatus, methicillin-resistant Staphylococcus aureus (MRS), E. coli, Pseudomonas aeruginosa, Klebsiella pneumonia, and Vancomycin-resistant Enterococci (VRE) at the tested concentrations ranging from 8 to 200 \u03bcg/mL. The identification and tentative characterization of the secondary metabolites were conducted using LC-QToF analysis. This method helps in the putative characterization of sesquiterpene lactones, flavonoids, coumarate derivatives, and aliphatic compounds. The developed method identified 43 compounds, of which the majority were sesquiterpene lactones, such as eudesmanolides, germacranolides, and guaianolide derivatives, followed by flavonoids. The proposed LC-QToF method helps develop dereplication strategies and understand the major class of chemicals before proceeding with the isolation of compounds.Phytochemical investigation of the aerial parts of  Artemisia L. is the largest genus belonging to the family Asteraceae + and [M + Na]+ adduct precursor ions. The tentative characterization of compounds was processed based on the molecular features, such as accurate mass, fragment ions , and precursor ion molecular formula. The representative base peak chromatograms (BPC) in negative and positive modes, along with LC-DAD profiles at 210 nm, are shown in The secondary metabolites from aerial parts of Artemisia species. To perform characterization using LC-QToF, a few sesquiterpene lactones, e.g., santonin (compound 19), were isolated from aerial parts of A. heptapotamica in this study. Santonin mass fragmentation initiated with loss of water molecules, resulting in m/z 228.1219 [M + H-H2O]+. The following loss of -CO and sequential loss of -C2H4 and -CH4 resulted in m/z 201.1271, 173.0954, and 157.0648, respectively. Further loss of -CO ion resulted in the opening of the lactone moiety. Further loss of ketene moiety (-C2H2O) from the opened six-membered ring resulted in the formation of m/z 115.0542. Fragments with the least molecular weight (m/z 105.0698 and 91.0544) were useful in determining the backbone skeleton of the molecule. These characteristic fragments matched with the reported santonin fragmentation pathway \u2212 and fragment ions are shown in 25 showed m/z 509.1284 [M + H]+ with a C23H24O13 molecular formula. The corresponding fragment ions resulted in m/z 347.0762 [M + H-Glc]+, 331.0434, and 289.0329, respectively. The compound was characterized as 3,4,5,7-tetrahydroxy-3-methoxyflavone 7-O-\u03b2-D-glucopyranoside. Similarly, the flavone derivatives were characterized based on their fragmentation pathways. Along with hydroxy/methoxy flavone derivatives, simple aglycone flavonoids such as quercetin (compound 27) and luteolin (compound 26) were identified. Quercetin showed m/z 303.0491 [M + H]+ and m/z 301.0354[M-H]\u2212 with fragment ions at m/z 153 and 151, respectively. Furthermore, luteolin showed m/z 287.0548 [M + H]+ and 285.0406 [M-H]\u2212 in both positive and negative ionization modes. In addition, one quercetin derivative (compound 33) was identified with m/z 317.0658 [M + H]+ with fragment ions at m/z 301.0343 (such as quercetin aglycone), 274.0465, and 137.0233. The compound was characterized as quercetin 3-O-methyl ether.Using the isolated reference compounds and exact mass measurements, fourteen flavonoid compounds were identified. Most of the compounds are flavone derivatives. There are two quercetin derivatives, along with quercetin and luteolin. Flavonoids have established mass fragmentation patterns based on the literature previously reported ,23,24,25m/z 165.0546 [M + H]+ and 163.0398 [M-H]\u2212 with fragment ions at m/z 119.0502 [M-H-CO2]\u2212 in the negative mode of ionization. The compound was characterized as p-coumaric acid (compound 37). In addition, an aliphatic compound was identified with chemical formula C10H18O3. Compound 38 was tentatively characterized based on the exact mass at m/z 185.1186 [M-H]\u2212 and corresponding fragment ion at m/z 167.1077 [M-H-H2O]\u2212. Compound 39 was identified as 3,4,5-tri-caffeoylquinic acid with a precursor ion at m/z 677.1524 [M-H]\u2212. The corresponding fragment ions were at m/z 515.1198 [M-H-Glc]\u2212, 353.0822 [M-H-Glc-Glc]\u2212, and 191.0572 [quinic acid]  , respectArtemisia species.The isolated compounds, as well as chromatographic peaks with distinctive fragment ions, were confirmed based on the reported literature studies whose corresponding references are listed in 1H and 13C NMR spectra were recorded on a Bruker Avance 400 MHz instrument. HR-ESI-MS was taken on BrukerBioApex-FTMS with electron spray ionization. Solvents used in this work, e.g., n-hexane, dichloromethane (DCM), ethyl acetate (EtOAc), methanol (MeOH), and ethanol (EtOH), were purchased from Fisher Scientific, USA. Deuterated solvents purchased from Cambridge Isotope Laboratories, Inc., Tewksbury, MA, USA, including methanol-d4 (CD3OD), chloroform-d3 (CDCl3), and pyridine-d5 (C5H5N-d5), were used for nuclear magnetic resonance (NMR) spectroscopic analyses. Acetonitrile, methanol, and formic acid of HPLC-certified grade were used for LC-MS analysis, and water was purified using a Milli-Q system . Column chromatography (CC) was performed using silica gel 60 . Thin-layer chromatography (TLC) analyses were carried out using pre-coated silica G plates w/UV254 . An ultraviolet lamp  was used for visualization of spots on thin-layer chromatograms at 254 and/or 365 nm. Spots were visualized by spraying with 2% vanillin  in sulfuric acid\u2013ethanol followed by heating at 110 \u00b0C.A. heptapotamica was collected in October 2021 from Kokpek village, Almaty, Kazakhstan. Identification and authentication were performed by Dr. Danilov Mikhail Petrovich. The sample was stored with the voucher number 0000723 in the Main Botanical Garden of the Institute of Botany and Phyto-introduction, Almaty, Kazakhstan. The same sample was assigned with the NCNPR number 25173 and stored in the National Center for Natural Product Research Botanical Repository, University of Mississippi, USA.The whole plant of A. heptapotamica were first dried in the shade and then crushed into small pieces. The dried plant material (1.65 kg) was extracted by maceration with 95% methanol three times at room temperature and was concentrated under reduced pressure to yield 253.35 g of the total extract. The total extract was mixed with a small amount of distilled water and successively fractionated with n-hexane and ethyl acetate. The fractions were concentrated under reduced pressure to produce n-hexane (29.1 g) and ethyl acetate fractions (58.74 g).The collected aerial parts of n-hexane and EtOAc  and finally washed with MeOH, affording 6 fractions. Fraction F-4 (395.9 mg) was subjected to silica gel CC , using DCM\u2013MeOH gradient mixtures to increase the polarity gradually in 2% MeOH till 80 % MeOH and finally 100% MeOH were produced, yielding 37 subfractions. The seventeenth subfraction (21.1 mg) afforded a mixture of compounds 3 and 4. The eighteenth subfraction (12.9 mg) produced a mixture of compounds 2\u20134. The nineteenth subfraction (16.5 mg) produced a mixture of compounds 2\u20134. The twentieth subfraction (6.2 mg) furnished a mixture of compounds 1\u20134.The EtOAc (58.74 g) fraction was subjected to fractionation using VLC silica gel CC using 5 (45 mg). The eighteenth subfraction was precipitated to give compound 6 (27 mg). Subfraction F-5-21 (1.26 g) was purified on silica gel CC using DCM\u2013MeOH gradient mixtures to gradually increase the polarity in 2% MeOH till 80% MeOH was produced, affording compound 7 (8.4 mg). Subfraction F-5-24 (342.8 mg) was rechromatographed over silica gel CC using DCM\u2013MeOH gradient mixtures to gradually increase the polarity in 2% MeOH till 80% MeOH was reached, producing a mixture of compounds 7 and 8 (14.5 mg) and 9 (24.4 mg). Subfraction F-5-26 (450.1) was purified on silica gel CC using DCM\u2013MeOH gradient mixtures to gradually increase the polarity in 2% MeOH till 80% MeOH was reached, producing compound 10 (8.2 mg).Fraction F-5 (14.3 g) was subjected to silica gel CC , using DCM\u2013MeOH gradient mixtures to gradually increase the polarity in 2% MeOH till 80% and finally 100% MeOH were produced, affording 29 subfractions. The eleventh subfraction was crystalized to produce compound The antimicrobial activity of the total extract and different fractions was evaluated using the method reported by Samy et al. .About 25 mg of extract was sonicated in 1.0 mL of methanol for 5 min, followed by centrifugation for 15 min at 7000 rpm. The clear filtered supernatant solution was used for analysis.TM HSS C18 column . The mobile phase consisted of water with 0.1% formic acid (A) and acetonitrile with 0.1% formic acid (B) at a flow rate of 0.23 mL/min. Analysis was performed using the following gradient elution: 15% B to 40% B in 30 min, then to 100% B in the next 15 min. Each run was followed by a 5 min wash with 100% B and an equilibration period of 15 min with 85% A/15% B. Two microliters of the sample were injected. The column temperature was 40 \u00b0C.The liquid chromatographic system was an Agilent Series 1290, and separation was achieved on an Acquity UPLC2) flow rate, 13 L/min; drying gas temperature, 325 \u00b0C; nebulizer pressure, 30 psi; sheath gas temperature, 300 \u00b0C; sheath gas flow, 11 L/min; capillary voltage, 3500 V; nozzle voltage, 0 V; skimmer, 65 V; Oct RF V, 750 V; and fragmentor voltage, 125 V. All the operations, acquisition of data, and analysis of data were controlled using Agilent MassHunter Acquisition Software ver. A.10.1 and processed with MassHunter Qualitative Analysis software ver. B.07.00. Each sample was analyzed in positive and negative modes over the range of m/z 50\u20131700 and an extended dynamic range. Accurate mass measurements were obtained by employing ion correction techniques using reference masses at m/z 121.0509 (protonated purine) and 922.0098  in positive ion mode, while m/z 112.9856 (deprotonated trifluoroacetic acid-TFA) and 1033.9881 (TFA adducted HP-921) were used in negative ion mode. Samples were analyzed in all-ion MS\u2013MS mode, where experiment 1 was carried out with a collision energy of zero and experiment 2 with a fixed collision energy of 45 eV.The mass spectrometric analysis was performed with a QToF-MS-MS  equipped with an ESI source with Jet Stream technology, using the following parameters: drying gas , which matched with the authentic samples of A. heptapotamica.Ten compounds were isolated from the ethyl acetate fraction of"}
+{"text": "Immune responses after COVID-19 vaccination should be evaluated in different populations around the world. This study compared antibody responses induced by ChAdOx1 nCoV-19, CoronaVac, and BNT162b2 vaccines. Blood samples from vaccinees were collected pre- and post-vaccinations with the second and third doses. The study enrolled 78 vaccinees, of whom 62.8% were women, with the following median ages: 26 years\u2014ChAdOx1 nCoV-19; 40 years\u2014CoronaVac; 30 years\u2014BNT162b2. Serum samples were quantified for anti-RBD IgG and anti-RBD IgA and anti-spike IgG by ELISA. After two vaccine doses, BNT162b2 vaccinees produced higher levels of anti-RBD IgA and IgG, and anti-spike IgG compared to ChAdOx1 nCoV-19 and CoronaVac vaccinees. The third dose booster with BNT162b2 induced higher levels of anti-RBD IgA and IgG, and anti-spike IgG in CoronaVac vaccinees. Individuals who reported a SARS-CoV-2 infection before or during the study had higher anti-RBD IgA and IgG production. In conclusion, two doses of the studied vaccines induced detectable levels of anti-RBD IgA and IgG and anti-spike IgG in vaccinees. The heterologous booster with BNT162b2 increased anti-RBD IgA and IgG and anti-spike IgG levels in CoronaVac vaccinees and anti-RBD IgA levels in ChAdOx1 nCoV-19 vaccinees. Furthermore, SARS-CoV-2 infection induced higher anti-RBD IgA and IgG levels in CoronaVac vaccinees. Several preventive strategies have been developed and continue to be generated to control SARS-CoV-2 transmission, the most important being vaccines . CurrentMost vaccines developed against SARS-CoV-2 induce an immune response toward the receptor-binding domain (RBD), a portion of the spike protein present at the S1 region, which is essential for binding the virus to angiotensin-converting enzyme-2 (ACE2) on the cell surface and to establish cellular infection . Thus, aAfter COVID-19 vaccination, IgG and IgA are mostly involved in virus neutralization, especially IgA since this Ig can neutralize SARS-CoV-2 at the mucosal surface entrance . Some stThe anti-SARS-CoV-2 vaccines comprise platforms of inactivated viruses , proteinThe present study was evaluated and approved by the Research Ethics Committee of the Hospital das Cl\u00ednicas, Universidade Federal de Goi\u00e1s, under the number CAAE: 30804220.2.0000.5078 and is in accordance with Resolution 466/2012 of the National Health Council, which regulates research involving human beings. The collection of data and blood samples was conducted after the individuals selected for the sample had agreed to participate and had signed the informed consent form.Blood samples were collected from 78 volunteers before immunization (ChAdOx1 nCoV-19 and BNT162b2) and after the administration of the second doses of ChAdOx1 nCoV-19, CoronaVac, and BNT162b2 vaccines, and after the third dose (booster) with BNT162b2 vaccine. In addition, nasopharyngeal swab samples were collected from individuals belonging to the ChAdOx1 nCoV-19 and BNT162b2 vaccine groups at the pre-vaccine collection and 1 month after the second dose to assess whether these individuals could be infected with SARS-CoV-2 during the period.Participants were recruited by phone or in person while in the queue for vaccination, and during the vaccination of health professionals at the Instituto de Patologia Tropical e Sa\u00fade P\u00fablica (IPTSP), Universidade Federal de Goi\u00e1s (UFG), Goi\u00e2nia-GO, Brazil. The inclusion criteria were individuals at least 18 years old, vaccinated with two doses of CoronaVac, ChAdOx1 nCoV-19, or BNT162b2, and those who may have received the third dose, but not necessarily.Primary pre-vaccine samples were collected at the queues for vaccination, in a physical space adjacent to the line, right before the first dose was administered. Post-vaccination samples were collected in three different periods: 1 month and 4\u20136 months after the second dose, and 1 month after the third dose. Samples were collected 1 month after the second and third doses because the peak of the antibodies occurs around four weeks after vaccination ,30,31,32\u00ae Viral RNA Mini Kit  was used, following the manufacturer\u2019s protocol. Samples were submitted to real-time polymerase chain reaction post reverse transcription (RT-qPCR) after the RNA extraction, using the Promega GoTaq\u00ae Probe 1-Step RT-qPCR System, according to the manufacturer\u2019s protocol [For Ribonucleic acid (RNA) extraction, the commercial QIAampprotocol . The priprotocol  was usedg for 10 min at room temperature. All samples were processed on the day of collection. Serum was separated and stored in 2 mL polypropylene cryotubes in a \u221280 \u00b0C freezer for subsequent enzyme immunoassay ELISAs.After blood collection from vaccinated individuals, whole blood was centrifuged at 600\u00d7 For the ELISAs, 96-well high-binding polystyrene half-area plates  were coated with 50 \u00b5L per well with the RBD protein, expressed according to Amanat et al. (2020) , at a co2 > 0.5. For all tests, p < 0.05 was considered significant.All analyses of individual samples were conducted using GraphPad Prism version 9 . The variables distribution patterns were evaluated via the Kolmogorov\u2013Smirnov and Shapiro\u2013Wilk tests. For paired comparisons between groups, the non-parametric Wilcoxon Matched-Pairs Signed Rank test was used. Paired analyses were assigned to samples from the same individuals at different collection times. Unpaired groups were analyzed with the non-parametric Mann\u2013Whitney U test. The non-paired analyses were attributed to a total group. For frequency calculations, we used the Fisher exact test. Multiple group comparisons were analyzed by running non-parametric Kruskal\u2013Wallis statistical tests and were corrected using Dunn\u2019s and Dunnett\u2019s methods. Spearman correlation test was used for association analyses. The existence of correlation was adopted for rn = 78) were divided into three groups according to the vaccines they had received: ChAdOx1 nCoV-19 (n = 33), BNT162b2 (n = 27), and CoronaVac (n = 18). The ChAdOx1 nCoV-19 group was composed of 33 individuals, 23 (69.7%) women and 10 (30.3%) men. The BNT162b2 group was composed of 27 individuals, 11 (40.7%) women and 16 (59.3%) men. Finally, the CoronaVac group was made up of 18 participants, 15 (83.3%) women and 3 (16.7%) men (Study participants (.7%) men . Therefo.7%) men . ParticiWhole blood collection periods comprise T1 (pre-vaccine), T2 (1 month post-second dose), T3 (4\u20136 months post-second dose or pre-third dose), and T4 (1 month post-third dose or booster dose). All participants who had their samples collected at T4 received the booster dose, mostly BNT162b2. Of 78 participants, 56 received the third dose. This information can be better observed in Only the pre-vaccine nasopharyngeal samples from the ChAdOx1 nCoV-19 group were tested via reverse transcription followed by the real-time polymerase chain reaction (RT-qPCR), and all samples were negative for SARS-CoV-2 RNA. In addition, a rapid test assay to detect IgM/IgG  was performed on all serum samples collected , and only a few samples were positive. Some of them belonged to individuals who reported SARS-CoV-2 infection during the application of the questionnaire. Therefore, we believe that the rapid test positivity was due to a previous SARS-CoV-2 infection.p = 0.0027, median\u2014pre-vaccine: 0.6250, post-vaccine: 0.8310; anti-RBD IgG, p < 0.0001, median\u2014pre-vaccine: 0.8990, post-vaccine: 10.16; anti-spike IgG, p < 0.0001, median\u2014pre-vaccine: 1.549, post-vaccine: 10.75; BNT162b2: anti-RBD IgA, p < 0.0001, median\u2014pre-vaccine: 0.6460, post-vaccine: 4.023; anti-RBD IgG, p < 0.0001, median\u2014pre-vaccine: 0.9560, post-vaccine: 11.80; anti-spike IgG, p < 0.0001, median\u2014pre-vaccine: 1.254, post-vaccine: 11.22   A,D,G.p < 0.0001, median\u2014ChAdOx1 nCoV-19: 0.7890; BNT162b2: 3.731; IgG, p = 0.0268, median\u2014ChAdOx1 nCoV-19: 9.991; BNT162b2: 10.87) and CoronaVac . Individuals immunized with CoronaVac exhibited lower anti-RBD IgG levels compared to those with ChAdOx1 nCoV-19   and CoronaVac   E. This w: 11.28) H.p < 0.0001, median\u2014ChAdOx1 nCoV-19: 1.634; BNT162b2: 6.217) and IgG , and in comparison, to CoronaVac, for anti-RBD IgA . In addition, comparisons made in relation to the previous collection period (4\u20136 months after the second dose or before the third dose) showed differences between anti-RBD IgA and IgG levels, after the booster dose, for the ChAdOx1 nCoV-19    C and Corr: 1.925 C; anti-Rr: 11.07 F), with : 10.80) I.p < 0.0001). Individuals administered with BNT162b2 showed a higher frequency of anti-RBD IgA production compared to those administered with ChAdOx1 nCoV-19 (p < 0.0001) and CoronaVac (p < 0.0001) at 4\u20136 months after the second dose (p = 0.0030) (All study vaccinees (100%) produced detectable levels of anti-RBD and anti-spike IgG after each vaccine dose. The frequency of individuals who produced anti-RBD IgA at post-vaccination collection times is shown in ond dose B. At 1 m 0.0030) C.p < 0.0001; T1 vs. T3, p = 0.0028, median\u2014T1:0.6250, T2: 0.8310, T3: 0.7890, and T4: 1.634)   (p < 0.0001) for anti-RBD IgA, and anti-spike IgG . The longitudinal analysis performed for BNT162b2 (p = 0.0313). For the CoronaVac group    , T2 (1 month post second dose vaccine), T3 (4\u20136 months post second dose vaccine), and T4 (1 month post third dose vaccine). The results of this evaluation for the ChAdOx1 nCoV-19 group A,D,G rev: 1.634) A and IgG: 10.59) D, and an: 10.76) G. The da: 10.76) A  C and IgG: 11.07) F and for: 10.80) I.p = 0.0165, median\u2014men: 11.03; women: 10.63)  .p = 0.0089; IgG, p = 0.0146) and women also vaccinated with ChAdOx1 nCoV-19  (p = 0.0145)  ) (p = 0.0080) (median (BNT162b2 men: 11.49)) (ChAdOx1 nCoV-19 women: 10.46)). At 4\u20136 months after the second dose, the group of men vaccinated with BNT162b2 exhibited higher anti-RBD IgA levels compared to men (p = 0.0030) and women (p = 0.0003) immunized with ChAdOx1 nCoV-19 (p = 0.0273) and men (p < 0.0001) in the BNT162b2 group   ) (p = 0.0014) and men (p = 0.0024) in the BNT162b2 group and compared to men in the ChAdOx1 nCoV-19 group (p = 0.0448) (median (ChAdOx1 nCoV-19: men: 10.05)  (CoronaVac: women: 5.224)) (p = 0.0003) and women (p = 0.0182) vaccinated with BNT162b2  (CoronaVac: women: 9.895)) (p = 0.0021) and men (p = 0.0297) vaccinated with ChAdOx1 nCoV-19  (BNT162b2: men: 9.279)) (The comparison of sexes between vaccine groups at post-vaccination collection periods  indicate 0.0001) A,D. In a 11.90)) A. The ev 11.90)) G, reveal nCoV-19 B. At the0.6920)) B. Women  5.224)) E. Women  9.895)) H. At 1 m 9.279)) C. The an 9.279)) F,I.Furthermore, we stratified the data of vaccinees in three age groups\u201418\u201330 years, 31\u201350 years, and >50 years old\u2014to evaluate the influence of age on antibody production.p = 0.0062) (median (ChAdOx1 nCoV-19: 0.9070) (BNT162b2: 4.135)). Individuals vaccinated with BNT162b2 also presented higher anti-RBD IgA levels (p = 0.0188) (median (BNT162b2: 18\u201330 years: 4.135) (ChAdOx1 nCoV-19: >50 years: 0.7880)) and anti-spike IgG (p = 0.0075) (median (BNT162b2: 18\u201330 years: 12.20) (ChAdOx1 nCoV-19: >50 years: 10.09)) than those older than 50 years vaccinated with ChAdOx1 nCoV-19 (p = 0.0038) and those older than 50 years (p = 0.0460) vaccinated with ChAdOx1 nCoV-19 (p = 0.0084) (p = 0.0379) in the CoronaVac group and compared to individuals aged 18\u201330 years in the ChAdOx1 nCoV-19 group (p = 0.0369)   (CoronaVac: 31\u201350 years: 0.7570)) (p = 0.0002) and ChAdOx1 nCoV-19 (p = 0.0405) compared to those aged 31\u201350 years in the CoronaVac group (p = 0.0218) (median (ChAdOx1 nCoV-19: 18\u201330 years: 9.872)  (CoronaVac: 31\u201350 years: 4.402)). Analysis of anti-spike IgG levels showed that individuals aged 18\u201330 years vaccinated with BNT162b2 presented higher levels of this antibody compared to those aged 31\u201350 years vaccinated with CoronaVac (p = 0.0030) (median (BNT162b2: 18\u201330 years: 12.12) (CoronaVac: 31\u201350 years: 9.761))  B. Those 0.7570)) B. Assessac group E. Furthe 9.761)) H.p = 0.0296), and those aged 18\u201330 years (p = 0.0142), both in the ChAdOx1 nCoV-19 group  (BNT162b2: 18\u201330 years: 7.252)) (p = 0.0259) (median (ChAdOx1 nCoV-19: 10.21) (BNT162b2: 12.08)) (p = 0.0347) (median (BNT162b2: 12.24) (ChAdOx1 nCoV-19: 10.30)) ) C. The as 12.08)) F. The an 10.30)) I.p = 0.0096, median\u2014comorbidities: 10.67; no comorbidities: 12.06)  , none of: 12.06) . This wa: 12.06) .p = 0.0180, median\u2014COVID-19+: 1.673; COVID-19\u2212: 0.7415) and CoronaVac  groups compared to na\u00efve individuals (uninfected)    (n = 3).We next determined serum antibody levels of vaccinees with a history of COVID-19, infected with SARS-CoV-2 at any moment, before or during the study. Of the 22 infected individuals, 13 were infected with SARS-CoV-2 before any vaccine dose was administered and nine were infected after one, two, or three vaccine doses. Due to the reduced number of individuals who had the infection, we were unable to analyze them according to the period in which they presented COVID-19. For anti-RBD IgA, individuals who were infected with SARS-CoV-2 at any moment showed higher antibody levels at 4\u20136 months after the second dose of ChAdOx1 nCoV-19 (nfected) A. The sa: 3.446) B. The ev: 3.446) C. We did: 3.446)  for anti: 10.86) . At 1 mo: 10.88) . However2 = 0.5979, p = 0.0013)  D, after  0.0013) E,F.In this study, we investigated the specific humoral immune response against RBD and spike proteins after COVID-19 vaccination with three vaccine formulations: ChAdOx1 nCoV-19, BNT162b2, and CoronaVac. The data showed that, after the primary vaccination regimen (two-dose regimen), all vaccinees showed an increase in anti-RBD IgG and IgA and anti-spike IgG levels and were able to maintain these antibody levels for 4\u20136 months after two doses. BNT162b2 induced greater production of anti-RBD IgG and IgA and anti-spike IgG compared to ChAdOx1 nCoV-19. These findings are similar to those of Zhang et al. (2022), who detected higher anti-spike and anti-RBD IgG levels induced by mRNA vaccines compared to adenovirus and recombinant protein vaccines . The booThe longevity of the immune response induced by vaccines may vary according to the vaccine platform used, among other factors. We found positive levels of anti-RBD IgG and IgA and anti-spike IgG antibodies up to 4\u20136 months after the second dose for the three vaccine formulations evaluated. Similarly, in other studies using sera from individuals immunized with mRNA vaccines such as mRNA-1273 (Moderna), neutralizing and binding antibody activity against SARS-CoV-2 variants was detected six months after the primary vaccination regimen ,42. GreaThe use of booster doses has already proven to be essential in improving the immune response, especially against SARS-CoV-2 variants . Many stTo assess whether host factors, such as sex, age, and comorbidities, could influence the response to the vaccines, these factors were evaluated to observe changes in anti-RBD IgG and IgA and anti-spike IgG production since some host characteristics can interfere with the development of immune responses against SARS-CoV-2 . No diffThe next evaluation conducted according to biological characteristics was the effect of age on antibody production. Aging has already been shown to be responsible for a decrease in cellular immune function due to the process known as immunosenescence ,53. It cAlthough the presence of comorbidities may hinder the induction of immune responses by different vaccines ,57,58, iOur research demonstrated higher production of anti-RBD IgG and IgA in individuals with a history of SARS-CoV-2 infection compared to na\u00efve individuals, during the collection period 4\u20136 months after the second dose, suggesting that the infection could have potentiated antibody production, especially for individuals in the CoronaVac group. This hypothesis can be corroborated by a study conducted by Padoan et al. (2021), which showed increased anti-S-RBD IgG levels in individuals previously exposed to SARS-CoV-2 and immunized with two doses of BNT162b2 vaccine compared to na\u00efve individuals . Chua etSome limitations of our study include the small size of the cohort and the differences between age and sex in each vaccine group. These limitations could have influenced the analysis of antibody production, but most of the results obtained in this work, such as the longevity of the antibody response and higher antibody levels induced by the heterologous regimen, are similar to those of other studies ,44,45,61In conclusion, two doses of the BNT162b2, ChAdOx1 nCoV-19, and CoronaVac vaccines were sufficient to induce detectable levels of anti-RBD IgG and IgA and anti-spike IgG antibodies. The heterologous booster dose with BNT162b2 increased the levels of anti-RBD IgA and IgG and anti-spike IgG for CoronaVac and anti-RBD IgA for ChAdOx1 nCoV-19 vaccinees. In addition, individuals who presented the SARS-CoV-2 infection at any time during the study had higher anti-RBD IgA and IgG levels at 4\u20136 months after the second dose, compared to uninfected individuals. Further studies involving larger cohorts should be conducted to investigate and find answers to unresolved questions."}
+{"text": "Correction to: Lossy Image Compression in a Preclinical Multimodal Imaging Study\u00a0https://doi.org/10.1007/s10278-023-00800-5The corresponding author for this paper should be Andreas Walter; details are below.Andreas WalterCentre of\u00a0Optical Technologies, Aalen University, Aalen, Germanyandreas.walter@hs-aalen.deThe original article has been corrected."}
+{"text": "Discovery of molecular mechanisms of primary osteoporosis development is fundamental to understand the pathogenesis of musculoskeletal diseases in general and for identifying key links in the genetic and epigenetic regulation of bone remodelling genes. The number of identified molecular genetic markers for osteoporosis is increasing but there is a need to describe their functional interactions. These interactions have been determined to be associated with the control of expression of a number of transcription factors and the differentiation of mesenchymal stem cells through the pathway of osteoblastogenesis or adipogenesis, and monocytic precursors through the pathway of osteoclastogenesis. The results of epigenetic studies have significantly increased the understanding of the role of post-translational modifications of histones, DNA methylation and RNA interference in the osteoporosis pathogenesis and in bone remodelling. However, the knowledge should be systematised and generalised according to the results of research on the role of epigenetic modifiers in the development of osteoporosis, and the influence of each epigenetic mechanism on the individual links of bone remodelling during ontogenesis of humans in general, including the elderly, should be described. Understanding which mechanisms and systems are involved in the development of this nosology is of interest for the development of targeted therapies, as the possibility of using microRNAs to regulate genes is now being considered. Systematisation of these data is important to investigate the differences in epigenetic marker arrays by race and ethnicity. The review article analyses references to relevant reviews and original articles, classifies information on current advances in the study of epigenetic mechanisms in osteoporosis and reviews the results of studies of epigenetic mechanisms on individual links of bone remodelling. Primary osteoporosis (OP) is an age-associated disease ofmultifactorial aetiology, which is based on a violation ofthe balance of bone remodelling, leading to a decrease inthe level of bone mineral density (BMD) and a violationof the structure of bone microarchitectonics, which resultin the appearance of an incorrect spatial structure of thespongy and cortical bone . Bone massis reduced to 26 % in individuals predisposed to OP. Inroughly 80 % of women, the mineral content in the spinefalls below the threshold value at the age of 60\u201370 years,and at 85 years \u2013 in more than 90 % .According to statistics, the frequency of fractures in womenis 33 %, in men, 20 % .Until recently, OP was diagnosed only by secondarysigns, such as low height and bone pain . In 1940, the American endocrinologist F. Albright,describing postmenopausal osteoporosis, suggestedthat it developed due to estrogen deficiency . On this basis, clinicians developed the now obsoleteconcept of two forms of OP, one associated withestrogen deficiency during menopause and the other withcalcium deficiency and skeletal ageing, both of which arecharacteristic of both sexes .Current evidence defines OP as a musculoskeletal diseaseassociated with profound metabolic changes not only inbone, but also in whole-body homeostasis: micro-nutrientand endocrine dysregulation, as well as a complex interactionof genetic, endogenous and environmental factors thatcontribute to a complex disease phenotype . Risk factors with regards to OP are assumedto be the female sex, being of the Caucasoid or Mongoloidrace , early menopause, old age, familyhistory, insufficient insolation, comorbidities with impairedbone matrix micronutrient absorption, smoking, alcoholabuse, sedentary lifestyle, taking hormonal drugs andrheumatoid arthritis. DNA testing has not been introducedinto diagnostic practice because significant populationdifferences in the frequency distribution of risk markersprevent the development of universal test systems, despitethe existence of significant and validated genetic markersfor OP .The first multicentre molecular genetic studies basedon the genome-wide association search (GWAS) methodconfirmed that OP is associated with genes for local andsystemic regulation of bone cell function. The lion\u2019s shareof risk markers, specifically, single-nucleotide polymorphisms,have been identified not in exons but in intronsand promoters of regulatory genes, transcription factor,receptor (ESR2) and growth factor (FGF2) genes .Certain markers have been identified in non-coding RNA(ncRNA) sequences, including microRNA . Significant levels of associationswith fractures have been identified among the genesof systemic and local regulators ,transmembrane receptors, as well as WNT signalling genes,nuclear transcription factors  and enzymesthat produce or inactivate local bone regulators .Bioinformatics studies have concluded that the main enrichmentpathways by means of the functional affiliation ofgenes associated with histone modifications and microRNApatterns are significantly associated with the regulation ofRUNX2, FGF2 and SOX9 transcription factors. Moreover,they are also associated with the regulation of mesenchymalstem cell (MSC) differentiation .Thus, particular epigenetic factors in the pathogenesis ofosteoporosis have been identified. However, at present, it ismore interesting to consider the molecular pathogenesis ofOP in terms of individual functional links in the regulationof bone remodelling, in particular how RANK/RANKL/OPG, WNT, RUNX2 transcription factor and others areepigenetically regulated.Controlled differentiation of osteoblast and osteoclast precursors(mononuclear phagocytes) is necessary to maintainthe balance of bone remodelling .Osteoclast activity is primarily regulated by the RANK/RANKL/OPG (receptor-activator nuclear factor k\u03b2/RANKligand/osteoprotegerin) system. RANKL is produced byosteoblasts and its binding to RANK on the osteoclast surface activates the expression of osteoclastogenic genes.The role of gene polymorphisms of this system in increasedrisk of fracture and the formation of low BMDhas previously been shown . Inthe RANKL gene sequence, two CpG sites were detected:one in the upstream sequence with 18 CpG sites, localizedat a distance of 14,415 pairs of nucleotides (bp) from thetranscription start site of the TSS I major isoform and onein the downstream sequence with 59 CpG sites, whichstarts at 260 and ends at 615 pairs of nucleotides of theTSS I isoform. In the OPG gene sequence, one island of56 sites spanning from \u2013402 to +850 nucleotide pairs fromthe transcription site of the TSS isoform was observed. In , a groupof patients from Guangzhou Medical University Hospitalwith osteoporotic fractures had significantly higher RANKLgene mRNA levels from femoral bone cells compared tocontrols, with downregulated methylation status .Dysregulation of the expression of these genes is knownto be a key link in the development of steroid-inducedosteonecrosis of the femoral head and was noted to be associatedwith increased DNA methylation levels of OPGand RANK genes and with decreased levels in the RANKLgene . Previously, J. Delgado-Calle et al.(2012) performed a comparative analysis of the expressionlevels and methylation profile of the RANKL and OPGgenes in bone tissue samples from patients with osteoporoticfemoral neck fractures and osteoarthritis (OA) ofthe hip joint, which obtained remarkable results. RANKLexpression levels were significantly higher in patientswith fractures ( p = 0.012), while no significant changes inOPG gene expression levels were observed. The ratio ofRANKL ligand to OPG was higher in bone samples withOP . Differentialmethylation analysis revealed that the upstream promoterregion of the RANKL gene was strongly methylated inall samples, while individual CpG junctions of the genewere equally hypomethylated in the comparison groups.There is evidence of the contribution of miRNA tothe regulation of the RANK/RANKL/OPG system. In2014, C. Chen et al. published results measuring miR-503microRNA levels in peripheral blood cells, the overexpressionof which in CD14+ mononuclear cells inhibitedRANKL-induced osteoclastogenesis. In CD14+ cells frompostmenopausal women with osteoporosis, the baselinelevel of miR-503 was lower than in normal controls andhad no change after induction by RANKL factor, suggestingthe direct role of miR-503 in the regulation ofRANK expression . miR-142-3p andmiR-21- 5p are potential biomarkers of OP as they have ahigh affinity with the OPG gene mRNA and are involvedin the regulation of several signalling pathways involved inbone formation . Thus, theepigenetic regulation of the RANK/RANKL/OPG systemis dynamic and dependent on the DNA methylation statusand a number of microRNAs.Transcription factor 2 (RUNX2) plays a crucial role inosteoblast differentiation. Gene expression is predominantlyhigh in the early stages of bone cell developmentwhen MSCs are differentiating into osteoblast precursors,but naturally decreases at the osteocyte maturation stage.The gene contains several functional regions: activationdomain, runt domain, PST domain, etc. . In terms of epigenetic regulators of RUNX2,microRNAs are well studied. In particular, miRNA-194modulates MSC differentiation  andaccelerates osteoblast differentiation by regulating RUNX2nuclear translocation by way of STAT1 signal transducer. miR-133a-5p inhibits RUNX2 geneexpression at the transcription and translation level by bindingto the 3\u2032-untranslated mRNA site .During the early stages of osteoblast maturation, miR-125b affects RUNX2 expression by affinity binding to the3\u2032-untranslated region of the gene, indirectly participatingin the formation of a complex with Cbf\u03b2 that inhibits differentiationof these cells. Using microarray technology,P. Garmilla-Ezquerra et al. (2015) discovered a significantreduction in miR-187-3p gene expression levels andinduction of miR-518f in bone with low BMD. In theirresearch, Y. Zhang et al. (2017) observed the involvementof miR-221 in the formation of low mineral density throughregulation of RUNX2 activity based on bioinformaticsanalysis . Several microRNAs affecting the activity of thisfactor are presented in Table 1.Phosphorylation of RUNX2 mobilises chromatin regulatoryfactors and accelerates MSC maturation. RUNX2 isphosphorylated at specific serine residues 301 and 319, inducingosteocyte maturation through MAPK-dependentsignalling  and BMP2-sensitive transcription . Phosphorylationof S104 leads to prevention of RUNX protein degradation. The ERK-MAPKsignalling pathway plays a crucial role in the regulationof RUNX2 gene expression and bone formation . MKK6 is a protein kinase with dual specificity thatparticipates in the MAP kinase signal transduction pathwayand promotes RUNX2 phosphorylation . Inaddition, parathormone, which is one of the main regulatorsof blood calcium levels, activates the phosphorylation ofthe RUNX2 factor by means of the PKA signalling pathway.This process is associated with the activation of the promoter of the MMP13 gene, which plays an essentialrole in bone resorption .Post-translational modification of histones, in particularhistone methylation, plays an important role in bone formation.The so-called JUMONJI protein is considered to bea transcription factor and is encoded by the JARID2 gene.The domain containing 3 JMJD3 is a histone demethylasespecifically catalysing the removal of trimethylation ofhistone H3K27me3. JMJD3 was found to inhibit the differentiationof RUNX2 osteoblasts. Conversely, inhibitionof JMJD3 activity decreases RUNX2 promoter activitywhile increasing H3K27me3 activity in promoter regions.Histone acetylation affects the state of chromatin compactionby neutralising the positive charge of histone tailsand reducing the electrostatic interactions of histone tailswith deoxyribonucleic acid. Studies have revealed thatosteoporosis induced by glucocorticoid therapy leads todecreased acetylation of H3K9/K14 and H4K12 in theregulatory regions of the RUNX2 and OSX genes andincreases the hyperacetylation of H3K9/K14 and H4K12in the PPAR\u03b32 regulatory region in bone marrow-derivedMSCs from osteoporosis. The transcriptional activity of theRUNX2 factor gene is enhanced by P300 acetyltransferaseand nicotinamide phosphoribosyltransferase (NAMPT),which, in turn, promote osteogenic differentiation of MSCsby H3K14 acylation and MC3T3-E1 cells through H3K9acylation, respectively .Thus, an extremely significant number of regulatorsof RUNX2 gene expression have been identified, whichis a promising therapeutic gene model in culture studies,as blocking or enhancing the activity of this gene andmonitoring its expression level during the induction ofosteogenic lines allows the identification of key switchesof mesenchymal stem cell differentiation.The WNT signalling pathway is one of the most importantsystems regulating embryonic development and cell differentiation.This pathway represents one of the centrallinks in the control of bone development and remodelling.Among the various genes involved in this system, methylationof the SOST (encoding sclerostin) gene promoterhas been comprehensively studied in osteoblast cultures.Sclerostin produced by osteocytes inhibits WNT signallingand reduces the rate of bone formation. DNA methylationof the gene is necessary for the transition of osteoblasts toosteocytes . In women withprimary OP, SOST methylation is elevated in iliac bonecells whilst sclerostin levels are decreased and the WNTpathway is enhanced .Several studies have shown that histone deacetylationunder the regulation of the WNT6, WNT10B, WNT10A andWNT1 genes inhibits WNT signalling and increases the riskof primary osteoporosis . Thus, elevatedlevels of HDAC5 deacetylase reduce SOST gene expressionin osteocytes, contributing to bone mass loss. Deficiency ofthis deacetylase is associated with the acetylation of histoneH3 lysine 27 as well as the interaction of MEF2C with theSOST gene enhancer, therefore suggesting the significantrole of sclerostin in the regulation of osteocyte maturation. High levels of the zeste 2 homologueenhancer methyltransferase (EZH2) have been demonstratedto suppress osteogenic differentiation of MSCs,while low ones decrease the levels of the H3K27me3 tagnear the transcription start site of osteogenesis genes, includingWNT10B . EZH2 increasesH3K27me3 levels at the WNT1, WNT6 and WNT10Apromoters and inhibits WNT signalling .Many regulatory microRNAs are associated with WNTsignalling. miR-433-3p inhibits DKK1 (Dickkopf-1)gene expression, enhancing osteoblast differentiation.Dickkopf-1 acts as an antagonist of the WNT signallingpathway and enhances bone resorption .miR-139-5p induces WNT signalling via the inhibition ofNOTCH1 . R.E. Makitie et al. (2018)screened a specially designed panel of 192 microRNAsin patients with a genetically determined WNT signallingdisorder with a heterozygous missense mutation c. 652 T>G(p. C218G) in exon 4 of the WNT1 gene. It was determinedthat miR-22-3p, miR-34a-5p and miR-31-5p levels werelower in mutation carriers compared to controls .Another common inhibitor of osteogenic differentiationis miR-31, the level of which drops in MSCs differentiatinginto osteoblasts. This was confirmed by S. Weilner et al.(2016), who observed increased levels of this microRNA in plasma in elderly patients with OP . miR-31 isreleased from extracellular vesicles of endothelial cellsand inhibits osteogenesis in stromal stem cells by bindingto the Freisled 3 protein. Furthermore, decreased levels ofmiR-199a-5p result in glucocorticoid-mediated inhibitionof osteogenesis . More recently, L. Duanet al. (2018) identified that high levels of miR-16-2* maycontribute to the development of primary OP: knockdownof this microRNA may promote RUNX2 activation. ThismicroRNA has an affinity for the WNT5A gene mRNA. miR-148a-3p has been revealed toenhance both osteoclastogenesis  andadipogenesis in osteogenic progenitor cells . Plasma levels of this microRNA are significantlyhigher in patients with OP compared to controls withoutOP . miR-30e is another importantmicroRNA in the pathogenesis of OP, playing a role inregulating adipocyte and osteoblast differentiation throughthe inhibition of LRP6 . Thus, the WNTsignalling pathway is regulated by a complex epigeneticsystem, notably microRNAs.The role of non-coding RNAs in bone remodellingMicroRNAs are considered to be the most studied epigeneticfactors in osteoporosis .They are conventionally divided into two classes: those thatpromote bone formation or bone resorption. In particular,several microRNAs that slow down the progression of OPhave been identified. For example, miR-33-5p is a mechanosensitivemicroRNA that positively regulates osteoblastogenesisby way of inhibition of HMGA2 high mobilitygroup proteins . miR-96 enhances osteogenicdifferentiation by inhibiting phosphorylation ofepidermal growth factor receptor EGFR and the expressionof major osteoblast factors RUNX2 and OSTERIX. miR-216a enhances bone formation byregulating the c-Cbl-mediated PI3K/AKT pathway . Table 2 shows the microRNAs separated bydirection of action in bone remodelling .miR-124 is a positive regulator of adipogenic and neurogenicdifferentiation, while being a negative regulator ofmyogenic and osteogenic differentiation. It directly targetsthe DLX3, DLX5 and DLX2 homeobox genes . In one pharmacogenetic study, patients with OP,after three months of treatment with the parathormoneanalogue Teriparatide, had reduced expression levels ofmiR-33 and one year later, miR-133a. Simultaneously,there was a general tendency: the increase in the level ofBMD increased with a decrease in the level of expressionof these microRNAs. Post-transcriptional regulation ofDKK-1 changes due to a decrease in miR-33 microRNAlevels and the action of parathyroid hormone, which leads toa decrease in the negative impact of DKK-1 on an alternativeregulatory mechanism that improves optimal controlof WNT signaling..Of interest are the results of studies on the effects oflong non-coding RNAs on the regulation of Sirtuins,nicotinamide adenine dinucleotide-dependent deacetylases.These molecules have a broad spectrum of action and areassociated with longevity and resistance to age-relateddiseases. It was established that SIRT1 gene expression isinversely related to HIF1A-AS1 ncRNA expression, whileHOXA-AS3 ncRNA interacts with EZH2 and is requiredfor RUNX2 trimethylation of lysine-27 H3 (H3K27me3)factor. Hence, HOXA-AS3 is important for bone formationin general .The long ncRNA HOTAIR reduces protein expressionand inhibits WNT signalling. DKK1 reduces the proteinlevels of C-myc, \u03b2-catenin, HOTAIR and RUNX2, whichtheoretically counteracts the regulatory effect of HOTAIR. If the expression level of p21 ncRNA islow, WNT signalling becomes more active due to increasedE2 secretion, which ultimately increases the rate of boneformation . Also, reduced levels of H19ncRNA reduce the level of DKK4 gene expression . AK045490 ncRNA levels are significantlyelevated and inhibit bone formation by inhibiting nucleartranslocation of \u03b2-catenin and suppressing TCF1, LEF1 andRUNX2 expression . Similarly, AK016739ncRNA inhibits osteogenic differentiation as it can reducethe expression and activity of osteoblastogenesis transcriptionfactors .Inhibition of UCA1 ncRNA promotes bone formationvia activation of the BMP-2/(Smad1/5/8) pathway in osteoblasts. As a result, microRNAs andlong ncRNAs remain among the most studied epigeneticfactors involved in the pathogenesis of primary OP butrequire further systematisation.The mechanisms of the relationship between bone andadipose cell formation are complex and remain an area ofactive research. Works performed on the cell cultures ofosteoblasts and MSCs convincingly show an inverse relationshipbetween differentiation of bone marrow MSCs intoadipocytes or osteoblasts. An imbalance between adipogenesisand osteogenesis has been proposed as a mechanismfor the development of OP, but obesity itself is not alwaysa predictor of an increased risk of osteoporosis.Post-translational modifications of histones play a keyrole in this system. Among them, histone methylation iscrucial, in particular in chromatin reorganisation. In particular,lysine methylation in H4K20, H3K27 and H3K9 isassociated with decreased levels of transcription, whereasmethylation of H3K79, H3K36 and H3K4, with active genetranscription .However, concerning osteogenic inducers, the homeoboxgene HOXA10, which contributes to osteogenic clone determinationby enriching the trimethylation of the 4th lysineresidue in histone, activating RUNX2, alkaline phosphataseand osteocalcin, thus stimulating bone cell maturation, isimportant . It is known that the combinationof methylation and demethylation can function as anepigenetic switch of osteogenesis to adipogenesis based onEZH2 activity, catalysing the trimethylation of histoneH3on lysine 27 key regulatory genes (such as RUNX2). Simultaneously,removal of this tag by lysindemethylase 6Ainhibits adipogenesis and induces osteoblastogenesis. Proteinsthat can be targeted by EZH2 and are involved in MSCswitching are HDAC9c and HDAC .A direct correlation was realised between increased levelsof EZH2 and decreased levels of HDAC9c gene expression. The methyltransferase activity ofEZH2 is reduced by phosphorylation and is associated withosteogenic induction It is acknowledged that during osteocyte aging there isan accumulation of adipose tissue in the bone marrow and,at the same time, the number of mesenchymal stem cellsincreases in the intercellular phase. From this perspective,it is interesting that overexpression of miR-1292 acceleratesthe senescence of human adipose-derived stem cellsand inhibits bone formation via the Wnt/\u03b2-catenin signallingpathway, while miR-10b inhibits adipose stem celldifferentiation via the TGF-\u03b2 pathway .Several bone-associated cells, including multipotentbone mesenchymal stem cells, osteoblasts that form bonetissue and osteoclasts that break it down, are in a symbioticrelationship throughout life. A growing body of evidencesuggests that epigenetic cell modifications induced byaging contribute to impaired bone remodelling and leadto osteoporosis. A variety of epigenetic mechanisms areinvolved, including DNA/RNA modifications, histonemodifications, microRNAs (miRNAs) and long non-codingRNAs (dnRNAs), and chromatin remodelling . Thus, epigenetic mechanisms can switch the directionof mesenchymal stem cell differentiation betweenosteoblastogenesis and adipogenesis.Molecular genetic determinants of endophenotypes ofosteoporosis, such as fracture risk and BMD levels, can beconverged through the whole epigenome. In the researchthey conducted, J.A. Morris et al. (2017), leveraging thetechnological capabilities of Infinium HumanMethylation450,performed a whole-genome methylome analysis,measuring site-specific DNA methylation in 5515 individualsof European descent. Following a meta-analysisof their results, they were able to identify the CpG sitecg23196985, which was significantly associated with lowMPCT adjusted for multiple comparisons without regard togender( pBH = 1.30 \u00d7 10\u20132) and in females (pBH = 3.41 \u00d7 10\u20135).The CpG cg23196985 site is localized to the 5\u2032-translatedregion of the hepatic carboxylase 1 gene CES1, which isexpressed in the liver and peripheral blood . J.G. Zhang et al. (2015), performing transcriptomeanalysis based on Affymetrix GeneChip Human Exon 1.0ST Array and microRNA analysis on Capitalbio Cor. microarrays,as well as methylome sequencing in patientswith low MPCT hip and controls, identified the mostenriched functional molecular pathways associated withOP or MPCT variability: a network of 12 interacting genesand 11 microRNAs. Among the genes are AKT1, STAT5A,PIK3R5, etc., while among the microRNAs, miR-141 andmiR-675 \u2013 their levels correlate with the expression ofthese genes and global DNA methylation status .D. Cheishvili et al. (2018) performed a full-epigenomicanalysis in women without OP and in women with earlypostmenopausal OP. Genes in which CpG sites with significantlevels of differential methylation were identifiedwere ZNF267, ABLIM2, RHOJ, CDKL5, PDCD1, ABRAand HOJ . At the human femurlevel, DNA methylation profile studies using pyrosequencingand qRT-PCR-based gene expression studies provedthat DNA methylation status was inversely correlated withthe expression of the iNOS and COL9A1 genes, but notcatabolic genes including MMP13 and IL1B. Significantdemethylation of the osteocalcin gene promoter was alsoshown between the embryonic and adult stages of development,demonstrating the importance of DNA methylation atthe tissue level . It is anticipated that theresults of these studies will confirm the whole-epigenomeapproach as being sufficiently robust to allow large-scalestudies of women at risk of developing OP.Despite the great advances and the wide range of workperformed in the study of the epigenetics of primary osteoporosis, understanding remains fragmentary. The researchdescribes key epigenetic regulators of bone remodelling,but it is difficult to build a coherent picture of the pathogenesisof osteoporosis from these data, common to all keypathogenetic processes in bone tissue.It is recognised that the key epigenetic changes in osteoporosisare converging on the regulation of MSCs differentiationand that the multi-step system of activity regulationof the transcription factor RUNX2, sclerostin,DKK1 protein, RANKL-RANK-OPG system and factorsinvolved in the regulation of WNT signalling is of greatimportance. A separate issue is the systematisation andcreation of a unified genetic network of a vast number ofregulatory microRNAs that affect key signalling pathwaysand the transcription factors associated with osteoporosis.However, these microRNAs influence the ultimate risk ofosteoporosis indirectly and it remains to be understoodhow they can be systematised for relevance and functionalinvolvement in pathogenesis due to their dynamism anddifferent levels depending on tissue specificity.Approaches necessary to implement early diagnosis ortargeted therapies for osteoporosis still need to be developed.The task is complicated by experimental data frommethylome studies, in which genes other than critical regulatoryfactors, such as RUNX2, sclerostin or DKK1, wereobserved to be key and most important. 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J. 2021;10(1):e0101.DOI 10.15275/RUSOMJ.2021.0101.Yang D., Okamura H., Nakashima Y., Haneji T. Histone demethylaseJmjd3 regulates osteoblast differentiation via transcription factorsRunx2 and osterix. J. Biol. Chem. 2013;288(47):33530-33541. DOI10.1074/jbc.M113.497040Yang K., Tian N., Liu H., Tao X.Z., Wang M.X., Huang W. LncRNAp21promotes osteogenic differentiation of mesenchymal stem cells inthe rat model of osteoporosis by the Wnt/\u03b2-catenin signaling pathway.Eur. Rev. Med. Pharmacol. Sci. 2019;23(10):4303-4309. DOI10.26355/EURREV_201905_17935.Yang M., Pan Y., Zhou Y. miR-96 promotes osteogenic differentiationby suppressing HBEGF-EGFR signaling in osteoblastic cells.FEBS Lett. 2014;588(24):4761-4768. DOI 10.1016/J.FEBSLET.2014.11.008.Yang Y., Yujiao W., Fang W., Linhui Y., Ziqi G., Zhichen W., Zirui W.,Shengwang W. The roles of miRNA, lncRNA and circRNA in the developmentof osteoporosis. Biol. Res. 2020;53(1):40. DOI 10.1186/s40659-020-00309-z.Yin C., Tian Y., Yu Y., Wang H., Wu Z., Huang Z., Zhang Y., Li D.,Yang C., Wang X., Li Y., Qian A. A novel long noncoding RNAAK016739 inhibits osteoblast differentiation and bone formation.J. Cell. Physiol. 2019;2347(7):11524-11536. DOI 10.1002/jcp.27815.Yu W., Wang H.L., Zhang J., Yin C. The effects of epigenetic modificationson bone remodeling in age-related osteoporosis. Connect.Tissue Res. 2022;1-12. DOI 10.1080/03008207.2022.2120392.Zhang J.G., Tan L.J., Xu C., He H., Tian Q., Zhou Y., Qiu C., Chen X.D.,Deng H.W. Integrative analysis of transcriptomic and epigenomicdata to reveal regulation patterns for BMD variation. PLoS One.2015;10(9):0138524. DOI 10.1371/journal.pone.0138524Zhang R.F., Liu J.W., Yu S.P., Sun D., Wang X.H., Fu J.S., Xie Z.LncRNA UCA1 affects osteoblast proliferation and differentiationby regulating BMP-2 expression. Eur. Rev. Med. Pharmacol. Sci.2019;23(16):6774-6782. DOI 10.26355/eurrev_201908_18715.Zhang W., Wu Y., Shiozaki Y., Sugimoto Y., Takigawa T., Tanaka M.,Matsukawa A., Ozaki T. miRNA-133a-5p inhibits the expressionof osteoblast differentiation-associated markers by targetingthe 3\u2032 UTR of RUNX2. DNA Cell Biol. 2018;37(3):199-209. DOI10.1089/dna.2017.3936Zhang Y., Gao Y., Cai L., Li F., Lou Y., Xu N., Kang Y., Yang H.MicroRNA-221 is involved in the regulation of osteoporosis throughregulates RUNX2 protein expression and osteoblast differentiation.Am. J. Transl. Res. 2017;9(1):126-135."}
+{"text": "Enterobacterales (MDR-Ent) has reached worrisome levels worldwide. In this context, Escherichia ruysiae is a recently described species mostly found in animals. However, its spread and impact on humans is poorly understood. A stool sample from a healthy individual living in India was screened for the presence of MDR-Ent using culture-based methods. Colonies were routinely identified using matrix-assisted laser desorption ionization\u2013time of flight mass spectrometry (MALDI-TOF MS) and phenotypically characterized by broth microdilution. Illumina and Nanopore whole-genome sequencing (WGS) platforms were implemented to generate a complete assembly. E. ruysiae genomes deposited in international databases were used for a core genome phylogenetic analysis. An extended-spectrum \u03b2-lactamase (ESBL)-producing E. coli strain (S1-IND-07-A) was isolated from the stool. WGS confirmed that S1-IND-07-A was indeed E. ruysiae, belonged to sequence type 5792 (ST5792), core genome (cg) ST89059, serotype O13/O129\u2212:H56-like, clade IV phylogroup, and possessed five virulence factors. A copy of blaCTX-M-15 and five other antimicrobial resistance genes (ARGs) were detected in a conjugative IncB/O/K/Z plasmid. A database search identified 70 further E. ruysiae strains from 16 countries . The core genome phylogeny revealed five major STs: ST6467, ST8084, ST2371, ST9287, and ST5792. Three out of the seventy strains possessed important ARGs: OTP1704 , SN1013-18 , and CE1758 . These strains were of human, environmental, and wild animal origin, respectively. E. ruysiae may acquire clinically important ARGs and transmit them to other species. Due to its zoonotic potential, further efforts are needed to improve routine detection and surveillance across One Health settings.Gut colonization with multidrug-resistant IMPORTANCEEscherichia ruysiae is a recently described species of the cryptic clades III and IV of the genus Escherichia and is commonly found in animals and the environment. This work highlights the zoonotic potential of E. ruysiae, as it has been shown to colonize the human intestinal tract. Importantly, E. ruysiae may be associated with conjugative plasmids carrying clinically relevant antibiotic resistance genes. Therefore, it is important to closely monitor this species. Overall, this study highlights the need for improved identification of Escherichia species and continued surveillance of zoonotic pathogens in One Health settings. Escherichia is comprised of 3 monophyletic species , including cryptic clades I, II, III and IV (E. ruysiae), and V (E. marmotae)  corresponding to specific host/source niches where this pathogen can be found, such as commensal E. coli of the gastrointestinal tract in humans (groups A or B1) and animals (group B1) and the environment  , such as those encoding extended-spectrum \u03b2-lactamases , which compromise the treatment (The genus armotae) \u20133. FurthB2 or D) . On thisreatment .E. ruysiae has been recently reported from animal and environmental sources, but it is still very rarely isolated in humans , unless a PCR-based approach  or whole-genome sequencing (WGS) are used  blaCTX-M-14-possessing Escherichia cryptic clade IV strain (OPT1704) was recently isolated from the stool of a traveler to Asia  isolated from a fecal sample of a healthy individual living in India. To investigate the spread of E. ruysiae in humans, we searched for other E. ruysiae genomes in international databases and compared the results using core genome phylogenetic analysis. Furthermore, we investigated the distribution of the associated ARGs, plasmids, virulence factors, and serotypes characteristic of strains isolated from various human and nonhuman samples.In this work, we characterized a E. ruysiae strain isolated from a person living abroad. To contextualize our findings, the second aim was to search global databases for other reported E. ruysiae strains isolated from different sources to perform a core genome phylogeny analysis. Finally, we examined the associated ARGs, plasmid replicons, virulence, and serotype distribution of all E. ruysiae strains included in this study to evaluate the potential of E. ruysiae as a human pathogen.The first aim of this study was to characterize a rare ESBL-producing E. coli . As shown in Screening of the individual\u2019s stool yielded a third-generation cephalosporin-resistant strain (S1-IND-07-A) that was identified by MALDI-TOF MS as E. ruysiae . This was also supported by in silico phylotyping, which determined that the species belonged to Escherichia cryptic clade IV.Using WGS, the species was confirmed as blaCTX-M-15, aadA1, dfrA1-like (99.79% identity), qnrS1, sul2, and tet(A). In the chromosome, mutations encoding for the quinolone resistance-determining region substitution (S83A) in GyrA were identified, together with the mdf(A)-like ARG (91.32% identity) . Overall, the genotypic screening results were consistent with the observed phenotype (S1-IND-07-A was of sequence type 5792 (ST5792) and core genome (cg) ST89059. Its genome consisted of a 4.4-Mb chromosome and 2 circular plasmids of 105.8\u2009kb (IncB/O/K/Z) and 1.5\u2009kb [Col(MG828)]. Most ARGs were carried by the IncB/O/K/Z plasmid (p1-S1-IND-07-A): henotype .in silico serotyping revealed that S1-IND-07-A carried O13/O129-like  and H56-like  antigens. Several virulence factor genes were also identified, including those encoding colonization factors (fimH), fitness (iutA and kpsMII), and toxins (pic and sat) (Table S1). Of note, p1-S1-IND-07-A carried the colicin Ib virulence factor gene cib, which is known to enhance virulence  in, for example, E. coli and Salmonella enterica (Table S1)  \u201313.E. ruysiae strains of the same sequence type and core genome sequence type (ST5792 and cgST89059). We also included strains isolated from different sources to identify potential ARG hot spots and evaluate the pathogenic potential of E. ruysiae in humans. Overall, we included a total of 70 E. ruysiae genomes with complete metadata  from the NCBI  and Enterobase  databases.To investigate the origin of S1-IND-07-A, we performed a database search for other E. ruysiae strains belonging to the cryptic clades III (n\u2009=\u200936) and IV (n\u2009=\u200935), detected between 1989 and 2022 , environmental , and human sources . The strains originated from 16 countries, mainly from the United States (n\u2009=\u200931) and the United Kingdom (n\u2009=\u200916).As a result, along with our strain, we identified a total of 71 n\u2009=\u20099), ST8084 (n\u2009=\u20098), ST2371 (n\u2009=\u20096), ST9287 (n\u2009=\u20094), and ST5792 (n\u2009=\u20094)  were observed. Overall, they consisted of strains isolated from poultry in the UK (C17-1 and C10-7), companion animals in Australia (MVC117 and MVC118), and the environment in the United States  \u201317.\u2013E. ruysiae strains from clade IV carried \u03b2-lactamase (bla) genes: CE1758 (blaCMY-2), OPT1704 (blaCTX-M-14 and blaLAP-2), and SN1013-18 (blaCTX-M-15 and blaTEM-1). The blaCTX-M-14 and blaLAP-2 genes in OPT1704 were associated with an IncI plasmid, while the blaCMY-2, blaCTX-M-15, and blaTEM-1 genes in CE1758 and SN1013-18 were unspecified (As depicted in mdf(A) and  sitABCD. Furthermore, many replicon sequence types were detected in both clades III and IV, most frequently Col and IncFI type  or O36-like (n\u2009=\u200910) and H42-like (n\u2009=\u200915), H52-like (n\u2009=\u200911) and H56-like (n\u2009=\u200910), while clade IV strains were O36-like (n\u2009=\u200911) or O13/O129-like (n\u2009=\u20098) and H56-like (n\u2009=\u200920). Furthermore, many virulence factors were also detected, but only a few were clinically relevant, as they were related to host colonization (fimH), fitness (iutA and kpsMII), toxins , and effectors (aaiC)  and pic and sat . Both estB and astA genes have been reported in enterotoxigenic E. coli strains in both diseased pigs and pigs with diarrhea, while pic and sat have been found in enteroinvasive E. coli strains . Compared to p1-S1-IND-07-A, the other plasmids were missing an IS26 flanking region containing tet(A) and sul2 and including a disrupted class I integron (missing sul1) region associated with the dfrA1-aadA1-qacE\u03941-IS26-\u0394blaTEM-1 cassette. Similar integron regions have been reported in hyperepidemic carbapenemase-producing E. coli strains also carrying blaTEM-30-associated IncI1 plasmids from colonized employees of a Swiss veterinary clinic (In our is study . Howevery clinic .blaCTX-M) and multiple hosts   \u201327. In f J53d-R1 , thus coEnt in the gut, which may include zoonotic diseases that can be transmitted between humans, animals, and the environment  and transfer them to other clinically important bacteria. Therefore, the application of WGS surveillance methods  is essential for the discovery and characterization of novel Escherichia species with zoonotic potential and the ability to serve as reservoirs for resistance plasmids. In this context, it is crucial to develop and improve routine laboratory identification techniques such as MALDI-TOF MS to distinguish between Escherichia species.In the present work, we showed that Enterobacterales (MDR-Ent) on his return to Switzerland in 2022 as part of an ongoing prospective screening study (https://data.snf.ch/grants/grant/192514). The stool sample was collected by the volunteer in a sterile container and sent within 1\u2009day to the Institute for Infectious Diseases in Bern for processing (see below). No hospitalization or diarrhea was reported, but previous antibiotic use (not specified) was noted in two independent episodes 2 and 4\u2009months before arrival in Switzerland. The study participant provided written consent, and the study was approved by the ethical committee of the Canton Bern, Switzerland (2020-01683).A healthy 62-year-old Swiss man living in India for 1.3\u2009years was screened for intestinal carriage of multidrug-resistant A stool sample (~50\u2009\u03bcg) was preenriched for 6\u2009h in 10\u2009mL Luria-Bertani (LB) broth supplemented with cefuroxime . The preenrichment was then diluted to a 0.5 McFarland standard, and a 100-\u03bcL aliquot was plated overnight at 36\u2009\u00b1\u20091\u00b0C on a CHROMID ESBL agar plate (bioM\u00e9rieux). The resulting colonies were subcultured on MacConkey II agar  overnight at 36\u2009\u00b1\u20091\u00b0C for further processing.Enterobacterales .Species identification of overnight-grown colonies was conducted using MALDI-TOF MS . Antimicrobial susceptibility testing (AST) was performed by broth microdilution implementing the Sensititre GNX2F panel (Thermo Fisher Scientific). Results were interpreted according to the 2023 European Committee on Antimicrobial Susceptibility Testing (EUCAST) v13.0 breakpoints for https://github.com/usadellab/Trimmomatic) and Porechop v0.2.4 (https://github.com/rrwick/Porechop), respectively. Long reads were filtered by read quality using Filtlong v0.2.1 (https://github.com/rrwick/Filtlong) . A complete genome assembly was generated using the hybrid pipeline from Unicycler v0.4.8 (https://github.com/rrwick/Unicycler), and its completeness  was confirmed with an independent long-read-only assembly using Flye v2.9-b1768 (https://github.com/fenderglass/Flye) (parameters: \u201c\u2013nano-raw\u201d and \u201c\u2013iterations 5\u201d). Genome assembly coverage was calculated using QualiMap v2.2.2-dev .Genomic DNA for WGS was extracted from strain S1-IND-07-A using the Invitrogen PureLink microbiome DNA purification kit (Thermo Fisher Scientific), and the quality was assessed using NanoDrop and Qubit 3 (Thermo Fisher Scientific). Short-read WGS was performed on a NovaSeq 6000 platform , while long-read WGS was conducted on a MinION sequencer (Oxford Nanopore) using a rapid barcoding library prep (SQK-RBK004) and a FLO-MIN 106D R9 flow cell, as described previously \u201331. In b\u2013E. coli number 1 configuration), cgMLSTFinder v1.2 (parameter: E. coli [Enterobase]), SerotypeFinder v2.0, and VirulenceFinder v2.0 (parameters: 60% minimum coverage and 95% minimum identity), respectively, from the Center for Genomic Epidemiology (https://www.genomicepidemiology.org/). In addition, the phylogroup was determined using the ClermonTyping tool (Clermont Type) available at Enterobase (https://enterobase.warwick.ac.uk/) (Escherichia/Shigella database). Finally, the species ID was confirmed using the Type (Strain) Genome Server (TYGS) (https://tygs.dsmz.de/) and by ANIb (based on BLAST) using JSpeciesWS (https://jspecies.ribohost.com/jspeciesws/) with the genome assembly.The genome assembly of S1-IND-07-A was screened for ARGs, plasmid replicon sequences, sequence type (ST), core genome ST (cgST), serotype, and virulence using ResFinder v4.1, PlasmidFinder v2.1 (parameters: 60% minimum coverage and 50% minimum identity), MLST v2.0 (parameter: https://isfinder.biotoul.fr/) with the BLASTn algorithm. Circular BLASTn plasmid comparisons were performed using BLAST Ring Image Generator v0.95 (https://brig.sourceforge.net/) with the most similar plasmids retrieved from NCBI BLASTn (https://blast.ncbi.nlm.nih.gov/Blast.cgi) and PLSDB v2021_06_23_v2 (https://ccb-microbe.cs.uni-saarland.de/plsdb/) on 27 January 2023.The genome assemblies were automatically annotated using the NCBI Prokaryotic Genome Annotation Pipeline. Insertion sequences (ISs) were manually confirmed using ISfinder  and Enterobase (https://enterobase.warwick.ac.uk/species/index/ecoli)  databases on 30 January and 2 March 2023, respectively. The species IDs of all the retrieved genomes were confirmed using the TYGS tool as described above.GCA_902498915.1) as the mapping reference. Genome regions with recombination were detected and removed using Gubbins v2.3.4 (https://github.com/nickjcroucher/gubbins). Furthermore, IS regions were identified with ISEScan v1.7.2.3 (https://github.com/xiezhq/ISEScan) and subsequently used to mask single nucleotide variants (SNVs) localized within these regions . A maximum likelihood core genome phylogeny (unrooted) was inferred using IQ-TREE v2.1.2 (http://www.iqtree.org/) with the general time reversible (GTR) model with ascertainment bias correction (parameters: GTR\u2009+\u2009ASC), 1,000 ultrafast bootstraps , and the SH-aLRT test . The tree was visualized and annotated using iTOL v6.7.2 (https://itol.embl.de/). Unless otherwise noted, all software analyses described above were performed with default parameters.A core genome alignment was performed using Snippy v4.4.5 with strain OPT1704 (GenBank assembly accession number E. coli recipient strain J53d-R1 was conjugated with the donor S1-IND-07-A as previously described (blaCTX-M group I primers (The rifampicin-resistant escribed . In shor primers . ExperimCP112983 to CP112985) and under BioProject accession number PRJNA903117.The complete genome sequence of S1-IND-07-A has been deposited at GenBank (accession numbers"}
+{"text": "Healthcare personnel (HCP) are at risk for acquiring and transmitting coronavirus disease 2019 (COVID-19) at work. Vaccination against COVID-19 is a primary strategy to prevent transmission in healthcare settings. Our objectives were to describe HCP reporting employer COVID-19 vaccine requirements, identify differences across occupations and settings, and describe policies.In a national opt-in internet panel survey of HCP in April 2022 to assess influenza and COVID-19 vaccination coverage, respondents were also asked about employer vaccination requirements. We weighted responses to the U.S. HCP population by age, sex, race, ethnicity, work setting, and U.S. census region. We assessed differences using two-tailed t tests with significance set at p< 0.05. We used multivariable logistic regression to identify factors associated with reporting employer requirements.Of 3,618 responding HCP, 60.0% reported employer requirements for the COVID-19 primary series. Of those, 95.5% reported completing the primary series vs. 81.9% without requirements (p< 0.001). In addition, 24.0% of HCP reported employer requirements for a booster dose. Of those eligible, 90.8% reported receiving a booster vs. 60.2% without a requirement (p< 0.001). Employer requirements for the primary series varied by occupation [46.1% (assistants/aides) to 77.5% (nurses)] and work setting [40.4%  to 75.4% ]. Of 2,157 HCP reporting a primary series requirement, employer accepted reasons for not being vaccinated included medical (56.8%), religious (50.9%), philosophical (10.0%), all three reasons (16.7%), and no reason (14.9%). Factors independently associated with no reported primary series requirements included working as an assistant/aide, in a non-clinical or other clinical occupation, in ambulatory care, long-term care/home health, or other clinical setting, and in the Midwest and South.Most HCP in this sample reported employer requirements for the COVID-19 vaccination primary series although requirements were less common for boosters. Differences by healthcare setting exist. Implementing workplace strategies, including vaccine requirements, may lead to higher COVID-19 vaccination coverage.All Authors: No reported disclosures"}
+{"text": "Wikstroemia indica, a plant belonging to the Thymelaeaceae family, was investigated by LC-MS/MS analysis. As a result, 21 daphnane diterpenoids (1\u201321) in the stems of W. indica were detected. Among these, six major compounds  were isolated and their structures were unequivocally identified through a comprehensive analysis of the MS and NMR data. For the minor compounds , their structures were elucidated by in-depth MS/MS fragmentation analysis. This study represents the first disclosure of structurally diverse daphnane diterpenoids in W. indica, significantly contributing to our understanding of bioactive diterpenoids in plants within the Thymelaeaceae family.Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) has emerged as a powerful tool for the rapid identification of compounds within natural resources. Daphnane diterpenoids, a class of natural compounds predominantly found in plants belonging to the Thymelaeaceae and Euphorbiaceae families, have attracted much attention due to their remarkable anticancer and anti-HIV activities. In the present study, the presence of daphnane diterpenoids in  Liquid chromatography coupled with high-resolution tandem mass spectrometry (LC-HR-MS/MS), usually equipped with an electrospray ionization source, has high adaptability across a broad spectrum of compounds, offering high mass accuracy and sensitivity. Moreover, it provides information-rich fragmentation through product ion spectra, thereby potentially revealing details about the molecular formula and structure of diverse secondary metabolites found in plants . The contrans-fused 5/7/6-tricyclic skeleton, have garnered attention for their diverse biological activities, including anticancer +) were observed in positive ion mode, while deprotonated molecular ions ([M\u2013H]\u2212) and/or formate adduct ions ([M + HCOO]\u2212) were observed in negative ion mode. (2) In the product ion spectrum obtained from the protonated molecular ion as a precursor ion, a diagnostic ion at m/z 253 (C17H17O2) or 269 (C17H17O3) was observed in the positive ion mode . Th. Th12, 1e G (12) , wikstro + appeared with greater intensity in the mass spectrum of peak 3  + (1), m/z 1028.4264 [M + NH4]+ (2), m/z 585 [M + H\u2013H2O]+ (3), m/z 547.2888 [M + H]+ (4), m/z 655.3469 [M + H]+ (5), m/z 719.3062 [M + H]+ (6), m/z 533.3104 [M + H]+ (7), m/z 775.3690 [M + H]+ (8), m/z 691.3133 [M + H]+ (9), m/z 721.3203 [M + H]+ (10), m/z 775.3687 [M + H]+ (11), m/z 912.4147 [M + NH4]+ (12), m/z 667.3107 [M + H]+ (13), m/z 673.2999 [M + H]+ (14), m/z 533.3107 [M + H]+ (15), m/z 557.3109 [M + H]+ (16), m/z 675.3151 [M + H]+ (17), m/z 639.3527 [M + H]+ (18), m/z 643.3458 [M + H]+ (19), m/z 639.3525 [M + H]+ (20), and m/z 585.3409 [M + H]+ (21). All data collected in the profile mode were acquired and processed using Thermo Xcalibur 4.1 software.The LC-MS/MS analysis was performed using the same instruments and column as in previous experiments . For LC W. indica through a combination of LC-MS guided isolation and MS/MS fragmentation elucidation. The investigation revealed that W. indica contained structurally diverse daphnane diterpenoids, including orthoester daphnane type, polyhydroxy daphnane type, and macrocyclic daphnane orthoester type compounds. The application of MS/MS fragmentation elucidation for structural analysis enabled the rapid and precise identification of these diterpenoids within crude plant extracts. This methodology holds great promise for future research endeavors aimed at discovering bioactive diterpenoids from plants of the Thymelaeaceae family.This study represents the first comprehensive identification of 21 daphnane diterpenoids from the stems of"}
+{"text": "Correction: Respiratory Research (2022) 23:291 10.1186/s12931-022-02195-3Following the publication of the original article , it was The original article has been corrected.Additional file 8: Table S1. The RECORD statement\u2014checklist of items, extended from the STROBE statement, that should be reported in observational studies using routinely collected health data.Additional file 9: Table S2. Disease names adopted as inclusion criteria.Additional file 10: Table S3. Disease names adopted as exclusion criteria."}
+{"text": "Mycobacterium marinum strain, which was isolated from skin tissue of a wound infection. This strain was subjected to short- and long-read sequencing. Its complete genome contains a single 6,393,703-bp circular chromosome. Phylogenomic analysis of all M. marinum genomes assigned this strain to cluster I.We report the complete genome sequence of a  Mycobacterium marinum is an opportunistic human pathogen  were filtered using NanoFilt v2.8.0 (de novo using Canu v2.2 (https://github.com/nanoporetech/medaka).Genomic DNA was extracted using a QIAamp minikit. Large DNA fragments were recovered using a Blue Pippin recovery system. Barcodes were added using PCR-free EXP-NBD104, adapters were ligated using a SQK-LSK109 connection kit (Oxford Nanopore), and long-read sequencing was performed using the Nanopore PromethION platform. Raw Nanopore signals were subjected to base calling, demultiplexing, and adapter trimming using Guppy v6.3.9  with a sanu v2.2 , Flye v2https://github.com/usadellab/Trimmomatic) and were used to polish the Nanopore assembly with two alternating rounds of Polypolish v0.5.0 . Paired reads  were trimmed using Trimmomatic v0.39 , M. pseudoshottsii JCM15466T (AP018410.1), M. shottsii JCM12657T (AP022572.1), and M. ulcerans Agy99T (CP000325.1) was 98.56%, 98.36%, 97.70%, and 98.09%, respectively, as determined using FastANI v1.33  rotated to start with NI v1.33 . Neverthm 050012 , 18, conM. marinum genome assemblies (n = 51) and SRA reads (n = 88) from NCBI. SRA reads were trimmed using Trimmomatic and assembled de novo using SPAdes v3.15.3 (n = 26) were removed. All genomes (n = 113) were identified as M. marinum as described above. Single nucleotide polymorphisms (SNPs) were called using M. marinum CCUG20998T as a reference using Snippy v4.6.0 (https://github.com/tseemann/snippy) with --ctgs option and Gubbins v3.2.1 with a general time-reversible (GTR) model plus gamma distribution and a 100-bootstrap test (M. marinum strains belong to two major clusters (I and II), consistent with previous findings  and SRR23294010 (Illumina reads), and GenBank nucleotide accession number CP118910.Complete genome sequences of"}
+{"text": "Scientific Reports 10.1038/s41598-023-33920-7, published online 27 April 2023Correction to: The original version of this Article contained errors in the Materials and methods section, under the subheading \u2018Software and hardware specification\u2019, where links to used resources were omitted and the present link was incorrect.https://docs.nvidia.com/deeplearning/frameworks/tensorflow-release-notes/rel_21-03.html).\u201d\u201cThe sMRI preprocessing was done through the SPM12 v7771 software. All further steps used Python 3.8.5 and Tensorflow 2.4.0. The machine learning experiments were performed on an NVIDIA DGX-2 server, within a Docker virtual machine containing 4 CPUs @2.7Ghz and 1 GPU TESLA V100-SXM3-32 GB. All source codes are available at Github (now reads:https://www.fil.ion.ucl.ac.uk/spm/software/spm12/). All further steps used Python 3.8.5 and Tensorflow 2.4.0 (https://docs.nvidia.com/deeplearning/frameworks/tensorflow-release-notes/rel_21-03.html). The machine learning experiments were performed on an NVIDIA DGX-2 server, within a Docker virtual machine containing 4 CPUs @2.7Ghz and 1 GPU TESLA V100-SXM3-32 GB. All source codes are available at Github .\u201d\u201cThe sMRI preprocessing was done through the SPM12 v7771 software (The original Article has been corrected."}
+{"text": "Adherence to antiretroviral therapy (ART) is critical for persons with HIV (PWH) for achieving durable virologic suppression and minimizing drug resistance. This study aimed to characterize ART adherence and its effect on quality of life (QoL) among PWH using real-world data.Data were drawn from the Adelphi HIV Disease Specific Programme\u2122, a cross-sectional survey of physicians and PWH in the U.S., collected between July-2021 and March-2022. At consultation, each of 60 physicians reported demographics, clinical characteristics, ART adherence and satisfaction on 10 consenting PWH. These PWH were then invited to complete questionnaires reporting QoL . Physicians categorized adherence as completely adherent (C-Adh), mostly adherent (M-Adh), or less adherent (L-Adh). Comparisons were made using Chi squared test.Of 578 PWH with physician-reported adherence status; mean [SD] age 44.9 [12.8] years, 70.4% cisgender male, 50.5% White. A total of 389 (67%) were C-Adh, 155 (27%) M-Adh, and 34 (6%) L-Adh. L-Adh (55.9%) and M-Adh (46.5%) PWH had the higher proportion of ART side effects compared to C-Adh (29.3%); p< 0.001. Also, L-Adh (75.8%) and M-Adh (50.8%) were more likely to report current HIV-associated symptoms than C-Adh PWH (29.3%); p< 0.001. C-Adh PWH were perceived as very satisfied with their ART (68.4%), compared to M-Adh 39.4% and L-Adh 23.5%; p< 0.001. Reasons for PWH skipping ART included forgetting, difficulties integrating into routine, and ART side effects . QOL survey was completed by 229 PWH. QoL was higher for C-Adh PWH, measured by PozQol score mean,  and EQ-5D score mean .About one-third of PWH who were not fully adherent to ART, and this was correlated with having poorer QoL. These findings highlight the need to better understand factors leading to sub-optimal ART adherence, to provide targeted interventions to improve quality of life.Girish Prajapati, M.B.B.S., MPH , Merck & Co., Inc.: Employee|Merck & Co., Inc.: Stocks/Bonds Bekana K. Tadese, PhD, MPH, Merck and Co Inc: Employee"}
+{"text": "We report the genome sequences of two genetic variants of grapevine rupestris stem pitting-associated virus (GRSPaV) from Idaho, USA. The coding-complete, positive-strand RNA genome of 8,700 nucleotides contains six open reading frames characteristic of foveaviruses. The two Idaho genetic variants belong to GRSPaV phylogroup 1. Foveavirus (family Betaflexiviridae) (Vitis vinifera L. and its hybrids (\u2013\u2013Grapevine rupestris stem pitting-associated virus (GRSPaV) has flexuous filamentous particles and belongs to the genus viridae) . GRSPaV viridae) . The vir hybrids \u20134. The vhybrids \u2013, 5, 6; ihybrids \u2013\u20139.V. vinifera reference genome using bowtie2 v2.4.4 in local mode  from British Columbia, Canada. The same Riesling sample also yielded three additional contigs of 2,514, 2,893, and 506 nt, which exhibited 98.3%, 98.6%, and 98.8% identity, respectively, to the GRSPaV isolate AMCF clone 3 (GenBank accession number MG938311) from France  contig (GRSPaV-ID1) that spanned six ORFs and, based on the BLASTn program , exhibited 98.5% identity to the GRSPaV isolate 12G412  under BioProject accession number PRJNA931750.The coding-complete genome sequences of GSPaV-ID1 and GSPaV-ID2 are available in GenBank under accession numbers"}
+{"text": "Invasive fungal infections present a significant risk to human health. The current arsenal of antifungal drugs is hindered by drug resistance, limited antifungal range, inadequate safety profiles, and low oral bioavailability. Consequently, there is an urgent imperative to develop novel antifungal medications for clinical application. This comprehensive review provides a summary of the antifungal properties and mechanisms exhibited by natural polyketides, encompassing macrolide polyethers, polyether polyketides, xanthone polyketides, linear polyketides, hybrid polyketide non-ribosomal peptides, and pyridine derivatives. Investigating natural polyketide compounds and their derivatives has demonstrated their remarkable efficacy and promising clinical application as antifungal agents. Candida, Cryptococcus, and Aspergillus species . Th. ThFusar B1 (87) . The C. toxicity . Parnafud D (93) , analogstructure . Parnafu 5 \u03bcg/mL  (Table 194)  shows antifungal activity against C. albicans, C. neoformans, and S. cerevisiae, with MIC values of 2, 2, and 15.6 \u03bcg/mL, respectively [94) has been shown to possess fungicidal activity against C. albicans, C. neoformans, and S. cerevisiae with minimum fungicidal concentrations of 4, 4, and 15.6 \u03bcg/mL, respectively [95) (S. cerevisiae (MIC > 200 \u00b5M), indicating the importance of the aldonic acid group for the antifungal activity of khafrefungin (94) [96) (94), indicating that the aldonic acid group not only enhances the water solubility of khafrefungin (94) but also plays a role in its antifungal activity [97) (94), highlighting the essentiality of the configuration of the 4-methyl group for the antifungal activity of khafrefungin (94) [94) under acidic conditions forms a six-membered lactone derivative (98) (S. cerevisiae (MIC = ~10 \u00b5M) to that of native khafrefungin (94) [The polyketide chain skeleton was catalyzed by type I PKSs and then subjected to complex post-modifications, including glycosylation, to form linear polyketide . There a94) , isolateaffected . Khafrefectively . Additioectively . Notablyely [95)  greatly gin (94) . The pre94) [96)  has beenactivity . Furtherity [97)  has beengin (94) . Additioive (98)  that exhgin (94)  Table 194 (S. ceBacillus laterosporus PNG 276 [99) (62) were administered to human diploid fibroblasts, with basiliskamide A (99) exhibiting minimal cytotoxicity at a concentration of 100 \u03bcg/mL, while cytopathic effect was observed with amphotericin B (62) only 12.5 \u03bcg/mL [99) demonstrated antifungal activity against C. albicans and A. fumigatus, with MIC values of 1 and 2.5 \u03bcg/mL, respectively, whereas basiliskamide B (100) (101) (99), inhibits the growth of C. albicans at a concentration of 25 \u03bcg/mL and has no effect on the growth of A. fumigatus at a concentration of 50 \u03bcg/mL [101) exhibits an additional methylene group compared to the linear polyketide chain of basiliskamide A (99), potentially accounting for the diminished antifungal activity of YM-45722 (101) [Basiliskamide A and B are isolated from  PNG 276 . Basilis276 [99)  and amph.5 \u03bcg/mL . Further B (100)  exhibite0) (101) , an anal50 \u03bcg/mL  (Table 122 (101) .FKS1 and FKS2 genes results in the inhibition of biosynthesis of \u03b2--D-glucan, an important component of the fungal cell wall, which destroys the integrity of fungal cell wall and disrupts the osmotic balance, ultimately leading to fungal death [Candida and Aspergillus species, but they are ineffective against C. neoformans [0 (102) (103) (0 (102) are discussed in this section. Pneumocandin B0 (102) is isolated from the filamentous fungus Glarea lozoyensis [0 (102) is a lipopeptide composed of myristic acid and a hexapeptide ring. PKSs catalyze the assembly of 10, 12-dimethylmyristic acid, and then, catalyzed by a series of enzymes, the polyketide intermediate localizes to NRPSs to acylate the 4, 5-dihydroxyornithine of pneumocandin B0 (102), initiating cyclic hexapeptide elongation.Echinocandins are novel lipopeptide antifungal products synthesized by a heterozygous pathway of non-ribosomal peptides synthases (NRPSs)-PKSs. Compared with azole and polyene antibiotics, echinocandins have a completely different mechanism of action and exert their antifungal effect by destroying the cell wall. Non-competitive binding of echinocandins to the catalytic subunits of \u03b2--D-glucan synthetase encoded by the al death ,83. Echioformans   , the lea2) (103) , is assezoyensis ,88. Pneu104) and 1097 (105) (Burkholderia ambifaria 2.2N [S. cerevisiae (MIC = 0.4 \u03bcg/mL), C. albicans (MIC = 12.5 \u03bcg/mL), and A. niger (MIC = 12.5 \u03bcg/mL) [S. cerevisiae (MIC = 1.6 \u03bcg/mL), A. niger (MIC = 1.6 \u03bcg/mL), and C. albicans (MIC = 12.5 \u03bcg/mL). Burkholdine 1215 (106), 1119 (107), and 1213 (108) (Burkholderia ambifaria [106) exhibits potent antifungal activity against S. cerevisiae (MIC = 0.15 \u03bcg/mL), C. albicans (MIC = 0.15 \u03bcg/mL), and A. niger (MIC = 0.15 \u03bcg/mL) [107) exhibits potent antifungal activity against S. cerevisiae (MIC = 0.1 \u03bcg/mL), C. albicans (MIC = 0.4 \u03bcg/mL), and A. niger (MIC = 0.1 \u03bcg/mL) [108) exhibits potent antifungal activity against S. cerevisiae (MIC = 2.0 \u03bcg/mL), C. albicans (MIC = 31.0 \u03bcg/mL), and A. niger (MIC = 2.0 \u03bcg/mL) [106) and 1119 (107) containing 2,4-diaminobutyric acid adjacent to the 3-OH-Tyr structure is higher than that of burkholdines 1229 (104), 1097 (105), and 1213 (108) containing Asn adjacent to the 3-OH-Tyr structure [106) and 1119 (107) is significantly higher than hemolytic activity, but the antifungal activity of burkholdines 1229 (104), 1097 (105), and 1213 (108) is equivalent to hemolytic activity [107), assigning an Asn replaced a 3-OH-Asn and attaching a \u03b2-xyloside glycosyl portion replaced a \u03b1-xyloside glycosyl portion compared to burkholdine 1215 (106), shows weaker activity against C. albicans (MIC = 0.4 \u03bcg/mL) and stronger activity against A. niger (MIC = 0.1 \u03bcg/mL). Burkholdine 1213 (108), attaching an Asn replaced a 2,4-aminobutyric acid of burkholdine 1119 (107), shows weaker activity against both C. albicans (MIC = 31.0 \u03bcg/mL) and A. niger (MIC = 2.0 \u03bcg/mL). Burkholdine 1229 (104), assigning a 3-OH-Asn replaced an Asn of burkholdine 1213(108), shows stronger activity against C. albicans (MIC = 12.5 \u03bcg/mL) and weaker activity against A. niger (MIC = 12.5 \u03bcg/mL). Burkholdine 1097 (105), removing the \u03b2-xyloside glycosyl portion of burkholdine 1229 (104), shows the same activity against C. albicans (MIC = 12.5 \u03bcg/mL) and stronger activity against A. niger (MIC = 1.6 \u03bcg/mL) (Burkholdines 1229 (97 (105)  are isolria 2.2N . Burkhol5 \u03bcg/mL) . Burkhol13 (108)  are isolmbifaria . Burkhol5 \u03bcg/mL) . Burkhol1 \u03bcg/mL) . Burkhol0 \u03bcg/mL) . The anttructure . The antactivity . Burkhol6 \u03bcg/mL) .109) , C. utilis (MIC = 5 \u00b5g/mL), C. neoformans (MIC = 2 \u00b5g/mL), S. cerevisiae (MIC = 20 \u00b5g/mL), T. mentagrophytes (MIC = 15 \u00b5g/mL), T. rubrun (MIC = 3.9 \u00b5g/mL), M. gypseum (MIC < 0.12 \u00b5g/mL), Blastomyces derniatitidis (MIC < 0.12 \u00b5g/mL), and Hormodendrum pedrosoi (MIC = 15 \u00b5g/mL) [109) has demonstrated significant potent efficacy in vivo, as evidenced by its ability to effectively treat T. mentagrophytes hyphae infection in guinea pigs. Specifically, a hydrophilic ointment containing 0.5% funiculosin had a cure rate of 97% within 10 days [109) to guinea pigs and rabbits was negligible, as evidenced by the survival of guinea pigs injected intraperitoneally with 500 mg/kg of the compound. However, it should be noted that funiculosin is toxic to mice and rats, with an LD50 of 5\u20137 mg/kg for both oral and intraperitoneal administration [Funiculosin (109) , a neutrsum THOM , has beesum THOM ,93 and w5 \u00b5g/mL) ,94 (Tabl110) and skyrin (111)  inhibits the growth of C. albicans ATCC 10231 (MIC = 6.25 \u03bcg/mL), A. niger ATCC 13496 (MIC = 12.5 \u03bcg/mL), and skyrin (111) inhibits the growth of C. albicans ATCC 10231 (MIC = 12.5 \u03bcg/mL) [112)  (Fusarium oxysporum (N17B) and exhibits potent antifungal activity against C. albicans with a MIC of 0.78 \u03bcg/mL [113) inhibits mammalian phosphatidylinositol 3-kinase in vitro and in vivo and membrane-associated phosphatidylinositol 4-kinase of the fungus Schizosacchromyces pombe [114) and B (115) (Microsphaeropsis species RA10-14 collected from the South China Sea [114) and B (115) have great broad-spectrum antifungal activity against C. albicans, Colletotrichum truncatum, Gloeosporium musarum, and Pestalotia calabae (MICs = 1.56\u20133.13 \u03bcg/mL) [116) and C (117) (Amycolatopsis species M39 [116) shows antifungal activity against C. albicans ATCC 10231 (MIC = 10 \u03bcg/mL) and S. cerevisiae ATCC9763 (MIC = 5 \u03bcg/mL) [117) shows weaker antifungal activity against C. albicans ATCC 10231 (MIC = 25 \u03bcg/mL) and S. cerevisiae ATCC9763 (MIC = 20 \u03bcg/mL) [118), hakuhybotrol (119), cladobotric acid A (120), cladobotric acid F (121), cladobotric acid E (122), cladobotric acid H (123), and pyrenulic acid A (124) , B (126), C (127), and D (128) , B (126), C (127), and D (128) show antifungal activity against C. neoformans H99 with MIC values of 8 \u03bcg/mL, 4 \u03bcg/mL, 4 \u03bcg/mL, and 8 \u03bcg/mL, respectively (Emodin (in (111)  are anths ZH-154 . Emodin 5 \u03bcg/mL) . HippolaL) [112)  is a fur\u03bcM [113)  is a fur78 \u03bcg/mL . Wortmanes pombe . Microke B (115)  are isolhina Sea . Microke3 \u03bcg/mL) . Macrote C (117)  are macrcies M39 . Macrote5 \u03bcg/mL) . Macrote0 \u03bcg/mL) . F2928-1 A (124)  are isola FKA-73 . These clentulus . Campafu D (128)  are isolectively .In conclusion, this review provides a comprehensive overview of natural antifungal polyketides encompassing various subclasses such as polyethers, macrolides, xanthones, linear polyketides, anthraquinone, polyphenols, pyridine derivatives, furan derivatives, pyranan derivatives, monophenyl derivatives, macrolactam polyketides, hybrid polyketide non-ribosomal peptides, and other polyketides. Additionally, this review discusses the origin, in vitro and in vivo antifungal activities, structure\u2013activity relationship (SAR), safety profile, mechanism of action, and the impact of structural modifications on the SAR of these polyketides. Previous studies on polyketides have demonstrated the substantial antifungal properties exhibited by certain natural polyketides, such as amphotericin B and caspofungin. This observation suggests that polyketide lead compounds hold considerable potential for the future treatment of fungal infections. Given the remarkable antifungal activities displayed by natural polyketides, this class of compounds has garnered significant interest as a potential therapeutic avenue for fungal infections in the future. Currently, the predominant research emphasis lies in synthesizing novel polyketides, while the clinical utilization of pre-existing polyketide compounds as antifungal medications remains limited. Consequently, further comprehensive and meticulous clinical investigations are imperative to substantiate their efficacy in the future. Moreover, the antifungal properties of unnatural polyketide compounds can potentially be harnessed through combinatorial biosynthesis. Numerous PKSs facilitate the production of primary polyketide compounds, which lack biological activity until they undergo modification by PKS post-modifying enzymes, thereby presenting a promising avenue for exploring new antifungal drugs. Polyketides\u2019 chemical composition and fungicidal properties can be altered by utilizing various post-modification enzymes, including cyclase, aromatase, glycosylase, and halogenase. Unconventional antifungal polyketides have been synthesized by modifying the modules, domains, and subunits of PKSs and employing site-directed mutagenesis techniques. Furthermore, the enhancement of antifungal polyketide production or the acquisition of novel antifungal polyketides can be achieved by combining initiation substrates and elongation units from different hosts and implementing targeted modifications. Only a small number of antifungal natural polyketide compounds have been identified. Many habitats of microorganisms, plants, animals and marine organisms have not been explored, and many antifungal polyketide products urgently need to be discovered. Natural polyketides from microorganisms such as fungi are scarce and difficult to obtain. The solubility, safety, and in vivo bioavailability of natural polyketides should also be considered. Semisynthetic components of natural products will play an important role in developing antifungal candidates in the future."}
+{"text": "Passive immunization via maternal antibodies (Ab) is a key mechanism for infant protection in early life. This study analyzed the landscape of passive immunization induced by SARS-CoV-2 infection and/or vaccination by comparing the magnitude and durability of the transferred Ab.Prospective multicenter observational cohort study of SARS-CoV2-infected and/or vaccinated pregnant women and their infants. We collected maternal and cord blood samples at delivery and neonatal/infant samples at delivery, and at 1, 2, 6 and 12 months (mo) of age. RBD and Spike IgG Ab titers were measured by ELISA.10 ng/ml and from infected and vaccinated mothers [4.61 (4.27-4.93)] log10 ng/ml were higher than those from infected mothers [2.20 (0.10-3.30)] log10 ng/ml, p< 0.001. These differences were observed until 6 mo of age, when only 1/13 (7.7%) infant from infected mothers had detectable maternal Ab compared to 14/15 (93.3%) and 8/8 (100%) from the vaccinated, and infected and vaccinated groups, respectively. At 12 mo none of the infants born to infected mothers had detectable maternal Ab, while in the vaccinated group (n=11), 3 (27.3%) had persistent Ab, and 4 (36.4%) suspected infection. At 12 mo in the infected and vaccinated group (n=6) 3 had no Ab, and 3 had suspected infection. The median [IQR] placental Ab transfer ratio was higher in the vaccinated only group (2.94 [1.34-3.74]) than the infected only group (1.19 [0.33-2.52]) p< 0.01. Linear regression analyses demonstrated that the mean RBD Ab transfer ratio for infants from vaccinated vs infected women was 1.76  higher after adjusting for trimester of infection or vaccination. Similar results were observed with Spike Ab.215 mothers  and 174 infants  were enrolled. At birth RBD Ab titers median [IQR] of infants from vaccinated mothers [4.28 (3.48-4.80)] logDemographic and clinical characteristics of the study populationm; median. IQR; interquartile range. N/A; not applicable.Infants from vaccinated mothers demonstrated maternal Ab up to 6 mo with higher titers than infants born from infected mothers. Antibodies generated by vaccination were transferred more efficiently than those generated by infection regardless of the trimester in which occurred.Asuncion Mejias, MD, PhD, MsCS, Astra-Zeneca: Advisor/Consultant|Merck: Grant/Research Support|Pfizer: Advisor/Consultant|Sanofi-Pasteur: Advisor/Consultant Octavio Ramilo, MD, AstraZeneca: Honoraria|Bill & Melinda Gates Foundation: Grant/Research Support|Janssen: Grant/Research Support|Merck: Advisor/Consultant|Merck: Grant/Research Support|Merck: Honoraria|NIH: Grant/Research Support|Pfizer: Advisor/Consultant|Pfizer: Honoraria|Sanofi: Advisor/Consultant|Sanofi: Honoraria"}
+{"text": "Candida, Aspergillus, and Pneumocystis spp. in BMT.Rezafungin (RZF) once weekly (QWk) is a next-generation echinocandin in development for treatment of candidaemia and invasive candidiasis (IC) and for prevention of invasive fungal disease caused by The Phase 3 ReSTORE treatment trial (NCT03667690) demonstrated RZF QWk noninferiority to caspofungin (CAS) QD. This sub-analysis evaluated efficacy and safety outcomes in patients identified as immunocompromised (immuno-c) during the trial.As previously described, adults with confirmed candidaemia and/or IC were randomized to RZF QWk (400 mg Wk1 then 200 mg QWk) or CAS QD for \u226514 days (\u22644 weeks) with optional oral fluconazole stepdown for CAS. In this sub-analysis, immuno-c patients were those with prior and/or concomitant use of immunosuppressants  and/or medical history ongoing at screening of neutropenia, BMT, SOT, lymphoma or leukaemia.Of 187 patients (mITT population), 90 were immuno-c as defined. Results are for those with data available for analyses: Day 14 global cure\u2014Immuno-c: RZF, 51.1% (23/45); CAS, 55.6% (25/45); \u2013No immuno-c: RZF, 66.7% (32/48); CAS 65.3% (32/49). Day 5 mycological eradication\u2014Immuno-c: RZF, 64.4% (29/45); CAS, 51.1% (23/45); no immuno-c: RZF, 72.9% (35/48); CAS, 71.4% (35/49). Among immuno-c, more had \u22651 AE (96.9%) and \u22651 SAE (64.6%) versus no immuno-c (79.0% and 45.0% respectively).In this sub-analysis of ReSTORE, immune-c status reduced efficacy rates overall but did not change efficacy rate differences between RZF and CAS."}
+{"text": "Wheat (Triticum aestivum L.) is a staple food and major source of dietary calories in Pakistan. Improving wheat varieties with higher grain yield and disease resistance is a prime objective. The knowledge of genetic behaviour of germplasm is key. To achieve this objective, elite wheat varieties were crossed in 4 by 3, line \u00d7 tester design, and tested in 2019 in a triplicate yield trial to estimate genetic variance, general and specific combining ability, mid-parent heterosis and stripe rust (Puccinia striiformis L.). High grain 3358 kg\u00b7ha\u20131 was recorded in F1 hybrid (ZRG-79 \u00d7 PAK-13). Analysis of variance (ANOVA) revealed significant genotypic variance in grain yield. Broad sense heritability (H2) was recorded in the range of 28 to 100 %. General combining ability (GCA) significant for grain yield in parents except FSD-08 and PS-05 was recorded, while specific combining ability (SCA) was recorded to be highly significant for grain yield only in two crosses (ZRG-79 \u00d7 NR-09 and ZRG-79 \u00d7 PAK-13). Mid-parent heterosis was estimated in the range of \u201328 to 62.6 %. Cross combinations ZRG-79 \u00d7 PAK-13 depicted highly significant mid-parent heterosis (62.6 %). Highly significant correlation was observed among spike length, spikelets per spike, plant height and 1000-grain weight. Rust resistance index was recorded in the range of 0 to 8.5. These findings suggest exploitation of GCA for higher grain yield is important due to the presence of additive gene action and selection in the filial generations will be effective with improved rust resistance, while cross combinations ZRG-79 \u00d7 PAK-13 high GCA are best suited for hybrid development. Wheat (Triticum aestivum L.) is an important cereal cropworldwide playing a crucial role in the daily dietary and nutritionalrequirement not only for human beings but also foranimals. It is the major food for one third of world populationand its chief use is the flour for making bread. It is grownaround all continents. Increasing human population, climatechange and global pandemics have an overwhelming impacton food security, especially wheat on crop with current inadequategenetic improvement of wheat to meet future demand.In Pakistan, wheat is grown in an area of 9.2 million ha withthe production of around 25.5 m tonnes and hardly meets the total requirement of the country. But thisfigure is continuously under fluctuation because of stagnantyield of cultivars, disease impact, drought, and floods. Apartfrom these factors, injudicious selection of parental selectionfor a breeding program without prior knowledge of geneticbehaviour in germplasm and lack of indigenous breedingprograms for genetic improvement of wheat is another constraintin the yield.Genetic recombination in germplasm by hybridization isa robust conventional breeding tool for obtaining transgressivesegregants and genetic variation, which provides meansof selection of ideotypes. Gene action and combining abilityanalysis are a most reliable biometric procedure for the studyof genetic behaviour of yield and yield-related components. General combining ability is the averageperformance of genotypes in a series of cross combinations,while specific combining ability is the performance of a particulargenotype in a specific cross combination. Mode ofselection depends based on genetic action in traits of interest.In self-pollinated crops, especially in wheat, plant breedersare usually interested in selection of segregants having additivegene action with high specific combining ability. Additivegene action boosts yield and yield components by cumulativeaddition of genes. Dominance genetic variance exploits heterosisin cross combinations and specific combining abilityprovides the presence of dominant or non-additive gene actionin a particular trait  and provides optimalparental identification . Equalmagnitude of both general and specific combining ability ina breeding population means preponderance of both additiveand dominant genes for the traits of interest; selection in thiscase is most effective for variety development . The term combining ability was first introduced andfurther refined as general combining ability (GCA) and specificcombining ability (SCA) by Sprague and Tatum (1942).GCA distinguishes between the mean performances of parentsin cross combinations whiles SCA is the deviation ofindividual crosses from the average performance of the parentsinvolved. GCA and SCA represent the additive and nonadditiveportions of genotypic variance respectively . The estimates from GCA and SCA provide anassessment of relative merits of the individual genotypes incross combinations to guide selection and testing schemes.Thus, line \u00d7 tester analysis is among the genetic-statisticalapproaches developed to assist in selection of parents based ontheir combining ability and the potential to produce promisingsegregating populations . According toGCA and SCA impacts, positive values are desirable for mostcrop plants characteristics, such as growth and yield-relatedattributes. Negative GCA and SCA impacts, on the otherhand, are desirable for characters where minimum values areessential and appealing, such as early flowering.Heterosis is a phenomenon where F1 hybrids are superiorin traits as compared to their parental genotypes. There areseveral theories that explain the genetic basis of heterosis,including over-dominance, dominance, and genetic balance.The over-dominance theory of heterosis, first proposed byShull and East (1908), suggests that heterozygous individuals,since they carry two different alleles, have an advantage overhomozygous individuals as they carry two identical alleles fora particular gene. This advantage is thought to mean that thetwo different alleles can supplement with each other, leadingto a vigorous phenotype in F1 hybrids. The dominance theory,presented by Jones (1917), suggests that hybrid vigour iscaused by dominant alleles that are more valuable than therecessive alleles. According to this theory, F1 hybrids accedetwo copies of the dominant allele, resulting in a vigorousphenotype. The third heterosis theory is the \u201cLerner\u2019s geneticbalance theory\u201d, suggested by Lerner (1954), that describesthat heterosis is the result of a balance between the expressionof genes that promote growth and those that hamper growth.In F1 hybrids, the expression of growth-promoting genes isincreased, whereas the expression of growth-retarding genesis decreased, leading to better growth and development.Heterotic studies for increasing wheat grain yield have beenan interest of early wheat researchers. Mid-parent heterosisis the percent of the increase or decrease in the F1 value ascompared to the average value of both parents for any metrictrait. In the early green revolution era Pal and Alam (1938)reported mid-parent heterosis (MPH) in wheat. After thegreen revolution and introduction of semi-dwarf wheat varieties,various wheat researchers reported MPH heterosisin wheat . Barbosa-Neto et al. (1996) reported MPH in soft redwinter wheat in the range of \u201320 to 57 %. Liu et al. (1999),Dreisigacker et al. (2005), Basnet et al. (2019) reported MPHin CIMMYT wheat varieties in the range of 9.5 to 14 %.Wheat crop faces numerous challenges that cause yieldlosses, including stripe rust (Puccinia striiformis f. sp. tritici), which is a major disease in areas where cool to mild warmtemperature prevails during the months of February and Marchin the wheat-growing season. Under conducive environmentalconditions, disease causes yield losses ranging from 10 to70 % depending upon susceptibility of genotypes . Development of cultivars containing genetic resistanceis the most cost-effective and environmentally friendlystrategy to mitigate yield losses by stripe rust . Stripe rust spores continue to mutate and evolve newvirulent races causing damage to previously resistant cultivars. Wheat crop in Pakistan has faced severedamage caused by stripe rust pathogen in recent years . Due to climate change and rapid mutation instripe rust pathogen, new races overwintering on alternativehost barberry in hilly areas at high altitudes evolve . Under these circumstances, the already resistantgenotypes become susceptible .There are two types of resistance mechanism against rustpathogens in wheat, vertical resistance, and horizontal resistance.Vertical resistance is conferred by a single gene toa specific pathogenic race of rust, while horizontal resistanceinvolves the use of multiple genes that provide broad spectrumdisease resistance against multiple pathogenic races of rust.There are several resistance genes present in the Pakistanibread wheat varieties that confer resistance against yellow rust,which include Yr5, Yr10, Yr15, Yr17, and Yr18. Qamar et al.(2014) reported Lr34/Yr18 gene complex that confers broadspectrum resistance against yellow rust and leaf rust in mostof Pakistani wheat varieties. Intikhab et al. (2021) reportedthe presence of Lr46/Yr29 gene complex in Punjab-2011 andPirsabak-2005 cultivars that confer resistance against striperust. Khan S.N. et al. (2022) reported the presence of Yr17 andYr5 gene complex in Pakistani wheat varieties Punjab-2011and Pirsabak-2005. Utilization of these resistance sources inthe breeding program for development of varieties resistantagainst stripe rust is an ultimate objective to ensure high yieldon sustainable basis.Various biometrical techniques and breeding designs areused for genetic evaluation and genetic behaviour of germplasmto be utilized in crop breeding programs, but line \u00d7 testeranalysis is an efficient mating design providing reliable informationabout GCA and SCA that ultimately depicts the modeof gene action in a particular trait . GCAand SCA are important to apprehend the genetic architectureof quantitative traits and create the road map for initiation ofan efficient breeding program .Several studies investigating the GCA and SCA effectshave been conducted in wheat. Zhao et al. (2013) reportedsignificant effects for both GCA and SCA for yield and itscomponents and inferred that selecting parental genotypeswith high GCA and SCA effects could lead to the developmentof high-yielding wheat hybrids. Similarly, researchers assessedthe GCA and SCA effects in spring wheat and durum wheatF1 hybrids by using line \u00d7 tester model for combining abilityestimate and concluded that GCA effects were more importantthan SCA effects for grain yield and yield-related traits, andselection of parental genotypes with high GCA effects couldincrease the prospective yield of wheat hybrids . They found thatboth GCA and SCA effects were significant for grain yieldand its components and suggested that selecting parents withhigh GCA and SCA effects could lead to the development ofhigh-yielding wheat hybrids. Selecting parents with high GCAand SCA effects can improve the yield potential and diseaseresistance of wheat hybrids, and the use of line \u00d7 tester designscan provide valuable information about the genetic effects ofparents and their hybridsThe objectives of this study is to elucidate the general andspecific combing ability, heterotic potential, and stripe rust(Puccinia striiformis f. sp. tritici) resistance behaviour ofindigenous elite wheat varieties and their breeding population.Experimental site and plant material. The research was carriedout at the experimental site of a wheat research program,National Agricultural Research Center, Islamabad Pakistanduring 2017\u20132018 wheat growing season. The soil type of thesite is clay loam from 0 to 20 cm, and at the 20\u201340 cm depthit is moderate clay loam. Five widely adopted approved wheatvarieties were used as lines  and three widelyadopted, registered and approved varieties for rainfed areasof Pakistan were used as a tester, namely, NARC-2009,Pakistan-2013 and Borlaug-2016 (Table 1). These testers arewidely adopted and due to their ability to withstand rainfed anddrought-prone areas of Pakistan their leaves have the abilityto stay green during high terminal heat and drought stress.Field experiment and crossing scheme. Eight parents werehybridized to produced 15 F1 cross combinations accordingto line \u00d7 tester crossing fashion as described by Kempthorne(1957) during 2017\u20132018 wheat growing season and crossingwas conducted during March 2018. 15 cross combinationsand seven parents were planted in Randomized CompleteBlock Design (RCBD) with three replications during 2018\u20132019 wheat growing season. In every replication, parents andF1 hybrids were sown in 1 m length with row-to-row spacing25 cm and plant-to-plant spacing 15 cm. The experimentwas conducted in an irrigated field and a total of 6 irrigationswere applied after sowing to harvesting time. Recommendeddoses of fertilizers, i. e. 120 kg N \u00b7 ha\u20131 and 80 kg P \u00b7 ha\u20131, wereapplied. Half of the fertilizers were used at the time of soilpreparation, the second half was applied at the time of tillering,and weedicides were used for eradication of broad leaves and narrow leavesweeds respectively according to the doses mentioned by themanufacturer. Herbicide was applied before the jointing stageof the crop. Leaf area was measured when leaves were fullyturgid and green.Data collection. Grain yield and some yield-related parameterswere measured in parents and hybrid combinations.Grain yield per plant was measured in grams and 1000-grainweight was measured after counting 500 grains of each wheatgrain sample on a counting tray once and the second samplewas repeated for the other 500 grains.Canopy temperature was measured by using a portablethermal gun . Readingsfor canopy temperature were taken at three Feeks stages like booting, kernel water ripening and grainmilking stages . All readings were taken at the angle of 30\u00b0 and above 50 cm of the crop canopy,avoiding land temperature by pointing thermal gum only atthe canopy. The observations were taken between 11:00 amand 14:00 pm under stagnant air conditions and clear sky asdescribed by . Observations for Normalizeddifference vegetative index (NDVI) were recorded 50 cmabove the canopy by using a hand-held Green Seeker with anoptical sensor unit  at three stages ofbooting and grain filling between 11:00 hours to 14:00 hourswith clear sky . Values of NDVI rangefrom \u20131  to +1 (the strongestgreen vegetative stage) .Statistical analysis. Data for other traits  were recorded from6 randomly selected plants. Data recorded were arranged inmean data and subjected to Analysis of Variance (ANOVA)accordingto Steel and Torrie (1980) and Line \u00d7 Tester analysis,according to Kempthorne (1957), combining ability andgene action were studied  byusing R Package agricolae . Genotypic variance and phenotypicvariance were estimated as mentioned by Almutairi (2022) inMS Excel 2016, by using the following formula:Environmental variance was estimated according to Comstockand Robinson (1952). Broad sense heritability was calculatedby using the following formula as described by Burtonand Devane (1953):Heterosis was estimated in percentage increase or decreaseof the F1 hybrids value over mid-parental value by followingthe formula as described by Fonseca and Patterson (1968):Disease observations and scoring. Observations for striperust were recorded at the time of appearance of disease anddata were recorded when rust pathogen was fully developedon leaves of a susceptible check cultivar and leaves\u2019 surfacewas fully covered with rust\u2019s spores. Disease observation wasrecorded in three replicates of each parental line and F1 hybridsaccording to the Cobb Scale method as described byPeterson et al. (1948). The severity of disease was expressedas the percentage of leaf area covered, and 0 % score wasgiven when there was no infection on the leaf and 100 %score was considered when the leaf area was fully coveredwith rust spores and infection. Readings of percent severitywere recorded with the following descriptions for scoringand response values: , response values, coefficient of infection(CI), average coefficient of infection (ACI), country averagerelative percentage attack (CARPA) and rust resistanceindex (RRI) according to Akhtar et al. (2002). The followingformula was used for the calculation of RRI:RRI was calculated by considering the scale of 0 to 9 fromCARPA, where 0 represents a most susceptible genotypeand 9 represents a highly resistant response of the genotypeto rust pathogen.Analysis of variance (ANOVA)Analysis of variance (ANOVA) results presented in Table 1show that the lines  had statistically significant differencesfor all the trails. The testers  showed statisticaldifferences for days to heading, plant height, peduncle length,spike length, days to maturity, grains per spike, 1000-grainweight and grain yield, while non-significant results forflag leaf area, tillers per plant and spikelets per spike wereshown. Interaction of line \u00d7 tester was significant in case ofplant height, flag leaf area, peduncle length, days to maturity,grains per spike and 1000-grain weight. Parents  used in this study provided a broad range of expressionfor various characters as shown in Table 2. There weresignificant differences ( p \u2264 0.05) among the means of genotypes(Table 3) for days to heading (DH), highly significant( p \u2264 0.01) for plant height (PH), flag leaf area (FLA), tillers perplant (TPP), peduncle length (PL), spikelets per spike (SPS),days to maturity (DM), grains per spike (GPS), thousand grainweight (TGW), grain yield (GY) and NDVI value.The values for days to heading (DH) were maximum in thetester  PAK-13 (119 days) and minimum in the lines PB-11, PS-05 and MRJ-08 (117 days). DH are thekey indicator of earliness in crop production. Plant breedersare keen to create new varieties of wheat genotypes with earlymaturity. So, early heading is a desirable trait. Delayed headingleads to a reduction in yield  and earlyheading increases the grain filling duration, which ultimatelyresults in high yield . Plant height (PH)was the highest in the female parent FSD-08 (103 cm) andthe lowest in the male parent NR-09 (83 cm). Minimum PHis preferred due to expected lodging losses. Similarly, thetester NR-09 (83 cm) can be assessed for developing droughttolerant variety with reduced plant height for future breedingprograms. Likewise, minimum flag leaf area (FLA) is alsodesirable for drought tolerance due to reduced transpirationlosses from a reduced area exposed to sunlight. The testers PAK-13 and BOR-16 showed the minimum values offlag leaf area: 29.57 and 29.83 cm2, respectively. Pedunclelength was longest in the female parent PS-05 (16.67 cm) andshortest in the male parent NR-09 (7.20 cm). PS-05 producedthe maximum grain yield (2531.8 kg \u00b7 ha\u20131) while minimumgrain yield (1696.1 kg \u00b7 ha\u20131) was recorded in ZRG-79.Mean performance of parents and their cross combinationsMean performance for line, testers and cross combinationsfor days to maturity (DH) ranged from 117 to 119 days. Theparental lines FSD-08, PB-11, PS-05, MRJ-08, ZRG-79 andBOR-16 were revealed to have 117 DH, while NR-09 andPAK-13 had 118 and 119 days to heading, respectively (seeTable 2). Among F1 hybrids Zargoon-79 \u00d7 Pakistan-2013 had119 days for heading while the rest of the cross combinationsshowed 117 DH. The grand means for parents, crosses, linesand testers were 117.67, 117.56, 117.27 and 118.33, respectively.The coefficient of variance 0.67 % obtained for DHwas also in the acceptable range.Average minimum plant height was recorded in NARC-2009 (83 cm) followed by cross combination of Punjab-2011 \u00d7Pakistan-2013 (86 cm), and maximum plant height of 104 cmwas recorded in the cross-combination Faisalabad-2008 \u00d7Borlaug-2016 followed by one of parent viz. Faisalabad-2008(103 cm). Grand mean, coefficient of variance (CV) and leastsignificant variance (LSD) for plant height of lines, testers andtheir parental combinations was revealed to be 95.92 cm, 3.64and 5.73, respectively.The cross-combination Punjab-2011 \u00d7 Pakistan-2013showed minimum value (26.8 cm2) for flag leaf area followedby the lines Pakistan-2013 (29.5 cm2) and Borlaug-2016(29.8 cm2). Maximum leaf area was recorded in the linePunjab-2011 (39.2 cm2) followed by the F1 combination,Faisalabad-2008 \u00d7 Pakistan-2013 (36.9 cm2). Grand mean,CV, LSD and standard error for leaf area of lines, testers andcross combinations was recorded as 10.46, 5.7 and 2.0 cm2respectively.Maximum 13 tillers per plant (TPP) was recorded in thetester Pakistan-2013 followed by 12.3 tillers in Borlaug-2016while minimum 7.6 tillers were observed in the female parentPunjab-2011 followed by Pirsabak-2005 (8.33). In thecross combinations a maximum of 10 tillers was recorded inPunjab-2011 \u00d7 NARC-2009 and Miraj-2008 \u00d7 NARC-2009and grand mean for TTP was recorded as 9.49 with CV, LSDand SE 11.66, 3.61 and 1.27 respectively.Maximum peduncle length was observed in the line Pirsabak-2005 while minimum peduncle length was recordedin the tester parent NARC-2009. Grand mean for pedunclelength was observed to be 11.96 cm with CV 9.7 % and LSD(\u03b1 0.05) value 1.9.Mean performance for spike length (SL) was observed12.36 cm in parents and their cross combinations. MaximumSL was observed in cross combinations Miraj-2008 \u00d7Pakistan-2013 followed by Miraj-2008 \u00d7 Borlaug-2016, butminimum SL was observed in the parental line Faisalabad-2008.Grand mean for spikelets per spike (SPS) for parents andcross combination was recorded as 20.33 with a maximumof 21.9 spikelets observed in Pakistan-2013 in addition to theF1 hybrid combination of Miraj-2008 \u00d7 Pakistan-2013 and inthe Pirsabak-2005 \u00d7 Pakistan-2013. Grains per spike (GPS)was recorded maximum in parental line Punjab-2011. Averagethousand grain weight was calculated to be 34.52 withhigher TGW in Faisalabad-2008 (43.73 g) and the crosscombination of Faisalabad-2008 \u00d7 Pakistan-2013 (43.53 g),and lower value for TGW was depicted by the parental lineNARC-2009 (22.47 g). Average grain yield was obtainedin all the parents and cross combinations (2344.5 kg \u00b7 ha\u20131),while average GY was higher in crosses as compared to theparents\u2019 grain yield, the maximum was recorded in the crosscombination ZRG- 79 \u00d7 PAK-13 (3358 kg \u00b7 ha\u20131), followed byPB-11 \u00d7 NR-09 (2820 kg \u00b7 ha\u20131), while minimum grain yieldwas recorded in the cross combination of ZRG-79 \u00d7 NR-09(1372 kg \u00b7 ha\u20131).Maximum normalized differences in vegetative index(NDVI) value was observed in the parental line PS-05 (0.73)followed by PB-11 \u00d7 PAK-13 and PS-05 \u00d7 NR-09 with thesame value. Average NDVI value for parents and crosses wasrevealed to be 0.67; lines, testers and crosses also containedsimilar values for NDVI.There were significant differences among the means ofcrosses combinations for almost all the traits studies exceptDH, TPP an SL. The lines  also depictedhighly significant differences in all the parameters under considerationexcept SL. The testers  also revealedhighly significant differences for all the traits except for FLA,SL, SPS, and NDVI. Interaction of lines \u00d7 testers depictedhighly significant differences in their mean performance forthe traits of PH, FLA, PL, DM, GPS, TGW and NDVI value.Estimates of genetic variance componentsEstimation of genotypic variance, phenotypic variance, environmentalvariance, variance due to general combining ability,variance due to specific combining ability and variance dueto GCA over SCA is mentioned in Table 4.Phenotypic variance was depicted more as compared togenotypic variance in some traits, i. e. PH, FLA, SL, DM,and GY, while only GY showed high environmental variance.Broad sense heritability (H2) was estimated in the rangeof 28.11 % (GY) to 100 % (CT). PH, PL, GPS, TGW, NDVIvalue and CT showed broad sense heritability of more than91 %, while DH and GY depicted less heritability. The traitswith high genetic variance, low environmental variance andhigh broad sense heritability have preponderance of additivegenes and these are stable characters and selection in the filialgenerations can be made by keeping eye on these traits. Grainyield (GY) and DH attained low broad sense heritability andshowed that environmental influence is more important forthe expression of these traits. Selection in the filial generationshould be made for these traits by considering diseaseincidence and drought proxy parameters, i. e. NDVI and CTvaluesProportional contribution of lines, testers,and their interactions to total varianceProportional contribution of total variance for yield andyield-related metric traits for lines, testers and their crosscombinations was estimated . For DH, PH, TPP andCT it was recorded to be higher as compared to the testersand combinations of both lines and testers. Contribution ofL\u00d7T to total variance was recorded as high in FLA, PL, SL,SPS, DM, GPS and NDVI value, while variance contributionof testers to TGW and GY was estimated higher as comparedto the lines and L\u00d7T combinations.General combining abilityGeneral combining ability (GCA) estimates for all the traitsare given in Table 5. Both positive and negative GCA effectswere observed for lines and testers. For DH, the value of GCAeffects ranged between 0.00 and 0.56. As a good general combiner,significant positive (0.56) and negative (\u20130.56) GCAeffectswere observed for lines Zargoon-79 and Pirsabak-2005,respectively. Similarly, in the testers, positive and significant(0.51) GCA effect was observed for Pakistan-2013 only (seeTable 5). Patel et al. (2020) demonstrated ( p \u2264 0.01) significantnegative and desirable GCA effects in lines and non-additivegene action was primarily involved in days to heading.For plant height, negative general combining ability effectsare more important since more emphasis is placed upon selectionfor short stature in segregating the population because itultimately turns out that a short stature line is more responsiveto fertilizer and tolerant to lodging. In this study, GCA effectsranged between \u20135.82 and 3.24 for PH. Significant positive(3.07) and negative (\u20135.82) GCA effects were observedfor the lines Pirsabak-2005 and Punjab-2011, respectively.Similarly, highly significant positive (3.24) was estimated forthe tester BOR-16 and highly significant but negative (\u20132.56)GCA effects were observed for the testers PAK-13, respectively.These results are in accordance with the results of.For flag leaf area (FLA), negative general combining abilityeffects are more important because FLA is much influencedby transpiration losses due to disclosure to sunlight, whicheventually affects the grain yield. Hence, more emphasis isretained on the selection of genotypes with smaller FLA.From that, among the female parents, Pirsabak-2005 andPunjab-2011 showed a highly significant negative GCA effect:\u20132.16 and \u20132.10, respectively. On the other side, no significantGCA effects were observed among the testers for FLA. Theseresults confirm the findings of .In case of tillers per plant (TPP), GCA effects ranged between\u20130.58 and 0.98. As a good general combiner, highlysignificant positive (0.98) GCA effects were observed only forthe line MRJ-08 while there were no significant GCA effectsamong the testers for TPP. To begin with, TPP is a significantyield-boosting characteristic that contributes to increased grainyield. A higher number of tillers per plant confirms optimalplant populations and as a result higher grain yield . For this point of view, the female line MRJ-08showed better performance. These findings are in accordancewith the results of .GCA effects ranging between \u20131.16 and 1.39 were observedfor peduncle length (PL). Highly significant positive (1.39)and negative (\u20130.76) GCA effects were observed for the lines FSD-08 and ZRG-09, respectively. In the same way,highly significant positive (0.65) and negative (\u20131.16) GCAeffects were observed for the testers BOR-16 and PAK-13, respectively. Likewise, in PH, shorter PL is preferred becausean increase in PL ultimately increases the PH and we prefera plant with short stature. In current study, two female parents,ZRG-79 (\u20130.76) and MRJ-08 (\u20130.70), showed negative generalcombining ability. Also, one male parent, PAK-13, showedsuperior general combining ability for this trait. So, it can beconcluded that the above-mentioned parents are desirable foruse in the breeding program. The findings of  supported the results.Greater spike length (SL) and larger number of spikeletsper spike (SPS) are essential for enhanced yield. Among parents,one line , MRJ-08, showed significant positivevalues(0.77) for SPS. One tester, Pakistan-2013, exhibitedhigh GCA for SPS. These results were quite close to thefindings of . Number of grains per spike (GPS) is also animportantfactor for enhanced grain yield. Therefore, positiveGCA effects are more important due to positive contributionof grain yield. Among male parents, only NR-09 showedpositive and higher values (3.26) of GCA effects for GPS.Among female parents, MRJ-08 and PS-05 showed positiveand higher values, i. e. 2.90 and 2.02 respectively. It shouldbe noted that values of male parents were higher than thoseof female parents. These findings match with the results of. These results are different fromthe findings of Nazir et al. (2005).For grain yield per plant (GY), only one female parentMRJ-08, and among the male parents, BOR-16 and NR-09,exhibited positive general combining ability effects. Similarresults were also found by .Specific combining abilitySpecific combining ability (SCA) estimates for all the traitsare given in Table 6. Both positive and negative SCA effectswere observed among the crosses.As for SCA effects for DH, all the fifteen crosses were ofnon-significant nature with positive and negative magnitude(see Table 6). The result indicates the involvement of bothadditive and non-additive genetic effects in the inheritance ofDH, with greater proportion of additive genetic effect. Lineswith maximum SCA effects can be used in development ofhybrid cultivars. Only six among fifteen crosses depicted negativeSCA effects for plant height. If parents with tallness arethe ideal ones, then the crosses FSD-08 \u00d7 NR-09, FSD-08 \u00d7PAK- 13, PB-11 \u00d7 NR-09, PB-11 \u00d7 PAK-13, PS-05 \u00d7 BOR-16and MRJ-08 \u00d7 BOR-16 would be considered good. However,the remaining crosses exhibited higher SCA effects. Thesefindings confirmed the results of . Furthermore,non-additive type of gene action is detected for PH and supportedby . Also, our results concur withAli F.K.H. and Abdulkhaleq (2019) for plant height.GCA effects for flag leaf area range from negative \u20133.80 topositive 3.33. Roughly 50 % of the crosses showed smallervalues of SCA effects for flag leaf area, which is desirable.As less flag leaf area is required for drought tolerance, thecrosses with significant SCA effects, i. e. FSD-08 \u00d7 BOR-16and PB-11 \u00d7 PAK-13 may be used in a future breeding programbecause they have high negative SCA values contributingtowards minimum FLA. However, the remaining crossesexhibited higher positive SCA effects for FLA. Comparableresults have also been stated by .Negative SCA effects are needed to reduce the pedunclelength (PL). In this study, two crosses showed significantlynegative SCA effects. FSD-08 \u00d7 NR-09 and MRJ-08 \u00d7 BOR- 16are the best hybrids for reduced PL. Similar results were reportedby .In case of spike length (SL), all the fifteen crosses were ofnon-significant nature with positive and negative magnitude(see Table 6). For a number of SPS, positive specific combiningability effects were shown in 6 out of 15 crosses but onlytwo crosses, FSD-08 \u00d7 NR-09 and PB-11 \u00d7 BOR-16, havesignificant GCA effects. These hybrids performed best andcan be suggested for future breeding programs. These resultsare in the conformity with those of .For grain yield per plant, SCA effects found varied muchamong crosses. The poorest cross with respect to SCA forgrain yield per plant was ZRG-79 \u00d7 PAK-13 whereas the crossthat appeared to be the best and the most promising specificcombination was ZRG-79 \u00d7 NR-09. Positive specific combiningability effects were displayed in 8 out of 15 crosses.But only ZRG-79 \u00d7 NR-09 showed such significant positiveeffects among crosses. Similar results were also reported by.Mid-parent heterosis estimation for grain yieldMid-parental heterosis (MPH) for GY was estimated for15 F1 hybrids . F1 hybrids ZRG-79 \u00d7 PAK-13 showedhigher mid-parental value (62 %) followed by FSD-08 \u00d7PAK- 13, ZRG-79 \u00d7 BOR-16, and PB-11 \u00d7 NR-09, whichrevealed mid-parent heterosis value above 30 % . Cross combinations PS- 05 \u00d7 PAK- 13,MRJ- 08 \u00d7 NR-09, MRJ-08 \u00d7 PAK-13, FSD- 08 \u00d7 NR- 09 depictedmid-parental heterosis value more than 15 %. Threecross combinations, ZRG-79 \u00d7 NR-09, MRJ-08 \u00d7 BOR- 16and PB-11 \u00d7 BOR-16, depicted negative heterosis.Cross combinations with more than 30 % mid-parentalheterosis can be used in hybrid breeding in wheat. Heteroticstudies for increasing wheat grain yield has been an interestof early wheat researchers. Pal and Alam (1938) reportedmid-parent heterosis in the pre-green revolution era. After theintroduction of semi-dwarf wheat in the post-green revolutionera, various wheat researchers reported mid-parent heterosisin wheat, i. e. . Barbosa-Neto et al. (1996) reported MPH in red softwinter wheat in the range of \u201320 to 57 %. Liu et al. (1999),Dreisigacker et al. (2005), Basnet et al. (2019) studied MPHin CIMMYT wheat varieties and reported MPH in the rangeof 9.5 to 14 %. Parental lines and tester used in present studieshave CIMMYT background and the majority of the genotypesexhibited similar results for MPH. However, crosses combinationZRG-79 \u00d7 PAK-13 has one indigenous parent ZRG-79and exhibited a high percentage of MPH. These finding candemonstrate that crosses among parents with CIMMYT backgroundhave low heterotic potential and additive gene actiongoverned the GY potential in these cross combinations andselection in the filial generation will be key for transgressive segregants, but in case of crosses among indigenous parentsand genotypes with a CIMMYT parent it will be good sourceof hybrid breeding.Correlation study of agronomic traitsCorrelation study among yield and related traits under rainfedconditions of eight parents and 15 wheat crosses is mentionedin Figure 3. High significance was observed between PH andTGW with the value of 0.9 ( p < 0.001), SL and SPS had acorrelation coefficient value of 0.72 ( p <0.001) followed byDH and DM with 0.57 ( p <0.01). PL also showed highlysignificant and positive correlation with TGW and PH . PL and TGW also revealedsignificant but negative correlation with DH \u20130.6 and \u20130.5,respectively ( p < 0.01).Positive and significant correlation among PH and PL withTGW showed that the higher the plant height the higher thethousand grain weight and peduncle length. Careful considerationshould be made while selecting the genotypes with stiffand strong stem girth to avoid lodging. Correlation betweenSL and SPS revealed that an increase in spike length leadsto an increase in spikelets per spike, genotypes with longspikes will be a good selection criterion for increasing yielddue to the increase in number of spikelets per spike. Positiveand significant correlation among DH and DM depicted thatgenotypes with early DH would mature earlier, so selectionof genotypes with early flowering is good for early maturityand short duration variety development. Significant but negativecorrelation between TGW and DH indicated that a delayin days to flowering leads to a reduced TGW and vice versa.TGW showed negative correlation with TPP and these findingsare in line with the results of Almutairi (2022). Low correlationof GY with other parameters in wheat was also reportedby Gowda et al. (2010).Stripe rust responses of parental linesand their cross combinationsThe response to stripe rust (Puccinia striiformis f. sp. tritici)on parental lines used in the study and their offspring(crosses) is recorded for disease scoring, coefficient of infection(CI), average coefficient of infection (ACI), country averagerelative percentage attack (CARPA) and rust resistanceindex (RRI) (Table 7). All the parental lines showed moderateresistant (MR) to highly resistant (R) reaction against striperust (Pst). The female parents (lines) FSD-08, PB-11, PS-05,MRJ-08 and ZRG-79 showed 20M, 20M, 5M, 30M and 40Mscores respectively, while the pollen parents (testers) viz.PAK-13, BOR-16 and NR-09 depicted 10MR, 5R and 40Mresponse against stripe rust. All these parents showed a slowrusting response against rust pathogen that is under the controlof multiple genesCross combinations of these parental lines showed a variedresponse, moderately resistant to moderately susceptible reaction against stripe rust. The F1 hybrids combinations PS- 05 \u00d7PAK-13, PS-05 \u00d7 BOR-16 and PS-05 \u00d7 NR-09 showed 10MR,10MR and 5MR reaction, the crosses FSD-08 \u00d7 PAK- 13,FSD-08 \u00d7 BOR-16, PB-11 \u00d7 PAK-13, PB-11 \u00d7 BOR-16, andMRJ-08 \u00d7 BOR-16 showed 20M reaction, the cross combinationMRJ-08 \u00d7 PAK-13 showed 30M reaction, while the restof the crosses showed moderately susceptible to susceptiblereaction against stripe rust.Average coefficient of infection (ACI) for the parentsPS- 05, PAK-13, BOR-16 and FSD-08 was recorded as 2.0,6.7, 4.3 and 14.0 respectively and these varieties revealeda very good level of resistance against stripe rust. Rust resistanceindex (RRI) of these parents was also high (ranged 5.8to 8.5), which indicated a good resistance response of thesevarieties. Among the cross combinations, PS-05 \u00d7 BOR-16,MRJ-08 \u00d7 PAK-13, PB-11 \u00d7 BOR-16, PB-11 \u00d7 PAK-13 andPS-05 \u00d7 NR-09 depicted ACI values of 4.0, 8.3, 12.0, 13.1, and14.7, respectively. These F1 hybrids had a resistant responseto stripe rust. RRI value of the F1 hybrids  was higher (ranging from 5.6 to 8.1).The higher the RRI value and the lower the ACI valuemeans of genotypes with a resistant response to the diseasepathogen and under the influence of slow rusting genes, theslower the disease progress and the lesser the yield losses.Genotypes with higher RRI values (>5.0) represent moderatelyresistant to highly resistant response against rust pathogen.The parental genotypes viz. PS-05, PAK-13 and BOR-16had higher values for RRI showing a highly resistant response against stripe rust pathogen.Cross combinations revealed an intermediate responseagainst stripe rust as compared to parents, especially testes,and resistant genes are under the control of additive geneaction. These results indicate that repeated backcross can bea better strategy for accumulation of resistant genes in thesecross combinations. Selection in these cross combinations byfollowing backcrosses with recurrent parents is efficient fordisease resistance in the filial generations. These results arevery much in line with the findings of Afzal et al. (2009) andMahmoud et al. (2015).According to these findings, it can be concluded that highergeneral combining ability and low broad sense heritabilityfor grain yield suggest the presence of additive genes, andexploitation of general combining ability for high grain yieldis important due to presence of additive gene action, and selectionin the filial generations and family rows will be effective. 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+{"text": "Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacterales with resistance to carbapenems or third-generation cephalosporins. The aim of the present study was to investigate the in vitro activity of the novel siderophore cephaloporin cefiderocol (CID) and four comparator \u03b2-lactam-\u03b2-lactamase-inhibitor combinations and to give insights into the genetic background of CID-resistant isolates. In total, 301 clinical Enterobacterales and non-fermenting bacterial isolates were selected for this study, including randomly chosen isolates  and challenge isolates . Isolates displayed CID MIC50/90 values of 0.12/0.5 mg/L (set I) and 0.5/1 mg/L (set II). Overall, the CID activity was superior to the comparators against A. baumannii, Stenotrophomonas maltophilia and set II isolates of P. aeruginosa. There were eight CID-resistant isolates detected (MIC > 2 mg/L): A. baumannii (n = 1), E. cloacae complex (n = 5) and P. aeruginosa (n = 2). Sequencing analyses of these isolates detected the acquired \u03b2-lactamase (bla) genes blaNDM-1,\u00a0blaSHV-12 and naturally occurring blaOXA-396, blaACT-type and blaCMH-3. In conclusion, CID revealed potent activity against clinically relevant organisms of multidrug-resistant Enterobacterales and non-fermenters. Antimicrobial resistance poses a global threat to public health. Of great concern are  Acinetobacter baumannii (carbapenem-resistant), Pseudomonas aeruginosa (carbapenem-resistant) and the order of Enterobacterales (carbapenem-resistant and ESBL-producing) [Stenotrophomonas maltophilia [The emergence of antibiotic-resistant bacteria has been described as one of the biggest threats to global health and food safety ,2. It isoducing) . Variousoducing) ,7. New aoducing) ,9. Howevtophilia ,9. P. aeruginosa [A. baumannii (CRAB), P. aeruginosa and S. maltophilia, as well as carbapenem-resistant Enterobacterales [Klebsiella pneumoniae or OXA-23 in A. baumannii), show the weak hydrolysis of CID [P. aeruginosa and Enterobacter cloacae complex, which could otherwise cause resistance development under therapy [Cefiderocol (CID) is a novel parenteral cephalosporin carrying a catechol moiety at the 3-position side chain . It has ruginosa . It is tcterales , with his of CID . Further therapy ,15,16. C therapy ,18. In vitro data of CID from Germany are scarce. In our previous study published in 2020, CID was found to inhibit 97.2% of a randomly chosen collection of 213 Gram-negative clinical isolates of different species at the investigational susceptibility breakpoint of \u22642 mg/L . The isoThe present study aimed (I) to investigate the in vitro activity of CID against Gram-negative pathogens recovered from patients during a more recent multicentre surveillance study conducted by the PEG in 2016/17 and (II) to compare it with the susceptibility against novel \u03b2-lactam\u2013\u03b2-lactamase-inhibitor (BL-BLI) combinations. 50 and MIC90) and the number and percent of susceptible and resistant isolates were calculated with the available breakpoints of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) (P. aeruginosa) of \u22642 mg/L or the non-species-related pharmacokinetic\u2013pharmacodynamic (PK/PD) breakpoint of \u2264 2 mg/L was applied .MIC distribution data of CID are presented in (EUCAST) . OverallEnterobacterales isolates, CID inhibited 98.2% (109/111)  was determined for a respiratory E. cloacae complex isolate (PEG-16-51-23) that originated from a nosocomial infection. Sequencing revealed that this isolate encoded the extended-spectrum \u03b2-lactamase SHV-12. There were two more isolates detected with CID MICs > 2 mg/L: an NDM-1-encoding P. aeruginosa (PEG-16-96-12) and another E. cloacae complex (PEG-16-75-70) harbouring blaSHV-12 (both MICs 4 mg/L). Among the 111 109/111) . In compP. aeruginosa isolates, with 89.7% of the isolates being inhibited at the respective breakpoint (R > 8 mg/L). The susceptibility rates of P. aeruginosa against the other agents were comparable to that seen with the Enterobacterales: 98.3% (CID), 96.6% (IMR), 94.8% (CTV) and 91.4% (CTT)  was more active than CTT (1/4 mg/L), CTV (2/8 mg/L), IMR (0.5/2 mg/L) or MEV (1/\u2265 16 mg/L). Of the nine A. baumannii isolates, all were inhibited by CID at \u22640.5 mg/L. Based on the MIC50/90 values, all comparator compounds were less active with MIC50 values of 0.5\u2013\u2265 16 mg/L and MIC90 values of \u226516 mg/L. Two OXA-23-encoding A. baumannii isolates (PEG-16-19-60 and PEG-16-22-42) were resistant to IMR (R > 2 mg/L) and exhibited MEV MICs of 16 mg/L. Among the S. maltophilia (n = 17) isolates, CID MICs ranged from \u22640.03 to 2 mg/L, with MIC50/90 values of 0.06 mg/L and 0.5 mg/L. In comparison, all four comparators were less active in S. maltophilia with MIC50/90 values of \u226516 mg/L. Compared to Enterobacterales, MEV was less effective against the 58 4% (CTT) . Based o50/90 and susceptibility/resistance rates, where applicable).The results of set II isolates are displayed together with set I in P. aeruginosa) of \u22642 mg/L or the non-species-related PK/PD breakpoint (A. baumannii) of \u22642 mg/L was applied. There were five isolates detected with MICs > 2 mg/L A. baumannii , E. cloacae complex  and P. aeruginosa . Whole-genome sequencing analysis detected the presence of carbapenemase gene blaNDM-1 in the A. baumannii isolate together with disrupted oprD and piuA genes. The resistant P. aeruginosa isolate harboured blaOXA-396, a variant of the chromosomal class D OXA-50-group in this species, together with the class A \u03b2-lactamase PDC-8, and carried a disrupted oprD. The three E. cloacae complex isolates were only positive for class C ACT-type \u03b2-lactamases or CMH-3, which naturally occur in E. cloacae.The CID MICs ranged from \u22640.03 to 32 mg/L, with an overall susceptibility rate of 95.3% (101/106) if the specifies-specific EUCAST susceptibility breakpoint , 51.3% (CTV), 46.2% (IMR) and 35.9% (MEV), while CID inhibited 97.4% (38/39) of P. aeruginosa isolates. Among the CRAB isolates, CID revealed the most potent activity compared to the other compounds with MIC50/90 values of 0.12/2 mg/L as opposed to \u226516/\u226516 mg/L.Among the 53 Enterobacterales isolates, 100% were susceptible to MEV, 98.1% were susceptible to both CTV and IMR and 77.4% were susceptible to CTT. With a susceptibility rate of 94.3% (50/53), CID exhibited slightly lower activity compared to CTV and IMR in Enterobacterales. Overall, the comparator agents were similar or less effective in the 39 The CID MIC distributions of ESBL-producing isolates, carbapenemase-producing isolates and colistin-resistant isolates from set I and II are summarized in 50/90 values of 0.5/1 mg/L. There was no difference in CID MIC distribution with regard to different resistance genes, with the exception of two blaNDM-1-encoding isolates (A. baumannii and P. aeruginosa) and two SHV-12-encoding E. cloacae complex isolates with CID MICs > 2 mg/L. With the exception of CTT, the comparator compounds revealed similar activity against ESBL-producing isolates compared to CID. The MIC50/90 values were 0.25/0.5 mg/L for CTV, 0.12/0.25 mg/L for IMR and \u22640.06/0.12 mg/L for MEV. The CID susceptibility in 47 ESBL-encoding isolates ranged from \u22640.03 mg/L to \u226564 mg/L, with MICThe overall CID susceptibility of the 47 ESBL-encoding Enterobacterales isolates was 95.7% (45/47). CTV, IMR and MEV were able to inhibit 100% of the isolates at their respective breakpoints, with only the CTT susceptibility being lower at 85.1%. 50/90 values were 0.5/2 mg/L. Fourteen of fifteen carbapenemase-producing A. baumannii isolates (93.3%) were inhibited at a CID concentration \u2264 2 mg/L. Regarding carbapenemase-producing P. aeruginosa isolates, 91.7% (11/12) exhibited CID susceptibility. CID-resistant isolates of both species (A. baumannii PEG-16-19-65 and P. aeruginosa PEG-16-96-12) encoded blaNDM-1 and revealed MIC values of the comparator substances of 16 mg/L each. With the exception of one A. baumannii isolate, all carbapenemase-encoding isolates were susceptible to colistin. The colistin-resistant isolate (MIC 8 mg/L) was susceptible to CID but resistant to all of the other tested compounds. Overall, the comparators revealed reduced activity in carbapenemase-encoding isolates compared to CID with MIC50/90 values greater or equal to their highest concentration tested. Among the 30 carbapenemase-producing isolates, CID MICs ranged from \u22640.03 to 32 mg/L, with 93.3% (28/30) of the isolates being inhibited at a concentration of \u22642 mg/L. The MIC50/90 values of 0.25/1 mg/L. Based on MIC50/90 values, CID activity was comparable to CTT (0.5/2 mg/L), CTV (0.5/4 mg/L) and IMR (0.25/1 mg/L). The distribution of MEV MICs revealed a lower MIC50 value compared to the other compounds (MIC50/90s: \u22640.06/2 mg/L). All colistin-resistant P. aeruginosa isolates (n = 14) were inhibited by CID and the comparator substances. The two colistin-resistant A. baumannii isolates (PEG-16-36-64 and PEG-16-50-50) were inhibited by a CID concentration < 2 mg/L but revealed different MIC results with regard to the comparator substances. PEG-16-36-64 was susceptible against all the other substances (MIC range 0.25\u20130.5 mg/L), while the blaOXA-23-like-positive PEG-16-50-50 exhibited MIC values of 16 mg/L against CTT, CZA, IMR and MEV.In colistin-resistant isolates (n = 37), CID MICs ranged from \u2264 0.03 to 4 mg/L, with MICA. baumannii and P. aeruginosa, as well as Enterobacterales species with acquired resistance against carbapenems or third-generation cephalosporins, all possessing a high risk of severely limited treatment options. In contrast to resistance against third-generation cephalosporins, carbapenem resistance is still rarely encountered in Enterobacterales species such as E. coli and K. pneumoniae in Germany. For example, the surveillance study originated by the Paul-Ehrlich-Society for Infection Therapy in 2016/17 revealed resistance rates of 0% against imipenem and meropenem in 571 E. coli isolates and 1.6% (5/318) against both substances in K. pneumoniae isolates. According to the annual surveillance data for Germany reported to the European Centre for Disease Prevention and Control (ECDC), the rates of carbapenem-resistant E. coli and K. pneumoniae isolates were 0.0% and 0.8% in 2021  compared to resistance rates against third-generation cephalosporins of 9.1%  and 10.4% (678/6538) (Surveillance Atlas of Infectious Diseases (europa.eu); data source: invasive isolates). In Acinetobacter spp. and P. aeruginosa, carbapenem-resistance was more frequently detected with 4.3% (26/605) and 14.8% (425/2864), respectively.The WHO has designated antimicrobial resistance as one of the top ten global public health threats. Of great concern are carbapenem-resistant non-fermenting Gram-negative bacteria such as A. baumannii, MBL-producing organisms and S. maltophilia, which is intrinsically resistant against multiple antimicrobial agents, including carbapenems [The antimicrobial agents compared in this study are considered promising compounds in the treatment of infections with the above-mentioned organisms when no other options are available. In contrast to the comparators, CID possesses activity against a variety of Gram-negative species and \u03b2-lactamases of all Ambler classes, including OXA-encoding bapenems . Unlike 50/90s 0.12/0.5 mg/L) and 95.3%  at \u22642 mg/L  were inhibited by CID at a concentration of \u22642 mg/L, while there was only one isolate detected among fourteen CRAB isolates (set II) with an MIC of >2 mg/L. However, due to the small sample size of S. maltophilia and A. baumannii, these data should be considered with caution. Naas et al. investigated a total number of 103 S. maltophilia and 161 A. baumanni isolates and reported comparable CID activity against both species and almost identical MIC50/90 values of 0.06 mg/L and 0.25 mg/L [S. maltophilia [50/90 values of 0.06/0.25 mg/L. However, studies on CRAB yielded conflicting results. A recent study by Mushtaq et al. investigated 99 A. baumannii isolates encoding various OXA-\u03b2-lactamases, with the majority being OXA-23 (n = 41), as well as NDM enzymes (n = 20) [A. baumannii (n = 25), while isolates harbouring OXA-58 or OXA-23 were all susceptible (n = 75) [Our study observed more potent CID activity against parators . Similar.25 mg/L . Karlowstophilia . They deA. baumannii and P. aeruginosa) and SHV-12 (E. cloacae complex), or the naturally occurring \u03b2-lactamases such as class C ACT-type, CMH-3 (both E. cloacae), as well as PDC-8 together with class D OXA-396 (P. aeruginosa)  . The cor0.5 mg/L ,27. In oruginosa ,29. FurtblaNDM-1 might be an exception to this and requires further investigation. CID activity was superior to the comparators against A. baumannii, S. maltophilia and challenge isolates of P. aeruginosa. In addition to ESBL-producing isolates, the majority of CP-producing and colistin-resistant isolates were inhibited at a CID concentration \u22642 mg/L, indicating good activity of CID in clinically relevant organisms. In conclusion, CID revealed potent activity against Enterobacterales and non-fermenting Gram-negative bacterial isolates from Germany. The association of CID non-susceptibility with a particular resistance determinant seemed to be unlikely, while the presence of In total, 301 Gram-negative bacterial isolates were investigated in this study. All isolates were obtained from patient samples collected at 22 German microbiological laboratories during a multicentre surveillance study conducted by the PEG in 2016/17. The majority of laboratories were affiliated with tertiary-care medical centres. Two sets of isolates were selected: random samples (set I) and challenge organisms (set II).E. coli (n = 52), K. pneumoniae (n = 34) and E. cloacae complex (n = 25) isolates. The non-fermenting bacteria included A. baumannii (n = 9), P. aeruginosa (n = 58) and S. maltophilia (n = 17) isolates. Set I included 195 isolates  which encompassed 111 Enterobacterales and 84 non-fermenting bacteria which were obtained from the respiratory tract (n = 117) and from blood culture (n = 78). Isolates were randomly selected; routine clinical isolates, also including ESBL- and carbapenemase-producing isolates; as well as colistin-resistant isolates. The Enterobacterales included E. coli and five K. pneumoniae isolates were ESBL producers, which were investigated via PCR/Sanger sequencing or whole-genome sequencing in different reference laboratories as part of the PEG study. All of the ESBL K. pneumoniae isolates as well as five ESBL E. coli isolates carried the blaCTX-M-15 gene. Additionally, three of these K. pneumoniae isolates also encoded SHV-40 (n = 1) or SHV-28 (n = 2). The remaining ESBL E. coli isolates carried blaCTX-M-1 (n = 4) or blaCTX-M-27 (n = 1). Few isolates of set I encoded carbapenemases: K. pneumoniae (n = 1) encoded VIM-1, A. baumannii (n = 2) encoded OXA-23 and P. aeruginosa (n = 1) encoded NDM-1. Seven Isolates were colistin-resistant (E. cloacae (n = 3), E. coli (n = 1) and K. pneumoniae (n = 3)).Among the Enterobacterales, ten A. baumannii and P. aeruginosa isolates with a meropenem MIC above 8 mg/L and Enterobacterales that were either ertapenem-resistant (R > 0.5 mg/L) and/or possessed a meropenem MIC of >0.12 mg/L . In contrast to set I, isolates of set II were especially enriched for ESBL- and carbapenemase-producing isolates as well as colistin-resistant isolates. The investigation of resistance genes was part of the multicentre study conducted by the PEG and was performed via PCR/Sanger sequencing or whole-genome sequencing in different reference laboratories [A. baumannii isolates of this set encoded the following \u03b2-lactamases: NDM-1 (n = 1), OXA-58 (n = 1), or OXA-23 (n = 11). The E. coli isolates of this set produced CTX-M-1 (n = 3), CTX-M-14 (n = 2), CTX-M-15 (n = 9), CTX-M-15 plus CTX-M-27 (n = 1), CTX-M-27 (n = 3), or CTX-M-55 (n = 1). P. aeruginosa isolates of this set encoded the following \u03b2-lactamases: GIM-1 (n = 2), IMP-7 (n = 2), IMP-13 (n = 1), VIM-1 (n = 2), VIM-2 (n = 3) and VIM-5 (n = 1). K. pneumoniae isolates of this set encoded the following \u03b2-lactamases: CTX-M-3 (n = 1), CTX-M-15 (n = 10) and VIM-1 (n = 1). One E. cloacae complex isolate encoded OXA-48.Set II comprised 53 Enterobacterales and 53 non-fermenting bacteria with either confirmed carbapenemases or ESBL genes or with phenotypic colistin resistance or \u201emeropenem non-susceptibility\u201d. \u201eMeropenem-non-susceptible\u201d isolates included ratories ,31. A. bThe verification of species identification was performed via matrix-assisted laser desorption ionization\u2013time of flight (MALDI-TOF) mass spectrometry .5 CFU/mL (range 2\u20138 \u00d7 105). Panels were incubated at 35 \u00b1 1\u00b0C for 18 \u00b1 2 h. The MICs were read visually and, as far as possible, interpreted according to the species-specific clinical breakpoints approved by the EUCAST , I  and R (resistant) [P. aeruginosa of \u22642 mg/L (S) and >2 mg/L (R) were applied. For other species, the EUCAST-approved pharmacokinetic\u2013pharmacodynamic  breakpoints were applied (\u22642 mg/L (S) and >2 mg/L (R)). Reference strains E. coli ATCC 25922 and P. aeruginosa ATCC 27853 were used for quality control. The following antimicrobial agents were tested with the noted ranges: ceftazidime\u2013avibactam (CTV) (0.12/4\u20138/4 mg/L), ceftolozane\u2013tazobactam (CTT) (0.25/4\u20138/4 mg/L), imipenem\u2013relebactam (IMR) (0.03/4\u20138/4 mg/L), meropenem\u2013vaborbactam (MEV) (0.25/8\u20138/8 mg/L) and cefiderocol 0.06\u201332 mg/L). MICs were determined using the broth microdilution procedure with geometric twofold serial dilutions according to the international standard ISO 20776-1 [ mg/L. MIsistant) . For CIDIsolates with CID MICs > 2 mg/L were sent to the International Health Management Associates (IHMA) for whole-genome sequencing.p < 0.05 was assumed. The statistical significance of differences in susceptibility rates was judged by comparing 95% confidence intervals (CIs). Intervals were constructed using the Newcombe\u2013Wilson method without continuity correction. If no rate was contained in the CI of the other one, significance of"}
+{"text": "Tenacibaculum haliotis strain RA3-2T , isolated from Korean wild abalone . As the only strain for this Tenacibaculum species worldwide, the information is of use for comparative genomic analyses delineating Tenacibaculum species.Here, we present the draft genome sequence of  Tenacibaculum isolates are associated with fish (Tenacibaculum crassostreae from the Pacific oyster (Crassostrea gigas)  (Most ith fish . Two mola gigas)  and Tenahannai) .T. haliotis strain, RA3-2T. A sample of RA3-2T was obtained from the Japanese Biological Resource Centre in glass ampoules (L-dried). The strain was grown on marine agar 2216 (BD Difco). The plates were incubated at 20\u00b0C for 3\u2009days .DNA was extracted from three colonies using the InstaGene matrix (Bio-Rad) per the manufacturer\u2019s instructions. The 16S rRNA gene was PCR amplified using the universal primer pair 27F and 1492R (The NucleoSpin soil kit (Macherey-Nagel GmbH & Co. KG), purified genomic DNA, and the NanoDrop Lite spectrophotometer were used to measure the DNA concentration (Thermo Fisher Inc.). The genome was sequenced by Macrogen. A DNA library was prepared using the TruSeq Nano DNA kit and sequenced using the Illumina NovaSeq 6000 platform  to generate 31,942,898 paired-end reads with an average length of 151\u2009bp.https://www.bioinformatics.babraham.ac.uk/projects/fastqc/). The next-generation sequencing reads were preprocessed using Geneious Prime v.2020.2.2 software , with a total of 2,977,857\u2009bp and 30.7% G+C content. The quantitated assembly quality indicated a minimum contig length of 1,346\u2009bp, a maximum length of 486,103\u2009bp, and an average length of 102,684\u2009bp. The draft genome sequence contained 2,728 coding DNA sequences (CDSs) with protein, 2,794 genes, 8 pseudogenes, and 61 RNAs , as determined by the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v.6.2 (https://github.com/ncbi/pgap) using the best-placed reference protein set, with GeneMarkS-2+ as the annotation method. The predicted tRNAs were verified using tRNAscan-SE v.2.0  rea v.5.2.0 . The assin silico genome analysis, genes were discovered with predicted associations to secretion systems (type I [T1SS] and T9SS), as detected using TXSScan . Iron-related protein families were identified  using FeGenie v.1.0 (Using ie v.1.0 . Using tie v.1.0 .JAOBTH000000000. The version described here is JAOBTH010000000. The sequences are publicly available under the BioProject accession number PRJNA877611, BioSample accession number SAMN30712836, and SRA accession number SRR22752080. The GenBank accession number for the 16S rRNA gene sequence is OP381101.This whole-genome shotgun project was deposited at DDBJ/ENA/GenBank under the accession number"}
+{"text": "Introduction: Varicella-zoster virus (VZV) reactivation was not listed as a side effect of any vaccine until introduction of COVID-19 vaccines. Since COVID-19 vaccines were introduced, several case reports described the link between COVID-19 vaccination and VZV reactivation. In our study we aimed to investigate how often patients developed VZV reactivation after mRNA COVID-19 vaccine compared to influenza vaccine.We used the TriNetX platform and its global health research network containing aggregated de-identified information of electronic health records on over 100 million patients from scores of healthcare organizations. Three patient groups were identified:st mRNA COVID-19 vaccine 12/20/2020-12/20/2022Patients receiving their 1nd mRNA COVID-19 vaccine 12/20/2020-12/20/2022Patients receiving their 2Patients receiving influenza vaccine 12/20/2018-12/20/2022Propensity Score Matching was used for age, gender, and Zoster vaccine status.We examined the prevalence of VZV reactivation in each group during the first 21 days following vaccination.st mRNA COVID-19 vaccine or the Influenza vaccine, 0.04% in the COVID-19 vaccine group and 0.15% in the influenza group, odds ratio (OR) 0.24 (95% CI 0.20-0.29), were diagnosed with VZV reactivation within 21 days. Among 445,382 patients in each matched group receiving either their 2nd mRNA COVID-19 vaccine or their influenza vaccine, 0.04% in the COVID-19 vaccine group and 0.13% in the Influenza group, OR 0.27 (95% CI 0.22-0.32), were diagnosed with VZV reactivation within 21 days. There was a lower risk of VZV reactivation after 1st or 2nd dose of mRNA COVID-19 vaccine than influenza vaccine.Out of 444,016 patients in each matched group receiving either their 1st or 2nd dose of mRNA COVID-19 vaccination is smaller than VZX reactivation after influenza vaccine. The risk of VZV reactivation after mRNA COVID-19 vaccine is very low and hesitance to receive COVID-19 vaccine due to concern of VZV reactivation is not substantiated.The risk of VZV reactivation after the 1David C. Kaelber, MD, PhD, MPH, FAAP, FACP, FACMI, FAMIA, Becton, Dickinson and Company: Advisor/Consultant|Dynavax Technologies Corporation: Advisor/Consultant|Merck Sharp & Dohme Corporation: Advisor/Consultant"}
+{"text": "Under many kinds of stress, eukaryotic cells rapidly decrease the overall translation level of the majority of mRNAs. However, some molecular mechanisms of protein synthesis inhibition like phosphorylation of eukaryotic elongation factor 2 (eEF2), which are known to be functional in animals and yeast, are not implemented in plants. We suggest that there is an alternative mechanism for the inhibition of protein synthesis in plant cells and possibly, in other eukaryotes, which is based on the discrete fragmentation of 18S rRNA molecules within small ribosomal subunits. We identified four stress-induced small RNAs, which are 5\u2019- and 3\u2019-terminal fragments of 18S rRNA. In the present work, we studied the induction of 18S rRNA discrete fragmentation and phosphorylation of the \u03b1-subunit of eukaryotic initiation factor 2 (eIF2\u03b1) in germinated wheat embryos in the presence of glyphosate, which imitates the condition of amino acid starvation. Using northern and western blotting, we have shown that stress-induced 18S rRNA fragments started to accumulate in wheat embryos at glyphosate concentrations that did not evoke eIF2\u03b1 phosphorylation. It was also found that cleavage of 18S rRNA near the 5\u2019-terminus began much earlier than eIF2\u03b1 phosphorylation, which became noticeable only at higher concentration (500 \u03bcM) of glyphosate. This result suggests that discrete fragmentation of 18S rRNA may constitute a regulatory mechanism of mRNA translation in response to stress and may occur in plant cells in parallel with and independently of eIF2\u03b1 phosphorylation. The identified small 5\u2019- and 3\u2019-terminal fragments of 18S rRNA that accumulate during various stresses may serve as stress resistance markers in the breeding of economically important plant crops. Protein biosynthesis is a very energy-intensive process,so under stress conditions, the translation of most cellularmRNAs is inhibited in order to save energy and resources andto ensure preferential synthesis of stress proteins. Several molecularmechanisms of protein synthesis inhibition have beendescribed in mammalian and yeast cells. One of these mechanismsis the eukaryotic translation elongation factor 2 (eEF2)phosphorylation, which is carried out by a highly specific proteinkinase in response to a sharp decrease in cytosolic ATPconcentration levels. Phosphorylation inactivates mammalianeEF2 by preventing it from binding to the ribosome . However, plants do not exhibit endogenous kinaseactivity for eEF2 either under normal conditions , or under stress .The second mechanism known in animals to reduce the levelof mRNA translation is triggered under conditions of aminoacid starvation and is mediated by eIF4E-binding proteins(4E-BPs), which prevent eIF4E from binding to the m7Gcapstructure of mRNA . However,no clear homologs of these eIF4E-BPs have yet been foundin plants , nor were anyorthologues of the 4E-BPs genes found in plant genomes.Another important mechanism of eukaryotic protein synthesisinhibition is the phosphorylation of the \u03b1-subunit ofeukaryotic initiation factor 2 (eIF2\u03b1) by specific protein kinases.This process in mammalian and yeast cells leads tothe blocking of GDP \u2192 GTP exchange protein eIF2B and toa sharp inhibition of the mRNA translation initiation . However, recycling of the ternary complex inplant cells can occur without the participation of eIF2B , and eIF2\u03b1 phosphorylation in plant systemsin vitro does not lead to strong inhibition of protein synthesis. In addition, of the four protein kinases that phosphorylate theeIF2\u03b1 in mammalian cells, only pGCN2-kinase was foundin plants, and the phosphorylation of eIF2\u03b1 in plants is not auniversal response to all stress types .Thus, the mechanisms of protein biosynthesis suppressiondue to the phosphorylation of translational factors, which arewell described for mammals and yeast, are either used to alimited extent or are not realized at all in plant cells. We suggestthat another mechanism of protein synthesis inhibitioncan function in plants, which is triggered by certain abiotic andbiotic stresses. In our understanding, this mechanism is associatedwith the cleavage at certain sites of the 18S rRNA as partof 40S ribosomal subunits (40S RS). Previously, we describedthe process of 18S rRNA cleavage, leading to 5\u2032-terminal fragmentsformation of 132\u2013134 nt. and of 54\u201357 nt. , as well as a 3\u2032-terminalfragment of 100 nt. . Our data arequite consistent with the data of full-transcriptome analysis,which showed that breaks in 28S-, 18S-, and 5.8S-rRNA donot occur randomly, but discretely, which leads to the fact thatsome fragments of ribosomal RNA are detected in the cell significantlymore often than other fragments .The process of RNA cleavage is widely used by cells duringthe processing of ribosomal RNA from their precursor duringribosome biogenesis . In addition, in proandeukaryotic organisms, the mechanism of protein biosynthesissuppression is realized due to the cleavage of the 28SrRNA molecule from the large (60S) ribosomal subunit alongthe sarcin-ricin loop with the cleavage of the 3\u2032-terminal EndorRNA-fragment . Toxins of plants , fungi (\u03b1-sarcin) and bacteria (Shiga toxin) actthis way . Possibly similar endonucleasesand/or glycosylases  are activatedin plant cells during stress, but targeting 18S rRNA in 40S RSinstead of 28S rRNA in 60S RS, and thus leading to temporaryor permanent suppression of mRNA translation.In this work, we have shown that in the case of glyphosatemediatedamino acid starvation, when the only specific eIF2\u03b1kinase of plants (pGCN2-kinase) is activated, in addition toplant eIF2\u03b1 phosphorylation, another protective mechanismis triggered in plant cells, namely, discrete fragmentation of18S rRNA. It was shown that the accumulation in plants of18S rRNA 5\u2032-terminal fragments of 75 nucleotides (75nt-5\u203218S) and 134 nucleotides (134nt-5\u203218S) begins earlier thanthe activation of pGCN2 kinase and becomes noticeable atrelatively low concentrations of glyphosate when plant eIF2\u03b1phosphorylation does not occur at all.Plant material and treatment. Wheat (Triticum aestivum L.cv. Kazakhstanskaya 10) seeds were sterilized in 70 % (v/v)ethanol for 2 min, then in 2 % (w/v) NaOCl for 20 min, andwashed thoroughly with sterile water. Seeds were germinatedat 26 \u00ba\u0421 on sterile filter paper soaked in water. After 18 hours,viable embryos were isolated by spatula from swollen seedsand placed in 1 % glucose solution containing 50 U/ml penicillin,50 \u03bcg/ml chloramphenicol, and 50 \u03bcg/ml nystatin. Afterthis, embryos were divided into equal portions (1 g), whichwere subjected to treatment with glyphosate (simulation ofamino acid starvation) or without any additives (control).Synthesis of probes. DIG-labeling of de novo synthetizedoligodeoxyribonucleotides 5\u203218S (5\u2032-ACAAGCATATGACTACTGGCAGGATCAACCAGGTA) and 3\u203218S (5\u2032-CAATGATCCTTCCGCAGGTTCACCTACGGAAACCT)was carried out using DIG Oligonucleotide 3\u2032-End LabelingKit (Roche) according to the manufacturer\u2019s manual. Probes(5\u203218S-DIG and 3\u203218S-DIG) were used for northern blottingNorthern blotting. Total RNA was extracted from planttissues with Tri-reagent (Sigma Aldridge) and analyzed on10 % PAGE with 8 M urea in Tris-borate buffer . RNAs were blottedto a nylon membrane (Roche) equilibrated in 0.1x TBEusing a semi-dry blotter (Sci-Plas) at 250 mA for 30 min.The membrane was dried and irradiated with UV light for2 min at 10 mJ/cm2 in a crosslinker (UVP). Hybridization ofDIG-labeled probes and subsequent chemiluminescent banddetection was performed with DIG Luminescent DetectionKit for Nucleic Acids (Roche) according to the manufacturer\u2019sprocedure. The hybridization temperature was 55 \u00b0C.Anti-Digoxigenin-AP Fab fragment conjugates (Roche) wereused to detect bound DIG-labeled probes. The blots weredeveloped using a commercial alkaline phosphatase substrateCSPD (Roche).SDS-polyacrylamide gel electrophoresis (SDS-PAGE)and immunoblotting. Frozen embryos were ground toa powder in a mortar and then homogenized in Laemmlisample buffer . Proteins were separated by12.5 % SDS-PAGE with 0.1 % SDS. The separated proteinswere transferred to a nitrocellulose membrane (GVS) thatwas afterwards stained with Ponceau S (Sigma-Aldrich). Theantibodies against human phospho-eIF2\u03b1 (S51) produced inrabbit  were used for theimmune-detection of phosphorylated T. aestivum (Ta) eIF2\u03b1(TaeIF2(\u03b1P)). Then horseradish peroxidase-conjugated antirabbitsecondary antibodies produced in donkey  were used.The effect of glyphosate concentration on 18S rRNA fragmentationin wheat embryos. Since the mechanisms ofmRNA translation inhibition mediated by 4E-BPs and eEF2Kare not implemented in plants, it is believed that the mainresponse in plants to amino acid starvation is eIF2\u03b1 phosphorylationwith pGCN2 kinase . To testwhether the process of discrete fragmentation of 18S rRNAis also induced under these conditions, the herbicide glyphosatewas used. Glyphosate targets 5-enolpyruvoylshikimate3-phosphate synthase, which catalyzes the key penultimatereaction in the shikimate pathway .Therefore, it inhibits the synthesis of many aromatic plantmetabolites including the amino acids tryptophan, tyrosine,and phenylalanine and leads to pGCN2 kinase activationand phosphorylation of the plant eIF2\u03b1 .Germinated wheat embryos were treated with glyphosate atvarious concentrations, after which the content of 18S rRNAsmall fragments and the phosphorylation status of TaeIF2\u03b1were assessed in their cells. The results are present in Figure 1.Phosphorylation of TaeIF2\u03b1 becomes noticeable only atrelatively high concentrations (0.5 and 5 \u03bcM) of glyphosate, at which wheatembryos stopped to grow . Theappearance of 3\u2032-terminal fragments 100nt-3\u203218S and 70nt-3\u203218S was observed at the same concentrations of glyphosate. At the same time,5\u2032-terminal fragments of 18S rRNA, 134nt-5\u203218S and 75nt-5\u203218S, began to accumulate in noticeable amounts even atvery low concentrations (5 \u03bcM) of glyphosate .The results of semi-quantitative optical densitometry analysisfor this experiment are presented in Table 1. Since the 134nt-5\u203218S fragment can be a precursor of 75nt-5\u203218S, and the100nt-3\u203218S fragment can act as a precursor for 70nt- 3\u203218S,it is reasonable to estimate the sum of these small 18S rRNAfragments.The dynamics of glyphosate influence on 18S rRNA fragmentationin wheat embryos. Then, we assessed how quicklywheat embryos respond to glyphosate treatment by measuringthe time dependence of TaeIF2\u03b1 phosphorylation and of discretefragmentation of 18S rRNA. For this, a glyphosateconcentrationof 500 \u03bcM was chosen, which induced quite effectivephosphorylation of TaeIF2\u03b1, as well as a significantincrease in the content of 18S rRNA small fragments: 134nt-5\u203218S, 75nt-5\u203218S, 100nt-3\u203218S and 70nt-3\u203218S . The results of the experiment are shown in Figure 2.The results of semi-quantitative optical densitometry analysisof the data presented in Figure 2 are shown in Table 2.Data presented in Figure 2 and Table 2 show that TaeIF2\u03b1phosphorylation begins 45 min after the start of glyphosatetreatment , and TaeIF2\u03b1P becomes quite noticeable after 60 minof such treatment .The 3\u2032-terminal fragmentation of 18S rRNA is observedafter 3 hours after the start of glyphosate treatment: fragments100nt-3\u203218S and 70nt-3\u203218S become detectable as faintlyvisiblebands on track 7 of Figure 2, b (right panel). Theamount of these 3\u2032-coterminal fragments is significantly lowerthan after 10 hours of the same treatment with glyphosate.As for fragmentation from the 5\u2032-terminus of 18S rRNA,the amount of both 5\u2032-coterminal fragments, 134nt-5\u203218Sand 75nt-5\u203218S, is significantly increased as early as by the15th min after the start of treatment of wheat embryos withglyphosate . Notably, the amountof the fragment 134nt-5\u203218S is higher than that of 75nt-5\u203218Sfragment during 30\u201345 min of incubation with glyphosate.By 60 min of incubation their amounts become almost equaland after that, the amount of 75nt-5\u203218S fragment becomeshigher (by 90 min) and even obviously prevalent by 180 min. Similar interrelation can be seenin Figure 1, b (right panel) regarding the applied concentrationsof glyphosate.These observations suggest that cleavage at 134th nucleotidemay happen more quickly and this site is more susceptibleat the beginning of stress. The cleavage site at 75th nucleotidebecomes more prevalent with an increase of stress durationand severity. The cleavage sites at the 3\u2032-terminal segmentof 18S rRNA occur only at very high severity and durationof stress. Therefore, there seemingly exist several differentmechanisms for the cleavage at 5\u2032- and 3\u2032-termini of 18SrRNA, which may result in several different consequencesfor the functioning of 40S RS.No phosphorylation of eIF2\u03b1 was observed in plants underosmotic and oxidative stresses , heatshock and during unfolded protein response in plants . At the same time, during these stresses a significantdecrease in the translation level of most mRNAs is observedwith exception only for those templates that are responsiblefor the synthesis of stress proteins . Most likely, in plants, other mechanismsof protein biosynthesis suppression are realized,than eIF2\u03b1 phosphorylation . In addition,eIF2\u03b1 phosphorylation is not the only possible mechanism ofresponse to some types of stress in different eukaryotic cells.For example, when yeast cells are exposed to harsh ultravioletlight, phosphorylation of eIF2\u03b1 is observed, as well as a suppressionof the overall level of protein synthesis. However,inhibition of mRNA translation upon exposure to UV lightoccurs even in cells containing a mutant form of eIF2\u03b1 that isnot capable of phosphorylation .We postulate that the process of discrete fragmentationof 18S rRNA observed under glyphosate mediated amino acid starvation may lead to a decrease in the level of mRNAtranslation. This molecular mechanism can be realized inparallel with the known mechanism of translational regulationmediated by eIF2\u03b1 phosphorylation and independentlyof it.Understanding the molecular mechanisms of plant adaptationto stresses can make it possible to increase the efficiencyof breeding work to obtain genetic lines and varieties ofeconomically important plant species that are characterizedby increased resistance to certain stresses.This paper presents data indicating that in plant cells the imitationof amino acid starvation induces, in addition to eIF2\u03b1phosphorylation, another cellular response that involves thecleavage of the 18S rRNA molecule with the formation ofdiscrete 5\u2032- and 3\u2032-terminal fragments. At the same time,3\u2032-terminal fragments of 18S rRNA appear only at lethalconcentrations of glyphosate and after a prolonged period ofstress (3 hours or more). In contrast, 5\u2032-terminal fragments of18S rRNA began to accumulate in wheat embryos at relativelylow glyphosate concentrations, at which wheat embryos couldcontinue development, and already 15 min after the start ofglyphosate treatment. Thus, the process of 18S rRNA fragmentationin wheat embryo 40S RS is triggered even underconditions where eIF2\u03b1 phosphorylation does not occur. Wesuggest that such cleavage of the 18S rRNA molecule, whichis activated during amino acid starvation, may result in eitherglobal or selective suppression of mRNA translation.The authors declare no conflict of interest.Altschuler M., Mascarenhas J.P. Heat shock proteins and effects of heatshock in plants. Plant Mol. Biol. 1982;1(2):103-115. DOI 10.1007/BF00024974.Baird Th.D., Wek R.C. Eukaryotic initiation factor 2 phosphorylationand translational control in metabolism. Adv. Nutr. 2012;3:307-321.DOI 10.3945/an.112.002113Ballard D.J., Peng H.-Y., Das J.K., Kumar A., Wang L., Ren Y.,Xiong X., Ren X., Yang J.-M., Song J. Insights into the pathologicroles and regulation of eukaryotic elongation factor-2 kinase. Front.Mol. Biosci. 2021;8:839. DOI 10.3389/fmolb.2021.727863.Browning K.S., Bailey-Serres J. Mechanism of cytoplasmic mRNAtranslation. Arabidopsis Book. 2015;13:e0176Chen Z., Sun Y., Yang X., Wu Z., Guo K., Niu X., Wang Q., Ruan J.,Bu W., Gao S. Two featured series of rRNA-derived RNA fragments(rRFs) constitute a novel class of small RNAs. PLoS One. 2017;12:e0176458. DOI 10.1371/journal.pone.0176458Echevarria-Zomeno S., Yanguez E., Fernandez-Bautista N., Castro-Sanz A.B., Ferrando A., Castellano M.M. Regulation of translationinitiation under biotic and abiotic stresses. Int. J. Molec. Sci. 2013;14:4670-4683. DOI 10.3390/ijms14034670.Endo Y. Mechanism of action of ricin and related toxins on the inactivationof eukaryotic ribosomes. Cancer Res. Treat. 1988;37:75-89.DOI 10.1007/978-1-4613-1083-9_5.Gallie D.R., Le H., Caldwell C., Tanguay R., Hoang N.X., BrowningK.S. The phosphorylation state of translation initiation factorsis regulated developmentally and following heat shock in wheat.J. Biol. Chem. 1997;272:1046-1053. DOI 10.1074/jbc.272.2.1046.Gallie D.R., Le H., Caldwell C., Browning K.S. Analysis of translationelongation factors from wheat during development and followingheat shock. Biochem. Biophys. Res. Comm. 1998;245:295-300. DOI10.1006/bbrc.1998.8427.Henras A.K., Plisson-Chastang C., O\u2019Donohue M.F., Chakraborty A.,Gleizes P.E. An overview of pre-ribosomal RNA processing ineukaryotes.Wiley Interdiscip. Rev. RNA. 2015;6(2):225-242. DOI10.1002/wrna.1269.Hernandez G., Altmann M., Lasko P. Origins and evolution of themechanisms regulating translation initiation in eukaryotes. TrendsBiochem. Sci. 2010;35(2):63-73. DOI 10.1016/j.tibs.2009.10.009.Immanuel T.M., Greenwood D.R., MacDiarmid R.M. A critical reviewof translation initiation factor eIF2\u03b1 kinases in plants \u2013 regulatingprotein synthesis during stress. Funct. Plant Biol. 2012;39(9):717-735. DOI 10.1071/FP12116.Kamauchi S., Nakatani H., Nakano C., Urade R. Gene expression inresponse to endoplasmic reticulum stress in Arabidopsis thaliana.FEBS J. 2005;272(13):3461-3476. DOI 10.1111/j.1742-4658.2005.04770.x.Kast A., Klassen R., Meinhardt F. rRNA fragmentation induced bya yeast killer toxin. Mol. Microbiol. 2014;91(3):606-617. DOI10.1111/mmi.12481.Knutsen J.H., R\u00f8dland G.E., B\u00f8e C.A., H\u00e5land T.W., Sunnerhagen P.,Grallert B., Boye E. Stress-induced inhibition of translation independentlyof eIF2\u03b1 phosphorylation. J. Cell Sci. 2015;128(23):4420-4427. DOI 10.1242/jcs.176545.Laemmli U.K. Cleavage of structural proteins during the assembly ofthe head of bacteriophage T4. Nature. 1970;227(5259):680-685.DOI 10.1038/227680a0.Lageix S., Lanet E., Pouch-Pelissier M.N., Espagnol M.C., Robaglia C.,Deragon J.M., Pelissier T. Arabidopsis eIF2\u03b1 kinase GCN2 is essentialfor growth in stress conditions and is activated by wounding.BMC Plant Biol. 2008;8:134. DOI 10.1186/1471-2229-8-134.Padgette S.R., Kolacz K.H., Delannay X., Re D.B., LaVallee B.J., TiniusC.N., Rhodes W.K., Otero Y.I., Barry G.F., Eichholtz D.A., PeschkeV.M., Nida D.L., Taylor N.B., Kishore G.M. Development, identificationand characterization of a Glyphosate-tolerant soybean line.Crop Sci. 1995;35(5):1451-1461. DOI 10.3389/fpls.2016.01009Ruberti C., Kim S.J., Stefano G., Brandizzi F. Unfolded protein responsein plants: one master, many questions. Curr. Opin. Plant Biol.2015;27:59-66. DOI 10.1016/j.pbi.2015.05.016.Shaikhin S.M., Smailov S.K., Lee A.V., Kozhanov E.V., Iskakov B.K.Interaction of wheat germ translation initiation factor 2 with GDPand GTP. Biochimie. 1992;74(5):447-454. DOI 10.1016/0300-9084(92)90085-s.Smailov S.K., Lee A.V., Iskakov B.K. Study of phosphorylation oftranslation elongation factor 2 (EF-2) from wheat germ. FEBS Lett.1993;321(2-3):219-223. DOI 10.1016/0014-5793(93)80112-8.Zhang Y., Wang Y., Kanyuka K., Parry M.A., Powers S.J., Halford N.G.GCN2-dependent phosphorylation of eukaryotic translation initiationfactor-2alpha in Arabidopsis. J. Exp. Bot. 2008;59(11):3131-3141. DOI 10.1093/jxb/ern169Zhanybekova S.S., Polimbetova N.S., Nakisbekov N.O., Iskakov B.K.Detection of a new small RNA, induced by heat shock, in wheat seedribosomes. Biochemistry (Moscow). 1996;61:862-870.Zhigailov A.V., Alexandrova A.M., Nizkorodova A.S., Stanbekova G.E.,Kryldakov R.V., Karpova O.V., Polimbetova N.S., Halford N.G.,Iskakov B.K. Evidence that Phosphorylation of the \u03b1-subunit ofeIF2 does not essentially inhibit mRNA translation in wheat germcell-free system. Front. Plant Sci. 2020;11:936. DOI 10.3389/fpls.2020.00936.Zhigailov A.V., Polimbetova N.S., Borankul R.I., Iskakov B.K. Investigationof discrete fragmentation of 18S rRNA within 40S ribosomalsubparticles of plant cells. Vestnik KazNU. Biological Series.2013;2:81-87. (in Russian)Zhigailov A.V., Polimbetova N.S., Doshchanov Kh.I., Iskakov B.K.Detection in plant cells of a new 75-nucleotide cytoplasmic RNAcorresponding to the 5\u2032-terminal fragment of 18S RNA. VestnikKazNU.Biological and Medical Series. 2014;1:191-194. (in Russian)Yu Ch.-Y., Cho Y., Sharma O., Kanehara K. What\u2019s unique? The unfoldedprotein response in plants. J. Exp. Botany. 2021:erab513.DOI 10.1093/jxb/erab513."}
+{"text": "Gordonia phage Nebulosus was isolated from soil on Gordonia terrae and is a siphovirus. The genome is 52,175 bp in length, has 62% GC content, and encodes 96 protein-coding genes. Nebulosus encodes a partitioning system, ParABS, which is likely involved in lysogeny maintenance.The temperate  Gordonia terrae 3,612 on PYCa agar plates, and incubated at 30\u00b0C for 2 days. Plaques were purified after seven rounds of plaque purification using standard methods  (A DNA phenol-chloroform extraction method was performed on a high-titer lysate . DNA wasGTGGTTA) . The genGTGGTTA) , 7.http://cobamide2.bio.pitt.edu) and PECAAN (https://blog.kbrinsgd.org/) (http://phages.wustl.edu/starterator) (\u2013The Nebulosus genome was auto-annotated using GLIMMER v3.02 and GeneMark v2.5 within DNA Master v.5.23.6 (gd.org/) , 12. Traterator) . Putativterator) \u201315. Threerator) \u2013, 17. A merator) \u2013 (7). Neberator) \u2013.Streptomyces kanamyceticus kanamycin biosynthetic gene cluster  downstream from the DNA polymerase I (gp46) with strong HHpred hits to a domain of unknown function DUF6197) found in a 7 found i"}
+{"text": "In the published article, there was an error in the legend for in vivo. Scale bar, 500\u03bcm.\u201d\u201cHucMSC-Exosomes accelerate wound healing. (A) Schematic overview of the study design. (B) Microscope image of huc-MSCs. The scale bar is 500 \u00b5m. (C) Flow cytometry analysis of HLA, CD29, CD34, CD45, CD73, CD90 and CD105 expression on the huc-MSC surface. (D) Transmission electron microscopy image of hucMSC-Exosomes. Scale bar, 100 nm. (E) Density plot of hucMSC-Exosomes. (F) Western blot of hucMSC-Exosome markers (G) Representative images of wound closure process of hucMSC-Exosomes treatment and PBS control. The scale bar is 4\u00a0mm. (H) Wound closure rate of hucMSC-Exosomes treatment and PBS control group. Two-tailed unpaired t-test (n=6). Error bar: mean standard deviation. ns, not significant, *p < 0.05, ***p < 0.001, ****p < 0.0001. (I) H & E and Masson staining of wounds seven days after injury. Scale bar = 1mm. (J) Uptake of hucMSC-Exosome by skin wound The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way. The original article has been updated."}
+{"text": "Opportunities for PrEP continue to be missed in key populations. We established a PrEP navigation program (\u201cSNAPS\u201d) with the goals to (1) increase PrEP uptake among groups disproportionately impacted by HIV and (2) preserve and improve PrEP adherence in an NYC safety-net hospital setting.SNAPS consisted of 5 components: (1) Surveillance of clinical sites where STI testing is high but PrEP use is rare e.g., ED and Women\u2019s Health Clinic (WHC), (2) Navigation for PrEP-eligible individuals (3) Accelerated follow-up with PrEP experts, (4) Point-of-care counseling and lab testing, and (5) Seamless longitudinal care. SNAPS launched 6/2019 with 2 full-time navigators. One year pre- vs post-SNAPS implementation we compared the sociodemographic profiles of PrEP initiators, their site of enrollment, and their medication possession ratios (MPRs), a proxy for PrEP adherence. Those on PrEP were a mixture of urgent and continuity care adults initiating PrEP at a safety-net hospital system. Bivariable analyses, employing Chi-square and t-test statistics, were conducted to compare sociodemographic profiles and MPR pre- vs post-SNAPS.We analyzed data on 274 (n=147 pre-SNAPS and n=127 post-SNAPS) individuals on PrEP. Compared to the pre-SNAPS period, post-SNAPS individuals were more likely to be cisgender women (33.9% vs 13.6%), Black or Latinx (84.3% vs 49.6%), uninsured (35.4% vs 29.3%), and Spanish speaking (58.3% vs 17.0%) (Table 1). Post-SNAPS individuals were more likely to be started on PrEP from the ED (51.2% vs 0%) or WHC clinic (10.2% vs 0%). Mean MPR for post-SNAPS was significantly lower than pre-SNAPS, 0.64 vs 0.89 (p < 0.001). Among post-SNAPS individuals, MSM were more likely to have higher MPRs compared to MSW and WSM individuals (Table 2). Furthermore, no difference in MPR was noted by race, preferred language, or insurance type.SNAPS successfully identified and linked key populations historically missed for PrEP opportunities. Efforts to improve SNAPS should target medication adherence.All Authors: No reported disclosures"}
+{"text": "Aeromonas caviae is an increasingly recognized etiological agent of acute gastroenteritis. Here, we report five draft genomes of A. caviae isolated from suspected cholera cases during the 2022\u20132023 cholera outbreak in Malawi. Aeromonas species cause a range of clinical infections, including gastroenteritis, bacteremia, septicemia, peritonitis, pneumonia, and wound infections  were identified as A. caviae by sequencing.The eviously . Genomiceviously . Of 68 iAeromonas species. The non-human reads were assembled using SPAdes v3.14.0 (A. caviae were identified by scanning the assemblies against PubMLST typing schemes (https://github.com/tseemann/mlst). The reads of A. caviae isolates from Malawi and those obtained from GenBank were mapped to the A. caviae reference genome strain 8LM (GenBank accession: CP024198) (https://github.com/tseemann/snippy) to generate pseudo-whole-genome alignments. Single nucleotide polymorphisms (SNPs) were identified in the alignment using snp-sites v2.5.1 Aeromonass v2.5.1 . Coding s v2.5.1 . Genome"}
+{"text": "Background: Breast cancer (BRCA) represents the most frequent diagnosed malignancy in women worldwide. Despite treatment advances, BRCAs eventually develop resistance to targeted therapies, resulting in poor prognosis. The identification of new biomarkers, like immune-related long non-coding RNAs (lncRNAs), could contribute to the clinical management of BRCA patients. In this report, we evaluated the LINC00426 expression in PAM50 BRCA subtypes from two clinical independent cohorts (BRCA-TCGA and GEO-GSE96058 datasets).Methods and results: Using Cox regression models and Kaplan-Meier survival analyses, we identified that LINC00426 expression was a consistent overall survival (OS) predictor in luminal B (LB) BRCA patients. Subsequently, differential gene expression and gene set enrichment analyses identified that LINC00426 expression was associated with different immune-related and cancer-related pathways and processes in LB BRCA. Additionally, the LINC00426 expression was correlated with the infiltration level of diverse immune cell populations, alongside immune checkpoint and cytolytic activity-related gene expression.Conclusion: This evidence suggests that LINC00426 is a potential biomarker of immune phenotype and an OS predictor in PAM50 LB BRCA. In 2020, breast cancer (BRCA) was the most frequent diagnosed malignancy and the leading cause of cancer-related death in women worldwide . BRCA isLncRNAs are a class of non-protein-coding transcripts greater than 200 nucleotides in length. Within a cell, lncRNAs are key players in a wide range of biological functions like regulation of gene expression, chromatin modification, genomic imprinting, transcriptional and translational processing . PreviouLINC00426 is a human lncRNA gene which contains 38,105 bases in length and is in the 13q12.3 region of the DNA antisense strand  of public databases. Using Cox regression models and Kaplan-Meier survival analyses, we found that LINC00426 expression is associated with OS in LB BRCA patients from both cohorts. Differential gene expression (DGE) and gene set enrichment analyses (GSEA) revealed that LINC00426 is associated with different immune-related and cancer-related pathways and processes in LB BRCA. Additionally, the LINC00426 expression correlates with immune-cell infiltration, expression of immune checkpoint genes (ICG) and cytolytic activity-related genes (CARG). These data suggest that LINC00426 is a potential biomarker of immune phenotype and an OS predictor in PAM50 LB BRCA.https://www.cbioportal.org/) and GDC Data Portal . Female patients who received neoadjuvant treatment and/or were lacking OS data were excluded, leaving a total of 927 patients analyzed in this study . The raw expression data were normalized to transcripts per million (TPM) and log2(TPM+1). A validation cohort (GSE96058) of 3,052 patients  was obtained from GEO database (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE96058). Again, female patients were lacking OS data and/or samples with label \u201crepl\u201d were excluded. For both datasets, patients were stratified in groups of low and high expression of LINC00426 by PAM50 BRCA subtypes, based on the lower (25%) and upper quartile (75%), respectively.Clinical information and raw RNA-seq expression data from BRCA patients in different PAM50 subtypes were obtained from TCGA database through cBioPortal  and survminer (version 0.4.9). The absolute number of patients at risk by time (in months) was determined through the survfit command and n.risk option from survival (version 3.4.0). Univariate analyses were performed through Cox proportional hazards regression models to identify clinicopathological variables associated with OS of patients stratified by PAM50 BRCA subtypes. Multivariate analyses were performed using Cox proportional hazards regression models and OS predictor variables, statistically significant, obtained via univariate analyses. Hazard ratios (HR) and 95% confidence intervals were obtained for each clinicopathological variable. These analyses were performed via survival (version 3.4.0) and survminer (version 0.4.9). p values < 0.05 were considered statistically significant.Considering the OS data (in months), Kaplan-Meier survival analyses were performed through the log-rank test in patients stratified by PAM50 BRCA subtypes, according to the low and high expression of LINC00426. These analyses were performed using the R packages DESeq2 (version 1.38.1) (2FoldChange (LFC) > 1.5 and <\u22121.5 with adjusted p values < 0.05. Volcano plots were generated using the R package EnhancedVolcano (version 1.16.0). The list of differentially expressed genes was used to perform Gene Ontology (GO) over-representation analyses for biological processes and molecular functions, using the R package clusterProfiler (version 4.6.0)  . Raw coun 4.6.0) . Kyoto EGSEA (version 4.1.0) with default parameters and 1,000 permutations. Gene sets with nominal p values < 0.05 and FDR <25% were considered statistically significant, according to the GSEA User Guide instructions for sample size and phenotype permutations (https://www.gsea-msigdb.org/gsea/doc/GSEAUserGuideFrame.html?Interpreting_GSEA).Using the normalized RNA-seq expression data from groups of patients with low and high expression of LINC00426, a GSEA  was perfhttps://cibersortx.stanford.edu/). This analysis was performed using the normalized gene expression data with 1,000 permutations. p values < 0.05 were considered statistically significant.Using CIBERSORTx, an analytical software based on transcriptome deconvolution method to infer the cell-type-specific gene expression and cell type abundance from RNA-seq data, the relative abundance of 22 tumor-infiltrating immune cell populations was determined: naive B cells, memory B cells, plasma cells, CD8 T cells, naive CD4 T cells, memory CD4 T cells (resting), memory CD4 T cells (activated), T follicular helper cells, regulatory T cells, gamma-delta T cells, NK cells (resting), NK cells (activated), monocytes, M0 macrophages, M1 macrophages, M2 macrophages, dendritic cells (resting), dendritic cells (activated), mast cells (resting), mast cells (activated), eosinophils and neutrophils    . Similarnd PRF1) .http://gepia2.cancer-pku.cn/#general)  . The OS GraphPad Prism (version 8.3.0) and R package ggpubr (version 0.5.0). The non-parametric Kruskal-Wallis test was performed to identify differences in the LINC00426 expression between PAM50 BRCA subtypes. The non-parametric Mann-Whitney U test was performed to determine differences in the infiltration level of immune cell populations, ICG and CARG expression between groups of BRCA patients with low and high expression of LINC00426. Spearman correlation analyses were performed to determine statistical relationships between the LINC00426 expression and the infiltration level of immune cell populations, ICG and CARG expression . p values < 0.05 were considered statistically significant.Statistical analyses were performed through n = 927) found high expression of LINC00426 in non-luminal compared to luminal BRCA subtypes. Significant differences were identified between LA and BL (p = 0.0063), LB and HER2-enriched (p = 0.0089), LB and BL (p = 0.0004) subtypes. No significant difference was observed in the LINC00426 expression between LA and HER2-enriched subtypes (p = 0.0878) (p = 0.28), HER2-enriched (p = 0.21) and BL BRCA patients (p = 0.12) (p = 0.01) (p = 0.024), with no significance observed in LA (p = 0.288), HER2-enriched (p = 0.246) and BL (p = 0.133) subtypes  . The cli = 0.12) . However = 0.01) , respectsubtypes . The OS subtypes . These rsubtypes .CLEC6A, IFNG, PLA2G2D, DCD and GNAT3) and 224 genes were upregulated     . Subsequd NELL1) , alongsi < 0.05) . The KEG < 0.05) .p-value < 0.05 and FDR <25%) with eight immune-related gene sets  and seven cancer-related gene sets    . In contDR <25%) . Altogetp = 0.0242), plasma cells (p = 0.0324), CD8 T cells (p < 0.0001), memory CD4 T cells (resting) (p = 0.0220), memory CD4 T cells (activated) (p < 0.0001), gamma-delta T cells (p < 0.0001), M1 macrophages (p < 0.0001) and increased infiltration of memory B cells (p < 0.0001), NK cells (resting) (p < 0.0001), M0 macrophages (p < 0.0001), M2 macrophages (p < 0.0001), mast cells (resting) (p = 0.0018), mast cells (activated) (p = 0.0005) and eosinophils (p < 0.0001). These immune cell infiltration patterns are reverted in patients with high expression of LINC00426 (p > 0.05)  . These rPDCD1, PDCD1LG2, CD274, CTLA4, LAG3, TIGIT and IDO1) and CARGs  expression in the PAM50 LB subtype from the BRCA-TCGA cohort. Spearman correlation analyses indicated that LINC00426 expression, positively and significatively, correlates with PDCD1 (R = 0.797), PDCD1LG2 (R = 0.736), CD274 (R = 0.618), CTLA4 (R = 0.821), LAG3 (R = 0.639), TIGIT (R = 0.901), IDO1 (R = 0.738), GZMA (R = 0.878), GZMB (R = 0.749), PRF1 (R = 0.830), ICG (R = 0.843) and CARG signatures (R = 0.845) (p < 0.001) (p < 0.0001) (p < 0.001)  . Concord 0.0001) . Further< 0.001) . These rn = 3,052). The clinicopathological characteristics of PAM50 BRCA patients from this cohort are described in p < 0.0001), LA and BL (p < 0.0001), LB and HER2-enriched (p < 0.0001), and LB and BL BRCA subtypes (p < 0.0001) (p = 0.98) (p = 0.042) (p = 0.005) and BL (p = 0.005) BRCA patients in the GEO-GSE96058 cohort  . Kaplan- = 0.98) , while t= 0.042) . In cont8 cohort . These f8 cohort . Multiva8 cohort . In addi8 cohort .p = 0.0003), plasma cells (p = 0.0225), CD8 T cells (p < 0.0001), memory CD4 T cells (resting) (p < 0.0001), memory CD4 T cells (activated) (p < 0.0001), gamma-delta T cells (p = 0.0004), M1 macrophages (p < 0.0001) and increased infiltration levels of memory B cells (p = 0.0049), NK cells (resting) (p < 0.0001), M0 macrophages (p = 0.0059), M2 macrophages (p < 0.0001), mast cells (resting) (p < 0.0001), mast cells (activated) (p < 0.0001) and eosinophils (p < 0.0001) (p = 0.0020), regulatory T cells (p < 0.0001), NK cells (activated) (p < 0.0001) and increased infiltration of naive CD4 T cells (p = 0.0137), dendritic cells (activated) (p = 0.0038) and neutrophils (p < 0.0001) (p > 0.05)  . In addi 0.0001) , when co 0.0001) . The inf > 0.05) .PDCD1 (R = 0.763), PDCD1LG2 (R = 0.687), CD274 (R = 0.648), CTLA4 (R = 0.770), LAG3 (R = 0.664), TIGIT (R = 0.807), IDO1 (R = 0.737), GZMA (R = 0.783), GZMB (R = 0.705), PRF1 (R = 0.763), ICG (R = 0.792) and CARG signatures (R = 0.773) (p < 0.001) (p < 0.0001) (p < 0.001) (We corroborated that LINC00426 expression positively correlates with the expression of < 0.001) . The exp 0.0001) . Also, t< 0.001) . These rThe expression of lncRNAs vary between different cancer types and can promote or antagonize tumor progression ; therefoIn this study, we found that LINC00426 expression is a consistent OS predictor in PAM50 LB BRCA in the BRCA-TCGA and GEO-GSE96058 cohorts, in contrast to other subtypes. Particularly, the low and high expression of LINC00426 was associated with reduced and increased OS in LB BRCA patients, respectively. Interestingly, a previous study showed a similar prognostic behavior for LINC00426 in LUAD and NSCLC . In contPrevious reports demonstrated that LINC00426 promotes LUAD progression and doxorubicin resistance in OSA, suggesting a potential oncogenic role of LINC00426 in these cancers . ConversSeveral studies demonstrated that tumor-intrinsic factors, like dysregulations on diverse oncogenic pathways, modulate the host\u2019s anti-tumor immune response depending on the cancer type and cellular context . We idenCCL2 through epigenetic pathways and hnRNPL binding to the promotor region of CCL2, which results in the recruitment of tumor-associated macrophages to the TIME of bladder cancer, promoting lymphatic metastasis via VEGF-C excretion . In addition, previous studies showed relationships between lncRNAs, including FENDRR and BCAR4, with the expression of cytokines and chemokines in cancer, which could modulate the infiltration of immune cells to the TIME . Chen etxcretion . FurtherGZMA and PFR1) are important to determine the functional status of local anti-tumor immune response  and CARGs . We suggest that LINC00426 could be involved, directly or indirectly, in the regulation of ICGs and CARGs expression in PAM50 LB BRCA. Studies demonstrated that the lncRNAs XIST, TSIX and MALAT1 regulate the PD-L1 expression in BRCA through ceRNA networks  and cytolytic activity markers (i.e., response . Reportsresponse  with Xiain ccRCC . We idennetworks . AdditioAlthough the importance of the immune response was reported in LB BRCA , there aThe main limitations of our study are related to its retrospective nature and bioinformatics approach based on transcriptomic data, limiting the mechanistic conclusions of LINC00426. Validation of these findings is needed through methodologies like multiplex immunofluorescence or flow cytometry. Future studies focused on LINC00426 in PAM50 LB BRCA are needed that include experimental approaches to gain a wide understanding about the exact functional role of LINC00426. Despite these limitations, we conclude that LINC00426 is a potential biomarker of cancer immune phenotype whose expression has a consistent and an OS prognostic value in PAM50 LB BRCA patients in two independent cohorts, which suggest a potential use for immunotherapies selection in patients, but further analyses are mandatory to confirm this hypothesis."}
+{"text": "Prostate Cancer and Prostatic Diseases 10.1038/s41391-022-00555-0, published online 03 June 2022.Correction to: In Table The original article has been corrected.Page 4: Last sentence before the section of Survival analysis.p < 0.0001; Table In patients with pre-existing CVD and receiving ADT for \u22656 months, a 70% lower risk of composite CV events was determined in GnRH antagonisttreated patients than GnRHa-treated patients (aHR 0.30; 95% CI, 0.16\u20130.54;"}
+{"text": "The correct name is: Yuelong Yan. The correct citation is: Yan Y, Zhang C, Hao J, Wang X-L, Ming J, Mi L, et al. (2019) DPPA2/4 and SUMO E3 ligase PIAS4 opposingly regulate zygotic transcriptional program. PLoS Biol 17(6): e3000324."}
+{"text": "Anaerostipes hadrus strains BA1 and GIF7 were isolated from a healthy man. The complete genomes\u2019 sizes are 2,946,270 bp (BA1) and 2,907,308 bp (GIF7), with high average nucleotide identity (ANIb = 100%) and alignments \u226596.86% between strains. Conversely, both strains share 97.47% (ANIb) identity and \u226477.36% alignments to A. hadrus ATCC 29173T. Anaerostipes of Bacillota phylum and Lachnospiraceae family are known most prominently for producing butyrate  was first isolated in 1976  agar (Oxoid) were sub-cultured in BHI broth (4 mL) for DNA extraction. Cells were lysed using MP Biomedicals bead-beating tubes on a Fast-Prep 24/5G at 6.0 m/sec for 40 s followed by DNA purification using a Promega Maxwell-16-FFS kit without modifications to the manufacturer\u2019s protocol. Paired-end 150-bp reads were sequenced on an Illumina NovaSeq without modifications to TruSeq Nano DNA library prep kit. Default software parameters were applied unless stated otherwise. Trimmomatic v.0.39 was used to trim and remove adapters from Illumina reads . Bo. BoA. haentities . A high entities ."}
+{"text": "J Clin Endo Metab Case Reports. 2023; 1(1): 10.1210/jcemcr/luac017), a production error occurred in the PDF version of Figure 1 regarding patient information.In the above-named article by Raven LM, Greenfield JR, and Muir CA (The article has been corrected online.The publisher regrets the error.10.1210/jcemcr/luac017doi:"}
+{"text": "Applying data-reduction techniques to extract meaningful information from electronic performance and tracking systems (EPTS) has become a hot topic in football training load (TL) monitoring. The aim of this study was to reduce the dimensionality of the internal and external load measures, by a principal component approach, to describe and explain the resultant equations for TL monitoring during a standard in-season microcycle in sub-elite youth football. Additionally, it is intended to identify the most representative measure for each principal component. A principal component analysis (PCA) was conducted with a Monte Carlo parallel analysis and VariMax rotation to extract baseline characteristics, external TL, heart rate (HR)-based measures and perceived exertion. Training data were collected from sixty sub-elite young football players during a 6-week training period using 18 Hz global positioning system (GPS) with inertial sensors, 1 Hz short-range telemetry system, total quality recovery (TQR) and rating of perceived exertion (RPE). Five principal components accounted for 68.7% of the total variance explained in the training data. Resultant equations from PCA was subdivided into: (1) explosiveness, accelerations and impacts (27.4%); (2) high-speed running (16.2%); (3) HR-based measures (10.0%); (4) baseline characteristics (8.3%); and (5) average running velocity (6.7%). Considering the highest factor in each principal component, decelerations (PCA 1), sprint distance (PCA 2), average HR (PCA 3), chronological age (PCA 4) and maximal speed (PCA 5) are the conditional dimension to be considered in TL monitoring during a standard microcycle in sub-elite youth football players. Current research provides the first composite equations to extract the most representative components during a standard in-season microcycle in sub-elite youth football players. Futures research should expand the resultant equations within training days, by considering other well-being measures, technical-tactical skills and match-related contextual factors. This isimpacts) . Otherwiimpacts) . The resimpacts) . Combiniimpacts) .Additionally, the emergent tracking tools appears to have created confusion in dose-response considerations given the data analysis requirement to extract relevant information from large amounts of data . This kiPrincipal component analysis (PCA) is one of the most used data-reduction techniques to extract redundant information from TL data in football . Using an = 20), U17 (n = 20) and U19 (n = 20) sub-elite youth football academy  15  was assured to perform PCA analysis   within a little pocket on the upper back between both scapulae of a custom-made vest (The training data eligibility considered the following inclusion criteria: (a) youth football players aged between 13 and 20 years old (and U19) ; (b) youand U19) ; (c) traand U19) ; (d) traand U19) , 2022a. and U19) . For ETLade vest . All metade vest .n = 41), MD-2 (n = 38), and MD-1 (n = 44)  (Using a \u201cmatch day minus format\u201d (MD), the weekly microcycle included the training sessions MD-3 (Tuesday), MD-2 (Wednesday), and MD-1 (Friday). The number of observation for each training day was: MD-3 ((n = 44) , 2021b. (n = 44) . All tra 52\u201366%) .i.e., MD-3, MD-2 and MD-1)  was categorized in accordance with nd MD-1) . Small, \u00ae, Northern Ireland) (\u00ae devices was good (bias 5%) (\u22121)), maximal running speed (MRS (m\u00b7s\u22121)), relative high-speed running (rHSR (m): 19.8\u201325.1 km\u00b7h\u22121) distance (m), high metabolic load distance (HMLD (m) > 25.5 W\u00b7kg\u22121), number sprints (n) and average sprint distance (SPR (m) (>25.1 km\u00b7h\u22121)) (m), dynamic stress load (DSL (a.u.)), number of ACC (>3 m\u00b7s\u22122) and number of decelerations (DEC < 3 m\u00b7s\u22122)  coupled with accelerometer (100 Hz), magnetometer (10 Hz) and gyroscope (100 Hz) (STATSports ApexIreland) . With a Ireland) . The accbias 5%) . The ETL3 m\u00b7s\u22122) , 2022a (3 m\u00b7s\u22122) .\u00ae TM HR band  was used to capture HR-based measurements utilizing a 1 Hz short-range telemetry system (max), average heart rate (HRmean), percentage of HRmax (%HRmax) and individual players\u2019 training impulse (TRIMP) were monitored   , 2022a  .\u00ae  with minimum clothing. Height (cm) was collected using an electronic stadiometer . Players\u2019 height (m), weight (kg) and sitting height (cm) were recorded by the average of three measurements to the nearest 0.1 using international units (IU). Body mass index (BMI) was calculated by dividing weight by the square of height (kg/m2). BMI\u2019s cut-offs used were: underweight < 18.5 kg/m2, normal 18.50\u201324.99 kg/m2, overweight \u2265 25 kg/m2, obese \u2265 30 kg/m2 (n = 52), mid-PHV (n = 65) and post-PHV (n = 207).Players\u2019 individual characteristics were collected by height (m), weight (kg), chronological age (years), sitting height (cm) and experience level (years). Anthropometric measures were measured using standard the International Society for the Advancement of Kinanthropometry (ISAK) guidelines . Body ma30 kg/m2 . Relativ30 kg/m2 . Based o30 kg/m2 . Sub-elii.e., anthropometric and maturational status) and the ITL measures  of the first principal component. The loadings must have a sum of squares of exactly one. This is due to the possibility of a considerable variation when loadings are of a great magnitude.n), along which data varies the most  have mean values equal to zero and standard deviations equal to one  and its weighted load vector (d vector .r < 0.4 (p < 0.05. Data are presented as the mean \u00b1 SD using JASP software (jasp-stats.org).A data reduction technique was conducted using a principal component analysis (PCA) with 95% confidence intervals (95% CI) . Monte Cr < 0.4 . Weightir < 0.4 . Kolmogovia HRmax, AvHR, %HR and TRIMP. The fourth PCA explained 8.3% of the baseline outset . The fifth PCA explained 6.7% of the accumulated TL . Constantly, PHV, relative age, experience level and BMI were excluded from the PCA (r < 0.4).i.e., PHV, relative age, experience level and BMI). Also, KMO\u2019s criteria reported a sampling adequacy of sampled data, reporting a considerable proportion of the variance as result of the underlying factors (KMO = 0.73). Furthermore, significant Barlett Sphericity test was significant (p < 0.001).The weightings (eigenvectors) of the PCA analysis are represented by a path graph in The resultant equations from extracted principal component are presented in The aim of this study was to reduce the dimensionality of the internal and external load measures, by a PCA approach, in order to describe and explain the resultant equations for TL monitoring during a standard microcycle in a sub-elite youth football players. Additionally, it is intended to identify the most representative measure for each principal component. After data reduction, five principal components were extracted from TL dataset explaining 68.7% of the total variance. The TL measures with the highest weight in each PCA were DEC, SPR distance, average HR, chronological age and MRS.i.e., HRmax, AvHR, %HRmax and TRIMP), confirming the correlation between HR-based measures and ETL outcomes  explosiveness, ACC and impacts (27.4%); (2) HSR (16.2%); (3) heart bate-based measures (10.0%); (4) baseline characteristics (8.3%); (5) average running velocity (6.7%). Considering the highest representative factor in each principal component, the variables considered were DEC (PCA 1), SPR distance (PCA 2), average HR (PCA 3), chronological age (PCA 4) and MRS (PCA 5). In football, outcomes . The fououtcomes . In lineoutcomes . Also, toutcomes . Indeed,outcomes . Effectioutcomes , 2018. Loutcomes , 2022c. outcomes , 2020.i.e., eigenvalues between 1.0% and 68.0%)  . Albeit,d 68.0%) .Current applied PCA determine the resultant equations from individual-based principal components, expressing by major component weightings . Indeed,i.e., small-sided and conditioned games), training day , age group  and maturational bands   . Additioost-PHV) . Also, tost-PHV) . Howeverost-PHV) . Furtherost-PHV) . In geneost-PHV) . Furtherost-PHV) . Finallyost-PHV) , 2021b. ost-PHV) .Current resultant composite equations can be applied to relative contribution of the ITL and ETL measures for monitoring and management load in sub-elite youth football.Data reduction techniques decrease the redundant information and dimensionality of the training data, expressing in the following principal components: explosiveness and impacts, high-speed running, heart bate-based measures, baseline characteristics and average running velocity.Considering the highest factor in each principal component, DEC (PCA 1), sprint distance (PCA 2), average HR (PCA 3), chronological age (PCA 4) and maximal speed (PCA 5) are the conditional dimension to be considered in TL monitoring during a standard microcycle in sub-elite youth football players.Maturational status should be carefully considered in the TL monitoring together with relative age effect, chronological and baseline characteristics.Self-perception and practice experience may affect the variance explained by perceived exertion and pacing behavior.Training intensity and volume can be more accurately measured by current resultant composite equations and/or most representative factor for a standard microcycle in sub-elite youth football players.Futures research should expand the resultant equations for TL monitoring in sub-elite youth football with well-being, technical-tactical and match-related contextual factors.Using a PCA approach, five principal components could be applied to extract to describe and explain resultant equations for TL monitoring during an in-season standard microcycle in sub-elite youth football. Current research provides the first composite equations to extract the TL in this specific population expressed as explosiveness and impacts, high-speed running, HR-based measures, baseline characteristics and average running velocity. Considering the highest factor in each principal component, DEC (PCA 1), SPR distance (PCA 2), average HR (PCA 3), chronological age (PCA 4) and maximal SPR (PCA 5) are the conditional dimension to be considered in TL monitoring during a standard microcycle in sub-elite youth football players.Future research should expand the resultant equations within the microcycle, by considering other well-being measures, technical-tactical factors and match-related contextual factors.10.7717/peerj.15806/supp-1Supplemental Information 1The individual data of the variables selected for this study.Click here for additional data file."}
+{"text": "Arthrobacter globiformis B-2979 culture. MrAaronian has > 99% nucleotide identity with cluster AW arthrobacteriophages Michelle, Stayer, Sloopyjoe, and StarLord.Arthrobacteriophage MrAaronian contains a 54,509 bp DNA genome with 87 predicted protein-coding genes. MrAaronian has siphovirus morphology and was collected from a flowerbed soil sample in Poughkeepsie, NY, and isolated on an  Arthrobacter group of bacteria is unusual since they appear as Gram-negative rods in young cultures and as Gram-positive cocci in older cultures  coordinates: 41.72201 N, 73.930158 W]. The sample was washed with peptone-yeast calcium (PYCa) liquid media, and the supernatant was passed through a 0.22 \u00b5m syringe filter. The filtrate was incubated with host biformis . High-tiry Guide .Bacteriophage genomic DNA was extracted using a Promega Wizard DNA Clean-Up System , and sequencing libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit . Libraries were sequenced by Illumina MiSeq at the Pittsburgh Bacteriophage Institute to generate 607,351 total single-end reads of 150-base read length. A single bacteriophage contig with 6,670\u00d7 coverage was assembled using Newbler v2.9 and checked for completeness and genomic termini using Consed v29 (https://discover.kbrinsgd.org), and DNA Master v5.23.6 (http://phagesdb.org/DNAMaster/). Consistent with other cluster AW bacteriophages, no tRNA genes were identified using Aragorn v1.1 (12) and tRNAscan-SE v2.0 (BLASTn  query of-SE v2.0 . TmHmm a-SE v2.0  assessed-SE v2.0  and HHPr-SE v2.0 , which i-SE v2.0  showing -SE v2.0 . All too"}
+{"text": "Nacubactam is a new non-\u03b2-lactam diazabicyclooctane \u03b2-lactamase inhibitor that inhibits penicillin binding protein 2 (PBP2) of Enterobacterales and acts synergistically as a \u03b2-lactam enhancer when combined with \u03b2-lactams. Nacubactam is under development for the treatment of serious Gram-negative infections. We evaluated the in vitro activity of cefepime-nacubactam against carbapenem-resistant Enterobacterales (CRE) clinical strains isolated at six medical facilities in Japan.Activity of cefepime-nacubactam (1:1 ratio) was tested against 376 CRE (MIC of meropenem or imipenem: \u22652 mg/L). The MICs were evaluated by the broth microdilution method according to the CLSI using frozen-panels  containing various antimicrobial agents. The percentage susceptible (%S) for cefepime-nacubactam was calculated using the CLSI susceptible breakpoint of cefepime.Escherichia coli, 27; Klebsiella spp., 89; Enterobacter spp.; 56; others, 12), the MIC50/MIC90 and %S were 8/32 mg/L and 4.9%S for meropenem, 4/32 mg/L and 29.3%S for imipenem, 32/ >128 mg/L and 31.1%S as 8 mg/L for cefepime and 2/4 mg/L and 95.7%S for cefepime-nacubactam, respectively. Against non-CPE , the MIC50/MIC90 and %S were 0.5/8 mg/L and 53.6%S for meropenem, 2/8 mg/L and 12.0%S for imipenem, 2/ >128 mg/L and 66.7%S as 8 mg/L for cefepime and 0.25/4 mg/L and 99.0%S for cefepime-nacubactam, respectively. The MIC50/MIC90 and %S of cefepime-nacubactam was 2/4 mg/L and 94.9%S against 156 MBL-producing CRE, and 0.5/2 mg/L and 100%S against non-MBL-producing CRE including 14 KPC and 8 OXA-producing strains and 6 strains producing other carbapenemases. Nacubactam alone showed limited activity against CPE  and non-CPE .The collection included 184 carbapenemase-producing Enterobacterales (CPE) strains and 192 carbapenemase-non-producing Enterobacterales (non-CPE) strains. Against CPE (In vitro activity of cefepime-nacubactam against CRE isolated in JapanCefepime-nacubactam demonstrated potent activity against both CPE and non-CPE/CRE clinical strains. This is the first study to report the activity of cefepime-nacubactam against carbapenemase-non-producing CRE, which accounts for 80% of CRE in Japan.Katsunori Yanagihara, MD, PhD, FUJIFILM Toyama Chemical Co., Ltd.: Commissioned research|KYORIN Pharmaceutical Co.,Ltd.: Commissioned research Hiroshige Mikamo, M.D, Ph.D, Asahi Kasei Pharma Corporation: Grant/Research Support|Merck Sharp & Dohme: Honoraria|Pfizer Inc.: Grant/Research Support|Pfizer R&D Japan: Honoraria|Sumitomo Pharma Co., Ltd.: Grant/Research Support|Sumitomo Pharma Co., Ltd.: Honoraria Yohei Doi, MD, PhD, bioMerieux: Advisor/Consultant|FujiFilm: Advisor/Consultant|Gilead: Advisor/Consultant|Gilead: Honoraria|GSK: Advisor/Consultant|Meiji Seika Pharma: Advisor/Consultant|Moderna: Advisor/Consultant|Moderna: Honoraria|MSD: Advisor/Consultant|MSD: Honoraria|Shionogi: Advisor/Consultant|Shionogi: Grant/Research Support|Shionogi: Honoraria"}
+{"text": "T and NL03-T5-1, were isolated from a soil sample collected from the Nanling National Forests, Guangdong Province, PR China. The two strains were Gram-stain-positive, aerobic, rod-shaped and had lophotrichous flagellation. Strain NL03-T5T could secrete extracellular mucus whereas NL03-T5-1 could not. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the two strains belong to the genus Cohnella, were most closely related to Cohnella lupini LMG 27416T (95.9% and 96.1% similarities), and both showed 94.0% similarity with Cohnella arctica NRRL B-59459T, respectively. The two strains showed 99.8% 16S rRNA gene sequence similarity between them. The draft genome size of strain NL03-T5T was 7.44 Mbp with a DNA G+C content of 49.2 mol%. The average nucleotide identities (ANI) and the digital DNA\u2013DNA hybridization (dDDH) values between NL03-T5T and NL03-T5-1 were 99.98% and 100%, indicating the two strains were of the same species. Additionally, the ANI and dDDH values between NL03-T5T and C. lupini LMG 27416T were 76.1% and 20.4%, respectively. The major cellular fatty acids of strain NL03-T5T included anteiso-C15:0 and iso-C16:0. The major polar lipids and predominant respiratory quinone were diphosphatidylglycerol (DPG) and menaquinone-7 (MK-7). Based on phylogenetic analysis, phenotypic and chemotaxonomic characterization, genomic DNA G+C content, and ANI and dDDH values, strains NL03-T5T and NL03-T5-1 represent novel species in the genus Cohnella, for which the name Cohnella silvisoli is proposed. The type strain is NL03-T5T (=GDMCC 1.2294T = JCM 34999T). Furthermore, comparative genomics revealed that the genus Cohnella had an open pan-genome. The pan-genome of 29 Cohnella strains contained 41,356 gene families, and the number of strain-specific genes ranged from 6 to 1649. The results may explain the good adaptability of the Cohnella strains to different habitats at the genetic level.Two strains, designated NL03-T5 Cohnella, which lies within the family Paenibacillaceae of the order Bacillales, was first proposed by K\u00e4mpfer et al. with the description of Cohnella thermotolerans as the type species , and catalase activity was determined by bubble production after mixing cells with 3% H2O2. Gliding motility was checked by observing the edges of colonies formed on 1:6-diluted R2A and using the hanging drop technique as described by Bernardet et al. (2002) [w/v), CM-cellulose , Tween 20  and 40  were examined as described by Son et al. [The morphological features of strains NL03-T5. (2002) . Hydrolyn et al. . Other pT and its related species were harvested from R2A agar after being incubated for 4 d at 30 \u2103. Fatty acid methyl esters were extracted using the Sherlock Microbial Identification System (MIDI) protocol version 6.1 and analyzed by gas chromatography  as previously described [For cellular fatty acids, polar lipids, and respiratory quinones analysis, strain NL03-T5escribed . The polescribed . Respiraescribed  and analT and NL03-T5-1 were sequenced on the Illumina HiSeq platform at Shanghai Majorbio Bio-Pharm Technology Co., Ltd. . Sequencing reads were assembled into contigs and scaffolds by applying SPAdes version 3.11.1 with default parameters [http://rast.nmpdr.org/, accessed on 1 August 2023) with default parameters. The genes encoding carbohydrate active enzymes (CAZymes) were identified using the dbCAN2 meta server  with HMMER annotation , and biosynthetic gene clusters (BGCs) were annotated using antiSMASH bacterial version 6.1.1  with default parameters, respectively. All 27 reference genomes of the genus Cohnella were downloaded from the NCBI database. A pan-genome analysis was performed using the bacterial pan-genome analyses tool (BPGA) pipeline [https://www.bic.ac.cn/ImageGP/, accessed on 1 August 2023). The genomic DNAs of strains NL03-T5rameters , and thepipeline . The dathttps://ggdc.dsmz.de/, accessed on 1 August 2023) [www.ezbiocloud.net/tools/ani, accessed on 1 August 2023) [Cohnella was reconstructed based on 92 up-to-date core genes using the software UBCG version 3.0 [Overall genome relatedness indices (OGRI) including the digital DNA\u2013DNA hybridization (dDDH) and average nucleotide identity (ANI) values were calculated using the Genome-to-Genome Distance Calculator (GGDC) (st 2023)  and Orthst 2023) , respectsion 3.0  with a mT and NL03-T5-1 showed 99.8% 16S rRNA gene sequence similarity. In the EzBiocloud and NCBI databases, the two strains were closely related to the species of the genus Cohnella and showed the highest similarities with C. lupini LMG 27416T (95.9% and 96.1%), and they both exhibited 95.3% and 94.0% similarities with C. abietis HS21T and C. arctica NRRL B-59459T, respectively. The 16S rRNA gene phylogenetic trees based on the ML, NJ, and ME methods  NaCl  NaCl . StrainsT and its reference species C. abietis HS21T, C. lupini LMG 27416T, and C. arctica NRRL B-59459T are given in T contained a large amount of anteiso-C15:0 (50.7%) and iso-C16:0 (21.2%), and small amounts of C16:0 (7.2%), iso-C15:0 (5.5%), anteiso-C17:0 (4.0%), iso-C14:0 (3.7%), iso-C14:0 (1.9%), anteiso-C13:0 (1.6%) and C14:0 (1.3%). The prominent fatty acids were similar to other species of the genus Cohnella, indicating that strain NL03-T5T is a member of the genus Cohnella. However, strain NL03-T5T contained a higher amount of iso-C16:0 than its closely related species C. abietis HS21T, and it contained a lower amount of C16:0 and a higher amount of iso-C15:0 and iso-C16:0 than C. lupini LMG 27416T and C. arctica NRRL B-59459T. Moreover, strain NL03-T5T did not contain alcohol-C16:1\u00a0\u03c97c (6.5%) or C16:1\u00a0\u03c911c (7.0%) whereas C. lupini LMG 27416T did. These significant differences in the fatty acids clearly distinguished strain NL03-T5T from other related species. The major polar lipids of strain NL03-T5T were DPG and PE, which were in common with its closely related species. Otherwise, four unidentified amino phospholipids (APLs) were also detected in strain NL03-T5T , and higher than C. abietis HS21T (44.8%) , amino acids and derivatives (17.3%), protein metabolism (9.3%) and cofactors, vitamins, prosthetic groups, and pigments (8.9%)  and carbohydrate-binding modules (CBMs) constituting the most abundant families  . The diss (8.9%) . In addi strains . The antfamilies . Additiomination , indicatCohnella strains comprised 41,356 gene families. The core genes were present in all 29 genomes, accessory genes were present in 2\u201328 genomes, and unique genes were present only in one genome. The numbers of core genes, accessory genes, and unique genes were 492 (1.2%), 19,166 (46.3%), and 21,698 (52.5%), respectively  in the power-law regression function. New gene distribution and gene family distribution of the 29 Cohnella strains are shown in Cohnella strains to different habitats through gene gains or losses during frequent evolutionary changes at the genetic level. The pan-genome of the 29 ectively a. The nuectively b, sugges genomes c. The cuT and NL03-T5-1, from a soil sample collected from the Nanling National Forests, PR China. The two strains showed 99.8% 16S rRNA gene sequence similarity. The ANI and dDDH values between them are 99.98% and 100%. These results indicate the two strains are of the same species. However, strain NL03-T5T could secrete extracellular mucus whereas NL03-T5-1 could not  [Cohnella strains contained 41,356 gene families, and the numbers of core genes, accessory genes, and unique genes were 492, 19,166, and 21,698, respectively (Cohnella strains might be associated with their colonized environments.The genus er (HGT) . The panectively a,c. TettCohnella silvisoli .w/v) NaCl. Oxidase and catalase are -negative. It can hydrolyze Tween 20, but not Tween 40, casein, or CM-cellulose. Additionally, it can hydrolyze gelatin or not. The major fatty acids (>10%) include anteiso-C15:0 (50.7%) and iso-C16:0 (21.2%). The predominant polar lipid is DPG. The main respiratory quinone is MK-7. The API ZYM test result was positive for alkaline phosphatase, esterase, lipase, leucine arylamidase, acid phosphatase, naphthol-AS-BI-phosphohydrolase, \u03b1-galactosidase, \u03b2-galactosidase, \u03b1-glucosidase, \u03b2-glucosidase, N-acetyl-\u03b2-glucosaminidase and \u03b1-mannosidase; but negative for valine arylamidase, cystine arylamidase, trypsin, \u03b1-chymotrypsin and \u03b1-fucosidase. The API 20NE test results were positive for gelatin hydrolysis, 4-nitrophenyl-\u03b2-d-galactopyranosidase and \u03b2-glucosidase or not; but negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase and urease. It can assimilate glucose, L-arabinose, mannose, mannitol, N-acetyl-D-glucosamine, maltose, gluconate, adipate, malic acid, citric acid, phenylacetic acid and capric acid or not.Cells are aerobic, Gram-stain-positive, lophotrichous flagellation, rod-shaped, 1.2\u20132.0 \u03bcm long and 0.4\u20130.6 \u03bcm in diameter after incubation on R2A agar for 4 d. Colonies on R2A agar are white-cream, circular, convex with extracellular secretions or not. Growth occurs at 15\u201337 \u2103 and at a pH range from 5.0 to 8.5 . Cells can tolerate 0.5% (T (=GDMCC 1.2294T = JCM 34999T), which was isolated from a soil sample collected from the Nanling National Forests, Guangdong Province, PR China. The genomic DNA G+C content of the type strain is 49.2 mol%.The type strain is NL03-T5"}
+{"text": "Correction: BMC Genomics 24, 212 (2023)https://doi.org/10.1186/s12864-023-09310-8Following publication of the original article , it was Additional file 1: Supplementary table 1. List of candidate genes for WES analysis. Supplementary table 2. Annotation of candidate variants identified in the 22 eoRCC patients.Additional file 2: Supplementary Table 8."}
+{"text": "TERT (rs2736100), CCDC26 (rs4295627), CDKN2A/B (rs4977756) and RTEL1 (rs6010620) gene polymorphisms as the risk factors specific to glioma. However, the data samples of previous genetic-epidemiological studies are modest to determine whether they have definite association with glioma.Previous genetic-epidemiological studies considered The study paid attention to systematically searching databases of PubMed, Embase, Web of Science (WoS), Scopus, Cochrane Library and Google Scholars. Meta-analysis under 5 genetic models, namely recessive model (RM), over-dominant model (O-DM), allele model (AM), co-dominant model (C-DM) and dominant model (DM) was conducted for generating odds ratios (ORs) and 95% confidence intervals (CIs). That was accompanied by subgroup analyses according to various racial groups. The software STATA 17.0 MP was implemented in the study.TERT gene rs2736100 polymorphism, CCDC26 gene rs4295627 polymorphism, CDKN2A/B gene rs4977756 polymorphism and RTEL1 gene rs6010620 polymorphisms increased the risk of glioma in Caucasians to different degrees. In Asian populations, the CCDC26 gene rs4295627 polymorphism and CDKN2A/B gene rs4977756 polymorphism did not exhibit a relevance to the risk of glioma. It is suggested to cautiously explain these results as the sample size is small.21 articles were collected. According to data analysis results, in four genetic models  TERT (rs2736100), CCDC26 (rs4295627), CDKN2A/B (rs4977756) and RTEL1 (rs6010620) genes in glioma might increase risk of glioma, but there are ethnic differences. Further studies evaluating these polymorphisms and glioma risk are warranted.The current meta-analysis suggested that the SNP of  Based oTERT), 8q24.21 , 9p21.3 , and 20q13.33  (TERT) is crucial for telomerase activity in preserving telomeres and cellular immortality (CCDC26) downregulates telomerase activity and increases apoptosis. Cyclin-dependent kinase inhibitor 2A/B (CDKN2A/B) can partially control the cell cycle. Regulator of telomere elongation helicase 1 (RTEL1), as a DNA helicase, can inhibit homologous recombination for directly maintaining the genomic stability  . Telomerortality . In conttability . These gTERT (RS2736100), CCDC26 (RS4295627), CDKN2A/B (RS497756), and RTEL1 (rs6010620).In this meta-analysis, we expanded sample size by enrolling up 21 articles and used five genetic models to examine the relationship between glioma and SNPs of 2The meta-analysis in the study was performed as per the Preferred Reporting Items for Systematic reviews and Meta-Analysis version.2.1Literature search was carried out comprehensively in eight databases, namely PubMed, Embase, WoS, Scopus, Cochrane Library, and Google Scholars. The MeSH term were as follows: \u201c OR (SNP) OR  OR (SNP) OR (SNP) OR (SNPs) OR (variation) OR (mutation)) AND ((Gliomas) OR (Glioma) OR  OR  OR  OR  OR (Mixed Glioma) OR  OR  OR  OR (Astrocytoma) OR (Glioblastoma) OR (Diffuse Intrinsic Pontine Glioma) OR (Ependymoma) OR  OR (Ganglioglioma) OR (Medulloblastoma) OR (Oligodendroglioma) OR (Optic Nerve Glioma)) AND (rs xxxxxxx)\u201d. In addition, we screened relevant reviews and references from previous meta-analyses to increase the number of other eligible studies. The retrieval process was performed by two researchers independently. Any discrepancy was adjudicated by a senior investigator.2.2TERT gene, rs4295627 polymorphism of CCDC26 gene, rs4977756 polymorphism of CDKN2A/B gene and rs6010620 polymorphism of RTEL gene with glioma susceptibility;(1) case-control or cohort study and genome-related study regarding the association of rs2736100 polymorphism of (2) studies containing allelic and genotypic data or ORs and 95% CIs for susceptibility;(3) appropriate statistical methods and reliable data with clear and unambiguous expression of results, allowing calculation of the ORs and 95% CIs;(4) no overlapped data, and studies involving the same data or overlapped by the same authors, only the one that had the largest sample size or the most recently published.2.3(1) review, commentary, abstract, and case report types;TERT (rs2736100), CCDC26 (rs4295627), CDKN2A/B (rs4977756) and RTEL1 (rs6010620);(2) articles with original literature content did not involve (3) articles were unrelated to glioma research;(4) articles with insufficient data, such as those lacking gene frequency information or ORs with 95% CIs;(5) articles with non-human study subjects.2.4Each study covered the data of: the first author\u2019s name, publication year, ethnicity , number of cases and controls with relevant genotypes, ORs and 95% CIs. In addition, the genetic model used in the article was checked. We applied the Newcastle-Ottawa Scale for evaluating the literature quality. Two researchers performed independent data extraction and evaluated the literature quality. A senior investigator took charge of discrepancy arbitration.2.5p<0.05 reported statistical significance). Second, we measured the association degree of TERT (rs2736100), CCDC26 (rs4295627), TERT (rs2736100) and RTEL1 (rs6010620) with glioma risk under five genetic models using the ORs and 95% CIs, which were the RM, O-DM, AM, C-DM and DM.STATA 17.0 MP was implemented for all statistical analyses in this study. First, we performed the Hardy-Weinberg Equilibrium (HWE) test .We merged the ORs and 95% CIs that satisfied the criteria under the same models. In addition, in all genetic models, subgroup analyses were carried out according to ethnic differences and the 33.1We initially searched 640 literature and eventually enrolled 21 eligible articles covering Caucasian, Asian, and others , 17\u201336. 3.23.2.1TERT gene rs2736100 polymorphism, allele G is the risk gene (TERT gene rs2736100 polymorphism led to increased glioma risk in 5 genetic models (AM (G vs T): OR=1.29, 95% CI: 1.25 to 1.34; RM (GG vs TT+TG): OR=1.31, 95% CI: 1.18 to 1.44; DM (TG+GG vs TT): OR=1.52, 95% CI: 1.34 to 1.73; C-DM (GG vs TT): OR=1.69, 95% CI: 1.56 to 1.82; C-DM (GT vs TT): OR=1.40, 95% CI: 1.31 to 1.49; O-DM (TT+GG vs TG): OR=0.93, 95% CI: 0.88 to 0.98). When stratifying the analysis according to ethnicity, we obtained similar results in Caucasians, although in the O-DM results only showed a trend of association with glioma (AM (G vs T): OR=1.28, 95% CI: 1.23 to 1.33; RM (GG vs TT+TG): OR=1.29, 95%CI: 1.13 to 1.47; DM (TG+GG vs TT): OR=1.51, 95% CI: 1.29 to 1.76; C-DM (GG vs TT): OR=1.64, 95% CI: 1.50 to 1.80; C-DM (GT vs TT): OR=1.37, 95% CI: 1.27 to 1.49; O-DM (TT+GG vs TG): OR=0.93, 95% CI: 0.87 to 1.00). Among Asians, we included only one study, and the results were similar to those of Caucasians. In the study of a multi-ethnic population, we included the study of Wrensch and Safaeian which revealed the relevance of the TERT gene rs2736100 polymorphism to increased glioma risk.In the isk gene Figure\u00a023.2.2CCDC26 gene rs4295627 polymorphism, allele T was the risk gene : OR=0.92, 95% CI: 0.81 to 1.03. In one study, the RM (GG vs TT+TG): 0.52; 0.25~1.05; DM (TG+GG vs TT): 0.86; 0.60 to 1.23; C-DM (GG vs TT): 0.50; 0.24 to 1.05; C-DM (GT vs TT): 0.95; 0.65 to 1.38; O-DM (TT + GG vs TG): 0.97; 0.67 to 1.40.3.2.3CDKN2A/B gene rs4977756 polymorphism, allele T was the risk gene : OR=1.03, 95% CI: 0.91 to 1.17. In one study, the RM (GG vs AA+AG): 1.12; 0.53 to 2.34; DM (AG+GG vs AA): 0.96; 0.66 to 1.39; C-DM (GG vs AA): 1.09; 0.52 to 2.32; C-DM (GA vs AA): 0.94; 0.63 to 1.38; O-DM (AA+GG vs AG): 1.08; 0.73 to 1.58. In other study, RM (GG vs AA+AG): 0.68; 0.25 to 1.81; DM (AG+GG vs AA): 0.98; 0.60 to 1.58; C-DM (GG vs AA): 0.69; 0.25 to 1.87; C-DM (GA vs AA): 1.04; 0.63 to 1.73; O-DM (AA+GG vs AG): 0.93; 0.56 to 1.52.3.2.4RTEL1 gene rs6010620 polymorphism, allele T was the risk gene : OR=1.35, 95% CI: 1.22 to 1.48; RM (GG vs AA+AG): 2.04; 1.61 to 2.57; DM (AG+GG vs AA): 1.35; 1.20 to 1.52; C-DM (GG vs AA): 2.41; 1.09 to 3.04; C-DM (GA vs AA): 1.22; 1.07 to 1.38; O-DM (AA+GG vs AG): 0.93; 0.83 to 1.05.3.3Our study adopted Egger\u2019s Test for evaluating the literature publication bias and the results are shown in In rs2376100, there was publication bias in AM (p=0.469). In rs4295627, publication bias existed in dominant, co-dominant and O-DM (DM (TG+GG vs TT): p=0.426; C-DM (GT vs TT): p=0.436; O-DM (TT+GG vs TG): p=0.427). In rs4977756, publication bias existed in O-DM (p=0.111). In rs6010620, publication bias existed in the recessive, dominant and O-DM (RM (GG vs AA+AG): p=0.010; DM (AG+GG vs AA): p=0.014; O-DM (AA+GG vs AG): p= 0.461).4Our study is one of the first ones with conducting the first meta-analysis with the largest sample size as far as we concern and we performed detailed analyses of multiple associated SNPs under five genetic models. Besides, stratified analyses were carried out considering ethnicity. This indicates reliable meta-analysis results.From the meta-analysis, allele G in the SNP at the rs2736100 locus differentially elevated glioma risk, which conforms to studies by Di Stefano . In partTERT is the most important component in regulating telomerase activity. A few scholars found the relevance of TERT expression to glioma grade and patients\u2019 prognosis  presented shorter telomeres relative to people who possessed TG genotype  and p15 (CDKN2B) oncoproteins which play multiple roles in cellular stress recognition, senescence regulation, differentiation as well as apoptosis in the developmental and proliferative phases of cells  regions of genes. Intron SNPs possibly facilitate aberrant exon inclusion or deletion by altering mRNA splicing and the formation and addition regarding nonsense transcripts (RTEL1 gene rs6010620 on 20q13.33 is a vital candidate genetic variant that has been widely reported (RTEL1 was a direct target of miR-4530. The abnormal expression of RTEL1 could lead to obvious reversion of the miR-4530 overexpression in glioma cell lines. The miR-4530/RTEL1 axis acts as a latent treating target specific to gliomas (RTEL1 has been linked to the development of astrocytoma and Hoyeraal-Hreidarsson syndrome (RTEL1 can maintain genomic stability by studying its ability to maintain telomere homeostasis and promote DNA replication, there are still a lot of questions that shall be highlighted. Nowadays, researches have not confirmed the regulation of RTEL1 and its recruitment to impact the replication forks and telomeres (nscripts , 57. Thereported . As repo gliomas . Not onlStudying the association between the risk of glioma and SNPs is definitive, because it could help clinicians use such polymorphisms as pragmatic molecular biomarkers of glioma risk, assist drug developers innovate relevant targeted therapeutic agents for the benefit of patients, and allow for basic researchers to explore the molecular mechanisms of glioma pathogenesis and accurately regulate glioma progression. According to meta-analysis results, racial differences exist in the risk of glioma, but there is a paucity of studies on Asian ethnicity, which requires researchers to conduct relevant studies. Exploration of SNPs at other loci and risk of glioma is also necessary.5Some limitations should be noted when interpreting the current results. The study also presents some limitations, which shall be noted during the interpretation of current studies.First, there were scarce studies on Asian populations. Studies on results in Asian populations should be interpreted with caution.In addition, the absence of more specific individual information about interaction within genes and that between gene and environment prohibited us from conducting a more precise analysis.Finally, some unavoidable publication bias in the results of meta-analysis should also be taken into account.6TERT gene rs2736100 polymorphism, CCDC26 gene rs4295627 polymorphism, CDKN2A/B gene rs4977756 polymorphism and RTEL1 gene rs6010620 polymorphism might all increase the risk of glioma development, but there are ethnic differences. It is necessary to well develop case-control studies with large sample sizes and a focus on more races or glioma types to overcome the limitations described earlier to make the conclusions more accurate.In summary, our study suggested that the All authors contributed to the article and approved the submitted version. YQW: Writing articles, searching literature and extracting literature. JZho and ZT: Analyzing the quality of articles. JZha: Analyzing Data. XC: Providing methodology. SL: Editing of articles."}
+{"text": "Computer-aided detection (CAD) using artificial intelligence (AI) to analyze chest radiographs is an important tool for community tuberculosis (TB) screening in high burden countries. Most current algorithms use a universal cut-off score to select individuals for confirmatory TB testing; however, using a tailored cut-off based on client demographics (age and sex) may improve performance.Community-based TB screening was conducted using portable X-ray with CAD  as part of a cluster-randomized trial in Uganda. Individuals scoring above a specified threshold were offered sputum Xpert Ultra testing. This threshold was initially set at 0.5 (range: 0-1) but was later lowered to 0.2 and then 0.1 for research purposes. For clients with scores \u2265 0.1 who did not undergo Xpert testing, we used multiple imputation to infer Xpert-positive status. We assumed that those with X-ray scores < 0.1 would be Xpert-negative if tested, but considered 0.1% or 0.5% prevalence of Xpert-positive TB in sensitivity analysis. We compared the sensitivity and specificity of using a universal screening threshold of 0.5 versus tailored thresholds based on client age and sex.A total of 15,375 individuals were screened using AI-interpreted digital X-ray, of whom 1,748 (11.4%) had valid sputum Ultra results; 157 (9.0%) tested positive. Assuming that people with qXR scores < 0.1 would test negative on sputum Ultra, the manufacturer-recommended universal threshold of \u2265 0.5 had an estimated sensitivity of 75.8% (95%CI: 70.3-81.6) and specificity of 95.3% (95% CI 95.2-95.4). A sex-differentiated threshold  had similar specificity but significantly higher sensitivity: 81.3% (95% CI 76.3-86.3). Sex-stratified thresholds also outperformed a universal threshold when assuming higher positivity among people with qXR scores < 0.1. Stratification by age did not significantly improve accuracy.Using differentiated CAD score thresholds based on client sex could improve the accuracy of digital X-ray for TB screening. Future research should validate these findings in other populations and explore the value of incorporating additional client characteristics into TB screening algorithms.All Authors: No reported disclosures"}
+{"text": "Correction to: Translational Neurodegeneration (2023) 12:5 10.1186/s40035-023-00337-1Following publication of this article , three eCorrection 1:136. Crocker TF, Brown L, Lam N, Wray F, Knapp P, Forster A. Information provision for stroke survivors and their carers. Cochrane Database Syst Rev. 2021;11:CD001919.Should be corrected as follows:136. Menni C, Valdes AM, Polidori L, Antonelli M, Penamakuri S, Nogal A, et al. Symptom prevalence, duration, and risk of hospital admission in individuals infected with SARS-CoV-2 during periods of omicron and delta variant dominance: a prospective observational study from the ZOE COVID study. Lancet. 2022;399:1618\u201324.Correction 2:Lina and colleagues reported that 51% of patients with VITT present with cerebral venous sinus thrombosis (CVST) (95% CI 36\u201366%) [188].\u201d should be corrected to \u201cPalaiodimou and colleagues reported that 51% of patients with VITT present with cerebral venous sinus thrombosis (CVST) (95% CI 36\u201366%) [188].\u201d\u201c188. Palaiodimou L, Stefanou MI, Katsanos AH, Aguiar De Sousa D, Coutinho JM, Lagiou P, et al. Cerebral venous sinus thrombosis and thrombotic events after vector-based COVID-19 vaccines: a systematic review and meta-analysis. Neurology. 2021;97:e2136\u201347.Correction 3:Shi and colleagues found that APOE \u03b54/\u03b54 genotype induces an increased rate of SARS-CoV-2 infection than the APOE \u03b53/\u03b53 genotype. Moreover, APOE4 astrocytes exhibited a more severe response following SARS-CoV-2 infection. This study provides the first insight into a possible APOE-mediated mechanism of COVID-19 vulnerability and severity [211].\u201d should be corrected to \u201cIn human induced pluripotent stem cell (iPSC)-derived neurons and astrocytes, Wang and colleagues found that APOE \u03b54/\u03b54 genotype induces an increased rate of SARS-CoV-2 infection than the APOE \u03b53/\u03b53 genotype. Moreover, APOE4 astrocytes exhibited a more severe response following SARS-CoV-2 infection. This study provides the first insight into a possible APOE-mediated mechanism of COVID-19 vulnerability and severity [211].\u201d\u201cIn human induced pluripotent stem cell (iPSC)-derived neurons and astrocytes, 211. Wang C, Zhang M, Garcia G Jr, Tian E, Cui Q, Chen X, et al. ApoE-isoformdependent SARS-CoV-2 neurotropism and cellular response. Cell Stem Cell. 2021;28:331\u201342.e5.The original article  has been"}
+{"text": "This cohort study evaluates the incidence of COVID-19 and hospitalizations across variant eras in 2021 and 2022 among vaccinated solid organ transplant (SOT) recipients. Understanding longitudinal COVID-19 outcomes on a population scale has implications for future counseling and risk prediction; thus, we quantified changes in COVID-19 burden among transplant recipients, hypothesizing reduced disease severity in recent variant eras among SARS-CoV-2 vaccine uptake.Solid organ transplant recipients experienced severe COVID-19 outcomes before vaccination and high breakthrough rates despite primary vaccine series.STROBE reporting guideline and was approved by Johns Hopkins University, with waiver of documentation of informed consent because of no more than minimal risk intervention.Transplant recipients reporting 1 or more SARS-CoV-2 vaccination within a national prospective cohort were surveyed following immunoprophylaxis events to ascertain incident COVID-19 (self-reported positive molecular or antigen test result) and hospitalizations between January 2021 and December 2022, with additional cohortwide surveys in January and June 2022 and unsolicited reporting  were compared between post-BA.1 and pre-Delta, Delta, and BA.1 waves using Poisson regression. Two-sided P\u2009<\u2009.05 was significant. Stata/SE, version 17 was used for analysis.Cohort characteristics, COVID-19 incidence rate, and hospitalization ratio were reported by variant era: pre-Delta (January to May 2021), Delta (June to December 2021), and Omicron BA.1 (January to March 2022), BA.2 (April to June 2022), and BA.4-BA.5-BQ.1 (July to December 2022). Monthly transplant recipient COVID-19 incidence rate was plotted against US population case counts.P\u2009<\u2009.001) and similar if less than 2 years vs 2 years or more from transplant  and in lung vs nonlung recipients . Transplant recipient COVID-19 incidence paralleled US population case counts  responded to 1 or more survey. Vaccinations increased over time . COVID-1e counts , A, peakP\u2009=\u2009.02) and lung transplant . Hospitalization ratios (per 100 incident infections) were 14.1 for pre-Delta; 11.8, Delta; 9.2; BA.1; 2.3, BA.2; and 3.5, BA.4-BA.5-BQ.1 (P\u2009=\u2009.75) and less than 2 years vs 2 years or more from transplant  yet higher in lung vs nonlung recipients .There were 37 COVID-19-related hospitalizations among 35 of 464 participants (7.5%). Hospitalized participants more often reported stronger immunosuppression , with a similar ratio of serious COVID-19 disease . Of 16 reported deaths, 5 were COVID-19 related (1 post-BA.1); 6, non-COVID-19 related (3 post-BA.1); and 5, unknown cause (4 post-BA.1).Hospitalization ratio comparing post-BA.1 vs pre-Delta, Delta, and BA.1 was 0.27 (95% CI, 0.11-0.69; In this study, 19.7% of participants reported incident COVID-19, 7.5% of whom required hospitalization. Transplant recipient COVID-19 incidence followed US case count trends, with higher incidence in later Omicron waves potentially due to home test capture and/or increased susceptibility to SARS-CoV-2 variants.4 improved therapeutics,5 home testing access, and/or differing viral pathogenicity.6 Limitations include sampling and recall bias due to observational study design, lack of SARS-CoV-2 sequencing or longitudinal testing frequency, and incomplete death ascertainment.COVID-19-related hospitalization among transplant recipients markedly decreased after the BA.1 wave, likely owing to changing population immunity,"}
+{"text": "Time to appropriate antimicrobial therapy is the most important predictor of survival among patients with sepsis due to Gram-negative bacteria (GNB). Infections due to multidrug-resistant (MDR) pathogens often result in inappropriate empiric treatment (tx). The GenMark ePlex identifies organisms and resistance markers within 1.5 hours from blood culture positivity compared to 48-72 hours by conventional methods.We are conducting a single-center study to assess outcomes before and after ePlex implementation. The pre-intervention period included patients with GN bacteremia from January to June 2021. Patients were excluded if they received comfort-only care within 48 hours, had polymicrobial GN bacteremia, or were infected by an organism not on the ePlex panel. Primary outcome is time to in-vitro active therapy from blood culture collection.Table 1). 16% of patients were infected by MDR GNB and 21% received inactive empiric tx. Overall median (IQR) time to in-vitro active tx was 3.9 (1\u201319) hours, but was 39.5 (13.6-71.9) hours among those who received inactive empiric tx. Median (IQR) time to first antibiotic modification was 2.7 (1.0-3.4) days. 38% of patients were transitioned to oral antibiotics at a median (IQR) of 4.2 (3.1-6.1) days. The median (IQR) duration of tx was 15 (10-17) days; 4.5% of patients were treated with \u2264 7 days. Median (IQR) length of hospitalization was 15.5 (7-36) days. 7.8% were re-admitted within 30 days (Table 2). In-hospital and 30-day mortality rates were numerically higher among patients who received inactive empiric therapy  when compared to patients who received active empiric therapy  .208 patients were identified during the pre-intervention period; 154 met inclusion criteria. Median age was 64 years, 49% were male, and 28% were immunocompromised. Median (IQR) Charlson Comorbidity index and Pitt Bacteremia scores were 6 (4-8) and 2 (1-8), respectively (Results Table 1: Patient demographics, underlying conditions and infection and treatment characteristics in the pre-intervention groupResults Table 2Results Figure 1Our pre-intervention data highlight opportunities to improve the management of GN bacteremia. Implementation of ePlex is likely to decrease the proportion of patients treated with inactive therapy, shorten time to optimal tx and reduce lengths of stay.Ryan K. Shields, PharmD, MS, Allergan: Advisor/Consultant|Cidara: Advisor/Consultant|Entasis: Advisor/Consultant|GSK: Advisor/Consultant|Melinta: Advisor/Consultant|Melinta: Grant/Research Support|Menarini: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Pfizer: Advisor/Consultant|Roche: Grant/Research Support|Shionogi: Advisor/Consultant|Shionogi: Grant/Research Support|Utility: Advisor/Consultant|Venatorx: Advisor/Consultant|Venatorx: Grant/Research Support"}
+{"text": "Arthritis is a leading cause of activity limitations, disability, and chronic pain, and is associated with dispensed opioid prescriptions, substantially contributing to health care costs.During 2019\u20132021, 21.2% of U.S. adults (53.2 million) reported diagnosed arthritis. Approximately one half (52.2%\u201362.4%) of adults aged \u226565 years with self-reported diagnosed dementia, chronic obstructive pulmonary disease, stroke, heart disease, diabetes, or cancer also had a reported diagnosis of arthritis.These prevalence estimates can be used to guide public health policies and activities to increase equitable access to physical activity opportunities within the built environment and other community-based, arthritis-appropriate, evidence-based interventions.Arthritis includes approximately 100 conditions that affect the joints and surrounding tissues. It is a leading cause of activity limitations, disability, and chronic pain, and is associated with dispensed opioid prescriptions, substantially contributing to health care costs. Combined 2019\u20132021 National Health Interview Survey data were analyzed to update national prevalence estimates of self-reported diagnosed arthritis. An estimated 21.2% (18.7% age-standardized) of U.S. adults aged \u226518 years (53.2 million) had diagnosed arthritis during this time frame. Age-standardized arthritis prevalences were higher among women (20.9%) than men (16.3%), among veterans (24.2%) than nonveterans (18.5%), and among non-Hispanic White (20.1%) than among Hispanic or Latino (14.7%) or non-Hispanic Asian adults (10.3%). Adults aged \u226545 years represent 88.3% of all U.S. adults with arthritis. Unadjusted arthritis prevalence was high among adults with chronic obstructive pulmonary disease (COPD) (57.6%), dementia (55.9%), a disability (54.8%), stroke (52.6%), heart disease (51.5%), diabetes (43.1%), or cancer (43.1%). Approximately one half of adults aged \u226565 years with COPD, dementia, stroke, heart disease, diabetes, or cancer also had a diagnosis of arthritis. These prevalence estimates can be used to guide public health policies and activities to increase equitable access to physical activity opportunities within the built environment and other arthritis-appropriate, evidence-based interventions.Arthritis includes approximately 100 conditions that affect the joints and surrounding tissues. If not managed properly, arthritis can result in severe pain, activity limitations, and disability  is an annual, nationally representative household survey of the noninstitutionalized U.S. civilian population.NHIS sample sizes and response rates for 2019, 2020, and 2021 were 31,997 (59.1%), 21,153 (48.9%), and 29,482 (50.9%), respectively.Approximately 53.2 million (95% CI = 52.1\u201354.4) or 21.2% (18.7% age-standardized) of U.S. adults aged \u226518 years had diagnosed arthritis . Age-sta2; 56.8%), dementia (56.1%), diabetes (54.0%), or cancer (52.2%) also had diagnosed arthritis  were aged \u226565 years, and 40.0% (21.3 million) were aged 45\u201364 years. Age-standardized prevalence of arthritis was higher among adults with dementia (47.8%),rthritis  Figure)2; 56.8%)During 2019\u20132021, 53.2 million (21.2%) U.S. adults aged \u226518 years had diagnosed arthritis. Approximately one half of adults aged \u226565 years with a chronic disease also reported diagnosed arthritis. Consistent with previous NHIS"}
+{"text": "Andixius Emeljanov & Hayashi, 2007 are described and illustrated from China. These are A.flagellihamus Wang & Chen, sp. nov., A.gracilispinus Wang & Chen, sp. nov., A.productus Wang & Chen, sp. nov. and A.truncatus Wang & Chen, sp. nov. Photographs of the new species and an identification key to all Andixius species are provided.Four new species of the genus  Andini  consists of 129 species in three genera worldwide  and A.venustus  . A.longispinus and A.trifurcus, to the genus. Later, A.cultratus and A.lingulatus.The planthopper tribe orldwide . Within A.flagellihamus Wang & Chen, sp. nov., A.gracilispinus Wang & Chen, sp. nov., A.productus Wang & Chen, sp. nov. and A.truncatus Wang & Chen, sp. nov., which are described here. Hence, the number of Andixius species is now 10, with nine species occurring in China.Recent study of some Chinese specimens has found four new species, The morphological terminology follows IEGU).The type specimens are deposited in the Institute of Entomology, Guizhou University, Guiyang, Guizhou Province, China , Japan (Ryukyu Islands).A.cultratus Wang, Zhi & Chen, 2020; China (Guangdong).A.flagellihamus Wang & Chen, sp. nov.; China (Xizang).A.gracilispinus Wang & Chen, sp. nov.; China (Xizang).A.lingulatus Wang, Zhi & Chen, 2020; China (Guangxi).A.longispinus Zhi & Chen, 2018; China (Yunnan).A.nupta Emeljanov & Hayashi, 2007; Japan (Ryukyus).A.productus Wang & Chen, sp. nov.; China (Xizang).A.trifurcus Zhi & Chen, 2018; China (Yunnan).A.truncatus Wang & Chen, sp. nov.; (Guangxi).A.venustus ; China (Taiwan).Taxon classificationAnimaliaHemipteraCixiidae\ufeffWang & Chensp. nov.4A130AF2-7552-5EC5-BFB7-D1E5DCAE7E08https://zoobank.org/5BB7E534-9C4C-4507-8FA5-CA5C1D3D5646Holotype: \u2642, China: Xizang Province, Medog County, Beibeng Town , 15 August 2020, Yongjin Sui leg.; Paratypes: \u2642, same data as holotype.n = 2).Body length: male 6.65\u20137.00 mm  Fig. .flagellihamus, referring to the 1-hooked spinose process arising from the apex of the endosoma.The specific name is derived from the Latin adjective This species can be distinguished from the other species of the genus by the following characters: basal right side of periandrium with a U-shaped spinose process; basal ventral margin of periandrium with a long spinose process, apex curved upwards, forming a hooked process; apex of endosoma with a hooked spinose process.Taxon classificationAnimaliaHemipteraCixiidae\ufeffWang & Chensp. nov.BECD8207-9CC0-51A5-91D1-51B9D3F7165Chttps://zoobank.org/4CF579B2-3F5B-4926-99D8-59E7F8E16E85Holotype: \u2642, China: Xizang Province, Bom\u00ea County, Yigong Town, Tongmai Village , 18\u201320 August 2020, Yongjin Sui leg.; Paratypes: \u2642, same data as holotype.n = 2).Body length: male 5.63\u20135.82 mm  Fig. .gracilispinus, referring to the one long spinose process arising from the apical right side of the ventral margin of the periandrium.The specific name is derived from the Latin adjective A.gracilispinus sp. nov. are similar to A.venustus Tsaur & Hsu, 1991 in appearance, but differs in: (1) basal left side of ventral margin of periandrium with a triangular laminal process, of which middle right side concaved heavily, forming two large processes (A.venustus with a spinose process in the same position); (2) near apical right side of ventral margin of periandrium with a long spinose process, slightly curved ; (3) basal dorsal margin of periandrium with a grooved laminal process (without process in A.venustus).Male genitalia of Taxon classificationAnimaliaHemipteraCixiidae\ufeffWang & Chensp. nov.F58ABCA6-C31E-5A1D-A365-33C0937A010Chttps://zoobank.org/AE3FAE97-597C-41D8-92B8-7C8A2CCB5464Holotype: \u2642, China: Xizang Province, Medog County, Damu Town, 80K , 18 August 2020, Yongjin Sui leg.; Paratypes: 6\u2642\u2642 2\u2640\u2640, same data as holotype.n = 7), female 7.78\u20137.90 mm (n = 2).Body length: male 5.71\u20136.90 mm  Fig. .productus, referring to the one long spinose process arising from the apical ventral margin of the periandrium.The specific name is derived from the Latin adjective A.productus sp. nov. are similar to A.trifurcus Zhi & Chen, 2018, but differs in: (1) periandrium with an expanded laminal process around the left side and dorsal margin of periandrium ; (2) basal ventral margin of periandrium with laminal process, of which ventral margin with two processes (A.trifurcus with three long spinose processes in the same position); (3) right side of ventral margin of periandrium with three spinose processes (right side of middle part of periandrium with a spinose process in A.trifurcus); (4) middle part of periandrium with a thick and long spinose process (without process in A.trifurcus).Male genitalia of Taxon classificationAnimaliaHemipteraCixiidae\ufeffWang & Chensp. nov.6AAFC595-BABB-5FB0-862A-EE78D8C83C99https://zoobank.org/1EE35403-E65B-43C5-B171-54C7CC313B23Holotype: \u2642, China: Guangxi Province, Longsheng County, Huaping National Natural Reserve , 18 July 2020, Xiaoya Wang, Yongjin Sui, Zhicheng Zhou and Jing Wang leg.; Paratypes: 9\u2642\u2642 5\u2640\u2640, same data as holotype.n = 10), female 7.25\u20138.86 mm (n = 5).Body length: male 6.56\u20137.20 mm  Fig. .truncatus, referring to the ventral margin of periandrium with a long, broad laminal process having a truncated apex.The specific name is derived from the Latin adjective Andixius species by the following characters: forewing general black-brown, with an irregular yellowish-white spot slightly below stigma and near claval fork; ventral margin of periandrium with a long and broad laminal process, apex truncated, margin with small teeth; endosoma curving ventrally in right angle, base of dorsal margin with a long spinose process.This species can be distinguished from the other Andixius once again emphasizes the need for further study on the group based on male genitalia whenever possible , of which only the latter two genera and 18 species occur in China  forming a short common stalk, while ScP, RP and MP emerge independently or very close to the basal cell in the other Andini genera. The outer edge of the apical half of the fore coxae is extended and smoothly protruding in Parandes, but the outer edge of the apical half of the fore coxae is straight and does not extend in Andes.Currently the tribe in China . A compa"}
+{"text": "Figure 2 as published. In the legend for Figure 2, \u201c(N) Brown adipose tissue (BAT).\u201d is a duplicate and needs to be deleted because the BAT is explained in the legend for Figure 2I. The corrected legend appears below.In the published article, there was an error in the legend for (A) Body weight (BW). (B) BW gain. (C\u2013E) Organ wet weight. (F) Inguinal white adipose tissue (iWAT). (G) Perirenal white adipose tissue (pWAT). (H) Epididymis white adipose tissue (eWAT). (I) Brown adipose tissue (BAT). (J) iWAT/BW ratio. (K) pWAT/BW ratio; (L) eWAT/BW rati. (M) BAT/BW ratio. (N) Representative rat liver images of hematoxylin and eosin (H and E) and Oil Red O staining per group (X200). (O) Representative iWAT, pWAT, eWAT, BAT. One-way analysis of variance (ANOVA) was conducted for the group comparison. n = 8, data are presented as mean \u00b1 SEM.*p < 0.05, **p < 0.01, ***p < 0.001 vs. MOD group. EPC, P. cocos ethanol extract; CON, normal diet control group; MOD, high-fat diet group; FC, Fenofibrate capsules; EPC-L, low-dose P. cocos ethanol extract; EPC-H, high-dose P. cocos ethanol extract.].\u201cFIGURE 2 | EPC ameliorated MAFLD in rats. Furthermore, there was an error in"}
+{"text": "Translational Psychiatry 10.1038/s41398-023-02499-y, published online 09 June 2023Correction to: The reference numbers in the Figs. 2 and 3 were given not correct. The figures have been corrected.Corrected Fig. 2Corrected Fig. 3"}
+{"text": "Bacterial nitrogen (N) fixation in alder nodules is a key process providing nitrogen to nutrient-limited arctic biomes. Here, 45 prokaryotic metagenome-assembled genome (MAG) sequences from root nodules of arctic alder are reported. Alnus viridis subsp. fruticosa, an N-fixing deciduous alder shrub that is an important contributor to N cycling in high-latitude ecosystems , which is located 103\u2009km from Nome, Alaska. At this location, alder is found in two distinct communities, short and dispersed alder growing in lowland areas (savanna) and tall alder growing as dense shrublands along the hillslope (shrubland) . Sequence processing and metagenome-assembled genome (MAG) generation were performed via metaWRAP v1.1  of \u22651\u2009kb. Contigs were binned with MaxBin2 v2.2.5 (https://tinyurl.com/4dtt38mt) by minimum information about a metagenome-assembled genome (MIMAG) standards  . DNA froRAP v1.1 . DefaultRAP v1.1 , resulti2 v2.2.5  to detertandards . GTDB-Tktandards  with thetandards , was use1.1 N50, ,779\u2009bp oPRJNA830531, and nodule metagenomes were deposited under SRA accession no. SRR18933204 for alder savanna and SRR18933205 for alder shrubland. GenBank accession numbers for the MAGs are listed in https://figshare.com/articles/dataset/Quality_metrics_for_metagenome-assembled_genomes_of_Alnus_viridis_spp_Fruticose_nodules/22096520/1.MAGs were deposited in the National Center for Biotechnology Information (NCBI) BioProject database under accession no."}
+{"text": "Citrus sinensis leaves and roots were investigated. Our findings indicated that increased pH mitigated Cu toxicity-induced alterations of HRMs, and Cu toxicity increased low-pH-induced alterations of HRMs. Increased pH-mediated decreases in ABA, jasmonates, gibberellins, and cytokinins, increases in (\u00b1)strigol and 1-aminocyclopropanecarboxylic acid, and efficient maintenance of salicylates and auxins homeostasis in 300 \u03bcM Cu-treated roots (RCu300); as well as efficient maintenance of hormone homeostasis in 300 \u03bcM Cu-treated leaves (LCu300) might contribute to improved leaf and root growth. The upregulation of auxins (IAA), cytokinins, gibberellins, ABA, and salicylates in pH 3.0 + 300 \u03bcM Cu-treated leaves (P3CL) vs. pH 3.0 + 0.5 \u03bcM Cu-treated leaves (P3L) and pH 3.0 + 300 \u03bcM Cu-treated roots (P3CR) vs. pH 3.0 + 0.5 \u03bcM Cu-treated roots (P3R) might be an adaptive response to Cu toxicity, so as to cope with the increased need for reactive oxygen species and Cu detoxification in LCu300 and RCu300. Increased accumulation of stress-related hormones (jasmonates and ABA) in P3CL vs. P3L and P3CR vs. P3R might reduce photosynthesis and accumulation of dry matter, and trigger leaf and root senescence, thereby inhibiting their growth.The effects of copper (Cu)\u2013pH interactions on the levels of hormones and related metabolites (HRMs) in  Citrus orchards due to long-term and heavy application of Cu-containing fungicides against fruit and foliar diseases and pests [Like other heavy metals, micronutrient copper (Cu) has high phytotoxicity at high concentrations . Currentnd pests ,3. Sincend pests . Root grnd pests ,6,7.3 (GA3), 1-naphthylacetic acid , indole-3-acetic acid (IAA), jasmonic acid (JA), salicylic acid (SA), 6-benzyladenine (BAP), and kinetin can mitigate Cu toxic inhibition on plant growth by reducing Cu uptake, inhibiting Cu translocation from roots to shoots, improving nutrient status, maintaining cellular redox homeostasis and/or preventing oxidative damage [Phytohormones, namely cytokinins (CKs), gibberellins (GAs), auxins (AUXs), abscisic acid (ABA), ethylene (ETH), jasmonates (JAs), salicylates (SAs), and strigolactones (SLs) play a role in plant Cu tolerance . Evidence damage ,15,16,173, and GA4 in maize (Zea mays) root apex, concluding that Cu toxicity-induced alterations of hormonal homeostasis at the root apex contributed to the strong root growth inhibition. Reckova et al. [Citrus grandis leaves, Cu excess led to increased levels of indole-3-lactic acid (ILA), IAA, ABA, L-tryptophan (TRP), cis-zeatin-9-glucoside (cZ9G), and total AUXs; and decreased levels of 5-deoxystrigol (5DS) and N6-benzyladenine-7-glucoside (BAP7G) [Raphanus sativus) hypocotyls [3 in Arabidopsis thaliana roots, and dihydrozeatin riboside (DHZR) and t-ZR in A. thaliana shoots [Populus deltoides), sunflower (Helianthus annuus), and maize leaves and roots [P. deltoids) [Cu toxicity had great impacts on hormone biosynthesis and concentrations in various plant tissues (organs). Matayoshi et al.  observeda et al.  reported (BAP7G) . Other rpocotyls ; trans-za shoots ; ABA in nd roots ,23,24; aCitrus sinensis roots, concluding that increased pH-induced decrease in total CKs, and increases in IAA, SA, total JAs, JA and methyl jasmonate (MEJA) in Al toxic roots might confer Al-tolerance by reducing Al uptake, increasing the Al-triggered release of malate and citrate and Al sequestration in vacuole, and maintaining the homeostasis of nutrients and the balance between ROS and MG biosynthesis and removal. Thus, hormones might play a role in elevated pH-mediated mitigation of Cu toxicity in plants. Currently, such data are very rare.Evidence shows that high pH can counteract the adverse impacts of Cu toxicity on plants ,25. MostCitrus are mainly planted in acid soils with high Cu bioavailability [C. sinensis leaves and roots. The objectives were (a) to elucidate how Cu\u2013pH interactions affect the abundances of HRMs in leaves and roots, and (b) to test the hypothesis that hormones play a role in elevated pH-mediated amelioration of Cu toxicity.lability . Here, wWe tested 88 HRMs in leaves , 55 of wWe detected higher concentrations of iP7G, tZOG, 2MeScZR, DHZ7G, cZ9G, BAP7G, and total CKs, and lower concentrations of DHZR and IPR in P3CL vs. P3L; higher concentration of IPR in P5CL vs. P5L and P3L vs. P5L; and higher concentrations of iP7G, tZOG, DHZ7G, cZ9G, and total CKs, and lower concentrations of DHZR and IPR in P3CL vs. P5CL. BAP9G was detected only in P3CL and P5CL, and its level was similar between the two. K9G, IP, and cZ were not detected in P5L, P5L, and P3CL, respectively, and their levels were similar among the other three detected treatments .We observed higher concentrations of IAA-Glu, TRP, IAA, IAA-Trp, IAA-Val, MEIAA, ICAld, ILA, IAN, and total AUXs in P3CL vs. P3L; higher concentration of IAA-Trp in P5CL vs. P5L; lower concentration of IAA-Val in P3L vs. P5L; and higher concentrations of IAA-Glu, TRP, IAA-Trp, IAA, IAA-Val, MEIAA, ICAld, ILA, and total AUXs in P3CL vs. P5CL. IAA-Leu and IAGlc were not detected in P3CL and P5CL, respectively, and their concentrations did not significantly differ among the other three detected treatments. TRA and IAM were detected only in P3CL .24, ABA, ABA-GE, total ABAs, SAG, SA, total SAs, ST, and total SLs, and lower concentrations of H2JA, OPDA, and 5DS in P3CL vs. P3L; higher concentration of JA-ILE in P5CL vs. P5L; higher concentrations of H2JA and OPDA in P3L vs. P5L; and higher concentrations of ABA, SAG, total SAs, ST, and total SLs, and lower concentrations of H2JA, JA-Val and 5DS in P3CL vs. P5CL  and roots (56) to Cu\u2013pH interactions. PC1 and PC2 accounted for 43.25% and 14.05% for leaves, and 39.35% and 21.62% for roots .We detected 25 upregulated HRMs, upregulated total CKs, total AUXs, total ABAs, total SAs, and total SLs, and seven downregulated HRMs in P3CL vs. P3L; 18 upregulated HRMs, upregulated total CKs, total AUXs, total SAs, and total SLs, and seven downregulated HRMs in P3CL vs. P5CL; six upregulated and one downregulated HRMs in P5CL vs. P5L; five upregulated and one downregulated HRMs in P3L vs. P5L; 28 upregulated HRMs, upregulated total CKs, total AUXs, and total SAs, and six downregulated HRMs in P3CR vs. P3R; 25 upregulated HRMs, upregulated total AUXs, total CKs, total GAs, and total SAs, and three downregulated HRMs in P3CR vs. P5CR; 11 upregulated HRMs, 17 downregulated HRMs, and downregulated total CKs, total JAs and total GAs in P5CR vs. P5R; and seven upregulated HRMs, 17 downregulated HRMs, and downregulated total CKs, total JAs and total GAs in P3R vs. P5R . These rA. thaliana root tips. Exogenous application of 1-NAA mitigated Cu excess-induced reduction in primary root growth, while application of 1-N-naphthylphthalamic acid  promoted Cu excess-induced reduction in primary root growth. Choudhary et al. [A. thaliana cotyledons and root apices, and decreased hypocotyl and primary root lengths and cotyledon area, concluding that endogenous hormonal balance and signal transduction played a role in Cu excess-triggered severe morphological responses. Sofo et al. [A. thaliana root and shoot growth, increased IAA concentration in roots, and did not affect IAA concentration in shoots. Wang et al. [A. thaliana seedlings. Wu et al. [C. grandis leaves, concluding that Cu toxicity-induced upregulation of auxin biosynthesis, transport, and levels might contribute to leaf Cu-tolerance by enhancing photosynthesis and water use efficiency (WUE). Ouzounidou and Ilias [Pisum sativum) seedlings Cu toxicity by decreasing Cu concentrations in shoots and roots and providing a thiol redox state to protect the proteins against oxidation. In Brassica juncea, foliar spray of IAA-mediated alleviation of Cu toxicity involved the increase in antioxidant capacity and the decreases in ROS, malondialdehyde (MDA), and electrolyte leakage levels in Cu toxic plants. Additionally, IAA application can improve plant nutrient status; maintain leaf function and photosynthesis; reduce root cell death; and ultimately increase Cu toxic plant biomass [A. thaliana roots, which might contribute to Cu toxicity-induced inhibition of primary root elongation. TRP can act as a precursor for the biosynthesis of IAA and melatonin, which play a role in plant Cu-tolerance [We obtained upregulated total AUXs, TRA, IAM, TRP, IAA-Trp, IAA-Glu, MEIAA, IAA, ICAld, ILA, IAN, and IAA-Val, and downregulated IAA-Leu in P3CL vs. P3L; upregulated total AUXs, MEIAA, ICAld, IPA, TRA, IAA-Glu, IAA-Phe-Me, IAA, OxIAA, ICA, TRP, and IAA-Asp, and downregulated IAA-Val and IAA-Leu in P3CR vs. P3R; upregulated IAA-Trp and downregulated IAGlc in P5CL vs. P5L; and upregulated IAA-Glu, OxIAA, IAA-Asp, and IAA, and downregulated ICA, ICAld, and IAM in P5CR vs. P5R . Auxin hy et al.  observedy et al.  observedo et al.  observedg et al.  found thu et al.  identifind Ilias  found thnd Ilias  observed biomass . Yuan et biomass  showed tolerance . Collectolerance . The incWe detected upregulated total AUXs, IAGlc, TRA, IAM, TRP, IAA-Glu, MEIAA, ILA, IAA, IAA-Trp, ICAld, and IAA-Val, and downregulated IAA-Leu in P3CL vs. P5CL; upregulated total AUXs, IAA, ICA, TRA, MEIAA, ICAld, IPA, IAA-Phe-Me, and TRP in P3CR vs. P5CR; downregulated IAA-Val in P3L vs. P5L; and upregulated IAA-Leu and IAA-Val, and downregulated IAN, ICA, and IAM in P3R vs. P5R , suggestNicotiana tabacum) plants had decreased shoot (leaf) growth, but enhanced root growth, concluding that CKs played an opposite role in regulating shoot and root growth. Foliar application of two CKs (kinetin and BAP) conferred castor (Ricinus communis) Cu-tolerance by reducing Cu level in shoots and improving antioxidant ability [A. thaliana roots might contribute to Cu toxicity-induced inhibition of root growth [A. thaliana seedlings. Massot et al. [Phaseolus vulgaris) prior to Al-induced inhibition of root growth. Al-induced rapid increment of CKs might contribute to root growth inhibition either directly or indirectly by influencing hormone homeostasis. Plant metallothioneins (MTs) have been shown to play a role in Cu-tolerance by lowering Cu toxicity-induced oxidative damage and binding Cu [C. sinensis seedlings, Al toxicity led to increased and decreased total CKs concentrations in pH 3.0- and pH 4.0-treated roots, respectively. Increased pH lowered the accumulation of CKs in roots, thereby ameliorating Al toxicity-induced inhibition of root growth [As shown in  ability . Observat growth . Sofo ett growth  reportedt et al.  observednding Cu . Thomas nding Cu  suggestending Cu . The elending Cu ,26. In Ct growth . Taken tC. sinensis roots [Atractylodes lancea roots was greater at pH 5.3 than at pH 6.0 [We detected upregulated total CKs, DHZ7G, iP7G, cZ9G, and tZOG, and downregulated DHZR, IPR, and cZ in P3CL vs. P5CL; upregulated total CKs, pT, K9G, 2MeSiP, BAP9G, cZ, IP, 2MeSiPR, tZOG, tZ, and cZROG, and downregulated tZR in P3CR vs. P5CR; upregulated K9G, IP, and IPR in P3L vs. P5L; and upregulated 2MeSiP, tZ and pT, and downregulated total CKs, tZR, BAPR, tZOG, IPR, BAP, and BAP7G in P3R vs. P5R . This agis roots , and that pH 6.0 . These fPhaseolus coccineus and Allium cepa root growth, and maize leaf growth, and that JA synthesis inhibitors-namely ibuprofen (IB) and salicylhydroxamic acid (SHAM), mitigated Cu toxicity-induced growth inhibition in P. coccineus roots and maize leaves, but not in A. cepa roots. Using metabolome and transcriptome analyses, Hu et al. [Cucumis melo) via repressing JA biosynthesis. Taken together, increased pH prevented Cu toxicity-induced changes in JAs in leaves, thus alleviating leaf Cu toxicity; while Cu toxicity lowered the levels of JAs in pH 4.8-treated roots, thus improving root growth.As shown in u et al.  demonstrWe detected downregulated H2JA and JA-Val in P3CL vs. P5CL; upregulated JA-ILE in P3CR vs. P5CR; upregulated H2JA and OPDA in P3L vs. P5L; and downregulated total JAs, JA-ILE, OPC-4, JA-Val, JA, and OPDA in P3R vs. P5R , suggestC. grandis leaves [Populus x euramericana), B229 and PE19/66. Cu toxicity increased ABA concentrations in PE19/66 leaves and roots, and in B229 leaves, but had no significant impacts on ABA concentrations in M1 leaves and roots, and in B229 roots. Obviously, Cu toxicity impacts on ABA concentrations depended on Cu concentration, pH, tissue (organ), and genotype.As shown in s leaves , and mais leaves . Cu exces leaves . Kebert s leaves  investigo/ABS) in P3CL vs. P3L, but not in P5CL vs. P5L [Penicillium funiculosum LHL06 mitigated the synergistic toxicity of heavy metals on soybean (Glycine max) by reducing uptake of heavy metals, accumulation of ABA and JA, and oxidative damage due to decreased H2O2 accumulation and enhanced antioxidant system in roots. Zehra et al. [2O2 production rates in pH 3.0-treated leaves and roots, but not in pH 4.8-treated leaves and roots [ABA can increase non-photochemical quenching (NPQ), the first line of defense to protect photosystem II (PSII) reaction centers against photo-oxidative damage . The upr vs. P5L . The inc vs. P5L . Bilal e vs. P5L  indicatea et al.  reporteda et al. . A studynd roots . Taken tVicia faba). The critical pH, below which net H+ exudation and root growth ceased, were 3.5 for maize and 4.00 for broad bean at 1 mM Ca2+. With the decrease in pH of the medium, both H+ exudation and root growth decreased gradually. Additional ABA in root medium led to increased H+ exudation and root growth for maize at pH 4.0, but decreased H+ exudation and root growth for broad bean at pH 4.1. Yang et al. [C. sinensis roots decreased with the decrease in pH. Taken together, low pH increased ABA concentration and decreased H+ exudation in RCu300, thus reducing root growth, but not in RCK. This agreed with the report that low pH affected root growth more at 300 \u03bcM Cu than at 0.5 \u03bcM Cu [We detected upregulated ABA in P3CL vs. P5CL; upregulated ABA in P3CR vs. P5CR; and downregulated ABA in P3R vs. P5R . The uprg et al.  observedSalvia officinalis [Oryza sativa) [Gossypium spp.) [As shown in icinalis , bean [4icinalis , rice (O sativa) , sunflow sativa) , and cotum spp.)  by reducum spp.) ,50.We detected upregulated SAG and total SAs in P3CL vs. P5CL and P3CR vs. P5CR, and downregulated SA in P3R vs. P5R , suggest24 in P3CL vs. P3L, upregulated GA9 and GA24 in P3CR vs. P3R, and downregulated GA1 and total GAs in P5CR vs. P5R. Saleem et al. [3 mitigated Corchorus capsularis Cu toxicity by enhancing growth and photosynthesis, and reducing oxidative damage. Additionally, GA3 increased Cu accumulation in roots, stems, and leaves. Similar results have been obtained for pea [4 is biosynthesized from GA24 via GA9 [24 in P3CL vs. P3L and GA24 and GA9 in P3CR vs. P3R might be an adaptive strategy to Cu toxicity, while the downregulation of GA1 and total GAs in P5CR vs. P5R and unaltered GAs in P5CL vs. P5L agreed with the finding that raised pH lessened Cu accumulation in Cu toxic roots, stems, and leaves [As shown in m et al.  observed for pea  and sunf for pea . Bioacti via GA9 . Thus, t7, GA1, GA24, GA9, and total GAs in P3CR vs. P5CR, and upregulated GA7 and downregulated GA1 and total GAs in P3R vs. P5R. The upregulation of GA7, GA1, GA24, GA9, and total GAs in P3CR vs. P5CR might be an adaptive response to low pH, while the downregulation of GA1 and total GAs in P3R vs. P5R agreed with the report that low pH increased Cu concentrations in roots, stems and leaves less under 0.5 \u03bcM Cu than under 300 \u03bcM Cu [As shown in Brassica juncea) was reduced by ethephon (ethylene source) and sulfur, which resulted in reduced oxidative damage, upregulated antioxidant system, and elevated photosynthesis. Both ethephon- and sulfur-treated plants displayed more tolerance to Cd. S-mediated alleviation can be reversed by aminoethoxyvinylglycine (ethylene biosynthesis inhibitor). Thao et al. [We obtained upregulated ACC in P5CR vs. P5R . Flora eo et al.  suggesteWe identified downregulated ACC in P3CR vs. P5CR, and upregulated ACC in P3R vs. P5R , suggestHordeum vulgare) Cd-tolerance by reducing Cd uptake and oxidative damage due to reduced H2O2 accumulation and elevated concentrations of ascorbate and reduced glutathione and activities of antioxidant enzymes in leaves and roots. Mostofa et al. [d10 and d17 displayed less tolerance to arsenic stress accompanied by increased arsenic concentration, decreased sequestration of arsenic in vacuole, and enhanced oxidative damage in roots. The upregulation of ST in P5CR vs. P5R and the downregulation of ST in P3CR vs. P5CR agreed with the findings that increased pH-mediated amelioration of Cu toxicity involved reduced Cu uptake and oxidative damage in roots, and that low pH caused oxidative damage and a greater increase in Cu uptake in RCu300, but not in RCK [We detected upregulated ST in P5CR vs. P5R and downregulated ST in P3CR vs. P5CR . Qiu et a et al.  indicatet in RCK . HoweverCitrus sinensis (L.) Osbeck cv. Xuegan) seedlings were transplanted to 6 L pots (two seedlings per pot) containing sand, and then cultivated in a greenhouse under the natural conditions at Fujian Agriculture and Forestry University, Fuzhou with an annual average relative humidity, temperature, and sunlight of ~ 76%, 20 \u00b0C and 1600 h, respectively [2 \u00d7 pH 3.0 or 4.8 (adjusted by 1M HCl) until dripping (~500 mL per pot). Cu concentrations and pH levels of solutions were chosen based on our previous reports [2, and then stored in a \u221280 \u00b0C freezer until extraction of HRMs.Plant material culture and treatments referred to Zhang et al. . Six weeectively . After s reports . Each trhttps://www.metware.cn/, accessed on 1 June 2022) for assay of hormones. After frozen samples were ground into powder in liquid N2, 50 mg of the powder was transferred to 2 mL tube containing 1 mL of methanol:water:formic acid  and ten \u03bcL of internal standard mixed solution (100 ng mL\u22121). After 10 min of vortex, the mixture was centrifuged at 16,000\u00d7 g for 5 min at 4 \u00b0C. The yielding supernatant was evaporated to dryness in N2 flow, dissolved in 100 \u03bcL of 80% methanol (v/v), and then filtered through a 0.22 \u03bcm filter. The filtrate was used for HRMs assay.Equal amounts of frozen leaves (roots) from five seedlings from different pots were mixed as one biological replicate. There were three biological replicates per treatment. Samples were sent to Wuhan MetWare Biotechnology Co., Ltd.  [The concentrations of HRMs in the filtrate were determined using an ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) system  .p < 0.05 using DPS 7.05 . PCA was made with SPSS statistical software .Data were analyzed by two-way ANOVA (two (Cu levels) \u00d7 two (pH levels)), followed by the least significant difference (LSD) at Citrus Cu toxicity, the mechanisms by which different hormones alleviate Cu toxicity are unclear and require further research.Our results showed that elevated pH prevented the alterations of HRMs levels caused by Cu toxicity, while Cu toxicity aggravated the changes in HRMs levels caused by low pH. A model for the role of hormones in elevated pH-mediated mitigation of Cu toxicity in leaves and roots was proposed . Elevate"}
+{"text": "Infection caused by Streptococcus pneumoniae, mainly invasive pneumococcal disease (IPD) and pneumococcal pneumonia (PP), are a major public health problem worldwide. This study investigated population-based incidence and risk of PP among Catalonian persons\u2009\u2265\u200950 years-old with and without specific underlying conditions/comorbidities, examining the influence of single and multi-comorbidities in the risk of suffering PP.Population-based cohort study involving 2,059,645 persons\u2009\u2265\u200950 years-old in Catalonia, Spain, who were retrospectively followed between 01/01/2017-31/12/2018. The Catalonian information system for development of research in primary care (SIDIAP) was used to establish baseline characteristics of the cohort (comorbidities/underlying conditions), and PP cases were collected from discharge codes (ICD-10: J13) of the 68 referral Catalonian hospitals.Global incidence rate (IR) was 90.7 PP cases per 100,000 person-years, with a 7.6% (272/3592) case-fatality rate (CFR). Maximum IRs emerged among persons with history of previous IPD or all-cause pneumonia, followed by haematological neoplasia (475.0), HIV-infection (423.7), renal disease (384.9), chronic respiratory disease (314.7), liver disease (232.5), heart disease (221.4), alcoholism (204.8), solid cancer (186.2) and diabetes (159.6). IRs were 42.1, 89.9, 201.1, 350.9, 594.3 and 761.2 in persons with 0, 1, 2, 3, 4 and \u2265\u20095 comorbidities, respectively. In multivariable analyses, HIV-infection , prior all-cause pneumonia , haematological neoplasia , chronic respiratory disease  and prior IPD  were major predictors for PP.Apart of increasing age and immunocompromising conditions , history of prior IPD/pneumonia, presence of chronic pulmonary/respiratory disease and/or co-existing multi-comorbidity  are major risk factors for PP in adults, with an excess risk near to immunocompromised subjects. Redefining risk categories for PP, including all the above-mentioned conditions into the high-risk category, could be necessary to improve prevention strategies in middle-aged and older adults. Young children, individuals with at-risk or immunocompromising conditions and elderly people support the greatest burden of pneumococcal disease, with higher incidence and mortality in these persons  registered in the electronic primary care medical records of each cohort member at baseline:Chronic pulmonary/respiratory disease: it included chronic bronchitis/emphysema (J41-J44), asthma (J45-J46) and/or other chronic pulmonary diseases .Chronic heart disease: it included congestive heart failure (I50), coronary artery disease  and/or other chronic heart diseases .Diabetes mellitus (E10-E14).Chronic liver disease: it included chronic viral hepatitis (B18), cirrhosis (K74) and/or alcoholic hepatitis (K70)).Alcoholism .Smoking (F17).Anatomic or functional asplenia .Primary immunodeficiency (D80-D84).HIV infection (B20-B24).Chronic renal disease: it included nephrotic syndrome  and severe chronic renal failure (N18-N19 with glomerular filtration rate\u2009\u2264\u200930 ml/min).Cancer: it included solid organ or haematological neoplasia (C00 to C97) diagnosed within previous 5 years.Immunosuppressive therapy: it included long-term immunosuppressive medication and/or radiotherapy in the previous 12 months (coded according to specific SIDIAP codes)."}
+{"text": "Outpatient parenteral antimicrobial therapy (OPAT) is administering intravenous antibiotics in the outpatient setting. Herein we wish to understand the Atlanta VAMC (AVAMC) home OPAT population, infection details, and risk factors for hospital readmission, adverse drug events (ADEs), and clinical failure.Medical records of AVAMC veterans receiving home OPAT  January 1, 2019 and June 30, 2022 were reviewed. Data collected included: demographic and co-morbidity data, OPAT indication/infection type, microbiological information, presence of surgical debridement, and antimicrobial selection and duration of therapy.Clostridioides difficile infection. Other information include modification of OPAT plan (drug or duration), patient follow-up visit occurrence, re-hospitalization, and clinical resolution of infection. Univariate regression analyses were conducted on a subset of patients with musculoskeletal (MSK) infections .Outcomes included ADEs such as venous access events, laboratory abnormalities, and S. aureas was the predominant bacteria isolated (20%). See included table for full details.A total of 149 veterans were reviewed, with median age 66 years, 92% male, and 44% white race. Median CCI score was 5, and 80/149 (54%) patients had diabetes. The majority of infections were bone infections (30%), followed by diabetic foot osteomyelitis (24%). Methicillin-sensitive In the full cohort, 100/149(68%) had clinical resolution at 6 months and 20/149 (13%) had ADE during therapy. Rehospitalization occurred in 29 (20%) pts. Diabetes was associated with increased risk of failure -- Risk Ratio (RR) 1.68 (95% CI 1.01-2.77). Patients with clinic follow-up had a RR of 0.39 (95% CI: 0.21-0.73) for rehospitalization versus pts without follow-up.For pts with diabetes in the MSK group (N=97), risk of failure was 1.50x higher than those without diabetes (95% CI: 0.67-3.34).The RR for rehospitalization was 0.23 (95% CI: 0.06-0.82) for patients who received follow-up compared to patients who did not receive follow-up.AVAMC OPAT population demographic data, N = 149The AVAMC OPAT cohort is an older group with largely MSK-related infections and MSSA. Few ADEs occurred, and clinical failure remained high.All Authors: No reported disclosures"}
+{"text": "GRT-R910 , a self-amplifying mRNA (samRNA) vaccine expressing the spike protein plus T-cell epitopes of SARS-CoV-2 (D614G variant), was tested in a phase 1 study as a booster in healthy adults.This phase 1 open-label, dose escalation study enrolled healthy adults who previously received an approved mRNA COVID-19 vaccine series. Groups of 10 adults aged 18-60 years were boosted with GRT-R910 at 3 or 6 mcg. Adults > 60 years were boosted with GRT-R910 at 3, 6, or 10 mcg. All participants > 60 years in the 6 and 10 mcg dose groups received prior mRNA COVID-19 boosters; none in other groups had prior boosts. Study boosts occurred at least 112 days after completion of primary series/boost of authorized mRNA COVID-19 vaccine or prior SARS-CoV-2 infection. Solicited local and systemic reactogenicity events were collected for 7 days, unsolicited adverse events for 28 days, and serious adverse events (SAEs) for 366 days post-vaccination. Humoral  are being assessed at multiple time points over 1 year after study vaccination. Participants who self-reported SARS-CohV-2 infection or receipt of non-study COVID-19 booster during the study were censored in the immunogenicity analyses.No severe local reactogenicity events were observed. Overall, out of 48 enrolled across all groups, 8 reported at least 1 severe systemic reactogenicity event (figure 1). Most severe systemic reactogenicity events were transient, with most graded severe for 1 day or less. No SAEs have been reported. Neutralizing antibody responses remain durable up to 1 year after 3 and 6 mcg boosts in adults 18-60 years (figure 2) and up to 6 months after 3, 6, and 10 mcg boosts in adults > 60 years .GMT= geometric mean titer; Boxes and horizontal bars denote interquartile range (IQR) and median AUC, respectively. Whisker endpoints are equal to the maximum and minimum values below or above the median +/- 1.5 x IQR. All participants in the 3 \u03bcg GRT-R910, 18-60 yo and 6 \u03bcg GRT-R910, 18-60 yo groups received a mRNA two dose primary COVID-19 vaccination series prior to enrollment. None were previously SARS-CoV-2 infected.GMT= geometric mean titer; Boxes and horizontal bars denote interquartile range (IQR) and median AUC, respectively. Whisker endpoints are equal to the maximum and minimum values below or above the median +/- 1.5 x IQR. All participants in the 3 \u03bcg GRT-R910, > 60 yo group received a mRNA two dose primary COVID-19 vaccination series prior to enrollment. All participants in the 6 \u03bcg GRT-R910, > 60 yo and 10 \u03bcg GRT-R910, > 60 yo groups received a mRNA two dose primary COVID-19 vaccination series plus a mRNA booster vaccine prior to enrollment. Two participants each in the 6 \u03bcg GRT-R910, > 60 yo and 10 \u03bcg GRT-R910, > 60 yo groups had a previous COVID-19 infection at enrollment (green dots). D1, D15, D29 testing for Groups 9 and 10 are planned against BA.4/5.While transient systemic reactogenicity with GRT-R910 as a booster was observed, no safety signals were identified. Preliminary immunogenicity data demonstrate durable neutralizing antibody responses for 6-12 months in both younger and older age groups. Forthcoming T cell response data will aid in assessing the immunogenicity of this novel vaccine.Nadine Rouphael, MD, Icon, EMMES, Sanofi, Seqirus, Moderna: Advisor/Consultant Anna Wald, MD, MPH, Aicuris: Advisor/Consultant|Bayer: Advisor/Consultant|Curevo: Participation on Data Safety Monitoring Board|GSK: Grant/Research Support|Sanofi: Grant/Research Support|UpToDate: Royalties or licenses Pedro Garbes, MD, Gritstone bio, Inc.: Employee|Gritstone bio, Inc.: Employee|Gritstone bio, Inc.: Stocks/Bonds|Gritstone bio, Inc.: Stocks/Bonds Karin Jooss, PhD, Gritstone bio: employee|Gritstone bio: Stocks/Bonds Andrew Allen, MD, PhD, Gritstone bio: Board Member|Gritstone bio: Ownership Interest|Gritstone bio: Stocks/Bonds Amanda Eaton, MBA, Moderna: Grant/Research Support David M. Koelle, MD, Curevo Vaccines: Board Member|MaxHealth LLC: Board Member|Sanofi Pasteur: Grant/Research Support Daniel F. Hoft, MD, PhD, Moderna: Advisor/Consultant|Poolbeg: Advisor/Consultant"}
+{"text": "Coccidioidomycosis is an endemic mycosis in the southwestern United States. While most infections are mild or asymptomatic, severe cases are associated with high morbidity and mortality. We aimed to evaluate the clinical outcomes of patients admitted to the intensive care unit (ICU) with proven culture-positive coccidioidomycosis.Coccidioides spp. culture within a multi-healthcare system between 10/01/2017 \u2013 07/01/2022. Clinical information was collected by chart review.After institutional review board approval, we retrospectively included ICU patients with positive Coccidioides spp. culture, 145 patients required ICU stay. The median age was 51 (32-62), with 100 (68.9%) male, 56 (38.6%) White non-Hispanics, and 25 (17.2%) Blacks. Fifty-seven (39.3%) received azole antifungals before hospitalization. Seventy-two (49.7%) had another fungal, bacterial, or viral infection during their hospitalization, and 41 (28.3%) had extrapulmonary coccidioidomycosis. The majority, 131 (90.3%), received antifungal therapy during their hospital stay. The median hospital length of stay was 18 days (10-37). Almost half (48.3%) died during the study period, with liver cirrhosis and age ( >60 years) resulting in statistically significant mortality, 12/13 (92.3%), p-value < 0.001, and 31/43 (72.1%) p-value =0.001, respectively. Mechanical ventilation and/or intravenous vasopressor support was associated with a high risk of mortality 66/113 . Failure to receive antifungal therapy before ICU admission was associated with increased mortality, 46/70 .Of the 392 hospital encounters with positive Patients with coccidioidomycosis admitted to the ICU face an increased risk of mortality. Risk factors associated with ICU mortality include advanced age, cirrhosis, and delay in the administration of antifungal therapy. Future studies are needed to evaluate these risks further.Tirdad Zangeneh, DO, AiCuris: Grant/Research Support Mohanad Al-Obaidi, MD, MPH, La Jolla Pharmaceuticals: Honoraria"}
+{"text": "Immunotherapeutic targets in multiple myeloma (MM) have variable expression height and are partly expressed in subfractions of patients only. With increasing numbers of available compounds, strategies for appropriate choice of targets (combinations) are warranted. Simultaneously, risk assessment is advisable as patient\u2019s life expectancy varies between months and decades.We first assess feasibility of RNA-sequencing in a multicenter trial . Next, we use a clinical routine cohort of untreated symptomatic myeloma patients undergoing autologous stem cell transplantation  64 months) to perform RNA-sequencing, gene expression profiling (GEP), and iFISH by ten-probe panel on CD138-purified malignant plasma cells. We subsequently compare target expression to plasma cell precursors, MGUS (n=59), asymptomatic (n=142) and relapsed (n=69) myeloma patients, myeloma cell lines (n=26), and between longitudinal samples (MM vs. relapsed MM). Data are validated using the independent MMRF CoMMpass-cohort .de novo (LfM-HRS) defined risk scores. LfM-HRS delineates three groups of 40%, 38%, and 22% of patients with 5-year and 12-year survival rates of 84% (49%), 67% (18%), and 32% (0%). R-ISS and RNA-sequencing identify partially overlapping patient populations, with R-ISS missing, e.g., 30% (22/72) of highly proliferative myeloma.RNA-sequencing is feasible in 90.8% of patients (GMMG-MM5). Actionable immune-oncological targets (n=19) can be divided in those expressed in all normal and >99% of MM-patients , those with expression loss in subfractions of MM-patients , aberrantly expressed in MM , and resistance-conveying target-mutations e.g., against part but not all BCMA-directed treatments. Risk is assessable regarding proliferation, translated GEP-  and RNA-sequencing based assessment of risk and targets for first choice treatment is possible in clinical routine. Clinical signs and symptoms relate to displacement of normal hematopoiesis, generation of osteolytic bone disease, and renal impairment . TreatmeMM is treated by combination treatment whenever possible , 13: effThe situation is different for immune-oncological therapies in development or recently approved. Bispecific antibodies or CAR-T cells against e.g., BCMA, GPRC5D, or FCR5H show single agent remission rates of 60-80% , 30\u201338. Presence or absence and height of target expression is an evident selection criterion for treatment choice, as shown, e.g., for CD38  and GPRCin situ hybridization [iFISH])  incorporating clinical factors  and molecular alterations in malignant plasma cells , t assessed by interphase fluorescence [iFISH]) . Innovat[iFISH])  or \u201cscor[iFISH]) \u201352.de novo risk score by RNA-sequencing (termed LfM-HRS). Findings are validated in the independent MMRF CoMMpass-cohort (n=767 patients).In this manuscript, we first assess applicability of RNA-sequencing in the GMMG-MM5 multicenter phase III clinical trial setting (604 patients) based on our low-input RNA-sequencing protocol . Secondl22.16 cells (Patients (n=604) were included in the prospective GMMG MM5-trial , 54 betw6 cells .iFISH using cytospins from CD138-purified plasma cells was performed centrally . Data could be obtained for 556/573 patients with available bone marrow aspirates (97%) and 556/559 patients with available CD138-purified plasma cells, respectively (99.5%). The median proportion of malignant plasma cells determined per iFISH, i.e., the highest percentage of a chromosomal aberration, was 95% .Samples for RNA-extraction followed by quality control were collected over two weeks and subjected to GEP by DNA-microarrays. In total, n=458 transcriptome datasets are available, i.e., 81.9% of patients with available CD138-purified plasma cells. Of these, two patients were excluded from further analysis for not fulfilling the trial\u2019s inclusion criteria. Gene expression profiling could not be performed in 53 cases due to low RNA quality (9.5%) and further 48 cases (8.6%) in which not enough RNA was available .Using our standardized RNA-sequencing protocol  , asymptomatic , symptomatic, therapy-requiring myeloma , relapsed myeloma , as well as healthy donors (n=10) as comparators were included in the study approved by the ethics committee  after written informed consent. For patient characteristics, see + memory B-cells (n=4) were FACS-sorted and polyclonal plasmablasts (n=4) were in vitro differentiated as described  and myeloma cells were purified using anti-CD138 microbeads . Peripheral blood CD27escribed . A totalRNA-sequencing data of 52/142/535/69 patients with MGUS/AMM/MM/MMR and 26 HMCLs were used. GEP-data of 534 patients with MM were used for score definition and validation .2.3Independent validation of risk assessment and target identification was performed using the Multiple Myeloma Research Foundation (MMRF) CoMMpass trial (NCT01454297), i.e., n=767 previously untreated myeloma patients with RNA-sequencing data available (release 13).2.42.4.1RNA was extracted using the Qiagen AllPrep DNA/RNA kit  according to the manufacturer\u2019s instructions. Quality control and quantification was performed using an Agilent 2100 bioanalyzer .2.4.2RNA-sequencing was performed as published . In brie2.4.3Gene expression profiling (GEP) using U133 2.0 plus arrays  was performed as published , 56\u201359. 2.5iFISH analysis was conducted on CD138-purified plasma cells using probes for chromosomes 1q21, 5p15, 5q31 or 5q35, 8p21, 9q34, 11q22.3 or 11q23, 13q14.3, 15q22, 17p13, 19q13, IgH-breakapart, as well as translocations t, t, and t according to the manufacturer\u2019s instructions  and data were analyzed as published .2.6https://www.r-project.org/) and Bioconductor (https://www.bioconductor.org/) versions 3.3.2 and 3.4 (score implementation) and versions 3.4.4 and 3.6 .GEP and RNA-seq analysis were performed as previously described , 51, 58 Overall (OS) and event-free (EFS) survival were investigated for symptomatic multiple myeloma undergoing high-dose therapy using Cox\u2019s proportional hazard model as published .Survival curves were computed with nonparametric survival estimates for censored data using the Kaplan-Meier method . DiffereEffects were considered statistically significant if the P-value of corresponding statistical tests was below 5%. For comparisons of parameters , or ADC, antibody-radio-conjugates, bispecific antibodies, or CART-products in trials .Based on the expression pattern, we divided targets in those expressed in normal and almost all (>99%) malignant plasma cell samples , those constitutively expressed in all normal plasma cells with expression loss in a subfraction of malignant plasma cells , and targets aberrantly expressed in malignant plasma cells, i.e., not expressed in normal bone marrow plasma cells . CD70 is expressed in half of BMPC decreasing to 24.5% in MM. Of assessed targets, CD1B is not expressed, CD25 at very low frequency and expression height only (2.9%). Results for expressed targets are summarized in RNA-sequencing allows detection of somatic variants and mutated transcripts; e.g., for BCMA, we found in 25% of patients coding non-synonymous single nucleotide variants present  with a median allele frequency of 1 in patients harboring the aberration. If present, a resistance conveying mutation would thus have been detected but expectedly could not, as none of the patients has been treated with BCMA-targeting agents prior to analysis.Targets expressed in all myeloma patients show high inter-patient variation, e.g., 6.8 (CD38), 6.9 (BCMA), 9.9 (GPRC5D), and 10.9 (FCRH5) log-fold variation . Of targets aberrantly expressed in malignant plasma cells, i.e., not expressed in BMPC , NY-ESO1 shows significantly higher expression from MGUS to MMR. Regarding a potential use of immune-oncological agents in early-stage myeloma, we assessed differences between AMM to MM to MMR, (instead of MGUS-AMM-MM-MMR) yielding similar results , BCMA, GPRC5D, FCRH5, TACI, CD74, CD44, CD37 and CD79B, seven show significant lower expression from MGUS to MMR , i.e., SLAMF7 (CS1), BCMA, GPRC5D, FCRH5, TACI, CD74, and CD37. If adjusted for multiple testing by Benjamini-Hochberg correction, significance is maintained for BCMA, FCRH5, TACI, CD74, and CD37. Targets with expression loss in a subfraction of malignant plasma cells of previously untreated patients  are all significantly lower expressed from MGUS to MMR , a change in expression of any of the investigated antigens could be found in 56 patients (88.9%) with both losses and gains occurring ; median OS was 35 vs. 86 vs. 130 months , we \u201ctranslated\u201d these. The RS-, UAMS70- and SKY92-score delineated a population of 10%, 19%, and 13% of high risk and 67% of medium risk (Rs-score) patients. High- (medium) risk patients showed significantly inferior EFS and OS, i.e. the RS-score - RSGEP and RNA-sequencing based scores showed a concordance of 71.7% - 92.5% regarding patients identified as high risk (3.4de novo generated a RNA-sequencing based proliferation index (RPI). In comparison to normal bone marrow plasma cells or non-proliferating memory B-cells, malignant plasma cells showed a significant and stage-dependent increase from early disease MGUS vs. asymptomatic vs. symptomatic, therapy-requiring multiple myeloma , to 2nd (61%), 3rd 38%, 4th(15%) and 5th line (1%). Furthermore, use of \u03b3-secretase inhibitors  . This first relates evidently to targets not expressed in all myeloma patients, as being either lost in a subfraction of malignant plasma cells, i.e., BAFF-R, CD19, CD20, and CD22, or gained, i.e., NY-ESO1/2, MUC1, and CD30. Secondly, it relates to high inter-patient variation of target expression. Considering targets expressed in all myeloma patients, 10.9 log-fold (FCRH5) differences were found. With reported relation of expression height and response for CD38  and GPRCacestat)  in case acestat) . It exemdifferent BCMA-targeting agents. As downregulations and mutations discussed here occur primarily under selection pressure of the respective treatment and, in part, different aberrations occur in a subclonal manner . Genes with stage-dependent loss of expression, like CD19, CD22, and BAFF-R, showed high dynamics with both gains and losses of expression occurring. The cancer testis antigens MUC1 and NY-ESO1/2 were predominantly gained in MMR. These findings are in line with a subclonal architecture in myeloma leading to clonal tides  and spat4.3Introduced in myeloma research in 2002  and 2011\u00ae, Signal Genetics\u2122   , suggestart.org)  or the Mart.org) . But thea priori evident.Immediate usefulness would be perceived if either response could be predicted, or targets selected in for personalized treatment. Despite factors associated with response have been identified , 25, it GEP/NGS-based approaches would be significantly fostered by the use of appropriated clinical trial designs, especially for regulatory and approval purposes , 95 espe5RNA-sequencing is applicable in 90% of patients (comparable to iFISH), is of overall equal predictive power as the current gold standard R-ISS, but identifies a further fraction of myeloma patients, e.g., with highly proliferative myeloma cells. It allows personalized target identification for immune-oncological drugs based on presence and height of expression. RNA-sequencing used for \u201ceducated first guess\u201d could be considered as \u201cIOnc drug and risk advisor\u201d in analogy to other decision tools.https://www.ebi.ac.uk/arrayexpress/, E-MTAB-4715; https://www.ebi.ac.uk/arrayexpress/, E-MTAB-4717; https://www.ebi.ac.uk/arrayexpress/, E-MTAB-5212; https://www.ebi.ac.uk/arrayexpress/, E-TABM-937; https://www.ebi.ac.uk/arrayexpress/, E-TABM-1088; https://www.ebi.ac.uk/ena, PRJEB37100; https://www.ebi.ac.uk/ena, PRJEB36223.The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: Patients were included in the study approved by the ethics committee of the medical faculty of the Ruprecht Karls University Heidelberg, Germany,  after written informed consent. The studies were conducted in accordance with the local legislation and institutional requirements. The participants provided their written informed consent to participate in this study.ME-R: Conceptualization, Formal Analysis, Investigation, Methodology, Validation, Writing \u2013 review & editing. SB: Formal Analysis, Writing \u2013 review & editing. VB: Writing \u2013 review & editing, Investigation, Methodology. HS: Investigation, Writing \u2013 review & editing. UB: Investigation, Writing \u2013 review & editing. CS: Investigation, Writing \u2013 review & editing. MH: Investigation, Writing \u2013 review & editing. KW: Investigation, Writing \u2013 review & editing. TH: Investigation, Writing \u2013 review & editing, Methodology. MR: Investigation, Writing \u2013 review & editing. HG: Investigation, Writing \u2013 review & editing, Funding acquisition. AJ: Investigation, Writing \u2013 review & editing. KM: Investigation, Writing \u2013 review & editing. EB: Investigation, Writing \u2013 review & editing. EM: Investigation, Writing \u2013 review & editing. KD: Investigation, Writing \u2013 review & editing. JM: Investigation, Writing \u2013 review & editing. KV: Investigation, Writing \u2013 review & editing. AS: Conceptualization, Formal Analysis, Funding acquisition, Investigation, Supervision, Writing \u2013 original draft. DH: Conceptualization, Formal Analysis, Funding acquisition, Investigation, Methodology, Supervision, Validation, Writing \u2013 original draft."}
+{"text": "Bloodstream infections (BSIs) are major causes of morbidity and mortality for which prior analyses and validated scoring tools have attempted to stratify risk. This study investigated institutional and epidemiologic predictors of mortality with Gram-negative BSIs to better identify targeted antimicrobial stewardship efforts to improve outcomes.All patients with aerobic, monomicrobial, Gram-negative BSIs admitted and discharged from our academic medical institution between July 2022 and March 2023 were included. Data collected included patient demographics, microbiological data, clinical management, length of stay, ID consultation, and clinical outcomes . Univariate and multivariable logistic regression models were performed to identify variables that were primary drivers for inpatient mortality.E. coli (37.8%), K. pneumoniae (11.2%), P. aeruginosa (10.0%), S. marcescens (8.5%), and E. cloacae (6.2%). The majority (65.3%) were community acquired infections with 41.7% in the ICU, and 46.7% received an infectious diseases consult. The overall mortality was 14.7% with the highest mortality due to P. aeruginosa (34.6%) followed by S. marcescens (27.3%), K. pneumoniae (13.8%), E. cloacae (12.5%), and E. coli (8.2%). Overall mortality was significantly associated with several parameters in a univariate analysis (Table 1). In multivariable regression analysis hospital-acquired infection (0=0.002), cancer (p=0.012), qPitt > 2 (p< 0.001), SBP< 100 or vasopressor use at 72-96 hrs after positive blood culture (p=0.017) was associated with increased risk of mortality. The only parameter to reduce mortality within the multivariable regression analysis, by more than two-fold, was ID consultation (p=0.022).Overall, 259 patients  were identified. The most common organisms were Parameters Associated with Mortality in Univariate AnalysisThe only factor found to significantly reduce mortality was formal ID consultation. ID consultation should be considered part of routine care for all patients with Gram-negative bacteremia.Katie B. Olney, PharmD, BCIDP, The Society of Infectious Diseases Pharmacists (SIDP): Grant/Research Support"}
+{"text": "Prior literature establishes bidirectional associations between suicide and substance use disorders (SUDs), particularly opioid use disorder (OUD). However, the context of mental health services utilization remains under-investigated. This analysis examined patterns of mental health services utilization in patients with SUDs and suicidality, identified associated risk factors, and evaluated the impact of patient engagement on subsequent mental health outcomesSee above.Electronic health records (EHRs) derived from 7 health systems across New York City between 2010-2019 were analyzed. Suicidality was identified as any ICD-9/10 diagnosis of suicide attempt, suicidal ideation, or self-harm injury. SUDs were identified as any opioid, cannabis, cocaine, hallucinogen, inhalant, sedative/hypnotic/anxiolytic, amphetamine, or other substance abuse or dependence. Quasi-Poisson regression adjusted for age, gender, and chronic diseases was used to model associations between OUD exposure and the frequency of encounters and estimate the relative risk (RR) of significant covariates.A total of 6977 adults with suicidality and any comorbid SUD were selected, including 2203 (31.6%) with a diagnosis of OUD and 4774 (68.4%) without a diagnosis of OUD. Most patients were male (54.8%) and aged between 25-64 years (79.3%). Many (61.3%) had over 3 chronic diseases, including depression (80.8%), hypertension (60.6%), anemia (43.0%), and hyperlipidemia (41.9%). Compared to patients with other SUDs, those with OUD had higher odds of self-harm injury [OR: 1.26 (95% CI: 1.13-1.41)], depressive disorders [1.47 (1.29-1.67)], anxiety disorders [1.65 (1.48-1.84)], psychotic disorders [1.23 (1.11-1.37)], personality disorders [1.30 (1.16-1.48)], and post-traumatic stress disorder [1.37 (1.20-1.57)]. Patients with OUD were more likely to utilize all-cause outpatient (RR: 1.16), emergency department (ED) (RR: 1.43), and inpatient (RR: 1.60) services (p<0.001). Among OUD patients, males were less likely to have outpatient visits (RR: 0.79) and inpatient hospitalizations (RR: 0.88), and older age was protective against ED admissions (RR range: 0.62-0.71). Additionally, individuals with OUD were more likely than those with other SUDs to have SUD-related encounters, as well as suicide-related ED admissions and inpatient hospitalizations (p<0.0001). Those who had more mental health outpatient visits were less likely to have suicide-related ED admissions (RR: 0.85), however this association was weaker among younger or male patients with comorbid OUD.Among suicidal adults with comorbid SUDs, those with a diagnosis of OUD were more likely to utilize mental health services and have psychiatric comorbidity. Males and older adults were less likely to utilize services.None Declared"}
+{"text": "Serum anti-spike IgG and neutralizing antibody against SARS-CoV-2 are recognized as immune correlates of protection (ICP) for COVID-19. However, there are limited data available on the ICP after the introduction of BA.4/5 bivalent vaccine. Thus, our study aimed to investigate the antibody response as correlates of protection against COVID-19, and the effects of BA.4/5 bivalent booster vaccine.A prospective cohort of healthcare workers was enrolled at Asan Medical Center, a 2,700-bed tertiary care hospital in South Korea between December 2022 and January 2023. Study participants had received a booster vaccination and either planned to receive a bivalent BA.4/5 vaccine or not. Blood samples were collected from study participants. Immunological assessments were performed using ELISA to measure variant-specific SARS-CoV-2 S1-IgG antibodies, and virus reduction neutralization test (VRNT) to measure neutralizing antibody levels.A total of 483 participants were enrolled, of whom 45 (9.3%) underwent subsequent infection after study enroll, while 438 (90.7%) did not. Of these 483 participants, 166 (34.4%) received the BA.4/5 bivalent booster vaccination. There was a significant difference in Wuhan-Hu-1 S1-IgG, BA.5 S1-IgG and neutralizing antibody against BA.5 levels between individuals with and without subsequent infection . While Wuhan S1-IgG and BA.5 S1-IgG (both P < 0.001) were found to be independent protective factors against subsequent SARS-CoV-2 infection, neutralizing antibody against BA.5 did not show significant protective effects. Hybrid immunity was robust predictive factor for subsequent infection in all analysis based on each immune marker . The predictive sensitivity of neutralizing antibody for subsequent infection was improved from 80.0%  to 90.0%  when combined with hybrid immunity and 100%  with bivalent vaccine.Taken together, humoral immune responses seem to be reliable markers for immune correlates of protection against SARS-CoV-2 infection, and hybrid immunity showed independent protective effect from subsequent infection.All Authors: No reported disclosures"}
+{"text": "Globally 20-50% population suffer from periodontal diseases. Bidirectional link exists between periodontal diseases and comorbidities. Limited evidence is available on antibiotic effectiveness in patients of dental infections with comorbidities.This retrospective, multi-centric, observational electronic medical record study gathered real world Indian evidence to assess effectiveness of cephalexin-clavulanic acid  fixed-dose combination (FDC) compared to amoxicillin-clavulanic acid (amoxicillin CV) FDC and cefuroxime in adults with dental infections. Following data represents effectiveness in subset of patients with and without comorbidity.Overall, 89% (316/355) patients were without comorbidity ; while 11% (39/355) patients had comorbidities .Time to overall clinical improvement was less in patients without comorbidities compared to those with comorbidities .In patients without comorbidity, 98.13% (157/160), 100% (70/70), and 97.67% (84/86) showed overall clinical improvement within 10 days, in cephalexin CV, cefuroxime, amoxicillin CV groups, respectively.In patients with any comorbidity, 100% (14/14) patients showed improvement in cephalexin CV against 94% (16/17) in cefuroxime and 87% (7/8) in amoxicillin CV group within 10 days (p >0.05).Overall clinical improvement in patients with and without comorbidities on Day 5 and 7 is presented in Table. In metabolic and cardiovascular disorders subsets, 100% patients showed improvement within 10 days in cephalexin CV group, while 92% (12/13) and 100% (5/5), respectively in cefuroxime group; and 88% (7/8) and 67% (2/3), respectively in amoxicillin CV (p >0.05).In patients with dental infections, presence of comorbidities delays overall clinical improvement. All patients with comorbidity treated with cephalexin CV showed overall clinical improvement within 10 days of treatment. No significant difference was observed between treatments for overall clinical improvement.Archana Karadkhele, MBA, Sun Pharma Laboratories Limited: Full time employee Shruti Dharmadhikari, MSc, Sun Pharma Laboratories Limited: Full time employee Chintan Khandhedia, MD, Sun Pharma Laboratories Limited: Full time employee Neeraj Markandeywar, MD, Sun Pharma Laboratories Limited: Full time employee Amey Mane, MD, Sun Pharma Laboratories Limited: Full time employee Suyog Mehta, MD, Sun Pharma Laboratories Limited: Full time employee"}
+{"text": "Pseudomonas aeruginosa high-risk clones pose severe threats to public health. Here, we characterize the imipenem/relebactam (IR) resistance mechanisms in P. aeruginosa high-risk clones sequence type 235 (ST235) and ST463 in China. Minimum inhibitory concentrations (MICs) were determined, and Illumina short-read sequencing was performed for 1,168 clinical carbapenem-resistant P. aeruginosa (CRPA) isolates. The gene copy number and expression level were analyzed by Illumina sequencing depth and reverse transcription-quantitative PCR, respectively. Resistance conferred by blaGES-5 was evaluated by cloning experiments. ST463 and ST235 accounted for 9.8%  and 4.5%  of total isolates, respectively, and showed high frequencies of extensively drug-resistant and difficult-to-treat resistant phenotypes. The overall IR-resistant rate in CRPA was 21.0% . However, the IR resistance rate was 81.7% (94/115) in ST463-PA and 52.8% (28/53) in ST235-PA. Of the ST463 isolates, 92.2% (106/115) were Klebsiella pneumoniae carbapenemase-producing P. aeruginosa (KPC-PA), and all 94 IR-resistant ST463-PA produced KPC-2. Compared to IR-susceptible ST463 KPC-2-PA, IR-resistant ST463 KPC-2-PA exhibited significantly higher blaKPC-2 copy numbers and expression levels. In ST463 KPC-2-PA, 16 mg/L relebactam resulted in additional fourfold reductions in imipenem MIC50/90 values compared to 4 mg/L relebactam. In ST235, 1.9% (1/53) carried blaIMP carbapenemase and 54.7% (29/53) carried blaGES carbapenemase. Other than the IMP producer, all 27 IR-resistant ST235-PA produced GES-5. Cloning experiments revealed that imipenem resistance in blaGES-5-carrying PAO1 transformants was generally unaffected by relebactam. In conclusion, IR-resistant CRPA isolates in China were mainly distributed in P. aeruginosa high-risk clones ST463 and ST235. The major underlying IR resistance mechanisms were blaKPC-2 overexpression and blaGES-5 carriage. Pseudomonas aeruginosa (CRPA) has emerged as an urgent public health issue. Although P. aeruginosa isolates usually exhibit great clonal diversity, the dissemination of certain high-risk clones is an essential step for CRPA to maintain an antibiotic-resistant phenotype. Among the high-risk clones detected globally, sequence type 235 (ST235) has been identified across five continents and is considered the most widespread  5. The hP. aeruginosa. In a multicenter study involving 4,927 MDR-PA isolates from adult patients, although 13.5% of samples produced metallo-\u03b2-lactamases (MBLs), nearly 70% of isolates were still susceptible to IR (Imipenem/relebactam (IR), a novel \u03b2-lactam-\u03b2-lactamase inhibitor (BLBLI) combination, has been developed and applied clinically to treat CRPA infections. Relebactam exerts strong inhibitory effects on many class A and class C \u03b2-lactamases and is almost unaffected by OprD inactivation or efflux pump extrusion , 10. Conle to IR .in vitro activity of IR against P. aeruginosa, the IR resistance mechanisms in CRPA high-risk clones remain largely unexplored. In this study, we evaluated the IR resistance profiles in clinical CRPA isolates from China, with a particular focus on the IR activity against ST235 and ST463 CRPA strains. Notably, we identified a surprisingly high IR resistance rate in ST235 and ST463 CRPA isolates. The underlying IR resistance mechanisms were further explored in detail.Currently, although many studies emphasize the A total of 1,168 clinical CRPA isolates, defined as resistant to either meropenem or imipenem according to the Clinical and Laboratory Standards Institute (CLSI) breakpoints , were inEscherichia coli ATCC 25922, P. aeruginosa ATCC 27853, and Klebsiella pneumoniae ATCC 700603 and ATCC BAA-1705 were used as quality controls. For all antimicrobials except fosfomycin, CLSI breakpoints  phenotype were defined according to the criteria established by Magiorakos et al. and Tamma et al. respectively  of amikacin, imipenem, meropenem, IR (relebactam fixed at 4 mg/L), ceftazidime, cefepime, ciprofloxacin, levofloxacin, piperacillin/tazobactam, aztreonam, fosfomycin, and polymyxin B were determined using the broth microdilution method with Thermo Scientific Sensititre System. To facilitate comparison between different antimicrobials in ST463 KPC-2-producing and ST235 GES-5-producing strains, the MICs of imipenem and IR (relebactam fixed at 4 mg/L or 16 mg/L) were simultaneously obtained by the agar dilution method described in CLSI standard M07 . Escheriakpoints  for P. aakpoints  for Enteectively , 15.De novo assembly was accomplished using shovill version 0.9.0 (https://github.com/tseemann/shovill). Multilocus sequence typing (MLST) was performed, and antimicrobial resistance genes were detected using mlst version 2.19.0 (https://github.com/tseemann/mlst) and ABRicate version 1.0.0 (https://github.com/tseemann/abricate). The oprD gene of clinical strains was compared with that of P. aeruginosa PAO1. OprD inactivation was confirmed if any of the following criteria were met: (1) truncation or absence of the oprD gene; (2) mutations leading to an OprD frameshift; and (3) mutations leading to the creation of a premature stop codon in OprD.The genomic DNA of all CRPA isolates was extracted using QIAamp DNA Mini Kit  and underwent short-read sequencing . blaKPC-2 copy number was further calculated based on WGS. For each sample, a SAM alignment file was obtained by mapping Illumina raw reads to references  using bowtie2 version 2.2.5 (https://github.com/BenLangmead/bowtie2) with the option \u201c-I 0 -X 800 --fast -q.\u201d After transforming the alignment results into BAM files, the sequencing depths of the blaKPC-2 gene and seven MLST alleles were analyzed by BEDTools version 2.30.0 (https://github.com/arq5x/bedtools2) with the option \u201cbedtools coverage -mean.\u201d Finally, the blaKPC-2 copy number of each strain was calculated as the depth of blaKPC-2 divided by the average depth of the seven MLST alleles.For KPC-2-producing strains, the P. aeruginosa isolates, genome assemblies were first annotated using Prokka version 1.14.6 (https://github.com/tseemann/prokka). Core gene alignment was then obtained by pangenome analysis using Panaroo version 1.2.9 . Reverse transcription was conducted using PrimeScript RT Reagent Kit . Quantitative PCR was performed in biological triplicates using TB Green Premix ExTaq . Each biological triplicate was assayed in technological triplicates. The housekeeping gene rpoD was chosen as the endogenous reference for normalization. The primers used for quantitative PCR are listed in Table S1.To compare the expression levels of blaGES-5 and blaGES-15 genes with the same upstream promoter sequences were amplified from clinical strains R16-15 and R20-99, respectively. The cloned sequences were homologously recombined into the pGK1900 plasmid. The constructed plasmids were then transformed into both E. coli DH5\u03b1 and P. aeruginosa PAO1. Transformants verified by Sanger sequencing were eventually selected for antimicrobial susceptibility testing. The primers used for cloning experiments are listed in Table S1.The P < 0.05.Statistical analysis was performed using the Fisher\u2019s exact test, the Mann\u2012Whitney U test, and the Kruskal\u2012Wallis test in SPSS version 25. The results were considered statistically significant when P. aeruginosa from other STs. The frequency of XDR/PDR phenotype was 88.7% (102/115) for ST463-PA and 73.6% (39/53) for ST235-PA ; the frequency of DTR phenotype was 90.4% (104/115) for ST463-PA and 52.8% (28/53) for ST235-PA . In contrast, the proportions of XDR/PDR-PA and DTR-PA in CRPA isolates from other STs were only 24.4%  and 24.5% , respectively. Therefore, the resistance status of ST463-PA and ST235-PA was significantly more severe.Among the 1,168 CRPA isolates included in this study, MLST analysis revealed 285 different STs (excluding 69 isolates belonging to unknown STs). In 21 out of these 285 STs, the CRPA strain numbers were more than nine . ST463  and ST235  were found to be the most prevalent STs. The resistance phenotypes of ST463-PA and ST235-PA to traditional antipseudomonal agents were compared with those of 50/90 of 16/>128 mg/L; however, only 245/1,168 (21.0%) isolates were resistant to IR with an MIC50/90 of 2/32 mg/L  isolates were resistant to imipenem with an MIC/32 mg/L . The add90 value ; Fig. 1.The phylogenetic tree was then constructed for IR-resistant isolates, and the clonal characteristics were analyzed. Among 245 IR-resistant isolates, 62 different STs were identified (excluding five strains with unknown STs). In 16 of these 62 STs, the IR-resistant strain numbers were more than one . As shown in P. aeruginosa (non-CPPA), while 106/115 (92.2%) isolates were KPC-producing P. aeruginosa (KPC-PA). The identified KPC alleles included blaKPC-2  and blaKPC-33 . In ST235-PA, in addition to 23/53 (43.4%) non-CPPA isolates, 54.7% (29/53) were carbapenemase-type GES-producing P. aeruginosa (GES-PA) and 1.9% (1/53) were IMP-15-producing P. aeruginosa. The carbapenemase-type GES alleles consisted of blaGES-5  and blaGES-15 .To further describe the molecular epidemiology and interpret the IR resistance data of ST463 and ST235 isolates, genomic analysis was subsequently conducted. As listed in Table S2, the strong association between class A carbapenemase types and STs could be clearly distinguished. In ST463-PA, 9/115 (7.8%) isolates were non-carbapenemase-producing blaKPC-33-carrying P. aeruginosa were IR intermediate, while 94/104 (90.4%) blaKPC-2-carrying P. aeruginosa were IR resistant  blaGES-5-carrying P. aeruginosa were IR resistant  esistant ; in ST23esistant . Overall50 and MIC90 values. However, with 4 mg/L relebactam, restoration of imipenem susceptibility was only achieved in 9/104 (8.7%) ST463 KPC-2-PA isolates, while the majority still exhibited high-level imipenem resistance (64 mg/L to 256 mg/L). Therefore, we proposed that 4 mg/L relebactam might be insufficient for inhibiting blaKPC-2 expression in ST463.IR resistance mechanisms were further explored in ST463 KPC-2 producers. As shown in 50 and MIC90 values were achieved by 16 mg/L relebactam  ST463 KPC-2-PA isolates became susceptible or intermediate to imipenem. Notably, with 16 mg/L relebactam, only three ST463 KPC-2 producers still exhibited high-level imipenem resistance (MIC: 64 mg/L). In comparison to 4 mg/L relebactam, additional fourfold reductions in imipenem MIClebactam .blaKPC-2 copy numbers were compared between ST463 KPC-2 producers showing different IR resistance phenotypes. Compared with ST463 KPC-2-PA isolates susceptible or intermediate to IR, IR-resistant ST463 KPC-2-PA isolates exhibited significantly higher blaKPC-2 copy numbers     . In stra 0.0001) . Besides < 0.01) . AltogetblaKPC-2 expression level was quantitively analyzed. In total, 18 blaKPC-2-carrying ST463 isolates were selected from three phenotypic groups . These strains shared similar genetic elements surrounding blaKPC-2 and the same blaKPC-2 promoter sequences. As shown in blaKPC-2 expression level differed significantly among these three groups , showing that blaKPC-2 overexpression was related to the attenuated inhibitory effects of 4 mg/L relebactam.Finally, to further verify our hypothesis, the relative mexE and mexY genes in ST463 KPC-2-PA, only the expression levels of mexA and mexD genes were analyzed. Interestingly, under 16 mg/L relebactam, compared with isolates susceptible to imipenem (MIC: 1 mg/L), ST463 KPC-2-PA isolates resistant to imipenem (MIC: 32 mg/L to 64 mg/L) displayed much lower mexA and mexD expression levels . This suggested that the IR resistance phenotype differences among ST463 KPC-2-PA could not be attributed to efflux pump overexpression. With regard to OprD inactivation, among imipenem-susceptible and imipenem-nonsusceptible ST463 KPC-2-PA (under 16 mg/L relebactam), the proportions of OprD-inactivating strains were 36.4% (4/11) and 96.8% (90/93), respectively (P < 0.0001). This significant difference indicated the potential contribution of OprD inactivation to imipenem resistance, which further led to IR resistance in ST463 KPC-2-PA.Although additional decreases in imipenem MICs were achieved when relebactam was increased from 4 mg/L to 16 mg/L, the majority  of ST463 KPC-2-PA were still not susceptible to imipenem. Previous studies have identified IR resistance associated with mutations in OprD and regulators of MexAB-OprM , 20. Thu50/MIC90 values or the cumulative inhibition curves of imipenem, suggesting that IR resistance in ST235 GES-5-PA could not be attributed to blaGES-5 overexpression.Mechanisms of IR resistance were also investigated in detail for ST235 GES-5-PA. As illustrated in blaGES allelic variants on imipenem and IR resistance were analyzed. The imipenem MICs were generally higher in ST235 GES-5 producers (32 mg/L to 128 mg/L) than in ST235 GES-15 producers (16 mg/L). The IR resistance rate was 100% (27/27) for blaGES-5-carrying ST235-PA isolates, with MIC50/MIC90 values of 32/32 mg/L. In contrast, all blaGES-15-harboring ST235-PA isolates were susceptible or intermediate to IR, exhibiting MIC50/MIC90 values of 4/4 mg/L. The above data demonstrated the potential involvement of blaGES-5 carriage in IR resistance.Subsequently, the impacts of blaGES-5 carriage  was acquired. In another in vitro study  were published. In one in vitro multistep selection study . Under these circumstances, clinical treatment choices would be greatly constrained.Generally, IR resistance related to KPC enzymes has seldom been reported. Until 2022, four studies investigating IR resistance development in KPC-producing on study , IR-resisistance \u201332, GaP. aeruginosa in previous studies . According to Hujer et al.\u2019s study (Although GES producers were found among IR-resistant  studies , 33, the\u2019s study , GES-20 exoU+ genotype and various resistance-related mutations. In a study on P. aeruginosa bacteremic pneumonia, a 29.8% increase in 5-day mortality and a 34.4% increase in 30-day mortality were noted in patients infected with ST235-PA (exoU and exoS genes, the ST463 clone also exhibits a hypervirulent phenotype. In a recent study by Zhang et al. (Galleria mellonella larvae infection model. The included ST463-PA isolates showed hypervirulence comparable to that of ST235-PA. Moreover, in a clinical retrospective cohort study conducted in East China (The poor outcome associated with ST235 and ST463 CRPA, mainly caused by their hypervirulence and MDR/XDR/PDR profiles, is concerning. Generally, hypervirulence is defined as the ability to cause higher mortality and more severe acute infections . ST235 CblaKPC-2 overexpression as the major driver of high-level IR resistance in ST463-PA. In addition, we also confirmed that blaGES-5 carriage mediated IR resistance in ST235-PA. Overall, this epidemiological survey systematically deciphered the IR resistance mechanisms in P. aeruginosa global high-risk clone ST235 and regional high-risk clone ST463.In conclusion, our data highlighted the distinct IR resistance characteristics between CRPA isolates from high-risk clones and other STs. Of particular concern is the association of high-risk clones with IR resistance phenotype and XDR/DTR phenotype, which may worsen clinical outcomes and restrict therapeutic options. Using whole-genome analysis methods and quantitative PCR, we identified"}
+{"text": "Author Correction: Genome Biol 24, 187 (2023)https://doi.org/10.1186/s13059-023-03023-7Following publication of the original article , the autThese errors do not affect the main results and conclusions of the paper. The original article  has beenAdditional file 2: Supplementary Methods [165-182]. Fig. S1. Imputation accuracy of individual samples for sites with MAF > 1%. Fig. S2. Median copy-number across the genome for wolves. Fig. S3. Repeatmasker classification of SINE variation. Fig. S4. Distribution of variation across the genome for breed and other dogs . Fig. S5. Nucleotide diversity along chr26. Fig. S6. Signature of a retrogene detected at the TEX2 locus."}
+{"text": "Correction : Environmental Microbiome (2020) 15:3 10.1186/s40793-019-0348-0Microviridae viruses found in the metagenomes in the Abstract (Results), Results , Discussion , and Conclusions. Since the article\u2019s publication it has come to light that the sequences associated with the Microviridae viruses belong to PhiX 174 (Genbank Accession Number NC_001422), a member of Microviridae family. These sequences were a contamination originating from the use of PhiX DNA as a control in Illumina sequencing platforms [Microviridae virus(es). After removing PhiX 174-associated sequences from the metagenomic data, the most abundant viral sequences were affiliated with Myoviridae, a viral family which includes phages that are suspected to affect Chlorobi [The original article briefly discusses Additional file 1. Supplementary Materials, Methods and Results. Figure S1.Natural blooms in Trunk River. Figure S2.Color and appearance ofsamples from all holes, depths, and timepoints. Figure S3.Filters thatwere used for biomass measurements and spectral analysis. Figure S4.Total cell count of three samples . Figure S5. Depthprofile representation of chemical data presented in Fig.2. Figure S6. Physicochemistry. Iron, nitrate, ammonium, acetate, Ca2+, and K+measurements. Figure S7. Individual diversity indices of all samples. Figure S8. Trajectories of community structure in hole A, E and K. Figure S9. Relative sequence abundance of the 20 most abundant clades onphylum, class, order, family and genus level, as well as the 20 mostsequence abundant ASVs (amplicon sequence variants). Figure S10.Relative sequence abundance of Chlorobiales ASVs. Figure S11.Relativechange of ASV abundance between surface (V1) and deeper layers (V2-4). Figure S12.Chlorobiales phylogeny. Figure S13. Circular map ofmetagenome-assembled genomes (MAGs). Figure S14. Chlorobiales phy-logenomics. Figure S15. Protein comparison of Bin 6. Figure S16.Pro-tein comparison of Bin 10. Figure S17. Genes involved in sulfurcycling. Figure S18. CRISPR arrays and cas genes predictions Bin 6.Figure S19. CRISPR arrays and cas genes predictions Bin 10. Figure S20. Relative sequence abundance of viral family-level clades. Table S1. Over-view of sequencing output and diversity indices. Table S2. Genome sta-tistics. Table S3. Average nucleotide identity (ANI) comparisons. Table S4. Oxidative phosphorylation and chlorophyll biosynthesis genes of Bin6 and Bin 10. Table S5. CRISPR-Cas system information for eachmetagenome-assembled genome."}
+{"text": "Moreover, Patients with AoAC showed a lower incidence of first-pass successful recanalization  and a higher incidence of postprocedural hemorrhage  than those without AoAC. On multivariate analysis, AoAC was independently associated with first-pass successful recanalization . Conclusions: AoAC on chest radiography can be used as a preoperative predictor of successful first-pass recanalization in patients undergoing EVT for AIS.Background: Vascular conditions can affect the recanalization rates after endovascular thrombectomy (EVT) for acute ischemic stroke (AIS). Chest radiography can assess the conditions of the aortic arch based on the presence or absence of aortic arch calcification (AoAC). The aim of this study was to investigate the relationship between AoAC on chest radiography and first-pass successful recanalization . Methods: We compared the rate of first-pass successful recanalization between patients with and without AoAC. A total of 193 patients with anterior circulation occlusion who underwent EVT between January 2017 and December 2021 were included. Results: AoAC was observed in 80 (41.5%) patients. Patients with AoAC were older (74.5 \u00b1 7.78 vs. 63.9 \u00b1 12.4 years,  Several randomized clinical trials have established the clinical efficacy of endovascular thrombectomy (EVT) for acute ischemic stroke (AIS) ,2,3,4,5.Recent retrospective studies have reported a relationship between the number of device passes and clinical outcomes and documented that recanalization with fewer device passes is associated with better clinical outcomes ,13,14,15Intracranial atherosclerotic stenosis (ICAS) and tortuous vascular anatomy are common causes of EVT failure ,18,19. IAtherosclerosis is a diffuse progressive disease characterized by vascular calcification, which is associated with disease progression ,21,22. CThe aim of this study was to investigate the relationship between AoAC on chest radiography and first-pass recanalization after EVT for AIS.n = 31) and (2) posterior circulation occlusions (n = 29). Eventually, 193 consecutive patients were included  symptomatic AIS, (2) large vessel occlusions confirmed using magnetic resonance (MR) angiography or computed tomography (CT) angiography, (3) less than 24 h from symptom onset to treatment, and (4) at least one-half mismatch between cerebral blood flow and cerebral blood volume map with MR perfusion imaging. A total of 253 patients who met the inclusion criteria were included. From the 253 patients, 60 were excluded because of (1) tandem occlusions , the imaging data and medical records of patients were reviewed. Data regarding the baseline characteristics of the patients, treatment characteristics, and radiological and clinical outcomes were obtained from the medical records. Baseline characteristics included age, sex, past medical history, occlusion side and site, etiology of stroke, AoAC on chest X-ray, type of aortic arch, National Institutes of Health Stroke Scale (NIHSS) at admission, onset-to-door time, and onset-to-puncture time. Treatment characteristics included intravenous thrombolysis, balloon guide catheter, EVT technique, and procedural time. Radiological and clinical outcomes included TICI grades, postprocedural hemorrhage, and the 3-month modified Rankin Scale (mRS) score.The occlusion sites were divided into three categories: (1) internal carotid artery (ICA), (2) M1 segment of the middle cerebral artery (MCA), and (3) M2 segment of MCA. The M1 and M2 segments were defined as the horizontal and vertical segments of MCA, respectively. The etiology of stroke was classified based on the Trial of ORG 10172 in Acute Stroke Treatment (TOAST) criteria. Two experienced radiologists (7 (KHR) and 6 (JB) years of experience) independently evaluated all chest X-rays in a binary manner. During the analysis, all radiologists were blinded to the radiological and clinical outcomes after EVT. AoAC assessments on chest radiographs are shown in There were three neurosurgeons who specialized in neurointerventions at our hospital. When EVT was planned for large vessel occlusion, it was performed by one of these three neurosurgeons. The procedures were performed under local anesthesia via femoral access. The primary EVT modality was at the surgeon\u2019s discretion. The use of a balloon guide catheter  was based on the surgeon\u2019s discretion.p-value of <0.05 was considered statistically significant. Multivariable logistic regression analysis was used to evaluate factors affecting first-pass successful recanalization and good clinical outcomes after EVT for AIS. Variables with a p-value of <0.20 in the univariate analysis were included in the multivariate logistic regression analysis. The interobserver agreement for each chest X-ray feature and the presence of AoAC between the two radiologists was assessed using the Cohen kappa statistic. The degree of agreement was classified using kappa values according to the Landis and Koch recommendation as follows [Baseline characteristics, treatment characteristics, and radiological and clinical outcomes were compared between patients with AoAC and those without, between patients first-pass successful recanalization and those without, and between patients with good clinical outcomes and those without. Categorical variables were analyzed using the chi-squared test or Fisher\u2019s exact test. Continuous variables were analyzed using Student\u2019s t-test or the Mann\u2013Whitney U test. A  follows : kappa vn = 37, 19.2%), cardioembolic occlusion , and unknown . AoAC was observed in 80 (41.5%) patients, while aortic arch types 1, 2, and 3 were observed in 46 (23.8%), 99 (51.3%), and 48 (24.9%) patients, respectively. ICAS was observed in 49 (25.4%) patients. Intravenous tissue plasminogen activator was administered to 78 (40.4%) patients. A balloon guide catheter was used in 34 (17.6%) patients. As first-line EVT strategies, stent retrievers, catheter aspiration, and a combination technique were used in 44 (22.8%), two (1.04%), and 147 (76.2%) patients, respectively. The total number of EVT attempts was one (n = 76), two (n = 43), three (n = 28), four (n = 26), five (n = 12), six (n = 3), seven (n = 2), eight (n = 2), and 12 (n = 1). Successful recanalization was achieved in 162 (83.9%) patients, and first-pass successful recanalization was achieved in 69 (35.8%) patients. Good clinical outcomes were achieved in 110 (57.0%) patients.A total of 193 patients  were analyzed. Of the 193 patients, 41 (21.2%), 119 (61.7%), and 33 (17.1%) had occlusion in the ICA, M1 and M2 segments, respectively. Stroke etiologies included atherosclerotic occlusion (p < 0.001) and were more likely to have hypertension (52/80 (65.0%) vs. 43/113 (38.1%), p < 0.001) and atrial fibrillation (45/80 (56.3%) vs. 39/113 (34.5%), p = 0.003). Aortic arch type 3 was more frequently observed in patients with AoAC (34/80 (42.5%) vs. 14/113 (12.4%), p < 0.001). Patients with AoAC had more EVT attempts . Procedural time was significantly longer in patients with AoAC . Successful recanalization (60/80 (75.0%) vs. 102/113 (90.3%), p = 0.005) and first-pass successful recanalization (15/80 (18.8%) vs. 54/113 (47.8%), p < 0.001) were achieved less frequently in patients with AoAC. Postprocedural hemorrhage was observed more frequently in patients with AoAC (36/80 (45.0%) vs. 31/113 (27.7%), p = 0.015). Although ICH was observed more frequently in patients with AoAC (22/80 (27.5%) vs. 17/113 (15.0%), p = 0.045), the frequency of SAH did not differ significantly between the two groups (14/80 (17.5%) vs. 13/113 (11.5%), p = 0.293). Patients with AoAC had a higher incidence of good clinical outcomes (37/80 (46.3%) vs. 73/113 (64.6%); p = 0.013). On multivariate logistic regression analysis for factors affecting AoAC on chest radiography, including age, sex, hypertension, atrial fibrillation, occlusion site, and NIHSS at admission, age (adjusted odds ratios (OR): 1.106, adjusted 95% confidence interval (CI): 1.063\u20131.151; p < 0.001), and hypertension  were independently associated with AoAC on chest radiography (iography .p = 0.030). Patients with first-pass successful recanalization showed a lower incidence of AoAC on chest X-ray (15/69 (21.7%) vs. 65/124 (52.4%), p < 0.001). Intravenous thrombolysis was administered more frequently in patients with first-pass successful recanalization (35/69 (50.7%) vs. 43/124 (34.7%), p = 0.033). Patients with first-pass successful recanalization had a shorter procedural time . Postprocedural hemorrhage was more frequently observed in patients with first-pass successful recanalization (11/69 (15.9%) vs. 56/124 (45.5%), p < 0.001). Patients with first-pass successful recanalization had a higher incidence of both SAH and ICH (4/69 (5.8%) vs. 23/124 (18.5%), p = 0.016) (7/69 (10.1%) vs. 32/124 (25.8%), p = 0.009). Good clinical outcomes were achieved more frequently in patients with first-pass successful recanalization (53/69 (76.8%) vs. 57/124 (46.0%), p < 0.001). Multivariate logistic regression analysis for factors affecting first-pass successful recanalization after EVT for AIS, including sex, hypertension, occlusion site, AoAC on chest X-ray, and intravenous thrombolysis revealed that AoAC on chest X-ray  and intravenous thrombolysis  were independent factors ( factors .p = 0.003). Hypertension was observed less frequently for patients with good clinical outcomes (47/110 (42.7%) vs. 48/83 (57.8%), p = 0.043). Left-side occlusion was observed less frequently for patients with good clinical outcomes (42/110 (38.2%) vs. 46/83 (55.4%), p = 0.020). Good clinical outcomes were less frequently achieved for patients with ICA occlusion than for those with M1 or M2 occlusions (16/110 (14.5%) vs. 25/83 (30.1%), p = 0.012). Patients with AoAC achieved good clinical outcomes less frequently than those who did not (37/110 (33.6%) vs. 43/83 (51.3%), p = 0.013). Patients with good clinical outcomes had fewer EVT attempts , shorter procedural time , higher rates of successful recanalization (103/110 (93.6%) vs. 59/83 (71.1%), p < 0.001), and first-pass successful recanalization (53/110 (48.2%) vs. 16/83 (19.3%), p < 0.001). Postprocedural hemorrhage (26/110 (23.9%) vs. 41/83 (49.4%), p < 0.001) and ICH (13/110 (11.8%) vs. 26/83 (31.3%), p = 0.001) were observed less frequently in patients with good clinical outcomes; however, the frequency of SAH did not differ significantly between the two groups (13/110 (11.8%) vs. 14/83 (16.9%), p = 0.402). Multivariate logistic regression analysis of factors affecting good clinical outcomes after EVT for AIS, including age, sex, hypertension, diabetes mellitus, atrial fibrillation, occlusion side, occlusion site, AoAC on chest X-ray, aortic arch type 3, intravenous thrombolysis, total number of EVT attempts, procedural time, successful recanalization, first-pass successful recanalization, postprocedural hemorrhage, and ICH revealed that left-side occlusion , first-pass successful recanalization , successful recanalization , and ICH  were independent factors ( factors .p < 0.001).The interobserver agreement showed moderate agreement between the two radiologists  but not atherosclerotic occlusion (TOAST 1) . These results suggest that AoAC on chest X-ray may reflect atherosclerotic changes in a type 3 aortic arch but not intracranial atherosclerotic changes. Previous studies have reported that the pathophysiology of ICAS is different from that of extracranial atherosclerosis [Our results showed that AoAC on chest radiography was significantly associated with a type 3 aortic arch . Recent studies have reported that first-pass recanalization is strongly associated with favorable clinical outcomes ,16,33,34We believe that the ability to predict first-pass successful recanalization, which indicates the procedure\u2019s difficulty before EVT for AIS, could be helpful in deciding the strategy for EVT. Observation of AoAC on preprocedural chest radiography allows neurointerventionalists to prepare for the possibility of longer procedural times and multiple EVT attempts. If the neurointerventionalists can predict the difficulty of the procedure in advance, they can select the techniques or devices that they are familiar with rather than unfamiliar new techniques or new devices; this can help achieve better radiological and clinical outcomes.During the initial examination of patients with AIS, brain CT angiography or MR angiography is performed, and brain CT angiography provides more information about AoAC than chest X-rays. Therefore, chest X-rays may be less useful when brain CT angiography is taken. However, neurointerventionalists try to check brain CTA focusing on intracranial lesions or carotid lesions, and they do not try to check brain CTA focusing on AoAC. Although our study was based on chest X-ray, we can suggest that we have to check AoAC on brain CT angiography as well as intracranial lesions or carotid lesions.First, this was a retrospective, single-center study, and selection bias cannot be ruled out. Second, the sample size was small. Therefore, the statistical power may be low. Third, we demonstrated a significant relationship between AoAC on chest radiography and first-pass successful recanalization after EVT for AIS; however, this was not a causal relationship. Fourth, AoAC was assessed based on the presence of calcification in a qualitative rather than quantitative manner. The impact of AoAC on first-pass successful recanalization should be evaluated quantitatively or semi-quantitatively using more detailed AoAC grades. More evidence with larger sample sizes is required to confirm these preliminary results.Our results suggest that AoAC on chest radiography is a useful predictor of first-pass successful recanalization after EVT for AIS; however, further evidence is required."}
+{"text": "Telomeres are the terminal regions of chromosomes that ensure their stability while cell division. Telomere shortening initiates cellular senescence, which can lead to degeneration and atrophy of tissues, so the process is associated with a reduction in life expectancy and predisposition to a number of diseases. An accelerated rate of telomere attrition can serve as a predictor of life expectancy and health status of an individual. Telomere length is a complex phenotypic trait that is determined by many factors, including the genetic ones. Numerous studies  indicate the polygenic nature of telomere length control. The objective of the present study was to characterize the genetic basis of the telomere length regulation using the GWAS data obtained during the studies of various human and other animal populations. To do so, a compilation of the genes associated with telomere length in GWAS experiments was collected, which included information on 270 human genes, as well as 23, 22, and 9 genes identified in the cattle, sparrow, and nematode, respectively. Among them were two orthologous genes encoding a shelterin protein (POT1 in humans and pot-2 in C. elegans). Functional analysis has shown that telomere length can be influenced by genetic variants in the genes encoding: (1) structural components of telomerase; (2) the protein components of telomeric regions (shelterin and CST complexes); (3) the proteins involved in telomerase biogenesis and regulating its activity; (4) the proteins that regulate the functional activity of the shelterin components; (5) the proteins involved in telomere replication and/or capping; (6) the proteins involved in the alternative telomere lengthening; (7) the proteins that respond to DNA damage and are responsible for DNA repair; (8) RNA-exosome components. The human genes identified by several research groups in populations of different ethnic origins are the genes encoding telomerase components such as TERC and TERT as well as STN1 encoding the CST complex component. Apparently, the polymorphic loci affecting the functions of these genes may be the most reliable susceptibility markers for telomere-related diseases. The systematized data about the genes and their functions can serve as a basis for the development of prognostic criteria for telomere length-associated diseases in humans. Information about the genes and processes that control telomere length can be used for marker-assisted and genomic selection in the farm animals, aimed at increasing the duration of their productive lifetime Telomeres are the terminal regions of chromosomes that ensuretheir stability and represented by evolutionary conservedtandemly repeated DNA sequences  of several kb in length . For example, theirlengths in humans at birth are 10\u201315 kb .3\u02b9 terminal end of a telomere is a single-stranded guanine-richDNA region (150\u2013200 nucleotides), whose end interacts withthe double-stranded region to form the so-called T-loop at thetelomere end. T-loop formation and stabilization are ensuredby a shelterin complex . This structure prevents recognitionof a chromosome terminal region by repair proteins.DNA polymerase is unable to fully replicate the 3\u02b9-end ofa linear DNA during cell division, which leads to a loss of50\u2013200 nucleotides of the telomeric sequence at each celldivision . Telomere shortening can also befacilitated by other factors and processes 1,such as oxidative stress, inflammation, UV irradiation, effectsof toxic agents, DNA replication errors, etc. . These factors are likely to producedifferent effects depending on cell types and the organism\u2019sdevelopment stage and species .http://vavilov.elpub.ru/jour/manager/files/Suppl_Ignatieva_Engl_27_3.pdf.Supplementary Materials are available in the online version of the paper:Telomere shortening initiates cellular senescence. Activationof DNA damage response signaling pathways results ina cell cycle arrest, which may in turn lead to apoptosis andeventually \u2013 to progressive tissue degeneration . The datacollected at the cellular level in vitro and in model organismslay the foundation for using the telomere length as a predictorof life expectancy and health status of an individual. Indeed,studies in humans , mice , sheep , cattle , wild birds , and other animals haveshown that shorter telomere length may be associatedwithreduced life expectancy. The studies in humans discovered anassociation between the telomere length and cardiovasculardiseases, cancer, diabetes, inflammation, and other pathologicalstates .Telomere shortening is prevented by telomerase, a specializedribonucleoprotein complex acting as a reverse transcriptase.In humans, telomerase is active in almost all thecancer cells studied , in blastocyst, in mostsomatic tissues at 16\u201320 weeks of development (except forbrain cells), and ovary and sperm cells at all ontogeneticstages (except for mature spermatozoids and oocytes) .Telomerase activity is controlled by the proteins regulatingexpression of telomerase components, their movementto various cell compartments, processing, and assembly aswell as by the proteins maintaining stability of the telomerasecomplex or, on the contrary, activating its degradation .The main stages of telomerase biogenesis are presented inFigure 2. The examples of proteins affecting the telomeraseactivity are presented in Suppl. Material 2. In addition, thetelomerase activity is also affected by the shelterin  and CST complex .Telomere length is a complex phenotypic trait determinedby multiple factors including genetic ones. The meta-analysisof heritability data for this trait performed in eighteen vertebratespecies showed the averaged heritability index of 45 %.The studies showed the value of 52 % in humans, 42 % \u2013 inHolstein cattle breed, 35 % \u2013 in hamadryas baboons, and5 % \u2013 in sheep .The problem of genetic basis of telomere length regulationis of interest for many researchers. The information on telomerase components and proteins involved in telomere lengthregulation  canbe found in The Telomerase Database (http://telomerase.asu.edu/) . Joyce and co-workers (2018)presented a set of 80 human genes with telomere-related functions.The GWAS data also indicate a polygenic nature of telomerelength heritability. For instance, the GWAS Catalog (https://www.ebi.ac.uk/gwas/) cites 99 human genes that either includeor neighbor the telomere length-associated allelic variants.One of the largest GWA studies presents the data on 138 humangenomic loci, whose allelic variants are associated withtelomere length .In addition to the telomere length association data gatheredusing GWA studies in various human population samples,the data obtained in other animal species also appear to be of interest. However, these studies are rather scarce and areavailable only for Holstein\u2013Friesian cattle , house sparrow nestlings , andC. elegans .The objective of this review was to characterize the geneticbasis of telomere length regulation using the GWAS datacollected in various human populations and to compare themwith the results of similar experiments in other animal species.For this purpose, (1) the data on genes identified in GWAStelomere length experiments were systematized; (2) functionalannotation of genes was performed, and the set of biologicalprocesses affecting telomere length was identified.The data on telomere length-associated genes were obtainedfrom the papers available in the PubMed database (https://pubmed.ncbi.nlm.nih.gov/) using such keywords as \u2018telomerelength\u2019 and \u2018GWAS\u2019. Functional annotation of genes wasperformed using information obtained from the papers presentingGWAS data, PubMed, The Telomerase Database (http://telomerase.asu.edu/) queries, and the DAVID knowledgebase(https://david.ncifcrf.gov/) .Human genes identified through GWA studiesPubMed queries produced 18 scientific papers presenting theresults of identifying telomere length-associated polymorphicloci in human genome based on GWAS data. These paperswere analyzed, and the data on 270 telomere length-associatedgenes were collected . Most genes (262)were identified in European-ancestry population samples, thedata on 15 genes were obtained from the studies in SoutheastAsian population samples , five genes were identified as a result of trans-ethnicmeta-analysis of Singaporean Chinese and European ancestrydata , and one gene was found in AfricanAmericans .The data on functional significance in the context oftelomere length regulation were presented by the authors ofGWA studies for 52 genes out of 270 .The fact that the data on gene significance in the context oftelomere length regulation were unavailable for a number ofloci reflects the capabilities and limitations of GWAS methodology.Most loci identified by GWAS and associatedwith the trait of interest are located in intergenic regions. Asa rule, in these cases, the set of candidate genes includes thenearest genes, whose functional significance is often difficultto interpret. To identify the mechanisms and genes, throughwhich intergenic variants affect the studied traits, additionalexperiments are required. For example, it was shown thatT-to-C substitution of rs1421085 in the intron of FTO geneaffects the expression of IRX3 and IRX5, whose transcriptionstart sites are far away (~520 and ~1160 kb) from rs1421085.Main functional groupsof human telomere length-associated genesA functional classification was performed for a set of 52 humangenes for which there was information about their functionalsignificance in the context of telomere length regulation. As a result, several functional groups ofgenes have been identified Genes encoding telomerase components: TERC is thetelomerase RNA component acting as a matrix for DNA strandextension at the telomere end and TERT is a reverse transcriptaseenzyme subunit .Genes encoding shelterin proteins: components of thiscomplex  can bindto both double-stranded and single-stranded telomeric DNAregions , stabilize them, protect them from exonucleases, reduce telomerase access, and inhibit the proteinsactivated by damaged DNA and involved in double-strandedbreak repair . TheGWAS data on telomere length association were obtained forthe genes coding for five out of six shelterin proteins  .Genes encoding CST proteins: CTC1, STN1, TEN1. CSTcomplex acts as a telomerase negative regulator at the late Sto G2 phase of the cell cycle  .Genes encoding proteins involved in telomerase biogenesisand regulating its activity. One of these genes, ZCCHC10,encodes a protein regulating telomerase synthesis at transcriptionallevel: ZCCHC10 suppresses TERT transcription. Processing and assembly of a telomeraseRNA subunit involves DKC1, NAF1, and SHQ1 , ribonuclease PARN, exoribonuclease DIS3, thecomponent of a nuclear exosome targeting (NEXT) complex,ZCCHC8 , SMUG1 , and CELF4/BRUNOL4 . Noncanonicalpolymerase TENT4B/PAPD5 ,trimethylguanosine synthetase TGS1 , andEXOSC10 RNA exosome component cause a decrease in the level of active TERC. The assemblyof telomerase nucleoprotein complex involves ATPaseRUVBL1/pontin  and telomerase-associatedprotein TEP1 . Two proteins (WRAP53/WDR79/TCAB1 and NOLC1/NOPP140) provide telomeraseaccumulation in Cajal bodies, the small nuclear organelleswhere processing of small nuclear and nucleolar RNAs andassembly or ribonucleoprotein complexes occur . Telomerase activityis modulated by activator protein SMG6/EST1A, which alsobinds to a single-stranded DNA , and PMLprotein, whose isoform PML-IV suppresses telomerase activity.Genes encoding proteins regulating functional activityof shelterin proteins. CSNK2A2 and CSNK2B are the subunitsof casein kinase which phosphorylates TERF1, increasingits binding to telomeres .ATM serine/threonine kinase, on the contrary, decreasesTERF1 binding to the telomeric DNA . PeptidaseUSP7 and ubiquitin ligase SIAH1 activate proteasomaldegradation of POT1 and TERF2, respectively . ADP ribosylases PARP1 and PARP2 reduce the DNAbinding activity of TERF2 .Genes encoding proteins involved in telomere replicationand/or capping: (1) enzymes RRM1 and TYMS involved insynthesis of deoxynucleoside triphosphates (dNTP) and thymidylatesrequired for DNA synthesis ; (2) helicases RTEL1 and MCM4; (3) RPA1 and RPA2, the subunitsof the RPA complex capable to unfold G-quadruplex structuresthat may block DNA replication ;(4) HNRNPA1 promoting telomere capping after DNA replication.Genes encoding proteins affecting the alternative telomerelengthening pathway. This telomerase-independentmechanism  includes the recombinationbetween telomeric regions of two DNA molecules. Three genes identified inGWA studies were attributed to this group . These genes encode SMC6 which activates ALT and its two inhibiting proteins: ATRX withchromatin remodeling activity and SLX4 endonuclease .Genes encoding DNA damage response proteins: (1) peptidaseSENP7 ; (2) chaperone protein BAG6; (3) DCAF4 interacting with CUL4-DDB1ligase ; (4) RFWD3 interacting withRPA protein (replication protein A) .Genes encoding subunits of RNA exosomes: EXOSC6,EXOSC9  and MPHOSPH6 . These proteins are functionally significant, becauseit is known that TERC may be subjected to 3\u02b9-processing,and the RNA-exosomes are involved in this process .Human candidate genes identified in more than one studyAs mentioned above, we have analyzed 18 papers on identifyingtelomere length-associated human genome loci based onGWA studies and collected the data on 270 such genes . Notably, only 16 genes were identified inat least two studies .The most frequently identified genes were the ones encodingboth telomerase components (TERC and TERT ) and STN1encoding a component of the CST complex . Three genes POT1, TERF1,and TERF2 encoding components of the shelterin complexwere mentioned in 4, 3, and 2 publications, respectively.Three more genes DCAF4, RTEL1, and NAF1 controlling theDNA damageresponse, telomere replication, and telomerasebiogenesis were identified in four studies. ATM, PARP1,MPHOSPH6, RFWD3, SENP7, and TYMS were identifiedin 3 or 2 papers.Most of 16 genes listed above were identified in populationsamples of different ethnic origin: (1) TERC in three ethnicgroups, namely Europeans, Bangladeshis, and SingaporeanChinese; (2) DCAF4, MPHOSPH6, and TYMS in Europeansand as a result of the trans-ethnic meta-analyses (SingaporeanChinese+Europeans); (3) TERT, STN1, POT1, RTEL1, NAF1,TERF1, ATM, PARP1 in two ethnic groups, namely Europeansand Singaporean Chinese.Identification of the genes related to telomerelength regulation according to DAVIDUsing DAVID, we found the terms from the GOTERM_BP_DIRECT dictionary that were significantly (FDR < 0.05) associatedwith the list of 270 human genes presented in Suppl.Material 3. Sixteen terms indicating biological processes thatdirectly control telomere length are presented in Suppl. Material6, and the remaining fifteen terms are listed in Suppl.Material 7. There were 30 genes associated with the termsfrom the first group , with two of them (SIRT6 and TP53) previously not recognized as biologicallyinterpretable  without comment on their functional significance inthe context of telomere length regulation). The analysis ofscientific papers showed that proteins encoded by both genescan function in the subtelomeric regions of chromosomes, which means theycould be indirectly involved in telomere length regulationThen, the genes associated with the second group ofGOTERM_BP_DIRECT terms identified at FDR< 0.05  were analyzed. Among them, 29 geneswere found that had no biological interpretation .This group of 29 genes included the above mentioned SIRT6and TP53, as well as BRCA1, SAMHD1, and BRCC3 associatedwith the maximum number of GO terms . Apparently, the genes from the list thusobtained may also be of interest in the context of telomerelength regulation.Telomere length-associated genesfound in other animal speciesGenome-wide search for telomere length-associated loci andgenes was carried out in three animal species: cattle (Bostaurus), sparrows (Passer domesticus), and nematodes (Caenorhabditiselegans).A GWA study to investigate the species Bos taurus wascarried out on 702 animals of the Holstein\u2013Friesian breed. The study of the DNA isolatedfrom the whole blood of cows sampled at birth showed sixtelomere length-associated polymorphic loci, and three additionalloci were identified when analyzing the DNA from bloodsamples at the first lactation. An analysis of the quantitativetrait loci (QTL) corresponding to the identified genetic variantsrevealed 14 candidate genes at birth and 9 at the first lactation. The authors wereunableto find any data on direct involvement of the identifiedgenes in processes associated with telomere length regulation.NUP93 nucleoporin gene encoding a nuclear pore componentwas considered a potential regulator, because it was shownearlier in yeast that nucleoporins facilitated silencing of genesin proximity of telomeric regions .The recently published results of GWA study  in house sparrow (Passer domesticus) nestlings madeit possible to identify 22 candidate genes . According to the authors, the genes ofinterest in the context of telomere length regulation seem tobe as follows: (1) WNT9B encoding a protein component ofWnt/ \u03b2-catenin signaling pathway due to \u03b2-catenin involvementin Tert activation in embryonic stem cells of mice; (2) CDCA4,GH, and GHRHR regulating cell proliferation, apoptosis, andbody growth; (3) RHOF involved in cytoskeletal organization;(4) RNF34 (E3 ubiquitin-protein ligase RNF34) regulatingubiquitination; (5) AQP1 due to involvement of aquaporinprotein in transport of nitrogen oxide and active forms ofoxygen,which increases oxidative stress which can in turnaffect telomerase activity; (6) SCN4A, because its expressionin human stem cells correlates with telomere length.Our analysis showed that none of the candidate genes identifiedin cattle (23 genes) and house sparrows (22 genes) (see theTable) had orthologs among the 270 genes identified based onGWA studies in humans and presented in Suppl. Material3.The study in C. elegans  produced9 candidate genes .One out of nine genes, pot-2, is orthologous to POT1 encodinga shelterin complex component in humans. The authorsassume that another gene ZK1127.4 may also be involved intelomere length regulation, because BCCIP encoded by anorthologous human gene interacts with BRCA2 involved inDNA replicationIn general, when comparing sets of candidate genes identifiedin humans and three other animal species, almost noorthologous genes are detected, which may be due to speciesspecificfeatures of telomere length regulation, some peculiaritiesof regulation at various ontogenetic stages, and differencesin sampled tissues or gender of the studied individuals.In the present paper, a compilation of telomere length-associatedgenes identified based on GWA studies and includingthe data on 270 human genes , as wellas 23, 22, and nine genes identified in cattle, house sparrow,and nematode (see the Table) is presented. The analysis offunctions of 52 human genes with functional interpretationavailable  showed that telomerelength may be affected by variants of genes encoding: (1) thestructural components of telomerase; (2) the protein componentsof telomeric chromosome regions (shelterin complexand CST complex); (3) the proteins involved in telomerasebiogenesis and regulating its activity; (4) the proteins regulatingfunctional activity of shelterin subunits; (5) the proteinsinvolved in telomere replication and/or capping; (6) the proteinscontrolling the alternative telomere lengthening pathway;(7) DNA damage response and repair proteins; (8) RNA exosomecomponentsCandidate human genes identified by several researchgroups in population samples of different ethnic origin aredetermined: genes encoding telomerase components (TERCand TERT ) and STN1 encoding a subunit of CST complex. It seems that polymorphic loci that affect thefunctions of these genes can potentially be the most reliablepredisposition markers for telomere length-associateddiseases.Comparison of the data obtained from GWA studies inhumans  with the results of similarexperiments obtained for other animal species  confirmed and expanded the understandingof the complex polygenic nature of telomere lengthregulation. In addition, a pair of orthologous genes encodinga shelterin protein (POT1 in humans and pot-2 in C. elegans)was identified; this finding demonstrates the high biologicalsignificance of this gene in various species.Systematized data on genes and their functions may lay thefoundation for development of prognostic criteria for humanpathologies explicitly associated with telomere length. 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+{"text": "Citrobacter braakii strain GW-Imi-1b1 was isolated from a hospital wastewater sample in Greifswald, Germany. The genome comprises one chromosome (5.09\u2009Mb), one prophage (41.9\u2009kb), and 13 plasmids (2 to 140.9\u2009kb). The genome harbors 5,322 coding sequences, shows a high potential for genomic mobility, and includes genes encoding proteins for multiple drug resistances.The imipenem-resistant  Citrobacter strains thrive in soil, water, and gut microbiota of humans and animals. They cause nosocomial infections and show various degrees of antibiotic resistance  on Mueller-Hinton agar .sistance . CitrobaC. braakii GW-Imi-1b1 was extracted using the MasterPure complete DNA purification kit , from a strain that had been isolated by repeated streaking and regrowth in 10\u2009mL Mueller-Hinton broth (Carl Roth GmbH & Co.) at 37\u00b0C. Illumina paired-end sequencing libraries were prepared with the Nextera XT DNA sample preparation kit  and sequenced using a MiSeq system and reagent kit v3 (2 \u00d7 300 cycles). Nanopore sequencing library preparation was performed with 1.5\u2009\u03bcg high-molecular-weight DNA, a ligation sequencing kit 1D (SQK-LSK109), and the native barcode expansion kit (EXP-NBD104). Sequencing was performed for 72 h using a MinION Mk1B system with a SpotON R9.4.1 flow cell  and MinKNOW v22.05.5 and Guppy v6.2.1 (hac mode) software.The manufacturer\u2019s recommendations were followed unless otherwise stated. Genomic DNA of NC_001422) leftovers. Furthermore, long reads were treated with Porechop v0.2.4 (https://github.com/rrwick/Porechop) and adjusted to 100\u00d7 coverage with Filtlong v0.2.1 (https://github.com/rrwick/Filtlong), and chimeric reads were removed with unicycler_scrub of Unicycler v0.4.9 (C. braakii GW-Imi-1b1. Unicycler v0.5.0 (de novo hybrid assembly of plasmids (confirmed by BLAST)  . Finallyy BLAST)  and Polyy BLAST) , employihttps://figshare.com/articles/figure/Supplementary_Figure_1/22133567]), and 13 circular plasmids (0.2 to 140.9\u2009kb). The coverages of all genetic entities were calculated with Qualimap v2.2.2 (Citrobacter braakii ASM207534v1 (GenBank accession number GCF_002075345.1), with a FastANI . Genome analysis with Resfams revealed the presence of 27 genes involved in antibiotic resistance, including resistance to \u03b2-lactams, aminoglycosides, quinolones, macrolides, nitroimidazoles, streptomycin, and chloramphenicol and efflux pumps .The genome contains one circular chromosome (5.1 Mbp), one circular (pro)phage . The whole-genome project of Citrobacter braakii GW-Imi-1b1 has been deposited in GenBank under the accession numbers CP115723 (chromosome), CP115737 (phage), CP115724 (plasmid 1), CP115729 (plasmid 2), CP115730 (plasmid 3), CP115731 (plasmid 4), CP115732 (plasmid 5), CP115733 (plasmid 6), CP115734 (plasmid 7), CP115735 (plasmid 8), CP115736 (plasmid 9), CP115725 (plasmid 10), CP115726 (plasmid 11), CP115727 (plasmid 12), and CP115728 (plasmid 13), with BioProject accession number PRJNA524094 and SRA accession numbers SRR23033312 (Oxford Nanopore Technologies MinION reads) and SRR23033313 (Illumina MiSeq reads).The assembled genomic entities , nucleotide and amino acid sequences, and GenBank records are accessible via Figshare ("}
+{"text": "Pseudomonas aeruginosa (PSA) is a common cause of acute pulmonary exacerbations in adult cystic fibrosis (CF) patients. In this setting, culture isolates are often multidrug resistant (MDR) or exhibit difficult-to-treat resistance (DTR) due to frequent antimicrobial exposure, resulting in the use of last-line or novel therapies  using a collection of clinical PSA isolates recovered from adult CF respiratory specimens.herapies . Cefiderherapies \u20137. TheA total of 31 adult CF PSA isolates were selected from an internal repository . These rin vitro potency compared with I-R, C-T, and CZA (50/MIC90 of CFDC was 0.25/16 \u00b5g/mL, respectively. The MIC50/MIC90 of I-R, C-T, and CZA was 2/32, 1/128, and 2/128 \u00b5g/mL, respectively (50 compared with the other novel agents (CFDC MIC50/MIC90 of 1/128 \u00b5g/mL; 75% susceptible per CLSI; Overall, CFDC retained the greatest  and CZA . The MICectively . Of notein vitro potency against PSA compared with C-T, CZA, and I-R. However, changes in target binding sites of PSA-derived AmpC \u03b2-lactamases may result in an increase in CFDC MICs (Antibiotic management of pulmonary exacerbations in the setting of CF is challenging as the etiologic agents often exhibit MDR. CFDC has enhanced activity against MDR PSA and is, therefore, an attractive option for intervention; albeit, CFDC resistance may occur , 13. TheFDC MICs . FurtherFDC MICs . Our stu"}
+{"text": "Response to NACT was evaluated using CA125 response and chemotherapy response score (CRS). Compared with the non-responders, the responders displayed a larger proportion of tumors showing increase in the infiltration of CD20+ cells (P = 0.046) and in the M1/M2 ratio (P = 0.038) as well as fewer tumors showing increase in the infiltration of CD56bright cells (P = 0.041). No association was found between pre-NACT TIME and response to NACT. Density of pre-NACT CD8+ cells was positively associated with longer progression-free survival (PFS) (P = 0.011) and overall survival (OS) (P = 0.048). Post-NACT CD20+ and CD163+ macrophages (M2) infiltrates were associated with prolonged (P = 0.005) and shortened PFS (P = 0.021), respectively. Increase in the density of CD4+ T cells was predictive for longer PFS (P = 0.022) and OS (P = 0.023). In the multivariate analysis, high density of CD8+ cells pre-NACT (P = 0.042) were independently associated with improved OS.Little is known about the association between efficacy of neoadjuvant chemotherapy (NACT)/survival and the dynamic change of tumor immune environment (TIME) during treatment in epithelial ovarian cancer (EOC). This study investigated the TIME landscape of treatment-naive EOC tumors using multiplex immunofluorescence and associated the TIME before and after platinum-based NACT with treatment efficacy and prognosis in 33 patients with advanced EOC. NACT significantly increased the density of CD8 Ovarian cancer is the most lethal gynecologic malignancy . Epithel+ cytotoxic cells, and CD3+ T cells] or deleterious  to ovarian cancer patients  of treatment-na\u00efve tumor has allowed for identifying components of immune contexture that are beneficial . Serous carcinoma was the predominant histologic subtype , and most of the patients had FIGO stage III disease . Almost all the tumors were grade 3 . Approximately half of the patients had lymph node metastasis. The median CA125 level was 1632.0 U/mL at baseline. Seventy-six percent of patients received two cycles of NACT . CA125 response to 1 (57.6%) Table\u00a01.+ T cells, significantly increased    . No significant difference was found in the density of other immune cell subsets, including CD3+, CD4+Foxp3+, CD4+, CD68+CD163- and PD-1+CD8+ . We also observed a significant increase in the infiltrates of PD-1+ cells (P = 0.048), PD-L1+CD68+ macrophages (P = 0.017), CD20+ B cells (P = 0.033), and CD56dim (P = 0.022) , PD-1+ cells (P = 0.011), PD-L1+CD68+ macrophages (P = 0.023) and PD-1+CD8+ cells (P = 0.035) in the stoma of CRS2 responders (+Foxp3+ Treg cells (P = 0.036) and PD-L1+CD68+ macrophages (P = 0.022) in the stroma and decreased M1/M2 ratio (P = 0.016) in the tumor of CRS1 non-responders (+ cells (P = 0.046) and in the M1/M2 ratio (P = 0.038) as well as fewer tumors showing increase in the infiltration of CD56bright cells (P = 0.041)  and M2 macrophages  in the tumor  and PFS   (P = 0.034) (Data not shown), and lower density of CD68+CD163+ M2 macrophages  in the post-NACT tumor were found to predict better PFS  and OS (log-rank P = 0.023) , and OS  and tumoral CD8+ pre-NACT  were independently associated with improved OS. The difference in PFS between pre-NACT CD8high and pre-NACT CD8low patients was marginally significant (P = 0.054)  Table\u00a03.+ T cell, CD20+ B cells, and PD-L1+CD68+ macrophages. The biopsies pre-NACT from patients with good response to NACT had significantly increased infiltration of CD8+ T cells and CD20+ B cells compared with tumor specimens post-NACT. Infiltrates of CD8+ T cells pre-NACT, CD20+ B cells and CD163+ macrophages post-NACT, and marked change in CD4+ T cells during NACT are associated with prolonged survival.In the current study, we reported that the infiltration of immune cells in the tumor lesions pre-NACT and post-NACT was associated with response to NACT and long-term survival benefits. With mIF, we analyzed multiple immune cell subsets closely associated with cancer dissemination in a cohort of patients with paired pre-NACT and post-NACT specimens. We show that NACT altered the balance of immune cell subsets of T cells, B cells, and macrophages, showing significantly elevated density of CD8+ T cells, CD3+ T cells, CD20+ B cells, and PD-1+ cells  were associated with prolonged and shortened PFS, respectively. There is evidence that a high M1:M2 ratio post-NACT is predictive of improved PFS and OS  for the publication of any potentially identifiable images or data included in this article.GC: Acquisition of data, writing-review and editing. DH: Methodology, writing-review and editing. JL: Data curation, formal analysis, writing-review and editing. XZ: Data curation, formal analysis, writing-review and editing. ZZ: Data curation, formal analysis, writing-review and editing. BZ: Data curation, formal analysis, writing-review and editing. TB: Writing\u2013original draft, writing\u2013review and editing. LC: Conceptualization, resources, supervision, writing\u2013review and editing. SC: Conceptualization, resources, supervision, writing\u2013review and editing. SW: Resources, data curation, writing-review and editing. LZ: Conceptualization, resources, supervision, methodology, project administration, writing\u2013review and editing. All authors contributed to the article and approved the submitted version."}
+{"text": "Correction: Arthritis Res Ther 25, 143 (2023)https://doi.org/10.1186/s13075-023-03129-0Following publication of the original article , the autThe sentence currently reads:17 studies were included , indicating that multiple sclerosis patients are more likely to suffer from periodontal diseases and TMD [7]. TMD and periodontal disease might be the first symptoms of several rheumatic diseases [8].The sentence should read:Unilateral mastication due to PD could induce pain and structural changes in the temporomandibular joint [7]. In addition to clinical observational studies, two symptoms often occur together in many diseases, such as rheumatic diseases  and multiple sclerosis [10].Updated references:J Periodontal Implant Sci 2017, 47(4):211-218.7. Jeon HM, Ahn YW, Jeong SH, Ok SM, Choi J, Lee JY, Joo JY, Kwon EY: Pattern analysis of patients with temporomandibular disorders resulting from unilateral mastication due to chronic periodontitis. Journal of clinical medicine 2018, 8(1).8. Gualtierotti R, Marzano AV, Spadari F, Cugno M: Main Oral Manifestations in Immune-Mediated and Inflammatory Rheumatic Diseases. Reumatol Clin (Engl Ed) 2020, 16(4):262-271.9. Gonz\u00e1lez-Ch\u00e1vez SA, Pacheco-Tena C, Campos Torres RM, Qui\u00f1onez-Flores CM, Reyes-Cordero G, Caraveo Frescas TJ: Temporomandibular and Odontological Abnormalities in Patients with Rheumatoid Arthritis. Evidence-based dentistry 2021, 22(1):44-45.10. Al-Ansari A: Is there an association between multiple sclerosis and oral health? The original article  has been"}
+{"text": "Predictors of SOF/VEL/VOX failure were genotype 3, active HCC, baseline cirrhosis, and prior SOF/VEL. Baseline RAS mutation and ribavirin supplementation were not associated with higher SVR12. Treatment discontinuation because of drug-related adverse events was uncommon . In summary, SOF/VEL/VOX is efficacious and safe for retreatment in DAA-experienced CHC patients, even with RAS mutation. Our findings support SOF/VEL/VOX as a first-line rescue treatment for DAA-experienced CHC patients. About 5% of chronic hepatitis C (CHC) patients experienced treatment failure with direct-acting antiviral (DAA) treatment. The global data on the practice and treatment outcomes of Sofosbuvir/Velpatasvir/Voxilaprevir (SOF/VEL/VOX) in DAA-experienced CHC patients remains sparse. We performed a systematic review and meta-analysis to evaluate the efficacy and safety of SOF/VEL/VOX as a salvage treatment in DAA-experienced CHC patients. We searched five electronic databases from inception to 31 January 2023. The study outcomes were SVR12 and treatment-related adverse effects, with subgroup analysis performed based on genotype, cirrhosis, HCC, prior SOF/VEL exposure, and region. We identified and analyzed data from 24 studies (2877 DAA-experienced CHC patients); 17.2% had prior SOF/VEL exposure, 25% received ribavirin with SOF/VEL/VOX, and 42% had pre-treatment resistance-associated substitution (RAS) testing performed. Eastern Mediterranean had a higher pooled SVR12 than the America and Europe regions ( Chronic hepatitis C (CHC) infection affects 56.8 million people globally, with about 1.5 million new infections every year, resulting in 400,000 deaths each year . VirologTreatment options remain limited in DAA-experienced CHC patients who fail to achieve SVR12. Retreatment of CHC patients with DAA failure could be challenging because of the emergence of resistance-associated substitution (RAS). Current guidelines recommend Sofosbuvir/Velpatasvir/Voxilaprevir (SOF/VEL/VOX) as a salvage treatment for DAA-experienced CHC patients who failed to achieve SVR12 . SOF/VELThe global data on practice and treatment outcomes of SOF/VEL/VOX remained lacking, especially in regions such as Asia and Africa. Although some studies reported a lower SVR12 among CHC patients with liver cirrhosis or genotype 3, which is prevalent in south Asia ,10, suchWe followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guideline for data extraction and reporting . With thWe included all studies that met one of the following inclusion criteria, regardless of publication dates and publication status. Our inclusion criteria were as follows: (1) DAA-experienced, adult CHC patients (age > 18 years old) treated with 12 weeks of SOF/VEL/VOX, with or without Ribavirin, and (2) reported data on either SVR12 or adverse events. Our exclusion criteria were (1) paediatric CHC patients; (2) animal studies; (3) studies that did not report study outcomes; (4) case reports or case series with less than 10 patients; (5) reviews, editorials, or guidelines; and (6) clinical trials. Three authors  independently performed the initial screening of the title and abstract identified in the primary search for eligibility. Any discrepancy in the article selection was resolved by discussion and consensus with the senior author (W.Y.J.). The data from each study were independently extracted using a standardized form. We contacted the corresponding author to verify any missing information. The data extracted included demographics of the study participants (age and genotype of hepatitis C virus), location, sample size, RAS testing at baseline, SVR12, concurrent use of ribavirin (RBV), and treatment-related adverse events requiring discontinuation of treatment.The outcomes studied were as follows: (1) SVR12 (both the intention-to-treat (ITT) and per-protocol (PP) analysis) and (2) frequency of adverse events leading to treatment discontinuation. The ITT analysis included all subjects treated with SOV/VEL/VOX, while the PP analysis only included SOF/VEL/VOX-treated CHC patients with available SVR12 results. Subgroup analysis was performed based on region, HCV genotype, baseline cirrhosis status, prior SOF/VEL exposure, hepatocellular carcinoma, RAS mutation, and concurrent use of ribavirin. p-value of less than 0.05 was considered to be statistically significant. Study heterogeneity was assessed by the 2I% statistics. We used Review Manager Software version 5.3  to perform our meta-analysis. The effect measures were estimated using the odds ratio (OR) for categorical outcomes and mean difference (MD) for continuous outcomes together with the respective 95% confidence interval (95%CI). The meta-analysis was performed using the random-effects model. A The quality of all included studies was independently assessed by three authors using the Risk Of Bias In Non-Randomized Studies of Interventions (ROBINS-I) tool (non-randomized studies)  and the . Most studies had either a low risk [From an initial total of 522 citations identified using our search strategy, 92 full texts were reviewed. We excluded 68 citations for the following reasons: clinical trial (n = 41), treatment-na\u00efve CHC patients (n = 8), case report or study with less than 10 patients n = 17), and missing data (n = 2). Finally, a total of 24 studies were included for analysis . Most st, and mislow risk ,21,22,23low risk ,31,32,33A total of 2887 DAA-experienced CHC patients from 24 studies were included. All patients received 12 weeks of SOF/VEL/VOX, with or without ribavirin. The patient characteristics of the included studies are summarised in p = 0.0006), Western Pacific region (five studies: 94.7% (95%Cl: 91.3\u201397.1%), p = 0.019), and Europe region (nine studies: 95.8% (95%Cl: 94.5\u201396.7%), p = 0.04) (p < 0.001) and GT3 infection . The real-world studies had a lower pooled SVR12 than the clinical trials (87.7% (95%CI: 86.4\u201389.2%) vs. 98.2% (95%CI: 95.9\u201399.2%), p < 0.0001). The overall SVR12 for DAA-experienced CHC patients treated with SOF/VEL/VOX based on PP and ITT analysis was 95% (95%CI: 94.0\u201395.8%) and 88% (95%CI: 86.7\u201389.5%), respectively. The SVR12 in the Eastern Mediterranean region (two studies: 98.2% (95%Cl: 96.0\u201399.3%)) was significantly higher than the region of Americas ,  = 0.04) . The higI2 = 7%)  and non-GT3 DAA-experienced CHC patients treated with SOF/VEL/VOX was 86.1% (95%CI: 82.3\u201389.3%) and 94.9% (95%CI: 93.4\u201396.1%), respectively. GT3 HCV infection was associated with a lower odds of SVR12  .I2 = 0%) (I2 = 3%)  and 97.7% (95%CI: 95.5\u201399.0%), respectively. Liver cirrhosis was associated with a lower odds of SVR12  A. DecompI2 = 3%) B.I2 = 54%)  and 91.7% (95%CI: 90.0\u201393.5%), respectively. Prior SOF/VEL exposure was associated with a lower odds of SVR12  A. I2 = 0%)  and 92.8% (95%CI: 90.3\u201395.3%), respectively. Active HCC during SOF/VEL treatment was associated with a lower odds of SVR12  B.I2 = 34%)  and 91.6% (95%CI: 85.1\u201398.1%), respectively. The odds of SVR12 were similar among DAA-experienced CHC patients receiving SOF/VEL/VOX, with or without baseline RAS testing  .I2 = 55%)  and 96.2% (95%CI: 94.0\u201398.5%), respectively. The odds of SVR12 were similar among DAA-experienced CHC patients receiving SOF/VEL/VOX, with or without ribavirin  .From a total of 1283 DAA-experienced CHC patients received SOF/VEL/VOX, the pooled risk of severe adverse events that occurred during SOF/VEL/VOX treatment was 1.94% (95%CI: 1.2\u20132.8%). The types of serious adverse events are summarised in In this meta-analysis, we found that SOF/VEL/VOX is an efficacious and safe salvage therapy among DAA-experienced CHC patients. The pooled SVR12 was high and treatment discontinuation due to treatment-related SAE was uncommon. Predictors for a lower SVR12 rate by SOF/VEL/VOX were genotype 3, active HCC, baseline cirrhosis, decompensated cirrhosis, and prior SOF/VEL exposure. Meanwhile, the presence of baseline RAS mutation and addition of ribavirin to SOF/VEL/VOX was not associated with higher SVR12. The higher overall SVR12 among the Eastern Mediterranean studies is likely attributed to a lower proportion of patients with GT3 infection and liver cirrhosis. SOF/VEL/VOX, being a protease-inhibitor based DAA, was discouraged in patients with decompensated cirrhosis because of potential toxicity from delayed drug clearance. Among 34 decompensated CHC patients treated with SOF/VEL/VOX in five different studies ,31,32,33Current EASL guidelines recommend the use of RBV with SOF/VEL/VOX in patients with a higher risk of treatment failure; however, the benefits of routine addition of RBV remain unclear . We founThis meta-analysis provides a comprehensive global perspective on the effectiveness and safety of SOF/VEL/VOX among DAA-experienced CHC patients. Prior to this, information on practice and treatment outcomes of SOF/VEL/VOX use among Asian and African population was limited. We acknowledge that there are limitations when interpreting our findings. Most studies did not report the compliance and potential drug interaction among patients receiving SOF/VEL/VOX. The number of patients within subgroups of decompensated cirrhosis was small because the current guidelines discourage the use of SOF/VEL/VOX in these patients. Nevertheless, the results of this meta-analysis provide important data to support the recommendation of using SOF/VEL/VOX as a first-line option among DAA-experienced CHC patients and could be considered level 1 evidence. To conclude, SOF/VEL/VOX is a well-tolerated and highly effective rescue therapy among DAA-experienced CHC patients who failed to achieve SVR12. Predictors of treatment failure include GT3, liver cirrhosis, and active HCC. Safety data of SOF/VEL/VOX among decompensated cirrhosis patients should be investigated in further studies."}
+{"text": "Cryptosporidium spp. are important zoonotic agents of bovine diarrhea. However, little is known about microbiota and short-chain fatty acids (SCFAs) changes in yaks infected with Cryptosporidium spp. Therefore, we performed 16S rRNA sequencing and detected the concentrations of SCFAs in Cryptosporidium-infected yaks. Results showed that over 80,000 raw and 70,000 filtered sequences were prevalent in yak samples. Shannon (p<0.01) and Simpson (p<0.01) were both significantly higher in Cryptosporidium-infected yaks. A total of 1072 amplicon sequence variants were shared in healthy and infected yaks. There were 11 phyla and 58 genera that differ significantly between the two yak groups. A total of 235 enzymes with a significant difference in abundance (p<0.001) were found between healthy and infected yaks. KEGG L3 analysis discovered that the abundance of 43 pathways was significantly higher, while 49 pathways were significantly lower in Cryptosporidium-infected yaks. The concentration of acetic acid (p<0.05), propionic acid (p<0.05), isobutyric acid (p<0.05), butyric acid (p<0.05), and isovaleric acid was noticeably lower in infected yaks, respectively. The findings of the study revealed that Cryptosporidium infection causes gut dysbiosis and results in a significant drop in the SCFAs concentrations in yaks with severe diarrhea, which may give new insights regarding the prevention and treatment of diarrhea in livestock.Diarrhea is a severe bovine disease, globally prevalent in farm animals with a decrease in milk production and a low fertility rate.  The long-haired ruminant yak is a plateau bovine species living in the 3000-5000\u00a0m high-altitude regions and is mostly found on the Qinghai Tibet plateau . DiarrheEscherichia coli, Salmonella spp., and Cryptosporidium spp. have been commonly observed in infected cattle  with equal number of negative samples (n=4) were sequenced and divided into infected (INF) and healthy (H) groups, respectively.Fecal samples (n=40) were collected from free-ranged yaks in Xining, Qinghai  and examined for sted PCR  and posiThe extraction of total genomic DNA was performed by utilizing a commercial TIANamp Stool DNA Kit (Tiangen Biotech (Beijing) Co., Ltd, China) according to the product\u2019s specifications. Fecal DNA concentration, purification, and quality examination were performed through NanoDrop 2000 UV-Vis spectrophotometer  and 1.2% agarose gel electrophoresis, respectively. Then the hypervariable regions of bacterial 16S rRNA gene (V3-V4) were amplified using primers 338F and 806R as described in a previous study . All PCR\u00ae OnePot II DNA Library Prep Kit for Illumina\u00ae  according to the product\u2019s instructions, and sequenced through the Illumina NovaSeq platform . Quality control of sequencing data was performed by employing QIIME2 (https://docs.qiime2.org/2019.1/) to generate amplicon sequence variant (ASV) (p<0.05 and log2 (FoldChange) > 2), clustering heatmap (with Z-score > 0.5 or < -0.5) and evolutionary tree (p<0.05) methods to reveal differences in bacterial abundance among yak samples   and taxont (ASV) . Analysi samples . Microbi samples , and parpost-hoc test to measure the individual differences in microbial function between the yak groups with a p<0.05 as statistically significant.The potential KEGG Ortholog (KO) functional profiles of yak microbiota was predicted with PICRUSt  by annotvia t-test.The concentrations of SCFAs in fecal samples were detected by employing GC-MS , and theP values < 0.05 were considered as statistically significant.The differences between different yak groups were calculated by the chi-square test piloting IBM SPSS Statistics (SPSS 22.0). p<0.01) and Simpson (p<0.01) were both significantly higher in group INF than in group H , Proteobacteria (8.97%), and Actinobacteria (8.72%) in group H, while Firmicutes (56.38%) and Bacteroidetes (29.83%) were the main phyla in group INF (Clostridia (51.13%) and Bacilli (17.28%) were the primary classes in healthy yaks, while Clostridia (51.13%) and Bacteroidia (29.83%) were the major classes in infected yaks , Lactobacillales (8.20%), and Bacillales (8.10%) were the primary orders in healthy yaks, while Clostridiales (51.04%) and Bacteroides (29.83%) were the main orders in infected yaks (Ruminococcaceae and Lachnospiraceae in groups H and INF (Pseudomonadaceae Pseudomonas (6.13%), and Lactobacillus (6.00%) were the dominating genera in healthy yaks, while unclassified (69.25%), Prevotellaceae Prevotella (5.13%) and Arthrobacter (2.45%) were the main genera in infected yaks (Veronii (6.11%), and Alactolyticus (1.66%), while unclassified (95.15%), Flavefaciens (1.50%) and Veronii (1.12%) were the main bacteria in group INF  and evolutionary tree (top 50 abundance) with heat map analysis were plotted. The results revealed that at the order level, infected yaks showed an abundance of Bacteroidia and Deltaproteobacteria, while healthy animals showed abundance of Bacilli, Erysipelotrichi, Betaproteobacteria, Alphaproteobacteria, and Nitriliruptoria as expressed in the clustering heatmap. The evolutionary tree also showed an obvious abundance difference in Betaproteobacteria, Fibrobacteria, SJA_176, 4C0d_2, Nitriliruptoria, Clostridia, and Bacilli between groups H and INF , Bacteroidetes (p<0.0001), Armatimonadetes (p<0.0001), Fibrobacteres (p<0.01), and Synergistetes (p<0.01) were visibly higher in infected yaks, while Cyanobacteria (p<0.0001), Proteobacteria (p<0.0001), Armatimonadetes (p<0.0001), Euryarchaeota (p<0.0001), Actinobacteria (p<0.01), Firmicutes (p<0.01), and Elusimicrobia (p<0.05) were significantly lower (p<0.0001), Prevotellaceae_Prevotella (p<0.0001), CF231 (p<0.0001), L7A_E11 (p<0.0001), BF311 (p<0.0001), Desulfovibrio (p<0.0001), Succiniclasticum (p<0.0001), Desemzia (p<0.0001), Anaerovorax (p<0.0001), Pseudobutyrivibrio (p<0.0001), Acinetobacter (p<0.0001), Fibrobacter (p<0.0001), Ruminococcaceae_Ruminococcus (p<0.0001), Anaerorhabdus (p<0.0001), Treponema (p<0.0001), Selenomonas (p<0.001), Clostridium (p<0.001), Shuttleworthia (p<0.001), Dehalobacterium (p<0.001), TG5 (p<0.01), unclassified (p<0.01), Anaerostipes (p<0.01), Syntrophomonas (p<0.01), Brachymonas (p<0.01), Pyramidobacter (p<0.01), SHD_231 (p<0.05), Butyrivibrio (p<0.05), Desulfobulbus (p<0.05), RFN20 (p<0.05), and Anaerofustis (p<0.05) were significantly higher in infected yaks, while Turicibacter (p<0.0001), Lactobacillus (p<0.0001), Sporosarcina (p<0.0001), Ralstonia (p<0.0001), Akkermansia (p<0.001), Streptococcus (p<0.001), Methylobacterium (p<0.01), Adlercreutzia (p<0.01), Faecalibacterium (p<0.01), Roseburia (p<0.01), Paenibacillus (p<0.01), Methanosphaera (p<0.01), Pseudomonadaceae_Pseudomonas (p<0.01), Peptostreptococcaceae_Clostridium (p<0.01), Slackia (p<0.01), Cupriavidus (p<0.01), Halomonas (p<0.01), Gemmiger (p<0.01), Dietzia (p<0.01), Blautia (p<0.05), Agrobacterium (p<0.05), Nesterenkonia (p<0.05), Sanguibacter (p<0.05), Phascolarctobacterium (p<0.05), Actinomycetospora (p<0.05), Bifidobacterium (p<0.05), SMB53 (p<0.05), and Dorea (p<0.05) were significantly lower in infected animals  were found between healthy and infected yaks, with 119 higher and 116 lower abundance enzymes in INF yaks , propionic acid (p<0.05), isobutyric acid (p<0.05), butyric acid (p<0.05) and isovaleric acid was significantly lower in infected yaks, respectively, while there was no significant difference of valeric acid and caproic acid between H and INF groups  and Simpson (p<0.01)   can be found below: The animal study was reviewed and approved by ethics committee of Nanjing Agricultural University.KL and QW, research idea and methodology. HD, XC, XZ, and CZ, reagents, materials, and analysis tools. KL, writing-original draft and preparation. KM, MF-E-A, ZB, QW, JZ, SN, and KL, writing-review and editing. KL, JZ, and QW, visualization and supervision. All authors contributed to the article and approved the submitted version."}
+{"text": "L265P and MYD88other.This study aims to investigate the clinical and molecular differences between diffuse large B-cell lymphoma (DLBCL) patients with MYD88L265P and MYD88other were investigated.DLBCL patients with MYD88 variations were collected from the Cancer Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College (CHCAMS), and Suzhou Municipal Hospital from February 6th, 2007 to May 20th, 2022. Clinicopathological parameters and treatment outcomes between MYD88L265P, while 54 were MYD88other. MYD88L265P was more common in non-GCB subtype than MYD88other . Besides, MYD88L265P was significantly related to higher proportion of testicle/ central nervous system involvement , PIM1 mutation , and PIM1 hypermutation , compared with MYD88other. Compared with MYD88L265P, MYD88other were more likely to have higher percentage of advanced stage , extranodal site\u2009\u2265\u20092 , elevated LDH , positive CD10 expression , BCL-6 translocation , and NOTCH pathway gene alteration . In non-GCB DLBCL subtype, patients with MYD88other were significantly associated with worse progression free survival (PFS) than those with MYD88L265P when treated initially with R-CHOP/R-CHOP-like regimen (P\u2009=\u20090.010).A total of 132 patients with MYD88 variations from a cohort of 475 DLBCL patients were included, among which, 78 were MYD88L265P and MYD88other are likely to be two subgroups with different clinical and molecular characteristics. The survival of patients with MYD88other is not superior than those with MYD88L265P, even poorer when focusing on the non-GCB subtype.The findings of this study indicate that DLBCL patients with MYD88The online version contains supplementary material available at 10.1007/s00432-023-04714-1. A total of 132 patients with MYD88 variations were included for analysis. The inclusion criteria were as follows: (1) patients were histologically diagnosed with DLBCL based on the World Health Organization classification of Tumors of Hematopoietic and Lymphoid Tissue (2008)  performance status (PS), number of extranodal involvement sites, lactate dehydrogenase (LDH) level, International Prognostic Index (IPI) score, bulky disease, B symptoms, primary site, testicle/central nervous system (CNS) involvement, relapse, bone marrow/peripheral blood involvement, cell of origin (COO) type, first-line therapy, protein expression of CD5, CD10, CD20, BCL-2, BCL-6, c-MYC, MUM-1, Ki-67, and PD-L1, genetic alteration of CD79, TP53, BCL-2, BCL-6, c-MYC, PIM1, SGK1, cell cycle-related genes , NOTCH pathway-related genes , JAK-STAT pathway-related genes , PI3K pathway-related genes , immune-related genes , epigenetics-related genes , RAS pathway-related genes . Any gene occurred alteration in the pathway will be defined as \u201cpathway gene alteration\u201d.Formalin-fixed paraffin-embedded (FFPE) tissues were obtained from included patients. The protein expression of CD5, CD10, CD20, BCL-2, BCL-6, c-MYC, MUM-1, and Ki-67 were evaluated by immunohistochemistry (IHC) using associated antibodies . The Programmed death-ligand 1 (PD-L1) expression was detected using the 22C3 anti-PD-L1 rabbit monoclonal antibody . The COO classification was confirmed according to the Hans algorithm  was performed on three-micrometer-thick FFPE tumor tissues using Vysis LSI CMYC/BCL2/BCL6 Dual Color Break Apart Rearrangement Probe  according to the manufacturer\u2019s instructions. Assessment of FISH signals was performed using Zeiss Axio Imager M2 epifluorescence microscope . Fifty tumor cells were counted, and the percentage of tumor cells with split-signal over 15% indicated the translocation of c-MYC, BCL-2, or BCL-6.Genomic DNA was extracted from the collected FFPE DLBCL tissues using the QIAamp DNA FFPE tissue kit . DNA concentration was quantified using Qubit dsDNA HS Assay Kit . DNA fragmentation was conducted using Covaris S2 Ultrasonicator . Fragments of 200\u2013250\u00a0bp were selected by AMPure beads . End repair, phosphorylation, adaptor ligation, hybridization with capture-probe baits, hybrid selection with magnetic beads, and polymerase chain-reaction amplification were subsequently processed. Two capture panels were adopted, one consisted of 112 commonly altered genes in lymphoma and hematologic malignancies, the other covering 413 frequently mutated genes in DLBCL. There were 101 overlapping genes between the two panels. Capture-based targeted sequencing was performed on a Next Seq500 Sequencer  with pair-end reads at Geneplus-Beijing  or Burning Rock Biotech . Detailed sequencing procedure was performed as our previous study described .Chi-squared test or Fisher\u2019s exact test when appropriated was adopted for the comparisons between categorical variables. Kaplan\u2013Meier survival curve and log-rank test were performed for comparing the PFS and OS. L265P, while 41% (54/132) were MYD88other. Among patients with MYD88other, the p.S219C (14/54), p.L273P (10/54), and p.S243N (6/54) were the top three frequently occurred mutations . Among the patients with MYD88 variation, 59% (78/132) occurred MYD88L265P and MYD88other variants . A total of 21 patients eventually occurred death. No statistically significance was observed in PFS and OS between MYD88L265Pand MYD88other   , which is consistent with previous reports  domain, which is the same domain as MYD88L265P resides. As regards the ability to activate NF-\u03baB, MYD88wild\u2212type presents modest activity, MYD88L265P has strong activity, as does MYD88M232T and MYD88S243N, while MYD88S222R and MYD88T294P exert intermediate effect. Their results suggested MYD88other can contribute to the constitutive NF-\u03baB activation in ABC DLBCL as with MYD88L265P  and NFKB2/p52  Below is the link to the electronic supplementary material."}
+{"text": "R. Soc. Open Sci.10: 230437 (Published online 6 September 2023). (https://doi.org/10.1098/rsos.230437)et al. [Corrigendum to the last five papers in the reference list of Bandeli et al. : referenWith Veasey 2020 re-inserted (and two other references swapped in position) the correct numbering for the last few references (now [101\u2013105]) is below:Panthera tigris altaica) as a model species. Animals10, 1536. (doi:10.3390/ani10091536)101. Veasey JS. 2020 Can zoos ever be big enough for large wild animals? A review using an Expert Panel Assessment of the psychological priorities of the Amur tiger 102. Kirchgessner ML, Sewall BJ. 2015 The impact of environmental, social, and animal factors on visitor stay times at big cat exhibits. Behavioural Brain Research33, 229\u2013240. (doi:10.1016/S0166-4328(89)80118-4)103. Cahusac PM, Miyashita Y, Rolls ET. 1989 Responses of hippocampal formation neurons in the monkey related to delayed spatial response and object-place memory tasks. et al. 2017 Suite of simple metrics reveals common movement syndromes across vertebrate taxa. Movement Ecology5, 12. (doi:10.1186/s40462-017-0104-2)104. Abrahms B Scientific Reports6, 36361. (doi:10.1038/srep36361)105. Tidi\u00e8re M, Gaillard JM, Berger V, M\u00fcller DW, Bingaman Lackey L, Gimenez O, Clauss M, Lema\u00eetre JF. 2016 Comparative analyses of longevity and senescence reveal variable survival benefits of living in zoos across mammals. In the text, this means that in the final two paragraphs of the Discussion, reference [101] should be [103], reference [103] should be [104], and reference [104] should be [105].This has been corrected on the publisher's website."}
+{"text": "The signal pathway of actin remodeling, including LIM-kinase 1 (LIMK1) and its substrate cofilin, regulates multiple processes in neurons of vertebrates and invertebrates. Drosophila melanogaster is widely used as a model object for studying mechanisms of memory formation, storage, retrieval and forgetting. Previously, active forgetting in Drosophila was investigated in the standard Pavlovian olfactory conditioning paradigm. The role of specific dopaminergic neurons (DAN) and components of the actin remodeling pathway in different forms of forgetting was shown. In our research, we investigated the role of LIMK1 in Drosophila memory and forgetting in the conditioned courtship suppression paradigm (CCSP). In the Drosophila brain, LIMK1 and p-cofilin levels appeared to be low in specific neuropil structures, including the mushroom body (MB) lobes and the central complex. At the same time, LIMK1 was observed in cell bodies, such as DAN clusters regulating memory formation in CCSP. We applied GAL4 \u00d7 UAS binary system to induce limk1 RNA interference in different types of neurons. The hybrid strain with limk1 interference in MB lobes and glia showed an increase in 3-h short-term memory (STM), without significant effects on long-term memory. limk1 interference in cholinergic neurons (CHN) impaired STM, while its interference in DAN and serotoninergic neurons (SRN) also dramatically impaired the flies\u2019 learning ability. By contrast, limk1 interference in fruitless neurons (FRN) resulted in increased 15\u201360 min STM, indicating a possible LIMK1 role in active forgetting. Males with limk1 interference in CHN and FRN also showed the opposite trends of courtship song parameters changes. Thus, LIMK1 effects on the Drosophila male memory and courtship song appeared to depend on the neuronal type or brain structure. Memory formation and forgetting serve as the basis of behavioralplasticity. Whereas memory is a specific process ofinformation acquisition, storage and retrieval by the nervoussystem, active forgetting is defined as \u201ca mechanism or seriesof mechanisms to remove memories that become unused\u201d. Associative memory formation andactive forgetting occur in both mammals and invertebrates,including Drosophila melanogaster , whichis a well-known object of classical genetics. Having a shortlife cycle and relatively simple nervous system, the fruit flymakes it easy to perform genetic analysis of the molecularbasis of behavioral and cognitive processes.There are several experimental techniques to form associativememory in Drosophila, including short-term memory(STM) and protein synthesis-dependent long-term me-mory(LTM). The most widely used technique is classical Pavlovianlearning with negative electroshock reinforcement,or olfactory aversive learning (OAVL), which revealed genesresponsible for different types of memory .More natural is conditioned courtship suppression paradigm(CCSP) .GAL4 \u00d7 UAS binary expression system  is usedto study the effects of specific genes on memory processes.The fine neural organization of the mushroom bodies (MB),a principle structure responsible for associative olfactorylearning in Drosophila, was evaluated in detail. The MB outputneurons (MBON) are the main effectors of MB, whereasspecific clusters of dopaminergic neurons (DAN) regulate theactivity of MB \u2013 MBON synaptic contacts . Among them are aSP13 DAN of the protocerebralanterior medial cluster (PAM), which innervate \u03b35 area ofMB, playing a crucial role in CCSP learning and memory.The molecular and neural mechanisms of active forgettingimplicate the activity of DAN and Rac1-dependent signalpathways . Small GTPases of the Rho family,including Rho and Rac, regulate neuronal actin polymerizationduring the Drosophila nervous system development. Rho viaits effector ROCK or Rac/Cdc42 via its effector Pak activateLIM-kinase 1 (LIMK1), which phosphorylates Drosophilacofilin (twinstar) protein, blocking its actin-depolymerizationactivity and inhibiting axon growth. Rac also acts through Pakindependentpathway to antagonize LIMK1 and promote axongrowth . In addition to its role in neurogenesis,Rac is crucial for both interference-induced and passiveforgetting in OAVL paradigm. PAK/LIMK1/cofilin pathwayprobably acts downstream Rac1 . Forgettingspecific types of memory depends on different signal proteins.Forgetting in OAVL paradigm is caused by several DANof the protocerebral posterior lateral 1 (PPL1) cluster, whichinnervates some MB structures, such as pedunculus, lowerand upper stalk. Memory acquisition and forgetting are regulatedby different dopamine receptors, dDA1 and DAMBrespectively . Coincidence of conditionedand unconditioned stimuli creates a memory trace in MBON-\u03b32\u03b1\u02b91, probably inhibiting the MB > MBON-\u03b32\u03b1\u02b91 synapses.The unconditioned stimulus alone activates DAN-\u03b32\u03b1\u02b91,which in turn disinhibit MB > MBON-\u03b32\u03b1\u02b91 synapses andcause forgetting . DAN that innervate theMB \u03b1\u03b1\u02b9 tip induce the interference-based forgetting throughthe scaffold protein Scribble, binding together Rac1, PAK3and cofilin .Whereas multiple data prove the importance of DAN andactin-remodeling signal pathway for forgetting in OAVL paradigm,there is virtually no data for molecular mechanisms ofmemory decay in CCSP. Effects of LIMK1-dependent signalcascade on CCSP learning and memory were firstly shown forthe temperature-sensitive mutant agn ts3, with LIMK1 increasein the adult brain compared to the wild type Canton-S (CS).Temperature rise leads to a decrease in agn ts3 LIMK1 level,simultaneously restoring its learning ability and 3 h memory,which are drastically impaired in the norm . agn ts3 has multiple polymorphisms within and nearlimk1 gene, as well as a changed profile of microRNA expression,and can serve as a model object for Williams syndrome. Thetemporal profile of STM learning index (LI) was assayedin CCSP for agn ts3, as well as for the wild-type strains withlimk1 polymorphisms, CS and Oregon R. Only CS was able tolearn and store memory up to 24 h .The behavioral effects of LIMK1 changes in agn ts3 do notgive information about specific cell types, where LIMK1 canbe involved in learning and memory. In this study, we performedthe analysis of memory decay for several DrosophilaGAL4 \u00d7 UAS strains with neuronal type-specific limk1 RNAinterference. LIMK1 distribution in the Drosophila brainstructures was studied in detail using confocal microscopy. Theeffect of limk1 interference on fly memory ability dependedon both neural type and memory form. LIMK1 also appearedto be involved in regulation of male courtship song: limk1interference in different neuronal types specifically affectedsome song parameters.Drosophila strains. Fly strains were provided by the ResearchCenter \u201cBiocollection of Pavlov Institute of Physiology RASfor the study of integrative mechanisms of the nervous andvisceral systems\u201d. The strain numbers (#) are given in accordancewith the Research Center and Bloomington DrosophilaStock Center, USA . The following strainswere used:1. \u0421anton-S (\u0421S) \u2013 the wild-type strain with limk1 polymorphisms.2. agn ts3 \u2013 temperature-sensitive mutation on CS geneticbackgroundwith limk1 polymorphisms, characterized bylearning and memory defects.3. Strains expressing GAL4 in specific neuronal types:#6794: w[*]; P{w[+mC]=nrv2-GAL4.S}8 P{w[+mC]=UAS-GFP.S65T}eg[T10]. GAL4 and green fluorescentprotein (GFP) are expressed in nervous system under Nrv2regulatory element;#6793: w[*]; P{w[+mC]=ChAT-GAL4.7.4}19BP{w[+mC]=UAS-GFP.S65T}Myo31DF[T2]. GAL4 andGFP are expressed in cholinergic neurons (CHN) underChAT and VAChT regulatory elements;#7009: w[1118]; P{w[+mC]=Ddc-GAL4.L}Lmpt[4.36].GAL4 is expressed in dopaminergic (DAN) and serotoninergic(SRN) neurons;#30027: w[1118]; P{w[+mW.hs]=GawB}fru[NP0021].GAL4 is expressed in fruitless neurons regulating matingbehavior.4. Act-GAL4: w[1118]; P{w[+mC]=} 25FO1/CyO, y[+];Canton-S background. GAL4 is expressed in the wholebody under actin promoter.5. Strains with UAS-dependent limk1 suppression:#26294: y[1] v[1]; P{y[+t7.7] v[+t1.8]=TRiP.JF02063}attP2. The strain expresses interfering RNA against limk1(RNAi) under UAS .#36303: y[1] v[1]; P{y[+t7.7]=CaryP}attP2. The controlstrain with genetic background identic to that for #26294,but lacking RNAi (limk1\u201c+\u201d).6. Strains with GFP gene regulated by UAS:#32186: w[*]; P{y[+t7.7] w[+mC]=10XUAS-IVS-mCD8::GFP}attP40.#32202: w[*]; P{y[+t7.7] w[+mC]=10XUAS-IVS-GFPWPRE}attP2.To induce limk1 RNA interference in specific neuronaltypes, a strain carrying GAL4 activator expressed under tissue-specific promoter was crossed to UAS strain #26294. The\u0441ross product of a GAL4 strain and #36303 strain served asa control.Flies were raised on the standard yeast\u2013raisin medium at25 \u00b1 0.5 \u00b0C and a 12:12 daily illumination cycle. For behavioraltests, experimental males were collected without anesthesiaand kept individually. 5\u20136-day-old males were used in experiments.Females (CS) were collected as virgins and broughttogether with CS males for fertilization in CCSP one daybefore experiment.Antibodies. Primary antibodies: Rat anti-LIMK1 multi-specificmonoclonal ; mouse anti-Drosophila cysteine string protein (CSP);rabbit anti-Drosophila tyrosine hydroxylase (TH) ; rabbit anti-GFP .Secondary antibodies: Goat anti-mouse Alexa Fluor 488, donkey-anti-rat Alexa Fluor 594, goat anti-rabbit AlexaFluor 633 .RNA extraction and RT-PCR analysis of limk1 expression.The level of limk1 expression was assayed usingsemi-quantitative PCR in complex with reverse transcription(RT-PCR). Flies were anesthetized by freezing. 10 male fliesor 70 male heads were homogenized in 300 \u03bcl TRI reagent. Total RNA was extracted from homogenatesaccording to the manufacturer\u2019s protocol. The quality ofRNA was checked by 1.5 % agarose gel electrophoresis. 1 \u03bcgRNA was reverse-transcribed by MMLV reverse transcriptase according to the manufacturers\u2019 protocol,using random hexamer primers and RNAse inhibitor . Semi-quantitative PCR was performed on a StepOnePlus  using qPCRmixHSSYBR+LowROX  containing direct andreverse primers (0.5 mM each). Baseline and cycle thresholdvalues were determined by automatic analysis using StepOnesoftware v2.3 . rpl32 transcriptwas used as an internal control. The predesigned limk1 primers were used to bind all five limk1 cDNA isoforms,both premature and mature forms, as primers do notspan the exon-intron borders. The relative limk1 transcriptlevel was calculated using the comparative \u0394\u0394Ct method. Thenumber of biological replicates (independent RNA extractionswith reverse transcription) was 3\u20135, the number of technicalreplicates was 3.The primer sequences were the following:rpl32:Forward: 5\u2032-TATGCTAAGCTGTCGCACAAATGGC-3\u2032Reverse: 5\u2032-GTTCTGCATGAGCAGGACCTCCA-3\u2032limk1:Forward: 5\u2032-GTGAACGGCACACCAGTTAGT-3\u2032Reverse: 5\u2032-ACTTGCACCGGATCATGCTC-3\u2032PCR parameters:1. 1 cycle: 95 \u00b0\u0421 \u2013 5 min.2. 45 cycles. 95 \u00b0\u0421 \u2013 20 s, 60 \u00b0\u0421 \u2013 20 s, 72 \u00b0\u0421 \u2013 20 s, 77 \u00b0\u0421 \u201315 s (detection).3. Melting curve: 95 \u00b0\u0421 \u2013 15 s, 60 \u00b0\u0421 \u2013 1 min, 60\u201395 \u00b0\u0421.Immunofluorescent staining of Drosophila brains.5\u20136-day-old imago males were anesthetized by freezing. Thebrains were prepared in PBS buffer (pH 7.5) using needlesharptweezers , fixed in 4 % paraformaldehydein PBS for 1 h at RT and stained according to , without a freezing stage. Antibodies were dilutedin PBT  as 1:200, foranti-CSP \u2013 1:20. Previously, for better staining of brains, weincreased the time of incubation with primary antibodies upto 5 days . Here, the incubation wasperformed at 4 \u00b0\u0421 for 3 days (with primary antibodies) orovernight (with secondary antibodies). Brains were mountedwith Vectashield mounting medium containing DAPI .Protein distribution analysis in the brain by confocalmicroscopy. Brains were scanned frontally using laser scanningconfocal microscopy .Scanning was performed using X63 objective at differentdepths (z-step 2 \u03bcm). Images were analyzed using Fiji software.The brain structures were visually mapped using theDrosophila brain online atlas . To measurethe average level of LIMK1 inside the brain structures, theaverage signal intensity was measured in three small squareareas (~10 \u00d7 10 \u03bcm) within each of the structures. The averagevalues were obtained and normalized to the average structureintensity for the given brain. Colocalization Threshold analysis was performed to measure co-localization of LIMK1 withneurospecific markers. To prepare figures, auto contrast functionwas used for each optical slice.Learning and courtship suppression tests in Drosophilamales. Flies learning and STM were estimated in CCSP, asdescribed in . In the case of long termmemory (LTM), learning was performed by placing flies infood-containing glasses  for5 h . Courtship index (CI) and learningindex (LI) were estimated at the following time pointsafter learning: for short-term memory (STM) analysis: 0 min(learning), 3 h; for STM decay analysis: 15, 30, 60 min,24 h; for LTM analysis: 0 min, 2 days, 8 days. In all groups,naive males (without mating experience) served as a controlto calculate LI:where CIN is the middle CI for naive males, and CIT is themiddle CI for males after training. The naive and trainedmales were the same age. The decrease in LI compared toLI (0 min) was considered a time-dependent memory decay.The decrease in LI for a mutant strain compared to that for thewild-type strain CS was considered a strain-specific impairmentof learning or memory.Courtship song analysis. The 5-day-old imago malecourtship song was recorded as in . A naive male of the studied line and a fertilizedfemale (CS) were placed together in a Perplex chamber witha latticed bottom on top of a microphone. The chamber wasplaced in a foam box in a soundproof room. The sounds wererecorded for 5 min using Audacity software . The sound signals were filtered to excludenoises, obtaining signals within 100\u2013800 Hz. The level ofnoise was decreased using a standard Audacity plugin. Thesoftware Drosophila courtship song analysis (DCSA)  was used to automatically detect pulse and sinesong components.The results of analysis were manually edited. The meanvalues of the song parameters were calculated for each fly.The following parameters were estimated: pulse song index, pulse song initiation frequency, sine song index , sinesong frequency , interpulse interval ,period of song pulse train , intertrain interval ,train duration (TrainDur), pulse number in train (PulseN), sinesong duration , sine song amplitude ,IPI variance (Var(IPI), ms2). Per is the time between the startsof the neighboring trains. ITI is the time between the end ofthe previous and the start of the next train.Statistical analysis. Analysis of LIMK1 mRNA level wasperformed using two-sided t-test, Social Science Statisticonline resource ( p < 0.05). Analysis of LI and courtship songparameters was performed using two-sided randomizationtest at significance level \u03b1 of 0.05 (n = 20), using DrosophilaCourtship Lite software , with 10000 iterations. The program is freelyavailable from the author upon request. Randomization testwas reported to be better for LI comparison than t-test orsome nonparametric tests . Courtshipsong parameters were also analyzed using two-sidedMann\u2013Whitney U-test. Python 3 scripts were used to drawthe box plots charts.limk1 RNA level in Drosophila UAS \u00d7 GAL4 hybridsTo check that GAL4 really induces limk1 RNA interferencein 26294 strain, we compared limk1 RNA level in the UAS (f)> GAL4 (m) hybrids. Females with and without transgenicRNAi for limk1 suppression were crossed to Act-GAL4 males, expressingGAL4 in the whole body. The level of total limk1 RNA wasapproximately 2-fold lower in the hybrid with limk1 interference.These data confirmed the efficiency of RNAi-dependentlimk1 suppression in 26294 strain (limk1-KD) upon its activationby GAL4. At the same time, there were no differences forlimk1-KD > 6794 and limk1\u201c+\u201d > 6794, where RNA expressionwas measured in heads and was regulated by neuronaltype-specific GAL4 . Thus, limk1 RNA differencesafter neural type-specific limk1 RNA interference might belocal or too low to be detected in the whole Drosophila heads.LIMK1 distribution in the Drosophila brainWhen studying LIMK1 distribution, we focused on thecentralpart of the Drosophila brain, without the optic lobes(OL), mainly at the level of the superior medial protocerebrum(SMP) and gamma-lobes (\u03b3L) of MB. Here the PAMclusters of DAN are located , responsiblefor the Drosophila courtship learning and memory . Additionally, the area including the central complex(CC) and calyx  surrounded by Kenyon cells (KC)was studied. The CSP-positive neuropil structures and tyrosinehydroxylase (TH)-positive DAN cell bodies and processesserved as landmarks in describing the LIMK1 distribution.The following description is given for the wild-type strainCS 1.https://vavilovj-icg.ru/download/pict-2023-27/appx9.pdfSupplementary Materials are available in the online version of the paper:The distribution of DAN clusters corresponded to thatdescribed in . PAM clusters were clearlyobserved near SMP, with processes extending towards thecentral part of the brain. The processes formed glomerularstructures around the MB horizontal lobes , probablybeing the synaptic endings innervating the correspondareas. The structure #3 was located above \u03b3L, the structures#5 and 8 \u2013 in the \u03b2\u2032L area, the commissure #9 was seen in thecentral part of the brain. TH signal was relatively low in \u03b2L. PPL1, PPM2 and other DAN clusterswere observed around Cal, sending their processes to the differentbrain areas .LIMK1 was concentrated in the neuropil structures of theanterior part of the brain, such as SMP and AL. The LIMK1-positive granularity was observed inside SMP, between the \u03b2\u2032Ltips (#8) and around the ellipsoid body (EB) of CC .LIMK1 level was also high in thin tissue layers adjacentto neuropil and some neural tracts, such as #12 around thegreat commissure, #13 around wedge (W) and #15 near esophagus(ES), morphologically resembling glia . LIMK1 signal was lower in cell bodies of the neuronssurrounding AL (ALCB), probably being the cell bodies of theprojection neurons, as well as in KC surrounding Cal. Here,LIMK1 was mainly concentrated in the cytoplasm, beyondthe nuclei. LIMK1 level was significantly decreased in all theMB lobes and pedunculus (Ped), as well as in the CC structures,whereas in Cal and the protocerebral bridge (PB) it wasrelatively high . LIMK1 and TH colocalizationwas observed in SMP, AL, Cal, the TH-positivecells and processes, and in glomerular densities, such as #3,5 and 6 .To check that the antibody specifically binds to LIMK1, thedistribution of LIMK1 main product p-cofilin was assayed inCS. The pattern of p-cofilin distribution was generally similarto that for LIMK1 . The level of p-cofilinwas low in MB  and CC . In contrast to LIMK1, p-cofilin wasmainly concentrated in the cell nuclei in the peripheral area ofthe fly brain, such as Kenyon cells around Cal, as well as in PB,subesophageal ganglion (SEG), and cell bodies surrounding AL. p-Cofilin was also localized in diffuse layers within thebrain structures, such as EB, probably formed by glia. Thep- cofilin-enriched cells were found in SEG, forming structureswith two glomerular branches (*), and around CC structures,fan-shaped body (FB) and EB (**).Several GAL4 activators were used to initiate limk1 RNAinterference. Both 6793 and 6794 strains specifically expressthe green fluorescent protein (GFP) under GAL4 promoter.In strain 6794, GAL4 was reported to be expressed in OL,thoracic ganglion, different nerves, and cortex glia . In 6794 > limk1-KDhybrid, GAL4-driven GFP expression was detected in glialikecells, surrounding the neuropil structures, such as AL,SMP, CC and its parts, as well as in the MB lobes, Ped andsome KC . GFP level was higher in \u03b1Land \u03b2L compared to \u03b1\u2032L, \u03b2\u2032L and \u03b3L. The signal was lowerin Cal and virtually absent in most neuropil structures, suchas AL, SMP, CC and others. Thus, limk1 interference shouldoccur in Cal and some glia cells, where the levels of bothGAL4 and LIMK1 were relatively high. Similar distributionwas observed in the control 6794 > limk1\u201c+\u201d strain . In the strain 6793, GAL4 is expressedin cholinergic neurons (CHN), with GFP signal in OL, ALwith the surrounding interneurons, the parts of CC, the greatcommissure (GC), Cal and the mechanosensory area of SEG. In the strains 6793 > limk1- KDand 6793 > limk1\u201c+\u201d , we observeda strong GFP signal in cell bodies surrounding SMPand AL, as well as in some KC, several neuropil structures, and GC. In all the studied strains, LIMK1distribution character appeared to be similar to that in CS.To check that GAL4 is active in 7009 and 30027 strains,we crossed them to strains expressing GFP under GAL4 promoter.In 7009 > 32186 hybrid, we observed a prominent GFPsignal in cell clusters near SMP, morphologically similar tothe TH-positive PAM clusters. Some cells might be SRN, butthey constitute the minority of the observed neurons in thisarea . The processesof PAM neurons extended to the horizontal MB lobes, including\u03b3L, and the densely innervated \u03b2\u2032L tip (#5) connected by a commissure, and to a much lesser extent the \u03b2L tip . EB was surrounded by the GFP-positive processesextending from different parts of the brain. The GFPpositiveDAN around Cal were also observed. Hence, GAL4activator of 7009 should suppress limk1 inside DAN, includingPAM neurons, which regulate memory storage in CCSP. Thefruitless-positive neurons (FRN) are responsible for matingbehavior. In 30027 > 32202 hybrid, we observed GFP insome KC, in the cell bodies located near SMP and AL, andglomerular structure, forming a ring-like structure around Ped. Similar structures were describedin . The distribution of LIMK1 in the hybridstrains with and without limk1 knockdown in the above neuronswas similar to that in CS .The normalized intensity of LIMK1 signal was calculatedfor several brain structures . The LIMK1 relative levelsin specific structures were very similar for the CS brain andthe average brain of all the strains. The biggest differenceswere observed for the TH-positive glomerular structure #6(TH+(6)), which is possibly responsible for memory formationin CCSP. In the average brain, ALCB had the normalizedLIMK1 level about 1. Compared to them, AL, SMP andTH+(6) structures had the higher LIMK1 level, whereas theMB lobes, EB and Ped had the lower LIMK1 level. In agn ts3,AL and ALCB had the higher LIMK1 level compared to CS,whereas most of the rest studies structures had lower LIMK1level. This corresponds to more contrast LIMK1 staining inagn ts3 relative to CS . There were no prominentdifferences after limk1 knockout, except for severalstructures with minor changes. The interstrain differencesmight be local or beyond the resolution of the method.3 h STM differs in hybridswith and without limk1 interference3 h STM was estimated for limk1-KD (f ) > 6794 (m) and thecontrol limk1\u201c+\u201d (f ) > 6794 (m) hybrids. In both cases, weobserved the decrease of courtship index (CI) after learning,with its partial recovery after 3 h. The box plot height wasminimal for CS and rather big for UAS \u00d7 GAL4 hybrids,showing that the value of courtship suppression significantlyvaried for individual flies. All strains were capable to learn inCCSP, with learning index LI (0 h) immediately after trainingof about 60\u201370 % . The CS LI was still high after 3 h,indicating STM preservation, in agreement with . The strain with limk1 interference also preservedSTM: although its LI (3 h) was only about 20 %, it did notstatistically differ from that for CS, as well as from LI (0 h). Inthe control strain, LI (3 h) decreased compared to LI (0 h) anddid not differ from zero, indicating the impaired STM. Thus,while 3 h memory storage or retrieval was impaired in thecontrol strain, limk1 interference seems to improve 3 h STM.At the same time, it did not affect the impaired 8 day LTM,with only minor positive effect on 2 day LTM .Neuron type-specific limk1 interferencedifferentially affects STM dynamicsTo investigate the dynamics of STM decay in different strainswith limk1 interference, we performed LI analysis immediatelyand 24 h after learning . To exclude the possible effectof eye color on learning and memory abilities, we appliedGAL4 (f ) > UAS (m) crossing scheme, where both the strainwith limk1 knockdown and the control strain had the samewild-type eye color. For 6794 activator (MB and glia), thecontrol strain showed nearly the same LI within 24 h, whereasthe strain with limk1 interference demonstrated a steeperforgetting curve. Hence, 6794 > limk1-KD showed high LIafter learning, but seemed to increase the speed of memoryforgetting on the interval 0\u201330 min. limk1 knockdown in CHN(6793) was associated with significant decrease of LI within60 min after training.For DAN and SRN activator (7009), both the strain withlimk1 interference and the control strain showed nearly thesame dynamics of STM decay, except for 30\u201360 min period.limk1 interference was associated with a dramatic defect onlearning: LI did not differ from zero. LI (24 h) was negativein both hybrids, possibly being the effect of sensitization:males did not suppress the courtship activity but courted moreactively some time after training. For FRN activator (30027),the effect of limk1 knockdown was the opposite to that ofMB/glia and CHN activators: limk1 interference decreasedthe speed of forgetting, and LI (30 min) did not differ fromzero. Thus, the effect of limk1 interference on STM dynamicsappeared to depend on the neuronal type.LIMK1 interference in CHN and FRN neuronsdifferentially affects courtship song parametersFinally, we studied the influence of limk1 interference on themale courtship song parameters. The hybrids with CHN andFRN drivers were studied . There were no interstraindifferences in interpulse interval (IPI), the species- and population-specific parameter , and IPI variance(Var(IPI)), the marker of neurodegenerative processes. limk1 interference in CHN(6793) decreased the pulse song index and frequency , increasing the mean period (Per), intertrain interval (ITI),train duration (TrainDur), sine song duration (Sdur) and trainpulse number (PulseN). On the contrary, in the strain with FRNactivator (30027), limk1 interference resulted in PFr increase,as well as Per, ITI, SInd and SDur decrease. limk1 knockdownby two different activators had the opposite effects on PInd,PFr, Per, ITI and SDur, leveling the initial differences betweenSInd, TrainDur and PulseN. Thus, limk1 interference in CHNseemed to decrease the rate of switching from the singingmode to silence mode and back, resulting in longer trains andITI, while limk1 interference in FRN neurons generally hadthe opposite effect.In mammals, LIMK1- and cofilin-dependent actin remodelingis widely involved in regulation of synaptic processes,such as exocytosis, receptor trafficking and remodeling ofdendritic spines. These processes underlay long-term potentiation(LTP), long-term depression (LTD) and different forms ofmemory. LIMK1 also affects gene expression through CREBand LTM formation. Deregulation of LIMK1-dependentactin remodeling is involved in multiple pathologies, such asAlzheimer\u2019s and Parkinson\u2019s diseases, Williams syndrome,schizophrenia, and autism .In mature neurons, actin is enriched in both pre- and postsynapticstructures, such as dendritic spines, regulated byRho signaling pathway. The action of LIMK1 on actin polymerizationand memory processes is rather complex, beingdependent on the mode of LIMK1 regulation (transient orlong-term overexpression) and cofilin level. While the activecofilin destabilizes fibrillar actin, in high concentrations itincreases actin polymerization and nucleates actin filamentsin dendritic spines during long-term potentiation . Thus, it is hard to predict the integral effect of LIMK1 and cofilin activity on memory processes.It was crucial to check the behavioral effects of Drosophilalimk1 suppression in specific types of neurons.Using paraffin section staining, both LIMK1 and its productp-cofilin were shown to be homogeneously distributed in thebrain neuropil, with maximum level in CC . Our results of the confocal microscopy analysis gavea different picture of LIMK1 distribution, quite similar for allthe studied strains. We have shown a specific LIMK1 decreasein MB, which is responsible for associative learning, as well asin CC, involved in higher movement control . This puts under question the role of LIMK1 inthe aforementioned processes. However, LIMK1 was presentin the cell bodies and processes of DAN, which interact withMB and CC  and regulate memory andforgetting. The observed p-cofilin distribution resembled thisfor LIMK1: its level was low in the MB lobes and CC. The lowlevel of p-cofilin in the MB lobes had been previously shown . In contrast to LIMK1, p-cofilin level waslow in Cal formed by PN and KC terminals and high in cellnuclei. The latter corresponds to its functioning in the cell, ascofilin phosphorylation is necessary for its translocation intothe nucleus .The effectiveness of limk1 suppression at the RNA level wasconfirmed using Act-GAL4 activator in the whole Drosophilabody. GAL4 was also active in specific brain areas of the correspondingstrains. However, we failed to quantitatively checkthe changes of limk1 expression in Drosophila UAS \u00d7 GAL4hybrids with neuronal-specific GAL4 expression. The decreasein LIMK1 level might be local or too small. limk1interference might also induce the compensatory activationof LIMK1 translation.To study limk1 knockdown effects on memory, we usedCCSP modification applied by :training was performed with the mated female. In this case,courtship learning results from the rise of sensitivity to theantiaphrodisiac cis-vaccenyl acetate (cVA) due to unsuccessfulcourtship. cVA is not required for learning, being necessaryfor memory performance. aSP13 DAN, which innervate thefru-positive tip of \u03b3L, are necessary and sufficient for courtshiplearning . 24 h memory consolidationrequires the prolonged aSP13 stimulation and Orb2 dimerizationin some \u03b3 neurons . \u03b1/\u03b2 neurons areinvolved in LTM processes . Hence, other DAN innervating \u03b1/\u03b2L of MB,including PAM and PPL1 cells , may alsobe involved in LTM.The behavioral differences were observed after limk1 interference,e. g., the restoration of 3 h STM for limk1-KD >6794 strain. GAL4 drivers themselves affected memoryabilities,which were generally decreased compared to CS.The drivers also significantly affected the forgetting curves.Thus, we studied the effects of limk1 interference relative toa control strain with the same GAL4 driver. We applied twodifferent crossing schemes \u2013 UAS (f) > 6794 (m)  and reverse . In the first case, the control UAS > limk1\u201c+\u201dhybrid had bright eyes due to v[1] recessive allele, in contrastto UAS > limk1-KD hybrid with the wild type dark red eyes.The observed 3 h STM differences are unlikely to be associatedwith the differences in eyes pigmentation, as v[1] fliesshowed a normal 3 h STM and 2 day LTM in CCSP , while both forms of memory were impaired in thecontrol strain. However, memory retention depends on parentaffect, with some paternal epigenetic factors affecting STMstrength . For 6794 > limk1-KD strain,we did not see STM difference from the control strain, thoughlearning ability slightly increased after limk1 knockdown . Thus, when studying LIMK1 effects on learning andmemory, it is necessary to consider the crossing direction.Acetylcholine is the major excitatory neurotransmitter inDrosophila. Among CNH are: PN forming synapses on KCof MB , the MB intrinsic neurons thatare responsible for olfactory memory, expressing ChAT andVAChT , and the \u03b1/\u03b2 core neurons requiredfor LTM consolidation . In the hybridswith 6793 driver (GAL4 expressed in CHN), GFP level wasspecifically high in \u03b1/\u03b2L compared to the other MB lobes.Here, limk1 interference resulted in faster STM forgetting.This contradicts the cofilin role in active forgetting shown inOAVL, where cofilin was proposed to be regulated by LIMK1. The involvement of LIMK1 and cofilinin forgetting may occur locally, within specific neuronalpopulations or synaptic terminals. At the same time, LIMK1may be crucial for memory storage and retrieval in CCSP.The glutamatergic MBON M6 neurons serve for STM output:aSP13 DAN prolongs potentiation of \u03b3L \u2013 M6 synapses . Some cholinergic MBON appeared to regulatethe Drosophila visual appetitive memory .As the extrinsic MB cells responsible for CCSP memory weresimilar to those used for appetitive memory , the decrease in 60 min STM might occur due to limk1suppression in some of these neurons.The hybrids with 7009 driver (DAN and SRN) showedgenerally low CI values and negative LI values 24 h afterlearning. Males of these strains had pale pink eyes because ofdefects of eyes pigmentation, due to non-complete w[1118]rescue. w[1118] males demonstrated low courtship activityand success, presumably due to some defects of sexual developmentand maturation . However, thecontrol 7009 > limk1\u201c+\u201d strain had normal LI up to 60 minafter training. limk1 knockdown by 7009 driver was associatedwith dramatic defects of learning and memory: LI just aftertraining did not statistically differ from zero. Thus, LIMK1-dependent signaling in DAN and SRN seems to be importantfor learning and memory in CCSP.limk1 knockdown by 30027 driver (FRN) decreased theforgetting rate in the time interval 30\u201360 min. This correspondsto the role of actin-remodeling pathway in forgettingin OAVL paradigm . LI of the controlstrain did not differ from zero starting from 30 min afterlearning, while limk1 knockdown increased it. In males, FRNare responsible for courtship behavior. There are ~1500 FRNin the Drosophila brain, including sensory organs, lateralhorn, lateral protocerebrum, SMP arch and motor controlcenters. Together they provide multisensory integration toregulate the male courtship process . Some CHN and DAN are also Fru-positive, suchas ~300 \u03b3L neurons and aSP13 DAN located in SMP, whichregulates courtship learning and memory. The activity of frugene was reported to decrease upon LTM formation in CCSP. Hence, suppression of some FRN activitymay be associated with memory prolongation and consolidation.In addition to memory processes, limk1 interference affectedsome parameters of the male courtship song. As well as forcourtship memory, we observed the opposite functional effectsof limk1 knockdown in FRN and CHN. FRN of the P1 classinitiate Drosophila courtship behavior and trigger courtshipsong. pIP10 neurons possibly convey the P1 signal to thoracicdPR1 and vPR6 neurons, proposed to be the parts of a centralpattern generator (CPG), which defines the time and shape ofthe pulse song. vPR6 possibly encode IPI . Pulse and sine CPG either contain FRN or interactwith them. As sine and pulse song normally do not overlap,the mutually inhibitory mechanisms must exist, switching between quasilinear and relaxation modes of oscillation forsine and pulse song, respectively. Some descending interneuronsmay control the type of the song, while the others triggersinging or terminate the song .Indeed, we observed the opposite changes of PInd/PFr andSInd/SDur upon limk1 interference in CHN and FRN, movingthe balance toward the sine and pulse song, respectively. Theincrease in PFr after limk1 knockdown in FRN might indicatethe negative role of LIMK1-dependent signaling on activityof the pulse CPG or the upstream brain centers, which switchthem from active to silent mode. \u0421\u0421 is important for control ofstability of pacemakers, which regulate the rhythmic structureof courtship song. PB destruction leads to sound signal distortions,FB and EB destruction additionally decreases sine andpulse trains . CC includes a large numberof neuronal types, such as CHN, DAN, SRN, and others. CHNare present in FB, EB, No and PB ,similarly to what we observed in our research. Hence, theyprobably play some role in regulation of male singing. Theopposite effects of limk1 interference in CHN and FRN mayindicate a specific role of LIMK1 in courtship controllingnetwork, whereas the other parts of the brain possibly havea total antagonistic effects on its activity. Alternatively, CHNand FRN fru neurons may differ in some aspects of regulationof LIMK1-dependent signaling pathway.In summary, we have shown that effects of limk1 interferencein Drosophila male courtship memory and song depend onboth the neuronal type and specific behavioral parameter.limk1 interference in CHN and FRN had generally oppositeeffects, whereas its suppression in DAN and SRN impaired theflies\u2019 ability to learn. Using activator strains with a narrowerpattern of GAL4 expression would help to better localize thebrain structures, where LIMK1 regulates memory and forgettingin CCSP. 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+{"text": "To date, etravirine (ETR) is the only NNRTI indicated in people living with HIV (PLHIV) after virologic failure (VF) to first-generation NNRTIs. Data about susceptibility to ETR, doravirine (DOR) and rilpivirine (RPV) after VF in PLHIV from Latin American countries are scarce. The aim of this study is to determine the prevalence of resistance to ETR, DOR, and RPV in PLHIV who experienced VF to a first-line NNRTI-based regimen, and factors associated with high-level resistance (HLR) to ETR.Retrospective cohort study. Prevalence of resistance to ETR, DOR and RPV was assessed by analyzing genotyping tests of adult PLHIV who experienced VF to a first-line NNRTI-based regimen, in Buenos Aires, Argentina (2010 to 2020). Stanford HIVdb v9.4 was used to interpret resistance profiles. We defined resistance as both intermediate resistance (IR) and HLR interpretations. A sample size of 125 subjects was calculated to allow estimation of the primary endpoint with sufficient precision (95%CI). A cross-sectional analysis was carried out to identify risk factors associated with HLR to ETR.2=4.5, p=0.034).N=125; 81.6% were male. Median age at VF was 39.5 years (IQR: 34.0-35.0); 82,4% received efavirenz and 17.6% nevirapine. Duration of VF: 245 days (IQR 133-413). Median HIV-1 viral load (VL): 11255 c/mL (3606-45100); median CD4+ cell count: 192 cell/uL (86-333). Seventy-seven samples (61.6%) had ETR resistance-associated mutations (RAMs); 25 (20%) had mutations associated with the highest levels of reduced susceptibility . Most frequent ETR RAMS were: 100I (16%), 190A (16%) and 181C (12%); DOR RAMs: 100I (16%), 188L (8.8%) and 106M (4.8%), and RPV RAMs: 100I (16%), 181C (11.2%) and 138A (7.2%). Prevalence of resistance to ETR was 44.8% , resistance to DOR: 64%  and to RPV: 52% . ETR maintained susceptibility to DOR resistant strains in 27.2% of the cases. We found a statistically significant association between HIV-1 VL >10000 c/mL and the risk of developing HLR to ETR (XIn our cohort, resistance to ETR after VF to a NNRTI-containing regimen was lower than to DOR and RPV. HLR was uncommon and associated with high HIV-1 VL. Almost one third of DOR-resistant strains remained susceptible to ETR.Ver\u00f3nica Mingrone, GSK/ViiV: Educational Courses|Janssen Pharmaceutical Companies: Educational Courses Eliana Loiza, n/a, GSK/ViiV: Educational Courses|Janssen Pharmaceutical Companies: Educational Courses Norma Porteiro, n/a, GSK/ViiV: Advisor/Consultant|GSK/ViiV: Grant/Research Support|GSK/ViiV: Educational Courses|Janssen Pharmaceutical Companies: Advisor/Consultant|Janssen Pharmaceutical Companies: Grant/Research Support|Janssen Pharmaceutical Companies: Educational Courses Ezequiel C\u00f3rdova, n/a, GSK/ViiV: Advisor/Consultant|GSK/ViiV: Grant/Research Support|GSK/ViiV: Educational Courses|Janssen Pharmaceutical Companies: Advisor/Consultant|Janssen Pharmaceutical Companies: Grant/Research Support|Janssen Pharmaceutical Companies: Educational Courses"}
+{"text": "These results should be taken into consideration for the elderly patients under treatment.COVID-19 inactivated vaccine-induced humoral responses in patients with lung cancer (LCs) to SARS-CoV-2 wild-type (WT) strain and variants BA.4/5 after the primary 2-dose and booster vaccination remained unknown. We conducted a cross-sectional study in 260 LCs, 140 healthy controls (HC) and additional 40 LCs with serial samples by detecting total antibodies, IgG anti-RBD and neutralizing antibodies (NAb) toward WT and BA.4/5. SARS-CoV-2-specific antibody responses were augmented by the booster dose of inactivated vaccines in LCs, whereas they were lower than that in HCs. Enhanced humoral responses waned over time after triple injection, notably in NAb against WT and BA.4/5. The NAb against BA.4/5 was much lower than WT. Age\u2009\u2265\u200965 was risk factor for immunization of NAb to WT. Undergoing treatment resulted in a lower antibody response than those without and radiotherapy was a also risk factor for seroconversion of NAb to WT. Lower lymphocyte counts contributed to a lower titer of IgG anti-RBD and NAb against BA.4/5 in LCs than HCs. Specifically, total B cells, CD4The online version contains supplementary material available at 10.1186/s13045-023-01443-3. Since SARS-CoV-2 spread all over the world, patients with lung cancer (LCs) had an estimated case fatality rate of more than 30%, compared with 0.7% to 8.0% in general population . As the In addition, the Omicron variant harboring 30\u201340 mutations in the viral spike protein produced high immune evasion . HomologP\u2009<\u20090.05) than the second in LCs. A durability of total antibodies was found   in LCs, but it decreased faster than HCs and the result showed it was 3.8-fold higher in HCs than LCs after 180\u00a0days post-booster vaccine (P\u2009=\u20090.0003)  in LCs   in LCs   and Omicron BA.4/5 in LCs (P\u2009=\u20090.0339). The lower immunization may be explained by lower number total B cells, CD4+T cells and CD8+T counts in LCs as their correlation to the humoral response (Additional file Moreover, our results suggested undergoing various anticancer therapies influenced the antibody response of post-booster vaccine. LCs received radiotherapy generated lower level of total antibodies than other therapeutic strategies post-booster inactivated vaccines. The IgG anti-RBD antibodies titer, NAbs against WT and Omicron BA.4/5 were significantly lower in patients receiving various therapies than those without (Fig.\u00a0Overall, our study revealed strengthened humoral responses post-booster vaccine among LCs, albeit lower than HCs. However, the booster dose failed to establish a potent and durable antibody response for Omicron BA.4/5, which gives rise to the risk of breakthrough infections of Omicron variants, especially in those old and undergoing anticancer therapies. Given the lower antibodies in LCs receiving various active anticancer therapies, further studies are needed to determine whether increased dosage, mixing vaccine types or additional doses enhance immunogenicity.Additional file 1: Methods.Additional file 2: Table S1. Matched demographics between 260 patients with LC and 140 HCs. Table S2. Antibody Response to SARS-CoV-2 inactivated Vaccine between 260 patients with LC and 140 HCs. Table S3. Demographics and clinical characterization of 40 patients with LC with sequential samples. Table S4. Comparative analysis of neutralizing effect responses to SARS-CoV-2 WT and Omicron variant BA.4/5 in 260 patients with LC. Table S5. Antibody response to SARS-CoV-2 inactivated vaccination in 260 LCs and 140 HCs aged\u2009<\u200965 and\u2009\u2265\u200965\u00a0years. Table S6. Antibody response to SARS-CoV-2 inactivated vaccination in 260 LCs receiving various treatment regimens. Table S7. Risk factors associated with seropositivity of SARS-CoV-2 antibodies in 144 LCs received booster vaccine.Additional file 3: Figure S1. SARS-CoV-2 antibodies response in 40 LC patients after the second or booster dose of inactivated vaccine. Total antibodies against SARS-CoV-2. Concentrations of IgG anti-RBD antibodies. Inhibition rates of NAb against SARS-CoV-2 WT. Inhibition rates of NAb against Omicron BA.4/5.Additional file 4: Figure S2. Comparison of neutralizing effect responses to SARS-CoV-2 WT and Omicron variant BA.4/5 in LCs. The figures show the median and quartiles. *P\u2009<\u20090.05, **P\u2009<\u20090.01, ***P\u2009<\u20090.001 and ****P\u2009<\u20090.0001. LC, lung cancer; WT, wild type.Additional file 5: Figure S3. Comparison of lymphocytes counts in 260 LCs and HCs. The figures show the median and quartiles. **P\u2009<\u20090.01.Additional file 6: Figure S4. Correlation of biological variables and magnitude of SARS-CoV-2 antibodies after the booster dose."}
+{"text": "The aim of this prospective multicentre, French observational study was to describe the conditions of Ceftolozane/Tazobactam (C/T) use in hospital settings, and outcomes. This sub-analysis focuses on the cystic fibrosis (CF) patients included in the overall cohort.Adult patients suffering from CF having received at least one dose of C/T and followed up as per routine clinical practice until stop of C/T were included in this analysis. Additional data related to CF were collected.Between October 2018 and December 2019, 63 patients with CF were enrolled from 28 sites. Mean age was 33.6 years and 44.4% were males. 12 patients (19.0%) received a lung transplant, 46.0% had comorbidities with diabetes (34.9%) being the most frequent. Most patients (69.8%) had normal renal function, almost one-third (28.6%) were immunosuppressed. One-half of patients presented with class 2 mutation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene. None of the patients received CFTR modulators due to non-availability during the study period.About one-half of patients (52.4%) experienced intolerance/allergy to antibiotics; of these, 78.1% was associated with Ceftazidime.Pseudomonas aeruginosa was the most predominant pathogen representing 89.9% (71/79) of the isolates. C/T showed a very high susceptibility rate of 91.5% across those strains (Table 1). C/T demonstrated higher susceptibility rates than other B-lactam agents (Table 2).Microbiology results showed that Treatment duration until complete cure had a median of 15 days and occurred in 44.4% of patients. Treatment duration until partial cure or end of treatment was a median of 15 days and occurred in 46% of patients (Table 3). Only 2 patients experienced an adverse event leading to a discontinuation of treatment.Treatment with C/T had a positive impact on pulmonary function measured by FEV1 (Table 4) with a mean increase after treatment from 1.33L (range 0.5 - 4.1L) to 1.47L (range 0.6 - 4.6L).Pseudomonas aeruginosa.These results suggest that C/T is an effective and safe option for the treatment of CF patients with a potential positive impact on FEV1 when treating bacterial infections mainly caused by Xavier Bourge, PharmD, Merck: Employe Brune Akrich, MD, MSD: Employee David Boutoille, MSD France: Board Member Isabelle Brassac, Scientist, MSD France: salary Carole Mackosso, n/a, MSD France: MSD employee Jean-Fran\u00e7ois Timsit, MD, merck: Advisor/Consultant Bernard Castan, MD, ADVANZ: SPEAKER MODERATOR|BIOMERIEUX: SPEAKER MODERATOR|GILEAD: Advisor/Consultant|MSD France: Board Member|MSD France: SPEAKER MODERATOR|SANOFI: Advisor/Consultant|SHIONOGI: Advisor/Consultant Pierre-Regis Burgel, MD,PHD, ASTRA-ZENECA: Honoraria|BOEHRINGER INGELHEIM: Honoraria|CHIESI: Honoraria|GSK: Honoraria|INSMED: Honoraria|NOVARTIS: Honoraria|PFIZER: Honoraria|VERTEX: Grant/Research Support|ZAMBON: Honoraria"}
+{"text": "Burkholderia cepacia complex (Bcc) and Burkholderia gladioli strains. The addition of taniborbactam to cefepime shifted cefepime minimum inhibitory concentrations toward the provisionally susceptible range in 59% of the isolates tested. Therefore, cefepime-taniborbactam possessed similar activity as first-line agents, ceftazidime and trimethoprim-sulfamethoxazole, supporting further development.The novel clinical-stage \u03b2-lactam-\u03b2-lactamase inhibitor combination, cefepime-taniborbactam, demonstrates promising activity toward many Gram-negative bacteria producing class A, B, C, and/or D \u03b2-lactamases. We tested this combination against a panel of 150  Pseudomonas aeruginosa, including strains producing VIM-2  producers (Taniborbactam (formerly VNRX-5133) is a novel bicyclic boronic-acid \u03b2-lactamase inhibitor  being deng VIM-2 \u201311, 13roducers , 14\u201317Burkholderia cepacia complex (Bcc) is a group of >20 related non-fermenting pathogens (Burkholderia gladioli can cause chronic infections in the immunocompromised and those with cystic fibrosis (CF) with 8%\u201310% of CF patients acquiring Bcc or B. gladioli by end stage, leading to increased morbidity and mortality  and AmpC (class C) \u03b2-lactamases  as well as \u03b2-lactam-\u03b2-lactamase inhibitor combinations   , while tic acid) . Severalin vitro , 28\u201330isolates , 28, 29.ambifaria, arboris, cenocepacia, cepacia, contaminans, diffusa, dolosa, multivorans, pseudomultivorans, pyrrocinia, seminalis, stabilis, ubonensis, and vietnamiensis) and 10 B. gladioli from the Burkholderia cepacia Research Laboratory and Repository (University of Michigan) . Minimum inhibitory concentration (MIC) results were interpreted using Clinical Laboratory Standards Institute (CLSI) breakpoints, where available (P. aeruginosa of susceptible (\u22648 \u00b5g/mL), intermediate (16 \u00b5g/mL), and resistant (>32 \u00b5g/mL)   , 25, 31 vailable . Cefepimsistant > \u00b5g/mL (3nd lung) \u201317 andnd lung) , 34. Theli panel , 32. Of 50 value decreased by fourfold from 32 to 8 \u00b5g/mL  shifted the MICs toward the provisionally susceptible range; the MICge panel . MoreoveblaPenA , 31. Poiocepacia . Alteratdomallei ; efflux  species .Burkholderia species with MIC50 of 4 \u00b5g/mL compared to 32 \u00b5g/mL for cefepime. The addition of 8 \u00b5g/mL of vaborbactam further increased the potency of meropenem with meropenem-vaborbactam yielding an MIC50 of 2 \u00b5g/mL. Levofloxacin performed similarly to cefepime alone with 26% vs 23% of isolates testing susceptible, respectively. Notably, only 4/150 strains (3%) possessed high MIC values to cefepime, cefepime-taniborbactam, meropenem, meropenem-vaborbactam, and levofloxacin. The rank order of potency by percent susceptibility or percent provisional susceptibility (\u226416 \u00b5g/mL for cefepime-taniborbactam) against these Burkholderia spp. was as follows: meropenem-vaborbactam (89%S) > cefepime-taniborbactam (69%S) > meropenem (55%S) > levofloxacin (26%S) > cefepime (23%S)  but much lower than relebactam and vaborbactam 3.2 and 38 \u00b5M, respectively (koff rate was for taniborbactam. These results demonstrate that the efficacy of cefepime-taniborbactam is largely the result of the taniborbactam inhibition of PenA1 \u03b2-lactamase. Taniborbactam and avibactam inhibited PenA1 in vitro with relatively similar potencies, while relebactam and vaborbactam exhibited weaker inhibitory kinetic profiles in comparison. In summary, cefepime-taniborbactam will be a welcome addition to the antibiotic arsenal due to its broad-spectrum activity against Enterobacterales and P. aeruginosa producing class A, B, C, and D \u03b2-lactamases. With broader analyses, cefepime-taniborbactam may also be valuable to treat infections caused by Burkholderia species.The parameters for inhibition of the ectively  (22, 29,ectively . Moreove"}
+{"text": "On September 1, 2022, the Advisory Committee on Immunization Practices recommended a bivalent mRNA COVID-19 booster dose for persons who had completed at least a primary COVID-19 vaccination series \u22652 months earlier. Early data showed high effectiveness of a bivalent booster in preventing COVID-19-associated hospitalization within 45 days of receipt; however, little is known about the durability of this protection.Data from the Investigating Respiratory Viruses in the Acutely Ill (IVY) Network were used to conduct a case-control analysis measuring bivalent vaccine effectiveness (VE) against COVID-19\u2013associated hospitalization over time. During September 8, 2022\u2013April 1, 2023, immunocompetent, hospitalized adults aged \u226565 years with COVID-19-like illness were enrolled at 25 hospitals in 20 U.S. states. COVID-19 case-patients tested positive for SARS-CoV-2 by a nucleic acid or antigen test within 10 days of illness onset, while control-patients tested negative for SARS-CoV-2 during the same interval. Multivariable logistic regression was used to measure absolute and relative bivalent VE adjusted for age, sex, race and ethnicity, admission date, and U.S. Health and Human Services region. Unvaccinated patients and patients who received 2\u20134 doses of monovalent-only mRNA vaccine were used as the reference group for absolute and relative VE, respectively. Bivalent VE was calculated for 7\u201389 days and 90\u2013179 days from booster dose receipt to illness onset.A total of 2,787 immunocompetent, hospitalized adults aged \u226565 years were enrolled in the IVY Network during the study period . Absolute VE of a bivalent booster dose against COVID-19-associated hospitalization was 58% (95% CI=42%\u201370%) after 7\u201389 days and 27% (95% CI= -7% to 50%) after 90\u2013179 days. Relative VE of a bivalent booster dose was 54% (95% CI=41%\u201364%) after 7\u201389 days and 19% (95% CI= -8% to 39%) after 90\u2013179 days .Bivalent mRNA vaccination provided moderate protection against COVID-19-associated hospitalization within 90 days of receipt among adults aged \u226565 years, with waning protection after 90 days. Additional booster doses could improve protection against COVID-19-associated hospitalization among older adults.Adit A. Ginde, MD, MPH, AbbVie: Grant/Research Support|Faron Pharmaceuticals: Grant/Research Support Ithan Peltan, MD, Asahi Kasei Pharma: Institutional support|Regeneron: Institutional support Emily T. Martin, PhD, MPH, Merck: Grant/Research Support Matthew Exline, MD, Abbott Labs: Honoraria|Regeneron: Grant/Research Support Adam S. Lauring, MD, PhD, Roche: Advisor/Consultant|Sanofi: Advisor/Consultant"}
+{"text": "Our objective was to assess their Kp clinical isolates were selected. Minimum inhibitory concentrations (MICs) were determined by broth microdilution in triplicate. Time-kill analyses were used to test CZA and MVB alone (1 and 4x MIC) and in combination with colistin , fosfomycin , gentamicin , meropenem , and tigecycline  against 1x108 cFu/mL. 24 hour log-kill was calculated as log cFu/mL decrease from time 0.15 KPC-ompK36 WT, IS5, or GD. 27% and 73% were KPC-3 and KPC-2, respectively. 100% were susceptible to CZA and MVB. None were resistant to COL. 3 were non-susceptible to GEN, which were all IS5 (Table 1).5 isolates each had Figure 1). Mean log-kills did not vary for CZA + COL, GEN, or TGC against ompK36 mutants vs WT; however, log-kills were significantly lower for CZA + FOS  or MEM  against ompK36 mutants compared to WT. Synergy rates were reduced from 100% against WT to 80%, 60%, and 40% against mutant isolates for CZA + COL, MEM, and FOS, respectively .In combination with CZA 1x MIC, each of COL, GEN, and MEM were bactericidal against all isolates; mean \u00b1 standard error (SE) log kills were -7.39 \u00b1 0.3, -6.58 \u00b1 0.93, and -5.81 \u00b1 0.21, respectively . Mean \u00b1 SE log kills were significantly greater against porin mutants compared to WT for MVB + COL , FOS , or TGC . Rates of synergy were also higher against porin mutants vs WT for MVB + FOS or COL .MVB 1x MIC in combination with COL, FOS, GEN, and TGC were bactericidal 67, 27, 80, and 47% of the time, with corresponding log kills of -3.96 \u00b1 1.1, -1.56 \u00b1 0.83, -5.05 \u00b1 0.82, and -2.53 \u00b1 0.28, respectively  and MVB (meropenem-vaborbactam) at 1X MIC, stratified by ompk36 status: wild-type (WT) versus mutant (MUT). Bactericidal is defined as -3 log kill from time 0 at 24 hours and indicated by the dotted horizontal line. COL = colistin; FOS = fosfomycin; GEN = gentamicin; MEM = meropenem; MIC = minimum inhibitory concentration; TGC = tigecycline.Figure 2Proportion of bactericidal, synergistic, and antagonistic outcomes for ompK36 wild-type  versus mutant  KPC-Kp isolates tested against CZA (ceftazidime-avibactam) or MVB (meropenem-vaborbactam) at 1X MIC as single agents and in combination with colistin (COL), gentamicin (GEN), tigecycline (TGC), meropenem (MEM), or fosfomycin (FOS). Bactericidal activity was defined as -3 log kill from time 0 at 24 hours; synergy and antagonism were defined as -2 or +2 log kill at 24 hours compared to most active single agent, respectively.Porin mutations negatively impact CZA synergy with other antibiotics compared to WT, though 24-hour log kills for COL, FOS, GEN, and MEM remained below the bactericidal threshold. Conversely, porin mutations potentiate synergy between MVB and COL, FOS, or TGC.Ryan K. Shields, PharmD, MS, Allergan: Advisor/Consultant|Cidara: Advisor/Consultant|Entasis: Advisor/Consultant|GSK: Advisor/Consultant|Melinta: Advisor/Consultant|Melinta: Grant/Research Support|Menarini: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Pfizer: Advisor/Consultant|Roche: Grant/Research Support|Shionogi: Advisor/Consultant|Shionogi: Grant/Research Support|Utility: Advisor/Consultant|Venatorx: Advisor/Consultant|Venatorx: Grant/Research Support"}
+{"text": "Scientific Reportshttps://doi.org/10.1038/s41598-019-41543-0, published online 21 March 2019Correction to: This Article contains errors. In Figure 6C the OS image for SCR siRNA is incorrectly duplicated as the Figure 8C OS image for CNT. The correct Figure"}
+{"text": "Kosakonia pseudosacchari RX.G5M8, a putative methylotroph, was isolated from garden soil in Hong Kong. Its complete genome, a single chromosome of 4,953,935 bp (GC content 53.91%), was established through hybrid assembly. Kosakonia, designated in 2013 after reassignment of four species of Enterobacter  was vortexed with 900 \u00b5L 0.9% (wt/vol) saline and a 10-\u00b5L aliquot used to inoculate 1 mL of a minimal salts medium (MSM)  containihttps://github.com/FelixKrueger/TrimGalore) , giving 2,137,624 read pairs  totaling ~534 Mbp. Long-read libraries, prepared from the same extracted DNA using the Rapid Barcoding Kit SQK-RBK004 (without size selection), were sequenced via Oxford Nanopore\u2019s Spot-ON Flow Cell (vR9.4.1) and MinION sequencer, with MinKNOW v20.06.2 software and basecalling by Guppy v6.1.5 high-accuracy mode. The final long-read data set, trimmed by Filtlong v0.2.1 (https://github.com/rrwick/Filtlong) , totaled 63,710 reads (984 Mbp) with mean length 15,448 bp . Default parameters were used for all software unless otherwise specified.A paired-end short-read sequencing library (NEB Next Ultra DNA Library Prep Kit) was sequenced via the NovaSeq 6000 platform using an SP PE250 flowcell and v1.5 Reagent Kit. Reads were quality filtered and trimmed using TrimGalore! v0.6.7 (Kosakonia pseudosacchari strain BDA62-3 (CP063425), with an average nucleotide identity of 98.67%.Unicycler v0.4.3  combinedRX.G5M8 possesses a glutathione-dependent formaldehyde detoxification pathway for methanol metabolism . The nit"}
+{"text": "The primary objective of this study was to characterize meropenem (MERO) pharmacokinetics (PK) in critically ill infants and children with septic shock to develop dosing recommendations to achieve therapeutic drug exposure in this complex population with a spectrum of renal function from augmented renal clearance (ARC) to acute kidney injury (AKI).Children \u22654 weeks old hospitalized with septic shock requiring fluid resuscitation and pressors, receiving MERO 20 mg/kg IV every 8 hr as standard-of-care, were enrolled and studied prospectively from 2019 to 2023. Population PK modeling was used to derive Bayesian post-hoc PK parameters (NONMEM 7.3). Renal biomarkers  were evaluated as covariates. Estimated glomerular filtration rate (e-GFR) was calculated by the modified Schwartz equation.A total of 304 MERO serum concentrations were included from 26 participants. Median age was 12.6 yr (range 1.2-19.6); weight 38.4 kg (10-98); SCr 0.36 mg/dL (0.09-2.57); CYS 448.9 mg/d (178.3-1824.1), and NGAL 176.05 ng/mL (29.3-1000). Median MERO serum concentration was 11 mcg/mL . A two-compartment model with allometrically scaled weight on clearance (0.75) best described the data. Significant covariates  were e-GFR, CYS, NGAL, age, diagnosis of appendicitis and albumin. Using the final model, the median CL was 0.15  L/hr/kg and V1 0.20  L/kg, with GFR 139  mL/min/1.73 m2 . The distribution of MERO concentrations, CL and GFR across quartiles documents wide variations in renal function observed in septic shock, from AKI to ARC .Meropenem Serum Concentration, Meropenem Renal Clearance, and Glomerular Filtration Rate, by QuartilesWide variation in MERO plasma exposure was documented in children with septic shock across the spectrum of renal function. GFR-based dosing recommendations may optimize MERO dosing and improve outcomes in this population.This work was supported by NICHD grant 1 R01 HD095547-01Edmund Capparelli, PharmD, Melinta: Advisor/Consultant"}
+{"text": "The physicochemical properties of silver nanocomposites (MTP(BTP)/Ag NCs) were fully elucidated using spectroscopic and microscopic tools. The antibacterial activity of the nanocomposites was screened against six multidrug-resistant pathogenic strains, comparable to ampicillin and ciprofloxacin commercial drugs. The antibacterial performance of BTP was more substantial than MTP, notably with the best minimum inhibitory concentration (MIC) of 0.0781 mg/mL towards Bacillus subtilis, Salmonella typhi, and Pseudomonas aeruginosa. Among all, BTP provided the clearest zone of inhibition (ZOI) of 35 \u00b1 1.00 mm against Salmonella typhi. After the dispersion of silver nanoparticles (AgNPs), MTP/Ag NCs offered dose-dependently distinct advantages over the same nanoparticle with BTP; a more noteworthy decline by 4098 \u00d7 MIC to 0.1525 \u00d7 10\u22123 mg/mL was recorded for MTP/Ag-1000 against Pseudomonas aeruginosa over BTP/Ag-1000. Towards methicillin-resistant Staphylococcus aureus (MRSA), the as-prepared MTP(BTP)/Ag-1000 displayed superior bactericidal ability in 8 h. Because of the anionic surface of MTP(BTP)/Ag-1000, they could effectively resist MRSA (ATCC-43300) attachment, achieving higher antifouling rates of 42.2 and 34.4% at most optimum dose (5 mg/mL), respectively. The tunable surface work function between MTP and AgNPs promoted the antibiofilm activity of MTP/Ag-1000 by 1.7 fold over BTP/Ag-1000. Lastly, the molecular docking studies affirmed the eminent binding affinity of BTP over MTP\u2014besides the improved binding energy of MTP/Ag NC by 37.8%\u2014towards B. subtilis-2FQT protein. Overall, this study indicates the immense potential of TP/Ag NCs as promising nanoscale antibacterial candidates.Newly synthesized mono- and bis-thioureidophosphonate (MTP and BTP) analogues in eco-friendly conditions were employed as reducing/capping cores for 100, 500, and 1000 mg L Recently, the emergence of antibiotic-resistant microbes has been seen as a global health concern affecting many people\u2019s lives . Regardi\u22121. The  nuclear magnetic resonance (NMR) spectra were consecutively recorded at 400, 101, and 162 MHz in DMSO-d6 using a Bruker Avance III HD, 600 MHz-NMR spectrometer probe, and BBFO cryoprobe . The elemental analysis of samples  was undertaken for the raw TP derivatives (MTP and BTP) by CHNS Vario EL III Elementar analyzer, Germany. The phosphorus content (%) was estimated via the chemical digestion in sulfuric/nitric solution and photometric measurements at \u03bbmax = 410 nm. Melting points (m.p) were recorded using  without correction. The surface images of nanocomposites were obtained using transmission electron microscope (TEM) by JEOL, JEM-2100 Tokyo, Japan. The crystalline properties of nanocomposites were obtained using powder X-ray diffraction (XRD) analysis using Shimadzu 6000 X-ray diffractometer with Cu K\u03b1-radiation (\u03bb = 1.54 nm) at 25 \u00b0C in the 2\u03b8 range of 10\u201380\u00b0 operating at a scan rate of 1 deg/min. X-ray photoelectron spectroscopy (XPS) was conducted using a Perkin Elmer PHI 5600, Perkin Elmer Instruments, Waltham, MA, USA, with analytical zone\u2019s diameter of 1 mm and indium sheets for sample deposition using Al K\u03b1 X-rays radiation source (200 W). The photometric assays of antibacterial results were measured by (Infinite F50 Robotic absorbance microplate reader) in the wavelength range of 400 to 750 nm. The zeta potential charge of silver nanocomposites was determined via dynamic light scattering using Zetasizer Nano ZS, Malvern Instruments Ltd., Malvern, UK. Scanning electron microscopy (SEM) was performed using JSM 6390 LA, JEOL, Tokyo, Japan, with a 15 KV accelerating voltage.The Fourier transform infrared spectroscopy (FT-IR) analysis has been accomplished for all samples on Bruker Alpha II spectrometer, Bremen, Germany, in the wavenumber range 4000\u2013400 cm3) were purchased from Aladdin . Solvents, such as acetonitrile (CH3CN), ethanol, and methanol were obtained from a commercial supplier  and used without further drying and purification. Deionized water was used to prepare all solutions and suspensions.Thiourea  and triphenyl phosphite  were obtained from Macklin . Terephthalaldehyde , pyridinium trifluoromethanesulfonate  (as protic ionic liquid), and silver nitrate   with the aid of a pyridinium trifluoromethanesulfonate (pyridinium triflate) as a Lewis acid protic ionic liquid-based catalyst. In the meantime, thiourea /,  terephthalaldehyde, triphenyl phosphite /, and pyridinium triflate / in 5 and 10 mL of CH3CN were progressively incorporated, affording MTP and BTP in a molar ratio of 1:1:1 and 2:1:2, respectively. Further, both solutions were magnetically stirred overnight under ambient conditions and the reaction was monitored via qualitative thin layer chromatography (TLC) using hexane\u2013methylene chloride (3:1) as eluent mixture. Eventually, the final products were filtered off under vacuum, washed with methanol, dried, and kept in a desiccator for 2 days giving MTP and BTP in good yields.The synthesis process included preparation of two different systems of mono and bis thioureidophosphonate (TP) products, where the involved components  were dissolved in acetonitrile (CHDiphenyl ((4-formylphenyl) (thioureido)methyl) phosphonate, denoted as mono-thioureidophosphonate (MTP):\u22121 (NH)str. + (NH2) str. overlapped, 3057.58 cm\u22121 , 2894.63 cm\u22121 , 1698.02 cm\u22121 , 1590.99 cm\u22121, (NH+NH2) bending, 1525.42 cm\u22121 (>C=C<) phenyl rings, 1488.78 cm\u22121 (>C=S) asym., 1194.69 cm\u22121 (-P=O), 944.95 cm\u22121 (P-O-C), 760.78 cm\u22121 (P\u2013C), and 684.61 cm\u22121 (>C=S) sym. 1H-NMR; \u03b4 ppm: 5.50 , 6.63\u20137.99 , 9.40\u20139.52 , 9.76 , and 10.04 . 13C-NMR ; \u03b4 ppm: 54.59 and 56.16 ppm (P-C\u2013H), 115.24, 120.22, 120.53, 125.39, 127.96, 128.92, 129.90, 149.80 (Ar-C), 183.84 (>C=S), and 192.73 (-CHO). 31P-NMR ; \u03b4 14.08 ppm. Elemental analysis calc. (%) for C21H19N2O4PS: C, 59.15; H, 4.49; N, 6.57; P, 7.26; S, 7.52; and O, 15.01; meas. (%): C, 57.85; H, 4.27; N, 6.34; P, 6.97; and S, 6.99.Light-beige solid, yield: , m.p: 160\u2013163 \u00b0C, FT-IR: 3457.74\u20133311.18 cmTetraphenyl ) bis(phosphonate) denoted as bis-thioureidophosphonate (BTP):\u22121 [(NH)str. + (NH2) str. overlapped], 3057.58 cm\u22121 , 2895.59 cm\u22121 , 1591.95 (>NH + -NH2) bending, 1527.35 cm\u22121 (>C=C<) phenyl rings, 1489.74 cm\u22121 (>C=S) asym., 1195.65 (-P=O), 947.84 (P-O-C), 761.74 cm\u22121 (P\u2013C), and 761.74 cm\u22121 (>C=S) sym. 1H-NMR; \u03b4 ppm: 5.93 , 6.54\u20137.61 , 9.21\u20139.33 , and 9.38 ppm . 13C-NMR ; \u03b4 ppm: 54.50 and 56.06 (P-C\u2013H), 115.26, 120.29, 120.86, 125.40, 127.50, 129.33, 129.84, and 149.94 (Ar-C), and 183.83 (>C=S). 31P-NMR ; \u03b4 ppm: 14.85 ppm. Elemental analysis calc. (%) for C34H32N4O6P2S2: C, 56.82; H, 4.49; N, 7.82; P, 8.62; S, 8.92; and O, 13.36; found: C, 54.97; H,4.36; N, 8.18; P, 8.83; and S, 7.91.Light-yellow solid, yield: , m.p: 135\u2013137 \u00b0C, FTIR: 3310.21 cm3 solution of 100, 500, and 1000 mg L\u22121 concentrations. Then, the prepared solutions were magnetically stirred (200 rpm) for 12 h, at ambient conditions. Afterward, the attained colloidal solutions were centrifuged, and the settled nanoparticles were washed with ethanol/deionized water and then oven-dried at 50 \u00b0C for further characterization ..36].Well dispersed AgNPs were produced via one-pot synthesis method using thioureidophosphonates (TPs) as dual-functioning reducing and capping agents. This affords spherical AgNPs different dispersion patterns based on the reaction conditions of AgNPs precursor concentrations and the structure of TPs .\u22121) illustrated meaningful changes in the characteristic absorption bands of capping agent (MTP)  was also involved in the interaction and reduction of Ag+ ions in all concentrations of 100, 500, and 1000 mg Ag L\u22121, respectively. Further, the aliphatic (H-C-P) bond was dramatically shifted from \u03bd = 2895 cm\u22121 to \u03bd = 2914 cm\u22121 in MTP/Ag-1000. On the other side, the FT-IR spectra of BTP and its  AgNP dispersions displayed similar changes, with a substantial shifting of amine functional groups from \u03bd = 3310 cm\u22121 to \u03bd = 3306, 3305, and (3320\u20133428) cm\u22121, consecutively  a. Notablcutively b. Hence,cutively . Basicalg Ag L\u22121 .+ ions, while the higher density of chelating centers in BTP significantly improved the reduction rate of Ag+ ions (with deeper color); thus, some smaller AgNPs were attained with the BTP surface (The morphology and dispersion nature of developed silver nanoparticles (AgNPs) were scrutinized using microscopic techniques. Accordingly, the TEM imaging for MTP/Ag-500 displayed good dispersion of spherical AgNPs with an average mean size range of 6.8\u201320.1 nm a\u2013c. On t surface d\u2013f. The Moreover, the crystallinity of the as-synthesized TPs after AgNPs functionalization was investigated compared to that of raw capping agents using XRD analysis. First, broad XRD patterns were individually centered at 2 theta (2\u03b8) = 21.7\u00b0 with (110) diffraction phases, reflecting the amorphous structure of both raw MTP and BTP analogues. The emergence of broad patterns stemmed from the strong inter- and intra-molecular hydrogen bonding in raw TP cores . Further+ ions incorporation and formation of AgNPs, X-ray photoelectron spectroscopy analysis (XPS) was carried out for MTP/Ag-500. The XPS results of MTP/Ag-500 were studied based on evaluating the changes in binding energies (BEs), atomic percentages (%), electronic properties, and chemical composition of MTP before and after Ag+ ion interactions. Five elemental signals of C 1s (at BE: 285.84\u2013287.23 eV), N 1s (400.72\u2013401.63 eV), O 1s (533.02\u2013534.81 eV), P 2p (134.32\u2013134.57 eV), and S 2p (163.19\u2013164.19 eV) were mainly evolved, consecutively  eV, besides considerable changes in the atomic percentages (%) of raw elemental functional groups that were estimated from 0.72\u201325.57%. Basically, these notably affected functional groups were ordered, respectively, in a descending mode according to changes in BEs as follows: C\u2013S (2p1/2) > C\u2013C, C\u2013H, and C=C > HC=O and -OH (H2O) > C\u2013S (2p3/2) > P\u2013C and P\u2013O (2p1/2) > >NH and -NH2 > C=S > C\u2013N > \u03c0\u2013\u03c0* sat, benzene rings, and C=S > C-OH, P\u2013O, and C\u2013O\u2013C > C\u2013O, C\u2013N, C\u2013P, and C=O  on the MTP, the main signal deconvolution assessed that Ag+ ions may be sorbed in their hydrated form . In this regard, the changes that occurred in BEs after AgNPs decoration were consistent with the results discussed before.To validate the most significant changes that occurred to raw functional groups after Agcutively . Regardiectively b 62]. H. H+ ions and C=O . FurtherStaphylococcus aureus (MRSA), Streptococcus mutans, and Bacillus subtilis) and Gram-negative  bacteria as well. Moreover, zone of inhibition diameters, minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), time\u2013kill kinetics, antiadhesion activity, and antibiofilm assay with morphological investigation were assessed for all developed compounds.The antibacterial potency of synthesized thioureidophosphonates (TPs) and their fabricated AgNPs-based composites was evaluated against several strains. The antibacterial activity using the agar well diffusion method was conducted against six clinical bacterial strains and their ATCC references of Gram-positive (methicillin-resistant Staphylococcus aureus (MRSA), Streptococcus mutans (S. mutans), and Bacillus subtilis (B. subtilis), compared to Ampicillin and Ciprofloxacin as reference antibiotics  resulted in higher MIC values = 1.25 mg/mL for both analogues. MTP and BTP recorded MICs at lower concentrations of 0.625 and 0.3125 mg/mL against S. mutans (ATCC-35668) and B. subtilis (ATCC-6633), progressively. The zone of inhibition (ZOI) was measured for each strain with DMSO as a negative control. Including the diameter of each well (6 mm) before treatment, MTP gave ZOIs in a range of 28 \u00b1 1.0, 30 \u00b1 1.00, and 24 \u00b1 1.00 mm, whereas the treatment with BTP increased the ZOIs width towards MRSA, S. mutans, and B. subtilis strains, respectively. Both TPs with MBC/MIC ratio of 2(+) have bactericidal properties, unlike Ampicillin and Ciprofloxacin. TPs loaded with silver nanoparticles (AgNPs)  were used to investigate the antibacterial properties towards three strains accompanied with a dose-dependent improvement in susceptibility. A clear MIC decline to very minute concentrations was appreciated from 0.156 to 0.1525 \u00d7 10\u22123 mg/mL with the three strains. The MIC of MTP significantly dropped by (4 \u00d7 MIC) after treating MRSA and S. mutans with MTP/Ag-100. Afterward, MTP/Ag-500 and MTP/Ag-1000 showed increased bactericidal properties against both strains, i.e., 16 and 2049 times sequentially. The treatment of MTP/Ag- to B. subtilis increased the potential bactericidal activity by 16, 128, and 1024 times, respectively. BTP/Ag-100 showed a more significant decline in MIC values by two and eight times against MRSA and B. subtilis, compared to MTP/Ag NCs. BTP/Ag-500 had less biocidal effects by 4 and 16 times that of MTP/Ag-500 on MRSA and B. subtilis, consecutively. This reflected that BTP and AgNPs have less synergistic interactions than MTP. However, BTP/AgNPs had the same bactericidal activity as MTP/AgNPs against S. mutans in 100 and 500 dispersions. Noteworthy, MTP and BTP loaded with 1000 mg L\u22121 resulted in a decrease in MICs by 2049 times against MRSA while BTP/Ag-1000 showed higher MIC against S. mutans and B. subtilis by only 1023 and 512 times, respectively , 38 \u00b1 1.00 (MTP/Ag-500), and 42 \u00b1 1.00 mm for MTP/Ag-1000, respectively. Likewise, ZOIs exhibited a gradual increase from 37 \u00b1 1.00 in BTP to 40 \u00b1 1.00 (BTP/Ag-100), 43 \u00b1 0.60 (BTP/Ag-500), and 49 \u00b1 0.100 mm (BTP/Ag-1000). MTP and BTP gave higher ZOIs with diameters of 35 \u00b1 1.00, 38 \u00b1 1.00, and 40 \u00b1 1.00 mm and 36 \u00b1 1.00, 41 \u00b1 1.00, and 41 \u00b1 1.00 mm, respectively, for B. subtilis , Salmonella typhi (S. typhi), and Serratia marcescens (S. marcescens) than previously screened Gram-positive bacteria and reference antibiotics  against P. aeruginosa than MTP, indicating higher susceptibility towards Gram-negative strains. Both MTP and BTP compounds had excellent inhibitory properties against S. typhi and S. marcescens, as their MIC ratio was reduced fourfold. Regarding the Ampicillin and Ciprofloxacin, 5\u00d7 and 20\u00d7 MIC reductions by MTP were recorded against S. typhi, while 5.0 and 1.2\u00d7 MIC were dropped with S. marcescens. On the contrary, BTP decreased the MICs of Ampicillin and Ciprofloxacin by 20 and 80 times towards S. typhi, and to 20 and 5 times against S. marcescens, respectively. Furthermore, the selectivity of these TP derivatives was determined against P. aeruginosa (ATCC-27853), S. typhi (ATCC-6539), and S. marcescens (ATCC-13880). Meanwhile, the MICs obtained by inoculating both ATCC isolates of P. aeruginosa and S. marcescens with MTP were 1.25 mg/mL for S. typhi (ATCC-6539). The selectivity of MTP was more effective, affording an MIC value of 0.625 mg/mL. BTP had two lowered MIC values against P. aeruginosa (ATCC-27853), S. typhi (ATCC-6539), and S. marcescens, with 0.3125 mg/mL for P. aeruginosa and 0.625 mg/mL for S. marcescens (ATCC-13880). On the other hand, ZOI results of MTP and BTP against P. aeruginosa, S. typhi, and S. marcescens were 28 \u00b1 1.00, 28 \u00b1 1.00, and 35 \u00b1 1.00 mm for MTP, while 33 \u00b1 0.60, 35 \u00b1 1.00, and 38 \u00b1 1.00 mm for BTP, consecutively. Therefore, MBC values ranged from 1.25 to 0.156 mg/mL, resulting in a significant bactericidal efficacy against all bacterial strains with MBC/MIC ratio = 2(+). Notably, AgNPs improved the antibacterial susceptibility of TP cores, lowering MIC values from 0.01953 mg/mL to 7.629 \u00d7 10\u22125 mg/mL. Basically, MTP/Ag-100 minimized the MIC concentration of raw MTP by around 32 times towards P. aeruginosa, and 16 times for both S. typhi and S. marcescens, successively. MTP/Ag-500 reduced MICs by 256 times for P. aeruginosa and 128 times for S. typhi and S. marcescens compared to raw MTP. For MTP/Ag-1000, MTP displayed an elevated antibacterial activity towards P. aeruginosa, S. typhi, and S. marcescens. Nevertheless, the net antibacterial activity of BTP was higher than MTP, and the combinatorial interactions of AgNPs with MTP were more tunable, unlike BTP. In the meantime, BTP/Ag-100 lessened the MIC ratio to raw BTP by fourfold towards P. aeruginosa and eightfold against both S. typhi and S. marcescens. BTP/Ag-500 had a remarkable bactericidal effect on S. typhi up to 128 times. Then, the BTP/Ag-1000 reduced the dose of MIC to 512 times with P. aeruginosa and 1023 times with both S. typhi and S. marcescens, respectively. Pertinently, BTP loaded with AgNPs manifested less powerful bactericidal potency than MTP capped AgNPs  started to dramatically decline after 1 h of treatment (10) and (1.8 log10) CFU/mL after 18 h of incubation, respectively. However, once MRSA was treated with TP/Ag NCs, the picture was changed where the MTP/Ag-100 induced the killing potential with cell count reductions to (2.2 log10) and (0.9 log10) CFU/mL for MTP/Ag-500, consecutively. More significant reductions of the cultural population of MRSA were achieved to (1.1 log10) and (0.3 log10) CFU/mL for BTP/Ag-100 and BTP/Ag-500, progressively. Despite the higher killing efficacy of BTP/Ag NCs than MTP/Ag NCs, the combination between AgNPs and MTP brought more bactericidal advantages over BTP, which was observed from the gradual improvement of killing (%) between dose 100 and 500 for both AgNPs formulations. Remarkably, the entire population was reduced and killed within 8 h, for both MTP (BTP)/Ag-1000 composites. Interestingly, the bactericidal rates towards MRSA were previously confirmed by both TP/Ag-1000; where both reduced MBC values to the same degree of 2049 times  of MRSA (ATCC-43300) suspensions for 18\u201324 h, using vancomycin hydrochloride (VAN) as a positive control, and the negative control was tryptic soy broth (TSB). Antiadhesion activity was quantitatively estimated against MRSA (ATCC-43300) on the microplate reader at \u03bbmax = 570 nm, validating an increase in MRSA (ATCC-43300) detachment in a dose-dependent manner within 1.25, 2.5, 5.0, and 10 mg/mL  than MTP, which could be attributed to its superior hydrophobic characteristics as the non-polar parts (aromatic rings) exceeded the polar ones, based on its molecular formula (M.F), namely, C34H32N4O6P2S2, comparable to MTP with M.F of C21H19N2O4PS. Moreover, the electrostatic repulsion forces assessed between the negatively charged MTP/Ag NCs and MRSA (ATCC-43300) surface played a critical role . Ad. Ad6 CFUactivity . On thisStaphylococcus aureus (MRSA) is one of the most opportunistic and resistant pathogens, being able to produce an extracellular polymeric substance (EPS) inducing biofilm growth prior to the occurrence of cross infection [MRSA (ATCC-43300) as a Gram-positive bacterium with reference to vancomycin hydrochloride (VAN) as a positive control. The biofilm disintegration activity (%) was validated by the microtiter plate assay using a semi-quantitative analysis based on crystal violet staining. Regarding 96-well plates treated with raw MTP and BTP, the biofilm (violet color circle) was slightly eliminated, while the gradual removal was notably viewed by six TP/Ag NCs, (max \u2248 570 nm with respect to the negative control of tryptic soy broth (TSB). The results showed a significant reduction in biofilm growth when treated with raw TPs, even after 24 h of incubation. The biofilm inhibition (%) was promoted with the gradual increase in the concentrations of tested compounds in the 1.25, 2.5, 5.0, and 10.0 mg/mL (MRSA (ATCC-43300). The studied TP cores (\u03b1-aminophosphonate derivatives) are considered structural bio-isosteres of \u03b1-amino acids (peptides) with a common amine functional group, but the carboxylic groups in \u03b1-amino acids are replaced with related phosphorous-containing moieties such as phosphonic acid. However, the reported data about their antibiofilm mechanism of action towards MRSA biofilm are directed for hybrid \u03b1-APs; thus, the inhibitory actions of mentioned \u03b1-APs analogues were proposed [Methicillin-resistant nfection . The ant/Ag NCs, . Therefo.0 mg/mL . Yet, frproposed ,73. Firsproposed . Moreoveproposed . It was proposed ,76.+ ions and reactive oxygen species (ROS), which increased the toxic effects on the integrity of bacterial cell wall. Additionally, ROS can cause the total impairment of envelope-bound proteins, deoxyribonucleic acid (DNA), respiratory coenzymes, and antioxidants such as glutathione (GSH) [MRSA (ATCC-43300) in the next section [After AgNPs functionalization, biofilm inhibition capability (%) was promoted for both MTP and BTP in all studied concentrations , consecutively. The most effective and synergistic concentration needed for the disruption of MRSA (ATCC-43300) biofilm was observed at 5.0 mg/mL for both TP/Ag NCs. For 10.0 mg/mL, the biofilm eradicating ability for both MTP(BTP)/Ag-1000 reached maximum values at 79.5 and 83.6%, progressively . Nonethene (GSH) . RemarkaMRSA (ATCC-43300) was also qualitatively verified through scanning electron microscopy (SEM) after incubation by 100 \u00b5g/mL of raw compounds and their TP/Ag-1000. The two upper images (MRSA (ATCC-43300) surface on which aggregating grape-like colonies were developed to form the biofilm. On the contrary, low percentages of biofilm clusters were present on the surfaces of MRSA (ATCC-43300) after treatment with raw MTP and BTP, indicating that these compounds could prevent bacterial biofilm formation and induce cell death. Moreover, almost no biofilm matrices could be observed on MRSA treated with MTP and BTP decorated with AgNPs, validating the potent biofilm disintegration ability of TP/Ag NCs. In summary, these observations were in line with antibiofilm investigation results and other studies referring to the ability of synthesized AgNPs to severely interfere with cell wall (biofilm) components, thus hampering metabolic functions, and thereby, MRSA (ATCC-43300) cells are killed [The biofilm inhibition of r images  represene killed .MRSA and B. subtilis) and Gram-negative (P. aeruginosa and S. typhi) bacterial proteins with respective codes of PDB: 4DKI, 2FQT, 5ZHN, and 3ZQE, in addition to confirming their illegibility as antibacterial agents with the experimental work. Furthermore, the docking results for the prepared raw compounds (MTP and BTP) in all downloaded proteins, along with type of chemical bonding and the amino acids involved in the interaction with the protein, were estimated. Additionally, values recorded for binding affinity and root mean square deviation (RMSD) were tabulated. There are different types of interactions, including hydrogen and hydrophobic H-pi interactions, with the binding sites of all the investigated proteins  for MRSA, with values of \u22127.9776 kcal/mol and \u22127.9074 kcal/mol, respectively. Amino acids involved in the interaction between BTP and the B. subtilis protein were GLY127 , GLY127 , and GLU57 , which are the same amino acids involved in the interaction of the co-crystallized ligand and the protein  and TYR446   and thereby for biofilm growth [Molecular docking studies were conducted for the prepared TP compounds to investigate their binding affinity for Gram-positive  , while Tcutively . It was 1.2593 \u00c5 . Interesproteins . These bm growth ,82; thusIt is pertinent to note that in this study, the structural and functional diversity of raw TP surfaces afforded highly tunable surface interactions and synergy, with AgNPs giving remarkable antibacterial activity. Additionally, the molecular docking studies were in good coherence with most experimental results for TPs and TP/Ag NCs. Based on the above considerations, some structural modifications may be applied in the future to exploit these compounds in various applications such as antibacterial textile fabrics, antibacterial food packaging films, and antibacterial thermoplastic polymer nanocomposites ,84,85.p > \u22122.5 [50 value was 0.335 and 0.285log \u03bcg/L for MTP-AgNPs and BTP-AgNPs, respectively. The anticipated scores are summarized below.Absorption, distribution, metabolism, and excretion (ADME) analysis is very helpful in simplifying clinical trials, especially in the early stage of drug design. Intestinal absorption, skin sensitization, and oral bioavailability are absorption parameters considered in drug discovery . An intep > \u22122.5 ; howeverp > \u22122.5 ; it measp > \u22122.5 , MTP-AgNp > \u22122.5 . Regardip > \u22122.5 ,88. Comp+ ions affording TP/Ag NCs, the physicochemical properties of which were thoroughly investigated through FT-IR, -NMR, elemental analysis, and TEM, XRD, and XPS analyses, respectively. The antibacterial properties of the as-prepared raw cores were studied via tuning different concentrations of silver nitrate, i.e., 100, 500, and 1000 mg L\u22121. Both raw Mono-TP and Bis-TP analogues proved to be bactericidal towards all isolates. In a dose-increasing manner, time\u2013kill assay results manifested the record-breaking bactericidal effect of both MTP(BTP)/Ag-1000 towards all MRSA cells within 8 h. At high concentrations, the antifouling characteristics of evaluated compounds were influenced by the surface electrostatic repulsion with MRSA (ATCC-43300), while hydrophobicity played a critical role at lower concentrations. Targeting the biofilm of MRSA (ATCC-43300) was confirmed, with net activity of 79.5 and 83.6% for MTP/Ag-1000 and BTP/Ag-1000, progressively, at highest used concentration. Nonetheless, higher net antibiofilm activity was observed in BTP/Ag NCs; the simpler MTP structure tuned the surface work function with Ag+ ions, giving better colloidal AgNPs and a gradual improvement in the biofilm disruption % after the nanoloading. The docking results were in line with the experimental work, where the best results were estimated for raw BTP towards B. subtilis-2FQT protein, recording binding energy (BE) of \u22127.9776 Kcal/mol. Interestingly, the progress in BE of MTP/AgNPs to \u22127.6668 Kcal/mol could stem from the exerted Ag-acceptor bonds with both amino acids (SER80 and GLU57) of B. subtilis-2FQT protein. Further, in silico pharmacokinetics assessment was conducted as a step to predict the potential safety and toxicity of the TP/Ag NCs, which revealed AMES toxicity for one of the compounds with neither hepatic toxicity nor skin sensitization. In summary, this study has provided a good design for the synthesis and development of a new generation of nano-based aminophosphonate composites as promising antibacterial candidates, although their safety should be carefully evaluated for each specific application; further studies are needed to fully understand the toxicity mechanisms and the optimal conditions for their pharmaceutical application.Two potent antibacterial agents were facilely synthesized in a one-pot green Kabachnik\u2013Fields reaction and then exploited as reducing/capping surfaces for Ag"}
+{"text": "Weight gain associated with integrase strand transfer inhibitors (INSTI) has been well documented among patients with HIV. However, recent reports suggest that the observed weight gain among patients who switch to INSTIs may be associated in part with their pre-switch regimen.We conducted retrospective analyses of a 50% sample of all treatment-experienced patients who switched to second-generation INSTI containing regimen (bictegravir/dolutegravir) at the Duke Adult Infectious Diseases Clinic  between 1 January 2014 and 31 December 2021. Covariates included sex at birth, age, race, ethnicity, pre-switch regimen, body mass index (BMI), CD4 count and HIV-1 viral load. All weights documented within 24 months from regimen switch were included. Outcomes of interest were percent change in weight and absolute weight change. Ordinary least squares was used to obtain average percent weight change from baseline, adjusted for covariates.Overall, 339 persons were included in the analyses. Cohort demographics were: median age 51.2 , 73% male sex at birth, 56% Black, 4% Hispanic ethnicity. At regimen switch, median CD4 count was 671 (IQR 460-935), and 78% had a viral load < 200 copies/cc. Mean weight at regimen switch was 87.8 kg (standard deviation [SD] 22.1kg), and mean total weight change over 24 months post-switch was +1.3 kg (SD 7.1 kg). Overall, 28% of cohort members gained >5% of baseline body weight within 24 months of switch. Hispanic ethnicity  and BMI < 20  were associated with post-switch weight gain. Only efavirenz-containing pre-switch regimens were significantly associated with weight gain over 24 months post-switch; EFV/FTC/TDF , all other EFV-containing regimens (6.3% [95% 0.1-12.4]).Pre-switch regimens containing efavirenz were significantly associated with weight gain after switch to second-generation INSTI-based regimens, consistent with findings from non-US based cohorts.Nwora Lance Okeke, MD MPH, Gilead Sciences: Advisor/Consultant"}
+{"text": "PurposeMarijuana use has been increasing in the adolescent population. Our objective was to examine the prevalence of marijuana use among a sample of adolescents and young adults, determine an association with risk-taking behaviors, identify reported medical symptoms, and delineate common beliefs about marijuana use.MethodsA questionnaire was administered to a sample of patients aged between 12 and 23 years old presenting to the emergency department of Penn State Hershey Medical Center, Hershey, Pennsylvania. Data were stratified by marijuana users and non-users, and further stratified by traditional  and non-traditional  use.ResultsThe analysis was based on 200 questionnaires. Thirty-nine percent (n=78) reported marijuana use. Marijuana users were more likely to report previous sexual intercourse , as well as the use of alcohol , cigarettes , prescription pain medications , and cocaine . Users more likely reported texting while driving  and experienced physical or electronic victimization due to bullying . Users were more likely to report gastroesophageal reflux disease (GERD), attention deficit disorder (ADD), anxiety, and depression. The most common symptoms associated with marijuana use were anxiety (65.4%), headache (61.6%), nausea/vomiting (53.8%), cough (51.3%), and abdominal pain (47.4%). Sixty-nine percent of respondents believed marijuana was \u201csafer than other drugs\u201d.ConclusionBased on our sample, we identified risk-taking behaviors, medical symptoms, and beliefs associated with marijuana use. Healthcare professionals may use these data to provide screening and anticipatory guidance to adolescents who use marijuana and consider marijuana use in their differential diagnosis. According to the 2021 Monitoring the Future study, marijuana use by adolescents declined from the late 1990s to the late 2000s but has since begun to rise again, peaking in 2020 with legalization both recreationally and medically in many states [Despite these negative consequences, many benefits of marijuana use have been cited. Clinical indications for medical marijuana vary by state but can include epilepsy and seizure disorders, cancer, HIV/AIDS, neurodegenerative disease, chronic pain, nausea, and post-traumatic stress disorder .In recent years, CBD has quickly grown in popularity in the form of oils, tinctures, and vapes as a treatment for a wide variety of diseases ranging from cancer to epilepsy, anxiety, pain control, and palliative care, with forms of epilepsy being the only FDA-approved indication to date . Many ofWhile there are many studies documenting different adverse effects associated with marijuana use -12, therWe conducted an observational, self-administered questionnaire-based study between August 2019 and September 2020, utilizing a sample of 200 patients presenting to the emergency department at the Penn State Hershey Medical Center, Hershey, Pennsylvania, aged 12 and 23 years. We excluded patients presenting with altered mental status, severe pain, developmental delay, acute psychiatric illness, or respiratory distress. Patients were also excluded if they were under 18 years of age, did not have a guardian present, were non-English speakers, or had previously completed the questionnaire.Once potential subjects were identified, the study team approached the patient and their guardian (for those less than 18 years old), obtained verbal consent, and provided the questionnaire and writing utensil. The questionnaire included 35 questions, focusing on demographics , risk-taking behaviors , self-esteem  , medicalData were stratified by whether participants were marijuana users or non-users, and marijuana users were further stratified by traditional and non-traditional use. Marijuana users were those who reported marijuana use; non-users were those who did not report marijuana use. Traditional marijuana users were classified as those reporting using vapes, pipes, or edibles, and non-traditional marijuana users were those using oils or concentrates, topical creams, or others.Descriptive statistics were generated, including means and standard deviations for continuous variables. This was an exploratory survey to find out more information about marijuana, and therefore, no formal sample size calculation was performed. Comparisons between categorical variables were analyzed using contingency table analysis; significance levels were determined by Chi-square tests. Data were stratified by marijuana users and nonusers, and marijuana users were further stratified by traditional and nontraditional use. The internal consistency of Likert scale questions was assessed by Cronbach's alpha, which was found to be 0.89. No adjustments for multiple tests were made. Tests were two-sided and based on a significance criterion of p<0.05. A multivariable logistic regression was then run, including all variables that were significant in the univariate analyses. All analyses were performed with the use of the SAS statistical package, version 9.4 . The Pennsylvania State University Institutional Review Board approved this study .Data analysis was performed on 200 completed questionnaires. The mean age of respondents was 17.5 years; 62.5% were female, and 66% identified as White (Table Marijuana use was reported in 39% (n=78) of respondents: 59% (n=46) were traditional, and 41% (n=32) were non-traditional. Marijuana users were less likely to live in a two-parent household than non-users (30.8% (95% CI: 22-42) vs. 54.1% (95% CI: 45-63); p= 0.0055).When stratified by frequency of use, compared to non-users, users were also more likely to have had previous sexual intercourse (79.5% (95% CI: 69-87) vs. 32.8% (95% CI: 25-41.6); p=<0.0001), and have had coitarche before age 18 (53.8% (95% CI: 43-64.5) vs. 21.3% (95% CI: 15-29.5); p=<0.0001)  vs. 10.7%% (95% CI: 6-17.5); p=<0.0001), cigarettes use (41% (95% CI: 30-52) vs. 8.2% (95% CI: 4-14); p=<0.0001), cigars use (30.8% (95% CI: 22-42) vs. 3.3% (95% CI: 1-8); p=<0.0001), chewing tobacco use (18% (95% CI: 11-28) vs. 0.8% (95% CI: 0-4); p<0.0001), prescription pain medications without a prescription (20.5% (95% CI: 13-31) vs. 4.1% (95% CI: 1-9); p=0.002), and cocaine use (14.1% (95% CI: 8-24) vs. 0.8% (95% CI: 0.01-5); p=0.0017)  (24.4% (95% CI: 16-35) vs. 9.0% (95% CI: 5-15.6); p=0.0164), anxiety (57.7% (95% CI: 47-68) vs. 35.2% (95% CI: 27-44); p=0.007), attention deficit disorder (ADD) (29.5% (95% CI: 20-40) vs. 10.7% (95% CI: 6-17.5); p=0.003), and depression (51.3% (95% CI: 40-62) vs. 27.9% (95% CI: 21-36); p=0.002). There were no significant differences between users and non-users who reported the following symptoms during the previous six months: chest pain, racing heart, difficulty breathing or coughing, dizziness, abdominal discomfort, nausea or vomiting, headache, tremors, sleep disturbances, or dehydration. However, there was a statistically significant increase in reports of anxiety in users as compared to non-users (66% (95% CI: 55-76) vs. 47% (CI: 38-56); p=0.009) (Table The most common symptoms associated with marijuana use were anxiety (65.4%), headache (61.6%), nausea/vomiting (53.8%), cough (51.3%), and abdominal pain (47.4%); there was no significant difference in symptom reports between traditional and nontraditional users.Vapes (25.6%) and pipes (25.6%) were the most common forms of marijuana use reported, and 61.6% of respondents reported using marijuana recreationally Table .With respect to perceived beliefs, 64% of respondents believed cannabis products are \u201cless addictive than other drugs,\u201d 69% believed they are \u201csafer than other drugs,\" 48% believed they \u201chelp to treat my medical illness\u201d and 33% believed that marijuana use is \u201cjust for fun.\" When asked to report a myth regarding cannabis products, respondents reported many common phrases, including \u201cit is a gateway drug\u201d (n=11), \u201cit is addictive\u201d (n=9), and \u201cit can kill you\u201d (n=7).After a multivariable logistic regression of all variables that were significant in the univariate analyses, the variables that ended up being significant in the final model were: bullying, sex, cigars, weakness, ADD, and anxiety , and anxiety complaints. More research is necessary to ensure the safe and appropriate use of marijuana as a plant-derived medication for adolescents and adults alike. Discovering key clinical symptomatology will aid physicians in identifying adolescents using marijuana. Furthermore, physicians should consider marijuana use in adolescents and young adults with high-risk behaviors presenting with anxiety-related complaints or other common symptoms associated with marijuana use, such as headache, nausea/vomiting, cough, and abdominal pain."}
+{"text": "The last few years have witnessed dramatic advances in our understanding of the structure and function of the mammalian mito-ribosome. At the same time, the first attempts to elucidate the effects of mito-ribosomal fidelity (decoding accuracy) in disease have been made. Hence, the time is right to push an important frontier in our understanding of mitochondrial genetics, that is, the elucidation of the phenotypic effects of mtDNA variants affecting the functioning of the mito-ribosome. Here, we have assessed the structural and functional role of 93 mitochondrial (mt-) rRNA variants thought to be associated with deafness, including those located at non-conserved positions. Our analysis has used the structural description of the human mito-ribosome of the highest quality currently available, together with a new understanding of the phenotypic manifestation of mito-ribosomal-associated variants. Basically, any base change capable of inducing a fidelity phenotype may be considered non-silent. Under this light, out of 92 previously reported mt-rRNA variants thought to be associated with deafness, we found that 49 were potentially non-silent. We also dismissed a large number of reportedly pathogenic mtDNA variants, 41, as polymorphisms. These results drastically update our view on the implication of the primary sequence of mt-rRNA in the etiology of deafness and mitochondrial disease in general. Our data sheds much-needed light on the question of how mt-rRNA variants located at non-conserved positions may lead to mitochondrial disease and, most notably, provide evidence of the effect of haplotype context in the manifestation of some mt-rRNA variants. Aminoglycoside antibiotics (AGs) are some of the most commonly prescribed antibacterials, despite their well-known capacity to cause toxic side effects to the kidneys and inner ear . AlthougOur understanding of the mito-ribosome has advanced much more slowly than that of other ribosomal systems. In particular, the issue of the maintenance of mito-ribosomal fidelity, defined as the accuracy of decoding during mitochondrial translation, could not be tackled until very recently. Despite this, evidence obtained mostly from studies with chimeric mito-bacterial ribosomes, as well as in yeast, has been successfully used to ascertain that decreased mito-ribosomal fidelity was behind the deleterious effect caused by the two known ototoxic variants . The recIn this analysis, we used a total of 83 and 9 variants, identified in deafness patients, that mapped to the mito-ribosomal small and large subunit (SSU and LSU) mt-rRNAs, respectively. The variants originated from 35 studies reported in the literature (80 variants), MITOMAP (2 variants), and this work (1 variant)  Ruiz-Pe. Figure The low conservation of most variants made the comparison to heterologous structures largely useless, implying that the type of evidence used in our previous heterologous inferential analysis (HIA) studies with exceptionally rare mt-rRNA variants could not be raised in this case . In the More than half of the 92 variants  could cause structural distortions, possibly resulting in some degree of defective mito-ribosomal function and constituting valid candidates for deafness-inducing variants under the new theoretical framework for the evaluation of the pathogenic potential of mt-rRNA variants. Forty-one variants, including the nine mapping to 16S mt-rRNA, were deemed likely silent and two unclear . TertiarThis section contains the structural description of all 49 variants regarded as potentially non-silent. Given the number of variants analyzed, most accompanying figures are provided as The existence of pathogenic mutations in several mt-SSU mito-ribosomal proteins is a good indication that variants in mt-rRNA residues interacting with these proteins may also lead to deleterious phenotypes. Early binding proteins are also of particular interest, as they constitute key nucleation sites for the assembly of the mt-SSU. Hence, mt-rRNA variants that interfere with the binding of these proteins could lead to defective mito-ribosomal assembly and other functional defects. All these variants will be discussed in this section.Escherichia coli S12 and mitochondrial MRPS12/uS12m  variant has been detected in several studies targeting hearing-impaired patients (22U (m.669U) base pairs with 281A (m.928A) in h3 of the SSU  and 4 (m.651) of h1 is visible (22U>C (m.669U>C) variant would replace a Watson:Crick U:A base pair with a C\u2022A mismatch at the junction between helices h1 and h3 and in an rRNA region that is surrounded by proteins MRPS12/uS12m and MRPS5/uS5m, the potential for a fidelity mutation is high.The patients . Positio the SSU . A hydro the SSU , in clos the SSU . Positio the SSU , with tw the SSU  is also  the SSU , one of  visible . As the 283G>A (m.930G>A) has been found to be associated with deafness in three studies (283G (m.930G) lies in the single-stranded stretch linking h3 and the functionally important central pseudoknot (helices h1 and h2) to h19 (283G (m.930G) has been modeled 2.39\u00a0\u00c5 away from Arg 47 of MRPS12/uS12m (283G (m.930G) to MRPS12/uS12m makes the 283G>A (m.930G>A) variant constitute a good candidate for a pathogenic residue. However, in light of its abundance in the population  to h19  (Brink e) to h19 . In the ) to h19 , the O6 12/uS12m , whose e12/uS12m . However12/uS12m . Despite139G>A (m.786G>A) rare variant was detected by The 178U>A (m.825U>A) and 180A>G (m.827A>G) are both haplotype markers that were found in association with deafness in several studies (178U (m.825U) and 180A (m.827A) are located in the single-stranded RNA stretch linking helices h5 and h15 (178U (m.825U) and 179A (m.826A) (178U (m.825U) and the N7 of 180A (m.827A) (178U (m.825U) is located \u223c3\u00a0\u00c5 away from the factor, indicating that this region might have an important role during translocation. In addition to this, 180A (m.827A) is an almost universally conserved residue (position 364 in E. coli 16S rRNA) (2 group at position N3 due to the 180A>G (m.827A>G) base change might result in a slight structural distortion of the region, perhaps by interfering with its interaction with helix h3. A similar structural argument could be made for the 178U>A (m.825U>A) variant in that the insertion of a bulkier adenosine at this position could also slightly induce the distortion of the local structure. Hence, the potential for a fidelity variant exists in both cases.The variants  studies . In the  and h15 , a regio and h15 . Several and h15 . Additio(m.826A) . These M(m.826A)   . Notably(m.827A) . The bas(m.827A) , thus ph(m.827A) . Positio(m.827A) . Since t(m.827A) , as judg(m.827A) . Specifi6S rRNA) , althoug295A>G (m.942A>G) variant was found in two patients with hearing loss (295A (m.942A) is located at the 5\u2032 end of h20 (295A>G (m.942A>G), namely, 400A (m.1047A) and 401C (m.1048C), where an A>G and a C>U transition were, respectively, identified in patients with hearing loss (401C (m.1048C) is modeled as part of a water-mediated, hydrogen bonding interaction with one of the phosphate oxygens of NAD (401C (m.1048C) O2\u2032 to NAD O2\u2032 and a hydrogen bond between spermine N1 and the RNA backbone at position 400A (m.1047A) (not shown). 400A (m.1047A) is unpaired and its base stacks onto that of 401C (m.1048C) (295A (m.942A) and 400A (m.1047A) OP1 (295A (m.942A) also forms water-mediated hydrogen bonds with the backbone of h19\u00a0at positions 487\u201388 (m.1134\u201335). Notably, position 487G (m.1134G) is within 3\u00a0\u00c5 of MRPS12/uS12m (295A (m.942A) is the Watson:Crick base pairing formed by positions 294G (m.941G) and 486C (m.1133C)  . A hydro47A) OP1 . The sam12/uS12m . Adjacen295 (m.942) is involved in the stabilization of mito-ribosomal structure via direct (and water-mediated) hydrogen bonds, namely, in the interaction between helices h19, h20 in the neighborhood of protein MRPS12/uS12m. Guaran et al. found the 295A>G (m.942A>G) variant together with the 35delG/35delG allele in the gene GJB2, encoding the protein connexin 26, which is the most common cause of genetic sensorineural deafness in Caucasian patients (295A>G (m.942A>G) variant to the observed symptoms is unclear in this patient. No GJB2 mutations were reported together with 295A>G (m.942A>G) in the other study (400A>G (m.1047A>G) and 401C>U (m. 1048C>U) variants, the situation is uncertain. The base of 401C (m.1048C) only interacts via a water-mediated hydrogen bond with NAD (401C (m.1048C) and NAD is of any relevance to mitochondrial translation is unknown. The abundance of the 401C>U (m. 1048C>U) variant, itself a haplotype marker, clearly indicates that the C>U base change is well-tolerated at this position. In the case of 400A (m.1047A), its base is not involved in any hydrogen-bonding interaction, prompting us to regard it as a silent polymorphism. Despite all this, due to the proximity of 401C (m.1048C) and 400A (m.1047A) to MRPS12, a potentially pathogenic role cannot be completely ruled out for these residues.Of the three variant bases, only patients . Hence, er study . As for with NAD . Whether304G>A (m.951G>A) variant is a haplotype marker that is associated with deafness in several studies (304G (m.951G) is base-paired to 396C (m.1043C) in the distal part of h20 (304G (m.951G). As the G>A base change at position 304 (m.951) would disrupt a base pair interaction next to sites of MRPS12/uS12 that are heterologous equivalents of known fidelity mutations, the potential of the variant to cause a fidelity phenotype is considered high.The  studies . Positiot of h20 , a regiot of h20 , near tht of h20   variant was found in heteroplasmy in a patient with non-syndromic hearing loss while the haplotype marker 460U>C (m.1107U>C) was identified as a potential deafness-causing variant in three subjects from a cohort of aminoglycoside-induced and non-syndromic hearing-impaired Chinese pediatric patients (459C (m.1106C) displays a potential contact with the backbone of protein MRPS37/mS37 (459C>U (m.1106C>U) variant, the results of this loss are uncertain.The patients . In the patients . The basS37/mS37 , which hS37/mS37 . Althoug460U (m.1107U) with the N6 of 478A (m.1125A), two nucleotides away from the central domain pseudoknot that connects helices h19 and h25 of 12S mt-rRNA (shown in red in 460U (m.1107U) O2\u2032 to 290U (m.937U) OP1 is 3.217\u00a0\u00c5, possibly indicating a direct tertiary interaction with the central domain pseudoknot (E. coli and were shown to be deleterious (460U (m.1107U) is not involved in any direct interactions with other residues except for a water-mediated hydrogen bond with position 947A (m.1594G) near the 3\u2032end of the molecule  suggests that the 460U>C (m.1107U>C) variant could be a simple polymorphism (but see below).A tertiary interaction is observed between the backbone at position eudoknot . Mutatioeterious . Despitemolecule . The imp459C>U (m.1106C>U) and 460U>C (m.1107U>C) cannot be ruled out.Considering all of this, as the 5\u2019 side of the central domain pseudoknot is adjacent to h19, which, in turn, wraps around MRPS12/uS12m, it is possible that even small distortions in this region could be transmitted through this protein and result in fidelity defects. As a result, the potential for the creation of slight fidelity phenotypes by the variants During the early assembly, mito-ribosomal proteins MRPS16/bS16m and MRPS18B/mS40 interact with the nascent 12S mt-rRNA transcript  base change was found in two patients with non-syndromic hearing loss (76A (m.723A) is involved in a triple base interaction with the 144G:150C (m.791G:797C) base pair, thus bringing together helices h7 and h12 (145C (m.792C) of h12, where a C>U base change was identified in two patients with non-syndromic hearing loss (145C (m.792C) is canonically base-paired to 149G (m.796G), closing the terminal loop of h12 (The ing loss . Positio and h12 . A seconing loss . Positiop of h12 .76A>C (m.723A>C) variant, the base change would prevent the base triple interaction, while the C>U base change at position 145C (m.792C) would result in a structurally disfavored C:G/U\u2022G exchange at a terminal base pair  and the 98A>G (m.745A>G) variants were identified in a patient with idiopathic, sensorineural hearing loss and a patient with profound hearing impairment, respectively (95U (m.742U) base-pairs to 85A (m.732A) in h7 (98A (m.745A) is involved in a base quadruple interaction involving the 82U:97A (m.729U:744A) base pair and position 83A (m.730A), both of them in h7 (95U (m.742U) and 98A (m.745A) are structurally important in the context of h7 and that this helix is part of the MRPS16/bS16 binding site, a disruptive effect beyond the local structure of h7 cannot be ruled out.The ectively . PositioA) in h7 . Positioem in h7 . Althougem in h7 , known tem in h7 . Given t209A>G (m.856A>G) variant was identified in four patients with non-syndromic hearing loss (209A (m.856A) is base-paired to 183U (m.830U) at the proximal end of helix h15 (209A>G (m.856A>G) variant is considerable.The ing loss . The varing loss  and Lebeing loss . Positioelix h15 . Hydrogeelix h15 . In addielix h15 . The 209During assembly, the 5\u2032 and 3\u2032 domains are brought together by the strong interactions established by the 5\u2032-domain binders MRPS16/bS16m, MRPS18B/mS40, and MRPS22/mS22 and the 3\u2032-domain binders MRPS27/mS27 and MRPS34/mS34 . The imp62G>A (m.709G>A), which has been frequently associated with hearing impairment (63U>C (m.710U>C) and 65C>A (m.712C>A) variants (62G (m.709G) is modeled, making two hydrogen bonds to Arg 48 of MRPS34/mS34 , 53A (m.700A), and 65C (m.712C) stabilizes the structure of the RNA stretch separating helices h6 and h6a and provides a recognition surface for the late-binding protein MRPS26/mS26 (pink in 62G>A (m.709G>A) clearly implies that the variant must be easily accommodated in the SSU structure. Despite this, the G>A base change is expected to disrupt the hydrogen bonding network observed in the structure, involving tertiary and quaternary contacts and likely leading to some degree of distortion. Similarly, the pyrimidine-to-purine replacement caused by the 65C>A (m.712C>A) variant is expected to disrupt the non-canonical base pair between 65C (m.712C) and 53A (m.700A). With this in mind, both variants must be regarded as potentially pathogenic.Three deafness-associated variants have been identified in the single-stranded stretch connecting helices h6 and h6a in the secondary structure map of the mt-SSU, namely, the haplotype defining variant pairment , as wellvariants  (Koningsvariants . The bas pink in  (Bogenha pink in . The abu62G (m.709G) is 63U (m.710U), whose base is stacked between 62G (m.709G) and Arg 30 of MRPS26/mS26 (63U (m.710U) is involved in a water-mediated hydrogen bonding interaction with 62G (m.709G) N7 (U>C base change at position 63 (m.710) is not expected to bring about any disruptive effect, as shown by the fact that a C is found at the equivalent position in the S. scrofa mito-ribosome (not shown) (Adjacent to S26/mS26 . 63U (m.709G) N7 . Despitet shown) .878C>G (m.1525C>G) variation was found by our group in a patient with profound hearing loss (878C (m.1525C) is located at the very distal end of h44 (878C (m.1525C) has been modeled, making a hydrogen bond to the backbone of protein MRPS27/mS27 (878C (m.1525C) would, in principle, argue for a silent phenotype, the rarity of the C>G variant, together with the existence of pathogenic mutations in MRPS34/mS34  (545C>A/U (m.1192C>A/U) (546U>C (m.1193U>C) (815G>A (m.1462G>A) (545C (m.1192C) and 815G (m.1462G) form a Watson:Crick base pair at the distal end of h28 (546U (m.1193U) (545C (m.1192C) and the functionally important \u03b2-hairpin of protein MRPS7/uS7m is less than 3\u00a0\u00c5 (not shown)  , 545C>A/92C>A/U) , 546U>C 1193U>C) , and 8151462G>A) . Positiod of h28 . This red of h28 , which id of h28 . Contactm.1193U) . Notablym.1193U) , adjacenm.1193U) . Additiot shown) .542U>C (m.1189U>C) variant was identified in several independent studies (542U (m.1189U) maps to the single-stranded stretch of h28 (542U (m.1189U) is nearly on the same plane as the 545C:815G (m.1192C:1462G) base pair but slightly tilted, allowing it to use its O2 to form a hydrogen bond with the neighboring 544C:816G (m.1191C:1463G) base pair (542U (m.1189U) O4 to the NH2 of 782C (m.1429C) (542U>C (m.1189 U>C) variant. 542U>C (m.1189U>C) was found together with the 141A>G (m.1811A>G) variant in 16S mt-rRNA and the R127H allele in gene GJB2 (present in heterozygosity) (The  studies . In the h of h28 . In the ase pair . The tilm.1429C) , a contaygosity) , both ofygosity) .In addition to elongation, this region of the mito-ribosome is also implicated in translation initiation. 545C:815G (m.1192C:1462G) base pair and the tertiary interactions of 542U (m.1189U) are important to stabilize the structurally complex and dynamic junction between helices h28 and h29. Of the three variants affecting the 545C:815G (m.1192C:m.1462G) base pair, 545C>A (m.1192C>A) and 815G>A (m.1462G>A) would replace a Watson:Crick base pair with either an A\u2022G or C\u2022A mismatch, while 545C>U (m.1192C>U) would result in a disruptive U\u2022G wobble at the end of a helical segment (546U (m.1193U) makes no hydrogen bonds which, together with the fact that a C is found in the S. scrofa structure at this position (position 544 in structure 5AJ4), strongly argues for a silent phenotype associated with the 546U>C (m.1193U>C) variant.All this evidence strongly suggests that the  segment . For all596U>C (m.1243U>C) was found in association with deafness in several studies (596U (m.1243U) maps to helix h32 of 12S mt-rRNA, where it base pairs to 699A (m.1346A) (596U:699A (m.1243U:1346A) base pair by the U>C base change at position 596U (m.1243T) could affect the interaction of h32 and MRPS14/uS14m in this region. Although the abundance of the 596U>C (m.1243U>C) variation implies that the base change is well-tolerated, whether it could lead to slight phenotypic effects remains uncertain.The haplotype marker  studies . Positiom.1346A) . Arg 36 m.1346A)  forms a 663C>U (m.1310C>U) variant has been identified in a Japanese patient with inherited sensorineural hearing loss (663C (m.1310C) maps to h38 of 12S mt-rRNA, where it base pairs with 668G (m.1315G) (663C (m.1310C) would create a disfavored U\u2022G wobble base pair at the end of a helical segment (The ing loss . Positiom.1315G) . Its O2\u2032m.1315G) . Late-bim.1315G)  also conm.1315G)  and the  segment  with the735A>C (m.1382A>C) variant was identified in two families with hearing loss (735A (m.1382A) is base-paired to 723U (m.1370U) at the proximal end of h41 in the head of the mt-SSU (735A (m.1370U:m.1382A) base pair, the pathogenic potential of this variant must be considered.The ing loss . Positioe mt-SSU . A contae mt-SSU  to the Re mt-SSU  establise mt-SSU . As the 796U>C (m.1443U>C) variant was identified in a non-syndromic hearing-impaired Chinese pediatric subject (796U (m.1443U) maps to the loop that caps h43 of 12S mt-rRNA (796U (m.1443U) establishes a hydrogen bond with the RNA backbone at position 799 (m.1446) (7 (m.1443-4) are visible in the structure. The adjacent position 797A (m.1444A) makes a tertiary interaction by forming a triple base interaction with 585A (m.1232A) of h31 and 753G (m.1401G) of h42 (796U (m.1443U) (not shown) (796U>C (m.1443U>C) must be considered significant.The highly rare  subject . Positio mt-rRNA . The N3 (m.1446) , an inte(m.1446)  and the ) of h42 . This ant shown) . Given a805U (m.1452U) and 806A (m.1453A), presumably associated with hearing impairment, map to h43 of 12S mt-rRNA (805U (m.1452U), while MRPS9/bS9 and MRPS14/uS14m (beige in 806A (m.1453A). Position 805U (m.1452U) forms a wobble base pair with 791G (m.1438G) (791 (m.1438), which is present in only 5% of the GenBank sequences (791/805 (m.1438/1452) base pair. As a result, both the 805U>C (m.1452U>C) and the 791G>A (m.1438G>A) variants should be considered silent, as they would result in the exchange of the wobble configuration seen in the structure for a Watson:Crick base pair. Preceding the 791/805 (m.1438/1452) base pair lies position 806A (m.1453A), which forms a Watson:Crick base pair with 790U (m.1437U) (806A (m.1453A) (The sites of variation  mt-rRNA  (Padma a mt-rRNA . Protein mt-rRNA  contact beige in  do the sm.1438G) , anotherm.1438G) . In factequences . These dm.1437U) . A deafnm.1453A)  that wouThe early binding proteins MRPS2/uS2m and MRPS23/uS23, together with MRPS28/bS1m, bind as a cluster to the solvent side of the 12S mt-rRNA molecule . Pathoge469A>G (m.1116A>G), 471A>G (m.1118A>G), and 472U>C (m.1119U>C) variants were found in four hearing-impaired patients (471A (m.1118A) and 472U (m.1119U) are located in the loop capping h25 (471A (m.1118A) has been modeled within 3\u00a0\u00c5 of Arg 297 of protein MRPS5/uS5m (beige in ram residue V336 within the same protein (not shown) (472U (m.1119U), its base has been modeled, making a water-mediated hydrogen bond, through its O2, to the base and the sugar of 479A (m.1126A) and 480A (m.1127A), respectively (472U (m.1119U) would be unaffected by a U>C base change, thus making 472U>C (m.1119U>C) a good candidate for a silent variant. In this particular patient, the presence of 62G>A (m.709G>A) together with 472U (m.1119U) could offer a better explanation for the observed deafness, as explained previously (469A>G (m.1116A>G) variant is different in that its base is part of a base triple with the 467U:480A (m.1114U:1127A) that would be likely affected by the base change. Additionally, Arg 100 of MRPS2/uS2m (orange in the structure) is within hydrogen bonding distance of the RNA backbone at position 469A (m.1116A). The density around 469A (m.1116A) supports these interactions. Hence, despite the peripheral location of the 469A (m.1116A) variant, the structural evidence does not permit its scoring as a silent mutation.The patients . Positioping h25  toward tbeige in  and \u223c13-t shown) . Despitet shown)  suggestsectively . In addiectively . All theeviously . The sit103 (m.750) is an A in the Cambridge mtDNA reference sequence (103 (m.750) is a G that forms a Watson:Crick base pair to 77C (m.724C) in h7 (103G (m.750G) is visible (103G (m.750G), the introduction of an A\u2022C mismatch at this position in h7 is expected to cause some level of structural distortion whose overall consequences cannot be predicted. Hence, the 103G>A (m.750G>A) base change is another example of a haplotype marker with a potentially pathogenic role.Position sequence . Howeversequence . Base chsequence . In the C) in h7 . A conta visible . Althoug122G>A (m.769G>A) and the 123C>U (m.770C>U) variant were described in the context of deafness in two studies (122G (m.769G) and 123C (m.770C) map to the GNRA tetraloop capping h11 (123C>U (m.770C>U) variant would not change the GNRA consensus, 122G>A (m.769G>A) would do so, likely disrupting the structure of the tetraloop and, perhaps, the binding of MRPS17/uS17m. Given the role of MRPS17/uS17m as an early binding protein, the possibility of disruptive potential in the case of the 122G>A (m.769G>A) variant must be considered.The haplotype marker  studies . Positioping h11  , 358U (m.1005U), 360G (m.1007G), and 361A (m.1008A) map to h23 of 12S mt-rRNA (358U (m.1005U) is involved in a water-mediated, non-canonical base pair with 336C (m.983C) and a quaternary contact with Asn 104 of MRPS11/uS11m (pink in 360G (m.1007G) and 361A (m.1008A) form Watson:Crick base pairs with residues 334C (m.981C) and 333U (m.980U), respectively. Lys 6 of MRPS21/bS21 contacts the RNA backbone at positions 359U (m.1006U) and 360G (m.1007G) (light green in 333U>C (m.980U>C) and 360G>A (m.1007G>A) would both result in a disruptive C\u2022A mismatch, while the 361A>G (m.1008A>G) would give rise to a less damaging U\u2022G wobble. In the case of 358U>C (m.1005U>C), the hydrogen bonding pattern observed in the structure would not be maintained, possibly causing some degree of structural disruption. All variants have been identified in MITOMAP with moderate-to-high frequency (two are haplotype markers) (Positions  mt-rRNA  (Li et a mt-rRNA . The bas pink in . Positiomarkers)  (Ruiz-Pemarkers) , indicat341G (m.988G) and 343U (m.990U) of h23 were found in the studies of 341G>A (m.988G>A) and 343U>C (m.990U>C) would disrupt the base hydrogen bonding pattern observed in the two high-resolution structures used in this analysis  variant was found in several patients with sensorineural and non-syndromic hearing loss (448U (m.1095U) base-pairs to 413A (m.1060A) in h24 of 12S mt-rRNA (448U (m.1060A:1095U) base pair by the U>C transition has the potential to affect the functioning of bridge B7b and cause translational defects. Results of biochemical analyses with cybrids carrying this variant in heteroplasmy were consistent with decreased mitochondrial function and increased AG susceptibility  variant was identified by Rydzanicz et al. in a patient with non-syndromic and aminoglycoside-induced hearing loss and by Chen et al. in a Uyghur patient with non-syndromic deafness (856G (m.1503G) is near the highly conserved bridge mB5, which according to lower-resolution structures of the human mito-ribosome, involves positions 858A (m.1505A) and 892C (m.1539C) in h44 of 12S mt-rRNA and MRPL14/uL14m in the LSU  and 890C>U (m.1537C>U) also map to h44 of 12S mt-rRNA and the neighborhood of mB5 (861C>U (m.1508C>U) has been identified in a patient with profound hearing loss without previous exposure to AGs (890C>U (m.1537C>U) variant, it has been identified in two patients with non-syndromic hearing impairment (890C (m.1537C) establishes two hydrogen bonds, one with the N3 of the adjacent 889A (m.1536A) and the other with the O2\u2032 of 864C (m.1511C) (Variants d of mB5 . Positioe to AGs . This ree to AGs . As for pairment . The amim.1511C) .861C>U (m.1508C>U) or 890C>U (m.1537C>U) variants could affect the proper functioning of mB5. Regarding 856G>A (m.1503G>A), its high frequency in the population clearly indicates that an A must be tolerated at this position. However, the loss of a quaternary interaction involving 856G (m.1503G) and 941U (m.2611T) of 16S mt-rRNA observed in the 2.2-\u00c5 cryo-EM human mito-ribosomal structure  variant was found in homoplasmy in pediatric subjects with non-syndromic hearing loss (533U>G (m.1180U>G) and 827G (m.1474G) of h28 form a wobble base pair (533U\u2022827G (m.1180U\u2022m.1474G) wobble lies position 826C (m.1473C), where a C>U transition was found in a patient with severe hearing loss (826C (m.1473C) forms a Watson:Crick base pair with 534G (m.1181G) (533U (m.1180U) lies the universally conserved 532G (m.1179G)  , and 534G (m.1181G)  in the cytoplasmic ribosome confer lethal phenotypes (533U (m.1180U) and 534G (m.1181G) , compensatory mutations restoring base pairing potential also restored growth (533U>G (m.1180U>G) variant would replace a U\u2022G wobble with a G\u2022G mismatch, its disruptive potential must be considered high. Hence, the use of the new structural data corroborates our previous assignment as an expectedly disruptive mutation. In the case of 826C>U (m.1473C>U), the transition would replace a Watson:Crick G:C with a G\u2022U wobble, creating a highly non-isosteric U\u2022G/G\u2022U tandem wobble combination (826C>U (m.1473C>U) variant is well tolerated, as judged by its high frequency (The very rare ing loss . We haveing loss  and descase pair . Next toing loss . Positiom.1181G) . On the nd 100%) , which ind 100%)   . Strikinl growth , but witin vitro . Mutagenenotypes . In the d growth . As the bination . Althougrequency , subtle 579C>G (m.1226C>G) variant was found in a pediatric subject with non-syndromic hearing loss (579C (m.1226C) is universally conserved (579C (m.1226C) is located at the single-stranded junction connecting helices h31 and h32 (579C (m.1226C) canonically base-pairs with 570G (m.1217G), also universally conserved  and adjacent residues were mutated, they resulted in moderate-to-strong growth defects and elicited fidelity phenotypes (579C (m.1226C)  , we cons579C (m.1226C). The 684A>G (m.1331A>G) base change was identified in one Han Chinese pediatric subject with aminoglycoside-induced and non-syndromic hearing loss (684A (m.1331A) maps to h34, it is brought into the neighborhood of 579C (m.1226C) by a tertiary interaction in the form of a trans-Watson:Crick base pair, with the base of 568U (m.1215U) (684A>G (m.1331A>G) must be considered a potentially pathogenic variant.A second variant maps to the neighborhood of position ing loss . Althougm.1215U)  (Leontism.1215U) . This in910A>C (m.1557A>C) variation was identified in a Russian individual suffering from deafness . Position 910A (m.1557A) maps to a single-stranded stretch of h44 (910A>C (m.1557A>C) variant in our previous analysis of highly rare variants mapping to 12S mt-rRNA, performed in the absence of high-resolution mito-ribosomal structures (910A (m.1557A) and 911A (m.1558A) in monitoring the geometry of A-site decoding is evident from the 2.2-\u00c5 structure of the mito-ribosome  was found in two aminoglycoside-induced hearing-impaired subjects (951G (m.1598G) maps to the single-stranded stretch near the 3\u2032end of the molecule (951G (m.1598G) would impact its interaction with 921U (m.1568U), possibly resulting in a distortion of the mRNA channel and leading to phenotypic effects. These effects, however, must not be drastic, as the 951G>A (m.1598G>A) base change has been found frequently in the population  and 908A>G (m.1555A>G) of 12S mt-rRNA. Positions 847C (m.1494C) and 908A (m.1555A) normally form a C\u2022A mismatch at the penultimate helix of human mt-12S rRNA , which ie trauma . However (76A>C (m.723A>C) variant. Position 76A (m.723A) is involved in a triple base interaction that brings together helices h7 and h12 near the recognition sites for MRPS16/bS16 and MRPS25/mS25, both known to harbor pathogenic mutations (76A>C (m.723A>C) is expected to disrupt this triple base pair interaction, possibly affecting the helical arrangement of part of 12S mt-rRNA and the binding of MRPS16/bS16 and MRPS25/mS25. However, the triple base interaction is not conserved, as it is absent in the S. scrofa mito-ribosome  and 908A>G (m.1555A>G) affect structurally non-conserved positions, forming a mismatch in the human mito-ribosome t-rRNAs) . This tyutations  (Miller utations . The potribosome . This exThe possibility that haplotype context could affect the manifestation of mt-rRNA variants was recently proposed by our group  and has 401C (m.1048C) (401C (m.1048C) and the NAD molecule is of any relevance to mitochondrial translation remains unknown.As mentioned in the introduction, AGs antibiotics are an important class of external agents capable of affecting the penetrance of mt-rRNA variants. HIA studies conducted by our group with highly rare mt-rRNA variants reportedly in association with diseases showed that an important number of them mapped to the vicinity of the binding sites of known ribosomal antibiotics . As a rem.1048C)  (Itoh etm.1048C) . The prem.1048C) . Despite910A>C (m.1557A>C) is a good example, as it has the strongest disruptive potential. Base changes at position 910A (m.1557A) are expected to completely abolish mito-ribosomal translation by directly interfering with genetic decoding (The growing realization in the field that mtDNA disease and, more specifically, deafness can be brought about by variants with low-profile phenotypes, such as those affecting mito-ribosomal fidelity (reviewed in accompanying study by decoding . Unfortudecoding .In summary, we have provided a much-needed assessment of the structural and functional role of suspect mt-rRNA variants identified in the context of deafness. Besides highlighting the key role of the primary sequence of mt-rRNA in the etiology of deafness, this research provides a new framework to understand how mt-rRNA variants, particularly those located at non-conserved positions, may lead to deafness. Hence, the results presented here constitute an important push toward the final elucidation of the role of mt-rRNA in mitochondrial deafness and mitochondrial disease. Such elucidation will eventually require biochemical evidence obtained with mutant mitochondria carrying mito-ribosomes with altered mt-rRNAs.Structural analysis was performed with Chimera X , essenti"}
+{"text": "The Supporting Information file legends are incorrect. Please view the correct Supporting Information file legends below.S1 Fig. CFTR presence and localization in the normal human pancreas. CFTR immunostaining using CFF-217  of normal pancreas from donors of different ages: 2 months (A-B), 6 months (C-D), 2 months (E-F), 7 months (G-H), 30 years (I-J), and 39 years (K-L). CFTR immunoreactivity is shown in ductal and islet endocrine cells. Shown in B, D, F, H and J are consecutive sections immunolabeled by using the pan-endocrine marker synaptophysin (SYN) to confirm CFTR expression in the pancreatic islets. A larger magnification of the section area in the inset in K is shown in L. Scale bars represent 50\u03bcm.https://doi.org/10.1371/journal.pone.0242749.s001(TIF)S2 Fig. Immunohistochemistry for CFTR. Normal human pancreas tissues obtained from three donors aged 6 months , 11 months  and 4 years  are immunostained by using anti-human CFTR antibodies CFF-432 (A-C), CFF-412 (D-F) and CFF-570 (G-i). CFTR expression (brown) occurs in both duct (green squares) and islet endocrine cells (red squares). Higher magnifications of the areas noted by green and white inserts i.e., ductal and islet areas, respectively, are shown under each image. Scale bars represent 100\u03bcm.https://doi.org/10.1371/journal.pone.0242749.s002(TIF)S3 Fig. CFTR antibodies not-suitable for immunohistochemistry. Normal human pancreatic tissues stained with the indicated CFTR antibodies. A. CFF-596 antibody generates non-specific immunostaining in the CFTR-negative control human tissue with the G542X truncation mutation. B-C. CFF-660 (B) and CFF-450 (C) antibodies do not generate specific immunostaining in the positive control ductal cells. D. Antibody Ab4067 does not generate immunostaining signal in the normal human pancreas. E-J. Pancreatic tissue from CftrWT (E-G) and CftrG542X mice (H-J) co-immunolabeled by using the mouse anti CFTR-specific antibody MrPink (J and H), insulin  and glucagon .https://doi.org/10.1371/journal.pone.0242749.s003(TIF)S4 Fig. Original blots used in Fig 4.https://doi.org/10.1371/journal.pone.0242749.s004(TIF)"}
+{"text": "Pseudomonas aeruginosa has not been fully elucidated. This study evaluated the in vitro activity of novel \u03b2-lactam/\u03b2-lactamase inhibitor combinations against Pseudomonas aeruginosa clinical isolates, determined how avibactam restored ceftazidime activity, and compared the activity of ceftazidime-avibactam (CZA) and imipenem-relebactam (IMR) against KPC-producing P. aeruginosa. Similar high susceptibility rates for CZA, IMR, and ceftolozane-tazobactam (88.9% to 89.8%) were found for 596 P. aeruginosa clinical isolates from 11 hospitals in China, and a higher susceptibility rate to ceftazidime than imipenem was observed (73.5% versus 63.1%). For CAZ-NS and IPM-NS isolates, susceptibility rates for CZA, ceftolozane-tazobactam, and IMR were 61.5% (75/122), 54.9% (67/122), and 51.6% (63/122), respectively. For CAZ-NS, IPM-NS but CZA-susceptible isolates, 34.7% (26/75) harbored acquired \u03b2-lactamases with KPC-2 predominant (n\u2009=\u200919), and 45.3% (34/75) presented overexpression of chromosomal \u03b2-lactamase ampC. Among 22 isolates carrying KPC-2 carbapenemase alone, susceptibility rates to CZA and IMR were 86.4% (19/22) and 9.1% (2/22), respectively. Notably, 95% (19/20) of IMR-nonsusceptible isolates had an inactivating mutation of oprD gene. In conclusion, CZA, ceftolozane-tazobactam, and IMR exhibit high activity against P. aeruginosa, and CZA is more active than IMR against CAZ-NS and IPM-NS isolates as well as KPC-producing P. aeruginosa. Avibactam overcomes ceftazidime resistance engendered by KPC-2 enzyme and overexpressed AmpC.The role of novel \u03b2-lactam/\u03b2-lactamase inhibitor combinations in ceftazidime-nonsusceptible (CAZ-NS) and imipenem-nonsusceptible (IPM-NS) IMPORTANCE The emergence of antimicrobial resistance poses a particular challenge globally, and the concept of P. aeruginosa with \u201cdifficult-to-treat\u201d resistance (DTR-P. aeruginosa) was proposed. Here, P. aeruginosa clinical isolates were highly susceptible to three \u03b2-lactamase inhibitor combinations, CZA, IMR, and ceftolozane-tazobactam. The combination of KPC-2 enzyme and nonfunctional porin OprD contributed to IMR resistance in P. aeruginosa, and CZA was more active than IMR in fighting against KPC-2-producing P. aeruginosa. CZA also showed good activity against CAZ-NS and IPM-NS P. aeruginosa, primarily by inhibiting KPC-2 enzyme and overproduced AmpC, supporting the clinical use of CZA in the treatment of infections caused by DTR-P. aeruginosa. Pseudomonas aeruginosa is a major cause of serious infections in humans, which often become notoriously refractory to treatment and lead to considerable morbidity, mortality, and socioeconomic impacts worldwide. In this scenario, \u03b2-lactam antibiotics remain one of the mainstays of therapy. One interesting phenomenon is the higher in vitro susceptibility rate of this pathogen to ceftazidime than to carbapenems  was proposed in 2018  and fluoroquinolones (ciprofloxacin and levofloxacin)  is commonly responsible for resistance to ceftazidime via structural mutations or overexpression. The intrinsic resistance-nodulation-division family of efflux systems  are also critical features associated with \u03b2-lactam resistance  . Mechani diverse . Mutatio diverse . Chromossistance , and theP. aeruginosa strains with inhibitory Ambler class A, C, and some D \u03b2-lactamases. IMR exhibits an antimicrobial spectrum similar to that of CZA except for inhibition activity against OXA-48 enzyme. Ceftolozane, a novel oxyimino-aminothiazolyl cephalosporin, is characterized by its stability in PDC hydrolysis. In previous studies, CZA and CT showed excellent in vitro activity against P. aeruginosa with susceptibility rates of 86.5% and 88.5%, respectively, and IMR demonstrated a susceptibility rate of 90.8% (P. aeruginosa according to the guidance of the Infectious Diseases Society of America (IDSA) , ceftolozane-tazobactam (CT), and imipenem-cilastatin-relebactam (IMR), have recently been approved and cast a new spotlight on the treatment of multidrug-resistant (MDR) Gram-negative bacterial infections. CZA expands the therapeutic coverage of ceftazidime against of 90.8% . These cP. aeruginosa isolates are emerging in China. The resistant gene blaKPC-2 is commonly located in plasmids with mobile elements and is frequently detected in sequence type 463 (ST463) isolates  to 100% (16/16)  , 12. Howin vitro activities of CZA, CT, and IMR against clinical P. aeruginosa isolates collected from multiple medical centers in China and uncovered the extent to which avibactam restored the activity of ceftazidime in ceftazidime-nonsusceptible (CAZ-NS) and imipenem-nonsusceptible (IPM-NS) P. aeruginosa. This study found high in vitro activities of CZA, CT, and IMR against P. aeruginosa clinical isolates. The combination of KPC-2 carbapenemase and nonfunctional porin OprD led to IMR resistance in P. aeruginosa, and CZA was more active than IMR in fighting against KPC-2-producing P. aeruginosa. CZA also retained good activity against CAZ-NS and IPM-NS P. aeruginosa isolates, as avibactam overcame the ceftazidime resistance engendered by acquired \u03b2-lactamase KPC-2 and overproduced chromosomal \u03b2-lactamase AmpC.This study investigated the P. aeruginosa isolates. The MIC50 and MIC90 for CZA were 2\u2009mg/L and 16\u2009mg/L, respectively, and avibactam restored the susceptibility of ceftazidime in 61.4% (97/158) of CAZ-NS isolates. Moreover, higher susceptibility to CAZ than to IPM was observed . The most active agents against 596 P. aeruginosa isolates were colistin  and amikacin (90.8%), and susceptibility rates to the remaining six agents ranged from 58.2% (aztreonam) to 78.2% (cefepime) (P. aeruginosa accounted for 11.9% (71/596).Overall, susceptibility rates to CZA, IMR, and CT were 89.8%, 88.9%, and 88.9%, respectively, among the 596 efepime) . DTR-P. P\u2009<\u20090.001) but relatively weaker correlations between the CZA and imipenem MICs  among the 596 P. aeruginosa clinical isolates  ceftazidime-susceptible (CAZ-S) and imipenem-susceptible (IPM-S) isolates, 98 (16.4%) CAZ-S but IPM-NS isolates, 36 (6.0%) IPM-S but CAZ-NS isolates, and 122 (20.5%) CAZ-NS and IPM-NS isolates  of these were susceptible to CZA  of e to CZA . Among te to CZA . In addiP. aeruginosa isolates displayed nonsusceptibility to imipenem, 78.6% (173/220) of IPM-NS isolates were susceptible to CZA, including 98 (44.5%) CAZ-S isolates and 75 (34.1%) CAZ-NS isolates (While 36.9% (220/596) of isolates .For 122 CAZ-NS and IPM-NS isolates, susceptibility rates for CZA, CT, and IMR were 61.5% (75/122), 54.9% (67/122), and 51.6% (63/122), respectively.To illuminate how avibactam overcomes ceftazidime resistance, mechanisms of ceftazidime nonsusceptibility were investigated using whole-genome sequencing (WGS) and quantitative real-time PCR (qRT-PCR) among 75 CAZ-NS, IPM-NS but CZA-S isolates .n\u2009=\u200919), VIM-2 (n\u2009=\u20091), and GES-5 (n\u2009=\u20091), as well as coexistence of GES-5 and PER-1 (n\u2009=\u20091). The VIM-2-carrying isolate had a CZA MIC of 8\u2009mg/L. Extended-spectrum \u03b2-lactamases (ESBL) were detected in four isolates (excluded the one coexisting with GES-5): PER-1 (n\u2009=\u20092) and OXA-101 (n\u2009=\u20091), as well as coexistence of PER-1 and OXA-101 (n\u2009=\u20091) (Table S2).Among these 75 isolates, 26 (34.7%) carried horizontally acquired \u03b2-lactamases  and maniampC overexpression phenotype  were witnessed in 21 isolates, including a premature stop codon (n\u2009=\u20096) and amino acid substitution G154R (n\u2009=\u20093) in ampR, a frameshift mutation (n\u2009=\u20097) in ampD, and a premature stop codon (n\u2009=\u20091), frameshift mutation (n\u2009=\u20093), and amino acid substitution T428P (n\u2009=\u20091) in dacB. A total of 17 subtypes of PDC were present in the 75 isolates (Table S3), with PDC-8 (n\u2009=\u200918) being the most common, followed by PDC-5 (n\u2009=\u200916), PDC-1 (n\u2009=\u20098), and PDC-3 (n\u2009=\u20096). Additionally, overexpression of the efflux pump gene mexAB-oprM was detected in 35 isolates  isolates, including frameshift mutations (n\u2009=\u200953) and premature stop codons (n\u2009=\u200912) (Inactivating mutations in (n\u2009=\u200912) .blaPER-1. These isolates did not harbor carbapenemases, nor did they show inactivating mutations in oprD. In total, 59.1% (13/22) of isolates displayed ampC overexpression, and 50.0% (11/22) of isolates exhibited alteration of regulatory factors contributing to ampC overproduction, including a premature stop codon (n\u2009=\u20091), frameshift mutation (n\u2009=\u20091), and amino acid substitution D135N/Y (n\u2009=\u20094) in ampR, a premature stop codon (n\u2009=\u20094) and frameshift mutation (n\u2009=\u20091) in ampD, and a premature stop codon (n\u2009=\u20091) in dacB. A total of 10 subtypes of PDC were detected in these 22 isolates (Table S3), with PDC-3 (n\u2009=\u200910) being the most prevalent, followed by PDC-8 (n\u2009=\u20092), PDC-16 (n\u2009=\u20092), and PDC-63 (n\u2009=\u20092).Among the 22 CAZ-NS, IPM-S, and CZA-S strains, only one isolate carried an ESBL-encoding gene, n\u2009=\u200917), ST16 (n\u2009=\u20092), ST238 (n\u2009=\u20091), ST244 (n\u2009=\u20091), and an undefined ST (n\u2009=\u20091). Notably, their susceptibility rates to CZA, CT, and IMR were 86.4% (19/22), 0%, and 9.1% (2/22), respectively.Among the 158 CAZ-NS isolates, 22 carried KPC-2 carbapenemase and did not carry metallo-\u03b2-lactamase. They were assigned to five STs, ST463  of IMR-nonsusceptible isolates had an inactivating mutation of the imipenem . The oneimipenem .P. aeruginosa clinical isolates were highly susceptible to three \u03b2-lactam/\u03b2-lactamase inhibitor combinations, CZA, IMR, and CT. In total, 89.8% of P. aeruginosa clinical isolates from China were susceptible to CZA with MIC50/MIC90 of 2/16\u2009mg/L, and avibactam restored the susceptibility to ceftazidime in 61.4% of CAZ-NS isolates. Similar results were found in a study in Spain, in which antimicrobial activity of CZA was demonstrated against 1,445 P. aeruginosa isolates, with a susceptibility rate of 94.2% and MIC50/MIC90 of 2/8\u2009mg/L  was found for 2,623 P. aeruginosa isolates from Asia, and relebactam restored imipenem susceptibility from 0% to 64.6% for 805 IPM-NS isolates (P. aeruginosa (in vitro activity (88.9%) was also observed for CT in the 596 isolates, with an MIC50/MIC90 of 0.5/8\u2009mg/L. Nevertheless, variation was noted in the susceptibility to CT in different regions, which ranged from 89.1% in Europe to 98.2% in North America  of IPM-NS isolates being susceptible to ceftazidime. Susceptibility  to ceftazidime was also reported for carbapenem-resistant P. aeruginosa (CRPA), as evidenced by data from the Antimicrobial Testing Leadership and Surveillance (ATLAS) program in the Asia-Pacific region from 2015 to 2019 . A global study reported a susceptibility rate to CZA of 75.9% for 29 KPC-2-positive clinical P. aeruginosa isolates, while another group from China reported a CZA susceptibility rate of 49.7% for 151 KPC-2-positive clinical P. aeruginosa isolates . Therefore, the combination of KPC-2 enzyme and nonfunctional porin OprD leads to IMR resistance in P. aeruginosa , and CZA is more active than IMR in fighting against KPC-2-producing P. aeruginosa.Among 22 isolates carrying KPC-2 carbapenemases alone, 95% (19/20) of IMR-nonsusceptible isolates had an inactivating mutation of the imipenem  and furtimipenem . AdditioP. aeruginosa, supporting the clinical use of CZA in the treatment of infections caused by DTR-P. aeruginosa. We also found that 45.3% (34/75) of isolates demonstrated overexpression of AmpC in CAZ-NS and IPM-NS but CZA-S isolates, indicating that overexpressed AmpC could be inhibited by avibactam. Consistently, the addition of avibactam increased ceftazidime susceptibility for 46 AmpC-hyperproducing clinical isolates from 10.9% to 76.1%  , few of which could contribute to CZA resistance  were active against the majority of P. aeruginosa isolates were collected from 11 hospitals across China from July 2018 and February 2019 (Table S4) (P. aeruginosa isolates were predominantly from respiratory tract specimens (61.7%), followed by urinary tract (12.6%) and blood specimens (4.0%).A total of 596 nonduplicate clinical able S4) . The 596P. aeruginosa  (www.eucast.org). P. aeruginosa ATCC 27853 served as a quality control strain.Antimicrobial susceptibility was tested by the agar dilution method, except that antimicrobial susceptibility to colistin was tested by the broth microdilution method. Susceptibility to all antimicrobial agents was interpreted according to the Clinical and Laboratory Standards Institute (CLSI) breakpoints , except https://github.com/ablab/spades) after trimming. The presence of resistance genes was explored using the Comprehensive Antibiotic Resistance Database (CARD) webtool (https://card.mcmaster.ca/home). Multilocus sequence typing (MLST) was ascertained according to the guideline on P. aeruginosa on the MLST website (https://pubmlst.org/organisms/pseudomonas-aeruginosa). A core genome phylogenetic tree was generated using kSNP and visualized using iTOL  was performed after extraction from a fasta file.The CAZ-NS isolates were sent for WGS performed with an Illumina NovaSeq system. The raw reads were assembled using SPAdes  were determined by qRT-PCR for clinical isolates as described previously (rpsL as the internal reference. Isolates were considered to overexpress AmpC, MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY-OprM when the transcriptional levels of ampC, mexB, mexC, mexE, and mexY were at least 10-, 3-, 10-, 10-, and 10-fold higher, respectively, than those of the PAO1 strain (The transcription levels of the gene eviously . Briefly1 strain .Data were analyzed and visualized using R software v4.1.3. Spearman rank-order correlation was used to evaluate associations between antibiotic MICs.PRJNA891673.The genomic sequence data have been submitted to the NCBI GenBank database with BioProject number"}
+{"text": "A recent review indicated that physical activity (PA), facilitated by organisations/clubs, may reduce alcohol consumption in early to mid-adolescence. Our study aims were to examine these factors, and identify how health determinants may influence association.2 tests achieved p<.10.Cross-sectional secondary data analysis using UK cohort data from ALSPAC. Ages and sample-size: Time-point (TP) 1: 13-14 (n\u2009=\u20091824), TP2: 15-16 (n\u2009=\u20091334). Variables: minutes per day spent in moderate to vigorous PA\u2009>\u2009=3 days of accelerometer wear. Ancillary PA variables: Club-type (CT); and frequency of attending club (FAC) collected age 15 only. Risk of alcohol-related harm (RARH) categorised as: no current risk, increasing risk, and at risk (AR). Ordinal regression was conducted at each TP, using PA as a covariate. Explanatory variables (EVs): Psychosocial health (PSH) - Cluster membership numbers generated through K-means cluster analysis (uniquely represented at each TP); Socioeconomic status (SES); Educational attainment; BMI; Smoking status, and Gender. EVs entered into regression model if preliminary Xp<0.002). At TP2: OR 1.24 . TP1: CT (age 15) was not statistically significant at age 13. However, \u2018sports club only\u2019 (SCO), had greatest RARH  vs. no CT. All EVs retained statistical significance at p<.05, excepting IDACI (p\u2009=\u2009.854) and gender (p\u2009=\u2009.917). TP2: Although CT was not statistically significant, SCO had the greatest RARH . RARH was reduced if attending club most evenings vs. no attendance . Statistically significant EVs were PSH  and smoking . Model fit \u03c72 331.010, df 11, p<0.001. Pearson .455. Deviance .218. Nagelkerke .251. -2LL test of parallel lines with significance .321.Regression showed a positive association between PA and RARH at both TPs. Odds of being AR at TP1 were 1.31 greater for each 30\u2009minute increase in PA (95%CI 1.10-1.57; The relationship between PA and alcohol consumption is complex. While facilitated PA can provide many benefits for adolescents, potential unwanted consequences may be an increase in risk-behaviours like alcohol consumption. Further research is needed for greater comprehension of this association."}
+{"text": "Kalamiella piersonii clinical isolate URMC-2103A041 from human bacteremia was determined using the hybrid assembly of short- and long-read sequencing chemistry. The genome contains a 3.93 Mb chromosome, three circular plasmids, and one linear plasmid.The complete genome of  Kalamiella piersonii is a Gram-negative Enterobacterales in the Erwiniaceae family. First described from the International Space Station environment, a recent phylogenomic analysis of Erwiniaceae has proposed reclassification as Pantoea piersonii comb. nov ]. Biochemical identification  produced Pantoea spp. (98.2% identification). The BD Phoenix NID panel identification was Shigella flexneri. Identification was resolved by 16S rRNA V1-V3 sequencing (MicroSeq 500 16S rDNA Sequening Kit) . Susceptibility was determined using zone diameter breakpoints for Enterobacterales   identified the organism as Pantoea spp., URMC-2103A041 was Voges-Proskauer- and ONPG-positive, fermented mannitol, arabinose, and rhamnose, but was arginine dihydrolase positive . DNA was extracted following the protocol for \u201cPretreatment for Gram-Negative Bacteria\u201d . DNA shearing and size selection were not performed. DNA was quantified with the QuantiFluor dsDNA system (Promega).For short reads, paired-end sequencing following library preparation  was performed on a MiSeq instrument . Unless otherwise specified, sequences were analyzed with tools available on the Galaxy server , using default parameters . Reads w50: 11,125 bp) for hybrid assembly  and polishing (Quast v5.2.0) , SQK-RBK004]. Long-read libraries were sequenced , and base calling was performed with Guppy v6.1.5 (MinKNOW v22.05.5). Filtering  generated 93,023 reads (read N v5.2.0) , 14.CP115895.1), three circular plasmids , and one linear plasmid . Annotation was added by the NCBI Prokaryotic Genome Annotation Pipeline (The hybrid assembly genome was 4,990,348 bp with 57.02% GC content; it consisted of one circular chromosome (3.93 Mb, GenBank: Pipeline 17."}
+{"text": "We conducted surveillance of respiratory syncytial virus (RSV) genomic sequences for 100 RSV-A and 27 RSV-B specimens collected during November 2022\u2013April 2023 in Arizona, USA. We identified mutations within prefusion F-protein antigenic sites in both subtypes. Continued genomic surveillance will be critical to ensure RSV vaccine effectiveness. Respiratory syncytial virus (RSV), an RNA virus of the family Pneumoviridae, causes acute respiratory infections, primarily in children, adults with severe lung disease, and elderly persons , panel AThe 2 major RSV subtypes, RSV-A and RSV-B, have distinct antigenic characteristics in the P, N, F, and G proteins , panel Bhttps://www.illumina.com). We used Trim Galore version 0.6.10  to quality filter and adaptor trim sequencing reads, mapped the reads to RSV-A and RSV-B reference sequences (GISAID accession nos. EPI_ISL_412866 and EPI_ISL_165399) using Burrows-Wheeler Aligner version 0.7.17-r1188 (https://bio-bwa.sourceforge.net), and generated consensus sequences using SAMtools version 1.17 (https://github.com/samtools/samtools/releases). We assembled 92 RSV-A (GA2.3.5 genotype) and 24 RSV-B (GB5.0.5a genotype) complete genome sequences and 8 RSV-A (GA2.3.5) and 3 RSV-B (GB5.0.5a) partial genome sequences  . We performed genomic surveillance to determine RSV strains circulating in Arizona during the 2022\u20132023 season. We performed 2 \u00d7 150-bp paired-end next-generation sequencing using a hybrid capture enrichment panel  , panel Ahttps://clades.nextstrain.org) showed that RSV-A sequences from Arizona clustered in clade GA2.3.5, indicating >3 independent introductions of RSV-A into Arizona  is monovalent and Abrysvo , bivalent (The US Food and Drug Administration has approved 2 RSV vaccines for persons bivalent . RSV-B Sbivalent  Figure 2Although RSV remains a substantial clinical burden, approved RSV vaccines reduce the risk of lower respiratory tract illness. By tracking RSV evolution, we can improve design of vaccine formulations to improve effectiveness. Our study was limited because we lacked understanding of potential functional consequences from mutations in F-protein antigenic sites. Although the G protein is under greater selective pressure and has higher mutation rates (Additional information about a study of respiratory syncytial virus in Arizona, USA, 2022\u20132023"}
+{"text": "Escherichia coli strains isolated from the recto-anal junction of slaughter-age Irish sheep. In silico serotyping and genome analysis determined that each of the strains harbored a Shiga-toxin subtype, a complete locus of enterocyte effacement, and a rare O-island 122.This study describes the hybrid genome assemblies of four Shiga toxin-producing  Escherichia coli (STEC) are significant foodborne zoonotic pathogens with primary virulence determinants being the bacteriophage encoded Shiga toxins (Stx) and the intimin adhesion protein  and trimmed using Porechop (v0.2.4) for ONT or Fastp (v0.20.1) (de novo hybrid assemblies were constructed with Unicycler (v0.4.9)  with 16 mg/L of novobiocin (Oxoid) and incubated at 41.5\u00b0C for 24 h. RT-PCR was performed on DNA extracted using InstaGene matrix (Bio-Rad) as described by Macori et al.  to confiv0.20.1)  for Illu(v0.4.9)  and rang(v0.4.9) .https://github.com/phac-nml/ecoli_vf) and a customized database containing nle gene sequences (https://github.com/macgenomegue/nle_database). Protein-coding sequences, tRNAs, and insertion sequence elements were predicted using Prokka (1.14.6) (Genome annotation was conducted using the ABRIcate tool (v1.0.1) , and vir(1.14.6) , tRNA-sc(1.14.6) , and ISE(1.14.6) , respect(1.14.6)  webtool."}
+{"text": "Staphylococcus aureus infections.As daptomycin (DAP) CL does not scale proportionally with weight, fixed-dose DAP has been proposed to optimize efficacy and mitigate toxicity. This study evaluated systemic exposure of a fixed dose (750 mg) of DAP in patients with S. aureus infection were eligible for enrollment. DAP 750 mg was infused over 30 minutes every 24 hours. At steady state, concentrations were obtained pre-dose (0.5 hours) and 1, 12, and 24 hours after the start of infusion. Concentrations were measured using a validated LC-MS assay. Pharmacokinetic (PK) analysis was performed using first-order equations and stratified by weight: low (< 65 kg), normal (65-85 kg), and high ( > 85 kg). Therapeutic AUC24 (\u2265 666 mg*hr/L) and elevated trough (Cmin > 24.3 mg/L) were the efficacy and safety targets, respectively.Adult non-ICU patients with CrCl \u2265 30 mL/min admitted to our academic medical center warranting DAP for a e 0.078 h-1 , CL 0.87 L/h , AUC24 861 mg*hr/L , and Cmin 10.5 mg/L . Therapeutic AUC24 was achieved in 14/22 (64%) and elevated Cmin occurred in 7/22 (32%), with no differences seen among weight groups. However, differences were noted between males (n=10) and females (n=12) with significantly higher CL (median [IQR]) noted in males  vs. females  (p=0.027). Hence, median [IQR] AUC24 was significantly lower in males at 594 mg*hr/L  vs. 1513 mg*hr/L  in females (p=0.021). Therapeutic AUC24 was achieved in 40% of males versus 83% of females, while elevated Cmin was seen in 20% of males and 32% of females.Overall, 22 patients  were enrolled with no differences observed in baseline characteristics . PK parameters (median [IQR]) did not differ by weight with Vd 9.9 L , K24 and Cmin) was not impacted by weight. These results support fixed, rather than weight-based, dosing of DAP in non-ICU patients with normal renal function. Notably, gender differences were observed with males having significantly higher CL, likely warranting higher doses of DAP to ensure optimal exposure.DAP PK and exposure : Grant/Research Support"}
+{"text": "Streptomyces sp. strain ICN988, an actinomycete isolated from Gorgonia. The assembled genome consists of 6,122,654 bp with a GC content of 73%. A comprehensive analysis revealed 19 biosynthetic gene clusters.This study elucidates the draft genomic sequence of  Actinomycetes produce many secondary metabolites, including antibiotics, antitumor agents, and immunosuppressive compounds, and are accountable for over half of the antibiotics used clinically . Marine Streptomyces sp. strain ICN988 was isolated from a Gorgonia sea-fan by-catch collected from Colachel coastal area, Tamil Nadu, India (8\u00b010'22.0\"N 77\u00b014'55.2\"E) on 28 December 2018. Initially washed with seawater to remove surface deposits, Gorgonia\u2019s holdfast was scraped aseptically, and the sand was suspended in 2% NaCl to prepare serial dilutions, then spread on Actinomycete Isolation Media (AIM) agar plates (supplemented with filter-sterilized 10 \u00b5g/mL nystatin and 50 \u00b5g/mL cycloheximide). Streptomyces sp. strain ICN988 (S5K) was one of the actinomycetes found after 7 days of incubation at 30\u00b0C. Genomic DNA was isolated from 5-day-old axenic ICN988 aerial mycelia using the NucleoSpin Tissue Kit (Macherey-Nagel). The 16S rRNA was PCR amplified with 63F forward primer and 1387R reverse primer. The actinomycete\u2019s 16S rRNA gene (GenBank: OP585661) was closely related (98.34% similarity) to Streptomyces ardesiacus NBRC 15402 (GenBank: NR_112454) via NCBI-BLAST and (96.60% similarity) to Streptomyces coelicoflavus NBRC 15399 (GenBank: AB184650) via EzBioCloud database . Sequence libraries were created using Nextera XT DNA Library Preparation Kit (Illumina). The whole-genome sequencing was obtained using Illumina HiSeq 4000 System (paired-end 150 bp). A total of 18,237,528 bp reads were obtained with 2,753,866,728 total read bases. The GC content was 71.01%, and Q30 was 88.73%. The tools such as Trimmomatic (v 0.36) for adapter trimming  (The genomic DNA was extracted from a 7-day-old AIM agar plate culture using HiPurA trimming  utilized(v3.0.2)  revealed(v3.0.2)  identifiStreptomyces sp. strain ICN988 was antagonistic to methicillin-resistant Staphylococcus aureus ATCC 33591 in agar-plug and metabolic extract disc diffusion assays.Through antiSMASH v7.0.0beta1-67b538a9 , 13, 30"}
+{"text": "Correction:J Exp Clin Cancer Res\u00a042, 169 (2023)https://doi.org/10.1186/s13046-023-02723-zFollowing publication of the original article , an erro6*, Youtao Xu1* and Rong Yin1,4,5,7*Guoren ZhouThe correction do not affect the overall Conclusion of the article. The original article has been corrected."}
+{"text": "Bacillus thuringiensis SS2 consists of 426 contigs assembled at the scaffold level, totaling 5,030,306\u2009bp, and contains 5,288 putative PATRIC protein-coding genes, including genes responsible for total benzoate consumption, degradation of halogenated compounds, heavy metal tolerance/resistance, biosynthesis of secondary metabolites, and microcin C7 self-immunity protein.The draft genome sequence of strain  Bacillus thuringiensis is a Gram-positive, aerobic, rod-shaped, mesophilic bacterium that has been reported to metabolize persistent organic pollutants (POPs), pesticides, and heavy metals (B. thuringiensis SS2 was identified during a screen for dioxin-degrading bacteria from dibenzofuran (DF)-polluted waste dump soil obtained with a site with a history of incineration in Cele, Isolo, Lagos, Nigeria , and 2,7-dichlorodibenzo-p-dioxin  (MH714859) and biochemical characterization (7-diCDD) . It was rization .B. thuringiensis SS2 was extracted using the Quick-DNA fungal/bacterial miniprep kit , from a Luria-Bertani culture that had been grown overnight at 28\u00b0C. The DNA was quantified using a QuantiFluor double-stranded DNA (dsDNA) system with a Qubit 3.0 fluorometer. Whole-genome sequencing (WGS) was performed through Illumina NextSeq paired-end sequencing. The library preparation and 2\u2009\u00d7\u2009150-bp paired-end sequencing were performed using the Nextera XT Index kit v2 (FC-131-2001 to FC-131-2004) and a NextSeq 500 system  at Quadram Institute Bioscience  , forming 426 contigs . The longest contig was 120,678\u2009bp, with an N50 value of 22,410\u2009bp. RASTtk identified 5,288 PATRIC protein-coding sequences and 1,174 hypothetical proteins, 66 tRNA genes, and 4 rRNA genes; the closest relative for B. thuringiensis SS2 is B. thuringiensis 97-27 (serovar konkukian).The genome sequence of B. thuringiensis SS2 revealed putative genes linked to the degradation of dioxin and other POPs similar to those of Serratia marcescens SSA1, which was isolated from the same burnt dump site. These genes include putative genes for total consumption of benzoate via hydroxylation and halogenated aromatic compound  degradation, among others  and copper-translocating P-type ATPase (EC 3.6.3.4) genes, suggesting heavy metal tolerance and resistance. Other putative genes, including those for microcin C7 self-immunity protein (mccf), macrocin O-methyltransferase, and CAAX amino-terminal proteins, could indicate this strain\u2019s diverse metabolic adaptations to stress in its environment.WGS of g others . B. thurB. thuringiensis SS2 at PATRICbrc (BV-BRC) can be viewed at https://www.bv-brc.org/view/Genome/1428.1921. The draft genome sequence was deposited in GenBank under the accession number JAOWLZ000000000; project data are available under BioProject accession number PRJNA888024, with BioSample accession number SAMN31205655 and SRA accession number SRR21870348. The 16S rRNA gene sequence is available under GenBank accession number MH714859.The genome annotation of"}
+{"text": "Microsphaeropsis, taxonomically classified within the kingdom fungi, phylum Ascomycota, subphylum Deuteromycotina, class Coelomycetes, order Sphaeropsidales, and family Sphaeropsidaceae, exhibit a ubiquitous distribution across various geographical regions. These fungi are known for their production of secondary metabolites, characterized by both structural novelty and potent biological activity. Consequently, they represent a significant reservoir for the advancement of novel pharmaceuticals. In this paper, a systematic review was present, marking the analysis of secondary metabolites synthesized by Microsphaeropsis reported between 1980 and 2023. A total of 112 compounds, comprising polyketones, macrolides, terpenoids, and nitrogen-containing compounds, were reported from Microsphaeropsis. Remarkably, among these compounds, 49 are novel discoveries, marking a significant contribution to the field. A concise summary of their diverse biological activities was provided, including antibacterial, antitumor, and antiviral properties and other bioactivities. This analysis stands as a valuable reference, poised to guide further investigations into the active natural products derived from Microsphaeropsis and their potential contributions to the development of medicinal resources. Microsphaeropsis, taxonomically classified within the phylum Ascomycetes, subphylum Deuteromycotina, class Coelenterata, order Sphaeropsidales, and family Sphaeropsidaceae in the fungal taxonomy [Microsphaeropsis was not accurately recognized, leading to the misplacement of many fungi within the genus Coniothyrium. In 1980, Sutton meticulously elucidated the characteristics of the Microsphaeropsis genus. He subsequently reclassified species previously assigned to the genus Coniothyrium under Microsphaeropsis and renamed the genus [Microsphaeropsis have been unearthed and taxonomically elucidated. As of now, the repository of the Species Fungorum database (https://speciesfungorum.org/Names/Names.asp (accessed on 2 November 2023)) registers a total of 54 species within the genus Microsphaeropsis. It is evident that fungi within the genus Microsphaeropsis exhibit remarkable biodiversity, with a high likelihood of containing structurally novel and biologically active secondary metabolites.taxonomy , are comtaxonomy . For a lhe genus ,4. SubseMicrosphaeropsis genus by Sutton in 1980, various structural categories of compounds, including polyketides, terpenoids, nitrogen-containing compounds, macrolides, and others, were isolated from 2 known and 17 unknown species of Microsphaeropsis, and these demonstrated excellent antibacterial, antifungal, cytotoxic, and other activities  and three known compounds: (4S)-4,8-dihydroxy -3,4-dihydro-1(2H)-naphthalen-1-one (2), (4S)-4,6,8-trihydroxy-3,4-dihydro-1(2H)- naphthalen-1-one (3), and chrysogeside D (94). Notably, compounds 3 and 94 were discovered for the first time in Microsphaeropsis. The research team conducted evaluations of the antifungal activity of these compounds. Compound 1 and compound 94 exhibited moderate antifungal activity against A. tenuissima with minimum inhibitory concentrations (MICs) at 64 \u03bcg/mL [Qin et al. 2017) successfully isolated four compounds from the endophyte fungus  successf77), along with the known compounds (R)-mellein (8), -hydroxymellein (9), -hydroxymellein (10), and 4,8-dihydroxy-3,4-dihydro-2H-naphthalen-1-one (11), were successfully isolated from the fungus Microsphaeropsis sp. H5-50, which inhabits the marine sponges M. incrustans. Notably, both compounds displayed significant antifungal activity against E. repens and U. violacea in agar diffusion assays [The novel compound microsphaeropsisin M. arundinis PSU-G18, found within the plant Garcinia hombroniana [110), and seven new phthalides, microsphaerophthalides A-G (29\u201335), were identified, alongside eleven known compounds . Compound 44 exhibited remarkable antifungal activity, with an IC50 value of 8 \u03bcg/mL. Additionally, microsphaerophthalides A (29) and microsphaerophthalides E (33) displayed moderate antifungal activity against M. gypseum SH-MU-4 and C. neoformans, both with MIC values of 64 \u03bcg/mL. Decaspirone (57), isolated from the endophyte fungus Microsphaeropsis sp. 7291, demonstrated significant antifungal activity against M. violaceum, evidenced by inhibition zone diameters measuring 18 mm [Sommart et al. (2012) isolated nineteen compounds from the endophyte fungus broniana . Among t112), was successfully isolated from the endophyte fungus Microsphaeropsis sp. 7820, found inhabiting the plant Salsola oppositifolia. Remarkably, compound 112 exhibited moderate antifungal activity against M. violaceum, resulting in inhibition zone diameters measuring 9 mm [A novel polychlorinated triphenyl diether, microsphaerol (ing 9 mm .74) and microketides B (75), from the fungus Microsphaeropsis sp. RA10-14, and both compounds exhibited significant antifungal activity against Candida albicans, Colletotrichum truncatum, Gloeosporium musarum, and Pestalotia calabae [74) exhibited IC50 values of 1.56 \u03bcg/mL, 1.56 \u03bcg/mL, 3.13 \u03bcg/mL, and 1.56 \u03bcg/mL, respectively. Microketides B (75) displayed equal IC50 values of 3.13 \u03bcg/mL.Liu et al. (2020) isolated two novel metabolites, microketides A , were isolated from Microsphaeropsis sp. F2076 and Microsphaeropsis sp. F2078. Notably, the four microsphaerins, A, B, C, and D, exhibited significant antibacterial activity against Methicillin-resistant Staphylococcus aureus (MRSA), with IC50 values of 3 \u03bcM, 3 \u03bcM, 5 \u03bcM, and 1 \u03bcM, respectively. To delve deeper into their antibacterial activity, microsphaerins D (48) was selected for further investigation, encompassing Gram-positive and Gram-negative bacteria. Results revealed that it demonstrated moderate antibacterial activity against various Gram-positive bacteria; however, it displayed inactivity against Gram-negative bacteria, except for K. pneumoniae [Four metabolites, namely microsphaerins A\u2013D (eumoniae .1 (56), palmarumycin M2 (58), papyracillic acid C (61), microsphaeropsins A (78), and microsphaeropsins B (62), were successfully isolated, alongside three known compounds, decaspirone (57), papyracillic acids A (59), and papyracillic acids B (60), from the endophyte fungus Microsphaeropsis sp. 7291, which resides within the plant Pilgerodendron uviferum. Of significance, compounds 56, 57, and 58 exhibited moderate antibacterial activity against B. megaterium, resulting in inhibition zone diameters of 6 mm, 6 mm, and 7 mm, respectively [Five new metabolites, namely palmarumycin Mectively .Microsphaeropsis sp. 8875 residing in the plant Lycium intricatum [64\u201366), in addition to two known compounds, citreorosein (67) and emodin (68). Moreover, from a Microsphaeropsis sp. 7177 strain sourced from the plant Zygophyllum fortanesii, they identified two new metabolites, 3,4-dihydrofusidienol A (70) and microsxanthone (73), along with three known compounds, fusidienol A (69), 8-hydroxy-6-methyl-9-oxo-9H-xanthene-1-carboxylic acid methyl ester (71), and methyl 3,8-dihydroxy-6-methyl-9-oxo-9H-xanthene-1-carboxylate (72). Remarkably, all these compounds exhibited significant antibacterial activity, particularly against the Gram-negative bacterium E. coli. Microsphaerol (112), derived from the endophyte fungus Microsphaeropsis sp. 7820, displayed moderate antibacterial activity against B. megaterium and E. coli, evidenced by inhibition zone diameters measuring 9 mm and 8 mm, respectively [Krohn et al. 2009) conducted a comprehensive study resulting in the isolation of several metabolites from the endophyte fungus 09 conducectively . 74) and microketide B (75), were successfully isolated from the fungus Microsphaeropsis sp. RA10-14, which inhabits the gorgonian Anthogorgia ochracea from the South China Sea. Both compounds exhibited significant antibacterial and antifungal activities, with a notable emphasis on their antibacterial effects. Specifically, compound 74 demonstrated significant antibacterial activity against B. subtilis, B. megaterium, E. aerogenes, K. rhizophila, and P. aeruginosa, all with equal MIC values of 0.19 \u03bcg/mL, mirroring the potency of ciprofloxacin [Two novel metabolites, microketide A , was isolated alongside seven known compounds from the fungus Microsphaeropsis sp. BCC 3050. This fungal strain was collected from the lichen Dirinaria applanata. The known compounds in this collection include preussomerins E (18), F (19), G (20), H (21), I (17), deoxypreussomerins A (22), and bipendensin (23). Remarkably, all compounds, except compound 23, exhibited activities against M. tuberculosis, and compounds 16, 17, and 18 showed moderate antibacterial activity, with IC50 values of 25 \u03bcg/mL, 12.5 \u03bcg/mL, and 25 \u03bcg/mL, respectively. Compounds 19, 20, 21, and 22 showed significant antibacterial activity, with IC50 values of 3.12 \u03bcg/mL, 3.12 \u03bcg/mL, 6.25 \u03bcg/mL, and 1.56 \u03bcg/mL, respectively [94), sourced from the endophyte fungus Microsphaeropsis arundinis PH 30472, showed moderate antibacterial activity against E. coli [The new isomer, 3\u2032-O-demethylpreussomerin I ((21), I 1, deoxypr79), B (80), and C (81), were isolated alongside three known compounds from the endophyte fungus M. arundinis E12-2112, which resides within the plant Ulmus macrocarpa. Among the known compounds, one isochroman-1-one, arundinone A (27), a polyoxygenated benzofuran-3(2H)-one dimer, arundinone B (28), and 1\u03b2-hydroxy-\u03b1-cyperone (82) were identified. Significantly, compound 82 displayed moderate antibacterial activity against S. aureus with an value of 11.4 \u03bcg/mL [Three novel ent-eudesmane sesquiterpenoids, namely arundinols A (.4 \u03bcg/mL .R)-1\u2032--1\u2032-oxobutan-3\u2032-ylacetate (4), was isolated, alongside six known compounds, (R)-1--3-hydroxybut -anone (5), 1--2-buten-1-one (6), (R)-6-hydroxy-2-methyl-4-chrom -anone (7), modiolide D (84), modiolide E (85), and modiolide A (86). These compounds were obtained from the endophyte fungus M. arundinis, residing within the plant Paepalanthus planifolius [5 exhibited moderate cytotoxic activity against murine breast adenocarcinoma (LM3), with IC50 values of 36.83 \u00b1 4.86 \u03bcg/mL, while compound 6 displayed moderate cytotoxic activity against human breast cancer (MCF-7), with IC50 values of 33.95 \u00b1 3.62 \u03bcg/mL [A new metabolite, , through alterations of the culture medium for the fungus M. olivacea F010 [98 demonstrated significant cytotoxic activity against murine leukemic cells (L1210), achieving an inhibition ratio of 90% [Keusgen et al. (1996) successfully isolated a cerebroside, namely cea F010 . Remarkao of 90% .99), C (101), and E (103), in conjunction with two known compounds, TAN-1496 B (100) and D (102), were successfully isolated from the fungus Microsphaeropsis sp. FL-16144. Extensive research has revealed that these compounds act as specific inhibitors of calf thymus Topo I [Three novel metabolites, TAN-1496 A (s Topo I . Notablys Topo I . Additios Topo I .87) and macrosphelide B (88), from the fungus Microsphaeropsis sp. FO-5050 [50 values of 3.5 \u03bcM and 36 \u03bcM, respectively. Notably, in vitro cell growth assays showed that these compounds had no discernible effect on the growth of cells, including P388 leukemia, human prostate tumor cells, and L929 fibroblast cells, even when administered at a high dose of 200 mg/kg for 5 days [89) and macrosphelide D (90) from the fungus Microsphaeropsis sp. FO-5050 [89) and macrosphelide D (90) exhibited moderate cytotoxic activity, with IC50 values of 67.5 \u03bcM and 25 \u03bcM, respectively. Fukami et al. (1998) continued their exploration of the fungus Microsphaeropsis sp. FO-5050 and made further discoveries [91), macrosphelide K (92), and 6-epi-5\u2019-hydroxymycosporulone (12). Although these compounds displayed cytotoxic activity, their IC50 values exceeded 100 \u03bcg/mL. Collectively, these findings underscore the exceptional potential of the fungus Microsphaeropsis sp. FO-5050 as a source of novel macrolides with cytotoxic properties. These substances hold high promise for future development in the treatment of leukemia.It has been widely reported that cell adhesion molecules play pivotal roles in numerous physiological and biochemical processes, ranging from cancer ,42,43 to FO-5050 . To explr 5 days . Takamat FO-5050 . Macrospcoveries . They is18, 19, and 20 possess the capability to inhibit Ras farnesyl-protein transferase [16\u201321 have demonstrated significant cytotoxicity against a spectrum of cell lines, including human epidermoid carcinoma (KB cells), human breast cancer (BC-1 cells), and African green monkey kidney fibroblast (vero cells). Compound 28, sourced from the endophyte fungus M. arundinis E12-2112, has exhibited moderate cytotoxicity against human bladder carcinoma cells (T24) and human lung carcinoma cells (A549), characterized by IC50 values of 35.4 \u03bcg/mL and 81.6 \u03bcg/mL, respectively [Singh et al. (1994) previously reported that compounds nsferase . Seephonnsferase . Furtherectively .49), graphislactone A (50), ulocladol (51), botrallin (52), 2,5-diacetylphenol (53), 7-hydroxy-2,4-dimethyl-3(2H)- benzofuranone (54), and enalin (55), were isolated from the endophyte fungus M. olivacea, which resides within the plant Pilgerodendron uviferum. Remarkably, graphislactone A (50) and botrallin (52) exhibited significant activity against acetylcholinesterase (AChE), characterized by IC50 values of 8.1 \u03bcg/mL and 6.1 \u03bcg/mL, respectively. However, all these compounds displayed no discernible antibacterial or antifungal activities [Seven known compounds, namely butyrolactone I (tivities .76), from the fungi Microsphaeropsis sp. FO-5050 [76 holds great promise as a novel class of anti-influenza virus drugs, representing a potential breakthrough in the realm of influenza treatment.Fukami et al. (2000) isolated a novel anti-influenza virus antiviral, known as 10-norparvulenone ( FO-5050 . Interes95), was isolated from the endophyte fungus Microsphaeropsis sp. MF6057, found within the plant Prosopis glandulosa. Compound 95 demonstrated an IC50 value of 71 \u03bcM in binding to a cloned human B2 receptor with 3H-bradykinin, marking a noteworthy development in the pursuit of potential treatments [Bradykinin, an endogenous peptide, exerts its effects through specific cell receptors, giving rise to a plethora of physiological reactions, including pain, allergic responses, and muscle contractions. Bradykinin antagonists hold the potential to inhibit these physiological reactions, offering prospects for treating conditions such as inflammatory edema, rhinitis, and asthma. In a significant discovery, a novel non-peptide bradykinin-binding inhibitor, L-755,807  and microsphaerones B (97), were successfully isolated from the fungus Microsphaeropsis sp. KMPB W-22. Interestingly, neither of these compounds exhibited significant cytotoxicity. Additionally, they displayed either no activity or only moderate activity against S. littoralis and A. salina [Two novel . salina .83 and 104) and three fresh 1,3,6,8-tetrahydroxyanthraquinone congeners (13\u201315) were successfully extracted from the fungus Microsphaeropsis sp. KMPB W-22. Remarkably, all these compounds, with the exception of compound 104, exhibited moderate inhibitory activity against protein kinases such as PKC, CDK4, and EGF-R, encompassing an IC50 range of 18.5\u201354.0 \u03bcM [Two novel betaenone derivatives  first reported on the in vitro activity of preussomerins against lciparum . Compoun44, sourced from the endophyte fungus M. arundinis PSU-G18, has exhibited significant antiplasmodial activity, boasting an IC50 value of 9.63 \u03bcg/mL. Notably, it also demonstrates significant radical scavenging capabilities, with an IC50 value of 0.018 mg/mL [1 (56), decaspirone (57), and papyracillic acids A (59), all originating from the endophyte fungus Microsphaeropsis sp. 7291, have demonstrated anti-algal activity against C. fusca. Their inhibitory effects are reflected in inhibition zone diameters of 6 mm, 13 mm, and 7 mm, respectively [112, derived from the endophyte fungus Microsphaeropsis sp. 7820, has displayed anti-algal activity against C. fusca, characterized by an inhibition zone diameter of 8.5 mm [64\u201373, apart from 8-hydroxy-6-methyl-9-oxo-9H-xanthene-1-carboxylic acid methyl ester (71), have demonstrated anti-algal activity against C. fusca [74) and microketides B (75), isolated from Microsphaeropsis sp. RA10-14, have exhibited moderate antiphytoplankton activity against P. helgolandica, showcasing IC50 values of 12.3 \u03bcg/mL and 16.8 \u03bcg/mL, respectively [Compound 18 mg/mL . In anotf 8.5 mm . Additioectively .Microsphaeropsis genus, and their activities have yet to be determined. For instance, Liu et al. (2018) isolated six known metabolites, including butyrolactones I (24), butyrolactones IV (25), aspernolide D (26), fumiquinazolines L (105), fumiquinazolines N (106), and notoamide D (107), from the fungus Microsphaeropsis sp. CF09-11, sourced from marine sediment [24\u201326 and 105\u2013107) were obtained from Microsphaeropsis for the first time. In a unique discovery, uncommon methyl-branched unsaturated fatty acids, 10-methyl-9Z-octadecenoic acid (108) and glyceride (109), were isolated from the endophyte fungus M. olivacea, which inhabits the marine sponge Agelus sp. This marks the first report of fungal fatty acids with methyl branches on a cis-double bond [63), papyracillic acids A (59), and papyracillic acids B (60), were isolated from the endophyte fungus Microsphaeropsis sp., originating from the plant Arbutus unedo. Importantly, this research led to the establishment of the absolute configuration of massarigenin A (63) [Furthermore, several compounds were isolated from fungi within the sediment . It is nble bond . Addition A (63) .Microsphaeropsis in 1980, fifty-four different species have been included. Currently, this genus has yielded an impressive array of secondary metabolites, with 112 secondary metabolites reported, of which 49 (41.96%) were identified for the first time. These compounds are distributed across various structural classes, including 76 polyketones (67.86%), 14 nitrogenous compounds (12.5%), 10 macrolides (8.9%), 7 terpenoids (6.25%), and 5 other compounds (4.46%), indicating polyketones are the predominant structural type  Employ non-directional activation strategies to activate the biosynthesis of secondary metabolites, such as the OSMAC (one strain many compounds) strategy [Microsphaeropsis has not been reported in the literature so far. Thus, whole gene sequencing and analysis will be the focus of further work. (iii) Expand the scope of activity screening of existing compounds and delve into their mechanism of action to verify their potential for pharmaceutical applications. For instance, microbetides A (74), a novel polyketone compound, demonstrates remarkable antibacterial activity against Gram-positive bacteria and Gram-negative bacteria, with an IC50 of 0.19 \u03bcg/mL, comparable to ciprofloxacin. Polyketone 10-norparvulone (76) exhibits the potential to effectively inhibit virus replications, positioning it as a promising candidate for a new anti-influenza drug. Macrolide compounds like macrosphelide A (87) and macrosphelide B (88), characterized by their unique chemical structures and cytotoxicity against leukemia, hold promise as novel anticancer drugs.Fungi occupy an exceptionally pivotal role in drug discovery . Many restrategy , co-cultstrategy , and epistrategy . (ii) Hastrategy , transcrstrategy , the hetstrategy , DNA-assstrategy , and ribstrategy . However"}
+{"text": "Escherichia coli (ECOL) and Enterobacter cloacae complex (ENTCC) demonstrate a range of MEM MICs. The objective of this study was to assess the in vitro killing activity of dose-optimized MEM against CRE with or without KPC that demonstrate variable MEM MICs and to determine if vaborbactam potentiates the killing activity of MEM against non-CP CRE.KPC producing CRE are common in the US; however non-carbapenemase producing (CP) CRE are as common. KPC- and non-CP Table); isolates demonstrated MEM MICs ranging from 1 to 8 mg/L. MEM standard (1g q8h over 30 minutes) and optimized (2g q8h over 3 hours) regimens were tested in 1-compartment and hollow fiber infection models (HFIM); targeted exposures were confirmed by LC-MS/MS. In HFIM, MEV (4g q 8h over 3 hours) was also tested. Resistant subpopulations were selected by agar plates containing MEM at 4x baseline MIC.8 ECOL and 8 ENTCC were characterized by whole-genome sequencing  or ENTCC . Bacterial regrowth occurred more rapidly against KPC compared to non-CP isolates. For MEM optimized regimens, 72-hour log-kills decreased as MEM MICs increased against non-CP ECOL; optimized MEM regimens were not bactericidal against KPC ECOL with MICs \u22652 mg/L. For ENTCC, optimized MEM resulted in greater log-kills against non-CP than KPC isolates at each MIC level. Across all non-CP and KPC isolates with MEM MICs \u22642 mg/L (n=4 each), mean log-kills at 72 hours with optimized MEM were -4.43 and -0.26 logs, respectively (P=0.04). Colonies selected on MEM containing agar demonstrated a median (range) MEM MIC of 64 mg/L (16 \u2013 >256 mg/L). Next, we compared optimized MEM and MEV in 10 day HFIMs against isolates with MEM MICs of 4mg/L. MEV demonstrated potent bactericidal killing against KPC isolates. Unexpectedly, MEV was also bactericidal against both non-CP isolates, and potentiated MEM killing against ECOL0174 .Standard MEM regimens were largely not bactericidal against ECOL . Optimized dosing MEM 2g q8h, 3 hour infusion (green). Limit of detection is 0.7 log colony forming units (CFU)/mL, denoted by dashed line. BMD = broth microdilution. KPC = Klebsiella pneumoniae carbapenemase.Standard dosing MEM 1g q8h, 30 minute infusion (red). Optimized dosing MEM 2g q8h, 3 hour infusion (green). Lower limit of detection is 0.7 log CFU/mL, denoted by dashed line. BMD = broth microdilution. KPC = Klebsiella pneumoniae carbapenemaseStandard dosing meropenem 1g q8h, 30 minute infusion (red). Optimized dosing meropenem 2g q8h, 3 hour infusion (green), or meropenem-vaborbactam 4g q8h, 3 hour infusion (purple). Lower limit of detection is 0.7 log CFU/mL. MIC = minimum inhibitory concentration.in vitro killing of CR ECOL and ENTCC is dependent on MEM MICs and presence of KPC. Even at low MICs, optimized MEM exposures were ineffective against KPC isolates. Against non-CP CRE, killing decreases as a function of increased MEM MIC. MEV may play an additive role in killing against non-CP isolates that are not killed by MEM alone.Our data show Ryan K. Shields, PharmD, MS, Allergan: Advisor/Consultant|Cidara: Advisor/Consultant|Entasis: Advisor/Consultant|GSK: Advisor/Consultant|Melinta: Advisor/Consultant|Melinta: Grant/Research Support|Menarini: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Pfizer: Advisor/Consultant|Roche: Grant/Research Support|Shionogi: Advisor/Consultant|Shionogi: Grant/Research Support|Utility: Advisor/Consultant|Venatorx: Advisor/Consultant|Venatorx: Grant/Research Support"}
+{"text": "In the original publication , there wThere was an error in the original publication. We examined correlations among biomarkers, which should have been analyzed by Spearman\u2019s correlation method instead of Pearson\u2019s correlation.A correction has been made to Section 2, \u201cResults\u201d; Section 2.4, \u201cCorrelations between Major CSF Proteins in CSF of CN, MCI, and AD\u201d; and Section 2.5, \u201cCorrelations between Major CSF Proteins and AD Biomarkers\u201d.rs = 0.86) and L-PGDS (rs = 0.47) levels (Table 1), while GlcNAc-Tf also correlated with L-PGDS (rs = 0.57). In contrast, Man-Tf levels did not correlate with Sia-Tf or TTR levels. High correlations were thus found among Man-Tf, GlcNAc-Tf, and L-PGDS in the CN group. In MCI, Man-Tf levels correlated well with GlcNAc-Tf (rs = 0.73) and L-PGDS (rs = 0.66) levels, while GlcNAc-Tf correlated well with L-PGDS (rs = 0.69). Man-Tf levels were negatively, but weakly, correlated with TTR (rs = \u22120.36) levels but not with Sia-Tf. Again, high correlations were observed among brain-derived proteins in the MCI group. In AD, Man-Tf levels correlated well with GlcNAc-Tf (rs = 0.79) and L-PGDS (rs = 0.62) levels but not with Sia-Tf or TTR. On the other hand, GlcNAc-Tf levels correlated well with L-PGDS (rs = 0.61) levels. These results suggest that the brain-derived proteins showed significant correlations with each other across all clinical groups.Correlations of the CSF proteins were also analyzed for the CN, MCI, and AD clinical groups. In CN, Man-Tf levels correlated with GlcNAc-Tf (rs = 0.56), tau (rs = 0.56), A\u03b240 (rs = 0.73), and A\u03b242 (rs = 0.52); GlcNAc-Tf vs. p-tau (rs = 0.76), tau (rs = 0.79), A\u03b240 (rs = 0.76), and A\u03b242 (rs = 0.49); L-PGDS vs. p-tau (rs = 0.84), tau (rs = 0.79), A\u03b240 (rs = 0.79), and A\u03b242 (rs = 0.52). The A\u03b242/A\u03b240 ratio showed negative correlations with Man-Tf (rs = \u22120.15), GlcNAc-Tf (rs = \u22120.34), and L-PGDS (rs = \u22120.44); however, none of these were statistically significant. In contrast, neither Sia-Tf nor TTR levels correlated with any AD markers. Taken together, the brain-derived CSF proteins correlated significantly with p-tau, tau, A\u03b240, and A\u03b242 levels in the CN group but the blood-derived proteins did not. In MCI, CSF protein levels also correlated with AD markers as follows: Man-Tf vs. p-tau (rs = 0.71), tau (rs = 0.53), A\u03b240 (rs = 0.45); GlcNAc-Tf vs. p-tau (rs = 0.65), tau (rs = 0.54), and A\u03b240 (rs = 0.43); L-PGDS vs. p-tau (rs = 0.49) and tau (rs = 0.41). The A\u03b242/A\u03b240 ratio showed negative correlations with Man-Tf (rs = \u22120.24), GlcNAc-Tf (rs = \u22120.45), and L-PGDS (rs = \u22120.23) but none were statistically significant. Overall, in the MCI group, the brain-derived proteins correlated moderately with AD markers. In AD, p-tau showed a moderate, though significant, correlation with the CSF proteins (rs = 0.51~0.55) but not with other AD markers. TTR negatively correlated with p-tau (rs = \u22120.42) and tau (rs = \u22120.72), while Sia-Tf correlated with tau (rs = \u22120.41). These results suggest that AD markers tend to correlate with the brain-derived CSF proteins in the AD and MCI groups; however, with the exception of p-tau, their correlation declined concomitantly with AD progression.Our previous study demonstrated that Man-Tf and p-tau were co-expressed in hippocampal neurons in the AD brain and that CSF Man-Tf levels correlated well with p-tau levels in MCI and AD. Other CSF proteins, such as Man-Tf, GlcNAc-Tf, L-PGDS, TTR, and Sia-Tf, were therefore examined for their correlation with AD markers (Table 2). In CN subjects, the CSF proteins showed significant correlations with AD markers as follows: Man-Tf vs. p-tau (The authors state that the scientific conclusions are unaffected. This correction was approved by the Academic Editor. The original publication has also been updated."}
+{"text": "Serum UA , NT-proBNP levels (1090.4 (675.2\u20132664.9) vs. 759.0 (260.3\u20131474.8), p = 0.034), and MLHFQ scores (21.0 (14.0\u201347.0) vs. 14.5 (4.5\u201325.5), p = 0.012) were significantly higher, whereas 25(OH)D concentrations (17.6 (15.1\u201328.2) vs. 22.7 (19.5\u201333.8), p = 0.010) were lower in subjects with severely reduced LVEF. Also, 25(OH)D was independently associated with LVEF in univariate and multiple regression analysis, maintaining its significance even after adjusting for confounders such as age, NT-proBNP, the presence of chronic coronary syndrome, hypertension, and anemia. According to our current findings, 25(OH)D is closely associated with LVEF, further supporting the need to establish correct vitamin D supplementation schemes and dietary interventions in HF. The changes in LVEF, 25(OH)D, serum UA, and albumin levels in HFrEF and HFmrEF indicate a similar pathophysiological background.Vitamin D emerged as an important prognostic biomarker in heart failure (HF), with currently highly debated therapeutic implications. Several trials on vitamin D supplementation in HF showed improvements in left ventricular (LV) remodeling and function and health-related quality of life (HRQoL), which did not translate into mid- to long-term beneficial effects regarding physical performance and mortality. We addressed total 25-hydroxyvitamin D (25(OH)D), serum albumin, and uric acid (UA) levels, focusing mainly on vitamin D deficiency, as potential markers of LV systolic dysfunction in HF with reduced and mildly reduced ejection fraction . Seventy patients with LVEF < 50% were comprehensively evaluated using ECG, echocardiography, lung ultrasound (LUS), blood sampling, and the six-minute walk test (6MWT). HRQoL was also assessed using the Minnesota Living with Heart Failure Questionnaire (MLHFQ). Statistically significant positive correlations were found between LVEF, 25(OH)D, serum UA, and albumin, respectively ( Vitamin D is a pleiotropic hormone showing substantial cardiovascular involvement beyond its regulatory effects on calcium and phosphate homeostasis. Vitamin D deficiency is frequently associated with cardiovascular disease  and is i3 2D3), the biologically active form of vitamin D, exerts its genomic and rapid, non-genomic effects via vitamin D receptors (VDRs) . R. R2 usinEchocardiography was performed using a Philips Epiq7 ultrasound machine  and a Philips X5-1 xMATRIX array transducer . Clinical recommendations for multimodality cardiovascular imaging were applied ,28,29. LThe lung ultrasound (LUS) protocol consisted of an eight-zone evaluation, four on each hemithorax. Semi-recumbent positioning of patients facilitated the assessment of B-lines. The same cardiac probe was used for lung imaging as well, and depth was adjusted individually. The following threshold for abnormality (in terms of lung congestion) was applied: \u22653 B-lines per zone .Blood samples were collected after twelve-hour fasting , and centrifuged for 10 min at 3000 rpm. Laboratory tests were performed, including total-, LDL-, and HDL-cholesterol, serum triglycerides, UA, creatinine, albumin, serum iron, ferritin, and CRP using an Architect C4000 ; plasma fibrinogen was assessed with Sysmex CA-1500 . N-terminal pro-B-type natriuretic peptide (NT-proBNP) concentrations were measured using an electrochemiluminescent method on an Elecsys 2010 . Complete blood count was assessed using tripotassium ethylenediaminetetraacetic acid precoated tubes on a Mindray BC6000 .Quantitative determination ofthe total 25(OH)D in patients with HF was conducted using a competitive electrochemiluminescence protein binding assay on a Mindray CL-900i Chemiluminescence Immunoassay Analyzer . Patients were divided into tertiles based on serum 25(OH)D levels . None of the participants were taking vitamin D supplements in the two months prior to the evaluation. Dietary vitamin D intake from food sources and sun exposure were not assessed. The reference range for this assay was 30\u2013100 ng/mL.2 test. 25(OH)D concentrations were log-transformed for analysis to improve normal distribution. p-values < 0.05 were regarded as statistically significant. Microsoft Excel 2016  and GraphPad Prism 9.5.0  were used for data curation and processing.The Kolmogorov\u2013Smirnov and/or Shapiro\u2013Wilk normality tests were used for the assessment of data distribution. Paired t-test and Pearson\u2019s correlation were performed for variables showing normal distribution. Data following non-Gaussian distribution were processed using the Mann\u2013Whitney U test and Spearman\u2019s rank correlation analysis. Categorical variables were quantified for absolute and relative frequency, and 2 \u00d7 2 contingency tables were analyzed with the \u03c7p = 0.019). Patients with HFrEF had higher body mass index (BMI) (p = 0.025), LA volume index (LAVI) (p = 0.032), LVEDV index (LVEDVI) (p < 0.0001), LV end-systolic volume index (LVESVI) (p < 0.0001), lower stroke volume index (SVI) (p = 0.0002), longer-lasting heart disease (p = 0.041), compared to patients with HFmrEF. Also, study participants with HFmrEF showed better quality of life (p = 0.012). Serum UAd and NT-proBNP concentrations were significantly increased , whereas 25(OH)D levels were significantly decreased (p = 0.010) in HFrEF. Loop diuretic therapy was more frequently prescribed to patients with HFrEF. None of the participants took serum UA-lowering medication at the time of evaluation.Seventy patients  with a mean age of 66 \u00b1 11 years were included in the study. p < 0.01) (Both log 25(OH)D and serum UA showed significant differences and an inverse relationship in the two study groups ( < 0.01) .p = 0.023), LVEF , platelet count , serum albumin , and HDL-cholesterol concentrations . Statistically significant negative associations were reported between serum 25(OH)D and UA levels , LVESVI , and LVGLS . No significant relationship was found connecting 25(OH)D to CRP , 6MWT distance , or NYHA class . Patients with 25(OH)D levels in the first tertile vs. the third tertile associated type II diabetes mellitus (DM) more frequently, but it did not reach statistical significance .Also, when considering 25(OH)D levels in all patients, significant positive correlations were found with diastolic blood pressure values , BMI (p = 0.006), and E/e\u2019 ratio (p = 0.037). LVEF also showed strong negative associations with NT-proBNP (p = 0.029), serum UA levels (p = 0.009), GGT activity (p = 0.006), and thyroid-stimulating hormone (TSH) concentrations (p = 0.040). Interestingly, LVEF was not linked to 6MWT distance, age, duration of heart disease, or duration of HF. Significant positive correlations were found between LVEF and both components of blood pressure\u2014systolic (p = 0.003) and diastolic blood pressure (p = 0.006), and also serum 25(OH)D levels (p = 0.008).p = 0.036 and p = 0.030, respectively). In the second model, log 25(OH)D maintained its predictive value, even after adjusting for the presence of chronic coronary syndrome (CCS).Multivariate linear regression models (A and B) were constructed in the overall study population for the prediction of LVEF in HF . Log 25(p = 0.027) (Model C). Furthermore, log 25(OH)D predicted HFrEF, maintaining its significance when corrected for age, NT-proBNP tertiles, GGT tertiles, the presence of hypertension, CCS, anemia, and hypothyroidism (p = 0.036) (Model D).Nonlinear logistic regression models (C and D) were also set up to predict LV systolic dysfunction . In the 2D is released into the bloodstream, circulating in both protein-bound  and free forms [2D deficient mice developed glomerular (podocyte) injury, albuminuria, and subsequent kidney dysfunction, which were reversible upon exposure to 1,25(OH)2D [Over the last few decades, especially during the COVID-19 pandemic, vitamin D received widespread attention, mainly because of its complex biological activity, regulating gene transcription, and modulating signaling pathways . Vitaminee forms . The remee forms . Moreove25(OH)2D . Zhang e25(OH)2D . Our pat25(OH)2D ,37, vita25(OH)2D .+ currents proved to be an important cause of electrophysiological remodeling in cardiac hypertrophy, which was reversed upon exposure to 10 nM of calcitriol [2D. Calcitriol, at concentrations above 30 nM, reduced the proliferative capacity, caused mild hypertrophy, and altered cardiomyocyte morphology [p = 0.010). Currently, there is no guideline recommendation regarding screening for vitamin D deficiency and correct supplementation in HF. Both experimental and clinical trials linked vitamin D deficiency to the excessive activation of the RAAS with increased expression of the renin gene, higher plasma renin activity, and elevated plasma aldosterone and angiotensin II levels [2D increases intracellular calcium concentrations and decreases intracardiac filling pressures, thus enhancing systolic and diastolic ventricular function [p = 0.036 and p = 0.030; p = 0.027 and p = 0.031). Contrary to the results published by Khan et al., we found no significant relationship between 25(OH)D levels and the functional status of patients with HF, reflected by the 6MWT distance and NYHA class [p = 0.061). Vitamin D deficiency was previously linked to DM by regulating inflammation and inducing insulin resistance [A multiple inhibitory role regarding vitamin D in HF has been proposed: it reduces myocardial fibrosis, pathological remodeling, and atherosclerosis, by down-regulating low-grade vascular inflammation . The suplcitriol . Vitaminlcitriol . VDRs werphology . The sizrphology . VDR knorphology . There irphology . Other arphology . Our patI levels . Vitaminsistance . Cassanosistance .3 administered weekly and 400 mg of calcium citrate given BID for 6 months, failed to demonstrate the beneficial effects of cholecalciferol supplementation on physical performance in HF patients with a mean LVEF of 39.2 \u00b1 13.2%, and already on guideline-directed medical therapy [3 daily during a 3-year follow-up. Long-term intake of moderately high doses of vitamin D had no impact on mortality but was associated with a significantly greater need for mechanical circulatory support device implants in patients with baseline circulating 25(OH)D levels \u2265 30 nmol/L, who showed increased in-study concentrations > 100 nmol/L [Several trials addressed the potential benefits and pitfalls of vitamin D supplementation in HF but showed inconsistent results. The VINDICATE study enrolled vitamin D-deficient patients with HF and LVEF \u2264 45% already on optimal medical treatment. After 12 months of well-tolerated high-dose cholecalciferol supplementation (4000 IU daily), LV structure and function showed statistically significant improvements, reflected by the EF, end-diastolic, and end-systolic diameters, and volumes. Unexpectedly, the reversal of LV remodeling was not associated with improved 6MWT distance at 1 year, and further clinical outcomes were not assessed . Another therapy . The mor0 nmol/L . In a ca0 nmol/L . High pl0 nmol/L . A recen0 nmol/L ,54. Vita0 nmol/L .As seen, the trials investigating the effects of vitamin D in HF used different supplementation schemes  and study outcomes, which might explain the inconclusive results.p = 0.005). We also found that patients with 25(OH)D deficiency were prone to develop hyperuricemia. The association might be explained by the severity of LV systolic dysfunction, which is associated with lower levels of vitamin D [Serum UA also sparked controversy regarding its status as a potential cardiovascular risk factor, considering its pro-oxidant\u2013antioxidant activity. Elevated levels were linked to increased risk for cardiovascular disease, including HF . Moreoveitamin D ; both viitamin D ,24. Alsoitamin D . Treatmeitamin D ,64. The itamin D . In patiitamin D . The URRitamin D . It shouWe hypothesize that the associated changes in LVEF, 25(OH)D, serum UA, and albumin levels might indicate similar pathophysiological backgrounds in HF with systolic dysfunction. One possible explanation might rely on the pleiotropic biological impact of inflammation. Abundant evidence already linked low-grade systemic inflammation to HF . TherefoVitamin D, serum UA, and albumin are already established prognostic biomarkers in HF. We found 25(OH)D to be an independent marker of LVEF using univariate and multiple regression analysis. Significant correlations were identified between LVEF, 25(OH)D, serum UA, and albumin, suggesting potential molecular interactions. Although there is currently a plethora of information regarding vitamin D deficiency in HF, its precise mechanism, clinical impact, and management still need clarification."}
+{"text": "Electroconvulsive therapy (ECT) is one of the few non-pharmacological stimulation treatment which is cost effective, effecious and lifesaving in various psychiatric disorders. Although myths and misconceptions prevailed in a society undermine the usefulness of such treatment.Attitude towards Electroconvulsive therapy (ECT) among psychiatric patients.It was a descriptive cross-sectional study conducted at the Department of Psychiatry and Behavioural sciences, Jinnah Postgraduate Medical Centre (JPMC), Karachi from 22-Oct-2019 to 21-April-2020 and a total of 250 psychiatric patients were enrolled.Methode; Attitudes toward ECT were assessed using ECT attitude questionnaire6 (Annexure III). A 15 items questionnaire, each item has three alternatives based on which responses were categorized into positive, negative, or ambivalent attitudes. Mean score was calculated for each.Patients who were given 8 positive answers out of 15 were labeled as having a positive attitude. Patients who were given 8 negative answers out of 15 were labeled as having a negative attitude. Patients who were given 8 I don\u2019t know answers out of 15 were labeled as having ambivalent attitude.Age 18-65 yearsEither genderPsychiatric patients, having awareness regarding their nature of illness and could give consent for study.Patients with duration of illness >3 months.Psychiatry patients who have no awareness regarding their illness.Patients with impaired cognitiveForty-four (45.83%) patients had positive attitude, 36 (37.50%) had negative attitude and 16 (16.67%) had ambient attitude.Further stratification was also performed on the basis of educational status, occupational status, duration of illness, psychiatric diagnosis, and previous experience of ECT. There was no significant association was found of these variables with attitude regarding ECT.Mean age was 39.58\u00b112.48 years included in this study. There were 55 (57.29%) female and 41 (41.71%) male patients. There were 72 (75.00%) patients were household workers, 04 (4.17%) students, 06 (5.25%) unskilled labour, 3 (3.13%) skilled labour, 10 (10.42%) professionals and just 01 (1.04%) were law enforcement worker. 19 (19.79%) patients were diagnosed with schizophrenia, 62 (64.58%) were diagnosed with unipolar depression and 15 (15.63%) were diagnosed with bipolar disorder. Source of ECT information was 11 (11.46%) electronic media, 09 (9.38%) print media, 19 (19.79%) social media and 57 (59.38%) was from health professionals. Forty-four (45.83%) patients had positive attitude, 36 (37.50%) had negative attitude and 16 (16.67%) had ambient attitude.Knowledge regarding electroconvulsive therapy (ECT) was low in psychiatric patients in Pakistan. Only 45.83% patients showed positive attitude towards ECT.Carney S, Geddes J. Electroconvulsive therapy. Br Med J. 2003;326:1343-4Gangadhar BN., Kapur RL., Kalyanasundaram S. Comparison of electroconvulsive therapy with imipramine in endogenous depression: a double blind study. Br J Psychiatry.1982;141:367\u201371.Kellner CH., Fink M., Knapp R., Petrides G., Husain M., Rummans T., et al. Relief of expressed suicidal intent by ECT: a consortium for research in ECT study. Am J Psychiatry. 2005;162:977\u201382.Baghai T C, Moller HJ. Electroconvulsive therapy and its different indications. Dialogues Clin Neurosci. 2008 Mar; 10(1): 105\u201317.Weiner RD., Coffey CE., Folk J., Fochtmann LJ., Greenberg RM., et al. American psychiatric association committee on electroconvulsive therapy, The practice of electroconvulsive therapy. 2nd ed. Washington, DC: American Psychiatric Association; 2001None Declared"}
+{"text": "Although the survival rate of patients who undergo surgery for gastric cancer has greatly improved, still many patients have a poor prognosis. This retrospective study aimed to investigate the predictive ability of the PNI-IgM score, a combined prognostic nutritional index (PNI), and immunoglobulin M (IgM), on the prognosis of patients undergoing surgery for gastric cancer.340 patients with gastric cancer who underwent surgery from January 2016 to December 2017 were selected. The PNI-IgM score ranged from 1 to 3: score of 1, low PNI (< 48.45) and low IgM (< 0.87); score of 2, low PNI and high IgM, or high PNI and low IgM; score of 3, high PNI and high IgM. We compared the differences in disease-free survival (DFS) and overall survival (OS) among the three groups, while univariate and multivariate analyses calculated prognostic factors for DFS and OS. In addition, the nomograms were constructed based on the results of multivariate analysis to estimate the 1-, 3- and 5-year survival probability.P = 0.053) and the PNI-IgM score group 3 . In stratified analysis, PNI-IgM score 1 had a worse prognosis in the age < 60 years group and CA724 < 2.11 U/m group.There were 67 cases in the PNI-IgM score 1 group, 160 cases in the PNI-IgM score 2 group, and 113 cases in the PNI-IgM score 3 group. The median survival times of DFS in the PNI-IgM score group 1, the PNI-IgM score group 2, and the PNI-IgM score group 3 were 62.20 months, not reached, and not reached, and 67.57 months vs. not reached vs. not reached in three groups for OS. Patients in the PNI-IgM score group 1 had a lower DFS than the PNI-IgM score group 2 (HR = 0.648, 95% CI: 0.418-1.006, PNI-IgM score is a novel combination of nutritional and immunological markers that can be used as a sensitive biological marker for patients with gastric cancer who undergo surgery. The lower the PNI-IgM score, the worse the prognosis. According to the World Health Organization, gastric cancer was the fifth leading cancer in the world, with nearly 1.09 million new cases of gastric cancer worldwide in 2020. Gastric cancer is the fourth leading cause of cancer deaths, with almost 770,000 deaths worldwide in 2020 \u20135. In reThe nutritional status of patients with gastric cancer, which could predict the progression of the treated cancer, has been identified an important factor \u201311. It wA large number of previous researches have pointed out that composite indicators of nutrition and immunity, such as Glasgow Prognostic Score (GPS), platelet-to-lymphocyte (PLR), and Controlling Nutritional Status (CONUT) score could predict the prognosis of gastric cancer patients , 27. HowIn this study, we evaluated the predictive effect of the PNI-IgM score on efficacy and prognosis in 340 patients with gastric cancer who underwent surgery. To further validate the PNI-IgM score, we performed a subgroup analysis and created nomograms.This is a retrospective study, so the Ethics Committee of Harbin Medical University Cancer Hospital waived informed consent. In total, we collected 340 consecutive patients with gastric cancer who received surgery at Harbin Medical University Cancer Hospital between January 2016 and December 2017 and were tested for lymphatic subsets and specific proteins. Statistical analysis of 340 patients and their clinical information was implemented according to the Helsinki Declaration and its amendments. All patients were included according to the following criteria: (1) all patients underwent surgical treatment; (2) all patients had no chronic disease; (3) all patients were tested for lymphatic subsets and specific proteins; (4) all patients did not display inflammatory response. (5) Patients with gastric cancer combined with other primary malignant tumors were excluded. Patients without complete clinical information and regular review after surgery were exclusion criteria. The flow chart of clinical case selection is shown in Patients were followed up by telephone or outpatient visit, every 3-6 months during the first 2 years, and every 6-12 months from the 3rd to 5th year, and annually thereafter. Disease-free survival (DFS) was comprehended as the period from the first day of surgery date to the date of disease progression. The evidence of progression was obtained by chest and abdomen X-ray or computed tomography. DFS was also defined as the date of death, death from any cause, or the date of withdrawal from the follow-up. Overall survival (OS) was described as the period from the first day of surgery date to the date of death, the date of withdrawal from the follow-up, or the time of the last follow-up. Electronic medical record system was used to acquire patients\u2019 clinical and pathological information.9/L). The cut-off point was obtained by the receiver operating characteristic (ROC), which was based on OS for the prediction of patients\u2019 death. The area under the ROC Curve (AUC) was used to evaluate the predictive ability of PNI and IgM. The optimal cut-off values of PNI and IgM with the highest Youden index were obtained.The peripheral venous blood was collected in fasting state after admission in all patients. The counts of peripheral lymphocytes (L) were measured and analyzed by an automatic blood analyzer (BACKMAN COULTER LH750), the levels of peripheral albumin were measured and analyzed by an automatic blood analyzer (ADVIA-2400), and the levels of peripheral IgM were measured and analyzed by a specific protein analyzer (IMMAGE800). PNI was calculated as follows: PNI = albumin (g/L) + 5 \u00d7 total lymphocyte counts  and 95% confidence interval (CI). The Cox proportional hazards regression model was constructed to analyze independent prognostic factors for DFS and OS. The nomograms were also constructed to predict the 1-, 3-, and 5-year survival probability for DFS and OS. The calibration curve analysis was used to assess the prognostic predictive ability of nomogram. All statistical analyses were completed through the R 4.1.3 , IBM SPSS Statistics 25 . Finally, we considered two-sided P values < 0.05 as statistical differences.Continuous variables are presented as means with standard deviations or as medians with interquartile ranges. Categorical variables were expressed as percentages. The comparison between continuous variables used the t-tests, one-way ANOVA, Kruskal-Wails rank sum test. We used the Chi-square test or Fisher\u2019s exact test to compare the discrepancies between categorical variables. The Kaplan-Meier survival curve was used to compute the survival rate and the Log-rank test to compare the survival time difference. Univariate and multivariate analyses were performed using the Cox proportional hazards model. Variables that achieved statistical significance at P = 0.006), weight loss (P = 0.012), fatigue (P < 0.001), pTNM stage (P < 0.001) and tumor size (P = 0.003). The one-way ANOVA Kruskal-Wails rank sum test showed that PNI-IgM score was related to age (P < 0.001), BMI (P = 0.012), bleeding volume (P = 0.012). The detailed clinical characteristics of all 340 cases grouped by PNI-IgM score are displayed in The median age of patients was 60 years, and there were 105 women (30.9%) and 235 men (69.1%) in all two groups\u2019 cases. The optimal cut-off value of PNI was 48.45. The optimal cut-off value of IgM was 0.87 g/L. According to the optimal cut-off values of PNI and IgM, all patients were divided into three groups: PNI-IgM score of 3 (n = 113): high IgM (\u2265 0.87) and high PNI (\u2265 48.45); PNI-IgM score of 2 (n = 160): high IgM (\u2265 0.87) and low PNI (< 48.45), or low IgM (< 0.87) and high PNI (\u2265 48.45); PNI-IgM score of 1(n = 67): low IgM (< 0.87) and low PNI (< 48.45). The Chi-square test or Fisher\u2019s exact test showed that PNI-IgM score was related to melaena (P = 0.017), \u03b3- GT (P < 0.001), TBIL (P < 0.001), TP (P < 0.001), ALB (P < 0.001), GLOB (P < 0.001), UA (P = 0.002), ALP (P = 0.027), WBC (P < 0.001), RBC (P < 0.001), IgA (P < 0.001), IgG (P < 0.001), KAP (P < 0.001), LAM (P < 0.001) and CD3 +/CD4 + CD8 + cells (P = 0.004).In this study, we also collected patients\u2019 nutritional and hematological parameters before surgery, including total protein (TP), PALB, alanine aminotransferase (ALT), aspartate transaminase (AST), glutamyl transpeptidase (\u03b3- GT), lactate dehydrogenase (LDH), total bilirubin (TBIL), globulin (GLOB), urea, creatinine (CREA), urate (UA), alkaline phosphatase (ALP), monocyte (Mono), eosinophils (Eosi), basophil (Baso), red blood cell (RBC), platelet (P), carcinoembryonic antigen (CEA), carbohydrate antigen 199 (CA199), carbohydrate antigen 724 (CA724), carbohydrate antigen 125II (CA125II). We also collected lymphatic subsets and specific proteins, including immunoglobulin A (IgA), immunoglobulin G (IgG), transferrin (TRF), light-chain immunoglobulin (KAP), heavy-chain immunoglobulin (LAM), KAP/LAM, CD3 + cells (T cells), CD3 +/CD4 + cells (Th cells), CD3 +/CD8 + cells (CTL cells), CD4 +/CD8+ cells, CD3 +/CD4 + CD8 + cells, CD19 + cells (B cells), CD3 -/CD16 + CD56 + cells (NK cells), CD3 +/CD16 + CD56 + cells (NKT cells). We analyzed their relationship to the PNI-IgM score by Kruskal-Wails rank sum test (P = 0.001), BMI (P = 0.043), tumor size (P < 0.001), pTNM stage (P < 0.001), radical resection (P < 0.001), TBIL (P = 0.020), Eosi (P = 0.032), RBC (P = 0.032), CA724 (P < 0.001), CA125II (P = 0.003), IgG (P = 0.028), PNI-IgM score (P < 0.05). And the prognosis factors of patients in this study for OS were age (P = 0.001), melaena (P = 0.048), tumor size (P < 0.001), pTNM stage (P < 0.001), radical resection (P < 0.001), TBIL (P = 0.016), Eosi (P = 0.030), RBC (P = 0.031), CA724 (P = 0.004), CA125II (P = 0.004), IgG (P = 0.036), PNI-IgM score (P < 0.05). The multivariate analysis indicated that age (P = 0.036 vs. P = 0.035), pTNM stage (P < 0.001 vs. P < 0.001), radical resection (P = 0.006 vs. P = 0.001), and CA724 (P = 0.027 vs. P = 0.015) were both independent prognostic factors for DFS and OS. In addition, melaena (P = 0.003) was the independent prognostic factor for OS  and in CA724 < 2.11U/m group   or the PNI-IgM score 3 . Patients with the PNI-IgM score 1 have a shorter OS than patients with the PNI-IgM score 2  or the PNI-IgM score 3  , 65.4% (95% CI: 54.7%-78.3%), 53.1% (95% CI: 41.9%-67.3%). The PNI-IgM score 1 group\u2019s median survival time for OS was 67.57 months, the 1-, 3-, and 5-year OS probability was 89.5% (95% CI: 82.4%-97.2%), 69.0% (95% CI: 58.6%-81.3%), 55.6% (95% CI: 44.5%-69.4%). The PNI-IgM score 2 group\u2019s median survival time for DFS and OS were both not achieved. The 1-, 3-, and 5-year DFS and OS probability were 89.3% (95% CI: 84.6%-94.2%), 74.0% (95% CI: 67.4%-81.3%), 68.2% (95% CI: 61.1%-76.1%); 89.9% (95% CI: 85.4%-94.7%), 77.8% (95% CI: 71.6%-84.6%), 69.7% (95% CI: 62.8%-77.3%), respectively. The PNI-IgM score 3 group\u2019s median survival time for DFS and OS were both not achieved. The 1-, 3-, and 5-year DFS and OS probability were 93.8% (95% CI: 89.4%-98.4%), 80.5% (95% CI: 73.4%-88.4%), 80.5% (95% CI: 73.4%-88.4%); 95.6% (95% CI: 91.9%-99.4%), 82.0% (95% CI: 75.1%-89.4%), 81.0% (95% CI: 74.0%-88.7%), respectively. To further determine whether the PNI-IgM score could predict the prognosis of gastric cancer patients. The ROC curves were based on OS for the prediction of patients\u2019 death. For the traditional clinicopathologic factors, including ALB, L, PNI, and IgM, each feature and the combined PNI-IgM score were plotted, and the point with the highest AUC was illustrated on the ROC curve. The PNI-IgM score exhibited a higher prognostic accuracy for DFS and OS than PNI and other clinicopathological risk factors (P = 0.626) or the PNI-IgM score 3 . Patients with the PNI-IgM score 1 also have a shorter OS than patients with the PNI-IgM score 2 group  or the PNI-IgM score 3 group . Patients in the advanced pTNM stage had lower DFS  and OS  than those early pTNM stage patients  group (221 patients) and advanced pTNM stage (III + IV) group (119 patients). The median survival time for DFS and OS in the early pTNM stage group were both not reached. The 1-, 3-, and 5-year DFS and OS probability were 98.2% (95% CI: 96.4%-100.0%) vs. 98.2% (95% CI: 96.4%-100.0%), 89.2% (95% CI: 85.2%-93.5%) vs. 89.9% (95% CI: 86.0%-94.0%), 86.7% (95% CI: 82.2%-91.4%) vs. 87.4% (95% CI: 83.1%-92.0%). The median survival time for DFS and OS in the advanced pTNM stage group were 30.90 months and 40.27 months. The 1-, 3-, and 5-year DFS and OS probability were 76.9% (95% CI: 69.6%-84.9%) vs. 79.6% (95% CI: 72.7%-87.2%); 45.7% (95% CI: 37.2%-56.2%) vs. 54.1% (95% CI: 45.7%-64.0%); 34.7% (95% CI: 26.6%-45.4%) vs. 39.0% (95% CI: 30.9%-49.3%). Patients with the PNI-IgM score 1 have a shorter DFS than patients with the PNI-IgM score 2 (HR = 0.814, 95% CI: 0.356-1.860, This study found that age, radical resection, CA724, and pTNM stage were the independent prognostic factors for DFS. By constructing Cox proportional hazard regression model, age, melaena, radical resection, CA724, and pTNM stage were the independent prognostic factors for OS. Based on the results of multivariate analysis, the nomograms to predict the 1-, 3-, and 5-year survival probability for DFS and OS were established (Gastric cancer is a common malignant tumor in China, with the third highest incidence and mortality rate . AlthougOur study is the first to assess the association between the PNI-IgM score, which a composite indicator of immunity and nutrition, clinicopathological factors and survival. It also demonstrated that the PNI-IgM score predicted prognosis among gastric cancer patients who underwent resection. We found that low PNI and IgM were associated with poor patient prognosis. This study found that age, pTNM stage, radical resection, and CA724 were independent prognostic factors for DFS and OS. In addition, an independent prognostic factor for OS was melaena. In the Stratified analyses, we found age <60 years and CA724 < 2.11 U/m had shorter OS in PNI-IgM score 1.Although many studies have confirmed the prognostic association of PNI and IgM with gastric cancer \u201335 and oThis study also has some limitations. First, this was a single-region, single-center retrospective study with limited sample size and potential selection bias. Second, we only selected patients with gastric cancer who underwent surgery. Third, the cut-off value of PNI and IgM is usually derived from the ROC curve, and the optimal cut-off value is uncertain. Therefore, multi-regional, multi-center, and larger sample size studies are needed to validate our findings.In conclusion, our study found that the PNI-IgM score is a valid scoring tool for patients with gastric cancer who received surgery. Patients in the PNI-IgM score 1 group had a worse prognosis than those in the PNI-IgM score 2 group, and the PNI-IgM score 3 group. Therefore, the PNI-IgM score could be used as a biomarker to develop better treatment strategies for patients undergoing surgery for gastric cancer.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.This study was approved by the ethics committee of Harbin Medical University Cancer Hospital. All patients provided written informed consent before the study.Writing-original draft and Writing-review & editing: ZD, HaS; Data curation and Investigation: RZ, GD, and HP; Methodology and Supervision: YZ, RH; Resources, Funding acquisition and Project administration: YX, and HoS. All authors contributed to the article and approved the submitted version."}
+{"text": "Kidney function can alter drug clearance. This is pertinent for tenofovir-based pre-exposure prophylaxis (PrEP) regimens where drug concentrations confer protection against HIV. Among transgender (TG) individuals whose kidney function is dynamic due to use of gender affirming therapy (GAT), it is unclear if there is a relationship between tenofovir diphosphate dried blood spots (TFV-DP DBS) and direct measures of glomerular filtration rate (mGFR) The objective of this study was to quantify this relationship and compare if mGFR differs between TG persons with perfect/imperfect adherence to tenofovir alafenamide/emtricitabine (TAF/FTC).A prospective cohort study was performed among TG individuals on TAF/FTC for PrEP. Inclusion criteria were age \u226518 years old, TG identity, HIV negative, use of PrEP for \u226512 weeks, and willing to receive iohexol for direct measurement of GFR. TFV DBS concentrations were evaluated after being on \u226512 weeks of TAF/FTC. Perfect adherence to TAF/FTC was defined as a TFV-DP DBS of >1800fmol/punch . Iohexol clearance was used for mGFR by subcutaneously administering 0.5ml iohexol. Plasma iohexol concentrations were assayed 3h post-injection. Pearson\u2019s correlation coefficients were computed to quantify the relationship between TFV-DP DBS and iohexol clearance. Median iohexol clearances were compared between those with perfect/imperfect adherence.2=0.10, 95% confidence interval, CI: -0.29 \u2013 0.45, p=0.62). TFV-DP DBS >1800fmol/punch (perfect adherence) was observed in 22 participants. Median (95% CI) iohexol clearance between those with and without perfect adherence did not reach significance: 78 (68-96) vs 69 (60-83)mL/min, respectively (p=0.26).Among the 28 individuals included, median  age was 33.0 (26.3 \u2013 36.8) years. Three quarters were on GAT and over two thirds (67.9%) were assigned male at birth (AMAB). Median  iohexol clearance and TFV-DP DBS concentrations were 76 (65 \u2013 95)ml/min and 2611 (2114 \u2013 3240), respectively. There was no significant correlation between TFV-DP DBS and iohexol clearance (rNo significant relationship was observed between iohexol clearance and TFV-DP DBS among TG individuals taking TAF/FTC for PrEP.Sheldon Morris, MD, Gilead: Grant/Research Support"}
+{"text": "Skin and soft tissue infections (SSTIs) are very common bacterial infections. Numerous clinical trials have compared antibiotics for SSTI treatment. However, trials typically have non-inferiority designs that give little insight as to which antibiotic classes have superior efficacy.We performed a systematic literature review and a network meta-analysis of published SSTI treatment trials. Using standardized key words, we searched PubMed and Embase for SSTI clinical trials published from 1/1/66 to 5/31/22. We excluded trials on diabetic foot infections, non-generalizable populations , and studies with outcomes not involving clinical resolution or cure. Abstracts with relevant clinical trial data were pulled and, if inclusion criteria met, had manuscript data abstracted. Two reviewers independently performed abstract and manuscripts reviews and data abstraction. For the network meta-analyses, the comparator antibiotic class was glycopeptides. Two analyses were performed using intention to treat (ITT) and clinically evaluable (CE) populations. Clinical trial quality was measured using the PEDro score.We reviewed 5,469 abstracts, of which 260 were pulled for data abstraction and 93 contained analyzable trial data . Clinical trial quality varied widely . We identified 26 analyzable antibiotic classes or combinations for comparison. In the ITT model, cure rate for oxazolidinones (OR 1.28 [95% CI 1.10-1.49]), flucloxacillin plus clindamycin (OR 1.61 [95% CI 1.03-2.53]), lipopeptides (OR 1.66 [95% CI 1.04-2.66]), and tetracyclines OR 1.69 [95% CI 1.24-2.30]), significantly differed from glycopeptides.  In the CE model, cure rate for oxazolidinones (OR 1.29 [95% CI 1.01-1.64]) and tetracyclines (OR 2.05 [95% CI 1.37-3.06]) significantly differed.We found that most (22/26) antibiotic classes or combinations used for SSTI treatment had similar efficacy to glycopeptides. However, oxazolidinones, lipopeptides, flucloxacillin plus clindamycin, and tetracyclines may have superior efficacy compared to glycopeptides. Treatment guidelines may wish to favor these latter classes for patients at high risk of treatment failure.Amy Y. Kang, Pharm.D., BCIDP, Paratek: Grant/Research Support Elliot I. Miller, BS, Epic Systems, Verona, WI: Employee Loren G. Miller, MD MPH, ContraFect: Grant/Research Support|GSK: Grant/Research Support|Medline: Grant/Research Support|Merck: Grant/Research Support|Paratek: Grant/Research Support"}
+{"text": "A safe and effective method to protect against cytomegalovirus (CMV) infection is a public health priority. mRNA-1647 is an investigational mRNA-based vaccine against CMV consisting of 6 mRNA sequences encoding 2 CMV antigens (glycoprotein B and the pentameric glycoprotein complex) in lipid nanoparticles in a lyophilized presentation.This phase 2, randomized, observer-blind, placebo-controlled, dose-finding trial of mRNA-1647 was conducted in CMV-seronegative and -seropositive healthy adults aged 18-40 years (NCT04232280). In Part 1, males and females were randomized 3:1 to receive mRNA-1647  or placebo at Months 0, 2, and 6. In Part 2, females were randomized 3:1 to receive mRNA-1647 100 \u00b5g or placebo at Months 0, 2, and 6. Primary endpoints were safety throughout the study, including solicited adverse reactions (ARs) up to 7 days after each dose, and neutralizing antibody (nAb) titers against epithelial cell infection and against fibroblast infection up to 12 months after last dose. Parts 1 and 2 results are combined.Figure). Robust nAb responses against epithelial cell infection were observed after each mRNA-1647 dose and sustained through the end of the study (12 months after last dose); nAb titers against epithelial cell infection in seronegative mRNA-1647 100-\u00b5g recipients exceeded the geometric mean titer of all seropositive recipients at baseline.Of 315 adults randomized, solicited ARs following dose 1 were reported in 54.7% (placebo), 84.4% (50 \u00b5g), 87.5% (100 \u00b5g), and 91.1% (150 \u00b5g) of seronegative adults and 59.3%, 94.4%, 94.6%, and 94.4%, respectively, of seropositive adults. The most common local and systemic solicited ARs across serostatus groups after dose 1 were pain  and fatigue , respectively. Similar trends in ARs were observed after doses 2 and 3. nAb titers against epithelial cell infection and against fibroblast infection were observed in all mRNA-1647 groups (Antibody-Mediated Immunogenicity of mRNA-1647 by CMV Serostatus(A) nAb titers against epithelial cell infection and (B) nAb titers against fibroblast infection are presented from day 1 through month 18 for CMV-seronegative (colored solid lines) and CMV-seropositive (colored dashed lines) treatment groups. Data are presented by serostatus, treatment group, and visit as GMT and corresponding 95% CI. Data are from the Per-Protocol Set. The solid black lines represent nAb GMTs against epithelial cell infection (GMT = 4575.7) and against fibroblast infection (GMT = 4215.5) for all CMV-seropositive participants at baseline (n = 87). Doses 1, 2, and 3 were administered on D1, M2, and M6, respectively, as represented by an arrow and syringe.CMV, cytomegalovirus; D, day; GMT, geometric mean titer; M, month; nAb, neutralizing antibody.mRNA-1647 was generally safe and well-tolerated and induced antigen-specific immune responses at all dose levels in both CMV-seronegative and -seropositive participants. These results guided selection of the 100-\u00b5g dose for the mRNA-1647 phase 3 trial.Lori Panther, MD, MPH, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Sandeep Basnet, MD, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Richard Leggett, DO, Crossroads Clinical Research: Contract employee James Peterson, MD, Moderna, Inc.: Received payment as a study investigator Jiang Lin, PhD, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Kai Wu, PhD, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Heather Lee, BS, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Roxane Hasselbeck, BA, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Andrew Natenshon, MA, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds Jacqueline Miller, MD, Moderna, Inc.: Employee|Moderna, Inc.: Stocks/Bonds"}
+{"text": "Asaia bogorensis strain SC1 isolated from a human-blood-fed Aedes aegypti mosquito crop. Metabolic pathway characteristics of aerobic respiration were present in the genome, along with multiple putative antibiotic resistance mechanisms.Understanding microbe-host interactions is key to combating disease transmission by mosquitoes. Here, we report the genome sequence of  Asaia bogorensis strain SC1, isolated from a human-blood-fed Aedes aegypti mosquito crop. Importantly, Asaia sp. is dominant in the crop of Ae. aegypti  Aedes aegypti mosquito crop. Briefly, the crop was dissected from a mosquito using ethanol-flamed forceps, homogenized in 1\u00d7 PBS pH = 7.4, and inoculated on Luria-Bertani (LB) agar at 30\u00b0C. To ensure a pure culture, individual colonies were sequentially streaked on fresh LB plates three times. DNA was extracted from overnight LB broth cultures  using the Qiagen DNeasy PowerSoil Kit  and quantified via Qubit . For long-read sequencing, the Native Barcoding Kit 24 V14 Kit  was used to barcode samples and prepare libraries. DNA was not sheared or size selected prior to library prep. Sequencing was performed with an R10.4.1 flow cell (FLO-MIN114) and minION device under high-accuracy mode (280 bp/s). Basecalling was performed using Guppy 6.4.6, and reads with quality scores <7 were removed. Reads were filtered with Filtlong 0.2.1 (ng 0.2.1  to removcy mode 20 bp/s. Bl FLO-MIN4 and minng 0.2.1 . Potenting 0.2.1 . Taxonomng 0.2.1  and the ng 0.2.1 .Asaia bogorensis (96.36% average nucleotide identity). GhostKOALA identified complete pathways for central carbon metabolism, including glycolysis and pyruvate oxidation, TCA cycle, and the pentose phosphate pathway. A complete oxidative phosphorylation pathway was annotated. Interestingly, pyruvate decarboxylase (pdc) was absent, but other putative fermentation genes, aldehyde dehydrogenases  and alcohol dehydrogenase (adhA), were present. GhostKOALA identified genes potentially conferring resistance to vancomycin, beta-lactams, and cationic antimicrobial peptides. The genome sequence of Asaia bogorensis strain SC1 from the mosquito crop provides a valuable resource for further investigating host-microbe interactions of public health interest.The SC1 genome is 3.4 Mbp with 59.69% GC content . GTDB id"}
+{"text": "Rf) genes. Accordingly, mechanisms underlying the control of the wide phenotypic variation in pollen number for breeding germplasm should be elucidated. This study aimed to identify complete linkage DNA markers responsible for male sterility at the MS-P1 region based on fine mapping. Two P-class pentatricopeptide repeat (PPR) family genes were identified as candidates for Rf based on predicted mitochondrial localization and higher expression in a male fertile variety/selected strain than in a male sterile variety. Eleven haplotypes (HT1\u2013HT11) at the MS-P1 region were defined based on genotyping of DNA markers. Association analysis of diplotypes at the MS-P1 region and the number of pollen grains per anther (NPG) in breeding germplasms harboring Kishu-cytoplasm revealed that the diplotypes in this region influenced NPG. Among these haplotypes, HT1 is a non-functional restorer-of-fertility (rf) haplotype; HT2, a less-functional Rf; HT3\u2013HT5 are semi-functional Rfs; and HT6 and HT7 are functional Rfs. However, the rare haplotypes HT8\u2013HT11 could not be characterized. Therefore, P-class PPR family genes in the MS-P1 region may constitute the nuclear Rf genes within the CMS model, and a combination of the seven haplotypes could contribute to phenotypic variation in the NPG of breeding germplasms. These findings reveal the genomic mechanisms of CMS in citrus and will contribute to seedless citrus breeding programs by selecting candidate seedless seedlings using the DNA markers at the MS-P1 region.In citrus breeding programs, male sterility is an important trait for developing seedless varieties. Sterility associated with the male sterile cytoplasm of Kishu mandarin (Kishu-cytoplasm) has been proposed to fit the cytoplasmic male sterility (CMS) model. However, it remains undetermined whether CMS in citrus is controlled by interactions between sterile cytoplasm and nuclear restorer-of-fertility ( These s Tanaka) . TherefoRf). The male sterile phenotype is a result of interaction between the CMS-associated gene and a non-functional restorer-of-fertility nuclear gene (rf)  Rf2 being the first  family  . To eluche first . Among t) family . These P) family , with ea) family . In partstigated . Three nstigated ; however1 populations of a cross between different varieties and selected strains harboring the Kishu-cytoplasm (MS-P1). This reduced NPG is linked to the haplotype block derived from kunenbo in the MS-P1 locus (Rf are located in the MS-P1 locus and to determine the influence of a combination of haplotype blocks (including Rf) on the phenotypic variations pertaining to male sterility and fertility in individuals with Kishu-cytoplasm.Male sterility and fertility segregate in the Fytoplasm . Specifiytoplasm . In addiytoplasm . These rytoplasm  and haveP1 locus . TherefoMS-P1 region within the locus using fine mapping, (2) identify candidates for Rf (Rf-MS-P1) through bioinformatic and transcriptional analysis, (3) define the number of allelic haplotypes at the MS-P1 region among the founders; and (4) reveal the association between NPG and the combination of allelic haplotypes (diplotype) at the MS-P1 region. Hybrid varieties, selected strains, and individuals in F1 populations (breeding germplasms) harboring the Kishu-cytoplasm as well as the various diplotypes in each individual were evaluated. Two mitochondrial-targeted PPR family genes at the MS-P1 region showed significant transcriptional differences between sterile and fertile varieties during flower development, indicating that they played an important Rf role in the CMS model. Assessing the combination allelic haplotypes, including the two PPR family genes, would enhance our understanding of the wide phenotypic variations observed in citrus male sterility and fertility associated with a sterile cytoplasm.To elucidate the molecular mechanism underlying male sterility in Kishu-cytoplasm and assess if it fits into the CMS genetic model, the present study aimed to: (1) identify the 1 populations, Okitsu No. 46 \u00d7 \u2018Kara\u2019 (O46-K), \u2018Sweet spring\u2019 \u00d7 Okistu No. 56 (SS-O56) and \u2018Harehime\u2019 \u00d7 Okistu No. 63 (H-O63), and the varieties/selected strains used in this study were maintained in the Division of Citrus Research, Institute of Fruit Tree and Tea Science, NARO   S2. The and 2013 . The NPG1 populations and the 85 varieties/selected strains through a modified protocol using cetyltrimethylammonium bromide and a high-salt precipitation solution . Six DNA markers  in the MS-P1 locus were used in a previous report (MS-P1 locus (C. clementina genome v1.0 (JGI) (https://phytozome-next.jgi.doe.gov/info/Cclementina_v1_0) (https://mikan.dna.affrc.go.jp/) were identified using BLAST (https://mikan.dna.affrc.go.jp/blast/) (https://archive.gramene.org/db/markers/ssrtool) (https://bioinfo.ut.ee/primer3/)  , was obtna_v1_0) . The cor/blast/) . The idessrtool) . The pririmer3/) . Dimericrimer3/) , as descrimer3/) .MS-P1 locus were identified based on the genotype segregation pattern. A graphical genotype of the MS-P1 locus was constructed for the physical map of C. clementina genome v1.0 between 5,153,708 and 19,761,190 bp in scaffold 8. The male sterile genotypes within the MS-P1 locus in recombinant individuals were identified through comparisons with the genotype of male sterile varieties/selected strains  were obtained from Phytozome (https://phytozome-next.jgi.doe.gov/info/Cclementina_v1_0) (Zea mays) Rf2 (U43082), rice (Oryza sativa) Rf2 (AB583697), rice Rf17 (Os04g0475900), and sugar beet (Beta vulgaris) Rf1 (AB646135), all of which belong to the non-PPR family of Rf, were used as the query to perform BLAST analysis against the sequences in the Mikan Genome DB under default settings. The protein sequences of the PPR family at the MS-P1 region were analyzed using TargetP v.2.0  (https://urgi.versailles.inra.fr/predotar/) (https://ihg.helmholtz-muenchen.de/ihg/mitoprot.html) (https://ppr.plantenergy.uwa.edu.au/)  (https://prosite.expasy.org/scanprosite/), and phylogenetic trees were constructed using the neighbor-joining method of MEGA v.10.0.5 (https://www.megasoftware.net) with protein sequences of rice (Oryza sativa) Rf1a (DQ311053), rice Rf1b (DQ311054), rice Rf4 (KJ680249), Sorghum (Sorghum bicolor) Rf1 (Brassica napus) Rfp1 (KX671967), Petunia (Petunia hybrida) Rf-PPR592 (AY10271), Ciclev10030242m, and Ciclev10030361m. Bootstrap values were calculated through a 1,000-permutation test.The gene locus, annotations, and protein sequences at the na_v1_0) . The DNAetP-2.0) , Predotaedotar/) , and Mitot.html)  to prediedu.au/) . Proteinserver/) , while Plor) Rf1 , ChineseC. clementina genome v1.0 (JGI) (https://phytozome-next.jgi.doe.gov/info/Cclementina_v1_0) using HISAT2 v.2.1.0. Transcripts per million (TPM) values were obtained to measure gene expression using StringTie v.2.1.1. TPM values were imported to Subio Platform v.1.24.5853  for normalization. Statistical analysis of the comparisons between \u2018Shiranuhi\u2019 and KyOw14 was carried out using the \u201ccompare 2 groups\u201d tool of the Subio platform. All RNA-seq data was deposited in the DDBJ Sequence Read Archive under accession number DRA015326.Bulked stamen from a sterile selected strain (KyOw14) and fertile variety (\u2018Shiranuhi\u2019) were collected seven days before flowering (DBF) and 1 DBF in a field from the Division of Citrus Research, Institute of Fruit Tree and Tea Science, NARO , as two biological replicates. Total RNA was isolated using an RNeasy Plant Mini Kit , while the RNA-seq analysis was carried out by Hokkaido System Science Co., Ltd. . RNA-seq libraries were generated from the total RNA using the NEBNext Ultra RNA Library Prep Kit for Illumina . The 150 bp paired-end sequencing of RNA-seq libraries was performed using the NovaSeq 6000 system , and the sequenced reads were trimmed using fastp v.0.22.0 and mapped to https://phytozome-next.jgi.doe.gov/info/Cclementina_v1_0)  . Quantitative PCR was performed QuantStudio3 Real Time PCR System  using Power SYBR Green PCR Master Mix (Thermo Fisher Scientific) under 10 min at 95\u00b0C, followed by 40 cycles of 15 s at 95\u00b0C and 60 s at 60\u00b0C. The primers for the gene expression of Ciclev10030242m , Ciclev10030361m , and Ciclev10029947m  were designed at a specific region or 3\u2019UTR referred by Phytozome (na_v1_0) . The expGGATCAT) . SpecifiThe sources of cytoplasm in the 85 varieties/selected strains were determined referring to the pedigree chart .1 populations   Table S1d H-O63) , althougd H-O63)  of genotee chart .https://www.jichi.ac.jp/saitama-sct/SaitamaHP.files/statmedEN.html)  .MS-P1 locus between 33.5cM (TSRF161) and 69.1cM (SSR08B66) on linkage group 8 (LG8) in the corresponding linkage map of Okitsu No. 46 \u00d7 Okitsu No. 56  , 66 functional genes, including 10 PPR family genes, were predicted on the 920 kb genomic region, corresponding to the newly refined MS-P1 region (Rf2 (U43082), rice Rf2 (AB583697), rice Rf17 (Os04g0475900), or sugar beet Rf1 (AB646135), which are non-PPR families of Rf, were not found among the 66 genes here. Bioinformatic analysis using PPRfinder revealed that all PPR family genes at the MS-P1 region belonged to the P-class subfamily  should be expressed more in a male fertile variety/selected strain than in a male sterile variety/selected strain. Transcriptomic analyses of KyOw14 (sterile selected strain) and \u2018Shiranuhi\u2019 (fertile variety) stamens were performed at 7 and 1 DBF through RNA-seq analysis. RNA extracted from the stamens (two replicates) were used to build libraries for RNA-seq (MS-P1 region of KyOw14 and \u2018Shiranuhi\u2019 was performed (p< 0.05)  than in KyOw14 (sterile strain). In addition, the existence of Ciclev10030242m and Ciclev10030361m at the MS-P1 region provides evidence that molecular mechanisms underlying citrus male sterility fit into the general plant CMS model.The protein sequence analysis showed that Ciclev10030242m and Ciclev10030361m were highly similar proteins, with 86.66% overlap , ChineseRf or rf are located; therefore, we defined allelic haplotypes at the MS-P1 region among the founders of Japanese breeding germplasm. The nearest markers to candidates for Rf-MS-P1 were narrowed down to 00918-1 through fine mapping of the MS-P1 region  were genotyped (1 populations except for \u2018Willking\u2019 (see legend in MS-P1 locus (C. tangerina hort. ex Tanaka), Iyo (C. iyo hort. ex Tanaka), Ponkan (C. reticulata Blanco), or Willowleaf mandarin (C. deliciosa Ten.)  were investigated (MS-P1 region (p< 0.05) (MS-P1 region (p < 0.05) (p < 0.05)   corresponding to the MS-P1 region was derived from Willowleaf mandarin, which harbors HT6  to identify Rf-MS-P1 candidates was a reasonable strategy.The results here also suggest that plotypes Table\u00a02.bors HT6 ; therefoMS-P1 (MS-P1 region were not observed (data not shown). \u2018Nishinokaori\u2019 and \u2018Kara\u2019 exhibited an increased NPG, although \u2018Nishinokaori\u2019 harbored HT1/HT3 and \u2018Kara\u2019 harbored HT1/HT4, both of which are partial male sterile diplotypes  were detected in our previous study .The original contributions presented in the study are publicly available. This data can be found here: DDBJ, accession DRA015326 (SG acquired funding, designed the study, performed experiments, analyzed data, and wrote the original draft. SG, HF, TE, ToS, and TaS designed the study. HH, SO, and KN produced and managed materials. SG and TaS reviewed and edited this manuscript. All authors contributed to the article and approved the submitted version."}
+{"text": "Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, has infected >600 million people in the ongoing global pandemic. Several variants of the SARS-CoV-2 have emerged in the last >2 years, challenging the continued efficacy of current COVID vaccines. Therefore, there is a crucial need to investigate a highly cross-protective vaccine effective against variants of SARS-CoV-2. In this study, we examined seven lipopeptides derived from highly conserved, immunodominant epitopes from the S, N, and M proteins of SARS-CoV-2, that are predicted to contain epitopes for clinically protective B cells, helper T cells (TH) and cytotoxic T cells (CTL). Intranasal immunization of mice with most of the lipopeptides led to significantly higher splenocyte proliferation and cytokine production, mucosal and systemic antibody responses, and induction of effector B and T lymphocytes in both lungs and spleen, compared to immunizations with the corresponding peptides without lipid. Immunizations with Spike-derived lipopeptides led to cross-reactive IgG, IgM and IgA responses against Alpha, Beta, Delta, and Omicron Spike proteins as well as neutralizing antibodies. These studies support their potential for development as components of a cross-protective SARS-CoV-2 vaccine. Recent reports suggest that nAb responses, elicited by vaccines and/or natural infections, is an accurate predictor of protection against SARS-CoV-2 \u20135. Gener+/CD8+ T cells have shown to induce cross-reactive immunity that is resistant to mutations acquired by SARS-CoV-2 variants of concerns (VOCs) and effectively preventing VOC escape  and membrane (M) proteins, in addition to the spike protein . FurtherC escape . For a bin vitro and in vivo assays. We found that lipopeptides existed as larger micelle-like particles compared to their peptide counterpart, and stimulated APCs by the upregulation CD40, CD86 and HLA-DR molecules without the activation of TLR-2 and TLR-4 receptors. Intranasal immunizations with individual lipopeptides in mice generated stronger antigen-specific cellular, and mucosal as well as systemic humoral immune responses, compared to their native peptide immunizations. In addition, we provide evidence of the induction of cross-protective immunity upon mucosal immunization with the designed lipopeptides.In this study, we identified seven highly conserved, cross-reactive, immunodominant peptides from the S, N, and M proteins of SARS-CoV-2, MERS-CoV, SARS-CoV and other common cold coronaviruses that are predicted to contain epitopes for clinically protective B cells, helper T cells (TH) and cytotoxic T cells (CTL), and which also bind to multiple MHC class I and II molecules (covering >98% of human population) Table\u00a01 1, P3, P5, P7, P9, P11, and P13, ranged from 134.2-229.8 nm, 356.2-2792 nm, 79.32-309.9 nm, 1256- 4033 nm, 125.6- 1510 nm, 246.2-1856 nm, and 361-3554 nm, respectively , P4 (41.7%), P5 (7.1%), P6 (5.7%), P7 (9.6%), and P8 (17.3%) (3 (174.0%), P5 (145.5%), P6 (9.4%), P7 (15.9%), P8 (23.5%), P9 (13.9%), P10 (26.2%), P12 (4.1%), and P13 (2.4%) (5 (110.0%), P7 (7.9%), P8 (244.7%), P9 (400.7%), P10 (386.9%), P11 (332.1%), P12 (267.6%), P13 (340.0%), and P14 (289.7%) (1 (p=0.0028), P3 (p=0.0291), P5 (0.0053), P9 (p<0.0001), P11 (p=0.0015), and P13 (p=0.0005), and native peptide stimulation with P8 (p=0.002), P10 (p<0.0001), P12 (p=0.0056), and P14 (p=0.0029). THP-1 cells stimulated with lipopeptides, or peptides did not produce detectable levels of proinflammatory cytokines  or stimulate human TLR-2 and -4 receptors . We found that Pnd immunization, spleens were examined for antigen-specific proliferation responses using a colorimetric BrdU incorporation assay. Splenocytes from all immunized groups were also stimulated with ConA (T cell mitogen), which was used as a positive control  and P2   twice, 14 days apart. Eight days after the 21-P14, that were restimulated ex vivo with their respective native peptides (1 \u03bcg/ml). We found that splenocytes from lipopeptide immunizations, P1, P3, P9, and P13, produce higher levels of IFN-\u03b3, TNF-\u03b1, IL-2, and IL-4, compared to their respective peptide immunization groups , TNF-\u03b1 , and IL-2  significantly correlated with antigen-specific splenocyte proliferation responses , and serum samples collected from P5 (p<0.0001), P6 (p<0.0001), P7 (p=0.0041), P9 (p<0.0001), and P10 (p=0.0004) immunizations elicited a significantly higher anti-(S-N fusion)-IgA antibody response, compared to unimmunized controls (7 (p=0.0011) and P9 (p=0.0242) induced significantly higher anti-(S-N fusion)-IgA antibody titers, compared to their respective peptide immunizations. P5 induced a significantly higher anti-(S-N fusion)-IgA titer at a 1:2 dilution, compared to the P6 (p<0.0001) (5-P10) led to a significant anti-(N)-IgA antibody titer, compared to unimmunized controls -IgA titers compared to their respective peptide, with p<0.0001 and p=0.0347, respectively. Nucleocapsid-derived lipopeptide immunizations showed a strong mucosal anti-(S-N fusion)-IgA and anti-(N)-IgA antibody response compared to their respective nucleocapsid-derived peptide immunization. In contrast, membrane-derived lipopeptide and peptide (P11-P14) immunizations did not elicit anti-(M)-IgA antibodies (BAL fluid from P1-P14 immunized mice (1-P4) and nucleocapsid-derived (P5-P10) lipopeptide and peptide immunizations produced significant amounts of anti-(S-N fusion)-IgM antibodies (5-P10 immunized mice induced significantly higher anti-(N)-IgM antibodies, compared to the unimmunized controls (7 immunization induced a significantly higher anti-(S-N fusion)-IgM and anti-(N)-IgM antibody response, compared to its native peptide immunization . Also, P9 immunization induced a significantly higher anti-(N)-IgM antibody titer, compared to P10 (p<0.0001). Membrane-derived lipopeptide and peptide immunizations with P11 and P12 produced significantly higher anti-(M)-IgM antibody titers, compared to unimmunized controls (p<0.0001) -IgG antibody titres, compared to unimmunized controls (p<0.0001) (1 was significantly higher compared to its native peptide immunization (p<0.0001). Notably, P3 and P4 immunizations produced the highest anti-(S-N fusion)-IgG antibody titer compared to all other immunization groups. All nucleocapsid-derived lipopeptide and peptide immunizations have significantly higher anti-(N)-IgG antibody titres, compared to unimmunized controls (p<0.0001) (5 and P9 immunizations had significantly higher anti-(N)-IgG antibody titres, compared to their respective native peptide immunizations, with p=0.0036 and p=0.0089, respectively. Furthermore, we found that anti-(M)-IgG antibodies were only produced by P11 immunizations (1-P4), and nucleocapsid- (P5-P10) derived lipopeptide and peptide immunizations to their respective antigens.Lastly, we measured systemic IgG antibody responses against the S-N fusion, N, and M proteins of SARS-CoV-2. Immunizations with spike-derived and nucleocapsid-derived lipopeptides and peptides induced strong IgG antibody responses to their respective antigens. P1-P4) lipopeptides and peptides against SARS-CoV-2 VOCs: Alpha (B.1.1.7), Beta (B.1.351), Delta (B.1.617), and Omicron (B.1.1.529). P1 immunization showed significantly higher mucosal IgA antibody titers, compared to unimmunized controls and its respective peptide immunization, against all SARS-CoV-2 VOCs  , B.1.351 (p=0.0049), B.1.617 (p=0.0188), and B.1.1.529 (p<0.0001) variants of SARS-CoV-2   and B.1.617 (p=0.0088) variants , B.1.351 (p<0.0001), and B.1.1.529 (p=0.0059). Similarly, P3 immunizations induced significantly higher IgG antibody titers compared to P4, against the B.1.1.7, B.1.351, B.1.617, and B.1.1.529 variants (p<0.0001). Evidently, the spike-derived lipopeptide immunizations induced a more robust cross-variant IgG antibody response, compared to their respective peptide immunizations.From serum samples collected from P1-P4 immunizations were tested for their ability to block/neutralize viral binding to target host ACE2 receptor, using a cPass SARS-CoV-2 Neutralization kit. Serum from a non-spike lipopeptide immunization, P5, was used as a negative control. Serum antibody responses elicited by P1 and P3 immunizations showed a percent inhibition of 23.5% and 22.0%, respectively  had significantly higher neutralizing capabilities compared to their respective peptides . Next, we found that BAL antibody responses from P1 (23.7%) and P2 (20.3%) immunization showed significantly higher neutralizing capabilities compared to P5 , whereas P3 (12.1%) and P4 (12.2%) immunizations showed similar neutralization capability.Antibody responses in BAL and serum samples collected from Pectively Figure\u00a06+CD3-), helper T cells , and cytotoxic T cells  to identify functional changes among the immunization groups. Using clustering analysis, we identified cell clusters (Pop#) within B cells, TH and CTLs , TH cells with B cell and CTL helper function (3.0% vs. 0.0%), CTL helper function (43.9% vs. 12.7%), and B cell helper function (34.3% vs. 0.8%), and CTLs with effector (25.1% vs. 20.5%) and pleiotropic (46.2% vs. 33.6%) properties, compared to peptide immunizations , and higher percentage of regulatory B cells (45.0% vs. 31.1%), compared to peptide immunizations  activated APCs more efficiently, compared to their respective peptides, enabling the integration of the innate and adaptive systems without the need of an adjuvant. Furthermore, we found that lipopeptides did not stimulate TLR-2 and -4 receptors or produce detectable amounts of proinflammatory cytokines (The upregulation of antigen-presenting (HLA-DR) and costimulatory (CD40/CD86) molecules demonstrate enhanced capacity to present antigen on APCs and prime T cells. Interestingly, we found that lipopeptides derived from SARS-CoV-2 antigens led to higher upregulation of HLA-DR, CD40, and CD86 in comparison to the corresponding peptides and medium-alone control , 36. Thie system . Our resytokines . These p1, P3, P5, P9, and P13) induced significantly higher recall responses, upon ex vivo re-stimulation by corresponding lipopeptides and native peptides, compared to peptide immunizations  may drive T cells into anergic states. Anergic conditions are not desirable for a vaccine as it demonstrates the failure of our immune system to mount a response against the targeted antigen  and TH2 (IL-4) cytokines have been suggested to modulate immune responses towards cell-mediated or humoral immunity. IFN-\u03b3 functions to educate immune cells to recognize and eliminate pathogens, and associates with protective immunity against intracellular pathogens  marked the presence of long-lasting, antigen-specific cellular responses  and cellular (cytotoxic T cell clusters) immune responses  demonstrated a cross-reactive mucosal IgA and systemic IgM/IgG responses against four different SARS-CoV-2 VOCs   responses, compared to the corresponding peptide groups Figure\u00a04he blood . The indIn conclusion, our results clearly demonstrate that intranasal immunization with lipopeptides, even without any added adjuvant, lead to significant induction of mucosal antibody responses and systemic cellular and humoral immune responses. This study describes an important and innovative finding that opens new avenues for developing a broadly protective vaccine for SARS-CoV-2 virus and its variants, and potentially other heterologous coronaviruses.1, P3, P5, P7, P9, P11, and P13) and their corresponding native peptides , derived from highly conserved and functional regions of the S-, N- and M-proteins of SARS-CoV-2, were custom synthesized by Genscript Inc.  with >96% purity , and a standardized Sizing and Zeta report was generated by Zetasizer XPLORER software v1.10 to provide mean particle size for each sample.6 cells/ml) were seeded with P1-P14, at 1 \u03bcg/ml. The cultures were incubated in 5% CO2, at 37\u00b0C for 24 hours. The activation of THP-1 monocytes was determined by flow cytometry analysis. A total of 2\u00d7105 THP-1 cells from each culture was stained with co-stimulatory markers  using established procedures from Thermofisher , and subsequently, performed a percent of control calculation (Percent of Control =100 \u00d7 iMFIExp./iMFIControl) for graphing (THP-1 monocytes (ATCC TIB-202) were grown in medium containing RPMI-1640 (Gibco), 10% fetal bovine serum (FBS), 1% penicillin-streptomycin (P/S), and 1% L-glutamine, and maintained at a confluency between 70-80%. THP-1 cells  cell lines are stably transfected with human TLR2 and TLR4 genes. Upon TLR signalling, a secreted embryonic alkaline phosphatase (SEAP) reporter gene is induced, and SEAP is secreted, which is detected by a QUANTI-Blue\u2122 detection assay.5 cells/ml) and HEK-Blue\u2122 hTLR4 (2.5\u00d7105 cells/ml) cells were seeded with P1-P14, at 1 \u03bcg/ml and 10 \u03bcg/ml, in triplicates. The cultures were incubated in 5% CO2, at 37\u00b0C for 24 hours, and supernatants were collected to run QUANTI-Blue\u2122 detection according to the assay protocol. Using a DTX 880 Plate Reader (Beckman Coulter), optical density (OD) readings were taken of QUANTI-Blue\u2122 detection assays, at 620nm. Data was expressed as mean \u00b1 SEM of triplicate cultures.HEK-Blue\u2122 hTLR2 and HEK-Blue\u2122 hTLR4 cell lines were grown according to their respective handling procedures. HEK-Blue\u2122 hTLR2  for Health Sciences and were conducted according to the guidelines of the Canadian Council of Animal Care (CCAC). Four to six-week-old male C57BL/6 mice were purchased from Charles River Laboratory and housed in a pathogen-free animal facility (HSLAS) at the University of Alberta. Mice were immunized twice, 14 days apart with individual lipopeptides or peptides (10 \u03bcg/mouse), intranasally in a total volume of 30 \u03bcL (15 \u03bcL in each nostril). Mice were euthanized 8 days after second immunization and bronchoalveolar lavages (BAL), sera, lungs, and spleens were collected. Unimmunized mice were used as controls.5 cells splenocytes from immunized mice and 4\u00d7105 antigen presenting cells  were incubated with corresponding lipopeptides and peptides at various concentrations described in the figure legends  and isolated lung lymphocytes (1\u00d7105 cells) from immunized mice were stained with extracellular  and intracellular markers  using established protocols from Thermofisher  1X for 4h at 37\u00b0C and subsequently stained with extracellular and intracellular markers as mentioned above. Samples were run on Attune NxT flow cytometer  and analyzed by FlowJo v10.8 Software. Lung and spleen samples from immunized groups P1-P14 were concatenated to provide consistencies in clustering methods and creating tSNE (t-distributed stochastic neighbor embedding) plots. Using Phenograph and FlowSOM clustering algorithms, lung lymphocytes and splenocytes were classified into different clusters based on phenotypic markers that were expressed, and overlayed on tSNE plots. Violin Box was used to acquire MFI values of phenotypic markers that each cluster expressed, which was represented by heatmaps, created on Morpheus . The U-PLEX Biomarker Group 1 (mouse) assay was used to analyze cytokines profiles of IFN\u03b3, IL-10, IL-2, IL-4, and TNF\u03b1. The MSD plates were read on MESO QuickPlex SQ120 and analyzed on DISCOVERY WORKBENCH 4.0 Analysis Software.Cytokine concentrations were measured from supernatants collected from antigen-specific splenocyte proliferation cultures of PSerum and BAL samples were pooled from three immunized mice and ran in duplicates for each dilution, on 96-well plates. The plates were coated with SARS-CoV-2 nucleocapsid, membrane, a spike-nucleocapsid fusion protein, and the spike proteins of Alpha (B.1.1.7), Beta (B.1.351), Delta (B.1.617), and Omicron (B.1.1.529) variants . For detection of IgG and IgM in serum, and IgA in BAL, the antibody ELISA procedure described earlier was followed . The abs1-P4 immunized groups were tested for neutralizing antibodies. Triplicate samples were run using the cPass SARS-CoV-2 Neutralization Antibody Detection Kit according to the manufacturer\u2019s instructions. Serum and BAL samples were run at 1:9 dilutions and undiluted, respectively, and read using a DTX 880 Plate Reader (Beckman Coulter). Data are expressed as mean \u00b1 SEM of triplicates.Serum and BAL samples from PP \u2264 0.05 was used to indicated significance.Data were analyzed, and graphed using GraphPad Prism Software 9.4.1. Data was presented as mean \u00b1 SEM of 2-3 replicate values of 3 pooled mice and statistical differences were analyzed by one-way or two-way ANOVAs, adjusted for multiple comparisons. In addition, Spearman\u2019s test was performed to determine the correlations between cytokines and proliferation values. A The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The animal study was reviewed and approved by University of Alberta\u2019s Animal Care and Use Committee (ACUC) for Health Sciences (AUP00000212) and according to the guidelines of the Canadian Council of Animal Care (CCAC).RP, design, planning, execution of experiments, data analyses, writing first draft and editing. BA, conception, funding, design, planning, supervision, data analyses, writing and editing. All authors contributed to the article and approved the submitted version."}
+{"text": "The minimum inhibitory concentrations (MICs) were tested using a Trek Diagnostic System (Thermo Fisher Scientific). Susceptibility was determined by CLSI broth microdilution and interpreted with CLSI M100 (2021) breakpoints.From 2017-2021, 554 Gram-negative isolates were collected from 16 pediatric departments across China. GNB isolates from pediatric departments were mainly derived from bloodstream (n = 135), intraperitoneal (n = 129), lower respiratory tract (n = 198) and urinary tract (n = 92) infections (E. coli (34.30%) and K. pneumoniae (27.62%) were the most common pathogens, followed by P. aeruginosa (15.34%). Susceptibility of species with more than 20 isolates were showed in Table 1. The susceptibility of P. aeruginosa to C/T was 89.41%, which was the highest among beta-lactams and was second only to amikacin. The susceptibilities of E. coli and K. pneumoniae isolates to C/T were 92.63% and 58.17%, respectively. When exclude carbapenem-resistant E. coli and K. pneumoniae, the susceptibility to C/T increased to 96.17% and 86.14%. C/T showed similar activities to E. coli and K. pneumoniae isolated from pediatric patients < 1 year (Table 2). Thirteen P. aeruginosa isolates were collected from patients < 1 year and 13/13 were susceptible to C/T (Table 3).P. aeruginosa and E. coli obtained from pediatric patients in China showed a high susceptibility to C/T. When excluding carbapenem resistant isolates, C/T also showed good activities against K. pneumoniae.Pengcheng Li, M.D., MSD China: Employee of MSD China"}
+{"text": "Polybrominated diphenyl ethers (PBDEs) are commercially used flame retardants that bioaccumulate in human tissues, including breast milk. PBDEs produce endocrine and metabolic disruption in experimental animals and have been associated with diabetes and metabolic syndrome (MetS) in humans, however, their sex-specific diabetogenic effects are not completely understood. Our past works show glucolipid dysregulation resulting from perinatal exposure to the commercial penta-mixture of PBDEs, DE-71, in C57BL/6 female mice.As a comparison, in the current study, the effects of DE-71 on glucose homeostasis in male offspring was examined. C57BL/6N dams were exposed to DE-71 at 0.1 mg/kg/d (L-DE-71), 0.4 mg/kg/d (H-DE-71), or received corn oil vehicle (VEH/CON) for a total of 10 wks, including gestation and lactation and their male offspring were examined in adulthood.In vivo glucose challenge showed marked glucose intolerance (H-DE-71) and incomplete clearance (L- and H-DE-71). Moreover, L-DE-71-exposed mice showed altered glucose responses to exogenous insulin, including incomplete glucose clearance and/or utilization. In addition, L-DE-71 produced elevated levels of plasma glucagon and the incretin, active glucagon-like peptide-1 (7-36) amide (GLP-1) but no changes were detected in insulin. These alterations, which represent criteria used clinically to diagnose diabetes in humans, were accompanied with reduced hepatic glutamate dehydrogenase enzymatic activity, elevated adrenal epinephrine and decreased thermogenic brown adipose tissue (BAT) mass, indicating involvement of several organ system targets of PBDEs. Liver levels of several endocannabinoid species were not altered.Compared to VEH/CON, DE-71 exposure produced hypoglycemia after a 11\u00a0h fast (H-DE-71). An increased fast duration from 9 to 11\u00a0h resulted in lower blood glucose in both DE-71 exposure groups. Our findings demonstrate that chronic, low-level exposure to PBDEs in dams can dysregulate glucose homeostasis and glucoregulatory hormones in their male offspring. Previous findings using female siblings show altered glucose homeostasis that aligned with a contrasting diabetogenic phenotype, while their mothers displayed more subtle glucoregulatory alterations, suggesting that developing organisms are more susceptible to DE-71. We summarize the results of the current work, generated in males, considering previous findings in females. Collectively, these findings offer a comprehensive account of differential effects of environmentally relevant PBDEs on glucose homeostasis and glucoregulatory endocrine dysregulation of developmentally exposed male and female mice. Considerable evidence points to a potential contribution of endocrine- (EDCs) and metabolism-disrupting chemicals (MDCs) to the rapid increase in the incidence of these metabolic diseases . One claetration \u201314. Thervia exposed dams and accumulate in male and female offspring liver and brain  is informative in understanding the diabetic phenotype of T2D. The incretin GLP-1 is secreted by enteroendocrine L-cells and acts on pancreatic \u03b2-cells to promote postprandial insulin secretion. In T2D, this insulinotropic effect is either impaired or absent leading to hyperglycemia . ActivatHaving observed sexually dimorphic risk and susceptibility of perinatally exposed females to MetS , 29, thead libitum in glass water bottles. Procedures on the care and treatment of animals were performed in compliance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals and approved by the University of California, Riverside, Institutional Animal Care and Use Committee (AUP#20170026 and 20200018).C57BL/6N mice were obtained from both Charles River Laboratories  and Taconic Biosciences . Mice were group housed 2-4 per cage and maintained in a non-specific pathogen free vivarium on a 12\u00a0h light/dark cycle at an ambient temperature of 20.6-23.9 \u00b0C and relative humidity of 20-70%. Mice were provided rodent chow  and municipal tap water Dosing solutions were prepared as described previously , 32. In via the dam was accomplished as described previously  and ex vivo analysis of hepatic, adipose, adrenal, and endocrine parameters (Cohort 2). The DE-71 exposure and testing paradigm is shown in Perinatal PBDE exposure eviously . In brieeviously Figure\u00a01eviously , 20, 38,ex ratio . Male ofITT, was calculated using the formula (0.693 x 100) x t1/2-1 to determine in vivo insulin sensitivity from 0-30\u00a0min post insulin injection. Fasting blood glucose was determined from baseline values (t=0) obtained in IPGTT (11h) and ITT (9h). GTT and ITT sample sizes were different because ITT effect size was expected to be less (as estimated by our previous work (Mice were fasted overnight (ON) for 11h and then injected with glucose . Glucose was measured directly from tail blood (~1 uL) at time 0, 15, 30, 60, and 120\u00a0min post-glucose challenge. A calibrated glucometer and test strips  were used to measure plasma glucose concentrations. Seven days following IPGTT, an insulin tolerance test (ITT) was performed with Humulin R (Eli Lilly) bolus  on mice fasted ON for 9h. Tail blood was collected, and glucose was sampled in the same manner as IPGTT . For area calculations, the area under (AUC) or above the glycemia curve (inverse AUC) from 0-120\u00a0min post injection was used. The percent blood glucose reduction rate after insulin administration, Kous work ) and, thDuring sacrifice, under terminal isoflurane anesthesia, cardiac blood (0.3-1 mL) was collected and centrifuged at 16,000 x g for 20\u00a0min at 4 \u00b0C. After the addition of a cocktail of protease inhibitors and EDTA, the plasma samples were stored at -80\u00b0C until further use. The following organs were excised and weighed: liver, pancreas, spleen, adrenal glands and interscapular brown adipose tissue (BAT). Plasma, liver and adrenal samples were snap-frozen over dry ice and stored at -80 \u00b0C for later analysis of plasma hormones, adrenal epinephrine, liver endocannabinoids and enzymatic activity.via cardiac puncture (ad libitum fed state) was analyzed for metabolic hormones using commercially available kits according to manufacturer\u2019s instructions. Plasma insulin was measured using commercial ELISA kits  as previously described , 10 pmol d4-oleoylethanolamide , and 500 pmol d5-2-arachidonoyl-sn-glycerol . Following homogenization of samples, 2 mL of chloroform and 1 mL of water were added, followed by centrifugation at 2000 x g for 15\u00a0min at 4\u00b0C. The organic phase was extracted and the pooled lower phases were dried under N2 gas followed by resuspension in 0.1 mL methanol:chloroform (9:1). Analysis of endocannabinoids (EC) was performed via ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC/MS/MS) as previously described 6. The mixture was incubated at 0\u00b0C for 20\u00a0min and the oxidation reaction was stopped by the addition of 60 \u00b5L of a 9\u00a0N NaOH solution containing 4 mg/ml ascorbic acid . Fluorescence emission was determined at 520 nm (excitation wavelength at 420 nm) using a fluorescence plate reader (Promega). Epinephrine concentration  was converted from the mean fluorescence intensity units of each sample using calibration standards and polynomial curve fitting.Epinephrine content in adrenal glands was measured using a modification of the trihydroxyindole method as described previously . Brieflypost hoc testing for multiple group comparisons. For parametric ANOVAs, Tukey\u2019s post hoc test was used when both sample sizes and variance were equal, and a Dunnett\u2019s T3 test was used when sample size and variance were not equal. For non-parametric ANOVA, a Dunn\u2019s post hoc test was performed. For Two-way ANOVA, Tukey\u2019s, Sidak\u2019s and Holm-Sidak\u2019s post hoc tests were used. Differences were deemed significant at p<0.05. Data are expressed as mean \u00b1 standard error of the mean (s.e.m) unless indicated otherwise.Statistical analyses were performed using GraphPad Prism v.9.4.1. A one-way analysis of variance (ANOVA) was used to test the main effect of one factor. When normality assumption failed, as determined using a Shapiro-Wilk test, a Kruskal-Wallis H ANOVA was used. A Brown-Forsythe ANOVA was used if the group variances were significantly different. Data for fasting glycemia were analyzed by two-way ANOVA for main effects of exposure and fasting duration. ITT and GTT experiments were analyzed by repeated measures two-way or mixed model ANOVA to determine main effects of exposure and time. ANOVA was followed by Exposure effect F=9.34, p<0.001, Tukey\u2019s post-hoc VEH/CON vs L-DE-71, p=0.03 and lower in L-DE-71 vs H-DE-71, p=0.0003). Relative liver weight was 8% greater in H-DE-71 relative to L-DE-71 =2.824, p=0.0675, Tukey\u2019s post-hoc L-DE-71 vs H-DE-71 p<.05). The absolute and relative weights of pancreas and spleen were similar across groups.Exposure effect F= 6.65, p<0.01; Time effect F= 24.6, p<0.0001; Exposure x Time F= 0.76 ns; Sidak\u2019s post hoc test, 11\u00a0h: VEH/CON vs H-DE-71, p<0.01; 9\u00a0h vs 11\u00a0h, L-DE-71 p<0.01, H-DE-71, p<0.05 (We examined fasting blood glucose (FBG) after 9 and 11\u00a0h fasting using glycemia values from basal time points obtained in ITT and GTT experiments, respectively. , p<0.05 Figure\u00a02=4.767, p<0.05; Time effect F=330.2, p<0.0001; Time x Exposure F =3.112, p<0.01)  and 60\u00a0min post injection (p<0.05) and in L-DE-71 vs H-DE-71 at t=30 (p<0.01) and 60\u00a0min post injection (p<0.01).To investigate the effects of DE-71 on glucose tolerance, glycemia was measured during GTT over the 120\u00a0min post-injection time course =278.6, p<0.0001; Exposure effect F=9.39 p<0.001; Time x Exposure F=5.94, p<0.0001). Mean glycemia values for H-DE-71 were significantly different from VEH/CON at t=15 , 30 (p<0.01) and 60\u00a0min post injection (p<0.01) and from L-DE-71 at t=15 (p<0.01), t=30 (p<0.01) and 60\u00a0min post injection (p<0.001).Since FBG after an 11\u00a0h fast was lower in H-DE-71 relative to VEH/CON =5.21, p<0.05, Tukey\u2019s post-hoc VEH/CON vs H-DE-71, p<0.029, L-DE-71 vs H-DE-71, p<0.013; Percent basal glycemia AUC, Brown-Forsythe ANOVA: Exposure effect F=10.1, p<0.001, Dunnett\u2019s T3 post-hoc VEH/CON vs H-DE-71, p<0.004, L-DE-71 vs H-DE-71 p<0.0003 .The differences in magnitude and duration of glycemia are integrated using the area under the glucose curve, AUCTime effect F=56.2, p<0.0001; Exposure effect F=2.40, ns; Time x Exposure F=1.90, p<0.05) . When glycemia is expressed as a percent of baseline, L-DE-71 exposed mice displayed incomplete recovery or glucose clearance/utilization at 90 and 120\u00a0min after insulin injection =33.7, p<0.0001; Exposure effect F=8.014, p<0.001; Time x Exposure F=1.23, ns)  and 120\u00a0min (p<0.001). This is represented as a greater mean latency to reach the minimum insulin-induced hypoglycemia in L-DE-71 relative to VEH/CON =9.07, p<0.01; Dunn\u2019s post hoc VEH/CON vs L-DE-71, p<0.01)  was plotted using absolute =2.024, ns, Sidak\u2019s post-hoc VEH/CON vs L-DE-71, p<0.05) (ITTinsulin measured over the first 30\u00a0min post-injection (The inverse area under the glucose response curve (inverse AUCad libitum fed state) =3.684, p=0.0409; Dunnet\u2019s post-hoc VEH/CON vs L-DE-71 p=0.0473; Glucagon, One-way ANOVA: Exposure effect F=5.61, p=0.02; Dunnett\u2019s T3 post-hoc VEH/CON vs L-DE-71 p=0.0449) =16.06, p<.0001; Tukey\u2019s post-hoc VEH/CON vs L-DE-71 p=0.0001, VEH/CON vs H-DE-71 p=0.002). Adrenal weights were not different across experimental groups =6.82, p=0.061; Tukey\u2019s post-hoc VEH/CON vs L-DE-71 p=0.01, L-DE-71 vs H-DE-71 p=0.01) =19.51, p<.0001; Dunnett\u2019s post-hoc VEH/CON vs L-DE-71 p=0.002, VEH/CON vs H-DE-71 p<0.0001) (p=0.03).Exposure to DE-71 decreases glucose tolerance which may be due to enhanced hepatic glucose production. To examine this possibility, we tested the hypothesis that PBDEs increase the activity of GDH a hepatic gluconeogenic enzyme. We found that exposure to L- and H-DE-71 significantly <0.0001) Figure\u00a07Male F1 mice exposed to either dose of DE-71 displayed normal liver levels of the endocannabinoid (EC), anandamide , and related fatty acid ethanolamides, docosahexaenoyl ethanolamide (DHEA) and oleoylethanolamide (OEA), when compared to VEH/CON . The combination of DE-71 effects - exaggerated fasting hypoglycemia, glucose intolerance and incomplete glucose clearance/utilization in response to insulin challenge demonstrates the complex actions of PBDEs on glucose homeostasis in males. Previously, we have observed abnormal glucose metabolism and other parameters in their DE-71-exposed female siblings  activity increases energy expenditure by burning fat and increasing metabolic rate and can utilize glucose especially when stimulated by insulin . TherefoIt is important to note that developmental exposure to DE-71 ends at weaning, but that dysregulated glucoregulatory phenotypes in exposed male and female offspring are observed in adulthood . We specin vivo and ex vivo parameters measured in DE-71-exposed male offspring indicate broad, complex effects of developmental exposure to PBDEs on glucose homeostasis and glucoregulatory parameters involving various organ systems. Our results support human studies reporting the positive association between body burdens of PBDEs and T2D and MetS  (AUP# 20170026 and 20200018).Conceptualization, MC-C, EK. Methodology, MC-C, EK, ND, JK, BC. Validation, MC-C, EK, BC, PP, ND. Formal Analysis, EK, MC-C, PP, ND. Investigation, EK, BC, PP, JK, AB, MC-C, MD. Writing \u2013 Original Draft, EK, MC-C, AB. Writing \u2013 Review & Editing, MC-C, EK, AB. Visualization, EK. Resources, MC-C., ND. Data Curation, EK, MC-C. Supervision, MC-C, EK, ND. Project Administration, MC-C, EK. Funding Acquisition, MC-C, EK, ND. All authors contributed to the article and approved the submitted version."}
+{"text": "The COVID-19 pandemic has led to a considerable increase in Central-Line Associated Bloodstream Infections (CLABSI) and Catheter Associated Urinary Tract Infections (CAUTI). However, it remains unclear how this affected different population groups.This retrospective observational cohort study included CLABSI and CAUTI in a tertiary care facility. Information was collected on patient demographics, hospitalization, comorbidities, and COVID-19 status. Chi-square and Wilcoxon rank-sum tests were used for categorical and continuous variable comparisons. GEE models compared pre- and pandemic periods by interrupted time series analysis.From 1/1/2018 to 5/31/2022 98,791 patients had 151,550 hospital admissions. Of those, 17,796 patients had 29,483 central lines placed and 45,180 patients had 65,422 Foleys. 314 patients developed 338 CLABSI and 216 patients had 217 CAUTI. 1,552 patients tested positive for COVID-19 with 22 developing CLABSI and 14 CAUTI. The pre-pandemic downward trend in CLABSI and CAUTI was reversed during COVID-19 (p< 0.05).Black patients and those with other/unknown race had higher CLABSI per patient with central lines (p< 0.02) and higher device days (p< 0.0001) compared to white patients. Hispanic/Latino patients acquired more CLABSI per patient compared to non-Hispanic/non-Latino patients (p=0.04). During COVID-19 the already higher device days in Hispanic/Latino patients further increased compared to non-Hispanic/non-Latino patients (p< 0.0001).Black patients had higher CAUTI infection rates per patient with Foleys compared to white and other/unknown patients (p< 0.05). CAUTI per device days were also higher in black patients compared to white patients (p=0.02). No difference in CAUTI burden was detected for ethnic groups throughout the study period. Black and non-Hispanic/non-Latino patients had higher Foley usage days than other patients (p< 0.0001). Foley days decreased during COVID-19 (p=0.01).We detected health outcome disparities affecting black (CLABSI and CAUTI) and Hispanic/Latino (CLABSI) patients. Infections increased during the pandemic without altering race/ethnicity differences. Device utilization was significantly higher in affected groups.All Authors: No reported disclosures"}
+{"text": "In TANGO, switching to dolutegravir/lamivudine (DTG/3TC) was noninferior compared to continuing a tenofovir alafenamide (TAF)-based regimen, however switching from bictegravir (BIC)/emtricitabine (F)/TAF was not evaluated. Here, we present efficacy and safety of switching to DTG/3TC compared with continuing B/F/TAF in virologically suppressed adults through 24 weeks.DYAD (NCT04585737) is an ongoing, open-label clinical trial that randomized adults with HIV-1 RNA< 50 copies/mL and no prior virologic failure (2:1) to switch to once-daily fixed-dose DTG/3TC or remain on B/F/TAF. The primary endpoint is the proportion with HIV-1 RNA \u2265 50 c/mL at Week (W) 48 . A planned W24 interim analysis assessed noninferiority with a 6% margin.Overall, 222 adults  were randomized. At W24, 3 (2%) participants on DTG/3TC and 3 (4%) on B/F/TAF had HIV-1 RNA \u2265 50 c/mL  meeting noninferiority criteria. Through W24, 4 on DTG/3TC and 3 on B/F/TAF met confirmed virologic withdrawal (CVW) criteria and underwent resistance testing. 2/7 had treatment-emergent resistance. One B/F/TAF CVW developed M184M/I and G140G/S at W12; at study discontinuation (DC), HIV-1 RNA< 50 c/mL and the participant remained on B/F/TAF. One DTG/3TC CVW had no resistance on study genotype but underwent repeat genotypic testing outside the study (ordered by clinic provider) and had M184V at W12; at study DC, HIV-1 RNA< 50 c/mL on DTG/3TC and the participant was subsequently transitioned to DTG + darunavir/cobicistat (DRV/c). One non-CVW DTG/3TC participant developed M184V and K65R at W12 (genotype inadvertently collected at first episode of unconfirmed viremia); at study DC, HIV-1 RNA< 50 c/mL on DTG/3TC and the participant was subsequently transitioned to DTG+DRV/c. Drug-related adverse events (AEs) and withdrawals due to AEs occurred in 31 (21%) and 6 (4%) participants with DTG/3TC and 1 (1%) and 0 participants with B/F/TAF, respectively.DYAD demonstrated noninferior efficacy of switching to DTG/3TC vs. continuing B/F/TAF at W24. Higher AE rates in the DTG/3TC arm are likely consistent with the open-label nature of this switch study.Charlotte-Paige M. Rolle, MD, MPH, Gilead: Advisor/Consultant|Gilead: Grant/Research Support|Gilead: Honoraria|Janssen: Advisor/Consultant|ViiV: Advisor/Consultant|ViiV: Grant/Research Support|ViiV: Honoraria Federico Hinestrosa, MD, AbbVie: Honoraria|Gilead Sciences: Advisor/Consultant|Gilead Sciences: Honoraria|MSD: Honoraria|ViiV Healthcare: Advisor/Consultant|ViiV Healthcare: Honoraria Edwin DeJesus, MD, Gilead Sciences, Inc: Advisor/Consultant|Theratechnology: Advisor/Consultant|Theratechnology: Honoraria"}
+{"text": "Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis are frequent etiologic agents of CABP. Ceftaroline fosamil is a parenteral cephem approved treatment of patients with CABP caused by S. pneumoniae (including cases with concurrent bacteremia), methicillin-susceptible Staphylococcus aureus (MSSA), H. influenzae, and some species of Enterobacterales. In this study we report the in vitro activity of ceftaroline and comparators against isolates from community-acquired respiratory tract infections (CARTI) collected through a global surveillance program.Community-acquired bacterial pneumonia (CABP) is a frequent cause of patient morbidity and mortality. S. pneumoniae, H. influenzae, M. catarrhalis, MSSA, and methicillin-resistant S. aureus (MRSA) were tested. The isolates  originated from Asia/South Pacific ; Europe ; Latin America (671/8.5%); Middle East/Africa (659/8.4%); and North America (Canada only) (380/4.8%). Ceftaroline and comparator agent MICs were determined by CLSI M07 broth microdilution methodology. MICs were interpreted using 2023 CLSI M100 MIC breakpoints.Clinically relevant, non-duplicate, isolates cultured from respiratory specimens by clinical laboratories in 54 countries in 2018-2021 were collected by the ATLAS Surveillance Program central laboratory . Community-acquired infections were defined as those from patients < 48 hours in hospital. In total, 7,886 isolates of in vitro activity of ceftaroline and comparator agents is summarized in the table. Greater than 99% of S. pneumoniae and 100% of MSSA were susceptible to ceftaroline, including penicillin-nonsusceptible S. pneumoniae based on a dosage of 600 mg every 12h. Sixty-two (9.6%) MRSA were ceftaroline-susceptible-dose-dependent (MIC 2-4 \u00b5g/mL) based on a dosage of 600 mg every 8h administered over 2h, with the majority from (n) Thailand (9), S. Korea (7), and China (6). Four isolates, from S. Korea (2), China (1), and Ukraine (1) were resistant to CPT (MIC of \u22658 \u00b5g/mL). 99.6% of H. influenzae were susceptible to ceftaroline.The in vitro activity against current pathogens associated with CABP from a global collection.Ceftaroline demonstrated potent Meredith Hackel, PhD, Pfizer Inc.: Honoraria|Venatorx: Paid fees for conducting the study and abstract preparation Gregory Stone, PhD, Pfizer: Stocks/Bonds Daniel F. Sahm, PhD, Merck & Co., Inc.: Honoraria|Pfizer Inc.: Honoraria|Venatorx: Paid fees for conducting the study and abstract preparation"}
+{"text": "Dialysis, renal transplant (RTX) and autoimmune kidney disease (AKD) patients are at higher risk of COVD-19 and suboptimal humoral responses post primary SARS-CoV-2 vaccination. As such, these groups have been prioritised for further SARS-CoV-2 vaccine doses. Limited data are available on their immunogenicity.nd, 3rd and 4th SARS-CoV-2 vaccines. Endpoints were median anti-S IgG titres and seroconversion (anti-S IgG titre >1896 MFI).Patients on dialysis or with a RTX or AKD who received primary SARS-CoV-2 vaccination were prospectively recruited from three UK sites. SARS-CoV-2 IgG spike antibody (anti-S IgG) and nucleocapsid titres were measured with a Luminex assay post 2rd dose , with 27/58 (47%) of RTX and 17/46 (37%) of AKD patient non-responders seroconverting. Seroconversion among RTX and AKD patients overall improved to 78% (114/146) and 79% (122/155) respectively (p< 0.05), but not in dialysis patients post 3rd dose. 212/400 (53%) had serology post 4th dose , with 10/22 (46%) and 3/25 (12%) of RTX and AKD patient non-responders seroconverting. Anti-S IgG titre and seroconversion were similar within each group post 4th dose .628 patients  comprising 240 dialysis, 194 RTX, and 194 AKD patients were recruited, with seroconversion of 96%, 61% and 70% respectively post two vaccines, and median anti-S IgG titres (IQR) of 30604 (24393\u201331826), 5829 (455\u201328296) and 18711 (1233\u201330226). 400/627 (64%) had anti-S IgG titres post 3rd dose in a multivariable model were rituximab use within six months, mycophenolate, prednisolone, chronic kidney disease and AKD , and post 4th dose, rituximab alone (p< 0.001).Predictors of non-seroconversion post 3rd but not 4th dose improved anti-S IgG titres and seroconversion in renal patients. Immunosuppressive use, particularly rituximab, was associated with reduced serologic responses . Patients on immunosuppressives should therefore be prioritised for consideration of additional pre-exposure prophylactic agents.After primary SARS-CoV-2 vaccination, a 3Michael Chen-Xu, MBChB, MRCP, MPH, GSK: Grant/Research Support Rachel Jones, MD, GSK: Advisor/Consultant|GSK: Grant/Research Support|Roche: Grant/Research Support|Vifor Pharma: Advisor/Consultant Rona M. Smith, MD MRCP, GSK: Grant/Research Support|Union Therapeutics: Grant/Research Support"}
+{"text": "Remdesivir (RDV), a nucleotide analog prodrug that targets the viral RNA-dependent RNA polymerase Nsp12, is approved to treat COVID-19 in hospitalized and nonhospitalized patients. Obeldesivir (ODV), an oral mono-5\u2019-isobutyryl ester prodrug that is metabolized into the same active triphosphate as RDV, is being evaluated in Phase 3 clinical trials. The antiviral activity of RDV and ODV against previous Omicron subvariants (BA.1 to BQ.1.1) was maintained with respect to the ancestral WA1 strain. We assessed RDV and ODV antiviral activity against recent Omicron subvariants BF.7, BQ.1, XBB.1.5, CH.1.1, and XBF using clinical isolates and/or site-directed mutants (SDMs).Global Initiative on Sharing Avian Influenza Data (GISAID) EpiCoV database sequences. Structural analysis of identified substitutions was conducted on a prior cryo-electron microscopy-based model of the replication-transcription complex. Antiviral activity of RDV and ODV against subvariant clinical isolates was assessed by nucleoprotein ELISA in A549-hACE2-TMPRSS2 cells and by SDMs in the replicon systemThe prevalence of Nsp12 substitutions in Omicron subvariants was evaluated by analysis of 50 vs WA1). Evaluation of CH.1.1 and XBF by introducing variant-defining mutations into the replicon showed no change in in vitro susceptibility (< 1.8-fold change). Phenotyping of clinical isolates of BF.7, BQ.1, XBB.1.5, and CH.1.1 indicated no loss of RDV or ODV in vitro antiviral activity (< 1.3-fold change).Genomic analysis of > 2 million Omicron subvariant sequences revealed unique substitutions in Nsp12 vs WA1. No new defining substitutions in Nsp12 were found compared with earlier Omicron variants. The defining substitutions (\u2265 75% of sequences) included P323L , Y273H (BQ.1), and G671S . Less prevalent substitutions were observed with frequencies from 1-3.5%: L247F, T248I, V257F, N507I, A529V, F694Y; none had direct interaction with the incoming RDV nucleotide triphosphate or the viral RNA, except N507I. Phenotyping of individual substitutions using SDMs showed no loss of RDV or ODV susceptibility (< 1.2-fold change in ECRDV and ODV retained potent in vitro antiviral activity against all tested Omicron subvariants with potencies comparable to WA1.Lauren Rodriguez, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Jiani Li, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Dong Han, MS, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Ross Martin, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Jasmine Moshiri, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Nadine Peinovich, MPH, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds John P. Bilello, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Jason K. Perry, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Charlotte Hedskog, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds"}
+{"text": "Agrobacterium radiobacter strain MD22b was isolated from infected fruit from Vatan Farm, a dekhkan farm in Yangibog . The 5.7-Mbp draft genome sequence presented here shares homology with chromosomes 1 and 2, as well as with the Ti plasmid from agrobacteria. Agrobacterium radiobacter (Beijerinck and van Delden 1902) Conn 1942 (formerly Agrobacterium tumefaciens) . The fruit was rinsed three times in sterile distilled water and homogenized. Aliquots were spread onto Roy-Sasser agar (Agrobacterium-like colony was picked and identified as A. radiobacter by its API 20 NE (bioM\u00e9rieux) profile (1-4-6-7-7-4-4). DNA was extracted from cells cultivated in LB for 24\u2009h at 30\u00b0C with shaking using the GenElute bacterial genomic DNA kit (Sigma-Aldrich). Sequencing of the 16S rRNA gene using the primer combination 5\u2032-AGRGTTTGATYHTGGCTCAG-3\u2032 (27f_mod) and 5\u2032-TASGGHTACCTTGTTACGACTT-3\u2032 (1492r_mod) (A. radiobacter strain ATCC 23308 (GenBank accession number MT534521.1) using BLASTn v. 2.13.0 searches against the GenBank v. 252 nonredundant nucleotide database (https://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastn&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome). For genome sequencing by Eurofins Genomics, a NEBNext Ultra II DNA preparation kit was used, and Illumina NovaSeq 6000 S2 paired-end genomic sequencing was performed with a read length of 2\u2009\u00d7\u2009151\u2009bp, resulting in 10,125,972 reads with a Phred score of \u226528 and a total of 1,518,897,000 sequenced bases. Additional quality control was performed to remove any remaining adapter sequences using the Trim Reads tool in the CLC Genomics Workbench v. 20.1. Assembly was performed using the CLC de novo assembly tool, resulting in a total sequence length of 5,736,602\u2009bp and a total gapped length of 5,736,406\u2009bp, distributed in 38 contigs, with an N50 value of 267,803\u2009bp, coverage of 265\u00d7, and GC content of 59.44%. Unless otherwise noted, default parameters were used for all software. Annotation of the draft genome was done using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v. 6.1 (https://www.ncbi.nlm.nih.gov/genome/annotation_prok). The genome completeness was estimated at 100% using CheckM v. 1.0.18 (Agrobacterium with pairwise average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values of 97.97% and 83.8%, respectively, against the type strain A. radiobacter NCPPB 3001 using the ANI calculator (http://enve-omics.ce.gatech.edu) (https://ggdc.dsmz.de/ggdc.php#) (A. radiobacter K84 was performed, revealing significant homology with both chromosomes of strain K84 (repA and repB, and the plasmid Ti replication initiator gene, repC. The reported data will be useful for future understanding of the genetic diversity and virulence potential of A. radiobacter in Central Asia.ser agar  plates afile 1-4--7-7-4-4.rain K84 . A complA. radiobacter strain MD22b have been deposited in DDBJ/ENA/GenBank under accession numbers OP364099 and JANDHV000000000, respectively. The associated BioProject, SRA, and BioSample accession numbers are PRJNA856860, SRR21562299, and SAMN29594852, respectively.The partial 16S rRNA gene and whole-genome shotgun sequences of"}
+{"text": "Pantoea agglomerans has been isolated from various habitats including disease plants. Here, we present the genome of P. agglomerans strain NBBC-01 obtained from rotten potatoes that were infected by Ditylenchus desstructor. The genome information will prove advantageous in elucidating its ecological role.The Gram-negative bacterium  Pantoea agglomerans has been shown to promote plant growth  in Wuhan, China, which was infected with a plant-parasitic nematode Ditylenchus destructor  following the manufacturer\u2019s protocols. The DNA purity, concentration, and integrity were assessed with NanoDrop One spectrophotometer , Qubit v.4.0 Fluorometer , and 0.75% agarose gel electrophoresis, respectively. Size selection (20\u201330 kb) was carried out for long-read sequencing utilizing the PippinHT system . The DNA library was generated without DNA shearing utilizing a ligation sequencing kit ), Oxford, UK] and sequenced with a Nanopore PromethION sequencer instrument (ONT) on a R9.4.1 flow cell FLO-MIN106). Guppy v.4.4.2 (. Guppy vP. agglomerans DSM3493T (GenBank accession: GCA_900185875). The values of digital DNA\u2013DNA hybridization (P. agglomerans NBRC 102470 (GenBank accession: GCA_001598475) is 88.7%. Thus, NBEC-018 was identified as a P. agglomerans isolate.The high-quality short- and long-read sequences were assembled into a whole chromosome and plasmids using Unicycler v.0.4.9 . All sofdization , 13 betw"}
+{"text": "There is growing evidence that people with tuberculosis (TB) are at higher risk of mortality even after treatment completion. However, there are limited data on post-TB treatment mortality rates and causes of death in the United States (U.S.).Cohort study of all adults (age \u2265 18 years) diagnosed with TB in Georgia (U.S.) who finished treatment between January 1, 2008, and December 31, 2019. We obtained post-TB treatment mortality data from the Centers for Disease Control and Prevention National Death Index and general Georgia population death rates and causes of death for the same period from public databases. We calculated age- and sex-standardized mortality rates (SMRs) for the TB cohort and general Georgia population and the SMR ratio between these two groups .Among 3,183 patients alive at the time TB treatment was stopped, 2,391 (75.1%) were culture-confirmed, 625 (19.6%) were extrapulmonary, and 15 (< 1%) were rifampin and isoniazid resistant. The median age at TB diagnosis was 44 years (interquartile range (IQR): 32-57), 2,094 (65.8%) were male, and 1,557 (48.9%) were U.S.-born. 328 (10.3%) were co-infected with HIV. Most patients  completed treatment. Reasons for treatment non-completion included adverse events (n=11), patient declined treatment (n=13), and lost to follow-up (n=186). The median post-TB treatment follow-up was 6.4 years (IQR 3.4-9.0) and 233 (7.3%) patients died. Causes of death included cardiovascular disease and malignancy  and HIV . The age and sex adjusted SMR ratio of the TB cohort compared to the general Georgia population was 0.93 (95% CI: 0.78-1.09). The SMR ratio in a subgroup analysis restricted to U.S.-born persons was 1.6 (95% CI: 1.36-1.77).All data is shown as n (%) unless stated otherwise. Abbreviations: TB = tuberculosis, IQR = inter-quartile range 1. Other races = American Indian/Alaska Native , multiracial  2. Other risk factors = end-stage renal disease , post-organ transplantation , TNF-a antagonist therapy , immunosuppression (not HIV/AIDS) , incomplete LTBI , other ; missing data All rates are per 1000, . Abbreviations: TB = tuberculosis 1. Entire TB cohort: deaths n = 233, person-time = 18,440 person-years 2. Georgia population: deaths n = 900,874, person-time = 90,796,252 person-years 3. Race includes only non-Hispanic White and non-Hispanic Black categories: TB cohort (race standardization): deaths n = 212, person-time = 14,095 person-years GA population (race standardization): deaths n = 888,261, person-time = 85,630,800 person-years 4. Culture-confirmed TB cohort: deaths n = 182, person-time = 12,821 person-yearsOur data suggests the overall cohort of patients who finish TB treatment are not at higher risk of death compared to the general population. Analysis restricted to U.S.-born persons, however, did show a slightly higher risk of death post-TB treatment, which suggests the so-called \"healthy immigrant effect\" could have affected our post-TB treatment mortality estimates.All Authors: No reported disclosures"}
+{"text": "A simultaneous, high-throughput and sensitive method for analysing nine neonicotinoid pesticides (NEOs) and four metabolites (NEOms) in urine using a liquid chromatography-tandem mass spectrometry (LC\u2013MSMS) was developed. The method detection limit (MDL) and lowest concentration minimum reporting limit (LCMRL) of the nine NEOs were 0.0013\u20130.048 ng/ml and 0.0050\u20130.17 ng/ml, respectively. The MDL and LCMRL of the four NEOms were 0.0052\u20130.52 ng/ml and 0.011\u20131.6 ng/ml, respectively. Intermediate precision for the nine NEOs and four NEOms was 7.5\u201312.5% and 7.4\u201310.9%, respectively. Accuracy for the nine NEOs and four NEOms was 3.83\u20135.60% and 3.01\u201329.2%, respectively. The developed method was applied to analyse urine samples collected from participants of a large-scale birth cohort study, namely, the Japan Environment and Children's Study (JECS).\u2022The NEO and NEOm concentrations in 100 \u00b5l urine samples were analysed using a highly sensitive LC\u2013MSMS.\u2022Automated solid phase extraction in a 96-well plate was utilised to achieve high-throughput analysis.\u2022Intermediate precision and accuracy were less than 12.5% and 94.8\u201399.1%, respectively.  Specifications tableNeonicotinoid pesticides (NEOs) are agonists of neuronal nicotinic acetylcholine receptors 13C6) (ACE-IS), thiacloprid (pyridymethyl-13C6) (TCP-IS), sulfoxaflor  (SUL-IS), flonicamid  (FLN-IS), thiamethoxam  (THX-IS), dinotefuran (furylmethyl-13C5) (DIN-IS), clothianidin  (CLO-IS), imidacloprid (pyridymethyl-13C6) (IMI-IS), nitenpyram  (NIT-IS), acetamiprid-N-desmethyl  (dm-ACE-IS), TCP-amid (pyridymethyl-13C6) (TCP-amid-IS), CLO-desmethyl  (dm-CLO-IS) and IMD-olefin  (IMI-OF-IS), were purchased from Cambridge Isotope Laboratories, Inc. . Acetonitrile (purity 99.8%), ammonium acetate (guaranteed reagent), formic acid (purity \u226598.0%), methanol (purity 99.8%) and ultrapure water  were used for this analysis.The sample preparation method was based on our previous study TM 6500 mass spectrometer  was used to detect and quantify target analytes. The MSMS system was operated using electrospray ionisation positive mode, and multiple reaction monitoring was performed. An ACQUITY UPLC HSS T3  was used as an analytical column. The LC and MSMS parameters are listed in The Nexera X2 system  was used for separation. A Triple Quad. Sulfoxaflor (SUL) was a diastereomer. The two isomers were quantified separately as SUL-A and SUL-B. All samples that fell outside the calibration range were reanalysed following further dilution.The calibration range is shown in Table\u00a05A pooled urine sample was prepared using maternal samples collected from volunteers to create QC samples. Levels of analytes varied in actual samples, so we tried to create the QC sample to reflect such concentrations. The QC samples had the concentration of dinotefuran around 2 ng/ml. To double the concentration, NEOs and NEOms were fortified such that the concentrations of ACE, TCP, SUL-A, SUL-B, FLN, THX, DIN, CLO, IMI, NIT, dm-ACE, TCP-amid, dm-CLO and IMI-OF were 0.5, 0.5, 0.5, 0.5, 10, 2.0, 5.0, 5.0, 5.0, 5.0, 5.0, 1.0, 20 and 100 ng/ml, respectively. The QC samples were subjected to the same procedure as the unknown samples, with five replicates analysed in each analytical sequence. The lowest concentration minimum reporting limit (LCMRL) was calculated following the U.S. Environmental Protection Agency's instructions 2) higher than 0.990. Repeatability and intermediate precision were determined based on ISO 5725:1994 and 27148:2010, with QC sample measurements (n\u00a0=\u00a0100). QC for day-to-day analysis was determined using a Shewhart control chart  of 1.40 (0.27), 1.36 (0.31), 0.19 (0.06) and 1.34 (0.47) ng/ml for ACE, CLO, TCP and THX, respectively, were analysed in nine replicates. All measured concentrations fell within the certified ranges  samples (NMIJ CRM 7408-a) with certified concentrations  at 25\u00b0C (1.0125 g/cmd ranges .Table 7RNot applicable.Yukiko Nishihama: Conceptualization, Formal analysis, Writing \u2013 original draft, Visualization. Shoji F. Nakayama: Methodology, Software, Writing \u2013 review & editing, Supervision, Project administration. Tomohiko Isobe: Methodology, Investigation, Writing \u2013 review & editing.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper."}
+{"text": "Enterobacteriaceae bloodstream infections (CRE-BI) in children from Latin America and Caribbean (LAC ) is scarce.Systematic reviews about the burden and use of resource related to Carbapenem Resistant To analyze demographic, epidemiological, clinical, use of resource and microbiological data from CRE-BI in LACA systematic review following Cochrane methods, PRISMA/MOOSE statements was performed (PROSPERO 2022 CRD42022352504). We searched all databases  from 1/1/2012 to 9/30/2022. Independent selection, data extraction, and risk of bias assessment (RoB) were performed with COVIDENCE. Types of study included; observational/surveillance/experimental/quasi-experimental. Population: Inpatients < 19 years old, with CRE-BI. Outcomes: age, underlying conditions, previous . treatment and use of resources. Statistical analysis: proportion meta-analyses applying an arc-sine transformation and RoBTo, We conducted proportion meta-analyses and applied arc-sine. R studio was used25/858 screened studies included for full text assessment with 243 patients were included. The most represented countries were Brazil (30%), Colombia (26%) and Argentina (22%). Median age was 34.5 months (IRQ 14-85). Predominant resistance mechanisms: KPC (96%), NDM (58%), OXA (27%). Pool proportion were: Male 55%. Underlying conditions 100% (immunocompromised host 84%), Previous Colonization with CRE was 42% (CI95% 13-78) and broad spectrum antibiotic used was 88% (CI95% 46-98%), most commonly carbapenems 84% (CI95% 72-91%) Bacteremia secondary to intra-abdominal infection was 80%. Combination antimicrobial treatment was 96% (CI95% 22-100%). Median length of stay 22.5 (IQR 22-30 days). Overall case fatality ratio was 48% (IC95% 27-69%).CRE-BI cause morbidity and mortality mainly in pediatric population with underlying disease in LAC region. Considering its high impact in public health, it's mandatory to establish adequate policies and programs to improve antimicrobial stewardship.All Authors: No reported disclosures"}
+{"text": "Fig 1). Stimulation of mucosal antibodies and T cells, immune effectors also associated with protection against influenza, are anticipated to enhance VE.Adults over age 65 years exhibit increased susceptibility to influenza viruses, accounting for 70-85% of annual influenza-related US fatalities. Licensed vaccines mainly induce serum humoral immunity and remain the primary method of influenza prevention, but vaccine effectiveness (VE) remains particularly low in older individuals (Table 1). Demographics were comparable across cohorts (Table 2).A randomized, double-blind, double-dummy, placebo-controlled Phase 1b study (NCT05163847) was conducted at 6 US study sites with the investigational intranasal M2SR (M2 deficient Single Replication) and Fluzone High Dose (HD) vaccine that contained HA & NA from A/Cambodia/e0826360/2020 (Cam2020). Adults aged 65-85 years (n = 305) received either one administration of Cam2020 M2SR alone, Cam2020 M2SR along with HD, HD alone, or placebo . Moreover, M2SR coadministered with HD induced HAI and NAI antibodies in significantly higher proportion of subjects than HD alone . M2SR alone and M2SR coadministered with HD elicited significant mucosal secretory IgA (sIgA) antibodies against vaccine antigen, Cam2020, and drifted H3N2 Darwin2021 strain . Similarly, T cells positive for IFNg and GrzB secretion were significantly induced by M2SR only or by M2SR coadministered with HD . HD alone did not induce mucosal antibodies or T cell responses .Study vaccination was well-tolerated. Intranasal M2SR generated significantly increased serum HAI and NAI antibodies compared with placebo . These broad immune responses support the potential for the safe coadministration of M2SR with HD to enhance protection against influenza infection in this highly vulnerable age group.Coadministration of intranasal M2SR with intramuscular Fluzone HD resulted in a multi-faceted immune response in older adults aged 65-85 years old with a higher proportion of responders across all immune measurements compared with either vaccine alone. Coadministration of the two vaccines resulted in significant rises in serum HAI and NAI titers compared to HD alone (Joseph Eiden, MD, PhD, FluGen, Inc.: Advisor/Consultant Bryan Murray, M.D., FluGen, Inc.: Advisor/Consultant Renee Herber, B.S., FluGen, Inc.: Salary Roger Aitchison, ScM, FluGen, Inc.: Advisor/Consultant David Marshall, B.S., FluGen, Inc.: salary Michael Moser, Ph.D., FluGen, Inc.: salary Pamuk Bilsel, Ph.D., FluGen, Inc.: salary"}
+{"text": "Fusarium verticillioides is a filamentous fungus that causes plant diseases and harms human health through cancer-inducing mycotoxin and life-threatening Fusariosis. Given its threat to agriculture and public health, genome assembly of this fungus is critical to our understanding of its pathobiology and developing antifungal drugs. Here, we report a gap-free genome assembly of F. verticillioides using PacBio HiFi data and high-throughput chromosome capture (Hi-C) sequencing data. The assembled 42.0\u2009Mb sequence contains eleven gapless chromosomes capturing all centromeres and 19 of all 22 telomeres. This assembly represents a significant improvement over previous version on contiguity (contig N50: 4.3\u2009Mb), completeness (BUSCO score: 99.0%) and correctness (QV: 88.8). A total of 15,230 protein-coding genes were predicted, 6.2% of which are newly annotated genes. In addition, we identified three-dimension chromatin structures such as TADs-like structures and chromatin loops based on Hi-C data of ultra-high coverage. This gap-free genome of F. verticillioides is an excellent resource for further panoramic understanding mechanisms of fungal genome evolution, mycotoxin production and pathogenesis on plant and human host. Fusarium verticillioides, a filamentous fungus belonging to Fusarium fujikuroi species complex, causes Fusarium ear rot of maize, a major crop worldwide. Besides yield loss, various mycotoxins are produced during fungal infection of maize, reducing the quality of corn products. The best characterized F. verticillioides mycotoxins are fumonisins, a group of polyketide mycotoxins associated with esophageal cancer and neural tube birth defects in human populations consuming the contaminated maize products1. Although F. veticillioides is considered nonpathogenic to healthy human being, it can become a serious threat to individuals with compromised immune system such as those infected by undergoing organ transplants2. Human infection by F. verticillioides commonly known as Fusariosis has been a surging life threat to the immunocompromised patients due to limited options of antifungal drugs for treatment and emergence of multi-drug resistance3. Therefore, elucidation of molecular mechanisms underlying fungal pathogenesis and antifungal resistance in F. verticillioides is crucial to both agricultural safety and public health.F. verticillioides strain 7600 was released in 20104 with a contig N50 of 392.3\u2009kb. Recently, several updated versions of F. verticillioides genome assemblies are available in NCBI  genome database. Although these genome assemblies have since facilitated the genetic studies of fungal biological processes, they are highly fragmented with several hundreds to thousands of contigs. The fact that F. verticillioides has 11 chromosomes suggests the presence of gaps in these assembly versions. Furthermore, no telomere and centromere sequences have been reported in any F. verticillioides genome assembly available, leaving these essential and complex genomic regions unexplored. A complete genome sequence for F. verticillioides would enable accurate characterization of the fungal genome function, regulation and evolution, shedding light on mechanisms of growth, development, pathogenicity and mycotoxin production.Despite the importance of this fungus, its complete genome sequence has not been assembled and thoroughly analyzed, impeding dissection of molecular and evolutionary mechanisms underlying its pathogenesis, secondary metabolism and drug resistance. The first genome assembly of F. verticillioides, and update the genome annotations based on the improved genome assembly. We sequenced the genome of F. verticillioides strain 7600 to produce high-fidelity (HiFi) long reads of PacBio  single-molecule real-time (SMRT) sequencing, and Hi-C (high throughput chromatin conformation capture) data using Illumina pair-end sequencing. In total, we generated 4.1\u2009Gb (~96.7X coverage) PacBio HiFi raw reads with a N50 of 10.0\u2009kb. and 53.8\u2009Gb Hi-C data   and Flye7 to obtain draft genome assemblies which were individually polished using Nextpolish (v.1.4.0)8 followed by assembly merge using quickmerge (https://github.com/mahulchak/quickmerge)  contains 11 gap-free chromosomes  compared to the previous assembly Table\u00a0. CompariF. verticillioides contained all centromeres on 11 chromosomes  Hi-C data containing 95.8% valid interaction pairs after initial quality control  slant and stored in \u221280\u2009\u00b0C freezer. F. verticillioides 7600 mycelia and spores harvested from two-day old PDB (potato dextrose broth) culture in 150\u2009rpm shaker at 25\u2009\u00b0C were used to isolate high molecular weight DNA using CTAB (cetyltrimethylammonium bromide) method11. A total of 15\u2009\u00b5g purified genomic DNA were used to construct a standard PacBio SMRTbell library using PacBio SMRT Express Template Prep Kit 2.0 . The sequencing was performed using a PacBio Sequel II instrument at Biomarker Technologies Corporation .F. verticillioides was prepared from cross-linked chromatins of fungal mycelia using a standard Hi-C protocol12. The constructed Hi-C sequencing library was sequenced by a test run and examined for valid interaction read pair ratios using HiCPro (v.3.1.0)13 before going through high coverage sequencing. The library was sequenced by Illumina NovaSeq. 6000 to yield 10.5\u2009Gb (249.7 coverage) paired-end reads. The valid interaction pairs of Hi-C sequencing reads were used to anchor all contigs using Juicer (v.1.5)9, followed by using a 3D-DNA correction pipeline10 and manually correction with Juicebox (v.1.11.08)14. compartment A/B were analyzed using HiTC (v.1.40.1)15 and Cworld-dekker (https://github.com/dekkerlab/cworld-dekker), TADs-like structures and chromosome loop were identified by Juicer (v.1.5)9. Three-dimensional structure visualization of the whole genome using pyGenomeTracks (v.3.7)16.Hi-C library construction of 5, HiCanu (v.1.4)6 (parameters: -assemble -pacbio-hifi oeaErrorRate\u2009=\u20090.001), Flye (v.2.9)7 and NextDenovo (v.2.5.0) , to assemble, respectively, and then sorted the number of contigs of different assemblers in ascending order. Based on four assemblies we used quickmerge (v.0.3) (https://github.com/mahulchak/quickmerge) to produce a merged genome assembly, and finally used Juicebox14 to manually adjust misassemblies.To optimize the genome assembly strategy and take into account the differences of assembly algorithm between software, we used Hifiasm (v.0.16.1)F. verticillioides using Trizol  agents following manufacturer recommendation protocol. The RNA Nano 6000 Assay Kit of Agilent Bioanalyzer 2100 system  was used to evaluate the total RNA integrity. The total RNA used for library preparation first enriched the mRNA with polyA tail through Oligo (dT) magnetic beads. The mRNA was then subjected to sequencing library construction using Illumina True-seq transcriptome kit  with an insert size of 370bp\u2013420bp, and sequenced by an Illumina Novaseq. 6000 platform at Biomarker Technologies Corporation  to generate 150\u2009bp paired-end reads. RNA-seq data was checked for quality using fastp (v.0.23.2)17, mapped to F. verticillioides genome assembly using hisat2 (v.2.1.0)18, followed by calculating mapping ratios by samtools (v.1.15)19.Total RNA was extracted from the mycelia of de novo prediction and similarity aligmment to annotate it via RepeatModeler (v. 1.0.11)20 (parameters: -database -engine ncbi -pa) and softmasked genome by RepeatMasker (v. 4.1.2.p1)21. RepeatMasker\u2019s perl script (rmOutToGFF3.pl) converts various types of repeat sequence annotation results into a common generic feature format (GFF) version 3. Gene model prediction combined with the following three aspects of evidence: (a) ab initio prediction, (b) homologous protein, (c)RNA-seq evidence. During the ab initio prediction, we firstly trained the GeneMark-ET model for five rounds using BRAKER2 (v.2.1.6)22 (parameters:\u2013species\u2009=\u2009Fv\u2013fungus\u2013softmasking\u2013genome\u2013bam\u2013prot_seq\u2013prg\u2009=\u2009gth\u2013gff3\u2013rounds\u2009=\u20095), whose process employed GeneMark-ET23, NCBI BLAST24, DIAMOND25 and GenomeThreader26. We then trained the SNAP27 semi-HMM model for two rounds using MAKER28 . AUGUSTUS29 used the built-in Fusarium genome feature model. To provide homologous protein evidences for gene prediction, we downloaded the protein data of this species (anchored chromosomes) from the public database, including 7600 (NCBI Assembly ID: GCA_ 000149555.1), BRIP53590 (NCBI Assembly ID: GCA_003316995.2), BRIP53590 (NCBI Assembly ID: GCA_003317015.2) and BRIP14953 (NCBI Assembly ID: GCA_003316975.2). For transcriptome data, RNA-seq reads from the vegetative phase were firstly aligned to our genome assembly through hisat218 for BRAKER2 (v.2.1.6)22. Then, we performed reference-based assembly and de novo assembly of transcriptomes by Scallop (v0.10.5)30 and Trinity (v.2.8.4)31 , respectively. Transcripts obtained by two methods are de-redundant with CD-HIT (v.4.6)32 (parameters:-I -c 0.99 -T 50 -M 100000 -o). The above three evidences are integrated by MAKER (v.3.01.03)28 to predict the final gene model. Rfam/Infernal (v.1.1.4)33 (parameters: cmscan\u2013cut_ga\u2013rfam\u2013nohmmonly\u2013fmt 2 \u2013clanin \u2013tblout) and tRNAscan-SE (v. 2.0.9)34 (parameters: -E -X 20 \u2013f -m -b -j\u2013detail) are used to infer genome-wide non-coding RNAs. To compare the previous (NCBI: GCA_000149555.1) and our genome assembly, we performed the genome annotation on two genome assemblies by using the same software, parameters, and protein data.For repetitive sequences, we firstly use 35 with accession number of SRR2100352136, SRR2100352037, SRR2100351938, respectively. The gap-free genome assembly is deposited under the same BioProject at NCBI (GCA_027571605.1) and also in Genome Warehouse of National Genomics Data Center (https://ngdc.cncb.ac.cn/) at China National Center for Bioinformation under the accession number of GWHBQEB0000000039. Genome annotations including protein-coding regions, repeat sequence and ncRNA annotation files have been submitted to the online open access repository Figshare40.The raw PacBio HiFi sequencing data, Hi-C data and RNA-seq data have been deposited in the National Center for Biotechnology Information (NCBI) under the BioProject (PRJNA868307)14. We then aligned the species\u2019 mitochondrial genome to our assembly by megaBLAST24, which found no errors. Finally, we also used megaBLAST24 to aligned our genome assembly against a common database (ftp://ftp.ncbi.nlm.nih.gov/pub/kitts/contam_in_euks.fa.gz) to identify potentially contaminated sequences sequencing adaptor sequence (ftp://ftp.ncbi.nlm.nih.gov/pub/kitts/adaptors_for_screening_euks.fa) and nucleotide sequence database (remote mode), which again found no contamination.To get a nearly complete and error-free nuclear genome, we first manually corrected the assembly using Hi-C read alignment within the Juicebox41 (parameters: -W repetitive_k15.txt -t 104 -ax map-pb) and the quality value (QV) was assessed using Merqury (v.1.3)42 (parameters: k\u2009=\u200918 count). Second, the BUSCO 43 analysis was conducted to reflect the completeness of genome assembly. The final F. verticillioies gap-free genome assembly has a QV of 88.8, completeness of 99.7% and BUSCO score of 99%, suggesting the high accuracy and completeness of the assembly, respectively 44. This assembly has reduced the length of gaps from 90,816 in previous version to 0, and captured eleven centromeres and nineteen telomeres (TTAGGG) except missing three telomeres via trf (v. 4.09.1)45 (parameters: 2 7 7 80 10 90 2000 -d -m -l 2) from assemblies and raw sequences, one each at the end of Chr2 and Chr4 47 (parameters: intersect -wa -wb -f 1.0), which account for 99.5% of the old version genome and 93.6% of this study genome. Compared to previous version (NCBI: GCA_000149555.1), our assembly has corrected three major inversions (Fig.\u00a048 plot.The resolved fungal telomere and centromere regions have been well covered by PacBio HiFi reads that span these complex regions Fig.\u00a0 by IGV (hr4 Fig.\u00a0. There iools v.2.0.047 (paSupplementary Table 1"}
+{"text": "Correction:\u00a0BMC Public Health\u00a023, 960 (2023)https://doi.org/10.1186/s12889-023-15934-yFollowing publication of the original article , the autFig. 2 correct version of Fig. 1"}
+{"text": "Little is known about the first line induction chemotherapy cycles for HIV-associated diffuse large B-cell lymphoma (DLBCL) as these are less common than HIV-negative lymphoma. Currently, the optimal treatment cycles option remains undefined. Therefore, we performed a multi-center study to analyze the clinical characteristics and outcomes of HIV-associated DLBCL patients in different treatment modes in China.Totally 273 newly diagnosed HIV-associated DLBCL patients at eleven large academic centers from October 2008 to October 2021, were analyzed.p<0.001). Cox multivariate analysis showed that age \u226560 , high IPI score (3-5) , B symptoms , elevated LDH  and received less than 4 cycles chemotherapy  were independent risk factor for adverse prognosis based on overall survival (OS).In the entire cohort, the median age was 47 years  at lymphoma diagnosis, and 223 patients were male (81.7%). One hundred and ninety-four (71.1%) patients were germinal center B-cell-like lymphoma (GCB) subtype. Most patients  had elevated lactate dehydrogenase (LDH), and advanced Ann Arbor stage (78.9% 213/273) at diagnosis. High international prognostic index (IPI) score (3-5) at diagnosis was found in 65.2% (178/273) of patients. One hundred and fifty-five patients (56.8%) had extranodal involvement. The median CD4 cell count was 168/\u03bcl , of whom 174 (63.7%) had a CD4 cell count below 200/\u03bcl. The median follow\u2010up of our cohort was 10.1 (0.1-160) months. The overall 2-year OS rates 58.0%. Median OS times in the 0, 1-3, 4-6, and >6 cycles chemotherapy cohort were 7.1 months, 20.0 months, not reached, and not reached, respectively (Hazard Ratio (HR)=0.549, 95% Confidence interval (CI) 0.451-0.667; These results demonstrated that 4-6 cycles chemotherapy were significantly associated with improved outcomes in HIV-associated DLBCL patients. However, >6 cycles chemotherapy did not further improve the survival of patients. Human immunodeficiency virus (HIV)-associated lymphoma is a rare aggressive non-Hodgkin lymphoma. The incidence rate reported in 2017 was 100/100, 000 to 300/100,000 in population infected with HIV, becoming the highest incidence rate among HIV-associated cancers in the United States , regardless of chemotherapy regimen or dose intensity for HIV-associated DLBCL patients. This is because patients are easily complicated by opportunistic infections, resulting in an increase in mortality , 5. Withp<0.001)  received no anti-lymphoma chemotherapy because of fear of discrimination and poor financial situation. Median survival times in the patients who no received any anti-lymphoma chemotherapy was only 3.5 months, which was significantly lower than that of patients receiving chemotherapy  classification criteria.cART included two nucleoside reverse transcriptase inhibitors and one nonnucleoside reverse transcriptase inhibitor. The chemotherapy cohort included patients who received any of the following regimens: etoposide, doxorubicin, vincristine, cyclophosphamide and prednisone (EPOCH); rituximab, etoposide, doxorubicin, vincristine, cyclophosphamide and prednisone (R-EPOCH).18F-fluorodexyglucose positron emission tomography/computed tomography (PET/CT) or computed tomography (CT) were performed for radiological evaluation. Responses were classified according to standard criteria per each institution, typically aligning with Lugano criteria, which including complete response (CR), partial response (PR), stable disease (SD) and progressive disease (PD).Progression-free survival (PFS) was defined as the time from lymphoma diagnosis to disease progression, relapse or death from any cause. Overall survival (OS) was defined as the time from lymphoma diagnosis to last follow-up or death from any cause. Survival was estimated using Kaplan\u2013Meier curves and compared by the log-rank test. The Cox proportional hazards regression model was used in multivariate analysis to determine prognostic factors. All statistical tests were two\u2010sided, and p \u2264 0.05 was considered statistically significant. All statistical data were analyzed with GraphPad Prism 9.2-microglobulin (\u03b22-MG), and advanced Ann Arbor stage (78.9% 213/273) at diagnosis. High international prognostic index (IPI) score (3-5) at diagnosis was found in 65.2% (178/273) of patients. One hundred and fifty-five patients (56.8%) had extranodal involvement. Totally 53(19.4%) patients had bone marrow involvement, 24(8.8%) patients had central nervous system (CNS) involvement, 64 (23.4%) had poor Eastern Cooperative Oncology Group performance status (ECOG PS 2-4), and 150 (54.9%) had B symptoms at diagnosis. The median CD4 cell count at diagnosis was 168/\u03bcl , of whom 174 (63.7%) had a CD4 cell count below 200/\u03bcl at diagnosis. Among the Epstein-Barr virus (EBV) status, EBV load was elevated (5\u00d7103 copies/ml) in 104 (38.1%) patients. Of all patients, 37(13.6%) had positive HBsAg and 7(2.6%) were positive for anti-Hepatitis C virus (HCV) antibody.Two hundred and seventy-three newly diagnosed HIV-associated DLBCL were included in this study. The median age was 47 years  at lymphoma diagnosis, and 223 patients were male (81.7%). One hundred and ninety-four (71.1%) patients were germinal center B-cell-like lymphoma (GCB) subtype. Most patients  had elevated lactate dehydrogenase (LDH),  elevated serum \u03b22-MG, Ann Arbor stage, IPI score, extra-nodal involvement, bone marrow involvement, CNS involvement, CD4 cell count, and chemotherapy regimen. The 0 cycle chemotherapy cohort featured more cases of GCB . Baseline clinical characteristics are summarized in Patients were stratified based on the induction chemotherapy, where 38/273 (13.9%), 71/273 (26.0%), 73/273 (26.7%), and 91/273 (33.3%) were in the 0, 1-3, 4-6, and >6 cycles chemotherapy cohort, respectively. In the study, some patients received no anti-lymphoma therapy because of fear of discrimination and poor financial situation, some patients had less than four cycles of anti-lymphoma therapy also due to poor financial situation. This is because Central and Western China AIDS Lymphoma League (CALL) is located in southwest China, and the economic level of the patients is low. Patient characteristics were similar between the four cohorts, including gender, age, residence, educational level, HIV transmission route, cART therapy, years of HIV infection before lymphoma, ECOG-PS, the presence of B symptoms, elevated serum LDH, elevated serum \u03b2.In this cohort study, the median chemotherapy cycles were 4 , 38 (13.9%) received no anti-lymphoma therapy, 71 (26.0%) had less than four cycles of chemotherapy, 73 (26.7%) patients received four to six cycles of chemotherapy, and 91 (33.3%) patients received more than six cycles of chemotherapy. Of the 235 patients who received chemotherapy, 80 (34.0%) received EPOCH regimen and 155 (66.0%) received R-EPOCH regimen. 239 (87.5%) were administered cART. Totally 209 patients were evaluated for best treatment response at the end of treatment, including 65 patients who received EPOCH and 144 patients who received R-EPOCH. The results indicated an overall response rate (ORR) of 63.1% and 80.6%, respectively.CR rate were 23.1% and 42.4%, respectively  . Median PFS times in the 0, 1-3, 4-6, and >6 cycles chemotherapy cohort were 7.1 months, 10.1 months, 36.5 months, and 26.4 months, respectively (Hazard Ratio (HR)=0.713, 95% Confidence interval (CI) 0.600-0.847; p<0.001) . Median OS times were 7.1 months, 20.0 months, not reached, and not reached, respectively  . These results demonstrated that 4-6 cycles chemotherapy were significantly associated with improved outcomes in HIV-associated DLBCL patients. However, >6 cycles chemotherapy did not further improve the survival of patients.The median follow\u2010up of our cohort was 10.1 (0.1-160) months. Median PFS and OS were 17 months and 38.1 months respectively. The overall 2-year PFS and OS rates were 46.8% and 58.0%, respectively  . Median . Medianp=0.049) . Median OS times were 27.7 months and not reached. The overall 2-year OS rates were 52.6% and 68.5%, respectively (p=0.048) . These data suggested that rituximab administration was significantly associated with improved outcomes.Median PFS times in the no received rituximab and received rituximab groups were 16 months and not reached. The overall 2-year PFS rates were 41.0% and 53.0%, respectively (p=0.049) . Median p=0.006), high IPI score (3-5) , B symptoms , poor ECOG-PS , elevated LDH , Extra-nodal involvement  and received less than 4 cycles chemotherapy  were predictive of worse PFS. Age \u226560 , high IPI score (3-5) , B symptoms , elevated LDH  and received less than 4 cycles chemotherapy  were predictive of worse OS.Univariate analysis showed that advanced stage , B symptoms  and received less than 4 cycles chemotherapy  were independent risk factors for adverse prognosis based on PFS. Age \u226560 , high IPI score (3-5) , B symptoms , elevated LDH  and received less than 4 cycles chemotherapy  were independent risk factor for adverse prognosis based on OS .Cox multivariate analysis showed that high IPI score (3-5)  in May 2021. The CALL is a public welfare organization dedicated to the diagnosis and treatment of HIV-associated lymphoma patients. This is the first multi\u2010center retrospective study with HIV-associated DLBCL in eleven China academic centers from CALL from October 2008 to October 2021. The present study included 273 HIV-associated DLBCL cases, which is the largest cohort reported in China to date. To the best of our knowledge, this is also the first systematic study to explore the effect of chemotherapy cycles on prognosis in a large series of HIV-associated DLBCL.HIV-associated malignancies are divided into acquired immune deficiency syndrome-defining and non-acquired immune deficiency syndrome-defining cancers based on the coincidence rate. Kaposi\u2019s sarcoma, Non-Hodgkin\u2019s lymphoma and invasive cervical cancer are considered acquired immune deficiency syndrome- defining cancers. Other cancers, which including Hodgkin\u2019s lymphoma, hepatocellular carcinoma, oral and pharyngeal cancers, lung cancer, anal cancer, vulvar cancer and penile cancer, are considered non-acquired immune deficiency syndrome-defining cancers , 2. The The present cohort study showed that the median age was 47 years  at lymphoma diagnosis, and 223 patients were male (81.7%), in agreement with the overall male-to-female HIV incidence rate in China . CompareCurrently, there are no standard guidelines for treatment of HIV-associated DLBCL. In the pre-cART era, median survival times only 5 to 8 months for HIV-associated DLBCL patients. This is because patients are easily complicated by opportunistic infections, resulting in an increase in mortality , 5. In tp=0.03) and PFS  compared with HIV-negative patients (p=0.006) and median OS  compared with no receiving rituximab (p=0.049). The overall 2-year OS rates were 52.6% and 68.5%, respectively (p=0.048). The current data also suggested that rituximab administration was significantly associated with improved outcomes in HIV-associated DLBCL. In AMC034, rituximab combined with EPOCH was assessed, and the results were compared with R-CHOP in AMC010. The results showed that a higher CR rate (69%) was obtained for R-EPOCH compared with R-CHOP (47%)  . In the % . In thOP (47%) .p<0.001), and CR rate  were significantly higher in patients who received chemotherapy (p<0.001) were independent risk factor for adverse prognosis.The optimal treatment cycles option of first line induction chemotherapy for HIV-associated DLBCL remains undefined. Compared with HIV-negative DLBCL patients, HIV-associated DLBCL patients are difficult to accept long-term hospitalization chemotherapy, whether there is an appropriate reduction in the number of chemotherapy cycles, the survival of patients will not be affected? Wu D retrospectively analyzed the clinical features of 100 HIV-associated lymphoma patients, which including 66 HIV-associated DLBCL. 15 patients (15%) did not received chemotherapy. Compared to those who did not received chemotherapy, the 2-year OS rate (41.0% vs. 0%, otherapy . In our otherapy . Once thotherapy , 20\u201324. In summary, our multi-center study suggested that received less than 4 cycles chemotherapy predicted poor OS and PFS in HIV-associated DLBCL patients. Therefore, we recommend that patients receive at least 4 cycles of systemic chemotherapy. More high-quality randomized controlled studies were needed to test our findings.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Institutional Review Board. The patients/participants provided their written informed consent to participate in this study.CW, YW, JL, and HM conceived and designed the study, analyzed the data, and drafted and revised the paper. HZ, YC, and YL conceptualized and designed the study. All authors provided critical comments to the manuscript. All authors contributed to the article and approved the submitted version."}
+{"text": "Streptomyces; however, recent publications have reported terpene biosynthesis by members of the genera Actinomadura, Allokutzneria, Amycolatopsis, Kitasatosporia, Micromonospora, Nocardiopsis, Salinispora, Verrucosispora, etc. It should be noted that the use of genetically modified actinomycetes is an effective tool for studying and regulating terpenes, as well as increasing productivity of terpene biosynthesis in comparison with native producers. The review includes research articles on terpene biosynthesis by Actinomycetes between 2000 and 2022, and a patent analysis in this area shows current trends and actual research directions in this field.Terpenes and their derivatives  constitute the largest class of natural compounds, which have valuable biological activities and are promising therapeutic agents. The present review assesses the biosynthetic capabilities of actinomycetes to produce various terpene derivatives; reports the main methodological approaches to searching for new terpenes and their derivatives; identifies the most active terpene producers among actinomycetes; and describes the chemical diversity and biological properties of the obtained compounds. Among terpene derivatives isolated from actinomycetes, compounds with pronounced antifungal, antiviral, antitumor, anti-inflammatory, and other effects were determined. Actinomycete-produced terpenoids and meroterpenoids with high antimicrobial activity are of interest as a source of novel antibiotics effective against drug-resistant pathogenic bacteria. Most of the discovered terpene derivatives are produced by the genus  O-containing derivatives (terpenoids) are the largest  and structurally most diverse group of secondary metabolites derived from natural sources. Based on the number of isoprene units, terpene derivatives are classified into mono- (C10), sesqui- (C15), di- (C20), sester- (C25), tri- (C30), sesquar- (C35), and tetra- (C40) terpenes. Terpene derivatives are widely used in the food, cosmetics, and fragrance industries -1-epi-cubenol, firstly obtained by fermentation, was synthesized carbonyl and 2-[(2-chlorophenyl)amino]-2-oxo-acetyl, respectively, were identified among the metabolites of Streptomyces sp. SN194 [308 R=H309 R=CH3310 R1=R2=R3=H311 R1=R2=H, R3=CH3312 R1=H, R2=Cl, R3=H313 R1=OH, R2=R3=H314 R=H315 R=CH2OH316 R=CH(OH)2317 R=CHO318 R=COOH319320321 R=CH3322 R=H323 R1=H, R2=\u03b1OH, R3=H324 R1=H, R2=\u03b1OH, R3=CH3325 R1=H, R2=OCH3, R3=H326 R=CH3327 R=CH2OH328 R=H 329 R=CH3330 dixiamycin B331332333334335336337338339340341 R1=R2=H, R3=CH3, R4=H342 R1=R2=H, R3=CH2OH, R4=H 343 R1=R2=H, R3=R4=CH3344 R1=H, R2=OH, R3=CH3, R4=H345 R1=OH, R2=H, R3=CH3, R4=H346 KO-3988 , S. gris CB00830 , and Strp. SN194  synthesip. SN194 .308 R=H347 R= 348 R=349 R=351 R=NH2350 R=352 R=353 R=Streptomyces sp. T\u00fc6071 produced phenalinolactones A\u2013D (354\u2013357), tricyclic terpene glycosides, and their derivatives 359\u2013362, 365, and 366 [Streptomyces sp. T\u00fc6071 with inactivated oxygenase genes , dehydrogenase genes  and putative acetyltransferase gene (plaV) yielded phenalinolactone derivatives PL HS2 (364), PL X1 (363) PL HS6 (367), and PL HS7 (368) [354) and D (357) were identified as PL IM1 (370) and PL IM2 (369), respectively [S. coelicolor M512 resulted in the formation of the non-glycosylated derivative phenalinolactone E (358) [ and 366 ,237. TheS7 (368) . Later, ectively . Heterol E (358) . 371\u2013381), close structural analogues of phenalinolactones, were discovered by genome mining of diterpene synthases in Streptomyces sp. CB03234 and Streptomyces sp. CB03238. Tiancilactones are characterized by a highly functionalized diterpene backbone, which comprises chloroanthranilate and \u03b3-butyrolactone moieties, and exhibit antibacterial activity [\u03b3-lactone, namely trinulactones A (382) and B (383), were isolated from Streptomyces sp. S006 [1R2R3R4R354OH3-CO-CH3CH355OH3-CO-CHH356OH3-CO-CH2-O-CH3-CH357H3-CO-CH3CH358H3-CO-CHOH3CH359H3-CO-CH3CH360HHH3CH361H3-CO-CHH3CH362HHOH3CH363HH3CH364H3-CO-CH2-O-CH3-CH367 R=\u03b1OH368 R=O365 R=\u03b1OH366 R=O369 R370371 R1=Cl, R2=CH3, R3=OCH3372 R1=H, R2=CH3, R3=OCH3373 R1=Cl, R2=CH3, R3=OH374 R1=Cl, R2=CH3, R3=oxo375 R1=Cl, R2=H, R3=OCH3376377378379 R=H380 R=CH3381 R=H382 R=OH383Tiancilactones A\u2013K (activity . Two newsp. S006 .R1R2R3384), its isomer 385, and fusicomycin B (386) were separated from the fermentation broth of S. violascens YIM 100212 [387) and B (388) were obtained from Streptomyces sp. KCB17JA11 [389) is new meroterpenoid derived from Streptomyces sp. CNQ-027 consisting of an unprecedented dihydronaphthalenone polyketide linked to a bicyclic diterpenoid [384 R=385 R=386 R=387388389Fusicomycin A (M 100212 . Two newCB17JA11 . Actinorerpenoid .384 R=S. platensis MA7327 and S. platensis MA7339 were shown to synthesize platensimycin (390) and platencin (391), representatives of a new class of broad-spectrum antibiotics against Gram-positive bacteria, in particular S. aureus [ent-kaurene and ent-atiserene synthases in biosynthesis of 390 and 391, representing a new biosynthetic pathway for diterpenoids [ent-copalyl diphosphate synthase involved in the biosynthesis of 390 and 391 in S. platensis CB00739, was described. PtmT2 catalyzed the cyclization of GGPP to ent-CPP, which subsequently channeled into (16R)-ent-kauran-16-ol (392) or ent-atiserene (393) by two distinct type  diterpene synthases specific for biosynthesis of 390 or 391, respectively [S. platensis SB12002 and SB12600 produced 390 and 391 with yields of 323 \u00b1 29 mg/L and 255 \u00b1 30 mg/L, respectively, hundreds of times greater than those of wild-type strains [S. platensis SB12600, in addition to 391, accumulated eight new congeners, platencins A2\u2013A9 (394\u2013402) [390 using the mixed culture of S. hygroscopicus HOK021 (NITE P-02560) and Tsukamurella pulmonis TP-B0596 was patented (JP2019149945). Exemplified by 390 and 391, a method of searching for novel natural compounds based on the analysis of biosynthetic genes was proposed (WO2015200501). Data on the biosynthesis features and biological activity of natural and synthetic analogues of platensimycin and platencin were summarized in the reviews [. aureus ,246. Furrpenoids ,248,249.ectively . The met strains ,252 (US2394\u2013402) . A metho reviews ,255.N-containing aminobacteriohopanetriol (403), and adenosylhopane (405), as well as bacteriohopanetetrol (404) and ribosylhopane (406), were determined. Orf14 and orf18 of S. coelicolor A(3)2 responsible for the synthesis of 403 were identified [The intermediates of hopanoids biosynthesis, entified . Streptomyces sp. YIM 56130, triterpene glycoside soyasaponin I (407) [408) produced by S. argenteolus A-2 (FERM BP2065) has a unique structure consisting of a tetraterpene skeleton with 2-O-methylglucuronic acid and O-succinyl benzoate moieties [390391392393394 R1=OH, R2=H395 R1=H, R2=OH396 R1= R2=H397 R1=OH, R2=H398 R1=H, R2=OH399 R=OH400401 R=SCH3402 R=OCH3403 R=NH2404 R=OH405406407408Among the secondary metabolites of  I (407)  with a w I (407)  was obtamoieties .3903913Nocardiopsis, Amycolatopsis, Isoptericola, Saccharopolyspora, Salinispora, Kitasatosporia, Verrucosispora, etc. The compounds produced are represented mainly by sesqui- and diterpenes and their derivatives.Although most of the found actinomycete terpene derivatives are synthesized by streptomycetes, there is an increasing number of publications on terpene biosynthesis by representatives of the genera Nocardiopsis chromogenes YIM 90109, two new monocyclic germacradiene-type sesquiterpenoids germacradiene-9\u03b2,11-diol (409) and 11-hydroxy-germacradien-2-one  were identified along with the known geosmin-type sesquiterpenoid 46 [Kitasatospora setae KM-6054 [Micromonospora marina DSM 45555 [411) and (\u2212)-germacrene A (27), respectively. The ability to produce bicyclic 2-methylisoborneol (6) and geosmin (22) was described for Nocardia cummidelens and N. fluminea [S. avermitilis carrying the genes from Saccharopolyspora erythraea NRRL2338 yielded 2-methylisoborneol (6), while Micromonospora olivasterospora KY11048 synthesized 2-methyleneornane (412) [409410411412Among the secondary metabolites of enoid 46 . The TSs KM-6054  and MicrSM 45555  catalyzefluminea . The trane (412) .4094104413 and 414) were isolated from the culture medium of Amycolatopsis alba DSM 44262 [Isoptericola chiayiensis BCRC 16888, a new sesquiterpenoid isopterchiayione (415) was registered [Amycolatopsis benzoatilytica DSM 43387 catalyzing the formation of a bicyclic sesquiterpenoid (416) was described [Verrucosispora gifhornensis YM28-088 [Verrucosispora sp. FIM06031 produced bicyclic sesquiterpenoid cyperusol C (417) and FW03104 (418) (CN101898936), respectively. Two new monocyclic sesquiterpenoids (SM 44262 . Among tescribed . VerrucoYM28-088  and VerrStreptosporangium roseum DSM 43021 and Kitasatosporia setae KM-6054 afforded tricyclic sesquiterpenoids epi-cubebol (419) [420) and B (421) [Saccharothrix espanaensis DSM 44229 [E)-\u03b2-caryophyllene (93).413414415416417418419420421Terpene synthases from ol (419)  and new  B (421) ,266, resSM 44229  was incuMicromonospora marina DSM 45555 was functionally characterized to produce micromonocyclol (422), a new diterpene alcohol with a rare 15-membered ring [Mycobacterium tuberculosis H37Rv\u043d synthesized unique bicyclic diterpenoids, which presumably block the formation of phagolysosomes in human macrophages. The Rv3377c and Rv3378c genes proved to be responsible for synthesis of tuberculosinol (5(6),13(14)-halimadiene-15-ol, 423), 13R- (424) and 13S-isotuberculosinol (5(6),14(15)-halimadiene-13-ol, 425), and nosyberkol (426) (previously identified as edaxadiene). The analogs of Rv3377c and Rv3378c were found in the virulent strains of M. tuberculosis CDC1551 and M. bovis subsp. bovis AF2122/97, but did not occur in non-pathogenic strains [The TS from red ring . Mycobac strains ,271,272. strains .427) was firstly separated from the fermentation broth of Kitasatosporia griseola MF730-N6 (syn. Streptomyces griseolosporeus MF730-N6) in 1985 [ORF11 and ORF12 in S. lividans together with the GGDP synthase gene resulted in the formation of a new cyclic diterpene ent-clerod-3,13(16),14-triene  with a structure similar to 427 [429) to 428, accepted labdane-type diterpene diphosphates (+)-CDP (430), syn-CDP (431), (\u2212)-ent-CDP (432), as well as halimane-type diterpene diphosphate  and catalyzed the formation of corresponding derivatives (434\u2013437) [A bicyclic terpenoid terpentecin  .terp1 operon from Salinispora arenicola CNS-205 in E. coli led to the generation of isopimara-8,15-dien-19-ol (438). It should be noted that this terpenoid was not observed in pure cultures of S. arenicola CNS-205. Apparently, the terp1 operon was expressed under certain conditions, for example, in the presence of other marine organisms [Salinispora sp. PKU-MA00418 accepted CPP and syn-CPP and produced syn-isopimaradiene/pimaradiene analogues . Compound 439 possess a unique and previously unreported 6-6-7 ring skeleton [447) and B (448), were isolated from the culture medium of Verrucosispora gifhornenensis YM28-088 [Micromonospora haikouensis G039 [Microbispora hainanensis CSR-4 [449) and 2\u03b1-hydroxy-8(14),15-pimaradien-17,18-dioic acid (450) were identified, respectively.Heterologous expression of the biosynthetic rganisms . The terskeleton . New hydYM28-088 . Among ssis G039  and Micris CSR-4 , new ditActinomadura sp. SpB081030SC-15 [Actinomadura sp. KC 191 [451) and actinomadurol (452), rare bacterial C-19 norditerpenoids. A norditerpenoid k4610422 (453), originally discovered from a mesophilic rare actinomycete of the genus Streptosporangium, was isolated from the culture extract of a thermophilic actinomycete Actinomadura sp. AMW41E2 [422423424425426427428429430 R=\u03b1H431 R=\u03b2H432433434435436437438439440441442443444445446447448449450451452453030SC-15  and Acti. KC 191  synthesi AMW41E2 .4224234Catenulispora acidiphila DSM 44928 and Saccharopolyspora spinosa NRRL 18395 produced new di- and tricyclic catenul-14-en-6-ol (454), isocatenula-2,14-diene (455), isocatenula-2(6),14-diene (456) [457), B (458), and 2,7,18-dolabellatriene (459) [Diterpene synthases from ne (456) , and spine (459) , respectNocardia testacea NBRC 100365 and N. rhamnosiphila NBRC 108938 accepted GGPP, but not GPP, FPP, or GFPP as a substrate, which was converted by both enzymes in a tetracyclic diterpene phomopsene (460) [Allokutzneria albata DSM 44149 encoded four diterpene synthases that catalyze the formation of mono-, tri-, and tetracyclic compounds: new spiroalbatene (461), bonnadiene (462) and allokutznerene (463), and known compounds: cembrene A (164), thunbergol (464), phomopsene (460), and spiroviolene (200) [Terpene synthases isolated from ne (460) . Allokutne (200) ,288.465\u2013482) were found in the genus Frankia [483) was isolated from the cell walls of nonpathogenic mycobacteria [454455456457458459460461462463464Hopanoid lipids  (CN101921721). Saccharomonospora sp. CNQ-490 produced saccharoquinoline (485), meroterpenoid with drimane-type sesquiterpene unit [486) and B (487), were derived from Micromonospora sp. WMMC-218, a symbiont of marine ascidians Symplegma brakenhielmi [Mycobacterium tuberculosis H37Rv\u043d revealed that this enzyme catalyzed the formation of 1-tuberculosinyladenosine (488) and its two isomers, one of which was identified as 6N-tuberculosinyladenosine (489). Compounds 488 and 489 are specific diterpene nucleosides of pathogen of Mycobacterium tuberculosis and can serve as chemical markers of infection [Rv3377c-Rv3378c from M. tuberculosis H37Rv\u043d in M. kansasii led to the production of 1-tuberculosinyladenosine (488) [ene unit . Two newenhielmi . Furthernfection ,296,297.ne (488) .Nocardia brasiliensis IFM 0406 (now N. terpenica) to synthesize diterpene glycoside brasilicardin A (490) was first described in 1999 [490) displays a unique structure consisting of a diterpene skeleton with L-rhamnose, N-acetylglucosamine, amino acid, and 3-hydroxybenzoate components [N. terpenica IFM0406 and identified as brasilicardins B\u2013D (491\u2013493) [bra0-12), responsible for the synthesis of 490, in Amycolatopsis japonicum (A. japonicum::bcaAB01) led to the formation of four brasilicardin congeners, namely BraC (492), BraD (493), BraC-agl , and BraD-agl  [S. griseus::bcaAB01 (pRHAMO) transformant containing the biosynthetic cluster of brasilicardin A and a plasmid with a biosynthetic cassette for the generation of TDP-L-rhamnose resulted in increased yields of compounds 492 (1669 mg/L), 495 (926 mg/L), and a new metabolite (496) (15 mg/L). The target 490 was obtained through a five-step chemical modification of 494 [The ability of  in 1999 . Brasilimponents . Later, 491\u2013493) . The hetaF, 495) ,304,305.n of 494 .ato) gene cluster from Amycolatopsis tolypomycina NRRL B-24205 in S. albus led to the characterization of two unprecedented tricyclic sesterterpenoids atolypenes A (497) and B (498) [499), a new highly oxygenated unique tetracyclic 6-hydroxymeroterpenoid, was derived from Nocardiopsis sp. LGO5 [484485486487488489490 R=OCH3491 R=H492 R=OCH3493 R=H494 R=OCH3495 R=H496499497 R498 RCloning and activation of the atolypene ( B (498) . Terretosp. LGO5 .4844854Streptomyces, however, terpene biosynthesis by Allokutzneria, Amycolatopsis, Frankia, Kitasatosporia, Nocardia, Salinispora, Verrucosispora, etc., have been recently reported  contains more than 3000 strains of actinomycetes with a wide range of metabolic capabilities, which are promising for biocatalytic production of terpene derivatives [An extensive development of genome-sequencing technologies and bioinformatics tools have allowed the discovery of BCGs (including silent ones) in the genome of actinomycetes. That terpenoids and meroterpenoids are predominantly found among entified . In addirganisms ,50,310. olyspora , Nocardirdiopsis , Rhodocodococcus ,314, Salinispora , Verrucoosispora , and Actnomadura  have beeMF730-N6  and N. t IFM0406  revealed IFM0406 . Genomic IFM0406 ,310,321.ivatives ,325,326 epi-zizaene, pentalenene, cucumene, (E)-biformene synthases, and other TSs isolated from streptomycetes were characterized. In turn, genome mining of streptomycetes as producers of naphthoquinone-based meroterpenoids led to the discovery of unique prenyltransferase (PTase) and vanadium-dependent haloperoxidase enzymes (VHPO) [epi-zizaene synthase, the successful application of site-directed mutagenesis of the enzyme to control the range of the compounds produced was proved [Unlike the biosynthesis of well-studied secondary metabolites, such as polyketides and nonribosomal peptides, the prediction of terpene structures requires detailed understanding of the cyclization mechanisms and the structural characteristics of bacterial TSs ,327. In s (VHPO) ,183. Fors (VHPO) . It has s (VHPO) . By the s proved ,122 (WO2S. cinnabarinus PK209 and S. hygroscopicus HOK021 (NITE P-02560) synthesize the diterpene lobocompactol and the antibiotic platensimycin in the presence of the Gram-negative Alteromonas sp. KNS-16 [Tsukamurella pulmonis TP-B0596 (JP2019149945), respectively. The effectiveness of actinomycete terpenoids and meroterpenoids, namely pentalenolactone, albaflavenone, platensimycin, platencin, terpentecin, lavanducyanin, marinocyanins A\u2013C, furaquinocin L, 3-dechloro-3-bromonapyradiomycin A1, napyradiomycin A1, and merochlorin A, as promising antibiotics has been proven. This is true for cyslabdan, which enhances the action (1000-fold) of the antibiotic imipenem against MRSA. In addition to high antibacterial activity, many meroterpenoids, such as napyradiomycins B1, B3, B4, A80915A, B, C, furaquinocins A and B, murayaquinone, marinocyanin A\u2013C, and saccharoquinoline, exhibit a high cytotoxic activity against different cancer cell lines , which spread Streptomyces spores in the soil [S. coelicolor M145 spores, they synthesize albaflavenone, which may coordinate the development of the producer (quorum sensing) and/or play a role in the competitive repression of microflora (quorum suppression) in the natural environment [S. coelicolor A3(2) does not produce aminobacteriohopanetriol or produces this compound in negligible amounts. However, the triterpene generation increased sharply during the formation of an aerial mycelium and sporulation, which may be associated with structural changes in the membrane and protection against water loss [Micromonospora isolates revealed TS genes, which differ significantly from other groups of characterized bacterial TSs and may be useful as markers of the genus, while Mycobacterium tuberculosis H37Rv\u043d produced specific diterpene nucleosides, 1- and 6N-tuberculosinyladenosines, promising for development as specific diagnostic markers of tuberculosis.Actinomycete-derived terpenoids participate in specific interactions with macroorganisms , regulate the bacterial life cycle, perform protective functions, or serve as taxonomic markers. Bacterial terpenoids are often optical isomers of plant terpenoids and may represent two chemical communication channels that do not overlap even if the same habitat is occupied by prokaryotic and eukaryotic organisms producing terpenes . Soil-smthe soil . Accordithe soil . \u010cih\u00e1k eironment . In the ter loss . In addiDespite the significant (more than 300) number of publications on terpene biosynthesis by actinomycetes, the conducted patent analysis revealed only 26 patents in this research area . TerpenoThus, the synthesis of terpenes and terpenoids is an important pathway in the secondary metabolism of actinomycetes. The compounds produced may be promising therapeutic agents for the treatment of viral, inflammatory, cancerous, and other diseases in the future. Terpenoids and meroterpenoids synthesized by actinomycetes and possessing high antibacterial activity against drug-resistant pathogenic microorganisms may be useful for the development of new antibiotics. Further study of actinomycetes, accumulation of genetic information about this group of microorganisms, and employment of modern and development of novel tools of synthetic biology and genetic engineering will open prospects for creation of ideal \u201ccell factories\u201d using actinomycetes."}
+{"text": "C5 convertase cleaves C5 in common complement pathways yielding the anaphylatoxin C5a and C5b, part of the membrane attack complex (MAC) that mitigates bacterial infections . C5 can also be cleaved \u201cextrinsically\u201d by other enzymes. C5a is elevated in severe COVID-19 patients (Pt). Yet, inhibiting C5 cleavage increases infections without improved survival in critically ill COVID-19 Pt . In addition, eculizumab, a C5 blocker does not prevent C5a from being generated in COVID-19 Pt . Vilobelimab (Vilo), an anti-C5a monoclonal antibody that preserves MAC, was tested in a Phase 3 multicenter, double-blind, randomized, placebo (Plc)-controlled study for its effect on survival in critically ill COVID-19 Pt .COVID-19 Pt recruited worldwide  intubated within 48 hrs before treatment, received up to 6, 800mg infusions of Vilo or Plc on top of standard-of-care. The primary endpoint was 28-Day all-cause mortality with key secondary endpoints of 60-Day mortality and safety. Blood samples were assessed by ELISA at screening, Day 8 and at hospital discharge for Vilo and C5a levels. No meningococcal vaccination or prophylactic antibiotics were used in the study.vs 41.6% for Plc  with similar 60-Day results. On Day 8 after 3 infusions, mean Vilo trough levels were 21,799.3 to 302,972.1 ng/mL . C5a was highly elevated and comparable between groups at screening: Vilo mean 130.3 ng/mL, Plc mean 123.2 ng/mL. By Day 8, C5a levels were reduced 87% for Vilo  vs an increase for Plc (mean 129.8 ng/mL). Though post Day 8 sampling was sparse, C5a levels remained elevated for Plc and low for Vilo throughout the study. Serious AEs were 58.9% for Vilo and 63.5% for Plc. Treatment-emergent AEs were 90.9% for Vilo vs 91.0% for Plc. Infection incidence per 100 Pt days was comparable in both groups.Kaplan-Meier estimates showed a 28-Day mortality rate of 31.7% for Vilo Results from this Phase 3 study show that direct C5a inhibition by vilobelimab while presumably preserving MAC, as opposed to upstream C5 blockade, results in a survival benefit for critically ill COVID-19 Pt without increasing infections.Bruce P. Burnett, PhD, InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Employee Alexander Vlaar, MD, PhD, InflaRx: Advisor/Consultant Maria Habel, PhD, InflaRx GmbH: Employee Claus Thielert, PhD, InflaRx GmbH: Employee James Dickinson, MSc, InflaRx GmbH: Employee simon R\u00fcckinger, PhD, Metronomia Clinical Research GmbH: Advisor/Consultant Robert Zerbib, MSc, InflaRx GmbH: Advisor/Consultant Dorothee Neukirchen, PhD, InflaRx GmbH: Employee Renfeng Guo, MD, inflarx: Board Member|inflarx: I am an Inventor for Vilobelimab|inflarx: Ownership Interest|inflarx: Stocks/Bonds|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Board Member|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Founder|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Employee|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Ownership Interest Niels Riedemann, MD, PhD, InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Board Member|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Founder|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Employee|InflaRx GmbH, InflaRx Pharmaceuticals, Inc.: Ownership Interest"}
+{"text": "Eminium intortum (Banks & Sol.) Kuntze and E. spiculatum (Blume) Schott (Araceae). A total of 83 metabolites, including 19 phenolic acids, 46 flavonoids, 11 amino, and 7 fatty acids were identified by UHPLC-HRMS in the studied extracts for the first time. E. intortum flower and leaf extracts had the highest total phenolic and flavonoid contents . Significant radical scavenging activity (32.20 \u00b1 1.26 and 54.34 \u00b1 0.53 mg TE/g for DPPH and ABTS) and reducing power (88.27 \u00b1 1.49 and 33.13 \u00b1 0.68 mg TE/g for CUPRAC and FRAP) were observed in leaf extracts. E. intortum flowers showed the maximum anticholinesterase activity (2.72 \u00b1 0.03 mg GALAE/g). E. spiculatum leaves and tubers exhibited the highest inhibition towards \u03b1-glucosidase (0.99 \u00b1 0.02 ACAE/g) and tirosinase (50.73 \u00b1 2.29 mg KAE/g), respectively. A multivariate analysis revealed that O-hydroxycinnamoylglycosyl-C-flavonoid glycosides mostly accounted for the discrimination of both species. Thus, E. intortum and E. spiculatum can be considered as potential candidates for designing functional ingredients in the pharmaceutical and nutraceutical industries.The study aimed at the metabolite profiling and evaluation of antioxidant and enzyme inhibitory properties of methanol extracts from flowers, leaves, and tubers of unexplored  The search for bioactive natural compounds derived from medicinal plants is of great importance in modern medicine . NaturalEminium (Blume) Schott is a genus of the Araceae family, which includes over 300 species that grow mostly in tropical areas \u2212 at m/z 191.018 showed fragment ions at 173.008 [M-H-H2O]\u2212, 147.028 [M-H-CO2]\u2212, a base peak at m/z 111.07 [M-H-CO2-2H2O]\u2212, and was related to citric acid \u2212 at m/z 623.162 gave a base peak at m/z 299.056, corresponding to the concomitant loss of two hexoses, and was annotated as chrysoeriol 7-O-dihexoside. MS/MS spectrum of 54 [M-H]\u2212 at m/z 577.1568 showed fragment ions at m/z 431.098 [M-H-dHex]\u2212 and 413.088 [M-H-dHex-H2O]\u2212, and a base peak at m/z 269.045 [M-H-dHex-Hex]\u2212. The absence of the interglycosidic linkage breakdown is favored for 7-neohesperidoside. Thus, 54 was tentatively identified as apigenin 7-O-neohesperidoside -120]\u2212, [(M-H)-90]\u2212, and [(M-H)-30]\u2212. C-8 isomers were characterized by the base peak 0,2 X\u2212 [(M-H)-120]\u2212 as MS/MS of orientin (30), while C-6 isomers showed the base peaks [M-H]\u2212 as homoorientin (27), isovitexin (37), and chrysoeriol 6-C-hexoside (44), together with abundant fragment 0,2 X\u2212 [(M-H)-120]\u2212 (about 60%). In addition, the fragment ion 0,3 X\u2212 [M-H-90]\u2212 was favored by C-6 isomers (20 and 22 gave indicative ions at m/z 519.113 and 503.119 [(M-H)-90]\u2212, 489.103, and 473.109 [(M-H)-120]\u2212, 429.082 and 413.089 [(M-H)-120-60]\u2212, 399.072 and 383.077 [(M-H)-120-90]\u2212, 369.061 and 353.066 [(M-H)-2 \u00d7 120]\u2212, respectively. Based on 1,3 B\u2212 fragments at m/z 133.028 (20) and 117.033 (22), 20 and 22 were tentatively annotated as 6,8-C-diglucosides of luteolin and apigenin, respectively  and 0,3 X1 (\u221290) or 0,2 X1 (\u2212120) at m/z 357.062 and 327.051, respectively  was discernable by the fragment ions at m/z 341.067 ([(M-H)-132-90]\u2212 and 311.0561 ([(M-H)-132-120]\u2212, suggesting the presence of both O-pentosyl (X1) and C-hexosyl (X0) moieties. Additionally, the aforementioned structure was assigned on the basis of a series of fragment ions at 283.061 , 282.055 , 281.045 , and 237.055  (Among three isobars indicated as ectively . In the  X0/2CO) .29 with ([M-H]\u2212 at m/z 771.179 was acquired. In (-) ESI mode, 29 yielded prominent fragment ions at m/z 447.093 [(M-H)-2 \u00d7 162.05]\u2212, 357.062 [(M-H)-2 \u00d7 162.05-90]\u2212, 327.051 ([(M-H)-2 \u00d7 162.05-120]\u2212, suggesting the presence of both O-dihexosyl and C-hexosyl moieties. The absence of the interglycosidic linkage breakdown is favored for O-sophoroside. Thus, 29 was annotated as luteolin O-sophoroside-6-C-hexosyl-orientin. Compound 32 ([M-H]\u2212 at m/z 593.151) yielded prominent ions at m/z 413.088 ([(M-H)-(162.05 + H2O)]\u2212, 353.067 ([(M-H)-(162.05 + H2O)-60]\u2212, 311.056 ([(M-H)-(162.05)-120]\u2212, 293.047 [(M-H)-(162.05 + H2O)-120]\u2212, and 281.045 Y1\u2212/0,2 X0/CH2O indicating an O-hexosyl unit at the 2\u201dof the primary hexose [32 was assigned to 2\u2033-O-hexosyl-6-C-hexosyl-apigenin  and 45 ([M-H]\u2212 at m/z 799.210) shared the same fragmentation patterns yielding prominent ions at m/z 447.094 (34) and 431.098 (35), respectively, indicating a concomitant loss of hexose (162.05 Da) and sinapoyl residue  (m/z 205.049 [(sinapic acid-H)-H2O]\u2212, 190.027 [(sinapic acid-H)-H2O-CH3]\u2212, and 175.003 [(sinapic acid-H)-H2O-2\u00b7CH3]\u2212. The presence of a C-hexosyl moiety on the flavonoid skeleton was deduced from the relative abundances of the ions at m/z 327.051 [M-H-120]\u2212 (79.7%) (34) and 311.056 (100%) (45) suggesting a 6-C- and an 8-C-linkage, respectively. Thus, 34 was ascribed as luteolin O-sinapoylhexosyl-6-C-hexoside, while 45 was annotated as apigenin O-sinapoylhexosyl-6-C-hexoside (Two compounds 11H10O4) . Moreovehexoside .36 and 41 were discernable by the transition [M-H]\u2212\u2192Y0 resulting from the indicative loss of 324.085 Da (C15H16O8). Precursor ions gave a series of fragment ions involved in the C-glycosyl flavon pattern: m/z 353.064 , 341.067 , 311.056 , and 283.061  (m/z 161.023 [(caffeic acid-H)-H2O]\u2212 and 133.028[(caffeic acid-H)-H2O-CO]\u2212. Accordingly, 36 and 41 were assigned to apigenin O-caffeoylhexosyl-8-C-hexoside and chrysoeriol O-caffeoylhexosyl-8-C-hexoside (Two caffeoyl esters 2 X1/CO) . Caffeoyhexoside .O-hexosyl-8-C-hexoside  were deduced from the loss of feruloylhexosyl residue  and the subsequent transition of m/z 447.094/431.099/461.109\u2192327.051/311.056/341.067, respectively, arising from the hexose cross ring cleavage  (m/z 175.039 [(ferulic acid-H)-H2O]\u2212, 160.015 [(ferulic acid-H)-CH3]\u2212, and [(ferulic acid-H)-H2O-CH3-CO]\u2212 point out to the feruloyl moiety  . The ionl moiety   in the fragmentation patterns of the aforementioned compounds, accompanied with the ions at m/z 163.039 [p-coumaric acid-H]\u2212, 145.028 [(p-coumaric acid-H)-H2O]\u2212, and 117.033 [(p-coumaric acid-H)-H2O-CO]\u2212 confirmed the presence of a coumaroyl moiety (O-deoxyhexosyl-6-C-hexoside (60 and 62) were evidenced on the base of the loss of 292 Da (C15H16O6)    .25, 27, 30, 35, 37, 42, 47, 50, 52, 55, 63, and 64 was confirmed by comparison with reference standards.The identification of flavonoids 71 and 73), a dipeptide (74), and six amino acid hexosides were tentatively annotated , a series of fragment ions resulting from neutral losses were registered at m/z 241.119 [M-H-H2O]\u2212, 223.108 [M-H-2H2O]\u2212, 215.140 [M-H-CO2]\u2212, 197.1287 [M-H-H2O-CO2]\u2212, and a base peak at m/z 128.033, corresponding to the loss of leucine [M-H-131.096]\u2212 from the glutamyl residue. Thus, 74 was related to \u03b3-glutamyl-leucine [66, 67, 68, 69, 70, and 72 demonstrated base peaks, resulting from the loss of a hexose moiety, with the appearance of the corresponding amino acid residue. Hence, they were annotated as hexosides of glutamic acid, valine, tyrosine, leucine, phenylalanine, and tryptophan, respectively , three monounsaturated , and five polyunsaturated  free fatty acids were tentatively annotated in Eminium extracts (78\u201382) were dihydroxylated and three (75\u201377) contained three hydroxyl groups , l groups . Compounuwolffii .E. intortum and E. spiculatum methanolic extracts, we used six different methods including DPPH, ABTS cation, FRAP, CUPRAC, phosphomolybdenum, and metal chelate assays. As shown in E. intortum exhibited a much stronger antioxidant activity compared to the extracts of E. spiculatum, with values of 32.20 \u00b1 1.26 mg TE/g for leaves, 27.87 \u00b1 0.75 mg TE/g for flowers, and 26.90 \u00b1 1.61 mg TE/g for tubers. The methanolic extract of the tuber of E. spiculatum also showed good antioxidant activity with a value of 33.67 \u00b1 1.26 mg TE/g.To study the antioxidant activity of E. intortum and 86.27 \u00b1 2.74 for E. spiculatum. The methanolic extracts of the tubers and flowers of both plants showed lower antioxidant activity compared to the leaves.E. spiculatum and E. intortum showed very significant reducing activity, with values of 41.33 \u00b1 1.25 mg TE/g and 32.58 \u00b1 4.02 mg TE/g, respectively. The other extracts of both plants also showed significant results of antioxidant activity in this assay. In summary, our findings suggest that the extracts are rich in phenolic compounds with radical-scavenging activity and proton-donating ability. Polyphenols have been acknowledged for their antioxidant activity, which may account for their potential ability to prevent various diseases associated with oxidative stress. This assertion is supported by several studies [E. intortum and 61.55 \u00b1 3.97 mg EDTAE/g for E. spiculatum. It should be noted that this method is also highly specific.In the FRAP assay, a high absorbance indicates a high reducing power. The methanolic extracts of the tubers of both  studies ,28,29,30The antioxidant activity of a substance is influenced by how it interacts with radicals in the reaction medium. These interactions are facilitated by active molecules that trap the radicals, and previous studies have highlighted their importance ,32,33. TEminium genus. Alkofahi et al. [E. spiculatum extract against oxidative DNA damage using the 8-hydroxydeoxyguanosine assay, while no beneficial effect on alleviating oxidative damage was observed. Al-Ismail et al. [E. spiculatum ethanolic extract using DPPH, FRAP, and vegetable oil emulsion systems, where the extract exhibited significant properties with lower IC50 values. Additionally, Janat and Al-Thnaibat [E. spiculatum exhibited higher antioxidant activity in the phosphomolybdenum assay when compared to the aqueous extract.Limited literature exists regarding the antioxidant properties of the i et al.  conductel et al.  reportedThnaibat  reportedIn the intestinal tract, the breakdown of complex sugars into simple sugars is facilitated by two essential enzymes, namely \u03b1-amylase and \u03b1-glucosidase. The simple products, particularly glucose, are subsequently absorbed and can cause a rise in blood sugar levels. To manage diabetes, one potential therapeutic approach is to inhibit the enzymes responsible for carbohydrate hydrolysis, which can decrease postprandial blood sugar levels ,41. By iE. spiculatum also exhibited \u03b1-glucosidase inhibition, with values of 0.99 \u00b1 0.02 ACAE/g and 0.89 \u00b1 0.01 ACAE/g, respectively. Among E. intortum extracts, the leaf extract was only active against \u03b1-glucosidase (0.35 mmol ACAE/g). In addition, both tuber extracts showed no inhibitory effect on \u03b1-glucosidase. Our results demonstrate the potential of the studied leaf extracts to act as inhibitors of carbohydrate hydrolyzing enzymes. By delaying the absorption of dietary carbohydrates in the small intestine and reducing postprandial hyperglycemia, these inhibitors may be a valuable component in the development of antidiabetic drugs. This mechanism of action has been previously reported [reported ,46,47.Our study is the first to investigate the antidiabetic activity of the two plants, and we found that all studied extracts of the plants inhibited \u03b1-amylase activity, with values lower than 0.31 \u00b1 0.01 ACAE/g. The inhibitory effect of phenolic compounds on carbohydrate hydrolysis enzymes has been reported previously ,49,50, aAcetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are enzymes found primarily in nerve tissues and neuromuscular junctions. They are responsible for rapidly hydrolyzing acetylcholine, a neurotransmitter, into inert choline and acetate. Overexpression or excessive catalysis of AChE can cause neuronal disturbances and lead to neurological disorders. A potential strategy for neuroprotection is to inhibit AChE activity ,52,53. OE. spiculatum and E. intortum tubers exhibited strong inhibitory activity with values of 50.73 \u00b1 2.29 KAE/g and 48.13 \u00b1 0.24 KAE/g, respectively. Natural compounds capable of inhibiting tyrosinase activity are of great interest, with increasing demand in the fields of cosmetics and pharmaceuticals [In skin cells, tyrosinase plays a crucial role in the aging process, and inhibiting its activity is a key strategy for delaying skin aging. This enzyme catalyzes the initial two steps of melanogenesis, making it a rate-limiting factor in this process. Mutations in the tyrosinase gene or its absence can cause a reduction or cessation of pigmentation ,58. In oeuticals ,60. Numeeuticals ,63,64,65E. spiculatum. In contrast, the flower and leaves of E. intortum were grouped into Cluster III and IV, respectively  was mainly associated with antioxidant assays and AChE. The second component (PC2) contained amylase, BChE, and tyrosinase, while the third component (PC3) included glucosidase and DPPH B. Althouectively C.E. spiculatum, which contained high levels of chrysoeriol O-coumaroylhexoside-8-C-hexoside, luteolin-6,8-C-diglucoside, and apigenin 7-O-synapoylhexosyl-8-C-hexoside. The leaves and flowers of E. intortum were classified in Cluster II, which were characterized by high levels of luteolin 7-O-coumaroylhexosyl-6-C-hexoside and apigenin O-feruloylhexosyl-8-C-hexoside. Cluster III contained both tuber samples. Furthermore, we conducted a correlation analysis between the biological activities and individual components, and the results are shown in C-hexoside-8-C-pentoside was the main contributor to DPPH scavenging ability, while sinapic acid-O-dihexoside and isorhamnetin-3-O-rutinoside were positively correlated with ABTS and CUPRAC. Consistent with our findings, the compounds have been described as important antioxidants by several researchers [The classifications of the tested extracts based on their chemical components is presented in earchers ,67,68. Aearchers .Eminium species in Turkey (Eminium intortum (Banks & Sol.) Kuntze: Between K\u0131z\u0131ltepe and Mardin, 533 m, GPS: 37\u00b014\u203220\u2033 N, 40\u00b038\u203210\u2033 E; Eminium spiculatum (Blume) Schott: Derik, Mardin, 661 m, GPS: 37\u00b018\u203244\u2033 N, 40\u00b016\u203216\u2033 E). The plant specimens were identified by one of our co-authors, Dr. Ugur Cakilcioglu, and one specimen from the plants was deposited at the Harran University herbarium. Prior to extraction, the plant materials were carefully washed with tap and distilled water to eliminate any soil and contaminants. After being air-dried for 10 days (in shade at room temperature), the flowers, leaves, and roots were powdered.In the summer of 2021, we collected For extraction, we employed the maceration method, in which 5 g of plant material was mixed with 100 mL of methanol and left to macerate at room temperature for 24 h. After maceration, the extracts were filtered through Whatman filter paper, and the solvents were evaporated using a rotary-evaporator. To preserve the extracts, we stored them at 4 \u00b0C until analysis.p-coumaric, o-coumaric, ferulic acids, saponarin, homoorientin, orientin, rutin, isovitexin, isoquercitrin, luteolin 7-O-glucoside, kaempferol 3-O-rutinoside, kaempferol 3-O-glucoside, isorhamnetin 3-O-glucoside, apigenin 7-O-glucoside, quercetin, and apigenin. Chlorogenic and caffeic acids were supplied from Phytolab .Acetonitrile (hypergrade for LC\u2013MS), formic acid (for LC\u2013MS) and methanol  were purchased from Merck . The reference standards used for compound identification were obtained from Extrasynthese  for We determined the total phenolic and flavonoid content of the extracts using the Folin\u2013Ciocalteu and AlCl3 assays, respectively, according to Zengin and Aktumsek\u2019s protocol (2014). The results of these tests were reported in terms of gallic acid equivalents (mg GAE/g dry extract) and rutin equivalents (mg RE/g dry extract) .m/z range from 100 to 1000. The mass spectrometer parameters were as follows: spray voltage 3.5 kV (+) and 2.5 kV (\u2212); sheath gas flow rate 38; auxiliary gas flow rate 12; spare gas flow rate 0; capillary temperature 320 \u00b0C; probe heater temperature 320 \u00b0C; S-lens RF level 50; scan mode: full MS , and MS/MS . The chromatographic separation was achieved on a reversed phase column Kromasil EternityXT C18  at 40 \u00b0C. The UHPLC analyses were run with a mobile phase consisting of 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B). The run time was 33 min. The flow rate was 0.3 mL/min. The gradient elution program was used as follows: 0\u20131 min, 0\u20135% B; 1\u201320 min, 5\u201330% B; 20\u201325 min, 30\u201350% B; 25\u201330 min, 50\u201370% B; 30\u201333 min, 70\u201395%; 33\u201334 min 95\u20135% B. Equilibration time was 4 min [The UHPLC-HRMS analyses were carried out on a Q Exactive Plus mass spectrometer  equipped with a heated electrospray ionization (HESI-II) probe (ThermoScientific). The equipment was operated in negative and positive ion modes within the as 4 min . Data weas 4 min .We analyzed the extracts for a range of antioxidant and enzyme inhibitory activities, including, cupric reducing antioxidant capacity (CUPRAC), DPPH, and ABTS radical scavenging, metal chelating activity (MCA), ferric reducing antioxidant power (FRAP), phosphomolybdenum (PBD), and inhibition of amylase, tyrosinase, glucosidase, acetylcholinesterase (AChE), and butyrylcholinesterase (BChE). We employed the previously described methods to evaluate these activities . Each sap < 0.05). Then, Principal Component Analysis (PCA) and hierarchical clustered analysis (HCA) were performed to emphasize the distinct clusters in terms of their bioactivities. Furthermore, hierarchical clustered analysis (HCA) was performed to assess the (dis)similarity between samples in terms of their molecules. All used data were scale and molecules data and were log transformed before doing multivariate analysis. R v.4.2.3 statistical program was used for all analyses.All data were given as mean \u00b1 standard deviation (SD). Statistical analysis was performed by analysis of variance (ANOVA). A post hoc test (Tukey) was performed when the differences shown by data were significant  varied according to the plant parts used. Generally, the leaf extract of both species exhibited higher antioxidant effects compared to flowers and tubers. However, we obtained different results for each enzyme inhibition assay, and the leaf extracts provided good anti-amylase and anti-glucosidase actions. The extracts were rich in flavonoids. These findings provide a valuable scientific basis for evaluating the potential of the Eminium genus and suggest that the tested species could be considered as a source of natural bioactive agents for health-promoting applications. Nevertheless, additional research is necessary to elucidate the potential toxicity of both the extracts and the individual chemical constituents.In our study, we explored the detailed chemical composition and biological effects of two"}
+{"text": "Pseudomonas aeruginosa.To evaluate the efficacy of ceftazidime-avibactam (CZA) and aztreonam-avibactam (AZA) against bloodstream infections (BSIs) or lower respiratory tract infections (LRTIs) \u2013 caused by extensive drug-resistant or pan drug-resistant (XDR/PDR) P. aeruginosa. Whole-genome sequencing was used to analyze the resistance determinants of each isolate. Monte Carlo simulations (MCSs) were used to evaluate the probability of target attainment (PTA) and the cumulative fraction of response (CFR) of each CZA/AZA dosing regimen via traditional infusion (TI)/optimized two-step-administration therapy (OTAT).The two-fold dilution method was used to determine the minimum inhibitory concentrations (MICs) of CZA/AZA against XDR/PDR P. aeruginosa isolates producing IMP-45, VIM-1, or VIM-2 were inhibited by AZA at a concentration of 2 to 8 mg/L. All isolates producing IND-6 plus other serine \u03b2-lactamases were high-level resistant to CZA/AZA (MICs >64 mg/L). All simulated dosing regimens of CZA/AZA against BSIs-causing XDR/PDR P. aeruginosa achieved 100% PTA when the MIC was \u226432 mg/L.We found that XDR/PDR P. aeruginosa may carry some rare MBLs . P. aeruginosa producing IMP-45, VIM-1, or VIM-2. OTAT with sufficient pharmacodynamic exposure may be an optimal treatment option for XDR/PDR P. aeruginosa with a high-level MIC of CZA/AZA.AZA has been considered as an option for the treatment of infections caused by XDR/PDR  P. aeruginosa displays resistance to various antibiotics, making treatment challenging . CZA has good safety and was regarded as a vital treatment option for P. aeruginosa infections. CZA showed good capabilities against P. aeruginosa with a sensitivity rate ranging from 76.2% to 97.8% (P. aeruginosa infections ranged from 64.3% to 90.6% (Ceftazidime/avibactam (CZA) is a novel \u03b2-lactam/\u03b2-lactamase inhibitor (BL/BLIs). And it was approved by the Chinese National Medical Products Administration (CNMPA) in 2019 for the treatment of complicated intra-abdominal infections (cIAIs), hospital-acquired pneumonia (HAP), and ventilator-associated pneumonia (VAP), and in adult patients with limited treatment options for infections caused by the following gram-negative bacteria sensitive to this product: to 97.8% . In termto 90.6% .\u03b2-lactamases  against which ATM is ineffective. Avibactam (AVI) is a \u03b2-lactamase inhibitor with a wide enzyme inhibition spectrum, including Ambler class A , class C (AmpC), and class D (OXA-48 type) \u03b2-lactamase. The combination of ATM and AVI can potentially inhibit MBL-producing bacteria (Aztreonam (ATM) was the first monobactam antibiotic to be used in clinical therapy. It was approved by the U.S. Food and Drug Administration (FDA) in 1986 for treatment of various infections caused by sensitive aerobic gram-negative bacteria. ATM was stable to hydrolysis by Ambler class B Metallo-\u03b2-lactamases (MBLs) . Howeverbacteria .P. aeruginosa in this study. Whole-genome sequencing and bioinformatic analysis were used to identify resistance genes of each isolate. Besides, we combine the population pharmacokinetic parameters (PPKs) of CZA/AZA with minimum inhibitory concentration (MIC) distribution of XDR/PDR P. aeruginosa to evaluate the efficacy of various dosing regimens. Therefore, the objectives of our work are as follows. Firstly, our work aims to compare the in vitro activity of CZA and AZA against XDR/PDR P. aeruginosa. Secondly, our team wants to evaluate the relationship between resistance genes and CZA/AZA sensitivity rates of XDR/PDR P. aeruginosa. Thirdly, we assess the efficacy of CZA and AZA for the treatment of critically ill patients with BSIs/LRTIs caused by XDR/PDR P. aeruginosa.We performed antimicrobial susceptibility testing (AST) on XDR/PDR 22.1P. aeruginosa from critically ill patients admitted to the First Medical Centre of Chinese PLA General Hospital from January 2016 to November 2021. A total of 10 P. aeruginosa strains were categorized as XDR according to CLSI criteria. Moreover, 57 P. aeruginosa strains were categorized as PDR . Ceftazidime, avibactam, and aztreonam standards were purchased from MedChemExpress. The resistance rates of XDR/PDR Pseudomonas aeruginosa to cefoperazone-sulbactam, imipenem, ciprofloxacin, piperacillin-tazobactam, ceftazidime, levofloxacin, meropenem, tobramycin, amikacin and gentamicin were 89.6%, 100%, 89.6%, 100%, 97%, 89.6%, 100%, 97.1%, 100%, 100%, respectively.We collected 67 d as PDR . All P. 2.2P. aeruginosa. A fixed concentration of AVI at 4 mg/L, 8mg/L, and 16mg/L combined with 2-fold diluted CAZ and ATM were used in ASTs. The quality control strains of our tests were E. coli ATCC 25922 and P. aeruginosa ATCC 27853. The MICs are defined as the lowest concentration of antibiotics that inhibits the growth of bacteria. The definition of MIC50 is a drug concentration that inhibits the growth of bacteria by 50%. Similarly, MIC90 is a drug concentration inhibiting 90% of bacterial growth. MIC distributions of CZA/AZA against P. aeruginosa were represented by cumulative inhibition rates (CIRs). Besides, all experiments were conducted three times following Clinical and Laboratory Standards Institute (CLSI) standards.We used the broth two-fold dilution method to determine the minimum inhibitory concentrations (MICs) of CZA/AZA against XDR/PDR 2.3P. aeruginosa were subjected to whole-genome sequencing (WGS) using Illumina MiSeq short-read sequencing . Sequenced isolates were evaluated using FASTQC, version 0.11.6, and MultiQC, version 1.6. Trimmomatic, version 0.39, removed adapters and trimmed low-quality paired end reads. Comprehensive Antibiotic Resistance Database v.1.2.0  was used to identify drug resistance genes in the strains.67 2.4fT > MIC is the best indicator for assessing BSIs. Besides, %50fT > 5 \u00d7 MIC is the best indicator for assessing LRTIs. When combined with CAZ, %50fT > CT of 1 mg/L was considered the Pharmacokinetic (PK)/Pharmacodynamics (PD) target of avibactam. PK/PD targets of ATM for BSIs was %60fT > MIC. And PK/PD targets of ATM for LRTIs were %60fT > 5 \u00d7 MIC. As for avibactam, %50fT > CT of 2.5 mg/L was considered appropriate for guiding dosage selection for AZA (fT > n \u00d7 MIC equation was based on previous studies (Population pharmacokinetic (PPK) parameters of CZA and AZA were obtained from previously published articles . CAZ is  for AZA . Optimiz studies .2.5d, CL, t1/2) followed a log-normal distribution. All simulated dosing regimens via traditional infusion (TI)/optimized two-step-administration therapy (OTAT) were listed in MICi means each MIC value. p(MICi) means the percentage of each MIC value. A CFR \u2265 90% is adequate PD exposure for this dosing regimen.We conducted 10000-patient Monte Carlo simulations (MCSs) using Oracle Crystal Ball version.11.1.24. PK parameters  were listed in P. aeruginosa with an OXA-101 also produced OXA-850. CZA was as effective as AZA against these isolates. For isolates with an OXA gene plus IMP-45, VIM-1, or VIM-2, AZA was much more potent than CZA against these isolates, with all isolates being inhibited by a concentration of 8 mg/L. For isolates with an IND-6 plus more serine \u03b2-lactamases, all isolates produced CZA/AZA MICs of >64 mg/L. For XDR/PDR P. aeruginosa with other genotypes, the efficacy of CZA against these isolates did not differ significantly from that of AZA.The comparative MICs (mg/L) of CZA and AZA against 67 XDR/PDR 3.43.4.1The probability of target attainment (PTA) of each simulated dosing regimen for BSIs was listed in 3.4.2The PTA of each simulated dosing regimen for LRTIs was listed in 3.53.5.1via traditional infusion, the CFR was 95.63% for 4g q6h. If CZA was administered via OTAT, the CFR was 96.67% for 1.25g (0.5h) +1.25g (2h) q8h, 97.35% for 1.25g (0.5h) +1.25g (2h) q6h, 93.60% for 0.675g (0.5h) +0.675g (2h) q8h and 93.53% for 0.675g (0.5h) +0.675g (2h) q8h. If AZA was administered via traditional infusion, the CFR was 92.50% for 2.5g q6h and 96.36% for 4g q6h. If AZA was administered via OTAT, the CFR was 96.95% for 1.25g (0.5h) +1.25g (2h) q8h, 98.25% for 1.25g (0.5h) +1.25g (2h) q6h. The above dosing regimens are considered to provide adequate PD exposures for the treatment of CZA/AZA against XDR/PDR P. aeruginosa BSIs.The cumulative fraction of response (CFR) of each simulated dosing regimen for BSIs was listed in 3.5.2P. aeruginosa. CFRs were less than 90% for all simulated dosing regimens  + 1.25 g (2h)] q6h, 2.5 g  q8h, 4g  q6h, 4g  q8h).4P. aeruginosa has spread widely worldwide and the treatment of BSIs or LRTIs caused by XDR/PDR P. aeruginosa has become a tough problem . AVI is ineffective against MBL-producing isolates. However, the combination of AVI and ATM was a treatment option for MBL-producing isolates. Lee et\u00a0al. also found that the combination of ATM and CZA may be a treatment option for VIM-2-producing P. aeruginosa . IND-6 is a highly divergent IND-type MBL. It was first isolated from Chryseobacterium indologenes strain 597 in Burkina Faso   . Besidesutations . Mobile fections . Our worinfusion . Besideshe blood .P. aeruginosa was confined to a small sample size and northern China. Secondly, our study only focused on partial beta-lactamases (class B \u03b2-lactamases and class D \u03b2-lactamases). Therefore, large-scale animal or clinical trials are needed in the future to confirm the efficacy of CZA/AZA against BSIs caused by XDR/PDR P. aeruginosa.Our study had several limitations that should be noted. Firstly, the collection of P. aeruginosa are intrinsic, mutational, and horizontally acquired resistomes (P. aeruginosa drug resistant. The efflux pump was also considered in the study of this article (see the whole genome sequencing results in the The main resistance mechanisms in sistomes . We founP. aeruginosa harbouring IMP-45, VIM-1, or VIM-2. Secondly, OTAT with sufficient PD exposure may be an optimal treatment option for BSI caused by XDR/PDR P. aeruginosa with a high-level MIC of CZA/AZA.In conclusion, our work has the following results. Firstly, AZA was considered as an option for the treatment of XDR/PDR The original contributions presented in the study are publicly available. This data has been deposited into the NCBI repository under accession: PRJNA967114.Ethical approval was granted by the Chinese People\u2019s Liberation Army General Hospital.YK and JC designed and managed the project. YK performed all\u00a0the experiments and wrote the manuscript. LX analyzed bioinformatics analysis. JY provided technical guidance. All\u00a0authors contributed to the article and approved the submitted version."}
+{"text": "Two novel reassortant highly pathogenic avian influenza viruses (H5N1) clade 2.3.4.4b.2 were identified in dead migratory birds in China in November 2021. The viruses probably evolved among wild birds through different flyways connecting Europe and Asia. Their low antigenic reaction to vaccine antiserum indicates high risks to poultry and to public health. Since the Gs/GD/96-lineage highly pathogenic avian influenza virus (HPAIV) (H5N1) was identified in 1996, H5 HPAIVs have evolved into divergent clades and caused continuous outbreaks in birds (https://wahis.woah.org/#/event-management). Human cases of H5N1/H5N6/H5N8 infection were sporadically documented , highlighting the zoonotic risk of H5 HPAIVs. Since 2021, H5 HPAIVs have caused at least 9 outbreaks in wild birds rather than poultry in mainland China . However, large outbreaks of H5N1 HPAIVs in domestic poultry were reported during 2021\u20132022 in Europe and the United States  . In this study, we explored the genetic origin, spread patterns, and antigenicity of H5N1 viruses identified from 2 dead migratory birds in China.Starting during 2020\u20132021, a new wave of HPAIV H5N1/H5N8 clade 2.3.4.4b outbreaks was reported in wild and domestic birds in Eurasia (http://www.mabsky.com).We collected oral swab specimens and lung tissues from a dead whooper swan in northern China (Inner Mongolia) on November 3, 2021, and a deceased black swan in eastern China (Zhejiang) on November 15, 2021. We performed virus isolation in 10-day-old specific pathogen-free chicken embryos  and GISAID . We reconstructed phylogenetic trees for each gene of the 3 H5N1 isolates together with reference viruses from GISAID and the National Center for Biotechnology Information (https://www.ncbi.nlm.nih.gov/genomes/FLU/Database/nph-select.cgi), using the maximum-likelihood method with a general time-reversible model plus gamma distribution in RAxML 8.2.12 (https://cme.h-its.org/exelixis/web/software/raxml) , A/whooper swan/Northern China/11.03 IMEEDSAK2-O/2021 (Ws/NC/AK2-O/2021), and A/black swan/Eastern China/11.15 ZJHZ74-Lg/2021 (Bs/EC/74-Lg/2021). We deposited whole genomes in NMDC (e/raxml)  Table. We/raxml) .https://www.who.int/publications/i/item/manual-for-the-laboratory-diagnosis-and-virological-surveillance-of-influenza) to test the reactivities of antiserum of H5 Re-11/Re-13/Re-14 vaccines against these new H5N1 isolates and H5N8 HPAIVs identified in 2020 (http://www.moa.gov.cn/govpublic).We performed hemagglutination inhibitor (HI) assays ; however, they have only been identified in wild birds in mainland China. Compared with high HI antibody titers (256) between homologous antiserum and antigens of H5 vaccines, the recent H5N1/H5N8 viruses presented low HI titers (2\u201316) against Re-11/Re-13 antiserum  and Bs/2021-like (Bs/EC/74-Lg/2021) reassortants  Figure 4We reconstructed the genetic reassortment history for these H5N1 viruses  clade 2.3.4.4b reassortants in migratory birds, China.Additional information for study of novel avian influenza virus (H5N1) clade 2.3.4.4b reassortants in migratory birds, China."}
+{"text": "Integrase strand-transfer inhibitors (INSTIs) are associated with body weight gain among women living with HIV (WLH) in the Women\u2019s Interagency HIV Study (WIHS). The role of antiretroviral therapy (ART) pharmacokinetics in INSTI-associated weight gain is unknown. We report the relationship between INSTI plasma concentrations and weight change in WLH.Model-adjusted estimates of percent body weight change by integrase inhibitor plasma concentrationModel-adjusted estimates of percent body weight change with 95% confidence band (y-axis) by integrase inhibitor plasma concentration in ng/mL (x-axis). Models were adjusted for baseline age, race/ethnicity, baseline obesity (body mass index \u2265 30 kg/m2), history of prior non-nucleoside reverse transcriptase use, and self-reported antiretroviral therapy adherence (100% vs <100%). Abbreviations: RAL=raltegravir, DTG=dolutegravir, EVG=elvitegravirData from 2006-2017 were analyzed from virologically suppressed (< 200 copies/mL) WLH in the WIHS who switched/added raltegravir (RAL), dolutegravir (DTG), or elvitegravir (EVG) to ART. Percent body weight change was calculated using weights obtained 6-12 months pre- and post- INSTI switch/add. INSTI random plasma concentrations were measured with validated LC/MS-MS assays 6-12 months post INSTI switch/add. Area under the curve (AUC) was calculated using estimates from published models and updating individual-level parameters with plasma concentrations. Linear models assessed the relationship between plasma concentrations or AUC and weight change for each INSTI separately. Models were adjusted for age, race/ethnicity, baseline obesity, history of non-nucleoside reverse transcriptase use, and self-reported ART adherence.2 (\u00b18.8), and 57% reported 100% ART adherence. Mean percent body weight change was +2.3% (\u00b16.0) for RAL, +1.8% (\u00b19.0) for DTG, and +2.8% (\u00b110.5) for EVG. AUC was able to be calculated in 167 (95%) of women. In adjusted linear models, higher RAL plasma concentration was associated with greater percent body weight change (p=0.016), Figure 1. No association of drug AUC with body weight change for any INSTI was observed.176 WLH contributed plasma drug concentrations with a mean \u00b1SD 1.5 (\u00b10.1) years follow-up: 43 (24%) were on RAL, 87 (49%) on DTG, and 47 (27%) on EVG. Mean age was 51 years (\u00b1 9), 64% were Non-Hispanic Black, baseline BMI was 31.6 kg/mIn WLH, random RAL plasma concentrations, but not the RAL AUC, was associated with body weight gain. In contrast, the effect of DTG and EVG plasma drug exposure on short-term body weight change appears to be limited. Mechanisms contributing to body weight gain may be INSTI-specific. In addition to further pharmacologic assessments, other mechanisms to explain INSTI-associated weight gain should be examined.Cecile D. Lahiri, MD, MS, Merck: Grant/Research Support|Theratechnologies, LLC: Advisor/Consultant|Theratechnologies, LLC: Honoraria Julie B. Dumond, PharmD, Merck: Grant/Research Support Cyra Christina Mehta, PhD, MSPH, Merck: Grant/Research Support Igho Ofotokun, MD, MSc, FIDSA, Merck: Grant/Research Support Maria L L. Alcaide, MD, Discidium Biosciences: Board Member|Gilead: Honoraria|Merk & Co: Honoraria|Senhwa Biosciences: Honoraria|Virology Education: Honoraria Jordan Lake, MD, Gilead Sciences: Grant/Research Support|Theratechnologies: Advisor/Consultant Phyllis C. Tien, MD, MSc, Merck: Grant/Research Support"}
+{"text": "Rheumatology key messageVEXAS syndrome is venulitis with severe thickening of the walls of large veins.Dear Editor, VEXAS  syndrome is an often fatal, X-linked disease characterized by haematological dysplasia overlapping with severe autoinflammatory manifestations. Acquired somatic mutations of the ubiquitin-like modifier activating enzyme (UBA1) gene impairs ubiquitylation, causing activation of the innate immune system and release of pro-inflammatory cytokines, TNF-\u03b1, IL-8, IL-6, IFN-\u03b3 and IFN-inducible protein-10 (IP-10) that culminate in severe systemic inflammation , which is independent of age, disease duration, history of thrombosis and major organ involvement, disease activity, acute phase response and corticosteroid treatment [s.d. 0.1)] but not significant with APS [0.49\u2009mm (s.d. 0.1)] [Rheumatology online).Previous studies have shown that venous inflammation causes VWT . Varicosm s.d. 0.4 vs 0.27reatment . Additiom s.d. 0.4 vs 0.27The pathogenesis of organ inflammation in VEXAS syndrome is not entirely understood. UBA1 is essential for the initiation of ubiquitylation, a widely used post-translational protein modification mechanism that regulates many biological processes, including intracellular signalling, proteasomal/lysosomal protein degradation and antimicrobial defence . Consequkead168_Supplementary_DataClick here for additional data file."}
+{"text": "Correction to: Clinical Proteomics (2022) 19:4810.1186/s12014-022-09385-7Unfortunately, in the original publication of the article, the following errors were identified after online publication of the article.The Additional file In Abstract, line 11, the text that reads as \u201cphospho-heavy-labeled-spiketide FAIMS Stepped-CV DDA (pHASED)\u201d should read as \u201chospho-heavy-labeled-spiketide FAIMS stepped-CV DDA (pHASED)\u201d.The original article has been corrected.Additional file 2: Table S1. SBDS heavy-labeled phosphorylated peptide standards. Table S2. Common and unique phosphoproteins identifed across all four CVs based on PSM acquisition. Table S3. High confdence modifcation sites identifed in LFQ (p < 0.01). Table S4. High confdence modifcation sites identifed in pHASED (p < 0.01). Table S5. Unique and common phosphoproteins identifed in LFQ and pHASED datasets. Table S6. Phosphorylated master protein kinases identifed in LFQ dataset (p < 0.01). Table S7. Phosphorylated master protein kinases identifed in pHASED dataset (p < 0.01). Table S8. FLT3-D835 mutations associated with resistance to tyrosine kinase FLT3 inhibitors. Table S9. Kinase-Substrate analysis of LFQ dataset for resistant cells in comparison to FLT3-ITD (log2 fold change\u00b10.5). Table S10. Kinase-Substrate analysis of pHASED data?set for resistant cells in comparison to FLT3-ITD (log2 fold change\u00b10.5). Table S11. Canonical pathways identifed as signifcantly associated with LFQ dataset for resistant cells in comparison to FLT3-ITD. Table S12. Canonical pathways identifed as signifcantly associated with pHASED dataset for resistant cells in comparison to FLT3-ITD. Table S13. Kinase activity inferred by KSEA analysis of phosphorylation changes in pHASED dataset  for resistant cells in comparison to FLT3-ITD. Table S14. Mutation-specifc response to sorafenib. IC50 compared to FLT3-ITD. Table S15. Bliss Synergy scores for sorafenib in combination with KU-60019 at diferent doses. Table S16. Unique ATM substrates identifed with increased phosphorylation (log2\u22650.5) in pHASED dataset for resistant cells in comparison to FLT3-ITD. Table S17. Vector mutations in FLT3 gene."}
+{"text": "Correction to: Sport Sciences for Health 10.1007/s11332-022-01012-0The reference no. 4 has been incorrectly published in the original publicaiton. The complete correct reference [4] is given below.Coronavirus disease (COVID-19): the need to maintain regular physical activity while taking precautions. J Sport Health Sci 9(2):103.4. Chen P et al (2020)"}
+{"text": "Korean J Women Health Nurs 2023;29(1):66-75.https://doi.org/10.4069/kjwhn.2023.03.15This corrigendum is for correcting a reference that was mistakenly reported in the above article.4. Pereira-Kotze C, Feeley A, Doherty T, Faber M. Maternity protection entitlements for non-standard workers in low-and-middle-income countries and potential implications for breastfeeding practices: a scoping review of research since 2000. Int Breastfeed J. 2023;18(1):9. https://doi.org/10.1186/s13006-023-00542-8The corrected reference is as below. The authors apologize for any inconvenience that this may have caused.4. Brandhagen M, Lissner L, Brantsaeter AL, Meltzer HM, H\u00e4ggkvist AP, Haugen M, et al. Breast-feeding in relation to weight retention up to 36 months postpartum in the Norwegian Mother and Child Cohort Study: modification by socio-economic status? Public Health Nutr. 2014;17(7):1514-1523. https://doi.org/10.1017/s1368980013001869."}
+{"text": "Cytomegalovirus (CMV) infection is associated with significant morbidity and mortality among kidney transplant recipients (KTR). Given toxicities and costs of antivirals, there is growing interest in assays that measure CMV-specific T-cell immunity (TCI), which may predict protection against clinical infection. The Viracor\u00ae CMV TCI Panel (TCIP) uses flow cytometry and intracellular cytokine staining to measure %CMV-specific CD4+ and CD8+ T-cells; it is the only commercially available TCI assay in the US. In this investigator-initiated study, we evaluated the performance of the TCIP in predicting clinically significant CMV events. To our knowledge, this is the first prospective blinded clinical study of the TCIP.We enrolled consecutive donor (D) or recipient (R) CMV seropositive KTR, who had TCIP testing monthly until either discontinuation of valganciclovir prophylaxis or the primary outcome. We also enrolled KTR with low-level untreated DNAemia, or after completion of valganciclovir treatment, to evaluate assay predictive values for progression or relapse of CMV infection, respectively. The primary outcome was CMV DNAemia (tested at the discretion of the treating provider for symptoms or laboratory findings), prompting treatment initiation. Secondary outcomes were any DNAemia and DNAemia >1000 IU/mL. Treating providers were unaware of TCIP results.p=0.024), and the CMV protection ROC AUC was significant . The positive predictive values of both CD4+ and CD8+ T-cell positivity >0.2% for CMV protection were 96.3% (95%CI 81-99.9%) for the primary outcome, 92.6% (95%CI 75.7-99.1%) for any DNAemia, and 100% (one-sided 95%CI 87.2%) for DNAemia >1000 IU/mL.We included 46 KTR. Median age was 50.5 (range 18-74) years. 73.6% identified as men, 10.8% Black, 8.7% Hispanic; 41.3% had CMV D+/R- status. KTR with CMV events had significantly lower %CMV-specific CD8+ T-cells  (5R25AI140490) Dimitrios Farmakiotis, M.D., Astellas: Grant/Research Support|Merck: Grant/Research Support|Viracor: Advisor/Consultant|Viracor: Grant/Research Support"}
+{"text": "QTRL.caas-4A.1 and QTRL.caas-4A.2), TRA (QTRA.caas-4A and QTRA.caas-4D), and NRT (QNRT.caas-5B and QNRT.caas-5D) were identified and each explaining 5.94%\u20139.47%, 6.85%\u20137.10%, and 5.91%\u201310.16% phenotypic variances, respectively. Among these, QTRL.caas-4A.1 and QTRA.caas-4A overlapped with previous reports, while QTRL.caas-4A.2, QTRA.caas-4D, QNRT.caas-5B, and QNRT.caas-5D were novel. The favorable alleles of QTRL.caas-4A.1, QTRA.caas-4A, and QTRA.caas-5B were contributed by Doumai, whereas the favorable alleles of QTRL.caas-4A.2, QTRA.caas-4D, and QTRA.caas-5D originated from Shi 4185. Additionally, two competitive allele-specific PCR (KASP) markers, Kasp_4A_RL (QTRA.caas-4A) and Kasp_5D_RT (QNRT.caas-5D), were developed and validated in 165 wheat accessions. This study provides new loci and available KASP markers, accelerating wheat breeding for higher yields.Identifying loci for root system architecture (RSA) traits and developing available markers are crucial for wheat breeding. In this study, RSA-related traits, including total root length (TRL), total root area (TRA), and number of root tips (NRT), were evaluated in the Doumai/Shi4185 recombinant inbred line (RIL) population under hydroponics. In addition, both the RILs and parents were genotyped using the wheat 90K single-nucleotide polymorphism (SNP) array. In total, two quantitative trait loci (QTLs) each for TRL ( Wheat production is influenced seriously by abiotic stresses. Breeding higher-yielding and more stable accessions under abiotic stress is a crucial objective in modern wheat breeding . Root syPrevious breeding programs predominantly focused on aboveground traits such as disease resistance, grain quality, and harvest index, while the application of RSA traits has been limited due to the complexity of phenotypic evaluation . PreviouIn this study, we conducted linkage mapping for RSA traits using the wheat 90K assays in a biparental recombinant inbred line (RIL) population derived from the Doumai/Shi 4185 cross. The primary objective of this study is to uncover the genetic basis of these traits and develop available KASP markers for improving wheat RSA.2:6 RILs derived from the Doumai/Shi 4185 were used for evaluating RSA-related traits. A hydroponic experiment was conducted with three replicates in a greenhouse. There were 20 seeds from each line that were surface sterilized with10% H2O2 for 20\u00a0min. Subsequently, the cleaned seeds were placed in Petri dishes with moist filter paper. When the coleoptiles reached about 2\u00a0cm in length, wheat seedlings were transferred to plastic trays (53 \u00d7 27\u00a0cm) containing Hoagland\u2019s nutrient solution. Plastic trays were kept in 25\u00b0C with a 16-h light and 8-h darkness cycle. After 3 weeks, roots were evaluated for RSA-related traits .Three RSA-related traits, namely, total root length (TRL), total root area (TRA), and number of root tips (NRT), were assessed using the WinRHIZO root analysis system (LA6400XL). The roots were arranged systematically in a dish and scanned using the Expression 11000XL. Subsequently, the images were imported into WinRHIZO software and analyzed using a fixed threshold parameter of 40. Each accession was scored on five plants for RSA traits, and means of three replicates were obtained. Basic statistical analyses and frequency distributions were conducted using SAS v9.3  by CapitalBio Corporation. SNPs with missing data >20% or minor allele frequency (MAF) <0.5 were filtered for further analysis. The filtered SNPs were then analyzed using the BIN function of IciMapping v4.2  and grouThe inclusive composite interval mapping (ICIM) method using IciMapping v4.1  was applQTRL.caas-4A.1, QTRA.caas-4A, and QNRT.caas-5D) were converted to KASPs (http://www.polymarker.info/) (SNPs flanking QTL with higher PVE (to KASPs  and desir.info/) Table S2r.info/) .https://wheat.pw.usda.gov/GG3/). Genes, excluding hypothetical proteins, transposon proteins, and retrotransposon proteins with SNPs in the coding region, were considered as candidate genes. Quantitative real-time PCR (qRT-PCR) was conducted to test expression differences of the candidate genes between Doumai and Shi 4185. The roots were sampled for RNA extraction after phenotyping. RNA was extracted using the TRIzol method, and the cDNA was synthesized with the HiScript II 1st Strand cDNA Synthesis Kit. Primers were designed using the Primer Premier V5.0. PCR was conducted with a mixture of 20 \u03bcl, including 2 \u03bcl cDNA, 10 \u03bcl ChamQ Universal SYBR qPCR Master Mix, and 0.4 \u03bcl of each primer. qRT-PCR was conducted in the ABI StepOnePlus Real-Time PCR System with Tower, and the gene expression level was analyzed by the 2\u2013\u0394\u0394CT method. All assays were performed in two biological replicates and three technical replicates. TaActin1 was used as the internal control to normalize the expression levels of different samples.To identify candidate genes involved in the QTL for RSA-related traits detected in the Doumai/Shi 4185 RIL population, the genes located in the LD block region around the peak SNP  of each QTL were extracted from the wheat IWGSC v1.1 annotation (2 (range: 6.9 cm2\u201315.2 cm2), and 328.2 root tips (range: 94.7\u2013772.0). The standard deviation and coefficient of variation for TRL, TRA, and NRT were 18.1\u00a0cm (24.2%), 1.7 cm2 (16.4%), and 126.4 (38.5%), respectively. A significant correlation was observed between TRL, TRA, and NRT, with the correlation coefficient of 0.603 (P < 0.05) between TRL and TRA, 0.371 (P < 0.05) between TRL and NRT, and 0.312 (P < 0.05) between TRA and NRT.All three RSA-related traits exhibited continuous and significantly wide variation across the 262 RILs (QTRL.caas-4A.1 (BobWhite_c20306_147-Tdurum_contig54973_1510) and QTRL.caas-4A.2 (BS00059454_51-Kukri_c19883_816), respectively. These QTLs explained 5.94% (additive effect: 5.17) and 9.45% (additive effect: \u22127.12) of the total phenotypic variances  and QTRL.caas-4D (Ex_c6665_1067-wsnp_J-D_rep_c51623_35119179), respectively, explaining 7.10% (additive effect: 0.40) and 6.85% (additive effect: \u22120.39) of the total phenotypic variance. The favorable allele of QTRA.caas-4A TRA was contributed by Doumai, while the favorable allele of QTRA.caas-4D was originated from Shi4185, respectively. Two QTLs for NRT were detected on chromosomes 5B and 5D and named as QNRT.caas-5B (wsnp_Ex_rep_c67320_65870601-Tdurum_contig10191_996) and QNRT.caas-5D (IAAV6218-Kukri_c46526_103), respectively, explaining 5.91% (additive effect: 26.7) and 10.16% (additive effect: 39.1) of the total phenotypic variances. The favorable alleles of QNRT.caas-5B and QNRT.caas-5D were contributed by Doumai and Shi 4185, respectively  with higher phenotypic effects were used to develop KASP markers. Although attempts were made to develop a KASP marker for QTRA.caas-4A, it was unable to effectively distinguish between the two parental genotypes in the RIL population. Therefore, the marker did not yield conclusive results. Consequently, two KASP markers, Kasp_4A_RL  and Kasp_5D_RT , were successfully developed based on the tightly linked SNP markers  exhibited higher TRL compared to the unfavorable allele  at the P = 0.05 level  showed higher NRT than unfavorable allele  at the P = 0.05 level  encoded an ethylene-regulated nuclear protein (ERT2)-like protein; TraesCS4A01G436700 (QTRL.caas-4A.1) encoded a calcium-binding protein kinase (CDPK)-related kinase; TraesCS4A01G456300 (QTRL.caas-4A.1) encoded a cellulose synthase-like protein; TraesCS4A01G469400 (QTRL.caas-4A.2) encoded an F-box protein; and TraesCS4D01G092000 (QTRA.caas-4D) encoded an ABC transporter, whereas both the TraesCS5D01G378400 and TraesCS5D01G380200 for QNRT.caas-5D encoded E3 ubiquitin-protein ligase. The expressions of the seven candidate genes in Doumai and Shi 4185 were detected using qRT-PCR. Of these, TraesCS4A01G469400, TraesCS4D01G092000, and TraesCS5D01G380200 showed no significant differences between the parents, whereas TraesCS4A01G296500, TraesCS4A01G436700, TraesCS4A01G456300, and TraesCS5D01G378400 showed more than 1.4\u20133.9-fold higher expression in Doumai compared to Shi4185 , TRA (QTRA.caas-4A and QTRA.caas-4D), and NRT (QTRA.caas-5B and QTRA.caas-5D) TRA. Each of these QTLs accounted for 5.94%\u20139.47%, 6.85%\u20137.10%, and 5.91%\u201310.16% of the phenotypic variance, respectively.Optimization of crop root systems has long been proposed. However, genetic improvement of crop roots has been rarely attempted . A compr9 RIL lines and have identified 19 QTLs mainly distributed on chromosomes 1A, 2B, 2D, 3A, 3B, 3D, 5A, and 5D. Of these, the 5D loci (415.8 Mb\u2013457.6 Mb) overlapped with theQNRT.caas-5D (449.4 Mb\u2013454.1 Mb) identified in this study. IWB21309, 17.01 Mb) and 5B  for RSA traits were also different with the loci identified in this study . Furthermore, MQTL4A.1 (651.78 Mb\u2013705.73 Mb) and MQTL4A.6 (600.04 Mb\u2013691.14 Mb) significantly associated with root length, root surface, and root tips overlapped with QTRL.caas-4A.1 (702.3 Mb\u2013721.3 Mb) and QTRA.caas-4A (594.2 Mb\u2013602.9 Mb). Rht2/Rht10 19.18 Mb, SVP3-4D/BM1-4D 469.46 Mb, Vrn2-4D/ZCCT1-4D 509.43 Mb), 5B , and 5D   are different from the reported wheat genes associated with plant height and vernalization. Therefore, they represent novel genes associated with RSA-related traits. Above all, compared with the previous results and meta-analysis, QTRL.caas-4A.2, QTRL.caas-4D, QNRT.caas-5B, and QNRT.caas-5D were novel.0.00 Mb) . Based oQTRL.caas-4A.1 (702.3 Mb\u2013721.3 Mb) co-located influencing QTL clusters related to thousand kernel weight   co-located with regions affecting QTL clusters related to yield, including SN, KNS, TKW, FLW, and KL   . Additio65.6 Mb) . These rTraesCS4A01G436700 of QTRL.caas-4A.1 encodes the CDPK-related kinase. In plants, CDPKs play a critical role in various signaling pathways and are pivotal in root growth and development -like protein. Ethylene, a simple small molecule, plays a vital role in signal transduction, cell differentiation, division, and elongation (TraesCS5D01G380200, linked to QNRT.caas-5D, encodes an E3 ubiquitin-protein ligase, which plays essential roles in plant development (Four genes involved in the biological metabolism of plant hormones, cellulose, and the ubiquitin pathway were identified as high confidence candidate genes. Among these, elopment . Silencil length . TRA Anotivities . TraesCSongation . Ethylenongation . Additioelopment  and is celopment .Kasp_4A_RL and Kasp_5D_RT were successfully developed based on tightly linked SNP markers, proving to be valuable tools for MAS in breeding programs. Additionally, accessions with more favorable alleles, superior RSA traits and appropriate agronomic traits, such as Yumai 18, Jinhe 9123, Yumai 34, Bainong 3217, Lumai 6, Jinmai 61, Lumai 5, Lumai 23, and Yumai 57 are recommended as parental lines for improvement of RSA traits.Although conventional breeding has contributed to the enhancement of root system, the selection process is time-consuming and less efficient due to the challenges in field measurements of RSA traits . KASP ofIn this study, we identified six QTLs associated with wheat RSA traits and successfully developed two KASP markers that can be utilized in wheat breeding programs aimed at achieving higher and more stable yields. This research serves as a foundational step towards gene cloning and enhancement of wheat root systems.All datasets generated for this study are included in the article or YJ: Data curation, Funding acquisition, Writing \u2013 original draft. YW: Data curation, Software, Validation, Writing \u2013 original draft, Writing \u2013 review & editing. JL: Software, Validation, Writing \u2013 review & editing. FW: Formal Analysis, Resources, Software, Investigation, Writing \u2013 review & editing. XQ: Investigation, Project administration, Writing \u2013 review & editing. PL: Funding acquisition, Supervision, Validation, Writing \u2013 original draft."}
+{"text": "PrEP with T/C can prevent COVID-19 hospitalization and death in IC patients (pts) up to 6 months after injection. However, in the USA, authorization of T/C PrEP was paused in Jan 2023 due to loss of in vitro activity of T/C against dominant circulating SARS-CoV-2 variants, although loss of clinical efficacy is unclear. We investigated in vivo mechanisms of viral breakthrough in hospitalized IC pts with vs without prior T/C exposure.We analyzed remnant clinical SARS-CoV-2 PCR-positive swabs and sera from IC pts hospitalized at UPMC. SARS-CoV-2 variants and mutants were determined by whole genome sequencing and anti-RBD IgG levels by an enzyme immunoassay.Table 1). Median age was 67 yrs; 49% were male. IC conditions included organ transplant (23%) and hematologic cancer (32%) (Table 2). In IC patients with sequenced swabs, 21% received T/C (Table 3). Variant frequency mirrored national trends (Table 3). BA.5, XBB.1, and BF.7 were less common in T/C vs non-T/C pts . BA.2 and BQ.1 were more common in T/C vs non-T/C pts . The R346T and K444T/R/N mutations were more common in T/C vs non-T/C pts: 54% vs 41% and 37% vs 22% (Table 3). Anti-RBD IgG titers from 56% pts at the time of infection were higher in T/C vs non-T/C pts . COVID-19 mortality was numerically lower in T/C vs non-T/C pts . Mortality differences were consistent across variant epochs (Table 1).From Mar 28, 2022, to Mar 3, 2023, 72% (174/243) of swabs were successfully sequenced from 170 pts : Advisor/Consultant|LabSimply: Advisor/Consultant|Merck: Advisor/Consultant|Shionogi: Advisor/Consultant|Shionogi: Honoraria John W. Mellors, MD, AstraZeneca: Grant/Research Support|Gilead Sciences: Grant/Research Support"}
+{"text": "The identification and characterization of plasma proteins in drug resistant and drug sensitive in HIV-1 infected/AIDS patients were carried out using the SWATH-MS protocol. In total, 204 proteins were identified and quantified, 57 proteinswere differentially expressed, out of which 25 proteins were down regulated and 32 proteins were up regulated in drug resistant patients. Six proteins such as complement C4-A, immunoglobulin heavy variable 1-2, carboxylic ester hydrolase, fibulin-1,immunoglobulin lambda constant7, secreted phosphoprotein 24 were differentially expressed in individuals with drug resistant HIV as compared to individuals with drug sensitive HIV. Gene ontology of 57 differentially expressed proteins was analysedand documented. The analysis of HIV-1 infected human plasma is an attractive medium with which differentially expressed proteins could be identified in certain disease states. Proteomic techniques are widely used globally to identify differentiallyexpressed plasma proteins in response to HIV-1 infection. Sequential window acquisition of all theoretical mass spectra (SWATH-MS) is a novel technique conceptualized with the protein library and the individual protein was identified andquantified for biomarker discovery . TherefoAmmonium formate, formic acid, dithiothreitol (DTT) and iodoacetamide (IAA) were procured from Sigma, USA. Modified trypsin (sequencing grade) was procured from Promega, USA. Polysulfoethyl SCX cartridge  withcartridge holder was procured from Sciex, USA. Acetonitrile, Liquid phase-chromatography-mass spectroscopy grade water, was procured from Biosolve . Analytical grade chemicals were used for this study.The study focused on patients administered six first-line ART such as ZLE, ZLN, TLE, TLN, SLE and SLN patients and they were enrolled in the anti-retroviral therapy centre, Sarojini Naidu Medical College, Agra, India from December 2009 to November2016 as per the treatment guidelines provided by NACO, Govt. of India and the ethical guidelines formulated by the ICMR, Government of India. The age of the six patients varied from 32 years to 43 years. A patient information leaflet was used for datacollection  hsa04610: Complement and coagulation cascades - Homo sapiens (human)[2] hsa05322: Systemic lupus erythematosus - Homo sapiens (human)[3] hsa05133: Pertussis - Homo sapiens (human)[4] hsa05150: Staphylococcus aureus infection - Homo sapiens (human)[1] H00080: Systemic lupus erythematosus[2] H00102: Classic complement pathway component defects[3] H01649: SchizophreniaThe six-drug respondent and drug resistant patients were in first-line ART such as ZLE, ZLN, SLN, TLE, and TLN during their course of treatment in this period. The drug interactions with these proteins were analysed (www.dgidb.org). A totalThe serial number of the down regulated and up regulated proteins in the heat map is represented in the Gene symbol: 1.C4A 2. IGHM 3. APOA4 4.HBB, 5. SERPING1 6. PZP 7. IGHG2 8.CD5L 9.HPR 10. HBA1 11. JCHAIN 12. APOL1 13. IGHV1-2 14. IGHV3-9 15.IGHV5-51 16. C4B 17. BCHE 18. LPA 19. IGLV6-57 20. SERPINA1 21. IGKV2-24 22. IGLV3-10 23. FAT-1 24.BIN3 25. PROC 26. FN1 27. IGHA1 28. GSN 29. AFM 30. TTR 31. IGLV3-21 32. KRT1 33. IGLV3-19 34. IGKV3-15 35. C4BPB 36. FBLN1 37. IGKV1-5 38. F5 39. IGKV3-11 40.IGLV7-46 41. VNN1 42. IGLC7 43. IGHA2 44. IGHV3-13 45. IGHV1OR15-1 46. SPP2 47. CDKL3 48. IGLV10-54 49. CDC42BPA 50. SP5 51. BEX5 52. PEPD 53. IGHV6-1 54. IGHV1-45 55. CDH5 56. 3 57. SERPINA 10. The heat map of differentially expressed proteins was createdby the online tool (http://www.heatmapper.ca/expression).The gene list for a category of the molecular function of the differential expressed proteins are presented in the bar diagram. The number of genes involved in the different functional role is presented in serial number in the bar diagram1. Binding (GO:0005488), 2. Catalytic activity (GO:0003824), 3. Molecular function regulator (GO:0098772), 4. Molecular transducer activity (GO:0060089), 5. Transcription regulator activity (GO:0140110), 6. Transporter activity (GO: 0005215).The molecular function of the differential expressed proteins was created by the online tool PANTHER- gene analyst ( http://www.pantherdb.org ).The red colour dots represent the down-regulated proteins and black colour dots represent the up-regulated proteins . The volcano plot was created by Graph Pad Prism 8, USA( https://www.graphpad.com /scientific-software/prism/).The details description of the protein-protein interaction network nodes represents proteins: Splice isoforms or post translational modifications are collapsed i.e. Each node represents all the proteins produced by a single protein-coding gene locus.The edges represent protein-protein associations: Network status: number of nodes: 35, number of edges:80, average node degree: 4.57, average local clustering coefficient:0.453, expected number of edges: 7, PPI enrichment p-value:< 1.0e-16. Theprotein-protein interaction network was created by using the online tool STRING Version 11.0 (https://string-db.org).The differential expression and molecular function of the Serotransferrin and Apolipoprotein A1 was identified in HIV-1 patients treated with first line ART . Now, thWe are report that proteins identified in drug resistant and drug sensitive HIV-1 infected patients who are taking first line ART over 6 years by SWATH-MS. Plasma proteins such as complement C4-A, immunoglobulin heavy variable 1-2, carboxylicester hydrolase, fibulin-1, immunoglobulin lambda constant7, secreted phosphor protein 24 were differentially expressed in individual drug resistant HIV-1 subtype C patients. The role of these proteins is reported using gene ontology analysis. Thisstudy would be helpful for further implementation in the search for biomarker discovery in HIV-1/AIDS patients taking first line ART.The project \"Characterization of drug resistant HIV-1 mutants of Agra region, India by genomic and proteomic approaches\" funded for senior research fellowship of Sushanta Kumar Barik (File No. 80/990/2015-ECD-I). M. M. Alam, JALMA, ICMR and Rakesh KumarMishra, ART centre, S.N Medical College are acknowledged for providing technical help in the collection of blood samples."}
+{"text": "Kachinia Tong & Li, 2018 and Promolotra Tong & Li, 2020 are recorded from China for the first time. Two new species, Kachinialonglingsp. nov. (\u2642\u2640) and Promolotralushuisp. nov. (\u2642\u2640) are described. Descriptions, diagnoses, photographs and keys to Kachinia and Promolotra species are provided.The genera  Oonopidae Simon, 1890 includes 1893 extant species in 115 genera worldwide, mainly distributed in tropical regions  were taken under high vacuum with a Hitachi S-4800 after critical-point drying and gold-palladium coating. All measurements were taken using an Olympus BX51 compound microscope and are in millimeters. The terminology used in the text and figures follows SYNU) in Shenyang, China (curator: Yanfeng Tong).The specimens were examined using a Leica M205C stereomicroscope. Details were studied under an Olympus BX51 compound microscope. Photos were made with a Canon EOS 750D zoom digital camera (18 megapixels) mounted on an Olympus BX51 compound microscope. Endogyne were cleared in lactic acid. Scanning electron microscope images , Myanmar.Taxon classificationAnimaliaAraneaeOonopidae\ufeffTong & Zhangsp. nov.83FC57DD-B207-51D0-B931-DD9CFD1538A0https://zoobank.org/651BE492-19F3-480A-9EB7-CF6675FCFF21Holotype \u2642 (SYNU-640), China, Yunnan Prov., Baoshan City, Longling Co., Longxin Town, Xiaoheishan Nature Reserve, 16.02.2011, Z. Li & L. Wang; Paratypes: 1\u2640 (SYNU-641), 1\u2640 (SYNU-642), 4 \u2640 (SYNU-643-646), same data as for the holotype.K.mahmolae and K.putao by the strongly elevated epigastric region of the male . Habitus as in Fig. Female. As in male except as noted. Habitus as in Fig. The specific name is a noun in apposition taken from the type locality.Known only from the type locality.Taxon classificationAnimaliaAraneaeOonopidae\ufeffGenusTong & Li, 2020956B765C-34F2-50EC-8981-3DD686DC17FAPromolotrashankhaung Tong & Li, 2020 from Myanmar.Oonopinae Simon, 1890. According to Molotra Ubick & Griswold, 2011 by the heavily sclerotized dorsal and ventral abdominal scuta, the long spines on legs I and II, and the embolar region.The genus belongs to the subfamily Promolotrahponkanrazi Tong & Li, 2020, P.lushui sp. nov., P.shankhaung Tong & Li, 2020.China (Yunnan), Myanmar.Taxon classificationAnimaliaAraneaeOonopidae\ufeffTong & Zhangsp. nov.0111FE05-4B7A-59C2-8DCA-D3F41A907301https://zoobank.org/E02029CB-E8DA-471D-9B13-A5D39E3E20C0Holotype \u2642 (SYNU-647), China: Yunnan Prov., Lushui City, Pianma Town, 3.03.2011, Z. Li & L. Wang leg.; Paratypes: 2 \u2640 (SYNU-648-649), 3 \u2642 (SYNU-650-652), same data as for the holotype.The new species can be distinguished from congeners by the blunt end of the cymbiobulb . Habitus as in Fig. Female. As in male except as noted. Habitus as in Fig. The specific name is a noun in apposition taken from the type locality.Known only from the type locality."}
+{"text": "Nirmatrelvir/ritonavir administered twice daily for 5 days (5D) (within 5 days of symptom onset) resulted in a clinically and statistically significant (6% absolute and 86% relative) risk reduction in COVID-19 related hospitalization or all cause death through Day 28 [1]. COVID-19 rebound regardless of treatment has been reported . Here, an integrated analysis of EPIC-HR/SR  virology data was conducted to evaluate clinical resistance to nirm/r and if it is associated with viral load rebound (VLR) or progression to severe COVID-19 .10 copies/mL. The amino acid (AA) sequence was compared to Wuhan-Hu-1 [7] and Mpro substitutions were called if AAFREQ \u2265 10%. Emergent substitutions (ES) were called if observed post-baseline only and called treatment emergent substitutions (TES) if the ES was \u2265 2.5-fold more frequent, and had \u22653 additional occurrences, in nirm/r than placebo (PBO). Viral RNA load rebound (VLR) was defined as VL increase at D10 or D14 by \u2265 0.5 log10 copies/mL from D5 and resulting in a VL \u2265 3.0 log10 copies/mL.Next generation sequencing analysis was performed from nasal swabs  with a viral RNA level \u22653.0 logpro ES did not differ between treatment arms. Among those with sequence data available , nirm/r Mpro TES were observed in: T98I/del(n=3), E166V (n=3), and W207L/del (n=3) and within the Mpro cleavage sites: A5328S/V(n=5) and S6799A/P/Y (n=4). None of the TES were associated with progression to severe COVID-19. VLR was observed in  with Mpro TES observed in few VLR patients (3.4% in nirm/r and 0% in PBO). Among the Mpro TES identified, E166V is an in vitro resistance mutation . Figure 1 shows the VL trajectories of the 3 patients with E166V, one patient experienced VLR at D10, all effectively controlled the virus by D14 and did not experience severe COVID-19.In EPIC-HR/SR, the primary SARS-CoV-2 variant was Delta (91%) followed by Omicron (7.4%). The percentage of patients with Min vitro resistance substitution emerged in few patients, but was not associated severe COVID-19. Additionally, these data show VLR is not a result of treatment resistance to nirm/r driven by TES or any Mpro substitutions.In EPIC-HR/SR, an Mary Lynn Baniecki, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Shunjie Guan, PhD in Statistics, AbbVie: Stocks/Bonds|Pfizer: Stocks/Bonds Zhenyu Wang, Ph.D., Alkermes: Stocks/Bonds|Moderna: Stocks/Bonds|Pfizer: Stocks/Bonds Yan Chen, Ph. D, Pfizer, Inc: Stocks/Bonds Weihang Bao, PhD, Pfizer, Inc.: Employee of Pfizer|Pfizer, Inc.: Stocks/Bonds Wen He, PhD, Pfizer Inc.: Stocks/Bonds Elizabeth Dushin, PhD, Pfizer Inc.: Stocks/Bonds Craig Hyde, PhD, Pfizer: Employed by Pfizer|Pfizer: Stocks/Bonds Yuao Zhu, PhD, Pfizer, Inc.: Stocks/Bonds Rhonda D. Cardin, PhD, Pfizer, Inc: Stocks/Bonds Jennifer Hammond, PhD, Pfizer: Employee|Pfizer: Stocks/Bonds Sandeep Menon, PhD, Pfizer: Stocks/Bonds Charlotte Allerton, PhD, Pfizer: Stocks/Bonds Holly Soares, PhD, Pfizer: Stocks/Bonds"}
+{"text": "Enterococcus faecalis.The whole-genome sequence of strain K-4, isolated from grass silage in Thailand, which constitutes a chromosome and two plasmids, is 2,914,933\u2009bp long, has a GC content of 37.5%, and contains 2,734 predicted protein-coding genes. Average nucleotide identity based on BLAST+ (ANIb) and digital DNA-DNA hybridization (dDDH) values indicated that the strain K-4 was closely related to  Enterococcus comprises Gram-positive, non-spore-forming facultative anaerobic, spherical, or ovoid cells with low genomic GC content   were assembled de novo using Unicycler v0.4.8 (https://github.com/rrwick/Unicycler) (dnaA for the chromosome. The genome sequence was annotated with DFAST v1.2.6 (https://dfast.nig.ac.jp/)  and the Genome-to-Genome Distance Calculator 3.0 (http://ggdc.dsmz.de/ggdc.php)  and decoded on the Illumina MiSeq system. Overall, 1,192,876 reads with 299,930,918 bases were decoded with an average read length of 251.4\u2009bp and spot length of 602\u2009bp with 2\u2009\u00d7\u2009301-bp paired-ends reads. Low-quality bases  were trimmed, and short reads (<25\u2009bp) were removed using Platanus trim v1.0.7 (us_trim) . Data reus_trim)   . Averagegdc.php)  using reE. faecalis ATCC 19433T  with a sequence depth of ~88. Overall, 2,734 coding regions, 60 tRNAs, 12 rRNAs, and 1 CRISPR region were annotated. The DNA sequences of two plasmids, namely, pEK4S  and pEK4L , were also assembled and compared with their partial sequences using Blast  and DRX314299 (Illumina).Strain K-4 genome sequence and annotation data were deposited in DDBJ/GenBank under BioProject number"}
+{"text": "Professional and regulatory agency guidelines recommend that fluoroquinolones not be used as first-line therapy to treat uncomplicated UTI. Increasingly, \u03b2-lactams are being used to treat UTI despite limited evidence supporting their effectiveness. The study aim was to compare the efficacy of \u03b2-lactams and other commonly prescribed antibiotics for UTI to fluoroquinolones.< 30d, co-diagnosis where antibiotics were indicated, hospital discharge < 7d, scheduled urological procedure < 7d, pregnancy. Outcome: UTI related admission or outpatient return visit within 3-30d. Demographic, co-morbidity, vitals, laboratory, and prescription data were extracted. Propensity scores were used to balance treatments and GEE models were used to estimate outcomes.A multi-centered retrospective cohort study of outpatient visits with a UTI diagnosis between 2019-2021 was developed. Inclusion criteria: In-person visit in the Emergency Department or Primary/Urgent Care with ICD-10 documented UTI diagnosis and oral antibiotic prescription filled within 0-2 d. Exclusion criteria: Prior UTI There were 73,334 visits in 130 VA medical centers. Mean (S.D.) cohort age was 66.0 (15.8) y, 22.9% were female. Antibiotics prescribed N (%): \u03b2-lactams  27,385 (37.3%), nitrofurantoin 15,509 (21.1%), trimethoprim / sulfamethoxazole (TMP/SMX) 15,453 (21.1%), and fluoroquinolones 14,997 (20.4%). The overall 30-day clinical failure rate was 10,525 (14.4%).Comparative Effectiveness of Oral Antibiotics for Outpatient UTIsCommonly prescribed antibiotics for outpatient UTI were associated with a modest increase in 30-day clinical failure relative to fluoroquinolones. Effectiveness differences were most pronounced in male patients. Clinicians should carefully differentiate between complicated and uncomplicated UTI in males and consider preferentially prescribing fluoroquinolones.Karl Madaras-Kelly, PharmD, MPH, BioMerioux: Grant/Research Support"}
+{"text": "Vaccines against SARS-CoV-2 (COVID-19) proved beneficial for COVID-19 disease attenuation and preventing virus spreading. Cumulative reports of the rarity of antineutrophil cytoplasmic autoantibodies (ANCA)-associated vasculitis (AAV) raise concerns about its relationship with COVID-19 vaccination. Several case reports described ANCA-associated pauci-immune glomerulonephritis (ANCA-GN) following COVID-19 vaccination with some uniqueness. We systematically reviewed COVID-19 vaccine-induced ANCA-GN from PubMed, SCOPUS, and Cochrane library databases until 1 January 2023 according to PRISMA guidelines and presented our three cases. Twenty-six cases from 25 articles, including our 3 cases, were analyzed. Most cases were diagnosed following the second dose of the COVID-19 vaccine (59%) with a median (IQR) interval onset of 14 (16) days. The highest prevalence was related to the mRNA-type vaccine. Anti-myeloperoxidase (MPO) ANCA was far more common than the other ANCAs, with various positive autoantibodies. Fourteen cases  had extra-kidney AAV manifestation. Although severe kidney injury was observed in 10/29 (34%), remission was achieved in 89% (25/28) with no death. The mechanisms of the vaccine-inducing ANCA-GN were postulated here. Since ANCA-GN after the COVID-19 vaccine was rare, the benefit of the COVID-19 vaccine could outweigh the risk of ANCA-GN side effects in the pandemic era. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the ongoing global pandemic of coronavirus disease 2019 (COVID-19), with a cumulative confirmed case of 765 million, including a 1% death rate . In addiThe American College of Rheumatology/European Alliance of Associations for Rheumatology (ACR/EULAR) 2022 classified AAV into granulomatous polyangiitis (GPA), eosinophilic granulomatous polyangiitis (EGPA), microscopic polyangiitis (MPA), and renal-limited vasculitis (RLV) ,13. HoweTwo independent authors conducted systematic literature by searching relevant full-text articles in the PubMed, SCOPUS, and Cochrane library databases up to 1 January 2023. Keywords were  AND . The eligible articles included the full-text report in the English language of biopsy-proven ANCA-GN with pauci-immune deposit by immunofluorescence (IF) study (defined as negative or weak [\u22641+] staining of Ig and complement) or by electron microscopy (EM) (defined as no or faint electron-dense deposit [EDDs]) after COVID-19 vaccination. Patients\u2019 age, vaccine types, and the disease onset from the vaccination were not restricted. The exclusion criteria were AAV without GN, ANCA-GN with a non-available IF/EM report, and AAV presenting or having disease flare before the onset of vaccination. We also included our 3 GN-ANCA cases diagnosed and treated at King Chulalongkorn Memorial Hospital, Bangkok, Thailand, from January 2022 to December 2022. Data extraction was anonymized under the General Data Protection Regulation , and infData included patient demographics, vaccine types, vaccination numbers, the interval between the last vaccination and onset of AAV, the clinical manifestation of ANCA-GN and AAV, serum creatinine (Scr) at baseline and peak, serologies , kidney pathology finding, treatment, and outcomes after treatment.Severe acute kidney injury (AKI) was defined as dialysis requirement or peak Scr > 5.7 mg/dL. Outcomes were classified into remission with relapse, remission without relapse, and non-remission . Remission of ANCA-GN/AAV was defined according to the 2012 KDIGO Guidelines for Management of Glomerular Diseases  as the at-test. The continuous data were illustrated by using mean \u00b1 standard deviation (SD) for normal distribution data and median (interquartile range [IQR]) for non-normal distribution data. The categorical data were described as a ratio. Finally, the normally distributed variables were compared by A total of 2956 relevant records were initially obtained from the database search. After removing 312 duplicated records, 2644 record titles and abstracts of articles were first screened according to the inclusion criteria. 68 articles were eligible for the second screening; 43 articles (154 cases) were excluded ; 26 cases from 25 articles, including our 3 cases were recruited for the systematic review . The detThe median (IQR) age was 70 (22) years, with female predominance (59%). Most cases presented ANCA-GN after receiving the second vaccination (59%) with a median interval gap of 14 (11\u201327) days. The mRNA COVID-19 vaccine was the most common (55%), followed by the viral vector vaccine (31%) and inactivated COVID-19 vaccine (14%). The top-three most common presentations were AKI, abnormal urine sediments, and constitutional symptoms. Extra-kidney manifestations included pulmonary involvement  , neuromuscular involvement  , otologic/optic involvement , and Wallerian degeneration . Severe AKI was observed in 34% (10/29). MPO-ANCA/pANCA was positive in 79% (23/29). A discrepancy of ANCA serologies was observed in one case . A dual-positive ANCA (pANCA and cANCA) was detected in 3% (1/29). Autoantibodies were positive in 52% (13/25 cases), including ANA , Coombs antibody , cryoglobulin , and RF . The 2021 KDIGO Guidelines for Management of Glomerular Diseases  were adoWe described three cases of ANCA-GN following viral vectors (case #1) and mRNA (case #2 and #3) COVID-19 vaccines. Only one patient (case #2) was classified as microscopic polyangiitis (MPA) by ACR/EULAR 2022 . Case #19/L with a neutrophil predominance (83%), Scr 3.5 mg/dL, and albumin 2.5 g/dL. Urinalysis demonstrated similar profiles with dysmorphic erythrocytes. The urine protein-creatinine ratio (UPCR) was 7.5 g/g creatinine. pANCA was positive with negative MPO-ANCA and PR3-ANCA. Serologies were positive for ANA , RF 30 (0\u201320) IU/mL, and Coombs antibody (1+), while others were negative . Complement levels were within normal values. The kidney biopsy disclosed 24 glomeruli with 7 global sclerosis (GS) and 7 cellular crescents. Fibrinoid necrosis of arteries and severe tubulointerstitial inflammation were observed. Interstitial fibrosis and tubular atrophy involved 30% of cortical tissue. IF revealed IgG (1+), C3 (1+), Kappa (1+), and Lambda (1+). Focal trivial subepithelial and intramembranous EDDs with diffuse thickening of GBM were observed on EM. Hence, ANCA-GN without extra-kidney organ involvement was diagnosed. Three-day of 1 gm/day pulse methylprednisolone and 500 mg pulse cyclophosphamide was started. Unfortunately, her kidney function remained deteriorated, requiring maintenance hemodialysis.A 76-year-old female with type 2 diabetes mellitus, hypertension, and dyslipidemia presented with leg edema for 11 days prior to admission. She received the second COVID-19 vaccine (AZD1222) 3 months ago. A few weeks later, she developed chronic fever, weight loss, and productive cough. Several blood cultures were performed, yielding negative results. Non-contrast chest computed tomography (CT) revealed unusual interstitial pneumonia (UIP). Sputum cultures depicted normal flora. She denied a history of SARS-CoV-2 infection and had negative PCR for COVID-19. Her Scr was 1.5 mg/dL over the past year. Three-week before admission, her Scr increased to 1.7 mg/dL with 2+ proteinuria and erythrocyte of 10\u201320/HPF. A week later, she developed leg edema and foamy urine. Physical examination (PE) revealed temperature 38.9 \u00b0C, respiratory rate (RR) 20/min, blood pressure (BP) 165/91 mmHg, and bilateral 3+ leg pitting edema. Blood chemistries revealed hemoglobin 7.6 g/L, leukocyte count of 11.9 \u00d7 10Case #29/L with neutrophils predominance (83%), and platelet counts 386 \u00d7 109/L. Urinalysis depicted trace protein, erythrocytes 3\u20135 cells/HPF, and UPCR 0.5 g/g creatinine. pANCA and MPO-ANCA were positive, with negative PR3-ANCA and cANCA. Serologies were positive, including ANA , cryoglobulin, and 1+ direct antiglobulin test, while the others were negative . Complement levels were within normal values. Serum protein electrophoresis and immunofixation demonstrated polyclonal gammopathy. No structural kidney abnormalities were detected by ultrasonography. The kidney biopsy disclosed 13 glomeruli with 2 GS and 10 cellular crescents. Fibrinoid necrosis was found on the vasculitic medium-sized artery. Mild tubulointerstitial fibrosis was observed. IF revealed non-specific staining for all immunoreactants. Hence, AAV with ANCA-GN was diagnosed. She received plasma exchange \u00d77 times, pulse methylprednisolone, and oral cyclophosphamide. One year later, she remained well without dialysis requirements and active AAV manifestation. Her serum creatinine 2.3 mg/dL and negative erythrocyturia.A 69-year-old female presented with AKI while investigating anorexia, weight loss, and fatigue. She had hypertension for 2 decades and received 2 doses of AZD1222 and a COVID-19 vaccine booster with BNT162b2 (3 months ago). One month later, she developed anorexia, fatigue, a weight loss of 8 kg within 2 months, and established mixed conductive and sensorineural hearing loss. Chest X-ray and contrast computerized tomography (CT) revealed a subcentimeter pulmonary nodule. Two weeks after CT, her Scr increased from 0.7 to 2.4 mg/dL, and contrast-associated AKI was suspected. However, despite receiving an intravenous fluid infusion, she had a steadily increasing Scr. Her urinalysis revealed 1+ protein, leukocyte 50\u2013100/HPF, and erythrocyte 3\u20135/HPF. Ciprofloxacin was prescribed on suspicion of having a urinary tract infection. PE revealed temperature 38.0 \u00b0C, BP 120/60 mmHg, marked pale conjunctivae, and no leg edema. Blood chemistries were notable for Scr 7.1 mg/dL, albumin 2.5 g/dL, hemoglobin 6.6 g/dL, leukocyte counts 12.1 \u00d7 10Case #39/L with neutrophils predominance (80%), and platelet counts 209 \u00d7 109/L. Urinalysis depicted 2+ protein, leukocytes 30\u201350/HPF, erythrocytes 1\u20132/HPF, and UPCR 1.2 g/g creatinine. cANCA and MPO-ANCA were positive, with negative PR3-ANCA. Positive direct antiglobulin test at 1+ was detected, while others were negative . Complement levels were normal. Serum protein electrophoresis and immunofixation demonstrated polyclonal gammopathy. No structural kidney abnormalities were detected by ultrasonography. The kidney biopsy disclosed 20 glomeruli with 2 GS and 7 cellular crescents. Fibrinoid necrosis was noted in 6 glomeruli. Diffuse tubulointerstitial inflammation with abundant eosinophils and trivial tubulointerstitial fibrosis (<5%) were observed. IF revealed non-specific staining for all immunoreactants. ANCA-GN with eosinophilic interstitial nephritis was diagnosed. Due to the alteration of consciousness, magnetic resonance imaging (MRI) of the brain and orbits were performed, demonstrating Wallerian degeneration of the right corticospinal tracts and pons and bilateral optic neuritis. Intravenous pulse methylprednisolone and intravenous immunoglobulin were administered as per AAV. Temporary hemodialysis was initiated for 2 weeks. Subsequently, she remained well without AAV clinical flare with low-dose prednisolone and immunosuppressive agents; her Scr was 1.6 mg/dL with negative proteinuria and bland urine sediment.An 84-year-old female with underlying diseases of hypertension and dyslipidemia presented with a fever for one week prior to admission. She received complete primary series of AZD1222 and a booster dose of the COVID-19 vaccine with mRNA-1273 four months ago. One month later, she developed bilaterally severe sensorineural hearing loss and tinnitus. A few weeks before admission, she developed drowsiness, progressive dysphagia, decreased urine volume, and a weight loss of 8 kg. PE revealed BP 154/66 mmHg, temperature 38.5 \u00b0C, mildly pale conjunctivae, right eye episcleritis with afferent pupillary defect (RAPD), and 1+ leg edema. Blood chemistries were notable for Scr 5.1 mg/dL (baseline 0.9 mg/dL), albumin 2.7 g/dL, hemoglobin 9.6 g/dL, leukocyte counts 26.0 \u00d7 10ANCA-GN secondary to COVID-19 vaccination was more prevalent in the patients with mRNA vaccinations , older age, and female gender. The top three most common manifestations were AKI, abnormal urine sediment, and constitutional symptoms. MPO-ANCA was distinctly dominant compared to the other ANCAs. Concomitant autoantibodies were documented in 52%, particularly Coombs antibody, ANA, and cryoglobulin. Although one-third (34%) presented with severe ANCA-GN, most cases had good outcomes, with a remission rate of 89%. No death occurred. The higher immunogenicity and more common usage of the mRNA COVID-19 vaccine might explain its higher prevalence on ANCA-GN ,19. The Most AAVs have unknown etiology, known as primary AAVs; however, AAVs can be secondary to drugs  ,25,26,27in-vitro experiment and found that the influenza vaccine (RNA vaccine) was able to induce PR3-ANCA production from peripheral blood mononuclear cells (PBMCs), while the decoy molecule, inactivated split virion influenza vaccine 2007, could not [AAV/ANCA-GN induced by vaccination has been well documented in post-influenza vaccination ,32. The ould not . In addiould not ,34. Unfoould not . COVID-19 vaccine induces ANCA-GN and AAV, possibly through molecular mimicry, stimulating productions of MPO-ANCA and PR3-ANCA via the adaptive immune system, likewise inducing antiviral neutralizing antibodies ,37,38. TTreatment of ANCA-GN following the COVID-19 vaccines in this systematic review mostly complied with the 2021 KDIGO Guidelines , wherebyANCA-associated glomerulonephritis following the COVID-19 vaccine is rare, according to billions of vaccine doses that have been administered worldwide. The benefits of COVID-19 vaccination outweigh the risks of possible induced autoimmune diseases and ACNA-GN. Avoid unnecessary booster doses of the COVID-19 vaccine in high-risk patients  should warrant."}
+{"text": "South African Journal of Physiotherapy 78(1), a1611. https://doi.org/10.4102/sajp.v78i1.1611, there was an error regarding the affiliations for the authors. Instead of:In the published article, Anwar, S., Arsalan, S.A., Zafar, H., Ahmed, A., Gillani, S.A. & Hanif, A., 2022, \u2018Effects of breathing re-education on endurance, strength of deep neck flexors and pulmonary function in patients with chronic neck pain: A randomised controlled trial\u2019, Authors:\u20181https://orcid.org/0000-0002-6603-3185Sahreen Anwar1https://orcid.org/0000-0003-3492-7050Syed A. Arsalan1,2https://orcid.org/0000-0001-9318-3691Hamayun Zafar1https://orcid.org/0000-0002-1965-6224Ashfaq Ahmed3https://orcid.org/0000-0002-2996-0764Syed A. Gillani4https://orcid.org/0000-0002-2670-6402Asif HanifAffiliations:1Department of Physical Therapy, Faculty of Rehabilitation Sciences, University of Lahore, Lahore, Pakistan2Department of Physical Therapy, Faculty of Rehabilitation Sciences, King Saud University, Riyadh, Saudi Arabia3Department of Biostatistics, Faculty of Rehabilitation Sciences, University of Lahore, Lahore, Pakistan\u2019It should be:\u2018Authors:1https://orcid.org/0000-0002-6603-3185Sahreen Anwar1https://orcid.org/0000-0003-3492-7050Syed A. Arsalan1,2https://orcid.org/0000-0001-9318-3691Hamayun Zafar1https://orcid.org/0000-0002-1965-6224Ashfaq Ahmed3https://orcid.org/0000-0002-2996-0764Syed A. Gillani4https://orcid.org/0000-0002-2670-6402Asif HanifAffiliations:1University Institute of Physical Therapy, University of Lahore, Lahore, Pakistan2Faculty of Rehabilitation Sciences, King Saud University, Riyadh, Saudi Arabia3Faculty of Allied Health Sciences, University of Lahore, Lahore, Pakistan4University Institute of Public Health, University of Lahore, Lahore, Pakistan\u2019.In addition, there was an error on page 2. The following paragraph is updated as it was incorrectly formulated:The original incorrect wording:n = 34 in each group, Z1-\u03b1/2 = standardised level of significance = 95% = 1.96, Z1-\u03b2 = Power of test = 80% = 1.28, \u00b51 = Mean in control group = 4.60, \u00b52 = Mean in physiotherapy group = 5.40, \u03b412 = standard deviation in control group = 0.84, \u03b422 = standard deviation in physiotherapy group = 0.59. The primary outcome used to estimate sample size was vital capacity (VC) (Duymaz 2019). A difference of 0.5 in the standard deviation was regarded as a meaningful change .Here, The revised and updated wording:n = 15 in each group, Z1-\u03b1/2 = standardised level of significance = 95% = 1.96, Z1-\u03b2 = power of test = 80% = 1.28, \u00b51 = mean in control group = 4.60, \u00b52 = mean in physiotherapy group = 5.40, \u03b412 = standard deviation in control group = 0.84, \u03b422 = standard deviation in physiotherapy group = 0.59. The primary outcome used to estimate sample size was vital capacity (VC) (Duymaz 2019). A difference of 0.5 in the standard deviation was regarded as a meaningful change .Here, The authors apologise for these errors. The corrections do not change the study\u2019s findings of significance or overall interpretation of the study\u2019s results or the scientific conclusions of the article in any way."}
+{"text": "Nonspecific presentation and limited diagnostic solutions of acute infections and sepsis in emergency departments (EDs) result in early antimicrobial administration at a cost to antimicrobial stewardship. We validated the host response classifiers, IMX-BVN-3b and SEV-3b, to determine bacterial and viral infection status and illness severity. We also assessed antibiotic over- and underuse.We prospectively enrolled adult ED patients with suspected acute infections or sepsis with \u22651 vital sign abnormal across 7 sites. At enrollment, we surveyed treating physicians on infection probability and antibiotic prescribing. BVN/SEV-3b were calculated using NanoString nCounter\u00ae on PAXgene\u00ae blood samples. We compared BVN-3b likelihood of bacterial and viral infection to post-hoc clinical adjudication infection status and SEV-3b likelihood of severe outcomes to 7-day need for ICU care and 30-day mortality.Of 568 enrolled patients, 346 had consensus adjudications . The BVN-3b area under the receiver operating curve (AUROC) for bacterial infections was 0.82 (95%CI 0.77-0.87), compared to 0.76 (95%CI 0.71-0.81) for procalcitonin (p = 0.011). BVN-3b AUROC for viral infections was 0.89 (95%CI 0.84-0.95). SEV-3b AUROC was 0.78 (95%CI 0.69-0.86) and 0.88 (95%CI 0.73-1) for 7-day ICU care and 30-day mortality, respectively. Of the 346, 226 patients had a pre-lab result physician questionnaire; of these, 110 patients received antibiotics. Of these, BVN-3b would have corrected 71% of antibiotic prescribing errors (34/48 overprescriptions and 13/18 underprescriptions).BVN-3b and SEV-3b accurately identified bacterial and viral infections and risk status. If implemented in a rapid workflow, these tests may improve patient management and antibiotic prescribing, reducing healthcare costs in the ED.Arianna Beltran, BS, Inflammatix: Internship Uan-I Chen, MS, Inflammatix: Stocks/Bonds Edward A. Michelson, MD, Inflamatix: Research expense reimbursement for sponsored clinical trial Alexandra Weissman, MD, Inflammatix Inc: Advisor/Consultant Evangelos J. Giamarellos-Bourboulis, PhD, D(ABMM), Abbot Product Operations AG: Grant/Research Support|Abbot Product Operations AG: Honoraria|bioMerieux: Grant/Research Support|bioMerieux: Honoraria|Horizon 2020 European Program: Grant/Research Support|Horizon Health European Program: Grant/Research Support|Sobi AB: Advisor/Consultant|Sobi AB: Grant/Research Support|Sobi AB: Honoraria Oliver Liesenfeld, MD, Inflammatix Inc.: Ownership Interest Natalie N. Whitfield, PhD, D(ABMM), GenMark Dx/Roche: Employee|GenMark Dx/Roche: Stocks/Bonds|Inflammatix: Employee|Inflammatix: Stocks/Bonds"}
+{"text": "The 13-valent pneumococcal conjugate vaccine (PCV13) was introduced for infant use in the United States in 2010, replacing PCV7. A U.S. case-control study (2010\u20132014) demonstrated vaccine effectiveness (VE) for \u22651 dose of PCV13 at 86%; however, this study lacked statistical power to examine VE by number of doses and against individual serotypes.We used the indirect cohort method to estimate VE of PCV13 against vaccine-type invasive pneumococcal disease among children < 5 years in the U.S. from May 1, 2010 to December 31, 2019. We included IPD cases identified through CDC\u2019s Active Bacterial Core surveillance in 10 U.S. states; during 2010 \u2013 2014, we additionally included cases enrolled in a post-licensure matched case-control study from expanded sites. Cases and controls were defined as individuals with PCV13-type-IPD (VT-IPD) and non-PCV13-type-IPD (NVT), respectively; serotype 6C was categorized as VT. We used logistic regression to estimate the adjusted odds of prior PCV13 receipt, controlling for confounders identified a priori including age category, race/ethnicity, sex, state, year, and any immunocompromising and/or chronic conditions.>3 PCV13 doses; 47 cases (21.1%) and 53 controls (5.7%) had received no PCV doses. Serotypes 19A (N=96), 3 (N=60), and 19F (N=45) caused 90.1% (201/223) of VT-IPD. VE of >1 or \u22653 PCV13 doses against VT-IPD was 81.7%  and 87.8% (75.2\u201394.0%), respectively. VE of \u22653 PCV13 doses was 87.0% (75.8\u201393.0%), 54.6% (-8.8\u201381.0%), and 92.9% (74.4\u201398.0%) against serotypes 19A, 3, and 19F, respectively. VE was 87.6% (67.9-95.2%) for three primary doses before 12 months of age and 92.4% (78.2\u201397.2%) for three primary doses and a booster at 12 months of age or older.A total of 1,161 IPD cases were identified; 223 (19.2%) were VT cases and 938 (80.8%) were non-VT controls. Of those, 108 cases  and 600 controls  had received Vaccine effectiveness estimates against PCV13-type IPD among US children under five years of age, 2010 - 2019VE of \u22651 PCV13 dose against IPD was consistent with the previous estimate from the case-control study, and \u22653 doses appear to provide substantial protection. Among the most commonly circulating VT-IPD, PCV13 was protective against serotypes 19A and 19F IPD; protection against serotype 3 IPD did not reach statistical significance.Lee Harrison, MD, GSK: Advisor/Consultant|Merck: Advisor/Consultant|Pfizer: Advisor/Consultant|Sanofi: Advisor/Consultant Tamara Pilishvili, PhD MPH, GSK: Employed by GSK since February 2023. At the time of data collection and analysis for this work was employed by the CDC"}
+{"text": "T/C authorization in the US for PrEP of COVID-19 in IC individuals was initially based on a randomized trial (PROVENT). However, < 5% of enrollees in PROVENT were IC. We sought to assess real-world effectiveness of T/C PrEP among IC pts.We conducted a retrospective, observational, propensity-score matched cohort study at UPMC from Jan 1, 2022, to Mar 31, 2023. We assessed effectiveness at \u2264 6 months of T/C 600 mg  against chart confirmed COVID-19 hospitalizations and COVID-19\u2013related deaths among IC pts.Table 1). During the study period encompassing circulating variants both susceptible and resistant to T/C, there were 15 vs 18 COVID-19 hospitalizations across 377,832 person-days in T/C vs non-T/C pts (Table 2). Thus, 0.72% vs 0.87% of T/C exposed vs unexposed pts were hospitalized for COVID-19  . There was no difference in effectiveness by T/C dosing pattern (Table 2). Effectiveness was numerically higher in the BA.5 period, although there were few events in other variant eras (Table 2). There was a non-significant 50% lower risk of hospitalization in T/C patients with solid tumors or autoimmune conditions, and no significant difference by exposure status in patients with organ transplant or hematologic cancer (Table 2). There were 5 COVID-19 ICU admissions per exposure group, and 2 inpatient COVID-19\u2013related deaths (1 per group) (Table 3). Limitations included overall low event rate limiting power and an open healthcare system leading to possibility of missing hospitalizations due to pts seeking care elsewhere.Before matching, there were 2931 T/C vs 157,225 non-T/C pts. After matching, 2301 matched pairs remained . Most pts had moderate/severe IC conditions : Advisor/Consultant|LabSimply: Advisor/Consultant|Merck: Advisor/Consultant|Shionogi: Advisor/Consultant|Shionogi: Honoraria John W. Mellors, MD, AstraZeneca: Grant/Research Support|Gilead Sciences: Grant/Research Support"}
+{"text": "Their structures and stereochemistry were determined from analysis of NMR spectroscopic and circular dichroism (CD) data. Compound 2 represents the first report of naturally occurring arylnaphthalide lignan triglycoside. The cytotoxic activities of all isolated compounds were evaluated against A-549 and SMMC-7721 cell lines. Compounds 7\u201310 and 14\u201316 were more toxic than cisplatin in two tumor cell lines. This investigation clarifies the potential effective substance basis of D. versipellis in tumor treatment.One new dibenzyltyrolactone lignan dysoslignan A ( Arylnaphthalide lignans have received much attention due to their potent antiviral, antineoplastic, anti-inflammatory, and immunosuppressive properties . The repDysosma versipellis (Hance) M. Cheng ex Ying, belonging to the family of Berberidaceae, is widely distributed in the central/south regions of China + ; NMR data (DMSO-d6), see Dysoslignan A (2): white, amorphous powder; + ; NMR data (DMSO-d6), see Dysoslignan B (3): white, amorphous powder; UV (MeOH) \u03bbmax (log \u03b5) 204 (4.49), 225 (4.19), 264 (4.35), 326 (3.76), 363 (3.60) nm; IR (iTR)\u03bdmax 3367, 2989, 2946, 2833, 1741, 1608, 1520, 1467, 1420, 1348, 1274, 1213, 1186, 1115, 1090, 1027 cm\u22121; HR-ESI-MS (positive): m/z 385.0920 [M + H]+ , m/z 407.0737 [M + Na]+ ; NMR data (DMSO-d6), see Dysoslignan C (4): white, amorphous powder; UV (MeOH) \u03bbmax (log \u03b5) 202 (4.48), 225 (4.29), 263 (4.38), 312 (3.82), 355 (3.60) nm; IR (iTR)\u03bdmax 3410, 2939, 2839, 1745, 1605, 1535, 1465, 1352, 1244, 1131, 1094, 1030 cm\u22121; HR-ESI-MS (positive): m/z 383.0768 [M + H]+ ; NMR data (DMSO-d6), see Dysoslignan D (1 (1.0 mg) and 2 (1.0 mg) were dissolved in 1.0 mL of 2M HCl, and then hydrolyzed at 90 \u00b0C for 3 h. The HCl in the reaction mixture was removed under reduced pressure. The remaining reaction mixture was extracted with CH2Cl2. The water layers were directly analyzed by HPLC . The peaks at 13.15 and 14.27 min were coincided with D-glucose and D-galatose.The absolute configurations of the galatose and glucose moieties were determined by the previously reported method . Compoun1\u201318 were evaluated against human lung cancer A-549, hepatocellular carcinoma SMMC-7721 cell lines. The cells were cultured in RPMI-1640 medium, supplemented with 10% fetal bovine serum (FBS) at 37 \u2103 under 5% CO2 in a humidified atmosphere. Cell viability was assessed by conducting colorimetric measurements of the amount of insoluble formazan formed in living cells based on the reduction of 3--5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS). To be brief, 100 \u00b5L of cells were seeded into each well in a 96-well cell culture plate in advance. After 24 h, various concentrations of all test compounds were added. After the incubation for 48 h, MTS (20 \u03bcL) was added to each well, and the incubation continued for 4 h at 37 \u2103. The optical density at 492 nm was determined using a 96-well microtiter plate reader. The IC50 values were calculated by the Reed\u2013Muench method. Statistical analysis were performed by SPSS 20.0 software . All experiments were performed in triplicate.By the previously reported MTS method , the cytD. versipellis resulted in the isolation of one new dibenzyltyrolactone lignan dysoslignan A (1), three new arylnaphthalide lignans dysoslignan B\u2013C (2\u20134), along with fourteen known metabolites (5\u201318). Compound 2 is the first reported example of naturally occurring arylnaphthalide lignan triglycoside. All isolated compounds were tested for their in vitro cytotoxic activity against A-549 and SMMC-7721 cell lines using MTS assay. Among them, compounds 7, 14, and 15 were cytotoxic, with IC50 values of less than 1\u03bcM. Our research further demonstrated that the arylnaphthalide lignans are mainly responsible for the potent anticancer effect of D. versipellis.Further phytochemical studies on"}
+{"text": "Staphylococcus/Mammaliicoccus spp. isolates from contralateral healthy skin and DFU from 44 hospitalized patients.Diabetic foot infections (DFIs) are the most common complications of diabetic foot ulcers (DFUs), and a significant cause of lower extremity amputation. In this study we used whole genome sequencing to characterize the clonal composition, virulence and resistance genetic determinants of 58 S. aureus (n\u2009=\u200932) and S. epidermidis (n\u2009=\u200910) isolates were recovered from both DFUs and healthy skin, whereas, S. haemolyticus (n\u2009=\u20098), M. sciuri (n\u2009=\u20091), S. hominis (n\u2009=\u20091) and S. simulans (n\u2009=\u20093) were recovered exclusively from healthy skin. In contrast, S. caprae (n\u2009=\u20092) and S. saprophyticus (n\u2009=\u20091) were recovered only from DFUs. Among S. aureus isolates, MRSA were present with high prevalence , 18 of which (66.7%) were from DFUs and 9 (33.3%) from healthy skin. In contrast, the coagulase-negative Staphylococcus (CoNS)/Mammaliicoccus isolates (n\u2009=\u200926), in particular S. epidermidis and S. haemolyticus were more prevalent in healthy skin,  and , respectively. MLST, spa and SCCmec typing classified the 32 S. aureus isolates into 6 STs, ST672, ST80, ST241, ST1, ST97, ST291 and 4 unknown STs (STNF); 8 spa types, t044, t037, t3841, t1247, t127, t639, t937 and t9432 and 2 SCCmec types, type IV and type III(A). Among CoNS, the S. epidermidis isolates belonged to ST54, ST35 and ST640. S. haemolyticus belonged to ST3, ST25, ST29, ST1 and ST56. The sole M. sciuri isolate was found to carry an SCCmec type III(A). A wide range of virulence genes and antimicrobial resistance genes were found among our isolates, with varying distribution between species or STs. The pan-genome analysis revealed a highly clonal population of Staphylococcus isolates, particularly among S. aureus isolates. Interestingly, the majority of S. aureus isolates including MRSA, recovered from the healthy skin and DFUs of the same patient belonged to the same clone and exhibited similar virulence/resistance genotype.Staphylococcus/Mammaliicoccus spp. in DFIs, which could serve as a basis for further studies on these as well as other groups of pathogens associated with DFIs.Our study provides clinically relevant information on the population profile, virulence and antibiotic resistance of The online version contains supplementary material available at 10.1186/s12866-023-03087-2. Diabetes is a fast-growing global problem with huge social, health, and economic consequences . The prePeople with diabetes are at increased risk of long-term complications such as coronary heart disease, cerebrovascular and peripheral vascular diseases and diabetic foot ulcers (DFUs) . It was Microbial infections of the DFUs, termed diabetic foot infections (DFIs) are key contributors to the amputation risk . Limb loStaphylococcus are common colonizers of skin and mucous membranes of humans and animals, but also opportunistic pathogens capable of causing a wide range of infections. The staphylococci can be differentiated into Coagulase-Positive (CoPS) and Coagulase-Negative (CoNS), based on their ability to produce coagulase. Recently, five species among the CoNS , and belonging to S. sciuri group, were reclassified into the novel genus Mammaliicoccus, with Mammaliicoccus (M.) sciuri as the type species , Staphylococcus/Mammaliicoccus species were identified among the 58 isolates, including S. aureus , S. epidermidis , S. haemolyticus , S. simulans , S. caprae , S. hominis, S. saprophyticus and M. sciuri . The characteristics of the 58 isolates included in this study are shown in Table\u00a0Eight S. aureus isolates, 23/32 (71.9%) were recovered from DFUs and 9/32 (28.1%) from healthy skin. The 10 S. epidermidis isolates were recovered from 8 patients, including 6/10 (60%) from healthy skin and 4/10 (40%) from DFUs. All the S. haemolyticus (n\u2009=\u20098), M. sciuri (n\u2009=\u20091), S. hominis (n\u2009=\u20091) and S. simulans (n\u2009=\u20093) isolates were recovered exclusively from healthy skin, but S. caprae (n\u2009=\u20092) and S. saprophyticus (n\u2009=\u20091) were recovered only from DFUs.Among the 32 S. aureus isolates, 27/32 (84.4%) carried the mecA gene, and were therefore MRSA; 18/27 of which (66.7%) were from DFUs and 9/27 (33.3%) from healthy skin.Among the 32 S. aureus including MRSA isolates and CoNS were significantly associated with the source of isolation .Statistically, S. aureus were characterized based on the combination of MLST, SCCmec and spa typing. In silico determination of MLST revealed that S. aureus isolates belonged to 6 known STs including ST80 , ST241 , ST672 , ST1 , ST97 and ST291 , and 4 STNF .Clones of S. aureus isolates. The dominant one was t044 , followed by t037 , t3841 , t1247 and t127 , t639, t9432 and t937 .Eight spa types were identified among mec types were identified among MRSA isolates, type IV  and type III(A) . The type IV isolates were assigned to subtype IVc(2B)  and IVd(2B) .Two SCCMammaliicoccus isolates, SCCmec type III(A)  was detected in the sole M. sciuri isolate.Among CoNS/The dominant MRSA clone  was ST80- t044- IVc(2B), followed by ST241-t037- III(3\u00a0A)) ; whereas, ST672- t3841- IVd(2B) and STNF- t037- III(3\u00a0A) were each represented by 3/27 (11.1%) isolates. In addition, 2 other spa types were detected among ST80- IVc(2B) isolates, t1247  and t639 .2/5 (40%) of MSSA isolates belonged to ST1-t127, while ST97-t9432, ST291-t937 and STNF-t037 were each represented by one isolate .S. epidermidis isolates, 6/10 (60%) belonged to ST54, 2/10 (20%) to ST35 and 1/10 (10%) to ST640. 3/8 (37.5%) S. haemolyticus belonged to ST3 and 2/8 (25%) to ST25, while ST29, ST1 and ST56 were each represented by 1/8 (12.5%) isolate.Among The presence and distribution of the virulence genes are summarized in Tables\u00a02,S. aureus isolates including 42 adhesion genes and a large number of type 8 capsular polysaccharide, immune evasion and exoenzyme genes.A total of 116 virulence genes were detected among hlgA, hlgB, hlgC, hlb, hld and hla/hly), 13 staphylococcal enterotoxins (se) and staphylococcal enterotoxin-like toxins (sel) , with different carriage proportions ranging from 3.1 to 100%. ST1 isolates carried the highest number (n\u2009=\u20097) of se/sel genes, sea, seb, seh, sek, seq, selk and selq. In contrast, none of the se/sel genes were detected among the ST80 isolates.Thirty-five toxin-encoding genes were found among MRSA/MSSA isolates including 6 hemolysins  or a toxic shock syndrome toxin (tst) gene.Remarkably, none of the lukD, lukE) were detected in all S. aureus isolates , while lukF/lukS-PV were detected only in ST80 isolates . In addition, edinB gene encoding epidermal cell differentiation inhibitors was also detected only in ST80 isolates  and in ST291- t937 .The leukocidins genes , which was exclusively present in S. epidermidis isolates .On the other hand, the sole virulence factor found among the 26 CoNS/S. aureus ST or CoNS/Mammaliicoccus species was statistically significant. In contrast, no statistically significant association was found between the presence/absence of virulence genes and PEDIS grades or the source of isolates .The association of virulence genes with S. aureus is shown in Tables\u00a0mecA and blaZ, were detected among S. aureus isolates at frequencies of 84.4% (27/32) and 50% (16/32), respectively.The distribution of the genetic determinants of antibiotic resistance among the AME), aph(3\u2019)- III/ aph(3\u2019)- IIIa, were the most prevalent among S. aureus isolates . All ST80 isolates  were positive for ant(6)-Ia and aph(3\u2019)-III/ aph(3\u2019)-IIIa genes, and all ST241 isolates  were positive for 3 AME genes, ant(9)-Ia, aac(6\u2019)- aph(2\u2019\u2019) and aph(3\u2019)- III.The genes encoding aminoglycoside-modifying enzymes (S. aureus isolates. The msr(A) and mph(C) genes were detected in all the ST672 isolates , and the erm(A) in all the ST241 isolates .Four genes encoding resistance to macrolide-lincosamide-streptogramin B (MLSB) were detected in S. aureus isolates, tet(M) , tet(K)  and tet(38) . Remarkably, only isolates belonging to t037 and harboring SCCmec-III (ST241 and STNF) carried tet(M).Three genes encoding resistance to tetracycline were detected among fusB and fusC genes coding for fusidic acid resistance were detected in all the ST80-t1247  and ST1-t127 , respectively. The dfrG gene coding for trimethoprim-sulfamethoxazole resistance was detected only in ST241 isolates .The M. sciuri isolates. 18/26 (69.2%) carried both mecA and blaZ genes. Remarkably, the single M. sciuri isolate harbored both mecA and mecA1 . 9/10 (90%) S. epidermidis isolates carried both fusB and fosB genes. In addition, msr(A) and mph(C) were detected in S. epidermidis  and S. haemolyticus  isolates. Moreover, ermC was found in S. haemolyticus  and S. epidermidis  isolates. aac(6\u2019)- aph(2\u2019\u2019)/ aac(6\u2019)-Ie/aph(2\u2019\u2019)-Ia genes were detected in S .haemolyticus  and M. sciuri .As presented in Table\u00a0aph(3\u2019)-III/ aph(3\u2019)-IIIa  and tet(K)  genes were found exclusively in S. haemolyticus and in S. epidermidis, respectively. Genes conferring resistance to streptogramin (vat(B), vat(C) and vgb(B)), macrolides (vga(B) and vga(A)LC), kanamycin/neomycin (aadD), tetracycline (tet(L)), streptomycin (str) and to quaternary ammonium compounds (qacB) were detected only in S. haemolyticus isolates, at a frequency of one gene per isolate.The S. epidermidis , S. aureus , S. haemolyticus  and M. sciuri  . This is consistent with earlier findings that S. aureus isolates recovered from 4 distinct anatomical sites  of patients with type 2 diabetes were highly related in the same patient  was performed using Roary . Tree cop-value for describing possible associations between species/ST and virulence/resistance genes with PEDIS grades and the source of isolates. p-values\u2009<\u20090.05 were considered statistically significant.Data were analyzed using the Statistical Package for Social Sciences . Contingency tables were constructed and Chi-squared tests were used to calculate Below is the link to the electronic supplementary material.Supplementary Material 1Supplementary Material 2Supplementary Material 3"}
+{"text": "Correction to: Psychometrika 10.1007/s11336-022-09879-1Second line in Eq. (15): \u201cbelow Eq. (23): \u201cSecond line in Eq. (27): \u201cThe original version of the article contains the below listed errors. The following transcription errors have been corrected:"}
+{"text": "The HVTN 704/HPTN 085 clinical trial (AMP study) assessed the efficacy of VRC01 broadly neutralizing antibody infusions for HIV prevention. It offered access to oral pre-exposure prophylaxis (PrEP) at no-cost to all participants as standard of HIV prevention care. Understanding PrEP initiation among trial participants and describing its impact on study endpoints can guide future HIV prevention trial design.We characterized sociodemographic and geographic features of PrEP initiation (by self-report) using descriptive statistics. We used Cox models to identify factors associated with PrEP initiation and the association between PrEP initiation (as a time-dependent exposure) and HIV incidence.Of 2221 participants off PrEP at enrollment, 31.8% initiated PrEP during the trial. Median time to PrEP initiation was 60 days (range 1-673 days). Brazil had the highest proportion of PrEP initiation among their enrolled participants (83.2%) and Peru had the lowest (5.3%). In US sites, 54.2% initiated PrEP. Southern US sites had the highest proportion of PrEP initiation (68.8%) and the Midwest had the lowest (28.4%).In multivariate analysis, prior PrEP use [HR (95% CI) = 2.2 (1.3-4.0)] was associated with PrEP initiation. Participants from Switzerland [0.5 (0.3-1.0)] and Peru [0.08 (0.06-0.1)] had lower likelihood of PrEP initiation compared to the US, while participants from Brazil had higher likelihood [2.6 (2.0-3.3)]. Age, race, gender, and sexual orientation were not associated with PrEP initiation. HIV incidence rate was 3.6/100 person-years (PY) before PrEP initiation, compared to 1.2/100 PY after PrEP initiation. PrEP initiators had 58% less risk of acquiring HIV after adjusting for behavioral risk score and treatment arm. In the US, higher local PrEP-to-need ratio was also associated with lower PrEP initiation [0.9 per 5 units (0.8-1.0)].Providing PrEP access to AMP participants reduced HIV acquisition among PrEP initiators. Study endpoints were still met due to heterogenous regional PrEP uptake and lower PrEP effectiveness than expected. These data suggest that offering oral PrEP to HIV prevention trial participants as standard of care is a feasible option to consider for future efficacy HIV prevention trial design.Valeria D. Cantos, MD, Gilead Sciences: Grant/Research Support Srilatha Edupuganti, MD MPH FIDSA, Sanofi: Grant/Research Support Colleen F. Kelley, MD, MPH, Gilead Sciences: Grant/Research Support|Humanigen: Grant/Research Support|Moderna, Inc.: Grant/Research Support|Novavax, Inc: Grant/Research Support|Viiv Healthcare: Grant/Research Support"}
+{"text": "Phage-antibiotic combinations (PAC) have been proposed for high inoculum daptomycin-nonsusceptible (DNS) MRSA infections refractory to conventional therapy. We studied PAC with synergistic activity against two DNS MRSA clinical bloods isolates .9 CFU/mL) using: i) modified checkerboard (CB) minimum inhibitory concentration (MIC); and ii) 24h time-kill assays (TKA). Synergistic activity in CB assays was defined as either: a fractional inhibitory concentration (FIC) index \u2264 0.5 in modified CB assays; or a \u2265 2 log10 CFU/mL reduction by PACs vs the most active single-agent regimen. Significant differences between regimens were assessed by ANOVA with Tukey\u2019s post hoc modification (P < 0.05).PAC containing DAP and/or ceftaroline (CPT) (each at \u00bd MIC) plus a 2-phage cocktail  were tested at a high MRSA inoculum  plus either DAP or DAP + CPT demonstrated robust synergistic activity  vs. the next most effective regimen of CPT + Intesti13+Sb1 (P < 0.05 each). Against C37, Intesti13 + Sb1 (MOI of 1 and 0.1) with CPT or DAP + CPT were equally potent and effective regimens (-\u03947.14 log10 CFU/mL each), but neither were significantly better than the synergistic regimen of DAP + Intesti13 + Sb1 (-\u03946.65 log10 CFU/mL).By CB assay, synergistic activity was demonstrated with Intesti13 + Sb1 (MOI of 10 to 0.01) plus either DAP or DAP + CPT (FIC \u2264 0.5 for each combination). In 24h TKA vs These studies demonstrated positive phage-antibiotic synergy (PAS), with phage cocktail Intesti13 + Sb1 and either DAP or DAP + CPT. Addition of these phages to DAP or DAP + CPT combinations caused bactericidal and synergistic killing vs. CPT + phages. *, P<0.05. Abbreviations: CFU: colony forming units, GC: growth control, DAP: daptomycin, CPT: ceftaroline, MOI: multiplicity of infection, PAC: phage-antibiotic combination.These studies demonstrate positive phage-antibiotic synergy (PAS) with phage cocktail Intesti13 + Sb1 added to DAP, CPT, and DAP + CPT at an MOI of 1 and 0.1. Abbreviations: CFU: colony forming units, GC: growth control, DAP: daptomycin, CPT: ceftaroline, MOI: multiplicity of infection, PAC: phage-antibiotic combination.in vivo investigations of these candidate PACs, for treatment of high inoculum DNS MRSA infections  is warranted.The two-phage cocktail used (Intesti13 + Sb1) demonstrated impressive synergistic activity against two DNS MRSA isolates in combination with DAP or DAP + CPT. Further experimental Arnold S. Bayer, MD, Akagera Medicines: Grant/Research Support|ContraFect Corporation: Grant/Research Support Michael J. Rybak, PharmD, PhD, MPH, Abbvie, Merck, Paratek, Shionogi, Entasis, La Jolla, T2 Biosystems: Advisor/Consultant"}
+{"text": "Triticale  is of great interest as an insurance crop that can ensure the stability of the gross harvest of feed and food grains at a lower cost. In Western Siberia, only winter triticale varieties are cultivated, however, spring triticales are important for cultivation in regions not suitable for winter crops. To create spring varieties with high yields and good grain quality, it is necessary to study and enrich the gene pool, identify donors of economically valuable traits. One of the possible ways to solve this problem can be through the production of secondary hexaploid triticales with the involvement of the tetraploid wild-growing species of emmer wheat Triticum dicoccum (Schrank) Schuebl. The aim of this work was to create and study hybrids of emmer T. dicoccum (Schrank) Schuebl. with hexaploid triticale using genomic in situ hybridization for staining of meiotic chromosomes and analysis of plant productivity elements in F4\u2013F8. DT4, DT5, DT6 plants and the prebreeding F6 forms obtained from them \u2013 DT 4/168, DT 5/176 and DT 6/186 \u2013 were selected according to the characteristics of the productivity and the nature of the grain in the F4 hybrid population. The offspring of hybrids DT4 and DT5 and prebreeding forms DT 4/168 and DT 5/176 had an increased grain nature (over 750 g/l), but low productivity. The hybrid DT6 and the breeding form DT 6/186 obtained from it had high grain productivity , but, like the paternal form of triticale UK 30/33, had a reduced nature of the grain. In F8 DT 6/186 plants, 7 homologous pairs of rye chromosomes and from 27 to 30 wheat chromosomes were found in meiosis, which indicates the presence of a complete rye genome and two wheat \u0410\u0410\u0412\u0412 genomes. Rye chromosomes showed stable formation of bivalents in contrast to wheat chromosomes, which caused the presence of aneuploids in plant populations. Thus, hexaploid forms DT 4/168 and DT 5/176 with well-made smooth grain and high grain size were obtained, which can be used as a source of this trait for selection of food-grade triticale. DT 6/186 is a promising form for further breeding in order to obtain high-yielding forms of triticale. Triticale  is a wheat and rye hybridof a relatively short evolutionary history as an allopolyploidspecies. The first naturally created fertile wheat\u00d7rye hybridswere discovered in the late 1920s at the South-Eastern agriculturalexperimental station in Saratov . Theplants had intermediate traits and were described as a new botanicalspecies Triticum Secalotriticum saratoviense Meisterby G.K. Meister . Meister immediately predictedthe practical value of these intergeneric crossings. Thefirst man-made wheat\u00d7rye hybrids were obtained in 1888 by the German plant breeder W. Rimpau, who described 12 plants descending froma wheat\u00d7rye hybrid generally recognized as the first triticales. The cytological analysis of thefirst triticales developed in Russia and Germany showed thesomatic chromosome number of 56 (8\u0445) , which demonstrated the combinationof four genomes as follows: AABBDDRR, AABBDD,soft wheat, and RR rye.Octoploid triticales were of great interest for plant breedersdue to high seed set per spike, increased plant pathogen resistance,and resistance to environmental stresses. However, theprimary triticale lines suffered from meiotic errors  and high frequency of aneuploidy in earlier generations, which caused reduced fertility. Grain shriveling andlatenessof maturity in octoploid forms were also consideredas limitations for their introduction as cultivars. As a result,there have been global efforts to develop the technology forobtaining more promising lines with various valuable breedingtraits, which eventually produced hexaploid triticales.Primary hexaploid triticales of the AABBRR genome typewere obtained as crossings of tetraploid wheats  and S. cereale rye . However, undesirabletraits were not completely eliminated. Agronomicallyvaluable traits were improved by enriching the triticalegene pool with crossing experiments involving octoploidlines and commercial soft wheat varieties, as well as hexaploidtriticales with full rye genome and two wheat genomesidentified in the progeny of octoploid triticales as a resultof D-genome chromosome elimination . The crossing of two primaryoctoploid triticales produced hexaploid offspring, and breed-ingprograms were focused on hybridization of these octoploidswith the hexaploids identified in the progeny .Hexaploid triticales were more stable in terms of productivity. As a result, various recom-binantforms of secondary triticales were derived and karyotyped,and some of them having combined wheat-rye genomesshowed commercial value .Agronomic traits of triticales were further improved byintergeneric and interspecific crossings. Aegilops crassa , Ae. juvenalis , Ae. squarrosa and Ae. triaristata  , Agropyron intermedium ssp. trichophorum(2n = 42) , Hordeum parodii Covas, H. vulgare L. and T. monococcum L.  plants wereused in hybridization experiments with hexaploid triticales.Intergeneric polyploid triticales were also bred through hybridizationwith intermediate forms sharing at least one setof chromosomes (genome) with the triticale genome. Thesehybridization efforts produced plants with resistance genesagainst diseases. Lines isolated in the progeny of triticale hybrids(AABBRR) and amphidiploids  were resistant toyellow rust . Hybridization of a hexaploidtriticale and an amphiploid intermediate form  produced addition andsubstitution lines with the 3Sv Ae. variabilis chromosome carryingthe powdery mildew resistance gene Pm13 . Addition and substitution lines for chromosome 2Dwith the leaf-rust resistance gene Lr39 and semidwarf geneRht8, as well as chromosome 3D (or 3D/3B) with the Lr32 gene were isolated in the progeny of triticale \u00d7 amphiploid hybrids .Thanks to breeding achievements, triticale has becomea new economically significant cereal species characterizedby high grain and vegetative mass productivity, that could beused as forage and green feed . In the last three decades, its products have becomeincreasingly significant, which is demonstrated by increasingcrop acreage across the world, from 1,453,269 ha in 1994 to4,157,018 ha in 2016. Triticale grains are used to producebioethanol and food wrap, as well as various food products , andbran is used as the source of prebiotics and antioxidants foryoghurts. Food-grade triticale grains are comparable to wheatin macro- and micronutrient contents . Proteincontent in triticale grains is 1\u20131.5 % higher than in wheat and3\u20134 % higher than in rye, gluten content matches that in wheator is 2\u20134 % higher . However, triticaleunderperforms in test weight. This parameter is closely relatedto the plumpness and hardness of grains, as well as to theirsize and shape. Average test weight for wheat is 700\u2013810 g/l.At test weights below 740 g/l, flour yield tends to decreaserapidly as test weight goes down. Most spring triticales haveshriveling grains and low flour yields, which limits their usein bread making .Development of domestic high-productivity triticale varietieswith high grain quality requires further study and enrichmentof the gene pool, as well as identification of donors foreconomically valuable traits. One way to solve this problemis to obtain secondary hexaploid triticales using emmer wheatT. dicoccum (Schrank) Schuebl., a tetraploid wild-growingwheat with long, large, and plump grains.The goal of the present study was to breed new forms ofhexaploid triticales (\u0410\u0410\u0412\u0412RR genome type) with improvedtest weights by crossing emmer  with triticale and study their productivity and meioticstability with chromosome staining using genomic in situhybridization.Plant material. New forms of hexaploid triticale were obtainedby hybridization of emmer (T. dicoccum (Schrank)Schuebl.) and triticale . IntergenericF1 hybrid of emmer lines (L133 \u00d7 PKK) \u00d7 k-25516 (AABBgenome) was used as maternal plants. Awned semi-hullessemmer (L133 \u00d7 PKK) created by VIR researchers is characterizedby brittle spikes and low productivity, and awnlessemmer k-25516 was obtained at Siberian Research Instituteof Plant Production and Breeding (SRI PPB), ICG SBRAS from the population of awned emmer wheat from theVIR collection. The paternal plants were represented by a selectionform of hexaploid triticale UK 30/33 selected fromthe population of cytogenetically unstable octoploid triticaleUK30 (AABBDDRR genome) developed at the SRI PPB bycrossing the Ulyanovka soft wheat variety with the Korotkostebelnaya69 short stem rye with subsequent doubling ofchromosome number induced by colchicine water solution.The progeny of three F4 hybrids DT4, DT5, and DT6, andthree selection forms DT 4/168, DT 5/176, and DT 6/186 isolatedin F5 from hybrid populations DT4, DT5, and DT6 respectivelywas selected for the study. In 2020, their F6 progenywas seeded for research purposes along with F4 hybrids atthe nursery for distant wheat hybrids in the field of SRI PPB.Fluorescence in situ hybridization of meiotic chromosomes.To evaluate the meiotic stability in the selection forms,two most productive plants DT 6/186/156 and DT 6/186/165were selected in the F7 progeny of plant DT 6/186, and theirF8 seeds were seeded in the hydroponic greenhouse of theICG SB RAS in spring 2021. The chromosomal behavior ofthe progeny was studied using routine acetocarmine stainingprotocol and FISH staining (fluorescence in situ hybridization)following the technique described earlier .The meiocytes were analyzed at the diakinesis stage, metaphaseI (\u041cI), anaphase I (AI), and telophase II (TII). Theprobes used in the analysis were as follows: Aegilops tauschiipAet6-09 specific for centromere repeats in rice, wheat,rye, and barley chromosomes ; pAWRcspecific for centromere repeats in rye chromosomes , and rye genomic DNA. DNA repeat samples ofpAet6- 09 and pAWRc were the courtesy of Dr. A. Lukaszewcki.Centromere-specific probes were labeled with biotin 16-dUTPor digoxigenin 11-dUTP by means of polymerase chain reaction(PCR). The total rye DNA was labeled by Nick translationwith digoxigenin 11-dUTP. The probes were combined indifferent proportions and mixed with blocking wheat DNA.The preparations were mounted in Vectashield antifade solution(Vector Laboratories) slowing down fluorescence fadingand including 1 \u03bcg/ml DAPI  for chromatin staining. Allpreparations were analyzed using an Axio Imager M1 microscope, the images were recorded usinga ProgRes MF camera  at the Centerof Microscopic Analysis of Biological Objects, SB RAS andprocessed using Adobe Photoshop CS2 software.Analysis of economically valuable traits. Structuralanalysis of the plants was performed at the facility equippedfor metric measurements, threshing, and seed weighting. Asa result, the following data on productivity elements wereobtained: spike length; spike density; grain weight per spike;thousand kernel weight; grain number per spikelet; testweight measured using a microchondrometer ; and grain productivity per 1 m2. Statisticalprocessing of the results was carried out following thestandard technique . Significance of differencesbetween mean values of two samples was estimatedusing Student\u2019s t-test.The parental-line plants had different spike and grain morphology.The emmer plants (L133 \u00d7 PKK) had short, awned,brittle spikes and smooth, long grains . The k-25516emmer sample was an awnless plant with thin, long grains . Triticale UK 30/33 had a dense, awned spike and grains similar to those of soft wheat in shape but shriveling.Plants of three emmer\u00d7triticale F4 hybrids had differentspike morphology . The DT6 hybridhad a dense, awned spike. The plants of the remaining twohybrids had looser spikes. The spikes were sterile at the endsand therefore often susceptible to ergot.All hybrids had a hair neck similar to the paternal triticaleUK 30/33. This means that these genotypes have a gene responsiblefor manifestation of this trait localized in the longarm of chromosome 5R.Triticale UK 30/33 had a short spike, low test weight, andmedium grain productivity (see Table 1). Selection forms andhybrids, except for DT5, had a longer spike than the paternalform UK 30/33. Hybrid DT5 had low productivity due toflower sterility in the upper part of the spike and low thousandkernel weight. Hybrid DT4 had a high thousand kernelweight of 50 \u00b1 2 g, and the highest test weight of 806 \u00b1 14 g/l. However, its grain productivity, although higher than that ofDT5, was still not too high due to low seed set of spikes andspikelets. Flowers at the top of the spike were often sterile aswell. Hybrid DT6 was characterized by dense spikes and highgrain productivity (785 \u00b1 41 g/m2), as well as seed set of spikesand spikelets. The spikes were fertile along their full length.Selection forms DT 4/168, DT 5/176, and DT 6/186 haddifferent grain weights per spike, grain numbers per spike,test weights, and productivities. DT 4/168 had denser spikesand slightly higher seed sets of spikes and spikelets comparedto the DT4 hybrid it was obtained from. Breeding sampleDT 5/176 was more productive than the initial DT5 hybrid,but it had the lowest productivity among the three breedingsamples studied. Small spike size and sterility of 3\u20137 spikeletsin the upper part of the spike resulted in low productivityof hybrids DT5 and DT 5/176. The DT 6/186 line had thehighest seed set and productivity among the studied lines.Despite the lowest test weight values similar to triticaleUK 30/33 (706 \u00b1 25 g/l), grain productivity per plot reached822 \u00b1 74 g/ m2. DT 6/186 had a smoother overall morphologythan the initial hybrid DT6, but higher breeding value due toits higher grain productivity.Cytological analysis of the progeny of two plants (DT 6/186/156 and DT 6/186/165) from the highly productive DT 6/186line revealed instability in chromosome number and errors inchromosomal behavior in the first and second meiotic divisions.Chromosome staining using genomic in situ hybridizationin these plants showed 14 rye chromosomes formingbivalents, which implied the presence of seven homologouspairs . However, the bivalent chromosomes demonstratedpremature separation in some meiocytes (desynapsis),with rye chromosomes becoming univalent and distributinganomalously between the poles .Chromosome number in the discovered aneuploid plantsvaried from 2n = 41 to 2n = 44. Among the ten plants from theDT 6/186/156 progeny, only one plant had chromosome number2n = 42 , while there were no plants with euploidchromosome numbers in the progeny of DT 6/186/165.Univalents were discovered in the metaphase I , which were lagging at the equator during chromosomeseparation in anaphase I in 86.75 \u00b1 4.56 and61.32 \u00b1 2.81 % of cells in DT 6/186/156 and DT 6/186/165respectively . The lagging chromosomeswere divided into sister chromatids or broken at the centromere. Instability of chromosome separationduring the division resulted in micronuclei formation atthe tetrad stage . Micronuclei were observed in 60.29 \u00b1 3.14 and 72.16 \u00b1\u00b1 2.29 % tetrads in DT 6/186/156 and DT 6/186/165, respectively(see Table 2). Even the euploid plant with 2n = 42(DT 6/186/156) had micronuclei in 51.48 % tetrads, whichprevented us from considering this plant fully cytogeneticallystable.Plants with the minimum number of anomalous meiocytesin anaphase I and telophase II were selected from populationsDT 6/186/156 and DT 6/186/165 based on the results of theanalysis.Triticale  as an agricultural cropcombines wheat\u2019s high yield potential with rye\u2019s resistanceto biotic and abiotic stresses, which increases its adaptability to cultivation conditions in salty or highly acidic soils and inpresence of toxic heavy metals. Thanks to these traits, triticaleis of great interest as an emergency crop ensuring stable grossharvest of forage and food grains at lower costs . Despite today\u2019s applications of triticale grainsbeing mostly restricted to forage in animal husbandry and productionof feed and bioethanol, there is a rising interest in theuse of triticale grains in human food products. Triticale grainshave proven nutritional and dietary value , since they include not only proteins, carbohydrates,and fats, but also vitamins, minerals, and dietaryfibres (14\u201318 %) . Comparedto that of wheat, the protein from its grains has a richer aminoacid profile, particularly in indispensable amino acids, suchas lysine, threonine, and leucine . Triticale starch amounting to 3/4 of thekernel weight has a significantly lower amylose content comparedto rye and wheat , which ensures its betterdigestion by humans .To increase the share of triticales in production of breadsand pastries, in recent decades breeding efforts have beenaimed at increasing the quality of grains and finished products,which has resulted in development of triticale bread-makingvarieties , so State standards for triticaleflour have been developed (State Standard 34142-2017).The characteristics of winter triticale varieties also includeapplicability for bread making. Triticale breeding efforts inRussia and other countries are primarily aimed at developingwinter varieties . However, grainquality assessment of the samples from the spring triticalecollection showed that spring triticales have good potentialfor creating bread-making varieties . The samples with such improvedparameters as protein content, test weight, falling number,vitreousness, gluten quantity and quality, etc., are used inbreeding for improved grain quality .A trait such as high test weight can be transferred by distanthybridization. This parameter is closely related to geneticallydetermined parameters such as grain plumpness, hardness, andshape. In the present paper, tetraploid emmer wheat T. dicoccum(Schrank) Schuebl. with long, large, and plump grainswas used as the maternal line for hybridization with hexaploidtriticale . As a result, new hexaploidforms of triticale with AABBRR genome types were created,which is confirmed by the analysis of meiotic chromosomalbehavior using genomic in situ hybridization. Seven pairs ofrye chromosomes and 27 to 30 wheat chromosomes were observedin the plants, which implies the presence of a completerye genome and two wheat genomes. Three plants, DT4, DT5,and DT6, were isolated in the F4 population, the progeny ofwhich demonstrated test weight values significantly exceedingthose of the initial triticale line, and test weights in the progenyof DT 4/168, DT 5/176, and DT 6/186 forms from F6 werehigher or on par with those of the initial triticale line. Theseplants had different productivity values, with the highest onesin F4 recorded in the DT6 line (785 \u00b1 41 g/m2). The respectivevalue for DT 6/186, i. e., the progeny of the latter in F6,reached 822 \u00b1 74 g/m2. The DT 4/168 and DT 5/176 lines areof moderate interest for further research into improvement ofbread-making properties of triticale grains.The study of the meiotic behavior of rye and wheat chromosomesin the progeny of F8 plants DT 6/186/156 andDT 6/186/165 showed that they had not yet achieved cytogeneticstability, which was indicated by the discovered chromosomeseparation errors and the presence of aneuploids in thepopulations. Chromosome separation errors mostly occurredin wheat chromosomes due to their monosomy. Cytologicalinstability and aneuploidy in octoploid and hexaploid wheatryeallopolyploids have been an issue from the start , but secondary triticales turned outto be more cytogenetically stable than primary ones . Cytological study of triticales showed that the interactionbetween wheat and rye genomes in the cells of the sameplant resulted in physiological defects in cells persisting fordecades at least. Meiotic and mitotic errors were found bothin the triticale obtained by Rimpau in 1888  and in the triticales obtained later. Despite thecomplete set of chromosomes, meiotic univalents were foundin triticales with different ploidy levels . The studyinto meiotic chromosomal behavior in triticales in the presentand earlier papers demonstrated that while only bivalentswere present at the diakinesis stage, univalents appeared inmetaphase I as a result of to produce lagging chromosomes atthe equator . Presumably, aneu-ploidcells in triticales may emerge as a result of asynchronousseparation of rye and wheat chromosomes and their laggingin anaphase and telophase . Thepresumed cause of meiotic instability in the obtained amphidiploids is an imbalance in the genetic system of meioticpairing and the differences in cell cycles between wheat andrye .It is known for a fact that chromosome separation dependson kinetochore function . It was found thatkinetochore protein CENH3 produced by one of the parentsmaintained the function of other parental kinetochores in stablehybrids, despite the differences between DNA sequences ofcentromere regions in parental lines . It wasalso shown that cytogenetic stability in triticales could also belinked to increased expression of rye-specific CENH3 formsin a hybrid genome .Another possible cause of cytogenetic instability of theobtained triticales could be a nuclear-cytoplasmic incompatibility,because the hybrids are bred using an intergenericF1 hybrid of emmer lines (T. dicoccum (Schrank) Schuebl.)(L133 \u00d7 PKK) \u00d7 k-25516 (genome AABB) as the maternal form.The AABB genomes in paternal triticale forms originate fromsoft wheat, while the rye genome is used as the base. Selectionof genotypes of wheat varieties during triticale backcrossingmay lead to recovery of fertility and cytogenetic stability innew forms, as exemplified by alloplasmic soft wheat lines(H. vulgare)-T. aestivum .The result of this work was to create and study hybrids ofemmer T. dicoccum (Schrank) Schuebl. with hexaploid triticale.According to the characteristics of productivity and testweight values of the grain, hexaploid prebreeding forms F6were isolated \u2013 DT 4/168, DT 5/176 and DT 6/186. Thus,hexaploid forms DT 4/168 and DT 5/176 with well-filled,smooth grains and high test weights that can be used as thesource of said traits in food-grade triticale breeding havebeen obtained. The DT 6/186 line shows promise in terms ofbreeding for high yields. As a result of the analysis of meioticdivision in DT 6/186 plants, this sample is unstable forwheat chromosomes, therefore, aneuploids will occur in theoffspring. To restore fertility and cytogenetic stability of newforms of triticale with emmer cytoplasm and chromosomes, itis necessary to select the genotypes of wheat varieties duringbackcrossingThe authors declare no conflict of interest.Abdelkawy R.N.F., Turbayev A.Zh., Solov\u2019iev A.A. Technologicalproperties of spring triticale grain grown in the Central EconomicRegion, Russia. Vestnik Kurskoy Gosudarstvennoy SelskokhozyastvennoyAkademii = Bulletin of the Kursk State Agricultural Academy.2020;5:87-97. (in Russian)Ammar K., Mergoum M., Rajaram S. The history and evolution of triticale.In: Triticale Improvement and Production (FAO Plant Productionand Protection Paper. 179). Rome, 2004;1-10.Ayalew H., Kumssa T.T., Butler T.J., Ma X.-F. Triticale improvementfor forage and cover crop uses in the Southern Great Plains of theUnited States. Front. Plant Sci. 2018;9:1130. DOI 10.3389/fpls.2018.01130.Badaev N.S., Badaeva E.D., Bolsheva N.L., Maximov N.G., ZeleninA.V. 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+{"text": "Conversely, plasma cell and M2 macrophage levels were lower. The follicular helper T cells in tumor cell-enriched regions were negatively correlated with resting NK cells and positively correlated with activated NK cells. In immune cell-enriched regions, this relationship was reversed. We also explored the heterogeneity of HLA gene families, immune checkpoints, and metabolism-related genes in the three regions. In tumor cell-enriched regions, we obtained 19 prognosis-related metabolism genes via univariate cox analysis. We used multiplex immunofluorescence to verify the elevated expression of SLC8A1 and MDH1 in immune cell-enriched regions and tumor cell-enriched regions, respectively, both of which were associated with prognosis of NPC. In conclusion, we explored the spatial heterogeneity of the NPC tumor environment and found specific diagnostic and prognostic markers that can be used to differentiate tumor cell-enriched regions from immune cell-enriched regions in NPC.The heterogeneity of nasopharyngeal carcinoma (NPC) leads to mixed clinical outcomes. We collected 92 regions of interest from 41 biopsies of patients with untreated NPC and obtained their transcripts using GeoMx Digital Spatial Profiling (DSP) technology. Spatial heterogeneity was determined by measuring the expression of marker genes in tumor cell-enriched (PanCK-expressing), immune cell-enriched (CD45-expressing), and normal epithelial (Endo) regions. We screened 16 prognostic markers in tumor cell-enriched regions and 4 prognostic markers in immune cell-enriched regions. The levels of CD8 Nasopharyngeal carcinoma (NPC) is mainly prevalent in North Africa, the Middle East, and Southeast Asia. Its occurrence is mainly attributed to the local environment, genetics, and infection with Epstein-Barr virus (EBV) The tumor microenvironment is composed of tumor cells, immune cells, stromal cells, metabolites, cytokines, and the extracellular matrix The current study explored the spatial heterogeneity of NPCs in both composition and function. The digital spatial profiling (DSP) technology 2 tissue cores obtained from 58 patients using a TMArrayer (Pathology Devices). Regions consisting of high tumor content with large numbers of tumor infiltrating lymphocytes (TILS) were selected following preparation of the TMA block using H&E sections from the whole tissue block to ensure the presence of NPC cells in each sample. Subsequent DSP RNA assays were performed using 41 cores including 95 regions of interest . Three regions of interest (ROIs) were excluded from paired analyses due to loss of tissue integrity during staining. Finally, 38 cores containing paired tumor cell- and immune cell-enriched ROIs were included in the differential analysis.We collected paraffin-embedded tumor tissue from 58 NPC patients who underwent initial nasopharyngeal biopsy without treatment at the Affiliated Houjie Hospital of Guangdong Medical University. This study was approved by the Ethics and Scientific Committee of the Affiliated Houjie Hospital of Guangdong Medical University. A tissue microarray (TMA) was constructed with 1.5 mmAnother commercial NPC TAM  containing 126 cases of NPC tissue were used for Immunofluorescence analysis. Five cores were excluded from analyses due to loss of tissue integrity during staining. Extreme values  were excluded from survival analysis.NanoString GeoMx DSP RNA assays were performed at CapitalBio Technology  using the standard protocol. Slides were prepared following the Manual RNA Slide Preparation Protocol in the GeoMx DSP Slide Preparation User Manual . The Whole Transcriptome Atlas (WTA) probe reagent was added to slide. We stained the tissue with GeoMx Solid Tumor TME Morphology Kit  to distinguish various morphology: epithelial cells were positively stained with PanCK while immune cells were positively stained with CD45, and nuclear stained with SYTO13. The pathologist distinguishes normal epithelial cells from cancerous epithelial cells based on histological morphology. Regions of interest (ROIs) were selected and assessed by a pathologist and illuminated using UV light. The indexing oligonucleotides released from each ROI were collected and deposited into designated wells on a microtiter plate. DSP assay sequencing data were processed with the GeoMx NGS Pipeline (DND). After sequencing, the reads were trimmed, merged, and aligned to a list of indexing oligos to identify the source probe. The unique molecular identifier (UMI) region of each read was used to remove PCR duplicates and duplicate reads, thus converting reads into digital counts. The limit of quantitation (LOQ) was estimated as the geometric mean of the negative control probes plus two geometric standard deviations of the negative control probes. Targets that consistently fell below the LOQ were removed, and the datasets were normalized via upper quartile (Q3) normalization. We used the prcomp function to perform principal component analysis from the gene expression matrix and plotted it with the scatterplot3d package.Comparisons between the two groups were performed using the Mann-Whitney U test or Wilcoxon signed-ranks test. Genes of significance were defined based on a fold change > 1.5 and p-values <0.05. Gene ontology (GO) enrichment and KEGG enrichment analyses of DEGs were performed using clusterProfiler R-packages with Benjamini-Hochberg multiple testing adjustment.2 =0.89) for the Pearson correlation coefficient matrix. A topology overlap matrix (TOM) was constructed to perform hierarchical clustering with at least 50 genes per module (minClusterSize =50). The cutreeDynamic function was used to automatically cut the clustered modules and merge similar modules (abline = 0.25). The turquoise module contained 569 genes. The green module carried 267 genes and the blue module included 1010 genes. The genes of modules were extracted for GO (q =0.05) and KEGG (p =0.05) functional enrichment analysis. The R packages involved in these analyses were: clusterProfiler, org.Hs.eg.db, enrichplot, and ggplot2. Genes from the turquoise module (n=569), green module (n=267) and blue module (n=1010) were respectively submitted to the STRING database (http://www.string-db.org/). Parameter settings were as follows: network type (full STRING network), meaning of network edges (confidence), and minimum required interaction score (high confidence (0.700). Protein interaction data obtained from the STRING database were entered into the Cytoscape software. The cytoHubba plugin was used to screen core genes in the PPI network.Weighted correlation network analysis (WGCNA) of 2940 differential genes filtered from 18675 genes from matched 38 PanCK-expressing and 38 CD45-expressing regions was performed using the WGCNA and limma packages. The gene expression matrix was converted into a Pearson correlation coefficient matrix. The adjacency matrix was constructed based on the optimal power value  obtained from 38 PanCK-expressing regions was screened for 16 prognosis-related genes by univariate Cox analysis. The optimal cutoff value of each of these genes was determined according to the surv_cutpoint and surv_categorize functions. The expression of each gene was grouped according to the optimal cutoff value, followed by survival analysis with the log-rank test. The turquoise module (n=569) of 38 CD45-expressing regions was screened for four prognosis-related genes by univariate Cox analysis. The optimal cutoff value for each of these four genes was determined according to the surv_cutpoint and surv_categorize functions. The expression of each gene was categorized according to the optimal cutoff value, followed by survival analysis with the log-rank test. The R packages involved in the above process were survival and survminer.The relative content of 22 immune cells was calculated for each of the 92 ROIs  using the CIBERSORT algorithm. Immune cells were visualized in 90 ROIs  at P < 0.05. Differences in immune cell levels among the three regions  were identified using the Kruskal-Wallis test with the reshape2 and ggpubr packages. A set of genes related to immune function was obtained based on previous studies Differential expression of HLA genes in the three regions (92 ROIs) was determined using the limma, reshape2, ggplot2, and ggpubr packages via Kruskal-Wallis test. Differential expression of immune checkpoint-related genes in the three regions was explored using limma, ggplot2 and ggpubr packages via Wilcoxon test. Correlation analysis between immune checkpoint gene expression and immune cell levels was performed in PanCK- and CD45-expressing regions using the limma, reshape2, tidyverse, and ggplot2 packages with Spearman correlation coefficients. This analysis was visualized.http://www.gsea-msigdb.org/gsea/downloads.jsp). GSVA of metabolism-related genes was performed in 92 ROIs using the packages GSEABase, GSVA, limma, and pheatmap. Parameters were set to P < 0.05 and only 20 pathways were shown. A total of 134 genes were obtained by intersecting 944 metabolic genes with 2940 differential genes (PanCK- vs CD45-expressing regions) and plotting the Venn diagram based on the Bioinformatics (http://bioinformatics.psb.ugent.be/webtools/Venn/) website. Univariate Cox analysis (P < 0.05) of 134 genes was performed in 38 (PanCK-expressing) ROIs to screen for prognosis-related metabolic genes, survival analysis, and ROC curve plotting. A total of 171 genes were obtained by intersecting 944 metabolic genes with 2514 differential genes (PanCK-expressing vs Endo regions). Univariate cox analysis, survival analysis, and ROC curve plotting of 171 genes were performed in 38 (PanCK-expressing) ROIs. The R packages involved in univariate cox analysis, survival analysis, and ROC curve plotting were survivor, survminer, and timeROC.We obtained 944 genes related to metabolism based on the literature TMA section was deparaffinized to retrieve the antigen and block endogenous peroxidase, according to manufacturer's instructions. Tissue sections were blocked with 3% BSA in TBST for 30 min, and then incubated with the antibody for CD45  for 30 min. The antibody was detected using the corresponding secondary antibody tagged with HRP, before visualization using CY3-TSA. Subsequently, antigen was retrieved again to prepare the slides for the next antibody. All samples were stained sequentially with CK  visualized with FITC-TSA, MDH1  visualized with Opal 647 TSA, and SLC8A1  visualized with Opal 594 TSA. Slides were counterstained with DAPI for nuclei visualization for 10 min and coverslipped using the antifade mountant. All markers stained with multiplex immunofluorescence were reviewed by a pathologist.Single-cell transcriptome data (GSE162025) from 10 NPC tissues with 82,622 cells was analyzed using Seurat to deal with the expression matrix. By setting filtering parameters, we filtered out cells with mitochondrial gene ratio>10% to exclude cells in abnormal state, and filtered out cells with gene number<500 or >3000 to exclude data with poor sequencing quality and non-single cells, and finally obtained 75710 cells. We used the Harmony package for data integration and batch removal. We performed unsupervised clustering using the default parameters in the Seurat package and then annotated these cell populations. We used the FindAllMarkers function in the Seurat package to calculate the differential genes between cell populations (log2FC=0.25 and P<0.05). The obtained differential genes were sorted to get the top ten highly expressed genes with significant differences for each cell population, and the expression matrix of these genes was obtained using the FetchData function, and plotted using the corrplot package.All immunofluorescence slides were scanned using the Pannoramic Digital Slide Scanner (3DHistech) and images visualized in CaseViewer2.4 (3DHistech). TMA core images were analyzed in HALO (Indica Labs). Area Quantification FL V2.1 module in Halo V3.0.311.314 analysis software was used to quantify the positive area and colocalization-positive area of the target region, respectively. Extreme values  were excluded to minimize data variability.The study protocol is shown in Figure CDK1, CCNB1, MCM2, TOP2A, CDC6, DTL, TPX2, UBE2C, KIF2C, RFC4, TYMS, MCM4, NUSAP1, MCM7, CENPF, FOXM1, RRM1, PCNA, MRPL13 and MRPL12 as the core genes were used to construct the PPI network. Turquoise modular genes including CD4, CD8A, CD3E, CD247, ZAP70, LCK, CD3G, CD3D, FYN, VAV1, LCP2, CD28, ITK, PTPRC, LAT, CD2, CD80, PRKCQ, IL2RB and CTLA4 as the core genes were used to construct the PPI network , while the turquoise module was significantly positively correlated with CD45 expression (r=0.73 and P= 8e-14) Fig. A. A tota network C. EnrichCD27, CEP85L, DOK3, MAST4, SEC24A and UPK3B appeared to serve as protection factors. Conversely, the genes BZW2, DLL4, GTPBP4, LSM4, MBD3, PAICS, PALM2AKAP2, PAQR4, RUVBL1 and TGS1 represent risk factors; the higher the expression of these genes, the higher the risk of death in patients with NPC , CEP85L (P < 0.001), DOK3 (P <0.001), MAST4 (P<0.001), SEC24A (P <0.001), and UPK3B (P <0.001) in PanCK-expressing regions of NPC patients correlated with better prognosis. In contrast, the expression of BZW2 (P <0.001), DLL4 (P <0.001), GTPBP4 (P <0.001), LSM4 (P <0.001), MBD3 (P <0.001), PAICS (P <0.001), PALM2AKAP2 (P =0.001), PAQR4 (P <0.001), RUVBL1 (P =0.006) and TGS1 (P =0.124), correlated positively with worse prognosis in patients with NPC , CCL21 (P =0.012), FCGR2C (P <0.001), and SLC8A1 (P =0.003) was associated with poorer prognosis in patients with NPC . In PanCK-expressing regions, the genes NPC Fig. . A total+ T cells, follicular helper T cells, activated NK cells, monocytes, M0 macrophages, M2 macrophages, resting mast cells, and activated mast cells differed among the three groups . The relative levels of plasma cells were the highest in CD45-expressing regions, while the number of CD8+ T, follicular helper T cells, and activated NK cells were the highest in PanCK-expressing regions. M0 macrophages were the highest in PanCK-expressing regions, while M2 macrophages were the highest in CD45-expressing regions , and activated dendritic cells with CD40 (P <0.001) and IDO1 (P <0.01). Conversely, negative correlations were found between CD4 memory activated T cells and BTLA (P <0.01), activated mast cells and CD244 (P <0.01), and CD40LG (P <0.01), macrophages M1 and LAIR1 (P <0.01)  and TNFSF14 (P <0.001), macrophages M1 and CTLA4 (P <0.001), and B memory cells and CD40 (P <0.001). Negative correlations were found between CD8+ T cells and BTNL2 (P <0.01), plasma cells with TNFSF14 (P <0.001), resting NK cells with IDO1 (P <0.01), macrophages M0 and TNFSF4 (P <0.01), resting dendritic cells and CD27 (P <0.01), and B memory cells and CD276 (P <0.01). We performed cluster annotation of 75710 cells from 10 NPC tissues and obtained a total of 18 cell populations  are shown in Figure CD200 (P =0.04), CD40 (P =0.036) and ICOSLG (P =1.4e-05) was higher in PanCK-expressing regions than in CD45. The expression of CD244 (P =0.0032), CD160 (P =0.0075), CD80 (P =8.5e-06), CD48 (P =1.3e-07), CD40LG (P =1.4e-06), CD28 (P =0.0034), CD27 (P =2.1e-09), BTLA (P =0.00033), ICOS (P =0.00075), HAVCR2 (P =0.0044), CTLA4 (P =2e-06), NRP1 (P =1.4e-09), LAIR1 (P =1e-05), LAG3 (P =0.0054), PDCD1LG2 (P =0.0016), TNFRSF8 (P =0.017), TNFRSF4 (P =0.021), TMIGD2 (P =0.019), TIGIT (P =8.2e-05), and TNFSF18 (P =0.0033) was higher in CD45-expressing regions than in PanCK-expressing regions. In Endo regions, the expression of CD200 (P =0.0013), CD70 (P =0.00057), CD44 (P =0.029), CD40 (P =0.0062), CD276 (P =0.0043), ICOSLG (P =1.1e-05), TNFRSF9 (P =0.0016), and TNFSF4 (P =0.0038) was lower than in PanCK-expressing regions, while the expression of NRP1 (P =0.046), IDO1 (P =0.0056), and VTCN1 (P =0.0025) was higher than in PanCK-expressing regions (The expression of HLA genes differed in the three regions (P <0.05) Fig. A. The ex0 P =0.03 and ICOSIn PanCK-expressing regions, metabolic genes Figure A were ma-expressing regions revealed 11 prognosis-related metabolic genes. The expression of COMT (P = 0.012), DUT (P =0.040), GMPS (P =0.043), MDH1 (P = 0.039), MDH2 (P = 0.020), NME1 (P =0.029), PAICS (P =0.011), POLR2I (P =0.039), PTGS2 (P =0.034) and UCK2 (P =0.029) increased the risk of NPC, while PLCB2 (P = 0.048) was a protective factor , GMPS (P <0.001), MDH1 (P =0.001), MDH2 (P <0.001), NME1 (P <0.001), PAICS (P <0.001), POLR2I (P <0.001), and UCK2 (P =0.004), the worse was the prognosis of NPC. In contrast, a higher expression of PLCB2 (P <0.001) was associated with a better prognosis for survival in patients with in NPC  predicted 2-, 4-, and 5-year survival rates in patients with NPC with a high degree of accuracy. MDH1 expression is higher in PanCK than in CD45 (P < 0.001). Multiplexed immunofluorescence analysis revealed higher levels of MDH1 protein in PanCK than in CD45 (P < 0.001), and the higher the MDH1 protein level (P = 0.048), the worse was the prognosis of patients with NPC  Fig. B. Univartor Fig. C. In PanNPC Fig. D. The RONPC Fig. F.ADCY3 (P = 0.027), ADCY9 (P =0.028), CYP2J2 (P = 0.038), DNMT1 (P = 0.024), EPHX1 (P =0.022), MBOAT7 (P =0.018), POLD1 (P = 0.016) and UAP1 (P = 0.041) correlated positively with survival risk in patients with NPC , and were also metabolism-related genes. The expression of these genes correlated with a higher risk of death in patients with NPC.Higher expression of ADCY3 (P = 0.005), ADCY9 (P =0.008), CYP2J2 (P = 0.001), DNMT1 (P <0.001), EPHX1 (P <0.001), MBOAT7 (P <0.001), POLD1 (P = 0.001) and UAP1 (P < 0.001) was associated with worse prognosis in patients with NPC  B. Univarwith NPC C. Notablwith NPC D. The ROEither bulk-RNA seq or microarray sequencing was used to explore biomarkers of NPC before GeoMx DSP technology emerged CEP85L, KRT5, MAST4, MYO1G, SLA, SMARCC2, UPK3B, ATF5, BEX3, BIK, CADM4, CDK2AP1, CLDN1, DLL4, IGFBP2, NFE2L3, PTGS2, SMC1A, SUSD4 and VRK2) and 4 prognostic markers in immune cell-enriched regions . We also screened for metabolism-related genes that affect the prognosis of patients with NPC in tumor cell-enriched regions .Using DSP technology, we screened 20 prognostic gene markers located in tumor cell-enriched regions  associated with PDAC cell survival + T, CD4+ T-helper, na\u00efve T-cells, cytotoxic T-cells, exhausted T-cells, and Tregs) and B cells + and CD8+ T cell clusters in NPC were highly activated and depleted and co-expressed effector markers such as IL-2, GZMB, INFG, NKG7, GNLY, and GZMK. They also expressed depletion markers such as PDCD1, HAVCR2, LAG3, TIGIT, and CTLA4 + T, follicular helper T cells, activated NK cells, and M0 macrophages were higher in tumor cell-enriched regions than in immune cell-enriched regions. In contrast, plasma cells and M2 macrophages were more abundant in immune cell-enriched regions, suggesting that cellular immunity was predominantly exerted in tumor cell-enriched regions, while humoral immunity was mainly observed in the peritumor area. Interestingly, in tumor cell-enriched regions, follicular helper T cells were negatively correlated with resting NK cells and positively correlated with activated NK cells. In immune cell-enriched regions, the opposite was true, highlighting the need to use single-cell sequencing to further explore the interactions between these two cell types. In tumor cell-enriched regions, PDCD1 expression was negatively correlated with the levels of monocytes and CD4+ memory-activated T cells. The expression of LAG3 positively correlated with the levels of CD4+ memory-activated T cells and CD8 T cells. The expression of TIGIT positively correlated with monocyte levels and negatively correlated with follicular helper T cell levels. In immune cell-enriched regions, the PDCD1 expression negatively correlated with the number of eosinophils and CD4+ na\u00efve T cells. The expression of HAVCR2 negatively correlated with the number of gamma/delta T cells and positively correlated with the levels of CD4+ memory resting T cells. The expression of LAG3 positively correlated with the levels of CD4+ memory activated T cells, CD4+ memory resting T cells, M1 macrophages, and CD8+ T cells, but negatively correlated with the number of plasma cells. The expression of TIGIT was positively related with the levels of Tregs, CD8+ T cells, and CD4+ memory resting T cells, while negatively correlated with the number of plasma cells. The expression of CTLA4 positively correlated with the levels of CD8+ T cells, CD4+ memory resting T cells, activated NK cells, and M1 macrophages, while negatively correlated with the number of naive B cells. We therefore analyzed the relationship between these immune cells and depletion markers in locations that were complementary to previous studies. However, the specific mechanisms of interactions between cells at various spatial locations need to be further explored.The predominantly infiltrating lymphocytes in the NPC microenvironment are T (CD8Limitations of this study include the small sample size, which restricts the correlation between the tumor immune characteristics, treatment, and outcomes. Even though our study is small the strength is the paired samples. Another limitation is that the biomarkers we screened have not yet been validated in functional experiments, and the roles of these biomarkers still need to be further elucidated through in vivo and in vitro experiments in the future.In conclusion, we used DSP technology to explore the heterogeneity of the NPC microenvironment and its regional and metabolic prognostic markers and characterized the immune infiltration. The screened prognostic biomarkers may provide a theoretical basis for clinical diagnosis, targeted therapy, and new drug development.Supplementary figures and tables.Click here for additional data file."}
+{"text": "Metastatic rhabdomyosarcoma (RMS) is a challenging tumor entity that evades conventional treatments and endogenous antitumor immune responses, highlighting the need for novel therapeutic strategies. Applying chimeric antigen receptor (CAR) technology to natural killer (NK) cells may offer safe, effective, and affordable therapies that enhance cancer immune surveillance. in vitro and in a metastatic xenograft mouse model.Here, we assess the efficacy of clinically usable CAR-engineered NK cell line NK-92/5.28.z against ErbB2-positive RMS in vitro and significantly prolong survival and inhibit tumor progression in mice. The persistence of NK-92/5.28.z cells at tumor sites demonstrates efficient antitumor response, which could help overcome current obstacles in the treatment of solid tumors.Our results show that NK-92/5.28.z cells effectively kill RMS cells These findings encourage further development of NK-92/5.28.z cells as off-the-shelf immunotherapy for the treatment of metastatic RMS. AdoleDeveloping new therapeutic strategies to overcome tumor resistance remains a critical unmet need for these heavily pretreated patients. Chimeric antigen receptor (CAR)-engineered cells, which target and inhibit the proliferation and spread of tumor cells, are revolutionizing the treatment of r/r malignancies. However, this therapy can also lead to severe, albeit treatable, side effects, such as cytokine release syndrome (CRS), immune effector cell-associated neurotoxicity syndrome (ICANS), and secondary hemophagocytic lymphohistiocytosis/macrophage activation syndrome .The lack of suitable CAR target antigens, as well as the immunosuppressive nature of the tumor microenvironment (TME), and advanced-stage disease, renders CAR-T cells nonresponsive or exhausted against many solid tumors , 9. HoweHere, we employed a molecularly and functionally well-defined clonal derivative of the natural killer (NK) cell line NK-92 (NK-92/5.28.z) as effector cells \u201317, carrin vitro  aRMS xenograft model.To further develop the NK-92/5.28.z cell therapy towards clinical application as a consolidation treatment for children and young adults with r/r, metastatic, ErbB2-positive RMS tumors after re-induction radiotherapy/chemotherapy, we performed detailed preclinical 22.1The NK-92 cell line was derived from a 50-year-old patient with non-Hodgkin\u00b4s lymphoma . NK-92/5PAX3-FOXO1) derived from a patient with alveolar RMS was purchased from DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH). Cells were cultured in Roswell Park Memorial Institute (RPMI) 1640 medium with GlutaMAX (Gibco) containing 10% heat-inactivated fetal bovine serum (FBS) . For in vivo cell tracking, stable GFP/luciferase-expressing RH30 cells (RH30GFP/luc+) were generated by lentiviral transduction with the pSIEW-luc2 plasmid encoding enhanced green fluorescent protein (eGFP) and firefly luciferase linked by a T2A peptide (GFP/luc+ cells were enriched by fluorescence-activated cell sorting using a FACSAria II instrument (BD Biosciences).The aRMS cell line RH30 ( peptide . GFP-posPAX3-FOXO1), RMS127 (PAX3-FOXO1), and RMS335 (PAX7-FOXO1) were generated from tumor biopsies of patients  and subsequently stained with ErbB2 antibody for 20 minutes. After washing, 1x104 events were recorded by flow cytometry using FACSDiva software . Fluorescence minus one (FMO) controls were used as references. The actual ErbB2 receptor number was quantified using BD Quantibrite Beads (BD Biosciences) according to the manufacturer\u2019s instructions. ErbB2 expression was determined using FlowJo Software .Tumor organoid RMS cells were stained with phycoerythrin (PE)-conjugated antibody  for ErbB2 and subsequently analyzed with a FACSCanto 10c flow cytometer (BD Biosciences). Cells were stained according to the manufacturer\u2019s instructions. In brief, aliquots of 5x102.3The short-term toxicity of NK-92/5.28.z cells against tumor organoid RMS cells was determined using a europium release assay as reported elsewhere . In brie2.45 cells were washed in phosphate-buffered saline (PBS) and subsequently stained with ErbB2 antibody or an IgG1\u03ba isotype control  for 20 minutes according to the manufacturer\u2019s instructions. After washing, 1x104 events were recorded by flow cytometry using FACSDiva software . In addition, ErbB2-CAR surface expression of NK-92/5.28.z cells was assessed. For this purpose 2x106 either NK-92/5.28.z or NK-92 cells were washed once with DPBS and then incubated with Human TruStain FcX\u2122 (Biolegend) for 20 minutes at 4\u00b0C. After another washing step in DPBS, the cells were incubated with 10 \u00b5g recombinant ErbB2 with human Fc tag for 20 minutes. After two additional washing steps, cells were stained with APC antibody directed against the Fc tag (Biolegend), 410712 cells, washed again, and analyzed using a BD FACS Canto 10c instrument (BD Biosciences).The integrity of RH30 target and NK-92/5.28.z effector cells was verified. In brief, aliquots of 5x10in vitro: tumor cells were resuspended and were counted by Neubauer-improved hemocytometer. 5000 RH30GFP/luc+ cells per well were seeded in 200 \u03bcl of RPMI +10% FBS into ultralow attachment 96-well round-bottom plates without prior coating (Corning) and spun down. On day four of culture, 100 \u00b5l of supernatant was carefully removed, and 1x105 NK-92 or NK-92/5.28.z cells resuspended in 100 \u00b5l of NK-92 medium were added. Spheroids without effector cells were used as controls. Every three to four days, the medium was replaced by X-Vivo 10 medium with 5% FFP and 100 IU/ml IL-2. Spheroids were imaged on day 4, 5, 6, 8 and 10 after initial seeding using a Celigo Image Cytometer (Nexcelom Bioscience) with F-theta lens and AVT PIKE camera. The size of spheroids was quantified by the GFP signal using Fiji software (Version 2.3.0)  and were conducted according to the requirements of the German Animal Welfare Act.5 RH30GFP/luc+ cells resuspended in 100 \u00b5l of PBS were injected intravenously (iv) via the tail vein. Considering them as having an immanence risk for disease progression with then limited treatment options as previously shown  or NK-92/5.28.z cells . Both effector cell types were resuspended in X-Vivo 10 medium with 5% FFP and 100 IU/ml IL-2. Group size was set based on the experience from previous experiments with the same xenograft model.Twenty-seven female 10- to 12-week-old nonobese diabetic (NOD)/severe combined immunodeficient (SCID)/Il2receptorgamma\u2212/\u2212 (NSG) mice received sublethal irradiation with 2.5 Gy (Biobeam 2000) (d-1) according to the protocol for previously established metastatic RMS xenograft model closely resembling a clinical situation . One dayly shown , immunotin vivo grade VivoGlo luciferin (Promega) in 100 \u00b5l of PBS per mouse. Fifteen minutes later, images were acquired in dorsal and ventral positions and subsequently analyzed by Living Image In Vivo Imaging Software (Perkin Elmer). Uniform regions of interest were used for all mice, and total flux (photon/s) was used for measurement. The signals of dorsal and ventral images were combined into a luminoscore, and statistical analysis of the tumor burden was performed by one-way ANOVA (Tumor growth was monitored weekly by bioluminescence imaging (BLI) using an IVIS Lumina II system (Perkin Elmer). Mice were anesthetized by isoflurane inhalation and received subcutaneous injections of 150 \u00b5g ay ANOVA .Altogether, tumor-bearing mice were randomly divided into the following three groups:- Control: one course of six serial infusions with X-Vivo 10 media with 5% FFP and 100 IU/ml IL-2, n=56 cells, one course of six serial effector cell infusions, n=12- NK-92: 10x106 cells, one course of six serial effector cell infusions, n=10- NK-92/5.28.z: 10x10During the experiment, animals were monitored daily for disease symptoms, xenogenic graft versus host disease (GVHD) and other adverse effects of the NK cell therapies for a maximum of 105 days. Mice with visible signs of disease progression, discomfort or physical abnormalities were painlessly sacrificed by isoflurane anesthesia followed by cervical dislocation.2.5.1Peripheral blood (PB), BM, lung, liver, gut and spleen samples were isolated and analyzed for persistence of human tumor or effector cells: BM was flushed out of the femur and tibia of mice. PB and BM were incubated with red blood cell lysis buffer (RBC Lysis Buffer (10x), BioLegend) according to the manufacturer\u00b4s instructions and washed once with PBS. The organs were cut in representative halves, and one half was preserved in formaldehyde for fluorescence microscopy. The other half was incubated with collagenase D solution , filtered through a 70 \u00b5m cell strainer, and washed with PBS. Aliquots of the cell suspensions were analyzed by flow cytometry and quantitative polymerase chain reaction (qPCR). Sample identity was blinded for the executors of the consecutively analyses.2.5.2GFP/luc+ was assessed shortly before use in experiments with a FACS Canto 10c device (BD Biosciences). CAR expression of NK-92/5.28.z cells was analyzed using a chimeric ErbB2-Fc protein  after nonspecific Fc receptor blocking  and subsequent staining with an anti-IgG-Fc secondary antibody conjugated with allophycocyanin (APC) .GFP expression of RH30Cell samples from mouse organs were washed once in PBS and stained with phycoerythrin-cyanin 7 (PE/Cy7)-conjugated anti-human CD45 antibody .Analyses were performed with FlowJo Software .2.5.3Genomic deoxyribonucleic acid (DNA) from murine organs was extracted using the Extractme Genomic DNA Kit (BLIRT S.A.). The percentage of human cells in the different murine tissues was determined by a quantitative real-time approach specifically amplifying the human albumin gene. In a second step, the proportions of NK-92 or NK-92/5.28.z cells and tumor cells within the human cell fraction were determined by a human-specific short tandem repeat (STR) genotyping approach. The tumor burden of each mouse was quantified in organs previously described as RMS metastatic sites  , 32. For2.5.4For histological analyses, organ sections were fixed in 4% buffered formalin, paraffin embedded, cut, and stained with hematoxylin-eosin (HE). Immunohistochemistry (IHC) antibodies targeting CD56, MYOD1, DESMIN and MYOGENIN were used to evaluate the RMS phenotype of the tumors. The stained tissue sections were rated by a pathologist for the expression of each protein .2 fragment goat-anti-mouse conjugated to Alexa 647  and goat-anti-rabbit IgG conjugated to Alexa 488  were used, and nuclei were stained with 4\u00b4,6-eiamidino-2-phenylindole (DAPI) (Sigma-Aldrich). The localization of immune cells in the tumor tissue relative to blood vessels was determined by staining with rat-anti-mouse Meca32 antibody  and secondary goat-anti-rat Alexa546 antibody . The samples were examined with a BZ-X810 All-in-One Fluorescence microscope (Keyence) with basic lenses  and filter cubes Ex: 360/40 DM: 400 BA: 460/50 (DAPI), Ex: 470/40 DM: 495 BA: 525/50 (GFP), Ex: 620/60 DM: 660 BA: 700/75(Cy5).Immunofluorescence staining was used to observe immune cell infiltration in organs and tumors: organ slices were deparaffinized and rehydrated. After antigen retrieval and a blocking step, primary mouse-anti targeting human CD45 antibody  and rabbit-against GFP antibody  were added. For detection, secondary antibodies F(ab`)In addition, the stained tissue sections with immune cell infiltration were analyzed by a pathologist for xenogenic GVHD.2.6p< 0.05 (*), p< 0.01 (**), p< 0.001 (***) and p<0.0001 (****) were considered statistically significant.For statistical analyses and graphical presentation, GraphPad Prism software  was used. The results are given as the mean \u00b1 standard deviation (SD). Differences between different groups were evaluated by two-tailed Student\u00b4s t test or one-way ANOVA using the Bonferroni-Dunn (nonparametric) method. Overall survivals are given as median with 95% confidence interval, Differences between the survival of different treatment groups were analyzed by the log-rank (Mantel-Cox) test. Differences with 33.1in vitro assessment of efficacy, parental NK-92 or NK-92/5.28.z cells were tested against patient-derived tumor organoid aRMS cells in a 2D europium release assay. Maintaining actively growing rare patient-derived cells is challenging, but the tumor organoid RMS cells RMS102 (PAX3-FOXO1), RMS127 (PAX3-FOXO1) and RMS335 (PAX7-FOXO1) could be successfully passaged in vitro using previously published cultivation conditions  on day 10 (p = 0.0013) and NK-92-treated spheroids (p = 0.0425)  xenograft were used to assess homing, tumor invasion, persistence, and antitumor effects as well as the xenogenic cytotoxicity (GVHD) of parental NK-92 and NK-92/5.28.z cells. Sequential infusions of effector cells were given preemptively during the low tumor burden period, and mice were followed for 105 days (end of experiment) (p = 0.0101) and NK-92-treated (p = 0.0159) animals at the time of first in vivo response assessment on day +50 (p = 0.2120) . In contrast, NK-92/5.28.z cell therapy (81.0 \u00b1 8.8 days) significantly improved overall survival compared to that of the parental NK-92-treated (p = 0.0329) and untreated controls (p = 0.0366) . All detectable tumor lesions expressed CD56 , MYOD1 , DESMIN  and MYOGENIN .Tumor lesions were primarily observed in the livers and lungs of mice except in those with complete response to immunotherapy with NK-92/5.28.z cells. Multiple large (macroscopic) lesions were more likely to be present in the untreated and NK-92-treated mice vs. single small (microscopic) lesions in the NK-92/5.28.z-treated mice . In vivo treatment of mice with parental NK-92 cells had no significant effect, indicating that the observed therapeutic efficacy of NK-92/5.28.z cells was mainly mediated by the second-generation ErbB2-CAR construct.In summary, our data show that NK-92/5.28.z cells exhibit high cytotoxicity in r/r ErbB2-positive 2D patient-derived tumor organoid RMS cells and 3D aRMS tumor spheroid models in vivo expansion of NK-92/5.28.z cells upon irradiation may be addressed by repeated dosing and combination with checkpoint inhibition or other anticancer therapies that could further enhance or benefit from the immunomodulatory activity of the cells.NK-92/5.28.z cell therapy is already under safety evaluation in patients with recurrent glioblastoma, which is an advantage when considering to extend clinical application of this ready-to-use CAR-engineered product to other cancer indications such as metastatic aRMS. We therefore propose to use NK-92/5.28.z cells for consolidation therapy of r/r, metastatic ErbB2-positive RMS following re-induction radiotherapy/chemotherapy to benefit from their multiple endogenous cytolytic pathways in addition to direct CAR-mediated killing. Thereby, the limited life-span and lack of The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The animal study was approved by Regierungspr\u00e4sidium Darmstadt, Darmstadt, Germany; Gen.-Nr. TVA FK/1070. The study was conducted in accordance with the local legislation and institutional requirements.CH, LM, WW, EU, HB, MM and ER conceived and designed the experiments. CH, LM, HK, HB, EG, EU, MM and ER performed the experiments. CH, LM, HK, HB, EU, MM and ER analyzed the data. HK, HB, TT, WW, EG, EU, MM, MK, FH, JD, J-HK, PB contributed to reagents, materials and analysis tools. CH, LMM and ER wrote the manuscript. HK, HB, TT, WW, EG, EU, MM, M, FH, JD, J-HK, PB and MM revised the manuscript. J-HK and PB supervised the research. All authors contributed to the article and approved the submitted version."}
+{"text": "Correction: Implementation Sci 18, 31 (2023)https://doi.org/10.1186/s13012-023-01286-zThe original publication  of this In the second paragraph, Lyon and Bruns was incorrectly cited as reference 1, where it should have been cited as reference 70.The correct reference 70 is supplied below:70. Lyon AR, Bruns EJ. User-Centered Redesign of Evidence-Based Psychosocial Interventions to Enhance Implementation\u2014Hospitable Soil or Better Seeds? JAMA Psychiatry. 2019;76(1):3\u20134. doi:10.1001/jamapsychiatry.2018.3060The original article  has been"}
+{"text": "Ornithobacterium hominis type strain MSHR-COH1 (ATCC TSD-185/NCTC 14317), a bacterial species isolated from the human nasopharynx. Long-read sequencing reveals that the genome is 2,036,909 bp in length, with a GC content of 35.72%.We report the complete genome sequence of the  Ornithobacterium hominis is a species in the family Weeksellaceae, for which partial genomes were first described from nasopharyngeal samples from Thailand  (ATCC TSD-185), streaked on Columbia agar with 5% sheep blood , and incubated at 37\u00b0C for 72 h under humid microaerobic conditions  using the CampyGen atmosphere generation system  plus approximately 20\u2009mL water. All bacterial colonies were scraped into phosphate-buffered saline and immediately processed using the MasterPure complete DNA/RNA purification kit  following the manufacturer\u2019s protocol. The sequencing library was prepared with the 24-plex native barcoding kit v12 . No DNA shearing or size selection was employed.N50 value of 10,303 bp and an N90 value of 5,608 bp.The library was sequenced on an ONT MinION device with an R10.4 flow cell , with Guppy v6.4.6 base calling using the SUP model and adaptor/barcode removal. Reads were trimmed using Prowler (SRR8360295]) mapped with Minimap2 v2.24 , and 3 rRNA operons. Sites that may disrupt short-read assemblies include the rRNA operons, 450-bp noncoding sequences associated with putative Fibrobacter succinogenes domain genes, >500-bp rearrangement hotspot (RHS) domain sequences, and >1-kb sequences within putative type III fibronectin domain genes. Unusually, the dnaA gene is situated near the proposed genome terminus rather than near the origin of replication , Flye v2.9.2 (https://github.com/fenderglass/Flye), Trycycler v0.5.3 (https://github.com/rrwick/Trycycler), Ori-Finder 2022 (http://tubic.tju.edu.cn/Ori-Finder2022/public/index.php) (accessed 11 January 2023), Pilon v1.24 (https://github.com/broadinstitute/pilon), Minimap2 v2.24 (https://github.com/lh3/minimap2), and Proksee (https://proksee.ca) (accessed 27 April 2023).The following software was used: Guppy v6.4.6 (ONT), Prowler (commit ID c3041ba)  under BioProject accession number"}
+{"text": "Urinary tract infections (UTIs) are common healthcare-associated and community-acquired bacterial infections in children. Data on pediatric UTIs in the Gulf Cooperation Council (GCC) region  have not been collated. Our aim is to review the published literature on the risk factors, etiology, antimicrobial susceptibility, and treatment of pediatric (aged <18 years) UTIs from healthcare and community settings in the GCC countries. Up toEscherichia coli accounts for 80%\u201390% of pediatric UTIs (E. coli urinary isolates from children in the Gulf Cooperation Council (GCC) region  (Uropathogenic s (UAE)] . Furthers (UAE)] .The challenges of managing pediatric UTIs in the GCC region have not been documented in a single article. Therefore, this review will collate data on risk factors, pathogens, resistance phenotypes, and antimicrobial management practices for pediatric UTIs within the region. Our objective is to provide a comprehensive assessment of the current knowledge on pediatric UTIs to improve diagnostic accuracy and help clinicians make rational therapeutic choices for UTIs, to improve the health of children in the region.2.2.1.This review evaluated relevant studies published between 1st January 2011 and 31st March 2022 on the PubMed and Google Scholar databases. We excluded non-English language articles, case reports, letters, books, editorials, notes, and conference abstracts.The primary search terms were \u201curinary tract infection\u201d and \u201cpediatric\u201d, followed by secondary terms relating to the review topics shown in . A free 2.2.Following initial identification and screening of publication titles, 38 full-text articles remained . Nine articles  were excluded for not differentiating data for extraction.The final GCC study number totaled 28: 24 retrospective studies \u201334 and 43.3.1.The Saudi Pediatric Infectious Diseases Society (SPIDS) guidelines on community-acquired UTI (CA-UTI) recommend transurethral catheterization for collecting urine from infants and non-toilet-trained children to minimize bacterial contamination from the skin . Cathete3.2.SPIDS defines UTI as significant bacteriuria in a symptomatic patient (febrile and/or with urinary symptoms) and recurrent UTI as \u22652 episodes of symptomatic UTI within 12 months . Their tP\u2009=\u20090.001) (P\u2009=\u20090.044) and nitrite (P\u2009=\u20090.046) were significantly sensitive to detect E. coli UTI, compared with non-E. coli UTI , 2815, 2=\u20090.001)  26). Py. PyP\u2009=\u20090coli UTI  (26).3.3.SPIDS advocates renal and bladder ultrasonography as a safe, noninvasive procedure for the detection of abnormalities and renal infections in preference to micturating or voiding cystourethrogram (MCUG or VCUG) . Renal u3.4P\u2009<\u20090.001)] (The predominant symptom was fever (50.3%\u201386.6%), with a regional prevalence of 60.7%   15, 16,, 27, 32.\u20090.001)]  (32). Tw\u20090.001)] , 35.P\u2009=\u20090.002), whereas rates of abdominal pain and urinary symptoms were higher in older children [2\u201314 years (P\u2009<\u20090.001)] (P\u2009<\u20090.001) (Other common symptoms (>20% of each study cohort) were urinary frequency, urgency or dysuria, nausea, vomiting, and abdominal pain  15, 16,,16, 25. <\u20090.001)  32)..P\u2009=\u20090.00Blood C-reactive protein (>5\u2005mg/L) did not significantly change with patient age  or betwe4.4.1.4.1.1.The prevalence of pediatric UTI across four studies including adults was 18.6%\u201368.7% , 33. In 4.1.2.P\u2009<\u20090.001), P\u2009=\u20090.001)]. Boys presented with CA-UTI at a significantly younger age than girls  of UTI patients were girls. More girls than boys had a UTI in 15 studies  11, 12,, 36, 38; of UTI pan girls  (11).4.1.2.1In other studies, being uncircumcised was associated with UTI , 41. In 4.2.4.2.1.P\u2009=\u20090.018), antibiotic use (P\u2009=\u20090.015) and VUR (P\u2009=\u20090.019) were associated with UTI (P\u2009=\u20090.02) and duration of fever of >36\u2005h (P\u2009=\u20090.001), but not with the isolated uropathogen ..P\u2009=\u20090.01P\u2009=\u20090.05) (P\u2009=\u20090.025) and age >2 months (P\u2009=\u20090.006) (P\u2009<\u20090.0001)  were associated with UTI (\u2009=\u20090.05) . UTI amo=\u20090.006)  14). In. InP\u2009=\u20090\u20090.0001) .Following cardiac surgery, 7.0% of patients had UTIs  . Prolong4.2.2.E. coli, non-E. coli UTIs were associated with hospitalization >7 days (P\u2009=\u20090.042), male sex (P\u2009<\u20090.0001), younger age [<4 years (P\u2009=\u20090.01)], prior use of antibiotics (P\u2009=\u20090.011), or history of UTI (P\u2009=\u20090.012) (E. coli UTIs (E. coli species (E. coli (49%) and non-E. coli (51%) in patients with normal ultrasound were comparable (Compared with =\u20090.012)  28). Si. SiE. cooli UTIs  .4.2.3.E. coli, and ESBL-producing E. coli were associated with VUR in children (P\u2009=\u20090.044) ; in neonates than older children and infants (P\u2009=\u20090.016) (One study of adults and children in Saudi Arabia found that significantly more UTI patients had non-ESBL-producing than ESBL-producing =\u20090.044)  12). Th. ThE. co=\u20090.016) ; and in =\u20090.016) . In cont=\u20090.016)  22). An. AnE. co=\u20090.016)  and 40.64.2.4.P\u2009<\u20090.005)]   13, 15,, 32. The\u20090.005)] .5.5.1.E. coli was predominant (26.1%\u2013100% of isolates), with a regional prevalence of 64.9%   and 19.8% (37 of 187 K. pneumoniae), respectively (P\u2009=\u20090.05) (thogens) , 32\u201334. thogens)  11, 12,thogens) . The reg,thogens ectively , 32\u201334. \u2009=\u20090.05)  (11).Klebsiella spp. (predominantly Klebsiella pneumoniae) were the second most common uropathogens (3.8%\u201335.2% of isolates), with a regional prevalence for K. pneumoniae of 13.8%  (thogens) , 27, 30.Pseudomonas aeruginosa, Proteus mirabilis, Enterococcus spp., and Enterobacter spp.  and cefazolin (94.0%) in Bahrain and Saudi Arabia  (P\u2009<\u20090.001) for recurrent UTIs , 34E. coi Arabia  among E. to 2020  (34). Moeropenem . In Omanent UTIs .E. coli from children in Bahrain (E. coli) from Qatar were resistant to ceftriaxone (More than 96% of ESBL-producing  Bahrain  34) and andE. cotriaxone  36)..E. coli K. pneumoniae, >90% of isolates were susceptible to cefotaxime, ceftriaxone, and meropenem  was 26.9% among enterococci  from recurrent UTIs (P\u2009<\u20090.001) . CoE. coli, 83%) from Qatar were resistant to amikacin, with higher resistance to nitrofurantoin (13.0%), gentamicin (24.4%), and trimethoprim-sulfamethoxazole (59.7%) (E. coli from Bahrain was highest to nitrofurantoin and fosfomycin (71.0%\u2013100%) (No ESBL-producing isolates ( (59.7%)  36). Su. SuE. co0%\u2013100%)  33). Al. AlE. co0%\u2013100%) .K. pneumoniae, >90% of isolates were susceptible to the tested aminoglycosides . Th. ThE. co5.3.E. coli isolates (61.1%) harbored the blaCTX\u2212M\u2212G1 gene, whereas 46.2% of ESBL-producing K. pneumoniae carried blaSHV, blaTEM and blaCTX\u2212M\u2212G1 genes (Most pediatric ESBL-producing G1 genes  36). Th. ThE. coE. coli and K. pneumoniae (52.2% of isolates were urinary) (Similar findings were demonstrated by a study in Qatar of pediatric ESBL-producing urinary) . CTX-M gurinary) .E. coli; 30% K. pneumoniae) in Qatar revealed that carbapenem resistance was largely attributed to a limited number of OXA-48-like and NDM carbapenemases, suggesting that certain international high-risk clones associated with other carbapenemase genes such as blaKPC-type, have not spread locally , mostly a cephalosporin (43.5%) or penicillin (44.2%), namely amoxicillin-clavulanic acid (29.4%) or cefprozil (23.0%)  19). In. In19). 7.The evidence presented in this review indicates limited published data from the UAE and a lack of studies on HA-UTIs. Also, most studies specified a patient age of <14 years, while four studies included patients aged 14\u201318 years , 33. GCC8.E. coli (including ESBL-producing isolates). ESBL-producing uropathogens are an established cause of UTI in the region, whereas CRE organisms  are emerging  should be reserved for critically ill children, particularly those having a previous UTI caused by, or other risk factors for, ESBL-producing uropathogens. Alternatively, ceftazidime-avibactam, a novel cephalosporin/\u03b2-lactamase inhibitor combination that inactivates ESBL and OXA-48-type enzymes, could be considered a treatment option for ESBL pyelonephritis (Empirical use of aminoglycosides due to the high prevalence of uropathogens resistant to 3GC has been proposed in Kuwait . Gentamiountries , 36. Theountries , 48\u201351. ephritis .E. coli in our region.The treatment of UTIs caused by carbapenemase-producing uropathogens in our region is challenging. Although aztreonam is stable against NDM and 3GC against OXA-48 enzymes, none of these agents can be empirically used to treat UTI caused by NDM and OXA-48 producers as most isolates in our region co-produce CTX-M-type ESBL . It coulIn conclusion, our review showed that UTIs are increasingly caused by antimicrobial-resistant uropathogens in the pediatric population of GCC countries. Further epidemiological and clinical studies are needed to optimize diagnostic and antimicrobial stewardship strategies in pediatric UTIs in the GCC region."}
+{"text": "Correction: Microbiome 10, 226 (2022)https://doi.org/10.1186/s40168-022-01430-9Following publication of the original article , the autThe updated additional file is included here and original article has been updated.Additional file 1:Fig. S8.\u00a0L. reuteri improves metabolic control in DIO mice treated with CQA. HFD-fed mice were treated twice per week with L. reuteri + CQA  by oral gavage for 5 weeks. Related to Fig. 6. (a) GTT and AUC. (b) Serum HDL-C. (c) Serum LDL-C. (d) Liver weight. (e) Hepatic mRNA expression of lipid synthesis-related genes.  Representative FL-IR images and BAT temperature. (h) Relative mRNA expression of thermogenic genes in BAT. (i) Representative H&E (upper) and UCP1 (lower) staining of BAT sections, scale bar: 50 \u03bcm. n = 8/group. Data are presented as mean \u00b1 SD. *, p < 0.05; **, p < 0.01; and ***, p < 0.001. ns means not statistically significant."}
+{"text": "Dicranomyia (Erostrata) jejuensissp. nov. and D. (E.) koreanasp. nov., from Korea are described on the basis of morphology and mitochondrial COI sequences. DNA barcode sequences for other four D. (Erostrata) species from Korea are also provided for the first time. The identification key for all known D. (Erostrata) species is presented.Two new crane fly species,  Dicranomyia Stephens, 1829, is the largest genus of the Limoniidae and, as such, contains 1,136 species and 24 subgenera, including the subgenus D. (Erostrata) Savchenko, 1976. Twelve species of this subgenus have been reported from the Palearctic, Nearctic, and Oriental regions (D. (E.) globithorax has been reported in fungus on decaying logs  tabashii (as Limonia (Limonia) tabashii) from Suigen (= Suwon) in Korea, and Dicranomyia species of the subgenus Erostrata including three new species and suggested that Limoniacongesta Alexander, 1976 and Limoniastriopleura Edwards, 1919 be classified as members of subgenus Erostrata based on non-genitalic characters.D. (Erostrata) crane fly species are described from Korea, providing identification for all Korean members of the subgenus. A DNA barcode (COI) dataset for six Dicranomyia species of the subgenus Erostrata from Korea is also presented for the first time.Here, two new Crane fly adults were collected using insect nets or Malaise traps and preserved in 80% ethanol Table . Wings aThe terminologies used to describe the morphology generally follow KUEM \u2013 Korea University Entomological Museum, Seoul, Republic of Korea;NIBR \u2013 National Institute of Biological Resources, Incheon, Republic of Korea.Specimen depositories are as follows:COI sequences were amplified and sequenced following COI. All sequences were submitted to GenBank .Total genomic DNA was extracted from the leg muscle of using the DNeasy Blood & Tissue Kit  according to the manufacturer\u2019s instructions. COI sequences (Table D. (Erostrata) species (10 sequences), and the outgroup species D. (Dicranomyia) kandybinae Savchenko, 1987 (1 sequence). Phylogenetic analyses were conducted using the neighbor-joining (NJ) method and Kimura-2-parameter model (p-distances) in MEGA X, using the complete deletion option.DNA barcode analysis was performed using 11 er model , with 1,er model . SequencDicranomyia (Erostrata) canis Dicranomyia (Erostrata) cnephosa Dicranomyia (Erostrata) congesta Dicranomyia (Erostrata) cynotis Dicranomyia (Erostrata) globithorax Osten Sacken, 1869Dicranomyia (Erostrata) globulithorax Alexander, 1924Dicranomyia (Erostrata) jejuensis sp. nov.Dicranomyia (Erostrata) koreana sp. nov.Dicranomyia (Erostrata) melas Dicranomyia (Erostrata) reniformis Kato, Tachi & Gelhaus, 2018Dicranomyia (Erostrata) striopleura Dicranomyia (Erostrata) submelas Kato, Tachi & Gelhaus, 2018Dicranomyia (Erostrata) tabashii Dicranomyia (Erostrata) yazuensis Kato, Tachi & Gelhaus, 2018Limoniidae Speiser, 1909Family Limoniinae Speiser, 1909Subfamily Taxon classificationAnimaliaDipteraLimoniidae\ufeffSubgenusSavchenko, 1976C69BCB6D-4C3F-5FAE-A61B-87A541D8AD53Dicranomyia (Erostrata) Savchenko in Dicranomyiaglobithorax Osten Sacken, 1869 .Rostrum is very short or reduced. Number of palpomeres ranges from one to three. Wings have no patterns, even in stigmal region. Third and fourth tarsomere are slightly swollen. Internal sac or notch is located on the male seventh sternite. Gonocoxite has ventromesal lobe. Gonostylus is one paired, with one or two setae arising from small tubercle on outer surface.Taxon classificationAnimaliaDipteraLimoniidae\ufeff15076BEE-3A44-562E-8FC1-6CF43A4FF557https://zoobank.org/9659D5F8-C578-4FAA-B16A-6AF8A9AE39E5Holotype: Korea \u2022 \u2642; Jeju-do, Seogwipo-si, Namwon-eup, Sillye-ri, Iseungi-oreum Volcanic Cone; 33\u00b020.24'N, 126\u00b037.25'E; alt. 450 m; 4 Aug.\u20138 Sep. 2021; Y. J. Bae leg.; Malaise trap; GenBank: OM102981; CF21-0150H; NIBR.Paratypes: Korea \u2022 1 \u2640; same data as holotype, 14 Jul.\u20134 Aug. 2021; GenBank: OM102983; CF21-0151; KUEM \u2022 1 \u2642; same data as holotype; GenBank: OM102982; CF21-0150P; KUEM.Palpus is 3-segmented. Male seventh sternite has shallow V-shaped notch. Outer face of gonostylus has single seta arising from tubercle. Distal lobe of paramere has a hooked tip with a subapical process.Male (holotype). Body length 4.3 mm, wing length 4.6 mm, antenna length 0.9 mm. General body coloration pale yellow to yellowish brown . General body coloration brighter than male. Femur I 2.8 mm; II 3.2 mm; III 3.4 mm; tibia I 3.2 mm; I: 3.1 mm; III 3.4 mm; tarsus I 3.0 mm; II 2.6 mm; III 2.4 mm.Female terminalia  submelas.Adults of this species are found in deciduous forests with moss-covered rocks along intermittent, rocky mountain streams Fig.  and co-oAdults were collected from June through early September.Dicranomyia (E.) jejuensis sp. nov. is morphologically similar to D. (E.) yazuensis based on the male genital structures, but it can be distinguished by the following characters: pleuron entirely dull yellow ; palpus 3-segmented (vs 2-segmented); distal 1/2 of gonostylus tapered to tip ; posterior margin of male seventh sternite with shallow V-shaped notch (vs long triangular notch); distal part of paramere with hooked tip (vs straight tip).Taxon classificationAnimaliaDipteraLimoniidae\ufeff9F0ADF72-CF20-5AA1-80E4-B4473CD6363Chttps://zoobank.org/163BF75E-826C-4E64-AAE4-CFAA23F2E22DHolotype: Korea \u2022 \u2642; Jeju-do, Seogwipo-si, Hawon-dong, Mt. Hallasan; 33\u00b020.95'N, 126\u00b029.72'E; alt. 1220 m; 13 Jun.\u20134 Aug. 2021; Y. J. Bae leg.; Malaise trap; GenBank: OM102979; CF21-0148; NIBR.Paratypes: Korea \u2022 1 \u2642; Gyeonggi-do, Gapyeong-si, Buk-myeon, Jeokmok-ri, Garim-gyo (Br.); 37\u00b058.60'N, 127\u00b026.55'E; alt. 300 m; 25 Jul.\u20131 Aug. 2015; Y. J. Bae leg.; Malaise trap; published as D. (E.) tabashii by KUEM \u2022 2 \u2642\u2642, 1 \u2640; same data as for preceding; 2\u20138 Aug. 2015; published as D. (E.) tabashii by KUEM \u2022 1 \u2642; same data as for preceding; 23\u201329 Jul. 2016; published as D. (E.) tabashii by KUEM \u2022 1 \u2642; Gangwon-do, Inje-gun, Girin-myeon, Bangdong-ri, Mt. Bangtaesan; 37\u00b054.50'N, 128\u00b024.41'E; alt. 690 m; 30 Jul.\u201316 Sep. 2019; Y. J. Bae leg.; Malaise trap; KUEM \u2022 1 \u2642; Gyeonsangnam-do, Sancheong-gun, Sicheon-myeon, Jungsan-ri, Jungsan-ri Campsite, Mount Jirisan; 35\u00b018.63'N, 127\u00b045.09'E; alt. 700 m; 28 Jul. 2021; J. Kim, C. Lim, D. Lee, W. Lee leg.; sweeping; GenBank: OP081140; CF21-0152; KUEM.Palpus is 3-segmented. Center of male seventh sternite has a deep conical internal sac that has a wide, round entrance. Outer face of gonostylus has two setae arising from a small tubercle. Paramere is elongated and narrow, distally with a darkened tip.Male (holotype). Body length 3.5 mm, wing length 4.5 mm, antenna length 0.7 mm. General body coloration yellow . General body coloration lighter than male.Female terminalia  globulithorax on Mount Bangtaesan and with D. (E.) tabashii on Mount Jirisan.This species is found along intermittent mountain streams in moist mixed forests with grassy vegetation Fig.  and in wAdults are mainly active from July through August.D. (E.) koreana sp. nov. is similar to D. (E.) tabashii, but it can be distinguished by the following characters: palpus 3-segmented (vs 2-segmented); male seventh sternite with weakly darkened, conical internal sac with round entrance ; paramere with darkened tip (vs without). This species is also similar to another species, D. (E.) jejuensis sp. nov. based on the male genital structures, but it can be distinguished by the following characters: male seventh sternite with a deep, conical internal sac ; gonostylus with two setae from tubercle (vs a single seta); paramere without hook at tip (vs with hook).In terms of the shape of the male terminalia, D. (E.) koreana sp. nov. from Mount Bangtaesan differs from other materials of the species based on the shape of the seventh sternite internal sack  and paramere distal lobe (pointed tip). However, additional specimens are needed to determine whether this difference is due to intra- or interspecific variation.The male genitalia of COI sequences contained 190 variable sites, of which 156 were parsimony-informative. The interspecific divergences (p-distances) within subgenus D. (Erostrata) ranged from 11.54% to 16.42%, with a mean distance of 13.17% across the entire dataset (Table D. (E.) jejuensis sp. nov., 0.59% in D. (E.) koreana sp. nov., and 0.15% in D. (E.) tabashii. The maximum intraspecific genetic distance (0.59%) was much smaller than the minimum interspecific one (11.54%). The NJ tree   species. The present study identified two new species using both morphological and molecular data. According to the NJ tree  jejuensis sp. nov., D. (E.) koreana sp. nov., D. (E.) tabashii, and D. (E.) yazuensis can be distinguished from other members of their subgenus based on the shape and mesal face  of their gonostyli. Two hypotheses may be considered: i) this clade can be classified into morphological species groups, or ii) it can be elevated to a new subgenus. Additional materials are needed to more accurately reconstruct phylogenetic relationships within genus Dicranomyia.This is the first study to use DNA barcoding for the delimitation of the ree Fig. , the subD. (E.) tabashii by D. (E.) koreana sp. nov. Based on our observation, unknown cryptic species of crane flies could also be detected and identified using molecular data.Based on our morphological examinations of the materials, we also found that some specimens identified as"}
+{"text": "Granulomatous polyangiitis (GPA) is a rare autoimmune disease that can involve multiple systems throughout the body, including the ear, nose, upper and lower respiratory tracts. It is classified as an antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis. Telitacicept is a novel recombinant fusion protein targeting B-lymphocyte stimulator (BLyS). Telitacicept can inhibit the development and maturation of abnormal B cells by blocking BLyS, and inhibit the production of antibodies by abnormal plasma cells by blocking APRIL (A proliferation-inducing ligand), which is expected to become a new drug for the treatment of GPA. We report a 64-year-old man diagnosed at our hospital with GPA involving multiple systems including kidneys, lungs, nose and ears. Renal involvement was severe, with a clinical characteristic of rapidly progressive glomerulonephritis and a pathologic manifestation of crescentic nephritis with plasma cell infiltration. The patient was treated with hormones, immunoglobulins and cyclophosphamide (CYC) with the addition of telitacicept and a rapid reduction in hormone dosage. The patient\u2019s renal function improved significantly within a short period of time, and his hearing and lung lesions improved significantly. At the same time, he did not develop serious infections and other related complications. Our report suggests that short-term control of the patient\u2019s conditions is necessary in GPA patients with organ-threatening disease. Telitacicept combined with CYC and glucocorticoids may be an induction therapy with safety and feasibility. However, more clinical trials are needed to validate the efficacy and safety of the therapeutic regimen. Necrotizing vasculitis primarily involves small and medium-sized vessels and can involve life-threatening organs, such as alveolar hemorrhage. Rapidly progressive glomerulonephritis which manifests pathologically as crescentic glomerulonephritis, carries a high risk of progression to end stage renal disease (ESRD) or dialysis-dependent risk . GPA is Compared with normal people, naive B cells increased during AVV activity and GPA remission period , 5, and As we reported below, we observed favorable safety and efficacy of telitacicept in combination with conventional therapies  for the treatment of patients with severe, rapidly progressive renal impairment who with pathological manifestations of crescentic nephritis with plasma cell infiltration.22. He developed hearing loss and no purulent discharge in the external ear canal. No obvious abnormality was found in cardiopulmonary examination. Admission laboratory examination: Blood examination: white blood cells (WBC) 12.13 \u00d710^9/L (reference range 3.5-9.5*10^9/L), hemoglobin (Hb) 90 g/L (reference range 130-175 g/L); Urine examination: 5-10 red blood cells/HP were detected by microscopy (reference range 0-2/HP); 24-hour urinary protein quantification: 0.5-0.7 g/24h (reference range 0-150 mg/24h); Erythrocyte sedimentation rate (ESR): 29 mm/h (reference range 0-43.5 mm/h) C-reactive protein 118 mg/L (reference range<10 mg/L); Biochemical indicators: Urea 16.3 mmol/L (reference range 3.1-8.0 mmol/L), Creatinine (CRE) 382 \u03bcmol/L (reference range 57-97 umol/L), albumin 27 g/L (reference range 40-55 g/L). pANCA (+/-) (reference range -), MPO (++) (reference range -); Immunoglobulin indicators: IgG 19.9 g/L (reference range 7.0-16.0 g/L), IgA 1.35 g/L (reference range 0.7-4.0 g/L), IgM 0.436 g/L (reference range 0.4-2.3 g/L). Imaging examination: Lung CT: 1. Multiple nodules and spots in both lungs, and neoplastic lesions cannot be excluded; 2. Multiple lymph nodes enlargement in the mediastinum  see Figure\u00a013Systemic vasculitis diseases are considered as diseases with multisystem involvement. GPA commonly involves the ear, nose, upper and lower respiratory tracts . In 2022It has been shown that GPA is predominantly characterized by C-ANCA antibody positivity 73.3-76%) and PR-3 ANCA 74-82.2%) antibody positivity, and also by P-ANCA antibody positivity (9.8-13%) and MPO-ANCA antibody positivity (5-8.1%) .The datasets presented in this article are not readily available because this is a case report. Requests to should be directed to Liqi Huang  for the publication of any potentially identifiable images or data included in this article.LH: Investigation, Project administration, Writing \u2013 original draft. WL: Investigation, Visualization, Writing \u2013 original draft. YLiu: Investigation, Methodology, Writing \u2013 original draft. JZ: Resources, Writing \u2013 review & editing. YLi: Conceptualization, Resources, Writing \u2013 review & editing. ZZ: Conceptualization, Funding acquisition, Writing \u2013 review & editing. CT: Conceptualization, Funding acquisition, Writing \u2013 review & editing."}
+{"text": "Correction: Clinical Epigenetics (2023) 15:105 10.1186/s13148-023-01521-wThe wrong Additional file 4 was originally published with this article . It has The original article has been corrected.Additional file 4. Table S6. Application for NNLS model with 6 CpGs."}
+{"text": "Scientific Reports 10.1038/s41598-022-15180-z, published online 07 July 2022Correction to: The original version of this Article contained an error in the Acknowledgements section.\u201cI.I. acknowledges the partial financial support from the SONATA BIS project 2020/38/E/ST5/00176.\u201dnow reads:\u201cI.I. was partly funded by the NCN SONATA-BIS Program (UMO-2020/38/E/ST5/00176).\u201dThe original Article has been corrected."}
+{"text": "Dear Editor,. Some previous studies had suggested an association between higher interleukin(IL)-6 levels and mortality risk in prevalent HD patients. The influence of serum IL-6levels on nutritional status remains, however, to be elucidated. The aim of this studywas to evaluate the association between serum IL-6 levels and nutritional status, andits impact on short-term mortality in prevalent HD patients.Inflammation has been associated with increased mortality in hemodialysis (HD) patients+), and calcium (Ca2+). PTH was measured byelectrochemiluminescence and IL-6 levels were measured by ELISA. Baselinecharacteristics and laboratory data were compared among groups using the Student\u2019st-test for normally distributed continuous variables, Mann-Whitney U-test for skeweddistributed continuous variables, and Chi-square test for categorical variables.One-year all-cause mortality was assessed using standard survival methods. Statisticalanalysis was performed using SPSS (Version 23 for Mac OSX).We conducted a retrospective single-center study of prevalent HD patients between January2020 and January 2021. Patients with acute or chronic inflammatory or infectious diseaseand active malignancy were excluded. According to the mean IL-6 level measured atbaseline, patients were categorized into two cohorts: IL-6 \u2264 17pg/mL and IL-6 >17pg/mL. Baseline clinical and demographic data were collected. Laboratory parameterswere measured before the second HD session of the week, including hemoglobin (Hgb),ferritin, serum iron, transferrin saturation index (TSI), albumin, C-reactive protein(CRP), phosphorus (Pp = 0.045), had higher c-reactive protein(CRP) , lower serum iron, and lower transferrinsaturation index (TSI) .Those patients had also higher serum ferritin levels, although not statisticallysignificant . Regarding nutritional parameters, patients with IL-6 > 17 pg/mL had loweralbumin , lean mass index(LMI) , bodymass index (BMI) , and cholesterol , but without statistical significance. During the study follow-up period, 9(14.8%) patients died. One-year survival was 91.4% in the group with IL-6 \u2264 17 pg/mL and64.3% in those with IL-6 > 17 pg/mL .Using a Cox proportional hazards analysis, we observed that patients with IL-6 > 17pg/mL had an increased risk of all-cause mortality at one year compared to those withIL-6 \u2264 17 pg/mL .The mean age of the included 61 prevalent HD patients was 74.96 \u00b1 12.89 years, 34 (55.7%)were male, and 25 (41%) were diabetic. Mean IL-6 level was 17.3 \u00b1 28.19 pg/mL. Fourteenpatients (23%) had IL-6 > 17pg/mL. There was no gender, comorbidity, HD vintage, ormodality difference among groups. Patients with IL-6 > 17pg/mL were older , and cardiovascularmorbidity to be interrelated in HD patients, each additionally contributing to highermortality in prevalent HD patients."}
+{"text": "Correction: Psicol. Refl. Cr\u00edt. 36, 19 (2023) https://doi.org/10.1186/s41155-023-00262-2Following publication of the original article (Fitzpatrick et al., Maira Almeida Lopes to Ma\u00edra Lopes Almeida, Giana Frizzo to Giana Bitencourt FrizzoThe original article (Fitzpatrick et al.,"}
+{"text": "Long-acting cabotegravir and rilpivirine (CAB/RPV) offers a promising alternative to daily oral antiretroviral therapy (ART); however, system-level and individual challenges in wide scale implementation are anticipated. We describe a community, infusion center-based model (ICBM) for administration of CAB/RPV and associated clinical outcomes.This was a single-center, retrospective cohort study of adults with HIV who were referred for enrollment in the ICBM  from 3/1/22 to 2/28/23 . We investigated demographics, system-level implementation variables, individual factors, and clinical outcomes among CAB/RPV recipients.79 patients were referred for enrollment in the ICBM and 64 patients received at least 1 dose of CAB/RPV. Reasons for CAB/RPV not being administered include insurance barrier, N=2 (13.3%); changed mind, N=3 (20%); unable to reach, N=2 (13.3%), difficulty with collecting labs, N=1 (6.7%), and CAB/RPV started outside of study period, N=4 (26.7%). Of those who received at least one dose of CAB/RPV, 79.9% were male, 48.4% were African American, 42.4% were Medicaid beneficiaries, 67.2% reported mental illness and 39.1% reported alcohol or non-tobacco substance use. Other baseline factors are listed in Table 1. Mean time from referral to first injection was 38.8 days excluding oral lead in time. Mean treatment duration was 176 days (range 20-326 days). 211 maintenance injections were administered during the study period; 16 (7.6%) were outside of the injection window with 10 (62.5%) oral bridges administered. All patients were virally suppressed at end of study period. Overall, 1,122 interventions were completed . Table 2 describes interventions performed.Implementation of CAB/RPV requires a collaborative effort to address system-level and individual challenges. Utilizing existing infrastructure allows for resource optimization to engage vulnerable populations and enhance equitable access. Our program shows successful treatment with CAB/RPV in individuals with adherence barriers (Table 3) at community infusion centers.Carlos Malvestutto, MD MPH, Pfizer: Advisor/Consultant|ViiV Healthcare: Advisor/Consultant"}
+{"text": "Outpatient parental antibiotic therapy is safe and effective. Our institution is one of two tertiary public hospital pediatric centers in Singapore. We aim to describe characteristics and outcomes of pediatric OPAT (p-OPAT).We retrospectively reviewed p-OPAT patients from May 2013-Dec 2022. Clinically stable inpatients were referred to the pediatric infectious disease team for assessment, caregiver training, and weekly review. Two main delivery models were used; daily bolus/short infusion at an outpatient unit, and home self-administration via elastomeric device. Electronic medical records were analyzed for demographics, clinical and microbiological data, treatment, and outcomes. Institutional review board approval and written informed consent were waived.219 patients completed 249 p-OPAT episodes; 46.6% female, 53.3% male. Median age was 8 years 5 months (range 11 days-29 years). Patient ethnicities were 53.9% Chinese, 12.3% Indian, 20.5% Malay, 13.3% others.53/219 (24.2%) patients were immunocompromised; three had primary immunodeficiency, 50 had acquired immunodeficiency. 89/219 (40.6%) patients had chronic illness.176/249 (70.7%) episodes had only one pathogen identified (Tab 1). Pathogens were gram positive (21.7%), gram negative (38.2%), fungal (5.2%), mycobacterial (5.6%) (Tab 2). 18.9% episodes were culture-negative, 1.6% episodes had no cultures. Infection sites varied considerably; osteoarticular infection +/- bacteremia were most common (Tab 3). Ceftriaxone was the most frequent antimicrobial . Peripherally inserted central catheter was most commonly used .Complete success, partial success, and failure were seen in 174 (69.9%), 43 (17.3%) and 32 (12.8%) episodes respectively. Main adverse events (AE) were vascular site dermatitis, infection, and line dislodgement. 235 (94.4%) episodes achieved infection cure. Mean p-OPAT duration was 16.9 days (range 1-177 days), saving 4208 admission bed days.We describe one of the largest series of children receiving p-OPAT over 10 years and report good outcomes for a broad spectrum of infections. Vascular access issues were common AEs despite preventive measures.All Authors: No reported disclosures"}
+{"text": "The use of medical 3D printing has expanded dramatically for breast diseases. A writing group composed of the Radiological Society of North America (RSNA) Special Interest Group on 3D Printing (SIG) provides updated appropriateness criteria for breast 3D printing in various clinical scenarios. Evidence-based appropriateness criteria are provided for the following clinical scenarios: benign breast lesions and high-risk breast lesions, breast cancer, breast reconstruction, and breast radiation (treatment planning and radiation delivery).The online version contains supplementary material available at 10.1186/s41205-023-00171-1. Currently, medical 3D printing is performed for a variety of indications. In 2018, the RSNA 3D printing SIG published appropriateness criteria for medical 3D printing for various clinical scenarios [The RSNA Special Interest Group (SIG) on 3D Printing Guidelines Committee has initiated and revised several documents regarding the Clinical scenarios for which 3D Printing is considered an appropriate representation or extension of data contained in a medical imaging examination. This document highlights appropriateness of medical 3D printing for clinical utilization, research, scientific and informational purposes within breast diseases. This work is loosely modeled after the American College of Radiology (ACR) Appropriateness Criteria\u00ae  in that 1\u20133, rarely appropriate. There is a lack of a clear benefit or experience that shows an advantage over usual practice.4\u20136, maybe appropriate. There may be times when there is an advantage, but the data is lacking, or the benefits have not been fully defined.7\u20139, usually appropriate. Data and experience show an advantage to 3D printing as a method to represent and/or extend the value of data contained in the medical imaging examination.The SIG Guidelines Chairperson oversees the ratings via a vote among Special Interest Group members at in-person meetings. The results of the ratings follow the following 1\u20139 format (with 9 being the most appropriate):Clinical scenarios included in this document are stratified by histopathologic diagnosis and treatment. These include benign breast lesions and high-risk breast lesions, malignant breast lesions, breast reconstruction, and breast radiation (treatment planning and radiation delivery).th, 2022 SIG Appropriateness Committee Meeting. Afterwards, 2-week period for comments by SIG members was posted on the SIG\u2019s members-only online forum. All included studies were graded with a strength of evidence assessment according to ACR Appropriateness Criteria Evidence Document.An exhaustive English language PubMed literature search was performed (May 2022) which enabled the querying and retrieval of pertinent clinical documents regarding the appropriateness of 3D printing in each of the scenarios. The supporting evidence was obtained through structured searches, as detailed in each category. For each category, from the pool of total results, the number of publications considered \u201crelevant results\u201d was curated by consensus between physicians with expertise in 3D printing and breast care. Relevant publications that were not retrieved by the structured PubMed search were manually entered. The following categories were excluded because they were considered outside the project scope . All final components of this section were vetted and approved by vote of Special Interest Group members virtually at the July 20Benign breast lesions and high-risk breast lesions: Benign breast diseases are common and include a wide range of entities [entities . The mosentities . Fibroadentities . Benign entities , 4.High-risk breast lesions may confer an increased risk for breast cancer, may be associated with a higher risk for future breast cancer, and may be precursors in breast carcinogenesis. Management of high-risk lesions after core needle biopsy may include close imaging and clinical follow-up or excisional biopsy to evaluate for cancer \u20138. High-Surgical management of these entities may be needed in cases where cosmesis is altered or when symptom relief is needed. Surgical management may impact developing breast tissue in young women leading to alterations in its proper development . TherefoPubMed Search: ((3D printing) AND (fibrocystic change)) OR ((3D printing) AND (benign breast masses)) OR ((3D printing) AND (mastitis)) OR ((3D printing) AND ) OR ((rapid prototyping) AND (fibrocystic change)) OR ((rapid prototyping) AND (benign breast masses)) OR ((rapid prototyping) AND (mastitis) OR ((rapid prototyping) AND ). ((3D printing) AND ) OR ((3D printing) AND ) OR ((3D printing) AND (lobular neoplasia)) OR ((3D printing) AND ) OR ((3D printing) AND (papillary lesions) OR ((3D printing) AND (mucocele-like lesions)) OR ((rapid prototyping) AND ) OR ((rapid prototyping) AND ) OR ((rapid prototyping) AND (lobular neoplasia) OR ((rapid prototyping) AND ) OR ((rapid prototyping) AND (papillary lesions)) OR ((rapid prototyping) AND (mucocele-like lesions))PubMed Results: No results found.Preliminary rating: 1Malignant breast lesions: Breast cancer is the most common solid malignancy in women in the United States [d States . Approxid States , 12. Bred States . Understd States . They ard States . 3D prind States . This ind States \u201319. AddiPubMed Search: ((3D printing) AND (breast cancer) OR ((rapid prototyping) AND (breast cancer))PubMed Results: 14 [aSchulz-Wendtland R, Harz M, Meier-Meitinger M, et al. Semi-automated delineation of breast cancer tumors and subsequent materialization using three-dimensional printing (rapid prototyping). J Surg Oncol. 2017;115(3):238\u2013242.bSantiago L, Volk RJ, Checka CM, et al. Acceptability of 3D\u2010printed breast models and their impact on the decisional conflict of breast cancer patients: A feasibility study. Journal of surgical oncology. 2021;123(5):1206\u20131214.cBarth RJ, Krishnaswamy V, Paulsen KD, et al. A Patient-Specific 3D-Printed Form Accurately Transfers Supine MRI-Derived Tumor Localization Information to Guide Breast-Conserving Surgery. Annals of Surgical Oncology. 2017;24(10):2950\u20132956.dRao N, Chen K, Yang Q, Niu J. Proof-of-Concept Study of 3-D-Printed Mold-Guided Breast-Conserving Surgery in Breast Cancer Patients. Clin Breast Cancer. 2018;18(5):e769-e772.eKo BS, Kim N, Lee JW, et al. MRI-based 3D-printed surgical guides for breast cancer patients who received neoadjuvant chemotherapy. Scientific Reports. 2019;9(1):11,991.fFernandez RAS, Lau RWH, Yu PSY, Siu ICH, Chan JWY, Ng CSH. Use of custom made 3-dimensional printed surgical guide for manubrio-sternal resection of solitary breast cancer metastasis: case report. AME Case Rep. 2020;4:12.gWu ZY, Alzuhair A, Kim H, et al. Magnetic resonance imaging based 3-dimensional printed breast surgical guide for breast-conserving surgery in ductal carcinoma in situ: a clinical trial. Sci Rep. 2020;10(1):18,534.hOck J, Lee S, Kim T, et al. Accuracy evaluation of a 3D printing surgical guide for breast-conserving surgery using a realistic breast phantom. Computers in Biology and Medicine. 2021;137:104,784.iWu ZY, Kim HJ, Lee J, et al. Breast-conserving surgery with 3D-printed surgical guide: a single-center, prospective clinical study. Sci Rep. 2021;11(1):2252.jWu ZY, Kim GB, Choi S, Lee S, Kim N, Ko B. Breast-Conserving Surgery after Neoadjuvant Chemotherapy Using a Three-Dimensional-Printed Surgical Guide Based on Supine Magnetic Resonance Imaging: A Case Report. J Breast Cancer. 2021;24(2):235\u2013240.kWu ZY, Lee YJ, Shin Y, et al. Usefulness of 3-Dimensional-Printed Breast Surgical Guides for Undetectable Ductal Carcinoma In Situ on Ultrasonography: A Report of 2 Cases. J Breast Cancer. 2021;24(3):349\u2013355.lWu ZY, Kim GB, Lee S, Choi SH, Kim N, Ko B. Case Report: A 3D-Printed Surgical Guide for Breast-Conserving Surgery After Neoadjuvant Chemotherapy. Front Oncol. 2021;11:633,302.mLee HS, Kim HJ, Chung IY, et al. Usefulness of 3D-surgical guides in breast conserving surgery after neoadjuvant treatment. Sci Rep. 2021;11(1):3376.nSantiago L, Adrada BE, Caudle AS, Clemens MW, Black DM, Arribas EM. The role of three\u2010dimensional printing in the surgical management of breast cancer. Journal of surgical oncology. 2019;120(6):897\u2013902.ults: 14 , 18\u201329aSPreliminary Rating: 7Breast reconstruction: Breast reconstruction surgeries include either implant-based or autologous flap reconstructions. In autologous flap reconstructions, 3D printed models have been shown to facilitate the intramuscular dissection of perforator vessels by depicting the course and trajectory of the subfascial vascular tree and allowing the surgeon to view the model from various vantage points [e points , 30, 31.e points . 3D modee points , 34.PubMed Search: ((3D printing) AND (breast reconstruction) OR ((rapid prototyping) AND (breast reconstruction)PubMed Results: 11 [aJablonka EM, Wu RT, Mittermiller PA, Gifford K, Momeni A. 3-DIEPrinting: 3D-printed models to assist the intramuscular dissection in abdominally based microsurgical breast reconstruction. Plastic and Reconstructive Surgery Global Open. 2019;7(4).bChae MP, Rozen WM, McMenamin PG, Findlay MW, Spychal RT, Hunter-Smith DJ. Emerging applications of bedside 3D printing in plastic surgery. Frontiers in surgery. 2015;2:25.cChae MP, Hunter-Smith DJ, Rostek M, Smith JA, Rozen WM. Enhanced preoperative deep inferior epigastric artery perforator flap planning with a 3D-printed perforasome template: technique and case report. Plastic and Reconstructive Surgery Global Open. 2018;6(1).dChae MP, Hunter-Smith DJ, Spychal RT, Rozen WM. 3D volumetric analysis for planning breast reconstructive surgery. Breast Cancer Res Treat. 2014;146(2):457\u2013460.eHummelink S, Verhulst AC, Maal TJ, Ulrich DJ. Applications and limitations of using patient-specific 3D printed molds in autologous breast reconstruction. European journal of plastic surgery. 2018;41(5):571\u2013576.fDeFazio MV, Arribas EM, Ahmad FI, et al. Application of Three-Dimensional Printed Vascular Modeling as a Perioperative Guide to Perforator Mapping and Pedicle Dissection during Abdominal Flap Harvest for Breast Reconstruction. J Reconstr Microsurg. 2020;36(5):325\u2013338.gMehta S, Byrne N, Karunanithy N, Farhadi J. 3D printing provides unrivalled bespoke teaching tools for autologous free flap breast reconstruction. Journal of Plastic, Reconstructive & Aesthetic Surgery. 2016;69(4):578\u2013580.hChae MP, Hunter-Smith DJ, Chung RD, Smith JA, Rozen WM. 3D-printed, patient-specific DIEP flap templates for preoperative planning in breast reconstruction: a prospective case series. Gland Surg. 2021;10(7):2192\u20132199.iChen K, Feng CJ, Ma H, et al. Preoperative breast volume evaluation of one-stage immediate breast reconstruction using three-dimensional surface imaging and a printed mold. J Chin Med Assoc. 2019;82(9):732\u2013739.jOgunleye AA, Deptula PL, Inchauste SM, et al. The utility of three-dimensional models in complex microsurgical reconstruction. Arch Plast Surg. 2020;47(5):428\u2013434.kTomita K, Yano K, Taminato M, Nomori M, Hosokawa K. DIEP Flap Breast Reconstruction in Patients with Breast Ptosis: 2-Stage Reconstruction Using 3-Dimensional Surface Imaging and a Printed Mold. Plast Reconstr Surg Glob Open. 2017;5(10):e1511.ults: 11 \u201340aJabloPreliminary rating: 8Breast Radiation (Treatment planning and Radiation delivery): 3D printing can be used to design customized patient specific boluses and shields that allow homogeneous distribution of radiation dose to the area of interest while sparing adjacent normal tissue [l tissue . 3D prinl tissue PubMed Search: ((3D printing) AND (breast radiation) OR ((rapid prototyping) AND (breast radiation)PubMed Results: 8 [aPoulin E, Gardi L, Fenster A, Pouliot J, Beaulieu L. Towards real-time 3D ultrasound planning and personalized 3D printing for breast HDR brachytherapy treatment. Radiother Oncol. 2015;114(3):335\u2013338.bAristei C, Lancellotta V, Piergentini M, et al. Individualized 3D-printed templates for high-dose-rate interstitial multicathether brachytherapy in patients with breast cancer. Brachytherapy. 2019;18(1):57\u201362.cYang K, Park W, Ju SG, et al. Heart-sparing radiotherapy with three-dimensional printing technology after mastectomy for patients with left breast cancer. Breast J. 2019;25(4):682\u2013686.dHe C, Zhang S, Shi L. Three-Dimensionally-Precise Breast Conformal Device for IMRT in Breast Cancer Patients Treated With Breast-Conserving Surgery-A Pilot Randomized Controlled Trial. Technol Cancer Res Treat. 2020;19:1,533,033,820,971,563.eRobar JL, Moran K, Allan J, et al. Intrapatient study comparing 3D printed bolus versus standard vinyl gel sheet bolus for postmastectomy chest wall radiation therapy. Pract Radiat Oncol. 2018;8(4):221\u2013229.fPark SY, Choi CH, Park JM, Chun M, Han JH, Kim JI. A Patient-Specific Polylactic Acid Bolus Made by a 3D Printer for Breast Cancer Radiation Therapy. PLoS One. 2016;11(12):e0168063.gPoulin E, Gardi L, Fenster A, Pouliot J, Beaulieu L. A novel approach for real-time, personalized breast HDR brachytherapy treatment using 3D printing technology. Brachytherapy. 2014;13:S18.hPark K, Park S, Jeon MJ, et al. Clinical application of 3D-printed-step-bolus in post-total-mastectomy electron conformal therapy. Oncotarget. 2017;8(15):25,660\u201325,668.sults: 8 , 42\u201348aPPreliminary rating: 6Table Limitations of this work include its lack of objective data collection and inferential statistics. Although such an analysis would be desirable, it is not practical with most published breast related applications due to the small number of publications and patients. PubMed search terms, were based on prior search terminology from previously published guidelines . The RSNThis document provides updated appropriateness criteria for 3D printing in breast conditions. Adoption of common clinical standards regarding appropriate use, information and material management, and quality control are needed to ensure the greatest possible clinical benefit from 3D printing. With accruing evidence for in 3D printing, this consensus guideline document, created by the members of the RSNA 3D printing Special Interest Group, will provide a reference for clinical standards of 3D printing. The document will be periodically refined, based on expanding clinical applications and growing medical literature.Additional file 1."}
+{"text": "Chronic obstructive pulmonary disease (COPD) is a multifactorial disease of the respiratory system which develops as a result of a complex interaction of genetic and environmental factors closely related to lifestyle. We aimed to assess the combined effect of the PI3K/AKT/mTOR signaling pathway  and sirtuin  genes to COPD risk. SNPs of SIRT1 , SIRT3 , SIRT6 (rs107251), AKT1 (rs2494732), PIK3R1 , MTOR , PTEN  genes were genotyped by real-time polymerase chain reaction (PCR) among 1245 case and control samples. Logistic regression was used to detect the association of SNPs in different models. Linear regression analyses were performed to estimate the relationship between SNPs and lung function parameters and smoking pack-years. Significant associations with COPD were identified for SIRT1 (rs3818292) , SIRT3 (rs3782116)  and SIRT3 (rs536715)  under the dominant model, SIRT6 (rs107251) , PIK3R1: , rs831125 , rs3730089 ), PTEN: , and rs2735343 ). A significant genotype-dependent variation of lung function parameters was observed for SIRT1 (rs3818292), SIRT3 (rs3782116), PIK3R1 (rs3730089), and MTOR (rs2536). Gene-gene combinations that remained significantly associated with COPD were obtained; the highest risk of COPD was conferred by a combination of G allele of the PIK3R1 (rs831125) gene and GG of SIRT3 (rs536715) (OR = 3.45). The obtained results of polygenic analysis indicate the interaction of genes encoding sirtuins SIRT3, SIRT2, SIRT6 and PI3KR1, PTEN, MTOR and confirm the functional relationship between sirtuins and the PI3K/AKT/mTOR signaling pathway. Chronic obstructive pulmonary disease (COPD) is a multifactorialrespiratory system disease that affects the distalparts of the respiratory tract  and lungparenchyma with lung emphysema development . \u0421hronic obstructive pulmonary disease developsas a result of complex interaction between molecular geneticand environmental factors, closely related to lifestyle, andsmoking is considered the main cause of COPD . Published data suggest that the COPD pathogenesismay involve dysregulation of stress responses that inhibitcellular senescence, which includes a wide range of signalingcascades and their regulators .The PI3K/AKT/mTOR intracellular signaling pathway isa universal pathway controlling cell growth, metabolism, andproliferation . The key components of thissignaling pathway are phosphatidylinositol-3 kinase (PI3K),serine/threonine protein kinase (AKT), and serine/threoninekinase  . Signal transduction through the PI3K/AKT/mTOR signaling cascade is essential for cellular senescence.This signaling pathway is inhibited by the tyrosine phosphatasesPTEN (phosphatase and tensin homolog) and SHIP-1(inositol polyphosphate-5-phosphatase D). Both enzymeshave oxidation-sensitive cysteine residue in the active region.Oxidative stress is the main mechanism that causes acceleratedsenescence through its damaging effects on DNA and thePI3K/AKT/mTOR signaling pathway activation . In COPD and other age-associated diseases, the expressionof genes encoding endogenous antioxidant molecules isreduced, which leads to an increased level of oxidative stressand cellular senescence activation .NAD-dependent protein deacetylases from the sirtuins familyare considered as potential factors that decrease senescence.We aimed to assess the combined effect of the PI3K/AKT/mTOR signaling pathway  andsirtuins  genes on COPD risk.DNA samples were collected from unrelated subjects whowere Tatars in ethnicity and resided in the Republic of Bashkortostan.The study was approved by the Ethics Committeeat the Institute of Biochemistry and Genetics . All participants of this study providedwritten informed consent. The COPD group included 621patients (539 (86.79 %) males and 82 (13.21 %) females) witha mean age of 64.42 \u00b1 10.71 years. There were 510 (82.13 %)smokers and former smokers and 111 (17.87 %) nonsmokersin the COPD group. The smoking index was 45.34 \u00b1 23.84pack years in the smokers and former smokers. The controlgroup included 624 subjects (555 (88.94 %) males and 69(11.06 %) females) with a mean age of 59.67 \u00b1 12.31. Therewere 526 (84.29 %) smokers and former smokers and 98(15.71 %) nonsmokers in the group; the smoking index was38.75 \u00b1 24.87 pack years in the smokersIn all patients, spirometry was performed to assess lungfunction, including vital capacity (VC), forced vital capacity(FVC), forced expiration volume in the first second (FEV1),and the FEV1/FVC ratio. The group of patients had the followingparameters : FEV1 = 40.75 \u00b1 18.33;FVC = 45.01 \u00b1 18.22; VC = 49.32 \u00b1 14.34; FEV1/FVC =59.5 \u00b1 12.34. Inclusion and exclusion criteria for the COPDand control have been previously described .Genotyping. DNA was isolated from peripheral bloodleukocytes by phenol-chloroform extraction. The set includedSNPs of the following genes: SIRT1 ,SIRT3 , SIRT6 (rs107251), AKT1(rs2494732), PIK3R1 ,MTOR , PTEN 1.http://vavilov.elpub.ru/jour/manager/files/Suppl_Korytina_Engl_27_5.pdfSupplementary Materials are available in the online version of the paper:The SNPs were selected for the study based on the followingcriteria: functional effect of SNP on gene expressionor relation to non-synonymous substitutions, and/or associationswith complex human diseases; minor allele frequency (MAF) of \u2265 5 % in the European populations according tothe National Center for Biotechnology Information database(http://www.ncbi.nlm.nih.gov/projects/SNP/). The functionalsignificance of the SNPs was verified using RegulomeDBVersion 1.1 (https://regulomedb.org), SNPinfo Web Server(https://snpinfo.niehs.nih.gov), and HaploReg v3 . Data were presented in Supplementary Material 2.SNP genotyping was performed by real-time polymerasechain reaction (PCR) using commercial kits for fluorescencedetection  anda BioRad CFX96TM instrument . The methods of analysis were described in detailpreviously .Statistical analyses. Statistical analyses of the results wereperformed using the software packages IBM SPSS Statis-tics 22.0 and PLINK v. 1.07 . The methodswere described in detail previously .Association analyses of allele or genotype combinationswith COPD were carried out using the APSampler 3.6.1program (http://sourceforge.net/projects/apsampler/). TheBenjiamini\u2013Hochberg correction for multiple testing wasperformed using special software (http://www.sdmproject.com/utilinies/?show=FDR) to decrease the false discoveryrate (FDR) and to obtain \u0420cor- FDR. The linkage disequilibriumstructure LD (D\u2019) and haplotype frequencies were calculatedwith Haploview 4.2.Before analyzing the association of candidate gene alleleswith COPD, we verified whether the genotype frequencydistributions corresponded to the Hardy\u2013Weinberg equilibriumand evaluated minor allele frequencies (MAF) both in thecombined group of patients and healthy subjects and in eithergroup individually . All studiedSNPs were in Hardy\u2013Weinberg equilibrium in the controlgroup: SIRT1 (rs3758391) (PX-B = 0.24), SIRT1 (rs3818292)(PX-B = 0.47), SIRT3 (rs3782116) (PX-B = 0.5), SIRT3(rs536715) (PX-B = 0.75), SIRT6 (rs107251) (PX-B = 0.67),AKT1 (rs2494732) (PX-B = 0.2), PIK3R1 (rs10515070)(PX-B = 0.65), PIK3R1 (rs831125) (PX-B = 0.25), PIK3R1(rs3730089) (PX-B = 0.22), MTOR (rs2295080) (PX-B = 0.15),MTOR (rs2536) (PX-B = 0.24), PTEN (rs701848) (PX-B = 0.85),PTEN (rs2735343) (PX-B = 0.06).The groups of COPD patients and healthy controls differedsignificantly in the genotypes and/or alleles frequencydistributions of SIRT1 (rs3818292), SIRT3 , SIRT6 (rs107251), AKT1 (rs2494732), PIK3R1, and PTEN  (Table 1). Statistically significant results ofassociation analysis of the studied gene loci and COPD areshown in Table 2.An association of SIRT1 (rs3818292) with COPD was establishedin the dominant model .The risk of COPD was increased in heterozygous individuals. An association of COPD with theheterozygous genotype  and in thedominant model of SIRT3 (rs3782116) and in the dominant, log-additive  models and with the heterozygous genotype of SIRT3 (rs536715). It shouldbe noted that in both cases the COPD risk was associatedwith the frequent G allele (rs3782116 \u2013 OR = 1.21 95 % CI1.03\u20131.43 \u0438 rs536715 \u2013 OR = 1.58 95 % CI 1.32\u20131.91) andthe GG genotype (rs3782116 \u2013 OR = 1.44 95 % CI 1.16\u20131.81\u0438 rs536715 \u2013 OR = 1.99 95 % CI 1.58\u20132.51).We carried out a linkage disequilibrium analysis of thers3758391 and rs3818292 loci of the SIRT1, rs3782116 andrs536715 of the SIRT3, which showed the absence of linkagedisequilibrium between the loci of the SIRT1 gene  and the SIRT3 gene . Basedon the obtained data, haplotypes association analysis wasnot performed. Association of SIRT6 (rs107251) with thedevelopment of COPD was detected in the dominant model, but the association with the heterozygousCT genotype was more significant . The risk of COPD was associated with the CC ge-notype of SIRT6 (rs107251) (OR = 1.54 95 % CI 1.23\u20131.93).We have identified the association SNPs of the PIK3R1 gene with COPD. The risk ofCOPD was associated with heterozygous genotypes of thestudied SNPs of the PIK3R1 gene: rs10515070 , rs831125  and withthe GG genotype of rs3730089 .We showed the absence of linkage disequilibrium betweenrs10515070, rs831125, rs3730089 of the PIK3R1 gene:for rs10515070 and rs831125 , forrs10515070 and rs3730089 , forrs831125 and rs3730089 .Based on the obtained data, haplotypes association analysiswas not performed. Association of PTEN (rs701848)and COPD was established in the dominant , recessive  andlog-additive models Padj = 0.0015, OR = 1.35). We haveidentified the association of PTEN (rs2735343) with COPD inthe log-dominant model  and for theheterozygous genotype . Linkagedisequilibrium between the rs701848 and rs2735343 was notdetected , thus, haplotype associationanalysis was not performed.Association of the studied genes lociand quantitative phenotypes with lungfunction parameters and smoking indexLung function decline is a key clinical feature of airwayobstruction in COPD that indicates progression of the disease.We investigated the relationship between the studied genes lociand lung function parameters: Forced Vital Capacity (FVC),Forced Expiration Volume in 1 s (FEV1), and FEV1/FVCratio in COPD patients (Table 3). The heterozygous genotypeof PIK3R1 (rs3730089) (P = 0.013) and the TT genotypeof MTOR (rs2536) (P = 0.013) were associated with adecrease in the FVC value. Carriers of the SIRT3 (rs3782116)AA genotype exhibited a higher FVC value (P = 0.0015).Individuals that presented the AA genotype of SIRT1(rs3818292) (P = 0.017), the GG genotype of PIK3R1(rs3730089) (P = 0.025), and the AA genotype of SIRT3(rs3782116) (P = 0.0028) showed a significant increase in theirFVC. Meanwhile, carriers of the heterozygous genotypes ofSIRT1 (rs3818292) (P = 0.04), MTOR (rs2295080) (P = 0.025),and SIRT3 (rs3782116) (P = 0.016) exhibited a lower FVC value (see Table 3). The carriers of the heterozygous genotypeof PIK3R1 (rs831125) (P = 0.0082), the AA genotype ofSIRT1 (rs3818292) (P = 0.036) had a significantly highersmoking index.Analysis of gene-gene interactionsUsing the APSampler algorithm, we have identified gene-genecombinations significantly associated with \u0441hronic obstructivepulmonary disease. In order to identify significant interactionsof functionally related sirtuins genes, SIRT2 (rs10410544) wasincluded in the analysis . We obtained2324 patterns associated with \u0441hronic obstructive pulmonarydisease. Table 4 shows the results of the most significant genegenecombinations with PFDR less than 0.05 and OR morethan 2.0 for risk combinations) or less than 0.35 for protectivecombinations. A total of 19 gene-gene combinations fulfilledthe above-mentioned criteria. Nine patterns were associatedwith an increased risk of COPD; ten were protective. Allele Gand genotype GG of PIK3R1 (rs831125) contributed to themost significant combinations associated with COPD risk(four patters).The highest risk of COPD was conferred by the combinationof these variants of PIK3R1 (rs831125) with the GG genotypeof SIRT3 (rs536715) (OR = 3.45); and with the C alleleof PTEN (rs2735343) (OR = 3.06) and their combination:genotype GA of PIK3R1 (rs831125) together with the G alleleof SIRT3 (rs536715) and the C allele of PTEN (rs2735343)(OR = 2.86). The analysis on the gene-gene interaction of thestudied gene loci established an association of the T alleleof MTOR (rs2536) only in combinations with the G alleleof PIK3R1 (rs831125) (OR = 2.71). Three of the identifiedpatterns included the G allele of PIK3R1 (rs3730089) in combination with the GG genotype of SIRT3 (rs536715)(OR = 2.09), or with the CC genotype of SIRT6 (rs107251)(OR = 2.01).The most significant protective patterns included the A alleleor the AA genotype of PIK3R1 (rs831125) and the A alleleof PIK3R1 (rs3730089) in combination with the A allele orthe AG genotype of SIRT3 (rs3782116) and the A allele ofSIRT3 (rs536715) (see Table 4). Thus, the PIK3R1 (rs831125),PIK3R1 (rs3730089), and SIRT3 (rs536715) loci exhibited anallele-specific effect, when the G allele of PIK3R1 (rs831125),the G allele of PIK3R1 (rs3730089), and the G allele and theGG genotype of SIRT3 (rs536715) were part of risk patterns,and alternative alleles of the same polymorphic loci werepresent in protective patterns.As a result our study, significant associations between theSIRT1 (rs3818292), SIRT3 , SIRT6(rs107251) polymorphic variants and COPD were found.SIRT1 is the most studied member of the mammalian sirtuinfamily. It has been shown that SIRT1 plays an important rolein signaling pathways involved in cellular senescence andcell death . SIRT1 deacetylates manykey regulatory proteins and transcription factors involved inDNA repair, inflammation, expression of antioxidant genes,and cellular senescence, including the PI3K/AKT/mTORsignaling pathway genes, transcription factor FOXO3a, p21,p16, Klotho proteins .Increased expression of SIRT1 inhibits the TGF-\u03b21/SMAD3signaling pathway and impairs epithelial-mesenchymal transformation,which leads to a decrease in COPD-associatedairway remodeling . SIRT1 levels arereduced in peripheral pulmonary and peripheral blood mononuclearcells of patients with COPD .We found that the COPD risk is higher in heterozygouscarriers of SIRT1 (rs3818292). Moreover, this polymorphicvariant demonstrates an association with a decrease in FV\u04211, which reflects the progression of the disease. Functionalanalysis showed that SIRT1 (rs3818292) is in linkage witha SNP in the 5\u2032-untranslated DNA region (rs3740051) thatchanges the NFKB transcription factor binding site. We didnot associate SIRT1 (rs3758391) with COPD. The results ofour study are in agreement with the previously published datareported for the Han Chinese population .According to functional analysis, rs3758391 is located in thepromoter region of the SIRT1 gene, and the C variant disruptsbinding sites for several transcription factors and regulatoryproteins, affecting gene expression. The role of rs3758391in the development of age-associated diseases is well known.Mitochondrial dysfunction in respiratory epithelial cellsplays an important role in the pathogenesis of COPD . SIRT3 is the main mitochondrial deacetylaseregulating many enzymes involved in energy metabolism,respiratory chain components, the tricarboxylicacid cycle,ketogenesis, and fatty acid beta-oxidation .SIRT3 can directly control the production of reactive oxygenspecies (ROS) by deacetylating manganese superoxidedismutase (SOD2), the main mitochondrial antioxidant enzyme. SIRT3 is involved in regulatingthe activity of the DNA repair enzyme OGG1, which leadsto increased damage to mtDNA and apoptosis of alveolarepithelial cells . A number of studies haveshown the SIRT3 association with various complex diseases.We studied the association between two SIRT3 gene functionalpolymorphisms (rs3782116 and rs536715) and \u0441hronicobstructive pulmonary disease. Functional analysis showedthat rs3782116 is located in the region of binding sites forhsa-miR-328; polymorphic loci rs3782116 and rs536715 arelocated in DNA regions that bind regulatory proteins. Bothpolymorphic loci were associated with COPD in our cohort;the disease development risk was associated with the G allelesof rs3782116 and rs536715. It should be noted that the carriersof the homozygous rare allele A of rs3782116 of the SIRT3gene had higher values of VC and FVC1. The contributionof SIRT3 variants to COPD has not been studied previously.At the same time, effects of the SIRT3 gene SNPs have beenfairly extensively investigated in age-associated diseases inwhich oxidative stress and cellular senescence play a key role.We obtained significant associations of SIRT6 (rs107251)with COPD; the frequent C allele is associated with COPDrisk, while the heterozygous genotype has a protective effecton the disease development. rs107251 is located in the DNAregion that binds to the SOX8 regulatory protein and it is inclose linkage with rs350846, localized in the 3-non-translationalregion of the SIRT6 gene \u2013 a binding site for severalmiRNAs . SIRT6 participates in the regulationof genome stability, NF-kB signaling, glucose homeostasis,exhibits ADP-ribosyltransferase and histone deacetylase activity,plays a role in DNA repair and maintenance of telomericchromatin integrity . In the studyby N. Takasaka et al. (2014), a decrease in SIRT6 levels wasshown in respiratory epithelial cells of COPD patients due tocigarette smoke exposure, leading to cellular senescence anddisruption of autophagy processes. Association of the SIRT6gene SNPs with COPD has not been studied previously, butthere is evidence of their association with cardiovasculardiseases, which are often comorbid pathologies in COPD.The PIK3R1 gene encodes regulatory subunit 1 of phosphoinositide3-kinase, a key element of the PI3K/AKT/mTORsignaling cascade . We investigated threePIK3R1 gene polymorphic loci , which showed a significant association withCOPD in our studied group. Carriers of rare alleles of thesepolymorphic loci had a high risk of COPD. In addition, weinvestigated the relationship between rs3730089 genotypesand VC and FVC1 values; thus, heterozygotes have lowervalues, and these results are in agreement with associationanalysis. Functional analysis showed that an intronic variantrs831125 is located in a binding site for regulatory proteins;rs3730089 is a missense variant with a \u201cbenign\u201d effect accordingto the PolyPhen-2 database (http://genetics.bwh.harvard.edu/pph2/), located in a binding site for regulatory proteinsand affecting splice sites. SNPs of the PIK3R1 gene have notbeen evaluated for COPD before. Previously, an associationhas been observed between rs3730089 and type 2 diabetes.The phosphatase PTEN regulates the activity of phosphoinositide-3 kinase (PI3K) . Smokingas a major risk factor for COPD provokes oxidative stress,which, in turn, affects PTEN expression .We investigated two PTEN gene functional polymorphicloci; rs70184 is located in PTEN gene 3\u2032 region and changesbinding sites for hsa-miR-1252 and hsa-miR-1304 miRNA;rs2735343, located in the intronic region, affects binding sitesfor several regulatory proteins.Significant associations with COPD in our sample werefound with PTEN gene loci; thus, homozygous carriers ofthe rare C allele of rs701848 and heterozygous carriers ofrs2735343 had a significant risk of COPD development.Published data have demonstrated an association betweenPTEN (rs701848) and COPD; the risk was significantly reducedfor homozygous T allele carriers, which is consistentwith the data obtained for our sample .PTEN participates in the regulation of various biologicalprocesses, including cell proliferation, apoptosis, inflammatoryreactions, transcription, and genomic stability . Decreased levels of PTEN lead to activation of PI3Ksignaling and increased cell senescence in COPD . It has been shown that decreased PTEN activity inCOPD increases the activity of matrix metalloprotease MMP9in bronchial epithelial cells, which consequently contributesto the progression of inflammation and extracellular matrixdegradation .The analysis of gene-gene interactions revealed significantsynergy between polymorphic loci of genes encodingphosphoinositide-3-kinase (PIK3R1) and mitochondrialdeacetylase (SIRT3), which were present in most significantcombinations associated with an increased risk of \u0441hronicobstructive pulmonary disease. The C allele in the PTEN(rs2735343) was part of four informative combinations associatedwith a high risk of \u0441hronic obstructive pulmonarydisease.The results of polygenic analysis indicate the interactionof genes encoding sirtuins SIRT3, SIRT2, SIRT6 and PI3KR1,PTEN, MTOR and confirm the functional relationship betweensirtuins and the PI3K/AKT/mTOR signaling pathway.The obtained results of single locus and polygenic analysis indicatethe contribution of SIRT3 , SIRT6(rs107251) and PIK3R1 polymorphisms to COPD and interaction of genes enco-ding the key components of the PI3K/AKT/mTOR signallingpathway and sirtuins, and confirm the involvement of cellularsenescence mechanisms with COPD developmentThe authors declare no conflict of interest.Barnes P.J., Baker J., Donnelly L.E. Cellular senescence asa mechanism and target in chronic lung diseases. Am. J.Respir. Crit. Care Med. 2019;200(5):556-564. 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+{"text": "Even though NoShow\u2019s 52,825 bp genome is most similar to phages in cluster AB, its  Mycobacterium abscessus  revealed a siphovirus morphology , prepared for sequencing using the NEB Ultra II library kit, and sequenced on Illumina MiSeq (v3 reagents), yielding 59,426 single-end 150-base reads. Raw reads were assembled and checked for completeness using Newbler (v2.9) and Consed (v29) with default settings (MK279840), Muddy , and FF47  (Geneious Prime (2022.0.2)) and has the highest G + C content . NoShow possessed 11 bp 3\u2032 single-stranded genome termini (5\u2032-CGGAGAGGCTT-3\u2032) (NoShow\u2019s DNA was extracted by Phenol-Chloroform (settings . Within http://phages.wustl.edu/starterator), ARAGORN (v1.2.38) and tRNAscan-SE (v1.3) (The genome was annotated using PECAAN (v2021) , PhameraE (v1.3) , 11, HHPE (v1.3) , and BlaE (v1.3) , and theE (v1.3) . DefaultConsistent with cluster AB phages, NoShow lacks any identifiable immunity repressor or integrase functions and is, therefore, considered a lytic phage \u201318. Unli\u201321\u2013"}
+{"text": "The name of the first author was incorrectly expressed in the Citation. The correct Citation is: Winck FV, Arvidsson S, Ria\u00f1o-Pach\u00f3n DM, Hempel S, Koseska A, et al. (2013) Genome-Wide Identification of Regulatory Elements and Reconstruction of Gene Regulatory Networks of the Green Alga Chlamydomonas reinhardtii under Carbon Deprivation. PLoS ONE 8(11): e79909. doi:10.1371/journal.pone.0079909."}
+{"text": "AbstractDoryctopambolusgen. n. is erected to contain Doryctopambolus pilcomayensis , comb. n., which was previously placed within Pambolus (Pambolinae), as well as three new species, Doryctopambolus clebschisp. n., Doryctopambolus dominicanussp. n. and Doryctopambolus sarochensissp. n. Members PageBreakof this new genus are mainly characterised by the presence of at least one pair of conspicuous propodeal apico-lateral projections, which are similar to those present in all members of Pambolinae and in species of three Australasian doryctine genera. We generated DNA barcoding sequences for the three newly described species. We discuss the morphological similarity between species of the Australasian Echinodoryctes Belokobylskij, Iqbal & Austin and Doryctopambolus. A key for the described species of Doryctopambolus is provided.The new Neotropical doryctine genus  Eucharitidae, Figitidae, Ichneumonidae and Braconidae, among others. Within the cyclostome group of subfamilies belonging to Braconidae, the presence of acute tubercles or spines on the apical lateral region of propodeum has been considered as the main diagnostic character employed to distinguish members of the cosmopolitan Pambolinae , all belonging to the cyclostome subfamily Doryctinae, also have these propodeal projections, indicating that this feature has actually evolved on separate occasions within Braconidae.The presence of propodeal or scutellar spines is a common feature found in species of a number of hymenopteran parasitoid families, including mbolinae . HoweverPambolus described to date , comb. n. and three additional Neotropical species described herein. We also molecularly characterise the three newly described species with the DNA barcoding locus , Colecci\u00f3n Nacional de Insectos, Instituto de Biolog\u00eda, Universidad Nacional Aut\u00f3noma de M\u00e9xico (CNIN-IB UNAM), Museo Argentino de Ciencias Naturales \u201cBernardino Rivadavia\u201d, Buenos Aires, Argentina (MACN), and Museo de La Plata, Argentina (MLP). Terminology for body structure, sculpture and wing venation features follows Digital colour photographs of all the species included in this work were taken with a Leica\u00ae Z16 APO-A stereoscopic microscope and a Leica\u00ae DFC295/DFC290 HD camera, and were edited using the Leica Application Suite\u00ae program. Digital scanning electron microscope (SEM) photographs of DNA sequences belonging to the barcoding locus [~650 bp of the cytochrome oxidase I (COI) mitochondrial DNA gene; Nunes & Zald\u00edvar-River\u00f3ngen. n.urn:lsid:zoobank.org:act:2C42D9FA-392C-4F6E-8A6E-CA00EAE711C0http://species-id.net/wiki/DoryctopambolusPambolus pilcomayensis van Achterberg & Braet, 2004Doryctopambolus can be distinguished from members of most doryctine genera except Concurtisella bidens, Echinodoryctes, Fijispathius and Ryukuspathius by having the propodeum with at least one pair of conspicuous apico-lateral projections. SpePageBreakPageBreakcies of Doryctopambolus and Concurtisella bidens are the only Neotropical doryctine taxa reported to have these projections, though they mainly differ by their first subdiscal cell  and ovipositor length . Doryctopambolus differs from the Australasian Fijispathius and Ryukyuspathius mainly by the fore wing first subdiscal cell open at apex (closed in the later two genera) and the first metasomal segment not petiolate . Doryctopambolus is morphologically similar to the Australian Echinodoryctes , propodeum evenly curved and strongly rugose-areolate (globose and mostly smooth in Echinodoryctes), hind coxa without basoventral tubercle and all femora without dorsal protuberances (both present in Echinodoryctes).Species ofdoryctes . HoweverHead: head globose; antennal sockets distinctly separated from each other by at least 0.5 times its diameter; frons almost flat, without median carina or furrows; ocelli arranged in equilateral triangle; eye with distinct and sparse setae; gena and temple smooth; malar suture absent; first flagellomere slightly shorter than scape and pedicel combined, slightly longer than second flagellomere; antenna with 16\u201328 antennomeres; occipital carina meeting hypostomal carina before mandible. Mesosoma: Length of mesosoma about two times its maximum width; neck of pronotum fairly long; pronotal crest conspicuous; mesoscutum declivous anteriorly; mesoscutal lobes smooth and polished medially; notauli complete and strongly impressed; scutellar sulcus deep, with its height 0.8\u20130.9 times height of scutellar disc; precoxal sulcus complete and scrobiculate, as long as mesopleuron; prepectal carina coarse and complete; propodeum evenly curved and strongly rugose-areolate, with at least one pair of conspicuous apico-lateral projections; propodeal bridge absent. Legs: fore tibia with a row of 7\u20138 stout spines; middle tibia without spines; femora without dorsal protuberances; hind coxa without basal tubercle. Wings: partially reduced to well-developed wings; fore wing veins r-m and 2RS present; m-cu arising interstitial or slightly antefurcal with vein 2RS, cu-a distinctly postfurcal with vein 1M; first subdiscal cell open at apex; hind wing vein M+CU equal length of vein 1M; cu-a present, m-cu absent; stigma present on male hind wing. Metasoma: length of first metasomal tergum 1.3\u20131.6 times its apical width, apical width about 2.0\u20132.3 times basal width; basal sternal plate (acrosternite) about 0.33\u20130.5 times length of tergum; suture between second and third metasomal tergites absent; second metasomal tergite at least sculptured basally; third metasomal tergite usually smooth, sometimes sculptured basally; remaining metasomal tergites entirely smooth and polished; ovipositor about same length of metasoma.Small size, 2.2\u20133.6 mm; black to light brown species. Neotropical. Known from central Argentina to northern Venezuela, and from Dominican Republic in the Caribbean.Doryctopambolus and Doryctopambolus pilcomayensis comb. nov. are described and redescribed in this study, respectively. Four additional speciePageBreaks belonging to this genus, two from Cerro Saroche, Lara, Venezuela, and two from Argentina, were also identified. The two Argentinian species could not be described due to their bad state of preservation.Of the two species from Venezuela, one was represented by a single male and the other one by an incomplete female, and their allospecificity was corroborated with DNA barcoding sequences . The Parque Nacional Cerro Saroche is a natural reserve of about 32,294 h mainly composed of xeric vegetation with deciduous and semideciduous shrubs comb. n.Pambolus pilcomayensis van Achterberg & Braet 2004: pp. 341\u2013344.Doryctopambolus sarochensis sp. n. by having one pair of propodeal apico-lateral projections (two pairs in Doryctopambolus sarochensis sp. n.) and third metasomal tergite entirely smooth . Doryctopambolus pilcomayensis differs PageBreakfrom Doryctopambolus dominicanussp. n. and Doryctopambolus clebschi sp. n. by having the vertex striate (smooth in Doryctopambolus dominicanus sp. n. and Doryctopambolus clebschi sp. n.), first metasomal tergite not petiolate (at least 0.5 of tergum) and second metasomal tergite entirely sculptured (mostly smooth).This species distinguishes from Colour: head light to dark brown, antenna light brown, malar space light brown to yellowish, mandible and palpi light brown; propleuron and pronotum light brown, remainder of mesonotum brown to black; wings slightly infuscate, narrow banded with brown veins; metasoma brown to black, first two metasomal tergites darker; legs brown to dark brown, fore and middle coxae, trochanter, trochantellus and tarsi light brown to pale yellow; ovipositor and sheaths light brown with dark apex. Head: antero-dorsal surface of eyes bordered by groove; eyes setose; face costate-rugose laterally, central area slightly swollen and smooth; clypeus costate-rugose; malar space 0.8\u20131.0 times eye height; frons and vertex striate; temple in dorsal view 0.7 times eye width; antenna with 26 antennomeres. Mesosoma: length of mesosoma 2.0 times its maximum height; pronotum rugose; pronotal groove wide and scrobiculate laterally, dorsally smooth; propleuron costate-rugose; notauli deep and scrobiculate, meeting scutellum with parallel carinae; scutellum smooth and setose; scutellar sulcus with three parallel carinae; mesopleuron porcate dorsally, smooth near precoxal sulcus and ventrally; ventral portion of mesopleuron separated by shallow crenulate median groove with some rugosities near middle coxae; propodeum with one pair of conspicuous apico-lateral projections, longer than first flagellomere; median carina present only basally. Legs: hind coxa costate to costate-rugose dorsally. Wings: fore wing length 3.7\u20134.8 times its maximum width, r:3RSa:3RSb = 2:8-12:18-32; 2RS:3RSa:r-m = 6:8-12:2-3; m-cu arising interstitial or slightly antefurcal to vein 2RS; 1cu-a interstitial with 1M. Metasoma: length of first metasomal tergite 1.3 times its apical width, apical width about 2.0 times basal width costate-rugose, with two or three longitudinal carinae reaching apex of tergum; second metasomal tergite strongly costate; remaining metasomal tergites smooth and polished; basal sternal plate (acrosternite) about 0.33 times length of tergum; ovipositor nearly as long as metasoma.Females. Body length 3.0\u20133.6 mm, fore wing 2.0 mm, ovipositor 1.8 mm. Males. Essentially as females; body length 2.1\u20133.0 mm; body brown, with antennae, trochanter, trochantellus, tarsi, fore and middle coxae light brown to yellowish; face, frons and vertex slightly sculptured; stigma present in hind wing.Brazil: Goi\u00e1s State (GO), Itumbiara, Malaise trap, C.H. Marchiori col.: one female 24-I-1998, three males 28-I-1998, one female and one male 31-I-1998, two males 7-II-1998, three females and one male 28-II-1998, one female and one male 7-III-1998, one female 1-IV-1998, one female and one male 15-IV-1998, one female 08-XI-1998 (DCBU); Mato Grosso do Sul State (MS), Bodoquena, Parque Nacional da Serra da Bodoquena, Faz. Pitangueiras, Bandeja Am. 21 (yellow pan trap), Crepaldi RA et al. col: one female V-2009 (DCBU). Argentina: Misiones Province, Estaci\u00f3n Experimental Loreto, Dr. A. Ogloblin: one female 15-VI-1930 (MLP); Buenos Aires Province, Reserva Ecol\u00f3gica de Vicente L\u00f3pez, 34.492\u00b0S, 58.480\u00b0W: one male 16-I-2011 (MACN); one male, no data (MLP).Twenty-two specimens . PageBreakSouthern Brazil to Argentina.Doryctopambolus pilcomayensis near Buenos Aires can be explained by the courses of the Parana and Uruguay basins, which carry downstream many plant and animal species from tropical areas to higher latitudes.The presence of Zald\u00edvar-River\u00f3n & Mart\u00ednezsp. n.urn:lsid:zoobank.org:act:0992A72F-1163-48FB-A1BF-D5E59F7D117Fhttp://species-id.net/wiki/Doryctopambolus_clebschiDoryctopambolus clebschi is morphologically similar to Doryctopambolus dominicanus sp. n. However, Doryctopambolus clebschi differs from the latter species by having the face smooth medially and slightly rugose laterally (entirely rugose in Doryctopambolus dominicanus sp. n.), mesopleuron mostly smooth (mostly smooth-rugose), first metasomal tergite mostly smooth apically , and second metasomal tergite mostly smooth, slightly costate baso-laterally . These two species distinguish from the remaining two described species of Doryctopambolus, Doryctopambolus pilcomayensis comb. n. and Doryctopambolus sarochensis sp. n., by having the vertex smooth , first metasomal tergite distinctly petiolate (not petiolate), and second metasomal tergite partially smooth .. Body length 2.8 mm; fore wing 2.3 mm; ovipositor 1.7 mm. Colour: head brown, antennae brown, turning honey yellow to apex, palpi yellow; mesosoma and first metasomal tergites brown, second metasomal tergite honey yellow, remaining metasomal tergites light brown; wings hyaline; veins and stigma light brown; legs light brown to honey yellow; ovipositor and sheaths light brown to brown, apex strongly sclerotised and dark. Head: antenna with 16-17 antennomeres; eyes small, ovoid and setose; face smooth and glabrous medially, slightly rugose and pilose laterally, median area not swollen; clypeus slightly rugose; malar space 0.4 times eye height; frons and vertex smooth; temple in dorsal view 2.7 times eye width. Mesosoma: 1.8 times longer its maximum height; pronotum rugose laterally; pronotal groove wide and scrobiculate; propleuron slightly rugose; mesoscutal lobes entirely smooth and polished, sparsely pilose; notauli wide, deep and scrobiculate, meeting before scutellum in a longitudinally costate area; scutellum smooth, sparsely setose; scutellar sulcus with three parallel carinae; height of scutellar sulcus 0.8 times height of scutellar disc; subalar sulcus wide and scrobiculate, joining mesopleural sulcus, remaining area of mesopleuron mostly smooth and polished, slightly rugose near mesopleural sulcus; venter of mesopleuron smooth; propodeum with one pair of long and sharp apico-lateral projections, shorter than first flagellomere. Legs: hind coxa mostly smooth, slightly rugose ventrally. Wings: fore wing length 3.6 times its maximum width, r:3RS:3RSb = 2:3:10; 2RS:3RSa:r-m = 4:3:2; m-cu arising antefurcal with vein 2RS; 1cu-a interstitial with 1M; hind wing vein M + CU about equal length of vein 1M. Metasoma: length of first metasomal tergum 1.5 times its apical width, apical width about 2.3 times basal width, first metasomal tergite costate basally and medially, smooth apically; second metasomal tergite mostly smooth and polished, slightly costate baso-laterally; remaining metasomal tergites smooth and polished; basal sternal plate (acrosternite) about 0.5 times length of tergum; ovipositor about 1.1 times longer than metasoma.FemaleVariation. Females. Body length 2.6\u20132.8 mm.Males. Unknown.PageBreakHolotype. Female (CNIN IB-UNAM). Dominican Republic: A. Bermudez NP, La Ci\u00e9nega, Telostablones, 19.066\u00b0N, 70.863\u00b0W, 15-16-IX-2009, sweep, H. Clebsch col. . Paratype. One female (CNIN IB-UNAM), same data as holotype.Dominican Republic.This species is named in honour to our good friend and colleague Hans Clebsch, who collected the specimens assigned to this species.Zald\u00edvar-River\u00f3n & Mart\u00ednezsp. n.urn:lsid:zoobank.org:act:99C34338-DB4F-4AE2-A662-4DDA9957BA43http://species-id.net/wiki/Doryctopambolus_dominicanusD. Clebschi.See . Body length 2.7 mm; fore wing 1.8 mm; ovipositor 1.3 mm. Colour: head light brown, antennae light brown, turning honey yellow to apex, palpi yellow; mesosoma and first and basal two thirds of second metasomal tergites dark brown; wings hyaline; veins and stigma light brown; legs light brown to honey yellow; ovipositor and sheaths honey yellow, apex strongly sclerotised and dark. Head: PageBreak16 flagellomeres  about 0.5 times length of tergum; ovipositor 0.9 times as long as metasoma.FemaleHolotype.Female (CNIN IB-UNAM). Dominican Republic: Punta Cana , Malaise trap, Masner col. .Dominican Republic.The name of this species refers to the country where the holotype was collected, Dominican Republic.Nunes & Zald\u00edvar-River\u00f3nsp. n.urn:lsid:zoobank.org:act:9C8C7812-2519-4849-A2CB-D3EED92F7FFBhttp://species-id.net/wiki/Doryctopambolus_sarochensisDoryctopambolus sarochensis sp. n. distinguishes from the remaining species of the genus by having the vertex striate-rugose (striate in Doryctopambolus pilcomayensis comb. n. and smooth in the remaining two species), two pairs of propodeal apico-lateral projections, the basal ones small and blunt and the second pair long and distinctly truncate, and third metasomal tergite costate baso-laterally (entirely smooth in the other species).. Body length 3.0 mm; fore wing 2.0 mm; ovipositor 1.7 mm. Colour: head brown, malar space honey yellow, antenna honey yellow, turning brown at apex, mandible honey yellow, palpi yellow; mesosoma and metasoma dark brown to black; wings slightly infuscate; veins and stigma brown; fore leg honey yellow; middle leg honey yellow, with femur light brown; hind coxa and femur brown,PageBreak trochanter, trochantellus, tibia and tarsi honey yellow; ovipositor and sheaths honey yellow, apex strongly sclerotised and dark. Head: 23 antennomeres; eyes setose; face striate medially, striate-rugose laterally, with median area slightly swollen; clypeus costate-rugose; malar space 0.6 times eye height; frons and vertex striate-rugose; temple in dorsal view 0.8 times eye width. Mesosoma: two times longer its maximum height; pronotum rugose laterally; pronotal groove wide and scrobiculate; propleuron costate-rugose; mesoscutal lobes mostly smooth and shinning, rugose along notauli and at lateral edges; notauli narrow and scrobiculate, meeting before scutellum in a longitudinally costate-rugose area; scutellum smooth, with some setae; scutellar sulcus with three parallel carinae; height of scutellar sulcus 0.8 times height of scutellar disc; subalar sulcus wide, deep and scrobiculate, joining mesopleural sulcus in an inverted \u201cV\u201d shape; mesopleuron porcate dorsally, porcate-rugose laterally, smooth ventrally; venter of mesopleuron smooth; propodeum with two pairs of apico-lateral projections, the most basal one small and blunt, the apical one long and distinctly truncate, longer than first flagellomere. Legs: hind coxa strongly costate-rugose dorsally, poorly sculptured ventrally. Wings: fore wing length 3.8 times its maximum width, r:3RS:3RSb = 3:6:22; 2RS:3RSa:r-m = 8:6:5; m-cu arising antefurcal with vein 2RS; 1cu-a interstitial with 1M. Metasoma: length of first metasomal tergite 1.5 times its apical width, apical width about 2.0 times basal width, costate rugose with two dorsal carinae converging at apex; second metasomal tergite strongly costate; third metasomal tergite mostly smooth, costate baso-laterally, remaining metasomal tergites smooth and polished; basal sternal plate (acrosternite) about 0.33 times length of tergum; ovipositor as long as metasoma.FemaleVariation.Females. Body length 2.7\u20133.0 mm; antenna with 22\u201323 antennomeres. Males. Unknown.Holotype.Female (CNIN IB-UNAM). Venezuela: Lara, Parque Nacional Cerro Saroche, Ca\u00f1aote #3, 10\u00b011'083\"N, -69\u00b026'013\"W; 1000 msnm; R. Brice\u00f1o, R. Paz, W. Rom\u00e1n, D. Torres colls.; . Paratype. Onefemale (CNIN IB-UNAM), same data as holotype .NortheastVenezuela.The name of this species refers to the place where the type material was collected..Echinodoryctes tetraspinosus Belokobylskij, lqbal & Austin. One specimen. Paratype. Female. Australia: Prevelly Park, W of Margaret\u00a0River, 1 nov. 1984 W.A., I. & N. Lawrence, in marri nuts on ground. Doryctopambolus cf. pilcomayensis: Doryctopambolus sp. 1: one female (MLP): Argentina, Misiones, Estaci\u00f3n Experimental Loreto, Dr. A. Ogloblin, 25-VI-1931; one male (MLP): Argentina, Misiones, Estaci\u00f3n Experimental Loreto, Dr. A. Ogloblin, 8-VI-1933; Doryctopambolus sp. 2: one male (MACN): Argentina, Ciudad Aut\u00f3noma de Buenos Aires, Reserva Ecol\u00f3gica Costanera Sur, trampa de ca\u00edda, Mamani, Turienzo, Zapata, 19-I-2009; Doryctopambolus sp. 3 (CNIN IB-UNAM): Venezuela, Lara, Parque Nacional Cerro Saroche, Ca\u00f1aote, 10\u00b011'083\"N, 69\u00b026'013\"W; 1000 m; R. Brice\u00f1o, R. Paz, PageBreakW. Rom\u00e1n, D. Torres colls. ; Doryctopambolus sp. 4 (CNIN IB-UNAM): Venezuela, Carabobo, Palmichal, 10.2859N, 68.2399W, 931 m, 30-31-iii-07, YPT/64 plates, shade coffee/Orange grove plantations, H. Clebsch col. .Doryctompambolus, Doryctopambolus clebschi (two specimens), Doryctopambolus dominicanus (one specimen), Doryctopambolus sarochensis (two specimens) and the two undescribed species from Venezuela (one specimen each), was congruent with the interspecific variation observed in other braconid genera, ranging from 4.2 to 14%. The lowest interspecific genetic distance occurred between specimens of the two Dominican species, Doryctopambolus clebschi and Doryctopambolus dominicanus.Interspecific variation of the barcoding locus among the examined species of"}
+{"text": "Postoperative Atrial Fibrillation (POAF) is the most common arrhythmia in cardiac surgery. Its incidence ranges between 20-40%. The objective of this study was to determine the predictive factors of POAF that occurred in patients undergoing coronary artery bypass grafting (CABG), and the impact of this event on morbidity and mortality.This bi-directional and longitudinal cohort study was made with 3,010 patients undergoing CABG between June/2009 and July/2010, aged \u2265 18 years. We excluded 382 patients for chronic or paroxysmal atrial fibrillation, atrial flutter, or association of the surgery evaluated with other procedures.The incidence of POAF was 12.4% ; rate female: male ratio of 1:3; CRI (p=0.001), creatinine >/= 2.2mg/dl (p=0.002); COPD (p=0.001), ICC (p=0.004), EuroSCORE (p<0.001), blood transfusion (p<0.001), Peripheral Arterial Insufficiency (PAI) ; emergency surgery/emergency (p= 0.006), postoperative stroke (p<0.001) and RI postoperatively (p <0.001). Patients with POAF had longer postoperative hospital stay (p<0.001), higher mortality at one year (p <0.001) and a higher rate of readmission within 30 days (p = 0.004).The age, male gender, CRI, COPD, peripheral arterial disease, creatinine >/= 2,2mg/dl, CHF, EuroSCORE, blood transfusion, emergency surgery/emergency (p= 0.006), postoperative stroke (p<0.001) and RI postoperatively (p<0.001) variables were independently predictive of POAF event. Due to the strong correlation with the increased length of hospitalization, mortality and re-hospitalization in these patients, we must implement strategies for POAF prophylaxis and prevention."}
+{"text": "Computerized testing of neurocognitive function yields an accurate and reliable assessment . There iThe purpose of this study was to investigate neurocognitive function and recovery patterns in college students who incur migraine headaches compared to college students who do not.Volunteers (ages 18-29) completed computerized neurocognitive baseline (B) testing. Forty-four migraineurs incurring a migraine (M) were matched to 44 non-migraine (NM) controls for sex, age and education level. Verbal and visual memory, processing speed and reaction time were measured at 24 hours, 48 hours and 7 days post migraine.Repeated measures ANOVAs revealed declines in neurocognitive function of migraineurs in verbal memory , visual memory [md (24hr-B)M=-4.70 +15.61, NM=3.05+10.94; p=.041), and reaction time [md(24hr-B)M=.02\u00b1.09, NM=-.01\u00b1.04."}
+{"text": "Fragility fractures are a common and increasing cause of morbidity and mortality in the general population; risk factors for fractures are commoner in HIV . This study aims to determine the prevalence and associations of low bone mineral density (BMD) and high fracture risk (FR) in an HIV cohort, suggest screening and management guidelines.A cross-sectional study of 223 randomly selected HIV-infected outpatients was undertaken. Recruitment was stratified by gender for age groups 30-39, 40-49 yrs and \u226550 yrs. Osteopenia and osteoporosis were diagnosed according to the WHO criteria. Patients completed a detailed questionnaire including combination HIV drugs (cART) history, & a dual-energy X-ray absorptiometry (DEXA) of Lumbar spine & Left Hip. Investigations included serum Ca, phosphate, 25OH vit D, alkaline phosphatase (Alk P) PTH, albumin, sex hormone binding globulin (SHBG), testosterone, CD4, HIV RNA. BMD risk factors were recorded including previous fractures, smoking, malabsorption, alcohol consumption, BMI, chronic diseases, physical activity index and past medication. FRAX score (10yr probability major fractures), and remaining lifetime fracture probability (RLFP) were calculated. Controls were from the Twin Research Unit at Kings College London. Data were analysed using multivariate logistic regression.Demographics: 133(60%) were male, 106(48%) were Caucasian, 71(33%) had AIDS at diagnosis. 190(85%) were taking cART, of whom 50(26%) were on their first line therapy. Osteoporosis/osteopenia were present in 13%/39% of males, 11%/29% females, and was approximately 2.4/3.0 fold greater than age-matched controls. The overall mean 10 yr fracture risk was 3.16%. RLFP exceeded 1.0 in 76% HIV patients, and <20% controls.Factors associated with low BMD after multivariate analysis: adjusted OR(95 % CI)/p-value BMI 0.90, p<0.001, Alk P 1.01, p 0.05, testosterone 1.04, p 0.01, Initiated cART 3.61, p 0.01. The lack of association with age is notable, adjusted OR 1.08, p 0.35Reduced bone mineral density and subsequent fracture risk is much commoner in patients with HIV compared to controls, especially, and those taking cART, and occurred across the age ranges. Hence screening for BMD and risk factors for fragility fractures is indicated in patients with HIV at a younger age than in the general population, especially if they are on cART."}
+{"text": "Staphylococcus aureus Hospitalizations,\" by Matthew J Kuehnert et al., an error occurred. In Table 3, columns 3 and 5, the rates shown for hospitalization with S. aureus and MRSA-related discharge diagnoses were per 10,000 discharges, rather than per 1,000 discharges, as indicated.In \u201cMethicillin-resistant\u2013http://www.cdc.gov/eid/11/6/04-0831-t3.htmThe corrected table appears in the updated article at We regret any confusion this error may have caused."}
+{"text": "The article title is incorrect. \u201cThe correct title is: MicroRNA-130b Suppresses Migration and Invasion of Colorectal Cancer Cells through Downregulation of Integrin \u03b21.\u201dThe correct Citation is: Zhao Y, Miao G, Li Y, Isaji T, Gu J, et al. (2014) MicroRNA-130b Suppresses Migration and Invasion of Colorectal Cancer Cells through Downregulation of Integrin \u03b21. PLoS ONE 9(2): e87938. doi:10.1371/journal.pone.0087938"}
+{"text": "The Sanger Mouse Genetics Project was incorrectly abbreviated in the citation. The correct citation is: Migdalska AM, van der Weyden L, Ismail O, White JK, Sanger Mouse Genetics Project, et al. (2012) Modeling Partial Monosomy for Human Chromosome 21q11.2-q21.1 Reveals Haploinsufficient Genes Influencing Behavior and Fat Deposition. PLoS ONE 7(1): e29681."}
+{"text": "Granulicella mallensis MP5ACTX8T is a novel species of the genus Granulicella in subdivision 1of Acidobacteria. G. mallensis is of ecological interest being a member of the dominant soil bacterial community active at low temperatures and nutrient limiting conditions in Arctic alpine tundra. G. mallensis is a cold-adapted acidophile and a versatile heterotroph that hydrolyzes a suite of sugars and complex polysaccharides. Genome analysis revealed metabolic versatility with genes involved in metabolism and transport of carbohydrates. These include gene modules encoding the carbohydrate-active enzyme (CAZyme) family involved in breakdown, utilization and biosynthesis of diverse structural and storage polysaccharides including plant based carbon polymers. The genome of Granulicella mallensis MP5ACTX8T consists of a single replicon of 6,237,577 base pairs (bp) with 4,907 protein-coding genes and 53 RNA genes. T (= ATCC BAA-1857T = DSM 23137T), is the type strain of the species Granulicella mallensis [Granulicella, in subdivision 1 of Acidobacteria, was first described by Pankratov et al. in 2010 [Granulicella mallensis  was described along with other species of the genus Granulicella isolated from tundra soil [Granulicella species.Strain MP5ACTX8Acidobacteria is one of the most ubiquitous bacterial phyla found in diverse habitats and is abundant in most soil environments [Acidobacteria are phylogenetically and physiologically diverse [Acidobacterium [Terriglobus [Edaphobacter [Granulicella [Acidipila [Telmatobacter [Acidicapsa [Bryocella [Bryobacter [Blastocatella [Thermotomaculum [Holophaga [Geothrix [Acanthopleuribacter [Candidatus Koribacter versatilis\u2019 [Candidatus Solibacter usitatus\u2019 [Candidatus Chloracidobacterium thermophilum\u2019 [Acidobacteria are relatively difficult to cultivate with slow growth rates and typically require up to several weeks to develop visible colonies on solid media. Nevertheless, the phylogenetic diversity, ubiquity and abundance of this group suggest that they play important ecological roles in soils. The abundance of Acidobacteria has been found to correlate with soil pH [Acidobacteria being most abundant in slightly acidic soils. Our previous studies have shown that Acidobacteria dominate in the acidic tundra heaths of northern Finland [Acidobacteria [T was classified as a novel species of the genus Granulicella [Granulicella mallensis MP5ACTX8T , G. sapmiensis S6CTX5AT (96.2%) and G. arctica MP5ACTX2T (96.1%) and 94.6 \u2013 97.4% to G. rosea TPO1014T (94.6%), G. aggregans TPB6028T (96.0%), G. pectinivorans TPB6011T (96.1%), G. paludicola OB1010T (96.5%) and G. paludicola LCBR1 (97.4%). Phylogenetic analysis based on the 16S rRNA gene of taxonomically classified strains of family Acidobacteriaceae placed G. paludicola type strain OB1010 T as the closest taxonomically classified relative of G. mallensis MP5ACTX8T  at pH 3.5\u20136.5 (optimum pH 5) and at +4 to +28 \u00b0C (optimum 24\u201327 \u00b0C) [T forms opaque white mucoid colonies with a diameter of approximately 1 mm. Cells are Gram-negative, non-motile, aerobic rods, approximately 0.5\u20130.7 mm wide and 0.6\u20131.3 mm long. Growth observed with up to 1.5% NaCl (w/v) (T is shown in 4\u201327 \u00b0C) . On R2 aCl (w/v) . The celG. mallensis utilizes D-glucose, maltose, cellobiose, D-fructose, D-galactose, lactose, lactulose, D-mannose, D-ribose, raffinose, sucrose, trehalose, D-xylose, N-acetyl-D-glucosamine, glucuronate, glutamate, melezitose and salicin, but does not utilize D-arabinose, acetate, formate, pyruvate, malate, mannitol, D- or L-alanine, D-glycine, L-leucine, L-ornithine, gluconic acid, aspartate, dulcitol, butyrate, caproate, valerate, lactate, oxalate, propionate, fumarate, adonitol, methanol, ethanol, succinate, D-sorbitol or myoinositol, when grown using VL55 mineral medium with 100 mg yeast extract l-1. G. mallensis hydrolyzes aesculin, starch, pectin, laminarin and lichenan, but not gelatin, cellulose, xylan, sodium alginate, pullulan, chitosan or chitin on R2 medium. Strains show positive reaction for acid and alkaline phosphatases, leucine arylamidase, a-chymotrypsin, naphthol-AS-BI-phosphohydrolase, \u03b1- and \u03b2-galactosidases, \u03b1- and \u03b2-glucosidases, N-acetyl- \u03b2-glucosaminidase, \u03b2-glucuronidase, trypsin and valine arylamidase, but negative for \u03b1-fucosidase, \u03b1-mannosidase, esterase (C4 and C8), lipase (C14) and cystine arylamidase. Strain MP5ACTX8T reduces nitrate to nitrite. Strain MP5ACTX8T is resistant to the antibiotics erythromycin, chloramphenicol, neomycin, rifampicin, streptomycin, gentamicin, polymyxin B and penicillin, but susceptible to ampicillin, kanamycin, tetracycline, lincomycin, novobiocin and bacitracin [G. mallensis are iso-C15:0 (45.3%), C16:1\u03c97c (28.7%), iso-C13:0 (8.3%) and C16:0 (8.9%). The cellular fatty acid compositions of strain MP5ACTX8T were relatively similar to that of other Granulicella strains with fatty acids iso-C15:0 and C16:1\u03c97c being most abundant in all strains. Strain MP5ACTX8T contains MK-8 as the major quinone.The major cellular fatty acids in G. mallensis strain MP5ACTX8T was selected for sequencing in 2009 by the DOE Joint Genome Institute (JGI) community sequencing program. The Quality Draft (QD) assembly and annotation were completed on December 26, 2010. The complete genome was made available on Dec. 1, 2011. The genome project is deposited in the Genomes On-Line Database (GOLD) [T is deposited in GenBank (CP003130). e (GOLD)  and the G. mallensis MP5ACTX8T was cultivated on R2 medium as previously described [escribed . Genomicescribed .G. mallensis MP5ACTX8T (JGI ID 4088692) was generated at the DOE Joint genome Institute (JGI) using a combination of Illumina [The finished genome of Illumina  and 454 Illumina . The 454Illumina . The 454Illumina . The sofIllumina  was usedIllumina  was usedIllumina  or sequeGenes were identified using Prodigal  as part The genome consists of one circular chromosome of 6,211,694 bp in size with a GC content of 57.8 mol% and consists of 53 RNA genes . Of the Granulicella mallensis type strain MP5ACTX8T has the largest genome size of 6.2 Mbp. among the three tundra soil strains of subdivision 1 Acidobacteria [Granulicella mallensis identified a high abundance of genes assigned to COG functional categories for transport and metabolism of carbohydrates (9.1%) and amino acids (6.7%) and involved in cell envelope biogenesis (8.3%) and transcription (8.6%). Further genome analysis revealed an abundance of gene modules encoding for functional activities within the carbohydrate-active enzymes (CAZy) family [G. mallensis hydrolyzed complex carbon polymers, including CMC, pectin, lichenin, laminarin and starch, and utilized sugars such as cellobiose, D-mannose, D-xylose, D-trehalose. This parallels genome predictions for CDSs encoding for enzymes such as cellulases, pectinases, alginate lyases, trehalase and amylases. In addition, the G. mallensis genome contained a cluster of genes in the neighborhood of the cellulose synthase gene (bcsAB) which included cellulase (bscZ) (endoglucanase Y) of family GH8, cellulose synthase operon protein (bcsC) and a cellulose synthase operon protein (yhjQ) involved in cellulose biosynthesis. Detailed comparative genome analysis of G. mallensis MP5ACTX8T with other Acidobacteria strains for which finished genomes were available is reported in Rawat et al. [G. mallensis is involved in hydrolysis, the utilization of stored carbohydrates, and in the biosynthesis of exopolysaccharides from organic matter and plant based polymers in the soil. Therefore, we infer that strain G. mallensis may be central to carbon cycling processes in arctic and boreal soil ecosystems.bacteria . Genome ) family  involvedt et al. . The dat"}
+{"text": "Hepatic dysfunction has been associated with outcome of ICU patients. However, most scoring systems including APACHE II only marginally reflect acute liver dysfunction on admission. Indocyanine green (ICG) is eliminated by hepatobiliary excretion. Therefore, the ICG plasma disappearance rate (ICG-PDR) is used as a dynamic liver function test. ICG-PDR has been associated with mortality in several studies.A prospective study to compare prediction of 28-day mortality by ICG-PDR, other markers of liver function and scoring systems (primary endpoint). In the subgroup of patients with transpulmonary thermodilution (TPTD) monitoring , predictive capabilities of ICG-PDR were compared with cardiac index (CI), extravascular lung water index (EVLWI), global end-diastolic volume index (GEDVI) and pulmonary vascular permeability index (PVPI). ICG-PDR  and all other predictors were determined within 48 hours after admission. Statistics: IBM SPSS 20.P <0.001) and SOFA . Among markers of hepatic function on admission, ICG-PDR provided the largest AUC , which was larger than for GOT , bilirubin  and INR . Among TPTD parameters, only PVPI significantly predicted 28-day mortality , whereas CI, GEDVI and EVLWI were not predictive. Prediction of 28-day mortality by SOFA could not be improved by models including any hepatic parameter. By contrast, ICG-PDR was independently (P = 0.036) associated with mortality when included in a model (R = 0.58) with APACHE II. This model based on APACHE II and ICG-PDR provided the largest of all ROC-AUCs . ICG-PDR itself was independently associated with age (P = 0.025), but not with any other biometric parameter .A total of 154 patients , age 59 \u00b1 13 years, APACHE II score 16.0 \u00b1 5.7, SOFA score 7.6 \u00b1 4.2. Etiology: sepsis 14.4%, cirrhosis 28.8%, GI bleeding 8.9%, ARDS 17.8%, cardiogenic shock 4.1%, acute renal failure 3.4%, various 22.6%. The 28-day mortality was significantly predicted by APACHE II (ROC-AUC: 0.762; ICG-PDR on admission is an independent predictor of 28-day mortality. Predictive capabilities particularly of APACHE II can be improved by combination with ICG-PDR. Among TPTD-derived parameters, only PVPI provides significant prediction of mortality."}
+{"text": "The authors would like to add Matthias Gunther as the 6th author. The updated byline is: David A. Feinberg1,2,3*, Steen Moeller4, Stephen M. Smith5, Edward Auerbach4, Sudhir Ramanna1, Matthias Gunther1, Matt F. Glasser6, Karla L. Miller5, Kamil Ugurbil4, Essa Yacoub4The updated author contributions are: Conceived and designed the experiments: DAF SMS KU EY. Performed the experiments: DAF SM EA SR MG EY. Analyzed the data: DAF SM SMS EY KLM MFG. Contributed reagents/materials/analysis tools: DAF SM EA SR SMS MG MFG. Wrote the paper: DAF SM SMS KU EY KLM."}
+{"text": "Plasmopara viticola, is one of the most severe diseases of grapevine and is commonly controlled by fungicide treatments. The beneficial microorganism Trichoderma harzianum T39 (T39) can induce resistance to downy mildew, although the molecular events associated with this process have not yet been elucidated in grapevine. A next generation RNA sequencing (RNA-Seq) approach was used to study global transcriptional changes associated with resistance induced by T39 in Vitis vinifera Pinot Noir leaves. The long-term aim was to develop strategies to optimize the use of this agent for downy mildew control.Downy mildew, caused by P. viticola inoculation. RNA-Seq analysis resulted in the identification of 7,024 differentially expressed genes, highlighting the complex transcriptional reprogramming of grapevine leaves during resistance induction and in response to pathogen inoculation. Our data show that T39 has a dual effect: it directly modulates genes related to the microbial recognition machinery, and it enhances the expression of defence-related processes after pathogen inoculation. Whereas several genes were commonly affected by P. viticola in control and T39-treated plants, opposing modulation of genes related to responses to stress and protein metabolism was found. T39-induced resistance partially inhibited some disease-related processes and specifically activated defence responses after P. viticola inoculation, causing a significant reduction of downy mildew symptoms.More than 14.8 million paired-end reads were obtained for each biological replicate of T39-treated and control leaf samples collected before and 24\u2009h after The global transcriptional analysis revealed that defence processes known to be implicated in the reaction of resistant genotypes to downy mildew were partially activated by T39-induced resistance in susceptible grapevines. Genes identified in this work are an important source of markers for selecting novel resistance inducers and for the analysis of environmental conditions that might affect induced resistance mechanisms. Plasmopara viticola (Berk. and Curt.) Berl. and de Toni is a biotrophic oomycete that causes downy mildew in grapevine Release 3 [78] in control (C), Trichoderma harzianum T39-treated (T39), Plasmopara viticola-inoculated control (C+P.v.), and P. viticola-inoculated T39-treated (T39+P.v.) plants. Expression values in each biological replicate (numbered from 1 to 3) and each sequencing replicate (named A and B) are also listed. Click here for fileCorrelations between sequencing replicates of RNA-Seq analysis. Comparison of the expression levels of all grapevine genes, expressed as fragments per kilobase of transcript per million fragments mapped (FPKM), in the two sequencing replicates (named A and B) of (A) control (C) biological replicate no. 1; (B) C biological replicate no. 2 (B); (C) C biological replicate no. 3; (D) Trichoderma harzianum T39-treated (T39) biological replicate no. 1; (E) T39-treated biological replicate no. 2; (F) T39-treated biological replicate no. 3; (G) Plasmopara viticola-inoculated control (C+P.v.) biological replicate no. 1; (H) C+P.v. biological replicate no. 2; (I) C+P.v. biological replicate no. 3; (J) P. viticola-inoculated T39-treated plants (T39+P.v) biological replicate no. 1; (K) T39+P.v biological replicate no. 2; and (L) T39+P.v biological replicate no. 3. Click here for filePrincipal component analysis of grapevine treatments. Principal component analysis (PCA) is based on the expression values (FPKM) of all grapevine genes for each sequencing replicate (named A and B) of each biological replicate (numbered from 1 to 3) for control (C), Trichoderma harzianum T39-treated (T39), Plasmopara viticola-inoculated control (C+P.v.), and P. viticola-inoculated T39-treated (T39+P.v.) plants. Click here for filePearson\u2019s correlation coefficients between sequencing and biological replicates of RNA-Seq analysis. Pearson\u2019s correlation coefficients are based on the gene expression values (FPKM) of all grapevine genes in two sequencing replicates (A and B) of each biological replicate (numbered from 1 to 3) for control (C), Trichoderma harzianum T39-treated (T39), Plasmopara viticola-inoculated control (C+P.v.), and P. viticola-inoculated T39-treated (T39+P.v.) plants. Click here for fileExpression levels, clustering, and annotation results of differentially expressed genes. Gene expression values (FPKM) and standard errors are reported for differentially expressed  genes in control (C), Trichoderma harzianum T39-treated (T39), Plasmopara viticola-inoculated control (C+P.v.), and P. viticola-inoculated T39-treated (T39+P.v.) plants. Differentially expressed genes were identified by the DESeq package with a false discovery rate (FDR) of 5% and a fold-change greater than two in at least one pairwise comparison. Fold-changes, clustering results, and functional annotation are reported for each gene. Click here for filePrimer sequences of the grapevine genes analysed by real-time RT-PCR. Forward and reverse primer sequences of real-time RT-PCR analysis are reported for 24 selected grapevine genes. Grapevine Actin (TC81781)  and VATP16 (XM_002269086.1) [94] were used as constitutive genes for normalising the real-time RT-PCR data. Click here for fileExpression levels of grapevine genes that are members of the actin gene family. Gene expression values (FPKM) and standard errors are reported for grapevine genes belonging to the Actin gene family in control (C), Trichoderma harzianum T39-treated (T39), Plasmopara viticola-inoculated control (C+P.v.), and P. viticola-inoculated T39-treated (T39+P.v) plants. Click here for file"}
+{"text": "Corrigendum to Anthony L. Cunningham, Andrew N. Harman, Najla Nasr (2013) Initial HIV mucosal infection and dendritic cells. EMBO Mol Med 5 (658\u2013660) DOI: 10.1002/emmm.201202763The name of the second author in this article, \u201cAndrew N. Harman,\u201d missed the middle initial N. The correct author names are listed above."}
+{"text": "O-Sulfated Heparan Sulfate Recognized by the Antibody HS4C3 Contributes to the Differentiation of Mouse Embryonic Stem Cells via Fas Signaling.The word \"Contributes\" is misspelled in the article title. The correct title is: 3-O-Sulfated Heparan Sulfate Recognized by the Antibody HS4C3 Contribute to the Differentiation of Mouse Embryonic Stem Cells via Fas Signaling. PLoS ONE 7(8): e43440. doi:10.1371/journal.pone.0043440 The correct citation is: Hirano K, Sasaki N, Ichimiya T, Miura T, Van Kuppevelt TH, et al. (2012) 3-"}
+{"text": "Long-term potentiation (LTP) at thalamic input synapses to the lateral nucleus of the amygdala (LA) has been proposed as a cellular mechanism of the formation of auditory fear memories. We have previously shown that signaling via ERK/MAPK in both the LA and the medial division of the medial geniculate nucleus/posterior intralaminar nucleus (MGm/PIN) is critical for LTP at thalamo-LA synapses. Here, we show that LTP-inducing stimulation of thalamo-LA inputs regulates the activation of ERK and the expression of ERK-driven immediate early genes (IEGs) in both the LA and MGm/PIN. Further, we show that pharmacological blockade of NMDAR-driven synaptic plasticity, NOS activation, or PKG signaling in the LA significantly impairs high-frequency stimulation-(HFS-) induced ERK activation and IEG expression in both regions, while blockade of extracellular NO signaling in the LA impairs HFS-induced ERK activation and IEG expression exclusively in the MGm/PIN. These findings suggest that NMDAR-driven synaptic plasticity and NO-cGMP-PKG signaling within the LA coordinately regulate ERK-driven gene expression in both the LA and the MGm/PIN following LTP induction at thalamo-LA synapses, and that synaptic plasticity in the LA promotes ERK-driven transcription in MGm/PIN neurons via NO-driven \u201cretrograde signaling\u201d. Fear conditioning is a type of associative learning that is widely studied as a model of learning and memory across a variety of species. Fear conditioning has been extensively characterized at the behavioral level, particularly auditory fear conditioning, in which a tone  is paired with footshock . In brief, auditory fear conditioning is thought to involve transmission and integration of sensory information from CS and US pathways within the lateral nucleus of the amygdala (LA), where alterations in synaptic transmission are believed to encode key aspects of the learning \u20133. In suLong-term potentiation (LTP), an experimental model of synaptic plasticity, is widely believed to be a potential mechanism by which fear conditioning promotes synaptic alterations in the LA , 6. In sin vitro slice recording methods have pointed to a role for a number of intracellular signaling pathways in LTP at thalamic input synapses to the LA, including CaMKII  or 60\u2009min , or 60\u2009min (ERK1: t(5) = 1.847, P > .05; ERK2: t(5) = 0.434, P > .05) time points. Further, this effect was not observed in LFS controls  = 0.725, P > .05; pERK2: t(5) = 0.759, P > .05), or 30\u2009min (pERK1: t(7) = 1.076, P > .05; pERK2: t(7) = 0.300, P > .05) time points following LFS. The findings for the LA are presented in Figures  Figures , left. I Figures , right; t(5) = 2.897, P < .05; pERK2: t(5) = 2.596, P < .05) and 30\u2009min after stimulation (pERK1: t(3) = 5.655, P < .05; pERK2: t(4) = 3.747, P < .05). No significant differences were observed for the 60\u2009min time point (pERK1: t(4) = 0.405, P > .05; pERK2: t(4) = 0.073, P > .05)  = 0.520, P > .05; ERK2: t(5) = 0.731, P > .05), 30\u2009min (ERK1: t(4) = 0.661, P > .05; ERK2: t(4) = 0.648, P > .05), or 60\u2009min (ERK1: t(4) = 1.131, P > .05; ERK2: t(4) = 0.195, P > .05) time points. Interestingly, LFS controls were also observed to have elevated levels of phospho-ERK1 and phospho-ERK2 in MGm/PIN at 5 minutes (pERK1: t(5) = 3.193, P < .05; pERK2: t(5) = 4.073, P < .05), but not at 30\u2009min (pERK1: t(5) = 0.529, P > .05; pERK2: t(5) = 0.067, P > .05)  = 1.551, P > .05; ERK2: t(5) = 0.691, P > .05) or 30\u2009min (ERK1: t(4) = 0.224, P > .05; ERK2: t(4) = 1.611, P > .05) time points following LFS. The fact that both HFS and LFS induced increases in ERK activation in the MGm/PIN at 5\u2009min suggests that the elevated ERK phosphorylation in MGm/PIN at this time point was due to local electrical stimulation. Importantly, at the 30\u2009min time point only HFS promoted significant ERK activation in the MGm/PIN, suggesting that the enhanced ERK phosphorylation in MGm/PIN at this later time point is specifically associated with LTP. The findings for the MGm/PIN are presented in Figures  Figures , left. I Figures , right. In our first series of experiments, we observed significant increases in ERK activation in the LA and MGm/PIN following HFS of the thalamo-LA pathway. In the present experiments, we used a combination of Western blotting and immunohistochemistry to examine whether LTP-inducing stimulation of thalamic input synapses to the LA regulate the ERK-driven IEGs Arc/Arg3.1, EGR-1 and c-Fos in the LA and the MGm/PIN. As before, anesthetized rats received either HFS or LFS of thalamic inputs to the LA and were sacrificed by decapitation 2 hours later Figures , a time t(8) = 6.502, P < .05; c-Fos: t(8) = 3.901, P < .05; EGR-1: t(8) = 5.273, P < .05; t(8) = 1.294, P > .05; c-Fos: t(8) = 0.241, P > .05; EGR-1: t(8) = 2.822, P > .05; t(8) = 3.642, P < .05; c-Fos: t(5) = 3.403, P < .05; EGR-1: t(6) = 2.59, P < .05; t(8) = 1.163. P > .05; c-Fos: t(5) = 0.094, P > .05; EGR-1: t(6) = 1.720, P > .05; The results of our Western blotting experiments are depicted in Figures t(5) = 6.894, P < .05; t(5) = 0.124, P > .05; t(5) = 3.822, P < .05; c-Fos: t(5) = 4.144, P < .05; Figures t(5) = 0.709, P > .05; c-Fos: t(5) = 0.121, P > .05; Figures Immunohistochemical localization of the three IEGs after HFS and LFS in both the LA and MGm/PIN can be seen in Figures t(6) = 22.556, P < .05], EGR-1 , and c-Fos  proteins  = 1.913, P > .05; EGR-1: t(5) = 0.227, P > .05; c-Fos: t(5) = 0.395, P > .05]. Rats receiving HFS exhibited Arc/Arg3.1, EGR-1, and c-Fos labeled cells throughout the MGm and PIN, while very few labeled cells were observed in the MGv  = 4.075, P < .05; pERK2: t(4) = 3.904, P < .05; t(4) = 0.173, P > .05; pERK2: t(4) = 1.273, P > .05] or 7-Ni , however, significantly impaired HFS-induced ERK activation in the LA (t(4) = 5.026, P < .05; pERK2: t(4) = 3.640, P < .05; The findings are depicted in Figures n the LA . Interest(4) = 3.035, P < .05; pERK2: t(4) = 3.864, P < .05; t(5) = 1.405, P > .05; pERK2: t(5) = 1.348, P > .05], 7-Ni , or c-PTIO . In the MGm/PIN, we observed a significant increase in phospho-ERK1/2 activation 30\u2009min after HFS in vehicle-infused controls [pERK1: Thus, blockade of NMDAR-driven synaptic plasticity and NO signaling at the level of the LA impairs HFS-induced ERK activation not only in the LA, but also in the MGm/PIN. Further, extracellular release of NO in the LA is required for HFS-induced ERK activation in the MGm/PIN, but not in the LA. In this series of experiments, we examined whether blockade of NMDAR-driven synaptic plasticity and NO signaling in the LA impairs HFS-induced expression of the IEGs Arc/Arg3.1, c-Fos, and EGR-1 in both the LA and the MGm/PIN. As in the previous experiment, anesthetized rats were given intra-LA infusion of ifenprodil, 7-Ni, or c-PTIO prior to LTP-inducing stimulation of the thalamo-LA pathway . In addit(7) = 3.440. P < .05; c-Fos: t(7) = 2.405, P < .05; EGR-1: t(7) = 5.009, P < .05; t(7) = 0.377. P > .05; c-Fos: t(7) = 0.708, P > .05; EGR-1: t(7) = 0.306, P > .05; t(7) = 2.629. P < .05; c-Fos: t(7) = 3.783, P < .05; EGR-1: t(7) = 3.440, P < .05; t(7) = 0.211. P > .05; c-Fos: t(7) = 0.592, P > .05; EGR-1: t(7) = 0.139, P > .05; The findings for rats infused with ifenprodil are depicted in Figures t(7) = 2.374. P < .05; c-Fos: t(7) = 2.462, P < .05; EGR-1: t(7) = 2.402, P < .05; t(7) = 0.672. P > .05; c-Fos: t(7) = 1.101, P > .05; EGR-1: t(7) = 1.499, P > .05; t(7) = 3.066. P < .05; c-Fos: t(7) = 2.383, P < .05; EGR-1: t(7) = 2.687, P < .05; t(7) = 0.065. P > .05; c-Fos: t(7) = 0.025, P > .05; EGR-1: t(7) = 0.460, P > .05; The findings for rats infused with 7-Ni are depicted in Figures t(7) = 2.374. P < .05; c-Fos: t(7) = 2.462, P < .05; EGR-1: t(7) = 2.402, P < .05; t(7) = 2.545. P < .05; c-Fos: t(7) = 2.406, P < .05; EGR-1: t(7) = 2.509, P < .05; t(7) = 3.066. P < .05; c-Fos: t(7) = 2.383, P < .05; EGR-1: t(7) = 2.687, P < .05; t(7) = 1.241. P > .05; c-Fos: t(7) = 0.696, P > .05; EGR-1: t(7) = 1.600, P > .05; The findings for rats infused with c-PTIO are depicted in Figures t(7) = 7.972. P < .05; c-Fos: t(7) = 3.686, P < .05; EGR-1: t(7) = 4.599, P < .05; t(6) = 1.688. P > .05; c-Fos: t(7) = 0.631, P > .05; EGR-1: t(7) = 1.287, P > .05; t(7) = 7.972. P < .05; c-Fos: t(7) = 4.064, P < .05; EGR-1: t(7) = 4.901, P < .05; t(6) = 1.688. P > .05; c-Fos: t(7) = 1.101, P > .05; EGR-1: t(7) = 0.401, P > .05; The findings for rats infused with PKG inhibitor Rp-8-Br-PET-cGMPS are depicted in Thus, similar to the findings of our ERK experiments, blockade of NMDAR-driven synaptic plasticity and NO signaling at the level of the LA impairs HFS-induced IEG expression not only in the LA but also in the MGm/PIN. Further, extracellular release of NO in the LA appears to be required for HFS-induced IEG expression in the MGm/PIN, but not in the LALong-term potentiation (LTP) at thalamo-LA synapses has been proposed as a candidate cellular mechanism of the formation of auditory fear memories, yet little is known about the molecular mechanisms underlying LTP at this synapse. In the present study, we have examined the regulation of ERK and that of three different ERK-driven IEGs at both sides of the thalamo-LA synapse after LTP-inducing stimulation. We found that LTP-inducing stimulation at thalamo-LA synapses is accompanied by ERK activation and ERK-driven gene expression not only in the LA, but also in regions of the MGm/PIN that are presynaptic to the LA. Further, pharmacological disruption of either NMDAR-driven synaptic plasticity or NO-cGMP-PKG signaling at the level of the LA impairs ERK activation and IEG expression in each region. Collectively, these findings suggest that NMDAR-driven synaptic plasticity and NO signaling within the LA coordinately regulate ERK activation and ERK-driven gene expression in both the LA and the MGm/PIN following LTP induction at thalamo-LA synapses.A recent study in our lab showed that LTP-inducing stimulation of thalamo-LA inputs induces ERK activation in the LA, and that intra-LA infusion of ERK/MAPK inhibitor impairs LTP at thalamo-LA synapses . HoweverOur findings also revealed that LTP-inducing stimulation of thalamo-LA synapses regulates the expression of the ERK-driven IEGs Arc/Arg3.1, c-Fos, and EGR-1 in both the LA and MGm/PIN. Previous studies have extensively documented the role of Arc/Arg3.1 \u201332 and EIn contrast to Arc/Arg3.1, both EGR-1 and c-Fos are thought to behave as transcription factors, regulating the expression of late-response genes that are critical for long-term synaptic plasticity. Importantly, several studies have observed associative increases in the expression of c-Fos and EGR-1 in the LA after cued fear conditioning \u201340. FurtOur findings of enhanced activation of ERK-driven transcriptional regulation in both LA and MGm/PIN neurons is consistent with previous work that has shown that infusion of a MEK inhibitor into either the LA or the MGm/PIN impairs LTP in the LA , 23 and in vitro [While most widely studied in the hippocampus , 58\u201363 ain vitro , 18.\u223c2-3\u2009mm distance in such a rapid manner. However, previous reports have suggested that retrograde transport can occur very rapidly in neurons, between \u223c4\u20138\u2009mm/hr [In the present study, we used pharmacological methods to ask whether NMDAR-driven synaptic plasticity and NO-cGMP-PKG signaling may be regulating both ERK and ERK-driven transcription within the LA and the MGm/PIN. We showed that blockade of NR2B (via ifenprodil), NOS (via 7-Ni), or PKG (via Rp-8-Br-PET-cGMPS) significantly impaired HFS-induced activation of ERK and ERK-driven gene expression in both the LA and the MGm/PIN. Remarkably, however, blockade of extracellular NO signaling (via c-PTIO) significantly impaired HFS-induced activation of ERK and ERK-driven gene expression in the MGm/PIN, but not in the LA. These findings suggest that the ERK activation and downstream IEG expression in MGm/PIN following LTP is driven by NO \u201cretrograde signaling\u201d at the level of the LA. The identity of the signal that links NO release at the level of the LA with ERK activation and ERK-driven gene expression at the level of the MGm/PIN is currently unknown, as is how such a signal may propagate in a retrograde manner from synapse to nucleus across a \u20138\u2009mm/hr . Further\u20138\u2009mm/hr , or inhi\u20138\u2009mm/hr , which m\u20138\u2009mm/hr . The present findings are consistent with a revised model of the molecular events underlying synaptic plasticity at thalamo-LA synapses in which NMDAR-driven synaptic plasticity and NO signaling in LA neurons promotes pre- and postsynaptic alterations at thalamo-LA synapses via regulation of ERK-driven gene expression in MGm/PIN and LA neurons, respectively, . In thatIn support of this model, recent studies from our lab and others have shown that ERK-driven transcription in the MGm/PIN is required not only for fear memory consolidation , 67, 68 In summary, our findings suggest that NMDAR-driven synaptic plasticity and NO signaling within the LA coordinately regulate ERK activation and ERK-driven gene expression in both the LA and the MGm/PIN following LTP induction at thalamo-LA synapses, and further suggest that synaptic plasticity in the LA promotes ERK-driven transcription in MGm/PIN neurons via NO-driven \u201cretrograde signaling\u201d. These findings further extend what is known about the molecular basis of LTP within the LA, and provide additional evidence that studying LTP at thalamo-LA synapses may inform us about the molecular basis of fear memory formation in the amygdala."}
+{"text": "These brain and renal injuries were associated with increased gene expression of NADPH oxidase components, NADPH oxidase activity and oxidative stress in brain and kidney tissues as well as systemic oxidative stress. Treatment with the ARB, olmesartan (10 mg/kg/day) reduced blood pressure in saline-drinking KK-Ay mice and attenuated cognitive decline, BBB disruption, glomerular injury and albuminuria, which were associated with a reduction of NADPH oxidase activity and oxidative stress in brain and kidney tissues as well as systemic oxidative stress. Furthermore, a suppressive dose of azelnidipine (3 mg/kg/day) exaggerated these beneficial effects of olmesartan. These data support the hypothesis that a CCB enhances ARB-associated cerebrovascular-renal protective effects through suppression of NADPH oxidase-dependent oxidative stress in type 2 diabetes.Recent clinical trials have demonstrated that combination therapy with renin-angiotensin system inhibitors plus calcium channel blockers (CCBs) elicits beneficial effects on cardiovascular and renal events in hypertensive patients with high cardiovascular risks. In the present study, we hypothesized that CCB enhances the protective effects of an angiotensin II type 1 receptor blocker (ARB) against diabetic cerebrovascular-renal injury. Saline-drinking type 2 diabetic KK-A The beneficial effects of renin-angiotensin system (RAS) inhibition with angiotensin converting enzyme inhibitors (ACEIs) and angiotensin II type 1 receptor blockers (ARBs) have been demonstrated in hypertensive patients with high cardiovascular-renal risks including heart failure , diabetePrevious large-scale epidemiological studies have indicated that CKD is not related to the incidence of stroke . Howevery mice treated with the ARB, olmesartan [It has been highlighted that type 2 diabetes and hypertension are risk factors for cerebrorenal injury including cognitive decline ,22, BBB mesartan .y mice [Experimental protocols and animal care were performed according to the guidelines for the care and use of animals established by Kagawa University, Japan. The experiments were approved by the Animal Experimentation Ethics Committee at Kagawa University (No. 112). At the end of the experiment, organs were dissected under sodium pentobarbital anesthesia . Six-week-old male type 2 diabetic KK-Ay mice  and contn = 11) and saline-drinking C57BL6 mice . Type 2 diabetic KK-Ay mice were divided into four groups: tap water drinking KK-Ay mice ; saline-drinking KK-Ay mice ; saline-drinking KK-Ay mice treated with olmesartan  ; saline-drinking KK-Ay mice treated with olmesartan plus azelnidipine  . It has previously been reported that azelnidipine at 3 mg/kg body weight/day did not change blood pressure in KK-Ay mice [y mice .After a two week period of acclimatization and measuring basal parameters, 8-week-old mice were underwent combination treatments for 16 weeks. Control C57BL6 mice were divided into two groups: tap water drinking C57BL6 mice  was monitored in conscious mice by tail-cuff plethysmography . Urinary albumin and creatinine concentrations were measured by using commercially available assay kits  . PostpraAt the end of the experiment blood, brain and kidney samples were harvested under anesthesia with sodium pentobarbital . The brain and kidney tissues were harvested and fixed in 10% buffered paraformaldehyde or embedded in Tissue-Tek OCT compound , and remaining tissues were snap-frozen in liquid nitrogen. Small amounts of brain and renal cortical tissues were collected in RNAlater and stored overnight at 4\u00b0C RNAlater-treated samples were subsequently snap-frozen in liquid nitrogen and stored at -80\u00b0C until processing for RNA extraction and reverse transcription-polymerase chain reaction (RT-PCR) analysis.A passive avoidance test was performed to evaluate cognitive function as described previously . Detailsn = 6 per group) as previously described [BBB permeability was determined using the EB extravasation technique  and Mallory-Azan reagents to evaluate glomerular sclerosis and tubulointerstitial fibrosis, respectively ,32. The Glomerular podocyte injury was evaluated by immunohistochemical analysis of desmin and was performed as previously described ,31. To investigate oxidative stress in the brain and kidney tissues, DHE immunofluorescence staining was performed as previously described . DetailsNADPH oxidase-derived superoxide anion  generation was measured using lucigenin-enhanced chemiluminescence, as described previously . Details The mRNA expression in brain and renal cortical tissues were analyzed by RT-PCR using a LightCycler FastStart DNA Master SYBR Green I kit and an ABI Prism 7000 Sequence Detection System  as previously described ,31. The Plasma and urine 8-hydroxy-2\u2019-deoxyguanosine (8-OHdG) , plasma level of non-esterified fatty acid (NEFA), triglyceride (TG), total cholesterol (TCho) , and insulin  were measured using commercially available kits.post hoc test. Values of P < 0.05 were considered statistically significant. All values are presented as means \u00b1 S.E.M.. Statistical comparisons of differences among groups were performed using one-way repeated-measures analysis of variance (ANOVA), followed by the Newman-Keuls y mice showed elevated SBP compared with that in age-matched C57BL6 or C57BL6 + 0.9% NaCl mice   . In addi tissues . These dy mice, DHE staining was significantly increased in kidney tissues compared with that in C57BL6 mice Click here for additional data file.Table S1(DOCX)Click here for additional data file.Figure S1Body weight changes during the experimental period. KK-Ay mice showed higher body weight compared to C57BL mice. However, none of the treatments affected body weight gains in KK-Ay + 0.9% NaCl mice (n=11). aP < 0.05 vs. C57BL/6, bP < 0.05 vs. C57BL/6 + 0.9% NaCl.(TIF)Click here for additional data file.Figure S2NADPH oxidase subunits gene expression in brain tissues analyzed by RT-PCR. NADPH oxidase subunits gp91phox (A) and p47phox (B) mRNA levels in whole brain tissues. Saline-dinking KK-Ay mice showed upregulation of NADPH oxidase subunit mRNA levels in brain tissues, which were attenuated by treatment with olmesartan. Furthermore, the combination of olmesartan plus azelnidipine completely prevented these changes resulting in levels similar to that in C57BL6 mice (n=8). aP < 0.05 vs. C57BL/6, bP < 0.05 vs. C57BL/6 + 0.9% NaCl, cP < 0.05 vs. KK-Ay, dP < 0.05 vs. KK-Ay + 0.9% NaCl, eP < 0.05 vs. KK-Ay + 0.9% NaCl + olmesartan.(TIF)Click here for additional data file.Figure S3NADPH oxidase subunits gene expression in kidney tissues analyzed by RT-PCR. NADPH oxidase subunits gp91phox (A) and p22phox (B) mRNA levels in renal cortical tissues. In saline-dinking KK-Ay mice, superoxide production in renal tissues was associated with upregulation of NADPH oxidsase subunit genes expression. Treatment with olmesartan markedly attenuated these changes. Furthermore, the combination of olmesartan plus azelnidipine completely prevented these changes resulting in levels similar to that in C57BL6 mice (n=8). aP<0.05 vs. C57BL/6, bP<0.05 vs. C57BL/6 + 0.9% NaCl, cP<0.05 vs. KK-Ay, dP<0.05 vs. KK-Ay + 0.9% NaCl, eP<0.05 vs. KK-Ay + 0.9% NaCl + olmesartan.(TIF)Click here for additional data file."}
+{"text": "Salmonella enterica strains that are representatives of the S. enterica serovar Typhimurium complex in reference collection A (SARA) are closely related but exhibit differences in antibiotic resistance, which could have public health consequences. To better understand the mechanisms behind these resistances, we sequenced the genomes of two multidrug-resistant strains: SARA64 (Muenchen) and SARA33 (Heidelberg).The  Salmonella enterica is one of the most important bacterial enteric pathogens and has been implicated in food-borne illnesses worldwide .orldwide . Emergenorldwide , 3. Explorldwide , 5, consintI1) . Two of these strains, SARA64 and SARA33, exhibited resistance to ampicillin, chloramphenicol, tetracycline, streptomycin, sulfisoxazole, and kanamycin. SARA33 also showed resistance to gentamicin. Both strains were positive for the integrase found in integrons class I (intI1) ; howeverintI1) . In ordede novo assembled using CLC Genomics Workbench version 5.5.1 . The G+C mol% values of SARA64 and SARA33 were 52.0 and 52.1%, respectively, which are similar to the reported GC content for other Salmonella strains . The genomes were sequenced using an Ion Torrent (PGM) sequencing system with the 200-bp reads chemistry  at 30 to 40\u00d7 coverage, using an Ion PGM 200 sequencing kit, according to the manufacturer\u2019s instructions. Genomic sequence contigs for each strain were  strains . Strain http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html) (in silico multilocus sequence typing (MLST) (http://cge.cbs.dtu.dk/services/) -laa, aph(3\u2032)-la, strA, strB, and aadA1], sulfonamides (sul1 and sul2), beta-lactams (blaOXA), tetracycline (tetB), and phenicol (catA1), which explain its MDR phenotype. Some of these resistance genes were on a genomic island similar to GI-DT12 in Salmonella enterica serovar Typhimurium T000240 (intI1-blaOXA-aadA1-qacE\u03941-sul1). The genes strA, strB, and sul2 were located on a plasmid.These draft genome sequences were annotated using the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (ne.html) . Identitrvices/)  using thdatabase ; strain database . Antibiocreening . SARA64  T000240 , which caac(6\u2032)-ly, aadA5, aadB, aa(6\u2032)-33, and aadA1], sulfonamides (sul1 and sul2), beta-lactams (blaOXA-2 and blaTEM), and tetracycline (tetD). A novel integron cassette was identified that contained one hypothetical protein (hp) with unknown function, followed by aadA1 and aa(6\u2032)-33 genes [intI1-hp-aac(6\u2032)-33-aadA1].In contrast, SARA33 carried resistance genes for aminoglycosides [Salmonella enterica strains are now available in GenBank under accession numbers AUQD00000000 for strain SARA33 (2213-Heidelberg) and AUQE00000000 for strain SARA64 (2244-Muenchen).The draft genome sequences of these two"}
+{"text": "Immunohistochemical studies suggest that the rheumatoid nodule (RN) is a Th1 granuloma , yet theWe measured circulating cytokines of DMARD-na\u00efve early RA patients, and compared profiles of patients with RN to those without RN.A cross-sectional cohort of 149 DMARD-na\u00efve adults with early RA (symptom duration \u2264 2 years). Assessments included a smoking history, the presence of subcutaneous RN, the Simplified Disease Activity Index (SDAI), and X-rays scored using the modified Larsen method, Rheumatoid Factor (RF) and anti cyclic citrullinated peptides (aCCP) tests. Serum cytokines were measured using the Bio-Plex\u00ae suspension array system. Differences between patients with RN and those without RN were explored using univariate and multivariate techniques, correcting for SDAI by regression analysis.Of 149 patients , the majority were Black Africans (93%) and 34 (22.8%) had RN. Patients with RN were more likely to be males, but neither smoking history nor symptom duration was significantly different between the two groups. RN patients had severe RA with higher SDAI (p=0.04) and X-ray scores (p=0.004), and higher RF (p=0.005) and aCCP (p=0.04) titers. Cytokine profiles in the RN patients showed significant elevation of predominantly Th1 cytokines, with higher interferon-gamma (IFN-G) (p=0.05), interleukin (IL)-12 (p=0.02), and IL-2 (p=0.05). In addition, cytokines of macrophage  and fibroblast origin  were significantly higher in these patients. Multivariate analysis showed IL-7 (p<0.001) and IFN-G (p=0.001) to be independently significant.DMARD-na\u00efve early RA patients with RN have a unique serum cytokine profile, with significant elevation of predominantly Th1 and fibroblast cytokines."}
+{"text": "Diastolic dysfunction (DD) is common in patients with cardiovascular disease. Current cardiac magnetic resonance techniques for assessment of DD require the acquisition of tagged imaging sequences, and complex post-processing. A new post-processing software allows for rapid strain assessment using feature tracking analysis (FT)  based on conventional steady-state free precession (SSFP)-Cine sequences. The aim of this study was to compare tagging (TAG) with FT for assessment of diastolic function in patients with DD and healthy controls.20 healthy volunteers and 10 patients with echocardiographic diagnosed DD Grade II-III were investigated by the 2 techniques. Cardiovascular magnetic resonance imaging (Philips Intera 1.5T) using CSPAMM and SSFP-Cine sequences was performed for matched mid-ventricular short-axis slices. Each modality was analyzed offline using dedicated post-processing software  and early-diastolic strain rate (EDSr) was calculated from both datasets. EDSr derived from CSPAMM and SSFP-Cine data were compared and inter-observer and intra-observer variability assessed using Bland-Altman analysis and Pearson correlation coefficient.-1) was highly correlated with TAG EDSr (76.71+/- 17.45s-1), Bland-Altman analyses for method comparison yielded a mean difference of 1.4 (95% CI: 0.0 to 2.9). Limits of agreement (+/- 1.96 SD) were -14.9 and 12.0. In patients with DD FT EDSr (61.40+/- 15.60s-1) was not significantly different to TAG EDSr (57.60+/- 12.30s-1) (p=ns). The same was found in the control group FT EDSr (86.52+/- 10.55s-1) and TAG (86.27+/- 10.31s-1) (p=ns). Intra-observer variability yielded a mean difference of 1.2+/- 4.9 (FT) and 0.9+/- 3.5 (TAG). Inter-observer variability displayed a mean difference of 2.8+/- 5.84 (FT) and 0.3+/- 3.7 (TAG). Pearson correlation coefficient was (0.96 (FT); 0.97 (TAG)) for intra-observer variability and (0.94 (FT); 0.97 (TAG)) for inter-observer variability.For all measurements FT EDSr (78.15+/- 17.17 sAssessment of EDSr using a novel feature tracking technique shows good agreement with tagging derived data and may thus be used for rapid clinical determination of diastolic dysfunction. However, compared to feature tracking, tagging revealed lower intra - and inter-observer variability and may therefore remain the preferred technique for research applications.None."}
+{"text": "AbstractCerapachys Smith are keyed. Twelve species are recognized of which 6 are described as new. The species are: Cerapachys aitkenii Forel, Cerapachys aliisp. n., Cerapachys anokhasp. n., Cerapachys besucheti Brown, Cerapachys biroi Forel, Cerapachys indicus Brown, Cerapachys longitarsus (Mayr), Cerapachys nayanasp. n., Cerapachys schoedlisp. n., Cerapachys seemasp. n., Cerapachys sulcinodis Emery and Cerapachys wightisp. n. Geographic distribution and group affinities of the new species are discussed. A revised key to the Indian species is provided. The rare ergatoid queens of Cerapachys nayana, Cerapachys schoedli and Cerapachys seema are reported. Formed in response to selective pressures these ergatoid queens have a significant role in dispersal strategies and contribute much to our understanding of the biology of these ants.The Indianspeciesof the ant genus  Cerapachys includesmainly myrmecophagous ants which raid the nests of other ants for prey .The ant genus for prey . The genfor prey . Cerapacglobally . The triCerapachys in India is currently represented by 6 species  flank to the midpoint of the frontovertextal margin  to the posterior extension of the propodeal lobes = Weber\u2019s length measured from WL  to the dorsal edge of the mesosomaMH IIIAL  software. Later, images were cleaned as per requirement with Adobe Photoshop CS6. Description style and morphological terminology for measurements and indices follow l margin ; HW = Hesal view ; EL = Eye of eye ; SL = Sc condyle ; WL = Weal lobes ; MH = Memesosoma ; PrW = Psal view ; PL1 = Psal view ; PW1 = Psal view ; IIIAL =helcium) ; IIIAW =sal view ; IVAW = sal view ; IVAL = etergite ; CI = Ce Natural History Museum, London, U.K.BMNH Punjabi University Patiala, Ant Collection at Department of Zoology and Environmental Sciences, Punjabi University, Patiala, Punjab, India.PUACPageBreakhttp://zoobank.org/52540038-860B-4EA4-BA1E-40217FF6D555http://species-id.net/wiki/Cerapachys_alii10\u00b045'N, 76\u00b044'E, 118m a.s.l., 10.x.2011, Winkler method (coll. Shahid A. Akbar); Holotype in PUAC and paratype in BMNH.Holotype and 6 paratypes (worker): India, Kerala, Salim Ali Bird Sanctuary, Measurements (holotype in brackets): HL 0.46-0.51 (0.48); HW 0.37-0.39 (0.38); WL 0.47-0.49 (0.49); MH 0.28-0.31 (0.31); PrW 0.25-0.29 (0.25); PL1 0.16-0.20 (0.17); PW1 0.16-0.19 (0.18); IIIAL 0.20-0.22 (0.22); IIIAW 0.22-0.27 (0.23); SL 0.21-0.22 (0.22); IVAL 0.43-0.49 (0.49); IVAW 0.39-0.41 (0.41). Indices: CI 76-80 (79); SI 56-57 (57); PI 95-105 (105) (n=5).rd of its length.Head. Rectangular, longer than broad, sides converge anteriorly; vertexal margin concave, posterior lateral corners rounded. Parafrontal ridges prominent, raised. Eyes absent. Mandibles dentate; narrow, with strongly incurved apical tooth; anterior clypeal margin entire and projects forward as a low rounded transparent lobe or apron. Lateroclypeal teeth reduced. Antennae 9 segmented; scapes short, clavate, each falling short of posterior margin of head by 1/3Mesosoma. Stout, wider anteriorly; dorsal surface slightly convex, almost flat, the dorsal surface gently rounded along sides without any distinct margin. Declivous face of propodeum with cariniform margins across the top and along lateral margins.Metasoma. Petiole as long as broad, without overhanging dorsolateral margins. Anterior face transverse and posterior face shallowly convex. Subpetiolar process prominent, acute, posteriorly directed; no fenestra present. Postpetiole slightly longer than broad, lateral angles uniformly rounded. Gaster elongate; base of cinctus of first gastral tergite with cross ribs; sting exerted.Sculpture. Mandibles punctured. Head strongly foveate. Mesosoma, petiole and postpetiole with similar prominent foveate sculpture.Vestiture. Body with reduced white pilosity; moderate, decumbent or subdecumbent hairs distributed evenly throughout. Apical funicular segments and legs with small standing hairs.Colour. Dark red with mandibles, antennae and legs castaneous.PageBreakThe species is named in honor of Dr. Salim Ali, renowned Indian Ornithologist.Cerapachys alii can be easily separated from other species known from India. Only eight other known species of Cerapachys are reported to have 9 segmented antennae. These eight species are placed in the typhlus group and include; Cerapachys biroi Forel, 1907; Cerapachys cryptus Mann, 1921; Cerapachys edentatus Forel, 1900; Cerapachys fuscior Mann, 1921; Cerapachys papuanus Emery, 1897; Cerapachys pawa Mann, 1919; Cerapachys pusillus Emery, 1897 and Cerapachys typhlus Roger, 1861. The new species can be easily separated from all of them. Cerapachys cryptus and Cerapachys fuscior are larger species (HW > 0.70 mm) while Cerapachys alii is a smaller species (HW< 0.40 mm). Cerapachys typhlus has the postpetiole more than half as long as the succeeding gastric segment while in Cerapachys alii it is less than half as long as the succeeding gastric segment. Cerapachys papuanus, Cerapachys pawa and Cerapachys pusillus have the anterolateral shoulders of the first gastric segment abruptly rounded, accentuating the medium concavity that receives the postpetiole while in Cerapachys alii theanterolateral shoulders of the first gastric segments as seen from above broadly rounded and gradually widening caudad. Cerapachys biroi and Cerapachys edentatus predominantly have punctuate body sculpture while Cerapachys alii has predominantly foveate body sculpture. Cerapachys alii can also be confused with Cerapachys fragosus Roger, 1862 and Cerapachys coecus Mayr, 1897 which has similar prominent foveate body sculpture, however these two species are characterized by 11 segmented antennae while as Cerapachys alii has 9 segmented antennae.With its 9 segmented antennae Ecology. This subterranean species seems to be of rare occurrence as it was encountered only once during the extensive surveys in the region. The specimens were collected from a leaf litter sample taken from Salim Ali bird Sanctuary. A low-land evergreen forest area, located between the branches of Periyar river. The region is considered as the richest bird habitat on peninsular India.http://zoobank.org/2A8BE8D2-BED1-4DAC-A73F-E9AC2189FE23http://species-id.net/wiki/Cerapachys_anokha9\u00b0.30'N, 77\u00b0.16'E, 1003m a.s.l., 15.x.2011, hand picking method (coll. Shahid A. Akbar). Holotype in PUAC and paratype in BMNH.Holotype and 3 paratypes (worker): India: Kerala, Periyar tiger reserve, Thanikkudy, Measurements (holotype in brackets): HL 0.69-0.73 (0.72); HW 0.60-0.63 (0.63); EL 0.20-0.22 (0.22); WL 0.72-0.80 (0.80); MH 0.33-0.38 (0.38); PrW 0.42-0.47 (0.47); PL1 0.29-0.33 (0.33); PW1 0.38-0.41 (0.41); IIIAL 0.38-0.45 (0.41); IIIAW 0.44-0.55 (0.52); SL 0.29-0.32 (0.32); IVAL 0.85-0.92 (0.92); IVAW 0.57-0.64 (0.64). Indices: CI 86-87 (87); SI 48-51 (51); PI 124-131 (124) (n=4).PageBreakParafrontal ridges present. Eyes prominent, placed below midline of head. Mandible triangular with acute apices and sharp concave, edentate, masticatory margins; anterior clypeal margin with small apron shaped transparent structure. Lateroclypeal teeth small, blunt and projecting slightly inwards. Antennae 12 segmented; scape short and clavate, reaching up to 1/3rd of posterior margin of head.Head. Rectangular, longer than broad, widest at about mid-length; sides parallel; vertexal margin slightly concave, posterior lateral corners are weakly acute to rounded. Mesosoma. Stout, rectangular in dorsal view; dorsal surface convex, the dorsal surface gently rounded along sides without any distinct margin. Declivous face of propodeum lacking cariniform margins across the top and along sides.Metasoma. Petiole highly convex, broader than long, with traces of reduced dorsolateral margins, anterior and posterior faces continuous with dorsum. Subpetiolar process prominent, wedge like, with apex directed backward; no fenestra present. Postpetiole wider than long, uniformly rounded. Gaster elongate; base of cinctus of first gastral tergite with cross ribs; sting exerted.st gastral, with few cross ribs.Sculpture. Mandibles smooth and shining. Head with few small punctures. Sculpture on dorsal surface of mesosoma, petiole and postpetiole consist of very small, uniform punctures, distributed throughout the surface. Gaster mostly smooth, with few scattered punctures. Cinctus of 1Vestiture. Body covered with decumbent or subdecumbent hairs. Longer hairs are also present on postpetiole and gaster. Head also consists of few long hairs; apical funicular segments and legs with standing hairs.Colour. Black with mandibles, antennae and legs castaneous.The species epithet is Hindi for \u201cunique\u201d, in reference to its unique nature of propodeal declivity.PageBreakCerapachyinae. The new species show interesting variation in the form of the petiole. The petiolar node has the inferior as well as the superior posterolateral angles produced, but not sharply angular. The sides of the petiole could not be considered either immarginate (Cerapachys lineage) or marginate (Phyracaces lineage). This makes the placement of this species somewhat transitional between the two lineages and easily distinguishes it from other reported species of the genus. When using Cerapachys anokha comes close to singaporensis Viehmeyer, 1916. The two species however can be easily separated. Cerapachys singaporensis has the body arrayed with long pale hairs; and copiously pubescent, and the dorsal sides of the petiole strongly marginate, while Cerapachys anokha has only decumbent or subdecumbent body hairs, little pubescence and the dorsolateral sides of petiole are not marginate. Cerapachys anokha could also be confused with Cerapachys nayana which has similar habitat preferences and body colouration; however the two species can be easily separated: Cerapachys nayana has larger eyes (EL 0.24\u20130.27 mm), the declivous face of the propodeum has cariniform margins across the top, and the petiole has marginate dorsolateral sides; while Cerapachys anokha has smaller eyes (EL 0.20\u20130.22 mm), the declivous face of its propodeum lacks cariniform margins across the top, and the petiole is without marginate dorsolateral sides.This species is unique in having the declivous face of the propodeum lacking cariniform margins across the top and along the sides, features unique in described workers of the This species seems to be infrequent. It is from the Western Ghats. Four specimens were collected by handpicking from the Thanikkudy region of the Periyar tiger reserve. Which is a primary, undisturbed tropical moist evergreen forest. The area is situated at 1003 meters elevation. It is a shady place with little sunlight penetration.http://zoobank.org/46F8FF78-D753-4BEF-8FCB-7A83D4149E8Dhttp://species-id.net/wiki/Cerapachys_nayana11\u00b05'N, 76\u00b026'E, 897m a.s.l., 25.ix.2011, hand picking. Paratypes: 2 workers and 1 ergatoid queen with same data as holotype; 6 workers and 2 ergatoid queens, India, Karnataka, Gundlupet 11\u00b08'N, 76\u00b068'E, 800m a.s.l., 27.ix.2010, hand picking; 2 workers and 1 ergatoid queen, India, Kerala, Periyar tiger reserve, 9\u00b046'N, 77\u00b014'E, 1005m a.s.l., 10.x.2011, hand picking (coll. Shahid A. Akbar). Holotype in PUAC and paratype in BMNH.Holotype worker: India: Kerala, Silent valley national park, Measurements (holotype in brackets): HL 0.55-0.66 (0.66); HW 0.48-0.51 (0.51); EL 0.24-0.27 (0.25); WL 0.60-0.66 (0.66); MH 0.34-0.37 (0.34); PrW 0.33-0.38 (0.34); PL1 0.20-0.23 (0.23); PW1 0.25-0.29 (0.29); IIIAL 0.30-0.44 (0.44); IIIAW 0.37-0.47 (0.37); SL 0.21-0.27 (0.27); IVAL 0.58-0.61 (0.61); IVAW 0.47-0.52 (0.52). Indices: CI 77-87 (77); SI 44-53 (53); PI 125-126 (126) (n=10).PageBreakth of posterior margin of head.Head. Rectangular, longer than broad, widest at about its midlength; sides parallel, vertexal margin transverse to shallowly concave, posterior lateral corners weakly acute. Parafrontal ridges present but not raised, very low. Eyes large prominent. Mandibles subtriangular; masticatory margin without a row of small denticles. Lateroclypeal teeth small and reduced. Antennae 12 segmented; scapes short, reaching up to 4/5Mesosoma. Moderately stout, rectangular in dorsal view; dorsal surface flattened, bordered laterally by a distinct angle, but no margin. Declivous face of propodeum with cariniform margins across the top and along the lateral margins.Metasoma. Petiole broader than long, with strong overhanging dorsolateral margins. Anterior face concave while posterior face is transverse. Subpetiolar process small with stout acute apex, directed forward, located beneath anterior 1/3rd of the petiole; no fenestra present. Postpetiole sub trapezoidal, wider behind with the posterolateral angles uniformly rounded. Gaster elongate; base of cinctus of first gastral tergite with cross ribs; sting exerted.st gastral segment smooth and shining.Sculpture. Mandibles smooth and shining. Head with small punctures, spaced wider than their diameter, dorsum of head also with faint rugae in between the punctures. Similar sculpture on dorsal surface of mesosoma and petiole. Small continuous punctures produce a matt like appearance on the dorsum of the postpetiole. Gaster with similar matt-like appearance but less prominent. Cinctus of 1Vestiture. Body covered with moderate decumbent or subdecumbent hairs most prominent on postpetiole and gaster, head devoid of such hairs, only a few along sides; apical funicular segments with standing hairs.PageBreakColour. Black with mandibles, antennae and legs castaneous.HL 0.66-0.77; HW 0.55-0.60; EL 0.22-0.24; WL 0.79-0.82; MH 0.36-0.41; PrW 0.44-0.47; PL1 0.24-0.27; PW1 0.44-0.47; IIIAL 0.44-0.47; IIIAW 0.51-0.55; SL 0.18-0.22; IVAL 0.58-0.63; IVAW; 0.60-0.66. Indices: CI 77-83; SI 33-37; PI 174-183 (n=3).Like the workers of the same colony, but larger, with thicker body, especially mesosoma and gaster. Ocelli present on vertex, prominent. The pilosity is much more prominent compared to workers. Distinction between ergatoid queens and worker is vague, with size variation of workers very high.There is a considerable amount of size variation between individual specimens, the smaller workers are lighter in body colouration compared with larger specimens; the body sculpture and pilosity also differs between individuals.The species epithet is Sanskrit for \u201ceyes\u201d, in reference to the large eye size of the species.PageBreakCerapachys anokha from which it is separated by the combination of characters given in the diagnosis of the latter species. Cerapachys nayana is compared with Cerapachys longitarsus which also has marginate dorsolateral sides to the petiole; however, the two species can be easily separated. Cerapachys longitarsus has characteristic bicolouration, with the head brown, trunk red or brown, petiole and postpetiole light to dark reddish and the gaster brown or black, while Cerapachys nayana is uniformly black coloured with mandibles, antennae and legs castaneous. The new species also resembles the Philippines, Cerapachys luzuriagae  but can be easily separated from it. Cerapachys nayana is coloured black with the petiole broader than long, and with concave anterior and transverse posterior faces, the postpetiole with dense punctures and without dentition; while Cerapachys luzuriagae is reddish brown with the petiole as long as broad, with convex anterior and truncate posterior faces; postpetiole without dense punctures, and mandibles with prominent dentition.With themarginate dorsolateral sides of its petiole, this species is easily distinguished from other Indian species of its genus. The new species sharesmost characters with This species is widely distributed in the Western Ghats. It was collected from non-forested and forest habitats from small bushes, and foraging over dry soil surfaces.http://zoobank.org/38E7EC5E-E9F2-4FCC-B467-10E9208AA155http://species-id.net/wiki/Cerapachys_schoedli11\u00b05'N, 76\u00b026'E, 897m a.s.l., 25.ix.2011, Winkler. Paratypes: 13 workers and 3 ergatoid queens, same data as holotype; 2 workers, India, Kerala, Salim Ali Bird Sanctuary, 10\u00b045'N, 76\u00b044'E, 118m a.s.l., 6.xi.2011, Winkler; 10 workers, India, Kerala, Periyar tiger reserve, Manalar, 9\u00b035'N, 77\u00b018'E, 1630m a.s.l., 27.x.2011, hand picking (coll. Shahid A. Akbar). Holotype in PUAC and paratype in BMNH.Holotype worker: India. Kerala, Silent valley national park, Measurements (holotype in brackets): HL 0.64-0.68 (0.68); HW 0.44-0.46 (0.46); EL 0.07-0.11 (0.11); WL 0.62-0.69 (0.69); MH 0.41-0.48 (0.48); PrW 0.31-0.35 (0.35); PL1 0.24-0.27 (0.27); PW1 0.28-0.31 (0.31); IIIAL 0.28-0.32 (0.32); IIIAW 0.39-0.41 (0.41); SL 0.31-0.33 (0.33); IVAL 0.70-0.74 (0.74); IVAW 0.58-0.60 (0.60). Indices: CI 67-69 (67); SI 70-72 (72); PI 114-116 (114) (n=11).rd of its length.Head, rectangular, longer than broad; sides parallel; vertexal border transverse. Posterior lateral corners acute. Parafrontal ridges present, raised. Eyes medium sized, almost circular. Mandibles subtriangular; masticatory margins without a row of small denticles. Lateroclypeal teeth small. Antennae 12 segmented; scapes short, each falling short of posterior margin of head by 1/3Mesosoma. stout, humped in profile view; dorsal surface convex, continuous with sides, no lateral margins. Declivous face of propodeum with cariniform margins across the top and along the lateral margins.Metasoma. Petiole broader than long, and gently rounded towards the sides. Anterior and posterior faces transverse. Subpetiolar process stout, fenestra present. Postpetiole almost rectangular, wider behind, with the posterolateral angles not tuberculate but uniformly rounded. Gaster elongate; base of cinctus of first gastral tergite with cross ribs; sting exerted.Sculpture. Mandibles with small punctures. Head smooth and shining with some punctures. Mesosoma mostly smooth and shining with some punctures along the sides. Petiole, postpetiole and gaster with continuous punctures.PageBreakVestiture. Body covered with moderate, decumbent or subdecumbent hairs, most prominent on gaster; apical funicular segments and legs also with standing hairs.Colour. Bright yellowish orange to dark red.PageBreakHL 0.80-0.84; HW 0.59-0.63; EL 0.14-0.16; WL 0.88-0.92; MH 0.51-0.55; PrW 0.49-0.53; SL 0.41-0.43; PL1 0.29-0.31; PW1 0.38-0.41; IIIAL 0.44-0.50; IIIAW 0.57-0.59; IVAL 0.88-0.90; IVAW 0.86-0.88. Indices: CI 73-75; SI 68-69; PI 131-132 (n=3).Like the workers of the same colony, but larger, with more stout body, especially the mesosoma and gaster. Ocelli absent on vertex. Distinction between ergatoid queens and worker is vague, with size variation of workers very high.The species is named in the honor of the late Dr. Stefan Sch\u00f6dl.Cerapachys schoedli sharesmost characters with Cerapachys seema, from which it can be easily distinguished by the combination of characters given in the diagnosis of the latter species. The new species can also be compared with Cerapachys luteoviger Brown, 1975 which also has small punctures on the cephalic dorsum, with diameter lesser than the average distance separating them. However, the peculiar petiolar node  and rounded head shape of Cerapachys luteoviger easily separates it from Cerapachys schoedli, which has the petiolar node broader than long and the posterior lateral corners of the head acute.This species is aberrant in many characters, with the cephalic dorsum bearing small punctures with average diameter lesser than the average distance separating them, a highly shinning body and reduced body sculpture, which separates it from other reported Indian species. This species seems to be common in the Western Ghats; it was collected in non-forest as well as forest habitats in leaf litter and on dry soil surfaces.http://zoobank.org/AE131489-5514-422C-BE6E-5BDBCCA2F111http://species-id.net/wiki/Cerapachys_seema9\u00b035'N, 77\u00b018'E, 1630m a.s.l., 24.x.2011, hand picking. Paratypes: 4 workers, 3 ergatoid queens and 1 gyne, same data as holotype (coll. Shahid A. Akbar). Holotype in PUAC and paratype in BMNH.Holotype worker: India. Kerala, Periyar tiger reserve, Manalar, Measurements (holotype in brackets): HL 0.72-0.74 (0.74); HW 0.52-0.56 (0.56); EL 0.07-0.19 (0.19); WL 0.77-0.82 (0.77); MH 0.38-0.45 (0.45); PrW 0.37-0.40 (0.38); PL1 0.23-0.29 (0.26); PW1 0.36-0.41 (0.41); IIIAL 0.41-0.46 (0.41); IIIAW 0.51-0.54 (0.51); SL 0.33-0.41 (0.41); IVAL 0.70-0.74 (0.74); IVAW 0.62-0.69 (0.63). Indices: CI 72-75 (75); SI 63-73 (73); PI 141-157 (157) (n=9).rd the distance to the posterior margin of head.Head, longer than broad; sides converging posteriorly; vertexal margin transverse. Posterior lateral corners weakly acute. Parafrontal ridges prominent, raised. Eyes small. Mandibles subtriangular; masticatory margins deflexed and downcurved, with a row of small denticles. Lateroclypeal teeth prominent. Antennae 12 segmented; scapes clavate, reaching up to 2/3PageBreakMesosoma moderately stout, rectangular in dorsal view; dorsal surface flattened and gently rounded towards the sides. Declivous face of propodeum with cariniform margins across the top and along its lateral margins.PageBreakrd of the petiole; fenestra present. Postpetiole broader than long with posterolateral angles uniformly rounded. Gaster elongate; base of cinctus of first gastral tergite with cross ribs; sting exerted.Metasoma, Petiole broader than long, lacking dorsolateral margins. Anterior and posterior faces transverse. Subpetiolar process well developed, located below the anterior 1/3st gastral segment with prominent transverse ribs.Sculpture. Mandibles smooth with few punctures. Head with prominent punctures, spaced more widely than their diameter. Similar sculpture on dorsal surface of mesosoma. Petiole and postpetiole with larger punctures, forming a rugae-like surface on the dorsum. Gaster with similar sculpture to mesosoma. Cinctus of 1Vestiture. Whole body covered with dense decumbent or subdecumbent yellowish hairs, sides of head and mesosoma with fewer hairs; apical funicular segments and legs with standing hairs.Colour. Dark brownish black with mandibles, antennae and legs castaneous.HL 0.73-0.82; HW 0.55-0.60; EL 0.14-0.18; WL 0.81-0.86; PL1 0.27-0.29; MH 0.36-0.41; PrW 0.41-0.44; PW1 0.36-0.39; IIIAL 0.44-0.50; IIIAW 0.51-0.53; SL 0.39-0.42; IVAL 0.75-0.76; IVAW 0.72-0.77. Indices: CI 73-75; SI 70-71; PI 133-134 (n=3).Like the workers of the same colony, but larger, with a more stout body, especially the mesosoma and gaster. Ocelli present on vertex, prominent. The pilosity is much more prominent when compared with the workers. Distinction between ergatoid queens and workers is vague with size variation of workers very high.PageBreakHL 0.77; HW 0.55; EL 0.08; WL 0.93; MH 0.44; PrW 0.41; PL1 0.27; PW1 0.36; IIIAL 0.38; IIIAW 0.49; SL 0.38; IVAL 0.76; IVAW 0.73. Indices: CI 71; SI 69; PI 133 (n=1).Resembles the worker, with modifications expected for caste and the following differences; three prominent ocelli present on vertex, thicker body with heavy pilosity and prominent sculpture.The species epithet is Hindi for border, in reference to its type locality, Manalar, a place which marks border between Kerala and Tamil Nadu.Cerapachys schoedli. However the two species can be easily separated. Cerapachys seema has dull body colouration, sculpture much more prominent and course, pilosity denser, eyes not breaking the lateral margins of head and head almost oval, with anterior and posterior sections of the sides converging, while Cerapachys schoedli is brightly coloured, its sculpture and pilosity are reduced, its eyes break the lateral margins of the head and the head is rectangular with parallel sides.Thespecies is characterized by the punctures on the dorsum of the head being relatively small, separated, with their diameter smaller than the average distance separating them. The new species sharesmost characters with Manalar, part of Periyar tiger reserve, the type locality of this species is a fascinating green hill station (with plenty of leaf litter) surrounded on all sides by the tea gardens of Tamil Nadu. This species was found nesting beneath the marker stone on the border which separates Kerala and Tamil Nadu. It is presumed that the nest was in its initial stages of establishment as there were hardly any galleries and underground chambers. A single queen, 3 ergatoid queens and 7 workers were collected. This species seems uncommon in the Western Ghats range, since it was not encountered again from any other locality.http://zoobank.org/EB5CD657-F22C-4E1B-8352-995242B9531Dhttp://species-id.net/wiki/Cerapachys_wighti11\u00b05'N, 76\u00b026'E, 897m a.s.l., 25.ix.2011, Winkler (coll. Shahid A. Akbar). Holotype in PUAC and paratype in BMNH.Holotype and paratype worker: India. Kerala, Silent valley national park, Measurements (holotype in brackets): HL (0.69)-0.71; HW (0.58)-0.59; EL (0.05); SL (0.38)-0.40; WL (0.66)-0.70; MH (0.43)-0.47; PrW (0.33)-0.35; PL1 (0.28)-0.29; PW1 (0.32)-0.33; IIIAL (0.41)-0.44; IIIAW (0.47)-0.49; IVAL (0.71)-0.72; IVAW (0.66)-0.68 Indices: CI 83-(84); SI (65)-67; PI 113-(114).Head rectangular, longer than broad; sides rounding posteriorly, vertexal margin transverse, posterior lateral corners gently rounded, weakly acute. Parafrontal ridges raised, prominent. Eyes reduced. Mandibles subtriangular; masticatory margin without a row of small denticles. Lateroclypeal teeth reduced. Antennae 12 segmented; scapes short, clavate.PageBreakMesosoma stout, compact, rectangular in dorsal view; dorsal surface slightly convex, the sides gently rounded without any distinct margin. Declivous face of propodeum with the upper sides margined.Petiole broader than long, without strong overhanging dorsolateral margins. Anterior and posterior faces transverse. Subpetiolar process stout with hook like ventral margin; no fenestra present. Postpetiole sub trapezoidal, wider behind, posterolateral angles uniformly rounded. Gaster elongate; base of cinctus of first gastral tergite with cross ribs; sting exerted.st gastral with cross ribs.Sculpture. Mandibles punctured. Punctures on dorsum of head large, crowded, their diameter as large, or larger than, the average distance separating them. Mesosoma, petiole and postpetiole similarly sculptured. Gaster with smaller sized punctures compared with head, mesosoma and metasoma. Cinctus of 1Vestiture. Body with reduced pilosity; moderate decumbent or subdecumbent hairs. Mostly prominent on postpetiole and gaster. Apical funicular segments and legs with standing hairs.Colour. Dark reddish brown with mandibles, antennae and legs lighterThe species is named after botanist Robert Wight, who historically explored the area in 1847.Cerapachys wighti shares most characters with Cerapachys indicus, which also has large crowded punctures on cephalic dorsum. However the two species can be easily separated. Cerapachys wighti is smaller in size (HW 0.59 mm), has lighter body colouration and reduced eyes (EL 0.05 mm), while Cerapachys indicus is larger in size (HW 0.77 mm), with darker body colouration and large eyes (EL 0.24 mm).The new species can easily be separated from most of the Indian species on the basis of the large crowded punctureson its cephalic dorsum, with diameter as large, or larger than, the average distance separating them. PageBreakThe species seems to be of rare occurrence as it was encountered only once during the extensive surveys conducted in the area. It was collected from a litter sample taken near the Kuntipuzha river, which drains the entire length of the silent valley national park. With a pesticide free catchment area the region is rich in soil biota and ideal for cryptic ant species.Cerapachys from India. 12 species are recognized of which 6 are described as new. Partly for convenience the 12 Indian species are placed into arbitrary groups. Group I species with 12 segmented antennae viz., Cerapachys sulcinodis, Cerapachys anokha, Cerapachys schoedli, Cerapachys seema, Cerapachys indicus, Cerapachys aitkenii, Cerapachys wighti, Cerapachys longitarsus and Cerapachys nayana. Of the 9 species given above the first four i.e., Cerapachys sulcinodis, Cerapachys anokha, Cerapachys schoedli and Cerapachys seema, have the punctures on the dorsum of the head relatively small, separated, with their diameter smaller than the average distance separating them. Among these Cerapachys anokha, with the declivous face of the propodeum lacking cariniform margins, and Cerapachys sulcinodis, with the dorsal surface of the petiolar node with a smooth, median area are distinct species in the group. Cerapachys schoedli and Cerapachys seema are easily separated. Cerapachys seema has dull body colouration, sculpture much more prominent and coarse, pilosity denser and head almost oval, with the anterior and posterior sections of its sides converging, while Cerapachys schoedli is brightly coloured, with sculpture and pilosity reduced and the head rectangular with parallel sides. The next 3 species i.e., Cerapachys indicus, Cerapachys aitkenii and Cerapachys wighti, have the punctures on the dorsum of the head large, their diameter greater than the average distance separating them. Among these Cerapachys wighti has the smallest size (HW 0.59 mm) and relatively reduced eyes (EL 0.05 mm) wereas Cerapachys aitkenii and Cerapachys indicus are easily separated from each other on the basis of body sculpture and colouration. Cerapachys aitkenii has characteristic bicolouration and its body sculpture is foveate, wereas Cerapachys indicus is mostly piceous with bluish iridescent sheen and reduced sculpture. The remaining 2 species i.e., Cerapachys longitarsus and Cerapachys nayana are members of \u2018Phyracaces lineage\u2019 and easily recognized, with strong overhanging dorsolateral margins to the petiole. The two species are separated from each other on the basis of body colouration. Cerapachys longitarsus has characteristic bicolouration with head brown, trunk red or brown, petiole and postpetiole light to dark reddish and gaster brown or black, while Cerapachys nayana is uniformly black in colour, with mandibles, antennae and legs castaneous. Group II species have antennae with less than 12 segments viz., Cerapachys biroi, Cerapachys alii and Cerapachys besucheti. Among these Cerapachys besucheti has 11 segmented antennae while Cerapachys biroi and Cerapachys alii have 9 segmented antennae. Cerapachys biroi is characterized by its opaque body with closely spaced piligerous punctures, while Cerapachys alii has prominent foveate body sculpture.Herewe present a review of genus PageBreakCerapachys . Here we present ergatoid queens of three more species - Cerapachys nayana, Cerapachys schoedli and Cerapachys seema. In evaluating morphometric data of the three castes of Cerapachys seema i.e. worker, ergatoid queens and queen castes ) are excluded from this paper. The two species can be easily separates from other reported Indian species. Cerapachys browni shares mostaffinitieswith Cerapachys aitkenii but with black colour (unicolorous), rugo-reticulate sculpture and strongly constricted cintus of gaster. Cerapachys costatus with remarkable costate sculpture, which is not reported in any other Indian species.Two unpublished new species (PageBreak"}
+{"text": "AbstractThrasorinae are revised and Mikeius is transferred to Mikeiinae Paretas-Mart\u00ednez & Pujade-Villar, subfam. n., and Mikeius clavatus Pujade-Villar & Restrepo-Ortiz, sp. n., is described. Two new genera of Thrasorinae are erected: Cicatrix Paretas-Mart\u00ednez, gen. n., including Cicatrix pilosiscutum(Girault), comb. n. from Amblynotus, Cicatrix schauffi (Buffington), comb. n. from Mikeius, and Cicatrix neumannoides Paretas-Mart\u00ednez & Restrepo-Ortiz, sp. n.; and Palmiriella Pujade-Villar & Paretas-Mart\u00ednez, gen. n., including Palmiriella neumanni (Buffington), comb. n. from Mikeius, Thrasorus rieki Paretas-Mart\u00ednez & Pujade-Villar, sp. n., is also described. A phylogenetic analysis of 176 morphological and biological characters, including all these new taxa and all genera previously included in Thrasorinae, was conducted. All subfamilies were recovered as monophyletic, with the following relationships: Parnipinae (Euceroptrinae (Mikeiinae (Plectocynipinae (Thrasorinae)))). A worldwide key to the subfamilies of Figitidae is provided that includes the new subfamily, as well as a key to genera Thrasorinae.The Australian  Figitidae (Hymenoptera: Cynipoidea) are parasitoids of the larvae of other insects, principally cyclorraphous Diptera  on various trees and bushes. They are parasitoids of the gall inducers or other hymenopteran inhabitants in the galls with which they are associated , Mikeius Buffington , Myrtopsen R\u00fcbsaamen (eleven species: two Holarctic and nine Nearctic), and Scutimica Ros-Farr\u00e9 . Thrasorinae are characterized by the circumtorular impression  and the Queensland Museum (QM), as well as the type material of all species included in Mikeius Buffington, new questions arose regarding the taxonomy of Thrasorinae. First, an undescribed species of Mikeius was discovered (described herein); second, two species originally described in Mikeius were determined to render the genus polyphyletic, and new generic assignments are required; and third, phylogenetic analyses determined that the inclusion of Mikeius within Thrasorinae renders the subfamily paraphyletic with respect to Plectocynipinae. In response to these discoveries, Mikeiinae is described as a new subfamily to accommodate Mikeius, and species previously described in Mikeius are moved into other genera. In two cases, no current genus concept could accommodate these species, and the two new genera Cicatrix, gen. n., and Palmiriella, gen. n., are herein described. The goal of this study is to bring clarity to the taxonomic and phylogenetic relationships of these unusual groups of figitid wasps.Following the examination of many undetermined specimens of List of RepositoriesQueensland Museum, Brisbane, Australia (C. Burwell).QMAustralian National Insect Collection, CSIRO, Canberra, Australia .ANICPageBreakSpecimen illustration and observation. Environmental scanning electron micrographs (ESEM) were obtained at Barcelona University with the FEI Quanta 200 ESEM without any coating at 15 KV. Additional ESEM images were obtained either with a Hitachi TM3000 E-SEM, or an Amray 1810 SEM under a vacuum, using a lanthanum hexaboride electron source (LaB6) at 10 Kv, both housed at the National Museum of Natural History, Smithsonian Institution. Images were edited using Adobe CS4 Software . The terminology for morphological structures comes from Phylogenetic analysis. Twenty-two taxa were included in the phylogenetic analysis , so as to capture the morphological diversity of each genus. Parnips nigripes  was chosen as an out-group based on Parnips nigripes as the reference taxon, followed by 1000 bootstrap replicates, each replicate employing 100 TBR swapping replications.analysis , represeParetas-Mart\u00ednez & Pujade-Villarsubfam. n.urn:lsid:zoobank.org:act:9A0F4DEB-C4CE-44E2-BAAC-D86A88DC25CEhttp://species-id.net/wiki/MikeiinaeMikeius Buffington, 2008.Thrasorinae by the absence of a circumtorular impression  that induce galls on species of Acacia (Fabaceae)and Eucalyptus (Myrtaceae), although most of these host records await verification through isolated rearing .Associated with Australia.Mikeius Buffington, 2008.Pujade-Villar & Restrepo-Ortizsp. n.urn:lsid:zoobank.org:act:8D74319A-2A25-48A6-B857-80E6ABB2BE9Chttp://species-id.net/wiki/Mikeius_clavatusMikeius in having the antenna strongly clavate with the six terminal segments 1.5 times wider than previous segments  with the following specific characters.Length. Female 2.8 - 3 mm. Male unknown.Coloration. Head and mesosoma black, antenna yellowish, except scape, brown, metasoma pale brown. Legs pale yellow, except coxae, brown.Antenna. : 4(4): 5(3): 3(3): 3(3): 3(3.5): 4(5): 5(6): 5(6): 6(5): 5(6): 5(6): 7(4). Placoid sensillae from F7 to terminal segment.Antenna.  Female. PageBreakPageBreakMesosoma. Mesoscutum slightly striate. Notauli complete of uniform width. Antero-admedian lines weak. Median mesoscutal line very short. Scutellar foveae round to subquadrate, not delimited posteriorly. Mesopleural furrow absent.Forewing. Radial cell 2.4 times longer than wide.Metasoma. Base of T3 with an almost complete hairy ring.Mikeius clavatus P-V & R-O\u201d (red label). PARATYPE \u2640 (ANIC) with the following labels: \u201cW sidPageBreake Cobungra Hill 20km WbyN, Omeo Vic. 27 Feb. 1980, I.D. Naumann J. C. Cardale\u201d (white label), \u201cex alcohol collection\u201d (white label), \u201cAUST. NAT. INS. COLL.\u201d (green label), \u201cParatype Mikeius clavatus P-V & R-O\u201d (red label).HOLOTYPE \u2640 (ANIC) with the following label data: \u201cAUSTRALIA: Vict. Mt. Donna Buang, 1200m 11\u201317.i. 80, Eucalyptus-Nothofagus forest, A. Newton, M. Thayer\u201d (white label), \u201cflight intercept window/trough trap\u201d (white label), \u201cAUST. NAT. INS. COLL.\u201d (green label), \u201cHolotype Unknown.Victoria, Australia.The specific name refers to the strongly clavate antenna.Kovalev, 1994http://species-id.net/wiki/ThrasorinaeThrasorus Weld, 1944.Euceroptrinae by the absence of an areolet in the forewing and the absence of a lateral pronotal carina. Additional characters that distinguish Thrasorinae from other Figitidae can be found in the key to subfamilies below.Distinguished from other figitids by the presence of a circumtorular impression , 9A, B ; furtherThrasorus, In the redescription of Unknown.Australia, South America and North America.Cicatrix, gen. n.; Myrtopsen R\u00fcbsaamen, 1908; Palmiriella, gen. n., Scutimica Ros-Farr\u00e9, 2007; Thrasorus Weld, 1944.Paretas-Mart\u00ednezgen. n.urn:lsid:zoobank.org:act:F831C129-F846-4A87-A668-524A2EA64E19http://species-id.net/wiki/CicatrixCicatrix pilosiscutum (Girault), comb. n.Cicatrix neumannoides, sp. n., Cicatrix pilosiscutum (Girault), Cicatrix shauffi (Buffington), comb. n.Cicatrix, gen. n., is distinguished from Myrtopsen, Palmiriella, gen. n., and Scutimica by having T3 and T4 as separate sclerites  as well as Cicatrix neumannoides, sp. n., and Cicatrix schauffi (Buffington), comb. n.(Girault)comb. n.http://species-id.net/wiki/Cicatrix_pilosiscutumAmblynotus pilosiscutum Girault, 1929Melanips pilosiscutum (Girault) Weld, 1952Cicatrix neumannoides and Cicatrix schauffi by having female antenna with 11 flagellomeres ), much stronger carinae crossing the entire face .Length. Female 4.4 mm. Male unknown.As in generic description (see above) with the following specific characters: PageBreakColoration. Completely light brown except mesosoma, which is dorsally dark.Head. : 5(3): 10(3): 9(2.5): 8.5(2.5): 8(3): 8(3): 8(3): 7(3): 5(3): 5(3): 4.5(3): 7.5(3). Placoid sensillae absent on basal half of F1 to F4, scarce on dorsal half; abundant from F5 to F11.Mesosoma. Median mesoscutal impression absent. Scutellar foveae subtriangular.Amblynotus pilosiscutum \u2640, Type Girault\u201d , \u201cXyalophoroides pilosiscutum (Gir), E. F. Riek det 1953\u201d , QM Reg. No. T99348\u201d (yellow label), \u201cCicatrix pilosiscutum P-M det-2009\u201d (white label).HOLOTYPE \u2640 (QM) with the following labels: \u201c25. 10. 23, National Pk., Q. H. Hacker.\u201d (white label), \u201cHOLOTYPE\u201d (pink label), \u201cUnknown.Australia. Label data suggest the single specimen was taken in Royal National Park in Sydney.(Buffington)comb. n.http://species-id.net/wiki/Cicatrix_schauffiMikeius schauffi Buffington, 2008.Cicatrix neumannoides, sp. n., in having female antenna with 10 flagellomeres , but differs from Cicatrix neumannoides sp. n. by having a long median mesoscutal impression and subtriangular scutellar foveae  with the following specific characters: Coloration. Completely light brown.Head. : 4(3): 5(3): 4(3): 4(3): 4.2(3.1): 4.2(3.1): 4.3(3.3): 5.2(3.3): 4.6(3.3): 3.5(3.3): 6(4). Placoid sensillae present from F4, abundant from F6 through terminal segment.Antenna.  Female. Mesosoma.  with the following label data: \u201c23.36S 133.35E 32 km WNW of Alice Springs, NT 8 Oct. 1978 J:C: Cardale\u201d (white label), \u201cex alcohol collection\u201d (white label), \u201cAUST. NAT. INS. COLL.\u201d (green label). \u201cHOLOTYPE, Mikeiusschauffi, Buffington\u201d (red label), \u201cCicatrix schauffi P-M det-2009\u201d (white label).PageBreakUnknownCentral Australia.Thrasorinae, not in Mikeiinae. We transfer this species to Cicatrix gen. n., because it possesses all the diagnostic characters of that genus.The circumtorular impression present in this species indicates that it belongs in Paretas-Mart\u00ednez & Restrepo-Ortizsp. n.urn:lsid:zoobank.org:act:1A6D286B-94AF-43F8-945B-D9AF124EEC57http://species-id.net/wiki/Cicatrix_neumannoidesCicatrix schauffi, comb. n., having female antenna with 10 flagellomeres and a face with horizontal strigae only on the lateral areas , but differs from Cicatrix schauffi comb. n. by having short median mesoscutal impression and rounded scutellar foveae (Similar to r foveae .As in generic description (see above) with the following specific characters.Length. Female: 2.9 to 3.0 mm. Male unknown.Coloration. Shiny chestnut, scutum darker in center.Head. Frons and face with piliferous punctures; face with a few carinae from internal margin of eye reaching center of face.Antenna. Female. 10 flagellomeres, antennal formula: 6(2): 4(2.8): 6(2.5): 4.1(2.8): 4.1(2.8): 4(3): 4(3): 4(3): 4.8(3.1): 3.8(3.3): 3.5(3.3): 5.6(4). Placoid sensillae starting from F4, F4 to F6 are scarce, abundant from F7-F10.Mesosoma. , \u201cAUST. NAT. INS. COLL.\u201d (green label), \u201cMikeius neumanni Det. M. L. Cicatrix neumannoides P-M & R-O\u201d (red label)\u201d. PARATYPE \u2640 (ANIC) with the following labels: \u201cCrowea St. For. nr Pemberton W.A. Nov.-Dec. 1978 S.J. Curry Malaise trap open forest\u201d (white label), \u201cAUST. NAT. INS. COLL.\u201d (green label), \u201cParatype Cicatrix neumannoides P-M & R-O\u201d (red label)\u201d.HOLOTYPE \u2640 (ANIC) with the following labels: \u201cAUSTRALIA: NSW Peak Hill Range, Braidwood, Cooma Road, At top of pass. 30 December 1994. A. Sundholm & R de keyzer. On Acacia.Unknown; label data suggests an association with New South Wales and Western Australia, Australia.Mikeius neumanni in the collection at ANIC, he used only one specimen in his description of the taxon, designating it as the holotype. The species neumanni (based on PageBreakthe holotype) is transferred to Palmiriella, gen. n., below, and the second specimen, in addition to another specimen discovered in ANIC, belongs to Cicatrix.Although Pujade-Villar & Paretas-Mart\u00ednezgen. n.urn:lsid:zoobank.org:act:5F540007-4494-49BF-A619-F0A54F5CE41Ehttp://species-id.net/wiki/PalmiriellaPalmiriella neumanni (Buffington), comb. n., by present designation and monotypy.Palmiriella, gen. n., can be distinguished from other thrasorines by having the face smooth, without any sculpturing   , 4F. Addcutimica , with miyrtopsen ); from Tsoscutum  comb. n.http://species-id.net/wiki/Palmiriella_neumanniMikeius neumanni Buffington, 2008.Length. Female 3.2 mm. Male unknown.PageBreakColoration. Head and mesosoma black, antennae yellowish except scape, brown, metasoma medium brown. Legs light yellow except tibia and metatarsi, brown.Head : 4(4): 4(3): 4.5(3): 4.5(3): 4.5(3): 4(3): 4(3): 4(3): 3(3): 3(3): 3(3): 5(4). Placoid sensillae from F7 to terminal segment.Antenna . Female.Mesosoma , \u201cPalmiriella neumanni P-V & P-M det-2009\u201d (white label).HOLOTYPE \u2640 (ANIC) with the following labels: \u201cMt Nebo, S. E. Qld, 24. Xi. 1970, S. R. Monteith\u201d (white label), \u201cAUST. NAT. INS. COLL.\u201d (green label). \u201cHOLOTYPE, Unknown.Queensland, Australia.Weld, 1944Thrasorus pilosus Weld, 1944.Thrasorus pilosus Weld, Thrasorus rieki, sp. n., Thrasorus schmitdae Buffington.Paretas-Mart\u00ednez & Pujade-Villarsp. n.urn:lsid:zoobank.org:act:BCD3677F-EA0D-4D37-B62B-F093CEDF7B02http://species-id.net/wiki/Thrasorus_riekiThrasorus by having small scutellar foveae not clearly defined in posterior margin . PARATYPES: 4 \u2642 and 1 \u2640 (on the same pinned card as the holotype) with the same data as the holotype, \u201cParatype Thrasorus rieki P-M & P-V det-2009\u201d (red label); 1 \u2642 and 5 \u2640 (ANIC)  with the following labels: \u201cOut of Acacia galls ???? 19.1.16 QLD\u201d (handwritten below the label with the insects), \u201cAUST. NAT. INS. COLL.\u201d (green label), \u201cParatype Thrasorus rieki P-M & P-V det-2009\u201d (red label); 1 \u2640 (QM) with the following labels: \u201cAmblynotus berlesei \u2640 Girault types\u201d (white label handwritten), PageBreak\u201cHOLOTYPE\u201d (pink label), \u201cThrasorus berlesei (Gir) EF Riek det 1953\u201d (white label handwritten), \u201cQM reg. No. T99347\u201d (yellow label), \u201cParatype Thrasorus rieki P-M & P-V det-2009\u201d (red label).HOLOTYPE \u2640  with the following labels: \u201cOut of large galls on mullee acacia On 18\u20131-16\u201d (handwritten below the label with the insects), \u201cThrasorus berlesei (Grlt) Riek det\u201d , \u201csp 7 (berlesei) det ML Buffington 2006\u201d (white label), \u201cHolotype Acacia galls (based on label data).PageBreakUnknown host on Australia, Queensland.Cynipoidea.Named after E.F. Riek, who worked before us on Australian Amblynotus berlesei\u2019 by Girault. In ANIC, there are six specimens on one large card with a determination label placed by Riek, stating that taxon is \u2018Thrasorus berlesei (Grlt)\u2019. But as nomen nudum after Thrasorus rieki, sp. n., mixed with Chalcidoidea specimens.In the QM, there is one specimen labelled as \u2018Thrasorinae: the circumtorular impression . The portion of the plate that is reduced is the posterior part of the pronotal plate, or the portion of the plate dorsal to the submedial pronotal depressions. The arrow in Palmiriella , and T3-T4 fused into a syntergum . The primary difference between Thrasorus and Cicatrix is the sculpturing of the mesoscutum. Though the sculpturing on the mesoscutum can be variable in other groups of Figitidae, in the \u2018pool\u2019 of genera treated in this paper, mesoscutal sculpture is useful and unique character. Thrasorus is the only genus, not only among Thrasorinae but also among all the genera previously included in this subfamily , that aside from notauli, lacks sculpturing of any kind  in the mesoscutum; we believe that this character is enough to justify the separation of Thrasorus from Cicatrix and the other thrasorines.The morphology of the metasoma is a very important character and is frequently used in all Palmiriella from Scutimica and Myrtopsen are detailed in the diagnosis of the genus (see above). The combination of the smooth face , shape of syntergum T3-T4, shape of scutellum, shape of pronotum, and absence of sculpturing on pronotum, distinguish Palmiriella from Scutimica and Myrtopsen. The differences between Scutimica and Myrtopsen have already been remarked and discussed in The characters that differentiate Parnipinae (Euceroptrinae (Mikeiinae (Plectocynipinae (Thrasorinae)))). It is clear that Mikeius renders the Thrasorinae paraphyletic, supporting the description of Mikeiinae, and that Cicatrix and Palmiriella are distinct clades. Erecting a new subfamily for a single genus is not desirable, but the only alternative to this while respecting the clades recovered in the phylogenetic analysis would be grouping together Mikeius, Palmiriella, Thrasorus, Cicatrix, Scutimica, Myrtopsen, Plectocynips and Pegascynips in a single subfamily; we feel this grouping is undesirable from the standpoint of predictability, since these genera contain species possessing markedly different biological and morphological attributes, and still would lack a single common diagnostic character for all of them. As currently defined, each of the subfamilies recognized here has its own diagnostic character: long metatibial spur for Plectocynipinae, circumtorular impression for Thrasorinae and two carinae in median area of pronotum not forming a projected pronotal plate for Mikeiinae.PageBreakThe results of the phylogenetic analysis are summarized in Thrasorinae from Australia are one of the most poorly known groups of figitids. More field data and specimens would help to clarify the status of this group and some taxa described here. However, there is no single researcher in Australia dedicated to the study of Cynipoidea, and workers on Figitidae wanting to study the systematics PageBreakof this group must rely on \u2018rare\u2019 specimens coming from non-target collections while pursuing the sampling of other groups. The study we present here has been done with all the thrasorines and Mikeius that have been collected, curated, and deposited in museums worldwide.PageBreakThe"}
+{"text": "Over the last decade implementation of targeted therapy has improved the functional ability in JIA-children. Also, the attitude towards participation of JIA-children in sport has become less restrictive with growing evidence of the benefits of physical activity on e.g. disease, fitness and quality of life.To explore the habits in gym classes and sport in 10-16 year-old JIA-children compared to gender- and age-matched healthy controls.68 JIA-children , age 12.74 (\u00b11.70) years, disease duration 5.72 (\u00b14.17) years, and 118 healthy controls , age 12.36 (\u00b11.74) years, answered questionnaires on sport and exercise habits and functional ability (CHAQ38). For JIA: VAS-pain, Patient- and Physician-GA were noted.Mean CHAQ38 scores for JIA: 0.1896 (\u00b10.2025), controls: 0.0359 (\u00b10.0799) indicated only minimal functional impairment in both groups. Pain assessments correlated well to CHAQ38 scores in JIA .No significant differences were found between sport-active JIA-children and controls concerning type and number of sport activities or amount of time spent in sport. Significantly more JIA-children were not sport-active (38% (26/68) vs. 25% (29/118)), mainly due to joint pain (12/26) and getting short of breath/side-stitches (9/26).Significantly fewer JIA-children participated fully in gym (p<0.01) and reported pain (91% (62/68)) and difficulties with certain activities (p<0.01). Strategies mainly used when pain; a short break (81%), changing activity (58%), continuing despite pain (37%).JIA-children still participate less in sport activities and are more challenged in gym classes than healthy peers, despite close-to-normal functional ability. However, sport-active JIA-children do not differ significantly from sport-active healthy peers."}
+{"text": "Levels of brain natriuretic peptide (BNP) increase following CABG and predict post-operative outcomes. Release kinetics of BNP, Troponin-I (TnI) and CKMB after valve replacement are not well characterized.We assessed levels of these biomarkers 24 hours prior and 6,24, 48 hrs, 1 month following mitral/aortic valve replacement in 50 patients .Mean baseline BNP, TnI and CK-MB levels were 304.01 pg/ml, 0.03 ng/ml and 0.99 ng/ml. BNP initially decreased within 6 hours of surgery, and peaked at 24 hours; TnI and CKMB showed an early rise, with declining trends by 24 hrs. Peak BNP levels occurred in 90% patients by 24-48 hrs, while for TnI and CKMB this occurred in only 15-30%.Mean delta (peak-baseline) BNP, TnI, CKMB was 660.1pg/ml, 8.1ng/ml and 32.3ng/ml.At 1 month, levels of all biomarkers were not significantly different from baseline. Patients with higher baseline BNP more commonly had atrial fibrillation , higher right ventricular systolic pressure ,higher Euroscore II,longer inotrope duration , ventilator support time , longer ICU  and hospital stay . Inotrope duration>42 hrs, ventilation time>29 hrs and ICU stay>4 days was seen in 42%vs19%, 30%vs9% and 33%vs14% respectively in patients with baseline BNP >/< 200 pg/ml. Only baseline BNP was a significant predictor of inotrope duration (p=0.01) and ventilation time (p=0.02). Only 24 hour post-operative BNP and delta BNP were predictors of inotrope duration>42 hrs, ventilation time>29 hrs and ICU stay>4 days.Release kinetics of cardiac biomarkers following valve surgery are significantly different from each other. Of all the biomarkers, only BNP levels had an association with post-operative inotrope duration, ventilation time and ICU stay in patients undergoing valve replacement."}
+{"text": "Cell volume can be calculated as 1-DMF%*LV mass.In severe aortic stenosis (AS), hemodynamics and conventional indices do not fully explain symptoms, prognosis or treatment response. We hypothesize that diffuse myocardial fibrosis (DMF) is a key missing factor in AS. This can now be accurately measured non-invasively using equilibrium contrast CMR (EQ-CMR)  involvin(m)/DMF% (continuous variable or severity tertiles), or cell volume.63 severe AS patients with planned valve replacement underwent baseline and follow up EQ-CMR. Twenty normal controls were included. Baseline and follow-up assessment included NYHA, ECG, echocardiography , BNP and six minute walk test (6MWT). Follow up was at 6-months . EQ-CMR results were expressed as Vd(m):0.27\u00b10.04 vs 0.24\u00b10.04; DMF:17% vs 11%, p = 0.003) with a wide range (Vd(m):0.20-0.39; DMF:4-42%). Breathless patients had more DMF (NYHA class III/IV vs I/II: Vd(m):0.32\u00b10.03 vs 0.26\u00b10.04; DMF:15% vs 27%, p<0.001). DMF correlated with 6MWT  and aortic valve area . DMF only correlated with EF in patients with LV impairment . Severe DMF patients had worse diastolic function (p=0.029). In a multivariate analysis of all parameters classically associated with 6MWT distance, the only independent predictor was DMF (p=0.04). On univariate analysis there was a weak correlation with BNP and age.AS patients had more fibrosis than controls . However, only patients with severe fibrosis improved their exercise capacity (p=0.03). LVH regression  was shown to be cellular  rather than fibrosis (36g vs 34g p=0.572) resolution, figure In this first clinical EQ-CMR study of severe AS, DMF is higher when there is LV impairment, diastolic dysfunction and more severe stenosis. DMF is the single best predictor of pre-op exercise capacity and post-op improvement. EQ-CMR shows that at 6-month post valve replacement LVH regression is predominantly reduced cell rather than fibrosis volume. EQ-CMR for the non-invasive measurement of DMF appears to be a significant cardiological advance."}
+{"text": "One of the causes of septic mortality is a low cardiac output secondary to preload failure. Same patients demonstrate preload failure after aggressive volume replacement .t criteria.Ultrasound impulse-wave Doppler evaluation of transmitral flow: VmaxE, VmaxA, ejection time E,A; DT E wave, IVRT of LV. Ultrasound evaluation of end-diastolic and end-systolic LV volume, stroke volume  on Teichholz L. EDLVP = 1.06 + 15.15 \u00d7 VTI peakA/VTI peakE. Coronary perfusion pressure (CPP) = EDLVP - diastolic BP. We evaluate these parameters in 34 patients (age 28.1 \u00b1 8.0 months) with septic shock (SS) diagnosed according to Consensus 2002. Control (C) - 44 healthy children (age 40.7 \u00b1 8.5 months). Statistical analyses with The increase of VmaxA and decrease of VmaxE in patients of SS are demonstrated. IVRT and DT are less than in control group. We evaluated a decrease in E/A proportion. EDLVP in patients was more, and CPP lower, than in controls. See Table Pediatric SS accompanied with LV diastolic dysfunction, which decreases the effectiveness of volume restoration therapy, reduces preload and cardiac output."}
+{"text": "AbstractTetramorium tortuosum species group members encountered in the Afrotropical region, which we have placed in its own subgroup: the Tetramorium capillosum species complex. We re-describe the two previously known species Tetramorium capillosum Bolton and Tetramorium tabarum Bolton, and describe the new species Tetramorium hecatesp. n. The geographic distribution of the three species appears to be restricted to the equatorial rainforests of Central Africa. We provide a diagnosis of the Tetramorium capillosum species complex, an illustrated identification key to species level, and worker-based species descriptions, which include diagnoses, discussions, high-quality montage images, and distribution maps. Furthermore, we discuss biogeography and composition of the globally distributed Tetramorium tortuosum group.In this study we revise the taxonomy of the  Tetramorium Mayr represents one of the most species-rich ant genera, a group that is also widely distributed throughout most zoogeographical regions. However, in terms of species and species group numbers, the main diversity of the genus is found in the Afrotropical and Malagasy regions, from where around 220 Afrotropical and 84 Malagasy species are currently known , London, U.K.CASC California Academy of Sciences, San Francisco, California, U.S.A.MCZ Museum of Comparative Zoology, Cambridge, Massachusetts, U.S.A.MHNG Mus\u00e9um d\u2019Histoire Naturelle de la Ville de Gen\u00e8ve, Geneva, SwitzerlandNHMB Naturhistorisches Museum, Basel, SwitzerlandTetramorium capillosum and Tetramorium tabarum, together with a small amount of non-type specimens, are found in BMNH and MHNG.The material examined in this study is located in the collections of BMNH, CASC, and MCZ. More than 95% of the specimens examined belong to CASC and were sampled during ant inventories carried out in Central Africa from 1998 to 2001 . All images presented are available online and can be seen on AntWeb (http://www.antweb.org). The measurements were taken with a Leica MZ 12.5 equipped with an orthogonal pair of micrometers at a magnification of 100\u00d7, rarely 80\u00d7. Measurements and indices are presented as minimum and maximum values with arithmetic means in parentheses. In addition, all measurements are expressed in mm to two decimal places. The measurements and indices used in this study are the same as in All new type material and all imaged specimens can be uniquely identified with specimen-level codes affixed to each pin (e.g. CASENT0078328). In the presented descriptions we list all of the available specimen-level codes for the whole type series. It should be Head length (HL): maximum distance from the mid-point of the anterior clypeal margin to the mid-point of the posterior margin of head, measured in full-face view. Impressions on anterior clypeal margin and posterior head margin reduce head length.Head width (HW): width of head directly behind the eyes measured in full-face view.Scape length (SL): maximum scape length excluding basal condyle and neck.Eye length (EL): maximum diameter of compound eye measured in oblique lateral view.Pronotal width (PW): maximum width of pronotum measured in dorsal view.Weber\u2019s length (WL): diagonal length of mesosoma in lateral view from the postero-ventral margin of propodeal lobe to the anterior-most point of pronotal slope, excluding the neck.Propodeal spine length (PSL): the tip of the measured spine, its base, and the centre of the propodeal concavity between the spines must all be in focus. Using a dual-axis micrometer the spine length is measured from the tip of the spine to a virtual point at its base where the spine axis meets orthogonally with a line leading to the median point of the concavity.Petiolar node height (PTH): maximum height of petiolar node measured in lateral view from the highest (median) point of the node to the ventral outline. The measuring line is placed at an orthogonal angle to the ventral outline of the node.Petiolar node length (PTL): maximum length of the dorsal face of the petiolar node from the anterodorsal to the posterodorsal angle, measured in dorsal view excluding the peduncle.Petiolar node width (PTW): maximum width of dorsal face of petiolar node measured in dorsal view.Postpetiole height (PPH): maximum height of the postpetiole measured in lateral view from the highest (median) point of the node to the ventral outline. The measuring line is placed at an orthogonal angle to the ventral outline of the node.Postpetiole length (PPL): maximum length of postpetiole measured in dorsal view.Postpetiole width (PPW): maximum width of postpetiole measured in dorsal view.Ocular index (OI): EL / HW * 100Cephalic index (CI): HW / HL * 100Scape index (SI): SL / HW * 100Propodeal spine index (PSLI): PSL / HL * 100PageBreakPetiolar node index (PeNI): PTW / PW * 100Lateral petiole index (LPeI): PTL / PTH * 100Dorsal petiole index (DPeI): PTW / PTL * 100Postpetiolar node index (PpNI): PPW / PW * 100Lateral postpetiole index (LPpI): PPL / PPH * 100Dorsal postpetiole index (DPpI): PPW / PPL * 100Postpetiole index (PPI): PPW / PTW * 100Tetramorium ; anterior clypeal margin usually entire without median notch; frontal carinae very well developed and usually reaching posterior head margin; antennal scrobe present, weakly to very well developed; propodeal spines medium-sized to long, elongate-triangular to spinose; propodeal lobes short, triangular to elongate-triangular; petiolar node in profile nodiform, in profile as high as long to 1.3 times higher than long (LPeI 78 - 100), in dorsal view always longer than wide (DPeI 80 - 93); postpetiole sub-globular to moderately anteroposteriorly compressed; mandibular sculpture variable; cephalic sculpturation distinct, between frontal carinae longitudinally rugose to reticulate-rugose; mesosoma predominantly longitudinally rugose; petiolar node weakly to distinctly rugose, postpetiole ranging from unsculptured to longitudinally rugose; gaster unsculptured, smooth, and shiny; all dorsal surfaces of body with abundant, long, standing hairs; first gastral tergite without pubescence, and pilosity never short, dense, and appressed; sting appendage spatulate.Taxonomic notesPageBreakTetramorium tortuosum group are unlikely to be misidentified with species from the other three groups having 11-segmented antennae. The 26 species of the Tetramorium weitzeckeri group all have a squamiform or high nodiform petiolar node, which is always significantly wider than long. This node shape strongly contrasts with the shape observed in the Tetramorium tortuosum group since all three members have a nodiform node which is much longer than wide. The second species group, the Tetramorium angulinode group, is morphologically closer to the Tetramorium tortuosum group since both groups share a nodiform petiolar node. However, they can be clearly separated by the pilosity/pubescence patterns on the first gastral tergite. In the Tetramorium tortuosum group pubescence is absent and pilosity is long and mainly erect, whereas in the Tetramorium angulinode group pubescence and pilosity are usually present, dense, appressed to decumbent, and often pointed towards a longitudinal midline of the tergite. The synonymisation of Triglyphothrix under Tetramorium , and it seems appropriate to evaluate whether the Afrotropical species fit into one of these groups or deserve their own species complex. Based on the definitions of the complexes, the Afrotropical species cannot be members of the Tetramorium jedi, Tetramorium noeli, or Tetramorium smaug complexes due to a lack of sculpture on the forecoxae and the first gastral tergite. This would argue for placement in the Tetramorium andrei complex. Two reasons prevented us from doing so however. First, there is a difference in the development of the anterior clypeal margin, which is strongly medially impressed in the Malagasy Tetramorium andrei complex while the Afrotropical species treated in this study usually have an entire margin . The second reason not to place the African species into a Malagasy complex is the current uncertainty about whether the Tetramorium tortuosum group as a whole is a natural group of closely related species, an issue discussed below. Consequently, we propose to place Tetramorium capillosum, Tetramorium hecate, and Tetramorium tabarum in their own complex, the Tetramorium capillosum species complex.Biogeographic notes on the groupPageBreakTetramorium tortuosum group have a moderately restricted distribution range since they are only known from Equatorial rainforests in the Central African countries of Gabon, Cameroon, Democratic Republic of Congo, Central African Republic, and Uganda. Given all of the known African localities, the distribution of Tetramorium capillosum and Tetramorium tabarum appears fairly disjunctive. Most localities are located in the west of the distribution range in Gabon, Cameroon, and western parts of the Central African Republic, but few localities are found much further east in the northeastern Democratic Republic of Congo and northwestern Uganda. This represents a great gap between these two groups of localities. However, we think that this lack of occurrence records is very likely due to a sampling artefact since ant sampling has been relatively fragmentary in sub-Saharan Africa. The westernmost known distribution limit is the eastern coast of the Gulf of Guinea and the easternmost known locality appears to be the Budongo Forest in northwestern Uganda. It is unlikely that they occur further east, which is supported by an inventory of the myrmecofauna of the Kakamega Forest in Western Kenya , Tetramorium philippwagneri Hita Garcia, Fischer and Peters, and Tetramorium pinnipilum Bolton, are fairly successful, common, and relatively abundant species found in many equatorial rainforests. Also, the Tetramorium weitzeckeri group, with the exception of Tetramorium humbloti Forel that has invaded the Malagasy region, is restricted in distribution to the Afrotropical region. This fact indicates that it represents a relatively young and successful development within the Afrotropical Tetramorium fauna, supporting Tetramorium weitzeckeri group, there are many more Afrotropical species groups with 12-segmented antennae that perform well in rainforests, such as the Tetramorium bicarinatum, Tetramorium camerunense, and Tetramorium flabellum groups. This very strong competition in most modern-day rainforests might well explain the depauperate Tetramorium tortuosum fauna encountered in Africa. The species-rich Malagasy group fauna does not face the same competition by other genus members within their size and niche ranges, and with 22 species seems to have undergone a fairly successful radiation, mostly in the rainforests of eastern Madagascar  [MCZ] [examined]. Paratypes, seven pinned workers with same data as holotype  [examined].Holotype, pinned worker, GABON, Makokou, Non-type material. CAMEROON: Mbalmayo, XI.1993 (PageBreakN. Storck); Ndupe, 20.XII.1989 (A. Dejean); Sud, Bond\u00e9 Forest, N\u2019kolo village, 27.5 km 155\u00b0 SSE Elogbatindi, 3.2217N, 10.2467E, 40 m, rainforest, 12.IV.2000 (B.L. Fisher); Sud, Campo Reserve, 2\u00b036'N, 9\u00b056'E, 40 m, 25.X.1991 (D.M. Olson); Sud, P.N. Campo, 43.3 km 108\u00b0ESE Campo, 2.2825N, 10.2062E, 290 m, rainforest, 7.IV.2000 (B.L. Fisher); Sud, Res. de Faune de Campo, Massif des Mamelles, 15.1 km 84\u00b0E \u00c9bodj\u00e9, 2.59417N, 9.9595E, 180 m, rainforest, 4.IV.2000 (B.L. Fisher); CENTRAL AFRICAN REPUBLIC: Prefecture Sangha-Mba\u00e9r\u00e9, R\u00e9serve Sp\u00e9ciale de For\u00eat Dense de Dzanga-Sangha, 12.7 km 326\u00b0NW Bayanga, 3.005N, 16.1933E, 420 m, rainforest, 10.\u201317.V.2001 (B.L. Fisher); Prefecture Sangha-Mba\u00e9r\u00e9, Parc National Dzanga-Ndoki, Mab\u00e9a Bai, 21.4 km 53\u00b0NE Bayanga, 3.0333N, 16.41E, 510 m, rainforest, 1.\u20137.V.2001 (B.L. Fisher); DEMOCRATIC REPUBLIC OF CONGO: Epulu, 1.3833N, 28.5833E, 750 m, rainforest, 1.XI.1995 (S.D. Torti); GABON: La Makande, Foret de Abeilles, I.-II.1999 (S. Lewis); Makokou, rainforest, X.1972 (I. Lieberburg); Ogooue-Maritime, Aire d\u2019Exploit. Rationnelle de Faune des Monts Doudou, 24.3 km 307\u00b0NW Doussala, 2.2264N, 10.4097E, 375 m, rainforest, 6.-9.III.2000 (B.L. Fisher); Ogooue-Maritime, Aire d\u2019Exploit. Rationnelle de Faune des Monts Doudou, 25.2 km 304\u00b0NW Doussala, 2.2275S, 10.3945E, 640 m, rainforest, 14.III.2000 (B.L. Fisher); Ogooue-Maritime, Reserve des Monts Doudou, 25.2 km 304\u00b0NW Doussala, 2.2272S, 10.3945E, 630 m, coastal lowland rainforest, 13.-20.III.2000 (S. van Noort); Ogooue-Maritime, Reserve de la Moukalaba-Dougoua, 12.2 km 305\u00b0NW Doussala, 2.3167S, 10.5333E, 110 m, rainforest, 24.II.2000 (B.L. Fisher); Ogooue-Maritime, Reserve de la Moukalaba-Dougoua, 10.8 km 214\u00b0SW Doussala, 2.4227S, 10.5453E, 110 m, rainforest, 29.II.2000 (B.L. Fisher); Ogooue-Maritime, Reserve de la Moukalaba-Dougoua, 12.2 km 305\u00b0NW Doussala, 2.2833S, 10.4972E, 110 m, coastal lowland rainforest, 24.II.\u20133.III.2000 (S. van Noort); Woleu-Ntem, 31.3 km 108\u00b0ESE Minvoul, 2.08N, 12.4067E, 600 m, rainforest, 7.II.1998 (B.L. Fisher); UGANDA: Bunyoro District, Budongo Forest FS, 1.7264N, 31.5524E, 1081 m, 8.VII.2009 (W. Freund & T. Klug).Tetramorium capillosum from the remainder of the species group: eyes of moderate size (OI 23 - 25); antennal scapes moderately long (SI 80 - 83); petiolar node nodiform with anterodorsal and posterodorsal margins relatively rounded, posterodorsal margin situated higher than anterodorsal margin, dorsum convex; mandibles strongly longitudinally rugose; petiole and postpetiole usually with weak sculpture; whole body uniformly very dark brown to black.The following character combination clearly distinguishes (N=12). HL 0.79 - 0.89 (0.84); HW 0.76 - 0.84 (0.79); SL 0.62 - 0.69 (0.65); EL 0.18 - 0.21 (0.19); PH 0.41 - 0.51 (0.46); PW 0.60 - 0.68 (0.65); WL 1.02 - 1.19 (1.12); PSL 0.26 - 0.38 (0.30); PTL 0.31 - 0.37 (0.34); PTH 0.34 - 0.41 (0.37); PTW 0.28 - 0.33 (0.31); PPL 0.28 - 0.33 (0.30); PPH 0.37 - 0.43 (0.40); PPW 0.37 - 0.44 (0.41); CI 94 - 96 (95); SI 80 - 83 (82); OI 23 - 25 (24); DMI 55 - 62 (58); LMI 39 - 43 (41); PSLI 31 - 43 (35); PeNI 45 - 49 (47); LPeI 89 - 100 (94); DPeI 86 - 94 (89); PpNI 61 - 66 (63); LPpI 70 - 78 (75); DPpI 130 - 139 (135); PPI 127 - 138 (134).PageBreakPageBreak (DPeI 86 - 94). Postpetiole in profile relatively high and moderately anteroposteriorly compressed, approximately 1.2 to 1.3 times higher than long (LPpI 70 - 78); in dorsal view around 1.3 to 1.4 times wider than long (DPpI 130 - 139). Postpetiole in profile thinner and higher than petiolar node, in dorsal view approximately 1.3 to 1.4 times wider than petiolar node (PPI 127 - 138). Mandibles strongly longitudinally rugose; clypeus longitudinally rugulose, usually with three to five rugulae, median rugula better developed; cephalic dorsum between frontal carinae irregularly longitudinally rugose to reticulate rugose, posteriorly towards posterior head margin fully reticulate-rugose, anteriorly towards posterior clypeal margin more regularly longitudinally rugose; scrobal area unsculptured, smooth, and shining; lateral and ventral head longitudinally rugose to reticulate-rugose. Mesosoma laterally and dorsally strongly irregularly longitudinally rugose. Forecoxae unsculptured, smooth, and shining. Both waist segments with strong sculpture, mainly longitudinally rugose. Gaster unsculptured, smooth, and shining. Ground sculpture generally faint to absent everywhere on body. Whole body with abundant, very long, and fine standing hairs; first gastral tergite without appressed pubescence. Anterior edges of antennal scapes with suberect to erect hairs. Body of uniform very dark brown to black colour.Head longer than wide (CI 94 - 96); posterior head margin moderately concave. Anterior clypeal margin entire and convex. Frontal carinae strongly developed, approaching or ending at posterior head margin. Antennal scrobes narrow but very well-developed with clearly defined margins all around. Antennal scapes moderately long, not reaching posterior head margin (SI 80 - 83). Eyes of moderate size (OI 23 - 25). Mesosomal outline in profile weakly convex, moderately marginate from lateral to dorsal mesosoma; promesonotal suture and metanotal groove absent; mesosoma comparatively stout and high (LMI 39 - 43). Propodeal spines relatively long to very long, spinose, and acute (PSLI 31 - 43); propodeal lobes short, triangular to elongate-triangular, and usually acute. Petiolar node in profile rectangular nodiform, approximately as high as long to weakly higher than long (LPeI 89 - 100), anterior and posterior faces approximately parallel, posterodorsal margin situated higher than anterodorsal, anterodorsal and posterodorsal angles relatively rounded, petiolar dorsum convex; node in dorsal view approximately 1.1 times longer than wide Tetramorium capillosum is found throughout Gabon, southern Cameroon, eastern Central African Republic, and then much further east in the northeastern Democratic Republic of Congo and northwestern Uganda  either bicoloured or of uniform brown colour, whereas Tetramorium capillosum is noticeably larger  and of a very dark brown to black colour. However, both body size and colouration are often variable within the genus Tetramorium. Tetramorium capillosum also differs from Tetramorium hecate in antennal scape length, eye size, and petiolar node shape. In Tetramorium capillosum the antennal scapes are moderately long (SI 80 - 83), eyes are of moderate size (OI 24 - 25), and the petiolar node has anterodorsal and posterodorsal margins that are relatively rounded, with the posterodorsal margin situated higher than the anterodorsal. In contrast, Tetramorium hecate scapes are relatively short (SI 73 - 77), eyes are relatively large (OI 27 - 31), and the petiolar node is rectangular nodiform with anterodorsal and posterodorsal angles sharply defined and at about the same height. Furthermore, Tetramorium tabarum significantly varies from Tetramorium capillosum in eye size, propodeal spine length, and sculpture on mandibles and postpetiole. Tetramorium tabarum has much larger eyes (OI 27 - 31), much shorter spines (PSLI 22 - 25), and the mandibles and the postpetiole are unsculptured, whereas Tetramorium capillosum has smaller eyes (OI 24 - 25), much longer spines (PSLI 31 - 43), and conspicuously sculptured mandibles and postpetiole.Within the African Tetramorium capillosum remains morphologically remarkably stable without noticeable intraspecific variation.It should be noted that despite a relatively broad distribution range from the Gulf of Guinea to northwest Uganda, Hita Garcia & Fishersp. n.urn:lsid:zoobank.org:act:BEEEF558-1DDA-40AB-8D79-597684B38033http://species-id.net/wiki/Tetramorium_hecateHolotype, pinned worker, GABON, Province Estuaire, F.C. Mondah, 21 km 331\u00b0NNW Libreville, 0\u00b034.6'N, 9\u00b020.1'E, 10 m, littoral rainforest, sifted litter , collection code BLF01742, 24.II.1998 (B.L. Fisher) [unique specimen identifier CASENT0248334] [CASC]. Paratypes, 16 pinned workers with same data as holotype .2\u00b036'N, 9\u00b056'E, 40 m, 25.X.1991 (D.M. Olson); Mbalmayo, XI.1993 (N. Storck); Nkoemvon, 16.III.1980 (D. Jackson); Sud, P.N. Campo, 43.3 km 108\u00b0ESE Campo, 2.2825N, 10.2062E, 290 m, rainforest, 7.IV.2000 (B.L. Fisher); Sud, Res. de Faune de Campo, 2.16 km 106\u00b0ESE \u00c9bodj\u00e9, 2.5678N, 9.8443E, 10 m, littoral rainforest, 9.IV.2000 (B.L. Fisher); Sud, Res. de Faune de Campo, Massif des Mamelles, 15.1 km 84\u00b0E \u00c9bodj\u00e9, 2.5942N, 9.9595E, 180 m, rainforest, 4.IV.2000 (B.L. Fisher); Sud-Ouest, Bimbia Forest, 7.4 km 119\u00b0ESE Limbe, 3.9818N, 9.2625E, 40 m, rainforest, 14.IV.2000 (B.L. Fisher); GABON: Province Estuaire, Mondah Forest, near Libreville, 3.XII.1987 (J. Noyes); Province Estuaire, F.C. Mondah, 21 km 331\u00b0NNW Libreville, 0\u00b034.6'N, 9\u00b020.1'E, 10 m, littoral rainforest, 24.II.1998 (B.L. Fisher).CAMEROON: Campo Reserve, Tetramorium hecate differs from the other species of the group by the following character combination: antennal scapes relatively short (SI 73 - 77); eyes large (OI 27 - 31); petiolar node rectangular nodiform with anterodorsal and posterodorsal margins strongly angulate and situated at about the same height; mandibles unsculptured, smooth, and shining; petiole and postpetiole usually with weak sculpture; body colouration ranging from uniformly brown to head, mesosoma, waist segments yellowish to bright orange contrasting with very dark brown to black gaster.(N=12). HL 0.58 - 0.67 (0.63); HW 0.54 - 0.64 (0.60); SL 0.41 - 0.49 (0.45); EL 0.15 - 0.20 (0.17); PH 0.28 - 0.36 (0.33); PW 0.42 - 0.52 (0.47); WL 0.69 - 0.83 (0.77); PSL 0.12 - 0.26 (0.20); PTL 0.20 - 0.26 (0.24); PTH PageBreak0.22 - 0.27 (0.25); PTW 0.17 - 0.22 (0.20); PPL 0.19 - 0.22 (0.21); PPH 0.22 - 0.28 (0.25); PPW 0.24 - 0.29 (0.27); CI 93 - 96 (94); SI 73 - 77 (76); OI 27 - 31 (28); DMI 58 - 63 (61); LMI 41 - 44 (42); PSLI 21 - 38 (32); PeNI 39 - 46 (43); LPeI 89 - 100 (94); DPeI 81 - 89 (84); PpNI 55 - 62 (57); LPpI 80 - 91 (86); DPpI 120 - 130 (126); PPI 129 - 142 (135).PageBreakwaist segments laterally weakly, irregularly rugulose/rugose, dorsally mostly unsculptured, smooth, and shining. Postpetiole and gaster unsculptured, smooth, and shining. Ground sculpture generally faint to absent everywhere on body. Whole body with PageBreakabundant, long, and fine standing hairs; first gastral tergite without appressed pubescence. Anterior edges of antennal scapes with suberect to erect hairs. Body colouration relatively variable, ranging from bicoloured with head, mesosoma, legs, and waist segments yellowish to bright orange contrasting with very dark brown to black gaster to whole body uniformly brown.Head longer than wide (CI 93 - 96); posterior head margin weakly concave. Anterior clypeal margin usually entire and convex, sometimes with very small median notch only visible under higher magnifications. Frontal carinae strongly developed, approaching or ending at posterior head margin. Antennal scrobes well developed, moderately shallow, and with clearly defined margins all around. Antennal scapes relatively short, not reaching posterior head margin (SI 73 - 77). Eyes large (OI 27 - 31). Mesosomal outline in profile weakly convex, moderately marginate from lateral to dorsal mesosoma; promesonotal suture and metanotal groove absent; mesosoma comparatively stout and high (LMI 41 - 44). Propodeal spines usually long to very long (PSLI 30 - 38), elongate-triangular to spinose, and acute, rarely spines reduced, short, elongate-triangular, and acute (PSLI 20 - 21); propodeal lobes short, triangular to elongate-triangular, and acute. Petiolar node in profile rectangular nodiform, approximately as high as long to weakly higher than long (LPeI 89 - 100), anterior and posterior faces approximately parallel, anterodorsal and posterodorsal margins situated at about the same height, anterodorsal and posterodorsal angles well-developed and rectangular, petiolar dorsum flat; node in dorsal view around 1.1 to 1.2 times longer than wide (DPeI 81 - 89). Postpetiole in profile globular to subglobular, approximately 1.1 to 1.2 times higher than long (LPpI 80 - 91); in dorsal view around 1.2 to 1.3 times wider than long (DPpI 120 - 130). Postpetiole in profile appearing less voluminous than petiolar node, in dorsal view approximately 1.3 to 1.4 times wider than petiolar node (PPI 129 - 142). Mandibles unsculptured, smooth, and shining; clypeus longitudinally rugulose, usually with three distinct rugulae, median rugula better developed than remainder, rugulae often with cross-meshes; cephalic dorsum between frontal carinae irregularly longitudinally rugose to reticulate rugose, posteriorly towards posterior head margin well reticulate-rugose, anteriorly towards posterior clypeal margin more regularly longitudinally rugose ; scrobal area mostly unsculptured; lateral and ventral head longitudinally rugose to reticulate-rugose. Mesosoma laterally irregularly rugose, dorsally distinctly longitudinally rugose. Forecoxae unsculptured, smooth, and shining. Both Tetramorium hecate. The species epithet is a nominative noun in apposition, and thus invariant.The name of the new species is inspired by the ancient Latin and Greek goddess \u201cHecate\u201d or \u201cHekate\u201d, also known as the \u201ctriple Hecate\u201d or \u201cthree-faced Hecate\u201d, and refers to the morphological variation in colouration and propodeal spine length observed in Tetramorium hecate is forest leaf litter.The new species is currently only known from Cameroon and Gabon . All locTetramorium hecate is unlikely to be misidentified with the other two species of the group. The most obvious difference between Tetramorium hecate and Tetramorium capillosum and Tetramorium tabarum is the shape of the petiolar node. In the latter two the node is nodiform with relatively rounded anterodorsal and posterodorsal margins, and, in addition, the posterodorsal margin is situated noticeably higher than the anterodorsal margin. In contrast, the node of Tetramorium hecate is nodiform, with clearly rectangular anterodorsal and posterodorsal margins, situated at about the same height. The second-most important character is antennal scape length. Tetramorium hecate has the shortest scapes of the three species (SI 73 - 77), strongly contrasting with the longer scapes of the other two species (SI 80 - 86). Furthermore, Tetramorium hecate is also a much smaller species with much larger eyes and very different colouration than Tetramorium capillosum (see description of the latter for more details). The very well developed antennal scrobes with clearly defined margins all around also separate Tetramorium hecate from Tetramorium tabarum since the scrobes of the latter are shallow and without posterior and ventral margins.PageBreakTetramorium hecate to consist of three potential new species: a strongly bicoloured one with long propodeal spines  [MCZ] [examined].Holotype, pinned worker, DEMOCRATIC REPUBLIC OF CONGO, Epulu, 4.I.1949 (A. Dejean); Sud-Ouest, Bimbia Forest, 7.4 km 119\u00b0ESE Limbe, 3.9818N, 9.2625E, 40 m, rainforest, 14.IV.2000 (B.L. Fisher); CENTRALAFRICANREPUBLIC: Prefecture Sangha-Mba\u00e9r\u00e9, R\u00e9serve Sp\u00e9ciale de For\u00eat Dense de Dzanga-Sangha, 12.7 km 326\u00b0NW Bayanga, 3.005N, 16.1933E, 420 m, rainforest, 10.\u201317.V.2001 (B.L. Fisher); Prefecture Sangha-Mba\u00e9r\u00e9, Parc National Dzanga-Ndoki, 37.9 km 169\u00b0S Lidjombo, 2.3707N, 16.1725E, 360 m, rainforest, 20.\u201328.V.2001 (B.L. Fisher); GABON: Woleu-Ntem, 31.3 km 108\u00b0ESE Minvoul, 2.08N, 12.4067E, 600 m, rainforest, 17.II.1998 (B.L. Fisher).CAMEROON: Mvini, 21.XII.1988 ; eyes large (OI 27 - 31); petiolar node nodiform with anterodorsal and posterodorsal margins relatively rounded, posterodorsal margin situated higher than anterodorsal margin, dorsum convex; mandibles unsculptured, smooth, and shining; petiole with very weak sculpture and postpetiole completely unsculptured; head, mesosoma, waist segments yellowish to bright orange, gaster very dark brown to black.(N=10). HL 0.61 - 0.66 (0.63); HW 0.55 - 0.60 (0.57); SL 0.47 - 0.52 (0.49); EL 0.16 - 0.18 (0.17); PH 0.30 - 0.34 (0.32); PW 0.43 - 0.46 (0.44); WL 0.75 - 0.82 (0.79); PSL 0.14 - 0.16 (0.15); PTL 0.20 - 0.22 (0.20); PTH 0.24 - 0.27 (0.25); PTW 0.16 - 0.18 (0.17); PPL 0.19 - 0.21 (0.20); PPH 0.23 - 0.27 (0.25); PPW 0.24 - 0.27 (0.26); CI 90 - 92 (91); SI 84 - 86 (85); OI 27 - 31 (29); DMI 55 - 58 (56); LMI 38 - 43 (41); PSLI 22 - 25 (23); PeNI 37 - 39 (38); LPeI 78 - 82 (81); DPeI 80 - 85 (82); PpNI 56 - 59 (58); LPpI 79 - 85 (82); DPpI 124 - 130 (127); PPI 147 - 156 (152).PageBreakabsent; mesosoma comparatively stout and high (LMI 38 - 43). Propodeal spines relatively short to medium-sized, elongate-triangular to spinose, and acute (PSLI 22 - 25); propodeal lobes short, triangular to elongate-triangular, and acute. Petiolar node in PageBreakprofile rectangular nodiform, approximately 1.2 to 1.3 times higher than long (LPeI 78 - 82), anterior and posterior faces approximately parallel, posterodorsal margin situated higher than anterodorsal, anterodorsal and posterodorsal angles relatively rounded, petiolar dorsum convex; node in dorsal view approximately 1.2 times longer than wide (DPeI 80 - 85). Postpetiole in profile subglobular and moderately anteroposteriorly compressed, approximately 1.2 to 1.3 times higher than long (LPpI 79 - 85); in dorsal view around 1.2 to 1.3 times wider than long (DPpI 124 - 130). Postpetiole in profile appearing less voluminous than petiolar node, in dorsal view approximately 1.5 to 1.6 times wider than petiolar node (PPI 147 - 156). Mandibles unsculptured, smooth, and shining; clypeus longitudinally rugulose, usually with three rugulae, median rugula better developed; cephalic dorsum between frontal carinae with five to eight longitudinal rugae, most rugae running unbroken from posterior clypeal margin to posterior head margin, few rugae interrupted, but none with cross-meshes; scrobal area mostly unsculptured; lateral and ventral head longitudinally rugose to reticulate-rugose. Mesosoma laterally irregularly rugose, dorsally distinctly longitudinally rugose. Forecoxae unsculptured, smooth, and shining. Petiolar node laterally weakly to moderately longitudinally rugose. Postpetiole and gaster unsculptured, smooth, and shining. Ground sculpture generally faint to absent everywhere on body. Whole body with abundant, long, and fine standing hairs; first gastral tergite without appressed pubescence. Anterior edges of antennal scapes with suberect to erect hairs. Head, mesosoma, legs, and waist segments yellowish to bright orange, contrasting with very dark brown to black gaster.Head significantly longer than wide (CI 90 - 92); posterior head margin moderately concave. Anterior clypeal margin entire and convex. Frontal carinae strongly developed, approaching or ending at posterior head margin. Antennal scrobes developed, but shallow and without clearly defined posterior and ventral margins. Antennal scapes moderately long, not reaching posterior head margin (SI 84 - 86). Eyes large (OI 27 - 31). Mesosomal outline in profile weakly convex, moderately marginate from lateral to dorsal mesosoma; promesonotal suture and metanotal groove Tetramorium tabarum is known to occur in northern Gabon, western Cameroon close to the Gulf of Guinea, the southwest of the Central African Republic, and from the type locality in the northeast of the Democratic Republic of Congo , propodeal spine length (PSLI 22 - 25 vs. PSLI 31 - 43), and sculpture on mandibles and postpetiole, which is present and conspicuous in Tetramorium capillosum while absent in Tetramorium tabarum. The latter is also much smaller (WL 0.75 - 0.82) and bicoloured with a dark brown to black gaster contrasting with the remaining yellowish to orange body, which contrasts with the larger size (WL 1.02 - 1.19) and the uniformly very dark brown to black colouration of Tetramorium capillosum. Despite being often also bicoloured and within the same morphometric range, Tetramorium hecate is unlikely to be confused with Tetramorium tabarum. The antennal scapes are significantly longer in Tetramorium tabarum (SI 84 - 86) than in Tetramorium hecate (SI 73 - 77). More importantly, the petiolar node of Tetramorium tabarum has relatively rounded anterodorsal and posterodorsal margins with the posterodorsal margin situated higher than the anterodorsal, and the dorsum is convex, whereas in Tetramorium hecate the anterodorsal and posterodorsal margins are sharply defined and at about the same height. The varying development of the antennal scrobes is another difference. Tetramorium hecate possesses very well-developed scrobes with margins all around while the scrobes of Tetramorium tabarum are shallow without clear posterior and ventral margins.As already pointed out in the above descriptions of the other two species, Tetramorium tabarum is relatively limited, but it seems that intraspecific variation is very low.The material currently available for"}
+{"text": "Litopenaeus vannamei, and investigated their potential roles in WSSV replication using dsRNA-mediated gene silencing. Lvcaspase2-5 have the typical domain structure of caspase family proteins, with the conserved consensus motifs p20 and p10. Lvcaspase2 and Lvcaspase5 were highly expressed in muscle, while Lvcaspase3 was highly expressed in hemocytes and Lvcaspase4 was mainly expressed in intestine. Lvcaspase2-5 could also be upregulated by WSSV infection, and they showed different patterns in various tissues. When overexpressed in Drosophila S2 cells, Lvcaspase2-5 showed different cellular localizations. Using dsRNA-medicated gene silencing, the expression of Lvcaspase2, Lvcaspase3, and Lvcaspase5 were effectively knocked down. In Lvcaspase2-, Lvcaspase3- or Lvcaspase5-silenced L. vannamei, expression of WSSV VP28 gene was significantly enhanced, suggesting protective roles for Lvcaspase2, Lvcaspase3 and Lvcaspase5 in the host defense against WSSV infection. Apoptosis plays an important role in white spot syndrome virus (WSSV) pathogenesis, and caspases are central players in apoptosis. Here, we cloned four novel caspases (Lvcaspase2-5) from the Pacific white shrimp  Apoptosis plays a protective role in eliminating harmful cells and in the host response to viral infections ,2. When Interference with apoptosis by inhibiting the proteolytic activity of cysteine aspartic acid proteases (caspases) prolongs the life of virus-infected cells, resulting in enhanced viral replication and viral persistence . CaspasePenaeus monodon (Pm) caspase, inhibiting Pm caspase activity in vivo and in vitro . Lv. LvMjcasd 48 hpi . We alsospase2-3 . PmcaspaFigure S1A), Lvcaspase3 (D), Lvcaspase4 (B) and Lvcaspase5 (C) from L. vannamei.Nucleotide and deduced amino acid sequences of Lvcaspase2 ( The nucleotide (upper row) and deduced amino acid (lower row) sequences of Lvcaspase2-5 are shown. The initiation codon (ATG) and stop codon  are shown in bold. The caspase family p20 and p10 domains in Lvcaspase2-5 are shaded. (TIF)Click here for additional data file.Figure S2Domain architectures of shrimp caspases.The full-length protein sequences of shrimp caspases were subjected to the simple modular architecture research tool  to generate domain structures. The p20 and p10 domain are indicated as elliptical boxes, and the prodomain upstream of the p20 domain is indicated as a line. The initiator caspases have a long prodomain (> 90 amino acids) containing specific protein-protein interaction motifs that are necessary for their activation, whereas the effector caspases usually have a short prodomain of only 20-30 residues [residues .(TIF)Click here for additional data file.Figure S3A phylogenetic tree of Lvcaspase2-5 with other caspase family proteins.The full-length amino acid sequences of caspase family proteins from typical organisms were aligned using the ClustalX2.0 program . The rooted tree was then constructed by the \u201cneighbor-joining\u201d method and was bootstrapped 1,000 times using MEGA 4.0 software (http://www.megasoftware.net/index.html). The numbers at the nodes indicate bootstrap values. Lvcaspase2-5 are boxed in blue lines. Lvcasp1, L. vannamei caspase1 (Accession no. ABK88280); Lvcasp2, L. vannamei caspase2 (Accession no. KC660102); Lvcasp3, L. vannamei caspase3 (Accession no. KC660103); Lvcasp4, L. vannamei caspase4 (Accession no. KC660105); Lvcasp5, L. vannamei caspase5 (Accession no. KC660104); Pmcasp1, Penaeus monodon caspase1 (Accession no. AEW91437); Mjcasp3, Marsupenaeus japonicus caspase3 (Accession no. ABK62771); Pmcasp2, Penaeus monodon caspase2 (Accession no. ABO38430); Hscasp1, Homo sapiens caspase1 (Accession no. NP_001214); Mmcasp1, Mus musculus caspase1 (Accession no. NP_033937); Hscasp2, H. sapiens caspase2 (Accession no. AAH02427); Mmcasp2, M. musculus caspase2 (Accession no. NP_031636); Hscasp3, H. sapiens caspase3 (Accession no. NP_116786); Mmcasp3, M. musculus caspase3 (Accession no. NP_033940); Hscasp4, H. sapiens caspase4 (Accession no. NP_001216); Hscasp5, H. sapiens caspase5 (Accession no. NP_001129584); Hscasp6, H. sapiens caspase6 (Accession no. NP_001217); Hscasp7, H. sapiens caspase7 (Accession no. NP_001253987); Mmcasp7, M. musculus caspase7 (Accession no. NP_031637); Hscasp8, H. sapiens caspase8 (Accession no. NP_001073594); Hscas9, H. sapiens caspase9 (Accession no. NP_127463); Hscasp10, H. sapiens caspase10 (Accession no. AAD28403); Hscasp14, H. sapiens caspase14 (Accession no. NP_036246); DmIce, Drosophila melanogaster Ice (Accession no. NP_524551); Dmcasp1, D. melanogaster caspase1 (Accession no. AAB58237); Dmdream, D. melanogaster dream (Accession no. NP_610193); Dmdeath, D. melanogaster death executioner caspase (Accession no. NP_477462); DmNedd2, D. melanogaster Nedd2 (Accession no. NP_524017); Drcasp1, D. rerio caspase1 (Accession no. NP_571580); Drcasp2, D. rerio caspase2 (Accession no. NP_001036160); Drcasp3, D. rerio caspase3 (Accession no. NP_571952); Drcasp6, Danio rerio caspase6 (Accession no. NP_001018333); Drcasp7, D. rerio caspase7 (Accession no. NP_001018443); Drcasp7-2, D. rerio caspase7 like (Accession no. XP_002667104); Drcasp8, D. rerio caspase8 (Accession no. NP_571585); Drcasp9, D. rerio caspase9 (Accession no. NP_001007405); Ggcasp1, Gallus gallus caspase1 (Accession no. XP_003642432); Ggcasp2, G. gallus caspase2 (Accession no. NP_001161173); Ggcasp3, G. gallus caspase3 (Accession no. NP_990056); Ggcasp6, G. gallus caspase6 (Accession no. NP_990057); Ggcasp7, G. gallus caspase7 (Accession no. XP_421764); Ggcasp8, G. gallus caspase8 (Accession no. NP_989923); Ggcasp9, G. gallus caspase9 (Accession no. XP_424580); Ggcasp10, G. gallus caspase10 (Accession no. XP_421936); Ggcasp18, G. gallus caspase18 (Accession no. NP_001038154); Xlcasp1, X. laevis caspase1 (Accession no. NP_001081223); Xlcasp2, X. laevis caspase2 (Accession no. NP_001081404); Xlcasp3, Xenopus laevis caspase3 (Accession no. NP_001081225); Xlcasp7, X. laevis caspase7 (Accession no. NP_001081408); Xlcasp6, X. laevis caspase6 (Accession no. NP_001081406); Xlcasp8, X. laevis caspase8 (Accession no. NP_001079034); Xlcasp9, X. laevis caspase9 (Accession no. NP_001079035); Xlcasp10, X. laevis caspase10 (Accession no. NP_001081410); CeCED3, Caenorhabditis elegans CED3 (Accession no. NP_001255708); CeCSP1, C. elegans CSP1 (Accession no. NP_001022452); CeCSP2, C. elegans CSP2 (Accession no. NP_001023575).(TIF)Click here for additional data file."}
+{"text": "S/GSK1349572, a next-generation HIV-1 integrase inhibitor, has previously demonstrated potent antiviral activity in Phase 2a with once-daily, unboosted dosing. SPRING-1 is an ongoing dose-ranging study designed to select a dose to for Phase 3 evaluation.SPRING-1 is a Phase 2b, multicentre, partially-blinded study in therapy-na\u00efve adults, randomized 1:1:1:1 to 10mg, 25mg or 50mg of S/GSK1349572 or efavirenz(EFV) 600mg once-daily with either co-formulated TDF/FTC or ABC/3TC.205 subjects received study drug: 86% male, 20% non-white, 26%>100,000c/mL HIV-1 RNA, 67% TDF/FTC. Plasma HIV-1 RNA declined rapidly across all S/GSK1349572 doses with no differences in NRTI subgroups. Three protocol-defined virologic failures occurred, 1 on EFV (<1log10 decline by Week 4), and 2 on S/GSK1349572 (Week 4 and 24 rebound >400c/mL with no INI mutation detected). No dose-related clinical or laboratory toxicities were observed. More drug-related AEs of moderate-or-higher intensity were reported on EFV (20%) than S/GSK1349572 (6%) arms; none occurred in more than 1 S/GSK1349572 subject. The most frequent category of such events reported by subjects receiving EFV and S/GSK1349572 were gastrointestinal ; other frequent events on EFV were psychiatric (6%) and rash (4%) disorders. No SAE was considered related to S/GSK1349572. Six subjects (2: S/GSK1349572 and 4: EFV) withdrew due to AEs. Mean change from baseline in LDL cholesterol was +0.023mmol/L among S/GSK1349572 subjects and +0.468mmol/L among EFV subjects. S/GSK1349572 demonstrated low pharmacokinetic variability and drug exposure increased with dose. Table 1S/GSK1349572 administered once-daily without a PK booster was well tolerated with potent antiviral activity at all doses explored in SPRING-1. The greater CD4+ cell increases on S/GSK1349572 merit further observation and confirmation."}
+{"text": "Dear Editor,Escherichia coli O157 and Shigella dysenteriae, in children (E. coli serotypes (i.e. O104:H4). Thrombotic thrombocytopenic purpura (TTP) a closely related syndrome occurs more frequently in adults with more prominent neurological disorders (The hemolytic-uremic syndrome (HUS) is a serious disorder of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure that is frequently associated with Shiga toxins produced in enteric infections by isorders . Both ofisorders . Most pa i.e. O10:H4. ThroE. coli but the strain was not specified. Ultrasound examinations showed right and left kidney dimensions were 91 and 100 mm with bilateral multiple cysts. Intravenous antibiotic therapy with ciprofloxacin and imipenem was started. Patient\u2019s general condition significantly improved and fever subsided. Platelet count dropped to 88000/\u00b5L at the third day of treatment and then dropped to 7000/\u00b5L at the sixth day of antibiotic therapy. The following laboratory data were detected; serum lactic dehydrogenase (LDH): 1150 IU/L, hemoglobin: 7.5 mg/dL, WBC: 11000/\u00b5L, reticulocyte count: 4 (0.2-2/per high power field), total serum bilirubin: 2.3 mg/dL, direct billirubin; 0.5 mg/dL and serum creatinine rose to 6.2 mg/dL, D-dimer test was negative, serum ADAMTS 13 activity was 47%. Pipheral blood smear was positive for fragmented RBC (schistocyte). With the primary diagnosis of TMA, plasma exchange with fresh frozen plasma was started (1.5 L/d). After four days, platelet count rose to 105000/\u00b5L, LDH dropped to 580 U/L, and serum creatinine level returned to 4.0 mg/dL. During all those therapeutic periods, intensive antibiotic therapy was also continued in parallel with plasma exchange.A 65-year old woman was admitted to our nephrology unit with malaise, fever and left flank pain. Her oral temperature was 39\u00b0C, pulse rate was 100 per minute, and blood pressure was 120/70 mmHg. Initial laboratory tests revealed leukocytosis 31000/\u00b5L, serum creatinin: 4.7 mg/dL, hemoglobin: 9.4 mg/dL, MCV: 88 fL (80-99), MCH: 27 pg (27-31), MCHC: 31 (32-36), cholesterol: 124 mg/dL, triglyceride: 232 mg/dL, platelet count: 246000/\u00b5L, serum calcium: 8.6 mg/dL, serum phosphor: 6.3 mg/dL, i-PTH: 768 pg/mL, plasma Na: 142 meq/L, plasma k: 4.9 meq/L, AST: 11 (0-31 IU/L), ALT: 10 (0-41 IU/L), alkaline phosphatase: 326 (64-306 U/L), serum iron: 26 (40-165 \u00b5g/dL), TIBC: 247 (250-450 IU/L), and serum ferritin was 180 . Urinalysis showed numerous white and red blood cells (RBC), without any dysmorphic RBC. Serum protein electrophoresis result was negative for paraproteinemia. Blood culture was positive for E. coli bacteremia. However this should not discourage the administration of appropriate antimicrobial agents in this situation, as it may lead to death (Antibiotic therapy could increase the risk of full-blown HUS in children with STEC enteric infection . We obseto death . We foun"}
+{"text": "Orientia tsutsugamushi, is an endemic disease in Taiwan and may be potentially fatal if diagnosis is delayed.Scrub typhus, a mite-transmitted zoonosis caused by We encountered a 23-year-old previously healthy Taiwanese male soldier presenting with the right ear pain after training in the jungle and an eleven-day history of intermittent high fever up to 39\u00b0C. Amoxicillin/clavulanate was prescribed for otitis media at a local clinic. Skin rash over whole body and abdominal cramping pain with watery diarrhea appeared on the sixth day of fever. He was referred due to progressive dyspnea and cough for 4 days prior to admission in our institution. On physical examination, there were cardiopulmonary distress, icteric sclera, an eschar in the right external auditory canal and bilateral basal rales. Laboratory evaluation revealed thrombocytopenia, elevation of liver function and acute renal failure. Chest x-ray revealed bilateral diffuse infiltration. Doxycycline was prescribed for scrub typhus with acute respiratory distress syndrome and multiple organ failure. Fever subsided dramatically the next day and he was discharged on day 7 with oral tetracycline for 7 days.Scrub typhus should be considered in acutely febrile patients with multiple organ involvement, particularly if there is an eschar or a history of environmental exposure in endemic areas. Rapid and accurate diagnosis, timely administration of antibiotics and intensive supportive care are necessary to decrease mortality of serious complications of scrub typhus. Orientia tsutsugamushi, is an endemic disease in Taiwan and may be potentially fatal if diagnosis is delayed. It is an acute febrile illness characterized by a typical necrotic primary lesion (eschar), generalized lymphadenopathy, maculopapular skin rash, and nonspecific symptoms and signs. The incidence of eschar reported in patients with scrub typhus is low, < 25%, and only 5% are found over head, face and neck. An eschar in the external auditory canal has not been reported previously, so the diagnosis of scrub typhus was delayed with serious complications, including acute respiratory distress syndrome (ARDS) and multiple organ failure (MOF). In patients with scrub typhus, the incidence and mortality rate of ARDS are 11.1%-15.2% and 20%-25%, respectively. Reported here is a case of an eschar in the external auditory canal in scrub typhus complicated by ARDS and MOF in a 23-year-old previously healthy Taiwanese male soldier.Scrub typhus, a mite-transmitted zoonosis caused by 3 (reference range [RR]: 4000-11000/mm3) with 87.9% segmented neutrophils, hemoglobin 14.6 g/dL (RR: 14-16 g/dL), platelet counts 24 \u00d7 103/mm3 (RR: 140-400 \u00d7 103/mm3), blood urea nitrogen 25 mg/dL, creatinine 1.5 mg/dl (RR: 0.7-1.4 mg/dL), sodium 140 mEq/L, potassium 3.9 mEq/L, chloride 108 mEq/L, calcium 8.2 mg/dL, total protein 5.8 g/dL (RR: 6.0-8.0 g/dL), albumin 3 g/dL (RR: 3.5-5.0 g/dL), total bilirubin 4.8 mg/dL (RR: 0.1-1.2 mg/dL), direct bilirubin 2.7 mg/dL (RR: 0.0-0.2 mg/dL), C-reactive protein 16.23 mg/dL (RR <0.3 mg/dL), aspartate aminotransferase (AST) 368 IU/L (RR: 8-38 IU/L), alanine aminotransferase (ALT) 271 IU/L (RR: 4-44 IU/L), alkaline phosphatase (ALK) 324 IU/L (RR: 50-190 IU/L), lactate dehydrogenase 783 IU/L (RR: 120-240 IU/L), glucose 94 mg/dL, creatine phosphokinase 229 IU/L (RR: 10-160 IU/L), and a positive for Weil-Felix reaction with a Proteus OX-K titer of 1:1280 on day 11 of fever. Arterial blood gas analysis was pH 7.501, PaCO2 38.1 mmHg, PaO2 76 mmHg, HCO3- 30.1 mmol/l, and BEB 7.2 with a FiO2 of 60%. A central venous line was setup for monitoring his central venous pressure and fluid replacement because of hypotension. Chest x-ray , is a disease distributed throughout the Asia Pacific rim and endemic in South Korea, China, Japan, Taiwan, Pakistan, India, Thailand, Malaysia and northern Australia. It is an acute febrile illness characterized by a typical necrotic primary lesion (eschar), generalized lymphadenopathy, maculopapular skin rash, and nonspecific symptoms and signs such as fever, chills, headache, alerted sensorium, seizure, sorethroat, otalgia, cough, dysponea, vomiting, abdominal pain, jaundice, diarrhea, hepatosplenomegaly, abnormal bleeding, arthralgia and myalgia[Orientia tsutsugamushi [Proteus OX-K titer of 1:1280 was obtained on day 11 after exposure in our case. While all current scrub typhus tests have limitations and the Weil-Felix and immunofluorescent antibody (IFA) tests have low sensitivity and specificity[Scrub typhus, a mite-transmitted zoonosis caused by the intracellular Gram-negative bacteria, d myalgia-12. Unfod myalgia,12. Serid myalgia. In our d myalgia. Initiald myalgia. Acute rugamushi ,12. Abnougamushi ,8-10,12.ecificity,12. The ecificity,12. ScruWe recommend that scrub typhus should be taken into consideration in acutely febrile patients with varying degree of respiratory distress, impaired liver function, acute renal failure and hematologic involvement, particularly if there is an eschar or a history of environmental exposure in an endemic area-9,11,12.The authors declare that they have no competing interests.BJL took care of this patient in the intensive care units. CYC drew up the first draft of the report. SYH made a substantial contribution to draft the manuscript and revised the draft all over the course of submission. All authors read and approved the final manuscript.Written informed consent was obtained from the patient for publication of this study. A copy of the written consent is available for review by the Editor-in-Chief of this journal.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/11/79/prepub"}
+{"text": "Anopheles gambiae, Aedes aegypti, Culex quinquefasciatus, Ixodes scapularis, Pediculus humanus and Rhodnius prolixus. We observed that (1) disease vectors encode a lower percentage of ubiquitin-related genes when compared to Drosophila melanogaster, Mus musculus and Homo sapiens but not Saccharomyces cerevisiae; (2) overall, there are more proteins categorized as E3 ubiquitin ligases when compared to E2-conjugating or E1-activating enzymes; (3) the ubiquitin machinery within the three mosquito genomes is highly similar; (4) ubiquitin genes are more than doubled in the Chagas disease vector (R. prolixus) when compared to other arthropod vectors; (5) the deer tick I. scapularis and the body louse (P. humanus) genomes carry low numbers of E1-activating enzymes and HECT-type E3 ubiquitin ligases; (6) R. prolixus have low numbers of RING-type E3 ubiquitin ligases; and (7) C. quinquefasciatus present elevated numbers of predicted F-box E3 ubiquitin ligases, JAB and UCH deubiquitinases. Taken together, these findings provide novel opportunities to study the interaction between a pathogen and an arthropod vector. Protein regulation by ubiquitin has been extensively described in model organisms. However, characterization of the ubiquitin machinery in disease vectors remains mostly unknown. This fundamental gap in knowledge presents a concern because new therapeutics are needed to control vector-borne diseases, and targeting the ubiquitin machinery as a means for disease intervention has been already adopted in the clinic. In this study, we employed a bioinformatics approach to uncover the ubiquitin-mediated pathway in the genomes of  Vector-borne diseases are some of the most prevalent infectious illnesses worldwide. According to the World Health Organization, there are approximately 216 million cases of malaria alone, with over 1.2 million estimated deaths . OverallUbiquitin consists of a 76 amino acid protein that carries seven lysine (K) residues  . UbiquitDrosophila, it is established that developmental regulation by the evolutionarily conserved Notch receptor depends on ubiquitin [Drosophila because ubiquitination also regulates these pathways in disease vectors, such as: Anopheles gambiae, Aedes aegypti, Culex quinquefasciatus and Ixodes scapularis [In biquitin , and sombiquitin . Additiobiquitin . This imapularis .An. gambiae, Ae. aegypti, C. quinquefasciatus, I. scapularis, Pediculus humanus and Rhodnius prolixus. We compared our findings to datasets available for Saccharomyces cerevisiae, Drosophila melanogaster, Mus musculus and Homo sapiens [The lack of information regarding the role of ubiquitin in disease vectors is problematic since molecular interactions depend on this posttranslational modification. These biochemical signatures may lead to novel factors to control pathogen transmission and/or acquisition . For ins sapiens -13. Whil sapiens ,15, tran sapiens ,16-19 an sapiens ,20,21, iI. scapularis, An. gambiae, Ae. aegypti, C. quinquefasciatus, P. humanus and R. prolixus were downloaded from VectorBase [S. cerevisiae, D. melanogaster, H. sapiens and M. musculus were downloaded from http://www.yeastgenome.org [Complete protein datasets encoded within the genomes of ctorBase . Proteinnome.org , http://nome.org , and httnome.org , respectnome.org , and downome.org . This in<0.5. Importantly, observed discrepancies between our results and findings previously published for S. cerevisiae, D. melanogaster, H. sapiens and M. musculus [Using the selected Pfam domains, the hmmsearch program of the HMMER v3.0 package  was emplmusculus -13 were I. scapularis, An.gambiae, Ae. aegypti, and C. quinquefasciatus were aligned using the Multiple Sequence Alignment tool MUSCLE [ The resulting protein sequences from l MUSCLE  availabll MUSCLE . The ful For each aligned dataset, protein phylogenies were estimated with the maximum likelihood method, employing the PhyML program  availabl-like modifier (SUMO), neural precursor cell expressed developmentally downregulated gene 8 (NEDD8), autophagy-related protein 8 (ATG8), ubiquitin-related modifier-1 (URM1), homologous to ubiquitin 1 (HUB1), E1s (ubiquitin-activating enzyme 1 (UBA1), UBA2, UBA3, UBA1-like, autophagy-related E1-like enzyme (ATG7)), really interesting new gene (RING) , other E3s (homology to E6AP C-terminus (HECT), Cullin, U-box), and DUBs (ovarian tumor (OTU), Josephin, jun activation domain-binding protein (JAB), domain of unknown function (DUF) 862, Wss1p\u2010like metalloproteases (WLM), ubiquitin carboxyl-terminal hydrolase (UCH), Peptidase_C12, Peptidase_C48, and Peptidase_C54). For direct classification of ubiquitin-like proteins, we manually analyzed and categorized each protein encoded in the genomes of An. gambiae, Ae. aegypti, C. quinquefasciatus, and I. scapularis. Ubiquitin-like proteins analyzed were SUMO, URM1, NEDD8, HUB1 and ATG8. Proteins containing two or more Pfam domains were observed and recorded in Protein sequences belonging to specific subcategories of the five ubiquitination-components , ubiquitin-conjugating enzymes (E2), ubiquitin ligases (E3) and deubiquitinases) were identified based on the Pfam domain matches obtained from hmmsearch. Whenever possible, proteins were cataloged according to their major branching patterns in their respective phylogenies: ubiquitin/ubiquitin-like proteins , while P. humanus had the highest ratio at 3.13. R. prolixus had a %U of 2.25. When compared to four model species, all disease vectors possessed a much lower %U, with the model species ranging from 4.38 to 5.02. The exception was S. cerevisiae (%U = 2.24). Interestingly, D. melanogaster carried a much higher %U at 4.50.  The ubiquitin machinery regulates fundamental biological processes within eukaryotic cells . Thus, wI. scapularis (31), An.gambiae (40), Ae. aegypti (48), C. quinquefasciatus (40), P. humanus (35) and R. prolixus (129) (S. cerevisiae (7), D. melanogaster (30), H. sapiens (45) and M. musculus (41). Apart from S. cerevisiae (7) and R. prolixus (129), our analysis revealed a similar number of ubiquitin proteins across species. We further analyzed sequence similarities among ubiquitin family members  for An. gambiae, Ae. aegypti, C. quinquefasciatus, and I. scapularis from their multiple alignments  . We alsoignments . An. gambiae, Ae. aegypti, C. quinquefasciatus, and I. scapularis ( Ubiquitin-like proteins were first discovered in 1979 and named after the interferon-stimulated gene 15 (ISG15) . Since iapularis . UbiquitAn. gambiae (2), Ae. aegypti (2), C. quinquefasciatus (1), and I. scapularis (1)  . NEDD8 ih vector .An. gambiae, Ae. aegypti, C. quinquefasciatus, and I. scapularis , Ae. aegypti (2), C. quinquefasciatus (3), and I. scapularis (3) (URM1 is a ubiquitin-like protein that shares very little homology with ubiquitin . Four prapularis . HUB1 dih vector . ATG8 isaris (3) . I. scapularis (16), An. gambiae (29), Ae. aegypti (37), C. quinquefasciatus (30), P. humanus (19) and R. prolixus (26)  have a very similar number of E1 proteins, while I. scapularis and P. humanus have noticeably less E1s than the other disease vectors. We also analyzed the E1 number in S. cerevisiae (8), D. melanogaster (30), H. sapiens (42), and M. musculus (25)  domain. The E1 associated to ubiquitin is known as UBA1. UBA2 and UBA3 serve as activators for SUMO and NEDD8 respectively . The E1 xus (26) . The thrlus (25) . We furttimation . I. scapularis (32), An. gambiae (32), Ae. aegypti (36), C. quinquefasciatus (28), P. humanus (29) and R. prolixus (31)  . The numapularis . I. scapularis (151), An.gambiae (147), Ae. aegypti (161), C. quinquefasciatus (156), P. humanus (127) and R. prolixus (86) (3HisCys4. The zf-C3HC4 subcategory was the most common RING-type ligase within selected vectors: I. scapularis (100), An. gambiae (82), Ae. aegypti (84), C. quinquefasciatus (87), P. humanus (80) and R. prolixus (56)  . The numxus (56) -7. I. scapularis (28), An.gambiae (42), Ae. aegypti (43), C. quinquefasciatus (47), P. humanus (31) and R. prolixus (19) (I. scapularis (17), An. gambiae (12), Ae. aegypti (22), C. quinquefasciatus (12), P. humanus (16) and R. prolixus (11) (2HC2 motif and plays a role in DNA replication [I. scapularis (6), An. gambiae (11), Ae. aegypti (12), C. quinquefasciatus (10), P. humanus (6) and R. prolixus (8) (We report the number of zf-Apc11 proteins in xus (19) -7. RINGvxus (11) -7. Rtf2 lication . We analixus (8) -7. It isixus (8) .An. gambiae, Ae. aegypti, C. quinquefasciatus, and I. scapularis by phylogeny estimation , An. gambiae (17), Ae. aegypti (17), C. quinquefasciatus (23), P. humanus (14) and R. prolixus (16)  . The numtimation . I. scapularis (5), An.gambiae (6), Ae. aegypti (9), C. quinquefasciatus (4), P. humanus (7) and R. prolixus (8) (S. cerevisiae (4), D. melanogaster (14), H. sapiens (16), and M. musculus (11)  . The numlus (11) . We then ligases . SimilarI. scapularis (8), An.gambiae (10), Ae. aegypti (10), C. quinquefasciatus (12), P. humanus (7) and R. prolixus (8)  . Much liixus (8) . I. scapularis (43), An. gambiae (39), Ae. aegypti (49), C. quinquefasciatus (151), P. humanus (32) and R. prolixus (39) , D. melanogaster (46), H. sapiens (111), and M. musculus (104)  . We then species . I. scapularis (7), An. gambiae (9), Ae. aegypti (9), C. quinquefasciatus (9), P. humanus (7), R. prolixus (6), S. cerevisiae (2), D. melanogaster (12), H. sapiens (21), and M. musculus (18)  . In mostapularis , S10. I. scapularis (2), An. gambiae (1), Ae. aegypti (1), C. quinquefasciatus (2), P. humanus (2), R. prolixus (2), S. cerevisiae (0), D. melanogaster (1), H. sapiens (13), and M. musculus (10) , a neurodegenerative condition . We repolus (10) . All selapularis . I. scapularis (12), An. gambiae (10), Ae. aegypti (11), C. quinquefasciatus (18), P. humanus (10) and R. prolixus (8)  DUBs . This spixus (8) . Proteinapularis . I. scapularis (3), An.gambiae (1), Ae. aegypti (2), C. quinquefasciatus (3), P. humanus (2) and R. prolixus (11)  . The numlogenies . I. scapularis (2), An. gambiae (2), Ae. aegypti (0), C. quinquefasciatus (1), P. humanus (0), and R. prolixus (0)  . I. scapularis (25), An. gambiae (29), Ae. aegypti (28), C. quinquefasciatus (34), P. humanus (26) and R. prolixus (29) , Ae. aegypti (6), C. quinquefasciatus (8), and I. scapularis (5). ( DUBs containing the UCH domain have been known to release ubiquitin in polyubiquitin chains by hydrolyzing its C-terminus . We perfxus (29) . UCH-assris (5). . I. scapularis , An. gambiae , Ae. aegypti , C. quinquefasciatus , P. humanus , R. prolixus , S. cerevisiae , D. melanogaster , H. sapiens , and M. musculus   protease family ,52. Pept_C54: 5) . In mosth vector . Vector-borne illnesses threaten public health in the tropics. Environmental changes due to globalization and the lack of effective vaccines are also contributing to the spread of these maladies to temperate climates. Thus, novel treatments are necessary in the clinic. One effective therapeutic strategy recently used to treat cancer, neurodegenerative disorders and some infectious diseases is pharmacological intervention targeting (de)ubiquitination. However, the use of (de)ubiquitination as a therapeutic target to combat arthropod-borne diseases has not been established. A possible reason for this gap in translational research is the inherent complexity of vector-borne diseases. Another possibility is the lack of communication among scientists. Pharmacologists and chemists do not typically interact with vector biologists, and vector biologists have little incentive to interact with the drug development community. An. gambiae, Ae. aegypti, C. quinquefasciatus, I. scapularis, P. humanus and R. prolixus. Although independent work will be necessary to validate our bioinformatics analysis, empirical evidence suggests that the ubiquitin machinery is present in disease vectors. Recently, Severo et al., 2013 characterized a RING-type E3 ubiquitin ligase named x-linked inhibitor of apoptosis protein (XIAP) and showed the importance of ubiquitination for microbial pathogenesis in ticks [Here we took advantage of publicly available arthropod genomes and described the ubiquitin machinery of in ticks . Similarin ticks . Corroboin ticks . In summTable S1Source and release date of protein datasets.(PDF)Click here for additional data file.Table S2An.List of proteins in gambiae identified by the HMM search.(XLSX)Click here for additional data file.Table S3Ae. aegypti identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S4C. quinquefasciatus identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S5I. scapularis identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S6P. humanus identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S7R. prolixus identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S8D. melanogaster identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S9H. sapiens identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S10M. musculus identified by the HMM search.List of proteins in (XLSX)Click here for additional data file.Table S11List of protein identifications containing sequences matching two or more Pfam domains.(XLSX)Click here for additional data file.Figure S1I. scapularis, An.Ubiquitin and ubiquitin-like proteins in gambiae, Ae. aegypti, and C. quinquefasciatus. Protein sequences matched with ubiquitin and ubiquitin-like proteins were aligned using MUSCLE and a phylogeny was estimated with the PhyML software. Sequences with a bootstrap value lower than 50 were manually removed. Bootstrap values ranged from 0.64 - 0.86 for highlighted clustered categories . (TIF)Click here for additional data file.Figure S2I. scapularis, An.Ubiquitin and ubiquitin-like activating enzymes in gambiae, Ae. aegypti, and C. quinquefasciatus. Phylogeny of ubiquitin and ubiquitin-like activating enzymes. Matched sequences for ubiquitin and ubiquitin-like activating enzymes were aligned using MUSCLE and a phylogeny was estimated with the PhyML software. Bootstrap values ranged from 0.43 - 0.83 for highlighted clustered categories .(TIF)Click here for additional data file.Figure S3I. scapularis, An.Ubiquitin and ubiquitin-like conjugating enzymes in gambiae, Ae. aegypti, and C. quinquefasciatus. Protein sequences matched with ubiquitin and ubiquitin-like conjugating enzymes were aligned using MUSCLE and a phylogeny was estimated with the PhyML software. (TIF)Click here for additional data file.Figure S4An.Phylogeny of RING and RING-like ubiquitin ligases in gambiae and Ae. aegypti. Protein sequences matched with ubiquitin and ubiquitin-like ligases were aligned using MUSCLE and a phylogeny was estimated with the PhyML software. Proteins not listed under a specific subset were categorized as zf-C3HC4. Phylogeny of RING and RING-like ubiquitin ligases in (A) An. gambiae and (B) Ae. aegypti. Bootstrap values ranged from (A) 0.41 - 0.80 and (B) 0.41 - 0.83 for highlighted clustered categories .(TIF)Click here for additional data file.Figure S5C. quinquefasciatus and I. scapularis.Phylogeny of RING and RING-like ubiquitin ligases in  Protein sequences matched with ubiquitin and ubiquitin-like ligases were aligned using MUSCLE and a phylogeny was estimated with the PhyML software. Proteins not listed under a specific subset were categorized into the zf-C3HC4 classification. Phylogeny of RING and RING-like ubiquitin ligases in (A) C. quinquefasciatus and (B) I. scapularis. Bootstrap values ranged from (A) 0.43 - 0.83 and (B) 0.46 - 0.83 for highlighted clustered categories .(TIF)Click here for additional data file.Figure S6I. scapularis, An.HECT, Cullin and U-box ligases in gambiae, Ae. aegypti, and C. quinquefasciatus. Protein sequences matched with ubiquitin and ubiquitin-like ligases were aligned using MUSCLE and the phylogeny was estimated with the PhyML software. Sequences with bootstrap values lower than 50 were manually removed. Bootstrap values ranged from 0.35 - 0.88 for highlighted clustered categories .(TIF)Click here for additional data file.Figure S7An.Phylogenetic trees of F-box ubiquitin ligases in gambiae and Ae. aegypti. Protein sequences matched with F-box ligases were aligned using MUSCLE and the phylogeny was estimated with the PhyML software. Sequences with a bootstrap values lower than 50 were manually removed. Phylogenetic trees of F-box ubiquitin ligases in (A) An. gambiae and (B) Ae. aegypti. (TIF)Click here for additional data file.Figure S8C. quinquefasciatus and I. scapularis.Phylogenetic trees of F-box ubiquitin ligases in  Protein sequences matched with F-box ligases were aligned using MUSCLE and the phylogeny was estimated with the PhyML software. Sequences with a bootstrap values lower than 50 were manually removed. Phylogenetic trees of F-box ubiquitin ligases in (A) C. quinquefasciatus and (B) I. scapularis. (TIF)Click here for additional data file.Figure S9An.Phylogenetic trees of deubiquitinases in gambiae and Ae. aegypti. Protein sequences were aligned using MUSCLE and a phylogeny was estimated using the maximum likelihood method. Phylogeny of deubiquitinases in (A) An. gambiae and (B) Ae. aegypti. Bootstrap values ranged from (A) 0.43 - 0.87 and (B) 0.37 - 0.84 for highlighted clustered categories .(TIF)Click here for additional data file.Figure S10C. quinquefasciatus and I. scapularis.Phylogenetic trees of deubiquitinases in  Protein sequences were aligned using MUSCLE and a phylogeny was estimated using the maximum likelihood method. Phylogeny of deubiquitinases in (A) C. quinquefasciatus and (B) I. scapularis. Bootstrap values ranged from (A) 0.39 - 0.83 and (B) 0.37 - 0.78 for highlighted clustered categories .(TIF)Click here for additional data file."}
+{"text": "Candida species play an important complications in severely-ill hospitalized patients which may lead to therapeutic failure. During a 5-year period from 2006-2010, 7 Candida species and 14 unidentified strains were identified from 887 candidaemia of patients admitted at Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.Bloodstream infections due to Candida species were identified according to characteristics of chlamydoconidia production and carbohydrate assimilation/fermentation profile using 13/6 carbon substrates. Only a limited number of Candida were tested for antifungal susceptibility by agar gradient diffusion (Etest\u00ae)to amphotericin B and fluconazole.Candida albicans remains a major pathogen accounted for 50.3%, followed by C. tropicalis(27.4%), C. parapsilosis (12.5%), C.guilliermondii (6%), C. glabrata (1.8%), C. krusei (0.23%), C. rugosa (0.23%), and unidentified species(1.6%). From a total of 103 C. albicans, amphotericin B MIC ranged from 0.016-1ug/ml whereas fluconazole MIC ranged were between 0.023 to > 256ug/ml. Other species tested were C. tropicalis (73 strains) which yielded MIC range of 0.19-2ug/ml for amphotericin B and 0.5->256 ug/ml for fluconazole. 34 strains of C. parapsilosis gave amphotericin B ranged from 0.023-16ug/ml and 0.094->256 for fluconazole.From a total of 887 isolates, C. albicans remains the most prevalence Candida species causing candidaemia followed by C. tropicalis, C. parapsilosis and C. guilliermondii. Those non-albicans Candida seemed to express decrease susceptibility to both major antifungal agents like amphotericin B and fluconazole whereas a small number of C. albicans gave high MIC to fluconazole.None declared."}
+{"text": "Stress perfusion (SP) abnormalities predict prognosis in specific subsets of patients with coronary artery disease (CAD). However, results are more controversial in populations with stable CAD. We tried to assess prognostic value of SP cardiac magnetic resonance (CMR) in unselected patients with chronic CAD, beyond conventional risk factors such as left ventricle ejection fraction (LVEF).We considered consecutive patients undergoing CMR for definite/suspected chronic CAD. CMR included first-pass myocardial perfusion assessment after dipyridamole stress (0.56 mg/kg) and late gadolinium enhancement (LGE). CMR images were semi-quantitatively evaluated by two blinded operators. SP-CMR was judged positive if a perfusion defect \u2265 75% of myocardium wall thickness was detected in areas of myocardium without LGE. We also collected clinical and pharmacological history, atherosclerotic risk factors, ECG end Echocardiography data. Events in the follow-up , were recorded with outpatient/inpatient visits or with structured telephonic interviews. Univariate and multivariate Cox hazard analyses were performed, with continuous variables categorized according to cut-offs from the literature.We recruited 374 patients , between January 2002 and December 2006. During a follow-up period of 38\u00b121 months there were 30 deaths (8%). Moreover, 79 patients (21%) underwent revascularization procedures. Univariate analysis showed an association between SP-CMR and mortality . Stronger predictors of death were LVEF (19.1 [6.5-55.7]), LVWMSI (13.2 [4.6-38.2]), LVESV (13.1 [4.9-35.3]), LVEDV (9.2 [3.5-24.6]), LGE (8.3 [3.5-19.5]), anticoagulant (6.3 [2.7-14.3]), QTc interval (5.9 [2.9-12.2]) and loop diuretics . A significant univariate association was also found with LV diastolic function (4.4 [1.9-9.9]), aldosterone antagonist (4.2 [1.9-9.0]) and QRS duration (3.8 [1.8-7.9]). At multivariate analysis SP-CMR was found to be no more significant and independent predictors of death were only LVEF , QTc interval , non-sinusal rhythm  and LGE .A positive SP-CMR was the strongest predictor of non-CMR related revascularization procedures, at univariate as well as at multivariate analysis . Other independent predictors of revascularization need in the follow-up were only diabetes  and smoking habit .Our study showed lack of independent association between stress induced perfusion defects at CMR and mortality in a large group of unselected patients with stable CAD. Conversely, stress CMR was the strongest predictor of subsequent revascularization procedures unrelated to CMR itself."}
+{"text": "This study aimed at assessing left ventricular (LV), left atrial (LA) and right ventricular (RV) volumes in patients before and after MitraClip\u2122 implantation by cardiac magnetic resonance imaging (CMR).The MitraClip\u2122 is a novel device for percutaneous mitral valve repair. Recent studies demonstrated a reduction of LV volumes after MitraClip\u2122 implantation using echocardiography. CMR is currently the reference method to assess cardiac volumes but has not been used to assess LV remodeling after MitraClip\u2122 implantation so far.Twelve patients with functional (n=7) or degenerative (n=5) mitral valve regurgitation grade 3 to 4 underwent CMR at baseline (BL) before and at 6 month follow-up (FU) after successful MitraClip\u2122 implantation. CMR protocol consisted of short- and long-axis slices using a steady-state-free-precession cine sequence for the assessment of LV, LA and RV volumes.2; p=0.03) as well as LVESV (82 (54-91) vs. 69 (48-99) ml/m2; p=0.03) indices significantly decreased from BL to FU. No significant difference was found for RVEDV (94 (75-103) vs. 99 (77-123) ml/m2; p=0.91), RVESV (48 (42-80) vs. 51 (40-81) ml/m2; p=0.48) and LAV (87 (55-124) vs. 92 (48-137) ml/m2; p=0.20) indices between BL and FU.Minor device-related artifacts were observed, enabling reliable delineation of endocardial borders in all patients at FU. Figure CMR enables reproducible measurements of cardiac volumes in patients with implanted MitraClip\u2122 devices. Significantly decreased LV but unchanged LA and RV volumes were found at 6 month after successful MitraClip\u2122 implantation.No external funding."}
+{"text": "There was an error in the second author's name. The correct spelling is Sudhakar Agnihothram. The correct citation is: Varshney B, Agnihothram S, Tan Y-J, Baric R, Lal SK (2012) SARS Coronavirus 3b Accessory Protein Modulates Transcriptional Activity of RUNX1b. PLoS ONE 7(1): e29542."}
+{"text": "DAI are a serious public health problem worldwide. The PC and optimization of the HHA have proven to be excellent tools for DAI minimization or elimination.Assess the impact of PC and HHA on DAI in our AICU.A quasi-experimental study. We calculated the average rates of DAI using NHSN,CDC methodology in two periods:pre (Pa) and post intervention (Pb). Average rates wereexpressed as number of events per 1000 device days (DD). Study periods: ventilator associated pneumonia (VAP) Pa: January 2004-January 2010, Pb: February 2010-February 2013. Catheter-associated bacteremia (BACT) Pa: January 2004-June 2010, Pb: July 2010-February 2013. Catheter-associated urinary tract infection (UTI) Pa: January 2007-October 2011Pb: November 2011-February 2013. The Institute of Healthcare Improvement (IHI) proposed PC were implemented. PC adherence was assessed periodically. In October 2009, WHO campaign to improve HHA was implemented. Periodic measurements of HHA were performed. Statistical analysis: we compared average rates of HAI in both periods with Mann-Whitney test. We calculated the Incidence Rate Ratio (IRR) as Pb average incidence rate/Pa average incidence rate.NEU rate decreased from 9.88 (Pa) to 2.60 (Pb), P <0.001. IRR 0.26, 74% rate reduction. Attributable risk: 7.28/ 1000 DD. Cases avoided in Pb: 33.4 (4589 DD in three years). BACT rate decreased from 5.35 (Pa) to 2.34 (Pb), p 0.007. IRR: 0.44, 56% rate reduction. Attributable risk: 3.01/ 1000 DD. Cases avoided in Pb: 14.99 (4983 DD in 32 months). ITU rate decreased from 2.45 (Pa) to 1.30 (Pb), P 0.32. IRR: 0.53, 47% rate reduction. Attributable risk: 1.15/ 1000 DD. Cases avoided in Pb: 3.34 (2908 DD in 16 months). Adherence to PC and HHA ranged between 80 and 95%.PC implementation and HHA optimization in our AICU was associated with statistically significant decreases in VAP and BACT rates, and similar tendency of UTI rate. The significant number of cases averted, fully justifies the implementation of these tools.None declared."}
+{"text": "Once daily ritonavir-boosted darunavir (DRV/RTV) is a preferred antiretroviral regimen for treatment-na\u00efve patients. The pharmacokinetics (PK) of DRV/RTV may be influenced by patient demographics and co-medications. Also with an increased aging HIV population, investigations into the impact of older age on PK are important.0-24) and baseline CD4 cell count were 39yr (21-63), 74kg (57-105), 24kg/m2 (18-31), 4.35mg.h/L (2.27-10.99) and 500cells/mm3 (227-1129), respectively; 49 patients had undetectable viral load at time of study. PK sampling was performed at steady-state and between 1-3 PK curves were available per patient. Nonlinear mixed effects modelling (NONMEM v. VI 2.0) was applied to determine DRV PK parameters, interindividual and interoccasion variability and residual error. The following covariates were evaluated: age, weight, BMI, sex, ethnicity, RTV AUC0-24 and raltegravir co-medication (400mg twice daily). The model was validated by means of simulation and visual predictive check.Data were pooled from 3 DRV/RTV PK studies. In total 51 HIV-infected patients  stable on DRV/RTV  were included. Median age, weight, BMI, RTV area under the curve over 24h (AUCa 0.914h-1) and lag-time (0.358h) best described the data. Inclusion of a different apparent oral clearance (CL/F) and volume of distribution (V2/F) for one of the studies improved the fit . RTV AUC0-24 and age were significantly associated with DRV CL/F. An increase in age of 1yr produced a fractional decrease in DRV CL/F of 0.014, equivalent to a 14% reduction in CL/F with every 10yr increase in age. Based on the visual predictive check 94% of observed DRV concentrations were within the 95% prediction interval, indicative of an adequate model.A 2-compartment model with first-order absorption (k0-24 and age were significantly related to DRV CL/F. The impact of age requires further investigation and clarification over a wide age range, particularly in an elderly population.A population model describing the PK of once daily RTV-boosted DRV has been developed and validated. RTV AUC"}
+{"text": "Tocilizumab (TCZ), an IL-6 receptor inhibitor, improves arthritis and systemic symptoms associated with systemic JIA. It may be a valuable option in patients with JIA who show inadequate response to anti-TNF agents.To analyze the short-term effectiveness and safety of TCZ in patients with JIA refractory to anti-TNF agents.All patients who received TCZ due to refractory disease were included. All patients had exhibited anti-TNF primary or secondary failure. Data were retrieved from the Rheumatology Section data base. TCZ was administered intravenously at a dose of 8 mg/kg for patients > 30 Kg, 12 mg/Kg for patients < 30 Kg every 2 weeks. Efficacy endpoints included improvement (ACR Pedi30), disappearance of systemic symptoms, and reduction in corticosteroid dose.9 patients with JIA  were included. Median age at study entry was 10 years, disease duration 6 years. They received TCZ for 3 to 9 months. Patients had been refractory to etanercept (9), adalimumab (7) or infliximab (2). At baseline (medians): active joints: 11, joints with limited motion: 4, wellbeing (0-3): 0.1, disease activity (0-3): 0.45, ESR: 35 mm/h; CHAQ > 0.5 3 patients, fever/rash 1 patient. Six patients were receiving corticosteroids. All patients met improvement criteria, 7 before the 3rd TCZ infusion. At 1 month after TCZ therapy began: active joints: 4, joints with limited motion: 3, wellbeing: 0.05, disease activity: 0.24, ESR: 3 mm/h; CHAQ > 0.5 1 patient, fever/rash 0 patient. During the treatment, 4 patients reduced dose (< 50%) of corticosteroids. No side effects were recorded.Tocilizumab seems to be effective and safe with a rapid onset of action in children with anti-TNF refractory JIA."}
+{"text": "KRAS genotype in L-L and other or multiple metastatic (O/MM) MCRC patients treated with the FIr-B/FOx regimen was retrospectively evaluated.Bevacizumab (BEV) plus triplet chemotherapy can increase efficacy of first-line treatment of metastatic colorectal cancer (MCRC), particularly integrated with secondary liver surgery in liver-limited (L-L) patients. The prognostic value of the KRAS codon 12 and 13 and BRAF mutations by SNaPshot and/or direct sequencing. Fit MCRC patients <75 years were consecutively treated with FIr-B/FOx regimen: weekly 12-h timed flat-infusion/5-fluorouracil (TFI 5-FU) 900 mg/m2, days 1, 2, 8, 9, 15, 16, 22 and 23; irinotecan (CPT-11) 160 mg/m2 plus BEV 5 mg/kg, days 1, 15; oxaliplatin (OXP) 80 mg/m2, days 8, 22; every 4 weeks. MCRC patients were classified as L-L and O/MM. Activity and efficacy were evaluated and compared using log-rank test.Tumoral and metastatic samples were screened for KRAS wild-type (53%), 28 KRAS mutant (47%). At 21.5 months median follow-up, objective response rate (ORR), progression-free survival (PFS) and overall survival (OS) were, respectively: KRAS wild-type 90%, 14 months, 38 months; KRAS mutant 67%, 11 months, 20 months. PFS and OS were not significantly different. PFS and OS were significantly different in L-L compared to O/MM evaluable patients. In KRAS wild-type patients, clinical outcome of 12 L-L compared to 18 O/MM was significantly different: PFS 21 versus 12 months and OS 47 versus 28 months, respectively. In KRAS mutant patients, the clinical outcome of 13 L-L compared to 14 O/MM was not significantly different: PFS 11 months equivalently and OS 39 versus 19 months, respectively.In all, 59 patients were evaluated: 31 KRAS genotype wild-type and mutant does not significantly affect different clinical outcomes for MCRC patients treated with the first-line FIr-B/FOx intensive regimen. KRAS wild-type patients with L-L disease may achieve a significantly prolonged clinical outcome due to integration with secondary liver surgery, with respect to KRAS mutant patients.The  KRAS wild-type metastatic colorectal cancer (MCRC), reported overlapping activity and efficacy in phase III trials, ranging between objective response rate (ORR) 39% to 68%, progression-free survival (PFS) 7.2 to 10.6 months, overall survival (OS) 19.9 to 26.1 months [et al. [Triplet regimens consisting of chemotherapeutic drugs, or doublets plus bevacizumab (BEV)  or cetuximab  monoclonal antibody) in EGFR-overexpressing and 1 months . Phase I [et al. , proposeRAS, BRAF, PIK3CA genes, or loss of tumor suppressor function of PTEN, resulting in continuous activation of the RAS-mitogen-activated protein kinase (MAPK) or phosphoinositide 3-kinase (PI3K) pathways, characterize most colorectal cancers (CRC) [KRAS mutations represent an early event in colorectal tumorigenesis [BRAF mutations, prevalently c.1799 T>A (V600E) mutation, characterize 4.7% to 8.7% of CRC [Gain-of-function mutations of rs (CRC) -9. KRAS igenesis ,11 and oigenesis ,13, c.35igenesis , and codigenesis . They imigenesis . BRAF mu% of CRC -20.KRAS wild-type and mutant MCRC patients treated with irinotecan, 5-fluorouracil and leucovorin (IFL) plus BEV were 27.7 and 19.9 months, respectively [KRAS or BRAF wild-type compared to KRAS or BRAF mutant genotype was not significantly different, even though the hazard ratio (HR) was 0.64 and 0.38, respectively. A significantly better prognosis was reported only when KRAS/BRAF wild-type patients were compared with patients harboring mutations in the KRAS or BRAF genes (HR 0.51) [KRAS wild-type genotype significantly predicts a favorable clinical outcome of anti-EGFR or anti-vascular endothelial growth factor (VEGF) drugs added to doublet chemotherapy [the KRAS mutant genotype, BEV addition to IFL significantly prolonged PFS up to 9.3 months, without increasing OS and activity, compared to IFL [Clinical outcome  according to wild-type and mutant genotype assesses the prognostic relevance of a specific biomarker, potentially including the predictive role of effectiveness of treatment strategies. In randomized studies, the predictive relevance of wild-type or mutant genotype can also be specifically assessed by comparing experimental and control arms. The reported median OS values of ectively ,21. The HR 0.51) . KRAS wiotherapy ,21-23. Id to IFL ,21.KRAS genotype in MCRC patients enrolled in a previously reported phase II study [Here, we report a retrospective exploratory analysis evaluating the prognostic value of the II study  and in aII study .MCRC patients were enrolled in a previously reported phase II study  and in tL-oxaliplatin (OXP) [2/day, over 12 h (from 10:00 pm to 10:00 am), days 1, 2, 8, 9, 15, 16, 22 and 23; CPT-11 , 160 mg/m2, days 1, 15; BEV , 5 mg/kg, days 1, 15; l-OXP , 80 mg/m2, days 8, 22; cycles every 4 weeks.The FIr-B/FOx regimen was developed from previously reported doublet and triplet chemotherapy schedules ,25, consin (OXP) : TFI 5-FKRAS and BRAF genetic analyses were performed on paraffin-embedded tissue blocks from the primary tumor and/or metastatic sites. Genotype status was assessed for KRAS codon 12 and 13 mutations and BRAF c.1799 T>A (V600E) mutation by SNaPshot\u00ae multiplex screening for KRAS mutations and KRAS/BRAF mutations in 36 and 32 samples, respectively [KRAS mutations in 23 samples and to confirm detected mutations. After treatment with xylene thyocyanate and selection of tumoral cell clusters, DNA was isolated using the RecoverAll\u2122 Total Nucleic Acid Isolation Kit for FFPE Tissues  according to manufacturer's instructions. When considering the contamination of tumoral samples by non-malignant cells, a KRAS mutation in the tumor was defined as appearance of a mutant peak with a height of at least one-third compared to the wild-type.ectively ,27; direKRAS exon 2 and BRAF exon 15 were simultaneously amplified by polymerase chain reaction (PCR) using specific primers and purified using NucleoSpin\u00ae Extract II kit . PCR-amplified DNA was analyzed using the ABI PRISM SNaPshot Multiplex kit  and five primers including an additional tail at their 5' end allowing their simultaneous detection. Sense primers allowing the extension at nucleotides KRAS c.34G, c.35G, c.37G, c.38G and BRAF c.1799T were used and a multiplex SNaPshot reaction was performed as reported [KRAS exon 2 sequencing was performed from PCR-amplified tumor DNA using the Big Dye V3.1 Terminator Kit . Labeled products were separated using an ABI Prism 3130xl Genetic Analyzer . Data were analyzed using the GeneMapper Analysis Software version 4.0 .SNaPshot multiplex assay was performed as elsewhere reported ,27. Briereported . KRAS exKRAS genotype on clinical outcome of MCRC patients treated with FIr-B/FOx as first-line treatment. Moreover, patients were classified according to involved metastatic sites, L-L and O/MM [KRAS wild-type and mutant MCRC patients. Patients with L-L metastases were evaluated at baseline and every three cycles of treatment by a multidisciplinary team, consisting of a medical oncologist, liver surgeon and radiologist, to dynamically evaluate resectability defined according to resectability categories previously reported [A retrospective analysis was planned to evaluate prognostic relevance of and O/MM , to evalreported . ResectiKRAS wild-type versus mutant, L-L versus O/MM, and KRAS wild-type L-L versus O/MM, and KRAS mutant L-L versus O/MM MCRC patients [Clinical criteria of activity and efficacy were ORR, PFS and OS. ORR was evaluated according to Response Evaluation Criteria In Solid Tumors (RECIST) criteria ; patholopatients .KRAS wild-type and mutant genotypes was 31 (53%) and 28 (47%), respectively  and 19 (61%); KRAS mutant, 13 (46%) and 15 (54%). Table KRAS mutations detected in 28 patients: codon 12, 24 patients (85.7%), specifically c.35 G>A 15 patients (53.5%), c.35 G>T 7 patients (25%), c.34 G>A and c.35 G>C, 1 patient each; codon 13, 4 patients (14.2%), c.38 G>A 3 patients (10.7%) and c.37_39 dupl, 1 patient. A total of 32 tumoral samples (54%) were also analyzed for BRAF and no BRAF mutation was detected; 18 out of 31 KRAS wild-type MCRC patients were KRAS and BRAF wild-type; 14 out of 28 KRAS mutant MCRC patients were BRAF wild-type. EGFR protein expression was positive in 35 patients (59%) and negative in 24 patients (41%): among KRAS wild-type patients, positive in 23 patients (74%) and negative in 8 patients (26%); among KRAS mutant patients, positive in 13 patients (40%) and negative in 15 patients (60%).A total of 59 tumoral samples of 64 enrolled MCRC patients 92%) were available: 46 primary tumors and 13 metastases . Demographic and baseline features of patients were representative of the overall phase II study population . We observed 27 objective responses: 23 partial responses (77%) and 4 complete responses (CRs) (13%); 2 stable diseases (7%); 1 progressive disease (3%). Disease control rate was 97% (95% CI 90 to 100). Liver metastasectomies were performed in 11 patients (35%), 10 out of 12 L-L patients (83%). Median PFS was 14 months (1+ to 69+ months), 25 events occurred. Median OS was 38 months (1+ to 69+ months), 17 events occurred. Among the 18 KRAS/BRAF wild-type patients, ORR was 83% (95% CI 69 to 97), median PFS was 13 months (4 to 44 months), median OS was 31 months (8 to 66+ months). Among 27 evaluable KRAS mutant patients, ORR was 67% (95% CI 49 to 85). We observed 18 objective responses: 17 partial responses (63%) and 1 CR (4%); 4 progressive diseases (16%). Disease control rate was 85% (95% CI 71 to 99). Liver metastasectomies were performed in 7 patients (25%) out of 13 L-L patients (54%). Median PFS was 11 months (1+ to 60+ months), 20 events occurred. Median OS was 20 months (1+ to 60+ months), 17 events occurred. Overall, R0 liver resections made up 13 out of 18 liver metastasectomies (72%). Pathologic CRs were obtained in 2 patients (11%), both KRAS mutant patients, harboring codon 12 mutations, c.35 G>T and c.34 G>A, with multiple liver-only metastases. In one KRAS wild-type patient with single liver associated with lung metastases, double metastatic resections were performed. KRAS wild-type compared with mutant patients did not show significantly different PFS nor OS, even if OS seems to be favorable in KRAS wild-type patients  with FIr-B/FOx according to extension of metastatic disease  in KRAS ts Table . Among 2ts Table .KRAS wild-type patients, ORR in 12 L-L and 18 O/MM patients were 100% and 80%, respectively. Overall activity was 100% (ten liver metastasectomies and two cCRs) in L-L and 17% (one liver plus lung metastasectomy and two cCRs) in O/MM patients, respectively. Significantly different clinical outcome was confirmed in L-L compared to O/MM, respectively  while no liver metastasectomy nor cCR was obtained in O/MM patients. The comparison of PFS and OS in KRAS mutant L-L and O/MM patients was not significantly different: median PFS 11 months (3 to 60+ months) versus 11 months (1+ to 37 months), respectively; median OS 39 months (8 to 60+ months) versus 19 months (1+ to 59+ months), respectively  significantly increased median PFS up to 9.3 months, while ORR was equivalent to doublet arm , and median OS increased up to 19.9 months, even if not significantly [In ectively ,21,31,32ectively ,23,31-33patients . In KRASficantly .KRAS wild-type and mutant MCRC patients, BEV addition to triplet chemotherapy, according to FIr-B/FOx schedule, reported high activity and efficacy: ORR 90% and 67%, median PFS 14 and 11 months, median OS 38 and 20 months, respectively. A similar clinical outcome was also obtained in KRAS/BRAF wild-type patients. Equivalent efficacy was reported with FOLFOXIRI/BEV regimen: ORR 82% and 71%, median PFS 13.6 and 12.6 months, respectively [In ectively .KRAS wild-type and mutant patients, even if not significantly, while they were equivalent in the FOLFOXIRI plus BEV study [KRAS wild-type and mutant patients, respectively [KRAS/BRAF wild-type patients compared with patients harboring mutations in the KRAS or BRAF genes (HR 0.51) [KRAS wild-type and mutant MCRC patients treated with BEV-containing chemotherapy failed to significantly differentiate prognosis, as in the present study. Thus, intensive regimens adding BEV to triplet chemotherapy can further increase activity and efficacy in KRAS wild-type and mutant patients. Randomized studies would be able to properly evaluate this.Median PFS and OS values of MCRC patients treated with FIr-B/FOx were different in EV study . BEV addectively ,21. SignHR 0.51) . Direct KRAS wild-type L-L patients, accounting for 20% of fit MCRC patients, could gain 100% overall activity with an integrated medical and surgical approach, due to performed liver metastasectomies and long-lasting cCRs; median PFS 21 months and OS 47 months. A significantly favorable prognosis was demonstrated in KRAS wild-type L-L compared to O/MM patients, even if this represents a retrospective, exploratory analysis in a small cohort of MCRC patients. Using neoadjuvant cetuximab with either FOLFOX6 or FOLFIRI for unresectable colorectal liver metastases, metastasectomies were performed in 38% and 30% patients, respectively [KRAS wild-type L-L MCRC patients. In KRAS mutant patients, prevalently harboring c.35 G>A transversion (53.5%), integrated medical and surgical treatment failed to significantly increase PFS and OS in L-L compared to O/MM patients: median PFS was equivalent (11 months), in spite of 54% performed liver metastasectomies in L-L patients; median OS was 39 and 19 months, respectively. These data should be further evaluated in a larger cohort of MCRC patients. A proper multidisciplinary treatment strategy for KRAS mutant patients, showing different aggressiveness [The high activity of triplet chemotherapy plus BEV regimens correlated with increased resection rate of liver metastases and pathologic CR, particularly in L-L MCRC patients ,3,4,6. Wectively . Chrono-ectively . Furthersiveness , sensitiKRAS wild-type and mutant genotypes do not significantly affect the clinical outcomes of MCRC patients treated with the first-line FIr-B/FOx intensive regimen. KRAS wild-type patients with L-L disease may achieve significantly greater benefit from integration with liver metastasectomies compared to O/MM metastatic extension, with respect to KRAS mutant patients. The present findings should be verified in prospective trials of multidisciplinary strategies comparing clinical outcome according to KRAS genotype in patients with L-L and O/MM disease.The authors declare that they have no competing interests.Conception and design: GB, ER. Provision of study materials of patients: GB, AD, GC. Collection and/or assembly of data: all authors. Data analysis and interpretation: GB, KC, ER. Manuscript writing: GB, ER. Final approval of manuscript: all authors.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1741-7015/10/135/prepub"}
+{"text": "The study investigated cardiovascular changes in adult-onset growth hormone deficiency (GHD) and showed that patients with adult-onset GHD have a left ventricular mass index (LVMi) at or below the lower limit of normal, which improves with one year of growth hormone replacement.GHD causes cardiovascular problems, with loss of cardiac response to exercise and increased cardiac mortality, however the underlying processes are poorly understood.Ten patients with adult-onset GHD and age- and sex-matched controls (n=10) underwent CMR. Patients underwent scans before disease treatment and at twelve months after treatment. Cardiac parameters were calculated and indexed to body surface area (BSA). The comparison between groups was done using Mann-U-Whitney test and within the group using Wilcoxon test. The data are presented as median values.2, p=0.315; end diastolic volume index (EDVi) 69.2 vs. 76.3 ml/m2, p=0.1655; end systolic volume index (ESVi) 24.8 vs. 28.3 ml/m2, p=0.3527 and ejection fraction (EF) 64 vs. 63 %, p=0.8197. However, patients had an LVMi beneath the lower limit of normal when compared to published normal ranges (55.4 - 74.0 g/m2).Patients with GHD did not have significantly different left ventricle (LV) mass or volumetric parameters from controls: LV mass index (LVMi) 55.0 vs. 56.6 g/m2, p=0.393), ESVi  and EF  in the GHD group and controls.There were no differences between right ventricular (RV) EDVi  achieving statistical significance compared to pre-treatment values (p=0.0156).Patients with GHD on GH treatment for 1 year showed an increase in median insulin-like growth factor I (IGF-I) SDS from -1.83 to +0.40 (p=0.0068). There was no correlation between LVMi and IGF-I SDS . At one year the median LVMi moved into the previously published normal range (55.0 to 63.0 g/mPatients with adult-onset GHD have an LVMi at or below the lower limit of normal, which improves with one year of growth hormone replacement.Supported by departmental grants by Pfizer and Novartis."}
+{"text": "However, sudden cardiac death may be the first clinical manifestation. Hence early detection of myocardial abnormalities before overt left ventricular (LV) dysfunction is warranted for a better risk stratification.Seventeen paucisymptomatic LM carries , 14 paucisymptomatic DCM patients  with mild LV dysfunction (ejection-fraction 40%-55%) and 12 healthy controls  underwent complete clinical, echocardiographic, biohumoral and contrast-enhanced CMR assessment. Cine CMR was used to derive LV volumes, mass and function, and post-contrast CMR to detect late gadolinium enhancement (LGE) as an index of gross fibrosis. Eight LM patients, all DCM patients and all normal controls underwent pre- and post-contrast myocardial T1 mapping with gadolinium partition coefficient calculation, as an index of interstitial fibrosis.LM carriers presented a similar LV end-diastolic volume  and slightly reduced ejection-fraction (61\u00b16% p<0.001) than controls , while in DCM patients LV end-diastolic volume (102\u00b117 ml/m2) and ejection-fraction (49\u00b15%) were worse than both controls and LM carriers (p<0.001). Moreover, DCM patients presented worse diastolic dysfunction , NT-proBNP plasma levels  and Doppler-estimated pulmonary pressure  than LM carriers. Six (43%) DCM patients presented DE, representing 9\u00b13% of LV mass; similarly, seven (42%) LM carriers had DE, but with a larger extension . In LM carries DE distribution was patchy (2 pts) or midwall (5 pt), similarly to DCM patients . GPC reached a steady state 5 minutes after gadolinium injection. In DCM patients GPC was slightly higher than controls , while in LM carriers it was much higher (0.45\u00b10.07) than controls (p<0.01). Even excluding hyperenhanced myocardium, in LM carriers GPC remained higher than controls, while in DCM patients only slightly higher than controls.Myocardial fibrosis occurs early in LM carrier before LV dilatation and dysfunction and may represent an early phenotypic expression of cardiac involvement."}
+{"text": "Bloodstream infections (BSI) are important causes of morbidity and mortality. Most of all, when are caused by drug-resistant organisms (DR).To investigate the epidemiology, etiology, systemic response and treatment of DR-BSI.EPINE-EPPS project. Comparisons between groups were performed by means of the X2 test for categorical variables or analysis of variances (ANOVA) for continuous variables.A retrospective study was conducted about all BSI diagnosed in a secondary hospital during one year. The pattern resistant pathogen study was We included 60 patients  with 63 DR-BSI of which 71.5% were nosocomial and healthcare-associated BSI.Unknown and intravascular catheter-related DR-BSI accounted for 49.2% of cases. Among secundary infections, the source was 37.5% urinary track, 31.2% intra-abdominal and 15.6% respiratory track infections.S. aureus. Among DR-Gram-negative bacilli, 12.2% of enterobacteracea family produced extended-spectrum B-lactamasas. We found 5 DR-BSI caused by Acitetobacter carbapenem resistant and 3 DR-BSI by P. aeruginosa carbapenem resistant.Overall DR-BSI, DR-Gram-positive cocci were 55.6%. The most common isolated pathogens were staphylococcus coagulase-negative and Median time to diagnosis for DR-Nosocomial BSI was 14 days (IQR), 7-35 days after hospital admission. For Gram-negative was 11 days (7.5-31.5) and for Gram-positive 19 days (7-29).Only 31.7% of DR-BSI received appropriate initial empirical antimicrobial therapy versus 73.5% of non DR-BSI (p<0.001). More than one third (36.5%) of the episodes occur with significant systemic response (severe sepsis or septic shock). The crude mortality rate was 25.4 % (p<0.001). If the patient developed severe sepsis or septic shock crude mortality rose to 52.2%.Information about local epidemiology is important to develop prevention and control strategies in drug\u2013resistant microorganism and to improve the management of BSI.None declared"}
+{"text": "Remarkably, while glucose potently stimulated GIP release, fructose was without effect. Similar patterns were found in the mouse and rat, with both fructose and glucose stimulating GLP-1 secretion, whereas only glucose caused GIP secretion. In GLUTag cells, a murine cell line used as model for L cells, fructose was metabolized and stimulated GLP-1 secretion dose-dependently (EC50 = 0.155 mM) by ATP-sensitive potassium channel closure and cell depolarization. Because fructose elicits GLP-1 secretion without simultaneous release of glucagonotropic GIP, the pathways underlying fructose-stimulated GLP-1 release might be useful targets for type 2 diabetes mellitus and obesity drug development.Nutrients often stimulate gut hormone secretion, but the effects of fructose are incompletely understood. We studied the effects of fructose on a number of gut hormones with particular focus on glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP). In healthy humans, fructose intake caused a rise in blood glucose and plasma insulin and GLP-1, albeit to a lower degree than isocaloric glucose. Cholecystokinin secretion was stimulated similarly by both carbohydrates, but neither peptide YY While it has long been known that the presence of nutrients such as glucose, fat, and protein in the intestinal lumen stimulates secretion of many gut hormones, the effects of fructose are poorly understood. While some studies have shown that fructose does not stimulate insulin secretion in: 1) humans when perfused intraduodenally, 2) in rat pancreatic preparations, or 3) in isolated human and rat pancreatic islets  participated in the study, which was approved by the local ethical committee and conducted according to the principles of The Helsinki Declaration. All subjects had normal fasting blood glucose levels , and none had parents or siblings diagnosed with any type of diabetes. No subjects received medication known to interfere with glucose homeostasis. Each subject was studied on two occasions within 3 wk after the first day of study. Subjects were instructed to refrain from vigorous exercise and alcohol for at least 24 h before each study. Study days began at 0830 preceded by a 10-h overnight fast. Venous blood samples were collected at time \u221210, 0, 15, 30, 45, 60, 90, 120 min in prechilled EDTA (10.8 mg) tubes  through a polyethylene cannula placed in a cubital vein. At time 0 min, the seated, single-blinded, subjects drank a sugar solution containing 75 g fructose or glucose dissolved in 300 ml water (RT) within 2 min. For palatability, solutions were refreshed with lemon juice. Upon collection, blood samples were instantly chilled on ice and centrifuged  within 30 min. Plasma were stored at \u221220\u00b0C until analysis.Nine healthy volunteers  (data not shown). Rats were divided into weight-matched groups (n = 4/occasion) and given a bolus of either glucose or fructose  diluted in milli-Q water to a final concentration of 50% (wt/vol) or a matched volume vehicle (milli-Q water). Rats from the same cage received different treatments. Blood (200 \u03bcl) was collected into prechilled EDTA-coated capillary tubes  by sublingual vein puncture and instantly transferred onto ice. The zero sample was collected 10 min before bolus administration. At time 0 min rats were stimulated with either vehicle, glucose, or fructose, and blood was collected at time 15, 30, and 60 min. Blood glucose was measured instantly after collection, and samples were centrifuged  to obtain plasma, which was transferred to fresh Eppendorf tubes and immediately frozen on dry ice. Samples were stored at \u221220\u00b0C until analysis. At the end of the experiment rats were anesthetized by subcutaneous injection of hypnorm/midazolam  and killed by diaphragm perforation.Studies were approved by the Danish Animal Experiments Inspectorate (2013-15-2934-00833). Male Wistar rats were obtained from Taconic  and allowed at least 1 wk of acclimatization before the day of experiment. Rats followed a 12:12-h light-dark cycle with ad libitum access to standard chow and drinking water. Every day for a week up to the experiment, rats were handled to minimize stress-related responses on the day of study. Handling included restraint and gavage feeding with drinking water. Experiments were carried out on two occasions on nonfasted rats (294.9 \u00b1 3.8 g) just before their nocturnal feeding period (1700). Weight did not differ between groups or study occasions (time \u221230 min and again at time 0 min with fructose or glucose (3 g/kg) diluted in drinking water to a final concentration of 20% (wt/vol). At time 0 and 6 min, blood (75 \u03bcl) for hormone measurement was collected into prechilled EDTA-coated capillary tubes  supplemented with dipeptidyl peptidase 4 inhibitor  . After collection, blood samples were immediately transferred to Eppendorf tubes supplemented with 7.5 \u03bcl diprotinin A (1 mM) and centrifuged  within 30 min. Plasma was transferred to fresh Eppendorf tubes, snap-frozen on dry ice, and stored at \u221220\u00b0C until analysis. Blood for glucose measurements (3 \u03bcl) was obtained by tail puncture immediately after blood sampling and directly processed for blood glucose measurements as described below. After the first day of study (13 wk), the procedure was repeated. At both occasions, mice were randomly divided into glucose- and fructose-receiving groups. Weights did not differ between fructose and glucose groups at the respective days of study . At the end of the experiment, mice were killed by cervical dislocation.Studies were carried out with permission from the Danish Animal Experiments Inspectorate (2012-15-2934-00207). Female C57BL/6 mice were obtained from Charles River  and allowed 1 wk of acclimatization before the day of experiment. Mice  were fasted overnight but allowed free access to water. Mice were prestimulated with a gavage of 100 \u03bcl drinking water at 2 until 70\u201380% confluent, then trypsinized and plated  on a 24-well plate precoated with matrigel (1:100)  as described previously  supplemented with 0.1% (wt/vol) fatty acid-free BSA  and incubated for 2 h  with 250 \u03bcl test reagents  dissolved in saline buffer. All reagents were supplied by Sigma Aldrich. Supernatants were collected and centrifuged  to remove floating cells and debris and then either snap-frozen on dry ice and stored at \u221280\u00b0C or directly processed for GLP-1active measurement.GLUTag cells were cultured in T75 cell culture flasks in low-glucose (1.0 g glucose/l) DMEM 5564 medium  supplemented with 10% (vol/vol) FCS, 1% (vol/vol) penicillin/streptomycin, and 1% (vol/vol) glutamine. Cells were incubated at 37\u00b0C, 5% COeviously . The folIntracellular NAD(P)H concentration levels were measured using NAD(P)H's autofluorescence properties , 23. GLUtotal (GIP1\u201342 and GIP3\u201342) was measured with GIP-ELISA , whereas GLP-1 levels were measured by the same Millipore GLP-1 (active) ELISA kit as for the GLUTag cells. In both cases, data were acquired using a Biotek EL800 plate reader controlled by Gene5 software . For the mouse, GIPtotal concentrations were measured as described above, and GLP-1 levels were determined by the Meso Scale assay system . Data were obtained with a Sector Imager 2400 device controlled by MSD Discovery workbench software. Human plasma concentrations of total GLP-1, GIP, insulin (INS), glucagon (GCG), and intact neutrotensin1\u201313 (NTS1\u201313) were determined using in-house radioimmunoassays  with 125I-labeled tracers and hormone-specific antiserum, recognizing COOH-terminal epitopes, as described previously . Free and bound moieties were separated by addition of (bovine) plasma-coated charcoal . Supernatants were collected , and the radioactivity was counted in a Packard Cobra II Gamma Counter (GMI). Plasma polypeptide YY3\u201336 (PYY3\u201336) levels were measured by a commercially available RIA kit . Plasma cholecystokinin (CCK) concentrations were measured by RIA measuring all the tyrosyl-sulfated forms of CCK as described  read on a SpectraMax M5 plate reader controlled by SoftMax Pro 5.4.1 software . For both human, rat and mouse, blood glucose levels were measured by the glucose oxidase method immediately after sampling by a handheld Accu-chek Compact plus device . For the rat, plasma GIPescribed . For allt-test, paired Student's t-test, or one-sample t-test, testing values against a hypothetical value of zero, as indicated. P < 0.05 was considered significant. For the human study, baseline values are presented as the mean of the \u221215 and 0 min levels. EC50 value for fructose-stimulated GLP-1 secretion from GLUTag cells  (P < 0.001) with significantly higher area under the curve (AUC) values after oral glucose vs. fructose  . For both groups, blood glucose concentrations returned to basal levels by the end of the study period (2 h)  A. Both t, n = 9) B. Maximuod (2 h) A.P > 0.05, n = 9). There was a rise in insulin after both glucose and fructose ingestion , albeit glucose elicited a higher maximal response  (P < 0.05) (P < 0.001)  C. Insuli < 0.05) D. Insuli< 0.001) C.P > 0.05) (P = 0.17) (Glucagon levels did not change significantly at any time point after glucose or fructose intake ( > 0.05) E. Glucag = 0.17) F.n = 9)  with progressively increasing concentrations from baseline until 45 min (P < 0.001). Levels then tended to fall, reaching an elevated plateau phase from 60 to 120 min. GLP-1 also rose after oral glucose intake , reaching significantly elevated concentrations at 15 min (P < 0.01). Thereafter, levels remained elevated until 90 min (P < 0.05)  A. GLP-1 l 90 min A. Over t < 0.05) B.P > 0.05, n = 9) , reaching at plateau from 15 to 120 min , fructose did not significantly elevate GIP concentrations , also compared with basal levels (P > 0.05) (P < 0.0001)  C. While  > 0.05) C. GIP AU 0.0001) D.P > 0.05, n = 9) , but levels were slightly elevated at 60 and 90 min after fructose intake  (P > 0.05)  E. PYY co, n = 9) E. Plasma > 0.05) F.P > 0.05, n = 9)  with peak values occurring 15 min after ingestion for both treatments . Thereafter, CCK concentrations decreased for both treatments, albeit with a tendency toward higher values in the glucose treatment group at later time points (P > 0.05)  G. Glucose points G. CCK AU > 0.05) H.P > 0.05, n = 9) (P < 0.001), with similar peak values observed at 15 min for both treatments . NTS AUCs did not differ between treatments (P > 0.05)  I. Glucos > 0.05) J.P < 0.01, n = 7). At 60 min, blood glucose levels had normalized and were not different from basal levels (P > 0.05)  but not in the fructose-treated group   . In contrast, fructose stimulation did not elevate GIPtotal levels at any time points  (P < 0.001), but not after fructose , but GLP-1 AUCs were elevated by both glucose and fructose compared with control (P < 0.05)  A. Blood , n = 6) B. Blood , n = 6) A. GIPtot, n = 6) C. GIP AUfructose D. GLP-1  < 0.05) F.P < 0.05, n = 31) albeit glucose stimulation increased levels to a greater extent than fructose (P < 0.0001)  (P > 0.05). GIPtotal concentrations were elevated by glucose treatment , but fructose had no effect  (P > 0.05) (C).Responses to glucose and fructose were studied on two occasions separated by 13 wk. Blood glucose increased significantly for both groups compared with basal levels  A. GLP-1t n = 23) B to a si n = 23) C. Basal  > 0.05) , A\u2013C.P < 0.05, n = 43)  (P < 0.0001) in a dose-dependent manner, reaching a maximal response at 1 mM  . Addition of gliclazide (GLI) to induce ATP-sensitive potassium (KATP) channel closure did not potentiate fructose-induced GLP-1 secretion , but stimulation with GLI alone caused GLP-1 secretion to a similar degree as tolbutamide  (ATP channel opener diazoxide (340 \u03bcM) completely abolished fructose-induced GLP-1 secretion , which reached similar levels as during diazoxide exposure alone  (Metabolic responses to fructose were investigated by the NAD(P)H imaging technique. Representative traces from an experiment are shown in  n = 43) B. Stimul n = 43)  confirme n = 27) C. Fructo = 6\u201316) D. Again,= 14\u201315) F. Presen= 10\u201312) E.1\u201313 to a similar extent as glucose, and also enhanced insulin secretion, albeit to a lower degree than glucose. In contrast, neither fructose nor glucose altered glucagon or PYY3\u201336 levels. With a view to perform mechanistic studies regarding the differential responses of the incretin hormones, we also investigated GIP and GLP-1 responses to fructose in mice and rats. In both animals, fructose stimulated GLP-1 secretion to the same extent as isocaloric glucose, but, similar to humans, GIP secretion was not stimulated.The increased intake of dietary fructose in general and high-fructose corn syrup in particular has been suggested to play a role in the current obesity epidemic , 20. MorIn mice and humans, fructose intake also increased blood glucose levels, and a similar tendency was seen in the rat. It is generally accepted that fructose can be converted to glucose in the liver by mechanisms involving rapid phosphorylation of fructose to fructose 1-phosphate by fructose kinase, which by liver-type-B aldolase activity, yields the intermediate metabolites dihydroxyacetone phosphate and glyceraldehyde, with the latter being a substrate for glucose production by further metabolic activity. However, it has also been shown that the small intestine has capacity to transform fructose into glucose by the same pathway . AlthougATP channel closure, causing cell membrane depolarization  and a grant to J. J Holst from the Novo Nordisk Centre for Basic Metabolic Metabolism .The study was supported by a The authors declare no financial or other conflicts of interest.Author contributions: R.E.K., F.M.G., B.H., F.R., J.A.W., and J.J.H. conception and design of research; R.E.K., B.H., J.A.W., and J.F.R. performed experiments; R.E.K., B.H., and J.F.R. analyzed data; R.E.K., F.M.G., B.H., F.R., and J.J.H. interpreted results of experiments; R.E.K. prepared figures; R.E.K. drafted manuscript; R.E.K., F.M.G., B.H., F.R., J.A.W., and J.J.H. edited and revised manuscript; R.E.K., F.M.G., B.H., J.A.W., J.F.R., and J.J.H. approved final version of manuscript."}
+{"text": "Objective(s): In the current study, the effects of selected folk medicinal herbs were evaluated in D-galactose-induced aging in male mice.Materials and Methods: Male BALB/c mice were randomly divided into 12 groups composing sham, control, and treated groups. Aging was induced by administration of D-galactose (500 mg/kg/day for 6 weeks). A positive control group was assigned that received vitamin E (200 mg/kg/day). The extract of herbs was prepared, lyophilized, and used in this study. The herbs were administered by gavage for 4 weeks to D-galactose-aged animals at the selected doses (mg/kg/day) as follows: Zingiber officinale (250), Glycyrrhiza glabra (150), Rosmarinus officinalis (300), Peganum harmala (50), Aloe vera (150), Satureja hortensis (200), Teucrium scordium (200), Hypericum perforatum (135) and Silybum marianum (150). One group of animals was assigned as sham and not given D-galactose. Results: At the end of treatment, pro-inflammatory markers including tumor necrosis factor-\u03b1 (TNF-\u03b1), interlukine-1\u03b2 (IL-\u03b2), interlukine-6 (IL-6), NF-kappaB (NF-\u03bab), total antioxidant power (TAP), thiobarbituric acid reactive substances (TBARS) as lipid peroxidation (LPO) marker and male sex hormones i.e. testosterone and dehydroepiandrosterone-sulfate (DHEA-S) were measured in the blood.Conclusion: These data for the first time indicate significant anti-aging potential of examined herbs. Results showed that D-galactose induces a significant oxidative stress and promotes proinflammatory cascade of aging while all herbs more or less recovered these changes. Among 9 herbal extracts, Silybum marianum showed the best effect in restoring aging changes. Aging as a complex of natural circumstance is exhibited by an augmentation in the chance of illness and finally death. Although some theories have been proposed as the mechanisms of aging but the one relating aging and cellular oxidative stress have received more supports. Therefore, it can be said that reduced sex hormones and augmented quantity of oxidative stress parameters or inflammatory cytokines are main biochemical manifestations of aging -2. In acOne of the problems in testing anti-aging compounds is lack of suitable animal models. Although several models have been used so far but among them, typical mouse D-galactose-induced model of aging is the best one that gives closer results to clinical studies. D-galactose is a sugar that at higher levels converts to aldose and hydro-peroxide during the catalysis of galactose oxidase, culminated in the generation of free radicals . These mMany scientists and pharmaceutical companies try to develop a drug to reduce speed of human aging but no effective drug has been discovered yet. In the last decade the importance of folk medicine and herbal medicines have been revisited that resulted in developing many effective drugs for many human diseases. For instance, in the recent years, efficacy of herbal medicines in diseases like inflammatory bowel diseases -10, obesZ. officinale, G. glabra, R. officinalis, P. harmala, A. vera,S. hortensis, T. scor-dium, H. perforatum and S. marianum to test in D-galactose-induced model of mouse aging.We recently proved anti-aging potential of naturally-based drugs like IMOD and Angipars which have strong antioxidant power . On the Chemicals3-6H2O), D-galactose, and vitamin E (Trolox) were purchased from Merck (Germany). Rat specific tumor necrosis factor-\u03b1 (TNF-\u03b1), interlukine-1\u03b2 (IL-\u03b2), interlukine-6 (IL-6), NF-kappa B (NF-\u03bab) ELISA kits were purchased from BenderMed Systems (Austria). Testosterone and dehydroepiandrosterone ELISA kits were purchased from Dia Metra . Thiobarbituric acid (TBA), trichloroacetic acid (TCA), n-butanol, hexadecyltrimethyl ammonium bromide (HETAB), tri (2-pyridyl)-s-triazine (TPTZ), HCl, malondialdehyde (MDA), ferric chloride  were washed in a suitable bactericide (chlorh-exidine). The filets were grounded to a liquid, and the pulp was removed by filtering. The resultant gel was then freeze dried.Herbs were provided from the Research Institute of Medicinal Plants Karaj during June 2009 and were air-dried at room temperature. Samples were authenticated by a botanist (Y. Ajani), and voucher specimens were preserved in the central herbarium of medicinal plants (RIMP). The scientific names and tested parts of the herbal materials are detailed in AnimalsMale BALB/c mice  were provided from Tehran University of Medical Sciences (TUMS) animal house. The animals were housed in standard polypropylene cages with wired-net top in a controlled room  and were allowed free access to standard laboratory pellet diet and water during the experiments. All ethical issues on the use of animals were carefully considered and the study protocol was approved by TUMS review board with code number of 90-03-33-15668.Experimental designth group of animals was the sham group which was not given D-galactose. After 2 weeks, the 11 groups which had been given D-galactose were randomly divided into aging control group , positive control group  and herb-treated groups including 9 groups that each received 500 mg/kg D-galactose per 1 ml drinking water plus Z. officinale (250 mg/kg/day), G. glabra (150 mg/kg/day), R. officinalis (300 mg/kg/day), P. harmala (50 mg/kg/day), A. vera (150 mg/kg/day), S. hortensis (200 mg/kg/day), T. scordium (200 mg/kg/day), H. perforatum (135 mg/kg/day) and S. marianum (150 mg/kg/day), respectively by gavage for 4 weeks , lipid peroxidation (LPO) and male sex hormones including testosterone and dehydroepiandrosterone-sulfate (DHEA-S) were measured in the serum.Measurement of LPO 2SO4 (0.05 M). After addition of TBA (0.2% in sodium sulfate), the sample was heated for 30 min in a boiling water bath. Then, TBA reactive substances (TBARS) as LPO marker adducts were extracted by n-butanol and absorbance was measured at 532 nm as described in details in our previous work (LPO was measured by the reaction of thiobarbituric acid (TBA) with lipid peroxides. Samples were mixed with TCA (20%) and the precipitate was dispersed in Hous work . Data weMeasurement of TNF-\u03b1, IL-1\u03b2, IL-6 and NF-\u03babQuantitative detection of TNF-\u03b1, IL-1\u03b2, IL-6 and NF-\u03bab levels in serum were performed using an enzyme-linked immunosorbent assay rat specific ELISA kit according to each specific brochure. The absorbance of the final colored product was measured in 450 nm as the primary wave length and 620 nm as the reference wave length. TNF-\u03b1, IL-1\u03b2, IL-6 and NF-\u03bab levels were expressed as pg/mg.Measurement of TAP 3+ to Fe2+. Interaction of TPTZ with Fe2+ results in formation of a blue color with a maximum absorbance at 593. The whole procedure has been described in our previous study . Data were analyzed by one-way ANOVA followed by Tukey post-hoc test for multiple comparisons to ensure the variances of the data are distributed properly. A P< 0.05) and a significant decrease in TAP (P<0.05) were observed when sham group was compared with D-galactose-received aged group. P<0.05; 1.2\u00b10.05 vs. 2.5\u00b10.33, P<0.05; 27\u00b13.9 vs. 49.66\u00b13.4, P<0.05; 45.7\u00b12.4 vs. 97\u00b121.2, P<0.05). As shown in P<0.05; 1.2\u00b10.2 vs. 0.6\u00b10.08, P<0.05) in aged mice was lower than that in the sham.A significant increase in TBARS , and increasing TAP , , respectively. As shown in Z. officinale increased testosterone and DHEA-S  in aged mice.P<0.05), , 2. FiguEffects of G. glabra in aged miceG. glabra .G. glabra recovered D-galactose-induced increase in TNF-\u03b1, IL-6, IL-1\u03b2 and NF-kB , respectively. As shown in G. glabra increased testosterone and DHEA-S levels  in aged mice.D-galactose-induced elevation of TBARS and reduction of TAP , 2 were Effects of R. officinalis in aged miceR. officinalis treatment recovered D-galactose-induced rats by reducing TBARS , and increasing TAP  , respectively. As shown in R. officinalis increased testosterone,  and DHEA-S  in aged mice. P<0.05) , 2. FiguEffects of P. harmala in aged miceP. harmala . P. harmala recovered D-galactose-induced increase in TNF-\u03b1, IL-6, IL-1\u03b2, and NF-kB , respectively. As shown in P. harmala increased testosterone and DHEA-S  in aged mice.D-galactose-induced elevation of TBARS and reduction of TAP , 2 were Effects of A. vera in aged miceA. vera treatment recovered D-galactose-induced elevation of TBARS (P<0.05), and improved reduction of TAP (P< 0.05). A. vera recovered D-galactose-induced increase in TNF-\u03b1, IL-6, IL-1\u03b2, and NF-kB , respectively. As shown in A. vera increased testosterone and DHEA-S levels  in aged mice.Effects of S. hortensis in aged miceS. hortensis treatment recovered D-galactose-induced elevation of TBARS (P<0.05), and increased TAP (P<0.05). S. hortensis recovered D-galactose-induced increase in TNF-\u03b1, IL-6, IL-1\u03b2, and NF-kB , respectively. As shown in S. hortensis increased testosterone and DHEA-S  in aged mice.Effects of T. scordium in aged miceT. scordium . T. scordium recovered D-galactose-induced increase in TNF-\u03b1, IL-6, IL-1\u03b2, and NF-kB , respectively. As shown in T. scordium increased testosterone and DHEA-S  in aged mice.D-galactose-induced elevation of TBARS and reduction of TAP , 2 were Effects of H. perforatum in aged miceH. perforatum treatment recovered D-galactose-induced rats by reducing TBARS  and increasing TAP  , respectively. As shown in H. perforatum increased testosterone and DHEA-S  in aged mice. P<0.05) , 2. FiguEffects of S. marianum in aged miceS. marianum . S. marianum recovered D-galactose-induced increase in TNF-\u03b1, IL-6, IL-1\u03b2, and NF-kB , respectively. As shown in S. marianum recovered D-galactose-induced reduction of testosterone and DHEA-S  in aged mice.D-galactose-induced elevation of TBARS and reduction of TAP , 2 were Effects of vitamin E in aged miceP<0.05; 190\u00b113.1 vs. 120\u00b17.5, P<0.05). P<0.05; 1.4\u00b10.26 vs. 2.5\u00b10.33, P<0.05; 30.1\u00b12.2 vs. 49.66\u00b13.4, P<0.05; 57\u00b13.9 vs. 97\u00b121.2, P<0.05), respectively. As shown in P<0.05; 1\u00b10.16 vs. 0.6\u00b10.08, P<0.05) in aged mice.D-galactose-induced elevation of TBARS and reduction in TAP , 2 were In this study, for the first time, we analyzed the anti-aging potentials of nine famous herbs in a well-setup animal aging model using chronic administration of D-galactose. Our results showed that production of free radicals is the principal reason of up-regulation of pro-inflammatory cytokines and the main determinant involved in the D-galactose-induced aging model. Furthermore, these herbs dramatically diminished oxidative stress and proinflammatory cytokines in the aged mice. Supporting the mechanism of action of these herbs and the theory of oxidative stress in aging, vitamin E was used as the standard and showed the similar effects in examined markers of aging.Adapted from corresponding author\u2019s previous paper published in open access source .Interestingly, present results indicated improve-ment of testosterone and DHEA-S by herbs in the aged mice. Decline of steroid hormones with aging is already known and is believed a major contributor to elevation of pro-inflammatory markers .Recent studies have shown the mechanisms of action of anti-aging herbs in reducing aging process that is divided into four categories including anti-oxidant, anti-inflammatory, effect on memory/cog-nition/mood, and the sex hormones . This inGinger [Zingiber officinale Roscoe (Zingiberaceae)] and supplements derived from ginger like zingerone, shogaols and gingerols posse the abilities for the treatment of chronic inflammation. The protective effects of Z. officinale in lessening macromolecular damage in aged mice were shown in this study. Besides, recent study has shown that ginger extracts owns antioxidant activity  or licorice is the root of G. glabra from which a sweet flavor can be extracted. The results of this study showed that G. glabra has the protective effects in declining macromolecular damage in aged mice. It has been shown that G. glabra extract is the safest pigment-lightening agent with the fewest side effects  (Fam. Liliaceace) are the source of aloe vera gel. A. vera gel is greatly used in cosmetics and toiletries for its moisturizing and regenerating action. Also, the leaf of A. vera could assist cellular repairing, imbibition of foods, vitamins, minerals and vital nutrients  comprises 12 species, which possess antioxidant, anti-spasmodic, anti-nociceptive and anti-inflammatory properties -b-farnesene as the major components of T. scordium  Gaertn. . Interestingly, the present findings confirmed that S. marianum causes the best effects in improving antioxidant, anti-inflammatory and male sex hormones in aged mice. This effect is so important and should be considered as an advantage. This can be explained with current knowledge that among many medicinal plants, S. marianum, has been greatly used for centuries as a natural popular complementary medicine for the treatment of several diseases. The main indications for the use of silymarin are related to the hepatoprotection , a flavonoids complex known as \u2018milk thistle\u2019 is extracted from the fruit of otection , 63. Alsotection , 65. It otection . Excitinal cells . S. marianum showed the best effect in improving all the D-galactose-induced aging effects. Since all of the selected and examined herbs are already found safe in human and there are good information from traditional medicine, therefore, they can be supplemented into the diet of elderly people to reduce speed of aging. Testing the mixture of these herbs together or with other anti-aging products is among the plans of future.Taking collectively, the present results confirmed our hypothesis that the herbs with highest antioxidant power may reduce speed and rate of aging as evidenced by recovery of proinflammatory cytokines and sex hormones. Among tested herbs,"}
+{"text": "It has been reported in Mexico that genotypes H and G are the most common, although genotypes prevalence in HIV co-infected patients is unknown.We estimated the prevalence and identified the resistance pattern of HBV/H and G genotypes in HIV co-infected patients and compared them in mono-infected HBV patients.A cross-sectional prevalence and analytic study were realized. Risk factors, HIV or HCV co-infections, antiretroviral therapy (ART) experience, HBsAg, HBeAg, HBV viral load and mutations genetic analysis were collected; CD4+ cells count from HIV co-infected patients and HIV viral load were measured. We calculated the prevalence and exact 95% binomial confidence interval as well the Odds ratios (OR) and 95% confidence intervals to assess the relationship between risks factors and the risk of having HBV/H or G genotype.We enrolled 84 patients, 72 men and 12 women with 41 HIV co-infected patients. The distribution of HBV genotypes was: HBV/H 56 (66%), HBV/G 22 (26.1% [95% CI 17% to 36%]), HBV/F 4 (4.7%) and HBV/A 2 (2.3%). The most frequent mutations presented in 9 HIV co-infected patients and one mono-infected patient with ART experience were rtM204V and seven of them showed genotype G (7/9). Mono-infected HBV patients exposed more probability to HBV/H genotype than co-infected HIV patients OR 13.0 (CI 95% 3.40-49.79), P=0.0001 In contrast co-infected patients presented less possibility to have genotype H, 0.56 (CI 95% 0.42-0.75).Our results suggest that HBV/G genotype predominates in co-infected patients. As well, rtM204V and rtL180M mutations are common in HBV/HIV co-infected patients with genotype G and ART experience."}
+{"text": "Urinary tract infections are the main indication for antimicrobials in elderly.Despite the treat for resistance dissemination and therapy failure, clinicians are seldom informed on the per patient evolution of antimicrobial resistance in pathogens.Laboratory results were obtained from 13 voluntary diagnostic laboratories  in Belgium during the year 2005. Susceptibility profiles were done by Kirby Bauer disk diffusion according to CLSI. The first two urine samples from patients older than 65 year were included.E. coli (7188/1654), E. faecalis (1282/403), P. mirabilis (1230/313), K. pneumonia (673/173), P. aeruginosa (293/120), E. aerogenes (375/203), S. aureus (158/54), M. morganii (347/89), Group B streptococci (149/31), C. freundii complex (101/29). When comparing first versus second samples antibiograms for E. coli, a decrease in susceptibility was found for the following antimicrobial agents: cotrimoxazole -6.9%; nitrofurantoin -2.8%, fosfomycin 0.0%; ciprofloxacin -10.8%; cefuroxime -5.6%; amoxicillin-clavulanic acid -5.6%; ampicillin -10.5%. For E. faecalis, marked decreases were found for nitrofurantoin -2.4%; fosfomycin -2.2%; -ciprofloxacin -10.3%; and only mild decreases for amoxicillin-clavulanic acid 0.0%; and ampicillin -1.2%. For K. pneumoniae decreases were in the range of -2.9 to -4.1% for cotrimoxazole, ciprofloxacine, cefuroxime and amoxicillin-clavulanic acid, and was -12.4% for nitrofurantoin. For S. aureus and C. freundii no decrease (<-0.1%) was seen for nitrofurantoin and fosfomycin. For E. aerogenes, decreases of -18 and -12.5% were found for cotrimoxazole and fosfomycin, respectively. M. morganii showed in consecutive samples less susceptibility for cotrimoxazole (-16.2%), fosfomycine (-13.0%) and ciprofloxacin (-10.5%), while only a marginal decrease was found for nitrofurantoin (-0.5%).Following organisms were predominantly isolated (N first samples/N second samples): The resistance selection influence of consecutive samples depends on the antibiotic-bacterium combinations, and thus might be taken into account when empiric therapy guidelines for urinary tract infections in elderly are reviewed.None declared"}
+{"text": "In patients undergoing atrial fibrillation (AF) procedures, imaging of the left atrium (LA) and pulmonary veins (PV) is important for pre-ablation planning and to identify post ablation complications. Conventional MRA protocols use first-pass, non-gated sequences that require long breath-holds. Quality of non-gated MRA's can be challenging in sick or sedated patients. We developed a novel ECG-gated, respiratory navigated MRA sequence less dependent on patient compliance, which yields better clarity of LA anatomy.Eighty patients with AF underwent either conventional non-gated (n=40) vs. novel ECG-gated (n=40) MRA on a 3T Verio scanner . All MRA's were performed using 0.1 mmol/kg Multihance . A novel ECG-gated, respiratory navigated MRA was developed using 3D saturation recovery prepared, GRE sequence with fast contrast injection  followed by slow infusion . Saturation pulse was applied every heart beat and fat saturation was applied immediately before data acquisition during LA diastole. Additional scan parameters were: axial imaging volume, FOV =400x400x110, voxel size=1.25x1.25x2.5 mm, TR/TE=2.8/1.3ms, flip angle=15 degrees, TI=250ms, phase encoding direction: left to right. Typical scan time was 3-5 minutes.Conventional non-gated MRA were performed with contrast injection rate 2.0 mL/sec and continuous data acquisition during single breath-hold (14 sec.). Scan parameters included: axial imaging volume, FOV=400x263x120 mm, voxel size=1.25x1.25x2.5, TR/TE=2.8/1.1ms, flip angle=27 degrees.All MRA's were randomized and quality scores were determined by two experienced readers for contrast enhancement, border sharpness, and chamber detail of the PV's, LA, and LA appendage (LAA) .  in the non-gated cohort. Border sharpness scored 3.08 vs. 2.35 QS  respectively. Chamber detail was also assessed with specific anatomical positions, which all yielded superiority of ECG-gated MRA ; left atrial appendage 3.11 vs. 2.25 QS ; left atrium 3.08 vs. 2.35 QS .ECG-gated MRA improves image quality with better border sharpness and anatomical detail compared to conventional non-gated MRA. These advantages may translate into improved diagnostics in AF patients including detection of PV stenosis or following LA remodeling post-ablation.No disclosures."}
+{"text": "AbstractStenohya Beier, 1967 are described from China: Stenohya pengaesp. n.  and Stenohya huangisp. n. .The presence of Stenohya pengaesp. n. in the tree crown of Castanopsis fabri represents a new habitat for Neobisiidae. A key and a distribution map of the Chinese Stenohya species are also provided.Two new species of the genus  Stenohya Beier, 1967 is a small Asian pseudoscorpion genus of the family Neobisiidae Chamberlin, 1930. At present it includes 12 species , Stenohya curvata Zhao et al., 2011 (Yunnan Province) and Stenohya xiningensis Zhao et al., 2011 (Qinghai Province). species , of whicFagaceae, Styracaceae, Daphniphyllaceae, Lauraceae and Ericaceae  by sweeping vegetation with an entomological net. After examining the specimens in the laboratory, we found them to represent a new species, which is described here under the name Stenohya pengae sp. n. When we examined the pseudoscorpions collected by Prof. Fusheng Huang from Gushan Mountain, Fujian Province, China, we found another new Stenohya species, which is also described and illustrated in this paper as Stenohya huangi sp. n.Damingshan National Nature Reserve is located in the midwest of Guangxi Zhuang Autonomous Region, lying on the Tropic of Cancer, and possesses a rich subtropical primeval forest, which is home to many rare animals and plants. Daming Mountains are densely covered by jungle, including trees of the families ricaceae . In 2011b = basal; sb = sub-basal; st = sub-terminal; t = terminal; ib = interior basal; isb = interior sub-basal; ist = interior sub-terminal; it = interior terminal; eb = exterior basal; esb = exterior sub-basal; est = exterior sub-terminal; et = exterior terminal.The specimens are preserved in 75% alcohol and deposited in the Museum of Hebei University (MHBU). Permanent slide mounts were prepared by removing the chelicerae, pedipalps, leg I and leg IV from specimens with small needles and clearing overnight with lactic acid at room temperature. Drawings were made with the aid of a camera lucida mounted above the eyepiece of a compound microscope. Photographs were taken with a Leica M165 stereomicroscope. Terminology of trichobothria follows urn:lsid:zoobank.org:act:3E8D205C-B127-4BE0-AC87-B029B7F9719Fhttp://species-id.net/wiki/Stenohya_pengae23\u00b008'N, 108\u00b017'E], alt. 1250 m, 21 May 2011, Yan-qiu Peng leg. Tree-crown layer of Castanopsis fabri. Paratypes: 17 males and 25 females, same data as for holotype.Holotype male (Ps.-MHBU-GX110521), China: Guangxi Province, Nanning City, Daming Mountain , 24 February 1975, Fu-sheng Huang. Habitat unknown.PageBreakHolotype female (Ps.-MHBU-FJ750224), China: Fujian Province, Fuzhou City, Gushan Mountain [The specific name is a patronym in honour of Prof. Fu-Sheng Huang, who collected and donated the specimen.it and et at same level.Species with slender pedipalps  and slender legs IV (e.g. femur+patella 7.23 times as long as deep), with low numbers of the teeth (about 30) on movable chelal finger; trichobothria . Colour Carapace  smooth, Abdomen. Pleural membrane strongly granulate. Tergal chaetotaxy: 4: 12: 10: 10: 10: 10: 11: 11: 11: 10: 9, including at least 4 tactile setae on tergites VI\u2013XI; anterior genital sternite  with 22 PageBreakof femur with fine granulation; patella claviform, smooth; chelal fingers long and slender. Trichobothriotaxy: est, et and it grouped together distally; ist situated midway between isb and it, nearer to it than to isb. eb and esb situated on base of the hand, grouped very closely with ib and isb; b and sb closer to each other situated on the basal half, and st andPageBreakt closer to each other situated on the distal half of the movable finger. Fixed chelal finger with 63 pointed teeth of unequal length, movable finger with about 30 teeth which with 20 pointed teeth slightly unequal length in distal half, and 10 rounded teeth in basal half.Pedipalps . Apex ofCheliceral palm  with 7 sLegs . Tibia IDimensions (in mm) and ratios (in parentheses). Body length 4.2. Carapace 1.29/0.89 (1.45); diameter of anterior eye 0.10; diameter of posterior eye 0.09. Pedipalps: trochanter 0.59/0.26 (2.27), femur 1.58/0.25 (6.32), patella 1.38/0.26 (5.31), chela (with pedicel) 2.58/0.53 (4.87), chela (without pedicel) 2.42 (4.57), hand length (without pedicel) 1.04 (1.96), movable finger length 1.44 (1.38 times longer than hand without pedicel). Chelicera 0.67/0.38 (1.76), movable finger length 0.45. Leg I: femur 0.79/0.13 (6.08), patella 0.59/0.13 (4.54), tibia 0.63/0.10 (6.30), basitarsus 0.45/0.10 (4.50), telotarsus 0.43/0.10 (4.30). Leg IV: femur + patella 1.52/0.21 (7.24), tibia 1.22/0.12 (10.17), basitarsus 0.59/0.10 (5.90), telotarsus 0.79/0.10 (7.90).This species is known only from the type locality.Stenohya huangi sp. n. is only known from the female, but it can be easily separated from most other species of this genus by the proportions of pedipalpal femur and patella . Finally, the new species differs from Stenohya vietnamensis in having an epistome. patella . Two speme level  in Stenot and et : fig. 28Neobisiidae the lyrifissures near the trochanteral foramen of the pedipalpal coxa number 3 or 4  has 6 foramenal lyrifissures and 1 dorsal lyrifissure (PageBreakIn most r 3 or 4 . Having dorsally . Males o foramen : fig. 3 ifissure ."}
+{"text": "AbstractBambusananus cuihuashanensissp. n. , a new bamboo-feeding leafhopper species, is described and illustrated from Shaanxi Province of China. Checklist, host plants and distribution for each species of Bambusananus is given along with a key to all known species. Athysanini, was established by Van Duzee in 1892 and is the largest tribe of Deltocephalinae, including 228 genera and 1123 species  was established by Bambusananus furcatus Li & Xing, 2011, from Guizhou Province of China as its type species, and three new combinations: Bambusananus binotatus , Bambusananus bipunctatus  and Bambusananus maculipennis  were proposed at the same time. Bambusananus bipunctatus , Bambusananus maculipennis , Bambusananus furcatus Li & Xing, 2011, Bambusananus lii  and Bambusananus yangae Xing & Chen, 2013  and are currently known only from southern mainland China and Taiwan .In the present paper, terminology follows 1.Bambusananus bipunctatus  (in 1999) in Indocalamus hirsutissimus Z. P. Wang & P. X. Zhang and Qiongzhuea communis Hsueh & Yi) (Host plant. Bamboo (eh & Yi) .Distribution. China  .PageBreak2.Bambusananus maculipennis Chimonobambusa pachystachys Hsuch & W. P. Zhang and Qiongzhuea communis Hsueh & Yi) (Host plant. Bamboo (eh & Yi) .Distribution. China (Guizhou) .3.Bambusananus cuihuashanensis sp. n.Host plant. Bamboo.Distribution. China (Shaanxi) .4.Bambusananus furcatus Li & Xing, 2011Chimonobambusa angustifolia C. D. Chu & C. S. Chao) (Host plant. Bamboo (S. Chao) .Distribution. China (Guizhou) .5.Bambusananus lii Host plant. Bamboo .Distribution. China (Taiwan) .6.Bambusananus yangae Xing & Chen, 2013Indocalamus sp.) (Host plant. Bamboo (mus sp.) .Distribution. China (Guizhou) .http://zoobank.org/4B853422-EEC6-41EB-AD6B-B1570C4E92E2http://species-id.net/wiki/Bambusananus_cuihuashanensisChina: Shaanxi, Xi\u2019an, Cuihuashan , on bamboo, 37 Aug. 2008, J.-D. Li; paratypes: 2 \u2642\u2642, 4 \u2640\u2640, same data as holotype.Holotype: \u2642, The new species is named after its locality, Cuihuashan, Shaanxi Province, China.Body length (from apex of vertex to tip of forewings): male 4.75\u20134.85 mm (N = 2); female 5.75\u20135.90 mm (N = 4); forewing length: male 3.95\u20134.05 mm (N = 2); female 5.00\u20135.15 mm (N = 4).Crown  pale yelExternal features as in generic description. Crown shorter medially than width between eyes (0.54:1). Pronotum longer medially than crown (2.10:1). Scutellum shorter medially than pronotum (0.89:1). Forewing longer medially than width at widest part (3.38:1).Male pygofer  with basBamboo.PageBreakPageBreakChina (Shaanxi) .Bambusananus bipunctatus  in general appearance, but can be distinguished by: pygofer in lateral view with ventral margin broadly smoothly rounded, without notch at base of ventral process (with notch in Bambusananus bipunctatus); connective with stem slightly shorter than arms (longer in Bambusananus bipunctatus); aedeagal shaft with appendages longer, in lateral view with apex reaching to middle of aedeagus . This new species is also similar to Bambusananus lii , but can be distinguished by: upper area of frontoclypeus with a large kidney-shaped black marking (lacking in Bambusananus lii); ventral margin of pygoger without any lobe (with a lobe near middle in Bambusananus lii); appendages of aedeagal shaft mostly straight basally, apex directed laterally ; ventral margin of aedeagus without any teeth (with a row of teeth at middle in Bambusananus lii).This new species is similar to"}
+{"text": "We previously reported a cross-sectional association between the presence of human herpesvirus 8 (HHV-8) serum antibodies and screen-detected prostate cancer in men living in Tobago. In the same study population, we examined the association between HHV-8 seropositivity and incident prostate cancer discovered at later screenings.In 40-81 year-old men without prostate cancer discovered at initial digital rectal examination (DRE) and prostate-specific antigen (PSA) screening, a case-cohort design measured the association between baseline HHV-8 seropositivity (modified immunofluorescence assay for antibodies against HHV-8 lytic antigens) and incident prostate cancer detected at DRE and PSA screenings three or five years later.vs. 9.9% seropositive; crude odds ratio (OR) 3.96, 95% confidence interval (CI) 1.53-10.2; age-adjusted OR 2.42, 95% CI 0.91-6.47). HHV-8 seropositivity did not increase incident prostate cancer risk (age-adjusted hazard ratio (HR) 0.88, 95% CI 0.46-1.69).Analyses included 486 unique individuals, 96 incident prostate cancer cases, and 415 randomly selected subjects representing an at-risk cohort. By design, the random sub-cohort contained 25 incident prostate cancer cases. In the sub-cohort, the frequency of HHV-8 seropositivity increased across age groupings . HHV-8 seropositivity was higher in men with elevated (\u2265 4.0 ng/mL) than men with non-elevated PSA at initial screening . Co-occurrence of HHV-8 seropositivity and PSA elevation may explain cross-sectional association between HHV-8 and PSA screen-detected prostate cancer. Neisseria gonorrhoeae, Chlamydia trachomatis, human papillomavirus (HPV) type 18, and Treponema pallidum (syphilis)  measures of unadjusted and age-adjusted association between HHV-8 seropositivity measured at baseline (Wave 1) and prostate cancer detected later in time, at Wave 2 or Wave 3 [We used the chi-square test to evaluate 1) the statistical significance of differences between at-risk men with incomplete r Wave 3 . These mvs. 26.0% \u2265 60 years), smoking history (48.5% vs. 39.6%), personal cancer history (1.3% vs. 0.3%), Wave 1 PSA \u2265 4 ng/mL (13.5% vs. 5.5%), and Wave 1 DRE (64.7% vs. 76.3% negative and 20.0% vs. 10.3% missing).Table vs. 5.5% PSA \u2265 4 ng/mL), and more frequently positive Wave 1 prostate cancer screening (43.8% vs. 16.9% DRE or PSA positive). Case and sub-cohort HHV-8 seropositivity rates were 17.7% and 11.1%, respectively. When compared with 40-49 year-old sub-cohort men (3.5% HHV-8 seropositive), HHV-8 seropositivity was higher in 50-59 year-old sub-cohort men (13.6% HHV-8 seropositive) and higher yet in \u2265 60 year-old sub-cohort men (22.9% HHV-8 seropositive). HHV-8 seropositivity rates were lower in sub-cohort men with a history of smoking than those without (7.1% vs. 13.9%) and higher in sub-cohort men with a history of benign prostatic hypertrophy than those without (24.0% vs. 10.5%). HHV-8 seropositivity increased with Wave 1 PSA . HHV-8 seropositivity was higher in men with elevated (\u2265 4.0 ng/mL) than men with non-elevated PSA . HHV-8 seropositivity was higher in sub-cohort men with a positive than in men with a negative Wave 1 prostate cancer screen result (20.0% vs. 9.3% seropositive).Wave 1 HHV-8 serologic status was available for 415 (93.9% of 442) men in the sub-cohort and for 96 (88.1% of 109) men in the case group . Table vs. 3.5% and 50-59 years: 10.8% vs. 13.6%), except in the oldest age group (\u2265 60 years: 27.7% vs. 22.9%). In men with a non-elevated (< 4 ng/mL) Wave 1 PSA, the HHV-8 seropositivity rate was higher in the case group (17.1% vs. 9.9%). In men with an elevated (\u2265 4 ng/mL) Wave 1 PSA, however, the HHV-8 seropositivity rate was lower in the case group (19.2% vs. 30.4%). In the two age sub-groups with appreciable HHV-8 seropositivity, age-specific HHV-8 seropositivity rates were not consistently higher or lower in case than sub-cohort men with non-elevated Wave 1 PSA (50-59 years: 11.5% vs. 13.7% and \u2265 60 years: 28.1% vs. 20.4%) and consistently lower in case than sub-cohort men with elevated Wave 1 PSA (50-59 years: 9.1% vs. 12.5% and \u2265 60 years: 26.7% vs. 41.7%).Table Table Our previous study used an immunofluorescence assay to measure HHV-8 antibodies in 138 prostate cancer cases and in 140 age-matched controls . HHV-8 se.g., PSA \u2265 4 ng/mL, age-adjusted HR 0.39, 95% CI 0.10-1.63; Table A positive association between HHV-8 seropositivity and prevalent prostate cancer in a cross-sectional study  and an iFour comparative studies of HHV-8 and prostate cancer have appeared ,11,26,27Analyses restricted to the sub-cohort showed strong association 1) between HHV-8 seropositivity and increasing age, a result also seen in Tobago women  and manyStudy strengths include unique population and setting (predominantly African ancestry Tobago residents ) and a cOur prospective study could not demonstrate an association between HHV-8 seropositivity and incident prostate cancer. However, analyses uncovered a strong relationship between elevated HHV-8 seropositivity and PSA. The HHV-8 association previously observed with prevalent prostate cancer may signify enhanced detection of prostate cancer possibly caused by the effects of HHV-8 on PSA. In this context, the association we observed between HHV-8 seropositivity and PSA elevation deserves further study.HHV-8: human herpesvirus 8; DRE: digital rectal examination; PSA: prostate-specific antigen; OR: odds ratio; CI: confidence interval; HR: hazards ratio; HPV: human papillomavirus; PLCO: Prostate Lung Colorectal, and Ovarian Cancer Screening Trial.The authors declare that they have no competing interests.ACM participated in study design, data analysis and interpretation, prepared a first draft of the manuscript, and helped revise the final manuscript. FJJ completed immunofluorescence assays and helped revise the final manuscript. CHB conceived the study, acquired data, provided study coordination, and helped revise the final manuscript. JWW selected analytic methods and directed statistical analysis. ALP acquired data and provided study coordination. JLW participated in study design, data analysis and interpretation and helped revise the final manuscript. All authors read and approved the final manuscript."}
+{"text": "To determine the nosocomial acquisition rate of ESBL-E among patients and healthcare workers (HCWs) during an epidemic (March 2009 to Nov 2010) in an orthopaedics ward at HUG.Universal screening made by anal swab of all patients on admission and every 2 weeks if screening remained negative. 49 samples were collected from 41 HCW and 60 environmental samples were analysed. Molecular typing was performed on all ESBL-E isolates. If there was more than 97.5% similarity, strains were considered identical.Between March 2009 and November 2010, 1\u2019531 admissions occurred to the orthopaedic ward . Among 565 anal swabs, ESBL-E were detected in 204 samples from 45 patients.E. coli (n=39), Klebsiella pneumoniae (n=10), Enterobacter spp (n=8), Citrobacter spp (n=2), Morganella morganii (n=2), and Proteus vulgaris (n=1). Two different ESBL-E strains were detected in 6 patients, and 3 others carried three distinct isolates. The ESBL-E transmitted were E. coli (14 patients), K. pneumoniae (3 patients) and both in 2 patients.The ESBL-E found were Identical ESBL-E species with epidemiological links were found in 25 cases. Only 9 of these were attributable to the unit. Most positive patients (96% [43/45]) were colonized asymptomatically with ESBL-E.Among HCWs, 6 samples (12%) were positive. Transmission was only observed between patients, not HCWs.None of the environmental samples revealed presence of ESBL-E.Transmission of ESBL-E strains was only observed between patients. No transmission between HCWs and patients occurred. HCW screening and environmental sampling is not useful during ESBL-E carriage outbreaks.E. coli.The main ESBL-E transmitted was E. coli-ESBL carriers.ESBL-E transmission can occur in units with extended length of stay, questioning the new Swiss policy of abandoning contact precautions for None declared"}
+{"text": "In the Phase III, randomised, open-label ODIN trial, treatment-experienced HIV-1-infected adults with no screening DRV resistance-associated mutations received DRV/r 800/100mg qd or DRV/r 600/100mg bid (both arms + \u22652 NRTIs). At Week 48, 72.1% qd vs 70.9% bid patients achieved HIV-1 RNA <50 copies/mL , confirming non-inferiority of DRV/r qd. The relationship between DRV PK and efficacy and safety following treatment with DRV/r is explored.0h) and exposure  using a population pharmacokinetic model. Relationships between PK parameters and efficacy  were assessed using ANCOVAs. Relationships between PK parameters and occurrence of adverse events of interest and laboratory lipid abnormalities were evaluated using descriptive statistics.Sparse blood sampling for PK evaluations was taken at Weeks 4, 8, 24 and 48 to determine DRV trough concentrations (C0h was 1896 (184-7881) ng/mL for DRV/r qd and 3197  ng/mL for DRV/r bid. Median (range) AUC24h for DRV/r qd was 87,788  ng.h/mL and 109,401  ng.h/mL for bid. No relevant relationships were observed between DRV PK and efficacy: changes from baseline in HIV-1 RNA log10 copies/mL at Week 48 for pooled data by DRV AUC24h quartile ranges  were -2.06, -2.22, -2.19, and -2.08 log10 copies/mL, respectively. The % patients achieving HIV-1 RNA <50 copies/mL by these quartile ranges were 82.0%, 88.5%, 82.6% and 76.5% (observed data), respectively. In a logistic regression analysis adjusting for baseline viral load, AAG levels and number of sensitive NRTIs in the optimised background regimen, there were no relevant relationships between PK and virological response. No apparent relationships were observed between DRV PK and occurrence of rash-, cardiac-, GI-, liver-, lipid- and glucose-related AEs or laboratory lipid abnormalities.PK data were available for 280 DRV/r qd patients and 278 bid patients. Median (range) C0h and AUC24h compared to DRV/r 600/100mg bid; however, comparable efficacy between DRV/r qd and bid confirmed adequate DRV concentrations were achieved following qd dosing. No relevant relationships were observed between DRV PK and efficacy or safety at Week 48.Dosing with DRV/r 800/100mg qd resulted in lower C"}
+{"text": "Tricholepidion gertschi is the sole extant representative of the family Lepidotrichidae. Its phylogenetic position is of special interest, since it may provide crucial insights into the early phenotypic evolution of the dicondylian insects. However, the phylogenetic position of T. gertschi is unclear. Originally, it was classified among silverfish (Zygentoma), but various alternative relationships within Zygentoma as well as a sistergroup relationship to all remaining Zygentoma + Pterygota are discussed, the latter implying a paraphyly of Zygentoma with respect to Pterygota. Since characters of the head anatomy play a major role in this discussion, we here present the so far most detailed description of the head of T. gertschi based on anatomical studies by synchrotron micro-computer tomography and scanning electron microscopy. A strong focus is put on the documentation of mouthparts and the anatomy of the endoskeleton as well as the muscle equipment. In contrast to former studies we could confirm the presence of a Musculus hypopharyngomandibularis (0md4). The ligamentous connection between the mandibles composed of Musculus tentoriomandibularis inferior (0md6) is also in contact with the anterior tentorium. Phylogenetic analysis of cephalic data results in monophyletic Zygentoma including T. gertschi. Zygentoma are supported by the presence of a set of labial muscles originating at the postocciput, presence of an additional intralabral muscle, and four labial palpomeres. Character systems like the genitalic system, the mating behaviour, the segmentation of the tarsi, the overall body form, and the presence of ocelli which were proposed in other studies as potentially useful for phylogenetic reconstruction are evaluated.The relic silverfish  Trichlepidion gertschi occurs only in the coastal region of northern California. The species is characterized by a number of peculiarities with respect to all other extant silverfish species (= Euzygentoma) such as the presence of ocelli (in addition to compound eyes), 5-segmented tarsi [T. gertschi is the sister species to all other dicondylians (Zygentoma + Pterygota), sister species to all other Zygentoma, or a subgroup within Zygentoma . Moleed tarsi -7 and moed tarsi -13 disagT. gertschi is crucial to understand the evolution of several morphological characters in the stem lineage of Dicondylia, e.g. presence of a proventriculus [However, the phylogenetic position of triculus , sperm ctriculus ,21, the triculus ,14 and ttriculus  includintriculus .T. gertschi to remaining Zygentoma [T. gertschi with a strong focus on the documentation of mouthparts, endoskeleton and muscle equipment. We show that characters of the cephalic morphology indeed provide information on a sistergroup relationship of T. gertschi and the remaining studied Zygentoma.Available cephalic data did not help thus far to resolve the phylogenetic relationships of ygentoma ,14. Thissynchrotron radiation micro-computer tomography (SR-\u03bcCT), so that they could be used to complement the character matrix , Thermobia domestica  (Lepismatidae), Lepisma saccharina Linneaus, 1758 (Lepismatidae), and Atelura formicaria Heyden, 1855 (Nicoletiidae). The outer anatomy of T. gertschi was furthermore investigated with scanning electron microscopy (SEM).All specimens were fixed in Bouin's solution  and inveSR-\u03bcCT was done using the recommendations of Betz et al.  and the http://www.blender.org). Both software packages are distributed under the general public license (GPL). Tables and figures were edited with GIMP ver. 2.8, Inkscape ver. 0.48 and Scribus ver. 1.4.1 . A 3D model of the head of T. gertschi is available  the specimen was transferred in a series of steps into 100% ethanol, critical point dried , and sputter coated (Model Anatech Hummer VII). Microscopy was performed on a Hitachi S-2460 N scanning electron microscope using a rotatable sample holder . The terMachilis germanica was selected as the outgroup. Only unambiguous changes were mapped on the optimal trees. Character numbers and states are given in brackets using the following syntax: (character number : character state). The character matrix is derived from Blanke et al. [Parsimony analyses of 139 cephalic characters (see Appendix) and Bremer support (BR) calculations were carried out with TNT  using 1,e et al.  and is be et al. . Please 2. Among them, numerous (>500/100 \u03bcm2) small tubercles on the exoskeleton, possibly sensorial in function, cover the entire head : insertion directly anterior of 0md1 (muscle \"spt\" of Chaudonneret [0te2): the insertion in between the muscle bundles of 0md1 supports homologisation with the muscle \"spot\" of Chaudonneret [0te3): O, anterior side of the dorsal tentorial arm; I, frons near the dorsal part of the antennal base. M. posterotentorialis (0te4): absent. M. tentoriotentorialis longis (0te5): O, ventrolaterad on the anterior corpotentorium; I, mesad on the posterior tentorium. M. tentoriotentorialis brevis (0te6): O, anterior tentorium, along the gap towards the posterior tentorium; I, posterior tentorium, along the gap towards the anterior tentorium.Musculature: M. tentoriofrontalis posterior (donneret ); O, at donneret ; O, at tThe convex labrum Figures  is cover0lb1): O, mesally on the epistomal ridge; I, mesally on the inner basal wall of the labrum. M. frontoepipharyngalis (0lb2): absent. M. epistoepipharyngealis (0lb3): this muscle can easily be confused with the 0lb2. The origin of 0lb3 lies clearly only on the epistomal ridge. O, laterally on the epistomal ridge; I, basal epipharyngeal wall. M. labralis transversalis (0lb4): absent. M. labroepipharyngealis (0lb5): O, basal labral wall; I, basal wall of epipharynx. M. labrolabralis (0lb6): O, mesobasal labral wall in between the two bundles of 0lb5; I, medioapical area of labrum.Musculature: M. frontolabralis : O, mesally on the corpotentorium; I, anterior margin of the scapus. M. tentorioscapalis posterior (0an2): O, in the posterior part of the corpotentorium; I, posterior basal margin of the scape. M. tentorioscapalis lateralis (0an3): O, at the anterior base of the dorsal tentorial arms; I, dorsal basal margin of the scape. M. tentorioscapalis medialis (0an4): O, at the dorsal tentorial arms directly over 0te1; I, dorsal basal margin of the scape. M. frontopedicellaris (0an5): absent. M. scapopedicellaris lateralis (0an6): O, broadly on the dorsolateral base of the scape; I, dorsolateral base of the pedicellus. M. scapopedicellaris medialis (0an7): O, ventrolateral base of the scape; I, ventrolateral base of the pedicel. M. intraflagellaris (0an8): absent. M. interampularis (0ah1): absent. M. ampulloaortica (0ah2): absent. M. ampullopharyngealis (0ah3): absent. M. ampullofrontalis (0ah4): absent. M. frontofrontalis (0ah5): absent.Musculature: M. tentorioscapalis anterior  reaches into the lumen of the mandible directly posterad the mandibular ridge Figure . The twoThe mola is almost formed like a right-angled triangle in lateral view Figure , with th0md1): O, dorsal parts of the head, anterior of the postoccipital ridge and on this ridge; one muscle bundle also posterior of the postoccipital ridge; I, tendon originating from the proximal part of the posterior mandibular edge. M. craniomandibularis externus anterior (0md2): O, gena, anterior of the compound eye; I, tendon originating from the proximal part of the anterior mandibular edge, directly lateral to the anterior articulation complex. M. craniomandibularis externus posterior (0md3): O, directly posterior of the compound eye, with several bundles also at the postoccipital ridge; I, tendon originating from the anterior mandibular edge near the posterior articulation. M. hypopharyngomandibularis (0md4): O, small sclerite close to the loral arm of the hypopharynx; I, proximal inner side of the anterior wall of the mandible. M. tentoriomandibularis lateralis superior (0md5): O, ventrally at the anterior tentorial arm; I, anterior mandibular rim between 0md2 and 0md3. M. tentoriomandibularis lateralis inferior (0md6): O, the whole inner wall of the mandible except for the region near the incisivi and mola; I, in the same region of the other mandible and with a few muscle bundles also at the ventral anterior area of the corpotentorium. M. tentoriomandibularis medialis superior (0md7): O, at the ventral base of the anterior tentorial arms; I, proximal of the posterior articulation at the posterior mandibular rim. M. tentoriomandibularis medialis inferior (0md8): O, at the transition of the corpotentorium and the anterior tentorial arm below 0md7; I, mediodorsal wall of the mandibular cavity.Musculature: M. craniomandibularis internus  is three times longer than wide in overall shape Figure . The carThe stipes is composed of a narrow basistipes and a much larger mediostipes bearing the palpus, galea and lacinia Figure . The whoThe palpus is five-segmented and densely covered with trichoid sensilla. The first segment is one third as long as the second one. The third one is slightly longer than the second one, the fourth and fifth are as long as the second one. Each segment is slightly thinner than the preceding one. The fifth segment bears distally six special sensilla formed by a basal cylindrical segment densely covered with microtrichia and four to six tubular extensions on the tip of the cylindrical base Figures . These sThe galea is sickle-shaped Figure , distall0mx1): O, laterally at the postoocipital ridge; I, basal cardinal process. M. craniolacinialis (0mx2): O, distal of 0mx1 at the postoocipital ridge; I, cuticular tendon at the basal edge of lacinia together with 0mx6. M. tentoriocardinalis (0mx3): O, at the lateral part of the posterior tentorium; I, inner wall of the cardo. M. tentoriostipitalis anterior (0mx4): O, ventral on the corpotentorium; I, at the posterior stipital rim. M. tentoriostipitalis posterior (0mx5): O, lateral on the corpotentorium; I, at the basal inner wall of the stipes. M. stipitolacinialis (0mx6): O, posterior wall of the stipes, basal to the cardostipital ridge; I, basal proximal edge of lacinia, at a common cuticular tendon with 0mx2. M. stipitogalealis (0mx7): O, posterior wall of the stipes, basal to the cardostipital ridge next to 0mx6; I, basal edge of galea. M. stipitopalpalis externus (0mx8): O, posterior wall of the stipes, opposite of the palpus; I, posterior rim of the first palpomere of the maxillary palpus. M. stipitopalpalis medialis (0mx9): O, medially at the posterior wall of the stipes; I, ventral edge of palpomere 1. M. stipitopalpalis internus (0mx10): O, posterior inner wall of the stipes, opposite of the palpus; I, anterior rim of the first palpomere of the maxillary palpus. M. stipitalis transversalis (0mx11): O, outer stipital wall, near the palpus; I, inner stipital wall, near the palpus. M. palpopalpalis maxillae primus (0mx12): O, basal edge of palpomere 1; I, basal edge of palpomere 2. M. palpopalpalis maxillae secundus (0mx13): O, mesal edge of palpomere 2; I, mesal edge of palpomere 3. M. palpopalpalis maxillae tertius (0mx14): O, basal edge of palpomere 3; I, basal edge of palpomere 4; M. palpopalpalis maxillae quartus (0mx15): unclear.Musculature: M. craniocardinalis : O, posterior side of the postoccipital ridge; I, proximal area of the glossal base. M. postoccipitoglossalis lateralis (0la2): O, posterior side of the postoccipital ridge, right next to 0la1; I, dorsolateral area of the glossal base. M. postoccipitoparaglossalis (0la3): O, posterior side of the postoccipital ridge, lateral of 0la1 & 0la2; I, distal base of the paraglossa. M. postoccipitopraementalis (0la4): O, posterior side of the postoccipital ridge, right next to 0la1 & 0la2; I, inner proximal wall of the prementum. M. tentoriopraementalis (0la5): O, lateral area of the posterior tentorium; I, laterobasal edge of prementum. M. tentorioparaglossalis (0la6): O, lateral area of the posterior tentorium, right next to 0la5; I, paraglossa, close to the labial palpus. M. tentorioglandularis (0la7): O, lateral posterior area of the corpotentorium; I, labial gland. M. submentopraementalis (0la8): O, medially on the postmentum; I, mediobasal edge of prementum. M. postmentomembranus (0la9): unclear. M. submentomentalis (0la10): absent. M. praementoparaglossalis (0la11): O, distal on the basal edge of the prementum; I, basal edge of paraglossa. M. praementoglossalis (0la12): O, more mesally on the basal edge of the prementum, right next to 0la11; I, basal edge of glossa. M. praementopalpalis internus (0la13): O, mesally on the prementum; I, anterior basal edge of labial segment 1. M. praementopalpalis externus (0la14): O, mesally on the prementum, right next to 0la13; I, posterior basal edge of labial segment 1. M. praementomembranus (0la15): O, anterolateral area of the postmentum; I, anteromedial area of postmentum. M. palpopalpalis labii primus (0la16): O, mediobasal edge of labial palpomere 1; I, medial and distal edge of palpomere 2. M. palpopalpalis labii secundus (0la17): O, mediobasal edge of palpomere 2; I, basal edge of the palpomere 3.Musculature: M. postoccipitoglossalis medialis : O, frons, near the antennal base; I, oral arms of the suspensorial sclerites. M. tentoriooralis (0hy2): O, anterior tentorial arm, near the anterior tentorial pit; I, one muscle bundle on the oral arms of the suspensorial sclerites, one bundle at the lateral buccal wall: both bundles are well separated from 0hy1. M. craniohypopharyngealis (0hy3): O, posterior tentorial arms; I, suprasalivarial sclerite. M. postoccipitalohypopharyngealis (0hy4): O, posterior wall of the postoccipital ridge, together with 0la1 & 0la2; I, hypopharyngeal fulcrum. M. tentoriosuspensorialis (0hy5): O, anterior margin of the posterior tentorium; I, suspensorium of the hypopharynx. M. postmentoloralis (0hy6): O, anterior part of the postmentum; I, loral arm of the hypopharyngeal suspensorium. M. praementosalivaris anterior (0hy7): O, distolateral wall of the prementum, close to the labial palpus; I, lateral wall of salivarium. M. praementosalivaris posterior (0hy8): O, medially on the basal part of the prementum; I, posterior wall of salivarium. M. oralis transversalis (0hy9): unclear. M. loroloralis (0hy10): O, loral arm of suspensorial sclerite; I, loral arms of the suspensorial sclerites on the other side. M. lorosalivarialis (0hy11): O, hypopharyngeal suspensorium; I, loral arm of the hypopharynx. M. hypopharyngosalivaris (0hy12): O, loral arm of the hypopharynx, right next to 0hy11; I, salivarial orifice. M. anularis salivarii (0hy13): unclear.Musculature: M. frontooralis : two distinct muscle bundles. O, postclypeus; I, roof of the cibarium. M. clypeobuccalis (0bu1): O, clypeus, near the epistomal ridge; I, roof of the bucca. M. frontobuccalis anterior (0bu2): O, frons dorsal of the epistomal ridge; I, dorsal buccal wall. M. frontobuccalis posterior (0bu3): O, more posterior than 0bu2 on the frons; I, dorsal buccal wall, posterior of 0bu2. M. tentoriobuccalis lateralis (0bu4): absent. M. tentoriobuccalis anterior (0bu5): O, anteriormost part of the corpotentorium between the anterior tentorial arms; I, ventral wall of the bucca, directly behind the anatomical mouth. M. tentoriobuccalis posterior (0bu6): O, laterally on the dorsal wall of the corpotentorium; I, lateral wall of the foregut at height of the 0ph1. M. verticopharyngealis (0ph1): O, frons right next to the 0te4; I, dorsal wall of the foregut, posterior to the supraoesophagial ganglion. M. tentoriopharyngealis (0ph2): O, laterally on the dorsal wall of the corpotentorium right next to 0bu6; I, ventral wall of the foregut, beneath 0ph1. M. postoccipitopharyngealis (0ph3): absent. M. anularis stomodaei (0st1): ring muscle layer that covers the entire foregut. M. longitudinalis stomodaei (0st2): longitudinal muscle layer covering the entire foregut, right next to 0st1.Musculature: M. clypeopalatalis , presence of M. labroepipharyngealis (0lb5), M. verticopharyngealis (0ph1), M. tentoriopharyngealis (0ph2) and the five-segmented maxillary palpus.Generally, the monophyly of Dicondylia and Pterygota is well supported by molecular and morphological data ,41 even The monophyly of winged insects is strongly supported  was considered absent [Tricholepidion) exhibit an origin of this muscle exclusively on the tentorium. Our phylogenetic analysis corroborates monophyletic Zygentoma despite the inclusion of the mandible ligament in the character matrix (character 69) with high support . Euzygentoma and T. gertschi possess a remarkable set of extrinsic labial muscles originating from the postoccipital region and extending dorso-ventrally through the whole head into the labium. Archaeognatha and Pterygota clearly do not possess this set of labial muscles [gertschi and Euzygentoma, possess four labial palpomeres (103:3). Also, the intralabral muscle equipment is characterized by an additional muscle, the M. epistoepipharyngalis . In the present analysis the low number of ommatidia  is a homoplastic character since Grylloblattodea also show reduced eyes [Galloisiana yuasai [T. gertschi as sister to Euzygentoma.The position of ygentoma ,18, as sygentoma ,16, or aygentoma ,19 withid absent ,14 and dd absent ,44,45. Id absent ,44,45, w muscles ,44,46. Tced eyes  which hoced eyes ). A M. la yuasai . Except Tricholepidion as sistergroup to Odonata with low support while Giribet [Tricholepidion as sister taxon to Euzygentoma + Pterygota with low support. All other molecular works supported monophyletic Zygentoma [With few exceptions most molecular studies advocate monophyletic Zygentoma. Based on secondary structure alignment of the 18S unit Kjer  recovere Giribet , using aygentoma ,4,6,7.T. gertschi and Lepismatidae [T. gertschi Dallai et al. [T. gertschi resembles the ancestral state of insects with a 9 + 9 + 2 axonemal pattern and accessory tubules with 16 protofilaments. The sperm pairing in T. gertschi is a true fusion between two spermatozoa along the entire sperm head region, which is different from sperm aggregations in other Zygentoma [T. gertschi  in smatidae . Zrzav\u00fd smatidae  considersmatidae ). In a di et al. ,21 discoygentoma . Characteognatha ). The oveognatha ) and higeognatha .T. gertschi include similarities in the mating behaviour of T. gertschi and Lepismatidae [T. gertschi and some Nicoletiidae [Petrobiellus and Mesomachillis species [T. gertschi. The widening of the apical labial palpus segment is paralleled in males of several Machilidae and Meinertellidae  and, heT. gertschi, since these occur as well in several pterygote taxa and the phylogenetic significance is therefore unclear. Zrzav\u00fd [T. gertschi, it is conceivable that five-segmented tarsi already existed in the stem lineage of Dicondylia. Engel [Also, we interpret the five-segmented tarsi  not as a. Zrzav\u00fd  considera. Engel  even cona. Engel ).T. gertschi in its own family Tricholepidiidae because the extinct type species of the Lepidotrichidae, Lepidotrix pilifera Silvestri, 1912, might be more closely related to the Euzygentoma than to T. gertschi due to the apparent absence of ocelli [T. gertschi clearly possesses three ocelli, we consider this a weak argument for this classification, since loss of ocelli occurred several times within Dicondylia, e.g. in Xenonomia (= Notoptera + Mantophasmatodea), Phasmatodea [L. pilifera is mandatory. As for the head  we partThe authors declare that they have no competing interests.AB, BW and BM designed the study, AB and FW conceived the experiments. AB and FW conducted the SR\u03bcCT, AB the SEM and phylogenetic analysis. AB, MK and BW carried out the analysis of the raw data. AB, MK, BW, and BM wrote the manuscript. All authors read, revised and approved the manuscript.List of characters used for phylogenetic reconstruction0. Orientation of head: (0) orthognathous; (1) prognathous or slightly inclined; (2) hypognathous1. Number of ocelli: (0) 0; (1) 2; (2) 3.2. Compound eyes: (0) composed out of more than 80 ommatidia; (1) less than 80 ommatidia;3. Distance between eyes: (0) less than their own width; (1) greater than their own width; (2) eyes fused at single point; (3) eyes broadly fused along an eye seam4. Shape of vertex: (0) flat, not developed into large protuberance; (1) conical, or developed into a large transverse ridge5. Epicranial or coronal suture: (0) present; (1) absent6. Parietal ridge: (0) absent; (1) present.7. Postoccipital ridge: (0) present; (1) absent.8. Subgenal ridge: (0) absent; (1) present9. Pleurostomal ridge and circumocular ridge: (0) not in contact; (1) partly in contact10. Interantennal ridge: (0) absent; (1) present11. Shape of frons: (0) flat when seen from lateral; (1) outwardly bulged when seen from lateral12. Distinct convexity ventrad the antennal bases: (0) absent; (1) present13. Scutellum: (0) absent; (1) present14. x-shaped median apodeme on the frontal region: (0) absent; (1) present15. Clypeus: (0) not subdivided; (1) subdivided into ante- and postclypeus16. Postclypeus: (0) not enlarged; (1) enlarged17. Anteclypeus: (0) membranous; (1) sclerotised18. Adult mouthparts: (0) with function; (1) without function19. Oval sclerotization of labral base: (0) absent; (1) present.20. Tormae: (1) absent; (0) present21. Mesal extension of tormae: (0) present; (1) absent22. M. epistoepipharyngealis (0 lb3): (0) present; (1) absent23. M. labroepipharyngealis (0 lb5): (0) present; (1) absent24. M. labrolabralis (0 lb6): (0) present; (1) absent25. Insertion of antennae: (0) close to the anterior mandibular articulation with the pleurostomal and circumantennal ridges in contact (where applicable); (1) distinctly separated from the anterior mandibular articulation, pleurostomal and circumantennal ridges not in contact.26. Antennifer: (0) present; (1) absent27. Length of pedicel and scapus: (0) pedicel longer than scapus; (1) scapus longer than pedicel; (2) scapus and pedicel equal in length28. Oval scerite in membrane connecting scapus and pedicellus: (0) absent; (1) present.29. Size of first flagellomere: (0) not enlarged; (1) first flagellomere more than twice as long as second one.30. Antennal stridulatory organ: (0) absent; (1) present31. Areas of origin of antennal muscle 0an1: (0) anterior tentorial arms only; (1) anterior tentorial arms and tentorial bridge; (2) on dorsal tentorial arms only; (3) on dorsal arms and tentorial bridge; (4) anterior and dorsal tentorial arm.32. Areas of origin of antennal muscle 0an2: (0) anterior tentorial arms only; (1) anterior tentorial arms and tentorial bridge; (2) on dorsal tentorial arms only; (3) on dorsal arms and tentorial bridge; (4) tentorial bridge only; (5) dorsal and anterior tentorial arms.33. M. tentorioscapalis lateralis (0an3): (0) present; (1) absent34. M. tentorioscapalis medialis (0an4): (0) present; (1) absent35. Circumesophageal vessel ring branching off the dorsal aorta posterior to the brain: (0) present; (1) absent36. Ostia of dorsal vessel: (0) lips always present; (1) ostia with and without lips (excurrent ostia).37. Position and number of excurrent ostia within a segment: (0) one ventrolateral pair; (1) ventromedian.38. Antennal circulatory organs in adults: (0) present; (1) absent39. Antennal vessel wall: (0) uniform; (1) bipartite40. Contractibility of antennal ampulla: (0) absent (non-pulsatile); (1) present (pulsatile).41. M. interampullaris (0ah1): (0) absent; (1) present42. M. ampulloaorticus (0ah2): (0) absent; (1) present43. M. ampullopharyngealis (0ah3): (0) absent; (1) present44. M. ampullo-frontalis (0ah4): (0) absent; (1) present45. Connection of antennal ampulla to supraoesophageal ganglion: (0) absent; (1) present46. Oval nuclei in tissue connecting the antennal ampulla and supraoesophageal ganglion: (0) absent; (1) present47. Anterior and posterior tentoria: (0) seperated; (1) merged48. Transverse mandibular tendon: (0) present; (1) absent49. Processes of the anterior tentorial apodemes extending into the lumen of the mandibular base: (0) absent; (1) present50. Corpotentorium: (1) elongated; (0) slim.51. Apophyses on the anterior surface of the corpotentorium: (0) absent; (1) present52. Secondary anterior tentorial bridge (\u201cperforation of the corpotentorium\"): (0) absent; (1) present.53. Lateral lobes on the anterior tentorial arms: (0) absent; (1) present54. Cuticular dorsal tentorial arms: (0) absent; (1) present55. Trabeculae tentorii of posterior tentorial arms (0) present; (1) absent56. M. tentoriofrontalis posterior (0te1): (0) present; (1) absent57. M. posteriotentorialis (0te4): (0) present; (1) absent58. M. tentoriotentorialis longus (0te5): (0) present; (1) absent59. M. tentoriotentorialis brevis (0te6): (0) present; (1) absent60. Numbers of incisivi on the left mandible: (0) 2; (1) 3; (2) 5; (3) 0; (4) 1; (5) 461. Numbers of incisivi on the right mandible: (0) 2; (1) 3; (2) 4; (3) 5; (4) 0; 5 (1)62. Armament on the mesal side of the left mandible: (0) without teeth or ridges; (1) one tooth; (2) three ridges63. Dorsal cutting edge of the left mandible: (0) notched; (1) smooth64. Mandibular postmola: (0) absent; (1) present65. Anterior mandibular joint: (0) absent; (1) present66. Anterior mandibular joint: (0) cuticular hardening on the mandibular depression; (1) channel-joint (2) ball-and-socket joint67. Anterolateral part of the anterior mandibular articulation : (0) present; (1) absent68. Posterior mandibular joint: (0) cylinder-shaped (1) ball-and-socket joint69. Mandibular ligament: (0) present; (1) absent.70. M. craniomandibularis externus anterior (0md2): (0) present; (1) absent71. M. hypopharyngomandibularis (0md4): (0) present; (1) absent72. M. tentorio-mandibularis lateralis superior (0md5): (0) present; (1) absent73. M. tentorio-mandibularis medialis superior (0md7): (0) present; (1) absent74. Cardo: (0) present; (1) absent75. Division of stipes into basistipes and mediastipes: (0) present; (1) absent76. Galea: (0) present; (1) absent77. Distal field of trichomes on the galea: (0) undivided; (1) divided; (2) just a U-shaped seam78. Connection of lacinia and galea: (0) separated; (1) fused79. Shape of lacinia: (0) sickle-shaped; (1) chisel-shaped; (2) truncate; (3) short claw80. Mesally directed setae on lacinia: (0) present; (1) absent81. Lacinia: (0) free; (1) in galeal cavity82. Lacinial incisivi: (0) present; (1) absent83. Number of incisivi on lacinia: (0) 3; (1) 2; (2) 1; (3) more than 384. Dentisetae on lacinia: (0) present; (1) absent85. Proximal apodeme on the lacinia: (0) absent; (1) present86. Galeolobulus: (0) absent; (1) present87. Maxillary palpus: (0) 5-segmented; (1) 4-segmented; (2) 1-segmented; (3) 3-segmented; (4) 6-segmented; (5) 7-segmented88. Orientation of maxillary palpi: (0) ventrally oriented; (1) anteriorly or dorsally directed89. 0mx7: (0) present; (1) absent90. M. palpopalpalis maxillae primus (0mx12): (0) present; (1) absent91. Postmentum: (0) not subdivided; (1) subdivided into submentum and mentum92. Angle between submentum and mentum: (0) less than 60\u00b0 or absent; (1) more than 60\u00b093. Curvature of submentum: (0) absent; (1) curved in lateral view94. Median longitudinal tunnel of labium: (0) absent; (1) present95. Median cleft of prementum: (0) absent; (1) present96. Labium: (0) paraglossa and glossa seperated; (1) paraglossa and glossae completly fused97. Glossa: (0) present; (1) reduced98. Number of glossae: (0) 2; (1) 1;99. Number of paraglossae: (0) 2; (1) 1;100. Shape of paraglossa: (0) cylindrical, as wide as thick; (1) flat, wider than thick; (2) palpus-like.101. Relative length of paraglossae and glossae: (0) about equally long; (1) paraglossae twice as long or longer102. Orientation of labial palpi: (0) anterior or lateral; (1) ventral or posterior103. Number of labial palpomeres: (0) 3; (1) 1; (2) 2; (3) 4104. Shape of labial palpi: (0) approximately round in cross section; (1) dorsoventrally flattened105. Length of labial palpi: (0) longer than glossae; (1) about as long as the glossae106. Moveable hooks of labial palpi: (0) absent; (1) present107. M. postoccipitoglossalis medianus (0la1): (0) present; (1) absent108. M. postoccipitoglossalis lateralis (0la2): (0) present; (1) absent109. M. postoccipitoparaglossalis (0la3): (0) present; (1) absent110. M. postoccipitoprementalis (0la4): (0) present; (1) absent111. 0la5: (0) present; (1) absent112. Origin of M. tentoriopraementalis inferior 0la5 (M.29): (0) ventral apodeme; (1) posterior tentorial arms; (2) posterior tentorial arms (posttentoria) and postocciput.113. M. tentorioparaglossalis (0la6): (0) present; (1) absent114. Origin of M. tentorioparaglossalis (0la6): (0): tentorium; (1) basal edge of prementum115. M. tentorioglandularis (0la7): (0) present; (1) absent116. M. submentopraementalis (0la8): (0) present; (1) absent117. M. submentopraementalis (0la8): (0) one component; (1) two components118. M. postmentomembranus (0la9): (0) present; (1) absent119. M. submentomentalis (0la10): (0) absent; (1) present120. M. praementoparaglossalis (0la11): (0) present; (1) absent121. M. praementoglossalis (0la12): (0) present; (1) absent122. M. praementopalpalis internus (0la13): (0) present; (1) absent123. M. praementopalpalis externus (0la14): (0) present; (1) absent124. Hypopharynx overlapping paraglossae and glossae 0) absent; 1) present125. Shape of hypopharynx: (0) slope like; (1) distinctly flattened126. Superlinguae: (0) present; (1) absent127. Salivary glands and ductus: (0) present; (1) absent128. Connection of salivary ducts: (0) connected before opening, Y-shaped; (1) open separately129. M. frontobuccalis lateralis (0hy2): (0) present; (1) absent130. M. craniohypopharyngealis (0hy3): (0) present; (1) absent131. M. postmentoloralis (0hy6): (0) present; (1) absent132. M. praementosalivaris posterior (0hy8): (0) absent; (1) present133. M. lorosalivarialis (0hy11): (0) present; (1) absent134. 0hy12: (0) present; (1) absent135. M. frontobuccalis posterior (0bu3): (0) present; (1) absent136. M. tentoriobuccalis lateralis (0bu4): (0) absent; (1) present137. 0bu5: (0) present; (1) absent138. M. tentoriobuccalis posterior (0bu6): (0) present; (1) absent.139. Origin of M. tentoriobuccalis posterior 0bu6 (M.50): (0) anterior and/or posterior bridge, (1) pretentoriaHomologization of the muscular terminology with other authors. Abbreviations: +, muscle present ; -, muscle absent;/, muscle not dealt with by the author; ?, unclear homology.Click here for fileCharacter state matrix in excel format together with short descriptions of the charactes used: (?) refers to a missing or unclear character state, (-) to inapplicable characters.Click here for fileCharacter state matrix in nexus format: (?) refers to a missing or unclear character state, (-) to inapplicable characters.Click here for fileT. gertschi3D model of the head of . Please use the open source software Blender (http://www.blender.org) to view the model.Click here for fileImage stack of transversal slices through the head of T. gertschi packed in the \"avi\" movie format for easy browsing through the stack.Click here for fileComplete tree (strict consensus) showing all taxa. Character numbers above, character states below the nodes.Click here for file"}
+{"text": "In \u201cProbable Tiger-to-Tiger Transmission of Avian Influenza H5N1,\u201d by Roongroje Thanawongnuwech et al., an error occurred. The GenBank accession nos. for HA and NA gene initiated from influenza A virus (A/Tiger/ Thailand/CU-T3/04) are AY842935 and AY842936.http://wwwnc.cdc.gov/eid/article/11/5/05-0007_article.htm.The corrected article appears online at We regret any confusion these errors may have caused."}
+{"text": "S,6S,2R)-9,13-dimeth\u00adyl-5-methyl\u00adene-3-oxatricyclo\u00adtrideca-9,12-diene-4,11-dione], C15H16O3, is a guanolide isolated from Artemisia douglasiana. The fused-ring system contains a seven-membered ring that adopts a chair conformation, a fused planar cyclo\u00adpentenone ring and a five-membered lactone ring fused in envelope conformation. The absolute structure determined by X-ray analysis agrees with that previously assigned to this compound by NMR studies  and also with that of leucodine, a closely related guaianolide .Dehydro\u00adleucodin [systematic name: (1dero 1972. Tetra\u00adh al. 1988. J. Nat. DOI: 10.1107/S1600536811048938/bg2432Isup2.hkl            Structure factors: contains datablock(s) I. DOI: 10.1107/S1600536811048938/bg2432Isup3.mol            Supplementary material file. DOI: 10.1107/S1600536811048938/bg2432Isup4.cml            Supplementary material file. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:"}
+{"text": "Errors were introduced in the preparation of this article for publication: incorrect symbols were inserted in the legends of Tables S2, S3, S6, S7, and S8. The corrected legends are:Table S1. White-tailed Deer RRL Contig Table (3.66Mb XLS)Table S2. White-tailed Deer RRL Informative blastn Hits Table S3. White-tailed Deer RRL Putative SNPs Table S4. White-tailed Deer RRL Functional Annotation (1.07 Mb XLS)Table S5. White-tailed Deer RRL plus RSL (Pooled) Contig Table (7.99 Mb XLS)Table S6. White-tailed Deer RRL plus RSL (Pooled) Informative blastn Hits Table S7. White-tailed Deer RRL plus RSL (Pooled) Putative and Validated SNPs Table S8. White-tailed Deer RRL plus RSL (Pooled) Mitochondrial SNP Table Table S9. White-tailed Deer RRL plus RSL (Pooled) Functional Annotation (2.71 Mb XLS)"}
+{"text": "HIV-infected patients have a higher risk of developing cancer than the general population. Kaposi's sarcoma (KS), non-Hodgkin's lymphoma (NHL), primary CNS lymphoma (PCL) and invasive cervical cancers are considered AIDS-defining. An increased incidence in recent years, however, has been reported also for other malignancies after the introduction of HAART.We performed a case-control study to characterize all HIV-infected patients with both AIDS and non-AIDS-defining neoplasms observed among all consecutive patients followed at the Infectious Diseases Unit of Pescara General Hospital, since 1991 through 2012. All cases were matched with equinumerous controls without neoplasia homogeneous for age, sex and AIDS diagnosis.Out of 626 patients consecutively assisted since 1991, 57 cases of malignancy (9.1%) were observed. Of these, 45 (79.0%) occurred in males; mean age was 43.6\u00b19.3 years; 49 (86.0%) patients were diagnosed with AIDS. Tumors observed were: NHL, 17 (29.8%); SK, 13 (22.8%); HCC, 5 (8.8%); CPL, 6 (10.5%); Hodgkin's lymphoma, 4 (7.0%); solid tumors, 12 (21.1%), including 1 AIDS-defining tumor . Among these, 37 (66.1%) patients died; of them 14 (37.8%) had non-AIDS cancers. Cases were well matched with the 55 controls for sex (p=0.9), age (p=0.6) and AIDS diagnosis (p=0.6). In comparison with controls, CD4 nadirs were not different (153\u00b1151 in controls vs 136\u00b1154 cells/mmc), while CD4 at tumor diagnosis were very different between controls (463\u00b1283 cells/mmc) and cases . Among patients with malignancies, those who died had a non-significant reduction in CD4 counts (p=0.14); seemingly irrelevant were smoking status (p=0.9), working ability (p=0.4), HCV coinfection (p=0.4). Surprisingly, in patients co-infected with HBV, including HBsAg negative, antibody-positive subjects, tumors were significantly more frequent .Factors potentially relevant for carcinogenesis in the prolonged survival patients of the HAART era may include HBV coinfection in spite of the lack of active biochemical activity (HbsAg negative) in the majority of coinfected patients. The potential relevance of this finding deserves prompt assessment in a larger multicentric cohort."}
+{"text": "The third author's name appears incorrectly in the citation. The correct citation is: Talman V, Tuominen RK, Boije af Genn\u00e4s G, Yli-Kauhaluoma J, Ekokoski E (2011) C1 Domain-Targeted Isophthalate Derivatives Induce Cell Elongation and Cell Cycle Arrest in HeLa Cells. PLoS ONE 6(5): e20053. doi:10.1371/journal.pone.0020053"}
+{"text": "She was born small for gestational age with no catch-up growth. Congenital anomalies including right dysplastic kidney and large secundum atrial septal defect were present. She had delayed development with microcephaly and dysmorphism, including triangular facies, epicanthic folds, low set ears, micronagthia, and brachydactyly. Ophthalmological assessment showed astigmatism, myopia, and left exotropia requiring surgical correction. Audiology assessment showed bilateral mild conductive hearing loss. Thyroid function, morning cortisol, serum calcium and renal function tests were normal. Metabolic disease workup including lactate, plasma for amino acid, urine metabolic screen, VLCFA, transferrin isoelectrofocusing and clotting profile were normal. CT and MRI brain were normal. Serum IGF-1 at 13 years old was 62nmol/L (+ 0.5SD). Growth hormone study with glucagon showed peak growth hormone of 32mg/L. Bone age was 12 years at chronological age of 12-year-10month. Radiograph of both hands showed brachydactyly but no proximal implantation of 1st digit. Genetic study show normal karyotype, but MLPA and FISH later confirmed heterozygous terminal deletion of 15q26.2 with the genetic defect compatible with IGF-1 receptor mutation. Growth hormone therapy was refused by the patient and mother. Breast development started at 12-year-4month and menarche started at 13-year-5month of age. The growth spurt was absent with peak growth velocity of only 5.5cm/year. On the latest follow-up at 13-year-9month of age, her height was 128.8cm (-4.7SDS).A 5-year-9month old Chinese girl presented with severe growth retardation, height SDS -4.17 and body weight SDS -2.37. The mid-parental height was 151.2cm (3Our patient displayed typical phenotypic features of IGF-1 receptor mutation of heterozygous deletion of 15q26.2 with severe growth retardation. The condition could potentially benefit from growth hormone treatment according to recent literatures ["}
+{"text": "Other studies have reported high rates of depression and anxiety among human T-lymphotropic virus type 1 (HTLV-1) infected subjects, and have even suggested that HTLV-I causes psychiatric disease.We interviewed HTLV-I, HTLV-II and demographically similar HTLV seronegative blood donors with the Mini-International Neuropsychiatric Interview (MINI). Prevalences of major depression and generalized anxiety disorder in each group were calculated and compared to published U.S. population data. Adjusted odds ratios (aOR) and 95% confidence intervals (95% CI) controlling for educational achievement, alcohol intake and self-reported health status were calculated with multivariate logistic regression.Major depression was diagnosed in 5 (5.4%) of 93 HTLV-I positive subjects  and 17 (6.6%) of 256 HTLV-II positive subjects , compared to 12 (2.1%) of 585 HTLV seronegative blood donors. The prevalence of major depression among infected subjects was comparable to the 6.7% prevalence in the U.S. general population. Generalized anxiety disorder was diagnosed in 5 (5.4%) HTLV-I positive subjects  and 12 (4.7%) HTLV-II positive subjects (OR = 1.65 95% CI 0.68-4.01), compared to 15 (2.6%) seronegatives and 3.1% in the U.S. general population.We observed slightly higher prevalence of major depression and generalized anxiety disorder among HTLV-I and HTLV-II subjects that was not significantly elevated after controlling for health status and other confounding variables. Comparison to U.S. population data suggested that these findings are in part explained by a \u201chealthy blood donor effect\u201d among our controls."}
+{"text": "Improving trunk appearance is important for scoliotic patients. Selecting an appropriate rater is vital for effectively measuring this outcome. This study investigates whether a four-week intensive scoliosis-specific exercise programme results in improved patient body image and how this varies between patients, physiotherapists and an external scoliotic rater.82 patients  with IS and mean age 30.79 years (Range 10-81) were treated with a four-week intensive scoliosis-specific physiotherapy course (ScolioGold). Patients, 2 blinded physiotherapists and a blinded scoliotic rater rated patients\u2019 body-image before and after treatment. Body-image was assessed using a 0-10 scale, for 5 elements .Mean total scores before treatment were; patients 28.51 (SD8.76), physiotherapists 22.94 (SD6.01) and external rater 20.55 (SD7.27) and after treatment were; patients 15.46 (SD7.40), physiotherapists 13.73 (SD5.88) and external rater 6.61 (SD4.10). Differences in patients (P), physiotherapists (T) and external rater(E) body-image scores were found to have statistically significantly improved after treatment using Wilcoxon-signed rank . ICC scores between P&T, P&E and T&E were; fair (0.28), slight (0.19) and moderate (0.58) before treatment and fair (0.34), fair (0.28), moderate (0.59) after treatment.Statistically significant improvements between pre-post treatment scores substantiate intensive exercise methods (ScolioGold), in the treatment of IS-related negative body image. Significant variation between patients\u2019, physiotherapists\u2019 and external rater\u2019s scoring highlight the need for patients to rate body-image due to the subjectivity of this outcome measure, to ensure we adopt a client-centred care approach in our treatment goals to improve patient body-image."}
+{"text": "There was an error in the title. The correct title is: HIV-1 Tropism Determination Using a Phenotypic Env Recombinant Viral Assay Highlights Overestimation of CXCR4-Usage by Genotypic Prediction Algorithms for CRF01_AE and CRF02_AG. The correct citation is: Mulinge M, Lemaire M, Servais J-Y, Rybicki A, Struck D, et al. (2013) HIV-1 Tropism Determination Using a Phenotypic Env Recombinant Viral Assay Highlights Overestimation of CXCR4-Usage by Genotypic Prediction Algorithms for CRF01_AE and CRF02_AG. PLoS ONE 8(5): e60566. doi:10.1371/journal.pone.0060566."}
+{"text": "Therefore, many individuals are capable of mitigating the presence of autoantibodies and avoiding clinical disease. The objective of this study was to investigate unique features of SLE patients compared with autoantibody-positive (aAbn = 790) were screened by multiplex, bead-based assays for autoantibodies against: dsDNA, chromatin, ribosomal P, SS-A/Ro, SS-B/La, Sm, Sm/RNP, RNP, SCL-70, Jo-1, and centromere B. Sera from aAb+ individuals, matched aAb- controls and SLE patients were tested for 52 cytokines using multiplex bead-based assays and ELISAs. Hierarchical clustering was performed and Kruskal-Wallis tests with Dunn's multiple comparisons were compared including a false discovery rate. Immune cell phenotyping and phospho-flow cytometry was performed on peripheral blood mononuclear cells from aAb+ and aAb- healthy individuals. Unpaired t tests and Mann-Whitney tests were performed.Individuals  and matched aAb- healthy controls and SLE patients were selected for further analysis. While aAb+ healthy individuals displayed some similar cytokine patterns to SLE patients, they also displayed a suppressed cytokine profile that included decreased T-cell cytokines (IFN\u03b3 (P < 0.05), IL-5 (P < 0.05), IL-17F (P < 0.01)), decreased B-lymphocyte stimulator levels (P < 0.05), and increased IL-1 receptor antagonist levels (P < 0.001). An increased percentage of B cells was found in aAb+ healthy individuals compared with aAb- healthy controls (P = 0.039) that consisted of an increased percentage of memory B cells (P = 0.034) and a decreased percentage of transitional B cells (P = 0.028). Compared with aAb- healthy controls, B cells from aAb+ healthy individuals showed significantly increased pSTAT1 and pSTAT3 in response to IFN\u03b1 stimulation (P = 0.037 and P = 0.040), increased pSTAT1 in response to IFN\u03b3 stimulation (P = 0.018), and increased pSTAT3 in response to IL-21 stimulation (P = 0.041). CD4+ T cells from aAb+ healthy individuals showed decreased pSTAT3 in response to IFN\u03b3 and IL-2 stimulation (P = 0.018 and P = 0.005). IFN\u03b1, IFN\u03b3 and IL-6 stimulation significantly increased pSTAT signaling in monocytes from aAb+ healthy individuals.Of the screened individuals, 57 individuals were positive for at least one autoantibody (7.2%), with 33.3% being Native American, 57.9% European American, 8.8% African American, and 89.5% female. European American aAb+ healthy individuals exhibit features of inflammation and loss of immune tolerance, they are capable of suppressing these responses by regulatory mechanisms probably no longer functional in patients with autoimmune disease.Although aAb"}
+{"text": "There was an error in the third subheading of the Results section (\"African-Americans and Caucasians\"). The correct sentence is: Single locus tests of association in the African-Americans identified seven significant allelic associations at IL12B polymorphisms rs3212227 (p = 0.002), rs2421047 (p = 0.008), rs2288831 (p = 0.008), rs10631390 (p = 0.001), rs3212220 (p = 0.003), rs6894567 (p = 0.007), and rs17860508 (p = 0.002) and three significant genotypic associations at rs3212227 (p = 0.05), rs10631390 (p = 0.05) and rs6894567 (p= 0.03) (Table 2b).&rdquo;"}
+{"text": "Jatropha curcas leaf extracts.Drug-resistant HIV, a major global concern, warrants the development of novel anti-virals as alternative and inexpensive therapy. In the current study, we isolated potentially drug-resistant HIV and assessed previously unreported anti-viral activity of In vitro micro-co-culture was employed for virus isolation followed by drug susceptibility assays to determine resistance to Azidothymidine (AZT) and Lamivudine (3TC).Jatropha curcasleaves were extracted using Soxhlet apparatus. Methanolic (ME) and aqueous (AE) extracts were chosen for further study. Secondary metabolites were detected by High-Performance Thin Laye rChromatography and in vitro cytotoxicity established by MTT assay. Anti-viral activity was evaluated by p24 inhibitionin post- and pre-infection interaction studies.50 values ranging from 0.001418-82.73 \u00b5M AZT and 2.645-15.35 \u00b5M 3TC.Seven HIV isolates were obtained (isolation rate: 23.33%) with drug IC50 values were 32.07 mg/mL AE and 35.5 mg/mL ME.Tannins, flavonoids, saponins were detected in AE and flavonoids, saponins in ME while CC50 values were ranging from 0.0255-0.4137 mg/mL AE and 0.00073-0.1278 mg/mL ME; pre-infection studies (1 isolate) showed 100% p24 inhibition by ME and 97.19% p24 inhibition by AE at 25 mg/mL each.In post-infection studies (4 isolates), ICJatropha curcas leaf extracts showed effective anti-viral and probable entry inhibition activity against potentially drug-resistant HIV, which has not been reported earlier. We conclude that Jatropha curcasis a good candidate for anti-HIV therapy with further research.HIV isolates potentially resistant to AZT/3TC were obtained; genotypic drug resistance is being ascertained."}
+{"text": "Significance of blood glucose (BG) control in ICU patients, especially in terms of mortality reduction, has been recognized in recent years. However, the relationship between BG profile and the severity, as well as BG target, is not clearly elucidated. A preliminary study was performed in order to clarify the relationship between BG profile and the severity of ICU patients.r) between the abovementioned SOFA scores and BG parameters were calculated using two-dimensional and linear regression analysis .Forty-seven patients were studied. The following parameters were calculated during first week after ICU admission. (1) Mean and maximum value of SOFA scores . (2) Mean, standard deviation, maximum, minimum, and difference of BG levels , respectively). BG levels were measured basically every 6 hours with capillary blood. (3) Correlation coefficients ((1) rt and rl (rt/rl) between SOFAmax and BG parameters: BGsd (0.48/0.36), BGmax (0.47/0.33), BGd (0.47/0.35), BGm (0.30/0.22), BGmin (0.21/0.36). (2) (rt/rl) between SOFAm and BG parameters: BGsd (0.45/0.33), BGmax (0.45/0.28), BGd (0.45/0.30), BGm (0.28/0.17), BGmin (0.17/0.29).(1) Variable and maximum BG levels during the first week , rather than mean and minimum BG levels , were related to the severity. (2) Suppression of the higher variable and maximum BG levels was considered to link to better outcome. (3) On the other hand, part of the severe patients seemed to have the lowest of those variable and maximum BG levels, judging from two-dimensional or parabolic correlations between those BG parameters and SOFA scores."}
+{"text": "Reasons for screen failures are evaluated for three phase 1 HIV vaccine clinical trials recruiting healthy low-risk participants at the Perinatal HIV Research Unit: SAAVI102/HVTN073 and SAAVI103/HVTN086  and IAVIB003/HVTN091 (Ad26 and Ad35 ENV vaccines). Recruitment strategies involved a pre-screening programme, clinics and community outreach.Protocol-specific eligibility was determined using assessments of understanding, risk behaviour, medical history, physical examination, blood and urine testing, and for HVTN073 and 086, electrocardiograms. Descriptive analysis and multivariate logistic regression of age, trial group and gender were performed.Between June 2009-2012, 225 participants, median age 22 years(IQR:20-25) were screened. Overall 53% were ineligible, 60% of females vs. 51% of males (p=0.2). Site screening-to-enrolment ratios for 073, 086 and 091 were 2.1:1, 2.3:1 and 1.7:1 respectively. Medical abnormalities contributed 59% (n=70) of ineligibility reasons, chiefly urine abnormalities , abnormal ECG (n=12), raised liver enzymes (n=10), raised blood pressure (n=9), low white cells (n=8) and hepatitis B (HBsAg+ve) or C (anti HCV+) (n=7). Other criteria excluded 41%(n=49) e.g. incomplete screening before enrolment closure (n=16), high-risk sexual behaviors (n=15), inability to comply with protocol (n=11), enrolment in another study (n=3), substance abuse , and poor understanding (n=2). In multivariate analysis, increasing age  predicted ineligibility but gender did not . HVTN073&086 participants were more likely to be ineligible than HVTN091 .Screen failures in phase 1 vaccine trials in Soweto provide young people opportunities for care, especially through blood pressure, urine, risk behaviour and hepatitis B/C screening. Older participants and those in protocols stipulating ECG criteria were more likely to fail screening."}
+{"text": "Gillisia limnaea Van Trappen et al. 2004 is the type species of the genus Gillisia, which is a member of the well characterized family Flavobacteriaceae. The genome of G. limnea R-8282T is the first sequenced genome (permanent draft) from a type strain of the genus Gillisia. Here we describe the features of this organism, together with the permanent-draft genome sequence and annotation. The 3,966,857 bp long chromosome (two scaffolds) with its 3,569 protein-coding and 51 RNA genes is a part of the  Genomic Encyclopedia of Bacteria and Archaea project. T (= DSM 15749 = LMG 21470 = CIP 108418) is the type strain of the species Gillisia limnaea . The. TheG. ln 4.9 \u00b5m , which i strains -34. MotiT are iso-C15:0, anteiso-C15:0, iso-C15:1, iso-C16:0, C17:0 2-OH, iso-C17:0 3-OH, iso-C17:1 \u03c99c, anteiso-C17:1 \u03c99c and summed feature 3 (containing iso-C15:0 2-OH and/or C16:1 \u03c97c) [T.The principal cellular fatty acids of strain R-8282 Genomic Encyclopedia ofBacteria andArchaea project [This organism was selected for sequencing on the basis of its phylogenetic position , and is  project . The gen project  and the G. limnaea strain R-8282T, DSM 15749, was grown in DSMZ medium 514 (BACTO Marine Broth) [et al. 2009 [e Broth)  at 20\u00b0C.al. 2009  for optial. 2009 .The genome was sequenced using a combination of Illumina and 454 sequencing platforms. All general aspects of library construction and sequencing can be found at the JGI website . PyroseqGenes were identified using Prodigal  as part The genome consists of two scaffolds with 3,558,876 bp and 407,981 bp length, respectively, with a G+C content of 37.6% . Of the G. limnaea R-8282T revealed the presence of three rhodopsin genes related to proteorhodopsin  and xanthorhodopsin  protein-encoding sequences, whereas a third rhodopsin protein sequence  seems to be truncated. Another finding was a set of genes involved in \u03b2-carotene biosynthesis, together with a gene encoding a \u03b2-carotene 15,15'-monooxygenase , an enzyme that oxidatively cleaves \u03b2-carotene into two molecules of retinal, which is necessary for rhodopsin function. PRs and XRs are photoactive transmembrane opsins that bind retinal and which belong to the microbial rhodopsin superfamily [E. coli cells for ATP synthesis [Salinibacter ruber M31T [Bacteria [Archaea [Genome analysis of erfamily . When exerfamily . This prynthesis  as well ynthesis . In contynthesis ,52, thatber M31T ,54. Its ber M31T , resultiBacteria ,57 or ba[Archaea .Flavobacteriaceae  [Krokinobacter sp. 4H-3-7-5 (AEE18495) [Dokdonia [The gene encoding the putative XR (Gilli_2340) of strain R-8282HQ04368)  and KrokEE18495) , which wDokdonia ,66 , indicating that Gillisia sp. strain CBA3202 does not belong to the species G. limnaea.Interestingly, no rhodopsin-encoding sequence could be detected in the genome sequence of  CBA3202 , which w CBA3202 . Digital CBA3202  between Bacteroidetes phylum were frequently found attached to aggregates [T several genes were detected that are thought to be involved in gliding motility (gldA (Gilli_1140), gldB (Gilli_2923), gldC (Gilli_2942), gldD (Gilli_1840), gldE (Gilli_1841), gldF (Gilli_3447), gldG (Gilli_3446), gldH (Gilli_2158), gldI (Gilli_0258), gldJ (Gilli_1638), gldK (Gilli_2747), gldL (Gilli_2748), gldM (Gilli_2749), gldN (Gilli_2750), espA (Gilli_3049), espB (Gilli_3050), remB (Gilli_2697), sprA (Gilli_2693) and sprE (Gilli_2130)). This observation indicates the possible gliding motility of strain R-8282T, but has never been reported in literature.Compared to free-living bacteria, representatives of the gregates  and durigregates ,71. Theygregates ,73. In s"}
+{"text": "There was an error in the title.The correct version of the title is: Specific Inhibition of the Redox Activity of Ape1/Ref-1 by E3330 Blocks Tnf-\u03b1-Induced Activation of Il-8 Production in Liver Cancer Cell LinesThe correct citation is: Cesaratto L, Codarin E, Vascotto C, Leonardi A, Kelley MR, et al. (2013) Specific Inhibition of the Redox Activity of Ape1/Ref-1 by E3330 Blocks Tnf-\u03b1-Induced Activation of Il-8 Production in Liver Cancer Cell Lines. PLoS ONE 8(8): e70909. doi:10.1371/journal.pone.0070909"}
+{"text": "Eradicating cHCV is necessary for the subsequent management of patients with HIV and every HIV/HCV co-infected patient should be considered for treatment. We describe differences between cHCV mono-infected and cHCV/HIV co-infected patients in baseline factors and outcome of cHCV-treatment with peginterferon alfa 2a + RBV in the worldwide largest cHCV cohort.Noninterventional prospective multicenter German cohort, started January 2008 and still recruiting. Interim analysis of cHCV patients, stratified for cHCV mono-infection and cHCV/HIV co-infection. The results are based on a cross-sectional analysis of all available data in April 2010.2, na\u00efve/relapse/non-responder/re-infection: 86.4/3.8/4.5/5.3(CI), 88.0/6.1/5.3/0.6 (MI) %, source of infection (>1 answer possible): iv drug use 25.2(CI), 44.9(MI) %, sexual transmission 60.7 (CI), 4.1(MI)%, other 8.0 (CI), 24.0 (MI), unknown 13.1(CI), 33.0 (MI)%. 86.4 % of the co-infected patients received antiretroviral HIV-treatment (ART), 66.2 % of them had an HIV-RNA level below 50 copies/mL, median CD4-cells/\u00b5 count was 502. From those patients who finished treatment, 52.9% of the CI-Group and 67.7% of the MI-Group completed the planned course. Reasons for discontinuation (>1 answer possible) were non-response  and patient request . Other reasons were tolerability  and compliance issues .Treatment response rates, stratified by genotypes, were already available regarding RVR and EVR  and 4.993 cHCV mono-infected (MI). Main baseline- characteristics: 85.9% were GT1/4/5/6 patients in the CI-Group and 63.4% in the MI-Group, age was 41.0 (CI), 42.0 (MI) yrs, 89.7 (CI), 62.9 (MI)% were male, BMI was 22.8 (CI), 24.9 (MI) kg/mIn this preliminary analysis HCV/HIV co-infected patients seemed to respond similar according to RVR and EVR in GT1/4/5/6 as HCV-mono infected patients on HCV-treatment. Treatment discontinuation due to non-response and patient request was much more common in the co-infected group. Other reasons for discontinuation like tolerability and compliance are equal in both arms.A more detailed analysis, in particular the influence of the HIV-ART on HCV-therapy outcome, may help interpreting these data. An updated analysis will be presented."}
+{"text": "Radical prostatectomy (RP) is frequently responsible for erectile dysfunction (ED). Post-RP patients often show insufficient response or treatment failures to PDE5 inhibitor therapy. This study was undertaken to evaluate the acute effects of the soluble guanylate cyclase (sGC) stimulator, BAY 60-4552 and vardenafil administered alone or in combination on erectile responses to electrical stimulation of the cavernous nerve (ES-CN) in rats with cavernous nerve (CN) crush injury-induced ED.Male Sprague-Dawley rats underwent laparotomy  or bilateral CN crush injury (n=57). After 3 weeks of recovery, erectile function was evaluated in urethane-anesthetized rats following ES-CN at different frequencies. The acute effects of intravenous (iv) injection of vehicle, vardenafil 0.03 mg/kg, BAY 60-4552 0.03 mg/kg or 0.3 mg/kg, or a BAY 60-4552 0.03 mg/kg + vardenafil 0.03 mg/kg combination were evaluated in CN crushed rats.Bilateral CN crush injury followed by a 3-week recovery period decreased erectile responses to ES-CN by about 50%. In CN crushed rats, both iv vardenafil 0.03 mg/kg and BAY 60-4552 at the tested dosings (0.03 or 0.3 mg/kg) increased erectile responses to ES-CN to the same extent: \u0394ICP/MAP at 10Hz ES-CN was 20.9\u00b1 1.3 % after iv vehicle, 25.3\u00b1 3.3 % (P<0.001) after iv vardenafil, and 26.3\u00b1 4.9 % and 26.6\u00b1 5.2 % after BAY 60-4552 0.03 mg/kg (P<0.01) and 0.3 mg/kg (P<0.001) respectively. The combined iv administration of vardenafil and BAY 60-4552 in CN crushed rats exerted additive effects and totally restored erectile responses to ES-CN equivalent to sham rats .The present study supports the concept that the combined administration of a sGC stimulator, BAY 60-4552 and vardenafil provides additive beneficial effects. Thus this combination could become a novel treatment option in ED patients with cavernous nerve injury showing insufficient response or treatment failures to PDE5 inhibitors."}
+{"text": "AbstractNazeris Fauvel collected from Nabanhe Nature Reserve, Yunnan Province, are described under the names of Nazeris nabanhensis sp. n. and Nazeris caoi sp. n. The male sexual characters are described and illustrated. A key to the Nazeris species of Yunnan is provided. A map of the collecting sites is given.Two new species of the genus  Nazeris Fauvel (1873: 298) can be readily distinguished from other Paederinae by the labrum having four teeth at the front margin and the bi-lobed 4th tarsal segments. Up to the present, 51 species and subspecies of Nazeris have been recorded from China. Yunnan is a mountainous province located in Southwest China, from PageBreakwhich nine species of Nazeris have been described: Nazeris zhangi Watanabe & Xiao (1993: 130) from \u201cYu\u2019an-shan near Kunming City\u201d, Nazeris giganteus Watanabe & Xiao (1997:2) and Nazeris daliensis Watanabe & Xiao (1997: 7) from \u201cDiancang shan Mts., Dali shi\u201d, Nazeris alpinus Watanabe & Xiao (1997: 5) from \u201cMt. Yulongxue shan, Lijiang County\u201d, Nazeris jizushanensis Watanabe & Xiao (1997: 9) from \u201cMt. Jizu Shan, Binchuan County\u201d, Nazeris baihuaensis Watanabe & Xiao (2000: 312), Nazeris ishiianus Watanabe & Xiao (2000: 319) and Nazeris nomurai Watanabe & Xiao (2000: 316) from \u201cGaoligong Shan Mts., Baoshan area\u201d, Nazeris huanxipoensis Watanabe & Xiao (2000: 318) from \u201cHuanxipo, Tengchong Xian\u201d. Only two species have become known from the three countries adjacent to Yunnan : Nazeris coomani Jarrige (1948: 40) (redescribed by Nazeris odzisan Watanabe (1996: 1) from \u201cMt. Tam Dao, Vinh Phu Prov.\u201d (Vietnam).The genus Nazeris nabanhensis sp. n. and Nazeris caoi sp. n. The male sexual characters of the two new species are described and illustrated. A map .Nazeris species was not examined. The characters used in comparative remarks and keys are according to descriptions of ; postocular portion 1.96 times as long as eye length; punctation coarse, dense, and umbilicate; interstices reduced to narrow ridges. Antennae slender; relative length of each segment from 1 to 11: 42.0 : 15.0 : 29.0 : 23.0 : 22.5 : 22.0 : 19.0 : 17.0 : 15.5 : 14.0 : 20.0; relative width of each segment from 1 to 11: 12.0 : 7.5 : 6.0 : 6.0 : 6.0 : 6.0 : 6.0 : 6.0 : 6.0 :7.5 : 7.5.convex, oval, longer than wide (length/width = 1.20), narrower (pronotum/head = 0.93) and shorter (pronotum/head = 0.94) than head; prosternum with strong longitudinal median carina, which disappears behind anterior margin. Elytra shorter than wide (length/width = 0.91), distinctly shorter (elytra/pronotum = 0.74) and slightly narrower (elytra/pronotum = 0.97) than pronotum.PageBreakdian lobe in dorsal view tri-lobed, median part cone-shaped, two outer parts acute at apices, with little agnail in each outer side near apex; dorso-lateral apophyses slightly curved inward, distinctly widened near apex, extending beyond apices of median lobe.elongate, tergites without any microsculpture. Seventh sternite  trapezoiSeventh and 8th sternites simple. The other characters are similar to those of male.Nazeris daliensis Watanabe (1997: 7) from Yunnan Province in appearance, but it can be distinguished from the latter by the following characters: elytra slightly narrower than pronotum ; depth of excision of male 8th sternite nearly half of middle length of sternite ; apical part of median lobe of aedeagus in dorsal view tri-lobed . The new species can be distinguished from Nazeris coomani Jarrige (1948: 40) from Vietnam by head with umbilicate punctation (in Nazeris coomaniPageBreak punctation of head simple) and distinguished from Nazeris odzisan Watanabe (1996: 1) from Vietnam by elytra shorter than wide (in Nazeris odzisan elytra longer than wide); dorso-lateral apophyses of aedeagus extending beyond apices of median lobe (in Nazeris odzisan not extending to apices of median lobe).The new species is similar to The specific name is derived from the name of the type locality: Nabanhe Nature Reserve.urn:lsid:zoobank.org:act:3B251645-1B6B-4388-ACEC-4E5B42C4A858PageBreakParatypes 1 male, same data as holotype; 1 female, Jinghong City, Nabanhe Nature Reserve, Bengganghani, 1,900m, 1. V. 2009, Hu Jia-Yao & Yin Zi-Wei leg. SHNUC.CHINA: Holotype: Yunnan Prov.: male, Jinghong City, Nabanhe Nature Reserve, Bengganghani, 1,930m, 14. XI. 2008, Hu Jia-Yao & Tang Liang leg. Body length: 6.1\u20136.4 mm; forebody length: 3.3\u20133.6 mm. elongatesuborbicular, slightly longer than wide (length/width = 1.07); postocular portion 2.14 times as long as eye length; punctation coarse, dense, and umbilicate; interstices reduced to narrow ridges. Antennae slender; relative length of each segment from 1 to 11: 42.0 : 13.5 : 30.0 : 23.0 : 21.0 : 21.0 : 19.0 : 18.0 : 18.0 : 16.0 : 22.0; relative width of each segment from 1 to 11: 10.5 : 7.0 : 6.5 : 6.0 : 5.5 : 6.0 : 5.5 : 5.5 : 6.0 :6.5 : 7.0.PageBreak strong longitudinal median carina, which disappears behind anterior margin. Elytra slightly shorter than wide (length/width = 0.97), distinctly shorter (elytra/pronotum = 0.80) and slightly narrower (elytra/pronotum = 0.97) than pronotum.convex, oval, longer than wide (length/width = 1.19), narrower (pronotum/head = 0.88) and shorter (pronotum/head = 0.98) than head; prosternum withelongate, tergites without any microsculpture; densely and coarsely punctate. Seventh sternite  distinctSeventh and 8th sternites simple. The other characters are similar to those of male.Nazeris nabanhensis sp. n. from the same locality, but can be distinguished from the latter by the following PageBreakcharacters: postocular part more than twice as long as longitudinal diameter of each eye ; median lobe of aedeagus in dorsal view bi-lobed (in Nazeris nabanhensis tri-lobed); dorso-lateral apophyses of aedeagus not extending to apices of median lobe (in Nazeris nabanhensis extending beyond apices of median lobe). The new species can be distinguished from Nazeris coomani Jarrige (1948: 40) from Vietnam by head with umbilicate punctation (in Nazeris coomani punctation of head simple), and distinguished from Nazeris odzisan Watanabe (1996: 1) from Vietnam by elytra shorter than wide (in Nazeris odzisan elytra longer than wide); median lobe of aedeagus in dorsal view bi-lobed (in Nazeris odzisan not lobed).The present new species is similar in appearance to The species is named in honor of Mr. Guanghong Cao of Nabanhe Nature Reserve, who helped us a lot during field work."}
+{"text": "There was an error in the citation. The correct citation is: Aka JA, Lin S-X (2012) Comparison of Functional Proteomic Analyses of Human Breast Cancer Cell Lines T47D and MCF7. PLoS ONE 7(2): e31532. doi:10.1371/journal.pone.0031532"}
+{"text": "The fifth author's name was spelled incorrectly. The correct name is: Max Schelker. The corrected citation is:Raue A, Schilling M, Bachmann J, Matteson A, Schelker M, et al. (2013) Lessons Learned from Quantitative Dynamical Modeling in Systems Biology. PLoS ONE 8(9): e74335. doi:10.1371/journal.pone.0074335In equation 7, many additional factors of 2 were added incorrectly. Please see the correct equation 7 here: In equation 8, '2A_0' should only read 'A_0.' Please see the correct equation 8 here:"}
+{"text": "Neonatal mice developed neurological disease and pulmonary dysfunction after an infection with a mouse-adapted human Enterovirus 71 (EV71) strain MP4. However, the hallmark of severe human EV71 infection, pulmonary edema (PE), was not evident.To test whether EV71-induced PE required a proinflammatory cytokine response, exogenous pro-inflammatory cytokines were administered to EV71-infected mice during the late stage of infection.After intracranial infection of EV71/MP4, 7-day-old mice developed hind-limb paralysis, pulmonary dysfunction, and emphysema. A transient increase was observed in serum IL-6, IL-10, IL-13, and IFN-\u03b3, but not noradrenaline. At day 3 post infection, treatment with IL-6, IL-13, and IFN-\u03b3 provoked mild PE and severe emphysema that were accompanied by pulmonary dysfunction in EV71-infected, but not herpes simplex virus-1 (HSV-1)-infected control mice. Adult mice did not develop PE after an intracerebral microinjection of EV71 into the nucleus tractus solitarii (NTS). While viral antigen accumulated in the ventral medulla and the NTS of intracerebrally injected mice, neuronal loss was observed in the ventral medulla only.Exogenous IL-6, IL-13, and IFN-\u03b3 treatment could induce mild PE and exacerbate pulmonary abnormality of EV71-infected mice. However, other factors such as over-activation of the sympathetic nervous system may also be required for the development of classic PE symptoms. Enterovirus genus of Picornaviridae family. In general, EV71 infections are mild, such as hand, foot, and mouth disease and herpangina in young children. However, central nervous system (CNS) infections with life-threatening pulmonary and cardiac complications have occurred [Enterovirus 71 (EV71), a highly neurotropic, positive-sense single-stranded RNA virus, belongs to the occurred .Pulmonary edema (PE) and subsequent rapid onset cardiopulmonary failure are hallmarks of EV71 induced mortality . EV71-inOur previous studies showed that a mouse-adapted EV71 strain, EV71/MP4, could experimentally infect laboratory mice via oral (p.o.), intramuscular, and intracranial (i.c.) inoculation routes, resulting in CNS infection and death . Clinica7 PFU/ml. MP4 strains were tested by monoclonal antibodies for EV71 (mAb3324), EV71/coxsackievirus A16 (mAb3323), coxsackievirus A24 (mAb3302), coxsackievirus B3 (mAb3306), echovirus 9 (mAb3313), PV type 1 (mAb3331), PV type 2 (mAb3332), and PV type 3 (mAb3335)  using indirect immunofluorescence staining of infected RD cell cultures, and RT-PCR using specific primers for EV71. HSV-1/KOS, a clinical isolate strain of human simplex virus-1 (HSV-1)  was grown in Vero cells.Rhabdomyosarcoma (RD) cells  were maintained in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum, 2 mM L-glutamine, penicillin and streptomycin. Vero cells (ATCC) were maintained in DMEM containing 5% newborn calf serum, penicillin and streptomycin. Mouse-adapted EV71 strain MP4  was prop5 PUF/mouse) or HSV-1/KOS (2 \u00d7 102 PFU/mouse) through the fontanelles using a 26-gauge needle. Control mice were given culture medium. To systemically increase proinflammatory cytokine levels, mice were intraperitoneally (i.p.) injected at three days post-inoculation with recombinant mouse IL-6, IL-13, and IFN-\u03b3   2 or 3 times within 48 h. Control mice were given bovine serum albumin (BSA). Mice were observed twice daily for clinical signs and mortality. Lung function and body weight monitored daily for one week. Lung weight/body weight analysis, histopathology, and immunohistochemistry were performed after these experiments. For i.c. microinjection, EV71/MP4 was bilaterally microinjected into the nucleus tractus solitarii  of 8-week-old ICR mice in a volume of 0.2 \u03bcl (3 \u00d7 107 PFU/ml) over 10 min through a 27-gauge stainless steel cannula connected to a 10-\u03bcl Hamilton syringe  driven by an injection pump . Mice were observed twice daily for clinical signs and mortality for 5 days. Clinical disease was scored as follows: 0, healthy; 1, ruffled fur and hunchbacked appearance; 2, wasting; 3, limb weakness; 4, limb paralysis; 5, moribund and death. The Institutional Animal Care and Use Committee approved all animal protocols.Specific-pathogen-free, 7-day-old ICR mice  were i.c. injected with 10 \u03bcl of EV71/MP4 . Readings were collected for 3 min after 3 min of resting in the chamber.After anesthetization with pentobarbital sodium , blood was collected after axilla dissection. The levels of IL-6, IL-10, IL-13, and IFN-\u03b3 in serum and noradrenaline in plasma of EV71-infected mice were determined using ELISA kits  and a noradrenaline EIA kit , respectively according to the manufacturer's instructions. The detection limits for IL-6, IL-10, IL-13, IFN-\u03b3, and noradrenaline were 15.6, 31.25, 62.5, 31.25, and 0.027 pg/ml, respectively.Changes in vascular permeability in lungs were determined by measuring the increment of wet lung weight. Individual lung lobes were removed, blot-dried on gauze, and weighed using a balance with accuracy to 0.0001 gram. Data were expressed as the ratio of total wet lung weight to body weight.Animals were perfused with isotonic saline containing EDTA. The whole brains and lungs were removed and weighed. Half of the tissues were immersion fixed in 10% buffered formalin for 48 h, bisected, embedded in paraffin, and stained with hematoyxlin and eosin (H & E) or Nissl stain. The remaining tissue samples were frozen in a liquid nitrogen-cold hexane bath in 100% OCT compound . All samples were stored at -70\u00b0 C until assayed.The presence of EV71 virions in cryosections of frozen tissues  was visualized by a previously described method [U tests. The results are expressed as means \u00b1 standard errors of the means (S.E.M.). A P value of < 0.05 was considered significant.The clinical scores, lung weight/body weight ratio, and numbers of EV71-positive and NeuN-positive cells were analyzed using either the nonparametric one-way analysis of variance (ANOVA) or Mann-Whitney 5 PFU/mouse), 7-day-old mice did not gain weight and developed clinical symptoms with diffuse emphysema but no evidence of PE in the lung , IL-13 (0.15 \u03bcg/mouse), and IFN-\u03b3 (0.8 \u03bcg/mouse) to mice 3 days after an i.c. inoculation of EV71 (two doses within 48 h). At this time the animals developed mild clinical symptoms with the presence of virus in the CNS. All the mock-infected BSA-treated mice gained weight normally Figure  without P = 0.0054, Figure P = 0.0318, Figure 2 PFU/mouse) prior to the administration of IL-6, IL-13, and IFN-\u03b3. HSV-1 is a neurotropic virus and its infection is known to cause diffuse encephalitis in mice, mainly in tissues of the cerebrum and cerebellu [Weight loss in EV71-infected mice resulted in increased lung weight/body weight ratios, as compared with non-infected control mice . Immunostaining showed that EV71-positive cells accumulated at the ventral medulla (VM) over time Figure  and 4B. In the course of the development of a mouse model of human EV71 infection, we demonstrated that EV71 could spread from the gastrointestinal tract or muscle to and accumulate in the CNS, especially in the brainstem of mice after different routes of inoculation . FollowiClinical findings showed that increases in serum IL-1\u03b2, IL-6, IL-10, IL-13, TNF-\u03b1, and IFN-\u03b3 were associated with PE development in EV71 patients . In thisBased on the distribution of viral antigens and viral genomic sequences, EV71 is likely propagated throughout the CNS via motor pathways . Our preSurprisingly, PE did not developed in adult mice after a direct inoculation of EV71 to the NTS, an area known to contribute to the development of PE in rats upon injury ,25. In oBesides an over-stimulated proinflammatory response, sympathetic excitement has been proposed as another mechanism involved in the development PE in EV71 patients , other aCollectively, we demonstrated that IL-6, IL-13, and IFN-\u03b3 over-stimulation exacerbated pulmonary dysfunction and clinical symptoms, and provoked a mild PE in EV71-infected mice. A synergistic proinflammatory cytokine response and damage to specific brain regions, or more precisely specific neuronal cells, may be necessary for the development of EV71-induced PE.BSA: bovine serum albumin; EV71: Enterovirus 71; HSV-1: herpes simplex virus-1; PIF: peak of inspiratory flow; PEF: peak of expiratory flow; LC: Locus coeruleus; MRI: magnetic resonance imaging; MV: minute volume; NTS: nucleus tractus solitarii; PV: poliovirus; PE: pulmonary edema; TV: tidal volume.The authors declare that they have no competing interests.SWH participated in the design of the study, carried out the animal studies, performed the statistical analysis, and drafted the manuscript. YPL participated in the design of the study. YTH carried out the immunostaining studies. CHL carried out the intracerebral microinjection studies. JIC participated in design and coordinate the immunostaining studies. HYL participated in the design of the study. IJS participated in the design of the study. CKY conceived of the study, and participated in its design and coordination and helped to draft the manuscript. All authors read and approved the final manuscript.Figure 1S. Intracranial inoculation of EV71 resulted in CNS infection, clinical disease, and emphysema in mice. Seven-day-old ICR mice (n = 24) were intracranially inoculated with or without EV71/MP4 strain (4 x105 PUF/mouse). Body weight (A) and clinical disease (B) were then monitored daily after infection. Lung tissues were collected for general morphological analysis (H & E stain) . Clinical disease was scored as followed: 0, healthy; 1, ruffled hair, hunchbacked or reduced mobility. 2, wasting; 3, limb weakness. 4, limb paralysis. 5, death. Bars: 1.0 mm. Note moderate emphysema but not pulmonary edema was observed in the EV71-infected mice (arrowheads).Click here for fileFigure 2S. Exogenous IL-13 treatment exacerbated pulmonary dysfunction in EV71-infected mice. Seven day-old ICR mice (n = 12) were intracranially inoculated with EV71 (4 \u00d7 105 PFU/mouse) followed by an intraperitoneal injection IL-13 at days 3 and 4 post inoculation. Change in pulmonary functions of mice were monitored daily. Ctrl: culture medium; BSA: bovine serum albumin; PIF: peak of inspiratory flow; PEF: peak of expiratory flow; MV: minute volume; TV: tidal volume. Data represent means \u00b1 S.E.M. *, P < 0.05 and **, P < 0.01.Click here for file"}
+{"text": "Lipoic acid (LA) stimulates production of an immunomodulatory molecule, cAMP, that may have therapeutic benefit in multiple sclerosis (MS). The study sought to determine the bioactivity of two forms of oral LA, racemic (R/S-LA) and R-LA, to identify factors that may improve the therapeutic effectiveness of LA in MS.Participants met the following criteria: age 18-70 years and a definite multiple sclerosis diagnosis. Consenting participants were randomized to 2 groups: R/S-LA (n=20) or R-LA (n=8). Blood was collected at baseline, 5, 10, 15, 30, 60, 90, 120, 180, 240, and 300 minutes after ingestion of a single 1200 mg LA dose. Bioactivity was determined by measuring immune cell cAMP levels (pmol/mg) at baseline and 240 minutes post ingestion. Group differences were analyzed by repeated measures analysis of variance (cAMP) and t-test (pharmacokinetics).For R/S-LA, mean baseline cAMP levels were 8.6 (SE 0.83) pmol/mg protein, which increased to 14.1 (SE 1.7) 240 minutes post-ingestion. For R-LA, baseline cAMP levels were 5.7 (SE 0.54), which decreased to 4.7 (SE 0.41) 240 minutes post-ingestion. The mean change in cAMP levels were different between groups, p<0.01. The two groups showed no differences in serum LA levels at 240 minutes (p=0.29), but showed a difference in the mean AUC (min*mcg/ml): R/S-LA is 681.8 (SE 83.4) and R-LA is 389.5 (SE 43.9) (p<0.01). Tmax (minutes) between groups differed: R/S-LA was 81.0 (SE 8.9) and R-LA was 13.1 (SE 2.7); p<0.001.At 1200 mg oral dose and comparable serum LA levels at T240, R/S-LA showed higher cAMP levels compared to R-LA. Since R-LA had an earlier Tmax, it is plausible that increases in cAMP may have occurred at earlier undetected time-points. This pilot study warrants further investigation as differences between R-LA and R/S-LA in bioactivity and pharmacokinetics would impact clinical trial design and patient care."}
+{"text": "Elevated signal intensity (SI) in T2w images is often designated as area-at-risk (AAR) following acute/sub-acute MI. While AAR has been qualitatively associated with reductions in wall thickening (WT), quantitative information between the two is not available -3. QuantIn an animal model, quantify the:1) Effect of SI threshold on estimating AAR,2) Reduction in WT with AAR at increasing SI thresholds, and3) Correlation between AAR and percent scar.Acquisition Protocol: Basal, mid, and apical slices in short axis orientation were obtained in a pig (n=14) AMI model (LAD occlusion) at 3.0T :a) Cine SSFP: TR/TE/\u03b1 = 3/1.5 ms/45\u00b0; acquired temporal resolution: 12 ms.b) Delayed enhancement MRI (DE-MRI): 10 min after 0.2 mmol/kg of contrast, scar was visualized using a gated IR-TFE sequence with inversion delay (TI) adjusted to null normal myocardium.c) Dual-IR T2W imaging (BB): Effective TE/TR: 80 ms/2*RR interval; TSE readout.Data Analysis: Myocardial region was manually segmented from cine, DE-MRI, and BB images Figure  using MA1) Normalized wall thickness (nWT) per segment = (WTES - WTED)/( WTED), where ED = End-Diastole, ES-End-Systole.2) Segmental scar burden, defined as ratio of pixels designated as scar to total number of pixels in each segment of DE-MRI. Scar pixel is one with SI > [mean + 5*Standard Deviation (SD) of normal remote myocardium].3) Segmental AAR, defined as regions in BB images with SI > mean + n*SD of normal remote myocardium .1) As a share of total myocardium, AAR burden was significantly higher (43 to 57 %) than scar burden (30 %) (Table 2) Spatially, AAR overlapped, and extended beyond scar regions.3) Reduction in segmental nWTwas lower in AAR regions that did not overlap scar region compared to those segments that did(108 \u00b1 36 % vs 91 \u00b1 29 %, Figure In sub-acute AMI, AAR is significantly larger than scar. In non-overlapping regions of AAR and scar, nWT, while diminished compared to normal remote myocardium, is significantly better than in regions of scar.NA"}
+{"text": "The word \"Hydroxypropyl\" is misspelled in the article title. The correct title is: Enhancement of Naringenin Bioavailability by Complexation with Hydroxypropyl-\u03b2-Cyclodextrin.The correct citation is: Shulman M, Cohen M, Soto-Gutierrez A, Yagi H, Wang H, et al. (2011) Enhancement of Naringenin Bioavailability by Complexation with Hydroxypropyl-\u03b2-Cyclodextrin. PLoS ONE 6(4): e18033. doi:10.1371/journal.pone.0018033Hydroxypropyl is also misspelled in the fifth sentence of the abstract. The fifth sentence should read: Hydroxypropyl-\u03b2-cyclodextrin (HP\u03b2CD), specifically, increased the solubility of naringenin by over 400-fold, and its transport across a Caco-2 model of the gut epithelium by 11-fold."}
+{"text": "Topiramate is the only approved oral preventive drug for chronic migraine (CM). However, we usually use other preventive drugs of prevention of frequent episodic migraine. Zonisamide is a neuromodulator that has showed efficacy in prevention of CM.To evaluate the efficacy of zonisamide after three months of therapy in CM patients (IHS-2006 criteria) that present inefficacy/intolerance/contraindication to topiramate, sodium valproate, \u00e2-blocks and flunarizine.Doses ZNS: 50-200mg. Primary end-point: Number of migraine days-NMD: Ineffective response-Inef(reduction of days<50%), good-G(50-75%), excellent-Exc(>75%). Secondary end-points: number of migraine episodes-NME; number of headache days-NHD; drug overuse; consumption of triptans and NSAIDs; EVA and MIDAS score, effective mean dose of ZNS, adverse events.We prospectively included 27 patients (May 2011-May 2012), 85.2% women. Twelve patients (44.4%) presented 16 adverse events (100% mild). Four of them-14.8% left the therapy. NMD, in the 23 patients who finished the three months follow-up, was Exc-39.1%, G-43.5%, and Inef-17.4%; NME was Exc-30.4%, G-43.5%, and Inef-26.1%; and NHD was Exc-34.8%, G-39.1%, and Inef-26.1%. Drug overuse disappeared in 47.8% of overusers. The consumption of triptans and NSAIDs decreased 54.3%, and 65.7%. The EVA score decreased 3.7 points (56.2%), and MIDAS scale score decreased 14.6 points (48.7%). Effective mean doses of ZNS: 106.6mg/night.Zonisamide presents a good profile of efficacy (70.3% of responses) and an acceptable tolerance. It could be a useful therapeutic option in CM patients intolerants, refractory, or with contraindications for topiramate, and the other drugs preventive drugs of migraine."}
+{"text": "The proposed contrast-enhanced first-pass perfusion cardiovascular MR (CMR) protocol integrates some recent technical MRI advances towards quantitative analysis of perfusion CMR images: fast multi-slice pulse sequence , robust To evaluate an integrated first-pass perfusion cardiovascular MR (CMR) protocol designed to determine absolute contrast-agent concentrations in blood and tissues.1 shortening expected in blood and wall. First-pass perfusion CMR was performed in 7 volunteers . A signal-to-concentration model was applied to calculate Gd-DTPA concentrations in blood and tissues [1 relationship based on Bloch equation in the center of k-space. Gd-DTPA concentrations were calculated assuming: fast water exchange condition [1=3.8L.mmol-1.s-1[1 measured with a multi-point SR fit. TurboFLASH imaging parameters included: FOV=350mm\u00d7315mm, slice thickness=8mm, matrix=160\u00d7144, in-plane resolution=2.2mm\u00d72.2mm, TE/TR=1.2/2.4ms, flip angle 10\u00b0, temporal resolution=114ms, tSENSEx3, centric k-space trajectory, and receiver bandwidth=1008Hz/pix. Total image acquisition time was 523ms for the acquisition of 4 slices, namely aortic root and SA base, mid, and apex levels, with respective TD values 50-164-278-393ms. Contours for the blood and left ventricle were drawn manually, and the myocardium was divided into 6 (base-mid) or 4 (apex) standard segments.A multi-slice saturation recovery (SR) pulse sequence with sequential SR time delays (TD) after a non-selective saturation pulse  was impl tissues ,5. A proondition , longituRepresentative images at peak contrast in blood and myocardium are shown Fig.The proposed integrated first-pass perfusion CMR protocol at 3T produced AIF and myocardial wall Gd-DTPA concentrations consistent with previously published results. Future work includes evaluation of the integrated protocol in cardiac patients."}
+{"text": "Biventricular size and function have been studied extensively in patients after tetralogy of Fallot (TOF) repair, but little is known about atrial size and function. The atria play a crucial role in ventricular filling during diastole, and abnormalities in atrial size and function may reflect ventricular diastolic dysfunction. Diastolic dysfunction may precede systolic dysfunction and may therefore play an important role in the early detection of ventricular dysfunction.We assessed bi-atrial size and function in patients after TOF repair, and evaluated relationships with biventricular systolic and diastolic function, and clinical parameters.51 patients (21\u00b18 years) and 30 healthy controls (31\u00b17 years) were included and underwent magnetic resonance imaging. Patients also underwent exercise testing, and biomarker assessment. Bi-atrial and biventricular size, systolic and diastolic function were assessed from time-volume curves (figure 2 (patients) vs. 28\u00b18 ml/m2 (controls), p=0.001); RA reservoir function, RA early emptying fraction, and the RA early-to-late-emptying ratio (E/A ratio) were decreased (RA E/A ratio: 1.1\u00b10.5 (patients) vs. 1.9\u00b10.9 (controls), p<0.001). Patients had larger right ventricular (RV) volumes, lower biventricular ejection fraction and prolonged biventricular deceleration time (Dt) (RV Dt: 0.24\u00b10.10 (patients) vs. 0.13\u00b10.04 (controls), p<0.001). Left atrial (LA) maximal volume and LA early emptying fraction were decreased. Left ventricular (LV) E/A-filling ratio was increased (LV E/A-ratio: 4.2\u00b13.7 (patients) vs. 2.7\u00b11.3 (controls), p=0.008).In patients, right atrial (RA) minimal volume (min.vol.), RA pump function, and RA late emptying fraction were increased (RA min.vol.: 34\u00b18 ml/m2 slope: 32\u00b14 (patients with EDFF) vs. 28\u00b16 (patients without EDFF), p=0.014). Patients with abnormal RA emptying (reservoir function <30% and pump function >24%) had higher NT-proBNP levels  vs. 7\u00b17 pmol/l , p=0.002) and worse exercise capacity. RA min.vol. was positively associated with RV end-diastolic volume .Patients with end-diastolic forward flow (EDFF) had larger RA and RV size, higher RA pump function (RA pump function: 27\u00b18% (patients with EDFF) vs. 20\u00b17% (patients without EDFF), p=0.003), higher N-terminal prohormone brain natriuretic peptide (NT-proBNP) levels, and higher ventilatory response to carbon dioxide production (VE/VCOIn TOF patients with good clinical condition and moderate RV dilatation, abnormal bi-atrial function and abnormal biventricular diastolic function is common. Abnormal RA emptying was associated with impaired clinical outcome, as was the presence of EDFF. These parameters, together with RA enlargement, could serve as useful markers for clinically relevant RV diastolic dysfunction.This research was funded by the Netherlands Heart Foundation: grant 2006B095."}
+{"text": "ICU patients (ptes) account for a large proportion of HI. Its epidemiology allow us to identify key issues and prioritize interventions. We determined the incidence of HI in ICU in Uruguay and described specific locations, dispositive-associated rates and microorganisms.A national surveillance system for HI was implemented in 2006; prospective surveillance was performed using NNISS criteria. It is mandatory, to record online and send their data to the Ministry of Health. Data are audited and results published annually.S. aureus , A. baumanii , Ps. aeruginosa  and K. pneumoniae .We present 2010 results of medical-surgical ICUs. 53 hospitals reported. 13611 ptes were surveyed (99541 ptes-days). 2340 HI episodes were reported (density of incidence rate (ID) 23.5/1000 ptes-days and cumulative incidence 17.2%). Ventilator-associated pneumonia (VAP) (34%), bronchitis (B) (28%), catheter-associated urinary tract infection (CAUTI) (21%) and central-line associated bacteremia (CLAB) (8%) were the main infectious sites. VAP ID was 14.2/1000 ventilator-days (dispositive utilization (DU) 0.5, secondary bacteremia (SB) 5.3% and contributor mortality (CM) 19.7%), CLAB ID was 1.9/1000 catheter-days  17.7%) and CAUTI ID was 5.8/1000 catheter-days  3.7%, (CM) 5.1%). Microorganisms were Respiratory tract infections are the main problem. ICUs predominance of Gram Negative Bacilli in these infections may suggest the importance of exogenous sources. VAP and CAUTI rates are also high, diminish its incidence is priority for the national program.None declared."}
+{"text": "Thermus oshimai JL-2 and Thermus thermophilus JL-18 each have a circular chromosome, 2.07 Mb and 1.9 Mb in size, respectively, and each has two plasmids ranging from 0.27 Mb to 57.2 kb. The megaplasmid of each strain contains a gene cluster for the reduction of nitrate to nitrous oxide, consistent with their incomplete denitrification phenotypes.The strains  Thermus comprises >15 species of thermophilic bacteria, including the biotechnologically exalted Thermus aquaticus and the genetically tractable Thermus thermophilus. We reported previously the isolation of a large number of Thermus strains from the Great Boiling Spring system in the U.S. Great Basin, typified by Thermus oshimai JL-2 and Thermus thermophilus JL-18, and described their roles in incomplete denitrification in situ (The genus T. oshimai JL-2 and T. thermophilus JL-18 were sequenced using 454-GS-FLX Titanium and Illumina GA II (2 \u00d7 75 bp) methodologies, were assembled using Newbler version 2.3 (prerelease), and were annotated using Prodigal version 1.4, Gene Prediction Improvement Pipeline (GenePRIMP) at the Joint Genome Institute (JGI) in Walnut Creek, CA. The data are available at the Joint Genome Institute (JGI) Integrated Microbial Genomes (IMG) website ; and a smaller circular plasmid, pTHEOS02 . T. thermophilus JL-18 has a 1.9-Mb circular chromosome carrying 2,057 predicted genes; a circular megaplasmid, pTTJL1801 ; and a smaller circular plasmid, pTTJL1802 .The genomes of T. thermophilus JL-18 was similar to those of T. thermophilus HB8 and T. thermophilus HB27, apart from a few chromosomal rearrangements . T. oshimai JL-2 possesses two nitrate/nitrite transporters (narK1 and narK2), which is similar to T. thermophilus HB8 (T. thermophilus JL-18 possesses a single copy of narK. Genes encoding nitrite reductase (nirS and nirK in T. oshimai JL-2 and nirS in T. thermophilus JL-18) and nitric oxide reductase (norB and norC) were also identified in close proximity to the narGHIJK operon in the megaplasmids of both T. thermophilus JL-18 and T. oshimai JL-2. However, nitrous oxide reductase (nos) genes, which are needed for the conversion of nitrous oxide to dinitrogen, are absent, concurrent with the incomplete denitrification phenotype of these strains and the high flux of nitrous oxide reported at Great Boiling Spring (T. thermophilus HB8 and HB27 (sox gene cluster that includes a sulfite dehydrogenase gene (soxCD) essential for the chemotrophic growth of Paracoccus pantotrophus , CP003250.1 (T. oshimai JL-2 megaplasmid pTHEOS01), CP003251.1 (T. oshimai JL-2 plasmid pTHEOS02), CP003252.1 (T. thermophilus JL-18 chromosome), CP003253.1 (T. thermophilus JL-18 plasmid pTTJL1801), and CP003254.1 (T. thermophilus JL-18 plasmid pTTJL1802).The genome sequences from this study are available from GenBank under the following accession no:"}
+{"text": "However, its efficacy and indication are still controversial issues. Recently, randomized controlled studies are ongoing in other countries. Our hypothesis is that PMX-DHP therapy may improve the hemodynamic status in septic patients.Since 1994, Polymyxin B-direct hemoperfusion (PMX-DHP)  in our ICU were included in this retrospective observational study. Patients' clinical, microbiological and PAC data were collected from medical archives. PAC variables were compared between immediately before and immediately after PMX-DHP therapy. Values were expressed as mean \u00b1 SD. Data were analyzed by Wilcoxon signed-ranks test, chi-square test and Fisher's exact probability test. 2), and oxygen delivery and consumption  significantly improved after PMX-DHP therapy . The systemic venous resistance index (SVRI) (dyn second m2/cm5) and mixed venous oxygen saturation  were not statistically different before and after PMX-DHP therapy . The inotropic score and P/F ratio improved after the therapy .Sixty-three patients ) were studied. The mortality rate was 30.2% at 28 days after PMX-DHP. The APACHE II score and SOFA score on the day of PMX-DHP therapy were 20.2 \u00b1 14.8 and 7.3 \u00b1 3.8, respectively. Mean arterial pressure , cardiac index (CI; l/minute/m2, VO2, inotropic score and P/F ratio improved after PMX-DHP therapy, while SVRI and SVO2 did not change. PMX-DHP could contribute to oxygen delivery due to improve hemodynamic status, while decreasing inotropic agents in septic patients.MAP, CI, DO"}
+{"text": "The effects of adrenomedullin in circulatory shock states are controversially discussed: while its exogenous supplementation improved organ function and survival  in expern = 11) or the adrenomedullin antibody HAM1101 . Fifteen hours later animals were anesthetized, mechanically ventilated and instrumented for a consecutive 6-hour observation period. Colloid fluid resuscitation and continuous i.v. noradrenaline were titrated to maintain normotensive (mean blood pressure >60 mmHg) and hyperdynamic hemodynamics. Creatinine blood levels and clearance were assessed as surrogate for glomerular filtration [Immediately after cecal ligation and puncture to induce peritonitis, mice randomly received vehicle (ltration . All datP < 0.001), increased total diuresis  vs. 0.6  ml, P = 0.028) resulting in improved fluid balance  vs. 0.26 , P = 0.011) and kidney function  vs. 2.0  \u03bcg/ml, P = 0.006; creatinine clearance: 400  vs. 197  \u03bcl/minute, P = 0.006).Adrenomedullin antagonism decreased the noradrenaline requirements needed to achieve target hemodynamics  vs. 0.02  \u03bcg/g/hour, In resuscitated murine septic shock, early modulation of excess adrenomedullin activity via antibody HAM1101 improves cardiovascular catecholamine responsiveness, ultimately associated with attenuation of acute kidney injury."}
+{"text": "To the Editor, 9/L; ANC: 4.6 x 109/L; Hct: 6.9%; RBC count: 1.19 x 1012/L; RDW: 20.5; MCV: 58 fL; Plt count: 485 x 109/L; absolute reticulocyte count: 45.5 x 109/L . Peripheral blood smear showed hypochromic microcytic red cells, polychromasia, and nucleated red cells. Serum iron was 5 \u03bcg/dL, total iron binding capacity was 450 \u03bcg/dL, and ferritin was <1 ng/mL. A 3-year-old female toddler was referred to our pediatric emergency unit with a 2-week history of fatigue, anorexia, progressive pallor, and vomiting. Medical history showed that iron deficiency anemia was diagnosed one year before and oral iron-sulfate was given. She also had a one year history of intermittent vomiting. Her diet seemed adequate in iron-rich foods. Chest X-ray and abdominal ultrasonographic examination performed in a medical center were normal. Complete blood count findings were as follows: Hb: 1.7 g/dL; WBC count: 8.0 x 10She was hospitalized and given packed red cell transfusion. The following day she suddenly developed respiratory distress after breakfast. Chest X-ray showed a radiolucent lesion in the right paracardiac area . Congeni12/L; MCV: 70.5 fL; MCH: 21.9; MCHC: 31; RDW: 24; WBC count: 9.8 x 109/L; ANC: 4.3 x 109/L, Plt count: 453 x 109/L; absolute reticulocyte count: 11 x 109/L) [normal 50-150 x 109/L]). Peripheral blood smear showed hypochromic microcytic red cells. Esophagogastroduodenoscopy and esophageal biopsy showed erosions, gastroesophageal reflux, chronic inflammation, and hyperplasia of the epithelium. A proton pump inhibitor, domperidone, and anti-acid medications were started. Iron sucrose was administered twice weekly for 3 weeks and the Hb increased . Blood smear showed normal red cells. At the 2-year follow-up the patient had no gastrointestinal symptoms or anemia. At 6 weeks post surgery the patient had no gastrointestinal symptoms, but her Hb dropped to 7.9 g/dL and fecal occult blood test findings were positive (Hct: 25.6%; RBC count: 3.63 x 10The coexistence of Morgagni and Bochdalek defects is rarely reported . About 5CDH must be included in the differential diagnosis of severe iron deficiency anemia in the absence of such obvious causes as nutritional deficiency, melena, hematochezia, and malabsorption. Physicians must be aware that a history of intermittent vomiting and/or sudden onset respiratory distress in young children are indications for imaging of the upper gastrointestinal tract and thorax."}
+{"text": "Head and Neck Oncology and is not related to the content of BMC Medicine in any way.This comment relates to articles published in archived content of the journal  Head and Neck Oncology and is not related to the content of BMC Medicine in any way.This comment relates to articles published in archived content of the journal Head & Neck Oncology. In order to maintain the integrity of the BioMed Central portfolio of journals, we decided to cease publication of the journal with effect from 9th August 2012.While conducting an internal audit of publications between January and June 2012, BioMed Central discovered a number of apparent major irregularities in the content and editorial handling of the journal University College London (UCL) and University College London Hospital (UCLH) subsequently conducted a joint investigation of our concerns. This focused primarily on the actions of Editor-in-Chief Mr Colin Hopper, because neither of the other active Editors-in-Chief, Waseem Jerjes and Tahwinder Upile, were employees of UCL during the time covered by our audit, so an investigation of their actions would have been beyond the scope of UCL\u2019s investigation.Following their investigation UCL were satisfied that there was no evidence of research misconduct arising from any employee, honorary researcher or student in relation to the articles they were asked to investigate. They were also satisfied that there was no evidence of editorial or author misconduct on the part of Colin Hopper.In the absence of definitive conclusions about all of the concerns raised by its audit, BioMed Central has provided details of its findings in the form of a Publishers Note on relevant articles which will be updated if further information becomes available. If you are an author of a published article in this journal and have further questions, please contact info@biomedcentral.com.The affected articles are listed below:A review of the epidemiology of oral and pharyngeal carcinoma: updateDaniel M SamanHead & Neck Oncology 2012, 4:1http://www.headandneckoncology.org/content/4/1/1Randomized clinical trial of LigaSure versus conventional suture ligation in thyroid surgeryAnandi H.W. Schiphorst, Bas A. Twigt, Sjoerd G. Elias and Thijs van DalenHead & Neck Oncology 2012, 4:2http://www.headandneckoncology.org/content/4/1/2A decision support system for quality of life in head and neck oncology patientsJoaquim J Goncalves and Alvaro RochaHead & Neck Oncology 2012, 4:3http://www.headandneckoncology.org/content/4/1/3Etiology analysis and computed tomography imaging of a tonsillar inflammatory myofibroblastic tumor: report of an immunocompetent patient and brief reviewYun-Zhen Luo, Li-Bo Dai, Shui-Hong Zhou, Xing-Mei Luo, Jun Fan and Ling-Xiang RuanHead & Neck Oncology 2012, 4:4http://www.headandneckoncology.org/content/4/1/4cTNM vs. pTNM: the effect of not applying ultrasonography in the identification of cervical nodal diseaseWaseem Jerjes, Tahwinder Upile, Hani Radhi, Aviva Petrie, Jesuloba Abiola, Aidan Adams, Jacqueline Callear, Panagiotis Kafas, Syedda Abbas, Kartic Rajaram, Colin HopperHead & Neck Oncology 2012, 4:5http://www.headandneckoncology.org/content/4/1/5The effect of tobacco and alcohol and their reduction/cessation on mortality in oral cancer patients: short communicationWaseem Jerjes, Tahwinder Upile, Hani Radhi, Aviva Petrie, Jesuloba Abiola, Aidan Adams, Panagiotis Kafas, Jacqueline Callear, Ramin Carbiner, Kartic Rajaram, Colin HopperHead & Neck Oncology 2012, 4:6http://www.headandneckoncology.org/content/4/1/6Nimesulide inhibited the growth of hypopharyngeal carcinoma cells via suppressing Survivin expressionJia-Jun Tian, Su-Mei Lu, Liang Yu, Ju-Ke Ma, Ya-Kui Mu, Hai-Bo Wang and Wei XuHead & Neck Oncology 2012, 4:7http://www.headandneckoncology.org/content/4/1/7Primary squamous cell carcinoma of thyroid: a case report and review of literatureMutahar A Tunio, Mushabbab Al Asiri, Mosa Fagih and Rashad AkashaHead & Neck Oncology 2012, 4:8http://www.headandneckoncology.org/content/4/1/8The prognostic significance of p63 and Ki-67 expression in myoepithelial carcinomaYou-Hu Jiang, Bo Cheng, Ming-Hua Ge and Gu ZhangHead & Neck Oncology 2012, 4:9http://www.headandneckoncology.org/content/4/1/9Parapharyngeal space hemangiopericytoma treated with surgery and postoperative radiation- a case reportMuhammad Mohsin Fareed, Abdullah Suleiman Mazaed Al Amro, Rashad Akasha, Mansour A Assiry, Mushabbab Al Asiri, Mutahar Tonio and Yasir BayoumiHead & Neck Oncology 2012, 4:10http://www.headandneckoncology.org/content/4/1/10Knockdown of aberrantly expressed nuclear localized decorin attenuates tumour angiogenesis related mediators in oral cancer progression model in vitroNyla Dil and Abhijit G BanerjeeHead & Neck Oncology 2012, 4:11http://www.headandneckoncology.org/content/4/1/11A retrospective, deformable registration analysis of the impact of PET-CT planning on patterns of failure in stereotactic body radiation therapy for recurrent head and neck cancerKyle Wang, Dwight E Heron, John C Flickinger, Jean-Claude M Rwigema, Robert L Ferris, Gregory J Kubicek, James P Ohr, Annette E Quinn, Cihat Ozhasoglu and Barton F BranstetterHead & Neck Oncology 2012, 4:12http://www.headandneckoncology.org/content/4/1/12Clear cell chondrosarcoma of the head and neckSepideh Mokhtari and Abbas MirafshariehHead & Neck Oncology 2012, 4:13http://www.headandneckoncology.org/content/4/1/13Delay in pathological tissue processing time vs. mortality in oral cancer: Short communicationWaseem Jerjes, Tahwinder Upile, Hani Radhi, Aviva Petrie, Aidan Adams, Jacqueline Callear, Panagiotis Kafas, Colin HopperHead & Neck Oncology 4:14http://www.headandneckoncology.org/content/4/1/14The cost burden of oral, oral pharyngeal, and salivary gland cancers in three groups: commercial insurance, medicare, and medicaidJed J Jacobson, Joel B Epstein, Frederick C Eichmiller, Teresa B Gibson, Ginger S Carls, Emily Vogtmann, Shaohung Wang and Barbara MurphyHead & Neck Oncology 2012, 4:15http://www.headandneckoncology.org/content/4/1/15Photodynamic therapy in the management of potentially malignant and malignant oral disordersWaseem Jerjes, Zaid Hamdoon, Colin HopperHead & Neck Oncology 4:16http://www.headandneckoncology.org/content/4/1/16CO2 lasers in the management of potentially malignant and malignant oral disordersWaseem Jerjes, Zaid Hamdoon, Colin HopperHead & Neck Oncology 2012, 4:17http://www.headandneckoncology.org/content/4/1/17Expression of Glut-1, HIF-1alpha, PI3K and p-Akt in a case of ceruminous adenomaWan-Qin Shen, Ke-Jia Cheng, Yang-Yang Bao, Shui-Hong Zhou and Hong-Tian YaoHead & Neck Oncology 2012, 4:18http://www.headandneckoncology.org/content/4/1/18Systemic therapy in the management of metastatic or advanced salivary gland cancersAymen Lagha, Nesrine Chraiet, Mouna Ayadi, Sarra Krimi, Bassem Allani, Hela Rifi, Henda Raies and Amel MezliniHead & Neck Oncology 2012, 4:19http://www.headandneckoncology.org/content/4/1/19Tongue cancer in young patients: Case report of a 26-year-old patient.Aleksandra Crede, Michael Locher and Marius BredellHead & Neck Oncology 2012, 4:20http://www.headandneckoncology.org/content/4/1/20Reconstruction of scalp defects with the radial forearm free flapLarissa Sweeny, Brendan T Eby, J. Scott Magnuson, William R Carroll and Eben L RosenthalHead & Neck Oncology 2012, 4:21http://www.headandneckoncology.org/content/4/1/21The use of specific anti-growth factor antibodies to abrogate the oncological consequences of transfusion in head & neck squamous cell carcinoma: an in vitro studyTahwinder Upile, Waseem Jerjes, Sandeep Singh, Mohammed Al-Khawalde, Zaid Hamdoon, Hani Radhi, Colin HopperHead & Neck Oncology 2012, 4:22http://www.headandneckoncology.org/content/4/1/22Definitive radiotherapy for early stage glottic cancer by 6 MV photonsChi-Chung TONG, Kwok-Hung AU, Roger Kai-Cheong NGAN, Sin-Ming CHOW, Foon-Yiu CHEUNG, Yiu-Tung FU, Joseph Siu-Kei AU and Stephen Chun-Key LAWHead & Neck Oncology 2012, 4:23http://www.headandneckoncology.org/content/4/1/23Branchial cysts within the parotid salivary glandTahwinder Upile, Waseem Jerjes, Mohammed Al-Khawalde, Panagiotis Kafas, Steve Frampton, Angela Gray, Bruce Addis, Ann Sandison, Nimesh Patel, Holger Sudhoff, Hani RadhiHead & Neck Oncology 2012, 4:24http://www.headandneckoncology.org/content/4/1/24A patient with ulcerated calcifying epithelioma of Malherbe in the pinna: case reportTahwinder Upile, Waseem Jerjes, Fabian Sipaul, Ann Sandison, Panagiotis Kafas, Mohammed Al-Khawalde, Hani RadhiHead & Neck Oncology 2012, 4:25http://www.headandneckoncology.org/content/4/1/25Computed tomography and pathological findings of five nasal neurilemmomasJing Hu, Yang-Yang Bao, Ke-Jia Cheng, Shui-Hong Zhou, Ling-Xiang Ruan and Zhou-Jun ZhengHead & Neck Oncology 2012, 4:26http://www.headandneckoncology.org/content/4/1/26Metastatic rhabdomyosarcoma of the thyroid gland, a case reportMohamed T Hafez, Mohamed A Hegazy, Khaled Abd Elwahab, Mohammad Arafa, Islam Abdou and Basel RefkyHead & Neck Oncology 2012, 4:27http://www.headandneckoncology.org/content/4/1/27Squamous cell carcinoma of the oral cavity and the oropharynx in patients less than 40 years of age: a 20-year analysisSamuel E Udeabor, Majeed Rana, Gerd Wegener, Nils-Claudius Gellrich and Andr\u00e9 M EckardtHead & Neck Oncology 2012, 4:28http://www.headandneckoncology.org/content/4/1/28Structural validation of oral mucosal tissue using optical coherence tomographyZaid Hamdoon, Waseem Jerjes, Raed Al-Delayme, Gordon McKenzie, Amrita Jay, Colin HopperHead & Neck Oncology 2012 4:29http://www.headandneckoncology.org/content/4/1/29Solitary giant neurofibroma of the neck subjected to photodynamic therapy: case studyZaid Hamdoon, Waseem Jerjes, Raed Al-Delayme, Colin HopperHead & Neck Oncology 2012 4:30http://www.headandneckoncology.org/content/4/1/30Oral sex, cancer and death: sexually transmitted cancersTahwinder Upile, Waseem Jerjes, Mohammed Al-Khawalde, Hani Radhi and Holger SudhoffHead & Neck Oncology 2012 4:31http://www.headandneckoncology.org/content/4/1/31Enhanced patient reported outcome measurement suitable for head and neck cancer follow-up clinicsNaseem Ghazali, Derek Lowe and Simon N RogersHead & Neck Oncology 2012, 4:32http://www.headandneckoncology.org/content/4/1/32A patient with primary Burkitt's lymphoma of the postnasal space: case reportTahwinder Upile, Waseem Jerjes, Jesuloba Abiola, Panagiotis Kafas, Ann Sandison, Zaid Hamdoon, Mohammed Al-Khawalde and Hani RadhiHead & Neck Oncology 2012, 4:33http://www.headandneckoncology.org/content/4/1/33Positron emission tomography in the detection of occult primary head and neck carcinoma: a retrospective studyGabriel G Pereira, Joaquim C Silva and Eurico F MonteiroHead & Neck Oncology 2012, 4:34http://www.headandneckoncology.org/content/4/1/34Granulocyte colony-stimulating factor-producing squamous cell carcinoma of the lower gingiva: a case reportJun-ichi Kobayashi Dr., Akihiro Miyazaki Dr., Takashi Yamamoto Dr., Kenji Nakamori Dr., Rina Suzuki Dr., Takeshi Kaneko Dr., Naohiro Suzuki Dr. and Hiroyoshi Hiratsuka Prof.Head & Neck Oncology 2012, 4:35http://www.headandneckoncology.org/content/4/1/35Spinal metastasis in head and neck cancerGregory M Trilling, Hyongyu Cho, Mohamed A Ugas, Samerah Saeed, Asia Katunda, Waseem Jerjes and Peter V GiannoudisHead & Neck Oncology 2012, 4:36http://www.headandneckoncology.org/content/4/1/36Analysis of the compatibility of dental implant systems in fibula free flap reconstructionRamin Carbiner, Waseem Jerjes, Kaveh Shakib, Peter V Giannoudis and Colin HopperHead & Neck Oncology 2012, 4:37http://www.headandneckoncology.org/content/4/1/37p16 overexpression in malignant and premalignant lesions of the oral and esophageal mucosa following allogeneic hematopoietic stem cell transplantationYasumasa Kakei, Masaya Akashi, Hideki Komatsubara, Tsutomu Minamikawa and Takahide KomoriHead & Neck Oncology 2012, 4:38http://www.headandneckoncology.org/content/4/1/38Spinal metastasis in thyroid cancerSami Ramadan, Mohamed A Ugas, Richard J Berwick, Manisha Notay, Hyongyu Cho, Waseem Jerjes, Peter V GiannoudisHead & Neck Oncology 2012, 4:39http://www.headandneckoncology.org/content/4/1/39Feasibility of recruitment to an oral dysplasia trial in the United KingdomPaul C Nankivell, Janet A Dunn, Michael J.S. Langman and Hisham MehannaHead & Neck Oncology 2012, 4:40http://www.headandneckoncology.org/content/4/1/40Effect of human beta-defensin-3 on head and neck cancer cell migration using micro-fabricated cell islandsKevin Wang, Joanne H Wang, Harihara Baskaran, Russell Wang and Rick JurevicHead & Neck Oncology 2012, 4:41http://www.headandneckoncology.org/content/4/1/41"}
+{"text": "Cardiac MRI (CMR) is used to follow patients after TOF repair to assess pulmonary regurgitation (PR), pulmonary stenosis (PS) and right ventricular (RV) function. 4D flow-sensitive MRI techniques enable visualization of complex flow patterns ,2]. Wit. Wit2]. Analyze flow patterns in superior vena cava (SVC), inferior vena cava (IVC), right atrium (RA), right ventricle (RV), and main, right and left pulmonary arteries  using 3D radially-undersampled, 4D flow-sensitive MRI.3 volume, 1.0-1.25mm3 acquired isotropic spatial resolution, and VENC of 50-200cm/s. Scan time was approximately 8-12min using an adaptive respiratory bellows reading and 50% efficiency. Retrospective ECG-triggered cardiac gating was used with datasets reconstructed into 20 time frames. Flow visualization and analysis  on 1.5T and 3.0T clinical systems  after IRB-approval and obtaining consent. 4D flow-sensitive MRI data were acquired with a radially undersampled phase contrast (PC) sequence, PC VIPR . The. The4]."}
+{"text": "With newly available 32-channel cardiac coil, 3 tesla MRI system provides increased signal-to-noise radio (SNR), reduced imaging time and improved spatial resolution. This study sought to determine the diagnostic performance of 3T stress MRMPI with 32-channel cardiac coil in detecting clinical relevant coronary artery stenosis in comparison with invasive coronary angiography (ICA).Forty four patients  who were scheduled for ICA underwent stress MRMPI with 3T MRI  with a 32-channel cardiac receiver coil . The total amount of contrast medium  was 0.15mmole/kg with injection rate at 4ml/s. The MR protocol included gadolinium-enhanced stress first-pass perfusion , rest perfusion, and delayed enhancement (DE). Ischemia was defined as (1) a territory with perfusion defect at stress study but no DE, (2) territory with DE but additional peri-infarcted perfusion defect at stress study. Diagnostic performance was determined on a per-patient basis. Quantitative CA served as the reference standard.Coronary angiography depicted significant stenosis  in 22 of 44 patients (50%). No complications were observed during Stress perfusion 3.0 tesla MRI and dipyridamole infusion. For detection of significant coronary stenosis, 3T stress MRMPI provided sensitivity: 0.91(0.75-1.00), specificity: 0.86(0.72-1.00), positive predictive value: 0.87(0.74-1.00) and negative predictive value: 0.91(0.748-1.00). The overall diagnostic accuracy was: 0.88.Our study showed that 3T stress MRMPI using 32-channel cardiac coil demonstrated high diagnostic accuracy for detection of significant stenosis, having QCA as the reference standard. Further investigation is needed to assess if advances in technology and improvement in diagnostic accuracy for CAD detection will translate in better patient care and outcomes."}
+{"text": "Follow-up studies used flyback EPI to reduce image artifacts. More recently a spiral k-space trajectory was utilized for improved SNR.The original implementation of 2D cine DENSE (displacement encoding with stimulated echoes) employed a conventional EPI To evaluate and compare SNR efficiencies and strain results of these three techniques for 2D cine DENSE imaging.k-space trajectories, namely conventional bottom-up interleaved EPI, flyback bottom-up interleaved EPI, and interleaved spiral. They were compared in volunteers using a two-breath-hold protocol on a 1.5T Siemens Avanto system with a four-channel chest coil. In accordance with protocols approved by the local institutional review board, 5 healthy volunteers were imaged. Identical parameters included pixel size = 3.8 \u00d7 3.8 mm2, slice thickness = 8 mm, flip angle = 20\u00b0, cardiac phases = 15, displacement encoding frequency = 0.08 cycles/mm, two-point phase cycling, and through-plane dephasing frequency = 0.08 cycles/mm to suppress artifacts. Other parameters for conventional/flyback EPI included image matrix = 72 \u00d7 96, TE = 8.57/10.57 ms, TR = 17.69/21.19 ms, ETL = 8, segments = 16, sampling time per image = 99.8 ms, heartbeats per breath-hold = 21/20, and fat suppression by water excitation. For spiral, other parameters included image matrix = 96 \u00d7 96, TE = 1.08 ms, TR = 17 ms, interleaves = 6, interleaves per heartbeat = 2, sampling time per image = 66.8 ms, heartbeats per breath-hold = 14, and fat suppression by chemically-selective saturation pulses applied prior to displacement-encoding pulses. Based on the magnitude-reconstructed images, SNR efficiencies were calculated. Strain maps were also calculated.Cine DENSE pulse sequences were developed that employed different As shown in Figure Conventional EPI, flyback EPI and spiral cine DENSE produce similar strain results. Spiral cine DENSE provides improved SNR efficiency compared to the other two techniques."}
+{"text": "Liver fibrosis (LF) progression is fated to become one of the major long-term complications in HIV patients, even in those without HCV or HBV co-infections (HIV-mono-infected). The aim of this study was to assess LF progression in HIV-mono-infected patients and associated risk factors.Observational retrospective study. All HIV naive patients who started HAART from 1996 to 2006 were included. Concordance between FIB-4 and APRI scores was assessed using the weighted kappa coefficient. Rates of transition from lower classes to higher classes were estimated by Kaplan-Meier analysis. Cox regression models were applied to assess possible predictors both at baseline and during the follow-up.1,112 naive patients were selected. A moderate concordance between FIB-4 and APRI was demonstrated (K=0.573). For FIB-4, the incidence of transition to higher classes was 0.064 PYFU , while for APRI the incidence of transition was 0.099 PYFU . Viro-immunological control during HIV infection appeared to reduce the risk of both FIB-4 and APRI transitions. HIV-RNA <500 copies/ml  and higher CD4 T-cell counts only for FIB-4 (HR 0.881 p=0.0004 for 100 cells higher) during the follow-up were statistically protective. Among baseline variables, for FIB-4 transition, age \u2265 40 years (HR 1.037 p<0.0001) and higher FIB-4 values (HR 1.526 p=0.0038) were associated with increased risk of LF progression, while sexual risk factor for HIV acquisition resulted to be protective (HR 0.524 p=0.0314). For APRI, male gender (HR 1.390 p=0.017), higher GGT values (HR 1.015 p=0.014) and higher APRI values (HR 1.748 p=0.007) were independently associated with APRI transition. A sensitivity analysis demonstrated that DDX drugs as time-dependent covariates were associated with a significant risk of transition with FIB-4 (HR 1.662 p=0.0007) or APRI (HR 1.661 p=0.0001)Our data suggest that a better viro-immunological control of HIV infection may slow down fibrosis progression provided that DDX are avoided. Moreover our analysis provided a comprehensive feature of the risk factors that should be controlled in clinical practice"}
+{"text": "There is an error in the title. \"gp120\" was incorrectly labeled as \"gsp120.\"The correct title is: Synergistic Cooperation between Methamphetamine and HIV-1 gp120 through the P13K/Akt Pathway Induces IL-6 but not IL-8 Expression in AstrocytesThe correct citation is: Shah A, Silverstein PS, Kumar S, Singh DP, Kumar A (2012) Synergistic Cooperation between Methamphetamine and HIV-1 gp120 through the P13K/Akt Pathway Induces IL-6 but not IL-8 Expression in Astrocytes. PLoS ONE 7(12): e52060. doi:10.1371/journal.pone.0052060"}
+{"text": "We have previously shown that the tryptophan-catabolizing enzyme indoleamine-2,3-dioxygenase (IDO) critically impairs host defense against secondary bacterial pneumonia . Since iin vitro data, transgenic mice that conditionally express IDO in the airway epithelium upon doxycycline (dox) treatment and control mice were challenged with LPS (1 \u03bcg) or Klebsiella pneumoniae (104 colony-forming units) intranasally and sacrificed after 24 hours to count neutrophils in bronchoalveolar lavage fluid . Statistical analysis was performed by Student's t test or Mann-Whitney U test where appropriate. P < 0.05 was considered significant.Freshly isolated neutrophils were cultured in the presence or absence of tryptophan, kynurenine and 3-hydroxy-anthranilic acid. Apoptosis was identified by annexin V/propidium iodine staining . To confirm our P < 0.05), which was reversed by adding tryptophan. Conditional transgenic mice, which showed marked expression of IDO in the pulmonary compartment, had reduced neutrophil numbers in bronchoalveolar lavage fluid after challenge with K. pneumoniae  and LPS , which was associated with active caspase-3 staining in dox-treated mice, but not in control mice.Both kynurenine and 3-hydroxy-anthranilic acid enhanced apoptosis in freshly isolated neutrophils (60.3 \u00b1 8.7% and 45.5 \u00b1 1.7% respectively vs. 33.5 \u00b1 8.1% under control conditions, both in vivo and may impair host defense against secondary bacterial infections.Neutrophils undergo apoptosis in presence of kynurenine or 3-hydroxy-anthranilic acid and the absence of tryptophan. Pulmonary IDO expression, as occurs during influenza infection, enhances neutrophil apoptosis"}
+{"text": "In rheumatoid arthritis (RA) synovial tissue (ST) angiogenesis can be observed already in the earliest phase of disease. The chemokine CXCL12, which is induced via the non-canonical nuclear factor-kappaB (NF-kB) pathway, plays an important role in angiogenesis, lymphocyte transendothelial migration, and the homing of endothelial progenitor cells. Therefore, the non-canonical pathway, with its key mediator NF-kB inducing kinase (NIK), may play an important role in pathological angiogenesis and the perpetuation of synovial inflammation in RA.To study the role of non-canonical NF-kB signaling in pathological angiogenesis in RA.-/- mice in the aortic ring assay. Physiological  angiogenesis was evaluated by isolectin B4 staining of the retina followed by confocal microscopy. The contribution of NIK to synovial angiogenesis was studied in vivo in antigen-induced arthritis (AIA).Expression of NIK and CXCL12 in RA ST was evaluated using immunofluorescence microscopy (IF). Angiogenesis was studied in endothelial cells (EC) in vitro using the tube formation assay and ex vivo by comparing WT and NIK-/- mice showed normal TNF- and VEGF-induced microvessel outgrowth. In contrast, whereas non-canonical NF-kB stimuli induced microvessel outgrowth in WT mice (unstim 29.94 \u00b1 6.08 vs. LT 159.1 \u00b1 50.24 vs. LIGHT 110.3 \u00b1 17.68 (mm^2) p<0.05), no microvessel outgrowth was observed in aortic rings from NIK-/- mice (unstim 28.74 \u00b1 15.89 vs. LT 45.9 \u00b1 16.71 vs. LIGHT 43.41 \u00b1 15.73 (mm^2)). In line with this, NIK-/- mice exhibited normal developmental angiogenesis in the retina, but a 50% reduction in pathological angiogenesis in synovial inflammation .NIK, p52 and CXCL12 were highly expressed in EC in RA ST, mainly in small (newly formed) blood vessels. Stimuli that induce non-canonical NF-kB signaling (lymphotoxin (LT), LIGHT, and CD40L) significantly enhanced in vitro tube formation 2,5-fold (p<0.05), which could be completely blocked by siRNA targeting NIK or IKK\u03b1. Aortic rings from WT and NIK-/- mice exhibited normal developmental and VEGF-induced angiogenesis, but reduced pathological angiogenesis in AIA. These findings point towards an important role of the non-canonical NF-kB pathway in pathological angiogenesis associated with chronic  inflammation. This could be exploited for the development of future new therapies for RA.NIK is preferentially expressed in EC in RA ST and non-canonical NF-kB signaling in EC results in enhanced angiogenesis in vitro. NIK"}
+{"text": "There were errors in the article title.The correct version of the title is: The Long Coiled-Coil Protein NECC2 Is Associated to Caveolae and Modulates NGF/TrkA Signaling in PC12 CellsThe correct version of the citation is: D\u00edaz-Ruiz A, Rabanal-Ruiz Y, Tr\u00e1vez A, Gracia-Navarro F, Cruz-Garc\u00eda D, et al. (2013) The Long Coiled-Coil Protein NECC2 Is Associated to Caveolae and Modulates NGF/TrkA Signaling in PC12 Cells. PLoS ONE 8(9): e73668. doi:10.1371/journal.pone.0073668"}
+{"text": "To assess the feasibility of cardiac magnetic resonance imaging (MRI) and angiography (MRA) at 3.0T in pediatric patients with congenital heart disease (CHD) and to compare the technical and diagnostic performance with an age-matched and clinically comparable control group imaged at 1.5T.Forty-six patients  with suspected or known CHD were evaluated (9/2008-5/2011). SSFP cine imaging, time-resolved MRA (TR-MRA), and high resolution contrast-enhanced MRA (HR-MRA) were performed. Two independent observers analyzed the MRI data for image quality, thoraco-abdominal vessel definition, and artifacts.At 3.0T, 91% of SSFP cine images (k=0.55) and 97% (374 of 387) of vascular segments (k=0.49) were rated as good or excellent image quality with 72% of SSFP cine images having mild and 27% having moderate artifacts. At 1.5T, 92% of SSFP cine images (k=0.52) and 96% (344 of 356) of vascular segments (k=0.18) were rated as good or excellent image quality with 86% of SSFP cine images having mild and 14% having moderate artifacts. The SNR and CNR of SSFP images and HR-MRA were higher at 3.0T (p<0.001) with off-resonance artifacts being more prevalent at 3.0T (45% of images at 3.0T vs 27%). However, they rarely rendered the images non-diagnostic.Cardiac MRI & MRA at 3.0T are feasible in children with CHD. Both field strengths can be used successfully for cardiac and vascular imaging; deciding which to use depends on local availability and the importance of vascular (extra-cardiac) vs. intra-cardiac imaging.Siemens Research Grant"}
+{"text": "The word \"Splice\" is misspelled in the article title. The correct title is: Broad Expression Analysis of Human ANTXR1/TEM8 Transcripts Reveals Differential Expression and Novel Splice Variants.The correct citation is: Vargas M, Karamsetty R, Leppla SH, Chaudry GJ (2012) Broad Expression Analysis of Human ANTXR1/TEM8 Transcripts Reveals Differential Expression and Novel Splice Variants. PLoS ONE 7(8): e43174. doi:10.1371/journal.pone.0043174"}
+{"text": "AbstractPerkinsyllis augeneri  and Prosphaerosyllis chauseyensis Olivier et al., 2011 are new records for the Mediterranean Sea. Detailed information is given on the morphology, ecology and distribution of the species recorded for the first time in the studied areas. In addition, an update on the distribution of the genus Prosphaerosyllis San Mart\u00edn, 1984 in the Mediterranean is given and an identification key to the Mediterranean species is provided.The syllid fauna of three locations in Crete and Israel (eastern Mediterranean Sea) was studied, yielding 82 syllid species, many of which were found for the first time in the respective areas: Seventeen species were recorded for the first time on the Israeli coasts and 20 in Greek waters.  Syllidae are a highly diverse family of polychaetes with currently around 900 valid species belonging to over 80 genera (pers. obs.) and have recently received considerable taxonomic and phylogenetic research effort, including a high number of new taxon descriptions (e.g. The ons e.g. . Syllidsons e.g. .http://www.nagisa.coml.org), a field project of the Census of Marine Life ; b) soft-sediment samples from the Israeli coast have been obtained in the framework of a project focusing on the soft bottom benthos of Haifa Bay. In all samples, Syllidae were highly abundant and yielded many species recorded for the first time in the respective area, as well as a species new to science  32\u00d735cm, volume 20 l, penetration 20 cm. The sediment was preserved in buffered formalin 10% for 3\u20137 days, then sieved through a 250 \u00b5m mesh sieve and subsequently stored in 70% ethanol. In this study, only a subset of the collected material is presented.Specimens from Israel were collected on 31 May 2009 and 11 Oct 2009 in Haifa Bay,  from soft sediments of mixed grain sizes in shallow waters . SedimenSpecimens from Crete were collected in September 2007 and June 2008 from two sites in northern Crete characterized by a continuous hard bottom habitat with dense algal coverage and a moderate wave exposure . At eachSyllidae .This manuscript was prepared in a Virtual Research Environment (Scratchpads) allowing for rapid and simultaneous publication of the results in print as well as electronically in a semantically enhanced form . This puhttp://ipt.pensoft.net/ipt/resource.do?r=easternmedsyllids). The data are furthermore available in Darwin Core Archive format, a simple and extensible schema for sharing biodiversity data which has been developed by the Global Biodiversity Information Facility  to allow easy and rapid mobilisation of species occurrence data through the internet. Darwin Core Archives are essentially a set of text files stored together with an XML descriptor file which describes the structure of the data files. Data are described through the Darwin Core schema, allowing for their usage within the semantic web. This new type of data publishing allows data to be indexed and discoverable through global biodiversity infrastructures such as GBIF or other data repositories, allows data to be integrated and compared with other datasets and ensures proper accreditation of the data provider (http://www.datadryad.org) and can be accessed at doi: 10.5061/dryad.4b7k408g.The underlying dataset of this study has been published under a Creative Commons license according to the Pensoft Data Publishing Policies and Guidelines for Biodiversity Data  and are provider . AdditioSyllinae and Exogoninae, whereas the samples from Israel did not contain any specimens of Anoplosyllinae or Autolytinae, and 73% of the examined species belong to the small-sized Exogoninae , CELA-5d-08 (2 ind.) [coll. 12.6.2008]; CELB-10c-07 (1 ind.) [coll. 27.9.2007].Gal\u00e1pagos Islands (Pacific Ocean).Gal\u00e1pagos Islands, north-east Pacific, Atlantic, Mediterranean Sea: WB, AS. New record for the Greek coast.Zostera beds, in vermetid reefs.Shallow subtidal depths, in sandy and muddy sediments, among photophilic algae and Imajima, 1966http://species-id.net/wiki/Syllides_japonicusSyllides japonicus Imajima, 1966: 112, figs 36a\u2013h; Syllidesjaponicus: cf. Elounda, Crete, Greece: CELA-15a-07 (1 ind.) [coll. 26.9.2007]; CELA-5d-08 (1 ind.), CELB-15d-08 (1 ind.) [coll. 12.6.2008].Japan (Pacific Ocean).Japan, Australia , MediterPosidonia oceanica rhizomes.Shallow subtidal depths, in sandy and muddy sediments, on rocks with algal cover, among GenusMilne Edwards, 1845Myrianida fasciata Milne Edwards, 1845http://species-id.net/wiki/Myrianida_inermisAutolytus inermis Saint-Joseph, 1887: 237, pl. 12, fig. 117; Autolytus (Autolytides) inermis : Myrianida inermis : Elounda, Crete, Greece: CELB-1e-07 (1 ind.) [coll. 29.9.2007].Dinard, France (north-east Atlantic Ocean).North-east Atlantic, north-west Atlantic , north-eUntil 100m depth, on rocks among algae and hydrozooans, in coralligenous substrates .http://species-id.net/wiki/Myrianida_quindecimdentataAutolytus quindecimdentatus Langerhans, 1884: 249, pl. 15, figs 3a\u2013b; Autolytus lugens Saint-Joseph, 1887: 234, pl. 12, fig. 116; Fauvel, 1923: 318, fig. 122g; Odontosyllis longicornis Hartmann-Schr\u00f6der, 1960: 98, figs 101\u2013104.Myrianida quindecimdentata : Alykes, Crete, Greece: CALA-10c-08 (4 ind.) [coll. 17.6.2008]; CALA-1b-08 (1 ind.), CALB-1c-08 (1 ind.), CALB-1d-08 (1 ind.) [coll. 18.6.2008]. Elounda, Crete, Greece: CELB-5e-07 (1 ind.) [coll. 27.9.2007]; CELB-1a-07 (2 ind.), CELA-1d-07 (2 ind.), CELB-1e-07 (5 ind.) [coll. 29.9.2007]; CELA-10b-08 (1 ind.) [coll. 11.6.2008]; CELB-1a-08 (1 ind.), CELB-1b-08 (1 ind.), CELA-5d-08 (1 ind.) [coll. 12.6.2008].Madeira (Atlantic Ocean).PageBreakEast and west Atlantic , north-east Pacific, Red Sea . MediterPosidonia oceanica rhizomes, endobiontic in sponges http://species-id.net/wiki/Perkinsyllis_augeneriPionosyllis augeneri Hartmann-Schr\u00f6der, 1979: 98, figs 119\u2013125; 1980a: 52; 1981: 32, fig. 52 Non : 35; SanPerkinsyllis augeneri : Haifa Bay, Israel: ALA-IL-7 (7 ind.) [coll. 11.10.2009].Boone, west Australia.Australia, New Zealand. Mediterranean Sea: LB. New record for the Mediterranean Sea.Intertidal and shallow subtidal depths, in coarse coralline sand, in muddy sand and seagrass beds .Prostomium pentagonal with 4 eyes in trapezoidal arrangement, posterior pair closer together than anterior one. Palps longer than prostomium, basally fused. Antennae cylindrical, smooth, longer than prostomium and palps. Tentacular cirri similar to antennae but slightly longer. Dorsal cirri of some anterior segments slender, longer than body width, some shorter, in midbody alternating short and long cirri, posteriorly all shorter than body width. Parapodia with 9\u201310 falcigers per fascicle anteriorly, 6\u20137 posteriorly. Shafts smooth or slightly serrated. Blades with marked dorso-ventral gradation , coarsely serrated, with small subdistal tooth. After proventriculum, dorsal blades unidentate, elongated, spiniger-like, twice as long as anteriorly, ventral blades stout, with strong serration, especially basally. Dorsal simple chaeta first appearing on midbody, blunt, subdistally serrated. Ventral simple chaetae posteriorly, bidentate, equally sized teeth forming a right angle, some long spines subdistally. Paired aciculae anteriorly, single ones posteriorly, with rounded, slightly enlarged tip. Pharynx through 4 chaetigers, pharyngeal tooth located anteriorly. Proventricle through 5 chaetigers with ca. 20\u201322 muscle cell rows.Perkinsyllis augeneri has not yet been fully resolved. In recent molecular phylogenies the species groups either within Exogoninae or as a sister group, and forms a sister clade of Eusyllinae in all analyses , recorded for the first time in 2003 in Cyprus .GenusFauvel, 1923Pionosyllis gestans Pierantoni, 1903http://species-id.net/wiki/Parapionosyllis_elegansPionosyllis elegans Pierantoni, 1903: 236, pl. X, fig. 2: pl. XI, fig. 27.Parapionosyllis elegans : Haifa Bay, Israel: ALA-IL-7 (11 ind.), ALA-IL-10 (45 ind.) [coll. 11.10.2009].Gulf of Naples (western Mediterranean Sea).North-east Atlantic (Iberian Peninsula). Mediterranean Sea: WB, CB, AD, AS, LB. New record for the Israeli coast.PageBreakUntil 30 m depth , in mediGenusSan Mart\u00edn, 1984Sphaerosyllis xarifae Hartmann-Schr\u00f6der, 1960San Mart\u00edn, 1984http://species-id.net/wiki/Prosphaerosyllis_adelaeSphaerosyllis (Prosphaerosyllis) adelae San Mart\u00edn, 1984a: 376, figs 1\u20134.Prosphaerosyllis adelae : San Mart\u00edn: 2003: 220, fig. 116.Haifa Bay, Israel: ALA-IL-7 (11 ind.) [coll. 31.5.2009]; ALA-IL-7 (6 ind.), ALA-IL-10 (2 ind.) [coll. 11.10.2009].Balearic Islands (western Mediterranean Sea).Mediterranean Sea: WB, LB. New record for the eastern Mediterranean Sea.Posidonia oceanica rhizomes.Until 13 m depth, in coarse sands, among http://species-id.net/wiki/Prosphaerosyllis_campoyiSphaerosyllis campoyi San Mart\u00edn Acero, Contonente and Gomez, 1982: 175, fig. 2; Sphaerosyllis (Prosphaerosyllis) campoyi : Prosphaerosyllis campoyi : San Mart\u00edn, 2003: 222, figs 117\u2013118.Elounda, Crete, Greece: CELA-10a-07 (1 ind.) [coll. 27.9.2009]; CELA-10b-08 (1 ind.) [coll. 11.6.2008].Andalusia, Spain (western Mediterranean Sea).North-east Atlantic , Mediterranean Sea: WB, AS, LB. New record for the Greek coast.Until 70 m depth , on rockThe specimens agree well with the description of Olivier, Grant, San Mart\u00edn, Archambault & McKindsey, 2011http://species-id.net/wiki/Prosphaerosyllis_chauseyensisProsphaerosyllis chauseyensis Olivier et al., 2011, figs 1\u20133a, b.Haifa Bay, Israel: ALA-IL-8 (12 ind.) [coll. 31.5.2009]; ALA-IL-1 (23 ind.), ALA-IL-2 (4 ind.), ALA-IL-5 (1 ind.), ALA-IL-7 (73 ind.), ALA-IL-8 (24 ind.), ALA-IL-9 (68 ind.), ALA-IL-10 (99 ind.) [coll. 11.10.2009].Sphaerosyllis brevicirra Hartmann-Schr\u00f6der, 1960  (Typ), 29.3.56, coll. Remane/Schulz]); Prosphaerosyllis chauseyensis , C1AM et C3AV]).Chausey Islands, Normandy (north-east Atlantic).North-east Atlantic (Normandy), Mediterranean Sea: LB. New record for the Mediterranean Sea.Until 13 m depth, in medium to very coarse sand.Three specimens collected at Station ALA-IL-8 on 31 May 2009 with egg capsules attached near dorsal cirri on midbody chaetigers.The specimens from Israel agree well with the specimens from Normandy, however, the Mediterranean specimens differ from the Holotype in: a) Papillation pattern: each segment with one papilla between dorsal cirri and four papillae, situated dorso-laterally and ventro-laterally on each side of parapodium, most developed in posterior chaetigers, from mid-body additional papillae arranged in two very irregular lines along middle of dorsum, increasing in length towards posterior end (ca. 20 \u00b5m posteriorly). Ventrally 2 smaller  papillae in middle of ventrum at posterior end of each segment. Specimens from Normandy have an irregular papillation pattern, but papillation is more distinct laterally, as in specimens from Israel; b) Length of anal cirri: about 125 \u00b5m, ca. 2.5\u20133 times length of posterior dorsal cirri  the ana.Sphaerosyllis tetralix Eliason, 1920, from the Gulf of Elat and the Mediterranean Sea might in fact belong to Prosphaerosyllis chauseyensis. The description and illustrations agree with many characteristics of Prosphaerosyllis chauseyensis, including the characteristic papilla on the dorsal cirri. However, Ben-Eliahu reports the species to have palps widely separated anteriorly (fused in Prosphaerosyllis chauseyensis), doPageBreakrsum with four longitudinal rows of papillae (irregular rows in Prosphaerosyllis chauseyensis) and the proventriculum stretching through 4 chaetigers (5 in Prosphaerosyllis chauseyensis). The material of the species described by Ben-Eliahu could not be examined during this study, therefore it can only tentatively be assigned to Prosphaerosyllis chauseyensis.Individuals identified by http://species-id.net/wiki/Prosphaerosyllis_longipapillataSphaerosyllis longipapillata Hartmann-Schr\u00f6der, 1979: 106, figs 148\u2013150; 1982: 71; 1984: 23; 1985: 71; 1986: 43; 1991: 40;Prosphaerosyllis longipapillata : Haifa Bay, Israel: ALA-IL-7 (2 ind.) [coll. 11.10.2009].ComparativeProsphaerosyllis longipapillata .Broome, north-west Australia.Australia, Mediterranean Sea: LB. New record for the Israeli coast.Sargassum vulgare  description by having alternating rows of long and short papillae on the dorsum  from California. To determine the identity of the Mediterranean material and whether Prosphaerosyllis bilineata and Prosphaerosyllis longipapillata are different species or not, careful examination of all type material is needed.The specimens from Israel agree well with the material and description of \u00c7inar et al., 2003). However, both the material from Cyprus and Israel, as well as the description and illustrations of e dorsum , fig. 5.\u00c7inar, Dagli & A\u00e7ik, 2011http://species-id.net/wiki/Prosphaerosyllis_marmaraeProsphaerosyllis marmaraeHaifa Bay, Israel: ALA-IL-2 (3 ind.), ALA-IL-8 (12 ind.) [coll. 31.5.2009]; ALA-IL-7 (4 ind.) [coll. 11.10.2009].Prosphaerosyllis marmarae . Prosphaerosyllis laubieri .PageBreakErdek, Marmara Sea (eastern Mediterranean).Mediterranean Sea: LB, Marmara Sea. New record for the Israeli coast.Until 17 m depth, in muddy sand , in coarProsphaerosyllis laubieriProsphaerosyllis marmarae. Both species have eyespots, strongly papillated palps, short, retractile antennae and dorsal cirri, pharynx and proventriculum each through 4 segments and short (8\u201310 \u00b5m) blades of falcigers. These two species differ however in the following characteristics: a) Prosphaerosyllis laubieri has small, scattered papillae all over the dorsum, in Prosphaerosyllis marmarae they are restricted to the lateral margins, near the dorsal cirri; b) cirrostyles of antennae and dorsal cirri of Prosphaerosyllis marmarae are much shorter  than those of Prosphaerosyllis laubieri  and appear as small, retracted caps; c) dorsal cirri of Prosphaerosyllis laubieri possess a small papilla at distal end of cirrophore . Prosphaerosyllis riseri Perkins, 1981 from Florida shares with Prosphaerosyllis marmarae the shape of the dorsal cirri and antennae (short and strongly retracted), however, its palps are less densely papillated. Prosphaerosyllis sp. A , but might in fact belong to Prosphaerosyllis marmarae. The morphological characteristics of her specimens agree very well wth those of Prosphaerosyllis marmarae , retractile cirri, falcigerous blades short (7.8 \u00b5m), proventriculum longer than proboscis (through 4 segments), no discernible dorsal papillation). Differences can be found in the cutting edge of the falcigerous blades which are smooth in the Red Sea specimens, whereas those of Prosphaerosyllis marmarae are serrated. However, due to the size of the blades (8 \u00b5m) this is a feature difficult to observe under an optical microscope and might have been overlooked. The material of the species described by Ben-Eliahu was not examined during this study, therefore it can only tentatively proposed to be assigned to Prosphaerosyllis marmarae.Specimens from the Red Sea described by http://species-id.net/wiki/Prosphaerosyllis_xarifaeSphaerosyllis xarifae Hartmann-Schr\u00f6der, 1960: 103, figs 121\u2013124; 1979: 103, figs 139\u2013140; 1980b: 56; 1981: 37; 1984: 25; PageBreakSphaerosyllis sp.: Sphaerosyllis cf. xarifae :Sphaerosyllis (Prosphaerosyllis) xarifae :Prosphaerosyllis xarifae : Haifa Bay, Israel: ALA-IL-10 (5 ind.) [coll. 11.10.2009]. Elounda, Crete, Greece: CELA-10b-08 (1 ind.) [coll. 11.6.2008]; CELA-5c-08 (1 ind.) [coll. 12.6.2008].Sarso, Red Sea.Circumtropical, Mediterranean Sea: WB, CB, AS, LB. New record for both the Israeli and Greek coasts.Until 40 m depth, euryoceous, among photophilic algae, in sand, mud, seagrasses, calcareous substrates .Specimens from Israel agree well with the description of GenusMcIntosh, 1885Salvatoria kerguelensis McIntosh, 1885http://species-id.net/wiki/Salvatoria_alvaradoiPseudobrania alvaradoiSalvatoria alvaradoi : Alykes, Crete, Greece: CALB-10b-08 (5 ind.), CALB-10d-08 (2 ind.) [coll. 17.6.2008]; CALB-5a-08 (2 ind.) [coll 18.6.2008]. Elounda, Crete, Greece: CELA-15a-07 (3 ind.), CELB-20e-07 (1 ind.) [coll. 26.9.2007], CELA-10a-07 (1 ind.) [coll. 27.9.2007]; CELB-1a-07 (1 ind.) [coll. 29.9.2007]; CELB-10a-08 (1 ind.), CELA-10b-08 (4 ind.), CELB-10b-08 (3 ind.), CELB-10c-08 (1 ind.), CELA-20a-08 (1 ind.), CELA-20d-08 (3 ind.) [coll. 11.6.2008]; CELA-5a-08 (8 ind.), CELA-5c-08 (18 ind.), CELA-5d-08 (1 ind.), CELB-15a-08 (1 ind.), CELB-15c-08 (9 ind.) [coll. 12.6.2008].Balearic Islands (western Mediterranean Sea).Mediterranean Sea: WB, CB, AS, Sea of Marmara . New recPosidonia oceanica rhizomes, in sediments with much organic material.PageBreakUntil 20 m depth, among algae with much sediment, among Sard\u00e1, 1984http://species-id.net/wiki/Salvatoria_euritmicaPseudobrania euritmica Sard\u00e1, 1984: 10, fig. 1.Grubeosyllis euritmica : Salvatoria euritmica : Pionosyllis yambaensis Hartmann-Schr\u00f6der, 1990: 52, figs 18\u201322.Alykes, Crete, Greece: CALB-20b-08 (1 ind.) [coll. 17.6.2008]; CALB-1d-08 (4 ind.) [coll. 18.6.2008]. Elounda, Crete, Greece: CELA-15c-07 (2 ind.) [coll. 27.9.2007]; CELB-1b-07 (4 ind.), CELA-1d-07 (1 ind.) [coll. 29.9.2007]; CELA-10b-08 (1 ind.), CELA-20c-08 (1 ind.) [coll. 11.6.2008]; CELB-15d-08 (1 ind.) [coll. 12.6.2008].Strait of Gibraltar (western Mediterranean Sea).Caribbean Sea, Australia, north-east Atlantic , Mediterranean Sea: WB, AS, LB. New record for the Greek coast.Until 20 m depth, on hard substrates between algae, in seagrass beds, on coralligenous substrates.Pionosyllis yambaensis was synonymized with Salvatoria euritmica by http://species-id.net/wiki/Salvatoria_neapolitanaPionosyllis neapolitana Goodrich, 1930: 651, figs 1\u201312.Pseudobrania neapolitanaGrubeosyllis neapolitana : Salvatoria neapolitana : Pionosyllis subterranea Hartmann-Schr\u00f6der, 1956: 89 figs 6\u20139.Brania subterranea : Grubeosyllis subterranea : Elounda, Crete, Greece: CELA-15a-07 (2 ind.), CELB-20c-07 (2 ind.) [coll. 26.9.2007]; CELB-15a-08: (5 ind.), CELB-15c-08 (1 ind.) [coll. 11.6.2008]; CELB-1d-08 (1 ind.) [coll. 12.6.2008].Bay of Naples, Italy (western Mediterranean Sea).Circumtropical, Mediterranean Sea: WB, AS . New recUntil 20 m depth, in coarse sand, among photophilic algae.Pionosyllis subterranea was synonymized with Pionosyllis neapolitana and transferred to Grubeosyllis by Grubeosyllis with Salvatoria, which has priority over the former.http://species-id.net/wiki/Salvatoria_vieiteziPseudobrania vieitezi San Mart\u00edn, 1984b: 160, figs 31\u201332.Grubeosyllis vieitezi : San Martin 1991a: 718, fig. 2e\u2013f; Salvatoria vieitezi : Alykes, Crete, Greece: CALA-10d-08 (1 ind.), CALA-15c-08 (1 ind.), CALA-20c-08 , CALB-20c-08 (1 ind.), CALB-20b-08 (1 ind.) [coll. 17.6.2008]; CALA-1b-08 (2 ind.), CALB-1b-08 (1 ind.), CALB-5a-08 (1 ind.) [coll. 18.6.2008]; CALB-20e-07 (1 ind.) [coll. 18.9.2007]; CALA-5c-07 (1 ind.) [coll. 19.9.2007]. Elounda, Crete, Greece: CELA-20d-07 (3 ind.) [coll. 26.9.2007]; CELA-10b-07 (1 ind.) [coll. 27.9.2007]; CELA-20c-08 (1 ind.), CELA-20d-08 (7 ind.) [coll. 11.6.2008]; CELA-5d-08 (1 ind.), CELB-15a-08 (1 ind.), CELB-1b-08 (5 ind.) [coll. 12.6.2008].Balearic Islands (western Mediterranean Sea).North-east Atlantic , Caribbean, Mediterranean Sea: WB, CB, AS. New record for the Greek coast.Posidonia oceanica rhizomes.Until 30m depth, on rocky substrates among photophilic algae, as endobiont of sponges, among http://species-id.net/wiki/Salvatoria_yraidaePseudobrania yraidae San Mart\u00edn, 1984b: 156, fig. 30.Grubeosyllis yraidae : Salvatoria yraidae : Alykes, Crete, Greece: CALB-10b-08 (1 ind.), CALB-15a-08 (1 ind.), CALB-20b-08 (3 ind.), CALB-20d-08 (1 ind.) [coll. 17.6.2008]. Elounda, Crete, Greece: CELA-15b-07 (1 ind.), CELA-15e-07 (2 ind.) [coll. 26.9.2007]; CELA-5c-07 (4 ind.) [coll. 27.9.2007]; CELA-10b-08 (3 ind.), CELB-10b-08 (8 ind.), CELB-10c-08 (1 ind.), CELA-20a-08 (1 ind.), CELA-20b-08 (1 ind.) [coll. 11.6.2008]; CELB-15a-08 (6 ind.), CELB-15c-08 (4 ind.), CELA-15d-08 (5 ind.), CELB-15d-08 (5 ind.) [coll. 12.6.2008].PageBreakBalearic Islands (western Mediterranean Sea).Mediterranean Sea: WB, CB, AD, AS. New record for the Greek coast.Until 20 m depth, in sandy substrates, on rocks among algae.GenusClapar\u00e8de, 1863Sphaerosyllis hystrix Clapar\u00e8de, 1863Southern, 1914http://species-id.net/wiki/Sphaerosyllis_bulbosaSphaerosyllis bulbosa Southern, 1914: 20, plates I\u2013II, figs 2a\u2013g; Fauvel, 1923: 304, figs. 116h\u2013r; Sphaerosyllis (Sphaerosyllis) bulbosa : Haifa Bay, Israel: ALA-IL-7 (4 ind.), ALA-IL-10 (51 ind.) [coll. 11.10.2009].Ireland (Atlantic Ocean).North-east Atlantic, Arctic Sea , New CalUntil 70 m depth, in sandy or muddy sediments, on calcareous substrates.The examined material differs from the description of Perkins, 1981http://species-id.net/wiki/Sphaerosyllis_glandulataSphaerosyllis glandulata Perkins, 1981: 1123, figs 18\u201319; Sphaerosyllisglandulata:  cf. Haifa Bay, Israel: ALA-IL-7 (1 ind.) [coll. 31.5.2009]; ALA-IL-7 (47 ind.), ALA-IL-10 (19 ind.) [coll. 11.10.2009]. Elounda, Crete, Greece: CELA-15d-08 (1 ind.) [coll. 12.6.2008].Florida, Hutchinson Island.West Atlantic , China  MediterrUntil 120 m depth, in calcareous habitats and fine to coarse sands, among photophilic algae.Sphaerosyllis glandulata.The specimens from Israel differ from San Mart\u00edn\u2019s (2003) description in having papillated palps and a longer proventriculum . Other characteristics, especially chaetal ones, agree well with former descriptions of Somaschini & San Mart\u00edn, 1994http://species-id.net/wiki/Sphaerosyllis_gravinaeSphaerosyllis gravinae Somaschini and San Mart\u00edn, 1994: 358, figs 1\u20132; Haifa Bay, Israel: ALA-IL-8 (4 ind.) [coll. 31.5.2009].Zannone Island, Italy (western Mediterranean Sea).Mediterranean Sea: WB, AD, LB. New record for the eastern Mediterranean Sea.Shallow subtidal depths, in medium to coarse sands, among algae.Perkins, 1981http://species-id.net/wiki/Sphaerosyllis_tayloriSphaerosyllis taylori Perkins, 1981: 1140, fig. 26; Haifa Bay, Israel: ALA-IL-1 (1 ind.); ALA-IL-2 (33 ind.) [coll. 31.5.2009]; ALA-IL-7 (103 ind.), ALA-IL-10 (14 ind.) [coll. 11.10.2009].Florida, Hutchinson Island.North-east and north-west Atlantic , Pacific Ocean  (Li\u00f1ero-Arana and D\u00edaz-D\u00edaz 2011), Arctic Sea , MediterPosidonia oceanica rhizomes.PageBreakShallow subtidal depths, in muddy to coarse sands with organic material, on rocks among photophilic or calcareous algae, among San Mart\u00edn 1984http://species-id.net/wiki/Sphaerosyllis_thomasiSphaerosyllis thomasi San Mart\u00edn, 1984b: 250, fig. 59; 2003: 199, figs 103\u2013104; Haifa Bay, Israel: ALA-IL-7 (2 ind.) [coll. 11.10.2009].Balearic Islands (western Mediterranean Sea).Mediterranean Sea: WB, CB, AD, AS, LB. New record for the Israeli coast.Posidonia oceanica rhizomes.Shallow subtidal depths, in muddy to coarse sands, among The examined specimens agree well with the description of GenusLangerhans, 1879Opisthosyllis brunnea Langerhans, 1879Langerhans, 1879http://species-id.net/wiki/Opisthosyllis_brunneaOpisthosyllis brunnea Langerhans, 1879: 541, fig. 7; Elounda, Crete, Greece: CELA-1d-07 (1 ind.) [coll. 29.9.2007], CELA-5d-08 (1 ind.) [coll. 12.6.2008].Madeira (Atlantic Ocean).Circumtropical. Mediterranean Sea: WB, CB, AS, LB. New record for the Greek coast.Intertidal to shallow subtidal, on hard substrates , endobiont of sponges.GenusLamarck, 1818Syllis monilaris Lamarck, 1818Moore, 1908http://species-id.net/wiki/Syllis_alternataSyllis alternata Moore, 1908: 323; 1909: 321; Typosyllis alternata : Alykes, Crete, Greece: CALB-15c-07 (1 ind.) [coll. 18.9.2007]; CALB-1a-07 (1 ind.) [coll. 19.9.2007]; CALA-10d-08 (2 ind.), CALA-15d-08 (1 ind.), CALB-20b-08 (1 ind.), CALA-20b-08 (2 ind.), CALA-20c-08 (5 ind.), CALB-20d-08 (6 ind.) [coll. 17.6.2008]. Elounda, Crete, Greece: CELB-20c-07 (1 ind.) [coll. 26.9.2007]; CELB-1a-07 (4 ind.) [coll. 29.9.2007]; CELA-10b-08 (1 ind.), CELB-10b-08 (1 ind.), CELA-10c-08 (1 ind.), CELB-10c-08 (1 ind.), CELA-10d-08 (2 ind.), [coll. 11.6.2008]; CELB-1a-08 (1 ind.), CELA-5b-08 (1 ind.), CELA-5d-08 (2 ind.), CELB-15c-08 (5 ind.) [coll. 12.6.2008].Alaska (Pacific Ocean).East Pacific (Alaska to Panama), west Atlantic (North Carolina to Cuba) , Japan , IndonesPosidonia oceanica rhizomes, calcareous algae, corals and photophilic algae  compacta Gravier, 1900: 165, pl. 9, fig. 11, text-figs 35\u201338.Syllis compacta : Syllis golfonovensisSyllis golfonovensis Hartmann-Schr\u00f6der, 1962. : PageBreak CELA-10a-07 (1 ind.), CELA-10d-07 (1 ind.) [coll. 27.9.2007]; CELA-5c-07 (1 ind.), CELB-5d-07 (1 ind.) [coll. 29.9.2007]; CELA-5b-08 (1 ind.), CELA-5d-08 (2 ind.), CELB-15d-08 (3 ind.) [coll. 12.6.2008].Alykes, Crete, Greece: CALB-1e-07 (1 ind.), CALA-5e-07 (1 ind.) [coll. 19.9.2007]; CALA-15c-08 (1 ind.), CALA-20b-08 (1 ind.), CALB-20b-08 (1 ind.), CALA-20c-08 (1 ind.) [coll. 17.6.2008]. Elounda, Crete, Greece: CELA-15b-07 (1 ind.), CELA-15e-07 (3 ind.), CELB-20a-07 (2 ind.) [coll. 26.9.2007];Red Sea.Red Sea. Mediterranean Sea: WB, CB, AD, AS. New record for the Greek coast.Posidonia oceanica rhizomes.Shallow subtidal depths, on biogenic calcareous substrates, among photophilic algae and Syllis variegata Grube, 1860. Recent works  [coll. 17.6.2008]. Elounda, Crete, Greece: CELB-10a-08 (1 ind.) [coll. 11.6.2008].Canary Islands (Atlantic Ocean).North-east Atlantic (Canary Islands), Mediterranean Sea: WB, CB, AD, AS, LB. New record for the Aegean Sea.Until 115 m depth, on coralligenous substrates, among photophilic algae, endobiont of sponges.http://species-id.net/wiki/Syllis_gerundensisTyposyllis gerundensis Al\u00f3s and Campoy, 1981: 21, figs 1\u20133; Syllis gerundensis : Alykes, Crete, Greece: CALA-20b-08 (1 ind.), CALB-20b-08 (1 ind.) [coll. 17.6.2008]; CALA-5d-08 (2 ind.) [[coll. 18.6.2008]. Elounda, Crete, Greece: CELB-1e-07 (1 ind.) [coll. 29.9.2007]; CELB-1d-08 (1 ind.), CELA-5d-08 (3 ind.) [coll. 12.6.2008].Columbretes Islands, Spain (western Mediterranean Sea).Mediterranean Sea: WB, CB, AD, AS, LB.New record for the Aegean Sea.Posidonia oceanica rhizomes and photophilic algae, endobiont of sponges.Shallow subtidal depths, on calcareous grounds, sandy bottoms, among San Mart\u00edn & L\u00f3pez, 2000http://species-id.net/wiki/Syllis_jorgeiSyllis jorgei San Mart\u00edn and L\u00f3pez, 2000: 430, figs 4\u20136; Typosyllis lutea : Syllis luteaSyllis lutea Hartmann-Schr\u00f6der, 1960). : Haifa Bay, Israel: ALA-IL-7 (3 ind.) [coll. 11.10.2009]. Alykes, Crete, Greece: CALA-20c-07 (1 ind.) [coll. 18.9.2007], CALB-1a-08 (1 ind.) [coll. 18.6.2008]. Elounda, Crete, Greece: CELB-1a-08 (1 ind.), CELA-1c-08 (1 ind.), CELB-5d-08 (1 ind.) [coll. 12.6.2008].Columbretes Islands, Spain (western Mediterranean Sea).East Atlantic (Canary Islands), Mediterranean Sea: WB, CB, AD, AS, LB. New record for the Israeli coast.Posidonia oceanica rhizomes and photophilic algae.Until 145 m depth , on bioghttp://species-id.net/wiki/Syllis_pulvinataTyposyllis pulvinata Langerhans, 1881: 97, 104; Syllis pulvinata : Syllis (Typosyllis) truncata mediterranea Ben-Eliahu, 1977a: 10, fig. 2.Syllis mediterranea : San Mart\u00edn, 1984b; 209, fig. 8.Elounda, Crete, Greece: CELA-1b-08 (1 ind.), CELA-5c-08 (2 ind.), CELB-15d-08 (1 ind.) [coll. 12.6.2008].Canary Islands (Atlantic Ocean).North-east Atlantic (Cantabrian Sea to Canary Islands), Red Sea, Mediterranean: WB, CB, AD, AS, LB. New record for the Aegean Sea.PageBreakShallow subtidal depths, on calcareous substrates (vermetid reefs), among photophilic algae, endobiont of sponges.http://species-id.net/wiki/Syllis_tyrrhenaTyposyllis tyrrhena Licher and Kuper, 1998: 228, figs 1\u20134; Syllis tyrrhena : Elounda, Crete, Greece: CELB-10b-08 (1 ind.) [coll. 11.6.2008].Island of Elba, Italy (western Mediterranean Sea).Brazil , MediterUntil 13 m depth, in sandy substrates of mixed grain sizes , on rockSan Mart\u00edn, 1984http://species-id.net/wiki/Syllis_westheideiSyllis westheidei San Mart\u00edn, 1984b: 403, figs 108\u2013109; 2003: 436, figs 240\u2013241; Typosyllis westheidei : Typosyllis variegataSyllis variegata Grube, 1860). : Alykes, Crete, Greece: CALB-15d-08 (1 ind.) [coll. 17.6.2008].Balearic Islands (western Mediterranean Sea).Pacific Ocean , Red Sea, Mediterranean: WB, CB, AD, AS. New record for the Greek coast.Posidonia oceanica rhizomes and vermetid reefs.Shallow subtidal depths, on hard substrates, among photophilic algae, in GenusClapar\u00e8de 1864Syllis zebra Grube, 1860Clapar\u00e8de, 1868http://species-id.net/wiki/Trypanosyllis_coeliacaTrypanosyllis coeliacaPseudosyllis brevipennis Grube, 1863: 44, pl. 4, fig. 5.Haifa Bay, Israel, eastern Mediterranean Sea, Station ALA-IL-7 (1 ind.) [coll. 11.10.2009]. Alykes, Crete, Greece: CALA-10b-08 (1 ind.), CALB-10c-08 (1 ind.) [coll. 17.6.2008]; CALA-5a-08 (1 ind.), CALB-1d-08 (2 ind.), CALB-5a-08 (1 ind.) [coll. 18.6.2008]. Elounda, Crete, Greece: CELA-15b-07 (1 ind.), CELA-15c-07 (1 ind.) [coll. 26.9.2007]; CELB-5c-07 (1 ind.), CELA-10a-07 (1 ind.), CELB-10c-07 (1 ind.) [coll. 27.9.2007]; CELB-1a-07 (2 ind.), CELB-1e-07 (1 ind.) [coll. 29.9.2007]; CELB-10b-08 (1 ind.), CELA-15a-08 (1 ind.) [coll. 17.6.2008]; CELB-1b-08 (1 ind.), CELA-5b-08 (1 ind.), CELA-5c-08 (1 ind.), CELB-5c-08 (1 ind.), CELA-5d-08 (2 ind.) [coll. 18.6.2008].Gulf of Naples (western Mediterranean Sea).Circumtropical. Mediterranean Sea: WB, CB, AD, AS, LB. New record for the Israeli coast.Posidonia oceanica rhizomes, in vermetid reefs, in coarse sand.From infralitoral depths to 760 m, on hard substrates, among algae, corals, hydrozoans, sponges and Trypanosyllis coeliaca have in the past been identified as Pseudosyllis brevipennis Grube, 1863, but according to Pseudosyllis brevipennis is regarded as a synonym of Trypanosyllis coeliaca.Specimens from Greece have a faint or no visible trepan. Individuals without trepan but otherwise identical to Exogoninae  in the Mediterranean belongs to these doubtful records. Records of Sphaerosyllis brevicirra from the western MediterraneanPageBreak Sea by Prosphaerosyllis species  description but in fact absent  likewise do probably not belong to Prosphaerosyllis brevicirra: Ben-Eliahu\u2019s (1977a) redescription of the species based on material from the Gulf of Elat (Red Sea) differs in several aspects from Hartmann-Schr\u00f6der\u2019s (1960) descripPageBreaktion and from the type material. In particular, Ben-Eliahu does not mention or illustrate the papilla on the dorsal cirrus, her specimens have four eyes and one anterior pair of eyespots  and the proventriculum occupies 4 chaetigers (3 in Prosphaerosyllis brevicirra). According to the description and illustrations, the species might in fact belong to Prosphaerosyllis marmarae (see remarks for this species above). The record of Sphaerosyllis brevicirra from the Spanish Atlantic coast . The species Prosphaerosyllis brandhorsti  has been recorded in Italy by Prosphaerosyllis ranunculus . The presence of Prosphaerosyllis brandhorsti in the Mediterranean Sea has thus to be considered as doubtful. An identification key to the currently valid Mediterranean species of Prosphaerosyllis can be found below.The present study yielded a number of species reported for the first time in the respective areas, and a high number of the new additions belong to the subfamily ogoninae . This coall e.g. . The Exo species . These dt absent ), the ab species , are absic coast , though"}
+{"text": "Thrombosis, mainly venous, is a rare and well-recognized extraintestinal manifestation of inflammatory bowel disease (IBD). We describe a 25-year-old Caucasian man affected by ulcerative colitis and sclerosing cholangitis with an episode of right middle cerebral arterial thrombosis resolved by intraarterial thrombolysis. We perform a brief review of the International Literature. Thrombosis, mainlyh venous, is a rare and well-recognized extraintestinal manifestation of inflammatory bowel disease (IBD). Arterial Thromboembolic (TE) complications, and in particular strokes, occur less frequently . They usA 25-year-old Caucasian man was observed in the emergency room for a sudden left-sided hemiparesis characterized by sensitive impairment, confusion and bladder incontinence. NIH stroke scale score (NIHSS) impairment was 8. A diagnosis of ulcerative colitis (UC) and sclerosing cholangitis (SC) had been made 8 years before. He also referred an episode of acute pancreatitis 7 years before as a complication of mesalazine therapy. He had no familial history of cerebral vascular disease (CVD). At clinical examination the patient was normotensive and without fever. Blood tests showed iron deficiency anemia , raised white cell counts (WBC 12.470/mmc-n.v. 4000\u201311000/mmc), erythrocytes sedimentation (ERS) 56 (n.v. < 10), reactive C protein 7.6\u2009mg/L, (n.v.\u2014until 5\u2009mg/dL), gamma Glutamil trans-peptidase (297\u2009U/L-n.v. 11\u201349), gamma globulin 2.55\u2009g/dL, (n.v. until 1.8\u2009g/dL) antinuclear antibodies 1\u2009:\u2009640 (v.n. negative). An emergency brain angio-computed tomography (A-CT) evidenced thrombosis of the M2 tract of the right middle cerebral artery, without any (apparent) other brain lesions . After aThe association of inflammatory bowel disease (IBD) and thrombosis was, for the first time, described in 1936 by Bargen and Baker . Patters"}
+{"text": "The objective of this study was to evaluate the efficacy, tolerability, and safety of fosamprenavir/ritonavir (FPV/r) versus efavirenz (EFV), both in combination with abacavir/lamivudine (ABC/3TC), in a population that is often underrepresented in U.S. clinical trials.In this ongoing 96-week, open-label, prospective, randomized, multicenter study, we compared once-daily ABC/3TC 600 mg/300 mg with FPV 1400 mg/r 100 mg or EFV 600 mg in ARV-na\u00efve, HIV-1-infected subjects with entry viral load (VL) >5,000 c/mL, were HLA-B*5701 negative, and did not have major resistance mutations to study drugs. The primary endpoint was time to switch of third drug or time to development of any treatment-related Grade 3 or 4 adverse events (AEs). Results from the planned 48-week analysis are reported.3 in each group. Rate of treatment-related grade 2-4 AEs was lower in the FPV/r-arm  vs. the EFV-arm  primarily due to EFV-related rash and dizziness (8% each). Rates of treatment-related serious AEs and grade 3-4 lab abnormalities were similar between FPV/r vs. EFV. A total of 8 virologic failures occurred through W48. At failure, HIV PRO or RT treatment-emergent mutations were present in 4 of 5 EFV patients and 1 of 3 FPV/r patients selected an RT mutation. Median change from BL in total/HDL cholesterol ratio was unchanged in both groups but the FPV/r arm had larger changes in triglycerides (32 vs. 7 mg/dL) and in LDL cholesterol (22 vs. 11 mg/dL).SUPPORT enrolled 32% (32/101) women and 79% (80/101) non-Caucasians. Baseline and demographic characteristics were generally similar between groups. A total of 84 subjects (83%) completed study through W48. Eight patients met the primary endpoint: 3 (6%) and 5 (10%) on FPV/r and EFV, respectively. At W48, by ITT-Exposed missing-equals-failure analysis, 76% (39/51) and 82% (41/50) of subjects achieved VL <50 c/mL on FPV/r vs. EFV, respectively. Median change from baseline to W48 in CD4 cell count was 178 cells/mmThrough 48 weeks, in a diverse population, virologic/immunologic responses were not demonstrably different between FPV/r and EFV when given with ABC/3TC, but the EFV regimen had slightly more patients meeting the tolerability endpoint, treatment-related grade 2-4 AEs, virologic failures, and treatment-emergent mutations at failure."}
+{"text": "KRAS wild-type and mutant metastatic colorectal cancer (MCRC) patients (pts) treated with bevacizumab (BEV)-containing chemotherapy is not significantly different. Since specific KRAS mutations confer different aggressive behaviors, the prognostic role of prevalent KRAS mutations was retrospectively evaluated in MCRC pts treated with first line FIr-B/FOx, associating BEV to triplet chemotherapy.Prognosis of KRAS codon 12, 13 and BRAF V600E mutations by SNaPshot and/or direct sequencing. MCRC pts <75-years-old were consecutively treated with FIr-B/FOx: weekly 12 hour-timed-flat-infusion/5-fluorouracil , irinotecan plus BEV ; and oxaliplatin . Pts were classified as liver-limited (L-L) and other/multiple metastatic (O/MM). Progression-free survival (PFS) and overall survival (OS) were compared using the log-rank test.Tumor samples were screened for KRAS mutant pts: c.35 G > A, 15 (25.4%); c.35 G > T, 7 (11.8%); c.38 G > A, 3 (5%); other, 3 (5%). KRAS wild-type, 31 pts (52.7%). The objective response rate (ORR), PFS and OS were, respectively: c.35 G > A mutant, 71%, 9 months, 14 months; other than c.35 G > A mutants, 61%, 12 months, 39 months. OS was significantly worse in c.35 G > A pts compared to KRAS wild-type (P = 0.002), KRAS/BRAF wild-type (P = 0.03), other MCRC patients (P = 0.002), other than c.35 G > A (P = 0.05), other codon 12 (P = 0.03) mutant pts. OS was not significantly different compared to c.35 G > T KRAS mutant (P = 0.142).Fifty-nine pts were evaluated at a median follow-up of 21.5 months. KRAS c.35 G > A mutant status may be significantly associated with a worse prognosis of MCRC pts treated with first line FIr-B/FOx intensive regimen compared to KRAS/BRAF wild type and other than c.35 G > A mutant pts. KRAS genotype, wild-type or mutant, addresses the medical treatment of metastatic colorectal cancer (MCRC) patients (pts), consisting of triplet regimens combining chemotherapeutic drugs, or doublets plus targeted agents [KRAS mutant patients [KRAS wild-type and mutant pts [d agents . The addpatients ,3; anti-tant pts ,5. In litant pts -13.KRAS genotype can be assessed by evaluation of clinical outcome , overall survival (OS)) in wild-type and mutant pts, depending on differential tumor biological aggressiveness and predictive effectiveness of treatment strategies. The median OS of KRAS wild-type and mutant MCRC pts treated with irinotecan, 5-fluorouracil and leucovorin (IFL) plus bevacizumab (BEV) was 27.7 and 19.9 months, respectively [KRAS and BRAF wild-type pts were compared with pts harboring mutations in one gene. In KRAS wild-type pts and in BRAF wild-type pts compared to mutant, HR was 0.64 and 0.38, respectively, but did not reach statistical significance [et al. [KRAS wild-type and mutant genotypes [KRAS wild-type and mutant pts , but not significantly [KRAS wild-type pts with L-L disease may achieve a significantly greater benefit from integration with liver metastasectomies, with respect to KRAS mutant patients [The prognostic relevance of the ectively ,5. The henotypes . Median ficantly . L-L ptspatients ,13.KRAS wild-type genotype predicts favorable clinical outcomes when anti-EGFR or anti-VEGF molecules are added to doublet chemotherapy [KRAS mutant genotype significantly predicts prolonged PFS up to 9.3 months, while there was no increase in OS and activity [The otherapy ,5. The Kactivity ,5, in ptKRAS mutations occur in 35% to 45% of colorectal cancer (CRC), mostly in codon 12 (80%), c.35 G > A (G12D) and c.35 G > T (G12V) transversions, representing 32.5% [in vitro model proposed by Guerrero et al. [ng 32.5% ,15 and 2ng 32.5% ,16, respng 32.5% . These mng 32.5% . In the o et al. , codon 1o et al. . Codon 1o et al. .KRAS mutant tumors seems to confer worse clinical behavior. A multivariate analysis suggested that the presence of KRAS mutation significantly increased the risk of recurrence and death; the codon 12 c.35 G > T (G12V) mutation retained an independent increased risk of recurrence and death [KRAS mutations was not confirmed in other studies [The biological aggressiveness of codon 12 nd death , and signd death . The poo studies ,23.KRAS mutation in MCRC pts enrolled in a previously reported phase II study [We report a retrospective exploratory analysis evaluating the prognostic value of the prevalent codon 12 c.35 G > A (G12D) II study  and in aMCRC pts were enrolled in a previously reported phase II study  and in tFIr-B/FOx association consisted of 5-fluorouracil associated with alternating irinotecan/BEV or oxaliplatin, according to a previously reported weekly schedule .KRAS and BRAF analyses were performed on paraffin-embedded tissue blocks from the primary tumor and/or metastatic site. Genotype status was analyzed for KRAS codon 12 and 13 mutations and BRAF c.1799 T > A (V600E) mutation by SNaPshot\u00ae multiplex [KRAS mutations and KRAS/BRAF mutations in 36 and 32 samples, respectively; direct sequencing of the KRAS gene was performed in 23 samples. After treatment with xylene thiocyanate and selection of tumor cell clusters, DNA was isolated using the RecoverAll\u2122 Total Nucleic Acid Isolation Kit for FFPE Tissues  according to the manufacturer's instructions.ultiplex , for KRA\u00ae multiplex assay was performed as previously reported [KRAS exon 2 and BRAF exon 15 were simultaneously PCR-amplified and analyzed for the presence of mutations at KRAS nucleotides c.34G, c.35G, c.37G, c.38G and BRAF mutation at nucleotide c.1799T using the ABI PRISM SNaPshot\u00ae Multiplex kit  [xl Genetic Analyzer (Applied Biosystems). Data were analyzed using the GeneMapper Analysis Software version 4.0 (Applied Biosystems).SNaPshotreported ,25. KRASCA, USA) ,25. LabeKRAS exon 2 sequence reaction was performed from PCR-amplified tumor DNA, using the Big Dye V3.1 Terminator Kit (Applied Biosystems), and run on an automated sequencer .KRAS mutant genotype on the clinical outcome of MCRC pts treated with first line FIr-B/FOx. Pts were classified as L-L and O/MM [A retrospective analysis has been planned to evaluate the prognostic relevance of the prevalent codon 12 c.35 G > A (G12D) and O/MM . Clinicaand O/MM ; patholoand O/MM . PFS andand O/MM . The logand O/MM . PFS wasand O/MM . The resThe KRAS/BRAF genotype was evaluated in 59 pts, among 64 consecutive, unselected MCRC pts recruited in the phase II study and expanded clinical enrollment of the FIr-B/FOx regimen as the first line treatment of MCRC [KRAS wild-type and 28 (47%) as KRAS mutant [KRAS mutations was: codon 12, 24 pts (40.6%), specifically c.35 G > A (G12D), 15 pts (25.4%), c.35 G > T (G12V), 7 pts (11.8%), c.34 G > A (G12S) and c.35 G > C (G12A), 1 patient each; codon 13, 4 pts (6.7%), c.38 G > A (G13D), 3 pts (5%) and c.37_39 dupl, 1 patient , 8 (61.5%) and 19 pts (61%), respectively.Table KRAS mutant, other KRAS mutant, and KRAS wild-type pts was, respectively: L-L 6 pts (40%), 7 pts (54%), and 12 pts (39%); O/MM 9 pts (60%), 6 pts (46%), 19 pts (61%).Pts' distribution according to extension of metastatic disease in c.35 G > A KRAS wild-type and mutant pts at a median follow-up of 21.5 months were previously reported [KRAS mutant pts . We observed 8 partial responses (61%), 2 stable diseases (15%), and 3 progressive diseases (23%). Median PFS was 12 months (3-37 months): 12 events occurred and 1 patient (8%) was progression-free >12 months. Median OS was 39 months (8-59+ months): 8 events occurred and 5 pts (38%) were alive. Liver metastasectomies were performed in 5 pts out of 13 (38%) and out of 7 (71%) with L-L metastases; 4 R0 liver resections (57%). Two pathologic CRs were obtained (15%) in pts with multiple L-L metastases, harboring codon 12 mutations, c.35 G > T and c.34 G > A: 1 patient progressed at 17 months, 1 patient was progression-free at 35 months. Clinical outcome according to extension of metastatic disease, L-L and O/MM [Among 13 other than c.35 G > A and O/MM ,13, was:KRAS wild-type pts was previously reported [KRAS/BRAF wild-type pts [Activity and efficacy among 30 evaluable reported : ORR wastype pts , ORR wasKRAS mutant plus KRAS wild-type pts, ORR was 81% , median PFS was 13 months (1+-69+ months) and median OS was 34 months (1+-69+ months) , median PFS was 12 months (1+-60+ months) and median OS was 20 months (1+-60+ months). Among 7 c.35 G > T KRAS mutant pts, ORR was 57% , median PFS was 12 months (3-5 months) and median OS was 21 months (11-46+ months). Among 4 codon 13 KRAS mutant pts, ORR was 75% , median PFS was 12 months (7-37 months) and median OS was 44 months (8-59+ months). Among 3 c.38 G > A KRAS mutant pts, ORR was 67% , median PFS was 12 months (7-37 months) and median OS was not reached (8-59+ months).Among 44 evaluable other than c.35 G > A KRAS mutant pts compared to KRAS wild-type pts was not significantly different while OS was significantly worse (P = 0.002). In addition, c.35 G > A KRAS mutant pts compared to other than c.35 G > A KRAS mutant pts showed significantly worse OS (P = 0.05); other than c.35 G > A KRAS mutant pts compared to KRAS wild-type pts did not have different OS ; KRAS/BRAF wild-type pts (P = 0.03); and other codon 12 mutant pts (P = 0.03)   [The prognostic relevance of tant pts ,5,8,13. tant pts . BEV addKRAS mutation characterizes 10.3% of CRC and represents up to 30% of KRAS mutations [KRAS mutation and exhibited a high activity of the FIr-B/FOx intensive regimen (ORR 71%). Liver metastasectomies were performed in 13% of pts (33% of L-L disease), median PFS and OS were 9 and 14 months, respectively. In pts with the KRAS c.35 G > A mutation, activity and PFS were not significantly different, while OS was significantly worse compared to KRAS wild-type, KRAS/BRAF wild-type, and other codon 12 and 13 mutant pts. Median OS was not significantly different in other KRAS mutant compared to wild-type pts. This is the first report of a worse prognosis in KRAS c.35 G > A (G12D) mutant MCRC pts, treated with intensive triplet chemotherapy plus BEV.The prevalent c.35 G > A (G12D) utations . In the KRAS mutations may increase aggressiveness by the differential regulation of KRAS downstream pathways associated with higher AKT/protein kinase B activation, bcl-2, E-catherin, \u03b2-catenin, and focal adhesion kinase overexpression, and RhoA underexpression, whereas codon 13 KRAS mutant cells show increased sensitivity associated with increased activation of the c-Jun-NH2-terminal kinase I pathway [KRAS codon 12 with codon 13 mutations in CRC. RASCAL (Kirsten Ras in CRC) studies showed that the presence of the KRAS mutation significantly increased the risk of death by 26% [KRAS codon 12 mutations  were associated with inferior survival in patients with KRAS-wild-type/BRAF-wild-type cancers [Codon 12  pathway . Severalh by 26% ,21; the h by 26% , and up h by 26% . KRAS co cancers .BRAF and KRAS mutations can confer different biological aggressiveness and effectiveness of treatment strategies; the balance between aggressiveness and effectiveness can differentiate prognosis, that is, median OS. Comparison of median OS in pts with different genotypes can discriminate this net prognostic effect. Thus, specific mutations and treatment strategies  could be major parameters determining different prognoses in MCRC. The prevalent BRAF c.1799 T > A (V600E) mutation, characterizing 4.7% to 8.7% of CRC, demonstrated a negative prognostic effect compared to BRAF wild-type pts in MCRC pts treated with doublet chemotherapy alone or added to cetuximab, BEV and cetuximab plus BEV, with a median PFS of 5.6 to 8 months and median OS of 10.3 to 15.9 months [BRAF mutant MCRC pts [KRAS c.35 G > T mutation and other mutations were associated with a worse outcome when receiving chemotherapy plus cetuximab, compared with chemotherapy alone [In MCRC pts, specific 9 months ,31. The MCRC pts . Patientpy alone .KRAS c.38 G > A mutation (G13D) confers a significantly worse prognosis [KRAS mutations [KRAS wild-type pts [KRAS mutation in first line cetuximab-containing chemotherapy [KRAS c.38 G > A (G13D) mutant pts demonstrated a significantly favorable predictive effect of cetuximab-containing associations compared to other KRAS mutant MCRC, and significantly lower ORR, with no significantly different PFS and OS compared to KRAS wild-type pts [KRAS mutations and patient outcome. Opposite findings were reported when panitumumab was combined with first line oxaliplatin, whereas similar data were reported when it was combined with second-line FOLFIRI [In MCRC pts pre-treated with chemotherapy alone, the rognosis . Cetuximutations , and no type pts . Recentlotherapy : signifitype pts ,36. In p FOLFIRI .KRAS/BRAF mutations, specifically KRAS c.35 G > A (G12D), c.35 G > T (G12V), c.38 G > A (G13D) mutations and BRAF c.1799 T > A (V600E).Prospective studies should be developed to confirm the differential prognosis and predictive effect of chemotherapeutics and/or targeted agents in MCRC pts harboring KRAS c.35 G > A (G12D) mutant genotype has a significantly worse effect on the OS of MCRC pts treated with the first line FIr-B/FOx intensive regimen compared to wild-type pts or to pts harboring different other KRAS mutations, due to heterogeneous biological aggressiveness and the effectiveness of treatment strategies. The present findings should be verified in prospective trials of multidisciplinary strategies comparing clinical outcome in MCRC pts harboring specific mutations that differentially activate the downstream RAS-MAPK or PI3K pathways.The prevalent Anti-EGFR: anti-epidermal growth factor receptor; anti-VEGF: anti-vascular endothelial growth factor; BEV: bevacizumab; cCR: clinical complete responses; CR: complete response; CRC: colorectal cancer; CT: computed tomography; HR: hazard ratio; IFL: irinotecan, 5-fluorouracil, and leucovorin; L-L: liver-limited; MCRC: metastatic colorectal cancer; O/MM: other/multiple metastatic; ORR: objective response rate; OS: overall survival; PET: positron emission tomography; PFS: progression-free survival; pts: patients; RECIST: Response Evaluation Criteria in Solid Tumors.The authors declare that they have no competing interests.GB contributed to the conception and design of the study, in the provision of study materials of patients, in data analysis and interpretation and in the manuscript writing. ER contributed to the conception and design of the study, in data analysis and interpretation and in the manuscript writing. KC, TF, MT, EA participated in data analysis and interpretation. GB, KC, CF and ER provided clinical management and data on patients. DDG, AL, JCS provided molecular genetic analysis. All authors participated in the collection and/or assembly of data. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1741-7015/11/59/prepub"}
+{"text": "Previous studies suggest yoga is effective for mild to moderate chronic low back pain (CLBP) in mostly white higher socioeconomic status (SES) populations. However, little is known regarding yoga\u2019s optimal dose or its effectiveness for more severe CLBP in diverse lower SES populations.From September-December 2011, we conducted a 12-week RCT comparing once vs. twice-weekly standardized 75-minute hatha yoga classes for 95 adults with nonspecific CLBP. Recruitment and classes occurred in a large safety-net hospital and five affiliated community health centers in Boston, Massachusetts. Primary outcomes were mean low back pain intensity in the previous week (0-10) and back-related function . We used two-sample t-tests to compare once/week vs. twice/week mean change scores (baseline-12 weeks) for pain and MRMD. Analyses used the intention-to-treat principle.<$30,000, and 35% with high school education or less. Baseline pain intensity  and MRMD  were consistent with moderate-severe CLBP. Baseline characteristics of the once/week (n=49) and twice/week (n=46) groups were similar. Overall class attendance was 73% and 62% for the once/week and twice/week participants, respectively. Both groups practiced yoga at home on average 3-4 days/week. Each group experienced statistically significant (p<.0001) and clinically meaningful improvements in pain and function: Mean pain change scores for the once/week and twice/week groups were -2.1 (SD 2.7) and -2.4 (SD 2.2), respectively. Mean MRMD change scores for the once/week and twice/week groups were -5.2 (SD 6.5) and -4.9 (SD 4.4), respectively. There were no statistically significant differences between the two groups for pain or MRMD.Participants were on average 48 years old, 76% female, 82% non-white, 63% with annual household incomes Twelve weeks of either once or twice-weekly hatha yoga classes augmented by home practice were similarly effective for moderate to severe CLBP in a diverse predominantly lower SES population."}
+{"text": "In recent years a large amount of literature data emphasizes the interrelationship between pathogenic mechanisms of chronic hepatitis C virus (HCV) infection and lipid and glucid metabolism. In this study we aimed to characterize lipid and glucidic metabolic patterns in chronically HCV-infected patients and to evaluate the role of HCV on cardiovascular risk (CVR).1c), liver enzymes, and viral load (VL). Liver histology was assessed by Fibromax (Biopredictive).We conducted a cross-sectional analysis on chronic HCV-infected adult patients, monitored at the National Institute of Infectious Diseases \u201cProf. Dr. Matei Bal\u015f\u201d. Patients with diabetes mellitus, chronic alcohol consumption, other chronic liver diseases, HBV or HIV co-infections were excluded. We recorded demographic data, HCV infection history, personal and family history of CVR factors. We measured height and weight for body mass index, waist to hip ratio, blood pressure and we assessed the 10 years CVR using Framingham score. Blood samples were tested for lipid profile, serum glucose, glycosylated hemoglobin (HbA1c levels (p=0.022).Seventy-six patients with a median age of 51years (IQR 44.25-58.0) were included. Sex ratio was F:M=1.53. Median VL was 118500 IU/mL (IQR 0-600951). Twenty-five percent (19/76) of the patients had no fibrosis (F0), 51.3% (39/76) had hepatitis (F1-2), 6.5% (5/76) had a fibrosis score equivalent to transition to cirrhosis (F3), and 17.1% (13/76) had cirrhosis (F4). Mean serum cholesterol, LDL and triglycerides were 187 mg/dL (IQR 166-220), 119 mg/dL (IQR 93-147) and respectively 94 mg/dL (IQR 69.25-132.0). Patients with no fibrosis were more frequently younger and females , had higher cholesterol (p=0.014) and LDL levels (p=0.009) and lower VL (p=0.017) and CVR according to Framingham score (p=0.000). Patients with cirrhosis were more frequently males (p=0.033), and had higher viral load (p=0.011) and serum glucose (p=0.027). Fibrosis score correlated to age (p=0.000), VL (p=0.001), higher CVR (p=0.001), low LDL (p=0.034), high glucose (p=0.031). The VL correlated to HDL (p=0.023) and with lower HbAIn patients with chronic HCV infection although high fibrosis correlates to better lipid profiles, it also correlates to higher cardiovascular risk."}
+{"text": "The correct title is An error was introduced in the preparation of this article for publication. The word \"At2 Receptor Antibodies Are Nonspecific\". \"Commercially Available Angiotensin II ATThe correct citation is: 2 Receptor Antibodies Are Nonspecific. PLoS ONE 8(7): e69234. doi:10.1371/journal.pone.0069234Hafko R, Villapol S, Nostramo R, Symes A, Sabban EL, et al. (2013) Commercially Available Angiotensin II AT"}
+{"text": "This crystalline monohydrate addition product, formed by reaction of d-glucose and sodium hydrogen sulfite in water, forms a three-dimensional network through complex cation coordination and extensive inter\u00admolecular hydrogen bonding. Each of the independent mol\u00adecules has an open-chain structure with the carbon chains adopting a sickle-like conformation, similar to that found in the potassium salt , but there are significant differences in the patterns of complexation.The title salt, Na al. 2001. Carbohy DOI: 10.1107/S1600536812007210/su2377Isup2.cdxSupplementary material file. DOI: 10.1107/S1600536812007210/su2377Isup3.hklStructure factors: contains datablock(s) I. DOI: 10.1107/S1600536812007210/su2377Isup4.cmlSupplementary material file. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:"}
+{"text": "Several studies have described a high prevalence of 25[OH]D deficiency in HIV-infected patients, as well as its role in predicting disease progression.We performed a descriptive, cross-sectional study on a group of 68 HIV-infected patients monitored in the Regional HIV/AIDS Centre Mure\u015f, during December 2012-February 2013. We determined plasma levels of 25[OH]D. We estimated the proportion of patients with 25[OH]D deficiency/insufficiency and correlated 25[OH]D levels to HIV-RNA plasma viral load, CD4+ T-cells count and associated pathology. Data were compared to those obtained from a group of healthy, HIV-negative volunteers. Statistical analysis was performed using the GraphPad program.We studied 68 C-stage HIV-infected patients, 40 male: 28 female, with an average age of 26 and a median of 23. The average plasma 25[OH]D level was 30.94 ng/mL, median 30 ng/mL in HIV-positive patients compared to an average of 44.72 ng/mL and median 39.82 ng/mL in HIV-negative subjects (p=0.0109). 25[OH]D deficiency <20 ng/mL was found in 23.52% HIV-infected patients, while 26.47% HIV-positive subjects had medium levels of 25[OH]D insufficiency, between 20-30 ng/mL. Only 2 patients had severe 25[OH]D deficiency, below 10 ng/mL. We did not find any statistically significant correlations of 25[OH]D levels with CD4+ T-cells level, HIV-RNA plasma viral load or protease inhibitor therapy. Patients with HIV-tuberculosis co-infection had significantly lower levels of 25[OH]D (p=0.0454) than HIV-positive subjects without tuberculosis.25[OH]D plasma levels are lower in HIV-infected patients compared to the general population. Low 25[OH]D levels are reported in patients with HIV-tuberculosis co-infection. Low 25[OH]D levels may indicate unfavorable outcome in HIV-infected patients."}
+{"text": "There are various errors contained in the references 6, 7, 8, 9, 10, 25, 28, 29, 39, 40 and 43. A summary of issues is as follows:* References 6 - 10 contain incorrect information and numbering. They should read as follows:6. (1998) Control and surveillance of African trypanosomiasis. Report of a WHO Expert Committee. World Health Organ Tech Rep Ser 881: I-VI, 1-114.7. Chappuis F, Udayraj N, Stietenroth K, Meussen A, Bovier PA (2005) Eflornithine is safer than melarsoprol for the treatment of second-stage Trypanosoma brucei gambiense human African trypanosomiasis. Clin Infect Dis 41: 748-751.8. Kennedy PG (2008) The continuing problem of human African trypanosomiasis (sleeping sickness). Ann Neurol 64: 116-126.9. Pepin J, Milord F (1994) The treatment of human African trypanosomiasis. Adv Parasitol 33: 1-47.10. Schmid C, Richer M, Bilenge CM, Josenando T, Chappuis F, et al. (2005) Effectiveness of a 10-day melarsoprol schedule for the treatment of late-stage human African trypanosomiasis: confirmation from a multinational study (IMPAMEL II). J Infect Dis 191: 1922-1931.* Reference 25, 28 and 29 should be removed.* Reference 39 is incorrect and should read as follows:Lipinski CA (2004) Lead- and drug-like compounds: the rule-of-five revolution. Drug Discov Today: Technologies 4: 337-341.* Reference 40 is incorrect and should read as follows:Wager T, Verhoest P, Villalobos A (2010) Moving beyond Rules: The Development of a Central Nervous System Multiparameter Optimization (CNS MPO) Approach To Enable Alignment of Druglike Properties. ACS Chem Neurosci 1.* Reference 43 is incorrect and should read as follows:Sykes ML, Avery VM (2009) A luciferase based viability assay for ATP detection in 384-well format for high throughput whole cell screening of Trypanosoma brucei brucei bloodstream form strain 427. Parasit Vectors 2: 54."}
+{"text": "Staphylococcus aureus (MTCC 737), Pseudomonas aeruginosa (MTCC 424), Escherichia coli (MTCC 1687), and yeast-like fungi Candida tropicalis. p-Toluidine on treatment with ammonium thiocynate formed 2-benzothiazolamines (II), which on reaction with hydrazine hydrate formed a hydrazino derivative (III). Compounds GG4 to GG8 were synthesized by reacting the hydrazine derivative with different acetophenones. All the synthesized compounds were identified by IR and 1H-NMR, and antimicrobial activity was performed on the synthesized compounds. Presence of NO2, Br, OCH3, and Cl groups to the substituted benzothiazole enhanced the antibacterial and antifungal activities.In the present study, five new derivatives (GG4 to GG8) of benzothiazoles were synthesized and evaluated against  Since then, significant research has been carried out taking benzothiazole as the basic moiety. From the literature survey, it has been found that extensive work has been reported on 2-substituted benzothiazole derivatives in the past and evaluated for different activities, such as, antibacterial, antiprol1 NMR spectra on Bruker AC 30 of the NMR spectrometer 400 MHz.All the chemicals and solvents used during the experimental studies were of analytical grade and were procured from CDH, New Delhi and Sigma Chemicals, Mumbai. Melting points of all synthesized compounds were determined by using an open capillary tube and were uncorrected. Infrared (IR) data were recorded in KBr disks, on a Perkin Elmer R-IX FTIR spectrophotometer, and the Hp-Toluidine (5.35 g) was dissolved in a mixture of concentrated HCl (4.3 ml) and water (11.6 ml) by heating in a water bath. The contents were cooled and solid ammonium thiocyanate (3.5 g) was added. The mixture was heated on water bath for about 22 hours. The precipitated product was cooled and filtered, washed with water three to four times, and dried. It was recrystallized with aqueous methanol to get cream colored crystals. Yield: 88% (m.p: 130\u00b0C). IR: 3435 (N-Hstr), 2999 (Aliphatic C-Hstr), 1612 (N-Hben), 1462 (Aromatic C=Cstr), 1310 (Aromatic C-Hben) 1H-NMR: 3.35 , 7.24-7.11 , 2.30 .2SO4 was added to p-tolylthiourea (8.3 g) and the temperature of the mixture was raised to 80\u00b0C on a water bath. Next, 48% HBr (0.5 g) acid was added slowly and the reaction mixture was stirred for two hours and set at 80\u00b0C. It was then cooled to room temperature and the reaction mixture was slowly introduced to cold water and then adjusted to pH 9 or 10 by adding ammonia water. The whole mixture was stirred for one hour by heating at 70\u00b0C and then cooled to room temperature. The mixture was extracted twice with dichloromethane and the combined extract was dried with anhydrous sodium sulfate and evaporated, to obtain the title compound. Yield: 80% (m.p: 145\u00b0C). IR: 3395 (N-Hstr), 3261 (N-Hstr), 1462 (Aromatic C=Cstr), 1326 (Aromatic C-Nstr), 1253 (C-Sstr). 1H-NMR: 3.45 , 7.32-7.26 , 2.34 Fifteen milliliters of concentrated Hstr), 3162 (Aromatic C-Hstr), 3000 (Aliphatic C-Hstr), 1611.9 (N-Hben). 1H-NMR: 9.59 , 7.34-7.11 , 3.37 , 2.26 .2-Amino-6-methybenzothiazole (20 g) [0.82 mmol] and hydrazine hydrate (85%) [0.11 mmol] in 50 ml of ethylene glycol were refluxed by stirring for four hours (60\u00b0C). The color of the reaction changed to green and a homogeneous solution appeared. A white solid was precipitated at the end of the reflux period. The mixture was cooled and the product was filtered and then washed with water several times. It was air dried and recrystallized by using ethanol. Yield: 43% (m.p: 192\u00b0C). IR: 3434 (NHNHthin layer chromatography (TLC). On cooling, the solid was separated. It was filtered and washed with little water and recrystallized with absolute ethanol. Yield: 48% (m.p: 181\u00b0C). IR: 3428 (N-Hstr), 3087.8 (Aromatic C-Hstr), 1613.9 (C=Nstr), 823 (Aromatic C-Nstr). 1H-NMR: 8.77 , 7.36-7.06 , 2.38 , 2.69 .2-Hydrazino-5-methylbenzothiazole (1.5 mmol), 3-nitroacetophenone (2.2 mmol), and glacial acetic acid (2\u20133 drops) were taken in absolute ethanol (20 ml) and refluxed on a water bath for eight hours, till different spots appeared, on str), 3164 (Aromatic CHstr), 1612 (C=Nstr), 1581 (NHben), 699 (C-Brstr). 1H-NMR: 9.58 , 7.26-7.12 , 2.50 , 2.27 .2-Hydrazino -5-methylbenzothiazole (1.5 mmol), 4-bromoacetophenone (2.2 mmol), and glacial acetic acid (2\u20133 drops) were taken in absolute ethanol (20 ml) and refluxed on a water bath for eight hours till different spots appeared, on TLC. On cooling, the solid was separated. It was filtered and washed with little water and recrystallized with absolute ethanol. Yield: 52% (m.p: 189\u00b0C). IR: 3434 (NHstr), 3165 (Aromatic C-Hstr), 1612 (C=Nstr), 1581 (N-Hben), 1285 (Aromatic C-Nstr). 1H-NMR: 9.59 , 7.25-7.11 , 6.72 , 2.97 , 2.33 .2-Hydrazino-6-methylbenzothiazole (1.5 mmol), 4-Methoxyacetophenone (2.2 mmol), and glacial acetic acid (2\u20133 drops) were taken in absolute ethanol (20 ml) and refluxed on a water bath for eight hours till different spots appeared, on TLC. On cooling, the solid was separated. It was filtered and washed with little water and recrystallized with absolute ethanol. Yield: 41% (m.p: 169\u00b0C). IR: 3435 (N-Hstr), 3164 (Aromatic C-Hstr), 1612 (C=Nstr), 1582 (N-Hben), 800 (Aromatic C-Clstr). 1H-NMR: 9.59 , 7.25-7.11 , 3.40 , 2.26 .2-Hydrazino-6-methylbenzothiazole (1.5 mmol), 2,4-Dichloroacetophenone (2.2 mmol), and glacial acetic acid (2\u20133 drops) were taken in absolute ethanol (20 ml) and refluxed on a water bath for eight hours till different spots appeared, on TLC. On cooling, the solid was separated, and was filtered and washed with little water and recrystallized with absolute ethanol. Yield: 54% (m.p: 177\u00b0C). IR: 3434 (N-Hstr), 3163 (Aromatic C-Hstr), 1610 (C=Nstr), 1415 (Aromatic C=Cstr). 1H-NMR: 9.58 , 7.38-7.11 , 3.38  2.26 .2-Hydrazino-6-methylbenzothiazole (1.5 mmol), 2,4-Dimethoxyacetophenone (2.2 mmol), and glacial acetic acid (2\u20133 drops) were taken in absolute ethanol (20 ml) and refluxed on a water bath for eight hours, till different spots appeared, on TLC. On cooling, the solid was separated, and was filtered and washed with little water and recrystallized with absolute ethanol. Yield: 58% (m.p: 185\u00b0C). IR: 3433 (O-H and N-Hp-Toluidine on reacting with ammonium thiocyanate formed p-Tolylthiourea (I), which on reaction with hydrobromic acid yielded 2-benzothiazolamines (II). This on reaction with hydrazine hydrate formed hydrazino derivatives (III). The compounds (GG4 to GG8) were synthesized by reacting with hydrazine derivatives, with different acetophenones .The efficient synthetic route for the synthesis of benzothiazole derivatives is shown below . p-ToluiGram positive bacteria \u2014  Staphylococcus aureus (MTCC 737), Gram negative bacteria \u2014 Pseudomonas aeruginosa (MTCC 424), Escherichia coli (MTCC 1687), and yeast-like fungi Candida tropicalis. The concentration of the test compound used was 50 mg/ml. Ampicillin and Clotrimazole were taken as the standard drugs [Tables S. aureus [P. aeruginosa, E. coli [C. tropicalis [In the present study, the efficacy of five new compounds was detected against oli MTCC 687, and sa MTCC 44, Escher. aureus , P. aeru E. coli , C. tropopicalis , and witopicalis , respectS. aureus, E. coli, and C. tropicalis when tested at 50 mg/ml concentration taking ampicillin and clotrimazole as the standard. From the SAR studies, the presence of the electron withdrawing group  in compound GG4 was assumed to be responsible for the observed activity. From the above-mentioned results, it may be concluded that the derivatives of benzothiazoles possess moderate-to-potent antimicrobial activity[Compound GG4 showed significant activity against  activity5 when co"}
+{"text": "MRI is a common method for detecting breast cancer in women at high risk ,2 These BRCA, 65 family history) were managed at the Royal Marsden Hospital from 1994 to 2013. Following ethical approval, data were collected retrospectively for each presentation of breast cancer: method of presentation/diagnosis , age at diagnosis, cancer type, grade, size, presence of DCIS, lymphovascular invasion (LVI), nodal status and tumour subtype. Chi-squared and ANOVA analyses determined any association between the parameters, P < 0.05 was significant.A total of 125 high-risk patients with 134 breast cancers (69 P = 0.008); mean size 17, 29, and 34 mm (P = 0.076) respectively).The majority of cancers were high-grade (68%) invasive ductal carcinomas (78%) without LVI (76%). MRI-detected cancers were triple negative in 60% (P = 0.03), node negative in 100% (P = 0.005) with DCIS in 70% (P = 0.007). Mammography-detected cancers were luminal in 77% (P = 0.03), node negative in 77% (P = 0.005), with DCIS in 81% (P = 0.007). Symptomatic cancers were luminal in 54%, triple negative in 41%, node negative in 56% and DCIS positive in 51%.Ten breast cancers were MRI detected, 43 mammography detected and 81 symptomatic (mean age 41, 51, and 45 years (In this high-risk cohort, MRI detects small, triple-negative, node-negative cancers in younger women, while mammography detects larger, luminal, cancers in older women that may be node positive."}
+{"text": "AbstractZyginella menghaiensissp. n. (Hemiptera: Cicadellidae: Typhlocybinae: Zyginellini), is described from Chinaand a key to species of Zyginella from China is provided.A new species,  Zyginella was established by L\u00f6w in 1885. The genus belongs in the tribe Zyginellini of Typhlocybinae and consists of twenty-two species distributed in the Oriental, Palaearctic and Afrotropical Regions. Members of the genus can be distinguished by the distinct dark spot on the 3rd apical cell of the forewing .Recent taxonomic work on the genus includes L\u00f6whttp://species-id.net/wiki/ZyginellaZyginellaZyginella pulchra L\u00f6w, 1885. L\u00f6w, 1885: 346; PyramidotettixConometopius citri Matsumura, 1907. Synonymized by  Matsumura, 1932: 59; RemmiaRemmia orbigera Vilbaste, 1968. Synonymized by  Vilbaste, 1968: 91;  Forewing  with dis1 markedly distad of point of fusion of Cu1 with M3+4.Head  acutely Male pygofer  with sho Oriental region, Palaearctic region, Afrotropical region.Song & Lisp. n.urn:lsid:zoobank.org:act:090D3B2C-B357-4E11-9F81-AC71C63C6743http://species-id.net/wiki/Zyginella_menghaiensis Head and thorax yellowish brown; vertex with lateral margins with soft red tinge; eyes brownish grey; pronotum brownish with two longitudinal darker stripes; scutellum with basal triangles testaceous. Forewing  reddish Coronal suture  extendinAbdominal apodemes  slender,Pygofer lobe  broad, w Body length males 2.9~3.1 mm.Holotype, male, China: Yunnan Province, Menghai County, 23 July 2008, coll. YUE-HUA SONG. Paratypes: two males, same date as holotype.Zyginella tsauri Chiang, Hsu & Knight (1989), but the forewing has a large dark costal patch ( The new species is similar to al patch  and the al patch . The new species is named for its type locality: Menghai."}
+{"text": "Diffusion tensor magnetic resonance imaging (DT-MRI) enables 3D evaluation of whole heart microstructure. DT invariants evaluate microstructural remodeling by quantifying trace (increases with decreasing cellularity), fractional anisotropy , and tissue mode (decreases with increasing fiber disarray) . We have2, 0.5 x 0.5 x 0.75 mm resolution). Trace, FA, and mode were segmented into epicardial, midwall, and endocardial regions. Bootstrapped histograms with 95% confidence intervals (95%-CIs) of the de-correlated (via decimation by the auto-correlation length) and segmented invariant data were defined to make statistical comparisons of non-Gaussian datasets tractable. Two-group comparisons of median invariant data of each heart were used to test for significant differences (p < 0.05) between HF and CNTL in each transmural region.HF was induced in 10-12 month old New Zealand White female rabbits (N=8) with an epicardial pacing lead placed in the lateral LV wall and tachycardia pacing at 250 beats per minute (bpm) for 3 days, 300 bpm for 3 days, and 350 bpm for 3-4 weeks. Normal weight matched rabbits (N=5) served as controls (CNTL). Hearts were excised, formalin fixed, and DT-MRI was performed on a 7T scanner  (24 diffusion gradient directions, 6 nulls, TE/TR=30/500 ms, b-value=1000 s/mmFigure DT invariant data indentify statistically significant microstructural remodeling in the pacing induced HF model within epicardial, midwall, and endocardial regions.Kung - AHA Grant 12PRE9160024, Chen - NIH Grants P01HL78931, R01HL78932, a Medtronic-Zipes Endowment and the Indiana University-Indiana School of Medicine Strategic Research Initiative."}
+{"text": "Bifidobacteria, Lactococcus and Streptococcus, have been described. However, information for the prophage of Mycobacterium remains poorly defined.Prophages, integral components of many bacterial genomes, play significant roles in cognate host bacteria, such as virulence, toxin biosynthesis and secretion, fitness cost, genomic variations, and evolution. Many prophages and prophage-like elements present in sequenced bacterial genomes, such as In this study, based on the search of the complete genome database from GenBank, the Whole Genome Shotgun (WGS) databases, and some published literatures, thirty-three prophages were described in detail. Eleven of them were full-length prophages, and others were prophage-like elements. Eleven prophages were firstly revealed. They were phiMAV_1, phiMAV_2, phiMmcs_1, phiMmcs_2, phiMkms_1, phiMkms_2, phiBN42_1, phiBN44_1, phiMCAN_1, phiMycsm_1, and phiW7S_1. Their genomes and gene contents were firstly analyzed. Furthermore, comparative genomics analyses among mycobacterioprophages showed that full-length prophage phi172_2 belonged to mycobacteriophage Cluster A and the phiMmcs_1, phiMkms_1, phiBN44_1, and phiMCAN_1 shared high homology and could be classified into one group.Mycobacterium.To our knowledge, this is the first systematic characterization of mycobacterioprophages, their genomic organization and phylogeny. This information will afford more understanding of the biology of The online version of this article (doi:10.1186/1471-2164-15-243) contains supplementary material, which is available to authorized users. Mycobacterium prophages.Phages can be divided into virulent or temperate based on their relationship with the host. Temperate phage inserts and integrates into its host genome upon infection, and can reside as quiescent prophage. Prophage does not infect its host and maintains the dormant state . Whole-gMycobacterium tuberculosis H37Rv genome [M. ulcerans Agy99 genome [M. marinum M and two of them, phiMmar02 and phiMmar08, are full-length prophages [M. abscessus ATCC 19977 chromosome contains a full-length prophage and three prophage-like elements [M. abscessus subsp. bolletii BD genome [M. abscessus Strain 47J26 [M. abscessus M93 is described [M. massiliense Strain M172 contains putative mycobacteriophage [M. canettii STB-I [M. simiae strain DSM 44165 [Mycobacterium prophages remain to be characterized. Knowledge regarding their genomic composition, distribution can facilitate the elucidation of the biology of Mycobacterium.There is huge gap between the number of mycobacteriophages isolated and cognate prophages found within mycobacteria. To date, there are 3427 mycobacteriophages isolated and 448 of them with genome sequenced. They can be assembled into 20 clusters (A-T) and seven of them are singletons , 7. In crophages ; the M. elements ; prophagD genome ; two proin 47J26 ; a potenescribed ; M. massriophage ; a 55-kbii STB-I ; a 40-kbMycobacterium complete genomes sequences from GenBank, shotgun assembly sequences from Whole Genome Shotgun (WGS) databases, and searched for mycobacterioprophages in published literatures. Together, 33 prophages were described in detail, and 11 of them were previously undocumented prophages among Mycobacterium genomes. The genomes, gene contents, comparative genomics studies and the relationships among them were characterized.In this study, we screened all available Mycobacterium genomes. Secondly, the presence or absence of the integrase genes was tested to exclude negative results. Finally, mycobacterioprophage sequences were identified based on the homology between prophage ORFs (open reading frames) and known phage genes. Thirty mycobacterial complete genomes , 11 prophages can be considered as full-length prophage. The remaining 22 prophages were prophage-like elements. The result showed that small prophage-like elements were more prevalent than putative full-length prophages. The small prophage-like elements might be more stable due to mutational decay and loss of some genes somehow involved in genome excision. Small prophage-like elements were more stable and can be more easily detected than the full-length prophages. Through the tRNA search tool, 19 prophages were integrated into tRNA genes  to MAV_0841 (excisionase DNA binding protein), contains sixty-three ORFs  strain [Mycobacterium unique protein. This implies that MAV_0835 and MAV_0790 play a role in the physiology and pathogenicity of M.avium 104.In addition to ORFs similar to other phage genes, two ORFs show unexpected similarity to bacterial key proteins. MAV_0835 encodes type VI secretion protein IcmF (Intracellular Multiplication F), a core component of type VI secretion system in bacteria \u201332. Baserequency , and hasM.avium 104, extends from MAV_1484 (integrase gene) to MAV_1504 (Phage terminase) and contains 21 ORFs , response regulator receiver protein (MAV_1485), DNA primase/polymerase (MAV_1486), Y4cG protein (MAV_1493), transposase (MAV_1498) and phage terminase (MAV_1504). Other phiMAV_2 prophage ORFs similar to known bacterial functional proteins are also identified  to Mmcs_2908 (transglycosylase-like protein) and contains sixteen ORFs , whose predicted protein products are HNH endonuclease and DNA repair protein RadA, respectively. The lysogeny module consists of Mmcs_2921 (putative phage excisionase) and Mmcs_2923 (phage integrase).There are two prophage-like elements in attL and attR sites. Based on Blast-p, only 8 ORFs have sequence similarity to other phage genes at the amino acid sequence level and 4 can be assigned function, namely Mmcs_3802 (HNH endonuclease), Mmcs_3805 (phage major capsid protein), Mmcs_3814 (HNH endonuclease domain-containing protein), and Mmcs_3816 (phiRv1 integrase).The phiMmcs_2 prophage remnant inserts between Mmcs_3803 and Mmcs_3817. The prophage sequence contains 15 ORFs  to BN42_21185  and contains only eight ORFs , BN42_21178 (excisionase), BN42_21179 (DNA primase), BN42_21182 (phage prohead protease), and BN42_21183 (phage major capsid protein).PhiBN42_1, phiBN44_1, and phiMCAN_1 are found in M.canettii CIPT 140060008, flanked by a 22-bp repeat , MCAN_10521 (DNA-binding protein), MCAN_10541 (DNA primase), MCAN_10551 (HNH endonuclease), MCAN_10561 (phage terminase), MCAN_10571 , and MCAN_10601 (phage major capsid protein).Prophage phiMCAN_1 Figure\u00a0, which iM.smegmatis JS623, contains 13 ORFs , Mycsm_04296 (DNA-binding protein), Mycsm_04298 (DNA primase), Mycsm_04299 (HNH endonuclease), Mycsm_04302 (phage terminase), and Mycsm_04303 . Additionally, Mycsm_04293, whose protein product is similar to glycerate kinase, is also present in phiBN44_1.Prophage phiMycsm_1 Figure\u00a0, inserteM.sp. MOTT36Y, extends from W7S_04825 (integrase gene) to W7S_04880  and contains 12 ORFs , W7S_04845 (pantothenate kinase), and W7S_04855 (transposase).Prophage phiW7S_1 Figure\u00a0 integrathttp://phagesdb.org/blast/) and dot plot matrix of the genomes of full-length mycobacterioprophages and mycobacteriophage clusters (A-T and singletons) revealed that phi172_2 shared sequence similarity to cluster A  were firstly used for analyzing bacterial genome to find candidate prophages [DNA sequences of bacteria for analysis were downloaded from multiple databases, such as NCBI . PHAST (rophages . An interophages \u201320. Finarophages .http://lowelab.ucsc.edu/tRNAscan-SE/) [http://mbio-serv2.mbioekol.lu.se/ARAGORN/) [http://blast.ncbi.nlm.nih.gov/Blast.cgi) and the phagesdb.org site (http://phagesdb.org/blast/). Some data about mycobacteriophage genomes was downloaded from the phagesdb.org site (http://phagesdb.org/). DNAman was used to searching the flank of prophage to find attL and attR sites. Sequences were submitted entries to the GenBank sequence database by Sequin (http://www.ncbi.nlm.nih.gov/projects/Sequin/index.html). Comparative genomic analyses of prophage could be carried out by Blast-N for the global comparison of phiMmcs_1 (or phiMkms_1), phiBN44_1, and phiMCAN_1 and Geneious software for the dotplot of all the mycobacterioprophage-like sequences [http://embnet.vital-it.ch/software/ClustalW.html) or MEGA4 [Prophage sequence was annotated using a variety of programs including Glimmer . tRNA ancan-SE/)  and ARAGRAGORN/) . BLAST aequences . Multiplor MEGA4 .Additional file 1: Table S1: Mycobacterial genomes retrieved in this study. (DOC 56 KB)Additional file 2: Table S2: Database matches for phiMAV_1. (DOC 85 KB)Additional file 3: Table S3: Database matches for phiMAV_2. (DOC 44 KB)Additional file 4: Table S4: Database matches for phiMmcs_1. (DOC 40 KB)Additional file 5: Table S5: Database matches for phiMmcs_2. (DOC 38 KB)Additional file 6: Table S6: Database matches for phiMkms_1. (DOC 40 KB)Additional file 7: Table S7: Database matches for phiMkms_2. (DOC 39 KB)Additional file 8: Table S8: Database matches for phiBN42_1. (DOC 30 KB)Additional file 9: Table S9: Database matches for phiBN44_1. (DOC 34 KB)Additional file 10: Table S10: Database matches for phiMCAN_1. (DOC 33 KB)Additional file 11: Table S11: Ddatabase matches for phiMycsm_1. (DOC 34 KB)Additional file 12: Table S12: Database matches for phiW7S_1. (DOC 34 KB)Additional file 13: Figure S1-S11: Comparative genomic analyses of phi172_2 and cluster A (subcluster A1-A11) mycobacteriophage. (DOC 5 MB)Additional file 14: Figure S12: Comparative genomic analyses of phiMAB_1 and subcluster F1 mycobacteriophage. (DOC 549 KB)Additional file 15: Figure S13-S14: Comparative genomic analyses of phiMAB47J26_1, subcluster F1 and cluster N mycobacteriophage. (DOC 1 MB)Additional file 16: Figure S15-S17: Comparative genomic analyses of phiMAB47J26_2, cluster P, subcluster F1 and cluster N mycobacteriophage. (DOC 2 MB)Additional file 17: Figure S18-S19: Comparative genomic analyses of phi172_1, subcluster F1 and cluster N mycobacteriophage. (DOC 1 MB)"}
+{"text": "Shock-related hyperglycemia impairs mitochondrial function and integrity , ultimatn = 9) or imeglimin . Fifteen hours later animals were anesthetized, mechanically ventilated and instrumented for a consecutive 6-hour observation period. After a second imeglimin bolus, colloid fluid resuscitation and continuous i.v. noradrenaline were titrated to maintain normotensive and hyperdynamic hemodynamics. Then 2 mg/g/hour glucose was infused to induce hyperglycemia. Glucose oxidation and gluconeogenesis were derived from blood 13C6-glucose and mixed expiratory 13CO2/12CO2 isotope enrichment during continuous isotope infusion. Liver mitochondrial activity was assessed using high-resolution respirometry [Immediately after cecal ligation and puncture, mice randomly received s.c. vehicle  by increasing whole body glucose oxidation  vs. 51 % of infused isotope, P = 0.085), which coincided with partial restoration of gluconeogenesis  vs. 0.31  mg/g/hour, P = 0.032), liver mitochondrial activity  vs. 116  pmol O2/second/mg tissue, P = 0.003); maximal oxidative capacity  vs. 147  pmol O2/second/mg tissue, P = 0.064). Imeglimin increased liver HO-1, reduced liver Bax expression and attenuated NF-\u03baB activation .Imeglimin decreased blood glucose levels  vs. 192  mg/dl, Imeglimin improved whole body glucose utilization and gluconeogenesis, a well-established marker of liver metabolic capacity ,5, and a"}
+{"text": "Early diagnosis of sepsis and its differentiation from non-infective SIRS is very important. The links between inflammation and coagulation play an important role in the SIRS/sepsis process. We investigated hematological and biochemical parameters (including thromboelastography (TEG)) in patients after surgical resection of esophagus. The aim of our project was to find out whether there are any changes in these parameters that could help in differentiation between SIRS and sepsis.In our study we enrolled 38 patients (aged 41 to 74) undergoing esophagectomy. Blood samples were obtained in the morning before the operation and then every 24 hours for the next 6 postoperative days (POD). Blood samples were analysed for the following parameters: procalcitonin (PCT), C-reactive protein (CRP), IL-6, aspartate transaminase (AST), lactate, white blood count (WBC), D-dimers, antithrombin (AT), international normalised ratio (INR), activated partial thromboplastin time (APTT) and parameters of TEG.P < 0.002) only; POD 2: AST (P < 0.003), lactate (P < 0.006), D-dimers (P < 0.02), PCT (P = 0.03), IL-6 (P < 0.03), WBC (P < 0.03); POD 3: AST (P < 0.03), PCT (P < 0.02), IL-6 (P = 0.006), CRP (P < 0.04), WBC (P < 0.05); POD 4: AST (P = 0.006), PCT (P = 0.007), IL-6 (P < 0.02), CRP (P = 0.03), D-dimers (P < 0.05), INR (P = 0.03); POD 5: PCT (P < 0.003), IL-6 (P < 0.04), CRP (P < 0.04), AT (P = 0.03); and POD 6: PCT (P = 0.0001), CRP (P < 0.013), WBC (P = 0.03), TEG-LY30 (P < 0.04).Nine patients developed sepsis within 6 postoperative days. Five of them had pneumonia and in four patients the cause of sepsis was dehiscention of gastroesophageal anasthosmosis. Significant differences between patients with SIRS and patients with sepsis were found in the following parameters: 0-day (before operation): no significant differences; POD 1: differences in AST (Sequential measurement of biochemical and hemato-logical parameters, mainly AST, PCT, IL-6, WBC, CRP and D-dimers, can help in early diagnosis of sepsis in patients after extensive operation such as esophagectomy. On the contrary, TEG does not seem to be helpful in differentiation of SIRS/sepsis during the early postoperative period. However, it seems to be useful after the fifth postoperative day."}
+{"text": "There was an error in the title. The correct title is: The RNA Polymerase PB2 Subunit of Influenza A/HongKong/156/1997 (H5N1) Restricts the Replication of Reassortant Ribonucleoprotein Complexes. The correct citation is: Nakazono Y, Hara K, Kashiwagi T, Hamada N, Watanabe H (2012) The RNA Polymerase PB2 Subunit of Influenza A/HongKong/156/1997 (H5N1) Restricts the Replication of Reassortant Ribonucleoprotein Complexes. PLoS ONE 7(2): e32634. doi:10.1371/journal.pone.0032634"}
+{"text": "Pituitary adenoma in children is rarely reported. Acromegaly is one of clinical manifestation in GH releasing-pituitary adenoma. Recurrence of clinical manifestation after resection must be evaluated for possibility of pituitary adenoma relapse.Relapse of pituitary adenoma in children must be considered in the recurrence of clinical manifestations.N,male,15-yo, came to pediatric endocrinology outpatient clinic with the main complain of acromegaly and decreased of visual field which was getting worse since two weeks before. He was consulted to ophthalmology and neurosurgery outpatient clinic. MRI with contrast revealed pituitary adenoma. Laboratories results showed TSHS:0.9773(0.35-4.94)uIU/ml, prolactin:0.51(4.04-15.2)ng/ml, testosterone less than 2.50(boys:13-17:28-1110)ng/ml, growth hormone was more than 40,00(>10.0)ng/ml. He was performed transsphenoidal removal cystic tumor. Pathological result showed macroscopic: yellowish cystous mass;0.6x0.4x0.2cm whether microscopic: appropiate to pituitary adenoma, non chromophobe. After surgery, patient was given DDAVP nasal spray 10 microgram/day, L-thyroxin 100 microgram once daily. One year after surgery, patient complaint of acromegaly, decreased visual field, especially in right and left temporal side, cephalgia. On physical examination, body weight was 91.5kg, height was184.5 cm. There was hemianopsia bitemporal. Tanner stage was A2P4G4. MRI with contrast showed pituritary adenoma relapse. Bone age was normal with height percentage based on it is about 96.8%. Tanner Whitehouse showed adult height 186.4cm. Thorax X ray showed heart and lungs were normal. Laboratories results revealed IGF1:1359(237-996)microgram/L, FT4:1(0.89-1.76)ng/dl; TSHS:0.3(0.5-4.94)microIU/ml(12-18yo), testosterone:435.1(28-1110) ng/dl. Working diagnosis was pituitary adenoma relapse post tumor resection, panhypopituitarism, diabetes insipidus. Testosterone 150mg once per month was added."}
+{"text": "Herb combination has been very popular in traditional medical prescriptionssuch as Traditional Chinese Medicine (TCM). Persistent efforts and attempts have beenmade to dissect the action mode of TCM in recent years, which has provided certain evidencefor inter-herbal interactions. However, the interactions among different componentsin a single herb have been largely neglected.Curcuma WenyujinY.H.Chenet C Ling onMDA-MB-231 and MCF-7 breast cancer cell proliferation alone or in combination with afixed-dose-combination was investigated.In this experimental study, the interactions among different componentsof a single herb were explored. The effect of three main sesquiterpenes  isolated from \u0394\u03c8m) changes showed similar results. However, theydemonstrated complicated interactions on the expression of apoptotic-related proteinsand key signal transduction proteins.Furanodiene significantly inhibited cancer cell proliferation while germacrone andcurdione showed no effect. Germacrone enhanced furanodiene\u2019s anti-proliferative effect.Curdione showed no effect on furanodiene\u2019s anti-proliferative effect but partly reversed theanti-proliferative effect of germacrone and furanodiene combined. The morphological andmitochondrial membrane potential (Curcuma WenyujinY.H.Chenet C Ling may exist. The intra-herb interactions should be taken intoconsideration when attempts are made to interpret the art of TCM formulation or other similarrecipes.Unpredictable and complex interactions among different components in  Accumulated data have demonstrated that complementaryand alternative medicine (CAM)shows beneficial effect in the treatment of severalkinds of cancers -3. Tradisalvia miltiorrhiza. Results showed that the combinationyields synergy in the treatment of APL  model by investigatingthe interaction of three pure compoundsnamely tetraarsenictetrasulfide, indirubin, and tanshinoneIIA which were selected to represent threeherbs respectively: realgar, indigo naturalis, andt of APL .Furthert of APL -11. HoweCurcuma WenyujinY.H.Chenet C Ling Ling is a commonlyprescribed Chinese herb with anti-cancer potentials..Curcuma In this experimental study, germacrone, curdione,and furanodiene used in this experimentalstudy were purchased from the National Instisutesfor Food and Drug Control .The RPMI-1640 culture medium was obtainedfrom Gibco (USA). Fetal bovine serum (FBS),phosphate-buffered saline (PBS), penicillinstreptomycin(PS), and 0.25% (w/v) trypsin/1mMEDTA were purchased from Invitrogen (USA).3--2,5-diphenyl tetrazoliumbromide (MTT), 5, 5', 6, 6'- tetrachloro-1,1', 3, 3'-tetraethyl-benzimidazolylcarbocyanineiodide (JC-1), phenylmethanesulfonyl fluoride(PMSF) andprotease inhibitor cocktail were purchasedfrom Molecular Probes (USA). RIPA lysisbuffer was obtained from Santa Cruz (USA).Primary antibodies against Bcl-2, p-Bcl-2, Bclxl,Bax, Bad, Bok, Bim, caspase-9, cleaved caspase-9, PARP, NF-\u03baB, p38MAPK, p42/44MAPK,\u03b2-actin, and secondary antibodies were obtainedfrom Cell Signaling (USA). In this paper, A standsfor germacrone (14.3 \u03bcM), B stands for curdione(42.9 \u03bcM), C stands for furanodiene (42.9 \u03bcM),AB stands for A (14.3 \u03bcM) and B (42.9 \u03bcM) combined while ABC is A (14.3 \u03bcM), B (42.9 \u03bcM)and C (42.9 \u03bcM) combined together and the restfollows the same pattern.2.Human breast cancer cell lines, MCF-7 andMDA-MB-231 were obtained from ATCC (USA).Cells were cultured in medium containing RPMI-1640, antibiotics , and 10 % (v/v) heat-inactivated FBSat 37\u02daC under 5 % CO4) cells in100 \u03bcL medium were seeded in 96-well plates andtreated with A (14.3 \u03bcM), B (42.9 \u03bcM) and C (42.9\u03bcM) alone or in a combination for 24 hours. Thecellular morphology was observed with AxioCamHRC CCD camera (Carl Zeiss).Exponentially growing MCF-7  was monitored by JC-1 staining as previouslydescribed  using SPSS 11.5 software.Statistical differences were considered significantat p<0.05.Firstly, the effect of A, B and C alone on MDAMB-231 and MCF-7 cell proliferation was determined.As shown in figure 2, neither A nor B showedcytotoxic effect on both cell lines at 50 \u03bcM while Cdramatically inhibited both cell line proliferation.No obvious morphologic changes were observed aftertreating with A and B alone or in combination whileC significantly changed the cell morphology and decreasedthe cell number. AC, BC, and ABC inducedobvious morphologic changes . The cel\u0394\u03c8m is shown in figure3 (below), which is quite similar to those of MTTresults. Compared with the control group, no obviouschanges in the red fluorescence were observedin A, B and AB treated group. However, C, AC, BC,and ABC treated groups exhibited increased greenfluorescence. Furthermore, more green fluorescencewas observed in AC group than that of ABCgroup. These results showed that \u0394\u03c8m decreased inC, AC, BC, and ABC groups.The effect of A, B and C alone or in a combinationon MDA-MB-231 cell Both A and B showed no effect on Bcl-xl, Bcl-2and Bim expression, increased Bax andp-Bcl-2, anddecreased Bok expression. B decreased Bad expressionwhile A had no effect. C increased Bcl-2, Baxand Bim expression and decreased Bcl-xl, Bok andBad without affecting p-Bcl-2. AB decreased Bcl-xl,Bok and Bad expression, while increasing Bcl-2, p-Bcl-2, Bax, and Bim expression. AC decreased Bclxl,Bok and Bax expression, increased Bcl-2 andBim without affecting p-Bcl-2 and Bad expression.BC decreased Bok expression, increased Bax and-Bim expression while showing no effect on Bcl-xl,Bcl-2, p-Bcl-2 and Bad expression. ABC decreasedBcl-xl, increased Bax and Bim without affectingBcl-2, p-Bcl-2, Bok, and Bad expression .AC and BC decreased caspase-9 expression while Cand AC increased cleaved caspase-9 expression. It isinteresting to note that B, C, AB, AC, BC ABC couldsignificantly increase PARP expression, especially inAC, BC and ABC groups .A, B and C alone or in a combination showedno effect on NF-\u03baB expression. It is interesting tonote that A, B, C alone increased p42/44 MAPKexpression while in a combination showed no effect.A, C, AB, AC, BC, ABC showed no effecton p38 MAPK expression while B decreasedp38MAPK expression .AIDS), malaria and tuberculosis.Mixtures of compounds produced by plants mayprovide important combination therapies and provideclinical efficacy beyond the reach of single compoundbaseddrugs -negative cell line, while the latter is an ER positivecell line. MDA-MB-231 cells also demonstrateddrug resistant characteristics. Present results suggestthat although both cell lines respond equally to thesecompounds, the underlying mechanisms maybe quitedifferent, which needs further study to elucidate.Drug combination therapy, in which two or moreagents interact with multiple targets simultaneously, isconsidered a rational and efficient form of pharmacotherapydesigned to control complex diseases such ascancer . The fixeddrugs . The appeddrugs . Furthereddrugs , 5.Howeeddrugs -27. Littwenyujin. The MTT\u0394\u03c8m, an early marker for apoptosis. A and B alone or ina combination did not affect \u0394\u03c8m while C significantlydecreased \u0394\u03c8m suggesting that C might induce apoptosis.This was consistent with previous observationsin HL-60 cells (\u0394\u03c8m (\u0394\u03c8m and proliferation.Actually, our study showed that less than 50 \u03bcM Aexhibit no significant cytotoxic effect on breast cancercells (Morphological observations demonstrated similarresults. JC-1 staining is a common method to study the60 cells . Our reclls (\u0394\u03c8m , 18. Theercells . Althougercells , 29, no To further confirm the interactions among A, Band C at the molecular level, some apoptotic-relatedproteins and several key signaling molecularwere determined. Results demonstrated that A, Band C alone showed complex effects on the expressionof Bcl-2 family proteins. For example, Cincreased the anti-apoptotic protein Bcl-2 and thepro-apoptotic protein Bax, Bim at the same time.Also, it decreased Bok but increased the Bim protein,two other pro-apoptotic Bcl-2 proteins ,31. TheCurcuma WenyujinY.H. ChenetC Ling namely germacrone, curdione, and furanodiene.Therefore, intra-herb drug interactions should betaken into consideration when attempts are made tointerpret the art of traditional medicines.Taken together, the present study provides evidencethat there are complicated interactions among thethree components of"}
+{"text": "Due to an error introduced during the production process, the word \"Growth\" was misspelled in the article title. The correct title is: Exogenous Amino Acids Are Essential for Interleukin-7 Induced CD8 T Cell Growth. The correct citation is: Pearson C, Silva A, Seddon B (2012) Exogenous Amino Acids Are Essential for Interleukin-7 Induced CD8 T Cell Growth. PLoS ONE 7(4): e33998. doi:10.1371/journal.pone.0033998"}
+{"text": "Acinetobacter baumannii clinical strains belonging to sequence types ST-208 and ST-218 are reported in this study. They were isolated from tracheobronchial aspirate of mechanically ventilated adult patients admitted to the intensive care unit of a Spanish tertiary hospital during 2010 to 2011.The draft genome sequences of seven multidrug-resistant  Acinetobacter baumannii is among the leading etiologies of hospital-acquired infections, particularly in critically ill patients  isolated from tracheobronchial aspirates. Whole-genome sequencing was performed using the Illumina MiSeq platform with 300 bp paired-end reads, resulting in mean genome coverage of 25.2 \u00b1 5.5-fold (mean \u00b1 SD of the 7 strains). Reads were preprocessed with Trimmomatic v0.32 (http://www.usadellab.org/cms/?page=trimmomatic) (de novo assembled with ABySS assembler v1.5.2 (http://www.bcgsc.ca/platform/bioinfo/software/abyss)  and RAST server v1.0 (http://rast.nmpdr.org) (https://cge.cbs.dtu.dk/services/MLST/) (https://cge.cbs.dtu.dk/services/ResFinder/) (aac(6\u2032)-II, strB, and strA, were detected in pulsotype 1 strains, while pulsotype 2 strains carry aac(3)-Ia, aadA1, strB, and strA genes, and pulsotype 3 strains carry aac(6\u2032)-II, aac(3)-IIa, strB, and strA genes. Resistance genes blaOXA-58, blaOXA-66, and blaADC-25, which confer \u03b2-lactam resistance, were found in strains from pulsotypes 1 and 2; nevertheless, pulsotype 3 strains carry blaOXA-109 and blaADC-25 genes. Referring to sulfonamide resistance, all the strains have sul1 gene, and strains from pulsotypes 1 and 2 also possess sul2 gene. The gene tet(B), related with tetracycline resistance, was found in all the strains. Furthermore, point mutations in genes associated with quinolone resistance were found in all the strains using BLAST  (http://blast.ncbi.nlm.nih.gov/Blast.cgi)  , resultis/MLST/) , 7. MostLIWE00000000 (AbMDR-GLH1), LIZZ00000000 (AbMDR-GLH2), LJAA00000000 (AbMDR-GLH3), LJAB00000000 (AbMDR-GLH4), LJAC00000000 (AbMDR-GLH5), LJAD00000000 (AbMDR-GLH6), and LJAF00000000 (AbMDR-GLH8). The versions described in this report are the first versions, LIWE01000000, LIZZ01000000, LJAA01000000, LJAB01000000, LJAC01000000, LJAD01000000, and LJAF01000000.These whole-genome shotgun (WGS) projects have been deposited at DDBJ/EMBL/GenBank under the accession numbers"}
+{"text": "AbstractVigna subg. Ceratotropis (Piper) Verdc. represents a homogenous and distinct group of species with highly specialized complex floral characters. It is most diverse in Asia. India, with 24 species, represents a secondary center of species diversity of the subgenus.Vignapandeyana RD Gore, SP Gaikwad & SD Randive, is described from hill slopes of the northern Western Ghats of India. It resembles Vignayadavii Gaikwad et al. and Vignadalzelliana (Kuntze) Verdc. but differs from the latter in its dimorphic shoots  and densely hairy pods, from the former by its curved style, flattened style beak, foveolate seed coat and absence of standard protuberance and horn-like keel pocket in cleistogamous flowers.A new species,  Vigna Savi of the tribe Phaseoleae comprises about 90 species distributed in six subgenera (Vignasubg.Ceratotropis (Piper) Verdc. has its center of species diversity in Asia and is popularly known as Asian Vigna  Verdc.  Verdc. . MoreoveVigna on the hill slopes near Chalkewadi in Satara district of the Maharashtra State of India which has unusual dimorphic shoots and cleistogamous flowers. It was recollected in the subsequent two years for further studies of its vegetative and floral characters. Initially, the new species was confused with Vignayadavii Gaikwad et al., as both hold cleistogamous flowers. However, a critical study of floral structures, seeds, seed coat and type of seed germination have revealed that it represents an undescribed species of Vigna subg. Ceratotropis. This has been confirmed by the perusal of relevant literature .Type status:Other material. Location: continent: Asia; country: India; countryCode: IND; stateProvince: Maharashtra; municipality: Nasik District; locality: Kasara-Ghat near Igatpuri; verbatimElevation: 365 m; verbatimLatitude: 19\u00b041'02.1\"N; verbatimLongitude: 73\u00b029'58.3\"E; Event: eventDate: 10-11-2012; fieldNumber: RD Gore 1042a; fieldNotes: Twining herbs; leaves stipulate; stipules submedifixed; chasmogamous flowers yellow & Cleistogamous flowers white; pods falcate to straight; seeds well developed; Record Level: language: English; institutionID: Botanical Survey of India, Pune (BSI).Type status:Other material. Location: continent: Asia; country: India; countryCode: IND; stateProvince: Maharashtra; municipality: Nasik District; locality: Kasara-Ghat near Igatpuri; Identification: identifiedBy: SP Gaikwad & RD Gore; Event: eventDate: 10-11-2012; fieldNumber: SD Randive 322; fieldNotes: Twining herbs; flowers yellow; pods slightly hairy; Record Level: institutionID: Herbarium of Walchand College of Arts & Science, Solapur (WCAS).Type status:Other material. Location: continent: Asia; country: India; countryCode: IND; stateProvince: Maharashtra; municipality: Nasik District; locality: Saptshrungi hills ; Identification: identifiedBy: SP Gaikwad & RD Gore; Event: eventDate: 9-11-2012; fieldNumber: RD Gore 1040; fieldNotes: Twining herbs; flowers both chasmogamous (yellow) and cleistogamous ; Record Level: institutionID: Herbarium of Walchand College of Arts & Science, Solapur (WCAS).Type status:Other material. Location: continent: Asia; country: India; countryCode: IND; stateProvince: Maharashtra; municipality: Satara District; locality: Pasarnighat; Identification: identifiedBy: MM Aitawade; Event: eventDate: 21-10-2011; fieldNumber: SP Sutar 156; fieldNotes: Vignasilvestris but differs in pod & seed number; seeds rectangularClosely resembles with ; Record Level: institutionID: Kew herbarium (K); collectionCode: K000978011; source: http://apps.kew.org/herbcat/getImage.do?imageBarcode=K000978011Type status:Other material. Location: continent: Asia; country: India; countryCode: IND; stateProvince: Maharashtra; municipality: Satara District; locality: Pasarnighat; Event: eventDate: 21-10-2011; fieldNumber: SP Sutar 156; fieldNotes: Vignasilvestris but differs in pod & seed number; seeds rectangularClosely resembles with ; Record Level: institutionID: Herbarium of Shivaji University, Kolhapur (SUK).Type status:Other material. Location: continent: Asia; country: India; countryCode: IND; stateProvince: Maharashtra; municipality: Sangli; locality: Dandoba hills; Event: eventDate: 28-09-1989; fieldNumber: AN Londhe 170037; Record Level: institutionID: Botanical Survey of India, Pune (BSI).Chasmogamous flowers: yellow, 2-4 in axillary and terminal pseudo-racemes; peduncle 5-7.5 cm long, densely hairy with whitish-brown, 1-1.5 mm long, retrosely spreading hairs; pedicels 2-2.5 mm long, densely hairy as peduncle; bracts lanceolate, 2.5-3 x 0.8-1 mm, herbaceous, acute at apex, densely hairy; bracteoles 2, linear, 4-4.5 mm long, densely hairy. Calyx campanulate, ca 2 mm long; teeth triangular, 0.2-1.2 x 0.7-1 mm, sparsely hairy along margins. Standard petal yellow, asymmetrical, broadly ovate, 6-6.5 x 7.5-8.5 mm, emarginate at apex, central protuberance present inside (up to 0.9 mm long); claw ca 1 mm long. Right wing concealing the upper portion of the keel petals; claws 0.7-1 mm long; lamina obliquely obovate, 3.5-6.5 x 2.7-3.5 mm, notched at apex. Left wing claws 1-1.2 mm long; lamina obliquely obovate, 4-6.2 x 1.2-4 mm. Keel petals spirally incurved to left, 6-6.5 x 2.5-3.5 mm; horn-like pocket present (1.5-1.7 mm long) on the left side of keel petal. Stamens 9+1, included; staminal tube 4.5-5 mm long; free filament 8-9 mm long; anthers dorsifixed, 0.2-0.3 mm long. Pistil 1.5-1.8 cm long; ovary linear, 2.2-2.4 mm long, densely covered with long whitish hairs; style filiform, 1.2-1.3 cm long, \u2018S\u2019 shaped before stigma, beaked beyond stigma; beak 0.8-1 mm long, upwardly curved at apex. Pods ascending linear, cylindrical, 2-5 x 0.3-0.5 cm, straight or slightly curved, densely covered with brownish, 2-3 mm long spreading hairs, acute at apex. Seeds 4-10 per pod, broadly rectangular, 3-3.1 x 2-2.1 mm, brown, mottled with faint black patches, rounded or rectangular at both ends; seed coat porous with mesh-like reticulation; hilum well developed, rim-aril protruded, elliptic, 1.8-2 x 0.7-0.9 mm, whitish-brown. Germination epigeal; ecophylls petioles, simple and minutely pulvinous. Cleistogamous flowers: 2-5, whitish-yellow on leafless sub-aerial branches which produce on lower nodes of stem; pedicels 1-2 mm long; bracts lanceolate, 5-6 mm long, acute at apex, densely hairy; bracteoles 2, linear, 7-9 mm long, densely covered with 1-3 mm long bulbous based hairs. Calyx as in chasmogamous flowers. Standard petal symmetrical, broadly ovate, 4.5-5 x 5.5-6 mm, emarginate at apex, inside central protuberance absent; claws 0.3-0.5 mm long. Right wing petal slightly concealing the upper portion of the keel petal; claws 0.3-0.5 mm long; lamina obliquely ovate, 4.5-5 x 2-3 mm; auricle 0.1-0.2 mm long. Left wing petal spreading; claws 0.5-0.7 mm long; lamina ovate-elliptic, 4.9-5 x 2-2.7 mm, base oblique; auricle 0.3-0.5 mm long. Keel petals slightly curved, 4.8-5 x 1.5-2.5 mm. Stamens 9+1, dimorphic, out of 9 stamens in a bundle 4 short and 5 long; short stamens sterile with ca 1 mm long filaments, whereas long stamens fertile with ca 2 mm long filaments; staminal tube ca 1.5 mm long; free stamen sterile with ca 2 mm long filament; anthers dorsifixed 0.2-0.6 mm long. Pistil filiform, up to 6.5 mm long; ovary linear, 1.7-2 mm long, glabrescent or sparsely hairy; style filiform, 3-3.5 mm long, curved (not \u2018S\u2019 shaped), densely hairy before stigma with 0.4-0.6 mm long hairs, beaked beyond stigma; beak short, 0.3-0.5 mm long, straight, pointed at apex. Pods linear, cylindrical, straight, 2.5-4 x 0.3-0.4 cm, narrowed at both ends, sparsely hairy with short hairs (0.5 mm long), white hairs. Seeds 8-9 per pod, obliquely rounded, 3.5-4 x 3-3.5 mm, dark brown; seed coat foveolatewith mesh-like reticulation; hilum well developed, protruded, broadly elliptic, 2-2.2 x 0.4-0.7 mm, whitish. Germination epigeal; ecophylls petioles, simple and minutely pulvinous.  cm long, densely covered with retrorse or spreading bulbous based hairs; stipules elliptically lanceolate, medifixed, 7-9 mm long, 5-7 nerved, rounded at base, acute at apex, hairy on dorsal surface; stipels 2, linear-lanceolate, 2-3 mm long, acute or acuminate at apex, glabrous. Leaflets membranous, entire; lateral ones obliquely ovate or rhomboid to lanceolate, 2.2-5.5 x 1.5-2.3 cm, obliquely rounded at base, acute or shortly acuminate at apex, sparsely hairy; terminal leaflet ovate or rhomboid-lanceolate, 3-6.4 x 1.2-4 cm, base rounded , apex acute or shortly acuminate, sparsely hairy, margins entire or sometimes wavy; rachis 2-10 mm long, densely covered with whitish hairs. Flowers are of two kinds; the chasmogamous flowers present on leafy aerial shoots while cleistogamous flowers present on leafless subterranean shoots, which are close to soil surface.  cm long,August-October.Vignayadavii similis, ramis dimorphis, floribus cleistogamis vexillo sine processo carina sine marsupio corniformi, styli rostro applanato, seminum testa foveolata, hilo bene evoluto differt.The specific epithet honors Prof AK Pandey, Department of Botany, Delhi University, New Delhi (India), in recognition of his valuable contribution to the taxonomy of flowering plants of India.India, Maharashtra, Satara district, near Chalkewadi in Patan tahsil.Carviacallosa (Nees) Bremek., Crotalarianana Burm. f., Crotalariavestita Baker, Cajanuslineatus (Wight & Arn.) Maesen, Eragrostis spp., Pseudarthria spp., Nogradalzellii (Baker) Merr. and Themeda spp.It is a twining annual herb, grows on lateritic gravelly soil on hill slopes amonggrasses and herbs at about 1200 m altitude above mean sea level in Satara district of Maharashtra, India. The species has adapted to the monsoon seasonality. It thrives in humid climate with heavy rainfall during growth season. The seed germination takes place with onset of monsoon rain in the first week of June and the plant completes its life cycle with formation of seeds when rains ceasein mid October. The common associates of the species are Vigna sect. Ceratotropis while cleistogamous floral parts correspond to the species of V. sect. Aconitifoliae of the same subgenus. Thus, it shows a combination of characters of species of both sections. The new species shows morphological similarities with Vignayadavii Gaikwad et al. and Vignadalzelliana (Kuntze) Verdc. but differs in its dimorphic branches, foveolate seed coat and absence of standard protuberance and horn-like keel pocket in cleistogamous flowers. A comparative account between above mentioned three species is given in Table Interestingly, two types of shoots are observed in the species, one a normal aerial leafy shoot and the other subterranean (close to soil surface) leafless shoot producedatthe lower nodes of the stem. The later shoots produce cleistogamous flowers, which remain closed. They show differences in the structure of their floral parts as compared to chasmogamous flowers such as standard petal without central protuberance inside, keel petals without horn-like pocket, curved style and short style beak 0.3-0.5 mm long). Figs , 5, 6\u200b.  mm long."}
+{"text": "The publisher apologizes for the error. The correct citation is: Mart\u00ednez de Paz A, Sanchez-Mut JV, Samitier-Mart\u00ed M, Petazzi P, S\u00e1ez M, Szczesna K, et al. (2015) Circadian Cycle-Dependent MeCP2 and Brain Chromatin Changes. PLoS ONE 10(4): e0123693. doi:Please download this article again to view the correct version. The originally published, uncorrected article and the republished, corrected article are provided here for reference.S1 File(PDF)Click here for additional data file.S2 File(PDF)Click here for additional data file."}
+{"text": "Previous meta-analyses regarding the performance of interferon-gamma release assays (IGRAs) for tuberculosis diagnosis in children yielded contrasting results, probably due to different inclusion/exclusion criteria.vs. high income countries); including only microbiological confirmed TB cases; including only studies performing a simultaneous three-way comparison of the three tests, and including immunocompromised children.We systematically searched PubMed, EMBASE and Cochrane databases and calculated pooled estimates of sensitivities and specificities of QuantiFERON-TB Gold In Tube (QFT-G-IT), T-SPOT.TB, and tuberculin skin test (TST). Several sub-analysis were performed: stratification by background (low income vs. 92%) but not in low income countries (85-93% vs. 90%).Overall, 31 studies (6183 children) for QFT-G-IT, 14 studies (2518 children) for T-SPOT.TB and 34 studies (6439 children) for TST were included in the analyses. In high income countries QFT-G-IT sensitivity was 0.79 (95%IC: 0.75-0.82) considering all the studies, 0.78 (95%CI:0.70-0.84) including only studies performing a simultaneous three-way comparison and 0.86 (95%IC 0.81-0.90) considering only microbiologically confirmed studies. In the same analyses T-SPOT.TB sensitivity was 0.67 (95%IC 0.62-0.73); 0.76 (95%CI: 0.68 to 0.83); and 0.79 (95%IC 0.69-0.87), respectively. In low income countries QFT-G-IT pooled sensitivity was significantly lower: 0.57 (95%IC:0.52-0.61), considering all the studies, and 0.66 (95%IC 0.55-0.76) considering only microbiologically confirmed cases; while T-SPOT.TB sensitivity was 0.61 (95%IC 0.57-0.65) overall, but reached 0.80 (95%IC 0.73-0.86) in microbiologically confirmed cases. In microbiologically confirmed cases TST sensitivity was similar: 0.86 (95%IC 0.79-0.91) in high income countries, and 0.74 (95%IC 0.68-0.80) in low income countries. Higher IGRAs specificity with respect to TST was observed in high income countries (97-98% Both IGRAs showed no better performance than TST in low income countries. Diagnosis of paediatric tuberculosis infection remains a challenging issue. Tuberculin skin test (TST) has several limitations: sensitivity may be influenced by the child\u2019s age and immunologic status, bacille Calmette-Gu\u00e9rin (BCG) vaccination or non-tuberculosis mycobacterium-infections; in case of repeated tests a booster effect can occur and a double access to a health care facility is needed. Nevertheless, infiltrate measurement may be operator-dependent -4. InterMycobacterium tuberculosis antigens, which are absent in BCG and many non-tuberculosis mycobacteria, and displayed similar sensitivity and higher specificity than TST in adults  and summarized the results in forest plots. Random-effects meta-analysis was performed using MetaDiSc\u00ae, Meta-analysis of Diagnostic and Screening tests, Version 1.4 . StudiesFor the analysis of sensitivity, 31 studies (20 conducted in high income countries and 11 in low income countries) for QFT-G-IT, including 6183 children -44, 14 sThe specificity analysis included 17 studies, overall including 3844 children (11 conducted in high income countries and 6 in low income countries) using QFT-G-IT ,34,37,43Considering the whole study population, significantly higher sensitivities of QFT-G-IT and TST in high- than in low-income countries were observed, with no difference between them, while T-SPOT.TB sensitivity was particularly low in both settings   in high income countries, and 0.57 (95%IC 0.52-0.61) in low income countries, and TST pooled sensitivity was 0.78 (95%IC 0.74-0.82) in high income countries, and 0.67 (95%IC 0.64-0.70) in low income countries.Specificity of the three test was similar in low income countries while both QFT-G-IT and T-SPOT.TB displayed higher specificity than TST in high income countries. In particular, QFT-G-IT specificity was 0.97 (95%IC 0.96-0.98) in high income countries, and 0.85 (95%IC 0.82-0.88) in low income countries. T-SPOT.TB pooled specificity was 0.98 (95%IC 0.96-0.99) in high income countries, and 0.93 (95%IC 0.87-0.96) in low income countries. TST specificity was 0.92 (95%IC 0.89-0.93) in high income countries, and 0.90 (95%IC 0.87-0.92) in low income countries.Data regarding microbiologically confirmed cases were available in 16 studies (11 studies conducted in high income countries and 5 studies conducted in low income countries), overall including 3689 children for QFT-G-T ,37,40,44.In this analysis sensitivity of the three test was similar (0.81 (95%CI: 0.76-0.85) for QFT-G-IT, 0.80 (95%CI: 0.74-0.84) for T-SPOT.TB, and 0.79 (95%CI: 0.75-0.83) for TSTA sub-analysis in studies conducted in high income and low income countries was performed. In high income countries sensitivity of the three test was confirmed to be similar but IGRAs performance in low income countries was suboptimal. QFT-G-IT pooled sensitivity was 0.86 (95%IC: 0.81-0.90) in high income countries but only 0.66 (95%IC: 0.55-0.76) in low income countries. Even when excluding studies including HIV-infected children, QFT-G-IT pooled sensitivity in low income countries reached only 0.68 (95%IC: 0.55-0.76).T-SPOT.TB pooled sensitivity was 0.79 (95%IC: 0.69-0.87) in high income countries, and 0.80 (95%IC 0.73-0.86) in low income countries. TST sensitivity was 0.86 (95%IC 0.79-0.91) in high income countries, and 0.74 (95%IC 0.68-0.80) in low income countries table .Six of the available studies, including 618 children, evaluated all three tests simultaneously in the same children ,23,37,39Overall, 11 studies have assessed utility of IGRAs in paediatric populations including HIV infected children ,50,52-58Complete data for a specific subgroup-analysis were available in four studies ,34,43,50The meta-analytic estimate for sensitivity was very low and similar for QFT-G-IT and T.SPOT.TB: 0.47 (95%CI: 0.38-0.55) for QFT-G-IT and 0.54 (95%CI:0.49-0.59) for TST. The meta-analytic estimate for specificity was not performed since only data from 2 studies were available ,43. MetaSix studies, including 1733 children, included exclusively children aged \u2264 5 years ,42,59,60Debord and colleagues evaluated QFT-G-IT performance restrospectively in 19 French immunocompetent children (median age: 1.52 years) with active tuberculosis . The ratet al. \u2018s study [Although, in general, results in young children were encouraging, a specific meta-analysis for this subjects, could not be performed as complete data were not available in most studies, except for Detjen \u2018s study .vs. 70%). Other authors [vs. microbiologically confirmed TB cases  [Data on IGRAs\u2019 performance in children are accumulating. In previous meta-analyses, similarly to data reported in adults, higher IGRA specificity with respect to TST has been reported. However, the reported IGRA sensitivity ranged between 62% and 89% for T-SPOT.TB and 66% and 83% for QFT-G-IT ,5,9-11.  authors  performefor TST) .Our study is the first paediatric meta-analysis evaluating both QFT-G-IT and T-SPOT.TB performance by setting. Moreover, we were first to present a sub-analysis of studies performing a simultaneous three way comparison using all the tests in the same child, allowing to reduce potential bias due to individual differences.vs. 75%), where T-SPOT.TB seems to have lower sensitivity than the two other tests (67%). However this result was not confirmed including only ascertained cases, with a microbiological confirmation. In this sub-analysis T-SPOT.TB sensitivity reached 80% (95%CI: 59-90) while QFT-G-IT sensitivity decreased, but not significantly, to 66% (95%CI:55-76). This finding suggests caution when interpreting results from studies including probable and ascertained TB cases in children, for possible misdiagnoses.At a first glance, our meta-analytic results showed a higher sensitivity of QFT-G-IT than TST in high income countries (79% vs. 84%).In a further sub-analysis including only studies performing simultaneously the 3 tests, all performed in high income countries, overall including 618 children, no different sensitivity of both IGRAs and TST was observed, while a higher IGRAs specifity was confirmed  in children.Click here for file"}
+{"text": "This is a report of the complete genome sequences of plaque-selected isolates of each of the four virus strains included in a South African commercial tetravalent African horse sickness attenuated live virus vaccine. Orbivirus, family Reoviridae). The AHSV genome consists of 10 segments of double-stranded RNA (dsRNA) collectively encoding seven structural proteins and four nonstructural proteins (African horse sickness (AHS) is an arthropod-borne equine disease caused by African horse sickness virus (AHSV)  and serotype 6 (HS02/75) viruses, which are precursors of the respective AHS-ALV virus strains, are available from GenBank . The pairwise nucleotide sequence identity of AHSV-2/Labstr/ZAF/1998/OBP-252.1 and HS82/61 was 99.536%, while that of AHSV-6/Labstr/ZAF/1998/OBP-252.1 and HS02/75 was 99.350%.P = 7.56 \u00d7 10-156) composed of eight segments likely derived from AHSV7-tVP2 , and the other segments likely derived from HS39/63 (accession numbers KF860011 and KF860013).The genome sequences of the AHSV serotype 7 strain with a truncated VP2 protein (AHSV7-tVP2)  from whiP = 2.71 \u00d7 10-282), and the segments encoding VP4 and NS3 likely derived from an AHSV for which whole-genome sequence data are currently not available .The genome sequences of the AHSV serotype 8 (HS10/62) from which AHSV-8/Labstr/ZAF/1998/OBP-252.1 was derived are available from GenBank (accession numbers KF860026 to KF860035). Pairwise nucleotide sequence identity between AHSV-8/Labstr/ZAF/1998/OBP-252.1 and HS10/62 over four of the 10 segments was >99.9%, while that for the remaining segments was between 76.8% and 97.4%. Analysis using RDP4.56 (KT715601 to KT715610, KT715611 to KT715620, KT715621 to KT715630, and KT715631 to KT715640, respectively.The AHSV-2/Labstr/ZAF/1998/OBP-252.1, AHSV-6/Labstr/ZAF/1998/OBP-252.1, AHSV-7/Labstr/ZAF/1998/OBP-252.1, and AHSV-8/Labstr/ZAF/1998/OBP-252.1 sequences have been deposited in GenBank under accession numbers"}
+{"text": "The BSC infections increase mortality, complications, hospital stay and the costs. In our setting, BSC is one of the device-associated infections more common. The Alliance for Patient Safety, WHO, launched in 2004, the campaign for the prevention of infection associated to medical care though hand hygiene. The Working Group on Infectious Diseases of the Spanish Society of Intensive Care Medicine and Coronary Units (GTEI-SEMICYUC), developed the National Survey of Nosocomial Infection Surveillance (ENVIN) as a computerized record of the incidence of nosocomial infection for services or intensive care units (ICU). Were selected for monitoring those most serious and frequent nosocomial infections related to instrumentation, including ventilator-associated pneumonia , urinary tract infection associated with urethral catheterization and BSCWe want to show the evolution of BSC: central venous catheter (CVC) and arterial (CA) in our 18-bed unit, after the implementation of BZ program and compare it with spanish data.- Data from ENVIN 01.04.11 / 01.04.2015. 18 ICU beds.Rates: BSC/100 admitted, BSC/CVC-CA. Incidence density (ID): BSC/1000ds,BSC/1000 days CVC-CAN 4198 (patients admitted to ICU), 1659 patients with CVC-CA, 17575 days of stay(ds),11024 days of CVC, CA.- Training and educational campaign were imparted to professionals of our unit andAnesthesiology service (235 people). Some classes were presencial and another were by means of internet. Implementation of protocol for hand washing, technical skills about the insertion of CVC, cleaning and maintenance of catheters.- The main objective of BZ project was to reduce BRC DI < 4 episodes per 1000 CVCdays01.04.11-01.04.12: 3 BSC 0.29/100 admitted, 0.80/CVC-CA, 0.62/1000ds, 0.96/1000days CVC-CA. Germ A. Baumanii 33.33%, P.mirabilis 33.33%, S. Epidermidis33.33%. Sepsis 66.67%01.04.12-01.04.13: 7 BSC. 0.65/100 admitted, 1.61/CVC-CA, 1.42/1000ds, 2.1/1000days CVC-CA. Germ S. Coagulasa negativo (SCN). Severe sepsis 28.57%01.04.13-01.04.14: 5 BSC 0.44/100 admitted, 1.08/CVC-CA, 1.02/1000ds, 1.63/1000days CVC-CA. Germ S. Epidermidis. Sepsis 60%01.04.14-01.04.15: 3 BSC 0.31/100 admitted, 0.78/100 CVC-CA, 0.71/1000 ds,1.07/1000 days CVC-CA. Germ: S. epidermidis. Sepsis 100%.Spain, last year: N = 20799, 1.59/100 admitted, 2.51/100 pat CVC-CA, 2.10/1000 ds,1.69/ 1000 days CVC-CA. Germ S. Epidermidis. Sepsis 64.85%We are below the level of BSC set by the SEMICYUC with fewest BSC than the rest of the country through the implementation of the program BZ. In the last year, we have improved our numbers because the realization of program recommendations has been monitored more intensively since in the previous two years had been a relaxation and thereby increased the numbers of BSC.UCI San Cecilio"}
+{"text": "The correct name is: David J. Herren. The correct citation is: Herren DJ, Norman JB, Anderson R, Tremblay ML, Huby A-C, Belin de Chantem\u00e8le EJ (2015) Deletion of Protein Tyrosine Phosphatase 1B (PTP1B) Enhances Endothelial Cyclooxygenase 2 Expression and Protects Mice from Type 1 Diabetes-Induced Endothelial Dysfunction. PLoS ONE 10(5): e0126866. doi:The publisher apologizes for this error, which was introduced during the typesetting process."}
+{"text": "A 66-year-old male patient was admitted to the hospital with backache, fatigue, and paraesthesia and spasm in both legs. He had lower extremity numbness and bladder and bowel incontinence. Physical examination revealed the absence of bilateral lower extremity reflexes and lower extremity weakness. Magnetic resonance imaging showed a large mass extending from T8 to T9, fracture of T9, and compression of the spinal cord. Informed consent was obtained.Laboratory results at initial evaluation revealed the following: haemoglobin: 118 g/L, haematocrit: 33.5%, white blood cell count: 6.7x109/L, platelets: 192x109/L, blood urea nitrogen: 7.5 mmol/L, creatinine: 113.1 \u00b5mol/L, calcium: 1.9 mmol/L, total protein: 63 g/L; albumin: 31 g/L; and erythrocyte sedimentation rate: 59 mm/h. Protein studies by nephelometry revealed IgA of 4.25 g/L (reference range: 0.7-4 g/L) and lambda light chain of 3.98 g/L (reference range: 0.9-2.1 g/L). A small monoclonal spike was present upon protein electrophoresis. Urine immunoelectrophoresis documented no monoclonal light chain. Bone marrow aspirate and biopsy were performed and the patient underwent surgical decompression and stabilisation of the thoracic spine. The bone marrow aspirate and biopsy morphology showed infiltration with atypical, multilobated, cleaved, and monocytoid nuclei plasma cells . The biopsy material stained positive with lambda light chain and CD138."}
+{"text": "Microcystis aeruginosa is a typical algal bloom-forming cyanobacterium. This report describes the whole-genome sequence of a non-microcystin-producing strain of Microcystis aeruginosa, NIES-44, which was isolated from a Japanese lake. Microcystis aeruginosa is a typical algal bloom-forming cyanobacteria, one of several cyanobacteria known to produce the potent hepatotoxin microcystin  and combined into a hybrid assembly with the 454 reads using GS de novo assembler version 2.8 (454 Life Sciences). Gaps between the resultant 375 contigs were closed using NESONI version 0.118 (http://www.vicbioinformatics.com/software.nesoni.shtml) and Platanus version 1.2.1 (http://platanus.bio.titech.ac.jp/platanus-assembler). The draft genome was annotated using the RAST server (http://rast.nmpdr.org/rast.cgi), which predicted protein-coding sequences (CDSs). Whole-genome homology mapping of various strains, including M.\u00a0aeruginosa NIES-44, was performed using Gegenees version 2.2.1  with a paired-end library (400\u00a0bp) and a Roche/454 PE genome sequencer FLX  with a mate-paired library (8\u00a0kb). HiSeq reads were assembled M.\u00a0aeruginosa NIES-44 genome comprised 80 contigs (six scaffolds) and had a total length of 4,565,330\u00a0bp and a G+C content of 43.19%. It included 4,790 protein-coding sequences and 47 RNA-coding genes . The annotation revealed that 2,614 CDSs exhibited homology to genes with known functions, and the remaining 2,176 genes were identified as encoding hypothetical proteins of unknown function.The M.\u00a0aeruginosa NIES-44 were 99.73% homologous  to those of M. aeruginosa strain NIES-843 (mcy) and nonribosomal peptide synthetase gene clusters. Since M.\u00a0aeruginosa NIES-44 showed 64.69% homology to M.\u00a0aeruginosa TAIHU98, in which mcy gene cluster deficits have been previously reported (M.\u00a0aeruginosa NIES-843 (M.\u00a0aeruginosa NIES-44 has a characteristic gene structure.The 16S rRNA sequences of BBPA00000000 and refers to the first version that is described in this paper.This whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession number"}
+{"text": "High rates of albuminuria are observed among HIV-infected individuals on stable antiretroviral therapy (ART). Though pro-inflammatory and pro-fibrotic responses are described as components of albuminuria in the general population, it is unclear how these responses are associated to albuminuria in ART-treated chronic HIV. We investigated the relationship of monocyte subsets and urine inflammatory and fibrotic biomarkers to albuminuria in ART-treated HIV-infected participants.Cross-sectional analyses were performed on Hawaii Aging with HIV-cardiovascular disease study cohort participants who were required at entry to be \u226540 years old and on ART \u22653 months. Monocyte subpopulations were determined in banked peripheral blood mononuclear cells (PBMC) using multi-parametric flow-cytometry. Entry random urine samples were assessed for albumin-to-creatinine ratios (UACR). Urine samples were measured for inflammatory and fibrotic biomarkers using Luminex technology.low/+CD16++) monocytes were significantly elevated in subjects with albuminuria (p = 0.034) and were correlated to UACR . Elevated non-classical monocyte counts were significant predictors of worsening albuminuria, independent of traditional- and ART-associated risk factors . Urine TGF-\u03b21 and collagen-IV were significantly higher in albuminuric compared to non-albuminuric participants . Urine TGF-\u03b21 was significantly correlated with non-classical monocyte counts .Among 96 HIV-infected subjects with measured UACR , 18 patients (19%) had albuminuria. Non-classical (CD14Alterations in monocyte subpopulations and urine pro-fibrotic factors may play a role in kidney dysfunction during chronic HIV infection and warrants further study. Albuminuria is a strong and independent risk factor for renal and cardiovascular disease among the general and HIV-infected populations \u20137. There++CD16-), intermediate (CD14++CD16+), and non-classical (CD14+/lowCD16++) subsets  assessed to have moderately increased albuminuria (UACR 30-300mg/g) with only 2 (11%) having severe albuminuria (UACR >300mg/g). Demographics and clinical parameters are summarized in p <0.05), but not with CD4 T cells (r = 0.070), CD8 T cells (r = 0.150), activated CD8 T cells (r = -0.039), or with classical (r = 0.112), intermediate (r = 0.067), or transitional monocytes (r = 0.127).T cell and monocyte subset percentages and counts were compared between participants with and without albuminuria. Non-classical monocyte counts were significantly elevated in HIV-infected participants with albuminuria as compared to HIV-infected participants without albuminuria . No signp = 0.035, C.I. = 0.001\u20130.033; Hypertension, B = 0.463, p<0.001, C.I. = 0.235\u20130.691; HOMA-IR, B = 0.077, p = 0.010, C.I. = 0.019\u20130.136; Total cholesterol/HDL cholesterol ratio, B = 0.067, p = 0.046, C.I. = 0.001\u20130.132; Current use of Tenofovir, B = 0.215, p = 0.131, C.I. = -0.066\u20130.496; Current use of Ritonavir, B = -0.027, p = 0.827, C.I. = -0.274\u20130.220. We conclude that elevation of non-classical (CD14low/+CD16++) monocytes significantly predict worsening albuminuria in HIV-infected participants on stable ART, independent of traditional- and ART-associated risk factors.We assessed the predictive value of non-classical monocytes in a multivariable linear regression model, adjusting for traditional risk factors of age, hypertension, HOMA-IR, total cholesterol/HDL cholesterol ratio, and ART-associated risk factors of current use of Tenofovir and/or Ritonavir . Univari1 and collagen IV , while IP-10 (r = 0.008), MCP-1 (r = -0.141), IL-18 (r = -0.037), Cystatin C (r = -0.129), TGF-\u03b22 (r = 0.063), TGF-\u03b23 (r = -0.189), collagen IV (r = 0.032), and TIMP-1 (r = 0.029) did not show significant correlations. No correlations were seen between other monocyte subsets or T cell subpopulations and urine inflammatory or fibrotic biomarkers. We also assessed the correlations among the urine inflammatory and fibrotic biomarkers. TGF-\u03b21 strongly correlated with TGF-\u03b22 (r = 0.752) and TIMP-1 (r = 0.424). Furthermore, TGF-\u03b22 correlated with TGF-\u03b23 (r = 0.389), collagen IV (r = 0.617), and TIMP-1 (r = 0.730). Of the urine pro-inflammatory biomarkers, only urine MCP-1 correlated with TGF-\u03b22 (r = 0.520), TGF-\u03b23 (r = 0.362), collagen IV (r = 0.558), and TIMP-1 (r = 0.592).We assessed the correlation between the measured urine inflammatory and fibrotic biomarkers with non-classical monocytes. Urine TGF-\u03b2low/+CD16++) monocytes, independent of traditional and ART-associated risk factors. Treated HIV-infected individuals with albuminuria excrete higher levels of urine TGF-\u03b21 and collagen IV as compared to those without albuminuria, and increased non-classical monocytes are associated with increased urine TGF-\u03b21. Furthermore, urinary levels of TGF-\u03b21 are strongly associated with other urine fibrotic markers. It is important to note that although only 89% of the participants were virally suppressed, when participants with viral loads were excluded, all significant observations stated remained significant.Our study is the first report showing that albuminuria in HIV-infected individuals on stable ART is associated with increased levels of non-classical  and intermediate (CD14++CD16+) subsets, are generally characterized as a pro-inflammatory cellular compartment, being potent producers of TNF-\u03b1, IL-6, IL-1\u03b2 and IL-12 [+ monocytes in human inflammatory disease states may mediate further pro-inflammatory responses. Individuals with rheumatoid arthritis have been shown to have significantly higher CD16+ monocytes as compared to healthy controls [CD16nd IL-12 \u201351. Sevecontrols , 53. Elecontrols , 54. NonElevated levels of peripheral non-classical monocytes have also been described in HIV-infected individuals as compared to HIV-uninfected controls , 55, 56.Contrasting studies in the general population have shown non-classical monocytes in humans to play a role in the resolution of inflammation and the differentiation into M2 macrophages that aide in anti-inflammatory and wound healing responses . In myocDuring the resolution of inflammation, an important component of the response is TGF-\u03b2. TGF-\u03b2 is a multi-functional cytokine that regulates many cellular functions, including cellular growth and differentiation . In the This study is limited by its relatively small sample size and the lack of HIV-uninfected controls with measured UACR and phenotyped immunologic cellular subpopulations. However, the strengths of the study are the careful clinical and cardio-metabolic characterizations performed on HIV-infected groups, as well as detailed phenotypes of T cell and monocyte subpopulations and quantification of urine biomarkers that reveal discriminating associations in the HIV-infected group.low/+CD16++) monocytes is associated with worsening albuminuria in HIV-infected individuals on ART. This association is independent of traditional- and ART-associated risk factors of albuminuria. Furthermore, HIV-infected individuals with albuminuria excrete higher levels of urine TGF-\u03b21 and collagen IV as compared to those without albuminuria, and increased non-classical monocytes are associated with increased urine TGF-\u03b21. The role of non-classical monocytes in pro-inflammatory and/or pro-fibrotic responses in the kidney of HIV-infected individuals on ART warrants further study.In conclusion, elevation of non-classical (CD14"}
+{"text": "In mice, cryo plus checkpoint blockade facilitates tumor antigen release, T-cell priming, and improved survival . Here, wWomen with ESBC were treated 7-10 days preceding mastectomy with either cryo n=6), single-dose ipi at 10mg/kg (n=6), or cryo+ipi (n=6) , single- and intrCryo+ipi generated greater increases in peripheral Ki67+CD4+ (p=0.05), Ki67+CD8+ (p=0.05), ICOShiCD4+ (p=0.005), and ICOShiCD8+ (p=0.005) cells. The intratumoral T-effector/regulatory ratio was higher following cryo+ipi, but only when Ki67-gated (p=.01). Cryo+ipi generated greater increases in IL-2 (p=.01), IFN\u03b3 (p=.06), and IL-5 (p=.09). Despite negligible intratumoral changes by immunohistochemistry, cryo+ipi generated more high-magnitude (~1000 amplicon) clonal expansions by TCR sequencing (medians: 52 v. 3 clones).Cryo+ipi is associated with potentially favorable immunologic effects. Ki67-gating and TCR sequencing may identify intratumoral changes otherwise undetectable by flow or IHC."}
+{"text": "Desulfovibrio carbinoliphilus subsp. oakridgensis FW-101-2B is an anaerobic, organic acid/alcohol-oxidizing, sulfate-reducing \u03b4-proteobacterium. FW-101-2B was isolated from contaminated groundwater at The Field Research Center at Oak Ridge National Lab after in situ stimulation for heavy metal-reducing conditions. The genome will help elucidate the metabolic potential of sulfate-reducing bacteria during uranium reduction. Desulfovibrio carbinoliphilus subsp. oakridgensis FW-101-2B was isolated from groundwater of well FW-101 at The Field Research Center (FRC) at Oak Ridge National Lab (ORNL). The FRC is part of the Y-12 security complex, located in the Bear Creek drainage. ICP-MS analysis of FW-101 groundwater in late 2001 estimated uranium concentrations between 20 and 250\u00a0ppm, and chromium concentrations between 35 and 85\u00a0ppm. Previous studies have demonstrated reduction of nitrate and uranium levels in the subsurface upon bio-stimulation with ethanol at the FRC (1the FRC 13. Duringthe FRC 1.http://www.phrap.com) was used for genome finishing. Genes were identified using Prodigal (The genome was sequenced by 454\u00a0GS FLX Titanium and paired-end Illumina GAii (2\u00a0\u00d7\u00a035\u00a0bp). The pyrosequencing and Illumina reads were assembled using the Newbler (Roche) and Velvet  and Desulfovibrio vulgaris (63.3% G+C). The COG predictions categorize 581 of the 3,737 protein-encoding genes as pertaining to information storage and processing, 1,183 as cellular processes, 1,576 as metabolism genes, and 596 as poorly characterized functions. Sequencing detected 2 plasmids of different sizes and G+C content, pFW10101  and pFW10102 .Sequence determination revealed a 4.1-Mb genome with 66.5% G+C content, which is comparable to D. carbinoliphilus D41T. The ANIb values calculated by JSpecies (D. vulgaris Miyazaki (69.11%), D. vulgaris Hildenborough (67.45%), and D. vulgaris DP4 (67.41%) than Syntrophobacter fumaroxidans (64.68) and Desulfovibrio desulfuricans ATCC 27774 (60.93%). The small-subunit (SSU) rRNA gene and sulfite reductase gene (dsrAB) of FW-101-2B was most similar to D.\u00a0carbinoliphilus D41T at 99% and 92% similarity, respectively.FW-101-2B was most closely related to JSpecies  showed tD. carbinoliphilus D41T, physiological evidence would support classification of a new strain. Thus, we propose the classification as D. carbinoliphilus subsp. oakridgensis. This is the first genome sequence of a D. carbinoliphilus strain.Although FW-101-2B is phylogenetically very similar to ADFE00000000. The version described in this paper is the version ADFE00000000.2.The whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession no."}
+{"text": "Escherichia coli is an increasing problem especially in developing countries.Antimicrobial resistance among pathogenic E. coli collected from two unrelated geographical areas.To compare between resistance patterns of E. coli (n= 402) collected from hospitals in Khartoum state, Sudan and in Aseer region, Saudi Arabia were studied. Identification and antimicrobial susceptibility testing of isolates were performed following standard methods. Multi-drug resistance (MDR) was defined as non-susceptibility to \u2265 three antimicrobials.A descriptive comparative study was conducted between May 2010 and August 2011. E. coli isolates studied, MDR patterns were significantly higher among isolates from Sudan than Saudi Arabia [92.2% (214/232) vs. 70.6% (120/170)] (p = 0.000). The resistance rates of E. coli isolates were recorded as follows (Sudan and Saudi Arabia): High to moderate resistance to amoxicillin (97.7% and 94.2%), trimethoprim-sulfamethoxazole (88.3% and 82.5%), tetracycline (77.1% and 74.2%), amoxicillin- clavulanic acid (51.4% and 70%), ceftriaxone (64% and 52.4%) and ciprofloxacin (58.4% and 40%). Low resistance was to ceftazidime (35% and 20%), gentamicin (35% and 17.5%) and nitrofurantoin (22.4% and 11.7%). Resistance to amikacin was uncommon (1.9% and 5%). Significant differences (p < 0.05) in resistance rates of isolates between both countries in term to patient\u2019s gender and age. The most frequent MDR phenotypes among isolates were to 7(15.9%) in Khartoum state and to 3(20.8%) in Aseer region.Of the 402 E. coli isolates was observed in both regions. Continuous monitoring of resistance profiles, locally and international surveillance programs are required.Variation and emerging of antimicrobial resistance among pathogenic  Escherichia coli is a clinically significant bacterium because they are the most common species recovered in the clinical laboratories and has been incriminated in human infectious diseases  phenotypes make wide ranges of antimicrobials usefulness to treat most infections caused by this bacterium  as recommended by Clinical Laboratory Standard Institute , against 15 antimicrobial agents from different categories including: amikacin (30 \u03bcg), amoxicillin (10 \u03bcg), amoxicillin-clavulanic acid (30 \u03bcg), ceftazidime (30 \u03bcg), ceftriaxone (30 \u03bcg) cefuroxime (30 \u03bcg), chloramphenicol (30 \u03bcg), ciprofloxacin (5 \u03bcg), gentamicin (10 \u03bcg), nalidixic acid (30 \u03bcg), nitrofurantoin (50 \u03bcg), ofloxacin (5 \u03bcg), tetracycline (30 \u03bcg), tobramicin (10 \u03bcg) and trimethoprim- sulfamethoxazole (25 \u03bcg) . E. coli isolate was considered non-susceptible to an antimicrobial agent when it tested resistant, intermediate or non-susceptible when using clinical breakpoints as interpretive criteria, provided by the CLSI, (2011). MDR patterns of E. coli isolates were defined as non-susceptibility to at least one agent in three or more antimicrobial categories  software. Comparisons of antimicrobial resistance rate of isolates from both countries were done in term of setting, sex and age. The proportions were compared using the Chi-square test. A p-value of less than 0.05 was considered as statistically significant.E. coli isolates  were examined for antimicrobial susceptibility testing using double-disk diffusion method against 15 antimicrobial agents from different categories. Out of the 402 E. coli isolates tested for their antimicrobial susceptibility, 334 isolates were characterized as MDR strains from the two geographical regions. The majority of MDR E. coli were recovered from specimens of urine (n = 220) followed by wounds (n = 59) and vaginal swabs (n = 20) with low isolation rates from other specimens  were found to be MDR E. coli. These 214 MDR E. coli isolates were obtained from all age groups: 125 (58.4%) were from females and 89 (41.6%) were from males. Of these 214 MDR isolates, 168 (78.5%) were obtained from adult patients, whereas 46 (21.5%) from children.Out of 232 E. coli strains collected from hospitals in Aseer region, Saudi Arabia, MDR E. coli represented 120 (70.6%) of total isolates. These 120 isolates were obtained from females (n = 77) and males (n = 43). Eighty seven (72.5%) of the 120 isolates were from adults, whereas 33 (27.5%) were from children patients.Of the 170 E. coli sourced from Khartoum state, Sudan than that from Aseer region, Saudi Arabia isolates [92.2% (214/232) vs. 70.6% (120/170)] (p = 0.000).The occurrence of MDR patterns was found to be significantly higher among E. coli recovered from clinical specimens of patients in Kartoum state, Sudan and Aseer region, Saudi Arabia. The resistance rates among the strains from both countries were recorded as follows (Sudan and Saudi Arabia): High to moderate resistance rates of isolates were observed in amoxicillin (97.7% and 94.2%), cefuroxime (92.5% and 35.8%), trimethoprim-sulfamethoxazole (88.3% and 82.5%), tetracycline (77.1% and 74.2%), nalidixic acid (72% and 57.5%) ceftriaxone (64%). ciprofloxacin (58.4% and 40%), ofloxacin (55.1% and 38.3%), amoxicillin-clavulanate (51.4% and 70%). Low resistance rates of isolates were observed to ceftazidime (35% and 20%), gentamicin (35% and 17.5%), nitrofurantoin (22.4% and 11.7%), chloramphenicol (18.2% and 18.3%), and tobramicin (18.2% and 29.2%). Resistance to amikacin was uncommon (1.9% and 5%).E. coli isolates collected from Khartoum state, when compared to those from Aseer region. Strains from Khartoum state were more resistant to ceftazidime, ceftriaxone, cefuroxime, ciprofloxacin, gentamicin, nalidixic acid, nitrofurantoin and ofloxacin, whereas isolates fromAseer region were more resistant to amoxicillin-clavulanic acid and tobramicin. In general, Sudan sourced strains were more resistant to antimicrobial agents than those from Saudi Arabia.As shown in E. coli strains recovered from male patients in Khartoum state, Sudan and Aseer region, Saudi Arabia. Higher percentage of resistance was reported among isolates from males in Khartoum state than those from Aseer region for ceftriaxone (p < 0.001), cefuroxime (p < 0.001), gentamicin (p = 0.002), nitrofurantoin (p = 0.037), trimethoprim-sulfamethoxazole (p = 0.001). In contrast, higher resistance rates among isolates from Aseer region were observed for amoxicillin-clavulanic acid (p < 0.001) and tobramicin (p = 0.003).E. coli isolates recovered from female patients in Khartoum state to those fromAseer region  of antimicrobial drugs. Among the isolates collected from Khartoum state, Sudan the most prevalent MDR phenotypes were to 7 (15.9%), followed by 8 (11.7%) of tested antimicrobial agents. While among the strains originated from Saudi Arabia, the most frequent MDR patterns were to 3 (20.8%) followed by 5 (13.3%) and 4 (15.8%) of antimicrobial agents.The most frequent resistance profiles among Sudan strains were: AML-AMC-CXM-CRO-CAZ- CIP-OFX-NA-SXT-TE (8 strains) followed by AML-CXM-CRO-CAZ-CIP-OFX-NA-SXT-TE and AML-CXM-SXT-TE (7 strains), AML-CXM-CRO-CAZ-CIP-OFX-NA-SXT-TE-GN-TOB and AML-CXM- CRO-CAZ-F-SXT-C (6 strains each). Among Saudi Arabia strains, the predominant resistance profiles were: AML-SXT-TE (12 strains), AML-AMC-CXM-CRO-CAZ-CIP-OFX-NA-SXT-TE-TOB (10 strains), AML-AMC -SXT-TE (8 strains) and AML-AMC-NA-SXT-TE (6 strains).E. coli isolates recovered from two different unrelated geographical areas of Khartoum state in Sudan and Aseer region in Saudi Arabia. Antimicrobial resistance rates vary among regions and countries, with increasing rates in the many parts of the world  when comparing to a survey done in Sudan in the year 2000 in which MDR were recorded as 58%  among in Sudan , Saudi Ain Sudan , other din Sudan  and devein Sudan . The higin Sudan .E. coli isolates non-susceptible to fluoroquinolones have been reported in many countries as in Thailand (>50%), China (\u226570%) and India (>80%)  . In the esistant . The higesistant . Also, pE. coli strains collected from Sudan than Saudi Arabia for ceftriaxone (64% vs. 31.7%) (p< 0.001) and ceftazidime (35% vs. 20%) (p = 0.004). In a study carried out at two hospitals in Makka city, Saudi Arabia, resistance rates of E. coli isolates were recorded as 36.2 % for cefuroxime, 24.6 % for ceftazidime and 18.3% for ceftriaxone , trimethoprim-sulfamethoxazole (94.4% vs. 74.4%) and gentamicin (38.2% vs. 11.6%) whereas, higher resistance to amoxicillin-clavulanic acid and tobramicin were observed in Saudi Arabia sourced strains  than that from Sudan . These figures are indication of variations in resistance patterns in both countries with relatively higher rates in Sudan. The possible explanation is that this could be due to differences in prescribing patterns, types and quality of antimicrobial agents, as well as availability of drugs. Or perhaps due to better control strategies in the use of antimicrobial drugs, monitoring of resistance levels and updating prescribing policies in Saudi hospitals than in Sudan. This study indicated that there were higher significant differences in resistance levels of The common MDR phenotypes in our isolates were between amoxicillin/trimethoprim-sulfamethoxazole/cefuroxime/tetracycline/or nalidixic acid/amoxicillin-clavulanic acid. In the early study of antimicrobial resistance in bacterial isolates from patients with diarrhea and urinary tract infection in the Sudan, the most common MDR profile of isolates was reported to ampicillin, amoxicillin, tetracycline, trimethoprim-sulfamethoxazole, sulfonamide, and chloramphenicol . LikewisE. coli isolates is alarming and emerging in the both regions of Khartoum state, Sudan as well as in Aseer, Saudi Arabia, with different kinds of resistance patterns. Such high rates of resistance among isolates might be due to misuse and unnecessary prescription of antimicrobial drugs in both countries. Continuous monitoring and updating of antimicrobial resistance profile data as well as hospital policy for restriction and prudent use of antimicrobial drugs can reduce the spread of MDR strains. Antimicrobial resistance varied between settings, population and countries therefore, local and international surveillance programs in each region and setting are required.The study concluded that antimicrobial resistance of pathogenic"}
+{"text": "A reader noted a number of issues with Figure 1. The figure legend was:10.1096/fj.09-136481; Figure 1D, rotated) and Sallustio et al. Kidney International (2013) . Panels D and J from the PLOS ONE paper are identical to Figure 1D of Sallustio et al. Kidney International (2013), though flipped (doi:10.1038/ki.2012.413).Figure 1 panels D, E, & F  come from the same original images as panels J, K, & L (representing glomerular ARPC cells), respectively. Between E & K, there is a small overlap. Between panels D & J and F & L, respectively, the images have been rotated and there is a large overlap. Panels F and L are identical to panels in Sallustio et al. FASEB Journal (2010) , PAX2 (E), BMI-1 (F), and of glomerular ARPCs with Oct-4 (J), PAX2 (K), BMI-1 (L).10.1371/journal.pone.0087496. A correction is also being made to that article.In a subsequent PLOS ONE article, Figure 3 reproduced panels from Figure 1 of this article: Sciancalepore AG, Sallustio F, Girardo S, Gioia Passione L, Camposeo A, et al. (2014) A Bioartificial Renal Tubule Device Embedding Human Renal Stem/Progenitor Cells. PLOS ONE 9(1): e87496. doi:Fig. 1. Characterization of isolated glomerular and tubular ARPCs. Cytofluorimetric and immunofluorescence analysis of tARPCs showed the expression of: CD133 (A), CD24 (B), CD44 (C), Oct-4 (D), PAX2 (E), BMI-1 (F), Cytofluorimetric and immunofluorescence analysis of gARPCs showed the expression of: CD133 (G), CD24 (H), CD44 (I), Oct-4 (J), PAX2 (K), BMI-1 (L). CD106 expression in glomerular (M) and tubular (N) ARPCs. Original view: X63.S1 FileCharacterization of isolated glomerular and tubular ARPCs, showing immunofluorescence of tubular ARPCs with Oct-4 (D), PAX2 (E), BMI-1 (F), and of glomerular ARPCs with Oct-4 (J), PAX2 (K), BMI-1 (L).(ZIP)Click here for additional data file."}
+{"text": "Streptococcus dysgalactiae subsp. equisimilis) emerged as human pathogens in the late 1970s. We report here the draft genome sequences of four genetically distinct human strains of GCS-GGS isolated between the 1960s and 1980s. Comparative analysis of these genomes may provide a deeper understanding of GCS-GGS genome and virulence evolution.\u03b2-Hemolytic group C and group G streptococci (GCS-GGS;  Streptococcus dysgalactiae subsp. equisimilis can infect humans and other mammals (\u2013Large-colony-forming \u03b2-hemolytic isolates of Lancefield group C and group G Streptococci (GCS-GGS) identified as  mammals , 2. GCS  mammals \u20135. The tmammals \u2013, 6, 7. Tmammals \u2013, 9. The S. dysgalactiae subsp. equisimilis with 16S rRNA sequencing (GenBank accession numbers KP972460 to KP972463), and the genetic relatedness of the GCS-GGS isolates was determined via multilocus sequence typing (MLST) (GenBank accession numbers KT347549 to KT347565)   11, 12), 12S. dyhttp://www.ncbi.nlm.nih.gov/genome/annotation_prok/) and RASTtk pipelines (Assembled genomes were annotated using NCBI PGAP version 2.1 (rev. 462191) (ipelines  in conjuipelines . The 16Sipelines . AutomatThe draft genome sequences have been deposited as whole-genome shotgun projects at DDBJ/EMBL/GenBank under the accession numbers listed in"}
+{"text": "To evaluate the effect of surface coils for carotid MR imaging on PET quantification in a clinical simultaneous whole-body PET/MR scanner. A cylindrical phantom was filled with a homogeneous 2L water-FDG mixture at a starting dose of 301.2MBq. Clinical PET/MR and PET/CT systems were used to acquire PET-data without a coil (reference standard) and with two carotid MRI coils . PET-signal attenuation was evaluated with Osirix using 51 (PET/MR) and 37 (PET/CT) circular ROIs. Mean and maximum standardized uptake values (SUVs) were quantified for each ROI. Furthermore, SUVs of PET/MR and PET/CT were compared. For validation, a patient was scanned with an injected dose of 407.7MBq on both a PET/CT and a PET/MR system without a coil and with both coils. PET/MR underestimations were -2.2% (Siemens) and -7.8% (Machnet) for SUVmean, and -1.2% (Siemens) and -3.3% (Machnet) for SUVmax, respectively. For PET/CT, underestimations were -1.3% (Siemens) and -1.4% (Machnet) for SUVmean and -0.5% (both Siemens and Machnet) for SUVmax, respectively using no coil data as reference. Except for PET/CT SUVmax values all differences were significant. SUVs differed significantly between PET/MR and PET/CT with SUVmean values of 0.51-0.55 for PET/MR and 0.68-0.69 for PET/CT, respectively. The patient examination showed that median SUVmean values measured in the carotid arteries decreased from 0.97 without a coil to 0.96 (Siemens) and 0.88 (Machnet). Carotid surface coils do affect attenuation correction in both PET/MR and PET/CT imaging. Furthermore, SUVs differed significantly between PET/MR and PET/CT."}
+{"text": "Photobacterium  were isolated from bleached coral Madracis decactis (scleractinian) in the remote St Peter & St Archipelago (SPSPA), Mid-Atlantic Ridge, Brazil. Healthy M. decactis specimens were also surveyed, but no strains were related to them. The novel isolates formed a distinct lineage based on the 16S rRNA, recA, and rpoA gene sequences analysis. Their closest phylogenetic neighbours were Photobacterium rosenbergii, P. gaetbulicola, and P. lutimaris, sharing 96.6 to 95.8% 16S rRNA gene sequence similarity. The novel species can be differentiated from the closest neighbours by several phenotypic and chemotaxonomic markers. It grows at pH 11, produces tryptophane deaminase, presents the fatty acid C18:0, but lacks C16:0 iso. The whole cell protein profile, based in MALDI-TOF MS, distinguished the strains of the novel species among each other and from the closest neighbors. In addition, we are releasing the whole genome sequence of the type strain. The name Photobacterium sanctipauli sp. nov. is proposed for this taxon. The G + C content of the type strain A-394T (= LMG27910T = CAIM1892T) is 48.2 mol%.Five novel strains of  Photobacterium comprises 26 formally described species   were identified as P. phosphoreum, P. damselae and P. mandapamensis   and coram corals . P. jeann Brazil , whereasustralia , as welln Brazil . Photobapamensis . Coral mpamensis .Madracis decactis in the Brazilian St Peter & St Paul Archipelago (SPSPA) analyzed 403 isolates , and RNA polymerase alpha subunit (rpoA) were obtained as described previously (\u00ae). Through this method, the DNA was simultaneously fragmented and tagged with sequencing adapters. The library size distribution was accessed using the 2100 Bioanalyzer and the High Sensitivity DNA Kit (Agilent\u00ae). The accurate quantification of the library was accomplished using the 7500 Real Time PCR (Applied Biosystems\u00ae) and the KAPA Library Quantification Kit (Kapabiosystems\u00ae). Paired-end (2 \u00d7250 bp) sequencing was performed on a MiSeq (Illumina\u00ae) using the MiSeq reagent kit v2 (500 cycles). R1 and R2 reads were quality filtered (Q > 20) and 3\u2019 end trimmed with Prinseq v0.20.4 . The gene sequence data obtained in this study are available through the open access website TAXVIBRIO (http://www.taxvibrio.lncc.br/). The GenBank accession numbers for the 16S rRNA, recA, and rpoA genes and genome sequences are listed in \u00ae). Spectra were generated with mMass software v5.5.0  as described previously  medium at ambient temperature (\u223c27 \u00b0C) after 24\u201348 h incubation . Gene seeviously . Primerseviously . Sequenceviously . Trees weviously . The robeviously . For geneviously  was used v0.20.4 . Ray v. rameters . Generalrameters . In siliGGDC 2.0 . This onGGDC 2.0  to generGGDC 2.0 . DDH preGGDC 2.0  with logGGDC 2.0 . StrainsGGDC 2.0  towards e v5.5.0 . Type steviously  and by BPhotobacterium  confirmed that the isolates formed a distinct lineage related to P. rosenbergii and P. gaetbulicola  with their closest neighbours, respectively. These levels of similarity are below the cut-offs determined to define a species of the family Vibrionaceae  ranged from 99.8% to 100% based on recA. Their rpoA sequences were identical. Trees based on partial sequences of the housekeeping genes recA (855 bp) and rpoA (969 bp) also confirmed their phylogenetic position in the genus Photobacterium and revealed they constituted a separate branch, clearly indicating that they belong to a new Photobacterium species (T genome are supplied in In silico DDH (%) values between A-394T and P. angustum S14, P. damselae subsp. damselae CIP 102761, P. halotolerans DSM18316, P. leiognathi lrivu.4.1 and P. profundum 3TCK were 21.5 (\u00b12.34), 22.7 (\u00b12.37), 20.3 (\u00b12.31), 21.6 (\u00b12.35) and 20.6 (\u00b12.31) respectively. AAI between A-394T and P. leiognathi lrivu.4.1 CIP 102761 was 75%.16S rRNA gene sequence analysis revealed that the five isolates formed a tight monophyletic branch affiliated to the genus acterium . The fivacterium . P. rosetrarenae  and P. m marinum . The phybulicola . The novionaceae . The sim species \u2013S2. Gene18:0, and absence of C16:0 iso (P. rosenbergii (LMG 22223T), P. gaetbulicola (LMG 27839T) and P. lutimaris (LMG 25278T)  . MLSA wa 25278T) . Based oPhotobacterium sanctipauli .\u03b2-glucosaminidase, \u03b2-galactosidase and arginine dihydrolase; but negative for lipase (C14), valine arylamidase, cystine arylamidase, trypsin, \u03b1-chemotrypsin, acid phosphatase, \u03b1-galactosidase, \u03b2-glucuronidase, \u03b1-glucosidase, \u03b2-glucosidase, \u03b1-mannosidase, \u03b1-fucosidase, lysine decarboxylase, ornithine decarboxylase, H2S production, urease activity, acetoin production (Voges\u2013Proskauer) and gelatinase. Variable reactions were obtained for esterase (C4) (\u2212), esterase lipase (C8) (\u2212), tryptophane deaminase (w) and indole production (\u2212) . Reduces nitrate to nitrite but not to N2. Positive for fermentation/oxidation of glucose and mannitol but negative for inositol, sorbitol, rhamnose, saccharose, amygdalin and arabinose. Melibiose (+) gave variable reactions. D-Salicin, \u03b1-D-glucose, D-mannose, D-galactose are used as sole energy sources. Does not utilize dextrin, D-raffinose, glycerol, N-acetyl-D-galactosamine, D-glucose-6-PO4, D-aspartic acid, D-serine, gelatin, glycyl-L-proline, L-alanine, L-arginine, L-aspartic acid, L-glutamic acid, L-pyroglutamic acid, L-serine, pectin, L-galactonic acid lactone, mucic acid, quinic acid, D-saccharic acid, p-hydroxy-phenylacetic acid, methyl pyruvate, D-lactic acid methyl ester, citric acid, D-malic acid, bromo-succinic acid, \u03b3-amino-butyric acid, \u03b1-hydroxy-butyric acid, \u03b2-hydroxy-D,L-butyric acid, propionic acid, acetic acid and formic acid. The following reactions are variable within the species: citrate (\u2212), D-maltose (\u2212), D-trehalose (\u2212), D-cellobiose (w), gentiobiose (\u2212), sucrose (\u2212), D-turanose (\u2212), stachyose (\u2212), \u03b1-D-lactose (\u2212), D-melibiose (\u2212), \u03b2-methyl-D-glucoside (\u2212), N-acetyl-D-glucosamine (\u2212), N-acetyl-\u03b2-mannosamine (\u2212), N-acetyl neuraminic acid (\u2212), D-fructose (\u2212), 3-methyl glucose (w), D-fucose (w), L-fucose (w), L-rhamnose (\u2212), inosine (\u2212), D-sorbitol (\u2212), D-mannitol (\u2212), D-arabitol (\u2212), myo-inositol (\u2212), D-glucose-6-PO4 (\u2212), L-histidine (w), D-galacturonic acid (\u2212), D-gluconic acid (\u2212), D-glucuronic acid (\u2212), glucuronamide (w), L-lactic acid (\u2212), \u03b1-keto-glutaric acid (w), L-malic acid (\u2212), tween 40 (\u2212) and acetoacetic acid (w). Does not assimilate any of the substrates included in the API 20 NE system. The most abundant cellular fatty acids are summed feature 3 , C16:0 (21.4%), C18:1\u03c97c (11.6%), C14:0 (5.2%), C12:0 and summed feature 2 , C12:03\u2013OH(2.5%), C17:0 (1.6%), Iso-C17:0 (1.5%), Iso-C15:0 and C17:1\u03c98c (1.1%), and in minor amounts C13:0, C17:1\u03c96c, C18:0 and Unknown 12.484 (0.3\u20130.5%). The G + C content of the type strain (A-394T) is 48.2 mol%. The type strain is A-394T (= LMG 27910T = CAIM 1892T). It was isolated from the tissues of bleached Madracis decactis (Scleractinia) in St Peter & St Paul Archipelago, Brazil. SPSPASt Peter & St Paul ArchipelagoMLSAmultilocus sequence analysisAAIaverage amino acid identityDDHDNA-DNA hybridizationGGDCGenome-To-Genome Distance CalculatorFAMEfatty acid methyl ester analysesMALDI-TOFmatrix-assisted laser desorption/ionization time-of-flightColonies are small, beige, irregular shaped, with smooth and translucent edge and 1\u20132 mm in diameter after 24 h at 28 \u00b0C on MA under aerobic conditions. On TCBS colonies are green, round with a smooth border and 2\u20133 mm in diameter. Cells are small bacilli measuring 2\u20133 \u00b5m in diameter, Gram-negative, motile, facultative anaerobic, oxidase and catalase-positive. Grows well between 20 and 30 \u00b0C but not at 4 and 45 \u00b0C. No growth occurs in the absence of NaCl, but grows well under NaCl concentrations of 1%\u20138% (w/v). Grows at pH 6-11. Positive for alkaline phosphatase, leucine arylamidase, naphtol-AS-BI-phosphohydrolase, N-acetyl-10.7717/peerj.427/supp-1Table S1Upper GenBank accession numbers for the 16S rRNA gene, recA and rpoA housekeeping genes and genome sequences of Photobacterium sanctipauli sp. nov.; and for recA and rpoA of P. gaetbulicola LMG 27839T (data from this study). Lower Accession numbers for the Photobacterium strains\u2019 genomes used for GGD calculation (data publicly available at GenBank).Click here for additional data file.10.7717/peerj.427/supp-2Table S2Photobacterium sanctipauli sp. nov. and source information.Strains of Click here for additional data file.10.7717/peerj.427/supp-3Table S3T (= LMG 27910T = CAIM1892T) genome determined in RAST environment.Statistics and general features of A-394Click here for additional data file.10.7717/peerj.427/supp-4Table S41, P. sanctipauli (range profile of five strains); 2, P. rosenbergii LMG 22223T . The evolutionary distances were computed using the Kimura 2-parameter method. Phylogenetic analyses were conducted in MEGA5. Bootstrap values (>50%) shown are based on 1,000 repetitions. Vibrio maritimus R-40493T was used as outgroup. Bar, 2% estimated sequence divergence.Neighbour-joining phylogenetic tree showing the position of Click here for additional data file.10.7717/peerj.427/supp-7Figure S2P. sanctipauli sp. nov based on rpoA gene sequences (969 bp). The evolutionary distances were computed using the Kimura 2-parameter method. Phylogenetic analyses were conducted in MEGA5. Bootstrap values (>50%) shown are based on 1,000 repetitions. Vibrio maritimus R-40493T was used as outgroup. Bar, 1% estimated sequence divergence.Neighbour-joining phylogenetic tree showing the position of Click here for additional data file.10.7717/peerj.427/supp-8Figure S3A\u2212T394, A-373, A-379, A-397 and A-398) showing they are not clonal. The closely related type strains of P. rosenbergii (LMG 22223T), P. gaetbulicola (LMG 27839T) and P. lutimaris (LMG 25278T) were included in the analysis. The dendrogram was constructed using Pearson correlation coefficient and UPGMA.Comparison of the MALDI-TOF MS fingerprint profiles of the 5 novel strains (Click here for additional data file."}
+{"text": "To evaluate CT-morphological features of lung cancers detected in a CT lung cancer screening trial, and to determine the correlation between CT-morphological features and the histopathological diagnosis of screen-detected cancers.197 solid lung cancers (192 participants) detected in all four screening rounds of the Dutch-Belgian randomized lung cancer screening trial (NELSON) were included. CT-morphological features included nodule shape, margin, location, volume, and volume-doubling time (VDT). Based on histopathology, cancers were divided into four groups: adenocarcinoma (N=114), squamous cell carcinoma (N=37), large cell carcinoma (N=28) and neuro-endocrine cancers (N=18). Data were analyzed using ANOVA, Chi-square and Fischer\u2019s exact test.Mean participant age was 61.3 years , and 160/192 (83.3%) were male. In all four histopathologic groups, the majority of cancers had a spherical nodule shape (70.6-95.8%). Margins of malignant nodules were most often lobulated (39.3-45.9%) and spiculated (22.2-35.7%), without statistically significant difference between histopathological groups. Most cancers (63.5%) were located in the upper lobes, adenocarcinomas significantly more often  than other types of cancers (p=0.004). Adenocarcinomas had a higher mean VDT than large cell carcinomas . VDTs of other histopathological subgroups did not differ significantly.Only VDT and location in the upper versus lower lobes are associated with histopathological diagnosis of screen-detected lung cancers. No discrimination can be made between different histopathologic cancer types based on CT-morphological features alone."}
+{"text": "Combination antiretroviral therapy (cART) may affect vitamin D [25(OH)D], parathyroid hormone (PTH), bone mineral density (BMD) and bone turnover (BT). Reduced BMD and secondary hyperparathyroidism have been reported with tenofovir (TDF). We investigated the associations between TDF and bone markers, especially in 25(OH)D-deficient patients.2) were measured using dual-energy X-ray absorptiometry. BT was assessed by serum type 1 procollagen (P1NP) and carboxy-terminal collagen cross-links (CTX). Mann\u2013Whitney U tests compared serum markers and BT, and t-tests compared BMD according to TDF in all and 25(OH)D-deficient patients.In a single-centre longitudinal study investigating BMD in HIV-positive men, serum 25(OH)D, calcium, phosphate, PTH and alkaline phosphatase (ALP) were measured. Lumbar spine, non-dominant total hip and non-dominant femoral neck BMD  years, 94% white ethnicity, 93% MSM, diagnosed HIV positive for median 9.6  years, median CD4 547  cells/\u00b5L, HIV RNA <40 copies/mL in 87% (96% of those on cART). 25(OH)D (nmol/L) was normal (>75), insufficient (50\u201375), deficient (25\u201350) and severely deficient (<25) in 14%, 29%, 50% and 7%, respectively. Of 381 men on cART, 77% were currently on TDF. TDF was not associated with median calcium (p=0.69) or phosphate (p=0.52), but patients had higher  median ALP [81  vs. 73  IU/L, p=0.005) compared to non-TDF cART. There was no difference in the association between vitamin D and PTH according to whether someone currently was D deficiency, hyperparathyroidism, reduced BMD or increased BT, although patients on TDF had higher but normal ALP. We found no evidence to support additional monitoring of bone markers in patients on TDF regardless of 25(OH)D status."}
+{"text": "Age is considered an independent risk factor for short and long-term mortality of elderly ICU patients, when very elderly (\u226585 years) and mid-elderly (75-84 years) populations are compared to young elderly (65-74 years) ones. However, it is not yet clear if this trend remains when very elderly are compared to mid-elderly populations.a) to compare short  and long-term (6-months and 1-year) mortality of mid-elderly to very elderly patients andb) to evaluate the influence of patients' characteristics on mortality.Single center, retrospective, observational study in an 8-bed adult general ICU (January 2011-June 2014). Patients \u226575 years were divided into two age-groups, 75-84 and \u226585 years old. Characteristics on ICU admission were recorded. Patients hospitalized \u226448 hours were excluded. ICU, hospital, 6-months and 1-year after hospital discharge mortality were calculated. Univariate analysis for categorical variables was performed using Pearson\u2019s x2 or Fisher test and Student t-test for continuous data. Multivariate analysis of the time to ICU mortality was calculated using Cox regression model and for hospital, 6-months and 1-year mortality using logistic regression analysis. P value < 0.05 was considered significant.244 patients were included, 195 in the 75-84 and 49 in the \u226585 years group. Mortality rates for the two groups were: ICU, 74/195 (37.9%) vs. 24/49 (49%) (p = 0.08), hospital, 23/121 (19%) vs. 5/25 (20%) (p = 0.90), 6-months, 15/95 (17%) vs. 4/20 (20%) (p = 0.74) and 1-year, 22/80 (27.5%) vs. 7/16 (20%) (p = 0.19), respectively. in multivariable analysis, patients with malignancy as reason for ICU admission had increased ICU mortality risk . Patients with higher APACHE II score on ICU admission had more important 1-year mortality risk .More than one third of our ICU patients were mid- or very elderly. No difference was observed between the two age-groups considering short or long-term mortality. Malignancy as a reason for ICU admission and APACHE II score negatively influenced the ICU and 1-year mortality, respectively."}
+{"text": "H. pylori infection. H. pylori stimulate Wnt/\u03b2-catenin pathway by activating oncogenic c-Met and epidermal growth factor receptor (EGFR), or by inhibiting tumor suppressor Runx3 and Trefoil factor 1 (TFF1). H. pylori also trigger Wnt/\u03b2-catenin pathway by recruiting macrophages. Moreover, Wnt/\u03b2-catenin pathway is found involved in H. pylori-induced gastric cancer stem cell generation. Recently, by using gastroids, researchers have further revealed that H. pylori induce gastric epithelial cell proliferation through \u03b2-catenin. These findings indicate that Wnt/\u03b2-catenin is an oncogenic pathway activated by H. pylori. Therefore, this pathway is a potential therapy target for H. pylori-related gastric cancer.A section of gastric cancers presents nuclear \u03b2-catenin accumulation correlated with  Wnt/\u03b2-catenin pathway, also called canonical Wnt pathway, is crucial to embryo development and adult tissue homeostasis , 2. AberHelicobacter pylori (H. pylori) infection is a strong risk factor for gastric cancer. The underlying mechanisms include chronic inflammation in gastric mucosa, genetic and epigenetic alterations of tumor suppressor genes, activation of oncogenic signals, and generation of gastric cancer stem cells (CSC)  [H. pylori can cause DNA double-strand breaks directly [H. pylori-related gastric carcinogenesis [H. pylori induce gastric epithelial cell epithelial-mesenchymal transition (EMT), and generate potential cancer stem cells [H. pylori also stimulate some oncogenic pathways. Activation of epidermal growth factor receptor (EGFR) can resist H. pylori-induced gastric epithelial cell apoptosis [H. pylori-induced gastric carcinogenesis.) Figure . Chronic (BMDCs) , 10. H. directly , and caudirectly , 13 or bdirectly . Hypermeogenesis . H. pyloem cells , 17. H. poptosis , 19. Mor\u03b2-catenin exon 3 that encodes serine-threonine phosphorylation sites for the GSK3\u03b2 [\u03b2-catenin mutation [APC mutation [APC methylation [Investigations on gastric cancer specimens showed that about 20%\u223c30% gastric cancers presented nuclear \u03b2-catenin accumulation . Some muhe GSK3\u03b2 . These mmutation . In colomutation . Unlike mutation , 24 nor hylation  seemed tH. pylori to nuclear \u03b2-catenin accumulation. Nuclear \u03b2-catenin mainly localized in epithelial cells within the proliferative zone in antral glands, and appeared more frequently in H. pylori cytotoxin-associated gene A (CagA)-positive specimens, compared with either CagA-negative or uninfected patients [H. pylori could induce nuclear \u03b2-catenin accumulation in vivo and in vitro [H. pylori promoted gastric epithelial cell proliferation through \u03b2-catenin by using gastroids, three-dimensional organ-like structures [Several studies from a same group demonstrated the attribution of patients . CagA-poin vitro -28. Receructures .H. pylori induced phosphorylation of c-Met and gastric epithelial cell proliferation [H. pylori-induced gastric epithelial cell proliferation and atrophic gastritis [Aberrant activation of c-Met receptor occurred commonly in gastric cancers . Infectiferation , 32. Upoferation , 32. CD4astritis . C-Met-Pastritis . On the astritis . Interesastritis , 36, indH. pylori-related gastric cancer [H. pylori secretory protein HP0175 [H. pylori induced EGFR phosphorylation, and then activated PI3K/Akt pathway [H. pylori infection  was expressed in normal gastric mucosa , but free mucosa , 56. In nfection . TFF1 innfection . On the -catenin   [It has been well accepted that tumor-associated macrophages (TAMs) promote cancer development.  (MCP-1) , 59 or S (MCP-1)  in gastr (MCP-1) , 7  . The sup (MCP-1) . Macroph (MCP-1) . These oH. pylori. MiR-101 and miR-320 were inhibited by H. pylori through CagA [H. pylori [H. pylori infection with Wnt/\u03b2-catenin activation in gastric cancer (Table MicroRNAs (miRs) are small noncoding RNAs that can up- or downregulate the expression of oncogenes and tumor suppressors. Some miRs, such as miR-101, mir-124a, miR-203, miR-210 and miR-320, were downregulated by ugh CagA , 65. Hypugh CagA -68. The ugh CagA -73, indi. pylori -76. Thes. pylori -79. The H. pylori may activate Wnt/\u03b2-catenin pathway by affecting Wnt ligands, receptors or antagonists. H. pylori and TNF-\u03b1 could induce Wnt10a and Wnt10b expression in gastric cancer cells [H. pylori infection could also activate Wnt co-receptor LRP6, and result in nuclear \u03b2-catenin accumulation [SFRP4 and SFRP5 methylation was found to be positively correlated with H. pylori infection [H. pylori on these molecules are still unclear.Some evidence indicates that er cells , 81. H. mulation . Secretemulation . Actuallnfection . In addinfection , Wnt7a [nfection , Wnt7b [nfection  and Wnt nfection , were alH. pylori colonized stomach gland epithelium, and promoted stem cell-related gene expression and Lgr5(+) stem cell proliferation [H. pylori also induced gastric epithelial cell EMT to generate gastric cancer stem cells, and this process was also via CagA [H. pylori-induced EMT and stem cell generation remain largely unknown. Wnt/\u03b2-catenin pathway was important for gastrointestinal progenitor cell proliferation and differentiation [H. pylori-induced gastric stem cell proliferation [H. pylori induce gastric stem cell generation and proliferation at least partly via Wnt/\u03b2-catenin pathway.Gastric stem cells are implicated in gastric cancer initiation and progression. Via CagA, feration . H. pylovia CagA . The molntiation , 91. Actntiation , 93. Recntiation . Moreoveferation . These fH. pylori in gastric carcinogenesis. H. pylori can upregulate Wnt/\u03b2-catenin activator c-Met and EGFR, and downregulate Wnt/\u03b2-catenin suppressor TFF1 and RUNX3. H. pylori can also activate Wnt/\u03b2-catenin pathway by recruiting tumor-associated macrophages. Importantly, via Wnt/\u03b2-catenin pathway, H. pylori induced gastric stem cell generation and expansion, promoting gastric cancer initiation and progression.Increasing evidence demonstrates Wnt/\u03b2-catenin as a crucial pathway stimulated by H. pylori infection, c-Met, EGFR, TFF1, Runx3, or else? Can dysregulations of these molecules synergize in gastric cancer development? Which virulent factor of H. pylori plays a dominant role in Wnt/\u03b2-catenin activation, CagA, VacA or else? Given the complexities of H. pylori strains and host factors, more work should be done to find the answers. In addition, the effects of H. pylori on Wnt ligands, receptors and antagonists, and the roles of miRs in Wnt/\u03b2-catenin activation in gastric cancer, need to be further investigated.However, there are still some questions need to be answered. Which signal molecule plays a dominant role in Wnt/\u03b2-catenin activation under H. pylori eradication can reduce the risk of gastric cancer, but it can not completely prevent H. pylori-related gastric carcinogenesis. One of the reasons is that the activation of oncogenic pathway, such as Wnt/\u03b2-catenin, has happened before H. pylori eradication.Recently, a series of therapies antagonizing Wnt/\u03b2-catenin pathway have entered clinical trials. As Wnt/\u03b2-catenin pathway is essential for tissue homeostasis, it remains elusive about their clinical efficacy and safety . H. pylo"}
+{"text": "Childhood-onset Systemic lupus erythematosus (cLES) is an autoimmune disease with a wide spectrum of clinical manifestations, characterized by periods of activity and remission. Hematological and immunological abnormalities are commonly found. Laboratory evaluation, including the cytokine profile, assists in the diagnosis, determination of disease activity and may predict future damage caused by the disease.To determine the serum levels of Th1 (IL-12), Th2 (IL-6 and IL-10) and Th17 (IL-17) cytokines in cSLE and healthy controls. To evaluate their association whit disease activity, laboratory and treatment features and Mood and anxiety disorders.We included 63 consecutive cSLE patients [median age 18 years (range 11-25)] and 59 healthy controls [median age 20 years (8-33)]. Controls were age and sex-matched to cSLE patients. SLE patients were further assessed for clinical and laboratory SLE manifestations, disease activity [SLE Disease Activity Index (SLEDAI)], damage [Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index (SDI)] and current drug exposures. Total dose of corticosteroids and other immunosuppressant medications used since the onset of disease were calculated by data obtained by careful review of the medical charts.Mood and anxiety disorders were determined through Becks Depression (BDI) and anxiety inventory (BAI). Th1 (IL-12), Th2 (IL-6 and IL-10) and Th17 (IL-17) cytokines levels were measured by ELISA and compared by non-parametric tests.Serum IL-6 (p=0.001), IL-10 (p=0.006), IL-12 (p=0.027) and IL-17 (p=0.0001) levels were increased in cSLE patients when compared to healthy controls. IL-6 levels were significantly increased in patients whit active disease (p=0.008). IL-6 (p=0.032) and IL-12 (p=0.028) levels were significantly increased in patients with active nephritis. We observed that IL-17 was associated with migraine (p=0.045), IL-6 with thrombocytopenia (p=0.022) and IL-12 with the presence of anxiety (p=0.048). No association between cytokine levels and SDI scores or medication was observed.Cytokines play a central role in cSLE, IL-6 is associated with SLEDAI and may be a biomarker of disease activity, Th1 and Th2 responses may play a role in lupus nephritis and Th1 and Th17 may play a role in neuropsychiatric symptoms in cSLE. Longitudinal studies are necessary to confirm their ability to predict SLE related manifestations.K. Peli\u00e7ari: None declared, M. Postal: None declared, R. Barbosa: None declared, N. Sinicato: None declared, F. Peres: None declared, R. Marini: None declared, S. Appenzeller Grant / Research Support from: FAPESP  CNPQ"}
+{"text": "Bacillus decisifrondis E5HC-32T (DSM 11725T) is a Gram-positive, subterminal spherical spore-forming, strictly aerobic bacterium. Here, we report the 5,613,728-bp genome sequence of B.\u00a0decisifrondis E5HC-32T, which is the first genome information of this species and will provide useful information for genomic taxonomy and phylogenomics of Bacillus-like bacteria. T was isolated from soil in Australia and was named as Bacillus decisifrondis sp. nov. .The type strain E5HC-32sp. nov. . Its phyd pH\u00a08.4 . NotablyB.\u00a0decisifrondis E5HC-32T (DSM 11725T) was performed via the Illumina Hiseq 2500 system. Two DNA libraries with insert sizes of 500\u00a0bp were constructed and sequenced. After filtering of the 0.67-Gb raw data, 0.66-Gb of clean data was obtained, providing approximately 100-fold coverage. The reads were assembled via SOAPdenovo software version 2  utilizing GeneMark, Glimmer, and tRNAscan-SE tools (The annotation of the genome was performed using the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (SE tools . A totalSE tools .LIYL00000000. The version described in this paper is version LIYL00000000.1.This whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession no."}
+{"text": "Bacillus sp. SM1 strain MCC2138), which was isolated from marine coastal waters and has the ability to degum raw silk fabric as well as Ramie fiber. The genome comprises 1.76\u00a0Mb with a GC content of 34.5%.Reported here is the draft genome sequence of an amylase-, protease-, DNase-, oxidase-, gelatinase-, and catalase-producing, Gram-positive diplobacillus ( It could accumulate Pb, Cr, Ni, Cu, and Co  with a 2.0\u00d7 coverage generated on an Illumina platform, totaling 17,69,494\u00a0bp, with a largest contig length of 5,01,909 bp and T server  revealedThe important genes contained in the different contigs are as follows: ABC transporter ATP binding protein; redox-sensitive transcriptional regulator (AT-rich DNA binding protein); heat shock protein 60 family cochaperone GroES/GroEL; ATP pyrophosphatase subunit (EC 6.3.5.2); phosphate regulon transcriptional regulatory protein PhoB (SphR) (accession no. LXJX01000001.1); oligopeptide ABC transporter, periplasmic oligopeptide-binding protein OppA/B/C/D/F (TC3.A.1.5.1); hydrolase, tRNA; glycerate kinase (EC2.7.1.31); arginase (EC3.5.3.1); diadenylate cyclase spyDAC; bacterial checkpoint controller DisA with nucleotide-binding domain; phosphoglucosamine mutase ; programmed cell death toxin YdcE and YdcD; outer membrane lipoprotein receptor; Holo (acyl carrier protein) synthase (EC2.7.8.7); UV endonuclease UvdE family; DEAD box ATP-dependent RNA helicase CshA (EC3.6.4.13); t-RNA (accession no. NZ_LXJX01000007.1); ABC transporter permease protein YvcS; ABC transporter ATP-binding protein YvcR; beta-lactamase class A; secreted and spore coat\u2013associated protein 1; chitinase (EC 3.2.1.14); cell-division protein FtsW; microbial collagenase (EC 3.4.24.3); transcriptional regulator, TetR family (accession no. NZ_LXJX01000017.1); cysteine synthase (EC 2.5.1.47); chaperonin (heat shock protein 33); tRNA (Ile) lysidine synthetase (EC 6.3.4.19); AbrB family transcriptional regulator (SpoVT); transcription repair coupling factor; Fin, required for the switch from sigma F to sigma G during sporulation; peptidyl-tRNA hydrolase (EC 3.1.1.29) (accession no. NZ_LXJX01000019.1).LXJX00000000. The version described in this paper is the first version, LXJX01000000.This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession number"}
+{"text": "AbstractDulichiella pattaniensis is new to science, and Melitalati latiflagella Ren & Andres, 2012 has not been previously reported from Thai Waters. Dulichiella pattaniensis is characterized by male gnathopod 2 distolateral crown with 4 spines; pleonite/urosomite formula 7-7-7-5-6-2; pereopod 5-7 dactylus with 2 accessory spines. This combination of characters has not been recorded previously in the Dulichiella. The characters of the specimens are described and illustrated. All specimens are deposited in the Princess Maha Chakri Sirindhorn Natural History Museum, Prince of Songkla University, Thailand.Two species of melitid amphipod were collected from the Gulf of Thailand.  Rotomelita longipropoda Wongkamhaeng et al., 2013 was described. In this study, we describe a new melitid species Dulichiella pattaniensis sp. n., and our observations of Melita latiflagella Ren & Andress, 2012, which has not been previously reported in Thai Waters. Figures and descriptions of both species are provided.Melitid amphipods most commonly occur in coastal and freshwater areas. Thailand has a variety of aqueous habitats including coral reefs, seagrass beds, and mangrove forests, but only one melitid amphipod, , antennaA; , gnathopodG; , headHD; , lower lipLL; , mandibleMD; , maxillaMX; , maxillipedMP; , pereopodP; , pleopodPl; , telsonT; , uropodU; , urosomeUR; , upper lipUL; , rightr; , leftl; , male\u2642; , female\u2640. Specimens of different species were deposited into the Prince of Songkla University Zoological Collection (PSUZC).Amphipods were collected from some settlement plates in an artificial reef in Ban Pak Bang Ta Wa, Pattani Bay and from sediment of Lower Songkhla Lake . The sitStout, 1912http://species-id.net/wiki/DulichiellaPageBreakmales with bunches of long slender setae. Pereopod 7 basis in female fully expanded. Pleonites dorsally serrate. Uropod 3 inner ramus scale-like; outer ramus 4 to 5\u00d7 longer than wide, 2-articulate. Telson deeply cleft, lobes tapering distally to an acute point. Head antDulichiella spinosa Stout, 1912 (type by monotypy).Dulichiella appendiculata Say, 1818; Dulichiella australis Haswell, 1879; Dulichiella celestun Paz-Rios & Ardisson, 2014; Dulichiella cotesi Giles, 1890; Dulichiella cuvettensis Appadoo & Myers, 2005; Dulichiella fresnellii ; Dulichiella guinea Lowry & Springthorpe, 2007; Dulichiella lecroyae Lowry & Springthorpe, 2007; Dulichiella oahu Lowry & Springthorpe, 2007; Dulichiella pacifica Lowry & Springthorpe, 2005; Dulichiella pattaniensis sp. n.; Dulichiella spinosa Stout, 1912 (type species); Dulichiella takedai Tomikawa & Komatsu; Dulichiella terminos Lowry & Springthorpe, 2007; Dulichiella tomioka Lowry & Springthorpe, 2007; Dulichiella tulear Lowry & Springthorpe, 2007.PageBreakhttp://zoobank.org/212F51D8-E32A-4601-9E04-88292FBEB989http://species-id.net/wiki/Dulichiella_pattaniensis6\u00b051'55\"N, 101\u00b010'7\"E), artificial reef , 1 September 2010, Puttapreecha, R., PSUZC-CR-0192. Allotypes, \u2640 collected with holotype; PSUZC-CR-0193; Paratype, collected with holotype ).Holotype. \u2642, THAILAND, Lower Gulf of Thailand, Pattani Bay , flagellum can be 25-articulate . Antenna 2, Antenna 2 setiferous, peduncular article 2 cone gland not reaching end of article 3; article 5 subequal to 4, flagellum 9-articulate.Based on male holotype. Body length 6.3 mm (from tip of rostrum to apex of telson). Upper lip, (labrum) distally rounded. Lower lip, inner lobes well developed, pubescent. Mandible, both similar, left incisor 3 dentates, right incisor 4 dentate; left and right lacinia mobilis armed with 3 and 4 dentates respectively; palp slender with marginal setae, article 1 smooth, article 3 slightly longer than article 2. Maxilla 1, inner plate narrow with 2 apical plumose setae; outer plate with 8 apical serrate robust setae; palp 2-articulate, article 1 with 3 distal setae, article 2 with 4 apical robust setae and 4 apical setae. Maxilla 2, inner plate with mediofacial row of 29 setae and 14 apical plumose setae; outer plate broader than inner plate, distally setose. Maxilliped, inner plate borad, with 6 plumose marginal setae; outer plate margin with 11 conate robust setae, terminal with 4 plumose setae; palp 4-articulate, article 2-3 with marginal setae, article 4 tapering with fine marginal setae.Pereon.PageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakGnathopod 1 subchelate, smaller than gnathopod 2; coxa anterodistal corner not produced, posteroventral corner notch present, anterior margin straight; length ratio of articles from basis to dactylus 10: 3:4:7:5: 3; basis slender; merus\u2013propodus setose; palm slightly convex, defined by posterodistal corner, without posterodistal robust setae. Gnathopod 2 sexually dimorphic; left and right gnathopods unequal in size; coxa posteroventral corner notch present; (larger) length ratio of article from basis to dactylus 8:1:3:1:10:11; propodus distolateral corner crown with 4 rounded spines, palm straight, posterodistal corner produced, upturned, fit with dactylus; dactylus apically blunt;  subchelate; length ratio of article from basis to dactylus 9:3:4:6:6:4; merus with sharp posteroventral spine; carpus subequal to propodus; palm straight, without posteroventral spine. Pereopod 3\u20134 alike. Pereopod 5 basis posterior margin straight, posteroventral corner rounded; carpus and propodus sparsely setose; dactylus unguis anterior margin with accessory spines. Pereopod 6\u20137 alike, basis, merus, carpus, propodus with long marginal setae. Pereopod 6 basis posterior margin straight, minutely castelloserrate; dactylus unguis anterior margin with accessory spines. Pereopod 7 basis posterior margin straight, with posterior margin minutely castelloserrate, posteroventral corner; dactylus unguis anterior margin with accessory spines.Pleon. Pleonite/urosomite dorsal spine formula (7-7-7-5-6-2). Pleonites 1\u20133 with dorsal setae. Epimera 1\u20133 posteroventral margin without spines above posteroventral corner. Epimeron 3 posterior margin smooth, posteroventral corner with strongly produced acute. Urosomite 1 with spine at midline, no medial gape. Urosomite 2 with dorsal setae. Urosomite 3 with dorsal setae, with 2 dorsal spines. Uropod 3 inner ramus scale-like, much shorter than outer ramus; outer ramus much longer (more than 2\u00d7 length) than peduncle, 2-articulate. Telson with dorsal robust setae.Female. . Length, 7.4 mm. Gnathopod 1 coxa 4 anterodistal corner not produced, posteroventral corner notch present, anterior margin excavated. Gnathopod 2 equal, coxa subrectangular, palm crenulated, oblique with setae on margins. Pereopod 7 basis expanded, posterior margin slightly convex.This species is named after the type locality.Dulichiella pattaniensis sp. n., with pleonite/urosome formular of 7-7-7-5-6-2 has only Dulichiella cotesi, Dulichiella oahu, Dulichiella pacifica and Dulichiella tulear that share this characters. This new species can be distinguished from Dulichiella cotesi, Dulichiella oahu and Dulichiella tulear by having male gnathopod 2 (large) with 4 spines on distolateral crown while those three species have 3 spines. Dulichiella pattaniensis differs from Dulichiella pacifica in the following: the male large gnathopod 2 has a fourth spine on its distolateral crown that is not well developed vs. its well-developed fourth set of spines. The male gnathopod 1 has carpus longer than its propodus vs. a carpus subequal to its propodus. Pereopods 3-4 have a dactyli with 2 accessory spines vs. 3-4 dactyli with 1 accessory spine.Dulichiella appendiculata, Dulichiella cuvettensis, Dulichiella celestun, Dulichiella fresnellii, Dulichiella guinea, Dulichiella lecroyae, Dulichiella pacifica and Dulichiella takedai share this distinct character. Dulichiella pattaniensis can be distinguished from amphipods by having pleonite/urosome formular of 7-7-7-5-6-2 while Dulichiella appendiculata, Dulichiella cuvettensis, Dulichiella fresnellii, Dulichiella lecroye and Dulichiella pacifica pleonite/urosome formula 7-7-7-5-4-2, Dulichiella guinea 9-9-7-5-4-2 and Dulichiella celestun 9-9-9-5-6-2. Moreover, Dulichiella pattaniensis differs from Dulichiella cuvettensis, Dulichiella guinea and Dulichiella lecroyae by having 2 accessory spines on pereopods 3-4 dactyli vs. 1 accessory spine. A summary of these distinguishing characters are given in The new species also has four spines on the distolateral crown of the male gnathopod 2. Only 7 species, Ren & Andress, 2012http://species-id.net/wiki/Melita_latiflagella09\u00b018'39.5\"N, 99\u00b046'46.4\"E), 1 Feb 2012, Wongkamhaeng, K. PSUZC-CR-0191. .Lower Gulf of Thailand, Songkhla Lake ."}
+{"text": "Hyperoxia is frequently encountered in the intensive care unit (ICU) and during surgical procedures such as coronary artery bypass surgery (CABG). Higher oxygen concentrations intuitively provide a salutary oxygen reserve, but hyperoxia can induce adverse effects such as systemic vasoconstriction, reduction of cardiac output, increased microcirculatory heterogeneity and increased reactive oxygen species production. Previous studies in patients undergoing CABG surgery suggest reduced myocardial damage when avoiding extreme perioperative hyperoxia (>400 mmHg). Here, we compare moderate hyperoxia to near-physiological values.2s from moderate hyperoxia to near-physiological values decreases myocardial damage and organ injury.To investigate whether an oxygenation strategy towards lowering perioperative PaO2 target according to common practice (200-220 mmHg during cardiopulmonary bypass (CPB) and 130-150 mmHg in the ICU) versus a lower PaO2 target . The primary outcome was myocardial damage (CK-MB and Troponin-T), which was determined before surgery, at ICU admission and 2, 6 and 12 hours thereafter.In this single-center, open-label randomized-controlled trial, 50 patients scheduled for elective isolated CABG surgery were allocated to either a PaO2 during CPB was 220 mmHg, IQR (211-233) vs. 157 , respectively. In the ICU, weighted PaO2 was 107 (86-141) vs. 90 . Median maximum values of CK-MB were 25.8\u00b5g/L, IQR (20.3-32.6) vs. 24.9  and of Troponin-T 0.35 \u00b5g/L, IQR (0.30-0.46) vs. 0.42. Areas under the curve (AUC) of CK-MB  and 0.30\u00b5g/L/h (0.25-0.44) vs. 0.39  for Troponin-T. Cardiac Index, Systemic Vascular Resistance Index, and serum lactate levels (Lactatemax median 2 mmol/L IQR(1.4-2.6) vs. 2.2) were similar between groups throughout the ICU period.Baseline and surgery characteristics were not different between groups. The mean age of patients was 66 years (SD 8) vs. 68(6), respectively. Mean duration of CPB was 105 minutes (SD 24) vs. 108(28). Weighted PaOIn the present RCT, an oxygenation strategy towards near-physiological arterial oxygen tensions did not reduce myocardial damage in comparison to moderate hyperoxia. On the other hand, conservative oxygen administration did not lead to increased lactate levels.This investigation was supported by grants from ESICM NEXT Start-Up 2014 and ZonMW"}
+{"text": "To seek for possible differences in tryptase values during acute anaphylaxis induced by non-steroid anti-inflammatory drugs (NSAID) related to the suspected mechanism (selective vs non-selective reactions).Retrospective review of patients attending the Emergency Department (2009-2013) for a NSAID induced anaphylaxis and with a tryptase determination. Patients were split into 3 groups according to the allergy work-up ResultsSelective reactions (S) were defined by positive skin results and/or negative challenges with other NSAID; non-selective (NS) reactions were defined by the existence of symptoms with several NSAID or after positive challenge results. Those cases who did not comply with the previous conditions were non-conclusive (NC). We analysed: demographics, severity of the episode (Chi Square test), and rate of tryptase higher than 11.4 mcg/l (Chi Square test), median tryptase values and timing of sample obtention .86 patients were included: S group: 36 patients, NS group: 20 patients, and NC: 30 patients. Median ages were 48, 45 and 54 years, respectively. Severity of the episodes: S group: Mild: 22.2%, moderate: 30.6% and severe: 47.2%; NS group: Mild: 30%, moderate: 35% and severe: 35%; NC group: Mild: 16.7%, moderate: 26.7% and severe anaphylaxis: 56.7% (p=0.658). The most frequent eliciting drugs were metamizole , acetic derivatives  and propionic acid derivatives . The median of tryptase determinations were: S group: 13.4(2.75-98.6), NS: 12.35(3.85-53.6) and NC: 13.4mcg/l (2.82-57.1) (p=0.856). Median duration of symptoms: S group: 1.5 (0.33-12), NS: 1.75 (0.5-4); NC: 1.5 hours (0.5-6h) (p=0.582).Although there have been reports of differences in tryptase releases during anaphylaxis related to different allergens, we have found no differences among the different patterns of NSAID reactions that may help us to avoid future challenges for diagnostic confirmation."}
+{"text": "Rictor to impair mTORC2 activity in mouse mammary epithelia, which decreased Akt phosphorylation, ductal length, secondary branching, cell motility, and cell survival. These effects were recapitulated with a pharmacological dual inhibitor of mTORC1/mTORC2, but not upon genetic disruption of mTORC1 function via Raptor deletion. Surprisingly, Akt re-activation was not sufficient to rescue cell survival or invasion, and modestly increased branching of mTORC2-impaired mammary epithelial cells (MECs) in culture and in vivo. However, another mTORC2 substrate, protein kinase C (PKC)-alpha, fully rescued mTORC2-impaired MEC branching, invasion, and survival, as well as branching morphogenesis in vivo. PKC-alpha-mediated signaling through the small GTPase Rac1 was necessary for mTORC2-dependent mammary epithelial development during puberty, revealing a novel role for Rictor/mTORC2 in MEC survival and motility during branching morphogenesis through a PKC-alpha/Rac1-dependent mechanism.Akt phosphorylation is a major driver of cell survival, motility, and proliferation in development and disease, causing increased interest in upstream regulators of Akt like mTOR complex 2 (mTORC2). We used genetic disruption of  The protein kinase mTOR is frequently activated in breast cancers, where it enhances cancer cell growth, survival, and metastastic spread to distant organs. Thus, mTOR is an attractive, clinically relevant molecular target for drugs designed to treat metastatic breast cancers. However, mTOR exists in two distinct complexes, mTORC1 and mTORC2, and the relative roles of each complex have not been elucidated. Moreover, as pathways that regulate normal tissue growth and development are often highjacked to promote cancer, understanding mTOR function in normal mammary epithelial development will likely provide insight into its role in tumor progression. In this study, we assessed the role of mTORC1 and mTORC2 complexes in normal mammary epithelial cell branching, survival, and invasion. Interestingly, while mTORC1 was not required for branching, survival and invasion of mammary epithelial cells, mTORC2 was necessary for these processes in both mouse and human models. Furthermore, we found that mTORC2 exerts its effects primarily through downstream activation of a PKC-alpha-Rac1 signaling axis rather than the more well-studied Akt signaling pathway. Our studies identify a novel role for the mTORC2 complex in mammary morphogenesis, including cell survival and motility, which are relevant to breast cancer progression. Post-natal mammary epithelial morphogenesis is a complex process during which an extensively branched ductal network develops from a rudimentary epithelial bud . Branchimammalian target of rapamycin (mTOR) regulates cellular metabolism, protein and lipid synthesis, cell survival, and cytoskeletal organization, processes that are required for proper mammary morphogenesis. mTOR regulates these processes through phosphorylation of its target substrates, including translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1), p70S6 kinase (S6K), Akt, SGK1, and protein kinase C-alpha  . For some experiments, cells were maintained for 24 hour in Starvation Media [Growth Media without serum or EGF] prior to stimulation and/or analysis. PKC-alpha inhibitor GO6976  and adenoviral particles  were purchased. Cells were incubated with adenoviral particles (5 X 108 particle forming units/ml) for 3\u20135 hours at 37\u00b0C and cells were allowed to recover for 48 hours prior to experimental analysis.MCF10A and MCF10A Rictor2HPO4, 3.5 millimolar NaH2PO4, 7.7 millimolar NaN3, 0.1% bovine serum albumin, 0.2% Triton-X 100, 0.05% Tween-20] and stained with rabbit anti-pan-cytokeratin  and AF621-goat anti-rabbit (1:100), counterstained with TO-PRO-3 Iodide (Invitrogen), and imaged using the Vanderbilt Cell Imaging Shared Resource Zeiss LSM 510 confocal microscope and LSM Image Browser software. For E-cadherin staining, the primary antibody used was anti-E-cadherin (BD Transduction Laboratories) and visualized with anti-mouse Alexa 594 secondary antibody . Confocal images of 3D structures were visualized using an LSM 510 META inverted confocal microscope with a 20X/0.75 plan apochromat objective.Matrigel-emdedded organoids cultured on coverslips were fixed 8 minutes in 1:1 methanol:acetone at -20\u00b0C, permeabilized in 0.5% Triton-X 100/PBS for 10 min, blocked , sonicated 10 seconds, and cleared by centrifugation at 4\u00b0C, 13,000 x g for 5 min. Protein concentration was determined using BCA (Pierce). Proteins were separated by SDS-PAGE, transferred to nitrocellulose membranes, blocked in 3% gelatin in TBS-T [Tris-buffered saline, 0.1% Tween-20), incubated in primary antibody overnight and in HRP-conjugated anti-rabbit or anti-mouse for 1 hour, and developed using ECL substrate (Pierce). Antibodies used: alpha-actin ; AKT and S473 P-Akt ; S6 and P-S6 ; Rictor ; Raptor ; Rab11 ; PKC-alpha and T638/641 P-PKC-alpha ; Rac . GST-Pak-PBD effector pulldown assays were performed using reagents from Millipore as per manufacturer\u2019s protocol.Cells and tissues were homogenized in ice-cold lysis buffer [50 millimolar Tris pH 7.4, 100 millimolar NaF, 120 millimolar NaCl, 0.5% NP-40, 100 micromolar NaIn situ TUNEL analysis was performed on paraffin-embedded sections using the ApopTag kit . IHC on paraffin-embedded sections was performed as described previously [Mammary glands were whole-mounted on slides, cleared of adipose, and stained with hematoxylin as described previously . Sectioneviously  using: KMethanol-fixed PMECs were probed 1 hour with GST-PBD (Millipore) diluted 1:50 in PBS. GST (lacking PBD) was used as a negative control. Samples were washed then probed with AF488-conjugated anti-GST (1:100), stained with DAPI or AF621-phalloidin, and mounted.5) were added to upper chambers of Matrigel-coated transwells in starvation medium and incubated 5 hours to score migration in response to 10% serum-containing medium in the lower chamber. Filters were swabbed and stained with 0.1% crystal violet, [MECs  has been accredited by AAALAC International since 1967 (File #000020). The AAALAC Council on Accreditation's most recent review of VU's program was done in 2011 and resulted in \"Continued Full Accreditation.\u201d Isofluorane was used for anesthesia, as well as euthanasia. For human euthanasia, cervical dislocation was used following isofluorane overdose.S1 FigA. Hematoxylin stained whole mount preparations from 6 week old RictorWT mice and RictorMGKO mice. High magnification panels show irregular ductal tracts and increased staining density in RictorMGKO samples, indicative of multiple cell layers. Data are a representation of 11 independent animals/genotype. B. Hematoxylin stained whole mount preparations from 10 week old wild-type RictorWT mice and RictorMGKO mice show sustained length defects in the ductal tracts of mammary glands lacking Rictor. C. IHC for Ki67 or TUNEL in TEBs from 6 week old wild-type RictorWT mice and RictorMGKO mice. Average percent Ki67 and TUNEL+ nuclei (\u00b1 S.D.) was determined. D. Confocal analysis of primary organoids (PMOs) stained for E-cadherin (red) revealed multiple cell layers in acinar structures and poor lumen formation in Rictor-deficient (RictorFL/FL organoids infected with Ad.Cre) PMOs relative to control PMOs (RictorFL/FL organoids infected with Ad.LacZ), consistent with the phenotype in RictorMGKO epithelium in vivo.(TIF)Click here for additional data file.S2 FigRictorFL/FL mice and infected with Ad.GFP or Ad.Cre. A. Fluorescent imaging of organoids 24 hours post-infection. B. Confocal analysis of WT organoids infected with Ad.GFP. C. Organoids were fixed 10 days post-infection and subjected to immunofluorescent staining using a pan-cytokeratin antibody (red) to confirm epithelial identity and ToPro-3 nuclear marker (blue). While Ad.LacZ-infected RictorFL/FL organoids formed hollow lumens surrounded by an organized epithelial layer, Ad.Cre-infected RictorFL/FL organoids remained rounded and disorganized. Data are a representation of 3 independent organoid isolates. D. MCF10A parental or MCF10A-RictorZFN cells were assessed for invasion through Matrigel-coated transwell filters. The cells remaining in the upper chamber after 24 hours were fixed and stained with either DAPI (blue) or Annexin V-FITC (green). The average percent viable cells (\u00b1 S.D.) left in the upper chamber was quantitated using Image J, Student\u2019s T-test.Primary organoids (PMOs) were isolated from (TIF)Click here for additional data file.S3 FigA-C. PMECs and organoids from RictorFL/FL mice were coinfected with Ad.Cre and either Ad.LacZ or Ad.AktDD. A. Western analysis of PMEC lysates. B. Organoids photographed after 10 days in Matrigel culture. Average number of branches/organoid \u00b1 S.D. (right panel) is shown. N = 7 independent organoid isolates, analyzed in triplicate. Midline values indicate average, whiskers indicate S.D., Student\u2019s T-test. C. Colony size of organoids (\u00b1 S.D.) measured in pixel area. N = 7 independent organoid isolates, analyzed in triplicate, Student\u2019s T-test.(TIF)Click here for additional data file.S4 FigA. Immunofluorescent detection of P-PKC-alpha in mammary gland sections from 6 week old virgin RictorWT and RictorMGKO mice. B.In situ detection of GTP-only control (no PBD) via IF detection of GST-PBD  on mammary gland sections from 6-week old mice. C.WT PMECs cultured \u00b1 Rac inhibitor or RictorFL/FL PMECs infected with Ad.LacZ or Ad.Cre were assessed for GTP-bound Rac via IF detection of GST-PBD . D.RictorFL/FL PMECs infected with Ad.Cre or Ad.LacZ were probed with GST-PBD (green) and counterstained with phalloidin (red). Representative images are shown. E.RictorFL/FL PMECs infected with Ad.Cre or Ad.LacZ (\u00b1Ad.caRac1) were analyzed for TUNEL. F. Whole mount analysis of endogenous mammary glands harvested from WT recipient mice.(TIF)Click here for additional data file.S5 FigWT PMECs infected with Ad.LacZ or Ad.caRac1 were grown to confluence (3\u20135 days after infection) then wounded using a P200 pipette tip in the presence of DMSO or rapamycin. Monolayers were imaged after 24 hours and total wounded area (arbitrary units) remaining was measured at 24 hours after wounding. Values shown are the average \u00b1 S.D. N = 6 per time point.(TIF)Click here for additional data file.S6 FigA. Western analysis of whole mammary gland lysates from 10 week old mice. B. IF for CK8, CK14 and ZO-1 in mammary gland sections. C. IHC for Ki67 or TUNEL in TEBs from 6 week old wild-type RaptorWT mice and RaptorMGKO mice. Average percent Ki67+ and TUNEL+ nuclei (\u00b1 S.D.) was determined, Student\u2019s T-test.(TIF)Click here for additional data file."}
+{"text": "Cichorium intybus L. (chicory) in acute liver injury. Pathological observation, reactive oxygen species (ROS) detection and measurements of biochemical indexes on mouse models proved hepatic protective effect of Cichorium intybus L. Identification of active compounds in Cichorium intybus L. was executed through several methods including ultra performance liquid chromatography/time of flight mass spectrometry (UPLC-TOF-MS). Similarity ensemble approach (SEA) docking, molecular modeling, molecular docking, and molecular dynamics (MD) simulation were applied in this study to explore possible mechanisms of the hepato-protective potential of Cichorium intybus L. We then analyzed the chemical composition of Cichorium intybus L., and found their key targets. Furthermore, in vitro cytological examination and western blot were used for validating the efficacy of the selected compounds. In silico analysis and western blot together demonstrated that selected compound 10 in Cichorium intybus L. targeted Akt-1 in hepatocytes. Besides, compound 13 targeted both caspase-1 and Akt-1. These small compounds may ameliorate liver injury by acting on their targets, which are related to apoptosis or autophagy. The conclusions above may shed light on the complex molecular mechanisms of Cichorium intybus L. acting on hepatocytes and ameliorating liver injury.This study aimed at investigating the possible mechanisms of hepatic protective activity of  Cichorium intybus L.  is a perennial herb from the Asteraceae family with both alimentary use and medicinal use. It is 40\u2013100 cm in height, usually with bright blue flowers, rarely white or pink. It has been reported that fresh chicory typically contains 68% inulin, 14% sucrose, 5% cellulose, 6% protein, 4% ash, and 3% other compounds, while dried chicory contains approximately 98% inulin and 2% other compounds + ; 1H-NMR  \u03b4: 7.07 , 6.72 , 3.66 , 3.52 ; 13C-NMR  \u03b4: 173.1 (C-8), 156.2 (C-4), 129.9 , 124.5 (C-1), 114.9 , 50.9 (C-1\u02b9), 39.5 (C-7).Compound 13: colorless acicular crystal (MeOH); HR-ESI-TOF-MS 153.0549 [M + H]+ ; 1H-NMR \u03b4: 7.00 , 6.67 , 3.46 , 9.27 ; 13C-NMR  \u03b4: 176.1 (C-8), 157.4 (C-4), 131.3 , 126.8 (C-1), 116.2 , 41.1 (C-7).Compound http://sea.bkslab.org/) [http://www.geneontology.org) and KEGG Orthology (KO) (http://www.genome.jp/kegg/ko.html) [Target proteins of the small molecule compounds were obtained from the similarity ensemble approach (SEA) (ab.org/) . An assiko.html) ,23,24. Fhttp://www.pdb.org/pdb/home/home.do). Molecular docking was performed using the UCSF DOCK6.5 program , which utilized DOCK algorithm to address rigid body docking by superimposing the ligand on to a negative image of the binding pocket [We selected 2 effective small molecule compounds and 2 proteins as receptors. Before the docking, we downloaded initial three-dimensional geometric co-ordinates of the X-ray crystal structures of compounds and proteins from the Protein Data Bank (PDB)  simulations. MD simulation of the complex was carried out with the GROMACS 4.5.4   package 3, and a protease inhibitor cocktail . Following one freeze\u2013thaw cycle, the cell lysates were centrifuged at 10,000\u00d7 g at 4 \u00b0C for 20 min and boiled in sample buffer for 5 min. The proteins were then subjected to SDS\u2013PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) and transferred to a PVDF (polyvinylidene fluoride) membrane  using a semi-dry electroblotting system. After blocking with 5% skim milk in PBS, the membranes were incubated with primary antibodies (1:1000 dilution) at 4 \u00b0C overnight. The membranes were then washed with PBS containing 0.05% Tween 20 and incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies (1:5000 dilution) at room temperature for 1 h. After washing, the membranes were soaked in ECL solution  for 1 min, according to the manufacturer\u2019s instructions, and exposed to film . Other materials used for protein analyses were purchased from Sigma-Aldrich  [The expression levels of the selected target proteins (Akt-1 and caspase-1) in liver tissues were measured by applying western blot. Liver tissues were lysed in a buffer containing 1% Triton X-100, 50 mM Tris (pH 7.5), 10 mM EDTA, 0.02% NaNMO, USA) ,31,32.t-test. p < 0.05 was considered statistically significant.All the presented data and results were confirmed in at least three independent experiments. The data are expressed as means \u00b1 S.D. Statistical comparisons were made by One-way ANOVA and Student\u2019s Cichorium intybus L. and active compounds in Cichorium intybus L. The active compounds ameliorate liver injury by acting on Akt-1 and caspase-1, which are related to apoptosis or autophagy. In summary, our results may shed light on the complex molecular mechanisms of Cichorium intybus L. acting on hepatocytes and ameliorating liver injury.This study demonstrates the hepatic protective effect of"}
+{"text": "The oral antihyperglycemic agent metformin was associated with favorable outcome and smaller myocardial infarct size in patients with diabetes undergoing percutaneous coronary interventions (PCI) for ST-segment elevation myocardial infarction (STEMI) , 2. HoweTo determine the effect of chronic metformin treatment on cardiovascular morbidity and mortality in patients with diabetes presenting with STEMI subsequently undergoing PCI.From January 2004 until June 2013, all consecutive critically ill patients undergoing primary PCI for STEMI at the University Medical Center Groningen were included in a registry and 1-year follow-up was obtained. Our primary endpoint consisted of the composite endpoint of myocardial infarction, target vessel and target lesion revascularization, and all-cause mortality (MACE). The secondary endpoint, myocardial infarction size, was estimated using peak levels of creatine kinase (CK), the myocardial band of CK (CK-MB), troponin T, and high-sensitive troponin T (hs-troponin T). The effect of metformin on myocardial infarct size from the 2004-2010 cohort has been reported previously [In total, 4776 consecutive patients underwent primary PCI for STEMI, 719 (15%) diabetic patients were included in the final analysis and 215 (30%) patients used metformin at admission. MACE and mortality rates were 21% and 12% for patients with diabetes, 23% and 19% for metformin patients, 21% and 15% for patients on sulfonylurea, and 30% and 20% for patients on insulin, respectively. Metformin was not associated with reduced risk for MACE (adjusted hazard ratio (aHR): 1.19  0.78-1.81), P = 0.42) or survival benefit (aHR: 0.23 (CI95% 0.80-2.51), P = 0.23) compared to diabetic patients not using metformin. Insulin use was an independent predictor for MACE (aHR 1.73 (CI95% 1.13-2.65), P = 0.01) and all-cause mortality (aHR 1.81 (CI95% 1.03-3.21), P = 0.04). Baseline levels of CK, CK-MB, and hs-troponin T were comparable between both groups. Median (interquartile range) peak levels of CK, CK-MB, and hs-Troponin T were all non-significant lower in the metformin group (table Chronic metformin use in patients presenting with STEMI was associated with smaller infarct-size and not with lower MACE and mortality rates, as compared to other patients with diabetes."}
+{"text": "To evaluate whole-body diffusion weighted magnetic resonance imaging (WB-DWI) for staging of women with cancer during pregnancy.Twenty patients diagnosed with cancer during pregnancy underwent WB-DWI additional to conventional imaging in this prospective single centre study. Reproducibility of WB-DWI between 2 readers was evaluated using Cohen\u2019s \u03ba statistics and accuracy was compared to conventional imaging for assessing primary tumour site, nodal metastases and visceral metastases. Histopathology after surgery or biopsy was the primary reference standard.Ten patients had breast cancer, 3 lymphoma, 2 cervical uterine cancer, 1 ovarian borderline tumour, 2 colon cancer, 1 lung cancer and 1 a conjunctival tumour. The WB-DWI readers showed very good agreement for lesion detection, \u03ba = 0.94. With WB-DWI, reader 1 detected 38 of 41 malignant lesions, reader 2 thirty-nine lesions and conventional imaging 27. WB-DWI showed sensitivity of 95% (95% CI: 74-99) for both readers and specificity up to 99% (95% CI: 76-99) compared to 50% sensitivity (95% CI: 28-72) with 100% (95% CI: 97-100) specificity for conventional imaging. For staging distant metastases, WB-DWI sensitivities were 66.7% (95% CI: 13-98) and 100% (95% CI: 40-100) respectively for reader 1 and 2 with specificities of 94.1% (95% CI: 69-99) and 100% (95% CI: 40-100) compared to sensitivity of 33.3% (95% CI: 1.7-87) and specificity of 100% (95% CI: 77-100) for conventional imaging.WB-DWI is feasible for single-step non-invasive imaging based cancer staging during pregnancy showing additional value to conventional imaging procedures for detecting distant and nodal metastases."}
+{"text": "Protochlamydia sp. strain HS-T3, an environmental chlamydia. This bacterium is an amoebal endosymbiont, found in Acanthamoeba isolated from a hot spring in Japan. Strain HS-T3 readily grew in mammalian cells at 37\u00b0C, a characteristic not previously reported for environmental chlamydiae.Here, we report the draft genome sequence of high-temperature-adapted  Chlamydiae is a bacterial phylum and class comprising obligate intracellular bacteria. Two distinct lineages exist, pathogenic and environmental chlamydiae, and this divergence likely occurred 0.7 to 1.4 billion\u00a0years ago   , sexuallindness) , and comindness) . In conteduction . Most eneduction . Howevere, Japan .Protochlamydia HS-T3 was obtained using an Illumina HiSeq 2000 sequencer , with sequencing runs for paired-end sequences. The bacterial DNA libraries were prepared using a TruSeq DNA sample kit (Illumina). The genome was assembled into 34 contigs, ranging in size from 189 to 363,061\u00a0bp, using de novo sequence assembler software  (http://rast.nmpdr.org/) (http://www.genome.jp/kegg/) (The draft genome of MBL-EBI) . Gene prdr.org/) , and funp/kegg/) . The seqProtochlamydia sp. HS-T3 was 2,308,589\u00a0bp (2.3\u00a0Mb) with a G+C content of 38.74% and 950-fold genome coverage. The genome sequence contained 2,259 coding sequences, 40 tRNAs, and 2 ribosomal RNAs. Neighbor analysis using RAST revealed that the closest neighbor of Protochlamydia HS-T3 was Protochlamydia amoebophila UWE25 (NC_005861.1)  and type III general secretion gene clusters but lacked the genes encoding the type IV secretion system. In addition, 1,139 hypothetical proteins and 8 unknown proteins were identified.The draft genome sequence of 5 NC_00581.1 (6), 05861.1) . SimilarProtochlamydia HS-T3 has been deposited at DDBJ/EMBL/GenBank under the accession numbers BBPT01000001 through BBPT01000034 (34 entries). The version described in this paper is the first version.The draft genome sequence of"}
+{"text": "Previous studies have described improvements on lipid parameters when switching from other antiretroviral drugs to tenofovir (TDF) and impairments in lipid profile when discontinuing TDF. \u20133It is This was a randomized, crossover, double-blind, placebo-controlled clinical trial (NCT 01458977). Subjects with HIV-1 RNA <50 copies/mL during at least 6 months on stable DRV/r (800/100 mg QD) or LPV/r (400/100 mg BID) monotherapy, with confirmed fasting total cholesterol \u2265200 or LDL-cholesterol \u2265130 mg/dL and not taking lipid-lowering drugs were randomized to (A) adding TDF/FTCduring 12 weeks followed by 24 weeks without TDF/FTC, or (B) continuing without TDF/FTC for 12 weeks, adding TDF/FTC for 12 weeks and then withdrawing TDF/FTC for 12 additional weeks. Randomization was stratified by DRV/r or LPV/r use at study entry. All subjects received a specific dietary counselling. Primary endpoints were changes in median fasting total, LDL and HDL-cholesterol 12 weeks after TDF/FTC addition. Analyses were performed by ITT.46 subjects with a median age of 43 (40\u201348) years were enrolled in the study: 70% were male, 56% received DRV/r and 44% LPV/r. One subject withdrew the study voluntarily at week 4 and another one interrupted due to diarrhoea at week 24. Treatment with TDF/FTC decreased total, LDL and HDL-cholesterol from 235.9 to 204.9 (p<0.001), 154.7 to 127.6 (p<0.001) and 50.3 to 44.5 mg/dL (p<0.001), respectively. In comparison, total, LDL and HDL-cholesterol levels remained stable during placebo exposure. Week 12 total cholesterol (p<0.001), LDL-cholesterol (p<0.001) and HDL-cholesterol (p=0.011) levels were significantly lower in TDF/FTC versus placebo. Treatment with TDF/FTC reduced the fraction of subjects with abnormal fasting total-cholesterol (\u2265200 mg/dL) from 86.7% to 56.8% (p=0.001) and LDL-cholesterol (\u2265130 mg/dL) from 87.8% to 43.9% (p<0.001), which was not observed with placebo. There were no virological failures, and CD4 and triglyceride levels remained stable regardless of exposure.Coformulated TDF/FTC has an intrinsic lipid-lowering effect, likely attributable to TDF."}
+{"text": "In heart failure (HF) alveolar-capillary membrane is abnormal. Surfactant-derived proteins (SPs) and plasma receptor for advanced-glycation-end-products (RAGE) have been proposed as lung damage markers.Eighty-nine chronic HF and 17 healthy subjects were evaluated by echocardiography, blood parameters, carbon monoxide lung diffusion (DLCO) and cardiopulmonary exercise test. We measured immature SP-B, mature SP-B, SP-A, SP-D and RAGE plasma levels.th\u201325th interquartile range) Vs. 11.1 (6.4), p<0.01; SP-A, 29.6 (20.1) Vs. 18.3 (13.5), p\u200a=\u200a0.01; SP-D: 125 (90) Vs. 78 (58), p<0.01]. Immature SP-B, SP-A, SP-D and RAGE values were related to DLCO, peak oxygen consumption, ventilatory efficiency, and brain natriuretic peptide (BNP), whereas plasma mature SP-B was not. The DLCO Vs. immature SP-B correlation was the strongest one. At multivariate analysis, RAGE was associated to age and creatinine, SP-A to DLCO and BNP, SP-D to BNP, mature SP-B to DLCO and creatinine, and immature SP-B only but strongly to DLCO.Immature SP-B (arbitrary units), mature SP-A (ng/ml) and SP-D (ng/ml), but not mature SP-B (ng/ml) and RAGE (pg/ml) levels, were higher in HF than in controls [immature SP-B: 15.6 (13.1, 75Immature SP-B is the most reliable biological marker of alveolar-capillary membrane function in HF. Impairment of respiratory function is part of the chronic heart failure (HF) syndrome, being both lung mechanics and gas exchange altered. In all cases, RAGE and SPs have been linked to alveolar capillary membrane damage, but a comparative evaluation among RAGE and the different SPs available as markers of alveolar capillary membrane damage in HF has not been performed yet. We therefore analyzed the correlation between lung diffusion abnormalities, in terms of carbon monoxide total lung diffusion (DLCO), and RAGE and several SPs in a population of chronic stable HF patients and healthy controls, aiming to identify the ones that better correlates with gas diffusion.We studied HF patients in stable clinical conditions and healthy subjects. Patients belong to a group of individuals regularly followed up at our HF unit and were randomly recruited between February 2012 and November 2012, whereas healthy subjects were hospital staff employees or their relatives with gender and age similar to the HF patients.Study inclusion criteria for HF patients were New York Heart Association functional classes (NYHA) I to IV, echocardiographic evidence of reduced left ventricular systolic function , optimized and individually tailored drug treatment, stable clinical conditions for at least 2 months, capability/willingness to perform a maximal or nearly maximal cardiopulmonary exercise test (CPET). Patients were excluded if they had severe obstructive and/or restrictive lung disease, anemia (hemoglobin <11 g/dL), history and/or documentation of pulmonary embolism, primary valvular heart disease, pulmonary arterial hypertension, pericardial disease, exercise-induced angina, ST changes, severe arrhythmias and significant cerebrovascular, renal, hepatic and hematological disease.At enrolment, demographical and clinical data were collected. Before CPET, in both HF and healthy subjects echocardiographic evaluation, natriuretic peptide B (BNP) and blood samples measurements, standard pulmonary function tests, including DLCO, were performed.1) and lung vital capacity (VC) , and were standardized as percentages of predicted normal values. 1/VC lower than 60% was considered indicative of a severe obstructive ventilatory defect BNP test was performed on UniCel-DxI-800 Access immunoassay  with Triage-Biosite reagent . Forced expiratory volume in 1 second . 2), carbon dioxide production (VCO2) and ventilation (VE) were measured breath by breath and reported as average over 20 seconds. All tests were executed and evaluated by 2 expert readers blinded to plasma SPs and RAGE values. The anaerobic threshold (AT) was identified by a standard techniques. 2 was calculated as the slope of the linear relationship between VE and VCO2 from 1 minute after the beginning of loaded exercise to the end of the isocapnic buffering period.A CPET familiarization test was always performed. g for 10 minutes at 4\u00b0C, divided into aliquots and frozen at \u221280\u00b0C until assayed.Fresh blood (5 mL) was drawn into Vacutainer tubes containing citrate 0.129 mol/L as an anticoagulant. Plasma was immediately prepared by means of centrifugation at 1,500\u00d7versus the volume of the total proteins loaded and stained with MemCode. The values were also normalized versus the band volume of pooled plasma, loaded as control on each gel, and they are expressed as arbitrary units (AU). Inter-assay coefficient of variation was 12.1\u00b12.9%.The analysis of the immature forms of SP-B, which is detectable in the 3 predominant forms. with molecular mass ranging from 17 to 42 kDa, was performed by Western blotting on plasma samples, as previously described. The quantitative analysis of the levels of the mature form (8 kDa) of SP-B was performed by an ELISA purchased from Uscn Life Science Inc. . Inter-assay and intra-assay coefficient of variation were 11.6\u00b12.1% and 7.9\u00b11.5%, respectively.Plasma levels of SP-A, SP-D and RAGE were determined using commercially available ELISA kits . Measurements were performed in duplicate and the results were averaged. The intra-assay and inter-assay coefficients of variation for SP-A were <5% and <10%, Limit of Detection (LOD) is 0.16 ng/ml, and cut off level is 1 ng/ml. For SP-D the intra-assay and inter-assay coefficients of variation were <3% and <4%, LOD is 0.01 ng/ml, and cut off level is 1.56 ng/ml. The intra-assay and inter-assay coefficients of variation for RAGE were <6% and <8%, respectively, minimal detectable dose is 4.12 pg/ml, and the cut off level is 78 pg/ml.th percentile\u201325th percentile). Categorical variables were compared with \u03c72 test. Unpaired t-test or non-parametric Mann-Whitney test were used when appropriate for between-group comparison. One-way analysis of variance [ANOVA] followed by Kruskal Wallis test were used to compare data for the non-normally distributed variables.Unless otherwise indicated all data are expressed as means \u00b1 SD. Data with skewed distribution are given as median and interquartile range (752 (\u226515 mL/kg/min and <15 mL/kg/min), median VE vs VCO2 slope (\u226430 and >30), median LVEF (\u226436% and >36%) and median BNP (\u2264160 pg/ml and >160 pg/ml). Finally, given that it is well-reported a possible confounding role of smoke on the alveolar-capillary membrane 2 value and VE/VCO2 slope. To avoid distorted estimates of the effect, Pearson\u2019s correlations were limited to the population with HF. A Bootstrap method, Besides between healthy subjects and HF patients, statistical analysis was also performed by subdividing HF patients according to NYHA class (I\u2013II vs III\u2013IV), median DLCO (\u226580% vs <80%), median peak VO2) and 17 healthy subjects  . Active smokers were 30/89 and 7/17 in the HF and control group, respectively. HF etiology was cardiomyopathy due to ischemic coronary disease in 50 cases and not-ischemic in 39 cases. Among HF subjects, 44 were in NYHA class I\u2013II and 45 in class III\u2013IV. The LVEF in HF patients was 36\u00b19%; the median BNP was 160 pg/mL . Treatment in patients with HF included angiotensin-converting enzyme inhibitors in 53 cases (60%), AT1 blockers in 21 cases (24%), \u03b2-blockers in 81 cases (91%), diuretics in 59 cases (66%), antialdosteronic drugs in 43 cases (48%), amiodarone in 37 cases (42%), digoxin in 4 cases (5%), antiplatelet in 59 cases (66%), and anticoagulant in 24 cases (27%).A total of 106 subjects were evaluated: 89 patients with HF  levels. Only immature SP-B and mature SP-A were higher in patients with most severe HF considering all the above reported HF severity criteria . VO2ATKG-Oxygen uptake at anaerobic threshold (mL/min/Kg). VEVCO2slope-Ventilatory efficiency. NYHA-New York Heart Association functional class. ISCHAEMIC_ETIOLOGY-. ICD-implantable cardioverter-defibrillator . CRT-cardiac resynchronization therapy . BBLOCK-Betablocker . ACEI-Ace inhibitor . AT1 BLOCK-Angiotensin converting enzyme inhibitors . ANTIALDOSTERONIC-angiotensin type 1 receptor blockers . DIURETIC-. AMIODARONE-. DIGOXIN-. ANTICOAGULANT-. ANTIPLATELET-.PAZIENT ID-Identification number. HF-Heart failure patient . SPBIMMATURE-Immature SPB (AU). SPB-Surfactant protein B (ng/mL). SPA-Surfactant protein A (ng/mL). SPD-Surfactant protein D (ng/mL). RAGE-Plasma receptor for advanced glycation end products (pg/mL). AGE-Age (years). GENDER-. IPERTENSION-. DMII-Diabetes type II . weight-(Kg). height-(cm). BMI-Body mass index (kg/m2). FUMO-smoke . LVEF-Left ventricular ejection fraction (%). Vtd-TeleDiastolic Volume (mL). Vts-TeleSystolic Volume (mL). IM-Mitral insufficiency . severityIM-Mitral insufficiency severity (1\u20134). GradoIT-Tricuspidalic insufficiency severity. TAPSE-Tricuspid annular plane systolic excursion (mm). PAPs-Systolic Pulmonary artery Pressure (mmHg). Hb-Hemoglobin (mg/dL). Crea-Creatinine (mg/dL). Na-Sodium (mmol/L). K-Potassium (mmol/L). Uric acid-(mg/dL). BNP-brain natriuretic petide (pg/mL). VC-Vital Capacity (L). VC_P-Vital Capacity (% of predicted). FVC-forced vital capacity (L). FVC_P-forced vital capacity (%). FEV1-forced expiratory volume in 1 second (L). FEV1_A-forced expiratory volume in 1 second (L). FEV1FVC-FEV1/FVC ratio. Dladj_ass-carbon monoxide lung diffusing capacity (mL/mm Hg/min). Dladj_perc-carbon monoxide lung diffusing capacity (% of predicted). SBPBASAL-Systolic blood pressure (mmHg). DBPBASAL-Diastolic blood pressure (mmHg). HRBASAL-Heart Rate (bpm). LOAD-Load at exercise (watt). QRPICk-Respiratory exchange ratio at peak. sbpPEAK-Systolic blood pressure at peak exercise (mmHg). DBPPEAK-Diastolic blood pressure at peak exercise (mmHg). VO(XLSX)Click here for additional data file."}
+{"text": "E. coli (ESBLEc) increased.We conducted an annually bloodstream infection (BSI) survey into hospitals overlapping the Centre France region (2.6 million). Since 2005, the incidence of BSIs associated with ESBL-producing To improve the understanding of the pathway and the determination of the risk factors of ESBLEc-BSIs.For each BSI, were reported patient age, sex, recent hospitalization, living in nursing home, recent antibiotherapy, urinary catheterization, BSI source, death within 7days of diagnosis.BSI isolates were studied: antimicrobial susceptibility, determination of molecular mechanism associated with ESBL-production, genetic diversity of ESBLEc (MLST).E. coli BSI were identified, including 31 ESBLEc . Incidence of community acquired(CA)- and healthcare associated(HCA)-BSI were 0.47/100,000 and 0.040/1,000 PDs, respectively.During the survey , 443 For ESBLEc-CA-BSIs, male/female ratio was 1.4, median age 80, urinary BSI source in 50% of cases, recent antibiotherapy in 33 %. Most ESBLEc were resistant to fluoroquinolones (67%), SXT/TMP (67%). High genetic diversity (8 STs including 4 ST131).For ESBLEc-HCA-BSI, male/female ratio was 0.9, median age 75, urinary BSI source in 63% of cases (recent catheterization in 1/2), recent antibiotherapy in 58%. Most ESBLEC were resistant to fluoroquinolones (79%), SXT/TMP (63%). Low genetic diversity (9 STs including 7 ST131).Among BSI, ESBLEc-BSI were associated with healthcare (p=0.004), long-stay unit (p=0.018), recent antibiotherapy (p=0.002). ESBLEc were associated with resistance to fluoroquinolones, SXT/TMP and genta./tobramycine (p<0.001).Among ESBLEc-BSI, clinical determinants and BSI characteristics similar whatever the clonal group excepted for ST131 associated with long-stay unit (p=0.042).Among ST131-BSI, clinical determinants and BSI characteristics similar for ESBLEc and non ESBLEc excepted median age higher in ESBLEc (80/67).Recent antibiotherapy (and easy spread into long-stay units for ST131): likely the major risk factor for ESBLEc BSI.None declared."}
+{"text": "KRAS wild-type and mutant. Prognostic relevance of KRAS genotype was evaluated in patients unfit for FIr-B/FOx, treated with conventional medical treatments. Consecutive MCRC patients not eligible for FIr-B/FOx regimen due to age (\u226575 years) and/or comorbidities were treated with tailored conventional first-line treatments. KRAS codon 12/13 mutations were screened by direct sequencing. Activity and efficacy were evaluated and compared according to medical treatments, age (non-elderly and elderly \u226565 years), comorbidity stage , metastatic extension (liver-limited and other/multiple metastatic), and KRAS genotype, using log-rank. Selected first line treatments were medical in 37 patients (92.5%), and surgical in 3 patients (7.5%). Medical treatment regimens: triplet, 18 (45%); doublet, 15 (37.5%); mono-therapy, 4 (10%). At median follow-up of 8 months, objective response rate (ORR) was 37%, median progression-free survival (PFS) 7 months, liver metastasectomies 8% (liver-limited disease 37.5%), median overall survival (OS) 13 months. Triplet regimens failed to significantly affect clinical outcome, compared to doublet. According to KRAS genotype, ORR, PFS and OS were, respectively: wild-type 50%, 8 months, 13 months; mutant 25%, 6 months, 9 months. KRAS genotype wild-type compared to mutant significantly affected PFS, while not OS. KRAS c.35 G>A mutation (G12D) significantly affected worse PFS and OS compared to wild-type and/or other mutations. KRAS genotype, specifically the c.35 G>A KRAS mutation, may indicate poor prognosis in MCRC patients unfit for intensive medical treatments.First-line triplet chemotherapy plus bevacizumab (FIr-B/FOx) can improve efficacy of metastatic colorectal cancer (MCRC),  KRAS genotype  faces with different options and lines of treatment according to patients\u2019 fitness , extension of metastatic disease [liver-limited (L-L) or other/multiple metastatic (O/MM)], genotype \u20134. Firstgenotype ,5,6. Morgenotype \u20139.In clinical practice, a decision-making process including functional, nutritional, and co-morbidity status is required to tailor first line medical treatment . ElderlyKRAS mutations occur in 35\u201345% of colorectal cancer (CRC), mostly codon 12 (80%), prevalently c.35 G>A (G12D) transversion (32.5%)  , impairi (32.5%) . In the ild-type \u201339; BEV activity ,37.KRAS genotype, depending on differential tumor biological aggressiveness  mutant genotype may significantly affect worse OS of MCRC patients treated with FIr-B/FOx, compared to wild-type or different other mutations . Thus, it was not a clinical trial and approval by ethics committee and institutional review board was not necessary, because patients were treated with conventional treatments without any additional medical intervention out of the best common clinical practice. Patients had histological confirmed diagnosis of MCRC, age \u226518 years, PS \u22642. Criteria to define patients unfit, or not eligible for intensive regimens were: age \u226575 years; uncontrolled severe diseases; cardiovascular disease ; thromboembolic disease, coagulopathy, preexisting bleeding diatheses; proteinuria >1 g/24 h. Patients were classified according to Cumulative Illness Rating Scale (CIRS)  for administration e (CIRS) . Treatme2/die, over 12 h (from 10:00 pm to 10:00 am), days 1\u20132, 8\u20139, 15\u201316, 22\u201323; CPT-11 , 120\u2013160 mg/m2, days 1 and 15; l-OXP , 70\u201380 mg/m2, days 8 and 22; cycles every 4 weeks. Other triplet, doublet and mono-regimens were administered according to previously reported schedules , associated to weekly alternating CPT-11 or L-OXP : TFI/5-Fchedules ,42. TargKRAS codon 12 and 13 mutations by direct sequencing. KRAS exon 2 sequence was performed from PCR-amplified tumor DNA using the Big Dye V3.1 Terminator kit, electrophoresis in ABI PRISM 3130xl Genetic Analyzer, and analysis using the GeneMapper Analysis software version 4.0 .Genetic analyses were performed on paraffin-embedded tissue blocks from primary tumor and/or metastatic sites, as previously reported . GenotypKRAS genotype on clinical outcomes were evaluated. Patients were classified according to: metastatic extension, L-L and O/MM , 23 (59%) were wild-type and 16 (41%) mutant. Clinical features of the 37 patients who underwent first line medical treatments were , specifically c.35 G>A (G12D), 7 (19.4%), c.35 G>T (G12V), 6 (16.6%); codon 13, 2 (5.5%), c.37 G>T (G13V), 1 (2.7%) and c.38 G>A (G13D), 1 (2.7%).Forty patients unfit for intensive regimens, among 72 consecutive MCRC (56%), were treated with : medicalnts were : male/fents were : non-eldMedical treatments were tailored according to age and CIRS stage. Triplet regimens were administered in 18 patients (49%): non-elderly 6, young-elderly 4, old-elderly 8; CIRS primary 2, intermediate 10, secondary 6. Doublet regimens were administered in 15 patients (40%): non-elderly 3, young-elderly 3, old-elderly 9; CIRS primary 1, intermediate 4, secondary 10. Mono-regimens were administered in 4 patients (11%): young-elderly 1, old-elderly 3; CIRS primary 1, intermediate 1, secondary 2.KRAS wild-type patient (33%), with primary rectal tumor and a single L-L metastasis. Twelve patients (32%) received, at least, a second line treatment.Among the 37 patients who underwent medical treatments, 10 were not evaluable for activity: 7 (19%) did not receive at least 2 cycles of treatment; 3 were on-treatment. The intent-to-treat analysis of 27 patients showed ORR 37%  . We obseAmong 7 evaluable L-L patients, ORR was 71%; 3 performed liver metastasectomies (43%) and 1 cCR (14%); median PFS 11 months (3\u201313+ months); median OS 12 months (3\u201313+ months). Among 20 evaluable O/MM patients, ORR was 25%; median PFS 6 months (1\u201312 months); median OS 13 months (1+\u221223+ months). Clinical outcome (PFS and OS) in L-L compared to O/MM patients was not significantly different .Among 16 evaluable patients treated with triplet regimens , ORR wasMoreover, PFS and OS were not significantly different in non-elderly and young-elderly compared to old-elderly patients , and in primary and intermediate CIRS stage compared to secondary stage patients .KRAS wild-type patients evaluable for activity, ORR was 50%  . We observed 3 partial responses (25%); 5 stable diseases (42%); 4 progressive diseases (33%). Disease control rate was 67% . No liver metastasectomies were performed. Median PFS was 6 months (1\u201311 months), 12 events occurred (80%). Median OS was 8 months (3\u201318 months), 10 events occurred. KRAS wild-type compared with mutant patients showed significantly different PFS (p=0.043), but not OS   (Among 14 CI \u00b1 27) . We obset not OS . KRAS c.ctively) , and to ctively) . No diffpatients . PFS andctively) .Patients unfit for first line FIr-B/FOx intensive regimen, due to age (\u226575 years) and/or comorbidities, were prevalent (56%), mostly elderly (76%), particularly old-elderly patients (54%), prevalently PS 1\u20132 (59%), CIRS stage intermediate/secondary (89%), O/MM disease (79%). Most unfit MCRC patients were treated with triplet or doublet regimens , and some (19%) did not reach the first evaluation of activity at 2\u20133 months.Retrospective evaluations of doublets consisting of CPT-11 or OXP, associated to 5-FU or capecitabine in elderly patients eligible for clinical trials gained ORR 18\u201359.4%, PFS 4.9\u201310.0 months and OS 8.5\u201320.7 months ,31,46. BYoung-elderly patients eligible for FIr-B/FOx intensive regimen, prevalently characterised by PS 0 (89%) and intermediate CIRS stage (93%), reported ORR 79%, PFS 11 months, OS 21 months, equivalent to overall patients . A complKRAS wild-type and mutant (KRAS geno-type was reported as significantly affecting PFS and OS in patients treated with XelOx/BEV (KRAS c.35 G>A (G12D) mutant genotype may significantly affect worse OS, compared to wild-type or other mutations , PFS and OS were not significantly different in d mutant ,6,8,37 ad mutant . RecentlelOx/BEV . We receutations . PresentA mutant ,40, and KRAS genotype may indicate different PFS, and c.35 G>A KRAS mutant a significantly worse PFS and OS, compared to wild-type and other mutations. Present findings warrant prospective trials comparing clinical outcome in unfit patients, according to KRAS genotype.In conclusion, in MCRC patients unfit for first line intensive FIr-B/FOx regimen, tailored doublet and triplet medical treatments showed similar activity and efficacy, also according to age and comorbidities."}
+{"text": "Klebsiella quasipneumoniae subsp. similipneumoniae, KP_Z4175. This strain, isolated as part of a hospital infection-control screening program, is resistant to multiple \u03b2-lactam antibiotics, aminoglycosides, and trimethoprim-sulfamethoxazole.We report here the draft genome sequence of a multidrug-resistant clinical isolate of  Klebsiella quasipneumoniae, formerly Klebsiella pneumoniae phylogroup KpII, was recently taxonomically reclassified as a new sister species of K.\u00a0pneumoniae with two subspecies, K.\u00a0quasipneumoniae subsp. quasipneumoniae and K.\u00a0quasipneumoniae subsp. similipneumoniae  using the GGDC 2.1 software  generating 2 \u00d7 301-bp paired-end reads. A total of 15,256,306 reads were produced comprising 1,540,886,906 bases after adapter sequence trimming. K.\u00a0quasipneumoniae subsp. similipneumoniae strain KP_Z4175, antibiotic resistance genes were identified using ResFinder version 2.1 -IIc), two fluoroquinolone resistance genes (aac(6\u2032)Ib-cr and QnrB4), one macrolide-lincosamide-streptogramin B resistance gene (ere(A)), two phenicol resistance genes (cmlA1 and floR), one rifampin resistance gene (ARR-2), two sulfonamide resistance genes (sul1 and sul2), one tetracycline resistance gene (tet(D)), and one trimethoprim resistance gene (dfrA14). All identified resistance genes had nucleotide identities of 98.35 to 100% over 85 to 100% of the reference gene lengths. Broth microdilution testing using CLSI breakpoints for Enterobacteriaceae indicated that KP_Z4175 is resistant to gentamicin (MIC >64), tobramycin (=32), cefazolin (>64), ceftriaxone (>64), aztreonam (>64), and trimethoprim-sulfamethoxazole (>64). The isolate had intermediate resistance to ampicillin-sulbactam (=16) and pipericillin/tazobactam (=32), and was sensitive to ertapenem (\u22640.03), imipenem (=1), meropenem (\u22640.03), amikacin (=0.5), cefepime (=8), and ciprofloxacin (=1).To examine the antibiotic resistance profile of sion 2.1 . In addieumoniae , \u03b2-lactaLVCD00000000. The version described in this paper is version LVCD01000000.This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession number"}
+{"text": "Dysregulation of the nucleo-cytoplasmic transport of proteins plays an important role in carcinogenesis. The nuclear export of proteins depends on the activity of transport proteins, exportins. Exportins belong to the karyopherin \u03b2 superfamily. Exportin-1 (XPO1), also known as chromosomal region maintenance 1 (CRM1), mediates transport of around 220 proteins. In this review, we summarized the development of a new class of antitumor drugs, collectively known as selective inhibitors of nuclear export (SINE). KPT-330 (selinexor) as an oral agent is showing activities in early clinical trials in both solid tumors and hematological malignancies.The online version of this article (doi:10.1186/s13045-014-0078-0) contains supplementary material, which is available to authorized users. The nucleo-cytoplasmic transport of proteins plays an important role in maintaining normal cellular functions. The nuclear export of proteins depends on the activity of transport proteins, exportins. Exportin-1 (XPO1), also known as chromosomal region maintenance 1 (CRM1), mediates transport of around 220 proteins . XPO1 isXPO1 binds to the cargo proteins through a leucine rich nuclear export signal (NES) and transports the proteins through a membrane pore complex via a Ran-GTP gradient -15]  . The cliin vitro with most potency. However, its use in vivo is limited by poor pharmacokinetics . KPkinetics -[25]. Th50) of 150 nmol/L and inhibited pancreatic cancer cell lines while sparing normal human pancreatic ductal epithelial cells. The in vivo effects were noted using KPT-330 (selinexor) in subcutaneous and orthoptic pancreatic cancer models in mice. Oral administration of KPT-330 led to significant tumor growth inhibition when compared with control or gemcitabine treatment . EG. EG37]. l status . The antl status . In celll status . KPT-330l status . Moreovenografts .in vitro in RCC cell lines . Tr. Tr52]. hibitors . They cohibitors ,54]. Ho. Hoet alsistance ,63],. SI. SIin vi KPT-185 . FLT3 ge KPT-185 . SINE do KPT-185 ,74]. Ko. Koin viradation . Nutlin-radation . The add,. SI. SI79]. able IC50. In vivoet al. demonstrated successful use of KPT-185 in vitro and KPT-330 in vivo in Philadelphia chromosome positive ALL (Ph + ALL) and chronic myeloid leukemia blast crisis (CML-BC). Combination with imatinib led to synergistic effects . Wa. Wa81].  effects . SINE tr effects .et al. recently. They showed that KPT-330 treatment given to heavily pretreated, refractory/relapsed AML patients had no DLT. Out of the 32 evaluable patients, 4 (12%) showed complete response (CR) with hematological recovery, 1 (3%) showed marrow CR (mCR), mCR without hematological recovery was seen in 1 (3%) patient. Partial response (PR) was seen in 2 patients (6%). Eleven patients (34%) showed progression while 12 (37%) experienced stable disease after 30 days . Pr. Pr91]. L) cells . KPT-185L) cells ,93]. KP. KPet alL) cells . In a 37L) cells .in vitro and in vivo. Su. Suin vid in vivo. C-myc ad in vivo. A studyd in vivo. BRD4, ad in vivo,99]. BR. BRin vid in vivo. JQ1, a d in vivo. This reet al. conducted an in vitro study of KPT-185 in MCL and showed that SINE increases MCL cell apoptosis primarily by increasing nuclear p53 levels -[1-[1100]-. KPT dru3 levels . They vepression . London  effects . Gutierrpatients . These dMore and more targeted small molecule inhibitors are entering clinical application quickly ,84],,-113]. S. S113]. DL and KP designed the study. All authors have contributed to data preparation, drafting and revising the manuscripts. All authors have read and approved the final manuscript."}
+{"text": "Blood biomarkers are increasingly used to diagnose, guide therapy in, and risk-stratify community-acquired pneumonia (CAP) patients in emergency departments (EDs). How pre-analytic factors affect these markers' initial levels in this population is unknown.In this secondary analysis of consecutive ED patients with CAP from a large multicentre antibiotic stewardship trial , we usedP = 0.02) in patients with liver insufficiency; ProADM, + 13.2%  in patients with age above median; and CRP, -12.8%  with steroid pretreatment.Of 925 CAP patients , 25.5% had antibiotic pretreatment, 2.4%, corticosteroid pretreatment, 22.3% chronic renal failure, and 2.4% chronic liver insufficiency. Differences associated with pre-analytic factors averaged 6.1 \u00b1 4.6%; the three largest statistically significant changes (95% Cl) were: PCT, +14.2% (+2.1 to +26.4%, The influence of pre-analytic factors on the examined biomarkers is marginal and not clinically relevant. Our observations reinforce the concept ofusing biomarkers in algorithms with widely- separated cutoff values and overruling criteria considering the entire clinical picture, without adjustment for pre-analytic factors."}
+{"text": "The importance of maternal nutrition to offspring health and risk of disease is well established. Emerging evidence suggests paternal diet may affect offspring health as well.In the current study we sought to determine whether modulating pre-conception paternal B vitamin intake alters intestinal tumor formation in offspring. Additionally, we sought to identify potential mechanisms for the observed weight differential among offspring by profiling hepatic gene expression and lipid content.1638N mice  were fed diets containing replete , mildly deficient (DEF), or supplemental (SUPP) quantities of vitamins B2, B6, B12, and folate for 8 weeks before mating with control-fed wild type females. Wild type offspring were euthanized at weaning and hepatic gene expression profiled. Apc1638N offspring were fed a replete diet and euthanized at 28 weeks of age to assess tumor burden.Male Apc1638N offspring of different paternal diet groups. Although in female Apc1638N offspring there were no differences in tumor incidence or multiplicity, a stepwise increase in tumor volume with increasing paternal B vitamin intake was observed. Interestingly, female offspring of SUPP and DEF fathers had a significantly lower body weight than those of CTRL fed fathers. Moreover, hepatic trigylcerides and cholesterol were elevated 3-fold in adult female offspring of SUPP fathers. Weanling offspring of the same fathers displayed altered expression of several key lipid-metabolism genes. Hundreds of differentially methylated regions were identified in the paternal sperm in response to DEF and SUPP diets. Aside from a few genes including Igf2, there was a striking lack of overlap between these genes differentially methylated in sperm and differentially expressed in offspring.No differences in intestinal tumor incidence or burden were found between male ApcIn this animal model, modulation of paternal B vitamin intake prior to mating alters offspring weight gain, lipid metabolism and tumor growth in a sex-specific fashion. These results highlight the need to better define how paternal nutrition affects the health of offspring. B, C, D) Photomicrographs of Oil Red-O stained liver sections from DEF (22.7%), CTRL (22.3%) and SUPP (37.6%) offspring respectively. Images are from mice with the highest staining area in each group. Images taken at 100x and scale bar is 200 \u03bcm. Paternal diets: DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented.(DOCX)Click here for additional data file.S3 FigA) CpGs 1\u201314 are located at -359, -357, -347, -337, -334, -320, -314, -304, -296, -288, -280, -274, -267 and -264 bp from the start of Acaca1 exon 1 respectively. B) CpGs 1\u20139 are located at +36, +59, +50, +27, +14, +10, -3, -14, -40 and -44 bp from the start of Elovl6 exon 1 respectively. C) CpGs 1\u201319 are located -122, -113, -96, - 89, -77, -73, -64, -61, -52, -50, -48, -45, -41, -39, -33, -31, -28, -26 and -23 bp from start of Gpam exon 1 respectively.D) CpGs 1\u20138 are located -3775, -3766, -3754, -3706, -3702, -3699, -3685, -3678 and -3665 bp from start of H19 exon 1 respectively. Unless noted, for each CpG site, 2-Way ANOVA p (group) >0.05 and p (sex) >0.05. \u2018a\u2019 denotes p <0.05 for group effect and \u2018b\u2019 denotes p v0.05 for sex effect. Unless noted, repeated measures ANOVA for group  effect yielded a p>0.05. Paternal diets: DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented; M male; F female. n = 4 males and 8 females per group.DNA methylation was measured in target genes using bisulfite pyrosequencing. (DOCX)Click here for additional data file.S1 TableB denotes biotin modification of primer at 5\u2019 end of FWD or 3\u2019 end of RVS primer.(DOCX)Click here for additional data file.S2 TableValues are mean \u00b1 SEM. n = 10, 13 and 9 respectively.(DOCX)Click here for additional data file.S3 TableDMR ID is an arbitrary identifier for each differentially methylated region; CTRL, control diet; DEF, B vitamin deficient diet; m1, mean log2(636/532) for CTRL sperm; m2, mean log2(636/532) for DEF sperm; chr:position identifies the location of the DMR within the chromosome using mouse genome build mm9 coordinates; diff, mean log2(635/532) difference across the DMR; qval, q-value\u2014an adjusted p-value indicating the false discovery rate (FDR) calculated by the Wilcoxon rank sum-test. For genes with a qval < 0.1, less than 10% of genes are expected to be false positives; Gene symbol identifies the gene annotated to a DMR and its proximity in bases to the left (-) or right (+) of the DMR up to a distance of 1000 kb where all genome coordinates and gene symbols are derived from mouse genome build mm9. n = 8/gp.(XLSX)Click here for additional data file.S4 TableDMR ID is an arbitrary identifier for each differentially methylated region; CTRL, control diet; SUPP, B vitamin supplemented diet; m1, mean log2(636/532) for CTRL sperm; m2, mean log2(636/532) for SUPP sperm; chr:position identifies the location of the DMR within the chromosome using mouse genome build mm9 coordinates; diff, mean log2(635/532) difference across the DMR; qval, q-value\u2014an adjusted p-value indicating the false discovery rate (FDR) calculated by the Wilcoxon rank sum-test. For genes with a qval < 0.1, less than 10% of genes are expected to be false positives; Gene symbol identifies the gene annotated to a DMR and its proximity in bases to the left (-) or right (+) of the DMR up to a distance of 1000 kb where all genome coordinates and gene symbols are derived from mouse genome build mm9. n = 8/gp.(XLSX)Click here for additional data file.S5 Table* Values are mean \u00b1 SEM. Note that number weaned offspring is less than sired offspring due to maternal cannibalism that is common amongst first-time mothers. DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented.(DOCX)Click here for additional data file.S6 TableValues are mean \u00b1 SEM. Paternal diets, DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented. Sample size is in parentheses.(DOCX)Click here for additional data file.S7 TableSignificant differences according to Cuffdiff analysis combining male and female offspring (q<0.05). Expression values are Fragments Per Kilobase Of Exon Per Million Fragments Mapped (FPKM). CTRL, control; DEF, deficient . n = 10/gp.(XLSX)Click here for additional data file.S8 TableSignificant differences according to Cuffdiff analysis combining male and female offspring (q<0.05). Expression values are Fragments Per Kilobase Of Exon Per Million Fragments Mapped (FPKM). CTRL, control; SUPP, supplemented . n = 10/gp.(XLSX)Click here for additional data file.S9 TableCategories identified by Ingenuity Pathway Analysis\u00ae. DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented . P values represent the range of p values of the sub-categories of each category.(DOCX)Click here for additional data file.S10 Tablen = 5/gp.(XLSX)Click here for additional data file.S11 Table(DOCX)Click here for additional data file."}
+{"text": "Edwardsiella piscicida-like species is a Gram-negative facultative anaerobe that causes disease in some fish species. In this report, we present the complete and annotated genome of isolate LADL05-105, recovered from cultured tilapia reared in Louisiana, which contains a chromosome of 4,142,037\u00a0bp and no plasmids.An  Edwardsiella, first recognized in the 1960s  was designated the type strain of the recently proposed taxon, E.\u00a0anguillarum sp. nov.  and Pacific Biosciences (Pac-Bio) (113\u00d7 coverage). The PBcR module of wgs-8.2beta (Celera) (Genomic DNA from (Celera) , 10 was (Celera) .The circularized and completed genome was submitted to the National Center for Biotechnology Information Prokaryotic Genomes Annotation Pipeline (PGAP) for annotation and submission to GenBank. The genome data was also subjected to RAST , 14 annoE.\u00a0piscicida-like species genome consists of one circular chromosome with 4,142,037\u00a0bp and 58.8% GC content. PGAP annotation predicted 3,686 genes encoding 3,159 proteins and 99 tRNAs. RNAmmer (http://enve-omics.ce.gatech.edu/ani). The complete genome of LADL05-105 shares an ANI of 99.84% with Edwardsiella strain 080813, which was recently proposed as the type strain for Edwardsiella anguillarum sp. nov. (GenBank accession no. CP006664) (E.\u00a0piscicida-like sp. isolate EA181011 (GenBank accession no. CP011364) (E.\u00a0piscicida isolate C07-087 (GenBank accession no. CP004141)  (E.\u00a0tarda isolate FL95-01 (GenBank accession no. CP011359)  , 94.39% P001600) , and 84.P011359) . These AEdwardsiella piscicida-like sp. isolate LADL05-105 has been deposited in GenBank under the accession number CP011516.The complete genome sequence for"}
+{"text": "Controlling MRSA has been a challenge for Geneva University Hospitals (HUG), particularly after the introduction of ST228 SCCmecI hyperendemic clone in 1999.To describe HUG\u2019s secular trends of MRSA, infection control measures and predominant MRSA clones using Staphylococcal chromosomic cassettes (SCCmec) genotyping.nd HHC, periodic audits and feedback (2006). The intensive care unit performed MRSA screening on admission, discharge and weekly since 2004. Surveillance included: (1) incidence rates (IR) of hospital acquired (HA)-MRSA infection or colonization; (2) HA-MRSA bloodstream infections (BSI); (3) proportion of MRSA/S. aureus BSI; (4) IR of MRSA clinical cultures (CC); (5) proportion of SCCmec in MRSA strains, assessed by routine multiplex PCR assay. Representative isolates were grouped in MLVA clusters to evaluate genomic diversity and subjected to MLST.A multifaceted MRSA prevention program initiated in 1993 included patient screening, decontamination, surveillance, contact isolation, an alert system and a hospital-wide hand hygiene campaign (HHC); subsequently strengthened by: an educational campaign (2003), routine MRSA SCCmec genotyping (2005), a 2S. aureus BSI remained around 34% (2000-2009), declining to 18% (2014). SCCmecI strains declined from 83% (2005) to 32% (2014); SCCmecII and SCCmecIV were higher in non-acute settings.At HUG, from 2000-2014, 12,543 patients were MRSA-colonized or infected . From 2000-2007, annual rates of all indicators showed an increasing trend, declining since 2008. New HA-MRSA cases per 100 admissions increased from 1.36 to 2.00 (2006), and then declined to 0.29 (2014). Trends expressed by incidence density of cases per 1000 hospital-days: HA-MRSA, from 0.92 to 1.36 (2007) to 0.21 (2014); ICU-acquired HA-MRSA from 2.3 (2002) to 10.5 (2006) to 1.31 (2014); MRSA-CC rates from 0.68 to 1.44 (2008), to 0.24 (2014); HA-BSI from 0.049 to 0.07 (2009), to 0.016 (2014). The proportion of MRSA/A multifaceted prevention program and possible changes in biologic fitness of the ST228 SCCmecI clone helped to decrease endemic MRSA rates for the last 7 years.None declared."}
+{"text": "In sepsis, impaired function and loss of antigen-presenting cells are observed in secondary lymphoid organs , the sitTo study seven-day lymphocyte and immunoglobulin trajectory, alterations in B cell subsets and potential mechanisms in septic ICU patients.9/L, respectively.Adults with severe sepsis from community-acquired infections without documented immunosuppression were enrolled. Hypogammaglobulinaemia and absolute lymphopenia were defined as IgG < 6.1, IgM < 0.4, IgA < 0.8 g/L and lymphocytes < 1.2 \u00d7 10Flow cytometry [FACScalibur [BD Biosciences]; FlowJo software,] was used for:Identifying na\u00efve, transitional, IgM, IgG and IgA memory and plasmablasts using anti-human CD19-PerCpCy5.5, IgG-APC H7, IgM-V450, CD24-PeCy7 [all BD Biosciences], IgA-FITC, CD38-PE, Annexin V Apoptosis detection set PE-Cy7 [all eBioscience] and live-dead stain [Invitrogen]. ADDIN EN.CITE ADDIN EN.CITE.DATA .FMO and isotype controls were used to define population gates.B cell survival ligands  were measured using ELISA.\u00ae Human Apoptosis Array; false discovery rate = 5%].Differentially expressed genes in sepsis are reported  and T [100%] lymphocyte counts. Trajectory of significantly higher increment immunoglobulins and lymphocyte counts occur earlier in survivors compared to non-survivors.In sepsis  there wasPreferential apoptotic loss of memory B cells and plasmablasts, with apoptotic cells showing higher phosphorylated extracellular signal-regulated kinases [p-ERK fluorescence], but no differences in the phosphorylated B cell receptor linked kinases  and protein kinase B.BAFF/APRIL levels were normal.Fas and bcl-2 apoptosis regulator genes were up regulated.In sepsis, activation-associated B cell apoptosis and changes in secondary lymphoid organs deplete B cell memory and contribute to long-term immunosuppression in survivors.UK NIHR, Biomedical Research Centre at Guy\u00b4s and St Thomas\u00b4 NHS Trust & King\u00b4s College London"}
+{"text": "Recent nationwide shortages of low-calcium formula (LCF) suggest this problem may be widespread. CYP24A1 mutations have been identified as a potential cause of IIH.Consultations for infantile hypercalcaemia (IIH) have increased at Sydney Children\u2019s Hospital since guidelines for vitamin DTo determine if IIH is occurring more commonly, de-identified, first-measured serum calcium from all infants <6 months (n=5796) measured in our laboratory, were grouped by years 2005-2007 (n=1516), 2008-2010 (n=1945) and 2011-2013 (n=2335). In addition, we analysed 13 infants treated by our department for idiopathic infantile hypercalcaemia (IIH) from 2011-2013.2P<0.001). Rates of hypocalcaemia (<2.25mmol/L) fell steadily . Twelve mothers of our 13 infants with IIH received antenatal vitamin D3 supplementation. One infant also received 400 units/day Vitamin D3 post-natally. At diagnosis, median age was 13 days (range 4-50), 77% were breast-fed, 54% were symptomatic and 25% had nephrocalcinosis. Median initial calcium was 3.00mmol/L (range 2.84-4.03) and phosphate 2.04mmol/L (1.1-3.33). PTH was not elevated (median 1.0pmol/L [<0.3-3.1]), urinary calcium: creatinine ratio not suppressed , 25OHVitD low-normal (median 44nmol/L [17-218]) and 1,25(OH)2VitD elevated (median 232pmol/L [64-720]), in keeping with an abnormality in CYP24A1. In 7/10 infants with data available, treated with LCF for median 95 days (range 25-310), median PTH rose to 17.1pmol/L  with a trend to lower 25OHVit D  despite continued high-normal calcium levels (median 2.66mmol/L [2.11-2.75]).Rates of hypercalcaemia (>2.75mmol/L) increased from 2011 (1.1% vs 1.3% vs 8.7%, \u03c73 supplementation as an aetiological factor. IIH was associated with significant morbidity, including symptomatic hypercalcaemia and nephrocalcinosis. Treatment with LCF prevented further symptomatic hypercalcaemia, but resulted in elevated PTH. The biochemistry of our patients with IIH raises variations in Vitamin D metabolism or calcium set-point as potential associated factors.Concurrent changes in rates of hyper and hypo-calcaemia suggest antenatal vitamin D"}
+{"text": "A range of cytokines are altered during sepsis. Multiplex analysis of inflammatory mediators makes it easier to study a larger range of mediators, and combined with newer bioinformatical tools , this has the potential to bring new knowledge of the inflammatory process during sepsis.To investigate if there are differences in expression of multiple inflammatory mediators between septic patients with and without bacteraemia during the initial phase of hospitalization, in order to identify bacteraemic patients.All together 80 adult, immunocompetent patients with sepsis and confirmed bacterial infection were included prospectively at Haukeland University Hospital during the first 24 hours of hospitalization at the HDU, ICU or medical department. Luminex analysis of plasma was used to analyse 35 mediators; 16 cytokines, 6 growth factors, 4 adhesion molecules and 9 matrix metalloproteinases/ tissue inhibitors of metalloproteinases. Mann-Whitney U-test were used analysing differences between the two groups were we examined both p-values with (< 0.0014) and without (p < 0.05) Bonferronis correction. We performed unsupervised hierarchical clustering of the five statistically most significant mediators using J-express software.In 41 patients (51%) a positive blood culture was found, while the remaining 39 (49%) had proven bacterial cause by confirmed by other tests. Among these, 37 were gramnegative, 39 were grampositive, while 4 patients had mixed infections.We found that 16 of 35 mediators showed statistical differences between bacteraemic and non-bacteraemic patients; IL1ra (p = 0.0015), IL-10 (p = 0.0009), CCL2 (p = 0.0376), CCL4 (p < 0.0001), CCL5 (p = 0.0376), CXCL8 (p = 0.0026), CXCL11 (p = 0.0468), TNF\u03b1 (p < 0.0001), HGF (p = 0.0018), E-selectin (p = 0.0004), ICAM-1 (p = 0.0009), VCAM-1 (p < 0.0001), MMP-8 (p = 0.0311), TIMP-1 (p < 0.0001), TIMP-2 (p = 0.0054) and TIMP-4 (p = 0.0036).Using unsupervised hierarchical clustering (se figure), we found that the five mediators TNF\u03b1, CCL4, E-selectin, VCAM-1 and TIMP-1 gave a reasonable good discrimination of bacteraemia patients; the patients with bacteraemia were mostly clustered in two separate groups  showing higher levels of the mediators. Only 5 (12%) of the bacteraemic patients were clustered in the middle cluster, while most of the non-bacteraemia patients  were clustered in this group (Chi-square p < 0.0001).We find higher levels of inflammatory markers in sepsis patients with bacteraemia, and hierarchical clustering may aid early identification of patients with bacteraemia.Upper and lower cluster contains most bacteraemia patients (red) showing higher levels of inflammatory mediators (green). Middle cluster with lower levels of mediators (blue) contains most patients without bacteraemia. Mediators are clustered horizontally and patients vertically."}
+{"text": "Eosinophilic esophagitis (EoE) is an increasingly common cause of chronic and recurrent esophageal symptoms with significant impact on quality of life. Allergic responses to food and aeroallergens have been implicated in the etiology of this disease.Clinical and sensitization profile characterization of an adult population with EoE diagnosis (EoEd) followed in our Immunoallergology Department.Retrospective study using our EoE database (2009-2014) of patients (pts) \u2265 18 years.Characterization of demographic, symptomatic, laboratorial , endoscopic and histological factors and sensitization profile .We included 25 patients (pts) ; median age of symptoms onset and diagnosis: 27(13-66) and 30(16-67) years respectively; average between symptoms onset and EoEd:27(1-180) months. Follow-up for 25.3\u00b121.7(1-88) months. Reported symptoms-%pts: impaction-84,GERD-like symptoms 48, dysphagia-36, bloating-24, abdominal pain-16 and vomiting-12. Most frequent first symptom: impaction-48%pts. 60% of pts had personal history of atopy . Median peripheral Eos 388.8(40-980) and total IgE 163,8kU/L(5.6-436). The main baseline endoscopic findings (%pts): rings (43.5), furrows (26.1), hiatal hernia (21.7) and white plaques (21.7). 13% had microabcesses. After EoEd, allergic sensitization was identified in 88% pts ; food allergens ; mites (54.2); pollens (37.5). After therapeutic: all had symptomatic improved; 12.5% pts achieved Eos\u226415/HPF in esophageal biopsy, all with food allergy who underwent dietary eviction.The first and the most frequent symptom was food impaction. The prevalence of allergic sensitization was high. The potential severity of the symptoms justifies the importance of recognition and management of the disease. A multidisciplinary evaluation is essential in the clinical, diagnostic and therapeutic workup."}
+{"text": "In 2012, 69.6% of adults aged \u226540 years who ever had a cardiovascular event (73.2% of men and 65.4% of women) were taking low-dose aspirin to prevent or control heart disease. Non-Hispanic white men (75.9%) were more likely to be taking low-dose aspirin compared with Hispanic (60.7%) and non-Hispanic black men (60.6%). No statistically significant differences were oberved among women by race/ethnicity.Source: National Health Interview Survey, 2012 data. Available at http://www.cdc.gov/nchs/nhis.htm.Reported by: Renee M. Gindi, PhD, iuz2@cdc.gov, 301-458-4502; Brian W. Ward, PhD."}
+{"text": "Their structures were elucidated by comprehensive spectroscopic analyses including MS, IR, 1D and 2D NMR. All compounds were evaluated for their anti-AChE activities.Seven new monoterpene phenyl ethers, namely micranthumnins A-G (Supplementary material is available for this article at 10.1007/s13659-013-0007-x and is accessible for authorized users. Supplementary material, approximately 4.23 MB."}
+{"text": "There are errors in the \u201cOncomine data analysis\u201d section of the Materials and Methods. It should read:Oncomine and CCLE data analysisUsmg5 copy number profiles in a large subset of cancer cell lines . In the dataset, the log2  values were analyzed. The number of DNA copies ) were calculated as advised in Oncomine instructions.We used the Oncomine Cancer Genomics Data Analysis tool [30] and Cancer Cell Line Encyclopedia, CCLE [35] to mine Usmg5 copy number in cancers\u201d section of the Results. It should read:There are errors in the \u201cUsmg5 copy number in cancer cell linesUsmg5 copy number in a large panel of cell lines  of ll lines  Several There are errors in Usmg5 copy number in various cancer cell lines :R215. doi: 10.1158/1541-7786.MCR-08-010766. Hu X, Stern HM, Ge L, O'Brien C, Haydu L, Honchell CD, Haverty PM et al. Genetic alterations and oncogenic pathways associated with breast cancer subtypes. Mol Cancer Res. 2009 Apr;7(4):511\u201322. doi: 10.1186/1755-8794-3-2367. Lu X, Zhang K, Van Sant C, Coon J, Semizarov D. An algorithm for classifying tumors based on genomic aberrations and selecting representative tumor models. BMC Med Genomics. 2010 Jun 22;3:23. doi: 10.1158/1541-778668.Olejniczak ET, Van Sant C, Anderson MG, Wang G, Tahir SK, Sauter G et al. Integrative genomic analysis of small-cell lung carcinoma reveals correlates of sensitivity to bcl-2 antagonists and uncovers novel chromosomal gains. Mol Cancer Res. 2007 Apr;5(4):331\u20139. Doi: 10.1172/JCI3712769. Sos ML, Michel K, Zander T, Weiss J, Frommolt P, Peifer M et al. Predicting drug susceptibility of non-small cell lung cancers based on genetic lesions. J Clin Invest. 2009; Jun;119(6):1727\u201340. doi:"}
+{"text": "Beh\u00e7et's disease (BD) is an inflammatory disorder of unknown aetiology, unanimously recognized as both autoimmune and autoinflammatory disease. Indeed many of its classical manifestations overlap with those of monogenic autoinflammatory disorders. Clinically disease is characterized by multiple organ involvement, in particular by the \u201ctriple symptom complex\u201d, consisting of recurrent oral aphthosis, genital ulcers and recurrent bilateral uveitis. The abnormal activation of either innate and adaptive immunity, triggered by some microbial agents in genetically predisposed individuals, with consequent interaction of both T lymphocytes and activated neutrophils would seem to be involved in the disease onset. Therefore multiple cytokines may contribute to the pathological scenario of BD playing a pivotal role in the occurrence of the clinical manifestations.To determine serum levels of IL-8, IL-18, IFN-\u03b12a, IL-6, IFN-\u03b3, CXCL10, CXCL11, CXCL9 and serum amyloid-A (SAA) concentration in patients with BD, in comparison to healthy controls (HC), and to correlate their concentration with the status of disease activity.78 serum samples were collected from 58 BD patients . Serum cytokine levels of IL-8, IL-18, IFN-\u03b12a, IL-6, IFN-\u03b3, CXCL10, CXCL11 and CXCL9 were determined using a multiplex bead analysis as well as SAA was assessed by Enzyme linked-immunosorbent assay.In BD patients serum concentrations of IL-8 (p=0.0001), IL-18 (p=0.0058), IFN-\u03b12a (p=0.0181) and IL-6 (p=0.0233) were significantly higher than in HC. When BD patients were divided into active and inactive group, IL-8 and IL-18 resulted higher in both active- (p=0.0001 and p=0.012 respectively) and inactive-BD (p=0.0001 and p=0.0128 respectively) than in HC, while IFN-\u03b12a (p=0.0141) and IL-6 (p=0.0332) serum levels were significantly higher in active-BD than HC. Moreover, CXCL11 (p=0.0154) serum concentrations were significantly lower in inactive-BD than HC. We also compared serum cytokine profiles between BD patients with SAA serum levels \u226420 mg/L, >20 mg/L and HC. Interestingly, we observed that BD patients with SAA >20 mg/L showed higher levels of inflammatory markers than HC. Among these cytokines, IL-18, IFN-\u03b12a and IL-6 were higher in BD group with SAA >20 mg/L than HC, whereas IL-8 and CXCL9 levels were higher than in patients with SAA \u226420 mg/L and HC.BD patients exhibit elevated levels of specific inflammatory mediators, especially during active disease periods and in those patients with SAA serum levels >20 mg/L, thus suggesting a possible role of SAA in the induction of BD inflammatory manifestations."}
+{"text": "To determine the agreement of manual and semi-automatic (SA) diameter and volume measurements of nodules found in low-dose computed tomography lung cancer screening.3) in 1,498 Dutch-Belgian NELSON trial participants were used. Extrapolated volume based on semi-automatic (SA) maximum diameter and mean of maximum transversal and perpendicular diameter were compared to SA volume measurements by Bland-Altman plots. Analyses were repeated by margin  and shape . In 100 randomly selected nodules, diameters were measured manually by two independent radiologists, and compared to the SA diameters.Baseline data of 2,240 solid intermediate-sized nodules (volume 50-500mmMedian participant age was 59-years (interquartile range:8), 14.2% were women. Compared to SA volume, volume extrapolated from SA mean or maximum diameter led to mean overestimation of 47.2% : 44.7-49.7%) and 85.1% (95%-CI:81.2-89.0%), respectively. For irregular and non-spherical nodules, mean overestimation was higher; 161.7% (95%-CI:131.7%-191.8%) and 168.9% (95%-CI:155.2%-182.5%), respectively. Manual diameter measurement overestimated SA maximum diameter by \u226510% in 44% (44/100) and underestimated by \u226510% in 18% (18/100) of the nodules. Using a 10-mm criterion for referral, SA maximum diameter measurements of indeterminate nodules would have led to direct referral in 7.9% (177/2240). Manual measurements would have led to 31% (31/100) referrals.The agreement between manual and SA diameter, as well as between volume extrapolated from SA diameter and SA volume is poor. Applying manual and SA diameter measurement in CT lung cancer screening leads to a substantial shift in nodule stratification compared to SA volume measurements."}
+{"text": "HIV-infected patients show an increased risk of cardiovascular disease (CVD). In the general population, lipoprotein-associated phospholipase A2 (Lp-PLA2) appears to be an independent predictor of CVD. We aimed to study associations between Lp-PLA2 plasma levels and other risk factors for CVD in HIV patients.A cross-sectional, comparative study of two series of cases  was conducted. Eighty-seven percent HIV patients on antiretroviral therapy (ART), 72.4% with HIV-1 viral load <50 cop/mL. Inflammatory biomarkers  and internal carotid intima-media thickness (IMT) were measured and CVD risk  was calculated. Univariate and multivariable associations between these variables were performed.HIV patients presented higher Lp-PLA2 levels [276.81 ng/mL (209.71\u2013356.58)] than uninfected healthy controls [220.80 ng/mL (172.70\u2013256.90)], p\u22640.01. In univariate analysis of the global sample, only cigarette smoking was associated with higher Lp-PLA2 levels, p\u22640.001. In HIV group, female and smoker patients showed higher Lp-PLA2 levels, p\u22640.05. No significant association was found between Lp-PLA2 levels and another CVD risk factors, carotid IMT, Framingham and SCORE algorithms, ART, HIV-1 viral load neither and CD4+ T lymphocyte count. In multivariate analysis, cigarette smoking remained significantly associated with Lp-PLA2 levels .HIV-infected patients present higher Lp-PLA2 levels than healthy controls, and in this population, tobacco smoking is significantly associated with increased Lp-PLA2 levels. Smoking cessation should be a priority in CVD prevention in HIV-infected patients."}
+{"text": "Edwardsiella tarda is a Gram-negative facultative anaerobe that has been isolated from fish, reptiles, amphibians, and mammals, including humans. This is a report of the complete and annotated genome of isolate FL95-01, recovered from channel catfish . Edwardsiella tarda is a Gram-negative intracellular facultative anaerobic bacteria infecting a wide range of avian, reptilian, fish, and mammalian hosts across the globe  and Pacific Biosciences (PacBio) (88\u00d7 coverage) platforms. The longest 40\u00d7 coverage PacBio reads were error-corrected with the remaining PacBio data using the PBcR module within Celera Assembler version 8.2 beta , 16. TheThe circularized and complete genome was submitted to the NCBI Prokaryotic Genomes Annotation Pipeline (PGAP) for annotation and submission to GenBank. Furthermore, the genome was submitted for RAST , 20 annoE.\u00a0tarda genome consists of one circular chromosome with 3,620,701\u00a0bp and 57.3% GC content. PGAP annotation predicted 3,258 genes encoding 3,091 proteins and 101 tRNAs. RAST analysis predicted 505 subsystems with 3,318 coding sequences and 129 RNAs. RAST comparison of FL95-01 with Edwardsiella piscicida C07-087 showed 89 unique subsystems in FL95-01 related to carbohydrate metabolism, cell wall and capsule, and copper tolerance. Additionally, RNAmmer (http://enve-omics.ce.gatech.edu/ani). The complete genome of E.\u00a0tarda isolate FL95-01 shares an ANI of 83.4% with E.\u00a0piscicida isolate C07-087 (GenBank accession no. CP004141) (E.\u00a0piscicida isolate 080813 (GenBank accession no. CP006664)   , and 83.P001600) . Edwardsiella tarda isolate FL95-01 has been deposited in GenBank under the accession number CP011359.The complete genome sequence for"}
+{"text": "Right atrial thrombus (RT) provides a rationale for anticoagulation and substrate for embolic events. CMR is well validated for thrombus detection, but has yet to be used to assess prevalence and predictors of RA thrombus among at risk cohorts.The population comprised consecutive patients with central venous catheters undergoing CMR at Memorial Sloan Kettering Cancer Center . Delayed enhancement CMR (inversion recovery GRE) was used to identify RT; defined as a right atrial (RA) mass with avascular tissue characteristics (non-enhancing) on \"long TI\" (600msec) DE-CMR. Cine-CMR (SSFP) was used to quantify cardiac structure and function, including RA and RV function and chamber size. Clinical indices were categorized based on medical record review. Echo (if performed within 14 days of CMR) was retrieved from image archives and independently read for RT. Clinical records were queried for documented pulmonary embolus (PE) within 60 days of CMR.2/m2, p= 0.94), RA end-systolic area , RV end-diastolic volume , and RVEF . Cancer diagnosis , catheter depth , age and gender (both p=NS) were similar between groups. Transthoracic echo, attained 4.1\u00b13.8 days from CMR in 50% of the population, demonstrated high sensitivity (89%) but moderate specificity (75%) in relation to DE-CMR. Cine-CMR yielded similar sensitivity (82%) but improved specificity (97%) vs. the reference standard of DE-CMR (Table). 27% of patients (3/11) with RT on DE-CMR had PE; all occurred prior to DE-CMR (average of 14 days before). Conversely, no PEs occurred among patients without RT. Clinical embolic events were independent of RT size .50 cancer patients  with RA catheters were studied. CMR was performed for evaluation of a suspected RA mass (36%) or unrelated clinical indications (64%). RT was present in 22% (n=11); all had RT avascularity confirmed by dedicated \"long TI\" DE-CMR. Among affected patients, 63% had a solitary RT (36% multiple). Patients with RT had similar right-sided structure and function vs. those without RT based on RA end-diastolic area (10.2\u00b13.5 vs. 10.2\u00b12.1 cmCatheter associated RT occurs independently of right-sided structure or function, and is associated with clinical embolic events. Morphologic imaging by cine-CMR and echo provide limited diagnostic utility for RT as established by DE-CMR tissue characterization.None."}
+{"text": "AbstractParatenetus Spinola are reviewed and a key is presented for their identification. Five species are recognized, P. gibbipennis Motschulsky, P. fuscus LeConte, P. punctatus Spinola and two sp. n., P. exutus  and P. texanus . Two syn. n. are proposed: P. cribratus Motschulsky, 1868 with P. gibbipennis Motschulsky, 1868 and P. crinitus Fall, 1907 with P. fuscus LeConte, 1850. A lectotype is selected for Paratenetus punctatus Spinola. A type species is designated for Storthephora M\u00e4klin, 1875 .The North American (north of Mexico) species of the tenebrionid genus  Paratenetus was proposed by Spinola in Paratenetus lebasi from Colombia and Paratenetus punctatus from the United States of America. The last mentioned species was represented by three specimens originating from the collection of Baron Dejean who received them from John Eatton LeConte. Subsequently, the genus received very little attention. In North America, John Lawrence LeConte described a new species in Paratenetus in 1868 from the material he collected in Georgia. In The genus The purpose of this paper is to review the American species occurring north of Mexico and provide a key for their identification.The study is based on the examination of about 3110 specimens borrowed from the following collections: Atlantic Forestry Centre, Fredericton, New Brunswick. Reginald P. Webster.AFC American Museum of Natural History, New York, New York. Lee H. Herman.AMNH The Natural History Museum, London, United Kingdom. Maxwell Barclay.BMNH California Academy of Sciences, San Francisco, California. David H. Kavanaugh.CAS Canadian Museum of Nature, Gatineau, Quebec. Fran\u00e7ois G\u00e9nier.CMN Canadian National Collection of Insects, Archnides and Nematodes, Ottawa, Ontario.CNC Cornell University Insect Collection, Ithaca, New York. James K. Liebherr.CUIC Department of Entomology, University of New Hampshire. Donald S. Chandler.DENH Department of Biology, Eastern New Mexico University, Portales, New Mexico. Darren A. Pollock.ENMU Florida State Collection of Arthropods, Gainesville, Florida. Paul E. Skelley.FSCPageBreak Georgia Museum of Natural History, The University of Georgia, Athens, Georgia. E. Richard Hoebeke.GMNH Gerard J. Hilchie Collection, Edmonton, Alberta.GHC Wallis-Roughley Museum of Entomology, University of Manitoba, Winnipeg, Manitoba. Robert E. Roughley.JBWM Janet Ciegler Collection, West Columbia, South Carolina.JCC Lyman Entomological Museum and Research Laboratory, McGill University, Sainte-Anne-de-Bellevue, Quebec. St\u00e9phanie Boucher.LEMM Louisiana State Arthropod Museum, Louisiana State University, Baton Rouge, Louisiana. Matthew L. Gimmel.LSAM Museum of Comparative Zoology, Harvard University, Cambridge, Massachusetts. Philip Perkins.MCZ Museo Regionale di Scienze Naturali, Torino, Italy. Luca Picciau.MRSN Northern Forestry Centre, Edmonton, Alberta. Greg R. Pohl.NFC Royal Alberta Museum, Edmonton, Alberta. Mark Steinhilber.RAM Royal British Columbia Museum, Victoria, British Columbia. Claudia Copley.RBCM Rolf L. Aalbu Collection, Sacramento, California.RLAC Royal Saskatchewan Museum, Regina, Saskatchewan. Ronald R. Hooper.RSM Reginald P. Webster Collection, Charters Settlement, New Brunswick.RWC Snow Entomological Museum, University of Kansas, Lawrence, Kansas. Zachary Falin.SEMC Texas A&M University, College Station, Texas. Edward G. Riley.TAMU Strickland Museum, University of Alberta, Edmonton, Alberta. Danny Shpeley.UASM Spencer Entomological Museum, University of British Columbia, Vancouver, British Columbia. Karen Needham.UBC National Museum of Natural History, Smithsonian Institute, Washington, DC. Warren E. Steiner.USNM Zoological Museum, Moscow University, Moscow, Russia. Nikolay B. Nikitsky.ZMMUThe photographs were made with a Leica Digital DC500 Imaging Workstation using Zerene Stacker software and retouched with Adobe Photoshop software.For type specimens, complete verbatim label data are given with additional information enclosed within quotation marks; individual labels are separated by a slash (/).http://www.simplemappr.net/).The distribution maps were generated using the software SimpleMappr . The idea for the name came from the peculiar shape of the palpi and particularly the flattening of the first two labial palpomeres.(based on species treated only). Body short, convex, pubescent; elytra with slanting setae in addition to erect setae. Epistoma with clypeolabral membrane exposed. Eyes present, prominent. Gena not sulcate. Antenna with last three antennomeres abruptly expanded, forming a distinct, loose club. Labial palpi short, penultimate palpomere swollen, last palpomere narrow, more or less fusiform; last maxillary palpomere large, at least twice as large apically than basally. Pronotum with sides denticulate, each denticle with one or two stiff setae; surface with relatively coarse punctures. Procoxae moderately separated. Mesepimeron not closing mesocoxal cavity. Elytra without striae, with relatively coarse punctures; epipleuron distinct and relatively wide up to apex. Abdomen with distinct membrane along posterior edge of ventrites 3 and 4. Intercoxal process of first ventrite relatively wide, more or less rounded apically. Tibia not expanded apically. Metatarsomere 1 elongate, as long as next two tarsomeres combined; penultimate tarsomere deeply lobate dorsally; last tarsomere not arising at apex of penultimate tarsomere. Tarsal claw simple, not pectinate. Tarsal formula 5\u20135-4. Defensive glands absent.This genus currently includes 57 species  ranging PageBreakParatenetus in his Cl\u00e9rites Coryn\u00e9to\u00efdes (currently Cleridae: Korynetinae). Cryptophagidae. Paratenetus back in the family Tenebrionidae, and placed it in the tribe Heterotarsini, a position that was followed by several authors including Paratenetus, Prataeus and Anaedus seem to be near the Lagriidae on account of the similarity in their larval stages\u201d and Heterotarsini (except Heterotarsus Latreille) from the tenebrionids to the lagriids based also on the morphology of the larvae. The study of the ovipositor structures by Paratenetus within the subfamily Lagriinae rather than the subfamily Tenebrioninae. Heterotarsini (except Heterotarsus) in the lagriine subtribe Lupropina of the tribe Adeliini. Paratenetus in the lagriid subfamily Lupropinae. Paratenetus in the lagriid subfamily Goniaderinae and Goniaderini. On the other hand, Lupropini following Paratenetus but we accept, following Goniaderini of the subfamily Lagriinae within the Tenebrionidae.We did not investigate the taxonomic position of the genus Paratenetus is poorly known. Many of the specimens seen in this study were collected in leaf litter in forested areas or in nests of the tent caterpillar genus Malacosoma (Lepidoptera: Lasiocampidae). All three winged species have been collected at black light. Paratenetus species pupate on the inner surfaces of rolled dead leaves (in which the larvae live) either hanging on fallen tree branches or on the ground.The biology of members of Paratenetus: erect and slanting. The slanting setae are characterized as subdepressed when the angle between the base of the seta and the elytra is between 10 and 40\u00b0, semierect when the angle is between 40 and 60\u00b0, and suberect when the angle is between 60 and 80\u00b0.There are two types of setae on the elytra of Motschulsky, 1868http://species-id.net/wiki/Paratenetus_gibbipennisParatenetus gibbipennis Motschulsky, 1868: 193. Type locality: \u00abAtlanta, G\u00e9orgie am\u00e9ricaine\u00bb .Paratenetus cribratus Motschulsky, 1868: 193. Type locality: \u00abG\u00e9orgie am\u00e9ricaine\u00bb . syn. n.Paratenetus gibbipennis. It bears the following labels: \u201c[green round disc] / [small brick red square label] / type [handwritten] / Paratenetus gibbipennis Motch Am. b. Mobile [handwritten on a rectangular green label].\u201d The specimen is intact although many of the setae on the pronotum and elytra are gone. The provenance of the specimen is doubtful. In the key to the Paratenetus in his collection, Motschulsky\u2019s collection at ZMMU contains a single specimen, a female, under the name Paratenetus cribratus. It bears the following labels: \u201c[green round disc] / Atlanta [handwritten] / type [handwritten] / Paratenetus cribratus Motch Am. bor. Atlanta [handwritten on a rectangular green label].\u201d The specimen is missing the left antennomeres 3\u201311 and the posterior legs.Motschulsky\u2019s collection contains a single specimen, a male, under the name Paratenetus gibbipennis and Paratenetus cribratus on the account that the first species has the lateral denticles of the pronotum very short while the second species has strong denticles. From an examination of the types, we cannot sustain Motschulsky\u2019s affirmation; the denticles are basically of the same size on both specimens.Paratenetus fuscus differ from the other three species treated by having the metaventrite very short. Paratenetus gibbipennis differs from Paratenetus fuscus by having few short erect setae on the elytra.This species and PageBreakly transverse. Pronotum with maximum width near midlength or slightly anterior to midlength; punctures moderately dense, not subcontiguous even over lateral half. Elytra very convex; slanting setae subdepressed, erect setae very few, short. Metaventrite short, length along midline clearly shorter than length of abdominal ventrite 2 along midline. Male protibia with calcar near middle along ventral surface; male mesotibia with short, in some specimens very short, more or less perpendicular preapical protuberance. Parameres with sides more or less parallel towards apex, apex not particularly acute . Rennie (CNC). Brandon (RBCM). Telford (NFC). Winnipeg (RBCM). Ontario. \u201cJonction Hwy 17 & 71\u201d (CNC). Bainsville (LEMM). Prince Edward Co. . Lancaster (LEMM). Chaffeys Locks Biol. Station (CNC). Alfred (CNC). Long Sault (CNC). 10 km W North Gower (CNC). Nepean (CNC). Belleville (CUIC). Thwartway Island, St. Lawrence Is. Nat. Park (CNC). Point Pelee (CNC). 2 km SE Spencerville (CNC). 4 km SW Kanata (CNC). Ottawa (CNC). Constance Bay (CMN). 4 km N of Westport (CNC). Campden (CNC). Rondeau Prov. Park (CNC). Arnprior (CNC). Erieau (CNC). 7 km W. Carleton Place (CNC). Blackburn (CNC). Normandale (CNC). Pelee Island (CNC). Hamilton (CNC). Flint Hill, nr Kemptville (CNC). Trenton (CNC). Toronto . Milton (CNC). Quebec. Montreal (CNC). Rigaud (CNC). Gatineau . Blind Lake, Gatineau Park (CNC). Gatineau Park (DENH). Hudson Heights (CNC). Oka (CNC). Ormstown (CNC). United States of America. Alabama.PageBreakMobile Co.: Mt. Vernon (CUIC). Monroe Co.: Haines Island Park, 3.5 mi. W Franklin (FSC). Connecticut.Fairfield Co.: Westport (AMNH). Litchfield Co.: Torrington (DENH); Canaan (TAMU); Cornwall . Georgia.Clarke Co.: 5 mi W Athens (GMNH); Whitehall Forest (GMNH). Rabun Co.: Tally Mill Crk. at Hwy 28 (CNC); Satolah (CNC). Illinois. \u201cN. Ill.\u201d (MCZ). \u201cIll.\u201d (USNM). Maine.Cumberland Co.: Portland (CNC). Kennebec Co.: Monmouth (MCZ). Oxford Co.: Paris (MCZ). Massachusetts. \u201cMass.\u201d (USNM). Bristol Co.: Swansea (MCZ); Somerset (MCZ); Dighton (MCZ); Fall River (MCZ). Hampshire Co.: Mount Tom (MCZ). Middlesex Co.: Waverly (USNM); Framingham ; Lowell (MCZ); Sherborn ; Hopkinton (CUIC); Cambridge ; Newton (MCZ). Norfolk Co.: Brookline ; Sharon (CUIC). Michigan.Marquette Co.: Marquette (USNM). Oakland Co. (CUIC). Wayne Co.: Detroit . Minnesota.Crow Wing Co.: Brainerd (CNC). Hennepin Co.: Lake Minnetonka (CUIC). Missouri.Saint Charles Co.: St. Charles (MCZ). Nebraska.Cuming Co.: West Point (USNM). New Hampshire.Grafton Co.: Franconia ; 0.5 mi S Rumney (DENH); Hanover (DENH). Rockingham Co.: Hampton ; Odiorne Point State Park (DENH). Strafford Co.: Somersworth (DENH); Durham (DENH); 3 mi. E Durham (DENH). New Jersey.Union Co.: Union ; Roselle (USNM); Elizabeth (USNM). New York. \u201cS[taten] I[sland]\u201d (MCZ). Dutchess Co.: Bulls Head (AMNH). Oswego Co.: North Pond (CUIC); Oswego (CUIC). Queens Co.: Flushing, L.I. (CUIC). North Carolina.Haywood Co.: Round Knob (USNM). Henderson Co.: 14 mi NW Hendersonville (SEMC). Macon Co.: 3 mi NW Highlands (DENH). Montgomery Co.: 2 mi S Eldorado (DENH). Yancey Co.: Black Mountains (AMNH). North Dakota.Richland Co.: Mirror Pool (USNM). Ohio. \u201cOhio\u201d (MCZ). Pennsylvania. \u201cPenn\u201d (MCZ). Allegheny Co.: Allegheny (USNM). Rhode Island. \u201cR.I.\u201d (USNM). South Carolina.Calhoun Co.: Wannamaker NP, St. Matthews (JCC). Chester Co.: Leeds (JCC). Edgefield Co.: Sumter Nat. For. (DENH). Lexington Co.: West Columbia (JCC). Newberry Co.: Billy Dreher Island State Park (JCC). Pickens Co.: Nine Times (JCC). Union Co.: Sedalia (JCC). Tennessee. \u201cChilhowee Mountain\u201d (CMN). Blount Co.: Rt. 129 just below rd. at The Narrows Overlook, GSMNP (LSAM); Ace Gap, GSMNP (LSAM). Cocke Co.: Gabes Mtn., GSMNP (LSAM). Sevier Co.: 0.5 mi W Laurel Falls Trailhead, GSMNP (LSAM); Twin Creeks, GSMNP (LSAM); Grapeyard Ridge (LSAM). Texas.Blanco Co.: Cypress Mill (USNM). Virginia.Giles Co.: Bald Knob, Mountain Lake (USNM); 9 km N Mountain Lake (USNM). Lee Co.: Pennington Gap (USNM). Wisconsin.Bayfield Co.: Bayfield (USNM). Dane Co.: Madison (TAMU). Shawano Co.: Tilleda (FSC).We have seen 660 specimens from the following localities. Canada. Females are much more abundant in collections than males. Of 183 specimens randomly selected, 8 were males (4.4%) and 175 were females (95.6%). The males came from Georgia (n=1), Alabama (n=6), and Missouri (n=1). No males were found among the 160+ randomly selected specimens from Canada and the northern states.Specimens were collected in January (n=1), February (n=1), March (n=89), April (n=64), May (n=8), June (n=61), July (n=20), August (n=95), September (n=31), October (n=38), November (n=6) and December (n=2).Labels on specimens read \u201cin leaf litter\u201d (6 specimens); \u201cin leaf litter of black birch and shrubs around and on areas of exposed rock\u201d (71); \u201cforest litter sifting\u201d (2); \u201cforest litter\u201d (3); \u201cmoist forest berlese\u201d (1).LeConte, 1850http://species-id.net/wiki/Paratenetus_fuscusParatenetus fuscus LeConte, 1850: 223. Type locality: Lake Superior (inferred from the title of the book).Paratenetus crinitus Fall, 1907: 253. Type locality: \u00abTrout Spring [New Mexico]\u00bb . syn. n.Paratenetus fuscus. It bears the following labels: \u201c[pale green round disc] / Type 4684 [partially handwritten on a red square label] / P. fuscus Lec. [handwritten].\u201d The specimen is intact.LeConte\u2019s collection at MCZ contains a single male specimen under the name PageBreakParatenetus crinitus from one specimen now at the MCZ. It bears the following labels: \u201cTrout sp. N.M. May [handwritten] / crinitus. Type [partially handwritten] / M.C.Z Type 24612 [red square label] / H.C. Fall Collection.\u201d The specimen is intact.Fall described Paratenetus crinitus and mentioned that \u201cin crinitus the metasternum is almost as short as in fuscus, which species is, however, very distinct by its subinflated elytra, more rounded sides of the prothorax and absence of erect hairs on the upper surface.\u201d Obviously Fall did not study the syntype in LeConte\u2019s collection since the specimen bears many erect hairs. LeConte never mentioned that character in his description and obviously Fall misidentified LeConte\u2019s species. We have studied the type specimens of both species and find no structural differences to separate them.Paratenetus gibbipennis by the character states listed in the description.This species differs from Paratenetus gibbipennis except for the following: slanting setae on elytra less depressed, semierect, occasionally even suberect; erect setae numerous, in seven or eight rows; metaventrite slightly longer, length along midline subequal to slightly shorter than length of abdominal ventrite 2 along midline.Same character states as This species ranges from Quebec City to the Rocky Mountains in northeastern British Columbia, north to southern Northwest Territories, south to northern New Mexico, northeastern Kansas, and Maryland .Alberta. \u201cTp. 74, Rge. 25, W. 5 Mer.\u201d (CNC). \u201cTp. 11, Rge. 1, W. 5 Mer.\u201d (CNC). Waterton Park (CNC). Calgary . Castor (UASM). Edmonton . Stettler (CNC). Cochrane (CNC). 30 km W Cochrane (CNC). Cypress Hills (CNC). Waiparous (CNC). Jumpingpound Creek (CNC). Sundre (CNC). Elkwater (CNC). British Columbia. North Pine (UBC). Pouce Coupe (UBC). Manitoba. \u201cTp. 9, Rge. 16, W. 1 Mer.\u201d (CNC). Aweme . Brandon (RBCM). Sandilands (JBWM). Birds Hill Prov. Park (ENMU). Husavik (CNC). Northwest Territories. Louise Falls, Hay River (CNC). Simpson (CAS). Ontario. Prince Edward Co. . Pelee Island (CNC). Ottawa (CNC). Constance Bay (CMN). Trenton (CNC). Arnprior (CNC). Quebec. Chelsea (CNC). Rigaud (CNC). Sainte-Croix-de-Lotbiniere (LEMM). Cap Rouge (CNC). Saskatchewan. Lac La Ronge (CNC). Morse (RSM). Rosefield (RSM). Oxbow (USNM). United States of America. Colorado.PageBreakBoulder Co.: Boulder . Custer Co. (USNM). Douglas Co.: Castle Rock (CNC). El Paso Co.: Colorado Springs (USNM). Jefferson Co.: Lookout Mountain (CUIC). Connecticut.Fairfield Co.: Westport (AMNH). District of Columbia. \u201cDC\u201d . Illinois. \u201cN. Ill.\u201d (MCZ). Champaign Co.: Urbana (USNM). Iowa. \u201cIowa\u201d . Johnson Co.: Iowa City . Story Co.: Ames (USNM). Kansas. \u201cKs\u201d (USNM). \u201cKans\u201d (USNM). Douglas Co.: Lawrence (CNC). Shawnee Co.: Topeka (USNM). Maryland. \u201cMd.\u201d (CNC). Anne Arundel Co.: 6 km ESE Laurel (USNM). Massachusetts. \u201cMass\u201d (USNM). Essex Co.: Lynn ; Salem (USNM). Middlesex Co.: Sherborn ; Tyngsboro (MCZ). Norfolk Co.: Brookline (MCZ). Michigan.Marquette Co.: Marquette (USNM). Montana. \u201cMont.\u201d (CUIC). \u201cMontana\u201d (USNM). Dawson Co.: Glendive (USNM). Powder River Co.: Fort Howes (USNM). Rosebud Co.: Colstrip (USNM). Nebraska. \u201cNeb.\u201d (USNM). Red Willow Co.: McCook (MCZ). New Mexico. \u201cN. Mex.\u201d (USNM). San Miguel Co.: Trout Spring (MCZ). New York. \u201cN.Y.\u201d . Westchester Co.: Peekskill (CUIC). New York Co.: Central Park, L.I. (USNM). North Dakota.Richland Co.: Mirror Pool (USNM). Rhode Island. \u201cR.I.\u201d (USNM). South Dakota.Jackson Co.: Cottonwood (RLAC). Tennessee. \u201cTenn.\u201d (MCZ). Vermont. \u201cVt.\u201d (MCZ). Wisconsin. \u201cWis\u201d (MCZ). Sauk Co.: Spring Green (USNM). Wyoming.Converse Co.: 11 mi N Douglas (CNC). Laramie Co.: Cheyenne (USNM).We have seen 305 specimens from the following localities. Canada. Females are a little more common in collections than males. Of 45 randomly selected specimens, 28 (62%) were females and 17 (38%) were males.Specimens were collected in February (n=1), March (n=30), April (n=58), May (n=8), June (n=40), July (n=7), August (n=19), September (n=14), October (n=5) and November (n=4).Spinola, 1844http://species-id.net/wiki/Paratenetus_punctatusLatridius pubescensnomen dubium]. Type locality: United States (inferred from title of the paper). Say, 1826: 265 \u201d; the second, a male \u201cParalectotype Paratenetus punctatus Spinola Ekis 74 [handwritten]\u201d; and the third, a female \u201cLectotype Paratenetus punctatus (Spinola) Ekis 74 [handwritten]\u201d. The first two specimens correspond neither to our concept of Paratenetus punctatus nor to any other North American species we have seen. The specimens are in poor condition, with almost all the setae gone, but they appear to be conspecific. Although Spinola indicated that all three of his specimens came from the United States and were provided by \u201cMr. [John Eatton] LeConte,\u201d these two specimens may have originated from Mexico, Central America or South America. The third specimen, a small individual (3.2 mm), fits our concept of Paratenetus punctatus and is here selected as lectotype. The label \u201cLectotype Paratenetus punctatus Spinola des. Y. Bousquet 2012\u201d has been attached to the specimen.Paratenetus punctatus can be separated from the other North American species of Paratenetus by their large size (3 mm or more). The vast majority of specimens of the other species are less than 3 mm long. Otherwise, the species can be separated from Paratenetus exutus in having the antennomere 8 subquadrate, the pronotum wider clearly anterior to the midlength, the punctation on the pronotum coarser, the slanting setae on the elytra slightly longer and more erect and the protibia of the male with a calcar along ventral surface. From Paratenetus texanus, this species is best separated in having the pronotum widest anterior to the midlength and the punctures on the pronotum coarser and denser, in part subcontiguous along the lateral half.Many specimens of Paratenetus gibbipennis and Paratenetus fuscus; slanting setae semierect in the vast majority of specimens, suberect in some specimens, erect setae few. Metaventrite long, length along midline longer than length of abdominal ventrite 2 along midline. Male protibia with calcar near middle along ventral surface; male mesotibia with very short, preapical spine, oriented perpendicularly or obliquely to long axis of tibia. Parameres with sides more or less parallel to very slightly convergent towards apex; apex more or less rounded . Victoria Beach (JBWM). New Brunswick. Jackson Falls, Carleton Co. (RWC). 10 km NW New River Beach, Charlotte Co. (AFC). 12 km SSE Upper Napan, Northumberland Co. (RWC). Cranberry Lake Protected Natural Area, Queens Co. (AFC). Acadia Research Forest, Sunbury Co. . Charters Settlement, York Co. (RWC). Canterbury, York Co. (RWC). 15 km W Tracy, York Co. (RWC). 14 km WSW Tracy, York Co. (AFC). Ontario. Ottawa (CNC). Constance Bay (CMN). Flint Hill, nr Kemptville (CNC). Ad & Lennox Co. . Pelee Island (CNC). Leamington (CNC). Hastings Co. (CNC). Walsingham (CNC). Prince Edward Co. . Point Pelee (CNC). Rondeau Prov. Pk. (CNC). Arnprior (CNC). Chaffeys Locks (CNC). Northumberland Co. (CNC). Sudbury (CNC). Hamilton (CNC). Gordon Island (St. Lawrence Is. Nat. Pk.) (CNC). 13 km W of Mattawa (CNC). Toronto (CUIC). 2\u20135 km W Mallorytown Landing (CMN). Quebec. Rouville (CNC). Fort Coulonge (CNC). Parc Provincial d\u2019Oka (CNC). Berthierville (CNC). Parc de la Gatineau (CNC). Laniel (CNC). Ile-du-Grand-Calumet (Pontiac) (CNC). Rigaud (CNC). Montreal PageBreak(CNC). Parc de la Yamaska (CNC). United States of America. Arkansas.PageBreakPageBreakGarland Co.: 3 mi W Crystal Springs (SEMC). Connecticut.Fairfield Co.: Westport (AMNH). Litchfield Co.: Litchfield (AMNH). New Haven Co.: Middlebury (USNM); Hamden (CUIC). District of Columbia. \u201cD.Col.\u201d (USNM). Takoma Park (USNM). Washington (USNM). Florida. \u201cFla\u201d (USNM). Alachua Co.: Gainesville (FSC); Newnans Lake (RLAC). Duval Co.: Jacksonville (USNM). Escambia Co.: Pensacola (FSC). Hillsborough Co.: Tampa (USNM). Indian River Co.: south of Vero Beach (FSC). Levy Co.: 4 mi SW Archer (FSC). Marion Co.: Juniper Springs (FSC); Rainbow Springs (FSC); Ocala National Forest . Palm Beach Co.: Lake Worth (CUIC). Polk Co.: Lake Marion Creek (GMNH). Putnam Co.: 2.5 mi NE Florahome (FSC); Crescent City (USNM); Welaka Exp. Sta. . Saint Johns Co.: St. Augustine (MCZ). Santa Rosa Co.: 4 mi N Munson (LSAM). Seminole Co.: Lake Mary (MCZ). Volusia Co.: South Daytona (CNC); Daytona (USNM). Georgia.Charlton Co.: 2.8 mi N Saint George (FSC). Johnson Co.: 1 mi E Kite (FSC); Suwanee Canal Rec. Area (FSC). Montgomery Co.: 5 mi W Uvalda (GMNH). Rabun Co. (MCZ); Clayton (MCZ). Tattnall Co. (FSC). Union Co.: \u201cHerbert Reece Park\u201d (GMNH). Illinois.Knox Co.: Galesburgh (MCZ). Macon Co. (FSC). Indiana.Allen Co.: \u201cSchoaf Park\u201d (USNM). Howard Co.: \u201cNW Howard Co.\u201d (LSAM). Jasper Co.: Jasper/Pulaski St. Forest (USNM). LaPorte Co.: Michigan City (USNM). Monroe Co.: Bloomington . Porter Co.: Dunes St. Pk. (RLAC). Tippecanoe Co. ; \u201cMcCormick Woods\u201d (USNM). Iowa. \u201cIowa\u201d (USNM). Johnson Co.: Iowa City (USNM). Polk Co.: \u201cBrown WDS Psv\u201d (CUIC); W. Saylorville Lake . Kansas. \u201cKans\u201d (USNM). Cherokee Co.: 2 mi S Galena (SEMC). Crawford Co.: Pittsburg (SEMC); 2 mi W Pittsburg (SEMC). Douglas Co.: 2 mi NW of Baldwin (SEMC). Jackson Co.: 6.5 km W Mayetta (SEMC). Jefferson Co.: 1 km SW Perry State Park (SEMC). Johnson Co.: Overland Park Arboretum (SEMC). Labette Co.: Big Hill Reservoir (SEMC). Neosho Co.: 2 mi SE Erie (SEMC). Sedgwick Co.: 0.5 mi S of Derby (SEMC). Shawnee Co.: S of intersection Woodring Rd & 69th St (SEMC). Kentucky. \u201cKy\u201d (USNM). Marshall Co. (FSC). Louisiana.Caddo Parish: Jacobs Nature Park (LSAM). Claiborne Parish: Corney Lake (CNC). East Feliciana Parish: Idlewild Exp. Station (LSAM). Livingston Parish: Livingston (LSAM). Natchitoches Parish: Kisatchie Nat. For. (LSAM). West Feliciana Parish: Saint Francisville (CMN); Tunica Hills, 0.5 mi W Weyanoke (LSAM). Maine.Androscoggin Co.: Wales (MCZ). Cumberland Co.: Casco (CUIC). Franklin Co.: Dead River (USNM); Farmington (USNM). Kennebec Co.: Augusta (DENH); Monmouth (MCZ). Oxford Co.: Rumford (USNM); Bethel (AMNH). Penobscot Co.: Lee (DENH); Passadumkeag (CUIC). Piscataquis Co.: Greenville (USNM). Washington Co.: Beddington (USNM). York Co.: West Lebanon (DENH). Maryland.Allegany Co.: Piclic Ridge, 5 km SE Pratt (USNM); Fifteen Mile Creek (RLAC). Anne Arundel Co.: 8 km ESE Laurel (USNM); 6 km ESE Laurel (USNM); Edgewater (USNM); 6 km S Edgewater (USNM); 3 km WSW Bristol at Jug Bay (USNM); Odenton . Baltimore Co.: 4 km SW Cockeysville (USNM); Catonsville (USNM). Calvert Co.: Plum Point (USNM). Carroll Co.: Eldersburgh (USNM). Cecil Co.: Pleasant Hill (USNM); Port Deposit (USNM). Frederick Co.: 2 mi W Thurmont (USNM). Garrett Co.: Rock Lodge, 4 km SW Bittinger (USNM); 7 mi N Oakland (USNM). Montgomery Co.: Kensington (USNM); Potomac (USNM); Rockville (USNM); Plummers Island (USNM); Great Falls (USNM); Hughes Hollow area, 5 km W Seneca (USNM). Prince Georges Co.: Cheverly (USNM); Bladensburg (USNM); Takoma Park (USNM); Laurel (USNM); Priest Bridge (USNM); Oxon Hill (USNM); Beltsville (USNM); Greenbelt (Park) (USNM); Bowie (USNM). Somerset Co.: Shelltown (USNM). Talbot Co.: 3 km SE Easton (USNM); Wittman (USNM); McDaniel (USNM). Worcester Co.: Assateague Island (USNM). Massachusetts.Barnstable Co.: Cape Cod (CNC). Bristol Co.: Dartmouth (MCZ). Essex Co.: Nahant (MCZ). Hampden Co.: Springfield (USNM). Middlesex Co.: Lincoln (MCZ); Sherborn (MCZ); Framingham (MCZ); Hopkinton (MCZ); Tyngsboro (MCZ); Natick (MCZ); Cambridge (MCZ). Norfolk Co.: Brookline (MCZ). Plymouth Co.: Marion (USNM). Suffolk Co.: Boston (MCZ); Jamaica Plain (CUIC). Michigan.Shiawassee Co.: Rose Lake Wldlf. Exp. Station (USNM). Wayne Co.: Detroit (USNM). Minnesota.Hennepin Co.: Minneapolis (CNC). Houston Co.: Winnebago Cr. Vy., 3\u20134 m NE Eitzen (USNM). Saint Louis Co.: Duluth (MCZ). Mississippi.George Co.: Lucedale (CUIC). Greene Co.: Leakesville (CUIC). Lauderdale Co.: Marion (MCZ). Missouri.Barry Co.: Mark Twain Nat. For. (FSC). Boone Co.: Ashland Wildlife Ar. (TAMU). Clay Co.: Cooley Lake (FSC). Greene Co.: near James River (TAMU). Jackson Co.: Raytown (FSC). Oregon Co.: Mark Twain Nat. For. (FSC). Randolph Co.: 1 mi E Moberly (TAMU). New Hampshire. \u201cN.H.\u201d (USNM). Coos Co.: Gorham (CNC); Mt. Washington . Grafton Co.: Mt. Moosilauke (MCZ); Bedell Bridge St. Pk. (DENH); Bath (DENH). Hillsborough Co.: Antrim (MCZ). Merrimack Co.: Concord (DENH). Strafford Co.: 1 mi SW Durham (DENH); Dover (DENH). New Jersey. \u201cN.J.\u201d (AMNH). Atlantic Co.: Buena (MCZ). Bergen Co.: Fort Lee (AMNH). Burlington Co.: Wharton State Forest (TAMU); Pemberton (USNM); 7 mi E Batsto (USNM). Cumberland Co.: Rutgers Exp. Sta. (USNM). Essex Co.: South Orange (MCZ); Eagle Rock (USNM); Montclair (USNM). Gloucester Co.: Malaga (USNM). Monmouth Co.: Highlands (USNM). Monroe Co.: Delaware Water Gap (USNM). Morris Co.: Boonton (USNM). Ocean Co.: Lakehurst . Orange Co.: Greenwood Lake . New York. \u201cS.I.\u201d (USNM). Albany Co.: Delmar (CUIC); Rensselaerville (USNM). Clinton Co.: vic. Taylor Pond Campground (GMNH). Erie Co.: Buffalo . Essex Co.: New Russia (CUIC); Whiteface Mt. (USNM). Orange Co.: Greenwood Lake (CUIC); Fort Montgomery (CUIC); West Point (USNM). Putnam Co.: Brewster (CUIC). Rockland Co.: Nyack (CUIC). Saint Lawrence Co.: Rossie (USNM). Seneca Co.: Willard (USNM). Suffolk Co.: Huntington, Long Island (DENH); Southold, L.I. (CUIC); Wyandanch, L.I. (USNM); Bellport (USNM); Yaphank (USNM). Tompkins Co.: Ithaca ; Dryden (CUIC). Yates Co.: Seneca Lake (USNM). North Carolina. \u201cN.C.\u201d (MCZ). \u201cRound Knob\u201d (USNM). Brunswick Co.: Southport (FSC). Buncombe Co.: 6 mi S Asheville (SEMC). Burke Co.: Linville Falls (CNC). Columbus Co.: Lake Waccamaw (USNM). Gates Co.: 6 km ENE Corapeake (USNM). Haywood Co.: Cove Creek (JCC); Cataloochee Divide ; 9 mi W Waynesville (SEMC). Henderson Co.: Fletcher (FSC). Jackson Co.: Balsam (USNM). Macon Co.: Nantahala Gap (CUIC); Highlands . Mitchell Co. (USNM). Moore Co.: Southern Pines (USNM). New Hanover Co.: Wilmington (USNM). Swain Co.: 2.5 mi NNE Cherokee, GSMNP (SEMC); Andrews Bald, GSMNP (LSAM); Ekaneetlee Gap, GSMNP (LSAM). Transylvania Co.: Lake Toxaway (AMNH). Watauga Co.: 3 mi NW Blowing Rock (TAMU). Yancey Co.: Black Mountains . Ohio.Ashland Co.: Mohican St. Pk. (FSC). Champaign Co.: Cedar Swamp (FSC). Ottawa Co.: Fishery Bay, S. Bass Isl. (CUIC). Union Co. (CUIC). Oklahoma.Latimer Co. ; 5 mi W Red Oak . Pennsylvania. \u201cPen\u201d . Allegheny Co.: Allegheny (CUIC). Dauphin Co.: Dauphin (CUIC). Lehigh Co.: Lehigh Gap (USNM). Luzerne Co.: Hazleton (MCZ). Montgomery Co.: Abington (MCZ). Philadelphia Co.: Frankford (USNM). Pike Co.: Twin Lakes (USNM). South Carolina. \u201cS.C.\u201d (MCZ). \u201cShiloh\u201d (JCC). Georgetown Co.: Sandy Island (JCC). Richland Co.: Pontiac (JCC). Tennessee. \u201cChimney Camp, Gt. Smoky Mts.\u201d (CUIC). Blount Co.: Cades Cove, GSMNP . Cocke Co.: Davenport Gap, GSMNP (LSAM). Sevier Co.: Goshen Prong, GSMNP (LSAM); Chimney Tops Picnic Nature Trail, GSMNP (LSAM); Roaring Fork, GSMNP (LSAM); Brushy Mnt., GSMNP (LSAM). Texas.Brazos Co.: College Station (TAMU). Vermont.Bennington Co.: Manchester (MCZ). Chittenden Co.: Burlington (USNM). Orange Co.: 12 mi E Chelsea (TAMU). Virginia. \u201cMiddletown\u201d (MCZ). \u201cFranklin Park\u201d (USNM). Arlington Co.: Glencarlyn (USNM). Fairfax Co.: Vienna (USNM); Black Pond (USNM); Great Falls (USNM); Great Falls N.P. near Clay Pond (USNM); Great Falls N.P. near quarry site (USNM). Giles Co.: Mountain Lake, Univ. Va. Biological Sta. (USNM). Lee Co.: Pennington Gap (MCZ). Loudoun Co.: Middleburg (USNM). Louisa Co.: Gum Spring (USNM). Montgomery Co.: Blacksburg (CUIC). Nelson Co. (USNM). Page Co.: Skyland . Rockbridge Co.: Natural Bridge (USNM). Shenandoah Co.: New Market (USNM). Warren Co.: 7 km NNE Linden, summit of Blue Mt. (USNM). Alexandria (USNM). West Virginia.Greenbrier Co.: W. Sulphur (USNM). Jefferson Co.: Harpers Ferry (USNM); Shepherdstown (USNM). Pocahontas Co.: Cranberry Glades (USNM). Preston Co.: Aurora (USNM). Wisconsin. \u201cWis\u201d (MCZ). Bayfield Co.: Bayfield (USNM). Douglas Co.: Bennett (USNM). Sauk Co.: Sauk City (GMNH). Shawano Co.: Tilleda (FSC). Wood Co.: Griffith State Nursery (USNM). Wyoming.Weston Co.: Newcastle (USNM).We have seen 1215 specimens from the following localities. Canada. Paratenetus fuscus but is easily separated by the coarse, irregular punctation on the pronotum and by the longer metaventrite.This species varies in regard to the punctation and setae. The punctation on the pronotum is coarse and in most specimens free on the disc and very close, in part subcontiguous over the sides; in some specimens the punctation is denser, being subcontiguous on the disc and contiguous all over the lateral sides. The slanting setae on the elytra are usually semierect but in some specimens they are less inclined and the erect setae are difficult to distinguish. The erect setae are usually short and moderately numerous but in some specimens, they can be relatively long or much more numerous; in such case the species can be confused with PageBreakFemales are more common in collections than males. Of 220 randomly selected specimens, 169 (77%) were females and 51 (23%) were males.Specimens were collected in March (n=6), April (n=89), May (n=296), June (n=384), July (n=152), August (n=67), September (n=40), October (n=9), November (n=5), and December (n=2).Malacosoma americana on Prunus serotina at mixed forest edge, shale barren area\u201d (7 specimens), \u201cshaken from and reared in moldy frass in old nest of Malacosoma americana on Prunus serotina\u201d (13), \u201cbeaten from dead leaf clusters on cut branches of Carpinus caroliniana at forest edge\u201d (6), \u201cbeaten from dead leaf clusters on branches of fallen Populus deltoides\u201d (7), \u201cbeaten from dead leaf clusters on fallen broken branch of Tilia americana in shade, mixed forest\u201d (2), \u201cbeaten from dead hanging leaf clusters on fallen Ailanthus in mixed forest\u201d (6), \u201cshaken from dead leaves on fallen branches of Quercus rubra\u201d (4), \u201cin moldy leaf clusters on fallen branch of Quercus alba in shade\u201d (6), \u201cbeaten from dead leaves of wind-blown Quercus rubra\u201d (1), \u201cbeaten from dead leaf clusters on fallen branches of Quercus rubra in mixed forest\u201d (14); \u201cat black light in longleaf pine and mixed oak, sand barrens\u201d (23), \u201cin moldy leaves on fallen branches of Acer rubrum\u201d (4), \u201cat black light in oak & longleaf pine sand barren\u201d (5); \u201cat black light; open sandy gap in mixed forest\u201d (1); \u201cat black light in mixed deciduous forest\u201d (1); \u201cat black light in mixed hardwood and loblolly pine forest\u201d (1); \u201cat black light in mixed pine and hardwood forest\u201d (3); \u201cbeaten from dead leaf clusters on branches of Castanea out ca. 2 weeks earlier\u201d (5); \u201cat black light near mixed forest, farmed fields and tidal creek\u201d (4); \u201cbeach drift\u201d (1); \u201cfrom pile of moldy thatch\u201d (1); \u201cin moldy leaf clusters on cut branches of Prunus serotina\u201d (4); \u201cin moldy leaf clusters on cut branches of Morus\u201d (2); \u201cbeaten from dead leaf clusters on cut branches of Acer rubrum at mixed forest edge\u201d (10); \u201cin old nest of Malacosoma on Prunus\u201d (2); \u201cin dead leaves on branches of fallen oak\u201d (1); \u201cshaken from dead leaves on cut Sassafras\u201d (7); \u201cbeaten from dead leaf clusters on fallen branch Acer negundo at mixed forest edge\u201d (1); \u201cat black light in tree canopy, mixed broken forest and residential area\u201d (43); \u201cat black light in mixed hardwood forest near pond and river\u201d (8); \u201cat black light in mixed forest, bluff above river\u201d (2); \u201cin old tent Malacosoma americana\u201d (5); \u201cat black light\u201d (2); \u201cin old tent nest of Malacosoma americana with moldy frass, on Prunus serotina\u201d (1); \u201cshaken from dry leaf (Vitis sp.) nest of Sciurus carolinensis in vine tangle ca. 3 m above ground\u201d (1); \u201cat black light at edge of clearing in mixed forest near drying vernal pool\u201d (2); \u201cat black light in mixed forest near vernal pools\u201d (8); \u201cat black light sheet in open mature mixed forest near river\u201d (3); \u201cbeaten from dead leaf clusters on fallen branch of Liriodendron in mixed forest\u201d (9); \u201cbeaten ex spruce\u201d (1); \u201ccollected in tents Malacosoma americana\u201d (12); \u201cin web of Malacosoma\u201d (3); \u201con Pinus strobus\u201d (2); \u201cex. canopy trap\u201d (34); \u201cintercept trap\u201d (1); \u201cbeating dead leaves\u201d (8); \u201cbtng oak blowdown\u201d (2); \u201cleaf litter\u201d (1); \u201cdead moldy leaves\u201d (1); \u201cbeating veg.\u201d (2); \u201cbeating flowers\u201d (2).Labels on specimens read \u201cin overwintered nest remains of PageBreakBousquet & Bouchardsp. n.http://zoobank.org/E79EDDDF-59F8-4A43-880F-2A86CF8EB2F2http://species-id.net/wiki/Paratenetus_exutusHolotype (\u2642) labeled \u201cTabusintac, N.S. 20-VI-1939 W.J. Brown / Holotype Paratenetus exutus Bousquet & Bouchard CNC No. 24035.\u201d The specimen is deposited in the CNC.Manitoba. Ninette, 31-V-1958, J.F. McAlpine ; same locality, 30-V-1958, R.B. Madge . New Brunswick. Tabusintac, 19-VI-1939, W.J. Brown ; same data but 20-VI. 1939 or 22-VI-1939 . York Co., 14 km WSW of Tracy, S of Rt 646, 45.6741\u00b0N, 66.8161\u00b0W, 26 April-10 May 2010, R. Webster & C. MacKay coll. . York Co., 15 km W of Tracy off Rt. 645, 45.6848\u00b0N, 66.8821\u00b0W, 19\u201325 May 2009, R. Webster & M.-A. Gigu\u00e8re coll. . York Co., New Maryland Charters Settlement, 45.8430\u00b0N, 66.7275\u00b0W, 12 July 2005, R. P. Webster coll. ; same locality but 45.8340\u00b0N, 66.7450\u00b0W, 30 April 2005 . Queens Co., Cranberry Lake P.N.A., 46.1125\u00b0N, 65.6075\u00b0W, 24 April-5 May 2009, R. Webster & M.-A. Gigu\u00e8re coll. ; same locality but 3\u201313 May 2011, M. Roy & V. Webster coll. . Carleton Co., Jackson Falls, \u201cBell Forest\u201d, 46.2200\u00b0N, 67.7231\u00b0W, 28.April-9 May 2009, R. Webster & M.-A. Gigu\u00e8re coll. . Carleton Co., Wakefield Meduxnekeag Valley Nature Preserve, 46.1890\u00b0N, 67.6766\u00b0W, 8 June 2005, M. Gigu\u00e8re & R. Webster coll. ; same locality but 46.1935\u00b0N, 67.6825\u00b0W, 19 April 2995 . Albert Co., Shepody N.W.A., Germantown Section, 45.7101\u00b0N, 64.7542\u00b0W, 30 July 2004, R.P. Webster coll. . Sunbury Co., Acadia Research Forest, 45.9866\u00b0N, 66.3841\u00b0W, 8\u201313 May 2009, 13\u201319 May 2009, 19\u201325 May 2009, 16\u201324 June 2009, R. Webster & M.-A. Gigu\u00e8re coll. . Nova Scotia. St. Peters, 25-VII-1930, M.L. Prebble . Ontario. Alfred bog, 16.VI.1981, A. Davies . Quebec. Sainte-Catherine Portneuf, 29-VIII-1971, Claude Chantal . D[ivision de] R[ecensement] Bellechasse, St-N\u00e9r\u00e9e, 10.VII.1976, J.F. Landry . Cascapedia, 11.VI.1933, W.J. Brown .Paratypes from the following localities: exutus, -a, -um (deprived of) and alludes to the fact that the protibia of the male lacks the spinelike projection  found in the other American (north of Mexico) species.The specific name comes from the Latin participle Paratenetus punctatus and Paratenetus texanus in having the antennomere 8 transverse. The males are also easily recognized among the species treated here in having no calcar on the protibia and a relatively long apical spine, oriented more or less parallel to long axis of tibia, on the mesotibia.This species is best separated from PageBreaktwo abdominal ventrites in the vast majority of specimens, not or only slightly darker in a few specimens. Antennomere 8 transverse. Pronotum with maximum width at or very slightly anterior of midlength . Peace River (NFC). Manitoba. Aweme (CNC). New Brunswick. Fredericton (CNC). Nova Scotia. Kentville (CNC). Annapolis Royal (CNC). Portapique (MCZ). St. Peter\u2019s (AFC). Cape Breton . Grand River . Woodside (AFC). White Point Beach, Queens Co. (JCC). Ontario. Ridgeway (MCZ). Trenton (CNC). Prince Edward Co. (CNC). La Rose Forest, near Bourget (CNC). Quebec. Hull [= Gatineau] (CAS). Lac Duparquet (LEMM). Lac Labyrinthe [Abitibi] (LEMM). Laniel (CNC). Valcartier (CNC). Saskatchewan. Red Earth (RSM). Somme (RSM). United States of America. Alabama.PageBreakPageBreakConecuh Co.: 19 km NE Evergreen (USNM). Arkansas.Newton Co.: 12 mi. W Jasper (SEMC). Connecticut.Litchfield Co.: Cornwall . District of Columbia. Washington (USNM). Florida. \u201cFla\u201d (USNM). \u201cHaulover\u201d (USNM). Alachua Co.: Cross Creek (FSC); Gainesville (RLAC); nr. Paynes Prairie St. Pk. (FSC). Brevard Co.: Hatbill St. Pk. (FSC). Dade Co.: Everglades Nat. Pk. \u201cRoyal Palm Pk.\u201d (CMN); Everglades Nat. Pk., Royal Palm Hammock (FSC). Highlands Co.: Archbold Biological Station (TAMU). Lake Co.: Camp McQuarrie (FSC). Liberty Co.: Torreya St. Pk. (FSC). Putnam Co.: 2 mi. SW Interlachen (FSC). Volusia Co.: Enterprise (USNM). Illinois.Lake Co.: Grayslake (SEMC). Indiana.Monroe Co.: Bloomington (FSC). Iowa.Buchanan Co.: Independence (USNM). Polk Co.: Walnut Woods St. Pk. . Kansas.Bourbon Co.: 9 mi SW Ft. Scott (SEMC). Crawford Co.: 3 mi NE Pittsburg (SEMC). Jefferson Co.: 1 km SW Perry State Park (SEMC); University of Kansas Field Station, Nelson Ravine Forest (SEMC); The Falin Property, 1.5 km N jct. 94th St. & Kingman Rd. (SEMC). Marshall Co.: Alcove Springs State Park (SEMC). Neosho Co.: 2 mi SE Erie (SEMC). Osage Co.: Melvern Lake Project, Outlet Park (SEMC); Pomona Lake, Outlet Park (SEMC). Pottawatomie Co.: St. George (SEMC). Wabaunsee Co.: 10 mi SW Alma (SEMC). Kentucky. \u201cKy\u201d (USNM). Lousiana.East Baton Rouge Parish: LA 37 at Comite River (LSAM). East Feliciana Parish: Boy Scout Camp Avondale, E of Clinton (LSAM); 1.2 mi S Central (LSAM). Maine.Aroostook Co.: St. Francis (DENH); Crystal (USNM); Howe Brook (USNM); Portage (USNM); Clayton Lake (USNM); Ashland (USNM). Cumberland Co.: South Portland (CUIC). Franklin Co.: Oquossoc (DENH). Hancock Co.: Blue Hill (DENH); E. Orland (USNM). Kennebec Co.: Vassalboro (USNM); Augusta (USNM). Knox Co.: Friendship (USNM). Lincoln Co.: New Harbor (USNM); Bristol (USNM); Boothbay Harbour (USNM). Oxford Co.: Peru . Penobscot Co.: Lee (USNM); Springfield (USNM). Piscataquis Co.: Kokadjo (DENH); Dover-Foxcropt (DENH); Chesuncook (USNM). Somerset Co.: Caratunk ; Embden (USNM); Bingham (USNM); Brighton (DENH); Rockwood (USNM); Seboomook (DENH). Waldo Co.: Palermo (USNM). Washington Co.: Princeton ; Wesley ; Steuben (CNC). York Co.: West Lebanon (DENH). Maryland.Carroll Co.: Finksburg (USNM). Somerset Co.: Shelltown (USNM). Talbot Co.: Wittman (USNM); 3 km SE Easton (USNM). Michigan.Marquette Co.: Marquette (USNM). Wayne Co.: Detroit (USNM). Minnesota.Becker Co.: Itasca St. Pk. area (USNM). Crow Wing Co.: Lake Hubert (CNC). Sherburne Co.: Elk River (CNC). Mississippi.George Co.: Lucedale (CUIC). Greene Co.: Leakesville (CUIC). Missouri.Greene Co.: nr. James River (TAMU). Randolph Co.: 1 mi E Moberly (TAMU). New Jersey.Atlantic Co.: 5 mi. N Hammonton (RLAC). Cape May Co.: Anglesea (USNM). Ocean Co.: Lakehurst (CUIC). Salem Co.: Lake Hudson, near Deepwater (RLAC). Union Co.: Elizabeth (USNM). New York.Suffolk Co.: Yaphank, L.I. (USNM). Ulster Co.: West Park (CUIC); Slide Mt. (CUIC). North Carolina.Buncombe Co.: Oteen (USNM); 6 mi S Asheville (SEMC). Haywood Co.: 9 mi. W Waynesville (SEMC); Cataloochee, GSMNP (LSAM); Purchase Knob, GSMNP (LSAM). Henderson Co.: Fletcher (FSC). Swain Co.: Andrews Bald, GSMNP (LSAM); Clingmans Dome, GSMNP (LSAM). Yancey Co.: Black Mountains (AMNH). North Dakota.Richland Co.: Mirror Pool (USNM). Ohio.Fairfield Co.: Barnebey Center (RLAC). Franklin Co.: Worthington (RLAC). Hamilton Co.: Cincinnati (USNM). Highland Co. (FSC). Hocking Co.: Ward Township (RLAC). Pike Co.: Jackson Lake (RLAC). Preble Co.: Hueston Woods (RLAC). Ross Co.: Tar Hollow St. Pk. (FSC). Trumbull Co.: Phalanx (CUIC). Oklahoma.Latimer Co.: Red Oak . Pennsylvania.Fayette Co.: 5 mi. W. Ohiopyle (USNM). Tennessee.Cocke Co.: Albright Grove (LSAM). Sevier Co.: Ramsey Cascade Trail, GSMNP (LSAM); Goshen Prong, GSMNP (LSAM); Indian Gap, GSMNP (LSAM). Swain Co.: near Charlies Bunion, GSMNP (FSC). Texas.Colorado Co.: Columbus (USNM). Victoria Co.: Victoria (USNM). Virginia. \u201cFt. Monroe\u201d (USNM). Covington (FSC). Bath Co.: 9.6 km N Clifton Forge (CNC). Lee Co.: Pennington Gap (MCZ). Loudoun Co.: 3 km SE Lovettsville (USNM). Montgomery Co.: Caldwell Fields . West Virginia.Mingo Co.: Justice (CUIC). Pocahontas Co.: Cranberry Glades (USNM). Wisconsin.Bayfield Co.: Bayfield (USNM). Wood Co.: Griffith State Nursery (USNM).We have seen 416 specimens, including the type material, from the following localities. Canada. While almost all specimens from Canada and northern United States had the metaventrite distinctly darker than the first two abdominal ventrites, this is not the case with the specimens from the southern states. There is also variation in the width of the antennomere 8. Most specimens have that antennomere distinctly transverse, some specimens from the southern states (particularly Louisiana) have the antennomere 8 only slightly transverse.Females are more common in collections than males. Of 105 randomly selected specimens, 76 (72%) were females and 29 (28%) were males.Specimens were collected in March (n=9), April (n=38), May (n=84), June (n=58), July (n=79), August (n=40), September (n=22), October (n=5), November (n=3), and December (n=2).Acer rubrum\u201d (3); \u201cbeaten ex spruce\u201d (35); \u201cbeaten ex fir\u201d (10); \u201con Bumelia lanuginosa\u201d (1); \u201cex. spruce\u201d (1); \u201cex. canopy trap\u201d (15); \u201cex. FIT, near upper meadow\u201d (1); \u201cex. FIT, near lower meadow\u201d (3); \u201cex. canopy malaise, near lower meadow\u201d (9); \u201cex. canopy FIT, near lower meadow\u201d (3); \u201cmalaise trap\u201d (6).Labels on specimens read \u201cat black light near mixed forest, farmed fields and tidal creek\u201d (4 specimens); \u201cat black light at edge of mixed forest and open turf on hill\u201d (1); \u201cin moldy leaf clusters on cut branches of Paratenetus inermis Bsq. and Bouch.\u201d since it was the intended name. Unfortunately, we realized that the name was already used by Champion only after the specimens were returned to their respective collections.Most specimens of this species in collections are identified under the name \u201cPageBreakPageBreakBousquet & Bouchardsp. n.http://zoobank.org/E4FC7175-796F-4270-966D-93B4EE8E681Ahttp://species-id.net/wiki/Paratenetus_texanusHolotype (\u2642) labeled \u201cPort Isabel, Tex. 20.X.1982 Lot 2 BF&JL Carr / Holotype Paratenetus texanus Bousquet & Bouchard CNC No. 24133.\u201d The specimen is deposited in the CNC.Texas. Port Isabel, 17.X.1982, 20.X.1982, 30.III.1987, BF&JL Carr . 18 mi. E of Hebbronville, 25.III.1987, BF&JL Carr . Cameron Co., Brownsville, 19 July 1981, W.E. Steiner . Cameron Co., Palmito Hill Hist. Site, Hwy. 4 east of Brownsville, 12-X-1993, S.M. Clark . Cameron Co., 11 mi. W Boca Chica, 28 Sept. 1976, R. Turnbow . Hidalgo Co., Mission, Bentsen State Park, 17 (or 18) July 1981, W.E. Steiner . Hidalgo Co., Anzalduas Co. Pk., 19 Oct. 1985, Wappes & Downie . Bee Co., Beeville, 19 June 1974, W.E. Steiner .Paratypes from the following localities: The specific name derives from the name of the state of Texas where the species has been commonly collected.Paratenetus punctatus and Paratenetus exutus in having the punctures on the pronotum sparser, not subcontigous even on the lateral half. They can also be distinguished from most adults of Paratenetus punctatus by their smaller size and from most adults of Paratenetus exutus by the subquadrate antennomere 8 and metaventrite of same color as the first two abdominal ventrites.Members of this species can be distinguished from those of Paratenetus gibbipennis and Paratenetus fuscus; slanting setae subdepressed, erect setae short. Metaventrite long, length along midline longer than length of abdominal ventrite 2 along midline. Male protibia with calcar near middle along ventral surface; male mesotibia with short, preapical spine, wide at base and oriented perpendicularly to long axis of tibia. Parameres with sides distinctly convergent towards apex; apex markedly acute . Louisiana.Avoyelles Parish: Mansura (USNM). Cameron Parish: Holly Beach ; nr. Oak Grove (TAMU). Texas. \u201c60 mi SE Cotulla\u201d (CNC). \u201c15 mi SW Jct FR 3073 & Hwy 16\u201d (CNC). Anderson Co.: Elkhart (TAMU). Aransas Co.: Goose Island St. Park . Atascosa Co.: Pleasanton (USNM); Campbellton (TAMU). Bastrop Co.: Bastrop St. Pk. (FSC). Bee Co.: Beeville (USNM); Pettus (CNC). Bexar Co.: San Antonio (USNM). Brooks Co.: Falfurrias (CNC); 9 mi W Falfurrias (TAMU). Cameron Co.: Boca Chica ; 6 mi W Boca Chica Beach (TAMU); 6.7 mi W Boca Chica Beach (TAMU); Brownsville ; 4 mi ESE Brownsville (TAMU); 6 mi E Brownsville (TAMU); 10 mi E Brownsville ; 12.5 mi E Brownsville (TAMU); 13.5 mi E Brownsville (TAMU); W of Harlingen (TAMU); Main Reservoir near Brownsville (RLAC); Resaca de las Palomas St. Pk. (RLAC); Resaca de La Palma St. Pk. (TAMU); Sabal Palm Grove Wildlife Sanctuary ; nr. Southmost (USNM); ca. 2 mi E Los Fresnos (TAMU); Laguna Atascosa NWR (TAMU); 9.7 mi E jct Rt 1419 on hwy 4 (TAMU). Chambers Co.: Anahuac (USNM). Duval Co.: San Diego (USNM); Freer (TAMU); Sepulveda Ranch (TAMU); 3.5 mi S Realitos (TAMU). Fort Bend Co.: Brazos Bend St. Pk. (TAMU). Galveston Co.: Virginia Point (USNM); San Luis Pass (TAMU); 3.5 mi SW Jamaica Beach (TAMU); 7 mi SW Jamaica Beach (TAMU). Goliad Co.: Goliad (USNM). Hidalgo Co.: Santa Ana Nat. Wdlf. Ref. ; Bentsen Rio Grande Valley St. Pk. ; Anzalduas Park (TAMU); Delta Lake (TAMU). Jefferson Co.: 10 mi W Sabine Pass (TAMU). Jim Wells Co.: Ben Bolt (CNC); 1 mi N Ben Bolt (TAMU); Alice (USNM); 5 km W Alice (CMN); 1 mi N Premont (TAMU); 1.4 mi S Premont (TAMU). Karnes Co.: 1 mi NE Runge (TAMU). Kendall Co.: Boerne (USNM). Kenedy Co.: Sarita (CNC); 2 mi S Sarita (TAMU); 13 mi S Sarita (TAMU); 25.3 mi S Sarita (FSC); 31.8 mi S Sarita (TAMU); Armstrong (CNC); 1 mi S Armstrong (TAMU); Norias (TAMU); 5 mi N Norias (TAMU); 6 mi S Norias (TAMU); 8 mi S Norias (CNC); Loyola Beach, Baffin Bay (CNC); Baffin Bay (TAMU). Kleberg Co.: Kingsville ; Riviera ; Riviera Beach (CMN); Velederos Creek (TAMU). Live Oak Co.: 17 mi SW George West (TAMU). Nueces Co.: Corpus Christi . Refugio Co.: 8 mi E Refugio (TAMU); 7 mi S Woodsboro (TAMU). San Patricio Co.: Sinton (USNM); nr. Sinton (CNC); 3 mi N Sinton (TAMU); 7 mi N Sinton (TAMU); Welder Wildlife Refuge ; Welder Wildlife Refuge, 17 km NE Sinton (CMN); Lake Corpus Christi St. Pk. (LSAM). Starr Co.: 1.5 m E Rio Grande City (LSAM). Tyler Co.: 4 mi E Spurger (TAMU). Willacy Co.: 8 miles SW Port Mansfield (TAMU). Mexico. Chiapas. El Aguacero, 16 km W Ocozocoautla (CMN); 5 km E Ocozocoautla (CMN); 2 km S Chicoasen (CMN); Cinco Cerros (CMN). Nayarit. 15 mi N Tepic (CNC). Tamaulipas. Mpio.San Carlos, Cerro del Diente (TAMU).We have seen 515 specimens, including the type material, from the following localities. United States of America. The two specimens from Miami in Florida externally agree perfectly with those from Texas. One is a male and its genitalia are identical to those of specimens from Texas.Males are more common in collections than females. Of 106 randomly selected specimens, 42 (40%) were females and 64 (60%) were males.Specimens were collected in January (n=1), February (n=1), March (n=65), April (n=39), May (n=89), June (n=30), July (n=53), August (n=21), September (n=36), October (n=108), November (n=2), and December (n=6).Prosopis and Celtis forest, sandy soil\u201d (6 specimens); \u201con Celtis\u201d (1); \u201cex dry okra pod\u201d (1); \u201ccotton\u201d (1); \u201ccollected on Celtis\u201d (2); \u201cfallen fruit Yucca treculeana\u201d (1); \u201con flower Yucca treculeana\u201d (2); \u201con Acacia Berlandieri Benth.\u201d (1).Labels on specimens read \u201cat black light in PageBreakPageBreakParatenetus constrictus, Paratenetus corticarioides, Paratenetus nigricornis, Paratenetus punctulatus, Paratenetus tibialis, and Paratenetus villosus, and none of them are conspecific with those of Paratenetus texanus. The three species not seen are Paratenetus tropicalis Motschulsky, Paratenetus koltzei Pic, and Paratenetus mexicanus Pic.This new species occurs in Mexico and nine species have been reported from that country. We have examined the type material of the six species described by Champion and housed in BMNH, i.e.,"}
+{"text": "Protective effects of boswellic acid (BA) against acetaminophen (APAP)-induced hepatotoxicity in Balb/ cA mice were examined. BA, at 0.05 or 0.1%, was supplied for 4 weeks. Acute liver injury was induced by APAP treatment. Results showed that BA intake increased hepatic BA bioavailability. APAP treatment decreased glutathione (GSH) level, increased reactive oxygen species (ROS) and oxidized glutathione (GSSG) production; and lowered activity and protein expression of glutathione reductase (GR) and heme oxygenase (HO)-1 in liver. BA intake at both doses alleviated subsequent APAP-induced oxidative stress by retaining GSH content, decreasing ROS and GSSG formations, reserving activity and expression of GR and HO-1 in liver, and lowering hepatic cytochrome P450 2E1 activity and expression. APAP treatment enhanced hepatic levels of interleukin-6, tumor necrosis factor-alpha and monocyte chemoattractant protein-1. BA pre-intake diminished APAP-induced release of those inflammatory cytokines and chemokines. APAP upregulated hepatic protein expression of toll-like receptor (TLR)-3, TLR-4, MyD88, nuclear factor kappa B (NF-\u03baB) p50, NF-\u03baB p65 and JNK. BA pre-intake at both doses suppressed the expression of NF-\u03baB p65 and p-JNK, and only at 0.1% down-regulated hepatic TLR-3, TLR-4 and MyD88 expression. APAP led to obvious foci of inflammatory cell infiltration in liver, determined by H&E stain. BA intake at both doses attenuated hepatic inflammatory infiltration. These findings support that boswellic acid is a potent hepatoprotective agent. Acetaminophen  is a widely used analgesic and antipyretic drug. It is metabolized by hepatic cytochrome P450 system, especially CYP2E1, which leads to the overproduction of reactive free radicals and n-acetyl-p-benzoquinoneimine (NAPQI) , 4. In aet al. [et al. [Toll-like receptors (TLRs) are pattern recognition receptors mainly responsible for immuno modulation. The activation of TLR-3 and TLR-4 has been implicated in APAP-induced hepatic damage because both TLRs could stimulate the production of adaptor proteins including MYD88 and nuclear factor kappa B (NF-\u03baB), which in turn promote the generation of oxidants and inflammatory cytokines and chemokines , 8. Furtet al.  reportedBoswellia species including Boswellia carteri, Boswellia serrata and Boswellia sacra [via its anti-oxidative and anti-inflammatory activities [et al. [Boswellic acid  was synthesized and provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, China. APAP (98%) was purchased from Sigma Chemical Co. .Five- to six-week-old male Balb/cA mice were obtained from National Laboratory Animal Center . Mice were housed on a 12-h light-12-h dark schedule, and fed with water and mouse standard diet . Use of the mice was reviewed and approved by the China Medical University animal care committee (104-305).BA at 0.05 or 0.1 g was mixed with 99.95 or 99.9 g powder diet to prepare 0.05 and 0.1% BA diets. Mice were divided into five groups: normal group ; BA group (0.1% BA diet), APAP group ; BA-low- APAP group (0.05% BA diet plus APAP treatment); BA-high- APAP group (0.1% BA diet plus APAP treatment). After 4 weeks supplement, normal and BA groups were sacrificed, and the other three groups were treated by a single intraperitoneal injection of APAP (400 mg/kg body weight). APAP was dissolved in phosphate-buffered saline (PBS). Mice were killed with carbon dioxide after 24 h. The mortality of mice due to APAP injection was zero. After sacrificed, liver from each mouse was collected and weighted. There was no mice die before sacrificed. Blood was also collected, and serum was separated from erythrocyte immediately. Liver tissue, 0.2 g, was homogenized on ice in 2 ml phosphate buffer (pH 7.2), and the filtrate was collected. The protein concentration of serum and liver filtrate was determined by a commercial assay kit  with bovine serum albumin as standard.et al. [The HPLC method of Lozano-Mena et al.  was usedSerum activities of ALT and AST were determined by using commercial assay kits . CRP level (mg/l) was measured by an ELISA kit .Liver tissue was homogenized with cold PBS containing 0.05% Tween 20 and 1 mM EDTA. After centrifuging, supernatants were used for measurements. GSH and GSSG concentrations (nmol/mg protein) in liver were determined by commercial colorimetric GSH and GSSG assay kits . ROS level was quantified by using 2\u2019, 7\u2019-dichlorofluorescein diacetate. Fluorescence value was measured by using a fluorescence microplate reader at excitation and emission wavelengths of 485 and 530 nm, respectively. Relative fluorescence unit (RFU) was the difference in fluorescence values obtained at time 0 and 5 min. Results are expressed as RFU/mg protein. The activity (U/mg protein) of GPX and GR was assayed by using commercial kits purchased from EMD Biosciences Co. .Hepatic levels of IL-6, TNF-alpha and MCP-1 were measured by using cytoscreen immunoassay kits . The sensitivity of assay with the detection limit was 5 pg/ml for IL-6, and 10 pg/ml for TNF-alpha and MCP-1.The activity of CYP2E1 in freshly prepared liver microsome was estimated by colorimetrically measuring the formation of 4-nitrocatechol, a product from p-nitrophenol hydroxylation catalyzed specifically by CYP2E1. The protein concentration of CYP2E1 was measured by ELISA, and a rabbit anti-CYP2E1 antibody  was used for detection. The formed 4-nitrocatechol was expressed as nmol/mg protein.Hepatic tissue, 40 mg, was homogenized in buffer containing protease-inhibitor cocktail purchased from Sigma-Aldrich Chemical Co.  and 0.5% Triton X-100. Homogenate was then mixed with buffer , and followed by boiling for 5 min. Protein sample at 40 \u03bcg was electrophoresed on 10% SDS-polyacrylamide gel, and further transferred onto nitrocellulose membranes  for 1 h. After blocking with a protein solution containing 5% skim milk for 1 h, membranes were treated with monoclonal antibody  against heme oxygenase (HO)-1, CYP2E1 (1:1000), NF-\u03baB p50, NF-\u03baB p65, JNK (1:500), TLR-3, TLR-4, MyD88 (1:2000) at 4\u00baC overnight, and followed by incubating with horseradish peroxidase-conjugated antibody at room temperature for 3.5 h. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as a loading control, and the bands were quantified by an ATTO image analyzer .Partial liver tissue from each mouse was fixed in 10% phosphatebuffered formalin, and embedded in paraffin. Paraffin section at 5 mm thickness was cut and processed with hematoxylin-eosin (H&E) stain, and followed by examining under a light microscope for histological analysis. The severity of hepatic inflammatory injury was assayed according to the Ishak scoring system . The injThe effect of each treatment was analyzed from 10 mice (n = 10) in each group. Data were reported as means \u00b1 standard deviation (SD), and subjected to analysis of variance. Differences among means were determined by the Least Significance Difference Test with significance defined at P < 0.05.P > 0.05, Table P > 0.05). The intake of BA at both doses alleviated subsequent APAP-induced elevation of ALT, AST and CRP levels in serum (P & 0.05).BA intake at 0.1% increased hepatic BA content, and did not affect body weight, feed intake, water intake and liver weight (P & 0.05); but BA pre-intake attenuated APAP-induced GSH depletion and decreased hepatic GSSG and ROS production (P & 0.05). APAP raised GPX activity and reduced GR activity in liver (P & 0.05). BA intake failed to change hepatic GPX activity (P > 0.05); but significantly maintained hepatic GR activity (P & 0.05). As shown in Figure P & 0.05). BA intake did not affect GPX protein expression (P & 0.05); but significantly retained protein expression of GR and HO-1 in liver (P & 0.05). APAP treatment enhanced CYP2E1 activity and expression ; however, BA intake significantly suppressed subsequent APAP-induced elevation of CYP2E1 activity and protein expression (P & 0.05). APAP treatment also significantly increased hepatic release of TNF-alpha, IL-6 and MCP-1 (Table P & 0.05). BA intake lowered APAP-induced hepatic production of these cytokines (P & 0.05).As shown in Table P & 0.05). BA pre-intake at both doses down-regulated the expression of NF-\u03baB p65 and p-JNK (P & 0.05), and only at high dose suppressed hepatic TLR-3, TLR-4 and MyD88 expression (P & 0.05). BA did not alter NF-\u03baB p50 expression (P > 0.05).APAP up-regulated hepatic protein expression of TLR-3, TLR-4, MyD88, NF-\u03baB p50, NF-\u03baB p65, JNK and p-JNK . BA intake at 0.05 and 0.1% decreased hepatic infiltration by inflammatory cells and lowered Ishak inflammation scores (P & 0.05), in which 0.1% BA treatment exhibited greater anti-inflammatory effects than 0.05% BA (P & 0.05).Histological data revealed that BA intake at 0.1% only (without APAP) did not cause inflammatory stress and showed similar Ishak inflammation score as normal groups Figure . APAP trBoswellia species, increased its deposit in liver and protected liver against subsequent APAP induced oxidative and inflammatory injury. Besides increasing GSH retention, we found that BA effectively decreased CYP2E1 activity and expression, lowered ROS, TNF-alpha and MCP-1 production, as well as suppressed HO-1, TLR-3, TLR-4, NF-\u03baB p65 and p-JNK expression. In addition, our histological results indicated that BA improved diffuse ballooning degeneration and lobular inflammation caused by APAP. These findings support that BA is a potent hepatic protective agent.Our present study revealed that the pre-intake of BA, an active compound of APAP at high dose led to GSH depletion and GSSG accumulation . Our datvia its anti-oxidative activities, which definitely led to less stimulation for hepatic NF-kB activation and JNK phosphorylation. Our western blot data regarding hepatic protein expression of NFkB p65 and p-JNK agreed that BA intake at both doses limited the activation of these two signal pathways. Consequently, it is reasonable to observe lower hepatic levels of IL-6, TNF-alpha and MCP-1 in BA treated mice. The decrease in serum levels of ALT, AST and CRP in BA treated mice also agreed that BA intake declined APAP caused liver inflammatory stress. In addition, our histological data further supported that BA pre-intake effectively improved hepatic inflammatory injury.APAP overdose activated NF-kB and JNK pathways through stimulating ROS generation , 23. Theet al. [et al. [in vivo anti-inflammatory data of BA, and extended BA\u2019s mediating activity to TLRs and MyD88.TLRs play crucial roles in the pathological progression of inflammatory liver diseases because TLRs recognize endogenous damage-associated molecular patterns during inflammatory reactions . TLR-3 iet al.  reported [et al.  reportedBA is naturally present in many edible plants including vegetables and herbs , 30. Thevia maintaining hepatic GSH content, retaining activity and expression of GR and HO-1, decreasing inflammatory cytokines and suppressing protein expression of CYP2E1, NF-\u03baB p65, JNK, TLR-3 and TLR-4. These findings support that boswellic acid was a potent hepatic protective agent.Boswellic acid pre-intake protected mice liver against subsequent acetaminophen-induced oxidative and inflammatory injury"}
+{"text": "To control the double burden of communicable and non-communicable diseases (NCDs), in the developing world, understanding the patterns of morbidity and healthcare-seeking is critical. The objective of this cross-sectional study was to determine the distribution, predictors and inter-relationship of perceived morbidity and related healthcare-seeking behavior in a poor-resource setting.Between October 2013 and July 2014, 43999 consenting subjects were recruited from 10107 households in Malda district of West Bengal state in India, through multistage random sampling, using probability proportional-to-size. Information on socio-demographics, behaviors, recent ailments, perceived severity and healthcare-seeking were analyzed in SAS-9.3.2.Pri)=0.76, 95% confidence interval=0.71-0.83) and for Govt. healthcare provider(AORGovt)=0.80(0.68-0.95)], females [AORGovt=0.80(0.73-0.88)], Muslims [AORPri=0.85(0.69-0.76) and AORGovt=0.92(0.87-0.96)], backward castes [AORGovt=0.93(0.91-0.96)] and rural residents [AORPri=0.82(0.75-0.89) and AORGovt=0.72(0.64-0.81)] had lower odds of visiting qualified practitioners. Apparently less severe NCDs , gastrointestinal [AORPri=0.28(0.24-0.33) & AORGovt=0.69(0.58-0.81)], respiratory [AORPri=0.35(0.32-0.39) & AORGovt=0.46(0.41-0.52)] and skin infections [AORPri=0.65(0.55-0.77)] were also less often treated by qualified practitioners. Better education [AORPri=1.91(1.65-2.22) for \u2265graduation], sanitation [AORPri=1.58(1.42-1.75)] and access to safe water [AORPri=1.33(1.05-1.67)] were associated with healthcare-seeking from qualified private practitioners. Longstanding NCDs  and serious infections [typhoid: AORPri=2.86(2.04-4.03)] were also more commonly treated by qualified private practitioners. Potential limitations included temporal ambiguity, reverse causation, generalizability issues and misclassification.Recent illnesses were reported by 55.91% (n=24600) participants. Among diagnosed ailments (n=23626), 50.92% (n=12031) were NCDs. Respiratory ), gastrointestinal  and musculoskeletal  problems were predominant. Non-qualified practitioners treated 53.16% (n=13074) episodes. Older children/adolescents [adjusted odds ratio for private healthcare providers(AORIn this poor-resource setting with high morbidity, ailments and their perceived severity were important predictors for healthcare-seeking. Interventions to improve awareness and healthcare-seeking among under-privileged and vulnerable population with efforts to improve the knowledge and practice of non-qualified practitioners probably required urgently. Demographic ageing, unplanned urbanization and unhealthy lifestyles are the major contributors for the changing pattern of disease in recent years, from communicable to non-communicable diseases (NCDs), globally.\u20133 This eDespite remarkable progress in socio-economic development and having an overarching aim of addressing the health needs through several comprehensive programs, health outcomes in India remained poor. During 2012, approximately 60% deaths were attributed to NCDs  and 28% to communicable, maternal, perinatal and nutritional conditions in this country.,11 EvideIndividual healthcare-seeking pattern in a community is determined by complex interrelationships between socio-economic and physical environment along with individual characteristics and behaviors. Thus heaRelevant researches on morbidity and healthcare-seeking ever conducted in India were mostly limited to urban areas of southern and western part while eastern region remained largely understudied. Malda isHence, a community-based cross-sectional study was designed involving a representative population of Malda to understand the distribution of the perceived morbidity and healthcare-seeking behavior, their predictors and inter-relationship.The study protocol was reviewed and approved by the Ethics Committee of the National Institute of Cholera and Enteric Diseases, Kolkata. Written informed consent left thumb impression  was obtained from residents older than 18 years and from the guardians of residents aged 1 to 17 years. Written assent was additionally obtained from residents aged 12 to 17 years.Based on the 2011 census data, the urban area of the Malda district was divided into two broad urban administrative divisions termed as Municipalities . Each Municipality was further subdivided into smaller administrative units called Wards . Using probability proportional to size (PPS) determined by the total number of households in the Wards, 4 Wards in Old Malda and 12 Wards in English Bazar were selected randomly. The rural area of the district consisted of 3701 villages and 27 rural towns from which similarly using PPS, 25 villages/census towns were selected randomly. Using an exhaustive house-list of the urban and rural areas, each selected municipal ward and village/rural town was categorized into several segments (considered as Primary Sampling Unit: PSU), each having 125 households (defined as those who shared the cooking-pot in each dwelling). Next, 4012 urban and 6095 rural households (maintaining the population ratio) were selected from the whole district, through multistage random sampling, using PPS. Thus, 16 municipal wards in urban and 24 villages/towns in rural area were selected. In each selected ward/village from the list of segments two were selected randomly and all households were surveyed there after collecting written informed consent from the residents.All the individuals residing in the selected households were interviewed at home by trained interviewers, using a structured, pre-tested, bi-lingual  questionnaire. Information was collected on socio-demographic and related variables such as age, gender, religion, caste, education level and occupation of the household members, maximum education level among adults in the house, house ownership, residential area, type and location of water source, water treatment at home, material used for cooking and domestic light source. Housing type was classified as Kachha , Pacca  and Semi-pacca . Sanitation level of toilet use practices were categorized as poor (if the household had no toilet and the members used open space/field/jungle for defecation), good (for households having toilets with flush to piped sewer system/flush to septic tank) and all others  as average.Based on the information regarding household assets (enquired using an appropriate list of assets), number of cattle, goats/sheep, poultry, place for keeping them and the aforementioned household information, wealth index was calculated by using relative weights for each and then the cumulative wealth index scores were log-transformed and divided into quintiles of socio-economic status: SES  based on the percentile distribution.For all the members of the selected households, information regarding last three episodes of ailments that forced them to seek some healthcare services within last two months was collected. Occurrence, perceived severity and healthcare-seeking behavior  regarding specific NCDs like: acid peptic disorder (APD) or peptic ulcer disorder (PUD), chronic obstructive pulmonary disease (COPD), hypertension (HTN), diabetes mellitus (DM), anemia and osteoarthritis (OA) as well as communicable diseases like: gastroenteritis, respiratory tract infection (RTI), typhoid and skin infections were also collected.Thus between October 2013 and July 2014, 43999 individuals (with approximately 8% non-response) were recruited from 10107 households  and collected data were analyzed using Statistical Analysis System (SAS) version 9.3.2. Distribution of the socio-demographic characteristics, morbidity pattern and healthcare-seeking were determined by conducting descriptive analyses using survey frequency procedure to determine overall and stratified frequencies, proportions and corresponding 95% confidence intervals (95%CI). Bivariate and multivariate logistic regression analyses were next conducted to determine unadjusted (OR) and adjusted  odds ratios (AOR) as the measures of association (with corresponding 95%CIs) between study variables. Multinomial logistic regressions  were useAmong 43999 subjects, majorities were aged 18\u201340 yrs , male , Hindu , general caste  and educated up to secondary level . For 38.82% (n = 17080). Maximum adult education in the household was also up to secondary level, 95.73% (n = 42122) stayed in own house, 39.60% (n = 15888) were in sedentary work and 62.60% (n = 27543) lived in rural areas. Only 5.31% (n = 2336) were drinking safe water, 50.32% (n = 22140) had to bring drinking water from outside, 95.06% (n = 41825) were not doing any water treatment at home, 29.08% (n = 12794) were using gas/electricity for cooking, 27.20% (n = 11961) were living in pacca houses. Electricity was the source of lighting at home for 88.87% (n = 39098), regarding toilet use 30.63% (n = 13475) had good sanitary practices and overall 19.44% (n = 8553) belonged to upper SES. Overall and stratified  socio-demographic distribution are presented in Regarding the distribution of self-perceived most recent (within past 2 month) morbidity, 44.09% (n = 19399) did not suffer from any such recently while for 17.28% (n = 7605), 13.48% (n = 5929) and 6.25% (n = 2749) residents the most recent morbidity was related to respiratory, gastrointestinal and musculoskeletal system respectively. Among the most recent ailments, NCDs were 50.92% (n = 12031), 53.16% (n = 13074) episodes were treated by non-qualified practitioners, 34.02% (n = 8368) by qualified practitioner from private sector and only 12.82% (n = 3153) by qualified practitioner from Govt. sector. Non-qualified practitioners were treating more communicable diseases compared to NCDs [57.52% (n = 7194) vs. 42.48% (n = 5313)]. Based on last three healthcare-seeking episodes, among specific ailments (suffered or not), 19.01% (n = 6734) suffered from RTI, 8.18% (n = 2554) had PUD/APD, 6.45% (n = 1977) experienced gastroenteritis while 3.60% (n = 1070) had some skin problems. Among subjects visiting nonqualified practitioners, only 16.85% (n = 1551) perceived their ailments as severe while this fraction for private sector qualified practitioners, was 40.85% (n = 1829). 41\u201360 = 2.01(1.82\u20132.23), AOR>60 = 2.86(2.41\u20133.39)], COPD , HTN , DM , OA , gastroenteritis  and RTI .Association of socio-demographics with morbidity and healthcare-seeking are presented in Tables Compared to males, females had higher odds of suffering from APD [AOR = 1.60(1.45\u20131.77)], HTN [AOR = 1.53(1.28\u20131.83)], anemia [AOR = 16.26(10.75\u201324.59)] and OA [AOR = 2.58(2.07\u20133.22)] and lower odds for COPD [AOR = 0.59(0.48\u20130.73)] and DM [AOR = 0.73(0.57\u20130.92)]. Muslims suffered less from APD [AOR = 0.77(0.69\u20130.87)] and gastroenteritis [AOR = 0.86(0.74\u20130.99)] but more from DM [AOR = 1.40(1.06\u20131.85)], typhoid [AOR = 1.80(1.31\u20132.46)] and skin infections [AOR = 1.25(1.06\u20131.49)] than Hindus. With reference to general, backward castes suffered less from APD [AOR = 0.74(0.67\u20130.81)], HTN [AOR = 0.82(0.69\u20130.97)] and anemia [AOR = 0.77(0.60\u20130.98)] but more from typhoid [AOR = 1.93(1.40\u20132.67)].Higher Secondary = 0.57(0.47\u20130.70), AOR\u2265Graduation = 0.57(0.46\u20130.70)], COPD , anemia [AOR\u2265Graduation = 0.48(0.26\u20130.87)], OA , gastroenteritis  and RTI .Compared to illiterates, higher familial education was associated with lower likelihood of APD [AORHard workers (reference = Sedentary) were more prone to APD [AOR = 1.45(1.24\u20131.71)] and anemia [AOR = 1.89(1.17\u20133.04)] but less vulnerable to COPD [AOR = 0.53(0.40\u20130.69)] and HTN [AOR = 0.60(0.46\u20130.77)]. Rural residents, compared to urban, were less likely to have HTN [AOR = 0.54(0.43\u20130.67)] but more prone to OA [AOR = 1.47(1.15\u20131.87)], gastroenteritis [AOR = 1.76(1.50\u20132.07)], typhoid [AOR = 2.85(1.86\u20134.38)], RTI [AOR = 1.27(1.16\u20131.38)] and skin infection [AOR = 1.45(1.19\u20131.77)].Upper middle = 0.64(0.44\u20130.92), AORUpper = 0.59(0.40\u20130.88)], gastroenteritis [AORUpper = 0.72(0.60\u20130.86)], typhoid [AORUpper = 0.63(0.41\u20130.99)], RTI  and skin infections . Higher SES also seemed to be associated with higher odds of having HTN  and DM . , AOR>60 years = 4.25(3.61\u20135.00)], familial education , sanitation level regarding toilet use practices  and SES . Perception of severity was lower among hard-workers  and rural residents . ]. 5\u201318 = 2.51(2.22\u20132.83)], and older residents were less likely  to suffer from communicable diseases (reference = NCD). Compared to respective reference groups, females [AOR = 0.72(0.67\u20130.77)], residents having higher familial education [AOR = 0.71(0.62\u20130.83)] and higher SES [AOR = 0.84(0.75\u20130.92)] had lower likelihood of communicable diseases. Muslims [AOR = 1.18(1.09\u20131.28)], persons belonging to backward [AOR = 1.15(1.08\u20131.24)] caste, those who had higher individual education [AOR\u2265Graduation = 1.38(1.13\u20131.69)] and rural [AOR = 1.47(1.36\u20131.60)] residents suffered more from communicable diseases. , AORGovt = 0.80(0.68\u20130.95)], females [AORGovt = 0.80(0.73\u20130.88)], Muslim religion , backward caste [AORGovt = 0.93(0.91\u20130.96)], physically demanding occupation  and rural residence  were associated with lower likelihood of visiting qualified practitioners . Age > 40 years , higher individual [for higher secondary: AORPrivate = 1.42(1.19\u20131.69) and for \u2265 Graduation: AORPrivate = 1.30(1.06\u20131.59)] and familial education [for higher secondary: AORPrivate = 1.26(1.13\u20131.41) and for \u2265 Graduation: AORPrivate = 1.40(1.22\u20131.62)], better sanitary practices [for average practice: AORPrivate = 1.17(1.07\u20131.28) and for good practice: AORPrivate = 1.58(1.42\u20131.75)] and higher SES [for Upper middle: AORPrivate = 1.59(1.43\u20131.77) and for Upper: AORPrivate = 1.51(1.35\u20131.69)] were associated with higher odds of seeking care from qualified  practitioners. , AORGovt = 0.36(0.31\u20130.43)], OA , gastroenteritis , RTI , skin infections [AORPrivate = 0.65(0.55\u20130.77)]. Those who had COPD , HTN , DM , typhoid and NCDs  were more likely to visit qualified practitioners. Higher self-perceived disease severity  was also positively associated with visiting qualified practitioners.  of the participants suffered from some recent morbidity while respiratory, gastrointestinal and musculoskeletal diseases were most common. This observed burden of self-perceived morbidity was considerably higher than previously reported values (ranged between 27% and 48%) in similar settings.\u201329 StudiMore than half of the ailments were treated by non-qualified practitioners, which raised a few concerns. Only about 13% visited qualified physicians from Govt. sector. The scenario seemed similar to that of other parts of India, Vietnam and Bangladesh ,28,32 buAmong specific ailments, RTI was perceived to be the commonest, followed by APD, gastroenteritis and skin problem. Contrary to some other study, perceived burden of HTN and DM were found to be relatively lower. May be sWhile more than two third subjects considered their ailments as less severe, those who perceived the severity, visited qualified doctors especially in private sector. The perceived severity probably helped them to overcome the potential barriers  in better healthcare-seeking.,35,38,39Corroborating with prior observation in similar settings elsewhere, children and adolescents were less likely to suffer from NCDs like APD, COPD, HTN, DM, anemia and OA but more from RTI, gastroenteritis and skin infection.,35,36,40Similar to some previous observation, females had higher likelihood of having APD, anemia and OA but less likely to suffer from COPD and DM \u201328 but gSupporting some prior evidences  and contOccupation with hard work was associated with higher odds of APD and anemia but lower odds of COPD and HTN. Physical exertion, work environment and appropriate nutrition probably were the key factors. Negative association between physical activity and HTN was well-established in prior studies.Rural residents compared to urban were less prone to HTN  but they had higher likelihood of having OA, gastroenteritis, typhoid, RTI and skin infection most likely due to lifestyle related factors, less awareness, poor hygiene and inappropriate sanitation. Urban preponderance of HTN was also reported previously  althoughDrinking safer water was associated with higher perceived burden of HTN and DM. Subjects having better sanitary practices regarding toilet use were also suffering more from APD, HTN, DM and OA. Health awareness and knowledge as probably a confounder here that positively influenced both better practices  and improved perception. Reverse causation might also be a possibility (being diagnosed with the disease resulted in better sanitation and hygiene). Drinking safer water and practicing better sanitation regarding toilet use seemed to be also associated with lower likelihood of suffering from gastroenteritis, typhoid, RTI and skin infections.Alike prior studies, we also found that, residents having comparatively higher SES were less likely to suffer from anemia, gastroenteritis, typhoid, RTI and skin infections ,29 but sPerceived severity of ailments was higher among those with higher age, better familial education, improved sanitation and upper SES and lower among hard-workers and rural residents had. Higher severity of self-perceived morbidity among elderly was also reported previously. Thus perCompared to those aged between 18\u201340 years, 5\u201318 years age group were more likely, and older residents were less likely to suffer from communicable diseases than NCDs. Female gender, better familial education and higher SES were negatively associated with risk of communicable diseases. Muslim religion, backward caste, higher individual education and rural residents had higher odds of suffering from communicable diseases.Socio-demographic predictors of Healthcare-seeking behavior in our study were quite similar to those reported from other parts of the world as well as India with some variations. While elderly subjects commonly visited qualified private and govt. sector physicians, older chSubjects suffering from NCDs were more likely to visit qualified practitioners especially the private sector. Alike soSelf-perceived severity of ailments were positively associated with odds of visiting qualified practitioners more so in private sector and this finding also supported prior evidences.,36,40 ThDespite efficient sampling design, use of detailed questionnaire and robust analyses, our study had certain limitations. Like any other cross-sectional study, causal interpretation of the observed associations is not recommended. Due to the potential vulnerability to temporal ambiguity by design, some of our observations might have suffered from reverse causation. Although self-perceived morbidity and severity are currently being considered an efficient parameter for the estimation of health needs in communities worldwide, keeping the lower literacy and potential lack of awareness in mind, the reported self-perceived morbidity pattern should only be interpreted as perceived health need of the community, not the prevalence. Residual confounding due to variables not included in our analyses could also be an issue. Information bias due to misclassification of self-reported information should always be kept in mind, especially due to the potential for differential recall. But we do not consider those to be serious issues here because we only dealt with the recent ailments, hence recall period was short and in majority of cases, medical records were consulted. Although results of our study should be extrapolated beyond the study sample with caution, still we are not worried about the generalizability of our results due to the representative nature of our study sample and very low (<8%) non-response.In this poor-resource setting, most important predictor for healthcare-seeking was the perception regarding severity and nature of ailments, while age, gender, caste, religion, familial education, SES, residential area, sanitation and hygiene influenced the morbidity pattern and relevant healthcare-seeking. Keeping the high burden of self-perceived morbidity in mind, interventions to improve physical health, awareness and care-seeking practices targeting children, elderly, females, backward castes, minority groups, illiterates, rural residents and those having lower SES, poor sanitary practices and inadequate access to safe drinking water were required urgently. Simultaneously, efforts to improve the healthcare service delivery might consider implementation of intervention targeting improvement of knowledge and practice among non-qualified practitioners in poor-resource settings where seeking healthcare services from these practitioners seemed to be a common occurrence."}
+{"text": "Consequently, several molecular targets based on the sequences of housekeeping genes, non-functional RNA and structural ribosomal RNAs have been proposed for the detection and identification of mycobacteria in clinical or environmental samples. While certain of these targets were proposed as specific for this genus, most are prone to false positive results in complex environmental samples that include related, but distinct, bacterial genera. Nowadays the increased number of sequenced genomes and the availability of software for genomic comparison provide tools to develop novel, mycobacteria-specific targets, and the associated molecular probes and primers. Consequently, we conducted an M. tuberculosis H37Rv, only 11 proteins showed 80% to 100% of similarity with Mycobacterium spp. genomes, and less than 50% of similarity with genomes of closely related Corynebacterium, Nocardia and Rhodococcus genera. Based on DNA sequence alignments, we designed primer pairs and a probe that specifically detect the atpE gene of mycobacteria, as verified by quantitative real-time PCR on a collection of mycobacteria and non-mycobacterial species. The real-time PCR method we developed was successfully used to detect mycobacteria in tap water and lake samples.Among the 3989 predicted proteins from atpE gene can serve for highly specific detection and precise quantification of Mycobacterium spp. in environmental samples.The results indicate that this real-time PCR method targeting the  Mycobacterium genus is constituted of several pathogenic species, including the M. tuberculosis complex (MTC) responsible for tuberculosis , M. leprae responsible for leprosy, and non-tuberculous mycobacteria (NTM), which are environmental potentially pathogenic species causing mycobacteriosis [rrs, gyrA, gyrB, hsp65, recA, rpoB, sodA genes and 16S-23S internal transcribed spacer (ITS) genes, to detect and/or identify mycobacteria species by sequence analysis [Mycobacterium genus in clinical and environmental samples, several studies have proposed targeting different loci of the 16S rRNA gene [gyrB [rpoB[hsp65[Corynebacterium, Nocardia, Rhodococcus, Mycobacterium (CNM) group [Mycobacterium genus targets with high levels of specificity and sensitivity that will be useful for studying mycobacteria in their habitat.teriosis . Detectiteriosis . Identifanalysis . In ordeRNA gene , or othene [gyrB , rpoB[19poB[hsp65. NeverthpoB[hsp65. Indeed,poB[hsp65 which alM) group  and whicM. tuberculosis H37Rv), the number of mycobacterial sequences has considerably increased due to advances in sequencing capacity and the appearance of high throughput sequencing techniques [M. tuberculosis and M. bovis species), two strains of M. leprae, and eleven species and subspecies of pathogenic (P) and non-pathogenic (NP) NTM: M. abscessus (P), M. avium (P), M. avium subsp. paratuberculosis (P), M. gilvum (NP), M. marinum (P), M. smegmatis (NP), Mycobacterium sp. JLS (NP), Mycobacterium sp. KMS (NP), Mycobacterium sp. MCS (NP), M. ulcerans (P), M. vanbaalenii (NP), [M. intracellulare, M. kansasii, M. parascrofulaceum). This increasing number of completely sequenced mycobacterial genomes led to the development of the MycoHit software, which permits gene- and protein-level comparisons across mycobacteria species, [As new mycobacterial sequences are added into genetic databases, our knowledge of mycobacterial genomes is increasing and this may help to design new primers and probes that will be both specific and sensitive. Since the whole sequencing of the first mycobacterial genome in 1998  by Sangechniques . Today, ii (NP), -26. Morespecies, . This sospecies, . HoweverMycobacterium spp.. We compared in silico proteins of whole mycobacterial genomes with those of non-mycobacterial genomes using the MycoHit software, in order to find conserved sequences among mycobacteria that will not be shared with non-mycobacterial species. Based on the screening results a primer pair and a probe targeting the atpE gene were designed and tested by real-time PCR. This novel target proved to be totally specific and sensitive. It also offers the advantage of targeting a gene present as a single copy in the genome. Thus this new real-time PCR method appears promising for water quality survey, and should be useful for studying the ecology of mycobacteria in aquatic, terrestrial and urban environments.In this paper, we used this tool for screening sensitive and specific targets of rrs gene and ITS , and according to our strategy of genome comparison  code for proteins which present similar conformations in non-mycobacterial studied genomes , and also for the 12 non-mycobacterial genomes studied .Excluding n Figure\u00a0 most of M. tuberculosis H37Rv genome , and less than 50% similarity levels in comparison with genomes (n\u2009=\u200912) of the other CNM group genera. As a result, among the 3989 predicted proteins of M. tuberculosis H37Rv genome , hypothetical PE or PPE family proteins (loci Rv0285 and Rv3022c), proteins coded by esxG, esxH and esxR genes in M. tuberculosis H37Rv , and proteins such as a lipoprotein coding by lppM gene (locus Rv2172c), an oxidoreductase (locus Rv0197), and a small secreted protein (locus Rv0236A).Among the 3989 predicted proteins of e Figure\u00a0, about 5e Figure\u00a0B displaycmaA1 gene, lipoprotein coding by lppM gene, as well as PE, PPE and proteins coded by esx genes esxG, esxH and esxR, were highly conserved in studies MTC species (tuberculosis and bovis) but very polymorphous in the 14 NTM species studied , the oxidoreductase (locus Rv0197), and the small secreted protein locus Rv0236A), are the less polymorphous among the 14 NTM species studied  whereas quantification limits were estimated at about 100 genome equivalents. In the positive collection all 31 mycobacteria species were positively detected by the real-time PCR method. This collection includes NTM species, leprae species and MTC species as tuberculosis and bovis  , and evaR Figure\u00a0A. DetectatpE gene, shows a vast diversity of mycobacteria concentration, ranging from 104 to 106 ge/L in water column and neuston samples, and 105 to 106 ge/g DW (dry weight) in sediment samples. Comparison with the previously published methods targeting 16S rRNA [Mycobacteria quantification in lake samples by real-time PCR targeting 16S rRNA  shows a gyrA, gyrB, hsp65, recA, rpoB, and sodA genes are appropriate for identification purposes [gyrB [rpoB[hsp65[Helicobacter pylori show positive amplification with several Mycobacterium specific primer pairs [atpE real-time PCR method that we propose is just as specific, but more sensitive than the previously proposed rrs real-time PCR method which cannot detect some mycobacterial species [Although purposes ,4, our res [gyrB , rpoB[19poB[hsp65 genes, hpoB[hsp65. For exaer pairs . Prospec species .M. tuberculosis H37Rv, sorting proteins according to similarity requests and listing candidate proteins . Only the aptE gene could be used to design primers and a probe for mycobacteria detection. Concerning the other genes, the sequence polymorphism among NTM species did not allow designing molecular targets for Mycobacterium spp. detection. However, these genes could be of immunological or pathogenic importance. Indeed, PE and PPE family proteins represent 0.9 to 4.2% of the genome coding capacity of several mycobacteria [esx gene clusters, which encode ATP dependent specific secretion system [esx gene clusters are very small and polymorphous among genomes of the 11 NTM species compared  is exclusively conserved in the genomes of the 17 mycobacterial species studied , which codes for the ATP synthase protein subunit C. These results also showed that our strategy of target design based on MycoHit software , slow mycobacteria growth, clumping of mycobacterial cells, high hydrophobicity of mycobacteria and contamination of culture media by other fast growing environmental microorganisms [rganisms .atpE with previously described method targeting 16S rRNA, [atpE gene presents two major advantages over the method targeting rrs gene. First, the new method detects all the tested mycobacterial strains, while the method targeting rrs gene cannot detect isolates of M. celatum, M. heckeshornense, and M. leprae[atpE gene is present in a single copy in the Mycobacterium genomes, while the 16S rRNA gene is present either in 1 or 2 copies in the genome [Comparison of the method targeting 6S rRNA, , showed M. leprae. Second,e genome . When coin vitro as positive controls whereas more than 150 mycobacterial species have been described so far [atpE real-time PCR method using a large representative collection of mycobacterial species , including members of MTC (n\u2009=\u20092), M. leprae species (n\u2009=\u20091), slow growing NTM (n\u2009=\u200913), and rapid growing NTM (n\u2009=\u200915). Given the broad diversity of mycobacterial species we have tested in this study, we expect the method to be applicable to all species within the Mycobacterium genus. In addition, it is the first time that a sensitive and specific molecular target has been identified based on an in silico comparison of 16 mycobacterial (13 species) and 12 non-mycobacterial genomes (4 closely related species).One of the limitations of this study is that only 31 mycobacterial species were tested Archaea. Using these databases, MycoHit, or other new software, may then be used to design new primers for real-time PCR detection or quantification, for in situ hybridization and other molecular tools. With this approach we were able to design primer pairs and a probe that target specific mycobacterial atpE gene, and could be used to detect and quantify very specifically mycobacteria in environmental samples. Although the atpE gene may not be appropriate for microdiversity studies, it appeared to be very useful for specific detection of the genus Mycobacterium in environmental samples. More generally, genome comparison used here showed its utility to identify specific genera\u2019s targets, and could be used to identify specific proteins for antimicrobial design as previously emphasized [In conclusion, although our strategy did not take into account non-coding regions, such as insertion sequences, repetitive units, non-functional RNA, and structural ribosomal RNAs, the comparison of whole bacterial genomes for design of specific primers is a promising approach not only for mycobacteria but also for other cultured bacterial or archaeal groups for which whole sequenced genomes are accumulating in databases. Metagenomic libraries from environmental samples which are increasingly performed in microbial ecology studies  could alphasized .M. tuberculosis genes, presenting homologue genes in other mycobacterial genomes, and not presenting homologue genes in non-mycobacteria genomes, we used the MycoHit software version 14.17  and performed an alignment search with Stand Alone tblastn algorithm as previously described [M. tuberculosis H37Rv has been used as a reference of the Mycobacterium genus, because it is the most historically described mycobacterial genome [M. tuberculosis H37Rv, corresponding to the query sequences used in order to search for matches in the genomic DNA of other organisms , allowed identification of the conserved mycobacterial proteins presenting no homology in non-mycobacterial genomes  using the accession numbers: M. abscessus ATCC 19977 (CU458896.1) (P), M. avium 104 (CP000479.1) (P), M. avium subsp. paratuberculosis K10 (AE016958.1) (P), M. bovis subsp. bovis AF2122/97 (BX248333.1) (P), M. gilvum PYR-GCK (CP000656.1) (NP), M. marinum M (CP000854.1) (P), M. smegmatis MC2 155 (CP000480.1) (NP), Mycobacterium sp. JLS (CP000580.1) (NP), Mycobacterium sp. KMS (CP000518.1) (NP), Mycobacterium sp. MCS (CP000384.1) (NP), M. tuberculosis CDC1551 (AE000516.2) (P), M. tuberculosis H37Ra (CP000611.1) (NP), M. tuberculosis H37Rv (AL123456.2) (P), M. tuberculosis KZN 1435 (CP001658.1) (P), M. ulcerans Agy99 (CP000325.1) (P), and M. vanbaalenii PYR-1 (CP000511.1) (P). In order to avoid data lost during genome comparisons performed by MycoHit software, we have chosen to ignore some mycobacterial genomes. Since the number of coding proteins is much lower compared to other mycobacterial species, M. leprae Br4923 (FM211192.1) (P), and M. leprae TN (AL450380.1) (P) were ignored in the analysis  [M. bovis BCG Pasteur 1173P2 (AM408590.1) (NP) and M. bovis BCG Tokyo 172 (AP010918.1) (NP) were also not taken into account, because these vicinal genomes present mutations [M. intracellulare ATCC 13950 (ABIN00000000) (P), M. kansasii ATCC 12478 (ACBV00000000) (P) and M. parascrofulaceum BAA-614 (ADNV00000000) (P) were also not used during MycoHit proceedings, because their genomes were still not assembled at the moment we performed the first screening step of our analysis. Nevertheless, the genomes of M. leprae, M. bovis BCG, M. intracellulare, M. kansasii and M. parascrofulaceum were used during alignment of nucleic sequences of the most conserved proteins in mycobacterial genomes.In order to perform comparisons of pathogenic (P) and non-pathogenic (NP) mycobacterial genomes with 122/97 BX8333.1 (PMC2 155) ,35. GenoCorynebacterium aurimucosum ATCC 700975 (CP001601.1), C. diphtheriae NCTC 13129 (BX248353.1), C. efficiens YS-314 (BA000035.2), C. glutamicum ATCC 13032 (BX927147.1), C. jeikeium K411 (NC_007164), C. kroppenstedtii DSM 44385 (CP001620.1), C. urealyticum DSM 7109 (AM942444.1), Nocardia farcinica IFM 10152 (AP006618.1), Nocardioides sp. JS614 (CP000509.1), Rhodococcus erythropolis PR4 (AP008957.1), R. jostii RHA1 (CP000431.1), and R. opacus B4 (AP011115.1).We selected non-mycobacterial genomes of species from the CNM group using the following accession numbers: In order to check the homology of the selected mycobacterial sequences, the protein and DNA sequences of these selected proteins were aligned using the ClustalW multiple alignment of the BioEdit software 7.0.9.0 with 1000 bootstraps . Primer et al. protocol [M. avium, M. fortuitum, M. intracellulare and M. gordonae . Specificity and sensitivity were estimated against 30 non-mycobacteria (negative) strains and 31 mycobacteria (positive), respectively. The collection contained reference and environmental strains of mycobacteria, as well as, strains of the closely related CNM group, and other non-actinobacteria strains isolated from the environment [leprae species (n\u2009=\u20091), as well as species of slow growing NTM (n\u2009=\u200913), and rapid growing NTM (n\u2009=\u200915). TaqMan\u00ae real-time PCR were performed in duplicate using an ABI7500 real-time PCR system (Applied Biosystems), a Lifetech 7500 software version 2.0.6 (Applied Biosystems) and TaqMan fast virus 1-STEP Master Mix with 6-carboxy-X-rhodamine (ROX) (Applied Biosystems). The TaqMan\u00ae probes were labeled (Eurogentec) with the fluorescent dyes 6-carboxyfluorescein (5\u2032 end) and Black Hole Quencher (3\u2032 end). All reactions were performed in a 25 \u03bcl reaction mixture volume (2.5 \u03bcl of DNA) with 500 nM of forward primer, 500 nM of reverse primer, 50 nM of probe and 5 mM of MgCl2. Reverse transcriptase was inactivated immediately  according to the manufacturer instruction, and real-time PCR consisted in 40 cycles of denaturation (95\u00b0C for 3 s), annealing and extension (both steps at 60\u00b0C for 30 s). Determinations of cycle threshold were performed by setting the instrument\u2019s threshold line at 0.02 \u2206Rn units .Reproducibility, sensitivity and specificity of the new real-time PCR method were estimated using DNA from a previously described microorganism collection, and according to Radomski protocol . Reproduprotocol  were estironment . Mycobacet al. [et al. [In order to compare the new real-time PCR method to the culture method, 26 tap water distribution points in Paris (France) were sampled between April 2011 and July 2011, corresponding to 90 samples. Briefly, one liter of tap water was sampled in sterile plastic bottle, then centrifuged at 5000\u2009\u00d7\u2009g for 2h and finally re-suspended in 1 ml of water. Mycobacteria density was estimated by culture (Method A) in all these samples following the procedure previously described by Le Dantec et al. . In para [et al.   between Mycobacterium tuberculosis H37Rv (AL123456.2) proteins and proteins of targeted mycobacterial genomes and proteins of non-targeted genomes. Targeted mycobacterial genomes include M. tuberculosis H37Ra (CP000611.1), M. tuberculosis CDC 1551 (AE000516.2), M. tuberculosis KZN 1435 (CP001658.1), M. bovis AF2122/97 (BX248333.1), M. ulcerans Agy99 (CP000325.1), M. marinum M (CP000854.1), M. avium 104 (CP000479.1), M. paratuberculosis K10 (AE016958.1), M. smegmatis MC2 155 (CP000480.1), M. abscessus ATCC 19977 (CU458896.1), M. gilvum PYG-GCK (CP000656.1), M. vanbaalenii PYR-1 (CP000511.1), Mycobacterium sp. JLS (CP000580.1), Mycobacterium sp. KMS (CP000518.1), Mycobacterium sp. MCS (CP000384.1), and non-targeted genomes include Corynebacterium aurimucosum ATCC 700975 (CP001601.1), C. diphteriae NCTC 13129 (BX248353.1), C. efficiens YS-314 (BA000035.2), C. glutamicum ATCC 13032 (BX927147.1), C. jeikeium K411 (NC_007164), C. kroppenstedtii DSM 44385 (CP001620.1), C. urealyticum DSM 7109 (AM942444.1), Nocardia farcinica IFM 10152 (AP006618.1), Nocardioides sp. JS614 (CP000509.1), Rhodococcus erythropolis PR4 (AP008957.1), R. jostii RHA1 (CP000431.1) and R. opacus B4 (AP011115.1).Click here for fileProtein sequence alignment of conserved proteins in mycobacterial genomes. Sequences are from genomes of M. abscessus ATCC 19977 (CU458896.1), M. avium 104 (CP000479.1), M. avium subsp. paratuberculosis K10 (AE016958.1), M. bovis subsp. bovis AF2122/97 (BX248333.1), M. bovis BCG Pasteur 1173P2 (AM408590.1), M. bovis BCG Tokyo 172 (AP010918.1), M. gilvum PYR-GCK (CP000656.1), M. intracellulare ATCC 13950 (ABIN00000000), M. kansasii ATCC 12478 (ACBV00000000), M. leprae Br4923 (FM211192.1), M. leprae TN (AL450380.1), M. marinum M (CP000854.1), M. parascrofulaceum BAA-614 (ADNV00000000), M. smegmatis MC2 155 (CP000480.1), Mycobacterium sp. JLS (CP000580.1), Mycobacterium sp. KMS (CP000518.1), Mycobacterium sp. MCS (CP000384.1), M. tuberculosis CDC1551 (AE000516.2), M. tuberculosis H37Ra (CP000611.1), M. tuberculosis H37Rv (AL123456.2), M. tuberculosis KZN 1435 (CP001658.1), M. ulcerans Agy99 (CP000325.1) and M. vanbaalenii PYR-1 (CP000511.1).Click here for fileDNA sequence alignment of conserved proteins in mycobacterial genomes. Sequences are from genomes of M. abscessus ATCC 19977 (CU458896.1), M. avium 104 (CP000479.1), M. avium subsp. paratuberculosis K10 (AE016958.1), M. bovis subsp. bovis AF2122/97 (BX248333.1), M. bovis BCG Pasteur 1173P2 (AM408590.1), M. bovis BCG Tokyo 172 (AP010918.1), M. gilvum PYR-GCK (CP000656.1), M. intracellulare ATCC 13950 (ABIN00000000), M. kansasii ATCC 12478 (ACBV00000000), M. leprae Br4923 (FM211192.1), M. leprae TN (AL450380.1), M. marinum M (CP000854.1), M. parascrofulaceum BAA-614 (ADNV00000000), M. smegmatis MC2 155 (CP000480.1), Mycobacterium sp. JLS (CP000580.1), Mycobacterium sp. KMS (CP000518.1), Mycobacterium sp. MCS (CP000384.1), M. tuberculosis CDC1551 (AE000516.2), M. tuberculosis H37Ra (CP000611.1), M. tuberculosis H37Rv (AL123456.2), M. tuberculosis KZN 1435 (CP001658.1), M. ulcerans Agy99 (CP000325.1) and M. vanbaalenii PYR-1 (CP000511.1).Click here for file"}
+{"text": "T) was obtained in China from crickets  living in cropland deserted for approximately 10 years. The isolated bacteria were Gram-negative, facultatively anaerobic, oxidase-negative rods. A preliminary analysis of the 16S rRNA gene sequence indicated that the strain belongs to either the genus Erwinia or Pantoea. Analysis of multilocus sequence typing based on concatenated partial atpD, gyrB and infB gene sequences and physiological and biochemical characteristics indicated that the strain belonged to the genus Erwinia, as member of a new species as it was distinct from other known Erwinia species. Further analysis of the 16S rRNA gene showed SCU-B244T to have 94.71% identity to the closest species of that genus, Erwinia oleae (DSM 23398T), which is below the threshold of 97% used to discriminate bacterial species. DNA-DNA hybridization results (5.78\u00b12.52%) between SCU-B244T and Erwinia oleae (DSM 23398T) confirmed that SCU-B244T and Erwinia oleae (DSM 23398T) represent different species combined with average nucleotide identity values which range from 72.42% to 74.41. The DNA G+C content of SCU-B244T was 55.32 mol%, which also differs from that of Erwinia oleae (54.7 to 54.9 mol%). The polyphasic taxonomic approach used here confirmed that the strain belongs to the Erwinia group and represents a novel species. The name Erwinia teleogrylli sp. nov. is proposed for this novel taxon, for which the type strain is SCU-B244T (= CGMCC 1.12772T = DSM 28222T = KCTC 42022T).A bacterial isolate (SCU-B244 Our aim was to explore the relationship between pesticide resistance and symbionts. Among 274 isolates cultured from Teleogryllus occipitalis, 27 strains of genera Lysinibacillus, Pseudomonas, Sphingobacterium, Exiguobacterium and Staphylococcus could evidently degrade chlorpyrifos, a common insecticide used in this field for many years. One isolate (SCU-B244T) that could degrade chlorpyrifos was cultured on TSA (tryptone soy agar) medium in August 2012 and could not be identified to the species level. A polyphasic taxonomic approach was used to investigate the strain, with the results suggesting that SCU-B244T represents a novel species of the genus Erwinia.In a recent study, we focused on the culturable strains associated with crickets  was used as reference strain in this study.Three crickets  were added to 100 mL sterile 0.85% (w/v) NaCl solution in a 250 mL flask and shaken at 220 rpm for 30 min. The supernatant which contained bacteria was plated onto TSA plates  and subsequently incubated at 37\u00b0C for 5 days. Glycerol stock at -80\u00b0C was adopted for long term preservation of the isolates. Exponential phase cells cultured in TSB  medium with shaking  at 37\u00b0C were harvested for DNA G+C content and ANI analysis. Phase contrast and transmission electron microscopy were used to examine cellular morphology and motility after growth on TSA medium at 37\u00b0C for 24 h. Gram staining was performed as described by Gerhardt T, PCR amplification, primers used and DNA sequencing conditions of 16S rRNA gene were performed as previously described [www.ezbiocloud.net/eztaxon) by comparison with 16S rRNA gene sequence data. A neighbour-joining phylogenetic tree was constructed using the method of Saitou and Nei [DNA extraction from strain SCU-B244escribed  , with the highest similarity scores being 96.1% to 94.1%. Neighbour-joining and maximum-likelihood phylogenetic trees were constructed using the methods described above.The results of 16S rRNA gene sequence alignment on the EzTaxon server revealed that strain SCU-B244T, other neighbour-joining and maximum-likelihood phylogenetic trees based on 16S rRNA gene sequences were constructed, including the strain SCU-B244T and all culturable type strains of the genera Erwinia (Candidatus Erwinia dacicola) and Pantoea, based on the same methods described above.To refine the taxonomic position of SCU-B244atpD, gyrB, infB and rpoB gene sequences enables the differentiation of the phylogenetically related genera Erwinia, Pantoea and Tatumella. A good congruence has previously been observed between DNA-DNA hybridization and MLSA [atpD, gyrB, and infB, were amplified and sequenced with the primers described by Brady et al. [atpD, gyrB and infB gene sequences were constructed using the same method as described above. Strains from genera Erwinia, Pantoea and Tatumella were used in MLSA analysis, including SCU-B244T. The partial sequence of atpD, gyrB and infB gene of related strains were obtained from GenBank and the accession numbers are indicated on the figures.Multilocus sequence analysis (MLSA) of concatenated partial and MLSA \u201316. In ty et al. . NeighboT and Erwinia oleae DSM 23398T was conducted as described by De Ley et al. [260/280 = 1.8 to 1.9) was extracted using the Omega Bacterial DNA kit (E.Z.N.A.\u00ae)DNA-DNA hybridization between SCU-B244y et al.  with hybT was determined by whole genome sequencing data.The DNA G+C mol% content of strain SCU-B244et al.[API 20E, API 50CHE acid production tests and API ZYM enzymatic characteristics test (BioM\u00e9rieux) were performed according to the manufacturer\u2019s instructions. BIOLOG GN2 carbon-source utilization analysis was conducted according to the instruction manual. Cellular fatty acids analysis was also performed. Biomass for fatty acid analysis was prepared by scraping growth from TSA plates after 24h incubation at 37\u00b0C. Lipids were extracted using the method of Folch et al.. Total let al., The comT, Next Generation Sequencing (Illumina Miseq) was conducted at Majorbio Inc. and the data was used to calculate the Average Nucleotide Identity (ANI). DNA extraction method and purity were described above. ANI is a similarity measure between two genome sequences and it correlates well with DNA-DNA hybridization values. [www.ezbiocloud.net/ezgenome/browse_db). We used Orthologous Average Nucleotide Identity Tool  to calculate orthologous ANI values. This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession LLXO00000000. The version described in this paper is version LLXO01000000.For the genome sequencing of SCU-B244 values.  A value  values.  Genome dTeleogryllus occipitalis and a bacterial isolate (SCU-B244T) is herein described. Initial microbiological characterization of the strain revealed that the cells were Gram-negative, oxidase-negative, rod-shaped, catalase-positive and facultatively anaerobic, suggesting that the strain belongs to the family Enterobacteriaceae [In total 274 isolates belonging to 29 genera were cultured from eriaceae .T and strains of the genera Erwinia and Pantoea cluster together. The diagram shows the phylogenetic relationship between SCU-B244T and genera Erwinia and Pantoea.Neighbour-joining  and maxiErwinia and Pantoea, indicated that strain SCU-B244T belongs to the genus Erwinia and is most closely related to Erwinia oleae (DSM 23398T), which is consistent with the results of the initial phenotypic analysis.Further neighbour-joining  and maxiatpD, gyrB and infB gene sequences, strain SCU-B244T and Erwinia oleae (DSM 23398T) cluster together on a single branch, consistent with the previous results, suggesting strain SCU-B244T is most closely related to Erwinia oleae (DSM 23398T).In neighbour-joining  and maxiT with species of genera Erwinia and Pantoea were 95.43% (Erwinia psidii LMG 7039T) and 95.42%  respectively, phylogenic closest strain was Erwinia oleae (DSM 23398T) and the identity was 94.71%, which is lower than the 97% threshold that has been established to discriminate species. Strains showing less than 97% 16S rRNA gene identity are unlikely to have more than 60 to 70% DNA\u2014DNA relatedness [T is a novel species. The DNA\u2014DNA relatedness between SCU-B244T and Erwinia oleae (DSM 23398T) was 5.79 \u00b1 2.52%. The value is mean of six hybridizations \u00b1 SD, which is significantly lower than the 70% value considered to be the threshold for the delineation of bacterial species [T and related species are listed in The highest 16S rRNA gene identity of SCU-B244leae DSM 398T and T was 55.32 mol% which could be discriminated from Erwinia oleae (DSM 23398T), which has a G+C content of 54.7 to 54.9 mol%.The DNA G+C content of strain SCU-B244T strain can be discriminated from each recognized species of the genus Erwinia by at least three characteristics, and from the phylogenetically most closely related species, Erwinia oleae, by 10 API 50CHE characteristics (API 20E and API 50CHE (BioM\u00e9rieux) tests were also carried out and the results were compared to related strains . The reseristics .API ZYM enzymatic characteristics and BIOLOG GN2 carbon-source utilization results are presented as supplementary data  Tables.T and related species of genera Erwinia and Pantoea is shown in T contained C16:0 (28.2%), Summed feature 4 (12.4%), C11:0 3-OH (11.6%), Summed feature 7 (10.5%), Summed feature 3 (10.0%), C14:0 (6.4%) as the major fatty acids, while strain DSM 23398T contained C16:0 (38.8%), Summed feature 4 (32.7%), Summed feature 7 (9.1%), C12:0 (7.7%), C16:0 \u03949 cyclo (4.6%) as the major fatty acids (14:0 2-OH (3.7%) and C18:0 (3.7%) were detected from strain SCU-B244T while those were not detected from strain DSM 23398T. Fatty acid composition of SCU-B244T and reference strains data from Erwinia and Pantoea showing that most of the compositions were in the range of Erwinia and Pantoea [T belongs to group Erwinia and Pantoea.Fatty acid composition analysis of SCU-B244ty acids . Summed  Pantoea , which iT is a member of genus Erwinia. DNA-DNA hybridization results, ANI values and phenotypic data obtained in this study can separate this isolate from related species. In conclusion the bacterial strain isolated from a cricket (genus Teleogryllus) represents a novel species for which the name Erwinia teleogrylli sp. nov. is proposed. Strain SCU-B244T (= CGMCC 1.12772T = DSM 28222T = KCTC 42022T) is the type strain.Based on the genotypic data , it is clear that SCU-B244Erwinia teleogrylli .The colonies are milky white, circular and convex with entire margins. Cells are short Gram-negative, facultatively anaerobic rods that are oxidase-negative, catalase-positive and able to grow in 6% (w/v) NaCl. Growth pH range is 6 to 9, with optimum growth at 7. Nitrate is reduced to nitrite.2S and indole production, acid production from sorbitol and sucrose. According to API 50CHE tests (BioM\u00e9rieux), acid is formed from glycerol, L-arabinose, D-ribose, D-xylose, D-galactose, D-glucose, D-fructose, D-mannose, L-sorbose, L-rhamnose, D-mannitol, D-sorbitol, methyl-\u03b1D-glucopyranoside, N-acetylglucosamine, arbutin, esculin ferric citrate, salicin, D-cellobiose, D-maltose, D-melibiose, D-trehalose, gentiobiose, potassium gluconate, potassium 2-ketogluconate, potassium 5-ketogluconate. No acid is produced from erythritol, D-arabinose, L-xylose, D-ardonitol, methyl-\u03b2 D-xylopyranoside, dulcitol, inositol, methyl-\u03b1D-mannopyranoside, amygdalin, D-lactose, D-saccharose, inulin, D-melezitose, D-raffinose, amidon, glycogen, xylitol, D-turanose, D-lyxose, D-tagatose, D-fucose, L-fucose, D-arabitol, L-arabitol. Results obtained with API ZYM tests (BioM\u00e9rieux) gave positive enzymatic activity results for alkaline phosphatase, esterase (C4), esterase lipase (C8), acid phosphatase, naphthol-AS-B1-phosphohydrolase, \u03b2-galactosidase, \u03b2- glucosidase and N-acetyl-\u03b2- glucosaminidase, and negative results for lipase (C14), cystine arylamidase, trypsin, \u03b1-chymotrypsin, \u03b2-glucuronidase, \u03b1-glucosidase, \u03b1-mannosidase and \u03b1-fucosidase.Results obtained with API 20E tests (BioM\u00e9rieux) gave positive results for acid production from citrate, glucose, mannitol, rhamnose, melibiose, amygdalin and arabinose, and negative results for \u03b2-galactosidase, gelatinase, urease, arginine digydrolase, lysine decarboxylase and ornithine decarboxylase activity, HThe following substrates were utilized according to BIOLOG GN2 carbon-source utilization analysis: dextrin, N-acetyl-D-glucosamine, L-arabinose, D-fructose, D-galactose, \u03b1-D-glucose, maltose, D-mannitol, D-mannose, D-melibiose, \u03b2-methyl-D-glucoside, D-psicose, L-rhamnose, D-trehalose, turanose, pyruvic acid methyl ester, citric acid, formic acid, D-galactonic acid lactone, D-galacturonic acid, D-gluconic acid, p-hydroxy phenylacetic acid, \u03b1-keto glutaric acid, propionic acid, D-saccharic acid, L-alanine, L-alanyl-glycine, L-asparagine, L-aspartic acid, L-glutamic acid, glycyl-L-aspartic acid, glycyl-L-glutamic acid, L-proline, L-serine, inosine, uridine, thymidine, glycerol, D,L-\u03b1-glycerol phosphate, glucose-1-phosphate, glucose-6-phosphate.16:0 (28.2%), Summed feature 4 (12.4%), C11:0 3-OH (11.6%), Summed feature 7 (10.5%) and Summed feature 3 (10.0%), with other minor components, including C14:0 (6.4%), C17:0 \u03949 cyclo (4.9%), C12:0 (4.3%), C14:0 2-OH (3.7%), iso-C18:0 (3.6%), C18:0 14-methyl (1.9%), C14:1 \u039411 (1.1%), C8:0 2-CH2CH3 (0.6%), C15:0 (0.5%). The DNA G+C content is 55.32 mol%.Major fatty acids are CT (= CGMCC 1.12772T = DSM 28222T = KCTC 42022T), was isolated from crickets (genus Teleogryllus) sampled in Chengdu , Sichuan Province, China.The type strain, SCU-B244S1 Certification(PDF)Click here for additional data file.S2 Certification(PDF)Click here for additional data file.S1 FigErwinia teleogrylli sp. nov. and the first 66 hit strains at the EzTaxon server within the family Enterobacteriaceae. Bar, 0.5% nucleotide substitutions. Numbers at branching points are bootstrap percentage values based on 1000 replications. Only values >50% are shown.The diagram shows the phylogenetic relationship between (TIF)Click here for additional data file.S2 FigErwinia teleogrylli sp. nov. and the first 66 hit strains at the EzTaxon server within the family Enterobacteriaceae. Bar, 0.5% nucleotide substitutions. Numbers at branching points are bootstrap percentage values based on 1000 replications. Only values >50% are shown.The diagram shows the phylogenetic relationship between (TIF)Click here for additional data file.S3 FigErwinia teleogrylli sp. nov. and taxa related type species of the genera Erwinia and Pantoea except Candidatus Erwinia dacicola. Escherichia coli ATCC 11775T was used as the outgroup. Bar, 0.5% nucleotide substitutions. Numbers at branching points are bootstrap percentage values based on 1000 replications. Only values >50% are shown.The diagram shows the phylogenetic relationship between (TIF)Click here for additional data file.S4 FigErwinia teleogrylli sp. nov. and taxa related species of genera Erwinia, Pantoea and Tatumella. Bar, 0.5% nucleotide substitutions. Numbers at branching points are bootstrap percentage values based on 1000 replications. Only values >50% are shown.The diagram shows phylogenetic relationship between (TIF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file."}
+{"text": "There are a number of errors in the legends for Figs. 3, 4, and 5. The complete, correct legends for Figs. 3, 4, and 5 are:Figure 3. Wallerian degeneration in mice lacking GD3s: tissue infiltration by activated macrophages and Schwann cell proliferation. A-B; D-E: Longitudinal sections of lesioned sciatic nerves from adult WT and GD3s KO mice at 5 days after crush lesioning immunolabeled for NF-200  or F4\u201380  and imaged at the distal nerve stump. The nuclei were counterstained with DAPI. C and F: Histograms indicating the number of NF-200 fragments (C) and active macrophages (cells positive for F4\u201380) at the distal nerve stump (F). G-H; J-K: Longitudinal sections of lesioned sciatic nerves from WT and GD3s KO mice imaged at the distal stump 7 days after crush lesioning and immunolabeled for p-histone H3 (G-H) or double immunolabeled for Ki-67 and GFAP (J-K). The nuclei were counterstained with DAPI. I and L: Histograms indicating the number of cells positive for phistone H3 (I) or KI-67/GFAP (L) at the distal stump in each group of mice. M-O: Optical slices obtained by confocal microscopy from transversal sections of wildtype uninjured (M), wildtype injured (N) or GD3s Ko injured (O) sciatic nerves immunolabeled for GFAP at distal stump, 5 days after crush lesion. Bars: A-B, G-H = 100 mm; and D-E, J-K = 50 mm; M-O = 20 mm. Statistics: Mann-Whitney, ns, p>0.05. doi: 10.1371/journal.pone.0108919.g003Figure 4. Committed nerve regeneration in adult mice lacking GD3s is restored by administration of exogenous GD3 in vivo and in vitro. A: Longitudinal sections of sciatic nerves proximally or 1 mm or 3 mm distally immunolabeled for GAP-43 at 21 days after crush lesioning. B: Histogram indicating the axonal density in the regenerating nerves from WT, GD3s KO and GD3-treated GD3s KO mice. C: Images of P1 mouse DRG explants seeded on PDL/laminin coverslips. The DRG samples from WT, GD3s KO and GD3s KO exogenously treated with GD3 ganglioside were incubated for 5 days in vitro. GD3 was added on day 2 of the incubation. Low-magnification images of DRGs immunolabeled for Tuj-1. D: Histogram quantifying neurite growth. E: High-magnification images of neurites immunolabeled for R24  or CD-60b . The nuclei were counterstained with DAPI (white) Bars: A = 100 mm; C = 500 mm; and E = 20 mm. Statistics: ANOVA ***p<0.001; *p<0.01. doi: 10.1371/journal.pone.0108919.g004Figure 5. Integrin-\u03b21 expression but not calcium influx is modified in neurons lacking GD3s. A-C: Images of integrin-\u03b21 obtained by apotome microscopy of mice neonate (P1) DRGs from WT (A), GD3s KO (B) and GD3s KO with exogenous GD3 (C). Samples were cultured for 5 days. A\u2032\u2013C\u2032: High-magnification optical sections of neurites (DIC) labeled for integrin-\u03b21. A\u2033-C\u2033: High-magnification optical sections of neurites (DIC) double labeled for integrin-\u03b21 and CD60-b (9-O-ac. GD3). Yellow dots indicate colocalization of the two markers. D: DIC image of the field shown in C, illustrating DRG (lower right) and neurites extended. E, E\u2032: Histogram of quantitative analysis of the number of integrin- \u03b21 (E) or 9-O-Ac. GD3 (E\u2032) clusters along extended neurites. F and J: P1 postnatal DRGs were dissected, cleaned, dissociated and cultured for 48 h in the presence of 50 ng/ml NGF. Cell cultures from both WT (F) and GD3s KO (J) mice show typical neurons with extensive neurites and flat Schwann cells. The same fields are shown under fluorescence . Further, H and L show the same microscope field under fura-2 fluorescence, in SCCI experiments. Typical responses are shown for 4 cells in WT (I) or in GD3s KO mice (M) when stimulated with 50 mM KCl  or 100 mM ATP . As shown in WT , cells #4 and #7 are neurons (with large cell bodies), whereas cells 14 and 10 are flat, typical of Schwann glia. The same is observed for GD3s KO cells , where cells #17 and #10 are neurons, and cells #28 and #39 are glia. N, O: Histograms indicating maximum calcium influx when stimulated by KCl  or ATP . Bars: A-D = 500 mm; A\u2032-C\u2032, A\u2032-C\u2033 = 100 mm F-H, J-L = 20 mm. Statistics: ns, p = 0.760 Mann-Whitney; ***p<0.0001, *p<0.001 ANOVA. doi: 10.1371/journal.pone.0108919.g005"}
+{"text": "P. aeruginosa an important pathogen causing lower respiratory tract infections (LRTI) both in HIV and non-HIV population. Molecular characterization of Pseudomonas spp. helps in better understanding of their clonal distribution among these patient populations. Our study aims to discriminate and generate highly specific fingerprints using 16S-rDNA PCR and amplified ribosomal DNA restriction analysis (ARDRA) techniques.Pseudomonas spp. were subjected to 16SrDNA PCR using universal primers and amplicons digested with Hae III, Alu I and Rsa I for ARDRA analysis. Nucleic acid size confirmation of the digested amplicons was done using MultiNA Bioanalyser. Phylogenetic tree was constructed by maximum parsimonious method using MEGA 4.0. Representative isolates from the major clones were sequenced and submitted to genbank for accession numbers and genetic relatedness was identified by UPGMA using NTSYS 1.80 software. Mean genetic distance (GD) and intraspecies mean GD were calculated.Seventy-two isolates  of P. aeruginosa, P.putida, P.stutzeri, Alcaligenes feacalis strains by BLASTN analysis. The overall mean GD and intra-species GD of the various species was as follows: P. aeruginosa : 4.664, 16.663, 7.536, 0.733, 0.096, 2.402 and 0.221, 3.487, 2.010, 0.029, 2.728, 0.011;P.putida:11.904, 7.3 and 1.064, 1.371; P.stutzeri (JF303641) is 2.732, 0.036; Alcaligenes feacalis (JF303642) : 30.248, 5.377 respectively.Based on ARDRA banding pattern, 14 groups and 10 clones were obtained from the 72 pseudomonas isolates with the following accession numbers: JF279962-64, JF303639-45. Sequences showed 97-100% similarity with known Pseudomonas spp causing LRTI among HIV and Non HIV patients.The highly variable mean GD values amongst the isolates from different and within the same species indicates high genetic diversity among"}
+{"text": "Nucleic Acids Res. 2014 Dec 16;42(22):13488\u201399. doi: 10.1093/nar/gku1097Streptococcus pneumoniae strain ST556, as shown in Table - M. Spn556I - M. Spn556II 8TTYG)- M. Spn556III 7TATC).In this article, the authors have incorrectly designated three DNA methyltransferases in In addition, the recognition sequence of the type I system encoded by MYY570-MMY572 in Figure 3A should be AAG(N)8TTYG."}
+{"text": "Deep-sea fungi, the fungi that inhabit the sea and the sediment at depths of over 1000 m below the surface, have become an important source of industrial, agricultural, and nutraceutical compounds based on their diversities in both structure and function. Since the first study of deep-sea fungi in the Atlantic Ocean at a depth of 4450 m was conducted approximately 50 years ago, hundreds of isolates of deep-sea fungi have been reported based on culture-dependent methods. To date more than 180 bioactive secondary metabolites derived from deep-sea fungi have been documented in the literature. These include compounds with anticancer, antimicrobial, antifungal, antiprotozoal, and antiviral activities. In this review, we summarize the structures and bioactivities of these metabolites to provide help for novel drug development. Chromocleista sp. was described by Park et al. chromen-4-one (96), together with six known compounds (97\u2013100) -4(97\u2013100) , were isectively .101)  , was isoectively .102) and dihydrooxosorbiquinol (103)  against P388, A549, HL60, BEL-7402, and K562 cell lines [Two new bisorbicillinoids, named oxosorbiquinol (ol (103) , have be104\u2013106), and the known breviones (107\u2013110) (Penicillium sp. and display strong cytotoxic effects against MCF-7 cells (IC50: 7.44\u201328.4 \u03bcM). Compound 106 exhibits cytotoxic activity against A549 cells with IC50 of 32.5 \u03bcM [111\u2013113)  [111\u2013113) show 25.2%\u201344.9% cytotoxicity against HeLa at 10 \u03bcg/mL. Particularly, compound 111 displays a very strong cytotoxicity to HIV-1 replication in C8166 cells with an EC50 of 14.7 \u03bcM [Three new breviane spiroditerpenoids, breviones I\u2013K (107\u2013110)  have bee 32.5 \u03bcM . Similar111\u2013113) , have be. (2009) . These c 14.7 \u03bcM .d-Pro-d-Phe) (114), cyclo(d-Tyr-d-Pro) (115), phenethyl 5-oxo-l-prolinate (116), cyclo(l-Ile-l-Pro) (117), cyclo(l-Leu-l-Pro) (118), and 3\u03b2,5\u03b1,9\u03b1-trihydroxy--ergosta-7,22-dien-6-one (119)  , have be120), and a known compound, cholesta-7,22-diene-3b,5a,6b-triol (121)  , were idlymerase .122), ferrineoaspergillin (123), aflatoxin (124), and (11S)-hydroxyl aspergillic acid (125) (Aspergillus sp. 16-02-1. These compounds possess certain cytotoxicity against human cancer cell lines K562 (33.6%\u201343.6%), HL-60 (24.1%\u201353.3%), HeLa (18.8%\u201345.4%), and BGC-823 (36.2%\u201351.2%) at the concentration of 100 \u03bcg/mL [New aspergillic acid (id (125)  have bee126)  [A known antibacterial compound, xanthocillin X (126) , was iso2 \u03bcg/mL) .127) (Penicillium sp. F00120. It inhibits in vitro proliferation of mouse melanoma (B16), human melanoma (A375), and human cervical carcinoma (HeLa) cell lines with IC50 of 0.08, 0.06, and 0.12 mM, respectively [One new sesquiterpene quinone, named penicilliumin A (127) , was isoectively .128), oxisterigmatocystin B (129), and oxisterigmatocystin C (130), together with one known compound, 5-methoxysterigmatocystin (131)  , have beectively .132) and 6-deoxy-5a,6-didehydrogliotoxin (133), and five known metabolites (134\u2013138) , whereas compounds 136\u2013138 containing a disulfide or tetrasulfide bond show potential inhibitory activity against histone methyltransferase (HMT) G9a (IC50: 2.1\u20136.4 \u03bcM) [Seven gliotoxin-related compounds including two new metabolites, bis(dethio)-10a-methylthio-3a-deoxy-3,3a-didehydrogliotoxin (134\u2013138)  have bee\u20136.4 \u03bcM) .Antimicrobials are the most important drugs to protect human beings from infective diseases. Deep-sea fungi are one of the potential pools for screening antimicrobial metabolites, which can be developed into new drugs.139\u2013143), together with six known analogues (144\u2013148) (Emericella sp. SCSIO 05240. All of them show weak growth inhibition against bacteria. The inhibition zone of compounds 139 and 141 against Escherichia coli (ATCC 29922), Klebsiella pneumonia (ATCC 13883), Staphylococcus aureus (ATCC 29213), Enterococcus faecalis (ATCC 29212), Acinetobacter baumanii (ATCC 19606), and Aeromonas hydrophila (ATCC 7966) is 1\u20133 mm in diameter. Moreover, compound 143 displays broad antifungal activities (3\u20134 mm in diameter) against Fusarium sp., Penicillium sp., Aspergillus niger, Rhizoctonia solani, Fusarium oxysporium f. sp. niveum, and Fusarium oxysporium f. sp. cucumeris [Four new prenylxanthones, emerixanthones A\u2013D 144\u2013148) , were is , were i149\u2013163) (Spiromastix sp. These compounds exhibit significant growth inhibition (MIC: 0.1\u20138.0 \u03bcg/mL) against Gram-positive bacteria including Staphylococcus aureus, Bacillus thuringiensis, and Bacillus subtilis. In addition, compounds 154\u2013158 display potential inhibitory effects on methicillin-resistant bacterial strains of Staphylococcus aureus (MRSA) and Staphylococcus epidermidis (MRSE). Moreover, compound 158 inhibits the growth of the vancomycin-resistant bacteria of Enterococcus faecalis and Enterococcus faecium (VRE) [Fifteen new depsidone-based analogues, named spiromastixones A\u2013O (149\u2013163) , have beum (VRE) .et al. (2013) [164)  against Staphylococcus aureus, Bacillus subtilis, Vibrio sp. 385, Vibrio sp. 333, and Vibrio sp. 1758 [Liu . (2013)  have rep3) [164)  that wassp. 1758 .165)  , has bees aureus .et al. (2014) [Acremonium implicatum DFFSCS001 (AI001), Aspergillus westerdijkiae DFFSCS013 (AW013), Alternaria tenuissima DFFSCS003 (AT003), Cladosporium cladosporioides DFFSCS016 (CC016), Cladosporium sphaerospermum DFFSCS019 (CS019), Engyodontium album DFFSCS021 (EA021), Geomyces vinaceus DFFSCS022 (GV022), and Tritirachium sp. DFFSCS034 (TS034). These fungal species were isolated from sediments of the South China Sea [Loktanella hongkongensis and Micrococcus luteus, and one marine pathogenic bacterium. Based on bioassay-guided isolation technique, they have isolated five compounds (166\u2013170)   have evahina Sea , and alm166\u2013170)  from the.9 \u03bcg/mL .171)  , has beef 276 \u03bcM .et al. (2014) [172), and five known diketopiperazine derivatives, diketopiperazine V (173), brevianamide Q (174), brevianamide R (175), brevianamide K (176), and brevianamide E (177) , 53.7% (174), 46.2% (175), 61.4% (176), and 19.3% (177) at 13.9 \u03bcM, respectively [Kong . (2014)  have rep E (177) , which wectively .178 (Aspergillus westerdijkiae SCSIO 05233 and shows strong antifouling activity with an EC50 of 8.8 mg/mL [Compound 178  has been.8 mg/mL .p-hydroxyphenopyrrozin (179)   , with a 79 , withnd (180)  that wasalbicans .181) and penicitrinone A (182)  at 100 \u03bcM [Two citrinin type compounds, phenol A acid ( A (182) , with ant 100 \u03bcM .The fungi in deep-sea environments are very diverse and abundant, making them a versatile reservoir of metabolites with both new structures and bioactivities that can be of potential use, acting as leading compounds to synthesize new modern medicine. Although the research on deep-sea fungi is not as up-to-date as the research on fungi in other environments such as terrestrial soil, fresh water, and shallow marine areas due to difficulties in both sample collection and fungal cultivation methods, more and more fungi have been cultivated from the deep sea based on culture-dependent methods. These deep-sea fungi can provide a potential source for natural bioactive product screening and new drug discovery. Up to now, more than 180 new and/or bioactive secondary metabolites from deep-sea fungi with broad bioactivities, such as anticancer, antimicrobial, antifungal, anti-larval settlement, and antiviral, have been described in the literature. Most of the investigated bioactive compounds exhibit cytotoxic activity, then antimicrobial activity. These bioactive compounds not only help deep-sea fungi to defend themselves against predators in the natural ecosystem, but also have the potential of becoming treatments for human diseases and probes for new biological targets. Work to isolate fungi from deep-sea environments and characterize their bioactive metabolites is underway and is of increased importance due to the urgent need for new drugs to overcome emerging and drug-resistant diseases."}
+{"text": "Headache is a very common health problem among adolescents, causing school and social disability . This isThree hundred and seventy-six adolescents aged 11-15 years, living in the Pavia province , were recruited for this cross-sectional school-based study. They were assessed about their headache using a medical history questionnaire; headache diagnosis was made according to the new classification system of the International Headache Society (ICDH-3 Beta). Headache-related disability was assessed by means of PedMIDAS (Pediatric Migraine Disability Assessment Score).Of the 376 students enrolled 91 24.2%) had headache: 60.44% girls (n=55) and 39.56% boys (n=36). At the first diagnosis, the prevalence of any migraine was 28.6% (n=26), any tension-type headache (TTH) 60.4% (n=55), any medication-overuse headache 3.3% (n=3) and \u201cunclassifiable\u201d headache was 7.7% (n=7) .None declared."}
+{"text": "BMS-663068 is a prodrug of BMS-626529, an attachment inhibitor that binds directly to HIV-1 gp120, preventing initial viral attachment and entry into the host CD4+ T-cell. AI438011 is an ongoing, Phase IIb, randomized, active-controlled trial investigating the safety, efficacy and dose\u2013response of BMS-663068 vs. atazanavir/ritonavir (ATV/r) in treatment-experienced (TE), HIV-1-positive subjects. At Week 24, response rates across the BMS-663068 arms were consistent with ATV/r.50 100 nM) were randomized equally to four BMS-663068 arms  and a control arm (ATV/r 300/100 mg QD), with tenofovir disoproxil fumarate (TDF) + raltegravir (RAL). The complete safety profile through Week 24 is reported.Antiretroviral TE subjects  with susceptibility to all study drugs (including BMS-626529 ICn=196)). In the ATV/r arm, Grade 2\u20134 drug-related AEs occurred in 14/51 (27.5%) subjects and were mostly secondary to gastrointestinal and/or hepatobiliary disorders. Serious adverse events (SAEs) occurred in 13/200 (6.5%) and 5/51 (9.8%) subjects receiving BMS-663068 and ATV/r, respectively; most were secondary to infections and none were related to study drugs. The most common AE reported for BMS-663068 was headache , occurring in 5/51 (10%) subjects in the ATV/r arm; in the BMS-663068 arms, this was not dose-related. There were no deaths.In total, 251 subjects were treated . No BMS-663068-related adverse events (AEs) led to discontinuation. Grade 2\u20134 drug-related AEs occurred in 17/200 (8.5%) subjects across the BMS-633068 arms; however, these events were mostly single instances and no dose-relationship was seen. Similarly, no noticeable trend for Grade 3\u20134 laboratory abnormalities was seen and Grade 3\u20134 hematologic changes and liver chemistry elevations were uncommon (neutropenia, 2.5%; AST/ALT elevations, 1% (BMS-663068 was generally well tolerated across all arms, with no related SAEs or AEs leading to discontinuation and no dose-related safety signals. There were no trends for Grade 2\u20134 AEs or clinical laboratory abnormalities. These results support continued development of BMS-663068.Previously submitted at IDWeek, Philadelphia, PA, 8 October 2014."}
+{"text": "HIV-infected patients treated with Highly Active Antiretroviral Therapy (HAART) may be predisposed to hypertriglyceridemia, which gives rise to a highly atherogenic lipid profile known as atherogenic dyslipidemia (AD). We propose that genetic variability leaves some HIV-infected patients more predisposed to AD than others , 2.This was a cross-sectional, observational study conducted in 468 antiretroviral-treated HIV-infected patients attending at the outpatient clinic of a tertiary hospital over a 6-month period, who were classified as normolipidemic (n=173) or presenting with AD (triglycerides: 1.7 mmol/L and HDLc < 1.02 [men] or 1.28 mmol/L [women]) (n=148). Polymorphisms were identified in the APOA5, APOC3, LPL, CETP, HL, MTP, APOE, LRP5 and VLDLR genes.Atherogenic dyslipidemia was detected in 31% of patients, most of whom were men (77%). This group was also older and had higher levels of remnant lipoprotein cholesterol (RLPc) than normolipidemic patients. The polymorphisms rs328 in LPL, rs708272 in CETP and rs1800588 in HL were 10\u201340% significantly more frequent in normolipidemic patients. At least 1 of these polymorphisms was detected in 90% of normolipidemic patients; in AD patients, the percentage decreased to 75% (p=0.003). This effect was dependent on both the allele and the dose of HAART and independent of the regimen administered. The protective combination showed a trend towards higher HDLc (1.13 [0.40] vs 1.24 [0.23] mmol/L), lower triglycerides (2.23 [2.34] vs 1.89 [1.24] mmol/L) and lower RLPc (16.41 [11.42] vs 12.99 [11.69] mmol/L).Polymorphisms in LPL, CETP and HL protect HIV-infected patients from developing AD in a dose-dependent manner ."}
+{"text": "Kabuki syndrome (KS) is a multiple congenital anomalies/intellectual disability syndrome characterized by developmental delay, specific facial features, skeletal and visceral abnormalities. This syndrome is caused by mutations in MLL2 and KDM6A gene. Autoimmune abnormalities such as idiopathic thrombocytopenic purpura, hemolytic anemia, thyroiditis, and vitiligo have been described very rarely in patients with KS ,2,3. HerCase report: A female patient aged 7 7/12 years presented with short stature. There was no consanguinity between her parents. She was born as a term neonate weighing 2100 g as the product of a twin pregnancy and had no perinatal complications. On physical examination, the patient was noted to have large and low-set ears, broad and arched eyebrows, elongated palpebral fissures with eversion of the lateral third of the lower eyelid, as well as high and narrow palate. She also had other phenotypic malformations such as numerous vitiligo lesions of different sizes in the neck, brachydactyly, prominent fetal finger pads, and hyperlaxity in her joints. Her height was 116.3 cm (-1.85 standard deviation score [SDS]) and weight was 21.7 kg (-0.78 SDS). She had sensorineural hearing loss  and moderate mental retardation . She had a normal female karyotype . A clinical diagnosis of KS was considered.4) was 0.42 ng/dL (reference range: 0.91-1.92), Anti-microsomal antibody: 450.6 U/mL (reference range: 0-9), anti-thyroglobulin antibody: 2766 U/mL (reference range: 0-4)]. Thyroid ultrasonography demonstrated a rough pattern, consistent with thyroiditis. Levothyroxine (50 \u00b5g/day) replacement therapy induced a euthyroid state (TSH: 3.65 mIU/L and fT4: 1.15 ng/dL).Additionally, laboratory findings revealed autoimmune thyroiditis [thyroid-stimulating hormone (TSH): 242 mIU/L (reference range: 0.55-6.7) and free thyroxine (fTThe main causes of KS are point mutations with large intragenic deletions and duplications of the histone methyl transferase MLL2 gene ,4,5. We In several patients, KS was reported to be associated with autoimmune abnormalities such as idiopathic thrombocytopenic purpura, hemolytic anemia, thyroiditis, and vitiligo ,5. The aPeer-review: Internal peer-reviewed."}
+{"text": "Current semi-quantitative cardiovascular magnetic resonance (CMR) techniques are of limited diagnostic value in patients with suspected myocarditis. T1- and T2-mapping CMR are promising novel quantitative approaches to assess myocardial injury. This study evaluated the performance of T1 and T2 mapping CMR to identify patients with myocarditis in comparison with conventional CMR techniques.This study included 104 patients with myocarditis and 26 controls, who underwent CMR at 1.5 Tesla. The CMR protocol included conventional sequences to assess myocardial edema (T2w-STIR), Early- (T1w-TSE) and Late-Gadolinium-Enhancement (PSIR). T1 quantification was performed using the modified Look-Locker inversion-recovery (MOLLI) sequence before and 15 minutes after administration of 0.075 mmol/kg Gadolinium-BOPTA. T2 quantification was performed using a free-breathing, navigator-gated multi-echo sequence. T1, T2 and extracellular volume (ECV) maps were calculated with a dedicated plug-in written for the OsiriX software. The diagnostic performance was compared between conventional parameters and global myocardial T1 (native and post-contrast), ECV and T2.The ROC areas-under-the-curve (AUC) to discriminate between patients with myocarditis and controls were 0.58 (p = 0.19) for the signal-intensity ratio myocardium/skeletal muscle on T2w-STIR, 0.67 (p = 0.01) for early myocardial enhancement on T1w-TSE and 0.80 (p < 0.0001) for presence of Late-Gadolinium-Enhancement, respectively. Furthermore, the AUC to discriminate between patients with myocarditis and controls were 0.77 (p < 0.0001) for native global myocardial T1, 0.59 (p = 0.14) for post-contrast global myocardial T1, 0.85 (p < 0.0001) for global myocardial ECV and 0.79 (p = 0.0001) for global myocardial T2, respectively.T1 and T2 mapping provide a superior performance to identify patients with myocarditis compared to conventional CMR techniques. In particular, the estimation of global myocardial ECV by T1 mapping improves the diagnostic value of CMR in patients with clinically suspected myocarditis.Marija-Orlovic-Foundation."}
+{"text": "The correct name is Christiane Rollenhagen. The correct citation is: Rollenhagen C, Macura SL, Lathrop MJ, Mackenzie TA, Doncel GF, Asin SN (2015) Enhancing Interferon Regulatory Factor 7 Mediated Antiviral Responses and Decreasing Nuclear Factor Kappa B Expression Limit HIV-1 Replication in Cervical Tissues. PLoS ONE 10(6): e0131919. doi:"}
+{"text": "To evaluate whole body diffusion-weighted MR imaging (WB-DWI MRI) for detection, staging and operability assessment in recurrent ovarian cancer compared with CT.Fifty-one women suspected for recurrent ovarian cancer underwent 3-Tesla WB-DWI/MRI using 2 b-values (b=0-1000 s/mm\u00b2), T2- and contrast T1-weighted sequences in addition to CT. WB-DWI/MRI and CT were compared for per-patient detection of recurrence, per-site detection of disease extent including peritoneal, serosal, retroperitoneal, periportal and distant metastases and for detecting disease extent according to institutional operability criteria. Imaging findings were correlated with surgical/pathological findings or imaging follow-up for at least 6 months.According to the reference standard, recurrence was confirmed in 48/51 patients. WB-DWI MRI showed 94% accuracy for detecting recurrence, versus 78% for CT. Per-site analysis showed significantly higher sensitivity of WB-DWI MRI over CT for assessing disease extent of the peritoneum, small bowel and colon mesentery and serosa , retroperitoneal suprarenal lymphadenopathies and periportal lesions (both p=0.031). Following institutional operability criteria, WB-DWI/MRI showed better sensitivity for detection of disease extent compromising operability; mesenteric root infiltration (p=0.008), carcinomatosis of small bowel (p=0.002) and colon (p=0.016), high volumetric peritoneal disease load (p=0.004) and irresectable distant metastases (p=0.016). WB-DWI MRI correctly predicted complete cytoreduction in 93% patients undergoing cytoreductive surgery versus 40% for CT.WB-DWI MRI showed higher accuracy compared with CT for recurrence detection while improving the sensitivity for staging and operability assessment of disease extent. WB-DWI MRI may be most valuable to select patients for surgical resection."}
+{"text": "Background. Plasmid-mediated quinolone resistance (PMQR) has received considerable attention recently. Data analysis in Jawaharlal Institute of Postgraduate Medical Education & Research (JIPMER) revealed 75% of the Enterobacteriaceae isolates to be ciprofloxacin-resistant in 2012. Few reports regarding the prevalence of PMQR are available from India. Hence, the present study was carried out to ascertain the prevalence of PMQR genes among clinical isolates of ciprofloxacin-resistant Enterobacteriaceae in JIPMER.Methods. The study included 642 ciprofloxacin-resistant clinical Enterobacteriaceae isolates. JIPMER hospital\u2019s annual consumption data for fluoroquinolones were retrieved from the Department of Pharmacy. The test isolates were screened for the presence of qnr A, B, D, S and aac(6\u2032)-Ib-cr genes. PMQR-positive isolates alone were tested for the presence of class I (intI1) and class II (intI2) integrons. Randomly selected PCR amplicons were sequenced and analysed using MEGA software. A total of 30 PMQR strains chosen at random were assessed for the transferability of the PMQR genes.Results. A majority of the strains exhibited high MIC values with 106 strains exhibiting MIC values >256 \u00b5g/mL. The aac(6\u2032)-Ib-cr gene had the highest prevalence at 64% (414) while, qnrB and qnrS genes were present in 15% (97) and 10% (64) of the isolates respectively. None of the strains were positive for qnrA and qnrD. All PMQR-positive isolates were screened for class I (intI1) and class II (intI2) integrons. Class I integron was found to be predominant among the test isolates with a few of them carrying both the classes of integrons. Transferability of PMQR genes to transconjugants was identified.Conclusion. The incidence of PMQR genes in the tertiary-care setup of the JIPMER hospital was found to be high which could be probably due to the increased prescription of fluoroquinolones. Thus, there is a need for rational usage of fluoroquinolones. The ATCC strain E. coli 25922 served as the quality control in the antimicrobial susceptibility test.A total of 642 clinical isolates belonging to the family Enterobacteriaceae resistant to ciprofloxacin by Kirby-Bauer disc diffusion method subsequently confirmed by agar dilution MIC were part of the study. Only one positive culture per patient was included. Standard methods were followed for isolation and identification of the bacteria from clinical specimens like blood, pus, CSF, etc. . E. coliE. coli ATCC 25922 was included as the quality control. The lowest concentration of antibiotic at which the growth of bacteria had been completely inhibited was recorded as the MIC.The discs for amikacin (30 \u00b5g), ceftriaxone (30 \u00b5g), ceftazidime (30 \u00b5g), ciprofloxacin (5 \u00b5g) and gentamicin (10 \u00b5g), meropenem (10 \u00b5g) were prepared in-house using antibiotics in pure power form  whereas cefoperazone-sulbactam disc was procured from Microxpress Tulip Diagnostics Pvt. Ltd, India. In particular cases, bacteria were also tested for imipenem  and piperacillin-tazobactam . The antibiotic susceptibility of the test isolates was interpreted as per CLSI M100-S25. Similarly, Minimum Inhibitory Concentration (MIC) values were determined according to CLSI M100-S25 guidelines for ciprofloxacin alone by agar dilution method using Muller Hinton agar procured from Himedia Laboratories, India. JIPMER hospital\u2019s annual prescribing data for fluoroquinolones were retrieved from Department of Pharmacy, JIPMER.intI1) and class II (intI2) integrons. Electrophoresis and staining analysed the PCR products with ethidium bromide.DNA templates were prepared from the overnight inoculum of test strains grown on Nutrient HiVeg\u2122 Agar  resuspended in MiliQ water after three rounds of washing. Crude template DNA was prepared by boiling lysis method . The reahttp://blast.ncbi.nlm.nih.gov/Blast.cgi) against the GenBank database of the National Center for Biotechnology Information. The phylogeny of the randomly sequenced aac(6\u2032)-Ib-cr amplicons were determined based on the nucleotide substitutions per site, i.e., the number of \u2019nucleotide substitutions\u2019 within the given sequence divided by the length of the sequence using MEGA software  were randomly selected for assessing the transferability of the PMQR genes following a previously described method (E. coli (J53) AziR (sodium azide-resistant) as the recipient strain. Transconjugants were selected on MacConkey agar containing sodium azide (100 \u00b5g/mL) and ciprofloxacin (0.5 \u00b5g/mL) and confirmed based on the results of biochemical and antimicrobial susceptibility tests carried out for transconjugants, recipient and donor bacterial cells. Screening of the transconjugants by PCR assay determined the transferability of PMQR genes.PMQR-positive strains numbering 30 -Ib-cr. The qnrB and qnrS genes were present in 97 (15%) and 64 (10%) isolates respectively. None of the strains were positive for qnrA & qnrD, indicating the absence of these qnr alleles among the clinical isolates included in the study (aac(6\u2032)-Ib-cr, qnrB and qnrS among the clinical isolates was found to be 64.49, 15.1 and 9.96 respectively. The overall prevalence of aac(6\u2032)-Ib-cr, qnrB and qnrS genes among the clinical Enterobacteriaceae strains isolated from JIPMER hospital could probably fall in the range of 60.72\u201368.12, 12.5\u201318.04 and 7.82\u201312.47 respectively.Remarkably, out of the 642 strains, as many as 414 (64.5%) harboured he study . The proE. coli had the maximum frequency of aac(6\u2019)-Ib-cr(197) and qnrB(43) genes. E. coli constituted almost half of the total aac(6\u2032)-Ib-cr positive isolates. On the other hand, the frequency of the qnrS gene was highest among K. pneumoniae isolates (32) with Klebsiella spp. altogether accounting for more than half of the total qnrS gene identified (47). The most interesting fact was that PMQR genes were more predominant among E. coli, Klebsiella spp., and Enterobacter spp. The occurrence of qnr alleles among Citrobacter spp., Serratia spp., Proteus mirabilis and Providencia spp. were almost negligible with Proteus mirabilis alone carrying the qnrB allele -Ib-cr gene. None of the isolates harboured qnrB and qnrS simultaneously. E. coli, Klebsiella spp., Enterobacter spp. and Proteus mirabilis were the organisms carrying multiple PMQR genes but, the association of qnrS with aac(6\u2032)-Ib-cr was seen only in E. coli and K. pneumoniae. The presence of aac(6\u2032)-Ib-cr and qnrS genes were associated with high MIC values (\u226564 \u00b5g/mL) and these associations were statistically significant with p-values <0.0000001 and <0.006261 respectively.The majority of the strains were found to carry one of the PMQR genes. But 7% of the isolates were found positive for multiple PMQR genes. All these isolates carried either Out of 528 PMQR positive isolates 212 were found to carry class I integron whereas, 95 isolates were found to carry class 2 integron and 47 isolates were positive for both the classes of integrons. However, we must admit that the study neither included the integron sequence analysis nor screened the integron-positive isolates for the presence of contiguous resistance gene cassettes.aac(6\u2032)-Ib-cr, four were positive for qnrB, and two were positive for qnrS genes. It is interesting to note that one particular transconjugant was found positive for both aac(6\u2032)-Ib-cr and qnrB.Of the 30 PMQR positive strains included in the conjugation only 18 transconjugants were successfully achieved. Among the transconjugants 11 were positive for aac(6\u2032) Ib-cr, seven qnrB and four qnrS strains were sequenced but none them had novel mutations within the nucleotide sequence. We have submitted nucleotide sequences of eight aac(6\u2032) Ib-cr, two qnrB and one qnrS genes to GenBank and accession numbers assigned are: KR080535, KR080536, KR080537, KR080538, KR080539, KR080540, KR080541 and KR080543 for aac(6\u2032)-Ib-cr, KR080544 & KR080545 for qnrB and KR080546 for qnrS. All the identified PMQR genes were found to be closely related based on the pair-wise distance matrix value. The overall distance matrix for aac(6\u2032)-Ib-cr was found to be 2.642 whereas the pair-wise distance matrix for the qnrB gene sequences was found out be 1.255. The study did not attempt to identify variants of qnr genes. The pair-wise distance matrix of the aac(6\u2032)-Ib-cr gene has been summarised as percentages, i.e., the number of nucleotide substitution per 100 nucleotide in the A total of 25 qnr determinants are qnrA, qnrB and qnrS -Ib-cr genes among our isolates was found to be high. The prevalence of aac(6\u2032)-Ib-cr in previously published reports ranged between 7% and 40% . Therefore, there was a significant association between increased MIC and the presence of PMQR genes, opening up the possibility that PMQR genes could have contributed to high MICs among the test isolates. But, we must agree that the efflux pump activities of these test isolates were not elucidated and their QRDR mutation profile was also not identified. Thus, this particular finding of the study is inconclusive as to how far PMQR genes have contributed to the increase in MIC of a strain against ciprofloxacin. With future investigation of these clinical isolates for QRDR mutations and efflux mechanisms, the prominence of PMQR in fluoroquinolone resistance can be elucidated.It is known that QRDR mutations induce high-level MICs while, PMQR genes induce low-level MICs . HoweverIn the present study, we have elucidated the prevalence of the plasmid-mediated quinolone resistance genes among clinical Enterobacteriaceae isolates recovered from a tertiary care hospital in Puducherry, India. Resistance to fluoroquinolones has predominantly increased with a majority of the isolates exhibiting high MIC values. Therefore, to combat this increased fluoroquinolone resistance it would be appropriate to use fluoroquinolones rationally for treating gram-negative infections. However, the significant finding of our study is that the prevalence of PMQR genes in JIPMER hospital is very high.qnr gene are on prevalence rates from around the world and reports on mechanistic aspects at the molecular level are very few. Future research should focus more the molecular mechanism of the PMQR genes and its encoded proteins.Moreover, the majority of the literatures on 10.7717/peerj.1995/supp-1Data S1Click here for additional data file.10.7717/peerj.1995/supp-2Data S2Click here for additional data file.10.7717/peerj.1995/supp-3Supplemental Information 1Click here for additional data file.10.7717/peerj.1995/supp-4Supplemental Information 2Click here for additional data file.10.7717/peerj.1995/supp-5Supplemental Information 3Click here for additional data file.10.7717/peerj.1995/supp-6Supplemental Information 4Click here for additional data file.10.7717/peerj.1995/supp-7Supplemental Information 5Click here for additional data file.10.7717/peerj.1995/supp-8Supplemental Information 6Click here for additional data file.10.7717/peerj.1995/supp-9Supplemental Information 7Click here for additional data file.10.7717/peerj.1995/supp-10Supplemental Information 8Click here for additional data file.10.7717/peerj.1995/supp-11Supplemental Information 9Click here for additional data file.10.7717/peerj.1995/supp-12Supplemental Information 10Click here for additional data file.10.7717/peerj.1995/supp-13Supplemental Information 11Click here for additional data file.10.7717/peerj.1995/supp-14Supplemental Information 12Click here for additional data file.10.7717/peerj.1995/supp-15Supplemental Information 13Click here for additional data file.10.7717/peerj.1995/supp-16Supplemental Information 14Click here for additional data file.10.7717/peerj.1995/supp-17Supplemental Information 15Click here for additional data file.10.7717/peerj.1995/supp-18Supplemental Information 16Click here for additional data file.10.7717/peerj.1995/supp-19Supplemental Information 17Click here for additional data file.10.7717/peerj.1995/supp-20Supplemental Information 18Click here for additional data file.10.7717/peerj.1995/supp-21Supplemental Information 19Click here for additional data file.10.7717/peerj.1995/supp-22Supplemental Information 20Click here for additional data file.10.7717/peerj.1995/supp-23Supplemental Information 21Click here for additional data file.10.7717/peerj.1995/supp-24Supplemental Information 22Click here for additional data file.10.7717/peerj.1995/supp-25Supplemental Information 23Click here for additional data file.10.7717/peerj.1995/supp-26Supplemental Information 24Click here for additional data file.10.7717/peerj.1995/supp-27Supplemental Information 25Click here for additional data file.10.7717/peerj.1995/supp-28Supplemental Information 26Click here for additional data file.10.7717/peerj.1995/supp-29Supplemental Information 27Click here for additional data file.10.7717/peerj.1995/supp-30Supplemental Information 28Click here for additional data file.10.7717/peerj.1995/supp-31Supplemental Information 29Click here for additional data file.10.7717/peerj.1995/supp-32Supplemental Information 30Click here for additional data file.10.7717/peerj.1995/supp-33Supplemental Information 31Click here for additional data file.10.7717/peerj.1995/supp-34Supplemental Information 32Click here for additional data file.10.7717/peerj.1995/supp-35Supplemental Information 33Click here for additional data file.10.7717/peerj.1995/supp-36Supplemental Information 34Click here for additional data file.10.7717/peerj.1995/supp-37Supplemental Information 35Click here for additional data file.10.7717/peerj.1995/supp-38Supplemental Information 36Click here for additional data file.10.7717/peerj.1995/supp-39Supplemental Information 37Click here for additional data file.10.7717/peerj.1995/supp-40Supplemental Information 38Click here for additional data file.10.7717/peerj.1995/supp-41Supplemental Information 39Click here for additional data file.10.7717/peerj.1995/supp-42Supplemental Information 40Click here for additional data file.10.7717/peerj.1995/supp-43Supplemental Information 41Click here for additional data file.10.7717/peerj.1995/supp-44Supplemental Information 42Click here for additional data file.10.7717/peerj.1995/supp-45Supplemental Information 43Click here for additional data file.10.7717/peerj.1995/supp-46Supplemental Information 44Click here for additional data file.10.7717/peerj.1995/supp-47Supplemental Information 45Click here for additional data file.10.7717/peerj.1995/supp-48Supplemental Information 46Click here for additional data file.10.7717/peerj.1995/supp-49Supplemental Information 47Click here for additional data file.10.7717/peerj.1995/supp-50Supplemental Information 48Click here for additional data file.10.7717/peerj.1995/supp-51Supplemental Information 49Click here for additional data file.10.7717/peerj.1995/supp-52Supplemental Information 50Click here for additional data file."}
+{"text": "Phytophthora infestans and Solanum nigrum, S. villosum and S. scabrum. European J of Plant Pathol 120(3): 233\u2013240. doi: 10.5580/e27. The correct reference 41 is: Rohani ER, Ismanizan I, Noor NM (2012) Somatic embryogenesis of mangosteen. Plant Cell Tiss Org Cult 110(2): 251\u2013259. doi: 10.1007/s11240-012-0147-4.References 40 and 41 are incorrectly switched. The correct reference 40 is: Lebrecka R (2008) Host-pathogen interaction between"}
+{"text": "Sesquiterpene hydrocarbons (C15) not only share the structural properties thought to lend protective qualities to isoprene and monoterpene hydrocarbons, but also react rapidly with ozone, suggesting that sesquiterpenes may similarly enhance tolerance of abiotic stresses. To test whether sesquiterpenes protect plants against ozone, UVB light, or drought, we used transgenic lines of the wild tobacco Nicotiana attenuata. The transgenic plants expressed a maize terpene synthase gene (ZmTPS10) which produced a blend of (E)-\u00df-farnesene and (E)-\u03b1-bergamotene, or a point mutant of the same gene (ZmTPS10M) which produced (E)-\u00df-farnesene alone,. (E)-\u00df-farnesene exerted a local, external, and transient ozone-quenching effect in ozone-fumigated chambers, but we found no evidence that enhanced sesquiterpene production by the plant inhibited oxidative damage, or maintained photosynthetic function or plant fitness under acute or chronic stress. Although the sesquiterpenes (E)-\u00df-farnesene and (E)-\u03b1-bergamotene might confer benefits under intermittent heat stress, which was not tested, any roles in relieving abiotic stress may be secondary to their previously demonstrated functions in biotic interactions.Among the terpenes, isoprene (C Supernatants were evaporated to 250 \u03bcL in a rotary evaporator and analyzed using an Agilent 1200 HPLC/MS/MS. Sample (5 \u03bcL) was injected and eluted with a flow rate of 800 \u03bcL/min through an Agilent Zorbax Eclipse DBC18 1.8 \u03bcm column. A gradient of 0.05% formic acid and acetonitrile (ACN) began with 5% ACN from 0\u20130.5 min and progressed linearly to 100% ACN at 4.5 min followed by washing with 100% ACN from 5\u20135.5 min and 3 min re-initialization at 5% ACN. SA and ABA were quantified relative to the labeled internal standards using MS/MS monitoring for the ion pairs 140.9/97.0 ([2H4] SA), 136.8/93.1 (SA), 269/159.2 ([2H6]ABA), and 263/153.2 (ABA) as in Wang et al. [Frozen leaf tissue (100 mg) was extracted overnight at -20\u00b0C with 1.5 mL methanol containing the labeled internal standards [g et al. .For each experiment , treatment effects and genotype-treatment interactions were tested by two-way ANOVA using SPSS . Phytohormone data were log-transformed and moisture data arcsine-transformed for homoscedasticity. Complete statistical results are documented in E)-\u03b2-farnesene with ozone, which has been observed in smog chambers [E)-\u03b2-farnesene into empty chambers containing ca. 500 ppb ozone lowered within-chamber ozone concentrations by >100 ppb within a few minutes (E)-\u03b2-farnesene from chambers containing 350 ppb ozone (E)-\u03b2-farnesene.We tested whether the reaction of -\u03b2-farnesene-mediated ozone destruction in the leaf headspace would protect plants, we supplemented the ozone-susceptible N. tabacum cv. BelW3 with external (E)-\u03b2-farnesene diluted in acetonitrile and impregnated into cotton tampons. A dose of 2000 \u03bcg plant-1 h-1 visibly reduced ozone-induced foliar necrosis (-1 h-1 was not protective (E)-\u03b2-farnesene from the 2000 \u03bcg plant-1 h-1 treatment reached and was retained in the leaf, although the 10-fold greater leaf retention in the control fumigation suggested that most (E)-\u03b2-farnesene was destroyed in the headspace -\u03b2-farnesene with ozone could protect plants by destroying ozone in the headspace around leaves. Yet whether endogenous (E)-\u03b2-farnesene production would be sufficient to achieve leaf protection, either by acting in the headspace or within the leaf itself, remained unclear. To test for benefits of endogenous production, we transformed N. attenuata with two versions of a maize sesquiterpene synthase expressed behind a constitutive 35S promoter. The enzyme ZmTPS10 (TPS10 line) yielded nearly equal amounts of (E)-\u03b2-farnesene and (E)-\u03b1-bergamotene, whereas the point mutant Zmtps10M produced primarily (E)-\u03b2-farnesene (E)-\u03b2-farnesene: t(2.29) = 1.11, p = 0.36; (E)-\u03b1-bergamotene: t(2.35) = 1.21, p = 0.33). Emission of the TPS10- and TPS10M-catalyzed sesquiterpenes from the transformed N. attenuata lines was approximately twice as high as the sesquiterpene emission rate measured from herbivore-damaged maize plants, and exceeded undamaged plant emission by an order of magnitude [External supplementation indicated that the reaction of  = 734.3, p<0.001) and significant decreases in moisture content  = 25.95, p<0.001)) and photosynthetic rate  = 7.81, p<0.016), all common effects of ozone exposure [F = 0.88, p = 0.37; moisture: F = 0.07, p = 0.80; photosynthetic rate: F = 0.04, p = 0.85). Surprisingly, ozone treatment had no significant effect on three markers of oxidative stress in leaves: malondialdehyde (MDA), a product of lipid peroxidation  = 0.55, p = 0.47); oxidative radical absorbance capacity (ORAC)  = 0.49, p = 0.50); and rutin  = 0.16, p = 0.69), a flavonoid glycoside that dominates the leaf phenolic profile of N. attenuata and whose induction plays a role in UVB tolerance [Sesquiterpene synthase-transformed and WT reatment . Ozone-fexposure . A two-wolerance . A TPS10E)-\u03b2-farnesene\u2019s reactivity and protective potential from the ozone experiments with BelW3, and we hypothesized that protection against UVB exposure would accrue from within-leaf terpenes, we used the TPS10M line with the highest leaf (E)-\u03b2-farnesene content . To control for effects of transformation unrelated to sesquiterpenes, we included a second, independently transformed TPS10M line .To directly test for within-leaf antioxidant effects of sesquiterpenes, we exposed sesquiterpene synthase-transformed and WT plants to UVB irradiation. Since we had evidence of  = 12.73, p = 0.001), while chlorophyll fluorescence  = 55.20, p<0.001) and photosynthetic rate  = 24.88, p<0.001) were significantly decreased, but no significant differences emerged between responses of TPS10M and WT plants  = 0.62, p = 0.55; chlorophyll fluorescence: F = 0.79, p = 0.47; photosynthetic rate: F = 0.42, p = 0.66). MDA measurements did not show a significant treatment effect  = 2.45, p = 0.14). The daily UVB exposure was continued for a total of 8 weeks, leading to significant decreases in height  = 157.00, p<0.001) and seed capsule number  = 38.15, p<0.001), but again, no differences in responses to treatment appeared between genotypes  = 0.50, p = 0.61; seed capsule number: F = 0.16, p = 0.86).The morphological responses of both TPS10M and WT leaves, which assumed a curled appearance and crispy texture, testified to the severity of the UVB exposure . After 8E)-\u03b2-farnesene-mediated resistance to drought stress using the same two TPS10M lines used in the UV experiment. Within 3 d of treatment onset, all drought-treatment plants raised their leaves, a sign of water stress in N. attenuata. At the time of tissue harvest, 6 days into the treatment, abscisic acid (ABA) levels were double those in controls  = 13.39, p = 0.001) and photosynthesis was decreased by 80%  = 562.33, p<0.001), but responses did not differ between TPS10M and WT  = 0.60, p = 0.56; photosynthesis: F = 0.12, p = 0.89). Although shoot MDA did not show a treatment effect  = 1.07, p = 0.31), root MDA increased  = 12.57, p = 0.001), but to a similar extent in all lines  = 0.71, p = 0.50). Ion leakage also increased  = 8.26, p = 0.007). Increases tended to be less dramatic in the (E)-\u03b2-farnesene emitter TPS10M1 than in WT, but TPS10M2 was no different from WT in its response, nor was the genotype x treatment term significant in an ANOVA  = 1.53, p = 0.23). Total fresh mass after 6 d of treatment  = 50.86, p<0.001), plant height after 2 weeks of treatment  = 152.19, p<0.001), and seed capsule count after an 11 d drying period in both treatment groups  = 16.94, p<0.001) all showed significant declines with drought, but no differences between the responses of TPS10M and WT  = 1.72, p = 0.20; height: F = 0.53, p = 0.59; seed capsule number: F = 1.06, p = 0.35).We tested for (5) and monoterpene (C10) hydrocarbons have been demonstrated to protect plants from oxidative stress. We tested whether sesquiterpene (C15) hydrocarbons, many of which react rapidly with ozone, also confer such protection when externally supplemented or internally generated. After demonstrating the ozone-quenching properties of the sesquiterpene (E)-\u00df-farnesene in empty fumigation chambers ((E)-\u00df-farnesene protected plants of an ozone-sensitive N. tabacum cultivar, BelW3, from visible cell death symptoms of ozone exposure ((E)-\u00df-farnesene was not protective. To test for benefits of endogenous sesquiterpene production, we used N. attenuata transformed with a Z. mays sesquiterpene synthase gene (35S::TPS10) that produced an (E)-\u00df-farnesene and (E)-\u03b1-bergamotene blend, or a point mutant of the same gene (35S::TPS10M) that produced primarily (E)-\u00df-farnesene. Augmenting endogenous sesquiterpene production by engineering these genes into N. attenuata did not enhance tolerance to oxidative stress under conditions of ozone, UVB, or drought.Plant-produced isoprene -\u00df-farnesene to quench headspace ozone (-1 h-1 conferred visible benefits (Z. mays [Solanum tuberosum [(E)-\u00df-farnesene (20 \u03bcg plant-1 h-1) was not protective ((E)-\u00df-farnesene recovered from supplemented, ozone-fumigated leaves -\u00df-farnesene and (E)-\u03b1-bergamotene, in the ecological model plant Nicotiana attenuata. The transformants grew and developed normally and had no significant physical or chemical differences from WT besides emission of the target volatiles [(E)-\u00df-farnesene recovered from leaves of BelW3plants externally supplemented with 2000 \u03bcg/plant ((E)-\u00df-farnesene were mediated by within-leaf activity.External supplementation experiments do not mimic the within-leaf sesquiterpene distribution resulting from olatiles . Sesquitolatiles . Because\u03bcg/plant , our train planta benefit. Based on the mechanisms proposed for sesquiterpene function, sesquiterpenes can be hypothesized to reduce levels of reactive oxygen species (ROS), especially in lipophilic cellular compartments, and thereby reduce oxidative damage and alter ROS-mediated signaling cascades [N. attenuata [Given the demonstrated reactivity of sesquiterpenes with ozone, we expected that sesquiterpene production might reduce ozone concentrations in the leaf boundary layer, as suggested for isoprene , or provcascades . Visiblettenuata ; shorterE)-\u03b2-farnesene emitted at 20 \u03bcg plant-1 h-1 had a short lifetime in the chamber headspace and did not confer benefits to neighboring plants. Diffusion effects [It is possible that the proximity of WT and transgenic plants in the fumigation chambers could have enabled WT to benefit from transgenic neighbors\u2019 emissions. However, our experiments with BelW3 showed that -\u00df-farnesene emission increases in a number of Pinus species in response to high light levels [Z. mays grown under high light [(E)-\u00df-farnesene was protective against the effects of UVB. UVB treatment strongly affected leaf morphology -\u00df-farnesene is stimulated by herbivory and leaf excision [E)-\u03b2-farnesene conferred no protection against this stress at either the chemical or whole-plant level: the effects of drought on ABA accumulation, electrolyte leakage, photosynthetic rate, and reproductive fitness correlates were similar in N. attenuata TPS10M and WT plants.If sesquiterpenes can diffuse across membranes and quench reactive oxygen species artments . Correlaartments . Althougartments ,58, in w winters . In Citrt stress . In Z. mexcision , which iexcision . In our E)-\u03b2-farnesene.Although our results did not demonstrate that sesquiterpenes ameliorate oxidative stress, they also do not directly refute the antioxidant hypothesis for terpene-mediated abiotic stress tolerance , since tOur results also do not directly challenge the membrane stabilization-mediated thermotolerance hypothesis . SharkeyA. thaliana, but this conclusion has been questioned [Z. mays [Spinacea olearacea thylakoids [Ours is not the first study to report that terpene supplementation does not increase tolerance to abiotic stress, but our study withstands the criticisms directed at earlier studies that showed a lack of benefit from added isoprene. Loivam\u00e4ki and colleagues  found noestioned  because [Z. mays . Isoprenylakoids  or increylakoids , but theN. attenuata. This wild tobacco has weathered extremes of temperature, drought, and light stress over its evolutionary history in the deserts of the American southwest [N. tabacum, might facilitate detection of terpene function in abiotic stress resistance. Genetic transformation of N. tabacum has previously revealed a small but significant role of isoprene production [A subtle protective influence of sesquiterpenes might be undetectable against the stress-adapted background of outhwest  and has outhwest . Using moduction  in toleroduction , and mig(E)-\u00df-farnesene and (E)-\u03b1-bergamotene in resistance to abiotic stress. Previous work suggests that these two sesquiterpenes may instead function against biotic stresses. ZmTPS10 transformation of A. thaliana attracted the parasitic wasp, Cotesia marginiventris, to its Lepidopteran host [(E)-\u03b1-bergamotene was emitted from N. attenuata in response to herbivore-mediated jasmonate signaling [N. attenuata\u2019s herbivores [ZmTPS10-produced sesquiterpenes may also primarily mediate biotic interactions in their native maize, where emission is more responsive to herbivory than to light, temperature, and humidity [(E)-\u00df-farnesene and (E)-\u03b1-bergamotene in biotic interactions [Our results argue against utility of ran host . Similarignaling  and attrrbivores . ZmTPS10humidity . Bioassaractions .A. thaliana plants under heat stress [N. attenuata, such signaling seems unlikely, since TPS10 and TPS10M transformants were chemically indistinguishable from WT and did not influence the defense physiology of WT neighbors when the two genotypes were grown together in the same pot [Another role for sesquiterpenes might be as within- or between-plant signals. The enhanced growth of isoprene-emitting t stress , and thet stress  or leaf t stress , would bt stress  or primet stress . In N. asame pot . Effects(E)-\u00df-farnesene and (E)-\u03b1-bergamotene, including the highly ozone-reactive \u03b1-humulene or (E)-\u03b2-caryophyllene [Sesquiterpenes other than phyllene , might bphyllene , a diverS1 Fig(DOCX)Click here for additional data file.S2 Fig(DOCX)Click here for additional data file.S3 Fig(DOCX)Click here for additional data file.S1 Table(XLSX)Click here for additional data file."}
+{"text": "In a subgroup, advanced HPs  of a medium-sized teaching hospital. Adult patients with severe sepsis were included and received standard goal-directed therapy (Surviving Sepsis Guidelines). Every patient received an arterial line and a central venous line in the upper diaphragm position. A subgroup received pulse contour cardiac output (PiCCO)-guided resuscitation and PT measurements. Pearson correlation coefficients (PCCs) were calculated between HPs and SL, which were measured every 4 hours for the first 48 hours after inclusion. n = 11) and pneumosepsis (n = 7). Mean HPs (with SD and range) were respectively: MAP 73 mmHg , HR 101 beats/minute , CVP 12 mmHg , UP 55 ml/hour , SL 3.2 mmol/l , CI 4.1 ml/kg/minute , GEDI 871 ml/m2 , ELWI 11 ml/kg , PT 32.1 C , and SvO2 75% . Relevant and significant (P < 0.005) PCCs between HPs and SL were respectively: MAP -0.417, HR 0.195, UP \u22120.237, SvO2 \u22120.204 and PT \u22120.569. Figure Twenty-five patients 12 men) were included. Mean age was 68 years (30 to 93), mean APACHE II score 31 (20 to 42). The most frequent reasons for IC admission were abdominal sepsis (2 men wer2 are significantly correlated to levels of SL, but clinical value might be limited due to the relatively low correlation coefficients. In a small subgroup, PT is better correlated to the level of SL.The conventional HPs MAP, HR, UP and SvO"}
+{"text": "Association of two HLA class I variants with HIV-1 pretreatment viremia, CD4+ T cell count at the care-entry and CD4+ T cell nadir.414 HIV-positive Caucasians (30% women) aged 19-73 years were genotyped for HLA-C -35 (rs9264942) and HLA-B*5701 variants. HIV-1 viral load, as well as CD4+ T cell count at care-entry and nadir, were compared across alleles, genotypes and haplotypes.HLA-C -35 C/C genotype was found in 17.6% patients, C/T genotype in 48.1%, and T/T genotype in 34.3% patients. HLA-B*5701 variant was present in 5.8% of studied population. HIV plasma viremia in the group with C allele was significantly lower (p=0.0002) compared to T/T group , while CD4+ T cell count at baseline was notably higher among C allele carriers compared to T/T homozygotes  (p=0.0007). Moreover, CD4+ T cell nadir among patients with C allele [median: 205 (IQR:83.5-390) cells/\u03bcl] was significantly higher compared to T/T group [median: 133 (IQR:46-328) cells/\u03bcl] (p=0.006). Among cases with HLA-B*5701 allele, significantly lower pretreatment viremia and higher baseline CD4+ T cell count were found  compared to HLA-B*5701 negative individuals. The lowest viremia (mean: 3.85 log [SD:1.3]) HIV-RNA copies/ml and the highest baseline and nadir CD4+ T cell [median: 476 (IQR:304-682) vs. median: 361 (IQR: 205-574) cells/\u03bcl, respectively) were found in individuals with HLA-B*5701(+)/HLA-C \u201335 C/C haplotype.HLA-C -35 C and HLA-B*5701 allele exert a favorable effect on the immunological (higher baseline and nadir CD4+ T cell count) and virologic  variables. This protective effect is additive for the compound HLA-B*5701(+)/HLA-C -35 C/C haplotype. Human leukocyte antigens C (HLA-C) strongly influence immunological activity in chronic viral infections  as well as in autoimmune diseases e.g. Crohn\u2019s disease, autoimmune liver diseases, Graves\u2019 disease and psoriasis vulgaris ,2,3,4,5.HLA-C antigens play an important role in HIV control through two mechanisms: acting as ligands for killer immunoglobulin-like receptors (KIRs) presented on natural killer (NK) cells and directly by antigen  presentation to cytotoxic T cells ,6,7.inter alia, on the level of HLA-C expression  (p = 0.0007) .Similarly, the CD4+ T cell nadir among patients with at least one HLA-C -35 C allele was significantly higher compared to T/T homozygotes [median: 205 (IQR: 84\u2013390) cells/\u03bcl vs. median: 133 (IQR: 46\u2013328) cells/\u03bcl] (p = 0.006) .In patients with HLA-B*5701variant, significantly lower pretreatment viral load and higher CD4+ T cell count at baseline were observed in comparison with HLA B*5701 negative group  were found in the group with the compound HLA-B*5701(+)/HLA-C locus -35 C/C haplotype Figs  and 2f.Associations between HLA-B*5701 and HLA-C -35 rs9264942 variants, HIV-1 viral load, baseline CD4+ T cell count and nadir were also analyzed separately for the HIV/HCV coinfected and HIV monoinfected groups. Results were in line with the findings presented for the entire group. In the HIV/HCV coinfected patients, lower pretreatment viral loads were observed for the HLA-C -35 C allele compared to T/T homozygotes [mean: 4.4 (SD: 1.0) vs. mean: 4.86 (SD: 0.85) log HIV-RNA copies/ml for the T/T] (p = 0.04), and for the HLA-B*5701 (+) [mean: 3.4 (SD: 0.49)] compared to the HLA-B*5701 (-) cases [mean: 4.6 (SD: 0.95)]. In patients without HCV coinfection, HLA-B*5701 (+) genotype was associated with higher baseline CD4+ T cell counts [median: 537 (IQR: 361\u2013682) cells/\u03bcl] vs. median 208 (IQR: 61\u2013500) cells/\u03bcl for the HLA-B*5701 (-) (p = 0.012), higher CD4+ T cell nadir [median: 437 (IQR: 313\u2013628) cells/\u03bcl] vs. median 162 (IQR: 45\u2013386) cells/\u03bcl (p = 0.08) and lower pretreatment viral loads (mean: 3.87 [SD: 1.3] vs. mean: 4.97 [SD: 0.91] log HIV-RNA copies/ml (p = 0.004). The remaining associations were not significant when analyzed separately for HIV/HCV coinfected and HIV monoinfected subgroups.2 = 0.321, p<0.0001) than among T/T homozygotes   versus r2 = 0.268, p<0.0001 for HLA-B*5701(-)]   Fig  and 3b, (-)] Fig  and 3d. 019) Fig  and 3f.HCP5 locus favorable for the viremia set-point in Caucasians, was not confirmed for Afro-American population [Favorable effects of certain HLA class I alleles on the outcomes of HIV infection have been shown in many studies , 22, 23.pulation .Moreover, some studies were conducted in the populations infected with divergent HIV subtypes ,25. Our We found that the -35 SNP rs9264942 allele C is the strongest marker for the lower baseline HIV plasma viremia as well as higher CD4+ T cell count and CD4+ T cell nadirs. It is consistent with the previous studies presenting advantageous influence of this polymorphism , 22, 28.The novel finding of this report includes the observation that the HLA-B*5701(+)/HLA-C rs9264942 C/C haplotype is associated with the most favorable pattern of influence on HIV viremia and CD4+ T cell count among the analyzed variants. This haplotype is clearly linked with the lowest pretreatment plasma viral loads, the highest CD4+ T cell counts at care-entry, as well as with the highest nadir of CD4+ T cell count, and reflects delayed progression to immunodeficiency.We found that the coexistence of two protective HLA gene variants results in an additive protective effect on HIV infection. Some studies revealed that HLA-C variants independently control the HIV infection ,13, 22, The protective host genes exert immune pressure on the virus, which creates \u201cescape\u201d mutations. Such finding connected with C/C variant of rs9264942 was presented by Blais and colleagues . The freThe limitation of the study is related to the fact, that it was impossible to assess the time from infection to the care entry, therefore differences in HIV plasma viremia and CD4+ T cell counts related to the time of infection in association with investigated HLA variants were not analyzed.To conclude, we have confirmed the protective effect during the HIV infection of the analyzed HLA variants, including additive one for the compound haplotype. Analysis of genetic variants across various ethnic groups allows confirming the consistence of the genetic effect exerted by the investigated variants."}
+{"text": "It has been unclear whether thrombolytic-related asymptomatic hemorrhagic transformation (AHT) affects the clinical outcome. To answer this question, we examined whether thrombolytic-related AHT affect short-term and long-term clinical outcome.All data were collected from the Thrombolysis Implementation and Monitor of Acute Ischemic Stroke in China (TIMS-China) registry. The patients were diagnosed as having AHT group and non- hemorrhagic transformation (HT) group based on clinical and imaging data. The patients with symptomatic hemorrhagic transformation were excluded from this study. Thrombolytic-related AHT was defined according to European-Australasian Acute Stroke Study (ECASS) II criteria. 90-day functional outcome, 7-day National Institutes of Health Stroke Scale (NIHSS) score, 7-day and 90-day mortalities were compared between two groups. Logistic regression analysis was used to evaluate the effects of AHT on a short-term and long-term clinical outcome.904 of all 1440 patients in TIMS-China registry were enrolled. 89 (9.6%) patients presented with AHT after thrombolysis within 24-36h. These patients with AHT were more likely to be elder age, cardioembolic subtype, and to have higher National Institutes of Health Stroke Scale score before thrombolysis than patients without AHT. No significant difference was found on the odds of 7-day (95% CI:0.692 (0.218\u20132.195), (P = 0.532) or 90-day mortalities (95% CI:0.548 (0.237\u20131.268), P = 0.160) and modified Rankin Score(0\u20131) at 90-day (95% CI:0.798 (0.460\u20131.386), P = 0.423) or modified Rankin Score(0\u20132) at 90-day (95% CI:0.732 (0.429\u20131.253), P = 0.116) or modified Rankin Score(5\u20136) at 90-day (95% CI:0.375 (0.169\u20131.830), P = 0.116) between two groups.Thrombolytic-related AHT does not deteriorate short-term and long-term clinical outcome. Intravenous thrombolysis with alteplase (recombinant tissue plasminogen activator) is the most effective therapy for acute ischemic stroke ,2. HowevAll of data were collected from the Thrombolysis Implementation and Monitor of acute ischemic Stroke in China (TIMS-China) registry , a natioEligible patients received intravenous thrombolysis within 4.5-hour. Patients with incomplete data and thrombolysis time window>4.5h or unknown were excluded. Patients with NIHSS score \u226525were excluded. Patients with symptomatic HT were excluded from this study. Eligible patients were divided into two groups: the AHT group, which had asymptomatic hemorrhagic transformation; and the non-HT group, which had no thrombolytic-related hemorrhagic transformation .The registry of TIMS-China was approved by ethics committee of Beijing Tiantan hospital in 2006. The data from the TIMS-China registry was anonymous before access and analysis by all researchers and patient treatment was assigned and determined independent of all researchers. All patients had written informed consent before being entered in the registry.Follow-up CT scans for patients were collected at 24\u201336 hours and 7-day. The diagnosis of HT on brain images was determined independently by 2 neurologists blinded to clinical data. If there was a disagreement between 2 neurologists, a third neurologist was consulted, and a consensus decision was reached. Hemorrhagic transformation must be scored according to the ECASS II classification [Results were reported as mean \u00b1 SD or the frequency of categorical variables. Pearson chi-square (\u03c72) test was used for comparisons of categorical variables. Continuous variables were analyzed using t test or Mann-Whitney U test. Percentage proportions of outcome events were calculated by dividing the number of events by the total number of patients. For each outcome variable, a separate multivariable logistic regression was performed. Odds ratios (ORs) with its 95% confidence intervals (CI) were calculated by using non-HT group after thrombolysis as the reference group. Two-sided values of P less than .05 were considered statistically significant. All statistical analyses were performed using SAS software version 9.3 .904 patients (62.8%) were enrolled from a patient population of 1440 from 67 centers across TIMS-China. Our exclusion criteria were as follows: (1) incomplete neuroimaging workups (n = 40), (2) loss to follow-up (n = 36), (3) thrombolysis time window>4.5h or unknown (n = 312), (4) symptomatic HT (n = 24), (5) Patients with severe stroke with a NIHSS score >25(n = 93). It had been reported that 89(9.6%) patients were transformed to AHT in China , 24  and mRS score (0\u20132) in patients after thrombolysis when we analyze the data without any adjustment for covariates. After adjustment for baseline relevant factors, no significant difference was found on the odds of 7-day (95% CI:0.692 (0.218\u20132.195), (P = 0.532) or 90-day mortalities (95% CI:0.548 (0.237\u20131.268), P = 0.160) and modified Rankin score(0\u20131) at 90-day (95% CI:0.798 (0.460\u20131.386), P = 0.423) or modified Rankin score(0\u20132) at 90-day (95% CI:0.732 (0.429\u20131.253), P = 0.116) or modified Rankin score(5\u20136) at 90-day (95% CI:0.375 (0.169\u20131.830), P = 0.116) between two groups . In AHT There were remarkably low rates of asymptomatic hemorrhagic transformation 4.5%)  in the N.5%  in tPrevious some studies have shown that thrombolytic-related AHT does not have a negative effect on the short-term or long-term functional outcome. AHT after thrombolysis has a suggestive of vascular recanalization and effective treatment . MoreoveDifferent from mechanism of symptomatic HT, we speculate that AHT may link with small artery formation rather than large artery formation. It has been showed that thrombolysis after acute ischemic infarction may lead to artery reopening and collateral circulation reperfusion6 . MoreoveOur study may have the following limitations. (1) Our sample size was relatively small, and our data exclude severe stroke(NIHSS> or = 25), which maybe have a subtle effect (either positive or negative) of AICH outcome. (2) Another limitation is that the study did not follow the randomized, double-blind principles. Despite these limitations, our results have implications that post-thrombolytic AHT in China does have not a negative effect on clinical prognosis.S1 Table(DOCX)Click here for additional data file.S2 Table(XLS)Click here for additional data file."}
+{"text": "There is a lack of data on potential gender differences in the use of interventions to prevent and treat cardiovascular disease (CVD) in HIV-positive individuals. We investigated whether such differences exist in the D:A:D study.Follow-up was from 01/02/99 until the earliest of death, 6 months after last visit or 01/02/13. Rates of initiation of lipid-lowering drugs (LLDs), angiotensin-converting enzyme inhibitors (ACEIs), anti-hypertensives and receipt of invasive cardiovascular procedures  were calculated in those without a myocardial infarction (MI) or stroke at baseline, overall and in groups known to be at higher CVD risk: (i) age >50, (ii) total cholesterol >6.2 mmol/l, (iii) triglyceride >2.3 mmol/l, (iv) hypertension, (v) previous MI, (vi) diabetes, or (vii) predicted 10-year CVD risk >10%. Poisson regression was used to assess whether rates of initiation were higher in men than women, after adjustment for these factors.n=13,039; median (interquartile range) 34 (29\u201340) years) were younger than men  years, p=0.001), and were less likely to be current smokers , to have diabetes  or to have hypertension . Of 49,071 individuals without a MI/stroke at enrolment, 0.6% women vs. 2.1% men experienced a MI while 0.8% vs. 1.3% experienced a stroke. Overall, women received ICPs at a rate of 0.07/100 person-years (PYRS) compared to 0.29/100 PYRS in men. Similarly, the rates of initiation of LLDs (1.28 vs. 2.46), anti-hypertensives (1.11 vs. 1.38) and ACEIs (0.82 vs. 1.37) were all significantly lower in women than men (At enrolment, women (than men . As expethan men . Use of most CVD interventions was lower among women than men in the D:A:D study. Our findings suggest that actions should be taken to ensure that both men and women are monitored for CVD and, if eligible, receive appropriate CVD interventions."}
+{"text": "Pressure-support ventilation improves lung mechanics, blood gas exchange, hemodynamics, and work of breathing (WOB) in mild acute respiratory distress syndrome (ARDS) , 2. NeveT=6ml/kg) using two PEEP levels (2 and 5 cmH2O) in a mild ARDS model.To compare PSV and PCV target to protective tidal volume  instillation. After 24 hours, animals were anesthetized, tracheotomized, and their lungs were mechanically ventilated in PSV to achieve VT = 6 ml/kg. After baseline data collection, animals were randomly divided to four groups (n=8/group):2O (PCV-P2);PCV + PEEP = 2 cmH2O (PCV-P5);PCV + PEEP = 5 cmH2O (PSV-P2);PSV + PEEP = 2 cmH2O (PSV-P5).PSV + PEEP = 5 cmHThirty-two male Wistar rats (310 \u00b1 19 g) were submitted to intratracheal RS) and peak transpulmonary  pressures, and pressure-time product (PTP), as a surrogate of WOB, were evaluated.Animals were ventilated for 2 hours. Mean arterial pressure (MAP), arterial blood gases, peak airway . In accordance, PTP was lower in animals submitted to PEEP = 5 cmH2O compared to PEEP = 2 cmH2O during PSV .All animals showed better oxygenation along time, regardless of ventilator strategy. Animals submitted to PCV, regardless of PEEP, received more colloids to keep MAP>70 mmHg. Ppeak,In a mild ARDS model, pressure-support ventilation is associated to better hemodynamics, lung mechanics, and it seems to have a dependent effect of the adjusted PEEP level, as depicted by work of breathing.CNPq, FAPERJ, CAPES, PRONEX, MS-DECIT"}
+{"text": "Systolic myocardial strain is load dependent, but the CMR literature largely disregards effects of myocardial afterload on strain and strain rate. This may reflect well known limitations of conventional myocardial afterload assessment using wall stress analyses, which are based on erroneous assumptions about left ventricular(LV) geometry and/or myocardial material properties. Therefore, we compared the utility of a nongeometric afterload index(NGI) derived from LV pressure(P) volume(V) and mass, which requires no assumptions about material properties, to that of conventional noninvasive end-systolic circumferential stress(CWS) as determinants of CMR LV circumferential strain(CST), ejection fraction(EF) and strain rate(SR) in normals(NL) and patients with nonischemic dilated cardiomyopathy(CM).We obtained breath-hold volumetric short-axis SSFP cines, cuff systolic P, systolic duration, LVV, M and EF, feature-tracking global CST(TomTec Imaging Systems) and mean SR in NLsyrs) and CM  yrs, EF 27.2%(sd10.8%). CWS was calculated using Mirsky's formula(Biophys. J.1969) while NGI was determined as end-systolic PV/M.EF, CST and CSR were markedly reduced in CM compared to NL(EF 27.2%sd10.8)vs 58.4%(4.6), (-53%)p < 0.0001; CST -10.7%(5.3) vs -23.9%(4.3), (-55%), p < 0.0001);(CSR -32.1%/s(14.8) vs -65.7(14.9) p < 0.0001). But CWS was also markedly elevated in CM versus NL(CWS 307.6(9.2) vs 176.2(42.1)x 103 dyn/cm2,(+75%), p < 0.0001). Thus afterload excess due to adverse LV remodeling, may account for most EF, strain and strain rate reduction in CM. However, PV/M was more markedly increased than CWS, (162.6(sd48.9) vs 84.4(18.4)(+93%) p < 0.0001) and correlated more closely and significantly with EF, CST and CSR than CWS in the expected inverse relationship in both NL and CM subgroups(Table 0.8vs 58.Afterload excess due to adverse LV remodeling is an important determinant of reduced myocardial and LV chamber function in CM, making a major contribution to reductions in EF, CST and SR, but PV/M, a simple, nongeometric afterload index, is superior to conventional wall stress calculation as a quantitative afterload index.St. Francis Research Foundation."}
+{"text": "P1\u2009=\u20090.007; CR vs. SD: P2\u2009<\u20090.001; PR vs. SD: P3\u2009=\u20090.004), 95.7% vs. 88.7% vs. 70.2% , 92.0% vs. 87.4% vs. 74.3%  and 95.9% vs. 88.8% vs. 81.8% , respectively. Multivariate analysis identified that the tumour response to IC was an independent prognostic factor for DFS, OS and LRRFS.The prognostic value of the tumour response to induction chemotherapy (IC) for long-term survival outcomes after intensity-modulated radiation therapy in nasopharyngeal carcinoma (NPC) remains unknown. We retrospectively reviewed 1811 consecutive patients with newly diagnosed NPC treated using IMRT, and 399 eligible patients with pre- and post-induction chemotherapy magnetic resonance images were recruited. The clinicopathological features of patients with different tumour responses were compared using the Chi-square test or Fisher\u2019s exact test. Prognostic value was assessed using a multivariate Cox proportional hazards model. After IC, 101/399 (25.3%) patients had a complete tumour response overall (CR), 262 (65.7%) had a partial response (PR) and 36 (9.0%) had stable disease (SD). The 4-year disease-free survival (DFS), overall survival (OS), distant metastasis-free survival (DMFS) and locoregional relapse-free survival (LRRFS) rates for CR vs. PR vs. SD were 90.0% vs. 79.0% vs. 58.2% (CR vs. PR:  Nasopharyngeal carcinoma (NPC) is a cancer with an extremely unbalanced geographical distribution, with an age standardised incidence rate of 20\u201350 per 100,000 males in south China2345et al. reported that neoadjuvant docetaxel-cisplatin followed by CCRT provided a 3-year overall survival (OS) benefit in stage III-IVB NPC891011The value of additional induction chemotherapy (IC) before CCRT in advanced NPC has received intense investigation. In 2009, Hui et al. revealed that the unsatisfactory tumour response after induction chemotherapy could predict poor prognosis for patients with advanced-stage NPCHowever, despite these negative outcomes, IC may have potential clinical value. Previous studies reported that the response to chemotherapy correlate with clinical outcome1314Of the 1811 patients with newly diagnosed non-metastatic NPC treated between November 2009 and February 2012 at Sun Yat-sen University Cancer Center, and the 399 patients for whom both pre- and post-induction chemotherapy magnetic resonance (MR) images of the nasopharynx and cervical region were available were retrospectively analysed. This study was conducted in compliance with the institutional policy regarding the protection of patients\u2019 private information and approved by the Research Ethics Committee of Sun Yat-sen University Cancer Center. All the methods were carried out in accordance with the approved guidelines of Sun Yat-sen University Cancer Center. Written informed consent was obtained from all patients prior to therapy.Routine staging workup included a complete history, clinical examinations of the head and neck, direct fibre-optic nasopharyngoscopy, magnetic resonance imaging (MRI) of skull base and whole neck, chest radiography, whole-body bone scan, abdominal sonography, and positron emission tomography-CT if clinically-indicated. Immunoglobulin A antibodies against EBV viral caspid antigen (VCA-IgA) and Epstein Barr virus early antigen (EA-IgA) were quantified. All patients underwent dental evaluations before RT.th edition of the International Union against Cancer/American Joint Committee on Cancer (UICC/AJCC) staging systemPatients were restaged according to the 7b-values\u2009=\u20090 and 1000 s/mm2; three signal averages) were obtained before contrast injection. After intravenous administration of gadopentetate dimeglumine , axial and sagittal T1-weighted spin-echo and coronal T1-weighted fat-suppressed spin-echo sequences were performed .All patients underwent MRI of the region from the suprasellar cistern to the inferior margin at the sternal end of the clavicle using a head-and-neck coil with a 3 Tesla system . T1-weighted fast spin-echo images in the axial, coronal and sagittal planes (repetition time [TR]/echo time [TE]\u2009=\u2009650 ms/9 ms), T2-weighted fast spin-echo MR images in the axial plane (TR/TE\u2009=\u20092470 ms/90 ms) and a spin-echo echo-planar DWI sequence All patients underwent IMRT while immobilized using a custom head-to-neck thermoplastic cast with the patient\u2019s neck resting on a support. A high-resolution contrast planning CT scan was taken from the vertex to 2 cm below sternoclavicular joint . Target volumes were delineated slice-by-slice on treatment planning CT scans using an individualized delineation protocol that complies with International Commission on Radiation Units and Measurements reports 50 and 62. Prescribed doses were 66\u201372 Gy at 2.12\u20132.43 Gy/fraction to planning target volume (PTV) of primary gross tumour volume (GTVnx), 64\u201370 Gy to PTV of GTV of involved lymph nodes (GTVnd), 60\u201363 Gy to PTV of high-risk clinical target volume (CTV1), and 54\u201356 Gy to PTV of low-risk clinical target volume (CTV2). All targets were treated simultaneously using the simultaneous integrated boost technique.2) with 5-fluorouracil (1000 mg/m2) (PF), cisplatin (75 mg/m2) with docetaxel (75 mg/m2) (TP), or cisplatin (60 mg/m2) with 5-fluorouracil (600 mg/m2) and docetaxel (60 mg/m2) (TPF) every three weeks for two or three or more cycles. Concurrent chemotherapy was cisplatin weekly (30\u201340 mg/m2) or on weeks 1, 4 and 7 (80\u2013100 mg/m2) of radiotherapy.According to institutional guidelines, we recommended RT alone for stage I, concurrent chemoradiotherapy (CCRT) for stage II, and CCRT +/\u2212 IC/adjuvant chemotherapy (ACT) for stage III to IVA-B. IC consisted of cisplatin . The end points (time to first defining event) were disease-free survival (DFS), overall survival (OS), distant metastasis-free survival (DMFS), locoregional relapse-free survival (LRRFS), local relapse-free survival (LRFS) and regional relapse-free survival (RRFS).P\u2009<\u20090.05 was considered significant. Stata Statistical Package 12  was used for all analyses.The Chi-square test or Fisher\u2019s exact test were used to compare categorical variables and treatment failure patterns between the CR, PR and SD groups. Life-table estimation was performed using the Kaplan-Meier method and log-rank test. The multivariate Cox proportional hazards model was used to estimate hazard ratios (HRs) and 95% confidence intervals (CIs); age, gender, smoking, drinking, pathological type, T category, N category, concurrent chemotherapy, overall response were included as variables. All tests were two-sided; n\u2009=\u2009308)-to-female (n\u2009=\u200991) ratio was 3.4:1. The median age for the whole cohort was 45 years . The baseline characteristics of patients were list in For the 399 patients, the male  patients, PR for 262 (65.7%) patients, and SD for 36 (9.0%) patients. No patients had PD after IC. The local tumour response was a CR for 139/399 (34.8%) patients, PR for 195 (48.9%) patients and SD for 65 (16.3%) patients. Nineteen (4.8%) patients had N0 disease and were not included in the analysis of regional response. With regards to regional tumour response, 167/380 (43.9%), 166/380 (43.7%) and 47380 (12.4%) patients had a CR, PR and SD, respectively.P\u2009=\u20090.002; 58.3% vs. 40.1%, P\u2009=\u20090.038) and had advanced T category  compared to patients who achieved a CR or PR. Obviously, patients with CR received more cycles of induction chemotherapy compared with that of patients with PR (P\u2009=\u20090.009) or SD (P\u2009<\u20090.001). No significant associations were observed between any other clinicopathological feature and the overall tumour response.Significantly more patients with SD were previous smokers . By final follow-up, 21/399 (5.3%) patients had developed local failure, 23/399 (5.8%) patients developed regional failure and 51/399 (12.8%) patients developed distant failure.P\u2009=\u20090.022) and distant failure  after IMRT than patients with a CR (P\u2009=\u20090.054).With regards to overall tumour response after induction chemotherapy, patients with SD had higher rates of local failure . The local failure rates of the local PR and CR groups were almost significantly different . Patients with a regional CR after induction chemotherapy had a lower rate of distant failure after IMRT than patients with a regional PR  and regional SD . Additionally, patients with a regional CR had a lower rate of regional failure after IMRT than patients with a regional PR .Patients with local SD after induction chemotherapy had higher rates of local failure after IMRT compared to patients with a local CR , 95.7% vs. 88.7% vs. 70.2% , 92.0% vs. 87.4% vs. 74.3%  and 95.9% vs. 88.8% vs. 81.8% , 92.3% vs. 89.2% vs. 80.6% , 97.7% vs. 94.2% vs. 88.5%  and 90.5% vs. 85.7% vs. 85.9% , 93.7% vs. 88.7% vs. 71.5% , 97.5% vs. 91.1% vs. 93.1%  and 92.6% vs. 86.4% vs. 73.5%  is associated with a poorer prognosis in NPC, breast cancer and gastric cancerNPC is highly chemosensitive and radiosensitive. Although most patients with advanced NPC respond well to chemotherapy, recurrence of distant metastases is the major cause of treatment failure and has a poor prognosisDespite the outcome that unsatisfactory (PR or SD) tumour response after IC indicated poor prognosis, we should pay attention to the factors which may affect the result. Firstly, using the RECIST standardThe current study showed that patients with unsatisfactory tumour responses after IC have poorer prognosis compared with that of patients with satisfactory tumour responses. Therefore, we should pay more attention to patients with SD after induction chemotherapy in clinical practice. More intensive treatment regimens like higher radiation dose and adjuvant chemotherapy should be considered. Moreover, regular follow-up should also be provided to detect early recurrence and reduce the potential risk of distant failure.This study is limited by its retrospective nature and fact the follow-up time may have been insufficient, though we selected DFS as the major endpoint to address this shortcoming. Moreover, many other prognostic factors including pre-treatment plasma Epstein-Barr virus DNA load272830Tumour response to induction chemotherapy is an independent prognostic factor for patients with NPC receiving IMRT. Assessing tumour response after induction chemotherapy may assist prognostication and refine the treatment of high-risk patients with NPC. The outcomes in this current study need to be confirmed by prospective studies with large cohorts.How to cite this article: Peng, H. et al. The Tumour Response to Induction Chemotherapy has Prognostic Value for Long-Term Survival Outcomes after Intensity-Modulated Radiation Therapy in Nasopharyngeal Carcinoma. Sci. Rep.6, 24835; doi: 10.1038/srep24835 (2016)."}
+{"text": "Mucuna pruriens is the best known natural source of L-dopa, the gold standard for treatment of Parkinsonism. M. pruriens varieties are protein rich supplements, and are used as food and fodder worldwide. Here, we report L-dopa contents in seeds of fifty six accessions of four M. pruriens varieties, M. pruriens var. pruriens, M. pruriens var. hirsuta, M. pruriens var. utilis and M. pruriens var. thekkadiensis, quantified by HPTLC-densitometry. L-dopa contents varied between 0.58 to 6.42 . High and low L-dopa yielding genotypes/chemotypes of M. pruriens could be multiplied for medicinal and nutritional purposes, respectively. HPTLC profiles of M. pruriens seeds on repeated extraction (24\u2009h) in 1:1 formic acid-alcohol followed by development in butanol:acetic acid:water  showed consistent degradation of L-dopa (Rf 0.34\u2009\u00b1\u20090.02) into a second peak (Rf 0.41\u2009\u00b1\u20090.02). An average of 52.11% degradation of L-dopa was found in seeds of M. pruriens varieties. Since M. pruriens seeds and/or L-dopa are used for treatment of Parkinson\u2019s disease and as an aphrodisiac both in modern and/or traditional systems of medicine, the finding of high level of L-dopa degradation (in pure form and in M. pruriens extracts) into damaging quinones and ROS is very significant. Mucuna pruriens (L.) DC. is a climbing legume distributed across the tropics. Four varieties of the species have been documented so far from south India, of which M. pruriens var. pruriens is well distributed1M. pruriens var. hirsuta and M. pruriens var. thekkadiensis are restricted to the southern parts of the Indian peninsula, and M. pruriens var. utilis occurs only in cultivationM. pruriens var. pruriens and Mucuna pruriens var. utilis find importance as food, feed, cover crop and fodder and are extensively cultivated worldwide345M. pruriens varieties propagate mostly through their seeds. M. pruriens var. pruriens is best known as the natural source of the aromatic amino acid, L-3,4-dihydroxy phenylalanine (levodopa or L-dopa)  . L-dopa L-dopa and dopamine, generating semiquinones, quinones, oxygen radicals and other reactive oxygen species (ROS), play a role in neuronal cell death in PD20O-quinone products of L-dopa autoxidation are cytotoxic to cellular systems222324M. pruriens var. pruriens and L-dopa could recover spermatogenic loss which makes them the treatment of choice for infertility27PD is characterized by degeneration of dopaminergic neurons in the substantia nigra, and subsequent deficiency of the neurotransmitter dopamine in the brain areas. PD affects motor activities including writing and speaking abilities. Recent studies suggested oxidative stress, mitochondrial dysfunction and impairment of the ubiquitin-proteasome system as the major factors involved in pathogenesis of PD7891011et al., 2011 and Sundaram and Gurumoorthi, 2012 standardized protocols for HPTLC-based quantification of L-dopa in M. pruriens var. pruriens seeds29et al., 2008 , Behara et al., 2010 (4.83%), Raina and Khatri, 2011 (2.23\u20135.36%) and Raina et al., 2012 (3.29\u20135.44%) quantified L-dopa contents in M. pruriens var. pruriens seeds by HPTLC313233et al., 2007et al., 2011et al., 2008et al., 2008et al., 2010M. pruriens var. pruriens seeds (7.20%) and its formulations (4.20\u20135.60%) by spectroflourimetryStizolobium pruriens var. utilis (M. pruriens var. utilis) (3.9\u201310.6%)M. pruriens var. utilis (4.39\u20135.21%)M. pruriens var. pruriens (4.0\u20136.0%)et al., 2012 quantified L-dopa in M. pruriens var. pruriens formulations (3.0\u20136.0%) by HPLCet al., 2010 developed HPLC-based quantification of L-dopa in M. pruriens var. utiliset al., 2010 carried out HPLC-MS/MS quantification of L-dopa in rat plasmaet al., 1997 determined threshold L-dopa levels in plasma of patients with advanced PD by in vitro microdialysis-HPLCVachhani 1 2.23\u20135.% and Raiutilis 4.\u20135.21%M. M. pruriens seeds and formulations by chromatographic techniques  are on limited number of samples and involved prolonged, multistep extraction procedures in acidic media. Screening of more M. pruriens varieties/accessions could lead to the discovery of high (elite) and low L-dopa yielding accessions suitable for medicinal and nutritional purposes, respectively. Secondly, in our preliminary HPTLC profiling of M. pruriens extracts and L-dopa standard, we repeatedly detected labile L-dopa-based degradation signals. Most similar HPTLC/HPLC studies never recorded this degradation signal of L-dopa562931323334353638394041M. pruriens varieties viz., M. pruriens var. pruriens (21), M. pruriens var. hirsuta (3), M. pruriens var. utilis (5) and M. pruriens var. thekkadiensis (1), collected from various locations in Kerala in south India, (ii) L-dopa contents in second generation seeds of twenty-six accessions of M. pruriens var. pruriens (21) and M. pruriens var. utilis (5) grown in an Experimental Plot (EP) under identical ecological conditions, (iii) degradation patterns of L-dopa in M. pruriens seed extracts, (iv) quantification of L-dopa degraded products in seeds  of four M. pruriens varieties and (v) characterization of L-dopa degraded moieties by HPTLC, DART-MS and LC/EI-MS.Most L-dopa quantification studies in M. pruriens var. pruriens accessions varied from 0.89 to 6.42   to 3.34%. L-dopa contents in wild accessions of M. pruriens var. hirsuta ranged from 1.01 to 4.27%, and % SDP ranged from 0.01 to 1.40%. L-dopa contents in seeds of M. pruriens var. utilis wild accessions varied from 0.65 to 2.67%, and SDP contents varied from 0.12 to 3.82%. In EP grown accessions of M. pruriens var. utilis, L-dopa contents ranged from 1.33 to 3.97%, and SDP contents were zero to 0.77%. M. pruriens var. thekkadiensis wild accession showed 4.34% of L-dopa with SDP 0.01% (M. pruriens var. hirsuta and M. pruriens var. thekkadiensis in the Field Gene Bank (FGB). This is the first report of L-dopa quantification in M. pruriens var. hirsuta and M. pruriens var. thekkadiensis. This is also the first quantitative determination of degradation products of L-dopa in M. pruriens seeds.L-dopa contents in wild dr. wt.) . Percentdr. wt.) . In secoDP 0.01% . FruitinM. pruriens var. pruriens, M. pruriens var. hirsuta and M. pruriens var. utilis seeds, L-dopa contents did not show any positive correlation with altitudes of their collection locations   accessions , M. pruriens var. thekkadiensis  and M. pruriens var. hirsuta . Lowest L-dopa percentages were found in EP grown M. pruriens var. pruriens seeds   showed lowest degradation levels . Similarly, lowest L-dopa contents  were seen in seeds with high levels of degradation  (In wild %/2.96%) . M. prurcessions . Highest, 0.84%) . Again, , 2.75%) .M. pruriens accessions was 52.11% of total L-dopa content (138.45%)  are highly significant. Standard L-dopa also showed similar degradation into a second signal on HPTLC profile . Previous studies rarely mentionedM. pruriens seeds and formulations2931323334353638394041Total SDP 72.15%) in wild 30) and EP grown 26) 138.45%) . These d and EP g% in wild 138.45%)f 0.34\u2009\u00b1\u20090.02 (f 0.34\u2009\u00b1\u20090.02 (L-dopa) and a SDP at Rf 0.41\u2009\u00b1\u20090.02  showed two significant signals  at varying ratios  M. pruriens var. pruriens seed extract and decomposed L-dopa standard showed degradation patterns on HPTLC, DART-MS and LC/EI-MS  showed M+H+ signals at 123.06 (medium), 154.10 , 162.07 (medium), 198.09 , 199.09 (minor) and 224.11 (minor)  showed M+H+ signals at 129.08 (medium), 176.08 (major), 190.10 (major), 191.11 (minor), 192.13 (minor), 193.13 , 195.17 , 204.11 (major), 218.13 (medium), 244.12 (medium) and 245.13 (minor)  on HPTLC showed only one signal at R4\u2009\u00b1\u20090.02 . But, L-1\u2009\u00b1\u20090.02 . M. prurg ratios . Fresh (LC/EI-MS . On DARTt 198.09 . Fresh M (minor) . Decompo (minor) .M. pruriens var. pruriens extract (24\u2009h extracted) showed only major signals of L-dopa and dopamine. Decomposed M. pruriens var. pruriens extract did not show L-dopa and dopamine, instead showed a group of degradation signals at the M+H+ 190.10, 191.11, 192.13, 193.13 and 195.17. These DART-MS signals correspond to dopachrome, leucodopachrome, dopaquinone, other quinones and ROS , 20\u2009mM Tris buffer  and water at various time periods  were tested through HPTLC profiling. Time dependent degradation was observed in M. pruriens var. pruriens extracts and in standard L-dopa at these pH values , M. pruriens var. utilis (10), M. pruriens var. hirsuta (3) and M. pruriens var. thekkadiensis (1) accessions varied between 0.58 to 6.42 . M. pruriens var. pruriens elite accessions are 4283  and 4498   and M. pruriens var. hirsuta  (M. pruriens var. pruriens (4098) showed highest L-dopa content of 4.32%  and M. pruriens var. utilis  accessions, which showed less than 1% L-dopa, can be multiplied and utilized as protein-rich diets.Our study led to the discovery of elite genotypes/chemotypes in 56 accessions of four %, wild) . Other e, 4.27%) . Among tof 4.32% . These eM. pruriens seeds in the acidic medium of 1:1 formic acid-alcohol. On HPTLC profiling, seed extracts of all four M. pruriens varieties (collected from various wild locations and EP grown) showed L-dopa at Rf 0.34\u2009\u00b1\u20090.02 and a consistent second degradation peak at Rf 0.41\u2009\u00b1\u20090.02. This degraded peak was indentified as a labile mix of dopamine, dopachrome, leucodopachrome, dopaquinone and other ROS by HPTLC, DART-MS and LC/EI-MS. On an average, the degradation of L-dopa in a 24\u2009h extraction protocol was a high 52.11%, which is significant enough to cause adverse effects in biological systems.We consistently found degradation patterns of L-dopa in M. pruriens var. pruriens seed extract (4450) and L-dopa standard in acidic, neutral and water media showed time dependent degradation. L-dopa degradation rates in the acidic medium  were equivalent or even higher compared to Tris buffer  or water under identical extraction periods. In seven days, M. pruriens var. pruriens seed extract and L-dopa in these liquid media resulted in degradation with gradual appearance of a black deposit. In 30 days, both M. pruriens var. pruriens seed extract and L-dopa in these solvent media resulted in significant degradation and strong black deposits. The degradation rates of L-dopa in M. pruriens var. pruriens extracts and standard L-dopa were low at 4\u2009\u00b0C compared to room temperature.HPTLC profiles of M. pruriens seeds or L-dopa are administered for very long periods. Chronic L-dopa therapy in PD results in movement disorders or dyskinesia in most patients. M. pruriens seeds and its preparations are also used for the treatment of PD in the traditional medicinal system of Ayurveda since ancient times. Certain studies claimed that M. pruriens seeds are even more effective than L-dopa in PD treatment. Some literature reports caution insufficient evidence to recommend the clinical use of M. pruriens in the treatment of PD45M. pruriens seeds are also used in tonics for male vitality and virility in Ayurveda28Recent studies showed that L-dopa degradation products and dopamine adducts result in oxidative stress and cause selective cytotoxicity of neuronal cells inducing pathogenesis in PD1820212223242543M. pruriens seeds and/or L-dopa are used for treatment of PD and as an aphrodisiac both in modern and/or traditional systems of medicine, the finding of high level of L-dopa degradation (in its pure form and in M. pruriens extracts) into damaging quinones and ROS is very significant. Our finding of consistent degradation products suggests the need for careful review of the processing of M. pruriens seeds , L-dopa and their mode(s) of administration . Further studies are required to confirm the adverse effects of the degradation products from the \u2018cure\u2019  itself in PD patients and other users.Since M. pruriens varieties viz. M. pruriens var. pruriens (21), M. pruriens var. hirsuta (3), M. pruriens var. utilis (5) and M. pruriens var. thekkadiensis (1) were collected in January to April 2009 from various wild locations in Kerala in south India (M. pruriens accessions were recorded during field trips. Voucher specimens of these M. pruriens accessions were deposited at the Herbarium (TBGT) of Jawaharlal Nehru Tropical Botanic Garden and Research Institute (JNTBGRI). Seeds of these thirty M. pruriens accessions collected were dried and powdered (separately).Seeds of thirty accessions of th India . GPS cooM. pruriens seeds collected were initially planted in a FGB of the species established at JNTBGRI. First generation seeds of 21 accessions of M. pruriens var. pruriens and 5 accessions of M. pruriens var. utilis were collected from the FGB and planted in the EP in Randomized Block Design with two replications of each accession and one plant in each replication. M. pruriens var. hirsuta and M. pruriens var. thekkadiensis accessions did not produce fruits in the FGB. Therefore, these two varieties were not planted in the EP. M. pruriens var. pruriens and M. pruriens var. utilis accessions were maintained in the EP in uniform conditions. M. pruriens accessions planted in both FGB and EP were irrigated as and when required, and not supplemented with any external fertilizers. Second generation seeds of 21 accessions of M. pruriens var. pruriens and 5 accessions of M. pruriens var. utilis were collected in January to April 2011, dried and powdered (separately) (A second set of arately) .M. pruriens seed powder (2\u2009g each) was extracted with 1:1 formic acid-alcohol  at room temperature and the extract was filtered. Seed powder residue was then repeatedly extracted with 1:1 formic acid-alcohol , and extracts were filtered. This seed residue was again extracted with 1:1 formic acid-alcohol . Filtrates (of five extractions) were pooled, centrifuged  and made up to 100\u2009ml using 1:1 formic acid-alcohol. This M. pruriens seed extract (5\u2009ml) was concentrated on a rotary evaporator, and the extract weight was recorded. This (concentrated) M. pruriens seed extract was dissolved in 20\u2009ml 1:1 formic acid-alcohol and used for L-dopa quantification by HPTLC-densitometry. This extraction protocol was followed for quantification of L-dopa in seeds of all (56) M. pruriens accessions  made up of Linomat V sample applicator, twin-trough plate development chamber, TLC Scanner 3 and WinCATS Software 4.03. M. pruriens seed extract (5\u2009ml concentrated seed extract) was dissolved in 20\u2009ml of 1:1 formic acid-alcohol (see L-dopa extraction), 4\u2009\u03bcl of this solution was repeatedly applied to silica gel HPTLC plate  as 6\u2009mm wide bands with Camag Linomat V sample applicator, fitted with a microsyringe, in N2 flow . L-dopa standard was also applied along with M. pruriens seed extracts. HPTLC plate was developed upto 80\u2009mm in the twin-trough glass chamber pre-saturated for 30\u2009min with mobile phase butanol:acetic acid:water . Developed plate was scanned densitometrically at 282\u2009nm (deuterium lamp) using TLC Scanner 3 equipped with WinCATS software. L-dopa at Rf 0.34\u2009\u00b1\u20090.02 (n\u2009=\u200956) and a second degradation peak (SDP) at Rf 0.41\u2009\u00b1\u20090.02 (n\u2009=\u200956) were found in M. pruriens seed extract in butanol:acetic acid:water   it showed a clear second signal at Rf 0.41  . Similar Rf 0.41 . Other s signals . Solventf values  of the standard was reproducible, and was found to be same as the values observed for the peak (L-dopa) in M. pruriens seed extracts. Calibration curve was plotted between amount of standard L-dopa (fresh) versus average response (peak area) . Linearity of the calibration curve in the range 100-1000\u2009ng was ensured. Percentage L-dopa content(s)  in M. pruriens extracts were calculated from peak areas using the standard curve. Percentage of second degradation peak  which is a combination of labile molecules was also quantified based on L-dopa standard curve. Repeatability of sample application  was assessed by applying a sample solution  on a HPTLC plate developed up to 80\u2009mm under saturation conditions with butanol:acetic acid:water  as the mobile phase in the twin-trough glass chamber (previously saturated with the solvent for 30\u2009min). The spot (L-dopa) was scanned six times, % coefficient of variation was acceptable. Robustness of the method was checked by slightly altering the mobile phase composition and plate developing distance was checked. No considerable effect on the data was found. Recovery studies were carried out (in two modes) by the addition of L-dopa to pre-analyzed M. pruriens extracts and they were again analyzed (see Quantification of L-dopa). In the first mode, (i) M. pruriens var. pruriens seed powder  was extracted in the five-step protocol (24\u2009h) and (ii) standard L-dopa (10\u2009mg) was added initially to M. pruriens var. pruriens seed powder  and extracted in the five-step protocol (24\u2009h). (i), (ii) and (iii) fresh standard L-dopa (1\u2009\u03bcg/\u03bcl), in 1:1 formic acid-alcohol were loaded (4\u2009\u03bcl each) onto HPTLC plate, developed with butanol:acetic acid:water  and peak areas were measured at 282\u2009nm . % recovery of L-dopa was calculated from peak areas as 49.78%. In the second mode, M. pruriens var. pruriens seed powder  was extracted in the standard five-step protocol (24\u2009h). (i) M. pruriens var. pruriens extract in 1:1 formic acid-alcohol (4\u2009\u03bcl), (ii) M. pruriens var. pruriens extract in 1:1 formic acid-alcohol (4\u2009\u03bcl) and fresh L-dopa dissolved in 1:1 formic acid-alcohol  and (iii) fresh L-dopa dissolved in 1:1 formic acid-alcohol  were loaded onto HPTLC plate ((ii) co-spotted), developed and peak areas were measured. % L-dopa recovery was calculated as 99.30%. % residual standard deviations (RSD) were determined as 2.63 (L-dopa) and 3.58 (SDP). Limit of detection  and limit of quantification  were determined for both L-dopa  and SDP 48HPTLC-based quantification of L-dopa was validated in terms of precision, accuracy, repeatability and linearity. Specificity of the assays was tested by repeated application of standard L-dopa. RM. pruriens var. pruriens seed extract (Acc. No. 4450) prepared by the five stage extraction for 24\u2009h (fresh M. pruriens var. pruriens seed extract) and standard L-dopa (fresh) were suspended in 1:1 formic acid-alcohol and kept at room temperature for seven days with occasional stirring. These resulted in decomposed M. pruriens var. pruriens seed extract and decomposed L-dopa standard, respectively. Fresh M. pruriens var. pruriens seed extract, freshly prepared L-dopa standard , decomposed M. pruriens var. pruriens seed extract and decomposed L-dopa standard were applied onto silica gel plates  by HPTLC , developed in butanol:acetic acid:water  and scanned at 282\u2009nm  (zerland) .M. pruriens var. pruriens seed extract (100\u2009mg), decomposed M. pruriens var. pruriens seed extract (100\u2009mg) and decomposed L-dopa standard (26.1\u2009mg) were analyzed by DART-MS on an AccuTOF JMS-T100LC Mass Spectrometer having a DART . Samples were analyzed directly in front of the DART source. Dry He was used at a flow rate of 4 LPM for ionization at 350\u2009\u00b0C. Orifice 1 was set at 28\u2009V, spectra were collected, and the data from 6-8 scans were averaged (M. pruriens var. pruriens seed extract (80\u2009mg) and decomposed L-dopa standard (20\u2009mg) were subjected to LC/ESI-MS analysis on a Surveyor-LCQ Deca XP plus system  with Hypersil BDS C18 column , mobile phase: methanol-water 85:15, inj. vol.: 10\u2009\u03bcl, flow rate: 0.3\u2009ml/min, run time: 30\u2009min, LC detection: PDA/UV detector, 280\u2009nm and mass detection: electro spray ionization  seeds (1\u2009g each) were separately extracted with 20\u2009ml (each) of 1:1 formic acid-alcohol (strongly acidic), 20\u2009mM Tris-HCl, 20\u2009mM KCl  and water at room temperature and at 4\u2009\u00b0C. Similarly, standard L-dopa (5\u2009mg) was extracted (dissolved) in 10\u2009ml each of these three solvents at room temperature and at 4\u2009\u00b0C. These M. pruriens var. pruriens seed/L-dopa extracts (3\u2009\u03bcl each) were profiled using HPTLC-densitometry (as described in Quantification of L-dopa) at various time periods viz., 1\u2009h after initiation of extraction, 1, 7 and 30 days after initiation of extraction ."}
+{"text": "The purpose of this study was to determine if consumption of a powdered form of tart cherries derived from tart cherry skins . The lifters ingested the supplements one time daily (480 mg/d) for 10-d: 7-d pre-exercise, day of exercise, and 48-hr post-exercise. Subjects performed 10 sets of 10 repetitions at 70% of 1RM back squat exercises with 3-min recovery between sets, maintaining equivalent total work performed (p = 0.80) and average daily caloric consumption (p = 0.61) between groups. Isokinetic knee extension/flexion maximal voluntary contractions (MVCs) and fasting blood samples were taken pre-squat workout, 60-min following squat workout as well as after 24-h and 48-h of recovery and analyzed by MANOVA with repeated measures.23 resistance trained men  were matched based on relative maximal back squat strength, age, body weight, and fat free mass. Subjects were randomly assigned to ingest in a double blind manner capsules containing a placebo  or powdered tart cherries [CherryPUREPowdered tart cherry supplementation seemed to attenuate the drop from pre-lift measures in 3-repetition summation of isokinetic flexion work , extension work , and all work  through 60-min and 24-h of recovery compared to placebo as reported above by Cohen's d effect size, despite not being statistically significant. The overall MANOVA analysis revealed a significant Wilks' Lambda time (p < 0.001) interaction for all inflammatory markers, but no significant group \u00d7 time pro-inflammatory (p = 0.30) and anti-inflammatory (p = 0.45) effects. Univariate measures for pro-inflammatory markers reported significant main time effects for TNF-\u03b1 (p = 0.001), IL-1\u03b2 (p = 0.30), IL-6 (p = 0.023), and IL-8 (p = 0.018). Univariate measures for anti-inflammatory markers reported significant main time effects for IL-4 (p = 0.001) and IL-7 (p = 0.033) with IL-13 trending toward significance (p = 0.055). No significant group \u00d7 time effects were observed for any of the inflammatory markers, NT, or TBARS. Serum IL-1\u03b2 levels were significantly lower in TC compared to P (p = 0.048). Delta changes were assessed at all three recovery time points from the pre-lift marker measures. The overall delta MANOVA analysis revealed a significant Wilks' Lambda pro-inflammatory interaction across time (p = 0.001) and an anti-inflammatory time interaction trending toward significance (p = 0.070), but no significant group \u00d7 time pro-inflammatory (p = 0.44) or anti-inflammatory (p = 0.30) effects. TNF-\u03b1 (p = 0.010), IFN-\u03b3 (p = 0.042), IL-1\u03b2 (p = 0.031), IL-6 (p = 0.001), IL-8 (p = 0.025), IL-10 (p = 0.019), and NT (p = 0.018) demonstrated significant main effects on time, while IL-7 approached significance across time (p = 0.095). Serum IL-2 TC (p = 0.074) and IL-10 (p = 0.10) changes from pre-lift tended to be greater across the recovery time coupled with a tendency for IL-10 (p = 0.063) TC levels to also be greater compared to P. Contrarily, serum IFN-\u03b3 (p = 0.021) TC changes from pre-lift values were significantly smaller compared to P with specific differences at 24-h and 48-h post-lift.In accordance with previous TC juice supplementation research, the isokinetic performance results of this study indicate that short-term powdered TC consumption 7 days prior to, day of, and 2 days after a single bout of intense resistance exercise may help to attenuate the strength decrement over a 48-h recovery period. The seemingly better maintenance of strength during recovery with short-term powdered TC supplementation surrounding a single bout of resistance exercise did not, however, coincide with any definitive effect on markers of oxidative damage or inflammation. This may be due to the differences in resistance exercise metabolic demands, thus indicating the need for further mechanistic research."}
+{"text": "AbstractHoplolaimus Daday, 1905 belongs to the subfamily Hoplolaimine Filipiev, 1934 of family Hoplolaimidae Filipiev, 1934  with ten species, is characterized by lateral field distinct, with four incisures, excretory pore behind hemizonid; Hoplolaimus (Basirolaimus) with 18 species, is characterized by lateral field with one to three incisures, obliterated, excretory pore anterior to hemizonid, dorsal oesophageal gland quadrinucleate; and Hoplolaimus (Ethiolaimus) with four species is characterized by lateral field with one to three incisures, obliterated; excretory pore anterior to hemizonid, dorsal oesophageal gland uninucleate (Hoplolaimuspuriensis Ali, Shaheen & Pervez, 2009 has been described (Hoplolaimus reported in Vietnam, viz H.seinhorsti and H.chambus (The genus ev, 1934 . Daday e in 1905 . Hoplolaal crops \u200b. In 199ay, 1905 . Siddiqinucleate . Since tescribed . Up to nchambus .Hoplolaimusbachlongviensis sp. n. was isolated from banana soil in Bach Long Vi Island, Vietnam. The female of this species is described and illustrated below. Some diagnostic characters of this species include body slightly curved ventrally, offset lip region exhibiting three to four annules, lateral field reduced, pharyngeal glands with six nuclei, excretory pore anterior to hemizonid, epiptygma absent, intestine not overlapping rectum and male was not found. Hoplolaimusseinhorsti Luc, 1958 and H.chambus Jairajpuri & Baqri, 1973 were recorded . Soil nematodes were extracted using the decanting and modified Baermann tray method ; IEBR.Nema4050-2 .Hoplolaimusbachlongviensis sp. n. is similar to Hoplolaimusseinhorsti, H.chambus, H.columbus Sher, 1963 and H.pararobustus  Sher, 1963 by having excretory pore anterior to hemizonid, lateral field reduced, represented by interruptions of annules as a single incisure, often indistinct, pharyngeal glands with six nuclei (H.bachlongviensis sp. n. differs from H.seinhorsti by epiptygma absent vs present and number of longitudinal striations on basal ring 6 vs 8-12. It differs from H.chambus by male absent vs present, epiptygma absent vs present, intestine not overlapping rectum vs overlapping rectum. Hoplolaimusbachlongviensis sp. n. differs from H.columbus in having fewer tail annuli 9-13 vs 16-22; a=22-27 vs a=30-38; b=6-8 vs b=9.1-12.4; DGO=3-6 vs DGO=9-13; epiptygma absent vs present; hemizonid located about 7 annuli behind excretory pore vs 2-5 annuli and intestine not overlapping rectum vs overlapping rectum. Hoplolaimusbachlongviensis sp. n. differs from H.pararobustus by male absent vs present, intestine not overlapping rectum vs overlapping rectum, epiptygma absent vs present and sperm absent vs present.x nuclei . HoweverHoplolaimusbachlongviensis sp. n. is distinguished from H.sherivs two incisures in lateral field; having longer stylet 44-50 vs 40-45; having fewer longitudinal striations on basal ring 6 vs 20; hemizonid is conspicuous vs obscure; a=22-27 vs a=26-30; b=6-8 vs b=9.7-11.5.Hoplolaimusbachlongviensis sp. n. differs from H.puriensis by lateral field reduced, represented by a single incisure on the body, but often indistinct vs four lateral lines, longer stylet 44-50 \u00b5m vs shorter stylet 32-35 \u00b5m."}
+{"text": "Enterobacter cloacae strain S1:CND1 isolated from oil-contaminated soil in Guwahati, India. S1:CND1 contains 4,205 coding sequences and has a G+C content of 57.45%. This is the first report of the genome sequence of an E.\u00a0cloacae adapted to an oil-contaminated environment.We report here the 4.57-Mb draft genome sequence of hydrocarbon-degrading  Enterobacter cloacae is a Gram-negative, rod-shaped bacterium generally associated with nosocomial infections, and is considered one of the most difficult to treat among the Enterobacter sp. , along with 16 rRNAs, 77 tRNAs, 6 noncoding RNAs (ncRNAs), and 50 pseudogenes. Rapid functional annotation for CDSs of strain S1:CND1 was carried out with the RAST annotation server (s = 650 CDSs); amino acids and derivatives (s = 467); stress response (s = 157); respiration (s = 149); fatty acids, lipids, and isoprenoids (s = 138); DNA metabolism (s = 117); regulation and cell signaling (s = 150); protein metabolism (s = 168); RNA metabolism (s = 150); membrane transport (s = 177); virulence, disease, and defense (s = 100); cell wall and capsule (s = 196); and cofactors, vitamins, prosthetic groups, and pigments (s = 253). Genome annotation revealed the presence of hydrocarbon degradation genes as alkane-1-monooxygenase, alkanesufonate monooxygenase, naphthalene 1,2-dioxygenase, and quercetin 2,3-dioxygenase, thus underlining the extensive genetic adaptation of strain S1:CND1 to oil contamination.The genomic DNA for strain S1:CND1 was extracted using an Ultra-Clean Microbial DNA Isolation Kit  according to the manufacturer\u2019s protocol. Isolated genomic DNA was then sequenced with an Illumina HiSeq 2500, which generated 4,580,054 paired-end reads. After quality control measures, the reads were assembled using the  v. 3.81 , SPAdes  v. 3.81 , and Vel v. 3.81 , which gEscherichia coli 88.1467 (score = 501) as its closest neighbor, followed by E.\u00a0coli 88.0221 (score = 490) and E.\u00a0coli 89.0511 (score = 469). Enterobacter mori LMG 25706 (score = 344) was identified as the 18th-closest neighbor.A comparison of strain S1:CND1 with genomes in the RAST database identified E.\u00a0cloacae strain S1:CND1 has been deposited in DDBJ/EMBL/GenBank under the accession no. LUGN00000000. The version of the whole-genome sequence (WGS) described here is version LUGN01000000.This whole-genome shotgun sequencing project for"}
+{"text": "Thus, AvCystatin ameliorates enhanced RSV pathology without increasing susceptibility to, or persistence of, viral infection and warrants further investigation as a possible therapy for virus-induced airway disease.Respiratory Syncytial Virus (RSV) is a major pathogen causing low respiratory tract disease (bronchiolitis), primarily in infants. Helminthic infections may alter host immune responses to both helminths and to unrelated immune triggers. For example, we have previously shown that filarial cystatin (AvCystatin/Av17) ameliorates allergic airway inflammation. However, helminthic immunomodulators have so far not been tested in virus-induced disease. We now report that AvCystatin prevents Th2-based immunopathology in vaccine-enhanced RSV lung inflammation, a murine model for bronchiolitis. AvCystatin ablated eosinophil influx, reducing both weight loss and neutrophil recruitment without impairing anti-viral immune responses. AvCystatin also protected mice from excessive inflammation following primary RSV infection, significantly reducing neutrophil influx and cytokine production in the airways. Interestingly, we found that AvCystatin induced an influx of CD4 However, due to the proposed antiviral function of neutrophils, current practice suggests to target neutrophils under the umbrella of antiviral treatment .+ T cell derived IL-10. Collectively, our findings support the testing of AvCystatin as a potential novel agent to counter virus-enhanced respiratory inflammation.This is, to our knowledge, the first time that a recombinant parasite-derived immunomodulator has been shown to down-regulate virally-enhanced inflammation without impairing responses important for anti-viral immunity. We provide evidence that AvCystatin immunomodulation may act through the production of CD4All mouse experiments were ethically approved by the Imperial College Central Biological Services (CBS) ethics committee performed in accordance with approved UK Home Office guidelines (Project License No. PPL 70/6785).Mice were treated with 20 \u03bcg AvCystatin prepared as previously described  or PBS i5 PFU RSV 2 weeks thereafter. For rvv-G RSV models, mice were primed with vaccinia expressing RSV-G protein (3 x 106 PFU) by scarification of the rump and infected intranasally with 5 x 105 PFU RSV 14 days later. RSV titres were assessed by titration of lung homogenates on Hep-2 cell monolayers [RSV (strain A2) and rVV-G (vaccinia virus expressing the RSV G-protein) were propagated in Hep-2 cells (ATCC) as described previously . FI RSV nolayers . RSV L-gnolayers .BAL-accessible cells were obtained by repeated instillation of 1 ml of PBS/12 mM lidocaine via the trachea in sacrificed animals. Cytospin preparations were stained with hematoxylin and eosin and analysed by light microscopy (300 cells per slide). Lungs were homogenized and digested with collagenase D  for 30 min. Mediastinal  lymph nodes were dissociated and filtered through a 100 \u03bcm cell strainer.Flow cytometry was performed as previously described . Briefly6 PEC and infected with RSV i.n. 4 h later.Donor BALB/c were injected i.p. with 20 \u03bcg active or heat-inactivated AvCystatin and peritoneal exudate cells (PEC) recovered 20 h later in PBS/2 mM EDTA/0.2% BSA. Recipient mice were injected intravenously with 3x10Quantification of BAL chemokines and cytokines was performed using a 13-plex Luminex kit . Data were acquired with a Luminex 100  and Starstation software.P values of <0.05 were considered significant.GraphPad Prism software  was used to analyse data, generally shown as mean \u00b1SEM of 5 animals per group. Mann-Whitney t-test and 2 way ANOVA were used to compare data. S1 FigP values reflect Mann-Whitney t-test: * p<0.05, **p<0.01.A) Schematic of the FI RSV model: i.m. intramuscular; i.p. intraperitoneal; i.n. intranasal application. B) Weight loss in the FI-RSV model. Total cell number of eosinophil (C) and macrophages in the BAL (D). Viral load in the lungs measured by RSV L-gene copies (E). Representative data of 2 experiments, 5 mice per group. Error bars indicate SEM. (TIF)Click here for additional data file.S2 Fig+ T cells in the BAL (D and E) and lungs (B and C). Representative data of 2 experiments, 5 mice per group. Error bars indicate SEM. P values reflect Mann-Whitney t-test: * p<0.05, **p<0.01.Flowcytometric analysis of IL-10 intracellular cytokine content in RSV challenged or AvCystatin/RSV challenged mice is shown (A). Graphical visualization of IFN\u03b3 and IL-10 production by CD4(TIF)Click here for additional data file.S3 Fig+ CD4+ T cells (A) and the number of IL-10+ CD4+ T cells in the mLN (B) after AvCystatin treatment and RSV challenge. Representative data of 2 experiments, 5 mice per group. Error bars indicate SEM. P values reflect Mann-Whitney t-test: * p<0.05, **p<0.01.Total number of FoxP3(TIF)Click here for additional data file.S4 FigP values reflect Mann-Whitney t-test: * p<0.05.Relative expression of MUC5a in mice lungs after the vvG RSV model (A) or primary RSV model (B). Error bars indicate SEM. (TIF)Click here for additional data file."}
+{"text": "Ocimum sanctum) were explored and investigated for their diversity and antiphytopathogenic activity against widespread plant pathogens Botrytis cinerea, Sclerotinia sclerotiorum, Rhizoctonia solani and Fusarium oxysporum. 90 fungal isolates, representing 17 genera were recovered from 313 disease-free and surface sterilised plant segments (leaf and stem tissues) from three different geographic locations  during distinct sampling times in consequent years 2010 and 2011 in India. Fungal endophytes were subjected to molecular identification based on rDNA ITS sequence analysis. Plant pathogens such as F. verticillioides, B. maydis, C. coarctatum, R. bataticola, Hypoxylon sp., Diaporthe phaseolorum, Alternaria tenuissima and A. alternata have occurred as endophyte only during second sampling (second sampling in 2011) in the present study. Bi-plot generated by principal component analysis suggested tissue specificity of certain fungal endophytes. Dendrogram revealed species abundance as a function of mean temperature of the location at the time of sampling. Shannon diversity in the first collection is highest in Hyderabad leaf tissues (H' = 1.907) whereas in second collection it was highest from leaf tissues of Delhi (H' = 1.846). Mukteshwar  reported least isolation rate in second collection. Nearly 23% of the total fungal isolates were considered as potent biocontrol agent. Hexane extract of M. phaseolina recovered from Hyderabad in first collection demonstrated highest activity against S. sclerotiorum with IC50 value of 0.38 mg/ml. Additionally, its components 2H-pyran-2-one, 5,6-dihydro-6-pentyl and palmitic acid, methyl ester as reported by GC-MS Chromatogram upon evaluation for their antiphytopathogenic activity exhibited IC50 value of 1.002 and 0.662 against respectively S. sclerotiorum indicating their significant role in antiphytopathogenic activity of hexane extract. The production of 2H-pyran-2-one, 5,6-dihydro-6-pentyl from M. phaseolina, an endophytic fungus is being reported for the first time.Endophytic mycopopulation isolated from India\u2019s Queen of herbs Tulsi ( Endophytic fungi inhabit a unique biological niche and are categorized as highly diverse, polyphyletic group of primarily ascomycetous fungi, capable of colonizing tissues of plants asymptomatically without initiating any disease or overt negative symptoms , 2], , , 5]. 42. 425]. 4Ocimum sanctum is ubiquitous in Indian culture and tradition, called Tulsi in Hindi, . F. FO. sanThe GC-MS analysis of hexane extracts was carried on GC-MS (Agilent Technologies 7890A). For this 1mg of extract was dissolved 1 ml LR-grade dichloromethane . DB-WAX column (30 m \u00d7 250\u03bcm \u00d7 0.25 \u03bcm) was used with helium as a carrier gas at a flow rate of 1 ml/min at pressure of 11.654 psi. The GC oven temperature was kept at 120\u00b0C for 2 min and programmed to 5\u00b0C/min to 130\u00b0C for 1 min; 5\u00b0C/min to 150\u00b0C for 2 min; 2\u00b0C/min to 180\u00b0C for 3 min; 2\u00b0C/min to 200\u00b0C for 3 min; 5\u00b0C/min to 240\u00b0C for 20 min. Splitless injections were done with liquid injection method in this study. A library search was carried out using NIST library.2 placed in such a way that seven are put in periphery and one in the centre of the plate. The control experiment consisted of only methanol solvent in PDA medium , , , 39]..39].The similarity and of endophytic fungal assemblages among both tissues was compared using the following similarity indices:(QS), QS = 2a/(2a + b + c) where \u2018a\u2019 is the number of common species in both communities, while \u2018b\u2019 and \u2018c\u2019 are the number of species specified to each community under investigation, respectively .JS was calculated using the formula: JS = a/(a + b + c) where \u2018a\u2019 is the number of common species in both communities, while \u2018b\u2019 and \u2018c\u2019 are the number of species specified to each community, respectively .O. sanctum different plants parts in distinct sampling time, whilst cluster analysis deduced concurrence between species richness and temperature of geographical locations at the time of sampling. For this, software Unsrcambler X: version 10, CAMO, USA was used . M. Min vitPenicillium sp.), OSHL-5.2 (Alternaria sp.), OSHL-2.1 (M. phaseolina), OSHS-3.1(M. phaseolina), OSML-5.4 (Colletotrichum sp.), OSMS-2.2 (Aspergillus niger), OSMS-2.3 (Meyerozyma guilliermondii), OSDSL-9.8 (Fusarium proliferatum), OSDSS-2.5 (Chaetomium coarctatum), OSDSL-5.6 , OSHSS-5.2 (Sympodiomyces sp.), OSHSS-1.3 (Fusarium proliferatum) OSHSS-2.3 (Diaporthe phaseolorum) endophytic fungal isolates were multiplied on rice medium, and their crude ethylacetate extracts were subjected to intoxicated/poisoned food bioassay to investigate antiphytopathogenic activity of crude fungal extracts against all four phytopathogens. IC50 values of ethylacetate crude extracts of selected thirteen endophytic fungal isolates at five different concentrations was calculated by regression analysis are summarised as Chaetomium coarctatum recorded the highest activity against all phytopathogens with IC50 value of < 1mg/ml ranging from 0.262mg/ml\u2013 0.553 mg/ml. Second best activity was reported with Fusarium proliferatum obtained from Hyderabad with IC50 value ranging from 0.299 to 1.04 mg/ml.Based on the results of dual culture bioassay , followiFusarium proliferatum obtained from Hyderabad exhibited IC50 value of 0.51mg/ml against F.oxysporum which was stronger than another isolate of F. proliferatum recovered from Delhi in second sampling , OSHS-3.1 (M.phaseolina), OSDSS-2.5 (Chaetomium coarctatum), OSHL-5.2 (Alternaria sp.), OSML-5.4 (Colletotrichum sp.) and OSHSS-1.3 (F. proliferatum) were found positive for terpenoids in preliminary phytochemical screening. GC-MS chromatography of hexane extracts of these selected endophytic fungal isolates revealed presence of volatile compounds, fatty acids, and aliphatic constituents. GC-MS chromatogram of M. phaseolina (OSHL-2.1) showed presence of 2H-Pyran-2-one, 5, 6-dihydro-6-pentyl at RT 30.156, hexadecanoic acid at RT 53.017, linoleic acid at RT 64.986 (M. phaseolina (OSHS-3.1) showed presence 9,12-octadecadieonic acid, methyl ester at RT 61.108, trifluoroacetoxy hexadecane at RT 55.970 as major peaks reflecting strain variability (Chaetomium coarctatum (OSDSS-2.5) 2-fluorobenzoic acid at RT 38.644, 2,5-difluorobenzoic acid at RT 38.444, linoleic acid, ethyl ester at RT 64.900 were observed as major peaks (Alternaria sp. (OSHL-5.2) hexane extract, hexadecanoic acid at RT 52.990, 9,15- octadecadienoic acid, methyl ester at RT 61.036 were recorded as major peaks (Fusarium proliferatum (OSHSS-1.3)-major peaks were hexadecanoic acid at RT 52.992, linoleic acid at RT 64.902, oleic acid at RT 65.954 (Colletotrichum sp. (OSML-5.4) major peaks recorded were 7-hydroxy3- coumarin at RT 61.372, 9, 12-octadecadieonic acid at RT 61.052 . Rhizopus oryzae has been reportedly primary causal agent of tuber rot disease in sweetpotato in India and in this study has been isolated from Delhi and Hyderabad in first sampling season . I. IO. sanic fungi , 51], , , , and 7272.Additionally, cluster analysis inferred distribution and abundance of endophytic fungal species from different geographical locations as a function of temperature. It grouped together species richness of Delhi (second sampling) and Hyderabad (first sampling) when temperature ranged from 23\u201324\u00b0C into cluster 1, cluster 2 comprises of Delhi (first sampling) and Hyderabad (second sampling) when temperature ranged from 30\u201331\u00b0C and cluster 3 belongs to endophytic mycobiota of Mukteshwar where temperature range was 14\u201317\u00b0C during both sampling times. This revelation is in accordance with previous studies elucidating higher influence of temperature, precipitation and other climatic factors than geography on endophytic communities of plants , 74]..74].S. Sclerotiorum causes heavy yield losses up to the tune of 40% in Brassicas in India , ,  all the in India . Fusariuin India . Botryti disease . R. solahe world . Differeironment .M. phaseolina (OSHL-2.1) exhibited effective inhibitory activity against B. cinerea, F. oxysporum and R. solani with 4.504 mg/ml, 1.97mg/ml and 1.41mg/ml IC50 values respectively. Whereas hexane extract of M. phaseolina (OSHL-2.1) demonstrated highest activity against S. sclerotiorum with IC50 value of 0.38 mg/ml followed by F. oxysporum with IC50 value of 0.867mg/ml. Preliminary identification of bioactive metabolites were conducted on crude hexane extracts.GC-MS chromatogram of hexane extract of M. phaseolina illustrated presence of 2H-pyran-2-one, 5, 6-dihydro-6-pentyl at RT 30.152 and palmitic acid, methyl ester at RT 53.017, while that of C. coarctatum (OSDSL-2.5) illustrated Oleic acid at RT 61.675. Present study for the time reports antifungal activity of extracts of M. phaseolina and C. coarctatum. Antifungal activities of above mentioned pure compounds against plant pathogens resulted in remarkable revelation. Oleic acid methyl ester, palmitic acid methyl ester and 5, 6 Dihydro-2H-pyran-2-one-6 pentyl exhibited strong inhibitory activity against S. sclerotiorum with 0.547mg/ml, 0.662mg/ml and 1.002 mg/ml IC50 value while against F. oxysporum it was 0.27 mg/ml, 0.264 mg/ml and 0.27mg/ml respectively. Thereby, buttressing the fact that 2H-pyran-2-one, 5,6-dihydro-6-pentyl and palmitic acid, methyl ester are bioactive metabolites synthesised by endophytic fungus M. phaseolina which are responsible for its antiphytopathogenic activity against S. sclerotiorum. Antifungal activity of 6-Pentyl-2H-pyran-2-one and its analogs has been well documented in literature . R. RM. phalium spp . Anotheric fungi . The pro studies . Hexadecalbicans . The majNCBT 196 . Another. solani .Bipolaris maydis, Rhizoctonia bataticola and Chaetomium coarctatum in endophytic state harbored inside O. sanctum for the first time. In present study metabolites produced by M. phaseolina recovered from Hyderabad in first collection has exhibited promising antifungal activity against S. sclerotiorum and F. oxysporum broad spectrum phytopathogens. Metabolites originated from fungal endophytes hold promise to be further developed as greener and safer biocontrol agent in crop disease management. It can be concluded that fungal endophytes harbored inside leaf and stem tissues of Ocimum sanctum collected from different geographical locations in different sampling times hold great promise not only as biocontrol agents against broad spectrum and economically significant phytopathogens, but also as sustainable resource of novel antifungal secondary metabolites.The study reports noted plant pathogens such as S1 Fig(TIFF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file."}
+{"text": "Cancer patients are at risk for severe complications related to the underlying malignancy or its treatment, and therefore usually require admission to ICUs.To evaluate the clinical characteristics and outcomes in this subgroup of patients.Analysis of two prospective cohorts of cancer patients admitted to ICUs. We used multivariable logistic regression to identify variables associated with hospital mortality.P <0.001 for all analyses). ICU (47 % vs. 27 %) and hospital (63 % vs. 38 %) mortality rates were also higher in patients with complications (P <0.001). Chemo/radiation therapy-induced toxicity (6 %), venous thromboembolism (5 %), respiratory failure (4 %), gastrointestinal involvement (3 %) and vena cava syndrome (VCS) (2 %) were the most frequent complications. In multivariable analysis, the presence of cancer-related complications per se was not associated with mortality (odds ratio (OR) = 1.25 ). However, among the individual complications, VCS (OR = 3.79 (1.11-12.92)), gastrointestinal involvement (OR = 3.05 (1.57-5.91)) and respiratory failure (OR = 1.96 (1.04-3.71)) were independently associated with worse outcomes.Out of 2028 patients, 456 (23 %) had cancer-related complications. Compared with those without complications, they more frequently had worse performance status (PS) (57 % vs. 36 % with PS \u22652), active malignancy (43 % vs. 5 %), need for vasopressors (45 % vs. 34 %), mechanical ventilation (70 % vs. 51 %) and dialysis (12 % vs. 8 %) (The prognostic impact of cancer-related complications was variable. Although some complications were associated with worse outcomes, the presence of a severe acute cancer-related complication per se should not guide decisions to admit a patient to the ICU."}
+{"text": "Apelin (APLN) is a powerful inotrope, is widely expressed by the cardiovascular system with its receptor APJ-R, and should be considered as an alternative noncatecholaminergic support.Dobutamine (DOB) is the actual recommended b-adrenergic inotropic drug to support sepsis-induced myocardial dysfunction when cardiac output index is still low after preload correction. In this context, DOB cardiovascular response predicts outcome in septic shock. Alternative supportive and safer therapies are however mandatory because:Perform a comparative evaluation of APLN-13 (APLN active peptide) vs DOB in terms of hemodynamic efficacy, cardioprotection and outcome in a model of \u201csepsis-induced cardiac dysfunction\u201d.E. Coli 055:B5, 10-12mg/kg intraperitoneal).A rat model of LPS-induced myocardial dysfunction .0-6-18h.in vivo: echocardiography, final hemodynamics; urine output, weight and plasma volume variation, survival study, ex vivo: Langendorff dP/dt, in vitro: APJ-R and beta1-adrenergic receptor (AR) myocardial expressions, Pi3K/Akt/GSK3/mTOR activation-expression profiles, cTnI  and cleaved caspase-3 (apoptosis).Both drugs restored LPS-induced fall of left ventricular ejection fraction (LVEF) with dominant chronotropic impact and mean arterial pressure (MAP) restoration for DOB, and lower peripheral vascular resistances (PVR) for APLN. APJ-R but not beta1 AR myocardial expressions were upregulated by LPS challenge (p< 0.05). The deepness the induced LVEF drop, the higher the dP/dt response to APLN but not to DOB (p< 0.05). In 18h LPS-challenged hearts, Langendorff assays peak dP/dt responses were 3nM and 100nM for APLN and DOB, respectively (p< 0.05). Combining fluid resuscitation with APLN infusion declined urine output (UO) with plasma volume (PV) expansion, whereas DOB induced less PV expansion but more UO (p< 0.05). Survival proportions were clearly distinctive (p< 0.05).APLN further dampened down LPS-induced overphosphorylation/inhibition of Pi3K/Akt/mTOR/GSK3 and reduced injury/apoptosis (i.e. cTNI and cleaved caspase-3 expressions).APLN potentially offers distinctive mechanisms of hemodynamics, cardioprotective effects, and survival benefits, over DOB. Chemical optimization of APLN-13 with more extensive preclinical data, would pave the way for first phase clinical trials."}
+{"text": "Due to increasing resistance, alternatives to mupirocin and chlorhexidine for decolonization of MRSA carriage need to be evaluated.\u00ae) vs placebo in eliminating MRSA carriage at day 28 (D28) after the end of treatment.To evaluate the efficacy of polyhexanide  or placebo  applied to the anterior nares and skin for 10 days. The primary outcome was MRSA decolonization at D28 assessed by both intention-to-treat  and per-protocol (PP) analysis . Secondary outcomes included MRSA decolonization according to nasal MRSA carriage, safety and emergence of resistance.vs 22/75 (29.3%) in group P were MRSA-free at D28 . PP analysis confirmed the results with 19/53 (35.8%) decolonized polyhexanide-treated patients vs 17/56 (30.4%) in the placebo arm . In the subgroup of MRSA nasal carriers, PP analysis showed that 6/15 (40.0%) patients in group I vs 2/11 (18.2%) in group P were decolonized (P=0.40). Nine serious adverse events occurred in group I vs 12 in group P; none was attributable to study medication. Emergence of polyhexanide resistance was not observed.Of 2590 patients screened, 146 patients  were randomized between January 2011 and July 2014. Primary outcome was missing for 11 (7.5%) patients. ITT analysis showed that 24/71 (33.8%) patients in group I This study suggests that under real-life conditions a single polyhexanide decolonization course is marginally effective in eradicating MRSA carriage.C. Landelle: None declared, E. Von Dach: None declared, T. Haustein: None declared, A. Agostinho: None declared, G. Renzi: None declared, A. Renzoni: None declared, D. Pittet: None declared, J. Schrenzel: None declared, P. Fran\u00e7ois: None declared, S. Harbarth Grant/Research support from: a peer-reviewed research grant funded by Pfizer, Consultant for: the advisory boards of Destiny Pharma, bioMerieux, Novartis, and DaVolterra"}
+{"text": "COBI, a PK enhancer with no ARV activity is a more selective cytochrome P450 (CYP)3A inhibitor than ritonavir (RTV), does not induce CYP isozymes, and thus has less potential for drug-drug interactions. COBI boosts DRV PK as effectively as RTV in healthy volunteers.This 48-week, phase IIIb, open-label, single-arm, US multicentre study (NCT01440569) included HIV-infected treatment-nave and experienced adults with no DRV RAMs, viral load (VL) \u22651000 c/mL, eGFR \u226580 mL/min and genotypic sensitivity to investigator-selected N[t]RTIs. Patients received DRV/COBI 800/150 mg qd (as single agents) plus two fully active N[t]RTIs. The primary endpoint was any treatment-emergent grade 3 or 4 AEs through Week 24. We report 48-week safety, efficacy and PK/PD results in treatment-nave patients.10 c/mL and CD4+ 370 cells/mm3. Treatment-emergent grade 3 or 4 AEs regardless of causality were reported in 21 (7%) patients. AEs regardless of causality  were: diarrhoea (27%), nausea (23%), URTI (15%) and headache (12%). Sixteen (5%) patients had AEs leading to study drug discontinuation, most frequently rash (three patients), hypersensitivity and nausea (two patients each). Consistent with the known inhibition of tubular creatinine secretion by COBI, there was a mean increase from BL in serum creatinine by week 2 (0.09 mg/dL), remaining stable through week 48 (mean 0.10 mg/dL increase from BL). At week 48, 83% of patients achieved VL<50 c/mL; FDA Snapshot); median increase in CD4+ was 169 cells/mm3. Eight patients met the criteria for resistance testing. M184V was detected in one pt receiving FTC. New primary RAMs were not detected in the other seven patients. The mean population PK-derived DRV AUC24h was 100,620 ng.h/mL and C0h 2,105 ng/mL (n=281). There were no clinically relevant relationships between DRV exposure and virologic response, AEs or laboratory parameters.Of 313 ITT patients, 295 were treatment-nave (94%). In the treatment-nave cohort, 90% were male, 60% white and 294 (99.7%) received a TDF-containing regimen. Median baseline (BL) VL was 4.8 logThe DRV PK of DRV/COBI was consistent with historical data for DRV/RTV. DRV/COBI 800/150 mg qd plus two N(t)RTIs had an 83% response and was well tolerated through Week 48. These results are similar to published data for DRV/RTV 800/100 mg qd, and support the use of DRV/COBI 800/150 mg qd in treatment-nave patients."}
+{"text": "Methylobacter luteus, Methylobacter whittenburyi, Methylosarcina fibrata, Methylomicrobium agile, and Methylovulum miyakonense were generated. The strains represent aerobic methanotrophs typically isolated from various terrestrial ecosystems.Genome sequences of  Methylobacter whittenburyi (formerly \u201cMethylobacter capsulatus\u201d = UCM-B-3033), and Methylomicrobium agile (ATCC 35068) are methanotrophic bacteria commonly found in sediment samples from wetlands  have typically been obtained from meadows, dry hay, and cow mouth samples (\u2013Methylovulum miyakonense HT12T (= ATCC BAA-2070) was isolated from a forest soil (Methylosarcina fibrata AML-C10T (= ATCC 700909) was isolated from a landfill site , using the Illumina  and/or P17\u2013Prodigal  and GeneProdigal . AdditioProdigal  and MaGeProdigal  platformM. miyakonense HT12T genome (pxmABC)  or urea (urtABCDE/ureABCDEFG) as the sole source of nitrogen. M. miyakonense HT12T, M. luteus 98, and M. whittenburyi UCM-B-3033 possess the key genetic elements for nitrogen fixation (nifKDHWENX).Genome statistics and predicted core metabolic pathways are shown in T genome . A funct(pxmABC)  was founrogenase . Two typ protein . M. luteeductase  were ideMethylobacter spp.) produce cysts (M.\u00a0luteus 98 (Many methanotrophic species (including ce cysts . We wereuteus 98 , 30. Twouteus 98  were ideThe genome sequences have been deposited in GenBank under the accession numbers listed in"}
+{"text": "Simian hemorrhagic fever virus (SHFV) variant NIH LVR42-0/M6941 is the only remaining SHFV in culture, and only a single genome sequence record exists in GenBank/RefSeq. We compared the genomic sequence of NIH LVR42-0/M6941 acquired from the ATCC in 2011 to NIH LVR42-0/M6941 genomes sequenced directly from nonhuman primates experimentally infected in 1989. Arterivirus , causes viral hemorrhagic fever (VHF) in captive Asian macaques (reviewed in reference Macaca mulatta) inoculated with a whole blood suspension from a deceased SHFV-infected stump-tailed macaque (Macaca arctoides) (Simian hemorrhagic fever virus (SHFV), a member of the genus ctoides) . Resultsctoides) . Anotherctoides) .Chlorocebus aethiops; A095 and A230) and one rhesus monkey (I-618) on different days (d) postinoculation . Briefly, RNA was isolated with the omission of carrier RNA , and randomly primed double-stranded cDNA was synthesized , as previously described  by Vatter et al. . FurtherKM373784, KM371111, KM371105, KM371106, KM371107, KM371108, KM371109, and KM371110, respectively.The GenBank accession numbers of SHFV variant NIH LVR42-0/M6941 isolates KS_06_17_11, RJ_03_26_10, A095-d5, A095-d7, A230_d5, A230_d7, I618_d3, and I-618_d5 are"}
+{"text": "Migraine remains poorly treated with few effective preventive medications available.We evaluated the efficacy and safety of LY2951742, a fully humanized monoclonal antibody to Calcitonin Gene-Related Peptide (CGRP) for migraine prevention.Eligible subjects with 4-14 migraine headache days (MHD) per month were randomized in a double-blind manner to LY2951742 or placebo, administered subcutaneously, every other week over a 12-week period. The primary endpoint was the mean change from baseline in number of MHD in the last 28 day period (month three); secondary end points included the mean change in headache days, migraine attacks, and the 50% responder rate at month 3.Mean change in MHD (primary) was -4.2 vs. -3.0 for LY2951742 (n=107) and placebo (n=110), respectively (p = 0.003). LY2951742 was superior to placebo for all secondary endpoints including headache days -4.9 vs. -3.7 (p = 0.0117), migraine attacks -3.1 vs. -2.3 (p = 0.0051), and 50% responder rate, 70% vs. 45% . An exploratory endpoint of complete response (100% reduction in MHD in month 3) was 31.6% vs. 17.3%  for LY2951742 and placebo, respectively. Adverse events reported by approximately \u22655% of LY2951742-treated patients and more frequently with LY2951742 than placebo included upper respiratory tract infections, injection site pain, neck pain, abdominal pain, dizziness, injection site erythema, rash, hypertension and pain in extremity.Treatment with LY2951742 demonstrated significant separation from placebo on the primary and secondary endpoints. LY2951742 appeared to be safe and well-tolerated."}
+{"text": "Antiretroviral therapy (ART) initiation during treatment for tuberculosis (TB) improves survival in HIV/TB co-infected patients. Data on ART outcome for HIV/TB co-infected patients managed in primary health care in low-income regions is limited. We compared virological suppression rates, mortality and retention in care in HIV-positive adults receiving care in five Ethiopian health centres with regard to TB co-infection.HIV-positive ART-na\u00efve adults eligible for ART initiation were prospectively recruited from October 2011 until March 2013. At inclusion, all patients submitted sputum for microbiological TB testing . Virological suppression rates after six months of ART  with regard to TB status was the primary outcome. The impact of HIV/TB co-infection on VS rates was determined by multivariate regression analysis. Mortality and retention in care were analyzed by proportional hazard models.Among 812 participants , 678 started ART during the follow-up period . Median CD4 cell counts at ART initiation were 161 cells/\u00b5L  and 184  for TB and non-TB patients, respectively (p=0.05). No difference in retention in care between TB and non-TB patients was observed during follow-up; 25 (3.7%) patients died and 17 (2.5%) were lost to follow-up . Overall rates of VS at six months were 72.1% (<40 copies/mL) and 88.7% (<400 copies/mL), with similar results for subjects with and without TB co-infection (<40 copies/mL: 65/92 (70.7%) vs. 304/420 (72.4%), p=0.74; <400 copies/mL: 77/92 (83.7%) vs. 377/420 (89.8%), p=0.10, respectively). CD4 cell count increase during treatment was 87  and 103 cells/\u00b5L  for TB and non-TB patients, respectively, with no significant difference between the two groups (p=0.49).High rates of VS were achieved in adults receiving ART at Ethiopian health centres managed by non-physician clinicians, with no significant difference with regard to TB co-infection. These findings demonstrate the feasibility of combined ART and anti-TB treatment at primary health care level in low-income countries. This study is registered with clinicaltrial.gov, NCT01433796."}
+{"text": "Long-term prognosis after coronary artery bypass grafting (CABG) is related to the patency of coronary grafts, and pathogenesis of graft closure is linked to platelet aggregation. We analyzed the effect on late outcomes of postoperative dual antiplatelet treatment (DAT), maintained during the first year, compared to single antiplatelet treatment (SAT).A) Primary: Evaluation of adverse cardiovascular events: Hospital admission for acute coronary syndromes (ACS), unplanned target-vessel revascularization (UTVR), stroke and death of cardiovascular origin. B) Secondary: Evaluation of safety: analysis of bleeding events (BE).Retrospective study including all CABG patients during the years 2009-2010, with two years of clinical follow-up. Patients were classified in: A) SAT: daily 100 mg ASA. B) DAT: daily 100 mg ASA plus daily 75 mg clopidogrel.The study included 452 patients: 287 SAT (63.5%); 165 DAT (36.5%). 11.9% suffered a primary end-point event; 6.6% ACS; 4.4% UTVR; 1.5% stroke; 3.8% died during follow-up. Safety: 2 (0.4%) suffered a major BE, and 10 (2.2%) minor BE.DAT was associated with a reduction of the primary end-point from 14.6% to 7.3% (p = 0.020). ACS were reduced from 8.7% to 3.0% (p = 0.020). There were no differences in UTVR nor stroke. Mortality during follow-up was lower in DAT .A multivariate Cox proportional-hazards regression was performed; DAT was independently associated with the reduction of events .The greatest benefit of DAT was seen after Off-pump CABG  and in diabetic patients .DAT is associated with a reduction of late adverse cardiovascular events after CABG, especially in Off-pump CABG and in diabetic patients. DAT did not increase the risk of BE."}
+{"text": "Klebsiella pneumoniae strain IIEMP-3, isolated from Indonesian tempeh, is a vitamin B12-producing strain that exhibited a different genetic profile from pathogenic isolates. Here we report the draft genome sequence of strain IIEMP-3, which may provide insights on the nature of fermentation, nutrition, and immunological function of Indonesian tempeh. Klebsiella pneumoniae in Indonesian tempeh is intriguing since it produces vitamin B12 in tempeh   method showed that K. pneumoniae strain IIEMP-3 was determined employing a HiSeq 2000 platform  through a commercial service provider . A 350-bp paired-end library was constructed and sequenced, producing 10 million paired-end reads with read lengths of 100\u00a0bp.The nucleotide sequence of De novo assembly was processed using Velvet version 1.2.07 (n 1.2.07  with k-mn 1.2.07  resultedn 1.2.07  with 5 tn 1.2.07  with agghttp://www.ncbi.nlm.nih.gov/genome/annotation_prok). A total of 5,285 genes, 5,096 coding sequences (CDSs), 106 pseudogenes, 5 rRNAs, 77 tRNAs, and 1 noncoding RNA (ncRNAs) were identified.Genome annotation was performed employing the NCBI Prokaryotic Genome Annotation Pipeline (Klebsiella pneumoniae IIEMP-3 has been deposited in GenBank under the accession number LMAP00000000.The genome sequence of"}
+{"text": "A peer-reviewed Formal Comment by Prof. Backert and colleagues discussing results relating to this work has been published as Tegtmeyer N, Lind J, Schmid B, Backert S (2014) Helicobacter pylori CagL Y58/E59 Mutation Turns-Off Type IV Secretion-Dependent Delivery of CagA into Host Cells. PLOS ONE 9(6): e97782. doi:10.1371/journal.pone.0097782Following discussion among Associate Editor Matt Hodgkinson, the authors, and the handling editor Jun Sun, and in response to the issues raised in the Formal Comment, we are posting a correction to address: 1) confirmation of the strain mutations by PCR, sequencing, and RT-PCR; 2) clarification of the availability of the strains.The authors have provided DNA sequencing and RT-PCR data in supporting information files:cagL mutant (A), revertant, and amino acid replacement mutants (B).cagL mutant, revertant, and amino acid replacement mutants using PCR.H. pylori clinical strain Hp1033 wild type, cagL insertion mutant, revertant, and amino acid replacement mutants.cagL expression for Hp1033 wild type, cagL mutant and replacement mutants co-cultured with AGS cells at pH 7.4 for 1 hour.CagL antibody, and so they are not able to show protein expression.The authors have no Anti-H. pylori reported in the manuscript may be done following a formal request. This application is necessary to comply with the regulatory processes in the authors' institution and the customs/export regulations between Taiwan and other countries.Sharing of the strains/isolates of Table S1Primers used for sequencing and RT-PCR.Click here for additional data file.Figure S1cagL mutant (A), revertant, and amino acid replacement mutants (B).The diagram of construction of Click here for additional data file.Figure S2cagL mutant, revertant, and amino acid replacement mutants by using PCR. PCR Amplicons from wild type, revertants, and amino acid replacement mutants are 1.1kb (using primer cagL-5 & cagL-6) or 1.4kb (using primer cagIL-1 & cagL-6), from cagL insertion mutants are 2kb. (A) M: marker; w: Hp1033 wild type; lane1-16: Hp1033 cagL::cat. (B) lane1-12: 26695 cagL::cat; lane 13-23: J99 cagL::cat; lane 24: Hp1033 cagL-Y58/E59 revertant. (C) lane 25-29: Hp1033 cagL-Y58D/E59 amino acid replacement mutants; lane 30: Hp1033 cagL::cat ; lane 31: Hp1035 cagL::cat. (D) lane 32-35: Hp1033 cagL-Y58/E59K amino acid replacement mutants. (E) lane 36-39: Hp1033 cagL-Y58D/E59K amino acid replacement mutants ; lane 40: Hp1035 cagL-Y58D/E59K amino acid replacement mutants. Arrows indicate the clones selected in this study.Confirming Click here for additional data file.Figure S3H. pylori clinical strain Hp1033 wild type, cagL insertion mutant, revertant, and amino acid replacement mutants. Multiple Alignments were processed by MAFFT L-INS-1 (v6.850b) from EMBL-EBI website. The blue blocks indicate the region of chloramphenicol resistance gene. The yellow block indicates the position of amino acid 58 and 59 residues.The sequencing results of Click here for additional data file.Figure S4cagL mutant and replacement mutants co-cultured with AGS cells at pH 7.4 for 1 hour, cagL expression was examined by RT-PCR. There was no difference in CagL expressions among Hp1033 wild type, revertant, and amino acid replacement mutants. The 3rd lane indicated there was no RNA contamination.After Hp1033 wild type, Click here for additional data file."}
+{"text": "The publisher apologizes for the error.The word \u201cAlleviating\u201d is misspelled in the title. The correct title is: Penehyclidine Hydrochloride Pretreatment Ameliorates Rhabdomyolysis-Induced AKI by Activating the Nrf2/HO-1 Pathway and Alleviating Endoplasmic Reticulum Stress in Rats. The correct citation is: Zhao W, Huang X, Zhang L, Yang X, Wang L, Chen Y, et al. (2016) Penehyclidine Hydrochloride Pretreatment Ameliorates Rhabdomyolysis-Induced AKI by Activating the Nrf2/HO-1 Pathway and Alleviating Endoplasmic Reticulum Stress in Rats. PLoS ONE 11(3): e0151158. doi:"}
+{"text": "A. baumannii, K. pneumonia and P. aeruginosa strains, isolated from critically ill patients in a Greek ICU.The emergence of multidrug-resistant (MDR) pathogens is a major cause of infection-related mortality among critically ill patients. The synergistic effect between commonly used antibiotics against difficult to treat nosocomial MDR Gram-negative strains, if present, could provide a viable option as an alternative therapy. The aim of this study was to investigate the potential of antibiotic synergy against MDR A. baumannii, 41 K. pneumoniae and 64 P. aeruginosa strains, isolated during the period 2010 to 2013. All strains were resistant to carbapenems and showed reduced susceptibility or resistance to tigecycline or colistin (MIC >2), in accordance with CLSI guidelines. We evaluated double-drug combinations of carbapenem (CRB)/colistin (COL), tigecycline (TG)/COL, rifampicin (RIF)/COL, CRB/ gentamicin (GEN), CRB/amikacin (AMK) for A. baumannii, TG/COL, CRB/COL, piperacillin-tazobactam (PIP/TAZ)/GEN, CRB/GEN for K. pneumoniae and AMK/(PIP/TAZ), AMK/aztreonam (AZT), AMK/cefepime (CEF), AMK/CRB and CRB/COL for P. aeruginosa strains. In order to perform synergy tests, the E-test methodology  was used. Synergy was defined as a fraction inhibitory concentration (FIC) index \u22640.5, additive effect 0.5 to 1, indifferent or antagonistic effect >2 (Lorian definition).We tested 59 A. baumannii strains, the synergy effect of CRB/ COL was 55.9%, RIF/COL 38.9%, CRB/GEN 22%, CRB/AMK 20.3% and TG/COL 16.9%, respectively. Against 41 K. pneumoniae strains, synergy rates were: CRB/COL 43.9%, CRB/GEN 31.7%, PIP/TAZ/GEN 29.2% and TG/COL 24.4% respectively. Against 64 P. aeruginosa strains, synergy rates were: AMK/PIP/TAZ 64.6%, AMK/AZT 64.6%, AMK/CEF 58.3%, CRB/ COL 52%, AMK/CRB 25%.Against 59 MDR A. baumannii and K. pneumoniae strains tested was CRB/COL. The next most effective combination was RIF/COL and CRB/GEN respectively. No competitive effect was observed for RIF/COL combination in all cases tested. The most effective combinations for P. aeruginosa strains were AMK plus PIP/TAZ or AZT or CEF. The next most effective combination was CRB/ COL. We recommend implementation of an antibiotic synergy test for MDR pathogens as a routine antimicrobial test in the hospitals' microbiology laboratories, especially for critically ill patients, since some combinations seem to excel. Further studies are needed for the correlation of these combinations with clinical efficacy.The most effective combination for both the"}
+{"text": "A 48-year-old female presented with intermittent high-grade fever, chills, and severe myalgia for 4 days. There was no lymphadenopathy or hepatosplenomegaly. Investigations revealed hemoglobin concentration of 142 g/L; leucocyte count of 3.5x109/L with 54% neutrophils, 40% lymphocytes, 1% eosinophils, and 5% monocytes; and thrombocytopenia (platelet count of 55x109/L). Peripheral smear revealed numerous plasmacytoid lymphocytes and occasional cells with eccentrically placed nucleipacked with multiple prominent cytoplasmic vacuoles, morphologically consistent with Mott cells . MeanwhiNonmalignant reactive peripheral blood plasmacytosis can occur in tumors, autoimmune conditions, and infections . Polyclo"}
+{"text": "Delftia tsuruhatensis MTQ3 is a plant growth-promoting rhizobacterium (PGPR) isolated from tobacco rhizosphere. Here, we report the draft genome sequence of D.\u00a0tsuruhatensis MTQ3. Several functional genes related to antimicrobial activity and environment adaption have been found in the genome. This is the first genome sequence of D.\u00a0tsuruhatensis related to PGPR. Delftia tsuruhatensis, the type species of Delftia, is considered a plant growth-promoting rhizobacterium (PGPR)  \u20133, an or (PGPR) \u2013, 5, and  (PGPR) \u2013, 7. D.\u00a0tD.\u00a0tsuruhatensis MTQ3. Genomic DNA was extracted and sequenced using the Illumina HiSeq 2500 platform. The whole shotgun sequencing produced 8,206,833 paired-end reads with an average insert size of 580\u00a0bp (over 780-fold coverage). Another mate-paired (MP) library containing 3- to 5-kb inserts was constructed, and 13,462,994 paired-end reads were produced. All these raw data were then filtered by Trimmomatic v 0.30  (http://www.ncbi.nlm.nih.gov/genome/annotation_prok/).Here, we report the draft genome sequence of c v 0.30 , and Gapc v 0.30 . Final aD.\u00a0tsuruhatensis MTQ3 is phylogenetically related to D.\u00a0tsuruhatensis 391 (98.34%) (GenBank accession no. JNWH00000000), D. acidovorans CCUG 274B (97.96%) (AGYX00000000), D.\u00a0acidovorans SPH-1 (94.94%) (NC_010002), Delftia sp. 670 (94.30%) (JNWI00000000), D.\u00a0acidovorans CCUG 15835 (92.70%) (AGYY00000000), and Delftia sp. Cs1-4 (91.37%) (NC_015563).The genome consists of 6.27\u00a0Mb, with a G+C content of 66.79%. A total of 5,005 coding sequences (CDS), 13 pseudogenes, 72 tRNA genes, 1 noncoding RNA (ncRNA), and 10 rRNA genes were identified. Average nucleotide identity (ANI) analysis  revealedD.\u00a0tsuruhatensis MTQ3 were identified in the genome, such as the bacteriocin gene (AA671_02705), polyketide synthase (PKS) gene (AA671_12420), nonribosomal peptide synthetase (NRPS) gene (AA671_12430), balhimycin synthesis gene (AA671_12395), quinolinate synthesis gene (AA671_01765), and phenazine synthesis gene (AA671_00865). Some genes responsible for the processing and transport of antibiotics were identified in the genome, including antibiotic biosynthesis monooxygenase gene , macrolide transporter gene (AA671_00980), and antibiotic ABC transporter substrate-binding gene (AA671_25335).Many genes involved in the antimicrobial activity of D.\u00a0tsuruhatensis MTQ3 possesses genes with likely relevance to rhizosphere competence and adaptation in polluted environments. Specifically, quercetin dioxygenase  is likely a plant microbe-signaling bacterium involved in degrading plant-produced antimicrobial root exudates (In addition, exudates , 14. Proexudates . Genes rD.\u00a0tsuruhatensis MTQ3 has been deposited in DDBJ/ENA/GenBank under the accession number LCZH00000000. The version described in this paper is the first version, LCZH01000000.The whole-genome shotgun project of"}
+{"text": "Chondroitin sulfate proteoglycan 4 (CSPG4) is a membrane bound proteoglycan, composed of an N-linked 280kDa glycoprotein, and a single 450kDa chondroitin sulfate linked proteoglycan . CSPG4 e16 pediatric tumor lines were assessed for CSPG4 expression. CSPG4 was detected by flow cytometry in 69% of the cell lines assessed: osteosarcomas: 143b,HOS-MNNG,MG63; synovial sarcomas: SYO-I,HSSY-II, alveolar/embryonal rhabdomyosarcomas: Rh18c,Rh30/Rh36, medulloblastoma: DAOY, melanoma: Mel624,Mel1300. CSPG4 expression site density was estimated with BD PE Quantibrite beads by flow cytometry.Using two different single chain variable fragments (scFv) known to target CSPG4 (225.28 and TP41.2), two anti-CSPG4 second generation CAR constructs were synthesized by transient transfection of retroviral packaging lines. Both constructs have identical intracellular signaling components . Healthy donor T cells were transduced with viral supernatants, and assessed for CAR expression. Anti-tumor activity was measured by cytokine release and tumor specific lysis. Cell surface expression of the CSPG4-based CARs was detected by flow cytometry using conjugated Protein L to target the expressed scFv light chain. 225.28 and TP41.2 based CAR T cells induced 25% and 31% lysis of target pediatric tumor lines, respectively (E:T ratio of 20:1). On further investigation, TP41.2 based CAR T cells were able to specifically lyse CSPG+ cell lines 143b, Rh30, and Rh18c at 40%, 37%, and 16%, respectively (E:T ratio of 30:1). Additionally, the TP41.2 and 225.28 CAR T cells were assessed for Interferon gamma (IFNy) production by flow cytometry. Compared to mock T cells coincubated with varying tumor lines , the TP41.2 and 225.28 CAR+ T cells had significantly positive populations for IFNy production (p < 0.0001), predominantly in the CD8+ population.in vitro at significant levels (p = 0.01). Targeting CSPG4 with CAR redirected T cells represents a new strategy for eliminating these pediatric malignancies in an antigen specific, cell-mediated manner.High levels of CSPG4 are present on a variety of pediatric solid tumors. TP41.2 CSPG4-based CAR T cells target, generate significant levels of IFNy, and lyse osteosarcoma"}
+{"text": "Porphyromonas gingivalis is strongly associated with periodontitis. P.\u00a0gingivalis strain trafficking and tissue homing differ widely, even among presumptive closely related strains, such as W83 and A7436. Here, we present the genome sequence of A7436 with a single contig of 2,367,029\u00a0bp and a G+C content of 48.33%. Porphyromonas gingivalis is an oral bacterium that is associated with periodontal disease  and grown as previously described (escribed . Genomicescribed  (Roche).https://www.ebi.ac.uk/~zerbino/velvet/) (http://www.phrap.org/consed/consed.html) (\u2013http://metagenomics.anl.gov) (http://img.jgi.doe.gov/er) (https://geneprimp.jgi-psf.org) (GS-20 reads were assembled using Velvet version 0.7.63 (velvet/)  and Newbed.html) \u201324 and banl.gov)  and IMG-.gov/er) , then ampsf.org) .P.\u00a0gingivalis A7436 has approximately 57-fold coverage and contains a single contig of 2,367,029\u00a0bp (G+C content of 48.33%). A total of 2,078 genes were annotated, which included 2,011 predicted coding sequences (CDSs), 53 tRNAs, 12 rRNAs, and 1 transfer-messenger (tmRNA). There are 234 subsystems in the genome. 169 protein metabolism, 164 cofactors, vitamins, prosthetic groups, and pigments, 74 RNA metabolism, 87 DNA metabolism, 96 carbohydrates, and 19 membrane transport subsystem features were observed.The genome of P.\u00a0gingivalis A7436 genome was compared to P.\u00a0gingivalis strains W83, ATCC 33277, and TDC60 using RAST (The annotated ing RAST  and IMG-ing RAST . All-to-ing RAST , 17. HowP.\u00a0gingivalis strains.The availability of the A7436 genome expands our ability to compare observed behavior with genotype in a growing number of CP011995. The version described is the first version.This genome sequencing project was deposited in GenBank under accession no."}
+{"text": "We also accessed the proliferative role and downstream targets of Pak1 in endometrial cancer. Pak1 was expressed in cytoplasm whereas Pak4 and p-Pak4 were expressed in both cytoplasm and nucleus of endometrial tissues. In normal endometrium, significantly higher Pak1 (P = 0.028) and cytoplasmic p-Pak2 (P = 0.048) expression was detected in proliferative endometrium than secretory endometrium. Pak1, cytoplasmic and nuclear Pak4 and nuclear p-Pak4 was significantly overexpressed in endometrial cancer when compared to atrophic endometrium . Moreover, type I endometrioid carcinomas showed significantly higher Pak1 expression than type II non-endometrioid carcinomas (P<0.001). On the other hand, Pak1, Pak4 and p-Pak4 expression negatively correlated with histological grade  while p-Pak2 and cytoplasmic Pak4 expression inversely correlated with myometrial invasion . Furthermore, patients with endometrial cancers with lower cytoplasmic Pak4 expression showed poorer survival (P = 0.026). Multivariate analysis showed cytoplasmic Pak4 is an independent prognostic factor. Functionally, knockdown of Pak1, but not Pak4, in endometrial cancer cell line led to reduced cell proliferation along with reduced cyclin D1, estrogen receptor (ER\u03b1) and progestogen receptor (PR) expression. Significant correlation between Pak1 and PR expression was also detected in clinical samples. Our findings suggest that Pak1 and cytoplasmic p-Pak2 may promote cell proliferation in normal endometrium during menstral cycle. Pak1, cytoplasmic and nuclear Pak4 and nuclear p-Pak4 are involved in the pathogenesis of endometrial cancer especially in postmenopausal women. Pak1 promote endometrial cancer cell proliferation, particular in type I endometrioid carcinoma. Cytoplasmic Pak4 can be potential prognostic marker in endometrial cancer.p21-activated kinases (Paks) are serine/threonine protein kinases involved in biological events linked to malignant tumor progression. In this study, expression of Pak1, p-Pak2 Ser Endometrial cancer is the most common gynecological malignancy worldwide  and its p21-activated serine/threonine kinases (Paks) are effectors for the small Rho GTPases Rac1 and Cdc42 that play important roles in a variety of cellular functions, including cell morphogenesis, motility, survival, anchorage-independent growth and angiogenesis, all of which are prerequisite steps for tumor formation and tumor invasion . Based oSer20 expression in ovarian cancer was also detected , and cleared by centrifugation at 4\u00b0C. Protein concentration was determined by DC (detergent compatible) protein assay . 20 \u03bcg protein was resolved by SDS-PAGE, transferred to polyvinylidene difluoride membrane, and hybridized with corresponding antibodies . Mann-Whitney test was applied for comparison between two groups. Kruskal-Wallis rank test was applied for comparison among multiple groups. Survival analysis was performed by Kaplan-Meier analysis and log-rank test. P = 0.001) and secretory (P = 0.004) endometrium, atypical hyperplasia (P = 0.042) and endometrial cancers (P = 0.044) (P<0.05) was observed  . Signifiobserved . Althougobserved . SimilarP<0.05) except complex hyperplasia whereas nuclear p-Pak2 only showed significantly higher expression in proliferative endometrium than atrophic (P = 0.046) and secretory (P = 0.028) endometrium and endometrial cancers (P = 0.007) (P = 0.007) and pure atypical hyperplasia (P = 0.013) showed significant higher cytoplasmic p-Pak2 expression  and pure atypical hyperplasia   . Althougpression . Furtherctively) .P<0.05) (P = 0.047) and endometrial cancers (P = 0.036) (P = 0.026)  (P<0.05)  . SignifiUp-regulation of Pak1 and Pak4 mRNA and protein expression was also found in cancer cell lines  compared with normal endometrial cells by qPCR  and WestP<0.05), but not with other clinical parameters (P<0.05) (P<0.05) (P<0.05)  and patients\u2019 age at diagnosis  . There w(P<0.05) . Higher (P<0.05) . No signP = 0.026), but not Pak1, nuclear and cytoplasmic p-Pak2, nuclear Pak4, nuclear and cytoplasmic p-Pak4, resulted in a poorer survival in endometrial cancer patients .Kaplan-Meier-survival analyses revealed that lower expression of cytoplasmic Pak4 Click here for additional data file.S2 Table(DOC)Click here for additional data file.S3 Table(DOC)Click here for additional data file."}
+{"text": "Postoperative morbidity after sleeve gastrectomy is decreasing, but remains significant. Bleeding and surgical fistules remain the leading causes of morbidity and mortality. In several studies in postoperative care of obese patients, non-invasive positive pressure ventilation (NPPV) reduced the risk of lower respiratory tract infection and pneumonia , thereby2 above 90%. After period: standard treatment plus NPPV - all patients were submitted to a systematic postoperative protocol: NPPV was provided using an oxygen CIPAP system with 5 cmH2O. Statistical analysis: complication rates were compared using the chi-square test. P < 0.05 was considered statistically significant.A 4-year before-after study was conducted in a 19-bed intermediate care unit of a private hospital. Before period: standard treatment - all patients received oxygen supplementation to achieve SaOP <5%.A total of 857 patients were included. Inclusion characteristics were similar in the two groups: Before group - noNPPV: 352 patients, 2010 to 2011. Age: 40.58 \u00b1 10.94, BMI: 42.79 \u00b1 5.51, sex ratio F/M: 0.81. After group - NPPV: 504 patients, 2012 to 2013. Age: 40.81 \u00b1 11.24, BMI: 42.92 \u00b1 5.09, sex ratio F/M: 0.77. There is a significant between-group difference in the complication rate: Before group - noNPPV: 10 surgical fistula (2.84%) and six postoperative bleeding (1.70%); After group - NPPV: seven surgical fistula (1.39%) and three postoperative bleeding (0.6%). The overall complication rate fell from 4.54% to 1.98%. The chi-square statistic = 4.58. The number of degrees of freedom is 1. The value returned from the chi-square statistic is Systematic use of NPPV significantly improves morbidity in the postoperative care of sleeve gastrectomy."}
+{"text": "Burkholderia sp. strain MP-1 was isolated from pesticide-contaminated soil. Herein, we report the draft genome sequence of strain MP-1, which contains 168 contigs of 8,611,053 bp, with a G+C content of 62.55% and 7,631 protein-coding genes. Burkholderia spp. are widely distributed in nature and include a large variety of related environmental, clinical, and agribiotechnological species (Burkholderia now comprises 78 species (http://www.bacterio.net/burkholderia.html). A methyl parathion (MP)-degrading bacterial strain, designated as Burkholderia sp. strain MP-1 (LMG 27927 = MCCC 1K00250), was isolated from the contaminated soil of a former pesticide-manufacturing company in Jiangsu province of China. The 16S rRNA sequence of the newly isolated strain showed the highest similarity to that of Burkholderia grimmiae DSM 25160T (98.45%)  of 8,611,053\u00a0bp (131-fold coverage), with an average G+C content of 62.55%. Automatic gene annotation was performed by the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html). The genome comprises 7,791 genes, including 55 tRNA genes for all 20\u00a0amino acids, 3 rRNA genes, 1 small noncoding RNA gene, 101 pseudogenes, and 7,631 protein-coding genes (CDS) (with an average length of 950\u00a0bp), which give a coding intensity of 84.2%. In all, 4,853 proteins were assigned to Clusters of Orthologous Groups (COG) families , with a MiSeq system using paired-end sequencing technology. A total of 3,758,646 pair reads and 9,573 clean single reads  were assembled using Newbler 2.6. The genome of Burkholderia strains were investigated, such as that of Burkholderia pyrrocinia CH-67, whose genome contains genes encoding enzymes for aromatic compound degradation .The draft genome sequence of strain MP-1 has been deposited at GenBank under the accession no."}
+{"text": "Oncotarget. 2015; 6:15594-15609doi:10.18632/oncotarget.3709Present Figure 2: During the assembly of figure 2C, the same image was inadvertently used for both the non-transfected and empty vector control.Corrected Figure 2:"}
+{"text": "As the eighth leading cause of annual mortality in the USA, influenza A viruses are a major public health concern. In 20% of patients, severe influenza progresses to acute lung injury (ALI). However, pathophysiological mechanisms underlying ALI development are poorly defined. We reported that, unlike wild-type (WT) C57BL/6 controls, influenza A virus-infected mice that are heterozygous for the F508del mutation in the cystic fibrosis transmembrane conductance regulator (HETs) did not develop ALI. This effect was associated with higher IL-6 and alveolar macrophages (AMs) at 6 days postinfection (d.p.i.) in HET bronchoalveolar lavage fluid (BALF). In the present study, we found that HET AMs were an important source of IL-6 at 6 d.p.i. Infection also induced TGF-\u03b2 production by HET but not WT mice at 2 d.p.i. TGF-\u03b2 neutralization at 2 d.p.i. (TGF-N) significantly reduced BALF IL-6 in HETs at 6 d.p.i. Neither TGF-N nor IL-6 neutralization at 4 d.p.i. (IL-6-N) altered postinfection weight loss or viral replication in either mouse strain. However, both treatments increased influenza A virus-induced hypoxemia, pulmonary edema, and lung dysfunction in HETs to WT levels at 6 d.p.i. TGF-N and IL-6-N did not affect BALF AM and neutrophil numbers but attenuated the CXCL-1/keratinocyte chemokine response in both strains and reduced IFN-\u03b3 production in WT mice. Finally, bone marrow transfer experiments showed that HET stromal and myeloid cells are both required for protection from ALI in HETs. These findings indicate that TGF-\u03b2-dependent production of IL-6 by AMs later in infection prevents ALI development in influenza A virus-infected HET mice. It has been estimated that seasonal influenza infections result in 12 influenza-related deaths per 100,000 persons per year in the United States, most of which occur in the elderly , which is associated with poor prognosis . Once AL+ absorption via epithelial Na+ channels (ENaC) and Cl\u2212 absorption or secretion through the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel  were intranasally infected with 10,000 plaque-forming units (pfu)/mouse of egg-grown influenza A/WSN/33 (H1N1) in 50 \u03bcl PBS with 0.1% BSA  wk old w. In our Thin sections (3 \u03bcm) were prepared from formalin-fixed, paraffin-embedded lung tissue . IL-6 wail-6 mRNA was determined by the \u0394\u0394Ct method and normalized to the endogenous control 18s rrna, which was not altered by infection  at 2 d.p.i. by intraperitoneal (i.p.) injection . To neut137Cs irradiator (6/mouse) (in 0.5 ml PBS) were then transplanted by tail vein injection into recipient mice, which were placed on Baytril antibiotic water bottles for 2\u20133 wk peri-irradiation. This protocol resulted in no outward clinical signs.As in our previous studies, bone marrow-recipient mice were irradiated with 1,000 cGy given in two doses by a 2 saturation, heart rate, lung homogenate viral titers, lung wet:dry weight ratios, lung mechanics, and BALF inflammatory mediators were performed as in our previous studies . Gaussian data distribution was verified by the method of Kolmogorov and Smirnov. Differences between group means were analyzed by one-way ANOVA, with Tukey-Kramer multiple-comparison posttests. il-6 gene expression in AMs isolated from HET mice than AMs from WT mice at 6 d.p.i. (We reported previously that attenuation of cardiopulmonary dysfunction in HETs infected with the A/WSN/33 H1N1) influenza strain  was associated with exaggerated AM and IL-6 responses at 6 d.p.i. N1 influeA. In con6 d.p.i. C.Given the known role of TGF-\u03b2 as an anti-inflammatory cytokine  and an iTo determine whether the exaggerated IL-6 response to infection in HET mice was TGF-\u03b2 dependent, influenza A virus-infected mice were treated with a single dose of a neutralizing antibody to TGF-\u03b2 at 2 d.p.i. (TGF-N) . ControlTo determine whether altered TGF-\u03b2 and IL-6 responses to infection contribute to protection from influenza A virus-induced ALI, we examined the effects of TGF-N and IL-6-N on weight loss, viral replication, and cardiopulmonary function in influenza A/WSN/33 virus-infected mice at 6 d.p.i. We selected this time point because it would allow us to directly compare the downstream effects of treatment with anti-TGF-\u03b2 at 2 d.p.i. with those of treatment with anti-IL-6 at 4 d.p.i. Postinfection weight loss at 6 d.p.i. did not differ between untreated WT and HET mice A. IgG trIn the absence of treatment, lung water content (wet:dry weight) remained normal  in HET mice at 6 d.p.i. but was significantly elevated in WT controls . Both TGIn the absence of treatment or after treatment with IgG, static and dynamic lung compliance were both significantly higher  in infected HETs at 6 d.p.i. B. ProtecAs in our earlier studies, BALF from influenza A/WSN/33 virus-infected HET mice contained far greater numbers of AMs than WT controls at 6 d.p.i. A. HoweveWe previously reported that BALF IFN-\u03b3, IL-10, CC chemokine ligand 2 (CCL-2)/monocyte chemotactic protein 1 (MCP-1), CCL-5/RANTES, and CXCL-10/inducible protein 10 content did not differ between WT and HET mice at 6 d.p.i. . However, both treatments significantly reduced BALF CXCL-1/KC content in both mouse strains. In WT mice, both TGF-N and IL-6-N also significantly reduced BALF IFN-\u03b3 but increased IL-12 at 6 d.p.i. TGF-N, but not IL-6-N, also reduced BALF CCL-5/RANTES in WT mice at this time point. In contrast, neither treatment impacted BALF IFN-\u03b3 or CCL-5/RANTES in HET mice, and only IL-6-N significantly increased BALF IL-12 at 6 d.p.i. Treatment with nonspecific IgG had no effect on CXCL-1/KC, IFN-\u03b3, IL-12, or CCL-5/RANTES in either mouse strain.We previously demonstrated that HET AMs were necessary for protection from influenza A virus-induced ALI in HET mice . To deteDespite a 50% reduction in cell-surface CFTR expression and anion transport , we and Previous investigators have reported that an attenuated AM response contributes to increased influenza severity , 59, 64.We found no differences in viral replication kinetics and BALF neutrophil counts between WT mice, untreated or IgG-treated HETs, and antibody-treated HETs at 6 d.p.i. This indicates that, in our model, influenza-induced ALI is independent of these factors. Moreover, we did not find that development of ALI in HETs following TGF-N and IL-6-N was associated with development of a so-called \u201ccytokine storm\u201d , 33, 62.Some investigators have reported an association between high BALF IL-6 and increased influenza mortality , 39, 56.Greater influenza severity has also been linked to inadequate anti-inflammatory responses. TGF-\u03b2 is a potent inflammatory regulator , and inc1-subtype adenosine receptors, which induces increased CFTR-mediated Cl\u2212 secretion . A. Amer was supported by a Cystic Fibrosis Foundation Research Grant and The Ohio State University Public Health Preparedness in Infectious Diseases Program. I. Davis was supported by The Ohio State University Public Health Preparedness in Infectious Diseases Program and The National Heart Lung and Blood Institute at the National Institutes of Health (R01-HL102469).P. Woods was supported by No conflicts of interest, financial or otherwise, are declared by the authors.Author contributions: P.S.W., M.F.T., and N.M.C. performed experiments; P.S.W., M.F.T., N.M.C., and I.C.D. analyzed data; P.S.W. and I.C.D. interpreted results of experiments; P.S.W. and I.C.D. prepared figures; A.O.A. and I.C.D. edited and revised manuscript; A.O.A. and I.C.D. approved final version of manuscript; I.C.D. conception and design of research; I.C.D. drafted manuscript."}
+{"text": "This study was undertaken to determine virulence factors among clinical Enterococcus species by phenotypic and molecular methods.Enterococci are opportunistic pathogens causing severe urinary tract infections, surgical wound infections, bacteremia and bacterial endocarditis. Aggregation substance  was the predominant species obtained, followed by E. faecalis (73/157). 72/157(45.85%) strains were positive for hemolysin, 61/157(38.85%) gelatinase, 16/157(10.19%) strong and 66/157(42%) were moderate biofilm producers phenotypically. PCR results showed, 37/157(23.56%) cylA, 81(51.59%) gelE, 10(6.36%) hyl, 87(55.41%) asa1 and 78(49.68%) isolates were positive for esp genes. Only 25/72, 32/61, 6/16 and 7/16 isolates were phenotypically and genotypically positive for cylA +hemolysin, gelE +gelatinase, esp +biofilm and asa1 +biofilm, respectively. Interestingly, E. faecalis carried multiple virulent genes (>4 genes) when compared with E. faecium among our study isolates.asa1>gelE>esp were the predominant genes observed. Majority of E. faecalis isolates were strong and E. faecium were moderate biofilm producers.Hemolysin and gelatinase were the predominant virulence factors expressed phenotypically whereas,"}
+{"text": "Feature tracking strain (FTS) is a new technique to evaluate myocardial deformation from routinely acquired cardiac magnetic resonance (CMR) cine images, however, it has not been validated in single ventricle patients. The purpose of this study is to validate FTS against myocardial tagged harmonic phase (HARP) images, considered the reference standard in non-invasive deformation analysis.We retrospectively analyzed CMRs of 15 consecutive single ventricle Fontan patients  between HARP and FTS, Pearson r = 0.67, p = 0.006. Average GCS was -19 (95% CI -4.3 to -33.8) from HARP and -16.7 (95% CI -6.4 to -26.9) from FTS. FTS yielded lower values (bias -2.3) than HARP (Figure Feature tracking analysis has moderate agreement with grid-tagged HARP measurements of circumferential strain in Fontan patients, with a trend towards lower strain values via FTS. Further validation of FTS in a large sample is warranted.Dr Kevin K. Whitehead: NIH K23 Grant HL089647 from the National Heart, Lung and Blood Institute. Dr. Mark Fogel: NIH R01HL098252-01, \"Understanding mechanisms of Fontan failure and key predictors for patient outcome.\""}
+{"text": "The asymmetric unit consists of one-quarter of the mol\u00adecule (S site symmetry 2) and the complete mol\u00adecule has 2/m (Ch2) point symmetry with the C=C bond in an E conformation. The geometry of the title compound is compared to those of a chloro derivative and a mercury complex.The title compound, C DOI: 10.1107/S1600536814023319/hb7285Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S1600536814023319/hb7285Isup3.cmlSupporting information file. DOI: Click here for additional data file.10.1107/S1600536814023319/hb7285fig1.tif. DOI: The mol\u00adecular structure of DTCDD with displacement ellipsoids drawn at the 30% probability level. Symmetry codes: (i) 1-x, y, z; (ii) x, 1-y, 1-z; (iii) 1-x, 1-y, 1-z.Click here for additional data file.10.1107/S1600536814023319/hb7285fig2.tif. DOI: The unit-cell packing in DTCDD viewed down the b-axis.1030564CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:"}
+{"text": "Hepatitis C virus (HCV) represents one of the persistent viral infections afflicting humankind, and a significant proportion of chronic HCV disease progresses over time through liver fibrosis, cirrhosis and hepatocellular carcinoma (HCC). One potential mechanism underlying the chronic disease is believed to be viral escape from immune surveillance via upregulation of inhibitory molecules on immune cells by HCV. We investigated the diverse expression of various inhibitory molecules in PBMCs of healthy non-HCV controls and chronically HCV infected patients.The expression of inhibitory molecules on PBMCs was investigated in chronic HCV infected patients relative to healthy non-HCV controls using standard immunological and molecular methods. The serum levels of indoleamine 2, 3 deoxygenase (IDO) and cyclooxygenase-2 (COX-2) were also investigated.p\u22640.01), PD-1 (p\u22640.01), FOXP-3 (p\u22640.01), BLIMP-1 (p\u22640.01), CD160 (p\u22640.01), CTLA-4 (p\u22640.01), TRAIL (p\u22640.01), BTLA (p\u22640.01) and LAG-3 (p\u22640.01) with fold change of 1.3, 0.4, 14.6, 0.87, 6.6, 0.4, 14.7, 10.9 and 2.5 respectively in chronically HCV infected patients. The plasma IDO and COX-2 levels were significantly higher (p=0.001) in chronically HCV infected subjects relative to healthy control.The gene expression profile of chronically HCV infected patients was significantly different from control individuals. Our results showed upregulation of TIM-3 (The upregulation of inhibitory molecules on PBMCs in chronically HCV infected patients suggest the contribution of these molecules to immune cells impairment in HCV infection. Viral persistence and eventual progression following potential evasion of the host immune armory via viral impregnation of inhibitory immune biosignatures in HCV disease pathogenesis warrants further elucidation."}
+{"text": "Recenortality . We repoWe assessed outcome for HO patients at CC (primary outcome) and hospital discharge, and at 6-month and 1-year follow- up. Single variable logistic regression analyses, adjusted by age, gender and haematological diagnosis, and multivariate analyses were performed to identify independent predictors of outcome using STATA.P < 0.001), number of organs supported , P/F ratio , inotropic requirement , and IMV status  influenced unit survival at single variable analyses. At multivariate analysis, the P/F ratio  and IMV status  independently predicted outcome.A total of 225 HO patients were admitted to CC. Median age was 59 (interquartile range (IQR) 46 to 66) years. The most common haematological diagnoses on admission were acute myeloid leukaemia in 57 (25.3%) cases, non-Hodgkin lymphoma in 54 (24%) cases and multiple myeloma in 42 (18.7%) patients. Median APACHE II score was 21 (IQR 17 to 26). A total of 164 patients (72.9%) had at least one organ supported. Unit and hospital mortality rates were 34.7% (78 patients) and 49.3% (111 patients), respectively. At 6-month and 1-year follow-up, mortality increased to 63.1% (142 patients) and 70.5% (153 patients), respectively. The APACHE II score (OR = 0.93, 95% CI = 0.89 to 0.97, Organ failures and need for organ support correlated with outcome. P/F ratio and need for IMV were independent predictors of mortality, in agreement with previously published data . The CC"}
+{"text": "The two symmetry-related iodide anions bridge two LiI cations, forming an inversion dimer in which the Li2I2 plane is nearly perpendicular to the imidazol-2-yl\u00adidene ring, with a dihedral angle of 85.5\u2005(3)\u00b0. No hydrogen bonding is observed in the crystal.In the title binuclear complex, [Li DOI: 10.1107/S2056989015009822/xu5851Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989015009822/xu5851fig1.tif. DOI: The mol\u00adecular structure of the title compound with the atom-numbering scheme and 30% probability displacement ellipsoids.Click here for additional data file.10.1107/S2056989015009822/xu5851fig2.tifa . DOI: a axis.The packing diagram viewed along the 1402139CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:"}
+{"text": "The increasing prevalence of ESBL-E in the community is a cause of concern for hospitals. Early detection of ESBL-E carriers on admission could allow timely implementation of control measures or appropriate selection of antimicrobials.To describe the current prevalence of ESBL-E rates upon admission to 4 different services at HUG, in the context of a multicenter European study (R-Gnosis).Patients admitted to 4 different services were screened by rectal swabs on admission. From January 2014 through January 2015, patients admitted to 4 wards, including: Ortho1 (sport traumatology), Ortho 2 (septic); Geriatrics (2 wards) and patients undergoing elective colorectal surgery (ECS) were screened from April 2013-October 2014.E. coli ; K. pneumoniae  and other Enterobacteriaceae . Among K. pneumonia carriers on admission, 24/26 (92.3%), had a previous hospitalization less than 12 months before admission screening and 21/26 (80.8%) had the previous hospitalization within 3 months only. ESBL-E carriage was 83/981 (8.4%) and 61/430 (14.2%) for Ortho 1 and 2 respectively; Geriatrics, 42/371 (11.3%); and ECS, 41/354 (11.6%).Overall, from 2394 admitted patients, 2136 were screened on admission (89.2%). Median age was 67.3 years (SD\u00b120.9); 51.6% were male. Only 92/2136 (4.3%) had a previously known status of ESBL carriage. A total of 226/2136 (10.6%) patients were found to be ESBL-E carriers: E. coli. Patients admitted to septic orthopedics, geriatrics and ECS had a higher prevalence on admission. The majority of ESBL- Klebsiella pneumonia carriers had a recent history of hospitalization.Overall 10.6% of patients screened were ESBL-E carriers upon admission at HUG, mostly due to ESBL-producing None declared."}
+{"text": "Pediatric chronic nonbacterial osteomyelitis (CNO) is a sterile inflammatory bone disorder in which innate and adaptive immunity dysfunction involved. Unifocal and multifocal disease courses are known. The modern treatment modalities include non-steroid anti-inflammatory drugs (NSAIDs), steroids, sulfasalazine (SSZ), methotrexate (MTX), bisphosphonates and biologic drugs - TNF\u03b1 and IL1\u03b2-antagonists, with limited data.The aim of our study was to assess children with CNO and to evaluate efficacy of treatment modalities.Our cohort of CNO patients included 22 children, 8 boys and 14 girls. Monofocal disease course was in 9/22 children (40.9), multifocal in 13/22 (59.1) with mean 6 foci per patient. Histological confirmation was made in 13/22. Repeated MRI, CT and bone scintigraphy was performed in all patients. 3 patients have family history of autoimmunity  had comorbid autoimmune diseases (different types of JIA): 5 had monoarthritis, 1 arthritis with uveitis, 1 - psoriatic arthritis, 1 - polyarthritis PF neg, 6 had enthesitis-related arthritis (3 had ankylosing spondyloarthritis) and 1 had Crohn's disease. Spine involvement was in 5/22 (22.7). Onset age was 8.5  years, the right diagnosis delay was 3.6  months.Fever at onset, high painVAS and parental VAS scores highly correlated with risk of relapse disease course. Treatment: effectiveness of NSAID only 3/10 (30%), SSZ - 1/5 (20%), corticosteroids - 0/3 (short-term effect only), MTX - 4/7 (57.1%), pamidronate (PAM) with partial response 2/12 (16.7%) and with complete response - 10/12 (83.3%). Biologics - adalimumab and etanercept were effective in 3/4 (75%) patients, who fail to NSAID, MTX, PAM and SSZ. During disease course treatment lead to decreasing sings of disease activity, such as: parental VAS (p = 0.015), pain VAS (p = 0.026), MDVAS (p = 0.026), CRP (p = 0.0008), WBC (p = 0.006), ESR (p = 0.00024), PLT (0.014). The main effectiveness belonged to PAM (p = 0.003) and biologics (p = 0.07) in decreasing of pain VAS (-100% and -80%), parental VAS (-92% and -74%) and MD VAS . We calculated the cumulative probability of survival (event of interest: CNO flare) in the entire patient sample, depending the kind of treatment  obtained by the Kaplan-Meier method. Significant difference was proved comparing 3 therapeutical branches (p = 0.028). MTX treatment was effective (p = 0.04), as well as PAM (p = 0.01) than NSAID. Only flu-like syndrome during PAM treatment was in 10/12 (83.3%). No any others side effects were reported. All patients who had flu-like syndrome on first infusion had complete response to PAM, vice verse patients, who had no such complication had only partial response to this treatment.CNO is a group of chronic inflammatory conditions associated with different rheumatic diseases. The most effective treatment modalities were PAM, biologics and MTX. PAM was safety and can reach the rapid response and maintain long sustained remission.None declared."}
+{"text": "Nevirapine (NVP) induces cytochrome P450 3A4 by which rilpivirine (RPV) is metabolized. Switching NVP to RPV could result in decreased RPV exposure with subsequent virological failure and dyslipidemia because NVP is regarded as the least dyslipidemic, non-nucleoside, reverse transcriptase inhibitor. This trial evaluated the efficacy, pharmacokinetics, safety and cardiovascular risks of switching NVP to RPV.Prospective open label controlled trial. HIV-1 patients with HIV-1 RNA <50 copies/mL on once daily NVP, emtricitabine/tenofovir (FTC/TDF) switched to single tablet RPV/FTC/TDF. Eligible patients on NVP, FTC/TDF were controls. Primary endpoint was week 12 HIV-1 RNA <50 copies/mL by intention to treat analysis. Secondary endpoints were week 24 HIV-1 RNA <50 copies/mL, NVP and RPV pharmacokinetics, safety and fasting lipids, Framingham risk scores (FRS) and Adult Treatment Panel III (ATP-III) lipid goals.Of 189 eligible patients, we included 50 RPV switchers and 139 NVP controls. Week 12 HIV-RNA was <50 copies/mL in 46/50 switchers (92.0%) which was not different from the hypothesized 90% week 12 suppression rate (p=.431). Forty-four of 50 switchers had week 24 HIV-1 RNA <50 copies/mL compared to 126/139 controls . NVP plasma concentrations were below detection level in all at week 3. Mean week 1 RPV trough concentration was 0.083 mg/L and comparable to phase III trial data (p=0.747). Adverse events occurred in 36 switchers, the majority (82.0%) were grade one. Two switchers discontinued RPV for side effects. Significant changes over 24 weeks (p<0.001) were observed in switchers on total cholesterol , low density lipoprotein (LDL)-C  and high density lipoprotein (HDL)-C . The TC/HDL-C ratio increased 0.20  and systolic blood pressure decreased 6.0 mmHg . The median FRS did not change over 24 weeks . More patients achieved LDL-C  and TC  ATP-III treatment goals at week 24 on RPV.A NVP to RPV switch does not influence RPV exposure and results in adequate ongoing HIV-1 suppression. RPV could be an option for patients at risk for cardiovascular diseases."}
+{"text": "Mesorhizobium ciceri strain CC1192, an efficient nitrogen-fixing microsymbiont of Cicer arietinum (chickpea). The genome consists of 6.94\u00a0Mb distributed between a single chromosome (6.29\u00a0Mb) and a plasmid (0.65\u00a0Mb).We report the complete genome sequence of  Cicer arietinum (chickpea), a globally important grain legume, forms a N2-fixing symbiosis with soil bacteria (rhizobia) in the genus Mesorhizobium   , 7. Thushttp://www.dpi.nsw.gov.au/content/agriculture/resources/soils/australian-inoculants-research-group) and confirmed to fix N2 effectively with C.\u00a0arietinum, using established methods (http://www.bioinformatics.babraham.ac.uk/projects/fastqc), and adaptors were removed by comparison against a comprehensive in-house adaptor sequence library. PacBio subreads were assessed using in-house software, and reads were automatically error-corrected in the assembly process.Strain CC1192 was sourced from the Australian inoculant industry mother culture ( methods . CC1192  methods . Whole-gde novo using the hybrid approach of SPAdes assembler version 3.6.2 (http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html). The genome consists of 6,943,628 bp with an average GC content of 62.49%. There are 6,642 coding sequences distributed between a single circular chromosome of 6,295,397 bp and a single plasmid of 648,231 bp.The filtered Illumina and PacBio reads were assembled on 3.6.2 , with thon 3.6.2 . The asson 3.6.2  and annoM.\u00a0loti R7A (nif and fix) appear to be located on a 419-kb symbiosis island integrated within the chromosome of CC1192 . This region also harbors a type IV secretion system, conjugative relaxase, biotin and nicotinate biosynthetic clusters, proteins known to control excision and transfer of ICEMlSymR7A  and CP015063 (plasmid pMc1192).The nucleotide sequence of the complete genome of CC1192 has been deposited in GenBank under the accession numbers"}
+{"text": "E. coli that were engineered to constitutively express highly active E. coli beta-glucuronidase intracellularly to enhance the anticancer activity of CPT-11. The engineered bacteria, E. coli (lux/\u03b2G), could hydrolyze SN-38G to SN-38, increased the sensitivity of cultured tumor cells to SN-38G by about 100 fold and selectively accumulated in tumors. However, E. coli (lux/\u03b2G) did not more effectively increase CPT-11 anticancer activity in human tumor xenografts as compared to non-engineered E. coli. SN-38G conversion to SN-38 by E. coli (lux/\u03b2G) appeared to be limited by slow uptake into bacteria as well as by segregation of E. coli in necrotic regions of tumors that may be relatively inaccessible to systemically-administered drug molecules. Studies using a fluorescent glucuronide probe showed that significantly greater glucuronide hydrolysis could be achieved in mice pretreated with E. coli (lux/\u03b2G) by direct intratumoral injection of the glucuronide probe or by intratumoral lysis of bacteria to release intracellular beta-glucuronidase. Our study suggests that the distribution of beta-glucuronidase, and possibly other therapeutic proteins, in the tumor microenvironment might be an important barrier for effective bacterial-based tumor therapy. Expression of secreted therapeutic proteins or induction of therapeutic protein release from bacteria might therefore be a promising strategy to enhance anti-tumor activity.CPT-11 is a camptothecin analog used for the clinical treatment of colorectal adenocarcinoma. CPT-11 is converted into the therapeutic anti-cancer agent SN-38 by liver enzymes and can be further metabolized to a non-toxic glucuronide SN-38G, resulting in low SN-38 but high SN-38G concentrations in the circulation. We previously demonstrated that adenoviral expression of membrane-anchored beta-glucuronidase could promote conversion of SN-38G to SN-38 in tumors and increase the anticancer activity of CPT-11. Here, we identified impediments to effective tumor therapy with  Clostridium  (E. coli (lux) and E. coli (lux/\u03b2G) in tumors reached a plateau from days 3 to 7  or E. coli (lux/\u03b2G) displayed splenomegaly (E. coli induced an immune response. This immune response may cause the transient body weight loss (9.2%) on day 3 observed after systematic administration of E. coli (The time course of lux/\u03b2G)] . The lums 3 to 7 , demonst tissues . Determiectively . Mice innomegaly , suggest E. coli .E. coli and beta-glucuronidase in tumor sections from E. coli (lux/\u03b2G), E. coli (lux), or PBS-treated mice. DAPI (blue) stained DNA in both live tumor cells and E. coli. DAPI-stained E. coli were observed as tiny spots that formed a dense cluster under UV excitation (E. coli further confirmed the location of the bacteria. Anti-E. coli beta-glucuronidase antibody staining showed that E. coli (lux/\u03b2G) but not E. coli (lux) expressed beta-glucuronidase in the tumor microenvironment. 72.4% \u00b113.9% of E. coli (lux) and 84.9% \u00b1 9.2% of E. coli (lux/\u03b2G) were located in necrotic areas of tumors as distinguished by DAPI staining. E. coli (lux) and E. coli (lux/\u03b2G) didn\u2019t show significant differences in localization to tumor necrotic areas  could enhance the anti-tumor activity of CPT-11, NOD/SCID mice bearing established HCT116 human colon tumors were i.v. injected with PBS or E. coli. Four days later, the mice were i.v. injected with 10 mg/kg CPT-11 or vehicle on two consecutive days. Treatment of mice with either CPT-11 or E. coli (lux/\u03b2G) alone significantly suppressed tumor growth as compared to treatment with vehicle alone (E. coli (lux/\u03b2G) produced significantly greater antitumor activity that either individual treatment. However, this effect did not depend on bacterial expression of beta-glucuronidase because treatment of mice with CPT-11 and E. coli (lux) suppressed tumor growth to a similar extent (E. coli (lux/\u03b2G) or E. coli (lux) produced significant but similar suppression of tumor growth (E. coli (lux/\u03b2G).To determine whether le alone . Combinar extent . Similarr growth , indicatE. coli (lux/\u03b2G) or E. coli (lux) could be caused by in vivo induction of endogenous beta-glucuronidase expression in E. coli (lux) or by in vivo loss of beta-glucuronidase expression in E. coli (lux/\u03b2G), we i.v. injected E. coli (lux/\u03b2G) or E. coli (lux) into mice bearing established HCT116 tumors and then i.v. injected a fluorescence glucuronide probe, FDGlcU, to measure in vivo beta-glucuronidase activity (E. coli (lux/\u03b2G) displayed similar luminescence emission but higher fluorescence emission than E. coli (lux) (E. coli (lux/\u03b2G) expressed significantly more beta-glucuronidase activity in tumors than did E. coli (lux). We conclude that E. coli (lux/\u03b2G) and E. coli (lux) displayed the expected high and low beta-glucuronidase activity in tumors.To investigate if the similar anticancer activity observed for treatment with CPT-11 and either activity . FDGlcU li (lux) , indicatE. coli therapy failure. To investigate differences between bacterial and adenoviral beta-glucuronidase therapy, we first compared the enzymatic activities of E. coli beta-glucuronidase (produced in E. coli (lux/\u03b2G) and murine beta-glucuronidase (produced by Ad/m\u03b2G). E. coli beta-glucuronidase hydrolyzed the glucuronidase substrate 4-Nitrophenyl \u03b2-D-glucopyranoside (pNPG) faster than did murine beta-glucuronidase at pH 7 [E. coli beta-glucuronidase displayed about 250 fold greater enzymatic activity for the hydrolysis of SN-38G compared to murine beta-glucuronidase at pH 7 (E. coli (lux/\u03b2G) or intratumoral injection of Ad/m\u03b2G. Again, the total beta-glucuronidase activity in tumors was significantly greater in mice treated with E. coli (lux/\u03b2G) than those treated with Ad/m\u03b2G (E. coli (lux/\u03b2G) can deliver more beta-glucuronidase activity to tumors as compared to direct injection of tumors with Ad/m\u03b2G.We previously showed that intratumoral injection of Ad/m\u03b2G, which allows expression of murine beta-glucuronidase as a membrane-anchored form on the surface of infected cells, can enhance the anti-tumor activity of CPT-11 . Therefo at pH 7 . Similar at pH 7 . Next, wh Ad/m\u03b2G . These rE. coli (lux/\u03b2G) or i.t. with Ad/m\u03b2G. Fluorescence signals were apparent in the tumors of mice receiving either treatment (E. coli (lux/\u03b2G)  (E. coli (lux/\u03b2G) produced more beta-glucuronidase activity in tumors, the beta-glucuronidase activity generated by Ad/m\u03b2G can more effectively hydrolyze a systemically administered glucuronide probe in the tumors.To assess the functional accessibility of beta-glucuronidase to systemically administered drugs, we i.v. injected the fluorescent glucuronide probe FDGlcU and then imaging the fluorescence intensity in tumors of mice treated i.v. with reatment , but mic(lux/\u03b2G) . A relatnce from  by the bnce from . Tumors (lux/\u03b2G) . We concE. coli (lux/\u03b2G) suggested that the chemical probe delivered via the tumor blood supply may not effectively contact beta-glucuronidase in the tumor. To test this idea, we compared the fluorescence in tumors after systemic (i.v.) or direct intratumoral (i.t.) injection of FDGlcU to mice previously treated with E. coli (lux/\u03b2G) or Ad/m\u03b2G. The dose of FDGlcU was decreased by 100 fold for intratumoral injection groups to prevent saturation of beta-glucuronidase in the tumors. While mice treated with Ad/m\u03b2G displayed similar fluorescence after either i.v. or i.t. injection of FDGlcU, the mice treated with E. coli (lux/\u03b2G) exhibited more tumor fluorescence after i.t. injection of FDGlcU (E. coli (lux/\u03b2G) was significantly greater when FDGlcU was directly injected into tumors as opposed to after i.v, administration  to allow tumor colonization and then directly injected a mixture of lysozyme/DNase I into tumors to break the bacteria cell wall. This approach was used to release beta-glucuronidase from the bacteria because the enzyme is too large (~280 kDa) to be efficiently secreted (results not shown). Mice were first i.v. injected with FDGlcU on day 4 to measure the fluorescence generated by hydrolysis of FDGlcU by intact bacteria in the tumors. After 48 h, the lysozyme/DNase I mixture was injected into tumors and then FDGlcU was again i.v. injected and tumor fluorescence was imaged. Imaging of bacterial luminescence showed the number of bacteria in tumors on days 4 and 6 were not significantly different  slow uptake of glucuronide compounds into E. coli may reduce the rate of prodrug hydrolysis and 2) the preferential accumulation of E. coli in necrotic regions of tumors may hamper efficient contact of systematically administered drugs with beta-glucuronidase. Our study suggests that both the cellular and spatial distribution of beta-glucuronidase, and possibly other therapeutic enzymes, might be an important barrier for effective bacteria-based prodrug cancer therapy.Methods to enhance the selectivity and efficacy of cancer chemotherapy are needed to improve patient outcome. Bacteria that have been modified to express enzymes for tumor-selective activation of proactive anticancer agents have been investigated as a promising approach to achieve more selective cancer therapy ,48. Heree CPT-11 ,35. Conve CPT-11 ,37. ThesE. coli was selected in our study to deliver beta-glucuronidase to tumors. There are two major advantages of using E. coli as a delivery vehicle for prodrug activation. First, E. coli can selectively colonize solid tumors, providing high tumor/normal tissue colonization ratios of around 10,000 fold [E. coli to mice. By contrast, Salmonella provides tumor/liver and tumor/spleen bacterial ratios of 100~1000 [E. coli for tumors may help reduce prodrug activation in normal tissues. Second, unlike liposomes, antibodies or replication deficient viruses, bacteria can proliferate in tumors [E. coli. These advantages have been recognized as indicated by the increasing number of papers investigating E. coli for tumor imaging and tumor therapy [E. coli used in our study (DH5\u03b1) does not express enterotoxins or lipopolysaccharides and is considered to be non-pathogenic [E. coli were also able to accumulate in murine tumors in immune-competent mice with similar toxicity as observed in immune deficient mice, suggesting that E. coli (DH5\u03b1) may be a suitable delivery vehicle for clinical applications.000 fold ,18,38. I100~1000 ,14. Like100~1000 \u201324. The n tumors \u201351. The  therapy ,20,38,52thogenic . We usedE. coli [E. coli. We constitutively expressed E. coli beta-glucuronidase in E. coli (lux/\u03b2G). These bacteria expressed about 60 ng beta-glucuronidase per 107 c.f.u. of bacteria and could convert cytotoxic concentrations of SN-38 from SN-38G. Because E. coli beta-glucuronidase can hydrolyze SN-38G to SN-38 about 250-fold faster than murine beta-glucuronidase  as compared to mice treated with Ad/m\u03b2G (E. coli beta-glucuronidase did not increase CPT-11 antitumor activity beyond that achievable by treating tumors with wild-type E. coli (E. coli (lux/\u03b2G)  .E. coli (lux/\u03b2G) therapy differ in several important facets as indicated in E. coli preferred to colonize the margins between necrotic regions and live cells (E. coli (lux/\u03b2G) with systemically-administered drugs. Second, the enzyme location in the cells differs. Ad/m\u03b2G was designed to express beta-glucuronidase on the cell membrane of infected cells whereas E. coli (lux/\u03b2G) expresses beta-glucuronidase in the periplasmic space of the bacteria. Glucuronide drugs can directly interact with membrane-anchored beta-glucuronidase but require receptor-mediated transport into E. coli to interact with beta-glucuronidase [E. coli (lux/\u03b2G) expresses E. coli beta-glucuronidase. The enzymatic activity of E. coli beta-glucuronidase is much greater than mouse beta-glucuronidase  as compared to Ad/m\u03b2G. First, beta-glucuronidase is expressed intracellularly in E. coli (lux/\u03b2G) but is present on the surface of cells as a membrane anchored form after infection with Ad/m\u03b2G. Thus, glucuronide substrates must enter bacteria, presumably via a glucuronide transporter complex present on E. coli [et al [E. coli preferentially accumulated at the border between viable cancer cells and necrotic regions  significantly increased probe hydrolysis (E. coli (lux/\u03b2G) in the tumor by intratumoral injection of lysozyme and DNase I to release beta-glucuronidase also significantly increased hydrolysis of the glucuronide probe (We identified two impediments that may account for the relatively poor performance of  E. coli , to cont E. coli . The seci [et al , we obse regions . Salmonef tumors , suggestf tumors \u201358 whichf tumors ,60. The drolysis , consistde probe . Taken tE. coli, other bacteria such as Salmonella and Shigella also prefer to colonize in the necrotic regions of tumors [Besides f tumors . Thus, df tumors \u201367 or inf tumors  might re"}
+{"text": "AbstractSymplanella Fennah, S. hainanensissp. n., S. recurvatasp. n. and S. zhongtuasp. n., are described and illustrated from South China. A checklist and a key to species of genus Symplanella are provided.Three new species of the Oriental caliscelid planthopper genus  PageBreakSymplanella was erected by Symplanella breviceps Fennah, 1987) and was placed in the subtribe Augilina of the tribe Ommatidiotini of the family Issidae. Recently, the genus was transferred to the family Caliscelidae by Caliscelidae. Symplanella from China and described one new species, Symplanella unipuncta Zhang & Wang, 2009, and proposed one new combination, Symplanella brevicephala  (transferred from Symplana Kirby). To date, only three species, Symplanella brevicephala (China: Yunnan), Symplanella breviceps (Burma: Dawna Hills) and Symplanella unipuncta (China: Hainan), are included in the genus Symplanella.The genus Symplanella are described and illustrated from South China . The generic characteristics are redefined. A checklist and a key to known species of Symplanella are provided.In this paper three new species of the genus Terminology follows Fennah, 1987http://species-id.net/wiki/SymplanellaSymplanella Fennah, 1987: 244; Symplanella breviceps Fennah, 1987, by original designation.PageBreaktriangular, venation as shown in Vertex , 15, 27 Oriental Region (China and Burma).Symplanella brevicephala ; China (Yunnan).Symplanella breviceps Fennah, 1987; Burma (Dawna Hills).Symplanella hainanensis sp. n.; China (Hainan).Symplanella recurvata sp. n.; China (Guangdong and Guangxi).Symplanella unipuncta Zhang & Wang, 2009; China (Hainan).Symplanella zhongtua sp. n.; China (Yunnan)http://zoobank.org/FA3249B6-4106-4928-9201-F444D3E76BEDhttp://species-id.net/wiki/Symplanella_recurvataBody length including forewing: male 5.78\u20135.98 mm (N = 6), female 6.15\u20136.25 mm (N = 12); forewing length: male 4.90\u20135.15 mm (N = 6), female 5.30\u20135.40 mm (N = 12).PageBreakPageBreaksegment. Central area of vertex and pronotum, base of mesonotum with somewhat pale yellowish red. Procoxae, mesocoxae, metapleura, abdominal sternites laterally and pregenital sternite of female fuscous.General color light yellowish brown with somewhat green. Ocelli reddish brown, eyes black brown. Antennae with one black spot at apex of second Vertex including eyes narrower than pronotum (0.86:1). Vertex shorter in middle line than broad at base (0.60:1). Frons 1.28 times longer in middle line than widest part. Pronotum slightly longer in middle line than vertex (1.21:1). Mesonotum 1.24 times as long as vertex and pronotum together in middle line. Forewing longer in middle line than broad at widest part (4.71:1). Hindwing longer in middle line than broad at widest part (2.01:1), venation as shown in Anal segment of male in posterior view  nearly l23\u00b008'N, 113\u00b014'E), on bamboo , 22 Nov. 2006, X.-S. Chen; paratypes: 5 \u2642\u2642, 11 \u2640\u2640, data same as holotype; 1 \u2640, Guangxi, Daxin, Encheng, 4 May 2009, H.-R. Li.Holotype: \u2642, China: Guangdong, Guangzhou, Huanan Botanical Garden .Bamboo (South China (Guangdong and Guangxi) .Symplanella unipuncta Zhang & Wang, 2009 but differs in: i) anal segment in lateral view with one stout process at apical margin ventrally, which curves cephalad apically ; ii) posterior margin of pygofer without process (with one process in Symplanella unipuncta); iii) aedeagus without process at base .This new species is closely related to The new species is named after the strongly recurved tip of the anal tube.http://zoobank.org/C12B4D8F-BF4B-4939-977B-D13F7CEDDF9Fhttp://species-id.net/wiki/Symplanella_hainanensisBody length including forewing: male 5.45\u20135.62 mm (N = 2), female 6.00\u20136.30 mm (N = 8); forewing length: male 4.40 mm (N = 2), female 4.65\u20134.95 mm (N = 8).PageBreakPageBreakGeneral color dirty yellowish brown. Ocelli reddish brown, eyes black brown. Antennae with one black spot at apex of second segment. Frons and clypeus mostly dark brown. Central area of vertex and pronotum, base of mesonotum with somewhat pale yellowish red. Procoxae, mesocoxae, metapleura, abdominal sternites laterally and pregenital sternite of female, fuscous.Vertex including eyes as wide as pronotum. Vertex longer in middle line than broad at base (1.24:1). Frons 1.67 times longer in middle line than widest part. Pronotum shorter in middle line than vertex (0.74:1). Mesonotum 0.87 times as long as vertex and pronotum together in middle line. Forewing longer in middle line than broad at widest part (4.00:1).Anal segment of male in dorsal view  with med18\u00b047'N, 109\u00b052'E), on bamboo, 9\u201312 Apr. 2009, X.-H. Hou; paratypes: 1 \u2642, 8 \u2640\u2640, data same as holotype.Holotype: \u2642, China: Hainan, Diaoluoshan National Natural Reserve (Bamboo.South China (Hainan) .Symplanella breviceps Fennah, 1987, but can be distinguished from the latter in: i) frons mostly dark brown (stramineous in Symplanella breviceps); ii) vertex with anterior margin rounded (angulated in Symplanella breviceps); iii) posterior margin of pygofer with one spinous process dorsally (absent in Symplanella breviceps); iv) genital style in posterior view broad and short (narrow and long in Symplanella breviceps).This new species is similar to the type species from Burma, The new species is named after the type locality, Hainan Province, China.http://zoobank.org/B8FADA9C-2202-4485-928A-E17811BB1A59http://species-id.net/wiki/Symplanella_zhongtuaBody length including forewing: male 6.10\u20136.35 mm (N = 4), female 6.30\u20136.50 mm (N = 2); forewing length: male 5.15\u20135.30 mm (N = 4), female 5.20\u20135.40 mm (N = 2).PageBreakPageBreakGeneral color dirty yellowish brown. Ocelli reddish brown, eyes black brown. Antennae with one black spot at apex of second segment. Frons and clypeus mostly blackish brown. Central area of vertex and pronotum, base of mesonotum with somewhat pale yellowish red. Procoxae, mesocoxae, metapleura, abdominal sternites laterally and pregenital sternite of female fuscous.Vertex including eyes narrower than pronotum (0.98:1). Vertex shorter in middle line than broad at base (0.65:1). Frons 1.41 times longer in middle line than widest part. Pronotum as long in middle line as vertex. Mesonotum 1.45 times as long as vertex and pronotum together in middle line. Forewing longer in middle line than broad at widest part (4.45:1).PageBreakapical third abruptly narrowed, stick-like, apical margin rounded, each side with one spine-like process; phallobase lobe-like, each with one small spine at apex; aedeagus in lateral view , on bamboo, 28 July 2011, W.-B. Zheng and Z.-M. Chang; paratypes: 3 \u2642\u2642, 2 \u2640\u2640, data same as holotype.Holotype: \u2642, China: Yunnan, Xishuangbanna, Menglun (Bamboo.Southwest China (Yunnan) .Symplanella brevicephala , but can be distinguished by: i) frons and clypeus mostly black ; ii) posterior margin of male pygofer having one stout spinous process at middle, directed ventro-caudad ; iii) genital style in lateral view dorso-apical angle broadly rounded ; iv) aedeagal shaft mostly straight .This new species is similar to The name is derived from transliteration of the Chinese \u201czhongtu\u201d, meaning posterior margin of male pygofer having one stout spinous process at middle.Diversity of bamboo-feeding planthoppers. The current authors paid particular attention to the species of bamboo planthopper in field research and collected large quantities of specimens in the past twelve years. A number of new taxa or new records were found and some of them have been published (Bambusoideae (Delphacidae (78 species in 15 genera), Caliscelidae (three species in two genera), Cixiidae (two species in one genus) and Tropiduchidae (one species in one genus). The genus Symplanella with three known species and three new species described in this paper, represents the second bamboo-feeding genus in the tribe Augilini after Pseudosymplanella Che, Zhang & Webb, 2009  and the International Science and Technology Cooperation Program of Guizhou (No. 20107005)."}
+{"text": "A 10-year-old girl developed an enlargement of parotid and submandibular salivary glands and lymph nodes up to 2.5 cm associated with edema of her upper eyelids. In further follow-up, she developed non-palpable, vasculitic skin lesions. She received cephixime, that co-incided with partial regression of lymph nodes. Re-occurrence of vasculitic lesions with bilateral edema of upper eyelids resembling Miculitz disease was observed. At 7 years of age, she developed asthma requiring the use of bronchodilators. Clinical examination: a 10-year-old girl, pale skin, at the both calves irregularly shaped, livid, painless, vasculitic lesions, 2-3 cm in size. The upper eyelids are swollen. Parotid and submandibular salivary glands and submandibular lymph nodes are enlarged.To present clinical and laboratory investigations in IgG4-related disease in childhood.Routine laboratory tests including serum Ig concentrations, autoantibody screen, and histopathologic examination.Serum proteins were 101 g/l, albumins,38 g/l. Serum immunoglobulin IgA 1.92, IgM 0.38, IgG 42.2 g/l; IgG subclasses: IgG1 18.9 g/l (4.32-10.2), IgG2 17.05 g/l (0.72-4.3), IgG3 6.35 g/l (0.13-0.85), IgG4 9.02 g/l (0.02-0.93). Serum IgE 900 IU/ml . Coombs test negative. Autoantibody screening was negative. Serologic tests to HSV, EBV, CMV, hepatitis B and C, HIV were negative. Serum ACE was normal, 57 U/l. Neck ultrasound showed an enlargement of both parotid glands, lymph nodes and submandibular salivary glands up to 2.5 cm.Histopathathologic evaluation of lymph nodes, submandibular and parotid gland: the enclosed lymph node profiles show retained architecture with prominent reactive follicular hyperplasia. The intervening paracortex focally appears prominently hypovascular and contains a polymorphous lymphoid infiltrate comprising small lymphocytes, scattered immunoblasts, some plasma cells and focally prominent eosinophils.The salivary gland tissue shows abundant mononuclear inflammatory infiltrate with focal prominence of plasma cells and significant patchy sclerosis. The immunostainings including CD3, CD20 and CD79a show retained lymph node architecture. There is a reactive pattern of expression of Ki67, CDI0, bcl-2, bCl-G, CD21 and CD23. The immunostains also highlight fragmentation of some of the germinal centres. Most of the IgG staining plasma cells were of IgG4 positive phenotype.Bilateral, symmetrical, painless swelling of their lacrimal and salivary (parotid and submandibular) glands should raise suspicion on IgG4-related disease in childhood.None declared."}
+{"text": "After an acute myocardial infarction with ST-segment elevation (STEMI) treated with percutaneous coronary intervention (PCI), the left ventricle (LV) can undergo negative remodeling (R-). We aimed to investigate whether global longitudinal strain (SGL) of the left ventricle (LV) predicts remodeling.Transthoracic echocardiography with speckle tracking imaging (TTE-STI) was performed 2 to 3 days after primary PCI and 6 months later in patients with diagnosis of STEMI. LV R- criteria were: LVEF increase \u22645% and end-diastolic volume increase \u226515%. Logistic regression and ROC curve analysis was used for the statistical analysis.n = 35, 42%) and no LV R- patients . Diabetes mellitus  and TnI levels  showed higher incidence in LV R- patients. SGL was -12.5 \u00b1 5.6% in no LV R- patients and -6.5 \u00b1 3.4 in LV R- patients. In the regression analysis just LV SGL and SL in left anterior descending territory remained significant, OR: 1.85 (1.24 to 2.76) (P < 0.001) and OR: 1.63 (1.15 to 2.31) (P < 0.001), respectively. The analysis of ROC curves revealed that at the cutoff level of -12.46%, SGL identifies LV R- with a sensibility of 81% and a specificity of 86%   with STEMI at any LV localization and subjected to primary PCI were studied during 2012: LV R- patients (1 years wSGL assessment in the first days after primary PCI is useful in the prediction of LV R- independently of the myocardial infarction localization."}
+{"text": "Erratum: Drug resistance in multiple myeloma: latest findings and new concepts on molecular mechanisms1,2, Guoan Chen3, Hong Chang1,2,4Jahangir Abdi2186-2207Oncotarget. 2013; 4:PMID: 24327604http://www.impactjournals.com/oncotarget/index.php?journal=oncotarget&page=article&op=view&path%5B%5D=1497&path%5B%5D=1760The affiliation of the first author is incorrect in the published paper.1 Division of Immunopharmacology, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The NetherlandsThe corrected affiliation is provided here.1 Dept. of Laboratory Medicine & Pathobiology, University of Toronto, Ontario, Canada"}
+{"text": "Enterobacteriaceae-producing extended-spectrum \u03b2-lactamases (ESBLs). A retrospective case-control study was conducted to identify independent risk factors for ESBL-producing Escherichia coli and Klebsiella pneumoniae in non-hospitalized KTPs with UTI. Forty-nine patients suffering from UTI by ESBL-producing bacteria (ESBL-P) as case group and the same number of patients with UTI by ESBL negative (ESBL-N) as control-group were compared. Clinical data, renal function parameters during UTI episodes, UTI recurrence and relapsing rate, as well as risk factors for recurrence, molecular characterization of isolates and the respective antimicrobial susceptibility profile were evaluated. Diabetes mellitus (p <0.007), previous antibiotic prophylaxis (p=0.017) or therapy (p<0.001), previous UTI (p=0.01), relapsing infection (p=0.019) and patients with delayed graft function after transplant (p=0.001) represented risk factors for infection by ESBL positive Enterobacteriaceae in KTPs. Interestingly, the period of time between data of transplantation and data of UTI was shorter in case of ESBL-P case-group (28.8 months) compared with ESBL-N control-group (50.9 months). ESBL-producing bacteria exhibited higher resistance to fluoroquinolones (p=0.002), trimethoprim-sulfamethoxazole (p<0.001) and gentamicin (p<0.001). Molecular analysis showed that blaCTX-M was the most common ESBL encoding gene (65.3%), although in 55.1% of the cases more than one ESBL gene was found. In 29.4% of K. pneumoniae isolates, three bla-genes (blaCTX-M-blaTEM-blaSHV) were simultaneously detected. Low estimated glomerular filtration rate (p=0.009) was found to be risk factor for UTI recurrence. Over 60% of recurrent UTI episodes were caused by genetically similar strains. UTI by ESBL-producing Enterobacteriaceae in KTPs represent an important clinical challenge regarding not only hospitalized patients but also concerning outpatients.Urinary tract infection (UTI) is a common complication after kidney transplantation, often associated to graft loss and increased healthcare costs. Kidney transplant patients (KTPs) are particularly susceptible to infection by  Urinary tract infections (UTI) affect 5 to 36% of kidney transplant patients (KTPs), being the main infectious complication among such patient population \u20134, and oblaCTX-M [ESBLs are a large, rapidly evolving group of plasmid-enzymes that confer resistance to penicillins, first-, second-, and third generation cephalosporins, and aztreonam. They are inhibited by beta-lactamase inhibitors such as clavulanic acid \u201310. DurilaCTX-M . The prelaCTX-M \u201317. TherlaCTX-M \u201320, howeE. coli and K. pneumoniae comparatively to the respective non ESBL-producing counterparts. ESBLs encoding genes were identified. E. coli and K. pneumoniae isolates sequentially recovered from each patient were genotyped to assess about re-infection or bacterial persistence.The aim of this study was to identify risk factors, the susceptibility profile and recurrence associated to UTI caused by ESBL-producing This study was approved by Comiss\u00e3o de Etica para a Sa\u00fade (CES) do Centro Hospitalar de S\u00e3o Jo\u00e3o , signed by Prof. Manuel Pestana (Director of Nephrology Department). Patient records/information was anonymized and de-identified prior to analysis.E. coli and K. pneumoniae ESBL-producing bacteria (ESBL-P), as case-group, and 49 outpatients with UTI by the same species but ESBL negative (ESBL-N), as a control-group, was performed. All patients were kidney transplant patients (KTPs) from Centro Hospitalar S\u00e3o Jo\u00e3o, Porto  with documented UTI between January 2012 and August 2012.A retrospective case-control study enrolling 49 outpatients with UTI by 2) was registered [Demographic data  and clinical variables such as delayed graft function, the presence of urinary tract obstruction, bacteremia, fungemia, previous urinary tract infection, creatinine blood level (reference range 0.6\u20131.1 mg/dL), and the percentage of patients with low estimated glomerular filtration rate (eGFR) during UTI  and K. pneumoniae (n = 32) were recovered from 49 KTPs (case-group): 25 patients yielded each a single isolate, 16 patients two isolates each, 7 patients three isolates each, one single patient yielding four isolates. Forty-nine ESBL negative isolates (32 E. coli and 17 K. pneumoniae) were recovered from the 49 patients of control group. All isolates were analyzed by Vitek2 System  using GN card for identification and the AST-60 for antimicrobial susceptibility profile. ESBL presence was confirmed by disc diffusion method, according to the recommendations of Clinical Laboratory Standard Institute [Escherichia coli ATCC 25922 and Klebsiella pneumoniae ATCC700603 type strains, as recommended by CLSI protocol [Eighty-two isolates of ESBL-producing nstitute ; a doublprotocol  and plasblaCTX-M,blaTEM, and blaSHV) were screened by PCR multiplex, as previously described [blaSHV: 5\u00b4-ATCCACTATCGCCAGCAGG-3\u00b4 and 5\u00b4-TCATTCAGTTCCGTTTCCCAG-3\u00b4; blaTEM: 5\u00b4-GAGTATTCAACATTTCCGTGTC-3\u00b4 and 5\u00b4-GGGCGAAAACTCTCAAGGATC-3\u00b4; blaCTX-M: 5\u00b4-GTTGTTAGGAAGTGTGCCGC-3\u00b4 and 5\u00b4-GCCCGAGGTGAAGTGGTATC-3\u00b4. Primers amplified the internal fragments of ESBLs genes possessing different sizes. Multiplex PCR was performed in a 25 \u03bcl reaction mixture containing 1x Dream Taq buffer (Fermentas), 2.5 mM MgCl2, 10 mM dNTPs, 2.5\u201340 pmol of specific-group primers, 50\u2013150 ng plasmid DNA and 1 U of Dream Taq Polymerase (Fermentas). Amplification reactions were carried out in Mastercycler Realplex2 (Eppendorf) under the following conditions: initial denaturation at 95\u00b0C for 2 min, followed by 30 cycles of denaturation (95\u00b0C for 30 sec), annealing (59\u00b0C for 30 sec), extension (72\u00b0 for 30 sec) and a final extension step (72\u00b0C for 10 min). PCR products were run at 80 V, during 1 hour in a 2.5% agarose gel containing 0.002% Ethidium Bromide . ESBLs bacteria exhibiting well-characterized ESBL genes kindly gift by Dr. Rafael Cant\u00f3n  were used as positive controls.The most frequently ESBL genes , and involved the following steps: initial denaturation at 94\u00b0C for 1 min, followed by 45 cycles of denaturation (94\u00b0C at 1 min), annealing (34\u00b0C at 1 min), extension (72\u00b0C at 2 min), and a final extension step (72\u00b0C at 10 min). PCR products were separated by electrophoresis (60 V for 3 hours) in a 2.5% agarose gel. DNA fingerprints were compared by visual inspection with the Molecular Imager ChemiDoc XRS (BioRad). Clonality was assessed by visual inspection of the different fragments obtained, in a range of 2000\u2013200 bp for E. coli and 3000\u2013250 bp for K. pneumoniae. According to Wong et al. [Sequential isolates from case-group (ESBL-P)- 2 or more isolates (24 patients)\u2014were genotyped by RAPD. Distinct primers were used for E. coli   version 20. Continuous variables were summarized as mean value and standard deviation (SD). Categorical variables, summarized as percentages, were compared using Chi-square test (or Fishers \u2018exact test whenever was necessary) and Student\u00b4s Patient clinical data, including demographic characteristics, co-morbidities, clinical laboratory data, immunosuppressive therapeutic regimen, previous UTI and antibiotic exposure are summarized in Regarding immunosuppression, all patients showed adequate therapeutic levels; mean values were: 8.1 ng/mL for tacrolimus, 190.5 ng/mL for cyclosporine and 6.28 ng/mL for everolimus. Patient death occurred solely among ESBL-P group, during the 3 months after infection . No bactK. pneumoniae susceptibility to amikacin between both study groups  were detected by PCR multiplex in isolates from ESBL-P group (blaCTX-M (65.3%), either alone (18.4%) or in combination with another ESBL gene (46.9%), followed by blaTEM (61.2%) and blaSHV (40.8%). More than half of the case- group express more than one ESBL gene, being the association blaCTX-M- blaTEM the most frequent (28.5%), particularly among E. coli strains (37.5%). Among K. pneumoniae isolates, blaCTX-M-blaTEM-blaSHV (29.4%) was the most frequent association, followed by blaCTX-M-blaSHV (23.5%). Strains expressing a single ESBL gene, blaCTX-M (25%) and blaTEM (18.8%) corresponded most often to E. coli, while blaSHV was more common among K. pneumoniae isolates (17.6%) (ESBL genes (-P group . Interes (17.6%) . No corrE. coli and 63.6% in the case of K. pneumoniae) (see representative examples in Although the level of recurrence found for both groups was similar (around 40%), the relapsing percentage was significantly higher on control-group . The 48 Kidney transplant recipients display many risk factors for UTI and are considered a particular vulnerable population to such infections . Severalet al. study [blaCTX-M,blaTEM, and blaSHV) in E. coli and K. pneumoniae revealed that blaCTX-M was the most common ESBL gene. A higher predominance of blaCTX-M enzyme among ESBL positive E. coli was reported in Greece [blaCTX-M and blaTEM [E. coli and K. pneumoniae (57.3%) isolates exhibiting two or three genes was previously reported [blaSHV and blaTEM types of ESBLs have traditionally been responsible for serious health care related infections [blaCTX-M types have been mainly associated with community-onset UTI [blaCTX-M-producing bacteria and resistance to fluoroquinolones [Similar to Linares study  impairmel. study . Remarkan Greece . Conversn Greece . In Port blaTEM ,36. Repo blaTEM . A high reported , similarfections  while blnset UTI . The epinset UTI , being tinolones  such corE. coli and K. pneumoniae. Molecular epidemiology showed that blaCTX-M was the most common ESBL encoding gene, either alone or in association with other genes. Low eGFR and high blood creatinine are risk factors for UTI recurrence. The high co-resistance to other antibiotics (non-\u03b2-lactams) found from ESBL producing bacteria in UTI from KTPs, remains a serious clinical challenge.In conclusion, we have demonstrated that delayed graft function, diabetes mellitus, previous antibiotic exposure, antibiotic prophylaxis and relapsing UTI are independent risks factors for acquiring infections by ESBL-producing"}
+{"text": "Their structures and relative configurations were established by extensive spectroscopic analysis. A possible biosynthetic pathway for 1\u20134 was also proposed.Guajadials C-F (Supplementary material is available for this article at 10.1007/s13659-012-0102-4 and is accessible for authorized users. Supplementary material, approximately 2.05 MB."}
+{"text": "The correct citation is: Seed Ahmed M, Pelletier J, Leumann H, Gu HF, \u00d6stenson C-G (2015) Expression of Protein Kinase C Isoforms in Pancreatic Islets and Liver of Male Goto-Kakizaki Rats, a Model of Type 2 Diabetes. PLoS ONE 10(9): e0135781. doi:"}
+{"text": "MEDI4736 is an engineered human IgG1 that blocks PD-L1 binding to PD-1 and allows T-cells to recognize and kill tumor. MEDI4736 has single-agent activity and potential for further increased activity in combination. A comprehensive development programme is underway in NSCLC, as monotherapy and in combination.NSCLC data from 2 multicentre, open-label Phase I studies are reported. NCT01693562 evaluates safety and efficacy of MEDI4736 administered every 2 or 3 weeks. NCT02000947 evaluates safety and efficacy of MEDI4736 in combination with tremelimumab, a human IgG2 anti-CTLA-4 mAb, at 4-week intervals.NCT01693562: As of May 2014, NSCLC cohort included 155 patients (pts) . Median follow-up: 6 weeks (range 0-67). Treatment-related adverse events (TRAEs): 29% (Grade [Gr] \u22653: 3%); none led to treatment discontinuation. Most frequent TRAEs: fatigue (7%), nausea (5%), and vomiting (5%). No treatment-related colitis any Gr. No treatment-related Gr 3/4 pneumonitis or dyspnea. 58 pts had \u226512 weeks follow-up: 16% had partial response (as early as 6 weeks), duration of response ranged 5-54+ weeks, disease control rate 35%. PD-L1 positivity appears to enrich for response.NCT02000947: As of April, 2014, 12 pts treated at 4 dose-levels . No DLTs observed in any cohort during DLT observation period. Most frequent TRAEs: -\u2191amylase, abdominal pain, arthralgia, colitis, diarrhea, epigastric discomfort, fatigue and nausea. TRAEs \u2265Gr 3 noted in 3 pts: Gr 3 - \u2191AST/ALT & Gr 5 myasthenia (MG) (n = 1), Gr 3 diarrhea/colitis (n = 1), Gr 4 - \u2191amylase (n = 1). TRAEs led to discontinuation in two subjects: Gr 5 MG and Gr3 colitis. In 12 response-evaluable pts  support continued development in NSCLC. Additional monotherapy NSCLC studies: Phase II 'ATLANTIC' (NCT02087423), Phase III 'PACIFIC' following chemo-radiotherapy (NCT02125461), Combination: Phase I + gefitinib (NCT02088112), and Phase Ib + AZD9291 (NCT02143466). Monotherapy and combination: Phase III 'ARCTIC' vs standard-of-care."}
+{"text": "AbstractOphioninae is updated, based on the examination of about 800\u2013900 individuals in the South African and European museum collections. A robust interactive matrix key was built to provide quick and reliable identifications. The key is available online at http://www.waspweb.org. Two new species are described: Dicamptusmaxipolsp. n. and Enicospilusgauldetmitchellorumsp. n. Numerous new distribution and biological records are provided, and noticeable morphological intra-specific variations are detailed. Enicospilusbatus Gauld & Mitchell, syn. n. is considered as a junior synonym of Enicospilusluebberti (Enderlein).The revision of the Afrotropical  Ophioninae is one of the two major subfamilies of Ichneumonidae that have been extensively revised in the Afrotropical region. The revision of the Ophioninae is mainly due to The subfamily Ophioninae in the region.It is of note that very few amendments have been brought to their work since the revision was published, except three new species descriptions  and some Natural History Museum, London, UK (Gavin Broad).BMNH California Academy of Science, San Fransisco, USA (Brian Fisher).CASC Mus\u00e9um d\u2019Histoire Naturelle de La R\u00e9union, Saint Denis, France (Sonia Ribes).MHNR Mus\u00e9ul National d\u2019Histoire Naturelle, Paris, France (Claire Villemant).MNHN Mus\u00e9um Royal de l\u2019Afrique Centrale, Tervueren, Belgium (Eliane de Coninck).MRAC KwaZulu-Natal Museum, Pietermaritzburg, South Africa (Burgert Muller).NMSA Iziko South African Museum, Cape Town, South Africa (Simon van Noort).SAMChttp://www.waspweb.org.Specimens were point mounted on black, acid-free card for examination (using a Leica M205C stereomicroscope with LED light source), photography and long term preservation. Images were acquired using the Leica LAS 4.4 imaging system, which comprised a Leica\u00ae Z16 microscope with a Leica DFC450 Camera with 0.63\u00d7 video objective attached. The imaging process, using an automated Z-stepper, was managed using the Leica Application Suite V 4.4 software installed on a desktop computer. Lighting was achieved using techniques summarized in PageBreakThe terminology follows : body length, from torulus base to apex of metasoma (mm).B: fore wing length, from tegula base to wing apex (mm).F : shortest distance between eye and mandible / basal mandibular width.ML : distance between outer edges of tentorial pits / median height of clypeus.CT (post-ocellar line index): shortest distance between posterior ocelli / posterior ocellus longest diameter.POL (oculo-ocellar line index): shortest distance between eye and posterior ocellus / posterior ocellus longest diameter.OOL : length of flagellomere 1 (annellus excluded) / length of flagellomere 2.Fl1\u20132th flagellomere): length / width of flagellomere 20. .Taking into account the taxonomic updates post Gauld and Mitchell\u2019s revision, including the present one, we acknowledge here a total of 194 species of hioninae . A few ihioninae . Finallyhttp://www.waspweb.org.The matrix includes these 194 species and their known intra-specific variability. Furthermore, the dichotomous key provided in Taxon classificationAnimaliaHymenopteraIchneumonidaeRousse & van Noortsp. n.http://zoobank.org/8C1C347B-4AD0-4FB2-A126-242C3FEA947C(verbatim label data). HOLOTYPE \u2640: SOUTH AFRICA, W. Cape, West Coast Fossil Park, (5.5 km 270\u00b0 W Langebaanweg) 32\u00b057.759'S, 18\u00b005.519'E, 9\u201316 Oct 2002, S. van Noort, Malaise trap LW02-R4-M96, Rehabilitated slimes dam, SAM-HYM-P049469 (SAMC).Orange with inter-ocellar area, most of mesosoma and apex of metasoma black; mandible not twisted, with a central tuft of hairs; clypeus wide, long and flat in profile; antenna short and stout with 56 flagellomeres; mesosoma laterally coarsely punctate to rugose-punctate, dorsally densely and more finely punctate; mesoscutum with notaulus distinct and relatively long; mesopleuron with epicnemial carina not distinct above lower corner of pronotum; propodeum anteriorly densely punctate, posteriorly coarsely rugose-reticulate; disco-submarginal cell with fenestra developed but without distinct sclerite; fore tibia with dense and long spines on outer surface; fore tibial spur with a vestigial basal membrane.Dicamptus species in the world by the absence of distinct sclerites in the disco-submarginal cell; in the Afrotropical region, it seems related to Dicamptusneavei Gauld & Mitchell, 1978, which shares the dense spines on the tibia, the exceptionally reduced ocelli and a somewhat similar colour pattern; Dicamptusneavi is, however, a tropical species with shorter antennae, a stouter metasoma, and distinctly different alar indices with a distinct proximal sclerite in the disco-submarginal cell. In Gauld and Mitchell\u2019s key . B 20.8; F 11.5; ML 1.2; CT 1.2; OOL 2.0; POL 1.2; FI 20%; FColor. Orange interspersed with black; black: inter-ocellar area, entire mesosoma except for mesonotum and metanotum, base of tergite 1, tergite 5 and following, all coxae and trochanters except trochantelli; antenna orange, slightly darkening toward apex; wings hyaline, venation dark reddish to black except for pterostigma anteriorly light reddish.Head. Mandible short and stout, without longitudinal groove, with a central tuft of long hairs, upper tooth barely longer than lower tooth; malar line long; clypeus long and wide, coarsely and densely punctate, rather flat in profile, somewhat swollen medially and ventrally, ventral margin strongly impressed; face strongly transverse, densely and coarsely punctate; frons rather smooth, upper head densely punctate; gena moderately swollen behind eyes; occipital carina complete and strong; antenna short and stout with 56 flagellomeres.PageBreakMesosoma. Pronotum, mesopleuron and metapleuron coarsely and densely punctate, fading to rugose-punctate ventrally; anterior margin of pronotum simple; epicnemial carina short, indistinct above lower corner of pronotum; posterior transverse carina of mesosternum complete though ventrally weak; submetapleural carina not expanded anteriorly; mesoscutum densely and more finely punctate; notaulus long, moderate, distinct to anterior third of mesoscutum; scutellum densely punctate, carinate almost to apex; propodeum with anterior area densely punctate, anterior transverse carina complete, posterior area coarsely rugose-reticulate, abruptly declivous in profile and mid-posteriorly concave. Wings. Disco-submarginal cell with fenestra developed, without any distinct sclerite except a weak quadra centrally; Rs+2r hardly sinuate, slightly bent and thickened near pterostigma; Rs&M distal to cu-a by about its own width; hind wing with 7 hamuli. Legs. Fore tibia with dense and long spines on outer surface; fore tibial spur with a vestigial membrane basally to macrotrichial comb, membrane barely less than 0.1\u00d7 length of spur; hind coxa in profile 1.8\u00d7 as long as high; hind trochantellus mid-dorsally 0.2\u00d7 as long as wide; hind tarsal claws symmetrical with 8 pectinae.Metasoma. Slender; tergite 2 in profile 2.7\u00d7 longer than high; thyridium large, oval, separated from anterior margin of tergite 2 by 1.3\u00d7 its own length; ovipositor not reaching beyond metasomal apex.MALE. Unknown.Named after the unusually reduced ocelli, and as a result the large POL. Noun in apposition.South Africa (Western Cape).Taxon classificationAnimaliaHymenopteraIchneumonidaeRousse & van Noortsp. n.http://zoobank.org/5F861712-DBF0-41CC-9854-00DEB2913E86(verbatim label data).HOLOTYPE \u2640: Tanzania, Mkomazi Game Reserve, Ibaya Camp, 3.58S 37.48E, 18 April 1996, light trap, S. van Noort, open Combretum bushland, SAM-HYM-P015183 (SAMC).PageBreakYellow orange overall, head paler yellow; mandible with upper tooth distinctly longer than lower tooth; clypeus hardly convex in profile, its ventral margin barely concave and in-turned; occipital carinae complete; gena moderately swollen behind eye; ocelli moderately enlarged; antenna with 56 flagellomeres; pronotum unspecialized; mesopleuron and metapleuron closely and deeply punctate; epicnemial carina laterally indistinct; posterior transverse carina of mesosternum complete and noticeably strong; submetapleural carina slightly broadened anteriorly; notaulus vestigial; propodeum basally punctate, posteriorly coarsely and concentrically striate; fore wing without any sclerite in disco-submarginal cell; fore tibia with dense spines on outer surface; thyridium very shallow.Enicospilus in Afrotropical, Oriental and Australasian regions by the combination of the absence of alar sclerites and the dense spines on fore tibia. The swollen genae and the wing venation make it somewhat related to Enicospilusleucocotis, but this latter is strongly larger, with only sparse spines on tibia and slenderer antenna. In Gauld and Mitchell\u2019s key . B 18.8; F 11.5; ML 0.3; CT 1.6; OOL 0.1; POL 0.4; FI 50%; FColor. Yellowish orange overall with face and orbits paler yellow and apex of metasoma slightly infuscate.Head. Mandible basally constricted, apically parallel-sided and slightly twisted, with upper tooth distinctly longer than lower tooth (greatly worn by abrasion in holotype); outer mandibular surface sparsely setose, without longitudinal groove; labrum 0.3\u00d7 as long as wide; clypeus in profile hardly convex, its ventral margin barely concave and in-turned; clypeus and face finely and moderately densely punctate; gena moderately swollen behind eye; occipital carina complete; ocelli slightly enlarged; antenna with 56 flagellomeres.PageBreakMesosoma. Pronotum mid-dorsally long, anterior margin simple; mesoscutum densely punctate, notaulus vestigial; scuto-scutellar groove smooth; scutellum densely and shallowly punctate, barely longer than basally wide, carinate to near its apex; mesopleuron and metapleuron closely and deeply punctate, punctures arranged longitudinally but without distinct striation; epicnemial carina short, indistinct above lower corner of pronotum; submetapleural carina weakly broadened anteriorly; posterior transverse carina of mesosternum complete and strong; propodeum with anterior area finely, shallowly and densely punctate, anterior transverse carina complete, posterior area coarsely and concentrically striate Wings. Disco-submarginal cell with fenestra developed, without any distinct sclerite; Rs+2r sinuate; cu-a basal to Rs&M by 0.3\u00d7 cu-a length; hind wing with 6 hamuli and 1A basally straight. Legs. Fore tibia with numerous dense and long spines on outer surface, basally separated by far less than their own length; hind coxa elongate, in profile 2.4\u00d7 as long as high; hind trochantellus mid-dorsally 0.2\u00d7 as long as wide, its apical margin simple; hind tarsal claws symmetrical with 8 pectinae, pectinae long and acute.Metasoma. Slender; tergite 2 in profile 3.2\u00d7 longer than high; thyridium very shallow, elongate, separated from anterior margin of tergite 2 by 1.7\u00d7 its own length; ovipositor acute not reaching beyond metasomal apex.PageBreakMALE. Unknown.Enicospilusleucocotis. Let give Gauld what belongs to Gauld (updated after Mark 12:17).This species was probably mentioned in Tanzania.http://isodp.hof-university.de/fuzzyg/query/ and Google Earth http://www.google.com/earth/Provided are the verbatim label data. Only unambiguous identifications are listed. All geographical coordinates are also available on a separate file as Suppl. material Dicamptuspulchellus . The Gambia: 1\u2640 Kombo Nth district, Bilijo Forest Park, xi.1992, M. S\u00f6derlung coll., SAM-HYM-P049471 (SAMC).Euryophionlatipennis . South Africa: 1\u2640 Kwazulu-Natal, Itala Game Reserve, xii.1992, S. van Noort coll., SAM-HYM-P044187 (SAMC); 1\u2640 1\u2642 same label data except: xii.1999, SAM-HYM-P044163 and SAM-HYM-P044185 (SAMC); Zambia: 1 specimen [apex of metasoma lacking] Southern Province, Choma Nansa farm xii.1993, A.J. Gardiner coll., SAM-HYM-P044072 (SAMC).Laticoleuspalpalis Gauld & Mitchell, 1978. Kenya: 1\u2640 Eastern Province, Kenplains, x.1984, C.F. Dewhurst coll. (BMNH).Laticoleusunicolor . Botswana: 1\u2640 Xugana , xi.1979, B.H. Lamoral coll., SAM-HYM-P049474 (SAMC).Lepiscelusdistans . South Africa: 1\u2640 Kwazulu-Natal, Itala Game Reserve, xii.1999, S. van Noort coll, SAM-HYM-P044186 (SAMC); 1\u2640 Limpopo, junction Crocodile and Marico Rivers, ii.1918, R. Tucker coll., SAM-HYM-P006194 (SAMC); 2\u2642\u2642 Mpumalanga, Nelspruit, i.1939, R.F. Lawrence coll., SAM-HYM-P006193 (SAMC); Zimbabwe: 1\u2642 Essexvale, ii.1963, SAM-HYM-P006228 (SAMC).Skiapuscoalescens . The Gambia: 1\u2640 Kombo Nth district, Bilijo Forest Park, xi.1992, M. S\u00f6derlung coll., SAM-HYM-P049477 (SAMC).Enicospilusalbiger . Zambia: 1\u2642 South Luangwa near. Mfuwe, xii.2011, A. Gumovsky coll., SAM-HYM-P049484 (SAMC).Enicospilusbabaulti . Malawi: 1\u2640 Nyika National Park, Juniper forest, ix.1999, R.J. Murphy coll., SAM-HYM-P021341 (SAMC); South Africa: 1 specimen [metasoma lacking] ii.1917, C.J. Swierstra coll., SAM-HYM-P001398 (SAMC); Zimbabwe: 1\u2640 Chirinda forest, xi.1955, SAM-HYM-P006247 (SAMC).Enicospilusbebelus Gauld & Mitchell, 1978. Gabon: 1\u2640 Province Ogoov\u00e9\u2013Maritime, R\u00e9serve des Monts Doudou, iii.2000, S. van Noort coll., SAM-HYM-P041707 (SAMC).PageBreakEnicospilusbetanimenus . Ethiopia: 2\u2640\u2640 Adola, xi.1941, SAM-HYM-P047374 and SAM-HYM-P006253 (SAMC); Zimbabwe: 2\u2640\u2640 Bulawayo ii.1971, D.K.B. Wheeler coll, SAM-HYM-P006286 (SAMC).Enicospilusbicoloratus Cameron, 1912. Zimbabwe: 1\u2640 Matopos, ii.1963, SAM-HYM-P006265 (SAMC).Enicospiluscamerunensis . Mayotte: 1\u2640 Demb\u00e9ni, iii.2013, G. Cazenove coll. (MHNR).Enicospilusdivisus . Uganda: 1\u2642 Kibale National Park, Kanyawara, viii.2008, S.van Noort coll., SAM-HYM-P049506 (SAMC).Enicospilusdrakensbergi Gauld & Mitchell, 1978. Tanzania: 1\u2642 South Pare Mountains, alt. c. 1700m, xi.1995, S. van Noort coll., SAM-HYM-P014698 (SAMC).Enicospilusequatus Gauld & Mitchell, 1978. Central African Republic: 2\u2640\u2640 1\u2642 Pr\u00e9fecture Sangha-Mba\u00e9r\u00e9, R\u00e9serve Sp\u00e9ciale de For\u00eat Dense de Dzanga-Sangha, v.2001, S. van Noort coll., SAM-HYM-P049510\u2013P049512 (SAMC).Enicospilusfinalis Gauld & Mitchell, 1978. Central African Republic: 5\u2640\u2640 Pr\u00e9fecture Sangha-Mba\u00e9r\u00e9, Parc National de Dzanga-Ndoki, v.2001, S. van Noort coll., SAM-HYM-P049514 \u2013P049517 (SAMC); Mozambique: 1 specimen [apex of metasoma broken] Mt Gorongoza, ix.1957, SAM-HYM-P006229 (SAMC).Enicospilusoculator Seyrig, 1935. Zimbabwe 1\u2640 Tuli, v.1959, SAM-HYM-P006232 (SAMC).Enicospilushova Gauld & Mitchell, 1978. Uganda: 1\u2642 Kibale National Park, Kanyawara, viii.2008, S.van Noort coll., SAM-HYM-P049513 (SAMC).Enicospilusluebberti . 4\u2640\u2640 Botswana: 1\u2640 Xugana , xi.1979, B.H. Lamoral coll. (NMSA).Enicospilusmamatsus Gauld & Mitchell, 1978. South Africa: 1\u2640 Northern Cape, Sterboom farm, 1599 m, v\u2013vii 2010, S. van Noort, SAM-HYM-P054077 (SAMC).Enicospilusmnous Gauld & Mitchell, 1978. Tanzania: 3\u2640\u2640, Mkomazi Game Reserve, xi.1995, H.G. Robertson coll. and S. van Noort colls, SAM-HYM-P014159, SAM-HYM-P014161 and SAM-HYM-P014170 (SAMC); 2\u2640\u2640 same label data except: iv.1996, S. van Noort coll., SAM-HYM-P014156 and SAM-HYM-P014706 (SAMC).Enicospilusnesius Gauld & Mitchell, 1978. Central African Republic: 1\u2640 Pr\u00e9fecture Sangha-Mba\u00e9r\u00e9, Parc National de Dzanga-Ndoki, v.2001, S. van Noort coll., SAM-HYM-P054079 (SAMC).Enicospiluspallidus . Tanzania: 8\u2640\u2640, Mkomazi Game Reserve, xi\u2013xii.1995 and iv.1996, S. van Noort coll., SAM-HYM-P014157\u2013P0141578, SAM-HYM-P014171\u2013P014175 and SAM-HYM-P015200 (SAMC).Enicospiluspolemus Gauld, 1982. South Africa: 1\u2640 Kwazulu-Natal, Itala Game Reserve, xii.1999, S. van Noort coll., SAM-HYM-P044207 (SAMC); Tanzania: 1\u2640 Mkomazi Game Reserve, iv\u2013v.1996, S. van Noort coll., SAM-HYM-P015666 (SAMC).PageBreakEnicospilusquietus . Namibia: 1\u2640 Namib-Naukluft Park, x.1997, S. van Noort coll., SAM-HYM-P020721 (SAMC); 1\u2642 Otavi, xii.1918, R.M. Lightfoot coll., SAM-HYM-P006278 (SAMC); 1\u2640 Ondangua Ovamboland, 1921, K.H. Bernard coll., SAM-HYM-P006199 (SAMC); 2 specimens [metasomas lacking] Otjiperongo, i.1931, J.S. Brown coll., SAM-HYM-P047375 (SAMC).Enicospilusrubens . Madagascar: 1\u2640 Majunga Province, Maintirano District, iii.2008, M.Irwin and R.Harin\u2019Hala colls, MG-44-26 (CASC).Enicospilusrundiensis Bischoff, 1915. Namibia: 1\u2640 1\u2642 Kaross, 1925, SAM-HYM-P001381 (SAMC); 1\u2640 Warmbad, 1925, SAM-HYM-P001382 (SAMC); 1 specimen [apex of metasoma broken] Narubis, 1921, K.H. Barnard coll., SAM-HYM-P006277 (SAMC); 1 specimen [metasoma lacking] Otjiperongo, i.1931 J.S. Brown coll., SAM-HYM-P006276 (SAMC); Zimbabwe: 1\u2640 Harare, vi.1961, SAM-HYM-P006225 (SAMC).Enicospilusruscus Gauld & Mitchell, 1978. Kenya: 1\u2642 Nguruman, vii.2008, S. van Noort coll., SAM-HYM-P054106 (SAMC).Enicospilusbetanimenus . 2\u2640\u2640 from Zimbabwe SAM-HYM-P006286 (SAMC) ex Achaeacatella Guen\u00e9e, 1852 (Lepidoptera: Noctuidae).Enicospilusdubius . 2\u2640\u2640 from South Africa (SAMC SAM-HYM-P001508) ex Ctenoplusialimbirina (Guen\u00e9e) (Lepidoptera: Noctuidae).Enicospilusdolosus . 1\u2640 from Reunion  ex Anomisflava (Fabricius) (Lepidoptera: Noctuidae).Enicospilusleucocotis . 2\u2640\u2640 from South Africa (SAMC SAM-HYM-P046967 and SAM-HYM-P046968) ex Mesocelismontana (H\u00fcbner) (Lepidoptera: Lasiocampidae).Enicospilusmauritii . 1\u2642 from Reunion ex Callopistriamaillardimaillardi (Guen\u00e9e) (Lepidoptera: Noctuidae) feeding on Dryopterisbernieri (Pteridophyta: Dryopteridaceae) (idaceae) .Enicospilusluebberti . 1\u2642 from South Africa (SAMC SAM-HYM-P006196) ex Graphaniaatavistis (Hampson) (Lepidoptera: Noctuidae).Enicospilusbebelus Gauld & Mitchell, 1978. 1\u2642 from Central African Republic (SAMC SAM-HYM-P049492) and 1\u2640 from Gabon (SAMC SAM-HYM-P041707) with mesosoma interspersed with dark testaceous and black markings, and tergite 1 basally black; otherwise similar to original description.Enicospilusoculator Seyrig, 1935. 1\u2640 from Zimbabwe (SAMC SAM-HYM-P006232) with central sclerite totally absent, upper tooth twice as long as lower tooth, and numeric indices slightly different: FI 80%, AI 1.0, fore wing length 14 mm. Otherwise similar to PageBreakEnicospilusgrandiflavus Townes, 1973. 1\u2640 from South Africa (SAMC SAM-HYM-P049521) with entire head strongly darkened, nearly black. Otherwise similar to Enicospilusexpeditus . 1\u2640 1\u2642 from South Africa (SAMC SAM-HYM-P054068) with hind tarsal claws less pectinate than figured in Enicospilusluebberti . Numerous specimens from South Africa , Bostwana (NMSA), and Kenya (BMNH) showed the following non-correlated variations: inter-ocellar area and metasomal apex yellowish-orange to totally black, antenna with 48\u201362 flagellomeres, longitudinal groove on mandible more or less impressed, proximal sclerite more or less elongate and central sclerite variously sclerotized. These variations encompass the definition of Enicospilusbatus Gauld & Mitchell, 1978, syn. n., described on a single specimen, which is hereby recognized as a junior synonym of Enicospilusluebberti."}
+{"text": "Allergic rhinitis with/without conjunctivitis (AR/C) sufferers often rely on pharmacotherapy to relieve symptoms. Although the main goal of immunotherapy is long-term disease modification, reducing or eliminating the need for pharmacotherapy is also an important and desirable treatment goal.Ambrosia artemisiifolia; Merck/ALK-Abell\u00f3). Subjects with ragweed-pollen\u2013induced AR/C were randomized ~16 weeks before the 2010 pollen season to once-daily MK-3641  or placebo. During the trial, all subjects, whether taking MK-3641 or placebo, could use AR/C rescue medication, including oral/ocular antihistamines and intranasal/oral corticosteroids. We examined rescue medication use in all groups.Data were pooled from two trials that evaluated the efficacy and safety of short-ragweed sublingual immunotherapy tablet (SLIT-T), MK-3641  subjects receiving MK-3641 12 Amb a 1-U and 144 of 324 (44.4%) subjects receiving 6 Amb a 1-U used no rescue medication over the entire ragweed season, compared with 109 of 340 (32.1%) subjects receiving placebo. These differences represented 56% and 38% improvements over placebo. Similarly, during the peak ragweed season 173 of 311 (55.6%) subjects and 161 of 317 (50.8%) subjects in the 12 Amb a 1-U and 6 Amb 1-U groups, respectively, reported no rescue medication use, in contrast to 136 of 333 (40.8%) subjects receiving placebo. Fewer subjects taking 12 and 6 Amb a 1-U  used oral antihistamine than those taking placebo; 35% and 28% fewer subjects used ocular antihistamine; and 43% and 27% fewer subjects used intranasal corticosteroid .Compared with placebo, the SLIT-T treatment MK-3641 reduced rescue-medication use among subjects with ragweed-pollen\u2013induced AR/C.ClinicalTrials.gov Identifiers: NCT00783198; NCT00770315"}
+{"text": "Mevalonate kinase deficiency (MKD) is a rare autoinflammatory, autosomal-recessive defect on MVK gene. Clinical spectrum ranges from recurring febrile attacks to malformations and neurologic disorders. Gastrointestinal symptoms are cardinal. Severe gastrointestinal involvement has been described at the onset.To analyse severe gastrointestinal events (SGE) complicating MKD.Retrospective observational French cohort of MKD patients. SGE were defined as complicated inflammatory involvement, requiring an abdominal surgery and/or enteral/parenteral nutrition. Data were collected from clinical charts provided by the members of the Francophone Society for Paediatric Rheumatism and Inflammatory Diseases (SOFREMIP).25=13.7; P75=55.5); median MK activity: 2.2% . The significant co-morbidities found in SGE patients in comparison with the global cohort were: failure-to-thrive in 85.7% (6/7), pulmonary diseases in 37.5% (3/8) and feeding disorders in 28.6% (2/7) (p<0.05).From a 53-patient cohort, nine presented a SGE (17%). From these, disease onset median age was 1.0 months (0-12); one patient deceased (22 months) from a non-gastrointestinal event. Compound heterozygote mutations were found in 7/8, being Val377Ile the commonest (6/8). The main symptoms during febrile attacks were: diarrhoea , lymphadenopathy , skin lesions, joint pain , aphtous ulcers, abdominal pain , splenomegaly , hepatomegaly  and vomiting . Median mevalonic aciduria: 23.05 mmol/mol of creatinine , representing 43% (3/7) of patients with severe gastrointestinal disease: abdominal adhesions  and colitis/enterocolitis  were mainly found. 87.5% (7/8) needed surgery and 44.4% (4/9) required enteral/parenteral nutrition. Despite digestive resection, disease progression remained; two patients needed re-intervention due to surgical complications. Aphtous/ulcerative damage was the main endoscopic feature . The most consistent microscopic finding was lymphocytic infiltrates. IL-1 antagonists were the most used/effective treatment (4/9), resulting in with complete remission in all three patients with data available.MKD severe gastrointestinal involvement presentation has a non-negligible frequency. It usually appears as an aphtous/ulcerative disease involving any part of the digestive tract or as abdominal adhesions, frequently requiring surgery. The treatment with IL-1 antagonists resulted in complete remission in a majority of treated patients. Thus, MKD should be added to the list of monogenic early-onset inflammatory bowel disease."}
+{"text": "Lactobacillus brevis BSO 464 was sequenced and assembly produced a chromosome and eight plasmids. This bacterium tolerates dissolved CO2/pressure and can rapidly spoil packaged beer. This genome is useful for analyzing the genetics associated with beer spoilage by lactic acid bacteria.The genome of brewery-isolate  Lactobacillus brevis BSO 464  can resist the headspace pressure and dissolved CO2 in packaged beer, making it a virulent beer-spoiler (\u2013Brewery isolate -spoiler \u20133. Lb464De Novo assembler v2.5.3. Hawkeye (xl platform).Sequencing was performed using the Roche 454 FLX platform at the National Research Council Plant Biotechnology Institute . Two separate runs yielded 330,771 unpaired and 567,735 paired reads for ~30\u00d7 coverage. Reads were assembled using Newbler GS L.\u00a0brevis KB290  with nine plasmids .The Lb464 genome was assembled into a 2,503,991-bp circular chromosome (G+C content 45.7%) that had 8,461\u00a0bp cumulative of gaps due to transposase or repetitive regions not allowing PCR-based sequencing. Additionally, eight plasmids were assembled (G+C content ranged from 39.1% to 42.4%): pLb464-1 , pLb464-2 , pLb464-3 , pLb464-4 , pLb464-5 , pLb464-6 (5018 bp), Lb464-7 (2353 bp), and pLb464-8  (Pediococcus claussenii (hitA (horA (horC (Lb464 genome annotation by the NCBI PGAP pathway  producedaussenii , notablyii (hitA , horA (1tA (horA , and horrA (horC  on pLb46rA (horC . Based oLactobacillus brevis BSO 464 were deposited in GenBank under accession numbers CP005977, CP005978, CP005979, CP005980, CP005981, CP005982, CP005983, CP005984, and CP005985 for the chromosome and plasmids pLb464-1 to pLb464-8, respectively.The sequences for"}
+{"text": "NLRP3-gene encoding cryopyrin, leading to overproduction of IL-1\u03b2 and other NLRP3 inflammasome products. Myeloid-derived suppressor cells (MDSCs) represent a novel innate immune cell subset, are generated in tumor, infective, and proinflammatory microenvironments and are capable of suppressing T cell responses. Consequently, MDSCs are considered a key intermediary in balancing innate and adaptive immune responses, particularly under chronic disease conditions.Muckle-Wells syndrome (MWS) is caused by mutations in the We hypothesized that NLRP3 inflammasome-dependent factors induce the generation of MDSCs in MWS.highCD66bhighIL-4RainterHLA-DRlow neutrophilic cells in the PBMC fraction, according to previously established human MDSC analysis methods. The functionality of MACS-isolated MDSCs was assessed using polyclonal T cell proliferation and cytokine/chemokine secretion tests. Physician's global assessment of disease activity, CRP, ESR, and T helper cell subsets were determined at the same time points and correlated with MDSC levels. Serum samples of 22 MWS patients and 5 healthy controls were examined by multiplex technique for possible MDSC inducing factors.We studied granulocytic MDSC numbers in 25 MWS patients under anti-IL-1 therapy with canakinumab and 20 healthy controls. After Ficoll density gradient sedimentation, granulocytic MDSCs were characterized as CD33p = 0.0025), although clinical MWS-disease activity was generally low at time of examination. MDSCs were functionally competent, as they suppressed polyclonal T cell proliferation, Th1, Th2, and Th17 responses. MDSCs correlated directly with Treg/Th17 and Treg/Th1 ratios indicating an influence on T helper cell subsets. Multiplex assays revealed the established MDSC-inducing growth factors GM-CSF and VEGF elevated in MWS sera even under anti-IL-1 therapy with canakinumab.MWS patients under anti-IL-1 therapy displayed significantly elevated MDSC numbers  compared to healthy controls (mean 0.45 \u00b1 0.05%; range 0.12 - 1.04%; MWS patients under anti-IL-1 therapy display significantly elevated numbers of granulocytic MDSCs. Increased MDSCs in MWS might represent a novel autologous anti-inflammatory mechanism in autoinflammatory conditions and may serve as a future therapeutic target.N. Rieber Grant/Research Support from: NR, JKD, and DH obtained research grants from Novartis GmbH, Consultant for: NR and JKD took part in advisory boards for Novartis GmbH, A. Brand: None Declared, D. Neri: None Declared, T. Hall: None Declared, I. Sch\u00e4fer: None Declared, S. Hansmann: None Declared, J. K\u00fcmmerle-Deschner: None Declared, D. Hartl: None Declared."}
+{"text": "Pale-pink euhedral cobaltoan dolomite was associated with kolwezite [(Cu1.33Co0.67)(CO3)(OH)2] and cobaltoan malachite [2(CO3)(OH)2]. A crystal with a Co:Mg ratio of 1:5.6 (SEM/EDAX measurement), twinned on (11 -2 0) was used for crystal structural refinement. The refinement of the structural model of Reeder & Wenk  showed that Co is totally incorporated in the Mg site, with refined occupancy Mg0.83Co0.17, which compares with Mg0.85Co0.15 from chemical data. The Co substitution reflects in the expansion of the cell volume, with a pronounced increasing of the c cell parameter. A structural study has been undertaken on a cobaltoan dolomite, with chemical formula CaMgral. 1983, 68, 769 DOI: 10.1107/S2056989015003126/br2247Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989015003126/br2247fig1.tif. DOI: Micro photograph of the cobaltoan dolomite specimen, where pale pink cobaltoan dolomite is associated with pale green cobaltoan malachite.Click here for additional data file.10.1107/S2056989015003126/br2247fig2.tifx . DOI: x Cartesian rotation axis. Ca-centered octa\u00adhedra are cyan, whereas Mg-centered octa\u00adhedra are yellow; carbon and oxygen atoms are represented as green and red spheres, respectively.The crystal structure of cobaltoan dolomite, in a projection along [100], slightly tilted by 5\u00b0 about along the Click here for additional data file.10.1107/S2056989015003126/br2247fig3.tif. DOI: Coordination polyhedra in cobaltoan dolomite. Displacement ellipsoids are drawn at the 50% probability.1049359CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:"}
+{"text": "Aeromonas hydrophila-like arabinose-negative isolates from diverse sources  sampled in Valencia (Spain) during 2004\u20132005, were characterized by 16S rRNA gene sequencing and extensive biochemical testing along with reference strains of most Aeromonas species. These isolates and all reference strains of A. hydrophila subsp. dhakensis and A. aquariorum showed a 16S rRNA sequence similarity of 99.8\u2013100%, and they all shared an identical phenotype. This matched exactly with that of A. hydrophila subsp. dhakensis since all strains displayed positive responses to the Voges-Prokauer test and to the use of dl-lactate. This is the first report of A. hydrophila subsp. dhakensis recovered from environmental samples, and further, from its original isolation in India during 1993\u20131994. This was accurately identified and segregated from other clinical aeromonads  by using biochemical key tests. The API 20 E profile for all strains included in A. hydrophila subsp. dhakensis was 7047125. The prevalence of this species in Spanish sources was higher for water (9.4%) than for feces (6%) or eels (1.3%). Isolates recovered as pure cultures from diseased eels were moderately virulent (LD50 of 3.3\u00d7106 CFU fish\u22121) to challenged eels in experimental trials. They were all resistant to ticarcillin, amoxicillin-clavuranic acid, cefoxitin, and imipenem, regardless of its source. Our data point to A. hydrophila subsp. dhakensis as an emerging pathogen for humans and fish in temperate countries.Eight  Aeromonas (family Aeromonadaceae) is ubiquitous in aquatic ecosystems that include chlorinated drinking water, raw sewage, and natural waters  and free-living fish in such habitats  during 1993\u20131994 , A. hydrophila subsp. dhakensis exhibits high divergence from A. hydrophila subsp. hydrophila and A. hydrophila subsp. ranae  but 78 to 84% in the case of A. hydrophila subsp. dhakensis  in November 2004 ; and stry others .A. hydrophila subsp. dhakensis CECT5743, A. hydrophila subsp. dhakensis CECT5745, A. aquariorum MDC310, and A. aquariorum MDC317 was obtained by the Colecci\u00f3n Espa\u00f1ola de Cultivos Tipo (CECT) service . Bacterial genomic DNAs were extracted according to a method described previously (Escherichia coli numbering]) (Escherichia coli numbering) (2 (100 mM), 1.0 \u03bcL dNTPs (10 mM each), 1.0 \u03bcL each forward and reverse primers (50 \u03bcM), 0.5 \u03bcL Taq polymerase  and 5.0 \u03bcL template DNA (50 ng \u03bcL\u22121) in a total volume of 50 \u03bcL. PCR amplifications of the DNA templates were performed using a PTC-100 ThermoCycler (MJ Research). The conditions for 16S rRNA gene amplification were (i) 4 min at 94\u00b0C; (ii) 30 cycles of 1 min at 94\u00b0C, 1 min 30 s at 52\u00b0C and 2 min at 72\u00b0C; and (iii) a final elongation step of 10 min at 72\u00b0C. Amplified products were examined by agarose gel electrophoresis (1.2%) and ethidium bromide staining. Purified amplicons (Mo Bio Laboratories) were sequenced by the dideoxy method using the BigDye Terminator v. 3.0 Ready Reaction cycle sequencing kit and analyzed in an ABI PRISM 3730 sequencer (Applied Biosystems). Sequencing primers were the same as those used in the amplification reaction but diluted tenfold (5 pmol).The almost complete 16S rRNA gene sequence of strains ABF132, ABF144, ABF145, MA17, MA26, MA131, 133.341, 133.343, CECT4588, eviously . UniversAeromonas , A. aquariorum , A. hydrophila subsp. hydrophila (CECT839T), A. veronii bv. veronii (CECT4257T), A. veronii bv. sobria (CECT4835), A. sobria (CECT4245T), A. jandaei (CECT4228T), A. popoffii (CECT4995), A. bestiarum (CECT4227T), A. allosaccharophila (CECT4199T), A. eucrenophila (CECT4224T), A. encheleia (CECT4342T), A. trota (CECT4255T), A. enteropelogenes (CECT4487T), A. caviae (CECT838T), A. media (CECT4232T), A. schubertii (CECT4240T), and A. diversa (CECT4254T) were examined in 45 tests described by us as valuable traits for identifying Aeromonas  were used in all isolates in order to know their API 20 E code.Minimal inhibitory concentrations (MICs) of kanamycin (K), tetracycline (TET), nalidixic (NA), oxolinic (OA) acid, flumequine (UB), erythromycin (ERY), rifampicin (RD) and chloramphenicol (CHL) were determined by the microbroth dilution method . In addi8, 107, 106, 105, 104, 103, 102, 101 CFU mL\u22121), or with 0.1 mL PBS (controls). Each set of six eels was kept in a 20 L aquarium under the following conditions: i) dechlorinated tap water, ii) water temperature around 20\u00b0C, iii) oxygen concentration in water was above 90% saturation, and iv) fish were not fed. Mortality was recorded daily for 10 days and was only considered if the challenged bacterium was recovered as pure culture from the internal organs.For the challenge experiment, six young eels of around 10\u201320 g were challenged by intraperitoneal injection with each of the bacterial doses , but that recovered from the ulcer was not  was calculated , FJ230076 (A. sanarellii A2-67T), FJ230077 (A. taiwanensis A2-50T), AJ508765 (A. hydrophila subsp. dhakensis LMG 19562T), X60408 (A. caviae NCIMB 13016T), X60415 (A. trota ATCC 49657T), AJ508766 (A. hydrophila subsp. ranae LMG 19707T), X60410 (A. media ATCC 33907T), X60404 (A. hydrophila subsp. hydrophila ATCC 7966T), S39232 , X60412 (A. sobria NCIMB 12065T), FJ976900 (A. rivuli WB4.1-19T), X60417 (Aeromonas sp. ATCC 35941T), AJ224308 (A. popoffii LMG 317541T), X60411 (A. eucrenophila NCIMB 74T), AJ224309 (A. encheleia CECT 4342T), AY532690 (A. molluscorum 848T), X60406 (A. bestiarum CIP 7430T), AF134065 , X60405 , X60407 , AB027544 , X74680 , FM999971 (A. piscicola S1.2T), X60413 (A. jandaei ATCC 49568T), AF170914 (A. culicicola MTCC 3248T), FJ230078 , X60414 (A. veronii bv. veronii ATCC 35624T), AJ536821 (A. simiae IBS S6874T), X60416 (A. schubertii ATCC 43700T), GQ365710 (A. diversa CECT 4254T).EU085557 . These were compared with those of A. hydrophila subsp. dhakensis LMG 19562T and A. aquariorum MDC 47T and with those from other Aeromonas-type strains , in accordance with that reported solely for the type strain of these species , using both simple matching (SSM) (J) coefficients. Moreover, they were all clearly segregated from the other type strains of the Aeromonas species whose phenotypic profile was included in the numerical analysis ; \u03b2-haemolysis of sheep red blood cells; and acid production from d-fructose, d-galactose, maltose, d-mannitol, d-mannose, sucrose, and d-trehalose; however, they all were negative for: Gram staining; production of brown diffusible pigment; susceptibility to O/129 (150 \u03bcg) (Oxoid discs); growth with 6% (w/v) NaCl; hydrolysis of urea; and acid production from d-amygdalin, dulcitol, d-fucose, meso-inositol, melibiose, d-raffinose, d-sorbitol, and d-xylose.The ng (SSM)  and Jaccanalysis . All str cluster  showed tA. hydrophila subsp. dhakensis-A. aquariorum\u201d cluster exactly matched that described for A. hydrophila subsp. dhakensis  and -positive  simultaneously  and positive  for the utilization of dl-lactate   , which was obtained from Aeromonas strains  . Finally results  and 2. T strains . In addi jandaei , using b jandaei .Aeromonas hydrophila subsp. dhakensis was previously isolated in India (Aeromonas-positive samples was 17 (53.1%) and 20 (26.7%), respectively, with 32 patients suffering from Aeromonas gastroenteritis. The overall prevalence of A. hydrophila subsp. dhakensis in these Aeromonas-positive specimens was of 8.7% (6/69), although it was higher for water (17.7%) than for feces (6.25%) or eels (5%). Thus, we have found a wider distribution of A. hydrophila subsp. dhakensis in Spain in comparison with its unique previous finding in association with patients but not with water or fish (Aeromonas in these waters ranged from (102 to 104 CFU mL\u22121) in winter to (104 to 107 CFU mL\u22121) in summer , or by contact with them  were recovered from two siblings, a 1-year-old boy and a 6-year-old girl, who had acute gastroenteritis accompanied by bloody stools and high-grade (\u226539\u00b0C) fever and required antibiotic treatment. The fact that these clinical isolates were rather multi-resistant to antibiotics   . Fortunaisolates , as alsoA. hydrophila subsp. dhakensis  which have been recovered from a wild European eel suffering from hemorrhagic septicemia . The present results constitute the first report of A. hydrophila subsp. dhakensis from a temperate country, suggesting the worldwide distribution of this species.In summary, species"}
+{"text": "HTLV-1 induces a persistent infection leading to an increased production of inflammatory cytokines. HIV, another retrovirus that causes systemic inflammation, is associated with atherosclerosis. However, few is known about HTLV-1-infection and atherosclerosis. To determine the prevalence and risk factors associated with atherosclerosis in patients infected with HTLV-1. a cross-sectional study involving 54 HTLV-1-infected patients (24 asymptomatic and 30 HAM/TSP), was carried out at the CHTLV between 2012 to 2013. Sociodemographic and cardiovascular risk factors were evaluated . Patients were submitted to Doppler echocardiography of both carotid and vertebral arteries to measure intimal-media layer. The association between intima-media thickness (IMT) and HAM/TSP diagnosis, HTLV-1 proviral load, low-density lipoprotrein cholesterol (LDL-c) or ultrasensitive C-reactive protein was determined. The mean age of patients was 57.1\u00b112.3 years, 72.2% were women. Clinical atherosclerosis (IMT\u22651.5mm) was found in nine patients (16.7%) and subclinical atherosclerosis (IMT>1 and <1.5) in 10 patients (18.5%). Atherosclerosis was more frequent in women (41%) compared with man (20%). Median age was significantly higher in patients with atherosclerosis . The mean of LDL-c level was 140.11\u00b143.25 mg/dl (ranging from 49 to 287mg/dl). No significant differences were observed between the mean LDL-c level in individuals with subclinical (126.10\u00b138.95mg/dl) and with atherosclerosis diagnosis (144.\u00b143.87mg/dl), compared with individuals with normal IMT (143.21\u00b144.65mg/dl), (p=0.4175). There was no correlation between HTLV-1 proviral load and IMT. No association was found between HTLV-1 infection and clinical or subclinical atherosclerosis as well as with the risk factors for cardiovascular diseases. A positive association between atherosclerosis and age was observed."}
+{"text": "The LysE superfamily consists of transmembrane transport proteins that catalyze export of amino acids, lipids and heavy metal ions. Statistical means were used to show that it includes newly identified families including transporters specific for (1) tellurium, (2) iron/lead, (3) manganese, (4) calcium, (5) nickel/cobalt, (6) amino acids, and (7) peptidoglycolipids as well as (8) one family of transmembrane electron carriers. Internal repeats and conserved motifs were identified, and multiple alignments, phylogenetic trees and average hydropathy, amphipathicity and similarity plots provided evidence that all members of the superfamily derived from a single common 3-TMS precursor peptide via intragenic duplication. Their common origin implies that they share common structural, mechanistic and functional attributes. The transporters of this superfamily play important roles in ionic homeostasis, cell envelope assembly, and protection from excessive cytoplasmic heavy metal/metabolite concentrations. They thus influence the physiology and pathogenesis of numerous microbes, being potential targets of drug action. Rh. Rh20]. Using rigorous statistical criteria, we have expanded the LysE superfamily nearly four-fold. In addition to the LysE, RhtB and CadD families identified previously, this superfamily now includes the following families: NAAT, CaCA2, MntP, ILT, TerC, NicO, GAP and DsbD. Members of each of these families have been characterized and shown to play roles in transport of amino acids and resistance of heavy metal ions, along with cell surface maintenance. Most families include secondary carrier type transporters catalyzing heavy metal or amino acid efflux, but one family catalyzes amino acid uptake, another catalyzes heavy metal ion uptake, and a third catalyzes transmembrane electron transfer. GAP proteins have not been mechanistically characterized, but based on their inclusion in the LysE superfamily, we tentatively propose that GAP proteins operate as secondary carriers, where the energy source for lipid export is the proton motive force.Through sequence analyses, we were able to recognize a distinct pattern of homology. That is, LysE, RhtB, NAAT, CaCA2, MntP, ILT, TerC, NicO, GAP and DsbD proved to be homologous in 3 or more TMSs. The 3 TMSs that aligned are usually between the first 3 TMSs, the second 3 TMSs or both. This observation fits the predicted evolutionary pathway presented in According to the phylogenetic tree, amino acid exporter families RhtB and LysE branch close to each other, as suggested from previous studies . In contThis study suggests that members of the LysE Superfamily are involved in ionic homeostasis, protection from excessive cytoplasmic heavy metal/metabolite concentrations, cell envelope assembly and transmembrane electron flow. Many of the family members, however, are still poorly understood from functional and physiological standpoints. In continuing this project, genome context analyses will be conducted on members of each family. This will allow functional predictions, further promoting an understanding of the significance of these proteins. To date, no crystal structures exist for a member of this superfamily, and such studies will be crucial for understanding their mechanistic details. Thus, studies on the LysE superfamily remain in their infancy.S1 FigAlong with a step-wise description of the methods, the parameters for the programs used in major analyses are summarized.(TIF)Click here for additional data file.S2 Fig(A) LysE vs. RhtB. (B) RhtB vs. CadD. (C) LysE vs. CadD.(PDF)Click here for additional data file.S3 Fig(A) CadD vs. CaCA2. (B) LysE vs. CaCA2. (C) RhtB vs. CaCA2.(PDF)Click here for additional data file.S4 Fig(A) RhtB vs. MntP. (B) CadD vs. MntP. (C) CaCA2 vs. MntP.(PDF)Click here for additional data file.S5 Fig(A) CadD vs. ILT. (B) RhtB vs. ILT. (C) CaCA2 vs. ILT.(PDF)Click here for additional data file.S6 Fig(A) RhtB vs. TerC. (B) CadD vs. TerC. (C) LysE vs. TerC (D) MntP vs. TerC. (E) ILT vs. TerC. (F) CaCA2 vs. TerC.(PDF)Click here for additional data file.S7 Fig(A) LysE vs. NAAT. (B) RhtB vs. NAAT. (C) CadD vs. NAAT (D) MntP vs. NAAT. (E) TerC vs. NAAT.(PDF)Click here for additional data file.S8 Fig(A) RhtB vs. NicO. (B) CadD vs. NicO. (C) TerC vs. NicO (D) NAAT vs. NicO.(PDF)Click here for additional data file.S9 Fig(A) RhtB vs. GAP.(PDF)Click here for additional data file.S10 Fig(A) RhtB vs. DsbD. (B) CaCA2 vs. DsbD. (C) MntP vs. DsbD. (D) NAAT vs. DsbD. (E) GAP vs. DsbD.(PDF)Click here for additional data file.S11 Fig(A) LysE. (B) RhtB. (C) CadD. (D) CaCA2. (E) MntP. (F) NAAT. (G) NicO. (H) GAP. (I) DsbD. (J) ILT. (K) TerC.(PDF)Click here for additional data file.S12 FigGSAT comparisons between TMS#1\u20133 and TMS#4\u20136 for three CaCA2 homologues with assigned UniProt accession numbers. (A) Q2JWH3. (B) I7M883. (C) K4DX00.(PDF)Click here for additional data file.S13 FigGSAT comparisons between TMS#1\u20133 and TMS#4\u20136 for three ILT homologues with assigned UniProt accession numbers. (A) Q8YX33. (B) K9Q6B8. (C) J2KV33.(PDF)Click here for additional data file.S14 FigGSAT comparisons between TMS#1\u20133 and TMS#4\u20136 for three MntP homologues with assigned UniProt accession numbers. (A) A8SU47. (B) R9SLI6. (C) C6JCY1.(PDF)Click here for additional data file.S15 FigGSAT comparisons between TMS#1\u20133 and TMS#4\u20136 for three TerC homologues with assigned UniProt accession numbers. (A) A4IKQ1. (B) G8M4S7. (C) R9LI44.(PDF)Click here for additional data file.S16 Fig(A) LysE. (B) RhtB. (C) CadD. (D) CaCA2. (E) MntP. (F) ILT. (G) TerC. (H) NAAT. (I) NicO. (J) GAP. (K) DsbD.(PDF)Click here for additional data file.S17 Fig-20 by BLAST (Using UniProt) for each query sequence to improve the accuracy of aligning a small number of distantly related sequences. The bootstrap values are shown in blue text and located near each node.The Mafft-homologs function was set to retrieve 200 homologs at a threshold E-value of 1e(TIF)Click here for additional data file.S18 Fig(PDF)Click here for additional data file.S19 Fig(PDF)Click here for additional data file.S20 Fig(PDF)Click here for additional data file.S21 Fig(PDF)Click here for additional data file.S22 Fig(PDF)Click here for additional data file.S23 Fig(PDF)Click here for additional data file.S24 Fig(PDF)Click here for additional data file.S25 Fig(PDF)Click here for additional data file.S26 Fig(PDF)Click here for additional data file.S27 Fig(PDF)Click here for additional data file.S28 Fig(PDF)Click here for additional data file.S29 FigThe tree was generated using the SuperFamilyTree program and viewed using FigTree. It depicts the evolutionary relationship between the 11 different families in this study. Clustering indicates closer phylogenetic relationships. The tree is based on tens of thousands of BLAST bit scores generated with the SFT1 program where every protein was compared with every other protein included in the analysis. The SFT2 program was used to integrate all of the information to show the relationships of the eleven families to each other. Bootstrap values have been added in blue text and located near each node.(TIF)Click here for additional data file.S1 Supporting InformationThe corresponding zip file contains the FASTA files generated using Protocol1, for comparisons with Protocol2.(ZIP)Click here for additional data file.S2 Supporting InformationThe corresponding zip file contains the multiple sequence alignment (MSE) outputs generated using ClustalX, Mafft, and ProbCons. These MSEs have been used to generate (ZIP)Click here for additional data file.S3 Supporting InformationThe corresponding zip file contains the 100 trees generated from SFT, the consensus tree, the FASTA sequences used to generated the trees, and the newick file for the best tree generated from RAxML analyses (described in (ZIP)Click here for additional data file.S4 Supporting InformationThe corresponding zip file contains the FASTA files used to conduct MEME Suite analyses shown in Figs (ZIP)Click here for additional data file.S5 Supporting InformationThe corresponding PDF file contains the (PDF)Click here for additional data file.S6 Supporting InformationThe corresponding PDF file contains the (PDF)Click here for additional data file.S7 Supporting InformationThe corresponding PDF file contains the (PDF)Click here for additional data file."}
+{"text": "Thromboembolic complications (TEC) are very common and lethal in patients suffering from traumatic injury . The trat test, univariate and multivariate logistic regression analyses were performed.A retrospective chart review (2010 to 2013) was conducted on adult trauma patients that were admitted into a level 1 Trauma Centre in Toronto. Demographics, date of PTP initiation, date of TEC diagnosis (CT-PE/US Doppler), injury type and severity were collected. A comparison between early and late PTP initiation has been made with regards to TEC development. Student's P < 0.0005), had lower ISS , shorter length of stay (LOS) , more pelvic fractures , more head injury , less blunt trauma , lower incidence of TEC (5.3% (44) vs. 8.5% (42), P = 0.023), and lower mortality rate (1.5% vs. 7.5%). Univariate analysis showed LOS (P < 0.0005), ISS (P < 0.0005), time to PTP initiation (P = 0.0018) and blunt MOI (P = 0.0099) significantly associated with TEC events. Multivariate analysis, however, showed TEC events correlated only to LOS (P = 0.0001). Stepwise multiple logistic regression confirmed LOS as independently associated with TEC events .A total of 1,312 patients received PTP, 821 (62.5%) initiated early PTP (within 48 hours) while 491 (37.5%) initiated after 48 hours. The group that initiated early prophylaxis was younger (mean: 46 vs. 55, Mortality rates in patients with delayed PTP are higher. Our study shows LOS as the only independent predictor for TEC. However, this might not necessarily reflect causation. Delayed PTP appears not to be an independent predictor to TEC events in trauma patients, which favours current clinical trends when it comes to contraindicating early PTP initiation."}
+{"text": "A single-bolus non-ECG-triggered method was previously proposed to reduce the complexity of quantitative myocardial perfusion imaging . CompareData was acquired continuously with no ECG-triggering. 26 projections per slice were acquired following each SR-preparation in an interleaved manner. HYPR was used to reconstruct images with 5-fold undersampling. A sliding window was used to produce series of low resolution images in the basal slice with a temporal resolution of 44 ms. Triggering signal was derived retrospectively from the mean signal intensity in an ROI drawn around the heart. AIF was estimated using single-cardiac cycle T1 mapping [nd study, another 12 healthy volunteers underwent stress-rest studies using the proposed method. Mean myocardial perfusion reserve (MPR) was compared with previously published values.Ten healthy volunteers underwent rest perfusion MRI on a Siemens 3T Verio. A dual-bolus protocol was performed using a conventional clinical Cartesian sequence for comparison [Mean MBF values found using the ECG-triggered dual-bolus method and the non-ECG-triggered integrated T1 mapping method were 0.82 \u00b1 0.21 and 0.76 \u00b1 0.13 ml/min/g, respectively. There was no significant difference (p = 0.45) between them. The mean rest and stress MBF and MPR was 0.86\u00b10.5, 3.91\u00b11.1 ml/min/g, and 4.31\u00b11.3, respectively.A 3-slice, non-ECG-triggered, single-bolus quantitative perfusion MR method with integrated T1 mapping for AIF measurement produces similar MBF as the reference dual-bolus method with comparable ventricular coverage. Mean MPR is similar to those reported in literature. The proposed non-ECG-triggered technique improves ease-of-use, and has the potential to improve robustness to arrhythmias. This method may improve the clinical feasibility of quantitative myocardial perfusion imaging.NIH grant numbers T32 EB51705 and RO1 EB002623, NIBIB grant number EB002623, AHA Scientist Development Grant 14SDG20480123, GCRC grant MO1-RR00425, and Edythe L. Broad Women's Heart Research Fellowship UN55ES6580F."}
+{"text": "Moderate-to-severe allergic rhinitis (AR) is often poorly controlled. Patients remain symptomatic on treatment, despite multiple therapies. A more effective treatment is needed. We assessed the efficacy of MP29-02*  and fluticasone propionate (FP) in an advanced delivery system) during different seasons and for different symptoms and severities vs AZE, FP or placebo (PLA).Four thousand and twenty two moderate/severe SAR patients (\u226512 yrs) were randomized into 4 double-blind, PLA-controlled, 14-day, parallel-group trials to MP29-02*, AZE, FP or PLA nasal sprays (1 spray/nostril bid), during the Texas mountain cedar (MP4001), Spring (MP4002), Autumn (MP4004) and Spring/Summer (MP4006) seasons. Overall change from baseline (CFB) in reflective total nasal symptom score (rTNSS) was the primary endpoint. It was assessed by severity post-hoc in 2 ways . CFB in individual nasal and ocular symptom scores was also assessed.The response to MP29-02* was consistent across seasons; mean CFB -5.5, -5.5, -5.6 and -5.6 in each study (p<0.001 vs PLA). Nasal symptom relief was significantly greater with MP29-02* than with FP or AZE in all studies. In study MP4001 MP29-02 was approx. twice as effective as FP (relative difference (RD) 47%; p=0.0031) and 3 times as effective as AZE . For less severe AR (defined by median baseline rTNSS) the RD to MP29-02* was 42% vs FP (p=0.0188) and 64% vs AZE (p=0.0002), increasing to 49% and 70% vs FP (p=0.0436) and AZE (p=0.0035), respectively for more severe AR. When severity was categorized according to median baseline RQLQ the RD to MP29-02* was 60% vs FP (p=0.0244) and 69% vs AZE (p=0.0068) for those with less severe AR compared to 49% vs FP (p=0.0194) and 64% vs AZE (p=0.0013) for those with more severe AR. MP29-02* provided superior relief from all nasal and ocular symptoms than AZE or FP, which was particularly evident for congestion  & ocular itching .MP29-02* provides consistently superior symptomatic relief to an intranasal antihistamine or topical corticosteroid in AR patients regardless of season, symptom or severity, supporting MP29-02 as the drug of choice for AR.*Dymista"}
+{"text": "Erythropoietin treatment could augment the immune response and could be a tiger for systemic lupus erythematous flare-up and increasing the activity of renal and systemic lupus.Anemia is a common clinical finding in end-stage renal failure (ESRD). Systemic lupus erythematous (SLE) is an important cause of chronic kidney disease and anemia in lupus nephritis has different and complex causes, including; anemia of chronic disease (ACD), autoimmune hemolytic anemia (AHA), iron deficiency anemia (IDA), pure red cell aplasia (PRCA), pernicious anemia (PA), myelofibrosis, hemophagocytic histiocytic syndrome, thrombotic microangiopathy and drug induced myelotoxicity . Combina3/\u00b5l  and reticulocyte was 1%. Also, platelets count; 156\u00d7103/\u00b5l, serum creatinine: 7.63mg/dl, BUN: 84 mg/dl, uric acid: 7.5 mg/dl, phosphorous: 5.8 mg/dl, calcium: 8.4 mg/dl. Serum complements were as follow: C3: 68 mg/dl (90-180), C4: 10 mg/dl (10-40) and CH50: 76% (90-98). Further laboratory tests revealed; anti MPO\u2013ANCA (IgG): 0.8 u/ml , Anti PR3-ANCA(IgG): 6.0 U/ml , Anti-dsDNA: 42 U/ml, LDH: 800 IU/ml (< 500 ), AST: 36 IU/ml, ALT: 38 IU/ml (<40), serum Na: 139 meq/l, serum K: 5, 0 meq/l, total bilirubin: 0.33 mg/dl (0.1-1.2), direct bilirubin: 0.09 mg/dl (0.1-0.4). Antiphospholipid antibody (IgG/IgM) was negative. Stool examination was negative for occult blood. Also upper gastrointestinal endoscopy had normal results. Peripheral blood examination revealed a large number of teardrop red blood cells. Bone marrow aspiration was unsuccessful (dry tap), bone marrow biopsy was compatible with myelofibrosis. Renal allograft biopsy revealed diffuse lupus nephritis (lupus nephritis class IV) with cellular crescents. This situation was difficult to make a decision to start an intensive immunosuppressive therapy in the presence of marrow failure. Despite receiving adequate dialysis and blood transfusion and adjusted immunosuppressive therapy her general condition worsened and died two months after the admission.We recently admitted a 23 year-old-female with advance renal failure and oliguria. Physical examination on admission revealed a blood pressure of 130/90 mmHg and pulse rate of 105 minute/min. Cardiac examination showed a loud systolic murmur. Abdominal examination disclosed a palpable spleen. Hemodialysis was started with temporary jugular vein catheter. Laboratory examination revealed; hemoglobin: 4.1 g/dl, hematocrit: 17%, MCV: 78.6 fl, MCH: 24.4 pg/cell and MCHC: 31.1 g/dl. White blood cell count: 3.6\u00d710In SLE, there is an association between the level of inflammatory cytokines and intensity of anemia. Inflammatory cytokines inhibit erythropoietin production and erythroid progenitor cells proliferation. In severe cases, inflammatory cytokines lead to myelofibrosis ,2. RenalMRA is the single author of this paper.No financial support by any institution.None to declare."}
+{"text": "From 1999\u20132002 to 2009\u20132012, the prevalence of diabetes increased for non-Hispanic black and Mexican American adults, but remained stable for non-Hispanic white adults, increasing the disparity with the two minority populations. In 1999\u20132002, the prevalence of diabetes among non-Hispanic black (14.0%) and Mexican American (13.9%) adults aged \u226520 years was 1.6 times the prevalence among non-Hispanic white adults (8.5%). By 2009\u20132012, diabetes prevalence among Mexican American adults (20.5%) had increased to more than twice the prevalence among non-Hispanic white adults (9.1%); among non-Hispanic black adults (17.9%), the prevalence had increased to nearly twice that among non-Hispanic white adults.Source: Health, United States, 2014: with special feature on adults aged 55\u201364. Table 44. Available at http://www.cdc.gov/nchs/hus.htm.Reported by: Sheila J. Franco, sfranco@cdc.gov, 301-458-4331; Shilpa Bengeri."}
+{"text": "Doubts on the efficacy of exercise treatment for adolescents with Idiopathic Scoliosis (IS) still exists.To verify the effectiveness of exercises in everyday clinics.Prospective observational controlled cohort study nested in a prospective database started in March 2003.Setting: outpatient tertiary referral clinics.Participants: consecutive patients from start of the database to 31/12/2010. Inclusion criteria: IS; Risser 0-2; 11\u00b0 to 20\u00b0 Cobb; age 10 years or more; first evaluation. Exclusion criteria: consultations only; immediate prescription of a brace.Groups: Physiotherapic Specific Scoliosis Exercises - SEAS school ; Controls (CON: less than 15 min/week); Usual Physiotherapy (UP: other institutes/protocols).End-Of-Treatment (EOT): medical prescription, bracing, Risser 3.Failures: bracing for scoliosis; EOT above 30\u00b0.Statistical analysis: intent-to-treat (ITT: drop-outs included as failures) and efficacy (EA: only EOT patients). Relative Risk of failure (RR), 95% Confidence Interval (CI), and clinical and radiographic changes have been calculated.Out of 327 patients, 34 (10%) were excluded due to bracing at first evaluation. We included 293 adolescents: 145 PSSE, 95 UP, 53 CON, with no differences at baseline. Physicians prescribed bracing (failure) without differences among groups.Failures and drop-outs were 84 (28.7%) and 47 (16.0%) respectively: 21.4% and 18.6% in PSSE; 33.7% and 9.5% in UP; 39.6% and 20.8% in CON.Efficacy analysis (RR): CON vs PSSE 1.90 (IC 1.48-2.33); UP vs PSSE 1.42 (1.01-1.82); CON vs UP: not significant.Intent-to-treat (RR): CON vs PSSE 1.51 (1.21-1.80); CON vs UP 1.40 (1.08-1.72); UP vs PSSE: not significant.At the end of exercises, aesthetics (TRACE) improved statistically in PSSE (1.8 points out of 12) and UP (1.5), not in CON; only PSSE improvement was statistically better than CON.Patients performing UP or nothing (CON), compared to those treated with PSSE (SEAS), increase the risk of failure (bracing and/or 30\u00b0 at EOT) 1.9 and 1.4 times respectively (EA)."}
+{"text": "Evidence of the association between vitamin D, insulin resistance and oral disposition index (oDI) in obese children and adolescents is limited. We investigated serum 25(OH) D levels in obese children and adolescents in Zhejiang, China, and determined the relationship between serum 25(OH) D and glucose metabolism.A cross-sectional design was used. All together 348 obese and 445 non-obese children and adolescents (aged from 6-16 years old) were enrolled in this study. Obese children were divided into four subgroups: normal glucose tolerance (NGT), isolated impaired fasting glucose (IFG), isolated impaired glucose tolerance (IGT), combined IFG and ITG (IFG+ITG) according to the oral glucose tolerance test. We measured serum 25(OH) D levels and calculated the homeostasis model of insulin resistance (HOMA-IR), the whole body insulin sensitivity index (WBISI), the product of \u03b2-cell function and insulin sensitivity by the disposition index (DI).The levels of 25(OH)D in obese group were significantly lower than those of non-obese group; serum 25(OH)D level in obese with NGT group was higher than that of the other three subgroups, and it was significantly inversed with LogHOMA-IR , positively correlated with LogWBISI, LogHOMA0DI after control for age, sex, season, puberty stage . Obese patients with vitamin D deficiency have a significantly higher risk of disturbing the glucose metabolism, such as IFG, ITG, IFG plus ITG, either IFG or ITG, for its OR 3.198(95%CI 1.467-6.97), 5.443(95%CI 1.863-15.897), 5.560(95%CI 1.212-25.502), 4.007(95%CI 2.017-7.962).25(OH) D deficiencies or insufficiency are common in obese children and adolescents in Zhejiang, China. Obese patients with 25(OH) D deficiency (<30nmol /L) are at higher risk for abnormal glucose metabolism."}
+{"text": "This study determined the effects of eight weeks of heavy resistance training combined with branched-chain amino acid (BCAA) supplementation on body composition and muscle performance.Nineteen non-resistance-trained males resistance-trained (3 sets of 8-10 repetitions) four times/week for eight weeks while also ingesting 9 g/day of BCAA or 9 g/day of placebo (PLAC) on exercise days only . Data were analyzed with separate 2 x 2 ANOVA (p < 0.05).For total body mass, neither group significantly increased with training (p = 0.593), and there also were no significant changes in total body water (p = 0.517). Also, no training- or supplement-induced (p = 0.783) changes occurred with fat mass or fat-free mass (p = 0.907). Upper-body (p = 0.047) and lower-body strength (p = 0.044) and upper- (p = 0.001) and lower-body muscle endurance (p = 0.013) were increased with training; however, these increases were not different between groups (p > 0.05).When combined with heavy resistance training for eight weeks, 9 g/day of BCAA supplementation, half given 30 min before and after exercise, had no preferential effects on body composition and muscle performance."}
+{"text": "Mycobacterium abscessus complex, the third most frequent mycobacterial complex responsible for community- and health care-associated infections in developed countries, comprises of M. abscessus subsp. abscessus and M. abscessus subsp. bolletii reviously referred as Mycobacterium bolletii and Mycobacterium massiliense. The diversity of this group of opportunistic pathogens is poorly described.M. abscessus complex genomes found a pan-genome of 6,153 proteins and core-genome of 3,947 (64.1%) proteins, indicating a non-conservative genome. Analysing the average percentage of amino-acid sequence identity (from 94.19% to 98.58%) discriminates three main clusters C1, C2 and C3: C1 comprises strains belonging to M. abscessus, C2 comprises strains belonging to M. massiliense and C3 comprises strains belonging to M. bolletii; and two sub-clusters in clusters C2 and C3. The phylogenomic network confirms these three clusters. The genome length (from 4.8 to 5.51-Mb) varies from 5.07-Mb in C1, 4.89-Mb in C2A, 5.01-Mb in C2B and 5.28-Mb in C3. The mean number of prophage regions (from 0 to 7) is 2 in C1; 1.33 in C2A; 3.5 in C2B and five in C3. A total of 36 genes are uniquely present in C1, 15 in C2 and 15 in C3. These genes could be used for the detection and identification of organisms in each cluster. Further, the mean number of host-interaction factors  varies from 70 in cluster C1, 80 in cluster C2A, 74 in cluster C2B and 93 in clusters C3A and C3B. No significant differences in antibiotic resistance genes were observed between clusters, in contrast to previously reported in-vitro patterns of drug resistance. They encode both penicillin-binding proteins targeted by \u03b2-lactam antibiotics and an Ambler class A \u03b2-lactamase for which inhibitors exist.In-depth analysis of 14 published M. abscessus complex comprises three genomospecies, corresponding to M. abscessus, M. bolletii, and M. massiliense. The genomics data here reported indicate differences in virulence of medical interest; and suggest targets for the refined detection and identification of M. abscessus.Our comparative analysis indicates that The online version of this article (doi:10.1186/1471-2164-15-359) contains supplementary material, which is available to authorized users. Mycobacterium abscessus was long confused with Mycobacterium chelonae[Mycobacterium salmoniphilum[Mycobacterium immunogenum[Mycobacterium massiliense[Mycobacterium bolletii[Mycobacterium franklinii[Mycobacterium chelonae-abscessus complex. This complex is the third most frequent mycobacterial complex infecting humans in developed countries besides the Mycobacterium tuberculosis and Mycobacterium avium complexes [M. salmoniphilum[The non-tuberculous mycobacterium  chelonae. Other coniphilum, Mycobacmunogenum, Mycobacssiliense, Mycobac bolletii and Mycoranklinii altogethomplexes , 8. Biblomplexes . Not onlomplexes , 11 and omplexes \u201314 are ioniphilum, 15.M. abscessus comprises two subspecies named M. abscessus subsp. abscessus and M. abscessus subsp. bolletii. Later taxon accommodates isolates previously referred as M. bolletii or M. massiliense[M. abscessus and M. bolletii, it shares 99% sequence identity with M. massiliense. RpoB gene sequencing founded the description of recent species [M. massiliense from M. bolletii. In this report, the previous nomenclature M. abscessus, M. bolletii and M. massiliense forming the M. abscessus complex, has been retained for clarity.Current nomenclature is that the species ssiliense. This no species \u201319 but y species \u201322. Mult species  and mult species  differenM. abscessus, 13 M. massiliense and two M. bolletii genomes in the National Center for BioInformatics (NCBI) genome database provides new opportunities to assess the diversity of this species. Here, we review 14 complete published M. abscessus complex genomes and compare them with the re-annotated M. tuberculosis H37Rv genome  includes M. abscessus type strain and strains M93, 94, M152 and Go06; cluster 2 (C2) contains two subclusters: cluster 2A (C2A) includes M. massiliense type strain and strains M154 and M18; cluster 2B (C2B) includes strains 47\u00a0J26, M115, M172 and M139; cluster 3 (C3) includes two subclusters: cluster 3A (C3A) includes M. bolletii type strain and cluster 3B (C3B) includes M. bolletii strain M24 (Table\u00a0The average percentage of amino-acid sequence identity (AAI) of core proteins was determined as previously described . The AAIM. abscessus complex proteomes were further aligned using Mauve software [M. abscessus differently from the whole genome concatenated tree   and BLASC1 strains have been isolated from American and Malaysian patients suffering knee infection and lower respiratory infection, respectively , M. massiliense (C2) and M. bolletii (C3). Using an AAI <97% threshold would further determine two subspecies in M. massiliense (C2A and C2B) and in M. bolletii (C3A and C3B). Recent whole genome sequencing analyses of clinical isolates in Great Britain also clearly distinguished three clusters in agreement with the three here reported [M. abscessus complex, to recognize three genomospecies M. abscessus (C1), M. bolletii (C2), and M. massiliense (C3); and four unnamed subspecies C2A, C2B; C3A, C3B.Altogether, genomics analyses revealed a more heterogeneous structure of abscessus. It has abscessus, 35. Thereported . All theM. abscessus median GC% content is 64.2%, ranging from 62.7% (M. abscessus ATCC 19977) to 64.2% (strain Go 06). The GC% is not characteristic of the clusters as the median GC% content of C1, C2A and C3 is 64.2%, close to the median 64.1% GC% content in C2B.M. abscessus M154) to 5.51-Mb (M. abscessus M24) with a median of 5.07-Mb. The median of genome size is 5.07-Mb in C1, 4.89-Mb in C2A, 5.01-Mb in C2B and 5.28-Mb in C3. Differences in the genome size correlate with the number of prophage regions which are detected in 13/14\u2009M. abscessus genomes  has the smallest genome encoding no prophage whereas M. bolletii M24 (C3) has the largest genome encoding seven prophage regions  and phage tape measure protein. Both ends of this region encode putative phage integrases. M. abscessus genomes encode small prophage-like regions. However, only M. bolletii has been reported to produce a mycobacteriophage that we named Araucaria after we recently resolved its electron microscopy 3D structure [M. abscessus M94 genome harbours one particular pseudo-tRNA spanning the region 51,150-57,394 in contig 33, which is not observed in the other M. abscessus genomes [M. abscessus however, no such genes were identified but phages could be targeted for the differentiation between the three M. abscessus genomospecies.However, there is a significant 14.7% variation in the genome length from 4.8-Mb  genes conferring resistance to tetracyclyine and doxycycline; the number of tet(M) genes was correlated to the resistance to cyclines in Escherichia coli[M. massiliense was reported to be susceptible and M. abscessus and M. bolletii to be resistant to doxycycline [M. abscessus genomes encode resistance to fusidic acid, glycopeptides, MLS (Macrolide-Lincosamide-StreptograminB), phenicols, rifampicin, sulphonamide and trimethoprim. Also, M. abscessus genomes encode FolP homologs conferring resistance to cotrimoxazole, homolog of UDP-N- acetylglucosamine 1-carboxyvinyltransferase, a MurA protein conferring resistance to fosfomycin and homologs of 23S rRNA methylases conferring resistance to macrolides. Also, M. abscessus genome encodes an erm(41) gene which mutations were reported to confer clarithromycin resistance [In-vitro tests showed that M. massiliense clinical isolates could be distinguished from M. abscessus isolates for their susceptibility to ciprofloxacin [M. bolletii isolates were reported to be resistant to all quinolones [M. abscessus exhibiting high resistance to ciprofloxacin [M. abscessus mycobacteria encode qepA2, a plasmidic gene conferring quinolone resistance in gram-negative bacteria [M. abscessus mycobacteria were reported to be in-vitro resistant to penicillin, amoxicillin, cefoxitin, ceftriaxone, cefotaxime and imipenen [M. abscessus genomes encode an Ambler class A \u03b2-lactamase homologous to \u03b2-lactamases in gram-negative bacteria and to two \u03b2-lactamases in M. tuberculosis. \u03b2-lactamases inhibitors have not been evaluated against M. abscessus sensu lato mycobacteria.As all mycobacteria, sistance \u201344. M. aproteins ; and a msistance . M. abscsistance , 47. Indchia coli. Howeverycycline . M. abscsistance . In-vitrfloxacin  whereas inolones . A mutatfloxacin . This obfloxacin . Accordibacteria . M. abscimipenen . This coimipenen , 55. M. M. abscessus are ubiquitous environmental organisms in soil and water [M. chelonae, M. abscessus, M. massiliense and M. immunogenum were reported to survive within Acanthamoeba polyphaga tropohozoites and cysts [M. abscessus genomes encode factors implicated in host interactions. The mean number of genes encoding proline-glutamate (PE), proline-proline glutamate (PPE), 10-kDa lipoprotein antigen precursor (LpqH), Mammalian Cell Entry (MCE), oxidoreductase (Yrbe) and type VII secretion system is of 70 in C1, 80 in C2A, 74 in C2B and 93 in C3. In M. abscessus, rough colonies lack mmpL4 (a gene required for glycopeptidolipid biosynthesis) and lost surface colonization, replication into human macrophages and stimulation of innate immune response; these observations suggested that glycopeptidolipid was a virulence factor [M. avium subsp. avium[M. abscessus genomes encode MCE proteins similar to M. tuberculosis H37Rv. MCE operon promotes internalization of M. tuberculosis by mammalian cells [mce operons which correlated with pathogenicity [M. abscessus genomes encode 12 (C1) to 21  copies of Yrbe proteins. As for secretion systems, recent evidences showed that mycobacteria evolved specialized type VII secretion systems to transport extracellular proteins across the cell wall [M. tuberculosis[M. abscessus, our analyses indicate that ESX-3 and ESX-4 systems are conserved  lacks two proteins of the ESX-3 system and M. abscessus M93 (C1) lacks ESAT-6 like and CFP-10-like proteins secreted by the ESX-4 system. Interestingly, M. abscessus M18 (C2A) encodes ESAT-6 and CFP-10 proteins secreted by ESX-1 system. In addition, there are two or three PE and six  to 12 (M. bolletii M24) PPE proteins, which are reported to be involved in the virulence of M. tuberculosis[Actinobacteria  and pseudomonas (Pseudomonas aeruginosa and Burkholderia cepacia). Although distantly related, these bacteria share the same ecosystem as M. abscessus within cystic fibrosis microbiota.nd water  where thnd cysts . Accordie factor \u201358. Accoe factor  and biofe factor . Glycopesp. avium. M. abscan cells  and initan cells . The numgenicity , varies ell wall . Type VIerculosis, 66. In erculosis. Our anaM. abscessus has a non-conservative genome, suggesting the possibility of on-going transfer of additional genetic material. Unsurprisingly, M. abscessus has already acquired antibiotic resistance. Also, phages have mediated diversity and horizontal gene transfer which drived the rapid evolution of this complex. Indeed, gene transfers have driven the evolution of M. abscessus towards three different genomospecies M. abscessus, M. massiliense and M. bolletii; and the evolution of four different yet unnamed subspecies. Each genomospecies has its own specificities in terms of genome size, prophagome and genome content. We identified 66 genes uniquely present in each genomospecies; these genes could be used in refined detection and identification of M. abscessus organisms. These genomic differences support differences in host interactions and the clinical presentation of infection with M. massiliense (C2A and C2B) being more virulent than the two other genomospecies. Host-interaction factors may contribute to the ability of M. abscessus to colonize mammalian hosts where its respiratory tract habitat put it in close proximity to other serious opportunist pathogens which can act as donors of additional host-interaction factors.Our in-depth genomic analyses indicate that M. abscessus genomespecies will help understanding their pathogenesis factors and could reveal new, more specific targets for drug design and diagnosis tools.Here reported informations regarding differences between M. abscessus strains were downloaded from Genbank (Table\u00a0The whole genomes of 14\u2009http://www.uniprot.org/), the Clusters of Orthologous Groups (COG) [Protein sequences were predicted using prodigal software  to generps (COG)  and a hoM. abscessus homologous genes using orthoMCL [Proteome sequences were compared using by BlastP and pairwise alignments using ClustalW and the ANI was determined by the mean percentage of nucleotide sequence identity of core proteins . We clusorthoMCL  on the tM. abscessus proteomes were aligned using Mauve software [software  to infersoftware . The ortsoftware . Using tsoftware .http://treebase.org/treebase-web/home.html) repository, under the accession URL http://purl.org/phylo/treebase/phylows/study/TB2:S15632.The data set of Figure\u00a0http://purl.org/phylo/treebase/phylows/study/TB2:S15632?x-access-code=6fa2ebc53b96e3ae412a8df19187ab41&format=html.Reviewer access URL: The data sets of Figure\u00a0The data sets of Figure\u00a0A-AlignedM. abscessusgenome matrix constructed using Mauve software to infer phylogeny using the Neighbor-Net algorithm in the package SplitsTree4. B- The matrix of binary discrete characters  constructed using the orthologous group data found by orthoMCL to infer phylogeny using the Neighbor-Net algorithm in the package SplitsTree4. (XLS 28 KB)Additional file 1: The matrix used for Heatmap clusterisation ofMycobacterium abscessustype VII secretion system compared toMycobacterium tuberculosisH37Rv. (XLS 26 KB)Additional file 2:"}
+{"text": "Coinfection in patients with SARS-CoV-2 has been associated with greater complications. We describe the clinical characteristics and outcomes of 126 pediatric patients with COVID-19 and viral, bacterial, or fungal coinfection.We retrospectively reviewed and analyzed electronic data of all pediatric patients who tested positive for SARS-CoV-2 from April 16, 2020, to April 15, 2022, in our center. Confirmation of COVID-19 was based on positive RT-PCR. Viral coinfections (VC) were identified using a multiplex RT-PCR respiratory viral panel, bacterial coinfection (BC) was determined by positive bacterial culture  or clinical/radiological manifestations and antimicrobial assessment by a pediatric Infectious Diseases expert and fungal coinfection (FC) diagnosis based on Consensus definitions of invasive fungal disease.During the study period, among 400 pediatric patients with COVID-19, 126 children had coinfection. Children >10 years were the most affected age group. Underlying disease was present in 69%, hematological malignancies were the most common (17.5%). BC was detected in 76.9% (n=97), bacterial pneumonia (54.6%) was the main diagnosis, followed by oncologic patients with initial febrile neutropenia and posterior SARS-CoV-2 detection . Unusual BC as congenital syphilis w detected; acute appendicitis was the initial presentation of COVID-19 in 8 patients. VC was identified in 15.87% (n=20), prevailing rhinovirus (9.5%) and adenovirus (3.96%), One FC presented as proven pulmonary aspergillosis (0.8%). B-V and B-F coinfection were detected in 2 patients. Fever and cough were the most common symptoms, higher fever >40\u00b0C was mostly observed in the BC group (3%). Twenty-seven patients with BC (27.8%) were admitted to intensive care, with the OR 0.7 IR 95% (0.611\u20131.008), 4.1% died. One ICU admission was observed in the VC group (5%) and all VC cases resolved without complications.Pediatric patients with COVID-19 coinfection, especially BC were common in our center representing nearly one-third of the infected children, including unusual coinfections. BC was identified as a risk factor for ICU admission OR 0.7 IR 95% (0.611\u20131.008). Favorable outcomes were observed in most cases.All Authors: No reported disclosures."}
+{"text": "Bloodstream infection (BSI) following arterial aneurysm repair may signify vascular graft infection (VGI). The incidence of BSI and its implications in patients with arterial grafts have received scant attention with limited contemporary study. Therefore, the aim of the current investigation is to describe the incidence and epidemiology of BSI following arterial aneurysm repair in a population-based cohort.>18 years) residing in eight counties in southern Minnesota between January 1, 2010 and December 31, 2020. All repairs were performed at Mayo Clinic Rochester. Electronic health records were screened for initial BSI episode following aneurysm repair and complicating VGI.Retrospective case-cohort study using the expanded Rochester Epidemiology Project (e-REP) to identify all arterial aneurysm repairs performed in adults  overall, including 18.2 (95% CI 11.9-26.7) following open surgical repair and 10.2 (95% CI 5.8-16.6) following endovascular repair (p=0.148). Thirty-nine (92.9%) of the BSI cases were monomicrobial, with 18 (46.2%) due to Gram-positive organisms and 21 (53.8%) due to Gram-negative organisms. Of the 42 patients with BSI, 10 (23.8%) had VGI. VGI occurred in 8/18 (44.4%) monomicrobial Gram-positive BSI as compared to 2/21 (9.5%) monomicrobial Gram-negative BSI (p=0.013). The 3-year mortality rate following BSI was 52.3%.There was no statistically significant difference in incidence rates of BSI following open surgical repair and endovascular repair. Higher rates of VGI were seen with Gram-positive BSI as compared to Gram-negative BSI.Larry M. Baddour, M.D., Boston Scientific: Advisor/Consultant|Botanix Pharmaceuticals: Advisor/Consultant|Roivant Sciences: Advisor/Consultant|UpToDate, Inc.: Royalty payments - authorship duties."}
+{"text": "E. coli belonging to the serotype O177 is a rare strain found in ruminants, especially cattle. When compared to shiga toxin producing E. coli (STEC) O157 and non-O157 STEC  serotypes, the antimicrobial resistance, virulence factors, and genomic structure of E. coli O177 are poorly understood. Therefore, in this article, we present the whole genome sequence data of two aEPEC E. coli O177 isolates (E. coli O177_CF-154-A and E. coli O177_CF-335-B) generated using Illumina MiSeq platform. The raw data were generated, cleaned, and assembled using Trimmomatic and SPAdes. Genome data analysis yielded 5,112,402 and 5,460,435 bp, comprising contigs 101 and 191 with GC contents of 50.7% and 50.5% for E. coli O177_CF-154-A and E. coli O177_CF-335-B, respectively. Prokaryotic Genome Annotation Pipeline (PGAP) and Rapid Annotation using Subsystem Technology (RAST) showed that the complete genome of E. coli O177_CF-154-A contained 5040 coding sequences (CDS), 5146 genes, 4896 proteins, 90 RNAs, and 78 tRNA while that of E. coli O177_CF-335-B contained 5463 CDS, 5570 genes, 5230 proteins, 92 RNAs, and 80 tRNA for. A total of 426 and 425 subsystem features with 5190 and 5662 CDS were obtained for E. coli O177_CF-154-A and E. coli O177_CF-335-B, respectively. Several genes encoding virulence and antimicrobial resistance were identified in both genomes. Complete genome sequence data of both isolates have been deposited in the National Center for Biotechnology Information (NCBI), GenBank: accession numbers, VMKH00000000 (E. coli O177_CF-154-A) and VMKG00000000 (E. coli O177_CF-335-B). This data can be used as a reference for determining the virulence and antimicrobial resistance in E. coli O177 isolates from different sample sources.Atypical enteropathogenic  Moreover, these data give an extensive information on the virulence and antimicrobial resistance profile of this serotype, which may contribute to understanding and improving of scientific knowledge of this pathogenic strain.These data provide genomic features of \u2022E. coli O177 serotype from different environmental samples. In addition, these data can be used in public health to establish policy framework and strategy intended to curb antimicrobial resistance, especially in humans.The data may be used by researchers to develop new methods for detection of \u2022This genome can be used as a reference, especially for comparative genomic and epidemiological studies.1E. coli O177 isolates (E.\u00a0coli O177_CF-154-A and E. coli O177_CF-335-B) were obtained from cattle faeces in the North West province, South Africa (\u221227\u00b0 00\u2032 0.00\u2033 S 26\u00b0 00\u2032 0.00\u2033 E), E. coli O177_CF-154-A and E. coli O177_CF-335-B) are summarised in E. coli O177_CF-154-A and E. coli O177_CF-335-B, respectively. There were 5040 coding sequences (CDS), 5146 genes, 4896 proteins, 90 RNAs, and 78 tRNA for E. coli O177_CF-154-A genome, while E. coli O177_CF-335-B genome contained 5463 CDS, 5570 genes, 5230 proteins, 92 RNAs, and 80 tRNA. Furthermore, both genomes contained 2 CRISPR Arrays. Based on RAST annotation, there were 426 and 425 subsystem feature counts with 5190 and 5662 CDS in E. coli O177_CF-154-A and E. coli O177_CF-335-B, respectively. As depicted in Two atypical enteropathogenic 22.1E. coli O177 isolates were obtained from Antimicrobial Resistance and Phage Biocontrol Laboratory, Department of Microbiology. The isolates were selected based on the virulence and antimicrobial resistance profiles as described in the previous studies Two atypical enteropathogenic 2.2TM-Tissue MiniPrep Kit  following the manufacturer's instructions. The DNA concentration was determined using the NanoDropTM-Lite 1,000 spectrophotometer . After fragmentation, DNA libraries were constructed using the Nextera XT DNA library prep kit  following the manufacturer's instruction. The fragmented DNA was amplified using 12 cycles PCR, which adds the index sequences [index 1 (i7) and index 2 (i5)]. The PCR products were purified using 0.6\u00a0\u00d7\u00a0Agencourt AMPure XP beads (Beckman Coulter), and the quality was determined using 1.5% (w/v) agarose gel. Each library was diluted to 12 pmol. Samples were normalized to 4 nM using Nextra XT Library Normalization Beads (Illumina). Normalized libraries were pooled and 150 base paired-ends sequencing was performed with MiSeq Reagent V3 600-cycle kits on the Miseq instrument (Illumina).Genomic DNA was extracted from overnight cultures using the Zymo Research Genomic DNA2.3de novo genome assembly was carried out using SPAdes (v.3.13) 4  with all parameters  set at default Raw sequence data were generated and FASTQ files were obtained. The data were assessed for quality using FASTQC (v.0.11.5) and filtered for low quality reads and adapter regions using Trimmomatic (v.0.36) This study did not involve the use of human subjects or animal experiments.Peter Kotsoana Montso: Conceptualization, Methodology, Data curation, Writing \u2013 original draft, Visualization, Investigation, Software, Validation, Writing \u2013 review & editing. Victor Mlambo: Conceptualization, Methodology, Supervision, Software, Validation, Writing \u2013 review & editing. Collins Njie Ateba: Conceptualization, Methodology, Supervision, Software, Validation, Writing \u2013 review & editing.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper."}
+{"text": "Inonotus hispidus (Bull.: Fr.) P. Karst (IH) is an edible and medicinal parasitic mushroom. In this study, after a systematic analysis of its nutritional ingredients, the regulatory effects of IH on lipid metabolism were investigated in mice fed a high-fat diet (HFD). In HFD-fed mice, IH reversed the pathological state of the liver and the three types of fat and significantly decreased the levels of low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), triglycerides (TG), and leptin (LEP) and increased the level of high-density liptein cholesterol (HDL-C) in serum. Meanwhile, IH ameliorated liver damage by reducing alanine aminotransferase (ALT), aspartate aminotransferase (AST), interleukin (IL)-1\u03b2, IL-6, tumor necrosis factor-alpha (TNF-\u03b1), and plasminogen activator inhibitor-1 (PAI-1) levels in the liver and serum. Compared with HFD-fed mice, IH significantly modulated the gut microbiota, changed the relative abundances of microflora at different taxonomic levels, and regulated lipid levels. The results showed that 30 differential lipids were found. Results from Western blotting confirmed that IH regulated the nuclear factor erythroid-2 related factor 2 (Nrf2)/nuclear factor-kappa B (NF-\u03baB) signaling pathway and oxidative stress. This study aimed to provide experimental evidence for the applicability of IH in obesity treatment.Obesity is frequently associated with dysregulated lipid metabolism and lipotoxicity.  Obesity is a pathological condition requiring clinical intervention ; it may Obesity is frequently associated with dysregulated lipid metabolism and lipotoxicity , which mGrifola frondosa modulates ceramide levels and restores lipid metabolism by inhibiting the Toll-like receptor 4/NF-\u03baB signaling associated with inflammation and insulin resistance (IR) to treat obesity [Inonotus hispidus (Bull.: Fr.) P. Karst (IH) is an edible and medicinal parasitic mushroom belonging to the phylum Basidiomycota, class Agaricomycetes, and family Hymenochaetaceae. IH is mainly distributed in the Hebei Chengde, Shandong Linqing, Xiajin, and Xinjiang Aksu areas, and it prefers to live on mulberry, water willow, elm, poplar, and Japanese acacia. Previous investigations on IH have mainly focused on chemical composition analysis, artificial cultivation, mycelium fermentation, and pharmacological activity studies [Currently, weight loss occurs primarily through diet- and exercise-related strategies, which are often not sustainable . Bariatr obesity . Inonotu studies ,17,18,19 studies . IH petr studies . Extrace studies . HoweverIn this study, based on the detection of the main components of IH, combined with gut microbiota and lipid metabolomic analyses, the role of IH in regulating oxidative stress and inflammation, based on the Nrf2/NF-\u03baB signaling pathway, to alleviate obesity symptoms was explored in C57BL/6 mice with DIO. Our data provide experimental evidence for the applicability of IH in obesity treatment.IH fruiting bodies from Linqing, Shandong, were identified by Professor Yu Li. The general components , twenty kinds of amino acids, thirty-five kinds of fatty acids, seven kinds of minerals, six kinds of heavy metals, eight kinds of vitamins, and five kinds of nucleotides in IH were systematically detected, as we previously described ,23.n = 6/group), including an intragastrically HFD-fed group with 5 mL/kg of normal saline, an intragastrically simvastatin (SV) -treated group with 3 mg/kg of SV, and low- and high-dose intragastrically IH-treated groups with 500 and 1000 mg/kg of IH, respectively, daily for 8 weeks. The NCD mice were randomly divided into two groups (n = 6/group), including the vehicle-treated intragastrically NCD-fed group with 5 mL/kg of normal saline and the intragastrically IH-treated NCD-fed group with 500 mg/kg of IH daily for 8 weeks. Weekly measurements of body weight and plasma glucose levels were performed for all mice. After 8 weeks of drug treatment, the mice were euthanized using CO2 (the CO2 replacement rate was 30\u201370% of the container volume per minute). Peripheral blood was obtained by sampling the retro-orbital venous plexus. Organs , epididymal white adipose tissue (eWAT), inguinal white adipose tissue (iWAT), and perirenal white adipose tissue (pWAT) were dissected and weighed. The above tissue parts were stored at \u221280 \u00b0C for further biochemical analysis, and the remaining parts were fixed in a 4% tissue fixative  for subsequent pathological analysis.All experiments were performed in accordance with the guidelines of the Institutional Animal Ethics Committee of Jilin University (SY202106003). Thirty-six male C57BL/6JGpt mice (5 weeks old) from GemPharmatech Co., Ltd.  were maintained on either a normal chow diet  or a high-fat diet  under specific-pathogen-free (SPF) conditions on a 12 h light/dark cycle at a constant temperature (23 \u00b1 1 \u00b0C) and humidity (40\u201360%) for 8 weeks. To establish the DIO model for the high-fat diet study, 24 randomly selected mice were fed an HFD ad libitum during the entire experimental period. From week 9, the DIO mice were randomly divided into four groups . Serum was collected twice by centrifugation at 3500 rpm at 4 \u00b0C for 10 min. The levels of alanine aminotransferase (ALT) (MM-44625M1), aspartate aminotransferase (AST) (MM-44115M1), IL-1\u03b2 (MM-0040M1), IL-6 (MM-0163M1), TNF-\u03b1 (MM-0132M1), and plasminogen activator inhibitor-1 (PAI-1) (MM-0066M1) in the liver and serum, ROS (MM-43700M1), malondialdehyde (MDA) (MM-0897M1), and lysophosphatidylcholine (LPC) (MM-44698M1) in the liver, and high-density liptein cholesterol (HDL-C) (MM-44105M1), low-density lipoprotein cholesterol (LDL-C) (MM-43685M1), total cholesterol (TC) (MM-0632M1), triglycerides (TG) (MM-0631M1), and leptin (LEP) (MM-0622M1)  in the serum were measured using enzyme-linked immunosorbent assay (ELISA) kits.Liver tissue was homogenized in normal saline, and the protein concentration was determined using a Pierce\u2122 bicinchoninic acid (BCA) Protein Assay Kit  staining and Oil Red O staining were performed as described in our previous study . Fixed an = 3/group) was performed by 16S rRNA sequencing using an Illumina NovaSeq platform at Shanghai Personalbio Technology Co., Ltd. , as described in our previous study [A gut microbiota analysis of mouse cecum content in samples from NCD-, HFD-, and IH-treated groups (500 mg/kg) (us study . Sequencn = 3/group) was performed by liquid chromatography\u2013mass spectrometry (LC\u2013MS) at Shanghai Personalbio Technology Co., Ltd. , as described in our previous study [p \u2264 0.05) in metabolite levels and variable importance in projection (VIP) values \u2265 1.0 were regarded as the standard for differential lipids to filter out biomarkers.A plasma lipidome analysis of mouse serum from the NCD-, HFD-, and IH-treated groups (500 mg/kg)  buffer  containing protease and phosphatase inhibitors  and were homogenized using a high-throughput tissue grinder . After denaturation, 40 \u03bcg of protein was separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE)  and then transferred onto a polyvinylidene fluoride (PVDF) membrane . Membranes were blocked by Rapid Closure solution  and incubated with the primary antibodies overnight and then the secondary antibodies for 4 h at 4 \u00b0C. Finally, immunoreactive bands were visualized using an automated chemiluminescence image analysis system  and Ultra High Sensitivity enhanced chemiluminescence (ECL) kits . Protein expression levels were measured using ImageJ software  and normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Details regarding the antibodies used in this work are presented in p < 0.05.All values are presented as means \u00b1 SD. Biochemical indices were compared between different groups by one-way analysis of variance (ANOVA) followed by Tukey\u2019s test using BONC DSS Statistics 25 . Differences were considered statistically significant at The general nutritional composition of IH is 45.90% total dietary fiber, 25.50% total sugar, 15.90% protein, 9.90% ash, 9.22% total flavonoids, 6.18% moisture, 5.04% total polyphenols, 4.70% fat, 1.48% total triterpenes, 0.50% total saponins, 0.46% total alkaloids, 0.31% total sterols, etc. Among them, the total dietary fiber content was the highest. The glutamic acid content was the highest among the 20 amino acids detected. Seven minerals, including calcium (Ca), iron (Fe), zinc (Zn), selenium (Se), potassium (K), sodium (Na), and manganese (Mn), were detected, in addition to low concentrations of six heavy metals (lead (Pb), arsenic (As), mercury (Hg), cadmium (Cd), copper (Cu), and chromium (Cr)). Eight vitamins and five nucleotides were also detected . The corp < 0.05) (p < 0.05) (p < 0.001) (p < 0.01) (p < 0.01) (p < 0.05) (p < 0.05) (p < 0.001) without affecting other organs  A. Compar < 0.05) B and sup< 0.001) C, TC (p  < 0.01) D, and TG < 0.01) E in seru < 0.05) F. H&E st < 0.05) G. Among  < 0.05) . Compare < 0.05) . IH signr organs . IH alonr organs .p < 0.001) (p < 0.001) (p < 0.001) (p < 0.001) (p < 0.001) (p < 0.05) (p < 0.05). In the serum of HFD-fed mice, IH and SV showed the same effects on AST (p < 0.05) (p < 0.05) (p < 0.001) (p < 0.01) (p < 0.05) (p < 0.01) (p < 0.05). IH failed to influence the serum levels of ALT , high degrees of lipid deposition, and large amounts of lipid droplets (H&E staining) were improved by IH and SV A. Moreov< 0.001) B, AST (p< 0.001) C, IL-1\u03b2 < 0.001) D, IL-6 (< 0.001) E, and TN< 0.001) F in the  < 0.05) G in HFD- < 0.05) I, IL-6 ( < 0.05) K, TNF-\u03b1 < 0.001) L, and PA < 0.01) M. At 500 < 0.05) J and PAI < 0.01) M in HFD-s of ALT H. IH alos of ALT B\u2013M.Allobaculum, Adlercreutzia, Shigella, Dorea, Oscillospira, and Streptococcus and decreased Ruminococcus and Coprobacillus compared with the vehicle-treated HFD-fed mice  (p < 0.01)  D. IH and < 0.01) E.p < 0.01) (p < 0.05) (p < 0.001), IKK\u03b1 + \u03b2 (p < 0.001), and I\u03baB\u03b1  and significantly upregulated the expression levels of Nrf2 (p < 0.001), HO-1 (p < 0.01), and SOD-1 . Compared with the vehicle-treated NCD-fed mice, IH alone only increased the levels of Nrf2 (p < 0.01) and HO-1 (p < 0.05) and suppressed the activation of P-I\u03baB\u03b1 (p < 0.001) (The hepatic levels of ROS ( < 0.01) A and MDA < 0.05) B were de< 0.001) C.IH is rich in dietary fiber, which helps to improve hyperlipidemia by affecting lipid metabolism  and has Allobaculum, Dorea, and Oscillospira, facilitating the production of short-chain fatty acids (SCFAs) related to metabolic processes [Ruminococcus and Coprobacillus. SCFAs such as acetate, propionic acid, and butyric acid can reduce the generation of pro-inflammatory cytokines [Allobaculum can regulate hepatic lipid metabolic processes [Allobaculum plays a role in suppressing inflammatory responses by reducing the expression of p-IKK and TNF-\u03b1 [Dorea is negatively associated with inflammatory diseases [Oscillospira and Ruminococcus are inflammatory bacteria associated with inflammatory bowel disease [Oscillospira helps to maintain lipid homeostasis [Rosa Roxburghii Tratt, possessing hypolipidemic effects, can reduce the abundance of Coprobacillus [Streptococcus may be the main force for decomposing a large amount of cellulose [In obese mice, IH increased the abundance of the genera rocesses ,34,35 whytokines , displayytokines . SCFAs cytokines ,39. Dietytokines , which hytokines , Allobacrocesses , shows arocesses , and itsrocesses . Allobacnd TNF-\u03b1 , which hnd TNF-\u03b1 , Dorea idiseases . Both Os disease ,48, and eostasis . Rosa Robacillus . Moreoveellulose , which iellulose . Meanwhiellulose ,54, and ellulose .Metabolites of intestinal flora affect the process of lipid metabolism and host lipid composition . IH coulSOD and HO-1 [HO-1, a key target gene of Nrf2, exerts antioxidant effects by resisting endogenous and exogenous stimuli [Oxidative stress leads to increased lipid peroxidation and is closely associated with hyperlipidemia-related tissue damage ,63. Nrf2and HO-1 . SOD-1 iand HO-1 . HO-1, a stimuli . Nrf2/HO stimuli .Oxidative stress and inflammation are inextricably associated . Nrf2 caThe present study had certain limitations. In this study, we first reported the hypolipidemic effects of IH and analyzed the components involved. However, the specific active components possessing hypolipidemic activity were not confirmed; thus, further investigation is needed.In conclusion, IH regulated lipid metabolism through the Nrf2/NF-\u03baB signaling pathway, which is closely related to oxidative stress and inflammatory responses, in HFD-fed mice. Our study provides insights into the application of IH as a hypolipidemic agent and will facilitate its commercial application."}
+{"text": "In: Public health round-up. Bull World Health Organ. 2022 May 1; 100(5):296\u2013297, on page 297, middle column, the second sentence should read as follows: \u201cAs of 15 March, a total of 53 people were reported with suspected Yellow fever infections, six of whom died of the disease .\u201dhttps://dx.doi.org/10.2471/BLT.22.010522Public health round-up. Bull World Health Organ. 2022 May 1;100(5):296\u2013297."}
+{"text": "Serrated lesions (SLs), including sessile serrated lesions (SSL) and traditional serrated adenomas (TSA) have become subject of increased interest for their role as CRC precursors.Study aim was to evaluate the risk to develop total metachronous advanced neoplasia (T-MAN) at follow-up in patients with index SL compared to a matched cohort without SL.Patients 45-74y with SLs were identified through pathology database search. SL patients were matched 2:1 by sex; age; synchronous polyps ; timing of index, to patients without SL. Primary outcome was risk of T-MAN (advanced adenoma or high-risk SL) at follow-up. Secondary outcomes included risk of T-MAN stratified by synchronous polyps and SL characteristics.1425 patients were included . The SL group had greater risk of T-MAN compared to the non-SL group [Hazard-ratio (HR)=6.12 3.91-9.58)]. Patients with SL+HRA had higher risk of T-MAN compared to HRA alone [HR=2.62 (95%CI 1.45-4.71)], as well as patients with SL+LRA compared to LRA alone [HR=7.03 (95%CI 2.78-18.44)], and SL without adenoma compared to no-adenoma [HR=14.87 (95%CI 6.51-33.95)]. Presence of proximal SSL (HR=9.30), large SSL (HR=17.87) and proximal large SSL (HR=24.99), but not distal SSL, was associated with greater risk for T-MAN.Patients with SLs are at greater risk for developing T-MAN regardless of synchronous adenomas. Patients with SL and HRA, and those with large or proximal SSLs appear to be at greatest risk for T-MAN.OtherACGR. Djinbachian Grant / Research support from: Grant from the American College of Gastroenterology for the conduction of this project, M.-L. Lafontaine: None Declared, J. Anderson: None Declared, H. Pohl: None Declared, T. Dufault: None Declared, M. Boivin: None Declared, M. Bouin: None Declared, D. von Renteln Grant / Research support from: Daniel von Renteln is supported by a \u201cFonds de Recherche du Qu\u00e9bec Sant\u00e9\u201d (FRQS) career development award and has received research funding from ERBE, Ventage, Pendopharm, Fujifilm, and Pentax., Consultant of: Boston Scientific and Pendopharm,"}
+{"text": "Correction to: BMC Infectious Diseases\u00a0(2022)\u00a022, 897. https://doi.org/10.1186/s12879-022-07887-1Error 1:Following publication of the original article , the autError 2:Following publication of the original article , the aut"}
+{"text": "Candida auris is a globally spreading yeast pathogen causing bloodstream infections with high mortality in critically ill patients. The inherent antifungal drug resistance of most C. auris isolates and threat of multidrug-resistant strains create a need for adjunct immunotherapeutic strategies. While C. albicans candidemia was shown to induce immune paralysis and activation of inhibitory immune checkpoints, in vivo data on host responses to C. auris bloodstream infection are lacking as is an immunocompetent murine infection model to study the immunopathology and immunotherapy of C. auris sepsis. Therefore, herein, we developed an immunocompetent C. auris sepsis model by intravenously infecting C57BL/6 mice with 1.5\u2009\u00d7\u2009108 to 8\u2009\u00d7\u2009108 yeast cells of aggregate-forming (AR-0384) and nonaggregative (AR-0381) C. auris reference isolates. Both isolates caused reproducible, inoculum-dependent increasing morbidity, mortality, and fungal burden in kidney tissue. Notably, morbidity and mortality outcomes were partially decoupled from fungal burden, suggesting a role of additional modulators of disease severity such as host immune responses. Flow cytometric analyses of splenic immune cells revealed significant upregulation of the programmed cell death protein 1 (PD-1) on T cells and its ligand PD-L1 on macrophages from mice infected with C. auris AR-0384 compared to uninfected mice. PD-L1 expression on macrophages from AR-0384-infected mice strongly correlated with fungal tissue burden (Spearman\u2019s rank correlation coefficient [\u03c1] = 0.95). Altogether, our findings suggest that C. auris sepsis promotes a suppressive immune phenotype through PD-1/PD-L1 induction, supporting further exploration of PD-1/PD-L1 blockade as an immunotherapeutic strategy to mitigate C. auris candidiasis.IMPORTANCE Health authorities consider Candida auris to be one of the most serious emerging nosocomial pathogens due to its transmissibility, resistance to disinfection procedures, and frequent antifungal drug resistance. The frequency of multidrug-resistant C. auris isolates necessitates the development of novel therapeutic platforms, including immunotherapy. However, in vivo data on host interactions with C. auris are scarce, compounded by the lack of reliable immunocompetent mammalian models of C. auris candidemia. Herein, we describe a C. auris sepsis model in immunocompetent C57BL/6 mice and demonstrate reproducible and inoculum-dependent acute infection with both aggregate-forming and nonaggregative reference isolates from different clades. Furthermore, we show that C. auris sepsis induces upregulation of the PD-1/PD-L1 immune checkpoint pathway in infected mice, raising the potential of a therapeutic benefit of immune checkpoint blockade. Our immunocompetent model of C. auris sepsis could provide a facile preclinical platform to thoroughly investigate immune checkpoint blockade and combination therapy with antifungals. Candida auris  isolate bank   , which rre (MSS)  was quanre (MSS) \u201314, with10.1128/mSphere.00817-21.1TEXT\u00a0S1Candida auris inoculums and infection of mice, determination of fungal burden in kidney tissue, isolation of murine splenocytes, and fluorescent labelling of splenocytes. Download Text S1, DOCX file, 0.03 MB.Detailed descriptions of experimental procedures for preparation of Copyright \u00a9 2022 Wurster et al.2022Wurster et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 8 AR-0381 cells remained sublethal, and the mice developed only mild signs of distress  (8 and 8\u2009\u00d7\u2009108) of AR-0381 cells caused increasing morbidity  and 7-day mortality (33% and 44%)   and B. Iand 44%)  and B. Tand 44%) . Likewisectively .8 yeast cells of the aggregate-forming isolate AR-0384 caused robust morbidity  and 17% 7-day mortality     and B. Irs, 2.0)  and B. Hrs, 2.0) . This trrs, 2.0) , suggestC. auris infection (in vitro and in vivo studies yielded divergent (strain- and inoculum-dependent) results regarding the comparative pathogenicity and immunopathology of aggregate-forming and nonaggregative C. auris strains  and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) on T cells, natural killer (NK) cells, and natural killer T (NKT) cells was determined by flow cytometry as described in C. auris-infected mice harbored significantly higher frequencies of PD-1-positive T cells than uninfected mice  (To test whether =\u20090.015)  and C. I=\u20090.015) .C. auris-infected mice than in controls  (P\u2009=\u20090.01)   and C. I\u2009=\u20090.01) . Collect in mice  and patiThe main limitations of this pilot study include testing of only one isolate per aggregation phenotype and a limited number of animals tested per condition. Furthermore, our study focused solely on the induction of coinhibitory immune checkpoint molecules and did not dynamically capture the net state of pro- and anti-inflammatory immune signals in the bloodstream and infected tissues.C. auris infection model in immunocompetent C57BL/6 mice and demonstrate the induction of coinhibitory immune checkpoint signals after C. auris bloodstream infection. These data provide a theoretical framework for PD-1/PD-L1 blockade as a potential immunotherapeutic strategy to mitigate C. auris candidiasis. We and others previously demonstrated a therapeutic benefit of the PD-1/PD-L1 pathway blockade in murine models of invasive mold infections (C. albicans sepsis (C. auris infection model in cost-efficient C57BL/6 mice could serve as a facile preclinical platform to study checkpoint inhibitors as an investigational therapy for C. auris sepsis. Furthermore, our immunocompetent model could complement the published cyclophosphamide-immunosuppressed and/or neutrophil elastase-deficient murine C. auris sepsis models (C. auris in different host backgrounds.Despite these limitations, we herein describe a simple and reproducible fections \u201322 and Cs sepsis , 23, 24.s models  and allo10.1128/mSphere.00817-21.2TABLE\u00a0S1Table\u00a0S1, DOCX file, 0.02 MB.Antibodies used for flow cytometry. Antibody solutions were prepared in 100 \u03bcL of flow cytometry buffer per sample. Download Copyright \u00a9 2022 Wurster et al.2022Wurster et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the"}
+{"text": "Correction to: Eur J Nucl Med Mol Imaging (2022) 49:632\u2013651https://doi.org/10.1007/s00259-021-05603-wThe authors regret a typo in the units assessing hyperglycaemia, which should be defined as > 8.9 mmol/L, and not as erroneously mentioned in the published version as > 8.9 nmol/L."}
+{"text": "Gram-negative pathogens with multidrug resistance (MDR) or difficult-to-treat resistance (DTR) are increasingly common, resulting in limited treatment options. These resistant pathogens are often isolated from the respiratory tract or bloodstream and can result in significant patient morbidity and mortality. Imipenem/relebactam is a combination of imipenem with relebactam, a \u03b2-lactamase inhibitor of class A and C \u03b2-lactamases. We evaluated the activity of imipenem/relebactam and comparators  against gram-negative MDR and DTR isolates that were collected from patients with lower respiratory tract (RTI) or bloodstream infections (BSI) in the United States (US).In 2018-2020, 24 clinical labs participated in the global SMART surveillance program in the US, and each collected up to 100 consecutive aerobic or facultative gram-negative pathogens per year from patients with RTI and 50 from BSI. MICs were determined using CLSI broth microdilution and interpreted with CLSI breakpoints.P. aeruginosa (n=2018), 11.9% of E. coli (n=1952), 13.1% of K. pneumoniae (n=1080), 24.1% of E. cloacae complex (n=460), 19.7% of K. aerogenes (n=304), and 6.1% of S. marcescens isolates (n=304); DTR isolates were found among 7.1%, 0.1%, 2.4%, 1.5%, 0%, and 0.8%, respectively. Imipenem/relebactam was active against 61% of MDR P. aeruginosa, 43-53 percentage points higher than the studied comparator \u03b2-lactams, and against 49% of DTR P. aeruginosa, which were nonsusceptible to all studied commonly used \u03b2-lactams and levofloxacin (Table). Imipenem/relebactam was also active against 87-100% of MDR Enterobacterales, including against 97% of MDR K. pneumoniae, 18-96 percentage points higher than the studied comparator \u03b2-lactams, and against 85% of DTR K. pneumoniae.MDR isolates were found among 13.5% of collected in vitro data, imipenem/relebactam represents a promising treatment option in the US for patients with RTI or BSI caused by highly resistant gram-negative pathogens that pose substantial treatment challenges.Based on these Fakhar Siddiqui, MD, MBA, Merck & Co., Inc.: employee|Merck & Co., Inc.: Stocks/Bonds Karri A Bauer, PharmD, Merck & Co., Inc. Merck Research Laboratories: Stocks/Bonds Charles A. DeRyke, PharmD, Merck & Co., Inc. Merck Research Laboratories: Stocks/Bonds Katherine Young, M.S., Merck & Co., Inc.: Stocks/Bonds."}
+{"text": "Chronic inflammation, a hallmark of type 2 diabetes (T2D) and adiposity, increases the risk for age-related co-morbidities, including physical frailty. This study aimed to examine the effects of a mobile technology-enhanced behavioral lifestyle educational intervention on frailty and associated clinical, inflammatory, and laboratory outcomes in overweight older adults with T2D. Twenty participants  with T2D were recruited to complete a single-arm 6-month lifestyle intervention modified from the Action for Health in Diabetes (Look AHEAD) study, enhanced with technology for self-monitoring of diet and physical activity. Clinical assessments were collected at baseline and end of the study and analyzed with paired t-tests. Inflammatory cytokines  were quantified using a commercially available multiplex kit . Clinical lab analysis was performed by Quest Diagnostics . Eighteen participants completed the study . At baseline, 13 participants were pre-frail, 4 were frail based on Fried frailty criteria. Inflammatory cytokines and liver enzymes values were within normal limits. At follow-up, the following outcomes significantly improved: frailty score -44% (p=0.01), BMI -3% (P< 0.00), alanine transaminase -18% (p=0.03), aspartate aminotransferase -13% (p< 0.00), IL2 -33% (p=0.01), IL4 -31% (p=0.05), IFN\u03b3 -36% (p=0.03), GM-CSF -43% (p=0.03). No other significant differences were observed. The results suggest the efficacy of a technology-enhanced lifestyle intervention on frailty and associated clinical, inflammatory, and laboratory outcomes in overweight older adults with T2D."}
+{"text": "Ecdyonurusaurasiussp. nov., a micro-endemic species reported from several streams within the Aur\u00e8s Mountains (north-eastern Algeria), is described and illustrated at nymphal, subimaginal and imaginal stages of both sexes. Critical morphological diagnostic characters distinguishing the new species are presented, together with molecular affinities as well as notes on the biology and distribution of the species. Ecdyonurus Eaton, 1868 belongs to the Ecdyonurinae Ulmer, 1920, a subfamily with rather challenging and controversial taxonomy as genera delineation and phylogeny are still partially unsolved or in process  and Helvetoraeticus Bauernfeind & Sold\u00e1n, 2012 (15 species), according to the arrangement of setae on the superlingua, the number of bristles on the ventral side of the labrum and the number of comb-shaped bristles on the maxilla in nymphs, as well as the shape of the apical sclerite of the male genitalia.Ecdyonurusrothschildi Nav\u00e1s, 1929 and Ecdyonurusifranensis Vitte & Thomas, 1988, whereas one remains doubtful: Ecdyonurusvenosusvar.constantinicus Lestage, 1925, and the presence of Ecdyonurusvenosus  mentioned by Ecdyonurus.Currently, four taxa of this genus are reported from North Africa . Two of Ecdyonurusrothschildi from an oasis in Biskra Province, north-eastern Algeria, based on a male imago. The species was redescribed by E.aurantiacus  species group. Later, E.rothschildi, E.dispar  and E.aurantiacus nymphs. The species is now known from all Maghreb countries and is one of the most widespread species . We collected and reared fresh material at all stages. After critical observations and comparison with other Ecdyonurus species, we have clearly distinguished a new Algerian endemic species.The present study aims to examine BNP) and the Western Aur\u00e8s Massif   substitution model . Two independent analyses of four MCMC chains run for one million generations with trees sampled every 1000 generations were implemented, and 100 000 generations were discarded as a burnin after visually verifying run stationarity and convergence in Tracer ver. 1.7.2  and included 25% of parsimony informative sites. The COI gene tree grouped the five sequences of Ecdyonurusaurasius sp. nov. into a well-supported monophyletic clade, and was supported as a distinct species in the ASAP analysis  to 20.1% (mean distance to E.aurantiacus), with a minimum distance of 7.1% between GBIFCH01119302 / GBIFCH00673192 and EC-CH0 sequences.The Ecdyonurinae Ulmer, 1920Taxon classificationAnimaliaEphemeropteraHeptageniidae\ufeffDambri, Benhadji & Sartorisp. nov.11A25021-4118-5EC5-B4CF-3CFE86311118https://zoobank.org/0A552D79-3329-4CCA-9724-D01492F82D7BHolotype. Algeria \u2022 male imago in ethanol, with its corresponding nymphal and subimaginal exuviae, Wilaya de Batna, Charchar, 35\u00b024'22\"N, 6\u00b023'21\"E, 1340 m. a.s.l., 09 Nov. 2021, B. Dambri coll. (GBIFCH01128855) [MZL] \u2022 Paratypes. 1 male imago, with its nymphal and subimaginal exuviae (GBIFCH01128846), 1 female imago, with its nymphal and subimaginal exuviae (GBIFCH01128858), [MZL]; 6 female imagos [IB-US], same data as holotype; 1 male imago, 1 male subimago [IB-US], 2 female imagos, 7 male subimagos [FEEL-UB2], 06 Nov. 2021; 1 male imago [IB-US], 1 male imago [FEEL-UB2], 20 Oct. 2021; 1 male imago, with its nymphal and subimaginal exuviae (GBIFCH01119304), 1 female imago, with its nymphal and subimaginal exuviae (GBIFCH01128861) [MZL], 17 Oct. 2021; 1 female imago with its subimaginal exuvia, 1 female subimago (GBIFCH01128849) [MZL], 15 Oct. 2021; 1 female subimago, 1 male subimago [IB-US], 1 male subimago (GBIFCH01128853) [MZL], 2 nymphs [FEEL-UB2], 10 Oct. 2021; 3 nymphs [IB-US], 2 nymphs (GBIFCH01128857), 1 nymph on slide (GBIFCH01119301) [MZL], 09 Oct. 2021; 7 nymphs [IB-US], 15 nymphs [FEEL-UB2], 18 Jun. 2020; 15 nymphs [IB-US], 18 nymphs [FEEL-UB2], 5 nymphs (GBIFCH01128850) [MZL], 3 Mar. 2020; same locality, B. Dambri coll; 10 nymphs (GBIFCH00832138), 2 nymphs on slide (GBIFCH00673191-GBIFCH00673192), 1 male imago (GBIFCH00673193), 1 male imago, 1 female imago, 2 female subimagos (GBIFCH00832125), 23 Jun. 2019, same locality, L. Kechemir coll. et leg. [MZL]Other paratypes. Algeria \u2022 Wilaya de Batna, Berbaga, 35\u00b024'01N, 6\u00b024'31\"E, 1445 m. a.s.l., 1 male imago, with its nymphal and subimaginal exuviae (GBIFCH01119302), 1 female subimago with its nymphal exuvia (GBIFCH01128848) [MZL], 5 Nov. 2021; 1 male imago (GBIFCH01128852), 2 nymphs (GBIFCH01128847), 1 nymph on slide (GBIFCH01119303) [MZL], 13 nymphs [IB-US], 5 nymphs [FEEL-UB2], 4 Nov. 2021; 1 male imago [IB-US], 12 nymphs [FEEL-UB2], 30 Nov. 2020; 1 nymph [IB-US], 16 nymphs [FEEL-UB2], 03 May 2020; 1 male imago [IB-US], 10 nymphs [FEEL-UB2], 02 Mar. 2020, B. Dambri coll. Algeria \u2022 Wilaya de Khenchela, Yabous, 35\u00b021'11\"N, 6\u00b038'35\"E, 1420 m. a.s.l., 2 female imagos [IB-US], 2 female subimagos, 3 nymphs [FEEL-UB2], 22 Oct. 2021; 1 female subimago [IB-US], 2 nymphs [FEEL-UB2], 1 female imago with its subimaginal exuvia, 1 female subimago (GBIFCH01128854) [MZL], 13-14 Oct. 2021; 1 male imago with its subimaginal exuvia GBIFCH01128851), 1 female imago with is subimaginal exuvia (GBIFCH01128845) [MZL], 12 Oct. 2021; 5 nymphs [IB-US], 1 female imago, 2 male subimagos, 6 nymphs [FEEL-UB2], 09 Oct. 2021; 4 nymphs [IB-US], 2 nymphs [FEEL-UB2], 1 nymph (GBIFCH01128859) [MZL], 20 Jul. 2020; 2 nymphs [IB-US], 19 nymphs [FEEL-UB2], 1 nymph (GBIFCH01128856) [MZL], 02 Jun. 2020; 1 female subimago with its nymphal exuvia [IB-US], 8 nymphs [FEEL-UB2], 09 May 2020; 1 female subimago [IB-US], 15 nymphs [FEEL-UB2], 08 Mar. 2020; 1 female subimago with its nymphal exuvia [IB-US], 3 nymphs [FEEL-UB2], 23 Feb. 2020, B. Dambri coll. Algeria \u2022 Wilaya de Batna, Inoughissen, 35\u00b016'42\"N, 6\u00b032'34\"E, 1670 m. a.s.l., 1 nymph (GBIFCH01128863) [MZL], 07 Jul. 2020; 1 nymph (GBIFCH01128865) [MZL], 18 Apr. 2020, B. Dambri coll.35\u00b033'03\"N, 6\u00b000'22\"E, 1262 m. a.s.l.,1 nymph [IB-US], 17 Jun. 2020; 3 nymphs [IB-US], 10 nymphs [FEEL-UB2], 1 nymph (GBIFCH01128860) [MZL], 20 Apr. 2020, B. Dambri coll. Algeria \u2022 Wilaya de Batna, Bouailef, 35\u00b037'01\"N, 6\u00b011'17\"E, 1060 m, 1 nymph [IB-US], 08 Mar. 2020, B. Dambri coll.Algeria \u2022 Wilaya de Batna, oued Cha\u00e2ba, Aurasiusmons; aurasius is a noun in apposition.Aur\u00e8s mountains were coined by the Berber people as Awras, meaning tawny; translated by the Romans as Male imago Size: body length: 9.0\u20139.8 mm; forewing length 9.1\u201310.9 mm; cerci broken. General body color distinctly brown to reddish-brown . The new species presents more affinities with the two other North African endemics but can be distinguished from E.rothschildi by the much longer pronotal projections, the shape of the stout setae on the dorsal surface of femora (pointed in the latter), the shape of the gills  and the shape of the glossae ((inner margin rounded and convex in E.rothschildi). Ecdyonurusaurasius sp. nov. differs from E.ifranensis by the shape of the labrum (less broad in E.ifranensis), the shape of the stout setae on the dorsal surface of femora (pointed in E.ifranensis), and the shape of the glossae similar to E.rothschildi. In males, E.aurasius sp. nov. differs from E.rothschildi, E.dispar and E.aurantiacus by the compound eyes separated and not touching (character not stated in E.ifranensis description), from E.aurantiacus and E.dispar by the posterior margin of the basal sclerite smooth, and from E.ifranensis by the first transversal vein in the costal field surrounded by a dark brown maculation (the same in E.rothschildi), and by the shape of the posterior margin of the basal sclerite rounded (straight in E.ifranensis). It is also worth noting that E.aurasius sp. nov. differs from the two other North African species by the nervous ganglia tinted in purple in female imagos, whereas they are colorless in E.rothschildi and E.ifranensis.By the shape of the penis lobes and the posterolateral projections of the abdomen, E.dispar . The nymEcdyonurusaurasius sp. nov., as known so far, is restricted to the Aur\u00e8s region. The species has been recorded from only six localities in the Western Aur\u00e8s area; most habitats are located in the highest part of the streams, within altitudes ranging from 1010 to 1800 m a.s.l. These sites are represented by small mountain watercourses with gravel substrate  and the lowest one was observed at the Bouailef site .ate Fig. . The aveEphemeroptera species sporadically occurring in the same sites were Caenisluctuosa , Baetischelif Soldan, Godunko & Thomas, 2005 and Baetissinespinosus Sold\u00e1n & Thomas, 1983.The mature nymphs and subimagos (together with early-instar nymphs) were observed in May/June and another generation observed in September/October, thus suggesting a bivoltine life cycle. The other"}
+{"text": "Brain metabolic-sensory targets for modulatory glucose-sensitive endocrine and neurochemical signals remain unidentified. A hypothalamic astrocyte primary culture model was here used to investigate whether glucocorticoid receptor (GR) and noradrenergic signals regulate astrocyte glucose  and/or energy (5\u2032-AMP-activated protein kinase [AMPK]) sensor reactivity to glucoprivation by sex. Glucose-supplied astrocytes of each sex showed increased GLUT2 expression after incubation with the GR agonist dexamethasone (DEX) or norepinephrine (NE); DEX plus NE (DEX/NE) augmented GLUT2 in the female, but not in male. Glucoprivation did not alter GLUT2 expression, but eliminated NE regulation of this protein in both sexes. Male and female astrocyte glucokinase profiles were refractory to all drug treatments, but were down-regulated by glucoprivation. Glucoprivation altered AMPK expression in male only, and caused divergent sex-specific changes in activated, i.e., phosphoAMPK (pAMPK) levels. DEX or DEX/NE inhibited  or stimulated  AMPK and pAMPK proteins in both glucose-supplied and -deprived astrocytes. In male, NE coincidently up-regulated AMPK and inhibited pAMPK profiles in glucose-supplied astrocytes; these effects were abolished by glucoprivation. In female, AMPK profiles were unaffected by NE irrespective of glucose status, whereas pAMPK expression was up-regulated by NE only during glucoprivation. Present outcomes document, for each sex, effects of glucose status on hypothalamic astrocyte glucokinase, AMPK, and pAMPK protein expression and on noradrenergic control of these profiles. Data also show that DEX and NE regulation of GLUT2 is sex-monomorphic, but both stimuli impose divergent sex-specific effects on AMPK and pAMPK. Further effort is warranted to characterize mechanisms responsible for sex-dimorphic GR and noradrenergic governance of hypothalamic astrocyte energy sensory function. AMPK5\u2032-AMP-activated protein kinaseCaMKK\u03b2calcium/calmodulin-dependent protein kinase kinase-betaDEXdexamethasoneGCKglucokinaseGLUT2glucose transporter-2IIHinsulin-induced hypoglycemiaNEnorepinephrinepAMPKphosphoAMPKPP1protein phosphorylase-11The glucose-regulatory neural circuitry monitors cellular metabolic status and systemic energy substrate storage to exert appropriate control of central and peripheral effector motor functions \u20135. Many l-lactate for trafficking to neurons  and F = 19.86, p < 0.001; glucose status main effect: F = 16.63, p = 0.001; treatment main effect: F = 34.84, p < 0.001; glucose status/treatment interaction: F = 5.97, p = 0.006]) and GCK  = 15.22, p < 0.001; glucose status main effect: F = 95.43, p < 0.001; treatment main effect: F = 2.49, p = 0.097; glucose status/treatment interaction: F = 1.20, p = 0.342] and F = 6.70, p = 0.001; glucose status main effect: F = 36.47, p < 0.001; treatment main effect: F = 3.25, p = 0.050; glucose status/treatment interaction: F = 0.220, p = 0.881]) protein expression in each sex. For each sex, treatment groups including astrocytes supplied with 5.5\u2009mM glucose (G5.5), at left, are illustrated by gray bars, while glucose-deprived (G0) treatment groups, at right, are shown in white. Data indicate that DEX or NE alone each significantly increased GLUT2 levels in glucose-supplied male and female astrocytes . DEX and NE co-incubation (cross-hatched gray bars) reduced  or elevated  GLUT2 content relative to vehicle controls. In each sex, GLUT2 protein profiles were unaffected by glucoprivation (G0/V [solid white bars] versus G5.5/V). In glucose-deprived male and female astrocytes, GLUT2 profiles were elevated by DEX , but were refractory to NE ; DEX plus NE treatment (cross-hatched white bars) increased GLUT2 expression to levels measured after incubation with DEX alone. In each sex, hypothalamic astrocyte GCK content was decreased in response to glucoprivation, but was altered by any drug treatment when glucose was present or absent.Research outcomes described below were obtained using a characterized hypothalamic primary astrocyte culture model to address the question of whether hypothalamic astrocyte primary cultures established from male and/or female rats express glucose-sensitive plasma membrane and glycolytic pathway glucose sensors, and these nutrient gauges are regulated by glucocorticoid or noradrenergic input alone or by interaction of these regulatory stimuli. F = 37.09, p < 0.001; glucose status main effect: F = 38.86, p < 0.001; treatment main effect: F = 54.84, p < 0.001; glucose status/treatment interaction: F = 18.78, p < 0.001] and F = 7.32, p = 0.001; glucose status main effect: F = 15.07, p = 0.001; treatment main effect: F = 11.24, p < 0.001; glucose status/treatment interaction: F = 0.82, p = 0.499]) and pAMPK  = 19.16, p < 0.001; glucose status main effect: F = 13.28, p = 0.002; treatment main effect: F = 29.35, p < 0.001; glucose status/treatment interaction: F = 10.95, p < 0.001] and F = 21.65, p < 0.001; glucose status main effect: F = 3.68, p = 0.067; treatment main effect: F = 42.89, p < 0.001; glucose status/treatment interaction: F = 6.39, p = 0.002]) expression. Results show that glucose-supplied male astrocytes exhibited reductions in AMPK expression in response to DEX or NE alone; this negative response was exacerbated by DEX plus NE treatment. In the female, AMPK levels were elevated by DEX given alone or together with NE, but this protein was unaffected by NE alone. Glucose withdrawal suppressed AMPK content in male, but not female astrocytes. DEX alone or DEX plus NE decreased or stimulated AMPK expression in male versus female glucose-deprived astrocytes, respectively. Astrocyte pAMPK levels were diminished  or augmented  by DEX, according to sex. NE stimulated or had no effect on this protein profile in male versus female, respectively. In each sex, pAMPK content was affected similarly by DEX alone versus DEX plus NE treatment. Glucoprivation correspondingly up-  or down-  regulated pAMPK expression in male and female astrocytes. In glucose-deprived male glial cells, pAMPK profiles were refractory to DEX or NE, but were inhibited by DEX plus NE. Glucose-deprived female astrocytes, on the other hand, showed elevated pAMPK expression in response to all three treatments with the greatest magnitude of increase caused by DEX alone.The ultra-sensitive energy gauge AMPK monitors the cellular AMP/ATP ratio. Current work examined the premise that hypothalamic astrocyte total AMPK protein and/or activated, e.g., phosphorylated AMPK (pAMPK) protein profiles are affected by glucose withdrawal in a sex-specific manner, and the related question that AMPK activation may be differentially controlled by DEX or NE when glucose is present versus absent. Data presented in F = 7.26, p = 0.001; glucose status main effect: F = 6.05, p = 0.026; treatment main effect: F = 8.61, p = 0.001; glucose status/treatment interaction: F = 6.32, p = 0.005) or female  = 11.13, p < 0.001; glucose status main effect: F = 8.34, p = 0.011; treatment main effect: F = 19.51, p < 0.001; glucose status/treatment interaction: F = 3.67, p = 0.035) astrocyte CaMMK\u03b2 protein profiles. Data reveal that CaMMK\u03b2 expression in glucose-supplied astrocytes of either sex was insensitive to DEX or NE treatment alone or together. NE inhibited this protein in glucose-deprived male cells; this decline was amplified by combinatory NE plus DE treatment. In female astrocytes incubated without glucose, CaMMK\u03b2 content was increased by exposure to either DEX or DEX plus NE. F = 9.63, p < 0.001; glucose status main effect: F = 32.24, p < 0.001; treatment main effect: F = 2.05, p = 0.147; glucose status/treatment interaction: F = 8.34, p = 0.005) or female  = 6.82, p = 0.001; glucose status main effect: F = 37.94, p < 0.001; treatment main effect: F = 3.09, p = 0.057; glucose status/treatment interaction: F = 0.19, p = 0.903) astrocyte PP1 protein levels. NE alone had divergent effects on PP1 expression in male glucose-supplied (increased) versus glucose-deprived (decreased) astrocytes. NE plus DEX had no effect or suppressed PP1 profiles in male astrocytes when glucose was present or absent, respectively. Female astrocyte PP1 content was diminished during glucoprivation, but was insensitive to any drug treatment.AMPK activity is regulated in part by the upstream stimulatory kinase enzyme CaMMK\u03b2 as well as the inhibitory phosphatase PP1. It was of interest here to investigate whether action of DEX alone or in coordination with NE controls expression of these enzyme proteins in hypothalamic astrocytes. F = 17.04, p < 0.001; glucose status main effect: F = 38.47, p < 0.001; treatment main effect: F = 25.59, p < 0.001; glucose status/treatment interaction: F = 1.35, p = 0.295) or female  = 5.14, p = 0.001; glucose status main effect: F = 6.70, p = 0.016; treatment main effect: F = 5.44, p = 0.005; glucose status/treatment interaction: F = 4.31, p = 0.014) astrocyte GR protein expression. Data show that GR protein was decreased in G5.5 male astrocytes after incubation with DEX, NE, or DEX plus NE, and that the magnitude of this reduction was greatest in response to the latter combinatory dosing. Meanwhile, GR expression in female G5.5 astrocytes was unaffected by any of these treatments. Glucoprivation did not modify GR expression in astrocytes of either sex. Treatment with either DEX or DEX plus NE resulted in diminished GR protein profiles in G0 male and female astrocytes.Research here addressed the issue of whether male and/or female rat hypothalamic astrocyte primary cultures are directly receptive to glucocorticoid control by means of GR expression, and if so, whether this receptor protein profile is sensitive to glucose availability and regulated by glucocorticoid and noradrenergic stimuli in the presence versus absence of glucose. F = 10.83, p < 0.001; glucose status main effect: F = 22.84, p < 0.001; treatment main effect: F = 9.43, p = 0.001; glucose status/treatment interaction: F = 8.23, p = 0.002] and F = 23.78, p < 0.001; glucose status main effect: F = 0.27, p = 0.608; treatment main effect: F = 52.99, p = <0.001; glucose status/treatment interaction: F = 2.40, p = 0.106]), \u03b12-AR  = 20.78, p < 0.001; glucose status main effect: F = 77.61, p < 0.001; treatment main effect: F = 14.40, p < 0.001; glucose status/treatment interaction: F = 8.23, p = 0.002] and F = 14.31, p < 0.001; glucose status main effect: F = 5.52, p = 0.032; treatment main effect: F = 15.74, p < .001; glucose status/treatment interaction: F = 15.80, p < 0.001]), and \u03b21-AR  = 3.09, p = 0.029; glucose status main effect: F = 7.39, p = 0.015; treatment main effect: F = 2.02, p = 0.152; glucose status/treatment interaction: F = 2.74, p = 0.077] and F = 7.45, p < 0.001; glucose status main effect: F = 5.86, p = 0.023; treatment main effect: F = 7.13, p = 0.001; glucose status/treatment interaction: F = 8.36, p = 0.001]) protein expression. Data show that DEX alone inhibited \u03b11-AR protein, but did not regulate \u03b12-AR or \u03b21-AR levels in glucose-supplied male astrocytes. However, DEX alone stimulated each of these AR variant proteins in the female. Glucoprivation did not alter \u03b11-AR, \u03b12-AR, or \u03b21-AR expression profiles in the male, but up-regulated \u03b12-AR and \u03b21-AR levels in female astrocytes. In glucose-deprived male astrocytes, DEX decreased \u03b12-AR content, while NE suppressed \u03b11-AR and \u03b12-AR proteins. DEX plus NE had an equivalent effect to DEX alone on \u03b11-AR expression, but had greater inhibitory effects on \u03b12-AR levels compared to DEX or NE alone. Female glucose-deprived astrocytes showed augmentation of \u03b11-AR and \u03b12-AR, but not by \u03b21-AR by DEX. DEX plus NE increased \u03b11-AR levels to an extent similar to DEX alone, but did not alter \u03b12-AR expression. Both NE alone and NE plus DEX inhibited \u03b21-AR profiles in female astrocytes deprived of glucose.A critical objective of current work was to examine whether glucose status controls AR variant protein expression in male and female hypothalamic astrocytes, and to determine if DEX and NE act alone or synergistically to regulate these proteins during glucose availability versus deprivation. 4in vivo regarding neural control of glucose homeostasis in each sex.The current project used a hypothalamic astrocyte primary culture model to address the premise that these glia express glucoprivic-responsive glucose and energy sensing molecular biomarkers, and that glucose-sensitive hormone (glucocorticoid) and/or neurotransmitter (noradrenergic) stimuli govern sensor reactivity to glucose deficiency in a sex-specific manner. Outcomes show that in astrocytes of each sex, membrane glucose sensor GLUT2 and glycolytic pathway sensor GCK protein profiles are controlled by GR and AR input or by glucose, respectively. Yet, expression and activity of the energy sensor AMPK in these cells is subject to sex-dimorphic regulation by DEX, NE, and glucose. Results provide novel evidence that glucose modulates noradrenergic control of astrocyte GLUT2 (both sexes), AMPK , pAMPK (both sexes), and \u03b11- and \u03b12-AR  proteins. Evidence that co-administration of NE and DEX to male, but not female astrocytes attenuates (GLUT2) or exacerbates , effects of DEX alone infers that intersection of GR and AR signaling may be sex-specific. Ongoing work seeks to elucidate mechanisms that underlie differential male versus female astrocyte responses to GR or NE stimulation. There is also a need for understanding of functional implications of glucocorticoid and noradrenergic regulation of hypothalamic astrocyte metabolic sensory functions Present outcomes document expression of the characterized glucose sensors GLUT2 and GCK by hypothalamic astrocytes, and show that GR and AR input regulates the former, but not the latter protein in each sex. Evidence here that DEX treatment augments GLUT2 profiles in astrocytes of either sex, despite dissimilar regulatory effects of DEX on male (decreased) versus female (unchanged) astrocyte GR expression, infers that glucocorticoids may control GR input volume as well as post-receptor signaling in a sex-specific manner. GLUT2 ostensibly employs distinctive structural features to perform glucose transport versus monitoring functions . Data heThe characterized DEX dosage used here down-regulated astrocyte AMPK and pAMPK in male, but stimulated these proteins in the female, revealing bi-directional, sex-specific glucocorticoid regulation of AMPK expression and activity state in this brain cell type. This divergent hormone action may involve, in part, dissimilar GR signal volume in the two sexes as GR protein expression declined in DEX-treated male astrocytes, but was unaffected by this treatment in the female. DEX may also impose sex-contingent control of downstream post-receptor signaling, transcriptional, and/or post-translational events that govern astrocyte AMPK and pAMPK protein profiles. As the current study focused on a single DEX dose, there would be benefit from future investigation of whether varying GR ligand concentrations elicit proportionate modifications in AMPK and pAMPK levels over all or a segment of a comprehensive, physiological-like dosage range. There also remains a need to assess whether suppressive versus amplifying stimulatory effects of DEX on AMPK and pAMPK expression are associated with specific hormone concentration(s). Our findings that glucoprivation attenuates DEX-induced augmentation of male astrocyte pAMPK content infer that metabolic modulation of GR-mediated action on AMPK activation state may occur in this sex only. Noradrenergic control of this astrocyte energy sensor is evidently also sex-specific as NE incubation suppressed total AMPK protein and stimulated pAMPK levels in male astrocytes, yet had no effect either on protein profile in the female. Notably, glucoprivation was observed to elicit a directional shift in NE control  on male astrocyte AMPK, and to promote a loss or gain of NE regulation of pAMPK profiles in male versus female, respectively. These data point to sex-dependent metabolic modulation of noradrenergic governance of hypothalamic astrocyte AMPK activity.Current results show that neither DEX nor NE regulates the upstream stimulatory kinase CaMKK\u03b2 in the presence of glucose, yet during glucoprivation DEX elevates this protein profile in the female, while NE suppresses CaMKK\u03b2 expression in the male. PP1 levels in glucose-supplied astrocytes of either sex are similarly refractory to DEX, whereas NE enhances this protein in the male only. Notably, the direction of noradrenergic control of PP1 expression in male astrocytes is altered, e.g., switches from stimulatory to inhibitory when glucose is withdrawn. These outcomes support the novel concept that astrocyte energy status may determine whether glucocorticoid and noradrenergic signals are able to regulate astrocyte AMPK activity via upstream enzymes that govern phosphorylation, e.g., during glucose deficiency. DEX treatment of glucose-deprived female astrocytes stimulated CaMKK\u03b2 expression without affecting PP1 profiles, coinciding with up-regulated pAMPK. Thus, in this sex, CaMKK\u03b2 is likely required for glucocorticoid augmentation of AMPK activity. Interestingly, NE did not modify either CaMKK\u03b2 or PP1 expression, but also increased pAMPK profiles, apparently by kinase/phosphatase-independent mechanisms. Glucose-deprived male astrocyte CaMKK\u03b2 and PP1 profiles were resistant or inhibited by DEX, respectively, yet pAMPK expression was unaffected by either stimulus, inferring that potential augmentation of pAMPK levels by noradrenergic diminution of PP1 may be counter-balanced by signals that blunt AMPK activation.It is intriguing to observe that glucoprivic-associated diminution of astrocyte GCK expression coincided with up- or down-regulated pAMPK protein in male versus female, respectively. These findings infer that in the latter sex, augmenting effects of diminished glucose catabolism on sensor activity are apparently offset by regulatory signals that attenuate AMPK phosphorylation and/or decreased allosteric activation of AMPK due to energy production from non-glucose substrates.Present data also show that NE did not alter any astrocyte AR variant protein profile in either sex, but \u03b11-AR (both sexes), \u03b12-AR , and \u03b21-AR  proteins are responsive to DEX. The observed efficacy of DEX to up-regulate all three AR variant proteins in the female may contribute to unique effects of combinatory DEX plus NE treatment on specific astrocyte target proteins in that sex. Glucose stability evidently modulates DEX regulation of specific AR proteins in each sex, as glucoprivation abolishes DEX inhibition of \u03b11-AR profiles and allows DEX to suppress \u03b12-AR levels in the male, and in the female eliminates DEX stimulation of \u03b21-AR expression. Glucoprivic governance of noradrenergic regulation of AR protein profiles is, on the other hand, sex-specific as this metabolic stress allows inhibitory noradrenergic control of \u03b11-AR and \u03b12-AR proteins in male hypothalamic astrocytes only.in vivo models of glucose-sensitive glucocorticoid and noradrenergic signaling to the hypothalamus to understand, for each sex, hormonal and neurotransmitter regulation of astrocyte sensing and communication of cellular energetic stability or imbalance.In summary, current research presents unique proof that hypothalamic astrocytes express membrane and glycolytic pathway glucose-sensory biomarkers in addition to the energy sensor AMPK . Results"}
+{"text": "Existing chemotherapy treatments for breast cancer patients are high on toxicity. There are very limited options available for triple-positive breast cancer (TPBC) patients, and there have not been any major breakthrough for targeted therapy for triple-negative breast cancer (TNBC) patients. Therefore, there is a need to identify common therapeutic targets for breast cancer patients. In this manuscript, we compared the sphingolipid profiles of cancer cell lines representing TPBC and TNBC, and correlated these profiles with the proliferation and migration properties the of cell types. We then associated the sphingolipid profiles for each subtype specific cell line with transcriptional and translational expression of corresponding metabolizing enzymes. Our results suggested that ceramide kinase (CERK) that catalyzes the synthesis of ceramide-1-phosphates from ceramides is dysregulated in both cell types. We also showed that the targeting of CERK at transcriptional level by siRNA therapeutics or inhibiting the CERK activity by hydrogel-mediated delivery of chemical inhibitors can be an effective strategy to slow down the tumor progression. Therefore, CERK emerges as a potential therapeutic target that can be explored further for cancer therapy.Sphingolipids are key signaling biomolecules that play a distinct role in cell proliferation, migration, invasion, drug resistance, metastasis, and apoptosis. Triple-negative (ER\u2212PR\u2212HER2\u2212) and triple-positive (ER+PR+HER2+) breast cancer  subtypes reveal distinct phenotypic characteristics and responses to therapy. Here, we present the sphingolipid profiles of BT-474 and MDA-MB-231 breast cancer cell lines representing the TPBC and TNBC subtypes. We correlated the level of different classes of sphingolipids and the expression of their corresponding metabolizing enzymes with the cell proliferation and cell migration properties of BT-474 and MDA-MB-231 cells. Our results showed that each cell type exhibits a unique sphingolipid profile, and common enzymes such as ceramide kinase  are deregulated in these cell types. We showed that siRNA/small molecule-mediated inhibition of CERK can alleviate cell proliferation in BT-474 and MDA-MB-231 cells, and cell migration in MDA-MB-231 cells. We further demonstrated that nanoparticle-mediated delivery of CERK siRNA and hydrogel-mediated sustained delivery of CERK inhibitor to the tumor site can inhibit tumor progression in BT-474 and MDA-MB-231 tumor models. In summary, distinct sphingolipid profiles of TPBC and TNBC representing cell lines provide potential therapeutic targets such as CERK, and nanoparticle/hydrogel mediated pharmacological manipulations of such targets can be explored for future cancer therapeutics. Lipid reprogramming has emerged as a key hallmark for cancer pathogenesis ,2. SphinCeramide, the central player in the sphingolipid pathway, induces apoptosis and cell senescence in response to chemotherapy and radiation therapy A 9,10].,10.9,10]+ tumors), and are categorized as triple-positive (TPBC) phenotypes . A. A21]. Aenotypes . Therefoenotypes . In contenotypes . Chemothenotypes ,28.Each breast cancer subtype has a unique molecular portrait that influences its response to therapy, prognosis, and outcome. Earlier studies have reported the sphingolipid profile in luminal (ER+PR+) and TNBC (ER\u2212PR\u2212HER2\u2212) subtypes ,30. Howe\u00ae, Saint Louis, MO, USA. Fetal bovine serum (Cat#10270) was purchased from, Waltham, MA, USA. Matrigel (Cat#354234) was purchased from Corning Inc., Bedford, MA, USA. Penicillin-Streptomycin solution (SV30079) was purchased from Cytiva HyCloneTM, South Logan, OH, USA. Trypsin (Cat#TCL007), MEM Medium (Cat#AL081) and sodium bicarbonate (GRM849) were purchased from HiMedia Laboratories, Mumbai, India.Materials. Human breast cancer cell lines BT-474 and MDA-MB-231 were purchased from American Type Culture Collection . DMEM (Cat#D5648), DPBS (Cat#D5652), Crystal Violet Dye (Cat#C0775) were purchased from Sigma\u2013AldrichTM DNase (Cat#AM2238) was from Invitrogen, Vilnius, Lithuania. iScriptTM cDNA synthesis kit (Cat#1708891) and iTaq\u2122 universal SYBR\u00aeGreen Supermix (Cat#1725124) were purchased from Bio-Rad Laboratories, Hercules, CA, USA. RNase Inhibitor (Cat#AM2694), and PierceTM BCA protein assay kit (Cat#23227) were purchased from Thermo Scientific. Complete\u2122, Rockford, IL, USA. Protease Inhibitor Cocktail (P8340-1ML) was purchased from Sigma\u2013Aldrich\u00ae, Saint Louis, MO, USA. Sigma. CerK siRNA (Cat#L-004061-00-0005) and scrambled siRNA (Cat# D-001810-10-05) were purchased from Dharmacon Inc., Cambridge, UK. Ceramide Kinase Inhibitor, NVP231 (Cat#3960) was purchased from Tocris Bioscience, Bristol, UK.Lipofectamine 2000 (Cat#11668019) and TURBO\u00ae Polar Lipids, Inc., Alabaster, OK, USA. LASS1 (CERS1) (Cat#H00010715-A01), LASS2 (CERS2) (Cat#H00029956-M01A), LASS4 (CERS4) (Cat#H00079603-M01), LASS6 (CERS6) (Cat#H00253782-M01) were purchased from Abnova, Taipei, Taiwan. LASS5 (CERS5) (Cat#ab73289), CerK (Cat#ab155061), SPHK1 (Cat#ab109522), SPHK2 (Cat#ab264042), A-SMase (SMPD1) (Cat#ab83354), N-SMase1 (SMPD2) (Cat#ab131330), N-SMase2 (SMPD3) (Cat#ab199399), N-SMase3 (SMPD4) (Cat#ab133935), UGCG (Cat#ab124296), SMS1 (Cat#ab135365), SMS2 (Cat#ab237681), GBA1 (Cat#ab88300), GLB1 (Cat#ab96239), B4GALT6 (Cat#ab200639), Goat anti-mouse IgG (L+H) (Cat#ab6789) were purchased from Abcam, Cambridge, UK. ASAH1 (Cat#HPA005468), ASAH2 (Cat#ABS457), \u03b2-actin (Cat#A5541) were purchased from Sigma\u2013Aldrich\u00ae, Saint Louis, MO, USA. Goat anti-rabbit IgG-HRP was purchased from (Cat#sc-2004) Santa Cruz Biotechnology, Inc., Dallas, TX, USA. Ceramide/Sphingoid Internal Standard Mixture II (Cat#LM6005-1EA) was purchased from AvantiN,N\u2032-methylene bisacrylamide (Cat#M7279), glycine (Cat#G8898), MTT (Cat#M5655), triethyl ammonium bicarbonate buffer (Cat#T70408), and iodoacetamide (144-48-9) were purchased from Sigma\u2013Aldrich\u00ae, Saint Louis, MO, USA. Tris (Cat#MB029), NaCl (Cat#GRM853), polyoxyethylenesorbitan monolaurate (Tween\u00ae 20) (Cat#P7949), glycerol (Cat#GRM1027), bovine serum albumin fraction-V (Cat#GRM105) were purchased from HiMedia Laboratories, Mumbai, India. Paraformaldehyde (PFA) (Cat 81847) was purchased from Thomas Baker, Mumbai, India. Ammonium hydroxide (Cat#16227) and hydrochloric acid (Cat#29505) was purchased from Thermo Fisher Scientific, Waltham, MA, USA. MOPS free acid (Cat#MB0360) and acrylamide (Cat#AB1032) were purchased from Bio Basics Inc., Markham, ON, Canada. Sodium hydroxide (Cat#13913) and potassium hydroxide (Cat#84749) was purchased from Sisco Research Laboratories Pvt. Ltd., Maharashtra, India. Developer (Cat#4908216), Fixer (Cat#4908232) and XBT X-Ray film (Cat#6568307) were purchased from Carestream, Rochester, NY USA. Transwell Migration Plate (Cat#3464) was purchased from Corning Inc., Bedford, MA, USA. Nitrocellulose (Cat#HATF00010), Immobilon Western Chemiluminescent HRP substrate (Cat#WBKLS0500), ethanol (Cat#100983), glacial acetic acid (Cat#193402), chymotrypsin (11418467001), DMSO (Cat#276855), ethidium bromide (Cat#E8751), agarose (Cat#A9539), magnesium chloride (Cat#208337), sodium dodecyl sulfate (Cat#L3771), ammonium persulfate (Cat#A3678), Methanol (Cat#34966), chloroform (Cat#25669-1L), 2-propanol (Cat#34965), acetonitrile (Cat#34967), ammonium acetate (Cat#14267-25G), formic acid (Cat#56302-50ML), water (Cat#39253-4L) and ammonium formate (Cat#14266-25G) were purchased from Honeywell International Inc., Charlotte, NC, USA. ACQUITY UPLC BEH Shield RP18 column (Cat#186002854) was purchased from Waters\u2122 Ltd., Milford, MA, USA.2 in humidified incubator. Cell culture. Human breast cancer cell lines BT-474 and MDA-MB-231 were cultured in DMEM high glucose media with 10% Fetal bovine serum, 100 units/mL penicillin, and 100 \u00b5g/mL streptomycin. Cells were grown at 37 \u00b0C with 5% CO5 per well) were seeded in six-well plate in DMEM media with 10% FBS and 10% penicillin and streptomycin, and incubated at 37 \u00b0C in CO2 incubator for 24 h. At 80\u201385% confluency, siRNA-lipofectamine complexes in 1:3 ratio were incubated for 25 min in MEM media, and cells were transfected with these complexes. After 6 h of transfection, media was removed, and cells were incubated with antibiotic-free media containing 10% FBS for 36 h. CERK silencing was confirmed by Western blotting.siRNA transfections in cell culture. BT-474 and MDA-MB-231 cells were transfected with siRNA (targeting CERK or scrambled) using Lipofectamine 2000. BT-474 and MDA-MB-231 cells  (5000 cells/well) were used, and cell proliferation assay was performed following previously described method [For transwell migration assay, BT-474 and MDA-MB-231 cells or siRNA transfected cells (scrambled or CERK siRNA) transfected cells  were seeded in transwell inserts (4 \u00b5m pore size). We then placed the inserts in 24-well cell culture plates containing DMEM with 10% FBS and incubated for 24 h at 37 \u00b0C. We fixed the migrated cells in 4% paraformaldehyde (PFA) (~5 min), and permeabilized with methanol (20 min). Cells were then stained with 2% crystal violet dye (15 min) followed by PBS washing to remove the extra stain on the cell surface. Cells were counted manually and imaged using a Nikon microscope. Pellet collection for RNA and protein isolation. For RNA isolation, cells were grown in 100 mm cell culture plates for 90% confluency. For pellet collection, media was aspirated from the plates and washed two times with DPBS. Trizol (1 mL) was added to the plate, and plates were incubated for 5 min. After incubation, cells were scraped and transferred into 1.5 mL centrifuge tubes, and were used immediately or stored at \u221280 \u00b0C. For protein isolation, cells were similarly washed, scraped out in DPBS, centrifuged at 5000 rpm for 5 min, and used immediately or stored at \u221280 \u00b0C.Quantitative Real-Time PCR. Total RNA extraction and qRT-PCR studies were performed using previously described method . All priWestern Blotting. Protein expression analysis was performed by Western blotting as per previously described method . ProteinIsolation and quantification of sphingolipids using LC-MS/MS. Collection of cell pellets, lipid isolation, LC-MS/MS analysis, and absolute quantitation of sphingolipids was performed as per our previously published method .Animal studies. Animal experimental protocols were reviewed and approved by the Institutional Animal Ethics Committee of Regional Centre for Biotechnology (RCB/IAEC/2021/96). The experiments were carried out as per the guidelines issued by Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), Govt. of India.6 cells were injected subcutaneously in the flank. Once the tumor was palpable (20\u201330 mm3), tumor volume and weight of the mice were recorded after every two days. Tumor volume was calculated as per the formula, L \u00d7 B2/2 where L is the length of tumor and B is breadth of tumor. At the end of the experiment, tumors were excised, washed with ice-cold 1X PBS, and stored in Allprotect tissue reagent at \u221280 \u00b0C for further analysis. All tumor growth kinetic experiments were performed in female NOD SCID CB.17 mice. Prior to the cell injection, hair was shaved from flank of the mice. For comparative tumor kinetics experiments, respective cell lines (BT-474 and MDA-MB-231) were resuspended in FBS: Matrigel , and 3 \u00d7 103) were randomized into different groups with each group having 4\u20136 mice. Group 1 mice were left untreated, group 2 mice were injected intratumorally with scrambled siRNA, and group 3 mice were treated with target siRNA. The siRNA was injected in a volume of 40 \u00b5L with 300 ng of siRNA complexed with TAC6 polymer  at the tumor site [For siRNA experiments, mice with palpable tumor  were randomized into different groups with each group having 4\u20136 mice. Group 1 mice were left untreated, and group 2 mice were implanted subcutaneously with inhibitor (NVP-231)-loaded hydrogel [For inhibitor experiments, mice with palpable tumor .Visualization of sphingolipid metabolic pathways. Expression of genes involved in sphingolipid metabolism were quantified using RT-PCR. Protein levels were probed by Western blots and quantified using ImageJ, followed by normalization against actin. Lipidomics experiments were performed to quantify the metabolite (sphingolipid) levels. Log2 (fold changes) were calculated for MDA-MB-231 against BT-474 cells. The visualization was made using drawio, a cross-platform JavaScript based open source drawing application available on GitHub at We selected BT-474 and MDA-MB-231 breast cancer cell lines representing TPBC and TNBC subtypes. To quantify the levels of sphingolipid species in these two cell lines, we collected cell pellets, and isolated the total lipids enriched in sphingolipids. We performed qualitative and quantitative sphingolipidomics with absolute quantitation of 27 sphingolipid species including ceramides, glucosylceramides, lactosylceramides, sphingomyelins, ceramide-1-phosphates (C1P), sphingosine, and sphingosine-1-phosphate (S1P) using LC-MS/MS [p < 0.01), C18:0 , C22:0 , C24:0 , and C24:1  were elevated in MDA-MB-231 cells in comparison to BT-474 cells (p < 0.05), C18:0 , C20:0 , C22:0 , C24:0 , and C24:1 , are downregulated in MDA-MB-231 cells as compared to BT-474 cells (p < 0.05) (p < 0.05) species as compared to BT-474 cells (p < 0.05), C18:0 , C20:0 , C22:0 , and C24:0 , are less abundant in MDA-MB-231 cells as compared to BT-474 cells (p < 0.05), C18:0 , C20:0 , C22:0 , C24:0  and C24:1 , are elevated in MDA-MB-231 cells (p < 0.01) increase in levels of S1P   and C24:74 cells . We also74 cells C. It is 31 cells D. We alss of S1P  in MDA-Ms of S1P . TherefoSPT1) , SPT2 , delta 4-desaturase, sphingolipid 1 and 2 (DEGS1) , DEGS2 , and 3-ketodihydrosphingosine reductase (KDSR)  as compared to BT-474 cells , CERS2 , CERS3 , CERS4 , CERS5 , and CERS6  in MDA-MB-231 cells as compared to BT-474 cells , ASAH2 , alkaline ceramidase 1 and 3 ACER1 , and ACER3  are significantly upregulated in MDA-MB-231 cells as compared to BT-474 cells (p < 0.0005) in the expression of sphingosine kinase 1 (SPHK1) and a ~3.3-fold (p < 0.0005) increase in SPHK2 as compared to BT-474 cells that, again, positively correlates to higher S1P levels  in MDA-MB-231 cells (p < 0.01) decrease in the expression of S1P lyase (S1PL) in MDA-MB-231 cells over BT-474 cells.Among salvage pathway genes, expression of 74 cells . This inP levels B. Furthe31 cells B that coSMPD1) (p < 0.0001), SMPD2 , SMPD3 , and SMPD4  in MDA-MB-231 cells as compared to BT-474 cells (SMS1) (p < 0.0001), and downregulation of SMS2 (p < 0.0001) in MDA-MB-231 cells  in glucosylceramide synthase (UGCG) and a >50-fold increase (p < 0.01) in the expression of B4GALT6 in MDA-MB-231 cells in comparison to BT-474 cells  in glucosylceramidase 1 (GBA1) expression, responsible for breakdown of glucosylceramides, but we could not detect galactosidase (GLB1) expression due to low transcript levels. High expression of B4GALT6, thereby, support high lactosylceramide levels in MDA-MB-231 cells as compared to BT-474 cells in spite of high GBA1 expression (We observed a >100.0-fold increase (74 cells D. In spipression D.p < 0.05), CERS4 , CERS5 , but there was >2.0-fold (p < 0.05) decrease in the expression of CERS6 in comparison to BT-474 cells  gene expression in MDA-MB-231 cells as compared to BT-474 cells but it is not translated to protein expression, that might be due to different post-transcriptional and post-translational regulatory changes (p < 0.001) in MDA-MB-231 as compared to BT-474 cells is very well correlated with gene expression data and lower levels of sphingomyelins in MDA-MB-231 cells (p < 0.05), SMS2  and lower levels of SMPD4  as compared to BT-474 cells  increase in the expression of ASAH1 as expected from gene expression data, whereas there was no significant change in ASAH2 protein expression (p < 0.05), SPHK1 , and SPHK2  proteins in MDA-MB-231 cells as compared to BT-474 cells (p < 0.01) increase in UGCG expression in MDA-MB-231 cells in comparison to BT-474 cells, but there was a >7.5-fold decrease (p < 0.05) in the expression of B4GALT6 (p < 0.05) and low expression of GLB1  in MDA-MB-231 cells as compared to BT-474 cells  proliferation at 72 h as compared to MDA-MB-231 cells (p < 0.001) increase in migration of MDA-MB-231 cells as compared to BT-474 cells  were plated in 96-well plate, and proliferation index was determined by MTT assay. We observed that BT-474 cells exhibit >2.5-fold higher  of BT-474 cells after 72 h (p < 0.05) decrease in the number of colonies in BT-474 cells (p < 0.001) in cell proliferation (p < 0.001) (p < 0.0001) on knockdown of CERK by siRNA in MDA-MB-231 cells  E, and ce31 cells F.p < 0.05) decrease in cell proliferation (p < 0.01) decrease in number of colonies over untreated cells (p < 0.0001) decrease in cell proliferation of MDA-MB-231 cells on treatment with 1.0 \u00b5M of NVP-231 (p < 0.0001) decrease in number of colonies (p < 0.0001) decrease in number of migrated cells  decrease in tumor volume on final day (p < 0.0001) decrease in tumor volume on final day  decrease in tumor volume with NVP-231-Gel treatment on the final day as compared to untreated tumors (p < 0.05) decrease in volume of MDA-MB-231 tumors on final day , and worst prognosis, shorter survival and higher recurrence rate were positively associated to higher CERK expression among ER-negative cancers ,43. CERKPolymeric and lipid nanoparticles have emerged as effective delivery vehicles for gene therapeutics . In our CERK appears to be playing different roles in more aggressive metastatic breast cancer cell lines versus non-metastatic cell lines targeting cell migration and cell proliferation in a context-dependent manner. Hence, CERK can be a potential target for multiple breast cancer subtypes. However, more studies elucidating the molecular mechanism by which CERK controls these phenotypes and the extent of overlap in the pathways regulating them needs to be performed."}
+{"text": "Infections caused by multidrug-resistant (MDR) bacteria are an increasingly common public health threat associated with worse outcomes in immunocompromised patients. Eravacycline (ERV) has potent in-vitro activity against MDR Gram-negative and Gram-positive bacteria and has demonstrated non-inferiority to meropenem in the phase III IGNITE4 trial; however, the trial excluded immunocompromised patients. We aimed to evaluate clinical and safety endpoints of immunocompromised patients receiving ERV as definitive therapy.Multicenter, retrospective, observational study conducted from October 2018 to April 2022. Adult hospitalized immunocompromised patients treated with ERV for \u226572 hours were included. Immunocompromised patients were defined as having any of the following: chemo or radiation therapy < 30 days of hospital admission, HIV/AIDS with CD4 < 200, chronic steroids  age was 62 (53-70) and 61.6% were male. Hospital length of stay was 28 (13-42) days and 67% were admitted to the intensive care unit. SOFA and APACHE II scores were 3.5 (1-7) and 16 (11-20), respectively. Common infection sources were intra-abdominal (26%) and lower respiratory tract (18%); 24% were bacteremic. Most patients had cultured Enterobacterales (58.7%) and Enterococci (37%) spp. infections. Of those, 21.3% were CRE and 19% were VRE. Infectious diseases consult was obtained in 91.8% of cases. Time elapsed from index culture collection to ERV initiation was 4 (2-8) days and duration of ERV therapy was 7 (4-12) days. In total, 81.3% of immunocompromised patients achieved 30-day survival and 90.7% did not have 30-day infection recurrence. Probable drug-related adverse events occurred in 5.3% of patients .A majority of immunocompromised patients receiving ERV as definitive therapy achieved 30-day survival and did not experience infection recurrence. ERV use in immunocompromised subpopulations will benefit from studies tailored to their specific characteristics.Kimberly C. Claeys, PharmD, BioFire Diagnostics: Honoraria Bruce M. Jones, Pharm.D., FIDSA, BCPS, AbbVie: Advisor/Consultant|AbbVie: Honoraria|La Jolla: Honoraria|Melinta: Advisor/Consultant|Paratek: Honoraria|Regeneron: Honoraria."}
+{"text": "Respiratory syncytial virus (RSV) is a public health burden; no vaccine is currently available. An mRNA-based RSV vaccine (mRNA-1345) encoding the RSV prefusion stabilized F (preF) glycoprotein is under clinical investigation.A phase 1, randomized, observer-blind, placebo-controlled, dose-ranging study assessed safety and immunogenicity of mRNA-1345 in younger adults  and older adults  (NCT04528719). YA and OA were randomized to receive 1 dose of mRNA-1345  or placebo.In all, 74 YA participants  and 202 OA participants  received study injections. mRNA-1345 was well-tolerated in both groups, with lower reactogenicity observed in OA vs YA at higher doses. Injection site pain was the most frequent local solicited adverse reaction . Erythema and swelling were less frequent . Overall, 57.9-100% (YA) and 53.2-78.7% (OA) of mRNA-1345 and 40.0% (YA) and 45.5% (OA) of placebo groups reported \u2265 1 systemic SAR, most commonly headache, fatigue, myalgia, and arthralgia. As expected, neutralizing antibodies (nAbs) were present at baseline ; mRNA-1345 significantly boosted antibody titers through month (M) 1 in YA and OA, with comparable immunogenicity observed across age groups. M1 geometric mean fold rise (GMFR) for RSV-A nAbs were 20.0-22.3 (YA) and 12.1-16.6 (OA) and for RSV-B, nAbs were 11.7-14.4 (YA) and 8.7-12.6 (OA). M1 PreF binding antibody (bAb) GMFRs were 16.1-21.7 (YA) and 8.4-12.1 . Peak antibody titers declined through M6, but levels remained \u2265 4.1-fold above BL with minimal dose response. M6 GMFR for RSV-A nAbs were 7.0-9.6 (YA) and 4.1-5.8 (OA) and for RSV-B, nAbs were 5.0-8.9 (YA) and 4.5-5.5 (OA). M6 PreF bAbs GMFR were 5.9-7.0 (YA) and 4.1-4.7 (OA). Antibody decline over time was comparable in YA and OA cohorts.mRNA-1345 is well-tolerated in YA and OA. Antibody levels were boosted substantially above BL through M6 in both cohorts. These data support the continued development of mRNA-1345 as an RSV vaccine.Grace L. Chen, MD, MPH, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Runa Mithani, PharmD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Archana Kapoor, PhD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Sophia Lu, PhD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Laila El Asmar, PhD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Catherine A. Panozzo, PhD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Christine A. Shaw, PhD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Sonia K. Stoszek, PhD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Allison August, MD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds."}
+{"text": "Correction: Environ Health 20, 44 (2021)https://doi.org/10.1186/s12940-021-00726-xFollowing the publication of the original article , the autCV70  ultrasound imaging system using a linear array 7.5 MHz transducer .In brief, high resolution B-mode ultrasound carotid artery images were acquired using a Siemens Acuson However, the correct information should state:X 300  ultrasound imaging system using a linear arrayVF 10-5 (8.0 MHz) transducer .In brief, high resolution B-mode ultrasound carotid artery images were acquired using a Siemens Acuson"}
+{"text": "Brassica species, and they play important roles in defense due to their anti-nutritive and toxic properties. Here, we conducted a genome-wide association study of six glucosinolate metabolites (mGWAS) in rapeseed, including three aliphatic glucosinolates , one aromatic glucosinolate (m157 gluconasturtiin) and two indole glucosinolates (m165 indolylmethyl glucosinolate and m172 4-hydroxyglucobrassicin), respectively. We identified 113 candidate intervals significantly associated with these six glucosinolate metabolites. In the genomic regions linked to the mGWAS peaks, 187 candidate genes involved in glucosinolate biosynthesis  and novel genes  were predicted based on the mGWAS, combined with analysis of differentially expressed genes. Our results provide insight into the genetic basis of glucosinolate biosynthesis in rapeseed and should facilitate marker-based breeding for improved seed quality in Brassica species.Glucosinolates (GSLs) are secondary plant metabolites that are enriched in rapeseed and related  Brassica species, providing these plants with their pungent odor [Glucosinolates (GSLs) are secondary metabolites comprising sulfur and nitrogen that are specially produced in ent odor ,2,3. GSLent odor ,5. Howevent odor ,7,8,9,10\u03b2-D-thiosaccharide and (Z)-n-hydroxamic sulfate, but have variable R-side chain groups due to the precursor amino acids [\u03b3-glutamyl peptide (GGP1) and C-S lyase (Super root 1 (SUR1)), and then to the GSL core structure by glucosyltransferases (UGT74s) and sulfotransferases (SOTs) [O-methyltransferases (IGMTs) for indole GSLs [Generally, GSLs share the same basic structure, including no acids . The lenno acids ,13,14. Tno acids ,16, whicno acids ,17,18,19s (SOTs) ,22,23,24s (SOTs) . The corole GSLs ,27,28.Arabidopsis, the glucosinolate biosynthesis pathway is one of the best characterized specialized metabolite pathways [AtTSB1, AtCYP79B2, AtCYP79B3, AtCYP83B1 and AtST5a and so on), leading to the accumulation of GSLs [MYB51 and the GSL biosynthesis gene CYP83B1 to regulate the de novo biosynthesis of indole GSLs, but also regulates the expression of genes involved in side chain modification , thereby increasing the biosynthesis of indole GSLs [In pathways . Progrespathways ,32,33,34 of GSLs . The ali of GSLs ,37, whil of GSLs ,39. Seve of GSLs . Further of GSLs ,41. A reole GSLs .Brassica napus L.) is used worldwide as an oil crop and source of edible vegetable oil and feed meal. Therefore, breeding rapeseed varieties with low GSL levels in seeds is an important breeding goal. To date, many rapeseed resources with low seed GSL contents have been developed in polyploid rapeseed [B. napus seeds has been challenging. In this study, we performed a genome-wide association study for six glucosinolate metabolites in 143 rapeseed accessions using 239,945 SNP markers obtained by resequencing [B. napus with improved quality.Rapeseed , whereas the level of m172 showed a higher correlation with those of three aliphatic GSLs  than with that of m165 (Based on ultrahigh-performance liquid chromatography-heated electrospray ionization-tandem mass spectrometry (UPLC-HESI-MS/MS) analysis, we obtained six glucosinolate metabolites with high content in rapeseed at 35 days after flowering (DAF), including three aliphatic GSLs , one aromatic GSL (m157 gluconasturtiin) and two indole GSLs (m165 indolylmethyl-glucosinolate and m172 4-hydroxyglucobrassicin) a,b. Corr of m165 , indicat of m165 . The coe of m165 . Further of m165 , indicat of m165 . Therefo of m165 a, but al of m165 b.p-value < 4.17 \u00d7 10\u22126. The results of mGWAS for the six glucosinolate metabolites in two years (2017cq and 2018cq) and BLUP (best linear unbiased prediction) values  values  are summB. napus genome  in two years and BLUP values. In addition, qGSL-A01-4, qGSL-A02-2, qGSL-A03-3, qGSL-A05-2, qGSL-A06-1, qGSL-A06-3, qGSL-A09-6, qGSL-A09-7, qGSL-C04-2 and qGSL-C08-5 were repeatedly detected in at least one year or BLUP value. In these repeatedly detected interval regions, most of them were simultaneously associated with three glucosinolate metabolites  or any two of these  [Based on the physical positions of the significant SNPs, we mapped the intervals to the corresponding chromosomes of the reference genome h (v4.1)  and searBased on the known GSL biosynthesis pathway, we identified 64 candidate genes in the GSL biosynthesis pathway (including genes encoding enzymes in this pathway and their homologs) among the intervals, which showed differential expression during seed development in BnHG vs. BnLG . \u03b3-glutamyl peptidase 1 (GGP1), S-alkyl thiohydrogen oxidase lyase (SUR1), sulfotransferases , flavin-monooxygenase glucosinolate S-oxygenase 5 (FMOGS-OX5), oxoglutarate-dependent dioxygenase (AOP3) and indole-glucoside O-methyltransferases , were identified on the following chromosomes: A01 , A02 , A03 , A04 , A05 , A06 , A07 , A08 , A09 , C02 , C04 , C06  and C07 , respectively , methylthioalkyl malate synthase (MAM1), bile acid transporter (BAT5), cytochrome P450s , UDP-glucosyl transferases , glutathione S-transferases , ectively , indicatMYB34, MYB51, MYB122 and WRKY33, which are involved in GSL biosynthesis [B. napus.Previously studies have identified MYB transcription factors that regulate GSL biosynthesis in various plants ,38,50,69ynthesis ,70, were\u03b2-glucosidases (BGLUs), one ABC transporter family protein (PEN3) and six glucosidolate glucohydrolases (TGGs), respectively , three nitrile specifier proteins (NSPs), one glutathione gamma-glutamylcysteine transferase (PCS1), fifteen ectively . Correspectively . Among tectively .IMD1, MAM1, IGMTs, MYB34, MYB51, MYB122 and so on) and novel genes  were identified within the confidence intervals of GSLs. Our findings not only demonstrate the reliability of the association genetics approach, but also provide new insight into elucidating the biosynthesis of these GSLs in B. napus seeds.Altogether, numerous key homologous genes, including known genes  and BnaIMD1 (BnaA02g02020D), which are located in candidate intervals qGSL-A02-1 and qGSL-C02-11, associated with three aliphogenic GSLs and one aromatic GSL  and BanGS-OH (BnaA04g17900D), which participate in the side chain modification process of aliphatic GSL biosynthesis and showed higher expression profiles in BnHG vs. BnLG (CYP79 genes [CYP83 genes [GGP1 [SUR1 [UGT74 genes [SOT genes [BnaCYP79F1 (BnaA06g11010D), BnaCYP79B2 (BnaA08g16100D), BnaCYP83A1 (BnaA04g24160D and BnaC04g47910D), BnaGGP1 , BnaSUR1 (BnaA09g10030D), BnaUGT74B1 (BnaA09g29790D), BnaUGT74C1 (BnaC04g42530D), BnaSOT16 (BnaA07g31260D), BnaSOT17 (BnaA06g12720D) and BnaSOT18  , NSP genes (3), CAD1 (1), TGG genes (6) and NIT genes (3) (BnaSOT12 (BnaA02g27300D) is homologous to AtSOT12, which functions in flavonoid, brassinosteroid and salicylic acid activity, and is involved in plant responses to salt, osmotic stress and phytohormones [BnCYP81D11 (BnaA02g29380D and BnaA02g29390D), BnaCYP81D7 (BnaA03g23030D), BnaCYP81G1 , BnaCYP81F3 (BnaA08g15650D), BnaCYP81F4 (BnaA08g15660D) and BnaCYP81K1 (BnaC02g36740D), are located in the interval regions associated with glucosinolate metabolites and showed differential expression in BnHG vs. BnLG seeds  . In addienes (3) , which aBnaA08g16D, BnaCYP BnaA06g10D, BnaCYLG seeds . Among tynthesis ,103, butLG seeds . SimilarCYP81F2, IGMT1 and IGMT2 [A. thaliana [B. napus  biosynthesis, specifically the production of 4-methoxyindole-3-ylmethyl glucosinolate (4MI3G), by directly activating the expression of nd IGMT2 , while tnd IGMT2 ,70,105. thaliana . In thisB. napus . Our finB. napus seeds , 46 medium-glucosinolate content (45~100 \u03bcmol/g) and 27 high-glucosinolate content (>100 \u03bcmol/g), respectively  and low  seed glucosinolate content were selected from 143 B. napus accessions for transcriptome sequencing (RNA-Seq), respectively. The seeds of 20, 30 and 40 DAF were also collected and pooled from five or more individuals in BnHG and BnLG plants. All samples were stored at \u221280 \u00b0C until further analysis.In total, 143 ectively . All accThe total RNA was extracted from the seeds of 20, 30 and 40 DAF using an EZ-10 DNAaway RNA Mini-Preps kit  following the manufacturer instructions. Then, the qualified RNA samples were used for libraries construction and sequenced on Illumina Hiseq 2000 platform with 150 bp paired-end reads . The gene expression profiles were evaluated using FPKM (fragments per kilo base of exon model per million) values. Genes were considered as differentially expressed genes (DEGs) with a minimum 2-fold difference in expression (|log2FC| \u2265\u20091). g at 4 \u00b0C for 10 min. Then, the residues were repeatedly extracted. Eventually, the mixed liquid supernatants were used for UPLC-HESI-MS/MS analysis after being filtered by a 0.22 \u03bcm nylon filter. All experiments were performed in at least three replicates for each accession.The raw metabolites were extracted from fresh seeds described in our previous research , with miThe UPLC-HESI-MS/MS was performed using Dionex UltiMateTM 3000 UHPLC system  coupled to a Thermo Scientifific Q-Exactive System equipped with an S-Lens ionizer source , including the precolumn  and Acquity UPLCBEH C18 column . The parameters were as follows: the mobile phases A (0.1% formic acid) and B (0.1% acetonitrile aqueous solution), 37 \u00b0C column temperature, 0.3 mL/min flow rate and 10 \u03bcL injection volume, respectively. The mobile phase gradients are 0\u20132 min, 5% B\u201310% B; 2\u201310 min, 10\u201325% B; 10\u201313 min, 25\u201395% B; 13\u201316 min, 95% B; 16\u201316.5 min, 95\u20135% B; 16.5\u201321 min, 5% B. The mass spectrometry was detected in negative ion mode with a scanning range of 100 to 1200 (m/z), 3.5 kV ion source voltage, 350 \u00b0C capillary temperature, 35 sheath gas, 10 auxiliary gas and 0 backblow air, respectively.http://prime.psc.riken.jp/compms/msdial/main.htmL#MSP, accessed on 2 April 2022), and automatically converted by ABF  converter  [\u22121 . Raw data of UPLC-HESI-MS/MS were firstly treated with MS-DAIL ver4.1 MSP negative database (ch 2020) . Correspch 2020) ,110. Furch 2020) , and thehttp://www.eXtension.org/pages/61006, accessed on 14 October 2020), respectively. The content of six glucosinolate metabolites in the 2017 and 2018 growing seasons and resulting values of BLUP were used as phenotypes for GWAS, respectively. The heritability was calculated by using the multi-year repeated model. In addition, the quantitative data of glucosinolate metabolites were divided into 10 grades, and Shannon\u2013Wiener Diversity Index was used to calculate metabolites [t-test) were performed [The glucosinolate metabolites were detected in two consecutive years (2017 and 2018) with replications; we further obtained the best linear unbiased prediction (BLUP) of six glucosinolate metabolites per accession using a linear model using an R script  and GATK (version 3.2) to process local realignment and base quality detection for the alignment results, sequentially. Further, we then used AMtools mpileup (version 0.1.19\u201344428cd) and GATK to perform SNP calling. In total, 239,945 high-quality SNPs with a minor allele frequency (MAF) <5% were used for further analysis. The six models, including na\u00efve, Q, K, PCA, K\u2009+\u2009Q and K\u2009+\u2009PCA model, were applied to determine the statistical associations between phenotypes and genotypes. Quantile\u2013quantile (QQ) plots were used for false-positive correction for association analyses. In this study, genome-wide association analysis for six glucosinolate metabolites was carried out using the GLM with Q model and MLM with\u2009Q +\u2009K model by TASSEL 5.2.1 software. The population structure Q matrix was completed by admixture_linux-1.3.0 software [P\u2009 < \u2009P \u2009= \u20091/N , and the threshold of significance was set to p < 4.17 \u00d7 10\u22126.Detailed methods used for SNP genotyping and mapping were previously described ,47,48. I-bzh\u2019  using thsoftware . The sigbzh reference genome  [The significant interval regions were mapped to the Darmor-er 2022) , which wer 2022) . Finally"}
+{"text": "Correction: J Exp Clin Cancer Res 38, 435 (2019)https://doi.org/10.1186/s13046-019-1439-xFigure Figure Figure Figure Following publication of the original article , errors The corrections do not have any effect on the final conclusions of the paper.Additional file 1: Figure S1. JMJD2C promoted the metastasis of CRC LoVo cells. a-c Real time PCR and western blotting were performed to confirm the gene silencing and overexpressing efficiency for JMJD2C. LoVo was transiently transfected with shRNA/NT vector, shRNA/JMJD2C vector, empty overexpression vector, or JMJD2C overexpression vector. d Migration assays of LoVo cells transfected with shRNA/NT, shRNA/ JMJD2C, empty vector, or JMJD2C overexpression vector, respectively. e Numbers of migrated cells are shown as mean \u00b1 SD; n = 3. *, P < 0.05; **, P < 0.01 (t test)."}
+{"text": "Platythyreaclypeata species group is reviewed and three species, including one new species, P.homasawinisp. nov., are recognized. This species group is distinguished from the P.parallela species group by the reddish-brown body, the elliptical shape of the propodeal spiracle, the elongate antennal scape, and the distinctly narrowed posteriad space between frontal carinae. Platythyreahomasawinisp. nov., from Thailand and China, is described based on the worker caste. The type series of the new species was collected on the forest floor from dead wood in an advanced stage of decomposition. A key to the Oriental species of the genus Platythyrea based on the worker caste is provided.The  Platythyrea Roger, 1863  is a rarely collected group with recent reviews listing 39 species ; P.quadridenta Donisthorpe, 1941; P.tricuspidata Emery, 1900 and P.janyai Phengsi, Jaitrong, Ruangsittichai & Khachonpisitsak, 2018). Among them, five species are reported from Thailand, and they belong to two species groups (sensu Brown 1975): the P.clypeata group (P.clypeata and P.janyai) and P.parallela group   . Our recTHNHM). The specimens were compared with high-resolution images of holotypes and paratypes of closely related species  available on P.janyai were also examined. The holotype and paratypes of P.homasawini sp. nov. are deposited in THNHM.This study is mainly based on the material deposited in the Natural History Museum of the National Science Museum, Thailand  process vertically to a line intersecting the dorsal most point of the node.PL Petiole length: length of petiole measured in lateral view from the anterior articulation to the posterior articulation of petiole.PW Pronotal width: maximum width of pronotum measured in dorsal view.SL Scape length: straight-line length of the antennal scape, excluding the basal constriction or neck.SI Scape index = SL/HW \u00d7100.TL Total length: total outstretched length of the individual, from the mandibular apex to the gastral apex.Abbreviations of the type depositories are as follows:BMNHThe Natural History Museum, London, U.K.MHNGMus\u00e9um d\u2019Histoire Naturelle, Geneva, Switzerland.THNHM Natural History Museum of the National Science Museum, Thailand.Platythyrea species were made at the Microscopic Center, Faculty of Science, Burapha University with a LEO 1450 VP scanning electron microscope on gold coated specimens.Scanning electron microscope images of The species group can be characterized by the following characteristics: 1) body reddish brown; 2) space between frontal carinae distinctly narrowed posteriad , deposited in MHNG . Holotype of P.thwaitesi: alate queen from Sri Lanka, deposited in BMNH .THNHM-I-24983) and 1 dealate queen (THNHM-I-24982), Laos, Vientiane, Pak Ngum District, Ban Phang Dang, ca 300 m alt., 14.VI.2010, W. Jaitrong leg., Colony no. WJT-LAO-143 (THNHM-I-24981) \u2022 1 worker (THNHM-I-24984), Laos, Vientiane, Pak Ngum District, Ban Phang Dang, 14.VI.2010, Sk. Yamane leg., Colony no. LA10-SKY-128 \u2022 1 worker (THNHM-I-24981) Laos, Vientiane, Pak Ngum District, Ban Phang Dang, ca 300 m alt., 12.VI.2010, W. Jaitrong leg. \u2013 Thailand \u2022 9 workers (THNHM-I-02423 to THNHM-I-02431), eastern Thailand, Chachoengsao Province, Tha Takiab District, Khao Ang Reu Nai Wildlife Sanctuary, 27.IX.2002, W. Jaitrong leg., Colony no. WJT270902-01 \u2022 3 workers (THNHM-I-02432 to THNHM-I-02434), same locality, date and collector, Colony no. WJT270902-1 \u2022 6 workers (THNHM-I-02435 to THNHM-I-02440), eastern Thailand, Sa Kaeo Province, Khao Ang Reu Nei Wildlife Sanctuary, 26.VI.2003, W. Jaitrong leg., Colony no. WJT03-TH-228; 22 workers (THNHM-I-02441 to THNHM-I-02452) and 1 male (THNHM-I-02453), eastern Thailand, Chanthaburi Province, Soi Dao District, 14.V.2008, W. Jaitrong leg., Colony no. WJT08-E065 \u2022 14 workers  and 1 dealate queen, central Thailand, Nakhon Nayok Prov., Muang Dist., Nang Rong Temple, in rotting wood, 29.VIII.2018, W. Jaitrong leg., Colony no. WJT290819-09.Laos \u2022 10 workers (n = 10). and inhabits primary dry evergreen forest and disturbed forests. All colonies of this species were collected from dead wood on the forest floor in an advanced stage of decomposition.Sri Lanka, Vietnam, Laos (Vientiane), and Thailand  Fig. .Platythyreaclypeata is similar to P.gracillima, P.janyai, and P.prizo. However, P.clypeata can be easily separated from P.gracillima by the following characteristics (characters of P.gracillima in parentheses unless otherwise stated): 1) body size smaller ; 2) eye smaller and flat ; 3) clypeus narrow and rather convex (clypeus broad and rather flat); 4) declivity of propodeum concave ; 5) seen from back propodeal declivity rounded above ; 6) petiole laterally swollen ; 7) head finely punctate (head rather smooth).Platythyreaclypeata can be distinguished from P.prizo by the following characteristics (characters of P.prizo in parentheses unless otherwise stated): 1) head shorter ; 2) eye flat and small ; 3) eye without erect pubescence (eye covered with extremely fine short erect pubescence); 4) propodeum junction obtusely angulated (propodeum junction armed with a pair of short teeth or tubercles); 5) in profile, posterodorsal corner of petiole forming an acute angle (roundly convex).P.janyai (characters of P.janyai in parentheses unless otherwise stated) by 1) head relatively longer ; 2) eye clearly smaller ; 3) eye flat (eye convex); 4) head finely punctate with dense shallow foveae ; 5) in profile petiole slightly longer than high and in dorsal view its node slightly narrower posteriorly ; 6) ventral outline of petiole feebly concave (weakly convex) , northern Thailand, Chiang Mai Prov., Doi Saket Dist., Ban Mae Pong, 24.XI.2021, K. Homasawin leg., Colony No. WJT241121-01. Paratypes: 29 workers , same data as holotype.THNHM-I-02463), northern Thailand, Chiang Mai Prov., Muang Dist., restored forest, 8.V.2002, S. Sonthichai leg. \u2022 1 worker (THNHM-I-02454), western Thailand, Kanchanaburi Prov., Thong Pha Phum N.P., Natural Forest, 8.III.2005, W. Sakchooeong leg.China \u2022 1 worker, Yunnan Prov., Menghai County, Meng\u2019a Town, Papo Village, 1280 m, No. A97-2318, collected from secondary monsoon evergreen broadleaf forest, 10.IX.1997, Zhenghui Xu leg. \u2013 Thailand \u2022 1 worker . TL 7.72\u20137.80, HL 1.56\u20131.64, HW 1.08\u20131.12, SL 1.64\u20131.68, EL 0.16\u20130.20, PW 0.96\u20130.99, ML 2.80\u20132.84, PL 1.08\u20131.12, PH 0.60\u20130.64, DPW 0.40\u20130.44, CI 68\u201369, EI 15\u201319, SI 150\u2013159.holotype and paratypes). Head and Thailand (Chiang Mai and Kanchanaburi provinces) Fig. .The specific name is dedicated to Mr Kaisihanat Homasawin, who donated the type series.P.janyai but it can be separated from P.janyai by the following characteristics (characters of P.janyai in parentheses unless otherwise stated): 1) head weakly widening anteriorly (head not widening anteriorly); 2) dorsal outline of petiole weakly convex ; 3) posterior margin of petiolar node without a concavity in the middle (posterior margin of petiolar node with a concavity in the middle): 4) body surface with thin pubescence (body surface with thick pubescence); 5) head longer ; 6) antennal scape relatively long, 1/3 of its length extending beyond posterolateral corner of head ; 7) eye smaller and flat ; 8) seen from back propodeal declivity rounded above ; 9) dorsal outline of petiole weakly convex .The new species is similar to Platythyreahomasawini can be easily separated from P.clypeata by the following characteristics (characters of P.clypeata in parentheses unless otherwise stated): 1) head in full-face view, posterior margin weakly concave ; 2) antennal scape relatively long, 1/3 of its length extending beyond posterolateral corner of head ; 3) clypeus broad (clypeus narrow); 4) eye larger ; 5) mesosoma relative longer ; 6) mesopleuron not demarcated from mesonotum ; 7) in profile view, propodeal junction roundly convex ; 8) in dorsal view, posterior margin of petiole clearly convex ; 9) lateral face of head entirely micropunctate ; 10) gaster superficially shagreened (finely reticulate).Platythyreahomasawini can be distinguished from P.gracillima by the following characteristics (characters of P.gracillima in parentheses unless otherwise stated): 1) clypeus roundly convex (rather flat in P.gracillima); 2) petiole laterally convex; 3) seen from above longer than high ; 4) seen from above a little more than twice as long as broad); 5) mandible finely micropunctate (mandible finely and densely punctate); 6) head entirely finely micropunctate (rather smooth in P.gracillima).Taxon classificationAnimaliaHymenopteraFormicidae\ufeffPhengsi, Jaitrong, Ruangsittichai & Khachonpisitsak, 201880F1B0B2-8F43-5645-B08A-D5F34E0D516DPlatythyreajanyaiHolotype (THNHM-I-02392) and three paratypes (THNHM-I-02393 to THNHM-I-02395) workers, southern Thailand, Phatthalung Province, Si Banphot District, Riang Thong Waterfall, Khao Pu Khao Ya National Park, 28.IX.2007, W. Jaitrong leg., Colony no. WJT07-TH-2060 (THNHM-I-02392), deposited in THNHM (examined).THNHM-I-02465) \u2013 Thailand \u2022 2 workers, southern Thailand, Trang Province, Na Yong District, Khao Chong Botanical Garden, 7.XI.2014, W. Jaitrong leg., Colony No WJT071114-2 (THNHM-I-02421 to THNHM-I-02422) \u2022 5 workers, same locality and collector, 26.XII.2018, Colony no. WJT261218-1 (THNHM).Malaysia \u2022 1 worker, western Malaysia, Selangor, Ulu Gombak, 22.III.2013, F. Ito leg. ((n = 4).TL 6.63\u20136.96, HL 1.42\u20131.45, HW 1.06, SL 1.39\u20131.42, EL 0.20, ML 2.21\u20132.31, PL 0.73\u20130.79, PH 0.53, DPW 0.40, CI 72\u201374, EI 18, SI 131\u2013134.holotype and paratypes). Head and Malaysia  Fig. .Platythyreajanyai is similar to P.clypeata, P.homasawini and P.gracillima. However, P.janyai can be easily separated from P.gracillima by the following characteristics (characters of P.gracillima in parentheses unless otherwise stated): 1) body size smaller ; 2) eye relatively smaller ; 3) seen from back, propodeal declivity tapering above ; 4) petiole laterally convex, seen from above longer than broad . For differentiation of P.janyai and P.clypeata, see \u201cComparative notes\u201d under P.clypeata, and of P.janyai and P.homasawini, see under P.homasawini.Platythyrea are now known around the world. Among them, six species are found in Thailand, and they belong to two species groups : P.clypeata group  and P.parallela group .With this study, 40 valid species of the genus The shape of the frontal lobe, mandibular shape and dentition, and shape of mesosoma and petiolar node were characters used by Brown (1975) to distinguish the two species groups mentioned here. These morphological characters were confirmed and used by several authors who described new species after Brown (1975) . The preP.clypeata group can be distinguished from other congeners mainly by the following: body reddish brown; space between frontal carinae distinctly narrowed posteriad; propodeal spiracle opening elliptical, petiole longer than high, and posterior margin concave. The worker caste of Platythyrea species is generally monomorphic with little variation in size within species. Thus, worker body size can be used for separating large and small species. In this study, worker body size has been used to separate P.homasawini sp. nov., P.janyai, and P.clypeata.The Platythyreaclypeata is distinctly allopatric with P.janyai in geographic subregion in continental Asia . However, P.clypeata and P.janyai both were found in lowland (< 300 m a.s.l.). Platythyreaclypeata inhabits various habitats including plantations, secondary forests, and primary dry evergreen forests. Platythyreajanyai is confined to primary evergreen forests. Platythyreahomasawini sp. nov. is restricted to highland hill evergreen forests.tal Asia . It has"}
+{"text": "Cystobasidium ongulense has been reported from East Ongul Island near Syowa Station, East Antarctica, as a new basidiomycetous yeast species. This species has cold active lipases and cellulases that are active even at subzero temperatures. We report draft genome sequences of five Cystobasidium ongulense strains isolated from East Antarctica. Cystobasidium ongulense was isolated from East Ongul Island, East Antarctica, and reported as a new species of basidiomycete yeast  isolated from East Ongul Island and three strains  isolated from Inhovde, near Syowa Station.g for 5\u2009min at 4\u00b0C. The cell pellets were washed with sterile distilled water and precipitated by centrifugation again. The cell pellets were dried using a vacuum freeze dryer. The lyophilized cells were powdered in a mortar and used for genome extraction. The genomic DNA of five Cystobasidium ongulense strains was extracted and purified using a NucleoBond AXG100 column with a Nucleo buffer set III (TaKaRa Bio) according to the manufacturer\u2019s protocol. The concentration and purification of the genomic DNA were determined using a NanoDrop spectrophotometer (Thermo Scientific) and the Qubit double-stranded DNA (dsDNA) broad-range (BR) assay kit (Thermo Scientific). Then, the genomic DNA was digested using the microTUBE (Covaris), and the genomic library was constructed using the Illumina TruSeq DNA PCR-free library preparation kit (Illumina). The quality of the library was confirmed using the Quant-iT PicoGreen dsDNA assay kit, and the library size was checked using the Agilent 2200 TapeStation system . The paired-end sequence reaction was performed on a HiSeq 2500 instrument (Illumina). Adapters and low-quality bases were trimmed using fastp ver. 0.12.4 (\u2013ab initio gene predictions were carried out using AUGUSTUS ver. 3.4.0 (Each strain was cultured in yeast extract-peptone-dextrose (YPD) liquid medium (Difco) at 15\u00b0C for 5\u2009days. The cells from cultures were collected by centrifugation at 3,500\u2009\u00d7\u2009. 0.12.4 , annotat. 0.12.4 \u201311 automr. 3.4.0 . The resr. 3.4.0  and AGATr. 3.4.0 , Phobiusr. 3.4.0 , antiSMAr. 3.4.0 , and Eggr. 3.4.0  against r. 3.4.0 .C. ongulense strains is shown in A summary of the genomic analysis results of the five C. ongulense will help us understand how Antarctic fungi adapt to the extreme environment.We believe that the genomic information of PRJDB6568 with accession numbers BQUO00000000.1 (9A-2), BQUP00000000.1 (9A-5), BQUQ00000000.1 (051-20-1), BQUR00000000.1 (056-1-20Y-3), and BQUS00000000.1 (056-20-8). The raw reads are available under SRA accession numbers DRR118811, DRR118812 (9A-2), DRR118813, DRR118814 (9A-5), DRR118815, DRR118816 (051-20-1), DRR118817, DRR118818 (056-1-20Y-3), and DRR118819, DRR118820 (056-20-8).This whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under BioProject accession number"}
+{"text": "One of the main mechanisms of epigenetic regulation in higher eukaryotes is based on the methylation of cytosine at the C5 position with the formation of 5-methylcytosine (mC), which is further recognized by regulatory proteins. In mammals, methylation mainly occurs in CG dinucleotides, while in plants it targets CG, CHG, and CHH sequences (H is any base but G). Correct maintenance of the DNA methylation status is based on the balance of methylation, passive demethylation, and active demethylation. While in mammals active demethylation is based on targeted regulated damage to mC in DNA followed by the action of repair enzymes, demethylation in plants is performed by specialized DNA glycosylases that hydrolyze the N-glycosidic bond of mC nucleotides. The genome of the model plant Arabidopsis thaliana encodes four paralogous proteins, two of which, DEMETER (DME) and REPRESSOR OF SILENCING 1 (ROS1), possess 5-methylcytosine-DNA glycosylase activity and are necessary for the regulationof development, response to infections and abiotic stress and silencing of transgenes and mobile elements. Homologuesof DME and ROS1 are present in all plant groups; however, outside A. thaliana, they are poorly studied.Here we report the properties of a recombinant fragment of the ROS1 protein from Nicotiana tabacum (NtROS1),which contains all main structural domains required for catalytic activity. Using homologous modeling, we haveconstructed a structural model of NtROS1, which revealed folding characteristic of DNA glycosylases of the helix\u2013hairpin\u2013helix structural superfamily. The recombinant NtROS1 protein was able to remove mC bases from DNA,and the enzyme activity was barely affected by the methylation status of CG dinucleotides in the opposite strand.The enzyme removed 5-hydroxymethylcytosine (hmC) from DNA with a lower eff iciency, showing minimal activityin the presence of mC in the opposite strand. Expression of the NtROS1 gene in cultured human cells resulted ina global decrease in the level of genomic DNA methylation. In general, it can be said that the NtROS1 protein andother homologues of DME and ROS1 represent a promising scaffold for engineering enzymes to analyze the statusof epigenetic methylation and to control gene activity. DNA methylation is a dedicated mechanism of gene regulation,especially developed in higher eukaryotes. The 5-methylcytosine(mC) nucleobase, formed by cytosine methylationat the C5 position, serves as a reversible epigenetic mark thatplays an important role in the control of gene expression andthe protection of the genome from mobile elements. DNAmethylation occurs in a wide range of multicellular eukaryotes;however, its significance and functions in these organismsdiffer greatly .For example, in mammals, methylation most often occurs inCpG dinucleotides, while in plants, a significant proportionof mC occurs in trinucleotides CHG and CHH (where H is\u201cnot G\u201d). The consequences of mC for gene activity are mainlymediated by proteins containing methyl-binding domains thatform complexes with histone deacetylases or themselves havethe activity of histone-specific methyltransferases, chromatinremodeling factors, etc., which leads to chromatin condensationand transcription suppression . Recent studies have shown that an oxidizedderivative of 5-methylcytosine, 5-hydroxymethylcytosine(hmC), also plays an epigenetic role in the mammaliangenome . Unlike mC, hmC is enrichedin promoters and bodies of actively expressed genes and isconsidered an activating epigenetic marker Correct methylation of various sites in the genome is extremelyimportant, since the transcriptional activity of genesdepends on it. Errors in DNA methylation can have graveconsequences. In particular, in humans, global DNA demethylationor hypermethylation of tumor suppressor genes serve ascancer markers. Maintaining the status of DNA methylation incells requires a balance of methylation and active and passivedemethylation. The mechanisms of active demethylation ofthe genomic DNA in higher eukaryotes have only been discoveredin the last decade. In mammals, active demethylationis initiated by regulated damage to mC, which can occur intwo ways: either deamination of mC to T by AID/APOBECenzymes, or oxidation of mC to hmC and further derivatives(5-formylcytosine and 5-carboxycytosine) by TET familydioxygenases . The modified bases are further perceived bycellular repair systems as damaged and are removed by theDNA base excision repair pathway.Unlike mammals, plants have unique enzymes that directlyhydrolyze N-glycosidic bonds of mC nucleotides. These DNAglycosylases \u2013 DEMETER (DME) and REPRESSOR OFSILENCING 1   \u2013 are involved in theregulation of the methylation status of the sites in the plantgenome that determine gene imprinting during paternal ormaternal inheritance and silence or activate specific promotersduring plant development and stress response .After removal of mC, the resulting apurinic-apyrimidinic site(AP site) is cleaved either by the enzyme\u2019s own AP lyase activityor by the AP endonucleases APE1L or ARP, then a normalnucleotide is incorporated by one of the DNA polymerases,and the nick is ligated by the LIG1 DNA ligase. Interestingly,DME and ROS1 can also excise hmC, which is not regardedas an epigenetic base in plants . The localizationof demethylation by DME/ROS1 is regulated bysmall RNAs that bind to the enzyme itself or to the proteincomplex that contains it .In addition to DME and ROS1, the genome of the modelplant Arabidopsis thaliana contains three more genes that arehomologous to DME and ROS1: DEMETER-LIKE 2 (DML2),DEMETER-LIKE 3 (DML3), and AT3G47830, which has notbeen characterized thus far except for participation of DML2and DML3 in maintaining correct DNA methylation . All these proteins belongto the DNA helix\u2013hairpin\u2013helix (HhH) structural superfamilyof DNA glycosylases. Other members of this superfamily areinvolved in the removal of oxidized, alkylated and deaminatednucleobases from the genome . DME/ROS1 enzymes attract attention as potentialtools for targeted regulation of gene activity: for example, thepossibility of targeted DNA demethylation in human cells byA. thaliana ROS1 (AtROS1) fused to the RNA-guided Cas9protein has been shown , and A. thaliana DME (AtDME) was used to analyze the level ofmC in genomic DNA .In plants other than A. thaliana, there were few studies onthe role of DME-like proteins in active epigenetic demethylation;some data exist for rice, wheat, barley, and tomato . In 2007, a ROS1 homolog from Nicotiana tabacum(NtROS1) was cloned, and the recombinant proteinproducedin insect cell culture was shown to cleave methylatedtobaccogenomic DNA . None of these studiesincluded a detailed biochemical characterization of the protein.We have previously shown that the NtROS1 fragment correspondingto the minimal catalytically active AtROS1 fragmenthas the activity of 5-methylcytosine-DNA glycosylase.Here, in view of the potential value of plant demethylationenzymes as tools for genetic technologies, we characterizethe substrate specificity of the recombinant NtROS1 catalyticfragment on mC and hmC in different contexts of methylatedCpG dinucleotides and show that the expression of NtROS1in human cells causes a global decrease in DNA methylation.ProtoScript II reverse transcriptase, Q5 Hot Start High-FidelityDNA polymerase, Escherichia coli uracil DNA glycosylase,ClaI and SacI restriction endonucleases were purchasedfrom New England Biolabs (USA), and bacteriophage T4polynucleotide kinase, from Biosan .Oligonucleotides listed in the Table were synthesized at theSB RAS ICBFM Laboratory of Biomedical Chemistry usingcommercially available prosphoramidites . If necessary, the oligonucleotides were 32P-labeled atthe 5\u2032-end using \u03b3[32P]ATP (SB RAS ICBFM Laboratory ofBiotechnology) and T4 polynucleotide kinase.To build a model of the NtROS1 catalytic domain in theSwiss-Model program , the AtDME(AF-Q8LK56-F1-model_v1) and AtROS1 (AF-Q9SJQ6-F1-model_v1) templates from the AlphaFold database  were used.To obtain a catalytically inactive NtROS1 with theAsp1359Asnsubstitution, a pLATE31 plasmid with an insertencoding the catalytically active NtROS1 fragment (aminoacid residues 754\u20131796)  was mutagenizedusing primers D1359Nfwd and D1359Nrev (seethe Table) and the Q5 Site-Directed Mutagenesis Kit (NewEngland Biolabs). The mutation was confirmed by Sangersequencing. Wild-type NtROS1 and NtROS1 D1359N wereoverproduced and purified as described previously .To study the activity of NtROS1, double-stranded substrateswere obtained by annealing oligonucleotides C1, C2,M1, M2, H1, and H2 (see the Table). The reaction mixturecontained 50 nM substrate, 50 mM Tris\u2013HCl (pH 8.0), 1 mMEDTA, 1 mM DTT, 0.1 % bovine serum albumin, and 100 nMNtROS1. The mixture was incubated at 37 \u00b0C; aliquots werewithdrawn at various times (2\u2013300 min) and mixed with anequal volume of the stop solution . Ifnecessary, the aliquots were preheated for 2 min at 95 \u00b0C inthe presence of 0.1 M NaOH and neutralized with an equimolaramount of HCl. The reaction products were resolvedby electrophoresis in a 20 % polyacrylamide gel containing7.2 M urea, visualized by prosphorimaging using the TyphoonFLA 9500 system , and quantified usingthe Quantity One v4.6.3 software . The apparent rate constants were determined using theSigmaPlot v11.0 software  by nonlinearregression to the equation [P] = [P]max(1 \u2013 e\u2212kt ), where[P] is product concentration, [P]max is the maximum productconcentration, k is the reaction rate constant, and t is time.To assess the status of global DNA methylation uponNtROS1 expression in human cells, wild-type and D1359NNtROS1 coding sequences were cloned into the pIRES-eGFPpuroplasmid  at the SacI and ClaI restriction sites. HEK293 Phoenix cells (1.2\u00b7106) were transfected with5 \u03bcg of the plasmid by the calcium phosphate method andgrown in a monolayer in DMEM with 10 % fetal calf serum. After 24 and 48 h, the medium was changedwith the addition of 3 \u03bcg/ml puromycin. Transfection efficiencywas determined by flow cytometry  by detection of the fluorescence ofeGFP encoded by the same plasmid. NtROS1 expression inthe transfected cells was confirmed by reverse transcriptionPCR using \u03b2-actin mRNA as a control. Genomic DNA wasisolated from the cells (5\u00b7106) using a QIAamp DNA Mini Kit, and the relative content of mC wasdetermined using anti-mC antibodies . The resultswere compared using Student\u2019s t-test.Plant mC-specific DNA glycosylases are proteins of considerablesize: for example, AtDME and AtROS1, as well astheir DML2 and DML3 paralogs, are over 1000 amino acidresidues long . The extended N-terminal regions ofthese polypeptides are unstructured, although some of theirparts are necessary for enzyme activity. The C-terminal regionscontain a conserved HhH catalytic domain and an iron-sulfurcluster (FeS cluster), characteristic of many DNA glycosylasesthat recognize oxidative DNA damage, as well as an RNAbindingmotif  and a CXXCtype permuted zinc finger unique to the DME/ROS1 family. Unlike in all other HhH superfamily DNAglycosylases, the catalytic domain in DME/ROS1 proteinsis disrupted by a long non-conserved insert . Based on the literature data on the AtROS1protein, we previously cloned the NtROS1 cDNA fragmentencoding amino acid residues 754\u20131796 .This region contains all elements necessary for the catalyticactivity in AtROS1 .Since the structures of the DME/ROS1 family proteins arecurrently unknown, for a more detailed understanding of theorganization of the NtROS1 catalytic fragment, we carriedout homology modeling based on the AtDME and AtROS1models from the AlphaFold template collection . The two resulting models were almost identicalexcept for the structure of the long non-homologous regions.The disrupted HhH domain folded into the \u03b1-helical structurecharacteristic of DNA glycosylases of this superfamily,in which a DNA binding groove with the catalytic residuesLys1341 and Asp1359 is evident . In addition,several more peripheral \u03b1-helices buttress the HhH domain andthe FeS cluster and are obviously important for maintainingtheir structure. The FeS cluster, zinc finger, and RRM motifform separate structural elements that leave free access to theDNA binding groove . All disordered regions ofthe structure are also located on the side of the protein globuleopposite to the DNA binding groove.To analyze the catalytic activity and substrate specificity ofNtROS1, we performed a cleavage reaction of double-strandedoligonucleotides containing a CpG dinucleotide in which thecytosine was unmethylated, methylated, or hydroxymethylatedin one or both chains . The strand to be cleaved was32P-labeled at the 5\u2032-end. NtROS1 showed virtually no activityon the substrate containing an unmethylated CpG site, whichis consistent with the literature data claiming that the C5 positionof the cytosine must carry a substitution to be cleaved byAtROS1 . Activity towards mC andhmC was observed; however, its level differed markedly forboth types of substrates. Comparing the efficiency of cleavageof M1/C2 and M1/M2 substrates with H1/C2 and H1/H2 substrates (lanes 11 and 14), one cansee that the enzyme prefers to excise mC over hmC bothfrom CpG sites modified at only one strand and from fullymodified sites. Small differences in the cleavage of M1/C2,M1/M2 and M1/H2 substrates  indicatethat modifications in the complementary strand have littleeffect on the removal of mC, and hmC in the complementarystrand might even increase the cleavage. The enzymeshowed no activity against DNA substrates containing uracil or 8-oxoguanine. The NtROS1 D1359N mutant, as expected,exhibited no glycosylase activity against any substrate withvarious combinations of mC and hmC, which confirms theimportance of the Asp residue in the ROS1 catalytic centerfor the removal of modified bases from DNA.AtROS1 was reported to be an enzyme with a very lowturnover number , so it wasnot feasible to use steady-state kinetics to characterize theactivity of NtROS1. To determine the apparent reaction rateconstant, we used the conditions close to the single-turnoverkinetic regime ([E]0 > [S]0). Under these conditions, all DNAsubstrate rapidly binds to the enzyme, and the reaction rateis limited not by substrate binding or product release, butby the chemical step of the pseudo-first order reaction, thehydrolysisof the N-glycosidic bond of the modified nucleotide. Thus, following the accumulation of theproduct over time , one can estimate the reactionrate constant. The time courses for all substrates are shownin Fig. 3, b, and the rate constants calculated from these dataare summarized below:Substrate k\u00b7104, s\u22121*M1/C2 6.5 \u00b1 1.1*M1/M2 2.4 \u00b1 0.5*M1/H2 3.0 \u00b1 0.7*H1/C2 1.9 \u00b1 0.3*H1/M2 1.6 \u00b1 0.6*H1/H2 2.0 \u00b1 0.3*C1/C2 Not cleaved* 32P-labeled strand.In all cases, hmC was a worse substrate for NtROS1 comparedto mC. Based on these results, we conclude that NtROS1cleaves hemimethylated CpG sites most efficiently, while hmCin the context of HG/GM, on the contrary, is the worst substratefor this enzyme. The kinetic constants are consistent with thequalitative data on the relative cleavage efficiency for differentsubstrates .Many slow-turnover DNA glycosylases have lower APlyase activity in comparison with their DNA glycosylaseactivity. In this case, after the removal of the modified base,a part of the reaction product exists as an AP site for a longtime, and the true amount of the product can be revealed onlyupon treatment with alkali or nucleophilic amines . However, in the case of NtROS1, additional treatmentwith NaOH did not lead to a noticeable increase in theaccumulation of the reaction product . Apparently,the rate of the reaction catalyzed by NtROS1 is limitedby the hydrolysis of the N-glycosidic bond, which was alsosuggested for the enzyme from A. thaliana .To assess the ability of the NtROS1 protein to act asa demethylasewhen expressed in mammalian cells, we constructedplasmids based on the pIRES-eGFP-puro vectorencoding wild-type NtROS1 and its catalytically inactivemutant NtROS1 D1359N. Using anti-mC antibodies, we haveestimated the level of this epigenetic base in HEK293 cellsafter transfection with these plasmids. When cells were observedpost-transfection, the proliferation of cells with thepIRES-eGFP-puro-NtROS1 plasmid was reduced by about30 % compared to the control cells transfected with the plasmidwith no insert and the cells transfected with the plasmidencoding the catalytic mutant. The fraction of live cells wasthe same in all cases, which indicates that the cell cycle maybe slower in the presence of active NtROS1 due to the needto repair a large number of breaks introduced into DNA atmC residues. An analysis of the mC level revealed a ~2-folddecrease relative to control samples when wild-type NtROS1 was expressed and the absence of statistically significantchanges with the expression of NtROS1 D1359N . Ingeneral, it can be considered that the transient expression ofthe catalytic domain of 5-methylcytosine\u2013DNA glycosylaseNtROS1 in human cells indeed leads to the global erasure ofmC epigenetic marks. The amount of hmC in cells could notbe measured using anti-hmC antibodies, probably because thismodified base is about an order of magnitude less abundantthan mC .Thus, it can be concluded that NtROS1 is a DNA glycosylasespecific for 5-methylcytosine and, to a lesser extent,for 5-hydroxymethylcytosine. Its biological functions, as inthe case of AtROS1, most likely consist in the regulation ofmethylation status and gene expression during embryonicdevelopment  or in the response toinfections and abiotic stress ,and the regulation of silencing of transgenes and transposableelements . The mechanism of RNA-dependent addressing of DME/ROS1 family proteins to specific demethylation sites,which has not yet been elucidated, is of great interest. TheRRM motif in these polypeptides is homologous to the motifsresponsible for nonspecific interactions with RNA in manyother proteins  and, by its position in thestructure , could bind small RNAs complementaryto the DNA stretch 5\u2032 of the targeted mC. Zinc fingers ofthe CXXC type are used by many mC-recognizing proteins;however, they predominantly bind unmethylated DNA and arepresumably required for accurate positioning of the enzymein the presence of several methylation sites located at a shortdistance .The prospects for using NtROS1 and other proteins of theDME/ROS1 family as tools for genetic technologies largelydepend on the possibility of reducing the size of the catalyticfragment. A deletion of the long insert between the two partsof the HhH domain in AtROS1 fully preserves its activity, buta deletion of the C-terminal tail after the FeS cluster resultsin an inactive enzyme . Judging from thestructural models of AtROS1 and NtROS1, the HhH and RRMdomains interact with each other, and shortening the proteinhere may only be done by trimming the insert between them.In any case, DME/ROS1 proteins, including NtROS1, representa promising scaffold for enzyme engineering to analyzeepigenetic methylation status and control gene activity.The study of the demethylating DNA glycosylase ROS1 fromNicotiana tabacum reported in this work presents the onlybiochemical investigation of ROS1 beyond its homologuefrom Arabidopsis thaliana. 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DOI 10.1038/srep29565.Zemach A., Zilberman D. Evolution of eukaryotic DNA methylationand the pursuit of safer sex. Curr. Biol. 2010;20(17):R780-R785.DOI 10.1016/j.cub.2010.07.007.Zharkov D.O. Base excision DNA repair. Cell. Mol. Life Sci. 2008;65(10):1544-1565. DOI 10.1007/s00018-008-7543-2.Zhu J., Kapoor A., Sridhar V.V., Agius F., Zhu J.-K. The DNA glycosylase/lyase ROS1 functions in pruning DNA methylation patternsin Arabidopsis. Curr. Biol. 2007;17(1):54-59. DOI 10.1016/j.cub.2006.10.059."}
+{"text": "Several validated scales have been developed to measure frailty, yet it remains unknown how these measures are related. We used data from 7,070 community-dwelling older adults who participated in National Health and Aging Trend Study round 5 to construct a crosswalk among frailty measures. We operationalized the 60-item Frailty Index (FI), Study of Osteoporotic Fracture (SOF) Index, FRAIL Scale, Frailty Phenotype, Clinical Frailty Scale (CFS), Vulnerable Elder Survey-13 (VES-13), Tilburg Frailty Indicator (TFI), Groningen Frailty Indicator (GFI), and Edmonton Frailty Scale (EFS). Missing data, needed for the calculation of frailty scores, were imputed using multiple imputation by chained equations method. We then linked the scores of each frailty measure to FI using the equipercentile method, a statistical procedure that links different scales by equating percentile distributions. Participants considered frail on FI (cutpoint of 0.25) corresponded to the following scores on each frailty measure: SOF 1.3, FRAIL 1.7, Phenotype 1.7, CFS 5.3, VES-13 5.5, TFI 4.4, GFI 4.4, and EFS 5.8. Conversely, individuals considered frail on each frailty measure corresponded to the following FI scores: 0.37 (SOF), 0.40 (FRAIL), 0.42 (Phenotype), 0.21 (CFS), 0.19 (VES-13), 0.28 (TFI), 0.22 (GFI), and 0.37 (EFS). The CFS, VES-13, TFI and GFI each discriminates between non-frail and frail people in the pre- to mildly frail spectrum on the FI, whereas the SOF, FRAIL Scale, Phenotype, and EFS detect those in the higher frailty spectrum on the FI. Our results provide clinicians and researchers with a useful tool to convert and interpret frailty across scales."}
+{"text": "Retraction Note: J Exp Clin Cancer Res 37, 14 (2018)https://doi.org/10.1186/s13046-018-0681-yThe Editor in Chief has retracted this article. After publication, concerns were raised about Figs. 4d, 5e, and 6h. These were addressed in a Correction . Later,"}
+{"text": "Stem Cell Res Ther (2021) 12:456 https://doi.org/10.1186/s13287-021-02519-yThe authors have retracted this article. After publication, the authors noticed image duplication errors in Figs. 2A, 5K, 6B, S3E and S10B, some of which also overlapped with the authors' earlier article [All authors agree to this retraction."}
+{"text": "Lactobacillus pentosus (L. pentosus) plays a neuron-protective role. This study aimed to investigate the effects of L. pentosus on neurodegenerative diseases.  Neurodegenerative disease is a common neurodegenerative disorder. L. pentosus strain S-PT84. Reverse transcription-quantitative PCR (RT-qPCR) was applied to determine mRNA levels. Western blot was performed to detect protein expression. Cellular behaviors were detected using Cell Counting Kit-8 (CCK-8), flow cytometry, and TdT-mediated dUTP nick-end labeling (TUNEL) assay. The interaction between baculoviral IAP repeat containing 3 (BIRC3) and NLR family CARD domain containing 4 (NLRC4) was predicted by STING and verified by western blot.  Cells were treated with lipopolysaccharide (LPS) to establish neurodegenerative diseases model in vivo and with L. pentosus suppressed LPS-induced pyroptosis and promoted the cell viability of neurons. Additionally, L. pentosus suppressed the release of proinflammatory cytokines (interleukin 1 beta (IL-1\u03b2) and IL-18) and the protein expression of pyroptosis biomarkers (cleaved caspase1 (CL-CASP1) and N-terminal fragment gasdermin D (GSDMD-N)). Moreover, L. pentosus upregulated BIRC3, which induced the inactivation of NLRC4. However, BIRC3 knockdown alleviated the effects of L. pentosus and induced neuronal degeneration. L. pentosus may play a neuron-protective role via regulating BIRC3/NLRC4 signaling pathways. Therefore, L. pentosus may be a promising strategy for neurodegenerative diseases. Dementia is a common neurodegenerative disease, characterized by progressive cognitive degeneration, which induces huge burden on public health . Elder n\u03b2 and IL-18, and cell death [Pyroptosis is a form of programmed cell death executed by Gasdermin D (GSDMD)  The actill death . Previoull death \u201312. HoweLactobacillus pentosus (L. pentosus), a member of probiotics from intestinal flora, plays a neuron-protective role and mitigates aging- and scopolamine-induced memory impairment [Probiotics are considered to be a safe alternative therapy for many diseases, including cancer and neural disorders , 14. Accpairment , 17. How\u03baB) target gene activation to attenuate inflammation, which promotes the development of splenic marginal zone lymphoma [Baculoviral IAP repeat containing 3 (BIRC3) is a member of inhibitor of apoptosis proteins (IAPs) . BIRC3 slymphoma . Moreovelymphoma . However2.Human neuroblastoma cell lines SH-SY5Y were provided by American type culture collection (ATCC), USA. Cells were cultured in Dulbecco's modified Eagle's medium (DMEM) medium containing 10% FBS and 1% penicillin/streptomycin at 37\u00b0C with 5% CO5 cells) were incubated with 10\u2009\u03bcg/L of lipopolysaccharide (LPS) and/or L. pentosus strain S-PT84 .Cells . Then, PCR was performed using SYBR Premix Ex Taq . GAPDH served as loading control. Relative mRNA levels were calculated using the 2\u03b2, IL-18, procaspase1, cleaved caspase1, GSDMD-N, BIRC3, NLRP3, NLRC1, NLRP3, NLRC4, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) at 4\u00b0C overnight. Next day, the membranes were incubated with secondary antibodies. GAPDH was used as the loading control. Subsequently, the bands were captured using an efficient chemiluminescence (ECL) kit and analyzed using the ImageJ software.Total protein was extracted from SH-SY5Y cells. Protein concentration was measured using a bicinchonininc acid (BCA) kit. The protein was separated using 12% SDS-PAGE. Afterwards, the separated protein was moved onto polyvinylidene difluoride (PVDF) membranes, which was then sealed by 5% skimmed milk. The membranes were incubated with primary antibodies against IL-1Cells were collected and lysed. Then, cell lysates were centrifuged at 12,000 \u00d7 g and immunoprecipitated with specific antibodies. Afterwards, the proteins were coprecipitated and isolated using 12% SDS-PAGE. Immunoblotting was analyzed with antianalysis with the indicated primary antibodies against BIRC3, NLRC4, and GAPDH.3 cells/well). After incubated with L. pentosus for 48\u2009h, cells were cultured with Cell Counting Kit-8 (CCK-8) regents for 4\u2009h. Subsequently, optic density was determined using a microplate at 450\u2009nm.Cells were seeded into 24-well plates . Subsequently, a flow cytometry was used to calculate the pyroptosis rates: active caspase1\u2009+\u2009PI double positive cells/total cells\u2009\u00d7\u2009100%.Cells were collected and fixed with 4% paraformaldehyde. After washed with 5% PBS, cells were cultured with TUNEL solutions. Then, the cells were counterstained with DAPI. Finally, TUNEL positive cells were captured using a fluorescence microscope.t-test and one-way ANOVA. P < 0.05 dictated significant difference.All data were evaluated using GraphPad 6.0. and represented as mean\u2009\u00b1\u2009SD. The comparison was performed using Student's Lactobacillus pentosus (L. pentosus) on neuronal degeneration, cells were cultured with L. pentosus. As shown in To investigate the effects of L. pentosus, SH-SY5Y cells were exposed to LPS. As shown in Figures \u03b2 and IL-18, which was antagonized by L. pentosus. These results were consistent with that from western blot. L. pentosus treatment markedly alleviated the effects of LPS and suppressed the protein expression of IL-1\u03b2 and IL-18.Inflammation is a key factor for neuronal degeneration. To verify the effects of L. pentosus significantly decreased PI and active caspase1-positive cells induced by LPS. Additionally, LPS-mediated increase in TUNEL-positive cells was abated by L. pentosus treatment (L. pentosus on LPS-induced neuronal cell death, we determined the protein expression of pyroptosis biomarkers. LPS exposure significantly increased the protein expression of cleaved caspase1 and GSDMD-N (L. pentosus.Cellular functions were detected using flow cytometry and TUNEL assay. As shown in reatment . To furt GSDMD-N , which wL. pentosus treatment significantly increased BIRC3 mRNA levels compared with the LPS group (L. pentosus also increased the protein expression of BIRC3 (Previous studies report that BIRC3 plays a neuron-protective role. We then determined the expression of BIRC3 in neurons. As shown in PS group . Moreoveof BIRC3 .L. pentosus\u2009+\u2009sh-NC group. BIRC3 knockdown significantly suppressed the cell viability of SH-SY5Y cells (\u03b2 and IL-18 (Figures The rescue assay was performed to verify the roles of BIRC3 in neuronal degeneration. As shown in 5Y cells . Additio Figures .As shown in Figures We further investigated the underlying molecular mechanisms that BIRC3 inhibited neuronal pyroptosis. The online database STING showed that the potential genes interacts with BIRC3 . The resL. pentosus play a neuron-protective role. L. pentosus suppressed inflammatory response and pyroptosis of neuron cells. Moreover, L. pentosus upregulated BIRC3, suppressing the inactivation of NLRC4 inflammasome. Hence, L. pentosus may be a promising therapy for neurodegenerative diseases.In this study, \u03b2, and p-Tau to contribute to neuroinflammation and memory impairment [\u03b2 [L. pentosus treatment restored neuronal cellular functions, manifested by the increase in cell viability and decrease in pyroptosis rates. Previous studies evidence that L. pentosus promotes cognitive capability and alleviates aging-dependent memory impairment [L. pentosus may be a promising strategy for neurodegenerative diseases.The activation of inflammasomes increases cytotoxicity and contributes to the pyroptosis of neurons, which is a key factor for memory loss and cognitive impairment . For inspairment . High lerment [\u03b2 . NLRP1 drment [\u03b2 . These rrment [\u03b2 , which fpairment , 32. The\u03b2-induced neuronal apoptosis [L. pentosus alleviated LPS-induced downregulation of BIRC3. However, BIRC3 knockdown alleviated the effects of L. pentosus and promoted inflammation and pyroptosis of neurons. These results suggested that BIRC3 may play a neuron-protective role, which is consistent with previous studies [BIRC3 plays a vital role in neuronal function. For instance, NPD1-mediated upregulation of BIRC3 promotes neural cell survival . Additiopoptosis . Howeverpoptosis  as well poptosis . Hence,  studies .L. pentosus-mediated upregulation of BIRC3 contributed to inactivation of NLRC4 inflammasome, which suppressed neuronal pyroptosis.However, approximately 80% studies focused on the roles of BIRC3 cancer. The reports on its roles in neural disorders are limited. BIRC3 mainly exerts its neuron-protective functions via regulating caspases cascades, which suppresses the neuronal apoptosis . RecentlL. pentosus-induced upregulation of BIRC3 suppressed inflammatory response and pyroptosis of neurons via inactivating NLRC4 inflammasome. Therefore, L. pentosus may be an alternative strategy for neurodegenerative diseases.In conclusion,"}
+{"text": "Despite tremendous success of molecular targeted therapy together with immunotherapy, only a small subset of patients can benefit from them. Chemotherapy remains the mainstay treatment for most of tumors including non-small cell lung cancer (NSCLC); however, non-selective adverse effects on healthy tissues and secondary resistance are the main obstacles. Meanwhile, the quiescent or dormant cancer stem-like cells (CSLCs) are resistant to antimitotic chemoradiotherapy. Complete remission can only be realized when both proliferative cancer cells and quiescent cancer stem cells are targeted. In the present research, we constructed a cooperatively combating conjugate (DTX-P7) composed of docetaxel (DTX) and a heptapeptide (P7), which specifically binds to cell surface Hsp90, and assessed the anti-tumor effects of DTX-P7 on non-small cell lung cancer. DTX-P7 preferentially suppressed tumor growth compared with DTX in vivo with a favorable distribution to tumor tissues and long circulation half-life. Furthermore, we revealed a distinctive mechanism whereby DTX-P7 induced unfolded protein response and eventually promoted apoptosis. More importantly, we found that DTX-P7 promoted cell cycle reentry of slow-proliferating CSLCs and subsequently killed them, exhibiting a \u201cproliferate to kill\u201d pattern. Collecitvely, by force of active targeting delivery of DTX via membrane-bound Hsp90, DTX-P7 induces unfolded protein response and subsequent apoptosis by degrading Hsp90, meanwhile awakens and kills the dormant cancer stem cells. Thus, DTX-P7 deserves further development as a promising anticancer therapeutic for treatment of various membrane-harboring Hsp90 cancer types.The online version contains supplementary material available at 10.1186/s13045-022-01274-8. To the editor,To address off-target toxicity of conventional chemotherapy, one effort is to enhance the targeted delivery by conjugating therapeutic effector through a cleavable linker to a ligand specific to a drug target, and several conjugates of targeting agents have been approved for application in oncology , 2. HoweHerein, we designed and identified a peptide-conjugated drug comprising P7 and docetaxel (DTX) (namely DTX-P7)  and PI3K-Akt signaling pathway  Chemical structures of DTX and DTX-P7. DTX, molecular formula: C43H53NO14, molecular weight: 807.9 g/mol, white powder. DTX-P7, molecular formula: C84H118N8O25, molecular weight: 1,639.90 g/mol, white powder. b-c) Cell viability of DTX-P7 and DTX in A549 (b) and H1975 (c) cells following 48-h treatment. IC50 values: A549 cells, DTX-P7 11.4 nM, DTX 1.11 nM; H1975 cells, DTX-P7 0.62 nM, DTX 0.50 nM.Additional file 3: Supplementary Figure S2. Biodistribution of DTX-P7 in nude mice bearing A549 xenograft tumor. DTX-P7 was quantified by free DTX released from the conjugate. a) Plasma concentration of DTX and DTX-P7 in plasma samples throughout 72-h treatment. b-e) Distribution of DTX in the heart, liver, spleen, lungs, kidneys, and brain in 1 h (b), 2 h (c), 4 h (d) and 8 h (e) after DTX or DTX-P7 was administrated to mice implanted with A549 xenograft tumor. Data are given as mean \u00b1 SD (n = 3). * p < 0.05 vs. DTX group.Additional file 4: Supplementary Figure S3. Proteome changes induced in A549 cells by DTX-P7. a-c) Gene Ontology (GO) annotation analysis of A549 cell proteins that changed more than 1.5-ratio after DTX-P7 treatment, including altered proteins for biological process (a), cellular component (b) and molecular function (c) analyses. d) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation analysis of A549 cell proteins that changed more than 1.5-ratio after DTX-P7 treatment.Additional file 5: Supplementary Figure S4. Cell growth morphology of cancer stem cell-like A549/CD133+ cells and identification of cell surface Hsp90 in A549/CD133+ cells. a) Morphology of A549 and A549/CD133+ cells. b) Hsp90 expression levels were assessed in A549/CD133+ cells by cellular fractionation and Western blotting analysis. c) Immunofluorescence analysis of cell surface Hsp90 in A549/CD133+ cells. d) Immunofluorescence assays of FITC-labeled P7 binding to A549/CD133+ cells. Competition of P7 binding to A549/CD133+ cells by excess free P7.Additional file 6: Supplementary Figure S5. Effects of DTX-P7 on survival of A549/CD133+ cells and PKH26 staining of A549/CD133+ cells. a) Cell viability of DTX-P7 and DTX in A549/CD133+ cells following 48-h treatment. b) A549/CD133+ cells were stained by PKH26 followed by fluorescence detection at Day 0, 1, 6 and 10. c) Representative fluorescence-activated cell sorting profile of A549/CD133+ cells selected for sorting 10 days after PKH26 staining as compared with those of the unstained control (negative control) and Day 0."}
+{"text": "Fecal calprotectin (FC) is a sensitive marker of intestinal inflammation, and is used to both discriminate inflammatory bowel disease (IBD) from non-IBD patients and to monitor patients with IBD. It is unclear whether normal values established in adult patients are applicable in pediatrics.To evaluate FC\u2019s ability to differentiate IBD from non-IBD pediatric patients, and to understand factors influencing FC in pediatric IBD(pIBD).Stool FC samples collected on all patients<19 years of age in British Columbia(BC) from May 2020 to August 2022 were run using a Buhlmann ELISA at BC Children\u2019s Hospital(BCCH). The BCCH GI database identified patients with IBD. FC\u2019s ordered by adult IBD providers and patients awaiting endoscopy were excluded. The remaining samples were analysed as non-IBD. The sensitivity(Sn), negative predictive value(NPV) and false positive(FP) of FC were evaluated; comparisons were made using the Wilcoxon rank-sum test and chi-squared.3506 FC samples met inclusion criteria: 1853 IBD and 1653 non-IBD. 221 IBD samples were from prior to diagnosis, with median (IQR) FC 2615ug/g(1090-4183); median FC for non-IBD patients was 54ug/g(24-122). Using the Buhlmann \"normal\" cutoff of 80ug/g, the Sn was 0.991 (NPV 0.998) with a FP rate of 37%. Young patients were more likely to have FP's: <6yo's (n=305) had a FP rate of 42% vs 36% in those >6yo (n=1348)(p=0.035). With a FC cutoff of 160ug/g, Sn was 0.973 (NPV 0.996), with a FP rate of 20% . At a threshold of 250ug/g, Sn was 0.959 (NPV 0.994) with a FP rate of 13% . For patients <2yo, all 4 new IBD diagnosis had FC>1900ug/g. In the non-IBD population (n=69 samples <2yo), the FP rate was 52%, 30%, and 22% using a threshold of 80, 160, and 250ug/g, respectively.Evaluating FC as a disease-monitoring tool in IBD  found that at 6, 12, 18, and 24+ months post diagnosis, FC decreased from 750(159-1883), 505(110-1566), 351(87-1379), to 308ug/g(85-1129), respectively. Similarly, the proportion of FC\u2019s <250ug/g increased from 4.1% at diagnosis to 30%, 39.7%, 41.5%, 47% during the follow up period.Patients with UC/IBD-U had higher FC\u2019s, and were less likely to achieve FC<250ug/g. By 12 months post diagnosis, median FC of CD patients was 347ug/g(96-1150) and UC/IBD-U was 745ug/g(191-2017)(p=0.036), and at 18 months, CD 273ug/g(61-902) vs UC/IBD-U 932ug/g(144-2229)(p<0.001). At 2+yrs, median FC for CD patients was 259ug/g(76-1038) vs 387ug/g(108-1577) for UC/IC (p=0.017).Patients cared for by an IBD specialist had better FC outcomes vs those managed by non-IBD GIs: median FC at 12 months was 246ug/g(n=79) vs 677ug/g(n=153)(p=0.002), with 51% vs 34% achieving FC<250(p=0.014). Similarly, at 2+yrs post diagnosis, median FC was 243ug/g(n=437) vs 356ug/g(n=407)(p=0.02).Higher FC thresholds are likely required in younger populations compared to established adult cutoffs. In this pIBD cohort, <50% achieve FC levels <250ug/g.NoneNone Declared"}
+{"text": "We described emergency department (ED) visits by persons with multiple sclerosis (MS) in British Columbia, Canada (1 April2012 to 31 December 2017). We identified 15,350 MS cases using healthadministrative data; 73.4% were women, averaging 51.4\u2009years at study entry. Over4.9\u2009years of follow-up (mean), 56.0% of MS cases visited an ED . A diagnosis was documented for25,698 (69.3%) ED visits, and 18.4%  were infection-related.Inpatient admissions were reported for 20.4%  of all and 29.2%(1380/4725) of infection-related ED visits. Findings suggest that the ED plays asubstantial role in MS healthcare and infection management. For example,persons with MS had 41% more infection-related physician claims  versus a sex-, age-, andregion-matched non-MS population. However, relatively little is known about ED utilization by persons withMS.46 Here, we described overall andinfection-related ED use in an MS population.Emergency department (ED) presentations represent an important aspect of health careutilization in the general population.), including Medical Service Plan Billing Information (providing physician claims); the Discharge Abstract Database ; PharmaNet (for prescriptions filled at outpatient/community pharmacies); Census Geodata(providing socioeconomic status estimates); Registration and Premium Billing files ; Vital Statistics (capturing death dates); and the National Ambulatory Care Reporting System dataset  starting 1 April 2012).We performed a descriptive, population-based study in British Columbia (BC), Canada,using linked health administrative data  code, or first disease-modifyingdrug (DMD) prescription filled, or 1 April 2012 (start of the ED date). All includedpersons were \u2a7e18\u2009years old and BC residents for \u2a7e1\u2009year pre-study entry; follow-upended at the earliest of death, emigration, or 31 December 2017. Comorbidities weremeasured using a modified Charlson Comorbidity Index during the 1-year pre-studyentry.1618 MS cases everfilling a DMD prescription during follow-up were described.All MS cases were identified with a validated algorithm.n\u2009=\u20098603) with (vs without) comorbidity at study entrywere older (mean age\u2009=\u200956.5 (SD\u2009=\u200914.3) vs 49.5 (SD\u2009=\u200913.7) years) and averaged moreED visits/person/year (1.3 (SD\u2009=\u20092.4) vs 0.9 (SD\u2009=\u20091.9)). More than a quarter of MScases had \u2a7e3 ED visits ; of these 1383 (34.0%) of 4064 had \u2a7e1comorbidity at study entry. Most ED visits by MS cases did not require an ambulance; 28.8% required a ground ambulance  and>95% were of semi-urgent or higher priority .We identified 15,350 MS cases  women; Supplementary Table 2). These visits were made by 74.2%(6384/8603) of MS ED users. Of these, the most frequent primary diagnoses were\u201cabdominal pain/colic\u201d , \u201curinary tract infection\u201d, and \u201cMS\u201d . When combined, 18.4% of ED visits were infection-related with 32.2% (2056/6384) of MSparticipants having at least one such visit. Nearly one-third of infection-related ED visits led to hospitalization, while 20.4% of all ED visits made by 27.9% (2404/8603) of cases did so (Supplementary Tables 2 and 3).Diagnostic codes were reported for 69.3%  of ED visits (summarizedin Persons who filled \u2a7e1 DMD prescription(s) during follow-up (\u201cDMD users\u201d) wereyounger at study entry date than non-users (mean age\u2009=\u200941.9 (SD\u2009=\u200911.1) vs 53.8(SD\u2009=\u200913.3) years). However, the proportions accessing an ED at least once weregenerally similar , as was the study follow-up time (4.8\u2009years (SD\u2009=\u20091.6) for DMD usersvs 4.9 (SD\u2009=\u20091.6) for non-users).There were no clear patterns across the socioeconomic quintiles at study entryfor the MS cases: ever/never filling a DMD prescription during follow-up,ever/never being hospitalized subsequent to an ED visit, or for the five mostcommon ED diagnoses (data not shown).Over 50% of persons with MS had more than one ED consultation during our nearly6-year observation period; one-quarter visited an ED three or more times. Nearly 20%of ED visits were infection-related; one-third resulted in hospitalization, whereasone-fifth of all-cause ED visits did so. MS cases with \u2a7e1 (vs without) comorbidityat study entry averaged more ED visits/person/year. Our intentionally descriptivestudy has several limitations. While the overall burden of infection-related EDvisits was considerable in our MS population, this could be higher as diagnoses wereunavailable for one-third of all ED visits. Furthermore, our study lackedMS-specific clinical information, such as relapses, and a comparison to the generalpopulation. Our 1-year pre-study lookback period may have reduced the detection ofcomorbidities. Strengths of our study included the large cohort of MS casesidentified using a validated algorithm within a geographically defined populationwith universal healthcare coverage, including ED visits. Our results suggest thatthe ED visits by MS cases often lead to hospitalizations, perhaps more so forinfection-related visits. Further studies are necessary to better understand theimportance of ED use by persons with MS.Click here for additional data file.Supplemental material, sj-docx-1-msj-10.1177_13524585221078497 for Emergencydepartment use by persons with MS: A population-based descriptive study with afocus on infection-related visits by Jonas Graf, Huah Shin Ng, Feng Zhu, YinshanZhao, Jos\u00e9 MA Wijnands, Charity Evans, John D Fisk, Ruth Ann Marrie and HelenTremlett in Multiple Sclerosis JournalClick here for additional data file.Supplemental material, sj-docx-2-msj-10.1177_13524585221078497 for Emergencydepartment use by persons with MS: A population-based descriptive study with afocus on infection-related visits by Jonas Graf, Huah Shin Ng, Feng Zhu, YinshanZhao, Jos\u00e9 MA Wijnands, Charity Evans, John D Fisk, Ruth Ann Marrie and HelenTremlett in Multiple Sclerosis JournalClick here for additional data file.Supplemental material, sj-docx-3-msj-10.1177_13524585221078497 for Emergencydepartment use by persons with MS: A population-based descriptive study with afocus on infection-related visits by Jonas Graf, Huah Shin Ng, Feng Zhu, YinshanZhao, Jos\u00e9 MA Wijnands, Charity Evans, John D Fisk, Ruth Ann Marrie and HelenTremlett in Multiple Sclerosis Journal"}
+{"text": "The immunopathogenesis of chronic spontaneous urticaria (CSU) is poorly understood, but recent research suggests that patients can be divided into autoallergic and autoimmune subtypes. Given that not all patients can be controlled with current treatment regimens, including anti-IgE monoclonal antibodies, a better understanding of the immune pathways involved in CSU may enable the repurposing of monoclonal antibodies used for other dermatologic diseases . Therefore, we investigated the implicated immune cells and pathways by reanalyzing publicly available transcriptomic data.2 fold change| \u22651. Pathway analyses were conducted using ToppGene and KEGG. Cell-type enrichment was determined by CIBERSORT and xCell and was correlated with clinical characteristics.Microarray data of CSU and healthy control (HC) skin and blood were obtained from the Gene Expression Omnibus . Differentially expressed genes were defined as a false discovery rate <0.05 and a |logTh2  and Th17-related  pathways were upregulated in lesional compared to non-lesional and HC samples. In non-lesional versus lesional samples, CIBERSORT analysis revealed increased regulatory T-cells (Treg) and resting mast cells. xCell analysis established that Th1 and Th2 scores were not significantly different between lesional and HC samples. However, Th2 scores in both lesional and non-lesional samples correlated positively with disease severity. Few differentially expressed genes and pathways were identified between CSU and HC blood samples.Our results support the involvement of Th2 and Th17-related genes and pathways in CSU. Th2 scores associate with disease severity, which indicates the clinical relevance of these findings. Increased resting mast cell and Treg scores in non-lesional samples may suggest local suppression of wheal formation. Moreover, disease activity seemed to be restricted to the skin as there were limited findings from blood. Larger studies using next-generation sequencing will be helpful to confirm these results. Chronic spontaneous urticaria (CSU) is defined by the presence of wheals and/or angioedema occurring in the absence of specific external stimuli and persisting for \u2265 6 weeks . CSU is There has been a paradigm shift in the understanding of the pathogenesis of CSU and current evidence suggests that most CSU cases have an autoimmune etiology , 7. AutoSeveral studies support the involvement of dysfunction of adaptive cellular immunity in the immune pathogenesis of CSU , 11. Curin silico quantification of various cell-types from transcriptomic data of a heterogeneous cell population. CIBERSORT is one such RNA deconvolution technique that employs linear support vector regression to quantify relative proportions of cell types  (GSE57178) (Transcriptomic data acquired by microarray were obtained from the Gene Expression Omnibus (GEO) . From GiSE72542) , data waSE57178) , data onDifferentially expressed genes (DEGs) were determined using the GEO2R  online tgetGEO function from the GEOquery package . Select DEGs were presented by violin plots.DEG analysis was performed by The McGill Genome Centre, Montreal, Quebec. Datasets were analyzed separately. For each dataset, the normalized expression values were downloaded from the GEO database using the  package , and line  and 100 permutations to estimate the relative proportions of 22 cell-types . Cell abP-values and Q-values less than 0.05 were considered significant. All statistical tests were performed on RStudio version 1.4.1717 and graphs and figures were rendered using the Ggplot2 package , neutrophil degranulation (Q = 1.69E-23), innate immune system (Q = 4.50E-17), cytokine signaling in immune system (Q = 1.82E-14), staphylococcus aureus infection (Q = 2.85E-11), and signaling by interleukins (Q = 3.05E-11). Th2-related pathways including IL-4 and IL-13 mediated signaling (Q = 1.03E-13) and IL-18 signaling (Q = 2.05E-7) were upregulated. Additionally, IL-10 signaling (Q = 8.42E-14), an immunosuppressive pathway, and IL-12 mediated signaling (Q = 3.80E-3), a Th1 polarizing cytokine, were upregulated (Q = 1.04E-3), an activator directly upstream of the Th17 response, as well as IL-17 signaling (Q = 3.60E-6), and IL-6 mediated signaling events (Q = 8.28E-4)  .Q = 4.42E-15), cytokine-receptor interaction (Q = 3.05E-12), and TNF signaling (Q = 4.02E-8) (Q = 3.75E-8) and Th17 cell differentiation (Q = 0.017) (Upregulated KEGG pathways comprise staphylococcus aureus infection (4.02E-8) . In line= 0.017) . Genes u= 0.017) , whereas= 0.017) . IndividIL-4R was upregulated , cytokine signaling in immune system (Q = 1.70E-10), signaling by interleukins (Q = 2.60E-10), neutrophil degranulation (Q = 1.38E-9), IL-4 and IL-13 signaling (Q = 2.15E-9), IL-18 signaling (Q = 1.72E-4), IL-17 signaling (Q = 1.04E-3), IL-23 mediated signaling events (Q = 5.59E-4), IL-6 mediated signaling events (Q = 1.28E-3), and IL-12 mediated signaling events (Q = 2.57E-2). Additionally, the IL-4 mediated signaling events pathway (Q = 2.89E-2) was also upregulated , TNF signaling (Q = 9.20E-8), and staphylococcus aureus infection (Q = 5.06E-7) (Q = 9.20E-8) and Th17 cell differentiation (Q = 0.019) were upregulated  . Moreoveegulated . Upregulegulated  and Th17egulated  pathwaysQ = 0.033), Th17 cell differentiation (Q = 0.027), IL-5 signaling pathway and Th1/Th2 differentiation (Q = 0.027). Similarly, KEGG analysis demonstrated enriched Th1 and Th2 cell differentiation (Q = 0.045) and Th17 cell differentiation (Q = 0.045). These results must be interpreted with caution, however, because HLA-DRB3  was the only upregulated gene involved in each of these pathways.Five genes were upregulated and seven downregulated in the 20 CSU versus 10 HC blood samples. ToppGene pathway analysis of upregulated genes revealed enriched Th1 and Th2 cell differentiation (Q = 0.029) and CD8+ T-cells (Q = 0.0066) were decreased, whereas CD4+ resting memory T-cells (Q = 0.0081) were increased. Eosinophils were only detected in some lesional skin samples, but not statistically significantly greater than in HC samples. No significant differences were observed in B-cells, natural killer (NK) cells, macrophages, DCs, neutrophils, or Tregs.RNA deconvolution of 15 lesional and 12 HC skin samples was performed by CIBERSORT . In lesiQ = 0.020), CD4+ memory T-cells (Q = 0.039), common lymphoid progenitor cells (Q = 0.025), and class switched memory B-cells (Q = 0.015) had increased cell abundance scores in lesional compared to HC samples. No significant differences were observed in Th1 (Q = 0.30) or Th2 (Q = 0.55) cells; however, gamma delta T-cells (Tgd) had elevated cell abundance scores in lesional samples (Q = 0.015). Th2 scores in lesional samples correlated moderately with UAS7 (p = 0.044). Innate immune cells including M1 macrophages (Q = 0.022), mast cells (Q = 0.039), monocytes (Q = 0.015), and neutrophils (Q = 0.0055) had increased scores in lesional samples in contrast to HC samples. Similarly, total dendritic cells (DC) were elevated in lesional versus HC samples (Q = 0.0010), including activated dendritic cells (aDC) (Q = 0.0010), conventional dendritic cells (cDC) (Q = 0.047), immature dendritic cells (iDC) (Q = 0.0050), and plasmacytoid dendritic cells (pDC) (Q = 0.037). Interestingly, neurons (Q = 0.013) were decreased in lesional compared to HC samples. While basophils were detected in both lesional and HC samples, abundance scores were not significantly different.Cell abundance scores determined by xCell were compared between lesional and HC skin samples . CD4+ efQ = 0.039) in non-lesional samples. Interestingly, Tregs (Q = 0.014) and resting mast cells (Q = 0.014) were increased in non-lesional compared to lesional samples. Eosinophils were only detected in some lesional skin samples, but not statistically significantly greater than non-lesional samples. We did not detect any significant differences in B-cells, NK cells, macrophages, DCs, or neutrophils between lesional and non-lesional samples on CIBERSORT.CIBERSORT RNA deconvolution of 15 lesional versus 15 matched non-lesional samples  revealedQ = 0.012). Moreover, Th2 cell abundance scores of non-lesional samples correlated strongly with UAS7 scores (p = 0.0028), whereas Th1 cell abundance scores correlated negatively with UAS7 scores (p = 0.012). Basophils were detected in both groups but were not significantly different.xCell analysis of lesional versus non-lesional skin samples  revealedQ = 0.018) and decreased monocytes (Q = 0.018) in CSU blood compared to HC blood. No eosinophils were detected in either CSU or HC blood and no significant differences were detected in other cell-types. Although neutrophils were not enriched in CSU blood samples compared to HC blood, neutrophil CIBERSORT scores strongly positively correlated with UAS7 scores (p = 0.00099).RNA deconvolution using CIBERSORT of 20 CSU blood samples and 10 HC blood samples  revealedp = 0.007) and Th2 (p = 0.042) cell abundance scores correlated negatively with UAS7 scores. Basophils were only detected in CSU samples but not statistically significantly greater than in HC samples. Moreover, Th2 cells were significantly decreased in ASST+ samples compared to ASST\u2013 samples (p = 0.019), but no difference was observed in Th1 cells (p = 0.82).xCell analysis did not detect differences in cell-type abundance scores between CSU and HC blood samples. Among CSU blood samples, Th1 (Q = 0.014) and resting DCs (Q = 0.014) differed on CIBERSORT analysis. However, significant differences were identified in five cell-types by CIBERSORT and 16 cell-types by xCell in HC samples and six cell-types by CIBERSORT and 13 cell-types by xCell in non-lesional samples.The Gim\u00e9nez-Arnau et al.  and PateRecently, the role of Th2 dysregulation has been proposed as a potential driver of CSU pathogenesis in at least a subset of CSU patients , 31. Th2Th2 response is not only important for IgE antibody production, but also stimulates IgG1 complement binding and IgG4 antibodies underlying other autoimmune conditions . InteresIL-10 expression was shown to be increased in CSU lesions and a CSU mouse model overexpressing IL-10 exhibited increased pruritus and increased IL-2, IL-4, and IFN-\u03b3 expression, driven by JAK/STAT signaling  or whether it is an insufficient immunosuppressive response.We showed that the IL-4, IL-13, and IL-18 cytokine signaling pathways were upregulated in lesional CSU biopsies versus both HC and non-lesional samples, which suggest a role in the wheal response. Other studies have shown that cytokines that initiate the Type 2 response  , 34 are ignaling . It is uContrary to a previous study reporting increased Th2 cells in CSU lesions , we did IL-6) were also upregulated in lesional samples compared to non-lesional and HC samples. IL-17 and IL-23 have been shown to be elevated in the serum of CSU patients compared to controls and positively correlate with UAS7 scores . A Th1 response is usually associated with ASST positivity , but in In vivo and in vitro evidence indicate a role for Tregs in the suppression of mast cell degranulation via OX40-OX40L signaling  can be found in the article/PL, EN, IL, and CP designed the study. CP, PL, SG, and ML performed the data analysis. CP and SG prepared the figures and tables. EN, PL, IL, MB-S, and AG-A supervised the study. All authors contributed to writing and editing the manuscript.EN was a consultant and speaker for Novartis and has received an investigator-initiated research grant from Novartis. MB-S was a consultant for Novartis. AG-A was a medical advisor for Uriach Pharma, Genentech, Novartis, FAES, GSK, Sanofi\u2013Regeneron, Amgen, Thermo Fisher Scientific, Almirall, she has research grants supported by Uriach Pharma, Novartis, Grants from Instituto Carlos III- FEDER and performs educational activities for Uriach Pharma, Novartis, Genentech, Menarini, LEO-PHARMA, GSK, MSD, Almirall, Sanofi\u2014Regeneron, AVENE. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Globally, a high-salt diet (HSD) has become a threat to human health as it can lead to a high risk of cardiac damage. Although some studies investigating HSD have been carried out, the majority has been conducted in males, and there are few female-specific studies, thereby ignoring any effects of sex-specific damage on the heart. In this study, we determined how HSD induces different pathways of cardiovascular diseases through sex-specific effects on cardiac damage in mice.An HSD murine model of male and female C57BL/6J mice was fed with sodium-rich chow (4% NaCl). After 8 weeks, cardiac tissues were collected, and the whole gene transcriptome of the hearts of male and female mice was characterized and analyzed using high-throughput RNA sequencing. Immunohistochemistry staining was used to further assess the harmful effects of HSD on protein expression of genes associated with immunity, fibrosis, and apoptosis in male and female mice.HSD drastically altered the cardiac transcriptome compared to that of the normal heart in both male and female mice and had a sex-specific effect on the cardiac composition in the transcriptome. HSD produced various differentially expressed genes and affected different KEGG pathways of the transcriptome in male and female mice. Furthermore, we found that HSD induced different pathways of cardiovascular disease in the male mice and female mice. The pathway of hypertrophic cardiomyopathy is significantly enriched in HSD-treated male mice, while the pathway of dilated cardiomyopathy is significantly enriched in HSD-treated female mice. Finally, metabolism, immunity, fibrosis, and apoptosis in the mouse heart showed sex-specific changes predicting cardiac damage.Our results demonstrate that HSD adversely impacts cardiac structure and function by affecting the metabolism, immunity, fibrosis, and apoptosis in the murine heart and induces the mouse to suffer from sex-specific cardiovascular disease. This study provides a new perspective and basis for the differences in the pharmacology and interventional treatment of sex-specific cardiovascular diseases induced by HSD in men and women. High salt content in diet poses various health risks and contributes to the increase in disease prevalence in high-income countries , 2. SaltExisting evidence suggests that a high-salt diet (HSD) is related to cardiovascular disease , 12, chrTo investigate the effect of sex-specific damage of HSD on the structure and function of heart tissues, we generated an HSD murine model by administering male and female C57BL/6J mice with sodium-rich chow containing 4% NaCl. The whole gene transcriptome of the mouse HSD-treated heart was characterized and analyzed by high-throughput RNA sequencing. This study determined that HSD induces cardiac dysfunction and structural damage and possibly cardiovascular disease by inducing changes to metabolism, immune response, fibrosis, and apoptosis in the hearts of male and female mice. Our research may help to reveal the molecular mechanisms of physiological damage caused by HSD to cardiac tissue and is of great significance for the development of therapeutic intervention methods for HSD-induced cardiovascular-related diseases.A schematic of the experimental design is shown in http://www.gempharmatech.com). Before starting the experiment, all mice were placed in a 12-h LD cycle to adapt to the environment for 2 weeks. Male and female mice were fed a normal diet and HSD (4% NaCl) as the normal and high-salt groups, respectively. All experimental protocols were approved by the Institutional Animal Care and Use Committee of Henan Province People's Hospital.Twelve wild-type C57BL/6J male mice and female mice (6\u20138 weeks) were purchased from GemPharmatech Co., Ltd . Echocardiography of mice anesthetized with isoflurane was performed in M-mode. Fractional shortening (FS), left ventricular (LV) internal dimension at end-diastole (LVIDd), and diastolic left ventricular thickness (LVPWd) were collected.After 8 weeks following a normal diet or HSD, the hearts of mice were rapidly harvested and weighed after death by cervical dislocation. Tissues were immediately immersed in liquid nitrogen, transported, and stored in a freezer set to maintain \u221280\u00b0C.Immunohistochemistry (IHC) analysis was performed on murine heart samples according to a previously published protocol . ParaffiMasson staining of murine hearts was performed according to a previously published study . All carAll cardiac sections from CONMH, MH8W, CONFH, and FH8W groups were placed in xylene I-III  for 15\u201320 min and then rinsed with 75\u2013100% ethanol  for 5 min and rinsed with distilled water. The cardiac sections were stained with hematoxylin and eosin dye solution from a hematoxylin-eosin (HE) staining Kit . After the slices were dehydrated, they were mounted with neutral gum . Microscopy, image acquisition, and analysis of cardiac sections were performed using an orthotopic light microscope . For quantification of cardiac cell size, 20 400X field images were randomly selected in a double-blind manner and cell numbers in each field were counted by ImageJ software . Average cell size is calculated as the number of cells in the field divided by the area of the field.http://bgitechsolutions.com/sequencing). Frozen mouse heart tissue was broken up using a mortar and pestle to obtain a piece, macroscopically free of fatty infiltration, fibrosis, and blood. RNA was extracted using the RNA Easy Spin Column Kit  following the TRIzol RNA extraction protocol.All RNA extractions were performed by the Beijing Genomic Institute .The RNA library was prepared according to a previously published study  using thpost-hoc test was set as Tykey-Kramer with 0.95, and the effect size was set as Eta-squared. The other parameters were set to default values.Principal component analysis (PCA) was performed using the Statistical Analysis of Metagenomic Profiles (STAMP) software package . PCA washttp://www.genome.jp/kegg) and NCBI RefSeq GCF_000001635.25_GRCm38.p5 as the reference gene set. Pathways with q \u2264 0.05 were defined as significantly enriched in differentially expressed genes (DEGs). The P value formula used in this study is given as follows:Kyoto encyclopedia of genes and genomes pathway annotation was performed using BLASTALL software  against the KEGG database version 81.0  \u2264 0.05, as the threshold.where rrection , with q http://www.geneontology.org) and calculates the number of genes for each term. The hypergeometric test was then applied to determine the significance of the biological functions of the candidate genes in comparison to the different genetic backgrounds of this species. Referring to the software \u201cGO:: TermFinder\u201d , the calculation formula of this analysis is the same as the KEGG enrichment analysis, N is the number of genes with GO annotations, n is the number of candidate genes in N, and M is the number of genes annotated with a specific GO term in all genes. The calculated P-value was determined using the Bonferroni correction, with q  \u2264 0.05 as the threshold. GO terms that met this condition were defined as significantly enriched.The molecular function and biological process of genes were analyzed by Gene Ontology (GO) , 32. Sighttps://string-db.org/, version 11.5) (Protein-protein association networks (PPANs) for fibrosis, metabolism, immunity, and apoptosis were analyzed using STRING (on 11.5) . The netP < 0.05.GraphPad Prism software  was used to generate bar charts and scatter plots. The Venn diagram plotter, a network tool , was used to compare the number of transcripts between the normal and HSD-treated groups. The expression of transcripts in the normal and HSD groups was visualized using the pheatmap package in R . The data were analyzed using the two-way analysis of variance (ANOVA). The data are presented as mean values \u00b1 SEM and were considered statistically significant when P = 0.04) as well as a significant main effect of sex (P < 0.001) and HSD (P = 0.004) on body weight. These analyses revealed that the body weight of HSD-treated mice was significantly lower than that of the normal group in both male and female mice (P = 0.002) and sex (P = 0.024), but not sex \u00d7 HSD interaction (P = 0.067) on ratios of HW/BW (P < 0.001) and sex (P = 0.031), but not sex \u00d7 HSD interaction (P = 0.082) on FS; (2) a significant main effect of HSD (P = 0.003) and sex (P = 0.034), but not sex \u00d7 HSD interaction (P = 0.067) on LVIDd; (3) a significant sex \u00d7 HSD interaction (P = 0.012) as well as a significant main effect of sex (P = 0.044) and HSD (P = 0.006) on LVPWd. To study the effect of HSD on cardiomyocyte size, HE staining was performed. A two-way ANOVA demonstrated a significant sex \u00d7 HSD interaction (P = 0.037) as well as a significant main effect of HSD (P < 0.001) on cell size. In males, the cell size of cardiomyocytes  demonstrated a significant sex \u00d7 HSD interaction  and high-salt-treated (MH8W) groups, respectively ; for femQ value <0.05) was compared between the CONMH and MH8W groups, CONFH and FH8W groups, and CONMH and CONFH groups, respectively. As shown in Principal component analysis was used for data visualization to determine transcriptomic differences between the CONMH and MH8W groups, and CONFH and FH8W groups, respectively . The dimTo investigate the effect of excessive sodium intake on the higher expression genes, we compared the difference between the CONMH and MH8W groups, and CONFH and FH8W groups using Venn plots. As shown in P < 0.05) were identified as being unique to the CONMH group and were divided into four categories: (1) cellular processes: tight junction (P = 2.27E-03), focal adhesion (P = 8.22E-03), and lysosome (P = 4.15E-02); (2) environmental information processing: PI3K-akt signaling pathway (P = 2.89E-03), ECM-receptor interaction (P = 9.86E-03), and Rap1 signaling pathway (P = 1.37E-02); (3) human diseases: AGE-RAGE signaling pathway in diabetic complications (P = 2.39E-04), transcriptional mis-regulation in cancer (P = 3.41E-04), human T-cell leukemia virus 1 infection (P = 3.09E-02), and systemic lupus erythematosus (P = 2.85E-02); (4) organismal systems: oxytocin signaling pathway (P = 3.32E-04), complement and coagulation cascades (P = 3.15E-03), estrogen signaling pathway (P = 1.86E-03), platelet activation (P = 4.45E-03), relaxing signaling pathway (P = 5.37E-03), dopaminergic synapse (P = 6.44E-03), protein digestion and absorption (P = 1.34E-02), cholesterol metabolism (P = 1.85E-02), glucagon signaling pathway (P = 2.11E-02), circadian rhythm (P = 2.38E-02), glutamatergic synapse (P = 3.02E-02), cholinergic synapse (P = 2.90E-02), and adrenergic signaling in cardiomyocytes (P = 3.91E-02) (P < 0.05) were identified in the shared CONMH and MH8W groups and were divided into five categories: (1) cellular processes: autophagy-animal (P = 8.05E-10) and endocytosis (P = 1.02E-07); (2) environmental information processing: phosphatidylinositol signaling system (P = 1.46E-09) and TNF signaling pathway (P = 2.18E-08); (3) genetic information processing: ribosome biogenesis in eukaryotes (P = 5.72E-10), ubiquitin mediated proteolysis (P = 2.31E-09), and RNA degradation (P = 6.35E-08); (4) human diseases: herpes simplex virus 1 infection (P = 1.44E-10) and colorectal cancer (P = 1.80E-08); (5) metabolism: inositol phosphate metabolism (P = 8.35E-09) (P < 0.05) were identified as being unique to the MH8W group and were divided into four categories: (1) environmental information processing: ECM-receptor interaction (P = 1.09E-06), PI3K-Akt signaling pathway (P = 5.40E-04), and WNT signaling pathway (P = 3.63E-03); (2) human diseases: arrhythmogenic right ventricular cardiomyopathy  and dilated cardiomyopathy ; (3) metabolism: tyrosine metabolism (P = 3.96E-03), phenylalanine metabolism (P = 1.07E-02), histidine metabolism (P = 1.25E-02), and glycolysis/gluconeogenesis (P = 1.31E-02); (4) organismal systems: regulation of lipolysis in adipocytes (P = 5.40E-03)  . A total.35E-09) . While 1.40E-03) .P < 0.01) were identified as being unique to the CONFH group and were divided into three categories: (1) environmental information processing: AMPK signaling pathway (P = 8.81E-04), TNF signaling pathway (P = 2.13E-03), PI3K-Akt signaling pathway (P = 4.27E-03), and cytokine-cytokine receptor interaction (P = 7.40E-03); (2) human diseases: transcriptional mis-regulation in cancer (P = 2.66E-05), rheumatoid arthritis (P = 4.23E-04), hypertrophic cardiomyopathy , acute myeloid leukemia (P = 5.06E-03), measles (P = 7.51E-03), and systemic lupus erythematosus (P = 7.51E-03); (3) organismal systems: IL-17 signaling pathway (P = 3.24E-03), chemokine signaling pathway (P = 4.18E-03), intestinal immune network for IgA production (P = 4.33E-03), and adipocytokine signaling pathway (P = 5.38E-03) (P < 0.001) were identified in the shared CONFH and FH8W groups and were divided into six categories: (1) cellular processes: autophagy-animal (P = 1.33E-08), endocytosis (P = 1.63E-05), cell cycle (P = 1.69E-05), adherens junction (P = 8.86E-05), lysosome (P = 1.17E-04), apoptosis (P = 1.34E-04), signaling pathways regulating pluripotency of stem cells (P = 6.37E-04), and apoptosis-fly (P = 9.50E-04); (2) environmental information processing: phosphatidylinositol signaling system (P = 6.48E-08), TNF signaling pathway (P = 1.84E-07), notch signaling pathway (P = 2.33E-07), MAPK signaling pathway (P = 4.83E-07), ERBB signaling pathway (P = 5.01E-07), WNT signaling pathway (P = 2.14E-06), hippo signaling pathway-fly (P = 4.95E-05), RAS signaling pathway (P = 5.79E-05), rap1 signaling pathway (P = 7.87E-05), mTOR signaling pathway (P = 1.29E-04), NF-kappa B signaling pathway (P = 7.18E-04), and hippo signaling pathway-multiple species (P = 3.40E-10); (3) genetic information processing: ribosome biogenesis in eukaryotes (P = 6.93E-05), ubiquitin mediated proteolysis (P = 2.04E-04), RNA degradation (P = 2.59E-04), DNA replication (P = 3.89E-04), basal transcription factors (P = 4.25E-04), nucleotide excision repair (P = 4.46E-04), RNA transport (P = 9.61E-04), mismatch repair (P = 2.88E-06), non-homologous end-joining (P = 1.32E-05), base excision repair (P = 9.84E-05), spliceosome (P = 4.92E-04), Fanconi anemia pathway (P = 6.30E-04), and aminoacyl-tRNA biosynthesis (P = 8.04E-04); (4) human diseases: herpes simplex virus 1 infection (P = 3.40E-10), microRNAs in cancer (P = 7.55E-09), pathways in cancer (P = 1.32E-07), colorectal cancer (P = 6.54E-07), pancreatic cancer (P = 6.45E-06), hepatitis B (P = 1.19E-05), endocrine resistance (P = 1.93E-05), breast cancer (P = 6.12E-05), chronic myeloid leukemia (P = 1.23E-04), small cell lung cancer (P = 1.70E-04), prostate cancer (P = 1.95E-04), endometrial cancer (P = 1.96E-04), hepatocellular carcinoma (P = 2.80E-04), insulin resistance (P = 3.82E-04), AGE-RAGE signaling pathway in diabetic complications (P = 3.82E-04), non-small cell lung cancer (P = 4.08E-04), human papillomavirus infection (P = 5.64E-04), and choline metabolism in cancer (P = 6.17E-04); (5) metabolism: N-Glycan biosynthesis (P = 1.49E-08), inositol phosphate metabolism (P = 1.07E-07), pyrimidine metabolism (P = 2.00E-06), lysine degradation (P = 1.27E-05), terpenoid backbone biosynthesis (P = 5.54E-05), glycosylphosphatidylinositol (GPI)-anchor biosynthesis (P = 6.93E-05), one carbon pool by folate (P = 2.04E-04), purine metabolism (P = 2.59E-04), amino sugar and nucleotide sugar metabolism (P = 3.89E-04), other types of O-glycan biosynthesis (P = 4.25E-04), other glycan degradation (P = 4.46E-04), and seleno-compound metabolism (P = 9.61E-04); (6) organismal systems: axon guidance (P = 2.88E-06), FC gamma R-mediated phagocytosis (P = 1.32E-05), thyroid hormone signaling pathway (P = 9.84E-05), chemokine signaling pathway (P = 4.92E-04), B cell receptor signaling pathway (P = 6.30E-04), osteoclast differentiation (P = 8.04E-04) (P < 0.01) were identified as being unique to the FH8W group and were divided into five categories: (1) cellular processes: gap junction (P = 5.33E-06) and focal adhesion (P = 4.93E-05); (2) environmental information processing: ECM-receptor interaction (P = 6.80E-06), Rap1 signaling pathway (P = 3.88E-05), PI3K-Akt signaling pathway (P = 7.46E-05), calcium signaling pathway (P = 3.39E-03), MAPK signaling pathway (P = 4.49E-03), and WNT signaling pathway (P = 6.38E-03); (3) human diseases: melanoma (P = 4.34E-03), hypertrophic cardiomyopathy , dilated cardiomyopathy , herpes simplex virus 1 infection (P = 6.02E-03), and arrhythmogenic right ventricular cardiomyopathy ; (4) metabolism: tyrosine metabolism (P = 5.33E-03); (5) organismal systems: parathyroid hormone synthesis, secretion and action (P = 2.29E-04), platelet activation (P = 2.29E-04), relaxing signaling pathway (P = 3.79E-03), oxytocin signaling pathway (P = 4.36E-03), thyroid hormone signaling pathway (P = 6.22E-03), and circadian entrainment (P = 6.84E-03)  . A total.04E-04) . While 2.84E-03) .P < 0.01) were identified as being unique to the CONMH group and were divided into four categories: (1) cellular processes: endocytosis (P = 7.91E-03); (2) environmental information processing: calcium signaling pathway (P = 9.34E-03); (3) human diseases: hepatocellular carcinoma (P = 1.23E-03), pancreatic cancer (P = 6.12E-03), and human papillomavirus infection (P = 8.82E-03); (4) organismal systems: vitamin digestion and absorption (P = 5.66E-03), cholesterol metabolism (P = 6.60E-03), and oxytocin signaling pathway (P = 9.12E-03)  were identified as being unique to the CONFH group and were divided into three categories: (1) environmental information processing: PI3K-Akt signaling pathway (P = 8.93E-04), cytokine-cytokine receptor interaction (P = 5.96E-03), and WNT signaling pathway (P = 9.59E-03); (2) human diseases: transcriptional mis-regulation in cancer (P = 6.20E-04), rheumatoid arthritis (P = 2.92E-03), herpes simplex virus 1 infection (P = 7.40E-03), Kaposi sarcoma-associated herpesvirus infection (P = 8.08E-03), and microRNAs in cancer (P = 9.96E-03); (3) organismal systems: IL-17 signaling pathway (P = 8.99E-04), B cell receptor signaling pathway (P = 1.67E-03), osteoclast differentiation (P = 6.27E-03), and axon guidance (P = 6.74E-03)  . Top 10 .12E-03) , which e.74E-03) . AltogetQ = 3.17E-22), dilated cardiomyopathy , arrhythmogenic right ventricular cardiomyopathy , fluid shear stress and atherosclerosis (Q = 1.37E-12), and viral myocarditis (Q = 1.65E-03) (Q = 1.43E-07), peptide antigen binding (Q = 1.91E-05), beta-2-microglobulin binding (Q = 3.35E-05), signaling receptor binding (Q = 5.43E-05), integrin binding (Q = 5.43E-05), T cell receptor binding (Q = 5.43E-05), CX3C chemokine binding (Q = 3.67E-04), metal ion binding (Q = 3.67E-04), neuregulin binding (Q = 4.95E-04), protein-containing complex binding (Q = 6.58E-04), and TAP binding (Q = 9.87E-04); (2) transporter activity, voltage-gated calcium channel activity (Q = 1.91E-05). As shown in Q = 2.31E-06), cell adhesion (Q = 2.33E-05), heterotypic cell-cell adhesion (Q = 3.57E-04), endocardial cushion fusion (Q = 4.71E-04), integrin-mediated signaling pathway (Q = 2.31E-06), positive regulation of T cell mediated cytotoxicity (Q = 7.00E-05), positive regulation of cell migration (Q = 1.10E-04), and positive regulation of epithelial to mesenchymal transition involved in endocardial cushion formation (Q = 6.21E-04); (2) cellular process: mesodermal cell differentiation (Q = 4.31E-05), cell-cell adhesion in response to extracellular stimulus (Q = 1.15E-04), endodermal cell differentiation (Q = 3.76E-04), positive regulation of vascular smooth muscle cell proliferation (Q = 6.21E-04), and cellular response to amyloid-beta (Q = 7.42E-04); (3) developmental process: heart development (Q = 2.31E-06), inner ear development (Q = 2.44E-04), ventricular cardiac muscle tissue morphogenesis (Q = 4.41E-04), and atrial septum primum morphogenesis (Q = 6.21E-04); (4) immune system process: antigen processing and presentation of peptide antigen via MHC class I (Q = 2.33E-05), antigen processing and presentation of endogenous peptide antigen via MHC class Ib (Q = 2.33E-05), antigen processing and presentation of endogenous peptide antigen via MHC class I via ER pathway, TAP-independent (Q = 2.33E-05), and antigen processing and presentation of endogenous peptide antigen via MHC class I via ER pathway, TAP-dependent (Q = 6.21E-04); (5) localization: calcium ion transmembrane transport (Q = 8.76E-05) and cell migration (Q = 1.15E-04); (6) response to stimulus: response to drug (Q = 4.71E-04) and response to hypoxia (Q = 4.81E-04).Evidence from clinical trials indicates that excessive sodium intake is associated with cardiovascular disease , 39. To .65E-03) . The KEGQ = 1.77E-14), ARVC (Q = 9.55E-12), HCM (Q = 3.03E-11), fluid shear stress and atherosclerosis (Q = 1.25E-06), and viral myocarditis (Q = 2.08E-05) , RNA polymerase II activating transcription factor binding (Q = 1.20E-03), transcription factor binding (Q = 1.63E-03), histone methyltransferase binding (Q = 1.63E-03), metal ion binding , R-SMAD binding (Q = 4.48E-03), integrin binding , laminin binding (Q = 5.07E-03), disordered domain-specific binding (Q = 7.41E-03), peptide antigen binding , and protein binding (Q = 8.44E-03); (2) transporter activity: calcium- and calmodulin-responsive adenylate cyclase activity (Q = 6.81E-04), adenylate cyclase activity (Q = 1.63E-03), and phosphorus-oxygen lyase activity (Q = 4.26E-03). As shown in Q value 2.03E-04 vs. 2.33E-05), cell-matrix adhesion , and integrin-mediated signaling pathway ; (2) cellular process: cellular response to cadmium ion (Q = 4.43E-04) and cellular response to reactive oxygen species (Q = 5.75E-04); (3) immune system process: antigen processing and presentation of peptide or polysaccharide antigen via MHC class II (Q = 5.79E-06), antigen processing and presentation of exogenous peptide antigen via MHC class II (Q = 5.24E-05), antigen processing and presentation of peptide antigen (Q = 4.47E-04), and antigen processing and presentation (Q = 6.03E-04); (4) response to stimulus: response to drug . Collectively, these results demonstrate that excessive sodium intake can easily induce cardiovascular disease.To further elucidate the effect of excessive sodium intake, we next compared the expression of cardiovascular disease-related transcripts between normal and high-salt-treated hearts in female mice. As shown in .08E-05) . KEGG an.08E-05) . GO analQ < 0.05)  . As show < 0.05) , S5 and  < 0.05) .F = 9.958; P = 0.002) as well as a significant main effect of HSD , but not sex \u00d7 HSD interaction . As shown in P < 0.05). Further studies have found that fibrotic changes caused by HSD in the heart of female mice were approximately twice that of male mice (1.75 vs. 2.74%)  . CollectQ < 0.05). Similarly, the HSD had an adverse effect on the expression of 35 metabolism-related genes with significant differences in female mice, causing a serious disorder (Q < 0.05) (P < 0.001) were identified, in six categories: (1) amino acid metabolism: valine, leucine, and isoleucine degradation (Q = 9.10E-07) and glycine, serine, and threonine metabolism (Q = 6.04E-04); (2) carbohydrate metabolism: pyruvate metabolism (Q = 4.56E-06), glycolysis/gluconeogenesis (Q = 2.74E-05), propanoate metabolism (Q = 2.25E-04), fructose and mannose metabolism (Q = 3.04E-04), and butanoate metabolism (Q = 8.03E-04); (3) energy metabolism: biosynthesis of unsaturated fatty acids (Q = 1.56E-05); (4) global and overview maps: oxidative phosphorylation (Q = 1.45E-15), fatty acid metabolism (Q = 1.70E-09), and carbon metabolism (Q = 3.65E-06); (5) lipid metabolism: biosynthesis of amino acids (Q = 5.97E-04) and glycerolipid metabolism (Q = 8.03E-04); (6) metabolism of cofactors and vitamins: porphyrin and chlorophyll metabolism (Q = 7.80E-06) and ubiquinone and other terpenoid-quinone biosynthesis (Q = 1.03E-05) (P < 0.05) were identified, in seven categories: (1) amino acid metabolism: arginine and proline metabolism (Q = 2.31E-02) and lysine degradation (Q = 2.83E-02); (2) biosynthesis of other secondary metabolites: neomycin, kanamycin and gentamicin biosynthesis (Q = 3.27E-02); (3) carbohydrate metabolism: glycolysis/gluconeogenesis ; (4) energy metabolism: sulfur metabolism (Q = 4.88E-02); (5) lipid metabolism: ether lipid metabolism (Q = 2.14E-02), glycerolipid metabolism (Q = 2.83E-02), steroid hormone biosynthesis (Q = 4.18E-02), arachidonic acid metabolism (Q = 4.18E-02), and glycerophospholipid metabolism (Q = 4.31E-02); (6) metabolism of terpenoids and polyketides: insect hormone biosynthesis (Q = 2.60E-02); (7) nucleotide metabolism: purine metabolism (Q = 1.84E-02)  . To furt.03E-05) . For fem.84E-02) . The STR.84E-02) , but not.84E-02) . AltogetP = 0.198), and macrophage infiltration was increased by HSD treatment in the hearts of male and female mice . The STRING database was used to visualize the PPANs of these transcripts to further study the function and relevance of these immune-related genes. As shown in Q < 0.001)  . Collectvia the cytochrome C release pathway (Q < 0.05). As shown in Q = 5.43E-16) and activation of cysteine-type endopeptidase activity involved in the apoptotic process (Q = 2.60E-03) of biological process in male and female mice were identified (P = 0.032). The HSD significantly increased the expression of cleaved caspase-3 in the hearts of both male and female mice. Collectively, these results demonstrated that excessive sodium intake can induce cardiac apoptosis.Studies have shown that a prenatal HSD can induce apoptosis of cardiomyocytes in the heart of offspring mice  pathway  and can  pathway . In thisentified . To furtThe meaningful finding of this study is that HSD can adversely affect the mouse heart by changing metabolism, immunity, fibrosis, and apoptosis and can induce mice to suffer from sex-specific cardiovascular disease. The conclusions of this study are mainly based on the following findings: (1) HSD has a sex-specific effect on the body mass of mice, especially on the mass of heart tissue; (2) the cardiac transcriptome is different in male and female mice; (3) the HSD-treated cardiac transcriptome showed sex-specific differences; (4) compared with the normal group, HSD produced a different number of DEGs and affects different KEGG pathways in male and female mice; (5) HSD affects the expression of genes related to fibrosis, metabolism, immunity, and apoptosis in heart tissue of male and female mice; (6) KEGG pathway enrichment analysis showed that HSD induced HCM in male mice, while HSD induced DCM in female mice. These data help us understand the negative effects of HSD on the heart in different sexes.db/db mice show higher levels of pro-inflammatory cytokines and more immune cell infiltration in the kidneys than female db/db mice after HSD treatment for 4 weeks  can be found in the article/The animal study was reviewed and approved by Henan Province People's Hospital Institutional Animal Care and Use Committee.SH, XC, and HW: conceptualization, funding, resources, and supervision. SH and XC: methodology, investigation, analysis, visualization, and writing\u2014review and editing. All authors contributed to the article and approved the submitted version.Research support was provided by the National Natural Science Foundation of China (Grant Number 82101089 to SH), Natural Science Foundation of Henan Province (Grant Number 222300420361 to XC), the Key R&D and Promotion Special Program of Henan Province (Grant Numbers 212102311011 and 222102310120 to SH and 212102310477 to HW), the Henan Provincial Medical Science and Technology Research Joint Co-construction Project (Grant Numbers LHGJ20200064 to SH and SB201901087 to HW), the Basic Science Project for Youth of Henan Eye Institute/Henan Eye Hospital (Grant Number 20JCQN003 to SH), and the Doctoral Research and Development Foundation of Henan Provincial People's Hospital (Grant Numbers ZC20190146 to SH and ZC20200230 to XC).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Mycobacterium smegmatis mc2155 as a host. The genome of this temperate siphovirus is 75,632 bp long , and BLASTn alignment revealed 99.86% identity with the genome of L3 mycobacteriophage Samty.Subcluster L3 bacteriophage Finnry was isolated from soil collected in Charleston, South Carolina, using  Mycobacterium smegmatis mc2155 using enrichment at 37\u00b0C followed by two purification/amplification cycles in 7H9 top agar, as described in the SEA-PHAGES Discovery Guide  Science Education Alliance-Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) program  studied ry Guide . Althougile tail .TCGATCAGCC) were identified using PAUSE (https://cpt.tamu.edu/computer-resources/pause).To extract genomic DNA from high-titer lysates, the Promega Wizard DNA cleanup system was used, and a DNA library was prepared with the NEBNext Ultra II DNA library prep kit. Pittsburgh Bacteriophage Institute sequenced Finnry on an Illumina MiSeq system (MiSeq reagent kit v3) , and 771https://phagesdb.org/DNAMaster). Programs utilized to identify putative genes included GLIMMER v3.02 (https://seaphages.org/forums/topic/5398). Default parameters were used for other software.Annotation was performed with the PECAAN  workflowER v3.02 , PhameraER v3.02 , GeneMarER v3.02 , StarterER v3.02 , ARAGORNER v3.02 , and tRNER v3.02 . FunctioER v3.02 , HHpred ER v3.02 , and theER v3.02  (parametFinnry\u2019s genome contains 130 predicted protein-coding genes (51 with assigned putative functions), 9 tRNAs, and no transfer-messenger RNAs. Potential gene duplications include tandem duplication of the WhiB family transcription factor sequences gp79/gp80 (BLASTp indicated 37.66% identity and 79% query coverage) and displaced duplication of gp121/gp131 (BLASTp indicated 42.59% identity and 93% query coverage).\u221250) between Finnry and related actinobacteriophages, Phamerator was used  \u201320. To c\u2013was used . Finnry\u2019was used , 2 phamsGCS scores  and wholThe GenBank and SRA accession numbers for Finnry are presented in"}
+{"text": "This article has been corrected: Due to errors in figure preparation, the AZ-treated (72 hr) images , is an accidental duplicate of the \u2018CTRL\u2019 image in row 3, panel 1 of 1470-1489. https://doi.org/10.18632/oncotarget.28011Original article: Oncotarget. 2021; 12:1470\u20131489."}
+{"text": "C-X-C Motif Chemokine Receptor 4 (CXCR4) in CD4+ T cells in LBD. CSF protein levels of the CXCR4 ligand, C-X-C Motif Chemokine Ligand 12 (CXCL12) were associated with neuroaxonal damage in LBD. Furthermore, we observed clonal expansion and upregulated Interleukin 17A expression by CD4+ T cells stimulated with a phosphorylated \u03b1-synuclein epitope. Thus, CXCR4-CXCL12 signaling may represent a mechanistic target for inhibiting pathological interleukin-17-producing T cell trafficking in LBD.Recent studies indicate that the adaptive immune system plays a role in Lewy body dementia (LBD). However, the mechanism regulating T cell brain homing in LBD is unknown. Here, we observed T cells adjacent to Lewy bodies and dopaminergic neurons in post-mortem LBD brains. Single-cell RNA sequencing of cerebrospinal fluid (CSF) identified upregulated expression of  The immune system is implicated in the neurodegenerative process of Lewy body dementia. PDD is defined by changes in memory and behavior and afflicts patients in late stage Parkinson\u2019s disease (PD) . The syme models \u201313 and ie models . Moreovee models , 5, 14. P = 8.6X10\u22125; P = 0.0031; P = 6.33X10\u221213) and DLB (P = 4.02X10\u221213) presented with lower cognitive scores than PD-NCI patients  and patients with clinical DLB and PD  . Montrea+ T cells in close proximity to neuronal processes labeled by the dopamine enzyme tyrosine hydroxylase (TH) in the substantia nigra of PDD and DLB brains + glutamatergic neurons in the hippocampal CA2 region , a kinase essential for cytokine signaling, and the T cell activation gene Cluster Of Differentiation 69 (CD69) (C-X-C Motif Chemokine Receptor 4 (CXCR4) was also highly upregulated in PD-DLB CD4+ T cells  , 16 of C subtype . Highly 9 (CD69) . The che T cells . Moreove T cells . Quantifn PD-DLB . Thus, e+ T cells in PD-DLB prompted us to determine whether clonally expanded (i.e. antigen-specific) cells were distinct in PD-DLB. To assess clonal expansion, we performed single-cell T cell receptor sequencing (scTCRseq) on the same CSF cells as above , a marker of pro-inflammatory IL-17-producing (Th17) memory CD4+ T cells  and PD-NCI (P = 7.67X10\u221210) subjects  = 31.697, P = 5.18X10\u221212); P = 1.00X10\u22124) and PD-NCI (P = 8.30X10\u22123) subjects  = 9.161, P = 0.0002); P = 0.071); s = 0.40; P = 0.023), and these correlations were lesser in healthy  and PD-NCI  subjects (ANCOVA (F (P = 0.031); We next sought to determine whether levels of CSF CXCL12 were associated with cognitive impairment in PD. We measured CXCL12 in a cohort of age- and sex-matched healthy (n=84) and PD (n=79) subjects . This re+ T cells of the peripheral immune system and CSF. We performed scRNAseq on peripheral blood mononuclear cells (PBMCs) of the same subjects we analyzed by CSF scRNAseq and focused our analysis on CD4+ T cells  and Actin Beta (ACTB)  , which ra (ACTB) . We also T cells .Interleukin 17A (IL17A) in cells stimulated with \u03b1-synuclein  from each subject (IL17A-expressing cells co-expressed CD4 and some clonotypes also expressed the Th17-associated cytokine gene Interleukin 22 (IL22; +IL-17A+ T cells in the PDD substantia nigra, which were adjacent to TH+IL-17A+ neurons (P = 0.007; IL17A RNA expression in the brain, yet the gene encoding the IL17A receptor, IL17RA, was highly expressed in the midbrain (Notably, we also detected higher expression of ynuclein . IL17A e17 cells . To detepatients . We then subject . IL17A-e22 IL22; . We conf neurons . We alsomidbrain , suggestmidbrain . Finallymidbrain .IL17A, a pro-inflammatory cytokine involved in autoimmune diseases (In conclusion, these results implicate Th17 cell involvement in the degeneration of neurons in LBD. Notably, CXCR4 regulates cell migration , and antdiseases . In animdiseases , 32. Mordiseases . Thus, odiseases .Supplementary MaterialData S3Data S2Data S1Data S4"}
+{"text": "A prior study demonstrated suboptimal antibiotic prescribing in the emergency department (ED) at 77.4% for uncomplicated lower respiratory tract infections (LRTI), urinary tract infections (UTI), and acute bacterial skin and skin structure infections (ABSSSI). This study measured the effect of indication-based antibiotic order sentences (AOS) on prescribing.IRB-approved quasi-experiment of adults prescribed antibiotics in ED for uncomplicated LRTI, UTI, or ABSSSI from January - June 2019 (pre-group) or September - December 2021 (post-group). Exclusion: hospital admission, immunocompromised, active cancer, or prophylactic antibiotics. AOS are lean process, electronic discharge prescriptions retrievable by name or indication; AOS implementation occurred July 2021. Optimal prescribing was defined as the correct antibiotic selection, dose, and duration per local and national guidelines. Seven-day endpoints: antibiotic escalation, ED or hospital readmission, any outpatient contact, and reported adverse drug event (ADE). Descriptive and bivariate statistics performed. Variables considered for multivariable logistic regression had p< 0.2 or plausible association with optimal prescribing.294 patients included: 147-pre and 147-post. Patient characteristics are in Table\u00a01. Overall optimal prescribing improved from 12 (8.2%) to 34 (23.1%) (p< 0.001). Breakdown of optimal prescribing in pre- and post-groups: selection 90 (61.2%) vs 117 (79.6%) (p< 0.001), dose 99 (67.3%) vs 115 (78.2%) (p=0.036), duration 38 (25.9%) vs 50 (34%) (p=0.126). After adjustment, AOS were independently associated with optimal prescribing (Table\u00a02). Secondary endpoints: antibiotic escalation 10 (6.8%) vs 7 (4.8%) (p=0.662), hospital or ED readmission 12 (8.2%) vs 10 (6.8%) (p=0.658), outpatient contact 31 (21.1%) vs 28 (19%) (p=0.662), and ADE 2 (1.4%) vs 4 (2.7%) (p=0.684). Post-hoc analysis showed suboptimal uptake of AOS by ED prescribers.AOS is an efficient and promising antimicrobial stewardship strategy; provider re-education is needed to increase AOS uptake.All Authors: No reported disclosures."}
+{"text": "Nirmatrelvir coadministered with ritonavir (nirmatrelvir/r) is a COVID-19 treatment. This study evaluated nirmatrelvir/r in nonhospitalized, symptomatic adults with COVID-19 at high risk of progressing to severe disease. We report secondary efficacy endpoints associated with COVID-19\u2500related medical visits, including hospitalization details and oxygen support, as of the primary completion data cutoff .In this phase 2/3 double-blind, interventional study, adults with confirmed SARS-CoV-2 and symptom onset \u2264 5 days (d) were randomized 1:1 to receive nirmatrelvir/r 300 mg/100 mg or placebo (PBO) orally every 12 hours for 5 d. COVID-19\u2500related medical visits were collected through Day 28. Oxygen support for COVID-19 and details of COVID-19\u2500related hospitalization, including duration, intensive care unit (ICU) status, and mechanical ventilation, were assessed.Table\u00a01). In addition to fewer hospitalizations being reported with nirmatrelvir/r (n=8 [0.8%]) vs PBO (n=65 [6.2%]), pts receiving nirmatrelvir/r had fewer hospitalized d (Table\u00a02), with mean durations of 9.6  d with nirmatrelvir/r and 11.2  d with PBO in hospitalized pts. No pts in the nirmatrelvir/r group and 9 pts (0.9%) in PBO group were admitted to the ICU. No pts in the nirmatrelvir/r group received mechanical ventilation vs 3 pts in the PBO group. Fewer other COVID-19\u2500related nonhospital medical visits were reported with nirmatrelvir/r vs PBO (Table\u00a03). In the full analysis set, fewer pts required oxygen therapy for COVID-19 with nirmatrelvir/r (n=9/1120 [0.8%]) vs PBO (n=54/1126 [4.8%]).Of the 2246 patients (pts) enrolled globally from Jul to Dec2021, 2085  started treatment and met criteria for the modified intent-to-treat population . Fewer overall COVID-19-related medical visits were reported with nirmatrelvir/r vs PBO  vs pts receiving PBO. Clinical Trial: NCT04960202.Jennifer Hammond, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Heidi Leister-Tebbe, BSN, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Annie Gardner, MPH, MSPT, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Paula Abreu, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Weihang Bao, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Wayne Wisemandle, MA, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Wajeeha Ansari, MPH, Pfizer Inc.: Stocks/Bonds Rienk Pypstra, MD, MBA, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds James M Rusnak, MD, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds."}
+{"text": "Correction: BMC Musculoskelet Disord 24, 49 (2023)10.1186/s12891-023-06160-zhttp://ovidsp.ovid.com/ovidweb.cgi?T=JS&PAGE=reference&D=emed8&NEWS=N&AN=2009254927)\u201d instead of Chan et al. 2013.Following publication of the original article , the autFurthermore, the word \"Specific\" in Secondary outcomes is without the \"S\" in Table 1.The original article  has been"}
+{"text": "Urinary tract infections (UTI) are the second most prevalent microbial infection, impacting 150 million people globally every year. Traditionally, urine culture is considered the gold standard for UTI pathogen detection. However, molecular techniques, such as real time multiplex PCR tests targeting multiple pathogens, are becoming the alternative diagnostic tools for rapid detection of pathogens with the potential to provide quick diagnosis and enable targeted treatment. Due to the serious economic and healthcare utilization burden UTIs pose, early pathogen detection with a rapid turn-around-time to results has the potential to be instrumental for improving patient care and outcomes.A total of 300 deidentified patient samples that were previously tested via urine culture  were subjected to real time PCR molecular testing employing the nanofluidic Open Array \u00ae platform . Statistical analyses were performed using R version 3.6.0.Among 300 urine specimens studied, culture detected pathogens in 183 patient samples (61%), and 117 samples were deemed negative. Culture and PCR results demonstrated an overall agreement of 75.3% (n=226) with 59% positive (n=177) and 16.3% negative (n=49). Results for 24.7% samples (n=74) were discordant. A total of 2% (n=6) were culture positive and PCR negative, and 22.7% (n=68) were culture negative and PCR positive. Agreement between PCR and culture positive results was 0.97, 95%CI  (177/183). Among the PCR positive samples (n=245), 32.7% were poly-microbial (n=80) and 67.3% were mono-microbial (n=165). In comparison, among culture positive samples (n=183), 33.3% (n=61) were polymicrobial and 65.6% (n=120) monomicrobial.Our study demonstrates that multiplex real time PCR-based detection of UTI bacterial pathogens has positive agreement with the traditional urine culture method. PCR based testing has the potential to deliver quick pathogen identification to enable targeted treatment options and medication adjustments. Further studies will correlate semi-quantitative cfu/mL culture values with semi-quantitative copies/mL PCR values to further refine result reporting and test performance.Pallavi Upadhyay, PhD, HealthTrackRx: Stocks/Bonds Fahida Surar, B.S., HealthTrackRx: Salaried Employee Geun Kim, M.S., HealthTrackRx: Salaried Employee Jay Reddy, PhD, HealthTrackRx: Stocks/Bonds Barbara D. Alexander, MD, Astellas: Advisor/Consultant|HealthtrackRx: Advisor/Consultant|HealthtrackRx: Grant/Research Support|Scynexis: Grant/Research Support|UpToDate: Advisor/Consultant Kimberly Hanson, MD, MHS, FIDSA, HealthTrackRx: Advisor/Consultant|HealthTrackRx: Clinical Advisory Board Member Vijay Singh, PhD, HealthTrackRx: Stocks/Bonds."}
+{"text": "Enterobacterales order were detected in fecal samples among 580 patients during the period of 2017\u20132019. ESBL/carbapenemase/plasmidic AmpC producer rates were 28.8%, 2.4%, and 1.2%, respectively. A wide variety of ESBLs: CTX-M-15 (41%), CTX-M-3 (24%), CTX-M-27 (11%), and CTX-M-14 (4%) was found. The carbapenemases identified in this study were New Delhi metalo-\u03b2-lactamase (NDM)-1 (5.4%) and Klebsiella carbapenemase (KPC)-2 (1.5%). Most NDM-1 isolates also produced CTX-M-15/-3 and CMY-4 \u03b2-lactamases. They belonged to ST11 Klebsiella pneumoniae clone. The epidemiology typing revealed three main high-risk K. pneumoniae clones (26%)\u2014ST11, ST258, and ST15 and five main Escherichia coli clones\u2014ST131 (41.7%), ST38, ST95, ST405, and ST69. Sixty-one percent of ST131 isolates were from the highly virulent epidemic clone O25b:H4-ST131. Phylotyping revealed that 69% of E. coli isolates belonged to the virulent B2 and D groups. Almost all (15/16) Enterobacter isolates were identified as E. hormaechei and the most common ST type was ST90. Among all of the isolates, a high ESBL/carbapenemases/plasmid AmpC (32.4%) prevalence was observed. A significant proportion of the isolates (37%) were members of high-risk clones including two pan-drug-resistant K. pneumoniae ST11 NDM-1 producing isolates. Due to extensive antibiotic usage during COVID-19, the situation may worsen, so routine screenings and strict infection control measures should be widely implemented.The gastrointestinal tract is an important reservoir of high-risk Enterobacteria clones and a driver of antimicrobial resistance in hospitals. In this study, patients from six hospitals in four major Bulgarian towns were included in this study. Overall, 205 cefotaxime-resistant isolates (35.3%) of  Klebsiella pneumoniae and hospital-acquired infections [Enterobacterales species [K. pneumoniae, Escherichia coli, and Enterobacter cloacae complex have been globally reported [\u03b2-lactams are commonly used antimicrobials due to their safe profile and broad-spectrum activity ,2,3,4,5.fections . In the  species ,4,7. The species . These b species ,4,5,6. Ireported ,4,6. In reported . In addireported .The gastrointestinal tract can be an important reservoir of ESBL-/carbapenemase-producing enterobacteria in hospital settings  leading Fecal carriage of ESBL/carbapenemase producers has been widely reported all over the world ,10. DiffThe aim of this study was to evaluate the prevalence of \u03b2-lactamase production and clonal relatedness of fecal ESBL- and carbapenemase-producing enterobacteria, collected from hospitalized patients .TMKPC . Bacterial isolates were identified using routine biochemical identification and were confirmed by VITEK  or Phoenix . Hsp60 sequencing [Enterobacter spp. and Klebsiella oxytoca isolates.The study was conducted in six hospitals\u2014University Multiprofile Hospital for active treatment (UMHAT), Varna; UMHAT, Plovdiv; UMHAT, Pleven, and three hospitals in Sofia during the period of December 2017\u2013June 2019. The fecal samples were obtained from patients  during the routine diagnostic process and were additionally inoculated on selective MacConkey agar with 1 mg/L cefotaxime and on Chromagarquencing  was usedhttp://www.eucast.org/clinical_breakpoints/), last accessed on 10 August 2022. The following antibiotics were tested: amoxicillin/clavulanic acid 30 \u00b5g (AMC), ceftazidime 10 \u00b5g (CAZ), cefotaxime 5 \u00b5g (CTX), cefepime 30 \u00b5g (FEP), cefoxitin 30 \u00b5g (FOX), imipenem 10 \u00b5g (IMP), meropenem 10 \u00b5g (MEM), piperacillin/tazobactam 36 \u00b5g (PIP/TAZ), chloramphenicol 30 \u00b5g (CHL), gentamicin 10 \u00b5g (GEN), amikacin 30 \u00b5g (AMK), tobramycin 10 \u00b5g (TOB), trimethoprim/sulfamethoxazole 25 \u00b5g (SXT), ciprofloxacin 5 \u00b5g (CIP), levofloxacin 5 \u00b5g (LVX), fosfomycin 200 \u00b5g (FOS)(for E. coli isolates), and tigecycline 15 \u00b5g (TIG for E. coli isolates). Susceptibility to tigecycline in Klebsiella isolates was determined by the broth microdilution method . Susceptibility to colistin was initially tested with modified SuperPolymyxin medium [K. pneumoniae and E. coli that grew on this screening medium were tested with the broth microdilution method .Antimicrobial susceptibility testing was performed using the disk diffusion method on M\u00fcller-Hinton II agar and the microdilution broth method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines (Version 10). The susceptibility testing was carried out according to the clinical breakpoints of the EUCAST (Version 10) , a phenotypic confirmation of carbapenemase production was performed by the KPC, MBL, and OXA-48 Kit .Presumptive ESBL production was detected with the double-disk synergy method . PotentiE. coli K12:W3110 RifR lac/-/(1.2 \u00d7 107 CFU/mL). After overnight incubation at 37 \u00b0C, growth of the indicator strain on the gel localized the \u03b2-lactamase band by which the \u03b2-lactam had been inactivated.Production and number of \u03b2-lactamases were detected and analyzed by analytical isoelectric focusing (IEF) according to the method of Mathew  with modblaVIM, blaIMP, blaKPC, blaNDM, blaOXA-48), AmpC , and ESBLs  as previously described [blaSHV, blaCTX-M-1-group, blaCTX-M-9-group, blaCMY,blaDHA,blaKPC, and blaNDM. [All isolates were screened for the presence of carbapenemase-encoding genes , last accessed on 30 July 2022. A clone was defined as isolates showing 80% similarity.Clonal relatedness was investigated by ERIC PCR and Multilocus Sequence Typing (MLST). For Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR, ERIC-1 and -2a primers were used as described previously . GeneticK. pneumoniae MLST typing and Achtman scheme was applied for E. coli. For E. coli isolates, the assignment to allelic numbers and sequence types (STs) was performed according to the MLST database (https://bigsdb.web.pasteur.fr/ecoli/ecoli.html), last accessed on 30 July 2022. Detection of specific O25b-ST131 clone was performed with allele-specific PCR for pabB gene as previously described [Pasteur scheme was used for K. pneumoniae isolates, protocols and assignment to allelic numbers and sequence-types (STs) were carried out as described in the MLST database , last accessed on 30 July 2022. A clonal complex was defined as a group of two or more independent isolates that shared six identical alleles.For E. cloacae complex isolates, primers, protocols, and assignment to allelic numbers and sequence-types (STs) were carried out as described in the MLST database (https://pubmlst.org/organisms/enterobacter-cloacae), last accessed on 30 July 2022.For E. coli isolates as previously described [Phylotyping was applied for escribed .E. coli (n = 103); K. pneumoniae (n = 65); Klebsiella oxytoca (n = 8); E. cloacae complex (n = 16); Morganella morganii (n = 3); and Citrobacter freundii complex (n = 10).A total of 205 enterobacterial isolates resistant to cefotaxime were isolated from fecal samples, collected from 580 patients (35.3%) as follows: 61 cefotaxime-resistant isolates from 158 studied patients (38.6%) from University Hospital (UH)-Varna; 25 cefotaxime-resistant isolates from 71 patients (35.2%) from UH-Pleven; 58 cefotaxime-resistant isolates from 102 patients (56.9%) from UH-Plovdiv, and 61 cefotaxime-resistant isolates from 249 patients (24.5%) from three hospitals in Sofia. The isolates were identified as Hsp60 sequencing identified 15 E. cloacae complex isolates as Enterobacter hormaechei, subdividing them into three subspecies\u2014E. hormaechei spp. hoffmannii (n = 3); E. hormaechei spp. steigerwaltii (n = 10); and E. hormaechei spp. xiangfangensis (n = 2). One isolate was identified as Enterobacter kobei. Two isolates, initially identified as K. oxytoca, were reidentified by hsp60 sequencing as Klebsiella michiganensis.Klebsiella isolates to tigecycline was 93%. For E. coli, 17% and 3% resistance to tigecycline and fosfomycin, respectively, were found. Only five isolates of Klebsiella spp. (2.4%) were colistin nonsusceptible. The results are shown in K. pneumoniae isolates were determined as pandrug-resistant according to the criteria of Magiorakos et al. [The nonsusceptibility (resistance or intermediate susceptibility) rates in the collection of isolates were as follows: amoxicillin/clavulanic acid 93%, ceftazidime 86%, cefotaxime 100%, cefoxitin 35%, cefepime 95%, piperacillin/tazobactam 72%, imipenem 7%, meropenem 7%, tobramycin 68%, gentamicin 56%, amikacin 50%, trimethoprim/sulfamethoxazole 52%, ciprofloxacin 68%, levofloxacin 63%, and chloramphenicol 22%. The nonsusceptibility rates in the s et al. .E. cloacae complex and three C. freundii complex isolates were determined as possible inducible AmpC hyperproducers, demonstrating antagonism between amoxicillin/clavulanic acid and cefotaxime or ceftazidime.Disk diffusion synergy method (DDST) confirmed 171 isolates  as ESBL producers. Four K. pneumoniae isolates, suggesting class B carbapenemase production (zone around the disk with EDTA is \u22655 mm). Three isolates (two K. pneumoniae and one E. coli) demonstrated increased zones of inhibition by the disk containing meropenem and phenylboronic acid, suggesting class A (KPC) carbapenemase activity.The phenotypic test with meropenem and meropenem/EDTA disks was positive in 11 carbapenem-resistant blaSHV-1 gene was detected (which suggests possible SHV-1 hyperproduction). Three cefotaxime-resistant isolates, susceptible to carbapenems and with positive DDST did not produce a positive PCR reaction with any of the used ESBL group-specific primers.PCR and sequencing revealed the presence of genes, encoding ESBL in 167 (28.8%) isolates, obtained from 580 patients. In one isolate, only p < 0.0001). The ESBL producers among the isolates of E. coli and K. pneumoniae were 56.2% (94/167) and 34.1% (57/167), respectively.The prevalence of ESBL producers in Sofia hospitals was significantly lower  than in the other locations, 35.6% (118 ESBL producers/331 patients) (blaCTX-M-15 in 41% (84/205) of the isolates, blaCTX-M-3 in 24% (49/205), blaCTX-M-27 in 11% (22/205), and 8 isolates (3.9%) displayed blaCTX-M-14. Only one isolate was positive for blaCTX-M-9 and one for blaSHV-12 , except one isolate that showed blaCTX-M-14, were produced by E. coli strains. In contrast, blaCTX-M-3 was the prevailing ESBL among K. pneumoniae and K. michiganensis isolates. One K. pneumoniae isolate showed mixed sequences , GGT is a codon for glycine (specific for blaCTX-M-15)). All K. pneumoniae isolates were blaSHV positive. Of them, 29 representative isolates were sequenced, and blaSHV-1 (n = 21) and blaSHV-11 (n = 8) were identified. In 71 isolates, blaTEM was found and, later, 10 of them were confirmed by sequencing as blaTEM-1.Among the 205 investigated cefotaxime-resistant isolates, we observed solely laSHV-12 . blaCTX-K. pneumoniae, were carbapenemase producers. Thirteen of them coproduced ESBL or/and plasmid AmpC (K. pneumoniae isolates had blaNDM-1 together with blaCMY-4 and blaCTX-M-3/-15. Two K. pneumoniae and one E. coli isolates had blaKPC-2.Fourteen isolates (2.4%) from 580 patients, mainly mid AmpC . Ten K. E. coli  isolates showed two types of plasmid AmpC, blaDHA-1, n = 5 or blaCMY-2, n = 2. Thirteen isolates (2.2%) were AmpC hyperproducers (inducible or constitutive) (Seven itutive) .Thus, 188 isolates (32.4%) were found to carry ESBL (28.8%) or carbapenemases (2.4%) or plasmid AmpC genes (1.2%).Klebsiella spp. isolates, 24 representative isolates were subjected to IEF   exhibited two bands\u2014pI8.4 and pI8.8 (The isoelectric focusing was carried out in representative isolates according to the species and detected \u03b2-lactamase genes. Of 73 solates) . We confnd pI8.8   and did not show cefotaxime hydrolyzing bands corresponding to TEM and SHV ESBL \u03b2-lactamases. Three isolates with unidentified enzyme type gave bands with pI8.0 (two isolates) and pI9.2 (one isolate). They were without CTX hydrolytic activity.E. hormaechi, C. freundii complex and M. morganii isolates, 5 isolates were tested, and a single cefotaxime hydrolyzing band was detected, pIs corresponded with sequenced enzymes , ST11 , and ST37 being the dominant types. In addition, ST1198, ST280, ST34, ST15, ST258, ST17, ST253, and ST449 were also found. Six isolates demonstrated unique profiles . K. oxytoca and K. michiganensis isolates had unique ERIC profiles.Epidemiological typing revealed 18 ERIC clusters  among 65profiles . The isohospital . ST11 isE. coli isolates, 40 ERIC types were identified , ST38 , ST155 , ST405 , ST1196, and ST88 .Among entified . A totalE. coli (n = 43) was detected in five centers and was associated with the production of CTX-M-27 , CTX-M-15 , CTX-M-3 , CTX-M-14 , and KPC-2 carbapenemase, found in a single isolate ; D, n = 20 ; A, n = 12; and B1, n = 20. The association between MLST types and phylotypes is shown in The following E. cloacae isolates (n = 16) demonstrated 12 ERIC clusters and 6 ST types. The most common ST was ST90, associated with CTX-M-15 production (oduction .n = 580) was similar to that in Portuguese hospitals (24%) [E. coli and K. pneumoniae isolates  in Bulgaria in 2019, which was the highest in Europe and showed a stable trend during the last 5 years , last accessed on 10 August 2022. We also found increased nonsusceptibility rates in the tested ESBL fecal isolates to aminoglycosides (50%-68%) and quinolones (63\u201368%) in comparison to the rates found in 2015 (21%-57% for aminoglycosides and 15\u201327% for quinolones) [This study reveals a moderate rate of fecal colonization with ESBL producers 28.8%) among Bulgarian patients during the period 2017\u20132019. The rate of third-generation cephalosporin-resistant isolates (mostly due to production of ESBL/carbapenemase/plasmid AmpC) was high (35.3%), but lower than the rate observed in a pilot study on hospital fecal carriage in Varna city, Bulgaria in 2015 (42.5%) . A possils (24%)  and highls (24%) . The obs.8% amongnolones) . A similnolones) .E. coli (56.2% versus 69%) and an increased proportion of ESBL K. pneumoniae isolates (34.1% versus 20.4%) were found [E. coli have been reported in many studies [In comparison with our previous study in 2015, a decreased rate of ESBL-producing re found . Similarre found . The pre studies ,26,27,28E. coli, as well as in all other species except Klebsiella spp. CTX-M-15-producing isolates have been reported worldwide [Klebsiella isolates. Similar to CTX-M-15, CTX-M-3 was also widely distributed among enteric strains [E. coli isolates. CTX-M-9 and CTX-M-14 intestinal producers have been reported in Portugal, Spain, China, and Bulgaria [E. coli isolates. In Bulgaria, DHA-1 enzyme was identified for the first time in 2019 in an E. cloacae complex isolate obtained from a blood sample [The main \u03b2-lactamase type in our study was CTX-M-15 41%). It was the predominant ESBL in 1%. It waorldwide ,8,9. CTX strains ,26,27,28Bulgaria ,26. CTX-d sample .K. pneumoniae isolates and in a single E. coli isolate. All carbapenem-producing isolates were obtained from hospitalized patients in Sofia. Interestingly, the frequency of ESBL producers (19.7%) was not high among these patients. The coproduction of NDM-1, CTX-M-15/-3, and CMY-4 enzymes in nine K. pneumoniae isolates is a possible explanation for the high resistance rates, identified in the NDM-1 producing isolates, two of them being pandrug-resistant.An important finding in the current study is the detection of carbapenemase-producing isolates. Although the detection rate was low (2.4%), it is an indicator that the enteric tract may act as a reservoir for these problematic bacteria. The carbapenem resistance in the present collection of fecal isolates was associated with NDM-1 and KPC-2, detected in The rates of carbapenemase producers in Europe have steadily increased during the last years in both clinical and intestinal isolates ,33,34,35n = 17, 26%) among K. pneumoniae fecal isolates. Taking the second place, ST11 was represented in 17% of K. pneumoniae isolates, all obtained from patients in Sofia hospitals. Most NDM-1 carbapenemase-producers belonged to this clone, including two panresistant and the colistin-resistant isolates. This result is in concordance with our previous study from 2019 that detected a high level of ST11 blaNDM-1/blaCTX-M-15 or-3/blaCMY-4 positive isolates in the same hospitals [blaNDM-1/blaCTX-M-15/3/blaCMY-4 positive isolates have also been detected in the Czech Republic [Another important finding in the present study is the identification of the high-risk clones ST11, ST258, and ST15 , is another high-risk clone. It is also distributed worldwide and associated with KPC-2 production [ST258 oduction ,30,33,41oduction . ST11 anoduction ,40,42,43K. pneumoniae clone which has commonly been associated with the production of ESBLs, mainly CTX-M-15 [ST15 is another high-risk CTX-M-15 ,7,44, buCTX-M-15 . In the CTX-M-15 .K. pneumoniae clones found in this study were ST37 and ST17. These clones showed a sustained persistence in Bulgaria. These ST have been already identified in clinical and fecal isolates, associated with CTX-M-15 and KPC-2 production [Other intestinal multidrug-resistant oduction ,45. ST37oduction .In the present study, ST353 was presented by a high number of isolates (19%), mainly obtained from patients hospitalized in Plovdiv University Hospital(A). ST353 isolates were associated mainly with CTX-M-3 production. This ST type has been rarely reported. In studies from China and Colombia, authors reported ST353 isolates, producing KPC and OXA-48 ,47. ST11E. coli isolates with ST131 being the predominant, found in 41.7%. The second most common ST was ST38 (10%). ST405, ST95, and ST69 were presented by single isolates only. The isolates from these high-risk clones represented 57% (n = 59) of all the studied E. coli isolates. All of them belonged to the B2 or D phylogroups. This is an important finding as these phylogroups are associated with a prolonged fecal carriage [Five high-risk clones were observed among carriage . This win = 43) were representatives of phylogroup B2 and 60% (n = 26) belonged to the O25b:H4-ST131 clone. Our study showed that the O25b:H4-ST131 clone was the major clone among fecal E. coli isolates, harboring different ESBL genes, mainly blaCTX-M-15, but also blaCTX-M-3 and blaCTX-M-14, which is in concordance with other reports [E. coli isolates were detected in all centers except the Sofia hospital 1 (D), where NDM-1-producing ST11 K. pneumoniae was observed. In addition, the frequency of ST131 isolates in the present study was higher (42%) than the rate (35%) detected in 2015 [Almost all ST131 isolates  and the plasmid AmpC enzyme DHA-1. This type has also been reported as a high-risk clone, causing both nosocomial and community-acquired infections, mainly urinary tract infections [In our study, fections . The detfections .E. coli. Recently, it has been found that representatives of ST69 clone carry an intact locus of enterocytes effacement (LEE), coding second bacterial type III secretion systems involved in the pathogenesis of Gram-negative infections [E. coli were identified as ST405 (D phylogroup). This clone was previously detected in Bulgaria as a carrier of NDM-1 carbapenemases [E. coli ST405 is an emerging urosepsis pathogen, reported to carry blaCTX-M, blaNDM, and a number of virulent genes comparable with O25b:H4-ST131 [ST69 is an interesting lineage from the D phylogroup, identified in two isolates of fections . In our enemases . E. coliH4-ST131 .E. coli isolates belonging to B1 and A phylogroups, which have been reported as commensal gut bacteria [In addition to the isolates that represent highly virulent clones, we also detected bacteria ; the obsEnterobacter isolates, ST90 producing CTX-M-15 was detected as the dominant clone. Although no high-risk clones have been defined in Enterobacter spp. so far, ST90 isolates, producing different carbapenemases, were detected in many countries: VIM-1 in Greece, IMP-4 in Canada and UK, and NDM-1 in Romania [E. cloacae complex can be a candidate for an international high-risk clone.In the group of  Romania . Polish  Romania . So, we The fecal colonization with ESBL and/or carbapenemase producers from high-risk international clones, associated with significant virulence and invasive potential and multidrug or pandrug resistance, can be an important reservoir not only for difficult to treat nosocomial infections, but also for wide dissemination in the community. Given that this study was performed before the COVID-19 pandemic, we can assume that the increased antibiotic usage during the last three years has further worsened the situation.E. coli and 17 K. pneumoniae) were members of high risk clones (37%).Twenty six percent of the K. pneumoniae isolates were representatives of high-risk clones such as ST11, ST258 and ST15 K. pneumoniae isolates. A very worrying finding was the detection of two ST11 pandrug resistant isolates, coproducing NDM-1, CMY-4, and CTX-M-15. Among E. coli, five high risk clones (57% of E. coli isolates) were found . The investigated isolates were producers of a wide range of \u03b2-lactamases\u2014CTX-M-15, CTX-M-3, CTX-M-27 (reported for the first time in Bulgaria), CTX-M-14, NDM-1, KPC-2, and plasmid AmpC DHA-1 and CMY-2 enzymes. The detected high frequency of ESBL/carbapenemases-producing enteric bacteria before COVID-19 and the dramatically increased selective pressure during the pandemic period will negatively impact the antimicrobial resistance in clinically significant bacterial species. Further studies should closely monitor the future trends. The routine screening for colonization with MDR bacteria at hospital admission and during the hospital stay, especially in high-risk departments, as well as strict infection control measures should be widely implemented in Bulgarian hospitals to limit the further dissemination of problematic multidrug-resistant bacteria.In conclusion, a high rate of fecal colonization (32.3%) with ESBL/carbapenemase/plasmid AmpC producers among patients in Bulgarian hospitals was found. A high rate of ESBL producers (28.8%) was detected with a relatively low rate of carbapenemase producers (2.4%). Seventy six isolates (59"}
+{"text": "Bacteria and archaea are central to the production, consumption, and remineralization of dissolved and particulate organic matter and contribute critically to carbon delivery, nutrient availability, and energy transformations in the deep ocean. To explore environmentally relevant genomic traits of sinking-particle-associated versus free-living microbes, we compared habitat-specific metagenome-assembled genomes recovered throughout the water column in the North Pacific Subtropical Gyre. The genomic traits of sinking-particle-associated versus free-living prokaryotes were compositionally, functionally, and phylogenetically distinct. Substrate-specific transporters and extracellular peptidases and carbohydrate-active enzymes were more enriched and diverse in particle-associated microbes at all depths than in free-living counterparts. These data indicate specific roles for particle-attached microbes in particle substrate hydrolysis, uptake, and remineralization. Shallow-water particle-associated microbes had elevated genomic GC content and proteome nitrogen content and reduced proteome carbon content in comparison to abyssal particle-associated microbes. An inverse trend was observed for their sympatric free-living counterparts. These different properties of attached microbes are postulated to arise in part due to elevated organic and inorganic nitrogen availability inside sinking particles. Particle-attached microbes also were enriched in genes for environmental sensing via two-component regulatory systems, and cell-cell interactions via extracellular secretion systems, reflecting their surface-adapted lifestyles. Finally, particle-attached bacteria had greater predicted maximal growth efficiencies than free-living bacterioplankton at all depths. All of these particle-associated specific genomic and proteomic features appear to be driven by microhabitat-specific elevated nutrient and energy availability as well as surface-associated competitive and synergistic ecological interactions. Although some of these characteristics have been previously postulated or observed individually, we report them together here in aggregate via direct comparisons of cooccurring free-living and sinking-particle-attached microbial genomes from the open ocean. Thes10\u201316\u2013in situ by divers  and FL microbes found throughout the water column in the oligotrophic Pacific Ocean, we leveraged time series metagenomic samples recovered at Station ALOHA in the North Pacific Subtropical Gyre, generating a total of 407 mid- to high-quality MAGs from both sample types. FL microbes (captured on 0.2-\u03bcm-pore-size filters) were collected from surface waters to a depth of 4,000 m at Station ALOHA, and metagenomes were generated, assembled, and analyzed \u201345. In p\u2013To expand the genomic data from sympatric SPA and FL prokaryote communities throughout the water column, metagenomic data sets were prepared from time series samples collected from the surface to 4,000 m during Station ALOHA time series efforts and expeditions \u201347. Thes\u201343\u201310.1128/mbio.01569-22.6TABLE\u00a0S1Table\u00a0S1, XLSX file, 0.03 MB.Sample descriptions and metadata of metagenomes. Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the The FL and PA MAGs were classified into five sample types . FL MAGs10.1128/mbio.01569-22.7TABLE\u00a0S2Table\u00a0S2, XLSX file, 0.04 MB.Checkm statistics, taxonomic classification, and sample designation of the MAGs. Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the Proteobacteria, including Alphaproteobacteria, Gammaproteobacteria, Planctomycetota, Chloroflexota, Bacteroidota, and Verrucomicrobiota. Ten other bacterial phyla were each represented by only a single MAG. This included the poorly characterized phylum UBP17, formerly known as Cloacimonetes (WWE1), of which only a few MAGs have been included in the genome taxonomy database to date  FL and PA MAGs shared some taxa at higher classification levels, only nine genera were found in both the FL and PA data sets. These included MAGs closely related to Alcanivorax, Prochlorococcus_A, Thalassarchaeum, Henriciella, Idiomarina, Nitrosopumilus, Nitrosopelagicus, Roseibacillus_B, and Bythopirellula  identifiers (IDs) and clustered into 49,146 orthologous protein families. Nonmetric multidimensional scaling (NMDS) profiles based on the presence of KO IDs  and orth10.1128/mbio.01569-22.1FIG\u00a0S1FIG\u00a0S1, PDF file, 1.1 MB.Nonmetric multidimensional scaling (NMDS) plots of the recovered MAGs based on gene annotations. (A) NMDS plot generated based on the presence/absence of KEGG Orthology (KO) annotations. (B) NMDS plot generated based on the presence/absence of orthologous protein families. Shapes indicate the sample type designation of the MAG. Color indicates the taxonomic classification of the MAG. PA and FL designations are as described in the legend to Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the P value\u2009<\u20090.05)  , and S4.\u2009<\u20090.05) . For FL 10.1128/mbio.01569-22.2FIG\u00a0S2q value of <0.05 for Fisher\u2019s exact test. Download FIG\u00a0S2, PDF file, 0.5 MB.Venn diagram of significantly enriched genes. (A and B) Venn diagrams of significantly enriched genes based on KEGG Orthology (KO) annotations and orthologous protein family. Significantly enriched KOs and orthologous gene families were determined using Scoary with a Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 10.1128/mbio.01569-22.8TABLE\u00a0S3Table\u00a0S3, XLSX file, 0.4 MB.Scoary enrichment of KO IDs (A) and orthologous protein families (B) based on sample types. Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 10.1128/mbio.01569-22.9TABLE\u00a0S4Table\u00a0S4, XLSX file, 0.1 MB.Count tables of transporter genes (A), peptidase genes (B), CAZyme genes (C), and extracellular secretion system genes (D) included in MAGs. Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the The most enriched KO IDs identified by Scoary were significantly associated with the PA_Shallow MAGs A and B. Previous studies have shown that transporters of carbohydrates and energy sources are ubiquitous in heterotrophic prokaryotic communities throughout the open water column, from the surface to abyssal depths, and are crucial for organic matter transformation . To inve10.1128/mbio.01569-22.3FIG\u00a0S3FIG\u00a0S3, PDF file, 0.1 MB.Counts of MAGs encoding substrate-specific transporters. Genes encoding putative transporters were grouped based on their substrate specificity. Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 10.1128/mbio.01569-22.4FIG\u00a0S4FIG\u00a0S4, PDF file, 0.1 MB.Counts of MAGs encoding substrate-specific CAZymes. Genes encoding putative CAZymes were annotated against dbCAN (3.0) with default parameters  and filtCopyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the A greater proportion of MAGs encoding mannopine transporters were identified in the FL samples than in the PA samples (~6.2% versus ~1.2%) . The subGiven the higher prevalence and diversity of transporters in SPA MAGs, POC solubilization capabilities of both SPA and FL MAGs were investigated. Extracellular hydrolytic enzymes that include peptidases and carbohydrate-active enzyme (CAZymes), key components in particulate organic carbon hydrolysis, were examined in terms of their abundance, diversity, and function B and C.The SPA MAGs possessed a significantly higher count of genes encoding peptidases and CAZymes per genome than the FL MAGs  and B. ASince polysaccharides constitute a large fraction of DOC  and POC 59\u2013l-rhamnohydrolase (GH106), \u03b1-l-rhamnosidase (GH106), alginate lyase (PL6 and -14), endo-\u03b1-1,5-l-arabinanase (GH43), \u03b2-l-arabinofuranosidase (GH146), chitinase (GH18), and \u03b2-porphyranase (GH16). These CAZymes hydrolyze polysaccharides such as alginate, porphyran, ulvan, and xylan, which are structural components of algae, and chitin, a highly abundant polysaccharide in marine environments. These CAZymes also hydrolyze an assortment of polysaccharides, releasing sugars such as rhamnose, xylan, and arabinose, which are found enriched in spring phytoplankton blooms , was found to be enriched in FL MAGs in comparison to PA MAGs . This en63\u2013n blooms .l-fucosidase (GH29 and GH151), \u03b2-glucosidase , cellulase , cellobionic acid phosphorylase (GH94), \u03b1-N-acetylgalactosaminidase (GH109 and -114), \u03b2-galactosidase (GH2 and -42), and \u03b1-galactosidase . The enrichment of these CAZymes in SPA MAGs also correlates well with the spike in multiple sugars detected during summer phytoplankton blooms, such as mannan, fucose, glucose, galactosamine, and galactose , \u03b2-mannosidase (GH2), \u03b1-alactose .N-acetylhexosaminidase , which is involved in the hydrolysis of N-acetylated oligo/polysaccharides such as chitooligosaccharides and chitin, which are prevalent and abundant in marine crustaceans. MAGs containing genes for \u03b2-N-acetylhexosaminidase were present only within the Gammaproteobacteria and include the orders Arenicellales, Burkholderiales, Enterobacterales, Nitrosococcales, Pseudomonadales, UBA11654, and Xanthomonadales. Lastly, genes encoding laminarin endo-1,3-\u03b2-d-glucosidase (GH16 and -81) were found to be enriched in 25 SPA MAGs, which degrades laminarin, a polysaccharide abundant in microalgae such as diatoms  enable rapid transcriptional responses to environmental variations, including light, temperature, and nutrient availability . TCSs arExamination of the SPA and FL MAGs for genes encoding TCSs showed significantly higher representation of histidine kinases  in PA MAGs than in FL MAGs . In addiExtracellular protein secretion systems (ESSs) are widespread among bacteria and archaea and are central components in fimbria- and pilus-associated attachment and adhesion, nutrient transport, predation, cell-cell interactions, surface colonization, and pathogenicity in diverse environmental contexts and settings \u201374. The \u201310.1128/mbio.01569-22.5FIG\u00a0S5FIG\u00a0S5, PDF file, 0.1 MB.Frequency of different extracellular secretion systems in particle-associated versus free-living bacterial genomes in the water column. Genes encoding bacterial secretion systems were predicted using MacSysFinder with the TSScan reference database (v1.0rc1). Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the Prior work has suggested that deep-sea bacteria may have larger genomes than bacteria in the epipelagic zone , 76, andPA microbial communities are often viewed as \u201chot spots\u201d for microbial activity in comparison to their FL counterparts, based on their larger cell size and cell densities, higher enzyme activities, and overall rates of heterotrophic microbial metabolism , 12, 13.77\u201380\u2013in situ inorganic nitrogen availability and depth for FL bacterioplankton at Station ALOHA  had significantly larger genome sizes on average than FL microbes  . In termon ALOHA  44). No. Noin sion ALOHA . As for on ALOHA . These gon ALOHA .Recently, codon usage bias CUB) metrics in genes encoding ribosomal proteins have been used to estimate the maximum growth rates (minimum doubling times) of microorganisms in both laboratory and environmental settings \u201387. We t\u2013UB metric10.1128/mbio.01569-22.10TABLE\u00a0S5Table\u00a0S5, XLSX file, 0.03 MB.Estimated growth efficiencies based on MAG codon usage biases. Download Copyright \u00a9 2022 Leu et al.2022Leu et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the Prochlorococcus as well as motile heterotrophic copiotrophs like Idiomarina. These results further illustrate the distinctive microhabitat-specific phylogenetic, physiological, and ecological divergence between SPA and FL microbial communities  and energy , 61, 67.Previous metagenomic and metaproteomic studies of 0.2-\u03bcm-filtered samples throughout the water column reported that total peptidases and CAZymes decreased from epipelagic (0 to 200 m) to bathypelagic  microbial communities, while the percentage of extracellular enzymes and their diversity increased with depth . In our Given their particle-associated lifestyles, we postulated that SPA microbial genomes are also enriched in genes that promote environmental sensing and response, motility, attachment, colonization, and cell-cell interactions. Consistent with this hypothesis, both the shallow- and deep-water SPA microbes contained greater proportions of genes associated with two-component regulatory systems, extracellular secretion systems, and flagella than did FL microbes from the same depth horizons. The genomic distributions of TCSs support prior observations with respect to general metabolic types  and suggPrior gene-centric studies have demonstrated that genome size, GC content, and genome and proteome nitrogen content tend to positively correlate with environmental inorganic nitrogen availability among FL bacterioplankton populations , 81, 82.3\u2212, NO2\u2212) to reduce the energetic costs of nitrogen acquisition. Our data further suggest that as particles sink and nitrogen depletion ensues, SPA microbial assemblages undergo dynamic compositional shifts in response to the more nutrient-depleted particle microenvironments found at greater depths.SPA microbes, however, showed an opposite trend with respect to DNA GC content and proteome N-ARSC with increasing depth. Strikingly, shallow-water SPA microbes collected at 150 m had higher DNA GC and proteome nitrogen contents than did deeper, 4,000-m SPA microbes. The higher genome and proteome nitrogen content in shallow- versus deep-water SPA microbes is consistent with recent studies of the elemental composition and energy content (in Joules per unit mass of organic carbon)  of sinkiin situ maximum growth rates (minimum doubling times) of prokaryote environmental settings  at 37\u00b0C for 30\u2009min. Subsequently, 50\u2009\u03bcL of a proteinase K solution (0.8\u2009mg mL\u22121) was added, followed by the addition of 50\u2009\u03bcL of 10% SDS. Samples were incubated at 55\u00b0C for 2\u2009h. Final DNA purification was robotically performed using a Chemagen MSM I instrument with the CMG-1037 DNA saliva kit .Metagenomic sequence data from FL samples see \u201346. Brie\u2013Sequencing libraries were prepared and sequencing for all the above samples was performed as described previously , 45. Brihttps://sourceforge.net/projects/bbmap/) in two passes. The first pass used parameters \u201cktrim=r k\u2009=\u200923 mink\u2009=\u200911 hdist\u2009=\u20091 tbo tpe tbo tpe\u201d for Illumina sequencing adapters, and the second pass used parameters \u201ck\u2009=\u200927 hdist\u2009=\u20091 qtrim=rl trimq\u2009=\u200917 cardinality=t mingc\u2009=\u20090.05 maxgc\u2009=\u20090.95\u201d to remove phiX, low-quality bases, and sequences with unrealistically high or low GC. Additional low-quality bases and sequences were removed with Trimmomatic 0.38 (parameters: LEADING:10 TRAILING:10 MINLEN:100) (https://github.com/lh3/seqtk). The cleaned reads from each sample were assembled using SPAdes 3.11.1   . Unpaire,99,127) .https://github.com/wwood/CoverM). For each assembled metagenome, metagenome-assembled genomes were recovered using MetaBAT1 v0.32.5 (https://github.com/wwood/finishm).Mapping of quality reads was performed using CoverM v0.4.0 with default parameters  were reclassified as PA_Shallow (see Methods)  , since tMethods) .Sixty-one 4,000-m PA_Deep MAGs were reannotated as PA_Shallow based on their photic zone-dependent gene representation, well documented physiologies and lifestyles , and/or depth profile read mapping densities. These included the following MAGs: DT-Alcanivorax-1, DT-Alteromonas-1, DT-Bdellovibrionales-2, DT-Bdellovibrionales-3, DT-Caenarcaniphilales-1, DT-Chromohalobacter-1, DT-Cognatishimia-1, DT-Coraliomargarita-1, DT-Dinoroseobacter-1, DT-Ekhidna-1, DT-Epibacterium_A-1, DT-Erythrobacter-1, DT-Erythrobacter-2, DT-Flavobacteriaceae-1, DT-Flavobacteriaceae-3, DT-Flavobacteriaceae-4, DT-Flavobacteriales-4, DT-Gilvibacter-1, DT-Halieaceae-1, DT-Halioglobus-1, DT-Halomonas-1, DT-Halomonas-3, DT-Henriciella-1, DT-Henriciella-2, DT-Henriciella-3, DT-Hyphomicrobiaceae-1, DT-Idiomarina-1, DT-Idiomarina-3, DT-Ilumatobacteraceae-1, DT-Legionellales-1, DT-Micavibrionaceae-1, DT-Mycoplasmatales-1, DT-Mycoplasmatales-2, DT-Oleispira-1, DT-Oligoflexales-1, DT-Parvularculaceae-1, DT-Phaeodactylibacter-1, DT-Phycisphaerales-1, DT-Pseudoalteromonas-1, DT-Pseudobacteriovorax-1, DT-Pseudomonadales-2, DT-Psychroserpens-1, DT-Psychrosphaera-1, DT-Rhizobiales-4, DT-Rhodobacteraceae-1, DT-Rhodobacteraceae-2, DT-Rhodobacteraceae-3, DT-Richelia-1, DT-Rickettsiaceae-1, DT-Rickettsiales-1, DT-Rivularia-1, DT-Salinicola-1, DT-Saprospiraceae-1, DT-Saprospiraceae-2, DT-Shewanella-1, DT-Simkaniaceae-1, DT-Verrucomicrobiales-1, DT-Vibrio-1, DT-Winogradskyella-1, DT-Winogradskyella-2, DT-Xanthomonadales-1, and DT-Crocosphaera.https://github.com/Ecogenomics/GTDBTk). Briefly, marker genes were identified in each genome, aligned, concatenated, and classified with pplacer to identify the maximum-likelihood placement of each genome\u2019s concatenated protein alignment in the GTDB-Tk reference tree. GTDB-Tk classifies each genome based on its placement in the reference tree, its relative evolutionary distance, and FastANI distance.Classification of the MAGs was determined using GTDB-Tk  v.1.3.0 https://github.com/bbuchfink/diamond.git) against UniRef100 (accessed September 2019)  , clusterer 2019) , and Pfaer 2019)  and TIGRer 2019) . Genes wer 2019) .https://github.com/JessAwBryant/gene-characteristics.Amino acid sequences from the dereplicated genome set were used to calculate N-ARSC and C-ARSC values using custom scripts from Mende et al.  and avaiGenes were annotated against dbCAN (3.0) with default parameters  and filt\u221210. SignalP v5.0  domain annotations , 110. HiGenes encoding the bacterial secretion systems were predicted using MacSysFinder  and the https://www.protocols.io/view/viral-sequence-identification-sop-with-virsorter2-5qpvoyqebg4o/v3). Prophage genes were annotated using DRAM-v.py v1.2.4  ordination analysis with the Jaccard distance matrix using the metaMDS function in the Vegan v2.6-2 package in R , 119.To calculate the relative abundance, reads from each metagenomic data set were mapped to the dereplicated MAGs using CoverM v0.3.1 with the \u201ccontig\u201d command, a cutoff of 95% minimum identity, and a minimum aligned read length of 75% of each read. Coverage of each contig was calculated with the CoverM \u201ctrimmed_mean\u201d option, and the coverage for each MAG was calculated as the average of all contig coverages, weighted by their length. The relative abundance of each MAG in each metagenomic data set was calculated as its coverage divided by total reads in the sample multiplied by 100,000,000.The presence/absence of each KO ID or orthologous family in each genome was used as input for the program Scoary  to identP values that were affected by multiple testing were corrected for false discovery using the Benjamini-Hochberg procedure.The mean statistical significance of different proteins of interests and genomic characteristic metrics between sample types were determined through one-way analysis of variance (ANOVA), followed by the Tukey test using the R package multicompview v0.1-8 with the Tukey honestly significant different (HSD) function. All https://github.com/dparks1134/CompareM).Average amino acid identity (AAI) between the genomes was calculated using orthologous genes identified through reciprocal best BLAST hits by use of compareM v0.0.5 (http://bioinformatics.psb.ugent.be/webtools/Venn/). Figures were further refined using Adobe Illustrator.Figures were generated using pheatmap  and ggplPRJNA482655, and assembled MAGs are available under NCBI BioSample no. SAMN14675689 to SAMN14675809. Metagenomic reads produced from DNA extracted from shallow 150-m sediment trap samples (collected in 2015) (PRJNA358725. For Station ALOHA 0.2-\u03bcm-pore-size filter-collected metagenomes/MAGs, read sequence data and FL MAGs are available at NCBI SRA under BioProject no. PRJNA352737, and assemblies can be found under BioSample no. SAMN12604809.All data were deposited in the NCBI SRA archive as follows. Deep 4,000-m-trap metagenomes and MAGs are deposited under NCBI BioProject no."}
+{"text": "Invasive mucormycosis (IM) is associated with high mortality and morbidity. MAT2203 is an encochleated oral formulation of amphotericin B which has been shown to be safe and effective against murine aspergillosis and murine cryptococcal meningoencephalitis. We sought to compare the efficacy of MAT2203 to liposomal amphotericin B (LAMB) treatment in a neutropenic mouse model of IM.Rhizopus delemar 99-880 or M. circinelloides f. jenssenii DI15-131. Treatment with placebo (diluent control), oral MAT2203  or LAMB , began 16 h post infection. Survival (n=10-20/group from 1/2 experiments) through Day +21 and tissue fungal burden of lungs or brain (n=10/group) euthanized on Day +4 post infection  served as a primary and secondary endpoint, respectively.ICR mice were immunosuppressed with cyclophosphamide and cortisone acetate on days -2, -3 and +8, relative to infection with intratracheally instilled Rhizopus delemar infection, doses of MAT2203 5,15 mg/kg qd or 7.5 mg/kg bid, significantly prolonged median survival time (MST) and enhanced overall survival vs. placebo-treated mice . Importantly, MAT2203 treatments were as effective as LAMB . For mice infected with M. circinelloides, MAT2203 at 15 mg/kg, qd significantly prolonged MST and enhanced overall survival vs. placebo-treated mice . In both infection models MAT2203 treatment of 15 mg/kg or LAMB resulted in significant \u223c1.0-1.5 log reduction and \u223c2.0-2.2 log reduction in lung and brain fungal burden vs. placebo, respectively (Wilcoxon Rank Sum).For in vivo efficacy in treating R. delemar or M. circinelloides pulmonary infection in immunosuppressed mice, which was equivalent to the LAMB current standard of care. Continued investigation and development of MAT2203 as a novel, and oral formulation of amphotericin antifungal agent against mucormycosis is warranted.MAT2203 demonstrated Theresa Matkovits, PhD, Matinas BioPharma: Employee Jenel Cobb, PhD, Matinas BioPharma: Employee Raphael J. Mannino, PhD, Matinas BioPharma: Employee Ashraf S. Ibrahim, PhD, Matinas BioPharma: Advisor/Consultant|Matinas BioPharma: Grant/Research Support|SFunga: Grant/Research Support."}
+{"text": "Vaccination strategies that provide enhanced immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are needed. We evaluated the safety and immunogenicity of a bivalent omicron containing vaccine, mRNA-1273.214 (50 \u00b5g), administered as a second booster dose in adult participants.In this ongoing phase 2/3 trial, 50 \u00b5g of the bivalent vaccine mRNA-1273.214  or 50 \u00b5g of the authorized mRNA-1273 were administered as second boosters in adults who previously received a 2 dose (100 \u00b5g) primary series and a first booster (50 \u00b5g) dose of mRNA-1273 (\u2265 3 months prior). Primary objectives were safety and reactogenicity and immunogenicity 28 days post-booster dose.Figure). Safety and reactogenicity were similar for both vaccine groups.In participants with no prior SARS-CoV-2 infection who received booster doses of mRNA-1273.214 (n=334) or mRNA-1273 (n=260), neutralizing antibody (nAb) geometric mean titers ]) against omicron BA.1 were 2372.4 (2070.6\u22122718.2) and 1473.5 (1270.8\u22121708.4), respectively. The model-based GMT ratio (GMR [97.5% CI]) of mRNA-1273.214 compared to mRNA-1273 was 1.75 (1.49\u22122.04), meeting the pre-specified superiority criterion against omicron BA.1. The pre-specified criterion for non-inferiority against the ancestral SARS-CoV-2 strain was also met. Additionally, mRNA-1273.214 elicited higher GMTs (727.4 [632.8\u2212836.1]) than mRNA-1273 (492.1 [431.1\u2212561.9]) against omicron subvariants BA.4/BA.5 [GMR (95% CI) 1.69 [1.51\u22121.90])]. Binding antibody responses against alpha, beta, gamma, delta, and omicron were numerically higher in the mRNA-1273.214 group compared to mRNA-1273. mRNA-1273.214 GMTs were consistently higher across age (18-< 65 and \u2265 65 years) and pre-booster SARS-CoV-2 infection subgroups (The bivalent omicron containing mRNA-1273.214 elicited superior nAb responses against omicron 28 days post-immunization compared to mRNA-1273 regardless of age and prior SARS-CoV-2 infection; no new safety concerns were identified.Spyros Chalkias, MD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Stephen R. Walsh, MD, Janssen Vaccines: Grant/Research Support|Moderna, Inc.: Grant/Research Support|NIAID/NIH: Grant/Research Support|Sanofi Pasteur: Grant/Research Support Nichole McGhee, B.S., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Joanne Tomassini, Ph.D., Moderna, Inc.: Advisor/Consultant Xing Chen, Sc.D., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Ying Chang, M.S., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Andrea Sutherland, M.D., MPH, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds David Montefiori, Ph.D., Moderna, Inc.: Grant/Research Support Bethany Girard, Ph.D., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Darin Edwards, Ph.D., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Jing Feng, M.S., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Honghong Zhou, Ph.D., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Lindsey R. Baden, MD, Moderna, Inc.: Grant/Research Support|NIAID: Grant/Research Support Jacqueline Miller, MD, Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds Rituparna Das, M.D., Moderna, Inc.: Salary|Moderna, Inc.: Stocks/Bonds."}
+{"text": "Piper species  has been studied, along with their antifungal and phytotoxic activities. These EOs contained \u03b2-bisabolene/nerolidol (Pc), \u03b2-bisabolene/\u03b4-cadinene/caryophyllene (Pt), caryophyllene oxide (Pcs), bicyclogermacrene/10-epi-Elemol (Po), bicyclogermacrene/germacrene-D/apiol (Pd), caryophyllene/germacrene-D (Pa), germacrene-D (Pr), limonene/apiol (Ps), apiol (Psf), and apiol/bicyclogermacrene (Pm) as major components, and some are described here for the first time . A composition-based dendrogram of these Piper species showed four major groups . The spore germination effects  and phytotoxicity (Lolium perenne and Lactuca sativa) of these EOs were studied. Most of these Piper essential oils showed important activity against phytopathogenic fungi (except G1), especially against B. cinerea. Similarly, most of the essential oils were phytotoxic against L. perenne (except G1), with P. sancti-felicis (G4), P. casapiense (G2), and P. reticulatum (G2) being the most effective. Caryophyllene oxide, \u03b2-caryophyllene, \u03b2-pinene, limonene, \u03b1-humulene, and apiol were evaluated against B. cinerea,\u00a0with the most effective compounds being \u03b2-pinene, apiol, and limonene. This work demonstrates the species-dependent potential of essential oils from Peruvian Piper species as fungicidal and herbicidal agents.The chemical composition of essential oils (EOs) from ten Peruvian  Diseases caused by plant pathogens significantly contribute to annual loss in crop yield worldwide . The appIn plants, essential oils (EOs) play a protective role against herbivores, phytopathogenic fungi, and weeds. Essential oils also represent a new class of crop protectants due to their volatility and low toxicity to the environment and have been proposed as an alternative to synthetic pesticides ,7,8,9. APiper is found in tropical and subtropical areas, and in America, there are approximately 700 species [The Piperaceae family has approximately eight genera and 3000 species . The gen species ,13, with species ,15. This species , includi species ,17,18, a species ,20, inse species ,21.Piper EOs are characterized by the presence of monoterpene hydrocarbons , oxygenated monoterpenoids , sesquiterpene hydrocarbons , oxygenated sesquiterpenoids -nerolidol, caryophyllene oxide, \u03b1-cadinol, epi-\u03b1-bisabolol), and phenylpropanoids , among others [Piper essential oils have been described as being insect antifeedant, acaricidal, nematicidal, and herbicidal agents [Piper essential oils are a promising source of new potential biopesticide ingredients.g others ,22,23,24l agents ,21. TherPiper species for their biopesticidal potential, ten species native to the Peruvian Amazonian region  have been extracted by hydrodistillation to study the chemical composition of their EOs by GC-MS, along with their fungicidal activity against phytopathogens  and their phytotoxic effects (against Lolium perenne and Lactuca sativa) to assess their potential applications in phytopathogen and/or weed control.As part of an ongoing project on the bioprospection of Peruvian Piper coruscans; \u03b2-bisabolene (40.2%), \u03b4-cadinene (9.8%), caryophyllene (9.7%), germacrene-D (5.0%), and nerolidol (4.5%) for P. tuberculatum; caryophyllene oxide (10.2%), and caryophyllene (4.7%) for P. casapiense; bicyclogermacrene (7.9%), 10-epi-Elemol (7.3%), caryophyllene (6.3%), \u03b1-pinene (6.0%), \u03b2-pinene (5.1%), \u03b2-selinenol (4.9%), \u03b1-eudesmol (4.5%), and camphene (4.4%) for P. oblicuum; bicyclogermacrene (16.5%), germacrene-D (10.4%), apiol (8.9%), caryophyllene (6.8%), \u03b2-pinene (6.3%), \u03b1-cubebene (5.9%), and \u03b2-elemene (4.5%) for P. dumosum; caryophyllene (11.3%), germacrene-D (9.6%), \u03b1-humulene (6.6%), \u03b4-cadinene (6.6%), and (-)-\u03b2-copaene (5.8%) for P. anonifolium; germacrene-D (12.6%), bicyclogermacrene (8.1%), \u03b4-cadinene (6.0%), copaene (4.6%), and caryophyllene (4.5%) for P. reticulatum; limonene (38.5%), apiol (15.0%), caryophyllene oxide (8.4%), eudesma-3,7-(11)-diene and copaene (5.8%) for P. soledadense; apiol (76.1%), and caryophyllene (4.1%) for P. sancti-felicis and apiol (51.6%), bicyclogermacrene (9.0%), germacrene-D (6.7%), and myristicin (4.6%) for P. mituense.The chemical compositions of the essential oils are shown in P. sancti-felicis and P. mituense, which were characterized by phenylpropanoids, and P. soledadense, which was characterized by monoterpene hydrocarbons.The overall composition of these oils is shown in Piper species (P. coruscans (Pc) and P. tuberculatum (Pt), characterized by the presence of sesquiterpene hydrocarbons; (G2) P. casapiense (Pcs), P. obliquum (Po), P. dumosum (Pd), P. anonifolium (Pa), and P. reticulatum (Pr), characterized by sesquiterpenes; (G3) P. soledadense (Ps), with monoterpenes and sesquiterpenes; and (G4) P. sancti-felicis (Psf) and P. mituense (Pm), characterized by phenylpropanoids.A dendrogram based on the composition of the  species  showed fPiper essential oils against Aspergillus niger, Botrytis cinerea, and Alternaria alternate is shown in B. cinerea was the fungal species most susceptible to the action of Piper essential oils. The antifungal activity showed a pattern in accordance with the grouped EOs. P. coruscans and P. tuberculatum (G1) were not active. P. casapiense (G2), P. obliquum (G2), P. dumosum (G2), P. anonifolium (G2), and P. reticulatum (G2) were only active against B. cinerea with varying potencies, with P. obliquum and P. anonifolium being the most effective. P. soledadense (G3) inhibited the spore germination of A. niger and B. cinerea. P. sancti-felicis (G4) oil was active against all three fungal species with moderate effects, and P. mituense (G4) only acted on B. cinerea, probably due to the lower concentration in apiol of this oil.The antifungal activity (spore germination inhibition) of the Piper oils components tested against B. cinerea , \u03b2-pinene showed strong antifungal activity , followed by apiol  and limonene , with effective doses similar to the positive control  for apiol.Among the Lactuca sativa (dicotyledonous) and Lolium perenne (monocotyledonous) plants. The phytotoxic activity did not follow the grouping pattern observed for the antifungal effects. P. sancti-felicis (G4), P. casapiense (G2), P. mituense (G4), and P. reticulatum (G2),\u00a0effectively inhibited germination, leaf and root growth of L. perenne , P. obliquum (G2), P. dumosum (G2), and P. soledadense (G3)\u00a0inhibited leaf growth (>50%), followed by P. tuberculatum (G1) with a 50% inhibition.The essential oils were tested for phytotoxic effects on ne >50%, . P. anonP. sancti-felicis, P. casapiense, P. reticulatum, and P. mituense effectively inhibited the root growth of L. sativa (data not shown). P. soledadense reduced the root growth of L. sativa (data not shown). These results show strong selective herbicidal potential of the EOs tested against monocotyledonous plants.Piper essential oils present a wide variety of chemical compounds with important biological activities that may be of interest in agriculture, medicine, and food industries, among others. These oils play an important role in the defense of the plant against pests, and many studies have reported activities as insecticidal, antifeedants, phytotoxic and antifungal [tifungal ,26,27,28Piper species described here have been previously reported to show quantitative and qualitative chemical variations that can be attributed to environmental factors  [The essential oils from some of the umidity) ,29,30. AP. coruscans (Pc) and P. tuberculatum (Pt). P. coruscans EO had \u03b2-bisabolene (33.4%) and nerolidol (10.2%) as the main components, while this species collected in Ecuador contained \u03b2-caryophyllene (24.1\u201325.0%), \u03b1-humulene (11.6\u201312.0%), and caryophyllene oxide (9.3\u201310.9%) [P. tuberculatum studied here showed \u03b2-bisabolene (40.2%) as the main component followed by \u03b4-cadinene (9.8%), caryophyllene (9.7%), germacrene-D (5.0%), nerolidol (4.5%), copaene (4.2%), and \u03b2-elemene (3.3%). This Piper species (Pt) EO has been previously reported for plants collected from different locations. Pt collected in Venezuela gave an EO with \u03b1-farnesene (6.2%), humulene epoxide II (6.0%), 2-pentadecanone (4.1%), \u03b2-eudesmol (4.4%), 2-tridecanone (4.3%), ledane (3.6%), -farnesylacetone (3.6%), and \u03b1-cadinol (2.9%) [E)-caryophyllene (30.1%) [E-cariofilene (7.1%), and trans-4-muurola(14)-5-diene (9.9%) [(G1) 3\u201310.9%) . The EO l (2.9%) , while E (30.1%)  or myrissabolene .4% and nP. casapiense (Pcs), P. obliquum (Po), P. dumosum (Pd), P. anonifolium (Pa), and P. reticulatum (Pr): P. casapiense (Pcs), is reported here for the first time. The EO from P. obliquum had bicyclogermacrene (7.9%), 10-epi-Elemol (7.3%), caryophyllene (6.3%), and \u03b1-pinene (6.0%). However, the Po essential oil of Ecuadorian origin contained safrole (45.9%), \u03b3-terpinene (17.1%), and terpinolene (11.5%) [P. dumosum contained bicyclogermacrene (16.5%) and germacrene-D (10.4%). Similarly, the EO from Pd plants collected in the Brazilian Amazonia had bicyclogermacrene (16.2%), \u03b2-caryophyllene (15.9%), \u03b2-pinene (16.0%), and \u03b1-pinene (12.1%) [P. anonifolium studied here contained caryophyllene (11.3%), germacrene-D (9.6%), \u03b4-cadinene (6.6%), \u03b1-humulene (6.6%), and neoalloocimene (5.5%), while a previously reported EO from the Brazilian Par\u00e1 was composed of selin-11-en-4-\u03b2-\u03b1-ol (20.0%), \u03b2-selinene (12.7%), \u03b1-selinene (11.9%), and \u03b1-pinene (8.8%) [P. reticulatum studied here contained the phenylpropanoid apiol as the main component (15.0%) while \u03b2-elemene (24.6%) and \u03b2-caryophyllene (16.7%) were abundant in a Pr oil from the northern region of Brazil [(G2)  (11.5%) , while a (11.5%) . P. dumo (12.1%)  as the me (8.8%) . Furthere (8.8%) . The EO f Brazil .soledadense (Ps) is reported here for the first time.(G3) P. P. sancti-felicis (Psf) and P. mituense (Pm). The EO from P. sancti-felicis and P. mituense (Pm reported here for the first time) contained the phenylpropanoid apiol as the main component . \u03b4-3-carene (35.3%) and limonene (27.1%) were the main components of the EO from P. sancti-felicis collected in Choco, Colombia [(G4) Colombia ,25.Piper have been reported to have a wide range of biological properties [P. tuberculatum -\u03b2-ocimene 14% and \u03b2-caryophyllene 32.1%) [P. anonifolium  showed strong antifungal activity against Cladosporium cladosporioides and C. sphaerospermum [The essential oils studied here have shown important activities against phytopathogenic fungi (G2-4). Essential oils from the genus operties ,14,21,40operties ,38,39. Se 32.1%) , and P. ospermum .B. cinerea , \u03b2-pinene, limonene, and apiol showed strong antifungal activity. Therefore, \u03b2-pinene could contribute to the activity of Po and Pd EOs, while apiol and limonene could explain the effect of Ps, Psf, and Pm oils. \u03b2-caryophyllene (inactive) could be a synergist.The composition-based grouping of the EOs overlapped with the antifungal activity, suggesting that the presence of bicyclogermacrene, 10-epi-Elemol, germacrene-D, caryophyllene, limonene, \u03b2-pinene, and/or apiol could be responsible for significant antifungal effects. Among the oil components tested against Rhizoctonia solani, Fusarium oxysporum, Penicillium digitatum and Asperigallus niger [Botrytis cinerea [Aspergillus flavus, A. niger, A. fumigatus, and A. parasiticus [Botryodiplodia theobromae and Colletotrichum acutatum [Fusarium solani [Rhizoctonia solani and Helminthosporium oryzae [B. cinerea spore germination.(S)-Limonene has reported antifungal activity against us niger . Apiol s cinerea , Aspergiasiticus , Botryodacutatum ,46. The acutatum . Caryophm solani  and carym oryzae , but in P. sancti-felicis, P. mituense, P. casapiense, P. reticulatum, P. anonifolium, P. obliquum, P. dumosum, and P. soledadense) were phytotoxic to the monocotyledonous Lolium perenne, and this activity did not overlap with the composition-based groups, suggesting a multi-component phytotoxic action. This is the first report on the phytotoxic effects of these species, except for an EO from P. sancti-felicis that was not active and did not contain apiol [Piper species, including P. hispidinervum rich in safrole [P. dilatatum rich in apiol, and P. divaricatum rich in eugenol and methyleugenol [Most of the essential oils tested here  [Brassica campestris and Raphanus sativus [Echinochloa crusgalli, Lolium perenne, Amaranthus retroflexus, and Digitaria sanguinalis [Among the main compounds present in the  species , apiol i plants) , and car sativus  and the guinalis .2SO4. All investigated Piper species contained essential oils that range from 0.078 to 1.26% based on dry weight  of the selected Piperaceae species were collected in Iquitos, Loreto Department, Peru in different seasons. The taxonomic identification was carried out at the Herbarium Amazonense of the National University of the Peruvian Amazon, Iquitos, Peru. A voucher for each species has been deposited in the herbarium. All the plants were permitted for collection . The EOs extraction was performed by hydrodistillation using the dried aerial parts of the plants. The EOs were separated by decantation and dried over anhydrous Nay weight .Essential oils were analyzed by gas chromatography (GC) on a Shimadzu 2010 and gas chromatography-mass spectrometry (GC-MS) equipped with a mass spectrometer Shimadzu GCMS-QP2010-Ultra Mass Detector . The carrier gas was helium. The capillary column was a Teknokroma TRB (95%) dimethyl (5%) dimethylpolysiloxane (30 m \u00d7 0.25 mm ID and 0.25 \u00b5m phase thickness). Working conditions were as follows: injector temperature, 300 \u00b0C; column temperature 70\u2013290 \u00b0C, for 6 min, staying at 290 \u00b0C for 15 min, temperature of the transfer line connected to the mass spectrometer, 250 \u00b0C, and ionization source temperature 250 \u00b0C. The identification of compounds was performed with standard terpenes analyzed under the same conditions and by comparison of the mass spectra with those available in the library Wiley Mass Spectral Database , while relative area% has been used for quantification of all the peaks obtained in the chromatograms. The mass spectra and Kovats retention indexes obtained were compared with the literature reported ,54.Aspergillus niger, Alternaria alternata, and Botrytis cinerea came from the fungal collection of Instituto de Ciencias Agrarias-CSIC, Madrid, Spain where they are maintained. The antifungal activity of the essential oils was determined using a modified spore germination inhibition growth assay [5 cells/mL in NaCl 0.9% for A. niger and 1 \u00d7 107 cells/mL in distilled water for B. cinerea and A. alternate. Amphotericin B (5 \u00b5g/mL) was used as a positive control.The fungal species th assay . The essA. niger and 25 \u00b0C for B. cinerea). After the incubation process, 25 uL of an MTT (5 mg/mL) plus menadione (1 mM) solution in RMPIMOPS were added, the plates were incubated again for 3 h, the medium was removed, 200 \u00b5L of acidic isopropanol (95% isopropanol and 5% 1 M HCl) was added, and the plates were incubated for another 30 min. The absorbance was read at 490 nm in an Elisa reader. The IC50 values (the effective dose to give 50% inhibition) were calculated by a regression curve of % spore germination inhibition on log dose.The samples and spore suspensions (4 replicates) were placed on 96-well plates and incubated for 24 h .Lactuca sativa, and Lolium perenne seeds (40 seeds/test) in 12-well microplates, as described previously [L. sativa) or seven days (L. perenne), and the root length  was measured  at the end of the experiment. A nonparametric analysis of variance (ANOVA) was performed on root/leaf length data [These experiments were conducted with eviously . The essgth data ,21.https://www.statgraphics.com, accessed on 2 July 2022).The data were analyzed using STATGRAPHICS Centurion XIX  subjected to cluster analysis . The groups were chosen with a distance >2.Piper species as fungicidal and herbicidal agents based on their composition. A dendrogram based on the composition of the Piper species showed four groups: (G1) P. coruscans (Pc) and P. tuberculatum (Pt), characterized by the presence of sesquiteterpene hydrocarbons; (G2) P. casapiense (Pcs), P. obliquum (Po), P. dumosum (Pd), P. anonifolium (Pa) and P. reticulatum (Pr), characterized by sesquiterpenes; (G3) P. soledadense (Ps), with monoterpenes and sesquiterpenes; and (G4) P. sancti-felicis (Psf) and P. mituense (Pm), characterized by phenylpropanoids. The essential oils in G2-4 showed important activity against Botrytis cinereal and were phytotoxic against Lolium perenne.This work demonstrates the species-dependent potential of essential oils from Peruvian Considering the composition-based grouping of the EOs, we can conclude that the presence of bicyclogermacrene, 10-epi-Elemol, germacrene-D, caryophyllene, limonene, \u03b2-pinene, and/or apiol could be responsible for significant antifungal and herbicidal effects. \u03b2-pinene, apiol, and limonene showed antifungal activity, but not caryophyllene, suggesting that this compound could be a synergist.Piper germplasm bank and the domestication of selected species to grant a sustainable biomass source for the production of essential oils with biopesticidal activity.These findings have important implications for the development of a"}
+{"text": "The aim of this study was to determine the sensitivity (SE), specificity (SP), positive predictive value (PPV), and negative predictive value (NPV) for PC-local recurrence and metastases on a per region basis.Despite increasing use for the detection of biochemically recurrent prostate cancer (rPC), the diagnostic accuracy of positron emission tomography/computed tomography (PET/CT) with [18F]PSMA-1007 PET/CT for rPC were retrospectively analysed. Six body regions were defined: prostate fossa, pelvic lymph nodes (LN), retroperitoneal LN, supradiaphragmatic LN, bones, and soft tissue. A region was counted positive if at least one PSMA-positive lesion suspicious for PC was observed. Confirmation of a true-positive PSMA-avid lesion was defined as positive by histopathology, fall in serum prostate-specific antigen (PSA) (> 50%) after targeted therapy or confirmatory further CT, MRI, PET/CT, or bone scan imaging. Regions where additional imaging was able to confirm the absence of suspicious PC lesions or regions outside exclusively targeted RT with serum PSA decline (> 50%) were counted as true-negative regions. SE, SP, PPV, and NPV were calculated for all six regions.One hundred seventy-seven consecutive patients undergoing [n = 6) was too small for an accurate statistical analysis.The overall PET-positivity rate was 91%. Conclusive follow-up for affirmation or refutation of a PSMA-positive lesion was available for 81/152 patients on a per region basis. In this subgroup, overall sensitivity, specificity, PPV, and NPV were 95% (CI: 0.90\u20130.98), 89% (CI: 0.83\u20130.93), 86% (0.80\u20130.90), and 96% (CI: 0.92\u20130.98), respectively. On a per region basis, PPV was 97% (CI: 0.83\u20130.99) for local recurrence, 93% (CI: 0.78\u20130.98) for pelvic LN, 87% (CI: 0.62\u20130.96) for retroperitoneal LN, 82% (CI: 0.52\u20130.95) for supradiaphragmatic LN, and 79% (0.65\u20130.89) for bone lesions. The number of solid organ metastases  sensitivity and NPV on a per region basis. However, overall PPV was limited (86%), particularly for bone lesions (79%), which are a potential diagnostic weaknesses when using this tracer.The known high PET-positivity rate of [The online version contains supplementary material available at 10.1007/s00259-022-05693-0. Radioligands to the prostate-specific membrane antigen (PSMA) have become the gold standard for the staging of primary prostate cancer (PC) \u20134 and re68Ga]Ga-PSMA-11, a large number of alternative radioligands have become available, including but not limited to [68Ga]Ga-PSMA-I&T; [68Ga]Ga-THP-PSMA; and [18F]-labelled PSMA-radiotracers such as [18F]-rhPSMA-7, [18F]-DCFPyL, [18F]-JK-PSMA-7, or [18F]PSMA-1007 [18F]-labelled PSMA-radiotracers have numerous advantages over 68Ga-labelled ligands: [18F] is cyclotron produced with a longer half live and a lower positron energy compared to [68Ga] (0.65 MeV vs. 1.90 MeV), leading to improved spatial resolution [18F]PSMA-1007 suggest improved detection rates especially in local relapses and pelvic lymph node metastases in proximity to the urinary tract [18F]PSMA-1007 might be of benefit [68Ga]Ga-PSMA-11 can be an alternative PSMA-1007 in rPC. Hitherto, studies report only preliminary observations in small cohorts (n = 40) [18F]PSMA-1007 in a cohort of men undergoing PSMA-PET/CT for rPC in a clinical setting on a per patient and per region basis using a composite standard of truth (CSOT) for verification or refutation of imaging findings.Despite widespread adoption, few studies report the diagnostic accuracy of PSMA-1007 PET/CT in the University Clinic for Nuclear Medicine, Inselspital, Bern. The period was chosen to enable a minimum follow-up period of 12 months. Inclusion criteria were biochemical recurrence of PC (rPC) according to the ASTRO/AUA Guideline as rising PSA value after a PSA-Nadir after definitive treatment  PSMA-1007 was produced as previously described [18F]PSMA-1007 solution was given by intravenous bolus injection .[escribed . The [1818F]PSMA-1007. They were investigated on either Biograph-mCT PET/CT  (n = 90) or Biograph-VISION 600 PET/CT  PET/CT (n = 96) scanner. The examination protocols and reconstruction algorithms used are previously published PSMA-1007 PET/CT, but where histopathology of the PSMA-avid lesion was negative, where the PSA after targeted RT did not decrease by 50% or where post therapy imaging were not confirmatory. Regions where no PSMA-positive lesion was detected with additional confirmatory imaging or regions outside exclusively targeted RT with PSA decline were counted as true-negative (TN) regions. Patients with no histopathology, no further treatment or imaging, and therefore no composite standard of truth were not included in the final analysis  following exclusively targeted radiotherapy of a PSMA-positive lesion suspicious for PC were true positive (TP). The absence of PSMA-positive legions in a region where additional imaging revealed the presence of PC or where histopathology showed a positive result were counted as false negative (FN). A false-positive (FP) region was counted where a PMSA-positive lesion was observed in 8FPSMA-10 Figures .Fig. 2Co\u03b1) for \u03b1 = 0.05 PSMA-1007 in patients with rPC was 95.6% : 0.90\u20130.98).Of these 81 PSMA-PET/CTs in 73, at least one suspicious PSMA-avid lesion was detected, and therefore, the PET scan was rated as \u201cpositive\u201d. This difference in PET-positivity rate for this subgroup to the overall positivity rate was not statistically significant . On a region basis, PSMA-positive lesions were detected in the prostatic fossa in 51%, in pelvic LN in 48%, in retroperitoneal LN in 23%, in supradiaphragmatic LN in 16%, in bones in 53%, and in other metastasis (soft tissue lesions) in 7%  and 0.89 (CI: 0.83\u20130.93), PPV and NPV were 0.86 (CI: 0.80\u20130.90) and 0.96 (CI: 0.92\u20130.98), and positive and negative likelihood ratio (LR+/LR-) were LR+: 8.71 (CI: 5.69\u201313.34) and LR\u2212: 0.06 (CI: 0.03\u20130.12).n = 6) showed a PPV of 0.40 (0.12\u20130.77) and a NPV of 0.97 (CI: 0.86\u20130.99). Further details are given in Figure Sensitivity and specificity on a region basis were 0.94 (CI: 0.81\u20130.98) and 0.92 (CI: 0.64\u20130.99) for local recurrence, 0.93 (CI: 0.77\u20130.99) and 0.92 (CI: 0.0.73\u20130.99) for pelvic LN, 1.00 (CI: 0.77\u20131.00) and 0.94 (0.80\u20130.98) for retroperitoneal LN, 1.00 (CI: 0.70\u20131.00) and 0.94 (CI: 0.81\u20130.98) for supradiaphragmatic LN, 0.97 CI: 0.85\u20130.99) and 0.74 (CI: 0.57\u20130.85) for bone lesions, and 0.67 (CI: 0.21\u20130.94) and 0.92 (CI: 0.79\u20130.97) for soft tissue lesions. PPV and NPV on a region basis were 0.97 (CI: 0.83\u20130.99) and 0.86 (CI: 0.60\u20130.96) for local recurrence, 0.93 (CI: 0.78\u20130.98) and 0.92 (CI: 0.74\u20130.98) for pelvic LN, 0.87 (CI: 0.62\u20130.96) and 1.00 (CI: 0.89\u20131.00) for retroperitoneal LN, 0.82 (CI: 0.52\u20130.95) and 0.96 (CI: 0.81\u20130.99) for supradiaphragmatic LN, and 0.79 (CI: 0.65\u20130.89) and 0.96 (CI: 0.81\u20130.99) for bone lesions. The small number of soft tissue lesions (\u20130.99 andROC and corresponding area under the curves (AUC) were 0.70 \u00b1 0.03 overall. For local recurrence, AUC was 0.76 \u00b1 0.05, for pelvic LN 0.71 \u00b1 0.09, for retroperitoneal LN 0.55 \u00b1 0.17, and for supradiaphragmatic LN 0.51 \u00b1 0.14. For bone lesions, the AUC was comparatively lower at 0.63 \u00b1 0.06. Corresponding ROC curves are outlined in Figure 18F]PSMA-1007 was clinically first introduced in 2015 [18F]PSMA-1007, there are currently very few evidence-based recommendations supporting its use [18F]PSMA-1007 on a patient-level and a per region basis. In this retrospective analysis of 177 patients, we present the largest cohort of men undergoing [18F]PSMA-1007 for rPC with a confirmatory standard of follow-up PSMA-1007 PSMA-1007. No significant difference in the PET-positivity rate between the subgroup with CSOT and those without was observed (p = 0.21), implying no bias in the patients with follow-up data available.One strength of our study was the high rate of follow-up available for our patients (86%). Through scrutiny of clinical imaging reports, multi-disciplinary team meeting minutes, and clinical records, we are able to demonstrate the performance of this radiopharmaceutical under clinical conditions in a large cohort of men undergoing PSMA-PET/CT in an academic nuclear medicine centre. Nevertheless, in common to all studies with rPC, follow-up data which can be used to verify imaging findings is not available for all patients. For example, a watchful waiting strategy might be chosen by the patient and his treating physicians. Using a CSOT including histopathology, PSA decline (> 50%) after targeted RT, and further imaging after the PSMA-PET showing response to system therapies, we were able to confirm or refute PSMA-PET findings. With a high number of consecutively screened patients with rPC (n = 152) is not small, the relatively smaller number of supradiaphragmatic lymph nodes (n = 13) and solid organ metastases (n = 6) precluded an accurate analysis of diagnostic performance in these regions. Further studies, ideally of prospective design, are required to confirm these data. In order to reduce selection bias, patients were consecutively followed up. This heterogeneous cohort of patients at a variety of stages of rPC and prior treatment furnishes an overview of the radiotracer\u2019s performance under routine clinical conditions. Further studies, for example, directed at early stages of recurrence should be performed, particularly since the PPV, which is dependent upon pre-test prevalence [We highlight several weaknesses with our study. Our overall cohort size was limited by the introduction of the radioligand in our centre in Oct 2019 and the requirement to collect 12 months\u2019 of follow-up. Although our cohort of patients with follow-up Below is the link to the electronic supplementary material."}
+{"text": "COVID-19-related physical isolation, fear and anxiety determined de novo mental illnesses, by potentially facilitating the emergence of Hikikomori traits .The present study aims at screening a cohort of university students for the Hikikomori traits and assessing a set of psychopathological determinants associated with Hikikomori, particularly boredom and loneliness dimensions.A cross-sectional web-based survey was carried out by administering Hikikomori Questionnaire (HQ-11), Italian Loneliness Scale (ILS), Multidimensional State Boredom Scale (MSBS), Depression Anxiety Stress Scale (DASS-21) and Toronto Alexithymia Scale (TAS-20).1,148 respondents  were recruited. 70.7% declared to have experienced psychological distress. HQ-11 average total score was 18.4\u00b1SD=7.5 with statistically significant higher values in the males (p=0.017) and amongst students studying Informatics, Mathematics/Physics/Chemistry, Science of Communication and Engineering. The HQ-11 positively correlated with ILS (r=0.609), MSBS (r=0.415), TAS-20 (r=0.482) and DASS-21 (r=0.434).This study represents the first screening of the Hikikomori phenomenon in Italian university students. Hikikomori traits appear to be particularly represented in the Italian youth population and should be carefully investigated in future studies.No significant relationships."}
+{"text": "Salmonella have changed in recent years as multidrug-resistant (MDR) strains have become prevalent among various serovars. The recent expansion of MDR Salmonella enterica serovar Indiana sequence type 17 (ST17) poses an increasing threat to global public health, as 24.3% (61/251) of S. Indiana isolates in this study exhibited resistance to three clinically important antimicrobial agents: fluoroquinolones (ciprofloxacin), extended-spectrum \u03b2-lactams , and macrolides (azithromycin). Both the evolutionary histories and antimicrobial resistance (AMR) profiles of this serovar remain to be described. Bioinformatic analysis revealed multiple lineages have coexisted and spread throughout China. Specifically, emergence of a predominant lineage appears to be associated with accumulated various substitutions in the chromosomal quinolone resistance-determining regions (GyrA S83F D87N and ParC T57S S80R) (141 [56.2%]), as well as acquisition of an extended-spectrum \u03b2-lactamase (ESBL)-producing IncHI2 plasmid that has subsequently undergone extensive rearrangement and an IncX1 plasmid that contains mph(A), conferring resistance to azithromycin. Several other evolutionary events influencing the trajectory of this drug-resistant serovar were also identified, including sporadic acquisitions of blaCTX-M-carrying plasmids, along with chromosomal integration of blaCTX-M within subclusters. Most human isolates reside in clusters containing isolates from animals, mainly from chickens, indicating the close relationship of human isolates with those from food animals. These data demonstrate that MDR S. Indiana ST17 is already widespread and capable of acquiring resistance traits against the clinical important antimicrobial agents, suggesting it should be considered a high-risk global MDR pathogen. The complexity of its evolutionary history has implications for AMR surveillance, epidemiological analysis, and control of emerging clinical lineages.The genetic features of foodborne IMPORTANCE The emergence and worldwide spread of AMR Salmonella constitute great public health concerns. S. enterica serovar Indiana is a typical MDR serovar characterized by sporadic reports. However, comprehensive population genomics studies have not been performed on this serovar. This study provides a detailed and comprehensive insight into the rapid evolution of AMR in this important Salmonella serovar in the past 15\u2009years in eight provinces of China. We documented diverse contributory genetic processes, including stable chromosomal integrations of resistance genes, the persistence and evolution of mobile resistance elements within sublineages, and sporadic acquisition of different resistance determinants that occur at all genetic levels . There are different mechanisms of antimicrobial resistance in S. enterica serovar Indiana from those of other serovars. This study sheds light on the formation of MDR S. enterica serovar Indiana with chickens as its potential reservoirs and paves the way to curb its further expansion among food animals. Salmonella enterica (NTS) remains one of the important foodborne pathogens globally -resistant Salmonella spp. were listed by the WHO in 2017 as priority pathogens for which new antimicrobials were urgently needed , or changes in the food chain. Some clonal lineages of MDR Salmonella have shaped the epidemiology of the disease at a global level, as in the case for sequence type 34 (ST34) S. enterica serovar 4,[5],12:i:\u2212, ST313 S.enterica serovar Typhimurium, and ST198 S.enterica serovar Kentucky , extended-spectrum \u03b2-lactams , and macrolides (azithromycin [AZM]) approved by the FDA in the United States to treat infections caused by Salmonella (S. Indiana isolates can express resistance to carbapenems or colistin (CT), the last-resort antimicrobials, suggesting that these bacteria have now become a serious challenge for public health  of DNA gyrase gene (gyrA) and DNA topoisomerase IV gene (parC) conferred FQs resistance, along with plasmid-mediated quinolone resistance (PMQR) genes [oqxAB and aac(6\u2032)-Ib-cr]. In addition, diverse blaCTX-M genes mapped on mobile genetic elements (MGEs) were reported earlier and found to be integrated into the chromosome (as in the case of blaCTX-M-55) (Recent studies investigated the genetic basis underpinning resistance to FQs and extended-spectrum \u03b2-lactamases (ESBLs) in TX-M-55) , 23. HowS. Indiana isolates collected from human and food-related samples in eight provinces of China during 2006 to 2017. Using phenotypic susceptibility data and whole-genome sequencing (WGS) analysis, we determined the prevalence and mechanisms of resistance and identified potential drivers of variation among the AMR profiles described within different lineages.Here, we report on the genomic epidemiology and AMR features of 251 S. Indiana isolates from eight provinces in China, 120 isolates were cultured from clinical samples  taken during 2007 to 2017 and 131 isolates from food-related samples taken during 2006 to 2016 (see in silico multilocus sequence typing (MLST) as S. Indiana ST17 after whole-genome analysis.Out of the 251 confirmed 10.1128/msystems.00253-22.4TABLE\u00a0S1Salmonella Indiana isolates. Download Table\u00a0S1, PDF file, 0.01 MB.Source and basic information of Copyright \u00a9 2022 Du et al.2022Du et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the S. Indiana isolates (P < 0.05) (P < 0.05).Testing of the susceptibility of 251 isolates  to 13 anisolates , with 21isolates . The res < 0.05) . Further10.1128/msystems.00253-22.5TABLE\u00a0S2S. Indiana isolates against ciprofloxacin. Download Table\u00a0S2, DOCX file, 0.02 MB.MIC value related to the PMQR gene and substitution mutations in QRDR of 251 Copyright \u00a9 2022 Du et al.2022Du et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the n\u2009=\u200911) had S83F and D87N amino acid substitutions in GyrA along with two ParC amino acid substitutions. Furthermore, five PMQR genes were detected, including aac(6\u2032)-Ib-cr (n\u2009=\u2009146), oqxAB (n\u2009=\u200968), qnrS1 (n\u2009=\u20092), qnrD (n\u2009=\u20091), and qepA (n\u2009=\u20091). PMQR genes aac(6\u2032)-Ib-cr and oqxAB coexisted in 54 isolates  and were only detected with 4 amino acid substitutions in GyrA and ParC, with MICs of ciprofloxacin ranging from 8 to 256\u2009mg/L (qnrS1 and oqxAB coexisted only in two food-related isolates. Of the 97 isolates possessing amino acid substitutions in GyrA (S83F and D87G) and ParC (T57S and S80R), the detection rate (47.5% [57/120]) in human-derived isolates was higher than that identified in food-related isolates (30.5% [40/131]) (P < 0.01). However, of the 141 isolates possessing amino acid substitutions in GyrA (S83F and D87N) and ParC (T57S and S80R), the detection rate in human isolates (47.5% [57/120]) was lower than that in food-related isolates (64.1% [84/131]) (P < 0.01).The genomes of 251 isolates were screened for known genetic determinants of AMR, including mobile resistance genes and mutations within QRDRs . For mut256\u2009mg/L , while q10.1128/msystems.00253-22.6TABLE\u00a0S3S. enterica serovar Indiana isolates. Download Table\u00a0S3, DOCX file, 0.02 MB.Comparisons of the resistance genes of human and food-related Copyright \u00a9 2022 Du et al.2022Du et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the blaCTX-M subtypes were identified among the 138 ESBL-producing isolates  distributed in different provinces, including blaCTX-M-65 , blaCTX-M-14 , blaCTX-M-55 , blaCTX-M-15 , blaCTX-M-27 , and blaCTX-M-123 . The three dominant subtypes blaCTX-M-65/55/14 were detected in human and food-related isolates, while blaCTX-M-15/123 was found only in human isolates. Moreover, the prevalence rate of blaCTX-M-65 among isolates from \u22641-year-old patients (42.6% [20/47]) was significantly higher than that among isolates from other patients (11.1% [8/72]) (P < 0.01), which is in line with the phenotypic characteristics of isolates from these two patient groups. In general, most of the blaCTX-M variants have been detected in humans and chickens.Six mph(A) gene, which is highly associated with azithromycin resistance, had the highest detection rate of 33.5% . The mph(E) (n\u2009=\u20097), msr(E) (n\u2009=\u20097), erm(42) (n\u2009=\u20091), and erm(B) (n\u2009=\u20091) genes were also detected.For macrolides, the 26). The pairwise co-occurrence matrix of AMR genes is shown in arr-3 (conferring resistance to rifamycin), catB3 (conferring resistance to chloramphenicols), aac(6\u2032)-Ib-cr (conferring resistance to aminoglycosides), and blaOXA-1 (conferring resistance to penicillins), which co-occurred in 58.2% of genomes (146/251); the combination of arr-3, catB3, aac(6\u2032)-Ib-cr, and blaOXA-1 occurred with sul1 (sulfonamides) in 50.6% (127/251), aac(3)-Iva (aminoglycosides)/aph(4)-Ia (aminoglycosides)/tet(A) (tetracycline) in 45.8% (115/251), floR (chloramphenicols) in 43.4% (109/251), and sul2 (sulfonamides) in 36.7% (92/251) of genomes. These co-occurring genes were formed into resistance regions comprising IS26 and tet(A), sul1, arr-3, catB3, blaOXA-1, aac(6\u2032)-Ib-cr, aac(3)-Iva, aph(4)-Ia, and sul2 located in ps15D023-IncHI2, ps12177-CTX, pIndS104-CTX, and ps11011-CTX (aph(3\u2033)-Ib, aph(6)-Id, and aadA5 co-occurred in 19.1% (48/251) of genomes; the combination of aph(3\u2033)-Ib, aph(6)-Id, and aadA5 occurred with oqxAB in 14.7% (37/251), fosA in 12.0% (30/251), blaTEM-1 in 11.5% (29/251), and blaCTX-M-65 in 9.2% (23/251) of genomes. These were caused by the resistance region IS26-aph(3\u2033)-Ib-aph(6)-Id-sul2-IS26 co-occurring with other resistance genes. Salmonella genomic island 1 (SGI1), the widely reported original chromosomal integron containing an antibiotic resistance gene cluster and identified in several S. enterica serovars . Download FIG\u00a0S2, TIF file, 0.9 MB.Sequence comparison of IncHI2 plasmids in Copyright \u00a9 2022 Du et al.2022Du et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the blaCTX-M-positive isolates, 90 (65.2%) could be classified by the locations of blaCTX-M in the genome from the fragmented short-read assemblies. blaCTX-M-14 in 14 isolates (42.4% [14/33]) and blaCTX-M-55 from 9 isolates (30.0% [9/30]) were located on the chromosome. blaCTX-M-15 from 9 isolates (100% [9/9]) and blaCTX-M-65 from 58 isolates (96.7% [58/60]) were carried by plasmids. In addition, 76 blaCTX-M-positive isolates were from humans and 62 were food related, accounting for 63% (76/120) and 47% (62/131) of the two groups, respectively. Although the positive ratio of blaCTX-M was significant higher in human isolates (P < 0.05), among the six subtypes, only the positive ratio of blaCTX-M-15 was significant higher in human isolates .Among the 138 blaCTX-M among representative nonclonal isolates from different sources, genome sequences of five isolates  were successfully completed. The IndS102 and s15D023 isolates separately carried blaCTX-M-14 and blaCTX-M-55 on their chromosomes within different genetic contexts. Another three isolates carried blaCTX-M on IncHI2 plasmids ranging from 200 to 322 kbp and sharing similar core structures  with the specific genetic context IS26-ISEcp1-blaCTX-M-65-IS903B located in an MDR region . BLASTn analysis demonstrated pIndS104-CTX was most similar (99.99% identity at 100% coverage) to a locus on the chromosome of Chinese blaCTX-M-65-positve S. Indiana SI43, obtained from a spiral shell in China in 2010 , indicating pIndS104-CTX may have the ability to recombine with the chromosome entirely or evolve from ancestor clones like SI43. ps11011-CTX was 263,731\u2009bp in size and possessed 783 predicted coding sequences. ps11011-CTX showed high similarity to ps12177-CTX , originating from a human sample from China in 2006, and also to Escherichiacoli plasmids pE648CTX-M-65  and pECJS-B60-267 , as well as Klebsiellapneumoniae plasmid pHNHF1_NDM-99 . ps12177-CTX was 200,106\u2009bp in size (48.61% GC content), and it encoded an MDR region different from that described in ps11011-CTX , which might have resulted from the truncation of ISEcp1 by IS26.Although genetic contexts of  by IS26 . ISEcp1 mph(A) was positive in the chicken isolate IndS104 and was located on plasmid pIndS104_3_29k, a typical IncX1 plasmid of size 29,056\u2009bp. It was found to be homologous  to six similar plasmids that ranged from 34,764 to 222,492\u2009bp among Salmonella spp. and E. coli, as well as a chromosomal locus in a human S. Indiana isolate SI111 (CP050764) (mph(A)-bearing IncX1 plasmids were hypothetically mobilizable and could move into the chromosome via insertion sequences such as IS26. The typical IS26-mphR(A)-mrx-mph(A)-IS26 transposition unit was embedded in the IncX1 plasmid and other MDR plasmids such as IncHI2 (S. Indiana SI111 chromosome demonstrated a variant of mph(A)-bearing IncX1 plasmid recombined into the SI85 chromosome by IS26 to generate the chromosomal IncX1 segment in SI111 (26 in the transmission of mph(A) among plasmids and chromosomes. Detailed analysis of mph(A)-bearing contigs in the 84\u2009mph(A)-positive isolates showed that the core structure mphR(A)-mrx-mph(A) (n\u2009=\u200984) and seven additional different core structures were prevalent among these isolates (mphR(A)-mrx-mph(A) were not identified because of short fragmented assembled contigs based on Illumina short-read data.Among the five isolates  with complete genome sequences, P050764) , which is IncHI2 . Linear in SI111 . This hiisolates . However10.1128/msystems.00253-22.3FIG\u00a0S3mph(A)-harboring S. Indiana isolates. Seven different genetic structures are listed above. The numbers of isolates of the seven types are 1, 1, 6, 1, 11, 63, and 1, respectively. Boxes or arrows represent the ORFs. Red arrows represent the mph(A) gene. Blue arrows indicate mobile elements. mphR(A) and mrx are shown by green arrows. Light gray arrows represent the resistance gene and hypothetical protein. A triangle represents the truncated gene. Download FIG\u00a0S3, TIF file, 1.3 MB.Genetic environment comparison of 84 Copyright \u00a9 2022 Du et al.2022Du et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the S. Indiana emerged within the study period and were transmitted to those enrolled provinces. All 251 isolates harbored the mutations on gyrA and parC associated with the FQ resistance. According to core-genome-based phylogenies, we divided them into six lineages : lineage 1 included seven isolates with D87G in GyrA and T57S in ParC, lineage 2 included two isolates with S83F in GyrA and T57S in ParC, and lineage 3 included four isolates with S83F in GyrA and T57S and S80R in ParC. The other three later lineages with 4- amino acid substitutions were the dominant groups. Lineage 4 (n\u2009=\u200949) and lineage 5 (n\u2009=\u200948) had S83F and D87G in GyrA and T57S and S80R in ParC, while lineage 6 (n\u2009=\u2009141) had S83F and D87N in GyrA and T57S and S80R in ParC, which was the most common lineage (141/251 [56.2%]). Moreover, in the early stages of quinolone resistance evolution, most of the isolates with double-amino-acid substitutions in GyrA (D87G [lineage 1] or D87F [lineage 2]) and ParC (T57S) were susceptible to ciprofloxacin, except in two isolates that also carried PMQR genes (oqxAB and qnrS1), and the isolates from lineage 3 with three amino acid substitutions in GyrA (S83F) and ParC (T57S and S80R) exhibited low MIC values of ciprofloxacin , 76.7% (23/30), and 70% (42/60) of the isolates harboring each genotype, respectively (mph(A) appeared most frequently in lineage 6 (88.1% [74/84]), followed by lineage 5 (9.5% [8/84]), and only occurred once in lineage 2 and lineage 3, respectively. Furthermore, co-occurrence of mph(A) with diverse ESBL genes was observed . Both IncN and IncX1 replicons were dominantly distributed in lineage 6, with prevalence of 75.3% (55/73) and 98.5% (64/65). Based on the complete genome sequences of IndS104, mph(A) was located in an IncX1 plasmid. In addition, among the 251 isolates, mph(A) and IncX1 were both detected in 55 isolates belonging to lineage 6, and both were negative in 157 isolates. The distribution of mph(A) and IncX1 displayed a significant correlation with a calculated coefficient of 0.64 . The isolates carrying IncX1 plasmids might be more likely to carry mph(A) .Based on the 2,904 core genome single nucleotide polymorphisms (SNPs) obtained from 251 genomes, we performed phylogenic analysis and displayed the phylogenetic relationships of sequenced isolates . Multipllineages , which ifloxacin . In contfloxacin . Of noteectively . MoreoveSalmonella have changed significantly in recent years as MDR Salmonella strains have become prevalent among various serovars, such as S. 1,4,[5],12:i:\u2212, S. Kentucky, and S. enterica serovar London  -Ib-cr was the dominant genotype, with the carriage rate increasing from 20% in 2006 to 65% in 2017  of the ciprofloxacin-resistant Salmonella strains of various serovars, the oqxAB and aac(6\u2032)-Ib-cr combination was the predominant one in our study, which was located in the IncHI2 plasmid. This gene combination was also located in a nonconjugative plasmid, pCFSA244-1, mainly transmitted among S. Typhimurium isolates, and qnrS2-aac(6\u2032)-lb-cr-oqxAB elements were also found on the chromosome in S. enterica serovar Derby (oqxAB-aac(6\u2032)-Ib-cr-bearing IncHI2 plasmid found in S. Indiana was mobilizable by conjugation  . Furtherservoirs . Since 2servoirs . Our who E. coli . Recent  E. coli . It has  strains . This mijugation . The hig studies . This mi studies .S. Indiana, contributing to their cefotaxime resistance phenotypes   variant of blaCTX-M-15, blaCTX-M-55, possessing enhanced cephalosporin-hydrolyzing activity, was both prevalent in both isolates from humans and those from food-related samples, particularly in lineage 6, indicating potential higher adaptability of blaCTX-M-55. IncHI2 plasmids were typical MDR plasmids positive for blaCTX-M, mcr-1, class 1 integrons, and oqxAB, similar to those found in other Enterobacteriaceae  . Most blaCTX-M-55 genes in S. Indiana are located on the chromosome, which is different from its plasmid origin in other serovars, such as S.enterica serovars Enteritidis, Goldcoast, Typhimurium, and Choleraesuis  . FurtherE. coli) , 41. Furplasmids . The blaosporins . The occeraesuis \u201346.S. Indiana isolates harboring the plasmid-borne mph(A), mph(E), erm(42), and erm(B) genes. Moreover, in the present study, 24% of isolates were coresistant to azithromycin, ciprofloxacin, and cefotaxime, which is considerably higher than the levels reported in a Chinese national surveillance study (0.2%) from 2005 to 2011 (mph(A) and mph(E), raising concern about the spread of resistance among bacteria (mph(A)-positive isolates were mostly grouped into lineages 5 and 6 coexisting with diverse ESBL genes. This feature is highly related to the IncX plasmid, which has played an important role in the spreading of resistance genes, such as blaNDM (IncX3) and mcr-1 (IncX4 and IncX2) (\u2013Macrolide (azithromycin) resistance was determined in 36.3% 91/251) of  to 2011  and a Ch to 2011 . In the bacteria . In our d IncX2) \u201351. More of  to 2 IncX2) \u2013.Salmonella. The high prevalence of IncHI2 plasmids in S. Indiana is similar to that reported for S. Typhimurium  and food  collected previously by the National Health Commission Key Laboratory of Food Safety Risk Assessment from four provinces in China during 2006 to 2016. In total, isolates from eight provinces were included. Clinical fecal samples were enriched in selenite brilliant green sulfa enrichment broth for 18 h at 37\u2009\u00b1\u20091\u00b0C, and anatomical site samples  were enriched on blood agar plates for 18 h at 37\u2009\u00b1\u20091\u00b0C. Clinical sample isolates were purified as described previously , along with amplification of the invA gene by PCR. All isolates were identified to the serogroup level and then serotyped by slide agglutination with commercial Salmonella antisera  following the Kauffmann-White scheme at the National Health Commission Key Laboratory of Food Safety Risk Assessment in Beijing, China.A total of 251 confirmed eviously . S. enteuidebook . The isoS. Indiana isolates was performed by the agar dilution method and interpreted according to 2018 Clinical and Laboratory Standards Institute (CLSI) guidelines (https://eucast.org/). The antimicrobial susceptibilities of the following antimicrobials were assessed: ampicillin (AMP), cefotaxime (CTX), cefotaxime-clavulanic acid (CTX-CLA), chloramphenicol (CHL), ciprofloxacin (CIP), nalidixic acid (NAL), gentamicin (GEN), streptomycin (STR), imipenem (IPM), meropenem (MEM), tetracycline (TET), azithromycin (AZM), sulfonamide (SUL), and colistin (CT). The MICs were calculated. Multidrug resistance was defined as resistance to three or more classes of antimicrobials. Escherichia coli ATCC 25922 and Klebsiella pneumoniae ATCC 700603 were used as the quality control strains.Antimicrobial susceptibility testing (AST) of the idelines  and the in silico multilocus sequence typing (MLST) scheme was used to subtype the isolates using BLASTn and seven housekeeping genes: aroC, dnaN, hemD, hisD, purE, sucA, and thrA  and then sequenced using an Illumina HiSeq 2500 platform  to generate 150-bp paired-end reads from a library with an average insert size of 500 bp. Raw reads were filtered to remove low-quality reads by fastQC  and then62\u2013and thrA .blaCTX-M were determined after whole-genome sequence analysis. blaCTX-M-containing contigs were examined for plasmid Inc types using PlasmidFinder  was used as the reference in phylogenetic analysis. Illumina reads were mapped to the reference genome using Bowtie 2 v2.2.8, with single nucleotide polymorphisms (SNPs) identified using Samtools v1.9, and the data were combined as described previously . Multiple alignments of core genomes identified from the pairwise alignments with S. Indiana D90 were used as the input for Gubbins to detect and remove recombination sites (The complete genome sequence of eviously . The higon sites . Phylogeon sites  and seleon sites . The conon sites . The phyon sites .PRJNA850394.The genome sequences in this study were deposited into the National Center for Biotechnology Information database under BioProject accession no."}
+{"text": "Based on the pharmacokinetic-pharmacodynamic (PK-PD) breakpoints, cefepime-zidebactam inhibited 98.5% of Enterobacterales and 98.9% of Pseudomonas aeruginosa isolates, respectively. Against carbapenem-resistant and difficult-to-treat resistant Gram-negative bacilli, cefepime-zidebactam demonstrated better activity against Enterobacterales  and P. aeruginosa . Among the 379 carbapenem-resistant Enterobacterales isolates, the most common carbapenemase genes detected were blaKPC-2 (64.1%) and blaNDM (30.9%). Cefepime-zidebactam showed an MIC90 of \u22642\u2009mg/L for 98.8% of blaKPC-positive isolates and 89.7% of blaNDM-positive isolates. Ceftazidime-avibactam also showed efficient in vitro activity against Enterobacterales (93.6%) and P. aeruginosa (87.7%). Ceftazidime-avibactam was active against 97.5% of blaKPC-positive isolates and 100% of blaOXA-232-positive isolates. Cefepime-zidebactam inhibited 97.3% of Acinetobacter baumannii isolates with an MIC50/90 of 16/32\u2009mg/L. Our study systematically evaluated the in vitro activities of these new BLBLIs against a variety of Gram-negative bacilli, provided preclinical data for the approval of these BLBLIs in China, and supported cefepime-zidebactam and ceftazidime-avibactam as potential efficient therapies for infections caused by carbapenem-resistant Enterobacterales (CRE), carbapenem-resistant P. aeruginosa (CRPA), and DTR isolates.Novel \u03b2-lactam\u2013\u03b2-lactamase inhibitor combinations (BLBLIs) are in clinical development for the treatment of infections caused by carbapenem-resistant and difficult-to-treat resistant (DTR)  Gram-negative bacilli. This study evaluated the IMPORTANCEEnterobacterales, Pseudomonas aeruginosa, and Acinetobacter baumannii are the most common Gram-negative bacilli to cause nosocomial infections throughout the world. Due to their large public health and societal implications, carbapenem-resistant A. baumannii (CRAB), carbapenem-resistant P. aeruginosa (CRPA), and carbapenem-resistant and third-generation-cephalosporin-resistant Enterobacteriaceae were regarded by the World Health Organization (WHO) as a global priority for investment in new drugs in 2017. The present study showed the potent in vitro activity of these novel BLBLIs and other comparators against Gram-negative bacillus isolates, including carbapenem-resistant or difficult-to-treat resistant phenotypes. Polymyxins, tigecycline, and ceftazidime-avibactam (except for blaNDM-positive isolates) were available for the treatment of infections caused by CRE isolates. Currently, cefepime-zidebactam and other BLBLIs have not yet been approved for use in China. Here, our study aimed to evaluate the in vitro activities of BLBLIs against Gram-negative bacillus isolates, especially CRE, before clinical use. Carbapenem-resistant Gram-negative bacilli have rapidly increased worldwide in the last decade, which is related to the emergence and prevalence of plasmid-mediated extended-spectrum \u03b2-lactamases (ESBLs), AmpC cephalosporinases, and carbapenemases among these isolates, conferring resistance to \u03b2-lactam antibiotics, and make difficulties in empirical treatment for clinicians  were carbapenem-resistant Enterobacterales (CRE), including K. pneumoniae , E. coli , and Enterobacter cloacae .Among the tested P. aeruginosa (CRPA) and carbapenem-resistant A. baumannii (CRAB), respectively, and 11.9%  of Enterobacterales isolates and 8.6% (65/756) of P. aeruginosa isolates were difficult-to-treat resistant (DTR) isolates.For glucose-nonfermenting bacteria, 228/756 (30.2%) and 471/630 (74.8%) were carbapenem-resistant in vitro activities of cefepime-zidebactam, ceftazidime-avibactam, cefepime-tazobactam, ceftolozane-tazobactam, and other comparator agents against 4,042 clinical isolates are summarized in Tables 1Enterobacterales isolates  with an MIC50/90 of 0.06/1\u2009mg/L. A total of 98.5% of isolates were inhibited at the provisional cefepime-zidebactam pharmacokinetic-pharmacodynamic (PK-PD) breakpoint (\u22648\u2009mg/L), with 24 E. coli, 4 K. pneumoniae, 7 Proteus rettgeri, 3 P. mirabilis, 1 E. cloacae, and 1 Serratia marcescens isolates showing MICs of \u226516\u2009mg/L among the various genera of Enterobacterales. Besides cefepime-zidebactam, ceftazidime-avibactam was also active against all Enterobacterales clinical isolates with an MIC50/90 of 0.25/4\u2009mg/L. Among 171 ceftazidime-avibactam-resistant isolates, cefepime-zidebactam showed an MIC of 8\u2009mg/L or lower against 84.1% of the tested isolates (data not shown). Apart from cefepime-zidebactam and ceftazidime-avibactam, tigecycline (96.5% susceptible) and amikacin (90.4% susceptible) also displayed potent activity against Enterobacterales. The rate of susceptibility to cefepime-tazobactam was 85.8%, similar to those for polymyxin B (81.4% susceptible) and meropenem (85.6% susceptible), which showed good activity against the tested isolates. More than 60% of the Enterobacterales isolates were susceptible to ceftolozane-tazobactam (74.2% susceptible), imipenem (74.6% susceptible), piperacillin-tazobactam (75.5% susceptible), cefoperazone-sulbactam (69.8% susceptible), and ceftazidime (60.5% susceptible). The following other comparator agents showed limited activity: cefepime (55.3% susceptible), ceftriaxone (46.1% susceptible), aztreonam (54.7% susceptible), ciprofloxacin (42.1% susceptible), levofloxacin (48.1% susceptible), and trimethoprim-sulfamethoxazole (53.7% susceptible) (The eptible) .P. aeruginosa were highly inhibited by cefepime-zidebactam with an MIC50/90 of 2/8\u2009mg/L at a PK-PD breakpoint of \u226432\u2009mg/L (98.9% susceptible). The rate of susceptibility of P. aeruginosa to cefepime-zidebactam was similar to or slightly higher than those for ceftazidime-avibactam (87.7% susceptible), ceftolozane-tazobactam (90.2% susceptible), polymyxin B (95.6% susceptible), and amikacin (95.4% susceptible)  . The rat50/90 value of cefepime-zidebactam against 630 A. baumannii isolates was 16/32\u2009mg/L , blaNDM-positive (n\u2009=\u2009117), blaIMP-positive (n\u2009=\u20098), blaOXA-232-positive (n\u2009=\u20097), blaVIM-positive (n\u2009=\u20091), as well as carbapenemase-negative (n\u2009=\u20093) isolates  and polymyxin B (98.2% versus 96.9%), whereas the rates of susceptibility were 76.3% for ceftolozane-tazobactam and 68% for ceftazidime-avibactam as the most active comparators. Except for imipenem and meropenem, CRPA isolates were moderately resistant to other \u03b2-lactams, aztreonam, and fluoroquinolones, with susceptibility rates of 30% to 50%.90 values of >32\u2009mg/L.For CRAB, cefepime-zidebactam, tigecycline, and polymyxin B showed high susceptibility rates of 96.6%, 88.5%, and 96.6%, respectively, and amikacin and trimethoprim-sulfamethoxazole showed limited activity, with susceptibility rates of 18.7% and 23.1%, respectively. The MICs of other agents were higher, with MICEnterobacterales isolates with an MIC50/90 of 1/4\u2009mg/L at \u22648\u2009mg/L, and 74.7% of DTR Enterobacterales isolates were susceptible to ceftazidime-avibactam with an MIC50/90 of 2/>64\u2009mg/L. Only tigecycline (95.9% susceptible) and polymyxin B (92.4% susceptible) displayed greater in vitro activity than cefepime-zidebactam and ceftazidime-avibactam against all DTR Enterobacterales isolates  demonstrated greater in vitro activity than the above-described agents against DTR P. aeruginosa isolates  were blaNDM-5 positive, 12.4% (47/379) were blaNDM-1 positive, 2.1% (8/379) were blaIMP positive, 1.8% (7/379) were blaOXA-232 positive, and 0.3% (1/379) were blaVIM positive, respectively. Additionally, blaKPC-2 was mainly detected in K. pneumoniae , S. marcescens , Citrobacter freundii , and Morganella morganii . The highest prevalences of blaNDM-5 were 82.5% (33/40) in E. coli and 55.6% (5/9) in Klebsiella aerogenes isolates. blaNDM-1 was the predominant type of carbapenemase gene among E. cloacae  and P. rettgeri  isolates.In this study, 99.2% (376/379) of the CRE isolates had a single carbapenemase gene, and only 3 isolates were negative for all five common carbapenemase genes . Among tP. aeruginosa, and A. baumannii, which has substantially increased morbidity and mortality rates and caused nosocomial outbreaks , European Committee on Antimicrobial Susceptibility Testing (EUCAST), or U.S. Food and Drug Administration (FDA) clinical breakpoints of cefepime-zidebactam, so according to its PK-PD breakpoint (\u226432\u2009mg/L), P. aeruginosa had a rate of susceptibility to cefepime-zidebactam of 99.6%, whereas it was 98.9% in our study. The potent activity of cefepime-zidebactam against CRE, P. aeruginosa, and A. baumannii isolates harboring carbapenemase genes has been previously reported. In another study of a worldwide surveillance program, Sader et al. (n\u2009=\u2009153) had cefepime-zidebactam MICs of \u22648\u2009mg/L, similar to the results of this study (98.5%).In this study, 98.5% of \u226432\u2009mg/L , respect\u226432\u2009mg/L , the autr et al.  reportedE. coli, 0.6% to 1.0% for Enterobacter spp., 0.6% to 3.0% for Klebsiella spp., and 8.4% for P. aeruginosa (data not shown). In our study, we observed slightly higher DTR rates of 1.2% for E. coli, 0.04% to 0.5% for Enterobacter spp., 9.3% for Klebsiella spp., and 8.6% for P. aeruginosa. The differences in DTR rates between the 2 studies may reflect the characteristics of the strains among different regions and different specimen sources. Kadri et al. reported that mortality was significantly higher for DTR than for carbapenem-resistant, extended-spectrum-cephalosporin-resistant, or fluoroquinolone-resistant infections  (96% to 98.2%) and DTR isolates (96.9% to 97.2%).The DTR phenotype, a novel category in the study of Gram-negative bacteremia, focuses on treatment-limiting resistance to all first-line agents. The DTR phenotype was defined as an isolate that tests not susceptible (intermediate or resistant) to all \u03b2-lactam categories, including carbapenems and fluoroquinolones, and it was demonstrated that isolates that were not susceptible to first-line agents were associated with increased patient mortality and clinical failure. Karlowsky et al.  studied fections . The in blaKPC- or blaOXA-48-positive isolates. blaKPC-positive isolates showed a low rate of resistance to ceftazidime-avibactam (2.5%), but the majority (87.5% to 100%) of blaNDM-positive isolates were resistant to ceftazidime-avibactam. The major resistance mechanisms that confer reduced susceptibility to ceftazidime-avibactam are as follows: the production of metallo-\u03b2-lactamases (MBLs) such as NDM, VIM, or IMP; blaKPC variants; and the transposition of KPC with porin deficiency  and P. aeruginosa  isolates producing important \u03b2-lactamases, including MBLs (except for ceftazidime-avibactam), KPCs, and OXA-232, for which treatment agents are limited. The results from this study support the use of cefepime-zidebactam and ceftazidime-avibactam as potential therapies for infections caused by CRE, CRPA, and DTR isolates.We studied a recent nationwide collection of Gram-negative bacilli and observed that new BLBLIs, especially cefepime-zidebactam and ceftazidime-avibactam, demonstrated potent The study protocol was approved by the Institutional Review Board of Huashan Hospital, Fudan University (no. 2019-460).Klebsiella pneumoniae (n = 979), Escherichia coli (n = 900), P. aeruginosa (n = 756), A. baumannii (n = 630), Enterobacter cloacae (n = 172), Proteus mirabilis (n = 119), Serratia marcescens (n = 118), K. aerogenes (n = 103), Morganella morganii (n = 89), Citrobacter freundii (n = 84), Proteus vulgaris (n = 51), Proteus rettgeri (n = 29), and Klebsiella oxytoca (n = 12). Among the tested clinical isolates, 23.6% of the isolates were isolated from patients in the intensive care unit, followed by outpatient and emergency departments (18.5%), urology surgery (6.7%), respiratory medicine (5.6%), neurosurgery departments (4.2%), and other departments. A total of 33.9% of the tested isolates were isolated from sputum, followed by urine (22.5%), blood , secreta (7.7%), bronchoalveolar lavage fluid (3.9%), pus (2.8%), wound (2.7%), abdominal fluid (2.0%), bile (1.7%), shunt fluid (1.3%), drain (1.2%), and other sources (8.2%). Species identification was performed at each participating site and confirmed by the central laboratory using matrix-assisted laser desorption ionization\u2013time of flight mass spectrometry . Quality control was performed according to Clinical and Laboratory Standards Institute (CLSI) guidelines using E. coli ATCC 25922 and ATCC 35218, K. pneumoniae ATCC 700603, and P. aeruginosa ATCC 27853 for antimicrobial susceptibility testing.The China Antimicrobial Surveillance Network (CHINET) is a multicenter bacterial resistance surveillance program in operation since 2005 in China. In 2019, 46 hospitals in 28 provinces or cities collected up to 4,042 nonduplicate, clinically significant Gram-negative isolates from CHINET, including Enterobacterales . Cefepime-tazobactam MICs were interpreted using provisional breakpoints of \u226416\u2009mg/L for Enterobacterales and P. aeruginosa based on PK-PD studies  in the study. Quality control and test results were interpreted according to 2021 CLSI breakpoints . Difficult-to-treat resistance phenotypes were defined by testing resistance to all tested \u03b2-lactams (including carbapenems and \u03b2-lactamase inhibitor combinations) and fluoroquinolones .Enterobacterales (CRE) isolates were selected for analysis of carbapenemase. The five most common carbapenemase genes  were confirmed for all of the CRE isolates by PCR with specific primers and DNA sequencing, as described previously (Carbapenem-resistant eviously ."}
+{"text": "Deutereulophus Schulz, D.felixsp. nov. and D.daguisiensissp. nov., are described from China. A key to species of Deutereulophus known from China is provided.Two new species of  Deutereulophus Schulz is one of several small genera of the tribe Eulophini, subfamily Eulophinae (Hymenoptera: Eulophidae). Species of the genus are distributed in all zoogeographic regions, except for the Afrotropical region. Currently Deutereulophus contains 23 valid species (D.froudei (Girault), D.interruptus Zhu & Huang, D.marginatus Zhu & Huang, and D.tennysoni (Girault)  , 2002b.Deutereulophus (as Entedonomorpha Girault) was subdivided into two species groups by tennysoni-group and provides a key to all species occurring in China.The genus All specimens were collected by sweeping or yellow-pan trapping, and were dissected and mounted in Canada balsam on slides following the method of Terminology follows the F1\u20133 flagellomeres 1\u20133;MV marginal vein;OOL minimum distance between a posterior ocellus and corresponding eye margin;PMV postmarginal vein;POL minimum distance between posterior ocelli;SMV submarginal vein;STV stigmal vein.NEFU), Harbin, China.All type material is deposited in the insect collections at Northeast Forestry University : SMV = 36; MV = 36; PMV = 12; STV = 16.ing Fig.  2.1\u00d7 as Metasoma .felix, meaning lucky.Named after the Latin adjective Deutereulophusmarginatus, and can be separated using the key given above.The new species is similar to Taxon classificationAnimaliaHymenopteraEulophidae\ufeff08C5DC71-9949-5063-A04A-0D42C97F314Ahttps://zoobank.org/02F37BE0-42A2-4F4D-B908-7AF3845F0280Holotype, \u2640 , China, Hubei Province, Suizhou City, Daguisi National Forest Park, 12. VI. 2012, Guo-Hao Zu and Jiang Liu, by sweeping. Paratypes: 1\u2640 [on slide], same data as holotype.Head and mesosoma black. Vertex with scattered pits. Antennal scrobes smooth. Antenna yellowish-white with F3 dark brown, clava dark brown with apex yellowish-white. Female funicle 3-segmented, clava 4-segmented. Legs yellowish-white with procoxa and profemur dark brown. Face with raised reticulation. Metasoma dark brown. Mesoscutum strongly reticulate with large meshes. Sublateral grooves on mesoscutellum converging and meeting posteriorly. Propodeum with a raised triangular cup-shaped area anteromedially, median carina split and diverging posteriorly.Female. Length 2.4 mm, fore wing length 1.7 mm. Head and mesosoma black. Eyes gray. Ocelli pale yellow. Antenna yellowish-white with F3 dark brown, clava dark brown with apex yellowish-white. Mandibles dark brown. Petiole black. Metasoma dark brown. Legs yellowish-white with procoxa and profemur dark brown. Wings hyaline with veins brown.Head : SMV = 29; MV = 32; PMV = 12; STV = 10.ing Fig.  2.4\u00d7 as Metasoma .Named after the type locality, the Daguisi National Forest Park in Hubei Province.D.malabarensis Narendran, but can be separated from it by the following combination of characters: antenna yellowish-white with F3 dark brown, clava dark brown with apex yellowish-white ; sublateral grooves on mesoscutellum converging and meeting posteriorly ; propodeum with a raised triangular cup-shaped area anteromedially .The new species is similar to"}
+{"text": "Among current e-cigarette users, 14.5% reported that the brand they usually used was Puff Bar, followed by Vuse (12.5%), Hyde (5.5%), and SMOK (4.0%). Approximately one fifth (21.8%) of current e-cigarette users reported \u201csome other brand\u201d as their usual brand.https://stacks.cdc.gov/view/cdc/121630). Among current users of flavored pods or cartridges, the reported flavor types used were fruit (58.4%); menthol (53.9%); candy, desserts, or other sweets (30.3%); and mint (27.6%). Among current users of flavored tanks or mod systems, the reported flavor types used were fruit (69.6%); candy, desserts, or other sweets (47.7%); mint (40.1%); and menthol (35.2%).Among current e-cigarette users overall, 84.9% used flavored e-cigarettes; of these, the reported flavor types, in descending order of use, were fruit (69.1%); candy, desserts, or other sweets (38.3%); mint (29.4%); and menthol (26.6%). A similar pattern was observed among current users of flavored disposable e-cigarettes: fruit (75.2%); candy, desserts, or other sweets (40.4%); mint (29.6%); and menthol (16.7%) (Supplementary Table, In 2022, 2.55 million U.S. middle and high school students currently used e-cigarettes. Most reported using flavored products, and, among those students, approximately seven of 10 used fruit flavors. Disposable products were the most commonly reported device type. Further, among middle and high school students who used e-cigarettes, approximately four in 10 reported frequent use, and approximately one in four reported daily use. The use of tobacco products in any form, including e-cigarettes, by middle and high school students is unsafe. Sustained implementation of comprehensive tobacco prevention and control strategies at the national, state, and local levels,"}
+{"text": "We present a comprehensive data set that describes an anaerobic microbial consortium native to polychlorinated biphenyl (PCB)-contaminated sediments. Obtained from sediment microcosms incubated for 200\u2009days, the data set includes 4 metagenomes, 4 metatranscriptomes (in duplicate), and 62 metagenome-assembled genomes and captures microbial community interactions, structure, and function relevant to anaerobic PCB biodegradation. P < 0.0001).Polychlorinated biphenyl (PCB)-contaminated sediments threaten human and ecological health but often harbor PCB-transforming bacteria that help detoxify sediments \u20133. We es\u2013After 200\u2009days of incubation, DNA was extracted from single slurry samples (2\u2009mL) with a modified DNeasy PowerWater Sterivex kit protocol , and RNAMethanosarcina barkeri (GenBank accession number NZ_CP009530.1) and Methanobacterium subterraneum (GenBank accession number NZ_CP017768.1) were added to further deplete methanogenic archaeal rRNA sequences. RNA was sequenced on a single Illumina NovaSeq 6000 flow cell lane (2 \u00d7 150-bp paired-end reads).High-throughput DNA and RNA sequencing (4 metagenomes and 4 metatranscriptomes in duplicate) was performed at the Iowa Institute of Human Genetics (IIHG) . Indexed DNA libraries, prepared with the KAPA HyperPrep kit  using sheared DNA , were pooled and sequenced on separate lanes of an S Prime NovaSeq 6000 flow cell (2 \u00d7 150-bp paired-end reads). RNA libraries were indexed and rRNA depleted with the stranded total RNA preparation with Ribo-Zero Plus kit . To improve mRNA sequencing efficiency, supplemental rRNA probes developed from Metagenome sequencing yielded 97,279,994 (E2_1), 105,614,978 (E2_2), 102,655,932 (F4_1), and 103,672, 591 (F4_2) raw reads from each bottle. Metatranscriptome sequencing (RNA-seq) yielded 50,403,972 (E2_1 metatranscriptome-A), 60,356,358 (E2_1-B), 49,353,924 (E2_2-A), 45,023,913 (E2_2-B), 54,860,340 (F4_1-A), 47,603,523 (F4_1-B), 55,440,491 (F4_2-B), and 71,530,818 (F4_2-C) raw reads. After trimming and filtering of unassembled sequence reads with Trimmomatic (v0.39) .Dehalococcoides mccartyi) (Metagenome-assembled genomes (MAGs) were obtained by removing low-abundance k-mers from quality-filtered reads with khmer (v3.0.0) , coassem12\u2013ccartyi) , which wccartyi) , is expePRJNA743546. Metagenomic data are available under SRA accession numbers SRX11347095 to SRX11347098. Metatranscriptomic data are available under accession numbers SRX14430540 to SRX14430547. PCB data are available at Iowa Research Online (https://www.doi.org/10.25820/data.006156).Raw data files (24 fastq files) and MAGs are available under BioProject accession number"}
+{"text": "Actinomyces oris strain K20 was isolated from oral apical lesions. Here, we report the complete circular genome sequence of this strain, obtained by means of hybrid assembly using two next-generation sequencing datasets. The strain has a 3.1-Mb genome with 2,636 coding sequences. Actinomyces are ubiquitous in soil as well as human and animal microbiota  using NanoFilt v.2.3.0 , following the manufacturer\u2019s standard protocol. Paired-end (2\u2009\u00d7\u2009156-bp) reads were obtained using a MiSeq instrument (Illumina). The raw sequencing data  were processed using the FASTQ preprocessing program fastp v.0.19.5  (The strain was cultured overnight in heart infusion broth (BD Difco) at 37\u00b0C under aerobic conditions. Genomic DNA (gDNA) was extracted from the culture medium of this strain using the MagAttract high-molecular-weight (HMW) DNA kit . The obtained gDNA was subjected to long- and short-read sequencing. Long-read sequencing was performed using the GridION X5 sequencing platform ; 1.0\u2009\u03bcg unfragmented gDNA was used for library construction using a ligation sequencing kit SQK-LSK109; ONT). The prepared library was applied to an R9.4.1 flow cell . The base calling of long-read sequences using Dogfish v.0.9.6-3 generated 114,245 reads (1.1\u2009Gb), with an ; ONT. Thde novo genome assembly, the remaining long- and short-read data were processed using the Unicycler v.0.4.4 pipeline (https://dfast.ddbj.nig.ac.jp/), predicted 2,636 coding sequences, 9 rRNA genes, and 52 tRNA genes.For complete pipeline . Assemblpipeline . BlobToopipeline  was usedpipeline  until nopipeline , provideA. oris strain K20 has been deposited at DDBJ/EMBL/GenBank under the accession number AP025590. The associated BioProject and BioSample accession numbers are PRJDB13022 and SAMD00442649, respectively. The SRA accession numbers are DRR351395 (Illumina) and DRR351396 (Nanopore).The complete genome sequence of"}
+{"text": "We found that tricalcium phosphate-solubilizing bacteria, phytate-degrading bacteria, and gcd and bpp abundances were more abundant in silt plus clay  than in macroaggregate (250 to 2000\u2009\u03bcm) and microaggregate (53 to 250\u2009\u03bcm). Fertilization treatment and aggregate fractionation showed distinct effects on PSB number and P-cycling-related gene abundance. We found significantly negative correlation between gcd gene abundance and tricalcium phosphate-solubilizing bacterial number (Col-CaP) and dramatically positive correlation between bpp gene abundance and phytate-degrading bacterial number (Col-Phy). P fractions were responsible for PSB number and P-cycling-related gene abundance. The isolated Pseudomonas sp. strain PSB-2 and Arthrobacter sp. strain PSB-5 exhibited good performances for solubilizing tricalcium phosphate. The inoculation of Pseudomonas sp. PSB-2 could significantly enhance plant fresh weight, plant dry weight, and plant height. Our results emphasized distinct distribution characteristics of PSB and P-cycling-related genes in soil aggregates and deciphered a close linkage between PSB number and P-cycling-related gene abundance. Our findings might guide the isolation of PSB from agricultural soils and provide a candidate plant-growth-promoting bacterium for agro-ecosystems.Deciphering distribution patterns of phosphate-solubilizing bacteria (PSB) and phosphorus-cycling-related genes in soils is important to evaluate phosphorus (P) transformation. However, the linkage between PSB number and P-cycling-related gene abundance in soils, especially soil aggregates, remains largely unknown. Here, we estimated the numbers of PSB and abundances of P-cycling-related genes (i.e., IMPORTANCE Phosphate-solubilizing bacteria are responsible for inorganic P solubilization and organic P mineralization. Elucidating the linkage between phosphate-solubilizing bacterial number and P-cycling-related gene abundance is important to isolate plant-growth-promoting bacteria for agro-ecosystems. Our findings reveal differentiating strategies of phosphate-solubilizing bacteria in soil aggregates, and the deciphered P fractions show strong effects on distribution patterns of phosphate-solubilizing bacteria and P-cycling-related genes. Additionally, we isolated phosphate-solubilizing bacteria with good plant-growth-promoting ability. This study enriches our knowledge of P cycling in soil aggregates and might guide the production and management of farmland. Prenic acid . In contnic acid , 14. Phynic acid , 15. Phyin soils . ConsequAcinetobacter, Citrobacter, Massilia, and Pseudomonas) (21\u2013Bacillus)   , 21\u201324, acillus) . For insosum L.) . Consequosum L.) , 25, 26.gcd and bpp), (ii) estimate abiotic  and biotic  factors on numbers of PSB, and (iii) decipher the performance of PSB for promoting plant growth. Considering that P availability affects the abundances of P-cycling-related genes  circulation , 27 and 3\u201313\u2013ed genes , 30, we ed genes , 17, 31,5 to 8.77\u2009\u00d7\u2009108 CFU/g soil) was significantly higher in NPK  and N  fertilization treatments than that in CK (control without fertilizer), M (swine manure), and MN (combined swine manure and N fertilizer) fertilization treatments (P\u2009<\u20090.05). The phytate-degrading bacterial number (Col-Phy) (4.0\u2009\u00d7\u2009104 to 3.27\u2009\u00d7\u2009108 CFU/g soil) was remarkably higher in N and MN fertilization treatments than that in CK, M, and NPK fertilization treatments (P\u2009<\u20090.05). In five fertilization treatments, the Col-CaP and Col-Phy were basically slightly higher in silt+clay than that in macroaggregate and microaggregate  rather than aggregate fractionation  showed a significant effect on PSB number (Col-CaP and Col-Phy) (P\u2009<\u20090.05) but the opposite for MN fertilization treatment (P\u2009<\u20090.05). Col-CaP was slightly higher than Col-Phy in the same soil aggregate with N fertilization treatment (P\u2009>\u20090.05). These results indicated different distribution patterns of PSB in soil aggregates with different fertilization treatments.The numbers of PSB represented by CFU varied in different soil aggregates . The triCol-Phy) . Additiogcd gene (4.32\u2009\u00d7\u2009106 to 2.20\u2009\u00d7\u2009107 copies/g soil) was more abundant in M and MN fertilization treatments than in other fertilization treatments (P\u2009<\u20090.05), whereas the bpp gene (1.99\u2009\u00d7\u2009105 to 3.42\u2009\u00d7\u2009106 copies/g soil) showed no significant difference among five fertilization treatments (P\u2009>\u20090.05). Basically, gcd and bpp were more abundant in silt+clay than in macroaggregate and microaggregate with the same fertilization treatment  and aggregate fractionation  showed significant effects on abundances of P-cycling-related genes (gcd-harboring bacteria were significantly higher than that of bpp-harboring bacteria in the same soil aggregate with the same fertilization treatment (P\u2009<\u20090.01). These results reflected that fertilization treatment and aggregate fractionation might influence the distribution and abundance of gcd-harboring bacteria and bpp-harboring bacteria.The abundances of P-cycling-related genes varied in different soil aggregates . The gcdreatment . Consequed genes . In addigcd gene abundance was significantly negatively correlated with Col-CaP  (P\u2009<\u20090.05 or P\u2009<\u20090.01 or P\u2009<\u20090.001), whereas only OP was significantly correlated with Col-Phy (P\u2009<\u20090.05). Similarly, nonphosphorus nutrients  and P fractions  were significantly positively correlated with both gcd and bpp gene abundances (P\u2009<\u20090.05 or P\u2009<\u20090.01 or P\u2009<\u20090.001) (Linear regressions reflected that and bpp) . Nonphos<\u20090.001) . Accordi<\u20090.001) . Additiogcd abundance, bpp abundance, Col-CaP, and Col-Phy . Additionally, soil TC and P fractions, rather than gcd abundance, showed strong indirect and direct effects on Col-CaP, respectively  were identified as Bacillus, Pseudomonas, Massilia, Citrobacter, Arthrobacter, and Acinetobacter genera according to a phylogenetic tree based on 16S rRNA gene sequences  and soluble P levels of PSB-2 cultured in liquid National Botanical Research Institutes phosphate growth medium (NBRIP) media were significantly higher than that for PSB-5 at the same period , and growth tended to be stable after day 5 (P\u2009>\u20090.05). The soluble P levels of PSB-5 displayed a fast increase during 5\u2009days and presented slight fluctuation after day 5. In contrast, soluble P levels of PSB-2 exhibited significantly fast increases during 8\u2009days   affect abundances of P-cycling-related genes  \u20138, 34. Fcd genes . The abuicient P , 39, 40.icient P  and difficient P . Additioicient P . These ricient P , 43, 44,icient P , 14, 15.icient P . This phicient P .bpp gene abundance and phytate-degrading bacterial number, which is expected, and this might be because phytate is a relatively stable compound   , 16, 47.rganisms , 34, whirganisms . Organicsing CO2 , which fironment , 41, 50.ironment . Consequ studies , 42; to Pseudomonas sp. PSB-2 exhibited good performance for P-solubilizing, and the best P-solubilizing level was 59.98\u2009mg/L. The solubilizing capacity for Ca3(PO4)2 by Pseudomonas sp. PSB-2 is higher than that for Pantoea dispersa Cav.cy3 (<50\u2009mg/L)  (Serratia marcescens RP8 (974\u2009mg/L)   but lowe93\u2009mg/L)  and Serr74\u2009mg/L) . PSB in 74\u2009mg/L) , 14, 23,74\u2009mg/L) . The Pser peanut , Enterobhick pea , Acinetothaliana , and Psegeranium . The inogeranium . Previougeranium , 57. Therogenase , 57. Futgcd and bpp) in soil aggregates under different fertilization treatments. We found strong linkages between gcd gene abundance and tricalcium phosphate-solubilizing bacterial number, as well as between bpp gene abundance and phytate-degrading bacterial number. The phosphate-solubilizing bacteria Pseudomonas sp. PSB-2 and Arthrobacter sp. PSB-5 showed good performances for inorganic phosphorus solubilization. The phosphate-solubilizing bacterium Pseudomonas sp. PSB-2 could enhance growth of Chinese cabbage, showing significant increases in plant fresh and dry weight as well as plant height. Our findings extend the knowledge of mechanisms for distribution patterns of phosphate-solubilizing bacteria and P-cycling-related genes in soil aggregates and might guide the isolation of phosphate-solubilizing bacteria. Future work will use multiple techniques  to investigate P-cycling-related bacterial abundance and community composition and optimize condition for P solubilization by PSB and decipher mechanisms for P solubilization at both the gene and protein levels.In conclusion, we found distinct distribution patterns of phosphate-solubilizing bacteria and P-cycling-related genes  and winter wheat (Triticum aestivum L.) rotation since it was built in 1978. Five fertilization treatments were used, including CK, N, NPK, M, and MN  for soil aggregates by using quantitative PCR (qPCR). Primer bppF (5\u2032-GAC GCA GCC GAY GAY CCN GCN NTN TGG-3\u2032) and primer bppR (5\u2032-CAG GSC GCA NRT CAN CRT TRT T-3\u2032) were employed to amplify the bpp gene 2 or sodium phytate solid medium and incubated at 30\u00b0C for 5 days. The NBRIP contained 10 g/L glucose, 5 g/L Ca3(PO4)2 or sodium phytate, 0.25 g/L MgSO4\u00b77H2O, 5 g/L MgCl2\u00b77H2O, 0.2 g/L KCl, 0.1 g/L (NH4)2SO4, 2\u2009mL/L trace element solution, 0.2 g/L cycloheximide, and 18 g/L agar 2 were subcultured by picking and streaking five times to isolate pure colonies. We gained six strains  and identified them using simple 16S rRNA gene sequencing at Wuhan Qingke Innovation Biotechnology Co., Ltd. The universal primers 27F (5\u2032-AGA GTT TGA TCC TGG CTC AG-3\u2032) and 1492R (5\u2032-GGT TAC CTT GTT ACG ACT T-3\u2032) were used to amplify the 16S rRNA gene (Single colonies from the NBRIP containing CaRNA gene . The phy3(PO4)2 and incubated it at 30\u00b0C for 3\u2009days.The isolated PSB  were incubated in both solid and liquid NBRIP to estimate their solubilization potentials for tricalcium phosphate . We inoc7 CFU/mL) (Exp group). Each treatment had five replicates. Chinese cabbage (Shanghai Qing) seeds were purchased from China National Seed Group, precultivated in sterile nutritious soils, and allowed to grow to about 10-cm length of sprouts . The original physicochemical properties and processing of experimental potted soils were described previously . Two pot sprouts . Each sporimetry .If not otherwise stated, we estimated significant differences by using the one-way analysis of variance. Permutational multivariate analysis of variance was conducted using the function adonis in the vegan package of R. Canonical analysis of principal coordinates was used to estimate effects of soil physicochemical factors on PSB number and gene abundance using the capscale function in the vegan package. A structural equation model was built to reflect potential linkage among soil physicochemical factors, gene abundance, and PSB number using IBM SPSS Amos v.21. For the structural equation model (SEM), the PC1 value of the first axis of the principal-component analysis, accounting for 99.89% of the total variation, was applied as a proxy for representing P components ."}
+{"text": "Commiphora have been used in traditional medicines for centuries. More than 200 Commiphora species exhibit highly variable phytochemical compositions. A novel highly selective, sensitive, accurate HPLC-MS/MS method was developed and validated to quantify five characteristic phytosteroids and furanosesquiterpenoids, namely (E)-guggulsterone, (Z)-guggulsterone, curzerenone, furanoeudesma-1,3-diene, and myrrhone. The resulting contents and additionally GC analysis were used to classify and differentiate Commiphora oleogum resins of the species C. myrrha, C. erythraea, C. mukul, C. holtziana, C. confusa, and C. kua, as well as unspecified resins. Interestingly, a Commiphora sample from Ogaden, Ethiopia, comprised 446 ng/mg guggulsterones presumed to be unique to C. mukul from the Indian subcontinent. However, Commiphora from Ogaden differed considerably from C. mukul in respect to guggulsterones isomer\u2019s ratio. Moreover, the cytotoxicity of Commiphora extracts, essential oils, botanical drugs containing Commiphora, and pure compounds against the epidermoid carcinoma A431, malignant melanoma RPMI-7951 and SK-MEL-28 cells was investigated in vitro. Thereby, especially C. mukul extract and C. myrrha essential oil exhibited high cytotoxicity against skin cancer cells with IC50 of 2.9\u201310.9 \u00b5g/mL, but were less toxic to normal keratinocytes. In summary, Commiphora oleogum resins and its phytochemicals warrant further investigation aiming at chemotaxonomical classification as well as application in skin cancer treatment.Oleogum resins of the genus  Commiphora Jacq. of the Burseraceae family are shrubs or small trees with thorny branches and aromatic oleogum resin exudates with characteristic odors  Engl. (syn. C. molmol Engl.) predominantly growing in Somalia Commiphormponents . In factCommiphora oleogum resin or extract, namely Myrrhinil-Intest\u00ae and Gugulipid\u00ae were investigated. Additionally, two botanical drugs containing \u00ae is intended for use against gastrointestinal disorders, such as non-specific diarrhea, mild cramps, or flatulence. It contains chamomile flower extract (70 mg/pill), coffee charcoal (50 mg/pill), and powered C. molmol (syn. C. myrrha) oleogum resin (100 mg/pill) [C. myrrha analysis. However, Myrrhinil-Intest\u00ae contained with 0.456 \u00b5g/mg considerably less furanoeudesma-1,3-diene than the crude C. myrrha oleogum resin (87.7 \u00b5g/mg) (Myrrhinil-Intestmg/pill) . HPLC-MS7 \u00b5g/mg) a. \u00ae is considered a hypolipidemic drug containing an ethyl acetate extract of C. mukul [Z)-guggulsterone is the dominant isomer over (E)-guggulsterone with guggulsterone(ER/Z) = 0.61, which corresponds to the natural guggulsterone proportion in C. mukul [GugulipidC. mukul . ChemicaC. mukul ,22.Commiphora oleogum resin extracts and essential oils against the epidermoid carcinoma cell line A431 and the malignant melanoma cells lines RPMI-7951 and SK-MEL-28 were compared. The cell lines selected are rather resistant to treatment with the standard chemotherapeutic drugs cisplatin and 5-fluorouracil  between 8.4 and 21.4 \u00b5g/mL. Furthermore, Commiphora extracts were toxic for RPMI-7951 and SK-MEL-28 with IC50 = 2.9\u201311.5 \u00b5g/mL and IC50 = 10.9\u201323.4 \u00b5g/mL, respectively, which is comparable or even higher (particularly for SK-MEL-28 cells) than the cytotoxicity of cisplatin and 5-fluorouracil. Interestingly, the extracts obtained from C. mukul exhibited the highest cytotoxicity against all cell lines investigated. Statistical analysis confirmed higher cytotoxic efficacy of a C. mukul extract compared to extracts from C. erythraea, C. holtziana, C. kua, and C. from Tarraxo  to treat various conditions including cancers [C. myrrha, C. erythraea, and C. holtziana essential oils was shown to exhibit antiproliferative, proapoptotic and cytotoxic effect on human lung adenocarcinoma cells in vitro and in cancer xenografts in mice [Likewise, essential oils of ll lines . Essential cells . In Chin cancers . Particu cancers . Curzere in mice .C. mukul extract and C. myrrha essential oil were additionally tested for their cytotoxicity against non-carcinogenous human keratinocytes (MBU-IM). Here, the C. mukul extract and C. myrrha essential oil were less toxic to normal human keratinocytes than to malignant melanoma cells indicating their selectivity towards cancer cells -guggulsterone have been shown to induce caspase-dependent apoptosis in prostate cancer cells at concentrations > 10 \u00b5M [E)-guggulsterone exhibited cytotoxic effect on all three cancer cells lines with an IC50 = 12.6 \u00b5g/mL for A431, an IC50 = 3.7 \u00b5g/mL for RPMI-7951, and an IC50 = 11.1 \u00b5g/mL for SK-MEL-28. Similarly, (Z)-guggulsterone was also cytotoxic with an IC50 = 18.9 \u00b5g/mL for A431, an IC50 = 6.4 \u00b5g/mL for RPMI-7951, and an IC50 = 10.7 \u00b5g/mL for SK-MEL-28 - and (Z)-guggulsterone with regard to their toxicity against skin cancer cell lines were investigated. The combination index (CI) [E)- and (Z)-guggulsterones exhibited slight to moderate antagonistic interactions at effective doses ED50, ED75, and ED90. The only moderate synergistic interaction was observed in RPMI-7951 at ED50 . In addition to guggulsterones, C. mukul contains a further steroid substance group, termed guggulsterols [Because dex (CI) ,43, a qu at ED50 . This inlsterols . HoweverE)-guggulsterone, (Z)-guggulsterone, curzerenone, and furanoeudesma-1,3-diene were purchased from Sigma-Aldrich  and myrrhone from ChemFaces . Commiphora oleogum resins were provided by field experts  and subsequently deposited at the Herbarium of the Botanical Garden of Ulm University, Institute of Systemic Botany and Ecology, Germany (voucher: ULM-24224). For more precise sample information please see Commiphora botanical drugs Myrrhinil-Intest\u00ae (Batch No. 9100816) and Gugulipid\u00ae (Batch No. 1353619) were from Repha GmbH  and Natural Organics Inc. , respectively.All solvents and chemicals were of analytical reagent grade. The solvents used for the extraction, sample preparation, and HPLC-MS/MS analysis were MeOH, acetic acid , and ultrapure water  coupled to an Arium Pro station . The compounds  and evaporated to dryness by using a rotary evaporator.For extraction, 600 mg of freshly grounded n-hexane. The solvent of the combined organic phases was evaporated at 60 \u00b0C (water bath) with nitrogen stream.Essential oils were obtained by hydrodistillation of oleogum resins as previously described . ShortlyThe HPLC-MS/MS experiments were carried out on an Agilent 1260 Infinity HPLC system  coupled with an AB API 2000 triple quadrupole mass spectrometer  using an electrospray ionization ion source (ESI) in positive ionization mode. Devices were controlled and data were processed by means of Analyst 1.6.1 software .The chromatographic separation was performed using an analytical reversed-phase HPLC column  with a precolumn . v/v) (eluent A) and methanol (eluent B), both acidified with 0.2% acetic acid. Initial conditions were 32% eluent A and 68% eluent B followed by a linear gradient to 95% eluent B over 6.8 min, then 95% eluent B until 11.8 min. Thereafter, a linear gradient to initial conditions until 12.0 min and reequilibration continued until 17.0 min. In order to stabilize the chromatographic system, the column was kept at a temperature of 28 \u00b0C. The injection volume was set to 10 \u00b5L.By means of Design of Experiments (DoE) a novel method for chromatographic separation of guggulsterones was developed, in which the flow rate, the starting concentration of eluent B, and the gradient slope were optimized. Thus, the flow rate was 600 \u00b5L/min, the mobile phase consisted of methanol/water  and m/z 97.1 (product ion) as quantifier for (E)-guggulsterone and (Z)-guggulsterone. Moreover, m/z 313.3 (precursor ion) and m/z 109.1 (product ion) were used as qualifier for both guggulsterones. The ions m/z 231.0/83.0 (quantifier) and m/z 231.0/149.1  were used for curzerenone, m/z 229.0/159.0 (quantifier) and m/z 229.0/187.2  for myrrhone, and m/z 215.0/119.1 (quantifier) and m/z 215.0/105.2  for furanoeudesma-1,3-diene. The optimized source parameters and MS tune parameter are listed in The MS/MS detection was performed in multiple reaction monitoring (MRM) mode with Commiphora oleogum resins were not mixed with Boswellia oleogum resin, all samples were additionally analyzed for ten boswellic and lupeolic acids namely, \u03b1-boswellic acid (\u03b1-BA), acetyl-\u03b1-boswellic acid (\u03b1-ABA), \u03b2-boswellic acid (\u03b2-BA), acetyl-\u03b2-boswellic acid (\u03b2-ABA), 11-keto-\u03b1-boswellic acid (\u03b1-KBA), 11-keto-\u03b2-boswellic acid (\u03b2-KBA), acetyl-11-keto-\u03b1-boswellic acid (\u03b1-AKBA), acetyl-11-keto-\u03b2-boswellic acid (\u03b2-AKBA), lupeolic acid (LA), and acetyl-lupeolic acid (ALA). The boswellic and lupeolic acid analysis by HPLC-MS/MS was carried out as previously described [To ensure that escribed ,24.Commiphora species by GC-FID and GC-MS was carried out as described previously in detail [n-alkanes and mass spectra comparison with NIST17 libraries and in-house libraries. Moreover, 2-methoxyisofuranogermacrene and dihydropyrocurzerenone were identified by comparison of retention indices and mass spectra with expert literature [Analysis of essential oils in oleogum resins of different n detail ,45. The terature ,46. ResuThe skin cancer cell lines, epidermoid carcinoma cells A431, malignant melanoma cells RPMI-7951 and SK-MEL-25, and normal human keratinocytes MBU-IM were from the German Collection of Microorganisms and Cell Cultures  and cultured as recommended. 50 values were determined using SigmaPlot 14.0 software . Cells were seeded into 96-well plates and treated 24 h later using Tecan D300e digital dispenser . After 72 h incubation period, cell viability was analyzed by addition of 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide salt . Absorbance of the formed orange formazan dye was analyzed with an Infinite M1000 PRO Tecan plate reader (Tecan) at \u03bb = 450 nm with a \u03bb = 630 nm reference filter . For quaE)- and (Z)-guggulsterone in human skin cancer cell lines were carried out according to recommendations [Combination studies of cytotoxic effects of , SigmaPlot 14.0 software , and Valoo 2.10 software . All data were tested for normal distribution by the Anderson-Darling test and equality of variances by Levene\u2019s test. Sample groups were compared by one-way analysis of variance (ANOVA) and post hoc by Dunett\u2019s test. Comparison of two sample groups was carried out by Student\u2019s Commiphora by HPLC-MS/MS has been developed and validated. Additionally, essential oils of the respective oleogum resins were analyzed by GC. The phytochemical profiles were used to classify Commiphora oleogum resins of the species C. myrrha, C. erythraea, C. mukul, C. holtziana, C. confusa, and C. kua as well as unspecified Commiphora resins. Hence, patterns in the phytochemical composition were discovered assisting a chemotaxonomical differentiation among different Commiphora species. Interestingly, a Commiphora oleogum resin from the Ogaden region in Ethiopia comprised guggulsterones, which are unique for C. mukul from the India subcontinent. Considering the guggulsterones isomer\u2019s ratio and essential oil composition, Commiphora from Ogaden in Africa differs considerably from C. mukul suggesting that at least one African Commiphora species produces also phytosteroids such as guggulsterones. Moreover, the study provides evidence for cytotoxic efficacy of Commiphora extracts and essential oils against human epidermoid carcinoma and malignant melanoma cells in vitro. Here, especially C. mukul extract and C. myrrha essential oil exhibited the highest cytotoxicity against all three skin cancer cell lines investigated, but were less toxic to normal keratinocytes. Commiphora preparations and phytochemicals should be investigated more detailed regarding, for example, their systemic toxicity aiming at the development of new anticancer drugs.A highly, selective, and accurate method for the simultaneous determination of five phytosteroids and furanosesquiterpenoids in oleogum resins of the genus"}
+{"text": "Cytochrome P450 1B1 (CYP1B1) gene are a major cause of PCG. Current study was conducted to screen CYP1B1 gene in highly consanguineous PCG affected families from Pakistani population consistent with the autosomal recessive pattern of PCG inheritance.Primary congenital glaucoma (PCG) is a heterogeneous rare recessively inherited disorder prevalent in regions with high consanguinity. Disease phenotype is associated with increased intra ocular pressure and is a major cause of childhood blindness. Sequence variations in CYP1B1 gene was done for all enrolled families. In-silico analysis was performed to identify and predict the potential disease-causing variations.For this study, patients and controls  from 25 consanguineous families belonging to Punjab, Baluchistan and Khyber Pakhtunkhwa, Pakistan were recruited through ophthalmologists. DNA was isolated from collected blood samples. Genetic screening of Pathogenicity screening revealed sequence variants segregating with disease phenotype in homozygous or compound heterozygous form in eleven out of 25 analyzed families. We identified a total of sixteen disease causing variants among which five frameshift i.e., c.629dup (p.Gly211Argfs*13), c.287dup (p.Leu97Alafs*127), c.662dup (p.Arg222Profs*2), c.758_759insA  and c.789dup (p.Leu264Alafs*63), two silent c.1314G>A, c.771T>G and six missense variations c.457C>G (p.Arg153Gly), c.516C>A (p.Ser172Arg), c.722T>A , c.740T>A (p.Leu247Gln), c.1263T>A (p.Phe421Leu), and c.724G>C (p.Asp242His) are previously un reported. However two frameshift c.868dup (p.Arg290Profs*37), c.247del (p.Asp83Thrfs*12) and one missense variant c.732G>A (p.Met244Ile), is previously reported. Furthermore, six polymorphisms c.1347T>C, c.2244_2245insT, c.355G>T, c.1294G>C, c.1358A>G and c.142C>G were also identified. In the intronic region, a novel silent polymorphism i.e., g.35710_35711insT was found in homozygous state. All the newly detected disease-causing variants were negative in 96 ethnically matched controls.CYP1B1 gene. Genetic counseling was provided to families to refrain from practicing consanguinity and perform premarital screening as a PCG control measure in upcoming generations.Among twenty-five screened families, eight families  were segregating disease causing variants in recessive manner. Two families (PCG049 and PCG062) had compound heterozygosity. Our data confirms genetic heterogeneity of PCG in Pakistani population however we did not find molecular variants segregating with PCG in fifteen families in coding exons and intron-exon boundaries of  GLC3A [GLC3B [GLC3C [GLC3D [CYP1B1) at GLC3A and Latent Transforming growth factor-\u03b2-binding Protein-2 (LTBP2) at GLC3D have been reported to cause PCG [MYOC) [FOXC1) [TEK) [Glaucoma is characterized by impaired vision due to increased intraocular pressure, a primary risk factor for irreversible optic nerve damage. This disorder can be categorized according to etiology (primary glaucoma/secondary glaucoma), onset  and iridocorneal angle (open/close) . PrimaryGLC3A , GLC3B [A [GLC3B , GLC3C [B [GLC3C  and GLC3C [GLC3D  at positC [GLC3D . Among tause PCG . HoweverG [MYOC) , Forkhea [FOXC1) , and the1) [TEK)  have alsCYP1B1 gene has three exons out of which the last 2 codes for a 543 amino acid (a.a) protein [CYP1B1 in development of eye is uncertain however it is believed that due to mutations in this gene, generation of some important morphogens is affected leading to structural defects in trabecular meshwork and the aqueous humour outflow pathways [CYP1B1 gene including missense, small deletions, indels, gross deletions and regulatory mutations [CYP1B1 gene in PCG cases belonging to consanguineous Pakistani families, we enrolled and screened twenty-five families for CYP1B1 variants. Each family had at least one child affected with primary congenital glaucoma. protein . Cytochr protein . This me protein . Exact fpathways , 17, 18.utations . Studiesutations , 15, 20.http://haplopainter.sourceforge.net/about.html) [Based on clinical assessment provided by ophthalmologists, 25 diagnosed families of PCG were enrolled belonging to Khyber Pakhtunkhwa (KPK), Baluchistan and South Punjab. Clinical data, family history and blood samples of patients and each available family member was collected after informed written consent following the principles of world medical association of Helsinki . The stuut.html)  was usedet al., 2010 [Average 4ml of peripheral blood sample was taken from each participating individual and stored in 5ml EDTA (Ethylene Diamine Tetra Acetic acid) vacutainer. Extraction was performed using non-organic method of DNA extraction described by Kaul l., 2010 . To checet al., 2019 [et al., 2019 [https://www.mutationtaster.org/) [Amplification of coding regions and at least 50 base pairs of flanking non-coding regions was performed using primers reported previously by Afzal er.org/) . Furtherhttp://www.hgvs.org/) guidelines, Mutalyzer (2.0.35)  [https://varsome.com/) [https://www.genomnis.com/access-hsf) was used to determine pathogenicity due to disruption of splicing signals because of sequence variants.For significance of each variant and to check their nomenclature according to Human Genome Variation Society HGVS (zer.nl/)  was usedme.com/)  was usedhttp://genetics.bwh.harvard.edu/pph2/) [https://sift.bii.a-star.edu.sg/) [http://provean.jcvi.org/seq_submit.php) [https://folding.biofold.org/i-mutant/i-mutant2.0.html) [http://mupro.proteomics.ics.uci.edu/) [fwt-\u0394Gfmut) of protein structure that corresponds to stabilizing or destabilizing effect of variations. To check the conservation between different mammalian species, Clustal omega  [https://weblogo.berkeley.edu/) [https://www3.cmbi.umcn.nl/hope/) [PolyPhen-2 (Polymorphism Phenotyping v2) (u/pph2/) , SIFT (Sedu.sg/)  and PROVmit.php)  were use.0.html)  and MUprci.edu/)  softwareustalo/)  was usedey.edu/)  was usedl/hope/)  was usedIn this study, twenty-five PCG segregating consanguineous Pakistani families were enrolled. Among these families i.e., 09 belonged to Punjab province of Pakistan, 01 to Azad Kashmir whereas 12 and 03 belonged to Khyber Pakhtunkhwa and Baluchistan respectively. At the time of enrollment, seventeen families  had a single PCG affected individual, three families  had two, one family (PCG058) had three, three families  had four whereas family PCG050 had six affected members respectively .CYP1B1 coding regions and at least 50 base pairs of flanking noncoding region using DNA of each proband revealed thirteen novel disease-causing variations in coding regions according to mutation taster , (c.247del), (c.732G>A) and six reported polymorphisms (c.1347T>C), (c.1294G>C), (c.1358A>G), (c.2244_2245insT), (c.355G>T), (c.142C>G) were also found. Another polymorphism g.35710_35711insT was also found in intronic region in family PCG062 that has not been reported previously. Out of the novel disease-causing variants, five were frame shift variations c.629dup (p.Gly211Argfs*13), c.287dup (p.Leu97Alafs*127), c.662dup (p.Arg222Profs*2), c.758_759insA  and c.789dup (p.Leu264Alafs*63). Other novel disease-causing variants include six missense variants c.457C>G (p.Arg153Gly), c.516C>A (p.Ser172Arg), c.722T>A , c.740T>A (p. Leu247Gln), c.1263T>A (p.Phe421Leu), and c.724G>C (p.Asp242His) and two silent variations c.1314G>A and c.771T>G. In addition to disease causing variants, seven polymorphisms were also detected .In family PCG049, two disease causing variations were detected. A single nucleotide substitution i.e., c.457C>G was present in heterozygous condition, it changed arginine at position 153 to glycine and was deleterious according to PROVEAN and Polyphen-2 with a score of -5.21 and 1.00 respectively  Table 3Table 3. TTCGGC and Fas ESS site TGTTTC was broken. Two new ESS sites TTTTCG and GTTTTC were created for IIE and one TGTGTTTT for PESS (putative exonic splicing silencer).Insertion of thymine (T) in PCG050 in exon 2 at position 629\u2013630  changed CYP1B1 gene  that replaced aspartic acid at position 83 to threonine shifting reading frame and creating stop codon after 12 residues . I-MutanCTGTGGTTTTTGTC>CTGTGGTTTTAGTC changing consensus value (CV) from 50.91 to 78.78. CV for newly created site showed that it is not a very strong site (strong site CV> 80). PCG063 had two unreported mutations, a silent heterozygous mutation c.771T>G and a frameshift homozygous mutation c.789dup , Nomascus leucogenys (XP_003262792.2), Pongo abelii (XP_009235654.1) and Pan troglodytes (XP_001167556.1) with high similarity index to Homo sapiens   was founCYP1B1 coded protein is reported to cause abnormal development of ocular structures resulting in impeded outflow of aqueous humor and PCG phenotype [CYP1B1 gene varies among different populations i.e.; p.Ser476Pro is 44% prevalent in India, p.Arg469Trp, p.Arg368His, p.Arg390His, p.Gly61Glu and p.Glu173Arg are 70% prevalent in Iran, p.Gly61Glu, p. Arg390His and p.Glu229Lys are 80\u2013100% prevalent in Saudi Arabia however p.Arg330Phe and p.Arg390His are predominantly reported from China [High frequency i.e., 70\u2013100% of consanguineous marriages  is the mhenotype , 36. Datom China . Founderom China . Previouom China , 38, 39;CYP1B1 analysis in 25 cases enrolled through various regions of Punjab, Baluchistan and Khyber Pakhtunkhwa, Pakistan revealed a total of seven frameshift, seven missense and two silent disease-causing variations. Among seven frameshift variations five are novel however two are previously reported in patients of different ethnicities. The variant c.868dup (p.Arg290Profs*37) (rs67543922) was initially identified in PCG affected Pakistani family by Sheikh et al., 2014 [et al., 2015 [CYP1B1 conducted on population of Sindh and Punjab province of Pakistan by Rashid et al., 2019 [In present study l., 2014  and then7) rs675422 was inl., 2019  reportedet al., 2009 [et al., 2018 [Second reported frame shift variant c.247del (p.Asp83Thrfs*12) was initially identified in a study conducted on Indian population by Tanwar l., 2009 . Accordil., 2009 . Five hol., 2018  have shol., 2018 .CYP1B1 gene. Previously compound heterozygosity has been reported in developmental glaucoma, [et al., 2021 reported that two heterozygous mutations c.1310C>T (p.P437L) and c.3G>A (p.M1I) are responsible for glaucoma in a Chinese family [et al., 2019 identified compound heterozygosity  in two consanguineous families of PCG belonging to different ethnic groups of Pakistan [CYP1B1 with heterozygous TEK alleles in PCG cases [CYP1B1, recessive inheritance pattern and previously reported allelic interactions of two un linked genes for PCG phenotype [Missense disease-causing variants found in family PCG049, 052, 060, 062 and 067 and two silent disease-causing variants found in family PCG062, 063 are also previously unreported , Table 3laucoma,  and primlaucoma, . Cai et e family . In anote family . Waryah Pakistan . FurtherCG cases . In preshenotype , 13, 48 CYP1B1 disease causing variations in PCG patients [LTBP2, TEK, MYOC, FOXC1 and regulatory effect of cis-acting elements, splicing elements or possible modifiers [Due to epigenetic modifications and different environmental factors incomplete penetrance and increased variability could be observed in manifestation of patients , 49. We odifiers , 40, 50.et al., 2019 [All single nucleotide polymorphisms (SNPs) identified in current study except one present in intronic region i.e., g.35710_35711insT in homozygous state are previously reported. Four reported SNPs i.e., rs1056836 (c.1294G>C), rs1800400 (c.1358A>G), rs1056827 (c.355G>T) and rs10012 (c.142C>G) showed amino acid change while two polymorphisms rs1056837 (c.1347T>C) and rs4646431 (c.2244_2245insT) were silent. Most prevalent polymorphism (45%) c.1347T>C in present study was also reported in other studies conducted on PCG cases from Pakistani population . Afzal eCYP1B1 gene to PCG pathophysiology in our population.In conclusion we identified thirteen previously unreported and three reported mutations as well as six SNPs (one novel) in PCG probands born to parents having consanguineous marriages highlighting the autosomal recessive pattern of disease. Proper genetic testing and counseling should be provided to people in high consanguinity areas to help ophthalmologists in disease management and treatment. Mass screening and additional studies are required to better understand the heterogeneous pattern and contribution of"}
+{"text": "Corona virus disease-19 (Covid-19) has significantly affected organ transplantation with concerns regarding severe infection and mortality. Data on Covid-19 in renal transplant recipients (RTRs) is scarce from Pakistan. The aim of this study is find out the factors effecting mortality among Covid-19 patients in renal transplant recipients from the largest transplant center of Pakistan.All RTRs >18 years, with positive severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) polymerase chain reaction (PCR) and diagnosed as severe disease, between April to December 2020 were retrospectively reviewed. The severe disease was defined as O2 saturation < 94% at room air on admission. Survivors and non- survivors were compared. Demographics, immunosuppression, comorbid conditions, clinical features, laboratory investigations and graft function were noted.A total of 95 RTRs had severe disease. There was no difference in mortality between age, gender and co-morbid conditions among survivors and non-survivors. Both groups received similar immunosuppressive regimen. Intensive care unit (ICU) admission [16.5% vs 68.8% p< 0.001 OR 11.17 95% CI (3.3-37.6)] and high D-dimers >1.5\u00b5g/ml (p=0.052) at the time of admission were significantly associated with mortality. There was no association of graft function with mortality. Treatment with methyl-prednisolone was found to be significantly associated with survival [83% vs 43% P=0.02 OR 0.15 95% CI (0.05-0.49)]. (Table\u00a01) WHO grading of the disease is shown in figure 1, there was a 100% mortality among patients on mechanical ventilator.ICU admission and high D-dimers at the time of admission are the significant risk factors for mortality in patients with Covid-19 infection. There was no association of graft dysfunction with mortality. Steroids use has significantly improved survival in renal transplant recipients with severe Covid-19 infection.All Authors: No reported disclosures."}
+{"text": "Colitis-associated cancer (CAC) is a subtype of inflammatory bowel disease (IBD)-associated colorectal cancer. Huoxiang Zhengqi (HXZQ) is a classical Chinese herbal medicine and has been used to treat intestinal disorders, however, anti-CAC effects and underlying mechanisms of HXZQ have not been reported. An azoxymethane/dextran sulfate sodium-induced CAC mice model was used to investigate the anti-CAC effect of HXZQ. HXZQ significantly reduced colonic inflammation, suppressed the size and number of tumors, and reduced the levels of pro-inflammatory cytokines  and oxidative stress markers , and increased the levels of anti-inflammatory cytokines (IL-10 and IL-27) in CAC mice. Intestinal microbiota and serum metabolomics analyses indicated that HXZQ altered the gut microbial composition and the abundance of 29 serum metabolites in CAC mice. Additionally, HXZQ activated the nuclear factor-erythroid factor 2-related factor 2 (Nrf2) signaling pathway and increased the levels of antioxidants such as catalase (CAT), heme oxygenase-1 (HO-1), NAD(P)H quinone oxidoreductases-1 (NQO-1), and superoxide dismutase-1 (SOD-1). HXZQ inhibited the activation of the nuclear factor kappa-B (NF-\u03baB) signaling pathway and decreased the expression of NLR family pyrin domain containing 3 (NLRP3) by inhibiting the phosphorylation of inhibitor of nuclear factor kappa-B (I\u03baB), inhibitor of nuclear factor kappa-B kinase (IKK), and NF-\u03baB. In conclusion, HXZQ alleviated CAC in mice by modulating the intestinal microbiota and metabolism, activating Nrf2-mediated antioxidant response, and inhibiting NF-\u03baB-mediated NLRP3 inflammasome activation against inflammation. The present data provide a reference for the use of HXZQ as a therapeutic or combination agent for clinical CAC treatment. Colitis-associated cancer (CAC) is a subtype of inflammatory bowel disease (IBD)-associated colorectal cancer. IBD includes Crohn\u2019s disease (CD) and ulcerative colitis (UC) . InflammAtractylodes, regulates oxidative stress through the Nrf2 pathway and alleviates 2,4,6-trinitrobenzenesulfonic acid-induced acute colitis by influencing the composition of intestinal microbiota  that does not express BF enterotoxin to prevent DSS-induced colitis and prevent the formation of polyps in the model of CAC production, which affects the homeostasis of the intestinal microbiota . ROS canel of CAC.Prescriptions of Peaceful Benevolent Dispensary for approximately 900\u00a0years and has been used to treat intestinal disorders (Huoxiang Zhengqi (HXZQ) is a classic Chinese herbal medicine  that wasisorders . Recent isorders . Norfloxisorders . HoweverThe azoxymethane (AOM)/DSS-induced CAC mouse model mimics the pathology of CAC in humans and has been used for screening agents with anti-CAC properties and revealing the underlying mechanisms . In the Thirty-two 6-week-old male C57BL/6 mice (SCXK(Liao)2020\u20130,001, Liaoning Changsheng Biotechnology Co., Ltd. Benxi, China) were housed under specific pathogen-free conditions at the appropriate temperature (22 \u00b1 2\u00b0C) and humidity (50% \u00b1 10%) and kept on a 12/12\u00a0h light/dark cycle. The mice had unrestricted access to food and water. The experimental protocol complied with the ARRIVE guidelines and was approved by the Institution Animal Ethics Committee of Jilin University (SY202104007).n = 8) (serving as the model group) or 0.45 or 1.35\u00a0g/kg HXZQ  (serving as the HXZQ-treated groups) daily for 6 weeks. Another eight mice were intraperitoneally injected with normal saline on the first day, received normal drinking water for the entire experimental period, and received normal saline orally daily from the fifth to 10th\u00a0week (serving as the control [Ctrl] group)  on the first day, and their drinking water was changed to 2% DSS  at the second, fifth, and eighth week. From the fifth week and beyond, the mice were randomly divided into three groups and orally received normal saline (] group) . Six houThe fixed colon, liver, spleen, and kidney tissues were paraffin-embedded, sectioned, and dewaxed in xylene for 40\u00a0min, anhydrous ethanol for 10\u00a0min, and 75% ethanol for 5\u00a0min, and then washed with running water. The sections were stained with hematoxylin and eosin (H&E) and treated sequentially with ethanol and xylene for dehydration. The sealed sections were observed and analyzed using an ECLIPSE E100 upright optical microscope  .Paraffin slides of colorectal tumors were subjected to antigen retrieval after de-paraffinization and blocked with 3% bovine serum albumin. The slides were incubated overnight at 4\u00b0C with primary antibody for nuclear factor kappa-B factor kappa-B (NF-\u03baB) p65  in a wetn = 4) were used for 16S rRNA analysis of the gut microbiota. Nucleic acids were extracted from the contents of each cecum, and PCR amplification of the V3-V4 region of the bacterial 16S rRNA gene was performed. PCR products were quantified and 2 \u00d7 250\u00a0bp double-end sequencing was performed. Sequencing was performed at Shanghai Personal Biotechnology Co. Ltd. .The cecum contents obtained from Ctrl, Model, and HXZQ-treated mice (https://www.ncbi.nlm.nih.gov/sra/PRJNA860221). These analyses were performed as previously described (The 16S rRNA sequencing results were clustered into amplicon sequence variants (ASVs) using DADA2 with 100% similarity. Based on the ASV abundance data, a flower plot was generated, and alpha diversity indices and the weighted UniFrac distance matrix were calculated. Abundance data from each group of microbiota were used to generate a species composition heatmap and for LDA effect size (LEfse) analysis. Microbial functions were predicted using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States. The sequences of bacteria were uploaded to the NCBI Sequence Read Archive with accession number PRJNA860221  for vortex mixing. The supernatant obtained by centrifugation of the mixture was vacuum dried to a solid state. Before the analysis, the samples were dissolved in an aqueous solution of acetonitrile . Samples were separated on an ultra-high-performance liquid chromatography system  equipped with a ACQUITY UPLC BEH Amide 1.7\u00a0\u03bcm, 2.1\u00a0mm \u00d7 100\u00a0mm column  and were analyzed using a quadrupole-time of flight mass spectrometer . The separation conditions were identical to those reported in previous studies .p-value <0.05 as filtering criteria were used to draw heatmaps and correlation plots. The abundance data of the microbiota and metabolites with significant changes were jointly analyzed to obtain association heatmaps. These analyses were performed as described in our previous study , and score plots were plotted. The abundance of signature metabolites screened with OPLS-DA VIP >1 and us study .The collected tumor tissues were homogenized separately in phosphate buffer solution (PBS). Protein concentrations were determined using a Pierce\u2122 BCA Protein Assay Kit . IL-1\u03b1, IL-1\u03b2, IL-6, IL-17A, IL-23, IL-27 and TNF-\u03b1 in tumor tissues were detected using the LEGENDplex\u2122 Mouse Inflammation Panel (13-plex) with a V-bottom plate . Enzyme-linked immunosorbent assay (ELISA) kits  were used to measure IL-10 (#MM-0176M1), IL-21 (#MM-0688M1), granulocyte-macrophage colony-stimulating factor (GM-CSF) (#MM-0185M1), malondialdehyde (MDA) (#MM-0897M1), and ROS (#MM-43700M1) levels in tumors.Tumor tissues were lysed at low temperatures in Radioimmunoprecipitation assay (RIPA) Buffer  supplemented with 1% Protease and Phosphatase Inhibitor Cocktail . Cytoplasmic and nuclear proteins in tumors were isolated by NE-PER Nuclear and Cytoplasmic Extraction Kit . Total protein in each sample was measured using the Pierce\u2122 BCA Protein Assay Kit . The protein samples were separated using 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The proteins were transferred to PVDF membranes, and the membranes were blocked with western fast blocking solution  for 10\u00a0min at 25\u00b0C. The membranes were incubated with primary antibodies  for 12\u00a0hp-value less than 0.05 was considered statistically significant.All values are presented as mean \u00b1 standard error of the mean (S.E.M.) Biochemical indices were compared between Ctrl and Model using Student\u2019s t-test. Comparisons among Model, 0.45\u00a0g/kg HXZQ, and 1.35\u00a0g/kg HXZQ were performed with one-way analysis of variance (ANOVA) followed by a post hoc multiple comparisons (Dunnett) test using BONC DSS Statistics 25 software . A p < 0.05) (HXZQ remarkably reduced the number and size of colonic tumors in CAC mice without influencing their body weight ( < 0.05) . Compare < 0.05) . HXZQ fa < 0.05) .Candidatus_Arthromitus, Turicibacter, Dorea, Acinetobacter, Clostridium, and Desulfovibrio, which were suppressed by HXZQ. Compared to the model group, HXZQ resulted in an increased abundance of 25 genera  (p < 0.001)  . The abu LDA >2) . Compara< 0.001) .p < 0.05) (The score plots of OPLS-DA showed significant differences in metabolite levels between the model and HXZQ-treated mice . The lev < 0.05) .Phascolarctobacterium, Prevotella, Helicobacter, and Butyricimonas (p < 0.05) (Candidatus_Arthromitus and p-75-a5 (p < 0.05) (Serum metabolite concentrations were significantly correlated with changes in colony abundance. Indoxyl sulfate was negatively correlated with  < 0.05) . L-Gluta < 0.05) .p < 0.01) (p < 0.01) (p < 0.001) (p < 0.01) (p < 0.001) (p < 0.01) (p < 0.05) (p < 0.05) (p < 0.001) (p < 0.05) (p < 0.001) (p < 0.01) (Inflammation severity tended to correlate positively with CAC development. HXZQ do < 0.01)  and ROS  < 0.01) , and red< 0.001) , IL-1\u03b2 ( < 0.01) , IL-6 (>< 0.001) , IL-17A  < 0.01) , IL-21 ( < 0.05) , IL-23 ( < 0.05) , TNF-\u03b1 (< 0.001) , GM-CSF  < 0.05) . Corresp< 0.001)  and IL-1 < 0.01) .p < 0.05), catalase (CAT) (>400%) (p < 0.001), heme oxygenase-1 (HO-1) (>400%) (p < 0.05), NAD(P)H quinone oxidoreductases-1 (NQO-1) (>470%) (p < 0.001), and superoxide dismutase-1 (SOD-1) (>220%) (p < 0.01) in colorectal tissues (p < 0.05), p-NF-\u03baB (>52%) (p < 0.05), phosphorylated inhibitor of nuclear factor kappa-B kinase (p-IKK) (>33%) (p < 0.001), IL-1\u03b2 (>51%) (p < 0.01), IL-6 (>63%) (p < 0.001), TNF-\u03b1 (>40%) (p < 0.001), and NLRP3 (>29%) (p < 0.01) in colorectal tissues (p < 0.001)  ( tissues . HXZQ do tissues . HXZQ in< 0.001) . The res< 0.001) .IL10-/- mice spontaneously developed colitis, whereas upregulation of IL-10 alleviated CAC, which is consistent with our study  production and a strong positive correlation with the pro-inflammatory cytokine TNF-\u03b1 (Acinetobacter activates the NLRP3 inflammasome and consequently mediates IL-1\u03b2 production (Turicibacter and Desulfovibrio is increased under inflammatory conditions (Desulfovibrio has been reported to induce secretion of IL-6 and IL-8 from endothelial cells (Notably, changes in pro-inflammatory cytokine levels may also affect host\u2019s intestinal microbiota. Accordingly, IL-33-deficient mice developed a dysregulated gut microbiota , and alt colitis . Disrupt colitis , which a colitis . The eff colitis . HXZQ inne TNF-\u03b1 . Acinetooduction . The gronditions , and Desal cells .Intestinal microbiota directly influence host metabolism . HXZQ inNLRP3 transcription (Unsurprisingly, HXZQ activated Nrf2 signaling while increasing the levels of antioxidant enzymes such as CAT, HO-1, NQO-1, and SOD-1. Moreover, HXZQ inhibited the phosphorylation of I\u03baB\u03b1, IKK\u03b1/\u03b2, and NF-\u03baB as well as the expression of NLRP3. The accumulation of ROS during chronic inflammation leads to oxidative stress that aggravates CAC development . Nrf2 cocription .The present study has some limitations. The detailed relationship between HXZQ-mediated gut microbiota regulation and the anti-inflammatory and anti-oxidant effects that underlie its anti-CAC activity requires further investigation. Second, the main components involved in the therapeutic effect of HXZQ in the treatment of CAC need to be identified. Finally, the effects of HXZQ on other IBDs require further investigation.Altogether, HXZQ alleviates CAC by modulating the intestinal microbiota and metabolism, activating Nrf2-mediated antioxidant response, and inhibiting NF-\u03baB-mediated NLRP3 in mice. Our data provide a reference for the use of HXZQ as a therapeutic agent or a combination agent for clinical CAC treatment."}
+{"text": "Paenibacillus polymyxa DSM 365. The genome consists of a 5,788,318-bp chromosome, with a GC content of 45.48%. Annotation of the genome revealed a total of 5,246 genes . Gene function analysis indicated the ability to fix nitrogen (N2) and to produce value-added chemicals.We report the complete genome sequence of  Paenibacillus polymyxa DSM 365 is a Gram-positive plant growth-promoting rhizobacterium . To isolate DNA, cultures were grown overnight in tryptic soy broth at 30 \u00b0C and 200\u2009rpm. Genomic DNA was extracted using the Wizard high-molecular-weight (HMW) DNA extraction kit . Library preparation and sequencing were conducted by Novogene Inc.  using the Illumina NovaSeq 6000 platform. To prepare the library for sequencing, genomic DNA was randomly sheared into short fragments. The fragments were end repaired, adenine tailed, and ligated with Illumina adapters. The quantified libraries (350-bp size) were pooled and sequenced to produce 6\u2009Mb of paired-end 150-bp reads . In order to ensure accuracy and reliability, the reads were filtered using readfq software (v.10)  in 47 scaffolds, with a GC content of 45.48% and an average read coverage of 291\u00d7.acterium  with caphemicals \u20137. P. poe v.10) , 10, SPA , 10, SPe (v.10) , and ABye (v.10)  assemblye (v.10) . Assemble (v.10) . GapClose (v.10)  was usedGeneMarkS (v.4.10)  was used16\u2013P. polymyxa DSM 365 was submitted to the National Center for Biotechnology Information (NCBI) database using the Prokaryotic Genome Annotation Pipeline (PGAP) (v.6.0) (e\u22125). Coding genes were predicted by Augustus (v.2.7) (nif operon), sporulation, acetoin utilization, biosynthesis of siderophores, polyketides, exopolysaccharides, and butanediol were detected.The whole-genome sequence of  (v.6.0) . Homolog (v.6.0)  was used (v.2.7)  with homP. polymyxa DSM 365 has been deposited in GenBank under the BioProject accession number PRJNA809744, the BioSample accession number SAMN26200526, and the Sequence Read Archive (SRA) accession number SRR18173204. The whole-genome shotgun project has been deposited in DDBJ/ENA/GenBank under the accession number JAKVDC010000000.The annotated genome sequence of"}
+{"text": "Data on willingness to participate in population-based long-COVID studies are sparse. We invited all citizens of Essen aged 18-74 years with a positive SARS-CoV-2 PCR test between Mar-Aug 2020 and assessed COVID-related symptoms in responders \u223c1.5 years after infection.The invited population included 1282 infected citizens (48% women). At the time of testing 64% reported symptoms. We asked responders about past and current symptoms, hospitalization, smoking, sport, pre-existing conditions , subjective health status as compared to before infection, assessed BMI, and performed descriptive statistics.2), and more pre-existing conditions (23% vs 10%). Compared to before infection, 53% rated their current health worse, with a higher rate among inpatients (81%). After \u223c1.5 years, 55% still reported symptoms: 25% fatigue, 20% concentration disorder, 18% breathing problems, 13% odor and 11% taste disorders. Persistent symptoms were more common in inpatients than in non-hospitalized (69% vs 53%).We investigated 255 participants  \u223c20 month (median) after the PCR test. 95% reported symptoms at the time of testing: 67% fatigue, 58% taste disorders, 56% limb pain, 55% odor disorders, 54% headache, 50% cough, 43% fever; 10% needed hospitalization, 3% intensive care, 1.6% artificial ventilation. Compared to the non-hospitalized the formerly inpatients were more often male (62% vs 49%), older (56\u00b113 vs 49\u00b114 years), less often never smokers (42% vs 53%), had a higher BMI (31\u00b17 vs 28\u00b15 kg/mSymptomatic individuals are more likely to participate in a COVID19 follow-up study than asymptomatic ones. This may overestimate the number of individuals with long-term symptoms in population-based long-COVID study populations. However, persistent symptoms seem to be more likely in formerly inpatients compared to non-hospitalized individuals with former SARS-CoV-2 infection.\u2022\u2002Symptomatic individuals are more likely to participate in a COVID19 follow-up study than asymptomatic ones.\u2022\u2002Persistent symptoms seem to be more likely in formerly inpatients compared to non-hospitalized individuals with former SARS-CoV-2 infection."}
+{"text": "Endocrine disruption is an important factor in the development of endometrial cancer. Expression of miR-149-3p is observed in some cancer types, while its role in uterine corpus endometrial carcinoma (UCEC) is unclear.  The clinical and genomic data and prognostic information on UCEC were obtained for patients from the TCGA database. The Kruskal\u2013Wallis test, Wilcoxon signed-rank test, and logistic regression were used to analyze the relationship between clinical characteristics and miR-149-3p expression. Kaplan\u2013Meier survival curve analysis was used to study the influence of miR-149-3p expression and miR-149-3p target genes on the prognosis of UCEC patients. The TargetScan, PicTar, Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were used to determine the involvement of miR-149-3p target genes in function. Immune infiltration analysis was used to analyze the functional involvement of miR-149-3p. QRT-PCR was used to validate the expression of miR-149-3p in UCEC cell lines. P < 0.001), histological type (P < 0.001), histological grade (P < 0.001), tumor invasion (P=0.014), and radiation therapy (P=0.011). High miR-149-3p expression predicted poorer overall survival (OS) , progression-free interval (PFI) , and disease-specific survival (DSS) . Low expressions of miR-149-3p target genes, including ADCYAP1R1, CGNL1, CHST3, CYGB, DNAH9, ESR1, HHIP, HIC1, HOXD11, IGF1, INMT, LSP1, MTMR10, NFIC, PLCE1, RARA, SNTN, SPRYD3, and ZBTB7A, were associated with poor OS in UCEC. MiR-149-3p may be involved in the occurrence and development of UCEC via pathways including PI3K-Akt signaling pathway, Ras signaling pathway, AGE-RAGE signaling pathway in diabetic complications, focal adhesion, and MAPK signaling pathway. miR-149-3p may inhibit the function of CD8 T cells, cytotoxic cells, eosinophils, iDC, mast cells, neutrophils, NK CD56bright cells, NK CD56dim cells, pDC, T cells, T helper cells, TFH, Th17 cells, and Treg. miR-149-3p was significantly upregulated in UCEC cell lines compared with endometriotic stromal cells.  High expression of miR-149-3p in UCEC was significantly associated with age ( High expression of miR-149-3p was significantly associated with poor survival in UCEC patients. It may be a promising biomarker of prognosis and response to immunotherapy for UCEC patients. Uterine corpus endometrial carcinoma (UCEC) is one of the three main gynaecological malignancies, the incidence of which increases over time. Endometrial cancer is an epithelial tumor of the endometrium and a malignant tumor of the female reproductive system with a high incidence, posing a serious threat to the health of women worldwide , 2. It cMicroRNAs (miRNAs) are a unique class of endogenous and small noncoding RNAs that are approximately 18 to 25 nucleotides in length. They alter gene expression at the posttranscriptional level primarily through complete or incomplete base pairing with the 3\u2032 untranslated region (3\u2032UTR) of their target mRNAs. Translational repression and mRNA degradation are the 2 main pathways through which miRNAs direct gene regulation . There iIn oral squamous cell carcinoma, reduced levels of miRNA-149-3p lead to malignant progression and predict poor prognosis . IncreasThe present study examined the expression of miR-149-3p in UCECs using an online database and analyzed the relationship between expression levels and clinical characteristics. A survival curve was drawn to analyze the relationship between miR-149-3p expression level and overall survival (OS). Important contributions of miR-149-3p target genes to function were identified by TargetScan, PicTari, Gene Ontology (GO), and Kyoto Gene and Genome Encyclopedia (KEGG) analyses. The functionally significant involvement of miR-149-3p was analyzed by immune infiltration analysis. QRT-PCR was used to validate the expression of miR-149-3p in UCEC cell lines. The results of this study could provide new prognostic biomarkers for UCEC.https://portal.gdc.cancer.gov/) UCEC project. The miRNAseq data in RPM (reads per million mapped reads) format were log2-transformed.The analysis was carried out according to references , 17. R . R package was the ggplot2 package.Nomogram plot analysis was carried out according to literature , 20. R phttps://bioinfo.life.hust.edu.cn/miR_path/download.html). The UCEC prognosis-related genes were analyzed according to reference [miR-149-3p targets were obtained from Database TargetScan, miRanda, TarBase, miRTarBase, miR2Disease, miRecords, and miRWalk \u201325. UCECeference . R  [P value less than 0.05) were considered significant categories.The Database for Annotation, Visualization, and Integrated Discovery (DAVID) can provide a comprehensive set of functional annotation tools to facilitate understanding of the biological significance behind a large number of genes. GO and KEGG analyses were performed on the targets of miR-149-3p using the DAVID database (rf.gov/) \u201328. GO aThe analysis was performed according to reference . R  have been isolated from endometriotic tissue. Human UCEC cells Ishikawa and KLE were obtained from our laboratory. KLE cells were grown in F12. Ishikawa cells were grown in RPMI-1640. 10% fetal bovine serum was added to the medium to maintain the cell state. Add 1% antibiotics to the soil to prevent contamination. Cells were grown in a 37\u00b0C incubator containing 5% COeference . The priP values less than 0.05 were considered statistically significant.Statistical analysis was carried out according to reference . All staR0 (90.9%), 22 R1 (5.4%), and 15 R2 (3.7%). The histologic grade included 98 G1 (18.3%), 121 G2 (22.6%), and 316 G3 (59.1%). Tumor invasion (%) included 260 patients (<50) (55.3%) and 210 patients (\u226550) (44.7%). The menopause status included 34 pre (6.8%), 17 peri (3.4%), and 449 post (89.8%). The hormone therapy included 297 no (86.6%) and 46 yes (13.4%). The diabetes included 323 no (72.6%) and 122 yes (27.4%). The radiation therapy included 278 no (53.4%) and 243 yes (46.6%). The surgical approach included 207 minimally invasive (39.6%) and 316 open (60.4%). The age range was 57 to 71 years, with a median of 64 years.A total of 546 patients were analyzed in the present study . The clin\u2009=\u2009546) was significantly higher than that in normal tissues  (P=0.007)  . The areP=0.007) , suggestn\u2009=\u2009273) and low (n\u2009=\u2009273) expression groups. As shown in P < 0.001), histological type (P < 0.001), histological grade (P < 0.001), tumor invasion (P=0.014), and radiation therapy (P=0.011). As shown in P=0.048), age , histological type , histologic grade , tumor invasion , and radiation therapy .The characteristics of UCEC patients are shown in P < 0.001), PFI , and DSS  of UCEC patients (P < 0.001), clinical stage , primary therapy outcome , age , histological type , residual tumor , histologic grade , tumor invasion , and radiation therapy . As shown in P < 0.001), primary therapy outcome , radiation therapy , and hsa-miR-149-3p  were independently correlated with OS in multivariate analysis. The above data indicated that miR-149-3p is a prognostic factor, and increased miR-149-3p is associated with poor OS. A nomogram was constructed to predict the 1-, 3-, and 5-year survival probability of UCEC patients by combing the expression of miR-149-3p with clinical variables, as shown in The association between miR-149-3p expression and OS of patients with UCEC was evaluated by Kaplan\u2013Meier analysis, which indicated that the expression of miR-149-3p was correlated with poor OS , cytotoxic cells (P < 0.001), eosinophils (P < 0.001), iDC (P < 0.001), mast cells (P=0.002), neutrophils (P < 0.001), NK CD56bright cells (P < 0.001), NK CD56dim cells (P < 0.001), pDC (P < 0.001), T cells (P < 0.001), T helper cells (P=0.004), TFH (P=0.024), Th17 cells (P=0.006), and Treg (P < 0.001).As shown in Figures P < 0.001) (P < 0.001)  . The exp< 0.001) . These rThe development of type I endometrial cancer is associated with continuous estrogen stimulation of the endometrium without progestin antagonism. The endometrium lacks progesterone antagonism, and continuous stimulation by estrogen will result in a prolonged state of hyperproliferation, which will further develop into endometrial cancer. MiRNAs regulate the cell cycle and cell differentiation and migration, which may also act as tumor suppressor genes or oncogenes during tumorigenesis and tumor development . MiRNAs P < 0.001), histological type (P < 0.001), histological grade (P < 0.001), tumor invasion (P=0.014), and radiation therapy (P=0.011). UCEC expressed more miR-149-3p than normal tissue, especially in patients with age (>60), histological type (mixed and serous), histologic grade (G2 and G3), tumor invasion (\u226550%), or radiation therapy (yes). These phenomena suggested that miR-149-3p may be involved in tumor development and promote proliferation. And miR-149-3p was highly expressed in UCEC, and patients with high miR-149-3p expression had poorer OS , PFI , and DSS  of UCEC patients. Low expressions of miR-149-3p target genes, including ADCYAP1R1, CGNL1, CHST3, CYGB, DNAH9, ESR1, HHIP, HIC1, HOXD11, IGF1, INMT, LSP1, MTMR10, NFIC, PLCE1, RARA, SNTN, SPRYD3, and ZBTB7A, were associated with poor OS in UCEC. Lymphovascular space invasion (LVSI) has an independent influence on the poor prognosis of UCEC patients [In the present study, miR-149-3p was significantly correlated with age , histological type (P < 0.001), histological grade (P < 0.001), tumor invasion (P=0.014), and radiation therapy (P=0.011). miR-149-3p may be involved in the occurrence and development of UCEC via pathways, including PI3K-Akt signaling pathway, Ras signaling pathway, AGE-RAGE signaling pathway in diabetic complications, focal adhesion, and MAPK signaling pathway. Expression of miR-149-3p was correlated with immune infiltration in UCEC. This study partially elucidates the role of miR-149-3p in UCEC and provides a promising biomarker for prognosis and immunotherapy response in UCEC patients.miR-149-3p was highly expressed in UCECs and correlated with poorer OS compared with normal tissues. The high miR-149-3p expression in UCEC was significantly associated with age ("}
+{"text": "Substantial changes in access and delivery of primary HIV care occurred during the COVID-19 pandemic. To assess how care access changed during the COVID-19 pandemic, we estimated ED use among PWH in care 2017-2021 in the southeastern US.For each calendar year, among PWH in care in the UNC CFAR HIV Clinical Cohort (defined as having a clinic visit in the current or prior year), we estimated the percent of patients with \u2265 1 ED visit in a given year, overall and by age, gender, race/ethnicity, HIV viral load (VL), and CD4 count. We estimated risk ratios (RRs) comparing patient characteristics and years 2020-2021 vs. 2017-2019, using Poisson regression with generalized estimating equations to account for repeated measures.B-F, all P< 0.05).Among 2129 PWH in care 2017-2021 (N\u22481700-1800 in each year), 57% identified as Black, 31% White, 8% Hispanic, 26% women, with median age of 47 years (IQR 35-55). During the study period, there were 3645 ED visits over 8813 person-years, a rate of 41.4 ED visits-per 100 person-years(95% CI 36.8-46.5) per 100 person-years. The 845 PWH with at least one ED visit during the study period contributed a median of 2 visits each (IQR 1-5). The unadjusted probability of having \u22651 ED visit in a given year was higher among women vs. men , Black vs. White PWH , with VL \u2265 40 copies/mL , and with CD4 < 200  or 200-349  vs. \u2265 500 cells/\u03bcL; age was not associated with ED use. Compared with 2017-2019, the annual probability of having \u2265 1 ED visit was lower in 2020-2021, with RRs of 0.83 (95% CI 0.76-0.90) in unadjusted analyses and 0.80 (95% CI 0.71-0.90) after adjusting for demographics, VL, and CD4. There was also a significant unadjusted decrease for 2020-2021 vs. 2017-2019 among women, men, PWH who were Black, White, < 40 or 50-59 years old, and with CD4 >500 (Fig.\u00a0Among PWH in HIV care, ED use was higher among women, Black PWH, and PWH with poorly controlled HIV. ED use decreased 2020-2021 in most groups, indicating that PWH during the COVID-19 pandemic may be delaying seeking care for acute conditions, or accessing care in other ways. Work is ongoing to characterize reasons for ED visits across calendar years and examine the impact of reduced ED utilization among PWH.Joseph J. Eron, MD, Adagio Therapeutics: data safety monitoring committee|Gilead Sciences: Advisor/Consultant|Gilead Sciences: Grant/Research Support|Glaxo Smith Kline: Advisor/Consultant|Merck: Advisor/Consultant|ViiV Healthcare: Advisor/Consultant|ViiV Healthcare: Grant/Research Support."}
+{"text": "Gait speed is a predictor of overall health and mortality in older adults. Metabolomics may provide insights into biological mechanisms underlying gait speed. Herein, we examined the association between 193 lipid metabolites with gait speed in 1,717 adults (52.1% women) aged 82.0 \u00b1 14.5. Lipidomic analysis was performed using liquid chromatography-mass spectrometry. Gait speed was measured over 4-meters and slowness was defined as <0.8m/s. Logistic regression, adjusted for age, sex, field center, height, fasting-duration and familial-relatedness, were used to examine the association between log-transformed metabolites with slowness. A false discovery rate (FDR) of p<0.05 was employed to account for multiple comparisons. Gait speed was 0.83 \u00b1 0.32 and 53.4% had slowness. Three lipid metabolites were significantly associated with lower odds of slowness: an acylcarnitine, sphingomyelin and a ceramide non-hydroxy fatty acid-sphingosine. Our results potentially link lipids involved with mitochondrial beta-oxidation and nerve signal transduction to gait speed in older adults."}
+{"text": "Klebsiella sp. CTHL.F3a was isolated from kimchi (Korean fermented cabbage/vegetables). Its complete genome sequence , comprising a chromosome and a single plasmid, was established through hybrid assembly.The cellulolytic strain  Klebsiella oxytoca species complex (KoSC) may be encountered as human commensals and opportunistic pathogens  in Bushnell-Haas medium (BHM) (Six samples of fresh kimchi (Korean fermented cabbage/vegetables) from Hong Kong markets were screened for cellulolytic bacteria using carboxymethylcellulose (CMC) agar, as described previously . Twelve um (BHM)  containiN50, 12,200\u2009bp). Default parameters were used for all software unless otherwise specified.Paired-end short-read sequencing libraries, prepared using the NexteraXT DNA library preparation kit, were sequenced via the Illumina MiSeq platform with v3 chemistry (2 \u00d7 300 bp). Adapter sequences were removed, and 1,416,362 raw reads were quality filtered and trimmed using Trimmomatic v0.32  to give dnaA/repA on the forward strand), with genome coverage of 81\u00d7, which were submitted to NCBI PGAP v5.0 (The Illumina and Nanopore datasets were combined using Unicycler v0.4.3  to yieldGAP v5.0  and to PGAP v5.0  for annohttps://www.ezbiocloud.net/tools/ani) of 99.60% (Klebsiella sp. BDA134-6 (GenBank accession number CP064784) (bcs operons (Klebsiella pneumoniae recovered from the gut of a Chinese bamboo rat (Rhizomys sinensis) (The CTHL.F3a chromosome has an average nucleotide identity (by OrthoANIu online at f 99.60%  with ricP064784) . It cont operons , punctua operons . Phyre2  operons  finds thinensis) .https://bioinfo-mml.sjtu.edu.cn/KpVR/index.php) (Plasmid pCTHL.F3a was classified as IncFIB(K)_30 (clade II) by the KpVR Web-based tool (dex.php) . It contdex.php)  but no aKlebsiella sp. CTHL.F3a are available through NCBI under BioProject PRJNA758781, with GenBank accession numbers CP082360 (chromosome) and CP082361 (plasmid) and SRA accessions SRX12151552 (MiSeq) and SRX12151553 (MinION). Klebsiella sp.Complete genome sequences and raw sequence data for"}
+{"text": "Klebsiella pneumoniae (CR-cKp) and carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp) isolates in southwestern Iran. From 2019 to 2021, 136 (88.9%) cKp and 17 (11.1%) hvKp isolates were identified using biochemical tests and polymerase chain reaction (PCR). Antibiotic resistance, beta-lactamases, and clonal relatedness of carbapenem-resistant isolates were investigated using disk diffusion, PCR, and enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR), respectively. The different markers of hvKp isolates were as follows: string test , magA , rmpA , rmpA2 , iucA , and peg344 . Also, 55.1% (n = 75/136) of cKp and 47.1% (n = 8/17) of hvKp isolates were CR-cKp and CR-hvKp, respectively. All CR-hvKp  isolates were MDR. Colistin, tetracycline, and tigecycline were the most effective antibiotics. The occurrence of beta-lactamase genes in 75 CR-cKp and 8 CR-hvKp isolates was as follows: blaNDM , blaIMP , blaVIM , blaGES , blaOXA\u201348\u2013like , blaCTX\u2013M , blaSHV , blaTEM , blaFOX , blaDHA , blaCMY , blaLAT , and blaACT . ERIC-PCR showed a high diversity among isolates. In this study, the occurrence of MDR CR-hvKp isolates harboring blaNDM and blaGES was detected for the first time in southwestern Iran. To prevent the spread of CR-hvKp and reduce selection pressure, long-term surveillance and more effective treatment strategies should be implemented.This study investigated the molecular epidemiology of carbapenem-resistant classic  Klebsiella pneumoniae is one of the most important Gram-negative bacteria (GNB) causing a variety of community-acquired and nosocomial infections. It is estimated that about one-third of all Gram-negative infections are caused by this bacterium (K. pneumoniae (hvKp) characterized by hypermucoviscosity was first reported from a liver abscess with extrahepatic complications, including endophthalmitis, in Taiwan in 1986. HvKp has been implicated in community-associated infections in healthy people with the most common cases of pyogenic liver abscesses in Asia (acterium . Hypervi in Asia .Klebsiella pneumoniae (cKp) strains from hvKp. These include the string test, the regulator of mucoid phenotype A and A2 (rmpA and rmpA2), an aerobactin siderophore (iucA), the peg-344 gene, and the mucoviscosity-associated gene A (magA) (Some phenotypic and genotypic distinctive properties and determining factors differentiate classic A (magA) . The strA (magA) . TherefoK. pneumoniae. A recent systematic review and meta-analysis from Iran found a prevalence rate of 0.004\u201358% for carbapenem-resistant K. pneumoniae (CR-Kp) in different regions of the country  GNB, including  country . Accordi country . HoweverK. pneumoniae include the production of various beta-lactamases capable of hydrolyzing carbapenems as well as the reduced membrane permeability that occurs via loss or downregulation expression of outer membrane porins (OMPs)  . CR-Kp is (OMPs) .Klebsiella pneumoniae carbapenemase (KPC), oxacillinases (OXA)-type enzymes such as OXA-48-like carbapenemases, Guiana extended-spectrum beta-lactamase (GES), imipenem-hydrolyzing beta-lactamase (IMI) and metallo-beta-lactamases (MBLs) including Verona integrin-encoded MBL (VIM), New Delhi MBL (NDM), and imipenemase (IMP) play a major role in the development of carbapenem resistance in Enterobacteriaceae (K. pneumoniae (Pseudomonas extended resistance), and VEB (Vietnamese extended-spectrum beta-lactamases) enzymes  affiliated to the Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran. Ahvaz is the capital of Khuzestan province which located at 31\u00b020\u2019 north latitude and 48\u00b040\u2019 east longitude, and ranks the second-largest city (with an area of about 215 km2) after Tehran, the capital of Iran , non-repetitive clinical isolates of  of Iran . The afoK. pneumoniae isolates was performed using a series of standard biochemical tests including Gram stain, triple sugar iron (TSI) agar, sulfur-indole-motility (SIM) agar, lysine iron agar (LIA), ornithine decarboxylase, citrate utilization, and urea hydrolysis  and incubated aerobically for 18\u201324 h at 37\u00b0C. Primary identification of drolysis . All medK. pneumoniae isolates were confirmed by polymerase chain reaction (PCR) using specific primers for the 16S\u201323S internal transcribed spacer (16S\u201323S ITS) gene (130 bp), as previously described  to determine the size of PCR products compared to a 100 bp DNA ladder. The agarose gel was scanned using a UV light transilluminator . Confirmed isolates were stocked in tryptic soy broth (TSB)  containing 20% glycerol and stored at \u201380\u00b0C for ongoing analysis.The presumptive escribed . To extrescribed . All PCRescribed  for 30 smagA, rmpA, rmpA2, iucA, and peg344 were investigated by PCR using previously described primers , ampicillin/sulbactam (10/10 \u03bcg), aztreonam (30 \u03bcg), cefazolin (30 \u03bcg), ceftriaxone (30 \u03bcg), ceftazidime (30 \u03bcg), cefotaxime (30 \u03bcg), cefoxitin (30 \u03bcg), cefepime (30 \u03bcg), chloramphenicol (30 \u03bcg), ciprofloxacin (5 \u03bcg), ertapenem (10 \u03bcg), gentamycin (10 \u03bcg), fosfomycin (200 \u03bcg), imipenem (10 \u03bcg), meropenem (10 \u03bcg), piperacillin/tazobactam (100/10 \u03bcg), tetracycline (30 \u03bcg), tobramycin (10 \u03bcg), and trimethoprim/sulfamethoxazole (1.25/23.75 \u03bcg)  were tested on Mueller\u2013Hinton agar (MHA)  using the Kirby-Bauer disc diffusion method according to Clinical Laboratory Standard Institute (CLSI) procedures . Also, tectively . Any isoectively . Multiplectively . EscheriK. pneumoniae isolates that had breakpoints of \u226427 mm for cefotaxime (30 \u03bcg), \u226422 mm for ceftazidime (30 \u03bcg), and \u226425 mm for ceftriaxone (30 \u03bcg), were selected for primary ESBL screening  were considered as primary AmpC producers. These isolates were further confirmed by a combined disk method using a cefoxitin disk (30 \u03bcg) alone and in combination with phenylboronic acid (PBA) (400 \u03bcg) . The cefoxitin disk and cefoxitin/PBA disk were dispensed onto an MHA plate that had already been inoculated with the test isolate  by the standard disk diffusion method. Subsequently, the inoculated plates were incubated for 24 h at 37\u00b0C. The isolates were confirmed as AmpC producers when the zone diameter of cefoxitin/PBA increased by \u22655 mm compared to cefoxitin alone .E. coli ATCC\u00ae 25922\u2122. After 18\u201324 h incubation at 35 \u00b1 2\u00b0C, mCIM and eCIM results were interpreted as follows: carbapenemase positive (only mCIM positive): zone diameter of 6\u201315 mm, carbapenemase negative (mCIM negative): zone size \u2265 19 mm, MBL positive (both mCIM/eCIM positive): increase in eCIM zone size by \u22655 mm compared to zone size of mCIM  together with EDTA-modified carbapenem inactivation method (eCIM) . For mCI of mCIM . It shoublaCTX\u2013M, blaSHV, blaTEM, blaPER, and blaVEB), AmpC , carbapenemase , and MBLs  as previously described  was performed with the previously described primers in a final volume of 25 \u03bcl containing 1 \u03bcl of each primer, 3 \u03bcl of template DNA, 12.5 of master mix , and 7.5 \u03bcl of nuclease-free water using the Biorad S1000 thermocycler (USA) . FollowiP-value \u2264 0.05) of variables were evaluated with Chi-square and Fisher\u2019s exact tests  version 22.0 . Significant associations  males and 71 females (46.4%). All isolates showed the 130 bp band of the 16S\u201323S ITS gene in PCR and were confirmed as K. pneumoniae. The mean \u00b1 SD age of patients was 43.1 \u00b1 16.3 (10\u201381 years) for males and 36.9 \u00b1 12.7 (11\u201382 years) for females. Using phenotypic and genotypic criteria, 136 (88.9%) and 17 (11.1%) isolates were identified as cKp and hvKp, respectively. Of all hvKp isolates, 6 (35.3%), 17 (100.0%), and 6 (35.3%) were positive for the string test, gene markers, and both, respectively. The occurrence of hvKp gene markers was as follows: magA , rmpA , rmpA2 , iucA , and peg344 . The co-occurrence of two or more markers was detected in 5 (29.4%) isolates as follows: magA/rmpA2 (n = 1), rmpA2/iucA/peg344 (n = 3), and magA/rmpA/rmpA2/iucA/peg344 (n = 1).Using standard bacteriology tests, a total of 153 presumptive n = 83/153), including 90.4% (n = 75/83) cKp and 9.6% (n = 8/83) hvKp isolates. In other words, 55.1% (n = 75/136) of cKp and 47.1% (n = 8/17) of hvKp isolates were CR-cKp and CR-hvKp, respectively. Imipenem was the most effective carbapenem against K. pneumoniae isolates, with a susceptibility rate of 48.4% (n = 74/153), followed by ertapenem , and meropenem . Of 75 CR-cKp isolates, 60 (80.0%), 8 (10.7%), and 7 (9.3%) isolates were resistant to ertapenem/imipenem/meropenem, ertapenem/meropenem, and imipenem/meropenem, respectively. However, all CR-hvKp isolates were simultaneously resistant to meropenem/imipenem/ertapenem. The carbapenem-resistant isolates had MICs ranging from 0.03 to 64 \u03bcg/mL, MIC50 = 8 \u03bcg/mL, MIC90 = 32 \u03bcg/mL for ertapenem; MICs ranging from 0.03 to 64 \u03bcg/mL, MIC50 = 16 \u03bcg/mL, MIC90 = 32 \u03bcg/mL for imipenem; and MICs ranging from 8 to 64 \u03bcg/mL, MIC50 = 16 \u03bcg/mL, MIC90 = 32 \u03bcg/mL for meropenem. The detailed antibiotic resistance rates of cKp, hvKp, CR-cKp, and CR-hvKp isolates were summarized in n = 150/153), tetracycline , and tigecycline  and the highest resistance to ampicillin/sulbactam . Resistance rates to other antibiotics ranged from 29.4% (n = 45/153) for aztreonam to 64.7% (n = 99/153) for cefazolin. CR-cKp and CR-hvKp isolates showed resistance rates of more than 50.0% against gentamicin, tobramycin, amikacin, and piperacillin/tazobactam. Also, all hvKp isolates (n = 17) including CR-hvKp (n = 8) strains were susceptible to colistin, tigecycline, and tetracycline. There were no significant differences  in the antibiotic resistance patterns of the cKp with hvKp isolates and the CR-cKp with CR-hvKp isolates except for tigecycline  of all isolates were resistant against third-generation cephalosporins. Of 136 cKp, 1 (0.7%), 1 (0.7%), 1 (0.7%), 3 (2.2%), and 81 (59.6%) isolates were resistant to cefotaxime, cefotaxime/ceftazidime, ceftazidime/ceftriaxone, cefotaxime/ceftriaxone, and cefotaxime/ceftazidime/ceftriaxone, respectively. Of 17 hvKp, 11 (64.7%) isolates were simultaneously resistant to cefotaxime/ceftazidime/ceftriaxone. Of 75 CR-cKp isolates, 69 (92.0%) strains showed resistance against third-generation cephalosporin as follows: cefotaxime , ceftazidime/ceftriaxone , cefotaxime/ceftazidime , cefotaxime/ceftriaxone , and cefotaxime/ceftazidime/ceftriaxone . Also, all CR-hvKp were simultaneously resistant to third-generation cephalosporins. In total, 66.7% (n = 102/153) and 7.8% (n = 12/153) of isolates were MDR and XDR, respectively. None of the isolates were PDR. Of 75 CR-cKp, 84.0% (n = 63) and 16.0% (n = 12) were MDR and XDR, respectively. While, all CR-hvKp  isolates were MDR (K. pneumoniae isolates had 57 (A1\u2013A57) different antibiotypes (antibiotic resistance patterns) (n = 10), A4 , and A5  were the most frequent patterns. Also, MARI ranged from 0.1 to 1.0 and the majority of isolates  had MARI of \u22650.5.Using the carbapenem antibiotic disk diffusion test and the broth microdilution method, the overall resistance to carbapenems was 54.2% (ecycline . The carwere MDR . The 83 atterns) . A2 (12.n = 71/136), male patients , patients aged 26\u201341 years , urine samples , and men ward . Meanwhile, the highest occurrence of hvKp isolates was found in Golestan Hospital , male patients , patients aged 42\u201357 years , urine samples , and men ward  . Similar results were obtained for CR-cKp and CR-hvKp isolates (n = 41/75) than in males . Also, the CR-hvKp isolates were more prevalent in intensive care unit (ICU)  than other wards. However, the distribution of CR-cKp and CR-hvKp was not significantly different according to the various items  . Althougisolates . Neverthus items .n = 69/75) and 100.0% (n = 8/8) of CR-cKp and CR-hvKp were presumptive ESBL producers, respectively (n = 20/75) and 12.5% (n = 1/8) of CR-cKp and CR-hvKp were ESBL producers, respectively. All CDT positive isolates were also positive for at least one ESBL gene by PCR. The distribution of ESBL genes among CR-cKp and CR-hvKp was as follows: for CR-cKp: blaCTX\u2013M , blaSHV , blaTEM ; and for CR-hvKp: blaCTX\u2013M , blaSHV , and blaTEM   . None ofn = 59/75) of CR-cKp and 100.0% (n = 8/8) of CR-hvKp were presumptive AmpC producers, the confirmatory cefoxitin (30 \u03bcg)/phenylboronic acid method was positive in only 13.3% (n = 10/75) and 0.0% (n = 0/8) of CR-cKp and CR-hvKp isolates, respectively (n = 21/75) of CR-cKp and none of CR-hvKp isolates were AmpC producers. The PCR method detected a greater number of AmpC positive CR-cKp isolates compared with the phenotypic confirmatory test. The distribution of AmpC genes among CR-cKp was as follows: blaLAT , blaACT , blaFOX , blaDHA , and blaCMY . The blaACC was not detected in any isolate  resistance criteria, 78.7% (ectively . However isolate . The disn = 26/75) and 25.0% (n = 2/8) of CR-cKp and CR-hvKp were carbapenemase producers, respectively , blaGES . Moreover, the occurrence of carbapenemase genes in CR-hvKp isolates was as follows: blaOXA\u201348\u2013like , blaGES  , 34.7% (ectively . PCR shon = 2/8) . The blan = 18/75) and 25.0% (n = 2/8) of CR-cKp and CR-hvKp were MBL positive, respectively (n = 33/75) of CR-cKp and 25.0% (n = 2/8) of CR-hvKp isolates were MBL producers. The distribution of MBL genes among CR-cKp was as follows: blaNDM , blaVIM , and blaIMP . Moreover, 25.0% (n = 2/8) of CR-hvKp isolates carried blaNDM gene , 24.0%  of CR-cKp and 25.0% (n = 2/8) of CR-hvKp isolates. Also, the blaCTX\u2013M/blaSHV was the second most prevalent pattern, detected in 6.7% (n = 5/75) of CR-cKp and 12.5% (n = 1/8) of CR-hvKp isolates. The frequency of co-occurrence of various beta-lactamases was as follows: MBL/carbapenemase/AmpC , MBL/carbapenemase/ESBL , carbapenemase/ESBL/AmpC , MBL/carbapenemase and MBL/AmpC , ESBL/carbapenemase, ESBL/MBL, and ESBL/AmpC , and carbapenemase/AmpC .The different genotypes of beta-lactam resistance genes were shown in n = 62/75) and 13 singletons  with 57 different ERIC-types (E1\u2013E57), indicating high genetic diversity among the isolates (blaNDM positive). The remaining isolates had different beta-lactamase genotypes. The eight CR-hvKp isolates were also divided into two clusters (A and B) and two singletons with four ERIC -types (E1\u2013E4)  consisting of 2\u20137 isolates  . ElectroK. pneumoniae isolates, including 136 (88.9%) cKp and 17 (11.1%) hvKp isolates, were identified in different clinical samples. Previous reports from Iran by n = 14/52), n = 22/146), and n = 3/122) showed different prevalence rates of hvKp than the current study. However, n = 12/111) from Turkey reported almost the same prevalence of hvKp as in the current study. These discrepancies can be explained by the different nature and size of the samples studied and the detection method of the hvKp isolates. To date, there are no standard methods to differentiate hvKp from cKp strains, and several studies classified hvKp based on positivity of the string test and some virulence genes including rmpA, rmpA2, iucA, peg-344, and magA (n = 6/17) of hvKp had a positive string test, and all  were also positive for at least one of the studied genes. However, 64.7% (n = 11/17) of hvKp isolates carrying gene markers had a negative string test. This phenomenon has also been reported in previous studies from Turkey and Iran , rmpA , rmpA2 , iucA , and peg344 . Simultaneous occurrence of two or more genes was also found in five (29.4%) isolates. In a previous study from Turkey, string test, iucA, and magA were positive in three (25.0%), eight  of the hvKp isolates, respectively. However, the rmpA and peg344 genes were not detected  of cKp and 47.1% (n = 8/17) of hvKp isolates were CR-cKp and CR-hvKp, respectively and showed various resistance rates against ertapenem, imipenem, and meropenem ranging from 47.1 to 54.4%  revealed that a relatively high proportion (more than 50.0%) of cKp and hvKp isolates were resistant to aminoglycosides, beta-lactam combination agents (piperacillin-tazobactam and ampicillin/sulbactam), quinolones, folate pathway inhibitors, and different classes of cephalosporins. These findings were consistent with recent reports of increasing emergence of highly resistant XDR and PDR cKp and hvKp strains from different countries, including Iran, Turkey, Spain, India, Saudi Arabia, and Lebanon . Generalto 54.4% . Howevern = 3/75) of CR-cKp isolates were resistant to colistin. These observations coincided well with previous reports from Iran  and 16.0% (n = 12/75) of CR-cKp were MDR and XDR, respectively. While all CR-hvKp  isolates were MDR (n = 70/83) had a MARI of \u22650.5 and no PDR isolate was detected. In contrast to the current study, in previous reports by In this study, the majority of CR-cKp and CR-hvKp isolates had high resistance rates (more than 60.0%) against 18 of 23 tested antibiotics, so that 84.0%  and CR-hvKp  isolates were identified from urine samples, followed by blood samples for CR-cKp  and tracheal tubes  for CR-hvKp. Also, the CR-hvKp isolates were more prevalent in the ICU ward  than in other wards. However, the distribution of CR-cKp and CR-hvKp did not differ significantly by sample type, hospital wards, and other demographic variables (P > 0.05)  in this study. The reason for this observation was unclear.In this research, most CR-cKp  . These f > 0.05) . Howevern = 20/75) and 12.5% (n = 1/8) of CR-cKp and CR-hvKp were ESBL producers, respectively. The PCR assay confirmed the results of the CDT phenotypic test and all CDT positive isolates harbored at least one ESBL gene. Also, the co-occurrence of ESBL genes was detected in both CR-cKp and CR-hvKp isolates, which was in line with previous reports from different countries , blaSHV , blaTEM ; and for CR-hvKp: blaCTX\u2013M , blaSHV , and blaTEM  , blaSHV , and blaTEM . However, similar to the findings of the current study, the blaPER and blaVEB were not detected , blaTEM , and blaCTX\u2013M . These frequencies were higher than in the current study. Moreover, in contrast to this research, the prevalence of blaSHV was significantly higher (P = 0.048) in hvKp isolates than in cKp strains (n = 9) harbored blaSHV and blaCTX\u2013M, whereas blaTEM was not detected. In this study, the blaCTX\u2013M was the most frequent ESBL among carbapenem-resistant K. pneumoniae isolates that was in line with the previous report from Iraq  . None ofdetected . Also, T strains . In anotrom Iraq . In the al ESBLs . Antibioal ESBLs .n = 21/75) of CR-cKp isolates as follows: blaLAT , blaACT , blaFOX , blaDHA , blaCMY , and blaACC . There is evidence for the role of AmpC beta-lactamase in resistance to carbapenems and increasing their MIC (blaCMY\u20132 (60.7%) and blaDHA\u20131 (34.4%) were found in K. pneumoniae isolates  of CR-cKp and 25.0% (n = 2/8) of CR-hvKp, were carbapenemase producers using mCIM test. All mCIM positive isolates carried at least one carbapenemase gene. The distribution of carbapenemase genes among CR-cKp and CR-hvKp was as follows: blaOXA\u201348\u2013like  and blaGES . Also, the blaKPC and blaIMI were not detected in any isolate. This study was the first to detect the blaGES carbapenemase in CR-hvKp isolates from Iran. In previous studies from Iran, blaGES was not detected in hvKp isolates (blaGES gene (27.8%) was the third most prevalent carbapenemase among carbapenem-resistant K. pneumoniae isolates  was reported to be the most prevalent gene in carbapenem-resistant K. pneumoniae isolates.In this study, 34.7%  of CR-cKp and 25.0% (n = 2/8) of CR-hvKp isolates were MBL positive using the mCIM/eCIM positivity criteria. PCR showed that all mCIM/eCIM positive isolates carried at least one MBL gene. However, PCR detected a greater number of MBL positive isolates than the phenotypic confirmatory test. This may be due to the co-occurrence of carbapenemases and MBLs, resulting in a false-negative eCIM test (n = 33/75) of CR-cKp and 25.0% (n = 2/8) of CR-hvKp isolates were MBL producers. The blaNDM gene was the most frequently detected MBL gene in CR-cKp (41.3%) and CR-hvKp isolates (25.0%), followed by blaVIM (8.0%) and blaIMP (4.0%) for CR-cKp. These results were in good parallel with available data that identified the blaNDM (30.1%) as the most predominant MBL type among K. pneumoniae isolates from Iran, followed by blaVIM (10.6%) and blaIMP (4.5%)  of hvKp isolates (blaNDM and blaOXA\u201348 genes were frequently associated with hvKp isolates (n = 6/75) was the most co-occurrence pattern among CR-cKp isolates. In contrast, CR-hvKp isolates had only the ESBL co-existence pattern (blaCTX\u2013M/blaSHV). Contrary to these results, the co-occurrence of ESBLs and carbapenemases in hvKp isolates has been reported in previous studies from Iran  . Howeverisolates . In mostisolates . This phisolates . Anotherrom Iran .K. pneumoniae isolates, several expensive and time-consuming molecular methods such as pulsed-field gel electrophoresis (PFGE), multiple locus sequence typing (MLST), and ribotyping are available  based analysis for phylogenetic relatedness, pan-genome analysis and insights on the circulating plasmids and Inc groups especially among the colistin-resistant isolates.blaCTX\u2013M, blaSHV, blaNDM, and blaGES, in southwestern Iran. As a result of their hypervirulence coupled with multidrug resistance, these isolates pose a particular threat to healthcare systems. Hence, long-term surveillance and more effective treatment strategies should be implemented to prevent the spread of CR-hvKp and reduce selection pressure. Also, it is recommended to use a fast and cheap molecular method such as ERIC-PCR for primary evaluation of clonal relatedness of MDR isolates.This study was the first report of emergence of MDR CR-hvKp isolates harboring different beta-lactamases, including The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.This study was approved by the Ethics Committee of the Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran (ethics code: IR.AJUMS.REC.1398.489) according to the Declaration of Helsinki. All methods were performed in accordance with the relevant guidelines and regulations of the Ethics Committee of the Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran. Clinical samples were not collected as part of this study. The clinical samples were collected as routine clinical care and to check the presence of any infection for referred and admitted patients. As a result, written informed consent was waived by the Ethics Committee of Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.MSak performed the experiments, analyzed majority of the data, and wrote the whole manuscript. MSav, MH, and MA prepared all the necessary materials and performed some of the experiments. MSav, MH, and SSS contributed to the discussion of experimental results. MA designed and supervised the experiments at different stages. All authors contributed to the article and approved the submitted version."}
+{"text": "Similarly, Arabidopsis thaliana SAUR63 was able to increase growth of various organs, antagonize PP2C.D5 phosphatase, and increase H+-ATPase activity. Using a gain-of-function approach to bypass genetic redundancy, we dissected structural requirements for SAUR63 growth-promoting activity. The divergent N-terminal domain of SAUR63 has a predicted basic amphipathic \u03b1-helix and was able to drive partial PM association. Deletion of the N-terminal domain decreased PM association of a SAUR63 fusion protein, as well as decreasing protein level and eliminating growth-promoting activity. Conversely, forced PM association restored ability to promote H+-ATPase activity and cell expansion, indicating that SAUR63 is active when PM-associated. Lipid binding assays and perturbations of PM lipid composition indicate that the N-terminal domain can interact with PM anionic lipids. Mutations in the conserved SAUR domain also reduced PM association in root cells. Thus, both the N-terminal domain and the SAUR domain may cooperatively mediate the SAUR63 PM association required to promote growth.In plants, regulated cell expansion determines organ size and shape. Several members of the family of redundantly acting Small Auxin Up RNA (SAUR) proteins can stimulate plasma membrane (PM) H Plant organs reach their final shape and size after substantial cell expansion. Proton pumps at the plasma membrane promote cell expansion by acidifying the cell wall to loosen it, and by increasing electrochemical potential across the plasma membrane for solute uptake that maintains intracellular turgor. Plasma-membrane-associated proteins tightly regulate proton pump activity, in order for organs to grow to an appropriate extent. We have studied requirements for activity of one such regulatory protein in the model plant Arabidopsis called SAUR63. This protein is made rapidly in response to plant growth hormones, and it increases proton pump activity to promote organ growth. These activities depend on its binding to anionic lipids in the plasma membrane, and forced plasma membrane association of SAUR63 can increase growth. Many proteins in the same family are found within Arabidopsis and in all land plants, and likely differ in their affinity for the plasma membrane or in other properties. Further studies of other family members may show how such proteins regulate growth under diverse physiological contexts. Small Auxin Up RNA (SAUR) genes. SAUR genes are numerous in all land plants, and have evolved through extensive independent gene amplifications in multiple lineages ) of seedlings of indicated genotypes grown for 3d in darkness on plates with 0.5x MS medium. C,D) Root length (C) and root tortuosity index (D) of seedlings of indicated genotypes grown for 4d in long days on plates with 1x MS 1% Sucrose medium. E,F) Root length (E) and root tortuosity index (F) of seedlings of indicated genotypes grown for 4d in short days on plates with 0.5x MS medium. Graphs show means \u00b1 s.d. Letters in graphs indicate statistical classes based on Tukey\u2019s Honestly Significant Difference test. n from left to right: Panels A,B: 51, 24, 26, 28, 23, 25, 25; Panels C,D: 34, 17, 23, 14, 19, 15, 16; Panels E,F: 43, 21, 18, 17, 16, 15, 17. The same genotypes were measured in panels A-F, with genotype designations shown only in panels E and F. G,H,I) Hypocotyl epidermal cells of seedlings of indicated genotypes grown in short days for 2 days, visualized with the ML1:RFP shoot epidermis plasma membrane marker. Shown are z-stack confocal image projections of the near side of the hypocotyl. Scale bar, 0.1 mm. (TIF)Click here for additional data file.S4 FigA) Cotyledon area of seedlings of indicated genotypes grown on vertically oriented plates for 7d in long days in the absence (open bars) or presence (filled bars) of 1% sucrose. B) Root length of seedlings of indicated genotypes grown on vertically oriented plates for 4d in long days in the absence (open bars) or presence (filled bars) of 1% sucrose. C,F) Hypocotyl lengths (E) and hypocotyl tortuosity index  of seedlings grown on vertically oriented plates for 3d in darkness without sucrose. D) Cotyledon areas of P35S:SAUR63 and P35S:CBL11-12:SAUR6326-142 seedlings grown for 6d on vertically oriented MS 1% Suc plates. E) Hypocotyl lengths of P35S:SAUR63 and P35S:CBL11-12:SAUR6326-142 seedlings grown for 4d on vertically oriented 0.5x MS plates. Panels D and E show data for the three homozygous single-locus P35S:SAUR63 lines that differed most from wild type among seven lines analyzed. Graphs show means \u00b1 s.d. Letters in graphs indicate statistical classes based on Tukey\u2019s Honestly Significant Difference test. n, from left to right: panel A: 26, 20, 25, 15, 22, 23, 24, 20, 22, 23, 19, 19, 21, 19, 21, 20; panel B: 19, 16, 18, 14, 20, 20, 21, 16, 21, 17, 19, 16, 18, 15, 16, 17; panel D: 77, 18, 17, 17, 19, 20; panel E: 119, 25, 24, 27, 21, 24; panels C and F: 35, 36, 45, 43, 47, 38, 41, 47.(TIF)Click here for additional data file.S5 FigA) Appearance of genotypes used for crosses presented in A and in B) Hypocotyl lengths of seedlings of indicated genotypes grown for 4d in short days in the absence of sucrose. Plants measured were F1 progeny of crosses of transgenic lines with each other or with wild-type Columbia, and were hemizygous for the indicated transgene(s). C) Appearance of PEST:SAUR63:CerFP:HA and wild-type seedlings grown with estradiol and in the absence or presence of 15 mM LiCl. Seedlings were grown for 3d under control conditions, and then transferred to plates with estradiol and with or without 15 mM LiCl, and grown for an additional 3d before imaging. Dots mark positions of root tips at the time of transfer to estradiol plates. Scale bar, 5 mm. D) Root growth of indicated genotypes in the absence (open bars) or presence (closed bars) of 15 mM LiCl. Seedlings were grown without LiCl for 5d, transferred to plates containing 0 or 15 mM LiCl, and root growth over the next three days was measured. E) HPTS fluorescence ratios around root cells of indicated genotypes. Data are pooled from measurements taken on three different days, each normalized to the average of wild-type values on those days. Graphs show means \u00b1 s.d. Letters in graphs indicate statistical classes based on Tukey\u2019s Honestly Significant Difference test. n, from left to right: panel B: 28, 24, 24, 30, 26, 27, 26, 25, 26, 25, 24, 20, 24, 26, 25; panel D: 21, 27, 16, 23, 19, 25, 20, 30, 18, 28, 21, 30, 23, 31, 18, 29; panel E: 26, 13, 14, 16.(TIF)Click here for additional data file.S6 FigA-D)P35S:SAUR63:YFP:HA. E-H)P35S:SAUR6326-142:YFP:HA. I-L)PUBQ10:WAVE 138Y expressing a PM-localized YFP fusion protein. M-P)PUBQ10:WAVE 1Y expressing a cytoplasmically-localized YFP fusion protein. Q-T)PUBQ10:WAVE 9Y expressing a YFP fusion protein localized to the tonoplast. Shown are fluorescence confocal microscopy images of YFP , FM4-64 membrane staining , and both channels together  with vertical yellow lines indicating locations of quantitation of fluorescence intensity signals, scaled to the maximum signal along the line . Image color channel brightnesses were adjusted for visibility. Scale bar, 20 \u03bcm.(TIF)Click here for additional data file.S7 FigA-M) Confocal images showing YFP fluorescence of transgenic lines expressing the indicated fusion proteins behind the P35S promoter. Scale bar, 20 \u03bcm.(TIF)Click here for additional data file.S8 FigA-L) Confocal images showing YFP fluorescence of indicated SAUR63:YFP:HA variants expressed in transiently transformed N. benthamiana leaves. Scale bar, 20 \u03bcm.(TIF)Click here for additional data file.S9 FigA-E) Protein levels in multiple P35S:SAUR63:YFP:HA, P35S:SAUR6326-142:YFP:HA, and P35S:CBL11-12:SAUR6326-142:YFP:HA pooled T2 seedlings from different T1 transformants. The blot in panel E shows protein extracts from selected genotypes in panels A-D for side-by-side comparison in the same experiment. F,G) SAUR63:YFP:HA, SAUR6326-142:YFP:HA, SAUR631-25:YFP:HA, CBL11-12:SAUR6326-142:YFP:HA, and SAUR63m2:YFP:HA fusion proteins, detected by western blots in total (T), soluble (S) and microsomal (M) protein fractions. Lower panels show controls for loading or fractionation. \u03b1-HA detects SAUR63 fusion protein; \u03b1-AHA2 detects a membrane protein; \u03b1-UGPase and \u03b1-APX detect soluble proteins. Arrows indicate position of full-sized SAUR63:YFP:HA fusion proteins. FL, Full-length SAUR63:YFP:HA fusion protein. wt, wild-type Columbia lacking any transgene. In genotype designations, S63 is short for SAUR63. Letters and numbers after genotype designations indicate independent transgenic lines. H) Amount of SAUR63:YFP:HA and SAUR6326-142:YFP:HA proteins in whole seedling extracts at indicated times after start of cycloheximide treatment to block new protein synthesis. Fusion proteins were detected by western blots using anti-HA antibody. The Rubisco large subunit band from Ponceau S staining of the same gels is shown as a loading control. A repeat of this experiment gave a very similar result. In the lower gel, the larger band is the presumed intact SAUR6326-142:YFP:HA protein, and the lower band is a presumed smaller breakdown product. In genotype labels, S63 denotes SAUR63, letters and numbers after genotype names indicate distinct transgenic lines, FL denotes a strong full-length P35S:SAUR63:YFP:HA line used as a common standard line in most experiments, and wt indicates wild-type Columbia lacking any transgene.(TIF)Click here for additional data file.S10 FigA) Western blot showing presence of fusion proteins in extracts used in lipid blot experiments in 26-142:YFP:HA fusion proteins (upper arrow) and SAUR631-25:YFP:HA N-terminal domain fusion protein (lower arrow). B) Longer exposures of two lipid blots from C) Mock experiment in which extracts were incubated for 70 minutes in protein extraction buffer at the indicated temperatures, and then run on a gel for western blots. For both SAUR63:YFP:HA and SAUR6326-142:YFP:HA, similar amounts of protein are present after incubation at -20 C or after incubation at 22 C, as during the lipid blot binding experiment.(TIF)Click here for additional data file.S11 FigA)PEST:SAUR63NAAIRS:CerFP:HA lines grown on plates with estradiol  or without estradiol (wild-type Columbia and PEST:SAUR63:CerFP:HA only). B) Western blots of total protein in estradiol-induced PEST:SAUR63NAAIRS:CerFP:HA lines using \u03b1-HA antibody.(TIF)Click here for additional data file.S12 FigA-D)P35S:SAUR63:YFP:HA. E-H)P35S:SAUR6326-142:YFP:HA. I-L)P35S:SAUR63m9:YFP:HA. M-P)P35S:SAUR63m13:YFP:HA. Q-T)P35S:SAUR63m15:YFP:HA. Shown are fluorescence confocal microscopy images of YFP , FM4-64 membrane staining , and both channels together  with vertical yellow lines indicating locations of quantitation of fluorescence intensity signals, scaled to the maximum signal along the line . Image color channel brightnesses were adjusted for visibility. Panels A-D are the same as in (TIF)Click here for additional data file."}
+{"text": "Plasmodium vivax apical membrane antigen-1 (pvama-1) is an important vaccine candidate against Malaria. The genetic composition assessment of pvama-1 from wide-range geography is vital to plan the antigen based vaccine designing against Malaria.P. vivax positive malaria patients from different districts of Khyber Pakhtunkhwa (KP) province of Pakistan. The highly polymorphic and immunogenic domain-I (DI) region of pvama-1 was PCR amplified and DNA sequenced. The QC based sequences raw data filtration was done using DNASTAR package. The downstream population genetic analyses were performed using MEGA4, DnaSP, Arlequin v3.5 and Network.5 resources.The blood samples were collected from 84 pvama-1 (DI) in KP samples with majorly prevalent H-14 and H-5 haplotypes. Pairwise comparative population genetics analyses identified limited to moderate genetic distinctions among the samples collected from different districts of KP, Pakistan. In context of worldwide available data, the KP samples depicted major genetic differentiation against the Korean samples with Fst\u2009=\u20090.40915 , while least distinction was observed against Indian and Iranian samples. The statistically significant negative values of Fu and Li\u2019s D* and F* tests indicate the evidence of population expansion and directional positive selection signature. The slow LD decay across the nucleotide distance in KP isolates indicates low nucleotide diversity. In context of reference pvama-1 sequence, the KP samples were identified to have 09 novel non-synonymous single nucleotide polymorphisms (nsSNPs), including several trimorphic and tetramorphic substitutions. Few of these nsSNPs are mapped within the B-cell predicted epitopic motifs of the pvama-1, and possibly modulate the immune response mechanism.The analyses unveiled total 57 haplotypes of pvama-1 DI among the P. vivax isolates acquired from widespread regions of KP province of Pakistan. The information may implicate in future vaccine designing strategies based on antigenic features of pvama-1.Low genetic differentiation was observed across the The online version contains supplementary material available at 10.1186/s12879-022-07798-1. Plasmodium. The P. vivax and P. falciparum are predominant species responsible for malaria [P. vivax is most widely distributed human malaria parasite, endemic in tropical and subtropical countries of Asia, South Pacific, Central and South America, Middle East, and North Africa [Malaria is an acute febrile infectious disease caused by vector-borne apicomplexan parasites of the genus  malaria . The P. h Africa . Accordih Africa .P. vivax [Plasmodium species such as apical membrane antigen-1 (AMA-1), Circumsporozoite proteins (CSP), Merozoite surface proteins (MSP) and Duffy binding protein (DBP) are reported as potent malarial vaccine candidates\u2019 targets [Treatment and control of malaria have become a serious challenge due to drug resistance and lack of effective vaccines. The wide-range distribution, antigenic variation, relapsing and co-infection led to a collective interest towards the development of effective vaccine against P. vivax . The impP. vivax . Several targets .Plasmodium species as promising malaria vaccine candidate antigens [Plasmodium parasites [P. falciparum and P. vivax [The genetic composition assessment of vaccine candidates\u2019 loci is indispensable in modern-age to plan an effective vaccination strategy. There are ample of studies suggesting the AMA-1 of antigens . The AMAarasites , 9. The arasites \u201312. The arasites . The ectP. vivax \u201317. FurtP. vivax . This suPlasmodium vivax ama-1 (pvama-1) have been conducted in malaria endemic countries [pvama-1 genetic features from Pakistan. Particularly, no study till date is reported from remote malaria endemic regions of Khyber Pakhtunkhwa (KP) province of Pakistan. The current study was therefore designed to evaluate the genetic composition of pvama-1 among P. vivax isolates collected from widespread KP regions of Pakistan . Blood samples were obtained from 100 consented patients tested positive for pvama-1 was amplified by polymerase chain reaction (PCR) using the specific primers and amplification conditions as reported previously [Escherichia coli DH5\u03b1 competent cells, and positive clones were selected by colony PCR. The nucleotide sequence of cloned insert was analyzed by automatic DNA sequencing with M13 forward and reverse primers . The raw data was filtered for quality assessment using DNASTAR Lasergene package.A DNA fragment flanking the DI region of eviously , 29. Thepvama-1 sequence i.e. Sal-I (AF063138) and Genbank-deposited pvama-1 sequences from China Myanmar Boarder (KX495505\u2013KX495577), Iran (KF422636.1\u2013KF422681.1), Korea (KM230319.1\u2013KM230384.1), Myanmar (FJ157248.1\u2013FJ157285.1), Papua New Guinea (PNG) (KC702402.1\u2013KC702501.1), Sri Lanka (EF218679.1\u2013EF218701.1), Venezuela (EU346015.1\u2013EU346087.1), Thailand (FJ784891.1\u2013FJ784990.1), and India (EU282774.1\u2013EF025196.1). The comparative sequences analyses were performed using MEGA4 software suite [The DNA sequences data generated in the current study was analyzed in comparison with reference pvama-1 with a threshold score of\u00a0\u2265\u20090.5. The higher BepiPred score predicts higher binding affinity of epitopes with immune receptors. The non-synonymous SNPs (nsSNPs) mapping within the top predicted epitopes of pvama-1 was checked. The intrinsically unstructured regions (IURs) and RBC binding sites within the pvama-1 have previously been characterized [pvama-1. Additionally, the positive selection sites in B-cell epitopes were identified via the maximum likelihood method of Codeml [The BepiPred-2.0  server wcterized , 32; andf Codeml  implemenf Codeml .2 index via DnaSP [D* and F* indices were calculated via a sliding window method using DnaSP. The population genetics statistical analyses, including pairwise fixation index (Fst), analysis of molecular variance (AMOVA), haplotype frequencies, and nucleotide diversity based on Nei\u2019s net distance (DA) were computed using Arlequin v3.5 [The DnaSP v6.12 software package  was usedia DnaSP . The Tajuin v3.5 . The hapuin v3.5 .pvama-1, flanking the DI domain were amplified from genomic DNA of 84 P. vivax positive samples. The sequences data spanning the 322\u2013737 nucleotide positions of the reference pvama-1 sequence i.e. Sal-I (AF063138). The analyses identified a large numbers of single nucleotide polymorphisms (SNPs) in KP samples. Among these, the 68 were nsSNPs, i.e. causing amino acid substitutions, including 53 dimorphic, 10 trimorphic, 3 tetramorphic, and 2 pentamorphic nsSNPs. The two pentamorphic amino acid changes observed were R112K/T/E/S and S228D/N/R/K. The ten trimorphic amino acid substitutions include the N132D/G, A141E/G, E145A/G, K190E/Q, T191K/P, A199T/V, S209G/C, P210S/L, P223L/S, and V233L/P. While the three tetramorphic amino acid changes are K120R/S/G, E189N/K/G, and E227V/K/G. These amino acid substitutions were observed at varied frequencies in the KP samples. Among the 68 nsSNPS, 59 have previously been reported in literature for P. vivax isolates from different geographical origin. However, the rest of 9 nsSNPs were found specific to KP samples set of this study. These nsSNPs were observed at low frequencies, i.e. 1.1 to 1.19%. Few nsSNPs, including K120R, N132D, L140I, A141E, K190E, E227V, and S228D were commonly observed in KP samples, as well as in some other continental pvama-1 sequences with high frequency of 3.8\u2013100%  of 0.978\u2009\u00b1\u20090.008 . The difference between dN/dS ratio for pvama-1 DI region was also found negative (\u2212\u20090.05413\u2009\u00b1\u20090.02) in case of KP samples set.The 416\u00a0bp sequences of pvama-1 DI\u00a0 were compared to the global pvama-1 sequences deposited in Genbank. The values of K and \u03c0 observed for KP sequences were more or less similar to previously reported sequences from Iran and India, however different from the rest of global sequences , followed by Swabi and Kohat  samples. The lowest Fst was estimated between Swat and Bannu samples , followed by Swat and Hungo samples   and mean pairwise differences (\u03c0xy) was observed between Bannu and Swabi samples, i.e. congruent to the Fst result . The Korean samples showed significant genetic distinction in pairwise comparison to rest of the global samples as well. Meanwhile, least genetic differentiation was observed among KP, Iranian, and Indian samples . Likewise, higher variance component was noticed within the population group i.e. 3.67534 as compare to among populations  pattern of pvama-1 DI sequences of P. vivax isolates from KP, Pakistan. The LD index (R2) (Y-axis) plotted against nucleotide distance (X-axis) using a two tailed Fisher\u2019s exact test.Additional file 4: Fig S3. Pearson correlation plot of KP and global pvama-1 samples based on pairwise Fst values. The plot shows clustering and correlation between the groups in hierarchical order. The dark brackets and large sizes depict the minimum genetic distinction and high correlation.Additional file 5: Fig S4. The principle component analysis (PCA) of pvama-1 DI sequences. Different colors depict different populations groups. This include KP Pakistan (as a single group) and worldwide samples."}
+{"text": "Subjects and Methods. The coexpression of PD-L1/PD-1 with CXCR3/CD36 on circulating lymphocytes was analyzed by flow cytometry in 78 lymphoma patients before and after therapy and in 50 healthy controls. The concentration levels of IL-19 in serum were assessed by an ELISA. Results. PD-L1 and PD-1 were expressed on circulating CXCR3+ and CD36+ lymphocytes in lymphoma and were significantly higher in patients with extranodal involvement than in lymphoma patients without extranodal involvement (P < 0.001). Elevated IL-19 levels were observed in lymphoma patients and increased significantly in extranodal involvement (P < 0.001). High percentages of PD-L1+CXCR3+ and PD-1+CXCR3+ lymphocytes were associated with high LDH levels, hepatomegaly, lymphedema, advanced tumor stage, and recurrence. Furthermore, patients with splenomegaly and generalized lymphadenopathy had high percentages of PD-L1+CXCR3+ lymphocytes. In addition, levels of PD-L1/PD-1 coexpression with CXCR3 and IL-19 were significantly associated with bone marrow, lung, and lymph vessel involvement. Further analysis revealed that high percentages of PD-L1+CD36+ and PD-1+CD36+ lymphocytes were associated with lung and bone marrow involvement. Patients with high levels of PD-L1/PD-1 coexpression with CXCR3 and IL-19 had inferior event-free survival (EFS) compared with that in lymphoma patients with low levels. EFS was decreased in patients with high percentages of PD-L1+CD36+ and PD-1+CD36+ lymphocytes. When using the receiver operating characteristic (ROC) curve, the superiority of IL-19 (area under the curve (AUC): 0.993) and PD-L1+CXCR3+% (AUC: 0.961) to PD-1+CXCR3+% (AUC: 0.805), PD-L1+CD36+% (AUC: 0.694), and PD-1+CD36+% (AUC 0.769) was evident in the diagnosis of extranodal involvement, identifying lymphoma patients with extranodal involvement from patients without extranodal involvement. Conclusions. Coexpression of PD-L1/PD-1 with CXCR3/CD36 in circulating lymphocytes and serum IL-19 levels contributes to poor prognosis and might be potential markers for extranodal involvement in lymphoma.Many studies have demonstrated that PD-L1/PD-1 signaling is an immune evasion mechanism in tumors. PD-L1/PD-1 coexpression with CXCR3/CD36 in peripheral lymphocytes in lymphoma still needs to be clarified. The current study investigated PD-L1/PD-1 coexpression with CXCR3/CD36 in circulating lymphocytes, serum IL-19 levels, and their correlation with clinical outcome and extranodal involvement in lymphoma.  Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) are the two forms of lymphoma , 2. Bothy , 8. The y .T cells express the regulatory inhibitory protein programmed death-1 (PD-1), a transmembrane element . MacrophCXCR3, a G protein-coupled cell surface receptor (GPCR), is present on CD4+ and CD8+ lymphocytes surface and other cells, like epithelial cells . Still, Cluster differentiation 36 (CD36) is a scavenger receptor on the surface of monocytes, adipocytes, dendritic cells, macrophages, and lymphocytes \u201333. CD36Interleukin-19 (IL-19) is a member of the IL-10 family . EssentiTumors have developed various strategies, disrupting \u201cimmune checkpoints\u201d to get through the host's immune system . The stuA total of 78 lymphoma patients and 50 healthy volunteers participated in the study. Healthy controls included thirty-six males and fourteen females. Healthy volunteers' range in age was from 26 to 70 years old. Seventy-eight lymphoma subjects ranged in age from 11 to 81 years, including 39 men and 39 women. Patients received treatment as soon as the primary diagnosis is confirmed. Patients with lymphoma were followed, and the patients were separated into two groups based on how well they responded to treatment: group I with no extranodal involvement and group II with extranodal involvement. Lymphoma patients' clinical outcomes were evaluated according to Cheson response criteria  . AccordiA complete clinical examination was performed on all subjects to monitor lymphadenopathy and hepatosplenomegaly, and detailed history questionnaires were completed. To determine the performance, type, and stage of lymphoma, all patients underwent bone marrow aspiration and lymph node biopsy. The Ann Arbor classification has been considered for clinical staging . The modFollow-up for lymphoma patients was performed at the hematology clinic and by telephone. The posttherapy lymphoma status was evaluated in lymphoma patients by means of clinical examination and PET/CT scans. Patients will be evaluated every three months for the identification of lymphoma progression. The follow-up was only for 78 patients with lymphoma out of 92 patients. The follow-up could not be performed for these patients as some patients were referred to other centers, some have died, and some had rejected blood samples rewithdrawal. Event-free survival (EFS) measures the time between the end of therapy and the commencement of an event . Seven follicular cell lymphoma (FCL) and twenty-eight diffuse large B cell lymphoma (DLBCL) patients' subjects had CHOP therapy  . In contAll subjects provided peripheral samples, which were taken under very sterile conditions. Eight milliliters of blood were collected. For a complete blood count (CBC), 2\u2009mL of blood was put into an EDTA tube. For flow cytometric analysis, 2\u2009mL of blood was inserted into an EDTA tube. A plain tube was filled with 4\u2009mL of blood, then centrifuged for 5 minutes at 3000\u2009rpm. Serum was isolated and used to analyze liver function tests , and aspartate aminotransferase (AST)), lactate dehydrogenase (LDH), kidney function tests (serum creatinine and blood urea), and random blood sugar. The leftover serum was kept at -70\u00b0C until the serum IL-19 concentrations were measured.A computerized hematology analyzer was used to determine CBC . Renal function analysis (blood urea nitrogen and serum creatinine), liver function analysis , and random blood sugar were performed using an autoanalyzer . An automated ACE chemistry analyzer assessed LDH .PD-1/PD-L1 coexpression with CXCR3/CD36 was determined in collected blood samples. To identify the various immune cells, the following human monoclonal antibodies were used, as directed by the manufacturer: PD-1 , CXCR3 monoclonal antibody , PD-L1 , and CD36 monoclonal antibody . Unstained cells were utilized as a negative control for every patient. Negative isotypic controls were performed using other tubes. As isotype controls, monoclonal PE-conjugated IgG2a  and FITC IgG1 were used .The percentages of PD-L1+CXCR3+, PD-1+CXCR3, PD-L1+CD36+, and PD-1+CD36+ lymphocytes were calculated using flow cytometry BD FACSCanto II . In brief, a hundred microliters of anticoagulated-EDTA whole blood were stained with five uL of monoclonal antibodies and incubated at room temperature in the dark for twenty minutes. The cells were lysed using the lysing buffer and set aside for about ten minutes at room temperature in the dark. Cells were then washed two times with PBS and resuspended in 300\u2009uL of PBS solution. A minimum of 10,000 events were analyzed. Lymphocyte gating was carried out through the FSC/SSC plots (front scatter vs. side scatter technique) \u201360, and The concentrations of IL-19 in serum were assessed using an ELISA kit  following the manufacturer's instructions. In the 96 wells of the microtiter strips, a specific monoclonal for IL-19 was coated (sensitivity for IL-19: 1.3\u2009pg/mL). A microtiter plate reader was used to determine optical densities at 450\u2009nm.The data were analyzed by applying the SPSS application  version 25. Normally, quantitative data was analyzed by minimum and maximum range and mean and standard deviation (SD). The median and interquartile range (IQR) was utilized for quantitative nonparametric data, while percentage and number were employed for categorical data. Mann\u2013Whitney analysis was carried out to analyze quantitative nonparametric data between two groups. Kruskal-Wallis test was carried out to analyze nonparametric data between more than two groups, proceeded by pairwise comparisons between each two groups applying Bonferroni correction. Fisher's exact test, or the Chi-square test, was carried out to compare the qualitative data between groups. Association between continuous and qualitative ordinal variables was assessed by Spearman's correlation, while Pearson's correlation was performed for the association between 2 continuous variables.P values less than 0.05 were considered significant.The Kaplan-Meier analysis was carried out to assess EFS, comparing the survival curves using the log-rank test. The variables' cutoff point, area under the curve (AUC), specificity, and sensitivity were calculated using the receiver operator characteristic (ROC) curve. The study included 50 healthy individuals and 78 lymphoma patients. Normal controls included thirty-six males and fourteen females. Healthy volunteers ranged in age from 24 to 81 years old. Seventy-eight lymphoma subjects ranged in age from 11 to 81 years, including 39 males and 39 females. The criteria for all subjects are listed in (Supplementary Table N = 34) and patients with extranodal involvement (N = 34). Lymphoma patients with and without extranodal involvement revealed statistical significance regarding sex (P = 0.039), stage (P < 0.001), recurrence (P < 0.001), and death (P = 0.018). Among lymphoma subjects with extranodal involvement, 73.5% had a recurrence, and 20.6% died. 26.5% of patients with extranodal involvement presented with stage III diseases, while 58.8% had stage IV diseases. Only 2.9% of patients with extranodal involvement had stage I, and 11.8% had stage II   .P < 0.001). Interestingly, PD-L1+CD36+% and PD-1+CD36+% in newly diagnosed lymphoma subjects were significantly higher than in healthy volunteers   .n = 34) or without extranodal involvement (n = 44) (P < 0.001) (P < 0.001)  . Postthe< 0.001) . When co< 0.001) .P < 0.001, respectively)  . Moreover, lymphoma patients had higher posttherapy IL-19 levels than the normal controls   .P < 0.001) (P < 0.001) , which was significantly higher than that of subjects without extranodal invasion with a median of 46.5\u2009pg/mL (range: 33-137.5) (< 0.001) . Interes< 0.001) .P < 0.05). Furthermore, compared to the PR, recurrence, and refractory groups, the CR group had a significant reduction in posttherapy IL-19, CXCR3+%, PD-L1+CXCR3+%, and PD-1+CXCR3+% (P < 0.05). Additionally, PD-L1+CD36+% and PD-1+CD36+% were significantly lower in subjects with CR compared to the other groups. Contrarily, comparing the CR group to the other groups, there was a substantial rise in posttherapy CD36+%   .P < 0.001). However, no significant difference was observed when subjects with CR were compared to the treatment-refractory patients (P = 0.092 and P = 0.055). In addition, the CR patients' PD-L1+CD36+% and PD-1+CD36+\u2009% were significantly lower than those in the PR group (P = 0.037 and P = 0.002). However, no significant difference was detected between the CR group and the recurrence group or the refractory group (P > 0.05) (The CR group showed a significant reduction in pretherapy PD-1+CXCR3+% and posttherapy PD-L1+CXCR3+% compared to the PR group and recurrence group ( > 0.05) .P < 0.001). Additionally, the recurrence group's posttherapy IL-19 levels, PD-L1+CXCR3+, and PD-1+CXCR3+% were significantly higher than those of PR patients  (P = 0.022 and P = 0.002). However, no significant differences were observed when the treatment-refractory group compared to PR or recurrence groups (P > 0.05)  . Further > 0.05) .r = 0.344, P = 0.002; r = 0.375, P = 0.001; r = 0.315, P = 0.005, respectively). However, negative associations between CXCR3+%, PD-L1/PD-1+CXCR3%, and albumin levels were identified . Additionally, there was a negative association between PD-L1+CXCR3+% and hemoglobin   .r = 0.464, P < 0.001; r = 0.398, P < 0.001, r = 0.335, P = 0.003). Data revealed that CXCR3+%, PD-L1+CXCR3+%, and PD-1+CXCR3+% positively correlated with lymphoma stages   and general lymphadenopathy . Furthermore, CXCR3+% and PD-L1+CXCR3+% positively correlated with BCR-ABL . Additionally, CXCR3+%, PD-L1+CXCR3+%, and PD-1+CXCR3+% had a significant association with lymphoma recurrence  (P > 0.05)  . Moreove= 0.006) . CD36+%, > 0.05) .r = 0.349 and P = 0.002), hepatomegaly (r = 0.362 and P = 0.001), and splenomegaly (r = 0.231 and P = 0.042). Furthermore, pretherapy IL-19 levels were significantly associated with lymphedema, general lymphadenopathy, and recurrence   .r = 0.771, P < 0.001; r = 0.793, P < 0.001; r = 0.528, P < 0.001). CXCR3+%, PD-L1+CXCR3+%, and PD-1+CXCR3+% had a positive association with bone marrow involvement . Moreover, CXCR3+%, PD-L1+CXCR3+%, and PD-1+CXCR3+% were positively associated with lymph vessel involvement . Additionally, a significant association between CXCR3+%, PD-L1+CXCR3+%, PD-1+CXCR3+%, and lung involvement was identified   .r = 0.333, P = 0.003; r = 0.469, P < 0.001), while CD36+% had a negative association with extranodal involvement (r = \u22120.792 and P < 0.001). Of interest, PD-L1+CD36+% and PD-1+CD36+% were positively correlated with bone marrow infiltration . PD-1+CD36+% was positively correlated with lung involvement . CD36% was negatively correlated with bone marrow and lymph vessel involvement . Furthermore, percentages of CD36+ lymphocytes were correlated with spleen infiltration   .r = 0.848; P < 0.001). IL-19 levels are positively associated with bone marrow and lymph vessel involvement . Similarly, a positive association between posttherapy IL-19 levels and lung involvement was identified   .The effect of the markers on EFS was assessed using Kaplan-Meier statistics . SurvivaP < 0.001)   (P < 0.001) (P < 0.001)  . High pr< 0.001) . A prolo Figures .The effect of the immune markers on overall survival and recurrence-free survival was determined using Kaplan-Meier statistics (data not shown). Patients with low pre- and posttherapy CXCR3+%, PD-L1+CXC3+%, and PD-1+CXCR3+% do better than those with a high percentage. Furthermore, low PD-L1+CD36+% and PD-L1+CD36+% predicted a more prolonged survival and recurrence-free time.P < 0.001, respectively) . The pretherapy IL-19 cut-off was >209\u2009pg/mL, with specificity and sensitivity 75% and 91.18%  at a cutoff >49, >40, and >3, respectively . Posttherapy CD36+% had the best sensitivity of 100% and the best specificity of 95.45% . Posttherapy CD36+%, PDL-1+CD36+%, and PD-1+CD36+% had cutoff levels \u226437, > 11, and>3 , with a cutoff >280\u2009pg/mL for diagnosis of extranodal involvement. IL-19 showed 100% sensitivity and 97.73% specificity . The specificity and sensitivity were 97.73% and 100% at a cut-off >280\u2009pg/mL.PD-L1/PD-1 coexpression with CXCR3/CD36 in identifying patients with extranodal involvement was assessed using ROC curves. The AUCs of the pre- and posttherapy PD-L1+CXCR3+% were 0.981 and 0.961, respectively, with high specificity and sensitivity. The cut-off values were >33 and >40. CXCR3+% and PD-L1+CXCR3+% yielded the best sensitivity and specificity. Moreover, the ROC curve was assessed for posttherapy PD-L1/PD-1+CD36+ percentages. The AUCs were 0.694 and 0.769, with reduced sensitivity and specificity. Thus, according to the findings, PD-L1/PD-1+CD36+% is insufficient for identifying extranodal involvement in lymphoma patients. Furthermore, the pretherapy IL-19 ROC curves demonstrated a pattern of extranodal involvement (This study had some limitations: (1) the small size of the subjects in this study; (2) short follow-up time, longer follow-up, and multicenter collaborations are needed to confirm PD-L1/PD-1's role; (3) PD-L1/PD-1 coexpression with CXCR3/CD36 should be investigated in tumor tissue; (4) cell function activities such as differentiation, proliferation, apoptosis, and cytokine release were not carried out; future research will investigate these issues; (5) PD-L1/PD-1 coexpression with CXCR3/CD36 was not investigated in different lymphocyte subsets; further studies are required.In conclusion, PD-L1/PD-1 coexpression with CXCR3/CD36 and serum IL-19 may be involved in lymphoma extranodal involvement and have prognostic and predictive values in lymphoma. The findings could also shed light on the role of circulating CXCR3 and CD36-positive lymphocyte cells in lymphoma. Future clinical trials and research are required to create new treatments based on PD-L1/PD-1-induced lymphoma immune evasion mechanisms and host immune response regulation. The combination of PD-L1/PD-1 blockades, anti-CXCR3/CD36, and IL-19 monoclonal antibody therapy might start a new era for immunotherapy. PD-L1+CXCR3+ lymphocytes and serum IL-19 might play a more important role in poor clinical behavior in lymphoma."}
+{"text": "The second paragraph of Acknowledgments should read as follows: \u201cWe gratefully acknowledge funding from the following sources: Czech Science Foundation grants 20-23513S to H.H., 18-17529S to A.Z., and 20-04150Y to O.G.; Czech Ministry of Education grant OPVVV16_019/0000759; and Czech BioImaging grant LM2015062.\u201dVolume 6, no. 3, e00327-21, 2021,"}
+{"text": "Brassica rapa, we identified 28 putative HSP70 gene family members using state-of-the-art bioinformatics-based tools and methods. Based on chromosomal mapping, HSP70 genes were the most differentially distributed on chromosome A03 and the least distributed on chromosome A05. Ka/Ks analysis revealed that B. rapa evolution was subjected to intense purifying selection of the HSP70 gene family. RNA-sequencing data and expression profiling showed that heat and cold stress induced HSP70 genes. The qRT-PCR results verified that the HSP70 genes in Chinese cabbage (Brassica rapa ssp. pekinensis) are stress-inducible under both cold and heat stress. The upregulated expression pattern of these genes indicated the potential of HSP70 to mitigate environmental stress. These findings further explain the molecular mechanism underlying the responses of HSP70 to heat and cold stress.Heat shock proteins protect plants from abiotic stress, such as salt, drought, heat, and cold stress. HSP70 is one of the major members of the heat shock protein family. To explore the mechanism of HSP70 in  The heat shock transcription factor family strengthens plants under biotic and abiotic stress conditions; as a result, plants proceed with normal growth and development . TemperaArabidopsis, HSP70 deficient plants showed the stunted plant growth and abnormal leaf phenotype [Arabidopsis plant development under abiotic stress conditions [Brassica rapa family member. This vegetable-based family includes cabbage, Chinese cabbage, mustard, and turnip, which are sources of oil, amino acids, and proteins for humans and animals. These vegetables are highly susceptible to temperature stress [The HSP70 family, which encodes HSP70 proteins, has been identified and studied in many plants, such as Arabidopsis (17 genes), rice (26 genes), spinach 12 genes), and soybean (61 genes) . In Arabhenotype , double henotype . Further genes, ae stress  and are e stress .Brassica rapa has gained remarkable economic and nutritional value due to its heading trait, which makes it highly susceptible to stress. Under temperature stress, the leaves become pale and weakened due to chloroplast damage, and the plant becomes unable to properly regulate photosynthesis [Brassica rapa to date. We identified and explored the members of the HSP70 gene family, constructed their evolutionary tree and synteny correlations, and predicted protein structures with available bioinformatics software. Moreover, we analyzed the expression of these proteins by qRT-PCR to determine their responses to heat shock stress in Chinese cabbage (Brassica rapa subsp. Pekinensis). These findings will open up new gates for understanding the structure, function, and expression of HSP70 genes in B. rapa.ynthesis . Under sA. thaliana HSP70 family genes (AT3G12580.1) was obtained from the TAIR Arabidopsis genome database (TAIR-Home Page arabidopsis.org (accessed on 20 April 2022). This sequence was used as a query to retrieve the HSP70 genes in the Brassica database . Secondly, the HMMER 3.1  and BLASTP with the threshold e-value set to 1e\u22125 were performed using the Hidden Markov Model (HMM V 3.0) profiles of the HSP70 gene family (PF00012) were downloaded from the Pfam protein database  and used as the inquiry, The default limitation of HMMER 3.1 was set to 0.01. Finally, total of 28 HSP70 were retrieved in the Brassica rapa genome. B.rapaHSP70 proteins were further characterized by determining the molecular weight, number of amino acids, isoelectric point, and grand average of hydropathicity (GRAVY) through the ProtParam tool . The protein sequences were uploaded to the online database Wolf PSORT for the prediction of subcellular localization . Furthermore, the protein conserved domain was identified using the NCBI conserved domain online server  and motif analysis was performed by using MEME Suite . The gene structure was predicted by the Gene Structure Display Server (GSDS) , using the CDS and genomic sequences of 28 selected B.rapaHSP70. Furthermore, prediction of the subcellular location pattern of each B.rapaHSP70 was carried out using the WoLF PSORT server .Arabidopsis, B. oleracea, and B. napus were retrieved to construct the phylogenetic tree using MEGA X (V 6.06) software . The sequences were multiple aligned and employed using the neighbor-joining (NJ) method with 1000 bootstrap replicates [B. rapa with B. oleracea, B. napus, and Arabidopsis were developed using MCScanX to obtain collinearity files that were used to build dual synteny plots in TBtools .The protein sequences of HSP70s from plicates . SyntenyB. rapa [B. rapa and its closely related species through TBtools [Duplicated genes were retrieved using the Blast, MCScanX, and Advance Circos features of TBtools . For theB. rapa ,20. Synt TBtools .B.rapaHSP70 genes were classified, and the 2000 bp upstream of the start codon were downloaded from the BRAD database . The PlantCARE web-based tool  was used for further classification, and the results were presented using TBtools. The graphical construction of the 3-dimensional (3D) structure of B.rapaHSP70 proteins was done using the online web-based software PHYRE2 (PHYRE2 Protein Fold Recognition Server (ic.ac.uk), selecting the fold recognition method [Putative cis-elements of the 28 B.rapaHSP70 were used to determine the interaction of genes with microRNAs using the psRNATarget database , and the interactions were further prophesied by Cytoscape Software. Gene ontology (GO) annotation was performed to explore functional characteristics. The B.rapaHSP70 gene sequences were uploaded to the online eggNOG database . Then, the GO annotation data were handled by using  and graphical demonstration was given by using OmicShare .CDS sequences of B.rapaHSP70 genes in different tissues, and a heat map was constructed using TbTools [http://www.ncbi.nlm.nih.gov/geo/, accessed on 20 April 2022) under accession no. GSE43245. In addition, expression profiles of B.rapaHSP70 under cold and heat stress conditions were obtained by performing qRT-PCR.RNA-seq data were used to determine the expression of  TbTools . The datSeeds of Chinese cabbage were grown in pots containing soil: vermiculite mixture (3:1) in a controlled chamber  until they reached the five-leaf stage. Cold and heat stress conditions were applied to five-leaf seedlings. Seedlings were transferred to 4 \u00b0C for cold stress and 45 \u00b0C for heat stress under the same day/night and humidity conditions. Three biological replications were used in both stress conditions. Leaf samples were harvested after 0, 1, 3, 6, 12, and 24 h, placed in liquid nitrogen, and stored at \u221280 \u00b0C [B.rapaHSP70 gene expression; actin was used as the internal reference gene in Chinese cabbage. All primers used in this experiment are listed in \u2212\u0394\u0394Ct method was used to check the relative expression levels, and the results were graphically represented [Leaf samples were used to extract the RNA, and qRT-PCR was performed with three replicates to check resented .B.rapaHSP70-1\u2013B.rapaHSP70-28. These proteins were further analyzed for the HSP70 domain (PF00012) , equal number and similar motif are present within each group except in group 4 , and the longest was B.rapaHSP70-28 (894 aa). The genes that translate into HSP70 proteins were distributed on 10 chromosomes, A01\u2013A10. Among these, the maximum genes were distributed on chromosome A03 and the minimum on chromosomes A02, A04 and A05 , and all selected genes were named PF00012) . Conserv group 4 A. Detail group 4 . Gene st group 4 B. The am and A05 .B.rapaHSP70, B.nHSP70, B.olHSP70, and AtHSP70 were analyzed. Based on domains and a phylogenetic tree, 77 HSP70s were clustered into six major groups   and Arabidopsis.The evolutionary relationships between , and E) . The resB.rapaHSP70 genes in the B. rapa genome. We detected duplicated gene couples among the 28 B.rapaHSP70 genes identified in B. rapa , B. napus (AC), and B. oleracea (CC)\u00a0using collinearity analysis. In addition, a comparative synteny analysis of HSP70 gene pairs among B. rapa, B. napus, B. oleracea, and A. thaliana was conducted (B. rapa (such as Chinese cabbage) and B. oleracea (Cabbage) and B. napus (Mustard) was included due the presence of both sub genome (A and C). B.rapaHSP70 displayed the most collinearity with B. napus, followed by A. thaliana and B. oleracea. Results demonstrated that homologues genes from A genome of B. rapa are present in A as well as C genome of B.napus. Similar behavior was also observed in C genome of B. oleracea. These results suggest that, in addition to the whole genome duplication event, an independent duplication event also occurred during the evolution of these species.BraA10g033140.3C/BraA10g033130.3C, BraA09g038510.3C/BraA01g027130.3C, BraA01g038810.3C/BraA03g035360.3C, BraA08g029700.3C/BraA06g012060.3C, BraA03g019440.3C/BraA03g016490.3C, BraA03g051830.3C/BraA08g020460.3C, BraA02g003050.3C/BraA03g003930.3C, BraA08g021750.3C/BraA01g001130.3C, BraA03g047170.3C/BraA01g019900.3C, and BraA06g001280.3C/BraA04g004290.3C were determined A, which onducted B. We selB.rapaHSP70 genes was retrieved and uploaded to the PlantCare database to examine the cis-elements in the promoter region. The graphical representation  helices and beta (\u03b2) sheets, because proteins with similar structures often have similar functions  topology, thus preserving conserved signature fold of HSP70 family with central mixed \u03b2-sheet sandwiched between \u03b1-helices [B.rapaHSP70 protein family.With the advent of graphical visualization of genomic data, 3D protein structures also explain the properties, as well as facilitate the comparative studies, of proteins. All 28 unctions . Protein-helices . On the B.rapaHSP70 genes, we identified the miRNAs associated with the identified genes , cellular component (CC), and biological process (BP), were investigated to further explore the ed terms . Some ofed terms .The GO-MF (Molecular Function) enrichment results detected 19 enriched terms, namely, protein serine/threonine phosphatase activity (GO:0004722 and GO:0004674), manganese ion binding (GO:0030145), pectate lyase activity (GO:0030570), 1-phosphatidylinositol binding (GO:0005545), glycoprotein binding (GO:0005515), DNA-binding transcription factor activity (GO:0003700), calmodulin binding (GO:0005516), cobalt ion binding (GO:0050897), clathrin binding (GO:0030276), ATP binding (GO:0005524 and GO:0005524), oligopeptide transmembrane transporter activity (GO:0015198), oxidoreductase activity (GO:0016491 and GO:0016491), glutathione peroxidase activity (GO:0004602), 2,3-bisphosphoglycerate-independent phosphoglycerate mutase activity (GO:0046537), protein serine/threonine kinase activity (GO:0004674), beta-amylase activity (GO:0016161), and maltose biosynthetic process (GO:0000024).The GO-BP  enrichment results detected 26 enriched terms, namely, floral organ development (GO:0048437), oxidative stress responsive (GO:0006979), signal transduction (GO:0007165), signal transduction (GO:0006857), defense response (GO:0006952), pollen development (GO:0009555), male gamete generation (GO:0048235), signal transduction by cis-phosphorylation (GO:0007165), clathrin coat assembly (GO:0048268), regulation of transcription (GO:0006355), metabolic process (GO:0008152), salt stress responsive (GO:0009651), transmembrane transport (GO:0055085), plasma membrane organization (GO:0007009), glycolytic process (GO:0006096), abscisic acid responsive (GO:0009737), transmembrane receptor protein tyrosine kinase signaling pathway (GO:0007169), protein phosphorylation (GO:0006468), starch catabolic process (GO:0005983), cold stress responsive (GO:0009409), cadmium sensitivity/resistance (GO:0046686), regulation of meristem growth (GO:0010075), cadmium ion responsive (GO:0046686), temperature stimulus responsive (GO:0009409), maltose biosynthetic process (GO:0000024), and protein phosphorylation (GO:0006468).The GO-CC (cellular component) enrichment results detected 29 enriched terms, including mitochondrial protein-transporting ATPase activity (GO:0005739 and GO:0005739), plasma membrane (GO:0005886), chloroplast envelope (GO:0009941), endomembrane system (GO:0012505), nucleus (GO:0005634 and GO:0005634), chloroplast (GO:0009507), apoplast (GO:0048046), endomembrane system (GO:0012505), chloroplast stroma (GO:0009570), cytosol (GO:0005829), clathrin coat (GO:0030118), mitochondrial envelope (GO:0005740), vacuole (GO:0005773), and integral component of membrane (GO:0016021) .B.rapaHSP70 genes, we examined 8 tissues and organs of B. rapa at various growth phases based on RNA-seq data of B. rapa  under accession no. GSE43245. For instance, the expression patterns of most B.rapaHSP70 genes in the silique, callus, and flower were higher than those of other tissues  showed no transcript changes in any tissue/organ.To illustrate the transcript levels of the  tissues . Three BB.rapaHSP70 genes varied in various tissues and organs. In silique, the expression level of B.rapaHSP70-4, B.rapaHSP70-9, and B.rapaHSP70-24 showing higher expression, and that of B.rapaHSP70-19 was lowest. In callus, B.rapaHSP70-25 was highly expressed, and B.rapaHSP70-10, B.rapaHSP70-11, B.rapaHSP70-12, B.rapaHSP70-16, B.rapaHSP70-18, B.rapaHSP70-20, B.rapaHSP70-4, and B.rapaHSP70-4 was the least expressed. In callus, B.rapaHSP70-4, B.rapaHSP70-9, B.rapaHSP70-24 and B.rapaHSP70-25 are expressing highly and B.rapaHSP70-10, B.rapaHSP70-11, B.rapaHSP70-12, B.rapaHSP70-18, B.rapaHSP70-22 and B.rapaHSP70-28 are showing no expression change. In stem, B.rapaHSP70-4, B.rapaHSP70-9, B.rapaHSP70-24 and B.rapaHSP70-25 are highly expressing, whereas B.rapaHSP70-10, B.rapaHSP70-11, B.rapaHSP70-12, B.rapaHSP70-16, B.rapaHSP70-20, and B.rapaHSP70-26 showing no expression change. In roots, B.rapaHSP70-4, B.rapaHSP70-9, B.rapaHSP70-24, and B.rapaHSP70-25 are highly expressed, whereas B.rapaHSP70-10, B.rapaHSP70-11, B.rapaHSP70-12, B.rapaHSP70-16, B.rapaHSP70-20, and B.rapaHSP70-26 showing no expression change. In flower, B.rapaHSP70-4, B.rapaHSP70-9, and B.rapaHSP70-24 are highly expressed genes, whereas B.rapaHSP70-10, B.rapaHSP70-11, B.rapaHSP70-12, B.rapaHSP70-16, and B.rapaHSP70-22 showing zero expression change. Lastly, the expression was analyzed in leaf, according to the expression pattern the B.rapaHSP70-4, B.rapaHSP70-9, B.rapaHSP70-17, B.rapaHSP70-24, and B.rapaHSP70-25 showing higher expression whereas B.rapaHSP70-10, B.rapaHSP70-11, B.rapaHSP70-12, B.rapaHSP70-16, B.rapaHSP70-20, and B.rapaHSP70-26 showing no expression change. These findings suggest that these candidate genes may play diverse roles in regulating B. rapa growth processes  were upregulated under high-temperature stress at different time intervals, and most were highly expressed after 24 h of heat stress. The expression levels of 16 B.rapaHSP70s  were high at different time intervals under cold stress  as B. rapa. We identified 28 gene family members of HSP70 in B. rapa, which is higher than that found in Arabidopsis (11 AtHSP70) [The model plant B.rapaHSP70 were confirmed by the correlation between intron numbers and motif arrangements in combination with phylogeny. A phylogenetic tree was created to show the evolutionary relationships between B. rapa, B. napus, B. oleracea, and Arabidopsis. Recently, research has examined the evolutionary associations among these species [Notably, genes within the same phylogenetic subgroup had similar motif compositions and exon/intron structures . Genes w species ,34.B.rapaHSP70 family genes and we find all the duplicated B.rapaHSP70s are segmental duplicated.During the genome evolution process, gene duplications and chromosomal segments are major forces in the evolution of plant genome structure and content . Tandem A. thaliana were identified and subjected to promoter analysis to investigate the presence of temperature-associated (heat shock element (HSE)) and dehydration-associated cis-elements  [Previously, a total of 11 HSP70 genes from nt; DRE) . Based oB. rapa stress tolerance. Furthermore, GO enrichment analysis supports the findings of the HSP70 gene association with stress tolerance  [B.rapaHSP70 genes  in post-transcriptional gene regulation . In the nditions ,38. For nditions . Similarinensis) . These s70 genes .StHSP20 under heat stress. In another study, the results verified the involvement of B.rapaHSP70 proteins in thermotolerance [B. rapa and other plant species. In this study, we laid a foundation for recognizing signaling controlled by HSP70 proteins under biotic and abiotic stress conditions.The expression patterns of genes are associated with their function . Severalolerance . Until nB. rapa, and 28 putative members were identified. In silico analyses were performed, including gene structure, distribution, phylogenetic relationship, and syntenic studies, which helped to explore the evolutionary properties of the HSP70 gene family in B. rapa. In addition, targeted miRNAs, promotor cis-acting regulatory elements, and gene ontology (GO) were executed. The finding of all these analysis elucidating that B.rapaHSP70s are targeted by 34 different families of microRNAs, these genes are highly responsive to light, temperature and phytohormones, and B.rapaHSP70s are involved in the Biological, cellular and molecular functioning of B. rapa, respectively. Three dimensional protein structural knowledge can help improve crop properties, such as improving stress resistance and biomass yield. Furthermore, quantitative real-time PCR results indicated that HSP70 genes are strongly involved in heat and cold stress responses in B. rapa plants. To a large extent, all analyses performed on the HSP70 gene family will lay the foundation for further studies of molecular and physiological functions in Brassica crops.In the current study, we performed a genome-wide analysis of the HSP70 gene family in"}
+{"text": "Saussurea costus is a plant traditionally used for the treatment of several ailments. Our study accomplished the UPLC/T-TOF\u2013MS/MS analysis of a methanol extract of Saussurea costus roots (MESC), in addition to lipoidal matter determination and assessment of its in vivo hepatoprotective activity. In this study, we were able to identify the major metabolites in MESC rather than the previously known isolated compounds, improving our knowledge of its chemical constituents. The flavones apigenin, acacetin, baicalein, luteolin, and diosmetin, and the flavonol aglycones quercetin, kaempferol, isorhamnetin, gossypetin, and myricetin and/or their glycosides and glucuronic derivatives were the major identified compounds. The hepatoprotective activity of MESC was evaluated by measuring catalase activity using UV spectrophotometry, inflammatory cytokines and apoptotic markers using ELISA techniques, and genetic markers using PCR. Paracetamol toxicity caused a significant increase in plasma caspase 2, cytokeratin 18 (CK18), liver tumor necrosis factor-\u03b1 (TNF-\u03b1), interleukin 6 (IL-6), miRNA-34a, and miRNA-223, as well as a significant decrease in liver catalase (CAT) activity and in the levels of liver nuclear factor 1\u03b1 (HNF-1\u03b1), sirtuin-1, and C/ebp\u03b1. Oral pretreatment with MESC (200 mg/kg) showed a significant decrease in caspase 2, CK18, TNF-\u03b1, IL-6 and a significant increase in liver CAT activity. MESC decreased the levels of liver miRNA-34a and miRNA-223 and induced HNF-1\u03b1, sirtuin-1, and C/ebp\u03b1 gene expression. The histological examination showed a significant normalization in rats pretreated with MESC. Our findings showed that Saussurea costus may exert a potent hepatoprotective activity through the modulation of the expression of cellular cytokines, miRNA-34a, and miRNA-223. Saussurea costus  is considered one of the most important traditional Chinese medicinal plants. It is a rich source of various bioactive phytoconstituents, and its genus comprises about 300 species +. Luteolin-O-hexoside (21) exhibited a deprotonated molecule [M-H]\u2212 at m/z 447.0966; we then observed a subsequent loss of 162 amu for hexose at m/z 285.0520 [M-H-hexose]\u2212, while luteolin-C-hexoside (31) showed a protonated molecule [M+H]+ at m/z 449.1549. Luteolin-di-O-hexoside (13) exhibited a deprotonated molecule [M-H]\u2212 at m/z 609.1462; moreover, luteolin aglycone (40) was detected with a deprotonated molecule [M-H]\u2212 at m/z 285.039, and characteristic peaks at m/z 269.1614 [M-H-OH]\u2212 and 151.0980 indicated a dihydroxy-substituted A-ring [The glycoside diosmin (24) exhibited a protonated molecule [M+H]hexoside  exhibitem/z 577.1506 and a fragment at m/z 269.0386 [M-H-neohesperidose]\u2212. Apigenin-C-hexoside (22) showed a protonated molecule [M+H]+ at m/z 433.1232; however, apigenin-O-hexoside (32) showed a protonated molecule [M+H]+ at m/z 433.1851 and a fragment ion peak at m/z 271.1296 [M+H-hexose]+. Moreover, apigenin aglycone (34) was detected at m/z 269.0434 [M-H]\u2212 [O-rutinoside (10) showed a protonated molecule [M+H]+ at m/z 593.1631; on the other hand, acacetin aglycone (39) exhibited a deprotonated molecule [M-H]\u2212 at m/z 283.1895 and a production at m/z 253.1469 corresponding to [M-H-OCH3]\u2212. Baicalein-O-glycuronide (14) showed a protonated molecule [M+H]+ at m/z 477.0679.Rhoifolin glycoside (Apigenin neohesperidoside) (20) showed a deprotonated molecule [M-H] at hexoside  showed a+ at m/z 319.1651 [m/z 481.1810 [M+H]+ and also showed a fragment at m/z 319.0663 [M+H-hexose]+. Quercetin hexoside (19) showed a protonated molecule [M+H]+ at m/z 465.1058; The neutral loss of 162 amu of the hexose moiety was observed in MS2 fragmentations at m/z 303.0471 [M+H-hexose]+. Quercetin glucuronide (11) was detected at m/z 477.0659 [M-H]\u2212, and the neutral loss of 176 Daltons of the glycuronic moiety was confirmed by an ion at m/z 301.0332 [M-H-glycuronic acid]\u2212. Quercetin pentoside (15) showed a deprotonated molecule [M-H]\u2212 at m/z 433.1094; however, quercetin aglycone (28) was confirmed by an ion peak at m/z 303.0966 [M+H]+ \u2212 of kaempferol neohesperidoside (16) was detected at m/z 593.1527. Isorhamnetin-O-hexoside (25) was detected by the presence of a protonated molecule [M+H]+ at m/z 479.1228, a production at m/z 317.0612 corresponding to [M+H-hexose]+, and a characteristic fragment at m/z 285.0323 319.1651 . Gossype6 [M+H]+ . Kaempfe285.0323 . The dep+ at m/z 273.0884 and a characteristic fragment at m/z 147.0214 [M+H-C6H6O3]+ [\u2212 at m/z 301.2041. Formononetin-O-hexoside (29) showed a protonated molecule [M+H]+ at m/z 431.1681 and a fragment at m/z 269.1243 [M+H-hexose]+. The deprotonated molecule [M-H]\u2212 of formononetin aglycone (37) was detected at m/z 267.0712 and yielded a fragment due to the methyl group loss at m/z 252.0420 [C-hexoside (18) showed a deprotonated molecule [M-H]\u2212 at m/z 415.1944, while daidzein aglycone (27) was detected at m/z 255.0551.Naringenin aglycone (36) showed a protonated molecule [M+H]C6H6O3]+ . Hespere+ at m/z 579.1859 and a fragment ion at m/z 561.1873 [M+H-H2O]+. Petunidin-O-hexoside (26) showed a protonated molecule [M+H]+ at m/z 479.0961 and a fragment ion at m/z 317.0662 [M+H-hexose]+. Malvidin-O-hexoside (33) was detected at m/z 491.1155 [M-H]\u2212 and confirmed by a fragment at m/z 329.0696 for [M-H-hexose]\u2212.Procyanidin B2 (12) showed a protonated molecule [M+H]m/z 191.0349 [M-H]\u2212, and daphnetin (46), which was confirmed by a peak at m/z 179.0861 [M+H]+ with a fragment at m/z 163.0503. Aldehydes were tentatively identified as hexenal (47), showing a protonated molecule [M+H]+ at m/z 99.0437 and a fragment ion at m/z 83.0493, while cinnamaldehyde (48) showed a protonated molecule [M+H]+ at m/z 133.1001 and a fragment ion at m/z 117.0684 + at m/z 415.3205, in accordance with published data [Tentatively identified coumarins were scopoletin (44), that was detected in both modes at 117.0684 . The chehed data ,29.2 as in succinic and malic acids  [The common neutral loss of 44 Daltons was observed in MS2 fragmentations of phenolic and organic acids due to the loss of COctively) . The ideIn the present study, MESC was evaluated for its hepatoprotective activity in paracetamol-induced liver toxicity. Plasma caspase 2 and cytokeratin 18 (CK18) were elevated in the group Gp II in comparison with the control group. Hofer et al. reported that caspase-cleaved cytokeratin 18 (CK18-Asp396) values were increased significantly in Gp II . Caspasep < 0.01) in plasma caspase 2 and CK18 levels, corresponding to 346.39% and 139.31% increases, respectively, as compared to the levels in Gp I, considering the values in the control group as 100% (Toxicity by paracetamol (1 g/kg) led to a significant elevation ( as 100% . Pretreap < 0.01) in liver CAT level by 50.37%, compared to the levels in Gp I , resulting in inflammation ,41. MESCp < 0.05) were observed (p < 0.05) in C/ebp\u03b1, HNF-1\u03b1, and sirtuin-1 gene expression and a significant decrease (p < 0.05) of liver miRNA-34a and miRNA-223 genes expression compared to the levels in Gp II. MESC administration showed a significant increase (p < 0.05) in the expression of liver C/ebp\u03b1, HNF-1\u03b1, and sirtuin-1 genes by 404.35%, 179.55%, and 223.64%, respectively, as well as a significant decrease (p < 0.05) of liver miRNA-34a and miRNA-223 genes expression by 46.27% and 40.49%, respectively, compared with the levels in Gp II. Paracetamol overdose could suppress C/ebp\u03b1 gene expression in hepatocytes in vivo, which was accompanied by a significant accumulation of cytokines. MESC and silymarin protected rats against paracetamol-induced hepatotoxicity. Oxidative stress due to C/ebp\u03b1 gene expression depletion and uncontrolled ROS is known to be the primary pathogenic mechanism of paracetamol-induced liver toxicity, as well as the main inhibitor of hepatocytes protective factors. Our metabolomics results revealed that MESC is rich in flavonoid compounds that exerted pronounced antioxidative effects against paracetamol-induced depletion of C/ebp\u03b1 gene expression in the hepatocytes of Gp II rats [A significant decrease of the expression of HNF-1\u03b1, sirtuin-1, and C/ebp\u03b1 genes by 55.10%, 61.22%, and 77%, respectively, as well as a significant increase of the expression of liver miRNA-34a and miRNA-223 genes by 447.27% and 344.79%, respectively, in Gp II rats as compared with Gp I rats  demonstrated normal hepatocytes with no fibrosis or inflammation a, while S. costus; which supports the hepatoprotective activity of this plant [S. lappa root extract in rats [S. costus root extract pretreatment.Our LC\u2013MS/MS results showed that flavonoids were the major components of is plant . Coumariis plant . Tejaswi in rats ; moreove in rats ,57. ThisMethanol (HPLC grade), paracetamol (99%), silymarin (98.5%), and Tween 80 were purchased from Merk . Other reagents were of high analytical grade.Saussurea costus  Lipsch. roots were purchased from a local herbalist . Identification of the plant was performed by the Agriculture Research Center, Cairo, Egypt. In Soxhlet, powdered roots (1 kg) were extracted with methanol . The extract was left to cool, filtered, and then evaporated  [zerland) . The obtn-hexane (2 \u00d7 100 mL), filtered, and evaporated. The crude oil was investigated for its composition in a GC\u2013MS system  equipped with a mass spectrometer detector (5977A) at the National Research Centre, Cairo, Egypt. The GC was equipped with an HP-5MS column. Powdered roots (100 g) were macerated in m/z. The characterization of compounds was performed by the generation of the candidate formula with a mass accuracy limit of 10 ppm and also considering Rt, MS2 data, databases, and reference literature [The MESC was analyzed at the Proteomics and Metabolomics unit of the Children\u2019s Cancer Hospital Egypt 57357, Cairo, Egypt. The analysis was carried out using an Exion LC Triple TOF 5600+ system  operated at 40 \u00b0C and equipped with an X select HSS T3 C-18 column  and a precolumn . MESC (50 mg) was dissolved in solvent working solution (MilliQ water: methanol: acetonitrile\u201350:25:25), sonicated (10 min), then centrifugated . The stock sample (50 \u00b5L) was diluted with the working solvent (1000 \u00b5L). The phytoconstituents of MESC were analyzed using UPLC/T-TOF\u2013MS/MS in both negative and positive modes . Samplesterature . g, 10 min). The clear supernatant was tested for CAT using a Cayman Chemical Company kit . The UV spectrophotometric method for measuring catalase activity is centered on monitoring the change in 240 nm absorbance at high concentrations of hydrogen peroxide (30 mM). TNF and IL-6 levels in the liver were measured using a microplate reader at 450 nm . An anti-IL-6 or an anti-TNF-\u03b1 monoclonal antibody and a biotin-conjugated monoclonal anti-IL-6 or anti-TNF-\u03b1 antibody that binds to IL-6 or TNF-\u03b1 captured by the first antibody were used in an indirect sandwich enzyme-linked immunosorbent assay to evaluate IL-6 and TNF-\u03b1. Streptavidin\u2013HRP was added to the wells, followed by a substrate solution reacting with HRP. After stopping the reaction with acid, the absorbance was measured.Adult male albino rats  were purchased from the National Cancer Institute, Cairo University, Giza, Egypt. Rats were provided water and standard diet ad libitum, observed daily, and kept in polypropylene cages under normal environmental conditions (22 \u00b1 2 \u00b0C). The prophylactic potential of MESC against paracetamol hepatotoxicity was evaluated. Ethical approval was obtained by the ethic committee of the faculty of applied medical science . Animals and the treatment schedule (4 weeks) were as follows: Gp I  and Gp II (positive control); rats were orally given Tween 80  in saline daily; Gp III rats were orally treated with silymarin , daily . Gp IV r2, (1.5 mM), 0.2 mM of each dNTP, 0.4 \u03bcM of specific primers . RNA (1 \u03bcg) in reaction buffer was mixed with dithiothreitol (10 nmol/L), oligo (dT) primer (25 pg), 0.5 mmol/L of each deoxyribonucleoside triphosphate (dNTP), and 200 units superscript II Rnase H-Reverse Transcriptase. The reactions were kept at 42 \u00b0C for 2 min, 42 \u00b0C for 50 min, 70 \u00b0C for 15 min, and then chilled to 4 \u00b0C. The PCR reaction mixture consisted of PCR buffer, MgCl primers  for hepaThe liver pieces were fixed in formaldehyde solution (10%) and then were examined for histopathological changes.p < 0.05 was considered statistically significant). Statistical analyses were carried out using GraphPad Prism . ANOVA with posttest Tukey\u2019s multiple comparisons were performed. Values (n = 10) are presented as mean \u00b1 standard deviation for ELISA measurements and PCR analyses of gene expression  were characterized as belonging to different chemical classes, flavonoids being the major constituents besides organic acids, coumarins, and anthocyanidin. S. costus has a potent hepatoprotective activity through the modulation of cellular cytokines release and miRNA-34a and miRNA-223 expression. S. costus reduced the levels of caspase 2 and CK18, TNF-\u03b1, and IL-6 and increased catalase activity. Furthermore, it increased HNF-1\u03b1, sirtuin-1, and C/ebp\u03b1 expression and decreased miRNA-34a and miRNA-223 gene expression. S. costus minimized some negative symptoms and pathological changes. We hope that this study will attract attention towards this plant myriad chemical constituents and its great potential in health care.Herein for the first time, we were able to identify the chemical profile of"}
+{"text": "Worse sleep quality and increased inflammatory markers in women with schizophrenia (Sch) have been reported . However, the physiological mechanisms underlying the interplay between sleep and the inflammatory pathways are not yet well understood .Analyze the relationship between Neutrophil/Lymphocyte (NLR), Monocyte/Lymphocyte (MLR) and Platelet/Lymphocyte (PLR) ratios, and insomnia in Sch stratified by sex.Final sample included 176 Sch patients (ICD-10 criteria) . Assessment: PANSS, Calgary Depression Scale (CDSS), and Oviedo Sleep Questionnaire (OSQ) to identify a comorbid diagnosis of insomnia based on ICD-10. Fasting counting blood cell were performed to calculate ratios. Statistics: U Mann-Whitney, logistic regression.Insomnia as comorbid diagnosis was present in 22 Sch (12.5%) with no differences between sex , neither in their age. Female patients with insomnia showed increased NLR . However, no differences in PLR and MLR were found, neither in any ratio in males. Regression models using insomnia as dependent variable and covariates  were estimated. Females: presence of insomnia was associated with NLR [OR=3.564 (p=0.032)], PANSS-positive [OR=1.263 (p=0.013)] and CDSS [OR=1.198 (p=0.092)]. Males: only PANSS-positive [OR=1.123 (p=0.027)] and CDSS scores [OR=1.220 (p=0.005)] were associated with insomnia.NLR represent an inflammatory marker of insomnia in Sch but only in female patients. Improving sleep quality in these patients could help to decrease their inflammatory response.No significant relationships."}
+{"text": "Non-polio Enteroviruses (EV) are important neonatal CNS pathogens. Multiple EV genotypes have been detected in pediatric CSF, e.g. Echovirus (E) 6 and E30. CSF innate immune responses to EV genotypes remain poorly defined. Most data are from EV-A71 or E30 CNS infections and do not compare responses between these or other EV types. We sought to better define innate immune responses to EV genotypes in CSF.Salvaged standard of care CSF samples from \u22646 month olds  or EV PCR(-) controls) from Jan 2010 - Dec 2020 were tested in duplicate on a 21- cytokine bead panel (MilliporeSigma). EV positive samples were previously genotyped by sequencing the viral capsid gene. Cytokine levels calculated from the standard curve were compared by Kruskal-Wallis and post-hoc analysis (GraphPad Prism 8.4.3). Natural partitioning of participants was explored using principal component analysis and cluster analysis (IBM SPSS v27). The utility of cytokine signatures in predicting EV status was explored using discriminant analysis and ROC analysis (IBM SPSS v27).Data from 72 CSF with E6 (N=16), E9 (N= 9), E18 (N=9), and E30 (N=21) showed significant differences among EV genotypes vs. controls for 20 cytokines (IL-17 was excluded). Significant differences in cytokine levels in EV CSF vs controls were seen: E6 for all 20 cytokines; E9 for Fractalkine, IP10, and MCP1; E18 for Fractalkine and MCP1; E30 19 cytokines (not GM-CSF). PCA revealed only minor overlap of controls and EV positives; EV types overlapped, except E30, differing most from E9 and E18 but overlapping E6. The most important type-differentiating cytokines by PCA were MCP1, Fractalkine, IL-8, and IL-10. Patterns in DA resembled PCA; controls clearly separated from EV CSF, E30 being the most distinct. Overall, the discriminant model correctly classified EV type or controls at a 63% rate - highest for controls (94.1%) and E30 (74.1%). In the DA model, the most important cytokines were IP-10, IL-2, IL-1Ra, and Fractalkine. Discriminant scores had the largest area under the curve (AUC), 0.990. Among cytokines, IFNa2 had the largest AUC, 0.987.Preliminary data show significant EV-genotype differences in innate immune response to CNS infections. Cytokine patterns may serve as a key predictor for discerning EV genotypes.Brian R. Lee, PhD, MPH, CDC: Grant/Research Support|Merck: Grant/Research Support Christopher J Harrison, MD, Astellas: Grant/Research Support|GSK: Grant/Research Support|Merck: Grant/Research Support|Pediatric news: Honoraria|Pfizer: Grant/Research Support Rangaraj Selvarangan, BVSc, PhD, D(ABMM), FIDSA, F(AAM), BioFire: Grant/Research Support|Luminex: Grant/Research Support."}
+{"text": "A vascular system in plants is a product of aromorphosis that enabled them to colonize land because it delivers water, mineral and organic compounds to plant organs and provides effective communications between organs and mechanical support. Vascular system development is a common object of fundamental research in plant development biology. In the model plant Arabidopsis thaliana, early stages of vascular tissue formation in the root are a bright example of the self-organization of a bisymmetric (having two planes of symmetry) pattern of hormone distribution, which determines vascular cell fates. In the root, vascular tissue development comprises four stages: (1) specification of progenitor cells for the provascular meristem in early embryonic stages, (2) the growth and patterning of the embryo provascular meristem, (3) postembryonic maintenance of the cell identity in the vascular tissue initials within the root apical meristem, and (4) differentiation of their descendants. Although the anatomical details of A. thaliana root vasculature development have long been known and described in detail, our knowledge of the underlying molecular and genetic mechanisms remains limited. In recent years, several important advances have been made, shedding light on the regulation of the earliest events in provascular cells specification. In this review, we summarize the latest data on the molecular and genetic mechanisms of vascular tissue patterning in A. thaliana root. The first part of the review describes the root vasculature ontogeny, and the second reconstructs the sequenceof regulatory events that underlie this histogenesis and determine the development of the progenitors of the vascularinitials in the embryo and organization of vascular initials in the seedling root. Evolutionary formation of a vascular system in plants wasa necessary prerequisite for terrestrial colonization . Vasculature provides mechanical support, effectivetransportation of water, and mineral and organic compoundsas well as signal molecules and by this has enabled plants toreach enormous sizes and populate different territories. Thevascular system consists of two domains different in theirstructure and functions. These are xylem that provides watertransportation and delivers mineral compounds from the rootto above-ground organs; and phloem that conveys organiccompounds from photosynthesizing tissues rootward .In angiosperms, the mature xylem consists of (1) watertransportationvessels; (2) fibers to provide mechanical support;(3) parenchyma cells . The vesselsare the hollow tubes formed by the cells connected in a rawand having perforations in the anticlinal walls and pores in thepericlinal walls . The vessels and fibers are a productof the programmed death of the cells that have formed a lignifiedsecondary cell wall . Meanwhile, the living cellsof parenchyma perform a storage function, participating invessel lignification and regulating the water transport speed.The phloem, on the other hand, consists of (1) sieve tubesto transport organic substances; (2) companion cells; (3) fibersand sclereids to provide mechanical support, and (4) parenchymacells . Unlikethe lignified hollow vessels of the xylem, the sieve tubes area strand of living cells (sieve elements) communicating bysieve fields, anticlinal-wall regions with high numbers ofsmall pores. The sieve elements form a thickened non-lignifiedsecondary cell wall  and their main feature isthe lack most of the organelles including a nucleus, vacuole,rough endoplasmic reticulum, Golgi body, cytoskeleton,ribosomes whose presence could prevent substances transportation.The viability of the sieve elements is maintainedby companion cells \u2013 the parenchyma cells with large nucleiand mitochondria, directly contacting sieve elements. As for mechanical phloem elements \u2013 fibers and sclereids \u2013 they differfrom each other by the shape of their cells. While the formerare strongly elongated and pointed at the ends, the latter arejust slightly elongated.Organization of vascular system is different for differentorgans in different plant species at different stages of theirdevelopment . Nevertheless, the mechanisms determiningits developmentare quite conservative . Plant cells are not capable of migration, so duringmorphogenesis, the tissue and organ architecture is formedby regulating the sequence and orientation of cell divisions.In terms of its anatomy, the vascular system development hasbeen described in much detail , however, the molecular andgenetic mechanisms responsible for this process are muchless known. Our current understanding of these mechanismsis mainly based on the investigation of the model plant ArabidopsisthalianaIn the further sections of this review, we will provide a shortdescription of vascular tissue histogenesis in this plant speciesand reconstruct the corresponding sequence of regulatoryevents. We will describe the control of root vascular systemdevelopment in the embryo and seedling, i. e. the earlieststages of its formation. As for the mechanisms controllingvasculature development at later stages, their description canbe found in the recent reviews .GlossaryAmphicribral vascular bundle \u2013 a vascular bundle inwhich the phloem surrounds the xylem.Anticlinal \u2013 located in a plane perpendicular to the surfaceof a tissue or organ. Talking about anticlinal cell wallsor divisions we will mean an anticlinal plane perpendicularto the central axis of an organ.Anticlinal cell division \u2013 cell division in the anticlinalplane that leads to an increase in length.Asymmetric cell division \u2013 results in the formation oftwo daughter cells with different cell fates.Cortex \u2013 a cell layer surrounding the endodermis.Diarch vascular bundle \u2013 a vascular bundle whose phloemand xylem are located at different radii, wherein tworays of xylem are distinguished.Endodermis \u2013 the innermost cell layer surrounding thestele.Hypophysis is the upper cell of the suspensor, which acquiresits identity at the 16\u201332 cell stage; gives rise to thequiescent center  and the root cap.Periclinal \u2013 located in a plane parallel to the surface ofa tissue or organ.Periclinal cell division \u2013 cell division in the periclinalplain leading to an increase in the number of cell layersin the radial direction.Pericycle \u2013 parenchyma cell layer surrounding conductivetissues and forming the stele outer layer.Primary meristem \u2013 formed during embryogenesis.Procambium \u2013 indeterminate primary vascular meristemcells located between the xylem plate and phloempoles in the root of Arabidopsis thaliana.Provascular initials \u2013 four proembryo cells occurring atthe early globular stage to form the entire provascularmeristem of the root/hypocotyl, and only it.Provascular root/hypocotyl meristem \u2013 primary meristemfrom which the primary vascular system of theseorgans differentiates after embryo germination.Root apical meristem \u2013 primary root meristem to produceall cells of the root during its post-embryonicgrowth.Secondary meristem \u2013 formed during the postembryonicperiod.Stele  \u2013 primary conductive tissues locatedin the center of the axial organ, and surrounded bya pericycle.Suspensor \u2013 a structure at the base of an embryo thatconnects it to endosperm and consists of the descendantsof a two-celled pro-embryo basal cell.Vascular cambium \u2013 secondary vascular meristem toprovide root thickening.Xylem plate \u2013 a layer of primary xylem cells (or theirpredetermined precursors) located in the central planealong the root axisThere are primary (produced by the primary meristem) andsecondary (produced by the secondary meristem) vasculartissues.Development of root primary conductive tissuesAt the globule stage of A. thaliana embryogenesis the specificationof four provascular initials occurs. Provascular initialsundergo oriented divisions, finally giving rise to the provascularmeristem of the embryonic root and hypocotyl . Thecells of provascular meristem are not yet differentiated, but thecellular fate of some of them has already been determined \u2013after the embryo germination they give birth either to xylemor to phloem cells. The positions of these predetermined cellsin the provascular meristem matches that of the bisymmetric diarch organizationof the vascular system in the postembryonic root tip: in itstransverse \u2013 section, there is one layer of xylem precursor cellssurrounded on both sides by procambial cells that separate thefuture xylem from two files of phloem progenitor cells, whichlie in a perpendicular plane  .This structure is surrounded by pericycle cells that are alsoderived from provascular initials, so together they form a centralcylinder or a stele . It is noteworthy that theterminology designating the cells in developing root vascularsystem is rather blurred . In particular, theterm \u2018procambium\u2019 is applied to address either indeterminate cells of the primary vascular tissue in seedlings (and theirprogenitors) or the whole embryonic provascular meristem.Soon after germination, vascular elements start to differentiatein a hypocotyl stele and cotyledon veins, the provascularmeristem of the latter comes from the shoot apical meristem. From the hypocotyl, the processspreads upwards and downwards taking the epicotyl and root,respectively . In A. thaliana, these are the protophloemsieve elements adjacent to the pericycle that differentiatefirst, and since the cells surrounding them keep elongating,protophloem cells soon die to be functionally replaced bythe metaphloem sieve elements placed closer to the center ofthe stele . Later, theprotoxylem vascular elements are formed that are located atthe poles of the xylem plate and have annular or spiral thickeningsof the secondary cell walls. The last cells to differentiateare the metaxylem cells occupying the central position in thexylem plate and having pitted or reticulate lignin deposits.While the root grows in length, its new cells are producedthrough anticlinal division of the cells in the apical meristemlocated at the root tip . In A. thaliana,the root apical meristem is closed, i. e. different stem cells can produce not any but strictly limited set of celltypes and for each differentiated cell it is easy to trace whichstem cell it has originated from . Among the steleinitials those can be distinguished that give birth to (1) protoxylem;(2) metaxylem; (3) procambium and sieve elementsof proto- and metaphloem ;(4) only procambial cells; (5) pericycle . The mutual arrangement of initials correspondsto the diarch organization of young root vasculature, so thecell identity established in the embryo provascular meristem ismaintained in the root apical meristem. Here it is worth mentioningthat apart from the proto- and metaxylem, proto- andmetaphloem and procambium there are also companion cells.Some authors designate them more srictly as companion-likecells . These cells are adjacent to the sieveelements of proto- and metaphloem and possess a number ofmorphological and physiological characteristics of companioncells  but, unlike the latter, theydo not share a common initial with the proto- and metaphloemelements in the stem cell niche . Thecompanion-like cells differentiate when the protophloemsieve elements start functioning . InA. thaliana, the xylem and phloem parenchyma, fibers andtrue companion cells differentiate only during the secondarygrowth .In A. thaliana primary vascular system, periclinal divisionsof procambium cells are few, but after differentiation of theprimary vascular elements, these cells begin to actively dividepericlinally. The periclinal divisions also occur in the pericycle cells adjacent to the xylem plate. As a result, a closed cell ringforms around the xylem to give birth to the vascular cambium . It is noteworthythat only those procambium and pericycle cells in direct contactwith the xylem primary vessels give rise to the vascularcambium, i. e., have the properties of stem cells  while the descendants of other proliferating procambialcells differentiate into the phloem.Thus, the diarch root vasculature transforms into amphicribralone, in which the xylem is surrounded by thephloem with the cambium placed in between .Through asymmetric division, every initial is capable of producingphloem cells outwards and xylem cells inwards, sothe root gets thicker . In some species,e. g., in the vast majority of monocots, the cambium is notformed and no secondary growth is initiated. In this case, allprocambium cells get differentiated.The development of a multicellular organism is accompaniedby a gradual increase in the limitation of cellular potencies. Atthe first stage of this process predetermination or specificationoccurs, in other words, the fate of a totipotent cell is establishedin terms of the progenitor of what type of cells it will become.Meanwhile, the cell remains undifferentiated and can changeits fate under certain conditions. The process of cell identitydetermination involves the local accumulation of signalmolecules, which either activate or suppress the activity thegene networks inherent in specific cell types. In this case, animportant role is given to the non-cell-autonomous factors ableto move between cells and form gradients .Provascular stem cells specification at the early globularstage of embryogenesis is preceded by a series of cell divisionsand embryo polarity determination . The proper accomplishment of these processes isessential for the vascular tissue to begin its development fromthe right number of cells placed in the right positions. Planthormone auxin is a key regulator of embryogenesis, whoseheterogeneous distribution provides positional information,which directs embryo development . The main auxineffector in embryogenesis is transcription factor (TF) AUXINRESPONSE FACTOR 5 (ARF5)/MONOPTEROS (MP) and it is believedthat forming the auxin signal-distribution pattern is providedmainly due to feedbacks in regulation of phytohormonebiosynthesis, its polar intercellular transport and signalingpathway . As a result, at the early stagesof embryogenesis, auxin is accumulated in the apical cells todetermine the embryo polarity . Startingfrom the early globular stage (32 cells), its maximum is shiftedto the upper cells of the suspensor including the hypophysisthat later gives rise the quiescent center of the root apicalmeristem .Although the four provascular initials are only distinguishedat the early globular stage , the cellular identity of vascular tissue progenitors is determined in the fourinner cells of the lower layer of the proembryo as early as atdermatogen stage  . Via periclinaldivision at transferring to the 32-cell stage, they produceoutwards the ground tissue progenitors that lose the vascularidentity of their maternal cells  . A necessary condition for provascular-initial specification is ARF5/MP-dependent activation ofthe auxin signaling pathway, but meeting this condition aloneis not enough . Whileparticular auxin assistants remain unknown, its is suggestedthat this role is performed not by a single key regulator butby a multicomponent regulatory network, and TF G-BOXBINDING FACTOR 2 (GBF2) is believed to be one of itsmembers  . GBF2 is assumedto modulate ARF5/MP binding to target-gene promoters. It isworth mentioning here that the state, in which vascular systemprogenitors are uniformly specified is most likely transientwith no stable uniform cellular identity.As the oriented divisions of the provascular initials and theirdescendants continue, the hypocotyl and root vascular systemsbecome patterned through specification of particularcellular types. An important aspect at this stage is setting theboundaries for the cellular domains with different structuraland functional identities. By the end of embryogenesis, inthe embryo provascular meristem, the cell identity of all elementssuch as proto- and metaphloem, proto- and metaxylem,companion-like cells and procambium has been determinedas evidenced by the data on cell morphologyand expressionof marker genes  .In A. thaliana, the bisymmetry of the future root is believedto be predetermined already at the early globular stage by theextended contact between two provascular initials locateddiagonally relative to each other . This contactis probably formed due to the inaccurate match of cell divisionplanes in proembryo (at the four-cell stage) and is importantfor xylem plate formation . Startingfrom the early heart stage, auxin begins to be activelytransported into such contacting provascular cells from thecotyledon primordia located above them, while in other cellsthe hormone levels remain low . The local increase inauxin concentration is necessary for the specification of xylemprogenitor cells .At the same time, the cells rich in auxin begin to act as anorganizing center for the provascular meristem, coordinatingits growth through periclinal divisions and establishment ofbisymmetric organization . Auxin inducesthe ARF5/MP-dependent expression of TFs TARGETOF MONOPTEROS 5 (TMO5) and TMO5-LIKE1 (T5L1), which,forming heterodimers with the auxin-independent LONESOME HIGHWAY (LHW) TF , activatethe expression of cytokinin biosynthesis genes LONELYGUY3 (LOG3) and LOG4  . Simultaneously, auxin blockscytokinin signal transduction, increasing the expression ofgene ARABIDOPSIS HISTIDINE PHOSPHOTRANSFERPROTEIN 6 (AHP6 ) encoding a cytokinin signaling pathwayinhibitor , soa local cytokinin source is formed in xylem progenitors lackingcytokinin signalingThe high cytokinin level, on the one hand, limits auxin effluxfrom xylem progenitor cells by controlling the localizationof auxin transporter PIN-FORMED 1 (PIN1) on the cell membrane. On theother hand, cytokinin diffuses into neighboring cells followingthe concentration gradient. In these cells, in the absence of theinhibitor , cytokinin activates signalingcascade to stimulate periclinal divisions .Simultaneously, cytokinin signaling suppresses cell specificationinto xylem . This mechanismprovides for the radial growth of the provascular meristem,which is accompanied by spatial separation of the domainsfor increased auxin signal (cells obtain xylem identity) andcytokinin signal . Its sufficiencyfor self-organization of the bisymmetric pattern was confirmedusing a mathematical model .In early embryogenesis, provascular-meristem progenitorsbegin to express genes encoding peptide hormone CLAVATA 3(CLV3)/EMBRYO SURROUNDING REGION 25 (CLE25)and mobile TFs of the DNA BINDING WITH ONE FINGER(DOF) family united in the PHLOEM EARLY DOF (PEAR)group marking sieve-element progenitors in the postembryonicperiod . CLE25is expressed starting from a 64-cell embryo stage . Cytokinin-independent expression of PEAR1 isdetected already at a 16-cell stage, and starting from an earlyheart stage, this gene expression is activated by cytokinin. It is assumed that the CLE25 peptidebinding to the CLE-RESISTANT RECEPTOR KINASE(CLERK)-CLV2 receptor together with the PEAR1 TF contributeto the early specification of phloem progenitor cells.However, unlike that for xylem, the mechanism to initiatephloem development in embryogenesis remains unknown.Bisymmetric pattern in steleIn the postembryonic period, the stele cells progenitors maintainthe bisymmetric pattern established in embryogenesis,so some of the mechanisms regulating the cell dynamics andvascular-system element predetermination in provascular meristemkeep functioning even after germination. However, itcannot be said with complete certainty that these mechanismsare identical.In the apical meristem, auxin-rich xylem progenitors retainthe function of an organizing center, carrying out TMO5/LHW-mediated regulation of cytokinin levels in procambialcells  . The high contentof active cytokinin in xylem cells is maintained by TMO5/LHW-dependent activation of not only cytokinin biosynthesisgenes LOG3 and LOG4 but also of the BGLU44 gene encodinga \u03b2-glucosidase enzyme . Cytokinin response inxylem is blocked by auxin through AHP6 gene expression induction  as well as through limitingthe activity of TMO5/LHW by activating the ACAULIS 5(ACL5)\u2013SUPPRESSOR OF ACAULIS5 LIKE3 (SACL3)regulatory module blocking the formation of the TMO5/LHWheterodimer by competing with TMO5 for binding to LHW . Meanwhile,in xylem-adjacent procambial cells, the level of thecytokinin diffusing from the xylem is limited by TMO5/ LHWdependentactivation of CYTOKININ OXIDASE 3 (CKX3).The activation is mediated by the mobile SHORT ROOT(SHR) TF, encoded by TMO5/LHW target gene. The combinedaction of multidirectional regulatory modules ensuresthe stability of the pattern to short-term fluctuations in auxinconcentrations in xylem cells, while maintaining its sensitivityto slower/stable changes . What is interestingis that the SHR gene is important not only for the rootradial symmetry but also for the functioning of the quiescentcenter .TMO5/LHW-induced cytokinin activates the transcriptionof the DOF2.1 TF in the procambial cells surrounding thexylem pole, thus controlling their division  . It is worth noting that, besides xylem cells, it isdifferentiated phloem that transports the phytohormone andthus can be a source of cytokinin in the root apical meristem. However, mathematical modelinghas demonstrated that phloem cytokinin is not a fundamentalsource of the positional information for bisymmetric patternformation . At the same time, the highcytokinin content at the phloem poles arranges periclinaldivisions of procambium cells through activating the genesof mobile TFs of the DOF family united in the PEAR groupincluding PEAR1, PEAR2, TMO6, DOF6 . They create a concentration gradientand activate the periclinal divisions of the procambial cells surroundingthe phloem pole. HOMEODOMAIN LEU-ZIPPERclass-III (HD-ZIP III), TFs whose expression domain is set inthe central part of the stele (see below) limit the activity of thePEAR TFs , and PEAR1 activates the transcriptionof the genes belonging to the HD-ZIP III family, forminga negative feedback loop.Proto- and metaxylem predeterminationAs in embryogenesis, auxin is necessary for xylem cellspredeterminationin the root apical meristem. In proto- andmetaxylem predetermination, a key role is given to theSHR and miRNA165/166 mobile regulators . SHRis produced by xylem cells, from where the TF spreadstowards the periphery and, upon reaching the endodermis,activates the SCARECROW (SCR) TF, so they togetherinduce miRNA165/166 expression . MicroRNAs diffuse into neighboringcells, creating a concentration gradient towards the center ofthe root. In the stele, miRNA165/166 suppress the expressionof the genes encoding the TFs of the HD-ZIP III family, limitingit to the central domain . In such a way, themetaxylem cells are predetermined. Whether this mechanismworks in embryogenesis remains unknown, but this is a possibilitysince the PHABULOSA (PHB) TF of the HD-ZIP IIIfamily is expressed in the embryo root .Predetermination of phloem elementsThe phloem markers expressed in progenitors and induce thetissue development include a number of the DOF family TFs; strigolactonesignaling pathway suppressors SUPPRESSOR OF MAX21-LIKE 3 (SMLX3), SMLX4 and SMLX5 ; membrane proteins BREVIS RADIX (BRX), OCTOPUS(OPS), OPS-LIKE 2 (OPL2) ;phosphatase COTYLEDON VASCULAR PATTERN 2(CVP2) and its homolog CVP2-LIKE 1 (CVL1) ; the ALTERED PHLOEM DEVELOPMENT(APL) TF .The formation of protophloem elements is controlled byshifting the balance towards inducing or suppressing mechanismswith the central link connecting the opposing regulatorymodules being phloem-specific TFs of the DOF family. On the one hand, these TFs induce theexpression of phloem development activators, such as APLas well as their own genes, forming a positive feedback loop.On the other hand, DOFs induce the expression of CLE25,CLE26, and CLE45 signaling peptides migrating to neighboringcells where they trigger an inhibitory regulatory module. Interacting with the BARELY ANY MERISTEM(BAM) receptors and the CLAVATA3 INSENSITIVERECEPTOR KINASE (CIK) co-receptors, the CLE peptidesinduce the degradation of the DOF family TFs, suppressingthe formation of protophloem elements. The activity of theCLE peptide receptors can be additionally regulated, e. g., bythe MEMBRANE-ASSOCIATED KINASE REGULATOR 5(MAKR5)  or CORYNE (CRN)  regulators. The TFs of the DOF family activatethe expression of the genes encoding the OPS membraneprotein suppressing the BAM-CIK module .Properly positioned protophloem progenitor cells overcomethe inhibitory effect of CLE peptides due to the DOF TF accumulationdetermined by the positive feedback. Such a balancingmechanism makes it possible to repattern the phloemin case protophloem development has been disrupted . Here it should be noted that metaphloem developmentis probably regulated by other mechanisms and does notdepend on that of the protophloem .During phloem formation, the phloem/procambium stemcell divides anticlinally to produce a daughter procambiumand sieve-element progenitor to divide periclinally and forma procambium progenitor and a phloem sieve-element progenitor.The latter undergoes another periclinal division toproduce proto- and metaphloem progenitors . Companion-like cells are another product ofasymmetric division, but come from a different initial. Theseasymmetric cell divisions are controlled by a positional signal,a SHR-protein gradient whose migration into the endodermisactivates miRNA165/166 and induces asymmetric divisionsproducing companion-like cells, while SHR movement intothe phloem is necessary for the asymmetric divisions leadingto proto- and metaxylem formation .The vascular system of A. thaliana root is set at the earlieststages of embryogenesis. Wherein, the predetermination ofprovascular initials implies a labile, unstable, and reversiblespecification based on the physical arrangement of cells in theembryo and influenced by a complex regulatory network oftranscription factors. An interesting moment here is that bothxylem  and phloem  markers are jointly expressed by provascular initials inearly embryogenesis, but later they are separated into differentspatial domains in the provascular meristem and seedling.In A. thaliana, the vascular system is patterned by the timeof embryo maturation. Partially, the gene network that controlsthis process in embryogenesis continues to maintain thevascular system structure of the growing root of the seedlingand later during plant ontogenesis. This is associated withlocal accumulation of the molecular markers that are stablyexpressed in progenitor cells of a certain type. However,the factors working both in embryogenesis and during postembryonicdevelopment can act at these stages in differentwaysDespite the significant progress that has recently beenachieved in understanding the molecular and genetic mechanismsregulating vascular system development in plants,many questions remain open, in particular, those related tothe existence of parallel regulatory pathways and feedforwardloops. 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Neighboring parenchyma cells contribute to Arabidopsisxylem lignification, while lignification of interfascicular fibers iscell autonomous. Plant Cell. 2013;25(10):3988-3999. DOI 10.1105/tpc.113.117176.Stadler R., Wright K.M., Lauterbach C., Amon G., Gahrtz M., FeuersteinA., Oparka K.J., Sauer N. Expression of GFP-fusions in Arabidopsiscompanion cells reveals non-specific protein traffickinginto sieve elements and identifies a novel post-phloem domain inroots. Plant J. 2005;41(2):319-331. DOI 10.1111/j.1365-313X.2004.02298.x.Tanaka H., Dhonukshe P., Brewer P.B., Friml J. Spatiotemporal asymmetricauxin distribution: a means to coordinate plant development.Cell. Mol. Life Sci. 2006;63(23):2738-2754. DOI 10.1007/s00018-006-6116-5.Truernit E. Sieve elements and their cell neighbours in the Arabidopsisroot \u2013 roles and relationships. J. Plant Physiol. 2022;268:153569.DOI 10.1016/j.jplph.2021.153569.Tvorogova V.E., Osipova M.A., Doduyeva I.E., Lutova L.A. Interactionbetween transcriptional factors and phytohormones in regulationof plant meristems activity. Ekologicheskaia Genetika = EcologicalGenetics. 2012;10(3):28-40. (in Russian)Vera-Sirera F., De Rybel B., \u00darbez C., Kouklas E., Pesquera M.,\u00c1lvarez-Mahecha J.C., Minguet E.G., Tuominen H., Carbonell J.,Borst J.W., Weijers D., Bl\u00e1zquez M.A. A bHLH-based feedbackloop restricts vascular cell proliferation in plants. Dev. Cell. 2015;35(4):432-443. DOI 10.1016/j.devcel.2015.10.022.Verma S., Attuluri V.P.S., Robert H.S. An essential function for auxinin embryo development. Cold Spring Harb. Perspect. Biol. 2021;13(4):a039966. DOI 10.1101/cshperspect.a039966.Wabnik K., Robert H.S., Smith R.S., Friml J. Modeling framework forthe establishment of the apical-basal embryonic axis in plants. Curr.Biol. 2013;23(24):2513-2518. DOI 10.1016/j.cub.2013.10.038.Wallner E.S., L\u00f3pez-Salmer\u00f3n V., Belevich I., Poschet G., Jung I.,Gr\u00fcnwald K., Sevilem I., Jokitalo E., Hell R., Helariutta Y., Agust\u00ed J.,Lebovka I., Greb T. Strigolactone- and karrikin-independent SMXLproteins are central regulators of phloem formation. Curr. Biol.2017;27(8):1241-1247. DOI 10.1016/j.cub.2017.03.014.Weijers D., J\u00fcrgens G. Auxin and embryo axis formation: the ends insight? Curr. Opin. Plant Biol. 2005;8(1):32-37. DOI 10.1016/j.pbi.2004.11.001.Yang B., Minne M., Brunoni F., Pla\u010dkov\u00e1 L., Pet\u0159\u00edk I., Sun Y., Nolf J.,Smet W., Verstaen K., Wendrich J.R., Eekhout T., Hoyerov\u00e1 K.,van Isterdael G., Haustraete J., Bishopp A., Farcot E., Nov\u00e1k O.,Saeys Y., de Rybel B. Non-cell autonomous and spatiotemporalsignalling from a tissue organizer orchestrates root vascular development.Nat. Plants. 2021;7(11):1485-1494. DOI 10.1038/s41477-021-01017-6."}
+{"text": "Several antibiotics active against carbapenemase-producing organisms have recently been licensed for clinical use. These organisms usually have multiple integrative and conjugative elements with different resistance genes. The positioning of these agents for empirical use in sepsis caused by different carbapenemase-producing organisms is important in optimizing the likelihood of appropriate treatment before susceptibility testing is complete. Here we present the activities of meropenem-vaborbactam, imipenem-relebactam, eravacycline and tigecycline against NDM- and OXA-48-producing clinical isolates in a London teaching hospital.Klebsiella pneumoniae (17), Escherichia coli (10), Enterobacter aerogenes (3) and Serratia marcescens (1). NDM producers comprised K. pneumoniae (11), E. coli (10), Acinetobacter baumannii (3), Klebsiella oxytoca (1), Enterobacter asburiae (1), Enterobacter cloacae (1), Pseudomonas aeruginosa (1), Citrobacter koseri (1), Citrobacter freundii (1) and Leclercia adecarboxylata (1). MICs of meropenem/vaborbactam, imipenem/relebactam, eravacycline and tigecycline were determined using Etests. Interpretation was done using EUCAST breakpoints.Sixty-two previously characterized carbapenemase-producing Gram-negative clinical isolates were studied. These comprised 31 NDM and 31 OXA-48 producers. OXA-48 producers comprised OXA-48 producers: Eight of 17 K. pneumoniae, 8/10 E. coli and 2/3 E. aerogenes isolates were meropenem/vaborbactam susceptible. Of these, 2 K. pneumoniae, 5 E. coli and 1 E. aerogenes isolates were imipenem/relebactam susceptible. All imipenem/relebactam/susceptible isolates were also meropenem/vaborbactam susceptible. The S. marcescens isolate (1/1) was resistant to both antibiotics. The only isolates demonstrating susceptibility to the glycylcyclines were E. coli. Five of 10 were susceptible to both glycylcyclines. One of 10 was susceptible to one glycylcycline but resistant to the other. NDM producers: Five of 11 K. pneumoniae, 6/10 E. coli and 1/1 K. oxytoca, E. asburiae and C. koseri isolates were susceptible to meropenem/vaborbactam. Of these, two K. pneumoniae and the K. oxytoca (1/1) and E. asburiae (1/1) isolates were imipenem/relebactam susceptible. The L. adecarboxylata isolate (1/1) was meropenem/vaborbactam resistant but imipenem/relebactam susceptible. All P. aeruginosa, C. freundii, A. baumannii and E. cloacae isolates were resistant to both meropenem/vaborbactam and imipenem/relebactam. Seven of 10 E. coli isolates were tigecycline susceptible of which 6 were also eravacycline susceptible. One of 11 K. pneumoniae, 1/1 K. oxytoca and 1/1 L. adecarboxylata isolates were susceptible to both glycylcyclines. All other isolates were resistant to both glycylcyclines.E. coli than K. pneumoniae. Meropenem/vaborbactam provided better cover than imipenem/relebactam. Eravacycline and tigecycline showed similar activity to each other, but their activity was inferior compared with the \u03b2-lactam combinations, particularly against K. pneumoniae.In both OXA-48 and NDM producers, the activity of the four antibiotics tested was better against"}
+{"text": "Hcrt), tachykinin receptor 3 (Tacr3), cocaine and amphetamine-regulated transcript (Cart) and catecholamine-related biological processes; dopa decarboxylase (Ddc), histidine decarboxylase (Hdc), tyrosine hydroxylase (Th), and vasoactive intestinal peptide (Vip). In BACHD mice, few hypothalamic genes were differentially expressed compared to age-matched WT controls. However, GSEA indicated an enrichment of inflammatory- and gonadotropin-related processes at 10 months. In conclusion, we show that both wtHTT and mHTT overexpression change hypothalamic transcriptome profile, specifically mHTT, altering neuroendocrine circuits. In contrast, the ubiquitous expression of full-length mHTT in the BACHD hypothalamus moderately affects the transcriptomic profile.Structural changes and neuropathology in the hypothalamus have been suggested to contribute to the non-motor manifestations of Huntington\u2019s disease (HD), a neurodegenerative disorder caused by an expanded cytosine-adenine-guanine (CAG) repeat in the huntingtin (HTT) gene. In this study, we investigated whether hypothalamic HTT expression causes transcriptional changes. Hypothalamic RNA was isolated from two different HD mouse models and their littermate controls; BACHD mice with ubiquitous expression of full-length mutant HTT (mHTT) and wild-type mice with targeted hypothalamic overexpression of either wild-type HTT (wtHTT) or mHTT fragments. The mHTT and wtHTT groups showed the highest number of differentially expressed genes compared to the BACHD mouse model. Gene Set Enrichment Analysis (GSEA) with leading-edge analysis showed that suppressed sterol- and cholesterol metabolism were shared between hypothalamic wtHTT and mHTT overexpression. Most distinctive for mHTT overexpression was the suppression of neuroendocrine networks, in which qRT-PCR validation confirmed significant downregulation of neuropeptides with roles in feeding behavior; hypocretin neuropeptide precursor ( HTT) gene  is a fatal neurodegenerative disorder caused by a CAG repeat expansion in exon one in the huntingtin (TT) gene . The expTT) gene . The lenTT) gene . Current alleles , which sThe hypothalamus plays a primary role in the central-peripheral regulatory network that maintains body homeostasis, including regulating whole-body energy metabolism . MetabolTranscriptional dysregulation in the striatum is one of the hallmarks of HD . SpecifiAll the mice used in the study were housed in groups and maintained at a 12 h light/dark cycle with free access to a standard chow diet and water. All the experimental procedures performed on mice were carried out in accordance with the approved guidelines in the ethical permits approved by Lund University Animal Welfare and Ethics committee in the Lund-Malm\u00f6 region .Microarray profiling of the hypothalamic transcriptome was performed in Adeno-associated viral (AAV) vector-mediated groups of WT mice with targeted expression of wtHTT (AAV-HTT853-18Q) or mHTT (AAV-HTT853-79Q) fragments and BACHD mice that express full-length mHTT (97Q) . Both HDn = 5, 79Q: n = 8 and WT control: n = 5.AAV vector-mediated HD models achieve region-specific overexpression of HTT fragments in the brain through targeted injections using stereotactic surgery. AAV groups were assessed at 4 weeks post-injection since this was the earliest timepoint for a significant weight gain, as shown in previous studies . The vecn = 6 and WT: n = 6, and for 10 months of age: BACHD: n = 5 and WT: n = 3.Bacterial artificial chromosome-mediated transgenic mouse model is a transgenic mouse model of HD and ubiquitously expresses a full-length human mHTT  . In the Hypothalamic tissue was dissected on ice and snap-frozen in liquid nitrogen after a terminal dose of sodium pentobarbital  via intraperitoneal injection. Total RNA was extracted using the RNeasy Lipid Tissue Mini Kit  according to the manufacturer\u2019s instructions. RNA concentration and RNA quality measured in terms of RNA integrity number (RIN) were determined using the Agilent 2,100 Bioanalyzer . Samples with poor RIN (<7) were omitted from further analysis. Microarray analysis was performed on total hypothalamic RNA using the Affymetrix platform .Analyses were made using R v.4.1.1 . Raw .CEp-value followed by sorting it from the highest log2 (FC) to the lowest. The top 10 most upregulated and top 10 most downregulated significantly differentially expressed genes were identified, and their respective RMA-limma values were used to construct and color the heatmap. Heatmaps were generated using the pheatmap package in R1 (v. 1.0.12) (Limma output files for 18Q vs. WT and 79Q vs. WT were sorted based on adj.  1.0.12) . Hierarcp-value < 0.05. The 18Q vs. WT and 79Q vs. WT datasets were subsequently sorted into three gene sets: shared genes, unique genes for 18Q vs. WT, and unique genes for 79Q vs. WT. The shared and unique gene lists (ENTREZ IDs) were imported into DAVID2 . The reported p-values were further adjusted using the Benjamini-Hochberg procedure to correct for multiple testing. Outputs from GSEA can be found in ClusterProfiler (v.4.0.5) was used to perform GSEA . The funTo synthesize cDNA, 1 \u03bcg of RNA from each sample was reverse transcribed using SuperScript IV Reverse Transcriptase SuperScript IV kit  according to the manufacturer\u2019s instructions. Mouse qRT-PCR primers were designed using Primer3Plus software . qRT-PCRp-value < 0.05 considered statistically significant.Statistical analysis of qRT-PCR data was performed using Graphpad Prism 9 . . Data were analyzed using non-parametric Mann-Whitney U tests with a p-value in the limma datasets comparing injected mice to uninjected (18Q vs. WT and 79Q vs. WT) showed a skewed distribution with a preference for upregulated genes in the injected mice  of the microarray data showed a clear separation of the AAV-HTT groups from the WT samples . PCA of ted mice . Filteri5 cutoff .p < 0.05 cutoff, 1,422 variables remained for BACHD 2 months vs. WT  variables, and we further analyzed the data using the Database for Annotation, Visualization and Integrated Discovery (DAVID) (see text footnote 2) (p < 0.05) identified by limma between the 18Q vs. WT and 79Q vs. WT datasets (The 18Q vs. WT and 79Q vs. WT datasets had the highest number of significant (adj. tnote 2) . Previoutnote 2) . Furthertnote 2) . Therefodatasets , using tdatasets . DAVID Fn = 51 genes, (adj. p < 0.05) to 79Q vs. WT (adj. p > 0.05)] showed that the gene with the highest difference in log2(FC) was immunoglobulin heavy chain (X24 family) (Igh-VX24) followed by tripartite motif-containing 30D (Trim30d) and toll-like-receptor 7 (Tlr7) . Across all 51 genes, the mean difference with 95% CI was 0.135   > 0 and 61 with log2(FC) < 0 . Functio, 0.181) , 2.n = 12 genes, adj. p < 0.05) to 18Q vs. WT (adj. p > 0.05) showed a mean difference with 95% CI of -0.042 . In the top 5/10 cluster related to neuroactive ligands , we found a mean difference in log2(FC) of -0.067  between the 79Q vs. WT and 18Q vs. WT datasets, where the genes with the highest difference were proenkephalin (Penk) and tachykinin receptor 3 (Tacr3)   > 0 and 131 with log2(FC) < 0 . Among t: -0.24] , 2.Ldlr; log2(FC) 18Q vs. WT: \u22120.49, 79Q vs. WT: \u22120.54] (The shared gene list between 18Q vs. WT and 79Q vs. WT consisted of 410 genes with log2(FC) > 0 and 39 genes with log2(FC) < 0 . An immu: \u22120.54] , 2.p and log2(FC)] in the 18Q vs. WT and 79Q vs. WT datasets , and major facilitator superfamily domain-containing 2A (Mfsd2a) (Nms), Hdc, synaptic vesicle glycoprotein 2C (Sv2c), and Tacr3  , genes tnd Tacr3 . CompariNext, we elaborated further on the functional implications of the differences in the top 10 downregulated genes between 18Q vs. WT and 79Q vs. WT. GSEA of GO-BP with leading-edge analysis  was usedp-value < 0.05 for the 79Q vs. 18Q and BACHD datasets. By using GSEA, we may still identify biologically relevant pathways that exhibit noteworthy cross-correlation between genes with a subtle change in expression levels or weak statistical significance  was -0.156 . The leading- edge gene with the lowest log2(FC) was prolactin (Prl) at -1.63 followed by growth hormone (Gh) -1.02, Vip -0.59, the alpha subunit of glycoprotein hormones (Cga) -0.53 and hypocretin (Hcrt) -0.35  -0.35 , 4.Acot5) and vesicular glutamate transporter 1 (Slc17a7) that were respectively part of the fatty-acid related and \u201cGlutamatergic synapse\u201d pathways . None of the three pathways identified in 2 months old BACHD vs. WT were significant in GSEA-KEGG for 10 months old BACHD vs. WT. When comparing each leading-edge gene set in 2 months old BACHD vs. WT to the same genes in 10 months BACHD vs. WT, the mean log2(FC) difference was <0.2 for all three pathways. Instead, for 10 months old BACHD vs. WT, fourteen KEGG pathways were identified (H2-Aa), and Cd74 were among the leading-edge genes with the highest log2(FC) and difference from the 2 months of age vs. WT dataset . Pathways related to gonadotropin-releasing hormone (Gnrh), all NES > 0, were also found in 10 months old BACHD vs. WT, where the alpha subunit of glycoprotein hormones (Cga) was among the leading-edge genes with the highest fold change . Only one of the 14 KEGG pathways identified by GSEA had a negative NES of -1.95, indicating suppression; \u201cOxidative phosphorylation\u201d (mmu00190). The overall change of the 59 genes in the leading-edge gene set was -0.056  [mean log2(FC) with 95% confidence interval (95% CI)].For 2 months old BACHD vs. WT, GSEA-KEGG identified three pathways; \u201cGlutamatergic synapse\u201d , \u201cBiosynthesis of unsaturated fatty acids\u201d  and \u201cFatty acid elongation\u201d  . Among lentified . Immune-entified . HistocoHcrt) in mice with mHTT 79Q overexpression compared to wtHTT 18Q groups and uninjected WT controls  that was among the top suppressed processes in GSEA-GO BP consisted of 57 genes . We havecontrols . Here, imparison .Hdc as part of the top 10 downregulated genes and related leading-edge subsets in GSEA. Comparing GSEA-GO BP outputs between the 79Q vs. WT and 18Q vs. WT datasets showed a higher number of catecholamine-related processes that were only significant for the 79Q vs. WT dataset  or Growth hormone-releasing hormone (Ghrh)  . A matchransport .Cart, Tacr3, Hcrt, Vip, Th, and Ghrh, only Tacr3 was significantly downregulated compared to the age-matched WT controls.Next, we performed qRT-PCR of a subset of genes analyzed for the HTT-AAV groups in the BACHD 2 months group . Among CHdc mRNA have been reported in HD patients . Among leading-edge genes was Cga, which encodes the alpha subunit of glycoprotein hormones   . Furthereriphery , notableeriphery . Expresseriphery . Furthereriphery . Similareriphery . In lineeriphery . As HD periphery . TherefoStoml3 and Sv2c associated with dopaminergic signal transmission were among the top 10 differentially downregulated in 79Q vs. WT .RS-K, \u00c5P, ED, and MB designed the experiments from RS-K and \u00c5P. ED, AD, RS-K, and SL performed the experiments. ED, NA, AD, and RS-K analyzed the data. ED, RS-K, and AD wrote the first draft of the manuscript. All authors reviewed the manuscript and approved the final version."}
+{"text": "TM) can ameliorate liver fibrogenesis induced by thioacetamide (TAA) treatment. Our results showed that the TAA treatment caused lower body weight gains and enlarged livers, as well as higher serum ALT, AST, and ALP levels (p < 0.05). This liver inflammatory and fibrotic evidence was ameliorated (p < 0.05) by supplementing with GBHP01TM; this partially resulted from its antioxidant abilities, including decreased TBARS values but increased TEAC levels, reduced GSH contents and catalase/GPx activities in the livers of TAA-treated rats (p < 0.05). Additionally, fewer nodules were observed in the appearance of the livers of TAA-treated rats after supplementing with GBHP01TM. Similarly, supplementing GBHP01TM decreased fibrotic scars and the fibrotic score in the livers of TAA-treated rats (p < 0.05). Moreover, the increased hepatic IL-6, IL-1\u03b2, and TNF-\u03b1 levels after TAA treatment were also alleviated by supplementing with GBHP01TM (p < 0.05). Meanwhile, GBHP01TM could decrease the ratio of LC3B II/LC3B I, but upregulated P62 and Rab7 in the livers of TAA-treated rats (p < 0.05). Taking these results together, the CLH-based supplement (GBHP01TM) can be characterized as a natural agent against liver fibrogenesis.Chicken-liver hydrolysates (CLHs) have been characterized as performing several biofunctions by our team. This study aimed to investigate if a CLH-based supplement (GBHP01 Liver cancer ranks third among cancer categories globally, and approximately 830,000 people die with liver cancer every year . SimilarFood-derived protein hydrolysates have been suggested to provide functional properties such as antioxidant , lipid-lCandidates for functional-food ingredients should be either effective or low in cost. Regarding the poultry industry in Taiwan, approximately 10,000 metric tons of chicken livers (2.5% based on broilers\u2019 wt.) have been produced in recent years  and antiTM) containing 200 mg CLH powder and 20 mg hesperidin  per capsule was generously offered by Great Billion Biotech Co., Ltd., New Taipei City, Taiwan. The composition of free amino acids and imidazole-ring dipeptides in a GBHP01TM capsule were sent for analysis using an Amino Acid Analyzer  in the Food Industry Research and Development Institute . The total contents of free essential and non-essential amino acids and imidazole-ring dipeptides are 13.10, 65.04, and 0.07 mg per capsule (average 650 mg), respectively, where the total free branched-chain amino acid (BCAA) and taurine content are 5.77 mg and 51.67 mg per capsule ; (2) TAA: 100 mg TAA/kg BW (i.p.) + 1.5 mL pure H2O ; (3) TAA + SIL: 100 mg TAA/kg BW (i.p.) + 150 mg silymarin/kg BW in 1.5 mL pure H2O ; (4) TAA + GBHP01(1X): 100 mg TAA/kg BW (i.p.) + 133 mg GBHP01TM content/kg BW in 1.5 mL pure H2O ; (5) TAA + GBHP01(2X): 100 mg TAA/kg BW (i.p.) + 266 mg GBHP01TM content/kg BW in 1.5 mL pure H2O . It was reported that silymarin could ameliorate livre fibrogenesis via decreasing endoplasmic reticulum stress-related gene expressions, inflammatory cytokines, collagen accumulation, and matrix metalloproteinase (MMP-2 and MMP-9) activities, as well as increasing tissue inhibitors of metalloproteinase gene expressions (TIMP-1 and TIMP-3) in livers . Additionally, serum ALB values were reduced (p < 0.05) by TAA treatment, except in the TAA + GBHP01(2X) group. Focusing on serum lipid levels, no (p > 0.05) differences on TC levels were observed among groups. The TAA group had the lowest (p < 0.05) serum TG levels among groups, but a reverse (p < 0.05) effect was observed in silymarin and GBHP01TM-supplemented groups, while TAA + SIL and TAA + GBHP01(2X) groups even showed similar (p > 0.05) serum TG levels to the CON group.After 8 weeks of experiment, TAA treatment resulted in the lower body weight of rats (approximately a 12.19% reduction), while lower weight increases and smaller food intakes were observed in TAA-treated rats than in control rats ( < 0.05) . No , while the silymarin or GBHP01TM-supplemented group had similar (p > 0.05) TBARS to the CON group and no (p > 0.05) difference in TEAC levels among GBHP01TM-supplemented groups and the CON group. Besides, the decreased pattern of reduced GSH levels was observed in TAA-treated groups, but silymarin or GBHP01TM supplementation allowed for maintenance of the reduced GSH level in TAA-treated rats, especially in the group receiving 2X doses of GBHP01TM supplementation (p < 0.05). With regard to antioxidant enzymatic activities, SOD activities were not (p > 0.05) different among groups. The TAA group showed reduced (p < 0.05) CAT and GPx activities compared to CON group, while silymarin or GBHP01TM supplementation avoided reducing CAT activities, showing a result similar (p > 0.05) even to that of the CON group; reversed (p < 0.05) effects on GPx activities were only observed in GBHP01TM supplemented groups. In hepatic proinflammatory cytokines, the TAA group had higher (p < 0.05) hepatic IL-1\u03b2 , IL-6 , and TNF-\u03b1 (28.89% increase) than the CON group. GBHP01TM supplementation reduced (p < 0.05) these increased hepatic inflammatory cytokines of TAA-treated rats to levels similar (p > 0.05) even to those of the CON group, but silymarin supplementation only reduced the hepatic IL-6 level.As the results in TM supplementation. Via H&E stained observation  according to the H&E stains were increased by TAA treatments, while significant reductions (p < 0.05) in all three HAI scores were observed in the GBHP01TM supplementation group; however, silymarin only demonstrated a reduced pattern  score. Silymarin treatment did not (p > 0.05) attenuate the liver Metavir score in the TAA-treated rats. Treatment with 1X or 2X GBHP01 supplement decreased (p < 0.05) the scores in TAA-treated rats, although the scores were still higher (p < 0.5) than those of the CON group , but GBHP01TM supplement significantly downregulated (p < 0.05) this ratio, reaching levels even similar (p > 0.05) to those of CON (p < 0.05) by TAA treatment (TM supplementation significantly upregulated (p < 0.05) these two proteins in livers of TAA-treated rats; meanwhile, the P62 and Rab7 protein expression in the livers of the TAA + GBHP01(1X) and TAA + GBHP01(2X) groups were reversed to similar levels to those of the CON group (p > 0.05).With regard to autophagy modulation in livers of TAA-treated rats, the ratio of LC3B II/LC3B I of TAA group was almost 1.8 relative fold to that of CON group  and hesperidin, which are both characterized by antioxidant abilities , only 4.1 and 8.2 mg hesperidin/kg BW were given to rats. Hence, it was assumed that the amelioration of TAA-induced liver fibrosis by hesperidin can be considered negligible in this study. To summarize, this regulation probably mainly results from the bioactive ingredients in the CLHs of GBHP01TM supplements  . Accordin (IL10) , the revsurvival . Recentlsurvival  and incrsurvival . It was survival . It was survival . Besidessurvival . This phsurvival ,37. LC3Bsurvival , while Rsurvival . As the survival . Meanwhisurvival . Taurinesurvival . Similarsurvival . As explturation . Althougin/kg BW ,43,44. BTM) showed an ameliorating effect against liver fibrogenesis induced by TAA treatment  against liver fibrogenesis may be attributed to the bioactive ingredients in CLHs, including anserine, taurine, glutamic acid, aspartic acid, BCAAs, etc.In this study, chicken liver hydrolysate (CLH)-based supplements (GBHP01reatment . The hep"}
+{"text": "Anguilla japonica) are commercially important species, harvested extensively for food. Currently, this and related species (American and European eels) are challenging to breed on a commercial basis. As a result, the wild stock is used for aquaculture. Moreover, climate change, habitat loss, water pollution, and altered ocean currents affect eel populations negatively. Accordingly, the International Union for Conservation of Nature lists Japanese eels as endangered and on its red list. Here we presented a high-quality genome assembly for Japanese eels and demonstrated that large chromosome reorganizations occurred in the events of third-round whole-genome duplications (3R-WRDs). Several chromosomal fusions and fissions have reduced the ancestral protochromosomal number of 25 to 19 in the Anguilla lineage. A phylogenetic analysis of the expanded gene families showed that the olfactory receptors (group \u03b4 and \u03b6 genes) and voltage-gated Ca2+ channels expanded significantly. Both gene families are crucial for olfaction and neurophysiology. Additional tandem and proximal duplications occurred following 3R-WGD to acquire immune-related genes for an adaptive advantage against various pathogens. The Japanese eel assembly presented here can be used to study other Anguilla species relating to evolution and conservation.Japanese eels ( Fishes are highly diverse species living in many ecological habitats, including freshwater, estuarine, and the ocean . Over 99From the evolutionary perspective, eels are among the extant basal groups of teleost ray-finned fishes after the 3-round whole-genome duplication (3R-WGD) . The rayAnguilla japonica , was kept in a freshwater tank for a week with aeration. Blood sample was taken from the fish, snapped frozen in liquid nitrogen, and then stored at \u221280\u00b0C. Genomic DNA was extracted from the blood sample. DNA sequencing data were generated by different platforms, including Oxford Nanopore (ONT) long reads, PacBio continuous long reads (CLRs), Illumina short reads, Illumina mate-pair reads, 10\u00d7 Chromium linked reads, DNase Hi-C (Omni-C), and Bionano optical mapping .A market-purchased female Japanese eel, RRID:SCR_006255) 3.2.10. For PacBio CLR sequencing, the SMRTbell templates were prepared using Sequel Binding Kit 1.0 and sequenced on the PacBio Sequel System . For Illumina short reads and mate-pair sequencing, the libraries were prepared using the TruSeq DNA PCRFree Kit  and Nextera Mate Pair Library Preparation Kit  (gel plus), respectively. They were sequenced with 2\u00d7 150-bp reads on an Illumina HiSeq X Ten  instrument. The library for linked reads was prepared by a 10\u00d7 Genomics Chromium  system with the Chromium Genome library (v2) and sequenced with 2\u00d7 150-bp reads on an Illumina NovaSeq 6000  instrument. Dovetail Omni-C Kit  was used for Hi-C library preparation, which used NEBNext Ultra  enzyme and Illumina-compatible adapters. Biotin-containing fragments were isolated using streptavidin beads before PCR enrichment.\u202fThe library was sequenced with 2\u00d7 150-bp reads on an Illumina HiSeqX platform. The Bionano optical mapping was generated by 3 enzymes, 2 from Irys (Nt.BspQI and Nb.BssSI) and 1 from Saphyr (RRID:SCR_017992) (DLE1). We stretched and captured the images of fluorescently labeled DNA molecules in Irys and Saphyr G1.2 chips. The labeling distances were extracted from the images and recorded into the raw molecule files. Molecules over 150\u00a0Kbp were assembled into consensus maps using Bionano Solve for further analysis (The library for ONT long-read sequencing was prepared using the Ligation Sequencing Kit (LSK109) and sequenced using the Nanopore PromethION P48  sequencer with the flow cells (R9.4.1) and the basecaller version Guppy  v2 [RRID:SCR_017225) v2.5 [RRID:SCR_017016) v2.71 [RRID:SCR_017642) v1.4.16 [RRID:SCR_014731) v1.23 [MitoZ software (v2.4)  was used5880) v2 , Wtdbg2 6) v2.71  separate6) v2.71  to achie1) v1.23  for 2 roRRID:SCR_021172) v1.11.08 [We applied Tigmint v1.1.2  and ARKSv1.11.08  to extende novo approaches to annotate transposable elements (TEs) in the Japanese eel genome. For the homolog-based approach, RepeatMasker v4.0.7 [RRID:SCR_021169) v21.12 database [RRID:SCR_015247) v1.06 [de novo approach, RepeatModeler (RRID:SCR_015027) v1.0.8 was used to detect the TE families and repeat boundaries by integrating 3 complementary de novo repeat finding programs. RepeatMasker collected the union of these tools' results and annotated the genome accordingly.Tandem Repeats Finder v4.09  was applr v4.0.7  and Repedatabase  to the g7) v1.06  was usedde novo, homology-based, and transcriptome-based annotations. Maker (RRID:SCR_005309) v2.31.8 [Anguilla anguilla), zebrafish (Danio rerio), Indo-Pacific tarpons , Asian arowana (Scleropages formosus), and spotted gar (Lepisosteus oculatus), based on the phylogeny of teleost fishes [Three types of methods were used to annotate the protein-coding genes in the genome, including  v2.31.8  was adopt fishes .De novo annotation was performed using Augustus (RRID:SCR_008417) v3.2.1 [RRID:SCR_002127) v1 [RRID:SCR_015530) v2.1.0 [RRID:SCR_013048) v2.10.0 [) v3.2.1  and SNAP2127) v1  by train) v2.1.0  and asseGene functional annotation was performed by aligning the predicted gene sequences to protein sequences using BLAST v2.2.31  in the sRRID:SCR_015008) v5.1.2 [RRID:SCR_011980), to compare.BUSCO  v5.1.2  was usedRRID:SCR_010835) 1.3.1 [RRID:SCR_001598) [RRID:SCR_007891) v12 [tRNAscan-SE (5) 1.3.1  was used_001598) . The mic891) v12  to the gRRID:SCR_006086) v2.2.3 [Anguilla rostrata , A. anguilla , A. japonica (Japanese eel), M. cyprinoides , S. formosus , Gadus morhua , Oryzias latipes , D. rerio ,L. oculatus , Erpetoichthys calabaricus , Latimeria chalumnae , and Callorhinchus milii . We estimated the divergence times for single-copy orthologos using mcmctree in PAML (RRID:SCR_014932) package v4.8a [RRID:SCR_021162) website: D. rerio with O. latipes (180.0\u2013264.0 Mya), M. cyprinoides with A. anguilla (162.2\u2013197.3 Mya), C. milii with D. rerio (442.7\u2013515.5 Mya), and E. calabaricus with D. rerio (381.0\u2013407.0 Mya). To estimate gene family expansion and contraction, we used CAF\u00c9 (RRID:SCR_005983) v4.2.1 [OrthoMCL (v2.0)  was used) v2.2.3  to recon) v4.2.1  to modelRRID:SCR_011822) v2.2.26 [RRID:SCR_008493) v6.6.0 [RRID:SCR_001010) v2.2.26 to remove sequences that did not match genes already known in SwissProt and NR. InterProscan was used to determine the secondary structures of the predicted OR genes. Some genes were filtered due to lacking the 7 transmembrane domains. The maximum likelihood phylogenetic tree was reconstructed using IQ-TREE (RRID:SCR_017254) v2.2.0.3 [RRID:SCR_011811) v7.505 [We identified olfactory receptor (OR) genes using the pipeline described in GitHub , while c v2.2.26  was used) v6.6.0 . Using Ev2.2.0.3  based on) v7.505 .A. japonica, A. anguilla, A. rostrata, M. cyprinoides,and Lepisosteus oculatus. The numbers of nonsynonymous substitutions (Ka) and synonymous substitutions (Ks) were calculated using KaKs_calculator2.0 [MCscanX v1.5.1  and macrlator2.0 . In addilator2.0 , using sA. japonica (Japanese eel), M. cyprinoides (tarpon), and S. formosus . The ancestral teleosts karyotype (ATK) was constructed using zebrafish, S. formosus (arowana), and L. oculatus  [Ancestral eel/tarpon karyotype (AETK) was constructed using t-group) . This wat-group)  to obtait-group) .AB038556.2) of the NR database from NCBI  [In this study, MitoZ software was used to assemble and annotate the mitochondrial genome 16.686 Kb) of our sample to confirm the species' identity . The data matched with the Japanese eel mitochondrial genome , we identified 29,982 coding genes  loci in the 1,131 single-copy orthologs from the 12 fish species  and Cypriniformes  diverged from the Eloposteoglossocephala clade at 262.5 Mya. Above are fish groups that had undergone 3R-WGD. Compared to the out-groups, spotted gars, reed fish, coelacanths, and Australian ghost sharks underwent only 2 rounds of whole-genome duplication (2R-WGD).The orthology analysis of 12 species' coding genes identified 21,653 gene family clusters. The expansion and contraction of gene families reflect the evolution of organisms' adaptations to their environments. Ortholog analysis of genes from the 12 species  identified 21,652 gene family clusters. By removing gene families with too many (\u2265200) or too few (\u22642) genes, we achieved 129,862 genes to evaluate the expansion and contraction of gene families Fig.\u00a0. Among t2+ and K+ channel families were identified. Calcium and potassium play significant roles in neuronal excitability, muscle contraction, fertilization, and energy metabolism. Interestingly, the other expanded gene families include (i) the assembly of thick myosin filament in skeletal muscle, (ii) lipoprotein receptor\u2013related protein (metabolic and morphogenetic pathways), and (iii) isocitrate and isopropyl malate dehydrogenases family (carbohydrate and amino acid metabolism).Comparing the Japanese eel to the other 11 species, 433 gene families increased, with a total increase of 551 genes. On the other hand, a total of 943 genes were lost from 782 gene families . It is iA. japonica, A. Anguilla, and M. cyprinoides, respectively  had many paralogous pairs after splitting from the Osteoglossomorpha lineage . The obsely Fig.\u00a0. Additioely Fig.\u00a0. By idenThere are 21,249 duplicated genes identified among the 29,982 coding genes in the Japanese eel genome. Based on their duplication patterns, DupGen_finder  classified the duplicated genes into 5 categories: (i) 9,890 WGDs (46.54%), (ii) 1,420 tandem duplicates , (iii) 768 proximal duplicates , (iv) 3,975 transposed duplicates , and (v) 5,196 dispersed duplicates . We then calculated the Ks and Ka/Ks values for these 5 gene categories. Ks distribution indicates that TD and PD revealed additional duplication after 3R-WGD Fig.\u00a0. In addin) is 25 for tarpons, arowana, and zebrafish; 24 for medaka; and 23 for Atlantic cod. Japanese eels have a lower haploid chromosome number (n = 19). To assess the extent of interchromosomal rearrangements in Japanese eels, we reconstructed the karyotype of the common ATK and AETK  in tarpons. Comparatively, the 19 AETK chromosomes  underwent 24-fusion and 18-fission to form the 13 chromosomes  in Japanese eels (n = 19) in Japanese eels, of which Chr 1, and Chr 3\u20137 rearrangements are unique to Japanese eels and might play a role in speciation. Japanese eel's chromosomes  were derived from AETK's  with slight rearrangements. The patterns of chromosome rearrangements in Chr 8, 13, and 16\u201317 of Japanese eels were comparable with Chr 21, 6, 8, and 19 in tarpons. Without rearrangement, Japanese eel's chromosomes 10, 14, and 19 were equivalent to AETK's chromosomes 3, 6, and 22. In addition, 3 chromosomes in the Japanese eel  derived directly from AETK's chromosomes , which also correspond to tarpon's chromosomes , respectively. Figure\u00a0When comparing chromosome numbers of the fishes that had all undergone 3R-WGD, the haploid chromosome number  and zeta (\u03b6) group genes in the freshwater eels expanded enormously, comprising about 86% of the entire gene family. Delta (\u03b4) and \u03b6 belong to the type I genes , which a2+ channels were the significantly expanded gene families in Japanese eels. Genome studies suggest that the cellular functions of voltage-gated ion channels emerged early in Metazoan evolution [2+ during migration between waters with great variations of calcium contents. A gene expression study in marbled eel (Anguilla marmorata) showed high expression of voltage-gated Ca2+channels in brain, skin, and osmoregulatory tissues  and its response to changes in water calcium levels [2+ homeostasis, Ca2+ signaling coordinates various physiological processes, including skeletal muscle contractions, nervous system activity, and cardiac and reproductive functions. The expanded gene families of thick myosin filament in skeletal muscle imply enhanced coordination of muscle contraction and performance [The voltage-gated Cavolution , 85 in dm levels . Besidesformance , especiaformance . Additioformance , 90. Isoformance . The dupformance . These eformance . Notablyformance  and sturformance .n) of 26 [n = 24 or 25 in extant teleost species. In this study, we reconstructed the ancestral protochromosomes AETK (n = 25) to describe the cross-species chromosome collinearity and underpin the lineage-specific genome reorganization. The chromosome number of Anguillaspecies (n = 19) was reduced as compared with M. cyprinoides (n = 25) and S. formosus (n = 25). The Anguilliformes is made up of 15 families with remarkable karyotypic diversity [n = 19 and 21. The Anguilla lineage underwent a significant structural rearrangement upon their divergence from the common ancestor of tarpons (M. cyprinoides). The fusion and fission of their chromosome structure were the primary drivers of reducing the haploid chromosome number to 19.The acquisition of evolutionary novelty by WGD duplication and the subsequent fate change of duplicated genes is necessary for phenotype alteration, environmental adaptation, and speciation . The larn) of 26 . Evolutiiversity . The hapA. japonica whole genome sequencing and assembly are publicly available on NCBI databases under the accession number PRJNA852364. The gene models are available at Zenodo [GigaScience GigaDB database [The t Zenodo . All supdatabase .Supplementary Fig. S1. The alignment plot shows our mitochondrial genomes assembled and GenBank ID AB038556.2 of Japanese eels.Supplementary Fig. S2. Circos plot for the mitochondrial genome of Japanese eel. From outer to inner circles: protein-coding genes, rRNA, and tRNA; depth of Illumina short reads; and GC content.Supplementary Fig. S3. Multiplatform Japanese eel genome assembly.Supplementary Fig. S4. Genome comparison of Japanese (A. japonica) and European (A. anguilla) eels.Supplementary Fig. S5. Venn diagram of gene annotation based on 5 databases .Supplementary Fig. S6. KEGG-based gene function classification. The numbers represent how many genes are in the particular functions.Supplementary Fig. S7. KOG-based gene function classification. The numbers represent how many genes are in the particular functions.Supplementary Fig. S8. Length distribution of messenger RNA, CDS, exon, intron, and the number of exons in Japanese eel and the other species .Supplementary Fig. S9. Ks distributions of syntenic paralogs and orthologs. Ks value distribution is used to identify genome duplication and speciation.Supplementary Fig. S10. Comparative genomic analysis of Japanese eels (A. japonica), tarpons (M. cyprinoides), and arowanas (S. formosus).Supplementary Table S1. Genome sequencing platforms for A. japonica.Supplementary Table S2. A summary of contig statistics from the ONT long-read assembly.Supplementary Table S3. A summary of contig statistics after assembly error correction.Supplementary Table S4. Scaffolding by 10\u00d7 linked reads, Bionano optical mapping, and Hi-C.Supplementary Table S5. The completeness of A. japonica genome by BUSCO assessment.Supplementary Table S6. Statistical results for repeat sequences.Supplementary Table S7. A statistical analysis of the classification results for TE.Supplementary Table S8. Functional annotation of predicted genes from A. japonica.Supplementary Table S9. The completeness of A. japonica genes by BUSCO assessment.Supplementary Table S10. The average length of exons in Japanese eel and the 8 related fish species.Supplementary Table S11. GO enrichment analysis of the gene families expanded in the 3 freshwater eel genomes.Supplementary Table S12. GO enrichment analysis of the gene families expanded in the A. japonica genome.Supplementary Table S13. The karyotypes of M. cyprinoides (tarpons) and the common ancestor of eels and tarpons (AETK).Supplementary Table S14. The karyotypes of A. japonica (Japanese eel) and the common ancestor of eels and tarpons (AETK).giac120_GIGA-D-22-00177_Original_SubmissionClick here for additional data file.giac120_GIGA-D-22-00177_R1_Revision_1Click here for additional data file.giac120_GIGA-D-22-00177_Revision_2Click here for additional data file.giac120_Response_to_Reviewer_Comments_Original_SubmissionClick here for additional data file.giac120_Response_to_Reviewer_Comments_Revision_1Click here for additional data file.giac120_Reviewer_1_Report_Original_SubmissionChristiaan Henkel -- 8/9/2022 ReviewedClick here for additional data file.giac120_Reviewer_1_Report_Revision_1Christiaan Henkel -- 10/6/2022 ReviewedClick here for additional data file.giac120_Reviewer_2_Report_Original_SubmissionZhong Li -- 8/22/2022 ReviewedClick here for additional data file.giac120_Supplemental_Figures_and_TablesClick here for additional data file.AETK: ancestral eel/tarpon karyotype; ATK: ancestral teleosts karyotype; BLAST: Basic Local Alignment Search Tool; bp: base pair; BUSCO: Benchmarking Universal Single-Copy Orthologs; CLR: continuous long read; DSD: dispersed duplicate; Gb: gigabase; Kbp: kilobase pair; KEGG: Kyoto Encyclopedia of Genes and Genomes; Mb: megabase; Mbp: megabase pair; Mya: million years ago; NCBI: The National Center for Biotechnology Information; ONT: Oxford Nanopore; OR: olfactory receptor; ORF: open reading frame; PD: proximal duplicate; rRNA: ribosomal RNA; TD: tandem duplicate; TE: transposable element; TRD: transposed duplicate; tRNA: transfer RNA; WGD: whole-genome duplication; 2R-WGD: 2-round whole-genome duplication; 3R-WGD: 3-round whole-genome duplication; 4R-WGD: 4-round whole-genome duplication.The experimental plan and sequencing strategy were designed by C.K.C.W., E.L.Z., A.O.L.W., K.P.L., and T.F.C. Samples were collected by A.H.M.N. and H.T.W. Bionano optical mapping and data analysis were conducted by C.Y.L.C., E.Y.C.C., and J.Z. The sequencing data for genome assembly were analyzed by E.L.Z., H.W., B.W., J.J., E.Y.C.C., and T.F.C. The manuscript was written by C.K.C.W., H.T.W., E.L.Z., H.W., and A.O.L.W."}
+{"text": "Bifidobacterium bifidum DS0908 (DS0908) and Bifidobacterium longum DS0950 (DS0950). Treatment with DS0908 and DS0950 postbiotics significantly induced the expression of the brown adipocyte-specific markers UCP1, PPAR\u03b3, PGC1\u03b1, PRDM16 and beige adipocyte-specific markers CD137, FGF21, P2RX5, and COX2 in C3H10T1/2 mesenchymal stem cells (MSCs). In mice with high-fat diet (HFD)-induced obesity, both potential probiotics and postbiotics noticeably reduced body weight and epididymal fat accumulation without affecting food intake. DS0908 and DS0950 also improved insulin sensitivity and glucose use in mice with HFD-induced obesity. In addition, DS0908 and DS0950 improved the plasma lipid profile, proved by reduced triglyceride, low-density lipoprotein, and cholesterol levels. Furthermore, DS0908 and DS0950 improved mitochondrial respiratory function, confirmed by the high expression of oxidative phosphorylation proteins, during thermogenesis induction in the visceral and epididymal fat in mice with HFD-induced obesity. Notably, the physiological and metabolic changes were more significant after treatment with potential probiotic culture-supernatants than those with the bacterial pellet. Finally, gene knockdown and co-treatment with inhibitor-mediated mechanistic analyses showed that both DS0908 and DS0950 exerted anti-obesity-related effects via the PKA/p38 MAPK signaling activation in C3H10T1/2 MSCs. Our observations suggest that DS0908 and DS0950 could potentially alleviate obesity as dietary supplements.Probiotic supplements have promising therapeutic effects on chronic diseases. In this study, we demonstrated the anti-obesity effects of two potential probiotics,  The mice were administered DS0908 or DS0950 pellets  were fixed in 4% paraformaldehyde, embedded in paraffin, and cut into 10-\u03bcm sections. Standard H&E staining was performed using standard protocols. Five random fields of each section were evaluated, and the average adipocyte diameter was measured using the ImageJ software .Tbp was an internal control. We harvested C3H10T1/2 MSCs and extracted total RNA using an RNA Extraction Kit  following the manufacturer\u2019s guidelines. We used 1 \u03bcg RNA to synthesize cDNA using the Maxime RT PreMix Kit  on a Veriti 96-Well Thermal Cycler . qRT-PCR was performed using an iQ SYBR Green Supermix Kit  on a CFX96 Real-Time PCR Detection System (Bio-Rad). + with ImageLab (Bio-Rad). Anti-\u03b2-actin antibody was used as a loading control for each protein expression. Protein band intensities were quantified using the ImageJ software.Differentiated and treated C3H10T1/2 MSCs were prepared in RIPA lysis buffer supplemented with protease inhibitors. The total protein concentration was measured using the BCA Protein Assay Kit (Thermo Fisher Scientific). Equal amounts of protein samples were separated on a 4\u201320% sodium dodecyl sulfate-polyacrylamide gradient gel  and transferred onto polyvinylidene difluoride (PVDF) membranes. The PVDF membranes were then blocked and incubated with specific antibodies, as indicated in the figures. Immunoreactive protein bands were captured using the chemiluminescent ECL  assay on ChemiDoc XRSC3H10T1/2 MSCs were seeded on 6-well plates and grown to 80\u201390% confluence. The cells were then transfected with control siRNA , PKA\u03b1 siRNA  or p38 MAPK\u03b1 siRNA  oligonucleotide duplexes  using Lipofectamine 2000  according to the manufacturer\u2019s instructions. Transfection efficiency was determined using qRT-PCR and western blotting.t-test or two-way ANOVA  were used to distinguish the statistical significance levels, where *, **, ***, and ns indicates p < 0.05, < 0.01 and < 0.001, and no significance, respectively.All values are expressed as the average \u00b1 standard error mean (SEM). The experimental determinants were confirmed by using at least triplicate biological samples. Student\u2019s B. bifidum DS0908 (DS0908) and B. longum DS0950 (DS0950) promote brown adipocyte-, beige adipocyte-, and lipolysis-related marker expressions in 3T3-L1 pre-adipocytes. Treatment with culture supernatants of DS0908 and DS0950 reduced intracellular triglyceride (TG) accumulation during thermogenesis induction, confirmed by ORO staining and TEM analysis [Ucp1 (5.60- and 3.38-fold), Pgc1\u03b1 (12.85- and 7.02-fold) and Prdm16 (4.56- and 3.59-fold) mRNA expressions and UCP1 (2.41- and 2.32-fold), PPAR\u03b3 (3.12- and 5.93-fold), PGC1\u03b1 (12.06- and 1.98-fold) and PRDM16 (0.70- and 0.31-fold) protein expressions in C3H10T1/2 MSCs (Fgf21 (8.94- and 4.94-fold) and P2rx5 (6.96- and 10.80-fold), as well as the brown adipocyte-specific marker Cox2 (9.76- and 7.10-fold) in C3H10T1/2 MSCs. However, we observed no significant change in the Cd137 and Tbx1 mRNA expression levels , Psat1 (1.51- and 1.36-fold), Resistin (1.43- and 1.00-fold) and Serpina3k (0.59- and 0.51-fold) in DS0908 and DS0950-treated C3H10T1/2 MSCs (Ucp1 (137.37- and 124.96-fold), Prdm16 (25.60- and 267.22-fold) and Ppar\u03b3 (0.07- and 16.08-fold), as well as the beige adipocyte-specific marker Cd137 (0.85- and 5.90-fold) mRNA expressions  into seven groups as follows: NFD; (G1), HFD; (G2), HFD + BS (cell-free supernatant) of DS0908 (G3), HFD + BS (cell-free supernatant) of DS0950 (G4), HFD + BP  of DS0908 (G5), HFD + BP  of DS0950 (G6), and HFD + Rosiglitazone (G7). The HFD groups were fed a 45% HFD for four weeks to induce obesity in the animals  noticeably reduced insulin levels, suggesting improved insulin sensitivity . FollowiS133 phosphorylation , and acetyl-CoA carboxylase activity , 37, 38.ogenesis , 44. Theogenesis . We alsoB. infantis and its culture supernatants  can induce thermogenesis by promoting white adipocyte browning to beige and by improving brown adipocyte activity [EBF2 and FGF21 deletion blunt beige adipocyte formation, whereas their induction triggers thermogenesis in beige adipocytes [B. bifidum DS0908 and B. longum DS0950 increased mitochondrial UCP1 and OXPHOS protein expressions. In addition, mitochondrial biogenesis increases mitochondrial content, which is known to contribute to thermogenesis. PGC1\u03b1 reportedly improves mitochondrial biogenesis and oxidative phosphorylation, induced upon B. bifidum DS0908 and B. longum DS0950 supplementation [Bifidobacterium strains considered in the current study increased the PRDM16 level.Other approaches for ameliorating obesity include non-shivering thermogenesis and lipolysis activation in mature white adipocytes. Several studies demonstrated that activity , 9. Whitipocytes , 46. PPAipocytes , 21. Botipocytes , 29. Freipocytes . In our entation , 28. PRDentation . The BifaP2), Psat1, Resistin and Serpina3k; beige and brown adipocytes, by Cd137, Tbx1 and Fgf21, as well as Cox2 and P2rx5, expressions, respectively [B. bifidum DS0908 and B. longum DS0950 supplementation [e.g., AMPK, PKA, p38 MAPK and CREB) phosphorylation, further inducing downstream thermogenic (common beige and brown) protein  expressions [B. infantis, can inhibit adiposity via AMPK and PKA/p38 MAPK signaling activation [B. longum positively regulated thermogenesis via PKA signaling activation [e.g., PKA, P38 MAPK and AMPK) [B. bifidum DS0908 and B. longum DS0950 activated PKA/p38 MAPK signaling in C3H10T1/2 MSCs during the promotion of thermogenesis. It is worth mentioning that multiple studies have investigated crude culture supernatants of probiotics as viable components to understand their role in alleviating obesity [B. bifidum DS0908 and B. longum DS0950 pellets and culture supernatants both in vitro and in vivo, these findings could be firmly bolstered by identifying key metabolites and investigating their anti-obesity effects via browning activation.Mature white adipocytes are characterized by adipocyte-binding protein 4  -51. In o obesity . AlthougB. bifidum DS0908 and B. longum DS0950 pellets and culture supernatants ameliorate obesity by activating the thermogenic program. In vivo study revealed that they improved the plasma lipid profile, insulin sensitivity, and glucose metabolism without altering food intake, suggesting that B. bifidum DS0908 and B. longum DS0950 and their culture supernatants might mitigate the clinical onset of obesity.In summary, our study provides in vitro and in vivo evidence that http://jmb.or.kr.Supplementary data for this paper are available on-line only at"}
+{"text": "Retraction Note:J Exp Clin Cancer Res38, 223 (2019)https://doi.org/10.1186/s13046-019-1210-3The Editor-in-Chief has retracted this article. After publication concerns were raised regarding irregularities present in multiple figures in this article. There is overlap in Fig. 2A with Fig. 2A of a previously published article . Figure"}
+{"text": "Klebsiella quasipneumoniae subsp. similipneumoniae strain IF3SW-P1, isolated from the International Space Station, was sequenced using Oxford Nanopore Technologies. The genome lacks a megaplasmid typical of hypervirulent and multidrug-resistant Klebsiella strains but does contain a chromosomally encoded OqxAB efflux pump associated with carbapenem resistance.The 5.2-Mb circular genome of  Klebsiella pneumoniae were described as the novel species Klebsiella quasipneumoniae , carbapenem-resistant, and hypermucoviscous strains isolated from both hospital-borne and community-acquired infections  on the ISS on 4 March 2015  using a \u2013nano-hq and \u2013read-error 0.03 for the Q20+ data (Escherichia coli strain Q4552 plasmid pECQ4552_IHU08 (GenBank accession number CP077071.1) (blaKPC and Inc(FII), which are known to occur in Klebsiella species (Oxford Nanopore Technologies sequencing was performed using a GridION MK1 sequencer on a R10.4 flow cell (FLO-MIN112) with a library synthesized from Q20+ EA (early access) ligation reagents (SQK-LSK112-XL). The raw reads were base called using MinKNOW v29.10.8, with a mean quality score of 16.3 and a mode of 18.03. The genome was assembled, circularized, and polished using Flye v2.9 with the parameters 20+ data . The Fly77071.1) . Notably species .K. quasipneumoniae subsp. similipneumoniae by calculating the average nucleotide identity (ANI) using the EzBioCloud calculator compared to the two subspecies\u2019 type strains, K. quasipneumoniae subsp. quasipneumoniae 01A030T  and K. quasipneumoniae subsp. similipneumoniae 07A044T   . Strain 0T ANI, 9.63% and iutA, which encodes a ferric aerobactin receptor, although the gene encoding the associated siderophore aerobactin (iucA) is not present  and the Sequence Read Archive (SRR17974437). These data are also available at NASA GeneLab (GLDS-470).The genomic assembly and raw reads have been deposited at GenBank (accession number"}
+{"text": "The COVID-19 pandemic and lockdowns may adversely affect pregnancy outcomes due to disrupted healthcare provision and increased stress, anxiety and economic hardship. We assessed changes in perinatal outcomes in 2020 using population birth data in Europe.25 Countries in the Euro-Peristat Network implemented a federated analysis using routine national data. Countries generated anonymised aggregate data files using R scripts from individual-level data formatted to a common data model with 22 variables. We compared preterm birth, stillbirth, neonatal death and caesarean delivery rates in 2020 to 2015-2019 for 2 periods: full-year (FY) and pandemic (March-September [MS]). Data from October onward were not included in the MS period because potentially declining pandemic-related fertility may affect perinatal indicators. Country-specific relative risks (RR) for the periods, adjusted for linear trends, overall and by socio-economic (SES) group, were calculated and pooled using random effects meta-analysis.Preterm birth rates decreased slightly  0.95-0.99]; 0.98MS [0.96-1.00]) in 2020. Heterogeneity was high , with 5 countries experiencing significant declines. Neonatal mortality rates were unchanged (0.97FY [0.92-1.01]) while stillbirth rates were higher . Caesarean rates were slightly raised . Increases for stillbirth were more pronounced in the lowest (1.08FY [0.99-1.16]) versus highest SES group (1.05 FY [0.93-1.17]).In 2020, there was an unexpected decline in preterm birth in some countries, while increases in stillbirths and caesarean occurred in others. High country-level heterogeneity suggests that some government policies to mitigate the pandemic might have been more protective of pregnant women and newborns than others."}
+{"text": "E. coli isolates from suckling piglets with colibacillosis.Colibacillosis is a frequent enteric disease in the pig industry that causes significant economic losses. The objective of this study was to investigate the molecular characteristics of fluoroquinolone (FQ)-resistant E. coli isolates were tested in this study and all isolates showed multi-drug resistance (MDR) and mutations in quinolone resistance determining regions (gyrA or parC). Especially, FQ-resistant E. coli isolates with double mutations in both gyrA and parC were shown a high FQs minimum inhibitory concentration . Among 43 FQ-resistant E. coli isolates, 12 (27.9%) were showed plasmid-mediated quinolone resistance (PMQR) positive E. coli. Prevalence of PMQR gene, aac(6\u2019)-Ib-cr, qnrS, and qepA, were identified in 7, 3, and 2 E. coli isolates, respectively. We identified the following in PMQR-positive E. coli isolates: the tetracycline resistance genes tetD , tetE , tetA , and tetB ; \u03b2-lactamases\u2013encoding blaCMY-2 , blaTEM-1 , blaOXA-1 , blaSHV-1 , and blaAAC-2 ; and the chloramphenicol resistance genes ; the sulfonamide resistance genes sul1  and sul2 ; the aminoglycoside modifying enzyme gene aac(3)-II . The F4 , LT:STb:EAST1 , and paa  were most common fimbrial antigen, combinations of toxin genes, and non-fimbrial adhesins genes, respectively. All PMQR-positive E. coli carried class I integrons but only 4 isolates carried the gene cassette. The most prevalent plasmid replicon was FIB , followed by FIC, HI1, and N , respectively.A total of 43 FQ-resistant E. coli can serve as a reservoir of FQ resistant genetic determinants that can be transferred to pathogenic bacteria in humans or pigs, this represents a public health hazard.Because FQ-resistant The online version contains supplementary material available at 10.1186/s12866-022-02632-9. Escherichia coli (E. coli) in pigs is the most frequent enteric disease and an important cause of death in suckling piglets [Colibacillosis caused by  piglets . This di piglets . The useE. coli or other gram-negative bacteria. The World Health Organization (WHO) has classified FQs as \u2018\u2018critically important antimicrobials\u2019\u2019 because of their clinical importance in both human and animal medicine [E. coli have emerged in Korea after enrofloxacin was licensed to be used for veterinary purposes [FQs are broad-spectrum antimicrobials agents and have been used for the treatment of various infections caused by medicine . Becausepurposes .qnr-mediated inhibition of quinolone binding to DNA [qepA encoded efflux pump [aac(6\u2019)-Ib-cr mediated FQ acetylation [E. coli isolated from suckling piglets with diarrhea. Therefore, the purpose of this study was to investigate the molecular characteristics of FQ-resistant E. coli isolates from suckling piglets with colibacillosis.Multiple mechanisms are involved in resistance to FQ in Enterobacteriaceae. The major mechanisms of resistance to FQ involve mutations of chromosomal genes encoding DNA gyrase and/or topoisomerase IV , 6. In ag to DNA , 8, qepAlux pump , and thetylation , 11. Thetylation . Althougtylation \u201315, therE. coli isolates collected from each colibacillosis clinical case in suckling piglets from 2007\u20132018 were tested. The farms consisted of 42 different pig herds (50 to 100 sows per each herd). The aseptically collected intestinal contents and feces were inoculated on MacConkey agar containing 4\u00a0mg/mL of ciprofloxacin . Subsequently, suspected E. coli colonies were identified by VITEK II system . Thus, a total of 43 CIP-resistant E. coli were tested in this study.E. coli ATCC 25,922 was included as a quality control. Multidrug-resistance (MDR) was defined as acquired resistance to at least one agent in three or more antimicrobial classes [The disk diffusion method was performed according to the Clinical and Laboratory Standards Institute (CLSI) guidelines . The 19  classes .E. coli Reference Laboratory, Quebec, Canada).O-serogroup typing was performed using rabbit antisera purchased from SSI  with the slide agglutination technique of the Animal and Plant Quarantine Agency . A standard strain was obtained from Dr. J.M. Fairbrother  to identify mutations in 43 FQ-resistant E. coli isolates using primers and conditions described previously [http://www.ncbi.nlm.nih.gov/BLAST). PMQR genes -Ib-cr, and qepA) were detected by PCR amplification and sequencing analysis, as described in Table SPCR was carried out to amplify quinolone resistance determining regions (QRDRs) of the target genes -II, and ant(2\u2019\u2019)-I), \u03b2-lactam antimicrobials , chloramphenicols (cmlA and catA1), sulfonamides (sul1 and sul2), and tetracyclines . The virulence factor genes associated with the toxins , fimbriae , and non-fimbrial adhesins  were also confirmed by PCR as previously described [For detection of antimicrobial resistance genes and virulence genes, PCR was performed using DNA extracted from PMQR-positive escribed .E. coli isolates. The primers used in this study targeted 18 different replicons [For plasmid replicon typing and detection of integrons and gene cassettes, PCR was performed using DNA extracted from PMQR-positive eplicons  and clasE. coli J53 used as the recipient as previously described [To determine the transferability of PMQR and other genes, conjugation assays were performed using the broth mating method, with escribed . TranscoE. coli isolates from colibacillosis are shown in Table E. coli, all isolates showed MDR against 6 to 11 classes of antimicrobial agents. The rates of resistance to the various antimicrobial classes were as follows: aminoglycosides , penicillins , quinolones , tetracyclines , phenicols , \u03b2-lactam/\u03b2-lactamase inhibitor combinations , folate pathway inhibitors , cephems , monobactams , and polypeptides . The rate of resistance to 9 antimicrobial classes was the highest at 39.5%, and one (2.3%) FQ-resistant E. coli isolate was identified resistance to 11 antimicrobial classes.MDR patterns of FQ-resistant E. coli isolates are shown in Table gyrA amino acid substitutions were S83L (43 isolates), D87N (22 isolates), D87G (5 isolates), and D87E (2 isolates), and the parC mutations were S80I (23 isolates), S80R (9 isolates), E84A (5 isolates), S80K (3 isolates), S80N (1 isolates), S80W (1 isolates), E84G (1 isolates), A56C (1 isolates), and S57Q (1 isolates). The MIC ranges for CIP, ENR, and NOR were 4\u2013256\u00a0mg/mL, 8\u2013512\u00a0mg/ml, and 8\u2013512\u00a0mg/mL, respectively, and isolates with double mutations in gyrA were relatively higher than those of other isolates with single mutations in gyrA. In particular, FQ-resistant E. coli isolates with a high level of MICs range  were shown to carry double mutations in gyrA in combination with double mutations in parC. PMQR genes were detected in 12 (27.9%) of the 43 FQ-resistant E. coli isolates. The aac(6\u2019)-Ib-cr, qnrS, and qepA genes were identified in seven, three, and two FQ-resistant E. coli isolates, respectively. Among 12 PMQR-positive E. coli isolates, one isolate that showed the highest MICs for CIP (256\u00a0mg/mL), ENR (512\u00a0mg/mL), and NOR (512\u00a0mg/mL), also carried the PMQR gene aac(6\u2019)-Ib-cr.The molecular characteristics of 43 FQ-resistant E. coli isolates carried the following \u03b2-lactamase encoding genes: blaCMY-2 , blaTEM-1 , blaOXA-1 , blaSHV-1 , and blaAAC-2 . Tetracycline-resistance genes were detected in all PMQR-positive E. coli isolates as follows: tetD , tetE , tetA , and tetB . Two types of aminoglycoside-modifying enzyme genes were examined, but aac(3)-II was found only in 2 (16.7%) PMQR-positive E. coli isolates. The sul1 and sul2 sulfonamide-resistance genes were detected in 9 isolates (75.0%) and 10 isolates (83.3%), respectively. The cmlA chloramphenicol-resistance gene was found in 10 isolates (83.3%). Distributions of the virotypes are shown in Table E. coli isolates, all isolates were found to have class 1 integrons. Class 1 integrons contained four types of gene cassette arrangements, aadA1-dfrA1 (2 isolates), aadA1-aadA2- aadB (1 isolate), and aadA1-aadA2- aadB-cmlA6 (1 isolate). Eight isolates did not carry any of the gene cassettes. A total of 10 plasmid replicon types were identified in all 12 PMQR-positive E. coli isolates. The most common plasmid replicon was FIB , followed by FIC, HI1, and N , respectively. Transferability was only identified in 7 isolates among 12 PMQR-positive FQ-resistant E. coli isolates.The prevalence of antimicrobial resistance genes is shown in Table E. coli worldwide [E. coli isolates showed were MDR  with high levels of resistance to several antimicrobials: aminoglycosides (100.0%), penicillins (100.0%), tetracyclines (97.7%), and phenicols (90.7%). In particular, five isolates showed resistance to more than 10 classes. These results were consistent with previous studies showing co-association of resistance to other classes of antimicrobials and high MDR rates among FQ-resistant E. coli [Suckling piglets are vulnerable to colibacillosis for many reasons such as changes in environmental conditions, a decline in maternal antibody titers, and various stresses. Antimicrobials are used in intensive pig production systems to control infectious diseases. In particular, FQs are highly effective antimicrobial class with many advantages including high oral absorption, large volume of distribution, and broad-spectrum antimicrobial activity . Howeverorldwide ; thus, torldwide . In this E. coli , 27. Thi E. coli .E. coli and variety of O-serogroups has been associated with diarrhea [gyrA mutations, and 29 isolates (67.4%) were double amino acid substitutions (S83L plus substitution in aspartic acid 87). These results were consistent with previous studies showing that DNA gyrase is the primary target of FQ in gram-negative bacteria, and gyrA mutations are dominant mutations in E. coli [E. coli isolates had mutations at codon 80 in parC in the QRDRs, and the most common type of amino-acid substitution was S80I in parC. Previous studies reported that the substitution S80I was most frequently observed among substitutions in the QRDR of parC [E. coli isolates from humans [gyrA were relatively higher than those of other isolates with single mutations in gyrA. Notably, FQ-resistant E. coli isolates carrying double mutations in gyrA in combination with double mutations in parC were identified at high levels of MICs . These results are consistent with those of recent studies showing that the total number of point mutations in QRDR was positively correlated with the increased MIC.The O-serogroup is considered one of the major virulence factors of diarrhea \u201330. In tdiarrhea , 32. In  E. coli . Moreove of parC , 35. Them humans , 37. AlsE. coli isolates carried three types of PMQR genes, aac(6\u2019)-Ib-cr (7 isolates), qnrS (3 isolates), and qepA (2 isolates). These PMQR variants have been previously detected in E. coli from livestock, including in healthy animals and retail meats in the Czech Republic [gyrA and harbored qnrS in its plasmid. This result showed that PMQR genes play a role in FQ resistance [In this study, 12 FQ-resistant Republic , China [Republic , and theRepublic , as wellRepublic . Also, tRepublic . Howeversistance .E. coli isolates carried a variety of antimicrobial resistance genes such as blaCMY, blaTEM, blaOXA, blaSHV, blaAAC, tetA, tetB, tetD, tetE, aac(3)-II, sul1, sul2, and cmlA and harbored mobile elements such as integrons and gene cassettes at the same time. The bla genes hydrolyze the characteristic \u03b2-lactam ring and confer resistance to most \u03b2-lactam antimicrobials, including cephalosporins [bla positive-E. coli were detected at high levels [E. coli isolates harbored class 1 integrons and four isolates also contained gene cassettes aadA or dfrA or both genes. These genes are frequently detected in class 1 integrons isolated from animals and humans in Korea [E. coli isolates from suckling piglets may have acquired the genetic determinants of drug resistance, which could become a concern.The rise of antimicrobial resistance is thought to be closely associated with the widespread transfer of resistance genes between bacterial species. In this study, all 12 PMQR-positive osporins . Previouh levels . The preh levels . Also, ain Korea . TherefoE. coli isolates. The most common plasmid replicon was IncF plasmids including FIB, and FIC. IncF plasmids were associated with the important role in the spread of virulence and resistance to important classes of antimicrobials including quinolones, \u03b2-lactams, TEs, sulfonamides, chloramphenicol, and aminoglycosides among Enterobacteriaceae [Plasmids are small DNA molecules that are distinct from chromosomes and can provide beneficial effects to bacteria such as antibiotic resistance through horizontal gene transfer . In thiseriaceae .E. coli virulence factors is important for diagnosing and establishing preventative measures for colibacillosis [E. coli isolates, respectively. These LT, STa, and STb genes damage the vessel and cause edema and a high mortality in pigs [E. coli isolates were identified as having the paa gene and coexisting with the F4 gene. Although the specific role of the paa gene in the development of pathogenic E. coli has not yet been clearly defined, various virotypes may also appear as a result of horizontal gene transferability of the paa gene [The detection of cillosis . In this in pigs . Also, t in pigs \u201352. Prev in pigs , 53. In paa gene .E. coli isolated from suckling piglets with colibacillosis. All FQ-resistant E. coli isolates showed an MDR phenotype, and the most prevalent of the mutations were double point mutations in gyrA and a single mutation in parC. Also, FQ-resistant E. coli isolates with PMQR genes carried various antimicrobial genes and harbored mobile elements and plasmid replicons. Antimicrobial resistance may become a serious problem because many drugs are probably ineffective for the treatment of colibacillosis and resistance elements can be horizontally transferred on pig farms. Also, this represents a public health hazard because FQ-resistant E. coli can serve as a reservoir of FQ resistant genetic determinants that can be transferred to pathogenic bacteria in humans or pigs. These data support the critical need for comprehensive surveillance of antimicrobial resistance on pig farms.This study investigated the molecular characteristics of FQ-resistant Additional file 1:Figure S1-1. LC716475 (SSC-1) blast alignment. Figure S1-2. LC716476 (SSC-2) blast alignment. Figure S1-3. LC716477 (SSC-3) blast alignment. Figure S1-4. LC716478 (SSC-4) blast alignment. Figure S1-5. LC716479 (SSC-7) blast alignment. Figure S1-6. LC716480 (SSC-8) blast alignment. Figure S1-7. LC716481 (SSC-10) blast alignment. Figure S1-8. LC716482 (SSC-11) blast alignment. Figure S1-9. LC716483 (SSC-12) blast alignment. Figure S1-10. LC716484 (SSC-13) blast alignment. Figure S1-11. LC716485 (SSC-14) blast alignment. Figure S1-12. LC716486 (SSC-15) blast alignment. Figure S1-13. LC716487 (SSC-16) blast alignment. Figure S1-14. LC716488 (SSC-17) blast alignment. Figure S1-15. LC716489 (SSC-19) blast alignment. Figure S1-16. LC716490 (SSC-20) blast alignment. Figure S1-17. LC716491 (SSC-21) blast alignment. Figure S1-18. LC716492 (SSC-22) blast alignment. Figure S1-19. LC716493 (SSC-23) blast alignment. Figure S1-20. LC716494 (SSC-26) blast alignment. Figure S1-21. LC716495 (SSC-27) blast alignment. Figure S1-22. LC716496 (SSC-28) blast alignment. Figure S1-23. LC716497 (SSC-29) blast alignment. Figure S1-24. LC716498 (SSC-30) blast alignment. Figure S1-25. LC716499 (SSC-31) blast alignment. Figure S1-26. LC716500 (SSC-32) blast alignment. Figure S1-27. LC716501 (SSC-33) blast alignment. Figure S1-28. LC716502 (SSC-34) blast alignment. Figure S1-29. LC716503 (SSC-35) blast alignment. Figure S1-30. LC716504 (SSC-36) blast alignment. Figure S1-31. LC716505 (SSC-37) blast alignment. Figure S1-32. LC716506 (SSC-38) blast alignment. Figure S1-33. LC716507 (SSC-39) blast alignment. Figure S1-34. LC716508 (SSC-40) blast alignment. Figure S1-35. LC716509 (SSC-41) blast alignment. Figure S1-36. LC716510 (SSC-42) blast alignment. Figure S1-37. LC716511 (SSC-43) blast alignment. Figure S1-38. LC716512 (SSC-44) blast alignment. Figure S1-39. LC716513 (SSC-45) blast alignment. Figure S1-40. LC716514 (SSC-46) blast alignment. Figure S1-41. LC716515 (SSC-47) blast alignment. Figure S1-42. LC716516 (SSC-48) blast alignment. Figure S1-43. LC716517 (SSC-49) blast alignment. Additional file 2:Table S1. Primers used for PCR and DNA sequencing.Additional file 3:Table S2. History of samples and isolates separated from each sample."}
+{"text": "To date, 10 mcr variants, mcr-1 to mcr-10, have been described. mcr-8 was first identified in a carbapenem-resistant Klebsiella pneumoniae isolate  pathogens infections . Liu et  isolate . Unlike eumoniae , 5. In rfections . CurrentE. cloacae clinical isolate SD21 was recovered from the sputum sample of a male patient suffering from chronic obstructive pulmonary disease at a tertiary hospital in Shandong, China. The isolate was initially identified using mass spectrometry and confirmed by next-generation sequencing. MICs of 15 antimicrobial agents were determined by the broth microdilution method. The MIC of tigecycline was interpreted according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. And the interpretation of MICs of the remaining antimicrobials was based on the Clinical and Laboratory Standards Institute (CLSI) guidelines. This isolate exhibited a multidrug-resistant phenotype, with resistance to polymyxin B (>8\u2009\u03bcg/mL), cefmetazole, ceftazidime, cefotaxime, cefepime, tigecycline, ciprofloxacin, amikacin, and aztreonam (Table\u00a0S1). Conjugation assays were performed using the Escherichia coli J53 as the recipient strain  and long-read nanopore MinION (CP093914) and four plasmids, namely pSD21_mcr8 (CP093916), pSD21_266kb (CP093915), pSD21_54kb (CP093917), and pSD21_4kb (CP093918). According to multilocus sequence typing, SD21 was assigned as ST1718. Notably, a mcr-8 variant exhibiting 100% identity to mcr-8.2 was identified in pSD21_mcr8 (100 852\u2009bp). The plasmid pSD21_mcr8, belonging to IncFIA(HI1)/IncFII(K), harbored mcr-8.2 within the genetic context IS903B-orf-ISRor7-orf-dgkA-baeS-copR-ISEcl1-orf-mcr-8.2-orf-ISKpn26  from K. pneumoniae , showing 99.91% identity and 100% coverage to p2019036D-mcr8-345kb. Besides, the lack of an intact tra region may be one of the aspects limiting the mobilization of pSD21_mcr8. However, the lack of tra operon did not prevent the mobilization of mcr-8-carrying plasmids between K. pneumoniae and E. cloacae. There could be other underlying mobilization mechanisms that mediate the dissemination of mcr-8-carrying plasmids in different species.To investigate the genetic structure of -ISKpn26 . The mcrblaCTX-M-55, blaTEM-1B), fosfomycin (fosA3), aminoglycosides (aac(3)-IId, aph(3\u2032)-Ia, aph(6)-Id, rmtB, aadA22), macrolides (mph[A]), quinolones (qnrS1), sulfonamides (sul3), trimethoprim (dfrA14), tetracyclines (tet[A]), rifamycin (ARR-2), amphenicols (floR), and lincosamides (lnu[F]). Therefore, this plasmid enabled SD21 to exhibit resistance to multiple antimicrobial agents. The pSD21_266kb shared 99.99% identity with p16-6773.1 (CP039861.1) from Salmonella sp. . The other two plasmids in SD21 did not harbor accessory resistance genes.In addition to pSD21_mcr8, SD21 was found to harbor another resistance plasmid, designated pSD21_266kb. The pSD21_266kb with IncHI2/IncHI2A replicon type contained genes encoding resistance to \u03b2-lactams (mcr-8 in E. cloacae, identification of an E. cloacae isolate with mcr-8.2 indicates the further dissemination of the mcr-8 variant in different species. Furthermore, the mcr-8.2-bearing plasmid coexisted with a multidrug-resistant plasmid, which increased the difficulty of clinical treatment. In conclusion, we first reported an MDR E. cloacae strain with mcr-8.2 isolated from a patient in China. We hypothesize that the mcr-8.2-located transferable genetic elements may be the genetic basis for the transmission of mcr-8.2 among different species. Further research on epidemiology and transmission mechanism should be conducted to better understand the potential dissemination of mcr-8.Given that no reports have described the existence of CP093914, CP093915, CP093916, CP093917, and CP093918 under the BioProject PRJNA816615.The complete sequences of SD21 were deposited in the NCBI database with accession numbers"}
+{"text": "Retraction Note:J Exp Clin Cancer Res37, 202 (2018)https://doi.org/10.1186/s13046-018-0875-3The Editor-in-Chief has retracted this article because data presented in Figs. 2, 3 and 4 were duplicated from previously published articles. Specifically, a set of staining images in Fig. 2I had previously been published as Fig. 3E in . The act"}
+{"text": "To investigate clinical characteristics, management, and prognosis of Epstein-Barr virus (EBV)-positive lymphoma-associated hemophagocytic syndrome (LAHS) patients in real-world practice.This was a retrospective, single-center cohort study. EBV-positive LAHS patients diagnosed from January 2010 to December 2021 in our center were enrolled. Clinical characteristics, treatment, overall response rate (ORR), and overall survival (OS) were investigated. Univariate and multivariate analysis of potential factors were conducted.P=0.033, P=0.000, and P=0.004, respectively). Combined treatment of anti-hemophagocytic lymphohistiocytosis (HLH) and anti-lymphoma treatment was conducted in 24 patients; anti-HLH treatment was conducted in 18 patients; anti-lymphoma treatment was conducted in three patients; glucocorticoid treatment was conducted in one patient. ORR was 47.8%, and the median OS was 61  days for overall patients. Patients who received anti-HLH treatment and turned to anti-lymphoma treatment early displayed higher ORR and OS than those of anti-HLH patients . Elevated alanine aminotransferase level was the independent risk factor of EBV-positive LAHS prognosis.Of the 51 patients, 44 were T/NK cell lymphoma; five were B cell lymphoma; two were Hodgkin lymphoma. EBV-positive T/NK cell LAHS patients were significantly younger and showed lower fibrinogen levels and C-reactive protein levels than EBV-positive B cell LAHS patients (Prognosis of EBV-positive LAHS patients was poor. Anti-lymphoma treatment should be initiated as soon as HLH was rapidly controlled. EBV maind PD-L2 , 5. The nd PD-L2 . Howeveragenesis , 8.Hemophagocytic syndrome, also named hemophagocytic lymphohistiocytosis (HLH), is a rare but lethal hyperinflammatory syndrome. Aberrant activation of lymphocytes, monocytes, and macrophages leads to substantial release of inflammatory cytokines. HLH typically manifests itself as fever, hepatosplenomegaly, and multiple organ dysfunction syndrome (MODS). HLH can be categorized into primary HLH and secondary HLH. Primary HLH is characterized by early onset and cytotoxicity-related gene mutations. Secondary HLH mainly attacks adult patients secondary to malignancy, infection, autoimmune disease, pregnancy, and other triggers, with the incidence of 0.125 per 100,000 per year . Lymphomin situ hybridization. Besides antiviral treatment, induction treatment was divided into anti-HLH treatment, anti-lymphoma treatment, and the combination of both. Anti-HLH treatment consisted of HLH-1994 regimen, HLH-2004 regimen, and GED regimen (gemcitabine-etoposide-dexamethasone). Anti-lymphoma treatment was chemotherapy depending on different subtypes of lymphoma. Treatment response was evaluated according to the following conditions. Complete response (CR) required disappearance of all HLH-related symptoms, and HLH-related lab findings returning to normal range, including ferritin, soluble CD25 (sCD25), complete blood count, triglyceride, alanine aminotransferase (ALT), etc. Consciousness should recover to normal if central nervous system was involved. Partial response (PR) required normal body temperature and improvement by over 25% in at least two symptoms or lab findings, including sCD25 decrease by at least one third, ferritin and triglyceride decrease by at least 25%, ALT decrease by at least 50% if formerly over 400U/L, and neutrophil increase by 100% without transfusion. If neutrophil counts were formerly less than 0.5\u00d7109/L, they should exceed 0.5\u00d7109/L after treatment. If neutrophil counts were formerly over 0.5\u00d7109/L but less than 2\u00d7109/L, they should exceed 2\u00d7109/L after treatment. No response (NR) was defined as being unable to reach CR or PR. Overall response rate (ORR) was calculated as (CR+PR)/total patients \u00d7 100%. OS was defined as the interval between HLH diagnosis and all-cause death. The patients were followed up until December, 2021. This study was conducted with the approval of the West China Hospital ethics committee, according to Declaration of Helsinki.Patients consecutively diagnosed as EBV-positive LAHS from January, 2010 to December, 2021 at West China Hospital, Sichuan University were enrolled , 17. EBVt test and U test, respectively. Enumeration data were evaluated with \u03c7\u00b2 test. Survival analysis was assessed with Kaplan-Meier methods, and compared with Log rank test. Univariate and multivariate analysis of prognostic predictors was conducted with Cox proportional hazard model. It was regarded as statistically significant with P<0.05.Statistical analysis was conducted with SPSS 23.0 and GraphPad Prism 7.0 software. Measurable data with normal distribution and skewed distribution were evaluated with Fifty-one EBV-positive LAHS patients were enrolled in this study, including 35 (68.6%) male patients and 16 (31.4%) female patients. Median age at HLH diagnosis was 37 (13-64) years. Three (5.9%) patients were newly diagnosed as lymphoma, while 48 (94.1%) patients were relapsed/refractory lymphoma cases. Clinical characteristics of these patients at baseline were listed in P=0.033). Fibrinogen level and C-reactive protein (CRP) level were significantly lower in T/NK cell LAHS patients than B cell LAHS patients . Plasma EBV DNA level tended to be higher in B cell LAHS patients than in T/NK cell LAHS patients, but the difference was not significant. There was no significant difference in sex, IPI score, ECOG score, clinical manifestations, complete blood count, liver and kidney function, triglyceride, ferritin, sCD25, hemophagocytosis phenomenon, and the occurrence order of lymphoma and HLH between EBV-positive T/NK cell LAHS patients and EBV-positive B cell LAHS patients . EBV-positive LAHS patients displayed significantly higher hemoglobin and triglyceride levels at baseline than EBV-negative LAHS patients . T/NK-cell lymphoma tended to account for a higher proportion of EBV-positive LAHS patients (86.3%) than EBV-negative LAHS patients (69.8%), but the difference was insignificant  were significantly younger at HLH diagnosis than EBV-negative LAHS patients  patients who only received anti-HLH treatment, the median weeks of induction treatment was 4 (1-8). The majority of  patients received HLH-1994 regimen. HLH-2004 regimen  and GED regimen  were conducted in three patients each. Three (6.5%) patients only received anti-lymphoma chemotherapy. One ENKL patient and one ANKL patient received GLIDE regimen, and one DLBCL patient received chidamide-RCHOP regimen. Median number of chemotherapy cycles was 3 (2-8). One (2.2%) patient only received glucocorticoid treatment.For patients who were at higher risk or failed to achieve CR with a poor control of EBV infection, consolidation treatment was conducted, which included PD-1 monoclonal antibody treatment, chidamide treatment, and autologous or allogenic stem cell transplantation. For patients whose biopsy revealed positive PD-L1 staining of tumor cells, especially HL patients and DLBCL patients, PD-1 monoclonal antibody treatment was conducted. For patients who displayed aberrant epigenetic alterations, especially PTCL patients and double expression DLBCL patients, chidamide treatment was conducted. For young, fit, and high-risk patients with sufficient stem cells collected who were willing to receive transplantation, autologous or allogenic stem cell transplantation was conducted. Overall, seven (15.2%) patients underwent consolidation therapy. Three (6.5%) patients  received PD-1 monoclonal antibody treatment, of whom the DLBCL patient also received autologous stem cell transplantation after PD-1 monoclonal antibody treatment. Two (4.3%) patients  received autologous stem cell transplantation. One (2.2%) ANKL patient received allogenic stem cell transplantation. Two (4.3%) AITL patients received chidamide treatment.P=0.103). There was no significant difference in ORR among different treatment groups  patients, and PR was achieved in 18 (39.1%) patients. Twenty-four (52.2%) patients remained NR. ORR was 47.8%. In the combined treatment group, CR was reached in two (8.3%) patients, and PR was reached in 12 (50%) patients. NR was present in 10 (41.7%) patients. ORR was 58.3%. For patients who first received anti-HLH treatment and turned to anti-lymphoma chemotherapy subsequently, CR was achieved in two (10.5%) patients, and PR was achieved in 10 (52.6%) patients. Seven (36.8%) patients remained NR. ORR was 63.2%. For those who first received anti-lymphoma chemotherapy and turned to anti-HLH treatment subsequently, PR was achieved in two (40%) patients, and three (60%) patients remained NR. ORR was 40%. In anti-HLH treatment group, six (33.3%) patients achieved PR and twelve (66.7%) patients remained NR. ORR was 33.3% for the total anti-HLH treatment group, the HLH-1994 regimen subgroup, the HLH-2004 regimen subgroup, and the GED regimen subgroup. In anti-lymphoma treatment group, CR was achieved in two (66.7%) patients while one (33.3%) patient remained NR. ORR was 66.7%. One patient only received glucocorticoid treatment but remained NR. Patients who first received anti-HLH treatment and turned to anti-lymphoma treatment subsequently tended to show a higher ORR than those who received anti-HLH treatment only, but the difference was not significant . In terms of treatment group, median plasma EBV-DNA of patients after treatment in combined treatment group and anti-HLH group was 272.5 copies/mL (range 0-64800 copies/mL), and 3130 copies/mL (range 0-33800 copies/mL), respectively. For the three patients in anti-lymphoma group, median plasma EBV-DNA of patients after treatment was 0 copies/mL (range 0-50 copies/mL). For the only patient in glucocorticoid group, his plasma EBV-DNA level decreased from 158000 copies/mL to 88800 copies/mL after treatment. There was no significant difference in plasma EBV-DNA level after treatment among treatment groups.At baseline, plasma EBV-DNA of all patients was positive . After induction treatment, plasma EBV-DNA decreased significantly in all patients but one, who was a ANKL patient only receiving anti-HLH treatment but remaining NR . Plasma EBV-DNA of ten patients after treatment was negative. Another three patients had a plasma EBV-DNA level lower than 50 copies/mL after treatment. There was no significant difference in plasma EBV-DNA after treatment between T/NK-cell LAHS  and B-cell LAHS patients  patients received combined treatment. Sixteen (40%) patients first underwent anti-HLH treatment of 3.5 (1-9) weeks, and then turned to anti-lymphoma chemotherapy. Median number of chemotherapy cycles was 1 (1-8). Another five (12.5%) patients first received anti-lymphoma chemotherapy and turned to HLH-1994 regimen subsequently. The median number of chemotherapy cycles was 1 (1-3). The median weeks of HLH-1994 treatment was 4 (2-4). In the 16 (40%) patients who only received anti-HLH treatment, the median weeks of induction treatment was 4 (1-8). Two (5%) patients only received anti-lymphoma chemotherapy. Median number of chemotherapy cycles was 2.5 (2-3). One (2.5%) patient only received glucocorticoid treatment.Of the 40 evaluable EBV-positive T/NK cell LAHS patients, CR was achieved in three (7.5%) patients, and PR was achieved in 14 (35%) patients. Twenty-three (57.5%) patients remained NR. ORR was 42.5%. In the combined treatment group, CR was reached in two (9.5%) patients, and PR was reached in nine (42.9%) patients. NR was present in 10 (47.6%) patients. ORR was 52.4%. For patients who first received anti-HLH treatment and turned to anti-lymphoma chemotherapy subsequently, CR was achieved in two (12.5%) patients, and PR was achieved in seven (43.75%) patients. Seven (43.75%) patients remained NR. ORR was 56.25%. For those who first received anti-lymphoma chemotherapy and turned to anti-HLH treatment subsequently, PR was achieved in two (40%) patients, and three (60%) patients remained NR. ORR was 40%. In anti-HLH treatment group, five (31.25%) patients achieved PR and 11 (68.75%) patients remained NR. ORR was 31.25%. In anti-lymphoma treatment group, CR was achieved in one (50%) patient while one (50%) patient remained NR. ORR was 50%. One patient only received glucocorticoid treatment but remained NR. There was no significant difference in ORR among different treatment groups.P=0.033, We also compared treatment and response of EBV-positive LAHS patients and EBV-negative LAHS patients in our department. Treatment choice for these patients was significantly different, with a higher proportion of EBV-positive LAHS patients (52.2%) receiving combined treatment than EBV-negative LAHS patients  tended to display worse OS compared to patients whose plasma EBV DNA was no more than 105 copies/mL , but the difference was not significant (P=0.159) (P=0.060) (P=0.041) (P=0.003) (P=0.003) (P=0.006)  days, the median OS of the total cohort was 61 (95% CI 47.9-74.1) days. The median OS of EBV-positive NK/T cell LAHS patients was 65 (95% CI 52.7-77.3) days, while that of EBV-positive B cell LAHS or EBV-positive HL LAHS was not evaluable due to limited sample size. Patients whose baseline plasma EBV DNA exceeded 10P=0.001, P=0.004, and P=0.009, respectively). ENKL or ANKL tended to associate with worse OS (P=0.069). There was no significant association between plasma EBV-DNA level after treatment and OS. There was no significant relationship between other baseline characteristics and OS. When combining baseline ALT level, combined treatment of first anti-HLH regimen and subsequent anti-lymphoma regimen, treatment response, and ENKL or ANKL subtype in multivariate analysis, elevated baseline ALT level was the independent risk factor of OS (P=0.025). Combined treatment of first anti-HLH regimen and subsequent anti-lymphoma regimen and achieving CR or PR were independent protective factors of OS   Table\u00a04.P=0.001, P=0.016, P=0.044, P=0.017, and P=0.013, respectively). When combining these factors in multivariate analysis, lower ALT level, combined treatment of first anti-HLH regimen and subsequent anti-lymphoma regimen, and achieving CR or PR were independent protective factors of OS .Regarding EBV-positive T/NK cell LAHS patients, univariate analysis revealed lower ALT level, lower IL6 level, higher PT level, combined treatment of first anti-HLH regimen and subsequent anti-lymphoma regimen, achieving CR or PR were significantly related to improved OS , and achieving CR or PR were significantly related to improved OS . When combining both factors in multivariate analysis, only achieving CR or PR remained significantly related to improved OS , serving as protective factor of prognosis.We also compared prognosis of EBV-positive LAHS patients and EBV-negative LAHS patients in our department. OS of EBV-positive LAHS patients was similar to that of EBV-negative LAHS patients , Achievement Transformation Project (No. CGZH21001), 1.3.5 Project for Disciplines of Excellence, West China Hospital, Sichuan University (No. ZYJC21007), Translational Research Grant of NCRCH (No. 2021WWB03), and China Postdoctoral Science Foundation (No. 2021M692310).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Irritable bowel syndrome (IBS) includes diarrhea-predominant (IBS-D) and constipation-predominant (IBS-C) subtypes. We combined breath testing and stool microbiome sequencing to identify potential microbial drivers of IBS subtypes.2), methane (CH4), and hydrogen sulfide (H2S) levels were measured by gas chromatography, and baseline stool microbiome composition was analyzed by 16S rRNA sequencing. Microbial metabolic pathways were analyzed using Kyoto Encyclopedia of Genes and Genomes collection databases.IBS-C and IBS-D subjects from 2 randomized controlled trials (NCT03763175 and NCT04557215) were included. Baseline breath carbon dioxide, hydrogen  of stool methanogens, predominantly Methanobrevibacter, as well as higher absolute abundance of Methanobrevibacter smithii in stool. IBS-D subjects had higher breath H2 that correlated with lower microbial diversity and higher breath H2S that correlated with higher RA of H2S-producing bacteria, including Fusobacterium and Desulfovibrio spp. The predominant H2 producers were different in these distinct microtypes, with higher RA of Ruminococcaceae and Christensenellaceae in IBS-C/CH4+ (which correlated with Methanobacteriaceae RA) and higher Enterobacteriaceae RA in IBS-D. Finally, microbial metabolic pathway analysis revealed enrichment of Kyoto Encyclopedia of Genes and Genomes modules associated with methanogenesis and biosynthesis of methanogenesis cofactor F420 in IBS-C/CH4+ subjects, whereas modules associated with H2S production, including sulfate reduction pathways, were enriched in IBS-D.IBS-C subjects had higher breath CHM. smithii and H2S producers such as Fusobacterium and Desulfovibrio spp, respectively.Our findings identify distinct gut microtypes linked to breath gas patterns in IBS-C and IBS-D subjects, driven by methanogens such as  Irritable bowel syndrome (IBS) is estimated to affect 1 in 10 people globally, with a prevalence of 11.8%\u201314.0% in North America . Based o2) on the breath test (2S), may also be involved in diarrheal conditions. H2S is a gasotransmitter  and is involved in numerous functions throughout the body, including inflammation and mucosal repair in the gastrointestinal (GI) tract (2S) have been linked to colorectal cancer and ulcerative colitis  tract . However colitis ,8, which colitis . More reth IBS-D ,11, alth2 generated by syntrophic bacterial species for the generation of methane (4) is directly linked to slowing of intestinal transit in an animal model and an increased motility index in methane-producing IBS subjects  . Methano methane . Interessubjects  and may subjects ,16.2, CH4, and H2S) breath testing and stool microbiome sequencing to identify potential microbial drivers of clinical phenotypes in IBS.These findings demonstrate that changes in the gut microbiome are not uniform in IBS as a whole. Rather, different microbial compositions (microtypes) may account for the differing phenotypes of IBS. Identifying these microtypes may more clearly define possible microbial pathomechanisms in IBS in general. In this study, we combine 3-gas . IBS-C was diagnosed based on Rome IV criteria  based on Rome IV criteria . Subject2), H2, CH4, and H2S . Interpretation of breath test results was based on the North American Consensus for breath testing  lactulose breath testing using a system that allows the measurement of carbon dioxide  and stored at room temperature before DNA extraction. Stool form was classified according to the Bristol Stool Form Scale  stool samples were self-collected, immediately refrigerated, and then transported to the laboratory. For the IBS-C trial, only CHrm Scale .DNA extraction was performed using the MagAttract PowerSoil DNA KF Kit (Qiagen) with some modifications as described previously . ExtractMethanobrevibacter smithii and Methanosphaera stadtmanae, in stool from IBS-C and IBS-D subjects were determined by quantitative polymerase chain reaction (qPCR) using primers and probes targeting the beta subunit of RNA polymerase (rpoB) gene of each species (M. smithii stock culture and from M. stadtmanae DSM 3091 from the Leibniz Institute Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ)  were extracted using the same protocol, and standard curves with ten-fold serial dilutions was prepared for use as qPCR standards.Levels of 2 methanogenic archaeal species,  species . Assays http://links.lww.com/AJG/C678).Details of 16S sequencing and analysis protocols are provided in the Supplementary Digital Content , SAS 9.4 , RStudio , and GraphPad Prism 9 . Graph construction was performed using GraphPad Prism 9.02 (GraphPad Software). Significance was set at P < 0.05.The descriptive analysis is presented as mean \u00b1 SD. Categorical variables were compared with \u03c7http://links.lww.com/AJG/C678). Among IBS-C subjects, 58 (47%) were CH4 negative (IBS-C/CH4\u2212) and 66 (53%) were CH4 positive (IBS-C/CH4+) and considered to have IMO (4 positive. IBS-C/CH4+ subjects were significantly older than IBS-D (P = 0.037) and IBS-C/CH4\u2212 (P = 0.029) subjects. No differences in sex distribution or body mass index were identified between groups .A total of 171 IBS subjects were included (47 with IBS-D and 124 with IBS-C). Subjects' demographics and clinical characteristics are shown in the Supplementary Digital Content  (P < 0.0001).Stool form for all baseline stool samples was assessed using the Bristol Stool Form Scale. IBS-D subjects had a significantly higher average score (4.7 \u00b1 1.27), indicating looser, more watery stools, compared with IBS-C/CH2 was higher in IBS-D compared with all IBS-C subjects . Within IBS-C, 35.09% of IBS-C/CH4\u2212 subjects were positive for H2 SIBO, compared with 25.76% of IBS-C/CH4+ subjects  for HFigure 2 , as weres Figure b. Signifhttp://links.lww.com/AJG/C678). Breath H2 dynamics were markedly different between IBS-D subjects and IBS-C subcategories . H2 delta values (120 minutes after lactulose ingestion vs preingestion levels) were significantly higher in IBS-D subjects vs IBS-C/CH4\u2212 (P = 0.038) and IBS-C/CH4+ . By 15 minutes after lactulose ingestion, H2 levels were already higher in IBS-D vs IBS-C/CH4+ subjects  and remained significantly higher at all time points during the breath test , 15 (P < 0.0001), 30 (P = 0.011), and 75 (P = 0.037) minutes . CH4 dynamics were also different between IBS-C and IBS-D subjects , with higher CH4 levels in IBS-C/CH4+ vs IBS-D subjects at all time points during the breath test .In IBS-D and IBS-C/CH4+ subjects and 40 IBS-D subjects provided baseline stool samples. Of IBS-C/CH4+ subjects, 88.09% had detectable levels of the methanogen M. smithii compared with 17.94% of IBS-D subjects (P < 0.0001). M. stadtmanae was less abundant in stool and was only detectable in 10% of IBS-C/CH4+ and 7.69% of IBS-D subjects (P = 1). Absolute M. smithii abundance correlated positively with breath CH4 levels, regardless of time point during the breath test, but a higher correlation coefficient was obtained using the maximum CH4 level reached during the breath test . Absolute M. smithii abundance also correlated negatively with breath H2 levels at 105 minutes  and 120 minutes  .A total of 42 IBS-C/CH4+ and 40 IBS-D subjects) were also used for 16S rRNA sequencing. After denoising and removal of low-quality reads, a total of 3,780,543 reads were retained for taxonomic analysis . Microbial alpha diversity analysis was performed using 3 different indices, Chao1, Simpson index, and Shannon index. Chao1 is an estimator based on abundance, Simpson index gives more weight to common or dominant species , while Shannon index assumes all species (including rare species) are represented in a sample    . Interestingly, higher breath CH4 AUC correlated with higher stool microbial alpha diversity  . This al4+ and IBS-D subjects were evident even at higher taxonomic levels. The relative abundance (RA) of the archaeal phylum Euryarchaeota was higher in the stool microbiome of IBS-C/CH4+ subjects compared with IBS-D . Regarding bacterial taxa, the RA of phylum Firmicutes was 1.27-fold higher in IBS-C/CH4+ vs IBS-D (FDR P = 0.04), and the RA of phyla Tenericutes, Lentisphaerae, and Synergistetes were also higher in IBS-C/CH4+ vs IBS-D  . By contrast, the RA of phyla Bacteroidetes , Fusobacteria , Proteobacteria , Epsilonbacteraeota , and Spirochetes  were higher in IBS-D subjects vs IBS-C/CH4+ .Differences in microbial profiles between IBS-C/CH4+ subjects was characterized by higher RA of methanogenic archaea from families Methanobacteriaceae  and Methanomassiliicoccaceae  when compared with IBS-D subjects. The RA of genus Methanobrevibacter was higher in IBS-C/CH4+ subjects vs IBS-D , confirming the qPCR results. Bacterial families with higher RA in IBS-C/CH4+ subjects vs IBS-D included Anaeroplasmataceae , Flavobacteriaceae , Christensenellaceae , Enterococcaceae , and Ruminococcaceae , among others . Notably, the RA of family Methanobacteriaceae correlated positively with RA of these bacterial families in IBS-C/CH4+ subjects, indicating possible syntropic relationships between these microbes , Spirochaetaceae , Fusobacteriaceae , and Bacteroidaceae  . Most of these bacterial families negatively affected stool microbial diversity  and of Spirochaetaceae correlated with high breath H2S AUC  and an unknown genus from family Desulfovibrionaceae  . There was no significant difference in RA of genus Desulfovibrio in IBS-D vs IBS-C/CH4+ subjects after P value correction , but the RA of several Desulfovibrio OTUs was higher in IBS-D subjects vs IBS-C/CH4+ .At the genus level, 35 known and unknown genera had higher RA in the stool microbiome of IBS-D subjects when compared to IBS-C/CH4+ and IBS-D subjects , microorganisms encoding enzymes necessary for H2S and CH4 production correlated with breath H2S and CH4 levels in IBS subjects. RA of Fusobacterium and an unknown Desulfovibrio species correlated positively with AUC for H2S . RA of genus Methanobrevibacter correlated positively with breath CH4 levels at all time points (P < 0.0001) and with AUC for CH4 , consistent with the findings for stool M. smithii levels by PCR. Moreover, Methanobrevibacter RA correlated negatively with H2 levels at 105 minutes  and 120 minutes .Although RA of numerous microbial taxa was different between IBS-C/CH4+ subjects and included enrichment of Kyoto Encyclopedia of Genes and Genomes (KEGG) modules associated with methane production from CO2, methanol, and methylamine . The KEGG module predicting biosynthesis of F420, a cofactor used during methanogenesis (4+ subjects (P < 0.0001), and this module correlated with breath CH4 levels at all time points  and with CH4 AUC .Microbial metabolic pathway analysis further supported these associations. A signature associated with biomethanation was characteristic of the stool microbiome in IBS-C/CHogenesis , was als2S production, including dissimilatory and assimilatory sulfate reduction pathways . Although there were no direct associations between these pathways and breath H2S levels, the assimilatory sulfate reduction pathway correlated with H2 levels .A biochemical signature associated with sulfur metabolism was characteristic of the stool microbiome in IBS-D subjects due to enrichment of KEGG modules associated with H4 breath tests, breath CH4 levels were linked to higher stool levels of the methanogenic archaeon M. smithii, confirmed by both qPCR and sequencing. IBS-C/CH4+ subjects had a distinct gut microtype when compared to IBS-D, characterized by higher RA of the archaeal family Methanobacteriaceae (which includes M. smithii) that correlated with higher RA of specific H2-producing bacterial families, Ruminococcaceae and Christensenellaceae, which include known syntrophs of M. smithii. By contrast, IBS-D subjects were characterized by elevated breath levels of H2 and H2S. Breath H2S levels correlated with RA of gut bacterial H2S producers in IBS-D subjects, including genus Fusobacterium and an unknown species from genus Desulfovibrio, and the RA of family Fusobacteriaceae correlated with the RA of the H2-producing family Enterobacteriaceae. In addition, predicted microbial metabolic pathway analysis indicated enrichment of KEGG modules associated with methane production in IBS-C/CH4+ subjects and enrichment of KEGG modules associated with H2S production in IBS-D subjects. These findings suggest that increases in M. smithii and in bacterial H2S producers including Fusobacterium and Desulfovibrio species may contribute to the predominant constipation and diarrheal subtypes in IBS subjects, respectively.In this study, we identify breath gas profiles and associated gut microtypes characteristic of different IBS phenotypes. Specifically, in IBS-C subjects with positive CH2 nor CH4 is produced by human cells  correlated with the presence of SIBO  and breath CH4 in IBS-C/CH4+ subjects. Interestingly, there was also an inverse relationship between CH4 and H2 levels, which may be consistent with the syntropic relationship between fermenting bacteria and hydrogenotrophic methanogens  and a constipation phenotype. By contrast, IBS-D subjects are characterized by higher breath H2 and by higher breath H2S linked to increased prevalence of H2S-producing bacteria (predominantly Fusobacterium and Desulfovibrio spp) and a diarrhea phenotype. Furthermore, the predominant H2 producers were different in these distinct microtypes, with higher RA of Ruminococcaceae and Christensenellaceae in IBS-C/CH4+ and higher Enterobacteriaceae RA in IBS-D. Identification of these distinct microtypes may facilitate a better understanding of the relationship between the gut microbiome and the heterogeneous phenotypes of IBS and allow us greater precision in the development of targeted microbiome-based therapies.In conclusion, our data identify distinct gut microtypes linked to breath gas patterns in subjects with IBS-C and IBS-D. IBS-C subjects are characterized by detectable breath CHGuarantor of the article: Mark Pimentel, MD.Specific author contributions: Conceptualization: M.P., R.M. Formal analysis: G.L., M.J.V.M., J.W., A.R., M.P. Methodology: M.J.V.M., G.L., W.M., S.W., M.P. Investigation: M.J.V.M., G.L., G.P., M.L.P., G.M.B., M.S., S.A., D.C., S.W., C.C., M.R., A.H., A.F., B.C., N.P., A.R., M.P. Visualization: M.J.V.M., G.L. Funding acquisition: G.B., R.M., M.P. Project administration: R.M., M.P. Supervision: W.M., S.W., C.C., M.R., R.M., M.P. Writing\u2013original draft: M.J.V.M., G.L., G.B., W.M., J.W., M.P., Writing\u2013review & editing: M.J.V.M., G.L., G.B., W.M., J.W., A.R., R.M., M.P.Financial support: This study was supported in part by funds from The Monica Lester Charitable Trust (R.M.), The Elias, Genevieve, and Georgianna Atol Charitable Trust (R.M.), Synthetic Biologics, Inc (A.R.), Bausch Health (M.P.) and The National Philanthropic Trust (M.P.).Potential competing interests: M.P. is a consultant for Bausch Health, Ferring Pharmaceuticals Inc., and Vivante Health Inc. M.P. has received grant support from Bausch Health and Synthetic Biologics. R.M. has received grant support from Valeant Pharmaceuticals. A.R. is a consultant/speaker for and has received grant support from Bausch Health. In addition, Cedars-Sinai Medical Center has licensing agreements with Bausch Health and Gemelli Biotech. A.R., M.P., and R.M. have equity in Gemelli Biotech and M.P. has equity in Synthetic Biologics. All other authors report no conflicts of interest.Data availability: The data sets generated during the current study are available in the National Center for Biotechnology Information (NCBI) BioProject Repository https://www.ncbi.nlm.nih.gov/bioproject under BioProject PRJNA804225.\u2713 Irritable bowel syndrome (IBS) includes diarrhea-predominant (IBS-D) and constipation-predominant (IBS-C) subtypes.\u2713 The gut microbiome is associated with IBS, but the roles of specific gut microbial populations are poorly understood.2), hydrogen sulfide (H2S), and methane (CH4) are produced by gut microbes.\u2713 The gases hydrogen (H4 on the breath test is associated with IBS-C and correlates with increased predominance of methanogens, including Methanobrevibacter smithii.\u2713 Increased CH2 levels do not directly correlate with diarrhea, but H2S has recently been linked to a diarrhea phenotype.\u2713 H\u2713 Distinct gut microtypes are linked to breath gas patterns in IBS-C and IBS-D subjects.4+ IBS-C subjects, increased breath CH4 correlated with increased gut microbial diversity.\u2713 In CH2 correlated with lower microbial diversity and increased breath H2S correlated with increased predominance of H2S producers, including Fusobacterium and Desulfovibrio species.\u2713 In IBS-D subjects, increased breath H2 producers in IBS-C subjects were Ruminococcaceae and Christensenellaceae, which include known bacterial syntrophs for methanogens.\u2713 The predominant H4+ subjects and enrichment of pathways associated with H2S production in IBS-D subjects.\u2713 Predicted microbial metabolic pathway analysis indicated enrichment of pathways associated with methanogenesis in IBS-C/CH"}
+{"text": "Escherichia coli strain to modify chrysazin to chrysazin-8-O-\u03b1-l-rhamnoside (CR) and chrysazin-8-O-\u03b1-l-2\u2032-O-methylrhamnoside (CRM) using rhamnosyl transferase and sugar-O-methyltransferase. Biosynthesized CR and CRM were structurally characterized using HPLC, high-resolution mass spectrometry, and various nuclear magnetic resonance analyses. Antimicrobial effects of chrysazin, CR, and CRM against 18 superbugs, including 14 Gram-positive and 4 Gram-negative pathogens, were investigated. CR and CRM exhibited antimicrobial activities against nine pathogens, including methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive Staphylococcus aureus (MSSA) in a disk diffusion assay at a concentration of 40 \u00b5g per disk. There were MIC and MBC values of 7.81\u201331.25 \u00b5g/mL for CR and CRM against methicillin-sensitive S. aureus CCARM 0205 (MSSA) for which the parent chrysazin is more than >1000 \u00b5g/mL. Furthermore, the anti-proliferative properties of chrysazin, CR, and CRM were assayed using AGS, Huh7, HL60, and HaCaT cell lines. CR and CRM showed higher antibacterial and anticancer properties than chrysazin.Anthraquinone and its derivatives show remarkable biological properties such as anticancer, antibacterial, antifungal, and antiviral activities. Hence, anthraquinones derivatives have been of prime interest in drug development. This study developed a recombinant  Quinones are naturally occurring organic compounds found in higher plants, fungi, bacteria, and animals. They have a lot of structural varieties. Since they are found in different colors in nature, they are considered pigments . Anthraq3/P2O3Cl4 [Synthesis of anthraquinone derivatives is of great interest recently. There are various methods for the synthesis of anthraquinones derivatives, including intramolecular condensation of aryl and o-aroylbenzoic acid using fuming sulfuric acid, benzoyl chloride, concentrated sulfuric acid, benzoyl chloride, zinc chloride, and POCl/P2O3Cl4 . The che/P2O3Cl4 .Modification of anthraquinones can be performed by glycosylation, methylation, sulfation, prenylation, and so on. Glycosylation is an important process for increasing the solubility of hydrophobic compounds, improving stability, reducing toxicity, and modifying biological activities ,16. MethO-methyltransferase (OMT) that catalyzes the transfer of the methyl group of S-adenosyl-l-methionine (SAM) to hydroxyl groups [Methyltransferases are important enzymes in the modification of different substrate. However, methylation of sugar is very rare ,21. Genel groups ,22,23.Rheum palmatum L. (Polygonaceae) [Chrysazin  has been used as a medicine since ancient times. It can be found naturally. It is isolated from the root and rhizome of onaceae) . It has onaceae) ,25.E. coli sugar (thymidine diphosphate (TDP)-rhamnose) as a sugar donor, which will be used as a glycosyltransferase to conjugate to chrysazin and SAM as an O-methyltransferase to conjugate chrysazin rhamnoside. For the production of chrysazin derivatives, anthraquinone rhamnosyltransferase (7665) [Saccharothrix espanaensis and O-methyltransferase (ThnM1) [Nocardia sp. CS682 were cloned and heterologously expressed in E. coli. The recombinant strain E. coli was utilized for the production of chrysazin-8-O-\u03b1-l-rhamnoside (CR) and chrysazin-8-O-\u03b1-l-2\u2032-O-methylrhamnoside (CRM) from the substrate chrysazin (This study is based on the utilization of indigenous e (7665)  from Sac (ThnM1)  from Nochrysazin . Antican\u03b2-d-1-thiogalactoside (IPTG) was purchased from GeneChem Inc. . SAM was purchased from Sigma-Aldrich . Escherichia coli BL21 (DE3)  was used as an expression and biotransformation host. Luria\u2013Bertani (LB) broth medium and agar plates with appropriate antibiotics  were used for culture preparation, colony selection, and biotransformation. Pathogenic strains such as Staphylococcus. aureus CCARM 3640 (MRSA), S. aureus CCARM 3089 (MRSA), S. aureus CCARM 33591(MRSA), S. aureus CCARM 0205 (MSSA), S. aureus CCARM 0204 (MSSA), S. aureus CCARM 0027 (MSSA), S. aureus CCARM 3090 (MRSA), S. aureus CCARM 3634 (MRSA), S. aureus CCARM 3635 (MRSA), Bacillus subtilis ATCC 6633, Enterococcus faecalis 19433, Enterococcus faecalis 19434, Kocuria rhizophilla NBRC 12708, Micrococcus luteus, Escherichia coli ATCC 25922, Proteus hauseri NBRC 3851, Klebsiella pneumonia ATCC10031, and Salmonella enterica ATCC 14,028 were obtained from Professor Seung-Young Kim  [Chrysazin/Dantron was purchased from Tokyo Chemical Industry . HPLC-grade acetonitrile and water were purchased from Mallinckrodt Baker . All other chemicals used were of high analytical grade and commercially available. Isopropyl-E. coli, anthraquinone rhamnosyltransferase (7665) [S. espanaensis and rhamnose methyltransferase (ThnM1) [Nocardia sp. CS682 were taken to prepare E. coli S2 [E. coli S2 was generated by transforming pET32a \u00b1 7665(Am) CRISPRi-S1(Cmr) and pCDFDuet-metK-thnM1(Sm) into an E. coli strain harboring the rhamnose cassette piBR181-tgs.dh.ep.kr.pgm2.glf.glk (Km) [For the production of rhamnosylated and methoxy-rhamnosylated derivatives of chrysazin using engineered e (7665)  from S.  (ThnM1)  from Noc coli S2 . E. coliglk (Km) . RecombiE. coli S2 was cultured in a 5 mL LB medium supplemented with ampicillin, kanamycin, chloramphenicol, and streptomycin (each at 50 \u03bcg/mL) and incubated at 37 \u00b0C for 3 h. From the 5 mL seed culture, 500 \u00b5L was transferred into a flask containing 50 mL LB broth with respective antibiotics and cultured at 37 \u00b0C for around 4 h until the optical density of cells at 600 nm (OD600) reached 0.6\u20130.8. This culture had added to it 0.5 mm isopropyl \u03b2-d-1-thiogalactopyranoside (IPTG) to induce protein expression, followed by incubation at 20 \u00b0C for 20 h. To determine optimal substrate concentration, glucose concentration, and time, different concentrations  of chrysazin and different concentrations  of sterile glucose solution were fed into the cell culture after induction. After 20 h of adding IPTG, chrysazin dissolved in dimethyl sulfoxide (DMSO) at a final concentration of 400 \u00b5M was added along with 10% glucose. Samples (3 mL of culture broths) were withdrawn at 6, 12, 24, 36, 48, and 60 h for product analysis. After 48 h, compounds were extracted using a double volume of ethyl acetate  and a Soxhlet extractor. The Soxhlet extractor was kept still to separate the mixture for 4\u20136 h at room temperature after shaking. Ethyl acetate was removed under reduced pressure and dissolved in methanol. This sample was further analyzed by HPLC and mass spectrometry. To collect a sample to characterize structurally, the biotransformation experiment was performed using a fermenter (3 L of culture). The pure fraction of the compound was collected via preparatory-high-pressure liquid chromatography (prep-HPLC).A seed culture of 18 column  ). The binary mobile phase was composed of solvent A (HPLC-grade water + 0.1% trifluoroacetic acid) and solvent B . The total flow rate was maintained at 1 mL/min for the 30 min program. The ACN concentration began with 10%. A linear gradient from 10 to 50% for 10 min, 50\u201390% for 23 min, and 90\u201310% for 30 min was then used. The HR-QTOF ESI/MS was performed in positive ion mode using an Acquity mass spectrometer , which was coupled with a Synapt G2-S system (Waters). Purification of compounds was performed using a prep-HLPC instrument equipped with a YMC-Pack ODS-AQ C18 column,   and a connected UV detector (420 nm). Here, a 40 min binary program with implementation of 20% (0\u20135 min), 50% (5\u201310 min), 70% (10\u201320 min), 90% (20\u201325 min), 20% (25\u201330 min), and 10% (30\u201335 min) ACN at a flow rate of 10 mL/min was used. Purified products were pooled, dried, and lyophilized to remove water or moisture. Furthermore, the fully dried pure compound was dissolved in DMSO-d6 and subjected to a 700 MHz NMR spectrometer equipped with TCI CryoProbe (5 mm). From the extracted compound, a 20 \u00b5L volume was injected and directly analyzed by reverse-phase high-performance liquid-chromatography photo-diode array (HPLC-PDA) using a Thermo Scientific Dionex Ultimate 3000 ultrahigh-performance Liquid chromatography (UHPLC) system with a reverse-phase C1H NMR and 13C NMR) and two-dimensional NMRs (heteronuclear multiple quantum coherence (HMQC), rotating frame Overhauser enhancement spectroscopy (ROESY), and heteronuclear multiple bonded connectivity (HMBC)) were used as needed to elucidate the structure of the compound.One-dimensional NMRs , incubated at 37 \u00b0C in a humidified 5% CO2 overnight, and then treated with each compound after serial dilution  for 72 h. After that, 20 \u03bcL substrate solution (Promega) was added to each well. The plate was shaken for 5 min and kept in the dark for 10 min. Luminescence was measured using a multimode plate reader . IC50 values were analyzed using GraphPad Prism 5 .Three cancer cell lines , human gastric cancer cell line (AGS), and human leukemia cell line (HL60) and normal cell line (human keratinocyte cell line (HaCaT) were purchased from Korean Cell Line Bank . Huh7 cells were cultured in Dulbecco\u2019s modified Eagle\u2019s medium (DMEM)  and AGS cells were cultured in Roswell Park Memorial Institute 1640 medium (RPMI1640)  supplemented with 10% fetal bovine serum (FBS)  and 1% penicillin-streptomycin-amphotericin B . Human leukemia HL60 cells were cultured in RPMI1640 supplemented with 10% FBS, 1% penicillin-streptomycin-amphotericin B, and L-glutamine (2 mm) . HaCaT cell lines were grown in DMEM supplemented with 10% FBS, 100 \u03bcg/mL streptomycin, and 100\u2009\u03bcg/mL benzylpenicillin. All cells were maintained at 37 \u00b0C in a humidified 5% COStaphylococcus. Aureus CCARM 3640 (MRSA), S. aureus CCARM 3089 (MRSA), S. aureus CCARM 33591(MRSA), S. aureus CCARM 0205 (MSSA), S. aureus CCARM 0204 (MSSA), S. aureus CCARM 0027 (MSSA), S. aureus CCARM 3090 (MRSA), S. aureus CCARM 3634 (MRSA), S. aureus CCARM 3635 (MRSA), Bacillus subtilis ATCC 6633, Enterococcus faecalis 19433, Enterococcus faecalis 19434, Kocuria rhizophilla NBRC 12708, and Micrococcus luteus) and four Gram-negative bacteria  were used to test antibacterial activities of chrysazin and its derivatives. The paper disk diffusion assay on the Mueller\u2013Hinton agar (MHA) plate was carried out according to Clinical Laboratory Standard Institute (CLSI) guidelines and the Kirby\u2013Bauer method [8 colony forming units (CFU)/mL were spread onto MHA plates. Then, 40 \u00b5g/disk compounds were placed on the surface of inoculated agar plates using sterile paper disks of 6 mm . Samples were then incubated at 37 \u00b0C for 18\u201320 h. The zone of inhibition diameter was measured in millimeters for each pathogen. Dimethyl sulfoxide (DMSO) was used as a control for the zone of inhibition as all compounds were dissolved in DMSO.Fourteen Gram-positive bacteria , S. aureus CCARM 3089 (MRSA), S. aureus CCARM 33591(MRSA), S. aureus CCARM 0205 (MSSA), S. aureus CCARM 0204 (MSSA), S. aureus CCARM 0027 (MSSA), S. aureus CCARM 3090 (MRSA), S. aureus CCARM 3634 (MRSA), and S. aureus CCARM 3635 (MRSA). They were grown in Mueller\u2013Hinton Broth (MHB) . The broth dilution method was used to determine MIC [8 CFU/mL) and then diluted to 2.5 \u00d7 106 CFU/mL in MHB. After knowing the MIC, the MBC test was performed on a fresh MHB medium by inoculating cultured samples containing MIC compounds and experimental strains.The following nine strains were used in the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) tests: E. coli strain S2 was generated by engineering E. coli BL21 (DE3), which contained a sugar transfer cassette and a sugar methylation cassette. It was cultured and prepared for biotransformation as mentioned in E. coli strain S2 was analyzed by HPLC. The HPLC chromatogram of chrysazin was obtained with its standard retention time (tR) of 21.3 min. Two new peaks of CR and CRM were obtained at tR of 14.9 min and 16.3 min, respectively, with UV absorbance at 420 nm  and 2D NMR , as shown in Biotransformation was carried out through fermentation to collect CR and CRM for structure identification and further biological activity tests. The biotransformation reaction mixture was extracted with a double volume of ethyl acetate. The crude extract was subjected to preparatory-high-pressure liquid chromatography (prep-HPLC) for purification. After several rounds of prep-HPLC, purified compounds were obtained. The purified product was dried by lyophilization, dissolved in 400 \u00b5L of deuterated dimethyl sulfoxide, and analyzed by nuclear magnetic resonance (NMR) spectroscopy (700 MHz) including 1D NMR  was consistent with \u03b4 5.67 , in which the anomeric proton coupling constant (J = 1.1 Hz) confirmed that the conjugation of rhamnose moiety had an \u03b1-configuration. In addition, with 13C-NMR of CR, the anomeric carbon peak appeared at \u03b4 99.08 ppm, with other peaks appearing between 70 and 80 ppm along with a CH3 peak at 18.35 ppm. In the case of CRM, there was a methylation in the 2\u2032 -OH group of rhamnose in CR, where the OCH3 spectrum was visible in both 1H and 13C NMR at 3.5 ppm and 59.44 ppm, respectively. Furthermore, to confirm the sugar and sugar-O-methylation conjugation, two-dimensional (2D)-NMR analyses such as 1H-13C HSQC, 1H-13C HMBC, 1H-1H COSY, and 1H-1H ROESY experiments were performed. Similarly, in CR, HSQC showed a cross peak illustrating a correlation between the anomeric C-1\u2032 proton (\u03b4 5.67 ppm) and the anomeric carbon (\u03b4 99.00 ppm). Moreover, the C-8 signal appearing at \u03b4 157.18 ppm showed a direct correlation with the observed anomeric proton at \u03b4 5.67 ppm in HMBC (\u03b4 3.71 ppm) and the carbon (\u03b4 80.61 ppm), and HMBC showed a cross peak depicting the correlations between C-2\u2032 (\u03b4 80.62 ppm) and the protons of the methoxy group (\u03b4 3.50 ppm) (O-\u03b1-l-rhamnoside) and methylated derivative of CR  produced by E. coli S2 whole-cell biotransformation were new compounds.The  in HMBC . In the .50 ppm) . Results50) values of chrysazin for AGS, Huh7, and HL60 cells were 17.08, 30.53, and 22.24 (\u03bcM), respectively. Chrysazin-8-O-\u03b1-l-rhamnoside inhibited AGS, Huh7, and HL60 cells with 50% inhibitory concentration (IC50) values of 28.58, 21.28, and 14.68 (\u03bcM), respectively. Here, CR showed better anticancer activities in Huh7 and HL60 than chrysazin. In the case of AGS cells, it showed slightly lower anticancer activity. In the case of chrysazin-8-O-\u03b1-l-2\u2032-O-methylrhamnoside, it showed higher anticancer activities than chrysazin and CR. The 50% inhibitory concentration (IC50) values of CRM for AGS, Huh7, and HL60 cells were 7.513, 4.467, and 4.540 (\u03bcM), respectively. In the case of the HaCaT normal cell line, the IC50 values were >200 \u03bcM for chrysazin, CR, and CRM -2,5- diphenyltetrazolium bromide (MTT) colorimetric assay against three different cancer cell lines  and one  and CRM . These cS. aureus subsp.  were prepared at a concentration of 10 mg/mL. All compounds were added to each disk at a final concentration of 40 \u00b5g/disk (4 \u00b5L). Each disk was placed over Mueller\u2013Hinton agar (MHA) plates spread with bacterial strains. The diameter of the zone of inhibition was measured after 18\u201320 h. Results of disk diffusion assays revealed that chrysazin did not show any antibacterial activity against 18 different human pathogens tested. However, CR and CRM exhibited antibacterial activities against Gram-positive bacteria s subsp. . These rO-\u03b1-l-rhamnoside, and chrysazin-8-O-\u03b1-l-2\u2032-O-methylrhamnoside against nine different pathogenic bacteria were determined. MIC values of compound chrysazin, CR, CRM, and erythromycin as a positive control, against nine strains of Gram-positive bacteria S. aureus subsp were determined by the broth dilution method. The assays were performed in 96-well plates in duplicate with Mueller\u2013Hinton broth. As summarized in S. aureus CCARM 0205 (MSSA), S. aureus CCARM 0204 (MRSA), S. aureus CCARM 3640 (MRSA), S. aureus CCARM 3090 (MRSA), S. aureus CCARM 3634 (MRSA), S. aureus CCARM 0027 (MSSA), S. aureus CCARM 3089 (MRSA), S. aureus CCARM 3635 (MRSA), and S. aureus CCARM 33591(MRSA), with MIC values of 7.81\u20131000 \u00b5g/mL. After knowing the MIC value, we further analyzed MBC, the lowest concentration of a compound for killing inoculated bacteria. An MBC test was performed by inoculating cultured samples containing MIC compounds and experimental strains on a fresh MHB medium (Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for chrysazin, chrysazin-8-B medium . MBC valE. coli strain for the sustainable production of different derivatives of chrysazin. Chrysazin-8-O-\u03b1-l-rhamnoside and chrysazin-8-O-\u03b1-l-2\u2032-O-methylrhamnoside were novel compounds. We also evaluated their activities against three different cancer cell lines . CR and CRM exhibited higher cytotoxicities than their parental compound, chrysazin. More significantly, the evaluation of antibacterial activity revealed promising bioactivities of CR and CRM. This study establishes an engineered microbial platform that can be used to produce novel bioactive compounds. This microbial platform can be further fine-tuned for the production of novel derivatives of other anthraquinones or different compounds. Furthermore, the optimization of bioprocessing parameters and rational engineering of the host using various synthetic biological tools and metabolic engineering can be employed to enhance the production titer.In this study, we successfully engineered an"}
+{"text": "STING is a direct downstream target of let-7i after brain injury. Furthermore, the intranasal delivery of let-7i agomir can also effectively inhibit STING and is beneficial for inflammation resolution and neuronal survival in a mouse model of pial vessel disruption stroke. Consequently, let-7i agomir is a promising candidate for clinical application as a chemically engineered oligonucleotides-based therapeutic for brain injury.Overcoming the lack of drugs for the treatment of traumatic brain injury (TBI) has long been a major challenge for the pharmaceutical industry. MiRNAs have emerged as potential targets for progress assessment and intervention against TBI. The brain-enriched miRNA let-7i has been proposed as an ideal candidate biomarker for TBI, but its regulatory roles in brain injury remain largely unknown. Here, we find that the expression of let-7i is significantly downregulated in the early stages of a hippocampal stab wound injury. The noninvasive intranasal administration of let-7i agomir significantly improves cognitive function and suppresses neuroinflammation, glial scar formation, and neuronal apoptosis in TBI mice. Mechanically,  Traumatic brain injury (TBI) is one of the leading causes of death and permanent disability worldwide and is a risk factor for developing dementia and neuropsychological diseases . AlthougMicroRNAs (miRNAs) are small single-stranded non-coding RNA molecules (about 22 nucleotides) that govern the processes of RNA silencing and the post-transcriptional regulation of gene expression. As a single miRNA regulates the expression of multiple target genes involved in many cellular processes, manipulating the expression of a single miRNA may affect the entire gene network, thus changing the phenotype of complex diseases. Mounting evidence suggests that miRNAs may serve as potential targets for progress assessment and intervention against TBI because they control a range of physiological and pathological functions, such as cell proliferation, differentiation, apoptosis, and metabolism ,9.Let-7 is a family of evolutionary conserved miRNA that are highly expressed in adult tissues and generally serve as key regulators of developmental processes . Let-7i Here, we aimed to examine whether TBI affects let-7 expression in the brain and to assess whether the restoration of let-7 levels is beneficial for suppressing neuroinflammation and reducing neuronal apoptosis after TBI. Furthermore, the mechanism of let-7 targeting for TBI was also explored.C57BL/6J mice were obtained from the SPF  Biotechnology company and maintained in groups of 3~5 animals on a 12 h light/dark cycle, and they had free access to water and a standard mouse diet. Two-month-old male mice were assigned to undergo TBI or sham surgery. All animal procedures followed the ethical guidelines for the care and use of experimental animals, and all experiments were approved by the Animal Committee of the Institute of Zoology, Chinese Academy of Sciences.Hippocampal stab injury (HSI) was performed as previously described with minor modifications . Two-monPVD strokes were induced as previously described . BrieflyIntranasal administration of agomir-let-7i was conducted as described previously . Agomir-IL-6 forward TACCACTTCACAAGTCGGA, IL-6 reverse AATTGCCATTGCACAACTC; IL-1\u03b2 forward CCTCAAAGGAAAGAATCTATACCTG, IL-1\u03b2 reverse CTTGGGATCCACACTCTCC; TNF\u03b1 forward TTCTCATTCCTGCTTGTGG, TNF\u03b1 reverse TTGGGAACTTCTCATCCCT; human STING forward CACATCCACTCCAGGTACC, human STING reverse AGAAATAGATGGACAGCAGCA; mouse STING forward CTCATTGTCTACCAAGAACCC, mouse STING reverse TTCTTCCTGACGAATGTGC; let-7a forward ggcgTGAGGTAGTAGGTTGTATA, let-7b forward ggcTGAGGTAGTAGGTTGTGTG, let-7c forward ggccTGAGGTAGTAGGTTGTATG, let-7e forward ggcTGAGGTAGGAGGTTGTATA, let-7f forward ggcggTGAGGTAGTAGATTGTATA, let-7g forward ggcgTGAGGTAGTAGTTTGTACA, let-7i forward ggcTGAGGTAGTAGTTTGTGCT, common miRNA reverse GCAGGGTCCGAGGTATTC.Total RNA was isolated with TRIzol reagent  according to the manufacturer\u2019s instructions. RNA quality was assessed with the Thermo NanoDrop 2000 spectrophotometer to assess 260/280 and 260/230 nm ratios. All RNA samples met a 260/280 ratio >2.0 and 260/230 ratios in the range of 2.0\u20132.2. cDNA was generated from reverse transcription of 2\u03bcg total RNA using a Transcriptor First Strand cDNA Synthesis Kit . cDNA was quantified using the SYBR Green assay, and the relative gene expression levels were calculated against GAPDH or U6 by using the \u2206\u2206Ct method. The primers we used for qRT-PCR were as follows: w/v), 0.3%Triton X-100, and 0.2% sodium azide for 2 h at room temperature. The primary antibodies we used were as follows: anti-Iba1 , anti-GFAP , anti-NeuN . After overnight incubation at 4 \u00b0C with primary antibodies and washing with PBS for 30 min, brain sections were incubated with the secondary antibodies conjugated with Alexa Fluor 488 or 594 (1:500). Finally, sections were stained with DAPI and mounted on glass slides with adhesion anti-fade medium. Confocal images were obtained on a ZEISS 710 confocal laser-scanning microscope. Image analyses and quantification were performed using ImageJ software V1.53 .Mice were anesthetized and transcardially perfused with cold phosphate-buffered saline (PBS), followed by 4% paraformaldehyde (PFA) in PBS (pH 7.4). Brains were dissected out and postfixed in 4% PFA overnight. Brains were then equilibrated in 30% sucrose and sectioned into segments 40 \u03bcm-thick. Brain sections were washed three times (10 min each) with PBS and then blocked in 3% BSA  staining  was performed to detect the neuronal apoptosis. Briefly, hippocampal tissue sections were washed for 10 min with PBS and incubated in 2% BSA and 0.25% triton X-100 for 30 min at RT. Sections were then incubated with 200 \u03bcL TUNEL reaction mixture at 37 \u00b0C for 1 h, followed by 3 washes with PBS. Subsequently, sections were incubated with anti-NeuN  antibody overnight at 4 \u00b0C. Finally, sections were washed for 10 min with PBS 3 times and then incubated with the secondary antibodies conjugated to Alexa Fluor 488  at RT.Brain tissues were lysed with RIPA buffer . Protein samples were separated on 8\u201312% SDS-PAGE gels and transferred to polyvinylidene fluoride (PVDF) membranes . The PVDF membranes were then blocked in TBS-T containing 3% milk and incubated with primary STING antibodies  at 4 \u00b0C overnight. PVDF membranes were then incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies (1:5000) at room temperature for 2 h. Finally, the immunoreactive proteins were treated with enhanced chemiluminescence reagent . The 5200CE Tanon\u2122 Chemi-Image System was used to obtain the images of the blots, and the band intensity of the blots was analyzed using the software ImageJ.STING 3\u2032UTRs were amplified from genomic DNA by PCR and cloned into the dual luciferase reporter vector pmirGLO . Primers were used for the cloning 3\u2032UTRs of STING as follows: mouse STING forward: CTGTGGTCTCCACGATGACTTGA, mouse STING reverse: CACCCAGGTCTCCAACCTTTAAA; human STING forward: CAGTGGTCTCCAAGCCTCTG, human STING reverse: ATGGAACATGACCAGGAGCCA. Mutagenesis of the putative let-7i binding sites on STING CDS or 3\u2032UTR was performed using the Quick-Change II Site-directed Mutagenesis Kit  according to the manufacturer\u2019s protocol. The primers for subcloning the mutated CDS or 3\u2032UTR were as follows: mouse mutant STING forward: CATAatggagtGTTGGATGTTTGGCC, mouse mutant STING reverse: CCAACactccatTATGTCAGCAGTGTT; human mutant STING forward: TCACTGCCTatggagCCTCACG, human mutant STING reverse: TGAGGctccatAGGCAGTGATTATGA. All plasmid constructs were then verified by sequencing. Dual luciferase transfection assays were performed as previously described . A. A10]. AF = 101.2, p < 0.01; Sham vs. HSI(3 dpi), p < 0.01; Sham vs. HSI(7 dpi), p < 0.01; Sham vs. HSI(14 dpi), p = 0.986)  by using qRT-PCR. The expression of let-7i at 3 and 7 dpi was significantly lower than that in the sham group, and recovered to the same level as that in the sham group at 14 dpi (= 0.986) B. Furthehttp://www.targetscan.org, accessed on 1 October 2020). Gene Ontology (GO) term analyses  found that these predicted targets were related to biological functions involving transcription, regulation of transcription, transport, protein phosphorylation, pre-miRNA processing, protein ubiquitination, phosphorylation, and mRNA transport  = 101.2, p < 0.001; 0 h vs. 6h, p < 0.001; 0 h vs. 1 d, p < 0.001; 0 h vs. 2 d, p < 0.001; 0 h vs. 3 d, p = 0.790)  B. These TNF-\u03b1, IL-1\u03b2, and IL-6 in the hippocampi of agomir-let-7i-treated and scramble-treated mice at day 3 after HSI. Consistent with our expectation, the levels of TNF-\u03b1, IL-1\u03b2, and IL-6 were significantly elevated in the hippocampi of negative control mice after HSI  = 32.44, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p < 0.05; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.05), IL-1\u03b2  = 41.83, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p < 0.05; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.01), and TNF\u03b1  = 79.65, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p < 0.01; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.001) after HSI  = 42.42, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p = 0.195; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.001) and microglial branch number was greatly increased  = 32.44, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p = 0.741; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.001) in the agomir-let-7i-treated HSI group  = 43.44, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p < 0.01; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.05)  and Iba1 (a microglia marker) revealed a considerable reduction in the size of the glial scar after agomir-let-7i treatment in comparison to the scramble control, while the sham group did not reveal any glial scar formation in the hippocampus ( < 0.05) E,F. Thes+/NeuN+ cells was significantly decreased in the agomir-let-7i-treated group compared to that in the scramble-treated group  = 41.55, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p < 0.05; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.01)  staining of hippocampal tissue sections from sham, scramble-treated, and agomir-let-7i-treated mice at day 7 post-injury. The number of TUNEL < 0.01) A,B, indiGiven that agomir-let-7i could reduce neuronal apoptosis as well as inflammatory response and glial scar size in the hippocampus, we speculated that agomir-let-7i-treated mice might perform better than scramble-treated mice on learning and memory tests. To test this hypothesis, we conducted the rotarod test and the Barnes maze test to evaluate hippocampal integrity in HSI mice beginning at 15 dpi.F = 173.0, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p < 0.01; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.001)  B, suggesF = 26.36, p < 0.001; Sham vs. HSI + Scramble, p < 0.01; Sham vs. HSI + Agomir-let-7i, p < 0.05; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.01)  D. AltogeSTING (TMEM173) is known to play a pivotal role in responding to pathogenic DNA and self-DNA in the context of neurodegenerative and autoimmune disorders [STING mRNA  = 169.3, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p = 0.803; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.001)  = 35.86, p < 0.001; Sham vs. HSI + Scramble, p < 0.001; Sham vs. HSI + Agomir-let-7i, p = 0.436; HSI + Scramble vs. HSI + Agomir-let-7i, p < 0.01)  D. Moreov < 0.01) E. AltogeF = 159.2, p < 0.001; Sham vs. PVD stroke + Scramble, p < 0.001; Sham vs. PVD stroke + Agomir-let-7i, p < 0.001; PVD stroke + Scramble vs. PVD stroke + Agomir-let-7i, p < 0.001) and protein levels  = 360.4, p < 0.001; Sham vs. PVD stroke + Scramble, p < 0.001; Sham vs. PVD stroke + Agomir-let-7i, p = 0.138; PVD stroke + Scramble vs. PVD stroke + Agomir-let-7i, p < 0.001) of STING were significantly decreased in the cortex of agomir-let-7i-treated PVD stroke mice  A,B. Furtoke mice E,F. TherIn the present study, we found that TBI decreased let-7i and that the intranasal administration of agomir-let-7i reduced brain damage in TBI mice as well as in PVD stroke mice. Agomir-let-7i administration improved cognitive function in brain-injured mice. Mechanistically, agomir-let-7i administration suppressed neuroinflammation, glial scar formation, and neuronal apoptosis after brain injury, suggesting that agomir-let-7i may serve as a potential therapeutic candidate against injury-induced neuroinflammatory and neurodegenerative diseases, such as TBI and PVD stroke.Both TBI and PVD stroke have primary and secondary injury phases regardless of the severity of the insult. The primary injury encompasses mechanical damage to the brain tissues that release signals that activate microglia, astrocytes, and infiltrated peripheral immune cells to initiate the secondary injury, including inflammation, excitotoxicity, mitochondrial impairment, and neuronal cell death. The secondary injury phase evolves over minutes to days to months after the primary injury, suggesting that early interventions may ameliorate brain damage and stimulate neural regeneration and repair . The maiHuman TBI is a sophisticated disease process, and the diversity in the extent of injury as well as in pathoanatomical subtypes means that different patients are likely to experience different courses and outcomes of TBI. In addition, it is still difficult to generate animal models that can fully recapitulate all the pathophysiological aspects of human TBI. Let-7i has been considered as a promising serum biomarker for stroke and blast-induced TBI. For example, the expression of let-7i is upregulated in blood samples collected from patients with post-stroke cognitive impairment compared with patients with post-stroke cognitive normality . SimilarSTING) was a downstream target of let-7i in the brain, and the intranasal administration of agomir-let-7i could suppress the upregulation of STING, neuroinflammation, and glial scar formation in both TBI and PVD stroke mice. These findings are consistent with previous work that identified STING as a key regulator of inflammation in both rodents and human brain organoids and found that the inactivation of STING attenuates inflammation [Neuroinflammation plays an essential role in the pathophysiology of TBI and stroke. Although a low degree of neuroinflammation is initially beneficial for debris clearance and repair, a high degree of neuroinflammation elicits secondary injury that leads to chronic inflammation and neurodegeneration . Mechaniammation ,32,42,43ammation . Althougammation . Our datIn summary, we found that brain-enriched miRNA let-7i was significantly downregulated at the early stages of TBI in mouse brains. STING was a direct downstream target of let-7i, and agomir-let-7i could protect brain tissue from neuroinflammation, glial scar formation, and neural cell death after brain injury. Overall, our data suggest that the intranasal administration of agomir-let-7i is a potential therapeutic strategy for neurotrauma and PVD stroke."}
+{"text": "The rumen contains a complex microbial ecosystem that degrades plant materials, such as cellulose and hemicellulose. We herein reconstructed 146 nonredundant, rumen-specific metagenome-assembled genomes (MAGs), with \u226550% completeness and <10% contamination, from cattle in Japan. The majority of MAGs were potentially novel strains, encoding various enzymes related to plant biomass degradation and volatile fatty acid production. The MAGs identified in the present study may be valuable resources to enhance the resolution of future taxonomical and functional studies based on metagenomes and metatranscriptomes. This study was the first to use JB (arXiv.https://arxiv.org/abs/1303.3997) to the bovine reference genome ARS-UCD1.2/bosTau9. Filtered reads were assembled in SPAdes version 3.13.0 (arXiv.https://arxiv.org/abs/1303.3997). We binned MAGs with contigs using MetaBAT2 version 2.15  to prevent arbitrary mapping between similar genomes  with the following parameters: -m relative_abundance --min-read-percent-identity 0.95 -\u200d-\u200dmin-read-aligned-percent 0.75. We taxonomically classified RUG1ANI99% MAGs in GTDB-tk version 2.1.1  with GTDB release 207. We then built a phylogenetic tree in PhyloPhlAn version 3.0.60 , including Hungate1000 (strains (<99%) using the ANI outputs by dRep and GTDB-tk.Filtered reads were mapped against RUG1n 3.0.60 . To elucgate1000  using dRANI99% MAGs with Prodigal version 2.6.3 (Genomes (KEGG) database through GhostKOALA ( Carbohydrate-active enzyme (CAZyme) families, which are associated with cell wall degradation and essential for efficient lignocellulose processing in ruminants, were annotated using dbCAN2 (\u20135 to search cohesin (PF00963) and dockerin (PF00404) domains. We used PULpy  in 5,414 public Bacteroidetes genomes using PULpy. bioRxiv.https://doi.org/10.1101/421024) to predict polysaccharide utilization loci (PUL), linked gene clusters that encode the cell envelope-associated enzymes required for sensing, binding, and degrading polysaccharide substrates . Following dereplication at 99% ANI, we generated 146 nonredundant RUG13.2\u200d \u200dkb . A compaectively . TherefoANI95% MAGs, we identified 32 MAGs that were present (relative abundance >0) in >90% of the tested cattle (n=20), suggesting that they are core rumen bacteria in Japan , Actinobacteriota (10 MAGs), Firmicutes (9 MAGs), Verrucomicrobiota (7 MAGs), Spirochaetota (4 MAGs), Patescibacteria (3 MAGs), Proteobacteria (2 MAGs), Methanobacteriota (2 MAGs), Planctomycetota (1 MAG), Synergistota (1 MAG), Desulfobacterota_I (1 MAG), and Elusimicrobiota (1 MAG). In Bacteroidota MAGs, 10 RUG1ANI99% MAGs were classified as Prevotella, which is the most abundant bacterial genus in the rumen  with at least one gene related to acetate production . Firmicutes_C RUG1ANI99%These genes were absent from  MAGs. Rumen propionate is produced via the succinate and\u200d \u200dacrylate pathways . Prevotella  and GH77 (4-\u03b1-glucanotransferase), respectively, both of which are involved in starch degradation , GH3 (\u03b2-glucosidase), GH5 , GH13 (\u03b1-amylase), GH26 , GH32 (invertase), GH36 , GH43 (\u03b2-xylosidase), GH73 (lysozyme), GH94 (cellobiose phosphorylase), and GH97 (\u03b1-glucosidase).Among 146 RUG1radation . Further CAZymes , which iANI99% MAGs. The proteins containing cohesin and dockerin domains are summarized in ANI99% MAGs, including 2 Ruminococcus and 11 Ruminococcus_E MAGs. Sixteen dockerin-containing proteins carried CAZyme domains, mostly including GH families containing \u03b1-amylase . Among the 25 RUG1ANI99% MAGs, 13 had both cohesin- and dockerin-containing proteins, while 9 had only dockerin-containing proteins. Ruminococcus albus 8, which degrades cellulosic substrates, harbored no cohesion-containing protein. Therefore, we cannot rule out the possibility that RUG1ANI99% MAGs, which had no cohesion- and some dockerin-containing proteins, used an alternative mechanism for the immobilization of dockerin-containing enzymes onto carbohydrates, similar to Ruminococcus albus 8 . Ninety-eight and 80 PULs were identified in Cryptobacteroides MAG and Prevotella MAGs, respectively, and contained various CAZymes. Overall, the present results suggest that Cryptobacteroides and Prevotella are important for rumen function  in cattle in Japan.In  domains . The mosIn summary, we reconstructed 146 rumen-specific MAGs from cattle in Japan, with many being potentially novel. These MAGs are valuable resources for enhancing the resolution of future metagenome- and metatranscriptomic-based taxonomical and functional studies.Microbes Environ 37: ME22039.Sato, Y., Takebe, H., Oishi, K., Yasuda, J., Kumagai, H., Hirooka, H., and Yoshida, T. (2022) Identification of 146 Metagenome-assembled Genomes from the Rumen Microbiome of Cattle in Japan. https://doi.org/10.1264/jsme2.ME22039Supplementary Material 1Supplementary Material 2Supplementary Material 3Supplementary Material 4Supplementary Material 5Supplementary Material 6Supplementary Material 7"}
+{"text": "Correction: Immun Ageing 19, 57 (2022)https://doi.org/10.1186/s12979-022-00310-yFollowing publication of the original article , the autThe incorrect author name is: J\u00e9ssica S. de GuedesThe correct author name is: J\u00e9ssica de S. GuedesThe author group has been updated above and the original article  has been"}
+{"text": "ENDOGLIN (ENG) gene. Here, we generated induced pluripotent stem cells (hiPSCs) from a patient with rare mosaic HHT1 with tissues containing both mutant (ENGc.1678C>T) and normal cells, enabling derivation of isogenic diseased and healthy hiPSCs, respectively. We showed reduced ENG expression in HHT1 endothelial cells (HHT1-hiPSC-ECs), reflecting haploinsufficiency. HHT1c.1678C>T-hiPSC-ECs and the healthy isogenic control behaved similarly in two-dimensional (2D) culture, forming functionally indistinguishable vascular networks. However, when grown in 3D organ-on-chip devices under microfluidic flow, lumenized vessels formed in which defective vascular organization was evident: interaction between inner ECs and surrounding pericytes was decreased, and there was evidence for vascular leakage. Organs on chip thus revealed features of HHT in hiPSC-derived blood vessels that were not evident in conventional 2D assays.Hereditary hemorrhagic telangiectasia (HHT) is a genetic disease characterized by weak blood vessels. HHT1 is caused by mutations in the   \u2022Vessels from isogenic hiPSCs from HHT1 patients compared\u2022HHT1-hiPSC-ECs show defective vascular organization in 3D microfluidic chips\u2022HHT1-hiPSC-ECs show defective EC-pericyte interaction In this article, Orlova and colleagues describe 3D vessels on chip (VoCs) composed of hiPSC-ECs from a patient with hereditary hemorrhagic telangiectasia (HHT1). HHT1-hiPSC-ECs and healthy isogenic controls behaved similarly in 2D culture, but defective vascular organization and reduced pericyte coverage were evident in HHT1-hiPSC-ECs in VoCs. The model is thus a valuable tool for mechanistic studies and future drug discovery. ENG; HHT1), Activin receptor like kinase-1  or SMAD4 (HHT3), genes that mediate signaling by transforming growth factor \u03b2 (TGF-\u03b2) and bone morphogenetic protein (BMP) in vascular endothelial cells (ECs) (in\u00a0vitro makes them unsuitable as a renewable source of ECs for reproducibly modeling the disease in humans and for drug discovery.Hereditary hemorrhagic telangiectasia (HHT) is an inherited genetic disorder caused by autosomal dominant mutations in Endoglin (ls (ECs) . Phenotyls (ECs) . These als (ECs) . More sels (ECs) . To datels (ECs) . Medicalls (ECs) , thalidols (ECs) , itraconls (ECs) , and othls (ECs) . Geneticls (ECs) . Attemptls (ECs) . Blood ols (ECs) , but thein\u00a0vitro and (2) investigating defective endothelial-pericyte interactions.In the present study, we aimed to establish an efficient and scalable system that would recapitulate the formation of defective blood vessels in patients with HHT, based on\u00a0patient-derived human induced pluripotent stem cells\u00a0(HHT1-hiPSCs). We hypothesized that HHT1-hiPSCs might be useful for (1) identifying mechanisms underlying disease predisposition and modeling clinical features of HHT1 ENG (NM_001114753.2 (ENG):c.1678C>T; p.(Gln560\u2217)), which causes ENG haploinsufficiency (c.1678C>T and HHT1WT) (c.1678C>T-hiPSC-ECs compared with HHT1WT-hiPSC-ECs (ID1 expression was significantly upregulated in HHT1c.1678C>T-hiPSC-ECs after 2\u00a0h of TGF-\u03b2 treatment (hiPSC lines were generated from somatic tissue from a patient with HHT1 with a heterozygous nonsense mutation in ficiency . The pat HHT1WT) D\u2013S1G. HH HHT1WT) ; 2014b. iPSC-ECs A and 1B.iPSC-ECs B. ENG haiPSC-ECs A. HHT1-hreatment B.Figure\u00a0c.1678C>T-hiPSC-ECs  C. BarrieiPSC-ECs C and S2DiPSC-ECs C and S2DiPSC-ECs E.in\u00a0vitro was examined, as described previously .Finally, the ability to form 2D vascular networks eviously , 2014b. eviously F. Quantibranches G as wellc.1678C>T-hiPSC-ECs compared with HHT1WT-hiPSC-ECs  model was then examined A. PrimariPSC-ECs B and 2C,iPSC-ECs A. QuantiiPSC-ECs C and S3Dimilarly B. Furthee nuclei E and S3Fe nuclei . Notablyc.1678C>T-hiPSC-ECs compared with HHT1WT-hiPSC-ECs.Junctional integrity was examined by immunostaining of the microvascular networks with VEC and ZO1 D and 2E.c.1678C>T-hiPSC-ECs showed reduced pericyte coverage compared with HHT1WT-hiPSC-ECs   C\u2013S4E.Figc.1678C>T-hiPSC-ECs compared with HHT1WT-hiPSC-ECs  was added into the medium channel of the organ-on-chip device, and real-time videos of vascular segments pre-stained using fluorescent agglutinin were made . QuantifiPSC-ECs B and 4C.in\u00a0vitro model for the genetic vascular disorder HHT using hiPSCs derived from patients with mutations in the ENG gene (HHT1). The results showed that we likely captured the direct effects of reduced ENG protein on the EC surface without compensation or adaption mechanisms that normally occur in\u00a0vivo, notably in mutant mice  was used to knock down onic ECs . Completin\u00a0vivo  . The generation of the lines was approved by the Leiden University ethics committee under the P13.080 \u201cParapluprotocol: hiPSC.\u201d Patient samples, fibroblasts from skin biopsies, and erythroblasts isolated from peripheral blood were used for reprogramming. Reprogramming with episomal vectors was done as described, except that a newer generation of vectors without ere used . hiPSCs ere used . KaryotyRA-FISH) , and pluOne-way ANOVA and non-parametric Student\u2019s t test for unpaired measurements were applied as appropriate to test for differences in means between the groups. Detailed statistics are indicated in each figure legend. Data are expressed and plotted as the mean \u00b1 SD. Statistical significance is indicated in each figure legend. Statistical analysis was performed with GraphPad Prism 9.0.2.V.V.O., designed the research, established functional assays, performed experiments, and wrote the manuscript; D.M.N. and A.C., performed experiments in 3D vascular chips, did imaging, and performed quantification; X.C., performed EC differentiation; C.F., performed reprogramming experiments; F.v.d.H., conducted EC differentiation and isolation and FACS; F.L., assisted with quantification of microfluidic experiments; C.J.J.W., R.J.S., and H.-J.M., provided HHT patient samples; J.K.P.v.A., conducted genetic analysis; P.t.D. and F.L. helped analyze the data; C.L.M., designed the research and wrote the manuscript.The authors declare no competing interests."}
+{"text": "This study inventively combines epidermal growth factor receptor (EGFR) expression of the primary lesion and standardized uptake value (SUV) of positron emission tomography and computed tomography (PET/CT) to predict the prognosis of nasopharyngeal carcinoma (NPC). This study aimed to evaluate the predictive efficacy of maximum standard uptake value (SUVmax) and EGFR for treatment failure in patients with NPC.18F-FDG PET/CT of 313 patients with NPC. Time-dependent receiver operator characteristics was used for analyzing results and selecting the optimal cutoff values. Cox regression was used to screen out multiple risk factors. Cumulative survival rate was calculated by Kaplan\u2013Meier.This retrospective study reviewed the results of EGFR expression and pretreatment p\u2009=\u20090.0083), locoregional relapse-free survival (LRRFS) (p\u2009=\u20090.0077), distant metastasis-free survival (DMFS) (p\u2009=\u20090.013), and progression-free survival (PFS) (p\u2009=\u20090.0018) among the four groups. Patients in the EGFR-positive and SUVmax-T\u2009>\u20098.5 group had the worst survival, while patients in the EGFR-negative and SUVmax-T\u2009\u2264\u20098.5 group had the best prognosis. Subsequently, patients with only positive EGFR expression or high SUVmax-T were classified as the middle-risk group. There were also a significant difference in 3-year overall survival among the three risk groups (p\u2009=\u20090.034). SUVmax-T was associated with regional recurrence-free survival and LRRFS in multivariate analysis, whereas EGFR was an independent prognostic factor for LRRFS, DMFS, and PFS.The selected cutoff value of SUVmax-T was 8.5. The patients were categorized into four groups according to EGFR expression and SUVmax-T. There were significant differences in the 3-year local recurrence-free survival (LRFS) (The combination of SUVmax-T and EGFR expression can refine prognosis and indicate clinical therapy.The online version contains supplementary material available at 10.1186/s13014-023-02231-6. Nasopharyngeal carcinoma (NPC) is a highly aggressive malignant tumor believed to arise from nasopharyngeal epithelial cells . Accordi18F-fluorodeoxyglucose positron emission tomography and computed tomography (18F-FDG PET/CT) have been frequently used in pretreatment diagnostic evaluation and post-treatment monitoring because of their unique capability to image metabolically active lesions [The  weight)]. SUVmax-Radiotherapy dose and target volume delineation were performed according to the recommendations (Radiation Therapy Oncology Group) , 23. ThePatients with stage I were treated with radiotherapy alone, while patients with stage II-IV received radiotherapy combined with chemotherapy .After RT, follow-up was conducted once every 3\u00a0months for the first 2\u00a0years, once every 6\u00a0months in years 3 to 5, and annually thereafter. The final follow-up date was March 2022. Study endpoints included local recurrence-free survival (LRFS), regional recurrence-free survival (RRFS), locoregional relapse-free survival (LRRFS), distant metastasis-free survival (DMFS), progression-free survival (PFS), and OS.P-values\u2009<\u20090.05 indicated statistical significance.Uses IBM SPSS statistical software version 22.0 and R software version 4.0.5 for the statistical analysis. The optimal cutoff values was decided by Time-dependent receiver operator characteristic (ROC) analysis . Kaplan\u2013Meier methods were used to compute the survival analyses. Between-group differences in survival outcomes were assessed using log-rank tests. Cox regression was used to screen out multiple risk factors. All tests were two-tailed, and The clinical characteristics of all eligible patients are summarized in Table P\u2009=\u20090.075), 97.9% vs. 93.2% (P\u2009=\u20090.084), 94.7% vs. 87.6% (P\u2009=\u20090.032), 97.0% vs. 87.4% (P\u2009=\u20090.018), 91.8% vs. 76.7% (P\u2009=\u20090.0017), and 96.5% vs. 90.8% (P\u2009=\u20090.058), respectively .Among 1877 patients with NPC with IHC examination for primary lesions, EGFR was detected in 73.36% (1377 patients), and the proportion of patients showing negligible intensity (negative) of EGFR staining was 26.64% (500 patients). For the enrolled 313 patients, 241 (77.00%) showed positive EGFR expression, while 72 (23.00%) showed negligible expression (negative). The 3-year LRFS, RRFS, LRRFS, DMFS, PFS, and OS rates in the EGFR-negative group vs. EGFR-positive group were 96.9% vs. 92.2% (p\u2009=\u20090.0038), 96.3% vs. 95.3% (p\u2009=\u20090.21), 93.8% vs. 84.2% (p\u2009=\u20090.0063), 93.5% vs. 86.5% (p\u2009=\u20090.022), 85.8% vs. 75.5% (p\u2009=\u20090.018), and 94.4% vs. 93.5 (p\u2009=\u20090.066), respectively .The mean SUVmax-T was 10.21\u2009\u00b1\u20095.59 , and the mean SUVmax-N was 7.53\u2009\u00b1\u20095.55 . To further evaluate the prognostic value of SUVmax, time-dependent ROC analysis was used to determine the optimal cutoff values based on the 3-year survival outcome. The optimal cutoff value of SUVmax-T was 8.5 based on the 3-year LRFS , LRRFS (p\u2009=\u20090.0077), DMFS (p\u2009=\u20090.013), and PFS (p\u2009=\u20090.0018) .To better predict the prognosis of NPC, patients were divided into the following four groups based on SUVmax-T and EGFR expression: (a) EGFR negative and low SUVmax-T, (b) EGFR negative and high SUVmax-T, (c) EGFR positive and low SUVmax-T, and (d) EGFR positive and high SUVmax-T. There were obviously statistical difference in 3-year LRFS (18) Fig.\u00a0 among thp\u2009=\u20090.0029), LRRFS (p\u2009=\u20090.0026), DMFS (p\u2009=\u20090.005), and PFS (p\u2009=\u20090.00073), the 3-year OS also showed significant difference among 3 risk groups .Figure\u00a0p\u2009=\u20090.005) and LRRFS . Moreover, multivariable survival analysis revealed that EGFR was an independent prognostic factor for LRRFS , DMFS , and PFS .Six variables  were included in the univariate analysis for the six clinical endpoints, and the results are summarized in Additional file The most important prognostic factor for NPC is TNM clinical stage. However, there is considerable variability in outcomes among patients with the same TNM stage receiving the same treatment . Recent PET/CT is functional imaging, which is different from morphology and structure imaging. The parameters of PET can be used to characterize the burden of metabolically active lesions and biological aggressiveness in malignancies. SUV, TLG, and MTV are parameters that have been correlated with survival outcome \u201316. SUVmIn 2008, a retrospective study have shown SUVmax may predict DFS in NPC treated with CCRT and more aggressive treatment should be given to patients with higher SUVmax . A prospSeveral meta-analyses have shown that EGFR overexpression is significantly associated with poor OS and DFS. Thus, EGFR may serve as a potential prognostic predictor of NPC , 37, 38.p\u2009=\u20090.0083), LRRFS (p\u2009=\u20090.0077), DMFS (p\u2009=\u20090.013), and PFS (p\u2009=\u20090.0018) among the four groups. The K\u2013M curve revealed that patients in the EGFR-positive and SUVmax-T\u2009>\u20098.5 group had the worst survival, while patients in the EGFR-negative and SUVmax-T\u2009\u2264\u20098.5 group had the best prognosis. For patients in the EGFR-negative\u2009+\u2009SUVmax-T\u2009>\u20098.5 group and EGFR-positive\u2009+\u2009SUVmax-T\u2009\u2264\u20098.5 group, the cumulative survival curves were extremely close. This demonstrated that EGFR expression and high SUVmax-T may be adverse prognostic factors for NPC. Therefore, we further divided patients into three groups. Patients with positive EGFR expression or high SUVmax-T levels were defined as the middle-risk group. The 3-year OS also showed a significant difference among the three risk groups (p\u2009=\u20090.034). Moreover, SUVmax-T was associated with LRFS and LRRFS in multivariate analysis, whereas EGFR was an independent prognostic factor for LRRFS, DMFS, and PFS.To better predict the clinical outcomes of de novo NPC, we combined functional imaging data with molecular pathological data. Patients were categorized into four groups based on EGFR expression (negative or positive) and SUVmax-T (\u2264\u20098.5 or\u2009>\u20098.5). There were significant differences in 3-year LRFS ; B regional recurrence-free survival (RRFS); C locoregional relapse-free survival (LRRFS); D distant metastasis-free survival (DMFS); E progression-free survival (PFS); F overall survival (OS). Fig. S3: Kaplan-Meier curves in the low SUVmax-T (\u22648.5) group and the high SUVmax-T (>8.5) group. A local recurrence-free survival (LRFS); B regional recurrence-free survival (RRFS); C locoregional relapse-free survival (LRRFS); D distant metastasis-free survival (DMFS); E progression-free survival (PFS); F overall survival (OS)."}
+{"text": "Vigna unguiculata (lobia) production remains poorly understood. Thus, we aimed to isolate and characterize the soil microbes from the rhizosphere and develop novel microbial consortia for enhancing lobia production. Fifty bacterial strains were isolated from the rhizosphere soil samples of lobia. Finally, five effective strains  were identified and molecularly characterized by 16\u00a0S rDNA gene amplification. All selected strains showed positive plant growth promoting (PGP) properties in broth culture. Based on morphological, biochemical, and plant growth promoting activities, five effective isolated strains and two collected strains (Azospirillum brasilense MTCC-4037 and Paenibacillus polymyxa BHUPSB17) were selected. The pot trials were conducted with seed inoculations of lobia (Vigna unguiculata) var. Kashi Kanchan with thirty treatments and three replications. The treatment combination T3 (Pseudomonas sp. IESDJP-V2), T14 (Pseudomonas sp. IESDJP-V2\u00a0+\u00a0A. brasilense), T26 (Pseudomonas sp. IESDJP-V1+ B. cereus IESDJP-V4\u00a0+\u00a0P. polymyxa) and T27 (IESDJP-V1+ IESDJP-V5+ A. brasilense) were recorded for enhancing plant growth attributes, yield, nutritional content like protein, total sugar, flavonoid and soil properties as compared to control and others. The effective treatments T3 (Pseudomonas sp.), T14 (Pseudomonas sp. IESDJP-V2\u00a0+\u00a0A. brasilense), T26 (Pseudomonas sp. IESDJP-V1+ B. cereus IESDJP-V4\u00a0+\u00a0P. polymyxa) and T27 (IESDJP-V1+ IESDJP-V5+ A. brasilense) recorded as potential PGPR consortium for lobia production. The treatment of single (Pseudomonas sp.), duel (IESDJP-V2\u00a0+\u00a0A. brasilense) and triple combination (IESDJP-V1+ IESDJP-V4\u00a0+\u00a0P. polymyxa) and (IESDJP-V1+ IESDJP-V5+ A. brasilense) can be further used for developing effective indigenous consortium for lobia production under sustainable farming practices. These PGPR bio-inoculant will be cost-effective, environment-friendly and socially acceptable.The rhizosphere microbes play a key role in plant nutrition and health. However, the interaction of beneficial microbes and  The gThe application of microbial consortium or inoculum in single, dual, triple, tetra, penta and hexa, and more combinations are widely used for increasing sustainable agricultural productivity as well as enhancing soil fertility and health ,8. PGPR Vigna unguiculata L.) is an important seed legume crop that is broadly used as green vegetables in India and other countries of World , 10.13039/501100021098Design and Innovation Centre, BHU and Indian Institute of Technology (IIT-BHU) Varanasi [DIC-BHU/Project S-23 Approval/2016-17/693].Professor Jay Prakash Verma was supported by Data associated with this study has been deposited at NCBI-GenBank and got accession number of Pseudomonas sp. IESDJP-V1 (MH362754) and Pseudomonas sp. IESDJP-V2 (MH362755), S. marcescens IESDJP-V3 (MH362756), B. cereus IESDJP-V4 (MH362757), Ochrobactrum sp. IESDJP-V5 (MH362758).The authors declare no competing interests.No additional information is available for this paper."}
+{"text": "Pseudanabaena spp. are filamentous cyanobacteria widely distributed in temperate lakes. Though infrequent, they can form harmful algal blooms. Here, we present a high-quality metagenome-assembled genome of a Pseudanabaena sp. from a toxic, crimson cyanobacterial bloom in Lake Salubria, NY. Pseudanabaena spp. are filamentous, nonheterocystous cyanobacteria widely distributed in temperate lakes . In spring 2020, an algal bloom turned the lake a vivid crimson, concerning the community . Grab saN50 = 5,395\u2009bp).Filaments were filtered onto a 5-\u03bcm polycarbonate filter from which genomic DNA was extracted using standard phenol-chloroform methods (De novo assembly was conducted using Unicycler (v0.4.9b) in normal mode  . Illuminmal mode . CheckM mal mode . GTDB-Tkmal mode . The genmal mode . The 16SPseudanabaena sp. Salubria-1, consisting of 107 contigs with a total length of 7,138,199\u2009bp  and a GC content of 41.93%. The sequencing depth was ~191-fold. The genome was estimated at 99.1% complete with 3.3% contamination. The closest taxonomic placement (91.9% average nucleotide identity [ANI]) was to Pseudanabaena sp. strain UWO311, a red strain isolated from Dickson Lake, Ontario, Canada (Pseudanabaena/Limnothrix group (We recovered a MAG of bp N50 = 5,348\u2009bp JALQCR000000000. Reads are deposited in the NCBI Sequence Read Archive under accession numbers SRX15003418 and SRX15003417.The metagenome-assembled genome is deposited at DDBJ/ENA/GenBank under accession number"}
+{"text": "In this study, we aimed to investigate the effect of p62 on angiogenesis and microRNA (miRNA) expression profiles in acute myeloid leukemia (AML) exosomes.An Exiqon v19.0 microRNA MicroArray was used to profile miRNAs in exosomes derived from parental and p62-knockdown U937 cells. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases were used to predict the biological functions and potential mechanisms of differentially expressed miRNAs in AML exosomes. Endothelial cell tube formation assays using human umbilical vein endothelial cells (HUVECs) were performed to investigate the effect of AML exosomes on angiogenesis.P < 0.05). GO analysis indicated that miRNAs were most enriched in the intercellular pathways. Biological process analysis revealed that 1460 biological processes were associated with downregulated transcripts, including 19 pathways related to vesicles, and 1,515 pathways were upregulated, including 8 pathways related to vesicles. Molecular function analysis indicated that protein binding, transcription regulator activity, and DNA-binding transcription factor activity were enriched (P < 0.05). Pathway analysis indicated that 84 pathways corresponded to upregulated transcripts, and 55 pathways corresponded to downregulated transcripts (P < 0.05). We also found that exosomes derived from U937 cells promoted angiogenesis in HUVECs.We demonstrated that 2,080 miRNAs were expressed in exosomes derived from our cultured cell samples, of which 215 and 208 miRNAs were upregulated and downregulated, respectively, in p62-knockdown U937 cells (fold change \u2265 2, Our data suggest that exosomal miRNAs may play important roles in the pathogenesis of AML, which may be treated by p62 knockdown with exosomal miRNAs to inhibit angiogenesis. Acute myeloid leukemia (AML) is a fatal hematological malignancy with high recurrence rate. For patients receiving the most intensive treatment, the overall 5-year survival rate remains below 50%. For the remaining patients, the prognosis is even worse . AML is Exosomes are nanometer-scale extracellular vesicles containing many microRNAs (miRNAs) that are secreted from cells in both normal and pathological conditions . It has In this study, we constructed a miRCURYTM LNA Array (v.19.0) of miRNAs in exosomes derived from AML cells after p62 knockdown. The miRNAs in exosomes were analyzed by identifying signature miRNAs. We then investigated angiogenesis in human umbilical vein endothelial cells (HUVECs) exposed to exosomes derived from parental U937 cells, p62-knockdown U937 cells, or control cells. The data from these studies may shed light on the relationship between exosomal miRNAs and AML, further enhancing our understanding of AML progression.Our study may aid the development of potential biomarkers for the diagnosis and prognosis of AML progression.SQSTM1 gene (LV-SQSTM1-RNAi) and an empty recombinant adenovirus vector (Hu6-MCS-CMV-EGFP) were constructed. U937 cells were placed in a six-well plate. Polybrene  was used for the transfection. The transfection system included 1.8 mL RPMI-1640 with 10% fetal bovine serum, 10\u00a0\u00b5L LV-SQSTM1-RNAi or Hu6-MCS-CMV-EGFP, and 0.9 \u00b5L polybrene. After transfection for 24 h, the cell suspension was collected and centrifuged at 800 rpm for 5 min, the supernatant was discarded, and two mL of RPMI-1640 with 10% fetal bovine serum was added. After transfection for 48 h, fluorescence was observed. After culturing, 5 \u00b5g/mL puromycin was added, and the cells were screened for 15 days. After selecting the surviving cells, cell lines with clonal stability were cryopreserved and characterized through RT\u2013qPCR and western blotting.The human acute monocytic leukemia cell line U937 was purchased from the Bena Culture Collection  and stored in our laboratory. A recombinant lentivirus vector-mediated 4 cells/well in 96-well plates and allowed to grow for 12, 24, and 48 h. Next, 10\u00a0\u00b5L CCK-8 solution  was added to the cell suspension and incubated for 2 h. Absorbance was measured using a spectrophotometer at 450\u00a0nm. The experiment was repeated at least three times.U937 cells were plated at a density of 3\u20135\u00a0\u00d7\u00a010Flow cytometry analysis using an Annexin V-FITC/PI detection kit was used to compare the apoptosis rate of p62-control and p62 knockdown U937 cells. After 48 h of incubation, the cells were washed with phosphate-buffered saline and resuspended in 400 \u00b5L of 1\u00a0\u00d7 binding buffer. Thereafter, 5 \u00b5L Annexin V-FITC and 5 \u00b5L PI were added to the mixture and stained in the dark for 15 min at room temperature. Apoptosis was detected using flow cytometry  immediately after staining.P62-siRNA coated with lentivirus interfered with U937 cells to downregulate the expression of p62, with the empty virus vector used as a control. Two groups of cells were used as follows: p62-knockdown U937 cells and controls. The two groups of cells were cultured for 48 h in serum-free media. The supernatant was collected for exosome extraction via ultracentrifugation. Exosome shape and size were observed using electron microscopy, and exosomal markers were detected using western blot analysis.Total protein was extracted from U937 cells that had or had not been transfected with the p62-encoding gene using radioimmunoprecipitation assay lysis buffer . The protein concentration was determined using the BCA method. Equal amounts of protein samples were added to each well, separated using 10% SDS-PAGE, and transferred to a polyvinylidene chloride transfer membrane . The membrane was blocked with 5% skimmed milk for 2 h. It was then washed with TBST and incubated with primary antibodies against p62, TSG101, CD63, CD9, calnexin, and GAPDH  overnight at 4\u00a0\u00b0C. Thereafter, the membrane was incubated with anti-rabbit or anti-mouse horseradish peroxidase-conjugated secondary antibodies at room temperature for 2 h after washing three times with TBST. An enhanced chemiluminescence substrate (Thermo Fisher Scientific) was used to detect the protein bands. Image Lab software was used to detect and analyze the density of each band .TRIzol  was used to extract total RNA. A NanoDrop spectrophotometer  was used to measure RNA quality and quantity. RNA integrity was assessed using gel electrophoresis. After quality control, miRNA labeling was performed according to the instructions of the miRCURY\u2122 Hy3\u2122/Hy5\u2122 Power Labeling Kit . First, 1 \u00b5L RNA in 2 \u00b5L water was mixed with 1 \u00b5L CIP buffer and CIP . The mixture was then incubated at 37\u00a0\u00b0C for 30 min. The mixture was incubated at 95\u00a0\u00b0C for 5 min to stop the reaction. Then, 3 \u00b5L labeling buffer, 1.5 \u00b5L fluorescent label (Hy3TM), 2 \u00b5L dimethyl sulfoxide, and 2 \u00b5L labeling enzyme were added. The mixture was then incubated for 1 h at 16\u00a0\u00b0C, followed by 15 min at 65\u00a0\u00b0C to terminate the reaction. Hy3-labeled samples were hybridized on the miRCURYTM LNA array  according to the manufacturer\u2019s instructions. A total of 25 \u00b5L Hy3\u2122-labeled samples and 25 \u00b5L hybridization buffer were denatured at 95\u00a0\u00b0C for 2 min and then incubated on ice for 2 min. The hybridization system  was used with the microarray set at 56\u00a0\u00b0C for 16 to 20 h. After hybridization, the slides were washed several times using a washing buffer kit . Finally, an Axon GenePix 4000 B microarray scanner  was used to scan the slides.P value. Finally, hierarchical clustering was used to show the different miRNA expression profiles between the samples.GenePix Pro 6.0 software  was used to extract data by analyzing the imported scanned images. The samples were chosen to calculate normalization factors if the replicated miRNAs were averaged and for miRNAs with intensities \u226530. Median normalization was used to normalize the data. Normalized data = (foreground background)/median; the median was the 50% quantile of miRNA intensity, which was larger than 30 in all samples after background correction. After normalization, the miRNAs with significant differences between the two groups were determined according to the fold change and http://www.targetscan.org/) and mirdbV5 (http://mirdb.org/) are online sites for miRNA target gene prediction (TargetScan7.1 (ediction . In our http://www.geneontology.org) and KEGG (http://www.genome.ad.jp/kegg/) databases to study the potential organisms and signaling pathways of differentially expressed miRNAs. Differences were considered statistically significant at P\u00a0<\u00a00.05.We used the GO , seeded at 3\u00a0\u00d7\u00a0104 cells per well. Tubules were photographed using phase microscopy after incubation for 0, 3, and 6\u00a0h at 37\u00a0\u00b0C with 5% CO2.t-test, and statistical significance was set at P\u00a0<\u00a00.05.We used GraphPad Prism 6 to corroborate the statistical significance of the data for all graphs in this study. Values are presented as the means\u00a0\u00b1\u00a0standard deviation. Differences between groups were analyzed using Student\u2019s To generate p62-knockdown U937 cells, we used a lentivirus to transfect cells and observed the transfection efficiency by fluorescence microscopy. We then conducted RT\u2013qPCR analysis to determine the expression levels of the autophagy gene encoding p62 \u20131B and wP\u00a0<\u00a00.05).We observed exosome formation using electron microscopy after ultracentrifugation. Electron microscopy revealed exosomes as vesicles with a double-layer membrane structure   \u20134F, biol\u00a0<\u00a00.05)  and 6A. \u00a0<\u00a00.05) . As for \u00a0<\u00a00.05) . The miR\u00a0<\u00a00.05)  and 6A.http://www.targetscan.org/) and mirdbV5 (http://mirdb.org/) were employed to predict the potential target genes of miRNAs. We found that 2,018 genes co-expressed with upregulated miRNAs and 2,749 genes co-expressed with downregulated miRNAs in the two databases . Among the three groups of HUVECs+Exo(U937), HUVECs+Exo(p62-con) and HUVECs+Exo(p62-), the HUVECs+Exo(p62-) group grew slower than the other two groups (p\u00a0<\u00a00.05). In summary, the above results indicated that the fastest angiogenesis occurred in the presence of exosomes from U937 cells; the slowest angiogenesis occurred in the presence of exosomes from p62-knockdown U937 cells.HUVECs were inoculated in Matrigel and incubated with AML exosomes for a certain period of time to determine whether exosomes from AML cells could induce HUVEC tubular differentiation nificant . Microscnificant \u20137C. By cAlthough advances have been made in AML supportive care, prognostic risk stratification, and established therapies, patients with AML have poor long-term prognosis . The ideAccumulating evidence indicates that exosomes in tumors are oncogenic. Crosstalk between bone marrow tumors and endothelial cells can affect tumor progression in hematological tumors, and exosomes containing miRNAs are crucial in bone marrow angiogenesis promotion in hematological tumors. By delivering miR-365, exosomes can mediate the horizontal transfer of drug resistance in chronic myeloid leukemia cells . K562 ceP\u00a0<\u00a00.05). We used microarray technology to study the expression patterns of exosomal miRNAs derived from two U937 cell lines to further explore the relationship between exosomal miRNAs and p62. We identified 2,080 miRNAs, including 215 upregulated and 208 downregulated miRNAs in p62-knockdown U937 cells. To further validate microarray analysis results, we performed RT\u2013qPCR to validate the downregulated expression of miR-3064-3p and miR-339-5p in the same series of samples. Our future studies will involve identification of other miRNAs whose expression was the highest in our microarray results. The RT\u2013qPCR results were consistent with those obtained from microarray analysis. In addition, KEGG pathway analysis revealed that 84 pathways corresponded to upregulated transcripts, and 55 pathways corresponded to downregulated transcripts. For the downregulated transcripts, the most affected pathway was the \u201cTNF signaling pathway\u201d (Pathway ID: hsa04668), followed by the \u201cMAPK pathway\u201d (Pathway ID: hsa04010). As for upregulated transcripts, the \u201cPI3K\u2013Akt signaling pathway\u201d (Pathway ID: hsa04151) was the most enriched pathway.In our study, p62 knockdown in U937 cells inhibited cell proliferation and promoted apoptosis , VEGFC, GATA-binding protein 4 , matrix Although AML cells secrete angiogenic factors to remodel the vascular system and gain chemoresistance, anti-angiogenic drugs are generally ineffective in AML treatment. In conclusion, after a detailed examination of miRNA expression in exosomes derived from AML cells, we found that hsa-miR-3064-3p and hsa-miR-339-5p displayed downregulated expression in p62-knockdown cells, compared with control cells. In addition, we demonstrated that several differentially expressed exosomal miRNAs were closely related to multiple GO items and pathways involved in carcinogenesis, indicating that exosomal miRNAs play a key role in AML pathogenesis. This information may aid the development of potential biomarkers for diagnosis and prognosis of AML progression. In the present study, we also found that exosomes derived from AML cells promoted angiogenesis. However, the relationship between exosomal miRNAs and angiogenesis needs to be further investigated. These findings support the notion that promising new treatment strategies may be developed against AML, based on exosomal miRNA analysis.10.7717/peerj.13498/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.13498/supp-2Supplemental Information 2Click here for additional data file.10.7717/peerj.13498/supp-3Supplemental Information 3Click here for additional data file.10.7717/peerj.13498/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.13498/supp-5Supplemental Information 5Click here for additional data file.10.7717/peerj.13498/supp-6Supplemental Information 6Click here for additional data file.10.7717/peerj.13498/supp-7Supplemental Information 7Click here for additional data file.10.7717/peerj.13498/supp-8Supplemental Information 8Click here for additional data file.10.7717/peerj.13498/supp-9Supplemental Information 9Click here for additional data file.10.7717/peerj.13498/supp-10Supplemental Information 10Click here for 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+{"text": "Microbacterium elymi KUDC0405T was isolated from the rhizosphere of Elymus tsukushiensis from the Dokdo Islands. The KUDC0405T strain was Gram-stain-positive, non-spore forming, non-motile, and facultatively anaerobic bacteria. Strain KUDC0405T was a rod-shaped bacterium with size dimensions of 0.3\u20130.4 \u00d7 0.7\u20130.8 \u03bcm. Based on 16S rRNA gene sequences, KUDC0405T was most closely related to Microbacterium bovistercoris NEAU-LLET (97.8%) and Microbacterium pseudoresistens CC-5209T (97.6%). The dDDH  values between KUDC0405T and M. bovistercoris NEAU-LLET and M. pseudoresistens CC-5209T were below 17.3% and 17.5%, respectively. The ANI (average nucleotide identity) values among strains KUDC0405T, M. bovistercoris NEAU-LLET, and M. pseudoresistens CC-5209T were 86.6% and 80.7%, respectively. The AAI (average amino acid identity) values were 64.66% and 64.97%, respectively, between KUDC0405T and its closest related type strains. The genome contained 3,596 CDCs, three rRNAs, 46 tRNAs, and three non-coding RNAs (ncRNAs). The genomic DNA GC content was 70.4%. The polar lipids included diphosphatydilglycerol, glycolipid, phosphatydilglycerol, and unknown phospholipid, and the major fatty acids were anteiso-C17:0 and iso-C16:0. Strain KUDC0405T contained MK-12 as the major menaquinone. Based on genotypic, phylogenetic, and phenotypic properties, strain KUDC0405T should be considered a novel species within the genus Microbacterium, for which we propose the name M. elymi sp. nov., and the type strain as KUDC0405T . Microbacterium was classified by Orla-Jensen S [Microbacterium comprised 157 species, including Microbacterium aerolatum V-73T [M. agarici CC-SBCK-209T [M. album SYSU D8007T [M. algeriense G1T [M. amylolyticum N5T [M. aoyamense KV-492T [M. aquimaris JS54-2T [Microbacterium can be isolated from various sources, such as seawater, desert soil, maize rhizosphere, cow dung, and microfiltered milk. Members of this genus are Gram-stain-positive, rod-shaped, and have an optimum growth temperature of 20\u201330\u00b0C. Here, we report a taxonomic analysis of the novel bacterial strain, KUDC0405T, isolated from the rhizospheric soil of Elymus tsukushiensis, a plant native to the Dokdo Islands . During microbial diversity monitoring in April 2014, rhizospheric soil samples were collected from native plants of the Ulleungdo and Dokdo Islands . E. tsukushiensis var. trasiens (Hack.) Osada is native to the Dokdo Islands and is the dominant plant species on these islands, and its distribution is expanding [Brevibacterium iodinum KUDC1716 [Ochrobactrum lupini KUDC1013 and Novosphingobium pentaromativorans KUDC1065 [The genus Jensen S  based onum V-73T , M. agarU D8007T , M. algeicum N5T , M. aoyaxpanding . DespiteKUDC1716 , Ochroba-4\u201210-6) were prepared. A 100 \u03bcL aliquot dilution was plated onto R2A and 1/10 diluted tryptic soy agar (TSA)  and incubated at 25\u00b0C for 7 days. Morphologically different colonies were selected, and individual colonies were further purified by repeated streaking onto TSA media. The type strains used in this study were obtained from the China General Microbiological Culture Collection Centre (CGMCC) and the Korea Collection for Type Cultures (KCTC). The strains were cultivated on TSA at 25\u00b0C and maintained at -70\u00b0C in saline solution  supplemented with 15% glycerol (v/v).Plant samples were collected and stored as described previously . The samhttp://macrogen.com/) using the sequencing primers (518F and 800R) and an automated sequencer . The EzBioCloud server (https://ezbiocloud.net/) [The phylogenetics of the isolated strain was determined based on a comparative analysis of the 16S rRNA gene sequence. The 16S rRNA gene sequence was amplified and the PCR products were purified as described previously . UniversRathayibacter rathayi VKM Ac-1601T, which is not affiliated with the genus Microbacterium was used as an outgroup. Trees were rooted and constructed using MEGA-X in the Newick format.All 16S rRNA gene sequences of the closest type strains were aligned using CLUSTAL_W  and the et al. [et al. [T was sequenced using a MinION platform . The reads were assembled de novo using Flye (version 2.9) [T and closely related strains were calculated using the JSpeciesWS website (https://jspecies.ribohost.com/jspeciesws/) [http://enve-omics.ce.gatech.edu/) [http://ggdc.dsmz.de/distcalc2.php) [T and its two closest relatives, M. bovistercoris NEAU-LLET and M. pseudoresistence CC-5209T, using protein sequences annotated by Hyatt et al. [http://automlst.ziemertlab.com) [Chromosomal DNA was extracted in accordance with Sambrook et al.  and was  [et al. . The celion 2.9) . The aution 2.9)  and the ion 2.9)  were useion 2.9) . The aveciesws/) . The avech.edu/) . The dDDlc2.php) . The dDDt et al.  and the t et al. . A multilab.com) .T was observed with a Zentech digital camera  for cell morphology and size, using cells grown on TSA. The cells were treated with 1% osmium tetroxide 25\u00b0C for 1 h, and dehydrated with graded series of ethanol , followed by isoamyl acetate. After lyophilization, the samples were coated with platinum , and the cell morphology was observed using a field emission scanning electron microscope .The scanning electron micrograph of strain KUDC0405T and the reference strains (M. bovistercoris NEAU-LLET and M. pseudoresistence CC-5209T) at different temperatures  and different pH values . The pH values were adjusted as described by K\u00e4mpfer et al. [Growth capability of strain KUDC0405r et al.  in steriM. bovistercoris NEAU-LLET and M. pseudoresistence CC-5209T, which are related to KUDC0405T, were analyzed under the same conditions. The cell wall peptidoglycan was analyzed using an amino acid analyzer . To analyze the polar lipids, two-dimensional thin layer chromatography (TLC) analysis were used according to Minnikin et al. [T, and reference strains were incubated on TSA at 30\u00b0C for 7 days. To determine siderophore production by strain KUDC0405T, chrome azurol S (CAS) media were used as previously described [To determine hydrolysis of starch, urea, Tween 20, 40, 60, and 80, the isolate was cultured on TSA at 30\u00b0C for a week, as described by Cowan and Steel . The enzn et al. . The fatescribed , 40.T  was determined as previously described [M. bovistercoris NEAU-LLET, followed by M. pseudoresistence CC-5209T (97.58%), M. resistens NBRC 103078T (97.51%), M. oleivorans NBRC 103075T (97.51%), M. testaceum NBRC 12675T (97.51%), and M. paraoxydans NBRC 103076T (97.30%). A comparison of the preliminary 16S rRNA gene sequences revealed that strain KUDC0405T is related to members of the genus Microbacterium. In the Bayesian inference tree . The complete genome of strain KUDC0405T consisted of a circular chromosome . The genomic DNA G+C content was 70.4%, which is within the range reported for the Microbacterium genus. A total of 3,654 genes were identified, of which 3,018 were protein-coding genes and 52 were RNA genes . T and closely related strains. The genome of strain KUDC0405T displayed 256 subsystems according to genome annotation using RAST. Various metabolic genes were predicted for various metabolic processes, such as the metabolism of amino acids and derivatives (311 genes), carbohydrates (247 genes), cofactors, vitamins, prosthetic groups, pigments (153 genes), proteins (151 genes), nucleosides and nucleotides (106 genes), DNA (60 genes), virulence, disease, and defense (38 genes), membrane transport (33 genes), respiration (33 genes), and other metabolic processes. KUDC0405T did not appear to be motile and the genome contained no genes encoding proteins associated with motility. The OrthoVenn diagram revealed orthologous protein clusters of strain KUDC0405T, M. bovistercoris NEAU-LLET, and M. pseudoresistens CC-5209T  shared 1,510 orthologous protein clusters , T3PKS1 , T3PKS2, RRE-containing , and terpene  . In the case of ANIm, <20% of the genome was aligned for M. bovistercoris NEAU-LLET, and the alignment was assigned as suspicious by the software. In silico, AAI values were analysed at 64.7% and 65.0% in strain KUDC0405T, M. bovistercoris NEAU-LLET, and strain KUDC0405T and M. pseudoresistence CC-5209T, respectively. Also, GGDC results for strains KUDC0405T, M. bovistercoris NEAU-LLET, and M. pseudoresistence CC-5209T were calculated as 17.3% and 17.5% based on formula 2 (identities/HSP length). The ANIb, ANIm, AAI, and dDDH values of strain KUDC0405T compared with those of the closely related strains are presented in T represents a novel species of the Microbacterium genus.The complete genomes determined have been deposited in the NCBI GenBank database under accession number GCF_021582895  . The terT was gram-positive, non-spore forming, non-motile, and grew anaerobically on TSA. Colonies on TSA media were smooth, circular, yellowish-white, and the cells were rod-shaped (0.3\u20130.4 \u00d7 0.7\u20130.8 \u03bcm)  . Growth f plants , 42. Theand urea .T was MK-12. The polar lipids included diphospharidylglycerol, glycolipid, phosphatidylglycerol, an unidentified phospholipid, three unidentified aminolipids, and an unidentified lipid , ornithine (26.8%), alanine (25.4%), and glutamic (15.4%) as cell-wall peptidoglycans. The major fatty acids in KUDC0405T were anteiso-C17:0 (35.2%), iso-C16:0 (16.3%), and iso-C17:0 (8.0%), and the minor components included iso-C15:0 (4.0%), C16:0 (2.6%), anteiso-C15:0 (2.5%), and C18:0 (1.5%). T and the most closely related reference strains. M. bovistercoris NEAU-LLET and M. pseudoresistence CC-5209T presented anteiso-C17:0 and anteiso-C15:0 as major fatty acids. The major fatty acids in the genus Microbacterium were anteiso-C17:0.The predominant menaquinone in KUDC0405ed lipid . DiphospCC-5209T . Strain T represents a novel species of the genus Microbacterium, for which we suggest the name M. elymi sp. nov..To conclude, we suggest that strain KUDC0405T produces siderophores and contains glycine, ornithine, alanine, and glutamic acid as cell-wall peptidoglycans. The polar lipids were diphosphatydilglycerol, glycolipid, phosphatydilglycerol, and phospholipid; the major menaquinone was MK-12; and the major fatty acids were anteiso-C17:0 and iso-C16:0. The genomic DNA GC content was 70.4%.Cells are Gram-stain-positive, catalase- and oxidase- positive, non-spore forming, non-motile, facultatively anaerobic and rod-shaped (0.3\u20130.4 \u00d7 0.7\u20130.8 \u03bcm). Colonies are smooth, circular, yellowish-white, and 3.0\u20130.4 mm in diameter on TSA with growth for 2 days. Optimal growth occurs at 25\u201330\u00b0C, at pH 7, and 0.5\u20131.2%NaCl (w/v) on TSA media. Strain KUDC0405T (=KCTC 49411T =CGMCC1.18472T), was isolated from the rhizosphere of E. tsukushiensis collected from the Dokdo Islands, Republic of Korea. The GenBank/EMBL/DDBJ accession numbers for the partial 16S rRNA gene sequence and genome sequence of KUDC0405T were MT071892 and CP091139, respectively. NCBI accession number for genomes are GCF_021582895.The type strain, KUDC0405de novo assembly are as follows: genome size, 3,610,832 bp; number of contigs, 1; coverage, 119.0 \u00d7.General features of the genome http://jmb.or.kr.Supplementary data for this paper are available on-line only at"}
+{"text": "Flavobacterium sediminilitoris YSM-43T, isolated from a tidal flat in Yeosu, Republic of Korea. The whole genome consists of one circular chromosome of 3,913,692\u2009bp. A total of 3,599 genes were predicted, comprising 3,537 coding DNA sequences (CDSs), 50 tRNAs, 9 rRNAs, and 3 noncoding RNAs (ncRNAs).Here, we report the complete genome sequence of  Flavobacterium, the type genus of the family Flavobacteriaceae in the phylum Bacteroidetes, is a Gram-negative, yellow-pigmented, and rod-shaped bacterium. Commonly, Flavobacterium thrives in various habitats, including both terrestrial and marine ecosystems (Flavobacterium species have been published (https://lpsn.dsmz.de/genus/flavobacterium)  . Accordicterium) , two Flailitoris , 5. F. sT was cultivated on marine agar 2216 (BD). The cells were collected in a 5-mL Eppendorf tube for DNA extraction. Extraction of genomic DNA was performed using a genomic DNA extraction kit (RBC), following the manufacturer\u2019s instructions. A spin column  was utilized for DNA cleanup. The NanoDrop 2000 UV-visible (UV-vis) spectrometer was used for measuring the ratio of absorbance at 260/280\u2009nm and 260/230\u2009nm. The genomic DNA (gDNA) concentration was measured using a Qubit double-stranded DNA (dsDNA) high-sensitivity (HS) assay kit (Invitrogen) with a Qubit 2.0 fluorometer. The quantity and size distribution of the purified gDNA were calculated using Agilent 2200 TapeStation software (A.01.05)  were assembled de novo using Flye version 2.8.3 with the parameter \u201casm-coverage 100\u201d (https://github.com/PacificBiosciences/pbbioconda). Then, the genome was rotated using the fixstart method in Circlator version 1.5.5  (YSM-43A.01.05) . For bio N50, 7,54\u2009bp weredata set . As a resee.ca/) . To assisee.ca/) , eggNOG-see.ca/)  (Table\u00a01F. sediminilitoris YSM-43T has been deposited at GenBank under the accession number CP090145. The raw data have been deposited in the SRA under the accession number SRR17867805.The complete genome sequence of"}
+{"text": "The medical novelty of COVID-19 requires a comprehensive study of its impact on various areas of human health, including mental health.To study the spectrum and severity of psychopathological disorders in previously healthy patients of different age groups who have had moderate and severe COVID-19 pneumonia.Immediately after stabilization of the physical condition, patients completed the Symptom Checklist-90-R, designed to assess 11 parameters: somatization (SOM), obsessive-compulsive (OS), interpersonal sensitivity (INT), depression (DEP), anxiety (ANX), hostility (HOS), phobic anxiety (PHOB), paranoid ideas (PAR), psychoticism (PSY). Patients with cognitive impairment were excluded.The study involved 148 patients aged from 26 to 84 years. In the general sample, psychopathological symptoms were detected mainly on the SOM, DEP, ANX, HOS scales. To a lesser extent - on the INT and PAR scales; were practically not determined on the PSY and PHOB scales. Most of the symptoms are significantly more intense in patients over 46 years old (n = 129) compared with the younger population . Older patients according to SOM revealed 1.23 points (IQR 0.5) versus 0.85 (IQR 0.7) among young people, DEP - 0.88 (IQR 0.44) vs. 0.47 (IQR 0.44), ANX - 0.66 (IQR 0.44) vs. 0.43 (IQR 0.29), OS - 0.55 (IQR 0.5) vs. 0.31 (IQR 0.25) and HOS - 0.46 (IQR 0.34) vs. 0.29 (IQR 0.09).Patients recovering from severe COVID-19 pneumonia require psychiatric evaluation and subsequent differentiated psychotherapeutic rehabilitation, especially for the age group over 46.No significant relationships."}
+{"text": "Imaging with positron emission tomography (PET) plays a crucial role in patient selection prior to radioligand therapy and subsequent molecular response assessment. The presented study aims to investigate the role of quantitative uptake parameters on baseline 68Gallium-PSMA-11 PET/CT imaging as to their association with lesion response to radioligand therapy at individual tumor sites. Special emphasis is placed on the utility of PSMA-uptake thresholds for pretherapeutic prediction of lesion response.The prostate-specific membrane antigen (PSMA), a transmembrane protein frequently present on prostate cancer cells, has gained considerable interest as a target for both molecular imaging and therapy. Patients with metastatic castration-resistant prostate cancer can be successfully treated by delivery of beta particle-emitting 177Lu]Lu-PSMA-617. This study aims to quantify lesion-based response to RLT in relation to pretreatment standard molecular imaging metrics derived from [68Ga]Ga-PSMA-11 PET/CT. Sixty-one patients with mCRPC underwent [68Ga]Ga-PSMA-11 PET/CT imaging before and after a median of 4 (IQR 2\u20136) RLT cycles. Maximum and mean standardized uptake values , as well as tumor-to-liver ratio (TLR), were assessed. A median of 12 (IQR 7\u201317) lesions was analyzed per patient, resulting in a total of 718 lesions. Lesions with \u226530% SUVmax decline or falling below the blood pool uptake were considered responsive; \u226530% SUVmax increase marked lesion progression. Additionally, 4-point visual scoring was performed according to E-PSMA consensus. In total, 550/718 (76.6%) lesions responded to RLT, including 389/507 (76.7%) bone metastases and 143/181 (79.0%) lymph node metastases. Baseline SUVmax, SUVmean, and TLR values were associated with lesion response by a moderate but significant correlation . For the classification of lesion progression based on baseline PSMA uptake, receiver operating characteristics (ROC) found SUVmax, SUVmean, and TLR to have comparable discriminatory value . Of 42 tumor sites with baseline uptake below the liver (V-score < 2), 19/42 (45.2%) were responsive, 9/42 (21.4%) were stable, and 14/42 (33.3%) showed progression, leaving liver uptake a threshold with low prognostic value for the identification of RLT-refractory lesions (PPV 33%). This was observed accordingly for various liver uptake-based thresholds, including TLR < 1.5, <2.0 with a PPV at 24%, 20%, respectively. Standard uptake parameters quantified by routine baseline [68Ga]Ga-PSMA-11 PET/CT are moderately associated with post-treatment lesion response to [177Lu]Lu-PSMA-617. Commonly applied liver-based uptake thresholds have limited value in predicting refractory lesions at individual tumor sites.Baseline uptake on prostate-specific membrane antigen (PSMA)-targeted imaging is a prerequisite for radioligand therapy (RLT) with [ Inclusion criteria mandated that patients receive [68Gallium was obtained from a 68Ge/68Ga radionuclide generator . [68Ga]Ga-PSMA-11 was administered by intravenous injection and target activity per patient was 1.8\u20132.5 MBq/kg body weight. Whole-body images (vertex to mid-thigh) were acquired 61 \u00b1 12 min after tracer injection and PET acquisition time was 4 min per bed position. CT data were acquired for attenuation correction and anatomical localization using an X-ray tube voltage of 130 kV with a modulated tube current . Acquisitions were carried out on a Biograph 6 PET/CT scanner , with decay, scatter, and attenuation correction performed in accordance with the procedure guidelines set out by the joint EANM and SNMMI consensus statement Lu-PSMA-617 was administered by slow intravenous injection over 30\u201360 s, preceded and followed by 1000 mL of saline infusion. RLT was performed as an inpatient procedure at the nuclear medicine therapy ward in accordance with radioprotection regulations. Six cycles with an activity of 7.4 GBq per cycle were intended; administered activities were modified in patients with potential risks for toxicity.Radiolabeling of PSMA-617 with n detail . [177Lu]s). Receiver operating characteristics (ROC) analysis was applied to determine the ability of baseline PET/CT metrics  to predict treatment lesion progression at individual sites. The area under the curve (AUC) was calculated for all lesions and separately for bone metastases and lymph node metastases. Site-specific cutoff values for the detection of PL were calculated using Youden\u2019s J statistic. Various liver-based thresholds were tested as to their discriminatory value for lesion progression. Odds ratios (OR) were calculated with 95% confidence intervals (CI). Statistical analyses were performed with SPSS  and GraphPad Prism . All tests were two-sided, with p-values < 0.05 denominating statistical significance.Results are presented as median with interquartile range (IQR) and mean \u00b1 standard deviation (SD) for continuous variables. Categorical variables are reported as frequencies with respective percentages. Comparison of means was performed by a paired t-test for intraindividual analysis or by using a Mann-Whitney U test if data were not normally distributed. Association of categorical parameters was analyzed using non-parametric rank correlation (Spearman\u2019s correlation coefficient denoted with r68Ga]Ga-PSMA-11 PET/CT imaging and subsequently underwent a median of 4 (IQR 3\u20136) cycles of [177Lu]Lu-PSMA-617 given with a mean treatment activity of 6.9 \u00b1 1.4 GBq per cycle. Cumulative activity per patient was 29.0 \u00b1 17.5 GBq. Of all patients, 31/61 (50.8%) showed \u226550% PSA decline 12 weeks after treatment initiation, while 15/61 (24.6%) showed PSA progression based on PCWG3 criteria (\u226525% PSA increase).Overall, 718 lesions were included in the analysis, consisting of 507 bone, 181 lymph node, 22 visceral, and 8 primary/locally recurrent sites in 61 patients with mCRPC (median age 72 [IQR 67\u201378] years). This corresponded to a median 12 (IQR 7\u201317) lesions per patient. Patient characteristics at baseline are detailed in max of 14.11 (IQR 8.25\u201323.01) and SUVmean of 8.72 (IQR 5.09\u201314.39); tumor-to-liver ratio (TLR) was 3.36 (IQR 1.98\u20135.70). Of all lesions, 550/718 (76.6%) responded to RLT, consisting of 389/507 (76.7%) bone metastases and 143/181 (79.0%) lymph node metastases. There was no significant difference in mean SUVmax decline in bone vs. lymph node metastases (p = 0.23). Responding lesions (RL) had significantly higher SUVmax, SUVmean, and TLR values at baseline than stable (SL) or progressive lesions (PL), with an SUVmax at 16.01 (RL), 10.82 (SL), 5.11 (PL) (p < 0.001), SUVmean at 9.88 (RL), 6.62 (SL), 3.18 (PL) (p < 0.001), and TLR at 3.89 (RL), 2.55 (SL), 1.36 (PL) (p < 0.001). This was observed in both lymph node and bone metastases ; the relationship is shown in Details on lesion characteristics are provided in tastases , Table 268Ga]Ga-PSMA-11 PET/CT imaging are provided in The course of all lesions per patient is depicted in max, SUVmean, and TLR values. The area under the curve (AUC) was comparable for SUVmax, SUVmean, and TLR with 0.85, 0.87, and 0.83, respectively , with the corresponding SUVmean and TLR thresholds at 4.85  and 1.76 . While liver-based thresholds allowed the stratification of lesions based on their response category in a balanced manner, their value is limited for prognostication, as shown for various liver-derived thresholds in ROC analysis was performed to classify lesion progression based on pretherapeutic SUVectively . Cutoff max decline \u2265 30%) or non-detectability; 28/42 (66.7%) lesions were non-progressive (i.e. SL/RL). Lesions declining to an uptake below the blood pool after RLT (V-score = 0) had lower uptake values at baseline than lesions showing posttherapeutic V-scores \u2265 1 with SUVmax 10.63 vs. 14.89 (p = 0.002), SUVmean 6.78 vs. 9.15 (p = 0.009), and TLR 2.71 vs. 3.48 (p < 0.001) (p = 0.09).In addition to semiquantitative PET measurements, visual scoring (V-score) was noted for all lesions. The V-score change from baseline to posttherapeutic PET assessment was registered, as depicted in < 0.001) B. A mino68Ga]Ga-PSMA-11 PET/CT at baseline are moderately associated with lesion response to [177Lu]Lu-PSMA-617 RLT. Low uptake lesions were more frequently subject to progression, whilst addressable by RLT in a significant fraction of cases. Baseline metrics and liver-based thresholds had only limited value for single-lesion response prediction.This study indicates that routine PET parameters assessed by [177Lu]Lu-PSMA-617 RLT have proven that a higher density of the transmembrane glycoprotein PSMA on prostate cancer cells is associated with increased ligand internalization and treatment efficacy Ga-PSMA-11 PET/CT are moderately associated with lesion response to [177Lu] Lu-PSMA-617. Lesions with uptake values below the liver uptake remain non-progressive in the majority of cases examined in this cohort. Non-dominant tumor sites with low PSMA expression should thus not preclude patients from undergoing RLT.Standard uptake parameters quantified by routine baseline ["}
+{"text": "Dear Editor,TET2 is one of the most frequently mutated genes in myeloid malignancies . Mor. MorTET2MPN Fig.  6]. Col. ColTET2Nup98-HoxD13 (NHD13) transgenic mouse model, in which ~30% of mice develop AML. Interestingly, TET2 levels were lower in c-kit+ bone marrow (BM) cells of leukemia-transformed NHD13 mice relative to age-matched NHD13 mice, which developed MDS exclusively  or corresponding control (Tet2fl/fl) mice with NHD13 mice and monitored leukemia development following poly(I:C) treatment on both genotypes  counts, splenomegaly and hyper-cellularity in BM, while age-matched NHD13/Tet2-WT mice exhibited only cytopenia . At that time point, neither genotype showed signs of leukemia  population relative to those of Tet2-WT NHD13 mice, whereas the Lin-c-kit+Sca-1+ (LSK) population was unchanged by Tet2 deletion  from pre-leukemic NHD13/Tet2-KO or corresponding control NHD13 mice into lethally-irradiated secondary recipients to assess leukemogenicity. As expected, NHD13/Tet2-KO cell transplantation increased the percentage of CD45.2+ cells and WBCs in peripheral blood (PB) relative to NHD13/Tet2-WT cells  from WT or Tet2-KO mice  pool.To define mechanisms underlying MDS progression, we evaluated the mia Fig. , but Tetice Fig. . Importaon Figs.  and S2E.-KO Fig. . Within ice Fig. . Moreovells Fig.  or that nes Fig. . In the D13 Fig. . Tet2-KOlls Fig. . We nextls Figs. . By 16 w PB Fig. . Notablynts Fig. . Moreoveice Fig.  and obseTet2 loss in HSPCs can lead to hypermutagenicity [+ cells from pre-leukemic NHD13/Tet2-KO vs. matched NHD13/Tet2-WT mice. Relative to NHD13/Tet2-WT mice, we observed 271 newly acquired alterations and 199 alterations with increased variant allele frequency  in NHD13/Tet2-KO mice  and ranked them based on association with AML prognosis (http://precog.stanford.edu)  and mutation ratio (mutation ratio >0.5)  promoted the greatest increase in colony and cell number of NHD13 c-kit+ cells  Table . Accordi.5) Fig. . Pairwisons Fig. . To assesay Fig. . In the lls Fig. . Howeverity Fig. , indicatARIH2 function in leukemogenesis, we retrospectively analyzed GEO datasets and observed lower ARIH2 expression associated with shorter survival in MDS and AML patients  did not, and cells harboring the mutant exhibited a growth advantage relative to ARIH2WT cells  compared to those with TET2-WT  or ascorbate . Relative to vehicle-treated NHD13/Tet2-KO recipients, ASC treatment in NHD13/Tet2-KO mice significantly decreased WBC counts and the frequency of c-kit+ BM cells  from two TET2 mutant high-risk MDS patients (Table + cells (Fig. Given that vitamin C treatment mimics effects of toration , we treals Figs.  and S6A.mia Fig. . Notably KD Fig. . Moreovency Fig. . Finallyts Table  showed tlls Fig. .In summary, our results indicate that TET2 activity prevents further transformation of MDS HSPCs by decreasing the occurrence of secondary mutations, and that pharmacological enhancement of TET activity may represent an optimal strategy to block MDS malignant transformation.Supplementary data"}
+{"text": "Continuous emergence of the Omicron variant, along with its subvariants, has caused an increasing number of infections, reinfections, and vaccine-breakthrough infections, seriously threatening human health. Recently, several new Omicron subvariants, such as BA.5, BA.2.75, BA.4.6, and BF.7, bearing distinct mutation profiles in their spike (S) proteins, have significantly increased their capacity to evade vaccine-induced immunity and have shown enhanced infectivity and transmissibility, quickly becoming dominant sublineages. In this study, we found the S proteins of these Omicron subvariants to have 2- to 4-fold more efficient membrane fusion kinetics than that of the original Omicron variant (BA.1), indicating that these novel Omicron subvariants might possess increased pathogenicity. We also identified that peptide-based pan-CoV fusion inhibitors, EK1 and EK1C4, showed equal efficacy against membrane fusion mediated by S proteins of the noted Omicron subvariants and infection by their pseudoviruses. Additionally, either immune sera induced by wild-type (WT) SARS-CoV-2 RBD-based vaccine or BA.2 convalescent sera showed potent synergism with EK1 against both WT SARS-CoV-2 and various Omicron subvariants, further suggesting that EK1-based fusion inhibitors are promising candidates for development as clinical antiviral agents against the currently circulating Omicron subvariants. In late 2021, the Omicron variant B.1.1.529/BA.1 was first identified in South Africa and quickly spread to many countries ,4. SubseDifferent Omicron subvariants contain distinct mutation profiles, especially in their S proteins, well known to play a crucial role in mediating viral infection. Compared with WT SARS-CoV-2 S protein, Omicron BA.1 contains more than 30 spike mutations. In particular, K417N, N440K, G446S, S477N, T478K, E484A, Q493R, G496S, Q498R, N501Y, and Y505H mutations are known to occur in the Omicron BA.1 receptor binding domain (RBD) . Such mutations significantly alter the conformation of RBD in S protein and, hence, seriously threaten clinical vaccine efficacy . BA.2.12in vivo [Previous studies reported that S-mediated fusogenicity plays an important role in viral pathogenesis . For exain vivo . On the in vivo . NeverthAfter receptor engagement, heptad repeat 1 (HR1) and 2 (HR2) regions in the S2 subunit of coronavirus S protein interact to form a six-helix bundle (6-HB) structure, which, in turn, drives viral fusion with and entry into the host cell . Our preIn the current study, we found that the recently emerging Omicron subvariants showed strengthened fusion kinetics, particularly BA.2.75, BA.4.6, BA.5, and BF.7, compared to that of BA.1, indicating that they possess increased pathogenicity. Although these circulating Omicron subvariants carry distinct mutation profiles in S protein, their HR1 functional domains remain conserved. Accordingly, we found that our EK1-based fusion inhibitors  maintainWe first evaluated the fusogenicity of different Omicron subvariants. To mimic authentic viral fusion with target cells, as mediated by S protein, their S proteins were expressed on the membrane surface of 293T-GFP cells, as effector cells (293T-S-GFP)  of 142.02, 172.45 and 65.84 nM, respectively, while EK1C4 showed even more efficacy with IC50s ranging from 3.67 nM to 5.07 nM . On BA.21/2: 24h) and potential oral bioavailability by targeting SARS-CoV-2 HR1 in S2 subunit and RBD in S1 subunit [To further assess the efficacy of EK1-based pan-CoV fusion inhibitors against Omicron subvariants, we developed lentivirus-based nonreplicative pseudovirus systems for these Omicron subvariants. Such constructs can effectively mimic the entry process of authentic virus and are, therefore, widely used to evaluate antiviral agents . Both EK subunit . At the  subunit . These f subunit  and IPB2 subunit , respectCurrently, more than 12 billion vaccine doses have been administered globally , includiSince both RBD and HR1 are key in mediating viral entry , it is rMore importantly, WT-RBD-immunized mouse sera showed weak inhibitory activity against BA.2.12.1 and BA.2.75 with 31.4% and 33.1% inhibition, respectively, as did EK1 (100 nM) with 33.2% and 30.6% inhibition, respectively. However, when combined, they showed increased efficacy to 62.3% inhibition on BA.2.12.1 and 63% inhibition against BA.2.75  alone showed moderate inhibitory activity against the BA.2.12.1 sublineage with 64.6% inhibition, but little efficacy against BA.2.75, BA.4/BA.5, BA.4.6, BF.7, and WT SARS-CoV-2 . However, BA2-convalescent sera, when combined with low-dose EK1 (100 nM), showed significantly improved efficacy against BA.2.12.1, BA.2.75, BA.4/BA.5, BA.4.6, and BF.7 in the range of 57.9% to 83.0% inhibition ; HeLa and Caco2 cell lines were from the Chinese Academy of Science Cell Bank . 293T/ACE2 cells were preserved in our laboratory. All cell lines were cultured in Dulbecco\u2019s Modified Eagle\u2019s Medium (DMEM) with 10% fetal bovine serum (FBS). Plasmids, including pAAV-SARS-CoV-2-S-D614G-IRES-EGFP, pAAV-SARS-CoV-2-S-Delta-IRES-EGFP, pAAV-SARS-CoV-2-S-Omicron-IRES-EGFP, and PC-hACE2/horse_ACE2/cattle_ACE2/swine_ACE2/rabbit_ACE2/civet_ACE2/ bat_ACE2, were synthesized or preserved in our laboratory. Peripheral blood samples were collected from a convalescent BA.2 patient. Serum sample was isolated from centrifuged blood sample for inhibiting pseudovirus infection with 1:500 dilution. All collections were conducted according to the guidelines of the Declaration of Helsinki and approved by the Institutional Review Board of the Ethics Committee of Shanghai Fourth People's Hospital (2022095-001).Briefly, WT SARS-CoV-2 RBD-Fc (5 \u00b5g) formulated with an equal volume of Imject Alum adjuvant (Thermo Scientific) was used to vaccinate Balb/c mice (six-week-old) three times at two-week intervals, as previously described ,35. At tWestern blot was performed using an anti-SARS-CoV-2 S antibody and an anti-actin antibody, as previously described . BrieflyPlasmid pAAV-IRES-S-EGFP, encoding S protein and EGFP, was transfected into 293T effector cells (293T/S/GFP). Caco2 cells, naturally expressing human ACE2 receptors on the membrane surface, were used as target cells. 293T cells, transfected with plasmid pAAV-IRES-EGFP (293T/EGFP), were used as negative control. Effector cells (293T/S/GFP) were collected and resuspended. Free effector cells were added into target cells  for coincubation for indicated time at 37 \u00b0C and then observation under the fluorescence microscope.HIV-1 backbone-based Pseudovirus (PsV) bearing wild or mutant SARS-CoV2 S protein was produced, as previously described . Caco2 ct-test and ANOVA test. P values less than 0.05 were significant; **P\u2009<\u20090.01; ***P\u2009<\u20090.001. The concentration for half inhibition (IC50) was calculated by CalcuSyn software [Statistical analyses were carried out using GraphPad Prism 8.0. Analyses of independent data were carried out through Student\u2019s unpaired two-tailed software .Click here for additional data file."}
+{"text": "In 2019, the World Health Organization (WHO) endorsed thermal ablation (TA) for use within \u201cscreen-and-treat\u201d cervical cancer prevention programs in low- and middle-income countries (LMICs), including among women living with HIV (WLWH). We evaluated TA efficacy for treatment of biopsy-confirmed cervical intraepithelial neoplasia grades 2 and 3 (CIN2/3) among WLWH in western Kenya .Between August 2019 and November 2020, WLWH age 25-65 years underwent high-risk human papillomavirus (hrHPV) self-collection. hrHPV-positive women underwent colposcopy-directed biopsies, and thermal ablation treatment if eligible per WHO guidelines. Women with biopsy-confirmed CIN2/3 had colposcopy-directed biopsies at 12-months to determine treatment efficacy.Sixty-eight hrHPV-positive WLWH with biopsy-confirmed CIN2/3 at baseline; 14 CIN2, 54 CIN3, underwent thermal ablation. Mean age and parity were 41.2 years and 4, respectively. The mean CD4 count was 473.98 cells/mm3 and 96.9% had HIV viral suppression. Fifty-eight women (83.8%) have been seen for a 12-month follow-up visit, and pathology results are available for 54 (79.4%). Of these, 35 (66.0%) had successful treatment, defined as biopsy-confirmed CIN1 or normal findings 12-months following treatment, while 18 (34.0%) had treatment failure - persistent biopsy-confirmed CIN2/3. Treatment failure was 23.1% 95% CI (13.0 to 45.9) and 37.5%, 95% CI (22.1 to 52.0) among women with CIN2 and CIN3 at baseline, respectively.Hand-held thermal ablation devices are affordable, portable, easy to use, and hence highly scaleable within screen-and-treat programs in LMICs. However, our preliminary results, with rigorous disease status verification at both baseline and follow-up find higher than previously reported treatment failure rates for CIN3 among WLWH, a high-risk population for cervical cancer. If replicated by larger studies, this highlights a potential limitation of the current WHO cervical cancer elimination strategy, calling for better risk stratification in this population, and/or consideration of adjuvant therapy to prevent CIN2/3 recurrence following thermal ablation."}
+{"text": "Dysregulation of epigenetic mechanisms have been depicted in several pathological consequence such as cancer. Different modes of epigenetic regulation (DNA methylation (hypomethylation or hypermethylation of promotor), histone modifications, abnormal expression of microRNAs (miRNAs), long non-coding RNAs, and small nucleolar RNAs), are discovered. Particularly, lncRNAs are known to exert pivot roles in different types of cancer including breast cancer. LncRNAs with oncogenic and tumour suppressive potential are reported. Differentially expressed lncRNAs contribute a remarkable role in the development of primary and acquired resistance for radiotherapy, endocrine therapy, immunotherapy, and targeted therapy. A wide range of molecular subtype specific lncRNAs have been assessed in breast cancer research. A number of studies have also shown that lncRNAs may be clinically used as non-invasive diagnostic biomarkers for early detection of breast cancer. Such molecular biomarkers have also been found in cancer stem cells of breast tumours. The objectives of the present review are to summarize the important roles of oncogenic and tumour suppressive lncRNAs for the early diagnosis of breast cancer, metastatic potential, and chemotherapy resistance across the molecular subtypes. Epigenetic dysregulations have a crucial impact on the development and progression of human cancers, including breast cancer . Epigenep = 0.003) in malignant samples could separate it breast cancer samples from normal control  and correlated with advanced TNM stage (p = 0.002), poorer pathological differentiation (p = 0.004), and shorter overall survival (SOS) and disease-free survival (DFS) (p < 0.0001) in progesterone receptor (PR)  positive cancer tissues (p < 0.00001). Moreover, validation using gene expression omnibus data sets  and 100 breast cancer patients confirmed similar results  . A meta- tissues . Moreove results . Based o results . Higher  results . Additio results . Chen et results . Further results .via the regulation of SNCG (Synuclein Gamma) expression  during cancer progression. LncRNA HOXD-AS1 interacts with miR-421 and inhibits its expression leading to the upregulation of SOX4, a master regulator of EMT . Similarpression . High expression . Zheng epression . In addipression . Also, hpression . Si et apression . Further-5p axis .via increasing Checkpoint kinase 2 (CHK2) phosphorylation , PARP (cleaved-Caspase-3 and cleaved-poly adenosine diphosphate-ribose polymerase) had elevated expression in low expression lncRNA BANCR group . Liu et  pathway . Elevate pathway . Further pathway . The exp pathway .It was observed that lncRNAs can also regulate several cancers associated signalling pathways, including the activation of transcription factors, such as nuclear factor kappa B (NF-\u03baB). For example, overexpressed lncRNA NKILA bound to NF-\u03baB/I\u0138B masked its phosphorylation. This interaction prevented the over-activation of the NF-\u03baB pathway in inflammation stimulated breast epithelial cells . Accordivia modulation of miR-200b/axis/Wnt/\u03b2-catenin pathway  signalling pathway . Dysreguviz., lncRNA YIYA regulates CDK6 (cell division protein kinase 6) dependent phosphorylation of PFKFB3 (fructose bis-phosphatase PFK2), and thus can convert glucose 6-phosphate (G6P) to fructose-2,6-phosphate  gene in breast cancer (via modulating the expression of miR-4766-5p and SIRT1 (Sirtuin 1) genes (in situ hybridization) and Western blot assays in MCF-7 and MDA-MB-231 breast cancer cell. Luciferase reporter assay validated that RHPN1-AS1 inhibits miR-4261 and regulates the direct transcriptional target of c-Myc (Immunoprecipitation assays provided evidence that lncRNA H19 regulated the expression of t cancer . The res1) genes  Higher eof c-Myc .p = 0.0086)  . Huang a 0.0086) . Another systems . Accordi systems .Oncogenic lncRNA LINC02163 was found to be involved in breast cancer pathogenesis by mode of LINC02163/miR-511-3p/HMGA2 (high mobility group A proteins 2) axis . The finLi and et al. found that upregulation of lncRNA ZFHX4-AS1 suppresses FAT4 and increases YAP1 (yes-associated protein 1) and TAZ (Tafazzin) gene expression which is attributed to breast cancer cell proliferation . A reporp = 0.022), lymph node metastasis (p = 0.020), and higher Ki-67 positivity (p = 0.017). A multivariate analysis suggested that a low level of lncRNA EGOT acts as an independent prognostic factor for poor survival rate in breast cancer patients  (There are a number of lncRNA whose downregulation contributes in breast cancer development and progression . A meta-= 0.039) . Further= 0.039) . Yang et= 0.039) . Low relvia modulating expression of cyclin A2, CDK2, p-Akt, p-p44/42 MAPK, and p-p38 MAPK cancer signalling molecules (in-vivo and in-vitro model system induced the EMT process in cancer via regulation of PI3K (phosphatidylinositide-3 kinase)/Akt pathways. Therefore, MALAT1 may act as a promising therapeutic target for breast cancer metastasis via the PI3K-Akt pathway  expression. Thus, the results suggested that lncRNA MAGI2-AS3 has the potential to serve as an anticancer therapeutic candidate . Tumour olecules . Similarolecules . LncRNA olecules . Downreg pathway .A study by via the miR-190b-5p/MYLIP (Myosin regulatory light chain interacting protein) axis, thus providing evidence for potential therapeutic targets for breast cancer patients  and its binding with co-activator YAP1, leading to reduced metastatic ability . Anotherpatients . On the patients .in situ (DCIS) breast cells  identified lncRNA LINC00885 expression in both normal and ductal carcinoma st cells . Expressst cells . Overexpst cells . Similarst cells . Furtherst cells . In addist cells .Gene expression profiling deciphered the breast cancer into four distinct molecular subtypes such as Luminal, Her2+, Her2 enriched, TNBC, and basal like. Patients with same molecular subtype responded differently to targeted therapy and showed diverse clinical outcomes. However, the exact underlying mechanism for molecular heterogeneity remains to be elucidated. Many researchers have evaluated molecular subtype specific lncRNAs expression in breast cancers and suggested its involvement in cancer molecular heterogeneity . Computain-vivo and in-vitro systems  gene expression restoration . Further systems . Another systems . Greater systems . Presenco ELAVL1 . Beltr\u00e1no ELAVL1 . Higher o ELAVL1 . By asseo ELAVL1 . Another pathway . Wang et pathway . RNA imm pathway . Further pathway .via targeting miR-205 , and ALDH1 (Aldehyde dehydrogenase 1) marker\u2019s expression . Similar miR-205 . Microar miR-205 . LncRNAs miR-205 .Chemotherapy resistance is the major cause of cancer related deaths. LncRNAs have a key role in developing resistance against radiotherapy, chemotherapy, immunotherapy, and targeted therapy. Inhibition of lncRNA LINC02582 expression increased radiosensitivity miR-200c/LINC02582/CHK1 in breast cancer samples . LncRNA via the H19/SAHH/DNMT3\u00df (DNA (cytosine-5)-methyltransferase 3 beta) axis, contributed to tamoxifen resistance in breast cancer  . BermejoLong ncRNAs like H19 and XIST were discovered in the pre-genomic era, but were not fully characterized and explored until the early 2000s . Inventi"}
+{"text": "Sotrovimab, a monoclonal antibody with efficacy against SARS-CoV-2 including certain Omicron variants, has been used in treatment of mild-moderate COVID-19. Limited data exists regarding its use in pregnant women.Electronic medical record review of pregnant COVID-19 patients treated with sotrovimab from 12/30/21-1/31/22  was performed. Included were pregnant individuals \u2265 12 years, weighing \u2265 40 kg, with positive SARS-CoV-2 test (within 10 days). Those receiving care outside YNHHS or receiving other SARS-CoV-2 treatment were excluded. We assessed demographics, medical history, and Monoclonal Antibody Screening Score (MASS). Clinical outcomes assessed included emergency department (ED) visit < 24 hours, hospitalization, ICU admission, and/or death within 29 days of sotrovimab. Pregnancy and neonatal outcomes were assessed until 8/15/22.2. Sixty-three percent were Caucasian, 9% Hispanic, 14% African-American, and 9% Asian. Nine percent had diabetes and sickle cell disease. Five percent had well-controlled HIV. Eighteen percent, 46%, and 36% received sotrovimab in trimester 1, 2, and 3, respectively. No infusion/allergic reactions occurred. MASS values were < 4. Only 12/22 (55%) received complete primary vaccination ; none received a booster.Among 22 subjects, median age was 32 years and body mass index was 27 kg/mThere were no ICU admissions nor deaths. One subject was hospitalized for post-partum pyelonephritis; another had an ED visit for post-partum vaginal bleeding. Median gestational age at birth was 38.9 weeks. Nine percent had premature labor and premature rupture of membranes, respectively. Median infant birth weight was 3220 g. One neonate required an ICU stay due to prenatally diagnosed omphalocele (before sotrovimab) in a mother with congenital defect history. There were no abortions, fetal loss, or other birth/neurodevelopmental defects.Pregnant COVID-19 patients receiving sotrovimab at our center tolerated it well with good clinical outcomes. Pregnancy and neonatal complications did not appear sotrovimab-related. Though a limited sample, our data helps elucidate the safety and tolerability of sotrovimab in pregnant women.All Authors: No reported disclosures."}
+{"text": "Waning protection against emerging SARS-CoV-2 variants by pre-existing antibodies elicited because of current vaccination or natural infection is a global concern. Whether this is due to the waning of immunity to SARS-COV-2 remains unclear.We aimed to investigate the dynamics of antibody isotype responses amongst vaccinated na\u00efve (VN) and naturally infected (NI) individuals.n\u2009=\u2009100) in two phases: phase-I (P-I) at ~\u20091.4 and phase-II (P-II) at ~\u20095.3\u00a0months. Antibody levels were compared with those of unvaccinated and naturally infected subjects  at ~\u20091.7 (P-1) and 5.2 (P-II) months post-infection. Neutralizing antibodies (NTAb), anti-S-RBD-IgG, -IgM and anti-S-IgA isotypes were measured.We followed up antibody levels in COVID-19 messenger RNA (mRNA)-vaccinated subjects without prior infection  than the NI group at P-I, except for IgM. In the VN group, a significant waning in antibody response was observed in all isotypes. There was about an ~ 4-fold decline in NTAb levels (P\u2009<\u20090.001), anti-S-RBD-IgG , anti-S-RBD-IgM  and anti-S1-IgA . In the NI group, a significant but less steady decline was notable in S-RBD-IgM , and a much smaller but significant difference in NTAb  anti-S-RBD IgG . Unlike the VN group, the NI group mounted a lasting anti-S1-IgA response with no significant decline. Anti-S1-IgA, which were\u2009~\u20093-fold higher in VN subjects compared with NI in P-1 (P\u2009<\u20090.001), dropped to almost the same levels, with no significant difference observed between the two groups in P-II.The VN group elicited significantly greater antibody responses (Whereas double-dose mRNA vaccination boosted antibody levels, vaccinated individuals\u2019 \u2018boost\u2019 was relatively short-lived. The recent Coronavirus disease 19 (COVID-19) outbreak caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) has put the globe in an emergency state.,On 21 December 2020, Qatar initiated a mass COVID-19 vaccination programme, first utilizing the BNT162b2 vaccine and then the mRNA-1273 (Moderna) vaccine.,,When vaccination was ramped up, the country had two back-to-back waves from January to June 2021, mainly dominated by the B.1.1.7  and B.1.351 (or beta) variants.According to current evidence, patients vaccinated against COVID-19 would lose around half of their neutralizing antibodies (NTAb) ~\u2009108\u00a0days post-vaccination.Although vaccine-induced immunity is being extensively studied, the body of evidence for infection-induced immunity is very limited and insufficient to establish an antibody titre threshold that shows whether a person is protected from infection.n\u2009=\u2009100) who had received two doses of the two approved mRNA vaccines in Qatar: BNT162b2 and mRNA-1273. Levels were compared with those of unvaccinated but NI subjects (n\u2009=\u200940). Venous blood samples were collected from vaccines in two phases: phase 1 (P-I) and phase (P-II). For the VN group, P-I samples were collected at 1.4\u00a0months (median\u2009=\u20096\u00a0weeks) and P-II samples were collected at 5.3\u00a0months (median\u2009=\u200923\u00a0weeks) after the second dose. For the NI group, P-I samples were collected at ~\u20091.7\u00a0months (median\u2009=\u20097\u00a0weeks) and P-II samples were collected at 5.2\u00a0months (median\u2009=\u200922\u00a0weeks) post-infection with SARS-CoV-2.The study was reviewed and approved by the Institutional Review Board at Qatar University (QU-IRB 1537-FBA/21). The study\u00a0included VN participants , i\u2009\u2265\u20091.00 Positive (IgM antibodies to SARS-CoV-2 detected). Architect automated chemiluminescent assay  was used to test the samples for the previous infection by measuring the SARS-CoV-2 anti-nucleoprotein IgG antibodies (anti-N), considering that the IgG antibodies produced against the RBD on the spike protein are different from the IgG antibodies produced against the nucleoprotein of the virus. Therefore, the positive anti-N results of SARS-CoV-2 anti-nucleoprotein IgG antibodies indicate previous exposure to the whole virus.After blood collection, plasma was separated by centrifugation and heat-inactivated. Serological testings were performed using the automated analyser CL-900i\u00ae from Mindray Biomedical Electronics 20-22 to detect: (1) NTAb  were measured using the automated analyser CL-900i\u00ae. This assay is a competitive binding chemiluminescent immunoassay for the quantitative determination of SARS-CoV-2 NTAb that blocks the interaction between the receptor binding domain (RBD) of viral spike protein (bound on magnetic beads) and the enzyme-conjugated ACE2 surface receptor. Phosphate-buffered saline (PBS) was used to dilute the samples with readings higher than the mentioned range. The assay has a WHO conversion factor of 1\u00a0AU\u2009=\u20093.31\u00a0IU/mL, and the reference range is 10\u00a0AU/mL to 400\u00a0AU/ml. We recently evaluated this novel assay and reported high specificity and sensitivity against two reference methods.P-values were two-sided at a significance level of 0.05.GraphPad Prism software  was used to perform the statistical analysis. Continuous variables were summarized using geometric means and 95% confidence intervals (95% CIs). The collected dataset was subjected to the Shapiro\u2013Wilk normality test to evaluate the data\u2019s normality. Because of the absence of normal distribution, nonparametric tests were performed using Wilcoxon rank-sum test for pairwise group comparisons and Mann\u2013Whitney U to test for the differences between independent samples. In the different scatter plots, the central horizontal bar line shows the geometric mean titre, and the error bars show the 95% CIs. All n\u2009=\u2009100) had no previous history of infection  and received two doses of either BNT16b2 or mRNA-1273. After receiving the second dose in P-I and P-II, the median weeks were 6 (1.4\u00a0months) and 23 (5.3\u00a0months) weeks, respectively (n\u2009=\u200940) were unvaccinated COVID-19-recovered individuals. In P-I 1 and P-II, the median weeks were 7 (~ 1.7\u00a0months) and 22 (5.2\u00a0months) weeks post-COVID-19 infection, respectively. The NI group comprised 20% females and 80% males  and 26% (12/47), respectively. In P-II, 100% remained positive for anti-S-RBD IgG and 99% (99/100) were positive for NTAb antibodies. The anti-S1-IgA positivity rate dropped to 92% (86/93), and only 2% (1/47) remained positive for the IgM.Amongst the NI participants in P-1, the positivity rate for NTAb antibodies was 90% (35/39). For anti-S-RBD-IgG, IgM and IgA, the positivity rates were 100 (31/31), 43 (17/40) and 72% (23/32), respectively. In P-II, 90% (35/39) were positive for NTAb antibodies. However, the positivity rates dropped to 97 (30/31), 13 (5/39) and 66 (21/32) for anti-S-RBD-IgG, -IgM and -IgA, respectively.P\u2009<\u20090.001) in the levels of NTAb antibodies was observed, from the geometric mean 1328.4 (95% CI: 1086.5\u20131624.2) to 317.94\u00a0IU/mL (95% CI: 261.5\u2013386.5\u00a0IU/mL) (P\u2009<\u20090.001) was observed in the levels of anti-S-RBD-IgG antibody levels from the geometric mean 1950.93 (95% CI: 1617.5\u20132353.1) to 366.107 BAU/mL (95% CI: 294.4\u2013455.20) (P\u2009<\u20090.001) from the geometric mean 0.56 (95% CI: 0.4\u20130.8) to 0.09 (95% CI: 0.04\u20130.25) (P\u2009<\u20090.0001) from the geometric mean 6.30 (95% CI: 5.4\u20137.3) to 3.19 (95% CI: 2.6\u20133.9)  . A signi\u2013455.20) . IgM dec04\u20130.25) . The lev2.6\u20133.9) .P\u2009<\u20090.001), from the geometric mean 157.3\u00a0IU/mL, 95% CI: 108.7\u2013227.7 to 107.0\u00a0IU/mL, 95% CI: 80.42\u2013142.3 (P\u2009<\u20090.001) from the geometric mean 66.54 (95% CI: 36.81\u2013120.3) to 58.65 BAU/mL (95% CI: 37.06\u201392.80) (p\u2009=\u20090.005) (P\u2009<\u20090.001)  . IgM dec<\u20090.001) . No sign<\u20090.001) .,To the best of our knowledge, this is the first study to comprehensively evaluate the levels of SARS-CoV-2 neutralizing, anti-S-RBD-IgG, Anti-S-RBD-IgM and anti-S1-IgA antibodies in VN and unvaccinated NI individuals. In the current study, mRNA vaccination elicited significantly greater NTAb, anti-S-RBD- IgG and anti-S1-IgA, compared with natural immunity . These r,,However, despite the mRNA vaccination-boosted antibody levels, this \u2018boost\u2019 was relatively short-lived, with ~\u20092- to 6-fold significant waning observed in NTAb, anti-S-RBD-IgG, anti-S-RBD-IgM and anti-S1-IgA, 23\u00a0weeks post-full-vaccination . These rIn comparison with mRNA vaccine-induced immunity, natural infection elicited a significant but less steady ~\u20091\u20132-fold decline in NTAb, anti-S-RBD-IgG, and anti-S-RBD-IgM . InteresThese findings are similar to a recent report showing that natural infection exhibit a lasting IgA response.,,,This study had several limitations. It is known that recent variants, such as Omicron, need many folds of antibody titre to be neutralized compared to the original strains. That is, the mutations that emerged in different variants of the SARS-CoV-2 genome should have a significant influence on viral protein structures, shape, function and immunogenicity, which, in theory, should greatly affect the strength and the effectiveness of the immunological response and possibly duration of antibody\u2019s waning in infected patients. It would be great to study the antibody response and waning against different variants. However, it was not applicable to include variant-specific data because of the limited number of samples and lack of sequencing data. In addition, a variety of variables could influence the level of immune response elicited after infection. It should be noted that our NI group included only 45% symptomatic subjects, whereas the remaining were paucisymptomatic (20%), asymptomatic (20%) or with unspecified severity (15%), which could have affected our results. In those with more severe COVID-19, NTAb antibody titers were reported to rise faster and reach a greater peak.,Furthermore, several studies have shown a link between cycle threshold (Ct) and antibody titre, with lower Ct values linked with greater antibody titers at the population level.Despite these limitations, this study has several strengths that merit attention. First, most of the published studies have mainly focused on NTAb, IgG or IgM, whereas studies on IgA response are minimal, particularly amongst unvaccinated NI subjects. Second, in this study, we assessed anti-N antibodies, which is crucial to identify those who were exposed to a virus but were asymptomatic prior to vaccination, especially amongst those vaccinated with vaccines containing only S protein. In addition, despite the relatively small sample size across the analysed groups, we utilized strict inclusion criteria and included participants from a wide age range to achieve valid comparisons.Our findings provide important insights into the durability of vaccine- and natural infection-induced immunity. We evaluated the antibody responses of NTAb, anti-SRBD IgM, anti-S1-IgA and anti-SRBD IgG antibodies. Whereas double-dose mRNA vaccination elicited higher antibody titers compared with natural infection, this \u2018boost\u2019 was relatively short-lived in vaccinated individuals. In contrast, natural infection exhibited a less steady decline in NTAb antibodies, IgG, IgM and a lasting IgA response. Understanding the degree of waning immunity is crucial for policymaking, particularly regarding vaccination strategies, supporting the consideration of booster doses to sustain protection against COVID-19.Conceptualization, H.A.S., H.M.Y., L.J.A.-R. and G.K.N.; methodology, H.A.S., B.A.H., S.Y. and F.M.S., software, B.A.H., S.Y. and F.M.S; validation, H.A.S., B.A.H., S.Y. and F.M.S.; formal analysis, H.A.S., G.K.N., B.A.H. and S.Y.; Investigation, H.A.S., A.I., H.M.Y., L.J.A.-R. and G.K.N.; resources, H.A.S., N.L., H.Q., N.A.D., H.M.Y., L.J.A.-R and G.K.N.; data curation, H.A.S., B.A.H., S.Y. and G.K.N.; writing\u2014original draft preparation, H.A.S., S.Y. and G.K.N.; writing\u2014review and editing, H.A.S., S.Y., N.Y., D.W.A., N.A.D., H.M.Y., L.J.A.-R and G.K.N; visualization, H.A.S., B.A.H., S.Y., N.Y., D.W.A., H.M.Y., L.J.A.-R A.I. and G.K.N; supervision, H.A.-S., H.M.Y. and G.K.N.; project administration, H.A.S., H.M.Y., L.J.A.-R and G.K.N.; funding, G.K.N. All authors have read and agreed to the published version of the manuscript."}
+{"text": "Streoptomyces rimosus M527 is a producer of the polyene macrolide rimocidin which shows activity against various plant pathogenic fungi. Notably, the regulatory mechanisms underlying rimocidin biosynthesis are yet to be elucidated.rimR2, which located in the rimocidin biosynthetic gene cluster, was first found and identified as a larger ATP-binding regulators of the LuxR family (LAL) subfamily regulator. The rimR2 deletion and complementation assays were conducted to explore its role. Mutant M527-\u0394rimR2 lost its ability to produce rimocidin. Complementation of M527-\u0394rimR2 restored rimocidin production. The five recombinant strains, M527-ER, M527-KR, M527-21R, M527-57R, and M527-NR, were constructed by overexpressing rimR2 gene using the promoters permE*, kasOp*, SPL21, SPL57, and its native promoter, respectively, to improve rimocidin production. M527-KR, M527-NR, and M527-ER exhibited 81.8%, 68.1%, and 54.5% more rimocidin production, respectively, than the wild-type (WT) strain, while recombinant strains M527-21R and M527-57R exhibited no obvious differences in rimocidin production compared with the WT strain. RT-PCR assays revealed that the transcriptional levels of the rim genes were consistent with the changes in rimocidin production in the recombinant strains. Using electrophoretic mobility shift assays, we confirmed that RimR2 can bind to the promoter regions of rimA and rimC.In this study, using domain structure and amino acid alignment and phylogenetic tree construction, rim genes and binding to the promoter regions of rimA and rimC.A LAL regulator RimR2 was identified as a positive specific-pathway regulator of rimocidin biosynthesis in M527. RimR2 regulates the rimocidin biosynthesis by influencing the transcriptional levels of The online version contains supplementary material available at 10.1186/s12934-023-02039-9. Streptomyces, a genus of Gram-positive bacteria with three types of PKSs , is best known for producing polyketides [Fusarium oxysporum f. sp. cucumerinum [Polyketides, a large group of secondary metabolites synthesized by polyketide synthases (PKSs), exhibit various bioactivities, including antifungal (rimocidin), antibacterial (penicillin), antitumor (daunorubicin) properties \u20133. They yketides \u20136. Polyeyketides \u20139, They yketides , nystatiyketides , amphoteyketides , and rimyketides . For exaumerinum , is a prStreptomyces causes bottlenecks, leading to low production levels and long fermentation periods [Streptomyces is a complex process involving multiple levels [However, polyketide biosynthesis in  periods . Secondae levels \u201318, incle levels \u201321.Streptomyces antibiotic regulatory protein (SARP) family regulator, such as ActII-orf4, which regulates actinorhodin biosynthesis, and CcaR, which regulates clavulanic biosynthesis. These regulators are characterized by the presence of OmpR -like DNA-binding domains [Streptomyces natalensis [Streptomyces nodosus [Streptomyces noursei [Streptomyces ahygroscopicus [S. noursei ATCC 11,455 [S. natalensis [Streptmyces avermilitis [To date, the different types of regulators involved in polyene macrolide biosynthesis have been categorized as follows: (1)  domains . (2) PAS domains , 24. Thetalensis . AmphRIV nodosus , and Nys noursei . (3) Lar noursei  having ascopicus , and NysC 11,455 . (4) SARtalensis , and Ptermilitis . Recent rmilitis \u201335.Streptomyces rimosus M527, a rimocidin producer, was originally isolated by Lu et al. [S. rimosus M527. However, to the best of our knowledge, no pathway-specific rimocidin biosynthesis regulators are currently known.u et al.  and depou et al. , fermentu et al. , were apStreptomyces diastaticus var. 108 has been predicted, and its biosynthetic gene cluster has been published (GenBank Accession No. AY442225) [S. rimosus M527 was sequenced (GenBank Accession No: NZ_SADA00000000.1), a biosynthetic gene cluster responsible for rimocidin production (GenBank Accession No. MK300953) , rimR2-deleted and rimR2-complemented strains. Subsequently, rimR2 gene was overexpressed using different promoters  to improve rimocidin production. Furthermore, the regulatory mechanism of RimR2 was identified using electrophoretic mobility shift assays (EMSA).Although the rimocidin biosynthetic pathway in Y442225) , no pathS. rimosus M527 genome sequence, rimR2 gene (2757 nucleotides (nt)), located in the rimocidin biosynthesis gene cluster, encodes a protein with a predicted molecular mass of 97.3\u00a0kDa consisting of 918 aa. RimR2 protein contains a conserved nucleotide phosphate-binding domain  and an HTH DNA-binding domain   method  analysis revealed that the mutant M527-\u0394rimR2 could not produce any rimocidin, whereas a distinct rimocidin peak was clearly observed in the WT culture filtrates  located in the gene cluster. The mutant M527-\u0394rimR2 exhibited significantly lesser transcriptional levels of all the candidate rim genes than the WT strain  was performed to examine the effects of ain Fig.\u00a0, and therim genes. In this experiment, His6-tagged RimR2 protein was generated in E. coli BL21 (DE3) , showing an 81.8% increase compared with the WT strain (207.2\u00a0mg/l) , M527-R3, and M527-R4 were determined via a shake-flask experiment  is the consensus nucleotide sequence of the binding site of PAS-LuxR regulators has been revealed [S. rimosus M527 and found three matches similar to the sixteen conserved nucleotide sequences: (1) CTAGGGAATTCCCGAG, which was the most similar to the consensus sequence. It is located 103-bp upstream of open reading frame 18, which encodes putative GDP-mannose 4,6-dehydratase, but does not lie within its promoter region; (2) GCCAGGAATTCCCGCA, situated near the 3\u2032-end of the internal sequence of rimF, which encodes an aminotransferase, but does not lie within the putative promoter region of rimG encoding cytochrome P450 monooxygenase; (3) ACCGGAAAATCCTTAG, which is present in the intergenic region of rimE and rimD, 100-bp upstream of rimE but not within its putative promoter region. The locations of these three sequences suggest that they do not comprise the core elements for gene expression, which may explain the limited regulatory effect exerted by RimR1 on rimocidin production. The mechanism whereby RimR1 regulates structural genes in the rimocidin gene cluster will be elucidated in a future study.RimR1 shares high similarity with several well-studied transcriptional regulators of the PAS-LuxR family, for example, it shares 49.61% amino acid sequence identity with PimM from talensis . Notablytalensis . Surprisrevealed \u201347. UsinrimR1 promoter. Moreover, qRT-PCR revealed that rimR2 deletion decreased rimR1 expression and restored it to a level comparable with that in the WT strain M527 when it was complemented, suggesting that RimR2 indirectly regulates rimR1 expression. The relationship between RimR1 and RimR2 and their regulatory hierarchy are also worth investigating in a future study.The biosynthetic gene clusters encoding polyene macrolide antibiotics have been sequenced and multiple regulatory genes, usually organized in a hierarchical network, have been identified within them , 31, 42.S. rimosus M527. These two tetraenes differ in the aglycone moiety, with a propyl group in rimocidin and a methyl group in CE-108. As the elongation module is common for both rimocidin and CE-108 biosyntheses, RimR2 regulates both biosyntheses almost identically  and YrimR1R (5\u2032-GATGAAGCCCTCGACGACAC-3\u2032) were designed following the rimR1 gene sequence (GenBank accession no. MK300953).RNA extraction and the analysis of the transcriptional level of o et al. . qRT-PCRrimR2 coding sequence was amplified by PCR using primers PrimR2-F2/R3 . RimR2 protein was purified using nickel-NTA column (Qiagen) and eluted using imidazole. The inducible expression and purification of were performed according to standard manipulation method described by Sambrook and Russel [A 2757-bp DNA fragment harboring the rim genes were amplified by PCR using the biotin labeled primers  for the identification of clusters involved in rimocidin, and function annotation of biosynthetic gene cluster was listed in t test was used for statistical analysis.All experiments were carried out at least three times, and the results were expressed by the mean\u2009\u00b1\u2009standard deviation (SD). Students\u2019 Additional file 1: Table S1. Detailed information of RimR2 and some polyene macrolide biosynthesis regulators from other Streptomyces species in phylogenetic tree.Additional file 2: Figure S1. Construction of mutant S. rimosus M527-\u0394rimR2. Map of plasmid pWHU2653- \u0394rimR2. The sgRNA consists of the 20 nt target gene specific guide sequence of S. rimosus M527 (green) and the invariant scaffold RNA (yellow). Light blue parallelograms connect the identical UHA and DHA sequences on pWHU2653 and the S. rimosus M527 chromosome where homologous recombination can take place.Additional file 3: Figure S2. PCR verification of the mutant S. rimosus M527-\u0394rimR2. M: DL5000 DNA Marker. Lane 1, The PCR products of 2.8-kb rimR2 gene were amplified by using the primers PrimR2-F1/R1 from WT strain S. rimosus M527; Lane 2, The PCR products of 6.8-kb cassette containing 2.8-kb rimR2 gene and its 2.0-kb upstream and 2.0-kb downstream fragment were amplified by using the primers P1/P4 from S. rimosus M527; Lane 3-5, The PCR products of rimR2 gene were amplified by using the PrimR2-F1/R1 from three randomly mutant strains M527-\u0394rimR2; Lane 6-8, The PCR products of 4.0-kb cassette containing 2.0-kb upstream and 2.0-kb downstream fragment were amplified by using the P1/P4 from three randomly mutant strains M527-\u0394rimR2.Additional file 4: Figure S3. HPLC analysis of rimocidin production in the WT strain S. rimosus M527, in mutant S. rimosus M527-\u0394rimR2, and in the complemented strain S. rimosus M527-\u0394rimR2/pSET152::rimR2, and control strain S. rimosus M527/pSET152.Additional file 5: Figure S4. Purification and elution of RimR2 protein. M: Protein Marker; Lane 1, Purified His6-tagged RimR2 protein after affinity nickel-NTA column. Lanes 2-4, Eluted RimR2 protein with 250 mM, 300mM, 500mM imidazole.Additional file 6: Figure S5. Construction of recombinant plasmids for over-expression of rimR2 gene with different promoters.Additional file 7: Figure S6. Phenotypic\u00a0verification of recombinant strains recombinant strains harboring over-expression of rimR2 gene. Recombinant strains could grow on 2CMC agar medium containing 300 \u03bcg/ml apramycin, while control strain S. rimosus M527 did not. 2CMC agar medium was incubated at 28 \u00b0C for 4 days.Additional file 8: Figure S7. PCR analysis of apramycin (apr) gene from recombinant strains harboring over-expression of rimR gene. DL DNA 2000 marker was used (M). Lane 1: PCR product of apr gene from S. rimosus M527(negative control); lane 2: PCR product of apr gene from plasmid pSET152(positive control); lane 3-5: PCR product of apr gene from recombinant strains S. rimosus M527-ER; lane 6-8: PCR product of apr gene from recombinant strains S. rimosus M527-KR; lane 9-11: PCR product of apr gene from recombinant strains S. rimosus M527-NR; lane 12-14: PCR product of apr gene from recombinant strains S. rimosus M527-21R; lane 15-17: PCR product of apr gene from recombinant strains S. rimosus M527-57R.Additional file 9: Figure S8. HPLC analysis of rimocidin isolated from fermentation extracts of the recombinant strains S. rimosus M527-KR, S. rimosus M527-NR, S. rimosus M527-ER, S. rimosus M527-21R, S. rimosus M527-57R and WT strain S. rimosus M527.Additional file 10: Figure S9. Construction of recombinant plasmids of overexpression of rimR1/rimR2/rimR3 /rimR4 gene with permE* promoter.Additional file 11: Figure S10. Detection and comparison of rimocidin production (a) and cell dry weight (b) of WT strain S. rimosus M527(\u25cf), recombinant strains M527-R1(\u25a0), M527-R2(\u25b2), M527-R3(\u25bc), and M527-R4(\u25c6) in shake-flask culture experiment.Additional file 12: Figure S11. Detection and comparison of antifungal activities of WT strain M527, recombinant strains M527-R1, M527-R2, M527-R3, and M527-R4 against F. oxysporum f. sp. cucumerinum. Spore suspension (500 \u03bcl) of F. oxysporum f. sp. cucumerinum (1\u00d7106 cfu ml-1) was spread and inoculated on PDA medium at 28 \u00b0C for 1 d. A agar block (4 mm in diameter) containing actively growing WT strain M527, three random recombinant strains M527-R1(a), M527-R2(b), M527-R3(c), and M527-R4(d) was aseptically placed on aforementioned PDA medium containing pathogenic fungus at 28 \u00b0C for 3-4 d. The diameter of inhibition zone was measured as antagonistic activity. Plant-pathogenic fungus F. oxysporum f. sp. cucumerinum was used as indicator strain in antifungal activities assay.Additional file 13: Figure S12. Phylogenetic tree of RimR1 and other polyene macrolide biosynthesis regulators (PAS-LuxR).Additional file 14: Figure S13. Phylogenetic tree of RimR2, RimR3, RimR4 and other polyene macrolide biosynthesis regulators (LAL).Additional file 15: Table S2. The primers used for deletion or expression of rimR2 gene in this study.Additional file 16: Table S3. The primers used for EMSA assay in this study."}
+{"text": "Results indicated that LPS decreased the villus height (p < 0.0001), increased the crypt depth (p < 0.05), and lowered the villus height to crypt depth ratio (p < 0.0001), while sodium acetate/sodium butyrate supplementation caused a significant increase in the villus height (p < 0.001), decrease in the crypt depth (p < 0.01), and increase in the villus height to crypt depth ratio (p < 0.001), especially. In mice treated with LPS, it was found that the serum level of IL-1\u03b2, TNF-\u03b1 (p < 0.001), and MDA (p < 0.01) was significantly higher; however, sodium acetate/sodium butyrate supplementation significantly reduced IL-1\u03b2 (p < 0.001), TNF-\u03b1 (p < 0.01), and MDA (p < 0.01), respectively. A total of 19 genera were detected among mouse groups; LPS challenge decreased the abundance of Lactobacillus, unidentified F16, unidentified_S24-7, Adlercreutzia, Ruminococcus, unclassified Pseudomonadales, [Ruminococcus], Acetobacter, cc 1, Rhodococcus, unclassified Comamonadaceae, Faecalibacterium, and Cupriavidus, while increased Shigella, Rhodococcus, unclassified Comamonadaceae, and unclassified Pseudomonadales in group L. Interestingly, sodium acetate/sodium butyrate supplementation increased Lactobacillus, unidentified F16, Adlercreutzia, Ruminococcus, [Ruminococcus], unidentified F16, cc 115, Acetobacter, Faecalibacterium, and Cupriavidus, while decreased Shigella, unclassified Enterobacteriaceae, unclassified Pseudomonadales, Rhodococcus, and unclassified Comamonadaceae. LPS treatment upregulated the expressions of ZO-1 (p < 0.01) and NLRP3 (p < 0.0001) genes in mice; however, sodium acetate/sodium butyrate solution supplementation downregulated the expressions of ZO-1 (p < 0.05) and NLRP3 (p < 0.05) genes in treated mice. Also, the LPS challenge clearly downregulated the expression of Occludin (p < 0.001), Claudin (p < 0.0001), and Caspase-1 (p < 0.0001) genes, while sodium acetate/sodium butyrate solution supplementation upregulated those gene expressions in treated groups. The present study revealed that sodium acetate/sodium butyrate supplementation alleviated LPS-induced diarrhea in mice via enriching beneficial bacterium and decreasing pathogens, which could regulate oxidative damages and inflammatory responses via NLRP3/Caspase-1 signaling. The current results may give insights into the prevention and treatment of diarrhea.Diarrhea is a word-widely severe disease coupled with gastrointestinal dysfunction, especially in cattle causing huge economic losses. However, the effects of currently implemented measures are still not enough to prevent diarrhea. Previously we found that dropped short-chain fatty acids in diarrhea yaks, and butyrate is commonly known to be related to the epithelial barrier function and intestinal inflammation. However, it is still unknown whether sodium acetate/sodium butyrate could alleviate diarrhea in animals. The present study is carried out to explore the potential effects of sodium acetate/sodium butyrate on lipopolysaccharide-induced diarrhea in mice. Fifty ICR mice were randomly divided into control (C), LPS-induced (L), and sodium acetate/sodium butyrate -treated groups. Serum and intestine samples were collected to examine inflammatory cytokines, antioxidant levels, relative gene expressions  Diarrhea is a severe disease coupled with gastrointestinal dysfunction that has a global impact on fertility rate, milk production, and immunity in livestock . NowadayGut microflora is composed of millions of microorganisms that contribute remarkably to physiological processes, i.e., functions of nutrition absorption, metabolism, and immunity of the host by producing various metabolites . The anaLactobacillus plantarum alleviated diarrhea in a previous study by balancing gut microbiota and regulating SCFAs , and after 24 hours mice were euthanized to collect serum, small intestine, and rectum samples  were purchased from Qing Long Shan Dong Wu Fan Zhi . After 30 days of rearing, mice were randomly divided into five groups, namely control (C), LPS (L), and treatment groups . Group D (400: 200), B (300:300), and A (200: 400) were treated with 600 mg/kg sodium acetate/sodium butyrate solution  samples . All aniDuodenum, jejunum, and ileum samples from all the groups were preserved in 4% paraformaldehyde for at least 48 hrs and then processed for commercial H&E staining . On an Olympus CX23 microscope with an integrated digital imaging analysis system, histological slices were examined . The villus height and crypt depth were measured as depicted in the previous study .via commercial assay kits . The concentration of cytokines including interleukin 1 beta (IL-1\u03b2), interleukin 6 (IL-6), interleukin 10 (IL-10), and tumor necrosis factor-alpha (TNF-\u03b1) was measured by using commercial ELISA kits .Blood samples of mice were centrifuged at 4,000 g for 10 min and stored at \u201320\u00b0C for future analysis. Antioxidant capacity was examined by detecting the levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-px), total anti-oxidation capacity (T-AOC), and malondialdehyde (MDA) by utilizing commercial assay kits . Meanwhile, the NO concentrations were determined n = 4), L (n = 4), and A (n = 4) were extracted utilizing the fast DNA Stool Mini Kit  according to the manufacturer\u2019s specifications. The quantity and quality of all extracted DNA samples were examined by using NanoDrop 2000 UV-vis spectrophotometer  and agarose gel electrophoresis, respectively. Gene amplification of bacterial 16S rRNA gene was performed using the V3\u2013V4 regions primers 338F (5\u2032-ACTCCTACGGGAGGCAGCAG-3\u2032) and 806R (5\u2032-GGACTACHVGGGTWTCTAAT-3\u2032). Then all amplicon products were purified by employing Vazyme VAHTSTM DNA Clean Beads  and quantified using the QuantiFluor\u2122-ST . At last, all samples were sequenced by using the Illumina MiSeq platform  with MiSeq Reagent Kit v3.The total microbial genomic DNA was extracted from the rectum contents of each mouse. The samples from groups C (1 (via mafft (2 (via ggtree3. Krona species composition map was generated via KronaTools (v2.7)4 (via PERMANOVA and Adonis in QIIME2 (2019.4). Venn5, heatmap, metagenomeSeq, LEFSe 4 . Signifiq, LEFSe , OPLS-DAq, LEFSe , and ranq, LEFSe  was perfq, LEFSe  using Mevia denaturing formaldehyde gel electrophoresis and NanoDrop 2000 analyzer  to validate their integrity and concentrations, respectively. Then commercial SuperScript\u2122IV first strand cDNA synthesis kits  were used for translating RNA samples into cDNA under the guidance of the manufacturer\u2019s specifications. Finally, qRT-PCR for all groups was carried out by using 25 uL of reaction mixtures consisting of 2 uL of intestinal tissues cDNA, 12.5 uL of Hieff UNICON\u00ae Universal Blue qPCR SYBR Green Master Mix , 2uL of primers, and 8.5 uL nuclease-free water, then the procedure was performed in the StepOnePlus\u2122 Real-Time PCR System . All sample reactions were repeated three times and the method of 2\u2013\u0394\u0394CT was utilized for calculating gene relative quantification. All primer pairs used in the present study were synthesized by Sangon Biotech (China) and are shown in Intestinal tissue RNA extraction from all mouse groups was performed by utilizing TRIzol reagent . All of the RNA samples were examined via ANOVA, Student\u2019s t-test, Kruskal\u2013Wallis, and Dunn\u2019s test via IBM SPSS (22.0) software. Data presented as means \u00b1 SD and statistically significant are considered when P < 0.05.All the generated data were evaluated via H&E staining and found that LPS caused a decrease in the villus height (p < 0.0001), an increase in the crypt depth (p < 0.05), and it also lowered the villus height to crypt depth ratio (p < 0.0001). Whereas, sodium acetate/sodium butyrate supplementation resulted in a significant increase in the villus height (p < 0.001), decrease in the crypt depth (p < 0.01), and increase in the villus height to crypt depth ratio (p < 0.001), especially in the mice of groups B and A (The mice were weighed on a daily basis and the weight of the mice in group A was slightly higher than mice in other groups . The dia B and A .p < 0.05), while there was no marked difference between group L and treated groups D, B, and A, respectively. In group C, the serum level of mice induced by LPS was found prominently high for IL-1\u03b2 (p < 0.001), TNF-\u03b1 (p < 0.001), and MDA (p < 0.01), respectively. However, in the serum of groups D, B, and A, sodium acetate/sodium butyrate supplementation caused a remarkable decrease in IL-1\u03b2 (p < 0.001), TNF-\u03b1 (p < 0.01), and MDA (p < 0.01), respectively  and non-singleton (p < 0.05) was found between groups C and L (p < 0.05) , ANOSIM (p < 0.05), and PERMDISP (p < 0.05), respectively  . Beta diectively . UPGMA aectively . Signifiectively .Proteobacteria (83.02%) and Bacteroidetes (13.00%). While Firmicutes (83.36%), Bacteroidetes (12.80%), Firmicutes (45.35%), and Proteobacteria (46.88%) were the main phyla in groups C and A. At the Class level, the main classes were Gammaproteobacteria (81.28%) and Bacteroidia (12.99%) in group L, while Bacilli (74.72%), Bacteroidia (12.80%), Clostridia (8.08%), Gammaproteobacteria (45.90%), Bacilli (44.04%), and Bacteroidia (6.80%) were the dominant classes in groups C and A, respectively. At the order level, the primary classes were Enterobacteriales (55.60%), Pseudomonadales (25.65%), and Bacteroidales (12.99%), while Lactobacillales (72.68%), Bacteroidales (12.80%), Clostridiales (8.08%), Lactobacillales (43.52%), Enterobacteriales (41.07%), and Bacteroidales (6.80%) were the staple orders in groups C and A, respectively. At the family level, the dominating families in group L were Enterobacteriaceae (55.59%), Pseudomonadaceae (25.47%), and Bacteroidaceae (9.09%), while the main families were Lactobacillales (72.47%) and S24-7 (12.50%) in group C, whereas Lactobacillales (43.35), Enterobacteriaceae (41.07%), and Bacteroidaceae (4.84%) in group A, respectively. At the genus level, the major genera in group L were Shigella (54.62%), Pseudomonas (25.42%), and Bacteroides (9.09%), while Lactobacillus (72.45%), unidentified_S24-7 (12.50), Lactobacillus (43.34%), Shigella (40.71), and Bacteroides (4.84%) were the dominating genera in groups C and A , Bacteroides (9%), and Pseudomonas (8%) in group L, while Shigella (56%), Pseudomonas (26%), and Bacteroides (9%) in group A, and unidentified S34-7 (45%), unidentified Clostridia (15%), Lachnospiraceae (7%), and Turicibacter (7%) in group C, respectively  projected on the coordinate axis was clearly farther than groups A (light orange) and C (lavender), which revealed a difference between group L, and groups A and C, respectively  OTUs were shared in groups C and L, while 335 (11.96%) OTUs were shared in groups C and A . Then ASm genera . ASV/OTUs genera . It is d C and A . PCA anaectively . Also, Oectively .Bacteroidales and Lactobacillales were found significantly on the upside of the broken lines with decreased ASV 102 (p < 0.05), ASV 23 (p < 0.05), ASV 14 (p < 0.05), and ASV 53 (p < 0.05), increased ASV 73 (p < 0.05), ASV 1 (p < 0.01), ASV 194 (p < 0.05), ASV 50 (p < 0.05), ASV 18 (p < 0.01), and ASV 5 (p < 0.001) between groups C and A. Different genera like Bacteroidales, Lactobacillales, Clostridiales, Burkholderiales, Desulfovibrionales, Enterobacteriales, and Pseudomonadales were found significantly in groups C and L, with 107 decreased ASV and 86 increased ASV , family S24-7, order Clostridiales, class Clostridia, genus Gemella, order Gemellales, family Gemellaceae, phylum Tenericutes, class Mollicutes, family Lachnospiraceae, order RF39, family F16, class TMT-3, order CW040, phylum TM7, and genus Ruminococcus in group C (lavender), genus Faecalibacterium, family Acetobacteraceae, and order Rhodospirillales in group A (light orange)  .Clostridium, Ruminococcus, Lactococcus, Gemella, etc. Network analysis revealed that there were more edges between groups A and C than between groups L and C, which inferred a higher similarity between groups A and C , unidentified F16 (p < 0.0001), Adlercreutzia (p < 0.01), Ruminococcus (p < 0.05), [Ruminococcus] (p < 0.05), Acetobacter (p < 0.05), cc 115 (p < 0.05), and Cupriavidus (p < 0.05) in group L was obviously lower than group C, respectively. While Shigella (p < 0.05) was prominently higher in group L than in group C. Unidentified_S24-7 and unclassified RF39 were significantly higher in group C than in groups A (p < 0.05) and L (p < 0.05), respectively, while unclassified Enterobacteriaceae was significantly lower in group C than in groups A (p < 0.05) and L (p < 0.05), respectively. The abundance of unclassified Pseudomonadales (p < 0.05), Rhodococcus (p < 0.05), unclassified Comamonadaceae (p < 0.05), and Lysobacter (p < 0.05) in group L was conspicuously higher than group A, respectively. The abundance of Faecalibacterium in group L was clearly lower than groups A (p < 0.05) and L (p < 0.01), respectively. The abundance of Gluconacetobacter in group A was prominently higher than groups L (p < 0.05) and C (p < 0.01), respectively. The abundance of unclassified Bradyrhizobiaceae in group C was clearly higher than in group A (p < 0.05)  .In summary, different analysis methods include relative abundance taxa, classification levels tree diagram, GraPhlAn evolutionary tree diagram, Krona species composition diagram, Venn Diagram, Genera composition heatmap, PCA, OPLS-DA, metagenomeSeq analysis, LEfSe analysis, and random forests and network analyses demonstrated that LPS challenge changed microbiota composing in mice; however, sodium acetate/sodium butyrate supplementation could partly restore the gut microbiota in animals.p < 0.05), 9 (p < 0.01), and 20 (p < 0.001) significant different pathways between groups C and L, 37 (p < 0.05), 27 (p < 0.01), and 53 (p < 0.001) significant different pathways between groups A and C, respectively (p < 0.05), 15 (p < 0.01), and 51 (p < 0.001) significant different pathways between groups C and L, while one (p < 0.001) significant different pathway between groups A and C was examined  and NLRP3 (p < 0.0001) in mice of group L; however, sodium acetate/sodium butyrate solution supplementation downregulated the expression of ZO-1 (p < 0.05) and NLRP3 (p < 0.05) genes in treated mice. Furthermore, change in LPS clearly downregulated the expression of Occludin (p < 0.001), Claudin (p < 0.0001), and Caspase-1 (p < 0.0001) genes in group L, while sodium acetate/sodium butyrate solution supplementation upregulated those gene expressions in treated groups of D, B, and A (Relative gene expression of Zonula occludens 1 (ZO-1), Occludin, Claudin, Caspase-1, and NLRP3 were detected by employing qRT-PCR. LPS induction prominently upregulated the expression of ZO-1 , Phenylobacterium, and Aminobacter were positively related to inflammatory cytokines. Pseudomonas, Faecalibacterium, Gluconacetobacter, Rhodococcus, (Clostridium), and Gluconobacter were positively related to tight junction proteins, while Adlercreutzia, Candidatus Arthromitus, Lactococcus, Ruminococcus, Coprococcus, Acinetobacter, cc_115, Gemella, Anaeroplasma, Phenylobacterium, Lysobacter, and W22 were negatively related to tight junction proteins. Adlercreutzia, Ruminococcus, Coprococcus, cc_115, Roseburia, Gemella, and Anaeroplasma were negatively related to expressions of Caspase-1 and NLRP3 , intestine morphology indices, inflammatory cytokines, oxidative indices, and gene expressions was performed through Statistical Analysis System. Results showed that nd NLRP3 .Though various kinds of measures have been implemented so far to fight against diarrhea, there is still a long way to go . A recenIn the present study, mice were supplemented with sodium acetate/sodium butyrate before inducing diarrhea by utilizing LPS. Serve damage to intestines particularly jejunum, ileum, cecum, and colon was examined , which wTo explore the potential mechanisms, we detected the gene expressions of tight junction proteins in intestines. Among them Occludin and Claudins were recognized as important components of intestinal permeability . The expp < 0.01) was detected in group L; however, there was a significant decrease in MDA (p < 0.01) levels in sodium acetate/sodium butyrate supplemented groups D, B, and A, respectively (p < 0.0001) in mice in group L; however, sodium acetate/sodium butyrate solution supplementation downregulated the expression of NLRP3 (p < 0.05) genes in treated mice. Also, the LPS challenge clearly upregulated the Caspase-1 (p < 0.0001) gene in group L, while sodium acetate/sodium butyrate solution supplementation downregulated it in treated groups D, B, and A (p < 0.001) were found in group C, which was consistent with previous results found higher inflammatory factors and upregulation of NLRP3 in HUVEC cells , Acetobacter, cc 1, Rhodococcus, unclassified Comamonadaceae, Faecalibacterium, and Cupriavidus, while increased Shigella, Rhodococcus, unclassified Comamonadaceae, and unclassified Pseudomonadales in group L. Interestingly sodium acetate/sodium butyrate supplementation increased Lactobacillus, unidentified F16, Adlercreutzia, Ruminococcus, (Ruminococcus), unidentified F16, cc 115, Acetobacter, Faecalibacterium, and Cupriavidus, while decreased Shigella, unclassified Enterobacteriaceae, unclassified Pseudomonadales, Rhodococcus, and unclassified Comamonadaceae. Lactobacillus genus bacteria are probiotic microorganisms that have beneficial effects on to host .KL and QK: research idea, methodology, visualization, and supervision. XC, QK, XZ, CZ, PH, and HL: reagents, materials, and analysis tools. KL: writing\u2014original draft and preparation. MK, ZB, II, HA, QS, and KL: writing\u2014review and editing. All authors contributed to the article and approved the submitted version."}
+{"text": "Limited data exist regarding longer term antibody responses following three-dose coronavirus disease 2019 (COVID-19) vaccination, and the impact of a first SARS-CoV-2 infection during this time, in people with HIV (PWH) receiving suppressive antiretroviral therapy (ART). We quantified wild-type-specific, Omicron BA.1-specific and Omicron BA.5-specific responses up to 6 months post-third dose in 64 PWH and 117 controls who remained COVID-19-naive or experienced their first SARS-CoV-2 infection during this time.Longitudinal observational cohort.We quantified wild-type-specific and Omicron-specific anti-Spike receptor-binding domain IgG concentrations, ACE2 displacement activities and live virus neutralization at 1, 3 and 6 months post-third vaccine dose.Third doses boosted all antibody measures above two-dose levels, but BA.1-specific responses remained significantly lower than wild-type-specific ones, with BA.5-specific responses lower still. Serum IgG concentrations declined at similar rates in COVID-19-naive PWH and controls post-third dose . Antibody function also declined significantly yet comparably between groups: 6 months post-third dose, BA.1-specific neutralization was undetectable in more than 80% of COVID-19 naive PWH and more than 90% of controls. Breakthrough SARS-CoV-2 infection boosted antibody concentrations and function significantly above vaccine-induced levels in both PWH and controls, though BA.5-specific neutralization remained significantly poorer than BA.1 even post-breakthrough.Following three-dose COVID-19 vaccination, antibody response durability in PWH receiving ART is comparable with controls. PWH also mounted strong responses to breakthrough infection. Due to temporal response declines, however, COVID-19-naive individuals, regardless of HIV status, would benefit from a fourth dose within 6\u200amonths of their third. In British Columbia (BC), Canada, third doses of coronavirus disease 2019 (COVID-19) monovalent mRNA vaccines were introduced in November 2021, initially to individuals at risk of severe COVID-19 outcomes, including some people with HIV (PWH). Whether offered as part of a primary vaccine series or a \u2018booster\u2019, third doses help to maintain systemic immunity and enhance protection against infection by viral variants . DespiteLongitudinal monitoring of immune responses post-third dose in PWH is critical to inform the timing of future immunizations in this group. Though some data are available on initial immunogenicity to third COVID-19 vaccine doses in PWH \u201316, no sOur cohort was described previously . The preThis study was approved by the University of British Columbia/Providence Healthcare and Simon Fraser University Research Ethics Boards. All participants provided written informed consent.50/200\u200a\u03bcl in the presence of serial two-fold plasma dilutions (1/20 to 1/2560) and added to target cells in triplicate. Viral cytopathic effects (CPE) were recorded 3 days post-infection. Neutralization was reported as the highest reciprocal dilution able to prevent CPE in all three wells. Partial or no neutralization at 1/20 dilution was considered below the limit of quantification (BLOQ) and coded as a reciprocal dilution of 10.Assays were performed as previously described ,19. IgG-U test (unpaired data) or Wilcoxon test (paired data). Relationships between continuous variables were assessed using Spearman's correlation. In participants who remained COVID-19-naive, multiple linear regression was used to investigate the relationship between HIV infection and vaccine-induced immune measures using a confounder model that adjusted for variables that could influence vaccine responses or that differed in prevalence between groups. For Omicron-specific neutralization at 6\u200amonths post-third dose, multiple logistic regression was used because of the high proportion of results BLOQ. Included variables were: HIV infection (controls as reference group), age (per year), sex at birth , ethnicity (nonwhite as reference), number of chronic conditions , dual ChAdOx1 as the initial regimen [mRNA or mixed (ChAdOx1/mRNA) regimen as the combined reference group] [P less than 0.05 considered statistically significant. Analyses were conducted using Prism v9.2.0 .Continuous variables were compared using the Mann\u2013Whitney e group] ,17, thir+ T-cell counts of 645 [interquartile range (IQR) 473\u2013958] cells/\u03bcl, and median nadir CD4+ T-cell counts of 225 (IQR 95\u2013485) cells/\u03bcl, at enrolment. PWH were a median of 57 (IQR 42\u201365) years old and 90% men; controls were a median 47 (IQR 35\u201372) years old and 73% women. PWH had a higher proportion of white ethnicity  and more chronic health conditions [median 1 (IQR 1\u20133) compared with 0 (IQR 0\u20131) in controls]. More PWH (10%) than controls (<1%) received two doses of the recombinant viral vector ChAdOx1 vaccine as their initial immunization series. All third doses were monovalent mRNA vaccines, either BNT162b2 (30\u200a\u03bcg) or mRNA-1273 (50 or 100\u200a\u03bcg). Most PWH (69%) and controls (62%) received mRNA-1273, where, per local guidelines, all adults aged at least 70\u200ayears were eligible for a 100\u200a\u03bcg mRNA-1273 dose, as were PWH who met one or more of the following criteria: age at least 65\u200ayears, prior AIDS-defining illness, prior CD4+ T-cell count less than 200\u200acells/\u03bcl, prior CD4+ T-cell fraction 15% or less, any plasma HIV load greater than 50\u200acopies/ml in 2021, or perinatally acquired HIV [Characteristics of the 64 PWH and 117 controls, all of whom remained COVID-19-naive until at least 1 month post-third dose, are shown in Table P\u200a=\u200a0.4). Though SARS-CoV-2 variant information is unavailable for individual infections, most were likely Omicron BA.1 or BA.2 based on local epidemiology at the time [A total of 24 (38%) PWH and 45 (39%) controls experienced their first SARS-CoV-2 infection between 1 and 6 months post-third dose (one control experienced two infections during this period ). Based the time .10 BAU/ml in COVID-19-naive PWH and 3.67 (IQR 3.50\u20133.86) log10 BAU/ml in controls, respectively (P\u200a=\u200a0.5). Following this, wild-type-specific IgG responses declined comparably in COVID-19-naive PWH and controls. At 3 months, wild-type-specific IgG responses in PWH and controls declined to a median 3.48 (IQR 3.12\u20133.75) and 3.40 (IQR 3.21\u20133.61) log10 BAU/ml, respectively (P\u200a=\u200a0.5), while by 6 months, responses had declined to a median 2.96 (IQR 2.62\u20133.38) and 3.06 (IQR 2.82\u20133.24) log10 BAU/ml in PWH and controls, respectively (P\u200a=\u200a0.4) , whereas those in controls had declined to significantly lower than post-second dose levels (P\u200a<\u200a0.0001)  . One month post-third dose, for example, BA.1-specific IgG concentrations were 2.96 (IQR 2.71\u20133.32) log10 BAU/ml in PWH, which was 0.74 log10 BAU/ml lower than wild-type-specific concentrations at this time. By 6 months, BA.1-specific IgG concentrations had declined to 2.47 (IQR 2.14\u20132.59) log10 BAU/ml in PWH, which was 0.49 log10 BAU/ml lower than wild-type-specific concentrations at this time.Similarly, Omicron BA.1-specific IgG responses were comparable in COVID-19-naive PWH and controls at all post-third dose time points , as was male sex (P\u200a=\u200a0.012 for BA.1-specific responses) (Supplementary Table 1). In fact, adjusted IgG concentrations were slightly higher in PWH compared with controls, though this was not statistically significant. Of note, receipt of an mRNA-1273 third dose (rather than BNT162b2) was associated with stronger wild-type-specific IgG responses (P\u200a=\u200a0.029), though not BA.1-specific responses (P\u200a=\u200a0.13), 6 months post-third dose. Among PWH, we also observed no significant relationship between most recent or nadir CD4+ T-cell count and either wild-type-specific or BA.1-specific IgG concentrations at this time .We next performed multivariable analyses adjusting for sociodemographic, health and vaccine-related variables to identify variables associated with wild-type-specific and BA.1-specific IgG concentrations at 6 months post-third dose in the COVID-19-naive subgroup. These analyses revealed that a higher number of chronic health conditions \u2013 but not HIV infection \u2013 was associated with poorer IgG responses at this time . Estimated BA.1-specific IgG half-lives were also comparable, at a median 71 (IQR 49\u2013104) days for PWH compared with 74 (IQR 59\u201390) days in controls (P\u200a=\u200a0.8). Multivariable analyses confirmed that HIV infection was not associated with wild-type-specific or BA.1-specific IgG half-lives post-third dose (Supplementary Table 2). Among COVID-19-naive PWH, we initially observed a weak inverse correlation between recent CD4+ T-cell count and IgG half-life, but this was not significant after excluding an outlier with a long (>200\u200aday) half-life .We estimated the half-lives of wild-type-specific IgG following three-dose vaccination to be a median 66 (IQR 47\u201389) days in COVID-19-naive PWH compared to 72 (IQR 54\u201396) days in controls, a difference that was not statistically significant , which was 0.27 log10 BAU/ml higher than at 1 month post-third dose (P\u200a<\u200a0.0001). Importantly, the magnitude of these \u2018hybrid\u2019 IgG responses was comparable between PWH and controls at all post-infection time points tested . Notably, while most participants experienced a marked boost in antibody levels following SARS-CoV-2 infection, IgG responses in a minority of PWH and controls remained constant or even declined post-infection , while by 6 months, activities had decreased to a median 87% (IQR 67.5\u201398.1) in PWH versus 93.6% (IQR 81.5\u201397.8) in controls (P\u200a=\u200a0.3), levels that were comparable or lower than after two-dose vaccination   compared with 99.1% (IQR 97\u201399.6%) in controls , but these responses also declined similarly in COVID-19-naive PWH and controls , to 30.8% (IQR 23.0\u201354.1) in PWH and 25.9% (IQR 12.1\u201352.2) in controls (P\u200a=\u200a0.3), where the latter values were significantly lower than those observed after two-dose vaccination .BA.1-specific responses remained significantly lower than wild-type-specific responses at all time points . There was also no evidence that a low recent or nadir CD4+ T-cell count was associated with lower wild-type-specific or BA.1-specific ACE2 displacement activities 6\u200amonths post-third dose in COVID-19-na\u00efve PWH . This was consistent with prior observations at 1 month post-third dose, which we attributed to the observation that PWH with low nadir CD4+ T-cell counts were eligible for the higher (100\u200a\u03bcg) third mRNA-1273 dose [Multivariable analyses confirmed that, among COVID-19-naive participants, HIV infection was not associated with either wild-type-specific or BA-1-specific ACE2 displacement activities at 6\u200amonths post-third dose (Supplementary Table 3). Rather, having received a mRNA-1273 third dose was the only independent correlate of stronger wild-type-specific ACE2 displacement activity at this time (273 dose .P\u200a<\u200a0.0001), where activities at 6 months in this group were overall significantly greater than peak responses induced by three-dose vaccination (both P\u200a<\u200a0.0001) . It is again notable, however, that a minority of PWH and controls did not show appreciable increases in this activity following infection  , while by 6 months, neutralization had declined to a median 80 (IQR 35\u2013160) in PWH and 40 (IQR 20\u201380) in controls (P\u200a=\u200a0.006), values that were below the levels originally elicited by two-dose vaccination (both groups P \u2264 0.01). As reported previously [One month post-third dose, wild-type-specific neutralization activity in PWH  was slightly higher than in controls , though responses in PWH were not further impaired compared with controls. In fact, 1 month post-third dose, BA.1-specific neutralization was higher in PWH compared with controls: a median 60 (IQR 25\u2013160) and 40 (IQR 20\u201340), respectively (P\u200a=\u200a0.0005), though these values were five-fold to eight-fold lower than corresponding wild-type-specific responses. BA.1-specific neutralization subsequently declined rapidly; at 3 months, this was BLOQ in 67.4% of COVID-19-naive PWH and 80.5% of controls (P\u200a=\u200a0.2), whereas at 6 months, this was BLOQ in 82.4% of PWH and 92.2% of controls (P\u200a=\u200a0.2), levels that were significantly lower than peak responses after two-dose vaccination . In multivariable analyses, older age \u2013 but not HIV infection \u2013 was the only significant correlate of poorer BA.1-specific neutralization among COVID-19-naive individuals at 6-month post-third dose . We also observed no significant associations between CD4+ T-cell parameters and either wild-type-specific or BA-1-specific neutralization at 6 months among COVID-19-naive PWH .BA.1-specific neutralization was significantly lower than wild-type responses at all timepoints for all groups  .By contrast, individuals who experienced breakthrough infection showed significantly stronger wild-type-specific and BA.1-specific neutralization compared with their COVID-19-naive counterparts at 3 and 6 months , and 16-fold lower than that for wild-type   and wild-type   .Since the emergence of Omicron BA.1, even more immune evasive variants have developed, including Omicron BA.5 that has dominated globally \u201326. We, Our results demonstrate that antibody response durability following three-dose COVID-19 vaccination in COVID-19-naive PWH receiving suppressive ART is comparable to controls without HIV. As we reported previously ,17, initBy contrast, almost all participants who experienced their first SARS-CoV-2 infection (presumably BA.1 or BA.2 ) post-th+ T-cell counts and/or who are not receiving suppressive ART. Indeed, results from several studies indicate that PWH with CD4+ T-cell counts below 500\u200acells/\u03bcl mount weaker responses to the first [Our study has several limitations. Our observations may not be generalizable to PWH with low CD4he first \u201332 and she first \u201338 doseshe first , future he first \u201344.In conclusion, our observations confirm the humoral immune benefits of third COVID-19 vaccine doses in PWH receiving suppressive ART, and further reveal that the durability of third-dose responses is comparable to that in persons without HIV. Nevertheless, regardless of HIV status, individuals who remain COVID-19-naive will benefit from a fourth dose within 3\u20136\u200amonths of their third dose, as antibody concentrations and neutralization activities declined markedly over time in this group, and the ability of vaccine-induced responses to neutralize the dominant Omicron BA.5 variant were even poorer than BA.1. By contrast, the majority of individuals who experienced their first SARS-CoV-2 infection post-third vaccine dose showed significantly higher antibody activities than those induced by vaccination alone . These observations suggest that a slightly delayed fourth dose (e.g. to 3\u20136\u200amonths following infection) would optimally benefit this group. Further studies of hybrid response durability are required, as are direct comparisons with immune responses elicited by a fourth dose in COVID-19-naive individuals, particularly in light of new bivalent formulations that include wild-type and Omicron Spike antigens.This work is dedicated to the memory of our friend and colleague Hesham Ali who sadly passed away in July 2022. We thank the phlebotomists and laboratory staff at the BC Centre for Excellence in HIV/AIDS, the Hope to Health Research and Innovation Centre, St. Paul's Hospital, and Simon Fraser University for assistance. Above all, we thank the participants, without whom this study would not have been possible.Author contributions: M.A.B. and Z.L.B. are co-principal investigators and conceived the study, with C.T.C., C.C., A.H.A., V.L., M.H., M.G.R., R.G., S.G., J.S.G.M., M.H. and M.H. additionally contributing to study design and development. M.A.B., Z.L.B., M.G.R., A.H.A., C.T.C. and C.C. obtained project funding. M.A.B., Z.L.B., H.R.L., F.M. and P.K.C. designed experiments. H.R.L., R.M., P.K.C., Y.S., F.Y., S.S., E.B., N.M.-G., S.D., M.C.D., R.K., S.E., L.Y., B.G., F.H.O., G.U., J.T., P.S., L.B. and C.J.B. contributed to specimen collection, data collection, data curation and/or data analysis. H.R.L., Y.S., D.H., M.L.D., J.S., M.N., M.G.R., M.A.B. and Z.L.B. supervised the research, laboratory assays and/or contributed to project or cohort management. Z.L.B. performed statistical analyses with support from C.J.B.. N.P., C.F.L., M.G.R., W.D., C.J.B., and M.N. provided, generated and/or validated local Omicron isolates. H.R.L. wrote the first draft of the manuscript.Funding: this work was supported by funding from Genome BC, Michael Smith Health Research BC, and the BCCDC Foundation for Public Health through a rapid SARS-CoV-2 vaccine research initiative in BC award . It was also supported by the Public Health Agency of Canada (PHAC) through two COVID-19 Immunology Task Force (CITF) COVID-19 Awards . Additional funding was received from the Canadian Institutes for Health Research , the Coronavirus Variants Rapid Response Network , the Canada Foundation for Innovation through two Exceptional Opportunities Fund COVID-19 awards , a British Columbia Ministry of Health\u2013Providence Healthcare Research Institute COVID-19 Research Priorities Grant (to C.J.B. and C.F.L.) and the CIHR Canadian HIV Trials Network (CTN) (to A.H.A.). F.M. is supported by a fellowship from the CIHR Canadian HIV Trials Network. M.L.D. and Z.L.B. hold Scholar Awards from Michael Smith Health Research BC. F.Y. and E.B. were supported by SFU Undergraduate Research Awards. M.D. is supported by a CIHR Canada Graduate Scholarships-Master's award. G.U. and F.H.O. are supported by PhD fellowships from the sub-Saharan African Network for TB/HIV Research Excellence (SANTHE), a DELTAS Africa Initiative (grant # DEL-15-006). The DELTAS Africa Initiative is an independent funding scheme of the African Academy of Sciences (AAS)'s Alliance for Accelerating Excellence in Science in Africa (AESA) and supported by the New Partnership for Africa's Development Planning and Coordinating Agency (NEPAD Agency) with funding from the Wellcome Trust (grant # 107752/Z/15/Z) and the UK government. The views expressed in this publication are those of the authors and not necessarily those of PHAC, CITF, AAS, NEPAD Agency, Wellcome Trust, the Canadian or UK governments or other funders.https://doi.org/10.1101/2022.11.03.22281912Posted history: this manuscript is posted to MedRxiv: There are no conflicts of interest."}
+{"text": "Middle cerebral artery occlusion/reperfusion (MCAO/R) was created in mice. Oxygen-glucose deprivation/reoxygenation (OGD/R) was performed in brain microvascular vessel-derived endothelial cells (bEnd.3) to mimic ischemia/reperfusion injury in vitro. P-gp-specific siRNA and pharmacological inhibitor cyclosporine A were used to inhibit P-gp, whereas pcDNA3.1 was utilized to overexpress P-gp. Twenty-four hours after reperfusion, acute ischemic stroke outcome, BBB integrity and permeability, autophagic proteins and relative signaling pathways were evaluated. P-gp levels were markedly elevated in mouse brain and endothelial cells following MCAO/R and OGD/R, respectively. P-gp siRNA silencing or pharmacologically inhibiting (cyclosporine A) reduced infarct volume and brain edema, attenuated brain pathology, and improved neurological behavior in association with attenuated accumulation of neutrophils and macrophages, reduced expression levels of inflammatory cytokines (TNF-\u03b1 and IL-1\u03b2), matrix metalloproteinases (MMP-2 and MMP-9) and adhesion molecules (ICAM-1 and VCAM-1). P-gp silence also counteracted BBB leakage, restored the expressions of tight junction proteins , activated autophagic proteins , and diminished Akt/mTOR signal activity in mice following MCAO/R. In the endothelial cell OGD/R assay, P-gp silence downregulated the expressions of inflammatory cytokines and adhesion molecules, inhibited leukocytes adhesion and migration, increased tight junction protein levels, and activated autophagy, all were reversible by forceful P-gp expression. Additionally, treatment with an autophagy inhibitor (3-methyladenine) abolished protections against ischemic stroke and tight junction proteins reduction followed by P-gp silence. In conclusion, increased P-gp expression after ischemic injury resulted in BBB dysfunction and hyperpermeability by suppressing Akt/mTOR-induced endothelial autophagy.P-glycoprotein (P-gp) is expressed on brain microvessel endothelial cells of blood-brain barrier (BBB) and elevated after cerebral ischemia. In this study, we explored the influence and potential mechanisms of P-gp on BBB function in experimental ischemic stroke  Stroke is one of leading causes of disability and mortality worldwide, with ischemic stroke accounting for 87% of all stroke-related incidents . InflammP-glycoprotein (P-gp), also known as multidrug resistance transporter-1, is a membrane transport protein belonging to adenosine triphosphate (ATP)-binding cassette (ABC) transporter superfamily . As a maAutophagy is a self-eating cellular process to maintain cellular homeostasis and normal cellular functions , and is In this study, we investigated the effect and its underlying mechanisms of P-gp on BBB integrity in the mouse model of ischemic stroke. We found that elevated microvascular endothelial P-gp degraded endothelial tight junction, increased BBB hyperpermeability, accelerated brain inflammation and thus worsened ischemic stroke outcomes in association with increased Akt/mTOR activity and reduced endothelial autophagy.Male C57BL/6 mice weighting 22-25 g were purchased from Qinglongshan Animal Breeding Centre  and used in all experiments. All animals were housed and fed freely in a temperature-controlled room (22 \u00b1 2\u00b0C) with a 12 h light-dark cycle. All experimental procedures were performed under licenses that granted by China Pharmaceutical University Animal Experimental committee , in compliance with the National Institutes of Health  guidelines for the care and use of animals. Mice were randomly assigned into 6 treatments, including sham treatment, middle cerebral artery occlusion/reperfusion (MCAO/R) treatment, MCAO/R with negative control siRNA (NC siRNA) treatment, MCAO/R with NC siRNA and cyclosporine A treatments (NC siRNA + CsA), MCAO/R with P-gp siRNA treatment (MCAO/R + P-gp siRNA), MCAO/R with P-gp siRNA and 3-MA treatments (MCAO/R + P-gp siRNA + 3-MA). The treatment was administered in a blinded fashion. The researcher conducting the surgeries and behavior tests was unaware of the animal groups, and the researcher analyzing the data did not know the group condition.Mice were anesthetized by 3% isoflurane and maintained using 1.5% isoflurane  and placed in a stereotaxic frame  as previously described . The skiAutophagy inhibitor (3-MA) and P-gp inhibitor (CsA) were purchased from MedChemExpress LLC . 3-MA was prepared in normal saline immediately before use, and intracerebroventricularly injected at a dose of 15 \u03bcg per mouse 30 min prior to MCAO surgery . CsA wasFollowing 12 h fasting, MCAO/R was performed as described previously . Mice weTwenty-four hours after MCAO/R surgery, neurological score was graded as 0-4 based on Bederson\u2019s method , with hiThe whole brain was removed after mice were sacrificed at 24 h after reperfusion, and each brain was sliced into coronal sections (2 mm thickness). Brain slices were stained in TTC  at 37\u00b0C for 20 min, photographed with a digital camera, and analyzed using the ImageJ software  . HemisphTo estimate brain edema, brains were weighed prior to and 12 h after drying in a 110\u00b0C oven. Brain edema was calculated as percent of water content as following : Brain wTo evaluate the integrity of BBB , mice we2) at 37\u00b0C. The cells at 70-80% confluence was transfected with mouse P-gp (GGATCCAGTCTAATAAGAA) siRNA (50 pmol per well (6-well), RiboBio, Guangzhou, Guangdong, China) or P-gp pcDNA3.1 (+)  using the lipofectamine 2000 reagent  to silence or overexpress P-gp, respectively. The transfection efficiency was assayed 36 h after transfection by Western blot according to the manufacturer\u2019s recommendation. The control cells were transfected with the same volume of NC siRNA (RiboBio) or NC pcDNA3.1 (+) (Sangon Biotech).bEnd.3 cells were obtained from China Pharmaceutical University and cultured in high-glucose Dulbecco\u2019s Modified Eagle\u2019s Medium  supplied with 10% fetal bovine serum , D-Glucose (4.5g/L), glutamine (2 mmol/L), penicillin (80 U/mL), streptomycin (0.08 mg/mL) and pyruvate (1 mmol/L) in humidified air (5% CO2 and 5% CO2) at 37\u00b0C for 2 h [2) and maintained in serum-free high-glucose DMEM for 24 h. In control group, cells were cultured in the chamber with serum-free high-glucose DMEM and normal culture conditions for the same period. The cells were treated with an autophagy inhibitor 3-MA (1 mmol/L) 1 h prior to OGD stimulation [Confluent bEnd.3 cells were washed twice and subjected to OGD/R by replacing with serum-free low-glucose  DMEM in an anoxic incubator  was replenished every 2 d. Approximately 6-7 d after seeded, bEnd.3 monolayers were obtained. Leukocyte (1\u00d7106 cells in 0.2 mL of DMEM) were added to upper chamber and incubated for 4 h in bEnd.3 cells. Migrated leukocytes were counted from the lower chamber. Leukocyte transendothelial migration was quantitated as the number of leukocytes per square millimeter.Leukocyte transendothelial cell migration assay was performed as previously described  with sliBrains were harvested, fixed in 4% paraformaldehyde (PFA) for 24 h, dehydrated in graded ethanol, embedded in paraffin, and sectioned (6 \u03bcm). After deparaffinization and rehydration, sections were stained with Giles\u2019 hematoxylin and eosin (H&E) and imaged on a fluorescent inverted microscope .Paraffin sections (2 \u03bcm in thickness) were used for all staining. Briefly, following blocking endogenous peroxidase with PBS containing 3% hydrogen peroxide for 5 min, sections were stained with primary antibodies for neutrophils , macrophages , intercellular adhesion molecules-1 , vascular adhesion molecule-1 , and P-gp  at 4\u00b0C overnight. Then, sections were incubated with Immunohistochemical staining kit  for 1 h at room temperature. All stained sections were counterstained with Giles\u2019 hematoxylin and imaged on a fluorescent inverted microscope . To quantify P-gp expression levels, neutrophil and macrophages, randomly selected fields from the ischemic cortex were analyzed by using ImageJ software.Mice were anesthetized 24 h after reperfusion and perfused intracardially with cold 4% PFA followed by perfusion with PBS. Brains were fixed with 4% PFA and embedded in optimal cutting temperature compound (OCT), and sectioned (12 \u03bcm) followed by blocking nonspecific staining with 8% goat serum for 2 h at room temperature. For culture cells, the cells were fixed in 4% PFA for 20 min and incubated with solution containing 0.3% Triton X-100 in PBS (PBST) containing 8% goat serum for 2 h at room temperature for non-specific staining blocking and permeabilization .Tissue sections and cells were incubated with the primary antibodies against CD31  ICAM-1 (1:100), VCAM-1 (1:50), Claudin-5 , Occludin , Zonula occludens-1 , light chain 3  and glucocorticoid receptor  at 4\u00b0C overnight. Following 3 washes with PBST, sections or cells were incubated with Cy3-conjugated goat anti-mouse IgG  and FITC-conjugated goat anti-rabbit IgG  for 2 h at room temperature, washed with PBST 3 times and counterstained with 4, 6-diamidino-2-phenylindole  for 30 min at room temperature. Stained images were captured by a laser confocal microscope  or a fluorescent microscope , and analyzed using ImageJ software.Total RNA was extracted using the RNA isolater  and was transcribed into cDNA using a HiScript II Q RT SuperMix (Vazyme Biotech). Real time PCR was performed using quantitative PCR  with a fluorescent dye (SYBR Green I). mRNA levels were normalized to \u03b2-actin of the same samples and were reported as fold changes relative to sham or control treatment . The priTotal protein was extracted using RIPA lysis containing protease and phosphatase inhibitor cocktail . Protein concentrations were determined by BCA assay. Proteins were separated by SDS-PAGE and transferred onto polyvinylidene fluoride (PVDF) membranes. After blocking with 5% non-fat milk in Tris-buffered saline with 0.05% Tween 20 (TBST) for 1.5 h at room temperature, the membrane was incubated with antibodies against P-gp (1:500), Claudin-5 (1:500), Ocluddin (1:1000), ZO-1 (1:1000), LC3 (1:1000), mTOR , p-mTOR , Akt , p-Akt  and \u03b2-actin  at 4\u00b0C overnight. After 3 times washes with TBST, the blots were incubated with HRP goat anti-rabbit IgG (H + L)  or HRP-labeled goat anti-mouse IgG (H + L)  for 2 h at room temperature. The bands were visualized using the enhanced chemiluminescence (ECL) reagents (Affinity Biosciences) and were quantified with ImageJ software. The protein expression levels were expressed as the percentage of \u03b2-actin.To evaluate the function of P-gp on glucocorticoid transport, mice were injected with exogenous glucocorticoid  through femoral vein at 24 h after MCAO/R. Brains were harvested and ischemic hemisphere was separated at 5 min after the injection. Brain was homogenized with pure water with a mass/volume ratio of 1:3. Dexamethasone concentration in homogenates was determined by a UPLC-MS/MS system  and given as ng/g ischemic brain tissue.P<0.05. All data are presented as the means \u00b1 SD.All statistical analyses were performed using IBM SPSS Statistics 19.0 software. The distribution of all data was tested for normality via a One-Sample Kolmogorov-Smirnov test. Statistical differences among the groups were analyzed by One-Way Analysis of Variance (ANOVA) with post hoc Tukey tests if data were normally distributed and consistent with homogeneity of variance among the experimental groups. The Mann-Whitney test was used to the compare ranks if the data were not normally distributed or not consistent with homogeneity of variance. The difference was considered significant at P<0.01). Treatment with NC siRNA did not impact P-gp expression, infarct volume, and brain histopathology. Therefore, NC siRNA + MCAO/R mice were used to observe P-gp function in following experiments as model control. P-gp siRNA pretreatment reduced infarct volume, attenuated neurological impairment, and improved histopathological damage as compared to NC siRNA treatment after MCAO/R (P<0.01).Mice underwent the MCAO surgery were included for successful occlusion. P-gp expression was substantially increased at ischemic hemisphere 24 h following MCAO/R surgery. Treatment with P-gp siRNA significantly reduced P-gp expression in ischemic brain as compared to NC siRNA treatment , P<0.01.r MCAO/R , P<0.01.P<0.01).Ischemic stroke breaks down BBB, leading to the entry of serum proteins into brain fluid and edema formation . Twenty-P<0.05).MMP-2 and MMP-9 degrade collagen IV that is a major component of basal lamina, and ultimately disrupt BBB . In RT-PP<0.01). In tissue immunostaining, neutrophils (MPO) and macrophages (F4/80) were observed in the infarct site 24 h after MCAO/R. However, P-gp siRNA treatment substantially reduced the densities of both neutrophils and macrophages as compared to NC siRNA treatment (P<0.01).ICAM-1 and VCAM-1 are important for recruiting leukocytes into inflamed brain. ICAM-1 and VCAM-1 levels were significantly lower in the ischemic area of mice treated with P-gp siRNA than those in mice treated with NC siRNA , P<0.01.reatment , P<0.01.P<0.05, P<0.01). These results indicate that silencing P-gp reduced BBB permeability by increasing TJPs expression.In the CNS, tight junctions form a barrier to limit paracellular permeability. In Western-blotting analysis, the expression levels of TJPs including Occludin, Claudin-5, and ZO-1 were largely reduced in ischemic brain of NC siRNA-treated MCAO/R mice. In contrast, P-gp siRNA treatment restored the expression levels of all proteins in ischemic area . In immunofluorescence staining, P-gp knockdown also dramatically enhanced LC3 expression in endothelial cells (CD31+) (P<0.01).To determine the influence of P-gp on endothelial autophagy, we assessed the expression of microtubule-associated protein LC3 and autophagy adapter protein P62, two well established markers for autophagy activation , 39. In  (CD31+) , P<0.01.P<0.01). Glucocorticoid binds to glucocorticoid receptor (GR), induces GR nuclear translocation and thus inhibits Akt/mTOR activity [P<0.01).Akt/mTOR activity suppressed autophagy . As indiactivity . In ischP<0.05, P<0.01).To determine whether autophagy contributes to the suppression of ischemic stroke by P-gp inhibition, an autophagy inhibitor 3-MA was given to mice 30 min prior to stoke induction. 3-MA treatment did not alter P-gp expression in ischemic brain of P-gp siRNA-treated mice. However, inhibiting autophagy by 3-MA counteracted the effects of P-gp siRNA treatment on ischemic stroke outcome as indicated by enlarged infraction size, increased brain edema, worsened neurological defects and brain histopathology (P<0.05). Histologically, CsA treatment reduced infarct volume, brain edema, neurological behavior defects and brain histopathology induced by MCAO/R .Additionally, similar to P-gp siRNA silence, pharmacological inhibition of P-gp with CsA significantly attenuated P-gp expression levels in ischemic brain as compared to vehicle treatment , P<0.05.P<0.01). Preincubation with P-gp siRNA inhibited mRNA expressions of TNF-\u03b1, IL-1\u03b2, MMP-2, and MMP-9 in endothelial cells induced by OGD/R (P<0.05). Consistent with in vivo findings, P-gp knockdown substantially downregulated the expression of ICAM-1 and VCAM-1 on endothelial cells following OGD/R treatment (P<0.01). P-gp silence also mitigated leukocyte adhesion and transendothelial migration (P<0.01), while increased the expression levels of TJPs  in endothelial cells undergoing OGD/R .Under OGD/R condition, P-gp expression was markedly elevated in endothelial cells (bEnd.3 cells). P-gp knockdown dramatically reduced P-gp expression as compared to NC siRNA incubation , P<0.01.by OGD/R , P<0.05.reatment , P<0.01.igration , P<0.01,P<0.01) as compared to NC plasmid pcDNA3.1. P-gp overexpression increased mRNA expression levels of TNF-\u03b1, IL-1\u03b2, MMP-2, and MMP-9 following OGD/R treatment as compared to NC plasmid pcDNA3.1 (P<0.05). Additionally, P-gp overexpression also increased adhesion molecules expression, augmented leukocytes adhesion and transmigration , whereas reduced the expression levels of Claudin-5, Occludin, and ZO-1 in endothelial cells following OGD/R .To examine the effect of P-gp overexpression, endothelial cells were transfected with P-gp pcDNA3.1 plasmid to forcefully express P-gp or with pcDNA3.1 plasmid serving the NC. P-gp pcDNA3.1 transfection remarkably upregulated P-gp expression , P<0.01 pcDNA3.1 ; P<0.05.P<0.01). These data indicate that P-gp silence led to autophagy activation to protect endothelial cells against OGD/R-induced cellular injury. Further, P-gp silence largely diminished the phosphorylation of Akt and mTOR protein signaling (P<0.05), suggesting that P-gp silence may activate autophagy by inhibiting Akt/mTOR signaling activity.To assess the influence of P-gp on autophagy and potential mechanisms, endothelial cells were transfected with P-gp or NC siRNA followed by OGD/R treatment. In Western-blotting and immunofluorescence staining analyses, P-gp silence dramatically increased the the ratio of LC3-II/LC3-I as well as Beclin 1 levels, whereas reduced P62 levels ; P<0.01.ignaling , P<0.05,P<0.05, P<0.01). When endothelial cells were incubated with a small molecule inhibitor 3-MA, the upregulation of TJPs by P-gp silence was almost ablated . Expectedly, 3-MA treatment reduced the ratio of LC3-II/LC3-I and Beclin 1 expression (P<0.01), whereas increased P62 expression (P<0.01). These results indicate that autophagy activation by P-gp silence may protect OGD/R-induced endothelial dysfunction.Although OGD/R treatment dramatically reduced TJPs levels in endothelial cells, P-gp silence increased the suppression of TJPs expression following OGD/R as compared to NC siRNA treatment (pression , P<0.01,pression , P<0.01.in vivo and in vitro. P-gp silence or pharmacological inhibition alleviated ischemic stroke by improving the integrity and function of BBB. Therefore, P-gp overexpression caused BBB dysfunction and exacerbated stroke outcome by destroying TJPs and worsening inflammatory response.Despite advances in our understanding of P-gp in multiple drug resistance, the role and underlying mechanisms in BBB dysfunction induced by ischemic stroke remain largely unknown. In this study, we found increased expression of P-gp in experimental ischemic stroke both in vitro, or overexpressed P-gp by transfecting endothelial cells with P-gp pcDNA3.1. We showed that P-gp siRNA relived acute stroke injury, including reduction in infarct volume, improvement in neurological behaviors, and decrease in brain edema. Similar effects were also observed using a P-gp inhibitor CsA. Therefore, elevated P-gp expression induced by ischemic stroke is vital for aggravating acute stoke injury.P-gp is involved in the pathogenesis of certain CNS diseases by regulating inflammatory cytokine expression and immune cell infiltration , 12. ThoP-gp is mainly expressed on BMVECs of BBB that functions as a semipermeable barrier between the CNS and the peripheral circulation , 46. In TJPs including ZO-1, Claudin-5, and Occludin are important in regulating the integrity and permeability of BBB and are disrupted and redistributed after ischemic stroke , 54. We Autophagy is a crucial degradation pathway for maintaining cellular and energy homoeostasis and protecting cell death . AutophaIn conclusion, we demonstrated that elevated P-gp levels following ischemic stroke exacerbated BBB breakdown and brain inflammatory response by increasing Akt/mTOR activity and suppressing autophagy activation. Our findings help understand the role and underlying mechanisms of P-gp in brain inflammatory response and BBB integrity following ischemic stroke.www.aginganddisease.org/EN/10.14336/AD.2022.0225.The Supplementary data can be found online at:"}
+{"text": "Siphoviridae phage that infects Gordonia terrae 3612. The 68,128-bp genome of StarStruck has a GC content of 65.4% and contains 92 protein-coding genes, including the gene for a HicA-like toxin. StarStruck was assigned to subcluster CR2 based on >35% shared gene content with other cluster CR genomes in the Actinobacteriophage Database.Bacteriophage StarStruck is a lytic  Actinobacteria, are extremely abundant and diverse  icosahedral head and a 297.5-nm  flexible, noncontractile tail (n = 3 particles).Actinobacteriophages, which are viruses that infect bacteria of the phylum  diverse \u20134. By chdiverse \u2013, 6. Starrae 3612 . Soil exe assays . After ilysogens . The parCGCCGCGTAC). StarStruck shares >35% gene content with members of cluster CR in the Phamerator database Actino_Draft and was assigned to subcluster CR2  . Sequencster CR2 , 10, 11.http://cobamide2.bio.pitt.edu) and PECAAN (https://blog.kbrinsgd.org/) (http://phages.wustl.edu/starterator) analyses (The genome of StarStruck was autoannotated using GLIMMER v3.02 and GeneMark v2.5 within DNA Master v5.23.6 (gd.org/) , 13. Traanalyses . Putativanalyses , 15, 16.analyses , 18. Staanalyses . The laranalyses .Like many cluster CR phages, StarStruck encodes a HicA-like toxin (gp7) within the 12,000-bp region separating the small- and large-subunit terminases. Another interesting feature of StarStruck is the location of the lysin B (gp19) within this region, rather than adjacent to the lysin A genes (protease C39 domain [gp49] and glycosyl hydrolase domain [gp50]), which are located downstream of the minor tail proteins.ON456333 and the Sequence Read jhu7 (SRA) accession number SRX14816101.StarStruck is available at GenBank with the accession number"}
+{"text": "Pancreatic ductal adenocarcinoma (PDAC) is currently the most deadly cancer. Although characterized by 5\u201320% of neoplastic cells in the highly fibrotic stroma, immunotherapy is not a valid option in PDAC treatment. As CXCR4-CXCL12 regulates tumor invasion and T-cell access and PD-1/PD-L1 controls immune tolerance, 76 PDACs were evaluated for CXCR4-CXCL12-CXCR7 and PD-1/PD-L1 in the epithelial and stromal component. Neoplastic CXCR4 and CXCL12 discriminated PDACs for recurrence-free survival (RFS), while CXCL12 and CXCR7 discriminated patients for cancer-specific survival (CSS). Interestingly, among patients with radical resection (R0), high tumor CXCR4 clustered patients with worse RFS, high CXCL12 identified poor prognostic patients for both RFS and CSS, while stromal lymphocytic-monocytic PD-L1 associated with improved RFS and CSS. PD-1 was only sporadically expressed (<1%) in focal lymphocyte infiltrate and does not impact prognosis. In multivariate analysis, tumoral CXCL12, perineural invasion, and AJCC lymph node status were independent prognostic factors for RFS; tumoral CXCL12, AJCC Stage, and vascular invasion were independent prognostic factors for CSS. CXCL12\u2019s poor prognostic meaning was confirmed in an additional perspective-independent 13 fine-needle aspiration cytology advanced stage-PDACs. Thus, CXCR4-CXCL12 evaluation in PDAC identifies prognostic categories and could orient therapeutic approaches. Pancreatic ductal adenocarcinoma (PDAC) represents the fourth leading cause of cancer-related deaths in economically advanced countries . In the Patients with resectable PDAC  consecutThree-micrometer sections were cut from formalin-fixed paraffin-embedded (FFPE) tissue blocks. Sections were stained with hematoxylin and eosin for adequate tumor representation.2/field) for each ROI, on Olympus BX51 microscope . The evaluation was carried out by 3 qualified observers . For CXCR7 and CXCL12, staining extension was calculated for each case by the average percentage of positively stained cancer cells for the three ROIs. For CXCR4, due to heterogeneous staining patterns, the staining value was calculated by H-score. Staining intensity was based on membrane staining and rated as absent (0), weak/low (1+), and intermediate/moderate (2+), typically with cytoplasmic localization, strong/high (3+), typically cytoplasmic with membrane localization of CXCR4 staining. The % of positive cells at each staining intensity is obtained, and an H-score is determined by the sum of each intensity rating multiplied by its corresponding percentage. CXCR4 was scored as positive at H-score > 50, corresponding to PDAC with multiple subcellular signal accumulation (membrane and cytoplasm). CXCR7 and CXCL12 cells were rated positive when stained regardless of the cellular localization. CXCR7 was scored as positive with >20% positive cells [FFPE sections were dewaxed and rehydrated, and heat-induced epitope retrieval (HIER)  with appropriate Antigen Unmasking Solution was performed. After incubation with the appropriate serum for blocking non-specific background, FFPE tumor tissue slides were incubated overnight at 4 \u00b0C using primary antibodies: Mouse Monoclonal anti-human Anti-CXCR7/RDC-1 Antibody ((11G8), HIER citrate buffer pH6, 1:50 dilution, R&D Systems); Mouse Monoclonal anti-human Anti-CXCL12/SDF-1 Antibody ((79018), HIER citrate buffer pH6, 1:50 dilution, #MAB350 R&D Systems); two commercial CXCR4 antibody clones the mouse Monoclonal anti-human Anti-CXCR4/CD184 ((44716) HIER, citrate buffer pH6, 1:100 dilution, #MAB172 R&D Systems) and the rabbit monoclonal anti-human Anti-CXCR4 ((UMB-2) HIER, Tris-EDTA buffer, pH 9.0, 1:200 dilution, Abcam); two commercial PD-L1 antibodies; the rabbit monoclonal anti-human anti-PD-L1/CD274 ((E1L3N) 1:200; diluted; HIER citrate buffer pH6, #13684 Cell Signaling Technology and (22C3) ready to use with HIER Conditioning Solution (CC1) pH9 Ventana Medical Systems); Mouse monoclonal anti-human PD-1/CD279 ((NAT-105) ready to use, HIER Cell Conditioning Solution (CC1) pH9, Ventana Medical Systems). Stained pancreatic cancer cells were assessed in at least three regions of interest (ROI)/slide, recognized at low power (100\u00d7 magnification), and the cells enumerated in 5 consecutive, not-overlapping high-power fields , and p > 0.1 (remove variable). p-values less than 0.05 were considered significant. All statistical tests and graphs were conducted using SPSS version 20  and MedCalc 12.3.0 .Association between CXCR4, CXCL12, CXCR7, and PD-1/PD-L1 expression cancer cells and patients\u2019 clinic-pathological features were analyzed applying chi-square and Mann\u2013Whitney U tests for categorical and continuous variables, respectively. Recurrence-free survival (RFS) was set as the time from diagnosis to the recurrence or last follow-up. Cancer-specific survival (CSS) was set as the time from diagnosis to death for cancer. Kaplan\u2013Meier method was performed to estimate the survival curve and logrank test for statistical comparison. Cox proportional hazards regression was utilized to test the effect of multiple dichotomous covariates (risk factors) on RFS and CSS; The backward method for variable selection was applied in the final model with a conventional A population of twenty patients diagnosed with advanced PDAC, evaluated through endoscopic-US-guided pancreatic fine-needle aspiration cytology (FNAC), was considered a validation cohort. The samples were collected and analyzed at Pathology Unit, Department of Mental and Physical Health and Preventive Medicine, University of Campania \u201cLuigi Vanvitelli\u201d, Naples, Italy, from January 2020 to October 2021. Three-micrometer sections were stained for CXCL12 immunocytochemical evaluation following hematoxylin/eosin reviewing and histological confirmation. All cases were retrospectively reviewed, and the cytology on direct smears showed moderately to richly cellular samples composed of three-dimensional aggregates of epithelial cells with severe cytological atypia. The cyto-block (CB) sections were also re-examined, with a cytological morphology substantially similar to that observed on direct smears; when necessary, immunocytochemical (ICC) analysis was performed, which showed positivity in the neoplastic cells for CK19 and CA 19\u20139. In all 20 cases selected, the overall morphological analysis was consistent with the diagnosis of pancreatic adenocarcinoma (Category VI sec. Papanicolaou System). ImmunoCytoChemistry (ICC) was carried out on CB sections and stained for CXCL12 as previously described.Clinical pathological characteristics of the 76 patients are presented in CXCR4 was highly expressed in 25 out of 76 tumors (32.9%) . CXCR4 mRepresentative low\u2013moderate and high CXCR4 expressions were reported C\u2013H. CXCRCXCR7 was highly expressed in 26 out of 76 (34.2%) tumors and lowly expressed or undetectable in 50 tumors (65.8%). The CXCR7 mean expression was 13.3 \u00b1 16.7% . Unlike CXCL12 was predominantly identified in the membrane in 20 out of 76 (26.3%) tumors. The CXCL12 mean expression was 3.7 \u00b1 7.20% . RepresePD-1 and its natural ligand PD-L1 were evaluated in 76 PDAC. While PD-1 was not detectable in cancer cells, it was only sporadically expressed (<1%) in focal lymphocyte infiltrate (data not shown). PD-L1 was predominantly identified at the membrane of cancer cells in 29/76 (38.2%) PDAC. PD-L1 mean expression was 3.4 \u00b1 7.1 . FigureC\u2013H. Tumor = 0.42, p = 0.004). The relation among the evaluated markers and clinic pathologic characteristics revealed that CXCR4-positive tumor and CXCR4-positive tumor-infiltrating inflammatory cells were associated with vascular invasion (p = 0.0421 and 0.045) as expected [p = 0.002)  : 20.19\u201329.85); 33.60 months for cancer-specific survival CSS : 28.907\u201338.998). A total of 71 PDAC were analyzed for RFS; 58/71 (81.7%) patients experienced recurrence, with mean RFS durations of 17.30 months, and 13 (18.3%) did not experience recurrence up to 60.52 months. Out of 76, 4 patients were excluded for cancer-unrelated death; 72 PDAC were analyzed for CSS, 56/72 (77.8%) with mean CSS durations of 25.30 months, and 16/72 (22.2%) patients were alive at a mean of 62.65 months. Univariate analyses of RFS and CSS are summarized in p = 0.0024; CSS: 21.9 months vs. 36.6 months p = 0.08, not significant) (Patients with high CXCR4 expression had a significantly worse outcome (RFS: 11.76 months vs. 26.0 months ificant) A.n = 48) (RFS: 11.05 months vs. 33.70 months p = 0.0071) (p = 0.0001) and CSS  (p = 0.0001)  and CSS (p = 0.017)  in R0 patient\u2019s subgroup (p = 0.0441) but not CSS (p = 0.0605)  ( 0.0071) A. CXCL12 0.0024) B. Moreovsubgroup B. Patien 0.0605) C. PD-L1  0.0605) D.p = 0.016) and CSS (0.047) in R0 patients, , perineural invasion (p = 0.0107), and CXCL12 (p = 0.00002) but not CXCR4, predicted shorter poor RFS (p = 0.00007), vascular invasion (p = 0.0067), and CXCL12 (p= 0.0062) but not CXCR7 nor CXCR4 expression predicted shorter poor CSS . The median age was 72 \u00b1 10 years (range 50\u201385), with 8 (40%) male and 12 (60%) female, and 17 (85%) of patients \u226560 years old. At diagnosis, the majority of patients (85%) had an advanced stage. CXCL12 was detected in 70% (14/20) PDACs predominantly at the membrane and cytoplasm of cancer cells  , showing a median overall survival of 6 months. The patients with a high sharp, consistent distribution of CXCL12 expression showed short CSS with a median survival of 3 months, while low/negative CXCL12 expressing displayed short CSS with a median survival of 12 months (= 0.029) .p \u2265 0.05) as for NCCN 2022 guidelines. In our analysis, chronic pancreatitis was a \u201cpotential risk factor\u201d  not retained, for irrelevance or redundancy, in the multivariate analysis. As for PD-L1, early studies showed prognostic significance for overall survival with >5 or 10% cut-off [The role of the CXCR4-CXCL12-CXCR7 axis and PD-1/PDL-1 was addressed in tumor/stromal cells in 76 consecutive single-center patients undergone surgery between January 2014/March 2015 and followed for 5 years until January 2021. We demonstrated that the entire axis CXCR4-CXCL12-CXCR7 was overexpressed in PDAC neoplastic cells as compared to TME cells. Moreover, CXCL12 significantly correlated with poor prognosis in an unrelated cohort of 20 FNAC from PDAC patients. CXCR4 was expressed by acinar cells and islets of Langerhans, surrounded by extensive dense fibrotic stroma or desmoplasia . Trefoil cut-off ,50. Alth cut-off .MMR) proteins) and sample size. In agreement with the herein reported data, Diana et al. examined the prognostic value of PD-1 and PD-L1, together with CD8+ tumor-infiltrating lymphocytes (TILs) and FOXP3+ Tregs in 145 PDAC samples, describing PD-L1 expression not prognostic [Karamitopoulou reported PD-L1 expression in about one-third of 349 samples. The authors concomitantly analyzed PD-L1, PD-1, CD3, CD4, CD8, FoxP3, and CD68, reporting that PD-L1, present with T-cell infiltration, improved overall survival . Herein,ognostic . Wang etognostic . As relaPD-L1 was recently reported in stromal cells in PDAC , and als"}
+{"text": "Accurate polyp size measurement is important for guideline conforming choice of polypectomy techniques and subsequent surveillance interval assignments. Some endoscopic tools (forceps or endoscopic rulers [ER]) exist to help with visual size estimation. A virtual scale endoscope (VSE) has been developed that allows superimposing a virtual measurement scale during live endoscopies.Our aim was to evaluate the performance of VSE when compared to ER and forceps-based measurement.We conducted a randomized trial to evaluate the relative accuracy of size measurement of simulated colorectal polyps when using: VSE, ER, and forceps. Six endoscopists performed 60 measurements randomized at a 1:1:1 ratio using each method. Primary outcome was relative accuracy in polyp size measurement. Secondary outcomes included misclassification of sizes at the 5, 10, and 20mm thresholds.A total of 360 measurements were performed. The relative accuracy of biopsy forceps, ER, and VSE was 78.9% (95%CI=76.2-81.5), 78.4% (95%CI=76.0-80.8), and 82.7% (95%CI=80.8-84.8). VSE had significantly higher accuracy compared to biopsy forceps (p=0.02) and ER (p=0.006). VSE misclassified a lower percentage of polyps >5mm as \u22645mm (9.4%) compared to forceps (15.7%) and ER (20.9%). VSE misclassified a lower percentage of \u226520mm polyps as <20mm (8.3%) compared with forceps (66.7%) and ER (75.0%). 25.6%, 25.5%, and 22.5% of polyps \u226510mm were misclassified as <10mm with ER, forceps, and VSE, respectively.VSE had significantly higher relative accuracy in measuring polyps compared to ER or forceps assisted measurement. VSE improves correct classification of polyps at clinically important size thresholds.NoneR. Djinbachian: None Declared, M. Taghiakbari: None Declared, C. Haumesser: None Declared, M. Zarandi-Nowroozi: None Declared, M. Abou-Khalil: None Declared, S. Sidani: None Declared, J. Liu: None Declared, B. Panzini: None Declared, D. von Renteln Grant / Research support from: Daniel von Renteln is supported by a \u201cFonds de Recherche du Qu\u00e9bec Sant\u00e9\u201d (FRQS) career development award and has received research funding from ERBE, Ventage, Pendopharm, Fujifilm, and Pentax., Consultant of: Boston Scientific and Pendopharm,"}
+{"text": "All authors agree with this retraction.1. Kang, H., Ma, D., Zhang, J. et al. Long non-coding RNA GATA6-AS1 upregulates GATA6 to regulate the biological behaviors of lung adenocarcinoma cells. BMC Pulm Med 21, 166 (2021)."}
+{"text": "Cytochrome P450 family 8 subfamily B member 1 (CYP8B1) generates 12\u03b1-hydroxylated bile acids (BAs) that are associated with insulin resistance in humans.CYP8B1 in individuals without diabetes and identified carriers of complete loss-of-function (CLOF) mutations utilizing functional assays.To determine whether reduced CYP8B1 activity improves insulin sensitivity, we sequenced CYP8B1 mutation associated with lower fasting insulin in the AMP-T2D-GENES study. Exposure of primary human muscle cells to mutation-carrier CA/CDCA ratios demonstrated increased FOXO1 activity, and upregulation of both insulin signaling and glucose uptake, which were mediated by increased CDCA. Inhibition of FOXO1 attenuated the CDCA-mediated increase in muscle insulin signaling and glucose uptake. We found that reduced CYP8B1 activity associates with increased insulin sensitivity in humans.Mutation carriers had lower plasma 12\u03b1-hydroxylated/non\u201312\u03b1-hydroxylated BA and cholic acid (CA)/chenodeoxycholic acid (CDCA) ratios compared with age-, sex-, and BMI-matched controls. During insulin clamps, hepatic glucose production was suppressed to a similar magnitude by insulin, but glucose infusion rates to maintain euglycemia were higher in mutation carriers, indicating increased peripheral insulin sensitivity. Consistently, a polymorphic CLOF Our findings suggest that increased circulatory CDCA due to reduced CYP8B1 activity increases skeletal muscle insulin sensitivity, contributing to increased whole-body insulin sensitization.Biomedical Research Council/National Medical Research Council of Singapore.  Cyp8b1 show almost no 12\u03b1-hydroxylated BAs and elevated non\u201312\u03b1-hydroxylated BAs (Bile acids (BAs) are products of cholesterol catabolism, accounting for approximately 50% of daily cholesterol turnover in humans, and act as surfactants to promote intestinal lipid absorption . The priNR1H4) and membrane G protein\u2013coupled BA receptor  . Of these, 58 were predicted to be possibly or probably damaging by the functional prediction tool Polyphen 2.0 , and 41 were predicted to be damaging by SIFT . We generated all 100 variants in human CYP8B1 cDNA, quantified the product generated by each variant, and found a spectrum of defective CYP8B1 activities, classified as complete loss of function (CLOF) (<15% activity of wild-type), partial loss of function (PLOF) (15% to 85% activity of wild-type), or benign (>85% activity of wild-type)  . We idenCYP8B1 mutation carriers, and 41 age-, sex-, BMI-, and race-matched controls to our clinical study. P = 0.04) and APOB/APOA-I (P = 0.04) ratios, suggesting reduced risk for atherosclerosis. In addition, high-sensitivity C-reactive protein (hs-CRP) levels were decreased by approximately 50% in the mutation carriers (P = 0.06), suggesting reduced systemic inflammatory status and lower atherosclerotic risk in carriers  (P = 0.02), resulting in a 63% decrease in the CYP8B1 product/substrate ratio  ( 0.0002) . CYP8B1\u2019 0.0001) .P = 0.003) (P = 0.01), and both glycine- (P = 0.01) and taurine-conjugated (P = 0.048) CA were decreased (P = 0.03) (P = 0.01)  . Unconjuecreased . CDCA di = 0.03) . The CA/ = 0.01) . The BA CYP8B1 mutation carriers. Thus, we assessed whether CYP8B1 mutation carriers showed improved insulin sensitivity. Although fasting glucose was unchanged, fasting insulin was decreased by 28% in carriers  (P = 0.03) (P = 0.03), as well as QUICKI, another measure of insulin sensitivity (P = 0.04) , insulin levels were decreased in carriers  (P = 0.02) and the glucose infusion rate (GIR) was increased by 30% in carriers  (P = 0.02) (P = 0.04) and GIR by 25%  in carriers.12\u03b1-Hydroxylated BAs correlate with insulin resistance in humans , and 12\u03b1 = 0.03) . Accordi = 0.03) . The Mat = 0.04) , were in = 0.03) . During = 0.046) . The ins = 0.02) . The aboP = 0.38) (P = 0.48) (P = 0.58) (P = 0.19)  , glucone = 0.48) , and gly = 0.58)   , indicatCYP8B1 mutations have improved insulin sensitivity, we performed association analyses using the Type 2 Diabetes Knowledge Portal (http://www.type2diabetesgenetics.org), which enables association analyses between coding variation and glycemic traits in 45,231 exomes (CYP8B1 variants (R26X) was both a CLOF mutation (0.6% activity compared with wild-type CYP8B1), common in our study cohort , and present in several copies in the AMP-T2D-GENES data set. Association analyses showed that carriers of R26X had significantly lower fasting insulin levels after adjusting for BMI . These data are consistent with our findings of increased insulin sensitivity in the face of decreased CYP8B1 activity in the CYP8B1 mutation carriers.To confirm that humans with CLOF 1 exomes . Of the variants , only 1 P = 0.65) (P = 0.04) and GLP-1 during MMTT  were decreased  . Glucago = 0.65) . Both faecreased , in lineIRB being the predominant isoform in insulin target tissues, including muscle   (256) is cytoplasmic, whereas dephosphorylated FOXO1 is retained in the nucleus where it functions as a transcription factor (256)FOXO1 was decreased in muscle cells exposed to the carrier BA mix in response to insulin, suggesting increased nuclear localization and transcriptional activity of FOXO1 in carriers  (473)AKT was increased by 116% in muscle cells treated with the carrier CA/CDCA ratio in response to insulin  (P = 0.004)  , suggest= 0.002) . FOXO1 e = 0.04) . Phosphon factor . Phospho= 0.036) . Phospho= 0.036) . Phospho= 0.029) . Additio= 0.004) , confirm473)AKT was increased approximately 2-fold in muscle cells treated with the median carrier CA/CDCA ratio in response to insulin  (P = 0.009)  and lowest insulin sensitivities (CA/CDCA = 21:29). Thus, we assessed whether the median CA/CDCA ratio of the mutation carriers (CA/CDCA = 4.5:45.5) would increase muscle cell insulin signaling and glucose uptake when compared with the median CA/CDCA ratio of noncarrier controls (CA/CDCA = 8.3:41.7). Phospho(S 0.0008) . In line= 0.009) . These dCYP8B1 mutation carriers, suggesting that increased CDCA may be sufficient to increase muscle insulin sensitivity. Thus, we next assessed whether CDCA alone was sufficient to increase muscle insulin signaling. Exposure of muscle cells to CDCA, but not CA, increased phospho(S473)AKT in response to insulin   (Insr and Foxo1 expression (256)FOXO1 levels  (473)AKT levels  , indicatP = 0.8) . Moreovepression , and decP = 0.2) . To furtCYP8B1 mutation carriers (473)AKT or glucose uptake AKT/AKT ratio    , and on P = 0.6) . These dCYP8B1 improves insulin sensitivity in humans. Although BA synthesis involves several enzymes, and complex feedback and feedforward mechanisms are involved in the regulation of BA metabolism . It is possible that CDCA modulates intracellular signaling through as yet unidentified muscle-specific cell surface receptors or BA transporters. Our data suggest that CDCA modulates muscle insulin signaling through increasing muscle FOXO1 activity, and inhibition of FOXO1 reversed the increased CDCA-mediated muscle insulin signaling. How CDCA decreases muscle FOXO1 phosphorylation, thus increasing its nuclear retention and transcriptional activity, remains unclear. One possible mechanism by which CDCA may reduce phospho-FOXO1 is by increasing the activity of phosphatases acting on FOXO1. BAs do modulate other phosphatases such as Src-homology 2 domain\u2013containing tyrosine phosphatase 2 (SHP2)  .The reason for the potential selectivity of CDCA for skeletal muscle in regulating insulin sensitivity is unclear. However, FOXOs show tissue-specific protein interactions to modulate their functions in metabolic regulation . DistincCYP8B1 mutation carriers showed decreased total cholesterol/HDL-C and APOB/APOA-I ratios, suggesting that CYP8B1 inhibition may reduce type 2 diabetes without proatherogenic lipid changes. In addition, hs-CRP levels were decreased by approximately 50%, and hepatic fat was decreased by 30% in the CYP8B1 mutation carriers. These observations are consistent with those in \u2013/\u2013Cyp8b1 mice, which showed increased HDL-C, decreased LDL-C, and reduced atherosclerotic lesions when fed atherogenic diets , an analog of CDCA, was assessed in type 2 diabetics . Increasic diets , 27. Addgression , suggestCYP8B1 mutation carriers.The CYP8B1 product, 7\u03b1,12\u03b1-dihydroxy-4-cholesten-3-one, was not decreased in plasma of mutation carriers. However, downstream 12\u03b1-BAs were decreased by 52%, suggesting that the 12\u03b1-hydroxylase function of CYP8B1 in the liver was indeed reduced. Significantly increased unconjugated CDCA was also not observed in mutation carriers, although the ratio of CDCA and its conjugates to total BAs was increased. The reasons for this are unclear. In the mutation carriers, plasma BAs were quantified after an overnight fast. However, it has been shown that circulatory BA levels increase postprandially, with CDCA showing the largest increase (up to 5-fold) . Further\u2013/\u2013Cyp8b1 mice are viable, with no apparent adverse phenotypes, suggesting that the absence of CYP8B1 may not be harmful. We did not identify compound heterozygous or homozygous carriers of CLOF mutations, and almost all CLOF mutations were identified in only 1 or 2 heterozygotes. Thus, homozygous or compound heterozygous CLOF mutation carriers are likely to be extremely rare. One CLOF mutation, R26X, was found at 1.6% frequency in Malays. Assuming Hardy-Weinberg equilibrium, sequencing of approximately 13,000 Malays is required to identify a single R26X homozygote. Thus, it is unsurprising that we identified no homozygous or compound heterozygous CLOF mutation carriers. Additionally, in a large publicly aggregated database (gnomAD), no individuals harboring homozygous predicted CLOF mutations were reported, confirming that these individuals are extremely rare.We establish a fundamental role for CYP8B1-mediated changes in BA composition in the regulation of peripheral insulin sensitivity. We show that reduced activity of CYP8B1 is efficacious in increasing insulin sensitivity in humans, and mechanistically link CDCA signaling to muscle insulin sensitivity. We demonstrate here a target for future therapeutic intervention for diabetes.Detailed methods are provided in the supplementary material.CYP8B1 coding region was Sanger sequenced in population cohorts of nondiabetics from the Singapore Eye Research Institute and the Saw Swee Hock School of Public Health, National University of Singapore. Sequences were assembled in Sequencher (Gene Codes) and aligned to the human CYP8B1 reference sequence (GenBank AF090320.1). All nonsynonymous variants, frameshifts, and insertions/deletions were generated in human CYP8B1 cDNA and functionally characterized.The CYP8B1 cDNA inserted into pcDNA3.1(+) (Invitrogen) was used as the template for site-directed mutagenesis. The neomycin resistance cassette in pcDNA3.1(+) was replaced with green fluorescent protein (Gfp) in order to assess transfection efficiency of the variants. Mutagenesis primers were designed for each of the CYP8B1 variants, and PCR performed with conditions adjusted for each primer pair, amplifying the entire plasmid. PCR products were isolated from the template by Dpn1 digestion, transformed into DH5\u03b1 competent cells, and amplified plasmids were extracted by E.Z.N.A. Plasmid Mini Kit II (Omega Bio-tek Inc). The mutant CYP8B1 cDNAs were sequence confirmed, subcloned into the pcDNA3.1(+)-Gfp vector, and used for the in vitro assay. For the in vitro functional assay, HEK293T (ATCC) cells in 6-well plates were transiently transfected with the plasmids harboring wild-type CYP8B1, variant CYP8B1, or empty vector using X-tremeGENE HP (Roche). After 24 hours, fresh DMEM with 10% FBS containing 10 \u03bcmol/L of the CYP8B1 substrate 7\u03b1-hydroxy-4-cholesten-3-one  was added to the cells at 3 mL/well. After 4 hours, media were collected, centrifuged, and frozen until quantification of 7\u03b1,12\u03b1-dihydroxy-4-cholesten-3-one by LC/MS . Cells were washed, centrifuged, and frozen for Western immunoblotting.CYP8B1 mutations and age-, sex-, race-, and BMI-matched nonmutation carrier controls from the same cohorts were recruited at a ratio of 1 carrier to 2 controls for metabolic studies. Five participants only had 1 matched control. The studies of BAs and insulin phenotypes excluded individuals with BMI greater than 30, the World Health Organization definition of obesity, since obesity modulates insulin sensitivity. Individuals were also excluded if they had type 2 diabetes mellitus, renal impairment, elevated serum aspartate aminotransferase or alanine aminotransferase, chronic liver disease, or medications or previous gastrointestinal surgery that may alter glucose or BA metabolism  were measured in plasma following overnight fasts using ultraperformance liquid chromatography\u2013multiple reaction monitoring/mass spectroscopy (UPLC-MRM/MS) (University of Victoria-Genome BC Proteomics Centre) as described previously .After 8-hour fasts, venous blood was collected for plasma glucose, insulin, and GLP-1 measurements at 0, 15, 30, 60, 90, and 120 minutes after ingestion of liquid mixed meal .2H2O  for quantification of gluconeogenesis and glycogenolysis. To measure hepatic glucose production, primed-constant infusion of -glucose was performed. Insulin was infused at 40 mU/m2 body surface area/min for 180 minutes. Blood glucose was measured every 5 minutes. Blood for plasma insulin measurement was obtained every 30 minutes. Dextrose 20% (wt/vol) enriched with -glucose was infused at a variable rate to maintain blood glucose at 100 mg/dL with a coefficient of variation of less than 5%.Participants ingested 2 doses of CYP8B1 mutant R26X, selected because it was the most frequent in our study cohort, were performed using the publicly available Type 2 Diabetes Knowledge Portal (http://www.type2diabetesgenetics.org). Associations between R26X and fasting insulin, adjusted for BMI, age, and sex were calculated using the portal\u2019s Genetic Association Interactive Tool (GAIT), in which single-variant and gene-level association analysis can be conducted in 45,231 exomes from the AMP-T2D-GENES study.Association analyses for the CLOF Human adult skeletal muscle cells (Cell Applications) were differentiated following manufacturer\u2019s instructions and treated with a 50 \u03bcmol/L CA/CDCA mixture at the ratio of carrier or control, or dimethyl sulfoxide (DMSO) for 24 hours. To quantify AKT and FOXO1 phosphorylation subsequent to insulin stimulation, differentiated cells were treated with 100 nmol/L insulin . For glucose uptake assays, BA-treated cells were incubated with and without 100 nmol/L insulin, glucose uptake was quantified using fluorescence, and protein content was determined for normalization of glucose uptake.t tests. Non-normal data are reported as median (IQR) and were analyzed using the nonparametric Mann Whitney U test. Fisher\u2019s exact tests were used for categorical data, 1-way ANOVA was used for analyses of more than 2 groups, and 2-way ANOVA was used for repeated measures. All tests were 2-sided, and performed using GraphPad Prism 9.0. A P value of less than 0.05 was considered significant.All human data were first assessed for normality. Non-normal data were log transformed. Normal data are reported as mean \u00b1 SEM and were analyzed using parametric unpaired The human study received approval from the SingHealth Centralized Institutional Review Board. All participants provided written informed consent prior to the study.RRS, MRH, AKG, and HCT conceptualized the study. RRS and HCT acquired funding. RRS, HCT, FK, JLG, RMVD, RST, KAR, and CYC supervised the study. RRS wrote the original draft of the manuscript. SZ, RC, MC, DCM, BJC, KPS, THVD, JP, LJT, SVH, BR, PC, JLG, SC, JF, and XS conducted the investigation and analyses. JLG, AKG, FK, SC, JF, KAR, XS, HCT, and RRS provided resources. SZ, AKG, FK, CD, HCT, and MRH reviewed and edited the manuscript."}
+{"text": "Xanthomonas is a genus of gram-negative bacterium containing more than 35 species. Among these pathogenic species, Xanthomonas albilineans  is of global interest, responsible for leaf scald disease in sugarcane. Another notable Xanthomonas species is Xanthomonas sachari (Xsa), a sugarcane-associated agent of chlorotic streak disease.Xanthomonas strains was evaluated by disease index (DI) and Area Under Disease Progress Curve (AUDPC) in the susceptible inoculated plants (GT 46) and clustered into three groups of five highly potent, seven mild virulent, and twelve weak virulent strains. The highly potent strain  and its weak virulent related strain  were sequenced, assembled, and annotated in the circular genomes. The genomic size of JG43 was smaller than that of DD13. Both strains (JG43 and DD13) lacked a Type III secretory system (T3SS) and T6SS. However, JG43 possessed Salmonella pathogenicity island-1 (SPI-1). More pathogen-host interaction (PHI) genes and virulent factors in 17 genomic islands (GIs) were detected in JG43, among which six were related to pathogenicity. Albicidin and a two-component system associated with virulence were also detected in JG43. Furthermore, 23 Xanthomonas strains were sequenced and classified into three categories based on Single Nucleotide Polymorphism (SNP) mutation loci and pathogenicity, using JG43 as a reference genome. Transitions were dominant SNP mutations, while structural variation (SV) is frequent intrachromosomal rearrangement (ITX). Two essential genes (rpfC/rpfG) of the two-component system and another gene related to SNP were mutated to understand their virulence effect. The mutation of rpfG resulted in a decrease in pathogenicity.The virulence of 24 Xanthomonas strains and variations by 23 Xanthomonas strains. We sequenced, assembled, and annotated the circular genomes of Xal JG43 and Xsa DD13, identifying diversity detected by pathogenic factors and systems. Furthermore, complete genomic sequences and sequenced data will provide a theoretical basis for identifying pathogenic factors responsible for sugarcane leaf scald disease.These findings revealed virulence of 24 The online version contains supplementary material available at 10.1186/s12864-022-08900-2. Xanthomonas  is a large genus of gram-negative, yellow-pigmented bacteria associated with plants. The genus, which locates at the base of the Gamma proteobacteria, comprises 27 species that cause severe diseases in\u2009~\u2009400 plant hosts, including a wide variety of economically important crops, such as rice, citrus, banana, cabbage, tomato, pepper, and bean \u2009\u00d7\u2009100.Furthermore, the area under the disease-progress curve (AUDPC) value was calculated , 86.\\docyi is the severity of the symptoms at the ith observation; xi\u2014day at the ith observation, and n\u2014the total number of observations.where AUDPC is the area under the disease progress curve, X. albilineans JG43 was sequenced by Oxford Nanopore Technologies (ONT) and assembled using Canu (V1.5) [X. sacchari DD13 was sequenced using SMRT II sequencing technology , and a complete circular bacterial chromosome was assembled using HGAP software [The genome of u (V1.5) . Assemblsoftware . The gensoftware .https://www.repeatmasker.org/) [http://eddylab.org/infernal/) [http://lowelab.ucsc.edu/tRNAscan-SE/) [https://www.ebi.ac.uk/Tools/psa/genewise/) [http://www.pathogenomics.sfu.ca/islandviewer/) [http://phispy.sourceforge.net/) [Assembled genome was analyzed to identify the repeat sequences that were searched against the known repeat sequence database (Repbase) in the bacterial genome using RepeatMasker (V4.0.5) (er.org/) . Non-codfernal/) , while tcan-SE/) . The Gennewise/) . Genomicviewer/) , and proge.net/) .www.ncbi.nlm.nih.gov/refseq/about/nonredundantproteins/) [https://www.genome.jp/kegg/) [https://www.expasy.org/resources/uniprotkb-swiss-prot) and TrEMBL (http://www.bioinfo.pte.hu/more/TrEMBL.htm) [https://www.ebi.ac.uk/Tools/hmmer/) [https://www.blast2go.com/) [The genes were blasted against the databases of non-redundant proteins (oteins/) , Kyoto Ep/kegg/) , Swiss-PMBL.htm) . Pfam fu/hmmer/) . Functiogo.com/) , 101.http://www.cazy.org/) using HMMER software [http://www.cbs.dtu.dk/services/TMHMM/), Kohgpi (http://gpi.unibe.ch/) [http://www.cbs.dtu.dk/services/SignalP/) [https://card.mcmaster.ca/) using RGI in CARD Database [http://www.mgc.ac.cn/VFs/) [Carbohydrate EnZymes genes were annotated against Carbohydrate Active EnZymes Database (CAZyme) (software . Transmeibe.ch/) , and SigignalP/) . TransmeDatabase . The vircn/VFs/) .Xal JG43 by GATK software (https://gatk.broadinstitute.org/hc/en-us) [X. albilineans strains was sequenced using Illumina Novaseq 6000 platform, with an average coverage of 339\u2009\u00d7\u2009. The redundant reads (MarkDuplicates) were filtered by Picard software to ensure the detection accuracy of clean reads [X. albilineans, which were used to construct a phylogenetic tree. After removing ambiguous positions, a final dataset of 17,935 SNPs was generated for each sequence, which was aligned through the neighbor-joining method of MEGA7 software, utilizing the bootstrap value of 1000 replicates [Single nucleotide polymorphism (SNP) was called against the reference genome of c/en-us) . The genan reads . A totalplicates , 110.Xal genomes with Xal JG43, and the aligned reads were sorted using Samtools (V1.12) [Minimap2 (V2.17) was used to align the 23 sequenced  (V1.12) . BCFtool (V1.12) . BEDTool (V1.12) . Circos  (V1.12) .The single-base mutation was performed by the PNA-directed PCR clamping , and twopK18mobSacB plasmid to form a recombinant plasmid. The recombinant plasmid was digested, verified, and sequenced. The recombinant plasmid was introduced into the host bacterium by electro-transformation. The upstream and downstream of the target gene underwent single homologous exchange and double homologous exchange with the homologous fragment of the host bacterium. A 10% sucrose was used to screen double homologous exchange. Furthermore, internal and external primers were also used to screen double homologous exchange. Finally, the target gene was deleted , (c) and (d) show leaf scald symptoms after X. albilineans invade sugarcane; (b) Colony of X. albilineans isolated from diseased sugarcane plant; Right side: X. sacchari cause chlorotic streak disease. (a), (c) and (d) show chlorotic streak symptoms after X. sacchari infect sugarcane; (b) Colony of X. sacchari isolated from the diseased sugarcane plant. Fig.S2. Type III secretion system (T3SS) (a), and SPI-1 family (b) of six Xanthomonasspecies. Fig. S3. Type IV secretion system (T4SS) (a), T5SS and T6SS (b) of six Xanthomonas species. Fig. S4. Potential pathogenic factors of six Xanthomonas species, including CRISPR system, Lipopolysaccharide transport system protein, Glycogen, Type III secretion regulators, Two-component system regulators, Three-component system, and TALEs. Fig. S5. Verification of rpfC and rpfH mutations. (a) PCR amplification from the upstream and downstream 500 bp of rpfC. M: 2000 bp; Lane 1: rpfC Gene left arm; Lane 2: rpfC gene right arm. (b) Validation of enzymic fragment ligated with PK18mobsacB, a 500bp upstream and downstream fragment of rpfC gene. M: 5000 bp; Lane 1,2,3\uff1aValidation of rpfC recombinant plasmid fragment by enzyme digestion; Lane 4 not included in this experiment. (c) PCR amplified from mutants and its wild type JG43. M:1000 bp; Lane 1, 2, 3; PCR fragment amplified with mutants; Lane 4: PCR fragment amplified with JG43 as template; Lane 5: Water control; Lane 6, 7, 8: Internal primer verification of the target fragment missing in 123, none, which proves the successful deletion of rpfC gene; Lang 9: Internal primer fragment of PCR amplified with JG43 as template. (d)PCR validation of rpfHgene. M:1000 bp; Lane 1:Xcc8004; Lane 2: DD13; Lane 3: JG43; Lane 4: Water control;Lane 5: not included in this experiment. Fig. S6. rpf gene cluster of six Xanthomonas species. Fig. S7. PCR validation of single-base SNPs mutations. (a) PCR validation of single-base SNP mutations of candidate genes in FS 12. M:2000 bp; Lane 1: 1312440-G-C-L; Lane 2: 1312440-G-C-R; Lane 3: 1316566-A-G-L; Lane 4: 1316566-A-G-R; Lane 5: 1316572-A-G-L; Lane 6: 1316572-A-G-R; Lane 7: 1316840-G-A-L; Lane 8: 1316840-G-A-R; Lane 9: 1316855-T-C-L; Lane10: 1316855-T-C-R; Lane 11: 1316974-A-G-L; lane12: 1316974-A-G-R; lane13: 1317164-T-C-L; Lane 14: 1317164-T-C-R; Lane 15: 3055754-G-A-L; Lane 16: 3055754-G-A-R. (b) PCR validation of single-base SNP mutations of candidate genes in FS12 (Lanes 1 and 2), FS25 (Lanes 3 and 4), FS63 (Lanes 5 and 6) and NM10 (Lanes 7 and 8). M:2000 bp; Lane 1: 3055807-G-C-L; Lane 2: 3055807-G-C-R; Lane 3: 2749153-C-T-L; Lane 4: 2749153-C-T-R; Lane 5: 1508978-C-A-L; Lane 6: 1508978-C-A-R; Lane 7:1510223-A-C-L; Lane 8: 1510223-A-C-R. (c) SNP point mutation fusion fragment in FS12. M:2000 bp; Lane 1: 1312440-G-C; Lane 2: 1316566-A-G; Lane 3: 1316572-A-G; Lane 4:1316840-G-A; Lane 5: 1316855-T-C; Lane 6:1316974-A-G; Lane 7: 1317164-T-C. (d) SNP point mutation fusion fragment in FS12. M: 2000 bp; Lane 1:3055754-G-A; Lane 2:3055807-G-C; Lane 3: 3055836-T-G. (e) SNP point mutation fusion fragment in FS25 (Lane 1:2749153-C-T), FS63 (Lane 2: 1508978-C-A), and NM10 (Lane 3: 1510223-A-C). M:5000 bp.Additional file 2: Table S1. Basic information of Xal JG43 and Xsa DD13. Table S2. Repeat contents from genome sequence of Xal JG43 and Xsa DD13. Table S3. Genes in plasmid from the genome of Xal JG43. (.xls ) Table S4. Carbohydrate-active enzymes (CAZys) in Xal JG43 and Xsa DD13. Table S5. Comparative pathogenomics of X. albilineans JG43 and its related X. sacchari DD13. (.xls ) Table S6. Genomic island and prophages of Xal JG43 and Xsa DD13. (.xls ) Table S7. Resequencing of 23 X. albilineans strains. Table S8. Genomic variations (SNPs and SVs) obtained from 23 sequenced X. albilineans strains against JG43. Table S9. SNP mutations in 23 strains of X. albilineans. Table S10. Mutations at the DNA level in 23 strains of X. albilineans. Table S11. List of primers used in this study.Additional file 3."}
+{"text": "Scientific Reports 10.1038/s41598-022-18110-1, published online 18 August 2022Correction to: The Funding section in the original version of this Article was incomplete.\u201cYayasan Universiti Teknologi PETRONAS (YUTP), Cost Center 015LC0-278 received by D. L. C. C.\u201dnow reads:\u201cYayasan Universiti Teknologi PETRONAS (YUTP), Cost Center 015LC0-278 and NCRF 015MCO-030 received by D. L. C. C.\u201dThe original Article has been corrected."}
+{"text": "Correction: Inflamm Regen 42, 37 (2022)https://doi.org/10.1186/s41232-022-00217-7Following publication of the original article , authorsThe original version was:1*, Endo Yushiro1, Koga Tomohiro1,2, Yoshiura Koh-ichiro3 and Migita Kiyoshi4Atsushi KawakamiThe correct authorship has been updated in this Correction.The original article  has been"}
+{"text": "Concerns have been raised about ecological momentary assessment (EMA) acceptability among patients with schizophrenia spectrum disorders (SSD), which is of major relevance during the e-Mental health-focused COVID-19 pandemic.To investigate i) the levels of adherence to a passive smartphone-based EMA tool, the Evidence-Based Behavior (eB2), among SSD patients; and ii) putative predictors of this.Sample: SSD (F20-29-ICD10) outpatients, age 18-64, without financial incentives, recruited over 17/06/2019-11/03/2020 at the Hospital Universitario Fundaci\u00f3n Jim\u00e9nez D\u00edaz . Those who accepted the eB2 installation -users- and those who did not -non-users- were compared in sociodemographic, clinical, premorbid adjustment, neurocognitive, psychopathological, insight and metacognitive variables by a multivariable binary logistic regression model.Sample (N=77): n=41 males; age: 47.69\u00b19.76 years, n=24 users (31.2%). n=14 users (70%) had the eB2 installed at follow-up (median=14.50 weeks).2=25.296,df=6,p<0.001. Nagelkerke-R2=44.7%. Correctly classified: 76.9%, users:54.5%, non-users:88.4%.XAcceptability of a smartphone-based EMA application among SSD patients was low. Age (young) and good premorbid adjustment predicted acceptability. e-Mental Health methods need to be tailored for patients with SSD. Otherwise, these highly vulnerable individuals may be neglected by e-health-based services in the post-COVID-19 years ahead."}
+{"text": "We assessed cross-reactivity to BA.1, BA.2, and BA.5 of neutralizing antibodies elicited by ancestral, Delta, and Omicron BA.1 SARS-CoV-2 infection in mice. Primary infection elicited homologous antibodies with poor cross-reactivity to Omicron strains. This pattern remained after BA.1 challenge, although ancestral- and Delta-infected mice were protected from BA.1 infection. The SARS-CoV-2 Omicron variant  emerged nearly 2 years after the ancestral strain was identified  of SARS-CoV-2/Australia/Vic/01/20 , SARS-CoV-2/Australia/Vic/18440/2021 (Delta), and SARS-CoV-2/Australia/NSW/RPAH-1933/2021 (Omicron BA.1) strains in 7- to 9-week-old female K18hACE2 transgenic mice (2 TCID50), selected so that the mice would survive primary infection . We mock-infected 15 mice with phosphate-buffered saline (PBS). We collected blood on day 27 after primary infection and then challenged mice with 104 TCID50 of Omicron BA.1 virus. We collected lungs and nasal turbinates (NTs) 2 and 4 days after challenge; we weighed and monitored 5 mice per group for clinical signs for 14 days , survived without weight loss. The control group had mean virus titers of 102.6 (day 2) and 102.7 (day 4) in NTs and 103.7 (day 2) and 103.5 (day 4) TCID50/organ in lungs after Omicron BA.1 challenge. After primary infection, all Omicron BA1\u2013infected mice survived without major weight loss, but 1 ancestral strain\u2013infected and 5 Delta-infected mice died during days 8\u201313. After challenge with 10Consistent with other reports , panel CThe homologous responses were strongest to ancestral (geometric mean titer [GMT]\u00a0709), followed by Delta (GMT\u00a0129), and were lowest to BA.1 (GMT\u00a083) . The lowPrimary Omicron BA.1 infection did not induce heterologous neutralizing activity against ancestral, Delta, BA.2, or BA.5 viruses . In contDespite the absence of detectable BA.1 virus in the respiratory tract tissues after secondary infection in mice previously infected with ancestral or Delta , panel Chttps://doi.org/10.1101/2022.01.03.21268582). A boost occurred in preexisting SARS-CoV-2 neutralizing antibodies to ancestral and Delta but not in cross-reactivity to Omicron, probably because more epitopes are shared between ancestral and Delta than between those strains and Omicron. Serologic data from humans suggest that >3 exposures to ancestral strains as infection or vaccination or a combination are needed to induce cross-reactive antibodies to BA.1 (https://doi.org/10.1101/2022.05.10.22274906), we did not detect cross-reactive neutralizing antibodies after primary infection with ancestral and Delta strains. Protection from replication of the Omicron BA.1 strain despite the lack of cross-reactive neutralizing antibodies may be attributable to mucosal immunity or T-cell responses in ancestral strain\u2013infected and Delta-infected mice (Our observations are consistent with BA.1 being antigenically distinct from the ancestral and Delta strains (K. van der Straten K et al., unpub. data, Additional information about SARS-CoV-2 Omicron BA.1 challenge after ancestral or Delta infection in mice."}
+{"text": "Panax notoginseng (Burkill) F. H. Chen is not completely known. Morphological traits, N use and allocation, photosynthetic capacity and saponins accumulation were evaluated in two- and three-year-old P. notoginseng grown under different N regimes. The number and length of fibrous root, total root length and root volume were reduced with the increase of N supply. The accumulation of leaf and stem biomass (above-ground) were enhanced with increasing N supply, and LN-grown plants had the lowest root biomass. Above-ground biomass was closely correlated with N content, and the relationship between root biomass and N content was negatives in P. notoginseng (r = \u22120.92). N use efficiency-related parameters, NUE , NC (N content in carboxylation system component) and Pn (the net photosynthetic rate) were reduced in HN-grown P. notoginseng. SLN (specific leaf N), Chl (chlorophyll), NL (N content in light capture component) increased with an increase in N application. Interestingly, root biomass was positively correlated with NUE, yield and Pn. Above-ground biomass was close negatively correlated with photosynthetic N use efficiency (PNUE). Saponins content was positively correlated with NUE and Pn. Additionally, HN improved the root yield of per plant compared with LN, but reduced the accumulation of saponins, and the lowest yield of saponins per unit area (35.71 kg\u00b7hm\u22122) was recorded in HN-grown plants. HN-grown medicinal plants could inhibit the accumulation of root biomass by reducing N use and photosynthetic capacity, and HN-induced decrease in the accumulation of saponins (C-containing metabolites) might be closely related to the decline in N efficiency and photosynthetic capacity. Overall, N excess reduces the yield of root and C-containing secondary metabolites (active ingredient) in N-sensitive medicinal species such as P. notoginseng.Nitrogen (N) is an important macronutrient and is comprehensively involved in the synthesis of secondary metabolites. However, the interaction between N supply and crop yield and the accumulation of effective constituents in an N-sensitive medicinal plant  Nitrogen (N) is a determinant nutrient for plant biomass or crop yield . Yellow e.g., carboxylation and bioenergetics components), results in a lowed yield of N-excess Brassica campestris L.  R. Br. grown under low N condition (Datura stramonium L. is significantly increased with an increase in soluble sugar and proline content (primary metabolites) under N-excess condition  balance and consequently change the content of C- and N-containing secondary metabolites in the medicinal species . Excessibacum L. . Correspondition . The N-condition . UnexpecPanax notoginseng (Burkill) F. H. Chen (Sanqi in Chineses) is a perennial medicinal plant and a member of the Araliaceae family, which is a typically shade-tolerant and N-sensitive plants  enhances the accumulation of biomass and saponins though optimizing root architecture and N uptake efficiency in P. notoginseng , moderate nitrogen (MN) and high nitrogen (HN). We hypothesized that (i) root biomass of P. notoginseng might be reduced accompanying with HN-driven inhibition on photosynthetic capacity and NUE (N use efficiency); (ii) HN-driven decrease in saponin accumulation might be reflected by the C/N imbalance; (iii) N stress might reduce the yield of P. notoginseng.The present study aimed to shed light on an interaction between N availability and crop yield and saponins accumulation in the medicinal plant 2O) was 6.84, total N content was 0.17%, total phosphorus (P) was 0.23%, the available P content was 11.04 mg\u00b7kg\u22121, total potassium (K) was 0.24%, and the available K content was 127.32 mg\u00b7g\u22121.The study was conducted at the Yunnan Agricultural University teaching and experimental farm in Kunming, China , with an average annual rainfall and average annual temperature of about 1,006.7 mm and 14.5 \u00b0C, respectively. The properties of raw soil physical and chemical was determined as described by P. notoginseng, and the full sunlight irradiance is about 10%  provided one-year-old P. notoginseng seedlings. Subsequently, healthy and uniform seedlings were transplanted into a plastic flowerpot (30 cm \u00d7 25 cm \u00d7 20 cm) with each containing three rootstocks (\u22122), high nitrogen ) were designed (2O5), calcium superphosphate  and potassium sulfate (52% K2O), respectively. The same amounts of P (225 kg\u00b7P2O5\u00b7hm\u22122) and K (450 kg\u00b7K2O\u00b7hm\u22122) fertilizers were used in all treatments with the exception of the N fertilizer. Fertilization was applied in four times a year . In each pot, basal doses of P and K at the rates of 0.45 and 0.90 g, respectively , were applied at time while N was applied according to the treatments. N fertilizer rate 0 (LN), 0.45 (MN) and 0.90 (HN) g\u00b7pot\u22121 .A permeable black plastic net was used to create a shade-house for bout 10% . Meanwhibout 10% . Permeabotstocks . There wdesigned , and eacAt November, the two- and three-year-old plants were sampled from the experimental farm and then separated into root , stem and leaf in room. The length, width, and area of leaf were measured by LI-3000 leaf-area meter . Root tuber and stem diameter were measured by vernier caliper. Plant height, grown breadth, the length of main root and total root were determined as described by The samples were dried at 60 \u00b0C for 96 h. Dry matter was determined, and these results were used to calculate the percentage of biomass allocation into leaf , stem , roots , as well as root to shoot ratio (RSR). The root yield of per plant and economic yield (root yield of per hectare) were calculated based on root biomass data.P. notoginseng leaves were soaked. A standing period of 3 h was followed by a centrifugation of 3,000 g\u00b7min\u22121 for 10 min. A JASCO V-670 spectrophotometer  was used to measure absorbance at 665 and 649 nm wavelengths. Chl a, Chl b and Chl a/Chl b were analyzed as described by In 15 mL of acetone-ethanol mixture (2:1 v/v), 0.5 g of fresh 2 concentration of 10%, 25 \u00b0C, 500 \u03bcmol\u00b7photons\u00b7m\u22122\u00b7s\u22121 and 400 \u03bcmol\u00b7CO2\u00b7mol\u22121, respectively. Photosynthetic gas exchange parameters  were collected as previously described in LI-6400XT photosynthesis system  was used to determine photosynthetic gas exchange parameters. Set with a blue light ratio, temperature, photosynthetic photon flux density (PPFD) and COVcmax (maximum carboxylation efficiency), Jmax (maximum electron transfer rate), SLN and Chl contents, NC (N content in carboxylation system component), NB (N content in bioenergetics component) and NL (N content in light-harvesting systems component) were analyzed according to the method described by photo) = NB + NC + NL. Photosynthetic N use efficiency (PNUE) = Pmax (maximum net photosynthetic rate)/SLN.The leaf, stem and root N contents were determined by Kjeldagl method . AdditioP. notoginseng grown under different N regimes. The following equations were used to calculate N uptake and use efficiency (\u22121) = yield (underground dry weight)/plant N accumulation; NAE (kg\u00b7kg\u22121) = (yield with N application \u2013 yield without N application)/N rate; NUPE (kg\u00b7kg\u22121) = above-ground total N content/N rate; RNF (%) = /N rate \u00d7 100; NCR (%) = (yield with N application \u2013 yield without N application)/yield with N application \u00d7 100; NPFP (kg\u00b7kg\u22121) = yield with N application/N rate.Based on the biomass and N contents, N use efficiency (NUE), N agronomic efficiency (NAE), N uptake efficiency (NUPE), recovery of N fertilizer (RNF), N contribution rate (NCR), N partial factor productivity (NPFP) were calculated in ficiency : NUE  were purchased from Yuanye Bio-technology . Kit column  was used for the determination, and the mobile phase was acetonitrile (ACN)-water. Chromatographic conditions: elution with 0\u20135 min, 17\u201320% ACN; 5\u201320 min, 20% ACN; 20\u201345 min, 20\u201342% ACN; 45\u201350 min, 42\u2013100% ACN; set with flow rate, injection volume, monitoring wavelength and column temperature of 1.0 mL\u00b7min\u22121, 10 \u00b5L, 203 nm and room temperature, respectively. Total saponins are the sum of Rg1, Rb1, Re, Rd and R1. The HPLC chromatograms of P. notoginseng root grown in different N environments are shown in Dry root samples of 0.3 g were extracted in 100% methanol and sonicated for 30 min. The solution volume was fixed to 25 mL. Saponin contents were determined as described by n = 5 or 7). One-way analysis of variance was used to evaluate the effect of N treatment in a year by T-test using SPSS software (IBM SPSS Statistics). LSD-test was used to compare treatment means, with significant effects having P < 0.05. Plots were made using Origin 2021 and GraphPad 8.0 software. Pearson correlation coefficients were assessed using Origin 2021. Principal component analysis loading factors were assessed using Origin 2021.All data in the tables and figures were mean \u00b1 standard deviation (SD) of 5\u20137 independent biological replicates performed (P. notogisneng between MN and LN conditions (P > 0.05). The stem and leaf N content were higher in two-year-old P. notoginseng compared with three-year-old plants . For two-year-old P. notoginseng, main root length and total root length were increased by 31.24% and 11.10% in MN-grown plants compared with HN-grown P. notoginseng, respectively . The number of fibrous roots, length of fibrous root, total root length, and root volume declined with an increase in N application . Leaf biomass was increased by 145.45% and 125.00% in MN-grown plants compared with LN- and HN-grown P. notoginseng, respectively (P > 0.05). RMF, SMF and LMF were increased by 110.00%, 88.89% and 72.73% in MN-grown plants compared with HN-grown P. notoginsneng, respectively . Leaf biomass was increased by 127.62% and 54.60% in HN-grown plants compared with LN- and MN-grown P. notoginseng, respectively (P > 0.05). SMF and LMF were improved by 27.59% and 28.57% in HN-grown P. notoginseng compared with LN-grown plants . NUE was declined by 62.96% and 34.03% in two- and three-year old P. notoginseng grown under HN condition compared with MN conditions, respectively (P < 0.05). The minimum values of NAE, NUPE, NCR, and NPFP were obtained in the HN-grown P. notoginaseng (P < 0.05). RNF was increased by 29.57% in three-year-old P. notoginseng grown under HN compared with MN condition (There were considerable differences in N efficiency of ondition .P < 0.05), and SLN was higher in two-year-old plants compared with three-year-old plants (P. notoginseng in LN and MN conditions (P < 0.05).SLN increased with an increase in N application . CE (carboxylation efficiency) and Jmax were highest in two-year-old plants under MN condition. CE and Jmax were increased by 57.14% and 57.58% in MN-grown plants comparted with HN-grown P. notoginseng, respectively (Vcmax and \u0393* (carbon dioxide compensation point) variables, were not significantly different in two-year-old plants grown under LN and HN conditions , LSP (light saturating point), Vcmax, and Rd (dark respiration rate) were significantly declined in three-year-old plants under LN condition (P < 0.05). For three-year-old plants, the maximum values of Pmax, CE, \u0393*, Jmax, Vcmax, and Jmax/Vcmax were obtained in MN plants . HN induces the increase in NL, and NC was reduced by 13.79% in two-year-old plants grown under HN compared with MN  were not significantly different N regimes (P < 0.05). The minimum value of total saponins (%) were recorded in three-year-old plants grown under HN condition (P < 0.05). For three-year-old plants, the LN and HN-grown P. notoginseng showed 32.58% and 28.68% lower saponins yield of per plant than the MN ones  and SLN (r = \u22120.87). Root biomass was close positively correlated with N use efficiency (as reflected by NCR (r = 0.79), NPFP (r = 0.91) and NUE (r = 1.00)). There was little correlation between root biomass and RNF, plant height and leaf area. Stem and leaf biomass were close negatively correlated with SPAD and PNUE. N application was close negatively correlated with NAE (r = \u22120.87), NCR (r = \u22120.85), NPFP (r = \u22120.91) and NUPE (r = \u22120.75). NUPE was positively correlated with the root length (r = 0.66) and root tuber diameter (r = 0.59). In addition, Pn was negatively correlated with N application (r = \u22120.88), leaf area (r = \u22120.57), Chl contents (r = \u22120.75), SLN (r = \u22120.44) and leaf N content (r = \u22120.45). The relationship between saponins and Pn (r = 0.45), root biomass (r = 0.54) as well as NPFP (r = 0.57) were positive correlation in P. notoginseng. Saponins content was negatively correlated with Chl content (r = \u22120.67) and SLN (r = \u22120.64).Pearson correlation coefficients of 27 parameters were evaluated in  regimes . As showP. notoginseng biomass or saponins. In PC1, the weighting coefficients of biomass parameters , yield, NUE, SLN, SPAD and stem N content were larger . The cumulative contribution of PC1, PC2 and PC3 reached 84.80% . Thus, te larger . NUE, yie larger . In PC2,e larger . In PC3,e larger . LCP ande larger .Arabidopsis thaliana L. by increasing the length of total root and fibrous root (r = 0.66) and root tuber diameter  Previouoginseng , 4. HN iativa L. . Overalle.g., Dodonaea viscosa (L.) Jacq., Lolium perenne L. and Betula spp. (P. notoginseng are exposed to HN condition (r = \u22120.93) and stem (r = \u22120.72) N content . ula spp. , and shoula spp. . These rondition . It has ondition . However content . It is aMolinia caerulea (L.) Moench , and lower NAE and NCR were obtained in HN-grown P. notoginseng (Many studies have shown that N use is not positively related to N uptake . N conte) Moench . This isce N use , 4. Howeoginseng , Table 3L and Pn were recorded in LN-grown plants  and stem (r = \u22120.86) biomass  and yield (r = 0.53) were positively correlated with Pn  Fern.-Vill. grown under high N condition  Iljinsk.  is generally used as a traditional Chinese medicine  is recorded in three-year-old P. notoginseng grown HN condition  was recorded in HN-grown plants (Centella asiatica L. and Stevia rebaudiana (Bertoni) Hemsl.  Hemsl. ). Howeve) Hemsl. , 7E, 7F. quality . In shorP. notoginseng (P. notoginseng.A model was proposed to explain the interaction between high N and the accumulation of biomass and C-containing secondary metabolites in a N-sensitive medicinal species, such as oginseng . In conc10.7717/peerj.14933/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.14933/supp-2Supplemental Information 2Click here for additional data file.10.7717/peerj.14933/supp-3Supplemental Information 3Click here for additional data file."}
+{"text": "Ancestral SARS coronavirus-2 (SARS-CoV-2) and variants of concern (VOC) caused a global pandemic with a spectrum of disease severity. The mechanistic explaining variations related to airway epithelium are relatively understudied. Here, we biobanked airway organoids (AO) by preserving stem cell function. We optimized viral infection with H1N1/PR8 and comprehensively characterized epithelial responses to SARS-CoV-2 infection in phenotypically stable AO from 20 different subjects. We discovered Tetraspanin-8 (TSPAN8) as a facilitator of SARS-CoV-2 infection. TSPAN8 facilitates SARS-CoV-2 infection rates independently of ACE2-Spike interaction. In head-to-head comparisons with Ancestral SARS-CoV-2, Delta and Omicron VOC displayed lower overall infection rates of AO but triggered changes in epithelial response. All variants shared highest tropism for ciliated and goblet cells. TSPAN8-blocking antibodies diminish SARS-CoV-2 infection and may spur novel avenues for COVID-19 therapy. \u2022Airway organoids from different donors display distinct compositions of cell types\u2022Organoid biobank models the spectrum of the epithelium response to pathogens\u2022TSPAN8 is a conserved mediator of infection for SARS-CoV-2 variants Roose and colleagues generated a biobank of 20 airway organoids for modeling variations of airway epithelium response to SARS-CoV-2. They discovered Tetraspanin-8 (TSPAN8) as a facilitator of SARS-CoV-2 infection independently of ACE2-Spike interaction. Pre-treatment of airway organoids with a blocking TSPAN8 antibody decreased SARS-CoV-2 infection levels in airway organoids, suggesting TSPAN8 as a therapeutic target for COVID-19. While most SARS-CoV-2-infected individuals develop asymptomatic to mild disease, some develop a severe disease characterized by immune cell dysfunction (trans-membrane serine proteases (TMPRSS2) results in Spike protein activation and viral entry (Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) has caused a global pandemic of coronavirus disease (COVID-19) with more than half a billion cases worldwide  derived from adult stem cells of different individuals. We used this biobank to first optimize viral infection of AO with H1N1/PR8 influenza, and next performed a comprehensive analysis of SARS-CoV-2 infection with repeat infections. Spectral flow analysis of infected AO was used to assess cellular and functional responses of the epithelial cell compartment. Single-cell RNA sequencing (scRNA-seq) and Spectral flow enabled the discovery of Tetraspanin-8 (TSPAN8) as a conserved mediator of SARS-CoV-2 Ancestral (WA-1)-, Delta-, and Omicron-variant infection. Reductionist HEK293T cell-pseudo-virus approaches showed that TSPAN8 facilitates viral entry independently of the Spike-ACE2 interaction. We show that TSPAN8 is not an alternative entry receptor. Blocking TSPAN8 in airway epithelial organoids prior to infection is associated with a decrease in the viral load of AOs. Based on our TSPAN8 work in the context of cancer , we proplike cells , goblet-like cells  , three populations of cells expressing basal cell markers CD49f+CD271+, CD49fnegCD271+, CD49f+CD271+, and a\u00a0population of CD49fnegCD271negacTUBAnegMUC5ACnegTSPAN8neg cells. AOs derived from different donors displayed distinct cell-type compositions even though cultured in identical growth factors and environmental conditions  , pre-gobnditions G. Furthenditions H and\u00a0S1Anditions B. We gennditions C. Spectrnditions D and extnditions E revealenificant I\u20131K. So,high/CD271\u2212neg ciliated cells and acTUBAneg/MUC5AC+ goblet cells  A. ConfocSpectral flow analysis on five independent SARS-CoV-2 infections of 2522UL show that infections are consistent C and 3D To uncover the rules of infection with SARS-CoV-2, we selected a panel of 12 AOs A that ca+, or CD271+, or CD49fnegCD271neg cells  for Mock and SARS-CoV-2-infected organoids on 21 variables  A. PCA lA showed + cells\u00a0H upon SA+ cells\u00a0I\u2013S3L. Th+ cells\u00a0J, confir+ cells\u00a0. Surprison rates K and S3Gon rates , and theon rates C motivatWe infected four organoids  with SARS-CoV-2 WA-1 and performed scRNA-seq. Unsupervised clustering analysis based on most variable gene expression across all cells , regardlTSPAN8 mRNA reads were present in 64% of single cells positive for SARS-CoV-2 reads (size of the circle in +TSPAN8\u2212) from pre-goblet cells (MUC5AC\u2212TSPAN8+)  , allowinTSPAN8+) C but decTSPAN8+) A. As we TSPAN8+) J, most STSPAN8+) D, whereaTSPAN8+) E. FurtheTSPAN8+) F and S5BTSPAN8+) K, the prnfection G and S5Cnfection H and 5I.TSPAN8 expression decreases in airway brushes of acute illness in patients caused by non-SARS-CoV-2 respiratory viruses, while airway brushes from COVID-19 patients revealed preservation of TSPAN8 levels  numbers and influences their composition . EV fromjeroen.roose@ucsf.edu.A list of materials used and detailed methods are found on the Supplementary Materials section. Materials are available upon request.GSE211562.Resources: GEO: L.H. and J.P.R.: conceived the study. L.H., S.L., and K.K.: Spectral flow. L.H., K.K., S.L., O.M.G.: organoid biobank. L.H., M.M., and L.R.: BSL3 work. C.A., A.A.R., and A.J.C.: scRNA-seq. J.C.L.: statistical analyses. C.A., L.C.L.: scRNA-seq analyses. L.R.B. and D.J.E.: advice on airway populations. N.K.S. and M.K.: H1N1 virus. L.H., S.L., O.M.G., and K.B.: microscopy. L.H., S.L., K.B.: HEK239T cell line creation. J.Z.L., V.D., S.M., M.M.: patient samples. G.K., D.M.J., M.M., A.J.C.: funding. M.O.: SARS-CoV-2 virus, and SARS-CoV-2 pseudo-viruses. S.L., M.P.: pseudo-virus infections. J.R.K.: surgery airway samples, clinical data discussion. L.H., S.L., J.P.R.: manuscript writing. J.P.R.: funding. G.K.F., D.M.J., M.M., M.O., M.M., A.J.C., D.E., A.N.S., and J.R.K.: edits on the manuscript."}
+{"text": "Misgurnus anguillicaudatus), the most widely distributed species of the family Cobitidae, displays a mud-dwelling behavior and intestinal air-breathing, inhabiting the muddy bottom of extensive freshwater habitats. However, lack of high-quality reference genome seriously limits the interpretation of the genetic basis of specialized adaptations of the loach to the adverse environments including but not limited to the extreme water temperature, hypoxic and noxious mud environment.The loach (fos), a regulator of bone development, is positively selected in loach. Knockout of fos (ID: Mis0086400.1) led to severe osteopetrosis and movement difficulties, combined with the comparison results of bone mineral density, supporting the hypothesis that fos is associated with loach mud-dwelling behavior. Based on genomic and transcriptomic analysis, we identified two key elements involved in the intestinal air-breathing of loach: a novel gene (ID: mis0158000.1) and heat shock protein beta-1 (hspb1). The flavin-containing monooxygenase 5 (fmo5) genes, central to xenobiotic metabolism, undergone expansion in loach and were identified as differentially expressed genes in a drug stress trial. A fmo5\u2212/\u2212 (ID: Mis0185930.1) loach displayed liver and intestine injury, indicating the importance of this gene to the adaptation of the loach to the noxious mud.This study generated a 1.10-Gb high-quality, chromosome-anchored genome assembly, with a contig N50 of 3.83 Mb. Multiple comparative genomic analyses found that proto-oncogene c-Fos (Our work provides valuable insights into the genetic basis of biological adaptation to adverse environments.The online version contains supplementary material available at 10.1186/s12915-023-01517-1. Misgurnus anguillicaudatus), the most widely distributed species of the family Cobitidae, inhabits in the bottom of lakes, ponds, and other freshwater areas with humus-rich mud ; ugt, UDP-glucuronosyltransferase. Table S5. Primers used in this study. qPCR, quantitative PCR; # indicated T7 promoter sequences. Fos, proto-oncogene c-Fos; fmo5, dimethylaniline monooxygenase [N-oxide-forming] 5 (Mis0185930.1); hbb, hemoglobin subunit beta; hba, hemoglobin subunit alpha; atf6, cyclic AMP-dependent transcription factor; eif2ak3, eukaryotic translation initiation factor 2-alpha kinase 3; chop, DNA damage-inducible transcript 3 protein; muc2, mucin-2; Mis0185950.1, Mis0185940.1, Mis0185920.1, Mis0185970.1, Mis0186000.1, Mis0186010.1 (fmo5), dimethylaniline monooxygenase [N-oxide-forming] 5; Mis0115330.1, Mis0115330.1, Mis0135560.1 (ugt2a2), UDP-glucuronosyltransferase 2A2; Mis0072610.1 (ugt2a1), UDP-glucuronosyltransferase 2A1; npr2, atrial natriuretic peptide receptor 2; f9, coagulation factor IX; hspb1, heat shock protein beta-1; hyou1, hypoxia up-regulated protein 1; ptafr, platelet-activating factor receptor; scx, basic helix-loop-helix transcription factor scleraxis; galnt8, polypeptide N-acetylgalactosaminyltransferase 8; krt13, keratin, type I cytoskeletal 13 Cytokeratin-13; gp2, pancreatic secretory granule membrane major glycoprotein; smco3, single-pass membrane and coiled-coil domain-containing protein 3; ccl5, C-C motif chemokine 5; ifi44, interferon-induced protein 44; atf4, cyclic AMP-dependent transcription factor 4; ugt1a1, UDP-glucuronosyltransferase 1-1. Table S6. A summary of sequencing data used in genome assembly and annotation of Misgurnus anguillicaudatus.Additional file 2: Fig S1. Genome-wide Hi-C interaction map. Fig S2. Divergence distributions of four TE sequences predicted by the de novo method (a) and comparison of the distribution of several features in the final gene set for seven fish species (b). TE, transport elements; DNA, DNA transposons; LTR, long terminal repeats; LINE, long interspersed nuclear elements; SINE, short interspersed nuclear element. The seven fish species: Misgurnus anguillicaudatus, Carassius auratus, Cyprinus carpio, Danio rerio, Sinocyclocheilus graham, S. rhinocerous, and Triplophysa tibetana. Fig S3. The construction of maximum likelihood tree of fos gene and generation of the fos (ID: Mis0086400.1) knockout loach (fos\u2212/\u2212) by CRISPR/Cas9 technology. (a) Maximum likelihood tree ) of fos gene in fish species. Different branch colors represent different species. Red words indicate the knockout of fos gene in loach genome. (b) Schematic position of the CRISPR/Cas9 target site for fos gene knockout. (c) The transcription level of fos in livers of wild-type loach (WT) and fos\u2212/\u2212 loach. (d) A statistics of survival rate of fertilized eggs of WT loach and heterozygous F1 generation (F1 generation self-crossed). *** extremely significant difference (p < 0.001). hpf, hours post fertilization; fos, Proto-oncogene c-Fos. The gene in red color is a positively selected gene in the loach genome. Fig S4. Identification and expression analysis of air-breathing- and digestion/absorption- related genes in loach Misgurnus anguillicaudatus. (a) Maximum likelihood tree ) of hb gene family. Different color backgrounds represent different genes. Different branch colors and inner circle colors represent different species. hbb, hemoglobin subunit beta; hba, hemoglobin subunit alpha. (b) Expression levels of hbb and hba in posterior intestines of the loach under air exposure. (c) qPCR validation of RNA-seq data of loach posterior intestines under air exposure. (d) Maximum likelihood tree ) of air-breathing-related genes. Different branch colors represent different species. (e) Maximum likelihood tree of digestion/absorption-related genes ). Atp1a, sodium/potassium-transporting ATPase subunit; ryr, ryanodine receptor. Different color backgrounds represent different genes. Different branch colors represent different species. Npr2, atrial natriuretic peptide receptor 2; f9, coagulation factor IX; hspb1, heat shock protein beta-1; hyou1, hypoxia up-regulated protein 1; ptafr, platelet-activating factor receptor; scx, basic helix-loop-helix transcription factor scleraxis; galnt8, polypeptide N-acetylgalactosaminyltransferase 8; krt13, keratin, type I cytoskeletal 13 Cytokeratin-13; gp2, pancreatic secretory granule membrane major glycoprotein; smco3, single-pass membrane and coiled-coil domain-containing protein 3; ccl5, C-C motif chemokine 5; ifi44, interferon-induced protein 44; cldn5, claudin-5 Transmembrane protein deleted in VCFS; hspb1, heat shock protein beta-1; vegfr1 (flt1), vascular endothelial growth factor receptor 1; ctgf, connective tissue growth factor CCN family member 2. Fig S5. Identification of FMO and UGT gene families. (a) Maximum likelihood tree ) of FMO gene family. Different colors represent different FMO gene families and different branch colors represent different species. (b) Maximum likelihood tree ) of UGT gene family. Different colors represent different UGT gene families and different inner circle colors represent different species. Fig S6. qPCR validation of RNA-seq data from the drug stress trial. Fig S7. The expression levels of ugt genes in liver tissues of the loach under five drug stress. Mis115330.1 and Mis0135560.1 (ugt2a2), UDP-glucuronosyltransferase 2A2; Mis0072610.1 (ugt2a1), UDP-glucuronosyltransferase 2A1. Fig S8. Tissue expression analysis and the knockout of fmo5 (ID: Mis0185930.1) gene (fmo5\u2212/\u2212) in loach Misgurnus anguillicaudatus genome. (a) Tissue expression analysis of fmo5 (ID: Mis0185930.1) of the loach. (b) Schematic position of the CRISPR/Cas9 target site for fmo5 gene knockout. (c) The mRNA and protein expression levels of fmo5 in livers of wild-type (WT) and fmo5\u2212/\u2212 loach. Different letters above error bars indicate significant difference among different tissues (p < 0.05); *** extremely significant difference (p < 0.001). Fig S9. Survival rates of wild-type (WT) and fmo5deletion (fmo5\u2212/\u2212) loach under five drug stress."}
+{"text": "There is an error in reference 5. The correct reference is: Masashi K, Norihiro S, Hidetsugu N.Regular change in spontaneous preparative behaviour on intra-abdominal pressure and breathing during dynamic lifting. Eur J Pppl Physiol. 2014; 114(11):2233\u20139"}
+{"text": "Atrial fibrillation (AF) was more frequent in non-survivors (p < 0.0001), alongside a longer QTc interval (p = 0.0002), a lower Tp-e/QTc ratio (p = 0.0003), and right ventricular strain (p = 0.013). Remdesivir administration was associated with bradycardia development (p = 0.0005) but no increase in mortality rates. In a Cox regression model, AF (aHR 3.02 (95% CI 1.03\u20138.81); p = 0.042), QTc interval above 451 ms (aHR 3.24 (95% CI 1.09\u20139.62); p = 0.033), and right ventricular strain (aHR 2.94 (95% CI 1.01\u20138.55); p = 0.047) were associated with higher 28-day mortality risk. Conclusions: QTc interval > 451 ms, right ventricular strain, and AF are associated with higher mortality risk in SARS-CoV-2 hospitalized patients. ECG recording and its appropriate analysis offers a simple, quick, non-expensive, and validated approach in the emergency setting to guide COVID-19 patients\u2019 stratification.Background: Electrocardiogram (ECG) offers a valuable resource easily available in the emergency setting. Objective: Aim of the study was to describe ECG alterations on emergency department (ED) presentation or that developed during hospitalization in SARS-CoV-2-infected patients and their association with 28-day mortality. Methods: A retrospective, single-center study including hospitalized patients with SARS-CoV-2 was conducted. ECG was recorded on ED admission to determine: heart rhythm, rate, and cycle; atrio-ventricular and intra-ventricular conduction; right ventricular strain; and ventricular repolarization. A specialized cardiologist blinded for the outcomes performed all 12-lead ECG analyses and their interpretation. Results: 190 patients were included, with a total of 24 deaths (12.6%). Age ( Since the beginning of the SARS-CoV-2 global emergency in December 2019, more than 300 million cases and 5 million deaths have been recorded worldwide, and these numbers keep rising [Multimorbidity, including past cardiovascular or pulmonary disease history and older age above all, have been previously associated with severity of infection and mortality . On the 1Q3T3 sign or inferior leads T wave inversion, which reflects the associated lung involvement and is already linked to higher disease burden [Electrocardiographic abnormalities have been observed in 99% of elderly and critically ill patients infected with SARS-CoV-2 . These ie burden .Despite the bulky amount of data, a comprehensive analysis of ECG parameters on emergency presentation in COVID-19 patients is missing, as either attention is focused on specific ECG abnormalities, or solid evidence on alterations is still lacking. ECG recording represents the first step of the cardiological assessment and can prove essential for patients\u2019 risk stratification in the ongoing emergency frame, being a handy, inexpensive, and widely available tool.Therefore, this study aims to describe the prevalence and type of electrocardiographic alterations at emergency department (E.D.) arrival in subsequently hospitalized SARS-CoV-2-infected patients and to investigate the possible association between ECG parameters and 28-day mortality after adjusting for variables, including age, sex, comorbidities, and laboratory findings that could influence the endpoint.A monocentric, retrospective study was conducted at Azienda Ospedaliero Universitaria Policlinico Umberto I, a tertiary care hospital with 1235 beds, the seat of \u201cSapienza\u201d University of Rome Medical School, between March 2020 and January 2021.Patients over 18 years of age with SARS-CoV-2 infection confirmed by rapid antigen or molecular  nasopharyngeal swab test subsequently admitted from E.D. to Infectious Diseases COVID-19 hospital wards in the abovementioned period were initially included in the analysis, for a total of 531 patients. Underage or discharged patients, patients with no laboratory proven infection, or those admitted to wards other than infectious diseases ward (I.D.)  as well as patients receiving drugs potentially elongating the QT as well as >48 h of azithromycin or hydroxychloroquine were excluded. In addition, patients were excluded whether data were incomplete for study purpose or E.D. recorded standard twelve-lead ECGs were missing. Accordingly, the proposed criteria led to 341 excluded and 190 included patients .2/FiO2 ratio; potential ICU stay during hospitalization; in-hospital and 28-day mortality, length of hospital stay, and therapy administered against SARS-CoV-2. Patients\u2019 data were anonymously recorded from medical reports into an electronic spreadsheet for the following statistical analysis. These consisted of demographics; comorbidities included in the Charlson Comorbidity Index (CCI); plus systemic hypertension, AF, and asthma; vital signs recorded in E.D., including relative bradycardia (defined as copresence of body temperature \u2265 38.3 \u00b0C and heart rate (HR) < 90 bpm) [1Q3T3 or T3 alone pattern), and ventricular repolarization .All 12-lead ECG analyses and their interpretation were performed by a specialized cardiologist (M.C.G.) who was blinded for the outcomes. The following parameters were retrieved: heart rhythm, heart rate (expressed as bpm), and heart cycle , expressed as ms), atrio-ventricular and intra-ventricular conduction parameters , left posterior hemi-block (LPH), right or left bundle branch block (RBBB or LBBB)), morphological evaluation with particular emphasis on right ventricular strain . The need for informed consent was waived since all data were retrospectively extracted.Cardiovascular events during hospitalization were defined as the onset of new ischemic/embolic events, such as pulmonary thrombo-embolism by lung CT scan, acute cerebral ischemia, acute limb ischemia, or the development of myocardial infarction, Takotsubo syndrome, myocarditis.Heart rhythm disorders included the new onset of atrial fibrillation, supraventricular tachycardia, bradycardia, pairs of ventricular premature beats, and ventricular tachycardia.Relative bradycardia was defined as heart rate < 90 bpm and concomitant fever (tympanic temperature \u2265 38.3 \u00b0C) [Daytime bradycardia was defined as mean heart rate < 60 bpm recorded three times a day.1Q3T3 pattern  or negative T wave alone in leads V1\u2013V3 or II, III, or aVF with or without ST depression [Right ventricular strain was defined as the presence of Spression .The QT interval was defined as the interval from the onset of the QRS complex to the end of the T wave and expressed as ms. Corrected QT interval (QTc) was calculated according to Bazett formula: QTc = QT\u221a . In the QT dispersion, which reflects regional differences in myocardial refractoriness and predict cardiac dysrhythmias , was expFurthermore, regional differences in myocardial refractoriness were calculated also by means of Tp-e interval  . Measure\u00ae software, v. 15 (StataCorp); charts were generated using Microsoft Office\u00ae and Graphpad Prism\u00ae. Continuous data are expressed as median and interquartile range (IQR) values and categorical variables as numbers and percentage values. Categorical variables were compared using \u03c72-test or Fisher\u2019s exact test and continuous variables using Student\u2019s t-test or Mann\u2013Whitney U test as appropriate.Statistical analysis was performed using STATA2/FiO2, D-dimer, CRP, and lymphocytes count, expression of respiratory failure and inflammation during COVID-19, respectively, were performed. According to the reference values available at the laboratory of our hospital, abnormal levels of troponin corresponded to levels > 0.014 \u03bcg/L.In the subgroup of patients with troponin levels available, Spearman correlation analyses between troponin levels and PaOlogrank test. For all the statistical analyses, p < 0.05 was considered significant.Multivariate Cox regression models were used to determine the hazard ratios (HR) for mortality within 28 days from admission of the included variables accounting for covariables. Statistically and clinically relevant variables on univariate analysis were evaluated for the determination of the final multivariate model. To find the optimal cut-off of the QTc value associated with the highest sensitivity and specificity in the prediction of 28-day outcome, the Youden\u2019s Index was used. The resulting value was further inserted in the final model. Twenty-eight-day survival curves were plotted using the Kaplan\u2013Meier method and compared using the p < 0.0001), while no significant difference was observed regarding sex. No deaths were recorded within 48 h from E.D. arrival; therefore, this variable was not included in the study. Patients hospitalized during the first pandemic wave in Italy (February\u2013June 2020) were evenly distributed among the two compared subgroups (89/166 (53.6%) vs. 10/24 (41.7%); p = 0.273). This reduces possible bias in different diagnostic and therapeutic management of SARS-CoV-2 infection over time.Overall, 190 patients were included in the study, with 83/190 (44%) females . Median p < 0.0001). Besides hypertension, myocardial infarction (MI) was the most common cardiovascular comorbidity , followed by AF  and chronic heart failure (CHF) , which was more prevalent in the deceased subgroup, where AF was recorded in 7/24 patients , MI in 6 , and CHF in 3   patients). Calculated CCI was significantly higher in non-survivors (median (IQR), 8 (8\u201310)) when compared to survivors (3 (1\u20136); = 0.160) .p = 0.002), whereas dyspnea was the most prevalent among deceased although not statistically significant (16/24 (66.7%) vs. 85/166 (51.2%) patients; p = 0.164). Moreover, in the latter subgroup, median time between symptoms onset and E.D. presentation was significantly shorter than in survivors (median (IQR), 1.8 (0\u20135.5) vs. 6 (2\u20139) days; p = 0.039). Relative bradycardia was registered in 22/190 patients (11.6%) and, though more frequent among survivors (21/166 patients (12.6%)), resulted as not statistically relevant (p = 0.225).Fever was the most prevalent symptom in both the overall population (149/190 patients (78.4%)) and among survivors (136/166 (82%) vs. 13/24 (54.2%) patients; 2/FiO2 ratio on triage arterial blood gas was lower in non-survivors (median (IQR), 302 (243\u2013367) vs. 357 (314\u2013424); p = 0.0007), together with high white blood cells count (7180 (5070\u20138820) vs. 5755 (4502\u20137790) cells/\u00b5L; p = 0.009) and neutrophils count (5465 (3647\u20137662) vs. 4150 (3110\u20135995) cells/\u00b5L; p = 0.0038), anemia (median Hb (IQR) 11.3 (10.2\u201314.5) vs. 13.8 (12.7\u201314.9) g/dL; p = 0.0004), high D-dimer (1995 (1012\u20133198) vs. 777 (429\u20131469) U/L; p = 0.0012) and LDH (330 (268\u2013447) vs. 275 (213\u2013349) U/L; p = 0.0001), and low serum albumin (32 (30\u201335) vs. 38 (35\u201341) g/L; p < 0.0001). Regarding serum troponin T, the test was available on admission in 125/190 patients only and was higher in non-survivors (0.031 (0.021\u20130.04) vs. 0.012 (0.007\u20130.0245) \u03bcg/L; p = 0.084). Turning to laboratory tests, PaOp < 0.0001) and paroxysmal supraventricular complexes (PSVC) in 20.8% (5/24 (20.8%) vs. 9/166 (5.4%) patients; p = 0.0193). Mean RR interval was reduced, albeit not significantly, in non-survivors (median (IQR), 637.5 (570\u2013762) vs. 767 (664\u2013875) ms; p = 0.006) as well as its standard deviation (18 (11\u201330) vs. 20 (13.4\u201335) ms; p = 0.414). Regarding ventricular conduction parameters, the only significant difference between the subgroups was observed for the higher prevalence of left anterior hemiblock (9/24 (37.5%) vs. 28 (19.8%) patients; p = 0.0258) in non-survivors, who also recorded a higher occurrence of right ventricular strain as S1Q3T3 pattern (7/24 (29.1%) vs. 18/166 (10.8%) patients; p = 0.013) or as single-components inverted T wave in DIII (T3) (15/24 (62.5%) vs. 45/166 (27.1%) patients; p < 0.0001) and prominent S wave in DI (S1) (9/24 (37.5%) vs. 30/166 (18%) patients; p = 0.034). Median QTc interval duration was longer in non-survivors (436.8 (435\u2013487) vs. 428 (402\u2013447) ms; p = 0.0002), resulting in lower Tp-e/QTc ratio (0.2 (0.158\u20130.198) vs. 0.22 (0.211\u20130.233); p = 0.0003). Prolonged QTc was observed in 55 (28.9%) subjects, higher in non-survivors than survivors (39 (23.49%) vs. 16 (66.6%), p < 0.0001). Following Youden\u2019s index, the optimal cut-off of QTc differentiating 28-day survivors from non-survivors was 451 ms ).p < 0.0001) as new onset in-hospital AF although not relevant (3/24 (12.5%) vs. 8/166 (4.8%) patients; p = 0.1477). On the other hand, daytime bradycardia was more frequent in survivors (2/24 (8.3%) vs. 28/166 (16.9%); p = 0.284).p = 0.0045) and with systemic corticosteroids (17/24 (70.8%) vs. 72/166 (43.4%), p = 0.045), while administration of macrolides, HCQ, and prophylactic dose LMWH (4.000 UI/24 h s.q.) was uniform within the two subgroups, as shown in In the non-survivor subgroup, a higher rate of patients was treated with therapeutic dose LMWH (100 U/kg/12 h s.q.) (10/24 (41.7%) vs. 33/166 (19.9%) patients; p = 0.021 and 0.036, respectively) but not with right ventricular strain (p = 0.94) and tended to be associated with abnormalities at admission ECG (p = 0.08).Troponin levels at hospital admission was available in 125/190 (65.8%) subjects. Abnormal levels of troponin (>0.014 \u03bcg/L) were associated with AF and QTc , whereas a negative correlation was observed with PaO2/FiO2  and lymphocyte count   .logrank test analysis with p-values. Among electrocardiographic findings, AF on E.D. admission recording (p < 0.0001) or developed during hospitalization (p = 0.0409) or considered together as cumulative AF (p < 0.0001) were associated with lower 28-day survival rates for right heart strain (p = 0.0093) and QTc value > 451 ms (p < 0.0001). Relative bradycardia was not significantly different between survivors and deceased (p = 0.3148). In addition, age > 65 y (p = 0.0002), CRP over 4 mg/dL (p = 0.0023), D-dimer over 850 U/L (p = 0.0035), and serum albumin below 35 g/L (p < 0.0001) on E.D. admission laboratory tests were associated with lower 28-day survival rates.p < 0.05) and clinically relevant variables on univariate analysis were evaluated for the determination of hazard ratios (HRs) for 28-day mortality. Following Youden\u2019s index results, QTc value > 451 ms was considered in the final model. AF detection on E.D. arrival ECG or its in-hospital development (HR 3.02 (95% CI 1.03\u20138.81); p = 0.042), QTc > 451 ms (HR 3.24 995% CI 1.09\u20139.62); p = 0.033), and right ventricular strain (HR 2.94 (95% CI 1.01\u20138.55); p = 0.047) were associated with higher 28-day mortality risk after adjustment for age, sex, cardiac and pulmonary comorbidities , and laboratory tests  that proved clinically pertinent and could potentially influence the outcome  or inverted T wave on ECG recordings. This has already been described as a negative prognostic factor in patients with non-COVID19-related PTE [3 sign alone, indeed, was included in the right ventricular strain analysis as a strain mark and a recognized negative prognostic factor [2 \u2264 95% and RR > 20 bpm. Concerning laboratory tests on E.D. admission, low PaO2/Fio2 ratio, anemia, low serum albumin, leukocytosis with neutrophilia, elevated LDH, D-dimer, and CRP were prevalent in non-survivors, as previously described [Furthermore, hypoxic stress and lung damage, its related pulmonary hypertension and right ventricular heart strain, in addition to the high rates of pulmonary thromboembolism (PTE)  registerated PTE  and in oc factor . Our finc factor , who linescribed . Though escribed . As for escribed .A total of 15.8% of patients developed sinus bradycardia during hospitalization. This was not linked to higher mortality rates, as previously described , even thAutonomic dysfunction is shown throughout the lower heart rate variability registered in non-survivors as RR interval, which reflects the sympathovagal balance interacting with IL-6 and the ongoing pro-inflammatory boost ,33. This2/FiO2 (expression of respiratory failure), D-dimer, CRP, and lymphocytes (expression of inflammation during infection). Furthermore, abnormal troponin levels were associated with AF and QTc and, overall, tended to be associated with abnormal ECG on hospital admission. Taken together, these findings suggest that the possible myocardial damage, expressed by the troponin values, may be directly related to the infection itself rather than to a previous cardiac problem. Nevertheless, although troponin represents a marker of myocardial injury, it may be unreliable when considered alone, and therefore, it should be analyzed only when combined with additional clinical and laboratory parameters.Statistically significant correlations were found between troponin values and specific parameters of COVID-19, such as PaOThis study shows several limitations. Primarily, its retrospective design does not allow a confident generalizability. Despite controlling for demographic and clinical data, there could be other confounding variables not included in multivariate analysis, and the omission of radiologic parameters does not grant a strict clinical assessment of right heart strain and conceivable underlying pulmonary thromboembolism. Moreover, the temporal nexus between ECG findings and their development is missing, as we only analyzed the E.D. ECG, and certain abnormalities could have developed prior to SARS-CoV-2 infection; nevertheless, having these ECG abnormalities was associated with a worse prognosis. The potential association between ECG findings and serum troponin T levels could not be assessed, because of the few available TnT, as a former non-routine test on E.D. admission; nevertheless, only a statistical trend over significance was observed, and therefore, this variable was further excluded from the final model.Lastly, in consideration of the test duration and emergency situation experienced by our country during the study period, we did not have the possibility to routinely perform cardiac MRI to identify myocardial alterations in hospitalized patients and to comprehensively collect the frequency of post-COVID-19 cardiovascular sequelae. Nevertheless, the aim of the present study was to investigate the electrocardiographic features on hospital admission and in the emergency setting and not the cardiovascular sequelae, which, in our opinion, deserve per se additional investigations.To conclude, our study was conducted in an emergency, namely the first and second pandemic waves in Italy, prior to the development and approval of the anti-SARS-CoV-2 management strategies currently used. This supports the goal of investigate the untainted interaction of SARS-CoV-2 with heart rate and rhythm and, therefore, ECG, avoiding additional confounders such as new antiviral drugs and prophylactic treatments or the role of virus variants.This study demonstrates the association between older age, AF, QTc, and right ventricular strain recorded on E.D. admission ECG and higher mortality risk after adjusting for cardiopulmonary comorbidities and disease severity markers. These results endorse the role of the ongoing cardiovascular alterations in SARS-CoV-2 infection, likely related to the direct and indirect action of virus and cytokine storm on cardiomyocytes and unbiased by therapeutic strategies. ECG recording and its appropriate analysis offers a simple, quick, non-expensive, and validated approach in the emergency setting to guide COVID-19 patients\u2019 stratification in the ongoing pandemic frame, assisted by clinical and laboratory assessment."}
+{"text": "The somehow prophetic 2007 publication reviews \u201cnatural disasters, climate change and mental health considerations for rural Australia\u201d (2) and pinpoints central aspects of today\u2019s debate, namely anxiety and depression, vulnerability and resilience. In addition to problems of rural areas (2), the impact of urbanicity (3) will be discussed as well as the role of air pollution on psychiatric disorders (4). (1) UN Environment Programme. https://www.unenvironment.org/explore-topics/climate-change/about-climate-change Dec 22nd, 2020. (2) Morissey SA, Reser JP. Aust J Rural Health. 2007 Apr;15(2):120-5. doi: 10.1111/j.1440-1584.2007.00865.x. (3) Krabbendam L et al. Psychol Med. 2020 Mar 11:1-12. doi: 10.1017/S0033291720000355. (4) Kim SY et al. Sci Total Environ. 2020 Dec 8;757:143960. doi: 10.1016/j.scitotenv.2020.143960.According to the UN Environment Programme \u201cclimate change is one of the most pervasive and threatening issues of our time\u201d. \u201cIn many places, temperature changes and sea-level rise are already putting ecosystems under stress and affecting human well-being\u201d (1). The presentation wants to give an overview on how climate change can affect mental health. A search was performed on PubMed for the combination of \u201cclimate change\u201d and \u201cmental health\u201d. 281 publications were identified, the first being from 2007 (the only one in that year). In 2020, until Dec 22No significant relationships."}
+{"text": "During the COVID-19 pandemic, cancer patients are regarded as a highly vulnerable population. Given the unavoidable bias and unmeasured confounders in observational studies, the causal effects of cancers on COVID-19 outcomes are largely unknown. In the study, we tried to evaluate the causal effects of cancers on COVID-19 outcomes using the Mendelian randomization (MR) approach. No strong evidence was observed to support a causal role of cancer in COVID-19 development. Previous observational correlations between cancers and COVID-19 outcomes were likely confounded. Large and well-conducted epidemiological studies are required to determine whether cancers causally contribute to increased risk of COVID-19.p-values for the casual associations were all statistically insignificant: overall cancer , lung cancer , breast cancer , endometrial cancer , prostate cancer , thyroid cancer , ovarian cancer , melanoma , small bowel cancer , colorectal cancer , oropharyngeal cancer , lymphoma  and cervical cancer . Sensitivity analyses and multivariable MR analyses yielded similar results. In conclusion, cancers might have no causal effect on increasing COVID-19 risk. Further large-scale population studies are needed to validate our findings.Observational studies have shown increased COVID-19 risk among cancer patients, but the causality has not been proven yet. Mendelian randomization analysis can use the genetic variants, independently of confounders, to obtain causal estimates which are considerably less confounded. We aimed to investigate the causal associations of cancers with COVID-19 outcomes using the MR analysis. The inverse-variance weighted (IVW) method was employed as the primary analysis. Sensitivity analyses and multivariable MR analyses were conducted. Notably, IVW analysis of univariable MR revealed that overall cancer and twelve site-specific cancers had no causal association with COVID-19 severity, hospitalization or susceptibility. The corresponding  Coronavirus disease 2019 (COVID-19) is a global pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,4. ThereCancer patients are a vulnerable population during the COVID-19 pandemic ,6. CanceMendelian randomization, an epidemiological method, has been widely applied to assess the potential causal association between exposure and outcome ,15. Acco2, age \u00d7 gender, principal components and study-specific covariates by the original GWAS researchers. The COVID-19 outcomes included 1,683,768 participants  for susceptibility, 1,887,658 participants  for hospitalization, and 1,388,342 participants  for severity, respectively. The uninfected individuals served as the controls. All cases were confirmed by laboratory, self-reported, or physician diagnosis. The severe cases were defined as patients who died or required respiratory support with COVID-19 infection [The summary statistics in the genome-wide association studies (GWASs) for COVID-19 were sourced from the COVID-19 Host Genetics Initiative V5 , which eThe summary statistics of the GWASs for cancers were obtained from the UK biobank , InternaCovariates for multivariable MR analyses were included: BMI  , educatiAt the beginning of our study design, 24 site-specific cancers were considered. Overall cancer and 12 site-specific cancers were included, but another 12 types of cancer were not included due to insufficient SNPs . In p < 5 \u00d7\u200910\u22128). To ensure independence, SNPs were restricted by low linkage disequilibrium  using clumping [Appropriate SNPs used as IVs must be robustly associated with cancers ; otherwise, a random-effect model was applied. In addition, we used the \u201cleave-one-out\u201d validation to determine whether a single SNP had a significant independent influence on the MR estimation.In the univariable MR analysis, the IVW analysis was chosen as the primary approach to estimate the causal effects of cancers on COVID-19 outcomes ,38. We ap-value using the Bonferroni correction  was used. 0.0033 < p < 0.05 was regarded as suggestive evidence for a potential association. The \u03b2  and its SE (standard error) were calculated to reflect effect sizes. All statistical analyses were conducted in R v4.0.1  with the packages \u201cTwoSampleMR\u201d and \u201cMRPRESSO\u201d [We applied the random-effect IVW method to assess the causal effects of cancers on COVID-19 outcomes for the multivariable MR analyses, after controlling BMI, educational attainment, intelligence, smoking and alcohol consumption. Given the number of cancers and COVID-19 outcomes considered, a two-sided RPRESSO\u201d ,43.p = 0.07), ovarian cancer (p < 0.001), melanoma (p = 0.01) and cervical cancer (p = 0.08) (p = 0.34), lung cancer (p = 0.60), squamous cell lung cancer (p = 0.66), breast cancer (p = 0.43), ER+ breast cancer (p = 0.79), ER\u2212 breast cancer (p = 0.66), endometrial cancer (p = 0.79), prostate cancer (p = 0.54), thyroid cancer (p = 0.70), ovarian cancer (p = 0.62), melanoma (p = 0.79), small bowel cancer (p = 0.09), colorectal cancer (p = 0.85), oropharyngeal cancer (p = 0.31), lymphoma (p = 0.51) or cervical cancer (p = 0.25) on the COVID-19 severity  were included for COVID-19 severity. Severe COVID-19 cases were defined as patients who died or required respiratory support with COVID-19 infection. The effects of each SNP in cancers on COVID-19 severity can be found in  = 0.08) . Hence, severity .p = 0.03) and ovarian cancer (p = 0.01) . Although horizontal pleiotropy was observed in melanoma (p = 0.02), it showed no significant outlier. We conducted the \u201cleave-one-out\u201d analysis and found no potential SNP significantly biasing the results  . After r results . Taken t results . Results results .p = 0.06), ovarian cancer (p < 0.001) and cervical cancer (p = 0.04) (p = 0.42), lung cancer (p = 0.37), squamous cell lung cancer (p = 0.66), breast cancer (p = 0.74), ER+ breast cancer (p = 0.51), ER\u2212 breast cancer (p = 0.93), endometrial cancer (p = 0.24), prostate cancer (p = 0.17), thyroid cancer (p = 0.80), ovarian cancer (p = 0.96), melanoma (p = 0.45), small bowel cancer (p = 0.08), colorectal cancer (p = 0.79) or cervical cancer (p = 0.32) on COVID-19 hospitalization .  = 0.04) . The ranlization .p < 0.001; p = 0.78). The \u201cleave-one-out\u201d analysis showed no outliers (p = 0.04), estimates in the three analyses  and prostate cancer (p = 0.046) when adjusting for education attainment. A significant association was also found in small bowel cancer (p = 0.047) when adjusting for smoking. However, the associations of overall cancer, prostate cancer and small bowel cancer with COVID-19 hospitalization could not be replicated when intelligence , income  and alcohol consumption  were adjusted , prostate cancer (p = 0.06), thyroid cancer (p = 0.06), ovarian cancer (p < 0.001), melanoma (p = 0.05) and cervical cancer (p < 0.001) (p = 0.69), lung cancer (p = 0.96), squamous cell lung cancer (p = 0.08), breast cancer (p = 0.43), ER+ breast cancer (p = 0.30), ER\u2212 breast cancer (p = 0.18), endometrial cancer (p = 0.83), prostate cancer (p = 0.58), thyroid cancer (p = 0.28), ovarian cancer (p = 0.93), melanoma (p = 0.82), small bowel cancer (p = 0.19), colorectal cancer (p = 0.30), oropharyngeal cancer (p = 0.80), lymphoma (p = 0.37) or cervical cancer (p = 0.68) on COVID-19 susceptibility  were included for COVID-19 susceptibility. < 0.001) . Thus, wtibility .p = 0.02) and ovarian cancer (p < 0.001) . The \u201cleave-one-out\u201d plot showed one potential instrumental outlier (rs6983267) for colorectal cancer  . After rl cancer . Howeverl cancer  supporteDuring the COVID-19 pandemic, healthcare resources are extremely scarce, and there is an urgent need to allocate healthcare resources rationally . IdentifMR leverages the random allocation of genetic variants at conception, independently of confounders, to identify the causal effects that are substantially less confounded and not vulnerable to reverse causation ,15. We uAlthough many studies have generally shown positive correlations of cancers with the risk of COVID-19 ,45,46,47Risk factors may be correlated with COVID-19 outcomes, but not as a causal association. Previous MR studies have shown that many traditional risk factors have no causal association with COVID-19 outcomes, such as decreased lung function, chronic obstructive pulmonary disease, blood pressure, type 2 diabetes, chronic kidney disease, coronary artery disease, stroke and nonalcoholic fatty liver disease ,53,54,55MR design is less confounding than observational study, but limitations of this MR study need to be acknowledged. First, some types of cancer\u2014such as stomach cancer, pancreatic cancer, liver cancer and brain cancer\u2014were not included in the study because of insufficient SNPs . PotentiOverall, we used MR analysis to evaluate the causal effects of overall cancer and twelve site-specific cancers on COVID-19 severity, hospitalization and susceptibility. Results of the MR study did not suggest strong evidence to support the causal associations of any examined cancer with COVID-19 outcomes. Previous observational correlations of cancers with COVID-19 outcomes were likely confounded. More large-scale epidemiological studies are needed to validate our findings."}
+{"text": "Health impact of monoclonal gammopathy of undetermined significance (MGUS) and monoclonal B-cell lymphocytosis (MBL): findings from a UK population-based cohort. BMJ Open 2021;11:e041296. doi: 10.1136/bmjopen-2020-041296Lamb MJ, Smith A, Painter D, This article was previously published with an error in table 1. MGUS control numbers and percentages by gender were transposed and should read: males 11308 (51.6) and females 10620 (48.4)."}
+{"text": "Families with children may be at higher risk for influenza infection. Community transmission can suffer from underreporting as testing is often not performed. We studied the epidemiology of influenza in households with school-aged children using home-based sample collection.We conducted a remote household study surveilling respiratory viruses from November 2019-June 2021, in King County, Washington (WA), USA. Households with school-aged children were enrolled, mailed home specimen collection kits, and asked to self-assess for weekly acute respiratory illness (ARI) using remote survey platforms. Participants with ARI symptoms were prompted to complete serial illness surveys and self-collect/parent collect mid-turbinate nasal swabs. Samples were sent to a University of Washington study laboratory for RT-PCR influenza testing. Influenza rates were compared to WA Department of Health (DOH) reporting.A total of 1861 ARI events were reported among 992 adults and 869 children in 470 households; 75 influenza cases were detected (36 influenza A and 39 influenza B). The study participant median age was 32 years (0-84), 10 years (1-49) for influenza A, and 11 years (3-49) for influenza B cases. Overall 13% of households had an influenza case, of which 13 (22%) reported >1 case. A total of 81% of participants reported receipt of one dose of the 2019-2020 influenza vaccine, including 91% of influenza A and 90% of influenza B cases, and 84% received the 2020-2021 influenza vaccine. Like WA DOH, we observed a wave of influenza B cases followed by influenza A in 2019-2020. During influenza season 2020-2021, WA DOH reported 9 positive influenza tests and none observed in our study. Commonly, influenza case-patients reported were fever, cough, rhinorrhea, and fatigue. GI symptoms were more common in children than adults. Of the cases, 92% of influenza A and 78% of influenza B occurred in children.D0-Day of reported onset, D7-7 days after reported illness onset. No participants >49 years were positive for influenza. D0: 30 participants responded and of respondents, 13% <5 years, 47% 5-12 years, 3% 13-17 years, and 37% 18-49 years. D7: 31 participants responded and of respondents 13% <5 years, 48% 5-11 years, 3% 12-17 years, and 36% 18-49 years.D0-Day of reported onset, D7-7 days after reported illness onset. No participants >49 years were positive for influenza. D0: 28 participants responded and of respondents, 4% <5 years, 57% 5-12 years, 14% 13-17 years, and 25% 18-49 years. D7: 28 participants responded and of respondents, 4% <5 years, 57% 5-11 years, 18% 12-17 years, and 21% 18-49 years.Influenza illness in 2019-2020 was initially influenza B, and subsequently replaced by influenza A. Most cases were in children and adolescents, despite at least one dose of influenza vaccine. Symptoms were widely distributed and similar between influenza A and B. Influenza incidence in our cohort declined to zero with the rise of SARS-CoV-2 cases and widespread mitigation efforts.Janet A. Englund, MD, AstraZeneca: Advisor/Consultant|AstraZeneca: Grant/Research Support|GlaxoSmithKline: Grant/Research Support|Meissa Vaccines: Advisor/Consultant|Merck: Grant/Research Support|Pfizer: Grant/Research Support|Sanofi Pasteur: Advisor/Consultant Helen Y. Chu, MD, MPH, Cepheid: Reagents|Ellume: Advisor/Consultant|Gates Ventures: Grant/Research Support|Merck: Advisor/Consultant|Pfizer: Advisor/Consultant."}
+{"text": "T790M mutation, the major mechanism of acquired resistance to first-generation EGFR TKI. However, resistance to AZD9291 arises eventually and EGFRC797S mutation was reported to be a major resistance mechanism. Thus, it is highly valuable to develop novel EGFR fourth-generation inhibitors targeting C797S mutation to override the acquired resistance. In this study, we identified HCD3514 as a novel EGFR fourth-generation inhibitors targeting C797S triple mutation. It strongly inhibited EGFRL858R/T790M/C797S and EGFR19del/T790M/C797S mutations with IC50 values of 1.0 and 2.0 nM, respectively. HCD3514 dose-dependently inhibited the activation of EGFR in both engineered BaF3 cells and tumor cells harboring EGFRC797S triple mutant and thus effectively suppressed the proliferation of the cells. Moreover, HCD3514 caused a dose-dependent increase of apoptosis in C797S triple mutant cells accompanied by increased levels of cleaved caspase-3 and cleaved PARP. Furthermore, HCD3514 induced tumor growth inhibition in EGFR19del/T790M/C797S xenograft model as a single oral agent by decreasing the activation of EGFR. In addition to EGFRC797S triple mutations, HCD3514 also potently and selectively inhibited EGFRT790M double mutations (L858R/T790M and 19del/T790M). Collectively, HCD3514 is a highly selective and potent EGFR inhibitor against EGFRC797S triple mutations as well as EGFRT790M double mutations and is confirmed potently anti-tumor activity in preclinical models.Osimertinib (AZD9291), a third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI), has significantly improved the survival of non-small cell lung cancer (NSCLC) patients with EGFR WT), resulting in severe side effects Lung cancer is a malignant tumor with high incidence and mortality throughout the world, being responsible for almost one-quarter of all cancer-related deaths T790M/C797S resistance mutation has gained much attention.Consequently, the urgent need for highly selective third-generation EGFR TKIs that target both activating mutations and T790M resistance mutation leads to the development of osimertinib (AZD9291). AZD9291 exhibits remarkable efficacy in treating patients of T790M mutation-positive NSCLC in vitro and in vivo, but quite similar to that with EAI045 treatment, its antitumor efficacy is limited as a single agent T790M/C797S inhibitors showing potential antitumor activity of AZD9291-resistant triple mutant EGFR in vitro and in vivo, including TQB3804, LS-106 T790M. Typical examples include BI-4020 Currently, encouraging achievements have been made in the development of the fourth-generation EGFR TKIs, which could be summarized into two categories based on their discovery strategies. 1) Exploiting allosteric binding pocket that is remote from the location of C797S mutation. For example, EAI045, an allosteric inhibitor, is reported as the first fourth-generation EGFR inhibitor overcoming T790M and C797S resistance mutation T790M inhibitor AZD9291, we identified a potent fourth-generation EGFR inhibitor, HCD3514, which is effective as a single agent to overcome EGFR triple mutation (EGFRL858R/T790M/C797S and EGFR19del/T790M/C797S) and shows promising antitumor activity both in vitro and in vivo.In this study, through structural hybridization of brigatinib with EGFR2.BaF3 cells and PC-9 cells were obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ) and the European Collection of Authenticated Cell Cultures (ECACC), respectively. MRC-9 cells and GSE-1 cells were purchased from the American Type Culture Collection (ATCC). All the cells were maintained in RPMI-1640 medium (Gibco) or EMEM medium (Gibco) supplemented with 10% FBS (Gibco) at 37 \u00b0C in 5% COL858R/T790M/C797S, EGFR19del/T790M/C797S, EGFRL858R/T790M or EGFR19del/T790M and cells stably expressing these mutants were subsequently selected in medium supplemented with 1 \u03bcg/mL puromycin (Sigma-Aldrich). PC-9-OR cells harboring EGFR19del/T790M/C797S mutation were constructed by CRISPR/Cas9 genome-editing technology to simultaneously knock-in the T790M and C797S mutations into PC-9 cells harboring EGFR19delBaF3 cells were transduced with retroviruses harboring genes encoding EGFR1H NMR and 13C NMR spectra was recorded on a Bruker AV-400 spectrometer at 400 MHz and Bruker AV-600 spectrometer at 151 MHz, respectively, in CDCl3. Coupling constants (J) are expressed in hertz (Hz). Chemical shifts (\u03b4) of NMR are reported in parts per million (ppm) units relative to internal control (TMS). The first-order peak patterns are indicated as s (singlet), d (doublet), t (triplet), q (quadruplet). Complex non-first order signals are indicated as m (multiplet). The high-resolution ESI-MS results were recorded on an Applied Biosystems Q-STAR Elite ESI LC-MS/MS mass spectrometer. The purity of compound was determined by reverse-phase high-performance liquid chromatography (HPLC) analysis using an Agilent 1260 system (G1310B Iso pump and G1365D MWD VL detector) with an YMC Triart C18 reversed-phase column  at 254 nm. Elution was MeOH in water, and flow rate was 1.0 mL/min.1H NMR  \u03b4 8.43 , 8.41 , 8.35 , 7.96 , 7.43 , 7.33 , 7.24 , 7.06 , 6.87 , 4.58 - 4.42 , 3.60 - 3.53 , 3.41 , 2.79 - 2.63 , 2.62 - 2.43 , 2.43 - 2.36 , 2.34 , 1.99 - 1.90 , 1.79 - 1.67 , 1.28 , 1.26 , 1.24 .13C NMR  \u03b4 158.43, 158.22, 156.93, 150.71, 139.15, 134.79, 133.85, 130.39, 128.78, 125.49, 124.13, 123.55, 118.64, 117.55, 116.60, 113.01, 112.83, 108.99, 70.38, 61.96, 55.46 (2C), 50.81 (2C), 49.13, 48.94, 46.05, 28.80 (2C), 22.18 (2C), 8.14, 1.05. HRMS (m/z): [M+H] + calculated for C33H42ClN7O3S, 652.2831; found, 652.2814. HPLC purity: 98.1%. Brigatinib (cat. no. S8229) and AZD9291 (cat. no. S7297) were purchased from Selleck Chemicals.Compound HCD3514 was designed and synthesized by Ke Ding laboratory. L858R/T790M/C797S: cat. no. E10-12VG, EGFR19del/T790M/C797S: cat. no. E10-12UG, EGFRL858R/T790M: cat. no. E10-122DG, EGFR19del/T790M: cat. no. E10-122KG) or Eurofins Scientific (EGFRWT: cat. no. 14-531M). ELISA was used to evaluate the inhibitory activity of test compounds against EGFR, and the experiment was performed as described previously The recombinant EGFR proteins of the kinase domain were purchased from SignalChem Lifesciences (EGFRL858R/T790M/C797S was selected as the template to elucidate the binding mode of HCD3514. Protein structure was downloaded from Protein Data Bank (PDB 6LUD). The EGFR enzyme was defined as a receptor and the docking grid was centered on the ligand binding location. Docking simulations were performed in standard precision (SP) mode. Other parameters were set as default. After accomplishment of the molecular docking procedure, eight docking poses were scored and selected based on calculated energy.The molecular docking procedure was performed within Glide 7.9 software BaF3 cells, PC-9-OR cells, MRC-9 cells and GSE-1 cells were plated in 96-well plates in the corresponding medium and incubated overnight, followed by exposing to medium containing serial dilutions of compounds . After 72 h of drug treatment, cell counting kit-8 (CCK8) or sulforhodamine B (SRB) was added to each well and then measured by SoftMax Pro software at the absorbance of 450 nm or 560 nm Western blotting was performed as described previously L858R/T790M/C797S cells, BaF3-EGFR19del/T790M/C797S cells and PC-9-OR cells were seeded in 6-wells plates overnight, and exposed to indicated concentrations of compounds HCD3514 or brigatinib for 48 h. Then cells were collected and washed with PBS and measured by Annexin V-FITC Apoptosis detection kit . Signals were measured using FACS Calibur flow cytometer, and FlowJo software was used to analyze the data.BaF3-EGFR19del/T790M/C797S cells (2 \u00d7 106) were injected subcutaneously into BALB/c nude mice (Shanghai institute of medicine). After animals were randomly assigned to groups, tumor-bearing mice were orally administered vehicle, test compound HCD3514 or AZD9291 at the indicated doses once a day and tumor size were measured once per week. Tumor volume was calculated as length \u00d7 width2 \u00d7 0.5 (mm3) and the tumor growth inhibition (TGI) = [1-RTV (treated) / RTV (control)] \u00d7 100%. Mice were sacrificed and tumors were harvested for Western blot analysis. All animal study was approved by the Institutional Animal Care and Use Committee of the Shanghai Institute of Materia Medica and strictly performed according to the institutional ethical guidelines on animal care.BaF3-EGFRP < 0.05, **P < 0.01, ***P < 0.001.Data were presented as mean \u00b1 Standard Deviation (SD) and were analyzed by GraphPad Prism 8.0 software. Statistical analysis of the difference between vehicle and compound treated groups was compared by a two-tailed Student's t-test. Statistical significance was defined as *T790M/C797S, we incorporated important pharmacophore of brigatinib to the 4-indolyl-2-phenylaminopyrimidine scaffold of EGFRT790M inhibitor AZD9291 and developed a novel series of potent and mutant selective EGFRT790M/C797S inhibitor. Following extensive medicinal chemistry optimization by adding H-bond receptor of the indole ring and hydrophobic moiety on the benzene ring, we identified HCD3514 as a selective EGFRT790M/C797S inhibitor  and EGFR19del/T790M/C797S (IC50 = 2.0 \u00b1 0.3 nM) with a more than 78-fold selectivity over EGFRWT (IC50 = 156.0 \u00b1 6.9 nM). By comparison, HCD3514 possessed more potent kinase inhibitory activity against EGFR triple mutations than brigatinib  and more potent than AZD9291 (IC50 = 6.30 \u03bcM)  and gastric mucosal (GSE-1) cell lines. As shown in Supplementary M Figure A, indicain vitro anti-tumor activity of HCD3514 in EGFRT790M/C797S triple mutant cells through suppression of EGFR phosphorylation, leading to the inhibition of cellular proliferation.Thus, these results demonstrated the L858R/T790M/C797S cells, HCD3514 triggered significant apoptosis with apoptosis rates of 72.39% and 98.27% at the concentration of 1 \u03bcM and 3 \u03bcM, respectively  and cysteinyl aspartate specific proteinase-3 (caspase-3) play an important role in cellular processes, including apoptosis To gain deeper insight into the potential mechanism of apoptosis induction in EGFRin vivo in BaF3-EGFR19del/T790M/C797S xenograft mouse model. BALB/c nude mice bearing established BaF3-EGFR19del/T790M/C797S mouse xenograft tumors were daily oral treatment with HCD3514 (50 and 75 mg/kg), AZD9291 (25 mg/kg) or vehicle control. Consequently, HCD3514 resulted in an obvious growth inhibition of BaF3-EGFR19del/T790M/C797S mouse xenograft tumors in a dose-dependently manner with tumor growth inhibition (TGI) values of 22.53% and 37.73% at the dosage of 50 mg/kg and 75 mg/kg, respectively, while AZD9291 treatment did not show significant reduction of tumor growth in the xenograft model compared to the vehicle treated mice , as suggested by the molecular docking experiment showed in Figure L858R/T790M and EGFR19del/T790M mutations with IC50 values of 4.9 and 8.9 nM in biochemical, respectively. Moreover, HCD3514 showed antiproliferative activities in BaF3-EGFRL858R/T790M and BaF3-EGFR19del/T790M cells with IC50 of 0.66 \u03bcM and 0.52 \u03bcM, respectively, although the IC50 values of HCD3514 were higher than that of AZD9291. Furthermore, Western blot analysis revealed that HCD3514 dose-dependently suppressed the EGFR phosphorylation in both BaF3-EGFRL858R/T790M cells and BaF3-EGFR19del/T790M cells. Besides, we also evaluated the in vitro anti-tumor activity of HCD3514 in NCI-H1975 cells harboring EGFRL858R/T790M. HCD3514 also displayed an excellent antiproliferative potency in NCI-H1975 cells with IC50 of 0.49 \u03bcM and the expression of phospho-EGFR was down-regulated after treatment with HCD3514 in a dose-dependent manner. Thus, HCD3514 could target EGFRT790M double mutations as well as EGFRT790M/C797S triple mutations.In addition, we checked whether HCD3514 inhibits EGFR-double mutations  exploiting allosteric binding pocket by allosteric inhibitors. 2) developing ATP-competitive inhibitors by structural modification of ALK/EGFR inhibitor brigatinib or the third-generation EGFR TKIs.Oncogenic mutations in EGFR are prevalent in NSCLC. Though third-generation EGFR inhibitor AZD9291 has achieved a great clinical benefit in NSCLC patients, the EGFRT790M inhibitor AZD9291, we identified a new fourth-generation EGFR triple mutant-selective inhibitor HCD3514, which possessed a promising antitumor activity both in vitro and in vivo. Compared to AZD9291, molecular docking of HCD3514 identified an additional hydrogen bond interaction towards Lys745, which was proposed to compensate for the loss of covalent bound due to the C797S point mutation and ultimately enhanced the inhibitory efficacy of HCD3514 on EGFRC797S mutations. HCD3514 displayed a single digit-nanomolar kinase inhibitory activity against triple mutant EGFR, including EGFRL858R/T790M/C797S and EGFR19del/T790M/C797S, which showed superior biochemical activity than brigatinib. Importantly, HCD3514 exhibited a sufficient EGFRWT sparing window, suggesting its low toxicity. In addition, HCD3514 simultaneously inhibited the activities of both mutant forms of EGFR at the cellular level. It showed that both engineered BaF3 cells harboring EGFRL858R/T790M/C797S or EGFR19del/T790M/C797S were sensitive to HCD3514, as well as the PC-9-OR cells (EGFR19del/T790M/C797S expressing PC-9 lung cancer cells), with enhanced inhibition the phosphorylation of EGFR. Besides, cleaved PARP and cleaved caspase-3 levels were showed to increase in three C797S mutant cells, which was consistent with apoptosis induction in a concentration-dependent manner. Furthermore, unlike EAI045 or brigatinib which showed limited antitumor effect without combined with anti-EGFR antibody C797S triple mutations but also EGFRT790M double mutations.In this study, through structural hybridization of brigatinib with EGFRin vitro and in vivo and we also provided a novel chemical scaffold and a new lead compound that worthy of further investigation.In summary, our preclinical data demonstrated that HCD3514 was effective in overcoming both triple mutant and double mutant forms of EGFR-dependent resistance mechanisms Supplementary figures.Click here for additional data file."}
+{"text": "Assessing serological inflammation is difficult in tocilizumab (TCZ)-treated rheumatoid arthritis (RA) patients, as standard inflammation parameters, like erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP), are influenced by interleukin-6-receptor inhibition. Calprotectin in the serum, also named S100A8/S100A9, might be a more useful inflammation parameter in TCZ-treated patients.Sixty-nine RA patients taking TCZ were included. Serum-calprotectin levels were assessed, as well as ESR, CRP, need for a change in disease-modifying anti-rheumatic drugs due to RA activity (= active RA), and the RA clinical disease activity score (CDAI). Forty-five RA patients taking tumor-necrosis factor-inhibitors (TNFi) were investigated for the same parameters.P < 0.001). A calprotectin cut-off value of 1916.5 ng/ml resulted in a sensitivity and specificity of 80.0 %, respectively, for the detection of RA disease activity. Calprotectin values correlated with CDAI-scores . ESR and CRP were less suitable to detect RA activity in TCZ-treated patients. Also TNFi-treated patients with active RA had higher calprotectin values compared to not active RA . The calprotectin value with the best sensitivity and specificity for detecting RA activity was 3690.5 ng/ml among TNFi-treated patients.TCZ-treated patients with active RA had higher calprotectin values than not active RA patients (4155.5 [inter quartile range 1865.3\u20136068.3] vs 1040.0 [676.0\u20131638.0] ng/ml, Calprotectin in the serum can be a useful inflammation parameter despite TCZ-treatment.The online version contains supplementary material available at 10.1186/s13075-022-02887-7. Inflammation parameters in the serum are a useful tool to monitor disease activity in inflammatory rheumatic diseases. In rheumatoid arthritis (RA), mostly erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) are routinely assessed. CRP and acute phase protein synthesis in the liver is regulated by interleukin-6 (IL-6) , 2. TociCalprotectin, a heterodimer of the two calcium-binding proteins S100A8 and S100A9 , is a major cytosolic protein in monocytes and neutrophils, secreted during infections, malignancy, and inflammation , 6. CalpThe measurement of calprotectin in the serum as inflammation parameter in RA (without IL-6 receptor inhibitors) has been investigated and showed a correlation with radiographic damage scores and standard inflammation parameters . It alsoFew data is available for the use of calprotectin in RA with TCZ-treatment. Only one study of 33 patients is published , suggestOne hundred fourteen patients, fulfilling the 2010 ACR/EULAR classification criteria for rheumatoid arthritis , were inThe CDAI, comprising the sum of 28 swollen joints, 28 tender joints, patient global, and physician global assessment , was assSerum calprotectin was collected in serum-gel vials  and measured with EliA\u2122 Calprotectin 2 wells  with a Phadia\u2122 250 device . For each measurement cycle, a commercial negative and positive control (Thermo Fisher Scientific) was analyzed. Serum samples were centrifuged within 1h after blood collection and stored between 2 and 8\u00b0C until measurement. Stable calprotectin values can be measured with this assay within 7 days .ESR (reference range in the first hour 3\u20138 mm) and CRP (reference < 0.5 mg/dl) were measured in the central laboratory of the University Hospital of W\u00fcrzburg. CRP was detected by immunoturbidimetry .U tests were performed for continuous variables and Fisher\u2019s exact tests for categorical variables. Spearman\u2019s tests were used to calculate correlations. Receiver-operator characteristic (ROC) curves were calculated to detect cut-off values with the best sensitivity and specificity. SPSS Statistics v 28.0  was used for statistical analysis. For data collection Excel  was used. Figures were grouped by using Photoshop . Two-tailed P-values less than 0.05 were considered significant.Shapiro-Wilk tests were used to test for normal distribution. As normal distribution was mostly absent, medians with interquartile ranges (IQR) were shown. To test for differences between unpaired groups, Mann-Whitney Inflammation parameters were obtained from 69 RA patients. All patients received TCZ for at least 3 months. Forty-eight of 69 (69.6 %) were female; the median age was 64.0 years (range 33\u201389). Fifty-four of 69 (78.3 %) were rheumatoid factor (RF)-positive; 51/69 (73.9 %) were anti-citrullinated protein antibody (ACPA)-positive. All ACPA-positive patients were RF-positive. Fifty of 69 (72.5 %) received TCZ subcutaneously and 19/69 (27.5 %) intravenously. No concomitant DMARD was present in 43/69 (62.3 %) patients. Thirteen of 69 (18.8 %) received prednisolone , 12/69 (17.4 %) methotrexate (MTX), and 1/69 (1.4 %) lefunomide (LEF). The median activity parameters of the TCZ-treated patients were as follows: DAS28-ESR 1.5 (IQR 1.1\u20132.2), DSA28-CRP 1.7 (1.2\u20132.2), SDAI 5.0 (2.0\u20138.0), and CDAI 5.0 (2.0\u20138.0). The median inflammation parameters of the TCZ-treated patients were as follows: ESR in 1h 4.0 (2.0\u20136.0) mm, CRP 0.0 (0.0\u20130.1) mg/dl, and calprotectin 1072.0 (686.5\u20131916.5) ng/ml. Patients\u2019 characteristics are summarized in Table n = 115) compared to the active patients (n = 10) (1040.0 (676.0\u20131638.0) ng/ml vs 4155.5 [1865.3\u20136068.3] ng/ml, P < 0.001)  one group of patients, who continued their DMARD regimen (i.e. tocilizumab + concomitant medication) (not active), and (2) patients, who needed a change or escalation of their DMARDs due to inflammatory activity of RA (active). One hundred twenty-five calprotectin values were included in this study as from some patients more than one time point was measured. The calprotectin value in the serum was significantly lower in the not active RA patients ( one grouP < 0.001)   had a lower median calprotectin value compared to patients with CDAI-low activity (n = 67) . Patients with CDAI-low activity had lower median calprotectin levels than patients with CDAI-moderate activity (n = 17) . Patients with CDAI-moderate disease activity had lower median calprotectin levels than patients with CDAI-high activity (n = 2)   Fig. a and sigr = 0.301; P < 0.001), with DAS28-CRP , and with SDAI   with IL-6-dependent composite scores was detected: with DAS28-ESR  were included in this study. Patients\u2019 characteristics including sex, age, RF/ACPA-status, concomitant DMARDs, median activity scores, and median inflammation parameters are embedded in Table n = 43) had a lower serum-calprotectin (1845.0 [832.0\u20132569.0] ng/ml) than active RA patients (n = 10) receiving a change of their TNFi due to inflammatory activity of RA    Fig. d.r = 0.401; P = 0.001)  had a median calprotectin value of 1694.0 [773.5\u20132983.0] ng/ml, which was not significantly different from patients with CDAI-low activity (n = 26) . Patients with CDAI-low activity had lower median calprotectin levels than patients with CDAI-moderate activity (n = 5) . Patients with CDAI-moderate disease activity had numerically but not significantly lower median calprotectin levels than patients with CDAI-high activity (n = 2)  , with DAS28-CRP , and with SDAI   Fig. d. Patien Fig. d. P = 0.021, Fig. P < 0.001, Fig. In contrast to TCZ-treated patients, in TNFi-treated RA patients ESR and CRP showed differences between not active and active patients    and also found significant correlations between calprotectin values and RA-activity composite scores , which iDisease activity scores, including DAS28-ESR and DAS28-CRP, which contain serological inflammation parameters for their calculation (to a relevant extent), were lower in our TCZ-treated group than in the TNFi-group. In prior studies, RA patients receiving TCZ-treatment had higher DAS28-ESR remission rates than CDAI or SDAI remission rates, which was supposed to be due to the high weight of the ESR in the DAS28-ESR score . TherefoIt was formerly shown that RA patients with detectable TCZ trough levels (after intravenous application) did not have lower calprotectin values than patients with undetectable TCZ trough levels, in contrast to the CRP values, which were lower when TCZ was detectable . This suThe CDAI score is usually assessed directly after each patient\u2019s visit without having the inflammatory parameters available at that time. This was also the case in our study: The CDAI scores were assessed without knowing the respective calprotectin values (neither ESR nor CRP). Thus, the decision of the investigator on the CDAI score was not biased by the knowledge of the inflammatory parameters.Concerns over the pre-analytical factors of serum-calprotectin values have been raised, but recently it was demonstrated that serum-calprotectin is stable for 1 week when kept refrigerated . This siOur study is limited due to its retrospective design and single-center approach. Additionally, future longitudinal studies are needed to evaluate calprotectin as follow-up parameter and its usefulness in detecting treatment response. Most of our patients had a low disease activity with a median CDAI value of 5, so that high inflammation parameters were only included in our study to a limited extent.Additional file 1: Supplemental Figure S1 Correlation of interleukin-6-dependend composite scores including DAS28-ESR , DAS28-CRP  and SDAI  with calprotectin in tocilizumab (TCZ)-treated RA patients (a-c) and in tumor necrosis factor alpha-inhibitor (TNFi)-treated RA (d-f) patients."}
+{"text": "Salix species, including willow trees, are distributed in the temperate regions of Asian countries, including South Korea. Willow trees are used to treat pain and inflammatory diseases. Due to the medicinal properties of willow trees, pharmacological studies of other Salix spp. have gained attention; however, only a few studies have investigated the phytochemicals of these species. As part of our ongoing natural product research to identify bioactive phytochemicals and elucidate their chemical structures from natural resources, we investigated the marker compounds from indigenous Korean Salix species, namely, Salix triandra, S. chaenomeloides, S. gracilistyla, S. koriyanagi, S. koreensis, S. pseudolasiogyne, S. caprea, and S. rorida. The ethanolic extract of each Salix sp. was investigated using high-performance liquid chromatography combined with thin-layer chromatography and liquid chromatography\u2013mass spectrometry-based analysis, and marker compounds of each Salix sp. were isolated. The chemical structures of the marker compounds (1\u20138), 3-(4-hydroxyphenyl)propyl \u03b2-D-glucopyranoside (1), 2-O-acetylsalicin (2), 1-O-p-coumaroyl glucoside (3), picein (4), isograndidentatin B (5), 2\u2032-O-acetylsalicortin (6), dihydromyricetin (7), and salicin (8) were elucidated via nuclear magnetic resonance spectroscopy and high-resolution liquid chromatography\u2013mass spectrometry using ultrahigh-performance liquid chromatography coupled with a G6545B Q-TOF MS system with a dual electrospray ionization source. The identified marker compounds 1\u20138 were examined for their antimicrobial effects against plant pathogenic fungi and bacteria. Dihydromyricetin (7) exhibited antibacterial activity against Staphylococcus\u00a0aureus, inducing 32.4% inhibition at a final concentration of 125 \u03bcg/mL with an MIC50 value of 250 \u03bcg/mL. Overall, this study isolated the marker compounds of S. triandra, S. chaenomeloides, S. gracilistyla, S. koriyanagi, S. koreensis, S. pseudolasiogyne, S. caprea, and S. rorida and identified the anti-Staphylococcus aureus bacterial compound dihydromyricetin. Salix comprises approximately 500 species of deciduous trees and shrubs distributed in the temperate regions of Asian countries, including South Korea, and willow trees are the most representative plants of this genus  anthracene-induced phorbol ester-promoted skin carcinogenesis [S. caprea. Finally, S. rorida is a deciduous tree and a willow species native to Japan, northern China, Korea, and Russia. A recent phytochemical study revealed salicin as the major constituent of S. rorida and identified other constituents, including (+)-catechin, naringenin, salipurposide, aromadendrin, isosalipurposide, aromadendrin-7-O-\u03b2-D-glucopyranoside, and taxifolin-7-O-\u03b2-D-glucopyranoside, via liquid chromatography\u2013mass spectrometry (LC/MS) analysis [S. rorida has not yet been elucidated.l (DPPH) . Despiteivatives ,11, whicinkiller . A pharmrophages . In additivities . Howeverl crafts . S. koripriority , and fewin Korea . S. koreth decay . A pharmer cells . S. kore effects ,21,22. Dnd fever  Recent p effects ,25. Prevextracts ,24. S. cSalix species S. triandra, S. chaenomeloides, S. gracilistyla, S. koriyanagi, S. koreensis, S. pseudolasiogyne, S. caprea, and S. rorida. Ethanolic extracts of these Salix spp. were investigated using high-performance liquid chromatography (HPLC) combined with thin-layer chromatography (TLC) and LC/MS-based analysis, followed by the isolation of each marker compound of Salix spp. The chemical structures of marker compounds (1\u20138) were elucidated via nuclear magnetic resonance (NMR) spectroscopy and electrospray ionization (ESI) LC/MS analyses. Finally, the identified marker compounds 1\u20138 were tested for their antimicrobial effects. Herein, we report the isolation and structural characterization of marker compounds 1\u20138 as well as their bioactivity with respect to their antimicrobial effects.As part of our ongoing research projects to identify bioactive natural products and elucidate their chemical structures from natural resources ,30,31,32Salix species  were collected from Chungcheongnam-do, Chungcheongbuk-do, and Gangwon-do (South Korea), thoroughly dried, and mounted. Plant specimens  to obtain EtOH crude extracts.The twigs of eight pecimens  were madSalix species  facilitated the determination of the marker compounds for these species based on the amount of the component present in each extract. Phytochemical investigation of the crude EtOH extracts was performed via TLC and LC/MS-based analysis using column chromatography and HPLC to isolate the marker compounds (1) was isolated from S. triandra, 2-O-acetylsalicin (2) from S. chaenomeloides, 1-O-p-coumaroyl glucoside (3) from S. gracilistyla, picein (4) from S. koriyanagi, isograndidentatin B (5) from S. koreensis, 2\u2032-O-acetylsalicortin (6) from S. pseudolasiogyne, dihydromyricetin (7) from S. caprea, and salicin (8) from S. rorida propyl \u03b2-D-glucopyranoside (1) [O-acetylsalicin (2) [O-p-coumaroyl glucoside (3) [4) [5) [O-acetylsalicortin (6) [7) [8) [The marker compounds of side (1) , 2-O-aceicin (2) , 1-O-p-cside (3) , picein  (3) [4) , isogran [4) [5) , 2\u2032-O-acrtin (6) , dihydro (6) [7) , and sal [7) [8)  . Next, the marker compounds were tested for their antibacterial activity against the Gram-positive bacterium Staphylococcus aureus (HG003) and Gram-negative bacterium Escherichia coli (MG1655). Among the isolates, only dihydromyricetin (7) exhibited antibacterial activity against S. aureus, inducing 32.4% inhibition at a final concentration of 125 \u03bcg/mL with an MIC50 value of 250 \u03bcg/mL (1) and 1-O-p-coumaroyl glucoside (3) exhibited very weak activity against S. aureus, inducing only 1.9% and 1.3% inhibition, respectively . Twigs of S. chaenomeloides, S. koriyanagi, and S. caprea were collected from Inje-gun and those of S. rorida from Hwacheon-gun . The twigs of S. gracilistyla and S. koreensis were collected from Jecheon-si . Each material was authenticated by the author J.N.Yu. Voucher specimens of the materials  were deposited at the Hongcheon Institute of Medicinal Herbs, Hongcheon-gun, South Korea.In June 2021, twigs of S. triandra twigs (1.6 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (86.6 g). The extract (10.1 g) was dissolved in MeOH (100 mL) and applied to a reverse-phase (RP) Sep-Pak column with 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (5.2 g). The crude extract (1.0 g) was separated via preparative RP-HPLC  to obtain four fractions (P1\u2013P4). Fraction P2 (303.3 mg) was isolated via semipreparative RP-HPLC using 30% MeOH to obtain marker compound 1 .S. chaenomeloides twigs (1.8 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (144.6 g). The extract (10.3 g) was dissolved in MeOH (100 mL) and applied to an RP Sep-Pak column with 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (7.2 g). The crude extract (1.0 g) was separated via preparative RP-HPLC  to obtain five fractions (P1\u2013P5). Fraction P4 (68.2 mg) was isolated via semipreparative RP-HPLC using 20% MeOH to obtain marker compound 2 .S. gracilistyla twigs (1.0 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (163 g). The extract (11 g) was dissolved in MeOH (100 mL) and applied to an RP Sep-Pak column with 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (5.6 g). The crude extract (1.0 g) was separated via preparative RP-HPLC  to obtain five fractions (P1\u2013P5). Fraction P4 (468 mg) was isolated via semipreparative RP-HPLC using 23% MeOH to obtain marker compound 3 .S. koriyanagi twigs (2.0 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (167 g). The extract (10 g) was dissolved in MeOH (100 mL) and applied to an RP Sep-Pak column with 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (6.5 g). The crude extract (6.5 g) was subjected to HP-20 column chromatography, with eluting solvents of distilled water, MeOH, and acetone, to obtain three fractions (P1\u2013P3). Fraction P1 (2.6 g) was separated via preparative RP-HPLC  to obtain five fractions (P11\u2013P15). Fraction P13 (245 mg) was isolated via semipreparative RP-HPLC using 15% MeOH to obtain marker compound 4 .S. koreensis twigs (2.3 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (101 g). The extract (9.1 g) was dissolved in MeOH (100 mL) and applied to an RP Sep-Pak column using 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (7.9 g). The crude extract (7.9 g) was subjected to HP-20 column chromatography, with the eluting solvents distilled water, MeOH, and acetone to obtain three fractions (P1\u2013P3). Fraction P3 (582 mg) was separated via preparative RP-HPLC  to obtain four fractions (P31\u2013P34). Fraction P33 (582 mg) was isolated via semipreparative RP-HPLC using 43% MeOH to obtain marker compound 5 .S. pseudolasiogyne twigs (1.5 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (123.6 g). The extract (9.1 g) was dissolved in MeOH (100 mL) and applied to an RP Sep-Pak column using 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (4.0 g). The crude extract (4.0 g) was separated via preparative RP-HPLC  to obtain four fractions (P1\u2013P4). Fraction P4 (306 mg) was isolated via semipreparative RP-HPLC using 39% MeOH to obtain marker compound 6 .S. caprea twigs (1.5 kg) were dried at 35\u201345\u2103 in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (110.8 g). The crude extract (10.3 g) was separated using medium-pressure liquid chromatography (MPLC)  to yield seven fractions (P1\u2013P7). Fraction P4 (507 mg) was isolated via semipreparative RP-HPLC using 27% MeOH to obtain marker compound 7 .S. rorida twigs (1.5 kg) were dried at 35\u201345 \u00b0C in a plant-drying oven for one week, pulverized, and extracted with 80% ethanol (10 L) via sonication for 90 min three times at room temperature. The filtered ethanol extract was evaporated in vacuo to obtain the crude ethanol extract (143.3 g). The extract (10.5 g) was dissolved in MeOH (100 mL) and applied to an RP Sep-Pak column with 100% MeOH to remove the wax, lipids, and fatty acids, and the resultant residue was concentrated using an evaporator to obtain the crude extract (7.8 g). The crude extract (7.8 g) was separated by MPLC  to obtain four fractions (P1\u2013P4). Fraction P2 (218 mg) was isolated via semipreparative RP-HPLC using 15% MeOH to obtain marker compound 8 .B. cinerea (KACC40965), C. destructans (KACC 41077), F. solani (KACC 44891), F. asiaticum (KACC 46429), and R. solani (KACC 48921), were used for the in vitro antifungal activity assay. Fungi were cultivated on 25 mL of malt extract agar  at 25 \u00b0C for 14 d in the dark. Marker compounds (1\u20138) were evaluated for their antifungal activities in a 96-well microplate [6 cells/mL. The absorbance at 600 nm was measured every 12 h for 96 h. The experiments were performed in triplicate. The percentage of growth inhibition (GI%) was estimated using the following formula: GI% = 100 (Acontrol \u2212 Atest sample)/(Acontrol).Five plant pathogenic fungi provided by the Korean Agricultural Culture Collection . m KACC 469, and R.S. aureus (HG033) and the Gram-negative bacterium E. coli (MG1655) were used. Staphylococcus aureus strains were maintained aerobically in tryptic soy broth  or TSB 1.5% agar at 30 \u00b0C with shaking at 250 rpm. E. coli strains were maintained aerobically in lysogeny broth  or LB 1.5% agar at 37 \u00b0C with shaking at 250 rpm. The qualitative antibacterial activities of the marker compounds (1\u20138) were examined using the disk diffusion assay and MIC values [1\u20138). The marker compounds were prepared in DMSO (1%) at 100 \u03bcg/mL . The assay was performed with LB and TSB plates. Sterile beads were used to inoculate the surface of each plate to ensure homogeneous bacterial growth. Sample pellet disks were placed on the surface of the agar plates at equal distances. The inhibition was measured using a ruler after 24 h incubation at 30 and 37 \u00b0C. MIC assay was performed in a 96-well plate to examine the antibacterial effects of three marker compounds . Following serial dilution of the compounds  in 150 \u03bcL of Mueller\u2013Hinton broth, the culture was inoculated at an optical density (OD) of 0.002. After incubation at 30 and 37 \u00b0C for 24 h with shaking, bacterial growth was measured at OD600 using a spectrophotometer (BioTek Synergy HTX).For the antibacterial activity assay, the Gram-positive bacterium C values . The disSalix species  were phytochemically investigated, followed by the isolation of their marker compounds: 3-(4-hydroxyphenyl)propyl \u03b2-D-glucopyranoside (1) from S. triandra, 2-O-acetylsalicin (2) from S. chaenomeloides, 1-O-p-coumaroyl glucoside (3) from S. gracilistyla, picein (4) from S. koriyanagi, isograndidentatin B (5) from S. koreensis, 2\u2032-O-acetylsalicortin (6) from S. pseudolasiogyne, dihydromyricetin (7) from S. caprea, and salicin (8) from S. rorida. The chemical structures of these compounds were also determined via NMR spectroscopy and HR-ESI-MS analysis. Antimicrobial tests of the marker compounds (1\u20138) using plant pathogenic fungi and bacteria revealed that dihydromyricetin (7) exhibited antibacterial activity against S. aureus, inducing 32.4% inhibition at a final concentration of 125 \u03bcg/mL, with an MIC50 value of 250 \u03bcg/mL.In the present study, indigenous Korean"}
+{"text": "Ubiquitin-conjugating enzyme E2 T (UBE2T) is a potential oncogene. However, Pan-cancer analyses of the functional, prognostic and predictive implications of this gene are lacking.We first analyzed UBE2T across 33 tumor types in The Cancer Genome Atlas (TCGA) project. We investigated the expression level of UBE2T and its effect on prognosis using the TCGA database. The correlation between UBE2T and cell cycle in pan-cancer was investigated using the single-cell sequencing data in Cancer Single-cell State Atlas (CancerSEA) database. The Weighted Gene Co-expression Network analysis (WGCNA), Univariate Cox and Least absolute shrinkage and selection operator (LASSO) Cox regression models, and receiver operating characteristic (ROC) were applied to assess the prognostic impact of UBE2T-related cell cycle genes (UrCCGs). Furthermore, the consensus clustering (CC) method was adopted to divide TCGA-lung adenocarcinoma (LUAD) patients into subgroups based on UrCCGs. Prognosis, molecular characteristics, and the immune panorama of subgroups were analyzed using Single-sample Gene Set Enrichment Analysis (ssGSEA). Results derived from TCGA-LUAD patients were validated in International Cancer Genome Consortium (ICGC)-LUAD data.1 year\u2009=\u20090.720, AUC3 year\u2009=\u20090.700, AUC5 year\u2009=\u20090.630). The\u00a0CC method classified the TCGA-LUAD cohort into 4 UrCCG subtypes (G1\u2013G4). Kaplan\u2013Meier survival analysis demonstrated that G2 and G4 subtypes had worse survival than G3 . A comprehensive analysis of immune infiltrates, immune checkpoints, and immunogenic cell death modulators unveiled different immune landscapes for the four subtypes. High immunophenoscore in G3 and G4 tumors suggested that these two subtypes were immunologically \u201chot,\u201d tending to respond to immunotherapy compared to G2 subtypes (p\u2009<\u20090.001).UBE2T is highly expressed and is a prognostic risk factor in most tumors. CancerSEA database analysis revealed that UBE2T was positively associated with the cell cycle in various cancers. The risk signature of UrCCGs can reliably predict the prognosis of LUAD (AUCUBE2T is a critical oncogene in many cancers. Moreover, UrCCG classified the LUAD cohort into four subgroups with significantly different survival, molecular features, immune infiltrates, and immunotherapy responses. UBE2T may be a therapeutic target and predictor of prognosis and immunotherapy sensitivity.The online version contains supplementary material available at 10.1186/s12931-022-02226-z. To date, targeted therapy has become the first-line therapy for cancer patients harboring corresponding oncogenic driver mutations. However, drug resistance frequently occurs, resulting from de novo drug-resistant mutations, compensatory activation of collateral signaling pathways, or growth dominance of cells with preexistent drug-resistant genetic alterations . In the UBE2T gene is located in chromosome 1q32.1, encoding a protein product composed of 197 amino acids , we used a web-based analysis tool, the Gene Expression Profiling Interactive Analysis (GEPIA) (http://gepia.cancer-pku.cn/) [https://www.genome.gov/Funded-Programs-Projects/Genotype-Tissue-Expression-Project). The GEPIA included the re-computed TCGA and GTEx data that were processed from corresponding raw RNA-Seq datasets by the UCSC Xena project based on a uniform pipeline [http://ualcan.path.uab.edu/analysis-prot.html) [https://www.proteinatlas.org/) was used to explore the subcellular distribution of UBE2T in tumors [We downloaded the RNA-Seq expression data (FPKM format) of 33 different tumors from the TCGA database (pku.cn/) , to comppipeline . Batch eot.html) , 19 (Addot.html) , 19. Then tumors .Clinical data were fetched from the TCGA, including overall survival (OS), disease-specific survival (DSS), progression-free interval (PFI), and disease-free interval (DFI) survival data. Samples without survival information were excluded.  (http://biocc.hrbmu.edu.cn/CancerSEA/) database to examine the association of UBE2T with 14 primary tumor-related cellular activities, such as cell cycle, proliferation, and angiogenesis [UBE2T (FDR\u2009<\u20090.05 and correlation\u2009>\u20090.3). The correlations between UBE2T and scores of functional states were visualized using the \u201cggplot2\u201d package. Our previous publication demonstrated that UBE2T promoted autophagy in LUAD; therefore, we further investigated the functional relevance of UBE2T in LUAD at the single-cell level. We chose EXP0066 (GSE69405) and EXP0067 (GSE85534) . The correlation between gene expression and immune infiltration was estimated by the Pearson correlation test  with the same settings  method to identify unsupervised intrinsic groups with similar biological characteristics . This mehttps://bioinformatics.mdanderson.org/estimate/disease.html) [The stromal score, immune score, and tumor purity were compared among different UrCCG subtypes by the ESTIMATE  method (se.html) . Moreovehttps://tcia.at/home). The IPS is calculated based on four significant categories of tumor immunogenicity determinants, including effector cells, immunosuppressive cells, major histocompatibility complex (MHC) molecules (antigen processing), and checkpoints/immunomodulators. The IPS, ranging from 0 to 10, was calculated based on the z-score for the expression of related genes . For all analyses. A two-tailed p\u2009<\u20090.05 was regarded as statistical significance if not noted.https://portal.gdc.cancer.gov/) , comprising gene expression in normal tissues from GETX (https://www.genome.gov/Funded-Programs-Projects/Genotype-Tissue-Expression-Project). Overall, UBE2T expression was significantly upregulated in various cancers. Additionally, GEPIA-based analysis showed that UBE2T was differentially expressed in different stages in adrenocortical carcinoma (ACC), invasive breast carcinoma (BRCA), head and neck squamous cell carcinoma (HNSC), kidney chromophobe (KICH), ovarian serous cystadenocarcinoma (OV), kidney renal papillary cell carcinoma (KIRP), liver hepatocellular carcinoma (LIHC), LUAD, kidney renal clear cell carcinoma (KIRC), and thyroid carcinoma (THCA)  confirmed that UBE2T protein levels were significantly upregulated in LUAD, uterine corpus endometrial carcinoma (UCEC), renal cell carcinoma (RCC), OV, and BRCA  revealed both cytoplasmic and nuclear distribution of UBE2T  Fig.\u00a0A. For inv/) Fig.\u00a0B, we comE2T Fig.\u00a0E.Fig. 1TWe investigated the clinical relevance of UBE2T in the different tumor types using Kaplan\u2013Meier survival analysis Fig.\u00a0: Fig. S5http://biocc.hrbmu.edu.cn/CancerSEA/) [We next investigated whether UBE2T regulates some fundamental cancer-associated cellular processes by mining the CancerSEA database (cerSEA/) . UBE2T whttp://timer.comp-genomics.org/) revealed the association of the eight genes with critical immune cell infiltration . Compared with the G3 group, G2 and G4 groups had higher enrichment scores on tumor-related signaling pathways, including cell cycle, autophagy, mismatch repair, and MTORC1 signaling, but lower scores on the B cell receptor and p53 signaling pathway  [Solid tumor tissue includes malignant cells, normal epithelial, stromal cells, immune cells, and vascular cells. The ssGSEA was performed on LUAD samples to analyze the differences in immune cell infiltration and tumor purity among different UrCCG subtypes by ESTIMATE (se.html) . G3 and Given the importance of immune checkpoints (ICPs)  and immuhttps://tcia.at/home) to determine the sensitivity to immune checkpoint inhibitors for the four subgroups, the most comprehensive immune determinant to date [Finally, we used immunophenoscore (IPS) for LUAD patients from The Cancer Immunome Atlas  genes have been reported \u201340. ProgFurthermore, based on the differential expression patterns of UrCCGs, CC analysis divided the TCGA-LUAD cohorts into G1 to G4 subtypes, with significantly different survival. G3 subtype patients had better survival than other subtypes. The four subtypes exhibited different molecular, cellular, and clinical characteristics. Consistently, the G3 subtype with better survival had lower expression levels of UBE2T than the G2 and G4 subtypes. G3 subtype patients predominated in clinical stage I and M0 subgroups, while the fraction of G2 and G2 subtypes increased with advanced stage and metastatic status. The subtypes were also associated with tumor biomarkers such as NSE, KRT19, and CA125. Overall, the survival status predicted by UrCCGs agrees with the traditional TNM stage, tumor metastasis status, and tumor biomarkers.Regarding the molecular and cellular characteristics, compared with the G3 group, G2 and G4 groups had higher enrichment scores on tumor-related signaling pathways, including cell cycle, regulation of autophagy, mismatch repair, MTORC1 signaling, MYC-targets V2, unfolded protein response, nucleotide expression repair, PI3K/AKT/mTOR, reactive oxygen species pathway, but lower scores on B cell receptor pathway and p53 pathway. EMT played an essential role in tumor metastasis . With a We analyzed the relationship between different subtypes and immunity. Intriguingly, tumors with G3 and G4 subtypes had higher levels of immune cell infiltration than tumors with G2 subtype. First, ESTIMATE analysis indicated that patients with G3 and G4 subtypes had higher stromal-score and immune-score but lower tumor purity than G2 type. The ssGSEA was adopted to calculate stromal score and immune score for each tumor sample with \u201cstromal gene signature\u201d and \u201cimmune gene signature,\u201d respectively . Second,Moreover, with a published algorithm , we compIn conclusion, UBE2T is a critical oncogene. Moreover, UBE2T-related cell cycle genes could separate the LUAD cohort into four subgroups with significant differences in survival, immune cell infiltration, and immunotherapy sensitivity. Our findings demonstrated that UBE2T could be a therapeutic target, as well as a predictor of survival and immunotherapy in cancer treatment.https://portal.gdc.cancer.gov/. Accessed 20 Nov 2021.1. The TCGA database. https://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/. Accessed 11 Nov 2021.2. The Gencode (GENCODE.v32). http://gepia.cancer-pku.cn/. Accessed 15 Nov 2021.3. The Gene Expression Profiling Interactive Analysis. https://www.genome.gov/Funded-Programs-Projects/Genotype-Tissue-Expression-Project. Accessed 11 Sep 2021.4. The Genotype-Tissue Expression database. http://ualcan.path.uab.edu/analysis-prot.html. Accessed 10 Nov 2021.5. The UALCAN portal. https://www.proteinatlas.org/. Accessed 23 Oct 2021.6. The Human Protein Atlas. http://biocc.hrbmu.edu.cn/CancerSEA/. Accessed 5 Aug 2021.7. The Cancer Single-cell State Atlas. http://timer.comp-genomics.org/. Accessed 27 Nov 2021.8. The tumor immune estimation resource database. https://dcc.icgc.org/. Accessed 7 Nov 2021.9. The ICGC cohort. https://tcia.at/home. Accessed 21 May 2021.10. The Cancer Immunome Atlas. Additional file 1: Figure S1. The flowchart of the study.Additional file 2: Figure S2. Association between UBE2T gene expression and overall survival (OS) of 33 different types of tumors in TCGA database. A-K. Significant association between UBE2T and OS of ACC (A), BRCA (B), KIRC (C), KIRP (D), LGG (E), LIHC (F), LUAD (G), MESO (H), OV (I), STAD (J), and THYM (K).Additional file 3: Figure S3. Association between UBE2T gene expression and disease-specific survival (DSS) of 33 different types of tumors in TCGA database. A-J. The significant association between UBE2T and DSS of ACC (A), COAD (B), KICH (C), KIRC (D), KIRP (E), LGG (F), LUAD (G), MESO (H), OV (I), and PCPG (J).Additional file 4: Figure S4. Association between UBE2T gene expression and disease free interval (DFI) of 33 different types of tumors in TCGA database. A-J. Significant association between UBE2T and DFI of ACC (A), BRCA (B), KIRP (C), LIHC (D), LUAD (E), MESO (F), PAAD (G), PRAD (H), STAD (I), and THCA (J).Additional file 5: Figure S5. Association between UBE2T gene expression and progression-free interval (PFI) of 33 different types of tumors in TCGA database. A-K. The significant association between UBE2T and OS of ACC (A), KICH (B), KIRC (C), KIRP (D), LGG (E), LIHC (F), LUAD (G), MESO (H), PARD (I), STAD (J), and UVM (K).Additional file 6: Figure S6. Validation of discrepancies in tumor immune environment among the four UrCCG subtypes in ICGC-LUAD cohorts. A-C. Differences in the stromal score (A), immune score (B), and tumor purity (C) . D. Heatmap of 28 types of infiltrating immune cells. E. The fractions of 28 infiltrating immune cells were compared among G1-G4 subtypes.Additional file 7. Detailed methods for the construction of risk signature with UrCCGs, discovery and validation of the subtypes by the UBE2T-related cell cycle genes, and investigation of different tumor-associated cellular and molecular states and immunotherapy sensitivity among the four UrCCG subtypes."}
+{"text": "Staphylococcus aureus NCTC 6571-UB, a strain that was derived from S. aureus NCTC 6571. This strain was selected for sequencing in order to provide information on the genome dynamics and the acquired resistance genes for penicillin G, trimethoprim, and sulfamethoxazole resistance.We report the draft genome sequence of the laboratory strain  Staphylococcus aureus NCTC 6571 Oxford strain is a reference strain for penicillin sensitivity bioassays that was first deposited in the National Collection of Type Cultures (NCTC) by N. G. Heatley in 1943 .on assay  on Oxoidon assay . Therefode novo assembled using SPAdes v3.7 (The genomic DNA (gDNA) of NCTC 6571-UB was extracted from a 37\u00b0C overnight TS broth culture. Cells were lysed in Tris buffer (10 mM Tris-HCl pH 8.0) containing lysozyme. RNase A (0.1\u2009mg/mL), proteinase K (0.1\u2009mg/mL), and SDS (0.5% [vol/vol]) were subsequently added. gDNA was purified using SPRI beads  and sequenced by MicrobesNG . Multiple gDNA libraries were prepared using the Nextera XT library preparation kit , quantified using the Kapa Biosystems library quantification kit for Illumina, pooled, and sequenced with an Illumina NovaSeq sequencer (250-bp paired-end read setting). The raw data were quality filtered using Trimmomatic v0.36 , with a total of 2,661 coding DNA sequences, 58 tRNA genes, and 9 rRNA genes. The average coverage of the draft genome was 30\u00d7, and BUSCO analysis revealed 99.8% completeness. NCTC 6571-UB shared 99.86% 16S rRNA gene sequence similarity with the type strain S. aureus subsp. aureus DSM 20231 GenBank accession number AMYL01000007, but their genomes exhibited 97.5% average nucleotide identity (ANI) (in silico DNA-DNA hybridization (DDH) (accessed at https://ggdc.dsmz.de/ggdc.php#) .The sequencing resulted in 1,805,525 raw reads. The assembled draft genome of NCTC 6571-UB had a total length of 2,809,965\u2009bp, with a G+C content of 32.7%, and it consisted of 65 contigs (ty (ANI)  and 76.8NCTC 6571-UB harbored a prophage-associated metallo-\u03b2-lactamase superfamily domain protein (locus tag M3M53_RS02030), which could confer penicillin resistance . ComparePRJNA835436, with BioSample and Sequence Read Archive (SRA) accession numbers SAMN28102073 and SRR19138527, respectively. The whole-genome sequence is available in GenBank under the accession number JAMFMC000000000.1.The whole-genome sequencing project was deposited in GenBank under the BioProject accession number"}
+{"text": "VEGFA) and CXC chemokines have been shown to play vital roles in angiogenesis. Exploring the expression level, gene regulatory network, prognostic value, and target prediction of the CXC chemokine-VEGFA network in colon adenocarcinoma (COAD) is crucial from the perspective of tumor angiogenesis.  Tumor angiogenesis plays a vital role in tumorigenesis, proliferation, and metastasis. Recently, vascular endothelial growth factor A , TRRUST (version 2), LinkedOmics, and Metascape). CXCL1/2/3/5/6/8/11/16/17 and VEGFA were markedly overexpressed, while CXCL12/13/14 were underexpressed in patients with COAD. Moreover, genetic alterations in the CXC chemokine-VEGFA network found at varying rates in patients with COAD were as follows: CXCL1/2/17 (2.1%), CXCL3/16 (2.6%), CXCL5/14 (2.4%), CXCL6 (3%), CXCL8 (0.8%), CXCL11/13 (1.9%), CXCL12 (0.6%), and VEGFA (1.3%). Promoter methylation of CXCL1/2/3/11/13/17 was considerably lower in patients with COAD, whereas methylation of CXCL5/6/12/14 and VEGFA was considerably higher. Furthermore, CXCL9/10/11 and VEGFA expression was notably correlated with the pathological stages of COAD. In addition, patients with COAD with high CXCL8/11/14 or low VEGFA expression levels survived longer than patients with dissimilar expression levels. CXC chemokines and VEGFA form a complex regulatory network through coexpression, colocalization, and genetic interactions. Moreover, many transcription factor targets of the CXC chemokine-VEGFA network in patients with COAD were identified: RELA, NFKB1, ZFP36, XBP1, HDAC2, SP1, ATF4, EP300, BRCA1, ESR1, HIF1A, EGR1, STAT3, and JUN. We further identified the top three miRNAs involved in regulating each CXC chemokine within the network: miR-518C, miR-369-3P, and miR-448 regulated CXCL1; miR-518C, miR-218, and miR-493 regulated CXCL2; miR-448, miR-369-3P, and miR-221 regulated CXCL3; miR-423 regulated CXCL13; miR-378, miR-381, and miR-210 regulated CXCL14; miR-369-3P, miR-382, and miR-208 regulated CXCL17; miR-486 and miR-199A regulated VEGFA. Furthermore, the CXC chemokine-VEGFA network in patients with COAD was notably associated with immune infiltration.  Our results showed that  This study revealed that the CXC chemokine-VEGFA network might act as a prognostic biomarker for patients with COAD. Moreover, our study provides new therapeutic targets for COAD, serving as a reference for further research in the future. Colon cancer is a common malignant tumor of the digestive tract. The incidence and mortality of colon adenocarcinoma (COAD) are the third highest of all cancer types . Since tVEGFA) is a vital factor that plays an essential role in tumor angiogenesis and development [VEGFA inhibitor, has been used to treat advanced renal cell carcinoma. However, the side effects of sunitinib can be quite severe and include kidney and cardiovascular damage [VEGFA are heavily regulated during tumor angiogenesis. CXCL12 can promote a malignant phenotype by promoting the clonal growth of colorectal cancer cells and regulating the expression of VEGF and ICAM-1 [Chemokines are a family of small heparin-binding proteins 8\u201310\u2009kDa in size. Four subgroups exist within the chemokine family . The CXC subgroup has been shown to play a crucial role in angiogenesis in physiological and pathological settings . Recentlr damage . CXC cheVEGFA expression and the development and prognosis of COAD, as well as to provide new insights into targeted therapies for patients with COAD.Multiple online databases were used to explore the expression level, gene regulation network, prognostic value, and regulation targets of the CXC chemokine-VEGFA network in patients with COAD from an angiogenic perspective in this study. In addition, we aimed to identify the relationship between CXC chemokine and http://ualcan.path.uab.edu/analysis.html) is a free online database that provides analysis based on The Cancer Genome Atlas (TCGA) and MET500 cohort data [VEGFA, (2) dataset: COAD, and (3) threshold setting conditions: P value cutoff = 0.05. A Student's t-test was used for the comparative analysis [UALCAN , an open-access resource, provides analyses of specific human genes and proteins [VEGFA, (2) section: tissue and pathology, (3) tissue: colon and COAD, and (4) picture of tissue types: normal colon tissue and COAD. Data were obtained on February 14, 2022.The Human Protein Atlas (proteins . Screenihttp://gepia.cancer-pku.cn/index.html) is an analysis tool that delivers RNA sequencing expression data from 9,736 cancerous and 8,587 noncancerous samples [VEGFA), dataset (COAD), and threshold conditions  were set as screening criteria. The expression of CXC chemokines and VEGFA, as well as the pathological stage of COAD, was analyzed using a Student's t-test. The prognosis of patients with COAD was analyzed using the Kaplan\u2013Meier curve [GEPIA  is a free online database for visualizing, studying, and analyzing cancer genomic data [z-score threshold of \u00b12.0 was used to calculate mRNA expression z-scores for all samples (log RNA Seq V2 RSEM). CXC chemokines and VEGFA were the chosen genes [cBioPortal (mic data . The anaen genes \u201316. Datahttps://string-db.org/cgi/input.pl) is a free online database that helps researchers analyze all publicly available sources of protein\u2013protein interaction (PPI) data [Homo sapiens [STRING (PI) data . We crea sapiens , 15. Dathttp://www.genemania.org) is a free online database that creates PPI networks and analyzes gene function [VEGFA [GeneMANIA (function . The intn [VEGFA \u201316. Datahttps://metascape.org) is a free online gene function analysis tool that assists users in using current common bioinformatics analysis approaches to batch gene and protein analysis to predict function [Metascape (function . We condfunction \u201316. Datahttps://www.grnpedia.org/trrust/) is a free online database for human transcriptional regulatory networks [VEGFA) were chosen in this study [TRRUST (networks . We sougis study \u201316. Datahttp://www.linkedomics.org/) is a free database that provides methods for analyzing and comparing cancer multiomics data [VEGFA), and target dataset (RNA-seq) were chosen in this study [LinkedOmics (ics data . The \u201cLiis study \u201316. Datahttps://cistrome.shinyapps.io/timer/) is a free online platform for systematically analyzing tumor-infiltrating immune cells [TIMER (ne cells . The \u201cGene cells \u201316. DataCXCL1/2/3/5/6/8/11/16/17 and VEGFA were remarkably upregulated in (1) sex , (2) pathological stage (stage 1\u20134), and (3) sample type (COAD) (P < 0.05)  sex , (2) pathological stage (stage 1\u20134), and (3) sample type (COAD) (P < 0.05)  sex , (2) pathological stage (stage 2), and (3) sample type (COAD) (P < 0.05)  (CXCL8/11/14 expression were higher (P \u2264 0.05)   . Subsequ Figures  or when  < 0.05) .VEGFA was altered by 1.3% in COAD patients , CXCL3/16 (2.6%), CXCL5/14 (2.4%), CXCL6 (3%), CXCL8 (0.8%), CXCL11/13 (1.9%), and CXCL12 (0.6%), were found  signaling pathway . In addition, CXCL8 and VEGFA were found to be regulated by ZFP36 ring finger protein (ZFP36), X-box-binding protein 1 (XBP1), histone deacetylase 2 (HDAC2), activating transcription factor 4 (ATF4), E1A-binding protein p300 (EP300), early growth response 1 (EGR1), signal transducer and activator of transcription 3 (STAT3), and Jun proto-oncogene (JUN) (P < 0.01). Furthermore, CXCL1/5/14 and VEGFA were found to be regulated by Sp1 transcription factor (SP1) (P < 0.001). Breast cancer 1 (BRCA1) was the key transcription factor involved with CXCL1 and VEGFA in COAD patients (P < 0.01). Finally, estrogen receptor 1 (ESR1) and hypoxia-inducible factor 1 alpha subunit (HIF1A) regulated the functions of CXCL12 and VEGFA (P < 0.01).Potential transcription factors involved with the CXC chemokine-VEGFA network in COAD patients were identified . v-rel rCXCL1 were miR-518C, miR-369-3P, and miR-44. In addition, miR-518C, miR-218, and miR-493 were identified as potential miRNA targets that regulate CXCL2. Furthermore, we observed that CXCL3 was regulated by miR-448, miR-369-3P, and miR-221. miRNA target of CXCL13 is miR-423. Moreover, miR-378, miR-381, and miR-210 were identified as potential miRNA targets that regulate CXCL14. CXCL17 is regulated by miR-369-3P, miR-382, and miR-208. Furthermore, our results showed that miR-486 and miR-199A are potential miRNA targets that regulate VEGFA.The top three miRNA targets of the CXC chemokine-VEGFA network were obtained . The miRCXCL1/2/3/5/6/8/11/12/13/14/16/17 and VEGFA   = 0.8921, P = 4.226e\u2013132)    , CXCL2 (304e\u201390) , and ZC3882e\u201347) .CXCL2 expression, respectively                                      , CXCL1 (304e\u201390) , and ZC3735e\u201346) . Furtherectively . Among t Figures . Express26e\u2013132) , CXCL2 (01e\u2013119) , and ZC3446e\u201351) . Our resectively . Among t884e\u201368) , IL8 (PC632e\u201363) , and MMP269e\u201362) . Our resectively . Among t Figures . CXCL6 e689e\u201359) , MMP3 (P921e\u201355) , and IL8935e\u201353) . In addiectively . Among t Figures . CXCL8 e939e\u201376) , IL1B (P.25e\u201373) , and OSM368e\u201372) . Furtherectively . Among t Figures . CXCL11 99e\u2013101) , UBD (PC935e\u201362) , and IDO137e\u201360) . Moreoveectively . Among t Figures . CXCL12 835e\u201387) , SLIT3 (915e\u201386) , and SHE928e\u201384) . Our resectively . Among t Figures . CXCL13 598e\u201389) , SH2D1A 229e\u201380) , and SIR508e\u201380) . In addiectively . Among t Figures . CXCL14 .24e\u201358) , TNFSF11643e\u201341) , and COL338e\u201341) . Furtherectively . Among t Figures . CXCL16 175e\u201383) , FLII (P248e\u201340) , and NDE486e\u201339) . We alsoectively . Among t Figures . CXCL17 148e\u201321) , GPR110 333e\u201319) , and SEM753e\u201316) . Finallyectively . Among t Figures . VEGFA e639e\u201335) , CCNL1 (411e\u201330) , and CRE606e\u201327) .CXCL1 expression in COAD patients was positively associated with CD8+ T cell infiltration, neutrophils, and dendritic cells (P < 0.05) (CXCL1 expression (P < 0.01) (CXCL2 and CXCL3 (P < 0.001)     (CXCL17 expression (P < 0.001) (VEGFA expression (P < 0.01) ( < 0.05) . However < 0.01) . In addi Figures . However Figures . Further Figures . B cells Figures . The exp < 0.05) . B cells< 0.001) . CD4+ T  < 0.01) .VEGFA and CXC chemokines as important participants in angiogenesis, particularly tumor angiogenesis [VEGFA have been studied in a range of tumor types; however, findings are contradictory with regard to colonic adenocarcinomas [Tumor angiogenesis plays a vital role in tumorigenesis, proliferation, and metastasis. In recent years, studies have identified ogenesis , 26\u201328. rcinomas , 30. ThiCXCL1/2/3/5/6/8/11/16/17 and VEGFA was upregulated in patients with COAD compared with that in individuals without COAD. Patients with COAD also showed downregulated CXCL12/13/14 expression. The results were similar to those reported in a previous study in patients with COAD [CXCL5/6/12/14 and VEGFA were higher in patients with COAD than those in healthy individuals. Conversely, the promoter methylation levels of CXCL1/2/3/11/13/17 were lower in patients with COAD. Thus, we hypothesized that genetic methylation and alteration within the CXC chemokine-VEGFA network might be the leading cause of abnormal gene expression levels in patients with COAD.In this study, we also examined the potential correlation between pathological stage and differential expression of COAD. The expression of ith COAD  and contith COAD . This maCXCL9/10/11 and VEGFA expression and the pathological stage of COAD. Furthermore, the survival of patients with COAD was higher with low VEGFA or high CXCL8/11/14 expression levels. Therefore, the expression levels of CXCL8/11/14 and VEGFA may be potential prognostic indicators for COAD. CXCL8/11/14 and VEGFA promote tumor angiogenesis in different ways [We also observed a notable correlation between the ent ways \u201333. ThusVEGFA may promote cancer progression, and this could be through a potential interaction network. Genes in the network were mainly involved in cytokine receptor binding, chemokine and cytokine activity, leukocyte chemotaxis, and migration, all of which are closely related to angiogenesis. For instance, IL-8 (CXCL8) promotes tumor angiogenesis by binding to CXCR1 and CXCR2 receptors [The potential functions and interactions of the CXC chemokine-VEGFA network were further explored in this study. They were found to be complex and tightly connected. CXC chemokines and eceptors . In addieceptors . Collect\u03baB signaling pathway were highly involved in the CXC chemokine-VEGFA network in COAD patients, all of which are highly related to tumor angiogenesis [Furthermore, the functions of the CXC chemokine-VEGFA network in patients with COAD were mainly related to chemokine activity, cytokine activity, and growth factor activity, as demonstrated by GO enrichment analysis, all of which are closely related to tumor angiogenesis. More studies are needed to confirm the mechanism by which this happens. In this study, we further found through KEGG pathway analysis that the cytokine\u2013cytokine receptor interaction signaling pathway, IL-17 signaling pathway, and NF-ogenesis , 37. TheVEGFA. Studies have shown that RELA, NFKB1, HDAC2, SP1, ATF4, EP300, BRCA1, ESR1, HIF1A, EGR1, STAT3, and JUN regulate tumor angiogenesis, thus affecting tumor growth and prognosis [Mutated or altered transcription factors represent a unique class of drug targets that mediate aberrant gene expression, and the development of corresponding targeting drugs may impact future cancer treatments. Thus, the targets and regulators of the CXC chemokine-VEGFA network in COAD patients were further analyzed. The transcription factor targets of the CXC chemokine-VEGFA network in patients with COAD were identified. RELA, NFKB1, ZFP36, XBP1, HDAC2, SP1, ATF4, EP300, BRCA1, ESR1, HIF1A, EGR1, STAT3, and JUN were deemed crucial regulatory factors. Our results showed that these factors have potential functions in regulating tumor angiogenesis by targeting rognosis , 38\u201348. rognosis \u201352. In sVEGFA. Some of the genes with the highest correlation  were positively associated with tumor angiogenesis [VEGFA-related regulatory targets may serve as a viable therapeutic oncology approach.The correlation between CXC chemokine-VEGFA network expression and differentially expressed genes in COAD patients was explored in this study. We found that in patients with COAD, approximately 20,000 genes were negatively or positively correlated with CXC chemokine-VEGFA network expression. From these, we screened for genes with the highest correlation with CXC chemokines and ogenesis , 54. RegIn this study, we determined the expression levels and gene regulatory network of the CXC chemokine-VEGFA network, which plays a vital role in angiogenesis in COAD. We also identified new prognostic biomarkers and therapeutic targets. These findings provide insight into the study and treatment of COAD."}
+{"text": "Ceftazidime-avibactam (caz-avi), a novel \u03b2-lactam/\u03b2-lactamase inhibitor, is commonly utilized for carbapenem-resistant gram-negative infections (CR-GNI). However, the benefits vs risks of combining caz-avi with other agents are unclear.In this retrospective cohort study, inpatients with CR-GNI treated with caz-avi were identified at 9 U.S. hospitals. The impact of caz-avi monotherapy (MT) or combination therapy  was studied using logistic regression, controlling for baseline patient and hospital factors. The primary outcome was in-hospital mortality or discharge to hospice (death), and secondary outcomes were length of stay (LOS), resolution of infectious signs and symptoms , 90-day recurrent infection and future caz-avi\u2013resistant organism. An adjusted odds ratio (aOR) with 95% confidence interval (CI) was used to assess the primary and secondary outcomes.Klebsiella spp. (44.6%) followed by Pseudomonas aeruginosa (27.7%) (table 2). Concomitant gram-negative agents are shown in table 3. Overall, 92 (28.1%) patients died and CT (vs MT) displayed similar adjusted mortality risk  and LOS . CT (vs MT) was associated with greater odds of clinical response (aOR: 2.25 [95%CI:1.15-4.41]). Among survivors, similar rates of 90-day recurrent infection (50/154 (32.5%) were observed in CT vs 18/82 (22.0%) in MT group (p=0.09) and 5 (2.19%) patients had future infection with a caz-avi\u2013resistant pathogen (3 in CT and 2 in MT group).328/499 (65.7%) patients received caz-avi as targeted therapy for a CR-GNI. Overall patients treated with MT and CT were similar at baseline and had comparable baseline demographics although patients treated with CT were more likely to be in the ICU and receive a concomitant empiric in vitro-concordant antibiotic (table 1). The most common organism was Compared to patients with CR-GNI treated with caz-avi alone, those who received CT including caz-avy had similar survival and LOS but higher clinical response. The role of CT in the era of novel antibiotics warrants additional study.Helen W. Boucher, MD, American Society of Microbiology: Honoraria|Elsevier: Honoraria|Sanford Guide: Honoraria."}
+{"text": "Aspergillus fumigatus (AFM) is mainly associated with mutations in cyp51A and its promoter region or its homologue cyp51B. We evaluated the in vitro activity of isavuconazole (ISC), itraconazole (ITC), posaconazole (PSC), and voriconazole (VRC) against 660 AFM collected during 2017\u20132020.Azole resistance in cyp51 genes using whole genome sequencing.Isolates from Europe (EU), North America (NA), Latin America (LA), and Asia-Pacific (APAC) were identified by MALDI-TOF and/or sequencing and tested by CLSI broth microdilution. CLSI epidemiological cut-off values (ECV) were applied. A PSC ECV of 0.5 mg/L was used. Non-wildtype (NWT) isolates to azoles were screened for alterations in the cyp51 genes. Of these AFM, 29/32 (90.1%) were NWT to ITC, 25/32 (78.1%) NWT to ISC, 17/32 (53.1%) NWT to VRC, and 11/32 (34.4%) NWT to PSC. The most frequent alteration was CYP51A TR34/L98H, carried by 14 EU isolates, all NWT to ISC and ITC. Four isolates from NA carried the alteration I242V in CYP51A . One NA isolate carried the CYP51A alteration G448S  and one carried A9T (NWT to ISC). A single isolate from APAC carried a CYP51A G138C alteration and was NWT to all 4 triazoles. Multiple alterations in CYP51A were detected in 5 isolates: 4/5 NWT to ISC or ITC, 1/5 NWT to VRC, all WT to PSC. Alterations in CYP51B were noted in 7 isolates; 6/7 carried Q42L, 3 from NA , 2 from APAC , and 1 from EU . Among 34 NWT isolates without cyp51 alterations, 32.4% were WT to ISC, 47.1% WT to ITC, 85.3% WT to VRC, and 82.4% WT to PSC.Azoles had similar activities against 660 AFM isolates. Overall, AFM displayed WT MIC values to ISC (92.7%), ITC (92.9%), PSC (97.3%), and VRC (96.7%). Only 66 isolates (10.0%) were NWT to 1 or more of the azoles, and 32 harbored one or more alterations in the cyp51 alterations were detected in 32/66 NWT isolates. Only EU isolates harboured the environmental alteration TR34/L98H that was associated with a NWT phenotype to ISC and ITC. Alterations in AFM cyp51 can have variable effects on the in vitro activity of the azoles that are best delineated by testing all triazoles.The majority of AFM were WT to azoles. Ten different Michael A. Pfaller, MD, Pfizer: Grant/Research Support Cecilia G. Carvalhaes, MD, PhD, AbbVie: Grant/Research Support|Cidara: Grant/Research Support|Melinta: Grant/Research Support|Pfizer: Grant/Research Support Lalitagauri M. Deshpande, PhD, Melinta: Grant/Research Support|Pfizer: Grant/Research Support Paul Rhomberg, BS, MT(ASCP), Cidara: Grant/Research Support|Pfizer: Grant/Research Support Paul Rhomberg, BS, MT(ASCP), Cidara: Grant/Research Support|Pfizer: Grant/Research Support."}
+{"text": "Lenvatinib has shown promising efficacy in targeted therapies that have been tested to treat anaplastic thyroid carcinoma (ATC) in both preclinical and clinical studies. The aim of this study was to evaluate the efficacy and safety of lenvatinib in the treatment of patients with ATC.PubMed, the Cochrane Library, Embase, and ClinicalTrials.gov were searched for potential eligible studies from inception to February 1, 2022. The outcomes included partial response (PR), stable disease (SD), disease control rate (DCR), median progression-free survival (mPFS), and median overall survival (mOS). Effect sizes for all pooled results were presented with 95% CIs with upper and lower limit.Ten studies met the inclusion criteria. The aggregated results showed that the pooled PR, SD, and DCR were 15.0%, 42.0%, and 63.0%, respectively. The pooled mPFS and mOS were 3.16 (2.18\u20135.60) months and 3.16 (2.17\u20135.64) months, respectively. Furthermore, PFS rate at 3 months (PFSR-3m), PFSR-6m, PFSR-9m, PFSR-12m, and PFSR-15m were 52.0%, 22.5%, 13.9%, 8.4%, and 2.5%, respectively. Meanwhile, the 3-month OS rate (OSR-3m), OSR-6m, OSR-9m, OSR-12m, and OSR-15m were 64.0%, 39.3%, 29.7%, 18.9%, and 14.2%, respectively. The most common adverse events (AEs) of lenvatinib were hypertension (56.6%), proteinuria (32.6%), and fatigue (32%).This meta-analysis showed that lenvatinib has meaningful antitumor activity, but limited clinical efficacy in ATC.https://www.crd.york.ac.uk/PROSPERO/], identifier [CRD42022308624].PROSPERO [ Anaplastic thyroid carcinoma (ATC), a malignancy derived from undifferentiated thyroid follicular cells , accountLenvatinib is a multi-target antiangiogenetic broad-spectrum tyrosine kinase inhibitor (TKI) that can inhibit various signal receptors  \u201312. In aWe have registered our protocol on PROSPERO (registration number: CRD42022308624). This meta-analysis followed the Preferred Reporting Items for Systematic Review and Meta-Analysis (PRISMA) statement . The PRIPubMed, the Cochrane Library, Embase, and ClinicalTrials.gov were searched for potential eligible studies. The search was performed from inception to February 1, 2022. The search keywords were \u201cthyroid carcinoma, anaplastic\u201d and \u201clenvatinib\u201d and the search strategy in PubMed was as follows: Thyroid Carcinoma, Anaplastic [Mesh] OR Anaplastic Thyroid Carcinoma [Title/Abstract] OR Anaplastic Thyroid Carcinomas [Title/Abstract] OR Carcinoma, Anaplastic Thyroid [Title/Abstract] OR Carcinomas, Anaplastic Thyroid [Title/Abstract] OR Thyroid Carcinomas, Anaplastic [Title/Abstract] OR Thyroid Cancer, Anaplastic [Title/Abstract] OR Anaplastic Thyroid Cancer [Title/Abstract] OR Anaplastic Thyroid Cancers [Title/Abstract] OR Cancer, Anaplastic Thyroid [Title/Abstract] OR Cancers, Anaplastic Thyroid [Title/Abstract] OR Thyroid Cancers, Anaplastic [Title/Abstract] AND Lenvatinib [Mesh] OR 4-(3-chloro-4-((cyclopropylaminocarbonyl)amino)phenoxy)-7-methoxy-6-quinolinecarboxamide [Title/Abstract] OR N-(4-((6-carbamoyl-7-methoxyquinolin-4-yl)oxy)-2-chlorophenyl)-N\u2019-cyclopropylurea [Title/Abstract] OR 4-(3-chloro-4-(N\u2019-cyclopropylureido)phenoxy)-7-methoxyquinoline-6-carboxamide [Title/Abstract] OR lenvatinib mesylate [Title/Abstract])) OR (E7080 mesylate [Title/Abstract] OR monomethanesulfonate [Title/Abstract] OR lenvatinib mesylate [Title/Abstract] OR lenvatinib methanesulfonate [Title/Abstract] OR Lenvima [Title/Abstract] OR E-7080 mesylate [Title/Abstract] OR E 7080 [Title/Abstract] OR 4-(3-chloro-4-(((cyclopropylamino)carbonyl)amino)phenoxy)-7-hydroxy-6-quinolinecarboxamide [Title/Abstract] OR E-7080 [Title/Abstract] OR ER-203492-00 [Title/Abstract] OR E7080 [Title/Abstract] OR lenvatinib metabolite M2 [Title/Abstract]. No language, region, ethnicity, age, or payment restrictions were imposed during the search process.Inclusion criteria were as follows : studiesMethodological index for non-randomized studies (MINORS) evaluates single-arm studies . JBI CriTwo investigators independently made study selection. If there were any differences between them, the third author would discuss with them together. Information on the following characteristics of included studies was recorded: authors, study type, sample size, age, criteria for tumor response , adverse events (AEs), and reported endpoints.I2 statistic. When I2 \u2264 50%, use the fixed-effects model; otherwise, use the random-effects model. For pooled results with high heterogeneity, the sensitivity analysis was performed by excluding each study individually. Begg\u2019s test, Egger\u2019s test, and the trim-and-fill method were used to assess publication bias. p < 0.05 was considered statistically significant.Analysis of pooled PR, SD, and DCR, and of the pooled K-M curves of ATC patients treated with lenvatinib was performed using R version 3.6.3. Effect sizes for all pooled results were presented with 95% CIs with upper and lower limit. Heterogeneity between studies was examined using the Cochrane Q chi-square test and We initially identified 349 studies. Finally, our study included 10 studies, namely, 2 prospective studies , 18 and Two single-arm studies , 18 scorI2 = 59.0%, p < 0.01), and the pooled PR in subgroups was different , while the other subgroups showed a pooled PR of 11% .We extracted efficacy measures from each study which included in this meta-analysis , while the subgroup of the retrospective study showed a pooled SD of 36% , and the subgroup of the prospective study resulted in a pooled SD of 59%   and 59% , respectively. The total pooled DCR was 63%   on the estimated pooled PR and DCR , Tianjin Municipal Science and Technology Project  and Beijing-Tianjin-Hebei Basic Research Cooperation Project (20JCZXJC00120), The Science & Technology Development Fund of Tianjin Education Commission for Higher Education (2021ZD033), Tianjin Medical Key Discipline  Construction Project.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Correction:J ExpClin Cancer Res41, 299 (2022)https://doi.org/10.1186/s13046-022-02501-3Following publication of the original article , author This correction does not change the result, interpretation, and conclusions of the study. The original article has been corrected.Additional file 1: Supplementary Figure S5. Blank and plasma membrane impregnated MB morphology by SEM (25.13 K magnification). Scale bar = 1 \u03bcm."}
+{"text": "Nucleic acid reagents, including plasmid-encoded genes and small interfering RNA (siRNA), are promising tools for validating gene function and for the development of therapeutic agents. Native \u03b2-cyclodextrins (BCDs) have limited efficiency in gene delivery due to their instable complexes with nucleic acid. We hypothesized that cationic BCD nanoparticles could be an efficient carrier for both DNA and siRNA. Tetraethylenepentamine-coated \u03b2-cyclodextrin (TEPA-BCD) nanoparticles were synthesized, characterized, and evaluated for targeted cell delivery of plasmid DNA and siRNA. The cationic TEPA coating provided ideal zeta potential and effective nucleic acid binding ability. When transfecting plasmid encoding green fluorescent protein (GFP) by TEPA-BCD, excellent GFP expression could be achieved in multiple cell lines. In addition, siRNA transfected by TEPA-BCD suppressed target GFP gene expression. We showed that TEPA-BCD internalization was mediated by energy-dependent endocytosis via both clathrin-dependent and caveolin-dependent endocytic pathways. TEPA-BCD nanoparticles provide an effective means of nucleic acid delivery and can act as potential carriers in future pharmaceutical application. Gene expression could be modulated through the use of exogenous nucleic acids. The power of nucleic-acid-based drugs, including DNA molecules and small interfering RNAs (siRNAs), lies in their abilities to specifically enhance or silence genes of interest. The siRNA-based drugs were recently approved by US Federal Drug Administration , markingNonviral gene delivery vectors have been shown to have several advantages, such as ease of synthesis, mass production, versatile modification, and low immunogenicity . There aCationic polysaccharides can bind with siRNAs to form complexes, thereby protecting siRNA from degradation and neutralizing their negative charges . NanoparIn this study, we developed an easy and efficient strategy to generate TEPA-BCD. The 6-hydroxyl groups of BCD were activated with tosyl chloride. After nucleophilic displacement by ethylenediamine (EDA), TEPA-BCD nanoparticles were synthesized by the crosslinking reaction of EDA-BCD and TEPA via glutaraldehyde (GA). The synthesis of TEPA-BCD nanoparticles is demonstrated in \u22121 , 1036 cm\u22121 (C\u2013O vibration). Another two intense peaks around 1036 and 2900 cm\u22121 from C-O and C\u2013H stretching were also found in all BCD derivatives indicating the main BCD structure was maintained after the amine introduction .,45.44,45CPZ blocked clathrin-coated pit formation , and conDevelopment of siRNA delivery systems depends not only on their transfection efficiencies, but also on their gene-silencing effect. Although both Gen and MCD treatments showed no reduction in plasmid DNA transfection efficiency C, the geIn the intravenous delivery, the nanoparticles have to escape the various barriers such as kidney filtration, phagocytosis, and hydrolysis degradation. Sequentially, they should transport across vascular vessels, diffuse through the extracellular matrix, and reach the target cells . The enhIn summary, we developed a cationic BCD-based nanoparticle delivery system and applied it in the fibroblast 3T3 cell line and the epithelial ARPE cell line. Our results demonstrated the multifunctionality critical for internalization and activity of both gene-coded plasmid DNA and siRNA. When the TEPA-BCD/plasmid ratio was 1.71:1, the nanoparticles had a transfection efficiency of 97%, and cells maintained viability around 83%. When the TEPA-BCD/siRNA ratio was 96:1, it knocked down the fluorescent expression by 57%, yet the cell viability remained around 82%. TEPA-modified BCD demonstrated a nanometer size, positive surface charge, minimal cytotoxicity, great electrostatic interaction with nucleic acid, and efficient delivery. In both ARPE cells and 3T3 cells, TEPA-BCD entered cells through clathrin- and caveolae-mediated endocytosis. TEPA-BCD/plasmid DNA mediated high gene expression and TEPA-BCD/siRNA displayed efficient gene silencing. The TEPA-BCD provides a promising platform for delivery of therapeutic nucleic acids in vitro and warrants further development of its potential for in vivo delivery.BCD was a generous gift from Feng-Yuan Biotech . Acetone, methanol, and ethanol were purchased from Echo Chemicals . Green fluorescent protein (GFP)-expressing plasmid was obtained from Takara Bio . Fetal bovine serum (FBS) was purchased from Biological Industries . The siRNA against GFP gene  was purchased from MDBio . DNA and siRNA transfection controls (PolyJet and GenMute) are purchased from SignaGen . TEPA, p-tolylsulfonyl chloride, sodium chloride, and other chemicals were purchased from Sigma-Aldrich . All reagents were used without further purification.1H NMR  of BCD: \u03b4 = 5.69 , 4.84 , 4.44 , 3.62 , 3.33  ppm. 13C NMR  of BCD: \u03b4 = 102.04 (C-1), 82.01 (C-4), 73.51\u201372.51 , 69.63 , 60.37 (C-6) ppm. 1H NMR  of Tosyl-BCD: \u03b4 = 7.12 , 7.45 , 5.70 , 4.80 , 4.50 , 4.32 , 4.21 , 3.48 , 2.43  ppm. 13C NMR  of Tosyl-BCD: \u03b4 = 128.48 , 102.44 (C-1), 82.02 (C-4), 73.55\u201372.50 , 60.41 (C-6), 31.16 (Tosyl-CH3) ppm. 1H NMR  of EDA-BCD: \u03b4 = 5.70 , 4.84 , 4.44 , 3.48 , 2.1  ppm. 13C NMR  of EDA-BCD: \u03b4 = 101.82 (C-1), 81.09 (C-4), 73.04\u201371.78 , 60.25 (C-6), 42.04 (BCD-C-C-N) ppm. 1H NMR  of TEPA-BCD: 5.06 , 4.77 , 4.47 , 3.74 , 3.19 , 2.84 , 2.22  ppm. 13C NMR  of TEPA-BCD: \u03b4 = 102.10 (C-1), 81.24 (C-4), 73.20\u201372.08 , 60.15 (C-6), 45.22 (C-NH), 30.24 (cyclic ether C-C-O) ppm.Mono-6-deoxy-6-p-tolylsulfonyl (tosyl)-BCD and EDA-BCD were synthesized as described previously ,50. BrieE. coli cells. The host E. coli was grown in LB broth with kanamycin (50 \u00b5g/mL) and the plasmid DNA was isolated using Mini Purification Kit . The concentration and quality of nucleic acids were determined using a NanoDrop spectrophotometer  and the Beer\u2013Lambert law with an extinction coefficient of 50 \u03bcg/mL\u22121 cm\u22121 at 260 nm. Isolated plasmid DNA had OD 260/280 ratios of 1.80\u20131.95, indicating the quality was suitable for our application. Polyplexes were formulated by mixing 0.21\u20131.71 \u03bcg (10\u201380 \u03bcL) TEPA-BCD with plasmid DNA (1 \u03bcg) or siRNA  in 100 \u03bcL DMEM medium.We transformed plasmid GFP-C3 into DH-5\u03b1 To visualize morphology of BCD derivatives, diluted samples were adsorbed onto a 100 mesh copper grid and dried overnight, followed by imaging on a transmission electron microscope . The magnification power of TEM is 50,000. The chemical groups of modified BCDs were investigated using Fourier transform infrared spectroscopy  and an NMR spectrometer (Bruker AV III HD). The size and zeta potentials of the TEPA-BCD complexes were measured using ZetaSizer with 633 nm He-Ne Laser  and the folded capillary cell .ARPE cells  and 3T3 cells (a mouse embryonic fibroblast cell line) from the cell bank of BCRC  were used for GFP plasmid delivery experiments. DNA transfection was performed as previously reported . APRE-GF4 cells/well. When reaching 90% confluence, the cells were treated with polyplex and endocytic inhibitor for 3 h. Polyplexes were formulated by mixing 1.28 \u03bcg TEPA-BCD with plasmid DNA (1 \u03bcg) or siRNA (13.3 ng) in 100 \u03bcL DMEM medium. Subsequently, the transfection medium was replaced with fresh DMEM supplemented with 10% FBS and cells were incubated for another 24 h. Imaging was conducted on an automated image analyzer for analysis of fluorescence intensity and cell viability.The cellular uptake mechanisms of polyplex were examined using different endocytic inhibitors: chlorpromazine (30 \u00b5M), genistein (200 \u00b5M), sodium azide (10 mg/mL), methyl-BCD (10 mM), and monensin (3 \u00b5M). The cells were seeded in 48-well plates at 5 \u00d7 10t-test by Analysis Toolpak in Microsoft Excel. Statistical significance is represented as * p < 0.05.All cellular data were performed by two individual experiments. The Zetasizer results were the average from three individual experiments. The mean and standard deviation were calculated using Microsoft Excel. Statistical comparisons are performed using PROC ANOVA followed by the Tukey post hoc test using SAS 9.4 software  and unpaired two-tailed Student"}
+{"text": "In the United Kingdom(1) and internationally(2), help-seeking for domestic violence (DV) and domestic homicides have increased(3) during COVID-19 lockdown periods. Suspension and remote delivery of face-to-face clinical services, continuing healthcare and other support services limits opportunities for DV detection and disclosure.This presentation will summarise changes in DV incidence and help-seeking during COVID-19, their impacts on health and wellbeing, and present guidance for clinicians assessing and supporting survivors.World Health Organisation recommendations to Listen, Inquire, Validate, Enhance safety and Support (\u2019LIVES\u2019) survivors of DV remain the cornerstone of first-line support (4). Urgently-issued guidelines on safeguarding(5) and responding to DV during COVID-19(6) make a range of recommendations for clinicians supporting people experiencing DV.DV is an important social determinant of physical and mental health, with a range of potential fatal and non-fatal consequences. Despite the constraints of healthcare during a pandemic, attention to patients\u2019 risk of DV and its consequences is a crucial part of bio-psycho-social assessment and management planning. References: (1) Kelly, Morgan. Coronavirus: Domestic abuse calls up 25% since lockdown, charity says. 2020. https://www.bbc.co.uk/news/uk-52157620 (2) Graham-Harrison, et al. Lockdowns around the world bring rise in domestic violence. 2020. https://www.theguardian.com/society/2020/mar/28/lockdowns-world-rise-domestic-violence (3) Roesch, et al. Violence against women during covid-19 pandemic restrictions. BMJ 2020;369. (4) WHO. Responding to intimate partner violence and sexual violence against women: WHO clinical and policy guidelines. 2013. https://apps.who.int/iris/bitstream/handle/10665/85240/9789241548595_eng.pdf;jsessionid=E19DCC3CDAB9BE390EE6F8360C6F1D7E?sequence=1 (5) RCGP. COVID-19 and Safeguarding. 2020. https://elearning.rcgp.org.uk/pluginfile.php/149180/mod_resource/content/2/COVID-19%20and%20Safeguarding%20%286%29.pdf (6) IRISi. Guidance for General Practice teams responding to domestic abuse during telephone and video consultations. 2020. https://irisi.org/wp-content/uploads/2020/04/Guidance-for-General-Practice-Covid-19-FINAL.pdfNo significant relationships."}
+{"text": "Acute myeloid leukemia (AML) is a heterogeneous hematologic malignancy with high mortality and poor outcome, especially for elderly/unfit (age \u2265 60 years or unfit patients who are ineligible to receive intensive chemotherapy) with adverse genetic and molecular abnormalities in the newly diagnosed (ND) and refractory/relapsed (R/R) patients.Azacitidine (Aza), a hypomethylating agent, targets epigenetic gene silencing by inhibiting gene expression against malignant phenotypes. The complete remission (CR) rates of Aza combined regimens varied with different drugs, such as, Aza+Midostaurin (20.8%), Aza+Durvalumab 31.3%), Aza+Pracinostat (46%), Aza+Venetoclax (66.4%) in ND AML and Aza+Midostaurin (21.4%), Aza+Nivolumab (22%), Aza+Venetoclax 37.1%) in R/R AML  were enrolled between Jan 2020 and Dec 2021, including 72 ND and 40\u2009R/R Fig. . The 72 2/d on days 1\u20137 subcutaneous) was given in combination with the HAG regimen . Patients who did not achieve CR/CRi (CR with incomplete blood count recovery) following the first cycle could receive a second cycle at the same doses and schedule. The patients who did not reach CR/CRi after the second cycle were withdrawn from the study. Post-remission therapy for enrolled patients, the Aza+HAG regimen was further given 4\u20136 cycles or until the disease progresses, two of whom underwent allo-SCT .Induction therapy consisted of Aza . A higher CR/CRi rate of 79.2% in ND AML compared with 40.0% in R/R AML; and sAML (75.0%) obtained a similar high CR rate as de novo AML (80.4%)  achieved CR/CRi within the median time of 33.5d. Notably, 90.4% (66/73) of CR/CRi was obtained after the 1st cycle of Aza+HAG induction therapy. No statistical difference in CR/CRi rate between Aza+HAG (7-day)  vs Aza+HAG (14-day)  (4%) Fig. . In pati4%) Fig. . These dP\u2009=\u20090.0056; median RFS 5.21\u2009m, P\u2009=\u20090.0051)  . In patients with favorable, intermediate, or poor risk, median OS was not reached, 22.8\u2009m, and 17.3\u2009m  and relapse-free survival (RFS) in ND AML were 22.8\u2009m and not reached, respectively, which were longer than R/R AML  Fig. . De novo48) Fig. . In pati3\u2009m Fig. ; median 3\u2009m Fig. , respectP\u2009=\u20090.016) , respectively . The baseline demographic and characteristics were generally balanced between the two cohorts Table . The CR/16) Fig. . The medely Fig. . We alsoely Fig.  and S2. The most common non-hematological adverse event (AE) was infection (58.0%). Common non-hematological AEs of grade 3 or higher includes: infection (33.9%), hemorrhage (9.82%), fatigue (5.36%), hypokalemia (3.57%), cardiac arrhythmia (1.79%), and fever (1.79%). For the hematological AE, the median duration of neutropenia and thrombocytopenia were 11 d  and 16d , 10 d , and 16 d , 12d  and 17d  for total, 7-day, and 14-day schedule, respectively. No differences were observed between the 7-day and 14-day schedule. The early deaths within 4 weeks of the induction treatment occurred in 1.79%. No patients discontinued the induction therapy due to hematological or non-hematological toxicities Table . These dDNMT3A, TET2, and NPM1 , KIT , IDH1 , NPM1 , ASXL1 , DNMT3A , RUNX1 , FLT3  and TET2   and 87.4% (90/103) of patients had more than one gene mutation. The most frequently mutated genes were PM1 Fig. . We obse14) Fig.  (Table S14) Fig. . We alsoPM1 Fig. . OS and ion Fig. ; whereasype Fig. . These dIDH1 inhibitor Ivosidenib (60.9%) [IDH1-mutant ND AML or HMA\u2009+\u2009Venetoclax (71%) in IDH1/2-mutant AML [IDH1-mutant AML and the IDH1 mutant was dramatically reduced after the first cycle of Aza+HAG, suggesting that the patients with the IDH1 mutation could achieve a remarkable deep and durable remission upon Aza+HAG treatment.Recent studies reported the CR rates of patients treated with the Aza plus  (60.9%)  in IDH1-tant AML . This trIn summary, this trial demonstrated that the Aza+HAG regimen is a cost-effective first-line therapy with high efficacy and well tolerance for elderly/unfit AML. This trial provides a rationale for further expanding the patients for randomized clinical controlled studies and guiding suggestions for the clinical use of this novel combination therapy.Supplemental methods_tables_figures"}
+{"text": "Background and Objectives: To estimate the association between admission functional outcomes and exposure to physiotherapy interventions with mortality rate in intensive care unit (ICU) inpatients with cardiovascular diseases and new coronavirus disease (COVID-19). Materials and Methods: Retrospective cohort including 100 ICU inpatients (mean (standard deviation), age 75 (16) years) split into COVID-19+ or COVID-19\u2212. The association of in-ICU death with admission functional outcomes and physiotherapy interventions was investigated using univariable and multivariable regression models. Results: In total, 42 (42%) patients tested positive for COVID-19. In-ICU mortality rate was 37%, being higher for the COVID-19+ group : 3.15 (1.37\u20137.47), p = 0.008). In-ICU death was associated with lower admission ICU Mobility Scale score (0.81 (0.71\u20130.91), p = 0.001). Restricted mobility (24.90 (6.77\u2013161.94), p < 0.001) and passive kinesiotherapy (30.67 (9.49\u2013139.52), p < 0.001) were associated with in-ICU death, whereas active kinesiotherapy (0.13 (0.05\u20130.32), p < 0.001), standing (0.12 (0.05\u20130.30), p < 0.001), or walking (0.10 (0.03\u20130.27), p < 0.001) were associated with in-ICU discharge. Conclusions: In-ICU mortality was higher for inpatients with cardiovascular diseases who had COVID-19+, were exposed to invasive mechanical ventilation, or presented with low admission mobility scores. Restricted mobility or passive kinesiotherapy were associated with in-ICU death, whereas active mobilizations  were associated with in-ICU discharge in this population. Cardiovascular diseases (CVDs) are the leading cause of death globally, with estimated 31% mortality, representing about 17.9 million deaths every year . The BraThe hazards of hospitalization, particularly for older adults in the intensive care units (ICU), are a longstanding issue that ultimately favors a decline in musculoskeletal function and functional capacity . Higher Retrospective, single-center study. Data were obtained by the principal investigator through information previously contained in electronic medical records, examination reports, and notes of the health professional staff involved in the care of the patients. This study is reported following the REporting of studies Conducted using Observational Routinely-collected health Data (RECORD) statement . MinimumThis study retrospectively analyzed all data from February to November 2020 collected from patients consecutively hospitalized at the ICU of a primary-to-tertiary private hospital located in Curitiba, Paran\u00e1, Brazil.ICU admission criteria comprised at least of the following conditions: hemodynamic ; neurological ; or respiratory . Other causes for ICU admission included postoperative period of high-risk patients, postoperative period of large surgeries, postoperative cardiac surgery, postoperative neurological surgery, postoperative endovascular surgery, need for clinical monitoring, sepsis, septic shock, cardiac arrhythmia, vascular diseases of the heart, or acute renal dysfunction.Patients who had a primary diagnosis of CVD after a complete clinical exam and laboratory testing including laboratory blood tests, electrocardiogram, blood pressure, and/or echocardiography as prescribed, admission assessment by a physiotherapist, and tested for SARS-CoV-2 infection at admission were included. ICU admission was defined as an admission to the hospital\u2019s ICU for >12 h. Re-admissions of patients to the ICU within the study period were excluded from the analysis.All admission data were collected within <24 h of ICU hospitalization at the discretion of the medical staff and covered the required time for swab analysis. Data were collected retrospectively from electronic medical recordings regarding demographics, vital signs, laboratory, gasometry, presence of CVD and comorbidities, and drugs in continuous use. Date of hospital admission and discharge from the ICU or death were collected for computing the total length of ICU stay. The sample was divided into groups COVID-19+ or COVID-19\u2212 based on the ICU admission test, after a nasal and/or nasopharyngeal swab for SARS-CoV-2 by polymerase chain reaction method.Overall muscle strength was assessed by the Medical Research Council (MRC) scale, which uses a 6-point scale of 6 muscle groups bilaterally. Representative scores comprised the sum of points observed for each muscle group bilaterally, ranging from 0 (no muscle activity) to 60  .Mobility was assessed by the ICU Mobility Scale (IMS). The score varies between 0 expressing low mobility (patient who only performs passive exercises in bed) and 10 expressing high mobility  .All patients were exposed to physiotherapy interventions based on the hospital standards of usual care developed according to international and national recommendations ,27,28,292O and subsequent titration of ideal PEEP, provided they were clinically stable. When they needed oxygen therapy, it was performed using a low-flow system . Spontaneous prone was also used for at least 1 h. In the supine position [Ventilatory support was characterized using non-invasive mechanical ventilation, through an orofacial or facial interface connected to the mechanical ventilator in one or two pressure levels ventilation modes, or invasive ventilatory support . Patients diagnosed with acute respiratory distress syndrome used protective strategy ventilatory parameters, which may require alveolar recruitment through the prone position or recruitment through the gradual increase of positive end-expiratory pressure (PEEP) up to 35 cmHposition , the heaMobility activities were categorized as complete bed restriction; passive kinesiotherapy ; active kinesiotherapy ; assisted or active sitting out of bed; standing; and walking.The primary outcome was in-ICU mortality as well as admission functional assessments of MRC and IMS scores. In-ICU mortality was calculated from the admission date and confirmed using electronic medical records.No missing data occurred for exposures, in-ICU mortality, or admission IMS scores. Data were missing for the admission assessment of MRC scores in 18/100 participants due to sedation.p < 0.05 (two-tailed).Statistical analysis was performed in jamovi v. 1.8.1.0 and R project version 4.0.4 with packages after importing the electronic spreadsheet. Missing data in admission measurements were reported and assumed to be missing completely at random and univariate mean imputation was performed. Evidence of statistical significance was considered at Descriptive summaries were reported as mean (standard deviation (SD)) for continuous variables or absolute and relative frequencies (%) for categorical ones. Admission demographic data were compared between COVID-19+ versus COVID-19\u2212 groups using the linear model analysis of variance or Pearson\u2019s Chi-squared test for continuous and dichotomous variables, respectively.Univariable logistic regression analysis was performed to examine the association (odds ratio (OR) with 95% confidence interval (95% CI)) of exposure to physiotherapy intervention (ventilatory support and mobility) with group (COVID-19+ vs. COVID-19\u2212) and in-ICU mortality. Multivariable logistic regression model was fitted to determine independent factors associated with in-ICU mortality; all factors related to exposure to physical therapy were force-entered as a full model. Model fit was evaluated by Akaike information criterion (AIC) and C-statistic.2). The most common underlying CVD was hypertension (91/100), followed by a history of cerebrovascular disease (22/100), coronary artery disease (21/100), heart failure (16/100), and atrial fibrillation. Overall length of ICU stay was 9.7 (15.5) days, with patients with COVID-19+ showing longer ICU stay (14.5 (21.7) vs. 6.2 (6.8), p = 0.007). Between-groups comparisons show that patients with COVID-19+ showed higher admission MRC scores (48.3 (7.9) vs. 43.8 (10.4), p = 0.021) and IMS scores (5.5 (4.0) vs. 3.9 (3.8), p = 0.039). They were also younger (68 (16) vs. 80 (13) years, p < 0.001) and presented at admission with lower leukocytes count (9565 (5473) vs. 13,456 (6444) count/mcL, p = 0.002), lower partial pressure of carbon dioxide (PCO2) (31.7 (6.5) vs. 37.7 (8.5), p < 0.001), and lower bicarbonate (21.9 (4.5) vs. 23.8 (4.9) mEq/L, p = 0.044). Sedation at admission was more frequent in the COVID-19+ group . No statistical evidence of difference was observed between groups for severity of disease based on acute physiology and chronic health evaluation (APACHE II: 30.3 (4.8) vs. 30.2 (4.9), p = 0.917).Overall, the sample was composed of older adults (75 (16) years), balanced between sexes , and with most participants (40/100) classified as overweight (body mass index = 27.1 (5.1) kg/mp = 0.011) or prone position (22.80 (4.19\u2013425.35), p = 0.003), to alveolar recruitment (19.04 (4.97\u2013125.99), p < 0.001), awake prone (5.60 (1.27\u201339.05), p = 0.038), or length of stay (1.07 (1.02\u20131.13), p = 0.008), but not to non-invasive mechanical ventilation or oxygen therapy. Multivariable logistic regression analysis showed a good linear fit (C-statistic = 0.783), with COVID-19 remaining independently associated with exposure to alveolar recruitment (22.34 (3.56\u2013224.91), p = 0.002) or awake prone (13.41 (1.62\u2013228.22), p = 0.032).As related to ventilatory support , patientp = 0.029).As related to mobility during ICU stay , no statp = 0.008). As related to ventilatory support, in-ICU mortality was more likely in patients exposed to invasive mechanical ventilation in either supine (22.71 (8.28\u201370.76), p < 0.001) or prone position (4.74 (1.42\u201318.74), p = 0.016), oxygen therapy (8.75 (2.34\u201357.11), p = 0.005) or alveolar recruitment , p < 0.001). Also, in-ICU death was associated with longer length of stay (1.04 (1.01\u20131.09), p = 0.048)), lower admission IMS score (0.81 (0.71\u20130.91), p = 0.001) but not MRC score (p = 0.055). Multivariable logistic regression analysis showed an excellent linear fit (C-statistic = 0.920), with in-ICU death still independently associated with COVID-19+ , exposure to invasive mechanical ventilation (14.81 (2.97\u201397.70), p = 0.002) and admission IMS score (0.79 (0.63\u20130.96), p = 0.023).p < 0.001), passive kinesiotherapy (30.67 (9.49\u2013139.52), p < 0.001) and longer LOS (1.04 (1.01\u20131.09), p = 0.048) were associated with in-ICU death, whereas active kinesiotherapy (0.13 (0.05\u20130.32), p < 0.001), standing (0.12 (0.05\u20130.30), p < 0.001), or walking (0.10 (0.03\u20130.27), p < 0.001) were associated with in-ICU discharge. Multivariable logistic regression analysis showed an excellent linear fit (C-statistic = 0.947), with in-ICU death still independently associated with COVID-19+ (15.44 (2.80\u2013140.82), p = 0.005), restricted mobility (10.49 (1.74\u201395.85), p = 0.017), and passive kinesiotherapy (17.66 (2.32\u2013326.56), p = 0.017).As related to mobility during ICU stay, two patterns emerged: restricted mobility (24.90 (6.77\u2013161.94), The main findings suggest that in-ICU mortality is higher for inpatients with CVD who had a COVID-19+ test result, were exposed to invasive mechanical ventilation, or presented with low admission IMS scores, whereas restricted mobility or passive kinesiotherapy were associated with in-ICU death, active mobilizations  were associated with in-ICU discharge. Our findings contribute to the global discussion on the acute management of patients with COVID-19 by providing insights about the planning physiotherapy interventions aimed at reducing in-ICU fatality among patients with CVD.Demographic and clinical data from our sample in Brazil corroborate previous studies in other countries on the risk factors for hospitalization and mortality in patients with CVD and COVID-19: mainly older age, overweight, low lymphocyte count, and pre-existing comorbidities, among others ,9,31,32.Clinical algorithms  and consIn-ICU mortality was higher for inpatients with CVD who had a COVID-19+ test result, were exposed to invasive mechanical ventilation, or presented with a lower admission IMS score. Altogether, these characteristics can be understood as a proxy for disease severity. Interestingly, exposure to physiotherapy intervention showed two distinct effects on in-ICU mortality rate, whereas restricted mobility or passive kinesiotherapy were associated with in-ICU death, active mobilizations  were associated with in-ICU discharge. This finding corroborates previous studies showing improved mobility at hospital discharge and a higher probability of discharging home with increased frequency and longer mean duration of physical therapy visits in patients with COVID-19 admitted to acute care hospitals . ConsideThis study has some limitations. Due to the retrospective design, there were missing data for participants regarding admission assessment of functional outcomes. Clinical data at admission were collected within <24 h of ICU hospitalization and hence may differ from pre-admission status that required hospitalization. Moreover, physiotherapy interventions were delivered as per the rehabilitation team\u2019s clinical decision-making process. Whereas such lack of control in experimental factors may have influenced the delivered interventions in each group, such pragmatic approach most likely represents ICU routines in Brazil since they are based on national guidelines. Some CIs returned wide ranges, which suggests a large uncertainty about the effect  and that further information is needed. Nonetheless, assessment of goodness-of-fit of the models (Akaike information criterion and C-statistics) suggest acceptable model validity. The current sample is from a single center during the first \u2018wave\u2019 (February to November 2020) of cases in Brazil  when varIn-ICU mortality is higher for inpatients with CVD who had COVID-19+, were exposed to invasive mechanical ventilation, or presented with low admission mobility scores. The protective effects of routine physiotherapy interventions are highest when patients can perform active rather than passive kinesiotherapy. Patients with COVID-19 who further perform standing and walking activities appear to experience a higher survival effect."}
+{"text": "Tuberculosis (TB) remains a leading infectious cause of death and morbidity globally. Rifampin-resistance or intolerance requires prolonged treatment using less effective, more toxic regimens. Recent trials demonstrated that the all-oral six-month \u201cBPaL\u201d regimen, Bedaquiline, Pretomanid and Linezolid, is 90% effective. In these trials, linezolid-induced hematologic and neurologic toxicity was high using 1200mg daily, whereas lower exposure (dose or duration) reduced toxicity. Therapeutic drug monitoring (TDM) is used by U.S. TB experts to maintain a serum linezolid trough < 2ug/ml, which correlates with reduced toxicity. Since U.S. FDA approval in 2019, BPaL has been widely implemented for the treatment of rifampin-intolerant or resistant TB disease.We evaluated a cohort of patients with TB treated from October, 2019 through April, 2022, describing patient demographics, BPaL treatment dosing, and adverse events. TDM was performed for clinical purposes using liquid chromatography\u2013mass spectrometry to measure serum levels at trough and 2 and 6h post-linezolid dose. Clinical providers adjusted linezolid dose and dosing interval targeting a trough < 2ug/ml and peak of 12\u201326ug/ml.Among 64 BPaL patients, ages were 15\u201383 years, 22 (34.3%) were female, 6 (9.3%) U.S.-born, 4 (6.3%) HIV-infected. 50 (78.1%) had only pulmonary disease, 6 extrapulmonary, and 8 had both; 61 (91.0%) were culture-confirmed. Most (n=62) started linezolid 600mg daily. Linezolid was adjusted for 39 (66.1%) of the 59 patients with TDM; 18 had a trough >2ug/ml, 30 had dosing interval increased to thrice-weekly, and 17 had a dose increase. 52 (81.3%) patients completed BPaL and 12 remain on therapy. One 81-year-old female with diabetes, hypothyroidism, and B12 deficiency discontinued linezolid at 12 weeks for worsened neuropathy (linezolid trough=1.13ug/ml). She completed 26 weeks of bedaquiline/pretomanid and her symptoms returned to baseline.Use of BPaL with clinical and TDM monitoring has transformed treatment of rifampin-resistant or intolerant TB in the U.S. Patients previously sentenced to 18\u201324 months of treatment with 5\u20137 hard-to-tolerate medications and modest efficacy can now complete treatment in 6\u20139 months with little toxicity and exceptional cure rates.All Authors: No reported disclosures."}
+{"text": "Nocardia farcinica is an opportunistic pathogen that causes nocardiosis primarily in patients with compromised immune systems. In this study, we used the genetically tractable organism Caenorhabditis elegans as a model to study the innate immune responses to N. farcinica infection. We found that unlike other pathogenic bacteria such as Pseudomonas aeruginosa and Staphylococcus aureus, N. farcinica failed to kill adult worms. In another words, adult worms exposed to N. farcinica exhibited a normal lifespan, compared with those fed the standard laboratory food bacterium Escherichia coli OP50. Interestingly, deletion of three core genes  in the p38 MAPK/PMK-1 pathway reduced the survival of worm exposure to N. farcinica, highlighting a crucial role of this pathway for C. elegans in resistance to N. farcinica. Furthermore, our results revealed that N. farcinica exposure up-regulated the level of PMK-1 phosphorylation. The activation of PMK-1 promoted nuclear translocation of a transcription factor SKN-1/Nrf2, which in turn mediated N. farcinica infection resistance in C. elegans. Our results provide an excellent example that the integrity of immune system is key aspect for counteract with pathogenesis of N. farcinica. Nocardia farcinica, is an opportunistic pathogen under the genus of Actinomycetes widely distributed in the soil and rotten substances, causing lung infection through inhalation of bacterial particles [N. farcinica, suppress immune responses by inhibiting Notch signal pathway in RAW264.7 macrophages [N. farcinica in mammal. The mechanism of pathogenesis in N. farcinica infection is not fully understood.articles . Since tarticles . Nocardiarticles . Nocardiarticles . A recenCaenorhabditis elegans is a free-living, bacterivorous nematode that lives in the soil, well known as invertebrate without adaptive immune system as well as specific immune cells [C. elegans often encounters a variety of pathogens, thus it exerts a strong selective pressure to evolve and develop an effective innate immune system. C. elegans therefore represents a powerful invertebrate model to study the innate immune responses to nosocomial bacterial pathogens such as Pseudomonas aeruginosa, Staphylococcus aureus, Salmonella enterica, Salmonella typhimurium, and Serratia marcescens [Cryptococcus neoformans [Candida albicans [ne cells . In wildne cells . In the rcescens ,8,9,10, oformans  and Candalbicans .C. elegans have revealed a variety of the evolutionarily conserved signaling pathways in the innate immunity, such as the DAF-2 insulin-like signaling pathway [C. elegans to regulate innate immunity [C. elegans upon P. aeruginosa infection [P. aeruginosa infection [During the last two decade, studies using  pathway , the p38 pathway , and the pathway , the JNK pathway , the pro pathway , the G p pathway , the G p pathway , and the pathway . Of thesimmunity ,21. PMK-immunity . Meanwhinfection . Interesnfection . Under nnfection .N. farcinica in C. elegans. We discovered that the PMK-1/p38 MAPK signaling was required for the survival of worms upon N. farcinica exposure. Further studies indicated that PMK-1/p38 MAPK activated the transcription factor SKN-1, which in turn conferred resistance to N. farcinica infection in C. elegans.In this study, we screened the major signaling pathways that are involved in defense against N. farcinica in nematodes. However, we found that unlike worms exposed to P. aeruginosa, S. aureus, and E. faecium [N. farcinica exhibited a similar lifespan as those fed E. coli OP50, the standard laboratory food . In contrast, pmk-1(km25) mutants exhibited marked decrease in lifespan when exposed to N. farcinica, compared to worms fed on E. coli OP50 and WT (nsy-1(ag3) and sek-1(ag1) mutants exposed to N. farcinica also showed a significant reduction in lifespan, compared to E. coli OP50 diet and WT , sek-1(ag1), and pmk-1(km25) suppressed their mRNA levels , catalase (katG), and superoxide dismutase (sodF) are predicted with high expressivity in N. farcinica, which could defense against reactive oxygen species (ROS) [N. farcinica infection. Thus, ROS are unlikely to be involved in the activation of the p38 MAPK pathway in worms.We then asked if oli OP50 a,b. The osa PA14  and Cutium acnes . A previby PMK-1 . We thuspression c,d. As Fed genes , we thenA levels e. Taken es (ROS) . We thusC. elegans, SKN-1 confers resistance to Gram-negative bacteria P. aeruginosa PA14 and Gram-positive bacteria E. faecalis [N. farcinica infection, we used transgenic worms expressing SKN-1b/c::GFP+rol-6 to detect the nuclear translocation of SKN-1, which is an indicator of its activation [N. farcinica exposure. However, knockdown of nsy-1, sek-1 or pmk-1 by RNAi significantly reduced the nuclear translocation of SKN-1 in worms exposed to N. farcinica (gst-4 (encoding Glutathione S-transferase-4) is a target gene of SKN-1 [Pgst-4::gfp reporter gene to further demonstrate the activation of SKN-1 by N. farcinica infection. We found that the expression level of Pgst-4::gfp was dramatically increased in worms exposed to N. farcinica for 24 hours, The increase is abolished when worms subjected to skn-1, nsy-1, sek-1 or pmk-1 RNAi (skn-1(tm3411), nsy-1(ag3), sek-1(ag1), and pmk-1(km25) suppressed up-regulation of gst-4 expression induced by N. farcinica infection  mutants was comparable to that of the pmk-1(km25) mutant subjected to skn-1 RNAi upon N. farcinica infection  of N. farcinica and E. coli OP50 in the body of worms  worms, skn-1 (RNAi) worms, and control group worms. As black arrows pointing out, genetic inactivation of pmk-1 and skn-1 caused necrosis in the head of worms exposed to N. farcinica, but not to E. coli OP50 , known as key immune factor, mediates resistance to Mycobacterium. spp [N. farcinica in organisms ranging from C. elegans to mammals.In ruginosa ,38 S. au. aureus , Salmonelmonella , Yersinia pestis , and Mycbacteria . The p38bacteria  secreted O157:H7 . In mamm O157:H7 ,44, suggium. spp ,46. ThesP. aeruginosa, E. faecalis [Aeromonas dhakensis [N. farcinica infection. It has been reported that PMK-1 can phosphorylate SKN-1 at Ser-74 and Ser-340 [P. aeruginosa, E. faecalis and N. farcinica infection activates SKN-1 in a p38 MAPK-dependent manner. These results suggest that SKN-1 is one of downstream effectors of p38 MAPK that mediates C. elegans innate immune responses to both Gram-negative and Gram-positive pathogens. The molecular mechanism underlying SKN-1-mediated innate immunity remains unclear. Previously, we and others have demonstrated that autophagy plays an important role in C. elegans defense against a variety of pathogenic bacteria by repairing organismal insults [glp-1(e2141ts) mutants [hyl-1; lagr-1 mutants [N. farcinica infection via promoting autophagy. The opportunistic infections followed immunologic deficiency have mainly been seen in HIV infected patients, revealing that the environmental \u201charmless\u201d microorganisms could be life threaten, such as Penicillium marneffei, known as the third common opportunistic pathogen that causes penicilliosis in HIV patients. Likewise, N. farcinica is an opportunistic pathogen, with the majority of infections occurring in immunocompromised patients. In the current study, WT worms do not have a reduced lifespan when exposed to N. farcinica. These data suggested that like humans, WT worms with an intact innate immune system can resist to N. farcinica infection, but lacking key genes in the PMK-1/p38 MAPK pathway become susceptible to N. farcinica infection. In our studies, we also noticed that lack of p38 MAPK pathway and SKN-1 leads to significant increases accumulation of N. farcinica and obvious enlarged vacuoles in the head of worms. Thus, our finding emphasizes that the integrity of immune system is crucial for defense against N. farcinica infection in C. elegans.As a transcription factor, SKN-1 is involved in resistance to oxidative stress by upregulating a set of the Phase II detoxification genes . SKN-1 afaecalis , Mycobacfaecalis , and Aerhakensis . In the  insults ,20,48. S mutants . Meanwhi mutants . Thus, SN. farcinica to activate the p38 MAPK/PMK-1 pathway?However, our data demonstrate that ROS are not the signal component produced by N. farcinica. Several virulence factors of N. farcinica have been reported when the pathogen infected mammals. For instance, Nfa34810 protein of N. farcinica can stimulate macrophages to produce tumor necrosis factor alpha (TNF-\u03b1) and other immune related factors through the TLR4 pathway [N. farcinica IFM 11523, which isolated from Japanese patient, secretes two pathogenesis factors nocobactin NA-a (compound 1) and nocobactin NA-b (compound 2), which act as on the notch signaling inhibitors [What are the critical signal molecules from  pathway . BesidesC. elegans strains were maintained on NGM medium containing E. coli OP50 under standard condition. Strain used in this study include C. elegans Bristol N2, pmk-1(km25), mpk-1(n2521), let-60(ga89), jnk-1(gk7), hlh-30(tm1978), skn-1(tm3411), nsy-1(ag3), sek-1(ag1), skn-1b/c::gfp+rol-6(su1006), nlp-29p::gfp+col-12p::DsRed, gst-4p::gfp::NLS(pAF15) were kindly provided by the Caenorhabditis Genetics Center (CGC), which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Bristol N2 strain of C. elegans is used as non-genetic modified control in experiments, and labeled as wild-type (WT) in our study [The , mpk-1n21, let-60ur study .E. coli HT115 (DE3) strain carrying vector L4440 express dsRNA corresponding to targeted genes to perform double-stranded (ds)RNA-mediated RNA interference (RNAi). As experimental control, E. coli HT115 carrying empty vector (no dsRNA expression) were used, labelled as EV. All strains were grown overnight in LB medium with 100 \u03bcg/mL ampicillin at 37 \u00b0C. Cultured E. coli strains spread onto NGM plates contained 100 \u03bcg/mL ampicillin and 1 mM isopropyl 1-thio-\u03b2-D-galactopyranoside (IPTG). RNAi assay were performed according to pervious study [2 strain of C. elegans) fed on RNAi bacteria strains at 20 \u00b0C until reach to young adult. Then young adult worms were transferred on the BHI plates for further studies.Bacterial strains contain RNAi targeting genes were obtained from Ahringer library . E. colius study , in brieC. elegans strains were cultured on NGM plate with E. coli OP50 at 20 \u00b0C following the standard maintenance procedures [E. coli OP50 for further assays. To examine the effect of N. farcinica on worms\u2019 lifespan, synchronized L1 larvae were cultivated fed E. coli OP50 at 20 \u00b0C until the young adult stage. Then 50\u201360 worms were fed with either N. farcinica or laboratory standard diet E. coli OP50 on BHI medium containing 5\u2032-fluoro- 2\u2032-deoxyuridine (FUdR) (75 \u03bcg/mL) at 25 \u00b0C. The survival curves were plotted by scoring the dead worm at 24 h interval. Immobile worms unresponsive to touch were scored as dead. Three plates were analyzed per assay and all experiments were performed three times. The ocedures . Worms wocedures , eggs weskn-1b/c::gfp were fed with either N. farcinica or laboratory standard diet E. coli OP50 on BHI plates for 24 hours. Briefly, worms were mounted on slides from plates and imaged by using Nikon e800 fluorescence microscope. At least 30 worms were examined under each condition in three independent experiments.After young adult worms expressing After washed with M9 buffer, worms were homogenized in liquid nitrogen. Then the homogenate was lysed on ice for 30 min in lysis buffer RIPA . After centrifuged at 12,000 rpm for 15 min at 4 \u00b0C, the supernatant was obtained and used for Western blot analysis. The total protein extraction was loaded on 10% SDS-PAGE for electrophoresis. Proteins were then transferred to immobilon-PSQ transfer PVDF membrane . Primary antibodies were anti-phospho-p38 antibodies , and anti-\u03b1-tubulin antibodies . The secondary antibodies were peroxidase-coupled anti-rabbit IgG . Blots were developed using Super Signal chemiluminescence substrate . An imaging system (Amersham Imager 600) was used for documentation of the Western blot results. Band intensities were measured using ImageJ software (NIH).\u00ae Premix-Ex TagTM  on a Roche LightCycler 480\u00ae System . act-1 gene was used for an internal control. The primers used for PCR are listed in Total RNA was isolated from worms with TRIzol Reagent . Random-primed cDNAs were generated by reverse transcription of the total RNA samples with SuperScript II (Invitrogen). A real time-PCR analysis was conducted using SYBRnlp-29p::gfp or gst-4p::gfp were exposed to N. farcinica or E. coli OP50 for 24 h. Then the worms were mounted in M9 onto microscope slides. The slides were imaged using a Nikon e800 fluorescence microscope. Fluorescence intensity was quantified by using the ImageJ software (NIH). Mean value and standard errors were calculated based on more than 30 worms under each condition in three independent experiments.For detecting fluorescence in worms, analysis, synchronized young adult worms expressing either ROS was detected by transferring worms in to M9 buffer with DHE (3 \u03bcM) and stained for 3 h before mounting in M9 buffer onto microscope slide, examined by fluorescence microscope (Zeiss Axioskop 2 Plus) .N. farcinica colonized in worms, we fed young adult worms with either E. coli OP50 or N. farcinica for 96 h on BHI plates, then worms were transferred in to M9 buffer containing 25 mM levamisole hydrochloride , 50 \u03bcg/mL kanamycin and 100 \u03bcg/mL carbenicillin , and soaking for 30 min before washing with M9 buffer for three times. Worms were collected and grinded in PBS with 0.1% Triton, and serial diluted before placing on BHI ager for incubation, then bacterial colonies were counted to measure CFU.To ask if t-test. Differences in survival rates were analyzed using the log-rank test. Difference of CFU counting was assessed by two-tailed unpaired t-test. Data were analyzed using SPSSsoftware, version 26.0  and Graphpad Prism 8.The statistical significance of differences in gene expression and fluorescence intensity was assessed by performing a one-way ANOVA followed by a Student-Newman-Keuls test and two-tailed unpaired"}
+{"text": "Endoscopic Endonasal Surgery (EES) is an innovative surgical technique to remove brain tumors and lesions. Post-operative central nervous system (CNS) infections following EES are poorly described. The objective of this study was to define the epidemiology and characteristics of post-EES CNS infections.Adult patients who underwent EES between 1/2010 and 7/2021 were evaluated and included if microbiologically confirmed CNS infection occurred within 30 days of EES. Suspected contaminants, ventricular drain colonization, and pre-EES CNS infections were excluded.S. aureus, Enterobacterales, and P. aeruginosa. Among 20 patients with prior EES, pathogens included S. aureus (5/20), Enterobacterales (3/20), Enterococcus spp. (3/20) and polymicrobic infections (3/20). Overall, 35.1% (13/37) of patients developed CNS infection due to a pathogen susceptible to pre-EES prophylaxis. Among those colonized with MRSA at time of EES, 75% (3/4) developed MRSA CNS infection compared to 6.1% (2/33) of non-colonized MRSA patients (P=0.005). The overall 30-day mortality rate was 2.7% (1/37).Overall, 2005 patients underwent EES; 1.8% (37/2005) developed CNS infection. The median [IQR] age was 51 [42-60] years, 32.4% (12/37) were female, and 54% (20/37) had a prior EES. The most common indications for EES were tumor resection [67.6% (25/37)] and cerebrospinal fluid (CSF) leak repair [24.3% (9/37)]. Post-operative CSF leaks were documented in 70.3% (26/37) of patients and 24.3% (9/37) had an extra-ventricular drain or shunt in place for >48 hours at the time of infection. Ceftriaxone prophylaxis was prescribed in 64.9% (24/37) of cases and other regimens varied. The median [IQR] time from EES to diagnosis of CNS infection was 12 [6-19] days. The most common pathogens were A polymicrobic case was defined as >1 pathogen isolated from CSF (n=1) or from rhinocerebral tissue if CSF cultures were negative (n=11). Among polymicrobic cases (n=12), P. aeruginosa (n=5), Enterococcus spp. (n=4). and S. aureus (n=3) were predominant. Cases labeled as other consisted of Trichoderma spp, A. xylosoxidans, P. acnes, S. epidermidis, Peptostreptococcus spp.S. aureus, antimicrobial prophylaxis should ensure adequate coverage of this pathogen in addition to sinus flora, and programs may benefit from screening patients for MRSA colonization pre-EES. Our data also suggest that prophylaxis should target Gram-negative and other colonizing bacteria among patients with prior EES.CNS infection post-EES is rare and causative pathogens vary. Given the predominance of Ryan K. Shields, PharmD, MS, Infectious Disease Connect: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Roche: Grant/Research Support."}
+{"text": "Attention-Deficit/Hyperactivity Disorder (ADHD) is associated with alterations in both reinforcement sensitivity and affective processing but the nature of the associations of these characteristics is yet to be examined. We hypothesized that individual differences in the sensitivity of the Behavioral Approach System (BAS) would exhibit differential relations with affective network connectivity \u2013 involved in emotional regulation and salience monitoring \u2013 in youth at-risk for, relative to youth not at-risk for, ADHD.n = 125; Mage=16.24 years, SD = 1.09 years; 61.6% boys) were recruited as part of The Budapest Longitudinal Study of ADHD and Externalizing Disorders. Forty-nine were classified as at-risk for ADHD , defined as exhibiting \u22654 parent-rated symptoms of either domain on the ADHD Rating Scale-5. Participants completed a 10-minute resting-state functional Magnetic Resonance Imaging session, during which they were asked to focus their attention on a fixation cross, as well as various self-report assessments, including the Reinforcement Sensitivity Theory of Personality Questionnaire (RST-PQ).Adolescents  within a cluster based on functional similarity . Follow-up OLS linear regressions showed higher impulsivity scores predicted stronger functional connectivity between the (1) left amygdala-right insula  = 3.298, p = .005, adjusted R2=.101), (2) left amygdala-left insula  = 2.2, p = .048, adjusted R2=.055), (3) right amygdala-right insula  = 3.833, p = .002, adjusted R2=.121), and (4) right amygdala-left insula  = 3.064, p = .008, adjusted R2=.092) in at-risk youth, whereas an inverse relationship was apparent in not at-risk youth.Functional Network Connectivity analyses indicated an interaction effect between the RST-PQ BAS impulsivity subscale and at-risk status on functional connectivity between four affective network region-pairs (t(122)=\u22121.167, p = .246) or on functional connectivity ((1) t(122) = .383, p = .702; (2) t(122) = .195, p = .846; (3) t(122)=\u2212.107, p = .915; (4) t(122)=\u2212.206, p = .837).There was no main effect of group status on BAS impulsivity scores  as the role of the amygdala-insula connection in reward sensitivity appears especially relevant for a developmental phase and a diagnostic group linked to increased risk taking."}
+{"text": "Relative transformation efficiencies ranged from \u2212\u200944 to +\u200945% compared to Silwet\u00ae L-77. Surfactants S200, S240, and S279 demonstrated the greatest enhancement in transformation.Floral dip transformation of Arabidopsis has been used consistently for 20\u00a0years with little change in the protocol. Here we directly compare seven novel surfactants (BREAK-THRUThe online version contains supplementary material available at 10.1186/s13104-022-06251-5. Plant biotechnology offers tremendous promise for improving crops by increasing the sustainability and reducing the environmental impact of production. These technologies have led to crop plants with enhanced nutritional properties, abiotic stress tolerance, disease and pest resistance, and other agronomic traits, as well as crop tools for accelerated precision breeding. Although genetic engineering and genomic editing approaches have been applied successfully to various plant species, relatively few crops have an easy and efficient means of transformation, which restricts progress in research as well as agricultural deployment. Thus, additional techniques for broadly and efficiently applying transformation technologies to a wide range of crops and their constituent elite varieties are required.One of the earliest enhancements to transformation was the use of surfactants, which modify the surface properties of liquids to enhance their spreading, wetting, emulsifying, dispersing, or other characteristics. Surfactants have been used extensively in herbicide and pesticide formulations to increase their penetration. However, they must be used judicially, as surfactants may have inhibitory phytotoxic effects on plant tissue at high concentrations and stimulatory effects at low concentrations , 2.Agrobacterium-mediated transformation of Arabidopsis thaliana  in water: 25 (0.1%).BREAK THRU\u00ae-S200: Trisiloxane super-spreader (more water-soluble), stable within pH range 6\u20138, Static surface tension [mN/m] in water: 22 (0.1%).BREAK THRU\u00ae-S233: Trisiloxane super-penetrant  systemic products, stable within pH range 6\u20138, Static surface tension [mN/m] in water: 23 (0.1%).BREAK THRU\u00ae-S240: Trisiloxane super-spreader (liquid soluble), stable within pH range 6\u20138, Static surface tension [mN/m] in water: 22 (0.1%).BREAK THRU\u00ae-S279: Trisiloxane super-spreader (more oil-soluble), stable within pH range 6\u20138, Static surface tension [mN/m] in water: 21 (0.1%).BREAK THRUS301: Trisiloxane super-spreader, stable within pH range 6\u20138, Static surface tension [mN/m] in water: 22 (0.1%).\u00ae-SP133: Polyglycerol ester-based adjuvant , stable within pH range 4\u20139, Static surface tension [mN/m] in water: 29 (0.1%).BREAK THRU\u00ae compound see (www.evonik.com/break-thru) [For more details of the BREAK THRUak-thru) .We show an increase in transformation efficiency for Arabidopsis floral dip transformation using a published technique . HoweverAdditional file 1.\u00a0Statistics 1: Figure S1.Additional file 2.\u00a0PCR: Figure S2.Additional file 3.\u00a0Statistics 2: Table S1. Surfactant transformation efficiency. Table S2. Surfactant transformation statistics."}
+{"text": "Nature Communications 10.1038/s41467-021-25159-5, published online 09 August 2021Correction to: Nat. Commun. 12, 1\u201314 (2021). The correct reference for Ref. 11 is: Zhang, Y. et al. mTORC1 couples cyst(e)ine availability with GPX4 protein synthesis and ferroptosis regulation. Nat. Commun. 12, 1589 (2021). This has been corrected in the PDF and HTML versions of the Article.The original version of this Article contained an error in Ref. 11, which incorrectly gave the reference as: Zhang, Y. et al. mTORC1 couples cyst(e)ine availability with GPX4 protein synthesis and ferroptosis regulation."}
+{"text": "E.coli (STEC) infection which presents a characteristic triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. Preliminary in vitro and experimental animal studies demonstrated that Shigatoxins (STXs) induce the secretion of proinflammatory cytokines. Human neutrophil gelatinase-associated protein  has been reported as a marker of acute kidney injury. Dosing serum levels of IL-8, TNF-\u03b1, IL-6, IL-1\u03b2 and N-gal in children with STEC-associated infection and HUS would allow to establish the role of these cytokines as biomarkers of renal injury and severityHemolytic Uremic Syndrome (HUS) is a complication of Shigatoxin-producing Prospective study during 2017 \u2013 2020 was performed; three groups of patients < 18 years were included: bloody diarrhea (BD), HUS requiring dialysis (HUSD) and HUS and no dialysis requirement (HUSND), all of them with presence of STX in stool. Blood samples were collected at diagnosis (t1) and at 7 days (t2). An immunoassay was used for detection of TNF-\u03b1, IL-1\u03b2, IL-6, and IL-8 . An immunoassay  was used for N-gal detectionForty-nine children were admitted; 22 (49%) were male. Median age was 24 (IQR 13-36) months. Fourteen patients with BD, 24 patients with HUS: 12 HUSD and 12 HUSND. Eleven healthy children (HC) were included. At diagnosis, higher IL-8 values were found in HUSD vs. BD (p=0.0004), HUSND (0.003) and HC (p= 0.0043). Higher TNF-\u03b1 values were detected in HUSD vs. BD (P= 0.0001), HUSND (p= 0.0011) and HC (p= 0.0002). Higher TNF-\u03b1 values were found in patients with BD vs HC (p= 0.0055). HUSD exhibited higher IL-6 values as compared with BD (p= 0.0145) and HC (0.0106). N-gal concentrations were higher in HUSD vs. BD (P= 0.0120), HUSND (P= 0.0166) and HC (p= 0.0049). A decrease in IL-8 was observed in HUSD between t1 and t2 (p=0.0357). Interleukin 8, IL-6, and N-gal may be considered as potential serological markers of renal damage and dialysis requirement. TNF-\u03b1 concentrations are already increased in BD without HUS complicationsAll Authors: No reported disclosures."}
+{"text": "In the published article, the reference for Immunotherapy is an emerging tool used in cancer treatment (1) was incorrectly written as Ruhlmann CH, Iversen TZ, Okera M, Muhic A, Kristensen G, Feyer P, et\u00a0al. Multinational study exploring patients\u2019 perceptions of side-effects induced by chemo-radiotherapy. Radiother Oncol (2015) 117(2):333\u20137. doi: 10.1016/j.radonc.2015.09.014. It should be Mohanty R, Chowdhury CR, Arega S, Sen P, Ganguly P, Ganguly N. CAR T cell therapy: A new era for cancer treatment (Review). Oncol Rep (2019) 42(6):2183-95. Epub 2019/10/04. doi: 10.3892/or.2019.7335.In the published article, the reference for The fourth-generation CARs added genes encoding cytokines [interleukin (IL)-12 and IL-15] to be released by the CARs to improve CAR T-cell survival in a TME (32) was incorrectly written as Haso W, Lee DW, Shah NN, Stetler-Stevenson M, Yuan CM, Pastan IH, et\u00a0al. Anti-Cd22-chimeric antigen receptors targeting B-cell precursor acute lymphoblastic leukemia. Blood (2013) 121(7):1165\u201374. doi: 10.1182/blood-2012-06-438002. It should be Chmielewski M, Kopecky C, Hombach AA, Abken H. IL-12 release by engineered T cells expressing chimeric antigen receptors can effectively muster an antigen-independent macrophage response on tumor cells that have shut down tumor antigen expression. Cancer Res (2011) 71(17):5697-706. Epub 2011/07/12. doi: 10.1158/0008-5472.Can-11-0103.In the published article, the reference for The fifth-generation CARs build on the second-generation CARs by adding cytoplasmic structural domains from the IL-2 receptor beta chain and signal transducers and activators of transcription (STAT)3/5 binding pattern, triggering three signals including T-cell receptors , costimulatory factors , and cytokines  to improve the proliferation, survival, and antitumor activity of CAR T cells markedly (33) was incorrectly written as Chmielewski M, Kopecky C, Hombach AA, Abken H. IL-12 release by engineered T cells expressing chimeric antigen receptors can effectively muster an antigen-independent macrophage response on tumor cells that have shut down tumor antigen expression. Cancer Res (2011) 71(17):5697\u2013706. doi: 10.1158/0008-5472.Can-11-0103. It should be Kagoya Y, Tanaka S, Guo T, Anczurowski M, Wang CH, Saso K, et\u00a0al. A novel chimeric antigen receptor containing a JAK-STAT signaling domain mediates superior antitumor effects. Nat Med (2018) 24(3):352-9. Epub 2018/02/06. doi: 10.1038/nm.4478.In the published article, the reference for As conventional surgical treatment and antitumor drugs have limited effects, CAR T-cell therapy can provide targeted immunotherapy to patients with gastric cancer without developing drug resistance and effectively control the progression and metastasis of gastric cancer (62) was incorrectly written as Dur\u00e3es C, Almeida GM, Seruca R, Oliveira C, Carneiro F. Biomarkers for gastric cancer: Prognostic, predictive or targets of therapy? Virchows Arch (2014) 464(3):367\u201378. doi: 10.1007/s00428-013-1533-y. It should be Jiang H, Shi Z, Wang P, Wang C, Yang L, Du G, et\u00a0al. Claudin18.2-specific chimeric antigen receptor engineered T cells for the treatment of gastric cancer. J Natl Cancer Inst (2019) 111(4):409-18. Epub 2018/09/12. doi: 10.1093/jnci/djy134.In the published article, the reference for Animal models are constructed by gene editing to mimic specific biological characteristics of human diseases to introduce target genes or delete and modify endogenous genes (83) was incorrectly written as Hu W, Lazar MA. Modelling metabolic diseases and drug response using stem cells and organoids. Nat Rev Endocrinol (2022) 1\u201316. doi: 10.1038/s41574-022-00733-z. It should be Platt RJ, Chen S, Zhou Y, Yim MJ, Swiech L, Kempton HR, et\u00a0al. CRISPR-Cas9 knockin mice for genome editing and cancer modeling. Cell (2014) 159(2):440-55. Epub 2014/09/30. doi: 10.1016/j.cell.2014.09.014.The authors apologize for these errors and state that these do not change the scientific conclusions of the article in any way. The original article has been updated."}
+{"text": "Frequent users of Emergency Departments (EDs) are a diverse group accounting for disproportionate EDs visits. Psychiatric patients are more likely to visit EDs . EDs utilisation by psychiatric patients increased by 4.4% during COVID-19 pandemic.to determine frequent users characteristics within an Ottawa University Hospital, and assess Covid19 impact on overutilization of EDs compared to other hospitals.Retrospective study of repeat visits characteristics, data extracted from EMR database. Repeat visits defined as no less than 30 days first visit to any EDs. Period of observation: March 1st, 2018 - February 28th, 2021 Results.64% EDS visits for MH, 35% for addictions. More men (57%), age groups: 16-34 y.o. (41%), 34-64 y.o. (51%), 65 +y.o. (8%).Top presenting reasons: suicidality, self-harm, depression (40.5%). Anxiety, situational crisis (16%), bizarre behavior (12%).Most prevalent diagnoses: schizophrenia (28.7%), stress and anxiety (25.2%), personality disorders (13.5%) and depressive episode (10.6%). Only 35.1% admitted after repeat ED visits, 35.1% came by ambulance. Increase during peak pandemic exceeding 20%. Clearly pandemic created more pressures for MH services needs.Schizophrenia and personality disorders made most prevalent diagnostic groups. Even when patients are in acute needs, they do not always require hospitalization. Investigating what MH conditions that got more stressed by the Covid19 pandemic will be of interest.No significant relationships."}
+{"text": "Gossypium raimondii Ulbrich (short fibers) with cultivated diploid G. arboreum L and tetraploid G. hirsutum L. (long fibers); 2) G. hirsutum short fiber mutants, Ligon-lintless 1 (Li1) and 2 (Li2) with their near isogenic line (NIL), DP-5690 (long fibers). Chemical analyses showed that the short fibers commonly consisted of greater non-cellulosic components, including lignin and suberin, than the long fibers. Transcriptomic analyses also identified up-regulation of the genes related to suberin and lignin biosynthesis in the short fibers. Our results may provide insight on how high levels of suberin and lignin in cell walls can affect cotton fiber length. The approaches combining phenomic and transcriptomic analyses of multiple sets of cotton fibers sharing a common phenotype would facilitate identifying genes and common pathways that significantly influence cotton fiber properties.Fiber length is one of the major properties determining the quality and commercial value of cotton. To understand the mechanisms regulating fiber length, genetic variations of cotton species and mutants producing short fibers have been compared with cultivated cottons generating long and normal fibers. However, their phenomic variation other than fiber length has not been well characterized. Therefore, we compared physical and chemical properties of the short fibers with the long fibers. Fiber characteristics were compared in two sets: 1) wild diploid  Gossypium sp.) is the most economically important natural fiber in the world . Pe. PeArabiCW stage . Three Gi fibers .Li1 and Li2) was performed with total RNAs extracted from developing fibers at PCW stage (8\u201312 DPA) grown in greenhouse or cotton fields [Li1 and Li2 fibers were compared to their NIL, G. hirsutum DP-5690, using a 2-fold difference as a threshold. In the Li1 mutant fibers, 4,043 genes were up-regulated whereas 2,536 genes were down-regulated  was performed with the spectral region (1200\u20133000 cm-1) composed of IR peak bands of suberin, lignin, and cellulose (G. hirsutum Li2 < G. hirsutum Li1 < G. raimondii D5-31 < G. hirsutum TM-1 \u2248 G. hirsutum DP-5690 \u2248 G. arboreum A2-100. The three cultivated cottons of G. arboreum A2-100 (0.369), G. hirsutum DP-5690 (0.365), and G. hirsutum TM-1 (0.326) demonstrated similar positive PC1 scores with insignificant (p = 0.587) variations. In contrast, the other three short fiber cottons of G. hirsutum Li2 (-0.738), G. hirsutum Li1 (-0.279), and G. raimondii D5-31 (-0.043) had all negative scores that were significantly (p<0.0001) variable from the cultivated cotton species. Among the three short fiber cottons, the PC1 scores also showed significant (p<0.0001) variations. These results suggested that the chemical compositions of G. raimondii D5-31, G. hirsutum Li1, and G. hirsutum Li2 were different despite a sharing of suberin and lignin fractions in addition to cellulose.For an examination of the quantitative and statistical significances of the spectral features showing different chemical components among the fiber samples used in the 1ellulose . The anaellulose . The PC1G. hirsutum Li1  and Li2  fibers (G. hirsutum Li1 (880) and Li2 (376) mutants. To identify the commonly up-regulated orthologous genes between the diploid D5 genome and polyploid AD1 genome composed of AT and DT subgenomes, we compared the diploid G. raimondii 1,385 SEGs with non-redundant UGs in polyploid G. hirsutum Li1  and Li2  (T) and a peroxidase (Gorai.013G005800_AT) involved in lignin polymerization by oxidizing lignin monomers (monolignols) [T) required for suberin biosynthesis in Arabidopsis [Arabidopsis color mutant was also up-regulated [Of the UGs in ) fibers , there w  . Among t  . Consistlignols) , 53, 54 bidopsis  were foubidopsis  was alsoegulated . The othegulated , leucineegulated , FAD-binegulated , glutathegulated , PLAT/LHm Li1 3,1 and Li2 egulated , cytokinegulated , PSBP-doegulated , cyclic egulated , and NACegulated  were allG. raimondii and G. hirsutum Li1 and Li2 mutants were too short to be measured by HVI. Thus, we manually measured the maximum fiber lengths of the wet and relaxed cotton fibers from the chalezel end of cottonseeds [G. raimondii D5-6 (11.7 mm) and D5-31 (10.1 mm) as well as G. hirsutum Li1 (2.6 mm) and Li2 (8.1 mm) mutants were significantly shorter than those of cultivated diploid cotton, G. arboreum SXY1 (28.0 mm) and A2-100 (30.1 mm) as well as cultivated polyploid cotton, G. hirsutum TM-1 (41.7 mm), SG-747 (40.1 mm) and DP-5690 (38.7 mm) as shown in Figs G. hirsutum fibers. Lint fibers differentiate from ovule epidermis on the day of anthesis and they grow approximately 25~35 mm based on HVI measurements. In contrast, linter or fuzz differentiate from the ovule epidermis around 5 to 10 DPA and they do not grow longer than 15 mm [G. raimondii was often described as a lintless, non-fibered, or fiberless species [Li1 and Li2 mutants also contain \u201clintless\u201d [G. raimondii [G. hirsutum Li1 mutant [G. hirsutum Li2 mutant [G. raimondii and G. hirsutum Li1 and Li2 mutants can be classified as lint fibers according to the definition described by Lang [Physical properties of cultivated cotton fibers are generally assessed by a High Volume Instrument (HVI) which is defined by the International Cotton Advisory Committee as a standardized instrument for cotton fiber quality measurements . Cotton tonseeds . The maxan 15 mm . Wild di species , 69, 70.intless\u201d , 18. How1 mutant , and G. 2 mutant  all diff by Lang .G. raimondii and two G. hirsutum mutants produced fibers containing color pigments composed of lignin and suberin  and G. hirsutum Li1 and Li2 mutants (85.1~86.9%) fibers were lower than cultivated G. arboreum (95.6~100%) and G. hirsutum (95.8~98.0%) fibers  and G. hirsutum Li1 and Li2 (13.1~14.9%) fibers were substantially greater than the cultivated fibers (0~4.4%). These results were consistent with the previous reports showing different cellulose contents between green and white upland cotton [G. raimondii and cultivated G. arboreum [Chemical analyses using cellulose assay, ATR FT-IR spectroscopy, and mass spectrometry consistently showed suberin and lignin components in the three short fibers. Average cellulose contents of ers Figs  and 3B. d cotton  and funcarboreum .G. raimondii and G. hirsutum Li1 and Li2 mutants demonstrated the signature IR spectral peaks of suberin and lignin  than those of the other cultivated cotton fibers (0~ 0.6%). Recent studies suggest that lignin may play an important role in cotton fiber quality [The three short cottons of nin Figs  and 5B.  suberin . In natun fibers , 74, andn fibers . Suberinn fibers . In contn fibers . Generaln fibers . Our mas quality , 79.G. hirsutum Li1 (1623 cm-1), G. hirsutum Li2 (1610 cm-1), and G. raimondii D5 (1635 cm-1)  Figs  and 5B. -1) Figs .G. raimondii fibers, we used the RNA-seq data performed with the RNAs extracted from developing fibers of G. raimondii, G. arboreum, and G. hirsutum (Li1 and Li2) and their NIL DP-5690 were only performed with total RNAs extracted from developing fibers at PCW stage [G. raimondii fibers [G. raimondii D5 reference genome for analyzing transcriptomic profiles of the two sets because the G. raimondii genome sequence shows high homology (>96%) with the coding sequences of G. hirsutum AT and DT subgenomes. Thus, the D5 reference genome sequence has been successfully used for characterizing the Li1 and Li2 genomes by several groups [st set identified that genes involved in suberin and lignin biosynthesis were specifically expressed in G. raimondii fibers Click here for additional data file.S1 Table(XLSX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(XLSX)Click here for additional data file.S4 Table(XLSX)Click here for additional data file.S5 Table(XLSX)Click here for additional data file."}
+{"text": "Dear Editor,3. The intrinsic nature of MLKL and how it induces plasma membrane permeabilization, forming a huge pore or a channel, remain an interesting conundrum for a long time5. Our previous study demonstrated that MLKL forms cation channels and its channel activity is a primary effector of necroptosis6. In the field of MLKL, a major unsettled issue is, if MLKL does serve as channels, how it ignites the necrotic or non-necrotic pathogenic progresses5. Phosphatidylinositol 4,5-bisphosphate P2) is a necessary cofactor of various ion channels7. The physical interaction between MLKL and phosphatidylinositol phosphates (PIPs), including PIP2, facilitates MLKL-mediated liposome leakage and the necrotic membrane disruption10. These studies have been directed toward understanding whether PIP2 is a direct modulator for the MLKL channel activity. The potential effects of PIP2 on MLKL channels were thus evaluated electrophysiologically and the subsequent pathogenic influences were explored.Mixed lineage kinase domain-like protein (MLKL) emerged as executioner of necroptosisNT) is sufficient to induce oligomerization and trigger cell death9. Consistent with our previous study, MLKLNT protein exhibited channel activity, translocated onto plasma membrane, and caused cell death, similarly to full-length protein (MLKLFL) P2  was enhanced to 27.4%  and phosphatidylinositol 3,5-bisphosphate P2), phosphatidylinositol 4-phosphate (PI(4)P), 1,4,5-trisphosphate inositol (IP3) and diacylglycerol (DAG), phosphatidylinositol 3,4,5-trisphosphate (PIP3)11. Among these lipids, PI(4)P is the only one that increases MLKL channel open probability P2. An increased concentration of PIP2 induced more frequent and larger currents of MLKL channel P2 levels in live cells. Stimulation of the M1 muscarinic receptor can activate PLC-\u03b2 which hydrolyzes PIP2. PLC-\u03b2-PH-GFP (PLC-PH) construct, a PIP2 reporter, is used to monitor the PIP2 distribution on plasma membrane12. Here, MLKL, M1 receptor, and PLC-PH were co-transfected into HEK293 cells. With 5\u2009\u03bcM oxotremorine M (Oxo-M) treatment, the PLC-PH probe dissociated from the plasma membrane to cytoplasm, showing PIP2 hydrolyzation  in the N-terminal region of MLKL (MLKL)P2 Fig. . The ove.4% Fig. . These s.4% Fig. . Influenity Fig. . Collectnel Fig. . To furtion Fig. . Meanwhiion Fig. . These d2-induced augmentation of MLKL channel activity is correlated with cell death was examined. To elevate PIP2 concentrations in the plasma membrane, PIP5K, one of the key PIP2 biosynthetic enzymes, was transfected into L929 cells P. 1 Fig. . In addi. 1 Fig. . Similar2-sensitive ion channels, PIP2 binds with positive amino acid via \u201celectrostatic interaction\u201d13. We mutated the positive amino acids of MLKLNT to alanine and tested the channel activity  P2 sensitivity or abrogate the capability of killing cells after overexpression of PIP5K, suggesting that K22 and R34 may interact with PIP2 independently P. 2 Fig. . Notably4%) Fig. . Whetherls9 Fig. . Consisttly Fig. .14. Thus, we further explored the potential function of PIP2-enhanced MLKL channel activity in an inflammation model12  through String database (https://string-db.org), and further revealed the close relationship between PIP2, MLKL and LPS-induced inflammation -like receptor protein 3 (NLRP3) in a cell-intrinsic manner before cell lysis, but its working model has not yet been clarifiedins Fig. . To inveion Fig. .+ efflux in triggering inflammation has been proposed15. We therefore hypothesized that K+ efflux during inflammation is mediated by MLKL channels. A real-time, sensitive FluxOR\u2122 assay was performed to monitor the K+ efflux during the LPS-induced inflammation. We found that the K+ efflux gradually increased in a time-dependent manner  treated with LPS, Smac-mimetic compound (C) and a caspase inhibitor Q-VD-OPh (Q)14. Both the MLKL-mediated cytokine secretion and the LPS-induced K+ efflux were abrogated in MLKL-knockout (MLKL\u2013/\u2013) cells  of LPS significantly stimulated the secretion of inflammatory cytokines, but did not induce cell death, while the expression level of MLKL and PIP5K increased Fig. . Accordiner Fig. . The conlls Fig. . Whetherels Fig. . Since e2 in a dose-dependent manner for the first time Pime Fig. . PIuli Fig. . WhetherSupplementary informationSupplementary Tabel S1"}
+{"text": "Retraction of: Hua Liu, Jing Cheng, Heng Xu and Zhenzhen Wan. Lidocaine has antitumor effect on hepatocellular carcinoma via the circ_DYNC1H1/miR-520a-3p/USP14 axis. Open Life Sciences. Volume 16, Issue 1, doi: 10.1515/biol-2021-0072.https://doi.org/10.1515/biol-2021-0072) has been retracted due to the previously undisclosed conflict of interest between the authors. Authors apologize to the entire scientific community and Editorial team for any issues ensuant from this action.\u201cLidocaine has antitumor effect on hepatocellular carcinoma via the circ_DYNC1H1/miR-520a-3p/USP14 axis\u201d ("}
+{"text": "Pandemic coronavirus causes respiratory, enteric and sometimes neurological diseases. Proteome data of individual coronavirus strains were already reported. Here we investigated of SARS-CoV-2 ssRNA and protein of spike, envelope and membrane to determine stress adaptation profile. Thermodynamic properties, Physicochemical behaviour and, amino acid composition along with their RMSD value was analysed. Thermodynamic index of SARS-CoV2 spike, envelope and membrane ssRNA is unstable in higher temperature. Presence of higher proportion of polar with positive and negative charged amino acid residues into spike (S), envelope (E) and membrane (M) protein indicate the lower stress adaptability pattern. Our study represented several unstable pockets into S, E and M proteins of SARS-CoV-2 against different abiotic stresses, specifically higher in spike protein. Contact with heat through solvent may denature the architectural network of SARS-CoV-2 spike, envelope and membrane ssRNA and structural protein. The stress instability index of SARS-CoV-2 and the interactome profile of its transmembrane proteins may help to reveal novel factors for inhibiting SARS-CoV-2 growth. Zoonotic beta coronaviruses can lead to severe acute respiratory syndrome or mild\u00a0pneumonia . Coronav. The envelope (E) protein of SARS-CoV-2 contains 76 to 109 amino acids with a small (7-12 amino acids) integral hydrophilic tail in C- terminal side; whereas the transmembrane portion having a large N-terminal domain with \u03b1 helical nature [Coronavirus structural proteins include the spike (S)protein, envelope (E)protein, membrane (M)protein, and nucleocapsid (N)protein , 7. The l nature , 22 The l nature . The M pl nature , 24, 25.SARS-CoV-2 assemble promotes by spike protein which may influence by the expression of envelope and membrane proteins. Additionally D814 to G814 mutation into spike protein of SARS-CoV-2 increases the attachment with host which results enhancing infectivity , 27. A LStudy design: Our study's objective to evaluate and build up stress stability and instability of SARS-CoV-2 based on the ssRNA along with extracellular and transmembrane protein. SARS-CoV-2 Spike ssRNA Sequence were primarily obtained from GENBANK Database. Accession number of SARS-CoV-2 Wuhan: MT079854.1, Alpha: MZ314997, Beta: MZ314998.1, Delta: OK091006.1, Gama: MZ315141.1 and Omicron: OL672836.1 were retrieve for analysis. We focused on the physicochemical and structural properties of these three (the spike (S) protein, envelope (E) protein, membrane (M) protein) protein structures available on the PDB database 6CRV, 5X29 and 3I6G accordingly.Stability index of SARS-CoV-2 ssRNA Secondary Structures: Vienna RNA Web Services (http://rna.tbi.univie.ac.at/) predicted the secondary structure of the RNAs of SARs-Cov-2 [Rs-Cov-2 , 33. In Rs-Cov-2 .Structural Simulation with diversity: Variant of concerns (VOCs) were reported by CDC i.e alpha, beta, gamma, delta, and omicron from the five most heavily affected countries  also represented by \u201cNextstrain / ncov / open / global / 6m\u201d. The corresponding amino acid sequences were taken and modelled [0 with a single point charge (SPC) water model TIP3P with periodic boundary condition (PBC). Each protein was modeled using OPLS3e force where Na+ and Cl- ions were added to neutralize the charge. The system was energy minimized 2000 steps before a production run of 50 ns. RMSD and RMSF values of the backbone atoms of proteins were calculated for determining equilibrium parameters [modelled  by usingmodelled . Using trameters . The RMSrameters .Amino acid frequency and structural elements: We computed the frequencies of amino acid residues in the protein sequences of the SARS-CoV-2 proteome by using MEGA [ing MEGA . Three mBased on the genomic organization, thermodynamics stability indexes of the SARS-CoV2 was analysed . Virus sThe thermodynamic parameters for the SARS-CoV2 single-stranded RNA are as follows: N-terminal domain (NTD), Receptor binding domain (RBD), Fusion protein (FP), Heptapeptide Repeat 1 (HR1), Heptapeptide Repeat 2 (HR2), Transmembrane region (TM), Cytoplasmic domain (CD), Envelope (E) and Membrane (M) the melting temperature were resulted 70.5 \u00b0C, 72.7\u00b0C, 87.8\u00b0C, 70.9\u00b0C, 72.6\u00b0C, 70.5\u00b0C, 70.9\u00b0C, 74.6\u00b0C, 73.3\u00b0C; whereas the Gibbs free energy were tabulated -196.46 kcal/mol,-113.8 kcal/mol,-11.2 kcal/mol, -42.8 kcal/mol, -29.3 kcal/mol, -19.8 kcal/mol, -28.1 kcal/mol, -54.7 kcal/mol and, -180.4 kcal/mol; similarly enthalpy were studied -2011.7 kcal/mol, -1101.5 kcal/mol, -79.5 kcal/mol, -433.7 kcal/mol, -248.5 kcal/mol, -202.9 kcal/mol, -284.5 kcal/mol, -531 kcal/mol and, -1720.8 kcal/mol; the entropy were resulted -5852.7 cal/mol, -3184.5 cal/mol, -220.2 cal/mol, -1260.3 cal/mol, -822.8 cal/mol, -590.3 cal/mol, -826.6 cal/mol, -1535.7 cal/mol and, -4966.6 cal/mol. These parameters indicate that the SARS-CoV2 ssRNA is unstable accordingly .Amino acid usage patterns are influenced by natural selection pressure at the protein structural stability level; we have compared the amino acids robustness in the chosen proteome which are the most dominant factor in determining stability. Structural comparisons reveal that S, M & E of all the VOC\u2019s are similar. RMSD value of Spike, Membrane and Envelope 0.022\u00b10.01, 0.068 and 0.055 accordingly. Furthermore, some of the propensities of non polar amino acids  with polar amino acids ( i.e Thr and Ser) were higher in the case of S protein; whereas, the opposite was noticssed in E and M protein (Supplementary Table S2). The polar, non-polar, positive and negative charge amino acids distribution were plotted and displayed into the peptide of spike , envelop).The spike protein interface exhibits five thermo-unstable cavity that buries residues 155, 247, 248 250, 254 and 258 in pocket1, pocket2  and pocket3  represented turn structure whereas pocket4  and pocket5  exhibited sheet structure . Thermo-Fig. 3FE-protein represented several stress unstable and stable complexes. The heat unstable complex, pocket1  and pocket2  both have helix structure . WhereasM-protein interface cavity consistently contained cavities, few of them was stress unstable with stable pockets. The heat unstable complex; pocket1  showed sheet structure whereas pocket2  displayed helix structure . ThermosBiothermodynamics\u00a0is\u00a0the\u00a0exploration\u00a0of\u00a0the\u00a0forces\u00a0and\u00a0processes\u00a0that\u00a0cause\u00a0biological occurrence Gibbs\u00a0energy\u00a0is\u00a0the\u00a0driving\u00a0force\u00a0behind\u00a0all\u00a0natural\u00a0processes. The Gibbs free energy in growth is influenced by the chemical constitution of the organism and also depends on the environment, especially intermolecular interactions between reaction participants. By taking over the host translation machinery , virusesSTn) pockets and two stable (PST) pockets against heat, two unstable (PSALn) and four stable (PSAL) pockets against alkali, two unstable (PSAn) and stable (PSA) pockets against acidic conditions (ETn) pockets and one stable (PET) pocket against heat, one unstable  and two stable  pockets against alkali with an acidic condition  pockets and one stable  pocket against heat and acidic conditions; similarly it also represented one unstable (PMALn) and stable (PMAL) pocket against alkaline conditions  in the interaction site of human ACE2 with SARS-CoV-2 spike protein (Supplementary Table S3), which may be considered for further studies for the development of a new efficient anti-corona drug [Spike protein of SARS-CoV-2 binds significantly to human ACE2, TMPRSS2 and CD26 are already reported through several amino acid residual interactions. We found a heat unstable pocket (Pona drug , 48. Theona drug , 50. OurWe thank the Oriental Institute of Science and Technology, India for supporting our work.The authors declare that they have no conflict of interest.Supplementary Material 1Supplementary Material 2"}
+{"text": "There are limited data on the prevalence of doravirine (DOR)-associated drug resistance mutations in people with HIV (PWH) in Botswana. This cross-sectional, retrospective study aimed to explore the prevalence of DOR-associated resistance mutations among ART-na\u00efve and -experienced PWH in Botswana enrolled in the population-based Botswana Combination Prevention Project (BCPP).pol sequences were analysed for DOR-associated resistance mutations using the Stanford HIV drug resistance database, and their levels were predicted according to the Stanford DRM penalty scores and resistance interpretation. Virologic failure was defined as HIV-1 RNA load (VL) >400 copies/mL.A total of 6078 HIV-1C P < 0.01). Intermediate DOR-associated resistance mutations were observed in 106/1261 (7.8% [95% CI: 6.9\u201310.1]) in ART-na\u00efve individuals and 29/212 (13.7% [95% CI: 9.4\u20138.5]) among ART-experienced participants (P < 0.01). High-level DOR-associated resistance mutations were observed in 33/1261 (2.6% [95% CI: 1.8\u20133.7]) among ART-na\u00efve and 13/212 (6.1% [95% CI: 3.6\u201310.8]) among ART-failing PWH (P < 0.01). PWH failing ART with at least one EFV/NVP-associated resistance mutation had high prevalence 13/67 (19.4%) of high-level DOR-associated resistance mutations.Among 6078 PWH, 5999 (99%) had known ART status, and 4529/5999 (79%) were on ART at time of sampling. The suppression rate among ART-experienced was 4517/4729 (96%). The overall prevalence of any DOR-associated resistance mutations was 181/1473 ; by ART status: 42/212 (19.8% [95% CI: 14.7\u201325.4]) among ART-failing individuals (VL \u2265400 copies/mL) and 139/1261 (11.0% [95% CI: 9.3\u201312.9]) among ART-na\u00efve individuals (DOR-associated mutations were rare (11.0%) among ART-naive PWH but present in 62.7% of Botswana individuals who failed NNRTI-based ART with at least one EFV/NVP-associated resistance mutation. Testing for HIV drug resistance should underpin the use of DOR in PWH who have taken first-generation NNRTIs. The avaA newly approved third-generation NNRTI, doravirine (DOR), has been shown to be more potent, with distinct resistance patterns compared to the earlier-generation NNRTIs 22.1PWH aged 16\u201364 years in 30 communities in northern, central and southern parts of Botswana participated in the Botswana Combination Prevention Project (BCPP) between 2013 and 2018 2.2In this analysis, we included PWH who were either ART-na\u00efve or ART-experienced, who had HIV-1 viral load (VL) measurement at the first BCPP study visit and had available HIV-1 sequence. HIV-1 VL of participants was quantified using Abbott m2000sp/rt assay  with a range of 40\u201310 000 000 copies/mL 2.3https://genome.med.harvard.edu/) and through collaboration with PANGEA HIV consortium www.sanger.ac.uk/) with high-sequencing coverage using Illumina platforms MiSeq and HiSeq Both HIV-1 proviral DNA sequences and viral RNA sequences were generated by a long-range HIV genotyping protocol described elsewhere ,23. The 2.4Generated near full-length HIV-1 sequences were subtyped by online tools REGA version 3 2.5https://hivdb.stanford.edu/hivdb/by-sequences/). The level of DOR resistance was predicted according to the Stanford HIV DRM penalty scores and resistance interpretation . Only intermediate and high-level specific DOR-associated resistance mutations were reported. The list of mutations assessed are shown in DOR-associated resistance mutations were identified according to the lists of surveillance drug resistance mutations and major drug resistance mutations (DRMs) in the Stanford University HIV Drug Resistance Database algorithm 9.1  overall; (ii) ART-na\u00efve individuals; (iii) virologic failure (VF) on ART; and (iv) viral suppression (HIV-1 RNA \u2264400 copies/mL) on ART. The DOR prevalence was compared between groups (ii) and (iii), as well as between groups (iii) and (iv). The prevalence of specific DOR-associated resistance mutations was estimated within each group and compared among groups. We also assessed the presence of DOR-associated resistance mutations in a subset of participants on NNRTI-based regimens with at least one major efavirenz (EFV)/nevirapine (NVP)-associated resistance mutation .2.6http://www.hiv.lanl.gov/) pol gene. The adjustment for hypermutations was performed before the drug resistance analysis, as a part of quality control. HIV DRMs identified as hypermutations were not included in the prevalence of DOR resistance in this study.Guanine-to-adenine transitions (G-to-A) apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like (APOBEC)-induced hypermutations were screened in the viral sequences using the Hypermut program available in Los Alamos National Laboratory HIV Database tools  in NNRTI-failing individuals with at least one EFV/NVP-associated resistance mutation for detection of DOR-associated resistance mutations. A P value <0.05 was considered statistically significant. All the statistical analysis was done using STATA version 14 software.Patient demographics between ART-na\u00efve and individuals on ART experiencing VF were compared using Wilcoxon rank-sum test for continuous variables such as plasma log2.8The BCPP study was approved by the Institutional Review Board (IRB) at the U.S. Centers for Disease Control and Prevention and the Botswana IRB (HRDC), and it is registered at ClinicalTrials.gov (NCT01965470). All recruited participants provided written informed consent for participation.33.110 copies/mL) was similar to the median HIV-1 VL among individuals experiencing VF on ART . The study entry demographics of BCPP participants stratified by HIV-1 VL groups are in The median age at enrolment of the participants included in this analysis was 34 years , and study participants were mostly women (66%). Among 6078 participants, 5999 (99%) had known ART status (either \u2018on ART\u2019 or \u2018ART-naive\u2019). The majority of these participants, 4738 (79%), were on ART, whereas 1261 (21%) were ART-na\u00efve at the time of sampling. Among 4738 participants, a total of 4729 had HIV-1 VL data; 4517 (96%) were virally suppressed (HIV-1 VL \u2264400 copies/mL). The median HIV-1 VL among ART-na\u00efve individuals . A majority of the participants harboured intermediate DOR-associated resistance mutations with a lower prevalence of 106/1261 (7.8% [95% CI: 6.9\u201310.1]) in ART-na\u00efve individuals and 29/212 (13.7% [95% CI: 9.4-19.1]) among ART-experienced participants (P < 0.01). The prevalence of high-level DOR-associated resistance mutations was 33/1261 (2.6% [95% CI: 1.8\u20133.7]) among ART-na\u00efve individuals and 13/212 (6.1% [95% CI: 3.6\u201310.8]) among individuals experiencing VF on ART (P < 0.01). Of the two intermediate DOR-associated mutations reported in the study (V106M and Y188F), V106M was the most predominant mutation, 12/1473 (0.8%), and was common among individuals experiencing VF on ART 7/212 (3.3%) (P < 0.01). Seven high-level DOR-associated mutations predicted were V106A, Y188L, F227L and M230L, which occurred in 2/1473 (0.14%) participants, F227C and Y318F in 1/1473 (0.07%) each and the G190E 33/1473 (2.2%), which was the most predominant mutation. The prevalence of specific DOR-associated resistance mutations stratified by ART groups are summarized in A total of 1473 participants with viremia were analysed for DOR-associated resistance mutations. Of these, 1261 were ART-na\u00efve, whereas 212 were individuals experiencing VF on ART with HIV-1 VL >400 copies/mL. The overall prevalence of participants with DOR-associated resistance mutations was 181/1473 (12.3% [95% CI: 10.7\u201314.1]). Higher overall prevalence of DOR-associated resistance mutations was reported among individuals experiencing VF on ART, which was 42/212 (19.8% [95% CI: 14.7\u201325.4]) compared to 139/1261 (11.0% [95% CI: 9.3\u201312.9]) in the ART-na\u00efve population  were failing ART with at least one EFV/NVP-associated resistance mutation. A total of 42/67 (62.7% [95% CI: 50.0\u201374.2]) had a combination of DOR- and EFV/NVP-associated resistance mutations. The overall prevalence of intermediate DOR-associated resistance mutations was 29/67 (43.3% [95% CI: 31.2\u201356.0]) in this group, whereas the high-level DOR resistance was reported in 13/67 (19.4% [95% CI: 10.8\u201330.9]). Mutation V106M (7/67:10.4%) was the most predominant intermediate mutation, followed by Y188F. In the high-level resistance group, G190E was the most prevalent at 3/67 (4.5%), followed by equal proportions of V106A and M230L mutations at 2/67 (3.0%), and, lastly, F227C, F227L and Y318F, all occurring at a prevalence of 1/67 (1.4%) each. Individuals without EFV/NVP-associated resistance mutations had no DOR-associated resistance. Among 67 participants with at least one major EFV/NVP-associated resistance mutation, 38 had dual resistance with at least one major NRTI-associated resistance mutation. The prevalence of DOR-associated resistance mutations was higher in this group: 29/38 (76%) overall prevalence, 21 (55%) intermediate and 8 (21.1%) high-level resistance.3.410 copies/mL were not associated with DOR-associated resistance mutations in a small population of 67 participants . The prevalence of intermediate DOR resistance was 735/4517 (16.3% [95% CI: 15.2\u201317.4]) among individuals with viral suppression on ART vs 29/212 (13.7% [95% CI:9.4\u201319.1]) among individuals with VF on ART (P\u00a0=\u00a00.32) and high-level DOR resistance was 175/4517 (3.9% [95% CI: 3.3\u20134.5]) and 13/212 (6.1% [95% CI: 3.3\u201310.3]) among individuals with viral suppression and those experiencing VF on ART, respectively (P\u00a0=\u00a00.11), were not statistically different in the ART-experienced population. Mutations reported in the two groups were G190E (161/4729:3.4%), V106M (21/4729:0.4%), F227L (7/4729: 0.15%), Y188L (6/4729: 0.13%), Y318F (5/4729: 0.11%), V106A and M230L were 4/4729 (0.08%) each and F227C (4/4729: 0.04%). The prevalence of DOR-associated mutations is stratified by individuals with viral suppression and VF on ART in Supplementary Table S1.We also compared the prevalence of HIV-DRMs associated with DOR resistance among individuals with viral suppression on ART (HIV-1 VL \u2264400 copies/mL) compared to individuals experiencing VF on ART. The overall prevalence of mutations associated with DOR resistance was similar among individuals with viral suppression on ART, 910/4517 (20.1% [95% CI: 19.0\u201321.3]), compared to 42/212 (19.8% [95% CI: 14.7\u201325.8]) among those experiencing VF on ART (P < 0.01).Of all the 1091 participants with DOR-associated resistance, 230 (21.1% [95% CI: 18.7\u201323.6]) are likely to have specific DOR-associated resistance mutations and 861 (78.9% [95% CI: 76.4\u201381.3]) with nonspecific DOR-associated resistance mutations. Among 861 with nonspecific DOR-associated resistance mutations, 850 (98.7% [95% CI: 97.7\u201399.4]) and 11 (1.3% [95% CI: 0.6\u20132.3]) had intermediate and high-level DOR-associated resistance, respectively , confirming the opportunity for the use of DOR-containing regimens among PWH who are ART-na\u00efve in Botswana. However, the prevalence of DOR-associated resistance among ART-na\u00efve individuals (11.0%: 139/1261) in this study was statistically higher than the 2.9% prevalence of other NNRTI (first-generation)-associated resistance mutations previously reported in the BCPP study P < 0.01). Some studies reported higher resistance to other NNRTI-containing regimens when compared to DOR. A study from Greece, Italy and France reported a lower prevalence of DOR-associated resistance (1.4%) compared to first-generation NNRTI-associated resistance , with no association between HIV-1 subtype and presence of DOR-associated resistance mutations A significantly lower overall prevalence of DOR-associated resistance was reported among ART-na\u00efve compared to individuals experiencing VF on ART  of DOR-associated resistance mutations was among participants failing NNRTI-based therapy in our findings, this was lower when compared to 84.8% reported in a South African study The high-level DOR-associated resistance mutations G190E, F227L and Y318F all showed a similar prevalence among the three groups: ART-naive, individuals with VF, and individuals on ART with viral suppression. The similar prevalence of these mutations across the three groups highlights the importance of gaining a better understanding of their mechanisms, particularly in ART-na\u00efve and ART-suppressed individuals. It was difficult to determine if the presence of these mutations in ART-na\u00efve individuals was due to undisclosed ART use in the BCPP cohort EFV has been associated with an increased risk of developing high-level DOR-associated resistance mutations Because our study had a small number of participants on DTG-based ART, our findings cannot reveal whether DOR can be used as an alternative drug for DTG-failing participants with multi-resistance mutations. As a result, future studies should investigate DOR-associated resistance mutations in DTG-failing participants with multi-drug resistance.In conclusion, the prevalence of intermediate and high-level DOR-associated resistance mutations was low among ART-na\u00efve and ART-failing individuals, with the ART-failing group showing a statistically higher prevalence compared to the ART-na\u00efve group. Specifically, a higher prevalence was observed among PWH with EFV/NVP-associated resistance experiencing VF. Our results support the use of DOR among ART-na\u00efve patients in Botswana; however, we highly recommend HIV drug resistance testing among NNRTI-failing individuals before DOR-based regimen initiation.None declared."}
+{"text": "Scientific Reportshttps://doi.org/10.1038/s41598-022-10197-w, published online 15 April 2022Correction to: The original version of this Article contained an error in the References. The authors omitted the below Reference, which is listed below as Reference 50.et al. Adsorptive purification of CO2/H2 gas mixtures of spent disposable wooden chopstick-derived activated carbon: Optimal synthesis condition. Sep. Purif. Technol.291, 120948 (2022).50. Phadungbut, P. As a result, in the caption of Figure 1 (a),\u201cRepresentative SEM images (left) and HRTEM images (right) of the (a) biochar, (b) AC7-2 and (c) AC9-2 samples.\u201dnow reads:50, (b) AC7-2 and (c) AC9-2 samples.\u201d\u201cRepresentative SEM images (left) and HRTEM images (right) of the (a) biocharAs a result of the changes, the References have been renumbered.The original Article has been corrected."}
+{"text": "The SARS-CoV-2 Omicron variant has been rapidly spreading worldwide. We aimed to characterize Omicron severity by assessing in-hospital deaths and intensive care admissions in a large healthcare system in South Florida during an Omicron predominant surge.Laboratory-confirmed COVID-19 adult patients hospitalized during January 1\u201414, 2022 were retrospectively reviewed. Risks of in-hospital mortality and intensive care admission were estimated using logistic regression models. Analyses were stratified by age \u2265 65 years and vaccination status, and further adjusted for sex, comorbidities, and history of a previous COVID-19 infection.p< 0.001), with the median time from hospital admission to death being 13 days  . Patients aged \u2265 65 years had 2.6 times higher rates for in-hospital mortality  than those aged < 65 years, but were comparable for ICU admission . Past vaccination offered no protection against in-hospital mortality  or ICU admission  . In multivariable-adjusted models, patients aged \u2265 65 years had a higher in-hospital mortality than those aged < 65 years .500 consecutively hospitalized COVID-19 Omicron patients were included. The median age was 69  years, and 271 (54.2%) were women. The most common comorbidities were hypertension (65.5%), diabetes (32%), and chronic kidney disease (24%). 260 (52%) patients were fully vaccinated (defined as a patient who received 2-dose vaccines), and 32 (6.4%) were previously infected with COVID-19. 252 (50.4%) patients required supplemental oxygen, 54 (10.8%) required intensive care unit (ICU) admission, and 44 (8.8%) patients required mechanical ventilation. At study closeout of March 7, 2022, case fatality rates among patients aged 18\u201329 years, 30\u201339 years, 40-49 years, 50-59 years, 60-69 years, 70-79 years, and \u2265 80 years were 0%, 2.2%, 6.4%, 5.3%, 8.0%, 5.7%, and 15.4% respectively (This case series provides characteristics and outcomes of hospitalized adult patients with COVID-19 Omicron variant. Past COVID-19 vaccination did not impact ICU admission rate nor in-hospital mortality.All Authors: No reported disclosures."}
+{"text": "To explore why these standard treatments fail for some patients, we evaluated whether the variation in HPV oncoprotein levels among HPV+ OPSCCs affects mitochondrial metabolism, a source of antioxidant capacity. In cell line and patient-derived xenograft models, levels of HPV full-length E6 (fl-E6) inversely correlated with oxidative phosphorylation, antioxidant capacity, and therapy resistance, and fl-E6 was the only HPV oncoprotein to display such correlations. Ectopically expressing fl-E6 in models with low baseline levels reduced mitochondrial mass, depleted antioxidant capacity, and sensitized to therapy. In this setting, fl-E6 repressed the peroxisome proliferator\u2013activated receptor gamma co-activator 1\u03b1/estrogen-related receptor \u03b1 (PGC-1\u03b1/ERR\u03b1) pathway for mitochondrial biogenesis by reducing p53-dependent PGC-1\u03b1 transcription. Concordant observations were made in 3 clinical cohorts, where expression of mitochondrial components was higher in tumors of patients with reduced survival. These tumors contained the lowest fl-E6 levels, the highest p53 target gene expression, and an activated PGC-1\u03b1/ERR\u03b1 pathway. Our findings demonstrate that E6 can potentiate treatment responses by depleting mitochondrial antioxidant capacity and provide evidence for low E6 negatively affecting patient survival. E6\u2019s interaction with the PGC-1\u03b1/ERR\u03b1 axis has implications for predicting and targeting treatment resistance in OPSCC.Therapy with radiation plus cisplatin kills HPV  PGC-1\u03b1 (PGC-1\u03b1) . Our fin+ OPSCCs with survival outcomes annotation. In The Cancer Genome Atlas (TCGA), RNA-Seq data are available from 53 OPSCCs that express high-risk HPV transcripts  and intermediate tertiles (P = 0.043) (+ OPSCC cohort (n = 47) into tertiles by the identical methodology used for TCGA cases (P = 0.048). Wide variation in therapy was apparent from characteristics of the JHU and TCGA cohorts (https://doi.org/10.1172/jci.insight.159600DS1), making the relationship between survival and response to radiation plus chemotherapy unclear. Thus, we examined a subcohort of patients within the broader VU HPV+ OPSCC cohort (n = 37). Despite small sample size, dividing these cases into tertiles revealed better 3-year OS (P = 0.041) in the lowest versus the highest tertile  . This fiC cohort . DividinGA cases  showed bC cohort  who rece tertile . This as+ OPSCC PDX models previously established by us from treatment-naive patients  mice showed a positive linear correlation with mitochondrial mass defined by MT-CO1/B2M ratio  using the Cell Mito Stress Test on the Seahorse XF platform. A strong positive correlation was shown between mitochondrial mass and basal OCR in the cell lines  and cell line models for prediction of increased mitochondrial mass, mitochondrial function, and cisplatin response. RNA-Seq was performed for the 7 HPVpatients . Upregulrd curve    . The hig E6, E7) . This as E6, E7) . No othe E6, E7) . Althougial mass , whereasial mass . Similarll lines , whereasll lines . These f+ head and neck cancer lines with highest mitochondrial mass and lowest fl-E6 (SDHB (complex II), UQCRC2 (complex III), MTCO2P12 (complex IV), ATP5F1A (complex V), ACADM (acyl-CoA dehydrogenase medium chain-1), and MDH1   were reduced by fl-E6 overexpression (tert/1 keratinocytes expressing E7 (N-tert/E7 keratinocytes)  , and theenase-1) . Similarpression . To detenocytes) . Three bnocytes) , F and Gnocytes) , H and I-CO1/B2M  and basa-CO1/B2M . Increas and Broad GDAC Firebrowse repository. A total of 53 HPVpression . RNA-Seqescribed . RNA-SeqU cohort .n = 53) were aligned to HPV genomes using Bowtie2 and samples with alignment selected and identified as containing HPV 16, HPV 33, HPV 35, or HPV 56 genomes. Gene expression values were represented by the normalized reads of reads per kilobase of transcript per million mapped reads. HPV genome sequences and annotations with gene coordinates were obtained from RefSeq (https://www.ncbi.nlm.nih.gov/refseq/). Reads with mapped coordinates corresponding to regions specific for each gene were counted. Fl-E6 expression was quantitated in a region nonoverlapping with E6*. The probe region specific to E6* overlaps fl-E6 and thus represents total E6. E6* is calculated by subtracting fl-E6 from total E6. HPV 16 gene expression analysis used GenBank K02718.1, with the following gene coordinates for quantitation: E6: 227-408, total E6: 83-226, E7: 562-858, E2: 2814-3331, E4: 3332-3619, and E5: 3863-4099. HPV 33 gene expression analysis used GenBank M12732.1, with the following gene coordinates for quantitation: E6: 232-413, total E6: 109-230, E7: 573-866, E2: 2814-3325, E4: 3326-3577, and E5: 3854-4081. HPV 35 gene expression analysis used GenBank X74477.1, with the following gene coordinates for quantitation: E6: 233-414, total E6: 110-231, E7: 562-861, E1: 868-2713, E2: 2882-3293, E4: 3294-3584, and E5: 3814-4065. HPV 56 gene expression analysis used GenBank: X74483.1, with the following gene coordinates for quantitation: E6: 234-415, total E6: 102-232, E7: 572-889, E2: 2806-3221, E4: 3222-3577, and E5: 3854-4081.RNA-Seq reads within FastQ files for each TCGA OPSCC sample . Ribosomal RNA depletion was performed with Ribo-zero rRNA Removal Kit, HMR (Illumina Inc.). Library preparation was performed with the NEBNext Ultra II Non-directional Synthesis Module (New England Biolabs). Samples were sequenced on Illumina NextSeq High Output sequencer and aligned to combined genome from hg38 (human), mm10 (mouse), and the high-risk HPV genomes \u2014 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68 \u2014 using STAR with default parameters. HPV sequences in all samples mapped to HPV 16 exclusively. After STAR first-pass alignment, reads mapped to mm10 were removed, and reads aligned to human genome (hg38) and HPV genomes were retained. Raw counts for each viral transcript  were obtained from reads mapped to each gene based on the alignment file. Fl-E6 was computed as the ratio of the average coverage level in the first intron (approximate full-length E6 level) divided by the average coverage level in the first exon . RnaSTAR was used to identify the first introns for E6*, which are, in the format of donor-acceptor, 227-408 for HPV 16. The average coverage of read pairs within the intron  was divided by the average coverage of read pairs within exon1 , where coverage was computed by dividing the number of reads by the feature width. RNA-Seq data reported here were submitted to National Center for Biotechnology Information Gene Expression Omnibus (GSE193388).The 7 treatment-naive HPVNSG mice . The PDX\u2013/\u2013 (RRID:CVCL_HD97) cells are from ATCC. UM-SCC047 (RRID:CVCL_7759) and UMSCC104 (RRID:CVCL_7712) are from Sigma-Aldrich. UDSCC2 (RRID:CVCL_E325) and 93VU147T (RRID:CVCL_L895) cell lines were gifts from Silvio Gutkind  and Hans Joenje , respectively. Cell lines were authenticated using the Identify Mapping Kit (Coriell). They were cultured in DMEM-F12 (Gibco), 10% heat-inactivated FBS, 1% penicillin-streptomycin, and 1% nonessential amino acids. N/tert-1/E7 cells were cultured in keratinocyte serum-free medium (Thermo Fisher Scientific).SCC090 (RRID:CVCL_1899), SCC152 (RRID:CVCL_0113), SCC154 (RRID:CVCL_2230), HEK293(RRID:CVCL_0045), HCT116 (RRID:CVCL_0291), and HCT116 p53Total RNA was extracted using the RNeasy Plus Micro Kit (QIAGEN). DNase I  was used to remove genomic DNA from RNA samples. Reverse transcription was performed with oligo-dT plus random decamer primers using Superscript II (Thermo Fisher Scientific). QRT-PCR was performed with SYBR green master mix (Thermo Fisher Scientific) using Step-one Plus Real-Time PCR. Primers are in Fl-E6 was expressed transiently using the pLentiN-16E6no* plasmid (RRID:Addgene_37445) and its pLentiN vector control (RRID:Addgene_37444). Transient PGC-1\u03b1 overexpression was achieved with the pcDNA4 PGC-1\u03b1 plasmid (RRID:Addgene_10974). A total of 4 \u03bcg of each plasmid was delivered per well in 6-well plates using Lipofectamine 2000 (Thermo Fisher Scientific) and incubated for 72 hours. SiRNA silencing of PGC-1\u03b1 was performed using the mix of the 3 validated targeting siRNAs and scrambled negative control included in the PPARGC1A TriFECTa DsiRNA Kit (IDT) per supplier instructions. SiRNA was delivered at 50 nM concentration per well in 6-well plates using Lipofectamine RNAiMax (Thermo Fisher Scientific) and incubated for 72 hours.tert-E7 cells were similarly infected with the control and E6 viruses before blasticidin selection, propagation, and analysis of bulk cultures.The pLentiN-16E6no* plasmid containing full-length E6 and its pLentiN vector backbone control were used for stable fl-E6 expression. Lentiviral preparation and transduction were performed as described . BrieflyHPV 16 E6 mutant constructs 16E6, 16E6 (V42L), 16E6-8S9A10T, 16E6-I128T, and 16E6* are previously described . The pGLp-trifluoro-methoxyphenyl hydrazone, and 2 \u03bcM each of antimycin and rotenone (Sigma-Aldrich). Total protein per well was determined afterward with the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). Results were analyzed using XFe Wave software (Seahorse Bioscience), and 1 \u00d7 105 cells were used to measure NADPH and NADP+ levels using an NADP+/NADPH colorimetric quantitation kit (Sigma-Aldrich).Mitochondrial DNA content was determined by qPCR as described . OCR wasCell viability after 72-hour treatment with cisplatin, DMNQ, MnTMPyP, and/or MitoTEMPO (Sigma-Aldrich) was determined by WST-1 and/or BrdU assays (Sigma-Aldrich). Cisplatin dissolved in saline was administered intraperitoneally in vivo at 2 mg/kg. For irradiation, cells were harvested during exponential growth and plated at 2,000 cells/mL in 60 mm dishes containing 2 mL media. After 12 hours, they received 2, 4, or 6 Gy using a Gammacell 40 irradiator (Best Theratronics). Media were changed postirradiation. Colonies were stained with crystal violet (Thermo Fisher Scientific) and counted 10 days later.t tests. Pearson correlation coefficients were based on analyzing the means. Two-way ANOVA was used when comparing xenograft growth curves. Log-rank tests were used for Kaplan-Meier analyses. P values were calculated for multiple comparisons using a 1-way ANOVA corrected with Dunnett\u2019s or Holm-\u0160id\u00e1k procedure. Tests used are indicated in figure legends. Analyses were performed using Prism (GraphPad Software). A P value less than 0.05 was considered significant.When comparing 2 groups for cell viability or gene expression, significance was calculated using 2-tailed unpaired Mouse experiments were performed under Wistar Institute IACUC protocols 201166 and 201178 . PDXs were previously established under University of Pennsylvania IRB-approved protocol 417200 \u201cHead and Neck Cancer Specimen Bank\u201d  by signing a combined informed consent and HIPAA form for use of tissue for research. All clinical data shown are obtained from deidentified, publicly available data sets.MKS and DB conceived the study. DB, MKS, EAW, IMM, and BEW wrote and edited the manuscript and/or designed figures and tables. MKS, PR, LL, and VS performed experiments. PAG, PR, MKS, XW, XL, and BEW performed bioinformatic and/or statistical analyses. DB, EAW, PAG, DPK, and HN supervised the study and data analysis. DB, JBJ, RMB, XW, and XL analyzed clinical and/or pathologic data. All authors read and approved the final manuscript."}
+{"text": "H)-one represents a new template for AKA inhibitors, with antiproliferative activity against cancer cells. A quinazolin-4(3H)-one derivative was further designed and synthesized in order to improve the pharmacokinetic properties and antiproliferation activity against NSCLC cell lines. The derivative, BIQO-19 (Ethyl 6-(4-oxo-3-(pyrimidin-2-ylmethyl)-3,4-dihydroquinazolin-6-yl)imidazo pyridine-2-carboxylate), exhibited improved solubility and antiproliferative activity in NSCLC cells, including epidermal growth factor receptor\u2013tyrosine kinase inhibitor (EGFR-TKI)-resistant NSCLC cells. BIQO-19 effectively inhibited the growth of the EGFR-TKI-resistant H1975 NSCLC cells, with the suppression of activated AKA (p-AKA) expression in these cells. The inhibition of AKA by BIQO-19 significantly induced G2/M phase arrest and subsequently evoked apoptosis in H1975 cells. In addition, the combination of gefitinib and BIQO-19 exhibited synergistic antiproliferative activity in NSCLC cells. These findings suggest the potential of BIQO-19 as a novel therapeutic agent for restoring the sensitivity of gefitinib in EGFR-TKI-resistant NSCLC cells.Non-small cell lung cancer (NSCLC) is the most common lung cancer subtype. Although chemotherapy and targeted therapy are used for the treatment of patients with NSCLC, the survival rate remains very low. Recent findings suggested that aurora kinase A (AKA), a cell cycle regulator, is a potential target for NSCLC therapy. Previously, we reported that a chemical entity of quinazolin-4(3 Lung cancer is the leading cause of death from cancer, and constitutes almost 25% of all cancer-related deaths . Lung caAurora kinase is a serine/threonine kinase which plays an important role in cell cycle regulation . The higH)-one is a well-known core heterocycle pharmacophore used in medicinal chemistry that exhibits a variety of biological activities, including sedative, antiviral, antibacterial, anti-inflammatory, and anticancer effects pyridine-2-carboxylate) as an FL-4 derivative (H)-one derivative FL-4 +. Compound 5, 6-(6-aminopyridin-3-yl)-3-(pyrimidin-2-ylmethyl)quinazolin-4(3H)-one, was obtained as a white solid, yield 83.4%, ESI-MS: m/z 331.1 [M + H]+. BIQO-19 was separated as white crystals, yield 87.4%, m.p. 284\u2013286 \u00b0C. 1H-NMR  \u03b4 8.68 , 8.49 , 8.41 , 8.24 , 8.21 , 7.96 , 7.86 , 7.76 , 7.57 , 7.23 , 5.45 , 4.46 , 1.44 . 13C-NMR  \u03b4 164.50, 163.17, 161.06, 157.58, 148.15, 147.78, 144.54, 137.61, 135.54, 132.70, 128.76, 126.98, 126.59, 125.0, 123.8, 122.9, 120.2, 119.3, 117.6, 61.4, 51.7, 14.5. HRMS (ESI-QTOF) m/z Calcd. For C23H19O3N6 [M + Na]+ m/z: 449.1333, found: 449.1324. NMR and high resolution mass spectras are shown in The synthesis of the compound BIQO-19 is summarized in Culture media, fetal bovine serum, antibiotic-antimycotic solution , and trypsin-EDTA solution (1\u00d7) were purchased from Gibco . Gefitinib (CAS No. HY-50895) was purchased from MedChemExpress . Trichloroacetic acid (TCA), sulforhodamine B (SRB), crystal violet solution, and bovine serum albumin (BSA) were purchased from Sigma-Aldrich . Anti-aurora kinase A (#14475), anti-p-aurora kinase A (T288) (#3079), anti-p21 Waf1/Cip1 (#2947), anti-cdc2 (#9116), anti-cdc25C (#4688), anti-PARP (#9532), anti-Cleaved PARP (D214) (#5625), anti-Caspase-8 (#9746), and anti-Cleaved Caspase-8 (D374) (#9496) antibodies were obtained from Cell Signaling Technology .The ADMET properties of the FL-4 and BIQO-19 compounds were calculated with the ADMET descriptors of Discovery Studio 3.0. The ADMET descriptors parameter was set as the default. Each of the ADMET profile levels were categorized by following the criteria. ADMET_Solubility_Level: 0  < \u22128.0), 1  < \u22126.0), 2  < \u22124.1), 3  < \u22122.0), 4  \u2264 0.0), 5 ), 6 (warning: molecules with one or more unknown AlogP98 types). ADMET_BBB_Level: 0 , 1 , 2 , 3 , 4 (undefined).2.The human non-small lung cancer cell lines A549, NCI-H1975, NCI-H1299, and NCI-H1993 were purchased from the American Type Culture Collection . The human non-small cell lung cancer cell lines HCC827, PC9, HCC827-gef and PC9-gef were a kind gift from Dr. Jae Cheol Lee and Dr. Jin Kyung Rho . HCC827-gef and PC9-gef cells were subcultured in the presence of 1\u2009\u00b5M gefitinib. All cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 1% antibiotic-antimycotic. Cells were incubated at 37 \u00b0C in a humidified atmosphere with 5% COw/v) SRB in 1% acetic acid for 2 h at room temperature. The stained proteins were dissolved in 10 mM Tris solution (pH 10) and measured the absorbance at 515 nm with a VersaMax ELISA microplate reader .Cell viability was measured using the SRB method as described previously . BrieflyCells (500 cells/well) were seeded and cultured overnight. The compounds were treated for 48 h, the cells were washed with phosphate buffered saline (PBS), and continued incubation. Culture medium was replaced every three days for ten days. Cells were fixed with 4% paraformaldehyde solution and stained with 0.1% crystal violet.http://www.kmplot.com/lung, accessed on 6 April 2022). A total of 1925 lung cancer patient gene expression values and survival data were analyzed using multiple microarray datasets [The OS of patients with lung cancer and adenocarcinoma with different expression levels of Aurora Kinase A was analyzed through a Web-based meta-analysis Kaplan\u2013Meier plotter . Recombinant human aurora A protein (Active) (ab271368) and Recombinant human aurora B protein (Active) (ab51435) were purchased from Abcam . Myelin basic protein was selected as a substrate and staurosporine was used as a positive control. The ADP-Glo The cells were collected and lysed in sample loading buffer . Total protein concentrations were quantified using a BCA Protein Assay Kit . Quantified proteins were loaded onto SDS-polyacrylamide gels and separated by molecular weight via electrophoresis (PAGE). Then, proteins were transferred to polyvinylidene fluoride membranes ; 5% BSA in Tris-buffered saline containing 0.1% Tween-20 (TBST) was used for blocking. The membranes were blocked for 30 min at room temperature with mild shaking, then incubated with antibodies diluted in 2.5% BSA in TBST overnight at 4 \u00b0C with mild shaking. Following secondary antibody incubation and washing with TBST, the chemiluminescence signals were detected using the WEST-Queen detection system  and LAS-4000 .A molecular simulation was carried out using the Molecular Operating Environment . BIQO-19 was built in MOE using the Builder program and energy-minimized for docking simulation. The 3D structure of aurora A (PDB ID: 6C2T) was obtain from the PDB databank and optimized, 3-D protonated, and energy-minimized using the default parameters of MOE. The docking site was defined by the Site Finder application, including the residues within 10 \u00c5 of the cocrystallization ligand. The docking results were analyzed using Accelrys Discovery Studio Visualizer 4.5.TM Pro version 6.0 software .Cells were seeded and starved in serum-free medium overnight. After starvation, the cells were treated with compounds for the indicated times. After incubation, cells were harvested, washed with PBS, and fixed with 70% ethanol in PBS overnight at \u221220 \u00b0C. The fixed cells were washed with PBS, resuspended in 100 \u00b5g/mL of RNase A for 30 min at room temperature, and stained with 50 \u00b5g/mL PI. Fluorescence intensity was analyzed using a FACSCalibur flow cytometer  and BD CellQuestTM Pro version 6.0 software .Cells were treated with samples for 72 h and stained with an Annexin V-FITC apoptosis detection kit  according to the manufacturer\u2019s instructions. Briefly, cells were collected after 72 h treatment, washed with PBS, resuspended in 1\u00d7 binding buffer, and stained with Annexin V-FITC and PI in the dark for 15 min. After incubation, the cells were diluted with 1\u00d7 binding buffer and analyzed using an FACSCalibur flow cytometer  and BD CellQuestp < 0.05, ** p < 0.01, *** p < 0.001) was calculated using one-way analysis of variance coupled with Dunnett\u2019s t-test or Student\u2019s t-test.Data were obtained from three independent experiments, and are shown as the means \u00b1 standard deviation. Statistical significance -one derivative, BIQO-19, which is an aurora kinase A inhibitor that exhibits an improved ADMET profile and antiproliferative activity in NSCLC cells. The mechanisms of action for the antiproliferative activities of BIQO-19 in EGFR-TKI-resistant NSCLC H1975 cells involve aurora kinase A-mediated arrest at the G2/M phase of the cell cycle and the induction of apoptosis. Combined treatment with BIQO-19 and gefitinib exhibits synergetic activity with respect to inhibiting proliferation and inducing apoptosis in H1975 cells. Therefore, targeting aurora kinase A using a new class of inhibitors in combination with EGFR-TKIs represents a potential therapeutic strategy for treating patients with EGFR-TKI-resistant NSCLC."}
+{"text": "The geometric mean end-point titer (GMT) of anti-RBD-IgG in PLWH was also reduced, especially in patients with CD4 counts <200 cells/\u00b5L, regardless of age, gender, or HIV viral load. GMTs of anti-RBD-IgG in both PLWH and healthy controls declined gradually over time. Similar results were also observed in the anti-spike-IgG response. The frequency of RBD-specific MBCs in PLWH decreased (p<0.05), and then remained stable over time. Lastly, through multivariate analysis, we found the factors that predicted a less robust response to inactivated vaccines in PLWH were a low CD4 count and long time interval after vaccination. In conclusion, inactivated vaccines are well-tolerated in PLWH but with low immunogenicity. Therefore, SARS-CoV-2 vaccines and booster doses should be given priority in PLWH, especially in patients with low CD4 counts.It is important to know the safety and efficacy of vaccination in immunocompromised people living with HIV (PLWH), but currently, there is limited data on the inactivated SARS-CoV-2 vaccines\u2019 safety and immune responses in PLWH. In this prospective observational study, 139 PLWH and 120 healthy controls were enrolled and monitored for 21\u2013105 days after a two-dose vaccination. The safety, anti-receptor binding domain IgG (anti-RBD-IgG) and anti-spike-IgG responses, and RBD-specific memory B cell (MBC) responses were evaluated. The overall adverse events within seven days were reported in 12.9% (18/139) of PLWH and 13.3% (16/120) of healthy controls. No serious adverse events occurred in both groups. Overall, the seroprevalence of anti-RBD-IgG in PLWH was significantly decreased (87.1% vs. 99.2%; Trial registration:ClinicalTrials.gov identifier: NCT05043129.. The coronavirus disease 2019 (COVID-19) epidemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) began in 2019 and has continued to rage worldwide, bringing a heavy burden to global public health . Even th+ T cell loss after receiving an inactivated COVID-19 vaccine in a treatment-na\u00efve HIV-positive patient was reported by a recent study  vs. 278.6 [95% CI: 204.1\u2013380.4], p<0.05; 3 months: 93.3 [95% CI: 68.66\u2013126.8] vs. 150.8 [95% CI: 109.9\u2013206.8], p<0.05; 2 AU/mL, IQR [1.67\u20132.90 log2 AU/mL] vs. 2.64 log2 AU/mL, IQR [2.07\u20133.94 log2 AU/mL], p<0.01; 2 months: 1.42 log2 AU/mL, IQR [1.23\u20134.63 log2 AU/mL] vs. 2.17 log2 AU/mL, IQR [1.88\u20132.86 log2 AU/mL], p\u2009=\u20090.169; 3 months: 1.65 log2 AU/mL, IQR [1.38\u20132.01 log2 AU/mL] vs. 1.78 log2 AU/mL, IQR [1.21\u20132.29 log2 AU/mL], p\u2009=\u20090.786; To better understand the variation of humoral immune responses with passing time, we stratified three groups by time interval after full-course vaccination in the cross-sectional analysis. As expected, GMTs of anti-RBD-IgG gradually decreased over time in both PLWH and healthy controls. However, GMTs of anti-RBD-IgG in PLWH were sharply lower than that of healthy controls at every time point (1 month: 155.3 [95% CI: 128.3\u2013188.1] vs. 441.6 [95% CI: 355\u2013549.5], In summary, the weakened antibody response in PLWH kept on falling over time, whereas the impaired MBC response did not change over time.p<0.001; 2 AU/mL, IQR [1.72\u20132.98 log2 AU/mL] vs. 1.61 log2 AU/mL, IQR [1.41\u20132.03 log2 AU/mL]; p<0.001; To further explore the dynamic changes of antibody levels in PLWH, a longitudinal analysis was conducted in PLWH. Of the 96 PLWH observed after 1 month, 52 were followed up to the 6th month. As expected, GMTs of anti-RBD-IgG showed a clear downward trend in the 6th month compared with the 1st month (219.6 [95% CI: 179.3\u2013268.8] vs. 97.37 [95% CI: 83.99\u2013112.9]; Lastly, we wanted to investigate the risk factors related to inferior response to anti-RBD-IgG in PLWH. As shown in In this prospective study, we evaluated the safety, antibody responses, and RBD-specific MBC responses of inactivated SARS-CoV-2 vaccines in PLWH and healthy controls. Our results showed that inactivated vaccines were safe and well-tolerated in PLWH. The antibody and MBC responses waned in PLWH, especially in PLWH with CD4 counts <200 cells/\u00b5L. Therefore, PLWH should be vaccinated ahead of the healthy population.PLWH with SARS-CoV-2 have poor clinical outcomes, especially those who are immunosuppressed or do not receive ART ,19. HoweOur results showed that PLWH had lower anti-RBD-IgG and anti-spike-IgG titers than healthy controls 21\u2013105 days after vaccination, which is consistent with previous studies showing that PLWH had lower immune responses to the vaccine than healthy individuals . Previou+ T cells and a series of immune abnormalities [The level of immunosuppression is usually reflected by the CD4 cell counts . Usuallymalities . Howevermalities . In thismalities ,11. Thismalities . Of notemalities , hence, The MBCs produced during primary infection are quickly reactivated after a secondary infection, thus, preventing severe disease or death ,15. HencThere are some limitations in this study. First, few subjects participated in the follow-up until month 6 after full vaccination because the sporadic localized outbreaks of COVID-19 partially impeded travel. Second, the early stages of B and T cell responses were not evaluated, as we were mainly focusing on the significance of durable humoral immune responses. Therefore, further studies about the early stages of B and T cell responses in PLWH are needed. Third, the best correlation of antibody titers and vaccine efficacy in PLWH is currently unknown, and more large-scale population studies are needed. Nonetheless, we believe our results are particularly important and meaningful to clinicians.In conclusion, the inactivated SARS-CoV-2 vaccines are safe and well-tolerated in people living with HIV, with no serious adverse events reported. However, the antibody response and RBD-specific MBC response were weak, especially in PLWH with a low CD4 count. Therefore, SARS-CoV-2 vaccines and booster doses should be prioritized for this special population.Click here for additional data file."}
+{"text": "Trichoderma strains isolated from forest species of the Cerrado-Caatinga ecotone as biological control agents of crop pathogenic fungi. Nineteen Trichoderma strains were used to assess the antagonistic activity by in vitro bioassays against the plant pathogens Colletotrichum truncatum, Lasiodiplodia theobromae, Macrophomina phaseolina, and Sclerotium delphinii isolated from soybean, cacao, fava bean, and black pepper crops, respectively. All Trichoderma strains demonstrated inhibitory activity on pathogen mycelial growth, with maximum percent inhibition of 70% against C. truncatum, 78% against L. theobromae, 78% against M. phaseolina, and 69% against S. delphinii. Crude methanol extracts (0.5 to 2.0 mg mL-1) of Trichoderma strains were able to inhibit the growth of C. truncatum, except Trichoderma sp. T3 (UFPIT06) and T. orientale (UFPIT09 and UFPIT17) at 0.5 mg mL-1, indicating that the endophytes employ a biocontrol mechanism related to antibiosis, together with multiple mechanisms. Discriminant metabolites of Trichoderma extracts were unveiled by liquid chromatography-tandem mass spectrometry-based metabolomics combined with principal component analysis (PCA), which included antifungal metabolites and molecules with other bioactivities. These results highlight the biocontrol potential of Trichoderma strains isolated from the Cerrado-Caatinga ecotone against crop pathogenic fungi, providing support for ongoing research on disease control in agriculture.The indiscriminate use of chemical pesticides increasingly harms the health of living beings and the environment. Thus, biological control carried out by microorganisms has gained prominence, since it consists of an environmentally friendly alternative to the use of pesticides for controlling plant diseases. Herein, we evaluated the potential role of endophytic  Diseases caused by fungi are among the most harmful to plants due to their rapid spread and the ability to adapt to various environmental conditions . The mosResearch on alternative methods of controlling fungal diseases has been widely developed, often requiring the integrated implementation of several methods, known as integrated disease management (IDM) , 4. AmonA promising alternative method for plant pathogen control is based on the use of antagonistic microorganisms, such as endophytic fungi, capable of protecting their hosts from the action of pathogens , 9. ThesRecent research has revealed that although endophytic microorganisms have received global interest, there are still several gaps in knowledge, such as the different biomes explored . ConsideThe Cerrado and Caatinga biomes are recognized for their great importance. The Cerrado, also known as the Brazilian savanna, is one of the 25 biodiversity hotspots for conservation priorities in the world , 17, andTrichoderma species have been tried as BCA and used as an alternative to synthetic pesticides to control a variety of plant diseases + was identified as trigonelline (T. longibrachiatum (UFPIT02), Trichoderma sp. T2 (UFPIT05), T. koningiopsis (UFPIT07), Trichoderma sp. T5 (UFPIT11), Trichoderma sp. T6 (UFPIT13), and T. koningiopsis (UFPIT19) isolates. The MS/MS spectrum was characterized by the fragment of m/z 92.0496 [__HCOOH]+, referring to the carboxylic acid group +, identified as phomalone , T. longibrachiatum (UFPIT02), and T. orientale  isolates was identified as columbianetin  and Trichoderma sp. T6 (UFPIT13) isolates. The metabolite of m/z 419.1713 detected in T. orientale (UFPIT01), T. longibrachiatum (UFPIT02), Trichoderma sp. T6 (UFPIT13), and T. orientale  isolates were identified as syringaresinol , was identified as brefeldin-A (BFA) . The metabolite of m/z 453.1914 was identified as physodic acid , was identified as HT-2 toxin  and T. longibrachiatum (UFPIT02) isolates. The metabolite of m/z 163.0393 detected in all Trichoderma spp. was identified as 4-hydroxycoumarin , L. theobromae (B), M. phaseolina COUFPI 10 and COUFPI 11 (C), and S. delphinii COUFPI 209 and COUFPI 249 (D) paired with Trichoderma strains. Means followed by the same letter do not differ from each other by the Scott\u2013Knott test at the 5% probability level. The coefficients of variation (CVs) were 20.51% for L. theobromae, 15.99% for M. phaseolina COUFPI 10, 19.13% for M. phaseolina COUFPI 11, 10.68% for S. delphinii COUFPI 209, and 9.54% for S. delphinii COUFPI 249. Different lowercase letters indicate a significant difference between Trichoderma spp. Different capital letters indicate a significant difference between Trichoderma spp.Mycelial growth rate index (MGRI) of (TIF)Click here for additional data file.S2 Fig(TIF)Click here for additional data file.S3 Fig(TIF)Click here for additional data file.S4 Fig(TIF)Click here for additional data file.S5 Fig(TIF)Click here for additional data file.S6 Fig(TIF)Click here for additional data file.S7 Fig(TIF)Click here for additional data file.S8 Fig-1, 8.45% for 1.0 mg mL-1 and 9.36% for 2.0 mg mL-1.The coefficients of variation (CVs) were 4.65% for the concentration of 0.5 mg mL(TIF)Click here for additional data file.S9 FigTrichoderma spp. cultures generated using MetaboAnalyst. Con = Control, UFPIT01 = T1, UFPIT02 = T2, UFPIT03 = T3, UFPIT04 = T4, UFPIT05 = T5, UFPIT06 = T6, UFPIT07 = T7, UFPIT08 = T8, UFPIT09 = T9, UFPIT10 = T10, UFPIT11 = T11, UFPIT12 = T12, UFPIT13 = T13, UFPIT14 = T14, UFPIT15 = T15, UFPIT16 = T16, UFPIT17 = T17, UFPIT18 = T18, and UFPIT19 = T19.PC1 x PC2 (A) and PC1 x PC3 (B) loading plots of metabolic fingerprints of (TIF)Click here for additional data file.S10 Fig(TIF)Click here for additional data file.S11 Fig(TIF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file."}
+{"text": "Q2 and Q4 of non-HDL-C were associated with increased odds of SICH  compared to Q3. Q1 and Q2 of LDL-C was associated with increased odds of mortality  compared to Q3. In AIS patients who received IV tPA, low LDL-C was associated with increased odds of mortality while HDL-C may be protective against poor functional outcome.Contradicting evidence exists regarding the role of lipids in outcomes following intravenous (IV) thrombolysis with tissue plasminogen activator (tPA). Restricted cubic spline curves and adjusted logistic regression were used to evaluate associations of low-density lipoprotein cholesterol (LDL-C), non-high-density lipoprotein cholesterol (non-HDL-C), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C) and LDL-C/HDL-C ratio with poor functional outcome, symptomatic intracranial hemorrhage (SICH) and 90-day mortality, among 1004 acute ischemic stroke (AIS) patients who received IV tPA in a comprehensive stroke center. Quartile (Q) 1, Q2 and Q3 of HDL-C were associated with increased odds of poor functional outcome (adjusted odds ratio (adjOR) 1.66, 95% CI 1.06\u20132.60,  Stroke incidence and stroke-related mortality increased substantially from 1990 to 2019, and the stroke burden will continue to increase globally, especially in underdeveloped countries ,4, with In this study, we included consecutive patients who received IV tPA from September 2006 to June 2018. All these patients had no contraindications to IV tPA use. The study obtained ethics approval from the National Healthcare Group-Domain Specific Review Board (NHG DSRB Ref: 2010/00509). Patients were assessed by a neurologist for eligibility to receive intravenous thrombolysis according to institutional protocol and American Heart Association/American Stroke Association guidelines at a standard dose of 0.9 mg/kg body weight [LDL-C was calculated by the Friedewald equation (TC-HDL-C-TG/5) , while sThe primary outcome measured was poor functional outcome  of 3\u20136). Secondary outcomes measured were symptomatic intracranial hemorrhage (SICH) and 90-day all-cause mortality. SICH was based on the European Cooperative Acute Stroke Study (ECASS) II definition . The 90-p < 0.05.Analyses were performed using SPSS for Windows version 27.0  and R version 4.0.5. Restricted cubic splines with 5 knots were plotted to visually assess the univariate associations between the lipid parameters and the three outcomes. These served qualitative and descriptive purposes only. Since non-linear (U-shaped/inverse U-shaped/J-shaped/reverse tick) associations were found, lipid parameters were divided into quartiles for analysis. Descriptive statistics for continuous variables were presented as mean (SD) when normality and homogeneity assumptions were satisfied, otherwise as median (interquartile range) (IQR), and n (%) for categorical variables. Differences in continuous variables were assessed using 2 sample t-test when normality and homogeneity assumptions were satisfied; otherwise, Mann-Whitney U test was used where data was not distributed normally. Chi-square or Fisher\u2019s exact test was used for categorical variables. Covariates that were selected a priori for variable adjustment were gender, age, hypertension, atrial fibrillation, large vessel occlusion, diabetes mellitus, admitting NIHSS and admitting SBP. Logistic regression assessed the associations between lipid levels and outcomes, and with LAA. Results were presented as adjusted odds ratio (adjOR) with 95% confidence interval (CI). Statistical significance was set at two-sided This section may be divided by subheadings. It should provide a concise and precise description of the experimental results, their interpretation, as well as the experimental conclusions that can be drawn.1265 consecutive patients with ischemic stroke treated with IV tPA were analyzed and 1004 patients with valid lipid assessments were included in this study. Of 1004 patients, 589/986 (59.7%) were male, 586/883 (66.4%) were of Chinese ethnicity, 596/916 (65.1%) experienced a LVO and 190/1004 (18.9%) had AF. There were 657/1004 (65.4%) patients with history of with hypertension, 526/1004 (52.4%) with hyperlipidemia, and 306/1004 (30.5%) with diabetes mellitus. Median age was 66 (IQR 56\u201377) years while median admitting NIHSS was 15 (IQR 8\u201321). Median LDL-C, non-HDL-C, TC, HDL-C and LDL-C/HDL-C were 2.86 (IQR 2.18\u20133.50) mmol/L, 3.43 (IQR 2.70\u20134.19) mmol/L, 4.62 (IQR 3.86\u20135.36) mmol/L, 1.12 (IQR 0.95\u20131.32) mmol/L and 2.49 (IQR 1.84\u20133.30) respectively. In accordance with the TOAST classification, 322/975 (33.0%) had LAA stroke, 341/975 (35.0%) had cardioembolic (CE) stroke, 168/975 (17.2%) had small vessel occlusion (SVO), 10/975 (1.0%) had stroke of other determined etiology and 134/975 (13.7%) had cryptogenic stroke .p = 0.024) compared to Q1. Q2 and Q4 of non-HDL-C was significantly associated with increased odds of LAA  compared to Q1. Q3 of HDL-C was associated with increased odds of LAA  compared to Q4. Lastly, Q3 and Q4 of LDL-C/HDL-C were associated with increased odds of LAA  compared to Q1  count, neutrophils, platelets, admitting NIHSS and presence of LVO were significantly higher in the LAA compared to non-LAA group . Regardied to Q1 .Of 1004 patients, 479/995 (48.1%) experienced poor functional outcomes (mRS 3\u20136), 48/1003 (4.8%) suffered SICH and 117/990 (11.8%) died.  There waWhen evaluating the following relationships between lipid parameters and outcomes measured, variables adjusted for in the multivariate model include gender, age, hypertension, atrial fibrillation, large vessel occlusion, diabetes mellitus, admitting NIHSS and admitting SBP. Restricted cubic spline curves showed a \u2018reverse tick\u2019 relationship between LDL-C, non-HDL-C and TC with poor functional outcome .p = 0.028, adjOR 1.63, 95% CI 1.05\u20132.53, p = 0.027 and OR 1.56, 95% CI 1.01\u20132.44, p = 0.048 respectively) when compared to Q4. Q2 and Q4 of LDL-C/HDL-C ratio were associated increased odds of poor functional outcome  when compared to Q3  when compared to Q3. Similarly, Q2 and Q4 of TC were significantly associated with increased odds SICH  when compared to Q3. Q2 of HDL-C remained significantly associated with increased odds of SICH  when compared to Q1  when compared to Q3. Q2 of LDL-C/HDL-C was associated with increased odds of mortality  when compared to Q3. No significant associations were found to relate non-HDL-C, TC and HDL-C with mortality on multivariate analysis  compared to previous thrombolysis studies. However, this study was a single institution retrospective cohort study that solely evaluated intravenous thrombolysis patients. This may limit the generalisability of results to other cohorts, warranting more prospective studies on lipid parameters and ischemic stroke outcomes. We would like to acknowledge the possibility of Type I error in the multivariate analyses in which significant associations may no longer hold true after Bonferroni correction. Excessive correction of statistical level of significance may also increase the likelihood of Type II error as a trade-off to reduce Type I error, which may increase false negatives. The multivariate relationship between HDL-C and poor functional outcome is to be interpreted with caution as no significant relationship was found on univariate analysis, in which the possible reasons can be explained statistically by Lo et al. and Wang et al. ,39. HencIn AIS patients who received IV tPA, low LDL-C was associated with increased odds of mortality while HDL-C may be protective against poor functional outcome."}
+{"text": "Microbacterium foliorum NRRL B-24224. The genome has a length of 17,453\u2009bp and contains 25 total protein-coding genes, 20 of which were assigned functions. Based on gene content, Burgy was assigned to actinobacteriophage cluster EE.Burgy is a siphovirus that was isolated from compost soil near Fremont Township, Iowa, using  Actinobacteria have been identified throughout aquatic, soil, and animal microbiomes  , using standard methods ( plating . Transmi (n = 4) .M. foliorum (68.7%) (DNA was isolated from Burgy using the Promega Wizard DNA cleanup kit. The genome was sequenced using an Illumina MiSeq sequencer (v3 reagents) after the library was prepared using the NEBNext Ultra II FS kit, yielding 640,328 single-end 150-bp reads, which constituted ~5,500-fold coverage. Raw reads were assembled and checked for completeness using Newbler v2.9 with default parameters and Consed v29, respectively, as described previously (iorum 68.% (7). Buhttp://cobamide2.bio.pitt.edu/computer.htm), PECAAN (https://blog.kbrinsgd.org/), HHPRED (http://phages.wustl.edu/starterator), TMHMM v2.0  accession no. SRX14443487.The sequencing results for Burgy are available in GenBank with accession no."}
+{"text": "Clostridium botulinum is responsible for botulism, a potentially lethal foodborne intoxication. Here, we report the draft genome sequences of C. botulinum group II strains 202F (serotype F) and Hazen (serotype E). The genomes share many similarities, including multiple mobile genetic elements. Clostridium botulinum is an anaerobic, Gram-positive, spore-forming bacterium that produces the neurotoxin that causes botulism (C. botulinum strains are divided into four groups (I to IV)  , with moI to IV) , 5. HereC. botulinum strains 202F and Hazen were originally isolated from marine sediments in California, USA, in 1965  (Francisella sp. strain MA067296 plasmid (CP016929.1) was identified in 202F. Another plasmid  with 99% sequence homology to C. botulinum pCBI (CP006904.1)  indicative of a putative phage-like plasmid.Sequencing of strain 202F yielded 845,112 raw reads and 34 contigs with 30\u00d7 median coverage, while 629,334 raw reads and 80 contigs with 25\u00d7 median coverage were obtained for strain Hazen. The 202F and Hazen genomes comprise 3,829,425\u2009bp and 3,821,401\u2009bp, respectively, and both have 27.2% G+C content. Strain 202F contained 3,496 protein-coding sequences and 121 RNAs, while Hazen had 3,422 protein-coding sequences and 120 RNAs. A comparative analysis between the previously sequenced 202F genome (GenBank accession number C. botulinum strains 202F and Hazen have been deposited in DDBJ/ENA/GenBank under accession numbers NPMX00000000.1 and NPMY00000000.1 for the contigs and SRA accession numbers SRR17916278 and SRR17916277 for the raw reads, respectively.The genome sequences of"}
+{"text": "Residents and staff of continuing care retirement communities (CCRC) experienced many challenges during the COVID-19 pandemic including loss, social isolation, and staff turnover. This study examined factors that contribute to resilience in staff during the late stage of the pandemic using the Connor-Davidson Resilience Scale. Resilience scores ranged from 0 (low) to 100 (high). A total of 96 staff  were enrolled, and average age was 48.41 years (SD = 16.16). Average resilience in staff was 75.16 (SD = 11.81). Those under 35 years of age reported lower resilience scores (M = 67.38) compared to those 35-49 years of age (M = 76.65), 50-64 (M = 75.83), and 65 years and older (M = 82.71), p <.05. Staff who were married scored higher than those who were not (M = 76.63 vs 69.05), p < .05. Findings can inform professional development programs aimed at increasing coping skills in staff."}
+{"text": "Correction to: BMC Psychiatry 22, 106 (2022)https://doi.org/10.1186/s12888-022-03764-yFollowing publication of the original article , the aut5. Mangolian Shahrbabaki P, Dehghan M, Maazallahi M, Asadi N. Fear and anxiety in girls aged 7 to 11 years old and related factors during the coronavirus pandemic. Clin Child Psychol Psychiatry. 2022;27(1):259-68.The original article  has been"}
+{"text": "We studied SARS-CoV-2 genomes from travelers arriving in Hong Kong during November 2021\u2013February 2022. In addition to Omicron and Delta variants, we detected a BA.1/BA.2 recombinant with a breakpoint near the 5\u2032 end of the spike gene in 2 epidemiologically linked case-patients. Continued surveillance for SARS-CoV-2 recombinants is needed. We previously demonstrated the feasibility of testing incoming travelers for SARS-CoV-2 genomic surveillance (The SARS-CoV-2 Omicron variant (Pango lineage B.1.1.529) emerged in November 2021. Within a few weeks, subvariants BA.1, BA.1.1, and BA.2 were detected in varying proportions on different continents, but BA.1 initially was dominant (>100). Deduced genomes predominantly were Delta (n = 58) and Omicron  variants (Using our previously described next-generation sequencing method  Table 1.https://www.pfizer.com); patient 1 received the second dose on November 1, 2021, and patient 2 received the second dose on June 22, 2021.In our phylogenetic analysis, 2 additional nearly identical sequences formed a distinct branch in the Omicron clade  Figure 2https://www.medrxiv.org/content/10.1101/2022.03.03.22271812v2; T. Peacock, unpub. data, https://github.com/cov-lineages/pango-designation/issues/441).The distinct topology of viral sequences from these patients suggested that they were infected by a recombinant virus. To test that hypothesis, we used previously reported BA.1- and BA.2-defining single-nucleotide polymorphisms (SNPs) to analyze the genomes . We founWe further examined our sequence data to exclude the possibility of coinfection or contamination , panel AWe found no similar BA.1/BA.2 recombinant sequences in GISIAD or GenBank , suggesting a novel recombinant. The BA.1 region of this recombinant virus is genetically close to 3 BA.1 sequences detected in Europe and the United States , panel Bhttps://doi.org/10.1101/2020.08.05.238386; P. Colson et al., unpub. data; T. Peacock, unpub. data). The high transmissibility of Omicron (Emerging Omicron subvariants could allow vaccine breakthrough and widespread reinfection. Previous studies reported detection of SARS-CoV-2 interlineage recombinants at the same time as different SARS-CoV-2 lineages were cocirculating (Additional information recombinant BA.1/BA.2 SARS-CoV-2 virus in arriving travelers, Hong Kong, China, February 2022."}
+{"text": "S-gene) amplification (SGTF) in real-time reversetranscription\u2013polymerase chain reaction (RT-PCR) was used as a proxy indicator ofinfection with likely BA.5-related sublineages and S-gene targetpresence (SGTP) of infection with likely XBB/XBB.1.5-related sublineages . Specimenswith missing Ct values for N or ORF1ab were excluded.SARS-CoV-2\u2013positive specimens with either null or reduced amplification of thespike S-gene  were considered tohave SGTF  or SGTP (XBB/XBB.1.5-related); control-patients werethose who received a negative NAAT result. Tests among persons fulfilling any offollowing criteria were excluded from analyses: 1) presence of an immunocompromisingconditionAs of January 16, 2023, genomic sequencing data were available for a random subset ofICATT specimens with SGTP and collection dates through January 2, showing an increase inXBB.1.5 prevalence over time. During December 1, 2022\u2013January 2, 2023, XBB.1.5comprised 33% (495) of specimens exhibiting SGTP. During the interval December11\u2013January 2, XBB.1.5 accounted for 38% of sequenced ICATT specimens with SGTP(377), and during the interval December 18\u2013January 2, XBB.1.5 accounted for 43%of sequenced ICATT specimens with SGTP (252) were positive for SARS-CoV-2, including 10,596(78%) with SGTF (BA.5-related) and 3,052 (22%) with SGTP (XBB/XBB.1.5-related) . More coAcross age groups, VE was generally similar against BA.5-related infections andXBB/XBB.1.5-related infections. VE against symptomatic BA.5-related infection was 52%among persons aged 18\u201349 years, 43% among persons aged 50\u201364, and 37%among those aged \u226565 years . VEagaiThis report provides the first estimates of bivalent mRNA COVID-19 VE againstsymptomatic SARS-CoV-2 infection with XBB-related sublineages. These preliminaryestimates from national pharmacy testing conducted during December 1,2022\u2013January 13, 2023, showed relative bivalent booster dose VE  to be similar for XBB/XBB.1.5sublineage\u2013related infections and BA.5 sublineage\u2013related infections.VE estimates for both sublineages included in this analysis were similar toestimates from the same ICATT network published during a period of Omicron BA.5,BQ.1, and BQ.1.1 sublineage circulation in fall 2022 -gene target presence as a proxy for BA.2sublineages, including XBB and XBB.1.5, during December 2022\u2013January2023, the results showed that a bivalent mRNA booster dose providedadditional protection against symptomatic XBB/XBB.1.5 infection for at leastthe first 3 months after vaccination in persons who had previously received2\u20134 monovalent vaccine doses.Using spike (As new SARS-CoV-2 variants emerge, continued vaccine effectiveness monitoringis important. All persons should stay up to date with recommend COVID-19vaccines, including receiving a bivalent booster dose when eligible."}
+{"text": "Hepatitis C virus (HCV) is a risk factor that leads to hepatocellular carcinoma (HCC) development. Epigenetic changes are known to play an important role in the molecular genetic mechanisms of virus-induced oncogenesis. Aberrant DNA methylation is a mediator of epigenetic changes that are closely associated with the HCC pathogenesis and considered a biomarker for its early diagnosis. The ANDSystem software package was used to reconstruct and evaluate the statistical significance of the pathways HCV could potentially use to regulate 32 hypermethylated genes in HCC, including both oncosuppressor and protumorigenic ones identified by genome-wide analysis of DNA methylation. The reconstructed pathways included those affecting protein-protein interactions (PPI), gene expression, proteinactivity, stability, and transport regulations, the expression regulation pathways being statistically significant. It hasbeen shown that 8 out of 10 HCV proteins were involved in these pathways, the HCV NS3 protein being implicatedin the largest number of regulatory pathways. NS3 was associated with the regulation of 5 tumor-suppressor genes,which may be the evidence of its central role in HCC pathogenesis. Analysis of the reconstructed pathways has demonstratedthat following the transcription factor inhibition caused by binding to viral proteins, the expression of a numberof oncosuppressors  was suppressed, while the expression of others  was activated. Thus, the performed gene-network reconstruction has shown that HCV proteins can influencenot only the methylation status of oncosuppressor genes, but also their transcriptional regulation. The results obtainedcan be used in the search for pharmacological targets to develop new drugs against HCV-induced HCC. Liver cancer is the third leading cause of cancer-relateddeath in the world according to year 2020 statistics with over900,000 new cases of this pathology registered the sameyear around the world . Hepatocellularcarcinoma (HCC) has been the dominant type of primaryliver cancer, comprising about 90 % of all the cases . It may be caused by several risk factors suchas aflatoxin exposure, alcohol consumption; hepatitis B orC (HCV) virus infection, liver cirrhosis, non-alcoholic fattyliver disease, non-alcoholic steatohepatitis, metabolic syndrome,obesity, type II diabetes, and genetic predisposition.Currently, a lot of data has been accumulated on HCVassociation with impaired liver function, cirrhosis and HCCdevelopment . Having gotten into a humanbody, HCV seeks to exercise control over the biologicalprocesses occurring in host cells in order to increase its survivaland replication efficiency. In more than 70 % of thoseinitially infected, the disease takes on a chronic course, so thepatients experience progressive liver-tissue fibrosis and cirrhosisaccompanied by long-term inflammation . Using various mechanisms for infected cell cooptation,the virus can inadvertently lead to HCC development. At the same time, the molecular andgenetic mechanisms of virus-induced carcinogenesis remainunderstudied.In addition, HCC pathogenesis is associated with epigeneticmodifications and aberrant DNA methylation being a mediatorof epigenetic changes  thatcan serve as a biomarker for early HCC diagnosis .To establish the functional links between genes and to elucidatethe molecular mechanisms of biological processes, themethods for gene networks reconstruction have been widelyemployed. Previously, we developed the Associative NetworkDiscovery System (ANDSystem) software package designedto reconstruct gene networks based on the knowledge extractedfrom factual databases and scientific publications usingtext-mining techniques . The package has enabled one toreconstruct the molecular mechanisms of a number of pathologiessuch as preeclampsia , tuberculosis, comorbid conditions of asthma andhypertension , COVID-19 , HCV life cycle , etc.In the present study, ANDSystem was employed to reconstructthe regulatory pathways describing the potentialregulation mechanisms of the genes hypermethylated in HCCby HCV proteins. The analysis looked at the 32 genes knownto be hypermethylated HCC markers. Among the 7 types ofreconstructed regulatory pathways including protein-proteininteractions (PPI), gene expression, protein activity, stabilityand transport regulations, those responsible for gene expressionregulation turned out to be statistically significant. Ninemarker genes were identified that could potentially be subjectto regulation by HCV proteins, including three HCC suppressorgenes  that could be negativelyregulated and one apoptosis suppressor gene (TERT ) that canbe positively regulated.Genes hypermethylated in HCC. Information about thehypermethylated genes was taken from publications (Table 1).Only those genes were considered whose hypermethylationwas associated with HCC and confirmed through the analysisand meta-analysis given in the publications. The schematic ofthe data-processing algorithm can be seen in Figure 1.Regulatory pathways reconstruction in ANDSystem.The regulatory pathways were reconstructed using the ANDSystemsoftware package  thathad been designed to perform gene-networks reconstructionbased on automated analysis of scientific texts and factualdatabases. ANDSystem includes a knowledge base with morethan 40 million facts about molecular-genetic interactions,containing physical intermolecular interactions, gene expression,protein activity, stability and transport regulations. In thepackage, it is the the ANDVisio program that reconstructs andanalyzes gene networks using the Pathway Wizard functionperforming search calls to the knowledge base according toa given pattern. A schematic description of the used patternsis given in Table 2For instance, P4 means searching for all possible moleculargenetic pathways in the ANDSystem knowledge base thatsatisfy the following requirement: the first participant inthe pathway is the viral protein (Vp); the second is humanprotein (Hp); the third is a human gene from a list of target genes (Tg); the last member of the pathway is a Tg-encodedprotein (Tp). Further in the text, HCC marker genes will beregarded as target ones. Interactions between pathway participantsare represented by the following types: Vp and Hpare linked by PPIs; Hp and Tg are \u201cexpression regulation\u201dinteraction type (Exp reg), where Hp regulates Tg gene expression;Tg and Tp are interaction of the \u201cexpression\u201d interactiontype (Exp), i. e., the Tp protein is the expression product ofthe Tg gene. Examples of regulatory pathway reconstructionin ANDSystem using the patterns presented in the previouswork .Estimating the statistical significance of pathways. Thepattens from Table 2 were used to calculate the number ofmarker genes K participating in the regulatory pathways,as well as the number of such participants in the sample ofcontrol gene. The likelihood of observing the number K forrandom reasons was estimated using the standard hypergeometricdistribution and the hypergeom function from the SciPy 1.8.0 package (https://scipy.org). For the purposes ofstatistical processing, a group of genes proposed by Hoshidaet al. (2008) as a control to predict HCC outcomes based onthe expression level of genes was takenReconstruction of the potential regulatory pathwaysHCV proteins use to affect HCC marker genesA set of hypermethylated HCC marker genes was used toreconstruct the potential regulatory pathways through whichviral proteins could modulate the genes playing an importantrole in HCC pathogenesis (see Table 1). The set had beenbased on the published results of a genome-wide analysis ofDNA methylation and included 30 genes, the expression ofwhich, according to the studies, was reduced in hepatocellularcarcinoma, and two genes (WT1 and TERT ) with increasedexpressionTo reconstruct the regulatory pathways, the ANDSystemsoftware package was used. The search queries to the knowledgebase were formed using the pathway patterns presentedin Table 2. The patterns described different types of regulatorypathways determined by different combinations of moleculargeneticinteractions, including PPIs, gene expression, proteinactivity, stability, and transport regulationsAnalysis of the statistical significance of the pathwaysautomatically reconstructed by ANDSystem according to thegiven patterns showed that among the seven types of regulatorypathways analyzed, expression regulation ones turnedout to be statistically significant (P4 in Table 3). This patterndescribes the pathways including four participants: (1) viralproteins; (2) human transcription factors (TF) involved in PPIswith viral proteins; (3) marker genes presented in Table 1,whose expression is regulated by (2); (4) protein products ofmarker genes.The gene network describing the regulation pathways ofHCC marker genes included 8 HCV proteins, 7 intermediatehost proteins involved in PPIs with HCV proteins, and 9 genes whose aberrant expression correlated withHCC progression .Analysis of the regulatory pathwaysThe regulatory pathways involved 8 out of 10 HCV proteinsand 6 human genes, which protein products acting as the intermediateparticipants the viral proteins could form proteinheterocomplexes with. The latter included such genes oftranscription factors as STAT3 , NR4A1 (Nuclear receptor subfamily 4group A member 1), JUN (c-Jun/activator protein 1), BCL6(B-cell lymphoma 6 protein), transmembrane receptor NOTC1(Neurogenic locus notch homolog protein 1) and histonemethyltransferase SMYD3 (Lysine methyltransferase SETand MYND domain containing protein 3).Most of the viral proteins were associated with RUNX3 andWT1 regulation. Six of them  interacted with NR4A1 being a general expressionregulator of these two HCC marker genes.HCV protein NS3 (p70) interacted with the largest numberof expression regulators, and through these interactions itcould potentially regulate the expression of five tumor suppressorgenes and that of TERT.Now, let us consider the possibilities of implementing ofthese regulatory pathways in more detail.p8, p21, p68, gp32, p23, NS1/NR4A1/RUNX3, WT1. Thisregulatory pathway suggests six HCV proteins  can possibly affect HCC development by controllingthe activity of the RUNX3 and WT1 genes through theNR4A1 transcription factor. Indeed, NR4A1 directly interactswith the RUNX3 and WT1 promoters, suppressing RUNX3activity and activating that of WT1 . Both factors are involved in apoptosis regulation,hence, RUNX3 promotes activation of the extrinsic,TRAIL-induced apoptosis pathway , whileWT1 controls the mitochondrial  apoptosis pathwaythrough the regulation of the Bcl-2 anti-apoptotic proteingene, and, depending on a cell type, affects the expressionof the Bcl-2 gene in both positive and negative ways . It has been shown that in HCC, anincreased expression of the WT1 gene is observed, which isdue to hypermethylation of its promoter and correlates witha poor prognosis . Thesedata suggest that the role WT1 plays in HCC progression isassociated with blocked apoptosis.Experiments have demonstrated that HCV core proteininhibits at least the NR4A1 and RUNX3 genes expression ininfected cells , contributing to suppressing anexternal apoptosis pathway. The Y2H test (Two Hybrid Test)has shown NR4A1 can interact with such viral proteins asCORE, E1, E2, NS2, NS4A, and NS5B , but except for CORE, the effects of the other HCVproteins on TF activity have not been investigated yet.E1, NS3, Core, p23, NS1, p68/JUN, NOTC1, STAT3/TERT. Aberrant expression of the TERT gene, associated,among other things, with hypermethylation of its promoter, isa prognostic marker of HCC . TERT affects diseaseprogression through stimulation of cell proliferation due toreactivation of its gene expression in carcinoma cells . TERT activity has beenshown to also increase in HCV-infected cells, partly throughdirect interaction of the core protein with the enzyme , however, in general, the mechanisms HCVproteins affect TERT activity are not clear. This regulatorypathway suggests the involvement of the virus NS3, Core,E1, p23, NS1, and NS5B proteins in TERT gene expressionthrough interaction with the JUN (AP-1), STAT3, and NOTC1proteinsIndeed, experiments have shown that there is a possibilityto affect TERT expression through the AP-1 and STAT3 TFs being its direct regulators , as well as through the NOTC1 signaling pathway. Moreover, it has been demonstratedthat the HCV NS3 protein affects NOTC1 activity throughthe SRCAP transcription factor  and theexpression of AP1- and STAT3-regulated genes , however,the specific mechanisms of realization of these influences ininfected hepatocyte cells have not been practically examined.gp32, p70/JUN/WIF1. This regulatory pathway describesthe effect the NS3 and E1 HCV proteins have on WIF1 (Wntinhibitory factor 1) gene expression through interaction withthe c-Jun/AP-1 TF. WIF1 is a tumor suppressor that reducescell growth in HCC , and its expressionlevel is a prognostic indicator of the course of the disease.Experiments have demonstrated that there is both the possibilityof a direct effect of the NS3 and E1 proteins on theactivity of c-Jun/AP-1 , and the lattercan affect the expression of the WIF1 gene through interactionwith the DNMT1 methyltransferase (DNA methyltransferase1), suppressing WIF1 in gallbladder cancer cells . But what mechanisms of WIF1 gene suppressionare initiated in HCV-infected hepatocarcinoma cells remainsunknown.p70/STAT3/MGMT, DAPK1, SOCS1. This regulatorypathway is initiated by NS3 (p70) affecting the activity of theMGMT, SOCS, and DAPK1 genes through interaction withthe STAT3 TF. Proteins DAPK1 (Death-associated proteinkinase 1), MGMT (Methylated-DNA-protein-cysteine methyltransferase),and SOCS1 are considered tumor suppressors, and their low expressionin carcinomas correlates with disease progression .Experiments have demonstrated that NS3 can directly interactwith the STAT3 TF  involvedin the regulation of the expression of the abovementionedgenes , however, the effect of STAT3on MGMT, SOCS, and DAPK1 expression is not unambiguousand may be associated with cell specialization. As forthe mechanisms regulating the expression of these genes inHCV-infected hepatocarcinoma cells, they have not beenstudied yet.p70/BCL6/TP53. TP53 is a key activator of intrinsic apoptosispathway. The NS3 (p70) protein affects TP53 throughinteraction with the BCL6 (B-cell lymphoma 6 protein) TF.TP53 is a HCC marker gene of and its low expression correlateswith poor disease prognosis . BCL6 represses the TP53 gene in lymphoid cells,and its constitutive expression protects B lymphocytes fromDNA damage-induced apoptosis . However, the data describing the effectof HCV has on TP53 and BCL6 expression in these cells areassociated with a possible mutation induction and are mutuallyexclusive . Theinteraction of NS3 and BCL6 is discussed in ,but the specific mechanisms NS3 affects the TF activity havenot been studied yet.The studied set of hypermethylated HCC marker genes (seeTable 1) included 30 downexpressed and two over-expressedgenes. Using ANDSystem, the regulatory pathways by whichHCV proteins are able to influence the expression of thesemarker genes were reconstructed. The relationship betweenthe pathways and HCC-associated key biological processes isshown in Figure 3. According to the published data, the WT1,RUNX3, TP53, and SOCS1 genes are closely associated withapoptosis ,while the MGMT, TERT, RASSF1A, and WIF1 genes \u2013 withapoptosis and cell proliferation .The analysis showed the identified pathways could potentiallybe a part of the mechanism HCV proteins affect the activityof HCC marker genes. However, the effects the proteinshave on the function of human regulatory proteins during PPIformation are currently poorly understood. This fact preventsus from unambiguous interpretation of the reconstructedpathways, because what determines if a regulatory pathwayfunctions as an activator or suppressor of target gene expressionis whether or not the ability to regulate gene expressionremains in the host regulatory protein after its interaction withthe viral protein. Investigation of these effects requires furtherexperimental studies and computer molecular modeling.The literature describes the effects viral proteins can have onthe function of host proteins, e. g., the NS5A protein is knownto bind to SMYD3 in the cytoplasm and inhibit SMYD3 translocationto the nucleus . If the regulatoryproteins of a host organism are assumed to lose their ability toregulate gene expression due to the complexes formed withviral proteins, then the following effects can be expected:when considering the pathways regulating onco-suppressorexpression, four of the seven pathways inhibiting RASF1,RUNX3, WIF1, and DAPK1 will be suppressed, which canlead to their activation by HCV proteins, and that, in turn, willprevent carcinogenesis. In the remaining three pathways, theactivation of MGMT, SOCS1 and TP53 will be suppressed,which may possibly have a protumor effect. In the presentedpathways , WT1, MGMT, SOCS1 and TP53 areactivated by the corresponding factors ,while the expression of RASF1, RUNX3, WIF1, and DAPK1genes is negatively controlled .In the case of the TERT and WT1 genes that can be attributedto those with protumor activity, suppression of theWT1 gene can be expected and will lead to a negative effecton carcinogenesis. As for the TERT gene involved in apoptosissuppression and stimulating cell proliferation , according to our results , this gene was controlled through three different regulatorypathways. Its expression was activated by two pathwaysinvolving the STAT3 and NOTC1 host genes , and one of the pathwayssuppressed the expression involving c-JUN/AP-1 . The interaction with these host proteins couldlead to a blockage of these regulatory pathways. The pathwayinvolving c-JUN/AP-1 may be of particular interest in this respect, since inhibition of this TF by viral proteins (gp32 andNS3) could promote TERT activation. These assumptions arein good agreement with the data on differential gene expressionin acute HCV infection when the infected cells showedan increased TERT expression , so thispathway can be a promising pharmacological target.Thus, the considered assumptions lead one to conclude thepresence of multidirectional regulation of the observed expressionof HCC marker genes. This may indicate that not all regulatorypathways controlled by viral proteins can be attributedto HCC risk factors. However, the regulatory pathways thatensure the protumor activity of virus proteins undoubtedlydeserve additional study to understand the mechanisms ofvirus-induced HCC carcinogenesis. 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Low DAPK1 expression correlates with poor prognosisand sunitinib resistance in clear cell renal cell carcinoma.Aging ( Albany NY). 2020;13(2):1842-1858. DOI 10.18632/aging.103638.Takakura M., Kyo S., Inoue M., Wright W.E., Shay J.W. Function ofAP-1 in transcription of the telomerase reverse transcriptase gene(TERT ) in human and mouse cells. Mol. Cell Biol. 2005;25(18):8037-8043. DOI 10.1128/MCB.25.18.8037-8043.2005.Tan Y., Li Y. HCV core protein promotes hepatocyte proliferation andchemoresistance by inhibiting NR4A1. Biochem. Biophys. Res.Commun. 2015;466(3):592-598. DOI 10.1016/j.bbrc.2015.09.091.Tucci F.A., Broering R., Johansson P., Schlaak J.F., K\u00fcppers R. B cellsin chronically hepatitis C virus-infected individuals lack a virusinducedmutation signature in the TP53, CTNNB1, and BCL6 genes.J. Virol. 2013;87(5):2956-2962. DOI 10.1128/JVI.03081-12.Xu R.H., Wei W., Krawczyk M., Wang W., Luo H., Flagg K., Yi S.,Shi W., Quan Q., Li K., Zheng L., Zhang H., Caughey B.A., Zhao Q., Hou J., Zhang R., Xu Y., Cai H., Li G., Hou R., Zhong Z.,Lin D., Fu X., Zhu J., Duan Y., Yu M., Ying B., Zhang W., Wang J.,Zhang E., Zhang C., Li O., Guo R., Carter H., Zhu J.K., Hao X.,Zhang K. Circulating tumour DNA methylation markers for diagnosisand prognosis of hepatocellular carcinoma. Nat. Mater. 2017;16(11):1155-1161. DOI 10.1038/nmat4997.Yang C., Zhang Y., Wang J., Li L., Wang L., Hu M., Xu M., Long Y.,Rong R., Zhu T. A novel cyclic helix B peptide inhibits dendriticcell maturation during amelioration of acute kidney graft rejectionthrough Jak-2/STAT3/SOCS1. Cell Death Dis. 2015;6(11):e1993.DOI 10.1038/cddis.2015.338.Ye S., Zhao X.Y., Hu X.G., Li T., Xu Q.R., Yang H.M., Huang D.S.,Yang L. TP53 and RET may serve as biomarkers of prognosticevaluationand targeted therapy in hepatocellular carcinoma. Oncol.Rep. 2017;37(4):2215-2226. DOI 10.3892/or.2017.5494.Zhang C., Li J., Huang T., Duan S., Dai D., Jiang D., Sui X., Li D.,Chen Y., Ding F., Huang C., Chen G., Wang K. Meta-analysis ofDNA methylation biomarkers in hepatocellular carcinoma. Oncotarget.2016;7(49):81255-81267. DOI 10.18632/oncotarget.13221.Zhang H., Weng X., Ye J., He L., Zhou D., Liu Y. Promoter hypermethylationof TERT is associated with hepatocellular carcinomain the Han Chinese population. Clin. Res. Hepatol. Gastroenterol.2015;39(5):600-609. DOI 10.1016/j.clinre.2015.01.002.Zhu Z., Tran H., Mathahs M.M., Moninger T.O., Schmidt W.N. HCVinduces telomerase reverse transcriptase, increases its catalytic activity,and promotes caspase degradation in infected human hepatocytes.PLoS One. 2017;12(1):e0166853. DOI 10.1371/journal.pone.0166853.Zhu Z., Wilson A.T., Gopalakrishna K., Brown K.E., Luxon B.A.,Schmidt W.N. Hepatitis C virus core protein enhances Telomeraseactivity in Huh7 cells. J. Med. Virol. 2010;82(2):239-248. DOI10.1002/jmv.21644.Zong C., Qin D., Yu C., Gao P., Chen J., Lu S., Zhang Y., Liu Y., Yang Y.,Pu Z., Li X., Fu Y., Guan Q., Wang X. The stress-response moleculeNR4A1 resists ROS-induced pancreatic \u03b2-cells apoptosis via WT1.Cell Signal. 2017;35:129-139. DOI 10.1016/j.cellsig.2017.03.012.Zucman-Rossi J., Villanueva A., Nault J.C., Llovet J.M. Genetic landscapeand biomarkers of hepatocellular carcinoma. Gastroenterology.2015;149(5):1226-1239.e4. DOI 10.1053/j.gastro.2015.05.061."}
+{"text": "Klebsiella pneumoniae, a leading pathogen for mortality associated with antimicrobial resistance (AMR), was responsible for > 250,000 deaths in 2019. Genomic surveillance can guide the development of vaccines and new antibiotics against this pathogen. While AMR disproportionally affects low-and middle-income countries, limited genomic data are available from these countries. To close this gap, this study provides genomic characterization of K. pneumoniae blood isolates recovered in Peru.K. pneumoniae blood culture isolates were collected during an AMR surveillance study (VIRAPERU) from Jul 2017 to Oct 2019, from 15 tertiary hospitals from 13 regions of Peru. DNA extraction , DNA library  and genome sequencing  were conducted. De novo assembling (SPAdes v3.13.1), quality assessment and annotation (Nullarbor v2.0), and identification of species, ST group, K/O loci, AMR (Kleborate v2.0.1) were conducted. Phylogenetic trees were built with Microreact v.192.Consecutive non-duplicate K. pneumoniae isolates, 114 were recovered and confirmed to belong to the Klebsiella taxon. Six species were identified, the most prevalent being K. pneumoniae (106), followed by K. quasipneumoniae (3). Among carbapenem-resistant isolates (n=13), 10 (77%) carried the blaNDM-1 gene, while other three carried blaKPC-2 (1), blaIMP-16 (1) or no carbapenemase (1). Phenotypic co-resistance to colistin was present in 2 isolates, both negative for mcr-1 gene. The most common mechanisms of resistance to 3rd generation cephalosporins, quinolones and to aminoglycosides were CTX-M-15 (74.4%), qnrB (63.6%), aac(6')-Ib-cr (87.5%), respectively. Many other AMR genetic determinants were identified . Sixty ST groups were identified, but only 6 ST groups carried a carbapenemase gene . A wide diversity of K and O locus was also identified (54 distinct K-loci and 14 distinct O-loci).From 119 K. pneumoniae strains, carrying multiple AMR genes. Carbapenem resistance is principally a result of blaNDM-1 carriage, found across 6 specific ST groups.Bloodstream infections in Peru are caused by a wide diversity of Omai Garner, PhD, Beckman Coulter, Inc.: Clinical trial data collection funded by Beckman Coulter, Inc. Victor Nizet, MD, Cellics Therapeutics: Advisor/Consultant|Clarametyx Biosciences: Advisor/Consultant|Vaxcyte, Inc.: Advisor/Consultant|Vaxcyte, Inc.: Grant/Research Support."}
+{"text": "Liver transplanted (LT) patients for hepatocellular carcinoma (LT-HCC) or for other causes (LT-no-HCC) may develop post-transplantation malignancies. Although immune activation and senescence are frequently implicated in cancer development, no data is available on their possible role as biomarkers predictive of tumor onset in this setting. A total of 116 patients were investigated: the 45 LT-HCC patients were older than the 71 LT-non-HCC (p=0.011), but comparable for sex, HCV, HBV infection and immunosuppressive treatment. At baseline, the numbers of activated and senescent-like circulating cells were significantly higher in LT-HCC patients than in LT-no-HCC ones. After a median follow-up of 26.8 months, 6 post-transplant malignancies (PTM) occurred: 4 in LT-HCC (8.9%) and 2 in LT-no-HCC (2.8%) patients. Overall, subjects with high percentages of activated and exhausted T and B cells at baseline were at higher risk of PTM. Notably, within the LT-HCC group, a higher percentage of senescence-like T cells was also associated with cancer development. Moreover, patients with PTM had higher telomere erosion and higher levels of circulating PAMPs (16S rDNA) and DAMPs (mtDNA) when compared with matched patients without PTM. Overall, these findings suggest that immune activation and exhaustion may be useful to predict the risk of PTM occurrence, regardless of the cause of transplantation. In LT-HCC, T-cell senescence represents an additional risk factor for tumor onset. Liver transplantation is the treatment of choice for patients with end-stage liver disease, acute liver failure or hepatocellular carcinoma (HCC). In Italy, a total of 12,519 liver transplants were performed between 2010 and 2020  compared to liver transplanted patients for other causes (LT-no-HCC)  followed by chronic local inflammation \u201311. Afte+ T cells. Consistently, lower thymic output and shorter telomeres were associated with a higher risk of developing cancer in the same cohort , which have been implicated in both aging and cancer development , 16. In e cohort . Further outcome .Considering the possible clinical relevance of the availability of suitable biomarkers for the prediction of tumor onset in LT patients, in the present study we have characterized immune activation, exhaustion, and senescence markers in the blood of a prospective series of LT patients with the final goal to assess their possible usefulness in the definition of the risk of PTM development.3 = 4/3 x 3.14 x (radius of the tumor nodule in mm3)\u201d. Total tumor volume (TTV) was calculated as the sum of the tumor volume of each nodule. LT-HCC patients were followed for HCC recurrence through CT-scan or MRI every 3 months during the first year, and every 6 months thereafter, according to the European Association for the Study of the Liver/European Organization for Research and Treatment of Cancer (EASL-EORTC) clinical practice guidelines for the management of hepatocellular cancer . Patients\u2019 clinical information were recorded and included personal details , transplant details  score and alpha-fetoprotein levels at liver transplantation) and post-transplantation immunosuppressive schedule, considering modifications during follow-up. For LT-HCC patients, tumor characteristics were assessed through explant pathology evaluation for the following characteristics: number of HCC nodules, maximum nodular diameter, tumor differentiation and presence of macro/micro-vascular invasion. Vital tumor volume was quantified according to the following equation: \u201cTumor volume mmr cancer . As theyAfter a median follow-up of 26.8 months following liver transplantation, 6 patients developed a PTM (LT-PTM): 5 were DNMs and 1 was an HCC recurrence. DNMs were defined as neoplasms developing after transplantation in patients negative at pre-transplant screening for the HCC, or related pre-malignant lesions/conditions. DNMs were coded according to the International Classification of Diseases and Related Health Problems, 10th revision (ICD-10). Diagnosis of DNMs was established by histology on biopsies or surgical specimens of the tumor. Date of biopsy or surgical procedure was designated as the date of cancer diagnosis. All patients underwent a full pre-transplant screening aimed at excluding any type of cancer other than HCC, or related premalignant lesions/conditions, which was negative in all patients. Moreover, to avoid any accidental missing pre-LT diagnosis, lesions that appeared within 6 months after transplantation were excluded from the definition of DNM. In our cohort, the 5 DNMs were: infiltrating ductal carcinoma of the breast, ovarian cancer, larynx carcinoma, basal cell carcinoma of the external auditory canal, and prostate cancer. Among the 110 patients without PTM (LT-no-PTM), 26 subjects were selected as a matched (m)LT-no-PTM control group according to the following inclusion criteria: age, sex difference, immunosuppressive combined strategies, and a median time interval from baseline to follow-up. This study was approved by the Ethical Committee of Padova University Hospital (Prot. 4231/AO/17).Blood samples were collected in EDTA-containing tubes at baseline (at liver transplantation) and at follow-up . From samples at baseline and at closest time to the tumor onset in LT-PTM patients, or with a median time interval similar for mLT-no-PTM patients, peripheral blood mononuclear cells (PBMC) were isolated by centrifugation on a Ficoll-Paque  gradient. PBMC were cryopreserved in liquid nitrogen, and plasma samples at - 80\u00b0C, until use..Immunophenotyping was performed on cryopreserved PBMC. Cells were thawed, washed, stained for 20 min in the dark with the Live/Dead Fixable Near-IR Dead Cell Stain Kit  and the following labelled monoclonal antibodies (mAbs): anti-CD3 [fluorescein isothiocyanate (FITC)], anti-CD4 [peridinin chlorophyll protein (PerCP)], anti-CD38 [phycoerythrin (PE)], anti-HLA-DR [allophycocyanin (APC)], anti-CD279  [PE-Cy7], anti-CD57 [PE], anti-CD21 [BV421], anti-CD27 [PE-Cy7], anti-IgD [PE], anti-CD274  [BV421] ; anti-CD8 [VioGreen], anti-CD28 [APC], anti-CD19 [VioBright515], anti-CD10 [APC] . Cells were then washed and resuspended in phosphate-buffered saline supplemented with 1% paraformaldehyde. All samples were analyzed using LSRII Flow cytometer (Becton-Dickinson). A total of 50000 events were collected in the lymphocyte gate using morphological parameters (forward and side-scatter). Data were processed with FACSDiva Software (Becton-Dickinson) and analyzed using Kaluza Analyzing Software v.1.2 (Beckman Coulter) DNA was extracted from PBMC, using QIAmp DNA Blood Mini Kit  according to the manufacturer\u2019s instructions. Relative telomere length (RTL) was determined by monochrome quantitative multiplex real-time PCR, as previously described , 21.DNA extraction from 200 \u00b5l of plasma was performed using QIAamp DNA Mini Kit , and eluted in 50 \u00b5l of AE Buffer. A quantitative method based on real-time PCR assay was performed to quantify plasma levels of 16S rDNA with primer pair (forward 5\u2019-AGTTTGATCCTGGCTCAG-3\u2019 and reverse 5\u2019-GWATTACCGCGGCKGCTG-3\u2019) and probe (5\u2019-FAM-GCTGCCTCCCGTAGGAGT-BHQ-3\u2019), as previously described , 23. ResContinuous variables were summarized using median and interquartile range (IQR), categorical variables as frequencies and percentages. The median follow-up time was based on the reverse Kaplan-Meier estimator. Clinical characteristics at baseline were compared between LT-HCC and LT-no-HCC patients using the Kruskal-Wallis test, or Fisher exact test as appropriate. A Mann-Whitney test, adjusted for age, was used to address comparisons of each immunological parameter distribution between groups of interest. The impact of the immunological parameters on the probability of experiencing HCC or PTM was estimated in univariate logistic regression models fitted by penalized maximum likelihood to address low frequency data. Each covariate was considered as a categorical variable according to high and low levels. Optimal cut-points were selected using a criterion based on maximizing the Youden index, being the summary measure of the ROC curve, over all possible cut-points. The odds ratios (ORs) were reported with their 95% confidence interval (CI). ORs were also adjusted for age as continuous variable. Subgroup analyses were performed with explorative intent. All statistical tests were two-sided and a p value <0.05 was considered statistically significant. Statistical analyses were performed using the RStudio .vs 53.0 years (45.5-61.0), p=0.011.The patients\u2019 characteristics are summarized in + T cells and memory B cells were significantly higher in LT-HCC compared to LT-no-HCC patients (%CD8+CD38+HLA-DR+: 10.89 (5.61-18.52) vs 6.59 (4.26-9.25), p=0.003; %CD19+CD10-CD21-CD27+: 10.97 (5.59-20.68) vs 7.60 (3.00-13.72), p=0.040) (+ T cells (%CD4+CD38+HLA-DR+: 7.23 (4.11-14.12) vs 6.21 (3.49-9.23), p=0.092)  vs 7.54 (4.00-12.92), p=0.254; %CD4+PD-1+: 9.28 (5.66-13.50) vs 7.27 (4.54-10.34), p=0.222; %CD19+PD-L1+: 4.96 (3.12-9.75) vs 5.16 (2.18-9.00), p=0.797)  vs 5.92 (3.54-10.97), p=0.006; %CD4+CD28-CD57+: 3.80 (1.36-14.03) vs 1.80 (0.48-3.41), p=0.002; %CD19+CD27-IgD-: 12.20 (6.28-17.67) vs 6.59 (4.24-12.50), p=0.019)  following LT, 6 patients developed a PTM: 4 patients (8.9%) (3 DNMs and 1 an HCC recurrence) were in the LT-HCC group, and 2  were in the LT-no-HCC group Table\u00a01.To evaluate the role of immune activation, exhaustion and senescence as possible markers predictive of the risk of tumor development in this setting, we analyzed these immunological parameters at baseline in samples obtained from the 6 LT-PTM patients in comparison with those of 110 LT-no-PTM patients. At baseline, the two groups did not differ by age, sex, HCV and HBV infection  vs 7.14 (4.48-11.98), p=0.002; %CD4+CD38+HLA-DR+: 14.28 (10.53-16.84) vs 6.26 (3.58-9.77), p=0.0001; %CD19+CD10-CD21-CD27+: 25.57 (10.09-28.57) vs 8.55 (4.20-15.40), p=0.024) (The percentages of activated CD8p=0.024) Table\u00a03.+PD-1+: 19.81 (16.97-28.90) vs 7.78 (4.62-14.67), p=0.000; %CD4+PD-1+: 22.71 (17.96-27.89) vs 7.75 (4.47-10.85), p=0.000; %CD19+PD-L1+: 10.15 (7.82-19.75) vs 5.01 (2.23-9.09), p=0.015) (The percentage of circulating immune cells expressing markers of exhaustion was significantly higher in LT-PTM than in LT-no-PTM patients (%CD8p=0.015) Table\u00a03.+ T cells and B cells tended to be higher in the LT-PTM than in LT-no-PTM patients  vs 10.08 (5.33-12.83), p=0.002), while no difference was observed in senescent-like CD8+ and CD4+ T cells  copies/\u00b5l, p=0.008); since no difference occurred in the two groups during follow-up, this level still remained higher at follow-up (64.95 (29.51-106.15) vs 26.99 (11.27-58.74) copies/\u00b5l, p= 0.074) (vs 1822 (985-3984) copies/\u00b5l, p=0.096), but they become significantly higher at follow-up (3075 (2187-6244) vs 1668 (447-2424) copies/\u00b5l, p= 0.040)  vs 1.08 (0.80-1.63), p=0.391). Notably, at follow-up, the levels did not differ between the two groups (RTL: 0.79 (0.73-0.80) vs 0.77 (0.75-0.83), p=0.873), thus indicating that LT-PTM patients tended to have a higher telomere erosion during the same follow-up interval  vs -0.09 , p=0.091).At baseline, relative telomere lengths (RTL) on PBMC were longer, although not statistically significant, in LT-PTM than in mLT-no-PTM patients (RTL: 1.46 (1.03-1.63) de novo malignancies (DNMs) and HCC recurrence are the most common causes of long-term morbidity and death . The patients/participants provided their written informed consent to participate in this study.MRP, SS, MT, PP, DS, PB and ADR, designed the study. MRP, ER, FC, SG, performed the investigations. SS, DS, UC, PB, provided samples and reagents. PDB performed the statistical analysis. MRP wrote the manuscript. SS, MT, SG, PDB, PP, DS, UC, RD, PB and ADR supervised the project, reviewed and editing the manuscript. All authors contributed to the article and approved the submitted version.This work was supported by Ricerca Corrente IRCCS Centro di Riferimento Oncologico, Aviano: Italian Association for Research on Cancer; Grant number: AIRC IG No. 19112; and \u201cPathogenesis of EBV-driven post-transplant malignancies\u201d granted by Department of Surgery, Oncology and Gastroenterology (DiSCOG), University of Padova, Italy .The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Bifidobacterium lactis BL-99 (B.lactis BL-99) on the intestinal inflammation and functionsin the zebrafish models. After feeding for 6 hours, B.lactis BL-99 was fully retained in the larval zebrafishintestinal tract and stayed for over 24 hours. B.lactis BL-99 promoted the intestinal motility andeffectively alleviated aluminum sulfate-induced larval zebrafish constipation(p < 0.01). Irregular high glucose diet induced adultzebrafish intestinal functional and metabolic disorders. After fed withB. lactis BL-99, IL-1\u03b2gene expression was significantly down-regulated, and IL-10 andIL-12 gene levels were markedly up-regulated in this model(p < 0.05). The intestinal lipase activity was elevatedin the adult zebrafish intestinal functional disorder model afterB. lactis BL-99 treatment(p < 0.05), but tryptase content had no statisticalchanges (p > 0.05). B.lactis BL-99 improved the histopathology of the adultzebrafish intestinal inflammation, increased the goblet cell numbers, andup-and-down metabolites were markedly recovered after treatment ofB. lactis BL-99 (p <0.05). These results suggest that B. lactisBL-99 could relieve intestinal inflammation and promote intestinal functions, atleast in part, through modulating intestinal and microbial metabolism tomaintain intestinal health.This study was designed to explore the therapeutics and the mechanisms of apatented and marked gastric acid and intestine juice-resistant probiotics Probiotics consumption for health promotion and well-being has increased worldwide inrecent years  and variBacillus spp [Bifidobacterium,Lactobacillus, Bacillus, and several otherbacterial species [Aeromonas hydrophila, A.salmonicida, Yersinia ruckeri, andV. anguillarum to intestinal mucus fromrainbow trout (in vitro) [L. rhamnosus also counteracts zebrafishneurotoxicity caused by PFBS [The most studied probiotic candidates in aquaculture belong to the Firmicutes phylum,namely lactic acid-producing bacteria (LAB) and llus spp , 9\u201313. Allus spp , 15. Stu species \u201318. Thes species , 20. LAB species ,and somn vitro) . Dietaryn vitro) , and die by PFBS .Bifidobacterium, which makes it difficult to reach and colonizein the intestine through gastric juice [Bifidobacterium lactisBL-99  was originally isolatedfrom the intestines of a Chinese healthy infant [Inability to acid and gastrointestinal juice is a common property ofic juice . Bifidoby infant , 26 and y infant ,28. Thiy infant .B. lactis BL-99 has no exogenous antibioticresistance genes [B.lactis BL-99 was confirmed no dose-dependent mortality andtoxicity throughout multidose oral toxicity tests in mice and rats and thusgenerally recognized as safe (GRAS) status as a probiotic [In vivo experiments inmice showed that B. lactis BL-99 significantlypromoted the growth of intestinal Bifidobacteria and Lactic acidbacteria, and inhibited Desulfovibrio and/orEnterobacter, especially Helicobacter pylori and/orEscherichia-Shiga Bacteria [ce genes  and hasrobiotic . In vivoBacteria , 30. ThiBacteria .The research of probiotics on intestinal microbial balance, intestinal functions,inflammation, and intestinal metabolites, etc. mostly use traditional mammalianmodels. Conventional mammalian enteritis models are chemical-induced, for example,DSS  was used to induce mice colitis , and TNBDanio rerio) intestinal composition is similar to that ofhumans, e.g., connective tissue, external-longitudinal muscle and circular muscle,et al. [B.lactis BL-99 on the digestive enzymes, motility, inflammationand metabolites in the larval and adult zebrafish models.Zebrafish  and adult male zebrafishat 3.5 months post fertilization (3.5 mpf) were used in this study. Zebrafishwere maintained at 28\u00b0C in fish water . The zebrafish facility and the laboratory at Hunter Biotechnology, Inc.are accredited by the Association for Assessment and Accreditation of LaboratoryAnimal Care (AAALAC) International , 42, by B. lactis BL-99 was deposited in the ChinaCommon Microbial Culture Collection and Management Center (CGMCC 15650) on April26, 2018 [5\u2019-AGA GTT TGA TCC TGG CTC AG-3\u2019) and 1492R(5\u2019-GGT TAC CTT GTT ACG ACT T T-3\u2019) [B. lactis BL-99 culture was proliferatedwith De Man Rogosa Sharpe (MRS) medium  supplemented with0.05% (w/v) L-cysteine (MRSC) for 12\u201348 hrs at 37\u00b0C aerobically [B.lactis BL-99 was 1.5*1011 CFU / g and preservedat -80\u00b0C.26, 2018  andiden T T-3\u2019) . The stawww.dingguo.com). CM-DiI cell-labeling solution  and trizol reagent (cat. # 12183555) were bought from Thermo FisherScientific (China) Co., Ltd. FastKing RT Kit (With gDNase) (cat. # KR116-02) wasbought from TIANGEN BioTec (Beijing) Co., Ltd (www.tiangen.com), and iTaq Universal SYBR(R) Green Supermix waspurchased from BIO-RAD Co., Ltd. (www.bio-rad.com). Fish trypsin ELISA kit (item no. ml064285) wasbought from Shanghai Enzyme-linked Biotechnology Co., Ltd. (www.mlbio.cn). Lipase (LPS) kit (item no.A054-2-1) was bought from Nanjing Jiancheng Bioengineering Institute  (www.njjcbio.com).Tricaine methanesulfonate (cat. # 886-86-2) and aluminum sulfate (cat. #D1909026) were ordered from Shanghai Aladdin Bio-Chem Technology Co., Ltd, nile red (cat. # MKBP6198V) from Sigma-Aldrich , and glucose (lot. # 20201105) was purchased from Sinopharm ChemicalReagent Co., Ltd . Tissue cell fixation solution at 4%concentration (cat. # AR-0211-250 mL) was ordered from Beijing DingguoChangsheng Biotechnology Co., Ltd  containing 0.5% DMSO in PBS [B.lactis BL-99 were treated with larval zebrafish for itsretaining time determination in the intestinal tract and for its effects on theintestinal motility and digestion functions as described below. The dye istransferred from mother to daughter bacteria and fluorescent B.lactis BL-99 were clearly visible in the zebrafishintestinal tract.After collection, O in PBS  at 37\u00b0C B. lactis BL-99 at a density of2.42*108 CFU/mL at 28\u00b0C. The zebrafish intestinal fluorescentimages were taken periodically at the designated time points to determine theretaining time of this probiotics. After treatment of fluorescentB. lactis BL-99 for 24 hrs, the zebrafishwere transferred into fish water for 4 and 24 hrs, respectively, 10 zebrafishwere randomly selected from each group and at each time point for visualobservation and image acquisition under a fluorescent stereomicroscope , installed with a high-speed video camera .Quantitative image analyses were performed using image-based analysis, the retaining time and lasting period ofB. lactis BL-99 in the larval zebrafishintestine tract were calculated based on the fluorescent intensity. To protectfluorescent B. lactis BL-99 from light-induceddecomposition, experiments were carried out at a constant temperature (28\u00b0C) inthe dark. All experiments were performed in duplicate and repeated for at least3 times.Wild-type larval zebrafish at 5 dpf were distributed into 6-well microplates, 30 zebrafish per well in 3 ml fish water and treated withfluorescent B. lactis BL-99at concentrations of 2.42*106, 2.42*107 and2.42*108 CFU/mL, respectively, for 24 hrs. Domperidone was usedas a positive control drug. The zebrafish treated with aluminum sulfate and nilered only served as a model control. The zebrafish without any treatment wereused as a negative control. At the end of treatments, the zebrafish were imagedunder a AZ100 fluorescent stereomicroscope, installed with a high-speed videocamera. The therapeutic effects of B. lactisBL-99 on the larval zebrafish constipation were determined based on theintestinal fluorescent quantitative analyses.The larval zebrafish of AB strain at 5 dpf were distributed into a 6-wellmicroplate, 30 zebrafish per well in 3 ml fish water. The zebrafish constipationmodel was established by treatment with 1 \u03bcg/mL aluminum sulfate  at 28\u00b0C 6, 2.42*107 and 2.42*108 CFU/mL)were used for B. lactis BL-99 treatment.Untreated control zebrafish were examined in parallel. The adult zebrafish werehoused in a light and temperature-controlled aquaculture facility with astandard 14:10 h light/dark photoperiod. (1) Days 1\u20133 of the experiment: exceptfor the untreated control zebrafish, the resting groups were not fed and starvedfor 3 days. B. lactis BL-99 groups weretreated with this probiotic at 3 designated concentrations, respectively, asdescribed above in fish water every day during the daytime for 8 hrs and thenlived in fresh fish water; (2) Days 4\u201317: B.lactis BL-99 groups were treated with this probiotic duringthe daytime for 8 hrs and then transferred into 3% glucose in fish water for 16hrs. The model zebrafish were only treated with 3% glucose for 16 hrs and theuntreated control zebrafish were fed with brine shrimp twice a day. On the 18thday of the experiment, the zebrafish intestinal tissues were collected and theintestinal digestive enzymes, inflammatory and immunity factor genes andhistopathology were examined, respectively, and the interventional effects ofB. lactis BL-99 were assessed.Seventy-five male adult zebrafish of 3.5 mpf (months post fertilization)wild-type AB strain were transferred into 5 L beaker in a volume of 4 Lcontaining 15 zebrafish. In the initial tests, three concentrations, interleukin-10 (IL-10)and interleukin-12 (IL-12) were determined in the adultzebrafish intestines by real-time quantitative PCR (qPCR) [B.lactis BL-99 treatment, total RNA was extracted from 10homogenized zebrafish per group using trizol reagent. About 2 \u03bcg total RNA ofeach sample was used for cDNA synthesis using FastQuant RT Kit (With gDNase) andqPCR amplifications were carried out with a CFX Connect detection system  using the iTaq Universal SYBR Green Supermix in which there werethree technical or biological replicates. The qPCR protocol was 2 minutes at95\u00b0C-40 cycles of 5 seconds at 95\u00b0C-30 seconds at 60\u00b0C. Expression data wasnormalized against the expression of \u03b2-actin and the relative quantification ofeach gene mRNA among groups was calculated as follows: The relative expressionof RNA = 2^\u0394\u0394C(t); \u0394\u0394C(t) = \u0394C(t)Model\u2014\u0394C(t)Probiotics;\u0394C(t) = \u0394C(t)Target gene\u2014\u0394C(t)\u03b2-actin. The primers used inthis study were as follows: \u03b2-ACTIN-FOR:TCGAGCAGGAGATGGGAACC, \u03b2-ACTIN-REV:CTCGTGGATACCGCAAGATTC (GenBank accession numbers57934) [IL-1\u03b2-FOR: GTCACACTGAGAGCCGGAAG,IL-1\u03b2-REV GCAGGCCAGGTACAGGTTAC [IL-10-FOR:TTCAGGAACTCAAGCGGGAT, IL-10-REV:AAGAGCAAATCAAGCTCCCCC  [IL-12-FOR:AACTCCTACAAGCCCAGCAC, IL-12-REV:ACACTCGGTCGTCAAACGAA . Each primer pair was designed usingNCBI/Primer-BLAST.To explore the possible anti-inflammation and the intestinal immune mechanisms ofR (qPCR) . Brieflys57934) , 50,IL- 405770) , IL-10-F 553957) , IL-12-FB. lactisBL-99 on the intestinal functions of the adult zebrafish, ELISA kits were usedto determine the intestinal tissue lipase activity and trypsin content. Theoptical density (OD) values were measured by multifunctional microplate reader at wavelength 595 nm for the protein concentration,580 nm for the lipase activity, and 450 nm for the trypsin content,respectively. The lipase activity and trypsin content per gram of protein inzebrafish intestinal tissues were calculated based on the OD values.In order to evaluate the effects of B. lactis BL-99 intervention, weperformed the gut histopathological examinations on the adult zebrafish. At theend of the experiments, zebrafish intestinal tissues were fixed in 4%paraformaldehyde in 0.1 M phosphate buffered saline for 4 hrs at 4\u00b0C, dehydratedin graded series of ethanol solutions before paraffin embedding. Embeddedzebrafish intestines were longitudinally sectioned at 5 \u03bcm and stained withhematoxylin and eosin (H&E) [To confirm the intestinal damage caused by the irregular high-sugar diet, and theeffects of in (H&E) , 51.Thip < 0.05 and fold change \u2265 1.2 or fold change \u2264 0.8333and VIP \u2265 1; and statistically significant changes in at least two dose groups.Student\u2019s t test was used for the statistical analyses of the metabolites.Ten adult zebrafish whole guts from each group were used for the intestinalmetabolite extraction and the metabolomic analysis. Twenty-five mg intestinaltissues from each gut were homogenized with 800 \u03bcL pre-cold precipitation agent. After sonication on ice for10 minutes, let the mixture stand at -20\u00b0C for 120 minutes, followed bycentrifugation at 25000 g for 15 min at 4\u00b0C. Six hundred \u03bcL of supernatant wastaken and put in a freeze-drying machine to drain and reconstituted in 600 \u03bcL of10% methanol solution. After ultrasound and centrifugation, the supernatant waschromatographed using 2777C UPLC system , and the eluted smallmolecules were collected in positive and negative ion modes using Xevo G2-XSQTOF . Metabolite resonances were identified according to theinformation from the Human Metabolome Database (HMDB) and Kyoto Encyclopedia ofGenes and Genomes (KEGG). Significantly changed metabolites between the controland treatment groups were identified following the criteria below:p < 0.05,p < 0.01 and p < 0.001 were allconsidered statistically significant. For quantitative analysis, all data werepresented as mean \u00b1 SEM, and results were statistically compared between theprobiotics-treated and model zebrafish groups. All experiments were repeated forat least 3 times. Zebrafish natural death in untreated groups was \u2264 10%, and allintra- and inter-group coefficient of variation (CV) was \u2264 25%.One-way ANOVA followed by the Dunnett\u2019s test was used to compare differencesamong groups. All statistical analyses were performed using the GraphPadsoftware , and B.lactis BL-99 for 2, 6 and 24 hrs, the fluorescentintensities in the larval zebrafish intestinal tracts were 6.02\u00b1 0.866,10.7\u00b1 1.08 and 13.0 \u00b1 0.601 pixels, respectively. Comparing the fluorescentintensities between 24 hr and 2 hr feeding, p < 0.001,but p > 0.05 when comparing the fluorescent intensitiesbetween 24 hr and 6 hr feeding, suggesting that B.lactis BL-99 effectively retained in the larvalzebrafish intestinal tract after 6 hr feeding.As indicated in B. lactis BL-99from the treatment solutions and transferred the zebrafish into fresh fishwater for 0, 4 and 24 hrs, the larval zebrafish intestinal fluorescence was14.4\u00b1 1.31, 11.5\u00b1 1.58 and 10.5\u00b1 1.57 pixels ,indicating that the larval zebrafish constipation model was successfullyestablished. The positive control drug Domperidone significantly promotedthe intestinal motility . The dose-dependent intestinal fluorescent intensity decreases were found in theconstipation zebrafish treated with B.lactis BL-99 at 2.42*106 (476071 \u00b1 20633pixels), 2.42*107 (456847 \u00b1 15814 pixels) and 2.42*108CFU/mL (414652 \u00b1 11561 pixels), respectively .As demonstrated in IL-1\u03b2 gene expression. A concentration-dependentdownregulations of the IL-1\u03b2 gene expression was observedin the model zebrafish treated with 2.42*106, 2.42*107and 2.42*108 CFU/mL of B.lactis BL-99, and the decreases were 0.511\u00b1 0.055,0.691\u00b1 0.072 and 0.969\u00b1 0.049 folds, respectively, relative to the modelgroup .The purity of the extracted RNA (A260/A280) was in the range of 1.95\u20132.12. Asshown in IL-10 gene expression. After treatment withB. lactis BL-99 at the concentrationsof 2.42*106, 2.42*107 and 2.42*108 CFU/mL,IL-10 and IL-12 expression levels wereelevated to 3.96\u00b1 0.219, 1.27\u00b1 0.150 and 1.85\u00b1 0.176 folds and 1.01\u00b1 0.097, 1.51\u00b1 0.368 and 4.13\u00b10.745 folds , respectively, relative to the modelgroup .B. lactis BL-99 had no statisticallysignificant effect on the intestinal trypsin content, although it showed anincreased trend as indicated in As shown in B. lactis BL-99 treatmentat 2.42*107 CFU/mL led to the developed high villi, increasedgoblet cells and columnar epithelial cells, and the gut tissue morphologywas closely similar to that of normal zebrafish  adult zebrafish was taken and shown in ebrafish .B. lactis BL-99 with concentration of2.42*107 CFU/mL (BL-99-10-7) and the model group showedcomplete separation , and the degrees of aggregations amongthe BL-99-10-7 treatment groups were obvious. There were 106 positive-ionmetabolites and 218 negative-ion metabolites were statisticallysignificantly changed in the intestines as compared between normal and themodel zebrafish; and 213 positive-ion metabolites and 402 negative-ionmetabolites with significant differences between the model zebrafish and themodel zebrafish treated with B. lactisBL-99.As shown in B.lactis BL-99 (6B). As shown in Tables B.lactis BL-99. Among the 23 significantly differentintestinal metabolites, 13 metabolites were identified with the knownphysiological and pathological functions: citrulline, glycerol,CDP-Ethanolamine, gluconolactone, uridine, uracil, taurine, mesaconic acid,ureidosuccinic acid, orotic acid, 4-hydroxybenzaldehyde,bis-\u03b3-glutamylcystine and R-lipoic acid. The biological significances forthe remaining 10 metabolites below were not known or unclear yet: SAICAR,isonicotinic acid, GDP-d-mannuronate, 3-dehydro-L-gulonate,-3-hydroxybutane-1,2,3-tricarboxylic acid, cob (I) yrinate a,cdiamide, 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol glucuronide,s-(2-chloroacetyl)glutathione,2,4-diacetamido-2,4,6-trideoxy-d-mannopyranose, andcarbamazepine-o-quinone.Heat maps of significantly changed metabolites between the intestinalfunctional disorder zebrafish (model) and normal (untreated) zebrafish wereindicated in B. lactis has been confirmed as a gastric acid andintestinal juice tolerable probiotics [B.lactis BL-99 effectively preserved in the larval zebrafishintestinal tract after 6 hrs of feeding and stayed in the intestinal tract for over24 hrs. B. lactis BL-99 promoted the intestinalmotility and relieved the constipation in aluminum sulfate-induced larval zebrafishmodel. This patented and marked probiotics increased digestive enzyme lipaseproduction, regulated inflammatory and immune responses, and relieved intestinalinflammation in an irregularly high-glucose diet-induced adult zebrafish intestinalfunctional disorder model. These findings imply that B.lactis BL-99 could be an effective and probably potentmodulator of the intestinal functions for both physiological and pathologicalconditions.obiotics  that makL. casei SY13 and exploreits effects on gut microbial structure and diversity in mice, the authors found thatthe stable colonization of L. casei SY13 wasassociated with dosage and treatment days, and thus laid a foundation for studyinginteractions between L. casei SY13 and othermembers of the gut microbiota [B. lactis BL-99 to play itsfunctions in the intestinal health and the disease prevention and treatment.Orally administered probiotics encounter various challenges on their journey throughthe mouth, stomach and intestinal tract. The health benefits of probiotics arediminished mainly due to the substantial reduction of viable probiotic bacteriaunder the harsh conditions in the gastrointestinal cavity and the colonizationresistance caused by commensal bacteria . In a prcrobiota . The lonB. lactisBL-99 promoted the intestinal motility and relief constipation and increased thedigestive enzyme lipase production in the larval and adult zebrafish models,supporting the uses of this probiotics in preventing and treating dyspepsia andmotility disorders.In normal digestion, food is transited through the gastrointestinal tract by rhythmiccontractions called peristalsis. Slow gastrointestinal contractions could lead todigestive function disorders and constipation  that areIL-1\u03b2 geneexpression, reduced intestinal immune factors IL-10 andIL-12 gene levels, lessened intestinal lipase activity, damagedintestinal histology, and disordered intestinal metabolomics. AfterB. lactis BL-99 treatment, the adult zebrafishintestinal inflammation was alleviated, the intestinal immune responses wereenhanced, and the intestinal mucus barrier and histopathology were ameliorated.Sugar consumption has dramatically increased in the past few decades  and overB.lactis BL-99 treatment recovered these intestinal andmicrobiota metabolites to the levels similar or close to the normal controlzebrafish. These results suggest that B. lactisBL-99 could relieve intestinal inflammation and promote intestinal functions,probably at least in part, through modulating intestinal and microbial metabolism tomaintain intestinal health  and 5 intestinal microbiota-related metabolites were found statistically different in the intestines betweenthe high-glucose fed and untreated control zebrafish. These 11 metabolites, plus 2organic compounds bis-\u03b3-glutamylcystine and R-lipoic acid, were all significantlyincreased in the gut of 3% glucose-fed zebrafish. Surprisedly, l health . These aS1 File(XLS)Click here for additional data file."}
+{"text": "Venenivibrio stagnispumantis strain CP.B2T is a thermophilic, chemolithoautotrophic bacterium from the family Hydrogenothermaceae (phylum Aquificota), isolated from Champagne Pool in the Waiotapu geothermal field, Aotearoa-New Zealand. The genome consists of 1.73 Mbp in 451 contigs with a 30.8 mol% G+C content. Venenivibrio, within the phylum Aquificota (family Hydrogenothermaceae), is characterized by thermophilic microaerophilic hydrogenotrophs  of M\u0101ori iwi (tribe) Ng\u0101ti Tahu-Ng\u0101ti Whaoa. The V. stagnispumantis genome has previously been sequenced as part of the Genomic Encyclopedia of Bacteria and Archaea (GEBA) project KMG-II (mana whenua (customary rights) over the bacterium, its genome, and the location in which it was isolated.The bacterial genus notrophs \u20133. The tt KMG-II   was cultivated using a modified MSH medium  within a headspace consisting of N2/H2/CO2/O2 at ratios of 50:40:7.5:2.5 (vol/vol), respectively (\u22121) of CP.B2T was performed using a NucleoSpin Soil kit  according to the manufacturer\u2019s protocols. Whole-genome sequencing  was undertaken using the Illumina HiSeq 2500 platform with TruSeq DNA Nano (550) library preparation for paired-end 101-bp reads . Default parameters were used for all assembly, quality control (QC), and annotation software unless otherwise specified. A total of 3.8-Gbp raw sequences (37.32 million reads) were assembled using SPAdes v3.12.0 (JAPEIW010000000) using the GeneMarkS-2+ protein reference set. A functionally equivalent annotation of 47 scaffolds using the Integrated Microbial Genomes annotation pipeline v4.16.4 .The V. stagnispumantis CP.B2T, Sulfurihydrogenibium yellowstonense SS-5T, and Persephonella marina EX-H1T genomes gave <80% similarity for each, confirming the designation of Venenivibrio as a separate genus to sister genera Sulfurihydrogenibium and Persephonella.A pairwise average nucleotide identity (ANI) comparison using FastANI v0.1.3  of the VV. stagnispumantis CP.B2T genome were deposited in the European Nucleotide Archive under the BioProject accession no. PRJEB55610. The assembled genome was deposited in the Genomes Online Database (Ga0591133), for associated annotation with the Integrated Microbial Genomes & Microbiomes platform (2799112217). This Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under accession no. JAPEIW010000000. The version described in this paper is version JAPEIW010000000.1.Raw sequences for the platform  (IMG gen"}
+{"text": "Enterococcus spp. lower urinary tract infections (UTI). With this rationale, our clinical microbiology laboratory discontinued routine identification on Enterococcus urine isolates in 2012 and report \u201caminopenicillins are predictably reliable for uncomplicated enterococcal UTI\u201d. The study objective was to compare outcomes of patients treated with aminopenicillins (AP) versus non-aminopenicillins (NAP) for enterococcal cystitis.Aminopenicillins achieve urinary concentrations that overcome aminopenicillin resistance in E. faecalis, enterococcal UTI in past year, review of systems unavailable/altered mental status, urinary instrumentation except for stent/catheter, systemic infection, fever, genitourinary trauma, renal transplant. Primary endpoint: clinical and microbiologic success at 14 days, defined as resolution of symptoms without new symptoms and no repeat culture growth of index organism. Logistic regression evaluated characteristics associated with 14-day failure. Sample size of 178 calculated for non-inferiority using \u03b1=0.025, \u03b2=0.8, 15% non-inferiority margin.IRB approved, retrospective cohort of adults hospitalized with symptomatic enterococcal cystitis from 2013-2021. Exclusion: definitive 178 subjects included, 89 AP, 89 NAP . VRE identified: 73 (82%) AP and 76 (85%) NAP (P=0.50). Amoxicillin  and ampicillin  used most in AP. Linezolid  and fosfomycin  most common agents in NAP. 14-day clinical composite endpoint was no different between AP and NAP groups, respectively . No variables independently predicted failure (Table\u00a01). Non-inferiority analysis for AP vs. NAP for 14-day composite: mean difference 1.1% . Definite E. faecium subgroup: 14-day clinical composite success for was no different between AP and NAP groups, respectively .E. faecium. This stewardship strategy has implications to preserve VRE active therapies (e.g. linezolid) for serious infections.AP were non-inferior to NAP, supporting their use as first-line therapy for cystitis due to enterococci, including All Authors: No reported disclosures."}
+{"text": "Ocimum tenuiflorum L. This study evaluated the antibacterial activity of O. tenuiflorum extract and its interaction with antibacterial drugs against common mastitis pathogens including Staphylococcus aureus, coagulase-negative Staphylococci (CNS), Streptococcus agalactiae, and Escherichia coli. Anti-inflammatory activities in LPS-stimulated RAW264.7 macrophage cells were also studied. The O. tenuiflorum extract exhibited antibacterial activities against S. aureus, CNS, and S. agalactiae with minimum inhibitory concentration (MIC) ranging from 3.9 to 31.2 \u00b5g/mL and minimum bactericidal concentration (MBC) ranging from 15.6 to 500 \u00b5g/mL. Combinations of O. tenuiflorum with penicillin or amoxicillin-clavulanic acid showed synergistic effects against all tested strains but an additive effect with cefazolin and gentamicin. Pretreatment of the extract significantly decreased the expression of inflammatory molecules  generated by LPS in macrophages. Results suggested O. tenuiflorum effectiveness against various Gram-positive mastitis bacteria, with the potential to reduce antibacterial doses and combat inflammation.Mastitis is the most prevalent global illness affecting dairy cows. This bacterial infection damages and inflames the udder tissues. Several plant extracts have demonstrated synergistic antibacterial activities with standard drugs in mastitis treatment. Scant information exists on  Staphylococcus aureus (S. aureus), Streptococcus agalactiae , Escherichia coli (E. coli), Enterococcus spp., and coagulase-negative Staphylococci (CNS) -bis(4-methoxy6-nitro) benzene sulfonic acid hydrate (XTT), N-methyl dibenzopyrazine methyl sulfate (PMS) were obtained from Sigma-Aldrich . The 2x qPCRBIO SyGreen 1step Lo-ROX was obtained from PCR Biosystems . Tri-RNA Reagent was purchased from Favorgen . IL-6 and PGE2 quantitative sandwich ELISA kits were obtained from Abcam . The Griess reagent was purchased from Promega . All other reagents were obtained from Sigma-Aldrich unless otherwise describedLipopolysaccharide (LPS) from O. tenuiflorum were cleaned with distilled water twice. Then, 500 g of sample was dried at 60 \u00b0C and then powdered into fine powder. Two liters of 70% ethanol were added to powdered dried plants and allowed to remain for 24 h at room temperature. The mixture was filtered and evaporated using a rotary evaporator to eliminate the ethanol. The liquid was subsequently freeze-dried and stored at \u221220 \u00b0C until used.Leaves of v/v) formic acid). Authentic standards consisted of syringic acid (>97.0% T), sinapic acid , quercetin , naringenin , myricetin (>97.0% HPLC), luteolin (>98.0% HPLC), kaempferol (>97.0% HPLC), hydroxybenzoic acid , 4\u2013hesperidin , genistein (>98.0% HPLC), ferulic acid , (\u2212)-epigallocatechin gallate (>98.0% HPLC), 3,4-dihydroxybenzoic acid (\u226597% T), p-coumaric acid , cinnamic acid (>98.0% HPLC), chlorogenic acid , caffeic acid  and apigenin (>98.0% HPLC) from Tokyo Chemical Industry , vanillic acid (\u226597% HPLC), rutin (\u226594% HPLC), rosmarinic acid (\u226598% HPLC) and gallic acid (97.5\u2013102.5% T) from Sigma-Aldrich , galangin (\u226598.0% HPLC) from Wuhan ChemFaces Biochemical Co., Ltd.  and isorhamnetin (\u226599.0% HPLC) from Extrasynthese . The LC\u2013ESI-MS/MS chromatograms were shown in The phytochemical profile was analyzed using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) with the conditions and validation following a well-established protocol as previously reported  without S. aureus, S. agalactiae, CNS, and E. coli from clinical bovine mastitis cases were obtained from milk samples and collected from the Dairy Farming Promotion Organization of Thailand (D.P.O.), Saraburi, Thailand. Microorganism isolates were characterized based on their culture, morphological (Gram stain), and biochemical properties and sub-cultured on the selective medium following standard microbiological techniques -bis (4-methoxy-nitro) benzene sulfonic acid hydrate (XTT)-based assay to determine the concentration of 5 cells/well) or 6-well plates (1 \u00d7 106 cells/well) and then pretreated for 12 h with media containing different concentrations of O. tenuiflorum extract  before stimulation with LPS (10 ng/mL) for 24 h. Culture supernatants were collected from 96-well plates for interleukin-6 (IL-6), nitric oxide (NO), and prostaglandin E2 (PGE2). Cells were harvested from the 96 or 6-well plates for qPCR analysis.RAW 264.7 cells were cultured overnight in 96-well plates (1 \u00d7 10v/v) ratio with Griess reagent -ethylenediamine). After incubation for 10 min at room temperature, absorbance was measured at 540 nm using a microplate reader . A nitrite standard curve was used to calculate nitrite concentration in the supernatant.Nitric oxide generation was assessed by measuring the quantity of nitrite in the culture medium based on the Griess reaction . FollowiO. tenuiflorum extract on LPS-induced RAW 264.7 cell inflammation, the relative mRNA levels of IL-6, tumor necrosis factor-\u03b1 (TNF-\u03b1), interleukin-1\u03b2 (IL-1\u03b2), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were measured by RT-qPCR. Total RNA was extracted by homogenizing the cells in Tri-RNA Reagent according to the manufacturer\u2019s instructions. The RT-qPCR reaction mixture, qPCRBIO SyGreen 1-step Lo-ROX , was made according to the manufacturer\u2019s guidelines. Quantitative polymerase chain reaction (qPCR) was performed by qTOWER3 Real-Time PCR Systems . Thermal cycling conditions were used to amplify the target genes using the following parameters: reverse transcription step at 45 \u00b0C for 10 min, polymerase activation step at 95 \u00b0C for 2 min, denaturation step at 40 amplification cycles at 95 \u00b0C for 5 s, and the final step at 60 \u00b0C for 30 s. Relative levels of gene expression were normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA using the 2\u2212\u0394\u0394CT method [To investigate the effects of The levels of IL-6 and PGE2 in RAW 264.7 cell culture supernatants were measured using an enzyme-linked immunosorbent assay (ELISA) kit obtained from Abcam  and performed according to the manufacturer\u2019s instructions.p \u2264 0.05) was considered statistically significant.Statistical analysis was performed using GraphPad Prism ver. 5 software. The values are presented as mean \u00b1 standard deviation (SD) from at least three independent replicates. Statistical significance between groups was determined using a one-way ANOVA followed by Tukey\u2019s test. Statistical significance between groups was determined using a one-way ANOVA followed by Tukey\u2019s test. A probability of 0.05 or less (O. tenuiflorum ethanolic extract as follows: (i) O. tenuiflorum had antibacterial effects against Gram-positive bacteria including S. aureus, CNS, and S. agalactiae but not Gram-negative bacteria, (ii) O. tenuiflorum extract showed synergistic effects with penicillin or amoxicillin-clavulanic acid against all tested strains, while cefazolin and amikacin had an additive effect, and (iii) O. tenuiflorum extract showed anti-inflammatory activities, with reduced expression of inflammatory molecules in LPS-treated macrophages. However, further investigations on route and dosage for therapy in animal models are required.This study explored new antibacterial candidates from plants with multi-target abilities in mastitis treatment, covering antibacterial and anti-inflammation activities. Results demonstrated promising anti-mastitis properties of"}
+{"text": "JCI Insight. 2019;4(13):e125191. https://doi.org/10.1172/jci.insight.125191Original citation: JCI Insight. 2022;7(20):e165600. https://doi.org/10.1172/jci.insight.165600Citation for this corrigendum: The authors recently became aware that representative illustrations presented in n-3 DPA (n-3 DPA), including the gut-protective RvD5n-3 DPA (27) (Using liquid chromatography\u2013tandem mass spectrometry\u2013based (LC-MS/MS\u2013based) LM profiling, we identified mediators from all 4 major fatty acid bioactive metabolomes, including lipoxygenase- and cyclooxygenase-derived LMs that were identified in accordance with published criteria (33). The identity of each of the mediators was further corroborated by evaluating MS/MS spectra obtained in a subset of the samples analyzed and matching at least 6 diagnostic ions with those obtained for reference standards for each of these molecules, as shown for RvD5n-3 DPA  (23). Mun-3 DPA . This sh"}
+{"text": "Dear Editor,1 However, its function in subverting host translation machinery is still elusive.COVID-19 (Coronavirus Disease 2019) is causing an unprecedented public health crisis. Protein translation is crucial for virus lifecycle. Nucleocapsid (N) protein is among the most abundant SARS-CoV-2 proteins and highly conserved across coronavirus genus.2 , indicating that N mRNAs were much less efficiently translated after NPM1 depletion Fig. . SimilarIn IVT assays, ribosomes from 293T cells with exogenous N showed enhanced translation efficiency when translating N mRNAs, which was suppressed upon NPM1 knockdown Fig. , top row3 We therefore hypothesized that N protein functions via NPM1-binding snoRNAs. We first re-analyzed snoRNA expression in different tissues using public data from ENCODE,4 and found significant overlap of expression profile among lung, liver, and intestine (16/top 20, Supplementary Fig. 3 and their expression was highly correlated with ACE2, TMPRSS2, FURIN (Fig. 5 (Fig. NPM1 has been shown to function as RNA/DNA binding protein regulating 2\u2032-O-methylation of rRNA via direct binding of snoRNAs.RIN Fig. . Exploit. 5 Fig. . This wa. 5 Fig. . Besides. 5 Fig. . Knockdo. 5 Fig. , h. Besi. 5 Fig. .SNORD93 was predicted to target A576 on 18S rRNA for 2\u2032-O-Me modification Fig. , which iWe next explored strategy for SARS-CoV-2 prevention via targeting N protein-related translation machinery components. SARS-CoV-2 infection also enhanced 2\u2032-O-Me modification, as was detected by RTL-P assay (Supplementary Fig. In summary, we found that SARS-CoV-2 N protein enhances host translation efficiency by potentiating the host NPM1-snoRNA translation machinery. By interacting with NPM1 and snoRNAs, N protein significantly promotes ribosome translation efficiency, particularly for viral mRNAs, via enhancement of snoRNA-mediated 2\u2032-O-methylation on rRNAs (Supplementary Fig. Supplementary information"}
+{"text": "Gordonia rubripertincta NRRL B-16540. Survivors has a 45,436-bp genome encoding 69 predicted protein-coding genes, of which 32 have assigned functions. Based on gene content similarity to sequenced actinobacteriophages, Survivors is assigned to phage cluster CT.Bacteriophage Survivors is a siphovirus isolated from  Gordonia bacteria are opportunistic human pathogens with varied, important roles in bioremediation , using standard protocols from the Science Education Alliance-Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) Phage Discovery Guide . The soirphology . The caphttps://phagesdb.org/media/workflow/protocols/pdfs/PCI_SDS_DNA_Extraction_2.2013.pdf), prepared for sequencing using the New England BioLabs Ultra II library kit, and sequenced at the Pittsburgh Bacteriophage Institute using Illumina MiSeq (v3 reagents). There were 236,073 150-base single-end reads providing 735\u00d7 coverage. The raw reads were assembled and checked for completeness using Newbler v2.9 and Consed v29, respectively, using default settings (CGGTAGGCAT-3\u2032). Genome termini were determined through similarity to known phages, an analysis of read start buildups, and coverage levels across the genome (http://phagesDB.org), Survivors was assigned to cluster CT  (:1 (httpshttp://cobamide2.bio.pitt.edu), Glimmer (v3.02b) (http://phages.wustl.edu/starterator/), Phamerator (Actino_Draft v462)  (ft v462) , BLASTp ft v462) , HHPred ft v462) , ARAGORNft v462) , tRNAscaft v462) , TMHMM (ft v462) , and SOSft v462) . DefaultON970576 and has been assigned Sequence Read Archive (SRA) no. SRX14485091.Survivors GenBank accession no. is"}
+{"text": "Stolephorusbengalensis  or Stolephorusinsularis Hardenberg, 1933, revealed four distinct species, true S.bengalensis  and three new species, viz., Stolephoruseldoradosp. nov. , Stolephorusdiabolussp. nov.  and Stolephoruseclipsissp. nov. . Characters separating the four species include numbers of gill rakers on each gill arch and vertebrae and pelvic fin and dorsal-fin ray lengths. Two molecular markers  demonstrated the distinction of three of the species examined morphologically and enabled a reconstruction of their phylogenetic relationships. Each species was genetically divergent from the others by 3.5%\u20137.7% mean uncorrected distance in the mitochondrial cytochrome oxidase I gene.Examination of numerous specimens characterised by predorsal scute, long maxilla, indented preopercle and pelvic scute lacking a spine and previously identified as  Stolephorus Lacep\u00e8de, 1803 (Teleostei: Clupeiformes: Engraulidae), diagnosed by the presence of prepelvic scutes and an embedded urohyal and lack of postpelvic scutes, currently includes 37 valid species that preferentially inhabit marine and/or estuarine waters in the Indo-Pacific region S.insularis as Stolephorusbengalensis  and regarding the nominal species S.insularis as a junior synonym of Stolephorustri . However, subsequent re-examination of specimens, identified as S.bengalensis, in fact revealed the presence of four species.The anchovy genus c region . AmongstS.bengalensis and describe three new species of Stolephorus from specimens previously regarded as S.insularis or S.bengalensis. In addition to the morphological comparisons, complete mitochondrial cytochrome b gene and partial mitochondrial cytochrome oxidase I (COI) gene sequences from 31 specimens were used to estimate the genetic distinction of three of the latter (the fourth species unavailable) plus one unidentified, but related species from Segara Anakan Lagoon, Central Java, Indonesia  cytochrome b gene and partial (648 bp) COI gene. The cytochrome b gene sequences were published in COI gene was newly sequenced for 19 specimens of S.eldorado, including the holotype and several paratypes (Table COI sequences (available in GenBank) of S.diablocus , S.bengalensis (eight specimens from India), S.eldorado   species of PCR) amplification and sequencing of the COI gene followed standard protocols (COI gene used the following primers: forward COI_FishF1 (5\u2019-TCA ACC AAC CAC AAA GAC ATT GGC AC-3\u2019) and reverse COI_FishR2 (5\u2019-ACT TCA GGG TGA CCG AAG AAT CAG AA-3\u2019)  . PCR prob and COI sequences were determined separately by eye, requiring neither insertions nor deletions. The final alignment combining the two genes (for 31 specimens plus one outgroup) comprised 1788 nucleotide positions. Uncorrected pairwise genetic distances (i.e. p-distances) amongst species were calculated with MEGA X (ML) method of phylogenetic reconstruction using the general time-reversible model of nucleotide substitution with rate heterogeneity following a discrete gamma distribution (GTR + \u0413), using the software RAxML-NG (S.acinaces and the robustness of each node determined by bootstrap support (500 replicates).Alignments of the cytochrome h MEGA X . The relRAxML-NG  as impleRAxML-NG . The treTaxon classificationAnimaliaClupeiformesEngraulidae\ufeff67E69026-8C76-5B86-84F8-C5E877CDE39CAnchoviellabaganensisbengalensis Dutt & Babu Rao, 1959: 160  and the anal-fin origin located below the origin of the second to sixth dorsal-fin ray (vs. eighth to eleventh)  in having a predorsal scute and double pigment lines on the dorsum behind the dorsal fin, but differ in having deciduous body scales  and lacking a spine on the pelvic scute (pelvic scute with a hard posteriorly projecting spine)  . Stolephg spine) . CompariStolephorusbengalensis, S.diabolus sp. nov. and S.eldorado sp. nov. were divergent from each other by at least 3.5% COI-based mean uncorrected genetic distance (min-max = 3.5\u20137.7%) , forming clear intraspecific versus interspecific genetic gaps. The ML phylogenetic tree using COI and cytochrome b markers : IPMB-I 13.00001, 49.7 mm SL, Teluk Bahang, Penang, Malaysia.USMFC 82-0017, 43.7 mm SL, collected with the holotype; USMFC 82-0057, 4 specimens, 40.1\u201341.1 mm SL, estuary of Merbok River, Jeti Semeling, Malaysia; ZUMT 62056, 5 specimens, 28.5\u201338.4 mm SL, KAUM\u2013I. 163702, 36.3 mm SL, KAUM\u2013I. 163703, 36.4 mm SL, NSMT-P 143554, 36.4 mm SL, NSMT-P 143555, 36.6 mm SL, Singapore.14 specimens, 28.5\u201343.7 mm SL. Stolephorus with the following combination of characters: 1UGR 14\u201316 , 1LGR 20\u201323 (22), 1TGR 35\u201338 (38); 2UGR 10 or 11 (11), 2LGR 19 or 20 (20), 2TGR 30 or 31 (31); 3UGR 8 or 9 (9), 3LGR 11 or 12 (12), 3TGR 20 or 21 (21); 4UGR 6 or 7 (7), 4LGR 9 or 10 (9), 4TGR 15\u201317 (17); prepelvic scutes 5\u20137 (6); total vertebrae 39; long maxilla, posterior tip just reaching or slightly short of posterior margin of opercle; predorsal scute present; pelvic scute without spine; body scales deciduous; posterior border of pre-opercle concave, indented; paired dark patch on parietal area with little following pigmentation; distinct double pigment lines along dorsum posterior to dorsal fin; black spots below eye and on lower-jaw tip absent; anal-fin base long, 19.8\u201322.3% (mean 20.7%) of SL; maximum orbit diameter 8.1\u20138.7% (8.3%) of SL; third dorsal-fin ray short, 17.0\u201318.5% (18.0%) of SL; pelvic fin rather long, 9.6\u201311.3% (10.0%) of SL, its posterior tip not reaching to vertical through dorsal-fin origin when depressed in specimens > 40 mm SL; distance between posterior ends of supramaxilla and maxilla 5.7\u20136.4% (6.1%) of SL.A species of Data for holotype presented first, followed by data for paratypes in parentheses (if different). Counts and measurements, expressed as percentages of SL or HL, given in Tables Body uniformly pale white. A pair of distinct dark patches on parietal region, with little pigmentation on occipital area. No black spots below eye and on lower-jaw tip. Melanophores scattered on posterior margins of scale pockets on dorsum. Double pigmented lines dorsally posterior to dorsal fin. Melanophores scattered along bases of dorsal and anal fins. All fins transparent, melanophores scattered along fin rays of caudal fin and anterior parts of dorsal and anal fins.Stolephorusdiabolus sp. nov. is currently known only from the western coast of the Peninsular Malaysia (Merbok River Estuary and Penang) and Singapore ; 2TGR, 30 or 31 in S.diabolus [vs. 33 or more (rarely 30 or 31 in S.eldorado)]; 3TGR, 20 or 21 in S.diabolus [vs. 22 or more in the other three species (rarely 21 in S.eldorado)]; and 4TGR, 15\u201317 in S.diabolus (vs. 17 or more)  of SL in S.diabolus vs. 8.2\u20139.9% (8.9%) in S.eldorado; Fig. S.diabolus is distinguished from S.bengalensis by having a shorter third dorsal-fin ray , shortesis Fig.  and fewesis Fig. ."}
+{"text": "Bartonella spp. comprises emergent and re-emergent fastidious Gram-negative bacteria with worldwide distribution. Cats are the main reservoir hosts for Bartonella henselae and dogs represent opportunistic hosts for the bacteria. Even though ticks may also play a role in transmission, their competence as vectors for Bartonella spp. has not been totally understood. Considering only a few studies had a focus on screening Bartonella in animals, humans and ectoparasites in Portugal, this study aimed to address the molecular occurrence of Bartonella sp. in 123 stray cats, 25 stray dogs, 30 humans from Lisbon and 236 questing ticks within the country. Using a qPCR targeting the nuoG gene, it was possible to detect Bartonella sp. DNA on 20.32% of cat samples (25/123). From these positive samples, 13 sequences were characterized as B. henselae, 11 as B. clarridgeiae and 1 presented co-infection with both species. The absolute quantification of nuoGBartonella DNA in sampled cats ranged from 2.78 \u00d7 10 to 1.03 \u00d7 105 copies/\u00b5L. The sampled dogs, humans and ticks were negative. These results showed that B. henselae and B. clarridgeiae are circulating in stray cats from Lisbon. Additional and more extended studies should be conducted to determine the impact of such infections on humans, particularly those in constant and direct contact with cats. Bartonella spp. encompasses small, fastidious, and facultative Gram-negative intracellular bacteria with a global distribution -3\u2032 [\u00ae (IDT-Integrated DNA Technologies) encompassing the 83 bp B. henselae-nuoG gene fragment. The number of copies was determined by the following formula: Xg/\u03bcL DNA/[fragment length in bp \u00d7 660]) \u00d7 6.022 \u00d7 1023 \u00d7 plasmid copies/\u03bcL.A quantitative PCR targeting a fragment of the NADH ubiquinone oxireductase subunit G (BHQ1]-3\u2032  was carrnuoG-based qPCR gene were afterwards used in two cPCR assays. The citrate synthase gene (gltA) was targeted for amplification of a 767 bp fragment, using the primers 443F: 5\u2032-GCTATFTCTGCATTCTATCA-3\u2032 and 1210-R: 5\u2032-GATCYTCAATCATTTCTTTCCA-3\u2032 [ribC), using the primers BARTON-1: 5\u2032-TAACCGATATTGGTTGTGTTGAAG-3\u2032 and BARTON-2: 5\u2032-TAAAGCTAGAAAGTCTGG CAACATAACG-3\u2032 [gltA cPCR assay and cycling conditions consisted of an initial denaturation at 95 \u00b0C for 5 min, 35 cycles consisting of denaturation at 94 \u00b0C for 30 s, annealing at 55 \u00b0C for 30 s and extension at 72 \u00b0C for 1 min, followed by a final extension at 72 \u00b0C for 5 min. Ultrapure water was used as a negative control. All assays were conducted in a T100\u2122 Thermal Cycler . PCR products were analyzed by horizontal electrophoresis in 1.5% agarose gel stained with Green Safe Premium (NZYTech). Positive amplicons for both cPCR targeting the genes gltA and ribC were purified using the NZYGelpure kit (NZYTech) and Sanger sequenced at StabVida .Positive samples for the TTCCA-3\u2032 . The 25 TAACG-3\u2032 . ReactioE. coli competent cells (NZYTech) according to standard cloning protocols. Briefly, the purified amplified product was ligated into the plasmid pTZ57R/T. Chemically competent E. coli cells (NZYtech) were transformed, plated in LB agar (NZYtech) and incubated at 37 \u00b0C, overnight. Positive colonies were subcultured overnight on 3 mL of LB medium  at 37 \u00b0C and under 200 rpm of shaking. Finally, plasmid purification was carried out using the NZY Miniprep kit (NZYtech) following the manufacturer\u2019s recommendations, and 10 \u00b5L of purified samples together with 3 \u00b5L of M3 forward primer were sent to StabVida , for sequencing. Sequences were manually trimmed and submitted to BLAST [B. henselae gltA, B. clarridgeiae gltA, B. henselae ribC and B. clarridgeiae ribC), and a ClustalW multiple alignment was performed in Bioedit for each group of gene sequences. The number of haplotypes (h) and the haplotype diversity (Hd) was evaluated using the software DNA Sequence Polymorphism (DnaSP) v.6.12.03 [p-distance model with 1000 bootstrap replications. For the phylogenetic trees\u2019 construction, ClustalW multiple alignment was performed in Bioedit for each group of gene sequences (gltA and ribC), including representative sequences for each haplotype obtained in this study, together with other sequences selected from the GenBank (for gltA: Bartonella acomydis (AB444979.1), Bartonella alsatica (AF204273.1), Bartonella birtlesii (AF204272.1), Bartonella vinsonii subp. vinsonii (Z70015.1), Bartonella vinsonii subsp. arupensis (AF214557.1), Bartonella jaculi (AB444975.1), Bartonella rattaustraliani (EU111796.1), Bartonella japonica (AB242289.1), Bartonella coopersplainsensis (EU111803.1), Bartonella quintana (HQ014627.1), Bartonella koehlerae (AF176091.1), Bartonella henselae Human Australia (AJ439406.1), Bartonella henselae (L38987.1), Bartonella henselae Cat Flea Austria (MF374384.1), Bartonella henselae Cat Brazil (MH019304.1), Bartonella henselae Dog Chile (MG252490.1), Bartonella doshiae (AF207827.1), Bartonella rattimassiliensis (AY515124.1), Bartonella queenslandensis (EU111798.1), Bartonella elizabethae (Z70009.1), Bartonella schoenbuchii (AJ278184.1), Bartonella chomelii (AY254308.1), Bartonella capreoli (AF293392.1), Bartonella rochalimae (FN645459.1), Bartonella clarridgeiae Cat flea Chile (KY913636.1), Bartonella clarridgeiae Cat Brazil (MH019302.1), Bartonella clarridgeiae Cat Thailand (KX001761.1), Bartonella clarridgeiae (EU770616.1), Bartonella clarridgeiae strain 73 (FN645454.1), Bartonella baciliformis (DQ452947.1), Bartonella tamiae (DQ395177.1) and Brucella abortus (AE017223.1); and for ribC: Bartonella henselae Cat Brazil (HQ012583.1), Bartonella henselae fleas USA (AY953284.1), Bartonella henselae Cat Brazil (HM588661.1), Bartonella koehlerae (FJ832090.1), Bartonella quintana (AJ236917.1), Bartonella washoensis subsp. cynomysii (DQ825697.1), Bartonella heixiaziensis (KJ361664.1), Bartonella vinsonii subsp. arupensis (AY116631.1), Bartonella vinsonii subsp. vinsonii (AY116636.1), Bartonella vinsonii subsp. berkhoffii (AY116629.1), Bartonella alsatica (AY116630.1), Bartonella fuyuanensis (KJ361648.1), Bartonella doshiae (AY116627.1), Bartonella sp. Bat Kenya (HM363783.1), Bartonella bovis riboflavin synthase France (AY116637.1), Bartonella chomelii (AB290195.1), Bartonella capreoli (AB290194.1), Bartonella schoenbuchensis (AY116628.1), Bartonella bacilliformis (AJ236918.1), Bartonella clarridgeiae Cat China (EU571943.1), Bartonella clarridgeiae (AJ236916.1), Bartonella clarridgeiae Japan (AB292604.1), Bartonella clarridgeiae Cat Brazil (KR092386.1), Bartonella ancashensis (KP720649.1), Candidatus Bartonella ancashi Peru (KC886734.1), Brucella melitensis (CP008750.1)). No criteria regarding host or geographical localization were used to retrieve sequences from GenBank. Likelihood-mapping analyses were performed using the TREE-PUZZLE v5.3 program [Sequences were analyzed and manually cured using Bioedit Sequence Alignment Editor , in ordeto BLAST . Edited .6.12.03 . The hap.6.12.03  was usedlerae AF1091.1, BaBartonella spp. was not detected in ticks, dogs, and humans in the present study, but more studies need to be performed addressing this topic and putting emphasis on Bartonella spp. reservoirs and vectors. Regarding the dogs, surveys among endocarditis afflicted individuals can be carried out and similarly, studies conducted on humans should prioritize immune-compromised groups, as they are more prone to develop infections by Bartonella spp. About 20% of the cats sampled in Lisbon were positive for at least one Bartonella spp. and both B. henselae and B. clarridgeiae were identified in similar percentages. It is hoped that these results could bring attention to Bartonella sp. as an emerging pathogen in cats from Lisbon, and a potential threat for public health. As they are the main reservoir for B. henselae, preventive measurements should be implemented in order to avoid an outbreak of cat scratch disease in Lisbon.Bartonellosis is recognized as an emerging vector-borne disease that constitutes a threat to both animal and human health. Continuous and active surveillance to understand the epidemiological characteristics of bartonellosis is needed worldwide, including in Portugal."}
+{"text": "Escherichia coli is a reservoir of antimicrobial resistance genes (ARGs). Here, we report the draft genome sequence of an E. coli strain (31HGR-CBG) that was isolated from a urine sample in Tamaulipas, Mexico. 31HGR-CBG harbors multiple ARGs, including blaCTX-M-15 and class 1 integron. This strain also carries multiple virulence genes. Escherichia coli represents a threat to public health (E. coli (ExPEC) are genetically diverse and complex  approval nor informed consent was required. 31HGR-CBG was grown on Trypticase soy agar and CHROMagar Orientation medium and incubated overnight at 37\u00b0C. Standard biochemical tests were also performed.https://www.bioinformatics.babraham.ac.uk/projects/fastqc) and Trim Galore! v0.6.6  were used to evaluate quality and to trim the raw reads, respectively. Assembly was performed using SPAdes v3.15.2 (https://github.com/ablab/quast). Contigs smaller than 900\u2009bp were removed. Bacterial identification was confirmed by a BLASTN search (http://blast.ncbi.nlm.nih.gov) against the NCBI database and ribosomal multilocus sequence typing (rMLST)  at 37\u00b0C. DNA was extracted using the Wizard genomic DNA purification kit . DNA quantification was performed with the Qubit double-stranded DNA (dsDNA) HS assay kit in the Qubit 3.0 fluorometer . Libraries were constructed with the Nextera Flex library preparation kit and were sequenced using the MiniSeq sequencing system (150-bp paired-end reads). A total of 10,918,316 raw reads were generated. FastQC v0.11.3 (http://genomicepidemiology.org). ARGs were also predicted with the CARD v3.1.4 database using RGI v5.2.0 (Automatic annotation was performed by the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v5.2. Multilocus sequence typing (MLST) was executed with PubMLST v1 . SerotypI v5.2.0 . Phages I v5.2.0 . DefaultN50 value of 91,860\u2009bp, a GC content of 50.82%, and genome coverage of 220\u00d7. Annotation identified 4,939 genes, 4,850 coding sequences, and two CRISPR arrays. 31HGR-CBG belongs to sequence type 44 (ST44) and serotype 0101:H4.The genome was 4,981,584\u2009bp in size and was assembled into 141 contigs, with an aadA5, aph(6)-Id, aac(6\u2032)-Ib-cr, and aph(3\u2033)-Ib], extended-spectrum \u03b2-lactams (blaCTX-M-15 and blaOXA-1), phenicol (catB3), macrolides [mph(A)], sulfonamides (sul1 and sul2), tetracycline [tet(B)], trimethoprim (dfrA17), and quaternary ammonium compounds (qacE\u03941). In addition, plasmid replicons IncFIA, IncFIB, and IncFII were detected.Multiple ARGs were identified, including genes conferring resistance to aminoglycosides [gyrA, parC, and parE were found, indicating fluoroquinolone resistance. Virulence-associated genes and four intact prophages were identified.Chromosomal mutations in JAKJKJ000000000. The reads were deposited in the Sequence Read Archive (SRA) under the accession number SRR15258840.This draft genome has been deposited in GenBank under the accession number"}
+{"text": "Spinirta Jin & Zhang, 2020 (Araneae: Corinnidae) from Jiangxi Province, China are described here: S.sanxiandiansp. nov. (\u2642\u2640) and S.sishuishansp. nov. (\u2642). Detailed descriptions and photographs of the new species are provided.Two new species of  Corinnidae Karsch, 1880 has increased greatly in the past ten years . Measurements were taken with the AxioVision software (SE64 ver. 4.8.3) and are given in millimetres. Terminology of the male and female copulatory organs follows Specimens were examined using a Jiangnan SZ 6100 stereomicroscope with a Zoom Microscope System. Both male palps and female copulatory organs were detached and examined in 80% ethanol, using an Olympus CX43 compound microscope with a KUY NICE CCD camera. All specimens treated in this work are deposited in the Animal Specimen Museum, Life Science of College, Jinggangshan University . The abbreviations used in the text and figures are:ALE anterior lateral eye;AME anterior median eye;At atrium;CD copulatory duct;CO copulatory opening;CS cone-shaped spines;d dorsal;E embolus;EA embolic apophysis;FD fertilization duct;GA glandular appendage;MOA median ocular area;p prolateral;PLE posterior lateral eye;PME posterior median eye;PTA prolateral tibial apophysis;r retrolateral;RTA retrolateral tibial apophysis;Sp spermatheca;St subtegulum;v ventral;VTA ventral tibial apophysis.Taxon classificationAnimaliaAraneaeCorinnidae\ufeffGenusJin & Zhang, 2020921B6204-15E0-54B8-B299-06A67E4FBE76Spinirtajinyunshanensis Jin & Zhang, 2020. Type locality: Chongqing.Spinirta species, S.wuyishanensis Zhou, 2022 was recorded from Jiangxi Province in southeast China. It is worth mentioning that the female remains unknown.The genus includes 11 species, all of which are distributed in southern and southwest of China (WSC 2022). Currently, most of them are known only from females (three species) or males (four species) (WSC 2022). Most of China\u2019s nine species are recorded from southwestern China . Only onTaxon classificationAnimaliaAraneaeCorinnidae\ufeffLiusp. nov.E398719E-F60C-5419-B24C-A02AFD3BF57Ehttps://zoobank.org/6CEBCD02-A0B5-452B-A1AD-E0F681296427Holotype: 1 \u2642, China: Jiangxi Province, Ji\u2019an City, Qingyuan District, Donggu Town, Dawu Mountain, 26\u00b040'48.69\"N, 115\u00b025'7.79\"E, 1031 m, 25.X.2020, K. Liu et al. leg. (Cor-04). Paratype: 2 \u2640, 13.XI.2021, K. Liu et al. leg., other data same as holotype (Cor-03 and Cor-05).The specific name is derived from the type locality, Sanxiandian Temple in Dawu Mountain; noun in apposition.Spinirtasparsula Jin & Zhang, 2020  with a curved posterior part (vs. straight in S.sparsula) and the ear-shaped retrolateral tibial apophysis (RTA) without protruded base (vs. digitiform with a kidney shaped protruded base in S.sparsula). It also resembles S.sishuishan sp. nov. in having a thumb-like ventral tibial apophysis (VTA), a thick horn-like prolateral tibial apophysis (PTA) and a curved sperm duct (SD), but can be separated from it by the ear-shaped retrolateral tibial apophysis (RTA) (vs. shield-shaped in S.sishuishan sp. nov.), the anterior part of the tegulum with a broad lateral apophysis (vs. absent in S.sishuishan sp. nov.) and the sharp embolic apophysis in retrolateral view (vs. relatively blunt in S.sishuishan sp. nov.)  , the shield copulatory openings (CO) (vs. round in S.qizimeiensis), and the copulatory ducts (CD) extending from the anteromedial to the posterolateral part of the epigyne .The male of this new species is similar to that of cf. Fig. . The femMale. Habitus as in Fig. AME 0.32, ALE 0.31, PME 0.2, PLE 0.27, AME-AME 0.15, AME-ALE 0.07, PME-PME 0.3, PME-PLE 0.36, AME-PME 0.25, AME-PLE 0.47, ALE-ALE 0.87, PLE-PLE 1.45, ALE-PLE 0.2. MOA 0.74 long, front width 0.77, back width 0.72. Chelicera with three promarginal and five retromarginal teeth  thumb-like in ventral view. Retrolateral tibial apophysis (RTA) ear-shaped, nearly as long as tibial length, ventral surface with two lines of short cone-shaped spines (CS). Prolateral tibial apophysis (PTA) thick horn-like, strongly sclerotised, nearly as long as 1/3 of tibia. Tegulum with strongly sclerotized apex. Subtegulum (St) with many wrinkles on posterior surface. Sperm duct (SD) S-shaped in posterior part. Embolus (E) short, with thick base, forming a C-shape with short spine-like embolic apophysis (EA), nearly 3\u00d7 longer than embolic apophysis.Palp as in Fig. Female. Habitus as in Fig. AME 0.28, ALE 0.26, PME 0.19, PLE 0.24, AME-AME 0.16, AME-ALE 0.08, PME-PME 0.27, PME-PLE 0.31, AME-PME 0.26, AME-PLE 0.4, ALE-ALE 0.83, PLE-PLE 1.3, ALE-PLE 0.18. MOA 0.72 long, front width 0.66, back width 0.66. Abdomen: 5.55 long, 3.85 wide. Leg measurements: I 13.57 ; II 12.5 ; III 11.2 ; IV 14.77 ; spination  large, shield, covers equal or less than half of epigynal plate, anteromedially located. Copulatory openings (CO) very large, oval, located at anterolateral atrium. Copulatory ducts (CD) very broad, anteriorly touching, posteriorly slightly separated. Glandular appendages (GA) short, located at dorsal part of copulatory ducts, extending beyond medial part of copulatory ducts, directed anteriorly. Spermathecae (Sp) relatively broad, separated by 1/2 width of copulatory ducts. Fertilisation ducts (FD) directed anteriorly, shorter than spermathecal width.Epigyne as in Figs Spinirtasanxiandian sp. nov. may be the result of the influence of their development factors.The female specimens of this new species occur exactly in the same sites explored by the authors. They are identified as the same species based on appearance and epigyne. Variability was observed in the epigyne Fig. , which mKnown only from the type locality, Jiangxi Province, China Fig. .Taxon classificationAnimaliaAraneaeCorinnidae\ufeffLiusp. nov.0FA58825-87CA-51AD-843C-D1560BD78095https://zoobank.org/F06E9311-8FE0-4858-B870-22E9F59F3262Holotype: 1 \u2642, China: Jiangxi Province, Ganzhou City, Chongyi County, Sishui Mountain, near parking lot, 25\u00b027'11.73\"N, 113\u00b055'30.04\"E, 965 m, 2.X.2020, K. Liu et al. leg. (Cor-02).The specific name, derived from the type locality, is a noun in apposition.S.sanxiandian sp. nov. by the shield retrolateral tibial apophysis (RTA) (vs. ear-shaped), the anterior part of the tegulum lacking lateral apophysis (vs. present in S.sanxiandian sp. nov.) and the relatively blunt embolic apophysis (EA) in retrolateral view (vs. sharp in S.sanxiandian sp. nov.)  thumb-like in ventral view. Retrolateral tibial apophysis (RTA) shield in retrolateral view, nearly as long as tibial length, ventral surface with four lines of short cone-shaped spines (CS). Prolateral tibial apophysis (PTA) thick horn-like, strongly sclerotised, nearly as long as 1/3 of tibia. Tegulum with strongly sclerotized apex. Subtegulum (St) with many wrinkles on posterolateral tegulum. Sperm duct (SD) S-shaped in posterior part. Embolus (E) spine-like, with thick base, forming a C-shape with short blunt embolic apophysis (EA), nearly 4\u00d7 longer than embolic apophysis.Palp as in Fig. Female. Unknown.Known only from the type locality, Jiangxi Province, China Fig. ."}
+{"text": "This study aims to investigate the equivalence between Cambridge Neuropsychological Test Automated Battery (CANTAB) and Montreal Cognitive Assessment, Changsha version (MoCA-CS) as cognition measures for Chinese stroke survivors. Sixteen stroke survivors were recruited from stroke center in Shanghai, China. Participants completed Paired Associates Learning (PAL) task in CANTAB, MoCA-CS and questionnaire about demo-social characteristics. Pearson correlation and hierarchical regression were used for equivalence analysis. CANTAB-PAL task performances\uff08PAL First Attempt Memory Score, PALFAMS, PAL Total Attempts 2 Patterns, PALTA2\uff09were significantly associated with score of MoCA-CS and delayed recall (\u03b2=0.528, p=0.014; \u03b2=0.198, p=0.043; \u03b2= -3.885\uff0cp=0.017; \u03b2= -1.600,p=0.026, respectively\uff09after controlling for age, gender and education. The results suggest CANTAB-PAL is a reliable tool for measuring Chinese stroke survivors\u2019 cognitive function. Future studies are needed to establish the equivalence of CANTAB in multi-tasks in larger sample of Chinese stroke survivors."}
+{"text": "Escherichia coli (ETEC) strain harboring genes encoding colonization surface antigen 13 (CS13) and a heat-labile toxin. The ESEI_597 strain was isolated from an 8-month-old child living in Korogocho, Kenya, in 2013.Here, we report the draft genome of ESEI_597, an enterotoxigenic  Escherichia coli (ETEC) strain ESEI_597 was isolated in 2013 from a stool sample from an 8-month-old child residing in Korogocho, an informal settlement northeast of Nairobi, Kenya. A pea-sized fecal sample was emulsified in 5\u2009mL buffered peptone water . Approximately 10 \u03bcL of the overnight broth inoculum was plated on MacConkey agar (Oxoid), and a biochemically identified colony was subcultured in Mueller-Hinton agar (Oxoid) before being stocked at \u221280\u00b0C in tryptone soy broth (Oxoid) with 15% glycerol. All incubations were performed at 37\u00b0C for 18 to 24 h. Labile toxin (LT) and colonization surface antigen 13 (CS13) conventional uniplex PCR screening was performed using previously described primers (https://www.well.ox.ac.uk/ogc) and preprocessed using an automated protocol developed by the Modernising Medical Microbiology (MMM) Oxford Group. Read trimming to remove remnant adaptor sequences was performed using BBDuk, part of the BBTools package (www.ncbi.nlm.nih.gov/sra), with an automated step for removal of contaminant reads. The remaining reads were mapped (using Stampy v1.0.23) to an E. coli reference genome (GenBank accession AE014075.1) (http://www.bioinformatics.babraham.ac.uk/projects/fastqc), with all per-base-sequence quality flagged as passed.Enterotoxigenic 14075.1) . The quaN50 value of 140,019\u2009bp, a G+C content of 50.71%, and a total length of 4,937,264\u2009bp. ABRicate v1.0.1 (https://github.com/tseemann/abricate) was used to detect acquired antimicrobial resistance (AMR) genes with ResFinder v2.1 (https://github.com/tseemann/mlst) and the PubMLST database (https://pubmlst.org)  . Automataat operon, etpBAC operon, eatA, and tleA-like autotransporter are all absent. The cexE gene is present but truncated, with an LS-BSR value of 0.8893 (cshABCDEFGH) is 7,923\u2009bp long, and LT in ESEI_597 has been assigned to allele 29.ESEI_597 belongs to sequence type 155 (ST155) (Achtman scheme) ; plasmidf 0.8893 . The CS1The Kenya Medical Research Institute (KEMRI)/National Ethics Review Committee approved all procedures (protocol number 2507).E. coli ESEI_597 has been deposited in GenBank under the accession number JALDSW000000000. Other features of the sequence data are provided in The draft genome of"}
+{"text": "M. avium complex (MAC) strains in a chronic mouse infection model either as monotherapy or in combination with standard of care .Epetraborole (EBO) is a boron-containing oral inhibitor of bacterial leucyl-tRNA synthetase, an essential enzyme in protein synthesis; EBO demonstrates potent activity against nontuberculous mycobacteria. These studies evaluated oral doses (PO) of EBO against 5 M. avium 2285R evaluated EBO at 1, 10, 30, 100, 300 and 500 mg/kg PO once daily (QD) compared to 250 mg/kg CLR PO QD. C57BL/6 mice were infected with a pulmonary aerosol of 1x1011 CFU. Treatment was administered for 56 days starting on day 28 post-infection. The bacterial burden (CFU) in lungs was evaluated on days 1, 28 and 84 post-infection by plating serial dilutions of homogenates on Middlebrook 7H11 charcoal agar plates. An additional 4 strains of MAC were evaluated with EBO doses of 100, 200, 300 or 400 mg/kg QD compared with the SOC therapy for MAC  QD and SOC plus EBO 200mg/kg QD. Oral exposures of EBO were determined in a group of uninfected mice (Table\u00a01).A pilot chronic efficacy study against M. avium 2285R, a biofilm-forming strain, EBO at all doses tested was significantly better than CLR dosed at 250 mg/kg , and no CFU were detected on agar plates containing EBO (16 mg/L). In subsequent studies, SOC was compared to EBO in 4 additional MAC strains . Efficacy of EBO monotherapy was better than SOC against M. avium ATCC 700898, while it was as good as SOC with M. intracellulare 1956, M. intracellulareDNA00055, and M. intracellulare DNA00111 with CFU reductions ranging from 2 - 4.8 log10 compared to day 28 controls. In all four strains tested, 200 mg/kg EBO, which approximates the human oral equivalent dose of 500 mg, combined with SOC increased bacterial killing from 1.4 - 3.0 log10 CFU compared to SOC alone resulting in total lung CFU reductions of 4.6 - 5.6 log10.In a study with M. avium 2285R at day 84. EBO demonstrated potent in vivo efficacy against 5 MAC strains and significantly improved efficacy when combined with SOC, supporting further clinical development for EBO.In this chronic mouse lung infection model, no EBO resistance development was detected with Michelle S. DeStefano, n/a, AN2 Therapeutics: Grant/Research Support Carolyn Shoen, PhD, AN2 Therapeutics: Grant/Research Support Michael H. Cynamon, MD, AN2: Grant/Research Support|AN2: Grant/Research Support MRK Alley, PhD, ABBOTT LABS: Stocks/Bonds|ABBVIE: Stocks/Bonds|AN2 Therapeutics: Author on epetraborole patent|AN2 Therapeutics: Salary|AN2 Therapeutics: Ownership Interest|AVANOS MED INC: Stocks/Bonds|NABRIVA THERAPEUTICS PLC: Stocks/Bonds|NOVARTIS AG: Stocks/Bonds."}
+{"text": "Health care workers (HCWs) possess a potential risk to acquire and spread various infections. This study was planned to assess the immune status of HCWs in pediatric departments of two tertiary care hospitals in Northern India.In this cross-sectional study, HCW\u2019s (Indians), working in pediatric departments of these hospitals, over 6-months period (July18-Dec18) were enrolled after taking written consent and their 5-ml venous blood sample was collected. Ethical clearance was obtained from Institute Ethics committee, before enrolling subjects. Serum were tested for antibodies against diphtheria-toxin (DT), pertussis-toxin (PT), measles, mumps, rubella, varicella, hepatitis-B (HbsAb) and hepatitis-A using commercial IgG (quantitative) ELISA kits.A total of 160 HCW\u2019s (M:F=77:83), having mean age 30.6\u00b17.8 years, were enrolled. Out of them 106 (66.3%) were resident doctors, 31 (19.4%) nursing-staff, 18 (11.3%) faculty members, 3 (1.9%) research-staff and 2 (1.3%) paramedical-staff. In our study, antibodies (IgG) against DT were between 0.01 to 0.1 IU/ml in 78.1% (120/160); requiring a Tdap booster; while 7 out of 125 had titers < 0.01 IU/ml, which needed 3-doses of primary vaccination. Antibodies (IgG) against PT < 30 IU/ml were seen in 60.6% (97/160), which were considered as seronegative, as per kit recommendations.A total of 3% (5/160), 13.1% (21/160), 10% (16/160), 17.5% (28/160) had titers < 12 IU/ml for IgG antibodies against measles, mumps, rubella and varicella respectively; were considered unprotected. A total of 25% (40/160) had Anti-HBs antibody titers < 20 mIU/ml; which were low, therefore were advised to take one booster dose of Hep-B vaccine. A total of 15.6% (25/160) had IgG antibodies against hepatitis-A < 10 mIU/ml; were unprotected.Alarming proportions of pediatric-HCWs had low antibody titres against DT (78.1%) and PT (60.6%), necessitating a dose of Tdap. A total of 10%, 17.5% and 15.6% lacked protective antibodies against rubella, varicella and hepatitis-A. A quarter of screened population had low anti-Hbs titres, requiring boosting of immunity. Our study emphasizes the urgent need for improving immunization status of pediatric HCW\u2019s; as they continue to remain susceptible to various vaccine preventable infectious diseases.All Authors: No reported disclosures."}
+{"text": "Correction: BMC Cancer 22, 689 (2022)https://doi.org/10.1186/s12885-022-09740-9The results of colony formation assay which have been used in Fig. 4C(c) and Fig. 4D(c) were accidentally used again in Fig. The western blot result annotations \"Ctrl \"and \"MIR137HG\" were missed in Fig. Following publication of the original article , the autThe correct Fig."}
+{"text": "Scientific Reports, 10.1038/s41598-023-28540-0, published online 28 January 2023Correction to: The original version of this Article contained an error in Reference 37,et al.\u00a0Inclusive Wealth Report 2022 Executive Summary. .37. Barbier, E. B.\u00a0now reads:. UNEP. Inclusive Wealth Report 2022 Executive Summary. .37The original Article has been corrected."}
+{"text": "HFE p.C282Y/p.C282Y are incompletely characterized.Screening program participants with iron overload (IO) phenotypes without HFE p.C282Y and p.H63D genotyping.We studied white participants who had IO phenotypes without p.C282Y/p.C282Y in post-screening clinical examinations (CE). We defined IO phenotypes as a) elevated serum ferritin (SF) and transferrin saturation (TS) at screening and CE, and b) absence of IO treatment, anemia, transfusion >10 units, alcohol intake >30 g/d, hepatitis B or C, and pregnancy. We defined IO-related disease as elevated alanine or aspartate aminotransferase (ALT/AST) or swelling/tenderness of 2nd/3rd metacarpophalangeal (MCP) joints. All participants had HFE genotypes were 12.9 (p.C282Y/p.H63D), 3.0 (p.H63D/p.H63D), 1.9 (p.C282Y/wt), 0.9 (p.H63D/wt), and 0.5 (wt/wt) compared to 42,640 white screening participants without IO phenotypes or p.C282Y/p.C282Y. Regression on SF revealed positive associations: MCV ; swelling/tenderness of MCP joints ; and p.H63D/wt . IO-related disease  occurred in 19 participants . Median MCV was higher in participants with IO-related disease . Logistic regression on IO-related disease revealed a significant association with diabetes ).There were 32 men and 26 women (mean age 54\u00b116 y). Median food/supplemental iron intakes were 14.3/0.0 mg/d. Relative risks of HFE p.C282Y/p.C282Y, relative risks of HFE genotypes p.C282Y/p.H63D, p.H63D/p.H63D, and p.C282Y/wt were significantly higher than in 42,640 white screening participants with neither IO phenotypes nor p.C282Y/p.C282Y. SF was significantly associated with MCV, swelling/tenderness of 2nd/3rd MCP joints, and p.H63D/wt. IO-related disease was significantly associated with MCV and diabetes.In the present 58 screening program participants who had IO phenotypes without  HFE gene  [HFE alleles p.C282Y  and p.H63D  were discovered in referred white patients with hemochromatosis and iron overload (IO) in 1996 [The HFE alleles [HJV, chromosome 1q21.1), transferrin receptor 2 , hepcidin antimicrobial peptide , and solute carrier family 40 member 1 (ferroportin)  [Rare types of hemochromatosis, usually diagnosed in referred patients, are associated with novel  alleles  or deletHFE genotype and sex [Some population screening program participants who have elevated SF and elevated TS without p.C282Y/p.C282Y also have IO phenotypes due to hemochromatosis, although these participants have been incompletely characterized. A primary report of the Hemochromatosis and Iron Overload Screening (HEIRS) Study displayed separate initial screening TS and SF data of participants from a racially diverse population segregated only for  and sex . Those d and sex .HFE p.C282Y/p.C282Y who participated in the HEIRS Study post-screening clinical examinations (CE) [HFE genotypes other than p.C282Y/p.C282Y and identified variables significantly associated with IO phenotypes and IO-related disease. We discuss our observations in the context of previous reports.To learn more, we studied non-Hispanic whites who had IO phenotypes without ons (CE) , 8, 9. Gons (CE) , 8, 9. Wons (CE) . We compThe National Heart, Lung, and Blood Institute/National Human Genome Research Institute HEIRS Study evaluated the prevalence, genetic, and environmental determinants, and potential clinical, personal, and societal impacts of hemochromatosis and IO in a multiethnic, primary care-based sample of 101,168 adults enrolled during the interval 2001\u20132003 at four Field Centers in the US and one in Canada . The StuHFE p.C282Y and p.H63D allele-specific genotyping [The HEIRS Study recruited participants from a health maintenance organization, diagnostic blood collection centers, and public and private primary care offices in ambulatory clinics associated with five Field Centers . Ninety-notyping .HFE p.C282Y homozygotes (regardless of screening SF and TS), to all participants whose screening SF and TS exceeded study thresholds, regardless of HFE genotype, and to selected participants who had normal screening SF and TS and HFE wt/wt, defined as absence of p.C282Y and p.H63D. Median interval between screening and CE participation was 8 months [Invitations to participate in a post-screening CE were extended to all 8 months .HFE genotype. Each participant completed a questionnaire addressing medical history and medications [At CE, eligible participants were informed of their screening SF, TS, and ications  and a Unications . An HEIRications .HFE genotype based on p.C282Y and p.H63D allele-specific analyses [All testing was performed at the HEIRS Study Central Laboratory . At CE, a morning blood sample was obtained after an overnight fast to measure SF and TS , to confirm Elevated SF and TS were defined as SF >300 \u03bcg/L and TS >50% (men) and SF > 200 \u03bcg/L and TS >45% (women). Reference ranges for hemoglobin were 133\u2013177 g/L (men) and 117\u2013157 g/L (women) and for mean corpuscular volume (MCV) were 78\u2013105 fL (men) and 78\u2013106 fL (women) [Reflex testing for hepatitis B surface antigen and hepatitis C antibody was performed in all participants with elevated ALT or AST . Participants with elevated ALT or AST and positivity for hepatitis B surface antigen were defined to have viral hepatitis B . ParticiObtaining liver specimens by biopsy, estimating liver iron content using imaging techniques, and performing therapeutic phlebotomy were beyond the scope of the HEIRS Study.We defined IO phenotypes as the following: a) elevated SF and elevated TS at both screening and CE, and b) absence of IO treatment, anemia, erythrocyte transfusion >10 units, alcohol intake >30 g/d, hepatitis B or C, and pregnancy. We defined IO-related disease in participants with IO phenotypes according to modified criteria of Allen et al. : elevateHFE p.C282Y/p.C282Y who had IO phenotypes, who fasted overnight \u22658 h before CE, and whose data analyzed in this study were complete.We analyzed observations on all self-identified non-Hispanic white CE participants without Macrocytosis was defined as MCV greater than the respective sex-specific upper reference limits. Microcytosis was defined as MCV \u226477 fL.2.Estimates of dietary and supplemental iron and daily alcohol intake were taken from the Multi-Ethnic Dietary Questionnaire . We defiSelf-reported diabetes reported at screening was confirmed with medication reviews at CE . We defiPhysicians who performed physical examinations at CE recorded these manifestations as present or absent .The dataset for analyses consisted of complete observations on 58 CE participants with IO phenotypes. Age, SF, TS, hemoglobin, and MCV are expressed to the nearest integer. Descriptive data are displayed as enumerations, percentages, proportions, means (\u00b1 1 SD), or medians (range). Analyses of continuous data using d\u2019Agostino\u2019s and Shapiro-Wilk tests revealed that age and BMI values were normally distributed and thus these values are displayed as mean \u00b1 1 SD and were compared using Student\u2019s t tests (two-tailed). Continuous variables that were not normally distributed are displayed as medians (range) and were compared using Mann-Whitney U tests (unpaired data) or Wilcoxon\u2019s signed-ranks test (paired data). Percentages were compared using Fisher\u2019s exact test (two-tailed). The significance of the difference between two independent proportions was calculated using the z-ratio and two-tailed values of p. We compared screening and CE SF and TS data using Pearson\u2019s correlation coefficient.HFE genotypes other than p.C282Y/p.C282Y. We performed forward stepwise regression on SF at CE using a) all independent variables available at CE (except TS) and b) the first independent variable with p \u22650.05 as the stopping rule. TS was not used as an independent variable because its positive correlation with SF at CE was significant. We display standardized coefficients \u03b2 for each significant predictor variable in this regression.We computed relative risks of We performed logistic regression on IO-related disease using all available independent variables  for which values of p in univariate comparisons were \u22640.1500. We calculated odds ratios ) for significant independent variables.\u00ae , GB-Stat\u00ae , and GraphPad Prism 8\u00ae .We defined values of p <0.05 to be significant. Analyses were performed with Excel 2000HFE genotypes [HFE p.C282Y/p.C282Y [Screening observations in 43,453 non-Hispanic white participants  were evaluable for SF and TS levels and enotypes . Both sc/p.C282Y . ElevateHFE genotypes p.C282Y/p.H63D, p.H63D/p.H63D, and p.C282Y/wt in the present 58 CE participants with IO phenotypes were significantly greater than corresponding percentages in 42,640 non-Hispanic white screening participants without IO phenotypes (Percentages of enotypes . Relativenotypes . Aggregaenotypes .Detailed characteristics of 58 participants are displayed in HFE p.C282Y/p.H63D occurred in 18 participants (31.0%) and p.H63D/p.H63D in four participants (6.9%). Each of the genotypes p.C282Y/wt, p.H63D/wt, and wt/wt occurred in 12 participants. Two men and two women had SF >1000 \u03bcg/L . Macrocytosis was detected in one woman and microcytosis was detected in another woman. Median estimated daily iron intakes from food and iron supplements were 14.3 mg and 0.0 mg, respectively. Elevated ALT or AST occurred in 18 participants (31.0%). Swelling/tenderness of the 2nd/3rd MCP joints was observed in one participant (1.7%). Diabetes occurred in four participants (6.9%). 000 \u03bcg/L .HFE p.H63D/wt . This regression accounted for 32.0% of the variance of SF (regression ANOVA p = 0.0001).Forward stepwise regression on SF at CE using other CE observations (except TS) as independent variables revealed three positive associations: MCV ; swelling/tenderness of 2nd/3rd MCP joints ; and These data are displayed in There were 15 abstainers (25.9%) and 43 non-abstainers (74.1%). The difference between median MCV values of abstainers and non-abstainers was not significant (94 fL (86\u201397) vs. 96 fL (77\u2013107), respectively; p = 0.1369). All nine of 58 participants (15.5%) with MCV >100 fL were non-abstainers, although proportions of abstainers and non-abstainers with MCV >100 fL did not differ significantly .HFE p.C282Y/p.C282Y who also reported that they had not received treatment for IO. Our study design excluded non-Hispanic white CE participants who may have had elevated SF and elevated TS due to types of anemia that increase iron absorption, a history of erythrocyte transfusion >10 units, viral hepatitis B or C, or increased alcohol intake. Elevated SF without elevated TS is common in patients with obesity [SLC40A1 alleles [We evaluated observations in 58 non-Hispanic white participants in the HEIRS Study CE who had both elevated SF and elevated TS defined as IO phenotypes without  obesity , metabol obesity , diabete obesity , non-alc obesity , 19, inf obesity , maligna obesity \u201322, \"cla alleles , heredit alleles , and ben alleles . Thus, oHFE genotypes p.C282Y/p.H63D, p.H63D/p.H63D, and p.C282Y/wt in the present 58 CE participants with IO phenotypes were significantly greater than corresponding percentages in 42,640 non-Hispanic white screening participants without IO phenotypes. SF was significantly associated with MCV, swelling/tenderness of the 2nd/3rd MCP joints, and p.H63D/wt. IO-related disease was significantly associated with MCV and diabetes. IO and IO-related disease were not significantly associated with estimated intake of dietary or supplemental iron. Taken together, we infer that heritable factors in addition to common HFE genotypes other than p.C282Y/p.C282Y or acquired factors we did not study contribute to or account for IO phenotypes, IO-related disease, and other potential iron-related consequences of hemochromatosis in the present participants.Percentages of HFE p.C282Y/p.H63D was increased in the present participants, although we observed no significant association of IO-related disease with p.C282Y/p.H63D. Participants in an Australian population screening program with IO phenotypes and p.C282Y/p.H63D also had a low risk of IO-related disease  [Relative risk of 5 women) . In refe5 women) . In an a5 women) . In a me5 women) .GNPAT) p.D519G (rs11558492) occurred with greater prevalence in French and Australian subjects with HFE p.C282Y/p.H63D with higher SF than those with lower SF [CYBRD1) promoter polymorphism rs884409 was significantly higher in French patients with p.C282Y/p.H63D with higher SF than those with lower SF [HJV p.N196K  or HAMP -72C>T [HAMP p.G71D  did not modify SF and TS phenotypes [HFE iron-associated alleles probably account in part for the variable penetrance of p.C282Y/p.H63D phenotypes.Glyceronephosphate O-acyltransferase  [HFE coding region allele [HAMP Met50del IVS2+1(-G) and HAMP p.G71D, respectively [Relative risk of .9, 5.8) . In a me.9, 5.8) . Compounn allele , occurren allele , 36. Refn allele , 38. In ectively .HFE p.H63D/wt was increased in the present participants. In an analysis of 14 case-control studies of hemochromatosis, the pooled odds ratio of p.H63D/wt as a contributor to hemochromatosis IO was 1.9  [HFE alleles, e.g., p.I105T (c.314T>C) [HFE allele, e.g., p.E168Q (c.502G>C), in cis with p.H63D [Relative risk of .5, 2.5) . Rarely,.314T>C) , or withh p.H63D . In a meh p.H63D . It is pHFE p.C282Y/p.C282Y [Estimated median daily intake of dietary iron in the present participants is similar to that in adults in the U.S. not selected for IO phenotypes . SF in t/p.C282Y .HFE p.C282Y/p.C282Y were significantly higher than those of control subjects [HFE genotype [MCV was significantly associated with SF and IO-related disease in this study, after adjustment for other variables. In two previous reports, mean MCV values in referred hemochromatosis patients without subjects , 45. Incgenotype . In one genotype . By desigenotype ) and thagenotype ). We obsgenotype  and manygenotype , occurreHFE genotypes were p.H63D (n = 2) and p.H63D/p.H63D and wt/wt (one each). In 20,306 Canadian HEIRS Study participants linked to the Ontario Death Registry 9 years after screening, all-cause survival was significantly decreased in the 34 participants with screening SF >1000 \u03bcg/L and HFE genotypes other than p.C282Y/p.C282Y  [Four of the present 58 participants  had SF >1000 \u03bcg/L. Their 7 wt/wt) .HFE genotypes other than p.C282Y/p.C282Y in adults with IO phenotypes have meaningful clinical correlates. Some adults with IO phenotypes without p.C282Y/p.C282Y have hepatic steatosis [HFE hemochromatosis, or non-HFE \"modifier\" mutations. Guidelines for hemochromatosis management promulgated by expert groups in the U.S. [HFE genotyping. It is also recommended that first-degree relatives of persons diagnosed to have HFE hemochromatosis undergo TS/SF phenotyping and HFE genotyping [HFE alleles has disease-identification value other than to reveal IO risk.teatosis , 51 or cteatosis , 52, othteatosis . Adults the U.S.  and Eurothe U.S.  recommennotyping , 54. TheHFE p.C282Y/p.C282Y and unlikely to have other disorders that sometimes present with IO phenotype mimics. Another strength is the availability of screening and CE data sufficient to demonstrate a) significant differences in proportions and relative risks of HFE genotypes other than p.C282Y/p.C282Y and b) significant associations of IO phenotypes and IO-related disease with other potential iron-related consequences of hemochromatosis. An uncertainty of this study is that we may have excluded participants with \"classic\" SLC40A1 (ferroportin) hemochromatosis in whom SF but not TS is typically elevated [A strength of this study is that our selection criteria included only participants with IO phenotypes who were likely to have hemochromatosis in the absence of elevated .HFE genotype risk associated with IO phenotype because participants invited to the CE (other than those with HFE p.C282Y/p.C282Y or wt/wt) were selected on the basis of having both screening SF and screening TS values above HEIRS Study thresholds, not on the basis of HFE genotype [HFE p.C282Y and p.H63D. The HEIRS Study performed analyses to detect specific HFE alleles other than p.C282Y and p.H63D and specific HJV, TFR2, HAMP, SLC40A1 alleles in a small proportion of screening participants [In this study, we assessed genotype . The HEIicipants . None oficipants .HFE genotypes p.C282Y/p.H63D, p.H63D/p.H63D, and p.C282Y/wt were increased in screening program participants who had IO phenotypes without p.C282Y/p.C282Y. SF was significantly associated with MCV, swelling/tenderness of 2nd/3rd MCP joints, and p.H63D/wt. IO-related disease was significantly associated with MCV and diabetes. Novel HFE alleles, deleterious alleles of HJV, TFR2, HAMP, or SLC40A1, iron-related \"modifier\" alleles, or acquired factors could contribute to IO phenotypes and IO-related disease in some of the present participants.We conclude that proportions and relative risks of S1 Table(PDF)Click here for additional data file.S2 Table(PDF)Click here for additional data file.S3 Table(PDF)Click here for additional data file."}
+{"text": "Although convenient single-tablet antiretroviral regimens have been developed to treat HIV in recent years, some patients have continued to take a multi-tablet treatment, nevirapine extended-release (NVP XR) plus two nucleoside reverse transcriptase inhibitors (NRTIs) due to its excellent safety profile. This observational study examined the demographic and clinical characteristics of HIV-positive patients who switched to dolutegravir plus lamivudine (DTG/3TC) from NVP XR plus 2 NRTIs, following discontinuation of NVP XR from the Canadian market.Virally suppressed (< 50 cps/mL) HIV-positive adults \u226518 years who switched from NVP XR plus 2 NRTIs to DTG/3TC between 20 August 2019 and 30 April 2020 were retrospectively identified from Electronic Medical Records housed at Spectrum Health in British Columbia, Canada. Baseline demographic and clinical characteristics were summarized using descriptive statistics at the date of first DTC/3TC prescription. Virologic control, CD4 cell count, weight-related changes and exploratory characteristics related to metabolic syndrome were summarized at baseline and at 12 \u00b1 6-months post-switch using descriptive statistics. Reasons for treatment discontinuation were also captured.Sixty-nine patients were identified . Mean length of use (\u00b1SD) of NVP XR was 4.6 \u00b1 2.6 years. Sixty-three (91.3%) persisted on DTG/3TC at the 12-month timepoint post-switch with 61 (96.8%) virally suppressed < 50 cps/mL and all 63 (100%) virally suppressed < 200 cps/mL. Among persistent patients with CD4 cell counts available, mean CD4 cell count (\u00b1SD) remained stable, increasing slightly from 724.4 \u00b1 238.4 to 740.3 \u00b1 240.6 cells/uL. All six (8.7%) patients who discontinued DTG/3TC, discontinued due to tolerability and not effectiveness reasons.Our findings are the first to examine real-world use of single-tablet, 2-drug DTC/3TC among patients who switched from multi-tablet, 3-drug NVP XR plus 2 NRTIs in Canada. The majority persisted on DTG/3TC and remained virally suppressed at the < 50 cps/mL level 12-months post-switch. This, coupled with excellent tolerability, demonstrates the effectiveness of DTG/3TC in maintaining viral suppression among a unique and stable group of older men.Joss de Wet, MBChB CCFP FCFP, Gilead: Advisor/Consultant|Gilead: Board Member|Gilead: Grant/Research Support|ViiV: Advisor/Consultant|ViiV: Board Member|ViiV: Grant/Research Support Joann K. Ban, PharmD, MSc, GlaxoSmithKline Canada Inc.: Employee Gustavo Verdier, BSc, BPharm, MBA, GlaxoSmithKline: Stocks/Bonds|ViiV Healthcare ULC: Salary Juejing Ling, MSc, GSK: Employee of IQVIA , a company that receives consulting fees from GSK. Andrean Bunko, MPH, GlaxoSmithKline Inc.: GSK contracted with IQVIA for this and other projects. I am acting as an author in capacity of my employment with IQVIA. Michael McKimm, BSc, ViiV Healthcare: employee."}
+{"text": "Stenotrophomonas maltophilia with a 44,659-bp genome. This phage is closely related to Stenotrophomonas phage SM171, sharing 92% overall nucleotide identity as determined by BLASTn, and it shares 14 similar proteins  with some Pseudomonas phages from the genus Beetrevirus.Phage Suso is a temperate siphophage of  Stenotrophomonas maltophilia is emerging as a multidrug-resistant respiratory pathogen  and trimmed with FASTX-Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/). The trimmed reads  were used for assembly using SPAdes v3.5.0 . Annotation of the phage genome was performed using the Center for Phage Technology (CPT) Galaxy-Apollo platform  , in Wadsworth, TX, in September 2019, using reads 87,54 reads xy-pub) 79. Gene cxy-pub) 7 and Metaxy-pub) 7. tRNA gexy-pub) 7 and tRNAxy-pub) 7. Gene fuxy-pub) 7, TMHMM vxy-pub) 7, HHPred xy-pub) 7, and Sigxy-pub) 7. Genome-xy-pub) 7. All sofStenotrophomonas phage SM171 (MZ611865), sharing 92% overall nucleotide identity as determined by BLASTn. Besides this phage, Suso shares 14 similar proteins  with some Pseudomonas phages from the genus Beetrevirus (taxid 2560098), such as phages B3 (GenBank accession number NC_006548), JBD67 (GenBank accession number NC_042135), and vB_Pae_BR141c (GenBank accession number MK511065).Phage Suso has a siphophage morphology  possessiMZ326866. The associated BioProject, SRA, and BioSample accession numbers are PRJNA222858, SRR14095259, and SAMN18509666, respectively.The Suso sequence was deposited in GenBank with accession number"}
+{"text": "Mindfulness (being present in the moment without judgement) has been linked to greater caregiver emotional health. Recent mindfulness-based interventions report improved coping skills, mood, and reduced stress in dementia caregivers. In this cross-sectional study of 141 ADRD caregivers, we assessed whether the relationship between caregiver mindfulness and caregiver experience varies by caregiver gender, relationship to patient (spouse-vs-child), etiology (AD-vs-LBD), or stage (MCI-vs-dementia). A stratified univariate analytic approach was used. Four mindfulness parameters  were used: global score (GS), decentering (F1), positive (F2), and negative emotional regulation (F3). Outcomes included positive and negative appraisals of caregiving (PANAC), preparedness, care confidence, and depression. GS was linked to positive outcomes in male (rPANAC+=0.32/p=0.005), spouse caregivers (rPANAC+)=0.32/p=0.006 ) of ADRD patients regardless of etiology  and stage . Inverse relationships were observed with negative outcomes in male (rPANAC-=-0.46/p=0.002 and rdepression=-0.41/p=0.005), spouse caregivers (rPANAC-=-0.25/p=0.035 and rdepression=-0.30/p=0.009) of AD patients (rPANAC-=-0.25/p=0.043 and rdepression=-0.33/p=0.009) in early stages (rdepression=-0.41/p=0.001). F2 contributed to most relationships, with F3 and F1 significant in some but not all caregiver groups. Specifically, male spouse caregivers of AD patients regardless of stage may benefit from full-scope (F1-F3) programs while those of LBD patients from programs focused on improving emotional regulation (F2-F3). Wives of AD and LBD patients may in turn benefit from programs to improve positive emotional regulation (F2). Findings suggest that tailoring mindfulness-based interventions to specific caregiver groups may be effective in improving caregiver experience and mood."}
+{"text": "Rationale: Infection with the SARS-CoV2 virus is associated with elevated neutrophil counts. Evidence of neutrophil dysfunction in COVID-19 is based on transcriptomics or single functional assays. Cell functions are interwoven pathways, and understanding the effect across the spectrum of neutrophil function may identify therapeutic targets. Objectives: Examine neutrophil phenotype and function in 41 hospitalised, non-ICU COVID-19 patients versus 23 age-matched controls (AMC) and 26 community acquired pneumonia patients (CAP). Methods: Isolated neutrophils underwent ex vivo analyses for migration, bacterial phagocytosis, ROS generation, NETosis and receptor expression. Circulating DNAse 1 activity, levels of cfDNA, MPO, VEGF, IL-6 and sTNFRI were measured and correlated to clinical outcome. Serial sampling on day three to five post hospitalization were also measured. The effect of ex vivo PI3K inhibition was measured in a further cohort of 18 COVID-19 patients. Results: Compared to AMC and CAP, COVID-19 neutrophils demonstrated elevated transmigration (p = 0.0397) and NETosis (p = 0.0332), and impaired phagocytosis (p = 0.0036) associated with impaired ROS generation (p < 0.0001). The percentage of CD54+ neutrophils (p < 0.001) was significantly increased, while surface expression of CD11b (p = 0.0014) and PD-L1 (p = 0.006) were significantly decreased in COVID-19. COVID-19 and CAP patients showed increased systemic markers of NETosis including increased cfDNA (p = 0.0396) and impaired DNAse activity (p < 0.0001). The ex vivo inhibition of PI3K \u03b3 and \u03b4 reduced NET release by COVID-19 neutrophils (p = 0.0129). Conclusions: COVID-19 is associated with neutrophil dysfunction across all main effector functions, with altered phenotype, elevated migration and NETosis, and impaired antimicrobial responses. These changes highlight that targeting neutrophil function may help modulate COVID-19 severity. Coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) virus, was declared a global pandemic by The World Health Organization (WHO) on 11 March 2020 [Dysregulated virus induced host-immune responses are thought to be the primary cause of severe COVID-19 . NeutropAdvanced age is a recognised risk factor for severe COVID-19, including the development of ARDS ,7,8. AgeEmerging studies suggest that neutrophils are implicated in the pathogenesis of severe COVID-19 and every reporting study has described cell dysfunction (or inferred cell dysfunction through transcriptomics), which could contribute to tissue damage and secondary infection ,25,26,27These are important considerations. As neutrophil functions change with age, age matched controls are important in identifying pathological differences. Although approximately 12% of hospitalized COVID-19 patients require ICU support, the majority of hospital admissions and deaths occur on non-ICU wards ; understWe hypothesised that neutrophils from COVID-19 patients would exhibit diverse altered effector functions and changes to phenotype, with the degree of dysfunction associated with adverse patient outcomes.This study aimed to perform, for the first time, a comprehensive assessment of ex vivo neutrophil phenotypes and functions in a statistically powered cohort of hospitalized, non-ICU SARS-CoV2 infected patients compared to aged-matched controls and patients with non-COVID community acquired pneumoniae (CAP) and to investigate relationships with clinical outcomes.Recruitment is summarized in 2: FiO2 ratio (SF), converted to PaO2: FiO2 (PF) (SF = 57-0.61PF).COVID-19 patients were recruited within 48 h of hospital admission due to pneumonitis/pneumonia related to COVID-19. All patients had a positive COVID-19 PCR swab. No patients received novel treatments or were part of a COVID clinical medicinal trial on recruitment to this study. Exclusion criteria is listed in CAP patients were recruited within 48 h of hospital admission due to non-COVID-19 pneumonia. All patients had a negative COVID-19 swab. Exclusion criteria is listed in the Age matched controls (AMC) were either recruited from patients attending pre-booked face-to-face outpatient appointments or from hospital staff. AMC had no evidence of acute illness, including COVID-19, within the last two weeks, as assessed by a respiratory physician, and met the other exclusion criteria.S. pneumoniae and NETosis by the release of cell-free (cf) DNA  DNA . Plasma t-test or ANOVA. A Mann-Whitney U test for unpaired data, a Wilcoxon test for paired data, or a Kruskall-Wallis test was used to analyse non-normally distributed data. Data are presented throughout as median (IQR), with each n number representing a separate study participant. Significance was defined at p < 0.05. There were no corrections for multiple comparisons, but exact p values are given. A power calculation performed on isolated neutrophil NETosis data  suggested that 18 participants were required in each group . A Kolmogorov-Smirnov Test was used to determine data distribution. Normally distributed data were analysed using a student\u2019s 41 COVID-19 patients (mean age 71.5 years), two healthy AMC (mean age 70 years) and 26 CAP patients (mean age 67.5 years) were included in the study. Demographics are provided in p = 0.0332) and CAP  AMC vs. 40.63 (115.2) COVID-19, S. pneumoniae was significantly decreased in COVID-19 patients compared to AMC (Median fluorescence intensity (MFI): 8.0 (4.2) AMC vs. 6.6 (2.6) COVID-19, p = 0.0366) and CAP (MFI 9.8 (4.8), p = 0.0052, p = 0.3257, p = 0.9228, p = 0.6726, Neutrophil phagocytosis of p = 0.0091), and CAP patients (MFI: 285 (211) baseline vs. 357 (260) 30 min, p = 0.038), but not in COVID-19 patients (MFI: 44.1 (35) baseline vs. 65.3 (60) 30 min, p = 0.134, p < 0.0001, Cytoplasmic (c)ROS, and nuclear/mitochondrial (n/m)ROS were measured in resting neutrophils and following phagocytosis. Compared to unstimulated cells, cROS levels were elevated after phagocytosis in both AMC (MFI: 46.8 (28) baseline vs. 64.9 (51) 30 min, p < 0.0001), but not in COVID-19 patients (MFI: 18.9 (12) baseline vs. 21.2 (12), p = 0.0329) or CAP patients (MFI: 22.2 (21) baseline vs. 26.7 (17), p = 0.989, p < 0.0001) and CAP neutrophils  baseline vs. 32.0 (38), p = 0.0394, p = 0.0118, A larger increase in the level of cfDNA was detected in supernatants obtained from COVID-19 patient neutrophils post-PMA treatment compared to AMC (fold change in absorbance of neutrophils stimulated with PMA vs. vehicle control: 1.29 (0.32) AMC vs. 1.53 (1.66) COVID-19, p = 0.0396, p = 0.0186, p = 0.0322).At hospital admission, COVID-19 patients presented with higher concentrations of plasma cfDNA compared to AMC (621 ng/mL (1324) AMC vs. 1071 ng/mL (856) COVID-19, To determine whether neutrophils, via NETosis, were a source of the cfDNA, plasma samples were screened for the presence of CitH3, a protein that decorates the DNA backbone of NETs . Westernp < 0.0001, vs. 77.45% (34) CAP, p < 0.0001, Using NETs as a substrate, serum DNase activity was lower in COVID-19 patients at the time of hospital admission when compared to both AMCs and CAP patients (% degradation: 88.4% (93) AMC vs. 12.8% (49) COVID-19, To determine whether changes observed in neutrophil function were associated with phenotype, expression of key surface molecules were investigated by flow cytometry. This was compared to both AMC and CAP patients. A table of percentage receptor expression and MFI is shown in p = 0.0014, p = 0.0026, p = 0.046; MFI:1701 (1268) CAP, p < 0.0001). Compared to AMC, both the percentage of neutrophils expressing the activation marker CD11b (82% (15) AMC vs. 68% (44) COVID-19, p < 0.0001, vs. 7% (11) CAP, p < 0.0001, The percentage of neutrophils expressing CD54, a marker of reverse transmigration, was elevated in COVID-19 patients compared to both AMC and CAP patients (26% (37) AMC vs. 71% (21) COVID-19, p < 0.006, vs. 524 (264) and CAP, p < 0.0001, The percentage of neutrophils expressing CD66b, CD62L, CD10, CXCR2, CXCR4, CD11c and PD-L1 did not differ between AMC and COVID-19 patients see . The surThere was no association of neutrophil phenotypic marker expression with 4C severity score, or in survivors and non-survivors.p = 0.0322, p = 0.0273, p = 0.0420, On day three to five follow-up, relative to baseline readings, there was a decrease in CXCR2 expression: (MFI: 1960 (1601) day one vs. 1126 (1262), day three to five, p = 0.0322, p = 0.0273, p = 0.0420, On day three to five follow-up, relative to baseline readings, there was a decrease in CXCR2 expression: (MFI: 1960 (1601) day 1 vs. 1126 (1262), day three to five, p = 0.0296, CAP p = 0.0106, p < 0.0001, CAP p = 0.0032, p < 0.0001, CAP p < 0.0001, p < 0.0001, Compared to AMC, both COVID-19 and CAP patients showed elevated levels of IL-6 , p = 0.0156, Pre-incubation of COVID-19 neutrophils from cohort 2, with Pi3k delta inhibitor CAL101 and gamma inhibitor AS252434 significantly decreased PMA induced NETosis (PMA RFU 247 (228.5) vs. delta 207.5 (242.5), We present novel findings of COVID-associated neutrophil dysfunction across all main effector functions. In summary, compared to AMC and patients with CAP, systemic neutrophils from patients hospitalized with moderate severity COVID-19 demonstrated increased migration, impaired anti-microbial responses including reduced phagocytosis and nuclear/mitochondrial ROS generation after phagocytosis. Later/end phase neutrophil responses were increased, namely ex vivo NETosis with evidence of increased systemic NETosis, coupled with reduced DNase activity, which was also elevated in CAP. We also show an altered but distinct neutrophil phenotype, not compatible with a purely activated, immature, senescent or anti-inflammatory phenotype as described before (results summarised in Individually, as described in other studies, these changes to effector function could compromise aspects of the host defence. Collectively, these changes represent a clear mechanism for significant tissue damage. Poor phagocytosis would impede pathogen clearance, increasing the likelihood of secondary infection and amplifying inflammation. NETosis is implicated in tissue damage and thrombotic events in several disease settings ,38. The S. pneumoniae, the most common bacteria implicated in secondary infection in COVID-19 [Secondary infection in COVID-19 is associated with increased severity of lung disease and poorer outcomes ,41. ImpaCOVID-19 , alongsiCOVID-19 , may conElevated NETosis ,24,44 anThe collective pattern of neutrophil dysfunction in COVID-19 speaks of alterations to mechanosensing within these cells. Elevated migration and impaired phagocytosis could both be linked to reduced pseudopod extrusion, which is known to increase migratory speed . FurtherWe observed an altered neutrophil phenotype in moderate COVID-19, not compatible with previously described populations, and not in keeping with our AMC or CAP control cohorts. COVID-19 neutrophils expressed decreased levels of the activation marker CD11b and a lack of CD62L shedding, which has previously been observed in sepsis , alongsi+ CXCR2- neutrophils, confirming a report of reduced CXCR2+ neutrophils in ICU COVID-19 patients [Finally, we observed that COVID-19 neutrophils expressed elevated CD54, a marker of reverse transmigration, whereby neutrophils migrate from the tissues back into the circulation. These cells are capable of high levels of oxidative burst , which mpatients .The majority of COVID-19 hospitalisations and deaths occur in non-ICU wards , making Our data suggests a distinct cellular response in moderate COVID-19 which contributes to on-going immune mediated harm, but which may be modifiable using a targeted therapy, such as PI3K\u03b4 or PI3K \u03b3 inhibitors administered at this crucial point in disease progression.\u00ae to isolate neutrophils, and the authors combined data from multiple blood samples taken over eleven days of hospitalization. As we show changes in neutrophil phenotype and function over the three to five time course of our study, we suggest that combining time points obscures the complex changes occurring in this short-lived cell population. We also used opsonized S. pneumoniae for phagocytosis studies which may be phagocytosed by different mechanisms to S. aureus bioparticles [Our data complements studies which showed increased systemic NETosis in COVID-19 ,68, and articles , confounMore recently, Loyer et al., demonstrated the increased expression of CD11b and ROS production with decreased CD62L expression in a cohort of COVID-19 patients treated in the ICU in comparison with CAP and healthy controls . The autThis study was limited due to the safety measures required when handling biological fluids for a new infectious disease. All experiments were carried out within a BSL2 hood and methods were chosen based on tolerance to inactivation/fixation with 4% PFA. Our patients did not include an ICU group; however, mild-moderate disease affects a larger proportion of overall COVID-19 patients, and we believe it is this point in the patient pathway which holds most potential for successful intervention. Our AMC group highlights changes in COVID-19 and our CAP group highlights differences between disease types.Our study shows that moderate COVID-19 is associated with alterations in neutrophil phenotype, increased migratory capacity and NETosis, and impaired antimicrobial function, which contributes to the severity of COVID-19. Elevated NETosis in the lung is associated with disease severity, and elevated systemic NET production is likely to contribute to inflammation, which may drive ARDS associated damage and thrombosis. Targeting neutrophils and their downstream effectors may be beneficial in the treatment of COVID-19."}
+{"text": "Mycobacterium smegmatis mc2155. McGee has a genome 156,008\u2009bp long, containing 237 protein-coding genes, 31 tRNA genes, and 1 transfer-messenger RNA (tmRNA) gene. McGee shares high gene content similarity to phages in actinobacteriophage cluster C1.Mycobacteriophage McGee is a myovirus isolated from a wet soil sample collected at Manassas, VA, using  Mycobacterium smegmatis mc2 155 have been isolated and characterized, providing important insights into viral diversity and population structure . The soil sample was washed with 7H9 liquid medium, the wash filtered , and the filtrate plated in soft agar overlay containing Mycobacterium smegmatis mc2 155. McGee was purified with multiple rounds of plating. Negative-stain transmission electron microscopy revealed McGee to possess a Myoviridae morphology , and sequenced on an Illumina MiSeq instrument v3 reagents), resulting in 77,248 single-end 150-bp reads providing 73\u00d7 coverage. The raw reads were assembled and assessed using the programs Newbler v2.9 ( reagentshttp://cobamide2.bio.pitt.edu/) (http://phages.wustl.edu/starterator), whereas gene functional assignments were determined using BLASTp searches against the NCBI nonredundant v2.13.0 (13Genes were identified and annotated using DNA Master v5.23.6 (tt.edu/) , GLIMMER v2.13.0  and acti v2.13.0 , HHpred  v2.13.0 , and Pha v2.13.0 . Aragorn v2.13.0  and tRNA v2.13.0  were use2.13.0 1315 and TM2.13.0 13, 17. AllA total of 269 putative genes were identified in McGee, including 237 protein-coding genes, 31 tRNAs, and 1 tmRNA. Among those that could be assigned a function, the virion structure and assembly genes include a major capsid protein (gp101), capsid decoration protein , tail sheath protein (gp129), tail assembly chaperones , tape measure protein (gp134), minor tail proteins , and baseplate wedge protein . The genes involved in McGee\u2019s DNA metabolism include DNA helicase (gp188), thymidylate kinase (gp197), DnaC-like helicase loader (gp201), DnaB-like double-stranded DNA (dsDNA) helicase (gp202), DNA primase (gp204), DnaJ-like chaperonin (gp206), single-stranded DNA (ssDNA) binding protein (gp207), DnaE-like DNA polymerase III \u03b1 (gp208), RF-1 peptide chain release factor (gp209), RecA-like DNA recombinase (gp210), Holliday junction resolvase (gp210), RusA-like resolvase (gp214), Ro-like RNA binding protein (gp233), and multiple proteins with a helix-turn-helix DNA binding domain . Genes involved in lysis, lysin A, holin, and lysin B are encoded by gp252, gp253, and gp254, respectively. The largest gene in McGee is a hypothetical protein gp98  and the smallest gp64 (75\u2009bp long).https://phagesdb.org/), McGee was assigned to phage cluster C1 (Based on its gene content similarity (GCS) of at least 35% to phages in the Actinobacteriophage database (uster C1 , 18, 19.ON637764, while its raw reads have been deposited under Sequence Read Archive accession number SRX14483217.The complete genome sequence for McGee can be found at GenBank under accession number"}
+{"text": "However, the classification strategy and homology group of tet(X)-positive Acinetobacter spp. plasmids remain largely unknown. In this study, we classified them by genome-based replicon typing, followed by analyses of structural characteristics, transferability and in vivo effect. A total of 34 plasmids distributed in at least nine Acinetobacter species were collected, including three tet(X3)-positive plasmids and one tet(X6)-positive plasmid from our genome sequencing results. Among them, there were 28 plasmids carrying Rep_3 superfamily replicase genes and classified into six homology groups, consisting of GR31 (82.1%), GR26 (3.6%), GR41 (3.6%), GR59 (3.6%), and novel groups GR60 (3.6%) and GR61 (3.6%). Our tet(X3)-positive plasmids pYH16040-1, pYH16056-1, and pYH12068-1 belonged to the dominant GR31 group, whereas the tet(X6)-positive plasmid pYH12068-2 was unclassified. Structurally, all tet(X)-positive GR31 plasmids shared similar plasmid replication (repB), stability (parA and parB) and accessory modules [tet(X) and sul2], and 97.6% of plasmid-mediated tet(X) genes in Acinetobacter species were adjacent to ISCR2. Conjugation and susceptibility testing revealed pYH16040-1, pYH16056-1, and pYH12068-2, carrying plasmid transfer modules, were able to mediate the mobilization of multiple antibiotic resistance. Under the treatment of tigecycline, the mortality rate of Galleria mellonella infected by pYH16040-1-mediated tet(X3)-positive Acinetobacter spp. isolate significantly increased when compared with its plasmid-cured strain (p < 0.0001). The spread of such plasmids is of great clinical concern, more effects are needed and will facilitate the future analysis of tet(X)-positive Acinetobacter spp. plasmids.The rapid dissemination of plasmid-mediated  Acinetobacter species is a heterogeneous group of opportunistic pathogens and easily acquires antibiotic resistance genes (ARGs) (tet(X) genes have been reported in at least 10 different Acinetobacter species, including Acinetobacter baumannii, Acinetobacter gandensis, Acinetobacter piscicola, Acinetobacter schindleri, Acinetobacter johnsonii, Acinetobacter indicus, Acinetobacter towneri, Acinetobacter lwoffii, Acinetobacter pseudolwoffii, and Acinetobacter variabilis (tet(X3) and tet(X6) genes were detected with carbapenem resistance gene blaNDM\u20131 in A. baumannii, A. indicus, A. schindleri, and A. lwoffii isolates, posing a serious public health threat  Gram-negative and Gram-positive pathogens . Howevers (ARGs) . To dateriabilis . Worrisoh threat .Acinetobacter spp. genome and plays an important role in the horizontal transmission of ARGs, such as blaNDM\u20131 and blaOXA\u201323 , a series of plasmid classification schemes were developed based on replication initiator protein, mobilization protein and plasmid size (tet(X)-positive Acinetobacter spp. plasmids since the first mobile plasmid-mediated tet(X3) gene in 2019, and their homology groups remained to be analyzed (tet(X)-carrying Acinetobacter spp. plasmids by genome-based replicon typing, followed by analyses of structural characteristics, transferability, and in vivo effect.The plasmid is a self-replicating component of laOXA\u201323 . With thmid size . As the mid size . Howeveranalyzed . Herein,tet(X)-positive Acinetobacter spp. strains in China (tet(X3)-positive Acinetobacter spp. YH16040 and tet(X3)- and tet(X6)-positive Acinetobacter spp. YH12068 from pig; tet(X3)-positive Acinetobacter spp. YH16056 from soil; tet(X3)-positive A. pseudolwoffii YH18001 from human; tet(X4)-positive A. indicus Q22-2, Q85-2, and Q278-1 from migratory bird. In addition, a newly isolated tet(X6)-positive A. baumannii YC103 by CHROMagar\u2122 Acinetobacter plates  containing tigecycline (2 \u03bcg/mL) from duck in 2019 was also analyzed (tet(X)-harboring Acinetobacter spp. plasmids deposited at the NCBI database were collected by tblastn -positive Acinetobacter spp. isolates were sequenced by Oxford Nanopore , respectively. Combining the clean data with our previous Illumina HiSeq data , genome assembly was performed by Unicycler version 0.4.1 and corrected by Pilon version 1.12 (Genomic DNA of eight Seq data  as well ion 1.12 .tet(X)-harboring plasmids were annotated by Rapid Annotation using Subsystem Technology (RAST) version 2.0 -positive structures was conducted by Easyfig version 2.2.5 (All sion 2.0 . Plasmidsion 2.0 . Plasmidsion 2.0 . A maximsion 2.0 . Plasmidme group . Similaron 2.2.5 .tet(X)-mediated tigecycline resistance was evaluated by filter mating with rifampin-resistant Acinetobacter baylyi ADP1 and A. baumannii ATCC 19606 (tet(X) detection and PCR-based fingerprinting (Transferability of plasmid-borne CC 19606 . The putprinting . In paraprinting .tet(X3)-harboring GR31 plasmid pYH16040-1 was cured of Acinetobacter spp. YH16040 using sodium dodecyl sulfate (SDS) with adjustment (tet(X3) gene .The justment . An overX3) gene  and repl1 whereas eravacycline was uninterpreted with no breakpoint. Escherichia coli ATCC 25922 served as a quality control strain.Minimum inhibitory concentrations (MICs) of all strains were determined by broth microdilution and interpreted according to the Clinical and Laboratory Standards Institute guideline M100-Ed28 . The tesG. mellonella  were randomly grouped (16 per group), and then infected with GR31 plasmid-mediated tet(X3)-positive Acinetobacter spp. YH16040 or its plasmid-cured strain YH16040C  via the last left proleg. After incubation at 35\u00b0C for 2 h, the infected larvae were treated with tigecycline (2 \u03bcg/g) or phosphate-buffered saline (PBS) by injection into the last right proleg. Finally, the larvae were observed for survival rates per 24 h in the next 4 days. All in vivo experiments were performed in triplicate.As previously described , the heatet(X)-positive Acinetobacter spp. isolates successfully revealed four tet(X)-carrying plasmids. These included tet(X3)-positive plasmids pYH16040-1 (GenBank accession number: CP094542) from Acinetobacter spp. YH16040, pYH16056-1 (CP094546) from Acinetobacter spp. YH16056, and pYH12068-1 (CP094556) as well as a tet(X6)-positive plasmid pYH12068-2 (CP094557) from Acinetobacter spp. YH12068 (tet(X6)-positive chromosome cYC103 (CP054560) from A. baumannii YC103 was also obtained (tet(X)-positive strains A. pseudolwoffii YH18001, A. indicus Q278-1, A. indicus Q85-2, and A. indicus Q22-2 (tet(X)-harboring plasmids or chromosomes despite repeated attempts. By querying Nanopore raw data of A. pseudolwoffii YH18001 , A. indicus Q278-1 , A. indicus Q85-2 , and A. indicus Q22-2 , a repeated structure consisting of multiple copies of tet(X) genes was detected, respectively, which may lead to the failure of complete sequence assemblies.WGS analyses of eight  YH12068 . Meanwhiobtained . For theus Q22-2 , we failtet(X)-positive Acinetobacter spp. plasmids deposited at the NCBI database has been growing  subtypes located on plasmids (tet(X3) , tet(X5) , and tet(X6) , of which tet(X3) usually coexisted with tet(X6) . Besides tet(X) genes, large amounts of plasmid-mediated genes conferring resistance to tetracyclines, aminoglycosides, \u03b2-lactams, phenicols, sulfonamides, macrolides, lincosamides, and rifamycins were present (tet(X)-positive plasmids ranging from 42,489 to 332,451 bp were widely distributed in A. indicus , A. baumannii , A. towneri , A. schindleri , A. variabilis , A. pseudolwoffii , A. piscicola , Acinetobacter junii , Acinetobacter pittii , and unidentified Acinetobacter spp. strains -positive MDR Acinetobacter spp. plasmids.Since the first report in 2019 , the num n = 34; , includiplasmids , namely  present . Worrisotet(X)-positive Acinetobacter spp. plasmids carried replicase genes and all of them belonged to a Rep_3 superfamily (pfam01051). Therefore, plasmid classification was conducted based on nucleotide sequence alignment with already classified replicase genes (tet(X)-positive Rep_3 superfamily plasmids, a total of six homology groups were successfully identified, such as the dominant GR31 , GR26 , GR41 , and GR59 -positive plasmids pYH16040-1 (CP094542), pYH16056-1 (CP094546), and pYH12068-1 (CP094556) fell within the same group GR31, which has been sporadically detected with tet(X) genes in A. baumannii, A. indicus, A. schindleri, A. towneri, and other Acinetobacter spp. strains (tet(X6)-positive plasmid pYH12068-2 (CP094557) was unclassified due to the lack of replicase genes. In contrast, the replicase genes of tet(X3)- and tet(X6)-carrying plasmids pXMC5X702-tetX-145k (CP084302) and pYH12207-2 (CP048661) have < 75% nucleotide identities with existing homology groups GR1-GR59, and therefore were defined as novel groups GR60  and GR61 , respectively (tet(X)-positive Acinetobacter spp. plasmids.Replicon conserved domain analyses showed that 82.4% (28/34) of se genes . Among t strains . Howeverectively . We provAcinetobacter spp. isolates . In essence, the GR31 plasmids have been detected in 13 validly named Acinetobacter species, including A. baumannii , A. lwoffii , A. indicus , A. towneri , A. schindleri , and others  genes especially in the genera Acinetobacter.In order to evaluate the host range of plasmids belonging to GR26, GR31, GR41, GR59, GR60, and GR61, we conducted a blastn search against the NCBI database and confirmed they were mainly distributed in d others . The plaplasmids . Particuplasmids , but thetet(X)-carrying GR31 plasmids available in the NCBI database. A total of 23 plasmids belonging to GR31 were analyzed and they exhibited GC contents ranging from 39.1 to 46.1%. According to our WGS results, pYH16040-1 was 87,435 bp and harbored 94 putative open reading frames (ORFs), whereas pYH16056-1 was 98,709 bp consisting of 107 ORFs and pYH12068-1 was 100,866 bp consisting of 115 ORFs. Although they originated from different sources, pYH16056-1 and pYH12068-1 showed an average 76.5% nucleotide coverage and 99.8% nucleotide identity to pYH16040-1, with the encoding sequence insertion, deletion and rearrangement  and the accessory modules (tet(X)-positive GR31 plasmids owned a repB gene involved in plasmid replication, and parA and parB genes responsible for plasmid stability. However, only 21.7% (5/23) of them harbored plasmid transfer modules, including conjugal transfer (n = 3) and mobilization genes (n = 2). For accessory modules, they mainly consisted of ARGs  and heavy-metal resistance gene clusters .Predominantly, we compared the complete plasmid sequences of pYH16040-1, pYH16056-1, and pYH12068-1 with angement . Moreoveangement . Archety modules . Our restet(X) genes in Acinetobacter species, including tet(X3) (n = 28), tet(X6) (n = 12), and tet(X5) (n = 2), were further analyzed  and ISAba14 (n = 3). Similarly, the truncation by IS26 was found on GR60 (n = 2) and GR61 (n = 2) plasmids, one of which was also upstream truncated by ISAcsp12. Except the GR26 plasmid carrying a complete ISCR2-mediated transposition unit (n = 1), GR41 and GR59 plasmids were detected with truncation by IS3 (n = 1) and IS1006 (n = 1), respectively. In addition, our WGS results of A. pseudolwoffii YH18001 (JALHBG010000013), A. indicus Q278-1 (JALHBF010000002), A. indicus Q85-2 (JALHBE010000004), and A. indicus Q22-2 (JALHBD010000003) revealed the incomplete tet(X)-carrying structures were also related to ISCR2 (tet(X) variants and ISCR2 as well as truncation by other insertion sequences indicated the frequent recombination events.Genetic environments of all plasmid-mediated analyzed . The res process . For GR3to ISCR2 . The clotet(X3) genes were successfully transferred from Acinetobacter spp. YH16040  and YH16056  to the recipient A. baylyi ADP1 , but failed to A. baumannii ATCC 19606. In Acinetobacter spp. YH12068, the tet(X3)-carrying GR31 plasmid pYH12068-1 (CP094556) couldn\u2019t be transferred, which may be explained by the lack of entire conjugative transfer regions (tet(X6)-positive unclassified plasmid pYH12068-2 (CP094557) carrying plasmid transfer modules was transferred from Acinetobacter spp. YH12068 to A. baylyi ADP1 with a similar efficiency mentioned above. MICs of all transconjugants against tetracyclines increased by at least 32-fold when compared with A. baylyi ADP1, including tetracycline (\u226564 \u03bcg/mL), tigecycline (\u22654 \u03bcg/mL), the newly FDA-approved eravacycline (\u22652 \u03bcg/mL) and omadacycline  gene (CP040912) was also able to be transferred from human-derived A. baumannii to A. baumannii 5AB via electro-transformation, whereas the GR31 plasmid co-harboring tet(X3) and tet(X6) genes (CP044451) was from environmental A. indicus to A. baylyi ADP1 by natural transformation. The transferability of tet(X) genes between Acinetobacter species of human, animal and environment origins reminded us to consider the \u201cOne Health\u201d approach to prevent mobile tigecycline resistance.Conjugation experiments showed the GR31 plasmid-mediated  regions . Meanwhitet(X3)-mediated tigecycline resistance from Acinetobacter spp. YH16040, YH16056, and YH12068 to A. baylyi ADP1, as well as florfenicol resistance (128 \u03bcg/mL) from Acinetobacter spp. YH16040 and YH12068 and gentamicin resistance (16 \u03bcg/mL) from Acinetobacter spp. YH16056, which was consistent with the result of plasmid-mediated ARG mining (tet(X)-mediated tigecycline resistance and blaNDM\u20131-mediated carbapenem resistance between Acinetobacter species (tet(X) genes, as previously described for resistance determinants mcr-1 and blaIMP\u20131  was transferred with G mining . Recentl species . With thblaIMP\u20131 .tet(X3) gene, a plasmid-cured strain YH16040C [tet(X3)-negative] was obtained by serially passaging of tet(X3)-positive Acinetobacter spp. YH16040. For tigecycline, a significant decrease in MIC was detected (1 \u03bcg/mL) when compared with the parental isolate YH16040 , eravacycline (0.06 \u03bcg/mL), omadacycline (0.125 \u03bcg/mL), florfenicol (8 \u03bcg/mL) and sulfamethoxazole-trimethoprim (20 \u03bcg/mL) also exhibited at least 16-fold decrease. The result further confirmed the elimination of GR31 plasmid pYH16040-1 (CP094542) that co-harbored resistance genes tet(X3), floR and sul2  gene was evaluated using a G. mellonella model. 96 h later of tigecycline treatment (2 \u03bcg/g), the larval mortality rate of Acinetobacter spp. YH16040C significantly decreased to 37.5% while Acinetobacter spp. YH16040 was 87.5%  gene compromised the clinical effectiveness of tigecycline. Notably, the infection by Acinetobacter spp. YH16040 and Acinetobacter spp. YH16040C under the treatment of PBS resulted in a greater larval mortality rate (100%) than PBS only (12.5%) after 48 h (p < 0.0001), whereas no significant difference was observed between Acinetobacter spp. YH16040 and Acinetobacter spp. YH16040C -positive Enterobacteriaceae bacteria have also been constructed, and therefore G. mellonella was available for in vivo functional analyses of plasmid-mediated tet(X) genes.Subsequently, the tet(X)-carrying MDR plasmids in Acinetobacter species. Our results classified the tet(X)-positive Acinetobacter spp. plasmids of Rep_3 superfamily into six homology groups, including two newly assigned GR60 and GR61. To the best of our knowledge, this study first revealed a dominant GR31 plasmid mediating the horizontal transfer of tigecycline resistance gene tet(X) across different Acinetobacter species and contributed to the failure of tigecycline treatment in vivo. Accordingly, more efforts are needed to monitor and prevent the plasmid-mediated tet(X)-positive Acinetobacter spp. strains.Taken together, the data presented in this study highlighted the diverse distribution of The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/J-LH, Y-HL, and JS designed the study. CC, P-YH, C-YC, and QH performed the experiments. CC and P-YH analyzed the data. CC wrote the draft of the manuscript. All authors reviewed, revised, and approved the final report."}
+{"text": "Azadirachta indica L. and Melia azedarach L., belonging to Meliaceae family, have been shown to have medicinal benefits and are extensively employed in traditional folk medicine. Herein, HPLC analysis of the ethyl acetate fraction of the total methanolic extract emphasized the enrichment of both A. indica L., and M. azedarach L. leaves extracts with phenolic and flavonoids composites, respectively. Besides, 4 limonoids and 2 flavonoids were isolated using column chromatography. By assessing the in vitro antiviral activities of both total leaves extracts against Severe Acute Respiratory Syndrome Corona virus 2 (SARS-CoV-2), it was found that A. indica L. and M. azedarach L. have robust anti-SARS-CoV-2 activities at low half-maximal inhibitory concentrations (IC50) of 8.451 and 6.922 \u03bcg/mL, respectively. Due to the high safety of A. indica L. and M. azedarach L. extracts with half-maximal cytotoxic concentrations (CC50) of 446.2 and 351.4 \u03bcg/ml, respectively, both displayed extraordinary selectivity indices (SI>50). A. indica L. and M. azedarach L. leaves extracts could induce antibacterial activities against both Gram-negative and positive bacterial strains. The minimal inhibitory concentrations of A. indica L. and M. azedarach L. leaves extracts varied from 25 to 100 mg/mL within 30 min contact time towards the tested bacteria. Our findings confirm the broad-spectrum medicinal value of A. indica L. and M. azedarach L. leaves extracts. Finally, additional in vivo investigations are highly recommended to confirm the anti-COVID-19 and antimicrobial activities of both plant extracts.The leaves of  Drug resistance to infections in humans brought on by microorganisms including bacteria, viruses and fungi is a life-endangering phenomenon with considerable number of mortalities worldwide. Medicinal herbs were applied to produce a plenty of pharmaceuticals that are currently used to alleviate various health disorders or diseases. Numerous researches are performed to find out new antimicrobial treatment alternatives that really can prevent the spread of bacteria or eradicate them without endangering humans by conducting comprehensive screenings of medicinal herbs , 2.In December 2019, the SARS-CoV-2 virus has emerged as an unknown severe respiratory illness, namely corona virus illness 2019 (COVID-19) . Few weeAzadirachta indica L. (Neem) and Melia azedarach L. (China tree) are popular medicinal herbs that belong to the Meliaceae family. A. indica L, originated in India, is extensively scattered in all countries , 1H-NMR : 7.11 , 5.45 , 2.89 ,4.29 , 4.89, 4.95, 2.67 , 1.91, 1.64 , 7.39 , 6.21 , 6.93 , 1.70 , 2.15 , 1.10 . 13C-NMR : 158.2 (C-1), 126.4 (C-2), 206.1 (C-3), 41.3 (C-4), 50.4 (C-5), 69.8 (C-6), 76.9 (C-7), 43.7 (C-8), 40.2 (C-9), 41.3 (C-10), 16.1 (C-11), 28.3 (C-12), 45.7 (C-13), 72.8 (C-14), 57.5 (C-15),33.1 (C-16), 40.2 (C-17), 122.8 (C-20), 141.8 (C-21), 111.2 (C-22), 138.7(C-23), 21.51, 171.81 (OAC), 21.9 (C-methyls). From the formentioned spectral data, this compound was identified as 14,15-\u03b2-epoxynimonol + 612 for C35H48O9, 549 (100%), 449, 389 and 371. Additionally, 1H-NMR, and13C-NMR data were in agreement with previous studies + 694 for C40H54O10, 649 for ethoxyl group loss at C-12, then 449 for two tigloyl side chain loss, followed by 389(100%) due to acetyl group loss and subsequently water loss giving 371 which are in accordance to nimbolinin limonoids fragmentation pattern. The 1H and 13C NMR data were similar to  +, 552, 537 [M\u2013H2O + H]+. The data were consistent with the molecular formula C35H54O5. The 1H-NMR and13C-NMR data confirmed this compound as 3-\u03b1-tigloylmelianol which was previously identified from M. azedarach fruits by [Compound 6: yellow amorphous powder, m.p. 164\u00b0C. EI-MS (m/z): 462 (M+) for C22H22O11, in addition to 300, 285, 257, 152, 137, 116. IR data, UV absorbance in different shift reagents and 13C-NMR data characterized this compound as chrysoeriol 7-O-glucoside. This compound has been isolated for first time in the present study. The structures of the isolated compounds were illustrated in Fig 1.50), the half maximal inhibitory concentration 50 (IC50) and selectivity index (SI)(CC50/IC50) against SARS-CoV-2 were individually evaluated (Table 2). Both leaves extracts of A. indica and M. azedarach showed high safety in Vero E6 cells with CC50 values of 446.2 \u03bcg/mL for A. indica and 351.4 \u03bcg/mL for M. azedarach, respectively. Additionally, both extracts exhibited promising inhibition of SARS-CoV-2 replication in Vero E6 cells at potent IC50 concentrations of 8.541 \u03bcg/mL for A. indica and 6.922 \u03bcg/mL for M. azedarach . Interestingly, both extracts exhibited extraordinary selectivity indices (SI) (>50) (Table 2). The determined SI values are meaningfully greater than those recommended for further assessment of the bioactive compounds (> 10) [A. indica and M. azedarach indicate the bioactive potential of A. indica and M. azedarach leaves extracts. Compared to the reference drug control remedesivir , both plant extracts showed comparable safety and antiviral activities with slightly lower selectivity indices. These data demonstrate that both extracts can be considered as promising candidates for further in vitro and in vivo studies against SARS-CoV-2 virus and potentially other viruses.To appraise the antiviral efficacy of leaves extracts against SARS-CoV-2 virus, the half maximal cytotoxic concentration (CCs (> 10) , 39. TheA. indica extracts against the novel SARS-CoV-2 virus [et al. [To our knowledge, there is a few previous studies that assessed the antiviral activity of -2 virus , 41. In  [et al.  concludeBorkotoky and Banerjee , isolatein vitro viral inhibitory activity of M. azedarach L. leaves extract against SARS-CoV-2 which is so far not studied. Nethmini et al. found that M. azedarach L. could turn off the replication of herpes simplex virus, coxsackie virus, enterovirus, influenza A virus, and bovine rhinovirus in both epithelial and macrophage cell lines [In this study, we explored the ll lines .in vitro anti-SARS-CoV-2 effect in this study for both plant leaves extracts, the inhibitory zone assay was done to assess the in vitro antibacterial properties of the two leaves extracts against E. coli, P. aeruginosa, S. aureus, L. monocytogenes, and E. faecalis using agar well-diffusion assay. The obtained results indicated that 50 \u03bcg/ml of A. indica L. and M. azedarach L. displayed high potency for inhibiting a broad spectrum of examined Gram-negative (GN) species (.In addition to the Table 3). Results revealed that the size of ZOI of vancomycin against the selected pathogens; E. coli, S. enterica, P. aeruginosa, S. aureus, L. monocytogenes, and E. faecalis using disc diffusion assay were ranged between 7 and 11 mm as mentioned in Table 3).Simultaneously, two reference antibiotics drugs vancomycin (30 \u03bcg/disc) and ciprofloxacin (5 \u03bcg/disc) were applied in this study as a positive control . ResultsThese results show the tendency that the GN bacterial species were more susceptible to all tested plant extracts than the GP species. Conversely, the width of ZOI around discs was lower than those around wells. Our data are in agreement with previous study , in whicA. indica L. and of M. azedarach L. extracts were examined to estimate MIC values to assess their antibacterial potential during various exposure times  to different bacterial strains. As shown in Figs 4, the extracts demonstrated a notable bactericidal effect against all the tested bacterial strains. The detected MIC values depend on the concentrations (mg/mL) and exposure time (min). The consequences of MICs exhibited by the extract of M. azedarach displayed that the perfect antibacterial effects against E. coli (MIC = 50 mg/mL within 30 min), S. enterica (MIC = 25 mg/mL within 30 min), P. aeruginosa (MIC = 25 mg/mL within 30 min), S. aureus (MIC = 75 mg/mL within 30 min), E. faecalis (MIC = 25 mg/mL within 30 min) and L. monocytogenes (MIC = 75 mg/mL within 30 min). The measured MIC values of A. indica plant extract were lower in GN than GP bacteria. The enhanced antibacterial activity against GN could be due to variation in active compounds in M. azedarach extract and their penetration ability through bacterial cell wall and membrane [The different concentrations (10\u2013100 mg/mL) of membrane . This fiM. azedarach extract. The results represented in Fig 4 revealed significant antibacterial effects against all test bacteria. Also here, the calculated MIC values depended on the applied extract concentrations (mg/mL) and contact time (min). The results of MIC exhibited that the extract has clear bactericidal effect against E. coli (MIC = 50 mg/mL within 30 min), S. enterica (MIC = 25 mg/mL within 30 min), P. aeruginosa (MIC = 50 mg/mL within 30 min), S. aureus (MIC = 75 mg/mL within 30 min), E. faecalis (MIC = 75 mg/mL within 30 min) and L. monocytogenes (MIC = 100 mg/mL within 30 min). Interestingly, the results showed that the M. azedarach extract possess higher antibacterial activities against the tested GN bacteria in comparison to GP bacteria, which is matching with what we detected in case of A. indica plant extract except that MIC of Listeria monocytogenes was higher than S. aureus and E. faecalis .Using the same procedure and bacterial strains, we also assessed the antibacterial activity of Table 4. The results gained revealed that there was an exceptional positive correlation with strong significance between GP and GN species and the concentrations of all plant extracts. On the other hand, results perceived that a moderate correlation had been reported for E. coli.From statistical analysis results, the potential correlation with significance between various concentrations of particular extract towards all the targeted pathogenic bacterial strains was briefly reported in S. aureus, vancomycin-resistant enterococci, and Mycobacterium tuberculosis are widely documented as challenging hospital-associated infections to treat with known antibiotics. To this point, medicinal plants are recently applied to provide a supplementary treatment line against these bacterial infections [M. azadirachta L. and A. indica L. showed powerful antibacterial activities against various GP and GN species. The Meliaceae family has a substantial percentage of polyphenolic chemicals that could possess oxidative activities. A. indica L. phytochemicals have such a broad range of biological characteristics, especially antibacterial and cancer-fighting potential [M. azedarach L. towards numerous types of bacterial pathogens [As a public biohazard with millions of deaths every year, antibiotic resistance emerged recently due to the continuous prescription of improper antibiotics at non-lethal doses. Methicillin-resistant fections . M. azadotential . This bootential . Furtherathogens . Limonoiathogens .M. azedarach L. was established to be biocidal agent against P. aeruginosa, and E. coli, the lowest activity was documented in aqueous extract of M. azedarach against E. coli (8.5 mm) and S. aureus (8.2 mm). In previous study, E. coli showed the lowest sensitivity to methanolic extract (6 mm), and no activity was observed in 10, 20, 30 \u03bcg/ml of methanolic extract of M. azedarach L. [M. azedarach L. seed extracts efficiently manage illnesses caused on by both GP and GN species and reported that the that ethyl acetate fraction exhibited greatest suppression, followed by aqueous and methanol-based extracts, each of which reduced the proliferation of all the studied pathogenic strains. The petrol and benzene extracts displayed antimicrobial activity versus 15 pathogens, however, in comparison to the polarity extracts. The findings previously illustrate that seed extracts are powerful antibiotics that can effectively prohibit both disease-caused pathogens to human infections.The ethanolic extract of arach L. . PreviouA. indica L. and M. azedarach L. leaves were chemically characterized and were found to be rich in phenolic compounds and flavonoids. In vitro assessment of both extracts revealed that they have robust antiviral activity against SARS-CoV-2 as well as antibacterial effects against broader spectra of GN and GP bacteria. These data emphasize their broad-spectrum medicinal value and demand further in vitro and in vivo investigations as anti-SARS-CoV-2 candidate and antibacterial agents."}
+{"text": "Myasthenia gravis (MG) is an autoimmune disorder affecting neuromuscular junctions. Cytokines play important roles in facilitating the immune response and augmenting the pathogenic antibody production. The current study aims to sensitively characterize the serum levels of cytokines with very low concentration in generalized MG (gMG).Using ultrasensitive single-molecule arrays (SIMOA), we measured serum IL-2, IL-4, IL-5 and IL-12p70 in 228 participants including 152 immunotherapy-na\u00efve anti-acetylcholine receptor (AChR) subtype gMG from Huashan MG registry and 76 age-matched healthy controls. Subgroup analysis was then performed by stratifying patients according to the onset ages, MGFA classification, disease duration at baseline.P\u2009<\u20090.0001; 0.029\u00a0pg/mL versus 0.018\u00a0pg/mL, P\u2009=\u20090.0259; 0.215\u00a0pg/mL versus 0.143\u00a0pg/mL, P\u2009=\u20090.0007; 0.132\u00a0pg/mL versus 0.118\u00a0pg/mL, P\u2009=\u20090.0401). Subgroup analysis revealed that IL-2 levels were slightly elevated in gMG with MGFA II compared to MGFA III/IV , as well as elevated levels of IL-2  and IL-5  in late-onset gMG compared with the early-onset gMG. gMG patients with a long duration had a significant increased serum IL-12p70 than those with a short duration .Serum IL-2, IL-4, IL-5 and IL-12p70 levels were significantly elevated in gMG compared to controls (0.179\u00a0pg/mL versus 0.011\u00a0pg/mL, Serum IL-2, IL-4, IL-5 and IL-12p70 levels were increased in AChR subtype gMG using ultrasensitive measurement. Serum cytokines with very low concentrations may provide as potential biomarkers in stratifying gMG patients in future prospective cohort studies. Myasthenia gravis (MG) is characterized by fluctuating muscle weakness caused by autoantibodies against the acetylcholine receptor (AChR) and the muscle-specific tyrosine kinase (MuSK) , 2. RecePrevious studies on MG-relevant inflammatory cytokines mainly focused on Interleukin (IL)-17 (Th17 related), IL-21(Tfh related) and IL-6  \u201312. In cUltra-sensitive single-molecule arrays (SIMOA) provides an alternative and more sensitive method for detecting trace cytokines. , 17. HerA total of 228 participants were finally enrolled. Of these, 152 gMG patients  were immunotherapy na\u00efve with positive anti-AChR antibodies and 76 age-matched participants were healthy controls . The median anti-AChR antibodies titer for MG is 5.86  nmol/L and thymoma concurrence is 9.9%. We further divided 152 AChR subtype gMG patients into subgroups: (1) Clinical severity according to the Myasthenia Gravis Foundation of America (MGFA): mild group with MGFA II (n\u2009=\u2009100) and moderate to severe group with MGFA III/IV (n\u2009=\u200952); (2) Onset ages: early-onset MG  and late-onset MG ; (3) Disease duration: short disease duration group (n\u2009=\u200976) and long disease duration group (n\u2009=\u200976). Detailed information regarding each subgroup are listed in Table The levels for serum IL-2, IL-4 and IL-5 was higher in immunotherapy-na\u00efve gMG in comparison to HCs (measurable in 98.7% (150/152) versus 82.9% (63/76) for IL-2; 99.3% (151/152) versus 78.9% (60/76) for IL-4; 99.3% (151/152) versus 85.5% (65/76) for IL-5, respectively. For IL-12p70, the levels were equally matched in gMG and HCs (measurable in 83.6% (127/152) versus 84.2% (64/76)).P\u2009<\u20090.0001; 0.029 [0.016\u20130.045] pg/mL versus 0.018 [0.011\u20130.044] pg/mL, P\u2009=\u20090.0259; 0.215 [0.129\u20130.360] pg/mL versus 0.143 [0.077\u20130.243] pg/mL, P\u2009=\u20090.0007; 0.132 [0.092\u20130.202] pg/mL versus 0.118 [0.073\u20130.167] pg/mL, P\u2009=\u20090.0401) . In contrast, there were no differences in serum IL-4 levels , IL-5 levels , IL-12p70 levels  between these two subgroups . In parallel, serum IL-5 was also elevated in patients with late-onset gMG . In contrast, there were no difference in serum IL-4 nor IL-12p70 levels between these two gMG groups with different onset age  , while no significant differences in IL-2 , IL-4  and IL-5   and a negative correlation with disease course , baseline MG-ADL , QMG  and MG-QoL15 . In parallel, serum IL-5 had a positive correlation with age  and a negative correlation with QMG . Serum IL-4 was negatively correlated with disease course , while IL-12p70 was positively correlated with disease course . Multivariate analyses indicated that IL-4 levels had positive impact on QMG scores and MMT scores , while cytokines IL-2, IL-5 and IL-12p70 showed no statistically significant difference  score, MGFA-quantitative MG test (MGFA-QMG), MG quality of life 15-item (QoL-15) questionnaire, and MG manual muscle test (MMT). Serum IL-2 had a positive correlation with age . The other cytokines showed no statistically significant difference.Moreover, the correlations between baseline cytokine levels and the short-term prognosis at 12\u00a0months after enrollment were assessed. Baseline IL-12p70 was positively correlated with MG-ADL score at 12\u00a0months-follow up , which have been demonstrated decreased and impaired in MG patients , 29. OngRecent clinical studies had emerged regarding the difference in presentations and outcomes between the EOMG and LOMG cohorts. The majority of LOMG had ocular phenotype. The presence of neither anti-titin nor anti-MuSK antibodies points to an unfavorable outcome . Still tAlthough the concordant elevation in type 1 and type 2 cytokines were demonstrated in gMG, the correlations between baseline cytokines with the short-term clinical outcome revealed some difference. IL-12p70 promotes Th1 immunity thus imposes positive effect in longitudinal autoimmune response for MG. However, the correlations are relatively weak to make any conclusions.+ T lymphocytes and B cells are required to better delineate the role of cytokines in the immunological network. Third, given the current cost and the availability, SIMOA may not be adopted for routine diagnostics in a short time. Ultrasensitive measurement of serum cytokines with very low concentrations in a large longitudinal prospective MG cohort is anticipated in future investigations to monitor the outcome and therapeutic response.Several limitations in this study are needed to be addressed. First, the normal datasets for all age groups are not available for these cytokines measured by ultrasensitive analysis. Second, the concurrent measurement of peripheral CD4We confirmed the high levels of serum IL-2, IL-4, IL-5 and IL-12p70 in a cohort of AChR subtype gMG patients. In particular, further analysis revealed significant difference in serum levels of these cytokines in gMG subgroups divided by clinical severity, onset ages and the prior disease duration. These inflammatory serum cytokines may provide as disease biomarkers in stratifying gMG patients in future prospective cohort studies.This study was conducted in accordance with the ethical standards established in the 1964 Declaration of Helsinki and was approved by the Medical Ethics Committee of Huashan Hospital affiliated to Fudan University . Written informed consent was obtained from each participant.There are 1662 MG patients registered in National center for neurological disorders, huashan hospital, Shanghai from August 8, 2012, through May 31, 2021. MG mimicking diseases including Lambert-Eaton myasthenic syndrome, peripheral neuropathy, myopathies, and motor neuron diseases were excluded before the recruiting. The patients who have already enrolled in previous study for 18 cytokine measurement have been excluded in current study  and group comparisons were analyzed using Mann Whitney test as the data were previously tested for not following the normal distribution. Categorical variables were presented as No. (%) and analyzed by Chi-square test. Correlations between serum cytokines and clinical variables of MG cohort were analyzed using Spearman\u2019s method. Multiple linear-regression analyses were then further conducted. We considered a two-tailed adjusted"}
+{"text": "Itraconazole is first-line treatment for mild-moderate blastomycosis and consolidation of moderate-severe disease. Itraconazole is metabolized to 3 metabolites, including an active metabolite hydroxy-itraconazole. The sum of itraconazole and hydroxy-itraconazole levels > 1.0 mcg/mL is guideline recommended for treatment of invasive fungal infections; conversely, some experts suggest targeting a parent compound level alone > 1.0 mcg/mL. This study aims to compare clinical outcomes and adverse drug reactions (ADRs) of combined itraconazole and hydroxy-itraconazole levels > 1.0 mcg/mL versus itraconazole parent compound alone > 1.0 mcg/mL in patients with blastomycosis.Blastomyces infection who received itraconazole with at least one documented serum itraconazole level. The primary outcome was rate of partial or complete treatment response in patients with a combined itraconazole and hydroxy-itraconazole level > 1.0 mcg/mL versus itraconazole parent compound > 1.0 mcg/mL for > 75% of measured levels. ADRs attributable to itraconazole were compared between the groups. Treatment response rates and ADRs were compared between groups using two proportion z-tests.This study is a single-center, retrospective chart review of patients \u2265 18 years with probable or proven Total of 80 patients were included: 36 = combined itraconazole and hydroxy-itraconazole > 1.0, 32 = itraconazole parent alone > 1.0, 12 = 75% of all levels < 1.0. No statistically significant difference was observed between groups for blastomycosis rate of partial or complete treatment response . Significantly higher mortality was observed in patients failing to achieve itraconazole or itraconazole/hydroxy-itraconazole > 1.0 . There was no significant difference in total ADRs between the three groups (p=0.56).This limited evidence supports an itraconazole therapeutic target combining itraconazole and hydroxy-itraconazole > 1.0 for blastomycosis treatment.Paschalis Vergidis, MD, AbbVie: DSMB|Cidara: Grant/Research Support|Scynexis: Grant/Research Support Christina G. Rivera, PharmD, Gilead: Grant/Research Support|Gilead: Honoraria|Insmed: Honoraria."}
+{"text": "These insect-resistant inbred lines can be used as parents in maize breeding programs to develop new varieties.Maize seedlings contain high amounts of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA), and the effect of DIMBOA is directly associated with multiple insect-resistance against insect pests such as Asian corn borer and corn leaf aphids. Although numerous genetic loci for multiple insect-resistant traits have been identified, little is known about genetic controls regarding DIMBOA content. In this study, the best linear unbiased prediction (BLUP) values of DIMBOA content in two ecological environments across 310 maize inbred lines were calculated; and their phenotypic data and BLUP values were used for marker-trait association analysis. We identified nine SSRs that were significantly associated with DIMBOA content, which explained 4.30\u201320.04% of the phenotypic variation. Combined with 47 original genetic loci from previous studies, we detected 19 hot loci and approximately 11 hot loci  supported pleiotropy for their association with two or more insect-resistant traits. Within the 19 hot loci, we identified 49 candidate genes, including 12 controlling DIMBOA biosynthesis, 6 involved in sugar metabolism/homeostasis, 2 regulating peroxidases activity, 21 associated with growth and development [(auxin-upregulated RNAs (SAUR) family member and v-myb avian myeloblastosis viral oncogene homolog (MYB)], and 7 involved in several key enzyme activities . The synergy and antagonism interactions among these genes formed the complex defense mechanisms induced by multiple insect pests. Moreover, sufficient genetic variation was reported for DIMBOA performance and SSR markers in the 310 tested maize inbred lines, and 3 highly (DIMBOA content was 402.74\u2013528.88 \u03bcg g Zea mays), is an important agro-economical crop that is utilized globally as food, animal feed, and biofuel products with production of more than 1.14 billion tons in 2018 [Ostrinia furnacalis; Lepidoptera, Pyralidae) is one of the most destructive insect pests in maize. Newly hatched ACB larvae primarily feed on leaves during the whorl stage; subsequently, the 3rd or 4th instars bore into the stalk. This may cause yield losses of 10 to 30% in the outbreaks recorded in China [Rhopalosiphum maidis; Homoptera, Aphididae), especially in tropical and warmer temperature regions [Maize  provides a fine-mapping method that enables researchers to identify functional variation in a broader germplasm background . Both QT\u22121 FW (ZY19-Jiu1101) with an overall mean of 94.21\u03bcg g\u22121 FW; similarly, the average DIMBOA content of these seedlings in the E2 environment varied from 8.84 (T58) to 493.40 \u03bcg g\u22121 FW (RX20-1006) with an overall mean of 93.26 \u03bcg g\u22121 FW , umc2314  and umc2031  exhibited the maximum number of alleles  . It is wIn addition, the population structure of these inbred lines was analyzed by STRUCTURE software. When various groups  ranging from 1 to 12 were compared, the \u0394K reached the maximum value when K = 5, thus the 310 maize inbred lines were divided into five optimal groups A. The in\u22121 FW; accounted for 0.97%) had three genotypes, which were 19LX-1230, RX20-1006, and ZY19-Jiu1101; they were defined as high insect-resistant germplasms. Type II  had 15 inbred lines including F1227, M1005, F0501, LongF1008, 1201, M1009, PH1CRW, 1512, F3202, F1220, F3208, F2260, F1233, F2303, and F3210, and they were defined as moderate insect-resistant germplasms. Type III  had 27 genotypes, including ShanM3304, M1001, MeizaS2\u20133, Jizaoyu, M0803, F0306, M10202, ly6305, F2211, M1202, F0311, M1409Ying, F1501, M0124, M0822, M0105, F2502, M0505, PHHJC, 8723-2, M0306, F2222, F2213, M0824, M0125, 747, and F3226, and they were defined as insect-resistant germplasms. Type IV  included 52 moderate insect-susceptible germplasms. Type V  included 213 insect-susceptible germplasms . Type I (DIMBOA content in E1/E2: 467.11\u2013528.88/402.74\u2013493.40 \u03bcg grmplasms C. Furthermplasms B. The dahttps://www.maizegdb.org/data_center/map (accessed on 18 September 2022)) (p < 0.01) SSR loci associated with DIMBOA content in both environments (E1 and E2) by GLM and MLM, respectively; these SSR loci were in Bin 1.04, Bin 1.11, Bin 2.01, Bin 4.00, Bin 4.01, Bin 6.02, Bin 8.04, and Bin 10.04. The phenotypic variation explained by these SSR loci ranged from 4.30%  in the E1 environment to 20.04%  in the E2 environment via GLM, and ranged from 4.74%  in the E2 environment to 10.57%  in the E1 environment via MLM (p < 0.01) associated with BLUP values by GLM and MLM, respectively, which explained 5.41% \u201319.42%  by GLM, and explained 9.29% \u201313.70%  by MLM  of 186 SSRs on an IBM2 2008 Neighbors map frame (r 2022))  to buildr 2022)) . Then, t via MLM ; Table 1) by MLM ; Table 1Next, we attempted to obtain hot genetic loci for multiple insect-resistant traits of ACB and CLA to lay a foundation for fine mapping and candidate gene prediction, verification, and breeding application. First, we collected 47 original genetic loci for ACB-/CLA-resistant traits including DIMBOA content, tunnel length of corn borer (TL), aphid incidence rate (AIR), average aphid incidence grade (AIG), LFR, HO, tunnel length/number of holes of corn borer (TL/HO), and aphid resistance (AR) from previous studies , and comGRMZM2G085381 (bx1), GRMZM6G617209 (bx6), GRMZM2G441753 (bx7), GRMZM2G311036 (bx10), and GRMZM2G336824 (bx11) to examine their relative expression levels in maize seedlings of RX20-1006 (high insect-resistant line) and T58 (insect-susceptible line) in the E1 environment at the V6 stage using real-time PCR (RT-qPCR). The results showed that the relative expression levels of the five genes were significantly correlated with the DIMBOA content in the two maize genotypes , DIMBOA, and 6-methoxy-3h-1,3-benzoxazol-ne (MBOA) are multifunctional defense metabolites that can protect maize against insect pests feeding and pathogens ,32,33. Df 42\u201396% . Indeed,f 42\u201396% ,34,35, sf 42\u201396%  also repf 42\u201396%  further f 42\u201396% . Understanding the genetic basis of the multiple insect resistance of maize is critical to the control of combinatorial attacks of ACB and CLA in the field. In this study, we observed a key trait for multiple insect resistance, i.e., the DIMBOA content in two ecological environments and their BLUP values, to detect nine significant associated SSR markers using association mapping via GLM and MLM across 310 diverse maize inbred lines from Gansu Province, China; these nine SSR markers were located in Bin 1.04, Bin 1.11, Bin 2.01, Bin 4.00, Bin 4.01, Bin 6.02, Bin 8.04, and Bin 10.04, respectively ; Table 1r); r = 0.252) and TL/HO (r = 0.229) in 162 F3 maize population to ACB resistance [Interestingly, we identified 19 hot loci (Loci 1-Loci 19) involved in maize multiple insect resistance in the present study ; Table 3According to the physical interval of the above 19 hot loci controlling 8 insect-resistant traits and the GO annotations of corresponding genes in these hot intervals, a total of 49 candidate genes were identified ; they maGRMZM2G311036 (bx10), GRMZM2G336824 (bx11), and GRMZM2G023325 (bx12) were detected in Loci 1 (Bin 1.04), and they encoded DIMBOA-glucoside O-methyltransferase; GRMZM2G046163  was mapped in Loci 3 (Bin 1.11); GRMZM2G167549 , GRMZM2G172491 ), GRMZM2G063756 ), GRMZM6G617209 , GRMZM2G085054 -one 2-D-glucosyltransferase), GRMZM2G085381 , and GRMZM2G085661  were identified in Loci 8 (Bin 4.00\u20134.03); and GRMZM2G441753 -one (TRIBOA-Glc) O methyl transferase) was located in Loci 9 (Bin 4.04)  converted free indole to 2,4-dihydroxy-1,4-benoxazin-3-one (DIBOA) [bx1 was a modified form of the tryptophan synthase alpha subunit, and it was expressed constitutively in young seedlings, while igl was induced in more advanced stages of plant development and contributed to the blend of odors that attracted beneficial parasitoids [bx8 and GRMZM2G161335 (bx9) were involved in the glucosylation of DIBOA [bx6 was responsible for the hydroxylation step that converted DIBOA-Glc to TRIBOA-Glc, and this conversion likely took place in the cytosol [bx7, making DIMBOA-Glc [bx10, bx11, and bx12 were likely candidates for catalyzing the conversion of DIMBOA-Glc to HDMBOA-Glc [The 12 candidate genes were identified within three hot loci; namely, in 4.04) . The lgls larvae ,39,40. T (DIBOA) . The bx1asitoids ,39,40. T cytosol ,43. MethMBOA-Glc . In addiGRMZM2G150256 (mir2) and GRMZM2G150276 (mir1) were mapped within Loci 14 and encoded a maize insect resistance-cysteine protease (key defensive protein) against chewing insect pests in maize , Loci 5 (Bin 2.03\u20132.04), Loci 12 (Bin 5.03), Loci 17 (Bin 8.06), and Loci 18 (Bin 9.01), respectively . SimilarLOX genes, i.e., GRMZM2G017616 (LOX9) within Loci 1 (Bin 1.01\u20131.02), GRMZM2G040095 (LOX6) within Loci 4 (Bin 2.00\u20132.01), and GRMZM2G102760 (LOX5) within Loci 12 (Bin 5.03)  plays critical roles in plant defense against multiple insect pests and pathogens ,45,46; ain 5.03) . LOX6 wain maize .MYB transcription factor can interact with mRNA/proteins to form a fine regulatory network to activate the expression of downstream defense genes and induce insect-resistance defense response. Interestingly, the previous findings [MYB genes were validated within Loci 2 (Bin 1.04), Loci 5 (Bin 2.03\u20132.04), Loci 10 (Bin 4.05), Loci 13 (Bin 5.05\u20135.07), Loci 14 (Bin 6.02), Loci 15 (Bin 8.01\u20138.03), Loci 16 (Bin 8.04\u20138.05), and Loci 19 (Bin 10.04) in this study, respectively , GRMZM2G018716 (incw7), and GRMZM2G018692 (incw6) within Loci 4 (Bin 2.00\u20132.01), GRMZM2G139300 (incw1) within Loci 13 (Bin 5.05\u20135.07), and GRMZM2G123633 (incw3) within Loci 19 (Bin 10.04) were identified in this study  within Loci 16 (Bin 8.04\u20138.05)  catalyze the irreversible hydrolysis of sucrose into glucose and fructose and play important roles in sucrose partitioning, plant development, and defense responses to biotic stresses . The hydis study . Essmann04\u20138.05) . Thereby2O2 induces POD activity, which then oxidizes and polymerizes p-coumaryl/coniferyl-/sinapyl-alcohol into lignin monomers on the cell wall [ZmPrx35 as the prevailing POD was involved in defense against pathogens and insects. Similarly, we also identified Zm00001d024752 (POD21) and Zm00001d052335 (POD23) within Loci 19 (Bin 10.04) and Loci 11 (Bin 4.08), respectively  responsible for ubiquitin-conjugating enzyme 4, RMZM2G130224 responsible for restriction endonuclease type II-like superfamily protein, and GRMZM2G504910 responsible for tetratricopeptide repeat protein 27 homolog, were identified within Loci 4 (Bin 2.00\u20132.01), Loci 6 (Bin 2.04), and Loci 7 (Bin 2.05) in the present study, respectively (GRMZM2G130225 (near rs658849) and GRMZM2G102471 (near rs624256) were also identified to be involved in TL of ACB using GWAS analysis [In addition, three other candidate genes, i.e., ectively . Interesanalysis . In summary, according to the above studies, a possible molecular network underlying maize multiple insect-resistance to ACB and CLA was constructed , which cWhen assessing the genetic diversity in maize genotypes, SSR remains the preferred choice due to their co-dominant and multi-allelic nature, abundance, and loci specificity . In the In this study, the collected 310 elite maize inbred lines from different ecological environments  in Gansu Province, China . A totalv/v) was added to each tube. The tubes were rotated and placed in the dark for 12 h and then centrifuged at 12,000 rpm  for 20 min at 4 \u00b0C. Supernatants (600 \u03bcL) were slowly passed the corresponding Millex\u00ae needle filter and transferred into auto-sample vials for analysis by HPLC. Standard DIMBOA (CAS No.: 15893-52-4) was purchased  and was used to optimize the mass spectrometric parameters and fragment spectra.The DIMBOA content was determined using high-performance liquid chromatography . Namely, freeze-dried leaves (0.2 g per sample) were homogenized and weighted into screw-capped 10 mL polypropylene centrifuge tubes, and 5 mL of HPLC grade methanol-methanoic acid solution ) were used to perform SSR analysis. Fragments were separated using polyacrylamide gel electrophoresis. The polymorphism information content (PIC) value and Shannon-Wiener\u2019s index (I) value were determined as follows [http://web.stanford.edu/group/pritchardlab/structure_software/release_versions/v2.3.4/html/structure.html (accessed on 10 August 2022)) for the assessment of groups and genetic relationships among the 310 maize inbred lines. The project was run with the set parameters of the population admixture model, and the allele frequency correlated. The optimum group number was determined by \u0394K value [https://www.maizegdb.org/data_center/map (accessed on 18 September 2022)) using BioMercator v. 4.2 software (http://www.bioinformatics.org/mqtl/wiki/ (accessed on 18 September 2022)). The linkage disequilibrium (LD) values for r2 [https://tassel.bitbucket.io/ (accessed on 10 August 2022)), following a permutation test of 10,000. The K matrix and marker-DIMBOA content association mapping was completed in Tassel 3.0 software using the genotypic data of 186 polymorphic SSRs and phenotypic data on DIMBOA content for a set of 310 inbred lines. The association analysis was conducted by a GLM with Q matrix  [2) of the marker at p < 0.01 level and with the lowest false discovery rate (FDR). Using MaizeGDB (http://www.maizegdb.org/ (accessed on 20 September 2022)) and nucleotide and primer blast tools, the physical locations of associated SSR markers for DIMBOA content were determined on chromosomes.Genomic DNA was extracted from the 6th leaf of 310 inbred lines using the cetyltrimethyl ammonium bromide (CTAB) method . Then, a follows :(1)PIC=1tructure  was anal\u0394K value . A linkas for r2  and D\u2032 [s for r2  between ulation)  and an Mulation) . The asshttp://www.maizegdb.org/ (accessed on 23 September 2022)), NCBI (http://www.ncbi.nlm.nih.gov (accessed on 23 September 2022)), and CNKI (http://www.cnki.net (accessed on 23 September 2022)), we collected information on corresponding QTLs and associated markers for multiple insect-resistant traits from our results and previous studies via QTL analysis, GWAS, and association mapping. The hot genetic loci were the overlapping regions combining multiple genetic loci responsible for multiple insect-resistant traits or single genetic loci that explained the large R2 (10%) in 10 Mb physical intervals. Further, the physical map of all hot genetic loci and candidate genes involving multiple insect-resistant traits in these hot genetic loci regions was developed by BioMercator v. 4.2 software (http://www.bioinformatics.org/mqtl/wiki/ (accessed on 28 September 2022)) [https://agbase.arizona.edu/ (accessed on 6 October2022)) [Using public databases, i.e., MaizeGDB (r 2022)) . The funer2022)) .http://www.premierbiosoft.com/ (accessed on 9 October 2022)) , and cDNA was made using a kit , according to the manufacturer\u2019s instructions. RT-PCR was conducted using TransStart Tip Green qPCR SuperMix . The primers for five candidate genes ,64 were r 2022)) . Relativnce gene .http://www.R-project.org/ (accessed on 16 December 2021)) to calculate the BLUP of DIMBOA content values [http://www.ibm.com/products/spss-statistics (accessed on 16 December 2021)). The significance of the total and residual variances of DIMBOA content in 310 inbred lines under both ecological environments was estimated by a GLM for univariate data and by one-way analysis of variance (ANOVA). The broad-sense heritability , and r was the number of replications (r = 5). The CVg [The average DIMBOA content in 310 inbred lines from five biological replicates in each ecological environment were analyzed, respectively. A mixed linear model was fitted using the lmer function in lme4 package of R (t values . These d follows ,66,67:HB The CVg  of DIMBOIn summary, the DIMBOA confers significant resistance to ACB and CLA. In this study, SSR analysis revealed a wide genetic diversity in the 310 tested maize inbred lines from four type regions of China\u2019s Gansu Province, which is the largest maize seed production and breeding area in China. Population structure indicated that 294 inbred lines were successfully assigned to one or another group at a membership probability of \u2265 0.500. DIMBOA performance evaluation screened out 3 high and 15 moderate insect-resistant inbred lines, which can be used as parents in breeding programs to develop new maize varieties with multiple insect resistance. Using linkage mapping, we detected nine significant SSRs associated with DIMBOA content in both environments. We then combined the 47 original genetic loci for 8 multiple insect-resistant traits from previous studies to detect 19 hot loci. Among them, 11 hot loci were located in Bin 1.04, Bin 2.00\u20132.01, Bin 2.03\u20132.04, Bin 4.00\u20134.03, Bin 5.03, Bin 5.05\u20135.07, Bin 8.01\u20138.03, Bin 8.04\u20138.05, Bin 8.06, Bin 9.01, and Bin 10.04 regions, and they supported pleiotropy for their association with two or more insect-resistant traits. Further, the 49 candidate genes involved in DIMBOA biosynthesis, sugar metabolism/homeostasis, and other multiple insect-resistant defense mechanisms in maize were identified in all 19 hot loci, and their highly interconnected network may form complex maize, multiple insect, pest-induced defense mechanisms."}
+{"text": "Correction: BMC Nephrol 23, 348 (2022)https://doi.org/10.1186/s12882-022-02986-2Following publication of the original article , the autThe original article has been corrected.Additional file 1 Table\u00a01. Summary of literature on renal biopsy after intravitreal injection of vascular endothelial growth factor inhibitors."}
+{"text": "Sphaerotilus natans and Pseudomonas aeruginosa strains. Its genome consists of 61,858\u2009bp (64.3% GC) and 89 genes, including 32 with predicted functions. SN1 genome is very similar to Pseudomonas phage M6, which contains hypermodified thymidines. Genome analyses revealed similar base-modifying genes as those found in M6.Phage SN1 infects  S. natans ATCC 13338 as the host  using the host . An earlthe host . Host rand OT684 .S. natans ATCC 13338 in nutrient broth  and agitated at 30\u00b0C (P. aeruginosa PAO1 (HER1153) in TSB/TSA medium at 30\u00b0C using both plaque assays and lysis of liquid cultures. Species identification of the above two host strains was confirmed by 16S sequencing.Here, phage SN1 was amplified with its host  at 30\u00b0C . Cell deS. natans as host) using the phenol-chloroform extraction method (https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi) and blastp (https://blast.ncbi.nlm.nih.gov/Blast.cgi) (Phage genomic DNA was purified from lysate (n method . Libraryn method . Illuminn method . Trimmedn method . Two assn method  in combiast.cgi) , 9 usingast.cgi) . BioinfoPseudomonas phages with 95 to 98% nucleotide identity (73 \u2013 96% query cover). Interestingly, phage SN1 has 96.76% nucleotide identity (91% query cover) with Pseudomonas phage M6 genome, which contains hypermodified thymines . Half oON165687. Raw sequence reads are available under SRA number SRR18758685. Phage SN1 is available at www.phage.ulaval.ca.Genome sequence is available under GenBank number"}
+{"text": "Globularia L. have been used as healing agents for various ailments, with utilization of Globularia alypum L. being most frequently reported. The aim of this study was to evaluate the antidiabetic, antioxidant, anti-inflammatory, antibacterial and anticancer potential of G. alypum and three related species, G. punctata Lapeyr., G. cordifolia L. and G. meridionalis (Podp.) O.Schwarz, in relation to their phytochemical compositions. Globularin and verbascoside were identified using LC-PDA-ESI-MSn as the major metabolites of G. alypum with known biological activities. G. alypum demonstrated the greatest \u03b1-glucosidase inhibitory activity and DPPH radical scavenging activity (IC50 = 17.25 \u03bcg/mL), while its anti-inflammatory activity was not significantly different from those of related species. All investigated species showed considerable antibacterial activity against methicillin-resistant Staphylococcus aureus in the broth microdilution method (MIC = 1.42\u20133.79 mg/mL). G. punctata also showed antibacterial activities against Escherichia coli (MIC = 1.42 mg/mL), Bacillus subtilis (MIC = 1.89 mg/mL), B. cereus (MIC = 2.84 mg/mL) and Enterococcus faecalis (MBC = 5.68 mg/mL). G. punctata, G. cordifolia and G. meridionalis showed greater anticancer potential than G. alypum. Obtained results indicate investigated Globularia species could serve as sources of diverse bioactive molecules, with G. punctata having the greatest antibacterial potential.Species from the genus  According to the World Health Organization (WHO) statistics for 2021, the leading causes of global premature mortality from non-communicable diseases include cancer, cardiovascular diseases, diabetes, and chronic respiratory diseases. In 2019, 33.2 million people died solely from these diseases, which is 28% more deaths caused by the same four diseases than in 2000. Taken individually, there has been a 25% increase in the total global mortality from cardiovascular diseases (17.9 million deaths), a 37% increase in cancer mortality (9.3 million deaths), and a 10% increase in chronic respiratory diseases mortality (4.1 million deaths), while diabetes mortality has grown by 72% \u2212 and 153 [M\u2212H\u2212anhydroglucose]\u2212. In MS2, compound 50 was characterized by product ions at m/z 161 (\u2212330 Da) and 175 (\u2212316 Da). In MS3 of the first product ion, further loss of 28 Da (\u2212CO) was observed. The first loss in MS2 was attributed to simultaneous loss of hydroxytyrosol glucoside and a methyl group and the second to hydroxytyrosol glucoside. Loss of a methyl group was previously observed for the reference standard of ferulic acid \u2212. The compound was tentatively identified as galypumoside C (6\u2032-O-menthiafoloylverbascoside), whose presence in leaves of G. alypum was already reported \u2212. The formed radical aglycone product ion was further subjected to loss of one methyl group (\u221215 Da) in MS3 and major loss of 84 Da in MS4, attributed to the cleavage of three CO \u2212, 209 [M\u2013H\u2013anhydroglucose\u2212benzoylglucose]\u2212, 227 [M\u2013H\u2013anhydroglucose\u2212benzoylanhydroglucose]\u2212 and 371 [M\u2013H\u2013anhydroglucose\u2212benzoic acid]\u2212, indicated a similar structure to previously identified C-4 carboxylated iridoids. Second- and third-order mass spectra of compound 25 were comparable to MS3 and MS4 of compound 21, while major loss of 30 Da (\u2212CH2O) from the MS3 product ion present at m/z 165 was observed in MS4 together with minor fragment ions present at m/z 121 (\u2212C2H4O), 137 (\u2212CO) and 147 (\u2212H2O). These compounds were tentatively identified as 6\u2032-O-benzoyldeacetylasperulosidic acid glucoside and 6\u2032-O-benzoyldeacetylasperulosidic acid, keeping in mind other observed asperuloside-type iridoids and benzoylation of glucose at C-6\u2032 position, which was recorded for dumuloside in G. dumulosa \u2013, 163 [p-coumaric acid\u2013H]\u2013, 293 [M\u2013H\u2013glucose\u2013CO2]\u2013 and 355 [M\u2013H\u2013anhydroglucose]\u2013. Further MS/MS fragmentation pattern obtained in MS3 and MS4 were in accordance with those of asperuloside and besperuloside, which have acetic/benzoic acid attached at C-10 position. This compound, with UV \u03bbmax present at 191, 232 and 315 nm, comparable to those previously observed for asperuloside (191 and 239 nm) and p-coumaric acid  [O-(p-coumaroyl)-deacetylasperuloside.Compound G. alypum methanolic leaf extract indicated not only the previously described high globularin  content [G. alypum were glycosides of 6-hydroxyluteolin, the same as for other investigated species, while vicenin-2 was observed only in G. alypum, as well as the lignan diglucoside liriodendrin. Leaf extract of G. punctata contained high relative amounts of iridoids asperuloside, besperuloside, globularin, deacetylasperuloside, asperulosidic acid, and scandoside, while verbascoside, its isomers  and glycosylated derivatives  were observed as the major phenylethanoids. Solely G. punctata was characterized by acylated derivatives of 6-hydroxyluteolin, as previously reported [G. cordifolia and G. meridionalis were characterized by iridoids asperuloside and monomelittoside and their often benzoylated derivatives, with globularifolin (10-O-benzoylmonomelittoside) as the major compound, as previously reported [O-glucoside and demethoxycentaureidin 6,4\u2032-dimethyl ether were recognized as characteristic flavonoids. Other major constituents observed in investigated Globularia species included mannitol, sucrose, quinic acid  and fatty acid oxidation products that have already been reported [Obtained chromatogram of  content , but als content , do not reported ,40,42. Greported ,26,43. Mreported .Globularia species obtained by Soxhlet extraction was performed using LC-PDA-ESI-MSn -sophoroside, trichosanthoside B, trichosanthoside A, arenarioside, and besperuloside, with the iridoid asperuloside as the major compound. Other more prominent peaks included deacetylasperuloside and 10-O-(p-coumaroyl)-deacetylasperuloside, present also in G. cordifolia and G. meridionalis, as well as gardoside, caffeoylglucoside isomer and globusintenoside isomer present in all four species. For G. cordifolia and G. meridionalis, the major compounds were mannitol, sucrose, asperuloside and several phenylethanoids, including verbascoside, methoxyverbascoside isomer, isoverbascoside, globusintenoside isomer and benzoylrossicaside A isomer, while their distinctive peaks could be attributed to monomelittoside, 6\u2032-O-benzoylmonomelittoside and globularifolin. The major identified flavonoid characteristic for these two species was demethoxycentaureidin 6,4\u2032-dimethyl ether. Besides 6-hydroxyluteolin 7-O-glucoside, apigenin was detected in Soxhlet extracts of all investigated species.Identification of major constituents of methanolic aerial parts extracts of investigated -ESI-MSn , Table 4-ESI-MSn  or thoseG. alypum  using two different mobile phases . Blue zone of besperuloside, present only in G. punctata, was not clearly visible due to its overlapping with the brown zone of globularin. Shared yellow zones may be attributed to phenylethanoids present in all investigated species with verbascoside as the major shared component, while additional yellow zones observed for G. alypum and G. punctata correspond to the presence of their major unique phenylethanoids  that were detected using LC-MS  and glutathione peroxidase (GPx)) and two non-enzymatic biomarkers of oxidative stress (free thiol groups (-SH) and reduced glutathione (GSH)) in human hepatocellular carcinoma Hep G2 cells exposed to hyperglycemic conditions. Cell viability was evaluated using lactate dehydrogenase (LDH) and 3--2,5-diphenyltetrazolium bromide (MTT) assay. Antioxidant potential of aerial parts extracts was evaluated based on spectrophotometric measurement of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity and by performing thin layer chromatography (TLC) bioautography using the same free radical as spray reagent.p < 0.05), with G. alypum extracts being most effective (30.0\u201345.7% inhibition)  and reducing diabetic complications. This can partially be achieved by inhibiting the activity of \u03b1-glucosidase, the key enzyme responsible for hydrolytic cleavage of complex carbohydrates. Inhibition of \u03b1-glucosidase may retard the absorption of glucose and decrease postprandial blood glucose levels and, therefore, it is one of the key targets for treating type 2 diabetes mellitus (T2DM) . All fouibition) . This su50 = 0.52 mg/mL) and \u03b1-amylase (IC50 = 0.57 mg/mL) was reported for diethyl ether fraction of G. alypum aerial parts extract [G. alypum in the present and in related studies [Inhibition of \u03b1-glucosidase  (r = \u22120.791), was previously reported for G. trichosantha and G. orientalis [Clerodendrum bungei Stud., the phenylethanoids verbascoside (IC50 = 0.5 mM), leucosceptoside A (IC50 = 0.7 mM) and isoverbascoside (IC50 = 0.1 mM), which were found in all investigated Globularia species, exhibited stronger anti-\u03b1-glucosidase activities than the positive control (acarbose) [50 = 0.15 mM) [Globularia species. Hereby, many acylated phenylethanoids were only found in G. alypum, including galypumoside B, calceolarioside A, and calceolarioside B, of which the latter was recently identified also as a pan inhibitor of SARS-CoV-2 proteins [G. alypum, such as verminoside (caffeic acid) and specioside (p-coumaric acid). However, greater inhibition was observed for leaf extracts of G. alypum and G. meridionalis  . Inhibitientalis . In a stcarbose) . The obscoside IC = 0.5 mMproteins . Phenolidionalis , which cdionalis .-O-glucoside). In Origanum majorana L. leaves, 6-hydroxyapigenin (scutellarein) (IC50 = 12 \u03bcM) and 6-hydroxyluteolin (IC50 = 10 \u03bcM) were recognized as the most potent inhibitors of \u03b1-glucosidase, while weaker inhibitions were observed for 6-hydroxyluteolin 7-O-glucoside (IC50 = 300 \u03bcM) and flavones lacking the 6-hydroxyl substituent, apigenin and luteolin (IC50 > 500 \u03bcM) [G. alypum extract compared to those of extracts of related species may also be explained by presence of additional \u03b1-glucosidase inhibitors, such as vicenin-2 (IC50 = 270.53 \u03bcM) [The observed inhibitory effect of all four species may in part be connected to the presence of 6-hydroxyflavones  . The rec0.53 \u03bcM) .G. alypum [G. alypum as an antidiabetic could include prevention of diabetic complications through direct or indirect antioxidant activity, including radical scavenging activity [G. alypum methanolic leaf extract reduced glycemia and glycosylated hemoglobin levels, and improved the redox status, especially in the liver [The liver plays a vital role in blood glucose level regulation both in physiological and pathological states. In DM, liver is among the primary organs susceptible to hyperglycemia-induced oxidative stress, which may result in irreversible oxidative modifications of its macromolecules that may lead to abnormal glycogen deposition, non-alcoholic fatty liver disease, fibrosis, cirrhosis, hepatocellular carcinoma, and other liver abnormalities . Besides. alypum ,11,56, w. alypum , additioactivity ,57, enhaactivity ,59. For Globularia leaf extracts on biomarkers of oxidative stress have been evaluated in Hep G2 cells cultured under hyperglycemic conditions (p < 0.05). Increased GSH content was observed for G. alypum (+18.6%), G. punctata (+47.4%), and G. cordifolia extracts (+68.1%) at 1.0 mg/mL concentration and G. cordifolia extract at 0.5 mg/mL (+11.7%) (p < 0.05). All tested samples significantly increased free thiol groups content (+14.0\u201373.7%) and cell viability in the LDH assay (+22.2\u201376.7%) (p < 0.05), while in the MTT assay, a 21.6\u201325.4% increase was observed only for G. punctata (both concentrations) and G. cordifolia (c = 1.0 mg/mL) (p < 0.05). Overall, G. cordifolia reduced the pro-oxidant effects of hyperglycemic conditions observed in Hep G2 cells in all performed assays. Its favorable effect on oxidative status  has already been recorded in human keratinocytes [In the present study, effects of two different concentrations (0.5 and 1.0 mg/mL) of nditions . Treatmeinocytes .G. cordifolia was characterized by the highest condensed tannin content , and very good positive correlation between GPx activity and condensed tannin content    . Condens < 0.05) . The majpunctata , Table 3. alypum , was aspflavones , many ofr = 0.88, p < 0.01) as well as total phenolic content   than its phenylethanoids (IC50 =11.8\u201315.5 \u00b5M) and iridoids (IC50 = 28.2\u201376.0 \u00b5M), as well as than quercetin (IC50 = 7.8 \u00b5M) and butylated hydroxytoluene (IC50 = 30.0 \u00b5M) [Very good positive correlation was found between cell viability assessed by the LDH assay and flavonoid content ( < 0.05) . Accordiactivity ,35,64,65punctata . The latpunctata , contain30.0 \u00b5M) . In the G. alypum [Globularia [50 values ranged from 17.25 \u03bcg/mL  to 24.19 \u03bcg/mL (G. punctata). The obtained results are in accordance with our previous study, in which relatively higher antioxidant activities of both G. alypum leaf and flower extracts and relatively lower antioxidant activity of G. punctata flower extract were observed [50 value for G. alypum is comparable to previous reports for the diethyl ether fraction of its aerial parts (IC50 = 20.54 \u03bcg/mL) [50 = 15.58\u201327.54 \u03bcg/mL [50 = 25.65 \u03bcg/mL) and stems (IC50 = 22.11 \u03bcg/mL) [50 = 23.50 \u03bcg/mL) [50 value established for G. meridionalis is in accordance with that previously reported for methanolic extract of its aerial parts obtained by maceration (IC50 = 21.00 \u03bcg/mL), from whose methanolic fraction verbascoside (acteoside), isoverbascoside (isoacteoside) and apigenin 7-O-glucoside were isolated [50 values for DPPH radical scavenging activity and the estimated total phenolic content of aerial parts extracts  (50 = 22.9 \u00b5M) and calceolarioside B (desrhamnosyl isoacteoside) (IC50 = 26.2 \u00b5M) [G. alypum.Direct antioxidant activity of studied species was confirmed by the DPPH assay, which is the most frequently used assay for evaluation of antioxidant activity of . alypum ,67,68,69obularia ,30, to eobserved . Obtaine4 \u03bcg/mL) , methano54 \u03bcg/mL , IC50 = isolated . Excelle < 0.05) , which i26.2 \u00b5M) , found oGlobularia species\u2019 constituents [Globularia [G. alypum [50 = 58.1 \u00b5M) than ascorbic acid [Globularia species [G. punctata showed the lowest radical scavenging activity, the second method revealed at least three zones characteristic for this species showing prominent antiradical activity . Their Rf values matched those of three orange, fluorescent zones observed at 365 nm after NP/PEG treatment, presumably of flavone origin [G. punctata (Globularia elongata Hegetschw. (syn. G. punctata) [-O-sophoroside, 6-hydroxyluteolin 7-O-(6\u2032\u2032\u2032-O-caffeoyl)-sophoroside, and 6-hydroxyluteolin 7-O-(6\u2032\u2032\u2032-O-(p-coumaroyl))-sophoroside.To identify the constituents responsible for observed activity, TLC bioautography with DPPH used as spray reagent was performed as in earlier studies of antioxidant activity of different tituents ,65,67. Bobularia , as well. alypum ,67, a mo. alypum . Two dombic acid . The for species ,39,40,64e origin . Based opunctata  and lite2, PGD2, PGF2\u03b1, and PGI2) and thromboxane A2 [2 (cyclooxygenase site), which is afterwards reduced to PGH2 (peroxidase site) [G. alypum, G. punctata, G. cordifolia and G. meridionalis methanolic extract of aerial parts obtained by Soxhlet extraction was evaluated spectrophotometrically using two methods, one based on inhibition of peroxidase activity of COX-1 using N,N,N\u2032,N\u2032-tetramethyl-p-phenylenediamine dihydrochloride (TMPD assay) and the other based on inhibition of its cyclooxygenase activity assessed by the prostaglandin E2 assay (PGE2 assay). All tested species inhibited COX-1 activity  to 51.3%  in the TMPD assay (IC50 = 2.90 \u03bcM for indomethacin), and from 25.7%  to 40.6%  in the PGE2 assay (IC50 = 1.03 \u03bcM for indomethacin). The results obtained for G. alypum are comparable to those reported for methanolic leaf (5.33%) and flower extract (61.05%) of the same species tested at 33 \u03bcg/mL concentration using the TMPD assay [Cyclooxygenase isoenzymes COX-1 and COX-2 are first in the cascade of enzymes responsible for metabolism of arachidonic acid to prostaglandins (PGEoxane A2 . Due to se site) . Althougse site) . Anti-inactivity . ExtractPD assay .50 > 1 mM), one of the major metabolites found in all investigated species, was lower than that against COX-2 (IC50 = 0.69 mM) [Anisomeles indica (L.) Kuntze, of which verbascoside, isoverbascoside, calceolarioside A and apigenin were also recorded in investigated Globularia extracts, the latter compound gave the lowest docking score with COX-1 (\u22126.558) and COX-2 receptors (\u22128.441) in molecular docking analysis, which indicated its strong binding to the enzymes\u2019 active sites [G. alypum, for which the tendency of greater COX-1 inhibition was observed. Anti-inflammatory potential of investigated species could also be connected to their possible diminishing effects on other pro-inflammatory mediators , and cytokines such as interleukin-1\u03b2 (IL-1\u03b2), interleukin-6 (IL-6) and tumor necrosis factor-\u03b1 (TNF-\u03b1) [No correlation between the obtained results and assessed total phenolic or flavonoid content was found . Inhibit0.69 mM) . Among tve sites . Based ove sites , apigeni (TNF-\u03b1) ,79,80,81G. alypum and G. punctata, was recently reported to protect against glucose-induced podocyte injury by ameliorating apoptosis and inflammation through reduction of TNF-\u03b1, IL-6 and IL-1\u03b2 at 1\u201310 \u03bcM concentrations, which would suggest its preventive potential against diabetic nephropathy [G. punctata, G. cordifolia and G. meridionalis, significantly decreased the production of NO, PGE2, TNF-\u03b1, and IL-6, and inhibited the expression of inducible NO synthase (iNOS), COX-2, TNF-\u03b1, and IL-6 mRNA in LPS-induced RAW 264.7 cells [-O-benzoyl-monomelittoside), the major metabolite of G. cordifolia and G. meridionalis, was reported to significantly suppress expression of nuclear factor-\u03baB (NF-\u03baB) in LPS-stimulated cells at 200 \u03bcM concentration [G. alypum. The same compound, together with other above-mentioned compounds , as well as other major compounds identified in this study , were also reported for G. trichosantha [Catalpol, the iridoid present together with its many esters and/or derivatives (including globularin) in hropathy . The samhropathy . Similarhropathy , as wellhropathy . Asperul.7 cells . On the ntration , a lignahosantha ,64,83, whosantha , which mStaphylococcus aureus, a Gram-positive bacterium, is the main causative pathogen in diabetic foot infections, with methicillin-resistant S. aureus (MRSA) being the major multidrug resistant bacterium. Other pathogens may include Gram-positive bacteria, such as Streptococcus spp. and Enterococcus faecalis, and Gram-negative bacteria, such as Pseudomonas aeruginosa, Escherichia coli, Proteus spp., Enterobacter spp., and Klebsiella spp. [G. alypum, G. punctata, G. cordifolia and G. meridionalis was evaluated by testing the effect of methanolic extracts of aerial parts obtained by Soxhlet extraction against four Gram-positive  and three Gram-negative bacterial strains  using two complementary methods; well diffusion and serial broth microdilution method. In the first method, all four species showed notable inhibitory activity against S. aureus, with the greatest zone of inhibition recorded for G. alypum (25.0 mm), as well as low inhibitory activity against B. cereus  and 2.84 mg/mL . Low inhibitory activity of G. alypum against P. aeruginosa (MIC = 22.73 mg/mL) and moderate to good bactericidal/inhibitory activity of G. punctata against E. faecalis (MBC = 5.68 mg/mL) and E. coli (MIC = 1.42 mg/mL) were also confirmed. Comparable antibacterial activity of G. alypum methanolic extract against S. aureus (MIC = 2\u20134 mg/mL) was reported by previous studies [P. aeruginosa (MIC = 8 mg/mL) [The results of the serial broth microdilution method followed by sub-cultivation on agar plates mainly coincided with those of the first method . Assesse MBC = 5. mg/mL an studies , while g8 mg/mL) .G. punctata showed good antibacterial activity against B. cereus (MIC = 2.84 mg/mL) and B. subtilis (MIC = 1.89 mg/mL), while much lower inhibition against B. cereus was observed for related species (MIC = 11.36 mg/mL). The latter method possibly accounted for the antibacterial activity of less hydrophilic compounds of G. punctata, which might not have been observed earlier due to their lower solubility and consequential lower diffusion into the hydrophilic (agar) medium [r = \u22120.998, p < 0.01) was found between MIC values against B. cereus and flavonoid content , were equal to 8, 8 and 64\u2013400 \u03bcg/mL, respectively, while against S. aureus no inhibitory activity was observed [Opposite to the results of the diffusion method, in the second method, ) medium . Excelle content , indicatlucoside   and five clinical isolates of MRSA, were also tested (p > 0.05). Keeping in mind that these are the most common causative pathogens found in diabetic foot infections [G. alypum in the treatment of diabetes-associated foot ulcers. The anti-staphylococcal activity could partly be attributed to verbascoside, for which the assessed inhibitory activity (MIC = 227.2 \u03bcg/mL) corresponded to values previously reported against five MRSA strains (MIC = 64\u2013256 \u03bcg/mL) [S. aureus of all investigated species may also indicate contribution of other common constituents, such as isoverbascoside, rossicaside A, forsythoside A and 6-hydroxyluteolin 7-O-glucoside. Verbascoside (MIC = 60 \u03bcg/mL) was observed to have greater anti-staphylococcal activity than isoverbascoside (MIC = 130 \u03bcg/mL), while both compounds possessed lower inhibitory activities against E. faecalis (MIC = 100\u2013150 \u03bcg/mL), E. coli (MIC = 100\u2013250 \u03bcg/mL) and P. aeruginosa (MIC = 250 \u03bcg/mL) [G. alypum, also possess antibacterial potential against Gram-positive and Gram-negative bacteria [G. alypum against P. aeruginosa. The antibacterial potential of G. punctata against E. coli and E. faecalis, as well as S. aureus, may be connected to its characteristic phenylethanoids  and/or flavonoids. Excellent negative correlations were found between flavonoid content and MICs against MSSA MFBF 505  and MRSA MFBF 154 .Nine additional o tested . Observefections , the obs6 \u03bcg/mL) . Compara0 \u03bcg/mL) . Other pbacteria , which mGlobularia species was evaluated using the MTT assay based on their effects against MDA-MB-231 breast cancer cell line and A1235 glioblastoma cell line. Cytotoxic effect against MDA-MB-231 cell line was observed for extracts of G. punctata (34.48%), G. cordifolia (63.41%) and G. meridionalis (82.96%) (at the highest concentration used), while cytotoxic effect against A1235 cell line was observed for all investigated species (p < 0.05)  and the lowest for G. alypum (IC50 = 231.43 \u03bcg/mL)  . Greates3 \u03bcg/mL) .G. meridionalis, G. cordifolia and G. punctata may be connected to asperuloside and its derivatives (Oldenlandia diffusa (Willd.) Roxb. that contained asperuloside and deacetylasperulosidic acid also reduced MDA-MB-231 and MDA-MB-453 cell viability and suppressed their colony formation capacities [50 = 0.7 \u03bcg/mL), HL60 cells (IC50 = 11.0 \u03bcg/mL) and KB cells (IC50 = 104.2 \u03bcg/mL) was recorded for asperuloside [50 = 6.1 \u03bcg/mL) [50 = 10 \u03bcM) was reported [G. cordifolia and G. meridionalis. Very good negative correlations were established between observed cell viability and concentrations of flavonoids , iridoids , and total phenolics , while excellent correlation was found between MDA-MB-231 cell viability and concentration of condensed tannins  (50 = 0.159\u20130.258 \u03bcM) [The observed cytotoxic effects against MDA-MB-231 cells that were shown by ivatives  [102,103s IC50 = .2 \u03bcg/mL reported , could b< 0.001) . Cytotox.258 \u03bcM)  and a ra.258 \u03bcM) , the majCallicarpa nudiflora Hook. and Arn. against HeLa cells, A549 lung adenocarcinoma cells and MCF-7 cells resulted in isolation of flavonoids, phenylethanoids and iridoids, which included those found in investigated species, 6-hydroxyluteolin 7-O-glucoside, verbascoside and catalpol [50 = 1530 \u03bcg/mL) was also reported for G. alypum [Moreover, investigation of cytotoxic activity of catalpol . Weak cy. alypum .r = \u22120.95, p < 0.001), flavonoids  and condensed tannins , while very good negative correlation was observed between A1235 cell viability and iridoid concentrations   . Common thanoids ,111. Thi87 cells .2EDTA), 5,5\u2032-dithiobis(2-nitrobenzoic acid) , Dulbecco\u2019s modified Eagle\u2019s medium (DMEM), Type I \u03b1-glucosidase (isolated from Saccharomyces cerevisiae), glutathione (GSH), hematin, 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES), p-nitrophenyl-\u03b1-d-glucopyranoside (PNG), quercetin, N,N,N\u2032,N\u2032-tetramethyl-p-phenylenediamine dihydrochloride (TMPD), trichloroacetic acid, 2,3,5-triphenyltetrazolium chloride (TTC), trypsin-EDTA and vanillin from Sigma-Aldrich, St. Louis, MO, USA; and sodium diethyldithiocarbamate trihydrate (DDC) from VWR International, Radnor, PA, USA.Adrenalin bitartrate and indomethacin were purchased from Acros Organics, Geel, Belgium; glacial acetic acid from Alkaloid, Skopje, North Macedonia; asperuloside and catalpol from Carl Roth, Karlsruhe, Germany; hydrochloric acid from Carlo Erba, Emmendingen, Germany; arachidonic acid from Cayman Chemical Company, Ann Arbor, MI, USA; (+)-catehin, 2,2-diphenyl-1-picrylhydrazyl (DPPH), diphenylboric acid-\u03b2-ethylamino ester  and LC-MS grade formic acid from Fluka, Buchs, Schwitzerland; fetal bovine serum (FBS), minimum essential medium (MEM), penicillin-streptomycin, and trypan blue from Gibco, Gaithersburg, MD, USA; absolute ethanol from Gram-Mol, Zagreb, Croatia; verbascoside from HWI Analytik, R\u00fclzheim, Germany; ethyl acetate, Folin-Ciocalteu\u2019s reagent, D-(+)-glucose, polyethylene glycol 4000 (PEG 4000), potassium chloride, potassium phosphate monobasic, sodium carbonate decahydrate, sodium hydroxide, sodium phosphate dibasic heptahydrate, sulfuric acid and Tween from Kemika, Zagreb, Croatia; gentamicin sulfate and norfloxacin from Krka, Novo mesto, Slovenia; chloroform, copper(II) sulfate pentahydrate, gallic acid, LC-MS grade acetonitrile, methanol, M\u00fcller\u2013Hinton agar, M\u00fcller\u2013Hinton broth, sodium chloride and Tryptic soy agar (TSA) from Merck, Darmstadt, Germany; Tris-HCl buffer from Santa Cruz Biotechnology, Dallas, TX, USA; formic acid from Scharlau, Scharlab, Barcelona, Spain; acarbose, aluminum chloride hexahydrate, aucubin, 1-chloro-2,4-dinitrobenzene (CDNB), cyclooxygenase-1 (COX-1) from sheep, dimethyl sulfoxide (DMSO), 3--2,5-diphenyltetrazolium bromide (MTT), disodium ethylenediamine-tetraacetic acid , Grobnik field , Ba\u0161ke O\u0161tarije, Velebit  and Alan, Velebit . Plant material was identified by Prof. Kroata Hazler Pilepi\u0107. Voucher specimens are deposited in the Herbarium of the Department of Pharmaceutical Botany of the Faculty of Pharmacy and Biochemistry, University of Zagreb. Collection data are provided in Aerial parts of investigated v/v) methanol and extracted three times with chloroform. After chloroform removal, the remaining solvents were again evaporated at 30 \u00b0C and freeze-dried. All samples were stored at \u221220 \u00b0C until further use.Ultrasound-assisted extraction of powdered leaves (0.5\u20131.25 g) was performed using methanol (1:10) for 2 \u00d7 30 min. Obtained liquid extracts were filtered and evaporated to dryness using a rotavapor at 50 \u00b0C. Soxhlet extraction of powdered aerial parts (145\u2013280 g) was performed using methanol (1:5) for 8 h. Obtained extracts were evaporated to dryness using a rotavapor at 30 \u00b0C, re-suspended in 50% , was used for the analysis of methanolic leaf and aerial parts extracts composition. Zorbax SB-C18 column   was used as a stationary phase. Separation of constituents and data collection were carried out under the previously described conditions [m/z 50\u20132000. Electrospray ionization (ESI) was performed in negative ionization mode. MSn spectra (MS2\u2013MS4) were obtained by collision induced dissociation (CID) of the ion of the greatest intensity in the mass spectrum of lower order with helium as the collision gas and normalized collision energy set at 35%. Data acquisition and processing were performed using Thermo Xcalibur 2.2 . Compound identification was based on the comparison of chromatographic and spectral data to those of previously identified constituents found in methanolic extracts of aerial parts of the same Globularia species obtained by heating under reflux conditions [High-performance liquid chromatography-photodiode array detection-electrospray ionization-tandem mass spectrometry, HPLC-PDA-ESI-MSnditions . UV/Vis nditions .254 aluminum plates , 10 \u00d7 20 cm, thickness 0.20 mm, using chloroform-methanol-water = 61:32:7 (v/v/v), or ethyl acetate-methanol-water = 20:2:1 (v/v/v) as the mobile phase [w/v) ethanolic vanillin-5% (v/v) ethanolic sulfuric acid reagent (5\u201310 min at 100\u2013105 \u00b0C), to those of asperuloside, aucubin and catalpol reference standards (c = 0.5 mg/mL).Constituents of methanolic aerial parts extracts obtained by Soxhlet extraction (c = 50 mg/mL) were separated on thin-layer chromatography (TLC) silica gel 60 Fle phase . Sample v/v) DMSO were mixed with 50 \u00b5L Type I \u03b1-glucosidase from Saccharomyces cerevisiae  and preincubated at 37 \u00b0C. After 10 min, 50 \u00b5L PNG  was added. Absorbance was measured for 5 min at 405 nm against a blank in which PNG was replaced with phosphate buffer. Results are expressed as enzyme activity % in comparison to control (2% (v/v) DMSO), which was considered to give 100% enzyme activity. Acarbose was used as positive control.Inhibition of \u03b1-glucosidase activity was assessed using PNG as previously described . Methanov/v) FBS, 20 IU/mL penicillin and 20 \u00b5g/mL streptomycin at 37 \u00b0C and 5% CO2. After removal of culture medium, cells were washed with phosphate buffered saline (PBS), trypsinized using 0.25% trypsin-EDTA solution for 5 min at 37 \u00b0C, counted under a light microscope (Leitz Diavert) after 0.04% trypan blue staining using the B\u00fcrker\u2013T\u00fcrk counting chamber and seeded into six-well plates (1.67 \u00d7 106) with FBS-free medium or 96-well plates (2 \u00d7 105) (MTT assay). After 24 h incubation at 37 \u00b0C, cells were washed with PBS and incubated with methanolic leaf extract solutions (c = 0.5 and 1.0 mg/mL) in MEM with 20 mM glucose (hyperglycemic conditions) for 24 h at 37 \u00b0C. Normal control cells were kept in MEM with 5.56 mM glucose. Prior to cell treatment, samples were filtered using sterile Nalgene filter units of pore size 0.2 \u03bcm . After incubation and washing with PBS, the cells were lysed with 1% (v/v) Tween in PBS with the help of ultrasound (4 W) (Cole-Parmer 4710) for 15 s and centrifuged for 20 min at 14,000 rpm at 4 \u00b0C. The supernatant was stored at \u221280 \u00b0C until further use.Hep G2 cells obtained from European Collection of Authenticated Cell Cultures  were cultured in MEM supplemented with 10% (\u22121M\u22121). Content of free thiol groups (-SH) was evaluated spectrophotometrically using Ellman\u2019s reagent [\u22121M\u22121). For evaluation of GSH content, as previously described [v/v) trichloroacetic acid was added (100 \u00b5L), and the mixture was centrifuged . To obtained supernatant (100 \u00b5L), PBS  and DTNB (50 \u00b5L) dissolved in the same buffer were added. The production of yellow colored 5-thio-2-nitrobenzoic acid (TNB) was measured at 405 nm against reagent blank. Results were calculated using the molar extinction coefficient of the product . All results are expressed per mg protein. Protein concentrations were established fluorometrically using the Qubit Protein Assay Kit (Invitrogen).Glutathione peroxidase (GPx) activity was measured spectrophotometrically using the Glutathione Peroxidase Activity Colorimetric Assay Kit . Glutathione S-transferase (GST) activity was evaluated spectrophotometrically using CDNB as the substrate . The cel reagent . Reactioescribed , to depr\u22121M\u22121). Cell viability was also evaluated based on the assessment of their metabolic/mitochondrial function using the MTT assay as previously described [Viability of Hep G2 cells was evaluated spectrophotometrically using the cell lysate supernatant by measuring lactate dehydrogenase (LDH) activity as an indicator of cell membrane damage  with comescribed . After 250).Antiradical activity was assessed using the free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH) according to the previously described procedure . Methano254 glass plates (Merck), 20 \u00d7 20 cm, thickness 0.25 mm, using ethyl acetate-formic acid-glacial acetic acid-water  as the mobile phase [w/v) methanolic solution of DPPH [w/v) methanolic diphenylboric acid-\u03b2-ethylamino ester, and 5% (w/v) ethanolic polyethylene glycol 4000 (NP/PEG reagent) [Constituents of methanolic aerial parts extracts obtained by Soxhlet extraction (c = 20 mg/mL) were separated on TLC silica gel 60 Fle phase . Sample  of DPPH . Constitreagent) , to thatv/v) Tween and 300 \u03bcM DDC (c = 200 U/mL) and then in 0.1 M Tris-HCl buffer, pH 8.0 (1:100) just before the performed assay, while hematin (c = 1 mM) was dissolved in 0.01 M NaOH and afterwards diluted in 0.1 M Tris-HCl buffer, pH 8.0 (1:10). Sample  was preincubated with 0.1 M Tris-HCl buffer, pH 8.0 (20 \u03bcL), 72 mM adrenalin bitartrate (50 \u03bcL), 2 U/mL COX-1 (100 \u03bcL) and 100 \u03bcM hematin (10 \u03bcL) for 5 min at room temperature. The reaction was started by the addition of 100 \u03bcM arachidonic acid (10 \u03bcL). After 20 min incubation at 37 \u00b0C, the reaction was terminated by the addition of 10% (v/v) formic acid (10 \u03bcL). Concentration of produced PGE2 was evaluated using the Prostaglandin E2 EIA Kit-Monoclonal . After 18 h sample incubation with PGE2 tracer (PGE2-acethylcholinesterase conjugate) and PGE2 monoclonal antibody at 4 \u00b0C, wells were emptied and rinsed five times with wash buffer. After 90 min incubation with Ellman\u2019s reagent, absorbance was read at 405 nm using the iEMS Reader MF type 1401  and corrected for the absorbance at 620 nm. Blank absorbance (Ellman\u2019s reagent) and non-specific binding absorbance (absence of antibody) were subtracted from the readings. From absorbances obtained for samples and control (ethanol), PGE2 tracer binding % were calculated, which were inversely proportional to the PGE2 concentrations in the wells. Results are expressed as COX-1 inhibition %, calculated from established PGE2 concentrations. Indomethacin was used as positive control.Cyclooxygenase activity of COX-1 was evaluated spectrophotometrically according to previously described protocols ,119. COXPeroxidase activity of COX-1 was evaluated spectrophotometrically using the TMPD assay , with soBacillus cereus American Type Culture Collection  11778, B. subtilis ATCC 6633, Enterococcus faecalis ATCC 29212, Staphylococcus aureus ATCC 6538 and S. aureus ATCC 29213 among Gram-positive and Escherichia coli ATCC 10536, Klebsiella pneumoniae MFBF 10402 and Pseudomonas aeruginosa ATCC 27853 among Gram-negative bacterial species. Additionally, eight clinical isolates of S. aureus, including methicillin-resistant S. aureus  and methicillin susceptible S. aureus  were used. Bacterial strains were sourced from the Collection of Microorganisms, Department of Microbiology, Faculty of Pharmacy and Biochemistry, University of Zagreb. Inoculums were prepared from overnight cultures that were cultured on tryptic soy agar (TSA) at 37 \u00b0C, suspended in sterile physiological saline, with the optic density adjusted to 0.5 McFarland (1.5 \u00d7 108 colony-forming units (CFUs)/mL) using a densitometer . Right before cultivation in nutrient medium, 1 mL of each prepared suspension was dissolved with 9 mL physiological saline. Prior to cell treatment, sample stock solutions in distilled water (c = 50 mg/mL) were filtered through Chromafil cellulose acetate filters of pore size 0.22 \u03bcm .Antibacterial activity was evaluated using the following strains: d = 6 mm) and filled with 50 \u03bcL of methanolic aerial parts extract solution (c = 50 mg/mL) in distilled water. Plates were incubated at 37 \u00b0C for 18 h in the dark. Diameters of the zones of inhibition of bacterial growth (mm) were read from the diameters of transparent zones around the wells. Gentamicin and norfloxacin (c = 0.2 mg/mL) were used as positive controls.Well diffusion method was carried out according to the European Pharmacopoeia . Inoculuw/v) TTC (20 \u03bcL), which in the presence of metabolically active bacteria  gave red coloration/precipitate [Broth microdilution method was conducted in sterile plastic microtiter plates with 96 wells , in accordance with the Clinical and Laboratory Standards Institute (CLSI) recommendations . Two-folcipitate . This wa2, as previously described [3). Cells were incubated with methanolic leaf extract solutions in DMEM  for 24 h at 37 \u00b0C. Prior to cell treatment, samples were filtered through sterile filters of pore size 0.22 \u03bcm.Human MDA MB-231 breast cancer cells obtained from Dr. Sonja Levanat and human A1235 glioblastoma cells obtained from S.A. Aaronson  were culescribed . After rCell viability was assessed using the MTT assay as previously described . Prior t\u03b1-glucosidase activity, COX-1 inhibitory activity (PGE2 assay) and MDA-MB-231 and A1235 cell viability were performed in quadruplicate, and viability of Hep G2 cells in hyperglycemic conditions (MTT assay) in octuplicate, and the results are expressed as mean values and standard deviations. Concentrations that were observed to reduce the DPPH radical absorbance by 50% (IC50) were estimated using linear regression and those that reduced COX-1 activity and cell viability by 50% (IC50) using logarithmic regression.Phytochemical content, assessment of oxidative stress biomarkers and viability of Hep G2 cells in hyperglycemic conditions (LDH assay), antibacterial activity and COX-1 inhibitory activity (TMPD assay) were evaluated in triplicate and the results are expressed as mean values and standard deviations or, exceptionally, mean values . Assessment of r) was used to establish the relationship between observed biological activities and phytochemical content. In all tests, significance level \u03b1 was set at 0.05. Analyses were performed using GraphPad Prism 6.01 .Statistically significant differences were evaluated by using one-way analysis of variance (ANOVA), followed by Tukey\u2019s post hoc test (comparison between different species) or Dunnett\u2019s post hoc test (comparison between samples and control/hyperglycemia). Pearson\u2019s correlation coefficient (G. alypum and three related species, G. punctata, G. cordifolia, and G. meridionalis, considering different methods of extract preparation and different plant parts used. The bioactive compounds contained in investigated extracts of G. alypum and their observed biological activities are in accordance with the results of previous biological activity studies, as well as the reported traditional uses of this well-investigated medicinal plant, including its antidiabetic use, while those of related species suggest they too may have therapeutic potential. Observed antioxidant, anti-inflammatory, and antimicrobial activities of aerial parts extracts of investigated Globularia species support their use in cosmetics, while cytotoxicity assay results indicate further studies should preferably be carried out on G. cordifolia, G. meridionalis and G. punctata. The latter species also showed greater antimicrobial potential in comparison to G. alypum, which may be associated with its characteristic phenylethanoids and flavonoids. The paper displays how a combination of phytochemical and biological activity data obtained for extracts of several different species of the same genus may improve the understanding of their potential health benefits and facilitate the identification of compounds that are of possible interest for future biological activity studies. Future studies could focus more on the biological activities of major metabolites found in these species and elucidation of their underlying molecular mechanisms.The present study provides a greater insight into the phytochemical composition of"}
+{"text": "Proteus mirabilis to imipenem allows this pathogen evade surveillance for carbapenemases. Metallo-beta-lactamases of the VIM family are widespread among Enterobacterales in Europe. We identified an outbreak of P. mirabilis carrying 2 VIM enzymes among elderly patients from a medical center in Hungary.Nonsusceptibility of P. mirabilis isolates were received from Hungary during 2020 as part of the SENTRY Antimicrobial Surveillance Program. Isolates were susceptibility tested by the CLSI reference broth microdilution method. Carbapenem-nonsusceptible isolates were submitted to whole genome sequencing, screened for resistance mechanisms, and evaluated for core genome multi-locus sequence typing (cgMLST).A total of 16 P. mirabilis isolates from Hungary, 5 carbapenem-nonsusceptible, multidrug-resistant isolates (3 from urinary tract infections and 1 each from bloodstream infection and pneumonia) were identified from patients 66-92 years old. Four of these 5 isolates were listed as community acquired. All isolates were resistant to ceftriaxone , cefepime (16- > 32), imipenem ( > 8), gentamicin ( > 16), levofloxacin (16- > 32), nitrofurantoin ( > 64), trimethoprim/sulfamethoxazole ( > 4), and plazomicin ( > 128), but susceptible to meropenem (0.25-0.5) and ertapenem (0.03-0.25). Isolates carried blaVIM-4 and blaVIM-75 on a class 1 integron within IS26 and separated by aac(6\u2019)-IIc. IS26 was located on a compound plasmid carrying other resistance-encoding genes, including armA. The integron structure is identical to the blaVIM-4 and blaVIM-1-carrying integron described from a Vibrio cholerae isolated from a seagull in France in 2015 (KR262557). blaVIM-75 is a single amino acid variant (Q60R) of blaVIM-1. All isolates carried mutations in the QRDR . Based on cgMLST analysis, these 5 P. mirabilis isolates were highly similar, with only 7-19 SNPs detected.Among the 16 P. mirabilis isolates carrying blaVIM-4 and blaVIM-75 in a Hungarian medical center is a cause of concern. Surveillance should be performed to understand the spread of and treatment options for infections caused by carbapenem-nonsusceptible P. mirabilis.The outbreak of multidrug-resistant Lalitagauri M. Deshpande, PhD, Melinta: Grant/Research Support|Pfizer: Grant/Research Support Katalin Buri\u00e1n, MD, JMI Laboratories: Grant/Research Support Ilona D\u00f3czi Cs\u00e1nyi, MD, JMI Laboratories: Grant/Research Support Mariana Castanheira, PhD, AbbVie: Grant/Research Support|Cidara: Grant/Research Support|GSK: Grant/Research Support|Melinta: Grant/Research Support|Pfizer: Grant/Research Support|Shionogi: Grant/Research Support."}
+{"text": "Correction: BMC Med 20, 398 (2022)https://doi.org/10.1186/s12916-022-02595-8The labels of the two groups \u2013 response vs. non-response \u2013 were mistakenly switched.The HR in panel A was incorrectly stated as 0.38 instead of 0.308.The original article  containeThe figure has been updated in the original article to reflect these amendments."}
+{"text": "Stenotrophomonas maltophilia is a multidrug-resistant nosocomial pathogen that can cause life-threatening infections among immunocompromised populations. This report presents the complete 74,962-bp genome of S. maltophilia podophage Paxi, an N4-like phage sharing 85.3% nucleotide similarity to S. maltophilia podophage Pokken. Stenotrophomonas maltophilia is an environmentally ubiquitous and commensal bacterium, and it has also emerged as a nosocomial pathogen capable of causing life-threatening infections, especially among immunocompromised individuals  at 30\u00b0C, and phage propagation was performed using the soft agar overlay method (www.bioinformatics.babraham.ac.uk/projects/fastqc) and FASTX-Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/) to yield 84,373 trimmed reads, from which a contig was assembled with 80-fold coverage using SPAdes v3.5.0 (ATGGAGCCGGAGAGATCCTT-3\u2032 (forward) and 5\u2032-ACTTCATCAAGCGTGTCGGT-3\u2032 (reverse). The CPT Galaxy-Apollo phage annotation platform  was utilized for genome annotation , using otation 68. Structtation 6912. Gene tation 69\u201318. The Stenotrophomonas phage Pokken (GenBank accession number NC_049463.1) (Enterobacteria phage N4 (NC_008720.1), sharing 43 similar proteins  such as virion RNA polymerase (NCBI protein accession number YP_950528.1) and an SAR endolysin N-acetylmuramidase (YP_950539.1). Paxi was predicted by PhageTerm to contain 538-bp direct terminal repeats.Phage Paxi was determined to have a podovirus-like morphology  by viewi49463.1) , and BLAMZ326856. The associated BioProject, SRA, and BioSample accession numbers are PRJNA222858, SRR14095256, and SAMN18509291, respectively.The genome sequence for Paxi was deposited in GenBank under accession number"}
+{"text": "At all stages of f lowering, a decisive role is played by the family of MADS-domain transcription factors,the combinatorial action of which is described by the ABCDE-model of f lower development. The current volume ofdata suggests a high conservatism of ABCDE genes in angiosperms. The E-proteins SEPALLATA are the central hub ofthe MADS-complexes, which determine the identity of the f loral organs. The only representative of the SEPALLATA3clade in tomato Solanum lycopersicum L., SlMADS5, is involved in determining the identity of petals, stamens, andcarpels; however, data on the functions of the gene are limited. The study was focused on the SlMADS5 functionalcharacterization. Structural and phylogenetic analyses of SlMADS5 conf irmed its belonging to the SEP3 clade. Anin silico expression analysis revealed the absence of gene transcripts in roots, leaves, and shoot apical meristem,and their presence in f lowers, fruits, and seeds at different stages of development. Two-hybrid analysis showedthe ability of SlMADS5 to activate transcription of the target gene and interact with TAGL1. Transgenic plants Nicotianatabacum L. with constitutive overexpression of SlMADS5 cDNA f lowered 2.2 times later than the control; plantsformed thickened leaves, 2.5\u20133.0 times thicker stems, 1.5\u20132.7 times shortened internodes, and 1.9 times fewerf lowers and capsules than non-transgenic plants. The f lower structure did not differ from the control; however, thecorolla petals changed color from light pink to magenta. Analysis of the expression of SlMADS5 and the tobaccogenes NtLFY, NtAP1, NtWUS, NtAG, NtPLE, NtSEP1, NtSEP2, and NtSEP3 in leaves and apexes of transgenic and controlplants showed that SlMADS5 mRNA is present only in tissues of transgenic lines. The other genes analyzed werehighly expressed in the reproductive meristem of control plants. Gene transcripts were absent or were imperceptiblypresent in the leaves and vegetative apex of the control, as well as in the leaves and apexes of transgenic lines.The results obtained indicate the possible involvement of SlMADS5 in the regulation of f lower meristem developmentand the pathway of anthocyanin biosynthesis in petals. Throughout the plant\u2019s life cycle, its root and shoot apicalmeristems maintain a pool of pluripotent stem cells, whichgive rise to new organs: roots and leaves respectively, duringvegetative development and flowers during reproductionstage. At the reproductive stage, the shoot apical meristemof the angiosperms turns into the inflorescence meristem,which forms determined flower meristems . In all aspects of flowering, the MADS-domain familyof transcription factors (TFs) plays a key role according to thewell-known ABCDE flower development model .The ABCDE model is based on genetic and molecularstudies, primarily of model species Arabidopsis thaliana (L.)Heynh., Antirrhinum majus L., and Petunia\u00d7hybrida hort.ex E. Vilm. . Accordingto the model, the identity of flower organs is determinedby five classes of genetic activities: A and E \u2013 sepals; A, Band E \u2013 petals; B, C and E \u2013 stamens; C and E \u2013 carpels; C, Eand D \u2013 ovules. At the molecular level, the ABCDE-model isexplained by the so-called \u201cquartet\u201d model, according to whichMADS-TFs of ABCDE classes in various combinations formtetramers: for example, C/C/E/E \u2013 to determine carpel identity,or A/B1/B2/E \u2013 to specify petal identity . These tetramers activate or suppresstranscription of target genes . The current data suggest a high structural andfunctional conservatism of A, B, C, D, and E genes in floweringplants .The genes of the E-class, A. thaliana SEPALLATA , which are involved in determiningthe identity of all floral organs, deserve special attention . The knockout of onlyone of the SEP genes does not have a significant effect on theA. thaliana flower, while the sep1 sep2 sep3 triple mutationtransforms all the flower organs into sepals; a new flowerwith the same development pattern is formed instead of thepistil . The quadruple sep1 sep2 sep3 sep4mutation leads to the replacement of all flower organs withleaf-like organs .SEP proteins are the central hub in the formation of MADSTFquartets . Among SEPs, SEP3 isthe most functionally pleiotropic and interacts with almostall MADS-TFs responsible for the identity of flower organs. SEP3 gene simultaneous ectopic expression expressionwith the A-, B-, or C-class genes transforms leavesinto flower organs .During plant evolution, SEP genes are believed to havearisen later than other flower-related MADS-box genes, butat the same time they became key players in the origin offlowering plants, as well as in the domestication and breedingof crops . Therefore,their study in cultivated plants can expand the understandingof the role of these genes in determining economicallyvaluable traits.The tomato Solanum lycopersicum L. is one of the mostimportant vegetables and, at the same time, a model forstudying the fleshy fruit development and ripening. Thetomato genome has been sequenced and annotated (https://www.solgenomics.net/), and contains several SEP genes:TAGL2 (Solyc05g015750.2.1), SlMADS6/TM29/LeSEP1(Solyc02g089200.2.1), RIPENING INHIBITOR ( MADSRIN)(Solyc05g012020.2.1), SlMADS98/SlCMB1 (Solyc04g005320.2.1), SlMADS1/ENHANCER-OF-JOINTLESS-2(Solyc03g114840.2.1), SlMBP21/JOINTLESS-2 (J2)(Solyc12g038510.1.1) and SlMADS5/TM5/TDR5/LeSEP3(Solyc05g015750.3.1) .In addition to determining the flower organ identity, SEPproteins, together with MADS-TFs of the FRUITFULL (FUL)and AGAMOUS (AG) subfamilies, are actively involvedin the regulation of fruit ripening. This is clearly demonstratedin tomato, the fruit ripening of which is controlled byFUL1/FUL2, TOMATO AGAMOUS 1 (TAG1)/TOMATOAGAMOUS-LIKE 1 (TAGL1) and MADS-RIN . At thesame time, FUL2 and TAGL1 have been shown to play anadditional role in pistil initiation and early fruit development, which is likely tobe performed in combination with the tomato SEP3 homolog,SlMADS5 .SEP1-like gene TAGL2 was shown to be expressed atstages I (anthesis) and II of the tomato fruit development. Suppression of SEP1-like TM29 causesthe development of parthenocarpic fruits and the flower reversion. Tomato SEP4-likeSlCMB1 regulates ethylene biosynthesis and the accumulationof carotenoids during fruit ripening; suppression of SlCMB1leads to a change in the inflorescence architecture and anincrease in the sepal size . SEP4-likeSlMADS1 acts as a negative regulator of fruit ripening . SEP4-like SlMBP21 specifies the sepal size mediated by ethylene and auxin signaling, as well as the abscissionzone formation .SEP4-like MADS-RIN is the main regulator of fruit ripening:gene knockout leads to the formation of an unripe fruit, includingthe absence of carotenoid accumulation .The only representative of the tomato clade SEP3, TFSlMADS5, is involved in determining the identity of theorgans of the three inner flower whorls ,interacting with MADS-TFs of the SEP and AG subfamilies. Despite the SEP3 significance, thisgene variability has been characterized in cultivated and wildtomato species, and the SlMADS5 expression was observedin some organs and tissues .The aim of the present study was to characterize the functionof S. lycopersicum SlMADS5. SlMADS5 structural, phylogeneticand expression analysis confirmed its belonging to theSEP3-clade. Analysis in the yeast two-hybrid GAL4-systemshowed the SlMADS5 TF activator properties and itsinteraction with C-class MADS-TF. Transgenic Nicotianatabacum L. plants with SlMADS5 constitutive overexpressionexhibited a pronounced phenotype of reproductive developmentsuppression.Tomato S. lycopersicum cv. Silvestre recordo and tobaccoN. tabacum cv. Samsun plants were used in the study. Tomatoaccessions were grown under controlled greenhouse conditionsuntil flowering. Roots, leaves, flowers and ripe fruits werecollected separately. Tissues were grounded in liquid nitrogenand stored at \u201370 \u00b0C. Tobacco accessions were grown in vitroon a sterile MS medium in a climatic chamber  until the formationof 4\u20136 leaves.Total RNA was isolated from tomato  and tobacco  tissues using the RNeasy Plant Mini Kit, and used for cDNA synthesis . Genomic DNA wasisolated from leaf tissues by the standard potassium acetatemethod  and used for PCR tests forthe presence of a transgene in the plant genome.Primers for gene amplification, sequencing, and expressionanalysis were generated based on the MADS-box transcriptsof S. lycopersicum cv. Heinz and tobacco N. tabacumgenes available in the NCBI (http://www.ncbi.nlm.nih.gov/); NtLEAFY; NtWUSCHEL; NtAG ;NtPLENA : NtSEP1; NtSEP3 ; NtSEP2 ) so that forwardand reverse primers are separated by at least one intron andmatch all possible transcripts for each of the analyzed genes(Table 1). The primer sequences were additionally verifiedusing Primer 3 and BLAST (https://www.ncbi.nlm.nih.gov/tools/primer-blast/). Primers for CDS in-frame cloning intoplasmid vectors (GAL4 system) contained EcoRI and SalI  restriction sites at the 5\u2032 end.Full-length SlMADS5, TAG1, and FUL2 cDNAs wereamplified using the cDNA, isolated from S. lycopersicum cv.Silvestre recordo flowers; PCR conditions: initial denaturationat 95 \u00b0C for 5 min; 30 cycles of denaturation (94 \u00b0C for 30 s), annealing (55 \u00b0C \u2013 30 s) and synthesis (72 \u00b0C \u2013 1 min); finalsynthesis (72 \u00b0C \u2013 7 min). The PCR fragments of the expectedlength were purified using the MinElute Gel Extraction Kit, cloned into the pGEM\u00ae-T Easy plasmidvector  at EcoRI and Sal I sitesand sequenced (Core Facility \u201cBioengineering\u201d). Further, theSlMADS5, FUL2, and TAGL1 CDSs were cloned into hybridvectors pAD-GAL4 and pBD-GAL4cam : each gene was ligated in frame with the activatordomain (pAD) and DNA-binding domain (pBD) of the yeastTF GAL4. Recombinant pJ69-4a strains carrying each pADgeneand pBD-gene construct separately, as well as in pairspAD-gene + pBD-gene, were obtained. For plant transformation,SlMADS5 cDNA was cloned in a sense orientation intoa binary vector based on pBin19, under the control of theenhanced cauliflower mosaic virus promoter 35S and nopalinesynthase (NOS) terminator. With this construct, a recombinantagrobacterial strain AGL\u00d8 was obtained.For sequence structural analysis, the NCBI-CDD (http://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi), MEGA 7.0 and Phyre2 (http://www.sbg.bio.ic.ac.uk/phyre2/) were used. Sequence phylogeny was assessed inthe MEGA7, using Maximum Likelihood method based onthe JTT model.Gene expression analysis was performed in silico , as well as by quantitative (q) real-time (RT) PCRin two biological and three technical replicates. The kit \u201cReactionmixture for carrying out qRT-PCR in the presence ofSYBR Green I and ROX\u201d  and the CFX96Real-Time PCR Detection System  were applied. The qRT-PCR conditions were as follows:95 \u00b0C \u2013 5 min; 40 cycles .The reference gene actin-7 (XM_016658880.1)  was used for normalizing the expressionof tobacco genes. Statistical processing of the results wascarried out usingthe GraphPad Prism v. 7.02 (https://www.graphpad.com).The analysis of SlMADS5 interactions with TAGL1 andFUL2 proteins was carried out in vivo in a two-hybrid GAL4-yeast system using the Saccharomyces cerevisiae Pj69-4astrain, according to the HybriZAP-2.1-Hybrid cDNA Two-Hybrid Synthesis Kit protocol (Stratagene).Leaf explants of tobacco (N. tabacum cv. Samsun) weretransformed using Agrobacterium tumefaciens strain AGL\u00d8.To select transgenic regenerants, an MS medium containingkanamycin  for selection and carbenicillin(500 mg/L), which suppresses agrobacteria growth, was used.The rooted regenerants were adapted to the soil in greenhouseconditions and then tested for the presence of a transgene in thegenome by PCR with primers specific to the sequences of the5\u2032 end of the transgene and the NOS-terminator (see Table 1).To confirm the conservatism of the SlMADS5 function intomato (cv. Silvestre recordo), an analysis of its interactionswith MADS-TFs TAGL1 and FUL2, the interaction withwhich was and was not, respectively, shown earlier , was carried out.Structural analysis of the SlMADS5 protein was carried outin comparison with the known tomato, tobacco, and A. thalianaSEP homologs. The presence of the main domains characteristicof MIKCc type MADS-TFs was confirmed, namelythe highly conserved MEF2-like MADS-domain (1\u201376 aa),an I-region (77\u201392 aa), a conserved keratin (K)-like domain(93\u2013173 aa), and a variable C-region (174\u2013241 aa) .The performed phylogenetic analysis testified the belongingof SlMADS5 to the SEP3 clade .To characterize TF SlMADS5 functionally, we analyzedthe expression of the SlMADS5 gene in various tomato organsand the ability of SlMADS5 protein to activate gene transcriptionand interact with MADS proteins of the C and A classes.Also, transgenic N. tabacum model plants with constitutiveoverexpression of SlMADS5 cDNA were obtained.In silico analysis of the SlMADS5 expression pattern wascarried out in roots, leaves, vegetative shoot meristem, flowermeristem, flower (from bud to fully open and anthesis stage),fruits (4\u20138 days after anthesis), fruit skin and pulp (stages:Immature Green (IMG); Mature Green (MG); Breaker (BR),color change; Orange (OR); Red Ripe (RR)), and in seeds . SlMADS5 transcripts were notfound in roots, leaves, and the vegetative apical meristem. Atthe same time, SlMADS5 expression was shown in flowers(maximum \u2013 at the anthesis stage), fruits, fruit peel (maximumat MG and BR stages), fruit pulp , and seeds (maximum at IMG stage) .In vivo analysis in the yeast two-hybrid GAL4 systemshowed that TF SlMADS5 has the property of activating thetranscription of target genes, interacts with the C-class MADSprotein TAGL1, but does not interact with the A-class MADSprotein FUL2 (Table 2).The characterization of transgenic tobacco plants withSlMADS5 constitutive overexpression was performed. Independentregenerants T0 35S::SlMADS5 (18 plants) wereadapted to the greenhouse, tested by PCR for the presence ofa transgene expression cassette in the genome, and comparedwith the control (non-transgenic tobacco plants) during development.In comparison with the control, 35S::SlMADS5plants  bloomed much later . Also, 35S::SlMADS5 phenotype was characterizedby a 2.5\u20133.0 times thicker stem, 2.0 times shortened internodes, thickened and darker leaves, and 2.5 times fewerflowers and capsules. The 35S::SlMADS5 flower structure didnot differ from the control.Seeds of two transgenic T0 lines (S5-16 and S5-17) witha pronounced phenotype were planted in a greenhouse.T1 plants, which gave a positive PCR signal for the presenceof a transgene in the genome, bloomed 1.3\u20131.5 times laterthan the control, had a 35S::SlMADS5 phenotype, and formedflowers with magenta-colored corolla petals, in contrast tolight pink petals in the control.Seeds of lines T1 S5-16-6, S5-16-7, S5-17-1 and S5-17-4were planted on MS medium (Km 50 mg/l); the 3:1 ratio of thenumber of Km-resistant to Km-sensitive seedlings indicated aheterozygous state of the transgene and one copy of it in thegenome of transgenic lines. In seedlings, internodes were nearabsent, and only T2 plants of the S5-16-7 line (14 accessions)formed a noticeable stem and were adapted to the greenhouse(the rest of the plants died after transfer to the soil). PlantsT2 S5-16-7 demonstrated the 35S::SlMADS5 phenotype: theybloomed 2.4 times later than the control; formed thickenedstems and leaves, shortened internodes, and 2.3 times lessseed capsules.In T1 lines S5-16-7 and S5-17-1, in comparison with thecontrol, we analyzed the SlMADS5 expression, as well as theexpression of tobacco genes associated with reproductivedevelopment: NtLFY, NtAP1 (plant transition to flowering), NtWUS , NtAG,NtPLE, NtSEP1, NtSEP2, NtSEP3 (key genes for the identityof the flower meristem and flower organs). For the analysis,we used tissues of leaves and apical meristems  of transgenic and control plants.Expression of the SlMADS5 transgene was present onlyin the tissues of S5-16-7 and S5-17-1 plants. The expressionpattern of the remaining analyzed genes was similar: theirmRNA was absent or was minimal in the leaves of the controland transgenic lines, as well as in the S5-16-7 and S5-17-1apexes of undefined status. At the same time, these genes werehighly transcribed in the reproductive meristems of controlplants .In this study, a functional analysis of the SlMADS5 gene, theSEP3 homolog in tomato, was carried out. Structural analysis confirmed that SlMADS5 belongs to the SEP3clade, which may indicate the conservatism of its role in the reproductive development of tomato, namely, its participationin determining the identity of petals, stamens, carpels,and ovules.It is known that SlMADS5 is not expressed in tomato leavesand roots and is expressed in flowers and fruits . Also, SlMADS5 mRNA is present in the meristemdomains that correspond to the future three inner whorls ofthe tomato flower, as well as during organogenesis and inthe corresponding mature organs .A detailed in silico analysis of the SlMADS5 expression patterncarried out in this study revealed that SlMADS5 mRNAis absent not only in roots and leaves, but also in the shootapical meristems and flower meristems at early stages of development. Gene transcription is activated late inthe development of the flower meristem, and reaches a peakin an open flower and in the peel of an immature fruit . This corresponds not only to the well-known role ofSEP3 homologs in determining the differentiation of flowermeristem cells corresponding to the three inner whorls oforgans , but also suggests the active participation of SlMADS5 in the aspects of development andripening of tomato fruits and seeds.To characterize the SlMADS5 function, transgenic tobaccoplants with constitutive overexpression of SlMADS5 cDNAwere obtained. The phenotype of transgene overexpressiondoes not determine its function; however, it may indicate asimilarity with the already characterized homologs. Earlier,the effect of heterologous overexpression of SEP3 homologsof different plant species was studied mainly using transgenicA. thaliana plants, but there are works with the useof Nicotiana spp. plants. Tobacco, like tomato, belongs tothe Solanaceae family and has the same flower structure;therefore, in this study, a heterologous expression system intobacco was selected.Various effects of overexpression of SEP3 homologs havebeen described. Thus, SEP3 constitutive expression in A. thalianasignificantly accelerates flowering .In these plants, the APETALA3 (B-class) and AG (C-class)genes are transcribed ectopically .Overexpression of the P. \u00d7 hybrida SEP3-like gene FBP2leads to early flowering of the A. thaliana plants . Early flowering is caused by overexpression oftobacco SEP3-like gene NsMADS3 in N. sylvestris Speg. &Comes  and chrysanthemum SEP3-like geneCDM44 in N. tabacum .At the same time, no influence of overexpression of SEP3-homologous genes on the flowering time was also observed.Thus, homologous overexpression of FBP2 in P.\u00d7hybrida has no effect on plant vegetation period .Heterologous overexpression of Platanus acerifolia SEP3-likegenes in A. thaliana causes early flowering only in the caseof the PlacSEP3.2 gene, while overexpression of the secondgene, PlacSEP3.1, causes early flowering only in transgenictobacco plants .In the case of SlMADS5 constitutive overexpression, asignificant delay in flowering was observed, most likely associatedwith the incorrect development of the shoot apicalmeristem . Different effects of heterologous ectopicexpression of SEP3 homologs in transgenic plants maybe associated with structural differences in encoded proteinsequences responsible for binding to promoters of target genesor to partner proteins.Normally, traces of the A. thaliana SEP3 transcripts arefound in the inflorescence meristem, and gene expression isnoticeably activated only in the flower meristem parts, fromwhich petals, stamens, and carpels are subsequently formed. Therefore, thepresence of the TF SlMADS5 in tissues, where there shouldbe no tobacco SEP3 homologs, can lead to nonspecific proteinproteinand DNA-binding interactions of SlMADS5, whichcan disrupt the pattern of meristem development.To clarify the status of transgenic meristems S5-16-7 andS5-17-1, visually ready for flowering, we analyzed the expressionof genes whose activity is associated with the identity ofthe reproductive inflorescence and flower meristems (NtLFYand NtAP1) . Considering the results obtained , only the inflorescence meristem of thecontrol plant has reproductive status. The presence of a lowlevel of LFY expression in the vegetative apex of the controland in the S5-16-7 apex  suggests the initial stagesof the meristem transition to the reproductive state, since it hasbeen shown that in A. thaliana LFY begins to be expressed inthe flower meristem primordia at the periphery of the inflorescencemeristem .It is known that SEP3 is the central hub of the MADScomplexesin A. thaliana . TF SlMADS5also shows an exceptional ability to assemble tetrameric complexesof MADS TFs . The interactionof SlMADS5 with FUL2 and TAGL1 shown in this work (seeTable 2), as well as the role of FUL2 and TAGL1 in pistilinitiation and early fruit development , indicate the possible involvement ofSlMADS5 in determining the identity of the tomato pistil incomplex with FUL2 and TAGL1.One of the complexes, SEP3/SEP3/AG/AG, is requiredfor flower determination and completion of its development. This is due to a decrease in thenumber of stem cells because of the WUS gene suppressionwith the key participation of TF AG .Accordingly, in transgenic petunia plants with simultaneousoverexpression of SEP3-like FBP2 and D-class gene FBP11,where developmental arrest is observed at the cotyledonstage, transcription of AG-like FBP6 is activated and mRNAof WUS-like TERMINATOR is absent .This suggests the joint participation of SEP3, AG, and D-classgenes in the suppression of stem cells in the meristem.Taking into account the activation of AG expression inA. thaliana with SEP3 overexpression ,as well as the participation of SEP3 and AG in the suppressionof WUS transcription  and the interaction of TF SlMADS5 with theAG homolog TAGL1 (see Table 2), it can be assumed thatthe ectopically synthesized TF SlMADS5 is able to activatetranscription of the tobacco AG-like genes NtAG and NtPLEin transgenic shoot meristem. Subsequent formation of complexesSlMADS5/SlMADS5/NtAG/NtAG or SlMADS5/SlMADS5/NtPLE/NtPLE can lead to inhibition of meristemdevelopment due to the tobacco WUS-like gene NtWUSsuppression, since WUS plays a key role in determining thestem cell identity, the population of which is not supportedin plants with loss of WUS function .To test this possibility, we analyzed the expression ofSlMADS5, NtWUS, AG-like genes NtAG and NtPLE, as wellas SEP-like genes NtSEP1, NtSEP2, and NtSEP3. However,the presence of SlMADS5 ectopic expression did not leadto the activation of AG-like genes, and the expression ofNtWUS was significantly higher in the tissues of transgeniclines in comparison with the control (excluding the controlinflorescence meristem) . The latter can be a probablereason for the formation of significantly thickened, incomparison with the control, stem and leaves of transgenicplants of all 11 lines with the 35S::SlMADS5 phenotype  as a result of the increased number of stem cells andthe meristem overgrowth.It should also be noted that in transgenic plants, the anthocyanincolor of the flower corolla changed from pale pink(control) to magenta (35S::SlMADS5) . Previously,it was shown that the expression of the SEP-like geneMrMADS01 in Myrica rubra berries significantly increasesat the last stage of ripening, which allowed the authors tosuggest the involvement of this gene in the biosynthesis ofanthocyanins . Silencing the SEP-like genePaMADS7 in sweet cherry (Prunus avium) leads to a changein the content of anthocyanins in fruits . It canbe assumed that SlMADS5 is also involved in the regulationof anthocyanin biosynthesis in transgenic tobacco petals.Silencing of SlMADS5 gene leads to a change in the numberof flower whorls and the number of organs in whorls, as well asthe formation of green petals with signs of sepals, and sterileanthers and carpels with signs of sepals and petals, respectively, which may indicate the participation ofthe gene in determining the identity of tomato flower organs.Nevertheless, no complete homeotic transformation of certainflower organs was observed when SlMADS5 was suppressed.The data on the effect of SlMADS5 overexpression on thedevelopment of transgenic tobacco plants obtained in thisstudy also do not confirm the involvement of the gene indetermining the floral organ identity. 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+{"text": "The diagnostic performance of 18F-DCFPyL-PET/CT in localizing primary PCa within the prostate gland was assessed, allowing for PSMA-guided targeted-prostate biopsy.In primary prostate cancer (PCa) patients, accurate staging and histologic grading are crucial to guide treatment decisions. 18F-DCFPyL-PET/CT prior to robot-assisted radical prostatectomy (RARP). Two experienced and blinded nuclear medicine physicians assessed tumour localisation within the prostate gland on PET/CT, using a 12-segment mapping model of the prostate. The same model was used by a uro-pathologist for the RARP specimens. Based on PET/CT imaging, a potential biopsy recommendation was given per patient, based on the size and PET-intensity of the suspected PCa localisations. The biopsy recommendation was correlated to final histopathology in the RARP specimen. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) for clinically significant PCa  were assessed.Thirty patients with intermediate-/high-risk primary PCa were prospectively enrolled between May 2018 and May 2019 and underwent 18F-DCFPyL-PET/CT were 61.4%, 88.3%, 68.1% and 84.8%, respectively.The segments recommended for potential targeted biopsy harboured csPCA in 28/30 patients (93%), and covered the highest Gleason score PCa segment in 26/30 patient (87%). Overall, 122 of 420 segments (29.0%) contained csPCa at final histopathological examination. Sensitivity, specificity, PPV and NPV for csPCa per segment using 18F-DCFPyL-PET/CT with the RARP specimens, an accurate per-patient detection (93%) and localisation of csPCa was found. Thus, 18F-DCFPyL-PET/CT potentially allows for accurate PSMA-targeted biopsy.When comparing the PCa-localisation on The online version of this article (10.1007/s00345-020-03490-8) contains supplementary material, which is available to authorized users. Prostate cancer (PCa) is the most common cancer in men in the Western world , 2. HistBesides conventional imaging modalities such as mpMRI, novel imaging techniques including prostate-specific membrane antigen-positron emission tomography/computed tomography (PSMA-PET/CT) have been introduced. PSMA is significantly overexpressed in malignant prostate cells, correlates with higher tumour grades and represents a marker of tumour aggressiveness , 10. PSM18F-DCFPyL (PSMA) PET/CT imaging for the primary detection of PCa. The primary aim was to assess the accuracy of 18F-DCFPyL-PET/CT to localise primary PCa within the prostate gland, by comparing imaging results from 18F-DCFPyL-PET/CT to final histopathology of the robot-assisted radical prostatectomy (RARP) specimen. The secondary objectives were to investigate the ability of 18F-DCFPyL-PET/CT to provide a recommendation for potential targeted biopsy and to assess the diagnostic accuracy of determining local tumour stage (pT).This is the first prospective study on the accuracy of This was a prospective, non-randomised study in patients with diagnosed primary PCa. Pre-operative imaging results were compared to histopathology following RARP. All subjects signed informed-consent for the collection of their clinical data. The study has been approved by the ethical review board of the Amsterdam University Medical Centre (AUMC) (review number 2017.543). Patients were enrolled consecutively between May 2018-May 2019 in Amsterdam UMC, location VUmc.Patients had histologically proven, intermediate or high-risk, PCa, for which they underwent RARP , 15. Of 18F-DCFPyL which was synthesised under Good Manufacturing Practices conditions, as described by Jansen et al. [18F-DCFPyL (IQR 299\u2013324\u00a0MBq), and a median of 5.4\u00a0weeks (IQR 3.0\u20137.2) prior to surgery. Image-acquisitions were performed using a Philips Ingenuity TF -PET/CT system. No diuretics were administered prior to the scan. The scan trajectory included mid-thighs to skull-base, with 4\u00a0min per bed position. All PET scans were combined with a diagnostic CT scan , without contrast-enhancement. Images were corrected for decay, scatter, random coincidences, and photon attenuation.Patients were staged with n et al. , 18. PETImages were reconstructed with a BLOB-based Ordered-Subsets Expectations Maximization algorithm  ). The de18F-DCFPyL-PET/CT reading (>\u2009300 scans), in consensus. The readers used the 12-segment mapping model to demarcate the image-detected tumour extent (Appendix-1 in ESM) [max) of the suspected lesions. Finally, the readers indicated if radiological extra-capsular extension  or invasion into the seminal vesicles (rT3b) was suspected.Scan interpretation was performed blinded for the pathology results and other imaging by two nuclear medicine physicians  with ample experience in  in ESM) . For all in ESM)   guidelines . All spe18F-DCFPyL-PET/CT was matched to the histopathology results and the sensitivity, specificity, positive predicting value (PPV), and negative predicting value (NPV) were calculated on a segment basis. Correlation of 18F-DCFPyL-PET/CT with histopathology was considered if exactly the same segment was demarcated . Since there are no anatomical landmarks to delineate the segments, artificial segmentation can occur, causing a mismatch between the PET/CT- and histopathological findings while both correspond with the same lesion. Therefore, a second analysis of diagnostic accuracy was performed, in which PET correlation was also considered if there was a discrepancy of up to 1 region in the coronal or sagittal plane  [18F-DCFPyL-PET/CT to differentiate locally advanced disease (>\u2009rT3a) from prostate-confined disease (rT2). Numerical variables were summarised with median values and interquartile ranges (IQR); categorical variables with proportions (%). To compare medians of non-parametric data, the Mann\u2013Whitney-Wilcoxon test and the Kruskal-Wallis test were used (significance set at p\u2009<\u20090.05). Statistical analysis was performed with IBM\u00ae SPSS\u00ae Statistics for Windows\u00ae, version 26.The localisation of the detected prostate tumour by reement) , 24. RecA total of 30 patients was included in this study, having a median initial PSA-level of 11.1\u00a0ng/ml (IQR 5.8\u201322.4). According to EAU guidelines, 10/30 (33.3%) patients had intermediate-risk PCa and 20/30 (66.6%) had high-risk PCa . Pre-ope18F-DCFPyL-PET/CT to detect csPCa per segment with total agreement was 61.4% (95%CI 52.2\u201370.0%), 88.3% (95%CI 83.9\u201391.6%), 68.1% (95%CI 58.5\u201376.6%), and 84.8% (95%CI 80.2\u201388.5%), respectively (Appendix-2 in ESM). For near-total agreement, the sensitivity, specificity, PPV and NPV of 18F-DCFPyL PET/CT to detect csPCa per segment was 84.4% (95%CI 76.5\u201390.1%), 97.0% (95%CI 94.1\u201398.5%), 92.0% (95%CI 84.9\u201396.0%), and 93.8% (95%CI 90.3\u201396.1%), respectively. The area under the curve (AUC) of 18F-DCFPyL-PET/CT was 0.78 (95%CI 0.73\u20130.84) for the total agreement scores, and 0.85 (95%CI 0.80\u20130.90) for the near-total agreement scores (Appendix-3 in ESM). True positive segments had a median SUVmax of 8.26 (IQR 5.25\u201311.40), which was significantly higher than the median SUVmax of false-positive segments of 4.06 (IQR 3.56\u20135.10) (p\u2009=\u20090.02). The median SUVmax of true positive segments did not correlate with ISUP grade groups (p\u2009=\u20090.95) (Appendix-4 in ESM).All patients showed PSMA expression in the prostate. In 30 evaluated patients, 420 segments  could be used both for PET/CT and histopathological mapping evaluation. PCa was present in 129 of the 420 (30.7%) segments on histopathological examination, and csPCa was found in 122 of the 420 segments (29.0%) . The sensitivity, specificity, PPV and NPV of 18F-DCFPyL-PET/CT revealed csPCa in 28/30 (93.3%) patients. Moreover, it pinned the index PCa lesion in 26/30 (86.7%) patients. An example of potential 18F-DCFPyL-guided biopsy recommendation and concurrent histopathological examination of the RARP specimen is shown in Fig. max 8.62 (IQR 6.41\u201312.62). The recommendation for potential biopsy from the nuclear physicians that matched csPCA had a median SUVmax of 8.55 (IQR 6.34\u201313.79), and was significantly higher than the recommended potential biopsy segments that did not contain csPCa (median SUVmax of 3.10 (IQR 2.86\u20133.87) (p\u2009=\u20090.02).The primary potential biopsy recommendation by the nuclear medicine physician harboured csPCa in 24/30 (80.0%) patients and detected the index PCa lesion in 23/30 (76.6%) patients. When both the primary and secondary recommended segments would potentially be targeted, 18F-DCFPyL-PET/CT to detect locally advanced tumour growth (\u2265\u2009pT3a) was 35.7% (95%CI 14.0\u201364.3%), 93.8% (95%CI 67.7\u201399.7%), 83.3% (95%CI 36.5\u201399.1%), and 62.5% (95%CI 40.8\u201380.4%), respectively (Appendix-5 in ESM). For the detection of pT3a sub-stage, the sensitivity, specificity, PPV and NPV of 18F-DCFPyL-PET/CT were 20.0% (95%CI 3.5\u201355.8), 100% (95%CI 80.0\u2013100), 100.0% (95%CI 19.7\u2013100), and 71.4% (95%CI 51.1\u201386.0), respectively. For the detection of pT3b sub-stage, the sensitivity, specificity, PPV and NPV of 18F-DCFPyL-PET/CT were 75.0% (95%CI 21.9\u201398.7), 92.3% (95%CI 73.4\u201398.7), 60.0% (95%CI 17.0\u201392.7), and 96.0% (95%CI 77.7\u201399.8), respectively.Final histopathological analysis revealed pT3a in 10/30 (33.3%) patients, and pT3b in 4/30 (13.3%) patients. The sensitivity, specificity, PPV and NPV of 18F-DCFPyL-PET/CT imaging was used to locate primary PCa within the prostate gland, exploring the diagnostic potential of PSMA-based targeted biopsies. A total of 30 patients diagnosed with intermediate and high-risk PCa that underwent 18F-DCFPyL-PET/CT prior to RARP was analysed. When using a prostate-mapping model, the potential 18F-DCFPyL-PET/CT-based targeted biopsy recommendation detected csPCa in 28/30 (93.3%) patients. Moreover, it detected the index PCa lesion in 26/30 (86.7%) patients. Potentially, this makes PSMA-targeted biopsy a diagnostic tool that may adequately guide precision prostate biopsy. In biopsy-naive patients at increased risk of metastatic spread, and in whom staging imaging is mandatory (e.g. PSA\u2009\u2265\u200920), 18F-DCFPyL-PET/CT could potentially be used simultaneously to stage patients and to target PSMA-avid prostatic lesions suspicious for PCa.This is the first prospective study in which 18F-DCFPyL-PET/CT imaging demonstrated a moderate per segment-based sensitivity for the detection of csPCa of 61.4%, at a 88.3% specificity. The moderate sensitivity indicates that 18F-DCFPyL-PET/CT was not able to detect all localised csPCa. Segmentation of the prostate gland is problematic, as no clear anatomical landmarks are available to delineate the different segments within the prostate (Appendix-1 in ESM). A tumour located on the border of the apex and middle part of the prostate could be classified in different segments by the nuclear medicine physician and uro-pathologist, while in fact, they detected the same lesion. Therefore, the near-agreement score was introduced to approximate clinical reality. The sensitivity of the near-agreement score of 18F-DCFPyL-PET/CT imaging for the detection of csPCa was higher at 84.4% with a specificity of 97.0%.68Ga-PSMA-PET/CT and mpMRI prior to RARP [MpMRI has found a prominent place in the identification and localisation of PCa . Few stu to RARP . This st18F- PSMA-1007-PET/CT and mpMRI with subsequent RARP. Nine of the men were diagnosed with MRI-TBx and 1 with systematic biopsy.\u00a0Using a 36-segment mapping model, a similar assessment of agreement and near-total agreement was used. In 10 patients, 18F-PSMA-1007 PET/CT showed a high sensitivity , specificity , and accuracy  for the detection of csPCa. Although the specificity was similar, this study did however show lower sensitivity for PSMA-PET/CT compared to mpMRI . Above mentioned studies implicate that PSMA-PET/CT imaging performs at least equal to mpMRI to locate primary PCa. The rates of mpMRI might have been overestimated as at least a part of included patients in previously mentioned studies were diagnosed by MRI-TBx. Thus, selection bias may have been introduced.In another study, Kesch et al.  studied 18F-DCFPyL-PET/CT. So, similar to mpMRI, a substantial number of patients with\u2009\u2265\u2009pT3a was understaged by PSMA-based imaging [18F-DCFPyL-PET/CT of 93.8% is congruent with similar 68\u00a0Ga-PSMA studies (specificity\u2009>\u200990% for T3b disease) [There is therapeutic importance to distinguish between T2 and T3 disease  [disease) , 25, 26.18F-DCFPyL-PET/CT performs in a truly biopsy na\u00efve cohort of patients. The present study was set up to evaluate the capability of PSMA-PET/CT to guide targeted prostate biopsy for the detection of csPCa, not with the goal to discriminate between those who should be or should not be biopsied. Unfortunately, not all patients received a pre-operative mpMRI, limiting direct comparison to 18F-DCFPyL-PET/CT. Moreover, some of the patients who received a mpMRI had a longer interval between the mpMRI and the PET/CT scans due to the mpMRI being performed at the referring centre. Finally, since the RARP-specimen will always change shape (due to organ slicing and shrinking artefacts) when it is removed from the body, no truly exact anatomical correlation is possible. Therefore, the partial agreement score was used to correct for this pitfall.Our study has inherent limitations. Since the PET/CT resolution is confined at 5\u00a0mm, limited diagnostic accuracy for small PCa-foci is to be expected. A selection bias has been introduced due to the selection of patients with biopsy-confirmed csPCa. It is thus unclear how 18F-DCFPyL-PET/CT with the RARP specimen using anatomical mapping, an accurate per-patient localization (93%) of csPCa was found within the prostate. 18F-DCFPyL-PET/CT proves promising for PSMA-targeted biopsy and provides a moderate local staging ability.When comparing the localisation of PCa on Supplementary file1 (JPG 61 kb)Supplementary file2 (JPG 138 kb)Supplementary file3 (JPG 79 kb)Supplementary file4 (JPG 70 kb)Supplementary file5 (JPG 177 kb)Below is the link to the electronic supplementary material."}
+{"text": "Aegilops sharonensis, a wild relative of wheat, harbors diverse disease and insect resistance genes, making it a potentially excellent gene source for wheat improvement. In this study, we characterized and evaluated six wheat-A. sharonensis derivatives, which included three disomic additions, one disomic substitution + monotelosomic addition and two disomic substitution + disomic additions. A total of 51 PLUG markers were developed and used to allocate the A. sharonensis chromosomes in each of the six derivatives to Triticeae homoeologous groups. A set of cytogenetic markers specific for A. sharonensis chromosomes was established based on FISH using oligonucleotides as probes. Molecular cytogenetic marker analysis confirmed that these lines were a CS-A. sharonensis 2Ssh disomic addition, a 4Ssh disomic addition, a 4Ssh (4D) substitution + 5SshL monotelosomic addition, a 6Ssh disomic addition, a 4Ssh (4D) substitution + 6Ssh disomic addition and a 4Ssh (4D) substitution + 7Ssh disomic addition line, respectively. Disease resistance investigations showed that chromosome 7Ssh of A. sharonensis might harbor a new powdery mildew resistance gene, and therefore it has potential for use as resistance source for wheat breeding. Aegilops sharonensis Eig , a wild relative of wheat, is endemic to the coastal plains of Israel and southern Lebanon  substitution line (A. sharonensis 4Ssh (4D) substitution line by using a nullisomic backcrossing procedure. A. sharonensis amphiploid (genome AABBS1S1). A. sharonensis. A. sharonensis introgression lines. Recently, both A. sharonensis amphidiploids. However, there are very few reports on the isolation of wheat plants carrying individual A. sharonensis chromosomes. A. sharonensis, and produced three 1Ssh (1A) substitution lines, two 1Ssh (1B) substitution lines, three 1Ssh (1D) substitution lines and two 1Ssh (5D) substitution lines. Therefore, the set of wheat-A. sharonensis chromosome lines is still not complete, which greatly limits the mapping and utilization of excellent genes derived from this species in wheat.ion line . Later, A. sharonensis chromosome derivatives, including three disomic addition lines, one disomic substitution + monotelosomic addition line, and two disomic substitution + disomic addition lines, were identified by (Polymerase Chain Reaction) PCR-based landmark unique gene (PLUG) markers and fluorescence in situ hybridization (FISH) analysis. In addition, the infection types (ITs) of disease resistance, spike and grain characteristics of these wheat-A. sharonensis chromosome lines were also investigated to provide useful information for the possible subsequent development of wheat-A. sharonensis translocations for wheat genetic improvement.In this study, six wheat-Triticum aestivum cv. Jinan17 (JN17) and Jimai22 (JM22) were maintained at the Crop Research Institute, Shandong Academy of Agricultural Sciences in Jinan. T. aestivum cv. Chinese Spring (CS) was provided by Prof. Z. J. Yang, School of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu. The diploid A. sharonensis accession (TA1995) was provided by Mr. J. Raupp, Wheat Genetic and Genomic Resources Center, Kansas State University, United States. The CS-A. sharonensis amphiploid (JIC-31) and six unidentified CS-A. sharonensis chromosome lines  were kindly provided by Prof. S. M. Reader, John Innes Centre, United Kingdom.8 were synthesized by Chengdu Ruixin Biological Technology Co., Ltd. Probe sequences, the fluorochromes for probe labeling, FISH protocols and labeled DNA signal detection methods were according to 8 as a probe could be used to identify wheat chromosomes except 1A, 3D, 4D, 5D, and 6D, as described by EcoRI fragment isolated from bread wheat d JIC-31 . The karA. sharonensis chromosomes in lines JIC-32 to JIC-37 showed no signals associated with that probe. However, slightly different Oligo-pSc119.2-1 signals were found on both terminal regions of all A. sharonensis chromosomes , indicating that this pair of A. sharonensis chromosomes might be 1Ssh or 6Ssh.FISH using Oligo-pTa535-1 onto omosomes . In addie JIC-32 . MoreoveA. sharonensis chromosomes in JIC-32 to JIC-37, a total of 526 PLUG primer pairs were used to develop A. sharonensis chromosome-specific markers. As a result, fifty-one primer pairs could generate polymorphisms in A. sharonensis, the CS-A. sharonensis amphiploid, CS, JM22, and JN17. Among them, four, eight, nine, six, two, five, and seventeen belonged to chromosome homoeologous groups 1\u20137, respectively  substitution + 5SshL monotelosomic addition (A. sharonensis 6Ssh disomic addition (A. sharonensis 4Ssh (4D) substitution + 6Ssh disomic addition (sh (4D) substitution + 7Ssh disomic addition line  substitution + 5SshL monotelosomic addition, 6Ssh disomic addition, 4Ssh (4D) substitution + 6Ssh disomic addition and 4Ssh (4D) substitution + 7Ssh disomic addition lines were more elongated than that of CS. The CS-A. sharonensis 4Ssh disomic addition line (JIC-33) showed slightly elongated spikelets and overall longer spikes than that of CS. The CS-A. sharonensis 4Ssh (4D) substitution + 6Ssh disomic addition (JIC-36) showed shorter spikes and fewer spikelets per head than that of CS  showed slender grains and darker pericarp color than that of CS (sh disomic addition (JIC-33) showed smaller grains than those of CS.Grain morphologies of the six CS-at of CS  and the A. sharonensis 4Ssh (4D) substitution + 7Ssh disomic addition (JIC-37) was not obtained. The CS-A. sharonensis 4Ssh (4D) substitution + 7Ssh disomic addition (JIC-37) was nearly immune to powdery mildew, while CS and other CS-A. sharonensis chromosome lines tested were highly susceptible to powdery mildew (sh of A. sharonensis might carry powdery mildew resistant gene(s).Stripe rust, leaf rust, stem rust, and powdery mildew resistance tests showed that all the materials were moderately to highly susceptible to stripe rust, leaf rust, and stem rust  except ty mildew , suggestA. sharonensis chromosomes into wheat is difficult due to the presence of gametocidal (Gc) genes that control the preferential transmission of chromosome 4Ssh (A. sharonensis additions or substitutions (T. urartu-A. sharonensis amphiploid TA3398 in North Dakota in 1972 (unpublished). A. sharonensis chromosome 4S1 (some scientists defined the genome of A. sharonensis as S1S1). A. sharonensis Gc2 gametocidal gene (Gc2mut), which opened a way for introgression of genes from A. sharonensis into wheat. 1, 3S1, 5S1, 6S1, and 7S1 in a 4S1 (4D) background. A. sharonensis introgression lines which they then separated into six groups based on different substituted chromosomes belonging to definite homoeologous groups and different numbers of translocations. A. sharonensis amphiploid (genome AABBS1S1). A. sharonensis, and produced three 1Ssh (1A) substitution lines, two 1Ssh (1B) substitution lines, three 1Ssh (1D) substitution lines and two 1Ssh (5D) substitution lines.Transferring each pair of ome 4Ssh . Therefoitutions . Maan foA. sharonensis introgression lines are very rare. Furthermore, none to date has reported the production of wheat-A. sharonensis 2Ssh introgression lines. In this research, six wheat-A. sharonensis introgression lines were identified, including a CS-A. sharonensis 2Ssh disomic addition (JIC-32), a 4Ssh disomic addition (JIC-33), a 4Ssh (4D) substitution + 5SshL monotelosomic addition (JIC-34), a 6Ssh disomic addition (JIC-35), a 4Ssh (4D) substitution + 6Ssh disomic addition (JIC-36) and a 4Ssh (4D) substitution + 7Ssh disomic addition (JIC-37). Among these six introgression lines, four possessed chromosome 4Ssh, suggesting that chromosome 4Ssh of A. sharonensis was transmitted preferentially into wheat due to the gametocidal gene, which confirms the reports of preferential transmission of gametocidal chromosomes of earlier researchers  substitution + 5SshL monotelosomic addition (JIC-34) and the CS-A. sharonensis 4Ssh disomic addition line (JIC-33) were highly susceptible to powdery mildew, indicating that there were no powdery mildew resistance genes on chromosomes 4Ssh and 5SshL of A. sharonensis. However, the CS-A. sharonensis 4Ssh (4D) substitution + 7Ssh disomic addition (JIC-37) was nearly immune to powdery mildew (sh of A. sharonensis might carry new powdery mildew resistant gene(s).To date, more than 70 powdery mildew resistance genes have been permanently designated . Among ty mildew , suggestThe datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/CL conceived and designed the experiments. XW, ZHY, HW, JBL, and RH performed the experiments. WX, GL, and JG performed disease resistance testing. YZ, FL, DC, and AL analyzed the data. XW wrote the manuscript. HL, ZJY, JJL, and CL revised the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "We describe a case of metastatic pulmonary calcification in a 71-year-old male, images with  He was on diuretic therapy for chronic kidney disease for 4 years, followed by oral adsorptive carbon therapy for 13 months. He had been on hormonal therapy for prostate cancer (cT2cN0M0) for 9 months. He was hospitalized elsewhere for poor oral intake and general weakness. Laboratory studies at that time revealed the following : calcium 15.5 mg/dL (8.2\u201310.2 mg/dL); phosphorus 7.8 mg/dL (2.5\u20134.5 mg/dL); creatinine 3.67 mg/dL (0.7\u20131.2 mg/dL); total 25-hydroxyvitamin D level 12.06 ng/mL (approx. 30 ng/mL); and parathyroid hormone 15.71 pg/mL (15\u201365 pg/mL). 18F-fluorodeoxyglucose (FDG) PET/CT was performed to exclude the possibility of malignancy-related hypercalcemia. 18F-FDG) PET/CT images were acquired 1 h after intravenous injection of 238 MBq of 18F-FDG. The PET/CT images showed an increase in FDG uptake in the bilateral lower lungs  bone scan. It also revealed significantly increased diffuse uptake in the bilateral lower lung fields (Further examination with a bone scan was therefore recommended. Since the bone scan is a sensitive test for diagnosing metastatic pulmonary calcification, we performed a bone scan to identify metastatic pulmonary calcification and determine whether bone metastasis existed . The patg fields . The sca18F-FDG PET/CT to detect metastatic pulmonary calcification [18F-FDG PET/CT findings in patients with metastatic pulmonary calcification. In conclusion, in patients with chronic kidney disease, when hypercalcemia is present and PET/CT shows ground-glass opacity with mild FDG uptake, metastatic pulmonary calcification can be considered one of the differential diagnoses, though this is rare.Metastatic pulmonary calcification is a frequently underdiagnosed disease. Because usual imaging modalities such as chest radiographs and CT scan findings are not specific ,9. Only fication ,11. Howe"}
+{"text": "Biliary tract cancers (BTCs) are rare tumours with regional differences. Prognostic factors are poorly understood. Gemcitabine + platinum (GP) is the standard first-line chemotherapy in metastatic patients. We aimed to search for prognostic factors in patients with advanced disease in a cancer centre in South America.We conducted a retrospective analysis of patients with advanced BTC treated with chemotherapy. Variables were age (< or \u226570 years), Eastern Cooperative Oncology Group (ECOG) performance status (0/1 versus 2/3), gender, primary site (intrahepatic (IHC), extrahepatic (EHC), gallbladder (GB)), staging , metastatic sites, albumin (>3.5 g/dL versus <3.5 g/dL), biliary obstruction and first-line chemotherapy . Cox regression method was used to explore factors.p = 0.007), albumin < 3.5 g/dL (p = 0.001), biliary obstruction (p = 0.006), 5FU-based (p = 0.006) and single-agent (p < 0.0001) were associated with worse OS. ECOG performance status 2/3 (p = 0.058) and bone metastases (p = 0.051) were marginally related. In multivariate analysis, male (p = 0.003), bone metastases (p = 0.023), biliary obstruction (p = 0.001), 5FU-based (p = 0.016) and single-agent (p = 0.023) were independently associated with inferior OS.From 2010 to 2017, 104 patients were included. Median age was 62 years (32\u201386) and 22.1% were older than 70 years. Most patients had ECOG performance status 0/1 (63.4%), were female (51.9%) and were metastatic (82.7%). Bone metastases were found in 19.2%. Primary IHC, EHC and GB were 54.8%, 36.5% and 8.7%, respectively. GP was used by 79.8%. Median follow-up was 32.4 months. Median overall survival (mOS) was 11.4 months. In univariate analysis, male (In this retrospective study, we observed that male patients, bone metastases, biliary obstruction and regimens other than GP had worse survival. Larger studies should be conducted to confirm our findings. Biliary tract cancers (BTCs) are a rare group of malignances . Main suWe conducted a retrospective analysis of patients treated at a cancer centre in Brazil, from 2010 to 2017. Patients were included if they were 18 years of age or older, had histologic diagnosis of metastatic, recurrent or locally advanced unresectable BTC . All patients were deemed eligible to receive first-line chemotherapy by their treating physician. Exclusion criteria were absence of histologic confirmation, absence of data regarding systemic treatment and mixed histologies (hepatocellular-cholangiocarcinoma). Medical files were used as source of information. This study was approved by the local Ethics Committee (2680/19).Descriptive statistics were used for main demographic characteristics. Survival curves were estimated using Kaplan\u2013Meier method and compared with log-rank test. OS was defined as date of first chemotherapy cycle and death from any cause. Progression-free survival (PFS) was defined as date of first chemotherapy cycle and disease progression or death from any cause. Radiologic assessments of response and clinical benefit were performed according to local guidelines, typically by means of clinical examination every 2 weeks and computed tomography or magnetic resonance imaging every 8 weeks. Progressive disease was identified by the treating physician.2X tests were used to analyse categorical variables distributions between genders. To evaluate prognostic factors, univariate and multivariate analysis were performed using the Cox regression method. Age (<70 years versus >70 years), gender, staging , primary site (EHC versus IHC versus GB carcinoma), Eastern Cooperative Oncology Group (ECOG) performance status (0\u20131 versus 2\u20133), bone metastasis, baseline Carbohydrate Antigen 19-9 (CA19-9), albumin (<3.5 g/dL versus > 3.5 g/dL), biliary obstruction and first-line treatment (gemcitabine + platinum (GP) versus 5-Fluorouracil (5-FU) based versus monotherapy) were used in the univariate model. All tests were considered statistically significant with a two-sided p value of <\u20090.05. Statistical analysis was performed with IBM SPSS 20.Between July/2009 and March/2017, 104 patients were identified. Main demographics are shown in Regarding first-line chemotherapy , GP, 5-Fp = 0.32). Access to second-line was not statistically different among gender .Numerically, more female patients used GP as first-line . At time of analysis, seventy-six (73.1%) patients had died. Median OS (mOS) was 11.4 months (95% CI: 9.0\u201313.7) . Median p = 0.007), albumin < 3.5 g/dL (HR: 4.73 (95% CI: 1.96\u201311.4); p = 0.001), biliary obstruction (HR: 2.15 (95% CI: 1.24\u20133.71); p = 0.006), 5FU-based (HR: 2.64 (95% CI: 1.31\u20135.30); p = 0.006) and single-agent (HR: 6.27 (95% CI: 2.36\u201316.6); p < 0.0001) were associated with worse OS. Bone metastases (HR: 1.82 (95% CI: 0.99\u20133.34); p = 0.051) and ECOG performance status 2\u20133 (HR: 1.58 (95% CI: 0.98\u20132.53); p = 0.058) were marginally associated with prognosis.In univariate analysis, male patients (hazard ratio (HR): 1.88 (95% CI: 1.19\u20132.98); p = 0.003), bone metastases (HR: 3.53 (95% CI: 1.18\u201310.5); p = 0.023), biliary obstruction , 5FU-based (HR: 6.24 (95% CI: 1.41\u201327.6); p = 0.016) and single agent chemotherapy (HR: 7.22 (95% CI: 1.30\u201339.8); p = 0.023) were significantly and independently associated with inferior OS ; erior OS . mOS waserior OS .BTC is a rare gastrointestinal neoplasm but with increasing incidence over the past years, especially for IHC . SurgeryOur data bring relevant information about two aspects of metastatic BTC. The first is regarding the standard first-line treatment. Patients treated with GP regimens presented a more favourable outcome. GP was superior to Gemcitabine in the Phase III ABC-02 study and was established as the standard of treatment first-line therapy for metastatic biliary tract carcinomas . On the et al [p = 0.0031) and OS (13.0 versus 7.5 months (HR: 0.31 (95% CI: 0.15\u20130.63); p = 0.0013). However, in our study, 7.6% of patients were treated with FOLFOX in first-line and despite the small sample, outcomes were worse than those of patients treated with Gemcitabine combinations, reinforcing the role of GP. Clinical reasons not to employ GP were not accurately assessed in our analysis.Recently, the ABC-06 trial established FOLFOX as a new standard for second-line treatment of BTC . Based oet al  publisheSecond, female gender was significatively associated with better survival. We found that survival was almost doubled among female patients. In analogous tumours like metastatic pancreatic cancer, gender is a controversial prognostic factor , 14 . Hoet al [p = 0.037). Baton et al [p = 0.0002). Another retrospective large series from Korea with 740 BTC patients [p = 0.04).Clinical data endorses our findings. Bridgewater et al  describeon et al  publishepatients  also shop = 0.03) [Some aspects of BTC regarding access to therapeutics may help clarify this difference. Undergoing second and further lines of treatment is probably a strong prognostic factor. In a large study of second-line therapy, out of 378 patients that received first-line chemotherapy, only 96 patients (25%) received second-line. Female/male ratio was 31% and 21%, respectively ( = 0.03) . In our Currently, it is clear that there are striking differences in gastrointestinal cancer incidences among male and female patients. These differences are in part explained by exposition to risk factors such as smoking . NeverthDrug effects are also variable between genders. 5-FU based chemotherapy tends to be more toxic in women. A polled analysis of more than 28,000 patients, demonstrated that neutropenia and gastrointestinal toxicity were more likely to affect female patients . BenefitFinally, it is noteworthy that we have a growing body of evidence regarding molecular biology of BTC. Fibroblast Growth Factor Receptor (FGFR) fusions and Isocitrate Dehydrogenase (IDH) mutations are among targetable alterations in this scenario. Clinical trials have shown benefit of targeted therapies for such patients , 29. DesIn summary, our data identified prognostic factors related to outcome in metastatic BTC. Although FOLFOX has become the standard on the second-line, in our series patients receiving first-line treatment other than gemcitabine combination had worse prognosis. We also showed that bone metastasis, biliary obstruction and male gender were correlated with worse prognosis. These data could be useful for future trials in selecting patients with higher risk to more intensive and precise therapy. Further studies and multicentric collaborations are fundamental to validate our findings and also to understand the biology of this rare neoplasm.None.The authors declare that they have no conflicts of interest.Approved by local ethics committee (2680/19).Not applicable.All authors contributed to study concept and design; data acquisition; data analysis and interpretation; drafting of the manuscript; critical revision of the manuscript intellectual content and statistical analysis."}
+{"text": "Bacillus cereus strain HT18, isolated from forest soil, was 5,333,415\u2009bp long. The genome included 5,825 putative coding sequences and 35.2% GC content; the strain had 5 plasmids. Average nucleotide identity based on BLAST+ (ANIb) and digital DNA-DNA hybridization (dDDH) results showed that HT18 was 98.78% and 90.70% homologous, respectively, to B. cereus ATCC 14579T.The genome sequence of  Bacillus cereus group (phylum Firmicutes) comprises Gram-positive, spore-forming, facultative, anaerobic, rod-shaped bacteria with low-GC-content genomes . After colony isolation, the cells were cultured in LB broth at 37\u00b0C for 24 h. Then, genomic DNA was isolated using Marmur\u2019s method .http://platanus.bio.titech.ac.jp/pltanus_trim)  (dnaA first was performed with DFAST version 1.4.0 (https://dfast.ddbj.nig.ac.jp) (http://jspecies.ribohost.com/jspeciesws/) and the Genome-to-Genome Distance Calculator version 3.0 (http://ggdc.dsmz.de/ggdc.php)  and decoded on a MiSeq instrument (Illumina). A total of 1,488,012 reads with 431,000,004 bases were decoded with an average insert size of 641\u2009bp and spot length of 602\u2009bp with 2\u2009\u00d7\u2009300-bp paired ends. Low-quality bases (Q scores of <15) were trimmed, and short reads (<25\u2009bp) were removed using Platanus trim version 1.0.7 (gdc.php)  using thOne contig of the assembled genome sequence was 5,333,415\u2009bp long (35.2% GC content) with a sequence depth of approximately 80. A total of 5,825 coding regions, 107 tRNAs, 42 rRNAs, and 3 CRISPR regions were annotated. Five plasmid DNA sequences were also assembled .B. cereus strains WPySW2 and AFA01, and 99.9% and 99.9% homologous to the foodborne pathogen FORC021 (https://figshare.com/articles/dataset/suppl_Table_pdf/17111096). Although strain HT18 showed high homology to B. thuringiensis serovar Berliner ATCC 10792T, it was closely related to B. cereus ATCC 14579T when it was compared to reference strains of the B. cereus group. Therefore, strain HT18 was identified as a B. cereus strain.Based on the ANIb (%) and dDDH (%) results, strain HT18 was 99.9% and 100% homologous to both PRJDB11181, BioSample number SAMD00278679, DRA number DRA011610, accession numbers AP024504 to AP024509, and SRA numbers DRX266213 (Illumina) and DRX266214 (ONT).The genome sequence and annotation data for strain HT18 were deposited in DDBJ/GenBank under BioProject number"}
+{"text": "Wheatgrass Thinopyrum intermedium is a source of agronomically valuable traits for common wheat. Partial wheat\u2013wheatgrass amphidiploids and lines with wheatgrass chromosome substitutions are extensively used as intermediates in breeding programs. Line Agis 1 (6Agi2/6D) is present in the cultivar Tulaykovskaya 10 pedigree. Wheatgrass chromosome 6Agi2 carries multiple resistance to fungal diseases in various ecogeographical zones. In this work, we studied the transfer of chromosome 6Agi2 in hybrid populations Saratovskaya 29 \u00d7 skaya 10 (S29 \u00d7 T10) and Tulaykovskaya 10 \u00d7 Saratovskaya 29 (T10 \u00d7 S29). Chromosome 6Agi2 was identif ied by PCRwith chromosome-specif ic primers and by genomic in situ hybridization (GISH). According to molecular data, 6Agi2was transmitted to nearly half of the plants tested in the F2 and F3 generations. A new breeding line 49-14 (2n = 42)with chromosome pair 6Agi2 was isolated and characterized in T10 \u00d7 S29 F5 by GISH. According to the results ofour f ield experiment in 2020, the line had high productivity traits. The grain weights per plant (10.04 \u00b1 0.93 g) andthe number of grains per plant (259.36 \u00b1 22.49) did not differ signif icantly from the parent varieties. The number ofgrains per spikelet in the main spike was signif icantly higher than in S29 ( p \u2264 0.001) or T10 ( p \u2264 0.05). Plants werecharacterized by the ability to set 3.77 \u00b1 0.1 grains per spikelet, and this trait varied among individuals from 2.93 to4.62. The grain protein content was 17.91 %, and the gluten content, 40.55 %. According to the screening for fungaldisease resistance carried out in the f ield in 2018 and 2020, chromosome 6Agi2 makes plants retain immunity tothe West Siberian population of brown rust and to dominant races of stem rust. It also provides medium resistantand medium susceptible types of response to yellow rust. The possibility of using lines/varieties of bread wheatwith wheatgrass chromosomes 6Agi2 in breeding in order to increase protein content in the grain, to confer resistanceto leaf diseases on plants and to create multif lowered forms is discussed. Wild perennial common wheat relatives of the Thinopyrumgenus are broadly polymorphic. They can be sources ofcommercially valuable traits: resistance to fungal and viraldiseases ,tolerance of saline soils and drought, and high protein contentsin the grain . TheThinopyrum genus includes about 20 species of differentploidies: diploids, allotetraploids, allohexaploids, octoploids,and decaploids . The genetic poolsof two species are in the greatest use: elongate wheatgrassTh. elongatum (Agropyron elongatum) and intermediatewheatgrass Th. intermedium (Ag. glaucum). They becamedonors of genes for resistance to pests: Lr19, Lr24, Lr29,and Lr38 to brown rust; Sr24, Sr25, Sr26, Sr43, and Sr44to stem rust; Pm40 and Pm43 to powdery mildew; Bdv2to barley yellow dwarf virus; and Wsm1 to wheat streakmosaic virus .Viable wheat\u2013wheatgrass hybrids were first obtained byN.V. Tsitsin in 1930\u20131933. He crossed diploid, tetraploid,and hexaploid wheats to Ag. elongatum and Ag. glaucum and obtained octoploid forms of perennialand ratooning wheats known as intermediate wheat\u2013wheatgrasshybrids, IWWHs . Experiments on wheat hybridization to plants ofthe Thinopyrum genus were also carried out in the UnitedStates, Germany, Canada, and China. Various hybrid formswere obtained and annotated: partial amphiploids; highproteinaddition, substitution, and translocation lines andforms resistant to barley yellow dwarf virus, wheat streakmosaic virus, powdery mildew, yellow rust, brown rust,and stem rust .Partial wheat\u2013wheatgrass amphidiploids are used internationallyfor transferring valuable traits to commonwheat . In Russiatwo groups of common wheat cultivars resistant to fungalpests have been raised via IWWHs at the AgriculturalResearch Institute of the South-East and the Samara ResearchInstitute of Agriculture. In their genomes, wheatchromosome 6D is replaced by chromosome 6Agi fromwheatgrass Th. intermedium. Chromosomes 6Agi1 and6Agi2 are not identical, as they show different C bandingpatterns in Giemsa staining . In theformer case, 6Agi1 was inherited from substitution lineS29-Agro139-M2-2, obtained by crossing spring commonwheat Saratovskaya 29 to IWWH 139, and fromcv. Mnogoletka2. Then wheatgrass addition chromosomesrecombined with each other . The cultivarsraised in Samara inherited wheatgrass chromosome6Agi2 from substitution line Agis 1, obtained by crossingS29 to IWWH 644 .Since 1984, when Tulaykovskaya 5 was enlisted to theState Register of Selection Achievements, varieties withwheatgrass chromosome introgression bred in Samararetain their resistance to brown rust and powdery mildewin various ecogeographical regions of Russia . It has been shown thatthe Lr genes on chromosome 6Agi2 are not allelic to thegenes Lr9, Lr19, Lr24, Lr29, or Lr47, and the type of responseto inoculation with Puccinia triticina Eriks. isolatesconfirms their not being allelic to Lr19 or Lr38 . Testing of F2 and F3 hybrids of susceptiblevarieties with Tulaykovskaya 10 for brown rust resistanceshows that chromosome 6Agi2 houses a locus for resistanceto the West Siberian brown rust race .However, the copy number of resistance genes on 6Agi2is still unknown. The loci have not been mapped on thechromosome either.Molecular and cytogenetic markers are designed fordetectionof wheatgrass genetic material in the commonwheat genome . There are molecularmarkers specific to the Th. intermedium genome: simple sequencerepeats (SSRs) , markers designed on the base of expressed sequences (ESTs), and specificlocus amplified fragments (SLAFs) .There are several RFLP , SCAR, and ISSR  markersfor Pseudoroegneria spicata (St genome), designedfor identification of particular chromosomes of the St genome.The correspondence of wheatgrass chromosomes tohomoelogical common wheat groups is tested with uniquegene markers based on PCR (PLUG markers)  and SNP markers . Salina et al. (2016) designed markersspecific to the long and short arms of Th. intermediumchromosome 6Agi2Varieties bred in Samara are used in Russian breedingprograms . Thegoal of this work was to obtain breeding material with introgressedwheatgrass chromosome, test its commerciallysignificant indices, and investigate the transfer of Th. intermediumchromosome 6Agi2 present in cv. Tulaykovskaya10 by the example of a hybrid population with wheatcultivar Saratovskaya 29, which is a gold standard of grainquality. DNA markers specific to the long and short armsof 6Agi2 and genomic in situ hybridization (GISH) wereused to identify the chromosome.Plants. Experiments were conducted with spring commonwheat varieties Saratovskaya 29 (S29) and Tulaykovskaya10 (T10) and with their hybrids S29 \u00d7 T10  and T10 \u00d7 S29 (generations F2\u2013F6). The hybridgenerations were obtained by self-pollination of F1 hybrids.Varieties S29 and T10 belong to the mid-season group.Saratovskaya 29 is highly susceptible to leaf diseases. Tulaykovskaya10 is immune to brown leaf rust and mediumsensitiveto powdery mildew (https://samniish.ru/pshenicamyagkaya-yarovaya-sort-tulajkovskaya-10.html).Hybrids S29 \u00d7 T10, generations F2 and F3, and T10 \u00d7 S29,generations F2, F3 and F5, were grown in a hydroponicgreenhouse of the Laboratory of Artificial Plant Growth,Institute of Cytology and Genetics, Novosibirsk, in theautumn of 2017 and in the springs of 2019 and 2020,respectively. The temperature schedule was 22 \u00b0C in thedaytime and 16 \u00b0C at night. The light/dark schedule was16:8 h. Hybrid generations T10 \u00d7 S29 F4 and F6 were grownin the field in the Moshkovo raion of the Novosibirsk oblastin the summers of 2018 and 2020, respectively; localitycoordinates 55.14\u00b0 N and 83.63\u00b0 E.Fluorescence in situ hybridization (FISH). Mitoticchromosome slides for FISH were prepared as in Ivanova etal. (2019). Use was made of the Aegilops tauschii pAet6- 09probe specific to chromosome centromeric repeatsofrice, wheat, rye, and barley  andwheatgrass genomic DNA isolated from Th. intermediumplants. A DNA sample of the pAet6-09 repeat was kindlyprovided by Dr. A. Lukaszewski . All slides were examinedunderan Axio Imager M1 microscope .Images were captured with a ProgRes MF camera in the Shared Access Center for MicroscopyAnalysis of Biologic Objects, Siberian Branch ofthe RAS, and processed with Adobe Photoshop CS2.Plant DNA isolation. DNA was isolated from youngleaves of hybrids and control plants with a Genomic DNAPurification Kit  accordingto manufacturer\u2019s recommendations.PCR analysis. DNA samples were analyzed with primersMF2/MR1r2 (amplicon size 347 bp) to the long armof chromosome 6Agi2L of Th. intermedium, Te6HS476(amplicon size 200 bp) to the short arm of chromosome6Agi2S of Th. intermedium, and MF2/MR4 (ampliconsize 328 bp) to the long arm of chromosome 6DL. Theprimershad been designed at the Laboratory of Plant MolecularGenetics and Cytogenetics, Institute of Cytologyand Genetics . PCR was carried outin a Bio-Rad T-100 Thermal Cycler. The products wereresolved in 1.5 % agarose gel with ethidium bromide andvisualized with a Gel Doc XR+ gel documentation system.Assessment of commercially valuable traits. TheT10 \u00d7 S29 F4 progeny selected with molecular markers wastested for resistance to brown rust Puccinia triticina Eriks.and stem rust P. graminis Pers. in field experiments in2018. The F6 progeny selected by molecular cytologicalanalysis was tested for resistance to brown rust P. triticinaEriks., stem rust P. graminis Pers., and yellow rustP. glumarum Eriks.et Henn. in the field in 2020. The followingparameterswere recorded in generation F6 selectedby molecular and cytological methods in the field in 2000:the sprouting\u2013flowering interval, plant height, productivetillering, main spike length, number of spikelets in the mainspike, number of grains in the main spike, grain weight ofthe main spike, number of grains per spikelet in the mainspike, grain number per plant, grain weight per plant,1000 grain weight, and contents of protein and gluten inthe grain. Grains were sown on May 9, 2020, in plots of70 cm in width, 15 grains per row, and 25-cm intervalsbetween rowsThe degree of injury by fungal pests was assessed accordingto the CIMMYT scale .The contents of protein and gluten were measured with aninfrared OmegAnalyzer G . The timefrom the mass-scale appearance of sprouts till the first appearanceof yellow anthers in middle spikelets of spikeswas taken to be the sprouting\u2013flowering interval. Floweringdates were recorded in individual spikes. The significanceof differences between two mean values of two sampleswas assessed by Student\u2019s t testIdentification of wheatgrass chromosome 6Agi2in generations F2\u20133 of the S29 \u00d7 T10 and T10 \u00d7 S29 hybridswith chromosome-specific primersChromosomes 6Agi2 of wheatgrass and 6D of wheat werepresent in the F1 of S29 \u00d7 T10 and T10 \u00d7 S29 in the univalentstate. Therefore, their presence or absence in DNAsamples from generation F2 was tested by PCR with primersspecific to the wheatgrass chromosome. We tested 116and 45 DNA samples from F2 S29 \u00d7 T10 and T10 \u00d7 S29,respectively, and found samples with the absence of amplificationwith two primer pairs for the short and longarms of chromosome 6Agi2 and with amplification of themarker to chromosome 6D. Thus, there were no 6Agi2/6Dsubstitution in these samples, designated as wheat (w) type.The presence of chromosome 6D was also proven insamples with amplification of markers to either long orshort arm, being indicative of the presence of telocentrics. Altogether, 12 telocentrics forthe long arm and 29 telocentrics for the short arm were detected in samples of generations F2 and F3, and the ratioof telocentrics for the short and long arm depended significantlyon the cross direction. Telocentrics for the long armwere very rare in the T10 \u00d7 S29 cross.The presence of amplification fragments with two markersto the short and long arms pointed to the presenceof the whole chromosome 6Agi2. With regard to the presenceor absence of chromosome 6D, we suggest eitherfull 6Agi2/6D substitution (Ag type) or the heterozygousstate of the chromosome in the samples (H type).For further analysis, plants with amplification of markersto the short and long arm of the wheatgrass chromosomewere selected.Karyotyping of generation F5 of T10 \u00d7 S29 hybridsTo verify the presence of one or two wheatgrass chromosomesin chromosome sets and to confirm stable inheritanceof the substitution, we performed GISH of mitotic chromosomesat various self-pollination stages. The analysisof plants bearing substitutions according to PCR revealed42 chromosomes, of which two were whole wheatgrasschromosomes . Their long arms housed a largesubtelomeric heterochromatin block, which is consistentwith the locations of Giemsa C bands on chromosome6Agi2 in Tulaykovskaya 10 . Thecentromere-specific pAet6\u201109 repeat located on wheatgrasschromosomes showed weak signals, to demonstrate thepoor hybridization of the repeat to centromeric DNA ofwheatgrass chromosomes.In situ hybridization confirmed the stable inheritance ofthe 6Agi2/6D substitution through generations.Commercially valuable traitsin T10 \u00d7 S29 generations F5 and F6Tulaykovskaya 10 is present in the pedigrees of manymoderncommon wheat varieties. Its use in the breedingof new forms is based on its locus for brown leaf rustresistance, mapped on wheatgrass chromosome 6Agi2. Inspite of the replacement of chromosome 6D by alien chromosome6Agi2, the variety shows high grain yield, droughttolerance,and good baking quality (https://samniish.ru/pshenica-myagkaya-yarovaya-sort-tulajkovskaya-10.html).Three lines were raised from F4 plants of T10 \u00d7 S29 withidentified wheatgrass chromosomes: 33-2, 34-1, and 35-45.Analysis of the performance of T10, S29, and T10 \u00d7 S29F5 lines grown in a hydroponic greenhouse showed thatall the lines significantly outperformed T10 in all indices(Table 2). As compared to S29, the lines did not differ inproductive tillering; lines 34-1 an 35-45 did not differ ingrain number per plant or grain weight per plant; and inline 33-2, these indices were significantly lower. None ofthe lines outperformed S29 in 1000 grain weight; this indexwas significantly lower.We selected the most productive plants of generation F5of line 35-45 to analyze performance indices and the durationof the sprouting\u2013flowering interval in plants grown inthe field in 2020. Thus, daughter line 49-14 was selectedfrom the chosen segregating line 35-45.Phenological observations revealed the shortest sprouting\u2013flowering interval in line 49-14 (50.6 days), and in S29and T10 it was one day longer. The flowering durations inthe main spikes of individual plants were 11 days in 49-14,10 days in T10, and 9 days in S29.Comparison of performance indices in 49-14, S29, andT10 revealed no difference in main spike length, grainweight in the main spike, grain weight per plant, or grainnumber per plant (Table 3). Plants of line 49-14 weresignificantly taller than T10 but did not differ in height from S29. Productive tillering and main spike density in49-14 showed significant ( p \u2264 0.05) differences from thecultivars. The number of grains in the main spike in 49-14was significantly greater than in S29 ( \u0440 \u2264 0.001) or T10( \u0440 \u2264 0.05).The number of grains per spikelet in the main spike ofline 49-14 was significantly higher than in S29 ( p \u2264 0.001)or T10 ( p \u2264 0.05). Line 49-14 set 3.77 \u00b1 0.1 grains perspikelet on the average, and this trait varied among individualplants from 2.93 to 4.62 . The1000 grain weights in line 49-14 and T10 were significantly( \u0440 \u2264 0.001) lower than in S29.Grain quality analysis showed that S29, T10, and 49- 14had high contents of protein and gluten (see Table 3),characteristic of strong wheats. Grain quality in 49-14 wascomparable with S29 and T10.Screening of generations F4 and F6 of the T10 \u00d7 S29 crossfor resistance to fungal pathogensThe resistance of plants to brown rust and stem rust agentswas tested in the field in 2018 and 2020. Field resistanceto powdery mildew was not tested in those years, becauseweather conditions were unfavorable for the agent, asseen from the fact that the susceptible variety S29 wasnot injured.In tests of the resistance to the Siberian population ofthe brown rust agent P. triticina conducted in 2018, S29demonstrated the S (susceptibility) type of response, scoring4 with about 100 % damage of leaf surface .Tulaykovskaya 10 and F4 plants of T10 \u00d7 S29 showed theimmune type without P. triticina pustules .The hybrids tested and parental varieties produced aspecific response to stem rust. Plants of S29, T10, andF4 T10 \u00d7 S29 showed generally the immune response exceptfor a single case. One of the F4 plants showed a specific typeof interaction with the pathogen: occasional uredial pustuleswithout chlorosis (5S) . In practice, thedetected local but pronounced syndrome is interpreted asa sign of a rare virulent fungus race in the local population. As reported by Skolotneva et al.(2020), the stem rust population in the Novosibirsk oblastis highly heterogeneous, as it is formed by southern andwestern migrants.No signs of fungal diseases were detected in plants ofthe cultivars and line 49-14 at the stages of tillering andflowering in the field in 2020. Tests for plant resistanceto the brown rust population at the milky ripeness stageshowed type S (susceptibility) response in S29 plants, score 4 with about 100 % leaf damage, whereasT10 and 49-14 demonstrated the immune response with noP. triticina pustules .At the milky ripeness stage, on August 2\u20135, the start ofdamage of S29, T10, and 49-14 by the yellow rust agentP. striiformis was noted. The percentage of leaf area injury in S29 was 50 to 75 , corresponding to mediumsusceptibility (MS).Plants of T10 and 49-14 showed medium resistance(MR) and medium susceptibility (MS) to the yellow rustagent. The percentage of leaf area injury was 5 to 40, withchlorotic zones .No damage by stem rust was seen in plants of S29, T10,or 49-14 in the summer of 2020.Thus, the results of screening for resistance to a varietyof plant pathogens conducted in the field in different yearsindicate that chromosome 6Agi2 retains the immunity ofplants to the West Siberian brown rust population and immunityto dominant stem rust races. It also supports themedium resistant and medium susceptible types of responseto yellow rust agents.Breeding line 49-14 (2n = 42) was isolated from generationF5 of intervarietal hybrids T10 \u00d7 S29, with introgressionof a pair of wheatgrass chromosomes 6Agi2. It showshigh performance indices and immunity to West Siberianpopulationsof brown rust agents. The response of 49-14plants to the yellow rust agent varies from medium resistanceto medium susceptibility,probably because of thedifference in aggressiveness among the agent races. Stemrust injury was noted in only one plant and was interpretedas immunity to dominant stem rust races.Previously, it was demonstrated that the genetic materialof chromosome 6Agi2 in common wheat varietiesTulaykovskaya 5, Tulaykovskaya 10, Tulaykovskaya zolotistaya,Tulaykovskaya 100, and Volgouralskaya retains theresistance to brown rust populations typical of the Lowerand Middle Volga regions, Central and Ural regions, andWest Siberia . The damageof Tulaykovskaya 10 by brown rust in infection nurseriesof the Central Chernozem region reached 22 %, and thevariety was assigned to group II of epidemic resistance(moderately resistant ER II) . In Tatarstan,the damage of Tulaykovskaya 10 by stem rust was assessedas 5\u201310 % on the average, and the damage by powdery mildewscored 6; the type of response to brown rust remainedimmune . The susceptibility ofT10 to the powdery mildew population of the West Siberianregion was assessed as resistance. A genome-wideassociation search (GWAS) mapped the Pm6Agi2 gene onthe long arm of wheatgrass chromosome 6Agi2, and thisgene imparts resistance to the powdery mildew agent . In experiments in the Middle Volga region,T10 showed immunity to brown rust and medium resistance(20 % injury) to stem rust, yellow rust, and powdery mildew. Thus, T10 retains its immunity tobrown rust populations in various ecogeographical regions.It is medium susceptible to stem and yellow rusts but showsdiverse responses to the powdery mildew agent.The substitution of wheatgrass chromosome 6Agi2 for6D does not impair grain yield, grain quality, or droughttolerance ,although in some cases of using T10 as a resistance genedonor, plants with lower productive tillering and 1000 grainweight appeared among the offspring with chromosome6Agi2 . The contents of protein andgluten in line 49-14 were about the same as in S29 orT10, corresponding to the grain quality of strong wheats. Line 49-14 lagged behindthe parental varieties in productive tillering (S29), numberof spikelets in the main spike (T10), and 1000 grainweight (S29). In spite of lower productive tillering, fewerspikelets in the main spike, and lower 1000 grain weight,the indices grain weight per plant and grain number perplant in line 49-14 did not differ significantly from theparental varieties owing to the significantly higher grainnumber per spikelet in the main spike of 49-14 than inS29 ( \u0440 \u2264 0.001) or T10 ( p \u2264 0.05). Plants of 49-14 set3.77 \u00b1 0.1 grains per spikelet, the range of variation inindividual plants being 2.93\u20134.62, and up to 6 grains wereset in spikelets of the middle spike part. Spikelets werefan-shaped . This shape is a specific sign ofmultiflowered spikelets in wheat .Although common wheat has multiflowered spikelets,most of them set two or three grains. As the potential offorming more grains in wheat exceeds the actual yield byfar, many studies are dedicated to seeking tools to controlthis process. The genetic and physiological groundsof breeding for more grains in spikes and spikelets and,ultimately, more grains per unit area are extensively investigated. Analysis of thereproductive developmental stages of spikes, spikelets,florets, and grains, as well as of their genetic regulation, isthe best way to understand the formation of the trait \u2018grainnumber and spike fertility\u2019. The \u2018grain number per spikelet\u2019trait depends on the initiation of floret primordia, then onfloret survival at the next stage, and then on their efficientpollination. Normally, up to 12 floret primordia form atthe white anther stage, but later up to 60 % of the floretsmay remain underdeveloped . Thisapplies especially to apical (uppermost) florets of a spikelet.As reported by Kuperman (1969), the growth rates ofthe two lowest and upper floret apices are nonuniform atorganogenesis stage V; a spikelet may have up to five, lessoften, to seven florets. Lower florets very quickly formprimordia of generative organs, stamens, and the pistil.A delay in organ formation is observed in the third and,particularly, fourth, fifth, and subsequent florets. Pistilsmost often remain underdeveloped in the uppermost florets.Chromosomes 4A, 5A, 6A, 7A, 2B, 5B, 7B, and 7D bear QTLs responsible for the trait \u2018number of floret primordiaper spikelet\u2019 . Also, the correlation andcluster analyses performed in the same study infer thatthe number of grains per spikelet does not depend on themaximum number of floret primordia per spikelet . Hence, the number of grains in a spikelet isdetermined by the fertility of each floret .A QTL responsible for greater numbers of grains perspikelet was detected on the long arm of chromosome 2Ain GWAS of European common wheat varieties . Further studies of this locus mapped the GrainNumber Increase 1 (GNI1) gene, encoding a transcriptionfactor with the HDZip1 homeodomain. Its mutationcontributes much to greater numbers of fertile florets dueto upper florets of the spikelet . Supposedly,GNI1 was formed by gene duplication in wheatevolution, and its mutations were selected in domestication,as they increased the number of fertile florets, and,consequently, grains. Transcription factor ARGONAUTE1d(AGO1d ) also affects the grain number in the spikes ofcommon and durum wheats . AGO1dis important for the development of anthers and pollenat early developmental stages of wheat. Its malfunctionshortens the spike, reduces anther size, decreases pollenfertility, and thereby decreases the number of grains in thespike .The manifestation of traits in a plant is cumulatively affectedby the genotype, ambient conditions, and farmingtechniques. All these factors greatly influence quantitativetraits, including yield components . The day/night regime and solarspectrum are particularly important ambient factors atorganogenesis stages V and VI . Lowerintensities of the red and infrared radiation reduce thenumber of fertile florets, number of grains per plant, and1000 grain weight . The combinationsof environmental factors required for each developmentalstage stem from the conditions under which the species,varieties, and cultivars formed. With regard to their physiologicaldevelopmental features, cultivars S29 and T10belong to the Volga steppe and forest-steppe agroecologicalgroups, respectively, or morphophysiological type II  (https://samniish.ru/yarovaya_myagkaya_pshenica.html). Such varieties utilize mainly winter andearly spring precipitation in regions with water shortagein the second half of summer; that is, they are tolerant ofsummer drought. Cultivars bred in West Siberia belongto morphophysiological type V. The ecotype of Siberianforest-steppe wheats is determined by the climate: cold anddry April, May, and the first half of June; relatively ampleprecipitation in the second half of summer (July), and coldtemperatures in August. The delay in organogenesis stage Vallows much better use of late summer precipitation forthe formation of large spikes and multiflowered spikelets.Owing to developmental physiological features and highdrought tolerance, varieties of morphophysiological type IIcan be grown in steppe and forest-steppe regions of WestSiberia . Thus, the genotypes of S29 andT10 are environmentally flexible. In the climate of WestSiberian forest-steppe, they synchronize the metamericgrowth of spikelets to develop four, five, or more normalflorets in a spikelet.The genetic material of crested wheatgrass Agropyroncristatum is also beneficial for yield components. Additionlines with chromosome 6P of Ag. cristatum and,particularly, substitution lines 6P/6D show high productivetillering and significantly greater grain numbers inspikes and spikelets: up to 4.5 grains per spikelet . It has been inferred thatchromosome 6P houses genes controlling the numbers offlorets and grains in a spike and spikelet .Conceivably, chromosome 6Agi2 of Th. intermedium bearsgene(s) controlling the synchronous metameric growth ofspikelets in T10, whereas the additive manifestation of thetrait \u2018grain number per spikelet\u2019 is observed in line 49-14(T10 \u00d7 S29), where up to six normal florets develop ina spikeletSakuma et al., 2019) are means for improvingwheat grain yield.The authors declare no conflict of interest.Arbuzova V.S., Dobrovolskaya O.B., Martinek P., Chumanova E.V.,Efremova T.T. 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+{"text": "Wart (a disease caused by Synchytrium endobioticum) and golden cyst potato nematode (Globodera rostochiensis),which parasitize the roots of the host plant, cause signif icant damage to potato crop. Both of these diseasefactors are quarantined in the Russian Federation, and each registered variety is tested for resistance to their mostcommon races and pathotypes. The main method of opposing such diseases is by the development of resistant varieties.An important step in this process is the selection of resistant genotypes from the population and the estimationof the resistance of hybrids obtained by crosses during the breeding process. Conducting a permanent phenotypicevaluation is associated with diff iculties, for example, it is not always possible to work with pathogens, and phenotypicevaluation is very costly and time consuming. However, the use of DNA markers linked to resistance genes cansignif icantly speed up and reduce the cost of the breeding process. The aim of the study was to screen the GenAgropotato collection of ICG SB RAS using known diagnostic PCR markers linked to golden potato cyst nematode and wartresistance. Genotyping was carried out on 73 potato samples using three DNA markers 57R, CP113, Gro1-4 associatedwith nematode resistance and one marker, NL25, associated with wart resistance. The genotyping data were comparedwith the data on the resistance of the collection samples. Only the 57R marker had a high level of correlation  between resistance and the presence of a diagnostic fragment. The diagnosticeff iciency of the 57R marker was 86.11 %. This marker can be successfully used for screening a collection, searchingfor resistant genotypes and marker-assisted selection. The other markers showed a low correlation between the presenceof the DNA marker and resistance. The diagnostic eff iciency of the CP113 marker was only 44.44 %. Spearman\u2019scorrelation coeff icient  did not show signif icant correlation betweenresistance and the DNA marker. The diagnostic eff iciency of the NL25 marker was 61.11 %. No signif icant correlationwas found between the NL25 marker and resistance . The use of thesemarkers for the search for resistant samples is not advisable. Potato is one of the most important crops in the world andis the world\u2019s fifth largest staple food crop by volume . One of the possible reasons fora decrease in yield is the damage of potatoes by various factors.Especially dangerous for potatoes are golden potato cystnematode (Globodera rostochiensis) and potato wart (pathogen\u2013 Synchytrium endobioticum). They are quarantined in theRussian Federation. Data on resistance to G. rostochiensis andS. endobioticum are required when registering a potato varietyin the State Register of Selection Achievements Authorizedfor Use .Potato cyst nematode (PCN) can cause significant damageto the potato yield, which can reach 80\u201390 % . Today, 5 pathotypes of this pestare known in the world: Ro1, Ro2, Ro3, Ro4, Ro5 , while in Russia only the Ro1pathotype of PCN has been detected at the moment .Potato wart affects from 35  to 100 % of the yield. There are 43 wart pathogensin Europe today . Only a few varietiesaffected by this disease are registered in the State Registerof Selection Achievements .One of the main methods of dealing with these pests isthe development of resistant potato varieties. Accordingly,it is important to detect genes responsible for resistance toPCN, study their heritability, develop DNA markers linkedto these genes, and use genes in breeding in marker-assistedselection schemes.The potato has 7 loci of resistance to PCN on chromosomesIII ), V , H1 ,GroV1 ), VII ), X ),XI ). Four loci  provide partial resistance, while three others  give high resistance toone or more pathotypes . DNA markers havemade it possible to identify complex loci containing severalR-genes, including a locus containing two genes for PCN resistance, which was identified on chromosome Vin two different potato species .The H1 resistance gene is introgressed into breeding varietiesfrom Solanum tuberosum ssp. andigenum and S. vernei. This gene is dominant anddetermines resistance to pathotypes Ro1 and Ro4 of G. rostochiensis; according to other data, it determines resistanceto pathotypes Ro5 and Ro6 . This gene is located at the distal part of thelong arm of the V chromosome  and encodes the CC-NBS-LRR protein (coiledcoil/nucleotide-binding/leucine-rich repeat). The H1 gene isthe only nematode resistance gene for which Flora\u2019s geneto-gene interaction concept has been validated by classicalgenetic analysis . The H1 resistance gene corresponded to theAvr gene of golden potato cyst nematode G. rostochiensis.The GroV1 gene originates from the wild potato speciesS. vernei, is linked to the H1 locus ,and is responsible for resistance to the Ro1 pathotype ofG. rostochiensis .The Gro1 locus is localized on chromosome VII andcontains a family of genes Gro1-1, Gro1-2, Gro1-3, Gro1-4,Gro1-5, Gro1-6, Gro1-8, Gro1-10, Gro1-11, Gro1-12 andGro1-14, as well as a number of pseudogenes . J. Paal andcolleagues showed that the Gro1-4 gene is a monogenicdominant gene responsible for resistance to the Ro1 pathotypeof G. rostochiensis and encodes a protein belonging to theTIR-NB-LRR class of proteins. Gro1-4 was introduced into S. tuberosum from the wild potato S. spegazzinii .A number of loci of quantitative traits associated with resistanceto cyst nematodes were mapped in the potato genome:Gro1.2, Gro1.3, and Gro1.4 determining resistance to G. rostochiensiswere localized on chromosomes X, XI, and III. Inthis case, S. spegazzinii was the source of resistance .The Grp1 locus provides a broad spectrum of resistance toboth cyst nematodes G. rostochiensis and G. pallida. It hasbeen mapped to chromosome V  and determines resistance to the Ro5 pathotypeof G. rostochiensis .A significant number of diagnostic DNA markers have beendeveloped for the H1 gene. Among them are markers CD78, TG689 , N146, N195 , CP113 , TG689/TG689indel12 , 239E4left , EM15 (repulsion) and CMI (coupling) , 57R . Markers have also been designed forother genes and QTLs. For example, markers TG69 , SCAR-U14, and SCAR-X02 have been developedfor the GroV1 gene ; markers CP56 and St3.3.2 , CP56, CP51(c), GP516(c) were selectedfor the Gro1 locus . MarkersGro1-4  and Gro1-4-1  were designed forthe Gro1-4 gene. For Grp1-QTL, markers GP21 and GP179, TG432  have been developed.The TG63 marker was selected for Gro1.2-QTL . Markers Ssp75 and TG30 have been developedfor Gro1.3-QTL . The Ssp8 marker wasdesigned for Gro1.4-QTL .A number of genes for resistance to wart (S. endobioticum)have been found in potatoes. These are the followinggenes: Sen1, located on the XI chromosome ; Sen1- 4 mapped to chromosome IV ; locus Sen18-IX, located on chromosome IX; locusSen2/6/18- I, located on chromosome I ;locus Xla-TNL found on chromosome XI ; the Sen2 locus mapped to chromosome XI ; the Sen3 locus was mapped on chromosome XIin the same region as the Sen1 gene ; the authors suggested that Sen3 could be either a Sen1paralogue from the same cluster or an allelic variant of theSen1 gene.QTLs responsible for resistance to races 1, 2, 6 and 18 ofwart are found on other chromosomes: chromosome I (torace 2), chromosome II , chromosome VI , chromosome VII , chromosomeVIII , chromosome X , chromosome XI  .J.E. Obidiegwu and colleagues also found additional wartresistance loci on chromosomes I, IV, X, XI, and XII thatwere less influential than the main genes . Minor QTLs located on the chromosome X furtheraffect resistance to race 18 of wart .The Sen1 and Sen1-4 genes determine the resistance torace 1 of the potato wart pathogen; in both cases, resistanceis determined by the dominant alleles of the genes. The Sen1gene is located at the distal part of the long arm of chromosomeXI . However,it should be noted that J.E. Obidiegwu et al. (2015), usinggenome-wide association studies (GWAS), identified theSen1/ RSe-XIa multi-allelic locus on potato chromosome XIas the main factor of resistance to four S. endobioticum races . The Sen1-4gene is located on the long arm of chromosome IV at a distanceof 5 cM from the centromere .The Xla-TNL locus on potato chromosome XI is linked toresistance to races 18 and 6 and can be considered as one ofthe main factors of wart resistance .The Sen2 locus is mapped to chromosome XI and isa dominant monogenic locus that provides a high level ofresistance to eight races of S. endobioticum simultaneously:1 (D1), 2 (G1), 6 (O1), 8 (F1), 18 (T1), 2 (Ch1), 3 (M1) and39 (P1). The genetic and physical distances between the Sen1and Sen2 loci were indirectly estimated at 63 cM and 32 Mbp,respectively .Sen3 is a dominant monogenic locus of resistance to races 2,6, and 18 . Locus Sen18-IX (chromosomeIX) determines resistance to race 18 S. endobioticum,and locus Sen2/6/18-I (chromosome I) to races 2, 6, and 18.A. Ballvora et al. (2011) note that resistances to races 2, 6and 18 correlate with each other, but are inherited regardlessof resistance to race 1.Several markers have been developed to detect the dominantallele of the Sen1 gene: CP58, GP125 ,NL25 , Sti046, St_At5g16710, GP125 and GP259 . Also, using a genome-wide association studies,a haplotype-specific marker PotVar0067008 associated withSen1 was identified .To identify the Sen18-IX locus, markers GP129, GP101and STM3023b can be used. The Sen2/6/18-I locus can bediagnosed using markers STM2030, SC176, GP192, GP124,and GP194 . Markers Kc8103 and RK36,located on chromosome XI and linked to the Xla-TNL locus,have shown potential diagnostic value in determining resistanceto races 18 and 6 of S. endobioticum . Three markers, 5450_3, 2502_1, and 2502_3, linked tothe Sen2 locus were developed . It is possibleto use the markers chr11_1259552 and chr11_1772869to detect Sen3 .The aim of the study was to screen the GenAgro potato collectionof the Institute of Cytology and Genetics of the SiberianBranch of the Russian Academy of Sciences (ICG SB RAS)using known diagnostic PCR markers linked to resistance togolden cyst potato nematode and potato wart.Plant material. The research material was the collection ofvarieties and hybrids of potatoes named the \u201cGenAgro\u201d plantcollection of the ICG SB RAS. The collection was representedby 73 varieties and hybrids of potatoes (Solanum tuberosum)(Supplement 1)1. The plants were grown in the field on theterritory of the Michurinsky village, Novosibirsk region, fromMay to August 2017.http://vavilov.elpub.ru/jour/manager/f iles/Suppl_Totsky_Engl.pdfSupplementary Materials are available in the online version of the paper:Field tests were carried out according to the followingscheme: the number of rows for each genotype was two; thenumber of plants in a row \u2013 10; row length \u2013 3 m; distancebetween the rows \u2013 0.75 m; distance between the plants inrows \u2013 0.30 m; planting method \u2013 manually (by hand) on furrows,filling furrows with harrows; landing date is the thirddecade of May.Agrochemical characteristics of the soil: the content ofexchanged potassium 110.00 mg/kg; the amount of exchangedbases 24.19 mg-eq/100 g; hydrolytic acidity 3.23 mg-eq/100 g;exchanged acidity 5.60 mg-eq/100 g; humus content 2.67 %;the content of mobile phosphorus 5.14 mg/kg; the degree ofsaturation with bases (V) 88.20 %.Most of the data on resistance to PCN and potato wart weretaken from references, namely from the database of the StateRegister of Selection Achievements Authorized for Use , and from the EuropeanCultivated Potato Database (https://www.europotato.org/).Some of the samples and hybrids for which there were nopublished data on resistance were evaluated under experimentalconditions. Determination of resistance to PCN wascarried out in accordance with the methodology recommendedby OEPP/EPPO (2006) at the All-Russian Institute of PlantProtection. Potato wart resistance was evaluated accordingto the Glynn\u2013Lemmerzahl method as described in the EPPODiagnostic protocol for S. endobioticum at the Russian Potato Research Center.DNA isolation and PCR analysis. DNA was isolated fromthe skin of potato tubers using the DNeasy Plant Mini kit  according to the protocol. The concentrationand purity of the tested samples were determined using gelelectrophoresis and a Nanodrop 2000 apparatus.Several diagnostic markers most often used in breedingprograms were selected for genotyping (Table 1). These markerswere associated with R-genes that determine resistanceto race 1 of potato wart (S. endobioticum) and Ro1 pathotypeof potato cyst nematode (G. rostochiensis).Two markers, 57R and CP113, associated with the H1 resistancegene, and the Gro1-4 marker, associated with theGro1-4 resistance gene, were selected to identify PCN resistancegenes (see Table 1). The SCAR PCR marker CP113-5\u20192/CP113-3\u20192 was proposed by J. Niew\u00f6hner et al. (1995) basedon the RFLP marker CP113. Amplification of DNA of resistantgenotypes using this marker formed product with a 760 bplength. The 57R marker was proposed by L. Schultz et al.(2012). Amplification of DNA of resistant genotypes formedproduct with a 450 bp length. SCAR PCR marker Gro1-4 wasdeveloped by J. Paal et al. (2004) based on the RFLP markerGro1. Amplification of DNA of resistant genotypes formedproduct with a 602 bp length.The NL25 marker was proposed by R. Hehl et al. (1999)when mapping the Sen1 gene. C.A. Bormann et al. (2004)and C. Gebhardt et al. (2006) used this marker for markerassistedselection (see Table 1). Amplification produces oneor two fragments of 1200 or 1400 bp lenght. The presenceof the dominant Sen1 allele is determined by the presence ofa 1400 bp fragment.PCR was carried out in a 20 \u03bcL reaction mixture containing100 ng of DNA, 67 mM Tris-HCl (pH 8.8), 1.8 mM MgCl2,0.01 % Tween 20, 0.2 mM each dNTP, 0.25 \u03bcM forward andreverse specific primers, 1 unit Taq DNA polymerase.Two types of amplification programs (SSR55 and SSR60)represented the time-temperature profile of PCR. SSR55:(1) first cycle: 94 \u00b0C \u2013 2 min; (2) the next 45 cycles: 94 \u00b0\u0421 \u20131 minute, 55 \u00b0\u0421 \u2013 1 minute and 72 \u00b0\u0421 \u2013 2 minutes; (3) onecycle of 5 minutes at 72 \u00b0C (Gro1-4). SSR60: (1) first cycle:94 \u00b0C \u2013 2 min; (2) the next 45 cycles: 94 \u00b0\u0421 \u2013 1 minute, 60 \u00b0\u0421 \u20131 minute and 72 \u00b0\u0421 \u2013 2 minutes; (3) one cycle of 5 minutesat 72 \u00b0C .The analysis of the obtained PCR products was carriedout by electrophoresis in a 2 % agarose gel. The results weredocumented using a Molecular Imager Gel Doc XR System(BioRad) using UV light.Statistical processing of the data was carried out usingSpearman\u2019s correlation coefficient; for calculations, theSTATISTICA program was used. The diagnostic efficiency,sensitivity, specificity and predictive value were calculatedusing the MedCalc softwarehttps://www.medcalc.org/Diagnostic efficiency was defined as the proportion of correcttest results in the total number of test results, or the sum oftrue positive and true negative test results divided by the totalnumber of test results. The sensitivity was calculated as thenumber of resistant samples identified using a DNA markerdivided by the total number of resistant samples. Specificityis the number of susceptible samples identified by the DNAmarker divided by the total number of susceptible samples.Positive predictive value was defined as the proportion ofcorrect positive diagnostic test results.Among 73 samples selected for genotyping, 35 were resistantto PCN, 37 samples were susceptible, and in one sample,resistance to nematodes was unknown (Table 2). 69 sampleswere resistant to wart, 3 samples were susceptible to disease,the resistance of one sample was unknown (see Table 2).Genotyping of varieties and hybridsusing markers designed for resistance to PCNThe 57R marker is found in 85.7 % of resistant samples, aswell as in 13.5 % of susceptible ones . Some mismatches can be observeddue to the absence of linkage of the 57R marker with the H1resistance gene in a number of samples. The second reason forthe mismatches can be explained by the presence of other resistancegenes in samples that do not carry the 57R marker. Thediagnostic efficiency of the 57R marker, which is expressedas the percentage of true (both positive and negative) testresults to the total number of results obtained, was 86.11 %.The diagnostic sensitivity of the used marker, which shows thenumber of resistant samples identified using the DNA marker divided by the total number of resistant samples, was 85.71 %.The diagnostic specificity, which is the number of susceptiblesamples identified by the DNA marker divided by the totalnumber of susceptible samples, was 86.48 %. The predictivevalue of a positive result, showing the proportion of correctpositive diagnostic test results, was 85.71 %. Calculation ofthe Spearman correlation coefficient  showed a significant correlation betweenresistance and the 57R marker.The CP113 marker is found in only 48.6 % of resistant accessions,while the marker is present in 62.9 % of susceptiblegenotypes .These results can be regarded as the absence of linkage of themarker with the H1 resistance gene in many samples of the potatocollection. The diagnostic efficiency of the CP113 markerwas only 44.44 %. Diagnostic sensitivity was 48.57 %. Diagnosticspecificity accounted for 40.54 %. The predictive valueof a positive result, indicating the probability of resistancepresence if the test shows a positive result when CP113 markerwas used, was equal to 43.58 %. Spearman\u2019s correlation coefficient inthis case showed no significant correlation between resistanceand DNA marker. The use of such a marker when screeninga population to search for resistant samples is not advisable.29 samples were analyzed using the Gro1-4 marker. Thediagnostic fragment was amplified in only 5 samples. Correspondenceof the presence of the marker in the resistantsample was observed only in 1 case out of 5. In other cases,the marker was found in the samples susceptible to the disease.The data obtained show that when screening populationsfor resistance to PCN, it is advisable to use the 57R marker.Genotyping of varieties and hybridsusing markers linked to resistance to potato wart The NL25 marker is found in 62.3 % of resistant samples,however, the marker is present in two of the three susceptiblegenotypes . This can be explainedby the processes of crossing over and by the fact that in a numberof samples the linkage of the marker and the resistancegene is not observed; however, the small number of sensitivesamples does not allow sufficiently assessing the applicabilityof the marker for breeding. The marker is absent in 27 samplesand only in one case we observe the absence of a marker in thesusceptible sample, in the other cases the marker is absent inthe resistant samples. This can be explained by the presence ofanother resistance gene that is not linked to the NL25 marker.The diagnostic efficiency of resistance using the NL25 markerwas 61.11 %. The diagnostic sensitivity turned out to beat 62.31 %. The diagnostic specificity was only 33.33 %.However, the predictive value of a positive result, showingthe proportion of correct positive diagnostic test results, whenusing the NL25 marker was equal to 95.55 %. It should benoted that such results are associated with the fact that the setof samples contained only three sensitive samples, and twoof them showed the presence of the NL25 marker. Spearman\u2019scorrelation coefficient  in such situation showed the absenceof significant correlations.Despite the fact that the NL25 marker is often used inscreening and marker selection, a study in our set of samplesshowed that its use does not guarantee a reliable result.In our study, 13 resistant to golden potato nematode samplesthat had both markers (57R and CP113) linked to the H1nematode resistance gene were found. In addition, there are8 genotypes resistant to nematodes and wart and carryingboth the 57R and CP113 markers linked to the H1 nematoderesistance gene and the NL25 marker linked to the Sen1 wartresistance gene. There is also one sample (Safo) in the populationthat is resistant to wart and nematodes and carries all threemarkers 57R, CP113, Gro1-4, linked to nematode resistance,and marker NL25, linked to wart resistance.The NL25 marker linked to the Sen1 gene, which providesresistance to pathotype 1 of potato wart, is successfully usedin the practice of marker-oriented selection. So, C. Gebhardtand colleagues reported that after screening 17 plants in twofamilies of segregating populations using the NL25 marker, 14 genotypes with the marker were identified. All these plantswere found to be resistant to pathotype 1 S. endobioticum.Some were also resistant to pathotype 2 and/or pathotype 6.The effectiveness of this marker is also reported by O.Y. Antonovaand colleagues who analyzed 98 varieties using theNL25 marker. A diagnostic component was found in 95 studiedwart-resistant varieties, while it was not found in threesusceptible varieties. This shows a high level of correlationbetween the presence or absence of the marker and the resistanceand sensitivity of the genotype to wart, respectively.However, A. Khiutti and colleagues, when screening 52 genotypesusing the NL25 marker, found that 39 samples (bothsensitive and resistant genotypes) had the same nondiagnosticfragment, 12 genotypes did not have amplification of theNL25 marker fragments. Only 5 out of 52 genotypes hada diagnostic fragment indicating the presence of a resistancegene. Four of these five accessions were resistant, but onegenotype was found to be sensitive; most resistant genotypesdid not have a 1400 bp diagnostic fragment predicting a resistantphenotype .Our analysis also did not allow us to speak about the reliabilityof using the NL25 marker for screening resistantvarieties.Using the Gro1-4 marker in a segregating population, C. Gebhardtand colleagues found that all 45 plants carrying thismarker linked to the Gro1 gene were resistant to the Ro1 pathotypeof G. rostochiensis .C. Gebhardt and colleagues in 1993 found in a segregatingpopulation that the CP113 marker is linked to the H1 geneso strongly that it has zero recombination . However, D. Milczarek and colleagues (2011) reportedthat the CP113 marker was amplified for all tested varieties,resistant and sensitive, and was unsuitable for the selectionof resistant clones. A similar picture is observed in our work.The 57R SCAR marker was tested in a mapping population,where it was linked to the H1 locus and nematoderesistance . Later L. Schultzand colleagues reported that they analyzed two independentpopulations of 281 and 122 potato samples with knownresistance/sensitivity using the 57R SCAR marker. Whenscreening the first population, the 57R marker revealed a correspondencebetween genotype and phenotype, 89 out of90 resistant varieties had an allele associated with resistance.Only one resistant variety, in which no marker amplificationwas observed, became an exception. None of the 191 PCNsusceptible varieties had an allele predicting resistance. Thenanother independent population of 122 varieties was screened.All varieties showed complete correspondence between resistanceto G. rostochiensis and the presence/absence of the57R allele, corresponding to the presence of the resistancegene .O.Y. Antonova et al. (2016) identified the 57R markerin 33 (30.3 %) of 109 breeding varieties they studied. Theoverwhelming majority of the varieties with the diagnosed57R fragment were resistant or weakly affected by the nematode.The correspondence between resistance and the presenceof a diagnostic fragment was high \u2013 93.5 %. At the same time,only four genotypes with the Gro1-4 marker were identified:two resistant varieties, one weakly affected variety and onesusceptible. All these four varieties, along with the Gro1-4marker, also possessed the H1 gene markers \u2013 57R, TG689,N146, N195 .In the work of N.S. Klimenko et al. (2017) showed the presenceof the 57R marker in 24 out of 103 samples, while themarker was found in 15 resistant and 2 susceptible samples.It was shown that the correlation between the presence of atleast one marker of the H1 gene and the data on the nematoderesistance of varieties was +0.92 .T.A. Gavrilenko et al. (2018) showed that out of 39 samplesof the studied set of samples, 15 had a dominant allele ofthe H1 gene (based on a number of DNA markers), and twovarieties had dominant alleles of both H1 and Gro1-4 genes.At the same time, none of the markers was identified in theremaining 22 genotypes. Comparison of these results withresistance to G. rostochiensis (pathotype Ro1) showed thatall accessions with H1 gene markers are nematode resistant,while varieties affected by G. rostochiensis did not have thesemarkers . This high correlation shows the reliability of the markers used in the study, which can beused to select resistant samples.It should be noted that the saturation of the genotype withgenes of resistance to the nematode does not affect its economicallyvaluable traits. At the same time, there is a stronglink between the presence of the marker and resistance. So, inthe study of D. Milczarek and colleagues in 2014, the relationshipbetween the presence of markers TG689 and 57R linkedto the H1 gene, which determines resistance to the nematodeG. rostochiensis, and valuable agricultural traits is presented.Clones with these markers had a higher total yield of tubersand total starch yield than clones without markers. There wasno negative association between marker presence and quality.All 347 seedlings obtained after three crosses were genotypedusing both markers and phenotypically evaluated for resistanceto the Ro1 pathotype of G. rostochiensis. Of these, 316 (i. e.91 %) and 325 (94 %) clones were resistant and carried theTG689 or 57R markers .In general, our data on the 57R marker are quite close to theresults described above and confirm the high reliability ofthe work of this marker, which suggests the need to use thismarker when selecting samples resistant to PCN.The authors declare no conflict of interest.Antonova O.Y., Shvachko N.A., Novikova L.Y., Shuvalov O.Y., KostinaL.I., Klimenko N.S., Shuvalova A.R., Gavrilenko T.A. Geneticdiversity of potato varieties bred in Russia and its neighboring countriesbased on the polymorphism of SSR-loci and markers associatedwith resistance R-genes. Russ. J. Genet. Appl. Res. 2017;7(5):489-500. 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Development of a multiplex PCR method for simultaneousdetection of diagnostic DNA markers of five disease and pest resistancegenes in potato. Euphytica. 2011;180:347-355. DOI 10.1007/s10681-011-0381-6.Niew\u00f6hner J., Salamini F., Gebhardt C. Development of PCR assaysdiagnostic for RFLP marker alleles closely linked to alleles Gro1and H1, conferring resistance to the root cyst nematode Globoderarostochiensis in potato. Mol. Breed. 1995;1:65-78. DOI 10.1007/BF01682090.Obidiegwu J.E., Flath K., Gebhardt C. Managing potato wart: a reviewof present research status and future perspective. Theor. Appl. Genet.2014\u037e127(4):763-780. DOI 10.1007/s00122-014-2268-0.Obidiegwu J.E., Sanetomo R., Flath K., Tacke E., Hoferbert H.-R.,Hofmann A., Walkemeier B., Gebhardt C. Genomic architecture ofpotato resistance to Synchytrium endobioticum disentangled usingSSR markers and the 8.3 k SolCAP SNP genotyping array. BMCGenet. 2015;16:38. DOI 10.1186/s12863-015-0195-y.OEPP/EPPO Standards PM 7/28. Diagnostic protocols for regulatedpests: Synchytrium endobioticum. Bulletin OEPP/EPPO Bulletin.2004;34:213-218. DOI 10.1111/j.1365-2338.2004.00722.x.OEPP/EPPO Testing of potato varieties to assess resistance to Globoderarostochiensis and Globodera pallida. Bulletin OEPP/EPPO Bulletin. 2006;36:419-420. DOI 10.1111/j.1365-2338.2006.01032.x.Paal J., Henselewski H., Muth J., Meksem K., Men\u00e9ndez C.M., SalaminiF., Ballvora A., Gebhardt C. Molecular cloning of the potatoGro1-4 gene conferring resistance to pathotype Ro1 of theroot cyst nematode Globodera rostochiensis, based on a candidategene approach. Plant J. 2004;38(2):285-297. DOI 10.1111/j.1365-313X.2004.02047.x.Pajerowska-Mukhtar K., Stich B., Achenbach U., Ballvora A., L\u00fcbeckJ., Strahwald J., Tacke E., Hofferbert H.-R., Ilarionova E., BellinD., Walkemeier B., Basekow R., Kersten B., Gebhardt C. Singlenucleotide polymorphisms in the allene oxide synthase 2 geneare associated with field resistance to late blight in populations oftetraploid potato cultivars. Genetics. 2009;181(3):1115-1127. DOI10.1534/genetics.108.094268.Pineda O., Bonierbale M.W., Plaisted R.L., Brodie B.B., Tanksley S.D.Identification of RFLP markers linked to the H1 gene conferringresistance to the potato cyst nematode Globodera rostochiensis.Genome. 1993;36(1):152-156. DOI 10.1139/g93-019.Plich J., Przetakiewicz J., \u015aliwka J., Flis B., Wasilewicz-Flis I., Zimnoch-Guzowska E. Novel gene Sen2 conferring broad-spectrumresistance to Synchytrium endobioticum mapped to potato chromosomeXI. Theor. Appl. Genet. 2018;131:2321-2331. DOI 10.1007/s00122-018-3154-y.Prodhomme C., Esselink D., Borm T., Visser R.G.F., van Eck H.J.,Vossen J.H. Comparative Subsequence Sets Analysis (CoSSA) isa robust approach to identify haplotype specific SNPs; mappingand pedigree analysis of a potato wart disease resistance gene Sen3.Plant Methods. 2019;15:60. DOI 10.1186/s13007-019-0445-5.Prodhomme C., Peter G.V., Paulo M.J., Visser R.G.F., Vossen J.H.,van Eck J.H. Distribution of P1(D1) wart disease resistance in potatogermplasm and GWAS identification of haplotype-specific SNPmarkers. Theor. Appl. Genet. 2020;133:1859-1871. DOI 10.1007/s00122-020-03559-3.Ramakrishnan A.P., Ritland C.E., Blas Sevillano R.H., Riseman A. Reviewof potato molecular markers to enhance trait selection. Am. J.Potato Res. 2015;92(4):455-472. DOI 10.1007/s12230-015-9455-7.Rouppe van der Voort J., Lindeman W., Folkertsma R., Hutten R., OvermarsH., van der Vossen E., Jacobsen E., Bakker J. A QTL for broadspectrumresistance to cyst nematode species (Globodera spp.) mapsto a resistance gene cluster in potato. Theor. Appl. Genet. 1998;96:654-661. DOI 10.1007/s001220050785.Rouppe van der Voort J., van der Vossen E., Bakker E., Overmars H.,van Zandvoort P., Hutten R., Klein Lankhorst R., Bakker J. Two additiveQTLs conferring broadspectrum resistance in potato to Globoderapallida are localized on resistance gene clusters. Theor. Appl.Genet. 2000;101:1122-1130. DOI 10.1007/s001220051588.Schultz L., Cogan N.\u041e.I., McLean K., Dale M.F.B., Bryan G.J., ForsterJ.W., Slater A.T. Evaluation and implementation of a potentialdiagnostic molecular marker for H1-conferred potato cyst nematoderesistance in potato (Solanum tuberosum L.). Plant Breed. 2012;131:315-321. DOI 10.1111/j.1439-0523.2012.01949.x.Skupinov\u00e1 S., Vejl P., Sedl\u00e1k P., Domk\u00e1\u0159ov\u00e1 J. Segregation of DNAmarkers of potato (Solanum tuberosum ssp. tuberosum L.) resistanceagainst Ro1 pathotype Globodera rostochiensis in selected F1progeny. Rostlinn\u00e1 V\u00fdroba. 2002;48(11):480-485. DOI 10.17221/4399-PSE.State Register of Selection Achievements Authorized for Use for ProductionPurposes. Vol. 1. Plant Varieties . Moscow:Rosinformagrotech Publ., 2019. (in Russian)Toxopeus H.J., Huijsman C.A. Breeding for resistance to potato rooteelworm. I. Preliminary data concerning the inheritance and thenature of resistance. Euphytica. 1953;2(3):180-186. DOI 10.1007/BF00053725."}
+{"text": "Over 600,000 COVID-19 cases, including >7000 deaths reported to MN Dept of Health (MDH) by June 1, 2021. Clinical trials demonstrated high effectiveness of COVID vaccines. We assessed COVID-19 cases among fully vaccinated residents [vaccine breakthrough (VB) cases]. COVID-19 VB cases were MN residents with completed COVID-19 vaccination series \u226514 days prior to symptom onset or positive for SARS-CoV-2 by nucleic acid amplification or antigen test. COVID-19 cases were reported to MDH and COVID-19 vaccinations reported to the MN Immunization Information Connection (MIIC). COVID-19 cases were matched to MIIC to identify VB and interviewed; medical records of hospitalized cases were reviewed. Available VB case specimens underwent whole genome sequencing (WGS) at MDH or collaborating lab.Jan 19 \u2013 June 1, 2021, 2765 VB cases were reported among >2.45 million fully vaccinated residents and 147,445 COVID-19 cases. VB case median (MED) age was 52 y , 83% white, 65% female; MED age of fully vaccinated was 55 y , 77% white, 54% female. Of VB cases, 273 (10%) were hospitalized and 32 (1%) died . 2212 (80%) VB cases were interviewed; 60% reported symptoms; most common were fatigue (53%), rhinorrhea (49%), cough (42%), headache (41%). 35% reported a comorbidity.Of hospitalized VB cases, 120 had completed record reviews. 64 were admitted for COVID-19 related illness  including 27 admitted to ICU . 90% (108) reported a comorbidity, most common being chronic metabolic conditions (46%), obesity (45%), renal disease (31%) and chronic lung disease (26%); 27 were immunocompromised , including immunosuppressive therapy (15), hematological malignancy (9), other cancer (11), and organ transplant recipients (8).Of 604 VB case specimens, 79% were B.1.1.7, 9% B.1.427/429, 3% P.1, and 2% B.1.351; lineage distribution was similar to overall 24,157 MN SARS-CoV2 WGS data.Identified VB cases were 0.1% of those vaccinated and < 2% of total cases reported in the time period. COVID-19 vaccines are an important tool in preventing COVID-19. Additional surveillance, including WGS and case characteristics will be useful to monitor VB.Ruth Lynfield, MD, Nothing to disclose"}
+{"text": "Salmonella enterica serovar Typhi ISP2825, isolated in 1983 from a Chilean patient, is one of the major S. Typhi strains used for research, along with strains Ty2, CT18, and H58. The complete genome sequence of ISP2825, consisting of a 4,774,014-bp circular chromosome, will help us understand typhoid pathogenesis and evolution. Salmonella enterica serovar Typhi is the causative agent of the life-threatening systemic disease typhoid fever, which is a major cause of infection-mediated morbidity and mortality in countries of endemicity. Humans are the only known host of S. Typhi. Despite its narrow host specificity, S. Typhi has remained a highly successful pathogen since its emergence  and extensively drug-resistant (XDR) strains have become the dominant S. Typhi variants  overnight at 37\u00b0C, and its genomic DNA was extracted using the DNeasy blood and tissue kit , without processing additional fragmentation and size selection. The genomic DNA quality and quantity were monitored using a NanoDrop 2000 spectrophotometer  and a Qubit 4 fluorometer (Thermo Fisher Scientific). A genomic DNA library was prepared using a SQK-LSK110 ligation sequencing kit , followed by sequencing with two MinION Flongle flow cells (R9.4.1) using MinKNOW v21.06.0 (ONT). The combined raw reads from two Flongle flow cells were used for base calling using Guppy v5.0.11 (ONT). Fastq files having Q scores of \u22658 were collected, filtered using NanoLyse v1.2.0 (https://github.com/lh3/bwa) and polished sequentially using Racon v1.4.22 (-m 8 -x -6 -g -8 -w 500) (https://github.com/nanoporetech/medaka) (-m r941_min_hac_g507), and Homopolish v0.2.3 (-s bacteria.msh -m R9.4.pkl) (https://github.com/b-brankovics/fasta_tools) was used to set the start position of the polished assembly according to that of S. Typhi CT18 (GenBank accession number GCF_000195995.1) and Ty2 (GCF_000007545.1). The complete genome sequence was annotated using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP)  , Medaka 9.4.pkl) . Fasta_se (PGAP) , and itse (PGAP)  and AMRFe (PGAP) , respecte (PGAP) . Defaulte (PGAP) . OverallS. Typhi ISP2825 has been deposited at GenBank under accession number GCF_019645915.1 or CP080960.1, BioProject accession number PRJNA753482, BioSample accession number SAMN20695325, and Sequence Read Archive (SRA) accession number SRR15411315.The complete genome sequence of"}
+{"text": "Escherichia coli sequence type 1193 (ST1193) is an important cause of multidrug-resistant extraintestinal infections. Here, we report the complete genome sequence of strain AVS0096, isolated from river water in Switzerland in 2020. The genome consists of a chromosome (4.9\u2009Mbp), a multidrug resistance plasmid (101\u2009kb), and two small plasmids. Escherichia coli lineages sequence type 131-H30 (ST131-H30) and ST1193 are major causative agents of fluoroquinolone-resistant E. coli infections in humans  (\u2013blaCTX-M-27-carrying ST1193 isolate (AVS0096) obtained from river water in Switzerland.The pandemic  (ESBLs) \u20136. Where(ESBLs) \u2013, 6, few Enterobacteriaceae enrichment (EE) broth (BD) at 37\u00b0C for 24\u2009h. One loopful of the enrichment broth was spread onto Brilliance ESBL agar (Oxoid) and incubated at 37\u00b0C for 24\u2009h. Matrix-assisted laser desorption ionization\u2013time of flight mass spectrometry  was used for species identification. Susceptibility testing against 13 antimicrobial agents was performed using the disk diffusion method according to CLSI protocols (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/) and LongQC v1.2.0  and 274\u2009Mbp short-read  data using Unicycler v0.4.8 (AVS0096 was isolated in August 2020 from the river Lorze. The water sample 100\u2009ml) was filtered through a 0.45-\u03bcm membrane filter (Millipore). The filter was incubated in 10\u2009ml rotocols  and Pore0\u2009ml was r v0.4.8  and annor v0.4.8 . Resistar v0.4.8  and Plasr v0.4.8  databaseH64 clade, as determined using https://pubmlst.org/ and FimTyper v1.0   from E. coli WP3-W18-CRE-03 (ST1193), isolated from wastewater in Japan. According to the CLSI criteria, AVS0096 showed resistance against azithromycin (macrolide), ampicillin, cefazolin, and cefotaxime (\u03b2-lactams), ciprofloxacin (fluoroquinolone), streptomycin (aminoglycoside), tetracycline, and trimethoprim-sulfamethoxazole, confirming genotypically identified resistances.The complete genome of AVS0096 consisted of a 4,944,762-bp chromosome and the three plasmids pAVS0096-a , pAVS0096-b , and pAVS0096-c . AVS0096 belonged to the ST1193-per v1.0 , and carVS0096-a . A near-CP076344.1 (chromosome), CP076345.1 (plasmid pAVS0096-a), CP076346.1 (plasmid pAVS0096-b), and CP076347.1 (plasmid pAVS0096-c). The raw data were deposited in the NCBI Sequence Read Archive (SRA) under BioSample accession number SAMN19493560 and BioProject accession number PRJNA734472.The complete genome sequence of AVS0096 has been deposited in GenBank under the accession numbers"}
+{"text": "Candidemia is the second most common cause of healthcare-associated bloodstream infections in the US with mortality of approximately 25%. Studies demonstrate lower candidemia mortality with infectious diseases consultation (IDC). We evaluated effects of IDC on mortality and guideline-adherence at our institution to determine if mandatory IDC was warranted.Candida) between 1/1/2016-12/31/2019. Exclusion criteria included age < 19 years, polymicrobial blood culture, or death or hospice within 48 hours. Primary outcome was all-cause 30-day mortality. Secondary outcomes included guideline-adherence and treatment choice. Guideline-adherence was assessed with a modified EQUAL Candida score (Table 1). Descriptive statistics were performed.We retrospectively reviewed adults hospitalized with candidemia  were present in 66 (71.7%) patients and were the most common infection source (N=38 [41.3%]) followed by intra-abdominal (N=23 [25%]). The most isolated species were Candida glabrata (40/94 [42.6%]) and C. albicans/dublienensis (35/94 [37.2%]). 30-day mortality was 21.7%. IDC was performed in 84 (91.3%) cases. Outcomes are in Table 3. Mortality was not different between IDC vs no IDC (18 [21.4%] vs 2 [25%]); other comparisons were numerically different but not significant: repeat blood culture (98.8% vs 87.5%), echocardiography (70.2% vs 50%), CVC removal (91.7% vs 83.3%), and initial treatment echinocandin (67.9% vs 50%). All patients received antifungal therapy. IDC resulted in more ophthalmology consultations . Mean modified EQUAL Candida score was higher with IDC .Of 187 patients reviewed, 92 episodes of candidemia with 94 species of Table 2. Patient CharacteristicsAbbreviations. TPN: total parenteral nutrition, ICU: intensive care unit, AIDS: acquired immunodeficiency syndromeTable 3. OutcomesAbbreviations. NS: non-significant, CVC: central venous catheterIDC was common in candidemic patients and not associated with significant differences in outcomes. Current antimicrobial stewardship and consultation practices at our center do not warrant mandated IDC for candidemia.Trevor C. Van Schooneveld, MD, FACP, BioFire  Involved: Self): Consultant, Scientific Research Study Investigator; Insmed  Involved: Self): Scientific Research Study Investigator; Merck  Involved: Self): Scientific Research Study Investigator; Rebiotix  Involved: Self): Scientific Research Study Investigator"}
+{"text": "Analysis of 16S rRNA databases showed the preferences of Binatota to terrestrial and freshwater ecosystems, hydrocarbon-rich habitats, and sponges, supporting their potential role in mitigating methanol and methane emissions, breakdown of alkanes, and their association with sponges. Our results expand the lists of methylotrophic, aerobic alkane-degrading, and pigment-producing lineages. We also highlight the consistent encountering of incomplete biosynthetic pathways in microbial genomes, a phenomenon necessitating careful assessment when assigning putative functions based on a set-threshold of pathway completion.The recent leveraging of genome-resolved metagenomics has generated an enormous number of genomes from novel uncultured microbial lineages yet left many clades undescribed. Here, we present a global analysis of genomes belonging to Binatota (UBP10), a globally distributed, yet-uncharacterized bacterial phylum. All orders in Binatota encoded the capacity for aerobic methylotrophy using methanol, methylamine, sulfomethanes, and chloromethanes as the substrates. Methylotrophy in Binatota was characterized by order-specific substrate degradation preferences, as well as extensive metabolic versatility, i.e., the utilization of diverse sets of genes, pathways, and combinations to achieve a specific metabolic goal. The genomes also encoded multiple alkane hydroxylases and monooxygenases, potentially enabling growth on a wide range of alkanes and fatty acids. Pigmentation is inferred from a complete pathway for carotenoids  production. Further, the majority of genes involved in bacteriochlorophyll  Distinct strategies are employed for the analysis of the deluge of obtained genomes. Site- or habitat-specific studies focus on spatiotemporal sampling of a single site or habitat of interest. Function-based studies focus on genomes from single or multiple habitats to identify and characterize organisms involved in a specific process, e.g., cellulose degradation  or sulfa efforts , 10. The efforts , as well efforts , 13. As Candidate phylum UBP10 has originally been described as one of the novel lineages recovered from a massive binning effort that reconstructed thousands of genomes from publicly available metagenomic data sets . UBP10 hn\u2009=\u20092), Binatales (n\u2009=\u200948), HRBin30 (n\u2009=\u20097), UBA1149 (n\u2009=\u20099), UBA9968 (n\u2009=\u200934), UBA12105 (n\u2009=\u20091), and UTPRO1 (n\u2009=\u20097), encompassing 12 families and 24 genera  and 22 high-quality  genomes, as defined by MIMAG standards . Binatot4 genera . 16S rRN classes . RDP II-bacteria .10.1128/mBio.00985-21.7TABLE\u00a0S1Table\u00a0S1, XLSX file, 0.02 MB.Binatota genomes used in this study number, their GTDB classification, and the corresponding classification in Silva and RDP databases, the source from which they were obtained, and the calculated Binatota abundances in metagenomes with available contig coverage data. Download Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 10.1128/mBio.00985-21.8TABLE\u00a0S2Table\u00a0S2, XLSX file, 0.02 MB.Sequencing statistics for the genomic bins used in this study. Download Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 10.1128/mBio.00985-21.9TABLE\u00a0S3Table\u00a0S3, XLSX file, 0.02 MB.General genomic features of the studied genomes. Download Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the mno), typically associated with Gram-positive methylotrophic bacteria (Actinobacteria and Bacillus methanolicus)  , was thecyclales , was enccyclales . All latcyclales .mau) was more common, with mauA and mauB enzyme subunits encoded in Binatales, HRBin30, UBA1149, UBA12105, and UTPRO1 (mauC) is the most probable electron acceptor for methylamine dehydrogenase (mau cluster) carried the full complement of genes for methylamine oxidation via the indirect glutamate pathway  involved in the degradation of dimethyl sulfone to MSA with the concomitant release of formaldehyde. Three of these nine genomes also encoded alkane sulfonic acid monooxygenase (ssuD), which will further degrade the MSA to formaldehyde and sulfite. Degradation of DMS via DMS monooxygenase (dmoA) to formaldehyde and sulfide was encountered in 13 genomes . Further, one Binatales genome encoded the dso system (enzyme class [EC]: 1.14.13.245) for DMS oxidation to dimethyl sulfone, which could be further degraded to MSA as explained above , and dimethyl sulfide (DMS). Nine genomes  encoded dimethyl sulfone monooxygenase (S-transferase (dcmA) capable of converting dichloromethane to formaldehyde.One Bin18 genome encoded the specific dehalogenase/glutathione Proteobacteria, Verrucomicrobia, and \u201cCandidatus Methylomirabilis\u201d (NC10)  methanotrophs (bmoA sequences (putative butane monooxygenase gene A) from Actinobacteria  and SAR324 (\u201cCandidatus Lambdaproteobacteria\u201d) (Nocardioides sp. strain CF8 demonstrated its capacity to oxidize short-chain (C2 to C4) hydrocarbons, but not methane, via its CuMMO, and its genome lacked methanol dehydrogenase homologues  3D model (Protein Data Bank ID: 3RGB) revealed a heterotrimeric structure (\u03b13\u03b23\u03b33) with the 7, 2, and 5 alpha helices of the PmoA, PmoB, and PmoC subunits, respectively, as well as the beta sheets characteristic of PmoA and PmoB subunits , thought to coordinate the Cu cofactor, were identified in all TUSC-affiliated and SAR324/Actinobacteria-affiliated Binatota CuMMO sequences (Actinobacteria/SAR324-type Binatota genomes using Methylococcus capsulatus (Bath) PmoB subunit (Protein Data Bank [PDB] ID: 3RGB) predicted the binding pockets for Cu in Binatota sequences (Genes encoding copper membrane monooxygenases (CuMMOs), a family of enzymes that includes particulate methane monooxygenase (pMMO), were identified in orders Bin18 (2/2 genomes) and Binatales (9/48 genomes) (\u201d (NC10) \u201323  \u2013. In addiive site , 25. Phynotrophs  (2 Binatcteria\u201d) , 28  . Memberscteria\u201d) . Previoucteria\u201d) . Binatotmologues . Such dasubunits . Recentlsubunits . There hsubunits \u201333. Regaequences . Modelinequences .10.1128/mBio.00985-21.2FIG\u00a0S1Methylococcus capsulatus (PDB ID: 3RGB). The alignment is showing conserved residues (red highlight). Of particular importance are the three conserved histidine residues His33, His137, and His139 (shown with a blue rectangle), thought to coordinate the Cu cofactor. Numbering follows the Methylococcus capsulatus strain Bath PmoB subunit (PDB: 3RGB). Alignment was created using the ENDscript webserver (http://espript.ibcp.fr/ESPript/ESPript/). (B and C) Predicted Cu methane monooxygenase (PmoABC) 3D structure (grey) from a cluster 2 TUSC-affiliated Binatota genome  and an Actinobacteria/SAR324-affiliated Binatota genome , both superimposed on CuMMO from the model methanotroph Methylococcus capsulatus strain Bath (PDB: 3RGB) (green) with a global model quality estimate of 0.7 and 0.62, respectively, and a quaternary structure quality score of 0.57 and 0.55, respectively. Download FIG\u00a0S1, PDF file, 1.4 MB.(A) Alignment of the PmoB subunit of the 11 copper membrane monooxygenases predicted in Binatota genomes to PmoB from Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 1 oxidation to formaldehyde, formaldehyde oxidation to CO2, and formaldehyde assimilation. Formaldehyde generated by C1 substrates oxidation is subsequently oxidized to formate and eventually CO2. Multiple pathways for formaldehyde oxidation to formate were identified in all Binatota orders , the H4F-linked pathway , the glutathione-independent formaldehyde dehydrogenase (fdhA), and the glutathione-dependent formaldehyde . Also shown is the distribution of the NAD-dependent formate dehydrogenase (EC: 1.17.1.9) (fdh) for formate oxidation. (B) Overview of the pathways for formaldehyde assimilation via the serine cycle (left) and glyoxylate regeneration via the ethylmalonyl-CoA pathway and the glyoxylate shunt (GS) (right). Names of enzymes are shown in red and their distribution in the Binatota genomes from different orders is shown in the heatmap in panel C. glyA, glycine hydroxymethyltransferase [EC: 2.1.2.1]; sgaA, serine-glyoxylate transaminase [EC: 2.6.1.45]; hprA, glycerate dehydrogenase [EC: 1.1.1.29]; gck, glycerate 2-kinase [EC: 2.7.1.165]; ppc, phosphoenolpyruvate carboxylase [EC: 4.1.1.31]; pckA, phosphoenolpyruvate carboxykinase; mdh, malate dehydrogenase [EC: 1.1.1.37]; mtkA/B, malate-CoA ligase [EC: 6.2.1.9]; mcl, malyl-CoA/(S)-citramalyl-CoA lyase [EC: 4.1.3.24 4.1.3.25]; aceA, isocitrate lyase [EC: 4.1.3.1]; aceB, malate synthase [EC: 2.3.3.9]; phbB, acetoacetyl-CoA reductase [EC: 1.1.1.36]; croR, 3-hydroxybutyryl-CoA dehydratase [EC: 4.2.1.55]; ccr, crotonyl-CoA carboxylase/reductase [EC: 1.3.1.85]; epi, methylmalonyl-CoA/ethylmalonyl-CoA epimerase [EC: 5.1.99.1]; ecm, ethylmalonyl-CoA mutase [EC: 5.4.99.63]; mcd, (2S)-methylsuccinyl-CoA dehydrogenase [EC: 1.3.8.12]; mch, 2-methylfumaryl-CoA hydratase [EC: 4.2.1.148]; mut, methylmalonyl-CoA mutase [EC: 5.4.99.2]; mcmA1/A2, methylmalonyl-CoA mutase [EC: 5.4.99.2]. Abbreviations: PEP, phosphoenol pyruvate; OAA, oxaloacetate. Download FIG\u00a0S2, PDF file, 0.2 MB.Formaldehyde oxidation and assimilation capabilities encoded by Binatota genomes. (A) Heatmap of the distribution of formaldehyde oxidation genes in Binatota genomes from different orders. The heatmap colors (as explained in the key) correspond to the percentage of genomes in each order encoding a homologue of the gene in the column header. Shown are the different routes of formaldehyde oxidation, including the (myco)thiol-dependent formaldehyde dehydrogenase Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the Actinobacteria/SAR324-affiliated CuMMO to oxidize C1 to C5 alkanes and C1 to C4 alkenes as described above, some Binatota genomes encoded propane-2-monoxygenase (prmABC), an enzyme mediating propane hydroxylation in the 2-position yielding isopropanol. Several genomes also encoded medium-chain-specific alkane hydroxylases, e.g., homologues of the nonheme iron alkB (ladA homologues (enzyme class [EC]: 1.14.14.28)  known to have a broad substrate specificity for medium-chain-length (C3 to C10) mono- and dihaloalkanes, resulting in the production of their corresponding primary alcohol and haloalcohols, respectively    [EC: 1.1.1.80]; acmA, acetone monooxygenase (methyl acetate-forming) [EC: 1.14.13.226]; acmB, methyl acetate hydrolase [EC: 3.1.1.114]; aldehyde dehydrogenase (NAD+) [EC: 1.2.1.3]; acetaldehyde dehydrogenase (acetylating) [EC: 1.2.1.10]; acdAB, acetate-CoA ligase (ADP-forming) [EC: 6.2.1.13]; acs, acetyl-CoA synthase [EC: 2.3.1.169]; atoAD, acetate-CoA/acetoacetate CoA-transferase [EC: 2.8.3.8 2.8.3.9]; medium-chain acyl-CoA synthetase [EC: 6.2.1.2]; fadD, long-chain acyl-CoA synthetase [EC: 6.2.1.3]; pccA, propionyl-CoA carboxylase alpha chain [EC: 6.4.1.3]; epi, methylmalonyl-CoA/ethylmalonyl-CoA epimerase [EC: 5.1.99.1]; mut, methylmalonyl-CoA mutase [EC: 5.4.99.2]; mcl, malyl-CoA/(S)-citramalyl-CoA lyase [EC: 4.1.3.24 4.1.3.25]; mch, 2-methylfumaryl-CoA hydratase [EC: 4.2.1.148]; mct, 2-methylfumaryl-CoA isomerase [EC: 5.4.1.3]; meh, 3-methylfumaryl-CoA hydratase [EC: 4.2.1.153]; smtAB, succinyl-CoA:(S)-malate-CoA-transferase subunit A [EC: 2.8.3.22]; prpB, methylisocitrate lyase [EC: 4.1.3.30]; prpC, 2-methylcitrate synthase [EC: 2.3.3.5]; prpD, 2-methylcitrate dehydratase [EC: 4.2.1.79]; bcd, butyryl-CoA dehydrogenase [EC: 1.3.8.1]; acd, acyl-CoA dehydrogenase [EC: 1.3.8.7]; paaF, enoyl-CoA hydratase [EC: 4.2.1.17]; crt, enoyl-CoA hydratase [EC: 4.2.1.17]; paaH, 3-hydroxybutyryl-CoA dehydrogenase [EC: 1.1.1.157]; phbB, acetoacetyl-CoA reductase [EC: 1.1.1.36]; atoB, acetyl-CoA C-acetyltransferase [EC: 2.3.1.9]; fadJ, 3-hydroxyacyl-CoA dehydrogenase/enoyl-CoA hydratase/3-hydroxybutyryl-CoA epimerase [EC: 1.1.1.35 4.2.1.17 5.1.2.3]; fadA, acetyl-CoA acyltransferase [EC: 2.3.1.16]; dehH, 2-haloacid dehalogenase [EC: 3.8.1.2]; haloacetate dehalogenase [EC: 3.8.1.3]; glcDEF, glycolate oxidase [EC: 1.1.3.15]; (S)-2-hydroxy-acid oxidase [EC: 1.1.3.15]. Download FIG\u00a0S3, PDF file, 0.3 MB.(A) Heatmap of the distribution of various chain-length fatty acid and haloacid degradation genes in Binatota genomes. The heatmap colors (as explained in the key) correspond to the percentage of genomes in each order encoding a homologue of the gene in the column header. (B) Propionyl-CoA degradation pathways encoded by the Binatota genomes. The methylmalonyl-CoA (MMCoA) pathway is shown in blue, while the 2-methylcitrate pathway is shown in green. In some genomes, the MMCoA pathway seems to be functional but with a slight modification (shown in purple) that includes glyoxylate assimilation and regeneration. Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the actABCDEFG), and complex IV, as well as an F-type H+-translocating ATP synthase , 1f (22 sequences), 1i (1 sequence), and 1h (4 sequences) (2-tolerant hydrogenases) and methane (via pMMO) has been shown to occur in methanotrophic Verrucomicrobia to maximize proton-motive force generation and subsequent ATP production  . Simultaoduction . As well by pMMO  10.1128/mBio.00985-21.5FIG\u00a0S4cytb/cyt1) and/or alternate cytochrome III , while genes encoding cytochrome c oxidase activities (complex IV) belonged to different families, including family A , family C , and/or cytochrome bd (cydAB). Possible electron transfer proteins between complex III  and complex IV belonging to different cytochrome c families are shown. Also shown in panel A is the distribution of the three subunits of the type I respiratory O2-tolerant H2-uptake [NiFe] hydrogenase (hyaABC) in Binatota genomes. (B) Maximum-likelihood phylogenetic tree showing the classification of the hyaA genes carried by the Binatota genomes (magenta) in relation to other [NiFe] hydrogenases. The [Fe-Fe] hydrogenase of Methanobacterium formicicum was used as the outgroup. Bootstrap support (from 100 bootstraps) is shown for branches with >50% support. Download FIG\u00a0S4, PDF file, 0.3 MB.Electron transport chain in the Binatota. (A) Heatmap of the distribution of electron transport chain components in the Binatota genomes and electrons entry points from various substrates. The heatmap colors (as explained in the key) correspond to the percentage of genomes in each order carrying a homologue of the gene in the column header. All subunits of complexes I (NADH-quinone oxidoreductase [EC: 7.1.1.2]) and II (succinate dehydrogenase/fumarate reductase [EC: 1.3.5.1 1.3.5.4]) were encoded in all genomes but are shown here as single components for ease of visualization. Genes encoding quinone-cytochrome C reductase activities belonged to complex III  and oxygenated (xanthophyll) carotenoid biosynthesis capabilities. Carotenoids biosynthetic machinery in the Binatota included crtB for 15-cis-phyotene synthesis from geranylgeranyl pyrophosphate (PP), crtI, crtP, crtQ, and crtH for neurosporene and all-trans lycopene formation from 15-cis-phytone, crtY or crtL for gamma- and beta-carotene formation from all-trans lycopene, and a wide range of genes encoding enzymes for the conversion of neurosporene to spheroidene and 7,8-dihydro \u03b2-carotene, as well as the conversion of all-trans lycopene to spirilloxanthin, gamma-carotene to hydroxy-chlorobactene glucoside ester and hydroxy-\u03b3-carotene glucoside ester, and beta-carotene to isorenieratene and zeaxanthins , third bchE (magnesium-protoporphyrin IX monomethyl ester cyclase [EC: 1.21.98.3]), and fourth bchLNB  steps were identified in the Binatota genomes (bchM (magnesium-protoporphyrin O-methyltransferase [EC: 2.1.1.11]) and the fifth bciA or bicB  or bchXYZ (chlorophyllide a reductase [EC 1.3.7.15]) steps were absent (a (BChl a) formation from chlorophyllide a (bchXYZ (chlorophyllide a reductase [EC 1.3.7.15]) and bchF (chlorophyllide a 31-hydratase [EC 4.2.1.165]) were not identified, while genes encoding bchC (bacteriochlorophyllide a dehydrogenase [EC 1.1.1.396]), bchG (bacteriochlorophyll a synthase [EC: EC: 2.5.1.133]), and bchP (geranylgeranyl-bacteriochlorophyllide a reductase [EC 1.3.1.111]) were present in most genomes (c (BChl c) and d (BChl d) formation from chlorophyllide a (bciC (chlorophyllide a hydrolase [EC: 3.1.1.100]) and bchF (chlorophyllide a 31-hydratase [EC: 4.2.1.165]) or bchV (3-vinyl bacteriochlorophyllide hydratase [EC: 4.2.1.169]) were not identified, while genes for bchR [bacteriochlorophyllide d C-12(1)-methyltransferase (EC: 2.1.1.331)], bchQ [bacteriochlorophyllide d C-8(2)-methyltransferase (EC: 2.1.1.332)], bchU (bacteriochlorophyllide d C-20 methyltransferase [EC: 2.1.1.333]), and bchK (bacteriochlorophyll c synthase [EC: 2.5.1.-]) were identified  and 1,213 (IMG/M) 16S rRNA genes affiliated with the Binatota orders were identified  are shown on the y axis (B). Further subclassifications for each environment are shown for (C) terrestrial, (D) freshwater, (E) marine, (F) host-associated, and (G) engineered environments. Details, including GenBank accession number of hit sequences, are shown in Extended Data 3. Download FIG\u00a0S5, PDF file, 0.4 MB.(A) Maximum-likelihood phylogenetic tree based on the 16S rRNA gene representatives from six Binatota orders with representative hit sequences (number of sequences in parentheses following the order name) from the IMG and NCBI-nt databases identified by Blastn. Orders are color coded following the color scheme in Copyright \u00a9 2021 Murphy et al.2021Murphy et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the In addition to the 16S rRNA-based analysis, we queried the data sets from which a Binatota MAG was binned using the sequence of their ribosomal protein S3 and estimating the Binatota relative abundance as the number of reads mapped to contigs with a Binatota ribosomal protein S3 as a percentage of the number of reads mapped to all contigs encoding a ribosomal protein S3 gene. Results showed relative abundances ranging between 0.1 and 10.21% (average 3.84 \u00b1 3.21%) .Aplysina aerophoba. Analysis of the 16S rRNA data set suggests a notable association between Bin18 and sponges, with relatively high host-associated sequences , suggesting its widespread distribution beyond a single sponge species. The absolute majority of order Binatales sequences  were of a terrestrial origin  in Bin18 and Binatales , Firmicutes  (Verrucomicrobia (Candidatus Methylomirabilis\u201d (NC10)  is especially notable, given the global magnitude of methane emissions and the relatively narrower range of organisms   , Verrucomicrobia , and \u201cCa\u201d (NC10) . Further\u201d (NC10) , 44, for\u201d (NC10) , and met\u201d (NC10) , in the  [NC10])  capable chanisms . All 11 1 oxidation to formaldehyde, formaldehyde oxidation to CO2, and formaldehyde assimilation. Within the world of methylotrophs, a wide array of functionally redundant enzymes and pathways have been characterized that mediate various reactions and transformations in such modules. In addition, multiple combinations of different modules have been observed in methylotrophs, with significant variations existing even in phylogenetically related organisms. Our analysis demonstrates that such metabolic versatility indeed occurs within the methylotrophic modules of Binatota. While few phylum-wide characteristics emerged, e.g., utilization of serine pathway for formaldehyde assimilation, absence of H4MPT-linked formaldehyde oxidation, and potential utilization of PEP carboxykinase (pckA) rather than PEP carboxylase (ppc) for CO2 entry to the serine cycle, multiple order-specific differences were observed, e.g., XoxF-type methanol dehydrogenase encoded by Bin18 and Binatales genomes, MDH2-type methanol dehydrogenase encoded by UBA1149 genomes, absence of methanol dehydrogenase homologues in HRBin30 genomes, absence of methylamine oxidation in order UBA9968, and potential utilization of the ethylmalonyl-CoA pathway for glyoxylate regeneration by the majority of the orders versus the glyoxylate shunt by UBA9968.As previously noted , methyloprmABC, propane monooxygenase), medium- , and long-chain alkanes (ladA) identified  for their activation and conversion to central metabolites  as well as order-specific  habitat preferences ; Fig.\u00a0S5osystems , and we Within the phylum Binatota, it appears that orders HRBin30 and UBA1149 are abundant in thermal vents, thermal springs, and thermal soils, suggesting a specialization to high-temperature habitats . The preThe recovery of Binatota genomes from certain lakes could be a reflection of the high gaseous load in such lakes. Multiple genomes and a large number of Binatota-affiliated 16S rRNA sequences were binned and identified from Lake Kivu, a meromictic lake characterized by unusually high concentrations of methane . Biotica57\u201362\u2013Finally, the occurrence and apparent wide distribution of members of the Binatota in sponges, particularly order Bin18, are notable and could possibly be viewed in terms of the wider symbiotic relationship between sponges and their microbiome. Presence of hydrocarbon degraders , 67, incAlphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria (including methano- and methylotrophs) and Bacteroidetes, Deinococcus, Thermus, Deltaproteobacteria, Firmicutes, Actinobacteria, Planctomycetes, and Archaea, e.g., Halobacteriaceae and Sulfolobus. Here, carotenoids could serve as antioxidants  are not pigmented. Indeed, root-associated facultative methylotrophs of the genus Methylobacterium have traditionally been referred to as \u201cpink-pigmented facultative methylotrophs\u201d and are seen as an integral part of root ecosystems . While such a pattern is tempting to propose phototrophic capacities in the Binatota, the consistent absence of critical genes , coupled with our inability to detect reaction center-encoding genes, prevents such a proclamation. Identification of a single or few gene shrapnel from the chlorophyll biosynthesis pathway in microbial genomes is not unique. Indeed, searching the functionally annotated bacterial tree of life AnnoTree  could possibly be encoded by general methyltransferases (EC: 2.1.1.-), the missing bciC (EC: 3.1.1.100) could possibly be encoded by general hydrolases (EC: 3.1.1.-), and the missing bchF (EC: 4.2.1.165) or bchV (EC: 4.2.1.169) could possibly be encoded by general hydratases (EC: 4.2.1.-).Accordingly, we put forward three scenarios to explain the proposed relationship between Binatota and phototrophy. The most plausible scenario, in our opinion, is that members of the Binatota are pigmented nonphotosynthetic organisms capable of carotenoid production but incapable of chlorophyll production and lack a photosynthetic reaction center. The second scenario posits that members of the Binatota are indeed phototrophs, possessing a complete pathway for chlorophyll biosynthesis and a novel type of reaction center that is bioinformatically unrecognizable. A minimal photosynthetic electron transport chain, similar to that of antiacus , with thantiacus , it has  species . A third12. Such auxotrophies are common in the microbial world and could be alleviated by nutrient uptake from the outside environment  were downloaded as assemblies from NCBI. In addition, 128 metagenome-assembled genomes with the classification \u201cBacteria;UBP10\u201d were downloaded from the IMG/M database (April 2020). These genomes were recently assembled from public metagenomes as part of a wider effort to generate a genomic catalogue of Earth\u2019s microbiome (n\u2009=\u2009108) were retained for further analysis (All genomes classified as belonging to the Binatota in the Genome Taxonomy Database (GTDB) database , with the arbitrary cutoffs 56% and 68% for family and genus, respectively.Taxonomic classifications followed the Genome Taxonomy Database (GTDB) release r89 , 87 and 1, alkanes, and fatty acids metabolism, C1 assimilation, [NiFe] hydrogenases, electron transport chain complexes, and carotenoid and chlorophyll biosynthesis. To build the HMM profiles, Uniprot reference sequences for all genes with an assigned KO number were downloaded and aligned using Clustal-omega  were obtained from the pfam database . Additionally, HMM profiles were built for PscABCD (Chlorobia-specific), PshA/B (Heliobacteria-specific)  and reaction center type 2  , and theoflexota . The HMMoflexota , 99 was oflexota  to checkoflexota , 102, weoflexota . All ide2-tolerant H2-uptake [NiFe] hydrogenase large subunit (HyaA) were classified using the HydDB web tool (All sequences identified as belonging to the respiratory Oweb tool .Methylococcus capsulatus strain Bath (PDB: 3RGB) and to predict tertiary structure models. Predicted models were superimposed on the template enzyme in PyMol . Modeling of the active site was conducted similarly. The dicopper-binding site proposed for Methylococcus capsulatus strain Bath pMMO (3RGB) was used. Alignment of Binatota PmoB sequences with reference Methylococcus capsulatus strain Bath PmoB was performed with Clustal-omega . Two databases were searched: (i) GenBank nucleotide (nt) database (accessed in July 2020) using a minimum identity threshold of 90%, \u226580% subject length alignment for near full-length query sequences or \u226580% query length for non-full-length query sequences, and a minimum alignment length of 100\u2009bp and (ii) The IMG/M 16S rRNA public assembled metagenomes using a cutoff E value of 1e\u221210, percentage similarity\u2009of \u226590%, and either \u226580% subject length for full-length query sequences or \u226580% query length for non-full-length query sequences. Hits satisfying the above criteria were further trimmed after alignment to the reference sequences from each order using Clustal-omega and inserted into maximum-likelihood phylogenetic trees in FastTree . The ecological distribution for each of the Binatota orders was then deduced from the environmental sources of its hits. All environmental sources were classified according to the GOLD ecosystem classification scheme . We alsohttps://github.com/ChelseaMurphy/Binatota. Maximum-likelihood trees (https://itol.embl.de/shared/1WgxEjrQfEYWk. Maximum-likelihood trees for chlorophyll biosynthesis genes are available at https://itol.embl.de/shared/34y3BUHcQd7Lh.Genomic bins, predicted proteins, and extended data for od trees  can be a"}
+{"text": "OBJECTIVES/GOALS: Using multi-state discharge data, to identify predictors of frequent emergency department (ED) use among the homeless patients seen in emergent care, and to compare frequent versus less frequent homeless ED users for their risk of serious health services utilization outcomes. METHODS/STUDY POPULATION: Based on the State Emergency Department Database and the State Inpatient Database, homeless individuals  who made at least one ED visit in four states  in 2014. In this retrospective cross-sectional analysis, patient-level demographic and clinical factors were assessed as predictors for increased ED use. Risks of opioid overdose, opioid-related hospital admission/ED visit, in-hospital mortality, mechanical ventilation, and number of hospitalizations were compared between individuals with 4 or more vs. 2-3 vs. 1 ED visit(s), adjusting for potential confounders including hospital fixed effects . RESULTS/ANTICIPATED RESULTS: Higher rates of ED use were associated with Medicare coverage <65; primary diagnosis of alcohol abuse, asthma, or abdominal pain; and co-morbidity of alcohol abuse, psychoses, or chronic pulmonary disease. Individuals with \u22654 visits had significantly higher adjusted risk of opioid overdose (3.7% vs. 1.2% vs. 1.0%), opioid-related hospitalizations/ED visits (17.9% vs. 8.5% vs. 6.6%), mechanical ventilation (9.8% vs. 7.0% vs. 4.7%), and greater # of hospitalizations (3.2 vs. 1.3 vs. 0.8) compared to individuals with 2-3 or 1 ED visit. Individuals with \u22654 and 2-3 ED visits had similar but increased risks of in-hospital mortality compared to individuals with 1 ED visit (2.8% vs. 2.8% vs. 2.3%). DISCUSSION/SIGNIFICANCE OF IMPACT: Homeless patients who were high ED users were more likely to be hospitalized and have other adverse outcomes. These findings encourage targeted interventions (i.e. housing) for the high-utilizer homeless population to reduce the burden of serious outcomes and costs for the patient and society."}
+{"text": "BCOR and BCORL1. We report a distinct co-mutational pattern that suggests a role in disease progression rather than initiation, especially affecting mechanisms of DNA-methylation. Further, we found loss-of-function mutations of BCOR to be independent markers of poor outcomes in multivariable analysis. Therefore, loss-of-function mutations of BCOR need to be considered for AML management, as they may influence risk stratification and subsequent treatment allocation.Acute myeloid leukemia (AML) is a genetically heterogeneous disease. Clinical phenotypes of frequent mutations and their impact on patient outcome are well established. However, the role of rare mutations often remains elusive. We retrospectively analyzed 1529 newly diagnosed and intensively treated AML patients for mutations of BCL6 corepressor (BCOR) and its homolog, the BCL6 corepressor-like 1 (BCORL1), have been reported to be rare but recurrent mutations in AML. Previously, smaller studies have reported conflicting results regarding impacts on outcomes. Here, we retrospectively analyzed a large cohort of 1529 patients with newly diagnosed and intensively treated AML. BCOR and BCORL1 mutations were found in 71 (4.6%) and 53 patients (3.5%), respectively. Frequently co-mutated genes were DNTM3A, TET2 and RUNX1. Mutated BCORL1 and loss-of-function mutations of BCOR were significantly more common in the ELN2017 intermediate-risk group. Patients harboring loss-of-function mutations of BCOR had a significantly reduced median event-free survival : 1.005\u20132.134), p = 0.047), relapse-free survival (HR = 1.904 (95%-CI: 1.163\u20133.117), p = 0.01), and trend for reduced overall survival (HR = 1.495 (95%-CI: 0.990\u20132.258), p = 0.056) in multivariable analysis. Our study establishes a novel role for loss-of-function mutations of BCOR regarding risk stratification in AML, which may influence treatment allocation.Acute myeloid leukemia (AML) is characterized by recurrent genetic events. The  Acute myeloid leukemia (AML) is a genetically heterogeneous disease . In the BCOR) gene, and its homolog, the BCL6 corepressor-like 1 (BCORL1) gene, are located on chromosomes Xp11.4 and Xq26.1, respectively [The BCL6 corepressor  syndrome in heterozygous females, and prenatal death in hemizygous males [BCOR (mBCOR) and BCORL1 (mBCORL1) both occur in 4\u20136% of patients, and an association with poor outcomes has been suggested [Since BCOR is vital in ectodermal and mesenchymal differentiation, germline loss-of-function mutations in us males . Somaticus males ,16,17,18us males . It has us males ,20, chrous males , clonal us males , myelodyus males ,23,24,25us males ,26,27,28We retrospectively analyzed a multi-center cohort of 1529 AML patients. Eligibility criteria were newly diagnosed AML according to WHO definitions , AML200330). Remission and survival criteria were defined according to ELN2017 recommendations [AML was defined as de novo when neither previous malignancy nor previous treatment with chemo- and/or radiotherapy was reported. When myeloid neoplasms were documented prior to AML diagnosis, AML was defined as secondary (sAML). Prior exposure to chemo- and/or radiotherapy before the initial diagnosis defined therapy-associated AML (tMN). Early death was defined as death by any cause within 30 days of the initial diagnosis  using a TruSight Myeloid Sequencing Panel  was performed on pre-treatment bone marrow or peripheral blood, targeting 54 genes  associatn detail ,35. A DNWe compared categorical variables between groups using the chi-squared test, while continuous variables were compared using the Kruskal\u2013Wallis test. The Kaplan\u2013Meier method was used to estimate survival probabilities. The logrank test was used to compare survival time distributions between groups. Cox regression was used to estimate univariate and adjusted hazard ratios. For the binary endpoint of complete remission, logistic regression models were fitted to estimate univariate and adjusted odds ratios. A significance level of 0.05 was used to determine statistical significance. Calculations were performed in R 4.0.3.n = 1529), we found mBCOR in 71 (4.6%) and mBCORL1 in 53 (3.5%) patients. Twelve patients (0.8%) concomitantly harbored both mBCOR and mBCORL1. The median age for the entire cohort was 55 years (Interquartile range (IQR): 44\u201364). Median variant allele frequency (VAF) for mBCOR and mBCORL1 was 48% (range: 7\u2013100%) and 47% (range: 5\u2013100%), respectively (BCOR (87%) and mBCORL1 (83%) carried two or more additional mutations in other driver genes, while only one patient in each group had no other co-mutations detected by the panel , RUNX1 (30%), TET2 (23%), NRAS (20%), BCORL1 (17%), and STAG2 (17%). Low mutation frequencies were detected for other genes frequently mutated in AML, such as FLT3 , NPM1 (11%), and TP53 (4%). Similar to mBCOR, in mBCORL1 AML the majority of mutations had a loss-of-function effect , RUNX1 (34%), FLT3-ITD (25%), BCOR (23%), and TET2 (23%).However, due to the unknown effect of protein function, these mutations were classified as UFs (unknown functions) for sub-analysis. The co-mutational landscape of mBCOR AML F was chaect 60%, D and werect 60%, E were DNBCORL1 were more prevalent in females than in males , while for mBCOR no such association was observed. For both mBCOR and mBCORL1 no statistically significant associations were detected for age at diagnosis, the presence of a complex karyotype, hemoglobin levels, or platelet counts. With respect to disease status, the rate of sAML was significantly higher amongst patients harboring mBCOR than amongst wtBCOR patients , while no specific association with mutation type (LOF or UF) or mBCORL1 was found regarding AML type. Outcomes did not differ for patients with sAML and mBCOR or mBCORL1 compared to patients with de novo AML. In the ELN2017 intermediate-risk group we found a higher proportion of LOF of mBCOR compared to UF mBCOR and wtBCOR , as well as mBCORL1 compared to wtBCORL1 . Median white blood cell (WBC) counts in LOF mBCOR AML were significantly lower compared to UF mBCOR and wtBCOR (p < 0.001), while mBCORL1 showed no significant association with WBC. BCOR (LOF and UF) and mBCORL1, respectively.Mutations of BCOR and mBCORL1 were not associated with the rate of complete remission (CR) (mBCOR: OR = 0.781 (95%-CI: 0.469\u20131.301), p = 0.342 and mBCORL1: OR = 0.795 (95%-CI: 0.442\u20131.432), p = 0.445) or with ED30 (mBCOR: OR = 0.679 (95%-CI: 0.209\u20132.199), p = 0.518 and mBCORL1: OR = 1.288 (95%-CI: 0.454\u20133.649), p = 0.634). In contrast, mBCOR was associated with lower median measures of event-free survival (EFS) (2.8 months (95%-CI: 1.7\u20138.5) vs. 7.6 months (95%-CI: 6.8\u20138.5), HR = 1.485 (95%-CI: 1.147\u20131.922), p = 0.003; p = 0.026; p = 0.095; BCOR in general was not independently associated with EFS (HR = 1.243 (95%-CI: 0.89\u20131.736), p = 0.202), RFS (HR = 1.407 (95%-CI: 0.93\u20132.129), p = 0.106), and OS (HR = 1.216 (95%-CI: 0.846\u20131.748), p = 0.292).With respect to clinical outcomes, mBCOR, compared to UF mBCOR and wtBCOR, we found in both univariate and multivariable analysis significantly reduced median EFS (LOF mBCOR: 1.9 months (95%-CI: 1.4\u20138.0) vs. UF variant of mBCOR: 4.9 months (95%-CI: 1.676\u2013n.a.) vs. wild-type BCOR: 7.5 months (95%-CI: 6.8\u20138.5), multivariable HR of LOF compared to wtBCOR = 1.464 (95%-CI: 1.005\u20132.134), p = 0.047, BCOR: 8.8 months (95%-CI: 7.3\u201324.0) vs. UF variant of mBCOR: 17.7 months (95%-CI: 8.317\u2013n.a.) vs. wild-type BCOR: 18.7 months (95%-CI: 16.3\u201323.3), multivariable HR of LOF compared to wtBCOR = 1.904 (95%-CI: 1.163\u20133.117), p = 0.01, BCOR variants in univariate analysis (LOF mBCOR: 11.6 months (95%-CI: 8.5\u201333.2) vs. UF variant of mBCOR: 14.2 months (95%-CI: 11.045\u2013n.a.) vs. wild-type BCOR: 18.4 months (95%-CI: 16.7\u201321.4), HR of LOF compared to wtBCOR = 1.409 (95%-CI: 1.010\u20131.965), p = 0.044), and a strong trend for reduced median OS in a multivariable analysis (HR = 1.495 (95%-CI: 0.990\u20132.258), p = 0.056, DNMT3A, TET2, and RUNX1 did not significantly affect HR. However, in order to precisely account for multiple interactions, an even larger cohort is needed, due to the rarity of LOF BCOR.For patients harboring LOF mBCORL1 in general compared to wtBCORL1, median EFS (3.6 months (95%-CI: 1.9\u20139.8) vs. 7.5 months (95%-CI: 6.7\u20138.3), HR = 1.204 (95%-CI: 0.885\u20131.639), p = 0.236; p = 0.263; p = 0.654; BCORL1 showed significantly reduced EFS (LOF mBCORL1: 3.5 months (95%-CI: 1.1\u201310.1) vs. UF mBCORL1: 6.2 months (95%-CI: 1.9\u2013not reached) vs. wtBCORL1: 7.5 months (95%-CI: 6.7\u20138.3), HR = 1.521 (95%-CI: 1.045\u20132.213), p = 0.028); however, in a multivariable analysis adjusting for age, AML type, and ELN2017 risk, this was not statistically significant.In AML with mBCOR and mBCORL1 regarding outcomes in different ELN2017 risk groups. Since few patients with mBCOR and mBCORL1 were in the ELN2017 favorable  or ELN2017 adverse-risk groups , we focused on the ELN2017 intermediate-risk group . In ELN2017 intermediate-risk AML with mBCOR, median EFS and OS did not differ compared to wtBCOR, while there was a trend of lower median RFS in a univariate model (11.9 months (95%-CI: 7.0\u201324.0) vs. 14.8 months (95%-CI: 12.5\u201320.3), HR = 1.446 (95%-CI: 0.977\u20132.139), p = 0.065), as well as in a multivariable model adjusted for age and AML type (HR = 1.637 (95%-CI: 0.995\u20132.693), p = 0.052). In ELN2017 intermediate-risk AML with mBCORL1, median EFS and RFS did not differ, but there was a trend of lower median OS in a univariate model (14.9 months (95%-CI: 7.3\u201324.4) vs. 17.0 months (95%-CI: 14.0\u201320.6), HR = 1.401 (95%-CI: 0.954\u20132.058), p = 0.086), while in a multivariable model adjusted for age and AML type this difference was not statistically significant. There was no significant association between LOF of mBCOR or mBCORL1 and ELN2017 risk groups regarding outcomes.We did not find statistically significant differences in mBCOR and BCORL1. The respective proportions of mBCOR and mBCORL1 in the cohort were comparable to those reported in recent studies [BCOR with LOF and mBCORL1 to be more prevalent in patients in the ELN2017 intermediate-risk group [BCOR was significantly higher than in their wild-type counterparts, confirming previous reports [BCORL1 were significantly more prevalent in females than in males, as has been previously suggested [BCOR with LOF had significantly lower WBC, and a trend for lower peripheral blood blast counts. Mutations of BCOR frequently co-occurred with mutations of DNMT3A, RUNX1, TET2, NRAS, and BCORL1. Since both BCOR and DNMT3A function as epigenetic modifiers [BCOR and mutations of TET2, another epigenetic modifier [BCOR in our cohort, has been reported to induce MDS [BCOR and other essential regulators of normal myeloid development appears to be a factor that may promote leukemogenesis. Recent studies suggest, however, that perturbations of BCOR function alone do not suffice to induce malignant transformation [BCOR following an oncogenic event, e.g., MDS driver mutation triggers disease progression towards AML, as suggested by the higher frequency of sAML among mBCOR patients. Accordingly, in our cohort of patients with mBCOR AML, the majority had at least two co-mutations, while only one patient harbored no other co-mutations targeted by our panel (BCOR, co-mutations of mBCORL1 were RUNX1, DNMT3A, and TET2, as well as BCOR and FLT3-ITD, which were only rarely co-mutations of mBCOR. Again, the majority of patients harboring mBCORL1 had at least two other co-mutations, and only one patient had no other co-mutations revealed by our panel (BCOR, this suggests a potential interplay of impaired BCORL1 function with other dysfunctional mechanisms of DNA methylation, cell differentiation, and signal transduction in leukemogenesis.We analyzed a large cohort of newly diagnosed and intensively treated AML patients according to their mutational status of  studies ,21,26,27sk group . The pro reports . Mutatioodifiers , a synerodifiers ,40. In mmodifier  frequentduce MDS ,42. Thisduce MDS ,43. Therormation ,21,44, aur panel B. Similaur panel B. As forBCOR and mBCORL1 regarding CR rate and ED30, compared to wild-type patients. Interestingly, while we observed lower median EFS, RFS, and OS for both mBCOR and mBCORL1 in general, only LOF mutations of mBCOR were associated with significantly reduced EFS and RFS and a trend of reduced OS in multivariable testing adjusted for age, AML type, and ELN2017 risk. In MDS, Abuhadra et al. [BCOR, while general mutation status did not affect OS. Previous studies of mBCOR in AML have reported poorer outcomes to often be associated with distinct co-mutations; however, a significant association of LOF mutations of BCOR as independent markers of poor outcomes has not yet been reported in AML. Terada et al. [FLT3, and had intermediate-risk cytogenetics. Grossmann et al. [BCOR and a trend of reduced OS in normal-karyotype AML with no mutations in NPM1, FLT3-ITD, CEBPA, or MLL-PTD. However, in a validation cohort no association between mBCOR and OS was observed. Nevertheless, in both cohorts reduced EFS was detected [BCOR or SETBP1. Our findings underline the complexity of the mutational landscape of AML, where even mutational variants of rare mutations have to be considered in order to determine their clinical and prognostic effects. Future work needs to focus on the implementation of LOF mutations of BCOR in risk stratification tools for AML management.Regarding outcomes, we found no statistically significant differences between patients with ma et al.  recentlya et al.  reportedn et al.  reporteddetected . Recentldetected  reportedBCOR and mBCORL1 are rare but recurrent mutations in AML. While previous studies suggested poor outcomes for mBCOR in AML, especially in the context of co-occurring mutations, we found loss-of-function mutations of mBCOR to be independent markers of poor outcomes in AML, while mBCORL1 was not significantly associated with outcomes in multivariable testing.In conclusion, both m"}
+{"text": "Trimethoprim-sulfamethoxazole (TMP-SMX) is a high-bioavailability antibiotic associated with potentially serious adverse drug events (ADE). The objective of this study was to evaluate the safety of intravenous (IV) and oral (PO) high-dose TMP-SMX.\u2265 18 years old and > 72 hours of renally adjusted high-dose TMP-SMX defined as \u2265 5 mg/kg/day of TMP. Exclusion: prophylaxis. Endpoints during treatment: hyponatremia with sodium < 135 mmol/L, hyperkalemia with potassium > 5 mmol/L, serum creatinine increase of \u2265 0.3 mg/dL or 1.5-1.9 times from baseline, and fluid overload on physical exam. Descriptive and bivariate statistics were performed.IRB-approved retrospective cohort of hospitalized patients from January 2016 to November 2020. Inclusion: Stenotrophomonas maltophilia (52% IV and 18% PO) and Pneumocystis jiroveci (16.3% IV and 62% PO). Median (IQR) days of inpatient therapy: 6 (5-7.5) PO vs. 7.5 (6-11.3) IV. Median (IQR) days of total duration: 9 (6-21.5) PO vs. 12 (7.8-14) IV (p=0.93). IV group: 88% of patients received >1 liter of D5W daily. Median (IQR) liters of D5W daily was 1 (1-1.5). 56% had a diuretic added, and 38% had a diuretic dose increase. Majority of patients (78%) on IV were taking other oral medications. 100% patients experienced any adverse event with IV vs. 70% with PO . Most common ADE in both groups: hyponatremia, hyperkalemia, and elevated creatinine. Hyponatremia: 92% with IV and 32% with PO . Edema on physical exam, an ADE specific to IV TMP-SMX, was the third most common side effect in the IV group. Relative changes from baseline in sodium, potassium, and creatinine from those who experienced hyponatremia, hyperkalemia and elevated creatinine were listed in Table 2.Each group included 50 patients (Table 1). Intensive care unit patients comprised 82% IV TMP-SMX compared to 32% PO. Most common infection: respiratory tract 86% IV and 68.1% PO. Most common organisms were Table 1. Baseline and Clinical CharacteristicsTable 2. Adverse EffectsPatients on IV TMP-SMX therapy were more likely to experience an ADE compared to PO, likely driven by the high volume of free water. Most patients on IV TMP-SMX were on other PO medications, suggesting a missed stewardship opportunity for IV to PO conversion to reduce patient harm.Susan L. Davis, PharmD, Nothing to disclose Michael P. Veve, Pharm.D., Cumberland (Grant/Research Support)Paratek Pharmaceuticals (Research Grant or Support) Rachel Kenney, PharmD, Medtronic, Inc."}
+{"text": "Microbacterium foliorum NRRL B-24224. The 41,958-bp double-stranded DNA genome has 71 predicted protein coding genes and 1 tRNA. The lytic actinobacteriophage was extracted from soil samples collected in Stephenville, TX, and is related to cluster EB bacteriophages Didgeridoo and Lahqtemish.Microbacteriophage IndyLu was isolated from  Microbacterium foliorum NRRL B-24224 is a rod-shaped Gram-positive aerobic bacterium from the order Actinomycetales that contains no intact prophages or apparent antibacteriophage restriction-modification or CRISPR systems  (MH045566) and Lahqtemish (GenBank accession number MT889392). Auto-annotation using GLIMMER v3.02 (http://phagesdb.org/DNAMaster/), and PECAAN. Microbacteriophage IndyLu is predicted to contain 70 protein-coding genes and 1 tRNA coding for glutamine, identified using ARAGORN v1.2.38   showed gER v3.02 , 11 was ER v3.02 , DNA Mas v1.2.38  and tRNA v1.2.38 . Putativ v1.2.38 , 16 and OK318958. The raw reads are available in the SRA under accession number SRX12683423.The actinobacteriophage IndyLu genome sequence is available in GenBank under accession number"}
+{"text": "Theshort answer is to administer alpha-2 agonists slowly from admission orendotracheal intubation up to stabilized cooperative sedation. The \u201ctake home\u201dmessage is as follows: a) alpha-2 agonists are jointly sympathetic deactivatorsand sedative agents; b) sympathetic deactivation implies maintaining the strokevolume and iterative assessment of volemia. Evidence-based medicine shoulddocument our propositions.Cardiac, ventilatory and kidney management in the critical care setting has beenoptimized over the past decades. Cognition and sedation represent one of thelast remaning challenges. As conventional sedation is suboptimal and as thesedation evoked by alpha-2 adrenergic agonists  represents a valuable alternative,this manuscript covers three practical topics for which evidence-based medicineis lacking: a) Switching from conventional to cooperative sedation(\u201cswitching\u201d): the short answer is the abrupt withdrawal of conventionalsedation, immediate implementation of alpha-2 agonist infusion and the use of\u201crescue sedation\u201d (midazolam bolus[es]) or \u201cbreakthrough sedation\u201d  to stabilize cooperative sedation. b) Switching from conventional tocooperative sedation in unstable patients (e.g., refractory  Given the circulatory drawbacks in hypovolemic patients, only nicheindications are to be considered , which contradicts the\u201cone size fits all\u201d approach. Circulation is a major concern. In the setting ofsystolic15,16)( or diastolic17)( failure or cardiogenic pulmonary edema and a lowleft ventricular (LV) ejection fraction,18)( the sympathetic deactivation of capacitance(veins) and resistance vessels  is beneficial. Venous return is reduced, and ejectionimproves. In the hypovolemia scenario, alpha-2 agonists further reduce venous return and stroke volume (SV) and worsencirculatory distress .Alpha-2 adrenergic agonists  evoke \u201ccooperative\u201d, rousable sedation6,22-25)( or sleep26)( anti-inflammation31-35)$( and a reduced CCU stay.36), problems arise: a) how to switch from conventionalsedation to alpha-2 agonists (\u201cswitching\u201d), e.g., in agitated or unstable patients,refractory delirium tremens (DT), circulatory/ventilatory distress,etc.; and b) how can alpha-2 agonists be prescribed as first-line sedatives de novoupon admission? This manuscript addresses the parasympathetic vs. sympatheticsystems, circulation, and ventilation.As alpha-2 agonists interfere with the autonomic system 46-48). Our clinical practice spanning the period of 1980 -2020 in several countries  is summarized . Physiol49)emergence delirium is encountered following deep sedation. However,is this delirium related to the pathology itself, the CCUenvironment, or conventional sedation? Moreover, b) deep sedation, bordering GA (1),is used in clinical practice for ARDS or increased intracranialpressure49)( without evidence.50), paralysis and proning.51,52)( is missing.50)the absence of a control group under adequate spontaneous breathing50)( and b) the tendency toshorten57,58)( without strong evidence50), traumatic brain injury,As most groups use cooperative sedation after conventional sedation, i.e., only whenthe patient is recovering and ready for tracheal extubation (\u201cextubation\u201d),switching from conventional to cooperative sedation is examined first.- Hypovolemia: See below.&), sick sinus syndrome,atrioventricular block II or III without a pacemaker.- Bradycardia (- Liver failure (Child-Pugh C): Clonidine and dexmedetomidine areexcreted through the kidney and liver, respectively. Moreover,clonidine and dexmedetomidine are useful in the scenarios of liverand kidney failure, respectively. Nevertheless, a) clonidine can beadministered in the setting of acute renal failure if renalreplacement therapy (RRT) is used, and b) dexmedetomidine can beused in the setting of liver cirrhosis.Dexmedetomidine and clonidine are sympathetic inhibitors in healthy restingsupine volunteers. In the CCU, given the increased sympathetic activity, theynormalize sympathetic hyperactivity back toward baseline, i.e., sympatheticdeactivators, with the following contraindications: no clinical relevance but is only an invitro finding.63). In contrast, clonidine p.o.allows for convenient oral administration (nonintubated patient with DT),transitioning alpha-2 agonists from i.v. dexmedetomidine to p.o. clonidine toavoid alpha-2 agonist withdrawal, etc. Sedation is achieved within 30 - 60minutes in healthy volunteers after clonidine 300\u00b5g p.o.5,12) < +1 occursimmediately before initiation of dexmedetomidine infusion23). Rescue sedation is used to achieve-2 <RASS < +123)(using either a) fentanyl infusion, followed by a propofol bolus (25 -50mg)22)( or b) midazolam  and fentanyl.23).A \u201cceiling\u201d effect is reported with dexmedetomidine>1.5\u00b5g.kg-1.h-1.7) or propofol (25mg) to berepeated if necessary.65)( thesympathetic inhibition evoked by propofol68), blood pressure (BP) and cardiac output(CO).69) or a bolus of clonidine/dexmedetomidine. Consequently,severe bradycardia and hypotension may occur. To avoid such sideeffects, we used abrupt withdrawal. Abrupt withdrawal is performedduring the day shift only, starting in the earlymorning\u00a3. The prescription specifies the target (-2 <RASS < 0), the range of dose of dexmedetomidine (\u22641.5\u00b5g.kg-1.h-1), the rescue versusbreakthrough sedation ,and the supplementation (neuroleptics: see refractory DT).Progressive switching and circulation: Simultaneous administration ofconventional sedation and cooperative sedation combines thesympathetic deactivation evoked by alpha-2 agonists,69)( withoutsequelae.71) led to no changein mortality (SPICE III).72), while \u224875% of dexmedetomidine patients receivedpropofol, midazolam or both.72,73)( isuseless.73)( to reassess the outcome and makethis large series72)( or CCU72).Intraoperative opioid-free anesthesia was administered:13)(4\u00b5g.kg-1/15 minutes during the induction of anesthesia for livertransplant28) is usually sufficient to cover the firstpostoperative day, with provision for opioid-free analgo-sedation andnicardipine (0.5mg to be repeated if needed). Technology addresses volume  orperfusion . If, after volume adjustment,the perfusion pressure is a concern, adjuncts  areused to counteract sympathetic deactivation. An additive effect between theincoming alpha-2 agonist and the opioid69)( should be eliminated. ii) Ifconventional GA has been administered, low-dose alpha-2 agonists will beintroduced slowly  toeffect.Intraoperative conventional GA has been administered: given premedication with analpha-2 agonist23). Second, if needed afterthis interval, neuroleptics are administered. This avoids thecognitive side effects of benzodiazepines (below: refractory DT).Midazolam is used only as rescue sedation during the switch, e.g.,to facilitate nursing.We do not use-2 < RASS < +1 as others do:82) or low-dose neuroleptics added to high-dosealpha-2 agonists .Elderly patients appear less sensitive to the sedation evoked byalpha-2 agonists than young, muscular, combative, andaddictedThe required RASS (-2 < RASS < 0) deserves comments: 7)or Behavioral Pain Scale (BPS) score, opioids22,23)( (fentanyl 0.5 - 1\u00b5g.kg-1 every 15minutes) or nonopioid analgesics can be selected. However, a) alpha-2 agonistsevoke analgognosia8)( and preserve respiratory genesis.9-11), low-dose ketamine (50mg.d-1) and tramadol(\u201cweak\u201d opioid: 400mg.d-1). These doses are reduced by 50 - 75%after 24 - 72 hours. This may be a consequence of accumulation orthe indifference to pain evoked by the alpha-2 agonist followingsteady-state cooperative sedation.or lidocaine 0.5mg.kg-1.h-1 infusion (loading dose: 1mg.kg-1.h-1) orketamine (0.25mgkg-1.h-1) infusion or gabapentin  or pregabalin or carbamazepine or amitriptyline. Low-dose opioids are employed as rescue analgesics, ifneeded.Once steady-state cooperative sedation is achieved, antinociception isconsidered. Patients presenting with medical conditions require littleantinociception84,85) leads to hypertension and low HR (60 -70 beats per min in a 2-year-old child) without sequelae upon reduceddexmedetomidine administration.86)..In the setting of ambulatory cardiology, very high-dose alpha-2 agonists lead toresistant hypertension (clonidine 5400 - 6000\u00b5g.d-1).90-92)( low-dose dexmedetomidine(0.7\u00b5g.kg-1.h-1) was successfully supplemented with haloperidol innonintubated patients . Dexmedetomidine achieves \u224893% satisfactory sedationlevels  and halves the CCU stay.93). Thus, a combination of drugsmanages a complex neurochemical pattern.DT involves hyperactivity or hypoactivity of several central pathways. Thesignal-to-noise ratio95)(-2 \u2264 RASS \u2264 0) but\u201cfairly alert\u201d5)( or cognitivelyimproved24)( and thetemperature100)( and oxygen consumption(VO2)101-103) are insufficient to achieve-2 \u2264 RASS \u2264 0(stringent absence of restlessness) without tremor, neuroleptics are employed assecond-line agents. When hallucinations were prominent, haloperidol  is administered. In contrast, when agitation wasprominent, loxapine (100mg x 4 p.o. or via the nasogastric tube) is selected.Neuroleptics, then alpha-2 agonists, are tapered as soon as the absence ofrestlessness without tremor is ascertained for 24 hours.104)( or low-dose midazolam (0.5mg.h-1) may beconsidered.Refractory DT patients with Gayet-Wernicke disease required clonidine 4\u00b5g.kg-1.h-1+loxapine 400mgx4 to achieve-2 \u2264 RASS \u2264 0 andthe absence of tremor. To supplement a combination of high-dose alpha-2 agonist+ neuroleptic  and to avoid theadministration of higher doses of alpha-2 agonists + neuroleptics, baclofen (50- 150mg according to kidney function)93) and achieve quietness within 30 - 60 minutes. A similar regimenmay be used to transition from i.v. dexmedetomidine to oral clonidine,105)( up todiscontinuation.105) may be considered to initiate oral therapy or to transitionfrom i.v. dexmedetomidine to an oral alpha-2 agonist.Refractory DT in nonintubated patients106,107)( orrefractory111), the administration of alpha-2 agonistsis inadvisable in the setting of uncontrolled hemorrhage, septic or cardiogenicshock, etc. Indeed, for a short period of time, sympathetic activation is alifesaver in regards to control of the pathology, and exogenous vasopressorsand/or inotropes are required to maintain left ventricular perfusion pressureand/or contractility, in addition to endogenous sympathetic nervous activation.In contrast, AFTER controls acute cardioventilatory distress, and alpha-2agonists deactivate the prolonged sympathetic hyperactivity observed in the CCU.After circulatory optimization, normalized sympathetic hyperactivity towardbaseline may benefit metabolic syndrome, immunoparalysis, etc., e.g., in thefollowing settings: circulatory failure following cardiacsurgery108-110,112-114). Increasedpressor responsiveness to noradrenaline toward baseline follows.113,114)21,117)( Increased LV compliance17)( and vasomotorsympathetic deactivation in resistance vessels (arteries)15,16,118): PLR separates thevolume-responsive versus nonresponsive patients: thevolume-responsive patients are not necessarily in ahypovolemic state and do not necessarily need additionalvolume. Volume is minimized to prevent increased lungwater.122,123)( This may evokehypotension within the first 3 hours ofadministration125) or each incrementof alpha-2 agonist, absence of or minimal collapsibilityof the vena cava28,116,126)(O2 arteriovenousdifference127)( or superiorvena cava oxygen saturation (SsvcO2), carbondioxide (CO2) gap.- Second, the adequacy of CO and microcirculation areaddressed: diuresis, capillary refill, mottling,lactate,No hypovolemia: Following alpha-2 agonist administration, SVmaintenance is required:\u201cstart slow, goslow\u201d, borrowed from the administration of beta-blockers in heartfailure128)( (124)( (21). We propose this overtlycautious approach and termed it ure(128) . No alph) ((124) .(21)Drugs combining sedation and sympathetic deactivation modify the circulation andrequire the following: In summary, bolus alpha-2 agonist administration with simultaneous conventionalsedation administration or without the iterative assessment of volemia leads tosevere bradycardia and hypotension.51,52)( presents shortcomings.50)( are in their infancy.Established practice129) imposed onhypovolemia worsens circulatory distress.Switching from conventional to cooperative sedation in the setting of hypovolemiaand vasopressor administration addresses only one circulatory issue. In thesetting of ventilatory distress, circulatory distress is intermingled withventilatory distress:130): fevercontrol,131-133)( systemic acidosis136-138),10,61)( and uprightpositioning.139)( in 55).140,141). Rescue sedation isadministered if needed. Muscle relaxants are withdrawn immediately beforesteady-state cooperative sedation is established, with reassurance. Spontaneousbreathing is established as soon as84,148)(versus deep sedation).In summary, the management becomes analytical: administration of an alpha-2agonist allows one to separate the physiological versus controlled mandatory ventilation150), and switchingfrom conventional to cooperative sedation (see above).Switching in a patient presenting with acute cardioventilatory distress underconventional sedation in the CCU involves prioritizing between simultaneousissues beyond the scope of this manuscript: stabilized circulation (see above),stabilized ventilatory distress , PaO2/FiO2(P/F), HR, and systolic BP normalize. The patient is discharged withoutintubation.151)( These reports need replication.Dexmedetomidine (infusion: 0.7\u00b5g.kg-1.h-1) addresses agitation in patientstreated with NIV presenting with postoperative ventilatoryfailure.153-155)( or status asthmaticus (dexmedetomidine: 0.2- 0.7\u00b5g.kg-1.h-1).156)( Clonidine (4\u00b5g.kg-1 p.o.) achieves thesame effect.157) andtitrated to effect: stringent ataraxia is requested when psychogenic stimuli arepresented.A similar positive outcome was observed in the setting of severe bronchospasm(dexmedetomidine: 0.25 - 0.8\u00b5g.kg-1.h-1)158)(or early diffuse ARDS150)( are beyond the scope of this section.SARS-CoV-2-ARDS (COVID-ARDS) is an inflammatory disease leading to a highrespiratory drive and an inflammatory vascular disease of the pulmonarycapillaries. Low or medium PEEP is required, with tight control of temperature,agitation and inflammationSeptic shock143,159-161) or very highO2 flow allows one to buy time, expeditepreoxygenation162) prevents the circulatory collapse observed immediatelyafter intubation + positive pressure ventilation + PEEP in hypovolemicpatients.163). Second, iterative rescue sedation allowsone to stabilize incremental cooperative sedation. Finally, P-SILI and hypoxemiaare addressed (Table 155).The dose of dexmedetomidine is a function of the circulation . As an extended CCU stayis likely, immediate stable cooperative sedation is not warranted. First,stabilization of the circulation should be achieved (volume vs. vasopressorswhen the diastolic pressure is low148) achieves-2 < RASS< 0.Tolerance to the sedative effects of alpha-2 agonists develops overweeks165), titration of alpha-2agonists to effect, reassurance and additional neuroleptics.Muscle relaxation suppresses P-SILI and patient-to-ventilatordyssynchronyTwo issues deserve comment: 13) toisolated ventilatory distress and then to acute cardioventilatory distress. A \u201cstartslow-go slow\u201d approach is suggested. Neuroleptics supplement alpha-2 agonists, ifneeded, without benzodiazepines or propofol. Opioid-free analgesia is recommended.To avoid switching from conventional to cooperative sedation, alpha-2 agonistsshould be used as first-line sedatives.46), nociception, circulation . Evidencegathered from a randomized trial using a clear-cut design73,74)( may extend the preliminary outcomedata37-45)( and implement the present suggestions.In the critical care unit, alpha-2 agonists present intrinsicallyintertwined"}
+{"text": "Obsessive-Compulsive Disorder (OCD) is an anxiety disorder, which is the sixth largest contributor to non-fatal health loss globally. Coronavirus disease (COVID-19) pandemic, aside from its impact on physical health, has also had its effects on mental health. This study aimed to explore the frequency of new onset OCD symptomology in medical students amidst COVID-19 pandemic and its association with potential sociodemographic parameters.This cross-sectional study was conducted among medical students studying in Pakistani medical colleges. Data were collected after ethical approval from 1st January 2020 to 20th January 2020 during the second COVID-19 wave. Participants with a history of diagnosed psychiatric illness such as OCD, bipolar disorder, depression, anxiety and those taking relevant medications were excluded from the study. The online questionnaire included Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) and Revised Padua Inventory-Contamination Subscale (PI-CS), which were used to assess OCD symptoms and aversion for contamination respectively. Participants filled Y-BOCS twice, once for pre-pandemic score , and a second time during 2nd wave of COVID-19. Data were analysed using IBM SPSS v23.0 .The study included 711 participants  from over 46 medical colleges and over 44 cities of Pakistan. The mean pre-pandemic and mid-2nd wave Y-BOCS scores were 11.86 \u00b1 6.02 and 15.61 \u00b1 7.41 respectively. The mean PI-CS score was 17.27 \u00b1 9.17. Twenty five percent (n = 176) of students developed new onset OCD symptomology during pandemic, while seventy percent (n = 497) suffered from worsened Y-BOCS score during pandemic. New onset OCD symptomology was associated with age less than 20 years (p = 0.02), higher PI-CS score (p = 0.001) and studying in preclinical years (p = 0.002). Worsening of YBOCS score had significant association with female gender (p = 0.02), attending pandemic related awareness seminar (p = 0.027), studying in preclinical years (p < 0.001) and age less than 20 years (p < 0.001). High Padua scorers (16 and above) showed significant association with increase in YBOCS score (p < 0.000), age less than 20 years (p = 0.005), preclinical years (p = 0.001), frequency of engagements in pandemic related discussions (p = 0.001) and change in YBOCS score (p < 0.001).Our findings indicate that the prevalence of OCD symptomology increased during the COVID-19 pandemic as demonstrated by increased Y-BOCS scores.Femal medical students and students in preclinical years are more likely to suffer from psycological impact of COVID-19 pandemic and heightened concerns and fear for contamination."}
+{"text": "Lenacapavir (LEN) is a first-in-class HIV-1 capsid (CA) inhibitor in clinical development for treatment and prevention of HIV-1 infection. CALIBRATE is an ongoing, phase 2 clinical study evaluating subcutaneous (SC) or oral LEN, in combination with other antiretrovirals, in treatment-na\u00efve people with HIV-1. High rates of virologic success (HIV-1 RNA < 50 copies/mL) were achieved with LEN-based regimens by FDA Snapshot analysis at Week 28. Here, we present interim resistance analyses through Week 28. Participants were randomized (2:2:2:1) to treatment groups (TG) : SC LEN + oral daily emtricitabine/tenofovir alafenamide (F/TAF); at Week 28, participants switch F/TAF to oral TAF (TG-A) or bictegravir  (TG-B); oral daily LEN + F/TAF (TG-C), or oral daily B/F/TAF (TG-D). Genotypic analyses (population sequencing) of HIV-1 reverse transcriptase and integrase, and genotypic (deep sequencing)/phenotypic analyses for CA were performed at screening; genotypic and phenotypic analyses were conducted at confirmed virologic failure. CALIBRATE Study Design182 participants were randomized and dosed in TG-A to D . Most participants had subtype B HIV-1 (92.9%). Sequence analysis of baseline samples found 65% of amino acid residues were conserved with < 1% variation across CA overall, and 55% of residues were fully conserved. No mutations were detected at 6 positions in CA associated with reduced susceptibility to LEN in vitro; residues were fully conserved at 5 positions , and < 2% variation was observed at 1 position (T107). Three participants met the criteria for resistance analysis: 2 participants resuppressed to < 50 copies/mL while continuing treatment. One participant on SC LEN + F/TAF developed emergent resistance to LEN (Q67H+K70R) and emtricitabine (M184M/I), followed by resuppression after starting dolutegravir, zidovudine + lamivudine, tenofovir disoproxil fumarate. Emergent resistance to LEN was uncommon in treatment-na\u00efve participants receiving SC or oral LEN . These interim resistance findings support the ongoing evaluation of LEN for treatment and prevention of HIV.Laurie VanderVeen, PhD, Gilead Sciences  Nicolas Margot, MA, Gilead Sciences  Vidula Naik, MSc, Gilead Sciences  Silvia Chang, Masters, Gilead Sciences, Inc  Ross Martin, PhD, Gilead Sciences, Inc  Hadas Dvory-Sobol, PhD, Gilead Sciences  Martin Rhee, MD, Gilead Sciences  Christian Callebaut, PhD, Gilead Sciences"}
+{"text": "The vaccine is also licensed in ages 12 months and older in EU and certain other countries. We evaluated the safety and immunogenicity of MenACYW-TT compared to a licensed quadrivalent conjugate meningococcal vaccine (MenACWY-DT [Menactra\u00ae]) in Japanese children, adolescents and adults (2-55 years of age).MenACYW-TT [MenQuadfiA phase III modified double-blind, randomized study (NCT04368429) to evaluate the immunogenicity and safety of a single dose of MenACYW-TT versus MenACWY-DT was conducted in 360 participants (ratio 1:1) between ages 2 and 55 years in Japan. Serum bactericidal assays with human complement (hSBA) were used to measure antibodies against vaccine serogroups at baseline (Day 0) and 30 days post-vaccination (D30). Safety data were collected up to 30 days post-vaccination.Non-inferiority of immune responses for all four serogroups, based on percentages of participants achieving hSBA vaccine seroresponse as primary endpoint, was demonstrated for MenACYW-TT compared to MenACWY-DT at Day 30 in comparison to baseline: 85.6% vs 65.4% for serogroup A, 96.6% vs 62.6% for serogroup C, 87.4% vs 49.2% for serogroup W, and 97.7% vs 63.5% for serogroup Y. The proportions of individuals with hSBA titers \u2265 1:8 following MenACYW-TT administration were higher than those after MenACWY-DT administration for serogroups C (98.9% vs 81.0%), W (99.4% vs 91.1%) and Y (100 % vs 89.4%) and comparable for serogroup A (96.6% vs 92.7%). The hSBA GMTs were higher following administration of MenACYW-TT for all four serogroups. Immunogenicity results in participants 10 to 17 years of age and \u2265 18 years of age were comparable to those in the whole population (2-55 years). The safety profiles of MenACYW-TT and MenACWY-DT were comparable. There were no immediate adverse events (AEs), no AEs leading to study discontinuation, and no vaccine-related serious adverse events reported in the study.MenACYW-TT vaccine was well tolerated and demonstrated a non-inferior immune response compared to that for the licensed MenACWY-DT vaccine when administered as a single dose to meningococcal vaccine-na\u00efve children, adolescents, and adults in Japan.Osamu Matsuoka, MD, Sanofi Pasteur  Mugen Ujiie, MD, Sanofi Pasteur  Hitoshi Kikuchi, MD, Sanofi Pasteur (Scientific Research Study Investigator)Sanofi Pasteur (Research Grant or Support) Danaya Chansinghakul, MD, Sanofi Pasteur (Employee) Takahiro Inoue, MSc, Sanofi Pasteur (Employee) Kucku Varghese, PhD, Sanofi Pasteur (Employee) Nuchra Sirisuphmitr, MSc, Sanofi Pasteur (Employee) Tomoyuki Hashiguchi, MSc, Sanofi Pasteur (Employee) Betzana Zambrano, MD, Sanofi Pasteur (Employee) Takahiro Nakama, MD, Sanofi Pasteur (Employee) Carina Frago, MD, Sanofi Pasteur  Emilia Jordanov, MD, Sanofi Pasteur  Mandeep S. Dhingra, MD, Sanofi Pasteur"}
+{"text": "GRF gene family has been performed in cotton, and their functions in flowering are largely unknown.In plants, 14-3-3 proteins, also called GENERAL REGULATORY FACTORs (GRFs), encoded by a large multigene family, are involved in protein\u2013protein interactions and play crucial roles in various physiological processes. No genome-wide analysis of the GRF genes were identified in Gossypium herbaceum, G. arboreum, G. hirsutum, and G. raimondii, respectively, by genome-wide analyses and were designated as GheGRFs, GaGRFs, GhGRFs, and GrGRFs, respectively. A phylogenetic analysis revealed that these proteins were divided into \u03b5 and non-\u03b5 groups. Gene structural, motif composition, synteny, and duplicated gene analyses of the identified GRF genes provided insights into the evolution of this family in cotton. GhGRF genes exhibited diverse expression patterns in different tissues. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that the GhGRFs interacted with the cotton FLOWERING LOCUS T homologue GhFT in the cytoplasm and nucleus, while they interacted with the basic leucine zipper transcription factor GhFD only in the nucleus. Virus-induced gene silencing in G. hirsutum and transgenic studies in Arabidopsis demonstrated that GhGRF3/6/9/15 repressed flowering and that GhGRF14 promoted flowering.In this study, 17, 17, 31, and 17\u2009GRF genes were identified in cotton, and their gene and protein features, classification, evolution, and expression patterns were comprehensively and systematically investigated. The GhGRF3/6/9/15 interacted with GhFT and GhFD to form florigen activation complexs that inhibited flowering. However, GhGRF14 interacted with GhFT and GhFD to form florigen activation complex that promoted flowering. The results provide a foundation for further studies on the regulatory mechanisms of flowering.Here, 82 The online version contains supplementary material available at 10.1186/s12870-021-02923-9. The R package \u201cpheatmap\u201d was used to generate the expression patterns of the GhGRF genes in these tissues.To obtain the expression profiles of A490626) . The expGhGRF genes were independently cloned into the pMD19-T vector, and positive plasmids containing each gene, as identified by sequences analyses, were then introduced into the plant binary vector pCAMBIA2300-35S-OCS [35S:GhGRF3, 35S:GhGRF6, 35S:GhGRF9, 35S:GhGRF14, and 35S:GhGRF15. All of the constructs were introduced independently into Agrobacterium tumefaciens GV3101 and transformed into A. thaliana Col-0 using the floral dip method [The coding sequences of the five -35S-OCS  to generp method . FloweriBD-GhFT vector used in this study was previously published [GhFD and 17\u2009GhGRFs were amplified by PCR using gene-specific primers . The ublished . The codeviously .GhFD and five GhGRFs were amplified using gene-specific primers Additional file 4: Fig. S1. Chromosomal distributions of the Gossypium spp. GRF genes. (a) G. herbaceum. (b) G. arboreum. (c) G. hirsutum. (d) G. raimondii. Chromosomal locations were shown from top to bottom on corresponding chromosomes according to cotton genome annotation.Additional file 5: Fig. S2. Multiple amino acid sequence alignment of cotton GRF proteins. Amino acid sequence alignment of 82 GRF proteins from G. herbaceum, G. arboreum, G. hirsutum, and G. raimondii. Nine \u03b1-helices were marked as \u03b11-\u03b19.Additional file 6: Table S4. Seven conserved amino acid motifs and annotation of cotton GRF proteins.Additional file 7: Table S5. Syntenic gene pairs between G. hirsutum and other three cotton species.Additional file 8: Table S6. The duplicated GRF gene pairs from G. herbaceum, G. arboreum, G. hirsutum, and G. raimondii, respectively.Additional file 9: Fig. S3. Distributions of Ka/Ks values of cotton GRF gene pairs. (a) Ka/Ks ratios for paralogous genes in four cotton species. (b) Boxplot showing the Ka/Ks ratios for orthologous genes among cotton genomes. The center line in each box indicates the median, and the box limits indicate the upper and lower quartiles of divergence.Additional file 10: Fig. S4. The percentage of various responses about cis-acting elements on 2.0-kb promoter of the GRF genes in (a) G. herbaceum, (b) G. arboreum, (c) G. hirsutum, and (d) G. raimondii.Additional file 11: Fig. S5. Information of type, quantity, and location of various response elements in cotton.Additional file 12: Table S7. The FPKM value of GhGRFs in different tissues.Additional file 13: Fig. S6. Interaction of GhFT and each GhGRF protein. Plasmids transformed into AH109 yeast strains were screened on the -LT and -LTHA medium.Additional file 14: Fig. S7. Flowering times and the expression levels of the floral-meristem identity genes in the control and GhGRF silencing plants. (a\u2013e) Statistics of flowering times in the control and GhGRF silencing plants. qRT-PCR analysis of the expression levels of GhAP1 (Gh_D13G0878) (f\u2013j) and GhSOC1 (Gh_A11G0755) (k\u2013o). A cotton Ubiquitin7  gene was used as an internal reference gene. Values are means \u00b1 SD (n\u2009=\u20093). Asterisks indicate significant differences between control TRV:00 and TRV:GhGRFs silencing lines .Additional file 15: Fig. S8. qRT-PCR expression analysis of GhGRF, AtSOC1, and AtAP1. ACT2 (At3g18780) was used as an internal reference transcript. Values are means \u00b1 SD (n\u2009=\u20093). Asterisks indicate significant differences between Col-0 and the 35S:GhGRFs transgenic lines .Additional file 16: Table S8. PCR primers used in this study."}
+{"text": "Scrophulariae Radix (SR) has an important role as a medicinal plant, the roots of which are recorded used to cure fever, swelling, constipation, pharyngitis, laryngitis, neuritis, sore throat, rheumatism, and arthritis in Asia for more than two thousand years. In this paper, the studies published on Scrophularia buergeriana (SB) and Scrophularia ningpoensis (SN) in the latest 20 years were reviewed, and the biological activities of SB and SN were evaluated based on in vitro and in vivo studies. SB presented anti-inflammatory activities, immune-enhancing effects, bone disorder prevention activity, neuroprotective effect, anti-amnesic effect, and anti-allergic effect; SN showed a neuroprotective effect, anti-apoptotic effect, anti-amnesic effect, and anti-depressant effect; and SR exhibited an immune-enhancing effect and cardioprotective effects through in vitro and in vivo experiments. SB and SN are both known to exert neuroprotective and anti-amensice effects. This review investigated their applicability in the nutraceutical, functional foods, and pharmaceutical industries. Further studies, such as toxicological studies and clinical trials, on the efficacy and safety of SR, including SB and SN, need to be conducted. Scrophularia consists of more than 300 different herbs, and Scrophularia buergeriana (SB) and Scrophularia ningpoensis (SN) are representative plants plants of this genus. SB is called \u201cHyun-sam\u201d in Korea; it is a perennial plant with a strong fragrance that grows up to 1.8 m [Scrophulariae Radix (SR), specifically, SB and SN. The physiological properties discussed in this review have been verified using in vitro and in vivo studies, and these results are the scientific basis for the development of health foods or therapeutics.The genus to 1.8 m . It is nto 1.8 m ,5. SN isto 1.8 m . In thisLee et al.  and JeonMost of the compounds isolated from SN were found in the roots and isolated by solvent fractionation. Iridoids and iridoid glycosides ,27,28,29p < 0.01), IL-6 (SBE: p < 0.01), and MMP-9  mRNA expression levels in LPS-induced Raw 264.7 cells. In addition, SBE significantly suppressed the expression of MMP-9  and p65 phosphorylation . The mRNA expression levels of IL-6 and TNF-\u03b1 in LPS-induced Raw 264.7 cells were reduced by SBE treatment with Bay11-7085 (NF-\u03baB inhibitor).Shin et al.  reportedp < 0.05), IL-2 , and IL-4  in MOLT-4 cells. Moreover, IgG (p < 0.05) production in the SNU 265 human B cell line increased with 1 mg/mL SB treatment. The cells treated with SB and SB plus IFN-\u03b3 showed increased IL-12  production and induced NO  level and iNOS  expression in mouse peritoneal macrophages.Kim et al.  demonstrp < 0.01) treatment increased the cell viability in lymphocytes. In addition, ERS markedly decreased cAMP/cGMP (1 mg/mL: p < 0.05), IFN-\u03b3/IL-10 (1 mg/mL: p < 0.01), and MDA content (1 mg/mL: p < 0.01) and increased the SOD content (1 mg/mL: p < 0.01) in lymphocytes isolated from ICR mice spleen.Gong et al.  demonstrp < 0.001) suppressed TRAP-positive cell formation at 200 \u03bcg/mL concentration and had a resorption inhibition effect on mature osteoclasts. The resorption area (p < 0.001) was also observed to decrease after SB treatment.Nam et al.  reportedp < 0.01) in SH-SY5Y cells with glutamate-induced cell toxicity. Acetylcholinesterase activity  was decreased, and total glutathione content (SBE: p < 0.01) was increased in a dose-dependent manner. Glutamate-induced cell morphology changes and DNA fragmentation were measured using DAPI staining and TUNEL assays, and SBE was observed to reduce glutamate-induced fragmentation (SBE: p < 0.01).Lee et al.  demonstrp < 0.01), SOD-2 (SBE: p < 0.05), and GPx-1 (SBE: p < 0.01) expression levels, but it decreased Bax (SBE: p < 0.01) protein, cleaved caspase-3 , and cleaved PARP (SBE: p < 0.01) levels. SBE treatment also reduced the phosphorylation of p38 . In contrast, 500 \u03bcg/mL of SBE significantly (p < 0.01) increased B-cell lymphoma-2 (Bcl-2) expression levels.Increased antioxidant enzyme activity is known to protect nerve cells by reducing oxidative neuronal damage. SOD-1, SOD-2, and GPx-1 are antioxidant enzymes, and when reduced, they promoted the expression of apoptosis factors such as Bcl-2-associated X (Bax), cleaved caspase-3, and cleaved poly (adenosine diphosphate (ADP)-ribose) polymerase (PARP). SBE treatment markedly increased SOD-1  demonstrated the neuroprotection effects in PC12 cells. RSAE differently increased according to pretreatment hour. Pretreatment RSAE for 4 h did not affect the cell viability and pretreatment RSAE for 8 h (12.5 \u03bcg/mL: p < 0.0.05), 16 h (12.5 \u03bcg/mL: p < 0.05), and 24 h (6.25\u201325 \u03bcg/mL: p < 0.00001) statistically increased the cell viability. LDH (RSAE 12.5 \u03bcg/mL: p < 0.0001), MDA (RSAE 12.5 \u03bcg/mL: p < 0.001), and NO (RSAE 12.5 \u03bcg/mL: p < 0.01) levels are decreased and SOD (RSAE 12.5 \u03bcg/mL: p < 0.05), CAT (RSAE 12.5 \u03bcg/mL: p < 0.05), and GSH-Px (RSAE 12.5 \u03bcg/mL: p < 0.0001) activities are recovered with RSAE in oxygen-glucose deprived and reperfusion (OGD/R)-induced PC12 cells. JC-1 red fluorescence for mitochondrial membrane potential change detection was significantly increased by treating with 12.5 \u03bcg/mL RSAE in PC12 cells. OGD/R treatment induced MMP destruction and significantly suppressed the red/green fluorescence ratio, but it significantly (RSAE 12.5 \u03bcg/mL: p < 0.0001) enhanced the red/green fluorescence ratio.Meng et al.  reportedScrophularia ningpoensis water extract (RSN) presented the anti-apoptotic activity in HaCaT cells. RSN showed the IC50 value at 0.032 mg/mL concentration. TNF-a stimulation activates the NF-\u03baB pathway and induces inflammation. In addition, ERK upstream of NF-\u03baB affects cell proliferation and induces apoptosis. Pretreatment with 0.032 mg/mL RSN inhibited NF-\u03baB translocation induced by TNF-\u03b1, which was observed using immunofluorescence staining of HaCaT cells. RSN decreased ERK phosphorylation, and ERK increased gradually with dose and time. RSN did not affect the cell cycle phase in G1/G0, S, and G2/M (at 0.016\u20130.064 mg/mL for 6\u201396 h) in HaCaT cells, and a tumor-preferred effect was not detected in Colo 38, SK-Mel-28, and MRI-221 cells. These results suggest that RSN regulates ERK and NF-\u03baB signaling.Shen et al.  demonstrp < 0.05, SBE 1000 \u03bcg/mL: p < 0.001) and histamine  in RBL-2H3 cells. The release of pro-inflammatory cytokines, TNF-\u03b1 (p < 0.01) and IL-4 (p < 0.001), after antigen induction in RBL-2H3 cells was decreased after 1000 \u03bcg/mL SBE treatment. Furthermore, the cells treated with 100 and 1000 \u03bcg/mL SBE showed inhibition of ERK  and p38 phosphorylation .Kim et al.  reportedp < 0.01), macrophages (SBE: p < 0.01), neutrophils , lymphocytes number , and total cells (SBE: p < 0.01) in bronchoalveolar lavage fluid (BALF) and also decreased airway hyper-responsiveness (SBE 20 mg/kg with methylcholine (MC) 30 mg/mL: p < 0.05, SBE 40 mg/kg with MC 20 mg/mL: p < 0.05, SBE 40 mg/kg with MC 30 mg/mL: p < 0.01).Shin et al.  reportedp < 0.01), IL-13 , and IL-17  in BALF and total IgE  and OVA-specific IgE  levels in the serum decreased after SBE treatment. SBE treatment significantly decreased the inflammatory index and mucus production index  in the lung tissue. In the asthma model induced by OVA, SBE administration decreased MMP-9 (SBE: p < 0.01) expression and p65 phosphorylation (SBE: p < 0.01).The levels of pro-inflammatory cytokines, including IL-5 , as well as treatment with 3\u2013200 mg/kg KD-501  for 15 days significantly increased the step-through latency in the passive avoidance test. Acute and prolonged (for 15 days) treatment of 100 mg/kg KD-501 improved spatial memory ability by reducing escape latency in the Morris water maze test conducted for 4 days. The acetylcholinesterase activity of the cortex (KD-501: p < 0.05) and hippocampal  tissue was significantly increased by the acute administration of KD-501.Jeong et al.  reportedp < 0.001 in hippocampus and increasing glutathione reductase  and SOD  activities in the cortex and hippocampal tissue of amnesic mice. The prolonged (for 15 days) oral treatment with 100 mg/kg KD-501 showed antioxidant activity by decreasing GSSG/total GSH (KD-501: p < 0.001) and increasing glutathione reductase (KD-501: p < 0.001) and SOD (KD-501: p < 0.001) activities in the cortex and hippocampus.Acute treatment with 100 mg/kg KD-501 resulted in antioxidant activity by decreasing GSSG/total GSH (KD-501: These results showed that KD-501 may be used for the prevention and therapeutics of Alzheimer\u2019s disease.p < 0.05) in the passive avoidance test that decreased after scopolamine injection was significantly increased by treatment with 30 and 100 mg/kg SBE, and escape latency and swim distance were decreased in the Morris water maze test. Moreover, in the probe trial conducted on day 28, administration of SBE (30 and 100 mg/kg) significantly increased the crossing number (SBE: p < 0.01). The decreased acetylcholine  level was increased, while increased acetylcholinesterase (SBE 100 mg/kg: p < 0.01) activity was decreased in the hippocampus after SBE (30 and 100 mg/kg) administration.Lee et al. ,69 demonp < 0.01) expression level, and 30 and 100 mg/kg SBE administration significantly increased CREB (SBE: p < 0.01) phosphorylation. Furthermore, SOD-1 (SBE: p < 0.01) and SOD-2 (SBE 100 mg/kg: p < 0.05) expression levels were increased and IL-1\u03b2 (SBE 100 mg/kg: p < 0.01), IL-6 , and TNF-\u03b1 (SBE 100 mg/kg: p < 0.01) gene expression levels were decreased in mice with SBE. The group injected with scopolamine showed increased expression levels of Bax, cleaved caspase-9, and cleaved PARP and decreased expression level of Bcl-2. The SBE treatment decreased Bax (SBE: p < 0.01), cleaved caspase-9 (SBE 100 mg/kg: p < 0.01), and cleaved PARP (SBE: p < 0.01) expression levels and increased Bcl-2 (SBE 100 mg/kg: p < 0.01) expression level.Scopolamine injection reduced the BDNF expression levels and CREB phosphorylation. However, 100 mg/kg SBE treatment markedly increased the BDNF  and decreased escape latency and swim distance in the Morris water maze test. The crossing number  was also increased after SBE treatment. Glutathione reductase activity (SBE: p < 0.01) and SOD-1 (SBE 100 mg/kg: p < 0.01), SOD-2 (SBE 100 mg/kg: p < 0.01), and GPx-1 (SBE 100 mg/kg: p < 0.01) expression levels were decreased by \u03b2-amyloid injection and increased in the mice administered SBE. Furthermore, Bax (SBE: p < 0.01), cleaved caspase-9 (SBE: p < 0.01), and cleaved PARP (SBE: p < 0.01) levels were markedly decreased, and Bcl-2 (SBE 100 mg/kg: p < 0.01) expression level was increased after SBE treatment. The \u03b2-amyloid (SBE: p < 0.01) and phosphorylation of Tau (SBE: p < 0.01) were significantly decreased in the mice treated with SBE. According to the in vitro and in vivo studies, SBE improved spatial memory and cognitive ability by inhibiting cell apoptosis.The mice injected with \u03b2-amyloid showed decreased step-through latency in the passive avoidance test and increased escape latency and swim distance. SBE treatment (30 and 100 mg/kg) in these mice increased the step-through latency  and MDA (RSAE 2.4 g/kg: p < 0.01) and NO (RSAE 2.4 g/kg: p < 0.01) levels in the ischemic hemisphere. 2,3,5-Triphenyltetrazolium chloride (TTC) staining presented that the corrected infarct volume increased by MCAO/R operation was significantly decreased by RSAE (2.4 g/kg: p < 0.0001) administration. In the cortex and CA1 region, significantly increased neuronal damage due to MCAO/R was observed compared to the sham group. RSAE treatment significantly decreased neuronal damage by increasing neuron density in the ischemic cortex (RSAE 2.4 g/kg: p < 0.01) and hippocampus CA1 region (RSAE 2.4 g/kg: p < 0.01).Meng et al.  reportedp < 0.05) expression and increased Bcl-2 (RSAE 2.4 g/kg: p < 0.0001) expression in focal cerebral ischemia and markedly decreased the phosphorylation of ERK1/2 (RSAE 2.4 g/kg: p < 0.05), p38 MAPK (RSAE 2.4 g/kg: p < 0.01), and JNK1/2 (RSAE 2.4 g/kg: NS). According to the in vitro and in vivo studies, RSAE inhibits apoptosis and exhibits neuroprotective effects by regulating the MAPK pathway.The MCAO/R group showed increased cell apoptosis by an increase in Bax-positive cells and a decrease in Bcl-2-positive cells. However, RSAE significantly reduced Bax  remarkably decreased the number of escape failures and had an anti-depressive effect on the animal model.Xu et al.  demonstrThese results suggest that II could be used for the treatment of depression.Kim et al.  reportedp < 0.05, SBE 100 \u03bcg at 48 h: p < 0.01, SBE 100 \u03bcg at 72 h: p < 0.001, 1000 \u03bcg at 48, 72 h: p < 0.001) and weight  and decreased TNF-\u03b1  and IL-4 (SBE 1000 \u03bcg: p < 0.01) levels in ear homogenates in a dose-dependent manner.SBE application reduced ear thickness  has a ventricular remodeling effect. Left ventricular remodeling was induced in rats orally administered with 60, 120, and 240 mg/kg EERS for 14 weeks. The ventricular remodeling group showed decreased systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial pressure (MAP) when compared with the sham-operated control. The ERRS-treated rats showed increased SBP (CAL plus 120 mg/kg EERS: p < 0.05), DBP , and MAP , and their increased heart rate  was significantly reduced. The hemodynamic parameter left ventricular systolic pressure  was remarkably increased by ERRS. Increased by left coronary artery ligation (CAL), cardiac weight indexes such as the left ventricular weight index (LVWI)  and heart weight index (HWI)  and cardiocyte cross-section area  were decreased in the ERRS-treated group.Huang et al.  demonstrp < 0.01) and I/III collagen  in the interstitial space were remarkably decreased. The interstitial collagen volume fraction (ICVF)  and perivascular collagen volume fraction were also significantly reduced with ERRS treatment. The serum angiotensin II (Ang II)  concentration was decreased in the ERRS-treated rats. Endothelin (ET)-1 , atrial natriuretic peptide (ANP) , hydroxyproline (Hyp) , MMP-2 , and TNF-\u03b1  concentrations were decreased by ERRS administration. The rats with ventricular remodeling that were administered ERRS showed a significant decrease in angiotensin-converting enzyme (ACE) , ET-1 , and ANP  mRNA expression levels.The ERRS-treated group showed that types I and III collagen volume  possesses ventricular remodeling inhibition effects. Spontaneously hypertensive (SHR) male rats were used for the experiment and orally treated with 70, 140, and 280 mg/kg ACRS for 21 weeks. The cardiac mass indexes LVWI , HWI , and ICVF , perivascular collagen area (PVCA)  ratio, collagen I  and III  volumes, I/III ratio , serum norepinephrine , myocardium Ang II , and serum TNF-\u03b1 levels were significantly decreased by ACRS treatment in the SHR rats.Zhang et al.  showed tp < 0.01), TGF-\u03b21 , and ACE  mRNA expression levels and decreased phosphorylation of p44/42 MAPK , SANP/JNK , and p38 MAPK . These results show that ACRS inhibited ventricular remodeling through MAPK pathway regulation, and it may be used to prevent and treat heart failure.The ACRS-treated SHR rats showed reduced collagen I , HWI , myocardium Ang II , and Hyp (ERS-L and H: p < 0.01) decreased with the ERS treatment. To analyze the cardiocyte cross-section area (ERS-L and H: p < 0.01) by H&E staining, ERS administration was reduced similarly to that in the captopril group used as the positive control. In addition, PVCA (ERS-L and H: p < 0.01) and CVF (ERS-L and H: p < 0.01), which are increased by ventricular remodeling, were significantly decreased by the ERS treatment.Gu et al.  showed tp < 0.01) and III collagen contents  were significantly decreased and AT1R (ERS-L and H: p < 0.01), TNF-\u03b1 (ERS-L and H: p < 0.01), and TGF-\u03b21 (ERS-L and H: p < 0.01) mRNA expression levels were reduced in the myocardium by the ERS treatment.Subtypes I  was observed to significantly increase, and body temperature (ERS: p < 0.05), heart rate (ERS: p < 0.05), average speed (ERS: p < 0.01), and upright time (ERS: p < 0.01) were found to markedly decrease. Measurement of serum MDA and SOD levels and cAMP/cGMP suggested that ERS treatment decreased the MDA (ERS: p < 0.05) level and cAMP/cGMP (ERS: p < 0.05) and increased SOD (ERS: p < 0.05) level. The ERS-treated mice showed a statistical decrease in IL-6 (ERS: p < 0.01) expression level in the serum and Na+-K+ATP enzyme content (ERS: p < 0.01) in the liver homogenates. According to the in vitro and in vivo studies, ERS showed its immune regulation and antioxidant activities, and it can be used as dietary supplement for better health.Gong et al.  reportedIn this review, we summarized the biological effects of SB, SN, and SR based on the previous in vitro and in vivo studies. SB was administered orally at a minimum dose of 20 mg/kg to a maximum of 200 mg/kg (mice), SN was administered orally at a minimum dose of 5 mg/kg to a maximum of 2.4 g/kg (mice), and SR was orally administered at 2 g/kg (rat). SB showed anti-inflammation activity, immune-enhancing effects, bone disorder prevention activity, neuroprotective effect, anti-amnesic effect, and anti-allergic effect. SN exhibited neuroprotective effect, anti-apoptotic effect, anti-amnesic effect, cardioprotective effect, and anti-depressant effect; and SR showed immune-enhancing activity and cardioprotective effects. However, clinical trials need to be conducted to investigate the efficacy and safety of SR, including SB and SN.As per studies published on SB, SN, and SR in the last 20 years, these plants have been processed using water or ethanol as a solvent. This is thought to reduce side effects when administered orally, and will help increase their applicability to various formulations when developing health functional foods or therapeutics in the future.Taken together, we suggested the possibility of utilizing SB, SN, and SR for developing health functional foods or therapeutics for various applications on the basis of previously reported literature."}
+{"text": "Scientific Reportshttps://doi.org/10.1038/s41598-018-29965-8, published online 02 August 2018Correction to: The original version of this Article contained errors in the Reference list. The authors omitted the below paper, which is listed as Reference 22.Kirshenbaum AS, Akin C, Wu Y, Rottem M, Goff JP, Beaven MA, Rao VK, Metcalfe DD. Characterization of novel stem cell factor responsive human mast cell lines LAD 1 and 2 established from a patient with mast cell sarcoma/leukemia; activation following aggregation of FcepsilonRI or FcgammaRI. Leuk Res. 2003 Aug;27(8):677\u201382.As a result, in the Materials and Methods section,\u201cThe LAD2 human mast cell line was kindly provided by Dr. D. Metcalfe, (NIH Washington).\u201dnow reads:22.\u201d\u201cThe LAD2 human mast cell line was kindly provided by Dr. D. Metcalfe, (NIH Washington)Consequently, References 23\u201326 were incorrectly listed as References 22\u201325.The original Article has been corrected."}
+{"text": "Xanthomonas axonopodis pv. manihotis (Xam) seriously affects cassava yield. Genes encoding nucleotide-binding site (NBS) and leucine-rich repeat (LRR) domains are among the most important disease resistance genes in plants that are specifically involved in the response to diverse pathogens. However, the in vivo roles of NBS-LRR remain unclear in cassava (Manihot esculenta). In this study, we isolated four MeLRR genes and assessed their expression under salicylic acid (SA) treatment and Xam inoculation. Four MeLRR genes positively regulate cassava disease general resistance against Xam via virus-induced gene silencing (VIGS) and transient overexpression. During cassava-Xam interaction, MeLRRs positively regulated endogenous SA and reactive oxygen species (ROS) accumulation and pathogenesis-related gene 1 (PR1) transcripts. Additionally, we revealed that MeLRRs positively regulated disease resistance in Arabidopsis. These pathogenic microorganisms include Pseudomonas syringae pv. tomato, Alternaria brassicicola, and Botrytis cinerea. Our findings shed light on the molecular mechanism underlying the regulation of cassava resistance against Xam inoculation.Cassava bacterial blight (CBB) caused by  R genes) usually act as receptors of pathogen-encoded effector proteins, which are often secreted by pathogens directly into host cells  of NBS-LRR proteins usually contain the Toll/interleukin-1 receptor-like (TIR) domain, coiled-coil (CC) domain, or resistance to powdery mildew 8 (RPW8) domain, and the carboxyl terminus (C-terminus) contain a zinc-finger transcription factor-related domain containing the WRKY sequence (WRKY domain) , CC-NBS-LRR (CNL), and RPW8-NBS-LRR (RNL) proteins (Disease resistance genes (st cells . R genesomycetes . In the  species . Among tn plants . The ami domain) . Based oproteins .NBS-LRR-coding genes  genome  (Manihot esculenta) (Sorghum bicolor) (Triticum aestivum) (Gossypium hirsutum) (Zea mays) (Glycine max) (Vitis vinifera) (Dioscorea rotundata) (PR) genes (NB-ARC domain-containing (GbaNA1) gene impaired cotton resistance to Verticillium dahliae Vd991  genome , constit) genome . In the  indica) , cassavaculenta) , sorghumbicolor) , wheat (estivum) , cotton irsutum) , maize (ea mays) , soybeanine max) , grapeviinifera) , and yamtundata) . In recetundata) . ActivatR) genes . In cottae Vd991 . Similaraffected . NBS-LRRaffected . Yoodee  cassava , their rX. axonopodis pv. manihotis = X. phaseoli pv. manihotis)  and anthracnose disease (Colletotrichum gloeosporioides)  , cassavarioides) . Therefo viruses . A clustelection . HoweverNBS-LRR genes that showed high transcription level after pathogen infection attracted our attention. The expression levels of four chosen MeLRRs were significantly induced by exogenous application of SA treatment and Xam inoculation. Moreover, these genes positively regulated cassava resistance to Xam inoculation. The functional analysis of MeLRR genes will offer potential roles in genetic breeding for disease-resistant cassava.In this study, we analyzed the published transcriptome databases of cassava-pathogens interaction . Within NBS-LRRs in cassava, including both TIR-NBS-LRR and CC-NBS-LRR. Their transcript levels were analyzed through RNA-seq in response to CBSV and C. gloeosporioides infection  were both induced under CBSV and C. gloeosporioides infection and selected for further analysis. The four MeLRR proteins have typical leucine-rich repeats, which are named MeLRR1 (Manes. 11G053000.1), MeLRR2 (Manes. 03G071700.1), MeLRR3 (Manes. 13G036800.1), and MeLRR4 (Manes. 07G107800.1), located on chromosomes 11, 3, 13, and 7, respectively. MeLRR1, MeLRR3, and MeLRR4 belong to CC-NBS-LRR protein, while MeLRR2 is one of the TIR-NBS-LRR protein. Bioinformatics predicted that the MeLRR proteins were unstable and hydrophilic  leaves were infected with 35S:GFP or 35S:GFP-MeLRR1,\u22122,\u22123,\u22124 plasmid as described by 35S:GFP  of MeLRRs were cloned and inserted into the poly-cloning sites of the fusion expression vector pEGAD and fused upstream to a green florescence protein (GFP) fusion partner by the constitutive  35S:GFP .MeLRRs in response to SA treatment and Xam inoculation were analyzed by qRT-PCR . Under SA treatment, the expressions of MeLRR1, MeLRR3, and MeLRR4 were induced and peaked at 1 h post treatment (hpt), while MeLRR2 showed the highest level at 3 hpt . The partial sequences of MeLRR1 (453 bp), MeLRR2 (441 bp), MeLRR3 (433 bp) and MeLRR4 (423 bp) were individually inserted into pTRV2 plasmid to construct VIGS vector. At 14 days post-infection (dpi) in cassava infected with Agrobacterium GV3101 carrying the pTRV-MeLRR plasmids, qRT-PCR was performed to detect the target gene transcript level. The transcript level of the target MeLRR-1,-2,-3,-4 genes were significantly decreased in the MeLRR-silenced cassava leaves compared to the pTRV empty vector. The silencing efficiency of MeLRR-1,-2,-3,-4 was 46.33 (\u00b1 2.31)%, 15.28 (\u00b1 0.49)%, 30.22 (\u00b1 2.28)%, and 17.45 (\u00b1 0.87)% , respectively  in one VIGS line, the transcript levels of all four target genes were significantly decreased  gene and B. cinerea BcActA (XM_024697950.1) gene  gene  with theectively .AtICS1, AtPDF1.2, AtPR1, AtPR2, AtPR5, and AtTGA3 in WT and MeLRR overexpression lines upon A. brassicicola, B. cinerea, and P. syringae pv. tomato DC3000 infection , and its production in plants represents the successful recognition of pathogen infection and pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) and effector-triggered immunity (ETI) . In casse to CBB . TherefoXam regulate plant immune  effectors and type III effectors (T3Es) of t immune . Such asune , variety South China 124 (SC124), and N. benthamiana were cultivated in mixed soil  in a greenhouse with 16/8 h light/dark at 28/22\u00b0C, 60\u201370% relative humidity with irradiance of 130\u2013150 uE.m\u20132.s\u20131. A. thaliana ecotype Col-0 (Columbia-0) seedlings were cultivated in the mixed soil under fluorescent light (130\u2013150 uE.m\u20132.s\u20131) and were grown under 16/8 h light/dark at 22\u00b0C. For axenic growth, N. benthamiana, and A. thaliana seeds were sterilized  and sown on half-strength MS (Murashige and Skoog) medium  with 0.4% agar powder and 2% (w/v) sucrose. The seeds were grown in chambers under 16/8 h light/dark at 22\u00b0C and 130\u2013150 uE.m\u20132.s\u20131. For expression analysis, 4-week-old cutting seedlings of cassava leaves were sprayed with 5 mmol/L salicylic acid or Xam suspension for 0, 1, 3, 6, 12, or 24 h, and the bacterial solution was diluted to 4 \u00d7 108 colony-forming units/mL (cfu/mL) using 10 mmol/L MgCl2 with 0.05% Silwet L-77.Cassava  , Durio zibethinus (XP_022746274.1), H. brasiliensis , J. curcas , Populus alba , P. euphratica (XP_011001622.1), P. trichocarpa (RQO87881.1), Ricinus communis (EEF44774.1), Theobroma cacao (XP_01796 9995.1), Vernicia montana (AMM43068.1), V. vinifera (XP_010657.1), and Ziziphus jujuba (XP_024924720.1), respectively.The sequences of me v131) . The Proty index . Alignmety index . The 24 Arabidopsis and cassava gene transcripts were normalized to the AtAct2 gene (AT3G18780) and elongation factor 1\u03b1  using the comparative 2\u2013\u0394\u0394Ct method, respectively  of A. brassicicola were designed by Primer3Plus3 to find optimal primers . cDNA synthesis was performed using the Tiangen FastQuant RT kit  with 20-\u03bcl reaction mixture. qRT-PCR analysis was performed using UltraSYBR Mixture (low ROX)  in an ABI QuantStudio\u2122 6 flex Real-Time PCR System . The PCR cycling conditions were 95\u00b0C for 10 min, followed by 40 cycles at 95\u00b0C for 15 s and 60\u00b0C for 1 min. The ectively . The qRT primers , and theMeLRR1,-2,-3,-4 were amplified and cloned into the pEGAD vector (Promoter CaMV35S:GFP) via appropriate restriction enzyme digestion and T4 DNA ligase. The recombinant plasmids and empty vector were transformed into Agrobacterium GV3101. Then, the A. tumefaciens suspension was used to infect the leaves of cassava or tobacco as described by 8 cfu/mL of pathogenic bacteria Xam used for disease resistance assay, include number of Xam populations, MePR1 transcript level, and symptoms of cassava bacterial blight at 0 and 1 dpi, respectively.For overexpression, the full-length coding regions of MeLRRs and the online siDirect 2.04 tools  for 24 h at room temperature (The cassava or perature . The leaperature .P. syringae pv. tomato (Pst) DC3000 was streaked on LB medium with 50 mg/L of rifampicin at 28\u00b0C and shaken to OD600 reached 0.6. Thereafter, a fresh bacterial culture of Pst DC3000 was diluted to 4 \u00d7 108 cfu/mL in 10 mmol/L MgCl2 and 0.05% Silwet L-77 and then sprayed on 24-day-old Arabidopsis leaves. The A. brassicicola and B. cinerea strains were cultured on potato dextrose agar (PDA) medium with 2% (w/v) sucrose at 28\u00b0C. Conidia were suspended in distilled water for plant infection. Spore suspensions (about 4 \u00d7 106 spores/mL) of A. brassicicola and B. cinerea were sprayed on Arabidopsis leaves. The infected plants were grown in an incubator at 90% RH and 22\u00b0C. At 0, 2, and 4 dpi, the number of Pst DC3000 bacteria was determined, as well as the fungal actin gene transcript in leaves of Col-0 and mutants infected with B. cinerea and A. brassicicola  was set at p < 0.05. Each assay contained three independent replicates.Mean and standard deviations are displayed as representative values for data in the figures. Analysis of variance (ANOVA) with Duncan\u2019s test and Student\u2019s The original contributions presented in the study are included in the article/YW and HZ designed the research. HZ did most experimental works and wrote the manuscript. ZY, ZL, and YS did experimental works and database analysis. XL, JW, and GZ did experimental works. YW supervised this project. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic developing since the late 2019 and early 2020 has caused thousands of deaths and has had an enormous impact on our health systems and economies. Thrombotic complications associated with coronavirus disease 2019 (COVID-19) have been described. Acute limb ischemia is associated with complications related to cytokine storm syndrome and acute respiratory distress syndrome. that can result in significant morbidity and mortality. However, limited published data is available regarding thrombosis in coronavirus disease 2019 (COVID-19). Here we are presenting 2 cases of COVID-19 infection complicated by arterial thrombosis in the form of acute limb ischemia. \u2022COVID-19 pneumonia increases the incidence of acute limb ischemia.\u2022Thrombotic complications associated with coronavirus disease 2019 can result in significant morbidity and mortality.\u2022Patients with COVID-19 should receive pharmacologic thromboprophylaxis. On March 11, 2020, WHO declared coronavirus disease 2019 (COVID-19) as pandemic [The association of COVID-19 with coagulopathy has gained increasing interest recently. As per American Society of Hematology, some patients with severe coronavirus disease 2019 (COVID-19) have fulminant activation of coagulation and consumption of coagulation factors, which meets the criteria for disseminated intravascular coagulation as per International Society on Thrombosis and Hemostasis  which in22.180 years old patient, diabetic, hypertensive, presented to emergency department with complaints of shortness of breath and chest pain, with a discoloration and swelling of the right leg. Vital signs upon arrival were: heart rate at 113 per minute, blood pressure at 140/85\u00a0mmHg, oxygen saturation at 40% on room air and temperature at 38,6\u00b0 Celsius.The patient had bilateral crackles on lung exam and absent right popliteal, dorsalis pedis and posterior tibial pulses and the foot was swollen, cyanotic, and cold .Fig. 1Cy3 (reference: 4.000\u201310.000/mm3), platelets 239.000/mm3 (reference: 150.000\u2013400.000/mm3), potassium 5.1 meq/L , glucose 3,35g/l , creatinine 26,2 mg/l (reference: 6\u201313 mg/l), blood urea nitrogen 1,6 g/l , lactic acid 2,37 mmol/L (reference: 0.5\u20132.2 mmol/L), troponin 1150 ng/l (reference: less than 6 ng/l), D-dimer 35,2 (reference: less than 0.5), prothrombin time 67,%  1.24 (reference: less than 1), Creatine Phosphokinase (CPK) 2080 U/L  lactate dehydrogenase (LDH) 1497 U/L (reference: 140\u2013271 U/L), C-reactive protein (CRP) 239 mg/L (reference: less than 10 mg/L), ferritin 1642 ng/L (reference: 12\u2013300 ng/l), procalcitonin 2,88 ng/ml (reference: less than 2 ng/ml), calcium 9,1 mg/dl (reference: 8.6\u201310.3 mg/dl), and albumin 29 mg/l (reference: 35\u201350 mg/l).On initial laboratory evaluation, the following values were noted: hemoglobin 11.1 g/dl (reference: 12\u201316 g/dl), hematocrit 33,6% (reference: 36%\u201346%), white blood cells 27.680/mmChest X ray showed bilateral hazy infiltrates . ThoraciCovid-19 was diagnosed on the basis of reverse transcription polymerase chain reaction (RT- PCR) testing. Echocardiogram showed an ejection fraction of 55%. The patient was intubated and put under SDRA ventilation and was started on moxifloxacin, ceftriaxone, azithromycin, hydroxychloroquine, and therapeutic anticoagulation with heparin. He underwent a thigh amputation.2.269 years old patient, suffering from chronic obstructive pulmonary disease, presented to emergency department with complaints of shortness of breath and chest pain, with a discoloration and swelling of the left leg. Vital signs upon arrival were: heart rate at 130 per minute, blood pressure at 150/95\u00a0mmHg, oxygen saturation at 75% on room air and temperature at 38,1\u00b0 Celsius.The patient had bilateral crackles on lung exam and absent left popliteal, dorsalis pedis and posterior tibial pulses and the foot was swollen, and cold.Electrocardiogram showed sinus tachycardia 140 per minute.3 (reference: 4.000\u201310.000/mm3), platelets 345.000/mm3 (reference: 150.000\u2013400.000/mm3), potassium 4,8 meq/L , glucose 1,16 g/l , creatinine 7,25 mg/l (reference: 6\u201313 mg/l), blood urea nitrogen 0,56 g/l , lactic acid 2,37 mmol/L (reference: 0.5\u20132.2 mmol/L), troponin 9,5 ng/l (reference: less than 6 ng/l), D-dimer 12,96 (reference: less than 0.5), prothrombin time 78%  1.13 (reference: less than 1), Creatine Phosphokinase (CPK) 680 U/L  lactate dehydrogenase (LDH) 1040 U/L (reference: 140\u2013271 U/L), C-reactive protein (CRP) 269 mg/L (reference: less than 10 mg/L), ferritin 854 ng/L (reference: 12\u2013300 ng/l), procalcitonin 0,51 ng/ml (reference: less than 2 ng/ml), calcium 8,3 mg/dl (reference: 8.6\u201310.3 mg/dl), and albumin 33 mg/l (reference: 35\u201350 mg/l).On initial laboratory evaluation, the following values were noted: hemoglobin 15.1 g/dl (reference: 12\u201316 g/dl), hematocrit 42,5% (reference: 36%\u201346%), white blood cells 21.560/mmChest X ray showed bilateral hazy infiltrates. Thoracic and Abdominal CT angiography of the aorta with Iliofemoral runoff showed an occluded left anterior tibial artery and left peroneal artery with no flow to the foot .Fig. 4PrCovid-19 was diagnosed on the basis of reverse transcription polymerase chain reaction (RT- PCR) testing. Echocardiogram showed an ejection fraction of 60%. The patient was admitted to intensive care unit and put under high flow nasal oxygenation (Optiflow) and was started on azithromycin, hydroxychloroquine, and therapeutic anticoagulation with heparin. He underwent a thrombectomy of left common femoral artery, profunda femoris, superficial femoral artery, popliteal artery, anterior tibial artery, posterior tibial artery, and peroneal artery .Fig. 5Su3The COVID-19 pandemic is a fast-evolving situation. The spectrum of clinical manifestations of SARS-CoV-2 infection includes fever, myalgia, cough and dyspnea, and less frequently headache, diarrhea, nausea and vomiting . SeveritThe hypercoagulability of critically ill patients with COVID-19 became evident. In addition, some published epidemiologic data have shown increased thromboembolic events in those patients, such as unusual ischemic limbs and venous thromboembolism ,9. A higCui et al.  in a retAcute limb ischemia is a vascular emergency and can result in severe morbidity, including chronic pain, limb loss, and severe disability. A prompt diagnosis is a prerequisite for successful treatment. An observational study done in Italy revealed increased incidence of acute limb ischemia in patients with COVID-19 infections, they have noted the number of patients with acute limb ischemia during the peak of the pandemic has increased .The mechanism of thromboembolic complications associated with COVID-19 infection is not fully understood. COVID-19 related hypercoagulability is likely multifactorial-direct viral infection of the endothelial cell leading to diffuse endothelial inflammation, increased procoagulant factors such as factor VIII, von Willebrand factor, fibrinogen, and high inflammatory state associated with the cytokine storm leading to coagulation and fibrinolysis activation .The American Society of Hematology (ASH) recommends that all hospitalized patients with COVID-19 should receive pharmacologic thromboprophylaxis with LMWH or fondaparinux, unless bleeding risk exists, and full therapeutic-intensity anticoagulation in the appropriate clinical scenario .4We report 2 cases of arterial thrombosis in patients with COVID-19. Our case and review of literature reveals that health care providers should be aware of this unusual life-threatening manifestation of COVID 19 so that appropriate measures can be taken for the vascular emergency.Written informed consent was obtained from the patient for publication of this case report and any accompanying images.This is a case report.This study was not funded.Amine Bensaid: Stady concept, data collection, Data analysis, writing the paper. Imane Melhaoui: writing the paper. Oujidi Younes: Contributor. Inass Arhoun El Haddad: Data collection. Houssam Bkiyar: Supervision and data validation. Brahim Housni: Supervision and data validation.DR Amine Bensaid.Not commissioned, externally peer reviewed.No conflict of interest."}
+{"text": "Neisseria meningitidis serogroup B and at a university where outbreak and response activities had not occurred. All eligible students at the two universities received the serogroup B meningococcal factor H binding protein vaccine (MenB-FHbp); 5.2% of students  at one university received MenB-4C. A total of 1,514 meningococcal carriage isolates were obtained from 8,905 oropharyngeal swabs from 7,001 unique participants. Whole-genome sequencing data were analyzed to understand MenB-FHbp\u2019s impact on carriage and antigen genetic diversity and distribution. Of 1,422 isolates from carriers with known vaccination status , 1,406 (98.9%) had intact fHbp alleles (716 from MenB-FHbp-vaccinated participants). Of 726 isolates from MenB-FHbp-vaccinated participants, 250 (34.4%) harbored FHbp peptides that may be covered by MenB-FHbp. Genogroup B was detected in 122/1,422 (8.6%) and 112/1,422 (7.9%) isolates from MenB-FHbp-vaccinated and unvaccinated participants, respectively. FHbp subfamily and peptide distributions between MenB-FHbp-vaccinated and unvaccinated participants were not statistically different. Eighteen of 161 MenB-FHbp-vaccinated repeat carriers (11.2%) acquired a new strain containing one or more new vaccine antigen peptides during multiple rounds of sample collection, which was not statistically different (P\u2009=\u20090.3176) from the unvaccinated repeat carriers . Our findings suggest that lack of MenB vaccine impact on carriage was not due to missing the intact fHbp gene; MenB-FHbp did not affect antigen genetic diversity and distribution during the study period.Carriage evaluations were conducted during 2015 to 2016 at two U.S. universities in conjunction with the response to disease outbreaks caused by  Neisseria meningitidis (the meningococcus) is a major public health concern  caused by  concern . This ba concern , 3. RiskN. meningitidis  is commonly associated with carriage (\u2013N. meningitidis serogroup B strain NZ98/254 (derived from the MeNZB vaccine) with porin A variable region 2 (PorA-VR2) variant 4 as the major antigen  cause most IMD cases worldwide, with B, C, and Y predominating in the United States , 4, and carriage \u20138. Quadrarriage \u2013. Two proarriage \u2013. MenB-FHarriage \u2013, 11, cor antigen , 13. In iologist . Althougiologist , 16. Whiiologist , 8, 17.N. meningitidis serogroup B outbreaks nor MenB mass vaccination campaigns occurred  or undetermined genogroup  isolates either lacked or contained a truncated fHbp allele   analyzed, 635 (47%) were either capsule null (egative) . A higheegative) , was detegative) . All FHbegative) .10.1128/mBio.00855-21.2TABLE\u00a0S1Table\u00a0S1, PDF file, 0.03 MB.Number of carriage isolates by genogroup. Download Copyright \u00a9 2021 Marjuki et al.2021Marjuki et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the fHbp allele, with 28 unique FHbp peptides (19 belonging to A/v2-3 and nine to B/v1) being detected, and had higher sequencing diversity than other genogroups. FHbp B/v1 peptides were detected among 57/225 (25.3%) genogroup B isolates; all remaining genogroup B isolates contained FHbp A/v2-3. FHbp A05/3.45 was detected in 21/225 (9.3%) genogroup B isolates, and FHbp B24/1.1 was detected in 17/225 (7.6%) genogroup B isolates  contained an intact isolates .nhbA allele was detected in 1,331 (99.6%) carriage isolates, with 90 unique NhbA peptides being identified  carriage isolates had intact entified . NadA-3.entified .10.1128/mBio.00855-21.4TABLE\u00a0S3N. meningitidis carriage isolates. Download Table\u00a0S3, PDF file, 0.09 MB.Distribution and diversity of NadA among Copyright \u00a9 2021 Marjuki et al.2021Marjuki et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the fHbp and nhbA intact. Ninety-two (6.9%) carriage isolates contained all three intact fHbp, nhbA, and nadA genes, of which 44 (47.8%) belonged to capsular genogroup B.Overall, there were 1,320 (98.7%) isolates with both porA alleles that belonged to 207 PorA types, and 9/1,316 (<1%)  contained PorA-VR2 variant 4, included in the MenB-4C vaccine  harbored intact  vaccine .N. meningitidis isolates  were recovered from participants with known vaccination status  of students at OR. A total of 1,422 n status , with 7210.1128/mBio.00855-21.5TABLE\u00a0S4Table\u00a0S4, PDF file, 0.03 MB.Number of isolates from participants with known MenB vaccine records. Download Copyright \u00a9 2021 Marjuki et al.2021Marjuki et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the fHbp alleles: 716/1,422 (50.4%) from MenB-FHbp-vaccinated participants, 42/1,422 (3.0%) from MenB-4C-vaccinated participants, and 648/1,422 (45.6%) from unvaccinated participants. The distribution of FHbp subfamilies in MenB-FHbp-vaccinated and unvaccinated groups was analyzed for all three universities as well as each university  had intact iversity . OverallfHbp allele or contained a truncated fHbp allele, including nine isolates from MenB-FHbp-vaccinated participants and four isolates from unvaccinated participants at RI-1 and two isolates from OR (one each from MenB-FHbp-vaccinated and unvaccinated participants). There was no statistically significant difference in FHbp peptide distributions between vaccinated and unvaccinated participants among genogroup B isolates and non-B genogroups from each university.The distribution of FHbp peptides was assessed among carriage isolates from MenB-FHbp-vaccinated and unvaccinated participants at RI-1 and OR . OverallWhile FHbp B01/1.55 was not detected in any isolate in this study, FHbp A05/3.45 was present in 12 of 726 (1.7%) isolates (11 genogroup B and one undetermined genogroup) from MenB-FHbp-vaccinated participants (nine from RI-1 and three from OR), all of which were from six MenB-FHbp-vaccinated participants (each received either one or two vaccine doses). Other FHbp peptides , shown to be covered by MenB-FHbp \u201324, were\u2013P\u2009=\u20090.3176).Of 194 repeat carriers, 161 (83.0%) received MenB-FHbp, 3 (1.5%) received MenB-4C, and 30 (15.5%) did not receive a vaccine. Among 161 MenB-FHbp-vaccinated repeat carriers, 143 (88.8%) had 302 isolates that retained the same strain genotype and 18 (11.2%) had 40 isolates that acquired a new strain genotype . Among t10.1128/mBio.00855-21.6TABLE\u00a0S5N. meningitidis isolates recovered from repeat carriers who carried new strains during the carriage evaluation period. Download Table\u00a0S5, PDF file, 0.1 MB.Sequence typing profiles of Copyright \u00a9 2021 Marjuki et al.2021Marjuki et al.https://creativecommons.org/licenses/by/4.0/Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the P\u2009<\u20090.0001) and genogroup B isolates (P\u2009=\u20090.0225). Additionally, of the 42 isolates that acquired a new strain genotype, the FHbp peptide prevalence between the detected strains in the previous round and newly acquired strains in the following round changed from B09/1.13  to B16/1.4  and from p0010 and p0024  to p0021  for NhbA  carried by MenB-FHbp-vaccinated repeat carriers, 32 unique FHbp peptides were detected, with B16/1.4  being the most prevalent, followed by B09/1.13 . Within the 302 isolates that retained the same strain genotype, six each (2.0%) harbored FHbp A05/3.45 or B24/1.1. Among the 60 isolates (10 genogroup B isolates) from unvaccinated participants, 13 unique FHbp peptides were detected, with B16/1.4  being most prevalent. There was a statistical difference in FHbp peptide distribution between vaccinated and unvaccinated repeat carriers among all isolates  of carriage isolates. We found an equal proportion of carriage isolates with an intact fHbp gene recovered from vaccinated (95% received MenB-FHbp) and unvaccinated participants. Statistical analysis showed no significant difference in the distribution of FHbp subfamilies or peptides between vaccinated and unvaccinated populations, suggesting that the vaccine does not impact the overall FHbp diversity and distribution.Analysis of whole-genome sequencing data has allowed us to understand MenB vaccine antigens and assess the impact of MenB-FHbp on their genetic diversity and distribution in U.S. meningococcal carriage. Almost all carriage isolates (99%) contained an intact isolates , but theBordetella pertussis following the introduction of pertussis vaccines, possibly due to vaccine selective pressure   , 38 thanOverall, similar FHbp peptide distributions were observed among carriage isolates from both vaccinated and unvaccinated participants during the study period. Additional investigations over a longer period are warranted to adequately evaluate the vaccine impact on carriage.N. meningitidis serogroup B outbreak and response activities occurred. A total of 1,514 meningococcal isolates  were recovered from 8,905 oropharyngeal swabs collected from 7,001 unique individuals . N. meningitidis species was determined using real-time PCR targeting sodC .Isolates included in this study were collected during February 2015 to March 2016 from meningococcal carriage evaluations at three U.S. universities  following vaccination using standardized methods as previously described , 8, 19. ing sodC , and sering sodC , 41. A tNeisseria meningitidis serogroup B carriage in institutional settings during an outbreak.\u201dThese evaluations were considered public health evaluations and did not require CDC institutional review for human subjects\u2019 protection. Both evaluations were covered under project determination numbers 2015\u20136436 and 2015\u20136442 for \u201cEvaluation of De novo short-read assembly was carried out using SPAdes 3.7.0 (cnl) . Illumina reads were trimmed with cutadapt  to removes 3.7.0  with thees 3.7.0 ; isolate.0 (cnl) . Genome .0 (cnl)  against database  to deterdatabase . All unidatabase , 48, 49.While 1,514 carriage isolates were obtained, only one isolate from each participant who carried the same meningococcal strain in more than one round (repeat carriers) was included in the analysis to assess the MenB vaccine antigen diversity and distribution among the carriage isolates circulating in the three universities, resulting in a total of 1,337 isolates.To compare MenB vaccine antigen distribution between vaccinated (received \u22651 dose of MenB-FHbp or MenB-4C 14\u2009days prior to sample collection) and unvaccinated participants, all isolates from repeat carriers were included even if they appeared to be the same strain genotype (CC:ST:PorA:FetA); however, participants with unknown MenB vaccination records were excluded from the analysis (92 isolates from RI-1 and OR), resulting in a total of 1,422 carriage isolates. For phylogenetic analysis of MenB vaccine antigens, multiple-sequence alignment of peptides was generated by ClustalW  with CLCN. meningitidis serogroup B or non-N. meningitidis serogroup B. Isolates negative for both subfamilies A and B were excluded from this test due to being a low proportion (0.9%) of the overall isolate collection. Chi-squared goodness-of-fit tests were used to test for changes in distribution among N. meningitidis serogroup B and non-N. meningitidis serogroup B isolates. A Bonferroni correction was applied to correct for multiple comparisons. To assess proportions between vaccinated and unvaccinated repeat carriers who acquired a new meningococcal strain, a Fisher\u2019s exact test was performed; FHbp peptide distributions in all isolates and genogroup B isolates among these groups were analyzed using a likelihood ratio chi-squared test.Statistical analysis was performed using SAS version 9.4 . Fisher\u2019s exact test was used to assess changes in the distribution of FHbp subfamilies and peptides in isolates recovered from vaccinated and unvaccinated participants, stratified by university and whether the isolate was PRJNA533315.Sequence reads are available under NCBI BioProject no."}
+{"text": "FallsTalk is a one-month evidence-based falls prevention (FP) program that focuses attention on causes of an individual\u2019s falls and encourages new FP behaviors. We translated the program for family caregivers of persons with cognitive impairment or dementia (PwCID) to administer to the PwCID and examined the number of new FP behaviors (#newFPBs) reported during the intervention period by the Caregiver. Thirty-four Dyads (Caregiver+PwCID) were trained to conduct brief daily FP discussions together using paper (FTCGnoTech) or computerized  guidance. Dyads had FallsTalk training, daily discussions and weekly check-in calls. FTCGTech included discussion suggestions tailored to Caregiver concerns. To examine the use of technology, Poisson regression models compared #newFPBs between FTCGnoTech and FTCGTech and included covariates age (<80 or >=80), Mini-Mental Status Exam (MMSE >25 or <=25) and interactions. To evaluate the influence of Caregiver participation, #newFPBs reported by 115 non-demented clinical trial participants (no Caregiver- FT-ClTrNoCG) were compared with the Caregiver outcomes. Dyads using technology reported significantly more #newFPBs (MeanFTCGnoTech=5.34(SEM=0.68), MeanFTCGTech=8.46(SEM=0.76); p=.004) during the intervention month. A significant interaction was observed whereby Dyads with MMSE<=25 using technology, reported significantly more #newFPBs than the non-technology group (MeanFTCGnoTech=4.23(SEM=0.72), MeanFTCGTech=9.01(SEM=0.90); p=.047). Caregiver (n=34) involvement substantially increased #newFPBs (MeanFT-ClTrNoCG=1.39(SEM=0.15), MeanCaregiver=7.21(SEM=0.49); p<.0001), independent of technology. Across studies, participants or Caregivers for those with MMSE<=25 and <80yo reported significantly more #newFMBs (Mean=4.38(SEM=0.55) than those 80+yo (Mean=2.06(SEM=0.30); p=.0026). FallsTalk Caregiver provides an effective means to promote new Dyad FP strategies. The influence of Caregiver involvement and technology show promise in encouraging behavioral change to prevent falls."}
+{"text": "Escherichia coli and Klebsiella pneumoniae, as compared to 3GC-susceptible isolates of either species.Carbapenems are considered the drugs of choice for first-line treatment of severe infections caused by carbapenem-susceptible, extended-spectrum \u03b2-lactamases (ESBL)-producing Enterobacterales, while piperacillin-tazobactam has been recommended as an alternative for treatment of non-severe infections. Temocillin is stable to ESBL and AmpC enzymes and may thus represent another treatment option. This study assessed the in vitro activity of piperacillin-tazobactam and temocillin against third-generation cephalosporin (3GC)-resistant One hundred and nine isolates from hospitalized patients with bloodstream and urinary tract infections were tested. All isolates were collected during the resistance surveillance study of the Paul-Ehrlich-Society for Chemotherapy in 2016/17. Minimum inhibitory concentrations (MICs) were determined by broth microdilution according to the standard ISO 20776-1 and interpreted using EUCAST clinical breakpoints (version 11.0).E. coli, n=58; K. pneumoniae, n=21) were 3GC-resistant and 30  were 3GC-susceptible. Susceptibility to piperacillin-tazobactam was detected in 93.3% of 3GC-susceptible isolates (for both E. coli and K. pneumoniae) and in 79.3% and 57.1% of the 3GC-resistant E. coli and K. pneumoniae, respectively. In contrast, 3GC-susceptible isolates were 100% susceptible to temocillin as were 94.8% and 90.5% of the 3GC-resistant E. coli and K. pneumoniae, respectively.Seventy-nine isolates (E. coli and K. pneumoniae from bloodstream and urinary tract infection samples, with susceptibility rates exceeding those of piperacillin-tazobactam.In conclusion, temocillin demonstrated potent in vitro activity against carbapenem-susceptible, 3GC-resistant  Escherichia coli and Klebsiella pneumoniae is most commonly mediated by extended-spectrum \u03b2-lactamases (ESBLs), but also occurs through other mechanisms like plasmid-encoded AmpC-type enzymes . Mo enzymes , [2], ) and two were ceftazidime-resistant only (cefotaxime-susceptible [S+I]).Meropenem inhibited all isolates at the lowest concentration tested . Seventy-nine isolates were 3GC-resistant  and the number and percentage of isolates classified as susceptible (S or I) or resistant were calculated.MIC distribution data of temocillin and piperacillin-tazobactam are presented in Table 1 E. coli and K. pneumoniae. MIC50/90 values of piperacillin-tazobactam and temocillin were \u22641/4 mg/L and 4/16 mg/L, respectively, for E. coli, and \u22641/8 mg/L and 2/8 mg/L, respectively, for K. pneumoniae. One isolate each of E. coli and K. pneumoniae had a piperacillin-tazobactam MIC of \u2265128 mg/L. MICs of ceftazidime for both isolates were 1 mg/L as compared to MICs of \u22640.25\u20130.5 mg/L for the other 28 3GC-susceptible isolates.Susceptibility to piperacillin-tazobactam (MIC\u22648 mg/L) and temocillin (MIC\u226416 mg/L) among the 30 3GC-susceptible isolates was 93.3% and 100%, respectively, for both E. coli and K. pneumoniae to the other antibiotics tested are displayed in E. coli were observed for ampicillin (53.3%), cotrimoxazole (26.7%), and amoxicillin-clavulanic acid (20%), while the highest level of drug resistance in 3GC-susceptible K. pneumoniae, except for ampicillin, was observed for fosfomycin (33.3%). Two 3GC-susceptible K. pneumoniae were colistin-resistant.Susceptibility data of the 3GC-susceptible E. coli was 79.3% (95% CI: 68.9\u201389.7%) versus 94.8% (95% CI: 89.1\u2013100%), respectively, and among 3GC-resistant K. pneumoniae 57.1% (95% CI: 44.4\u201369.8%) versus 90.5% (95% CI: 83.0\u201398.0%), respectively. MIC50/90 values of piperacillin-tazobactam and temocillin were 2/32 and 8/16 mg/L mg/L, respectively, for E. coli, and 8/\u2265128 mg/L and 8/16 mg/L, respectively, for K. pneumoniae. Seventeen 3GC-resistant isolates  were resistant to piperacillin-tazobactam, but susceptible to temocillin, while one E. coli isolate was temocillin-resistant, but susceptible to piperacillin-tazobactam. Two isolates of either species were resistant to both antimicrobials  were detected for levofloxacin, tobramycin and fosfomycin in 3GC-resistant K. pneumoniae. One 3GC-resistant isolate each of E. coli and K. pneumoniae was colistin-resistant, and two 3GC-resistant E. coli were fosfomycin-resistant.Susceptibility data of the 3GC-resistant, carbapenem-susceptible E. coli and K. pneumoniae has become widespread in all parts of the world. According to data of the European Antimicrobial Resistance Network (EARS-Net), investigating invasive isolates, resistance to 3GC in E. coli rose from 1.7% in 2002 to 15.1% in 2019. 3GC resistance also increased in K. pneumoniae. EARS-Net reported an overall resistance rate of 31.3% for 2019, with extremely large differences in the level of resistance between individual countries (to >60%)  and one temocillin-resistant [MICs 32\u201364 mg/L]) without resistance to other \u03b2-lactams, found mutations in several genes, including a porin-encoding gene (ompD-like) and genes encoding for transport/membrane proteins . T. TE. coleumoniae . Temocillsewhere , 15], [, [E. col Table 2 , but the Table 2 . Seventy Table 2 . Thus, oic shock . It musthttps://doi.org/10.5061/dryad.931zcrjkc [Data for this article are available from the Dryad Repository: 31zcrjkc The investigation of the susceptibility of the test organisms to temocillin was supported by a grant from Eumedica S/A, Manage, Belgium, to Antiinfectives Intelligence GmbH.MK is a partner and CEO of Antiinfectives Intelligence GmbH, a research organization providing services to pharmaceutical companies. YP and GW declare that they have no competing interests."}
+{"text": "Streptomyces (68%), Micromonospora (6%), and Nocardiopsis (3%) are dominant producers of secondary metabolites. Additionally, alkaloids (37%), polyketides (33%), and peptides (15%) comprise the largest proportion of natural products with mostly antimicrobial activity and cytotoxicity. Furthermore, the data analysis and clinical information of SMs have been summarized in this article, suggesting that some of these actinomycetes with multiple host organisms deserve more attention to their special ecological status and genetic factors.The actinomycetes have proven to be a rich source of bioactive secondary metabolites and play a critical role in the development of pharmaceutical researches. With interactions of host organisms and having special ecological status, the actinomycetes associated with marine animals, marine plants, macroalgae, cyanobacteria, and lichens have more potential to produce active metabolites acting as chemical defenses to protect the host from predators as well as microbial infection. This review focuses on 536 secondary metabolites (SMs) from actinomycetes associated with these marine organisms covering the literature to mid-2021, which will highlight the taxonomic diversity of actinomycetes and the structural classes, biological activities of SMs. Among all the actinomycetes listed, members of  Actinomycetes are Gram-positive bacteria with a GC-rich linear genome and have proven to be a rich source of secondary metabolites (SMs) of broad structural diversity and biological properties . The oceThe objective of this article is to provide an overview of the natural products from actinomycetes associated with marine animals, marine plants, macroalgae, cyanobacteria, and lichens. The present review was not only summarizing the structural classes and biological activities of SMs but also highlighted the taxonomic diversity of actinomycetes, as well as the data analysis of integrated above information. Some of these metabolites with excellent activity are expected to become new drugs such as antibiotics, antineoplastic drugs, or anticancer drugs. Therefore, actinomycetes with multiple host organisms deserve more attention to their special ecological status and genetic factors.Streptomyces, Micromonospora, Microbacterium, and Nocardiopsis were abundant , respectively indole, another known compound indole-3-acetic acid (S16) was discovered from this strain indole (tnam Sea . In addis strain .73) and isoquinocycline B (74)  showed good antibiotic activity not only against the Gram-positive strain S. aureus SH1000, but also the Gram-negative strains efflux knockout (KO) mutant strains of K. pneumoniae ATCC10031 and A. baumannii ATCC 17978. It was also active against the human liver cancer cell line HepG2 with an IC50 of 13.3 \u00b5M ..Callyspo79) (S38), and aerugine (80) (S39) and furan-2-carboxamide (S40) from Streptomyces sp. OUCMDZ-1703 associated with a soft coral sample collected from the South China Sea. Thiazole derivatives 79 and 80 displayed moderate antibacterial activity against S. aureus and three methicillin-resistant strains MRSA082, MRSA111, and MRSA234 anthraquinone derivatives, urdamycinone E (in (134)  were ide E (135) , the pos50 \u03bcg/mL ,95.136\u2013141) (Streptomyces sp. CMS JV M18_3 [136\u2013139) has great antibacterial activities against MRSA and vancomycin-resistant Enterococcus faecium (VREF). In addition, compounds 136\u2013139 were active against HCT-116 human colon carcinoma [141) displayed anti-biofilm activity inhibiting Staphylococcus aureus biofilm formation, and compound 140 exhibited antimicrobial activities against some Gram-positive bacteria [Six dihydroquinone derivatives 136\u2013141)  were iso6\u2013141  wearcinoma . SF2415Bbacteria ,98.142) and B (143) (Nocardiopsis sp. KMF-002 was cultivated from an unidentified dark purple marine sponge. Nocatrione A (142) showed a significant protective effect against UVB irradiation in both NHDF cell lines, whereas nocatrione B (143) was active against UVB only in a specific NHDF cell line [In 2014, Min Cheol Kim et al. reported isolation and identification of two novel tetracenedione derivatives nocatriones A ( B (143)  from Nocell line .Actinokineospora sp. EG49 cultivated from the marine sponge Spheciospongia vagabunda afforded two new actinosporin analogs actinosporins C (144) and D (145) (146) and B (147) ( D (145) . At 1.25 B (147)  were als148)  [Microluside A (148)  is a uniectively . In 2015in (S72) .149) (S73), and a related analog (\u2212)-BE-52440A (150) -BE-52440A (150) showed cytotoxicity against NB4 and HL-60 cells with IC50 values of 1.7 and 1.8 \u03bcM, respectively. Naquihexcin A (149) bears a rare unsaturated hexuronic acid moiety and could inhibit the proliferation of an adriamycin-resistant human breast cancer cell line MCF-7 ADM with IC50 = 16.1 \u03bcM, indicating that the unsaturated hexuronic acid moiety could enhance the activity against the cancer cells [Three S-bridged pyranonaphthoquinone dimers naquihexcins A (149)  and B (S0A (150)  were proer cells .S74, identified as 3-hydroxy-2-methyl-4H-pyran-4-one , was isolated both from the Streptomyces sp. SBT348 and Rhodococcus sp. UA13 [S75 was discovered from the co-culture of sponge-derived Saccharomonospora sp. UR22 and Dietzia sp. UR66 [151\u2013153) (Nocardiopsis sp. HB-J378 associated with Theonella sp. Nocardiopsistin B showed the best anti-MRSA activity with the same MIC (3.12 \u03bcg/mL) as that of chloramphenicol, whereas nocardiopsistins A and C have a moderate anti-MRSA activity (MIC = 12.5 \u03bcg/mL) [151\u2013153), it was found that the ketone functional group at C-4 could enhance the anti-MRSA activity, while the hydroxyl group at C-3 weakened activity.Compound sp. UA13 ,45. Compsp. UR66 . In 2018151\u2013153)  separate5 \u03bcg/mL) . On compS76) was first isolated from Streptomyces rochei MB037 associated with marine sponge Dysidea arenaria collected at Yongxin Island in the South China Sea. And the co-culture with fungus Rhinocladiella similis 35 derived from gorgonian could enhance its production [154), together with three known actinosporins C, D, and G   exhibited potent antitrypanosomal activity and growth inhibitory activity towards Trypanosoma brucei strain TC221 [7-methoxy-2,3-dimethylchromone-4-one   were obtin TC221 .156 and 157)   were pro[158\u2013160  were iso and 162  showed iectively .146, 144) were isolated from the axenic cultures of strain EG49. Actinosporin A showed anti-trypanosomal activity and actinosporin C exhibited antioxidant activity. New actinosporins E\u2013H  (165)  (S78) [Actinosporins A and C   were pro4) (165)  and the  B (166)  (plant g167) (S79) with fully saturated eight-membered lactone ring were reported in 1991 from Streptomyces sp. PG-19 collected on the surface of Cortez gorgonian octocoral Pacifigorgia sp. Octalactin A demonstrated significant in vitro cytotoxicity toward B-16-F10 murine melanoma (IC50 = 7.2 \u00d7 10\u22123 \u00b5g/mL) and HCT-116 human colon tumor (IC50 = 0.5 \u00b5g/mL) [Isolation of two novel compounds Octalactins A (167)  and B (S5 \u00b5g/mL) .S80) was produced by a marine actinomycete Micromonospora sp. strain A5-1 obtained from soft coral Scleronephthya sp. in the East China Sea [Streptomyces sp. OUCMDZ-1703\u2020 associated with a soft coral collected from the South China Sea led to the discovery of two new chlorinated polyketides strepchloritides A and B (168 and 169) (S41). Polyketides 168 and 169 showed moderate cytotoxicity against MCF-7 tumor cells [A novel analog of jadomycin B, 7b,13-dihydro-7-O-methyl jadomycin B (hina Sea . Streptoand 169) , togetheor cells .S81) and two known anthracycline derivatives  were produced by coral-associated Streptomyces sp. SCSIO 41399. Compound 170 (Aranciamycin K (ound 170  exhibite171) and anthracimycin (172)  with MIC values less than 0.03 \u00b5g/mL. And anthracimycin B was also active against these four Gram-positive bacteria. In addition, anthracimycin displayed anti-tubercular activity against Mycobacterium tuberculosis [A novel gram-positive antibiotic anthracimycin B (in (172)  were isorculosis .173)  and HePG2 (hepatic carcinoma) in vitro [1-hydroxy-1-norresistomycin (HNM) (173)  was prodin vitro .174\u2013176)   were rep25 \u03bcg/mL .177)   was isolhrocytes ,110.178)   was founlomerase ,111.Streptomyces sp. #N1-78-1 from sea squirt Ecteinascidia turbinata in Puerto Rico led to the purification of bisanthraquinones 1 and 2 , as well as derivative 3 (181) (Staphylococcus aureus) and VRE , and these three compounds displayed cytotoxic activity against HCT-116 cells [The isolation of  3 (181) , the deh16 cells ,112.182, 183, S83, and S84) were produced by a strain of Micromonospora sp. derived from a Brazilian endemic ascidian Eudistoma vannamei. Compounds 182 and 183 . Halomadurones C (184) and D (185) (186)   showed p5) (186)  was also strains .187\u2013190) and nahuoic acid A (191)  with an unprecedented carbon skeleton was the first natural product inhibiting the SETD8 lysine methyltransferase and the first selective SETD8 inhibitor. Compounds 187\u2013191 showed weak antibiofilm activity against Shewanella onedensis MR-1 biofilm [S87, S88, 192\u2013194). Compounds E-G (192\u2013194)   were sep biofilm . In 2017192\u2013194)  had weaksubtilis .Streptomyces coelicoflavus strain HQA809, which is isolated from sea squirt Styela clava, produced germicidin (195) and 6-isopropyl group-3-ethyl-4-hydroxy-2-pyrone (196)  . Both twa salina .Streptomyces sp. PTY087I2 associated with styela canopus collected from Bastimentos Park, Bocas del Toro, Panama produced three naphthoquinone derivatives granaticin (197), granatomycin D (198), and dihydrogranaticin B (199)  . Co-cultathogens ,118.200\u2013204)  suppressed the proliferation of cancer cell lines, and metabolites 203 is the most active compound with IC50 values ranging from 0.59 to 3.39 mM. The 11-hydroxycurvularins 200 and 201 also showed antibacterial activity inhibiting the growth of Escherichia coli [205 . And the known metabolite 301 (Five curvularin macrolides (200\u2013204)  were isohia coli . Compounoli [205 , which wlite 301  with ant206), ochromycinone (207), and X-14881 C (208), together with a new angucyclines saccharothrixmicine A (209) (Saccharothrix espanaensis An 113 obtained from a marine mollusk specimen (Anadara broughtoni), which was collected from Peter the Great Bay, Sea of Japan, Russia. Compounds 206\u2013208 showed significant activity against C. albicans, B. subtilis, E. faecium, S. aureus, and Xanthomonas sp. pv. badrii. And Compound 209 exhibited activity towards Candida albicans and Xanthomonas sp. pv. Badrii [S89) and F (S90), together with gilvocarcins M (S91) and V (S92), have been isolated from an unidentified tunicate-associated Saccharopolyspora sp. SS081219 JE-28 [Three known analogs X- 14881 E ( A (209) , were pr. Badrii ,121. Mac19 JE-28 ,77.210) and I (211) were reported together with known violapyrones B (212) and C (213)  showed growth inhibitory activity against cancer cell lines at concentrations less than 26.12 \u03bcg/mL. Wherein compound 210 showed the highest cytotoxic activity against the HCT-15 cell line with a GI50 value of 1.10 \u03bcg/mL. Additionally, violapyrones B and C were demonstrated to have antibacterial activities. Therefore, it may be noteworthy that each compound has structural similarities, but showed different activities. The results suggested that the length of the aliphatic side chain and the position of the methyl group affected the activity. Furthermore, violapyrones having an isomethyl group in the alkyl side chain showed better activity than others [Isolation of two novel 3,4,6-trisubstituted \u03b1-pyrone derivatives violapyrones H ( C (213) , from Stn others .214) and PM100118 (215)   from StrTCC10231 .216, 217) (S93\u2013S97) were produced by the sea anemone-derived Streptomyces sp. ZZ406. New compounds 216 and 217 showed potent activity in inhibiting the proliferation of different glioma cells and downregulating the expressions of glioma metabolic regulators. Compound 216 was active against the proliferation of different glioma cells with IC50 values of 4.7 to 8.1 \u03bcM, high selectivity index (>12.3 to 21.3), and good stability in human liver microsomes [S98) and the known angucyclinone PD116740 (126) were isolated from the sea urchin-derived Streptomyces sp. HDa1 [Two new compounds   and fivecrosomes . A novelsp. HDa1 .11 (218) and Q12 (219)  were separated from the marine gastropod mollusk Batillaria zonalis-associated Streptomyces sampsonii SCSIO 054. In addition, four known anthraquinones chrysophanol (S101), 4-acetylchrysophanol (S102), islandicin (S103), and huanglongmycin A (S104) were also discovered. Julichrome Q12 (219) was found to display antibacterial activity against Micrococcus luteus and Bacillus subtilis with MICs of 2.0 and 8.0 \u03bcg/mL. What\u2019s more, compounds 218, 220\u2013222  , along w 220\u2013222  also sho64 \u03bcg/mL .223\u2013225) (50 0.63 \u03bcg/mL), and all three novel metabolites exhibited significant cytotoxicity against the 39 human cancer cell lines [Further investigation for metabolites of this strain has led to the isolation of three additional novel cytotoxic metabolites designated as halichoblelide A\u2013C (223\u2013225) . Halicholl lines ,126.226)  [Ochoa et al. reported the isolation of a new glycosylated polyketide phocoenamicin (226)  in 2017, 2.6 \u03bcM) .Most peptides from actinomycetes are circular and contain further rare structural elements, such as chromophore or unusual amino acids.Streptomyces sp. DA18 was separated from marine sponge Craniella australiensis collected at Sanya Island in the South China Sea, from which diketopiperazines (DKPs)    were disactivity .230)  and Trypanosoma brucei brucei (IC50 = 0.0032 \u03bcM). Additionally, it was active against 293T kidney epithelial cells (IC50 11.24 \u00b5M) and J774.1 macrophages (IC50 < 0.10 \u00b5M) [The cyclic depsipeptide valinomycin (230)  was isol0.10 \u00b5M) .231) (Nocardiopsis strain HB383 [232) . The compound displayed weak cytotoxic activity against human cervical carcinoma HeLa cells with an IC50 value of 49 \u03bcM [A diketopiperazine (231)  with weain HB383 . The new83 [232)  was puriof 49 \u03bcM .233) and JBIR-35 (234)   from Strd 2.5 mM .S106) and JBIR-57 (S107) were reported in 2011 from the new isolate Streptomyces sp. SpD081030SC-03, which was obtained from an unidentified sponge collected in Ishigaki, Okinawa, Japan [Isolation of two new peptides JBIR-56   were discovered in 2011 from Verrucosispora sp. strain WMMA107, which was separated from Chondrilla caribensis f. caribensis . 22\u2032-Deoxythiocoraline (236), thiochondrilline C (237), and 12\u2032 -sulfoxythiocoraline (238)  , which wne (238)  showed sectively .Streptomyces sp. strain RV15 was reported to produce four new cyclic lipopeptides cyclodysidins A\u2013D (S110\u2013S113) in 2012 [239 and 240) (S114), were isolated from Streptomyces M1087 associated with an unidentified sponge. The new compounds 239\u2013240 exhibited weak inhibition against the recombinant enzyme sortase B with EC50 values of 88.3 and 126.4 \u00b5g/mL [A sponge-derived  in 2012 ,133. Twoand 240) , along w.4 \u00b5g/mL .Streptomyces sp. NIO 10068 derived from a sponge collected from the western coast of India produced linear dipeptides proline\u2013glycine (S115) and N-amido-\u03b1- proline (S116) [241)  with MIC values of 0.25\u20130.5 \u03bcg/mL [e (S116) . Kocurin6) [241) , a new m.5 \u03bcg/mL ,137.242) (S117) in 2015, from sponge-associated Streptomyces sp. LS298. Quinomycin G (242) with a terminal double bond in one of the Ser groups exhibited moderate antibacterial activities against Staphylococcuse pidermidis, S. aureus, Enterococcus faecium, and E. faecalis with MIC values ranging from 16 to 64 \u03bcg/mL. In addition, it showed potent anti-tumor activities and the highest activity was observed against the Jurkat cell line (human T-cell leukemia) with an IC50 value of 0.414 \u03bcM [Xin Zhen et al. reported isolation and identification of two new metabolites quinomycin G (242)  and cycl0.414 \u03bcM .243) (Streptomyces sp. SBT348 exhibited significant cytotoxicity towards the human promyelocytic HL-60 and the human colon adenocarcinoma HT-29 cell lines [244) (S118) were isolated from Streptomyces sp. GKU 220 is associated with a marine sponge sample collected in Andaman sea, Ranong, Thailand. Rakicidin F (244) showed growth inhibitory activity against B. subtilis and E. coli at the dosage of 25 \u03bcg per disk [A new cyclic dipeptide petrocidin A (243)  was discll lines . New cyces [244)  and knowper disk .245)   was repo \u03bcg/disk . Four disp. G246 . And thr246 [246  and S120sp. G248 .247\u2013250) and actinomycin D (251) (Streptomyces sp. LHW52447 associated with Phyllospongia foliascens obtained from the Xisha Islands in the South China Sea. Actinomycins D1 (247) and D2 (248) introduced an oxazole unit into the central phenoxazinone chromophore and exhibited more potent activities against three strains of MRSA with MIC values of 0.125\u20130.25 \u03bcg/mL than that of actinomycins D3\u2013D4 (MIC = 0.5\u20131.0 \u03bcg/mL), which indicated that the incorporation of the oxazole unit would enhance the antibacterial activity. In addition, the cytotoxicity evaluation against human lung WI38 embryonal fibroblasts suggested that the incorporation of oxazole unit could decrease the cytotoxicity of actinomycins on human normal cells [Four new D-type actinomycin analogs actinomycins D1-D4 ( D (251)  were disal cells . The SARal cells ,142.Streptomyces sp. Call-36 isolated from sponge Callyspongia sp. collected in the Red Sea was reported to produce a new diketopiperazine actinozine A (252), cyclo(2-OH-D-Pro-L-Leu) (253), cyclo(D-Pro-L-Phe) (254) (L-Pro-L-Phe) (S123). Compounds 252 and 253 displayed potent activity against S. aureus and were moderately active against C. albicans. Compound 254 exhibited moderate and selective activity against HCT-116 with an IC50 of 32.7 \u00b5M, while cyclo (L-Pro-L-Phe) (S123) was inactive, indicating that the D/L configuration of Pro had an important effect on the activity [e) (254) , and cycactivity .255) (Nesterenkonia sp. MSA31 and active against multidrug-resistant Pseudomonas aeruginosa by inhibiting the phenotypic expression of virulence factors [256\u2013258)   was isol factors . Three a256\u2013258)  were obt235) is a thiodepsipeptide antitumor antibiotic isolated from Micromonospora sp. L-13-ACM2-092 is associated with a soft coral collected in the Indian Ocean off the coast of Mozambique. Thiocoraline had an inhibitory effect on DNA polymerase \u03b1. In addition, it displayed potent cytotoxicity and strong activity against Gram-positive bacteria [Thiocoraline   (229), bacillusamide B (261) (L-Pro-L-Leu) (246), and cyclo (L-Pro-L-Ile) (262)   from a sand MSSA ,147. Fou B (261) , cyclo (e) (262) , were send MCF-7 ,72.263\u2013265)  (263\u2013265)  was repoell line .266, 267) (S127\u2013S129) were discovered from Streptomyces sp. CNB-091 obtained from the surface of jellyfish Cassiopeia xamachana collected in the Florida Keys. Salinamides A and B exhibited moderate antibiotic activity against Gram-positive bacteria. Additionally, the results of phorbol ester-induced mouse ear edema assay showed that salinamides A and B displayed significant topical anti-inflammatory activity using [268)   and sality using ,150. In ng [268) , a new b269\u2013271  was separated from Micromonospora sp. ML1 isolated from a mollusk collected from the Indian Ocean coast of Mozambique [272) and H (273)  inhibitory activity with IC50 values of 7.50 and 7.37 \u03bcmol/L, respectively [Three known diketopiperazines 269\u2013271  producedolyticus . Thiocorzambique . Limazep H (273)  were disectively .S130 and the known valinomycin (230) in 2018, from sea anemone-associated Streptomyces sp. ZZ406. Valinomycin (230) was active against the proliferation of different glioma cells and downregulating the expressions of glioma metabolic regulators [Streptomyces sp. strains 22 and 34 that have been mentioned above. Compound 274    was sepaof 31 \u03bcM ,154.276, 277, S131\u2013S133) were isolated from Streptomyces sp. (ZJG1) cultivated from stony corals collected in the South China Sea. Compound 277 (276 (Five sesquiterpenes (ound 277  showed g277 (276  exhibiteS134) and B (278)   was repos aureus .Streptomyces sp. RKBH-B7 is associated with octocoral Eunicea. Guanahanolide A (279)   showed m280) (S135), two novel eunicellin diterpenoids, were reported in 2020 from the culture of Streptomyces albogriseolus SY67903 associated with the gorgonian Muricella sibogae collected at the South China Sea. Microeunicellol A exhibited cytotoxicities against several human cancer cell lines [Microeunicellols A (280)  and B (Sll lines .281\u2013283)   were disof 30 \u03bcM .284)  , a novel2 \u03bcmol/L .285) (S136) were isolated from sponge-associated Streptomyces sp. RM66. Manadoperoxide H exhibited antitrypanosomal activity against Trypanosoma brucei rhodesiense with an IC50 value of 0.375 \u03bcg/mL [Manadoperoxide H (285)  and acan75 \u03bcg/mL .S137) was purified from Micrococcus luteus R-1588-10 associated with sponge Xestospongia sp. collected at Noumea, New Caledonia. Additionally, the previously known synthetic 2,4,4\u2032-trichloro-2\u2032-hydroxydiphenylether (286)   was alsoalbicans ,160.287) (Streptomyces sp. (LA3L2). Two known compounds montagnetol (S138) and erythrin (S139) were also isolated from Streptomyces sp. (LA3L2) that is the first reported actinomycete to produce these lichen-related compounds. In addition, chromomycin A2 (S140), chromomycin A3 (S141), and chromomycin 02-3D (S142) were reported to be separated from Streptomyces sp. (LA3L1) [Isolation of a new cytotoxic metabolite (287)  characte (LA3L1) .290), B (291), and C (292), together with two known analogs X-14881 A (288) and X-14881 B (289)   , were pr B (293)  was alsoy modest ,121.Streptomyces sp. strain T03 derived from the sponge Tethya sp. led to the identification of butenolide (294) (trypanosoma activity against Trypanosoma brucei brucei (IC50 = 31.77 \u03bcM) [295) (Actinomadura sp associated with a sea squirt Ecteinascidia turbinate. It showed potent activity against Clostridium difficile NAP1/B1/027 [de 294) , which e4 , which1.77 \u03bcM) . EcteinaM) [295)  was obta1/B1/027 ,115.S143) and two known compounds S144 and S145 were isolated from Streptomyces microflavus strain No. HVG29 is associated with the marine sponge Hymeniacidon perlevis collected from the coast of Dalian (China). Compounds S143\u2013S145 were the first time to isolate deoxyuridine structures from S. microflavus associated with sponges [New nucleoside derivative ( sponges .Streptomyces sp. NIO 10068 derived from a marine sponge produced cinnamic acid (296)  , which wnt study .297, 298) (S146) from Solwaraspora sp. WMMB329 is associated with ascidian Trididemnum orbiculatum. The two novel compounds demonstrated antibacterial activity against MRSA and MSSA [In 2014, Ellis et al. reported isolation and identification of two novel trialkyl-substituted aromatic acids, solwaric acids A and B  , togetheand MSSA ,162.299, S147\u2013S149) and diphosphatidylglycerol (S150) were produced by Microbacterium sp. HP2 associated with sponge Halichondria panacea collected at the Adriatic coast, Rovinj, Croatia. The major compound 299  was reported in 2017 from Streptomyces griseorubens sp. ASMR4 is associated with an unidentified soft coral collected in the Red Sea at the Hurghada coast, Egypt. Additionally, along with metabolite S144, seven other known metabolites S152\u2013S158 were also discovered from the strain ASMR4 [Isolation of a new oxaphenalene derivative (in ASMR4 .S159) and tryptophan (S160) were produced by Rhodococcus sp. UA13 [S161) and 1,4-dihydroxy-2-naphthoic acid (S162) were isolated from the axenic culture of Saccharomonospora sp. UR22 [S160) and L-phenylalanine (S163) were produced by the sponge-associated Streptomyces sp. G246 [300  was produced by the sea anemone-derived Streptomyces sp. ZZ406 [Phenylacetic acid methyl ester (sp. UA13 . Two knosp. UR22 . In addisp. G246 . The kno246 [300  isolatedactivity . GTRI-02p. ZZ406 .S165) and thymidine-3-thioamine (S166) from sponge-associated Streptomyces sp. Call-36 [302)  transcriptional activation effect [In 2019, Shaala et al. reported the discovery of two new nucleosides thymidine-3-mercaptocarbamic acid   was prodE,8Z)/-5-oxo-6,8-tetradecadienoic acids   and the fish pathogen Tenacibaculum maritimum (preferred 304). In addition, compounds 303 and 304 displayed agonistic activity against peroxisome proliferator-activated receptors (PPARs) with an isoform specificity towards PPAR\u03b1 and PPAR\u03b3 [A pair of geometrically isomeric unsaturated keto fatty acids   were idend PPAR\u03b3 .305) (S167), actinopolysporin B (S168), and acanthosterol G (S169) was reported in 2021 from sponge-associated Streptomyces sp. RM66. Peroxidessethyl plakortide Z was active against solid tumor and L-1210 leukemia cell lines in vitro [Isolation of four metabolites ethyl plakortide Z (305) , seco-plin vitro .306, 109) were produced by an unidentified actinomycete strain CNC-837 obtained from the surface of the Caribbean brown alga Lobophora variegate. The new compounds, distantly related to antibiotics of the kijanimicin class, are potent inhibitors of topical PMA-induced edema in the mouse ear assay when administered either topically or IP [Two new anti-inflammatory compounds lobophorins A  and B  from the central Cantabrian Sea. Streptomyces cyaneofuscatus M-27 produced several antitumor antibiotics of the anthracycline family, of which two antibiotics were identified as daunomycin (307) and cosmomycin B (308) (309) (310)  . And it 8) (309) . In addi9) (310)  was sepaoperties .311)   was repoes T\u00fc 57 .312) (S170) were separated from the seaweed-associated Streptomyces sp. HZP-2216E obtained from sea lettuce Ulva pertusa, a traditional Chinese medicine. They are both existing as zwitterion and streptopertusacin A showed moderate activity against the growth of MRSA [A unique indolizinium alkaloid streptopertusacin A (312)  and a no312) S were sep of MRSA .S171\u2013S173), together with the known benzamides 2-acetamido-3-hydroxybenzamide (S174), 2-amino-3-hydroxybenzamide (S175), and 2-aminobenzamide (S176), were isolated from the Streptomyces sp. ZZ502 is associated with the seaweed Ulva conglobatea collected in the East China Sea [Three new compounds (hina Sea .S177 was reported in 2013 from Streptomyces cavourensis YY01-17 separated from the lichens grown in the Antarctic area [Isolation of a novel metabolite tic area .313 (S178\u2013S180) from Streptomyces sundarbansensis strain WR1L1S8 associated with brown algae Fucus sp. collected in Bejaia coastline, Algeria. Compound 313 stood out as the most active of the series, showing a selective antibacterial activity against MRSA with a MIC of 6 \u03bc\u039c [Polyketide 313  [2-hydro of 6 \u03bc\u039c .314) and B (315)  produced by Streptomyces cyaneofuscatus M-27 associated with Fucus spiralis was first isolated from Streptomyces or marine environments [Germicidins A ( B (315) , discoveronments .316)  , a 28-me0\u22125\u00b5g/mL .317), together with bafilomycin D(320), 9-hydroxybafilomycin D (321), and bafilomycin A1(322)  and 21,22-en-9-hydroxybafilomycin D (319) (S182) were separated. Compounds 318 and 319 had potent activity against the proliferation of glioma U251 and C6 cells with IC50 values of 0.12\u20131.08 \u03bcM. In addition, they were active against MRSA with MIC values of 12.5 mg/mL. The four bafilomycins of compounds 320\u2013322 and 317 showed potent activity in suppressing the proliferation of the four tested glioma cell lines with IC50 values of 0.35 to 2.95 \u00b5M. In addition, compounds 320\u2013322 were reported to have antibacterial, antifungal, insecticidal, and herbicidal activities. Bafilomycins D and A1  also exhibited potent activity in inhibiting vacuolar-type ATPase [A new compound of 23-O-butyrylbafilomycin D( A1(322) , was iso D (319) , togethee ATPase ,171.323)  and colon carcinoma (DLD-1), but not normal mammary fibroblasts [Desertomycin G (323)  was separoblasts .S183, S184) and Germicidins K, L , together with six previously reported metabolites wailupemycin D (S187), wailupemycin E (S188), enterocin/vulgamycin (324), 5-deoxy-enterocin (325), germicidin A (314) and germicidin B (315)  , was rep griseus .326\u2013328)   was repoor cells .329)   from Strectively .330) and imDKP (331) (Streptomyces praecox strain 291-11 separated from the rhizosphere of Undaria pinnatifida. The two compounds inhibited zoospores with a therapeutic ratio (LC50/EC50) of 17.7 and 21 and inhibited diatoms with a therapeutic ratio of 17.7 and 21, respectively [Two antifouling diketopiperazines bmDKP (KP (331)  were isoectively .S189) and cholest-4-en-3-one (S190) were discovered from Streptomyces sp. FX-58 is associated with marine plant Salicornia herbacea [332\u2013335)   produced.2 \u03bcg/mL .336) and 2-hepta-1,5-dienyl-3,6-dihydroxy-5-(3-methylbut-2-enyl) benzaldehyde (337)  benzaldehyde (de (337)  were repus iniae .S191) in 2013 together with a known compound (S192) from lichen-derived Streptomyces cavourensis YY01-17 [338) , hydrocinnamic acid (S193), and (E)-cinnamic acid(S194) were separated from Streptomyces ambofaciens BI0048 associated with red alga Laurencia glandulifera [Shan-Shan Su et al. reported the discovery of a novel compound ( YY01-17 . And a n17 [338)  was prodbacteria . Three kdulifera .S195), the compound responsible for the \u201cearth smell\u201d and beta-patchoulene (S196) used as a fragrance agent in the perfume industry, were two major metabolites of Streptomyces carnosus M-40 [339) and Linoleic acid (340)  , which wes T\u00fc 57 .Streptomyces (68%), Micromonospora (6%), and Nocardiopsis (3%) are the dominant producers , polyketides (33%), and peptides (15%) comprise the largest proportion of secondary metabolites, while roducers . Figure roducers .Approximately 64% of the SMs displayed various biological activities, especially antimicrobial activity and cytotoxicity . Interes9) is the earliest rifamycin antibiotic used in clinical application. Tetrodotoxin (105) has been widely used as an analgesic, sedative, antispasmodic, and local anesthetic in clinics. And daunomycin (307) has a good effect on acute myeloid leukemia. In addition, for these drugs already in clinical use, more clinical trials are underway for new diseases or new usages. .Molecules with excellent activities, which have been in clinical applications or have entered clinical trials, were listed in Metabolites from actinomycetes associated with marine organisms have proven to be an abundant source for the isolation of multiple potent bioactive metabolites with diverse structures. In this review, we attempt to discuss the significance of the special ecological status and genetic factors of these actinomycetes with multiple hosts. The chemical ecology underlying hosts\u2013actinomycetes interactions provide a great opportunity for the discovery of novel drugs. During the co-evolution, these actinomycetes and their specific hosts constructed a coordinated and relatively independent micro-ecological environment, in which SMs can be tolerated by the host and are the active inhibiting specific external invasion. Therefore, actinomycetes associated with various marine hosts play an important ecological role in producing novel medicinal active compounds. Currently, there are relatively few studies on these actinomycetes, but many secondary metabolites have been isolated with excellent bioactivities. Some of these metabolites have been used in clinical applications or have entered clinical trials where they are expected to become new drugs. There is no doubt that further exploration can be a useful strategy for discovering novel marine natural products. These actinomycetes, however, are difficult to be cultured under experimental conditions. Therefore, in-depth exploration of the ecology of these actinomycetes to continuously optimize culture conditions is crucial for further research. Meanwhile, the use of advanced bioinformatics technology for gene detection of uncultured actinomycetes and heterologous expression of the collected biosynthetic gene clusters will be another important pathway for research on SMs of marine organism-associated actinomycetes."}
+{"text": "The cave biodiversity of continental Portugal faces tremendous conservation challenges, mostly linked to their direct destruction and contamination infiltrating from the surface. Beetles are the most diverse insects and one of the most diverse arthropod groups in caves of Portugal.Carabidae) are the most diverse family of cave-adapted beetles in continental Portugal, followed by rove beetles (Staphylinidae). Beetles in caves of Portugal are mostly terrestrial and only one species is known to have evolved to live in groundwater. Trechus is the most diverse genus with four species, followed by Domene with three species and by Speonemadus and Iberoporus, both with one cave-adapted species. The aim of this contribution is to assess all endemic cave-adapted species of beetles from continental Portugal and to support their specific protection, to promote adequate management of surface habitats and the establishment of priority areas for conservation. The main biodiversity erosion drivers that are impacting the conservation of the studied species are pollution infiltrating from the surface, urbaniation, modifications of the natural habitat for touristic purposes and mining, quarrying and energy production infrastructures.We present the IUCN Red List profiles for the cave-adapted beetles from continental Portugal, all endemic to their respective geological units and massifs. Ground beetles (This document can be used in spatial planning and territory management in karst, based on the current scientific knowledge. Cave fauna has relatively low specific richness, but high conservation value for the global biodiversity of our planet . SubterrInsecta, Coleoptera) stand out, as a group that presents the greatest animal specific richness worldwide (Coleoptera (Carabidae (mostly Trechinae) and Leiodidae (mostly Cholevinae)  .Portugal is located in the western part of the Iberian Peninsula, has more than 2000 caves identified and is considered a hotspot of subterranean biodiversity . More thTrechusfulvus Dejean, 1831, captured by M.L.W. Schaufuss; the caves then explored remain unknown  consists of a matrix of unaggregated rock that can be found in scree slopes. Most of this sampling was performed under the framework of Master and Doctoral studies  and all 2) and the maps were created in the open source software QGIS 3.14.16, with the natural protected areas of Portugal layer (Extent of occurence (EOO) and area of occupancy (AOO) were calculated using the Geospatial Conservation Assessment Tool (GeoCAT) with an approximation to the standard IUCN 2 km \u00d7 2 km cells  or Photo(s): Fig. 1Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 2Basis of EOO and AOO: Known habitat extentBasis (narrative): The extent of occurrence (EOO) and area of occupancy (AOO) are both 4 km\u00b2.Min Elevation/Depth (m): 218Iberoporuspluto is a groundwater-adapted beetle known from a stream in a single cave, located in north-eastern Sic\u00f3 karst area. The cave stream, where it was found, flows in a subterranean system of approximately 15 km of horizontal implementation (Range description: entation .EOO (km2): 4Trend: Decline (observed)Justification for trend: A decline in EOO is inferred due to the degradation of the cave by anthropogenic impact as Soprador do Carvalho Cave is subject to recreational visitation and direct trampling on the subterranean stream.Causes ceased?: NoCauses understood?: YesCauses reversible?: YesExtreme fluctuations?: UnknownTrend: Decline (projected)Justification for trend: No decline in AOO has been observed, but it is inferred due to the decline and vulnerability of the habitat.Causes ceased?: NoCauses understood?: YesCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Iberoporuspluto occurs in a single cave, Soprador do Carvalho, in the Sic\u00f3 karst area in central Portugal and it is under moderate disturbance : Only one specimen  of this species is known from a single location in central Portugal.Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: FreshwaterHabitat specialist: YesHabitat (narrative): The specimen was collected in the bottom of a clay pool connected to a subterranean stream .Trend in extent, area or quality?: Decline (observed)Justification for trend: Soprador do Carvalho Cave is located in the vicinity of a village, agricultural fields and a quarry . It is aHabitat importance: Major ImportanceHabitats: 5.1. Wetlands (inland) - Permanent Rivers/Streams/Creeks 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 5.1. Wetlands (inland) - Permanent Rivers/Streams/Creeks 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 2.8 mmGeneration length (yr): 1Dependency of single sp?: UnknownIberoporus Castro & Delgado, 2001 is endemic to the Iberian Peninsula and exclusively stygobiont (Iberoporuspluto has extreme troglomorphic traits (eyeless and depigmentation) and unusually elongated legs not adapted for swimming (Ecology and traits (narrative): The genus ygobiont . Iberoposwimming .Justification for threats: The cave is explored for tourism and visitors step over the habitat where the species was found . Other tThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas3.2. Energy production & mining - Mining & quarrying6.1. Human intrusions & disturbance - Recreational activities9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas3.2. Energy production & mining - Mining & quarrying6.1. Human intrusions & disturbance - Recreational activities9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Measures should be taken to prevent infiltration of wastewaters from the village into the cave stream. The nearby quarry has been reported in the national media to be the source of the infiltration of small particles of quarry dust that have been deposited all over the gallery of Algarinho Cave by flood events. This type of slurry is known to perniciously impact groundwater quality  and AlgaConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management2.3. Land/water management - Habitat & natural process restoration4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management2.3. Land/water management - Habitat & natural process restoration4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Further investigation is needed about the distribution, ecology and life cycle of the species. Developing a management plan for this species is crucial. This plan will aid the conservation of the cave-adapted species of the Sic\u00f3 karst area.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Further investigation is needed about the distribution, ecology and life cycle of the species. Developing a management plan for this species is crucial. This plan will aid the conservation of the cave-adapted species of the Sic\u00f3 karst area.TrechusmachadoiScientific name: Species authority: Jeannel, 1941AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: CarabidaeFamily: T.fulvus-group of species.Taxonomic notes: This species belongs to the Figure(s) or Photo(s): Fig. 2Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 3Basis of EOO and AOO: Known habitat extent2.Basis (narrative): The extent of occurrence (EOO) and the maximum estimated area of occupancy (AOO) are both 4 kmMin Elevation/Depth (m): 389Max Elevation/Depth (m): 400Trechusmachadoi is a troglobiont beetle known only in the Alcobertas Cave and in a countinuous mesovoid shallow substratum (MSS) located in the Serra dos Candeeiros subunit of the Estremenho karst massif, central Portugal. The cave extends horizontally for approximately 210 m (Range description: ly 210 m  and the ly 210 m .EOO (km2): 4Trend: Decline (observed)Justification for trend: Despite intensive sampling in the type locality (cave), no specimens have been found in there since the species description. Only recent sampling in the MSS contiguous to the cave retrieved specimens.Causes ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: Decline (observed)Justification for trend: AOO decline has been inferred due to the vulnerability of the habitat.Causes ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 2Trechusmachadoi is known only from the Alcobertas Cave and from the adjacent MSS, located approximately 80 m from each other. Its distribution is likely to be confined to the subterranean habitats of the Serra dos Candeeiros subunit : So far, only one population is known from the Alcobertas Cave, which is also dispersed in the contiguous mesovoid shallow substratum at 0.5 m depth in scree slopes .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: NoSystem: TerrestrialHabitat specialist: YesTrechusmachadoi population to other parts of the subterranean network  in scree slopes close to the type locality (Habitat (narrative): The Alcobertas Cave was subject of a large anthropogenic intervention at the beginning of the 1970s, with the intention to transform it to receive mass tourism. During that process, a second entry was opened near the end of the gallery which induced significant changes in the climatology of the cave . Many ex network . The Alclocality . These slocality .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - Caves7.2. Caves and Subterranean Habitats (non-aquatic) - Other Subterranean HabitatsHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - Caves7.2. Caves and Subterranean Habitats (non-aquatic) - Other Subterranean HabitatsSize: 4.85 mm (lectotype)Generation length (yr): 1Dependency of single sp?: UnknownTrechusmachadoi is a troglobiont with reduced eyes and body depigmentation. It lives exclusively in subterranean habitats and is only known from a single cave and from scree slopes habitats in the Serra dos Candeeiros subunit of the Estremenho karst massif in central Portugal (Ecology and traits (narrative): Portugal .Justification for threats: Since the 1970s, this cave has been intensively explored for touristic activities. During that period, a second entrance has been opened, drastically changing the environment . The scrThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas3.2. Energy production & mining - Mining & quarrying3.3. Energy production & mining - Renewable energy6.1. Human intrusions & disturbance - Recreational activitiesThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas3.2. Energy production & mining - Mining & quarrying3.3. Energy production & mining - Renewable energy6.1. Human intrusions & disturbance - Recreational activitiesJustification for conservation actions: The habitats are protected under the EU \u201cRede Natura 2000\u201d , but theConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Further investigation is needed about the population size, extent of distribution, ecology and life cycle. It is urgent to develop a management plan for this species, which will consequently improve the conservation of further cave-adapted species of Serra dos Candeeiros subunit of the Estremenho massif.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Further investigation is needed about the population size, extent of distribution, ecology and life cycle. It is urgent to develop a management plan for this species, which will consequently improve the conservation of further cave-adapted species of Serra dos Candeeiros subunit of the Estremenho massif.TrechusgamaeScientific name: Species authority: Ribeira & Reboleira, 2009AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: CarabidaeFamily: T.fulvus-group\u201d species complex.Taxonomic notes: This species belongs to the \u201cFigure(s) or Photo(s): Fig. 3Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 4Basis of EOO and AOO: Known habitat extentBasis (narrative): The extent of occurrence (EOO) is 73.4 km\u00b2 and the maximum estimated area of occupancy (AOO) is 24 km\u00b2.Min Elevation/Depth (m): 250Max Elevation/Depth (m): 485Trechusgamae is a troglobiont beetle, known from five caves and from the mesovoid shallow substrate (scree slopes), all located in the Santo Ant\u00f3nio Plateau, the central subunit of the Estremenho karst massif (Range description: t massif .EOO (km2): 73.4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 24Number of locations: 6Trechusgamae was found in five caves: Algar de Marradinhas II, Algar das Gralhas VII, Algar do Pena, Algar da Arroteia and Algar do Ladoeiro, all located in the Santo Ant\u00f3nio Plateau, the central subunit of the Estremenho karst massif. Recently, a single specimen was found in the mesovoid shallow substrate at 0.5 m depth in scree slopes of F\u00f3rnea, which is also located in the Santo Ant\u00f3nio Plateau, showing that this species may also disperse through more superficial subterranean habitats : Amongst the six known localities with populations of eia Cave . Algar deia Cave . All theNumber of subpopulations: 6Trend: Decline (inferred)Justification for trend: All the subpopulations face threats derived from intensive quarrying activity, which changes land use and disturbs the natural processes of the habitat. The subpopulations from Algar do Ladoeiro, Algar das Marradinhas II, Algar da Arroteia and F\u00f3rnea face threats of pollution and land use disturbance due to the proximity of urbanised areas.Extreme fluctuations?: UnknownSevere fragmentation?: NoSystem: TerrestrialHabitat specialist: YesTrechusgamae was found in the deepest parts of the caves, from 50 to 95 m depth, all with high humidity levels (> 98%) and temperatures ranging from 13.5\u00baC (in Algar do Pena) to 17\u00baC (in Algar de Marradinhas II) (Habitat (narrative): nhas II) . The cavnhas II) . More renhas II) .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - Caves7.2. Caves and Subterranean Habitats (non-aquatic) - Other Subterranean HabitatsHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - Caves7.2. Caves and Subterranean Habitats (non-aquatic) - Other Subterranean HabitatsSize: 4.83\u20135.38 mm , 3.94\u20135.44 mm Generation length (yr): 1Dependency of single sp?: UnknownTrechusgamae was the only cave-adapted beetle collected in the caves and MSS of the Santo Ant\u00f3nio Plateau and it shows a strict subterranean lifestyle (Ecology and traits (narrative): ifestyle . Both adifestyle .Justification for threats: The distribution area of the species is all covered by the Natural Park of Serras d'Aire e Candeeiros. However, intense quarry activity is currently ongoing in the surrounding areas of the known localities. Algar do Pena is located 300 m from a quarry and Algar das Gralhas VII 168 m from the same quarry. Algar do Ladoeiro's entrance is 840 m from the closest urban areas, Algar das Marradinhas II is located 1.5 km from the nearest village and Algar da Arroteia is located 112 m from the closest house and 1.3 km from a quarry. Algar do Pena Cave hosts a laboratory and is open for visits upon previous booking. F\u00f3rnea is located 595 m from the closest house and 1.9 km from a quarry. The distribution area faces severe groundwater contamination , due to Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas3.2. Energy production & mining - Mining & quarrying6.1. Human intrusions & disturbance - Recreational activities9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas3.2. Energy production & mining - Mining & quarrying6.1. Human intrusions & disturbance - Recreational activities9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Although the habitat is protected by law under the \u201cRede Natura 2000\u201d , the speConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Additional investigation about the population size, extent of distribution, ecology and life cycle is required. The development of a management plan that will improve the conservation of this cave-adapted species in the Santo Ant\u00f3nio Plateau subunit of the Estremenho karst massif is fundamental to ensure its conservation and protection.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Additional investigation about the population size, extent of distribution, ecology and life cycle is required. The development of a management plan that will improve the conservation of this cave-adapted species in the Santo Ant\u00f3nio Plateau subunit of the Estremenho karst massif is fundamental to ensure its conservation and protection.TrechuslunaiScientific name: Species authority: Ribeira & Reboleira, 2009AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: CarabidaeFamily: T.fulvus-group\u201d species complex.Taxonomic notes: This species belongs to the \u201cFigure(s) or Photo(s): Fig. 4Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 5Basis of EOO and AOO: Known habitat extent2 and the maximum estimated area of occupancy (AOO) is 12 km\u00b2.Basis (narrative): The extent of occurrence (EOO) is 4 kmMin Elevation/Depth (m): 95Max Elevation/Depth (m): 307Trechuslunai is a troglobiont Carabidae known from three horizontal caves, located in Serra de Aire/S\u00e3o Mamede Plateau (Range description:  Plateau .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 12Number of locations: 3Trechuslunai was found in three caves of the Serra de Aire/S\u00e3o Mamede Plateau subunits. The southernmost distribution is the Almonda Cave, but this species is also known from the Contenda and Moinhos Velhos Caves' system : Three populations are known exclusively from the Serra d'Aire/S\u00e3o Mamede Plateau subunit of the Estremenho karst massif .Number of subpopulations: 3Trend: Decline (inferred)Justification for trend: The subpopulation in Almonda cave is subject to wastewater and pollution infiltration and the subpopulations of the Contenda and Moinhos Velhos Caves face heavy contamination derived from the village under which they are located.Extreme fluctuations?: UnknownSevere fragmentation?: NoSystem: TerrestrialHabitat specialist: YesTrechuslunai only occurs in the deepest parts of the caves, from 50 to 80 m depth. The three caves have high humidity levels and average temperature of 18\u00baC (Habitat (narrative):  of 18\u00baC . In Moin of 18\u00baC . The oth of 18\u00baC .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3.55\u20134.73 mmGeneration length (yr): 1Dependency of single sp?: UnknownTrechuslunai was the only troglobiont species captured in these caves. All known localities are caves that flood seasonally (Ecology and traits (narrative): asonally .Justification for threats: Almonda Cave is located 50 m from a factory that conducts the subterranean river into the building for industrial use of the water and at 420 m from the village centre. The surrounding village is also heavily populated by agricultural fields. Contenda and Moinhos Velhos caves are heavily contaminated, as their subterranean network extends below the village of Mira d'Aire. The entrance of Moinhos Velhos Cave is located in the village centre and it has been explored for tourism since the 1960s with several complementary touristic infrastructures built .Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.3. Agriculture & aquaculture - Livestock farming & ranching9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.3. Agriculture & aquaculture - Livestock farming & ranching9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: The Contenda and Moinhos-Velhos caves develop below the village of Mira d'Aire and infiltration of sewage is observed in the underground. Therefore an effort to improve sewage treatment is necessary in order to prevent wastewater run-off into subterranean gealleries and groundwaters. Almonda Cave is classified as Property of Public Interest (IIP) since 1993 and protected due to archaeological heritage . The arcConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Information about the population size, extent of distribution, ecology and life cycle of this species is scarce, therefore, further investigation is required. It is also necessary to develop a management plan that will improve the conservation of the species in the Serra de Aire/S. Mamede Plateau subunits of the Estremenho karst massif.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Information about the population size, extent of distribution, ecology and life cycle of this species is scarce, therefore, further investigation is required. It is also necessary to develop a management plan that will improve the conservation of the species in the Serra de Aire/S. Mamede Plateau subunits of the Estremenho karst massif.TrechustataiScientific name: Species authority: Reboleira & Ortu\u00f1o, 2010AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: CarabidaeFamily: T.fulvus-group\u201d species complex. It is recognisable by the shape of the aedeagus and has a slim body, rudimentary wings, reduced eyes  and depigmentation. This species is the most troglomorphic ground-beetle known from Portugal  or Photo(s): Fig. 5Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 6Basis of EOO and AOO: Known habitat extentBasis (narrative): The extent of occurrence (EOO) and the maximum estimated area of occupancy (AOO) are both of 4 km\u00b2.Min Elevation/Depth (m): 380Trechustatai is a cave-adapted hygrophilous Carabidae known only from one small cave, located in Serra do Montejunto (Range description: ntejunto .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Trechustatai only occurs in one cave, Algar do Javali, located in Serra do Montejunto. This species is geographically isolated : Only one population is known from Algar do Javali Cave, in Montejunto karst massif.Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: NoSystem: TerrestrialHabitat specialist: YesTrechustatai was only collected in the deep oligotrophic areas of the cave. It was never found in areas with high organic material content (bat guano accumulation zones). The cave is 10 m deep and extends for 80 m. Temperatures in the deepest zone of the cave ranged from 14.2 \u00baC in winter to 15 \u00baC in summer (Habitat (narrative): n summer .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 5.2\u20135.9 mm , 4.8\u20136 mm Generation length (yr): 1Dependency of single sp?: UnknownTrechustatai is the most troglomorphic carabid beetle from continental Portugal. Other caves of the area were sampled, but T.tatai was never collected elsewhere. The seasonal activity pattern of the beetle was studied during one year in the deepest zone of the cave and specimens were collected during winter, autumn and spring : d spring . The abshumidity .Eucalyptus intensive plantation, with direct impact on land use at the surface, pollution and groundwater depletion.Justification for threats: Algar do Javali Cave is located 1.6 km from a quarry with intensive extraction activity and 2.9 km from the closest village, which induces deep changes in land use at the surface and potential biotic exchange, such as introduction of invasive alien species. The cave entrance is located 50 m from a road and surrounded by Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.2. Agriculture & aquaculture - Wood & pulp plantations3.2. Energy production & mining - Mining & quarrying4. Transportation & service corridorsThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.2. Agriculture & aquaculture - Wood & pulp plantations3.2. Energy production & mining - Mining & quarrying4. Transportation & service corridorsJustification for conservation actions: Although this cave is protected by law through the \u201cRede Natura 2000\u201d , the speConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Some crucial steps necessary for the protection of the species are the development of a management plan for the conservation of this cave-adapted species in Serra do Montejunto and the promotion of further studies regarding population size, extent of distribution, ecology and life cycle.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Some crucial steps necessary for the protection of the species are the development of a management plan for the conservation of this cave-adapted species in Serra do Montejunto and the promotion of further studies regarding population size, extent of distribution, ecology and life cycle.SpeonemadusalgarvensisScientific name: Species authority: Reboleira, Fresneda & Salgado, 2017AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: LeiodidaeFamily: Speonemadusescalerai-group\" and is recognisable by the equal/subequal length of the 2nd, 4th, 5th and 7th antennomeres and a slightly transverse and hexagonal pronotum : Suppl. materials 7Basis of EOO and AOO: Known habitat extent2 and the maximum estimated area of occupancy (AOO) is 12 km2. The three caves are located along a 48 km straight line, with Algar\u00e3o do Remexido Cave being 23 km from Vale Telheiro and Vale Telheiro Cave being 25 km from Senhora Cave.Basis (narrative): The extent of occurrence (EOO) is 7.4 kmMin Elevation/Depth (m): 72Max Elevation/Depth (m): 269Speonemadusalgarvensis was collected in three caves in the southermost province of Portugal in the Algarve, being most likely endemic to the central and eastern parts of the Algarve karst massif, a region also known as Barrocal Algarvio (Range description: Algarvio .EOO (km2): 7.4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 12Number of locations: 3Speonemadusalgarvensis is known from three caves  in the Algarve karst massif : There are three populations known from Portugal, all from caves in the Algarve karst massif. The largest number of individuals was collected in Vale Telheiro Cave, followed by Senhora Cave and Algar\u00e3o do Remexido Cave .Number of subpopulations: 3Trend: Decline (inferred)Justification for trend: All populations are under risk due to wastewater infiltration derived from the urbanised areas in the region. The subpopulation of Algar\u00e3o do Remexido cave is threatened by agricultural pollution infiltration and the subpopulation from Senhora cave is threatened by industrial residue pollution.Extreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): Specimens were collected in three caves, 15 to 30 m deep, with high humidity levels and average temperatures of 17.8\u00baC , 18.8\u00baC (Senhora Cave) and 19.3\u00baC (Algar\u00e3o do Remexido Cave). This species is endemic to the Algarve karst massif .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 4\u20134.9 mmGeneration length (yr): 1Dependency of single sp?: UnknownSpeonemadusalgarvensis occurs exclusively in the Algarve and does not exhibit the typical troglomorphism found in other cave-adapted species, such as evident eye reduction, severe depigmentation and extreme body and appendages elongation, although it has only been collected in caves and never at the surface. This species was found to carry the ectoparasitic fungus of the order Laboulbeniales  attached to the cuticle and foresic acari (Ecology and traits (narrative): ic acari .Justification for threats: Algar\u00e3o do Remexido is located under agricultural lands, 370 m from the closest house and 1.7 km from the closest village. Vale Telheiro is located 290 m from the closest house and 745 m from the closest urbanisation. Senhora Cave is located 168 m from the closest house and 900 m from an industrial complex.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.3. Agriculture & aquaculture - Livestock farming & ranching9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.3. Agriculture & aquaculture - Livestock farming & ranching9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Although the habitat is protected under legislation by the \u201cRede Natura 2000\u201d , the speConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: The development of a management plan for the conservation of this cave-adapted species in the Algarve karst massif and the encouragement of more studies regarding population size, extent of distribution, ecology and life cycle are essential measures for the protection of the species.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: The development of a management plan for the conservation of this cave-adapted species in the Algarve karst massif and the encouragement of more studies regarding population size, extent of distribution, ecology and life cycle are essential measures for the protection of the species.DomenelusitanicaScientific name: Species authority: Reboleira & Orom\u00ed, 2011AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: StaphylinidaeFamily: Taxonomic notes: Individuals display troglomorphism, such as microphthalmia, lack of wings and body elongation .Figure(s) or Photo(s): Fig. 6Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 8Basis of EOO and AOO: Known habitat extentBasis (narrative): The extent of occurrence (EOO) and the maximum estimated area of occupancy (AOO) are both of 4 km\u00b2.Domenelusitanica was found in a single cave located in the Sic\u00f3 karstic massif (Range description: c massif .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Domenelusitanica was collected in Cer\u00e2mica Cave, located in the Sic\u00f3 karst area in central Portugal . This Ccave extends for 355 m : This species is known from a single population in central Portugal .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): Specimens were exclusively collected in the deepest zones of the cave (10 m deep), in high humidity levels and with average temperatures of 16.4\u00baC .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 9\u20139.48 mmGeneration length (yr): 1Dependency of single sp?: UnknownDomenelusitanica is included in the subgenus Lathromene, together with the other two Portuguese species of cave-adapted Domene: D.viriatoi and D.darinkae. This species is a predator troglobiont rove beetle, with reduced eyes, apterous, depigmented and elongated body and appendages : pendages . It is oel, 1946 .Eucalyptus plantations.Justification for threats: Cer\u00e2mica Cave is located 550 m from an animal farm, 3.5 km from the nearest village and 3.6 km from a quarry. It is surrounded by agricultural lands and Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.3. Agriculture & aquaculture - Livestock farming & ranching3.2. Energy production & mining - Mining & quarryingThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.3. Agriculture & aquaculture - Livestock farming & ranching3.2. Energy production & mining - Mining & quarryingJustification for conservation actions: The habitat is located in an \u201cRede Natura 2000\u201d area . PopulatConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: In order to build a sustainable conservation plan for the species in the Sic\u00f3 karst area, more information about population size, extent of distribution, ecology and life cycle is needed. The threats also need to be addressed and minimised, if possible, in order to improve the habitat quality.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: In order to build a sustainable conservation plan for the species in the Sic\u00f3 karst area, more information about population size, extent of distribution, ecology and life cycle is needed. The threats also need to be addressed and minimised, if possible, in order to improve the habitat quality.DomeneviriatoiScientific name: Species authority: Serrano & Boieiro, 2015AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: StaphylinidaeFamily: Taxonomic notes: This species displays body, leg and antennae elongation, microphthalmia and lack of wings .Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 9Basis of EOO and AOO: Known habitat extent2.Basis (narrative): The extent of occurrence (EOO) and the maximum estimated area of occupancy (AOO) are both 4 kmDomeneviriatoi was collected in two galleries of the Buraco da Moura Cave, located in the Serra da Estrela Mountain foothills (Range description: oothills .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Domeneviriatoi was collected from the Buraco da Moura Cave at the edge of the Estrela Mountain chain : This species is known from a single population in the western border of the Estrela Mountain chain, the highest mountain of continental Portugal .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): The cave is formed by granite blocks in the margins of the Cani\u00e7a stream at an elevation of 677 m. It extends for 150 m of underground passages .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 6.9\u20128.2 mm , 6.3\u20128.8 mm Generation length (yr): 1Dependency of single sp?: UnknownDomeneviriatoi is included in the subgenus Lathromene. Both adults and larvae of this species were observed foraging for preys in bat guano on the cave substrate : ubstrate . The cavJustification for threats: The cave entrance is located 127 m from the closest house, 530 m from a hydroelectric power station and 1.2 km from the closest village and is under anthropogenic disturbance due to tourism.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas6.1. Human intrusions & disturbance - Recreational activities7.2. Natural system modifications - Dams & water management/use9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas6.1. Human intrusions & disturbance - Recreational activities7.2. Natural system modifications - Dams & water management/use9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Buraco da Moura Cave was classified as a \u201cNational Important Underground Shelter for Bats\u201d therefore a decrease in human disturbance is expected . HoweverConservation action type: In PlaceConservation actions: 1.1. Land/water protection - Site/area protection5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 2.1. Land/water management - Site/area management4. Education & awarenessConservation action type: In PlaceConservation actions: 1.1. Land/water protection - Site/area protection5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 2.1. Land/water management - Site/area management4. Education & awarenessUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.1. Conservation Planning - Species Action/Recovery Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: A sustainable conservation plan for the species is only possible if more information about population size, extent of distribution, ecology and life cycle is collected.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.1. Conservation Planning - Species Action/Recovery Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: A sustainable conservation plan for the species is only possible if more information about population size, extent of distribution, ecology and life cycle is collected.DomenedarinkaeScientific name: Species authority: Magrini & Carotti 2019AnimaliaKingdom: ArthropodaPhylum: InsectaClass: ColeopteraOrder: StaphylinidaeFamily: Region for assessment: EuropeBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 10Basis of EOO and AOO: Known habitat extent2.Basis (narrative): The extent of occurrence (EOO) and the maximum estimated area of occupancy (AOO) are both 4 kmDomenedarinkae is a cave-adapted rove beetle known from an abandoned mine in northern Portugal : 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Domenedarinkae is known from a single horizontal artificial cave, Santa Isabel mine, located in the Mar\u00e3o Mountain chain in north Portugal : Only one specimen of this species is known from a single location in northern Portugal .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): The only known specimen was collected in the rocky debris along the main tunnel of the Santa Isabel mine . The geoTrend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 6.93 mm Generation length (yr): 1Dependency of single sp?: UnknownDomenedarinkae is included in the subgenus Lathromene. It is a predator and exhibits troglomorphisms, such as depigmentation, elongation of body and antennae and accentuated microphthalmia (Ecology and traits (narrative): hthalmia .Threat type: PastThreats: 6.3. Human intrusions & disturbance - Work & other activities7.3. Natural system modifications - Other ecosystem modificationsThreat type: PastThreats: 6.3. Human intrusions & disturbance - Work & other activities7.3. Natural system modifications - Other ecosystem modificationsJustification for conservation actions: Undisturbed areas in the surface of the mine need to be defined and established.Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awarenessConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awarenessUse type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Further information about population size, extent of distribution, ecology and life cycle is needed to better protect the species and the habitat. The mine, where the species was found, is an anthropogenic construction that clearly adversely affected the natural habitat of the species that should be the deep fissures and the mesovoid shallow substrate of the area. Therefore, it is recommended to sample these habitats in the area to understand the distribution of this species and to define new conservation priorities.Use type: InternationalEcosystem service type: Very importantResearch needed: 1.2. Research - Population size, distribution & trends1.3. Research - Life history & ecology2.2. Conservation Planning - Area-based Management Plan3.1. Monitoring - Population trends3.4. Monitoring - Habitat trendsJustification for research needed: Further information about population size, extent of distribution, ecology and life cycle is needed to better protect the species and the habitat. The mine, where the species was found, is an anthropogenic construction that clearly adversely affected the natural habitat of the species that should be the deep fissures and the mesovoid shallow substrate of the area. Therefore, it is recommended to sample these habitats in the area to understand the distribution of this species and to define new conservation priorities.http://iyck2021.org), an event organised by the International Union of Speleology to promote the awareness for the importance of caves and their habitats. Under this framework, a global initiative created the International Cave Animal of the Year  devoted to cave beetles. Within this initiative, different countries selected their own endemic species as a flag for advocating the conservation of subterranean ecosystems. Here, we offer information about the distribution  beetle from Portugal, was described, based on a female specimen and no further specimens have been found in a cave that has been constantly monitored for more than a decade. Its habitat, the Soprador do Carvalho Cave is under serious anthropogenic threats, such as groundwater contamination and touristic pressure ; 2: T.gamae (yellow circle); 3:T.lunai (pink circle); 4: T.tatai (red circle); 5: Iberoporuspluto (blue star); 6: Domenelusitanica (yellow diamond); 7: D.viriatoi (pink diamond); 8: D.darinkae (blue diamond); and 9: Speonemadusalgarvensis (pink triangle). (A) Detail of northern distribution, (B) Detail of central distribution and (C) Detail of southern distribution. In green are protected areas.1: File: oo_528602.tifhttps://binary.pensoft.net/file/528602A.S.P.S. Reboleira, R.P. Eus\u00e9bio0F7E68E8-AAD0-50EE-B880-0CEFA740227D10.3897/BDJ.9.e67426.suppl2Supplementary material 2Iberoporuspluto.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionIberoporuspluto distribution: Soprador do Carvalho Cave, Penela, Coimbra District.File: oo_560421.tifhttps://binary.pensoft.net/file/560421A.S.P.S. Reboleira, R.P. Eus\u00e9bioD7DABAF2-3B19-5E93-BD25-C30939739EB610.3897/BDJ.9.e67426.suppl3Supplementary material 3Trechusmachadoi.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionTrechusmachadoi distribution: Alcobertas Cave and mesovoid shallow substratum, Rio Maior.File: oo_560428.tifhttps://binary.pensoft.net/file/560428A.S.P.S. Reboleira, R.P.Eus\u00e9bioF33828A8-DED8-539A-8ADB-E01A34FF86B110.3897/BDJ.9.e67426.suppl4Supplementary material 4Trechusgamae.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionTrechusgamae distribution: (1) Algar da Arroteia Cave; (2) F\u00f3rnea (MSS); (3) Algar do Ladoeiro Cave; (4) Algar de Marradinhas II Cave; (5) Algar do Pena Cave; and (6) Algar das Gralhas VII Cave. All caves and MSS are located in the Santo Ant\u00f3nio Plateau, the central subunit of the Estremenho karst massif. (A) Detail of distribution.File: oo_560430.tifhttps://binary.pensoft.net/file/560430A.S.P.S. Reboleira, R.P. Eus\u00e9bio50F0176B-32A1-5094-B243-043C8F72D14210.3897/BDJ.9.e67426.suppl5Supplementary material 5Trechuslunai.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionTrechuslunai distribution: (1) Contenda and Moinhos Velhos Cave system; and (2) Almonda Cave, both located in the Estremenho karst massif.File: oo_560431.tifhttps://binary.pensoft.net/file/560431A.S.P.S. Reboleira, R.P. Eus\u00e9bio8999FE89-5F39-5920-80EA-13A029878CF110.3897/BDJ.9.e67426.suppl6Supplementary material 6Trechustatai.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionTrechustatai distribution: Algar do Javali Cave, Montejunto karst massif.File: oo_560436.tifhttps://binary.pensoft.net/file/560436A.S.P.S. Reboleira, R.P. Eus\u00e9bio3044C8C3-6105-508F-AB09-06C452FC863710.3897/BDJ.9.e67426.suppl7Supplementary material 7Speonemadusalgarvensis.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionSpeonemadusalgarvensis distribution: (1) Algar\u00e3o do Remexido Cave; (2) Vale Telheiro Cave; and (3) Senhora cavea, all located in the Algarve karst massif.File: oo_560437.tifhttps://binary.pensoft.net/file/560437A.S.P.S. Reboleira, R.P. Eus\u00e9bio2C303E65-CCAC-500D-B815-0FAE2AEA3BFE10.3897/BDJ.9.e67426.suppl8Supplementary material 8Domenelusitanica.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionDomenelusitanica distribution: Cer\u00e2mica Cave, Sic\u00f3 karst area.File: oo_560438.tifhttps://binary.pensoft.net/file/560438A.S.P.S. Reboleira, R.P. Eus\u00e9bio081BE9A5-9F66-5DA2-8DF0-7B8416C9E0E110.3897/BDJ.9.e67426.suppl9Supplementary material 9Domeneviriatoi.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionDomeneviriatoi distribution: Buraco da Moura Cave, Estrela Mountain chain.File: oo_560439.tifhttps://binary.pensoft.net/file/560439A.S.P.S. Reboleira, R.P. Eus\u00e9bio6A5B9DF4-9ACF-5EC2-B7F1-3228812F5DEC10.3897/BDJ.9.e67426.suppl10Supplementary material 10Domenedarinkae.Distribution of cave-adapted beetle Data typeSpecies distribution mapBrief descriptionDomenedarinkae distribution: Santa Isabel mine, Mar\u00e3o Mountain chain.File: oo_560442.tifhttps://binary.pensoft.net/file/560442A.S.P.S. Reboleira, R.P. Eus\u00e9bio"}
+{"text": "Ebola Virus Disease (EVD) outbreaks primarily occur in the HIV endemic setting of Sub-Saharan Africa. Transient increases in HIV viral load (VL), or blips, have been described following routine vaccinations. We characterized VL blips among PLWH enrolled in a phase 2 trial of a heterologous two-dose EVD vaccine.In EBL2003, adult participants with and without HIV were randomized 1:4 to receive placebo or vaccine. Part A in the US studied MVA-BN-Filo followed by Ad26.ZEBOV 14 days later. Part B in Africa evaluated this MVA/Ad26 regimen and also a schedule of Ad26.ZEBOV followed by MVA-BN-Filo 29 days later. VL was assessed at screening, pre-vaccination, and 21, 42, 180, and 365 days post dose 2. Participants with VL < 20 copies/mL at the first 2 visits who received both doses and had complete VL data through 42 days post dose 2 were evaluated. Blips were defined as a post-injection VL \u2265 20 copies/mL no later than 42 days post dose 2, with subsequent return to VL < 20 copies/mL.A total of 277 PLWH on antiretroviral therapy (ART) were assessed; 73.3% (203) had baseline virologic suppression, and 89.2% (181) of those received both doses with complete VL data for inclusion in the analysis. Overall, 19.9% (36) experienced blips: 20.0% (29) of vaccinees vs 19.4% (7) of placebo recipients (p=1.0). All baseline suppressed participants with post-injection viremia subsequently regained suppression. Among vaccinees, the mean blip VL was 192 copies/mL, and the mean blip duration was 56 days, which was not significantly different from placebo. Of all blips, only 2 were > 1,000 copies/mL. Blips occurred in 24.0% (25) of Ad26/MVA recipients, and 9.7% (4) of MVA/Ad26 recipients (p=0.07). A dose of Ad26 was associated with a blip in 6.9% (10) of recipients vs 13.1% (19) for MVA recipients (p=0.12). Regardless of regimen, dose 1 was associated with a blip in 8.3% (12) of vaccinees, compared to 11.7% (17) of vaccinees for dose 2 (p=0.43).Among successfully treated PLWH, we observed low magnitude post-dose HIV blips that were not more common in vaccine vs. placebo recipients and did not result in loss of virologic suppression. This data is favorable for the deployment of the EVD vaccines in this trial in areas of high HIV endemicity.Benjamin L. Custer, M.D., Alexion Pharmaceuticals (Shareholder)Armata Pharmaceuticals (Shareholder)Biomarin Pharmaceutical (Shareholder)Crispr Therapeutics (Shareholder)CVS Health Corp (Shareholder)Editas Medicine (Shareholder)Gilead (Shareholder)Glaxo Smith Kline (Shareholder)Hologic Inc (Shareholder)Merck (Shareholder)Mesoblast LTD (Shareholder)Pfizer (Shareholder)Sanofi (Shareholder)Unitedhealth Group (Shareholder)Vertex Pharmaceuticals (Shareholder) Georgi Shukarev, MD, Janssen (Employee) Auguste Gaddah, PhD, Janssen Pharmaceutica N.V (Employee) Kerstin Luhn, PhD, Janssen Vaccines and Prevention  Macaya Douoguih, MD, MPH, Janssen (Employee) Cynthia Robinson, MD, Janssen Vaccines (Employee)"}
+{"text": "Once-daily oral tenofovir-based combinations as pre-exposure prophylaxis (PrEP) have shown to be an effective biomedical HIV prevention strategy for populations at-risk of acquiring HIV-1. However, low adherence can lead to poor effectiveness. This study described the characteristics of commercially-insured US PrEP users.This retrospective study used IQVIA\u2122 PharMetrics Plus data (1/1/2015\u20133/31/2020) to identify adults newly initiated (index date) on emtricitabine/tenofovir disoproxil fumarate (FTC/TDF) or emtricitabine/tenofovir alafenamide (FTC/TAF) as daily PrEP. Users had \u22656 months of continuous enrollment pre-index (baseline); those diagnosed with HIV or with antiretroviral therapy (ART) use during baseline were excluded. User characteristics were described during the baseline period. For FTC/TDF users, proportion of days covered (PDC), persistence, treatment breaks, and switching were described during the follow-up period, which spanned from index to the earliest of disenrollment or end of data. Non-persistence was defined as a >90-day gap from last day of supply, with re-initiation after this gap indicating treatment break. For PDC and persistence, follow-up was censored at HIV infection, defined by both multi-class ART initiation and HIV diagnosis. In total 24,232 FTC/TDF and 1,187 FTC/TAF users were identified. Overall, mean age was 35.1 years and 94.5% were male (Table 1). Mean [median] length of follow-up was longer for FTC/TDF (504 [390] days) than FTC/TDF users (77 [70] days). On average, FTC/TDF users had 9.0 dispensings with 38.3 days of supply per dispensing over follow-up; 11.1% had \u22651 treatment break . Among those initiated on FTC/TDF, 10.8% switched to FTC/TAF. The mean PDC for FTC/TDF users at 6 and 12 months was 0.74 and 0.67, respectively, corresponding to 63.7% and 57.9% of patients with PDC \u22650.70 . Persistence to FTC/TDF at 6 and 12 months was 70.2% and 57.4%, respectively .Table 1. Baseline Demographics and Clinical Characteristics of PrEP Users by RegimenFigure 1. Proportion of Days Covered of FTC/TDF UsersFigure 2. Kaplan-Meier Persistence Rates of FTC/TDF UsersPatient characteristics of PrEP users are broadly similar between regimens, though switching from FTC/TDF to FTC/TAF is common. FTC/TDF users had lower real-world PDC and persistence than in recent clinical trials (DISCOVER and HPTN 083).Alan Oglesby, MPH, GlaxoSmithKline (GSK)  Guillaume Germain, MSc, ViiV Healthcare  Francois Laliberte, MA, Viiv (Research Grant or Support) Staci Bush, NP, GlaxoSmithKline (GSK)  Heidi Swygard, MD, ViiV Healthcare (Employee) Sean MacKnight, MScPH, Analysis Group (Employee) Annalise Hilts, BA, Analysis Group, Inc. (Employee) Mei Sheng Duh, MPH, ScD, ViiV Healthcare (Grant/Research Support)"}
+{"text": "The innate immune system is the first to respond to invading pathogens. It is responsible for invader recognition, immune-cell recruitment, adaptive-immunity activation, and regulation of inflammation intensity. Previously, two single-nucleotide polymorphisms of innate-immunity genes \u2013 rs5743708 (Arg753Gln) of the TLR2 geneand rs8177374 (Ser180Leu) of the TIRAP gene \u2013 have been shown to be associated with both pneumonia and tuberculosis in humans, but the data are contradictory among different ethnic groups. It has also been reported thatrs10902158 at the PKP3-SIGGIR-TMEM16J genetic locus belongs to a haplotype race-specifically associated with tuberculosis. Meanwhile, a gradient of its frequency is observed in Asia. The aim of this work was to assess the effect ofselection for the genotypes of the above-mentioned SNPs on the gene pools of populations living in harsh climaticconditions that contribute to the development of infectious lung diseases. We estimated the prevalence of thesevariants in white and Asian (Chukchis and Yakuts) population samples from Northern Asia and among patients withcommunity-acquired pneumonia (CAP). Carriage of the rs5743708 A allele was found to predispose to severe CAP, whereas the GG/CT genotype of rs5743708/rs8177374 proved to be protective againstit  in white patients. No association of rs10902158 with CAP  was foundamong whites. Stratification of CAP by causative pathogen may help eliminate the current discrepancies betweendifferent studies. No significant difference in rs5743708 or rs8177374 was found between adolescent and long-livedwhite samples. Carriage of the alleles studied is probably not associated with predisposition to longevity amongwhites in Siberia. Both white and Asian populations studied were different from Western European and East Asianpopulations in the variants\u2019 prevalence. The frequency of the rs8177374 T (Ser180Leu) variant was significantly higherin the Chukchi sample  relative to the East Asian populations. This result may confirm the hypothesisabout the selection of this allele in the course of human migration into areas with unfavorable climatic conditions. Innate immunity constitutes the first barrier against microorganisms and viruses by destroying infected cells and activatingadaptive immunity. Nonetheless, an excessive nonspecificimmune reaction (inflammation) may be life threatening because it can completely disrupt the functioning of vital organs.Community-acquired pneumonia (CAP) and pulmonary tuberculosis(PTB) are infectious diseases characterized by highmortality, and according to WHO, are ranked consistentlyamong the top 10 leading causes of death in the world (https://www.who.int/en/news-room/fact-sheets/detail/the-top-10causes-of-death).Pneumonia is an inflammatory lower-respiratory-tractdisease caused by viruses, bacteria, fungi, and parasites. Inaddition, it may be due to noninfectious processes or have acombined cause. For a long time, Streptococcus pneumoniaeinfection has been considered the main cause of CAP; however, it was shown recently that CAP develops mainly as aresult of viral infections  . Streptococcus pneumoniae, Haemophilus inf\u200aluenzae,Staphylococcus aureus, Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila and othermicrobes may be causative agents of bacterial pneumonia. Afterinvasion of the airway epithelium by pathogens, these cellsstart to produce reactive oxygen species, cytokines, and othermediators to recruit immune cells. Being most abundant inlungs, alveolar macrophages ingest bacteria and apoptoticcells and can present antigens on MHC II to other immunecells. Proinflammatory M1 macrophages produce cytokinesTNF\u03b1, IL-6, IL-1\u03b2, IL-12, and IL-23 to enhance inflammationfor elimination of the invaders. Anti-inflammatory M2 macrophages produce cytokines IL-4, IL-13, and TGF-\u03b2 to inducecompletion of the inflammatory reaction and remodeling ofdamaged tissue .Depending on the set of present chemokines and cytokines,different cells responsible for humoral and cellular immunityare attracted to the site of infection . Severepneumonias are more likely to develop in coinfections; ithas been demonstrated that a viral infection (in particularinfluenza) facilitates the development of pneumococcal infection by damaging the epithelium and reducing the amount ofa surfactant . Humanrespiratory syncytial virus, metapneumovirus, adenovirus, andinfluenza viruses A and B prefer a cold season . The seasonal increase in the incidence of pneumococcalpneumonia coincides with seasonal outbreaks of influenza;S. pneumoniae, H. inf\u200aluenzae, and S. aureus infections havebeen reported to be associated with significant influenzapandemics . PTB is a pulmonary infectious disease caused mainly byMycobacterium tuberculosis (Mtb). According to the WHO,approximately one-quarter of the world\u2019s population is estimated to be infected by Mtb, and 5\u201315 % of these peoplewill fall ill with active tuberculosis. In Russia, most of thesepatients (95 %) have PTB . The pathogenesisof pulmonary tuberculosis is based on Mtb survival afterphagocytosis by alveolar macrophages. These bacteria canmodulate a host immune response to protect the infectedcells, change their metabolism, induce IL-10, suppress IL-12and TNF\u03b1 synthesis, and to inhibit MHC II expression andantigen presentation. Mtb makes macrophages unresponsiveto interferon (IFN) \u03b3 and inhibits autophagy. It allows themycobacteria to establish a persistent or latent infection inmacrophages. Mycobacteria are believed to use the generalmechanism of negative feedback regulation that restrictsexcessive inflammation . With the lossof immunity-driven control over mycobacterial reproduction, foamy macrophages accumulate in granulomas, andlung tissue necrosis begins . Vitamin Ddeficiency is known to negatively affect the effectiveness ofthe immune response in tuberculosis .These data suggest that in Northern Asia, a region with lowtemperature and reduced insolation during most of the year, signs of purifying selection for genes associated with lunginfections may be noticeable. For many genes of innate immunity, an association with viral, bacterial, and autoimmune diseases has been proven. Despite differences in the pathogenesisbetween CAP and PTB, it has been shown that minor allelesof rs5743708 (the TLR2 gene) and of rs8177374 (the TIRAPgene) can have a pathogenic and protective effect, respectively, in both of these lung diseases in humans. Nevertheless,data obtained by different research groups are contradictory.Toll-like receptors (TLRs) play a pivotal role in host defense. Being membrane-anchored  orendosomal (TLRs 3 and 7\u22129) in human immune cells , they are involved in the recognition of structurallyconserved surface molecules of microorganisms and viruses aswell as viral nucleic acids . TLR2 participates in the recognition ofa large number of diverse lipoproteins and peptidoglycansof gram-positive and gram-negative bacteria, fungi, andvirus-infected cells. After ligand binding to the receptor, TIR(Toll-interleukin 1 receptor) domains of TLR2 and TLR1 orTLR2 and TLR6 dimerize via the formation of an extensivehydrogen-bonding network and hydrophobic interactions. Homodimerizationof the cytoplasmic domains of TLR2 does not induce TNF\u03b1production in vitro in murine macrophages, and the formation of the TLR2\u2013TLR2 dimer is not detectable even in thepresence of an agonist . Therefore, the existence of TLR2\u2013TLR2 homodimersin vivo is being questioned. After ligand binding, reorientationof the TIR domains and triggering of a cascade of intracellularreactions lead to the activation of proinflammatory NF-\u03baB andMAPK pathways, synthesis and a release of proinflammatorycytokines  and various chemokines into extracellular space, and the development of aninflammatory response at the pathogen entry site . In inflammatory monocytes,TLR2 induces type I IFN production in response to a viralligand . It is reported that prolongedstimulation of TLR2 (more than 24 h) causes PI3K/Aktpathway activation in alveolar macrophages. It limits theproduction of NF-\u03baB, TNF-\u03b1, and IL-12 and activates thesynthesis of anti-inflammatory IL-10. This mechanism is assumed to prevent excessive inflammation .The TLR2 gene is located in 4q31.3, has five exons, andexpresses few splicing isoforms, but all of the coding sequences are contained within exon 3. The protein consistsof 784 amino acid residues (aa) and includes extracellularleucine-rich repeat domains, which are primarily responsiblefor ligand recognition (aa 54\u2013524), followed by the leucinerich repeat C-terminal domain (aa 525\u2013579) and intracellularTIRdomain (aa 639\u2013782), which mediates downstream signaling (https://www.uniprot.org/uniprot/O60603). It is expressedconstitutively on macrophages and dendritic cells and can beinduced in epithelial cells or B-cells. Its overexpression inpatients with pneumococcal disease had been documentedSingle-nucleotide polymorphism (SNP) rs5743708 (of theTLR2 gene) causing the Arg753Gln substitution is locatedin the TIR domain of the protein. This SNP is associatedsimultaneously with resistance to Lyme disease  and with predisposition to tuberculosis , whereas the association withpredisposition to PTB is race-specific . It is believed that TLR2 signalingmay be nonessential to control acute tuberculosis but important during chronic tuberculosis . Ameta-analysis has shown the TLR2 rs5743708 minorallele to be associated with CAP, Legionnaires\u2019 disease, andpneumococcal disease; however, the data obtained in differentstudies are contradictory .The TIRAP (TIR domain-containing adaptor protein) genealso known as Mal (MyD88 adapter-like) encodes one of thefive adapter proteins that are involved in signal transductionfrom activated TLRs to protein kinases at the plasma membrane . It is located in 11q24.2, consistsof six exons, and encodes a protein of 221 aa. The TIRAPprotein includes an N-terminal PEST domain (aa 15\u201335) responsible for binding to special sites in the plasma membrane,followed by an AB-loop mediating MyD88 and TLR4 binding.A binding site for TRAF6 (TNF receptor-associated factor 6)is located within the region aa 188\u2013193 . TIRAP is expressed in many cell types , and its isoforms resulting from alternative splicing have unknown functions. There are different opinionsabout whether TIRAP forms a complex with the TIR domainof TLR6 for signal transmission; however, it has been proventhat TIRAP mediates TLR2 and TLR4 signaling by facilitating the recruitment of the MyD88 adaptor protein to theTLRs .Activation of NF-\u03baB, MAPK1, MAPK3, and JNK resultsin cytokine secretion and an inflammatory response. SNPrs8177374 (the TIRAP gene) is located in exon 5 and represents the Ser180Leu substitution in the encoded protein. It islocated close to the TLR-binding site of TIRAP. In carriersof this substitution, modulation of TLR1, TLR2, TLR4, andTLR6 but not TLR9 signaling has been shown . Ser180Leuin a heterozygous state has a protective effect against PTB andinvasive pneumococcal disease in white and African samplesand against malaria in African and Asian samples . Carriage of heterozygous Ser180Leuprotects children from pneumococcal lower-respiratory-tractinfections, whereas carriers of the homozygous 180Leu polymorphism alone or in combination with some TLR1 and TLR6polymorphisms may be susceptible to recurrent pneumococcal infections . Simultaneous carriage ofthe TIRAP 180Leu variant and some SNPs in the TLR4 geneas well as 180Leu homozygosity increases susceptibility tosevere hospital-acquired infections .The opposite results have been obtained as well. Thers8177374 T allele (180Leu) increases the risk of PTB in asample of Iranian population . A metaanalysis of nine published case-control studies did not reveal asignificant association of 180L with tuberculosis risk . There are controversial opinions about the mechanism behind the observed protective effect of Ser180Leuheterozygosity. They are based on differences in observed effects of the SNP at the level of proinflammatory cytokines.Depending on the model used, some research groups showedan increased level and others a decreased level  of cytokines after their induction in180 Leu/Leu carriers. Accordingly, homozygosity of the minorvariant of rs8177374 is thought to cause either an excessiveinflammatory reaction or the absence of an adequate immuneresponse. It is supposed that selection pressure on the TIRAPgene provides a balance between protection against excessiveinflammation and effective defense during infectious diseases.Besides the polymorphisms in genes TLR2 and TIRAP,in this paper, we focused on the PKP3-SIGGIR-TMEM16Jgene region. An association of its haplotypes with different types of tuberculosis has been shown among childrenin Vietnam and South Africa . It is believed that the impact of the haplotypes onimmunity is determined by SIGIRR , which is a negativeregulator of TLR signaling . Carriageof rs10902158 GG and rs7111432 AA in introns of PKP3and TMEM16J, respectively, acts additively with a vitamin Ddeficiency and \u201cpathogenic\u201d genotypes of rs5743708 (TLR2)and rs8177374 (TIRAP) on tuberculosis predisposition . rs10902158 located in intron 2of the PKP3 gene has been analyzed. Encoded desmosomalplaque protein plakophilin 3 is involved in intracellular adhesion . Of note, rs10902158 has a frequency gradient in Asia; according to the Genome Aggregation Database (GnomAD) (https://gnomad.broadinstitute.org/), it is absent in South Asia and is found with a frequency of~50 % in Southeast Asia. Nonetheless, functional significanceof genetic variants in noncoding parts of the PKP3-SIGGIRTMEM16J gene region, including rs10902158, is not clear.In this work, we analyzed the frequencies of rs5743708,rs8177374, and rs10902158 (for which conflicting data onthe association with respiratory infections have been reportedpreviously) in white and Asian samples from Northern Asiaand among CAP patients. According to the statistics of theMinistry of Health of the Russian Federation, NovosibirskOblast and Yakutia are characterized by an increased incidence of PTB, whereas Chukotka Autonomous Okrug isthe leader in both pneumonia and PTB morbidity in Russia . According to the WHO, thehighest death rate from pneumonia is observed before the ageof 5 and after 75\u201380 years (https://www.who.int/medicines/areas/priority_medicines/Ch6_22Pneumo.pdf). Therefore, weassumed that long-lived people of the Siberian Federal Districtmay differ from adolescents in the frequency of rs5743708 andrs8177374, and we assessed the prevalence of the pathogenicvariants in the sample of long-lived people.The study protocol was approved by the local Ethics Committee of the Institute of Internal and Preventive Medicine . Written informed consentto be examined and to participate in the study was obtainedfrom each patient. For individuals younger than 18 years, theinformed consent was signed by a parent or legal guardian.The white sample consisted of 451 adolescents  from Oktiabr\u2019skiidistrict of Novosibirsk (55\u00b001\u2032 N 82\u00b055\u2032 E) and 289 Russiansettlers in towns Tommot (58\u00b058\u203200\u2033 N 126\u00b016\u20320\u2033 E), Neryungri(56\u00b039\u203230\u2033N 124\u00b043\u203230\u2033 E), Ust-Nera (64\u00b034\u203205\u2033 N143\u00b014\u203210\u2033 E), and Yakutsk (62\u00b001\u203238\u2033 N 129\u00b043\u203255\u2033 E), wholived in Yakutia for more than 16 years or were born there.The adolescent sample was described previously . Asian samples  consisted of130Chukchis from the Kanchalan village (65\u00b010\u203241\u2033 N 176\u00b044\u203252\u2033 E) ofChukotka Autonomous Okrug and 132 \u00adYakuts  from the Kylayy village(63\u00b013\u203234\u2033 N 132\u00b008\u203206\u2033 E) and towns Tommot, Neryungri,and Ust-Nera of Yakutia. The sample of long-lived peoplewas collected in cities Novosibirsk, Tomsk, and Tumen andconsisted of 188 individuals  aged 90\u2013105, mean age 92.Ethnicity of individuals was identified using questionnaires and additional cross-examination with elucidation ofthe nationality of ancestors (at least in three generations).Persons of mixed origin were excluded from the analysis.The CAP patient sample (406 whites) was collected in offices of pulmonary hospitals of Novosibirsk and Yakutskin 2003\u20132005 before the COVID-19 outbreak. The sampleconsists of 120 patients with severe CAP  and 286 patients with nonsevere (mild to moderateseverity) CAP . The diagnosis ofpneumonia was made on the basis of radiologically confirmed\u201cfresh\u201d lung tissue infiltration and clinical data  in theabsence of an obvious diagnostic alternative. CAP of variousetiologies was regarded as severe if the CURB65 rating scaleindex was 4\u20135 points .Genomic DNA was extracted from peripheral blood leucocytes by the standard phenol\u2013chloroform method . Genotyping was performed using polymerasechain reaction (PCR) with an analysis of restriction fragmentlength polymorphism by electrophoresis in a 5 % polyacrylamide gel after visualization with an ethidium bromide solution.The rs5743708 SNP (TLR2) was identified by amplification of DNA with primers 5\u2032-GCCATTCTCATTCTTCTGG*AGC-3\u2032 and 5\u2032-GGGAACCTAGGACTTTATCGCA-3\u2032 (* denotes a nucleotide changed for restriction site creation). The168-bp PCR product was digested with the Pst I restrictionendonuclease  for 2 h at 37 \u00b0C. Thers5743708 A (753Q) allele was revealed by the presence offragments of 20, 45, and 103 bp, whereas the rs5743708 Gallele by fragments of 20 and 148 bp.The detection of rs8177374 (TIRAP) was performed byamplification of genomic DNA with primers 5\u2032-GGCTGCACCATCCCCCA*GC-3\u2032 and 5\u2032-CCGTTCCCCTTCTCCCT CCTGTAG-3\u2032 (* denotes a nucleotide changed for restrictionsite creation). The 162-bp PCR product was digested with theAccB7 I restriction endonuclease for 2 h at 37 \u00b0C. The rs8177374 T (180L) allele was identifiedby the presence of fragments of 21 and 141 bp, whereas incase of rs8177374 C, the PCR product was not cut.Primers 5\u2032-TGGCAAGGATTGGAGAACTC*C*TGTC-3\u2032and 5\u2032-CAGGGCCAGTGCCTCCCC-3\u2032 (* denotes nucleotides changed for restriction site creation) were used forthe amplification of the PKP3 intron 2 sequence containingrs10902158. The resulting 192-bp amplicon was digested withthe BstEN I restriction endonuclease  for 2 h at 65 \u00b0C. In the presence of the rs10902158 \u0410allele, the PCR product was not cut, whereas in the case ofthe rs10902158 G allele, fragments of 24 and 168 bp wereobserved.Statistical analysis was performed in the SPSS 16.0 software.Genotype distributions were consistent with the Hardy\u2013Weinberg equilibrium among all the population samples (data notshown). Minor allele frequencies for rs5743708, rs8177374,and rs10902158 are represented in Table 1.In the sample of Novosibirsk adolescents, allele frequenciesof rs5743708, rs8177374, and rs10902158 differed from thoseof the non-Finnish European sample in GnomAD .p = 0.013; \u03c72 = 19.541, p = 0; \u03c72 = 54.554, p = 0, respectively).For the frequency of the rs5743708 A (Arg753Gln) allele, there was a tendency for a decrease in Russian settlersin Yakutia and among long-lived people compared with theNovosibirsk sample.The rs8177374 T (Ser180Leu) frequency did not differamong the studied white samplesIn the sample of Russian settlers of Yakutia, males andfemales differed in the frequency of rs10902158 . Moreover, the frequency of this SNP among maleswas closer to that observed in non-Finnish Europeans according to GnomAD data, and the frequency among females wascloser to that observed in the Novosibirsk sample. Perhapsthere were sex differences during recent migration to Yakutiafrom different regions of Russia. Genetic analysis of a largersample and estimation of this SNP\u2019s frequency in westernregions of Russia are required for explaining the observeddifferencesThe two analyzed Asian samples differed from each otherand from GnomAD East Asian cohorts. Among Yakuts, thers5743708A (Arg753Gln) variant, which is very rare in otherAsian populations, was found at a frequency of 0.015\u00b10.007(mean\u00b1SD). Chukchis differed significantly from GnomADEast Asians in rs10902158 allele frequency (see Table 1).Frequencies of polymorphisms rs5743708, rs8177374, andrs10902158 were not different among adolescents and totalwhite CAP patient samples. By contrast, after the samplewas divided into patients with severe and nonsevere CAP,differences were found for rs5743708 .Next, genotype frequencies were estimated for rs5743708,rs8177374, and rs10902158 in the examined samples (exceptfor long-lived people regarding rs10902158). For the latterSNP (in the PKP3 gene), no difference in frequency wasdetectable within any group (data not shown). The observeddistribution of rs5743708 and rs8177374 genotypes amongthe studied samples is presented in Table 2.The frequencies of genotypes of rs5743708 and rs8177374did not differ among the following samples: adolescents ofNovosibirsk, long-lived people of Siberia, all patients withCAP, and all Russians in Yakutia. Possibly, the carriage of thestudied alleles is not associated with predisposition to longevity in Siberia and does not significantly affect the probabilityof resettling of migrants in more unfavorable climatic conditions at present. This notion is consistent with WHO findingsthat in Eastern Europe, in contrast to Western and CentralEurope, the pneumonia mortality rate does not increase significantly after age 80 (https://www.who.int/medicines/areas/priority_medicines/Ch6_22Pneumo.pdf). As for pneumonia,carriage of the rs5743708 A allele predisposed to severe CAP. The heterozygous genotype of rs8177374 in combination with the GG genotype ofrs5743708 had a protective effect against severe CAP .It was shown here that carriage of none of the three studiedSNPs, rs5743708, rs8177374, and rs10902158, is associatedwith the predisposition to CAP in total. By contrast, we foundthat the Arg753Gln variant of TLR2 predisposes to severeCAP, and the heterozygous Ser180Leu variant of TIRAP incombination with the 753 Arg/Arg variant of TLR2 has aprotective effect against it in the white population. These datapartially explain the contradictions in the data from differentresearchers. Most likely, the contribution of the alleles ofgenes TLR2 and TIRAP to CAP predisposition is determinedby pneumonia etiology. A substantial proportion of severepneumonia cases are known to be caused by combined viraland bacterial infections . TLR2 is responsible mainly forthe recognition of bacteria-associated molecular patterns;Ser180Leu of the TIRAP gene modulates signal transductiononly from TLR2 and TLR4 recognizing molecular patterns ofbacteria as well . Most likely, combinedand bacterial but not viral pneumonias are associated withTLR2 and TIRAP gene variants.The studied Asian ethno-geographical groups showed an increased frequency of the protective rs8177374 T (Ser180Leu)variant of TIRAP relative to neighboring East Asian populations. In the Chukchi sample, the difference was significant. It may be a consequence of the naturalselection that has promoted protection from excessive inflammation during pulmonary diseases. The hypothesis about theselection of the Ser180Leu variant along with the out-ofAfrica migration to a harsh environment has been advancedearlier . An increasedfrequency of the Arg753Gln variant of TLR2 and a decreasedfrequency of Ser180Leu of TIRAP as compared to non-Finnish Europeans may indicate higher genetic predispositionof the Siberian white population to PTB and severe CAP.Nevertheless, there are a lot of genes associated with CAPand PTB independently. Apparently, during the settlement ofpeoples in Northern Eurasia, the formation of gene pools hadbeen determined by the selection that facilitated adaptationto specific infections (Lime disease among them), parasites,and the climate. It would be interesting to determine why twomutations changing the same TLR2 signaling have oppositeeffects on the predisposition to severe CAP. One possible explanation is the difference in the structure and functions ofheterodimers TLR2\u2013TLR1 and TLR2\u2013TLR6 .Existing data on the roles of TLRs 1, 2, and 6 in the activation of proinflammatory and anti-inflammatory signaling areconflicting. Overexpression of TLR2 carrying the Arg753Glnvariant has been demonstrated to cause a significantly strongerimpairment of cytokine induction by TLR2/TLR1 ligands ascompared with TLR2/TLR6 ligands in the HEK293 cell line. In later papers, it has been shown thatthe Arg-to-Gln substitution at position 753 of TLR2 changesthe size, charge, and hydrophobic properties of this site andreduces the ability of TLR2 to form a heterodimer with TLR6. This SNP significantlyalters agonist-inducible association of TLR2 with adaptorproteins TIRAP and MyD88 and impairs NF-\u03baB signaling andIL-8 mRNA expression in the HEK293 cell line . Genes TLR1 and TLR2 have different expression activators . In the TLR2\u2013TLR1 dimer, TLR1and TLR2 are responsible for NF-\u03baB and MAPK pathways and for PI3K pathway activation, respectively; therefore, upregulation of proinflammatory cytokines is TLR1-dependent,whereas upregulation of type I IFN is TLR2-dependent . TLR2\u2013TLR6 binding disruption possibly causesan increase in the number of TLR2\u2013TLR1 dimers in whichTLR1 drives the activation of the NF-\u03baB inflammatory signaling cascade. On the contrary, the Ser180Leu variant of TIRAPweakens proinflammatory signal transmission. Besides,TIRAP acts as an adaptor protein for TLR4 homodimers.This receptor is primarily responsible for the recognitionof lipopolysaccharides of gram-negative bacteria and fungi. It is believed that TLR4 takes part notonly in MyD88-dependent proinf\u200alammatory signaling but alsoin MyD88-independent anti-inf\u200alammatory signaling. Weakening of TLR4 signaling through TIRAP probably enhances thesignaling through adapter proteins TRIF and TRAM, causingthe secretion of anti-inf\u200alammatory cytokines .It remains unclear why the SNPs in TLR2 and TIRAP havesimilar effects on the predisposition to or protection againstacute (CAP) and chronic (PTB) lung infections. Perhaps thisphenomenon is due to an impact on inflammation in both diseases. The severity of CAP is determined by life-threateningacute inflammation; in PTB, the development of chronic inflammation masks the infection from the host immune system.In Northern Asian populations, the observed difference inrs8177374 frequency may reflect consequences of naturalselection during the settlement of peoples on territories withunfavorable climatic conditions. As for pneumonia, carriage ofthe rs5743708 A allele (the TLR2 gene) predisposes to severeCAP; the heterozygous genotype of rs8177374 (the TIRAPgene) in combination with the GG genotype of rs5743708 (theTLR2 gene) has a protective effect against it. Stratification ofCAP by causative pathogen may help to eliminate the currentdiscrepancies among research groups. Regional differencesin a set of pathogens along with the genetic characteristics ofethno-geographical groups can determine the associations ofvarious genetic variants of innate immunity with the prevalence and severity of pneumonia.The authors declare no conflict of interest.Aguilera E.R., Lenz L.L. Inflammation as a modulator of host susceptibility to pulmonary influenza, pneumococcal, and co-infections.Front. Immunol. 2020;11:105. 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+{"text": "Correction to: Borderline Personal Disord Emot Dysregul 7, 19 (2020).https://doi.org/10.1186/s40479-020-00132-8Following publication of this article , it is rThe correct reference 35 should be:35. Green A, Charles K. Voicing the Victims of Narcissistic Partners: A Qualitative Analysis of Responses to Narcissistic Injury and Self-Esteem Regulation. SAGE Open. 2019;9(2). 10.1177/2158244019846693.The original article has been updated."}
+{"text": "The construction of a nanoimmune controlled-release system that spatiotemporally recognizes tumor lesions and stimulates the immune system response step by step is one of the most potent cancer treatment strategies for improving the sensitivity of immunotherapy response.Here, a composite nanostimulator (CNS) was constructed for the release of second near-infrared (NIR-II) photothermal-mediated immune agents, thereby achieving spatiotemporally controllable photothermal-synergized immunotherapy. CNS nanoparticles comprise thermosensitive liposomes as an outer shell and are internally loaded with a NIR-II photothermal agent, copper sulfide (CuS), toll-like receptor-9 (TLR-9) agonist, cytosine-phospho-guanine oligodeoxynucleotides, and programmed death-ligand 1 (PD-L1) inhibitors (JQ1). Following NIR-II photoirradiation, CuS enabled the rapid elevation of localized temperature, achieving tumor ablation and induction of immunogenic cell death (ICD) as well as disruption of the lipid shell, enabling the precise release of two immune-therapeutical drugs in the tumor region. Combining ICD, TLR-9 stimulation, and inhibited expression of PD-L1 allows the subsequent enhancement of dendritic cell maturation and increases infiltration of cytotoxic T lymphocytes, facilitating regional antitumor immune responses.CNS nanoparticle-mediated photothermal-synergized immunotherapy efficiently suppressed the growth of primary and distant tumors in two mouse models and prevented pulmonary metastasis. This study thus provides a novel sight into photo-controllably safe and efficient immunotherapy.The online version contains supplementary material available at 10.1186/s12951-021-01197-5. Tumor immunotherapy that mobilizes immune cells to fight malignant tumors is a promising therapeutic approach , 2. Dive. recently developed an activatable engineered immune device, which can be remotely controlled by near-infrared (NIR) light. In this scenario, NIR light excitation was used to break the photoresponsive linker, releasing active CpG oligonucleotides (ODNs) and precisely activating local intratumoral inflammation without obvious systemic side effects  in the primary tumors compared with the CNS0\u2009+\u2009L, CNSC\u2009+\u2009L, and CNSJ\u2009+\u2009L groups , CRT, and expression of high mobility group box 1 protein (HMGB1) were used to facilitate the uptake, processing, and presentation of tumor antigens in DCs. One day after treatment, the intratumoral ATP levels of various CNS nanoparticles plus laser irradiation were elevated and compared with the single PBS-treated and CNSely Fig.\u00a0a. MoreovPBS Fig.\u00a0b. In conals Fig.\u00a0c. These +CD80+CD86+)  via flow cytometric analysis to investigate the CNS-mediated synergized antitumor immune response  in the CNSD\u2009+\u2009L group were 1.6, 1.2, and 1.2-fold higher than those in the CNS0\u2009+\u2009L, CNSC\u2009+\u2009L, and CNSJ\u2009+\u2009L groups, respectively , is a key indicator of an antitumor immune response  = 100 \u00b5g mL-1) under the NIR-II laser irradiation  for 5 min. Fig. S3. The time constant for heat transfer from the system is determined by applying the linear time data from the cooling period of (a) versus the negative natural logarithm of driving force temperature. Fig. S4. Release of CpG from CNSD with or without laser irradiation  for 5 min . Fig. S5. Schematic illustration of the synthesis of ICG-loaded CNS nanoparticles  for tracing the nanoparticle trajectory. Fig. S6. UV-vis absorption spectrums of CNS0@ICG, CNSC@ICG, CNSJ@ICG, and CNSD@ICG. Fig. S7. Fluorescence intensity of 4T1 cells treated with PBS (control) or various ICG-loaded CNS nanoparticles ([ICG] = 20 \u00b5g mL\u22121) for 24 h via flow cytometry. Fig. S8. Relative mean fluorescence intensity (MFI) of CRT in Panc02 cells after different treatments. Fig. S9. The NIR fluorescence imaging of xenograft Panc02tumor-bearing C57BL/6 living mice at 0, 8, 24, and 36 h after systemic administration of CNS@ICG through tail-vein administration . The fluorescence images were collected with excitation at 710 nm and emission at 790 nm, and the tumors were marked by white circles. (b) The fluorescence intensity of tumor regions of mice at different post-injection times of (n = 3). Fig. S10. (a) NIR thermal photos of 4T1 tumor-bearing mice under laser irradiation at 24 h post-injection of PBS, CNS0, CNSC, CNSJ, and CNSD through tail-vein injection ; (b) Temperature elevation curves of tumors in 4T1 tumor-bearing mice after administration of Control (PBS), CNS0, CNSC, CNSJ and CNSD under NIR-II laser illumination . Fig. S11. Gating strategies for flow cytometry assay of matured CD80+CD86+ DCs in tumor-draining lymph nodes of Panc02 tumor-bearing C57BL/6 mice. Fig. S12. Western-blot analysis of JQ1-induced downregulation of PD-L1 in Panc02 cells (200 nM of JQ1 and 100 ng/ml of IFN-\u03b3). Fig. S13. Gating strategy for flow cytometry assay of CD4+ T cells and CD8+ T cells in distant tumors of Panc02 tumor-bearing C57BL/6 mice. Fig. S14. A volcano plot showing the up-regulated or insignificantly expressed or down-regulated genes when comparing the CNSD-treated group with the Control (PBS) group. Fig. S15. Body weights of Panc02 tumor-bearing C57BL/6 mice in different groups within 14 days of treatment (Control (PBS), CNS0, CNSC, CNSJ and CNSD, 0.2 mL, [CuS] = 300 \u03bcg/mL, n = 5). Fig. S16. Body weights of 4T1 tumor-bearing Balb/c mice in different groups within 14 days of treatment (Control (PBS), CNS0, CNSC, CNSJ and CNSD, 0.2 mL, [CuS] = 300 \u03bcg/mL, n = 5). Fig. S17. Representative histological H&E staining images of major organs  were collected from Panc02 tumor-bearing C57BL/6 mice in different treatment groups at the end of treatment (Control (PBS), CNS0, CNSC, CNSJ and CNSD, 0.2 mL, [CuS] = 300 \u03bcg/mL). The scale bar represents 50 \u03bcm. Fig. S18. Representative histological H&E staining photos of major organs  in healthy C57BL/6 mice before treatment (Control) and after tail-intravenous injection of CNSD  for 15 (Day 15) and 30 days (Day 30). The scale bar represents 50 \u03bcm. Fig. S19. The levels of (a) alanine aminotransferase (ALT), (b) aspartate aminotransferase (AST), (c) \u03b3-glutamyl transpeptidase (GGT), (d) urea, (e) creatinine (CREA), (f) red blood cells (RBC), (g) hemoglobin (HGB), (h) mean corpuscular hemoglobin (MCH), (i) hemoglobin concentration (MCHC), (j) red cell distribution width (RDW-SD), (k) red cell volume distribution width (RDW-CV), (l) platelet (PLT), (m) plateletcrit (PCT), (n) mean platelet volume (MPV), and (o) platelet distribution width (PDW) in the blood of healthy C57BL/6 mice before treatment (D0) and after tail-intravenous administration of CNSD  for 15 (D15), and 30 days (D30) (n = 3)."}
+{"text": "Vision loss is associated with restricted physical activity (PA), yet the relationship between multiple domains of vision measures and objectively measured PA, especially activity patterns, in mid-to-late life remains unclear. In 603 BLSA participants , best-corrected and presenting visual acuity (VA), contrast sensitivity, visual fields (VF), stereo acuity were assessed from 2015 to 2019. Free-living PA was assessed using a wrist-worn ActiGraph accelerometer for 7 days. Linear regression models showed that participants with vs. without best-corrected VA impairment had 29.3 fewer active minutes/day (p=0.03) and trended towards fewer activity counts (p=0.05), adjusting for sociodemographic and health characteristics. VF impairment was associated with 268,636 fewer activity counts (p=0.02), 46.2 fewer active minutes/day (p=0.02), and a 3% greater activity fragmentation (p=0.009). Older adults with visual impairment have restricted and more fragmented activity patterns. Longitudinal studies are warranted to examine causality between visual impairment and PA decline."}
+{"text": "The risk of cervical cancer is caused by persistent human papillomavirus (HPV) infection. Cervical cancer is the most frequent cancer among women. Our purpose was to investigate the association between TP53 215C>G (Pro72Arg), MDM2 -410T>G, and NQO1 609C>T gene polymorphisms with a high HPV load and the inf luence of gene-gene interactions on prolonged HPV infection. Eighty-nine women with a high HPV viral load and 114 healthy women were involved in a case\u2013control study. Genotyping for TP53 215C>G (Pro72Arg) and MDM2 -410T>G SNPs was carried out by allele-specif ic PCR and genotyping for NQO1 609C>T was performed by a TaqMan assay. Quantitative analysis of HPV DNA was performed by AmpliSens\u00ae HPV HCR screen-titer-FRT test system. Gene-gene interactions were analyzed using the multifactor dimensionality reduction (MDR) method. The study of separate SNPs of MDM2 -410T>G and NQO1 609C>T genes did not reveal any statistically signif icant difference in genotype and allele frequencies among women within the two groups. The frequency of the 215G (72Arg) allele and 215GG (72Arg/ Arg) genotype of the TP53 gene was signif icantly higher in the case group than in the control group . MDR analysis showed the signif icance of intergenic interactions of the three studied loci TP53 (rs1042522) \u2013 MDM2 (rs2279744) \u2013 NQO1 (rs1800566) for the formation of a high HPV load . Human papillomavirus (HPV) is implicated in the developmentof cervical cancer. A key critical step in papillomavirusrelatedcarcinogenesis is a persistent viral infection . There is heterogeneity in the development of humanpapillomavirus infection due to genetic variations, ethnicity,viral types involved in infection, viral load, and oncogenicexpression, as well as environmental, and hormonal, physiological,and nutritional factors . After HPV-infection, especially with high-riskHPV types ,HPV oncoproteins induce mutations in oncogenes, epigeneticmodifications, and chromosomal rearrangements . A disequilibrium in the relationship betweenvirus and host results in a decrease in the effectiveness of theimmune system, the imbalance between cellular and humoralimmune processes, as well as alteration in pro- and antiinflammatorycytokine levels, which increases the replicativeability of the virus . In addition, modifications that alter the stability of cellcycle proteins such as retinoblastoma protein (pRb), tumorsuppressor p53, result in uncontrolled cell cycle progressionand induce oncogenic transformation of cells .The TP53 tumor suppressor gene plays a crucial role inregulating DNA repair, apoptosis, and cell cycle control. Ithas been observed that most human tumors contain mutatedp53, with about 50 % of those mutations causing a reductionin DNA repair ability, irregular cell growth, and, eventually,progression to malignancy . Polymorphismsin the TP53 gene change p53 protein conformation,which leads to p53 degradation through a process mediatedby ubiquitin . The most widely studiedof the non-synonymous SNP TP53 Pro72Arg (rs1042522)replaces proline (Pro) with arginine (Arg) in the p53 proteindue to a substituted C to G base in the TP53 gene. Both variantshave the same binding affinity for DNA while their ability tobind components of the transcription factor is different. So, thetwo variants of the p53 protein are not functionally equivalent. The p53 is ubiquitinated in the proteasome,which is regulated by MDM2 via a ubiquitin-dependentdegradation pathway and NAD(P)H quinone oxidoreductase 1via a ubiquitin-independent degradation pathway . As a result, the levelof p53 is affected by MDM2 and NQO1 activity.Oncoprotein MDM2 is a negative regulator of the p53 tumorprotein . A functional SNP in theMDM2 gene promoter (-410T>G rs2279744) regulates MDM2protein expression. When T is replaced with G, this increasesthe affinity of the transcriptional activator Sp1, resulting inhigher MDM2 expression and subsequent suppression of thep53 pathway .The NQO1 enzyme can catalyze the reduction of variousquinones to hydroquinones by a two-electron reductionmechanism (NADH or NADPH) as a reducing cofactor. Thistwo-electron reduction prevents the formation of free radicals(semiquinones) that protect the cells from oxidative stress. The SNP of NQO1 at nucleotideposition 609C>T in exon 6 (rs1800566) with the proline toserine amino acid substitution at codon 187 induces a changeof enzyme activity. The homozygotes (TT ) genotype gives riseto an inactive enzyme NQO1, heterozygotes (CT ) have theenzyme displays mild activity, while the wild homozygotes(CC) have the highest activity of the NQO1 . Wild type NQO1 partially inhibits HPV E6-mediatedp53 degradation, although this does not occur with the mutanttype NQO1 .Thus, the efficiency of the cell cycle repair and controlsystem depends not only on the p53 protein. Also, the levels ofMDM2 and NQO1 proteins in the cell can affect the stabilityof the p53 protein and the activity of its degradation processes.However, human papillomavirus, as an exogenous factor, canbe an additional cause affecting the work of the repair system.Most of the studies on the association of SNPs of genes withHPV infection and cervical cancer are devoted to the analysisof individual nucleotide substitutions. There is practically nodata in the literature on the combined effect of polymorphicvariants of these three genes in the presence of HPV load.Our work aims to analyze the distribution of the polymorphismsof the TP53 gene (rs1042522), MDM2 gene(rs2279744), and NQO1 gene (rs1800566) in patients withHPV load versus HPV-negative women.Two hundred and three samples of epithelial cells scraped fromthe urogenital tract of women were used for molecular geneticstudies. The study equipment has been provided by the clinicaldiagnostic laboratory, Nauka . Thewomen were divided into two groups: women with a highHPV load (above 3 log of HPV genomes per 100 thousandhuman cells) (n = 89), and HPV-negative women (n = 114).All the women included in the study were over thirty yearsold. Criteria for women being included in the control group:a normal result of colposcopy, HPV-negative PCR-test. Thecomparative group of cases included women with symptomssuch as vaginal discharge, bleeding menstrual abnormalities,and HPV-positive PCR-test with an HPV load of morethan 3 log of HPV genomes per 105 human cells. The ethniccomposition of the women involved in the study groups was asfollows: Russians accounted for 86 %, Armenians accountedfor 9 %, and other nationalities of the Caucasian race \u2013 5 %.All women have given formal written consent to take partin the study. The study was approved by the Bioethics Committeeof the Academy of Biology and Biotechnology of theSouthern Federal University . All the tests for clinical experimentation were carriedout in line with the standards and ethical guidelines of theWorld Medical Association (Helsinki Declaration).The total DNA was isolated from scraping epithelial cells ofthe cervical canal of women according to the DNA-sorb-AM reagent kit protocol. The quantification ofDNA for high-risk HPV types  in biological material was analyzed accordingto the AmpliSens-HPV HCR screen-titre-FRT PCRkit  protocol. According to the kitmanufacturer\u2019s instructions and clinical reports, the viral loadis interpreted as follows: log \u2264 3 per 105 human cells \u2013 lowclinical significance, 3\u20135 log per 105 human cells \u2013 clinicallysignificance, risk of dysplasia; and > 5 log per 105 humancells \u2013 clinically significance, strongly probable dysplasia.Genotyping was performed for the SNP of TP53 215C>G(Pro72Arg) (rs1042522), MDM2 -410T>G (rs2279744) genesby allele-specific PCR and the SNP-express reagent  according to the kit protocol. NQO1 609C>T(rs1800566) was genotyped by a TaqMan genotyping assay.The amplification was carried out in a 25-ml reaction containing2 \u03bcl 25 mM MgCl2, 1 \u03bcl 10 mmol/L of the forward primer(5\u2032-CAG AGT GGC ATT CTG CAT TTC T-3\u2032) and reverse(5\u2032-CTG GAG TGT GCC CAA TGC TA-3\u2032) primers and0.5 \u03bcl mmol/L NQO1 wild-type (5\u2032-6FAM-CTT AGA ACCTCA ACT GA-MGBNFQ-3\u2032) and mutant (5\u2032-VIC-CTT AGAATC TCA ACT GAC A-MGBNFQ-3\u2032) probes, 0.5 \u03bcl Taqpolymerase(5 U/\u03bcl), 2.5 \u03bcl 2.5 \u043c\u041c of dNTP, 2 \u03bcl 10 \u00d7 PCRbuffer B, 12 \u03bcl ddH2O  and 3 \u03bcl DNA. Cyclingconditions were as follows: initial denaturation at 95 \u00b0C for10 min, followed by 40 cycles consisting of denaturationat 95 \u00b0C for 15 sec, then annealing at 54 \u00b0C for 60 sec. ThePCR products for NQO1 609C>T (rs1800566) were analyzedin real-time using RotorGene thermocycler. PCR productsfor the TP53 Pro72Arg and MDM2 -410T>G genes wereanalyzed by 3 % agarose gel horizontal electrophoresis andvisualized under the ultraviolet (UV) transilluminator GelDoc.To calculate the statistical data, the \u03c72 test was used to comparethe allele and genotype frequencies of the TP53 215C>G(Pro72Arg) (rs1042522), MDM2 -410T>G (rs2279744), andNQO1 609C>T (rs1800566) genes in the case group andcontrol group. The Hardy\u2013Weinberg equilibrium test wasperformed to determine the goodness-of-fit of the \u03c72 test withone degree of freedom by comparing the observed genotypefrequencies with the expected genotype frequencies. The SNPgenetic association was assessed by the \u03c72 test, odds ratio(OR), and its confidence interval (CI). A p-value <0.05 wasconsidered statistically significant. Statistical analyses wereperformed using GraphPad InStat 3.05 software.The analysis of intergenic interactions was performed usingthe MDR software  and by using the exhaustive search algorithm.All potential combinations of genotypes were evaluated withrespect to the risk of developing an HPV infection. Multilocusgenotypes are summed up in the MDR program into groups ofincreased and reduced disease risk, which reduces the dimensionof the number of calculated parameters. Using multiplecross-recalculations of the input primary data, the optimalmodel is selected for intergenic interaction, with the highestaccuracy and, accordingly, with the least error, to predict thepresence or absence of predisposition to the studied pathology.In 89 HPV-positive women, the average age was 40.1 \u00b1 7.3 yearsand 41.1 \u00b1 7.6 years in 114 HPV-negative women. Among 89HPV-infected women the minimum, middle, and maximumHPV DNA load were 3.2, 5.1, and 8.6 log of HPV genomesper 100 thousand human cells, respectivelyFrequency distributions for the three investigated TP53,MDM2, and NQO1 gene polymorphisms are given in Table 1.The polymorphic variants of MDM2 -410T>G and NQO1609C>T were not associated with a high HPV load. At thesame time females with a high HPV load had a significantlyhigher frequency of TP53 215G (72Arg) allele  and 215GG (72ArgArg)genotype  than healthy controls. The existence of multiple allelic variationsin genes that encode for protein molecules can lead to severalrelated changes in the genome and proteome function. Therefore,an analysis of intergenic interactions of allelic variantswas conducted.An analysis of intergenic interactions showed that the threelocusmodel of gene interaction has a prediction accuracy of64 % and cross-validation consistency (10/10) (Table 2). Interactionof TP53 (rs1042522) \u2013 MDM2 (rs2279744) \u2013 NQO1(rs1800566) genes is associated with the risk of high HPV loadamong women .A radial diagram demonstrates the contribution of polymorphismof each gene, both individually and in combination withothers for the three-loci. In the vertices of the diagram, thevalues of information for individual genes are indicated, onthe edges \u2013 the information value of the interaction of a pair ofgenes. The studied SNPs affect the formation of the viral loadto varying degrees. According to the model of loci interaction, the highest predictive potential is possessedby the SPN of the TP53 gene (2.26 %). The TP53 and MDM2loci have the greatest effect by intergenic interaction. A pronouncedsynergism was revealed between these loci \u2013 the totaleffect of the combination is 2.87 %. Its information value ishigher than the sum of its individual effects.Cervical cancer is the most common gynecological canceramong women and the high-risk HPV genotypes play a majorrole in abnormal lesion development and cervical malignant neoplasms . The presenceof a high viral load in HPV-positive women indicates that thevirus has not been entirely removed and will likely continueto replicate in the body cells for a long time. Long-term viruspersistence contributes to the incorporation of HPV DNAinto the human genome, the expression of E6/E7 oncogenicproteins, and the development of cancer .Human papillomatosis appears to be a polygenic disease,suggesting that recurrent, small-effect genetic variations canhave consequences for disease susceptibility . Tumor development is largely attributed to geneticvariations in the host\u2019s cell cycle control .The relationships between the TP53 gene (rs1042522), MDM2gene (rs2279744), NQO1 gene (rs1800566), and the risk ofhigh HPV load have been investigated in this studyIn our study, 43.8 % of women (89 out of 203) were positivefor high-risk HPV types. Our analysis revealed an associationof high viral load formation risk with 215G (72Arg)allele carriage and 215GG (72Arg/Arg) genotype of TP53 gene . On the contrary, 215C (Pro72)allele  and 215CC(72ProPro) genotype  showed protective effect compared to the controlgroup. The polymorphic variants of the p53 protein with Proor Arg in codon 72 have been shown to vary in the efficiencyof interaction with the E6 oncoprotein of HPV . The Arg variant is degraded by the E6 oncoproteinmore readily than the Pro variant. Therefore, the carriers ofthe Arg/Arg genotype have p53 protein more vulnerable toviral protein-induced degradation . Our resultsare consistent with several other studies suggesting that HPVpositivewomen are more vulnerable to cervical malignantneoplasms when having \u0422\u042053 72Arg/Arg genotype and \u0422\u04205372Arg allele .MDM2 promotes cell cycle progression through the activationof S-phase, via interaction with the retinoblastoma tumorsuppressor protein and the transcriptional factor E2F . MDM2 is one of the central nodes in the p53pathway regulation. It has been shown that even a small changein MDM2 level may affect the p53 pathway and, subsequently,cancer development . Our analysisshowed no statistically significant difference in the genotypes( p = 0.86) and allele frequencies ( p = 0.68) distribution ofMDM2 -410T>G gene polymorphism in two women groups.Our analysis showed no statistically significant differencein the genotypes ( p = 0.29) and allele ( p = 0.18) distributionof NQO1 609C>T gene polymorphism in the two groups ofwomen. In agreement with our results, J. Chansaenroj andhis coworkers showed no association of the NQO1 609C>Tpolymorphism with the risk of cervical cancer . At the same time, several studies reported arelationship between NQO1 609TT genotypes and the riskof cervical cancer . TheNQO1 gene (rs1800566) TT genotype is associated with nullenzyme activity and could influence cancer progression byreducing cytotoxic agents containing the quinone moiety.Favorable conditions for HPV persistence include multiplegenetic substitutions which result in gene expressionchanges. In our work, the analysis of gene-gene interactions(MDR) showed significant interaction of the polymorphic loci for increasedviral load (see Table 2). The interaction of the polymorphicvariantsfor the three loci of the genes TP53 215C>G (Pro-72Arg), MDM2 -410T>G, and NQO1 609C>T are associatedwith HPV viral load increase.A synergistic effect was revealed between the studied loci.That is, the combined effect of these loci is more pronouncedthan individual effects. Thus, we revealed an increased risk ofa high viral load in HPV infection in the case of a combinationof polymorphic variants of the TP53, MDM2, and NQO1genes. The risk may be due to disturbances in the work ofthe checkpoints of the cell cycle due to the activation of theprocesses of degradation of the p53 protein.The current study has several limitations. First, the smallsample size: our results should be verified in larger populationsas well as in other ethnic groups. Second, women with cervicalcancer were not included in our research. Comparison of thedifferent histological types of cervical cancer may also be warrantedfor future studies to determine whether the frequencyof TP53, MDM2, and NQO1 gene polymorphisms differ basedon the histological types of cervical cancer. Third, the influenceof epidemiologic risk factors such as smoking, alcoholintake, and sexual behavior or pathogenic factors like bacteriawith the risk of HPV infection was not included. It would beinteresting to analyze if TP53, MDM2, and NQO1 productionis associated with environmental or pathogenic factors.Our results demonstrate that the risk of high viral load formationis associated with TP53 215G (72Arg) allele andTP53 215GG (72ArgArg) genotype in HPV-positive women.Although the individual SNPs of MDM2 -410T>G andNQO1 609C>T genes did not reveal a statistically significantfrequency difference in our study, intergenic interactionsanalysis revealed significant interaction for all polymorphicvariants. This demonstrated that the infection developmentdepends on the synergistic effect of several polymorphismsthat induce changes in gene expression and represent an allelicload for HPV-positive cells. However, the role of thegenetic susceptibility to HPV infections and high HPV loadwith TP53 rs1042522, MDM2 rs2279744, NQO1 rs1800566polymorphisms requires further investigationThe authors declare no conflict of interest.Atia A., Abdullah A. 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+{"text": "In this study, we investigated the protective potential of B-LAP against diabetic nephropathy in streptozotocin (STZ) induced diabetic mice. Diabetes induction in mice was carried out by a single intraperitoneal injection of STZ. 2.5 mg/kg/day and 5 mg/kg/day doses of B-LAP were administered orally for twelve weeks and renal histoarchitecture, caspase-3, tumor necrosis factor-alpha (TNF-\u03b1), malondialdehyde (MDA), glutathione peroxidase (GPX), as well as urinary nephrin and neutrophil gelatinase-associated lipocalin (NGAL) were evaluated. Additionally, kidney levels of PI3K, phosphorylated (p)-Akt, p-mTOR, p-CREB, and SIRT1 were assessed in the present investigation. 5 mg/kg B-LAP significantly decreased urinary excretions of nephrin and NGAL. It also mitigated renal TNF-\u03b1 and MDA levels and simultaneously improved GPX activities. 5 mg/kg B-LAP improved renal function in diabetic mice as indicated by elevated values of creatinine clearance. While B-LAP elevated renal levels of SIRT1, it alleviated PI3K, p-Akt, p-mTOR, and p-CREB levels in the kidneys of diabetic mice.Collectively, these findings suggest B-LAP as a potential renoprotective agent in STZ-induced diabetic mice probably via modulating the PI3K/Akt/mTOR pathway. Diabetic nephropathy (DN) is the main cause of end-stage renal disease (ESRD) across the world, which develops in one-third of both type 1 and type 2 diabetic patients , IL-6, and tumor necrosis factor-alpha (TNF-\u03b1) levels via modulating NF-\u03baB and MAPK/ERK signaling pathways in vitro (\u03b2-LAPachone (B-LAP), the natural agent derived from the Lapacho tree, in vitro . Newer cin vitro -10. Morein vitro .The present investigation aimed to examine potential renoprotective effects of B-LAP in streptozotocin (STZ)-induced diabetic mice and to evaluate its impact on the PI3K/Akt/mTOR pathway in the kidneys.ChemicalsThe following commercially available chemicals were obtained: B-LAP  and STZ .Experimental designThirty male C57BL/6 mice were obtained from the Pasteur Institute of Iran. Diabetes induction was done by injecting a 50 mg/kg dose of STZ dissolved in citrate buffer (pH: 4.5) . InductiMice were assigned into 5 groups of 6 animals per each; 1- sham receiving B-LAP vehicle, i.e., 0.05% DMSO; 2- diabetic control mice (Diab); 3- Diab + low dose B-LAP (2.5 mg/kg/day); 4- Diab + high dose B-LAP (5 mg/kg/day); 5- Normal mice receiving 5 mg/kg/day B-LAP. Treatment doses of B-LAP were adopted according to a previously conducted similar study and the agent was administered via intra-gastric gavage . After 1Measurement of systolic blood pressureThe systolic blood pressure (SBP) readings were recorded a day before placing the mice in the metabolic cages using a non-invasive tail-cuff method . Animals were placed in a heated restrainer at 37 \u00b1 1 \u00b0C for 10 min during measurements. For each mouse, 3 blood pressures were measured and their average was taken as the SBP.Biochemical and immunochemical assessmentsIn order to calculate Ccr, first serum and urinary levels of creatinine were assayed via commercially available kits . Creatinine clearance (Ccr) was then calculated using the values for urinary creatinine, urine volume, serum creatinine, and body weight :Ccr (mL/min/kg) = (urine creatinine (mg/dL) \u00d7 urine volume (mL)) / (serum creatinine (mg/dL) \u00d7 1440 (min)) \u00d7 1000 / (body weight (g))Renal malondialdehyde (MDA) and glutathione peroxidase (GPX) activities were measured spectrophotometrically using the commercially available kits . Obtained values were normalized by renal total protein levels measured by the Lowry method.Tumor necrosis factor-alpha (TNF-\u03b1), nephrin, and neutrophil gelatinase-associated lipocalin (NGAL) levels were assayed by using enzyme-linked immunosorbent assay (ELISA) kits .Histopathological examinationsRenal tissue specimens were fixed in 10% neutral buffered formalin (NBF) and processed for paraffin sections of five-\u03bcm thickness. Sections were stained with hematoxylin and eosin (H&E) and were qualitatively examined under a light microscope .For immunofluorescence analysis, five-\u00b5m kidney tissue sections were cut and incubated with antibodies raised against caspase-3  after the antigens were retrieved enzymatically using 0.05% trypsin solution (Sigma) for 10 min. Subsequent to incubation with fluorescein isothiocyanate (FITC)-conjugated secondary antibody  solution, the slides were visualized using an immunofluorescence microscope . ImageJ software (version 1.41) was utilized to semiquantitatively analyze the data. For normalization, pixel intensities obtained from the treatment groups were divided by the corresponding pixel intensities recorded for the sham mice.Western blottingSodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed to electrophoretically separate the proteins. After the transfer of the separated proteins onto the PVDF membranes via electro-blotting, blocking was performed using a 5% skimmed milk solution for 60 min. Then, the membranes were incubated in caspase-3 , phosphoinositide 3-kinase (PI3K) , phosphorylated Akt (p-Akt) , phosphorylated mammalian target of rapamycin (p-mTOR) , phosphorylated cAMP response element-binding protein 1 (p-CREB-1) , Sirtuin 1 (SIRT1) , and \u03b2-actin  primary antibodies at 4 \u00b0C overnight. Blots were then incubated by the horseradish peroxidase-labeled secondary antibody solution at room temperature for 45 min and were visualized by Western Blotting Luminol Reagent (Santa Cruz). ImageJ software (version 1.41) was implemented to measure the pixel intensities of the visualized bands from the X-ray films. The band density of each protein was first divided by the band density of its respective \u03b2-actin loading control and the obtained values for study groups were then normalized by the values for the sham group.Statistical analysispost hoc comparisons using Statistical Package for the Social Sciences, . The statistical significance level was set at\u00a0P<0.05.Data were descriptively expressed as mean \u00b1 SD. The variables were analyzed by one-way analysis of variance followed by Tukey\u2019s test for General characteristics of the study groups have been presented in P<0.01)]. TNF-\u03b1 levels in the kidneys were declined by 23.4% after treatment with 5 mg/kg B-LAP (P<0.05)  .P<0.01), and 5 mg/kg B-LAP significantly reduced renal MDA levels to 11.62 \u00b1 1.19 nmol/mg protein (P<0.05)  compared with sham mice (4.42 \u00b1 0.92 nmol/mg protein) ((P<0.05) .P<0.01); and 5 mg/kg B-LAP significantly elevated its values to 15.62 \u00b1 1.36 U/mg protein (P<0.05) in diabetic mice. It should be underlined that healthy mice receiving 5 mg/kg B-LAP had higher activities of GPX in their kidneys (7.46 \u00b1 1.12 U/mg protein) compared with sham mice (5.21 \u00b1 1.06 U/mg protein) (P<0.05)  were significantly higher than in sham mice (5.21 \u00b1 1.06 U/mg protein) ((P<0.05) .P<0.01)]. Treatment with 5 mg/kg B-LAP reduced urinary levels of nephrin by 23.56% compared with diabetic mice (P<0.05). Urinary NGAL levels were significantly increased in diabetic mice (0.64 \u00b1 0.073 ng/mg Cr) compared with the sham group (0.23 \u00b1 0.042 ng/mg Cr) (P<0.01); and 5 mg/kg B-LAP reduced urinary NGAL levels to 0.53 \u00b1 0.055 ng/mg Cr (P<0.05)  .P<0.01). Its values were increased to 1.44 \u00b1 0.18 mL/min/kg after treatment with 5 mg/kg B-LAP (P<0.05)  in comparison with sham mice (2.21 \u00b1 0.23 mL/min/kg) ((P<0.05) .P<0.01) and 5 mg/kg B-LAP slightly, albeit statistically significantly, attenuated its levels compared with control diabetic mice (P<0.05)  .P<0.01) (P<0.05)  . PI3K anP<0.01) ; on the P<0.01) . Convers(P<0.05) .The current study showed that B-LAP, a plant-derived\u00a0naphthoquinone\u00a0with potential anti-inflammatory activity, protected the kidneys against the deleterious effects of STZ-induced diabetes. In addition to improving renal function as demonstrated by elevated creatinine clearance values, B-LAP successfully declined renal and urinary levels of tubular injury markers, including nephrin and NGAL. Ameliorating renal histoarchitecture, B-LAP reduced renal levels of p-mTOR as well as p-CREB and at the same time increased SIRT1 levels in the kidneys of diabetic mice. +; thereby, restored cellular NAD+ activates NAD+-dependent enzymes, including SIRT1 deacetylase. Deacetylated p65 subunit of NF-\u03baB protein complex is prone to degradation by the ubiquitin-proteasome system (UPS); therefore, increased activity of SIRT1 suppresses inflammation by reducing the expression of cytokines that are downstream to NF-\u03baB (B-LAP functions as the activator of NAD(P)H quinone dehydrogenase (NQO1), the enzyme that regenerates cellular NADH into NADto NF-\u03baB , 16. B-Lto NF-\u03baB . In lineto NF-\u03baB .et al., Huang Kui capsule (HKC) alleviates glomerular and tubular pathological changes associated with DN and simultaneously represses renal levels of p-Akt and p-mTOR in mice , in addition to repressing renal levels of PI3K and p-AKT, down-regulates transforming growth factor-beta (TGF-\u03b2) expressions in the kidneys and ameliorates albuminuria in type 2 diabetic rats (Our findings showed that B-LAP especially at 5 mg/kg attenuated elevated levels of PI3K, p-Akt, p-mTOR, and p-CREB in the kidneys of diabetic mice. The induction of the PI3K/Akt/mTOR signaling pathway contributes significantly to the pathogenesis of DN as it initiates the pathways involved in kidney fibrosis and inflammation . In agre in mice . Likewistic rats .CREB is a transcription factor that after phosphorylation is translocated into the nuclei of the renal cells and functions as the regulator of the expression of genes encoding several pro-fibrotic  and antiDemonstration of reduced MDA levels and elevated GPX activities in the renal tissues by B-LAP treatment is an indication of ameliorations in oxidative stress, the indispensable feature of DN both in mice and in humans . LikewisGlomerular basement membrane disintegration, as well as podocyte detachment, are two principal pathogenic features of DN . 5 mg/kgOur findings underline the protective actions of B-LAP on the renal complications of STZ-induced diabetic mice. Apart from reductions in the urinary indices of glomerular and tubular injury, creatinine clearance values, the indicators of renal function, were significantly improved in diabetic mice by one-month B-LAP treatment. Moreover, 5 mg/kg B-LAP confers protection against renal inflammation and oxidative stress in STZ-induced diabetic mice probably via modulating the activities of PI3K/Akt/mTOR and SIRT1 levels in the kidneys."}
+{"text": "We examined the in vitro activity of CAZ-AVI and comparators against presumed community-acquired  and hospital-acquired  isolates collected from pediatric patients as part of the ATLAS surveillance program.Ceftazidime-avibactam (CAZ-AVI) is a \u03b2-lactam/non-\u03b2-lactam \u03b2-lactamase inhibitor combination with Escherichia coli, Klebsiella spp., Proteus mirabilis) or meropenem MICs \u22652 \u00b5g/mL  or \u22654 \u00b5g/mL (Psa) were screened for \u03b2-lactamase genes.6654 non-duplicate isolates were collected in 52 countries in Europe (n=3423), Latin America (n=1323), Middle East/Africa (n=1177), and Asia/Pacific  from patients (newborn to 17 y) with lower respiratory tract , urinary tract , bloodstream , skin and soft tissue , and intra-abdominal  infections. Susceptibility testing was performed by CLSI broth microdilution and values were interpreted using CLSI 2021 breakpoints. CAZ-AVI was tested at a fixed concentration of 4 \u00b5g/mL AVI. Isolates with CAZ or aztreonam MICs \u22652 \u00b5g/mL (in vitro activity of CAZ-AVI exceeded that of meropenem and other tested \u03b2-lactams against Ent (97.8% susceptible (S)) and Psa (92.1% S) collected globally from pediatric patients (Table). Percentages of susceptibility to CAZ-AVI ranged from 95.4-99.2% among CA Ent from different infection types and were reduced 0.6-1.3% among HA isolates from LRTI, UTI, SSTI, and IAI. Susceptibility to CAZ-AVI was also similar (92.6-95.8% S) among CA Psa from different infection types and was reduced 1.2-7.0% among HA isolates. Larger differences in susceptibility were typically seen for the tested comparator \u03b2-lactams. For Ent, the lowest percentages of susceptibility to the tested \u03b2-lactams were observed among isolates from BSI, while the pattern was less clear for Psa.The Results TablePsa in pediatric patients.CAZ-AVI could provide a valuable therapeutic option for treatment of CA and HA infections caused by Ent and Krystyna Kazmierczak, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Sibylle Lob, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Gregory Stone, PhD, AztraZeneca Pfizer, Inc. (Employee) Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "Two severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants associated with increased transmission and immune evasion, P.1 and P.2, emerged in Brazil and spread throughout South America. Here, we report genomes corresponding to these variants that were recently detected in Uruguay. These P.1 and P.2 genomes share all substitutions that are characteristic of these variants. Betacoronavirus (family Coronaviridae) and the causative agent of the ongoing coronavirus disease 2019 (COVID-19) pandemic (Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel member of the genus pandemic . VOCs hapandemic , 5. Thespandemic , and theThe research described in this study was performed in adherence to the Declaration of Helsinki; no specific authorization was required, because the activities were conducted as part of a routine virological surveillance  by the Uruguayan official Institution for Surveillance of Influenza and Other Respiratory Viruses of the Ministry of Public Health (DLSP-MSP).The variants P.1 and P.2, which emerged in Brazil, have spread to other parts of South America , 8. HereTC) value of\u2009<18. RNA was extracted with a QIAmp viral minikit . Genome amplification was achieved using ARTIC 3 primers (https://artic.network/ncov-2019). First, cDNA strand analysis, Nextera DNA Flex library preparation, and 2\u2009\u00d7\u2009150-bp sequencing on an Illumina MiniSeq platform were performed following a previous report (http://cov-glue.cvr.gla.ac.uk/). Lineages refer to those assigned using the pangolin tool (https://cov-lineages.org). All tools were run with default parameters unless otherwise specified.Nasopharyngeal swab samples were collected in March 2021 in the Uruguayan Rivera department bordering Brazil and came from two symptomatic cases. The samples tested positive for SARS-CoV-2 using a standard quantitative PCR (qPCR) procedure ; both pas report . AdapterSample SARS-CoV-2/human/URY/374/2021 (P.1) has a sequence length of 29,835 nucleotides (nt), 1,240,211 total reads, 3,920\u00d7 mean coverage, and a 38.0% G+C content. Sample SARS-CoV-2/human/URY/380/2021 (P.2) has a sequence length of 29,858 nt, 1,007,202 total reads, 5,877\u00d7 mean coverage, and a 37.9% G+C content. Their genome sequences lack the outermost nucleotides (<20 nt) of the 5\u2032 and 3\u2032 untranslated regions (UTRs), which are not usually sequenced with the ARTIC protocol.nsp6 that is considered a P.1 genetic signature  and MW988205 . The raw reads and metadata were deposited under the BioProject accession number PRJNA634396 and SRA accession numbers SRX10652818 (SARS-CoV-2/human/URY/374/2021) and SRX10652819 (SARS-CoV-2/human/URY/380/2021).These genome sequences were deposited in GenBank under accession numbers"}
+{"text": "Dear Editor,TNFRSF8, is a transmembrane cytokine receptor of the tumor necrosis factor receptor (TNFR) superfamily and expressed on ~20% of DLBCL. To better understand the underlying mechanism of CD30 expression in DLBCL, we performed clinical, genomic and transcriptomic analysis in a cohort of 1048 patients with de novo DLBCL. A flow chart describing the cohort selection was displayed in Supplementary Fig. P\u2009<\u20090.001, Supplementary Fig. P\u2009=\u20090.002), as compared to CD30-DLBCL. In terms of IPI, 39% of CD30\u2009+\u2009DLBCL and 48% of CD30-DLBCL were categorized into low-risk group, 21% and 19% into low-intermediate risk group, 25% and 17% into intermediate-high risk group, and 16% and 16% into high-risk group, respectively (P\u2009=\u20090.040). Pathologically, CD30\u2009+\u2009DLBCL was significantly associated with increased percentage of non-GCB subtype (P\u2009=\u20090.042) and EBER positivity (P\u2009<\u20090.001), as previously reported [Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma with heterogeneous clinical, immunophenotypic, and genetic features. Surface antigens, such as CD20, CD22, CD79B, and CD19 are important accessible parts of lymphoma cells with immunotherapeutic potential. CD30, encoded by TNFAIP3 (P\u2009=\u20090.002), TNFRSF14 (P\u2009=\u20090.045), SOCS1 (P\u2009=\u20090.010), STAT6 (P\u2009=\u20090.007), CIITA (P\u2009=\u20090.007), CD58 (P\u2009=\u20090.037), KMT2C (P\u2009=\u20090.016), but lower mutation frequency in CD79B (P\u2009=\u20090.026) and MYD88 , cytotoxic T-cells (P\u2009=\u20090.030), exhausted T-cells (P\u2009=\u20090.004), T-helper 17 (Th17) cells (P\u2009=\u20090.001), regulatory T (Treg)-cells (P\u2009=\u20090.001), dendritic cells , and macrophages , with worst prognosis in JA subtype  and follicular helper T (Tfh) cells (P\u2009=\u20090.001), with a positive correlation between the signature genes for immune cell recruitment such as CCR6, CCL21, and CXCR4 , in parallel with immune checkpoint genes, such as PD-L1, TIM3, and LAG3  and CD8\u2009+\u2009T-cells  and CIITA (CIITAkd), were transfected into OCI-LY10. Meanwhile, SOCS1wt, SOCS1Q175H, CIITAwt, CIITAL807R, as well as a shRNA to knockdown TNFAIP3 (TNFAIP3kd), were transfected into SU-DHL-4. Quantitative real-time PCR and western blot were used to confirm the transfection efficiency , an anti-CD30 monoclonal antibody-drug conjugate has shown remarkable responses in first-line treatment of HL and CD30\u2009+\u2009peripheral T-cell lymphoma in combination with chemotherapy [TNFAIP3L147Q versus TNFAIP3wt OCI-LY10 , and TNFAIP3kd versus scramble SU-DHL-4 , and SOCS1Q175H versus SOCS1wt SU-DHL-4 , and CIITAL807R versus CIITAwt SU-DHL-4 . There are currently no plans to share data not included in this paper.Sequencing data is available at the National Omics Data Encyclopedia (NODE, Supplemental Materials"}
+{"text": "Drug-resistant tuberculosis (DR-TB), obesity, and malnutrition are growing public health problems in the world. However, little has discussed the impact of different BMI status on the emergence of TB drug resistance. We aimed to explore the drug-resistant profiles of DR-TB and its clinical predictors among underweight, overweight or obesity population.8957 newly diagnosed TB cases with drug susceptibility results and BMI data in Shandong China, from 2004 to 2019 were enrolled. Multivariable and univariable logistic regression models were applied to investigate the impact of BMI on different drug-resistance. Clinical predicators and drug-resistant profiles of DR-TB among obesity, underweight, normal TB group were also described.P\u2009<\u20090.05.Among 8957\u00a0TB cases, 6417 (71.64%) were normal weight, 2121 (23.68%) were underweight, 373 (4.16%) were overweight, and 46 (0.51%) were obese. The proportion of drug resistance and co-morbidity among normal weight, underweight, overweight, obese TB groups were 18.86%/18.25%/20.38%/23.91% (DR-TB), 11.19%/11.74%/9.65%/17.39% , 3.41%/3.06%/5.36%/0.00% , 4.21%/3.39%/5.36%/6.52% , 10.57%/8.44%/19.57%/23.91% (co-morbidity), respectively. Compared with normal weight group, underweight were associated with lower risk of streptomycin-related resistance , but contributed to a higher risk of MR-TB (isoniazid) (odds ratio (OR) 1.347, 95% CI 1.049\u20131.730; adjusted OR (aOR) 1.31, 95% CI 1.017\u20131.686), P\u2009<\u20090.05. In addition, overweight were positively associated with MDR-TB , isoniazid\u2009+\u2009rifampicin\u2009+\u2009streptomycin resistance : 1.061\u20133.577; aOR 2.113, 95% CI 1.141\u20133.912), Any isoniazid\u2009+\u2009streptomycin resistance , The higher risk of MDR-TB, isoniazid\u2009+\u2009rifampicin\u2009+\u2009streptomycin resistance, Any isoniazid\u2009+\u2009streptomycin resistance, and co-morbidity among overweight population implies that routine screening for drug sensitivity and more attention on co-morbidity among overweight TB cases may be necessary. In addition, underweight TB cases have a higher risk of isoniazid resistance. Our study suggests that an in-depth study of the interaction between host metabolic activity and infection of DR-TB may contribute more to novel treatment options or preventive measures, and accelerate the implementation of the STOP TB strategy. Mycobacterium tuberculosis (MTB), contributes to a largest number of deaths from infectious diseases (more than HIV/AIDS) [Tuberculosis (TB), is caused by IV/AIDS) , 2. AccoIV/AIDS) . AdditioIV/AIDS) . Nine peIV/AIDS) .Multidrug-resistant tuberculosis (MDR-TB) is defined as TB with resistance to at least isoniazid (INH) and rifampin (RFP), which is usually associated with longer hospitalization, more expensive treatment, and higher mortality . The bac2 was independent risk factor of latent tuberculosis infection  1.17, 95% CI 1.04\u20131.33) [The critical influences of diverse nutritional status on TB infection have been recognized for decades . Recent 04\u20131.33) . In summThis research intended to explore the association between BMI and primary DR-TB in the aspects as follows: (1) to describe the clinical characteristics of TB cases with four different BMI status ; (2) to illustrate the drug resistant profiles of TB cases subgroups stratified by BMI; (3) to analyze the relative risk of DR-TB including DR-TB , MDR-TB , mono-resistant tuberculosis , polydrug resistant tuberculosis , RFP-related resistance, INH-related resistance, streptomycin (SM)-related resistance, MR-TB (INH), INH\u2009+\u2009RFP\u2009+\u2009SM resistance (MDR3), INH\u2009+\u2009SM resistance (PDR2), Any INH\u2009+\u2009SM resistance among subgroups with different BMI; (4) to investigate the risk factors of DR-TB among TB cases subgroups stratified by BMI.The Ethics Committee of Shandong Provincial Hospital (SPH) and Shandong Provincial Chest Hospital (SPCH) approved for our study. Personal information of TB patients such as names were erased before data analysis and reporting. All methods were performed in accordance with relevant guidelines and regulations. Informed consent was obtained from all participants or, if participants are under 18, from a parent and/or legal guardian.2 [2) was 3.5% (95% CI 3.4\u20133.6) [Our study was conducted in Shandong, a coastal province in eastern China (36\u00b024\u2032N latitude and 118\u00b0 24\u2032 E longitude), with 100 million inhabitants and an area of 157,100\u00a0km2 . A large2 . The rat2 . As estiNontuberculosis mycobacteria (NTM) infection; (3) BMI information missing; (4) extra-pulmonary cases , meanwhile their basic demography and clinical characteristics were collected. Information of BMI, age, sex , drinking (yes or no), smoking (yes or no), cavity (yes or no), and co-morbidity were collected through questionnaire. Nutritional status indicators except BMI were not routinely collected. Exclusion criteria: (1) cases with previous TB history; (2) 2 were in the underweight range, between 18.5 and 24.9\u00a0kg/m2 were in normal weight range, between 25 and 29.9\u00a0kg/m2 were in the overweight range,\u2009\u2265\u200930\u00a0kg/m2 were in the obese range [BMI below 18.5\u00a0kg/mse range . Mono-rese range . Multidrse range . Polydruse range . MR-TB  of SPCH. Initially, at least two sputum specimens of each eligible patient were collected. Then, isolates were cultured in L\u00f6wenstein-Jensen (L-J) medium according to the standard protocol, and then these growing colonies were used for strain identification and phenotypic DST . DST forP value\u2009<\u20090.05. All data analyses were conducted in SPSS software (version 20.0).Categorical baseline demographic and clinical features of 8957\u00a0TB cases including age , sex (men or women), smoker or non-smoker, drinker or non-drinker, cavity (yes/no), co-morbidity (yes/no), and drug resistant profiles in four subgroups with different BMI status were compared by Pearson Chi-square test or Fisher\u2019s exact test. The age subgroups were divided according to previous publications , 22. BinAs shown in Table P\u2009<\u20090.05. Underweight TB cases had a higher proportion of asthma (0.85% vs 0.37%) and COPD (2.55% vs 1.78%) but with a lower rate of diabetes (3.58% vs 7.03%) than normal group, P\u2009<\u20090.05. There were no significant differences in sex, cavity, smoking and drinking between lower and normal BMI cases. Both these two groups have more males, but less drinkers, smokers, and cavity  or\u2009>\u200965 (34.64% vs 22.98%) age group, and they had a lower rate of co-morbidity (8.44% vs 10.57%), P\u2009<\u20090.05. In addition, overweight TB cases are less likely to be in 15\u201324 age group (6.18% vs 15.63%), P\u2009<\u20090.05. There were no significant differences in 0\u201314 age group, 65\u2009+\u2009age group, sex, smoking and drinking between overweight and normal cases (Table The proportions of 25\u201344 age group (33.33% vs 15.63%), 45\u201364 age group (39.78% vs 34.21%), cavity (51% vs 43.66%), co-morbidity (19.57% vs 10.57%), COPD (3.22% vs 1.78%), diabetes (13.94% vs 7.03%), hypertension (3.75% vs 1.82%), and cancer (0.80% vs 0.16%) were higher in The rates of females (32.61% vs 17.19%), co-morbidity (23.91% vs 10.57%), diabetes (19.57% vs 7.03%) were higher in obese TB cases than in normal cases, P\u2009<\u20090.05. There were no significant differences in age, smoking, drinking and cavity between obese and normal cases  or MDR3 (INH\u2009+\u2009RFP\u2009+\u2009SM) (3.22% vs 1.68%) cases among overweight group than normal group, P\u2009<\u20090.05. There were no significant differences in other drug resistant sub-types between abnormal weight groups and normal group.As shown in Table P\u2009=\u20090.028), but contributed to a higher risk of MR-TB (INH) . In addition, overweight was positively associated with MDR-TB , INH\u2009+\u2009RFP\u2009+\u2009SM resistance , Any INH\u2009+\u2009SM resistance . However, there were no statistical differences on drug resistance between obese group and normal weight group. In univariable and multivariable multinomial logistic regression models, we also found that overweight may be a risk factor for MDR-TB   and overweight cases . In addition, co-morbidity except COPD, diabetes, hypertension  were positively associated with DR-TB in underweight cases. Among normal weight group, male  or cavitary  pulmonary tuberculosis were more likely to have anti-TB resistance   and overweight (BMI\u2009=\u200924\u201326.9\u00a0kg/m2)  contributed to a lower risk of TB infection [2 was observed to be independently associated with LTBI  [Obesity has become a global public health problem, and it\u2019s reported that approximately 46% of adults and 15% of children China are overweight or obese . Plenty 04\u20131.33) . Interes04\u20131.33) , 26. Fur04\u20131.33) . Therefo04\u20131.33) , 30. The04\u20131.33) , and theOverweight TB cases had a higher possibility of suffering from co-morbidity including COPD, hypertension, diabetes, and cancer. Actually, previous studies have figured out that BMI was a reliable predictor of prevalent diabetes, hypertension, and COPD \u201333. A stPeople have found a significant association between underweight and higher risk of active TB, adverse TB treatment outcomes, tuberculosis-related mortality , 37\u201339. It has been observed that males were more likely to be affected with DR-TB among normal, underweight, or overweight population. However, there were inconsistent results in the previous literature about the influence of gender differences on vulnerability to MDR-TB \u201344. A stThis study had some strengths. Firstly, it\u2019s the first study to investigate the effect of BMI on drug resistance among newly diagnosed pulmonary TB cases. Secondly, the relative risk of numerous drug-resistant sub-types including DR-TB , MDR-TB , MR-TB , PDR-TB , RFP-related resistance, INH-related resistance, SM-related resistance, IMR-TB (INH), INH\u2009+\u2009RFP\u2009+\u2009SM resistance (MDR3), INH\u2009+\u2009SM resistance (PDR2), Any INH\u2009+\u2009SM resistance among underweight, overweight, obese were analyzed, and could provide clinical reference for future pathologic and molecular studies on different resistant MTB strains. Thirdly, the large amount (DST results from a province of 100 million people) and long time span (from 2004 to 2019) of our data guaranteed the reliance of our findings.This study also had some limitations. Firstly, drug resistance of second-line anti-TB drugs were not routinely examined in China unless at the initiative requirements of patients. Thus, the association between BMI and second-line anti-TB resistance remains to be discovered in future. Secondly, this was a retrospective study, and may have information bias. Thirdly, some obese patients can also be malnourished, since BMI is only one of prognostic factors for assessing malnutrition , 29, 45.Our study has important implications on the global triple epidemics of underweight, obesity, and DR-TB. The positive effect of overweight on MDR-TB, INH\u2009+\u2009RFP\u2009+\u2009SM resistance, Any INH\u2009+\u2009SM resistance, and co-morbidity implies that routine screening for drug sensitivity and more attention on co-morbidity among overweight TB cases may be necessary. Although patients with normal weight had a lower proportion of drug-resistance than overweight or obese patients in Shandong, screening for drug resistance cannot be ignored either because patients with normal weight accounted for 74.64%. In addition, we found underweight TB cases have a higher risk of INH resistance, which provides a reference for clinical rational use of drugs. Our study suggests that an in-depth study of the interaction between host metabolic activity and infection of DR-TB may contribute more to novel treatment options or preventive measures, and accelerate the implementation of the STOP TB strategy."}
+{"text": "In December 2020, B.1.1.7 lineage of SARS-CoV-2 was first detected in the United States and has since become the dominant lineage. Previous investigations involving B.1.1.7 suggested higher rates of transmission relative to non-B.1.1.7 lineages. We conducted a household transmission investigation to determine the secondary infection rates (SIR) of B.1.1.7 and non-B.1.1.7 SARS-CoV-2 lineages. From January\u2013April 2021, we enrolled members of households in San Diego County, CA, and Denver, CO metropolitan area (Tri-County), with a confirmed SARS-CoV-2 infection in a household member with illness onset date in the previous 10 days. CDC investigators visited households at enrollment and 14 days later at closeout to obtain demographic and clinical data and nasopharyngeal (NP) samples on all consenting household members. Interim visits, with collection of NP swabs, occurred if a participant became symptomatic during follow-up. NP samples were tested for SARS-CoV-2 using TaqPath\u2122 RT-PCR test, where failure to amplify the spike protein results in S-Gene target failure (SGTF) may indicate B.1.1.7 lineage. Demographic characteristics and SIR were compared among SGTF and non-SGTF households using two-sided p-values with chi-square tests; 95% confidence intervals (CI) were calculated with Wilson score intervals.552 persons from 151 households were enrolled. 91 (60%) households were classified as SGTF, 57 (38%) non-SGTF, and 3 (2%) indeterminant. SGTF and non-SGTF households had similar sex distribution  and age  and 31 years (IQR 15\u201345), respectively). Hispanic people accounted for 24% and 32% of enrolled members of SGTF and non-SGTF households, respectively (p=0.04). At least one secondary case occurred in 61% of SGTF and 58% of non-SGTF households (P=0.66). SIR was 52% (95%[CI] 46%-59%) for SGTF and 45% (95% CI 37%-53%) for non-SGTF households (P=0.18). SIRs were high in both SGTF and non-SGTF households; our findings did not support an increase in SIR for SGTF relative to non-SGTF households in this setting. Sequence confirmed SARS-CoV-2 samples will provide further information on lineage specific SIRs.All Authors: No reported disclosures"}
+{"text": "The elevated plus maze test was employed to evaluate anxiety-like behavior in rats, and in vivo microdialysis was used to measure the extracellular NE level in the BNST. In elevated plus maze tests, EtOHW rats but not EtOH-naive rats exhibited anxiety-like behavior when challenged with 7-minute mild restraint stress, which was, respectively, mitigated by prior intra-NTS infusion of the nitric oxide scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO), nonselective NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME), or selective neuronal NOS (nNOS) inhibitor 7-nitroindazole (7-NI). Each of these agents also decreased the plasma corticosterone levels in EtOHW rats. In in vivo microdialysis, prior intra-NTS infusion of carboxy-PTIO, L-NAME, or 7-NI attenuated the mild stress-induced NE release in the BNST of EtOHW rats. Additionally, EtOHW rats showed increased solitary nNOS gene and protein expression. Moreover, the anxiolytic effect of intra-NTS administration of 7-NI was abolished by subsequent intra-NTS administration of sodium nitroprusside. These results suggest that elevation of solitary nitric oxide signaling derived from nNOS mediates stress-precipitated anxiety and norepinephrine release in the BNST during protracted EtOHW.Ethanol withdrawal (EtOHW) alters the pattern of neurohormonal and behavioral response toward internal and external stimuli, which mediates relapse to alcohol use even after a long period of abstinence. Increased noradrenergic signaling from the nucleus tractus solitarius (NTS) to the bed nucleus of the stria terminalis (BNST) during EtOHW underlies withdrawal-induced anxiety, while nitric oxide synthase (NOS) inhibitors injected into the periaqueductal area attenuate EtOHW-induced anxiety. Therefore, this study investigated the involvement of NOS within the NTS in anxiety and increased norepinephrine (NE) release in the BNST during protracted EtOHW in rats exposed to a mild stress. Rats were intraperitoneally administered 3\u2009g/kg/day EtOH for 21 days followed by 28 days of withdrawal, and on the 28 Relapse is a major barrier in alcoholism therapy . EthanolElevated anxiety during EtOHW is the major negative emotional component for alcoholism relapse , and sev\u03b2-adrenergic antagonists reduced the withdrawal-associated conditioned place aversion . However, as also seen in Figures into open arms: F = 9.22, p < 0.001, EtOH/vehicle/non-AMRS group (n = 8) versus EtOH/vehicle/AMRS group (n = 8), p < 0.001, EtOH/vehicle/AMRS group versus EtOH/carboxy-PTIO/AMRS group (n = 8), p < 0.01; EtOH/vehicle/AMRS group versus EtOH/L-NAME/AMRS group (n = 8), p < 0.01; EtOH/vehicle/AMRS group versus EtOH/7-NI/AMRS group (n = 8), p < 0.01; EtOH/vehicle/AMRS group versus EtOH/L-NIO/AMRS group (n = 8), p > 0.05; %timespent in open arms: F = 8.05, p < 0.001, EtOH/vehicle/non-AMRS group versus EtOH/vehicle/AMRS group, p < 0.001, EtOH/vehicle/AMRS group versus EtOH/carboxy-PTIO/AMRS group, p < 0.01; EtOH/vehicle/AMRS group versus EtOH/L-NAME/AMRS group, p < 0.01; EtOH/vehicle/AMRS group versus EtOH/7-NI/AMRS group, p < 0.01; EtOH/vehicle/AMRS group versus EtOH/L-NIO/AMRS group, p > 0.05]. Moreover, at the same dose, intra-NTS 7-NI alone did not produce any significant behavioral changes in rats (data not shown).As shown in Figures F(drug) = 19.92, p < 0.001, F(stress) = 32.25, p\u2009<\u20090.001, F(drug \u00d7 stress) = 13.96, p < 0.01; saline/non-AMRS group (n = 6) versus EtOH/AMRS group (n = 6), p < 0.001; EtOH/non-AMRS group (n = 6) versus EtOH/AMRS group, p < 0.001; saline/AMRS group versus EtOH/AMRS group, p < 0.001]. However, as shown in F = 11.71, p < 0.001; EtOH/vehicle/non-AMRS group (n = 6) versus EtOH/vehicle/AMRS group (n = 6), p < 0.001, EtOH/vehicle/AMRS group versus EtOH/carboxy-PTIO/AMRS group (n = 6), p < 0.01], L-NAME , or 7-NI , but not L-NIO , inhibited the increased CORT secretion.Consistent with the results of the behavioral, as seen in n = 7): 4.25 \u00b1 0.32 (pg/15 uL); saline/vehicle/AMRS group (n = 7): 4.75 \u00b1 0.48; EtOH/vehicle/non-AMRS group (n = 7): 4.64 \u00b1 0.41; EtOH/vehicle/AMRS group (n = 7): 5.19 \u00b1 0.44; EtOH/carboxy-PTIO/AMRS group (n = 7): 4.38 \u00b1 0.37; EtOH/L-NAME/AMRS group (n = 7): 4.23 \u00b1 0.38; EtOH/7-NI/AMRS group (n = 7): 5.25 \u00b1 0.37; EtOH/L-NIO/AMRS group (n = 7): 4.81 \u00b1 0.37]. As seen in Figures F(treatment) = 0.07, p > 0.05, F(time) = 2.25, p > 0.05, F(treatment \u00d7 time) = 0.70, p > 0.05; in F(treatment) = 16.79, p < 0.001; F(time) = 25.63, p < 0.001, F(treatment \u00d7 time) = 9.55, p < 0.001; 15\u2009min: saline/vehicle/AMRS group versus EtOH/vehicle/AMRS group, p < 0.001; 30\u2009min saline/vehicle/AMRS group versus EtOH/vehicle/AMRS group, p < 0.05]. However, as shown in F(treatment) = 9.62, p < 0.001, F(time) = 87.92, p < 0.001, F(treatment \u00d7 time) = 4.53, p < 0.001; 15\u2009min: EtOH/vehicle/non-AMRS group EtOH/vehicle/AMRS group, p < 0.001, EtOH/vehicle/AMRS versus EtOH/carboxy-PTIO/AMRS group, p < 0.001; 30\u2009min: EtOH/vehicle/non-AMRS group versus EtOH/vehicle/AMRS group, p < 0.01, EtOH/vehicle/AMRS group versus EtOH/carboxy-PTIO/AMRS group, p < 0.01], L-NAME , or 7-NI , but not L-NIO , prevented these increases. In addition, intra-NTS L-NIO alone did not significantly affect NE release in naive rats (data not shown).No significant differences in the basal extracellular NE level in the vBNST were found between the groups , whereas the AMRS did not alter nNOS protein expression . However, the AMRS markedly enhanced phospho-nNOS levels in EtOHW rats  but not in saline-treated controls . Additionally, neither EtOHW nor AMRS significantly affected eNOS or phospho-eNOS protein expressions .As shown in F(drug) = 84.40, p < 0.001, F(stress) = 0.04, p > 0.05, F(drug\u00d7stress)\u2009=\u20090.06, p > 0.05; saline/non-AMRS group (n = 6) versus EtOH/non-AMRS group (n = 6), p < 0.001; saline/AMRS group (n = 6) versus EtOH/AMRS group (n = 6), p < 0.001]. The AMRS did not alter nNOS mRNA expression .In agreement with the Western blot data, as seen in into open arms: F = 15.87, p < 0.001; saline/vehicle/AMRS/vehicle group (n = 6) versus EtOH/vehicle/AMRS/vehicle group (n = 6), p < 0.001; EtOH/vehicle/AMRS/vehicle group versus EtOH/7-NI/AMRS/vehicle group (n = 6), p < 0.001; EtOH/7-NI/AMRS/vehicle group versus EtOH/7-NI/AMRS/SNP group (n = 6), p < 0.01; saline/vehicle/AMRS/vehicle group versus EtOH/7-NI/AMRS/SNP group, p < 0.001; %timespent in open arms: F = 11.17, p < 0.001; saline/vehicle/AMRS/vehicle group versus EtOH/vehicle/AMRS/vehicle group, p < 0.001; EtOH/vehicle/AMRS/vehicle group versus EtOH/7-NI/AMRS/vehicle group, p < 0.01; EtOH/7-NI/AMRS/vehicle group versus EtOH/7-NI/AMRS/SNP group, p < 0.05; saline/vehicle/AMRS/vehicle group versus EtOH/7-NI/AMRS/SNP group, p < 0.01].To further determine the involvement of solitary nNOS in AMRS-induced anxiety during protracted EtOHW, another cohort of EtOHW rats was sequentially treated with 7-NI and SNP and then tested in the EPM . As showThe results of the present study showed that 7-minute AMRS provoked anxiety-like behaviors, enhanced plasma CORT secretion, and sensitized NE release in the vBNST in rats treated with EtOH but not saline, at 28 days after the final dose of EtOH or saline. However, all of these behavioral, hormonal, and neurochemical abnormalities were attenuated by prior intra-NTS infusion of carboxy-PTIO, L-NAME, or 7-NI, but not by L-NIO. EtOHW elevated nNOS, but not eNOS, protein expression in the NTS, concomitant with an increased nNOS mRNA level, and the AMRS increased the phosphorylation rate of nNOS in the NTS of EtOHW rats. Moreover, intra-NTS injection of SNP after 7-NI administration abolished the expected anxiolytic action of 7-NI. Taken together, these results suggest a critical role of solitary nNOS in anxiety and vBNST NE release induced by acute mild stress during protracted EtOHW.The susceptibility to stress during protracted EtOHW alters neurotransmission responses to certain stimuli that are normally innocuous to provoke pathophysiological consequences , 27. To Both anxiety and plasma CORT secretion are closely associated with increased noradrenergic transmission in the BNST , 33. In The NTS primarily integrates and transmits visceral and external information to the forebrain, forming the autonomic-affective functional basis for the body. The gaseous molecule NO serves as both a neurotransmitter and neuromodulator, and is synthesized by three isoforms of NOS, i.e., nNOS, iNOS, and eNOS. We reported previously that systemic nicotine administration increased hypothalamic NE release via activation of both nNOS and eNOS in the NTS . HoweverIn summary, prior intra-NTS infusion of carboxy-PTIO, L-NAME, or 7-NI, but not L-NIO, attenuated the anxiety and vBNST NE release induced by 7-minute AMRS during EtOHW. EtOHW enhanced both nNOS protein and mRNA expression in the NTS but did not affect the eNOS protein level. These observations suggest that nNOS activity is promoted in the NTS during protracted EtOHW, which sensitizes the NTS-BNST noradrenergic response to stress and results in anxiety-like behavior in rats ."}
+{"text": "Long term sequelae across multiple medical domains, including the respiratory, psychiatric, and neurocognitive have been reported after COVID-19. Studies evaluating the impact of this symptom burden, however, are lacking. We aimed to describe the self-reported occurrence of symptoms and their effect on patient functioning six months after their acute hospitalization for COVID-19. From a historical cohort study of patients hospitalized for COVID-19 between March 8, and June 14, 2020, we identified patients discharged home. The purpose of the study was explained, and they were asked to consent to a telephone questionnaire. We used a modified version of a previously validated general symptom questionnaire (GSQ-30) to assess multi-system symptom burden. The Patient Health Questionnaire-2 (PHQ-2) was used to screen for major depression. Of the original 565 patients, 258 patients were discharged home (45%). Of these, 57 (22%) patients were able to be contacted and agreed to participate in the survey. The mean (SD) age of the respondents was 55.1 (14.8) years, and 37 (64.9%) were female. The most common symptoms at follow-up were fatigue (60.0%), dyspnea (57.1%), feeling irritable, sad or decreased pleasure (56.4%), and memory difficulty (56.4%). Females had a significantly higher GSQ score (0.02) than males. Patients ages < 60 years tended to experience similar, if not greater, impaired functioning (p=0.07) compared with those ages \u2265 60 years (Table 1). Females were more likely to be irritable or sad (p=0.007), not feel rested on awakening (p=0.04), have shooting, stabbing and burning pain (p=0.02), have discomfort with normal light and sound (p=0.04), and have memory difficulty (p=0.04) than males (Table 2). Table 1. Self-Reported Post-Acute Sequelae of COVID syndrome in adults younger than 60 versus adults at or older than 60 Years. SD: Standard deviation, ICU: Intensive care unit, ED: Emergency department, GSQ - General symptom questionnaire, PHQ-2: Patient Health Questionnaire-2Table 2. Self-Reported Post-acute Sequelae of COVID syndrome in female versus male adults. SD: Standard deviation, ED: Emergency department, GSQ - General symptom questionnaire, PHQ-2: Patient Health Questionnaire-2Our study describes the clinical burden of post-acute sequelae of COVID-19 (PASC) in four core domains: fatigue, neurologic, neuro-psychiatric and viral-like symptoms. Over 45% of patients ages < 60 years suffered impaired functioning, compared with 21.1% of patient\u2019s ages 60 years and above. Females had significantly higher GSQ scores than men which strongly corelates with the functional impairment among the females. Larger studies are needed to further validate our findings.All Authors: No reported disclosures"}
+{"text": "Caseins are major milk proteins that have an evolutionarily conserved role in nutrition. Sequence variations in thecasein genes affect milk composition in livestock species. Regulatory elements of the casein genes could be used to directthe expression of desired transgenes into the milk of transgenic animals. Dozens of casein alleles have been identified forgoats, cows, sheep, camels and horses, and these sequence variants are associated with altered gene expression and milkprotein content. Most of the known mutations affecting casein genes\u2019 expression are located in the promoter and 3\u2019-untranslated regions. We performed pronuclear microinjections with Cas9 mRNA and sgRNA against the first coding exon ofthe mouse Csn1s1 gene to introduce random mutations in the \u03b1-casein (Csn1s1) signal peptide sequence at the beginningof the mouse gene. Sanger sequencing of the founder mice identified 40 mutations. As expected, mutations clusteredaround the sgRNA cut site (3 bp from PAM). Most of the mutations represented small deletions (1\u201310 bp), but we detectedseveral larger deletions as well (100\u2013300 bp). Functionally most mutations led to gene knockout due to a frameshift or astart codon loss. Some of the mutations represented in-frame indels in the first coding exon. Of these, we describe a novelhypomorphic Csn1s1 (Csn1s1c.4-5insTCC) allele. We measured Csn1s1 protein levels and confirmed that the mutation has anegative effect on milk composition, which shows a 50 % reduction in gene expression and a 40\u201380 % decrease in Csn1s1protein amount, compared to the wild-type allele. We assumed that mutation affected transcript stability or splicing by anunknown mechanism. This mutation can potentially serve as a genetic marker for low Csn1s1 expression. Caseins are major milk proteins that have an evolutionaryconserved role in nutrition . Casein locushas been studied for a long time to understand the principleof gene regulation and hormone-induced expression . At thesame time, regulatory elements of the casein genes could beused to direct the expression of desired transgenes into milkof transgenic animals .This strategy is frequently employed to create \u201cenriched\u201dmilk with improved composition , or to achieve large scale productionof recombinant human proteins in mouse models  and in livestockspecies . Milk-specific signal peptidesare used in biotechnology to enhance recombinant proteinsecretion during lactation . During breeding, casein genes acquired many sequencevariations that can lead to altered gene expression and arecharacteristic of some goat and cow breeds . Many of thesesequence variants could be used as markers for breeding. Indairy industry casein composition is an important milk traitdirectly influencing quality of dairy products . Hypomorphic casein mutationare also associated with other traits such as litter size . For example, K. Wang with colleagues showedthat 11 bp del in the intron 8 of the goat Csn1s1 negativelyaffects the expression of the gene . Otherknown hypomorphic mutations are located in the promoterand 3\u2032-untranslated regions (UTRs) of casein genes . Novel CRISPR methods greatly facilitate genome editing infarm animals , including targeted transgeneintegration  and mutation modeling . The latter approach has potential toexplain the molecular mechanism of how hypomorphic mutations affect milk proteins. In this report, we used CRISPR/Cas9to create a set of mutations within a signal peptide sequenceof the \u03b1-casein (Csn1s1) gene in mice. One of the mutantswas chosen to study effects of a small in-frame insertion onthe Csn1s1 expression during lactation.Generation and genotyping of the Csn1s1 mutant mice.In vitro transcription and purification of the gRNA wereperformed with MEGAshortscript\u2122 T7 Transcription Kit(Thermo Fisher Scientific) and MEGAclear\u2122 TranscriptionClean-Up Kit (Thermo Fisher Scientific) according to themanufacturer\u2019s protocol. Cas9 mRNA (GeneArt\u2122 CRISPRNuclease mRNA) was purchased from Thermo Fisher Scientific. 50 ng/\u03bcL Cas9 mRNA and 25 ng/\u03bcL gRNA (5\u2032-GTGAGGATGAGGAGTTTCA-3\u2032) were mixed in RNase-free water,backfilled into an injection needle with positive balancingpressure  and injected into thecytoplasm of zygotes (C57BL/6 \u00d7 CBA background). Afterinjections, the embryos were cultured for 1 hour in drops ofM16 medium at 37 \u00b0C and an atmosphere of 5 % CO2. Theviable microinjected zygotes were transplanted the same day into oviducts of pseudopregnant CD-1 females (0.5 days aftercoitus). Isoflurane inhalation anesthesia was applied in theseexperiments.1.Primers for PCR were as follows: 5\u2032-GCGCATAACTAAGCATCTTATGCT-3\u2032 (forward primer), 5\u2032-TGACTTGGAGTTTTAGATTTGGACA-3\u2032 (reverse primer). Selected malemice founders were crossed with C57BL/6 females. Formutation c.4-5insTCC described in this paper, founder malewas crossed with two F1 heterozygous daughters. Offspringwas genotyped and two sibling females were selected foreach group  forfurther analysis.Mutations were detected using PCR and Sanger sequencingof the target region of the Csn1s1 exon 2 (Supplementary 1)http://www.bionet.nsc.ru/vogis/download/pict-2021-25/appx8.pdfSupplementary materials 1 and 2 are available at: All experiments were conducted at the Centre for Genetic Resources of Laboratory Animals at the Institute ofCytology and Genetics, SB RAS (RFMEFI61914X0005 andRFMEFI61914X0010). All experiments were performed inaccordance with protocols and guidelines approved by theAnimal Care and Use Committee Federal Research Centreof the Institute of Cytology and Genetics, SB RAS operatingunder standards set by regulations documents Federal HealthMinistry (2010/708n/RF), NRC and FELASA recommendations. Experimental protocols were approved by the BioethicsReview Committee of the Institute of Cytology and Genetics,SB RAS.Droplet digital PCR analysis. Total cellular RNA wasextracted from mouse mammary glands at day 8 of lactationusing TRI Reagent (Sigma-Aldrich). 1 \u03bcg of total RNA wasused to generate cDNA in a 20 \u03bcl reaction using RevertAidRT Kit (Thermo Fisher Scientific) with random hexamerprimers according to the manufacturer\u2019s instructions. DropletDigital PCR (ddPCR) was performed using a QX100 system(Bio-Rad) with primers and probes specific for the Csn1s1and Csn2 mouse transcripts (Supplementary 2). The primersand probes sequences were as follows: 5\u2032-TGTAGTGGATCAGGCACTGG-3\u2032 (Csn1s1 forward primer), 5\u2032-TCCTTGGAGACAATGGGCTT-3\u2032 (Csn1s1 reverse primer), 5\u2032-HEXCCAGTTCTCTGTTCAGCCCTTCCCACA-BHQ2\u20133\u2032(Csn1s1 probe), 5\u2032-AGGACTTGACAGCCATGAAGG-3\u2032(Csn2 forward primer), 5\u2032-ATGTTCAACAGATTCCTCACTGGA-3\u2032 (Csn2 reverse primer), 5\u2032-FAM-ATCCTCGCCTGCCTTGTGGCCCTTGC-BHQ1\u20133\u2032 (Csn2 probe). ddPCRreactions were set in 20 \u03bcl volumes containing 1\u00d7 ddPCR\u2122Supermix for Probes (no dUTP), 900 nM primers and 250 nMprobes, and 1 \u03bcl of 5000-fold diluted cDNA. ddPCR reactionsfor each sample were performed in duplicates. PCR was conducted according to the following program: 95 \u00b0C for 10 min,then 40 cycles of 95 \u00b0C for 30 s and 61 \u00b0C for 1 min, witha ramp rate of 2 \u00b0C per second, and a final step at 98 \u00b0C for5 min. The results were analyzed using QuantaSoft software(Bio-Rad). Concentrations of cDNA copies of Csn1s1 andCsn2 were derived from ddPCR and relative expression ofCsn1s1 to Csn2 was calculated for each animal.Milk and mammary gland protein analysis. Milk wasobtained from narcotized female mice at day 8 of lactationafter oxytocin administration . Themilk was collected with a pipette attached to an aspiration device, transferred into a microcentrifuge tube and stored at\u201380 \u00b0C. Inguinal mammary glands (MGs) were extracted fromthe same (euthanized) mice and stored at \u201380 \u00b0C. For protein extraction MGs were minced in Dounce homogenizers,resuspended in RIPA buffer  with protease inhibitor cocktail (1x Complete ULTRA(Roche), 1x PhosSTOP (Roche), 5 mM NaF (Sigma)). Thelysates were incubated on ice for 30 min and then centrifugedat 4 \u00b0C for 10 min at 10\u200a000 g. Supernatant was sonicatedand stored at \u201380 \u00b0C. Total protein concentrations for milkand MG lysates were determined with Pierce\u2122 BCA Protein Assay Kit (Thermo Fisher Scientific), according to themanufacturer\u2019s instructions. To prepare samples for SDSPAGE, milk or MG protein samples were mixed with RIPAand SDS-PAGE loading buffer (Bio-Rad) to a final concentration of 1 \u03bcg/\u03bcl and heated at 65 \u00b0C for 20 minutes. Thesamples (25 \u03bcg) were separated on a 12 % polyacrylamidegel and stained with Coomassie Blue G-250. ThermoFisherPageRuler\u2122 Prestained Protein Ladder (10 to 180 kDa) wasused as a protein molecular weight marker. Csn1s1, albuminand total protein concentrations were evaluated using QuantityOne (Bio-Rad) and ImageJ software.Coomassie-stained gel was used for wet transfer of theproteins to PVDF membrane .The membrane was blocked with 5 % milk in TBST  for 2 hoursand incubated with primary mouse anti-Csn1s1 antibodies(1:1000)  overnight at4 \u00b0C. Next day membrane was repeatedly washed with TBSTand incubated with secondary mouse HRP-antibodies (1:1000) at 25 \u00b0C for 2 hours.Immunodetection was performed with ECL substrate solution, according tothe manufacturer\u2019s instructions.Generation of the Csn1s1 mutant miceWe performed pronuclear microinjections with Cas9 mRNAand sgRNA against the first coding exon of the mouse Csn1s1gene . Cas9-induced mutations in this region couldpotentially affect signal peptide coding sequence and leadto altered milk composition. We screened founder mice by Sanger sequencing and identified multiple random mutations at the Cas9 cut site (presented in Supplementary 1).The sgRNA targeted the Csn1s1 site with high efficiencyas we detected 41 mutant alleles, 4\u20135 mosaic alleles  and only 4\u20135 wild-type allelesin 20 founder mice . Asexpected, mutations clustered around the sgRNA cut site(3 bp from PAM). Most of the mutations represented smalldeletions (1\u201310 bp), but we detected several larger deletionsas well (100\u2013300 bp). Of note, some of the unique mutationvariants had increased incidence rate. For example, 12 bpdeletion (GAAACTCCTCAT) arose independently four timesand another 10 bp deletion (CCATGAAACT) \u2013 three times(see Supplementary 1). We suspect this bias towards somevariants is caused by microhomology-mediated end-joining(MMEJ), since these two mutations are flanked with 3 and 2similar nucleotides  (see Supplementary 1). Although mutations were mostly deleterious forthe gene expression and led to a Csn1s1 knockout (KO) byframeshift, several in-frame mutation variants resulted insubtle changes in signal peptide coding sequence withoutKO (see Supplementary 1). We selected one of the mutants,tagged Csn1s1c.4-5insTCC, which had a 3 bp insertion followingthe start codon . To study gene expression andmilk composition we chose 6 female siblings  from the Csn1s1c.4-5insTCCline for milk and mammary glands collection. Mutation c.4-5insTCC leads to reducedexpression of the Csn1s1 geneWe estimated the Csn1s1 gene expression in mammary glandsof wild-type and mutant mice at day 8 of lactation usingdroplet digital PCR (ddPCR). We selected Csn2 (\u03b2-casein) asa reference gene as it has a similar expression profile in mammary gland (see Supplementary 2). ddPCR analysis revealedthat Csn1s1:Csn2 ratio was roughly 1:3 (0.338) in wild-typesiblings , which is in agreement with publisheddata . Heterozygous and homozygousCsn1s1c.4-5insTCC siblings showed lower Csn1s1 expressionwith ratios around 1:4 (0.248) and 1:6 (0.168), respectively, compared to wild-type siblings. We also usedfemales from the parental strains C57BL/6 and CBA as controls for normal caseins level . In rare cases, Cas9activity can provoke rearrangements near the target locus.We sequenced mutation-flanking regions, including theCsn1s1 promoter and surrounding introns (2.3 kb+1.1 kb)from homozygous mice (data not shown). We also sequencedtop two off-targets for the Csn1s1 sgRNA in the F0 founder. No mutations werefound in the examined sequences. Thus, we could confirmthat the 3 bp in-frame insertion in the first coding exon led toa 30\u201350 % decrease in the Csn1s1 gene expression. Milk protein composition in c.4-5insTCC mice We measured Csn1s1 protein levels and confirmed that mutation has a negative effect on milk composition. We collectedmilk and mammary glands from lactating females at the day 8of lactation. Csn1s1 knockout mouse was taken from anotherexperiment (\u201cKO\u201d) as additional control for the Csn1s1 levels.Separation of milk proteins on 12 % SDS\u2010PAGE resulted ina typical band pattern for mouse milk . In wildtype mice, Csn1s1 represents a major protein fraction andcorresponds to approx. 30 % of total milk protein . In homozygous mutants, loss of Csn1s1 could beobserved both at the Coomassie-stained gel  andafter western blotting . Csn1s1 levels fell down to30 % in homozygotes, both for milk and for mammary glandlysates . However, exact ratios varied depending on the control protein band used for calculations. For instance, we performed the following calculations for the milkCsn1s1: Csn1s1 (gel) vs total protein (gel) \u2013 40 % reduction; Csn1s1 (gel) vs albumin (gel) \u2013 70 % reduction; Csn1s1(western blot membrane) vs total protein (gel) \u2013 80 % reduction. This effect was even more pronounced in mammarygland lysates (intracellular casein levels): Csn1s1 (westernblot membrane) vs total gel \u2013 80 % reduction.We report a novel in-frame Csn1s1 hypomorphic mutationthat leads to a 50 % gene expression decrease in mice. Inmost cases, mutation effect is tied to disruption of a regulatory element  . Frame-shifting indels in the coding sequencecould initiate transcript surveillance pathway called nonsense-mediated mRNA decay (NMD) . Alternatively, in-frame mutationscan lead to exon removal by alternative splicing . Essentially, exon skipping could be promoted by internal exon splicing enhancersand suppressors (ESEs and ESSs) which are hard to predict, unliketypical splice site mutations . In ourmutant mice, promoter had no alterations as the mutationhappened in the coding sequence, quite far from a transcription start site. We assumed that it affected transcript stabilityor splicing by unknown mechanism. It should also be notedthat mutated Csn1s1 protein was still secreted in milk, thusthe function of N-terminal signal peptide was not criticallyaffected by the mutation. We demonstrated that CRISPR/Cas9 approach could be conveniently exploited to induce a spectrum of mutations in theCsn1s1 gene either by random mutagenesis, or, ideally, by aset of single-stranded oligo DNA nucleotides (ssODNs). Ourresults warn that careful examination of the gene\u2019s expressionis required in addition to protein analysis. The authors declare no conflict of interest.An L.Y., Yuan Y.G., Yu B.L., Yang T.J., Cheng Y. Generation of human lactoferrin transgenic cloned goats using donor cells with dualmarkers and a modified selection procedure. Theriogenology. 2012;78:1303-1311. DOI 10.1016/j.theriogenology.2012.05.027.Burkov I.A., Serova I.A., Battulin N.R., Smirnov A.V., Babkin I.V.,Andreeva L.E., Dvoryanchikov G.A., Serov O.L. Expression of thehuman granulocyte-macrophage colony stimulating factor (hGMCSF) gene under control of the 5\u2032-regulatory sequence of the goatalpha-S1-casein gene with and without a MAR element in transgenicmice. 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Short communication: Confirmation of candidate causativevariants on milk composition and cheesemaking properties in Montb\u00e9liarde cows. J. Dairy Sci. 2018;101:10076-10081. DOI 10.3168/jds.2018-14986.Sterne-Weiler T., Sanford J.R. Exon identity crisis: disease-causing mutations that disrupt the splicing code. Genome Biol. 2014;15(1):201.DOI 10.1186/gb4150. PMID 24456648. PMCID PMC4053859.Thompson B.A., Walters R., Parsons M.T., Dumenil T., Drost M.,Tiersma Y., Lindor N.M., Tavtigian S.V., de Wind N., Spurdle A.B.,InSiGHT Variant Interpretation Committee. Contribution of mRNAsplicing to mismatch repair gene sequence variant interpretation.Front. Genet. 2020;11:798. DOI 10.3389/fgene.2020.00798. PMID32849802. PMCID PMC7398121.Tuladhar R., Yeu Y., Tyler Piazza J., Tan Z., Rene Clemenceau J., Wu X.,Barrett Q., Herbert J., Mathews D.H., Kim J., Hyun Hwang T.,Lum L. CRISPR-Cas9-based mutagenesis frequently provokeson-target mRNA misregulation. Nat. Commun. 2019;10(1):4056.DOI 10.1038/s41467-019-12028-5. PMID 31492834. PMCIDPMC6731291.Uusi-Oukari M., Hyttinen J.M., Korhonen V.P., V\u00e4sti A., Alhonen L.,J\u00e4nne O.A., J\u00e4nne J. Bovine alpha s1-casein gene sequences directhigh level expression of human granulocyte-macrophage colonystimulating factor in the milk of transgenic mice. Transgenic Res.1997;6:75-84. DOI 10.1023/a:1018461201385.Wan Y.J., Zhang Y.L., Zhou Z.R., Jia R.X., Li M., Song H., Wang H.Z.,Wang L.Z., Zhang G.M., You J.H., Wang F. Efficiency of donor cellpreparation and recipient oocyte source for production of transgeniccloned dairy goats harboring human lactoferrin. Theriogenology.2012;78:583-592. DOI 10.1016/j.theriogenology.2012.03.004.Wang K., Yan H., Xu H., Yang Q., Zhang S., Pan C., Chen H., Zhu H.,Liu J., Qu L., Lan X. A novel indel within goat casein alpha S1 geneis significantly associated with litter size. Gene. 2018;671:161-169.DOI 10.1016/j.gene.2018.05.119.Wu X., Lin Y., Xiong F., Zhou Y., Yu F., Deng J., Huang P., Chen H.The extremely high level expression of human serum albumin in themilk of transgenic mice. Transgenic Res. 2012;21:1359-1366. DOI10.1007/s11248-012-9612-4.Yamaji D., Kang K., Robinson G.W., Hennighausen L. Sequential activation of genetic programs in mouse mammary epithelium duringpregnancy depends on STAT5A/B concentration. Nucleic Acids Res.2013;41:1622-1636. DOI 10.1093/nar/gks1310.Yu Z., Meng Q., Yu H., Fan B., Yu S., Fei J., Wang L., Dai Y., Li N.Expression and bioactivity of recombinant human lysozyme in themilk of transgenic mice. J. Dairy Sci. 2006;89:2911-2918. DOI10.3168/jds.S0022-0302(06)72563-2.Yuan Y.G., An L., Yu B., Song S., Zhou F., Zhang L., Gu Y., Yu M.,Cheng Y. Expression of recombinant human alpha-lactalbuminin the milk of transgenic goats using a hybrid pomoter/enhancer.J. Anal. Method. Chem. 2014;281031. DOI 10.1155/2014/281031.Yue X.P., Zhang X.M., Wang W., Ma R.N., Deng C.J., Lan X.Y.,Chen H., Li F., Xu X.R., Ma Y., Lei C.Z. The CSN1S1 N and F alleles identified by PCR-SSCP and their associations with milk yieldand composition in Chinese dairy goats. Mol. Biol. Rep. 2011;38:2821-2825. DOI 10.1007/s11033-010-0428-0.Zhou S., Cai B., He C., Wang Y., Ding Q., Liu J., Liu Y., Ding Y.,Zhao X., Li G., Li C., Yu H., Kou Q., Niu W., Petersen B., Sonstegard T., Ma B., Chen Y., Wang X. Programmable base editing of thesheep genome revealed no genome-wide off-target mutations. Front.Genet. 2019;10:215. DOI 10.3389/fgene.2019.00215."}
+{"text": "The kidneys are among the organs most affected by leukemic involvement in chronic lymphocytic leukemia (CLL) patients in postmortem autopsy series 63%-90%) ,2. Howev%-90% ,2.9/L (lymphocytes: 60%); platelets, 247x109/L. The blood \ufb01lm appearances were suggestive of CLL. The following biochemical tests were abnormal: blood urea nitrogen, 44 mg/dL ; serum creatinine (sCr), 6.16 mg/dL ; CKD-EPI estimated-glomerular filtration rate, 8 mL/min/1.73 m2 . His urine had no leukocytes or casts, with 3 erythrocytes/field and proteinuria in the non-nephrotic range (1779 mg/24 h). Serological tests for hepatitis B, hepatitis C, HIV, and autoimmune kidney disorders were negative. Serum-free light chain (sFLC) kappa was 317.5 mg/dL  and sFLC lambda was 22.2 mg/dL , but no monoclonal bands in serum or urine immunofixation were detected. Renal ultrasound showed normal-sized kidneys with no evidence of obstructive nephropathy. A renal biopsy was performed, which demonstrated heavy infiltration of diffuse monomorphic neoplastic lymphocytes in the interstitium (9/L). After a year of follow-up, his renal function had improved to sCr of 2.04 mg/dL, and a complete blood count revealed hemoglobin of 14.7 g/dL, leukocytes of 9.15x109/L, and platelets of 294x109/L.A 72-year-old white male without any renal disease presented with general malaise and weight loss. He had been diagnosed with stage B(1) B-CLL four years before and was treated with a bendamustine and rituximab protocol. Physical examination revealed general lymphadenopathy and hepatomegaly. Complete blood count findings were as follows: hemoglobin, 11.3 g/dL; leukocytes, 411.4x10rstitium . Six of rstitium  and 1C wBesides LI, renal impairment in CLL patients can be associated with prerenal azotemia, thrombotic microangiopathy, acute tubular necrosis, acute interstitial nephritis, uric acid nephropathy, light chain nephropathy, amyloidosis, obstructive nephropathy, glomerulonephritis, and cryoglobulinemia . LI of t"}
+{"text": "Correction to: J Foot Ankle Res 14, 15 (2021)https://doi.org/10.1186/s13047-020-00435-7Following publication of the original article , we wereThe original article has been corrected.\u00a0Additional file 2 has been replaced.Additional file 2. Development of the systematic podiatry protocol."}
+{"text": "A total of 50 neonates  participated in this study. LBW-VLBW neonates were further stratified as those with  and without  structural MRI (sMRI) abnormalities. TRUST and PC MRI studies were undertaken to determine OEF, CBF, and CMRO2. Ultimately, CMRO2 proved significantly lower (p = 0.01) in LBW-VLBW (vs term) neonates, both LBW-VLBW-a and LBW-VLBW-n subsets showing significantly greater physiologic deficits than term controls . CMRO2 and CBF in LBW-VLBW-a and LBW-VLBW-n subsets did not differ significantly (p > 0.05), although OEF showed a tendency to diverge (p = 0.15). However, OEF values in the LBW-VLBW-n subset differed significantly from those of term controls (p = 0.02). Compared with brain volume or body weight, these physiologic parameters yield higher area-under-the-curve (AUC) values for distinguishing neonates of the LBW-VLBW-a subset. The latter displayed distinct cerebral metabolic and hemodynamic, whereas changes were marginal in the LBW-VLBW-n subset  by comparison. Physiologic imaging may therefore be useful in identifying LBW-VLBW newborns at high risk of irreversible brain damage.Low birth-weight (LBW) and very low birth-weight (VLBW) newborns have increased risks of brain injuries, growth failure, motor difficulties, developmental coordination disorders or delay, and adult-onset vascular diseases. However, relatively little is known of the neurobiologic underpinnings. To clarify the pathophysiologic vulnerabilities of such neonates, we applied several advanced techniques for assessing brain physiology, namely T2-relaxation-under-spin-tagging (TRUST) magnetic resonance imaging (MRI) and phase-contrast (PC) MRI. This enabled quantification of oxygen extraction fraction (OEF), global cerebral blood flow (CBF), and cerebral metabolic rate of oxygen (CMRO Preterm newborns of low birth weight  or very low birth weight  present a substantial public health problem. In 2015, the worldwide prevalence was 14.6%, with 91% confined to low- and middle-income countries [primarily Southern Asia (24%) and sub-Saharan Africa (48%)]. More than 80% of perinatal deaths occur in LBW neonates .Although most LBW and VLBW preterm infants show catch-up gains in height and weight, they are still at increased risk of brain injuries , growth 133Xenon clearance  technologies have allowed noninvasive and quantitative measurements of critical neurophysiologic parameters, without need of contrast agents. The oxygen extraction fraction (OEF) of the brain is assessable using T2-relaxation-under-spin-tagging (TRUST) MRI technique , and glo2 using T2 prepared tissue relaxation inversion recovery (T2-TRIR) pulse sequences and ASL (2 and CBF in asphyxiated newborns with severe HIE (r = 0.88)  is another valuable means of perfusion MRI for this purpose. However, its signal-to-noise ratio (SNR) tends to be lower, the scan times lengthier, and there is a potential for confounding factors . In an earlier effort, infants with hypoxic ischemic encephalopathy (HIE) showed lower CBF and CMRO and ASL . Newborn = 0.88) . Our pre = 0.88) . Tortora = 0.88) . There h2 in a mixed group of LBW and VLBW newborns, comparing outcomes with those of term neonates.In the present study, we used TRUST MRI and PC MRI to determine OEF, CBF, and CMROn = 41) at birth weights >1,000 g but <2,500 g. Term neonates serving as controls (n = 9) had birth weights \u22652,500 g, were \u226537 weeks at birth, and showed no sMRI abnormalities. Population characteristics and results of blood gas analysis were listed in Our investigational protocol received approval from the Ethics Committee at Shengjing Hospital of China Medical University. Between June 2016 and December 2018, a total of 50 newborns accrued for study. The structural MRI pulse sequences performed included axial and sagittal T1-weighted imaging (TIWI), axial T2-weighted imaging (T2WI), and diffusion-weighted imaging (DWI). Congenital malformations, severe infections, or unusable MRI studies were grounds for exclusion. Subjects qualified as LBW-VLBW  obtained from all infants (LBW-VLBW and term) were viewed as Picture Archiving and Communication Systems (PACS) files by two neuroradiologists (YQ and XYS) with 10 years of experience, each blinded to group data. They independently scored MRI abnormalities for each infant, using a method similar to one already described  that focA 3T MRI system equipped with a phased-array head coil  was used for all MR scans. Prior to imaging procedures, a pediatrician sedated infants through nasal feedings of chloral hydrate (50 mg/kg). This is standard clinical practice at our institution. All newborns were well fed, visually monitored, and hearing protected during scans.2; and scan time, 30 s.Axial T1-weighted spin-echo images were acquired at the following settings: repetition time/echo time (TR/TE), 200 ms/2.3 ms; section thickness, 5 mm; field of view (FOV), 180 mm \u00d7 150 mm \u00d7 89 mm; matrix size, 224 \u00d7 162; and scan time, 34.4 s. For sagittal T1-weighted spin-echo images, the following settings were used: TR/TE, 250 ms/2.3 ms; section thickness, 5 mm; FOV, 230 mm \u00d7 230 mm \u00d7 107 mm; matrix size, 256 \u00d7 250; and scan time, 33.0 s. Axial T2WI used the following settings: TR/TE, 5,000 ms/80 ms; section thickness, 5 mm; FOV, 180 mm \u00d7 150 mm \u00d7 90 mm; matrix size, 112 \u00d7 112; and scan time, 40.9 s. Axial echo-planar imaging (EPI) DWI was performed as follows: TR/TE, 3,500 ms/63 ms; section thickness, 5 mm; FOV, 180 mm \u00d7 180 mm \u00d7 89 mm; b values, 0 and 1,000 s/mmOxygen extraction fraction was calculated as follows:where Ya and Yv signify arterial and venous oxygenation, respectively. Ya was measured peripherally, via pulse oximeter applied to neonatal toes. Yv was measured by TRUST MRI technique, given that blood T2 has a calibrationable relation with oxygenation level. This sequence utilizes a spin-labeling module to isolate pure venous blood signals, thereafter applying a series of T2-preparation pulses to modulate the MRI signal, the monoexponential fitting of which yields blood T2 . A T2-YvFor measuring CBF, PC MRI employed bipolar gradients to encode flow velocities of blood supplying major feeding arteries of the brain, specifically left and right internal carotid arteries (LICA and RICA) and left and right vertebral arteries (LVA and RVA), allowing quantitation of total blood flow to the brain. Flow values were then normalized to brain volume, expressing CBF in units of mL/100 g/min. Time-of-flight MR angiograms (TOF-MRAs) were first performed to allow visualization of above-referenced feeding arteries. Imaging slabs were positioned at center of epistropheus, with 60-mm saturation slabs placed above to suppress venous signals. Settings of TOF MRA were as follows: TR/TE, 20 ms/3.45 ms; flip angle, 18\u00b0; FOV, 90 mm \u00d7 90 mm \u00d7 20 mm; voxel size, 0.8 mm \u00d7 0.8 mm \u00d7 2 mm; and scan duration, 23.7 s.Based on MRA images, four PC MRI scans were performed by orienting imaging slices perpendicular to and centered on respective target arteries, as previously described . Scans oProcessing of PC MRI data again followed existing procedures . Manual 2 was calculated according to Fick\u2019s principle  on scan age, sex, and diagnostic group (independent variables). In regression analyses, paired groups were encoded as follows: LBW-VLBW (0) vs term (1) neonates; LBW-VLBW-a (1) vs LBW-VLBW-n (0) subsets; LBW-VLBW-a neonates (1) vs term controls (0); and LBW-VLBW-n neonates (0) vs term controls (1). LBW-VLBW newborns underwent MRI scans once their vital signs had stabilized, and ventilators were no longer required (3 weeks old on average). Term infants had good vital signs and were amenable to scanning at any time (1 week old on average). There were distinct chronologic differences in scan ages of these pairings. Due to remarkable disparities , it was 1 and b2 were based on results of an age- and sex-adjusted regression model similarly reported  and term neonates (n = 9), LBW-VLBW-a (n = 24) and LBW-VLBW-n (n = 17) subsets, LBW-VLBW-a neonates (n = 24) and term controls (n = 24), and LBW-VLBW-n neonates (n = 17) and term controls (n = 24). We also analyzed birth weight, brain volume, and cerebral physiologic parameters independently, generating receiver operating characteristic (ROC) curves to distinguish the LBW-VLBW-a subset from all other neonates and from the LBW-VLBW group overall. Inter-rater variations in MRI scores determined by YQ and XYS were assessed via intraclass correlation coefficient (ICC), rated as follows: 1.0\u20130.81, excellent; 0.80\u20130.61, very good; 0.60\u20130.41, good; 0.40\u20130.21, reasonable; and 0.20\u20130.00, poor. All computations were driven by standard software , setting significance at p < 0.05.where coefficients breported . Correctp = 0.89); CMRO2, 29.9 \u00b1 11.4 \u03bcmol/100 g/min vs 48.9 \u00b1 12.4 \u03bcmol/100 g/min (p < 0.001); CBF, 13.6 \u00b1 5.0 mL/100 g/min vs 19.5 \u00b1 3.8 mL/100 g/min (p = 0.002); and brain volume, 265.2 \u00b1 74.5 mL vs 354.1 \u00b1 52.0 mL (p = 0.001). Only OEF proved similar, the other parameters differing significantly by group.In LBW-VLBW and term neonates, measured parameters were as follows: OEF, 31.3 \u00b1 10.5% vs 30.8 \u00b1 6.2% (n = 41), including both LBW-VLBW-a and LBW-VLBW-n subsets, are detailed in p > 0.05). Fifteen LBW-VLBW newborns had required mechanical ventilation.Population characteristics and diagnostic outcomes  of the LBW-VLBW group (2 was significantly lower (p = 0.01) in LBW-VLBW (vs term) newborns, CBF (p = 0.06) and OEF (p = 0.67) did not differ significantly. Furthermore, CMRO2 (p = 0.48) and CBF (p = 0.63) failed to show scan age-related increases during this period of early development; and despite significant differences in birth weight, LBW-VLBW and term neonates did not differ significantly in brain volume.We first compared physiologic measures of LBW-VLBW and term neonates . Althoug2 rates relative to controls . Contrary to expectations, no significant differences in CMRO2 (p = 0.30), CBF (p = 0.50), or OEF (p = 0.15) recordings were evident across subsets upon linear regression analysis. Relative to term neonates, OEF was significantly lower (p = 0.02) in the LBW-VLBW-n subset.Next, we compared the two subsets of LBW-VLBW neonates (LBW-VLBW-a and LBW-VLBW-n) with term newborns, both displaying lower CMRO1, scan age; b2, sex), with 95% confidence interval (CI) of coefficients and respective p-values. Once corrected for scan age and sex, group comparisons of birth weight, CMRO2, CBF, OEF and brain volume produced similar results , the LBW-VLBW-a subset vs term controls , and the LBW-VLBW-n subset vs term controls . This suggests that observed differences in physiologic parameters (CMRO2 and CBF) of the LBW-VLBW-a subset and term controls stemmed from brain injuries and did not reflect scan age or sex. Corrected OEF values in the LBW-VLBW-n subset did differ significantly (p < 0.001) from those of term controls. There were no significant differences in corrected values of LBW-VLBW-a and LBW-VLBW-n subsets (p > 0.05). results . Correctp = 0.25], OEF , CMRO2 , CBF , and brain volume  in distinguishing the LBW-VLBW-a subset from term and LBW-VLBW neonates are shown in 2 demonstrated fair diagnostic performance in this regard.Receiver operating characteristic curves for birth weight . Some term neonates were also discounted due to abnormal laboratory results or MRI findings. Presently, the potential for neurocognitive deficits in these infants and the relevance of CMRO2 reductions on neurocognitive deficits. We will also bolster subject recruitment, further stratifying by birth weight  and sex, and foster a multicenter approach to ascertain neurocognitive thresholds for reductions in CMRO2. Owing to brain immaturity, especially in VLBW infants, there is exceptional vulnerability to injuries undermining the microstructural integrity and tract connectivity of periventricular white matter (a vascular watershed territory)  . Knut harritory) . We must2 at brain level. Finally, the image quality in It is unclear at this point whether administering an anesthetic, such as chloral hydrate, has any cerebrovascular hemodynamic ramifications. There is no direct evidence in studies of humans and animals that it increases CBF or lowers glucose metabolism . However2, and higher OEF tended to show minimal structural abnormalities. Physiologic imaging may be a useful means of identifying those LBW-VLBW newborns at high risk of developing irreversible brain damage.In the present study, we have demonstrated that structural damage in LBW-VLBW-a neonates is associated with metabolic and hemodynamic deficits of the brain. LBW-VLBW-n neonates with lower CBF and CMROThe raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The study protocol had ethical approval from the Ethics Committee of Shengjing Hospital of China Medical University (IRB2015PS28K). Written informed consent from the participants\u2019 legal guardian/next of kin was not required to participate in this study in accordance with the national legislation and the institutional requirements.YQ contributed study concepts and design, and providing post-processing assistance. Both authors participated in analysis of experiment results, drafting and editing of the manuscript, and have read and approved the final manuscript for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Clostridioides difficile infection (CDI) is the leading cause of life-threatening health care-related gastrointestinal illness worldwide. Phylogenetically appropriate closed reference genomes are essential for studies of C. difficile transmission and evolution. Here, we provide high-quality complete hybrid genome assemblies for the three most prevalent C. difficile strains causing CDI in Australia. Clostridioides difficile causes life-threatening diarrhea and health care-related gastrointestinal infections globally (C. difficile population distinct from that of the rest of the world (C. difficile strains causing C. difficile infection (CDI) in Australia, PCR ribotype 014 (RT014) , RT002 (11.8%), and RT056 (5.4%)  were selected from >1,500 isolates recovered from patients with symptomatic CDI, part of the ongoing nationwide longitudinal surveillance of CDI in Australia, the C. difficile Antimicrobial Resistance Surveillance (CDARS) study  for 48\u2009h . ONT sequencing was performed on a MinION Mk1C device  using an R9 generation flow cell following a DNA by ligation protocol (SQK-LSK109). Filtlong v0.2.0 (https://github.com/rrwick/Filtlong) was used to filter the low-quality reads , resulting in 2.28 (S-0352), 2.59 (S-0253), and 6.66 (S-0942) Gb of sequence data, respectively. The hybrid assembly of ONT and Illumina reads was performed using Unicycler v0.4.8  and annotated using the NCBI Prokaryotic Genome Annotation Pipeline v5.2  and at the ENA under BioProject accession number PRJEB41588 (Illumina sequence data); see The genome data are available at GenBank under BioProject accession number"}
+{"text": "Scientific Reports 10.1038/s41598-021-81366-6, published online 19 January 2021Correction to: The original version of this Article contained extensive errors in the Reference list.Reference 26 was incorrectly split into Reference 26 and Reference 27.\u201cTrainer, M. N. & Freud, P. J. High-Concentration Submicron Particle Size Distribution by Dynamic Light Scattering Power spectrum development with heterodyne technology advances biotechnology and nanotechnology measurements.\u201dnow reads:\u201cTrainer, M. N. & Freud, P. J. High-Concentration Submicron Particle Size Distribution by Dynamic Light Scattering Power spectrum development with heterodyne technology advances biotechnology and nanotechnology measurements. Microtrac, Inc. Application Note SL-AN-05 Rev B. (2009).\u201dAs a result, References 27\u201347 were incorrectly listed as References 28\u201348.The original Article has been corrected."}
+{"text": "The chemokine receptor 4 (CXCR4) and 7 (CXCR7) are G-protein-coupled receptors (GPCRs) activated through their shared ligand CXCL12 in multiple human cancers. They play a key role in the tumor/tumor microenvironment (TME) promoting tumor progression, targeting cell proliferation and migration, while orchestrating the recruitment of immune and stromal cells within the TME. CXCL12 excludes T cells from TME through a concentration gradient that inhibits immunoactive cells access and promotes tumor vascularization. Thus, dual CXCR4/CXCR7 inhibition will target different cancer components. CXCR4/CXCR7 antagonism should prevent the development of metastases by interfering with tumor cell growth, migration and chemotaxis and favoring the frequency of T cells in TME. Herein, we discuss the current understanding on the role of CXCL12/CXCR4/CXCR7 cross-talk in tumor progression and immune cells recruitment providing support for a combined CXCR4/CXCR7 targeting therapy. In addition, we consider emerging approaches that coordinately target both immune checkpoints and CXCL12/CXCR4/CXCR7 axis. Chemokines are small chemoattractant molecules that control cell migration, proliferation and survival in physiological and pathological processes including cancer . They ar\u2212/\u2212 knock-out mice die before birth -dependent phosphorylation and interaction with \u03b2 \u2013arrestin  GPCRs encoded on chromosome 2.1 , 17. CXCre birth . CXCR4 \u2212ogenesis . CXCL12 ogenesis . CXCL12-ogenesis . In addiogenesis . When CXarrestin . CXCR7 parrestin , angiogearrestin , neurogearrestin  and cardarrestin . Althougarrestin . Initialarrestin . Withoutarrestin . CXCR7, arrestin . CXCR7 carrestin . Studiesarrestin  or in hearrestin . CXCR4/Carrestin . Overallvia Epidermal Growth Factor Receptors (EGFRs) and Matrix Metallopeptidase 9 (MMP-9) (via angiogenesis (An active CXCL12/CXCR4 pathway is considered a feature of aggressive tumors  as it po (MMP-9) . Also CX (MMP-9) . CXCR7 a (MMP-9) . The pro (MMP-9) , 48. CXC (MMP-9) \u201351. In o (MMP-9)  while CX (MMP-9) . CXCR7 m (MMP-9) . In cerv (MMP-9) . In lung (MMP-9) . Accordi (MMP-9) . Convers (MMP-9) . CXCL12  (MMP-9) . In addi (MMP-9) . In neur (MMP-9) . In breaogenesis . Hence, ogenesis , EGFR , myeloid derived suppressor cells (MDSCs), and dendritic cells (DCs) to the tumor niche . CXCL12 via Raf-ERK pathway , CXCR4 is expressed in tumor endothelium sprouting tumor vessels  and CXCR pathway . CXCR7 e pathway  or IL-1b pathway , by lipo pathway  or durin pathway . CXCR7 i pathway  and surv pathway . It is s pathway , 84 and  pathway . CXCL12 and TECs . Thus, Celopment .DCs are the most potent antigen presenting cells (APCs) in the immune system . Immatur+CD25high FoxP3+) are CD4+ T cells with predominantly suppressive activity (5\u201310% of circulating CD4+ T cells in humans). Tregs impair immune effector cells function via cytokines, direct lysis, inhibitory receptors, metabolic disruption, IL-2 depletion or inducing an immunosuppressive microenvironment  potentiates migration toward vascular-associated CXCL12-positive cells in the BM. In lymphoma-bearing mice, TCXCR4 potentiates the effector function increasing tumor protection -\u03b3 respectively, particularly in macrophages (The only approved drug CXCR4 inhibitor is AMD3100 (known as Plerixafor or Mozobil)  while muin vivo . In prosin vivo . Other ain vivo  or CCX73in vivo . The antin vivo . Recentlin vivo . It bindrophages . Some anrophages . The cycrophages . A CXCR4rophages . Peptide+CD25\u2013Foxp3+IL2+CD40L+ helper-like cells (-L1) small interfering RNA (siPD-L1). The nanocomplex promotes T cell infiltration, decreases alpha-smooth muscle actin (\u03b1-SMA) and collagen, reduces MDSCs and Tregs recruitment (Recently, CXCR4 antagonists have been coupled to ICIs with the intent to remodel TME improving ICIs efficacy . Since tke cells . Inhibitke cells  while take cells . A nanocruitment . Thus, cImmuno-resistance and vascularization are acquired tumor features that contribute to cancer growth and metastasis. Among the different signaling pathways, directly or indirectly involved in cancer immune-resistance and angiogenesis, CXCR4/CXCR7/CXCL12 is crucial for participating in cancer migration, angiogenesis and immunosuppressive cell recruitment. Thus, the inhibition of the CXCR4/CXCL12 or CXCR7/CXCL12 axis is attractive in cancers overexpressing both receptors such as colorectal cancer , renal cSSa, CI, and SSc contributed in conception and design of the study. AMT, AC, FA, and GG supervised the study. SSa, CI, and SSc wrote and edited the manuscript. SSa and CI equally contributed. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Staphylococcus aureus (CA-MRSA) has become a major cause of S. aureus infections globally. We report the complete genome sequences of three of the earliest CA-MRSA strains isolated from remote Australian Indigenous communities in the Kimberley region of Western Australia.Initially reported in Western Australia in the 1980s, community-associated methicillin-resistant  Staphylococcus aureus (MRSA) is a leading cause of health care-associated infections worldwide and has emerged as a major cause of infections in the community . Each genome was also sequenced with the Illumina NextSeq 500 platform using the Nextera XT DNA library preparation kit (150-bp paired-end chemistry) and the NextSeq 500/550 kit v2.5 . Nesoni clip (github.com/Victorian-Bioinformatics-Consortium/nesoni) was used to remove adaptor sequences and to quality filter the reads. Illumina reads were mapped to the genome assembly with Minimap2 v2.17-r941 . The ONT SQK-RAD004 kit was used to create DNA sequencing libraries, which were loaded onto an ONT FLO-MIN106 flow cell and sequenced using ONT MinKNOW v20.10.3 and MinKNOW Core v4.1.2. Base calling was performed with Guppy v4.4.1+1c81d62 (ONT), using the dna_r9.4.1_450bps_hac.cfg model. NanoFilt . Unfilte.17-r941  was used.17-r941 . Genome .17-r941 . DefaultPRJNA703734 and PRJNA703736 and Sequence Read Archive (SRA) accession numbers SRX11246052, SRX11246051, SRX11247745, SRX11247744, SRX11246056, and SRX11246055, as indicated in GenBank accession numbers for the genome assemblies are provided in"}
+{"text": "First-generation COVID-19 vaccines are matched to spike protein of the Wuhan-H1 (WT) strain. Convalescent and vaccinee samples show reduced neutralization of SARS-CoV-2 variants of concern (VOC). Next generation DNA vaccines could be matched to single variants or synthetically designed for broader coverage of multiple VOCs.The synthetic consensus (SynCon\u00ae) sequence for INO-4802 SARS-CoV-2 spike with focused RBD changes and dual proline mutations was codon-optimized . Sequences for wild-type (pWT) and B.1.351 (pB.1.351) were similarly optimized. Immunogenicity was evaluated in BALB/c mice. Pre-clinical efficacy was assessed in the Syrian Hamster model.Figure 1. Design Strategy for INO-4802INO-4802 induced potent neutralizing antibody responses against WT, B.1.1.7, P.1, and B.1.351 VOC in a murine model. pWT vaccinated animals showed a 3-fold reduction in mean neutralizing ID50 for the B.1.351 pseudotyped virus. INO-4802 immunized animals had significantly higher (p = 0.0408) neutralizing capacity (mean ID50 816.16). ID50 of pB.1.351 serum was reduced 7-fold for B.1.1.7 and significantly lower (p = 0.0068) than INO-4802 (317.44). INO-4802 neutralized WT (548.28) comparable to pWT. INO-4802 also neutralized P.1 (1026.6) . pWT, pB.1.351 or INO-4802 induced similar T-cell responses against all variants. INO-4802 skewed towards a TH1-response. All hamsters vaccinated with INO-4802 or pB.1.351 were protected from weight loss after B.1.351 live virus challenge. 4/6 pWT immunized hamsters were completely protected. pWT immunized hamsters neutralized WT (1090) but not B.1.351 (39.16). INO-4802 neutralized both WT (672.2) and B.1.351 (1121) . We observed higher increase of binding titers following heterologous boost with INO-4802 (3.6 \u2013 4.4 log2-fold change) than homologous boost with pWT (2.0 \u2013 2.4 log2 fold change) .Figure 2. INO-4802 Induces Functional Humoral Immune Response Against SARS-CoV-2 Variants of ConcernFigure 3. INO-4802 Protects Hamsters Against Challenge With B.1.351 Live VirusFigure 4. Heterologous Boost with INO-4802 Induces Humoral Immune Response Against SARS-CoV-2 VariantsVaccines matching single VOCs, like pB.1.351 and pWT, elicit responses against the matched antigen but have reduced cross-reactivity. Presenting a pan-SARS-CoV-2 approach, INO-4802 may offer substantial advantages in terms of cross-strain protection, reduced susceptibility to escape mutants and non-restricted geographical use.Katherine Schultheis, MSc, Inovio Pharmaceuticals (Employee) Charles C. Reed, PhD, Inovio Pharmaceuticals  Viviane M. Andrade, PhD, Inovio Pharmaceuticals Inc. (Employee) Richa Kalia, MS, Inovio Pharmaceuticals  Jared Tur, PhD, Inovio (Employee) Blake Schouest, PhD, Inovio Pharmaceuticals (Employee) Dustin Elwood, PhD, Inovio Pharmaceuticals (Employee) Arthur Doan, n/a, Inovio (Employee) Patrick Pezzoli, BS, Inovio (Employee) Dinah Amante, BS, Inovio (Employee) David Weiner, PhD, Inovio  J Joseph Kim, PhD, Inovio (Employee) Laurent Humeau, PhD, Inovio Pharmaceuticals (Employee) Stephanie Ramos, PhD, Inovio Pharmaceuticals (Employee) Trevor R. F. Smith, PhD, Inovio  Kate Broderick, PhD, Inovio (Employee)."}
+{"text": "S. aureus (MRSA), vancomycin-resistant Enterococcus (VRE), ESBL- and carbapenemase-producing Enterobacteriaceae (E). We assessed T2R performance in detecting these resistant bacteria in whole blood (WB) and analyzed possible impact on time to appropriate Ab. Appropriate antibiotic (Ab) therapy of bloodstream infections (BSI) is often delayed by time to blood culture (BC) positivity, species (sp) identification and Ab sensitivity (sensi). The T2Resistance (T2R) Panel is a direct-from-blood (culture-independent) diagnostic that detects 13 genetic markers associated with methicillin-resistant We performed T2R using WB samples obtained from patients (pts) on the same day as BCs from July 2019-2020. Receipt of appropriate Ab was assessed at time of empiric, Gram stain-directed, MALDI-directed (sp identification) and sensi-directed therapy. T2R results were not available to care teams. Teams were notified of positive BCs. Stewardship optimized Abs based on sensi. S. aureus , Enterococcus , P. aeruginosa and others (n=10). 12 ESBL-E produced CTX-M 14/15. T2R sensitivity and specificity was 78% and 99%, respectively, compared to sequencing of resistance markers. Sensitivity was excellent for vanA/B, KPC (100% each), and CTX-M14/15 (92%); sensitivity was 58% for mecA/C. T2R detected resistance determinants in 3-7h. Median time to appropriate Ab was 16.3h, which was significantly longer for VRE (25.6h) and ESBL- or KPC-E (50.9h) BSIs than for T2R marker-negative bacteria . Pts with VRE or ESBL-/KPC-E BSI were less likely to received appropriate empiric Ab  than pts with T2R marker-negative BSI . Median times to achieve \u226580% appropriate Ab therapy of marker-negative, VRE and CTX-M/KPC-E BSIs were 15.5h (after Gram stain), 43.9h (after MALDI) and 63.5h (after sensi), respectively.BC from 103 pts grew 114 bacterial sp: E , Antibiotic TherapyThere was a significant delay in appropriate Ab therapy of BSIs, especially in pts infected with VRE and ESBL/KPC-E. T2R rapidly and accurately detected BSI caused by VRE and ESBL/KPC-E, and has the potential to significantly shorten time to appropriate Ab.Cornelius J. Clancy, MD, Merck (Grant/Research Support) Ryan K. Shields, PharmD, MS, Shionogi  Minh-Hong Nguyen, MD, Merck (Grant/Research Support)"}
+{"text": "NC_045512.2).We report a coding-complete genome sequence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strain SARS-CoV-2/BGD/GC001, isolated from a Bangladeshi patient with respiratory symptoms. Phylogenetic analysis assigned this strain to lineage B.1.1.7, which presented a total of 36 mutations in the spike and other genomic regions compared to strain Wuhan Hu-1 (GenBank accession number  Coronaviridae and genus Betacoronavirus (Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) belongs to the family onavirus . Lineagehttp://www.sanger.ac.uk/science/tools/smalt-0) and SAMtools v1.9   from the symptomatic patient was processed for nucleic acid isolation utilizing a QIAamp viral RNA minikit . SARS-CoV-2 was confirmed using a TaqMan real-time PCR (RT-PCR) assay . A cDNA ols v1.9 . De novo v3.11.1 . RATT wa45512.2) . The Nex45512.2)  and MEGA45512.2)  software45512.2) . DefaultNC_045512.2)  . Strain 45512.2) . MoreovePRJNA702998, BioSample accession number SAMN17993365, and GenBank accession number MW624725.1. The Illumina raw reads have been deposited in the NCBI Sequence Read Archive under accession number SRR13744683.The genome sequence of SARS-CoV-2/BGD/GC001 was deposited in the NCBI database under the BioProject accession number"}
+{"text": "Chimeric antigen receptor (CAR-T) T-cell therapy is a novel immunotherapy for cancer treatment in which patients are treated with targeted, genetically-modified T-cells. Common side effects include cytokine release syndrome, neurotoxicity, hypogammaglobulinemia, and increased susceptibility to infections. Long-term infectious outcomes are poorly characterized.We retrospectively examined patients who received CAR-T therapy at BIDMC & MGH from July 2016 to March 2020 and evaluated bacterial, fungal, viral, and parasitic infections at 3 months intervals to 1 year following cell infusion. The incidence, timing, and outcomes of the infectious complications were evaluated.In total, there were 47 patients; averaging 61.4 years of age (\u00b112 years). Primary indications for CAR-T therapy included diffuse large b-cell lymphoma (65%) and multiple myeloma (25%), chronic lymphocytic leukemia (2%) and mantle cell lymphoma (2%). Patients had received an average 4 \u00b1 2.9 lines of chemotherapy prior to CAR-T infusion; 19 subjects (40%) had a history of prior autologous stem cell transplant. All patients received acyclovir for antiviral prophylaxis and most received either trimethoprim-sulfamethoxazole  or atovaquone  for pneumocystis prophylaxis. In the first year, 35/47 (74.5%) of subjects experienced at least one infection with an infection rate of 84.4/10,000 person days. Median time to first infection was 59 days (range 1-338 patient days). 31/47 (66.0%) subjects had at least one bacterial infection, with pulmonary  sources being the most common site of infection. 13/47 (27.7%) of patients had a viral infection  and 6/47 (12.8%) had a proven or probable fungal infection. Death attributed to infection was noted in 2 subjects (4.3%), both related to COVID-19. Baseline IgG levels were significantly lower in the group with infections (p=0.028), while white blood cell count and absolute neutrophil counts were comparable.Table 1. Baseline Demographic, Clinical Characteristics, and Outcomes of 47 Recipients of CAR-T Cell Therapy by Infection StatusBMI: body mass index; DLBCL: diffuse large B-cell lymphoma; CLL: chronic lymphocytic leukemia; Flu/Cy: Fludarabine/cyclophosphamide; IVIG: intravenous immunoglobulin; WBC: white blood cell count; ANC: absolute neutrophil count; ALC: absolute lymphocyte count.Table 2. Characteristics of the 113 Infections in the 35 Subjects Who Developed InfectionsInfectious complications, particularly of bacterial etiology, are common in the first year following CAR-T therapy. These data may inform future prophylactic strategies in this patient population.Matthew Frigault, MD, Arcellx (Consultant)BMS (Consultant)Iovance (Consultant)Kite (Consultant)Novartis (Consultant) Jay A. Fishman, MD, Nothing to disclose Jon Arnason, MD, BMS/Juno (Advisor or Review Panel member)Regeneron (Advisor or Review Panel member)"}
+{"text": "Lung cancer is one of the most common types of cancer in the world. Although the mechanism of lungcancer is still unknown, a large number of studies have found a link between gene polymorphisms and the risk of lungcancer. The tumor suppressor p53 plays a crucial role in maintaining genomic stability and tumor prevention. MDM2is a critical regulator of the p53 protein. Despite the importance of p53 pathway in cancer, data on the contributionof SNPs of TP53 (rs1042522) and MDM2 (rs2279744) to the development of lung cancer are very contradictory. A metaanalysisthat collects quantitative data from individual studies and combines their results has the advantage of improvingaccuracy, providing reliable estimates, and resolving those issues in which studies on individual associationsare not effective enough. The aim of this study was to determine whether the TP53 (rs1042522) and MDM2 (rs2279744)polymorphisms confer susceptibility to lung cancer. A meta-analysis was conducted on the associations between theTP53 (rs1042522) and MDM2 (rs2279744) polymorphisms and lung cancer. A total of 51 comparison studies including25,366 patients and 25,239 controls were considered in this meta-analysis. The meta-analysis showed no associationbetween lung cancer and MDM2 (rs2279744) under any model. A noteworthy association of TP53 (rs1042522) withsusceptibility to lung cancer in overall pooled subjects was observed under three different models  and dominant). Stratification by ethnicity indicated an association between the TP53(rs1042522) and lung cancer in Asians and Caucasians. This meta-analysis demonstrates that the TP53 (rs1042522), butnot MDM2 (rs2279744) polymorphism may confer susceptibility to lung cancer. Cancerincidence rate varies in different regions of our planet, so thehighest incidence of lung cancer is observed in Eastern Europeand Central and East Asia .Lung cancer remains one of the most common forms of cancerin the world. Every year World Health Organization (WHO)includes lung cancer in the lists of the leading cause of deathworldwide. Thus, there were 2.1 million cases of lung cancerand 1.8 million deaths in 2018 (http://www.ensembl.org/).A large number of researches have been conducted tostudy the molecular base of lung cancer. One of the riskfactorsfor the development of pulmonary neoplasms isgenes polymorphisms. The main cause of carcinogenesis isdisordersin the regulation of cell cycle control. The tumorsuppressorgene TP53 plays an important role in regulatingthe cell cycle. p53 protein is known as the \u201cguardian of thegenome\u201d. p53 regulates many genes expression in responseto cellular stress induced by various adverse environmentalfactors . This protein plays a key rolein processes such as DNA repair, cell cycle arrest, apoptosisand senescence . MDM2 is a key regulatorof p53 protein activity and degradation. Polymorphicvariantsof the TP53 and MDM2 genes have been found invarious types of cancer, including lung cancer. Analysis ofthe literature data showed that polymorphisms of the TP53Arg72Pro (rs1042522) and MDM2 SNP309 (rs2279744) genescause an increased predisposition to tumor development.The TP53 (rs1042522) gene polymorphism is localized onchromosome 17 position 7676154 Genotype frequency in theCaucasian population GG: 0.074, CC: 0.503, CG: 0.423. Inthe East Asian population, GG: 0.173, CC: 0.345, CG: 0.482, bladder , colorectalcancer  and acute lymphocytic leukemia. However, no association was found betweenTP53 Arg72Pro and the risk of acute myeloid leukemia, oral squamous cell carcinoma , and esophagus cancer .http://www.ensembl.org/).The MDM2 SNP309polymorphism was also found to increase the risk of colorectalcancer , breast cancer  andliver cancer . But there was no associationwith prostate, urinary tract  and stomach.The polymorphic allele of the MDM2 gene rs2279744 islocated at 68808800 position on chromosome 12 Genotypefrequency in the Caucasian population TT: 0.404, GG: 0.113,GT: 0.483. In the East Asian population TT: 0.200, GG: 0.276,GT: 0.524 and MDM2 SNP309 (rs2279744) on the predisposition tothe development of pulmonary neoplasia. It was shown thatthe polymorphism of the TP53 Arg72Pro gene is associatedwith a high risk of small cell lung cancer among Spaniards. Similar data were found fornon-small cell lung cancer in Norwegians and Poles , squamous cell lung cancer in German residents , and lungadenocarcinoma in the Chinese population .Data on the contribution of MDM2 SNP309 to the developmentof lung cancer are very contradictory. Most studies haveshown an association of the MDM2 (rs2279744) mutant allelewith a high risk of lung tissue carcinogenesis . However, Pine et al.(2006) did not find that MDM2 SNP309 is associated withlung neoplasia in the European population.The data on the association of polymorphisms of the TP53genes Arg72Pro (rs1042522) and MDM2 SNP309 (rs2279744)with the development of tumors as a whole are very contradictory.Therefore, it would be interesting to perform a metaanalysison the association of TP53 Arg72Pro (rs1042522)and MDM2 SNP309 (rs2279744) with a risk of developinglung cancer in Asian and European populations.Search strategy. Search for relevant studies was conductedusing online databases, such as Scopus, PubMed and Web ofScience. The search strategy was performed using a combinationof the following keywords: \u201cTP53\u201d, \u201cMurine double minute2\u201d or \u201cMDM2\u201d, \u201cpolymorphism\u201d, \u201cSNP\u201d, \u201crs1042522\u201d,\u201crs2279744\u201d, \u201cArg72Pro\u201d, \u201ccodon 72 Arg\u201d, \u201cc.215C > G\u201d,\u201cSNP309\u201d, \u201cc.291 T > G\u201d \u201clung cancer\u201d, \u201cnon-small cell lungcancer\u201d, \u201cassociation\u201d.Inclusion and exclusion criteria. The eligible inclusioncriteria for the meta-analysis were (i) case-control study,(ii) identification of different histological types of lungcancer which was confirmed histologically or pathologically,(iii) having an available genotype for estimating anodds ratio (OR) with 95 % confidence interval (95 % CI),(iv) genotype frequencies in controls were consistent withthose expected from Hardy\u2013Weinberg equilibrium ( p > 0.05).The studies were excluded when (i) they were not casecontrolstudies, (ii) with duplicated data from previous articles,(iii) they were not original articles, e. g. review, (iv) inadequategenotype data were available.Data extraction and quality assessment. Two researchers(O.B. and A.K.) evaluated the eligibility of all retrieved studiesand extracted the pertinent data from the specified publicationsin standardized tables. The extracted data included:(i) the first author name, (ii) publication year, (iii) ethnicity,(iv) lung cancer patients and healthy controls sample size foreach studied polymorphism. Disagreement was resolved byconsulting with a third investigator (R.B.). The study qualitywas assessed in accordance with the Newcastle\u2013OttawaScale (NOS) .Statistical analysis. Hardy\u2013Weinberg equilibrium (HWE)in control population was assessed utilizing the \u201cCalculationof Chi-square test for deviation from Hardy\u2013Weinbergequilibrium\u201donline software (http://www.husdyr.kvl.dk/htm/kc/popgen/genetik/applets/kitest.htm). The statistical analysiswas performed using Comprehensive Meta Analysis version2.2.064 . Estimates weresummarized as ORs with 95 % CIs for each study. The heterogeneitywas evaluated by using the I2 index. An I2 valueof > 50 % was considered to indicate high heterogeneity . The random effects model for analysis was used in case high heterogeneity . Otherwise, the fixed-effectsmodel was used. Publication bias was measured via \u201cBegg\u2019sfunnel plot\u201d and \u201cEgger\u2019s linear regression\u201d method . A two-tailed p-value < 0.05 implied a statisticallysignificant publication bias.Studies included in the meta-analysis1.A total of 531 potential articles were identified from the databasessearch. After 236 duplicate records were removed, atotal of 295 potential articles were reviewed. Amongst thesearticles, 216 were excluded after titles and abstracts review.Afterwards, we excluded 28 studies for no case-control design.Finally, 51 studies with a total of 25,239 controls and25,366 cases that met the inclusion criteria were included inthis meta-analysis 1Supplementary Materials are available in the online version of the paper:http://www.bionet.nsc.ru/vogis/download/pict-2020-24/appx13.pdfCharacteristics of studies included in this meta-analysisA total of 37 articles that examined TP53 (rs1042522) associationwith lung cancer risk were determined. Two of thesearticles included data of two different sets (TP53 (rs1042522)and MDM2 (rs2279744))  and these sets were examined autonomously. Thus,the identified 37 articles encompassed case-controls studiesinvolving 16,229 lung cancer patients and 14,897 controls(Table 1). Among 37 articles, 20 studies were establishedin Asian populations and 17 in Caucasian populations. The genotype frequencies in controls of all studies were consistentwith those expected from HWE ( p > 0.05).Another 14 articles identified MDM2 (rs2279744) associationwith increased lung cancer risk were retrieved (Table 2).These 14 articles encompassed case-controls studies involving9,137 lung cancer patients and 10,342 controls. Among14 articles, 7 studies were established in Asian populationsand 7 in Caucasian populations. The genotype frequenciesin controls of all studies were consistent with those expectedfrom HWE ( p > 0.05).All estimated published articles were executed under accreditedgenotyping methods.Meta-analysis of the relationship betweenthe TP53 (rs1042522) polymorphism and lung cancer risksociatedwith lung cancer . And the association was statisticallysignificant under allele model (G versus C), homozygotemodel (GG versus CC)  and dominant model(GG+GC vs. CC)  ( p < 0.05). A summary ofmeta-analysis findings concerning associations between theTP53 (rs1042522) polymorphism and lung cancer risk isshown in Table 3.Further subgroup analysis was conducted on the associationbetween TP53 (rs1042522) polymorphism and the risk of lungcancer (see Table 3). After stratifying by ethnicity, this metaanalysisindicated an obvious association of TP53 (rs1042522)and lung cancer risk among Caucasians  and among Asians .Meta-analysis of the relationship betweenthe MDM2 (rs2279744) polymorphism and lung cancer riskIn this meta-analysis was shown no association MDM2(rs2279744) polymorphism with lung cancer . A summary ofmeta-analysis findings concerning associations between theMDM2 (rs2279744) polymorphism and lung cancer risk isshown in Table 4. Subgroup analysis detected no associationMDM2 (rs2279744) polymorphism with lung cancer.Between-study heterogeneities were found in all subjectsfor both polymorphisms TP53 (rs1042522) and MDM2(rs2279744) . Because of this the meta-analysiswas designed using \u201ca random effect model\u201d to establishpooled OR and corresponding 95 % CI for all models. Weperformed the meta-regression to explore the potential sourceof between-study. A big problem for meta-analysis is the disproportionatenumber of positive studies that leads to a biasin the publication. The funnel plot indicated some evidence ofpublication bias for Caucasians, but not for Asians in analysisof TP53 (rs1042522) and MDM2 (rs2279744) gene polymorphisms. The publication bias was observedfrom Egger\u2019s test ( p \u2264 0.05) also for Caucasian population.The tumor suppressor gene TP53 (previously named p53),is key regulator of a cell cycle network, apoptosis and DNArepair pathway. TP53 is one of the most carcinogenesis-associatedgenes. There were several studies assessing the effectsof TP53 polymorphisms on the risk of lung cancer, but theresults are very contradictory. For example, no associationsof the TP53 (rs1042522) polymorphism with lung cancerwere found in Jung et al.\u2019s (2008) article. But, increased risk to develop lung cancer was observed in association with thePro/Pro genotype variant in Chowdhury et al.\u2019s (2015) research.Mostaid et al. (2014) found that TP53 Arg72Pro andPro72Pro genotype significantly associated with increasedrelative risk of lung cancer. Our previous study also demonstratedthe association of genotype Arg72Pro of TP53 genewith lung cancer risk . Papadakis etal. (2002) demonstrated that subjects with Arg72Arg genotypeof rs1042522 had significantly increased lung cancer risk. Wecomprehensively searched the up-to-date electronic databasesto reveal the associations between TP53 genetic polymorphisms (rs1042522) and risk of lung cancer. The genome-wideassociation study (GWAS) is very popular method to detect avariation in SNPs with variation in common diseases. In 2017,data from a study of new loci of susceptibility to lung cancerwere published. The study identified RNASET2, SECISBP2L,NRG1, CHRNA2, OFBC1 and RTEL1 as candidate genes associatedwith lung cancer . The polymorphismsof TP53 (rs1042522) and MDM2 (rs2279744) weren\u2019tdetected in this GWAS .A total of 37 case-control comparisons for TP53 (rs1042522) were investigated in this meta-analysis. A noteworthy associationof TP53 (rs1042522) with susceptibility to lung cancer inoverall pooled subjects was observed under three differentmodels: the allele contrast, homozygote contrast (additive)and dominant model. Also, stratification analysis explained astrong evidence of this variant with risk of lung cancer amongAsians and Caucasian under allelic, homozygote (only forCaucasian) and dominant models. Moreover, the Arg72Arggenotype was associated with the obvious protective effect.Compared to TP53, whose role has been widely discussed inlung cancer developing, its main negative modifier \u2013 MDM2,has not been sufficiently studied. The data on the associationof polymorphism of MDM2 (rs2279744 or 309T > G) withthe risk of developing lung cancer as well as in the case ofTP53 (rs1042522) are contradictory. Thus, Enokida et al.(2014) did not found any association between polymorphismof MDM2 (rs2279744) and lung cancer risk. Chua et al. (2010)demonstrated that the MDM2 (rs2279744) TT rather than theGG genotype is associated with increased risk of lung cancerin Asian. But, the MDM2 TT genotype was associated witha decreased risk of developing NSCLC compared with thatof the MDM2 GG genotypes in Li G. et al.\u2019s (2006) research.A total of 14 case-control comparisons for MDM2 (rs2279744) wereinvestigated in this meta-analysis. There were no significantassociations between MDM2 (rs2279744) polymorphisms andlung cancer with regard to G allele vs. T allele: OR = 0.86,95 % CI 0.71\u20131.03, p = 0.1; homozygote model: OR = 0.86,95 % CI 0.71\u20131.03, p = 0.1; dominant model: OR = 0.90, 95 %CI 0.79\u20131.02, p = 0.5 and recessive model: OR = 1.10, 95 %CI 0.94\u20131.22, p = 0.276. The stratification analysis also didnot demonstrate the association of this polymorphism withrisk of lung cancer among Asians and Caucasian under allmodels. Thus, MDM2 (rs2279744) polymorphism does notaffect the risk of developing lung cancer.This meta-analysis has some limitations. First, heterogeneitylevel was high. But we tried to eliminate this effect usinga random effects model rather than a fixed effects model.Publication bias could also have biased the results, as studiesthat produced negative results may not have been published.Despite our use of Egger\u2019s regression test, we cannot eliminatethe possibility of bias. Second, the relative importance of theMDM2 (rs2279744) polymorphism during the developmentof lung cancer may vary between ethnic groups, but we wereonly able to perform ethnic-specific meta-analysis in Asiansand Europeans. Thus, our results are applicable to only theseethnic groups. Therefore, additional studies with other ethnicpopulations are warranted to assess the association betweenMDM2 (rs2279744) polymorphism and the risk of lung cancer.But, the present meta-analysis has also several strengths.We used a strong comprehensive search strategy, and had awell-defined inclusion and exclusion criteria. Reviewers performedthe study selection and extracted data independently.Moreover, we assessed the quality of the included studies bypredefined criteria and the score of included studies was high.Finally, all genotype data extracted from the studies werereported in the study. The advantage of this study over othermeta-analyzes is a more complete review of literature and theinclusion of recent data.In summary, this meta-analysis study indicated evidence ofassociation for TP53 (rs1042522), but not MDM2 (rs2279744)variants with lung cancer based on 51 case-control publishedstudies. Additionally, stratified analysis based on ethnicityobserved an obvious association of TP53 (rs1042522) bothamong Asian and European subjects under allelic, homozygoteand dominant models. However, polymorphism MDM2(rs2279744) may not impart susceptibility to lung cancer ineither Asians or Europeans.The authors declare no conflict of interest.Birgander R., Sjalander A., Rannug A., Alexandrie A.K., SundbergM.I., Seidegard J. P53 polymorphisms and haplotypes in lungcancer. Carcinogenesis. 1995;16(9):2233-2236. 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+{"text": "Complete characterization of bis(1H-indazol-1-yl)methane is given with 1H and 13C NMR, UV/Vis, FTIR, high resolution mass spectrometry and for the first time, single crystal X-ray diffraction. This simple, inexpensive pathway to yield exclusively bis(1H-indazol-1-yl)methane provides synthetic access to further investigate the coordination and potential applications of the family of bis(indazolyl)methanes.Synthetic access to poly(indazolyl)methanes has limited their study despite their structural similarity to the highly investigated chelating poly(pyrazolyl)methanes and their potentially important indazole moiety. Herein is presented a high yielding, one-pot synthesis for the 3 UV/Vis \u03bbmax (log \u03b5): 251 (3.54), 260 (3.51), 288 (3.52), 293 (3.52), 299 (3.50). IR (cm\u22121): 3106 (w), 3060 (m), 2957 (w), 2921 (w), 1617 (m), 1499 (m), 1463 (m), 1439 (w), 1353 (m), 1279 (m), 1197 (s), 1150 (w), 1005 (m), 906 (m), 828 (m), 758 (s), 736 (s).Formation of 2.2.2.2.1.H-indazole and 0.31 mmol of trans-dichlorotetrakis(pyridine)cobalt(III) chloride Cl was synthesized from the previous literature Cl. Complete characterization of 1L is presented including 1H and 13C NMR, UV/Vis, FTIR, high resolution mass spectrometry, and for the first time single crystal X-ray diffraction. Creating synthetic access and diffraction data will allow for expansion of our knowledge on the bis(1H-indazol-1-yl)methane family of ligands to be utilized in future synthetic, mechanistic, chelating and pharmaceutical applications.This article presents on a simple, high yielding one-pot synthesis producing exclusively bis(1Supplementary Information"}
+{"text": "Oncogene 10.1038/s41388-021-01973-5, published online 3 August 2021Correction to: In this article two text marking errors on the Fig. 3F and Supplementary Fig. The correct figures are given below.The original article has been corrected.Supplementary Fig. 3"}
+{"text": "Plasmodium parasite fertilization. A recombinant Plasmodium vivax P48/45 (Pvs48/45) protein expressed in Escherichia coli (E. coli) was highly antigenic and immunogenic in experimental animals and elicited specific transmission-blocking (TB) antibodies in a previous pilot study. Here, a similar Pvs48/45 gene was expressed in Chinese Hamster Ovary (CHO) cells and we compared its immunoreactivity with the E. coli product. Specific antibody titers were determined using plasma from Colombian individuals (n=227) living in endemic areas where both P. vivax and P. falciparum are prevalent and from Guatemala (n=54) where P. vivax is highly prevalent. In Colombia, plasma seroprevalence to CHO-rPvs48/45 protein was 46.3%, while for E. coli-rPvs48/45 protein was 36.1% (p<0.001). In Guatemala, the sero prevalence was 24.1% and 14.8% (p<0.001), respectively. Reactivity index (RI) against both proteins showed an age-dependent increase. IgG2 was the predominant subclass and the antibody avidity index evaluated by ELISA ranged between 4-6 mol/L. Ex vivo P. vivax mosquito direct membrane feeding assays (DMFA) performed in presence of study plasmas, displayed significant parasite transmission-blocking (TB), however, there was no direct correlation between antibody titers and oocysts transmission reduction activity (%TRA). Nevertheless, DMFA with CHO rPvs48/45 affinity purified IgG showed a dose response; 90.2% TRA at 100 \u03bcg/mL and 71.8% inhibition at 10 \u03bcg/mL. In conclusion, the CHO-rPvs48/45 protein was more immunoreactive in most of the malaria endemic places studied, and CHO-rPvs48/45 specific IgG showed functional activity, supporting further testing of the protein vaccine potential.P48/45 is a conserved gametocyte antigen involved in  P. falciparum is highly prevalent in sub-Saharan Africa, both P. falciparum and P. vivax coexist in vast regions of Latin America, Asia, and Oceania, with many countries displaying greater P. vivax prevalence  during the fertilization phase in the mosquito midguts  . These antigens have shown the capacity to elicit specific antibodies capable of blocking parasite transmission to mosquitoes and therefore have been identified as potential TB vaccine candidates  , 11. Othndidates . Among tclinical , 14, 15 clinical \u201319.Pvs48/45 is a cysteine-rich conserved protein expressed on the gametocyte surface of several Plasmodium species, and is involved in parasite fertilization  analyses. These antibodies also reduce parasite transmission to mosquitoes in ex-vivo direct DMFA  and Guatemala (GUA). Additionally, TB activity of anti-CHO-rPvs48/45 specific IgG was determined by ex-vivo DMFA.lization . A full-t manner . Additioect DMFA . Despiteproteins , 22, andP. vivax and P. falciparum are endemic  at the Centro Internacional de Vacunas  in June 2017 (code CECIV 1506-2017) as well as by the Ethics Committee of the GUA Ministry of Health (CNES-dq-005-2015). Additional approval was obtained in GUA from local community leaders before data collection. Written informed consent (IC) was obtained from each volunteer at enrollment. Minors with ages between 7-18 years signed an informed assent (IA) form, and parents or legal guardians gave the corresponding consent. Donors of P. vivax or P. falciparum infections as determined microscopically and confirmed by qPCR. Whole blood samples (5-15mL) were collected in heparin vacutainer tubes by arm venipuncture, and plasma immediately separated by centrifugation. In GUA, 54 patients with ages between 1-70 years of age (38.9% men) with P. vivax infections diagnosed by thick blood smears and confirmed by qPCR, were recruited by active case surveillance, and blood samples were collected as described above. Plasma samples were kept and stored at -80\u00b0C until used for serological characterization and for specific CHO-rPvs48/45 antibody affinity purification.In COL, 227 patients, men (56.8%) and women between 6-84 years of age, were recruited by passive surveillance and included in the study. Subjects were randomly selected among a larger group of patients harboring rPvs48/45 gene was expressed in E. coli, as described before  with sufficient viable cell culture biomass of suspension adapted CHO-Express\u2122 cells . All production cultures (post-transfection) were performed in serum-free, animal protein-free medium (low protein content). A non-malaria related protein, expressed in parallel during the production of the CHO-rPvs48/45 protein, was used as a control production to ensure cell culture and methods accuracy. Each production vessel was verified for critical parameters such as cell culture dynamics and recombinant protein production on samples from day 7 and/or day 14, to confirm the appropriate protein cultures. A total of 150 mg of this antigen was produced after a single purification step on IMAC-FPLC. Two buffer exchanges were performed: one to optimize binding of the protein construct to the IMAC column, and the other one, after elution, to remove excess imidazole. SDS-PAGE analysis of both E. coli- and CHO-rPvs48/45 proteins under both reducing (0.05 mol/L dithiothreitol) and non-reducing conditions together with immunoblot and mass spectrometry (LC-MS/MS) confirmed the protein identity. Immunoblot analysis was carried out using sera (1:200 dilution) of P. vivax semi-immune subjects, plasma from Aotus monkeys previously immunized with the E. coli-rPvs48/45 protein (1:200 dilution), and a E. coli-rPvs48/45 monoclonal antibody; normal human serum was used as control and LC/MS/MS analysis was performed using peptides generated by trypsin digestion. Peptides were separated and analyzed using a Qexactive-HFX coupled to U3000 RSLC (Thermofisher). Mass spectra were processed with MaxQuant (v1.6.10.43) using a database consisting of Uniprot entries of Cricetulus griseus , 250 classical contaminants and the sequence of the construction.The CHO-s\u2122 cells . BrieflyrPvs48/45 proteins\u2019 immunoreactivity, as described before , pH 7.4 at 4\u00b0C overnight. After plates were blocked with milk solution 5%, plasma samples diluted at 1:200 were added and incubated for 1 hour followed by incubation with alkaline phosphatase (1:1000) conjugated goat anti-human IgG antibody  for 1 hour. Reactions were revealed with para-nitrophenyl phosphate substrate (p-NPP) (Sigma Aldrich) and read at 405 nm wavelength . Cut-off points for ELISA were calculated as three SD above the mean absorbance value at 405 nm of sera from healthy adult volunteers who had never been exposed to malaria (n=60). The results were also expressed as reactivity index (RI) defined as OD values of tested samples divided by the cut-off value. P-value < 0.05 was considered significant.ELISA was used to determine the two d before , 26. BriP. vivax CHO-rPvs48/45 antibodies with ELISA reactive index > 5.0, as described elsewhere , followed by anti-CHO-rPvs48/45-specific antibodies purification using an NHS-activated Sepharose column  coupled with CHO-rPvs48/45 protein, as described previously. Affinity adsorption was performed using the purified total IgG, and elution fraction was neutralized by 1 mol/L Tris, pH 9.0. Specific IgG was then dialyzed against 1x PBS using an Amicon Ultra-4 Centrifugal Filter Unit (30 kDa membrane EMD Millipore). The final protein concentration of specific IgG was measured by DS-11FX+  and adjusted to 0.5 mg/mL (rPvs48/45-specific antibodies (Elisa Units - EU) as described before  of the CHO-rPvs48/45 recombinant protein competitor. The reaction was incubated for 30\u00a0min at RT and the reactivity of the antibody preparation was determined by ELISA as previously described  displaying ELISA reactivity indices (RI) > 5.0 and four plasma sera from the same regions with RI between 2.0-3.0  were selected to determine antibody avidity. Polystyrene microplates were coated with 1 \u03bcg/mL of the CHO-rPvs48/45 protein in duplicates. After overnight incubation plates were washed and blocked with 100 \u03bcL of PBS with 0.5% Tween (PBS-T) per well, 5% skim milk  for 2\u00a0h at room temperature (RT). Specific IgG at 10 \u03bcg/mL or plasma samples diluted at 1:200 in PBS-T 2.5% skim milk was added to the plates and incubated for 2h at RT. After washing, plates were incubated with different urea concentrations (from 0 to 7 mol/L) in duplicates for 15\u00a0min to determine antibody avidity. The reaction was revealed using an anti-human IgG alkaline phosphatase conjugate as described above. The urea concentration resulting in 50% of the original ELISA units (IC50) was calculated using linear regression  and anti-IgG2 . Then a peroxidase-conjugated goat anti-mouse IgG antibody  was added, and the reaction read at 450 nm.The lsewhere . BrieflyrPvs48/45 specific IgG antibodies at 10, 40, and 100 \u03bcg/mL was measured by a DMFA using P. vivax gametocyte-infected blood obtained from malaria patients, as previously described  were subsequently reconstituted with a pool of heat-inactivated male AB+\u00a0sera obtained from healthy donors purchased from Sigma (H5667 Sigma-Aldrich Inc) or with test samples. A total of ~100\u00a0Anopheles\u00a0albimanus\u00a0mosquitoes previously subjected to overnight fasting were fed for 15-20 minutes/cage with this mixture. Mosquito midguts were stained with 2% mercurochrome on day 7 post-feeding, and the number of oocysts per mosquito midgut counted.The TB activity of 143 plasma samples (1:2 dilution), and affinity-purified CHO-escribed . Brieflyp-values <0.05 were considered significant. Antibody responses  for both CHO-rPvs48/45 and E. coli- were compared by Fisher\u2019s exact test. Differences of RI between age groups were analyzed by Kruskal-Wallis, followed by Dunn\u2019s multiple comparison test. Correlation between %TRA of human sera and RI by Spearman test. Avidity (IC50) of P. vivax plasma samples and affinity-purified CHO-rPvs48/45 specific IgG were compared by Mann-Whitney tests , and ey tests .p-value for TRA was calculated using a zero-inflated negative binomial model as previously described (The percentage reduction in mean oocyst count/mosquitoes (TRA) was calculated using the following formula: [(Xc \u2212 Xa)/Xc)] \u00d7 100, where X is the arithmetic mean oocysts in control (c) and test (a) plasma or IgG. The 95% confidence interval and escribed .E. coli-rPvs48/45 gene without the signal peptide and GPI anchor was sub-cloned in the E. coli system as described elsewhere  inserted to\u00a0avoid inclusion bodies formation (rPvs48/45 has a shorter thioredoxin fragment (42aa), and a theoretical mass of 51,766.22  of samples and in 32% (n=90) to the E. coli-rPvs48/45 product (p<0.001) . In COL plasmas (n=227) the prevalence of specific antibodies was 46.3% and 36.1% to the CHO and E. coli products, respectively. In contrast, in GUA plasmas (n=54) it was 24.1% and 14.8%, respectively, indicating significant regional differences for both proteins (CHO and E. coli p<0.001). Also, samples from Tierralta, (COL), where P. vivax is highly prevalent , presented an immunoreactivity of 58.8% and 34.3% against the CHO and E. coli proteins, respectively. In contrast, samples from Tumaco, where P. falciparum is more prevalent  than P. vivax , ELISA showed 27.3% and 41.7%, respectively. Significant differences in prevalence among both proteins in samples from COL  and GUA samples (p<0.001) was observed. In contrast, in Tumaco (COL) E. coli was better recognized than CHO (p<0.001) (E. coli protein (p<0.001). In GUA, people with > 2.0 RI were 8% (n=1) and 0% (n=0) to CHO and E. coli protein, respectively. Fisher tests indicated that plasma samples with RI > 2 were significantly more frequent in Colombia than in GUA (p=0.0042) to CHO protein. In contrast, there was no difference between both countries to E. coli protein. To determine the impact of age on the RI, in both COL and GUA, samples were stratified into three groups: 0-14 years/old n=41 (14.6%); 15-30 year/old n=118 (42%); and more than >30 years/old, n=122 (43.4%). The mean RI presented an age-dependent increasing trend (rPvs48/45 protein (p<0.05). Individuals >30 years/old presented a RI to CHO mean of 1.45 whereas to E. coli, the mean was 1.13. Significant differences (p<0.05) were observed in the RI values between children (<15 years/old) and adults (>30) when the E. coli - rPvs48/45 protein was used . Regarding TRA activity, 93 of the 143 (65.03%) plasma samples examined by DMFA displayed >80% TRA; 51samples (54.83%) were reactive to the CHO- and 29 (31.18%) reactive to E. coli-rPvs48/45 protein (p>0.28 and E. coli p>0.63).From the total plasma samples studied (n= 281), ELISA assays indicated the presence of specific antibodies to the full-length CHO-p<0.001) Table\u00a01.Pvs48/45 protein. A total of 40 mg/mL of total IgG was obtained after protein G column purification and 500ug/mL of anti-CHO-rPvs48/45-specific antibodies after Sepharose column purification with specific antibody titers of 62.163 EU  (E.\u00a0coli-rPvs48/45 proteins (n=22)  using the CHO r2.163 EU . SpecifiE. coli-rPvs48/45 specific IgG and sera samples from malaria-endemic regions were incubated with different concentrations of urea. All data sets fitted into the linear regression model (R2>0.94), and the anti-rPvs48/45 specific IgG presented a similar IC50 (4.1 and 4.5 mol/L respectively) with plasma samples with RI between 2-4  and a slightly higher avidity with the HPV-3031 (5.5 mol/L), P-137 (5.6 mol/L), and P-72 (4.9 mol/L) of plasma samples with RI>5.0, except with the sample MPV-139 (3.9 mol/L)  in studies conducted in several countries in Africa where this species is almost exclusively transmitted and malaria transmission intensity is significantly higher than in Latin America  than in GUA  in the year 2017 . In this regard, in COL, immunoreactivity was higher and more prevalent in areas with the relatively higher transmission, e.g., in Tierralta , Department of Cordoba  the conformational differences between the 3D structures of the two proteins and, thus, recognition of different epitopes, 2) by the additional thioredoxin fragment added to the E. coli protein to improve its expression, 3) by potential glycosylation in the CHO protein.Although both proteins were highly immunoreactive, CHO was significantly better recognized by all age groups, except in Tumaco (COL) where Pvs48/45and nor the additional fragment are recognized by control sera.None of the constructs were recognized by sera from na\u00efve individuals indicating that neither full-length P. falciparum is significantly higher that P. vivax, yet, the response to rPvs48/45 was similar than in Tierralta (COL) and GUA suggesting a parasite cross-reactivity between the orthologous proteins in natural conditions. To better understand the sero-epidemiology of the rPvs48/45, we are currently studying malaria-endemic regions with different P. vivax/P. falciparum proportions.Another intriguing observation is that in Tumaco (COL) the prevalence of rPvs48/45 antibodies which confirmed the important role of this parasite antigen in TB immunity under natural conditions  and a low (~25kDa) molecular weight in the  pattern , 41. HerrPvs48/45 activity is independent of the classical complement; this issue requires further analyses.Although several IgG subclasses may contribute to the overall TB activity, the high prevalence of IgG2 suggests that anti-Pfs48/45 vaccine candidate  at the Centro Internacional de Vacunas  (code CECIV 1506-2017) Ethics Committee of the GUA Ministry of Health (CNES-dq-005-2015). Written informed consent to participate in this study was provided by the participants\u2019 legal guardian/next of kin.rPvs48/45 analysis: AK and GC. Contributed reagents/materials/analysis tools: MA-H, NC, and SH. Wrote the paper: MA-H, NC, GC, CL, and AK. All authors contributed to the article and approved the submitted version.Conceived and designed the experiments: MA-H and SH. Performed the experiments: NC, CE, KM, ES, AC, JR, and AM. Analyzed the data: MA-H, SH, GC, and CL. LC/MS/MS CHO-NIH/NIAID 1R01AI121237-01 sponsored this study and in part by the Intramural Research Program of NIAID, NIH.https://www.frontiersin.org/articles/10.3389/fimmu.2021.634738/full#supplementary-materialThe Supplementary Material for this article can be found online at: Click here for additional data file.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Pickering S, Batra R, Merrick B, et al. Comparative performance of SARS-CoV-2 lateral flow antigen tests and association with detection of infectious virus in clinical specimens: a single-centre laboratory evaluation study. Lancet Microbe 2021; published online June 30. https://doi.org/10.1016/ S2666-5247(21)00143-9\u2014This Article should have been published under a CC BY Open Access license. This correction has been made as of Aug 3, 2021."}
+{"text": "Mutant alleles of the Rht-B1 and Rht-D1 (Reduced height) genes are widely used in bread wheat breeding for the development of intensive-type cultivars. These genes and their f lanking regions have been sequenced and the point mutations leading to the nonsense codons  and various insertions  associated with a change in plant height have been described. DNA-markers based on the allele-specif ic PCR have been developed to identify single-nucleotide changes. However, the use of such technique imposes stringent PCR conditions, and the resulting data are not always unambiguous. An alternative can be found in the CAPS technology: it detects differences in sequences by digesting PCR products. In the absence of restrictases capable of digesting DNA at the point mutation site, restriction sites can be introduced into the primer sequence (derived CAPS). The aim of this study was to propose a system of CAPS-, dCAPS- and STS-markers for identifying alleles of the reduced height genes frequently used in breeding programs. Three CAPS have been developed to identify the Rht-B1b, Rht-D1b, Rht-B1p alleles, as well as two dCAPS for Rht-B1b, Rht-B1e. STS-markers for the insertion-containing alleles Rht-B1c, Rht-B1h and Rht-B1i-1 have been selected from publications. The proposed markers were tested during the genotyping of 11 bread wheat accessions from the VIR collection with the abovementioned mutant alleles and the wild-type Rht-B1a and Rht-D1a. The presence of nonsense mutations was also conf irmed by the resultsof allele-specif ic PCR. This marker system, along with the existing ones, can be used to identify dwarf ing alleles ofthe Rht-B1 and Rht-D1 genes in bread wheat for genetic screening of accessions from ex situ collections and/or formarker-assisted selection. The development of intensive-type short-stemmed wheat cultivarsis considered one of the key success factors in breadwheat breeding, primarily in implementing the Green Revolutioninitiative in the world\u2019s developing countries . The decrease in plant height notonly entailed higher resistance to lodging, with its favorableeffect on the efficiency of mechanized harvesting, but alsoincreased the number of grains per ear and their number per1 m2, which aggregately led to higher yields .At least 25 genes controlling plant height in bread wheat(Triticum aestivum L.) and related species were described: theyare known as Reduced height \u2013 Rht1\u2013Rht25. All these genesare in one way or another associated with the growth hormonegibberellin . Some of them,the so-called GA-sensitive genes Rht4\u2013Rht9, Rht12\u2013Rht20 andRht25, are apparently involved in the synthesis or degradationof gibberellic acid (GA). Other genes, GA-insensitive ones,such as Rht-A1, Rht-B1, and Rht-D1, determine the responseto this acid. For some genes , thenature of their response has not yet been clarified .The most widespread among GA-sensitive genes is Rht8,transferred in the early 20th century, together with the closelylinked photoperiod insensitivity allele Ppd-D1a of the Ppdgene (response to photoperiod ), from the Japanese cultivarAkakomugi first to Italian and later to many East and SouthEuropean cultivars . Thisgene does not exert any significant reducing effect on thecoleoptile length and, as a consequence, makes it possibleto sow seeds to a greater depth, which plays a decisive rolein maintaining the viability of seedlings under water deficitsor high temperatures .GA-insensitive genes were studied in more detail; they arelocated on the short arms of chromosomes of homeologousgroup 4 . Dominantalleles of these genes (wild-type) encode DELLA proteins,belonging to the family of GRAS proteins (transcription regulators);at their C-terminus, there is a conservative domainthat can bind to other transcription factors and thereby blocktheir function. That is why large amounts of DELLA proteinsin cells decelerate plant growth. There is a DELLA domain atthe variable N-terminus: it is capable of forming the GA\u2013GID1complex . Thiscomplex undergoes polyubiquitination and degradation inducedby proteasomes. Accordingly, a decrease in the amountof DELLA proteins in cells in the presence of GA reduces theirnegative effect on plant growth .A fairly large number of recessive and semi-dominant mutantalleles altering the stem length in different ways havebeen described for the Rht-B1 and Rht-D1 genes. These alleleshave been sequenced; the most thoroughly studied sequencesare presented by us in Supplementary material 11. The allelesRht- B1b (=Rht1), Rht-B1e ,Rht-B1p (=Rht17) and Rht-D1b (=Rht2) were shown to beassociated with single-nucleotide substitutions that lead tothe formation of premature stop codons . The phenotypic effectof such nonsense mutations varies from moderate  to strong  .http://vavilov.elpub.ru/jour/manager/files/Suppl_Porotnikov_Engl.pdfSupplementary materials 1 and 2 are available in the online version of the paper:The alleles Rht-B1h and Rht-B1i-1 have large (over 100 bp)insertions in the 5\u2032 flanking region, while Rht-B1c (=Rht3)is characterized by the presence of an insertion in the 5\u2032 untranslatedregion identical to that in Rht-B1h and, at the sametime, the presence of the Veju retrotransposon in the codingregion . Such insertions can lead to the formationof nondegradable proteins, so the growth of mutant plants isconstitutively repressed,more significantly than in the caseof nonsense mutations in the N-terminal coding region . For example, the Rht-B1c allelereduces plant height approximately by 60 % . However, insertions can not onlyreduce but also increase the height of plants  as, for example, in the case ofRht-B1i-1 . Besides, the alleles of \u201cstrongdwarfing\u201d, Rht-D1c (Rht10) and Rht-D1d (Rht Ai-bian 1a),reducing the height by 60\u201370 %, were identified in the Rht-D1gene; they turned out to be multiple copies of the mutant alleleRht-D1b . There are also other known allelesof the Rht-B1h\u2013o and Rht-D1e\u2013j genes, associated witheither nucleotide changes (missense mutations) or indels. Theyare identified in a large number of Chinese cultivars using theEcoTILLING method; however, their phenotypic effect hasnot yet been described. Mutant alleles of the Rht-A1 genewere also identified for the first time in Chinese cultivars .The alleles most frequently used in breeding programs areRht-B1b and Rht-D1b. Their source was the Japanese cultivarNorin 10. At the end of the 20th century, more than 70 % ofthe world\u2019s bread wheat cultivars contained these alleles . Later, however, it was shown thattheir occurrence depended on the region of the world. Rht-B1b was detected in 36.2 % of bread wheat cultivars from China,and Rht-D1b in 53.4 % . Meanwhile,the genotyping of 247 cultivars from the United States andCanada helped to identify these alleles in more than 90 %of them . Rht-D1b predominates in thegenotypes of European cultivars, while its occurrence in thecultivars registered after 1990 is 49 % .Widespread in Russia are cultivars with the Rht-B1e allele,obtained by mutagenesis in cv. Bezostaya 1; the mutantform is Krasnodarsky Karlik 1 . At present, semi-dwarf cultivars , homozygous for Rht-B1e alleles,are cultivated both in Russia and the ex-USSR countries onan area of more than 4 million hectares .The allele Rht-B1p is also promising for breeding: a stopcodon emerges in its DELLA domain due to the substitutionof cytosine for thymine at position 178 from the start codon.This mutation causes an up to 30 cm decrease in the heightof bread wheat plants, especially as far as the lower internodeis concerned, but it does not reduce the length of the ear .The sequencing of Rht-B1 and Rht-D1 alleles in variousbread wheat cultivars have led to the development of molecularmarkers for their identification. For example, STS markerswere obtained to identify the insertion-containing allelesRht-B1c, Rht-B1h and Rht-B1i-1 . Markers based on allele-specific PCR(AS-PCR), including real-time AS-PCR, are used to identifythe alleles Rht-B1b, Rht-B1e, Rht-B1p and Rht-D1b, carryingsingle-nucleotide substitutions .The widespread alleles Rht-B1b and Rht-D1b as well asthose of the Rht24 gene are identified on the basis of competitiveallele-specific PCR (KASP-markers), offering a possibilityto evaluate large numbers of bread wheat accessions atlow time costs .It should also be mentioned that AS-PCR results strongly dependon the reaction conditions, require several replicationsof the analysis, and call for strict observance of the author\u2019sprotocol, which is not always possible. The KASP analysis,in its turn, requires sophisticated equipment and expensivereagents, which are often unaffordable to small practice-orientedlaboratories.The use of CAPS (cleaved amplified polymorphic sequence)markers can be an alternative to AS-PCR: they are based onthe presence of a restriction site in the region with a singlenucleotidemutation (the site is absent in the wild type) or,contrariwise, on the disappearance of the site typical of thewild type in the mutant version . If restrictionsites are absent at the locations of the analyzed mutations,they can be produced purposefully through designing modifiedprimers, i. e., by the derived CAPS method, or dCAPS .Unlike AS-PCR, the CAPS and dCAPS marker techniquesare effortlessly reproducible and do not require stringentPCR conditions, while the results of such analysis are easilyinterpreted in agarose gels. It is possible to generate markersusing the basic PCR equipment. Previously, such markers weredeveloped for the Rht24 dwarfing gene .The objective of the present study was to develop CAPS anddCAPS markers for the analysis of single-nucleotide changesin Rht-B1 and Rht-D1, test STS markers for identification ofinsertions in these genes and, as a result, propose a markersystem for identifying the alleles most frequently used inbread wheat breeding.Plant material. Eleven bread wheat accessions from the VIRcollection with known alleles of the Rht-B1 and Rht- D1 dwarfinggenes (Table 1) served as the material for this study. Thecultivars Chinese Spring and Hongdongmai with wild-typealleles Rht-B1a and Rht-D1a were used as controls. Eachof the studied accessions was represented in the genotypingprocess by two or three individual plants as well as by bulkDNA sample, which was isolated from a total of 10\u201320 genotypes(seedlings).DNA extraction. DNA was extracted from 10-day-oldseedlings using a modified CTAB extraction technique .Sequences alignment. The sequences of different allelesof the Rht-B1 and Rht-D1 genes were aligned using MEGA X(https://www.megasoftware.net/), Unipro UGENE , and BioEdit Sequence Alignment Editor. Restriction sites were searched for using theGenScript Restriction Enzyme Map Analysis Tools .Primers development. Primers for the nested PCR andCAPS analysis were developed with the Primer3Plus software. Primer quality  was monitored using OligoAnalyzerTool, a web resource from Integrated DNA Technologies, Inc.. Primers for dCAPSmarkers were generated using the dCAPS Finder 2.0 software. The primers developed in the courseof this study and those supplied from published sources arepresented in Tables 2 and 3, and their locations are shown inFig. 1, a, b.PCR procedure: a) nested PCR. The nested PCR methodwas applied to enhance the specificity of the dCAPS analysis:the first PCR was performed with primers BF/VIR.B1R flankingthe region of point mutations in the Rht-B1 gene; afterthat, the resulting PCR product was used as a template for thesecond PCR with dCAPS  and CAPS(B1epF/B1pR) primers. The first round of nested PCR wascarried out in 25 \u03bcl of the reaction mixture containing 40 ngof total wheat DNA; 1\u00d7 reaction buffer; 1.5 mM of MgCl2;0.6 mM of each dNTP; 0.25 \u03bcM of both forward and reverseprimer, and 1 unit of Taq DNA polymerase . For higher specificity, the PCR programcontained the Touchdown function: the initial annealing temperaturewas 4 degrees higher than the design temperature anddecreased by 0.5 degrees per cycle for 8 cycles (see Table 3).Samples of the resulting amplification products (2 \u03bcl ofeach) were transferred into clean tubes, diluted 50 times withwater, and used as a template in the second stage of PCR.Another 10 \u03bcL of each PCR product was taken to control the success of PCR by agarose gel electrophoresis .The remainder (approximately 12 \u03bcl) was treated with therestriction enzyme BstV1I  to generate the CAPS marker for theRht- B1b allele.The second round of nested PCR was performed in 20 \u03bclof the reaction mixture containing 4 \u03bcl of the template; 1\u00d7reaction buffer; 2.5 mM of MgCl2; 0.3 mM of each dNTP;0.25 \u03bcM of both forward and reverse primer, and 1 unit of TaqDNA polymerase . The programs for each pair of primersare also presented in Table 3. Approximately 12 \u03bcl of theamplification mixture were taken for restriction analysis, andthe remainder was used for PCR control by electrophoresis;b) standard PCR. In the cases of CAPS markers for theRht-D1b allele and the markers detecting retrotransposonin the gene\u2019s coding region and insertions in the 5\u2032 flankingregion, PCR was performed under standard conditions. Thereaction mixture (20 \u03bcl) contained 40 ng of DNA; 1\u00d7 reactionbuffer; 2.5 mM of MgCl2; 0.3 mM of each dNTP; 0.25 \u03bcMof each primer, and 1 unit of Taq DNA polymerase ;the programs are presented in Tables 2 and 3;\u0441) allele-specific PCR. The conditions and the programs forAS-PCR corresponded to those recommended by the authorsof the primers .Restriction analysis. PCR products were treated with restrictionenzymes produced by SibEnzyme, using the manufacturer\u2019sprotocol (http://russia.sibenzyme.com).Fragment separation was done in horizontal agarosegels in the 1\u00d7 TBE buffer under the voltage of 5 V/cm. Thegels were stained with ethidium bromide and visualized inUV light.For the development of CAPS and dCAPS markers, thesequences from the NSBI databases were analyzed (https://www.ncbi.nlm.nih.gov/) for the following alleles of the dwarfinggenes: Rht-B1b, Rht-B1e, Rht-B1p and Rht-D1b. Also,the sequences of the wild-type alleles Rht-A1a, Rht-B1a andRht-D1a were retrieved as controls. The Genbank accessionnumbers for used sequences are given in Supplementary material1. Sequence alignment confirmed the presence of nonsensemutations in these allelic forms, which made it possible to startthe development of CAPS and dCAPS markers .A search was made for each nonsense mutation to identifyrestriction sites that would distinguish the target allele fromall others, including wild-type ones. The BstV1I (GCAGC)restriction enzyme, unable to digest the mutant GTAGC site,was selected for Rht-B1b. Similarly, BstHHI (GCGC) becamethe restriction enzyme for the Rht-B1p detection (mutantsite GCGT). On the contrary, the BstSFI restriction enzyme(CTRYAG) exclusively digested the mutant site (CTGTAG)contained in Rht-D1b. Thus, it was possible to develop suchCAPS markers as CB1b/BstV1I, CB1p/BstHHI and CD1b/BstSFI to identify the alleles Rht-B1b, Rht-B1p and Rht-D1b,respectively.We failed to identify restriction sites at the location of thenonsense mutation in the Rht-B1e allele. Hence, the dCAPSmarker dCB1e/HinfI was developed for it: the sequence of thereverse primer was modified so that the analyzed nucleotide,together with the 3\u2032 end of the primer, formed a GATTC restrictionsite, providing an opportunity to distinguish this mutationfrom all other alleles by means of the HinfI restriction. ThedCAPS marker dCB1b/Acc36I was additionally constructedto identify Rht-B1b .When performing PCR under standard conditions, with thegenomic DNA of bread wheat used as a template, we wereunable to obtain specific fragments for the dCAPS markers andthe CAPS marker CB1p/BstHHI (the data are not presented).We therefore applied the nested PCR method: the amplificationproducts of the BF/VIR.B1R primers, flanking the regionof localization of all analyzed point mutations in the Rht-B1gene, were used as a template for the second round .The developed markers were tested on a set of bread wheataccessions with known alleles of the dwarfing genes, and all of them demonstrated high efficiency in differentiating thewild-type Rht-B1a, Rht-D1a and mutant versions Rht-B1b,Rht-B1e, Rht-B1p and Rht-D1b .Concurrently, allele-specific primers retrieved from publishedsources were used to identify nonsense mutations inRht-B1b, Rht-B1e, Rht-B1p and Rht-D1b compared to thewild type . For this purpose, two pairs of primers were usedfor identification of each mutation: one of them detected themutant version, while the other spotted the wild type andall other alleles. It was shown for Rht-B1b, Rht-B1e andRht-D1b that the results of allele-specific PCR on the wholeagreed with the data of CAPS and dCAPS analyses. However,identification of the wild-type Rht-B1a and Rht-D1a alleleswith the primers BF/WR and DF2/WR2, respectively , involved certain difficulties: poor reproducibilityof results, and generation of weakly expressed fragments informs with Rht-B1b and/or Rht-D1b . In the case of Rht-B1p, allele-specific PCR under theconditions of this study turned out to be ineffective: afteramplification with the Rht-B1p-F/R1 primers , a specific product was generated both in the formswith mutant alleles and in those with the wild-type ones .The study also employed five pairs of STS primers  as a tool for identifyingmutations associated with the presence of a retrotransposonin the coding region (Rht-B1c) as well as with insertionsin the promoter region (Rht-B1i-1) and the 5\u2032 flanking region(Rht-B1h). The locations of these insertions are marked in thescheme of the Rht-B1 gene; it also shows primers for theirdetection .Two pairs of primers were used for the Rht-B1i-1 allele : one of them (B1i-MF1/MR1) in the presence ofan insertion produced a specific 330 bp fragment, while theother (B1i-MF2/MR2) amplified fragments of different sizesin genotypes with or without an insertion . Similarly, to detect the Rht-B1h allele, the Rht-B1h.F/R1 primers were used, resulting in a specific productof 247 bp, as well as the Rht-B1h.F/R2 primers, generatingfragments of different sizes  . Since Rht-B1h has a common insertion in the5\u2032 flanking region with Rht-B1c, the Rht-B1c-F1/R1 primer,specific for the retrotransposon sequence, was also used fortheir differentiation  . Additional evidence of the presence of a retrotransposonmay be found in the fact that no PCR productsare generated in genotypes with this insertion during the firstround of nested PCR with the BF/VIR.B1R primers: it can beexplained by a big distance between the primers .Our testing of STS markers showed a complete concordancebetween the presence of their diagnostic fragments and thecomposition of alleles present in the studied genotypes . The accessions Atlas 66 and Zheng 9023 containingRht-B1h yielded amplification products pointing to the presenceof an insertion in the 5\u2032 flanking region. In the accession Triumph carrying the Rht-B1i-1 allele, which increasesplant height, an insertion in the promoter region was detectedusing molecular markers, and in Tom Thumb (Rht-B1c), aretrotransposon in the coding sequence and an insertion inthe 5\u2032 flanking region were found. It should be mentionedthat when a retrotransposon was identified using the Rht-B1c-F1/ R1 primers, in addition to the formation of a fragment ofthe expected size, the emergence of nonspecific products ofa larger size was observed in Tom Thumb (Rht-B1c) and inall other genotypes .Assessing the system of the proposed molecular markersin its entirety, it should be kept in mind that it can be used togenerate a marker profile for each of the studied alleles of theRht-B1 and Rht-D1 genes, i. e., to get an unambiguous answerwhether one of the abovementioned alleles of the Rht-B1 andRht-D1 dwarfing genes is present in one or another genotype.Marker profiles for the alleles are presented in Table 4.As a result of this study, a system of molecular markers wasproposed for the Rht-B1 and Rht-D1 dwarfing genes to identifythe alleles most often used in bread wheat breeding. Thesystem is based on the developed CAPS and dCAPS markersof nonsense mutations in these genes, which were previouslydetected by allele-specific PCR . Five STS markersretrieved from published sources were used to identify insertions.The CAPS and dCAPS markers were tested during the genotypingof bread wheat accessions from the VIR collection,containing the mutant Rht-B1b, Rht-B1c, Rht-B1e, Rht-B1h,Rht-B1i-1, Rht-B1p and Rht-D1b alleles as well as those ofthe wild-type. The tests showed complete concordance ofthe obtained results with the expected ones. The presenceof Rht- B1b, Rht-B1e, Rht-B1p and Rht-D1b was also confirmedby allele-specific PCR with the primers widely usedin research and breeding programs .The main advantage of our molecular marker system liesin good reproducibility of results and their unambiguous interpretation.The CASP/dCAPS analysis faces no problemswith controlling the PCR reaction success, because amplificationproducts are formed in all genotypes, and differencesbetween alleles are pinpointed after treatment with restrictionenzymes. Besides, notwithstanding the high cost of restrictionenzymes, CASP/dCAPS analysis is less expensive, sincethere is no need to perform two independent PCRs in severalreplications to detect each allele. The procedure is conductedemploying standard PCR equipment and using agarose gel electrophoresis, so it can be carried out by small practiceorientedlaboratories. When any new point mutations in theRht-B1 and Rht-D1 dwarfing genes become known, a similarapproach to the development of CAPS/dCAPS markers canbe applied to identify them.The authors declare no conflict of interest.Antonova O.Yu., Klimenko N.S., Rybakov D.A., Fomina N.A., ZheltovaV.V., Novikova L.Yu., Gavrilenko T.A. SSR analysis of modernrussian potato varieties using DNA samples of nomenclatural standards.Biotekhnologiya i Selektsiya Rasteniy = Plant Biotechnologyand Breeding. 2020;3(4):77-96. DOI 10.30901/2658-6266-2020-4- o2. (in Russian)Bazhenov M.S., Divashuk M.G., Amagai Y., Watanabe N., Karlov G.I.Isolation of the dwarfing Rht-B1p (Rht17) gene from wheat and thedevelopment of an allele-specific PCR marker. Mol. Breed. 2015;35(11):1-8. DOI 10.1007/s11032-015-0407-1.Bazhenov M.S., Nazarova L.A., Chernook A.G., Divashuk M.G. Improvedmarker for the Rht-B1p dwarfing allele in wheat. 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Molecularmapping of Rht8 on the short arm of chromosome 2D of bread wheat(Triticum aestivum L.). Theor. Appl. Genet. 1998;96(8):1104-1109.DOI 10.1007/s001220050845.Kurkiev K.U., Tyryshkin L.G., Kolesova M.A., Kurkiev U.K. Identificationof Rht2 and Rht8 genes for semidwarfness in hexaploidtriticale with use of DNA markers. Vavilovskii Zhurnal Genetikii Selektsii = Vavilov Journal of Genetics and Breeding. 2008;12(3):372-377. (in Russian)Li A., Yang W., Guo X., Liu D., Sun J., Zhang A. Isolation of a gibberellin-insensitive dwarfing gene, Rht-B1e, and development ofan allele-specific PCR marker. Mol. Breed. 2012;30(3):1443-1451.DOI 10.1007/s11032-012-9730-y.Li A., Yang W., Lou X., Liu D., Sun J., Guo X., Wang J., Li Y., Zhan K.,Ling H.Q., Zhang A. Novel natural allelic variations at the Rht-1loci in wheat. J. Integr. Plant Biol. 2013;55(11):1026-1037. DOI10.1111/jipb.12103Lou X., Li X., Li A., Pu M., Shoaib M., Liu D., Sun J., Zhang A.,Yang W. 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+{"text": "Novel coronavirus 2019 (Covid19) caused by SARS-CoV2 can lead to significant morbidity and mortality. There is unclear association between Covid19 and bacteremia. Patient characteristics and outcomes are not well defined. This retrospective cohort study assessed this in patients with Covid19 and bacteremia.Patients with Covid-19 admitted to a tertiary care suburban academic medical center (UH) were assessed retrospectively by EMR chart review for co-morbidities, pre and in hospital factors, and outcomes as defined below. Bacteremias grouped into gram-negative or gram-positive with collation of each unique bacterial species (Table 1).Total 1398 patients with Covid19 hospitalized at UH during local peak of pandemic of whom 238 (17.02%) developed 264 bacteremias with gram-positive  and gram-negative organisms . Relevant characteristics (Table 2) 53% with immunomodulator therapy (steroids/Tocilizumab), mean length of stay 21.04days (SEM \u00b1 1.67) with day SARS-CoV2 PCR positivity -1.15days from hospitalization (SEM \u00b1 0.49) and day initial bacteremia 6.38 (SEM \u00b1 0.77), 55.4% required ICU admission, with 89% ICU admissions requiring mechanical ventilation. Most common co-morbidity  Hypertension 56.3% followed by Obesity (BMI >30) 45.8% and CAD/CHF 40.3%. Laboratory parameters (Table 3)significant for average difference (date bacteremia- date admission) for Procalcitonin 4.15ng/mL , CRP -0.934mg/dL , WBC 7.027 K/uL . These analyses excluded difference of 0 from hospital day 1 bacteremia. Average antibiotic number (1+ dose per antibiotic) 3.24 (SEM \u00b1 0.16) and total C difficile cases 3 (1.26%). Mortality rate34.45%.Relevant hospitalization characteristics, Covid19 and bacteremia co-infectionsRelevant laboratory parameters for patients with Covid19 and bacteremia co-infectionPatients with Covid19 and bacteremia had high mortality , 53% received immunomodulator therapy, possibly contributing to bacteremia development. With bacteremia increase in WBC and Procalcitonin, not CRP, noted. Most organisms CoNS, likely contaminants, gram positive bacteremias likely from indwelling lines. Only 3 C difficile infections identified. Trends noted in Procalcitonin rise, immunomodulator therapy, and low C difficile infection rates warrant further studies.Post-hospitalization OutcomesAll Authors: No reported disclosures"}
+{"text": "Staphylococcus aureus (MRSA), but there are limited patient outcomes in the setting of cystic fibrosis pulmonary exacerbation (CFPE). The study objective was to compare the efficacy and safety of TLV to vancomycin (VAN) in CFPE.Telavancin (TLV) is an advanced generation lipoglycopeptide with activity against methicillin-resistant Retrospective cohort conducted from 1/2011-6/2020. Inclusion criteria were: i) age \u226516 years, ii) hospitalized for CFPE with documented signs/symptoms of infection, iii) confirmed or suspected MRSA lower respiratory tract infection, iv) receipt of \u226548 hours of TLV or VAN. The primary outcome was 30-day CFPE-related readmission: infection recurrence, clinical worsening on treatment, or ADE requiring readmission. Secondary outcomes included adverse drug events (ADE) on therapy: acute kidney injury (AKI), rash, thrombocytopenias, cardiac abnormalities. P< 0.001). The median (IQR) time to TLV initiation from admission was 1 (0.8-1.4) days. 13 patients were readmitted within 30-days due to CFPE; 8 (15)% TLV vs. 5 (10%) VAN . Reasons for 30-day CFPE: TLV: 7 infection recurrence, 1 clinical worsening; VAN: 2 clinical worsening, 2 infection recurrence, 1 treatment-related ADE. When accounting for confounders, TLV was not associated with 30-day CFPE-related readmission . Patients who received VAN more commonly experienced an ADE while hospitalized (18%), most notably AKI101 patients were included: 52 (52%) TLV, 49 (49%) VAN. The median (IQR) age was 22 (21-27) years, 50% were women, and 86% were Caucasian. The majority (84%) of patients had some federal health insurance; 19% had private health insurance. 93% of patients used a maintenance cystic fibrosis (CF) medication, and 35% had previous CF-therapy compliance concerns. 62% had a previous positive culture for MRSA; 22 (43%) TLV patients had documented MRSA infection on admission compared to 41 (84%) VAN  ghassan wadi, MD, Cumberland (Grant/Research Support) Michael P. Veve, Pharm.D., Cumberland (Grant/Research Support)Paratek Pharmaceuticals (Research Grant or Support)"}
+{"text": "Yang LiuBJU Int. 2020 Jul;126(1):168-176.Percutaneous nephrolithotomy (PCNL) is the gold standard surgical treatment for large renal kidney stones >2.0 cm) according to EUA and AUA guidelines , 2. Howe.0 cm accSeveral authors have already reported favorable outcomes of mini-PCNL when compared to RIRS -6. A sim"}
+{"text": "Cited2 deletion has a minor impact on steady-state hematopoiesis, Cited2-deficient HSCs are severely depleted in young mice and fail to expand upon aging. Moreover, although they home normally to the bone marrow, they fail to reconstitute hematopoiesis upon transplantation. Mechanistically, CITED2 is required for expression of key HSC regulators, including GATA2, MCL-1, and PTEN. Hematopoietic-specific expression of anti-apoptotic MCL-1 partially rescues the Cited2-deficient HSC pool and restores their reconstitution potential. To interrogate the Cited2\u2192Pten pathway in HSCs, we generated Cited2;Pten compound heterozygous mice, which had a decreased number of HSCs that failed to reconstitute the HSC compartment. In addition, CITED2 represses multiple pathways whose elevated activity causes HSC exhaustion. Thus, CITED2 promotes pathways necessary for HSC maintenance and suppresses those detrimental to HSC integrity.Hematopoietic stem cells (HSCs) reside at the apex of the hematopoietic differentiation hierarchy and sustain multilineage hematopoiesis. Here, we show that the transcriptional regulator CITED2 is essential for life-long HSC maintenance. While hematopoietic-specific  \u2022Unperturbed hematopoiesis can be sustained long term while the HSC pool is depleted\u2022CITED2 promotes HSC survival but not quiescence under homeostatic conditions\u2022Mcl1 and Pten expressionCITED2 maintains the HSC pool by controlling  Cited2 deletion causes a progressive HSC loss under steady-state conditions without perturbing normal hematopoiesis. The authors show that CITED2 maintains HSCs by regulating the expression of Mcl1 and Pten. Finally, they indicate that CITED2 promotes multiple pathways necessary for HSC maintenance and suppresses those detrimental to HSC integrity to coordinate HSC function.Kranc, Guitart, and colleagues demonstrate that  Hematop humans) . The strCITED2 (CBP/p300-interacting-transactivator-with-an ED-rich-tail 2) is a transcriptional regulator that co-activates or represses multiple transcription factors, including AP-2 , HIF-1alMx1-Cre-mediated deletion of Cited2 (in which Mx1-Cre is induced by poly(I:C)-stimulated IFN-\u03b1 production) results in a rapid loss of HSCs via apoptosis and a resultant BM failure  or Trp53 (encoding p53) rescues depletion of Cited2-deficient HSCs. Another study -inducible Mx1-Cre-mediated Cited2 deletion results in loss of HSCs , compromises their reconstitution potential, and leads to a rapid BM failure upon myelotoxic stress. In this study, loss of quiescence, but not increased apoptosis, upon inducible Cited2 deletion is mediated at least in part by HIF-1alpha, as Hif1a deletion partially restores impaired quiescence of HSCs lacking Cited2 and improves their ability to reconstitute the HSC compartment upon transplantation  cells, LSKCD48\u2212CD150+ HSCs, Lin\u2212Sca-1\u2212c-Kit+ (LK) myeloid progenitors, and more-mature Lin\u2013 and differentiated Lin+ hematopoietic cell populations, and performed qRT-PCR. Cited2 was expressed in all compartments, with significantly higher expression in the HSC population compared with myeloid progenitors and more mature hematopoietic cell populations (Cited2 in long-term HSCs (LSKCD34\u2212CD135\u2212), MPP1 (LSKCD34+CD135\u2212CD150+CD48\u2212), MPP2 (LSKCD34+CD135\u2212CD150+CD48+), and MPP3 (LSKCD34+CD135\u2212CD150\u2212CD48+) populations, lymphoid-primed multipotent progenitors (LSKCD34+CD135+), which correspond to the MPP4 population, and CMP (LKCD34+Fc\u03b3RII/IIIlow), GMP (LKCD34+Fc\u03b3RII/IIIhigh), and MEP (LKCD34\u2212Fc\u03b3RII/IIIlow) compartments, we analyzed our SMART2-seq single-cell expression data in these populations  mice  analyses of 8- to 12-week-old Cited2CKO mice revealed unaffected WBC counts, with mild anemia and thrombocythemia  mice D, where CKO mice E. Surpricythemia F. Furthelularity G and unalularity H, as wellularity  and 1I.  numbers J. Notabl numbers F, mice lgenitors K. FinallBM cells L and 1M.or cells M. Taken Cited2 deletion on HSCs and primitive progenitor cells. We found that the total number of LSK cells  was not affected in young 8- to 12-week-old Cited2CKO mice , with an increase in MPP2 population and unchanged numbers of the MPP3-4 populations. Thus, despite a select reduction in absolute HSC and MPP1 cell numbers, Cited2CKO mice sustain largely unaffected unperturbed steady-state multilineage hematopoiesis.We next determined the impact of CKO mice A. MarkedCited2 deficiency on long-term HSC maintenance during unperturbed hematopoiesis. We aged mouse cohorts for 52\u00a0weeks and found that Cited2 deficiency had no impact on mouse survival (data not shown). Consistent with physiological HSC aging, during which the HSC pool undergoes expansion  and found that they also dramatically failed to reconstitute hematopoiesis  resulted in a partial rescue of the HSC and MPP1 cell pools compared with Cited2CKO mice  is a pro-survival BCL-2 protein family member, whose  failure , thus redeletion . Moreovedeletion . Given tnic mice , which oelements B. IntereCKO mice C. To invent mice D. Signifcl1 mice E and 4F.cl1 mice E and 4F.cipients G. Finallcolonies H, and whol cells I. TherefPten expression was decreased in Cited2-deficient HSCs , Cited2Het, PtenHet, and control mice  did not rescue HSC depletion -inducible ectively . Howevertor Mcl1 , whose hotential . Consistntenance . These dactivity . Indeed,Mx1-Cre deletion of Cited2 results in loss of quiescence of HSCs, a phenotype that is partially mediated by HIF-1alpha  is known to transiently alter HSCs , unlike Vav-iCre, which is constitutively expressed, and as such more accurately allows for gene deletion under steady-state conditions. Thus, it is possible that concurrent poly(I:C) administration and Cited2 deletion in the Mx1-Cre-mediated model may exhibit exacerbated phenotypes not seen in the Vav-iCre model.Poly(I:C)-inducible F-1alpha , 2014. NWhile in this study we focused on functional interrogation of the CITED2\u2192MCL-1 and CITED2\u2192PTEN axes, our work suggests that CITED2 is also likely to control other diverse pathways to coordinate HSC biology. Our analyses indicate that CITED2 represses multiple pathways downstream of c-MYC, E2F, and RUNX1 transcription factors and K-RAS and proinflammatory signaling pathways, whose upregulation has detrimental consequences for HSC integrity . FinallyCited2fl/fl , anti-CD4; biotin conjugated (cat. no. 553649), anti-CD5; biotin conjugated (cat. no. 553019), anti-CD8a; biotin conjugated (cat. no. 553029), anti-CD11b; biotin conjugated (cat. no. 553309), anti-CD45R/B220; biotin conjugated (cat. no. 553086), anti-Ter119; biotin conjugated (cat. no. 553672), anti-Gr-1/Ly-6G/C; biotin conjugated (cat. no. 553125), anti-CD34; FITC conjugated (cat. no. 553733), and streptavidin; BV421 conjugated (cat. no. 563259). BioLegend antibodies used were anti-c-Kit/CD117; APC conjugated (cat. no. 105812), anti-c-Kit/CD117; APC-Cy7 conjugated (cat. no. 105826), anti-Sca-1; APC-Cy7 conjugated (cat. no. 122520), anti-Sca-1; PB conjugated (cat. no. 108125), anti-CD48; PE conjugated (cat. no. 103406), anti-CD150; PE-Cy7 conjugated (cat. no. 115914), anti-CD135; APC conjugated (cat. no. 135310), anti-CD135; PE conjugated (cat. no. 135305), anti-CD16/32; APC-Cy7 conjugated (cat. no. 101328), anti-CD41; APC conjugated (cat. no. 133914), anti-CD105; PE conjugated (cat. no. 120408), anti-CD127; BV421 conjugated (cat. no. 135023), anti-TER-119; FITC conjugated (cat. no. 116206), streptavidin; PerCp conjugated (cat. no. 405213), anti-CD19; APC-Cy7 conjugated (cat. no. 115530), anti-CD45R/B220; APCCy7 conjugated (cat. no. 103224), anti-CD11b; APC conjugated (cat. no. 101211), anti-Gr-1/Ly-6G/C; PE-Cy7 conjugated (cat. no. 108416), anti-CD4; PE conjugated (cat. no. 130310), anti-CD8a; PE conjugated (cat. no. 100708), anti-CD45.1; FITC conjugated (cat. no. 110706), anti-CD45.2; PB conjugated (cat. no. 109820), anti-Ki67; FITC conjugated (cat. no. 652410). Annexin V; FITC conjugated (cat. no. 640906), and 7-AAD (cat. no. 420403) were purchased from BioLegend. DAPI was purchased from Life Technologies (cat. no. D1306).Mice received 30\u00a0mg/mL of NAC (Sigma) in drinking water for 4\u00a0weeks. The water bottles containing NAC were changed twice a week.6 BM cells stained first for LSK were resuspended in X-Vivo 15 medium (without phenol red) supplemented with 10% FCS, loaded with 5\u00a0\u03bcM MitoSox red (Invitrogen) for 20\u00a0min at 37\u00b0C and analyzed using FACS.For detection of mitochondrial super oxide, 3\u00a0\u00d7 10CFC assays were performed using MethoCult M3434 (STEMCELL Technologies). Colonies were tallied at day 10. For CFC replating, CFC1 cells were washed with IMDM then seeded in M3434.+/CD45.2+ recipient mice was achieved using a split dose of 11\u00a0Gy (two doses of 5.5\u00a0Gy administered 4\u00a0h apart) at an average rate of 0.58 Gy/min using a Cesium 137 GammaCell 40 irradiator. For transplantations 100 HSCs or 2,000 LSK sorted from BM of 8- to 10-week old adult mice mixed with 200,000 support CD45.1+ wild-type BM cells were injected into lethally irradiated CD45.1+/CD45.2+-recipient mice.Lethal irradiation of CD45.1+ BM were injected into CD45.1+ lethally irradiated recipients . After 18 h, recipients were sacrificed and BM CD45.2+ chimerism was analyzed by FACS.LSK cells sorted from CD45.2Actb expression.Gene expression analyses were performed as described previously . DiffereStatistical analyses were performed using GraphPad Prism software. p values were calculated using a Mann-Whitney U test.Cited2CKO and Cited2CTL animals (n\u00a0= 5 per genotype). On average, 6,200 cells per sample were collected and 53.7 million single-ended 85-bp reads per sample were sequenced. Reads were aligned to the GRCm38 mouse genome using HISAT2 v.2.1.0  were mapped to gene loci using the basic set of ENCODE genomic annotation from Ensembl v.91. Next, proximal promoters were defined as the region from 200\u00a0bp upstream to 100\u00a0bp downstream of the transcription start site. Proximal promoter coordinates were shuffled within chromosomes using the bedtools shuffle tool with the \u2013chrom flag to generate control DNA regions for motif analysis. Finally, DNA motifs overrepresented in promoters compared with control regions were identified by Homer v.4.10.in\u00a0vivo experiments, and data analyses and interpretation. L.N.L. and M.B. analyzed gene expression. K.J.C. and S.C. produced VavP-Mcl1 transgenic mice. A.T., J.D., A.V., A.B., C.M., E.G., C.M.-C., C.S., L.A., and J.C. helped with in\u00a0vivo experiments, FACS, and data analyses. D.O., B.G., and N.P.R. provided significant expertise to this study. K.R.K. and A.V.G. contributed equally to this work.K.R.K., A.V.G., and H.L. designed the experiments and wrote the paper. A.V.G. and H.L. performed The authors declare no competing interests."}
+{"text": "A terrible disease of the cardiovascular system, atherosclerosis, develops in the areas of bends andbranches of arteries, where the direction and modulus of the blood flow velocity vector change, and consequentlyso does the mechanical effect on endothelial cells in contact with the blood flow. The review focuses on topicalresearch studies on the development of atherosclerosis \u2013 mechanobiochemical events that transform the proatherogenicmechanical stimulus of blood flow \u2013 low and low/oscillatory arterial wall shear stress in the chains of biochemicalreactions in endothelial cells, leading to the expression of specific proteins that cause the progressionof the pathological process. The stages of atherogenesis, systemic risk factors for atherogenesis and its importanthemodynamic factor, low and low/oscillatory wall shear stress exerted by blood flow on the endothelial cells liningthe arterial walls, have been described. The interactions of cell adhesion molecules responsible for the developmentof atherosclerosis under low and low/oscillating shear stress conditions have been demonstrated. The activationof the regulator of the expression of cell adhesion molecules, the transcription factor NF-\u03baB, and the factorsregulating its activation under these conditions have been described. Mechanosensitive signaling pathways leadingto the expression of NF-\u03baB in endothelial cells have been described. Studies of the mechanobiochemical signalingpathways and interactions involved in the progression of atherosclerosis provide valuable information for thedevelopment of approaches that delay or block the development of this disease.Key words: atherogenesis; shear stress; transcription factor NF-\u03baB; RelA expression; mechanosensitive receptors;cell adhesion molecules; signaling pathways; mechanotransduction. Nowadays, cardiovascular disease is a major public healthissue. Moreover, atherosclerosis is one of the most commonpathologies of the cardiovascular system. The systemic riskfactors for the development of atherosclerosis include age,hypertension, diabetes mellitus, smoking, low physical activity,fatty diet, renal failure, increased level of fibrinogen, lowdensitylipoproteins, cholesterol and blood plasma C-reactiveprotein . The level of low-density lipoproteins(LDLs) is classified into a separate group of factorsthat account for the atherogenicity of the subfraction profileof apo-B-containing lipoproteins . The penetration of blood plasma LDLs throughthe endothelium in athero-susceptible areas of the arteries andtheir retention and accumulation in the extracellular matrix(ECM) of the subendothelial space initiates atherogenesis.LDLs are retained in the intima (mainly due to interaction withproteoglycans), undergo oxidation (formation of oxLDLs) andcause an inflammatory response \u2013 the infiltration of circulatingblood monocytes into the intima. In the intima, monocytesdifferentiate into macrophages, uptake oxLDLs and becomefoam cells .The development of atherosclerosis occurs in the followingstages: (i) adaptive intimal thickening, (ii) formation of fattystreaks, (iii) pathological intimal thickening (PIT), (iv) earlyfibroatheroma and (v) late fibroatheroma. In stage (i), smoothmuscle cells (SMCs) of the media migrate to the intima and secreteproteoglycans. The formation of fatty streaks in stage (ii)is accompanied by the accumulation of foamy cells (macrophagesloaded with lipids) in the intima. Lipid-loaded SMCsare less represented. The PIT process (iii) occurs with andwithout the infiltration of macrophages. In both cases, SMCsand extracellular lipid pools are present in the intima. The accumulationof SMCs occurs towards the lumen of the artery,and lipid pools accumulate close to the media. The formationof a fibrous cap that covers the necrotic core occurs in the laterstages of development of the atherosclerotic lesions, includingearly and late fibroatheroma . The cap includes SMCs,infiltrated macrophages, T-lymphocytes, as well as collagensand proteoglycans of the extracellular matrix. Programmedcell death, via apoptosis and necroptosis, plays an essentialrole in early fibroatheroma (iv) with the formation of foci ofnecrosis and cholesterol crystals. In late fibroatheroma (v),an extensive necrotic core that consists of cellular debris anda large number of crystals of free cholesterol and its esters isformed .The molecular and genetic processes of atherogenesis remainunclear. Numerous haemodynamic studies have shownthat, based on systemic risk factors, atherosclerosis developsmainly in the bends and branching of the arteries, where thereis a change in the nature of the blood flow . The haemodynamiccharacteristics of the effect of blood flow on the vessel wallsare wall shear stress (WSS), hydrostatic pressure and cyclicdeformation. WSS is the friction force that occurs whenflowing blood comes into contact with the inner wall of the artery. WSS on the arterial wall is described by :where \u03bc is the blood viscosity index, \u2192V(t) is the blood flowrate parallel to the vessel wall at time t and r is the radial coor-dinate.Studies conducted on animal models and observing patientsrevealed a regular maximum thickening of the intima and theformation of atherosclerotic plaques in areas with low WSS(< 10 dyn/cm2 in humans) and low/oscillatory WSS (with adeviation of the instantaneous WSS vector from its averagedirection). Such damage was minimal in areas of high WSS(>25 dyn/cm2 in humans). High values of WSS were realizedin the rectilinear sections of the arteries with laminar bloodflow. In the areas of branching and bending of the arteries nearthe walls, vortex flows were formed, leading to mechanicalstress on the walls, which was accompanied by pathologicaleffects. These flows were characterised by low and low/oscillatoryWSS .A study conducted on isolated segments of blood vesselsthrough which LDLs flowed demonstrated that the transport ofLDLs into the vascular wall increased with a decrease in WSSand, on the contrary, decreased with an increase in WSS . Patient-specific modelling of the subendothelialaccumulation of LDLs in the stenotic right coronary arteryalso showed an inverse relationship between the distribution ofWSS and the accumulation of LDLs .The zone of recirculating flow and low WSS correspondedto the maximum accumulation of LDLs, and in areas of highWSS, the accumulation of LDLs was low.To localise the segments of arteries with low and low/oscillatoryWSS and monitor the transformation of atheroscleroticplaques into a stable or unstable phenotype, computer modellingof blood flow in the vessels is being developed. Thiswill facilitate the identification of patient-specific fields andgradients of blood flow rates depending on the geometry ofthe vessels . To solve these issues, the Navier\u2013Stokesequations for an incompressible viscous fluid are used. Toreconstruct the geometric shape of the vessels, intravascularultrasound methods are used as well as X-ray microcomputertomography with the use of contrast agents . The ANSYS Fluent, OpenFOAM,FLUENT 6.0 and other software packages are widely used toconduct calculations via mathematical models of stationaryand unsteady blood flows in various areas of the arteries. Computermodelling of the distribution of WSS, which accountsfor patient-specific data on the geometry of blood vessels, isof high value for clinical practice.The molecules of cell adhesionand their interactions in the early stageof atherogenesisOxLDLs in the subendothelial space as well as low and low/oscillatory WSS cause pro-inflammatory activation of endothelialcells (ECs), which leads to the rolling of leukocytes inthe circulating blood flow to the endothelium, their adhesion and transendothelial migration (TEM). The mediators of thesecritical events of the early stage of atherogenesis are cell adhesionmolecules. These molecules are expressed on the surfaceof ECs and circulating blood cells (monocytes or leukocytesand platelets) and include platelet endothelial cell adhesionmolecule-1 (PECAM-1); intercellular adhesion molecule-1and -2 ; vascular cell adhesion molecule-1(VCAM-1); E-, L- and P-selectins; vascular endothelial (VE)cadherin; \u03b21 and \u03b22 integrins; proline-rich glycoprotein CD99and junctional adhesion molecule-A (JAM-A).Selectins (transmembrane glycoproteins) are expressed onthe surface of ECs , leukocytes(L-selectin) and platelets (P-selectin) . Early experiments conducted in a flow chamber with alaminar flow of monocytes on an EC monolayer  showed that the rolling of the monocytes to theECs, weak, reversible contact of the monocytes with ECs and slowing down of the rate along theendothelium determine the interactions of the L-selectin ofthe monocytes with glycoprotein ligands of the ECs and, to alesser extent, the P-selectin of the ECs with the glycoproteinligand PSGL-1 of the monocytes. E-selectin is not involvedin the process . Firm, irreversibleadhesion of the leukocytes to the ECs occurs during theinteraction of the leukocyte \u03b14\u03b21 (VLA-4) integrin with theendothelial immunoglobulin VCAM-1  and the interaction of the leukocyte\u03b1L\u03b22  integrinswith the endothelial immunoglobulin ICAM-1 .OxLDLs and lysophosphatidylcholine (a component ofoxLDLs) induce the expression of ICAM-1 and VCAM-1 onthe surface of cultured ECs and stimulate monocyte adhesion. A physiologicallylow WSS, created by the flow of leukocytes  on the EC monolayer,generates upward docking structures on the endothelium thatcontain ICAM-1 and VCAM-1 clusters within 1\u20132 minutes.These clusters surround the leukocytes and function as ananchor for them .In turn, the structures of VCAM-1 and ICAM-1 that surroundthe leukocytes stimulate the formation of lateral linear tracksof leukocyte \u03b21 (VLA-4) and \u03b22 (LFA-1) integrins that areoriented parallel to the ICAM-1 and VCAM-1 clusters . Moreover, 90% of the leukocytes thatare surrounded by the VCAM-1 and ICAM-1 clusters transmigrateto the subendothelial space, and the suppression ofVCAM-1 and ICAM-1 via inhibitors  significantly suppresses TEM. Regardless of theTEM pathway , the TEM process isassociated with the formation of a cupped traction structureby the VCAM-1/VLA-4 and ICAM-1/LFA-1 interactions ofendothelial and leukocyte cells, which guide and facilitateTEM .VE-cadherin plays an important role in the TEM of leukocytes.VE-cadherin is only expressed in ECs, localised mainlyin the intercellular contacts and plays an important role in the intercellular adhesion and barrier functions of the ECs . The adhesion of leukocytes to the endothelium inthe lateral intercellular contacts, preceding TEM, induces theformation of gaps in the intercellular distribution of VE-cadherinand the components of the VE-cadherin complex  at the sites of adhesion or transmigrationof the leukocytes. The gaps are formed as a result of lateraldisplacement of VE-cadherin in the membrane and facilitatethe TEM of the leukocytes. Following the completion of TEM,VE-cadherin moves in the opposite direction and closes thegaps (curtain opening and closing effect) . The lateral displacement of VE-cadherinin the membrane most likely occurs due to the destabilisationof the VE-cadherin bond with the actin cytoskeleton by thefollowing mechanism: the ICAM-1 and VCAM-1 clusters inthe sites of intercellular adhesion of the leukocytes to the ECsinduce the intracellular activation of the Src and Pyk2 tyrosinekinases and the phosphorylation of Tyr658 and Tyr731 of thecytoplasmic domain of VE-cadherin, which are involved in thelow-affinity binding of VE-cadherin to p120- and \u03b2-catenin,respectively. The weakening of these interactions disrupts theVE-cadherin bond with the actin cytoskeleton, destabilises theVE-cadherin or VE-cadherin cell-cell interactions and facilitatesthe lateral movement of the phosphorylated VE-cadherinin the membrane . p120-catenin regulatesthe phosphorylation of VE-cadherin and the paracellularTEM of leukocytes via a competition mechanism with theactivated Src and Pyk2 tyrosine kinases: the overexpressionof p120-catenin in the ECs leads to the absence of gaps in thedistribution of VE-cadherin and the blocking of the TEM ofthe leukocytes .The glycoprotein PECAM-1 (CD31) plays an importantrole in the TEM of leukocytes. In ECs, PECAM-1 is mainlylocalised in the intercellular contacts. The homophilic interactionsof this protein with adjacent ECs occur through theextracellular Ig-like domains IgD1 and IgD2 . PECAM-1 is also present in a distinct membranevesicularrecycling compartment adjacent to the lateral bordermembrane of the ECs . In the restingendothelium (in the absence of adhesive leukocytes), there isconstitutive membrane traffic between the lateral cell borderand the membrane-vesicular compartment  . In the presence of adhesive leukocytes onthe endothelium, directed kinesin-dependent migration of thePECAM-1-bearing LBRC membrane along the microtubulesof the ECs to the sites of para- and transmigration of the leukocytesoccurs, in addition to the surrounding of the leukocytesby the LBRC membrane. The LBRC provides non-ligatedPECAM-1 and CD99 in the ECs to interact with the homophilicligands  in the leukocytesand initiates the signals for further recruitment of theLBRC as the leukocytes move through the endothelial layer.The antibodies to PECAM-1 and CD99 block the TEM of theleukocytes . The recruitment ofthe LBRC to paracellular TEM sites precedes the formationof gaps in the intercellular distribution of VE-cadherin andis necessary for the formation of these gaps . In ApoE\u2013/\u2013 PECAM-1\u2013/\u2013 mice, the load of plaques inthe areas of the carotid artery with a low and low/oscillatory WSS was significantly less than in the control ApoE\u2013/\u2013 mice. A study on the relationship of singlenucleotide polymorphisms  in functionally important domainsof PECAM-1 in patients who were at risk of developingcoronary heart disease and myocardial infarction showed thatArg670Gly substitution can be a homozygous protector forthe development of myocardial infarction. This substitution islocalised close to Tyr663, the phosphorylation of which, underlow WSS conditions, initiates the signaling pathway of activationof the key transcription factor NF-\u03baB for the expressionof cell adhesion molecules . Val125Leuand Asn563Ser substitutions are not associated with the riskof coronary heart disease .Integrins also play an important role in cell adhesion. Theintegrins are a large family of receptors that are localised inthe plasma membrane and consist of 18 \u03b1 and 8 \u03b2 subunitsthat form 24 different heterodimers. The extracellular domainsof the integrins interact with ECM proteins  and ligands on the surface of other typesof cells, causing cell-substratum or cell-cell adhesion. Theintegrin-ligand interactions induce the activation of a variety ofsignaling pathways that modulate cellular behaviour, includingproliferation, shape, motility, survival or apoptosis, differentiation,protein phosphorylation, cytoskeleton organisation andgene expression. Many integrins are expressed in an inactivestate on the cell surface since the membrane-proximal highlyconserved sequences of the cytoplasmic domains of the \u03b1and \u03b2 subunits form a structural constraint that locks the conformationof integrins in an inactive, low-affinity state. Theactivation of integrins is often induced by intracellular signalsand regulatory factors that act on the cytoplasmic domains, aswell as phosphorylation. This alters the affinity of integrins forligands through conformational changes in their extracellulardomains, as well as clustering .The adhesion of leukocytes to the endothelium and theirinfiltration into the subendothelial space is enhanced bycytokines, chemokines and other factors. Thus, monocyticchemotactic protein-1 (MCP-1)  and interleukin-8 (IL-8) (producedby macrophages) are involved in the delay of peripheralcirculation monocytes and their adhesion and migration intothe arterial intima via interaction with monocyte receptors belongingto the CCR2 and CXCR2 types, respectively . IL-9, secreted mainly byCD45+CD3\u2013CD19\u2013 leukocytes in \u0410\u043f\u043e\u0415\u2013/\u2013 mice, stimulates theexpression of VCAM-1 in the ECs of the aorta of mice throughinteraction with the IL-9 receptor (IL-9R) and activation(phosphorylation) of the signaling protein and transcriptionactivator STAT3 . In the atheroscleroticaortas of \u0410\u043f\u043e\u0415\u2013/\u2013 mice, a high level of IL-17A expressionwas observed, as well as numerous IL-17A-producing CD4+T helper 17 (Th17) and \u03b3\u03b4+ T cells. IL-17A initiates the productionof several cytokines and chemokines by aortic cells,in particular, the pro-inflammatory chemokine CXCL1, whichactivates peripheral circulation monocytes and stimulates theiradhesion and migration to the aortic wall .Pro-atherogenic IL-17C, expressed mainly by aortic SMCsin \u0410\u043f\u043e\u0415\u2013/\u2013 mice, is involved in the recruitment of T cells and macrophages into the aortic wall .TGF-\u03b2-(H2O2-) inducible clone 5 (His-5), expressed on thesurface of ECs and SMCs, is involved in the formation ofstructures that are similar to the microvilli on the surface ofECs, which enhance the adhesion of monocytes to the ECs.Transcription factor NF-\u03baB is a key regulatorof the gene expression of cell adhesion moleculesunder conditions of physiologicallylow and low/oscillatory wall shear stressThe transcription factor NF-\u03baB positively regulates the expressionof cell adhesion molecules with the participation ofother transcription factors and coactivators. The VCAM-1 genepromoter has two NF-\u03baB sites that are required for transcriptionactivation . The C/EBP and NF-\u03baBsites were identified in the ICAM-1 gene promoter, and mutationsin the latter completely suppress the activation of theICAM-1 promoter . The ELAM-1 genepromoter includes a CRE/ATF site, three NF-\u03baB sites and threeHMGI(Y) sites; two of the HMG I(Y) sites are located withinthe NF-\u03baB sites (the interaction of HMG I(Y) and NF-\u03baBwith small and large DNA grooves, respectively). All threeNF-\u03baB sites are required for promoter activation and enhancethe affinity of NF-\u03baB and ATF-2 for the promoter . The promoter of the MCP-1 gene (monocytechemoattractant protein-1) contains the NF-\u03baB site and theAP-1 site, which are necessary for maximum induction of thepromoter . The core element GAGACC(SSRE) was identified in the pro-atherogenic platelet growthfactor (PDGF) promoter that stimulates the proliferation andmigration of SMCs ; it was found tointeract with NF-\u03baB .In ECs, the most common p50/p65 (RelA) heterodimer isNF-\u03baB. It is well known that, in the cytoplasm, latent NF-\u03baBis associated with an inhibitor of I\u03baB (mainly I\u03baB\u03b1) and isinactive. When ECs are stimulated by cytokines TNF\u03b1, IL-1 orbacterial lipopolysaccharide (LPS) that interact with specificEC receptors, signaling pathways are activated, leading to theactivation of the I\u03baB kinase (IKK complex), which specificallyphosphorylates I\u03baB. After the ubiquitination of I\u03baB and itsproteasomal degradation, free NF-\u03baB is translocated into cellnuclei (factor activation), interacts with elements of the DNAmajor groove and activates the expression of genes involved inthe immune response, cell survival, carcinogenesis and inflammation(with the participation of other transcription factorsand coactivators) .Early studies demonstrated that active NF-\u03baB and ICAM-1were present in ECs, macrophages and SMCs in atheroscleroticplaques of arteries of deceased patients but were absentin the intima or media of the healthy arteries of those patients. OxLDL triggers in the ECs in vitro andin vivo signaling pathways for IKK complex activation; thisincludes the activation of focal adhesion kinase (FAK) andribosomal S6 kinase (RSK) and leads to NF-\u03baB activation,VCAM-1 expression and monocyte adhesion to ECs . Using the model flow channel system  that is created by a laminar pulsating flow of theculture medium containing monocytes, in which an averagephysiologically low and uniformly distributed WSS acts on the EC monolayer, it was shown that a mechanical stimulus activates the IKK complex andNF-\u03baB and stimulates VCAM-1 expression and monocyteadhesion to the ECs . A model spatialgradient of averaged low/oscillatory WSS, close to the gradientof WSS in arterial branchings and bends, resulted in amore efficient activation of NF-\u03baB compared to uniformlydistributed low WSS .High activation of NF-\u03baB was observed in ECs that werecultivated based on a model of the calculated WSS profile in asite of the human carotid sinus  compared with a model of the calculatedWSS profile in the distal segment of the carotid arterybifurcation  orcells at rest . A study of the transcriptionalexpression of the VCAM-1 endothelium from the nature ofhaemodynamics and calculated profiles of WSS in the segmentsof the left and right coronary arteries showed that low/oscillatory WSS at the outer wall of the bifurcation of theleft anterior descending artery (atherogenic region) inducesa significantly higher expression of VCAM-1 compared to astraight region of the right artery , inwhich atherosclerotic plaques are largely unformed . The mapping of the endothelial expression ofNF-\u03baB/I\u03baB and the activation of NF-\u03baB in areas of the aorticarch of mice with high  and low  probability of atherosclerotic plaque formationdemonstrated that in the HP region with low/oscillatory WSS, the expression of NF-\u03baB/I\u03baB and the activationof NF-\u03baB significantly exceeded those in the LP region. S. Cuhlmann et al. (2011) also detectedincreased expression of the RelA subunit NF-\u03baB and a highernuclear localisation compared with the LP region in the HPregion of the aortic arch of mice. Moreover, this was correlatedwith increased expression of VCAM-1 and the accumulationof CD68+ macrophages in the HP regionThe gap-junction protein connexin 40 (Cx40), expressedin ECs under low WSS conditions, whose cytoplasmic C-terminusinteracts with I\u03baB\u03b1 and inhibits its phosphorylation,is a negative regulator of NF-\u03baB activation in areas with lowWSS . Conversely, mechanosensitive phosphatidicacid phosphatase (PPAP2B) regulates NF-\u03baB activity,expression of adhesion proteins, monocyte adhesion and TEMunder high WSS. PPAP2B shows increased expression inECs under atheroprotective flow characteristics  and shows decreased expression under atherogeniccharacteristics . PPAP2Bhydrolyses lysophosphatidic acid (LPA), a pro-atherogenicand thrombogenic glycerophospholipid in the blood and,thereby, blocks signaling pathways activated by the interactionof LPA with cellular LPA receptors (LPAR). LPA-LPAR1interaction activates the Rho kinase-NF-\u03baB signaling pathwayand the subsequent transcriptional expression of ICAM-1,VCAM-1 and E-selectin in ECs .LPA-LPAR1/2 interaction leads to an increase in the contractilityof ECs and the permeability of the endothelial monolayer. LPA is an extracellular signalingmolecule that is capable of interacting with at least six G protein LPA-LPAR1/2 interaction leads to an increase in the contractilityof ECs and the permeability of the endothelial monolayer. LPA is an extracellular signalingmolecule that is capable of interacting with at least six G protein-coupled cellular receptors, initiating intracellular signalingcascades  and exerting multiple effectson blood cells  and vascular wall cells. Therefore, in platelets, LPA induces a changein the shape, aggregation, and formation of platelet-monocyteaggregates. In ECs, LPA induces cell migration, expressionof VCAM-1, ICAM-1, E-selectin, chemokines (CXCL1),formation of actin stress fibres and cell contraction. In SMCs,LPA induces cell contraction, migration and proliferation.Additionally, LPA accumulates in the lipid-rich core of atheroscleroticplaques, and, when ruptured, enters the bloodstreamand activates platelets, leading to the formation of blood clots. Endothelial NO synthase (eNOS) isalso expressed under physiologically high WSS. Under theseconditions, the eNOS promoter is activated through the interactionof NF-\u03baB with the GAGACC element . eNOS produces NO, an inducer of I\u03baB\u03b1 expression.NO stabilises the NF-\u03baB\u2219I\u03baB\u03b1 heterotrimer in the cytoplasmand induces I\u03baB\u03b1 translocation into cell nuclei, leading to theinactivation of NF-\u03baB and the termination of NF-\u03baB-mediatedtranscription . Moreover, NO inhibitsI\u03baB kinase (IKK complex), a positive regulator of NF-\u03baBactivation .Mechanosensitive signaling pathwaysthat control the transcriptional expressionof the RelA subunit of NF-\u03baB in endothelial cellsThe WSS, created by the blood flow to the walls of the arteries,activates mechanosensitive signaling pathways in the ECs.Amechanosensor localised in the cell membrane perceives amechanical stimulus (WSS) and triggers intracellular signalingthat activates specific transcription factors, which regulatethe transcriptional expression of proteins. The internal bendof the aortic arch and the areas of arterial bifurcation, wherelow and low/oscillatory WSS are realized, are associated withthe localisation of atheroma. In the same areas, increased expressionof active c-Jun N-terminal kinase 1 (JNK1), whichbelongs to mitogen-activated protein kinases (MAPK), wasrevealed , as well as increased expres-sionand nuclear localisation of the RelA subunit of NF-\u03baB. The hypothesis that physiologicallylow WSS regulates the expression of RelA and NF-\u03baB targetgenes (VCAM-1 and others) through the JNK1-dependentpathway was proved by modelling the haemodynamics ofthe carotid artery in mice. The implantation of a tapered cuffaround the mouse carotid artery generates a laminar bloodflow with a low velocity and low WSS upstream of the cuffand disturbed, low velocity and low/oscillatory WSS downstreamof the cuff .This approach demonstrated that increased expression ofRelA and activated JNK1 is realised in regions with lowand low/oscillatory WSS. The transcription factor ATF2 isactivated by JNK1 (phosphorylation) and interacts with theRelA promoter sites to activate the promoter. Thus, in areaswith low/oscillatory WSS, the JNK-ATF2-RelA signalingpathway is implemented, which stimulates the expression ofRelA and NF-\u03baB target genes (VCAM-1). The pathway alsostimulates the accumulation of CD68+ macrophages, whichare an indicator of the development of arterial inflammation.The signaling pathway for the transcriptional expressionof RelA upstream of JNK1 includes the activation ofintegrins, which is initiated by the stimulation of the cellsurface mechanosensory complex and an integrin-dependentsignaling cascade, leading to the activation of JNK1. The cellsurface mechanosensory complex includes the PECAM-1 andVE-cadherin receptors and the vascular endothelial growthfactor 2 receptor (VEGFR2), which belongs to the receptortyrosine kinase subfamily . In this complex,PECAM-1 is a key mechanosensitive signaling moleculethat perceives a mechanical signal and converts it into a chainof intracellular biochemical reactions. WSS, acting on theextracellular domain of PECAM-1, affects the conformationof the cytoplasmic domain of PECAM-1 and the availabilityof Tyr663 and Tyr686 of this domain for phosphorylation.Phosphorylation is conducted by membrane-bound Fyn tyrosinekinase (family of Src tyrosine kinases), which is localisedin intercellular contacts near PECAM-1 . PECAM-1/PECAM-1 intercellular EC interactionsthrough the extracellular domains of PECAM-1  are required for efficient phosphorylation and thetriggering of intracellular mechanosensitive signaling . Activated PECAM-1 andVE-cadherin (which functions as an adapter and is associatedwith VEGFR2) facilitate the phosphorylation of VEGFR2 atTyr801 and Tyr1175 by the Src tyrosine kinase. In turn, thephosphorylated VEGFR2, through direct interaction withthe regulatory p85 subunit of the phosphatidylinositol-3-OHkinase (PI(3)K), phosphorylates PI(3)K .The activated PI(3)K stimulates the conformational activationof integrins through the conservative pathway of theassociation of phosphatidylinositol-3,4,5 triphosphate (PI(3)Kproduct) with pleckstrin homology (PH) domains of cytohesin-1 or cytohesin-like proteins. This is followed by the translocationof these proteins to the plasma membrane and theirassociation with the cytoplasmic domains of the \u03b2 subunitsof integrins, which is realized in various types of cells. Thisinteraction leads to conformational changes in the extracellulardomains of the \u03b1 and \u03b2 subunits of integrins and an increase intheir affinity for specific ECM proteins, as well as clustering. In athero-resistant areasof arteries, the ECM is rich in CL (IV) and LN. However, inatherogenic regions (low/oscillatory WSS conditions), theECM is enriched in the pro-inflammatory proteins FN andFG .Antibodies specific to ligated \u03b21 and \u03b23 integrins were usedto show that, under conditions of low/oscillatory WSS, thereis an increase in the binding of \u03b15\u03b21 integrin to FN  and of \u03b1v\u03b23 integrin to VN.Thus, the conformational activation of integrins and thedynamic formation of new bonds of integrins with specificligands occur \u2013 ECM proteins are realized .The composition of the ECM and the ligation of integrinsby the ECM proteins activate many intracellular signalingcascades \u2013 in this case, the Shc-Grb\u2219Sos-Ras-MAPKsignaling pathway. The cellular adapter protein Shc, activatedunder conditions of low/oscillatory WSS by tyrosinekinases Src and VEGFR2 close to the EC contacts (phosphorylationof Tyr239/240), forms an early unstable complex Shc\u2219VEGFR2\u2219VE-cadherin . Next, a stablecomplex Shc with an \u03b1v\u03b23 integrin that is ligated with FN orVN is formed . Additionally, Shc associates with the \u03b21 and \u03b25 integrinsthat are ligated with FN and VN . In this manner, Shc coordinates the intercellularcontact proteins (VE-cadherin) and integrin-ECM interactionsunder low/oscillatory WSS conditions. Membrane-associatedsmall G proteins Ras  function cyclicallybetween active Ras\u2219GTP and inactive Ras\u2219GDP forms, whichare a molecular switch of an intracellular signal in response toan extracellular stimulus . The complexesof phosphorylated Shc with the \u03b1v\u03b23, \u03b21 and \u03b25 integrinsformed on the cytoplasmic side of the plasma membrane areaccompanied by the association of Shc with the cytoplasmicGrb2\u2219Sos complex of the growth factor receptor-bound protein2 (Grb2) and the guanine nucleotide exchange factor Sos.This complex stimulates the rate of exchange of the GDPassociated with Ras, on the GTP  . In turn, activated Rasstimulates cytoplasmic Raf kinase  and recruits Raf to the inner surfaceof the plasma membrane through direct interaction with itsregulatory domains and the subsequent phosphorylation offour sites of the kinase domain . Raf,activated via a MAP kinase cascade, activates MAPK (JNK).However, the activation of Shc and membrane-associatedheterotrimeric G proteins leads to the stimulation of Ras under the conditionsof the action of WSS on the cells. In the absence of WSS,G proteins are activated via association with ligand-activatedreceptors: the \u03b1 subunit exchanges the GDP bound to it forGTP and dissociates from the \u03b2\u03d2 dimer. The \u03b1\u2219GTP and \u03b2\u03d2complexes become mediators of cellular event signaling untilthe \u03b1 subunit restores its inactive GDP-bound state (since ithas GTPase activity.) The reassociation of \u03b1\u2219GDP with the \u03b2\u03d2dimer provides an inactive G\u03b1\u03b2\u03d2 heterotrimer that is capableof entering a new activation cycle . Alterationsin the physical properties of membranes under WSSconditions affect the conformation and functions of membrane-associatedproteins and, as a result, the signaling pathways activated bythese proteins .Purified heterotrimeric G proteins in phospholipid liposomesloaded with [\u03d2-32P]GTP were shown to be activatedby the action of physiological levels of WSS on liposomes. Within 1 second, a physiologically lowWSS activated the G\u03b1q/11 and G\u03b1i3/0 subunits of G proteins inECs . The activated G\u03b1q and G\u03b2\u03d2 subunitsdissociated from them under conditions of a WSS gradientin a physiologically low range and initiated Ras activationand downstream MAPK signaling .Moreover, two primary basic mechanosensors, G\u03b1q/11 andPECAM-1, establish a mechanosensitive G\u03b1q/11\u2219PECAM-1complex, which is formed under conditions of laminar flow in vivo (atheroprotective straight region of the descending aorta of mice) and in vitro. Underoscillatory flow conditions (low/oscillatory WSS), leading tothe activation of G proteins, this complex is rapidly (within30 seconds) destroyed . The formation ofthe G\u03b1q/11\u2219PECAM-1 complex involves PECAM-1 extracellularIg-like domains 2 and 3, as well as a G\u03b1q/11 interactingreceptor that associates with G\u03b1q/11 and PECAM-1 Ig-likedomains 2 and 3 and serves as a bridge in the formation of thecomplex . The PECAM-1\u2219G\u03b1q/11 complexalso includes heparan sulfate proteoglycan, which associateswith the Ig-like domain 3 of PECAM-1 and mediates theformation of the PECAM-1\u2219G\u03b1q/11 complex .Computer modelling of the blood flow in the arteries makesit possible to determine the most atherogenic areas of thearteries, which are characterised by low and low/oscillatoryWSS. The transcription factor NF-\u03baB and the cell adhesionmolecules ICAM-1, VCAM-1 and E-selectin are the earliestmarkers of atherogenesis. Therefore, the processes involved inthe expression of these proteins under the conditions inducedby a mechanical stimulus (low and low/oscillatory WSS) onendothelial cells are of great interest. This review presents ananalysis of numerous studies that demonstrated how the activationof membrane-bound proteins that perceive a mechanicalstimulus (low and low/oscillatory WSS) triggers a cascadeof biochemical reactions that lead to the transcriptional expressionof NF-\u03baB, a key regulator of the expression of celladhesion molecules. This review also describes in detail themechanisms of interaction between the endothelial cell adhesionmolecules and blood leukocytes that are responsible foradhesion and the subsequent TEM of leukocytes during the initialstage of atherogenesis. 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+{"text": "Senegalia pennata subsp. insuavis (Lace) Maslin, Seigler & Ebinger, Citrus hystrix DC. and Solanum melongena \u2018Kermit\u2019 extracts exhibited high antioxidant activities. Moreover, Citrus hystrix DC. extract was a potent inhibitor against lipase, angiotensin-converting enzyme and butyrylcholinesterase, while Coriandrum sativum L. and Psophocarpus tetragonolobus (L.) DC. were potent anti-diabetic agents and Senegalia pennata subsp. insuavis (Lace) Maslin, and Seigler & Ebinger was a potent anti-glycation agent. Our data provide a comparative analysis of ten vegetables to encourage healthy food consumption and development to control NCDs in Thailand in the future.Non-communicable diseases (NCDs) are the leading global cause of death. The World Health Organization (WHO) has endorsed the consumption of fruits and vegetables because they are rich in phytochemicals that sustainably ameliorate the occurrence of NCDs. Thai food contains many spices and vegetables with recognized health benefits. Quality control of plant samples encountered a bottleneck in the field and comparative studies of plant control origins including species or cultivar identification, growing area and appropriate harvesting time are limited. To address this issue, all plant samples used in this study were cultivated and controlled by the Department of Agriculture, Ministry of Agriculture and Cooperatives, Thailand. The samples were phytochemically screened and determined their health-promoting bioactivities via antioxidant activities and inhibition of NCD-related enzymes including lipase (obesity), \u03b1-amylase and \u03b1-glucosidase (diabetes), angiotensin-converting enzyme (hypertension), as well as acetylcholinesterase, butyrylcholinesterase and \u03b2-secretase (Alzheimer\u2019s disease). The non-enzymatic reaction toward glycation was also evaluated. The results showed that  Non-communicable diseases (NCDs) including cancer, diabetes mellitus (type II), cardiovascular diseases, hypertension, and Alzheimer\u2019s disease (AD) are the leading cause of death worldwide. In 2012, the World Health Organization (WHO) reported that NCDs accounted for 68% of morbidity (38 million deaths from 56\u2009million deaths), while over 16 million 40%) were early deaths (<70 years) [0% were eFruits and vegetables are rich in fiber, vitamins, minerals, and phytochemicals including alkaloids, anthocyanins, glucosinolates, flavonoids, phytosterols, phenolic acids and terpenoids that are secondary plant metabolites, with pharmacological effects toward a wide range of ailments including NCDs . PhytochAllium cepa Aggregatum Group, Allium fistulosum L., Allium sativum L., Citrus hystrix DC., Coriandrum sativum L., Cymbopogon citratus (DC.) Stapf, Eryngium foetidum L., Psophocarpus tetragonolobus (L.) DC., Senegalia pennata subsp. insuavis (Lace) Maslin, Seigler & Ebinger, and Solanum melongena \u2018Kermit\u2019 against NCDs. Despite having nothing in common , these plants have been selected based on their regular and frequent usages in most Thai cuisines. All plant samples were vetted for their origin and quality by the Department of Agriculture, Ministry of Agriculture and Cooperatives, Thailand. Thailand is renowned for its mouthwatering cuisine, consisting of a wide variety of textures and aromas emanating from local vegetable ingredients. Scientific reports have identified the health benefits of vegetables used in Thai dishes including antioxidant , anti-inAllium cepa Aggregatum Group (A. cepa), Allium fistulosum L. (A. fistulosum), Allium sativum L. (A. sativum), Citrus hystrix DC. (Ci. hystrix), Coriandrum sativum L. (Co. sativum), Cymbopogon citratus (DC.) Stapf (Cy. citratus), Eryngium foetidum L. (E. foetidum), Psophocarpus tetragonolobus (L.) DC. (P. tetragonolobus), Senegalia pennata subsp. insuavis (Lace) Maslin, Seigler & Ebinger (Se. pennata), and Solanum melongena \u2018Kermit\u2019 (So. melongena). Seven flavonoids were detected: quercetin, kaempferol, hesperidin, luteolin, apigenin, delphinidin, and cyanidin  contained two flavonoids at different concentrations. Four extracts possessed only one flavonoid: A. fistulosum (kaempferol), Cy. citratus (luteolin), E. foetidum (kaempferol), and Se. pennata (apigenin). Interestingly, no flavonoids were detected in A. sativum and So. melongena extracts. An aqueous ethanolic extract of Co. sativum was rich in quercetin (166.16 mg/100 g dry weight (DW)), followed by A. cepa and Ci. hystrix , while kaempferol (4.44\u201347.97 mg/100 g DW) was detected in A. fistulosum, Co. sativum, and E. foetidum extracts, with A. fistulosum extract exhibiting the highest (47.97 mg/100 g DW). Hesperdin (453.47 mg/100 g DW) was solely detected in Ci. hystrix extract, luteolin (4.57 mg/100 g DW) in Cy. citratus extract, apigenin (3.46 mg/100 g DW) in Se. pennata extract, and delphinidin (15.77 mg/100 g DW) in P. tetragonolobus extract. Cyanidin was also found in P. tetragonolobus (43.02 mg/100 g DW) and A. cepa (13.35 mg/100 g DW).High-performance liquid chromatography (HPLC) was employed to determine the specific phytochemical profiles covering the flavonoids and phenolic acids of vegetable extracts including cyanidin . Among tp-coumaric acid, and ferulic acid were detected. Caffeic acid and p-coumaric acid were general phenolics as they were observed in six extracts with different concentrations. The highest content of caffeic acid (246.99 mg/100 g DW) was detected in So. melongena extract that also contained minute amounts of p-coumaric acid (2.16 mg/100 g DW). The highest content of p-coumaric acid (68.13 mg/100 g DW) was detected in Cy. citratus extract, which also possessed the highest content of ferulic acid (123.34 mg/100 g DW) and marginal amounts of caffeic acid (15.65 mg/100 g DW). The second most abundant caffeic acid (52.69 mg/100 g DW) was detected in E. foetidum extract, which also contained minute amounts of p-coumaric acid and ferulic acid . An aqueous ethanolic extract of P. tetragonolobus contained the most varieties of phenolic acids including caffeic acid (16.13 mg/100 g DW), vanillic acid (15.71 mg/100 g DW), 4-hydroxybenzoic acid (10.80 mg/100 g DW), and p-coumaric acid (3.85 mg/100 g DW), while 4-hydroxybenzoic acid was only detected in P. tetragonolobus extract. Other than being observed in P. tetragonolobus extract, vanillic acid was also found in Co. sativum but in lower amounts (3.73 mg/100 g DW). This extract also contained caffeic acid (23.81 mg/100 g DW) and p-coumaric acid (5.20 mg/100 g DW). An aqueous ethanolic extract of A. fistulosum was found to possess ferulic acid (23.13 mg/100 g DW) and p-coumaric acid (7.12 mg/100 g DW), while Se. pennata contained only one phenolic acid: caffeic acid (14.92 mg/100 g DW). Interestingly, no phenolic acids were observed in A. cepa, A. sativum, and Ci. hystrix extracts.For phenolic acid determination , five phSe. pennata exhibited the highest TPC, followed by Ci. hystrix, So. melongena, Cy. citratus, A. fistulosum, P. tetragonolobus, E. foetidum, A. cepa, and Co. sativum, respectively, while A. sativum exhibited the lowest (twelve times lower than Se. pennata). A spectrophotometric analysis indicated that TPCs of all vegetable extracts ranged from 1.23 to 15.33 mg gallic acid equivalent (GAE)/g DW . The aquSe. pennata extract exhibiting the highest DPPH radical scavenging activity and A. sativum extract the lowest. For the FRAP assay, results were consistent with the DPPH radical scavenging activities. Se. pennata exhibited the highest reducing ability (62.33 \u03bcmol TE/g DW) of ferric iron (Fe3+) to ferrous iron (Fe2+), while A. sativum exhibited the lowest reducing activity (3.25 \u03bcmol TE/g DW). On the other hand, Ci. hystrix and So. melongena possessed the highest ORAC activities (415.92\u2013418.32 \u03bcmol TE/g DW), while A. cepa exhibited the lowest (15.12 \u03bcmol TE/g DW) at approximately 27-fold lower than Ci. hystrix and So. melongena. Phenolics contribute to a wide range of health benefits, including being antioxidants. Antioxidant activities were investigated covering both hydrogen atom transfer (HAT) and single electron transfer (SET) mechanisms . The ferAll the vegetable extracts contained phenolic acids and flavonoids that contributed to their therapeutic potential, specifically against some NCDs. Therefore, all the extracts were tested for their therapeutic potential against critical enzymes involved in NCDs and non-enzymatic reactions involving anti-glycation properties. Ci. hystrix exhibited the highest lipase inhibition, while A. cepa exhibited the lowest.Lipase is a lipid-degradation enzyme, and lipase inhibitors prevent fatty acid accumulation as one characteristic of obesity . The resCo. sativum (58.4%), E. foetidum (31.2%), and Ci. hystrix (26.6%) showed inhibitory activities of more than 25%  converts angiotensin I to angiotensin II, leading to vasoconstriction and increased blood pressure ; therebyA. sativum, inhibited AChE activities in the range of 8.3\u201358.6% using extract concentration of 1 mg/mL , one type of dementia, have been attributed to (i) degradation of the neurotransmitter acetylcholine by acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), or (ii) accumulation of amyloid plaque formed during amyloidogenesis by \u03b2-secretase (BACE-1) . Therefo 1 mg/mL . Among tntration . The hig 1 mg/mL . Among tSe. pennata provided the most potent anti-glycation activities induced by both D-glucose and MG, while A. sativum and A. cepa exhibited poor ability to inhibit glycation reactions.The glycation reaction is a non-enzymatic reaction involving interaction between monosaccharides and amino acids, lipids, or nucleotides. Methylglyoxal (MG), as a by-product of glycolysis, can act as a potent protein-glycation inducing agent. Finally, glycation induced by either D-glucose or MG as advanced glycation end products (AGEs) contributed to several ailments including premature aging and diabetes . The resRelationships between vegetable extracts and TPCs, antioxidant activities, enzyme inhibitory activities and anti-glycation properties were investigated using principal component analysis (PCA) and hierarchical cluster analysis (HCA) to determine the particular characteristics of each vegetable extract. Information gained from these statistical analyses will be helpful to classify Thai vegetables according to their unique health-promoting characteristics. PCA results showed that TPCs, antioxidant activities, enzyme inhibitory activities, and anti-glycation properties of Thai vegetable extracts could be easily classified. . A. cepa and A. sativum (blue letters) were separated from others (red letters). A. cepa and A. sativum (Cluster 1) were separated from the others (cluster 2) and located far away from the centroid , while cluster 2 consisted of A. fistulosum, Ci. hystrix, Co. sativum, Cy. citratus, E. foetidum, P. tetragonolobus, Se. pennata, and So. melongena (red color). The outcome of HCA supported the PCA results , Allium fistulosum L. (A. fistulosum), Allium sativum L. (A. sativum), Citrus hystrix DC. (Ci. hystrix), Coriandrum sativum L. (Co. sativum), Cymbopogon citratus (DC.) Stapf (Cy. citratus), Eryngium foetidum L. (E. foetidum), Psophocarpus tetragonolobus (L.) DC. (P. tetragonolobus), Senegalia pennata subsp. insuavis (Lace) Maslin, Seigler & Ebinger (Se. pennata), and Solanum melongena \u2018Kermit\u2019 (So. melongena) were comparatively analyzed regarding their phenolic profiles (phenolic acids and flavonoids) and in vitro inhibitory activities against some NCDs. The health-promoting activities involved the inhibition of the key enzymes that control NCDs including lipase (obesity), \u03b1-amylase, and \u03b1-glucosidase (diabetes), angiotensin-converting enzyme (hypertension) and acetylcholinesterase, butyrylcholinesterase, and \u03b2-secretase (Alzheimer\u2019s disease) as well as the non-enzymatic anti-glycation reaction (premature aging). Results showed that among these ten plant extracts, A. cepa exhibited the strongest angiotensin-converting enzyme (ACE) inhibition, while A. fistulosum could effectively fight against \u03b2-secretase (BACE-1). Nevertheless, A. sativum seemed to be the least active extract against these NCDs-related enzymes, in which no inhibitory activities against \u03b1\u2013glucosidase, acetylcholinesterase (AChE), and BACE-1 were observed. However, high ACE inhibitory activity was observed in this plant extract, while other enzyme inhibitory activities were quite low (less than 50% inhibition). On the other hand, Ci. hystrix was the most active extract, which exhibited the strongest antioxidant activity and inhibitory activities against lipase, ACE and butyrylcholinesterase (BChE). Co. sativum was highly effective against \u03b1-amylase, while P. tetragonolobus exhibited the highest \u03b1-glucosidase inhibitory activity. Cy. citratus seemed to exhibit low to moderate enzyme inhibitory activities, with the exception of ACE inhibitory activity, which was more than 50% inhibition. E. foetidum exhibited the highest AChE inhibitory activity, while Se. pennata exhibited strong antioxidant activity and ACE inhibitory activity. Similarly, So. melongena was also a good source of antioxidants. From these results, it is of interest to group these plants according to their bioactivities and discuss the following topics in more detail; (i) Se. pennata exhibited the highest reducing and free radical scavenging abilities, while Ci. hystrix and So. melongena possessed the highest oxygen radical absorbance capacity; (ii) Ci. hystrix exhibited the highest lipase inhibition; (iii) Co. sativum and P. tetragonolobus were potential anti-diabetic agents with high inhibitions against carbohydrate-degrading enzymes; (iv) A. cepa, Ci. hystrix, and Se. pennata had the three highest ACE inhibitory activities; (v) Ci. hystrix and E. foetidum were potential anti-Alzheimer\u2019s disease (AD) agents with high inhibitions against acetylcholine degrading enzymes, whereas A. fistulosum acted against amyloid generating enzyme, and (vi) Se. pennata was a potential anti-glycation agent.Interest in indigenous plants for their health benefits in terms of disease prevention beyond nutritional benefits is increasing. Thai cuisine consists of various spices and herbs with unique aromas and flavors that also have health-promoting bioactivities. Copious literature exists on plant beneficial health characteristics, but control of plant origins is lacking, leading to the absence of a comparative analysis of these vegetables. Here, ten vegetables used in Thai cuisine including Se. pennata, Ci. Hystrix, and So. melongena exhibited high antioxidant capacities with the highest total phenolic contents (TPCs). Our results concurred with previous literature suggesting that TPCs and antioxidant activities in various plant extracts were strongly correlated [Se. pennata (or Cha-om in Thai) exhibit strong odor (some define as stinky) and are normally consumed as a blanched vegetable or mixed in an omelet and eaten with spicy sauce. In our study, Se. pennata exhibited high ferric reducing antioxidant power (FRAP) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activities, suggesting that antioxidants in this vegetable extract likely possess an ability to transfer one electron to any potential electron acceptors. A previous report suggested that methanolic extract of Se. pennata leaves exhibited high TPCs of 45.3 \u00b5g gallic acid equivalent (GAE)/mg dry extract with half maximal effective concentration (EC50) of 3.6 mg extract/mg DPPH [50) of 2.0 mg/mL [Ci. hystrix and So. melongena exhibited high oxygen radical absorbance capacity (ORAC), suggesting that the antioxidants in these vegetable extracts likely donate hydrogen atoms to free radicals. The fruit peel of Ci. hystrix  is commonly used in many Thai recipes (such as chili paste and curry) for its strong and unique aroma. Eggplants in Thailand can be classified into 22 species of Solanum, with 10 cultivars commercially available [So. melongena can be consumed as fresh or blanched vegetables and are the main ingredient in curry. Previous literature also reported high TPCs and antioxidant activities of these two vegetables. Ethanolic extracts of Ci. hystrix fruit peel were previously reported to exhibit TPCs of 0.32 mg GAE/mg extract and DPPH radical scavenging activity with IC50 of 0.09 mg/mL [So. melongena  exhibited DPPH radical scavenging activities of 17.52\u201346.13% [Among the vegetable extracts, rrelated ,28. Youn/mg DPPH , while i.0 mg/mL . Our stuvailable . Fruits 2\u201346.13% .Ci. hystrix but not on its anti-obesity property. Interestingly, in this study, fruit peel extract of Ci. hystrix was found to exhibit the highest lipase inhibition, an activity that retards lipid absorption and, thus, is related to the control of obesity. As the most abundantly found flavonoid in fruit peel of Ci. hystrix, hesperidin exhibited IC50 activity of 52.4 \u00b5M against porcine pancreatic lipase [50 of 6.1 \u00b5M [50 of 4 \u00b5M [Ci. hystrix, it was previously found that phenolic acids were generally less active against lipase inhibition than flavonoids [Ci. hystrix with the highest content of hesperidin and moderate amounts of quercetin possessed a potential bioactivity against lipase. Many authors have reported on the biological activities of c lipase , while af 6.1 \u00b5M . Compare of 4 \u00b5M , quercetavonoids ,35. TherCo. sativum and P. tetragonolobus effectively inhibited \u03b1-amylase and \u03b1-glucosidase, respectively. Leaves and young shoots of Co. sativum (coriander) are added to many Thai dishes including soup or stir fry meat as a decorated vegetable with a strong unique aroma. However, recently, coriander has become popular consumed as a fresh side dish vegetable along with other main dishes. Oral administration of the ethanolic leaf extract to mice under induced insulin deficiency resulted in lowered blood glucose [50 value 2.5-times lower than acarbose, a synthetic anti-diabetic drug [Co. sativum leaves inhibited \u03b1-amylase with 19% inhibition using extract concentration of 1 mg/mL [50 of 14.60 \u00b5M against pancreatic \u03b1-amylase, the most abundant flavonoid in Co. sativum leaves as quercetin exhibited IC50 of 12.7 \u00b5M [50 of 20.4 \u00b5M [Co. sativum is a good candidate as an anti-diabetic agent, even its seeds [P. tetragonolobus (or winged bean) is a Thai local vegetable normally consumed as boiled or blanched young bean pod with many spicy sauces. At present, no report on young bean pod of P. tetragonolobus regarding its anti-diabetic property is available. Nevertheless, major phenolics in P. tetragonolobus such as cyanidin and delphinidin effectively inhibited Saccharomyces cerevisiae \u03b1-glucosidase with IC50 values of 17.0 and 4.1 \u00b5M, respectively, compared to acarbose with IC50 of 0.53 \u00b5M [P. tetragonolobus also contained moderate contents of phenolic acids including 4-hydroxybenzoic acid, vanillic acid, caffeic acid and p-coumaric acid, while phenolic acids generally inhibited \u03b1-glucosidase with a lesser effect than flavonoids [As vegetable extracts with potential anti-diabetic properties,  glucose ,35,36. T 1 mg/mL . Compare 12.7 \u00b5M , while t 20.4 \u00b5M . With hits seeds ,40,41. L 0.53 \u00b5M . P. tetravonoids ,43,44. Cavonoids . A. cepa, Ci. hystrix and Se. pennata had the three highest ACE inhibitory activities. These results concurred with previous reports indicating that quercetin-rich onion skin extract decreased ambulatory blood pressure in patients under metabolic syndrome (overweight/obese/hypertension) [A. cepa as quercetin exhibited IC50 of 43 \u00b5M against rabbit lung ACE [Ci. hystrix, exhibited half inhibitory activity of quercetin using a concentration of 0.5 mM [Se. pennata (but in low amounts compared to other vegetables) with IC50 of 5.7 mM [Interestingly, all vegetable extracts effectively inhibited ACE at more than 50% inhibition using extract concentration of 1 mg/mL. Among these, tension) . The prelung ACE , while hf 0.5 mM . Caffeicf 5.7 mM .Ci. hystrix and E. foetidum acted as inhibitors for acetylcholine-degrading enzymes, while A. fistulosum showed promise as a potential extract inhibiting amyloid production. Ci. hystrix and E. foetidum were rich in hesperidin and caffeic acid, respectively, while the consumption of hespiridin-rich extract or caffeic acid avoided cognitive dysfunction and learning deficit in vivo [50 against AChE and BChE at 22.8 and 48.9 \u00b5M, respectively, and caffeic acid at 23.36 and 29.19 \u00b5M, respectively [E. foetidum, usually used in the famous sour soup called \u201cTom Yum\u201d was available; thus, future studies on the anti-AD properties of the caffeic acid rich extract of E. foetidum are required. When considering anti-AD via the inhibition of amyloid production, all extracts displayed mild to low BACE-1 inhibition. A. fistulosum and A. cepa gave the two highest anti-BACE-1 activities, albeit carrying different phytochemicals. A. cepa showed high quercetin, while A. fistulosum was high in kaempferol. Quercetin and kaempferol are well-known flavonoids exhibiting anti-BACE-1 properties with IC50 values at 5.4 and 14.7 \u00b5M, respectively [Co. sativum also possessed high quercetin (2.5-fold higher than A. cepa); however, the anti-BACE-1 activity was lower, indicating that multi-interaction of phytochemicals within the extract may display antagonist effects. For anti-AD properties, our data showed that using an extract concentration of 1 mg/mL,  in vivo ,49. Hespectively . Thus, hectively . Leaves Se. pennata may be a potential anti-glycation agent for both reactions. Se. pennata is typically fried with egg or with \u201cTom Yum\u201d soup. The glycation reaction and its AGEs relate to free radical productions [Se. pennata exhibited high antioxidant activity covering the SET mechanism due to its high phenolics content. It remains unclear which phytochemicals in Se. pennata exhibit this property because only trace amounts of caffeic acid and apigenin were observed, even though these two compounds were documented for their anti-glycation properties [The glycation reaction leads to the formation of advanced glycation end products (AGEs) that contribute to diseases such as diabetes, AD, and premature aging . The reaductions . Hence, operties ,55.Ci. hystrix). However, ACE inhibitory activity was different from other bioactivities since it was located in a different axis (PC3). Moreover, ACE inhibitory activities of all vegetable extracts were higher than 50%, even though the extract concentration used in this enzyme inhibitory assay was lower (0.2 mg/mL) than others (1 mg/mL in other enzyme inhibitory assays and 0.63 mg/mL in glycation reactions). It was previously reported that other than phenolics that could act as ACE inhibitors, small peptides could also act as effective ACE inhibitors as well [A. cepa and A. sativum exhibited low TPCs, antioxidant activities, and anti-glycation properties. Cluster 2 consisted of A. fistulosum, Ci. hystrix, Co. sativum, Cy. citratus, E. foetidum, P. tetragonolobus, Se. pennata, and So. melongena exhibited high TPCs, antioxidant activities, and enzyme inhibitory activities. Additionally, the principal component analysis (PCA) suggested that the activities that lied in the same axis were closely related to each other. For example, TPCs, antioxidant activities, \u03b1-glucosidase inhibitory activities, AChE inhibitory activities, and anti-glycation properties were on the same axis (PC1). The extract with relatively high TPCs and antioxidant activities would potentially exhibit relatively high anti-\u03b1-glucosidase, AChE, or glycation activities as well , Allium fistulosum L. (A. fistulosum), Allium sativum L. (A. sativum), Citrus hystrix DC. (Ci. hystrix), Coriandrum sativum L. (Co. sativum), Cymbopogon citratus (DC.) Stapf (Cy. citratus), Eryngium foetidum L. (E. foetidum), Psophocarpus tetragonolobus (L.) DC. (P. tetragonolobus), Senegalia pennata subsp. insuavis (Lace) Maslin, Seigler & Ebinger (Se. pennata), and Solanum melongena \u2018Kermit\u2019 (So. melongena) were collected following the recommendation of the Department of Agriculture, Ministry of Agriculture and Cooperatives, Thailand. The samples were deposited at the Bangkok Herbarium (BK), Bangkok, Thailand. Physical appearance of the edible part, harvesting time and the herbarium voucher specimen are provided in A. cepa (bulps), A. fistulosum (leaves), A. sativum (bulps), Ci. hystrix (fruit peel), Co. sativum (leaves), Cy. citratus , E. foetidum (leaves), P. tetragonolobus (whole fruits), Se. pennata (young leaves), and So. melongena (whole fruits) were cleaned with deionized (DI) water before freeze-drying using a Heto PowerDry PL9000 Freeze Dryer  for 3 days. The dry samples were then ground into fine powder using a Philips 600W Grinder  and expressed as CIELAB units (L* represented dark (0) to white (100), a* represented green (\u2212) to red (+), while b* represented blue (\u2212) to yellow (+)) as shown in Ten vegetables including v/v) aqueous ethanol (1:10 ratio) at 37 \u00b0C for two hours. The mixture was centrifuged at 3800 g for 15 min using a Hettich\u00ae ROTINA 38R centrifuge . The supernatant was collected, while the residue was repeatedly extracted with the same procedure twice. The supernatants from three extractions were pooled, and ethanol was removed by a rotary evaporator . The dried extracts were re-dissolved in DMSO, filtered through a 0.45 \u00b5M polytetrafluoroethylene (PTFE) membrane syringe filter, and kept at \u221220 \u00b0C until analysis. The dry samples were extracted using 80% (v/v) aqueous methanol (40 mL), 6 N HCl (10 mL) and 0.5 g/L tert-butylhydroquinone (tBHQ). Prior to injection into the HPLC system, the extract (10 mg/mL) was filtered through a 0.22 \u03bcM PTFE membrane. Milli-Q water (18.2 M\u2126.cm resistivity at 25 \u00b0C), HPLC-grade methanol, and HPLC-grade acetonitrile containing 0.05% (v/v) trifluoroacetic (TFA) were used as gradient mobile phases with a constant flow rate of 0.6 mL/min [p-coumaric acid , sinapic acid , and syringic acid (>97.0% T) were received from Tokyo Chemical Industry , while vanillic acid (\u226597% HPLC) and gallic acid (97.5\u2013102.5% T) were received from Sigma-Aldrich . The authentic flavonoid standards including quercetin , kaempferol (>97.0% HPLC), luteolin (>98.0% HPLC), hesperidin , naringenin , myricetin (>97.0% HPLC), and apigenin (>98.0% HPLC) were obtained from Tokyo Chemical Industry , while isorhamnetin (\u226599.0% HPLC), cyanidin (\u226596.0% HPLC), and delphinidin (\u226597.0% HPLC) were from Extrasynthese . The phenolic acids were detected at 280 nm and 325 nm, while flavonoids were detected at 338 nm and 368 nm. HPLC chromatograms were shown in To determine the phenolic profile, high-performance liquid chromatography (HPLC) was employed using an Agilent 1100 HPLC system equipped with a photodiode array detector and a Zorbax Eclipse XDB\u2013C18 column  as previously described . In brie6 mL/min . The auty is an area under the peak, a is a y-intercept, b is a slope of the calibration curve, and x is a standard concentration. LOD and LOQ were calculated using the following equation:aS is a standard deviation of the response (y-intercept), and b is a slope of the calibration curve. The intra-day precision was presented as a percentage of the relative standard deviation (%RSD) and calculated using the following equation:tRS is a standard deviation of the retention time, and tRMean is the mean of the retention time measured at all concentrations of each standard.Linear range, linear regression, correlation coefficients, limit of quantitation (LOQ), limit of detection (LOD), and relative standard deviation (RSD) of the standards were analyzed according to the protocol of Srinuanchai et al. 2019  as shownTotal phenolic contents (TPCs) were investigated using Folin\u2019s phenol reagent as formerly described . Gallic Antioxidant activities of the extracts were determined using 2,2-diphenyl-1-picrylhydrazyl (DPPH) together with ferric reducing antioxidant power (FRAP) and oxygen radical absorbance capacity (ORAC) assays as previously described . Trolox,TM HT 96-well UV-visible microplate reader and Gen5 data analysis software . To determine the enzyme inhibitory activities of the extracts against some NCDs, key enzymes that control obesity (lipase), diabetes (\u03b1-amylase and \u03b1-glucosidase), hypertension (angiotensin-converting enzyme), and Alzheimer\u2019s disease  were chosen for inhibitory reactions using the well-established protocols as previously described ,12,13,22Candida rugosa lipase , 50 \u00b5L of 0.2 mM 5-5\u2032-dithiobis, 10 \u00b5L of 16 mM 5,5\u2032-dithiobis(2-nitrobenzoic acid) (DTNB) and 40 \u00b5L of the extract (5 mg/mL). The inhibitory activity was visualized as a decline in enzyme kinetics at 412 nm.Briefly, the lipase inhibitory reaction consisted of 100 \u00b5L of 0.01 mg/mL p-nitrophenyl-\u03b1-D-maltopentaoside and 50 \u00b5L of the extract (4 mg/mL), while the \u03b1-glucosidase inhibitory reaction consisted of 100 \u00b5L of 0.1 U/mL Saccharomyces cerevisiae \u03b1-glucosidase , 50 \u00b5L of 2 mM p-nitrophenyl-\u03b1-D-glucopyranoside and 50 \u00b5L of the extract (4 mg/mL). The inhibitory activity was visualized as a decline in enzyme kinetics at 405 nm.The \u03b1-amylase inhibitory reaction consisted of 100 \u00b5L of 30 mg/mL porcine pancreatic \u03b1-amylase , 50 \u00b5L of 30 mM o-phthaldialdehyde and 50 \u00b5L of the extract (0.4 mg/mL). The inhibitory activity was evaluated using an excitation wavelength of 360 nm and an emission wavelength of 485 nm as an end-point assay.The angiotensin-converting enzyme (ACE) inhibitory reaction consisted of 3 \u00b5L of 0.5 U/mL rabbit lung ACE (\u22652 unit/mg), 30 \u00b5L of 3 mM hippuryl-histidyl-leucine, 15 \u00b5L of 20 mg/mL Electrophorus electricus AChE (1000 units/mg), 40 \u03bcL of 0.8 mM acetylthiocholine, 10 \u00b5L of 16 mM DTNB and 40 \u00b5L of the extract (5 mg/mL), while the butyrylcholinesterase (BChE) inhibitory reaction consisted of 100 \u00b5L of 0.5 \u00b5g/mL equine serum BChE (\u226510 units/mg), 40 \u00b5L of 0.4 mM butyrylthiocholine, 10 \u00b5L of 16 mM DTNB and 40 \u00b5L of the extract (5 mg/mL). The inhibitory activity was visualized as a decline in enzyme kinetics at 412 nm. The \u03b2-secretase (BACE-1) inhibitory reaction was studied using a BACE-1 fluorescence resonance energy transfer (FRET) assay kit  according to the manufacturer\u2019s recommendations. The inhibitory activity of the extract (20 \u00b5L of 5 mg/mL) was evaluated using an excitation wavelength of 320 nm and an emission wavelength of 405 nm as an end-point assay. The acetylcholinesterase (AChE) inhibitory reaction consisted of 100 \u03bcL of 20 ng The anti-glycation reaction induced by D-glucose consisted of 50 \u00b5L of 20 mg/mL bovine serum albumin  in 100 mM potassium phosphate buffer (pH 7.4) containing 0.02% (w/v) sodium azide, 25 \u00b5L of 1 M D-glucose and 25 \u00b5L of extract (2.52 mg/mL). For the anti-glycation reaction induced by methylglyoxal (MG), 25 \u00b5L of 4 mM MG was used instead of D-glucose. The reaction mixture was incubated at 37 \u00b0C for 2 weeks in the dark. The inhibitory activity was evaluated using an excitation wavelength of 330 nm and an emission wavelength of 410 nm as an end-point assay.A is the initial velocity of the control reaction with enzyme (control), a is the initial velocity of the control reaction without enzyme (control blank), B is the initial velocity of the enzyme reaction with extract (sample) and b is the initial velocity of the reaction with extract but without enzyme (sample blank). The percentage of enzyme inhibition using an end-point assay and anti-glycation reaction was evaluated using the same equation but changing from initial velocity to absorbance at a particular wavelength.The percentage of enzyme inhibition using enzyme kinetics was calculated using the following equation:\u00ae  to create a biplot. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) of TPCs, antioxidant activities and enzymatic and non-enzymatic inhibitory activities of Thai vegetable extracts were performed using XLSTATn = 3), with data expressed as mean \u00b1 standard deviation (SD). Statistical analysis was performed using the statistical package for the social sciences . Significant difference at p < 0.05 of more than two data was calculated using one\u2013way analysis of variance (ANOVA), followed by Duncan\u2019s multiple comparison test, while significantly difference at p < 0.05 of two data was calculated by Student\u2019s t-test. All experiments were evaluated in triplicate ("}
+{"text": "Ornithine aminotransferase (OAT) catalyzes transfer of the delta-amino group from L-ornithine to oxo-glutarate.In plants, this reaction biochemically connects urea cycle, proline cycle, and polyamine biosynthesis pathway.OAT activity is shown to be associated with biotic and abiotic stress responses and nitrogen metabolism, but itsphysiological role is still unclear. In our study, we decided to investigate transcriptional regulation of the OAT gene inArabidopsis thaliana under normal conditions and in response to various growth regulators. In the present work, thereporter gene construct containing the Escherichia coli \u03b2-glucuronidase gene (gus) under control of the A. thaliana OATgene promoter was introduced into the genome of A. thaliana ecotype Columbia plants using the floral dip method;GUS activity was assayed in different experimental conditions including hormone treatment, low and high nitrogen andsalinity. The GUS activity was analyzed histochemically. Plants were incubated with staining solution containing X-Gluc.We show that under standard growth conditions, the promoter is active during germination and in developing floralorgans. OAT promoter activity specifically activates in response to different forms of auxin , cytokinin(6- BAP), ethylene precursor (ACC), high nitrogen and salinity. Analysis of the OAT expression by qRT-PCR confirmed thepattern observed using the GUS reporter system. The OAT gene showed a significantly elevated expression in fourday-old seedlings and in plant roots in response to auxins and cytokinins. The analysis of the OAT promoter structurereveals cis-acting regulatory DNA elements associated with auxin regulation and abiotic stresses. The results of thestudy indicate that the OAT gene is involved in developmental processes and is regulated by auxin and cytokinins. The ornithine-\u03b4-aminotransferase (OAT) is a mitochondrialpyridoxal-5-phosphate (PLP)-dependent enzyme that transfersan amino group from ornithine to oxo-glutarate with formationof glutamate-1-semialdehyde (GSA) and glutamate . Although the biochemical function ofOAT is known, its biological role in plants is not fully understood.On the one hand, OAT is involved in metabolism of ornithine,which takes part in numerous biochemical processes inplants, such as arginine metabolism, synthesis of polyaminesand alkaloids . Onthe other hand, one of the products of the reaction mediated byOAT, namely GSA, is involved in proline production. It readilyinterconverts into the cyclic 1-pyrroline-5-carboxylate (P5C),an intermediate in the proline biosynthesis, in a non-enzymaticfashion . Proline is involved in plantstress response  anddevelopment . It has already beenshown in various experiments on several plant species thatoverexpression of the OAT gene is associated with increasedproline content and resistance to abiotic stresses . It is tempting to assume thatOAT might link biological processes related to proline, ornithineand P5C metabolism, such as nitrogen recycling, stressresponse, secondary metabolism, growth and development.We have previously shown that OAT overexpression in tobaccoincreases salt stress resistance. Interestingly, the levelof proline accumulation in OAT overexpressing lines did notdiffer from that of WT plants under both normal and stressconditions, suggesting that OAT might contribute to stressresistance through processes not related to proline synthesis. On a model of transgenic tobaccoplants expressing GUS under the control of putative Arabidopsisthaliana OAT promoter we showed that the promoter activityis associated with meristems and zones of active growth. This observation suggests that theOAT gene might be involved in developmental processes. Thepresent study aims to investigate transcriptional regulation ofthe OAT gene in A. thaliana under normal conditions and inresponse to various growth regulators.Development of transgenic Arabidopsis harboring AtOATpromoter construct. The 1844 bp region upstream of theOAT gene translation start  was clonedin the promoterless vector pBI101 with the formation of theP1844 construct . The resultingvector contains the expression cassette harboring the \u03b2-glucuronidase(gus) reporter gene under the control of putativeA. thaliana OAT promoter. A. thaliana plants ecotype Columbiawere grown at 22 \u00b0C in a long-day growth conditions (16 hof light and 8 h of dark). Construct P1844 was transformedinto Agrobacterium tumefaciens strain AGL0, which was usedto transform A. thaliana by floral dip method . T1 transformants were screened on 1/2 MS agar platescontaining 50 mg/L kanamycin, transferred to pots and grownto maturity until the T2 generation seeds were harvested.T2 seeds were germinated on 1/2 MS agar plates containing50 mg/L kanamycin and resistant plants were tested for thepresence of GUS activity by histochemical assay. Six independenttransgenic T2 lines showing the presence of GUS activityin seedlings were selected for further experiments. Plants fromthe selected lines were grown to maturity and T3 generationseeds were harvested. Thus, six independent T3 transgeniclines have been obtained.GUS staining. The histochemical staining method  was used to visualize GUS (Escherichia coli\u03b2-glucuronidase) activity in seedlings grown on agar platesand plant parts grown in soil (5-week-old plants). Wholeseedlings and different plant parts were incubated in X-Glucsolution Triton-X) for 24 h at 37 \u00b0C. Chlorophyll was removed byrepeated washing in 70 % (v/v) ethanol. GUS activity wasobserved using a ZEISS Stemi 2000-C microscope coupledwith an AxioCam HRc cameraExperimental treatments. Surface-sterilized seeds of sixindependent transgenic A. thaliana lines (T3) were germinatedon MS plates supplemented with 1 % sucrose, 0.7 % agar. Todetect promoter activity during germination, histochemicalassay was performed for seedlings at 3rd, 5th, 6th and 14th dayafter sowing (DAS) on plates. For experimental treatments,one-week-old seedlings were transferred to the same mediumsupplemented with the following growth regulators (fromSigma-Aldrich): auxins , cytokinins , gibberellic acid (10 \u03bc\u041c GA3),100 \u03bc\u041c abscisic acid, 1 m\u041c methyl jasmonate, ethyleneprecursor (50 \u03bc\u041c ACC), high nitrogen (10 mM NH4NO3),high salinity (200 mM NaCl). For low nitrogen treatment,MS NH4NO3-free medium (Duchefa Biochemie) was used.GUS activity was assayed after 1, 4, 6 and 8 days of treatment.For cold and heat treatment, two-week-old transgenicplants were used. For cold treatment, plates with seedlingswere incubated at +4 \u00b0C for 4 h, then for 2 h at 22 \u00b0C; forheat treatment, plates were incubated at +50 \u00b0C for 15 min,then 6 h at 22 \u00b0C.Gene expression analysis (RNA isolation and qRT\u2011PCR).Wild type Col-0 seed was surface sterilized with 12.5 %bleach  and 70 % ethanol and germinated on 1/2 MSmedium . To measure expression ofthe OAT gene during germination and early development,total RNA was isolated from whole seedlings at 4th, 7th and14th DAS. For experimental treatments, one-week seedlingswere transferred to 1/2 MS medium supplemented with differentgrowth regulators , and control, to 1/2 MSmedium. For each treatment, experiment was performed inthree biological replicates. There were 30 seedlings per eachbiological replicate. Total RNA was isolated from roots ofseedlings after 6 days of treatment with the RNeasy PlantMini Kit (Qiagen). RNA was treated with DNAse (QIAGENRNase-Free DNase Set). The concentration of RNA wasmeasured by NanoDrop 2000 (Thermo Scientific). The qualityof RNA was evaluated using Bioanalyzer 2100 (Agilent).First strand cDNA was synthesized from 1 \u03bcg of total RNAusing BIORAD iScript\u2122 Reverse Transcription Supermixfor RT-qPCR. For qRT-PCR analysis, cDNA was diluted tentimes. PCR was performed in a final volume of 15 \u03bcL: 3 \u03bcLof 5\u0445 Low Rox buffer (SibEnzyme), 0.15 \u03bcL of each primer(10 \u03bcM) and the taqman probe solution, 3 \u03bcL of diluted cDNA. The primers and probes were designed using IDT\u2019sPrimerQuest Tool (https://eu.idtdna.com/PrimerQuest/). Thecomparative threshold cycle method was used to determinerelative gene expression, with the expression of EF1-alfa andF-box (accession no. At1g13320 and At5g15710) serving asan internal control. The structures of primers and probes aregiven in Suppl. Table 11. The relative expression levels ofOAT mRNA in all the treated samples were quantified usingan Applied biosystems 7500 Real Time PCR System. Eachreaction was performed in three technical replicates usingthe following program of the qRT-PCR; 95 \u00b0C for 10 min;45 cycles of 95 \u00b0C for 15 s, 68 \u00b0C for 60 s. Statistical analysiswas performed using Student\u2019s t-test. p-values < 0.05 wereconsidered significant.http://www.bionet.nsc.ru/vogis/download/pict-2022-26/appx4.pdfSupplementary Tables 1\u20134 are available in the online version of the paper:Web tools used for cis-acting regulatory DNA elementssearch and expression data analysis. Search of cis-actingregulatory elements was performed using the PLACE database. Gene expression data from different microarray and RNA-seq experiments were extracted fromExpression Atlas and Arabidopsis eFP Browser Web tools.Tissue-specific promoter activation at different developmentalstages and under experimental treatments. StrongGUS staining was detected in hypocotyls and cotyledons ofseedlings at 3\u20134th DAS. At later stages, the GUS activity wasobserved only in cotyledons. In 6- and 14- DAS seedlings, theGUS activity was found only in the distal parts of cotyledons. During flower development, the GUS activity wasobserved in anthers, carpels and developing seeds of growingsiliques .To get a deeper insight into the transcriptional regulationof OAT, transgenic seedlings were subjected to experimentaltreatments including different concentrations of growth regulatorsand phytohormones auxins, cytokinins, gibberellin, ABA,methyl jasmonate, ethylene precursor (ACC), low and highnitrogen, high salinity, cold and heat stress . We observed tissue-specific OAT promoter activity inresponse to different forms of auxin ,cytokinin (6-BAP), and ACC. The strongest GUS activity wasobserved in response to 2,4-D in whole plant. Treatment withIAA and NAA caused GUS activation in root tips; treatmentwith 6-BAP \u2013 in the zone of root hairs. Treatment with ACC,salinity and nitrogen activated promoter along the whole root.OAT gene expression analysis. The qRT-PCR resultsshowed that transcript levels of the OAT gene are significantlyhigher in four-day-old seedlings, than at later developmentalstages . In experimental treatments, the OATgene showed significantly ( p \u2264 0.05) elevated expression inresponse to different forms of auxin  andcytokinin (6-BAP) in comparison to control conditions. Treatmentwith synthetic auxin 2,4-D led to 3-fold increase in OATexpression level in roots .Cis-acting regulatory DNA elements search and transcriptomicdata analysis. \u0421is-acting regulatory DNA elementssearch revealed putative transcription factors bindingsites, corresponding to different physiological processes, includinghyperosmotic and hypoosmotic stress response, auxinresponse, axillary bud dormancy control, specific regulation inontogenesis (Suppl. Table 3). Meta-analysis of microarray andRNA-seq data  shows that the OAT expression level changes in responseto cold, drought, heat, wounding, osmotic and salt stress. Expressionincreases in response to pathogens Botrytis cinerea,Pseudomonas syringae, Phytophthora infestans and someother infections. Altered OAT gene expression was observedin response to different hormones: it increased in response to3 h of treatment with ABA, methyl jasmonate, and decreasedin response to 3 h of treatment with brassinosteroids. The OATgene demonstrates high expression in seeds, siliques, embryos,senescent leaves, floral organs in A. thaliana (Suppl. Table 4).For more than a decade OAT has been considered an enzymeinvolved in metabolic response to different stress conditions,such as osmotic stress, pathogen attack and ROS production,nitrogen starvation, etc. . This enzyme belongs to the networkof nitrogen-metabolizing pathways in plants, affected byvarious environmental stimuli. It has been shown that plantsaccumulate proline during stress conditions . The results of Funck et al. (2008) and ourprevious study did not support the hypothesis of OAT contributionto proline accumulation. Instead, a specific role of the OATgene in plant developmental and growth processes under bothnormal and stress conditions is hypothesized . This study provides a deeper insight intothe role of the OAT gene in plant development.The important metabolic role of the OAT was clearly shownin an experiment where OAT-deficient plants failed to developwith arginine or ornitine as the sole nitrogen source . This result demonstrated that OAT is requiredfor utilization of arginine and ornithine. The present studydemonstrates high OAT promoter activity and elevated OATtranscript level during seed germination. These results are inagreement with available transcriptomic data . In arginine catabolism, OAT acts downstream ofarginase . Arginine is regarded as a majornitrogen storage compound in seeds. Urease and arginaseactivities increase sharply during germination in A. thaliana and other plant species . Taken together, these data provide evidence for OATinvolvement in nitrogen reorganization during seed germinationtogether with other enzymes of arginine catabolism.Our work also shows that the OAT gene promoter is activeduring inflorescence development. This observation isin accordance with recent findings showing that the OATenzymeplays a role in flower development and seed settingin rice . It has been reported that rice plantswith a mutated OAT gene (OsOAT mutants) have differentabnormalities in inflorescence and seed development. Themutant phenotype of the OsOAT mutant could be rescued byapplication of urea . Authors assumed thatOAT mediates arginase activity and plays a role in regulationof nitrogen reutilization, which is critical for developing tissues.Taking into account the association between the OAT geneexpression and proline accumulation , it can be assumed that OAT enzyme activity may also play a role in control of proline level duringinflorescence development. It has been reported that someproline metabolic enzymes can regulate a number of developmentalprocesses including flowering time , pollen development  and root growth . Prolineis known to be accumulated in reproductive organs of manyplant species . Ornithine to prolineconversion is mediated by the plant oncogene RolD , the overexpression of which stimulates floweringand affects inflorescence architecture in transgenic tobaccoplants . This study shows that the OATpromoter is active in inflorescences on different developmentalstages , suggesting that the OAT enzyme canconvert ornithine to proline directly or indirectly via argininecatabolism and glutamate production and might serve as aregulator of proline level during inflorescence development.Tissue-specific activation of the OAT transcription in rootsin response to auxin and cytokinin treatments, as well as thepresence of the auxin-responsive element in the OAT promoter(see Suppl. Table 3) allow us to assume specific regulation ofthe OAT gene during root growth and development. Recentfindings show the importance of nutrient and especially nitrogensignaling for root development and its interplay with hormoneregulation. Thus, cytokinins negatively regulate uptakeof nitrogen, but enhance nitrate distribution and translocation. Auxin level was shown to be elevated in rootsof plants growing on a low-nitrogen medium, while in roots ofplants growing on a medium with high nitrate concentrationthe auxin level was decreased. The reduction of auxin contentcorrelated with the degree of inhibition of root growth andlateral root development . The root growthregulation is associated with local reorganization of nitrogenmetabolism . Thereby, OAT may play animportant role in hormone-dependent fine-tuning of nitrogenmetabolism during the process of root development.The data regarding the OAT transcription activation in a widespectrum of experimental conditions, which was shown inour experiment and other studies, support the hypothesis thatornithine aminotransferase provides a link between differentbiochemical pathways of nitrogen conversion and contributesto the complicated signaling network regulating plantdevelopment.The authors declare no conflict of interest.Ballas N., Wong L.M., Theologis A. Identification of the auxin-responsiveelement, AuxRE, in the primary indoleacetic acid-inducible gene,PS-IAA4/5, of pea (Pisum sativum). J. Mol. Biol. 1993;233:580-596.DOI 10.1006/jmbi.1993.1537.Biancucci M., Mattioli R., Forlani G., Funck D., Costantino P., TrovatoM. Role of proline and GABA in sexual reproduction of angiosperms.Front. Plant Sci. 2015a;6:680. 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+{"text": "Estrogen receptor beta signaling in CD8+ T cells boosts T cell receptor activation and antitumor immunity through a phosphotyrosine switch. J Immunother Cancer 2021;9:e001932. doi:10.1136/jitc-2020-001932Yuan B, Clark CA, Wu B, This article has been corrected since it first published. The provenance and peer review statement has been added."}
+{"text": "DTG/3TC is a complete 2-drug regimen (2DR) for the treatment of HIV-1 infection. Non-inferior virologic efficacy has been proven over 3 years in treatment-naive people living with HIV (PLWH) and 2 years in a stable switch setting. TANGO, a randomized, open-label, non-inferiority study, evaluates efficacy and safety of switching to DTG/3TC in PLWH who are virologically suppressed  vs remaining on a 3- or 4-drug TAF-based regimen (TBR), stratified by baseline 3rd agent class. Week 144 analyses assessed non-inferiority (NI) with a 4% NI margin for Snapshot virologic failure (VF) and 8% for virologic success .P=0.044 (2-sided). Snapshot VS was high in both arms and demonstrated non-inferiority (Table). Zero pts on DTG/3TC and 3 (0.8%) on TBR met confirmed virologic withdrawal criteria with no resistance observed. Zero pts on DTG/3TC and 6 (1.6%) on TBR discontinued for lack of efficacy. Overall AE rates were similar between arms (Table). TC, LDL-C, and triglycerides improved with DTG/3TC, HDL-C improved with TBR, with no difference in TC/HDL-C ratio between arms. Changes in eGFR (cystatin C) and proximal tubular function marker were similar across arms. Adjusted mean change from BL in weight was 2.2 and 1.7 kg in the DTG/3TC and TBR arms, respectively, and proportion of pts with > 10% weight increase was similar across arms .Of 741 randomized/exposed pts , most pts entered the study on EVG/c (66%). For Week 144 Snapshot VF, switching to DTG/3TC was non-inferior to continuing TBR in the ITT-E analysis: 0.3% vs 1.3%; adjusted difference (95% CI): \u22121.1%  and superior to TBR in the per-protocol analysis: 0% vs 1.1%; adjusted difference: \u22121.1% ; Table. Efficacy and Key Safety Results for the ITT-E and Safety PopulationSwitching to the 2-drug regimen of DTG/3TC from a TAF-based 3- or 4-drug regimen resulted in high, non-inferior efficacy with zero confirmed virologic withdrawals and good tolerability over 3 years of treatment. DTG/3TC 2DR is a robust switch option with durable efficacy, good safety and tolerability, and a high barrier to resistance.Olayemi Osiyemi, M.D, Gilead Merck (Advisor or Review Panel member)ViiV Healthcare  Fiona Bisshop, MBBBS, Gilead (Grant/Research Support)ViiV Healthcare (Grant/Research Support) St\u00e9phane De Wit, MD, Gilead (Grant/Research Support)Janssen (Grant/Research Support)Merck Sharpe & Dohme (Grant/Research Support)ViiV Healthcare (Grant/Research Support) Joaqu\u00edn Portilla, MD, AbbVie Gilead Janssen Merck Sharpe & Dohme ViiV Healthcare  Jean-Pierre Routy, MD, FRCPC, ViiV Healthcare (Grant/Research Support) Mounir Ait-Khaled, PhD, ViiV Healthcare (Employee) Keith Pappa, PharmD, Glaxo Smith Kline (Shareholder)ViiV Healthcare (Employee) Ruolan Wang, Master of Science, ViiV Healthcare (Employee) Peter Leone, MD, viiv healthcare (Employee) Jonathan Wright, MSc, GlaxoSmithKline  Brian Wynne, MD, ViiV Healthcare  Jean A. van Wyk, MB,ChB, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Michael Aboud, MBChB, MRCP, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Kimberly Smith, MD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee)"}
+{"text": "The gut microbiota is associated with diverse age-related disorders. Several rejuvenation methods, such as probiotic administration and faecal microbiota transplantation, have been applied to alter the gut microbiome and promote healthy ageing. Nevertheless, prolongation of the health span of aged mice by remodelling the gut microbiome remains challenging.Akkermansia and the butyrate biosynthesis pathway in the rejuvenated mouse group. Furthermore, oral administration of Akkermansia sufficiently ameliorated the senescence-related phenotype in the intestinal systems in aged mice and extended the health span, as evidenced by the frailty index and restoration of muscle atrophy.Here, we report the changes in gut microbial communities and their functions in mouse models during ageing and three rejuvenation procedures including co-housing, serum-injection and parabiosis. Our results showed that the compositional structure and gene abundance of the intestinal microbiota changed dynamically during the ageing process. Through the three rejuvenation procedures, we observed that the microbial community and intestinal immunity of aged mice were comparable to those of young mice. The results of metagenomic data analysis underscore the importance of the high abundance of Akkermansia. Our results provide a rationale for developing therapeutic strategies to achieve healthy active ageing.In conclusion, the changes in key microbial communities and their functions during ageing and three rejuvenation procedures, and the increase in the healthy lifespan of aged mice by oral administration of Video abstractThe online version contains supplementary material available at 10.1186/s40168-021-01189-5. Health span is determined by the interactions between genetic and environmental factors , 2. In pAkkermansia muciniphila (AK) induces mucus production in the gut, which is critical for supporting intestinal integrity and other beneficial symbioses  or aged  mice were systemically treated with serum (100 \u03bcL per mouse) isolated from young mice by intravenous injection into the tail vein eight times (for 3 weeks) or sixteen times (for 6 weeks).Young ; (ii) hetero-chronic including hetero-young  and hetero-aged ; and (iii) iso-aged . After the suturing procedure, the mice were monitored daily for 6 weeks for signs of pain and distress. Body weight was recorded once per week. Circulatory exchange between parabiotic pairs was confirmed by injecting Evans blue dye /), and filtered through a strainer. Antibodies were purchased from BD Biosciences or BioLegend. BM cells were stained as previously described ; DAT, D-alanine transaminase [EC:2.6.1.21]; glutaryl-CoA dehydrogenase [EC:1.3.8.6]; lysine 2,3-aminomutase [EC:5.4.3.2]; beta-lysine 5,6-aminomutase [EC:5.4.3.3]; trans-2-enoyl-CoA reductase [EC:1.3.1.38]; acyl-CoA thioesterase YciA [EC:3.1.2.-]; PRPP, phosphoribosyl pyrophosphate; phosphoribosyl-ATP pyrophosphohydrolase [EC:3.6.1.31]; phosphoribosyl-AMP cyclohydrolase [EC:3.5.4.19]; phosphoribosylformimino-5-aminoimidazole carboxamide ribotide isomerase [EC:5.3.1.16]; imidazoleglycerol-phosphate dehydratase [EC:4.2.1.19]; histidinol-phosphate aminotransferase [EC:2.6.1.9]; histidinol-phosphatase (PHP family) [EC:3.1.3.15]; histidinol dehydrogenase [EC:1.1.1.23]; histidine ammonia-lyase [EC:4.3.1.3]; urocanate hydratase [EC:4.2.1.49]; imidazolonepropionase [EC:3.5.2.7]; GAD, glutamate decarboxylase [EC:4.1.1.15]; pectinesterase [EC:3.1.1.11]; pectate lyase [EC:4.2.2.2]; pectate disaccharide-lyase [EC:4.2.2.9]; oligogalacturonide lyase [EC:4.2.2.6]; glucuronate isomerase [EC:5.3.1.12]; tagaturonate reductase [EC:1.1.1.58]; altronate hydrolase [EC:4.2.1.7]; mannonate dehydratase [EC:4.2.1.8]; fructuronate reductase [EC:1.1.1.57]; L-xylulokinase [EC:2.7.1.53]; 3-dehydro-L-gulonate-6-phosphate decarboxylase [EC:4.1.1.85]; L-xylulokinase [EC:2.7.1.53]; L-ribulokinase [EC:2.7.1.16]; L-arabinose isomerase [EC:5.3.1.4]; xylulokinase [EC:2.7.1.17]; L-xylulose reductase [EC:1.1.1.10]; aldehyde reductase [EC:1.1.1.21]; D-xylulose reductase [EC:1.1.1.9]; rhamnulokinase [EC:2.7.1.5]; rhamnulose-1-phosphate aldolase [EC:4.1.2.19]; FabD, [acyl-carrier-protein] S-malonyltransferase [EC:2.3.1.39]; FabH, 3-oxoacyl-[acyl-carrier-protein] synthase III [EC:2.3.1.180]; FabG, 3-oxoacyl-[acyl-carrier protein] reductase [EC:1.1.1.100]; FabZ, 3-hydroxyacyl-[acyl-carrier-protein] dehydratase [EC:4.2.1.59]; FabK, enoyl-[acyl-carrier protein] reductase II [EC:1.3.1.-]; FabL, enoyl-[acyl-carrier protein] reductase III [EC:1.3.1.-]; FabF, 3-oxoacyl-[acyl-carrier-protein] synthase II [EC:2.3.1.179]; FabB, 3-oxoacyl-[acyl-carrier-protein] synthase I [EC:2.3.1.41]; YciA, acyl-CoA thioesterase [EC:3.1.2.-]; GSR, glutathione reductase (NADPH) [EC:1.8.1.7]; phospholipid-hydroperoxide glutathione peroxidase [EC:1.11.1.12]; PepA, leucyl aminopeptidase [EC:3.4.11.1]; PepD; dipeptidase D [EC:3.4.13.-]; PepN; aminopeptidase N [EC:3.4.11.2]; glutamate--cysteine ligase [EC:6.3.2.2]; GAD, glutamate decarboxylase [EC:4.1.1.15]; 2-hydroxyglutarate dehydrogenase [EC:1.1.99.2]; glutaconate CoA-transferase [EC:2.8.3.12]; ptb; phosphate butyryltransferase [EC:2.3.1.19]; buk; butyrate kinase [EC:2.7.2.7]; trans-2-enoyl-CoA reductase [EC:1.3.1.38]; acyl-CoA thioesterase YciA [EC:3.1.2.-]. h, Schematic summary shows key metabolic differences between young and aged group based on relative gene abundance profiles. Host metabolism is influenced by \u03b3-aminobutyric acid (GABA) neurotransmitter, affecting the brain (inducing satiety) GABA modulates inflammation. Fermentation of pectin by intestinal-specific bacteria produces butyrate. It affects host metabolism in several ways by acting on the G protein-coupled receptor (GPR) expressed by intestinal endocrine cells. Butyrate stimulates the release of glucagon-like peptide 1 (GLP-1) and peptide YY (PYY), affecting the pancreas (inducing insulin secretion) and brain (inducing satiety). Lipopolysaccharides (LPS) derived from the membrane of Gram-negative bacteria are pro-inflammatory compounds. AMUC_1100 derived from Akkermansia muciniphila improves intestinal barrier function by increasing goblet cell density and stimulating Toll-like receptor 2 (TLR2). AMUC_1100 exerts the beneficial effect on mucus layer regeneration, inflammation, and insulin sensitivity. Arrow heads indicate stimulation and bar heads indicate inhibition. Fig. S5. Parabiosis experiments restores intestinal function, canonical Wnt signalling target genes, and genes regulating ISC function. a, Verification of blood sharing between the parabiotic pairs using Evans blue. Representative photographs of the mice were taken at 0.5 and 24 h after the injection of Evans blue into the tail vein of one parabiont in a pair a week after surgery. The serum concentration of Evans blue in both mice in each pair was measured at 620 nm by spectrophotometry at 0, 0.5, 1, 24, and 48 h after injection. b, Gene expression profile of mucin in the colon. c, Gene expression profile of barrier-forming tight junction proteins in the colon. d\u2013g, Quantitative real-time PCR analyses for expression of canonical Wnt signalling target genes and genes regulating ISC function in ageing, cage model, serum injection model and parabiosis model. Data are means \u00b1 SEMs . ab means not sharing a common letter are significantly different at P < 0.05. h, Representative Ki67-stained pictures. Scale bar, 25 \u03bcm. i, quantification of Ki-67 positive cells per crypt base columnar cell in the colon. Data are means \u00b1 SEM. . Fig. S6. Dynamics of \u03b1-diversity indices including (a) Shannon diversity, (b) observed OTU, (c) Faith's phylogenetic diversity, (d) Pielou's evenness indices among samples of the co-housing, parabiosis, and serum injection groups. Statistical analysis was performed using Kruskal-Wallis test, young (week 20) versus rejuvenated mice group ; aged (W100) versus rejuvenated mice group . Abbreviations: Co-Y, young mice from co-housing experiments; Co-A, aged mice from co-housing experiments; Hetero-Y, young mice from heterochronic pairs; Hetero-A, aged mice from heterochronic pairs; Iso-Y, young mice from isochronic young pairs; Iso-A, aged mice from isochronic aged pairs; iv 8 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 16 times; iv 8 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 16 times. Fig. S7. Gut microbiome alteration in several rejuvenation models. a\u2013b, Principal coordinate analysis (PCoA) of \u03b2-diversity based (a) Jaccard distances, and (b) Bray-Curtis dissimilarity metric among samples of the co-housing (left), parabiosis (middle) and serum injection (right) groups of mice analysed . Each dot represents an individual mouse. c, Heatmap presenting the relative abundance (%) of the key prevalent bacterial taxa found in young and aged-mice. The red and blue genus indicates enriched bacterial taxa in young mice (W20) and aged mice (W100), respectively. Fig. S8. LEfSe analysis showing microbial genus that was significantly different in abundance between (a) young mice (W20) and Co-Y, (b) young mice and Hetero-Y, (c) Hetero-Y mice and Iso-Y, (d) Hetero-A and Iso-A, (e) aged mice and iv 16 (Y\u2192A), (f) young mice and iv 8 (Y\u2192Y), and (g) young mice and iv 16 (Y\u2192Y). Abbreviations: Co-Y, young mice from co-housing experiments; Hetero-Y, young mice from heterochronic pairs; Hetero-A, aged mice from heterochronic pairs; Iso-Y, young mice from isochronic young pairs; Iso-A, aged mice from isochronic aged pairs; iv 8 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 16 times; iv 16 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 16 times. Abbreviations: Co-Y, young mice from co-housing experiments; Co-A, aged mice from co-housing experiments; Hetero-Y, young mice from heterochronic pairs; Hetero-A, aged mice from heterochronic pairs; Iso-Y, young mice from isochronic young pairs; Iso-A, aged mice from isochronic aged pairs; iv 8 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 16 times; iv 8 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 16 times. Fig. S9. Parabiosis experiments induce corticosterone levels in plasma. Plasma corticosterone levels in (a) na\u00efve mice (young and aged mice) and (b) heterochronic parabiotic paired mice. All data are means \u00b1 SEMs . Fig. S10. Taxonomic composition and difference during rejuvenation process determined by metagenomic sequencing. Several microbial genera were significantly different in abundance between (a) Co-A and aged, (b) Co-Y and young, (c) Hetero-A and aged, (d) Hetero-A and Iso-A, (e) Hetero-Y and Young, (f) iv 8 (Y\u2192A) and aged, (g) iv 16 (Y\u2192A) and aged, (h) iv 8 (Y\u2192Y) and young, and (i) iv 16 (Y\u2192Y) and young. Bacterial taxon showing a significant abundance of change  was only shown with average fold-change value at the genus level. Red and blue indicate significantly increased bacterial taxa in young mice of 16s rRNA sequencing data and mice of 16s rRNA sequencing data, respectively. The Benjamini and Hochberg's FDR control method was used to correct for multiple comparisons. Abbreviations: Co-Y, young mice from co-housing experiments; Co-A, aged mice from co-housing experiments; Hetero-Y, young mice from heterochronic pairs; Hetero-A, aged mice from heterochronic pairs; Iso-Y, young mice from isochronic young pairs; Iso-A, aged mice from isochronic aged pairs; iv 8 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 16 times; iv 8 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 16 times. Fig. S11. Rejuvenation-associated changes in microbial functional potential and metabolism. a, LEfSe analysis of metabolic pathways in colon microbiota during each rejuvenation procedure. The two columns on the left indicate logarithmic LDA scores categorised by KEGG pathways, comparing young group, Co-A, Iso-A, Hetero-A, iv 8 (Y\u2192A), and iv 16 (Y\u2192A) to the aged group. The last column on the right shows logarithmic LDA scores categorised by KEGG pathways, comparing Co-Y, Iso-Y, Hetero-Y, iv 8 (Y\u2192Y), and iv 16 (Y\u2192Y) to the young group. The red and blue names of pathways indicate enriched metabolic pathways in young mice and in aged mice, respectively. Only KEGG pathways differ significantly during each rejuvenation procedure. b\u2013e, Shown horizontal bar plots indicate the fold-change of each enzyme involved in (b) pentose and glucuronate interconversions, (c) lysine degradation, (d) glutathione metabolism, and (e) fatty acid biosynthesis in young, Co-A, Hetero-A, iv8 (Y\u2192A), iv16 (Y\u2192A), Co-Y, Hetero-Y, iv8 (Y\u2192Y), and iv16 (Y\u2192Y). Red and blue KEGG reactions indicate high abundance in young mice and in aged mice, respectively. Abbreviations: Co-Y, young mice from co-housing experiments; Hetero-Y, young mice from heterochronic pairs; Hetero-A, aged mice from heterochronic pairs; Iso-Y, young mice from isochronic young pairs; Iso-A, aged mice from isochronic aged pairs; iv 8 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 8 times; iv 16 (Y\u2192Y), young mice treated with serum isolated from young mice by intravenously into the tail vein 16 times; iv 16 (Y\u2192A), aged mice treated with serum isolated from young mice by intravenously into the tail vein 16 times. The following genes are represented by enzyme name and EC number: OADH, 2-oxoglutarate dehydrogenase (OADH) [EC:2.3.1.61]; DAT, D-alanine transaminase [EC:2.6.1.21]; glutaryl-CoA dehydrogenase [EC:1.3.8.6]; lysine 2,3-aminomutase [EC:5.4.3.2]; beta-lysine 5,6-aminomutase [EC:5.4.3.3]; trans-2-enoyl-CoA reductase [EC:1.3.1.38]; acyl-CoA thioesterase YciA [EC:3.1.2.-]; GAD, glutamate decarboxylase [EC:4.1.1.15]; pectinesterase [EC:3.1.1.11]; pectate lyase [EC:4.2.2.2]; pectate disaccharide-lyase [EC:4.2.2.9]; oligogalacturonide lyase [EC:4.2.2.6]; glucuronate isomerase [EC:5.3.1.12]; tagaturonate reductase [EC:1.1.1.58]; altronate hydrolase [EC:4.2.1.7]; mannonate dehydratase [EC:4.2.1.8]; fructuronate reductase [EC:1.1.1.57]; L-xylulokinase [EC:2.7.1.53]; 3-dehydro-L-gulonate-6-phosphate decarboxylase [EC:4.1.1.85]; L-xylulokinase [EC:2.7.1.53]; L-ribulokinase [EC:2.7.1.16]; L-arabinose isomerase [EC:5.3.1.4]; xylulokinase [EC:2.7.1.17]; L-xylulose reductase [EC:1.1.1.10]; aldehyde reductase [EC:1.1.1.21]; D-xylulose reductase [EC:1.1.1.9]; rhamnulokinase [EC:2.7.1.5]; rhamnulose-1-phosphate aldolase [EC:4.1.2.19]; FabD, [acyl-carrier-protein] S-malonyltransferase [EC:2.3.1.39]; FabH, 3-oxoacyl-[acyl-carrier-protein] synthase III [EC:2.3.1.180]; FabG, 3-oxoacyl-[acyl-carrier protein] reductase [EC:1.1.1.100]; FabZ, 3-hydroxyacyl-[acyl-carrier-protein] dehydratase [EC:4.2.1.59]; FabK, enoyl-[acyl-carrier protein] reductase II [EC:1.3.1.-]; FabL, enoyl-[acyl-carrier protein] reductase III [EC:1.3.1.-]; FabF, 3-oxoacyl-[acyl-carrier-protein] synthase II [EC:2.3.1.179]; FabB, 3-oxoacyl-[acyl-carrier-protein] synthase I [EC:2.3.1.41]; YciA, acyl-CoA thioesterase [EC:3.1.2.-]; GSR, glutathione reductase (NADPH) [EC:1.8.1.7]; phospholipid-hydroperoxide glutathione peroxidase [EC:1.11.1.12]; PepA, leucyl aminopeptidase [EC:3.4.11.1]; PepD; dipeptidase D [EC:3.4.13.-]; PepN; aminopeptidase N [EC:3.4.11.2]; glutamate--cysteine ligase [EC:6.3.2.2]; GAD, glutamate decarboxylase [EC:4.1.1.15]; trans-2-enoyl-CoA reductase [EC:1.3.1.38]; acyl-CoA thioesterase YciA [EC:3.1.2.-]. Fig. S12. Administration of AK restores the expression of the canonical Wnt signalling target genes and ameliorates the senescence-related phenotype in haematopoietic system. a, Quantitative reverse transcription PCR (qRT-PCR) analyses for the expression of canonical Wnt signalling target genes and genes regulating ISC function. Data are presented as means \u00b1 SEMs . b, A representative fluorescence-activated cell sorting plot showing the frequencies of LT-HSCs, ST-HSCs, and MPPs among LSKs in the bone marrow of AK-treated and untreated aged mice. c, Percentages of LT-HSCs, ST-HSCs, and MPPs among LSKs. Data are presented means \u00b1 SEMs . d, Representative images and e, frequencies of neutrophils, T cells, and B cells in the blood of AK-treated and untreated aged mice. Data are presented as means \u00b1 SEMs . f, qRT-PCR analysis of the Icam1 mRNA purified from bone niches and lineage-negative and -positive cells of the aged mice treated with AK. Data are presented as means \u00b1 SEMs . Fig. S13. Effect of AK treatment in aged mice. a, Principal coordinate analysis (PCoA) of \u03b2-diversity based Jaccard distances and Bray-Curtis dissimilarity metric between AK-treated and untreated aged mice . The gut microbiome of AK-treated aged mice shows significant differences with control (Table S2). b, Taxonomic composition and difference determined by metagenomic sequencing. Microbial genera were significantly different in abundance between aged-AK and aged-vehicle groups. Bacterial taxon showing a significant abundance of change  was only shown with average fold-change value at the genus level. Red and blue indicate significantly increased bacterial taxa in young mice of 16s rRNA sequencing data and mice of 16s rRNA sequencing data, respectively. The Benjamini and Hochberg's FDR control method was used to correct for multiple comparisons. c, Analysis of differentially abundant microbial genus between AK-treated and untreated aged mice were analysed by LEfSe . d, LEfSe analysis of KEGG enzymes in colon microbiota between AK-treated and untreated aged mice. No metabolic pathway is enriched between aged-AK and aged-vehicle groups. e, Survival rate and f, body weight change of AK-treated and untreated aged mice. g, Schematic summary shows TLR 2 and Wnt signalling pathways can be activated via the high abundance of Akkermansia in the aged-AK group compared to the aged-vehicle group. Lipopolysaccharides (LPS) derived from the membrane of Gram-negative bacteria are pro-inflammatory compounds. AMUC_1100 derived from Akkermansia muciniphila improves intestinal barrier function by increasing goblet cell density and stimulating Toll-like receptor 2 (TLR2). AMUC_1100 exerts a beneficial effect on mucus layer regeneration, inflammation, and insulin sensitivity. Wnt signalling is involved in the development and renewal of intestinal epithelium and hematopoietic stem cells. Arrow heads indicate stimulation and bar heads indicate inhibition.Additional file 2: Table S1. Read statistics of 16s rRNA and metagenome sequencing.Additional file 3: Table S2. Alpha and beta diversity analysis of the analysed groups in this study.Additional file 4: Table S3. Taxa identified by LEfSe analysis in this study. \"LogMaxMean\" indicates the log of the highest-class average. \"Enriched in\" indicates the class with the highest mean if the taxa are discriminative. \"LDA\" indicates the logarithmic of the linear discriminant analysis (LDA) score. \"P-value\" indicates p-value of the pairwise Kruskal-Wallis test performed by LEfSe.Additional file 5: Table S4. Effect of parabiotic pairing on blood parameters.Additional file 6: Table S5. Relative abundance of phylotypes detected in ageing and rejuvenation samples.Additional file 7: Table S6. Relative abundance of phylotypes detected in AK administration samples.Additional file 8: Table S7. Frailty score used to develop a clinical frailty index in mice.Additional file 9: Table S8. Sequences of PCR primers used in this study."}
+{"text": "The corrected table is shown here.Page 5: cas type IIA system and the type II restriction-modification system LmoJ3 (24) were negatively associated with nonpersistence\u201d should read \u201cIn contrast, genes associated with the CRISPR-cas type IIA system and the type II restriction-modification system LmoJ3 (24) were associated with nonpersistence.\u201dPage 11, line 10: \u201cIn contrast, genes associated with the CRISPR-Page 15: References 24 and 32 are interchanged. The correctly numbered references are as follows:Listeria monocytogenes strains. Appl Environ Microbiol 78:2623\u20132630. https://doi.org/10.1128/AEM.07203-11.24. Lee S, Ward TJ, Siletzky RM, Kathariou S. 2012. Two novel type II restriction-modification systems occupying genomically equivalent locations on the chromosomes of Listeria monocytogenes clones with enhanced virulence. Appl Environ Microbiol 83:e01189-17. https://doi.org/10.1128/AEM.01189-17.32. Lee S, Ward TJ, Jima DD, Parsons C, Kathariou S. 2017. The arsenic resistance-associated Listeria genomic island LGI2 exhibits sequence and integration site diversity and a propensity for three"}
+{"text": "Cladosporium species have attracted considerable interest because of their ability to produce a wide array of metabolites, including alkaloids, macrolides, diketopiperazines, pyrones, tetralones, sterols, phenolics, terpenes, lactones, and tetramic acid derivatives that possess versatile bioactivities. Moreover, they produce diverse enzymes with biotechnological and industrial relevance. This review gives an overview on the Cladosporium species derived from marine habitats, including their metabolites and bioactivities, as well as the industrial and biotechnological potential of these species. In the current review, 286 compounds have been listed based on the reported data from 1998 until July 2021. Moreover, more than 175 references have been cited.The marine environment is an underexplored treasure that hosts huge biodiversity of microorganisms. Marine-derived fungi are a rich source of novel metabolites with unique structural features, bioactivities, and biotechnological applications. Marine-associated  Cladosporium (Cladosporiaceae) is one of the largest genera of dematiaceous hyphomycetes [Cladosporium species are frequent airborne molds, which can be isolated from almost every environment and geographic location, because their small conidia are easily dispersed [C. herbarum, C. cladosporioides, and C. sphaerospermum are its three major species [C. fulvum is the causal agent of tomato leaf mold [C. sphaerospermum isolated from Glycine max roots which can promote its growth [Cladosporium species have the potential to be used in various industrial processes [Cladosporium species have attracted considerable interest because of their ability to produce a wide array of metabolites, including macrolides, pyrones, phenolics, alkaloids, diketopiperazines, terpenes, sterols, quinones, lactones, and tetramic acid derivatives. These metabolites possess versatile bioactivities such as anticancer, antimicrobial, antiviral, insecticidal, antifouling, anti-malarial, anti-hyperlipidemic, and \u03b1-glucosidase and protein tyrosine phosphatase inhibiton [Cladosporium species derived from a marine habitat, including the structures and bioactivities of the reported metabolites, as well as the industrial and biotechnological potential of these species  even in a non-marine environment [The polycyclic aromatic hydrocarbons (PAHs) are volatile pollutants that can cause various environmental pollutions such as oceanic and freshwater contamination, which can take place during storage, use, or transportation of crude oil and its products. PAHs inhalation or ingestion through contaminated food and airborne contaminants leads to serious health disorders such as endocrine disruption, cancer, and reproductive and birth problems . Therefoironment .C. cladosporioides using the Buescher and Furmanski procedure after 10-day incubation and precipitation with (NH4)2SO4 and benzoate buffer at pH 4.0 [Pectinases are hydrolytic enzymes that are accountable for the hydrolysis of pectins. They are commonly found in fungi, bacteria, and plants. They have remarkable importance in the food industry such as vegetables and fruits processing, wine production, and olive oil extraction, as well as coffee, cocoa, and tea fermentation. They are utilized in the beverage industry to produce high yields due to improving clarification and pressing of concentrated fruit juices . Bastos t pH 4.0 .Cladosporium sp. isolated from the Antarctic macroalgae Ascoseira mirabilis and Georgiella confuens produced agarase that may have industrial importance in the extraction of agar or its byproducts such as bioactive galactose and oligosaccharides exist in the algal biomass to be utilized as substrates of 3rd generation bioethanol [Agarases and carrageenases can decompose algal biomass, producing carrageenans and agars that are the major components of the red algae cell wall. Furthermore, agarases hydrolyze agar, resulting in oligosaccharides that are employed as food additives with beneficial influences on human health ,105. Addoethanol .Cladosporium sp. isolated from Antarctic marine sponge had high xylanase potential when grown on wheat bran and pure xylans at lower temperatures that is a feature of cold-active enzymes [Cladosporium sp. could be convenient for many biotechnological processes, utilizing moderate- to low-temperature processes, especially those in food industries [Cladosporium sp. derived from Antarctic sponge. XynA is highly active on xylans with high arabinose content. Moreover, it is the most thermolabile endo-xylanase reported from filamentous fungus. Therefore, it could be a good alternative in some biotechnological operations to avoid heating, thereby reducing the costs [Xylan, the main component of hemicelluloses in the plant cell walls, represents about one-third of all renewable organic carbon on earth. Xylanases hydrolyze xylan to oligosaccharides that are further degraded to xylose. The latter is utilized for xylitol and bioethanol production. Xylanases have remarkable biotechnological influence in developing eco-friendly technologies in the pulp and paper industry and in food and feed industries, and for generating chemicals and liquid fuels from lignocellulose ,108,109. enzymes . Therefodustries . Gil-Durhe costs .2O2-dependent glycoprotein that needs Mn2+ for oxidizing aromatic dyes and mono-aromatic phenols [2 reduction to H2O [C. cladosporioides CBMAI 857 isolated from the Brazilian cnidarian Palythoa variabilis produced ligninolytic enzymes  with particular response to the various conditions of salinity and carbon sources. It possessed high values of MnP and laccase activities under salinity , indicating the potential use of this fungus for industrial applications and bioremediation of high-salt contaminated sites [The three main lignin-hydrolyzing enzymes that have great potential for industrial applications are LiP (lignin peroxidase), MnP (manganese-dependent peroxidase), and Lac (laccase) . LiP is  phenols . Laccasen to H2O . C. claded sites .C. cladosporioides CBMAI-857 associated with the coral Palythoa caribaeorum was tested for its RBBR decolorizing potential. It had efficient dye decolorization potential (93%) after 12 days in both liquid and solid media [Cladosporium sp. associated with the seagrass Posidonia oceanica produced tannases and ligninolytic enzymes at high salt concentrations. Its laccase and peroxidase activity was evident by the degradation of RBBR and Amaranth Red dyes [RBBR (Remazol Brilliant Blue R) and polymeric dyes decolorization has been assigned as an effective screening method for the fungi ability to degrade recalcitrant pollutants, including aromatic compounds such as PAHs. It was demonstrated that marine-derived fungi are often more effective than terrestrial fungi in treating various colored effluents because they are better adapted to perform under extreme conditions such as high salinity . C. cladid media . FurtherRed dyes .C. herbarum ER-25 possessed a high invertase potential and removed melanoidins from molasses through bio-adsorption and biodegradation mechanisms by Lac and MnP in the non-sterilized medium than in sterilized one at 5.5 pH and 20 \u00b0C. Therefore, this cold-adapted fungus can be used for molasses de-colorization [Invertase is a \u03b2-fructo-furanosidase that catalyzes sucrose conversion into fructose and glucose, giving invert syrup. This invert syrup is utilized in the beverage and food industries as a humectant in non-crystallizing creams, candies, artificial honey, and jam preparation . Molasserization .C. sphaerospermum obtained from deteriorated seaweed Ulva through SSF (solid-state fermentation) produced cellulase that had saccharification potential of seaweed biomass using green seaweed Ulva\u00a0fasciata. Therefore, this cellulase can be utilized for saccharification of cellulosic feedstock for bioethanol production from marine macro-algal feedstock [Cellulose is a main component of the plant material that is abundantly utilized for the production of alternative liquid fuels such as bioethanol. eedstock .Cladosporium sp. CBMAI 1237 isolated from Dragmacidon reticulatum, revealing the existence of ene-reductases [C. cladosporioides CBMAI-857 isolated from the Brazilian cnidarian Palythoa caribaeorum catalyzed the asymmetric bio-reduction of 1-(4-methoxyphenyl)ethanone to 1-(4-methoxyphenyl)ethanol [C. cladosporioides CBMAI-857 catalyzed the enantio-selective bio-reduction of different aromatic ketones at pH 7.0 and 32 \u00b0C [Biocatalysis is an eco-friendly process for renewable raw materials and clean energy production and for the remediation of environmental contaminants . Recentlductases . Additiond 32 \u00b0C .Cladosporium species are rich with diverse types of metabolites with varied structural features such as macrolides, fatty acids, pyrones, phenolics, alkaloids, diketopiperazines, terpenes, sterols, quinones, lactones, and tetramic acid derivatives. Their classification was carried out here according to the chemical nature. During our search, it was found that some of the reported metabolites had the same structures and molecular formulae with different nomenclature. On the other hand, some metabolites had the same names with different structures. Moreover, some metabolites did not have names, thus they are named here using the AUPAC system for nomenclature. Herein, the reported secondary metabolites from Cladosporium species, as well as their bioactivities have been discussed  of them are from C. sphaerospermum.Tetramic acids are five-membered heterocycles with a pyrrolidine-2,4-dione core that are formed by the fusion of polyketide units and amino acid . The terivatives . These sivatives . They arivatives ,128. Theivatives . Note th1, 2, 4, and 5) biosynthesized by C. sphaerospermum 2005-01-E3 obtained from deep-sea sludge had no activity towards influenza A H1N1 virus (3 exhibited anti-H1N1 activity (IC50 276.0 \u03bcM) in comparison to ribavirin (IC50 131.0 \u03bcM) [Mycobacterium tuberculosis in the disk diffusion method [2), C (3), F (5), and L (11) separated from C. sphaerospermum SW67 associated with Hydractinia echinat hydroid polyp were assessed for protection towards cisplatin-caused cell damage in LLC-PK1 cells [2 and 5 alleviated the LLC-PK1 cells damage induced by cisplatin (Conc. 25 \u00b5M). Compound 2 (Conc. 100 \u00b5M) recovered cell viability with 90.68% that was more than NAC , whereas 5 (Conc. 50 and 100 \u03bcM) increased cell viability by 77.65 and 85.60%, respectively. Thus, 2 may be a candidate for treating cisplatin-produced unwanted effects and/or to prohibited nephrotoxicity induced by anticancer drugs. It was proposed that the existence of the C-8 hydroxy group may be essential for the reno-protective effect towards cisplatin-produced toxicity in LLC-PK1 cells [The tetramic acid derivatives, cladosins A, B, D, and E (N1 virus . While 331.0 \u03bcM) . Moreoven method . MoreoveK1 cells . The co-K1 cells .5 and 6 from C. sphaerospermum 2005-01-E3 that did not have anti-influenza A H1N1, anticancer, and anti-tubercular, as well as no NF-\u03baB inhibitory activities [C. sphaerospermum WBS017 isolated from Fritillaria unibracteata var. wabuensis [7\u201310) and cladodionen (13) were isolated from sediment-derived C. sphaerospermum L3P3 and evaluated for cytotoxic capacity towards PC-3, MGC-803, SH-SY5Y, and HCT-116 cell lines using SRB method and against K562 and HL-60 using MTT method  was inactive. The results revealed that the C-8 absolute configuration and aniline moiety were essential for activity [In 2015, by OSMAC (one strain many compounds) technique, Yu et al. separated compounds tivities . Note thabuensis . CladosiT method . Compounactivity  , compared to etoposide (IC50 ranged from 1.76 to 2.27 \u03bcM) [, and 30 . Compoun2.27 \u03bcM) .C. sphaerospermum EIODSF 008 isolated from the deep-sea sediment collected from the East Indian Ocean yielded tetramic acid derivatives 13 and 22\u201328 (13 had cytotoxicity (IC50 28.6 \u00b5M) towards the HL-60 cell line [E. coli, M. luteus, and B subtilis [13 showed cytotoxic capacity towards HL-60, HeLa, HCT-116, and MCF-7 cell lines (IC50 ranged from 9.1 to 19.1 \u03bcM), compared to ADR (adriamycin) (IC50 ranged from 0.02 to 0.67 \u03bcM). However, it did not have antibacterial activities (conc. 100 \u03bcg/mL) against B. subtilis, P. aeruginosa, C. perfringens, S. aureus, E. coli, and C. albicans [14\u201321, new tetramic acid derivatives, were purified from the sea-sediment derived Cladosporium sp. acetone extract by Huang et al. in 2018. Compounds 14\u201316 are unusual 3-acyltetramic acids, having at C-3 of the pyrrolidine-2,4-dione core, a six-membered lactone ring, and hexyl-enic alcohol chain. They showed no obvious AchEI activity in the modified Ellman\u2019s enzyme assay [C. albicans and S. aureus in the broth micro-dilution method and no cytotoxic effect towards HL60, HepG-2, and MCF-7 cell lines in the CCK8 assay [nd 22\u201328 . They weell line . Additiosubtilis . Additioalbicans . Compounme assay . MoreoveK8 assay .Diketopiperazines (DKPs) are cyclic dipeptides, consisting of two amino acids with or without extra structural modifications in the DKPs nucleus . Their m32) and cyclo-(Phe-Pro) (33) were separated from the EtOAc extract of Cladosporium sp. F14 isolated from seawater and investigated for their anti-larval activity at conc. 50 \u00b5g/mL towards Bugula neritina and Balanus amphitrite larvae in the settlement inhibition assays [B. neritina settlement  and B. amphitrite settlement . Furthermore, 32 and 33 obviously prohibited L. hongkongensis growth , compared to streptomycin (MIC 250 \u00b5g/mL). The MICs of 33 towards Ruegeria sp. and M. luteus were 200 and 100 \u00b5g/mL, respectively, compared to streptomycin  [36) and B (37), and haematocin (38) purified from the sediment-derived Cladosporium sp. were moderately cytotoxic towards HepG2 cell line  [Cyclo-  . On the ctively) .Cladosporium species.Fungal alkaloids are nitrogen-containing metabolites that are derived from amino acid metabolism and the mevalonate pathway . Many st42\u201355, were separated from Cladosporium sp. PJX-41 isolated from mangrove and assessed for anti-H1N1 activity using CPE (cytopathic effect) inhibition assay , compared to ribavirin (IC50 87 \u03bcM), while 42\u201344, 46\u201348, 50, and 54 (IC50 100\u2013150 \u03bcM) had weak activity [Cladosporium sp. associated with Chondria crassicualis red alga afforded 56 that exhibited antioxidant potential (ED50 82.0 \u00b5M) more than oxybenzone  as evident by their UV-A protecting potential [S. aureus and S. aureus with MICs 31.0, 62.5, and 62.5, \u00b5g/mL, respectively [58, 68, and 70 separated from C. oxysporum were assessed for anti-plasmodial potential towards chloroquine-sensitive Plasmodium falciparum 3D7 [58 (conc. 3.13 \u00b5g to 25.0 \u00b5g) had an anti-plasmodial effect (EC50 24.8 \u00b5M), while 68 and 70 displayed no activity (EC50 > 25.0 \u00b5M), compared to artesunate (EC50 0.074 \u03bcM) in the SYBR Green I assay. Further, 58 (conc. ranged from 6.25 \u00b5M to 50.0 \u00b5M for 24 h) was investigated for apoptotic effect on 3D7-plasmodia strain by measuring the parasite \u0394\u03a8m . It induced loss of \u0394\u03a8m, leading to the release of cytochrome C from mitochondria to the cytosol resulted in parasite apoptosis. Therefore, it may provide a scaffold to apoptotic death in the stages of P. falciparum development [58, 68, and 70 had no anti-buruli ulcer activity against Mycobacterium ulcerans (IC50 \u02c3 10 \u00b5M), compared to rifampicin (IC50 \u02c2 1 \u00b5M) in the Resazurin microtiter assay [The glyantrypine-type alkaloids, on assay . Compounactivity . The mycotential . Furtherectively . The quiarum 3D7  , compared to curcumin . However, they showed moderate activity versus LNCap and LNCap , in comparison to curcumin  in the MTT assay [Cladosporium sp. HNWSW-1 associated with the mangrove plant Ceriops tagal biosynthesized compounds 74\u201376 that were assessed for their cytotoxic and \u03b1-glycosidase inhibitory effects , whereas 76 revealed cytotoxic potential towards BEL-7042 and Hela cell lines  in the MTT assay.They had significant activity towards HepG-2 and MCF-7 (ICTT assay . Cladosp effects . Compoun76 exhibited \u03b1-glucosidase inhibitory activity (IC50 78.2 \u00b5M), compared to acarbose (IC50 275.7 \u00b5M) in the glucose oxidase method [77) separated from Cladosporium sp. TPU1507 derived from marine sponge was assessed for its inhibitory effect towards PTP1B (protein tyrosine phosphatase) and TCPTP (T-cell PTP), using an enzyme-based assay [50 48 and 54 \u03bcM, respectively), in comparison to oleanolic acid (IC50 0.9 \u03bcM) [78, from a gorgonian-derived Cladosporium sp. collected from the South China Sea. It (IC50 0.76\u20133.11 \u03bcM) exhibited significant cytotoxicity towards HeLa, P388, HT-29, and A549 cell lines [B. cereus, T. halophilus, S. epidermidis, S. aureus, E. coli, P. putida, N. brasiliensis, and V. parahaemolyticus [Cladosporium sp. SCNU-F0001 isolated from a mangrove plant yielded a novel lactam macrolide named cladospamide A (79) that was evaluated for cytotoxic effect (conc. 50 \u03bcM) versus MDA-MB-435, A549, HCT116, HepG2, and BT549 in the MTT method and for antimicrobial potential (conc. 100 \u03bcg/mL) towards S. aureus, B. subtilis, E. coli, Salmonella ATCC 14028, and P. aeruginosa. Unfortunately, it exhibited no noticeable activity [Additionally, e method . Cladosped assay . It had  0.9 \u03bcM) . Cao et ll lines . On the olyticus . Cladospactivity .80) and B (81) purified from Cladosporium sp. SCSIO z015 broth did not have an obvious anti-biofilm activity towards S. aureus, E. coli, and B. subtilis [The new cyano-containing alkaloids, cladosporins A , compared to ascorbic acid (IC50 4.9 \u00b5M). Besides, they showed moderate toxicity towards brine shrine naupalii , compared with toosendanin (LC50 21.2 \u00b5M) in the brine shrimp lethality assay [84 and 85 from Cladosporium sp. JS1-2 isolated from the mangrove Ceriops tagal collected in the South China Sea. Compound 84 moderately prohibited the growth of Helicoverpa armigera Hubner newly hatched larvae (IC50 100 \u03bcg/mL), compared to azadirachtin (IC50 25 \u03bcg/mL). Further, they showed moderate antibacterial potential versus S. aureus with MICs 12.5 and 25.0 \u03bcg/mL, respectively, compared with ciprofloxacin (MIC 0.39 \u03bcg/mL) [In the DPPH assay, they also had no activity (ICty assay . In 20199 \u03bcg/mL) .91), together with 88 and 89 were separated from Cladosporium sp. FT-0012 was obtained from Pohnpei Island, Federated State of Micronesia, and assessed for antimicrobial activity using paper disks at conc. 10 \u00b5g/disk , while 88 was active (IZD 14.0 mm and IC50 17.0 \u00b5g/mL) towards X. campestris pv. oryzae. Moreover, 91 prohibited P. oryzae and M. racemosus growth  [The term \u201cmacrolides\u201d was first used to describe the natural antibiotics that have 12\u201316-membered macrocyclic lactone ring, functionalized by double bonds, and carrying different aminosaccharide and saccharide components . Among t \u00b5g/disk . Compounctively) .Cladosporium sp. F14 isolated from seawater yielded a nine-membered macrolide, 92 that had weak antibacterial potential towards M. smegmatis, E. coli, B. thuringiensis, S. aureus, and B. subtilis and weak cytotoxic potential toward A435, HeLa, K562, and A549 in the MTT method [C. herbarum isolated from Callyspongia aerizusa sponge yielded cladospolide B (89) and pandangolides 2\u20134 (95\u201397) that showed no antimicrobial potential versus S. aureus ATCC 25923, B. subtilis 168, E. coli ATCC 25922, and C. albicans in the agar plate diffusion assay [Cladosporium sp. IFB3lp-2 isolated from the mangrove forest of Hainan province of China yielded 88, 89, 93\u201396, 100, and 116 that had no significant activity against HCT-116, Coxsachievirus A16, A549, MD-MBA-231, HepG2, human enterovirus 71, A375, and SW1116 cell lines (conc. 20 \u00b5M) in the MTT assay [T method . C. herbon assay . MoreoveTT assay  , along with 89 that were evaluated for cytotoxic effect (conc. 50 \u03bcM) versus MDA-MB-435, A549, HCT116, HepG2, and BT549 in the MTT method and for antimicrobial potential (conc. 100 \u03bcg/mL) towards S. aureus, B. subtilis, E. coli, Salmonella ATCC 14028, and P. aeruginosa. Unfortunately, none of them exhibited noticeable activity [89 and 117\u2013120 from a gorgonian-derived Cladosporium sp. collected from the South China Sea. They showed no cytotoxicity towards HeLa, P388, HT-29, and A549 cell lines [B. cereus, T. halophilus, S. epidermidis, S. aureus, E. coli, P. putida, N. brasiliensis, and V. parahaemolyticus. Compounds 117\u2013119 exhibited antibacterial potential against all tested bacteria , however 89 and 120 had weak activity (MIC \u02c3 25.0 \u03bcM) [93 and 115 separated from Cladosporium sp. F14 at conc. 50 \u00b5g/mL had no anti-larval activity towards both B. neritina and B. amphitrite larvae in the settlement inhibition assays [98 and 99 and a known analog 93 from the rice culture EtOAc extract of C. cladosporioides associated with Bruguiera gymnorrhiza. Their configuration was established using ECD, modified Mosher\u2019s, and X-ray diffraction methods, as well as optical rotations to be 5R, 11R for 98; 11R for 99; and 3R, 5S, 11S for 93. They had weak AChEI activity (IC50 > 50 \u00b5M), in comparison to tacrine in the modified Ellman\u2019s method [C. cladosporioides MA-299 obtained from the mangrove plant B. gymnorrhiza yielded 12-membered thio-macrolides 96 and 101\u2013104 that were assessed for antimicrobial potential against E. tarda QDIO-2 and E. ictarda QDIO-9 (aquatic pathogens) and C. glecosporioides QDAU-2, B. sorokiniana QDAU-5, P. piricola Nose QDAU-15, and F. oxysporum f. sp. cucumerinum QDAU-8 (plant pathogenic fungi) in the microtiter plates assay. All metabolites revealed activity against C. glecosporioides (MIC 1 or 2 \u03bcg/mL), compared to amphotericin B (MIC 0.5 \u03bcg/mL). Moreover, 101 and 104 showed noticeable activity (MIC 1.0 \u03bcg/mL) towards with E. tarda and E. ictarda, respectively, compared to chloramphenicol (MIC 0.5 \u03bcg/mL), while 102 and 104 exerted obvious effectiveness (MIC 1.0 \u03bcg/mL) versus F. oxysporum f. sp. cucumerinum, compared to amphotericin B (MIC 0.5 \u03bcg/mL). The data revealed that sulfur substituent may influence the macrolides\u2019 bioactivities [107\u2013112 and the related formerly reported 93 and 101 isolated from mangrove-derived C. oxysporum HDN13-314 had no cytotoxic activity versus HCT-116, BEL-7402, HL-60, A549, L-02, HeLa, K562, MGC-803, MCF-7, PC-3, SH-SY5Y, and MDA-MB-231 (IC50 > 50 \u03bcM) [Additionally, activity . Cao et ll lines . Further25.0 \u03bcM) . The metn assays . In 2019tivities . The new> 50 \u03bcM)  , whereas 108 had the best effect (MIC 4 \u03bcg/mL) versus E. tarda [106) and G (108) and cladocladosin A (121), a macrolide with bicyclo 5/9-ring, were purified from C. cladosporioides MA-299 by Zhang et al. and assessed for antimicrobial effect versus various plant, human, and aquatic pathogenic microbes in the microtiter plates assay. All metabolites revealed activity (MIC ranging from 1.0 to 4.0 \u03bcg/mL) towards V. anguillarum and E. tarda (aquatic pathogenic bacteria) [Additionally, they exerted antibacterial activities versus the aquatic pathogens E. tarda . In 2020acteria) .108 and 121 exerted activity (MICs 4.0 \u03bcg/mL) towards H. maydis (plant-pathogenic fungus) and P. aeruginosa (aquatic-pathogenic bacterium), respectively [113 and 114, purified from Cladosporium sp. L037 isolated from the Okinawan marine brown alga Actinotrichia fragilis exhibited cytotoxic influence  towards L1210 murine lymphoma cells in the MTT assay [113 had antifungal potential against C. albicans, C. neoformans, A. niger, and N. crassa (MICs 8.4\u201316.7 \u00b5g/mL), whereas 114 exhibited antibacterial activity only towards M. luteus and inactive against the other microorganisms [Moreover, ectively . The newTT assay . Moreoverganisms .Butanolides and butenolides are five-membered \u03b3-lactones which may also be regarded as furan derivatives. They are an important class of structural motifs often encountered in various natural metabolites and synthetic targets . They ha122), purified from a soft coral-associated fungus Cladosporium sp. TZP-29, together with the formerly isolated derivative 126 showed no cytotoxic effect towards A-549, SMMC-7721, and HeLa cells in the SRB method [126 isolated from Cladosporium sp. IFB3lp-2 exhibited no significant activity against HCT-116, Coxsachievirus A16, A549, MD-MBA-231, HepG2, human enterovirus 71, A375, and SW1116 cell lines (Conc. 20 \u00b5M) in the MTT assay [B. cereus, T. halophilus, S. epidermidis, S. aureus, E. coli, P. putida, N. brasiliensis, and V. parahaemolyticus  [126 displayed no anti-larval activity towards both B. neritina and B. amphitrite larvae in the settlement inhibition assays [E. ictarda and Cytospora mandshurica Miura (MIC 8 \u03bcg/mL) [123, 124, and 127 and the known analog 126 separated from C. cladosporioides were assessed for AChEI activity using modified Ellman\u2019s method  [The newly separated C12-macrolide, cladospolide F  . Qi et an assays . Additios method . Only 12128, 130, and 141 isolated from Cladosporium sp. IFB3lp-2 EtOAc extract had no noticeable cytotoxicity versus HCT-116, Coxsachievirus A16, A549, MD-MBA-231, HepG2, human enterovirus 71, A375, and SW1116 cell lines (Conc. 20 \u00b5M) in the MTT assay [131 did not show any anti-larval activity towards both B. neritina and B. amphitrite larvae [132 did not have cytotoxic activity towards HCT-116, BEL-7402, HL-60, A549, L-02, HeLa, K562, MGC-803, MCF-7, PC-3, SH-SY5Y, and MDA-MB-231 (IC50 > 50 \u03bcM) [E. ictarda, E. tarda, and Cytospora glecosporioides (MICs ranging from 16 to 32 \u03bcg/mL) [129) separated from a soft coral-associated Cladosporium sp. TZP-29, together with the formerly isolated derivatives 130 and 131 had no cytotoxic effect towards A-549, SMMC-7721, and HeLa cells in the SRB method. Moreover, 129\u2013131 with IC50 ranged from 7.1 to 13.1 \u00b5M remarkably reduced the accumulation of lipid elicited by oleic acid (OA) in the HepG2 liver cells, in comparison to lovastatin as determined by oil-red O staining and intracellular triglyceride (TG) and total cholesterol (TC) quantification (The seco-acids TT assay . Compoune larvae . Moreove> 50 \u03bcM) , while i2 \u03bcg/mL) . Cladospfication .133, 134, and 138 and new fatty acids 135\u2013137, 139, and 140 isolated from C. cladosporioides OUCMDZ-187 obtained from the mangrove plant Rhizophora stylosa collected in Shankou, Guangxi Province of China showed no cytotoxic effects (IC50 > 50 \u03bcM) towards K562, A549, and HeLa cells in the SRB method [S. aureus CGMCC-1.2465, E. coli CGMCC-1.2389, E. aerogenes CGMCC-1.0876, P. aeruginosa CGMCC-1.1785, B. subtilis CGMCC-1.3376, and C. albicans CGMCC-2.2086 in the agar dilution method [Further, they exhibited potent lipid-lowering potential in HepG2 hepatocytes, revealing a promising anti-hyperlipidemic capacity . The newB method . Addition method .Tetralones comprise a bicyclic aromatic hydrocarbon and a ketone and are regarded as benzo-fused cyclohexanone derivatives. They played a substantial role as a starting material for the synthesis of a range of synthetic heterocyclic compounds and pharmaceuticals due to their potential reactivity and suitability . Additio152), a new dimeric tetralone bridged via C-C linkage, was separated from Cladosporium sp. KcFL6 derived from the mangrove plant Kandelia candel, together with 142\u2013144 , in comparison to NS-398 and indomethacin [A. baumannii ATCC-19606, S. aureus ATCC-29213, E. faecalis ATCC-29212, A. hydrophila ATCC-7966, E. coli ATCC-25922, K. pneumonia ATCC-13883, Fusarium sp., F. oxysporum f. sp. cucumeris, F. oxysporum f. sp. niveum, A. niger, and R. solani in the disc diffusion assay [Cladosporone A , compared to trichostatin A in the trypan blue-cell viability assay [142 had a remarkable anti-proliferative potential towards SW480, HT-29, and CaCo-2, in particular towards HT-29. It was revealed that HT-29 cells exposure to 142 produced G1/S phase cell cycle arrest, assisted by a vigorous p21waf1/cip1 expression, a significant down-regulation of CDK4, CDK2, cyclin E, and cyclin D1, and repression of CDK4 and CDK2 kinase activity [waf1/cip1 expression and inducing degradation of \u03b2-catenin, as well as impairing TCF/\u03b2-catenin pathway as evident by reduced cyclin D1 and c-Myc transcription. Finally, it induced the expression of E-cadherin, therefore antagonizing invasion and metastasis [C. cladosporioides HDN14-342 isolated from marine sediments yielded tetralone derivatives 143, 145\u2013147, 154, and 155 that were evaluated for cytotoxic activities towards HCT-116, HeLa, and A549 cell lines by SRB method and towards HL-60 and K562 cell lines by MTT method, in comparison to doxorubicin (IC50 0.2\u20130.8 \u00b5M). Compounds 146 and 147 were active towards K562, HeLa, and HCT-116 cell lines (IC50 ranging from 3.9 to 23.0 \u00b5M), while other metabolites had no activity (IC50 > 50.0 \u00b5M) [143 possessed no anti-allergic effect (IC50 > 200 \u00b5M) on RBL-2H3 cells, in comparison to loratadine (IC50 35.01 \u00b5M) using fluorometric assay [143) and cladosporols F-J (146 and 148\u2013151) from the marine algal-derived C. cladosporioides EN-399 and evaluated their cytotoxic activities towards H446, A549, HeLa, L02, Huh7, LM3, SW1990, and MCF-7 using MTT assay. Note that 143, 148, and 149 displayed cytotoxic activities towards most of the tested cell lines with IC50 ranging from 1.0 to 20.0 \u03bcM. Notably, 149 had cytotoxic effect towards LM3, A549, and Huh7 cell lines , compared to cisplatin (IC50 1.3 \u03bcM for A549 and 9.1 \u03bcM for LM3) and fluorouracil (IC50 6.2 \u03bcM for Huh7), whereas 143 exhibited cytotoxic activity (IC50 4.0 \u03bcM) towards H446 cell line, compared to adriamycin (IC50 4.0 \u03bcM). These results revealed that the existence of dihydro-1,4-naphthoquinone nucleus was important for the activity  and C-4 methoxyl strengthened the activity (148 vs. 151) [Compounds activity . It was tastasis . C. clad50.0 \u00b5M) . In 2020vs. 151) .E. coli, A. hydrophila, S. aureus, E. tarda, P. aeruginosa, M. luteus, V. alginolyticus, V. parahemolyticus, V. harveyi, A. brassicae, F. oxysporum, G. graminis, C. gloeosporioides, and P. piricolav using micro-plate assay. Compounds 143, 146, and 148\u2013151 showed inhibitory potential towards M. luteus, E. coli, and V. harveyi (MICs 4\u2013128 \u03bcg/mL). None of them had activity (MIC > 128 \u03bcg/mL) towards other tested microbes [143 and 145 from Cladosporium sp. JS1-2 isolated from the mangrove Ceriops tagal collected in the South China Sea [145 prohibited the growth of Helicoverpa armigera Hubner newly hatched larvae (IC50 150 \u03bcg/mL), compared to azadirachtin (IC50 25 \u03bcg/mL) [S. aureus with MIC 6.25 and 1.56 \u03bcg/mL, respectively, compared with ciprofloxacin (MIC 0.39 \u03bcg/mL) [150 and 153 that exhibited quorum sensing inhibitory potential towards Chromobacterium violaceum CV026  in the well diffusion assay [156 had no observable cytotoxic activity towards SF-268, NCI-H460, MCF-7, and HepG-2 (conc. 100 \u03bcM) in the SRB assay [157, in addition to 156, 158, and 159 isolated Cladosporium sp. JJM22 associated with the mangrove plant C. tagal had no cytotoxic effect (IC50 > 10 \u03bcM) versus HeLa cell line in the MTT assay, compared to epirubicin [158 exhibited noticeable antibacterial potential towards S. aureus, B. cereus, E. coli, V. alginolyticus, V. parahemolyticus, and MR S. aureus (conc. 20 \u03bcM) [160) and previously reported -3,4,8-trihydroxy-1-tetralone (159) were isolated from a sediment-associated Cladosporium sp. HDN17-58  , compared to ciprofloxacin [Moreover, their antimicrobial potential was assessed versus microbes . Bai et hina Sea . Compoun5 \u03bcg/mL) . Further9 \u03bcg/mL) . Cladospon assay . NeverthRB assay . The newirubicin   . One newHDN17-58 . Note thfloxacin .Perylenequinones comprise a class of natural products characterized by an oxidized pentacyclic core. They are dark-colored pigments isolated from diverse sources such as mold species, plants, and aphids . They re161\u2013164), were isolated from Cladosporium sp. KFD33  production by LPS (lipopolysaccharide) in RAW264.7 cells [Cladosporium sp. JJM22 yielded new naphthalene-chromane derivatives, cladonaphchroms A (169) and B (170), and related metabolites 165 and 168 that were assessed for antibacterial effectiveness versus S. albus ATCC-8799, E. coli ATCC-25922, B. subtilis ATCC-6633, Micrococcus tetragenus ATCC-13623, and M. luteus ATCC-9341, employing microplate assay. Compound 169 possessed significant potential against S. albus (MIC 1.25 \u00b5g/mL), compared to ciprofloxacin (MIC 0.6 \u00b5g/mL). Moreover, 169 and 170 demonstrated broad-spectrum antifungal activities (MICs 25.0\u2013100.0 \u00b5g/mL) towards P. parasitica var. nicotianae, A. brassicicola, B. oryzae, C. capsici, C. paradoxa Moreau, and D. medusaea Nitschke, compared to pochloraz (MICs 12.5\u201350.0 \u00b5g/mL) [166 had no cytotoxic effect (IC50 > 10 \u03bcM) versus HeLa cell line in the MTT assay and no antibacterial activity towards S. aureus, B. cereus, E. coli, V. alginolyticus, V. parahemolyticus, and MR S. aureus (conc. 20 \u03bcM) in the microplate assay [.7 cells   . Wu et ate assay .Xanthones are secondary metabolites commonly reported from plants, fungi, and lichen . They arC. halotolerans GXIMD 02502 associated with the coral Porites lutea yielded compounds 171\u2013177 that were evaluated for their cytotoxicity versus 22RV1 and C4-2B (prostatic cancer cell lines), as well as RWPE-1 . Among them, 171\u2013173, 175, and 176 revealed notable cytotoxicity versus C4-2B and 22RV1 cells (inhibitions ranged from 55.8% to 82.1% at conc. 10 \u00b5M), whereas 176 was the potent one . On the other hand, they exhibited nearly no cytotoxic effect versus RWPE-1 cell (inhibition < 27% at conc. 10 \u00b5M) [. 10 \u00b5M)  , along with the new metabolite, malettinin E (181) from Cladosporium sp. strain KF501 isolated from the German Wadden Sea , whereas 179\u2013181 exhibited weak antibacterial effect towards Xanthomonas campestris (IC50 28.3\u201337.9 \u03bcM), compared to chloramphenicol (IC50 2.1 \u03bcM) [Silber et al. reported the isolation of malettinins A\u2013C (dden Sea . These m 2.1 \u03bcM) .Bisnaphthopyrones are dimers, belonging to naphthopyrones. They have C13 basic skeleton (C6-C4-C3) that consists of naphthalene and pyrone cores .182), and the formerly isolated viriditoxin (183) and viriditoxin derivatives (184 and 185) were separated the sediment associated C. cladosporioides (183 was firstly reported from Aspergillus viridinutans [183 was the most potent one (IC50 0.1 \u03bcM), however 182 and 184 had a cytotoxic effect . However, 185 was ineffective [S. aureus ATCC-29213, with 183 being the most effective (MIC 0.023 \u03bcM) [The new binaphthopyrone, cladosporinone (orioides . Note thdinutans . They weffective . Note th.023 \u03bcM) .188\u2013190, 193, 200, 201, and 203 from Cladosporium sp. OUCMDZ-302 isolated from mangrove plant Excoecaria agallocha. They possessed no cytotoxic effect towards BEL-7402, A549, HeLa, K562, HL-60, and H1975 cell lines in the MTT and SRB methods. Whilst 201 and 203 showed radical scavenging activity against DPPH . None of these metabolites exhibited antimicrobial activities against E. coli, E. aerogenes, P. aeruginosa, B. subtilis, and C. albicans [192), had no \u03b1-glycosidase inhibitory effect and no cytotoxic activity towards SGC-7901, K562, Hela, and BEL-7042 cell lines in the MTT assay [186 and 205 displayed no cytotoxic effect (IC50 > 10 \u03bcM) versus HeLa cell line in the MTT assay, as well as no antibacterial potential towards S. aureus, B. cereus, E. coli, V. alginolyticus, V. parahemolyticus, and MR S. aureus (conc. 20 \u03bcM) in the microplate assay [C. halotolerans GXIMD 02502 associated with the coral Porites lutea yielded a new benzopyranone derivative, coniochaetone K (196) with unusual C-8 carboxyl, along with 194, 195, 197, and 198 that were evaluated for their cytotoxicity versus 22RV1, C4-2B, and RWPE-1 cell lines . On the other hand, they exhibited nearly no cytotoxic effect versus RWPE-1 and C4-2B cells [206 prohibited the growth of H. armigera Hubner newly hatched larvae (IC50 100 \u03bcg/mL), compared to azadirachtin (IC50 25 \u03bcg/mL) [S. aureus (MIC 6.25 \u03bcg/mL), compared with ciprofloxacin (MIC 0.39 \u03bcg/mL) [207) did not have obvious anti-biofilm activity towards S. aureus, E. coli, and B. subtilis [50 49.9 \u00b5M), compared to toosendanin (LC50 21.2 \u00b5M) in the brine shrimp lethality assay [210 possessed no anti-allergic effect (IC50 > 200 \u00b5M) on RBL-2H3 cells, in comparison to loratadine (IC50 35.01 \u00b5M) using fluorometric assay [211\u2013213 were separated from C. herbarum isolated from the sponge Aplysina aerophoba  with mortality rates 85 and 75% and 80 and 65%, respectively, while 213 did not have any activity. Besides, 213 showed growth inhibitory activity towards Spodoptera littoralis larvae  [211\u2013213 did not show any noticeable antimicrobial activity in the agar plate diffusion assay [Among them, 2B cells . Bai et 5 \u03bcg/mL) . Further9 \u03bcg/mL) . Cladospsubtilis . On the ty assay . Furtheric assay . \u03b1-Pyronerophoba . Compoun216 from C. cladosporioides that possessed no anti-allergic effect (IC50 > 200 \u00b5M) on RBL-2H3 cells, in comparison to loratadine (IC50 35.01 \u00b5M) using fluorometeric assay [C. tagal associated-fungus Cladosporium sp. JJM22 produced new cyclohexene derivatives, cladoscyclitols A\u2013D (218\u2013221) (219 (IC50 2.95 \u03bcM) revealed potent \u03b1-glucosidase inhibitory activity, compared to acarbose (IC50 2.35 \u03bcM) in the colorimetric assay [S. aureus ATCC-6538, E. coli ATCC-25922, B. cereu ATCC-6633, V. alginolyticus ATCC-3787, V. Parahemolyticus ATCC-17802, or MRSA CMCC-B-63303 in the micro-plate assay [223) and B (224), representing new azaphilone epimers, together with bicyclic diol (225) were separated from sea sediment-associated C. perangustum FS62 fungus. They had no observable cytotoxic activity towards SF-268, NCI-H460, MCF-7, and HepG-2 (Conc. 100 \u03bcM) in the SRB assay [In 2020, He et al. purified ic assay . The man218\u2013221) . Compounic assay . On the te assay . Perangu233 and 235 showed DPPH radical scavenging activity , in comparison to ascorbic acid (IC50 3.29 \u00b5M). Further, none of these compounds had antimicrobial potential versus P. aeruginosa, E. aerogenes, B. subtilis, E. coli, and C. albicans [232, 238, and 249 were separated from EtOAc extract of Cladosporium sp. F14 isolated from seawater and investigated for their anti-larval activity (conc. 50 \u00b5g/mL) towards B. neritina and B. amphitrite larvae in the settlement inhibition assays [232 had weak larvae settlement inhibition towards B. neritina and B. Amphitrite, respectively, whereas 238 and 249 showed weak inhibitory effects towards B. amphitrite and B. neritina larvae, respectively. In another larval settlement bioassay, 232, 238, and 249 inhibited B. neritina larval settlement  and B. amphitrite larval settlement . The larval settlement EC50 values of 249 towards B. amphitrite and 232 towards B. neritina were less than the US Navy program established standard requirement (EC50 25.0 \u00b5g/mL), revealing the potential of 232 and 249 as antifouling agents [232 obviously prohibited L. hongkongensis growth (IZD 8 mm and MIC 80 \u00b5g/mL), compared to streptomycin (MIC 250 \u00b5g/mL) [239 isolated Cladosporium sp. JJM22 associated with the mangrove plant C. tagal had no cytotoxic effect (IC50 > 10 \u03bcM) versus HeLa cell line in the MTT assay, compared to epirubicin [In the DPPH assay, albicans . The metn assays   . The ribirubicin .S. aureus, B. cereus, E. coli, V. alginolyticus, V. parahemolyticus, and MR S. aureus (conc. 20 \u03bcM) in the microplate assay [240 (IC50 2.05 \u03bcM) revealed potent \u03b1-glucosidase inhibitory activity, compared to acarbose (IC50 2.35 \u03bcM) in the colorimetric assay [S. aureus ATCC-6538, E. coli ATCC-25922, B. cereus ATCC-6633, V. alginolyticus ATCC-3787, V. Parahemolyticus ATCC-17802, and MRSA CMCC-B-63303 in the microplate assay [Cladosporium sp. associated with Chondria crassicualis red alga resulted in the separation of a phenol derivative, clavatol (241) that exhibited antioxidant capacity (ED50 227.0 \u00b5M) more than oxybenzone  as evident by their UV-A protecting potential [S. aureus [242 and 243 exhibited no observable cytotoxic activity towards SF-268, NCI-H460, MCF-7, and HepG-2 (Conc. 100 \u03bcM) in the SRB assay [247) did not have obvious anti-biofilm activity towards S. aureus, E. coli, and B. subtilis [50 16.4 \u00b5M), compared with ascorbic acid (IC50 4.9 \u00b5M). Besides, it showed moderate toxicity towards brine shrine naupalii (LC50 81.4 \u00b5M), comparing with toosendanin (LC50 21.2 \u00b5M) in the brine shrimp lethality assay [Additionally, it exhibited no noticeable antibacterial potential towards te assay . The newic assay . On the te assay . Phytochsubtilis , while ity assay .248 separated from Cladosporium sp. TPU1507 derived from marine sponge and assessed for inhibitory effect towards PTP1B and TCPTP using enzyme-based assay [50 27 \u03bcM) that was 2-fold weaker than on PTP1B (IC50 11 \u03bcM) [250), separated from C. herbarum isolated from Callyspongia aerizusa had no activity towards A. salina and HL-60 human leukemia cell line [251 from Cladosporium sp. OUCMDZ-1635 possessed no cytotoxic effect towards MCF-7, HeLa, HCT-116, HeLa, HCT-116, K562, and HL-60. Furthermore, it did not show antibacterial activity (conc. 100 \u03bcg/mL) against B. subtilis, P. aeruginosa, C. perfringens, S. aureus, E. coli, and C. albicans [252) prohibited the growth of H. armigera Hubner newly hatched larvae (IC50 150 \u03bcg/mL), compared to azadirachtin (IC50 25 \u03bcg/mL) [S. aureus (MIC 25.0 \u03bcg/mL), compared with ciprofloxacin (MIC 0.39 \u03bcg/mL) [253) and acetyl Sumiki\u2019s acid (254) exerted activity towards S. aureus and B. subtilis (IZDs 7 mm at conc. 5 \u00b5g/disk), whereas they had no activity towards C. albicans and E. coli [Compound ed assay . It show0 11 \u03bcM) . The newell line . In addi5 \u03bcg/mL) . Further9 \u03bcg/mL) . The fur E. coli .268) and six sterol derivatives: 256, 258, 260, 262, 263, and 267 from gorgonian-associated Cladosporium sp. WZ-2008-0042 [268 was reported in the same year by Pang et al. as new metabolites with the name cladosporisteroid B from Cladosporium sp. SCSIO41007 associated with Callyspongia sp. [50 values ranging from 0.11 to 0.17 \u00b5M) revealed antiviral activity against RSV  with therapeutic ratio (TC50/IC50) values ranging from 5.18 to 9.92, in comparison to ribavirin in the neuraminidase inhibition assay. This could be due to their binding to RSV GREs (glucocorticoid response elements) [268 had no noticeable antibacterial potential towards B. cereus, M. luteus, S. aureus, V. anguillarum E. coli, Shigella dysenteriae, B. subtilis, and V. Parahemolyticus, while 263 was moderately active (MIC 3.13 \u03bcM) towards S. dysenteriae [256, 261, 265, 266, and 268 separated from C. cladosporioides sea sediment-derived fungus possessed no anti-allergic effect on RBL-2H3 cells, in comparison to loratadine using fluorometeric assay [264) and new pregnanes, cladosporisteroid B (268) and cladosporisteroid C (269), along with 259, 265, and 270 from Cladosporium sp. SCSIO41007 isolated from Callyspongia sp. and assessed their antiviral activity towards EV71 and H3N2 using CCK-8 and CPE assays, respectively. Only, 268 (IC50 16.2 \u03bcM) had weak activity towards H3N2 compared to oseltamivir (IC50 34.0 nM). Moreover, they revealed no cytotoxic effect towards K562, MCF-7, and SGC-7901 in the CCK-8 assay [268 was purified from C. sphaerospermum EtOAc fraction by HPLC with the aid of LCMS and assessed for its influence on adipogenesis and lipid metabolism during maturation of adipocyte  using 3T3-L1 preadipocytes [Adipsin (adipocyte marker gene) expression. Further, it significantly upregulated ATGL  and reduced FASN and SREBP1  expression. Collectively, 268 facilitated lipid metabolism and disrupted adipogenesis via promoting lipolysis and prohibiting lipogenesis [A study conducted by Yu et al. in 2018 led to the separation of a new pregnane; 3\u03b1-hydroxy-7-ene-6,20-dione  from the culture of Cladosporium sp. isolated from intertidal marine sediment [E. coli ATCC-25922, B. subtilis ATCC-6633, and C. albicans ATCC-18804 in the agar diffusion method. It is noteworthy that this class of metabolites had been reported formerly from red algae  [S,3S,4E)-hepta-4,6-diene-2,3-diol (285) and -Undeca-3,8,10-trien-1,6-diol (286) were assessed for cytotoxic potential versus HeLa, BEL-7402, HL-60, A549, K562, and H1975 cell lines. Compound 286 had a cytotoxic effect versus H1975 cell line (IC50 10.0 \u00b5M), compared to ADR (IC50 0.38 \u00b5M). While both metabolites revealed no antioxidant and antimicrobial capacities [Gallo et al. reported for the first time from fungi the isolation of \u03b1,\u03b2-unsaturated aldehydes  ,156. Thepacities .Cladosporium sp. isolate N5 associated with Porphyra yezoensis red alga did not produce any pathogenic symptoms in the reinfection assay. Further, its EtOAc extract displayed no lethality to A. salina and had a moderate antimicrobial activity which indicated that Cladosporium sp. had no toxicity to the aquatic ecosystem and could be applied as a biocontrol agent [Cladosporium sp. EIODSF 008 EtOAc extract exhibited significant antibacterial potential towards E. coli, M. luteus, and B. subtilis (conc. 100 \u00b5g/disc) [Cladosporium sp. EN-S01 isolated from Sargassum cinereum brown algae showed anticancer activity towards MCF-7, HeLa, and DU-145 cell lines . The extract had greater cytotoxic activity and anti-proliferative towards MCF-7 and HeLa cell lines than towards DU-145 [C. cladosporioides KT384175 isolated from the seaweed Sargassum wightii possessed remarkable antioxidant potential that was comparable to ascorbic acid, as well as significant Fe3+ reducing power that could be referred to its phenolic contents. Moreover, it revealed anti-angiogenic potential as evidenced by the decrease in the number and length of blood vessel branches on CAM  in-vivo in the CAM assay. Further, C. cladosporioides extract (conc. 1.0 mg/mL) had lower wound healing potential than thalidomide (conc. 1.0 \u00b5g/mL) in the in vitro scratch assay using MCF-7 cells [Cladosporium sp. F14 can produce antifouling and antibiotic metabolites in the existence of xylose or glucose. Significantly, it showed higher antibiotic activity towards M. luteus, P. piscida, Rhodovulum sp., Ruegeria sp., V. fluvialis, and V. harveyi in the existence of a sugar carbon source than in its absence in the disc diffusion assay, even though the fungal cells were well-grown under both conditions. Moreover, it possessed antifouling potential as it reduced the attachment of B. neritina (bryozoan larvae) in the larval settlement assay [C. cladosporioides isolated from the seaweed S. wightii possessed noticeable antimicrobial potential towards E. coli MTCC-118, B. subtilis MTCC-441, S. aureus MTCC-7443, P. aeruginosa MTCC-424, and A. niger MTCC-281 with the highest growth inhibition towards S. aureus (IZD 12 mm) and least activity against B. subtilis (IZD 9.5 mm), compared to ampicillin  in the well diffusion method. Furthermore, they also had significant antioxidant potential comparable to ascorbic acid in the DPPH assay and moderate effectiveness in reducing power assay [C. halotolerans biomass isolated from the marine debris collected around Tarout Island showed a significant free radical scavenging effect (%inhibition 78% within 30 min incubation) in the DPPH assay. Moreover, it exhibited cytotoxic potential towards MCF-7 (IC50 34.27 \u00b5L/mL), compared to cisplatin (IC50 17.69 \u00b5L/mL) in the MTT assay, as well as an antifungal effect against A. niger  in the broth dilution method [Ding et al. stated that ol agent . In the \u00b5g/disc) . The EtOs DU-145 . Moreove-7 cells . The seant assay . The goler assay . Ameen en method .C. phlei  and C. cucumerinum  were isolated mainly from plant sources [Cladosporium species [seco-acids, macrolides, diketopiperazines, alkaloids, and tetramic acid derivatives were reported mainly from marine-associated Cladosporium species.From the comprehensive review of the available literature, it was noticed that  sources ,164,165. sources ,165. Add species ,169,170.Cladosporium species are of biotechnological and industrial relevance and could be considered as substantial enzyme producers. Their enzymes are active in harsh conditions such as extremely low temperatures and high salinity. Therefore, they can be utilized in various industrial and biotechnological applications. Besides, these species were found to be a wealthy pool covering a wide array of metabolites with various bioactivities. Over the past 22 years, 286 metabolites have been separated from marine-associated Cladosporium species isolated from various marine samples, including mangrove, sediment, sponges, corals, gorgonians, algae, bivalves, hydroids, and others  and C. cladosporioides   .The results revealed that alkaloids, macrolides, tetramic acid and pyrone derivatives, and phenolics are the major metabolites reported from this marine-associated fungal species . They coAlthough the structural diversity of these metabolites, they were insufficiently evaluated for their bioactivities. Most of them had been assessed for their antimicrobial, cytotoxicity, antiviral, and insecticidal activities .However, there are limited studies that focus on the mechanism of action of these metabolites. Many of the tested metabolites possessed no noticeable efficacy in some of the tested activities. Therefore, estimation of other potential bioactivities and derivatization of these metabolites, as well as the mechanistic and in vivo studies of the active metabolites should clearly be the target of future research.Cladosporium species such as co-cultivation of organisms and elicitors epigenetic, as well as, modifiers can be applied [Growing evidence has revealed that the activation of silent gene clusters has the potential to significantly enhance the discovery of new natural metabolites of high-therapeutic leads. Different strategies to awake the silent biosynthetic gene clusters of  applied ,173,174. applied ,175. The applied ,172,173. applied . Challen applied ,172,177. applied ,172,173. applied ,172,178. applied ."}
+{"text": "Cytopenias are rare complications of prolonged beta-lactam use; however, incidence and associated risk factors are not well described. Patients aged 18-64 years in the 2010-2016 IBM MarketScan Commercial Database discharged from the hospital on cephalosporin, penicillin, or carbapenem outpatient Parenteral Antimicrobial Therapy (OPAT) were included. The primary endpoint was hospital admission coded for neutropenia, leukopenia, or thrombocytopenia within the first 6 weeks post index discharge and within 7 days of beta-lactam discontinuation. Patients with history of malignancy and those who are on chemotherapy were excluded. Significant factors in univariate analysis were incorporated into a multivariable logistic regression model with sequential exclusion of variables with p > 0.1.A total of 35,102 patients received beta-lactam OPAT; median age was 52 years and 53.6% were male. The primary outcome occurred in 150 (0.43%) patients at a median of 19 days (IQR 10-28 days after index discharge), which included 63 (0.18%) neutropenia, 85 (0.24%) thrombocytopenia, and 23 (0.07%) leukopenia admissions. Factors independently associated with readmission cytopenias included chronic liver disease (OR 4.61 [CI 2.93-7.25]), valvular heart disease (2.69 [1.71-4.24]), receipt of vancomycin (2.10 [1.42-3.12]), or antifungal therapy (4.42 [2.01-9.68]); lower risk was associated with carbapenem therapy (0.49 [0.32-0.75]) and diabetes (0.48 [0.31-0.74]) (Table 1).Readmissions with cytopenias during beta-lactam OPAT were rare and carbapenem use was associated with lower risk compared to other classes of beta-lactams. Combination of beta-lactam with vancomycin was associated with an increased risk of cytopenias, and those patients might benefit from closer monitoring.Table 1. Factors Associated with Cytopenias during Beta-Lactams Outpatient Parenteral Antimicrobial Therapy (OPAT)Margaret A. Olsen, PhD, MPH, Pfizer"}
+{"text": "Post COVID Syndrome (PCS) is significant morbidity following COVID-19. This study aims to identify biomarkers that predict PCS in a Gulf Coast cohort known for poor health outcomes.Since March 2020 the study Collection of Serum and Secretions for SARS CoV-2 Countermeasure Development (aka ClinSeqSer) has been enrolling subjects with confirmed acute COVID-19, with initial visit at 1 month and follow up every three months from symptom onset. At follow-up, subjects complete symptom questionnaire, physical examination, nasopharyngeal swab/saliva collection, blood draw. Subjects with >= one symptom new since COVID are PCS, remainder are Non-PCS experienced at initial one month visit and six months or longer. Univariate and bivariate analysis was carried out to study significant associations of currently available dataset (N=60).Figure 1. Post-COVID SymptomsIncluded if \u201cnew since covid\u201d. For 60 subjects consented post-covid with completed questionnaire, results were analyzed. Most common symptoms reported were fatigue/tiredness or exhaustion (52%), muscle aches (38%), difficulty concentrating (33%) and headache (32%) as the most common symptoms during one month prior to their initial follow-up visit. The persistent symptoms experienced for six months or longer were fatigue/tiredness or exhaustion (25%), forgetfulness (22%), muscle aches (18%), and sleep difficulties (18%).Cohort is 36 (60%) female, 24 (40%) male, age group of 49 (82%) 18-64 years, 11 (18%) 65+ years, 33 (55%) African American, 27 (45%) Caucasian. Median follow-up time after symptom onset: 290 days. Study cohort reported fatigue (52%), myalgias (38%), difficulty concentrating (33%), headache (32%) as most common symptoms during first month from initial symptom onset. Persistent symptoms ( >=6 months) are fatigue (25%), forgetfulness (22%), myalgias (18%), sleep difficulties (18%). Bivariate analysis shows that gender , past stroke/transient ischemic attack (P=0.04), deep venous thrombosis (P=0.02), abnormal kidney function (P=0.01) associate with PCS. Convalescent antibodies  were measured and percentage inhibition of ACE2 spike interaction was recorded. Plasma inflammatory protein levels were measured using multiplex ELISA and Proximity Extension Assay technology during follow-up visit. Increased antibody ReSARS N IgG  response and higher convalescent IL-10 (P=0.04) was associated with PCS. Percent inhibition of ACE2: spike interaction was not associated (P=0.79) with PCS. Nasal swab/saliva SARS-COV-2 sequencing has not identified a specific SARS-CoV-2 virus mutation predictive of PCS. Table 1. Demographic and Clinical CharacteristicsThe bivariate analysis results showed that the gender , history of stroke or transient ischemic attack (P=0.0382), chest pain from narrow heart vessels (P=0.0479), deep venous thrombosis (P=0.0241) and abnormal kidney function (P=0.0142) were associated with Post-COVID syndrome.Table 2. Antibodies and ACE2 spike inhibition.The convalescent antibodies, ReSARS N IgG and S-RBD IgG were measured in U/mL and percentage inhibition of ACE2 spike interaction was recorded during follow-up visit for PCS vs Non-PCS subjects. The increased antibody ReSARS N IgG  response was associated with Post-COVID syndrome. Percent inhibition of ACE2: spike interaction was not associated (P=0.7932) with PCS.Table 3. Plasma inflammatory protein levels.Plasma inflammatory protein levels were measured using multiplex ELISA (MSD) and Proximity Extension Assay technology (Olink) recorded during follow-up visit for PCS vs Non-PCS subjects, revealing IL-10 (P=0.0379) was associated with development of PCS.This study identifies initial clinical and biomarker predictors of PCS in a cohort that is 55% African American.Figure 2. Antibody ReSARS N IgGReSARS N IgG measured in post-covid patients is significantly associated with post-COVID syndrome(P=0.0159). X axis: number of months from symptom onset to blood draw. Y axis: N IgG U/mL.Figure 3. Spike amino acid mutationsSpike amino acid mutations detected in SARS-CoV-2 from acute-phase respiratory isolates. Nasal swab/saliva samples were collected from subjects with acute COVID-19 at time of enrollment into ClinSeqSer, stored at -80\u00b0C followed by RNA isolation and SARS-CoV-2 qRT-PCR. Samples with Ct value of \u226430 were then sequenced using NextSeq (Illumina). All sequences are deposited on GISAID and under BioProject (ID PRJNA681020). X axis: subject ID, with ID number increasing chronologically. Y axis: amino acid position of each mutation moving from N- to C-terminus.Robert Garry, PhD, Zalgen Labs (Shareholder)"}
+{"text": "Ceftolozane/tazobactam (C/T), a cephalosporin\u2013\u03b2-lactamase inhibitor combination, is approved for treatment of complicated urinary tract infections, complicated intra-abdominal infections (cIAI), and nosocomial pneumonia in adults. Safety and efficacy of C/T in pediatric participants with cIAI was assessed.This phase 2 study (NCT03217136) compared C/T + metronidazole (MTZ) with meropenem (MEM) for treatment of cIAI. Age- and weight-adjusted dosing is summarized in Table 1. The primary objective was to evaluate the safety and tolerability of C/T + MTZ compared with MEM. A key secondary endpoint was clinical cure at end of treatment (EOT) and test of cure (TOC).Table 1. Summary of Dosing and Pharmacokinetic Sampling Schedule by Age CohortA total of 94 participants were randomized 3:1; 91 were treated with C/T + MTZ (n=70) or MEM (n=21) comprising the modified intent-to-treat (MITT) population. The clinically evaluable population included 78 participants at EOT  and 77 participants at TOC . The most common diagnosis and pathogen in the MITT population were complicated appendicitis  and Escherichia coli . The mean (SD) intravenous therapy/overall treatment duration was 6.4 (2.8)/9.3 (3.6) days and 5.8 (1.8)/9.0 (3.2) days for C/T + MTZ and MEM, respectively. In total, \u22651 adverse events (AE) occurred in 80.0% and 61.9% of participants receiving C/T + MTZ and MEM, respectively (Table 2), of which 18.6% and 14.3% were considered drug related. Serious AE occurred in 11.4% (8/70) and 0% (0/21) of participants receiving C/T + MTZ and MEM, respectively; none were considered drug related. No drug-related study drug discontinuations occurred. In the MITT population, rates of clinical cure for C/T + MTZ and MEM at EOT were 80.0% and 95.2%, and at TOC were 80.0% and 100%, respectively ; 6 of the 14 failures for C/T + MTZ were indeterminate responses scored as endpoint failures per protocol. In the clinically evaluable (CE) population, rates of clinical cure for C/T + MTZ and MEM were 89.8% and 100% at EOT, and 89.7% and 100% at TOC, respectively .C/T + MTZ was well tolerated in pediatric participants with cIAI, and rates of clinical success were high with C/T treatment. C/T is a promising new treatment option for children with cIAI.Carl-Christian A. Jackson, MD, Merck & Co. Inc. (Shareholder) Julia Lonchar, MSc, Merck Sharp & Dohme Corp.  Feng-Hsiu Su, MPH, MBA, Merck Sharp & Dohme Corp.  Jennifer A. Huntington, PharmD, Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA (Employee) Mekki Bensaci, PhD, Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA (Employee) Myra W. Popejoy, PharmD, Merck Sharp & Dohme Corp. (Employee) Matthew G. Johnson, MD, Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA (Employee) Carisa S. De Anda, PharmD, Merck Sharp & Dohme Corp.  Elizabeth G. Rhee, MD, Merck Sharp & Dohme Corp  Christopher Bruno, MD, Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA (Employee)"}
+{"text": "Sarcomas are highly heterogeneous in molecular, pathologic, and clinical features. However, a classification of sarcomas by integrating different types of pathways remains mostly unexplored.We performed hierarchical clustering analysis of sarcomas based on the enrichment scores of 14 pathways involved in immune, stromal, DNA damage repair (DDR), and oncogenic signatures in three bulk tumor transcriptome datasets.TP53 mutations, and the worst survival. We further validated the stability and reliability of our classification method by analyzing a single cell RNA-Seq (scRNA-seq) dataset. Based on the expression levels of five genes in the pathways of T cell receptor signaling, cell cycle, mismatch repair, focal adhesion, and calcium signaling, we built a linear risk scoring model (ICMScore) for sarcomas. We demonstrated that ICMScore was an adverse prognostic factor for sarcomas and many other cancers.Consistently in the three datasets, sarcomas were classified into three subtypes: Immune Class (Imm-C), Stromal Class (Str-C), and DDR Class (DDR-C). Imm-C had the strongest anti-tumor immune signatures and the lowest intratumor heterogeneity (ITH); Str-C showed the strongest stromal signatures, the highest genomic stability and global methylation levels, and the lowest proliferation potential; DDR-C had the highest DDR activity, expression of the cell cycle pathway, tumor purity, stemness scores, proliferation potential, and ITH, the most frequent Our classification method provides novel insights into tumor biology and clinical implications for sarcomas.The online version contains supplementary material available at 10.1186/s12967-022-03248-3. CDK4 and RB1-associated CNAs  prognosis among the sarcoma subtypes. Notably, DDR-C was likely to have the worst survival consistently in the three datasets  Fig.\u00a0B, while 05) Fig.\u00a0C.Fig. 3CU test, P\u2009<\u20090.001)   . These phenotypes are associated with tumor progression, immune evasion, drug resistance, and unfavorable prognosis , 29. Not01) Fig.\u00a0D. Stemne01) Fig.\u00a0E. In add05) Fig.\u00a0F. AltogeP\u2009=\u20090.047; hazard ratio (HR)\u2009=\u20091.72 and its 95% confidence interval (CI)   survival analysis by the multivariate Cox\u00a0proportional hazards model. We found that the subtype DDR-C remained a significant risk factor for DFS , while it showed no significant difference between Imm-C and DDR-C , while they were not significantly different between Imm-C and DDR-C  and/or increased CNAs . We founR-C Fig.\u00a0A. HomoloR-C Fig.\u00a0B. In addR-C Fig.\u00a0C. The G-R-C Fig.\u00a0D. OveralTP53 had a significantly higher mutation frequency in DDR-C than in Imm-C and Str-C \u2009=\u20092.0) . On the other hand, TP53 mutations were less frequent in Str-C than in DDR-C and Imm-C . This result could explain why Str-C was more genomically stable than the other subtypes because p53 plays an important role in the maintenance of genomic stability [DYNC2H1 and MDN1 were more frequently mutated in DDR-C than in Imm-C and Str-C      \u2009<\u20090.05]. In contrast, 124 and 180 genes had significantly higher methylation levels in Str-C and DDR-C, respectively, compared to other subtypes. Notably, most of these genes showed significant inverse correlations of their expression levels with methylation levels . These proteins included Syk, PREX1, Lck, 14-3-3_epsilon, PI3K-p85, Caspase-7_cleavedD198, PRDX1, Annexin-1, G6PD, Bax, ATM, p38, STAT5-alpha, Annexin_VII, Claudin-7, p90RSK, TIGAR, CD31, and GATA3  Fig.\u00a0. These pCD40LG, CDC25A, MSH2, FLT4, and ADCY2. The five genes were involved in five of the 14 pathways for clustering analysis, including T cell receptor signaling (CD40LG), cell cycle (CDC25A), mismatch repair (MSH2), focal adhesion (FLT4), and calcium signaling (ADCY2). ICMScore calculates risk score in a tumor as follows: ICMScore\u2009=\u20090.76\u2009\u00d7\u2009exp(CD40LG)\u2009+\u20090.73\u2009\u00d7\u2009exp(FLT4)\u00a0\u2212\u00a00.20\u2009\u00d7\u2009exp(ADCY2)\u2009+\u20091.97\u2009\u00d7\u2009exp(CDC25A)\u2009+\u20091.50\u2009\u00d7\u2009exp(MSH2), where exp(X) denotes the expression level of gene X in the tumor sample. To prove that ICMScore is an authentic risk factor in sarcomas, we compared ICMScores among the three sarcoma subtypes and analyzed their correlation with survival prognosis in sarcomas. In the three sarcoma datasets, As expected, ICMScores was significantly higher in DDR-C than in Imm-C and Str-C    to evaluate the prognostic risk of sarcomas based on the expression levels of five genes, including 05) Fig.\u00a0A. Surviv05) Fig.\u00a0B. These 05) Fig.\u00a0C.Fig. 8AUsing the pathway-based clustering method, we analyzed a scRNA-seq dataset GSE131309), which involved gene expression profiles in 6951 single cells from 12 sarcoma [advanced synovial sarcoma (SyS)] patients. The 6951 single cells included 4371 tumor cells, 90 B cells, 943 macrophages, 185 mastocytes, 102 NK cells, 235 CD4\u2009+\u2009T cells, 659 CD8\u2009+\u2009T cells, 206\u00a0T cells, 79 endothelial cells, and 81 cancer-associated fibroblasts (CAFs). Using the t-distributed stochastic neighbor embedding (tSNE) algorithm , we clus, which iWe performed hierarchical clustering of the 6951 single cells based on the enrichment scores of the 14 pathways identified three subtypes of these cells  exhibited the consistent expression pattern: Imm-C\u2009<\u2009Str-C\u2009<\u2009DDR-C    were significantly higher in Imm-C than in Str-C and DDR-C Fig.\u00a0D. The raR-C Fig.\u00a0D. It ind01) Fig.\u00a0E, suppor01) Fig.\u00a0F. It is TP53 mutations, and the worst survival. It is interesting to observe that there was no significantly different TMB between DDR-C and Imm-C, while their immune infiltration levels were significantly different. Two possible reasons could explain this observation: (1) DDR-C had more frequent arm-level copy number amplifications and deletions than Imm-C; and (2) DDR-C had higher ITH than Imm-C, since both CNAs [In this study, we proposed a novel classification method for sarcomas based on the enrichment scores of 14 pathways, which were involved in immune, stromal, DDR, and oncogenic signatures. In three datasets for bulk tumors and a scRNA-seq dataset, we reproducibly identified three sarcoma subtypes: Imm-C, Str-C, and DDR-C. Imm-C had the strongest anti-tumor immune signatures and the lowest ITH; Str-C showed the strongest stromal signatures, the highest genomic stability and global methylation levels, and the lowest proliferation potential; DDR-C had the highest DDR activity, expression of the cell cycle pathway, tumor purity, stemness scores, proliferation potential, and ITH, the most frequent oth CNAs  and ITH oth CNAs  are negaThe TCGA Research Network analyzed six types of adult soft tissue sarcomas: DDLPS, LMS, UPS, MFS, MPNST, and SS . We founBased on the enrichment scores of 14 pathways associated with immune, stromal, and DDR signatures, we classified sarcomas into three subtypes. The three sarcoma subtypes were characterized by different immune infiltration levels, stromal signatures, DDR activity, genome features, tumor progression phenotypes, and clinical outcomes. Our new classification method for sarcomas provides novel insights into tumor biology and clinical implications for this disease.Additional file 1: Table S1. A summary of the datasets analyzed.Additional file 2: Table S2. Pathways, immune signatures, and biological processes and their marker genes.Additional file 3: Fig. S1. Comparisons of the expression levels of human leukocyte antigen\u00a0(HLA) genes among the sarcoma subtypes. The one-way ANOVA test P-values are shown. * P\u2009<\u20090.05, ** P\u2009<\u20090.01, *** P\u2009<\u20090.001.Additional file 4: Table S3. Pearson correlations between the methylation levels and expression levels of the genes having significantly different methylation levels among the sarcoma subtypes.Additional file 5: Fig. S2. Heatmap showing top 30 genes with the most significant upregulation of methylation levels in each of the three subtypes.Additional file 6: Fig. S3. Positive correlations between the expression levels of 19 proteins significantly upregulated in Imm-C and immune signature scores in TCGA-SARC. * P\u2009<\u20090.05, ** P\u2009<\u20090.01, *** P\u2009<\u20090.001.Additional file 7: Fig. S4. Overlaps between our subtyping and other subtyping of sarcoma in TCGA-SARC. (A) Proportions of our subtypes in six types of adult soft tissue sarcomas: DDLPS, LMS (ULMS and STLMS), UPS, MFS, MPNST, and SS. (B) Proportions of our subtypes in the immune subtypes of sarcomas. (C) Proportions of the five subgroups (Clusters A-E) in our subtypes."}
+{"text": "Enterobacterales are considered an urgent threat for patients in healthcare facilities, causing infections with significant morbidity and mortality. Most isolates are multidrug resistant with limited treatment options, so combination therapy is an alternative. Recently, synergy with piperacillin/tazobactam (P/T) + meropenem (MP) was demonstrated against 7/10 (70%) KPC-producing Escherichia coli and 9/10 (90%) OXA-48-producing K. pneumoniae using time-kill assay . The aim of the present study was to further evaluate the combination of P/T + MP against KPC-producing Enterobacter cloacae, in addition to OXA-producing K. pneumoniae using our rapid ETEST MIC:MIC synergy method.Carbapenem-resistant K. pneumoniae  and 7 KPC-producing E. cloacae  were obtained from the CDC and FDA Antibiotic Resistance Isolate Bank. ETEST MICs for P/T and MP and our ETEST synergy method were performed in triplicate for each isolate. The summation fractional inhibitory concentration was calculated, and the mean value was interpreted as: < 0.5 synergy; > 0.5-1 additivity; > 1-4 indifference; and > 4 antagonism.14 carbapenemase-producing isolates: 7 OXA-48-like K. pneumoniae and synergy (6) or additivity (1) against all 7 KPC-producing E. cloacae. No antagonism was detected.MICs (\u00b5g/mL) ranged: MP, 0.5 to > 32 (14% susceptible) and P/T, 96/4 to > 256/4 . The combination of P/T + MP showed synergy (3) or additivity (2) against 5/7 (71%) OXA-producing K. pneumoniae and 7/7 KPC-producing E. cloacae, similar to previously published findings showing synergy in 7/10 KPC-producing E. coli and 9/10 OXA-48-producing K. pneumoniae using time-kill assay. Our ETEST synergy method is simple to use and should be evaluated more extensively. Regardless of the method used, results may or may not correlate in an in vivo setting. In vivo studies are needed.Using our ETEST MIC:MIC method, the combination of P/T + MP demonstrated synergy or additivity in 5/7 OXA-producing All Authors: No reported disclosures"}
+{"text": "Annexins as Ca2+/phospholipid-binding proteins are involved in the control of many biological processes essential for plant growth and development. In a previous study, we had shown, using a proteomic approach, that the synthesis of two annexins is induced in pea roots in response to rhizobial inoculation. In this study, phylogenetic analysis identif ied these annexins as PsAnn4 and PsAnn8 based on their homology with annexins from other legumes. The modeling approach allowed us to estimate the structural features of these annexins that might inf luence their functional activity. To verify the functions of these annexins, we performed comparative proteomic analysis, experiments with calcium inf lux inhibitors, and localization of labeled proteins. Essential down-regulation of PsAnn4 synthesis in a non-nodulating pea mutant P56 (sym10) suggests an involvement of this annexin in the rhizobial symbiosis. Quantitative RT-PCR analysis showed that PsAnn4 was upregulated at the early stages of symbiosis development, starting from 1\u20133 days after inoculation to up to 5 days after inoculation, while experiments with the Ca2+ channel blocker LaCl3 revealed its negative inf luence on this expression. To follow the PsAnn4 protein localization in plant cells, it was fused to the f luorophores such as red f luorescent protein (RFP) and yellow f luorescent protein (YFP) and expressed under the transcriptional regulation of the 35S promoter in Nicotiana benthamiana leaves by inf iltration with Agrobacterium tumefaciens. The localization of PsAnn4 in the cell wall or plasma membrane of plant cells may indicate its participation in membrane modif ication or ion transport. Our results suggest that PsAnn4 may play an important role during the early stages of pea-rhizobial symbiosis development. Annexins are of particular research interest due to their abilityto regulate various aspects of plant growth and development.Annexins belong to the evolutionarily conservedsuperfamily of proteins that are involved in Ca2+-dependentor Ca2+-independent binding with membrane phospholipids. Most annexinshave four putative annexin repeats of around 70 amino acids,with the conservative repeat GxGT-(38 residues)-D/E, whichconfers Ca2+/phospholipid-binding activity to these proteins. In addition,some plant annexins have motifs demonstrating F-actin bindingand peroxidase and ATPase/GTPase activities .Despite the general structural similarity of these proteins,the functions of annexins are diverse, and individual annexinsmay have specific activities. Annexins are involved in a widevariety of essential cellular processes, including the regulationof membrane organization, vesicle trafficking, cytoskeletaldynamics, exocytosis, cell cycle control, ion transport, andsignal transduction . Annexins as phospholipid-binding proteinsare being implicated in the fusion of membrane vesicles,as was shown for annexins from bell pepper and cotton . They are also involved inthe regulation of exocytosis, e. g., annexins in Zea mays rootcap cells . Moreover, annexins can functionas cationic channels activated by various stimuli in cells.Annexins can influence the Ca2+ influx in plant cells, as wasdemonstrated for a Capsicum annuum annexin, which hasCa2+-channel activity . The Arabidopsisthaliana annexin AtAnn1, which is expressed in root cells,exhibits pH-dependent cation-channel activity, while Z. maysannexins cause active conductivity of Ca2+ in lipid bilayersat slightly acidic pH . Since annexins can be Ca2+ sensors, these proteins arelikely to be involved in signal transduction; for example, theannexin from Triticum aestivum was suggested to be engagedin low-temperature signaling .Participation of annexins in the responses to cold, oxidative,and saline stresses is well-studied in plants . The annexinAtAnn1 from A. thaliana is involved in plant protectionagainst oxidative stress .The overexpression of AtAnn has been found to confer toleranceto drought and salt stresses and fungal attack in transgenicplants . Similarly,the overexpression of the wild tomato (Solanum pennellii)annexin SpAnn2 in cultivated tomato Solanum lycopersicumenhances drought and salt tolerance through the eliminationof reactive oxygen species (ROS) .Some annexins are also known to be activated in plantsduring interaction with plant-growth promoting bacteria and the development of mutualistic symbioses. During legume-rhizobial symbiosis,physiological changes occur, which are necessary for rhizobialinfection and nodule organogenesis, such as the stimulationof ion fluxes, membrane depolarization, ROS production,cytoplasm alkalinization, perinuclear calcium oscillations,and cytoskeletal rearrangements. In Medicago truncatula, thetranscription of MtAnn1 is activated directly by Nod factorsor inoculation with rhizobia in epidermal cells and later incortical cells . Studies using confocal microscopy showedGFP-labeled MtAnn1 to be localized in the cytoplasm, butprotein accumulation in response to inoculation occurred atthe periphery of the nucleus. MtAnn1 has been shown to beable to bind to the membrane phospholipid phosphatidylserine.Therefore, MtAnn1 is probably related to the eventsoccurring at the early stages of symbiosis, leading to bacterialinfection or nodule organogenesis .Transcriptome profiling of roots inoculated with rhizobiarevealed enhanced expression of MtAnn2, as well as MtAnn1. The expression of the MtAnn2 gene isassociated with cell division in the nodule primordium . Proteomic analysis revealed the MtAnn2protein presence in lipid rafts from root plasma membranepreparations . Another annexin MtAnn3was found to be important for root hair deformations inM. truncatula . The increased expression ofMtAnn1 and MtAnn2 is also associated with the early stagesof AM fungal symbiosis, which corresponds to the stages ofpre-infection and infection in this type of symbiosis . This may indicate the general role of these annexinsin the regulation of signaling pathways that lead to thedevelopment of two types of symbiosis.A protein homologous to MtAnn1 \u2013 PvAnn1 from Phaseolusvulgaris \u2013 is activated at the early stages of symbiosisdevelopment . The stimulation of Ca2+ ion transfer throughthe plasma membrane and ROS production caused by Nodfactors constitute an early response in the signal transductionpathway. Analysis of PvAnn1-RNAi transgenic roots inoculatedwith rhizobia showed a decrease in ROS productionand Ca2+ influx into the cells, which resulted in impairedprogression and decreased numbers of infection threads andnodules . Taken together, thesefindings point to the involvement of PvAnn1 in the regulationof signal transduction at early stages.Previously performed proteomic analysis in pea (Pisumsativum L.) allowed us to reveal two annexins, the synthesisof which was increased in response to inoculation with Rhizobium leguminosarum bv. viciae RCAM1026 in 24 h. In this work, searching in therecently released pea genome database using available codingsequences for annexin genes from M. truncatula andP. vulgaris revealed 15 annexins in pea. Phylogenetic analysisshowed the relationship among members of the annexin superfamilyin other legumes and allowed the identification oftwo previously revealed pea annexins responsive to rhizobialinoculation as PsAnn4 and PsAnn8 based on their homologywith the M. truncatula and P. vulgaris proteins. To verify thefunction of these annexins, we performed comparative proteomicanalysis using pea mutant P56 (sym10) unable to formsymbiosis and wild type cv. Frisson. The approaches employedincluded quantitative RT-PCR, experiments with calciumchannel inhibitors, and localization of labeled proteins.Plant material and bacterial strain. Pea Pisum sativum L.seeds cv. Frisson were sterilized with sulphuric acid for5 min, washed with water 3 times, transferred on 1 % wateragar plates and germinated at room temperature in the dark.4\u20135 days-old seedlings were transferred into pots with vermiculitesaturated with Jensen medium , grown in a growth chamber at 21 \u00b0\u0421 at 16 h light/8 h dark cycles, 60 % humidity. For experiments with inhibitor,the Ca2+ channels blocker LaCl3, the plants were grownin pots saturated with Jensen medium with 100 \u03bcM CaCl2 \u00d72 H2O. The Rhizobium leguminosarum bv. viciae strainRCAM 1026 (WDCM 966) was cultivated at 28 \u00b0\u0421 on TY agar medium with 0.5 mg/ml of streptomycin.Fresh liquid bacterial culture was grown in B\u2013 medium and the optical density of the suspensionat 600 nm (OD600) was adjusted to 0.5. Pea seedlingswere inoculated with 2 ml of R. leguminosarum bv. viciaeper plant. Pea roots  were harvested 1 dayafter inoculation (dai).Nicotiana benthamiana seeds were surface sterilized with10 % hypochlorite for 10 min, washed with water 5 timesand left for imbibition on a plate with sterile filter paper at4 \u00b0\u0421. All seeds were germinated in a large plastic box withsoil for seven days, and then transferred into individual potswith soil. Plants were grown at 23 \u00b0\u0421 with 16 h light/8 h darkcycles, 60 % humidity.http://www.clustal.org/omega/. The phylogenetic tree was generatedwith the Maximum Likelihood method using MEGA X https://www.megasoftware.net/ with 1000 bootstrap replicates. Thedomain composition of the corresponding encoded proteinswas assessed using PFAM https://www.sanger.ac.uk/science/tools/pfam.Phylogenetic analysis. Multiple sequence alignments wereperformed using Clustal\u03a9 https://salilab.org/modeller/9.20/release.html. Visualization of the three-dimensional structurewas obtained using the PyMol programhttps://pymol.org/2/. The three-dimensional crystal structure of theGhAnn1 G. hirsutum protein  was used asa template for building the model. To refine the model, the (VTFM) and the method of molecular dynamics in vacuum.The reliability of the model was calculated by the formula Protein homology modeling was performed in Modeller9.20 Isolation of total protein from pea roots. A modified methodwas used to isolate proteins from pea roots . 100 mg of the roots were ground in liquid nitrogen,then extraction buffer (0.1 M tris-HCl (pH 8.0), 30 % sucrose,10 mM dithiothreitol (DTT), 2 % sodium dodecyl sulfate(SDS), a mixture of protease inhibitors was added to the material and extraction was performed at+4 \u00b0\u0421. After centrifugation at 12 000 g for 15 min, the supernatantwas mixed in a 1:1 ratio with phenol (pH 8.0) , centrifuged at 12 000 g for 5 min.The upper phase was taken for precipitation of proteins. Fivevolumes of cold 100 mM ammonium acetate in methanol wereadded and incubated for 30 min at \u201320 \u00b0\u0421. After centrifugationat 12 000 g for 5 min, the pellet was washed twice with100 mM ammonium acetate in methanol and twice with 80 %acetone. The precipitate was dried in air and dissolved in thebuffer for isoelectric focusing (25 mM tris-HCl (pH 8.0), 9 Murea, 4 % CHAPS, 50 mm DTT, 0.2 % ampholytes ). Protein concentration was measuredusing Bradford assay .Two-dimensional differential gel electrophoresis. Twodimensionaldifferential gel electrophoresis (DIGE) of proteinswas performed using staining of samples with variousfluorescent dyes . The samples were conjugatedfor 30 min on ice with fluorescent dyes Cyanine 2or Cyanine 5 (Cy2 or Cy5) in various combinations. The incubationsolution contained 400 pM of each dye dissolvedin dimethylformamide for 30 min on ice. The reaction wasstopped by adding 10 mM L-lysine (Sigma-Aldrich), followedby incubation on ice for 10 min. After that, the controland experimental samples were mixed, DTT and ampholyteswere added. Passive in-gel rehydration with immobilizedpH gradient (Bio-Rad Laboratories) was performed overnightat room temperature. The total amount of sample applied to7 cm gel  was up to 100 \u03bcg.Isoelectric focusing (IEF) was performed in a Protean IEFsystem (Bio-Rad Laboratories) at a temperature of 20 \u00b0\u0421, thesamples were desalted at 250 V for 15 min, after which thevoltage was linearly increased to 4,000 V for 2 hours, thenIEF was carried out with increasing voltage up to 10 000 V.Before electrophoresis in polyacrylamide gel (PAGE), proteinrecovery was carried out in buffer with DTT  for10 min followed by alkylation in iodoacetamide buffer  for 15 min. The second direction of two-dimensionalelectrophoresis was carried out in tris-glycine bufferin 15 % polyacrylamide gel using a 4 % stacking gel. Afterseparation of proteins the gels were visualized using a laserscanner Typhoon FLA 9500 .Mass spectrometry. The proteins were rehydrated in trypsinsolution  on ice for1 h and then incubated for 1 h at 56 \u00b0\u0421. The peptides wereextracted from the gel with 50 % acetonitrile, 0.1 % formicacid. This solution was evaporated in vacuum concentratorCentriVap (Labconco) at 4 \u00b0\u0421 and dissolved in phase A . Mass spectrometry was performedusing Agilent ESI-Q-TOF 6538 UHD (Agilent Technologies)combined with high performance liquid chromatographAgilent 1260 (Agilent Technologies). Chromatographywas performed in system water \u2013 acetonitrile in the presenceof 0.1 % formic acid  in the gradient of acetonitrile (from 5 to 60 % phase Bfor 25 min and to 100 % phase B for 5 min) on Zorbax 300SBC18column 3.5 \u03bcm, 150 mm length (Agilent Technologies)with flow rate 15 \u03bcl/min.RNA extraction and quantitative reverse transcriptionPCR (RT-PCR). RNA extraction and RT-PCR were performedas described previously . Thequantitative RT-PCR analysis was performed on a CFX-96real-time PCR detection system with C1000 thermal cycler(Bio-Rad Laboratories). All primer pairs (Table 1) were designedusing the Vector NTI program and produced by theEvrogen company (www.evrogen.com). PCR amplificationspecificity was verified using a dissociation curve (55\u201395 \u00b0\u0421).mRNA levels were normalized against Ubiquitin and valueswere calculated as ratios relative to non-inoculated root expressionlevels. The data of two-three independent biologicalexperiments were analysed. Statistical analysis was conductedby Student\u2019s test ( p < 0.05) to assess the differences betweenvariants.Mass spectrometry. The proteins were rehydrated in trypsinsolution  on ice for1 h and then incubated for 1 h at 56 \u00b0\u0421. The peptides wereextracted from the gel with 50 % acetonitrile, 0.1 % formicacid. This solution was evaporated in vacuum concentratorCentriVap (Labconco) at 4 \u00b0\u0421 and dissolved in phase A . Mass spectrometry was performedusing Agilent ESI-Q-TOF 6538 UHD (Agilent Technologies)combined with high performance liquid chromatographAgilent 1260 (Agilent Technologies). Chromatographywas performed in system water \u2013 acetonitrile in the presenceof 0.1 % formic acid  in the gradient of acetonitrile (from 5 to 60 % phase Bfor 25 min and to 100 % phase B for 5 min) on Zorbax 300SBC18column 3.5 \u03bcm, 150 mm length (Agilent Technologies)with flow rate 15 \u03bcl/min.RNA extraction and quantitative reverse transcriptionPCR (RT-PCR). RNA extraction and RT-PCR were performedas described previously . Thequantitative RT-PCR analysis was performed on a CFX-96real-time PCR detection system with C1000 thermal cycler(Bio-Rad Laboratories). All primer pairs (Table 1) were designedusing the Vector NTI program and produced by theEvrogen company (www.evrogen.com). PCR amplificationspecificity was verified using a dissociation curve (55\u201395 \u00b0\u0421).mRNA levels were normalized against Ubiquitin and valueswere calculated as ratios relative to non-inoculated root expressionlevels. The data of two-three independent biologicalexperiments were analysed. Statistical analysis was conductedby Student\u2019s test ( p < 0.05) to assess the differences betweenvariants.Genetic constructs for plant transformation. To obtainthe pBIN19 vector for plant transformation, carrying the geneof interest, the coding sequence of PsAnn4 gene without stopcodonhas been amplified using cDNA as a template withcorresponding primers (see Table 1). Total RNA was isolatedfrom 2 dai pea roots of cv. Frisson. Amplification was doneusing Phusion Flash High-Fidelity PCR Master Mix (ThermoScientific). The amplified products were restricted with XbaIand EcoRI and subcloned in the pMON vector under 35Spromoter in the frame with the sequences encoding RFP orYFP and nopaline synthase terminator (Tnos). The inserts wereverified by sequencing. The cassette composed of the 35Spromoter, gene of interest fused with RFP or YFP and Tnoswas excised from pMON using Hind III, SmaI and cloned inthe pBIN19. All verified constructs were transferred into theAgrobacterium tumefaciens LBA4404.Transient protein expression in N. benthamiana leaves.A. tumefaciens strain LBA4404 was used for infiltration inN. benthamiana leaves. Bacterial culture was grown at 28 \u00b0Covernight, then centrifuged at 3000 g and resuspended in10 mM MES-KOH, 10 mM MgCl2 and 0.5 mM acetosyringoneup to culture density OD600 = 0.5. Bacterial cells wereinfiltrated into the leaves of 3-week-old N. benthamiana.Plants were analyzed 48\u201396 h after infiltration.ResultsPhylogenetic analysis of annexinsin pea and other legumesThe search of the sequences presumably coding for annexinsin legumes was performed using BlastX with 8 previouslyrevealed M. truncatula and 13 P. vulgaris nucleotidesequences encoding these proteins  as queries against differentplant sequence databases: https://phytozome.jgi.doe.gov/pz/portal.html for M. truncatula and P. vulgaris, http://www.kazusa.or.jp/lotus/ for L. japonicus, and the URGI databasev. 1 https://urgi.versailles.inra.fr/blast for P. sativum L.. As a result, we were able to identify 18 codingsequences (CDSs) for annexins in M. truncatula, 15 inP. sativum L., and 13 in L. japonicus (Table 2). Twenty-threegenes had been previously found to encode annexins in soybean. The coding sequences for annexinsfrom P. sativum were named based on their phylogeneticrelationships with the corresponding homologous sequencesfrom M. truncatula and P. vulgaris (see Table 2) .The phylogenetic analysis  was performed usingthe deduced amino acid sequences of annexins found andannotated for P. sativum along with those of other legumes and non-legumes , whichwere available in the Phytozome database v. 12.1 and otherdatabases.Based on our analysis, the previously found MtAnn1(Medtr8g038210) and PvAnn1 (Phvul.011g209300) clusteredin the subclade with proteins corresponding to P. sativumPsat4g147120 and Psat4g191080, named PsAnn1a andPsAnn1b (see Table 2). Revealed in M. truncatula MtAnn2(Medtr8g038220) and P. vulgaris PvAnn2 (Phvul.011g209200)clustered in the subclade with Psat4g191040, named PsAnn2.Two previously described pea annexins induced in rootsin response to rhizobial inoculation  were identified as proteins corresponding to Psat5g217440and Psat2g074960 coding sequences using a new databasehttps://urgi.versailles.inra.fr/blast for P. sativum (see Table 2). The phylogenetic analysis depicted anadditional branch in the phylogenetic group with MtAnn1/PvAnn1 and MtAnn2/PvAnn2, comprising MtAnn4(Medtr3g018780), PvAnn4 (Phvul.005g030100), and theirhomolog Psat5g217440, named PsAnn4 (identified by proteomicscreening) (see Table 2). Another previously foundpea annexin, Psat2g074960, might be closely related toMedtr5g063670 and Phvul.008G173100.1, defined as MtAnn8and PvAnn8 based on phylogenetic analysis (see Table 2).Analysis of the domain composition of pea annexinsand modeling of three-dimensional structureof PsAnn4 and PsAnn8Analysis of the domain composition of the correspondingproteinsin pea showed the presence of four typical domainsof plant annexins . This suggests that the annexin genefamily indeed comprises several members in pea. Althoughplant annexins have four putative annexin repeats, not all Ca2+-binding motifs in these repeats seem to be functional. In plantannexins, the Ca2+-binding site is highly conservative in thefirst (I) repeat but is not conservative in the second (II) andthird (III) repeats, while in the fourth (IV) repeat moderateconservatism is preserved .The crystal structure of the Gossypium hirsutum annexinGhAnn1 bound to calcium was obtained in an earlier study. Since PsAnn4 and PsAnn8 may be involvedin regulation of pea-rhizobial symbiosis, we modeled thethree-dimensional (3D) structure of these two annexins usingGhAnn1, with 50 % sequence identity for PsAnn4 and 78 %sequence identity for PsAnn8 as a template .The resulting 3D structures of PsAnn4 and PsAnn8 proteinsindicated the coordination of calcium ions in the first andfourth annexin repeats. In the first repeat of both proteins, thecalcium-binding site of the type II was coordinated by threecarbonyl oxygen atoms of the residues Phe-23, Gly-25, andGly-27, and carboxylate of Glu-67 in PsAnn4 and PsAnn8, as was shown earlier for GhAnn1.We suppose that the second calcium ion is bound in the loopof the fourth annexin repeat of PsAnn4 and PsAnn8 proteins. Itis coordinated in the binding site of type II by Ile-254, Lys- 256,and Gly-258 in pea annexins . The thirdcalcium ion (in the binding site of type III) is coordinated bytwo oxygen atoms of the residues Val-296 and Thr-299 andcarboxylate of Glu-304 in this protein   . However, in thefourth repeat of PsAnn4 protein, the Val-296 is replaced bySer and Glu-304 by Lys . This might potentially obstruct the binding of the calcium ion, as was shown in ourmodeling . Although we cannot rule out that thismight be due to low homology between PsAnn4 and GhAnn1,which was used as a template in the modeling, the results suggestthe potential difference in Ca2+ binding between PsAnn4and PsAnn8 proteins.Comparative analysis of protein patternsin wild-type and non-nodulating pea mutantTo verify whether the stimulation of synthesis of PsAnn4 andPsAnn8 proteins depends on Nod factor perception, the proteinpatterns were analyzed in wild-type pea cv. Frisson anda P56 mutant with a defective sym10 gene (which encodes aputative Nod factor receptor) .Two-dimensional differential in-gel electrophoresis-basedproteomics was used to characterize the pattern of proteindistribution . Two spots corresponding to the locationof the previously characterized annexins  were excised from the gel. Mass spectrometric analysisconfirmed their identity to annexins Psat5g217440 (PsAnn4)and Psat2g074960 (PsAnn8). Enhanced level of PsAnn4 wasfound in the inoculated roots of wild type pea plants (cv. Frisson)compared to the inoculated P56 mutant roots.The amount of PsAnn8 protein was also slightly higher inresponse to inoculation in the wild type than in the P56 mutant,but not as essential as for PsAnn4. In accordance with this,low amounts of PsAnn4 and PsAnn8 proteins were found inthe roots of the P56 mutant and didn\u2019t change in response toinoculation. This suggests that the up-regulation of both annexinsmay depend on Nod factor recognition in pea plantsand may be connected with the functioning of these annexinduring symbiotic interaction of plants with rhizobia at earlystages. Since the increase in the amount of PsAnn4 proteinwas more significant in response to inoculation, we focusedon this annexin in our next experimentsPsAnn4 expression pattern in response to rhizobialinoculation and treatment with Ca2+ inhibitorsThe PsAnn4 expression pattern in response to rhizobial inoculationwas analyzed in our experiments . A quantitativeRT-PCR analysis revealed that Rhizobium infectionenhanced the PsAnn4 gene expression at the early stages ofnodulation, starting from 1\u20133 days after inoculation up to5 days after inoculation, but thereafter their transcript levelsdid not significantly change upon nodule development . In our experiments the expression of another annexingene, PsAnn1a, the closest homolog of MtAnn1 genewas also analyzed . As it was expected, thePsAnn1a gene expression was primarily enhanced at theearly stages of symbiosis development and reached the highestlevels in the nodules. Similar pattern had been previouslyfound for MtAnn1 .Therefore, up-regulation of PsAnn4 expression may be relatedto the early stages of nodulation. The upregulation of thePsAnn4 transcription level was not as significant as it wasat the protein level, which implies that the regulation of thisannexin can be mainly achieved at the post-transcriptionaland translational level.To verify the influence of calcium inhibitors on the regulationof PsAnn4 gene, its expression level was estimated after plant treatment with the Ca2+ channel blocker LaCl3 .Two previously described as symbiosis-specific genes PsNINand PsEnod5 were also used in our experiments as a controlfor effective inoculation. In pea roots, the upregulation ofPsAnn4 expression in response to inoculation was revealedin 1 dai, corresponding with experiments on the dynamics ofthis gene expression upon nodulation. The significant decreasein the expression of PsAnn4 was found in our experiments inthe presence of LaCl3. Down-regulation of symbiosis-specificgenes PsEnod5 and PsNIN was also observed, which indicatedthe importance of Ca2+ influx for their regulation. Therefore,the influx of calcium ions into the cell, which is observed atthe early stages of symbiosis development, may affect theexpression level of PsAnn4 in pea roots .Subcellular localization of pea PsAnn4 annexinTo follow the PsAnn4 protein localization in plant cells, itwas fused to the fluorophores such as red fluorescent protein(RFP) and yellow fluorescent protein (YFP) at the C-terminusand expressed under the transcriptional regulation of the 35Spromoter in N. benthamiana leaves by infiltration with A. tumefaciens. The infiltration of constructs for thesynthesis of proteins fused with RFP and YFP allowed us tovisualize the protein in leaf tissues after transformation. In thecells of N. benthamiana leaves, PsAnn4 protein was localizedin the plasma membrane or in the cell wall. In addition, we alsoestimated the presence of PsAnn4 in different cell fractionsby Western-blot hybridization using anti-YFP or anti-RFPantibodies. PsAnn4-YFP was found in insoluble fraction ofleaf tissue pelleted at 36 000 g . It suggests thatPsAnn4 may be involved in cell wall or membrane modificationas well as in ion transport.DiscussionAvailable pea genome information  allowedus to determine the composition of the annexin genefamily in this legume. Database searches revealed 15 annexingenes in P. sativum L., 18 in M. truncatula as well as 13 inboth P. vulgaris and L. japonicus. Based on the phylogeneticanalysis of these annexins, close homologs can be identifiedamong these legume species .At present, only one pea annexin, p35, has been functionallycharacterized . The localization of thisannexin in root cells involved in active secretion suggests itsfunction in exocytosis. Subsequently, the use of antibodiesagainst this protein revealed its localization in epidermalcells of the leaf and stem . However,annexins involved in nodulation have not been characterizedin P. sativum. In contrast, in M. truncatula, two annexins,MtAnn1 (Medtr8g038210) and MtAnn2 (Medtr8g038220),demonstrated a high level of expression during nodulationand were found to be involved in controlling bacterial infectionand nodule organogenesis .Another annexin, MtAnn3 (Medtr4g097180), was found to beimportant for root hair deformations in M. truncatula . At the same time, close homologs of MtAnn1 \u2013PvAnn1 (Phvul.011g209300) and LjAnn1 (Lj0g3v0203419),which belong to the same phylogenetic group as MtAnn1, playimportant roles in the symbiotic process in P. vulgaris andL. japonicus .In our earlier work, two annexins activated at the earlystages of symbiosis development in pea were found using theproteomics approach . This approachmight be helpful for the identification of new regulators ofsignal transduction pathways at the initial stages of nodulationin pea. Our present analysis revealed that these two identifiedannexins of pea belong to different phylogenetic groups,defined as homologs of MtAnn4, PvAnn4 and MtAnn8,PvAnn8, respectively. Although PsAnn4, and MtAnn4 andPvAnn4 have high levels of homology with MtAnn1 andPvAnn1, they belong to another group of annexins basedon phylogenetic analysis. PsAnn8 belongs to a less studiedphylogenetic group. Therefore, two previously unknown annexinswere identified in our study. In addition to stimulationduring rhizobial inoculation, the dependence of PsAnn4 andPsAnn8 activation on the LysM-receptor-like kinase SYM10,encoding a putative Nod factor receptor, was revealed in thepresent study , which suggested that rhizobialsignaling molecules Nod factors may be important for theiractivation. It also suggests the participation of these twoannexins in the development of the symbiotic interaction ofplants with rhizobia.Phylogenetic analysis and prediction of the overall 3Dstructure of PsAnn4 and PsAnn8 proteins showed differencesin the Ca2+-binding motif in the fourth annexin repeat of theseproteins, and therefore, in the potential ability to bind calciumions. This can potentially influence the binding of these annexinsto phospholipids by means of a calcium bridge mechanism.It was predicted that three calcium ions were coordinatedin the first and fourth repeats, which is consistent with the dataof the canonical binding of the G. hirsutum annexin GhAnn1and animal annexins to the phospholipids of membranesusingthe mechanism of calcium bridges . Inthe predicted structures of Arabidopsis annexins , the canonicity of the Ca2+-bindingmotif in the first repeat and the presence of modified motifsin the fourth repeats of AtAnn1 and AtAnn3 were also shown,while AtAnn4 had no recognizable Ca2+ \u2013 or phospholipidbindingmotifs .Since the level of PsAnn4 synthesis in response to inoculationwas more significant in the roots of wild type pea plantscompared with mutant defective in symbiosis, we carried outthe analysis of this annexin in more detail. It was shown thatthe regulation of PsAnn4 annexin in pea could be achievedat the transcriptional level as well as post-transcriptionaland translational levels, probably. Significant activation ofMtAnn1 and MtAnn2 gene expression level was found in theroots of M. truncatula treated with Nod factors or inoculated with rhizobia . Meanwhile, theexpression of PvAnn1 in P. vulgaris was slightly upregulatedin developing nodules . However,a phosphoproteomic approach revealed that PvAnn1 wasa phosphorylated protein with enhanced levels of synthesisduring nodulation . Hence, theregulation of annexins involved in nodulation might be differentand is probably connected with different functions thatannexins fulfil in this processLocalization of annexins might differ depending on theirfunction. Some annexins show cytoplasmic and nuclear localization,while other annexins are associated with variousplant membranes, including the plasma membrane, endoplasmicreticulum, and nuclear membrane . Some annexins maybe embedded in the membrane in the form of monomers oroligomers. One of the distinctive characteristics of annexinsis their ability to change their cellular localization in responseto various stimuli. In our experiments, the localization ofannexin 4 (PsAnn4) in the cell wall or plasma membranewas shown, suggesting the participation of this annexin inprocesses associated either with membrane modification orion transport at the early stages of symbiosis establishment inpea. Similarly, the localization of the other annexin, MtAnn2,involved in nodulation in M. truncatula, was revealed to beassociated with the plasma membrane, particularly with lipidrafts from root plasma membrane preparations . In addition, the annexin PvAnn1 is essential forROS-dependent regulation of Ca2+ influx into the cells ofP. vulgaris, which strongly suggests the localization of thisprotein in the plasma membrane. Therefore, specific subcellularlocalization of annexins might be associated with theirfunction signal transduction at the early stages of symbiosis.In this study, phylogenetic analysis of the pea annexinsPsAnn4 and PsAnn8 was performed based on their homologywith annexins from other legumes. The modeling approachallowed us to estimate the structural features of these annexinsthat might influence their functional activity. To verify thefunctions of these annexins, we performed comparative proteomicanalysis, experiments with calcium influx inhibitors,and localization of labeled proteins. Essential down-regulationof PsAnn4 synthesis in a non-nodulating pea mutant P56(sym10) suggests an involvement of this annexin in the rhizobialsymbiosis. 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+{"text": "Background: Coronavirus disease (COVID-19) has spread rapidly since 2019. Approximately 15% of the patients will develop severe complications such as multiple organ disease syndrome related to cytokine release syndrome (CRS). Continuous renal replacement therapy (CRRT) can remove inflammatory cytokines through filtration or adsorption. We evaluated the effectiveness of CRRT in COVID-19 patients with CRS.Methods: This retrospective, multicenter, descriptive study included 83 patients with CRS from three hospitals in Wuhan.Results: In COVID-19 patients with CRS, the fatality rate was even higher in CRRT group (P=0.005). However, inflammatory markers such as C-reactive protein, neutrophil counts, and D-dimer decreased after CRRT (P<0.05). Results of Lasso model showed that tracheotomy (\u03b2 -1.31) and convalescent plasma (\u03b2 -1.41) were the protective factors. In contrast, CRRT (\u03b2 1.07), respiratory failure (\u03b2 1.61), consolidation on lung CT (\u03b2 0.48), acute kidney injury (AKI) (\u03b2 0.47), and elevated neutrophil count (\u03b2 0.02) were the risk factors for death.Conclusions: Our results showed that although CRRT significantly reduced the inflammation, it did not decrease the fatality rate of patients with CRS. Therefore, the choice of CRRT indication, dialysis time and dialysis mode should be more careful and accurate in COVID-19 patients with CRS. An outbreak of coronavirus disease COVID-19) in December 2019 in Wuhan, China, has developed into a global pandemic. COVID-19 is caused by infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) virus 9 in Dece. AlthougCytokine release syndrome (CRS) plays a vital role in the occurrence and disease progression of the COVID-19 . It has Although there are no standard diagnostic criteria for CRS, some treatments based on managing CRS have been widely applied clinically. These include antagonism at the IL-6 receptor (using the antagonist Tocilizumab), artificial-liver blood-purification therapy that can remove cytokines from the circulation, and continuous renal replacement therapy (CRRT) can remove inflammatory factors such as cytokines from the circulation via filtration and adsorption . TocilizOf the 83 cases included in our study, 67 were classified as critical, and 16 as non-critical. Of the 67 critical patients, 38 cases were treated with CRRT during hospitalization. A total of 45 patients died, and 36 were cured or improved. The outcome of two patients is unknown.The mean age of the patients was 67.3\u00b112.6 years. Almost three-quarters (73.5%) of the patients were male; 67 were critical cases, and 13.3% of the patients had a history of smoking. The underlying medical conditions of the patients included hypertension (55.4%), diabetes (24.1%), tumors (6%), and chronic obstructive pulmonary disease (1.2%). Pulmonary CT revealed that 51.7% of the critical patients had consolidation, and 73.3 % of non-critical patients had ground-glass changes in the lungs. The mean peak creatinine value during hospitalization was 133.2 pg/mL . AKI was present in 60.2%, MODS in 36.1%, and ARDS in 42.2% of the patients. Compared to the non-critical groups, the critical group had more patients with an IL-6 value >4000 pg/mL (29.9% vs.18.8%) .Patients in the critical group had a significantly higher WBC count (P=0.001), neutrophil count (P<0.001), neutrophil percentage (P<0.001), LDH (P<0.001), peak creatinine (P=0.01), high-sensitivity troponin I (P<0.001), CRP (P<0.001), ferritin (P=0.001) and procalcitonin (PCT) (P=0.043) than those in the non-critical group. Patients in the critical group had significantly lower lymphocyte counts (P=0.018) and albumin levels (P=0.015) than patients in the non-critical group .Treatments such as CRRT (P=0.001), antibiotics (P=0.002), hormones (P=0.004), globulin therapy (P<0.001), invasive mechanical ventilation (P<0.001) and non-invasive ventilation (P<0.001), were each more commonly used in critical patients than in non-critical patients. Moreover, critical patients had a higher incidence of AKI (P<0.001), respiratory failure (P<0.001), ARDS (P<0.001), MODS (P<0.001), gastrointestinal bleeding (P=0.004), acute liver dysfunction (P<0.001) and acute myocardial injury (P<0.001) .Over half of the critical patients (38/67) were treated with CRRT. Those who received CRRT had a lower blood platelets (PLT) (P=0.023), and lower albumin (P=0.041), a higher peak creatinine (P=0.018), and a greater incidence of MODS (P<0.001) than those who did not receive CRRT. More patients receiving CRRT also underwent tracheal cannulation (P=0.003) or invasive mechanical ventilation (P=0.002). Most patients who received CRRT (64.7%) had pulmonary consolidations on CT . Unexpec9g/L vs. 0.64  109g/L] followed the same pattern, demonstrating lower values in the CRRT patients. The CRP was higher in the CRRT than in the non-CRRT group [97.7  mg/L vs. 89.16  mg/L)]. Moreover, the incidence of AKI and ARDS in critical patients treated with CRRT was 81.1% and 71.4%, respectively, which was higher than the corresponding figures of non-CRRT . The blood oxygen saturation in patients who received CRRT was lower than in the non-CRRT group [90 % vs.93 %] and the lymphocyte count , and differences between the groups were determined using a two-sample t-test or Wilcoxon rank-sum test. Lasso and accurate logistic analysis were conducted to identify the factors related to death. The explanatory variables included treatments , respiratory failure, consolidation on lung CT, AKI, elevated neutrophil percentage, cough, dyspnea, hormone treatment, invasive mechanical ventilation, acute myocardial injury, White blood cell (WBC) count (x 10The data underlying this article are available in the article and its online supplementary material."}
+{"text": "Trichoderma are well-studied filamentous fungi generally observed in nature, which are widely marketed as biocontrol agents. The secondary metabolites produced by Trichoderma have gained extensive attention since they possess attractive chemical structures with remarkable biological activities. A large number of metabolites have been isolated from Trichoderma species in recent years. A previous review by Reino et al. summarized 186 compounds isolated from Trichoderma as well as their biological activities up to 2008. To update the relevant list of reviews of secondary metabolites produced from Trichoderma sp., we provide a comprehensive overview in regard to the newly described metabolites of Trichoderma from the beginning of 2009 to the end of 2020, with emphasis on their chemistry and various bioactivities. A total of 203 compounds with considerable bioactivities are included in this review, which is worth expecting for the discovery of new drug leads and agrochemicals in the foreseeable future. Moreover, new strategies for discovering secondary metabolites of Trichoderma in recent years are also discussed herein.Fungi play an irreplaceable role in drug discovery in the course of human history, as they possess unique abilities to synthesize diverse specialized metabolites with significant medicinal potential.  Trichoderma is a fungal genus that was first described in 1794 , and six known trichothecenes, trichodermol (9), trichodermin (10), trichoderminol (11), trichodermarin A (12) and trichodermarin B (13), and 2,4,12-trihydroxyapotrichothecene (14), were isolated from Trichoderma brevicompactum ADL-9-2, which was obtained as an endophyte of marine algae Chondria tenuissima (8), featuring a 2\u2019-N-acetylglucosaminyl moiety, represented the first aminoglycoside-bearing trichothecene. Chemical investigations of the marine fungus Trichoderma cf. brevicompactum TPU199 fermented with NaI afforded three new trichothecenes, trichobreols A\u2013C (15\u201317) , whereas compounds 16 and 17 were found only under NaI-containing culture conditions. Additionally, isolation of the same fungus yielded two new trichothecenes, trichobreols D (18) and E (19) (20) and B (21), two trichothecenes linked with octa-2,4,6-trienedioyl moiety, were isolated from the biofertilizer fungus T. brevicompactum (CGMCC19618) , 8-deoxy-trichothecin (23), and trichothecinol B (24) were yielded  . Interesd E (19) . HarzianCC19618) . From a  yielded .T. virens Y13-3, eight undescribed carotane sesquiterpenes, trichocarotins A\u2013H (25\u201332), along with the known compounds CAF-603 (33), trichocarane B (34), 7-\u03b2-hydroxy CAF-603 (35), and trichocarane A (36), were discovered (From the marine-derived fungus scovered  (Shi et scovered . Carotanscovered .37), containing a previously unrecognized site of an exocyclic olefin functionality at C-10, was obtained from the marine-derived fungus T. virens Y13-3 , and three known ones, i.e., aspergilloid G (41), rhinomilisin E (42), and rhinomilisin G (43), were characterized from the marine sponge-derived fungus Trichoderma sp. SM16 , was produced by the marine fungus T. harzianum (XS-20090075), which was isolated from soft corals -isocyclonerotriol (45) and (10Z)-isocyclonerotriol (46), which were characterized as the first example with an isomerized ring in cycloneranes (47), methyl 3,7-dihydroxy-15-cycloneranate (50), and 10-cycloneren-3,5,7-triol (51), as well as two structurally related cycloneranes, 9-cycloneren-3,7,11-triol (48) and (\u2013)-cyclonerodiol (49), were obtained from T. harzianum X-5, an endophyte of the marine alga Laminaria japonica  and 10(E)-cyclonerotriol (52), were obtained from T. longibrachiatum, an endophyte of the highly halophile Suaeda glauca , II (54), and III (55) (56), 3-oxoneomacrophorins I (57) and II (58), neomacrophorin VII (59), 5\u2032-epimacrophorin B (60), and 5\u2032-deoxyneomacrophorin IV (61), as well as four novel premacrophorin congeners, i.e., premacrophorin III (62), premacrophorindiol (63), premacrophorintriols I (64), and II (65), were isolated from the same fungus (56 and 57) or 2,3-epoxybenzosemiquinol substructures (58\u201360 and 62\u201364). Premacrophorins 62\u201365 carried acyclic isoprenoid side chains biosynthetically derived from neomacrophorins in the early stage, rather than the common drimane skeleton.Chemical exploration of an endophyte III (55)  (Hirose III (55) . They bee fungus . These mTrichoderma sp. 307 and aquatic pathogenic bacterium Acinetobacter johnsonii B2 afforded two undescribed furan-type isoeremophilane sesquiterpenes, microsphaeropsisins B (66) and C (67) , trichoacorenol (69), and trichoacorenol B (70) were isolated from marine-derived T. harzianum X-5, and they were structurally characterized as acorane sesquiterpenes (71), an unusual norsesquiterpene with a novel tricyclic-6/5/5--decane framework, was characterized to originate from T. longibrachiatum (72) and B (73), were produced by T. atroviride S361, an endophyte of Cephalotaxus fortunei  and B (75), were obtained from T. brevicompactum ADL-9-2, which was isolated from marine algae C. tenuissima , were isolated from T. asperellum residing in Panax notoginseng   and B (81)  . It is bterpenes . Trichodachiatum . Two newfortunei . Compounnuissima . Compounoginseng . The newe C (79) . Chemicad B (81) . InteresTrichoderma species (82\u201386), and three previously reported diterpenes, i.e., 3S-hydroxyharzianone (87), harziandione (88), and harzianol A (89), were isolated from the endophyte T. atroviride B7 of Colquhounia coccinea var. mollis (90), an undescribed harziane lactone, was isolated from an endophyte T. longibrachiatum A-WH-20-2 of marine algae Laurencia okamurai (91), was obtained from the soft coral-sourced T. harzianum (XS-20090075) by chemical epigenetic manipulation strategy , was produced by T. harzianum X-5 -dihydro-harzianone (93) and harzianelactone (94), were produced by Trichoderma sp. Xy24 from mangrove plant Xylocarpus granatum , as well as five new lactones, i.e., harzianones A\u2013D (97\u2013100) and harziane (101) (102), was isolated from Trichoderma erinaceum derived from Acanthaster planci  . Finallyr planci .103), a rare norditerpene, was produced by an endophyte T. citrinoviride cf-27 , was characterized from the marine algicolous fungus T. harzianum X-5 , characterized as a novel chlorinated cleistanthane diterpenoid, was isolated from the marine fungal strain T. harzianum (XS-20090075) (Trichoderma for the first time.Citrinovirin (de cf-27 . Compounanum X-5 . 104 and0090075) . TricyclT. harzianum KZ-20 afforded four new cyclodepsipeptides belonging to the destruxin family, i.e., trichodestruxins A\u2013D (106\u2013109), and two previously reported derivative, i.e., destruxin E2 chlorohydrin (110) and destruxin A2 (111) , trichomide B (113), and homodestruxin B (114), characterized as cyclohexadepsipeptides of the trichomide series, were produced by T. longibrachiatum (115) and halobacillin (116) were obtained from the endophyte T. asperellum (Bioassay-guided fractionation of the plant endophytic fungus A2 (111)  (Liu Z. A2 (111) . Destruxachiatum . Finallyperellum .Trichoderma sp. TPU199 is a producer of a series of diketopiperazines (117\u2013127) (117), gliovirin (118), and trichodermamide A (119). 117 and 118 possessed an unusual epipolythiodiketopiperazine (ETP) skeleton. Then, this strain with sodium halides added to the culture medium afforded the halogenated gliovirin-type ETPs DC1149B (120), DC1149R (122), and iododithiobrevamide (123). Subsequently, chlorotrithiobrevamide (124), the first trisulfide derivative in the type of ETP, was characterized. Furthermore, a highly modified dipeptide, dithioaspergillazine A (125), was obtained after the long time cultivation. Finally, two undescribed ETPs, i.e., 5-epi-pretrichodermamide A (126) and 5-epi-trithiopretrichodermamide A (127), were characterized under NaI-containing culture conditions. Pretrichodermamide G (128) was established as a 1,2-oxazadecaline ETP, and it was identified from the endophyte T. harzianum of Zingiber officinale (129), and a biogenetically related metabolite aspergillazin A (130) were produced by the marine-sourced T. harzianum D13 (129 and 130 were novel ETP derivatives with the sulfur bridge locating at different positions. Dehydroxymethylbis(dethio)bis(methylthio)gliotoxin (131) and -6-(para-hydroxybenzyl)-1,4-dimethyl-3,6-bis(methylthio)piperazine-2,5-dione (132), which were structurally characterized as two undescribed sulfurated diketopiperazines, were produced by an algicolous isolate of T. virens Y13-3  (133) (The marine fungus 117\u2013127)  (Yamazak117\u2013127) . Initialficinale . A rare anum D13 . Notablyns Y13-3 . The funo) (133) .T. virens FKI-7573 generated an undescribed N-containing compound, i.e., trichothioneic acid (134) (134 contained a heptelidic acid and an L-ergothioneine substructure. Ethyl 2-bromo-4-chloroquinoline-3-carboxylate (135) was produced by the soft coral-sourced T. harzianum (XS-20090075) in Czapek\u2019s medium  and B (137), isolated as stereoisomers originating from the PKS-NRPS mixed pathway, were isolated from T. gamsii, an endophyte of P. notoginseng (138) and B (139), were identified from Trichoderma sp. strain MF106 from the Greenland Seas (140), a nitrogen heterocyclic siderophore, was isolated from T. harzianum M10 -1-methyl-3,5-dioxopyrrolidin-2-ylmethyl]-3-methyl-butyric acid. Atrichodermone A (141) was a unique compound with a dimeric cyclopentenone framework that was discovered from the endophytic fungal strain T. atroviride  and 5\u2032-acetoxy-deoxycyclonerin D (143), were obtained from the marine fungus T. asperellum A-YMD-9-2   (Miyano id (134) . 134 cons medium . 135 wasoginseng . Two rarand Seas . Harziananum M10 . 140 wasroviride . 141 was-YMD-9-2 .144), a natural product, methyl-trichoharzin (145), and the known trichoharzin (146) and eujavanicol A (147) were produced by the marine fungus T. harzianum XS-20090075  was identified from a fungal strain of T. harzianum F031  and B (150) were isolated from an endophytic fungus T. spirale, and characterized as new octahydronaphthalene derivatives , 1,6-di-epi-koninginin A (152), 15-hydroxykoninginin A (153), 10-deacetylkoningiopisin D (154), and koninginin T (155), along with two previously reported derivatives, koninginin L (156) and trichoketide A (157), were produced by Trichoderma koningiopsis QA-3 (151\u2013153 were characterized as tricyclic polyketides with an octahydrochromene skeleton. Koninginins I (158), J (159) and K (160), which were structurally characterized as new koninginin-type compounds, were produced by T. neokongii 8722  and trichodermaketones A\u2013D (162\u2013165) (162 and 163 represented unprecedented tricyclic polyketides having a bistetrafuran skeleton. Trichoketides A (166) and B (167), two undescribed octaketides, were isolated from Trichoderma sp. TPU1237  (herein reported as a new compound) and M (168) were isolated from solid fermentation of T. koningii 8662  and M (168) was an oxygen bridge located between the C-10 and C-7 positions.Five new polyketides, sis QA-3 . Compoungii 8722 . Chemica162\u2013165) . 162 and TPU1237 . 166 andgii 8662 . A serie169) and trichoderone B (170), together with three previously reported cytochalasans, aspochalasins D (171), J (172), and I (173), were obtained from T. gamsii from P. notoginseng  and D (175), as well as three known cytochalasans, aspochalasins D (171), M (176), and P (177), were obtained from the abovementioned strain T. gamsii , was isolated from endophytic T. atroviride (179), was produced by the marine-sourced Trichoderma sp. HPQJ-34  and B (181), were found from the marine fungus T. harzianum (XS-20090075) (180 and 181 had a C9 polyketide framework with a \u03b3-lactone moiety. Trichoderpyrone (182), a novel cyclopentenone-pyrone mixed polyketide, was produced by T. gamsii , a new cyclopentenone, was obtained from the marine Trichoderma sp. , which was identified as a new isocoumarin derivative with a 6,8-dihydroxyisocoumarin moiety, was isolated from T. citrinoviride A-WH-20-3 (187) and 4,6-dihydroxy-5-methylphthalide (188), were produced by the fungus T. harzianum F031 (189 and 190) with a 1-hydroxyhepta/methoxyhepta-3,5-dien-2-one moiety were produced in the transformant of Trichoderma afroharzianum (191) and E (192), were obtained from dragonfly associated T. harzianum QTYC77 (193), elucidated as but-2-enoic acid 7-acetoxy-6-hydroxy-2- methyl-10-oxo-5,6,7,8,9,10-hexahydro-2H-oxecin-5-yl ester, was isolated from cultural filtrates of Trichoderma cremeum -7-hydroxy-de-O-methyllasiodiplodin (194) and (3R)-5-oxo-de-O-methyllasiodiplodin (195), were isolated from the cocultivation of Trichoderma sp. 307 and A. johnsonii B2 (196), and the known nafuredin A (197), were produced by marine fungus T. harzianum D13  . Compoun. gamsii . 182 haderma sp. . Compoun Fes1712 . Trichop-WH-20-3 . Previounum F031 . Two strarzianum . Two undm QTYC77 . A new 1 cremeum . Two undsonii B2 . An undeanum D13 .198) and B (199), were produced by a culture broth of T. polypori FKI-7382  and II (201). Notably, both of them were characterized from Trichoderma for the first time (202) and B (203), two new compounds with 4-(2-hydroxyethyl) phenol moieties, were isolated from an endophyte T. gamsii  associated with secondary metabolites are cryptic  or expressed at very low levels under general laboratory conditions . TherefoTrichoderma sp. TPU199 was found to produce a series of diketopiperazines under different conditions , 122 (Br derivative of 117), and 123 (I derivative of 177) when added with NaCl, NaBr, and NaI in culture medium, respectively. Moreover, TPU199 supplemented with DMSO yielded 124, a new trithio derivative of 120. A continuous study indicated that, with the long time cultivation, an undescribed modified dipeptide 125 was obtained. Finally, two undescribed ETPs 126 and 127 were characterized under NaI-containing culture conditions. It is undoubtedly proven that changing the culture conditions can activate cryptic metabolic pathways.The marine-derived fungus nditions  (Yamazaknditions . ChemicaTrichoderma sp. 307 and pathogenic bacterium A. johnsonii B2 yielded two undescribed sesquiterpenes (66 and 67) and de-O-methyllasiodiplodin (194 and 195) . This is the first report of cleistanthane diterpenoids isolated from Trichoderma species. This study provided solid example to show that it is efficient to activate the silent genes of Trichoderma species by chemical epigenetic manipulation.Microorganism coculture based on microbial interspecies competition is an efficient path to stimulate cryptic BGCs. Cocultivation of and 195) . HPLC anabolites . A histoT. afroharzianum, a laeA-like gene overexpression transformant was built (189 and 190). This study indicated that transcriptional control could be a considerable strategy in activating more secondary metabolites and enhancing the silent potential metabolism of Trichoderma species.The transcriptional control has also proven to be an effective approach. To activate the chemical potential of the endophytic fungus as built . Further1\u2013203 are listed in The producing fungus, environmental source, and bioactivities of compounds 1\u201314 were assayed for antifungal activity against Botrytis cinerea, Cochliobolus miyabeanus, Fusarium oxysporum f. sp. cucumerium, F. oxysporum f. sp. niveum, and Phomopsis asparagi  among these trichothecenes indicated that the acetoxy and methyl functionalities (compound 10) were necessary, while the epoxide moiety and the ether linkage were other possibilities   exhibited potent effect on Staphylococcus aureus, Bacillus subtilis, and Micrococcus luteus, with EC50s of 7.7, 7.7, and 9.9 \u03bcg/mL, respectively  was active against B. subtilis , Staphylococcus epidermidis (24.28 \u03bcM), and C. albicans (25.38 \u03bcM) (148) displayed mild activity against Colletotrichum gloeosporioides, with an MIC value of 128 \u03bcg/mL  was inactive against C. albicans (MIC > 125 \u03bcg/mL). However, it was active at 125 \u03bcg/mL when treated with 0.05 \u03bcg/mL ketoconazole  . Further10 \u03bcg/mL . The nov 8 \u03bcg/mL . The newectively . It seem.4 \u03bcg/mL . Cyclopeectively . The new5.38 \u03bcM) . Trichoh28 \u03bcg/mL . Polyket64 \u03bcg/mL . Trichodconazole . New iso32 \u03bcg/mL . Polyket32 \u03bcg/mL . The newd 7.0 mm . The newa solani . Nafuredectively .Amphidinium carterae, Heterocapsa circularisquama, Heterosigma akashiwo, and Prorocentrum donghaiense) of trichothecene derivatives 1\u201314 was evaluated. Notably, 10 featured the strongest effect, with IC50s of 1.7, 0.82, 0.91, and 1.4 \u03bcg/mL - and (10Z)-cyclonerotriol, the isomerization of the five-membered ring greatly suppressed their antimicroalgal activities , while the new proharziane diterpene 104 potently inhibited with IC50s of 1.2\u20134.3 \u03bcg/mL , H. akashiwo (9.1 \u03bcg/mL), and P. donghaiense (5.9 \u03bcg/mL)  .38\u201343 were evaluated against NCIH-460, NCI-H929, and SW620 cell lines  showed cytotoxicity against human adenocarcinoma cells (COLO 201) with an IC50 of 46 \u03bcg/mL  was observed to exhibit moderate cytotoxicity against NCI-H1975 , HepG2 (60.88 \u03bcM), and MCF-7 (53.92 \u03bcM) cell lines  , HepG2 (30.8 \u03bcM), and HeLa (33.9 \u03bcM)  , c. The 33.9 \u03bcM) . Cyclopeell line . The sel20) and B (21) decreased the shoot and root lengths of the dicot species Brassica chinensis and induced inhibitory effect of seed germination at 2 \u03bcg/mL . The results indicated that 20 and 21 possess potent herbicidal potential for dicotyledon and/or monocotyledon weeds. All of the isolated harzianes 95\u2013101 exhibited potent phytotoxicity at 200 ppm  lethal rates of 38.2 and 42.7% at 200 \u03bcg/mL  showed OH radical-scavenging and singlet oxygen-quenching ability in a dose-dependent manner, which was equivalent to those of positive controls (140) as a plant growth promoter was evaluated   .Trichoderma from 2009\u20132020. Their chemical structures were classified into terpenoids , cyclopeptides (104\u2013116), diketopiperazines (117\u2013133), alkaloids and other nitrogen-containing compounds (134\u2013143), polyketides , and other compounds (198\u2013203) according to their putative biogenetic sources. As shown in Trichoderma are considerable producing strains of novel terpenoids. It should be pointed out that some terpenoids, such as harzianes, are isolated exclusively from Trichoderma species. This review described 21 harziane diterpenes produced by Trichoderma. Considering their intriguing structures and bioactivities, much more attention should be devoted to this type of terpenoid in subsequent chemical studies.A total of 203 natural products were reported from Trichoderma comprises more than 340 species. Some of them are used as biocontrol agents, while some of them are promising producers of enzymes for industrial purposes. On the other hand, some Trichoderma species possess the unique capacity to synthesize various secondary metabolites with potent biological activities. In this review, a total of 17 identified species, including T. harzianum, T. brevicompactum, T. virens, T. gamsii, T. atroviride, T. longibrachiatum, T. asperellum, T. koningiopsis, T. koningii, T. citrinoviride, T. neokongii, T. spirale, T. afroharzianum, T. polypore, T. polyalthiae, T. erinaceum, and T. cremeum, are reported as the producing strains of the described metabolites. Among them, T. harzianum and T. brevicompactum were the most prolific strains, with 48 (23.76%) and 33 (16.34%) metabolites identified, respectively  have been characterized from T. brevicompactum.The genus ectively . The funTrichoderma is widely distributed and has been isolated in soils, decaying wood, and endophytes in the inner tissue of host plants. Previous studies have mainly focused on terrestrial species of Trichoderma. However, Trichoderma from the marine environment are unexploited. It would be useful to examine marine-derived Trichoderma species since they may be induced to produce specific metabolites in hyperhaline environments. Accordingly, in recent years, increasing attention has been devoted to marine Trichoderma. As shown in Trichoderma. From the above analysis, it can be concluded that marine environment and endophytes are more abundant sources of those productive strains.The genus 22 and 23 showed higher antifungal effect on C. lagrnarium  than the synthetic fungicide carbendazim  , anticancer (20.61%), and antimicroalgal (17.98%) activities were dominant in assessing the pharmacological potential of these metabolites . It shou2 \u03bcg/mL) . Cyclopeell line . The sel effects . These iTrichoderma from the beginning of 2009 to the end of 2020. As a result, a total of 203 metabolites are described herein, including their structural diversity and biological activities. Moreover, new strategies for discovering secondary metabolites of Trichoderma in recent years have also been discussed. Trichoderma has proven to be a treasure house of interesting secondary metabolites with medicinal importance. The biochemical studies of Trichoderma are untapped. Although a mass of metabolites have been isolated from Trichoderma species, the further excavation of those metabolites is worth expecting. By using new approaches to activate their silent gene clusters, including cultivation-based approaches, metabolomic profiling, and genome mining-based molecular approaches, an ever-increasing number of bioactive compounds will be obtained, which will be beneficial for the new drug discovery in the near future.In the present review, we offer a detailed summary of recently isolated metabolites from J-LZ and W-LT wrote this manuscript. Q-RH, Y-ZL, M-LW, L-LJ, CL, XY, H-WZ, and G-ZC collected and reorganized the literature data. X-XZ supervised the research work and revised the manuscript. All authors reviewed the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Uropathogen resistance, Fluoroquinolone-resistance (FQR) and Extended spectrum beta-lactamase (ESBL), has been observed to be emerging worldwide with prevalences above recommended thresholds for routine empirical treatment. We sought to determine recent resistance prevalence from a geographically diverse sample of US Emergency Departments (ED).Escherichia coli. We conducted a multi-center, observational cohort study utilizing a network of 15 geographically diverse US EDs. Patients \u2265 18 years of age with the primary international classification of diseases (ICD-10) diagnosis code of cystitis, pyelonephritis, or urinary tract infection (UTI) and were discharged home from the ED from 2018-2020 were included. We calculated descriptive statistics for uropathogens and susceptibilities. Logistic regression analysis was used to identify antimicrobial resistance risk factors associated with fluoroquinolone (FQ)-resistant E. coli was the most common pathogen (62.9%), followed by Klebsiella pneumoniae (13%) and Enterococcus species (5.8%). Across all sites, overall E. coli FQ-resistance prevalence was 22.1%, ranging from 10.5 to 29.7% by site. The prevalence of ESBL-producing uropathogen was 4.4%, ranging from 2.3% to 8.6% by site. Previous IV or oral antimicrobial use in the last 90-days and complicated vs. uncomplicated UTI were associated with FQ-resistant E. coli . Of the most prescribed oral antibiotics upon patients discharged from the ED, E. coli resistance to nitrofurantoin and cephalexin was 1.8% and 0.9%, respectively.Among 3,779 patients who met inclusion criteria, median age was 62.9 years (IQR: 41-77.6) and 76.3% were female. The most common diagnoses were complicated (40.9%) and uncomplicated cystitis (39.4%). Six hundred and forty-five (17%) patients reported receiving antimicrobials in the previous 90-days. E. coli is widely prevalent and ESBL-mediated resistance appears to be emerging across US sites highlighting the need for ongoing monitoring of antimicrobial resistance and, at some locations, modification of empirical treatments. FQ-resistant Brett Faine, PharmD, Spero Therapeutics (Research Grant or Support) Megan A. Rech, PharmD, MS, BCCCP, FCCM, Spero (Research Grant or Support) David A. Talan, MD, AbbVie (Consultant)GSK (Consultant)SPERO Therapeutics (Grant/Research Support)"}
+{"text": "Athelia (Fibularhizoctonia) sp. strain TMB , which forms termite-egg-mimicking sclerotia for which termites care. We further compare its repertoire of psilocybin gene homologs to homologs previously reported for Fibularhizoctonia psychrophila.Atheliales is a diverse order of crust-forming Basidiomycota fungi. Here, we report the draft genome of the \u201ccuckoo fungus,\u201d  Athelia (Fibularhizoctonia) sp. TMB strain TB5 , which forms termite-egg-mimicking sclerotia that termites tend. We further compare its repertoire of psilocybin gene homologs to homologs previously reported for Fibularhizoctonia psychrophila.Atheliales is a diverse order of crust-forming Basidiomycota fungi. Here, we report the draft genome of the \u201ccuckoo fungus,\u201d  Fibularhizoctonia psychrophila CBS 109695, associated with carrot spoilage  psilocybin gene cluster  is an ecologically diverse \u20134 order \u2013spoilage , 5, and Athelia (Fibularhizoctonia) sp. TMB  produces sclerotia  that chemically and structurally mimic Reticulitermes sp. eggs and receive care in over 70% of colonies of some Reticulitermes species . Sclerotia were cultured in potato dextrose broth at room temperature for 14\u2009days with shaking. Tissue was filtered through Miracloth, flash-frozen with liquid nitrogen, and pulverized with a mortar and pestle. Genomic DNA was immediately extracted using the DNeasy plant minikit (Qiagen). Short-read DNA libraries were prepared using the NEBNext Ultra DNA library preparation kit and were sequenced with a 150-bp paired-end format on a NovaSeq 6000 system (Illumina). Long reads were generated by fragmenting genomic DNA with a Covaris g-TUBE, preparing libraries using the SQK-LSK108 ligation sequencing kit, and sequencing the libraries on a MinION system (Oxford Nanopore Technologies) using an R9.4 flow cell. Sequencing generated 62,281,267 Illumina reads with an average coverage of 230.98\u00d7 and 228,721 MinION reads with a mean length of 3,983.38\u2009bp, a median length of 1,834\u2009bp, and a mean coverage of 10.94\u00d7. Coverage was calculated by mapping reads to the assembly with Bowtie 2 v2.4.1-2  data set (F. psychrophila expressed sequence tag (EST) and transcript data , Coniophora olivacea MUCL 20566 (https://mycocosm.jgi.doe.gov/Conol1), Coniophora puteana (https://mycocosm.jgi.doe.gov/Conpu1), Paxillus involutus ATCC 200175 (https://mycocosm.jgi.doe.gov/Paxin1), Pisolithus microcarpus 441 (https://mycocosm.jgi.doe.gov/Pismi1), Pisolithus tinctorius Marx 270 (https://mycocosm.jgi.doe.gov/Pisti1), Rhizopogon vesiculosus Smith (https://mycocosm.jgi.doe.gov/Rhives1), Scleroderma citrinum Foug A (https://mycocosm.jgi.doe.gov/Sclci1), Serpula himantioides MUCL 38935 (https://mycocosm.jgi.doe.gov/Serla_varsha1), and Suillus brevipes Sb2 v2.0 (https://mycocosm.jgi.doe.gov/Suibr2). Gene prediction was finalized using OrthoFiller v1.1.1 , C. olivacea MUCL 20566, F. psychrophila CBS 109695, Galerina marginata , Laccaria bicolor v2.0 (https://mycocosm.jgi.doe.gov/Lacbi2), P. croceum F 1598, Plicaturopsis crispa (https://mycocosm.jgi.doe.gov/Plicr1/Plicr1.home.html), Psilocybe serbica (https://mycocosm.jgi.doe.gov/Psiser1), and S. brevipes Sb2 v2.0.Illumina reads were trimmed using Trimmomatic v0.36  with the v1.0.11  and the  v1.0.11 . Genes w v1.0.11 , AUGUSTU v1.0.11 , Glimmer v1.0.11 , and Gen v1.0.11  via the  v1.0.11 . We refedata set , F. psycipt data , and pror v1.1.1  with defF. psychrophila and Athelia sp. TMB TB5 was calculated from the BLASTp . Protein domains were aligned (MAFFT v7.467 [Average amino acid identity (AAI) between e BLASTp  identitye BLASTp  with P. equences  were obtequences  domains T v7.467  with --aT v7.467 , -automaT v7.467 ). Close Athelia sp. TMB TB5 has a mean AAI of 82.58%  with F. psychrophila across 1,727 single-copy orthologs, consistent with substantial divergence time. Multiple copies of psilocybin decarboxylase, hydroxylase, and kinase in Athelia sp. TMB TB5 , and SRR12880627 (Illumina read library).The assembly and annotation of"}
+{"text": "Members of the interferon regulatory factor (IRF) gene family are crucial regulators of type I interferon signaling, which may play a role in the resistance of glioma to immune checkpoint blockade. However, the expression profiles, potential functions, and clinical significance of IRF family members remain largely unknown. Here, we examined IRF transcript levels and clinicopathological data from glioma patients using several bioinformatic databases, including ONCOMINE, GEPIA, TCGA, and cBioPortal. We found that IRF1, IRF2, IRF5, IRF8 and IRF9 were significantly upregulated in glioma compared to normal brain tissue. Higher IRF1, IRF2, IRF3, IRF4, IRF5, IRF7, IRF8 and IRF9 mRNA levels correlated with more advanced tumor grades and poorer outcomes. Moreover, although IRFs mutation rates were low (ranging from 0.5% to 2.3%) in glioma patients, genetic alterations in IRFs were associated with more favorable patient survival. Functional analysis showed that IRFs participated in glioma pathology mainly through multiple inflammation- and immunity-related pathways. Additionally, correlations were identified between IRFs and infiltration of immune cells within glioma tissues. Collectively, these results indicate that IRF family members, including IRF1, IRF2, IRF5, IRF8 and IRF9, may serve as prognostic biomarkers and indicators of immune status in glioma patients. Glioma is the most prevalent primary malignancy in the human brain and is characterized by high recurrence and lethality rates . AccordiRecent reports suggest that type I interferon signaling, which is regulated by interferon regulatory factors (IRFs), plays an important role in glioma resistance to immune checkpoint blockade , 6. The Recent advances in gene sequencing technology have enabled comprehensive analysis of IRF family members with existing bioinformatic tools. In this study, we performed an in-depth exploration of the expression patterns of IRF family members in glioma and evaluated their potential as prognostic biomarkers with the goal of improving molecular diagnosis and prognostic prediction for glioma patients.p = 0.008); two additional studies by Liang and Bredel found that IRF1 expression was increased 2.225- and 2.151-fold, respectively, in glioblastoma. IRF2 transcript levels were also higher in glioblastoma than normal brain tissues in two datasets from TCGA . The results of Sun\u2019s study suggested that IRF5 was increased 2.125-fold in diffuse astrocytoma (p = 1.33E-4) and 2.180-fold in anaplastic astrocytoma (p = 3.20E-5). Moreover, studies by Lee, Ramaswamy, and Bredel all found that IRF8 and IRF9 levels were significantly increased in glioblastoma or anaplastic oligoastrocytoma , IRF2 (p = 6.80E-18), IRF3 (p = 1.70E-23), IRF5 (p = 2.30E-08), IRF7 (p = 1.90E-29), IRF8 (p = 0.029), IRF9 (p = 4.80E-05) expression and pathological grade (p = 0.064). No correlation between IRF6 expression and pathological grade was observed (p = 0.49). For all of the identified correlations, expression increased as tumors progressed, suggesting that IRFs may play a role in glioma tumorigenesis and progression.Relationships between IRF family member expression and clinicopathological parameters of glioma patients were examined using data from the TCGA database. Among the 260 grade II, 267 grade III, and 173 grade IV glioma patients, significant correlations were observed between IRF1 , IRF2 (p = 2.22E-16), IRF3 (p = 1.43E-15), IRF4 (p = 0.004), IRF5 p = 5.84E-12), IRF7 (p = 6.85E-28), IRF8 (p = 0.001), and IRF9 (p = 0.000) transcript levels had significantly longer overall survival times   to 20.48% (17/83); mutations, deep deletions, and amplification were the most common types of alteration . For eac.237E-6) .We next explored potential co-expression among IRF family genes using data from TCGA glioma dataset. Pearson's correlation results revealed significant positive correlations between the following IRFs: IRF1 with IRF2 (r = 0.48) and IRF7 (r = 0.52); IRF2 with IRF1, IRF5 (r = 0.54), and IRF8 (r = 0.43); IRF3 with IRF7 (r = 0.41); IRF5 with IRF7 (r = 0.41) and IRF8 (R = 0.73); IRF7 with IRF9 (r = 0.74) . Little p < 1.0E-16; Next, we conducted a network analysis to examine potential internal interactions among IRF family genes as well as external interactions with other functionally related genes. PPI network analysis using STRING software revealed close protein-protein associations among the IRF family genes with 9 nodes, 32 edges, and an average node degree of 7.11  and 11 KEGG items were enriched. p < 0.01). Similar results were obtained for GBM. In addition, IRF1, IRF2, IRF6, and IRF9 were positively correlated with CD8+ T cell infiltration in LGG. However, IRF1, IRF5, IRF6, IRF7, IRF8, and IRF9 expression were negatively correlated with CD8+ T cell infiltration in GBM.Since the IRF family may regulate glioma progression and prognosis by participating in a wide range of inflammatory and immune responses, we undertook a comprehensive analysis of tumor immune infiltrates using the TIMER database. The results are shown in p = 0.002), CD8+ T cells (p = 0.042), IRF1 expression (p = 0.001), and IRF8 expression (p = 0.000) were significantly associated with the prognosis of LGG patients, while CD4+ T cells, dendritic cells (p =0.002), IRF1 expression (p = 0.01), IRF7 expression (p = 0.007), and IRF8 expression (p = 0.028) were associated with the prognosis of GBM patients , another online tool that contains RNA sequence expression data from 518 LGG samples, 163 GBM samples, and 207 normal brain samples [t-tests; p < 0.05 and fold change >2 were considered significant.We used a two-step analysis to assess IRF family member expression patterns in glioma patients. First, we examined mRNA level data from ONCOMINE, the largest public microarray database for genome-wide expression analysis . Data we samples . IRF expp-value <0.05 was considered significant.Correlations between IRF family member expression and clinicopathological characteristics and prognosis in glioma patients were evaluated using data derived from The Cancer Genome Atlas (TCGA) database, which contains both sequencing and clinical records for over 30 types of human cancers . A totalThe molecular characteristics and internal/external interactions of IRF family members were explored with multiple tools. Genetic alterations and their associations with patient prognosis were evaluated using cBioPortal, an online tool for visualization and analysis of multidimensional cancer genomics data . Co-exprp < 0.05. Additionally, associations between IRF family members and the activity of cancer pathways, including TSC/mTOR, RTK, RAS/MAPK, PI3K/AKT, hormone ER, hormone AR, EMT, DNA damage response, cell cycle, and apoptosis pathways, were explored with GSCALite, a web-based platform for gene set cancer analysis [To examine biological functions, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were conducted for IRF family members and functionally related genes using DAVID software (version 6.8) . Biologianalysis .The TIMER (Tumor Immune Estimation Resource) database is an immune infiltrate analysis tool for systematic evaluation of the different immune cells that infiltrate tumor tissue and their clinical significance . In our"}
+{"text": "In the SIDERO-CR-2014\u20132016 surveillance study, European clinical isolates comprising carbapenem-non-susceptible (CarbNS) Enterobacterales and MDR non-fermenters were tested against cefiderocol and comparators.Many carbapenem-resistant (CR) Gram-negative (GN) pathogens exhibit MDR, meaning few therapeutic options are available for CR-GN infections. Cefiderocol, a siderophore cephalosporin, has demonstrated Cefiderocol MICs were determined using iron-depleted CAMHB, and comparators using CAMHB, per recommended CLSI methodology. Carbapenemase gene profiles were determined using PCR.N\u2009=\u2009870) from 23 European countries comprised CarbNS Enterobacterales (n\u2009=\u2009457), MDR Pseudomonas aeruginosa (n\u2009=\u2009177) and MDR Acinetobacter baumannii (n\u2009=\u2009236). The most common carbapenemases were KPC (52%), OXA-48-like (19%), VIM (14%) and NDM (8%) in Enterobacterales, VIM (41%) in P. aeruginosa and OXA-23-like (57%) and OXA-24/40-like (37%) in A. baumannii. Most carbapenemase-producing isolates (65%) co-carried ESBLs. Approximately half of P. aeruginosa isolates were negative for carbapenemases, compared with 10% of Enterobacterales and 3% of A. baumannii. A similar proportion of Enterobacterales were susceptible to cefiderocol  compared with comparator antimicrobial agents, including colistin  and ceftazidime/avibactam . Of P. aeruginosa isolates, 98.3% were susceptible to cefiderocol (100% of VIM producers), similar to colistin (100%). Against A. baumannii, 94.9% had cefiderocol MIC \u22642\u2009mg/L and 93.6% of isolates were susceptible to colistin.Isolates (Cefiderocol demonstrated potent activity against CarbNS and MDR GN bacteria, including non-fermenters and a wide variety of MBL- and serine-\u03b2-lactamase-producing strains. Globally, the incidence of carbapenem-resistant (CR) GN bacteria is increasing,Cefiderocol is a novel siderophore cephalosporin, which was developed for the treatment of MDR GN bacteria, including those resistant to carbapenems. Cefiderocol has recently been approved in Europe for the treatment of infections due to aerobic GN organisms in adults with limited treatment options,,,Escherichia coli and Klebsiella pneumoniae, as well as meropenem-resistant Pseudomonas aeruginosa and Acinetobacter baumannii.The structure of cefiderocol is based around a cephalosporin backbone with the addition of a catechol moiety at the 3-position side chain. The cephalosporin core enables cefiderocol to act like other cephalosporins, binding primarily to PBPs and killing bacterial cells by inhibition of peptidoglycan cell wall biosynthesis. Cefiderocol differs from other cephalosporins in that the catechol moiety chelates ferric (Fe-III) iron, mimicking natural siderophores, allowing cefiderocol to exploit the bacteria\u2019s own active receptor-mediated iron transport system to cross the outer membrane.in vitro conditions are essential in order to mimic the hypoferremic conditions encountered by bacteria in the human body during infection.,,in vivo efficacySusceptibility testing of cefiderocol by broth microdilution requires an iron-depleted medium to promote the natural production of siderophores by bacterial cells. Iron-depleted In the SIDERO-CR surveillance study, CR and MDR clinical isolates of GN bacteria collected from patients between 2014 and 2016 were tested against cefiderocol and comparators using recommended CLSI broth microdilution methodology.JAC-AMR Online).,Full methodology for the SIDERO-CR study and molecular characterization using PCR has been published previously (see n\u2009=\u2009457), defined as having a meropenem MIC of \u22652\u2009mg/L. Isolates of P. aeruginosa (n\u2009=\u2009177) and A. baumannii (n\u2009=\u2009236) were included if they demonstrated an amikacin-resistant (MIC \u226532\u2009mg/L), ciprofloxacin-resistant (MIC \u22654\u2009mg/L) and imipenem-resistant (MIC \u226516\u2009mg/L) MDR phenotype.Included in the SIDERO-CR-2014\u20132016 European test set were carbapenem-non-susceptible (CarbNS) phenotypes of Enterobacterales isolates (A. baumannii (CLSI breakpoint \u22648\u2009mg/L),,P. aeruginosa are considered susceptible to cefiderocol at MIC \u22642\u2009mg/L (resistant >2\u2009mg/L).MICs were determined for cefiderocol, cefepime, ceftazidime/avibactam, ceftolozane/tazobactam, ciprofloxacin, colistin and meropenem by broth microdilution according to CLSI guidelines.in vitro samples were anonymized.Ethics approval was not required as all N\u2009=\u2009870) were from Italy [217 (24.9%)], Greece [128 (14.7%)] and Russia [91 (10.5%)].The majority of SIDERO-CR-2014\u20132016 European isolates (A. baumannii [135/385 (35.1%)] and P. aeruginosa (106/385 [27.5%]). K. pneumoniae (120/385 [31.2%]) was the most common Enterobacterales species in RTIs , followed by intra-abdominal infections , surgical site infections  and bloodstream infections . K. pneumoniae was the most common pathogen in UTIs [59/157 (37.6%)], SSIs [57/89 (64.0%)] and BSIs [47/85 (55.3%)] while A. baumannii was the most common in IAIs [46/125 (36.8%)].Respiratory tract infections (RTIs) were the most common isolate source [385 (44.3%)]; the majority were non-fermenters [241/385 (62.6%)], consisting of Is Table . Urinaryin vitro activity against a variety of CarbNS-GN bacteria in SIDERO-CR-2014\u20132016 European isolates. Overall, 772/870 (88.7%) had a cefiderocol MIC of \u22642\u2009mg/L; 547/634 (86.3%) of Enterobacterales and P. aeruginosa isolates were susceptible to cefiderocol and 224/236 (94.9%) of A. baumannii isolates had a cefiderocol MIC of \u22642\u2009mg/L.Cefiderocol exhibited 90\u2009=\u20094\u2009mg/L) (TableK. pneumoniae isolates (n\u2009=\u2009332), 82.8% were susceptible to cefiderocol, while 71.7% were susceptible to colistin and 88.9% to ceftazidime/avibactam.Of CarbNS Enterobacterales, 81.6% were cefiderocol susceptible (MIC/L) Table. The pro90 was 1\u2009mg/L against MDR P. aeruginosa and 98.3% of isolates were cefiderocol susceptible (MIC \u22642\u2009mg/L). Colistin demonstrated a similar level of activity (100% susceptible) to cefiderocol, while other comparators were active against <35% of isolates.The cefiderocol MICA. baumannii, the cefiderocol MIC90 was 1\u2009mg/L and 94.9% of isolates had a cefiderocol MIC of \u22642\u2009mg/L. Of the comparators, only colistin demonstrated activity (93.6% susceptible). The MIC ranges for cefiderocol were 0.004\u2009\u2212\u20098\u2009mg/L for P. aeruginosa and 0.03\u2009\u2212\u200932\u2009mg/L for K. pneumoniae, while the range against A. baumannii was somewhat wider (0.015 to >64\u2009mg/L).Against MDR P. aeruginosa isolates non-susceptible to cefiderocol (MIC >2\u2009mg/L) was 13.7% (87/634); these isolates consisted mainly of CarbNS K. pneumoniae (n\u2009=\u200957) and Enterobacter cloacae (n\u2009=\u200918) , Italy (n\u2009=\u20092), Denmark (n\u2009=\u20091), Portugal (n\u2009=\u20091) and Turkey (n\u2009=\u20091)].In total, the proportion of Enterobacterales and ) Figure . There wn\u2009=\u2009107; cefiderocol MIC90\u2009=\u20094\u2009mg/L; range: 0.06\u2009\u2212\u200932\u2009mg/L), as did 97.8% of ceftolozane/tazobactam-resistant P. aeruginosa  and all colistin-resistant isolates  , 76.9% were cefiderocol susceptible; all 15 colistin-resistant A. baumannii isolates had cefiderocol MICs of \u22642\u2009mg/L.Notably, 66.4% of ceftazidime/avibactam-resistant Enterobacterales remained susceptible to cefiderocol (L) Figure. Of coliP. aeruginosa isolates, VIM (41.2%) was the most common carbapenemase. OXA-23-like (57.2%) and OXA-24/40-like (37.3%) producers accounted for the majority of A. baumannii isolates. Most isolates produced multiple \u03b2-lactamases, e.g. 34/37 (91.9%) NDM-1 Enterobacterales also harboured ESBLs and OSBLs. A proportion of all isolates, phenotypically non-susceptible to meropenem, were negative for carbapenemase genes, with a greater proportion of P. aeruginosa (49.7%) being carbapenemase negative compared with Enterobacterales (9.8%) and A. baumannii (3.4%).In total, 28 subclasses of carbapenemase were identified, along with 37 ESBL (e.g. CTX-M) and older-spectrum \u03b2-lactamase  subclasses,n\u2009=\u2009157) and Greece (n\u2009=\u200997) provided the most CarbNS Enterobacterales isolates. Most of these isolates produced KPC  or VIM ; however, NDM was less prevalent in Italy (0.6%) than in Greece (5.2%). OXA-48-like carbapenemases ]. NDM-producing P. aeruginosa isolates were only apparent in Serbia, representing 6/10 samples.In countries providing \u22658 isolates, the majority of carbapenemase-producing MDR A. baumannii isolates from countries with \u22658 available isolates established that the majority produced OXA-23-like carbapenemases,Analysis of MDR 90 values were \u22644\u2009mg/L for all carbapenemases across all strains where \u226510 isolates were available for testing  and NDM  MBL producers. Isolates producing OXA-48-like (n\u2009=\u200985), OXA-23-like (n\u2009=\u2009135) and OXA-24/40-like (n\u2009=\u200988) carbapenemases had cefiderocol MIC90 values of \u22644\u2009mg/L.Cefiderocol MICing Table. Across 90 of KPC-producing isolates was 4\u2009mg/L, with 83.6% being cefiderocol susceptible at the EUCAST breakpoint of \u22642\u2009mg/L, but with 98.3% being cefiderocol susceptible at the CLSI breakpoint of \u22644\u2009mg/L. Ceftazidime/avibactam demonstrated potent activity against 97.9% of KPC-producing Enterobacterales by both EUCAST and CLSI breakpoints. Similarly, against OXA-48-like-producing Enterobacterales, only cefiderocol and ceftazidime/avibactam (both 88.2% susceptible) demonstrated efficacy in >75% isolates. Against VIM-producing Enterobacterales, only cefiderocol (79.0% susceptible) and colistin (93.5% susceptible) demonstrated notable activity. The proportion of susceptible NDM-producing Enterobacterales isolates was higher for cefiderocol (51.4%) and colistin (78.4%) versus all other comparators (<3%).The activity of cefiderocol and comparators by carbapenemase is summarized in TableP. aeruginosa, cefiderocol and colistin demonstrated potent activity, with 100% of isolates being susceptible to both agents. Cefiderocol and colistin were also active against non-carbapenemase-producing MDR P. aeruginosa, with 96.6% of isolates being cefiderocol susceptible and 100% being colistin susceptible.Against both VIM- and GES-producing A. baumannii producing OXA-23-like  and OXA-24/40-like  carbapenemases.Cefiderocol and colistin also demonstrated potency against P. aeruginosa and A. baumannii isolates is apparent in FigureThe wide variety of carbapenemases produced by European Enterobacterales, n\u2009=\u200939), followed by NDM (n\u2009=\u200919), VIM (n\u2009=\u200913), OXA-48-like (n\u2009=\u200910), OXA-23-like (n\u2009=\u20096) and OXA-24-like (n\u2009=\u20095). Twelve isolates carried either OSBLs/ESBLs or no known \u03b2-lactamase, with eight isolates harbouring the PER-type \u03b2-lactamase.A range of \u03b2-lactamases were identified across 99 isolates with cefiderocol MIC values of >2\u2009mg/L, including isolates with co-carriage of multiple \u03b2-lactamases. The most common carbapenemase was KPC  and carbapenemase-producing K. pneumoniae (n\u2009=\u2009107) were cefiderocol susceptible.Similar isolate studies involving cefiderocol have used provisional CLSI breakpoints. In a study including 1086 CR isolates from the USA (737 KPC producers), MICs were higher for isolates of Enterobacterales with \u03b2-lactamases compared with those without, but no clear association was found between the type of \u03b2-lactamase and the MIC.A key limitation of the SIDERO-CR study was the geographical spread and number of collection sites within countries. Isolates were selected by a limited number of sites per country based on the MDR/CR phenotype, therefore the relative frequency observed may not reflect the national prevalence. Additionally, the number of sites and isolates per country were not proportionate to population. Consequently, the isolate collection is not necessarily representative, as there may be considerable heterogeneity in the numbers and types of pathogens and mechanisms of resistance provided by different locales. However, from SIDERO-CR-2014\u20132016 isolate characterization, it is apparent that carbapenem resistance across Europe is associated with a diverse range of \u03b2-lactamases. Of the CR isolates included, 83.8% (729/870) produced at least one carbapenemase, with 65.2% (475/729) of carbapenemase-producing isolates also carrying ESBLs and/or OSBLs.K. pneumoniae with genes encoding for OXA-48 and NDM-1 has recently been reported in the Mecklenburg-Western Pomerania state of Germany.K. pneumoniae was investigated across eight Greek hospitals between 2013 and 2016.K. pneumoniae isolates were of a clonal type (ST11) similar to those identified in Bulgaria in 2015\u201316, indicating that the Balkan region is also at risk of increasing prevalence of MBL-producing Enterobacterales.K. pneumoniae infection has led to a shift in the carbapenemase landscape in Greece, with the incidence of MBLs increasing between 2015\u201317 (12.0%) and 2018 (51.1%), mainly due to VIM-producing K. pneumoniae becoming more prevalent.,,P. aeruginosa isolates resistant to ceftazidime/avibactam and 77.4% of all MDR P. aeruginosa isolates resistant to ceftolozane/tazobactam , recent reports have described an increasing proportion of non-KPC MBL-mediated resistance. A Rapid Risk Assessment report from the ECDC described an increase in NDM-containing carbapenemase-producing Enterobacterales in Italy,tam Table.A. baumannii.,P. aeruginosa and A. baumannii, but particularly in K. pneumoniae , mediated, to some extent, by transmissible mcr-1 resistance.K. pneumoniae) and 6.4% of A. baumannii, with 100% of P. aeruginosa isolates being colistin susceptible. These results do not necessarily reflect the incidence of colistin resistance in specific geographical locations or from specific infection sources; for example, the incidence of colistin-resistant A. baumannii was reported to be 47.7% in a set of A. baumannii RTI isolates from Greece, Italy and Spain (n\u2009=\u200965).In vitro assessment of CR-GN isolates from 18 hospitals in Greece demonstrated that \u223c40% of K. pneumoniae and A. baumannii isolates were resistant to colistin; 100% of these isolates had cefiderocol MICs of \u22644\u2009mg/L.Colistin, often considered a treatment of last resort, has a broad spectrum of GN activity and is frequently used for CR- and MDR-GN infections, particularly for MDR P. aeruginosa. Cefiderocol retained activity against the majority of isolates harbouring MBLs, with only 3.0% (3/99) of MBL-positive CarbNS Enterobacterales isolates having MIC values >4\u2009mg/L. The in vitro activity (MIC \u22642\u2009mg/L) of cefiderocol against a range of serine-\u03b2-lactamases varied from 100% in OXA-58-producing isolates to 83.6% in KPC producers. In addition, 91.5% of isolates with no carbapenemase and 97.5% with no \u03b2-lactamase had a cefiderocol MIC \u22642\u2009mg/L, demonstrating potent cefiderocol activity where the mechanism of resistance was not clear.In this study, the proportions of isolates susceptible to cefiderocol are similar to comparators in Enterobacterales and generally similar to colistin but greater than other comparators in P. aeruginosa isolates (98.3% cefiderocol susceptible), yet half carried no carbapenemase. The low prevalence of carbapenemases in SIDERO-CR-2014\u20132016 P. aeruginosa isolates aligns with recently reported data describing the prevalence of XDR P. aeruginosa; of 1445 P. aeruginosa isolates collected from 51 Spanish hospitals, 252 (17.3%) were classified as XDR and only 3.1% carried either carbapenemases or ESBLs.P. aeruginosa isolates in SIDERO-CR-2014\u20132016 is likely to be due to porin- and efflux pump-mediated mechanisms.in vitro activity against strains with alterations in outer membrane porins or overexpressed efflux pumps.,Cefiderocol demonstrated potency against the MDR in vitro activity against a broad range of CarbNS and MDR pathogens. The isolates tested included \u03b2-lactamase-producing strains from all Ambler classes, with cefiderocol demonstrating activity against the key MBLs VIM and NDM, as well as against clinically important serine-\u03b2-lactamases KPC, GES and OXA, and isolates co-carrying ESBLs and OSBLs. Consequently, cefiderocol represents a key addition to the limited armamentarium available for the treatment of infections caused by CR- and MDR-GN organisms and could be a particularly valuable and timely treatment option for when resistance is apparent but the mechanism is unknown.In conclusion, the SIDERO-CR European dataset, coupled with the rise of resistance to existing agents and the shift towards MBL- from serine-\u03b2-lactamase-producers, demonstrates the diverse and dynamic nature of the European carbapenemase landscape. In SIDERO-CR-2014\u20132016, cefiderocol exhibited potent dlaa060_Supplementary_DataClick here for additional data file."}
+{"text": "EGFR) T790M mutation on osimertinib efficacy remains unclear.Seventy-eight patients were studied with EGFR-mutated NSCLC and LM. Case data were collected and EGFR mutation status of circulating cell-free DNA from paired CSF, and plasma of 23 patients with LM was detected using droplet digital PCR. The median overall survival (mOS) was 8.08 months (95% CI: 6.07\u201310.09) in the study. Forty-four osimertinib-treated patients had an improved mOS of 13.15 (95% CI: 5.74\u201320.57) and a median progression-free survival (PFS) of 9.50 months (95% CI: 6.77\u201312.23) when compared with patients treated with first- or second-generation EGFR-TKI (mOS\u2009=\u20093.00 months (95% CI: 1.32\u20134.68) and median PFS\u2009=\u20091.50 months (95% CI: 0.00\u20133.14)). In the osimertinib group, mOS values for CSF with and without T790M mutation were 22.15 months (95% CI: 9.44\u201334.87) and 13.39 months (95% CI: 7.01\u201319.76), respectively, with no statistical differences. Regardless of the CSF T790M mutation status, osimertinib demonstrated significant efficacy against LM associated with NSCLC.Osimertinib has demonstrated promising efficacy against leptomeningeal metastasis (LM) associated with T790M-positive non-small-cell lung cancer (NSCLC). However, the effect of cerebrospinal fluid's (CSF's) epidermal growth factor receptor ( EGFR) is the most important driver gene in NSCLC. EGFR mutations occur in 10\u201320% of Caucasian patients with NSCLC but in 40\u201360% of Asian patients [EGFR mutations [EGFR tyrosine kinase inhibitor (EGFR-TKI) treatments [Leptomeningeal metastasis (LM) is defined as the spread of malignant cells within the leptomeninges and subarachnoid space, resulting in a devastating prognosis with limited treatment options . Non-smautations , 4. ThisEGFR mutation status is a critical prognostic factor in the treatment of NSCLC with LM. A secondary EGFR test is often required to determine the status of T790M drug-resistant mutation. However, it is difficult to obtain a second tumor tissue from most patients. Circulating cell-free DNAs (cfDNAs) are extracellular nucleic acids released by tumor cells that can be useful biomarkers for early diagnosis and prognosis [rognosis . Howeverrognosis , 7.EGFR-TKIs remains dismal [EGFR-TKIs, such as afatinib, could partially penetrate the blood\u2013brain barrier, they exhibit no obvious advantage as a treatment for LM [LM is often associated with an extremely poor prognosis, with a median OS (mOS) of 3\u201310 months after diagnosis . Currents dismal \u201313. AlthEGFR-TKI that selectively inhibits EGFR and EGFR T790M mutations. It is highly effective in both untreated and previously treated patients with EGFR-mutated NSCLC by prolonging OS and progression-free survival (PFS) [EGFR-mutated positive NSCLC [EGFR T790M mutation [Osimertinib is an irreversible third-generation al (PFS) \u201316. Accove NSCLC . In contEGFR T790M mutation status and osimertinib efficacy in patients with NSCLC [EGFR-TKIs and evaluate the factors influencing osimertinib efficacy.Few studies have focused on the relationship between CSF th NSCLC , 18. In EGFR-mutated NSCLC and LM at the People's Hospital of Zhengzhou University, recruited from January 1, 2014, to December 31, 2020. Inclusion criteria were: (1) space-occupying lesions of the lungs detected using computed tomography (CT) or whole-body positron emission tomography/computed tomography (PET/CT) and confirmed as NSCLC using histological biopsy, (2) symptoms and signs of the central nervous system metastases, (3) typical leptomeningeal enhancement on magnetic resonance imaging (MRI), and (4) cytologic identification of malignant cells within the CSF. All patients were scored using the Eastern Cooperative Oncology Group performance status (ECOG PS), CT, or PET/CT. In addition, the patient's histology, metastasis site, imaging, CSF parameters, and molecular profiling were collected. And no patients had complicated organ failure, infection in the central nervous system, and diseases of the autoimmune system or hematological system.This retrospective study involved patients with EGFR-mutated NSCLC and LM, 34 patients were treated with first- or second-generation EGFR-TKIs treatment, and 44 patients  were treated with osimertinib  analysis or next-generation sequencing.EGFR kit . All protocols of analysis were carried out in strict accordance with the manufacturer's instructions.Secondary T790M mutation analysis was performed using one of the following samples: biopsied tumor tissues, plasma, or CSF samples. Approximately 10\u2009mL of whole blood and 10\u2009mL of CSF (EDTA as the anticoagulant) were collected for the purpose of cfDNA extraction. The retrieved CSF was used for analysis within 4\u2009h of the lumbar puncture procedure. Plasma and CSF were centrifuged at 1,600\u2009\u00d7\u2009g and 10,000\u2009\u00d7\u2009g, respectively, for 10\u2009min at room temperature. CfDNA was extracted using the QiAamp circulating nuclear acid kit . Droplet digital PCR (ddPCR) was used to detect cfDNA using the Sysmex OncoBeam EGFR-TKI treatment to tumor progression. The cutoff date was December 31, 2020.OS for LM was defined as the time from LM diagnosis to death. PFS was calculated from the onset of p values were used for survival analysis at 95% confidence intervals (95% CI). Cox regression analysis was performed to estimate hazard ratios (HRs) and 95% CIs for OS. Statistical significance was set at p < 0.05.Statistical analyses were performed using SPSS software . Chi-square univariate analysis was used for continuous variables, and a non-parametric test was used for non-normal distribution variables. Kaplan\u2013Meier estimation and log rank EGFR-mutated NSCLC and LM were included. The median age was 61 years (range: 28\u201378), and 74.4% (58/78) were non-smokers. The ECOG PS scores at the time of LM diagnosis were: <1 in 23 (29.5%), 2\u20133 in 43 (55.1%), and 4 in 12 patients (15.4%). Twenty-five patients with NSCLC also presented with LM at initial diagnosis. The median time from NSCLC to LM diagnosis of the remaining 53 patients was 21 months (95% CI: 12.0\u201324.5). Statistically significant differences were not observed in age, sex, smoking status, histology, initial ECOG PS scores, and EGFR mutations.As shown in EGFR T790M mutation analysis. Of these samples, EGFR T790M mutation was identified in 28 cases.Among the 78 patients, 66 (84.6%) of them underwent secondary EGFR gene of cfDNA in 23 patients simultaneously, and T790M mutation was detected in 8 patients (Supplementary EGFR mutations in plasma and CSF samples were 56.5% (13/23) and 60.9% (14/23), respectively, but without any statistically significant difference (p=0.765).Plasma and CSF were used to detect the EGFR-TKI: 24 (45.3%) of them received gefitinib, 11/53 (20.8%) received erlotinib, 7/53 (13.2%) received icotinib, 5/53 (9.4%) received afatinib and gefitinib, and 6/53 (11.3%) received osimertinib. Moreover, 35/53 (66.0%) patients received cytotoxic chemotherapy, and 11/53 (20.8%) had undergone whole-brain radiation therapy (WBRT) for brain metastasis prior to LM  were treated with osimertinib at the time of LM diagnosis. The median duration of osimertinib treatment was 7.0 months (95% CI: 4.0\u20139.0). Thirty-four patients received first- or second-generation TKI treatment without osimertinib. Eighteen (52.9%) of them received gefitinib; 10 (29.4%) received erlotinib; 3 (8.8%) received icotinib; and 1 (2.9%) received afatinib and gefitinib. Twelve patients (35.7%) received cytotoxic chemotherapy; 4 (12.9%) received WBRT; 2 (6.5%) underwent IT chemotherapy; and 3 (9.7%) had VPS insertion .At the end of follow-up, 58 patients died from the disease, 15 were still alive, and 5 were lost to follow-up (6.4%). The 1-year OS rate in the study was 32.7% (19/73), with a mOS of 8.08 months , with a 1-year OS rate of 12.9%, which was shorter than that of patients treated with osimertinib (HR: 0.58 (95% CI: 0.44\u20130.77) and p \u2264 0.001; Among the 44 patients who received osimertinib treatment, 39 (88.6%) had positive clinical responses. Twenty-eight patients died, and 14 were still alive at the time of follow-up, with a mOS of 13.15 months (95% CI: 5.74\u201320.57). The 1-year OS rate was 53%. Among the patients who were treated with first- or second-generation Compared to the group treated with osimertinib, PFS of the group treated without osimertinib also was significantly shorter (PFS: 1.50 months (95% CI: 0.00\u20133.14) and HR: 0.57 month (95% CI: 0.44\u20130.75); p=0.564). Regardless of T790M status, the survival benefit of the osimertinib treatment group was better than that of the first- or second-generation EGFR-TKI treatment group (mOS\u2009=\u20093.00 months (95% CI: 1.32\u20134.68) and HR: 0.56 (95% CI: 0.41\u20130.78); Furthermore, we further divided the group treated with osimertinib into two subgroups according to T790M mutational status. The mOS for patients with and without T790M mutational status was 15.92 months (95% CI: 7.70\u201324.14) and 9.00 months (95% CI: 5.50\u201312.50), respectively, but without any significant difference (EGFR mutation in CSF cfDNA was detected in 23 patients of the osimertinib treatment group. Seven patients with T790M mutation in CSF had a mOS of 22.15 months (95% CI: 9.44\u201334.87), whereas 16 patients without T790M mutation had a mOS of 13.39 months (95% CI: 7.01\u201319.76). No statistical difference was found according to Kaplan\u2013Meier survival analysis  and 13.39 months (95% CI: 3.59\u201323.18), respectively. However, survival analysis revealed no significant differences between the two groups p=0.416; .p=0.001 and multivariate HR: 3.22 (95% CI: 1.74\u20135.95), p=0.021). Osimertinib treatment was identified as a significant independent favorable prognostic factor in patients with NSCLC and LM (univariate HR: 0.59 (95% CI: 0.45\u20130.78), p=0.000 and multivariate HR: 0.65 (95% CI: 0.49\u20130.87), p=0.004). Univariate analysis revealed a statistically significant correlation between WBRT and prognosis (0.45 (95% CI: 0.23\u20130.96), p=0.038). However, no significant associations were observed with concurrent brain metastases, initial EGFR and EGFR T790M mutations, CSF pressure, protein and glucose levels, and systemic and intrathecal cytotoxic chemotherapy , otherapy .EGFR-TKI drugs have resulted in significant survival benefits and improved quality of life for patients with EGFR mutations. Third-generation EGFR-TKIs, such as osimertinib, have also been found to extend the survival of patients with T790M mutation by more than 10 months [The treatment of lung cancer has gradually evolved from traditional therapies to gene-oriented personalized treatments. The first- and second-generation 0 months .EGFR\u2009+\u2009NSCLC, regardless of T790M mutation that confers drug resistance, with a median LM PFS of 9.50 months and a mOS of 13.15 months. Patients treated with only first- or second-generation EGFR-TKI after LM diagnosis had a median PFS of 1.50 months and a mOS of 3.00 months. Compared to first- or second-generation EGFR-TKIs, osimertinib showed a markedly improved survival benefit.Our study found that osimertinib is a clinically effective standard regimen in patients with LM associated with EGFR-TKIs for LM is limited because of the blood\u2013brain barrier. The first- and second-generation EGFR-TKIs have low CSF penetration, with an average CSF penetration of 1\u20133% for gefitinib and 3\u20136% for erlotinib compared to only 1% for afatinib [As mentioned, the efficacy of first-generation afatinib \u201313. Highafatinib . Althougafatinib .EGFR mutation NSCLC [EGFR-TKI therapy failure [Preclinical studies of osimertinib using mouse models of brain metastasis with 19 gene deletions (PC9) have demonstrated that a dose-dependent tumor reduction can be achieved . The AURon NSCLC . Several failure , 23. SimEGFR T790M screening failure in the BLOOM study might be a missed opportunity in revealing osimertinib activity in the CNS [EGFR-TKI-resistant NSCLC and without the T790M mutation should be further investigated.The excluded patients based on  the CNS . A recen the CNS . SimilarEGFR mutations detected using CSF cfDNA tended to be higher compared to plasma cfDNA [EGFR mutation rate based on CSF samples [in vivo, in vitro, and/or in silico along with other math tools and models, including meta-analysis [Due to the blood\u2013brain barrier, plasma cfDNA may not accurately reflect the actual state of mutations in intracranial tumors, whereas the rapid CSF circulation within the cerebral ventricle and spinal cord cavity indicates that CSF cfDNA may be a reliable biomarker for intracranial tumors. In patients with LM associated with metastasized lung carcinoma, the proportion of ma cfDNA . A simil samples , which i samples . Howeveranalysis , 26, resanalysis , 28, netanalysis , and molanalysis , 31.This study had several limitations. Firstly, this was a retrospective study involving a small number of patients. All the patients included in our study were confirmed using cytological findings, whereas patients diagnosed via clinical examination and imaging were not included, which may have contributed to selection bias. Secondly, the ddPCR method for detecting plasma and CSF cfDNA had low sensitivity, which could have potentially resulted in a lower detection rate. Therefore, more sensitive cfDNA detection assays, such as NGS, will be employed for further verification analysis.EGFR mutation status of circulating cell-free DNA from paired CSF, and plasma of 23 patients with LM was detected using droplet digital PCR. The median overall survival (mOS) was 8.08 months (95% CI: 6.07\u201310.09) in the study. Forty-four osimertinib-treated patients had an improved mOS of 13.15 months (95% CI: 5.74\u201320.57) and a median progression-free survival (PFS) of 9.50 months (95% CI: 6.77\u201312.23) when compared with patients treated with first- or second-generation EGFR-TKI (mOS\u2009=\u20093.00 months (95% CI: 1.32\u20134.68) and median PFS\u2009=\u20091.50 months (95% CI: 0.00\u20133.14)). In the osimertinib group, mOS values for CSF with and without T790M mutation were 22.15 months (95% CI: 9.44\u201334.87) and 13.39 months (95% CI: 7.01\u201319.76), respectively, with no statistical differences. Regardless of the CSF T790M mutation status, osimertinib demonstrated significant efficacy against LM associated with NSCLC.Case data were collected and"}
+{"text": "Kappaphycus alvarezii, Eucheuma denticulatum, Halymenia durvillaei (Rhodophyta), Caulerpa lentillifera, Caulerpa racemosa (Chlorophyta), Dictyota dichotoma and Sargassum polycystum (Ochrophyta). This review aims to highlight the therapeutic potential of North Bornean seaweeds and their nutraceutical profiling. North Bornean seaweeds have demonstrated anti-inflammatory, antioxidant, antimicrobial, anticancer, cardiovascular protective, neuroprotective, renal protective and hepatic protective potentials. The protective roles of the seaweeds might be due to the presence of a wide variety of nutraceuticals, including phthalic anhydride, 3,4-ethylenedioxythiophene, 2-pentylthiophene, furoic acid , eicosapentaenoic acid, palmitoleic acid, fucoxanthin, \u03b2-carotene (E. denticulatum), eucalyptol, oleic acid, dodecanal, pentadecane (H. durvillaei), canthaxanthin, oleic acid, pentadecanoic acid, eicosane (C. lentillifera), pseudoephedrine, palmitic acid, monocaprin (C. racemosa), dictyohydroperoxide, squalene, fucosterol, saringosterol (D. dichotoma), and lutein, neophytadiene, cholest-4-en-3-one and cis-vaccenic acid (S. polycystum). Extensive studies on the seaweed isolates are highly recommended to understand their bioactivity and mechanisms of action, while highlighting their commercialization potential.Malaysia has a long coastline surrounded by various islands, including North Borneo, that provide a suitable environment for the growth of diverse species of seaweeds. Some of the important North Bornean seaweed species are  Marine organisms have been widely used as sources of functional bioactive compounds over the years . Among mKappaphycus alvarezii (Doty) Doty, Eucheuma denticulatum (Burman) Collins et Harvey, Halymenia durvillaei Bory de Saint-Vincent, Caulerpa lentillifera J. Agardh, Caulerpa racemosa (Forssk\u00e5l) J. Agardh, Dictyota dichotoma (Huds.) Lamouroux, and Sargassum polycystum C. Agardh [Malaysia is part of the Coral Triangle, a geographical area in Southeast Asia and the Pacific that includes the oceans close to Indonesia, Malaysia, Philippines, Papua New Guinea, Timor-Leste and Solomon Islands. The temperatures of Malaysia\u2019s coastal waters make it ideal for the development and growth of a wide variety of seaweed species. The North Borneo region of Malaysia is one of the main seaweeds growing areas; it has a suitable environment for cultivating a diverse variety of seaweeds and is the only region of Malaysia where seaweeds are farmed commercially . In Nort. Agardh . Brief dSeaweeds are also known as sea vegetables and have been used for the treatment of various disorders ,14,15,16Inflammation is a recognized defensive mechanism evolved in high-level organisms in response to stressors that disrupt bodily homeostasis. Microbial infections, tissue stress and some traumas are examples of hazards that cause inflammation with common symptoms of fever, redness, swelling and pain ,24. OverK. alvarezii has been reported to have anti-inflammatory potential in asthma-induced rats. The extract changed circulating white blood cell levels, reduced mucin synthesis, and downregulated the expression of TNF-\u03b1, IL-4, nuclear factor kappa beta (NF-\u03baB), epidermal growth factor receptor (EGFR) and matrix metalloproteinase (MMP-9). The consumption of seaweed may be useful for asthma patients. The extract decreases bronchiole smooth muscle thickness for airflow facilitation and decreases asthmatic inflammation, lung eosinophil infiltration, and mucin production [oduction .C. lentillifera polysaccharides has been reported to have an inhibitory impact on lipopolysaccharide (LPS)-induced HT29 colorectal carcinoma cells, lowering the overproduction of TNF-\u03b1 and IL-1\u03b2, SIgA and mucin2 (related proteins), as well as decreasing TNF-\u03b1 and IL-1\u03b2 expression [C. racemosa polysaccharides have been reported to have anti-inflammatory potential and activate the hemoxigenase-1 (HO-1) pathway to sustain the production of hemoxigenase-1 enzyme, crucial for the prevention of inflammation [The anti-inflammatory activity of pression . C. raceammation . D. dichotoma extract at a concentration of 25 ug/mL inhibited the production of NO and PGE2, followed by a reduction in the expression of inducible nitric oxide synthase (iNOS) and COX-2 proteins, and iNOS and COX-2 mRNA in a dose-dependent pattern. COX-2 and iNOS are implicated in a variety of pathological processes, including inflammation. The solvent fractions of D. dichotoma extracts also decrease the mRNA expression of other cytokines including TNF-\u03b1, IL-1, and IL-6 in the murine macrophage cell line [In murine macrophage RAW 264.7 cells, the dichloromethane fraction of ell line .S. polycystum has been reported using a mouse model with the paw edema method, where the mouse paw was inflamed and the hexane fraction of seaweed extract at a concentration of 70 mg/kg b.w. was applied. The anti-inflammatory effect was measured by the decreased percentage of edema size. The hexane fractions of S. polycystum extract significantly reduced the edema size compared to untreated mice [Anti-inflammatory and analgesic activity from brown algae ted mice .Antioxidant phytochemical compounds can scavenge the reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the human body, and slow down or prevent the onset of oxidative stress-related diseases including cardiovascular diseases (CVDs), neurological diseases , Parkinson\u2019s disease, multiple sclerosis, amyotrophic lateral sclerosis (ALS), and depression), cancer , respiratory disease (chronic obstructive pulmonary disease (COPD) and asthma), rheumatoid arthritis, delayed sexual maturation, and kidney and liver diseases ,31,32,33K. alvarezii 1.63 and 225.00 TEAC and FRAP mM.mg/dry extract and 22.50 TPC mg PGE/g dry extract, E. denticulatum 1.54 and 153.97 TEAC and FRAP mM.mg/dry extract and 15.82 TP mg PGE/g dry extract, H. durvillaei 1.67 and 182.29 TEAC and FRAP mM.mg/dry extract and 18.90 TP mg PGE/g dry extract, C. lentillifera 2.16 and 362.11 TEAC and FRAP mM.mg/dry extract and 42.85 TP mg PGE/g dry extract, C. racemosa 2.01 and 355.36 TEAC and FRAP mM.mg/dry extract and 40.36 TP mg PGE/g dry extract, D. dichotoma 1.66 and 268.86 TEAC and FRAP mM.mg/dry extract and 35.23 TP mg PGE/g dry extract, S. polycystum 1.86 and 366.69 TEAC and FRAP mM.mg/dry extract and 45.16 TP mg PGE/g dry extract compared to butylated hydroxytoluene (standard) (3.84 and 615.71 TEAC and FRAP mM.mg/dry extract) [C. lentillifera has high antioxidation activities followed by C. racemosa, S. polycystum, H. durvillaei, D. dichotoma, K. alvarezii and E. denticulatum; while in terms of TP, S. polycystum has indicated high values followed by C. lentillifera, C. racemosa, D. dichotoma, K. alvarezii, H. durvillaei and E. denticulatum [Marine seaweeds from North Borneo are a good source of antioxidants ,11,34,35Pathogenic microbes including bacteria, fungi, viruses and parasites are responsible for the development of various diseases that arise in the community . SeaweedK. alvarezii against Staphylococcus aureus (S. aureus)  has been examined. Administration of the extract at a concentration of 200 mg/mL resulted in an inhibition zone of 10.03 mm [K. alvarezii was also effective against S. aureus, Staphylococcus epidermidis (S. epidermidis) , Pseudomonas aeruginosa (P. aeruginosa)  and Bacillussubtilis (B. subtilis)  bacterial strain at a concentration of 30\u201380% (w/v) with an inhibition zone of 11.93\u201314.85 mm, while ethyl acetate fractions of the ethanol extract of K. alvarezii at a concentration of 50% (w/v) inhibited the growth of S. aureus, S. epidermidis, P. aeruginosa and B. subtilis with inhibition zones of 7.14, 19.70, 0.73 and 18.30 mm, respectively. No antifungal activity of K. alvarezii extract and fractions have been published against Candida albicans  (C.P. Robin) Berkhout and Aspergillus niger (A. niger) (Tiegh) [K. alvarezii extract have been reported against Lagenidium spp and Haliphthoros fungal strains [Lagenidium thermophilum (L. thermophilum)  and Haliphthoros sabahensis (H. sabahensis)  are pathogenic to the eggs and larval stages of Scyllaserrata  and Scyllatranquebarica  (mangrove crabs) [K. alvarezii inhibited the hyphal growths of L. thermophilum IPMB 1401  and H. sabahensis IPMB 1402 [K. alvarezii  exhibited strong anti-influenza activity against a wide spectrum of influenza virus strains, including the newly evolving swine-origin H1N1-2009 influenza strain. The mechanism involved the direct binding of ECA-2 to the viral envelope protein hemagglutinin (HA) and inhibited influenza virus propagation [The antibacterial potential of the aqueous extraction of 10.03 mm . The eth (Tiegh) . On the e crabs) ,40. The pagation .E. denticulatum inhibited the growth of S. aureus with inhibition zones of 6.0\u201316.3 mm. Furthermore, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values for E. denticulatum extract against S. aureus were 10% and 15%, respectively. At 10%, minimum bacterial growth was observed, the number of bacteria greatly decreased from 3.0 \u00d7 107 to 1.5 \u00d7 102 CFU/plate, and turbidity levels also decreased; while at 15% of the extract, no bacterial growth was noticed [E. denticulatum extract has been reported against Aspergillus flavus (A. flavus) (Link) [E. denticulatum have been tested for in vitro antiviral activity against human herpes virus type 1 (HHV-1). Carrageenans indicated an antiviral impact by the inhibition of virus attachment and interference in a subsequent stage of the virus replicative cycle. HHV-1 viral DNA synthesis was reduced by 3 folds in cultures treated with sulphated polydigalactosides from E. denticulatum (0.75 mg/mL) [The ethanol extract of  noticed . No anti) (Link) . Sulphat5 mg/mL) .H. durvillaei has been reported to have antimicrobial effects. The presence of antimicrobial activities of H. durvillaei extract against pathogenic bacteria was determined using the disc diffusion method. The solvent extract of H. durvillaei inhibited the growth of P. aeruginosa (11.89 mm), S. aureus (12.22 mm), and Streptococcus pyogenes (S. pyogenes) Rosenbach, 1884 (10.67 mm), respectively. However, no fungicidal activity of the solvent extract of H. durvillaei has been reported against C. albicans [The extracts of albicans .C. lentillifera were tested against Methicillin-resistant S. aureus (MRSA) and neuropathogenic Escherichia coli K1 (E. coli K1). Moderate antibacterial activity of 62.17% against MRSA and poor antibacterial impact against E. coli K1 of 12.42% were demonstrated by C. lentillifera extract at a concentration of 250 \u03bcg/mL [C.lentillifera (0\u2013128 \u03bcg/mL) was recorded against the shrimp pathogenic bacteria Vibrio vulnificus  Farmer, 1980, V. alginolyticus  Sakazaki, 1968, V. parahaemolyticus  Sakazaki et al., 1963 or V. harveyi, , as compared with positive  and negative controls [C. lentillifera extract was tested against White Spot Syndrome Virus (WSSV)  [C. lentillifera extract yielded very good outcomes. Shrimps injected with WSSV and C. lentillifera (1\u201310 mg/mL) preincubated solutions exhibited significantly lower mortality of 0.0\u20136.7%, compared with the positive control (100%) (only WSSV-injected). This inhibitory effect was further confirmed by the reduction in viral loads of WSSV, and C. lentillifera (1\u201310 mg/mL) expressed significantly lower viral loads  than the positive control  [C. lentillifera against L. thermophilum and H. sabahensis have been reported as well. An ethanol extract of C. lentillifera inhibited hyphal growths of L. thermophilum IPMB 1401, L. thermophilum IPMB 1601 and H. sabahensis IPMB 1603 [The chloroform extracts of 50 \u03bcg/mL . Howevercontrols . The ant genome) ,47. WSSV genome) . The admtal DNA) . FungiciPMB 1603 .C. racemosa demonstrated antibacterial activity against MRSA and E. coli K1. The extract of C. racemosa, at a concentration of 250 \u03bcg/mL, displayed a high antibacterial effect of 97.7% against MRSA, but a weak effect of 19.90% against E. coli K1. A methanol extract of C. racemosa (250 \u03bcg/mL) also showed antibacterial activity of 61.54% and 42.91% against MRSA and E. coli K1 [C. racemosa has been reported to show antifungal activity against A. flavus. An ethanol extract of the seaweed demonstrated the strongest inhibitory power with a 30 mm diameter inhibition zone against A. flavus [C. racemosa was demonstrated against the Chikungunya virus (CHIKV) [Aedes aegypti and Aedes albopictus mosquitoes, which cause high fever, joint pain, back pain, vomiting, headache, kidney, liver, heart disease, etc. [C. racemosa was determined based on inhibition of the cytopathic effect caused by CHIKV on African monkey kidney epithelial (Vero) cells. Chloroform, ethyl acetate, ethanol, and methanol extracts (5 to 640 \u03bcg/mL) of C. racemosa showed a significant inhibition effect [A chloroform extract of  coli K1 , respect. flavus . The ant (CHIKV) . The virn effect .D. dichotoma at a concentration of 1.5 mg/disc were investigated for in vitro antibacterial and antifungal activities. The results indicated that the methanol extract inhibited the growth of B. subtilis (6.5 mm) and S. aureus (7.5 mm). The dichloromethane extract inhibited the growth of B. subtilis (7.0 mm), Enterobacter aerogenes (E. aerogenes)  (6.5 mm), E. coli (6.5 mm), Proteus vulgaris (P. vulgaris) (11.0 mm) and Salmonella typhimurium (S. typhimurium)  (7.0 mm), whereas the hexane extract inhibited the growth of B. subtilis (9.0 mm) and S. aureus (7.5 mm) only [D. dichotoma has been shown against Salmonella typhi (S. typhi), Klebsiella pneumoniae (K. pneumoniae)  Trevisan, 1887 and Shigella boydii (S. boydii)  [D. dichotoma has been reported against Mucor sp. and A. flavus [D. dichotoma (1.5 mg/disc) has been observed against C. albicans [D. dichotoma extract has been tested against herpes simplex virus (HSV) and coxsackievirus B3 (CVB3) [Methanol, dichloromethane and hexane extracts of mm) only . The antg, 1949) . The ant. flavus , while nalbicans . The ant3 (CVB3) . HSV bel3 (CVB3) ,55. The 3 (CVB3) .S. polycystum was tested against human pathogenic bacteria. The methanol extract of the seaweed resulted in the inhibition of P. aeruginosa (15 mm), K. pneumoniae (16 mm), E. coli (19 mm), and S. aureus (20 mm) [S. polycystum indicated no inhibition against B. subtilis or S. enteritidis. Similarly, no antifungal activity has been observed against A. niger [A solvent extract of  (20 mm) . HoweverA. niger .Cancer is one of the main causes of mortality in the world, and many research facilities are now focusing on the development of new anticancer medicines that could improve chemotherapy treatment and reduce mortality rates . The proK. alvarezii has been reported with anti-breast and anti-colorectal cancer potential. The anticancer activities were expressed with inhibitory concentration (IC50) value (\u00b5g/mL). An IC50 value of less than 100 is considered to indicate an active compound with anticancer properties. An ethanolic extract of K. alvarezii exhibited anticancer activity against human breast adenocarcinoma cell line MCF-7 with an IC50 of 75.7 \u00b5g/mL, while ethyl acetate and hexane extracts showed anti-colorectal cancer activity against human colorectal carcinoma cell line HCT-116 with IC50 values of 21.4 and 43.0 \u00b5g/mL, respectively [A solvent extract of ectively .E. denticulatum against Ehrlich carcinoma and Meth-A fibrosarcoma has been reported. Oral administration of the extract (1600 mg/kg b.w.) for 28 days resulted in the inhibition of Ehrlich carcinoma by 25% in tumor-implanted mice. Similarly, intraperitoneal administration of E. denticulatum extract (50 mg/kg b.w.) for 7 days resulted in the inhibition of Meth-A fibrosarcoma by 17% [H. durvelaei was investigated against four cancer cell lines . The results indicated that administration of H. durvelaei extracts reduced the growth of AGS and HT-29 cell lines by 27.17% and 1.47%, respectively [The antitumor activity of a by 17% . The antectively .C. lentillifera have been reported to show antitumor properties against human breast adenocarcinoma cell line MCF-7. Exposure to C. lentillifera oligosaccharides inhibited the growth of MCF-7 cells in a dose-dependent manner and induced apoptosis (triggered chromatin condensation and poly ADP-ribose polymerase degradation) [Oligosaccharides  obtained from adation) .C. racemosa have been reported to show antitumor activity in tumor-inoculated mice (H22 tumor). The results indicated that administration of C. racemosa polysaccharide fractions of different doses could significantly inhibit the H22 tumor. After 14 days of transplantation, the weight of tumors in mice without polysaccharide treatment increased to 1.02 g while the tumor weight in mice exposed to the polysaccharide at a dose of 100 mg/kg b.w./day by routine oral passage decreased to 0.47 g, and the tumor inhibition rate reached 53.9% [Polysaccharide fractions (coded as CRP) obtained from ed 53.9% .D. dichotoma were tested against seven different cancer cell lines including HCT-116, MCF-7, HepG2, A-549, PC-3, HeLa, and CACO in 96-well plates. The extract and fractions were applied at concentrations ranging from 0.86 to 100 \u03bcg/mL. The results demonstrated that D. dichotoma extract and fractions displayed significant anticancer effects against several cancer cell lines in a dose-dependent manner but were generally more selective against MCF-7 and PC-3 cell lines. The chloroform fraction was the most effective in MCF-7, PC3, and CACO cells  followed by the petroleum ether fraction against MCF-7 and PC-3  and the ethyl acetate fraction against HepG2 and CACO  [A methanol extract and fractions  of ctively) .S. polycystum at a concentration ranging from 25\u2013150 \u03bcg/mL has been reported against human breast adenocarcinoma cell line MCF-7 via cell viability assay. Treatment with the sulphated polysaccharides indicated the highest percentage of inhibition (90.4%) against the MCF-7 cell line at 150 \u03bcg/mL with an estimated IC50 of 50 \u03bcg/mL [S. polycystum polysaccharide induced apoptosis in colorectal cancer cell lines (HCT-15 cell) [The anticancer activity of sulphated polysaccharides (fucoidan) from 50 \u03bcg/mL . In anot15 cell) .The hypocholesterolemic effects of various marine algae and algal polysaccharides have been reported. The administration of extract significantly reduced serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglycerides (TG) ,20.K. alvarezii to the HCF diet dramatically decreased body weight (29.1%), LDL-C (49.3%), plasma TC (11.4%), TG (36.1%), plasma MDA level (10.7%), GSH-Px (13.49%), SOD (9.4%) and CAT (24.48%), and significantly increased HDL-C levels (55%), compared to rats fed the HCF diet only [After 16 weeks on high-cholesterol/high-fat (HCF) diets, male Sprague-Dawley rats weighing 260\u2013300 g had significantly higher body weight (b.w.), lipid peroxidation , end-product of lipid peroxidation), plasma low-density lipoprotein cholesterol (LDL-C), plasma total cholesterol (TC), plasma triglycerides (TG), erythrocyte glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) levels. The addition of 5% iet only . E. denticulatum played a vital role in the reduction of fat absorption by the body via inhibition of pancreatic lipase [E. denticulatum extract at a concentration of 3.8 mg/mL showed pancreatic lipase activity inhibition with an 83% reduction [c lipase . Inhibitc lipase ,66. An Eeduction . C. lentillifera was reported to show anti-obesity and cardiovascular protection activity. Supplementation with 5% C. lentillifera extract for 16 weeks in HCF-diet rats reduced body weight by 39.5%, significantly increased HDL-C levels by 48.7%, reduced plasma TC by 18.4%, LDL-C by 34.6% and TG by 33.7%, and lowered plasma MDA level by 9%, GSH-Px by 31.8% and CAT by 3.14%, compared to the corresponding levels in high-cholesterol-diet rats [Treatment of HCF diet rats with iet rats . S. polycystum significantly reduced body weight gain, plasma antioxidant enzymes and plasma lipid peroxidation to levels closer to those of healthy rats. Supplementation with 5% S. polycystum in rats fed a high-fat diet reduced body weight by 42.6%, significantly increased HDL-C levels by 16.2%, reduced plasma TC by 11.4%, LDL-C by 22% and TG by 7.69%, and decreased the plasma MDA level by 6.8%, GSH-Px by 43.4% and CAT by 15.7%, as compared to the corresponding levels in hypercholesterolemia and hypertriglyceridemia rats [The exposure of induced-hypercholesterolemia and -hypertriglyceridemia rats to mia rats .Millions of people die each year as a result of hepatic diseases across the world ,68. The K. alvarezii ethanolic extract administered for 25 days against lead acetate-induced hepatic injury in mice has been investigated. The extract at a concentration of 800 mg/kg b.w. reduced AST, ALT and ALP levels by 15.79%, 18.52% and 16.11%, respectively, compared to lead acetate-treated mice . Mice administered with ethanol extract of K. alvarezii (800 mg/kg b.w.) also demonstrated a significant (p < 0.05) elevation in SOD and GPx levels by 45.94% and 18.78%, respectively, and a significant (p < 0.05) reduction in MDA level by 22.83%, compared with lead acetate-treated mice. Histological observations of mouse hepatic tissues treated with K. alvarezii ethanolic extract indicated improved hepatic cell structure, blood congestion, and fatty degeneration compared to lead acetate-treated mice [The hepatoprotective activity of ted mice .C. lentillifera demonstrated hepatoprotection against acetaminophen  induced hepatic damage in juvenile zebrafish (aged 1\u20133 months). The administration of APAP to the control group at a concentration of 10 \u03bcM caused fish mortality; while the introduction of the methanol extract of C. lentillifera at concentrations of 10, 20 and 30 \u03bcg/l to tanks holding 10 \u03bcM APAP-treated groups reduced fish mortality. Histological observation by hematoxylin and eosin staining of zebrafish hepatic tissues exposed to 10 \u03bcM APAP and concurrently administered C. lentillifera extract indicated a reduction in hepatic necrosis, hepatocyte swelling, hepatocyte vacuolization and leukocyte infiltration in a dose-dependent manner, as compared to the control group treated solely with 10 \u03bcM APAP [A methanol extract of  \u03bcM APAP .C. racemosa at a concentration of 200 mg/kg b.w. was administered for 30 days on a daily basis to 40% carbon tetrachloride (CCl4) induced hepatic fibrosis rats . Intoxicated rats treated with water extracts of C. racemosa showed significant (p < 0.05) decreases in their high levels of AST (46.7%), ALT (82.2%), ALP (41.3%), LDH (25.8%) and total bilirubin (69.6%), as compared to CCl4 treated control rats [An aqueous extract of rol rats . S. polycystum was examined in acetaminophen  induced hepatic oxidative injured rats. The oral administration of S. polycystum extract in intoxicated rats at a concentration of 200 mg/kg b.w./day for 15 days reduced elevated levels of ALT (27.64%), AST (56.43%), LDH (43.38%), ALP (72.53%) and MDA (31.50%), compared to the levels in an APAP-administered control group [The protective effect of a solvent extract of ol group .Neuroprotection is a strategy for halting the progression of neurodegeneration . NeurodeK. alvarezii might be beneficial as a food supplement or medication for those who are prone to neurological disorders. In primary cultures of hippocampal neurons, the effects of K. alvarezii extracts on the development and complexity of neuronal cytoarchitecture were reported. A solvent extract of K. alvarezii with an optimal concentration of 1 \u03bcg/mL was added to primary cultures of fetal rat hippocampal neurons. The extract significantly elevated axonal length, number of secondary axonal collateral branches, length of primary dendrites and number of secondary dendritic branches by 58%, 8 folds, 68% and 2.6 folds, respectively, as compared to control [It was reported that an extract of  control .D. dichotoma has been reported. A methanol extract of D. dichotoma at a concentration of 1.3\u20136.5 mg/mL showed significant (p < 0.05) inhibition of cholinesterase enzyme (54.42%), compared to standard donepezil (cholinesterase inhibitor) (57.57%) at a concentration of 0.40\u20134.15 mg/mL [Alzheimer\u2019s disease is a common neurologic disorder, responsible for brain shrinkage and cell death. It is the most prevalent type of dementia and one of the top four causes of mortality in developed countries . So far,15 mg/mL . C. racemosa and S. polycystum at various concentrations (0.0125\u20130.2 mg/mL) have been determined. Solvent extracts of C. racemosa and S. polycystum indicated anti-acetylcholinesterase activities with IC50 values ranging from 0.086\u20130.115 mg/mL, while C. racemosa extracts indicated anti-butyrylcholinesterase activity with an IC50 value of 0.156 mg/mL [The acetylcholinesterase and butyrylcholinesterase inhibitory activities of 56 mg/mL .A summary regarding the protective nature of the above-mentioned North Bornean seaweeds is shown in Seaweed nutraceutical bioactive compounds have great potential in biomedical and pharmaceutical applications ,90,91,92The information was retrieved from multiple internet databases such as ScienceDirect, PubMed, Wiley, ACS publications, etc., and registers including theses and proceedings. Records were searched with keywords related to seaweed, North Borneo, distribution, taxonomy, bioactivity, secondary metabolites, and diseases. Around 250 records approximately from the year 2000 to 2021 were retrieved and screened. Among these, about 100 records were excluded due to being out of the scope of the review. Eventually, a total of 149 records were adopted for the current review paper, and data from organizations such as the World Health Organization were included as well.K. alvarezii), eicosapentaenoic acid, palmitoleic acid, fucoxanthin, \u03b2-carotene (E. denticulatum), eucalyptol, oleic acid, dodecanal, pentadecane, (H. durvillaei), canthaxanthin, pentadecanoic acid, eicosane (C. lentillifera), pseudoephedrine, palmitic acid, monocaprin (C. racemosa), dictyohydroperoxide, squalene, fucosterol, saringosterol (D. dichotoma) and lutein, neophytadiene, cholest-4-en-3-one, and cis-vaccenic acid (S. polycystum). In the current review, the protective effects of North Bornean seaweeds in terms of anti-inflammatory, antioxidant, antimicrobial, anticancer, anti-obesity and cardiovascular protection, neuroprotection, and renal protection as well as hepatic protection were described and followed by nutraceutical profiling. The protective roles of the seaweeds might be due to the presence of a wide variety of nutraceuticals including phthalic anhydride, 3,4-ethylenedioxythiophene, 2-pentylthiophene, furoic acid (K. alvarezii and E. denticulatum are widely cultivated, developed as a functional food source, and used for carrageenans production. Locally, C. lentillifera and C. racemosa are also consumed as a nutrition source. For future perspectives, studies on functional food development and cultivation techniques are highly recommended. Furthermore, extensive studies on the seaweed isolates are needed to understand their bioactivity and mechanisms of action, while highlighting their commercialization potential.Despite their excellent pharmacological characteristics, only"}
+{"text": "Increasing evidence emphasizes the implications of dysregulated apoptosis and autophagy cellular processes in coronary artery disease (CAD). Herein, we aimed to explore apoptosis- and autophagy-related long noncoding RNAs (lncRNAs) in peripheral blood of CAD patients. p value < 0.05, differentially expressed apoptosis- and autophagy-related mRNAs were screened between CAD and healthy blood samples. Also, differentially expressed lncRNAs were identified for CAD. Using the psych package, apoptosis- and autophagy-related lncRNAs were defined with Spearson's correlation analysis. Receiver operating characteristic (ROC) curves were conducted for the assessment of the diagnosed efficacy of these apoptosis- and autophagy-related lncRNAs.  The mRNA and lncRNA expression profiles were retrieved from the Gene Expression Omnibus (GEO) database. With \u2223fold\u2009change | >1.5 and adjusted  Our results showed that 24 apoptosis- and autophagy-related mRNAs were abnormally expressed in CAD than normal controls. 12 circulating upregulated and 1 downregulated apoptosis- and autophagy-related lncRNAs were identified for CAD. The ROCs confirmed that AC004485.3 (AUC = 0.899), AC004920.3 (AUC = 0.93), AJ006998.2 (AUC = 0.776), H19 (AUC = 0.943), RP5-902P8.10 (AUC = 0.956), RP5-1114G22.2 (AUC = 0.883), RP11-247A12.1 (AUC = 0.885), RP11-288L9.4 (AUC = 0.928), RP11-344B5.2 (AUC = 0.858), RP11-452C8.1 (AUC = 0.929), RP11-565A3.1 (AUC = 0.893), and XXbac-B33L19.4 (AUC = 0.932) exhibited good performance in differentiating CAD from healthy controls.  Collectively, our findings proposed that circulating apoptosis- and autophagy-related lncRNAs could become underlying diagnostic markers for CAD in clinical practice. Coronary artery disease (CAD), as a commonly diagnosed heart disease, contributes to the dominant cause of cardiovascular-related deaths . This diApoptosis and autophagy, as two types of programmed cellular deaths, are both involved in the development of CAD . Undue ahttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi) according to the following criteria: organism\u2014Homo sapiens; experiment type\u2014noncoding RNA profiling by array; and disease\u2014CAD. As a result, two datasets including GSE113079 and GSE69587 datasets were obtained for this study. The GSE113079 dataset included 93 CAD and 48 healthy blood samples based on the GPL20115 platform [The mRNA and lncRNA expression profiles of CAD patients and healthy controls were searched from the Gene Expression Omnibus  | >1.5 and adjusted heatmaps .https://www.kegg.jp/) [Genes in autophagy (entry: map04140) and apoptosis (entry: map04210) were obtained from the Kyoto Encyclopedia of Genes and Genomes database  . They wehttp://string-db.org/) [Physical or functional interactions between specified proteins were analyzed via the STRING online tool (db.org/) . Requiredb.org/) . Connectdb.org/) .p value < 0.05 with at least 50% of differentially expressed autophagy- and apoptosis-related mRNAs were considered as differentially expressed autophagy- and apoptosis-related lncRNAs.Spearson's correlation analysis between differentially expressed lncRNAs and differentially expressed autophagy- and apoptosis-related mRNAs was presented via the psych package in R. lncRNAs with correlation The expression of differentially expressed autophagy- and apoptosis-related lncRNAs was externally verified in blood samples from 5 CAD patients and 5 healthy controls in the GSE169256 dataset. Moreover, associations between their expression and clinical features (age) were analyzed by Spearson's correlation tests. Their expression was also compared between male and female patients.Based on the expression profiles of the differentially expressed autophagy- and apoptosis-related lncRNAs, relative operating characteristic curves (ROCs) were conducted via the pROC package in R in the GSE113079 dataset .To explore CAD-related mRNAs, we screened abnormally expressed mRNAs between 93 CAD and 48 healthy blood samples in the GSE113079 dataset. Firstly, we normalized the microarray data via the limma package Figures . 988 up-To find autophagy- and apoptosis-related mRNAs in CAD, we overlapped the abnormally expressed mRNAs and autophagy- and apoptosis-related mRNAs. As a result, 24 mRNAs were identified for CAD , as follCirculating lncRNAs have been considered as diagnosed biomarkers for CAD . Herein,Since the GSE69587 dataset has been standardized, this study no longer standardized the dataset. In total, 430 circulating lncRNAs were upregulated and 305 circulating lncRNAs were downregulated in CAD compared to healthy samples Figures . The topp < 0.05; p < 0.05). Meanwhile, JUN and ITPR3 had positive correlations with downregulated LOC338758 (both p < 0.05). Thus, these lncRNAs could be distinctly related to autophagy and apoptosis in CAD.We analyzed the correlation between 13 abnormally expressed circulating lncRNAs and autophagy- and apoptosis-related mRNAs. Herein, we found that PRKACA, PIK3R2, and NGF were positively related to the 12 upregulated lncRNAs , AJ006998.2 , H19 , RP11-247A12.1 , RP11-288L9.4 , RP11-344B5.2 , RP11-452C8.1 , RP11-565A3.1 , RP5-1114G22.2 , RP5-902P8.10 , and XXbac-B33L19.4  are as follows: AC004485.3 , and XXbac-B33L19.4 (AUC = 0.932) exhibited good performance to differentiate CAD from healthy controls. The above findings concerning circulating lncRNAs might possess effective diagnostic value on CAD, thereby reducing mortality. Among them, circulating H19 is correlated to risk of CAD among a Chinese cohort [On account of the shortcomings of current diagnostic markers on CAD, circulating lncRNAs appear to have attracted close attention. After verification, our data demonstrated that AC004485.3 (AUC = 0.899), AC004920.3 (AUC = 0.93), AJ006998.2 (AUC = 0.776), H19 (AUC = 0.943), RP5-902P8.10 (AUC = 0.956), RP5-1114G22.2 (AUC = 0.883), RP11-247A12.1 (AUC = 0.885), RP11-288L9.4 (AUC = 0.928), RP11-344B5.2 (AUC = 0.858), RP11-452C8.1 (AUC = 0.929), RP11-565A3.1  were distinctly upregulated in CAD compared to healthy controls. More importantly, they had good performance in distinguishing CAD from healthy individuals. Thus, these circulating lncRNAs could be promising diagnostic markers for CAD."}
+{"text": "Flavobacterium strains isolated from a remote Antarctic island, James Ross Island, were studied using a polyphasic taxonomic approach to determine their taxonomic position. Phylogenetic analyses based on the 16S rRNA gene and 92 core genes clearly showed that these strains formed two distinct phylogenetic clusters comprising three and five strains, with average nucleotide identities significantly below 90% between both proposed species as well as between their closest phylogenetic relatives. Phenotyping revealed a unique pattern of biochemical and physiological characteristics enabling differentiation from the closest phylogenetically related Flavobacterium spp. Chemotaxonomic analyses showed that type strains P4023T and P7388T were characterized by the major polyamine sym-homospermidine and a quinone system containing predominantly menaquinone MK-6. In the polar lipid profile phosphatidylethanolamine, an ornithine lipid and two unidentified lipids lacking a functional group were detected as major lipids. These characteristics along with fatty acid profiles confirmed that these species belong to the genus Flavobacterium. Thorough genomic analysis revealed the presence of numerous cold-inducible or cold-adaptation associated genes, such as cold-shock proteins, proteorhodopsin, carotenoid biosynthetic genes or oxidative-stress response genes. Genomes of type strains surprisingly harbored multiple prophages, with many of them predicted to be active. Genome-mining identified biosynthetic gene clusters in type strain genomes with a majority not matching any known clusters which supports further exploratory research possibilities involving these psychrotrophic bacteria. Antibiotic susceptibility testing revealed a pattern of multidrug-resistant phenotypes that were correlated with in silico antibiotic resistance prediction. Interestingly, while typical resistance finder tools failed to detect genes responsible for antibiotic resistance, genomic prediction confirmed a multidrug-resistant profile and suggested even broader resistance than tested. Results of this study confirmed and thoroughly characterized two novel psychrotrophic Flavobacterium species, for which the names Flavobacterium flabelliforme sp. nov. and Flavobacterium geliluteum sp. nov. are proposed.Despite unfavorable Antarctic conditions, such as cold temperatures, freeze-thaw cycles, high ultraviolet radiation, dryness and lack of nutrients, microorganisms were able to adapt and surprisingly thrive in this environment. In this study, eight cold-adapted  For a long time, Antarctica was considered an inhospitable environment with low biodiversity. Although it is a part of Earth\u2019s cryosphere which covers about 20% of the Earth\u2019s surface , it has Bacteroidetes   . Its mem05.2021) . In addic losses .Flavobacterium spp. in Antarctica is supported by their adaptation and coping mechanisms required for survival in harsh conditions including low temperatures, lack of nutrients or intense UV exposure, and freeze-thaw cycles that exert a strong evolutionary pressure on microbial cells . Microbiological sampling is conducted yearly on the James Ross Island (near the north-eastern extremity of the Antarctic Peninsula), Antarctica. Sampling is performed during the summer period when the majority of the island becomes an ice-free area. Antarctic summer season allows sampling of upper soil layers, permafrost, lakes and proglacial streams, which are all areas of intense microbial activities  sampled during the years 2010\u20132019  for parallel testing and comparison purposes. All strains were routinely cultivated on R2A agar plates at 20\u00b0C for 48 hrs.The present study describes a taxonomic investigation of eight  strains  isolatedtivities . Flavoba010\u20132019 . Water s3. The 16S rRNA sequences were submitted to the EzBioCloud server  software (v7.0) (Genomic DNA was extracted and purified using High Pure PCR Template Preparation Kit (Roche Diagnostics) according to the manufacturer\u2019s recommendations. The 16S rRNA genes were amplified and sequenced using universal bacterial primers pA (5\u2032-AGAGTTTGATCCTGGCTCAG-3\u2032) and pH (5\u2032-AAGGAGGTGATCCAGCCGCA-3\u2032) . PCR prod server  for inite (v7.0) . The maxde novo genome assembly. The MismatchCorrector was run to reduce short indels and the number of mismatches. The read coverage cut-off value was automatically computed by the software resulting in contigs and scaffolds. Quality assessment was performed by the QUAST tool  with 7 min of enzymatic fragmentation and final fragments length within the range of 550\u2013650 bp. Sequencing was performed on Illumina MiSeq platform with MiSeq Reagent Kit v2 (500-cycles) . The St. Petersburg genome assembler (SPAdes v3.11.1)  was usedAST tool  and BowtAST tool .Flavobacterium spp. The WGS data of type strains P4023T and P7388T were submitted to the Type Strain Genome server (TYGS)  values were calculated using the ANI calculator on the Kostas Laboratory website5 using reciprocal best hits (two-way ANI) (6 (The whole genome sequence (WGS) data were analyzed for further confirmation of the taxonomic status of the analyzed strains and for comparison with genomes of related r (TYGS)  and to tr (TYGS) . To furtpipeline  with defway ANI) . Proteinway ANI)  and subj ANI)  8 with CARD v3.1.1 database   and Operdatabase , NCBI Badatabase , MEGAResdatabase  and ARG-database . Furtherdatabase . The enzdatabase .T and P7388T, was also examined by transmission electron microscopy using a Morgagni 268D Philips (FEI Company) electron microscope.The colony morphology was determined on R2A agar after 48 h cultivation at 20\u00b0C. The cellular morphology of all strains was observed by light microscopy after Gram staining. The cellular morphology of the type strains, P40232PO4/0.1 M NaOH; pH 9.0\u201310.0, 0.1 M NaHCO3/0.1 M Na2CO3. The pH value of the TSB was confirmed after autoclaving.The growth at different temperatures  and the salt tolerance  were determined on R2A agar plates. The pH tolerance was assessed in Trypticase Soya Broth (TSB) inoculated with two drops of cell suspension of concentration as McFarland 2.0 with pH 5\u201310 at intervals of 1.0 pH unit at 20\u00b0C adjusted with the following buffer systems: pH 5.0\u20138.0, 0.1 M KHF. hercynium CCM 9054T, F. branchiicola CCM 9061T, F. chilense CCM 7940T, F. araucananum CCM 7939T, F. psychroterrae CCM 8827T and F. saccharophilum CCM 8770T were phenotypically characterized by the most relevant tests recommended for description of new taxa within the family Flavobacteriaceae (2S on Triple Sugar Iron Agar (HiMedia), hydrolysis of aesculin, starch   , phosphoe (ONPG) , nitratee (ONPG) , utilizae (ONPG)  and sodie (ONPG) . The pree (ONPG) .2, both on R2A agar plates. All above listed tests were inoculated using cells of analyzed strains grown on R2A agar at 20\u00b0C for 48 hrs. All tests were read daily for up to 7 days with exception of the tyrosine hydrolysis test (up to 10 days). The utilization of carbon sources, enzyme activities and other additional phenotypic characteristics were further tested using identification test kits GEN III MicroPlateTN (Biolog) with the protocol A, API 20 NE and API ZYM (bioM\u00e9rieux) according to the manufacturer\u2019s instructions.Capability of growth on different media was tested on R2A agar (Oxoid), Nutrient agar (Oxoid), Plate Count agar (Oxoid), Tryptone Soya agar (Oxoid), marine agar , MacConkIn vitro antimicrobial susceptibility was assessed using the disk diffusion method. Cell suspensions of concentration as 0.5 McFarland were prepared from cultures cultivated on R2A agar. Suspension containing each strain (100 \u03bcl) was spread on the Mueller-Hinton agar plates. Tested antibiotic disks  were as follows: ampicillin (10 \u03bcg), aztreonam (30 \u03bcg), carbenicillin (100 \u03bcg), cefixime (5 \u03bcg), ceftazidime (10 \u03bcg), cephalotin (30 \u03bcg), ciprofloxacin (5 \u03bcg), gentamicin (10 \u03bcg), chloramphenicol (30 \u03bcg), imipenem (10 \u03bcg), kanamycin (30 \u03bcg), co-trimoxazole (25 \u03bcg), piperacillin (30 \u03bcg), polymyxin B (300 U), streptomycin (10 \u03bcg) and tetracycline (30 \u03bcg). CLSI  and EUCAST  standards were strictly followed for cultivation and inhibition zone diameter reading , F. hercynium DSM 18292T (98.54%) and F. chilense LMG 26360T (98.39%). A proposed type strain for the second cluster, P7388T, showed the highest sequence similarities to the F. branchiicola 59B-3-09T (99.12%), F. araucananum DSM 24704T (99.08%) and F. psychroterrae CCM 8827T (99.04%). The 16S rRNA gene sequence similarities of strain P4023T fell below the cut-off value 98.65%, whereas strain P7388T reached similarities above this threshold. Nevertheless, several studies have already proved that the standard 16S rRNA gene sequence cut-off value is not sufficient for some genera, such as Streptomyces, Streptococcus or Bacillus  could not be assigned to a specific class of orthologous genes clusters (COGs) or with specific function  . Howeverhnsoniae .T contained 112 scaffolds with a draft genome size of 4,387,206 bp and the genomic G + C content 34.5 mol% . Prediction of PULs found seven putative loci involved in degradation of carrageenan, dextran, hemicellulose, chitin, pectin, starch and xylan, which implies the ability of strain P7388T to degrade various polysaccharide substrates.Functional annotation revealed that 485 genes (12.97%) could not be assigned to any COGs or with specific function  . The remT and P7388T. The analysis indicated that these strains harbor four (P4023T) and seven (P7388T) gene clusters potentially related to secondary metabolite production. Out of these, six clusters did not match any known BGCs deposited in the MIBiG database -non-ribosomal peptide synthetase (NRPS) BGC for strain P7388T that did not match any known BGCs   , 3. ThisF. hercynium DSM 18292T, F. saccharophilum DSM 1811T and F. pectinovorum DSM 6368T. Genome-mining focused on cold-adaptation confirmed that Flavobacterium spp. are well adjusted to environmental stress and harbor a significant number of genes associated with cold-adaptation regardless of their thermotype , transcription termination/antitermination factor (nusA), translation initiation factors IF1 and IF2 , polynucleotide nucleotidyltransferase (pnp), heat-shock cognate proteins (hsc), recombinase A (recA) and many others , a member of family Flavobacteriaceae, was found to substantially increase tolerance to freezing and thus considered a primary response to ensure freeze-tolerance of its hosts , chromosomal replication initiation ATPase (dnaA), DNA topoisomerases IV , transcription antitermination factor (nusA), translation factors IF-1, 2, 3  along with other cold-inducible genes were present in all genomes in one copy, regardless of the thermotype (rpoD) and sigma factor 24 (rpoE) were present in genomes only in one and two copies, respectively, although psychrotrophic Flavobacterium spp. were previously found to harbor multiple copies of these genes , superoxide-dismutase and peroxidase , or thioredoxin and peroxiredoxin reductases . Although all analyzed strains harbored peroxiredoxin genes in multiple copies , strain P4023T harbored multiple copies of superoxide-dismutase (sodA) and thioredoxin (trxA), which may be explained by its isolation from surface rather than deeper soil layers and therefore exposed to higher oxidative stress and stronger UV radiation. Similarly versatile microorganisms to flavobacteria are pseudomonads that colonize diverse Antarctic areas including soils, fresh or marine waters  by Pseudomonas extremaustralis 14-3T (Cold-adaptation is inseparably associated with oxidative stress response as a result of higher oxygen solubility at lower temperatures leading to increased levels of reactive oxygen species (ROS). Notably, e waters . Considee waters . Howeveris 14-3T . None ofT harbored a crtZ gene required for zeaxanthin biosynthesis  is a membrane light-driven protein acting as an outward Hon pumps . Both th retinal . PRs wer retinal  and inte retinal . Neverth. MED134 , which mFlavobacterium strains, since the type II CRISPR/Cas-system was found to be an adaptive immune system in other cold-adapted flavobacteria  were predicted in the P4023T genome likely to provide resistance against 16 different drug classes (T genome (in vitro antibiotic susceptibility testing. Correlation between resistome predictions and in vitro testing was found for resistance to aminoglycosides which is presumably encoded by aadS gene (MBP4138141.1) in the genome of P7388T, a widely spread bacterial adenylyltransferase conferring resistance to aminoglycoside antibiotics (JOHN-1 present in P4023T (MBP4142760.1) and P7388T (MBP4137930.1) genomes. This gene provides resistance to a broad spectrum of \u03b2-lactam antibiotics in Flavobacterium johnsoniae (in vitro susceptibility results. Additive effects on resistance to \u03b2-lactam antibiotics may result from the presence of putative OXA beta-lactamase OXA-29 (MBP4140625.1) in P4023T, known to hydrolyze penams and cephalosporines but is not effective against carbapenems , ciprofloxacin (5 \u03bcg) and chloramphenicol (30 \u03bcg). Disagreement between the phenotypic susceptibility pattern and ARGs computational prediction is a well-known controversy, especially in clinical bacteriology (in vivo vs. in vitro), testing of single strain culture vs. biofilm formation, presence of specific metabolites and growth factors, presence of specific phages or inadequate time/concentration of tested antibiotics that induce expression of resistance genes. Any of above-mentioned factors or even their combination may be also responsible for discrepancies observed among analyzed Antarctic Flavobacterium isolates, in particular considering extreme environment they colonize and presence of phages in their genomes.Although routine screening did not find any resistance genes, RGI prediction of the resistome from draft genomes of P4023 classes . InactivT genome , they maibiotics . All Antibiotics . This wihnsoniae , howeverbapenems . Both steriology . There aT additionally harbors a putative rosA gene (MBP4140897.1) encoding an efflux pump/potassium antiporter system. This efflux transporter has been detected in marine Flavobacterium spp. (Bacteroidetes (rosA gene is related to polymyxin B resistance and represents a part of a two component efflux antiporter system (RosAB) found in Yersinia enterocolitica and confers resistance to cationic antimicrobial peptides including polymyxin B , which is a well-known environment with resistant bacteria . AlthougT and P7388T were aerobic, Gram-negative rods with rounded ends, with average cell size 0.4\u20130.6 \u03bcm \u00d7 1.2\u20132.4 \u03bcm and 0.3\u20130.4 \u03bcm \u00d7 1.5\u20133.0 \u03bcm, respectively , Summed Feature 3 (C16:1 \u03c97c/C16:1 \u03c96c) (9.9%), anteiso-C15:0 (9.8%), iso-C15:0 (9.3%) and iso-C16:0 3OH (9.3%). The major fatty acid of the group comprising strain P7388T were iso-C15:0 (23.4%), Summed Feature 3 (C16:1 \u03c97c/C16:1 \u03c96c) (9.9%), iso-C17:0 3OH (8.5%) and iso-C15:0 3OH (8.4%). The proposed species differ not only qualitatively and quantitatively in the profiles of major fatty acids, but also quantitatively in minor fatty acids, especially iso-C14:0, iso-C16:1 H and Summed Feature 9 (C16:0 10-methyl/iso-C17:1). The overall fatty acids profile of both groups is in agreement with the genus description ] and minor amounts of putrescine [0.7 \u03bcmol (g dry weight\u20131)] and spermidine [0.2 \u03bcmol (g dry weight\u20131)]. The polyamine pattern of strain P7388T was similar with the major polyamine sym-homospermidine [20.5 \u03bcmol (g dry weight\u20131)] and minor amounts of putrescine [0.2 \u03bcmol (g dry weight\u20131)] and spermidine [0.3 \u03bcmol (g dry weight\u20131)]. Quinone systems, polar lipid profile and polyamine pattern are well in agreement with the genus description .L-tyrosine agar. Negative for hydrolysis of Tween 80, aesculin, ONPG, starch, DNA, CMC and agar. Does not produce lecithinase. Does not produce H2S. Positive for arginine dihydrolase, and negative for ornithine and lysine decarboxylases. Does produce acid from glucose and maltose in aerobic conditions. Negative for production of acid from fructose and xylose in aerobic conditions. Positive for utilization of glucose and maltose by API 20 NE. Negative for utilization of arabinose, mannose, N-acetylglucosamine, gluconic acid, capric acid, adipic acid, malic acid, citric acid and phenylacetic acid by API 20 NE. Positive for alkaline phosphatase, leucine arylamidase, valine arylamidase and acid phosphatase by API ZYM. Negative for esterase (C 4), esterase lipase (C 8), lipase (C 14), cystine arylamidase, trypsin, \u03b1-chymotrypsin, naphtol-AS-BI-phosphohydrolase, \u03b1-galactosidase, \u03b2-galactosidase, \u03b2-glucuronidase, \u03b1-glucosidase, \u03b2-glucosidase, N-acetyl-\u03b2-glucosaminidase, \u03b1-mannosidase, and \u03b1-fucosidase by API ZYM. Carbon source utilization ability via respiration determined by Biolog GEN III MicroPlate test panels is positive for D-maltose, \u03b1-D-glucose, D-glucose-6-PO4, gelatine, L-arginine, L-aspartic acid, L-glutamic acid, acetoacetic acid and acetic acid. Negative for D-trehalose, D-cellobiose, gentiobiose, sucrose, stachyose, D-raffinose, \u03b1-D-lactose, D-melibiose, \u03b2-methyl-D-glucoside, D-salicin, N-acetyl-\u03b2-D-mannosamine, N-acetyl neuraminic acid, D-mannose, D-fructose, D-galactose, 3-methyl glucose, D-fucose, L-fucose, L-rhamnose, inosine, D-sorbitol, D-mannitol, D-arabitol, myo-inositol, glycerol, D-fructose-6-PO4, D-aspartic acid, D-serine, L-alanine, L-histidine, L-pyroglutamic acid, L-serine, D-galacturonic acid, D-galactonic acid lactone, D-gluconic acid, D-glucuronic acid, glucuronamide, mucic acid, quinic acid, D-saccharic acid, p-hydroxy phenylacetic acid, methyl pyruvate, D-lactic acid methyl ester, L-lactic acid, citric acid, \u03b1-keto glutaric acid, D-malic acid, L-malic acid, bromo-succinic acid, Tween 40, \u03b3-amino-butyric acid, \u03b1-hydroxy-butyric acid, \u03b2-hydroxy-D,L-butyric acid, \u03b1-keto butyric acid, propionic acid and formic acid.Cells are Gram-stain-negative rods with rounded ends, cell size in range 0.4\u20130.6 \u03bcm \u00d7 1.2\u20132.4 \u03bcm, cells occurring in irregular clusters, occasionally singly or in pairs. Endospores are not formed. Does not produce a capsule. Negative for presence of flexirubin-type pigments. Does not adhere to agar. Yellowish colonies. Motile with gliding-activity. Catalase and oxidase positive. Growth occurs on R2A, PCA, TSA, marine agar NA, blood agar with 5% sheep blood, BHI, Mueller-Hinton and Endo agar. Does not grow on MacConkey agar. Grows in pH range 6\u20139 and temperature range 1\u201330\u00b0C with optimum growth at 20\u00b0C and pH around 7. Cells grow well in presence of 1% NaCl, and 2% NaCl inhibits growth. Does grow in microaerophilic conditions but growth in anoxic conditions is limited. Does not produce fluorescein on King B medium. No utilization of Simmon\u2019s citrate, malonate and acetamide. Negative for reduction of nitrates and nitrites. Negative for production of urease and indole. Positive for hydrolysis of gelatine, casein, and tyrosine. Does not produce brown diffusible pigment on 15:1 \u03c96c, Summed Feature 3 (C16:1 \u03c97c/ C16:1 \u03c96c), anteiso-C15:0, iso-C15:0 and iso-C16:0 3OH. Major respiratory quinone is MK-6 and major polyamine is sym-homospermidine. Polar lipid profile contains major amounts of phosphatidylethanolamine, an ornithine lipid, and two unidentified lipids  lacking a functional group, moderate amounts of unidentified lipid L1, unidentified glycolipid GL, minor amounts of an unidentified aminophospholipid (APL), and two unidentified lipids . The DNA G + C content of the type strain is 31.2 mol%.The major fatty acids are iso-CT (= CCM 9062T = LMG 31963T) was isolated in 2011 from the organic material of an abandoned bird nest located at the Lachman Cape . All characteristics listed in the species description are shared by all strains, except for the following strain-dependent test results: motility, production of acid from mannitol, and variable results observed by Biolog GEN III MicroPlate were utilization of dextrin, D-turanose, N-acetyl-D-glucosamine, glycyl-L-proline and pectin. A formal proposal of the species \u201cFlavobacterium flabelliforme sp. nov.\u201d is given in Type strain P4023Flavobacterium flabelliforme sp. nov. P4023T are MW691162 and JAGFBU000000000, respectively.The GenBank/EMBL/DDBJ accession numbers for the near full length 16S rRNA gene sequences and whole genome sequences for Flavobacterium geliluteum .L-tyrosine agar. Negative for hydrolysis of DNA and agar. Does not produce lecithinase. Does not produce H2S. Positive for arginine dihydrolase and negative for ornithine and lysine decarboxylases. Does produce acid from glucose, maltose and xylose in aerobic conditions. Negative for production of acid from mannitol in aerobic conditions. Positive for utilization of glucose, arabinose, mannose, N-acetyl-glucosamine, maltose, and hydrolysis of aesculin by API 20 NE. Negative for utilization of gluconic acid, capric acid, adipic acid, malic acid, citric acid and phenylacetic acid by API 20 NE. Positive for alkaline phosphatase, leucine arylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase and \u03b2-glucosidase by API ZYM. Negative for esterase (C 4), esterase lipase (C 8), lipase (C 14), valine arylamidase, cystine arylamidase, trypsin, \u03b1-chymotrypsin, \u03b1-galactosidase, \u03b2-galactosidase, \u03b2-glucuronidase, N-acetyl-\u03b2-glucosaminidase, \u03b1-mannosidase and \u03b1-fucosidase by API ZYM. Carbon source utilization ability via respiration determined by Biolog GEN III MicroPlate test panels is positive for D-trehalose, D-cellobiose, gentiobiose, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, \u03b1-D-glucose, D-mannose, D-glucose-6-PO4, glycyl-L-proline, L-aspartic acid, L-glutamic acid, D-galacturonic acid, acetoacetic acid and acetic acid. Negative for sucrose, stachyose, D-raffinose, \u03b1-D-lactose, D-melibiose, N-acetyl-\u03b2-D-mannosamine, N-acetyl neuraminic acid, 3-methyl glucose, D-fucose, L-fucose, L-rhamnose, inosine, D-sorbitol, D-mannitol, D-arabitol, myo-inositol, glycerol, D-aspartic acid, D-serine, L-alanine, D-gluconic acid, D-glucuronic acid, glucuronamide, D-saccharic acid, quinic acid, p-hydroxy phenylacetic acid, L-lactic acid, \u03b1-keto glutaric acid, D-malic acid, L-malic acid, bromo-succinic acid, \u03b3-amino-butyric acid, \u03b1-hydroxy-butyric acid, \u03b2-hydroxy-D,L-butyric acid, \u03b1-keto butyric acid, propionic acid, and formic acid.Cells are Gram-stain-negative rods with rounded ends, cell size in range 0.3\u20130.4 \u03bcm \u00d7 1.5\u20133.0 \u03bcm, occurring singly and in irregular clusters, occasionally in pairs. Endospores are not formed. Does not produce a capsule. Dark yellow to orange pigmented colonies. Flexirubin-type pigment present. Does not adhere to agar. Motile with gliding-activity. Catalase positive. Oxidase negative. Growth occurs on R2A, PCA, TSA, NA, blood agar with 5% sheep blood, BHI and Mueller-Hinton agar. Does not grow on marine and MacConkey agar. Grows in pH range 6\u20138 and temperature range 15\u201330\u00b0C with optimum growth at 20\u00b0C and pH around 7. Cells grow in presence of 0.5% NaCl, and 1% NaCl inhibits growth. Does grow in microaerophilic conditions but growth in anoxic conditions is limited. Does not produce fluorescein on King B medium. No utilization of Simmon\u2019s citrate, malonate and acetamide. Negative for reduction of nitrates and nitrites. Negative for production of urease and indole. Positive for hydrolysis of gelatine, aesculin, ONPG, starch, casein, tyrosine and CMC. Does not produce brown diffusible pigment on 15:0, Summed Feature 3 (C16:1 \u03c97c/C16:1 \u03c96c), iso-C17:0 3OH and iso-C15:0 3OH. Major respiratory quinone is menaquinone MK-6 and major polyamine is sym-homospermidine. Polar lipid profile contains major lipids phosphatidylethanolamine, an ornithine lipid, two unidentified lipids lacking a functional group , moderate amounts of unidentified lipid L1, minor amounts of lipids L2 and L6 and an unidentified glycolipid (GL). The DNA G + C content of the type strain is 34.5 mol%.The major fatty acids are iso-CT (= CCM 9064T = LMG 31962T) was isolated from water samples taken in 2016 from a small temporary lake . All characteristics listed in the species description are shared by all strains, except for the following strain-dependent test results: growth at 5 and 10\u00b0C, growth on Endo agar, hydrolysis of Tween 80, production of acid from fructose, presence of \u03b1-glucosidase in API ZYM and variable results observed by Biolog GEN III MicroPlate were utilization of dextrin, D-maltose, pectin, \u03b2-methyl-D-glucoside, D-salicin, D-fructose, D-galactose, D-fructose-6-PO4, gelatine, L-arginine, D-galactonic acid lactone, mucic acid, methyl pyruvate, D-lactic acid methyl ester, citric acid and Tween 40. A formal proposal of the species \u201cFlavobacterium geliluteum sp. nov.\u201d is given in Type strain P7388Flavobacterium geliluteum sp. nov. P7388T are MW691150 and JAGFBV000000000, respectively.The GenBank/EMBL/DDBJ accession numbers for the near full length 16S rRNA gene sequences and whole genome sequences for https://www.ncbi.nlm.nih.gov/genbank/, MW691162; https://www.ncbi.nlm.nih.gov/genbank/, JAGFBU000000000; https://www.ncbi.nlm.nih.gov/genbank/, MW691150; https://www.ncbi.nlm.nih.gov/genbank/, JAGFBV000000000.The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: SK performed phylogenetic, phylogenomic, genomic analysis, analysis of fatty acid-methyl esters, and drafted and finalized the manuscript. H-JB performed chemotaxonomic analyses of polar lipids, menaquinones, and polyamines. MB was responsible for whole-genome sequencing. MS-P was involved in genomic analysis of biosynthetic potential. MN was involved in genome assembly and quality of WGS data. DK performed electron microscopy. ES and IS performed morphological, physiological, and biochemical characterization including antibiotic susceptibility. SK drafted the manuscript with inputs of H-JB, MB, MS-P, and IS. All authors edited the draft manuscript and agreed to the final manuscript version for submission.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Investigation of the frequencies of functionally signif icant gene variants in the context of medical biology and gene geography is a relevant issue for studying the genetic structure of human populations. The transition from a traditional to an urbanized lifestyle leads to a higher incidence of civilizational diseases associated with metabolic disorders, including type 2 diabetes mellitus. The goal of the present paper is to analyze the frequencies of functionally signif icant gene alleles in the metabolic prof iles of indigenous Siberian peoples to identify the gene pool resilience, evaluate the susceptibility of various ethnic groups to metabolic disorders under changing environmental conditions, and predict the epidemiological situation that may occur in the near future. The study was performed in the monoethnic samples of eastern and western Buryats, Teleuts, Dolgans, and two territorial groups of Yakuts. A real-time PCR was used to determine the frequencies of single nucleotide polymorphisms (SNPs) G103894T, rs12255372, and C53341T, rs7903146 in the TCF7L2 gene. The results obtained were compared to the frequencies identif ied for Russians from Eastern Siberia and the values available in the literature. The frequencies of the polymorphic variants studied in the samples from the indigenous Siberian peoples place them in between Caucasian and East Asian populations, following the geographicgradient of polymorphism distribution. A signif icantly lower occurrence of type 2 diabetes risk alleles TCF7L2 (103894T)and TCF7L2 (53341T) in the samples of indigenous Siberian peoples compared to Russians was observed, which agreeswith their lower susceptibility to metabolic disorders compared to the newcomer Caucasian population. Taking intoaccount urbanization, a reduced growth in type 2 diabetes incidence may be predicted in indigenous Siberian peoples,i. e. Buryats, Yakuts, Dolgans, and Teleuts, compared to the newcomer Caucasian population. A further study of populationstructure with respect to different metabolic prof ile genes is required to better understand the molecular geneticfoundations of the adaptive potential of indigenous Siberian peoples Investigation into the peculiarities of the population geneticstructure of ethnic groups in the context of medical biologyand gene geography is a relevant issue in human genetics. Tobetter understand molecular genetic foundations of adaptivepotential that ethnic groups develop as they evolve under specificclimatic and geographic conditions and adapt to specificdietary patterns, it is important to analyze the frequencies ofthe candidate gene alleles proven to be functionally significantbased on studies in individual populations.Type 2 diabetes mellitus (DM2) is among the leading mortalityand disability factors in a working-age population . DM2 is a metabolic syndrome componentand above that is linked to increased risk of multiple associatedpathological states, primarily including cardiovasculardiseases  andchronic renal failureIncretin hormone secretion defect, a key element of DM2pathogenesis, is associated with TCF7L2 gene polymorphismsince it is this gene\u2019s product that regulates the production ofpancreatic \u03b2-cells from pluripotent stem cells and is involvedin glucose-stimulated insulin secretion .In addition, the gene also targets the brain, where TCF7L2determines the intensity of the anorexigenic effect and affectsthe central glucose homeostasis mechanism . In the liver, the gene is involved in the regulationof triglycerides and low- and very low-density lipoproteinexchange. It is also involved in gluconeogenesis and acts asan insulin resistance mediator .It was found that SNPs G103894T, rs12255372, andC53341T, rs7903146 in introns 3 and 4 of gene TCF7L2were associated with DM2 . The linkof TCF7L2 (103894T ) and TCF7L2 (53341T ) alleles withincreased risk of DM2 was demonstrated in a number ofpopulations around the world, including Russia . It was shown that the TCF7L2 (53341T )variant was linked to increased risk of DM2 compared toTCF7L2 (103894T ), with homozygous alleles showing highersusceptibility to the disease than heterozygous ones The TCF7L2 polymorphisms are also linked to BMI, totalbody fat volume, as well as subcutaneous and visceral fat. TCF7L2 (53341T )allele is associated with the risk of myocardial ischemia andmyocardial infarction as syntropic diseases with commonpathogenetic elements . Gene TCF7L2 is also linked to renal embryogenesis,i. e. its polymorphisms are associated with variousdegrees of chronic renal failure, a vascular complication ofDM2 . It was proved that TCF7L2 polymorphism inloci rs7903146 and rs12255372 was associated with risks ofgastric, breast, and colorectal cancer . The effect of natural selectionon locus rs7903146 in gene TCF7L2 was discovered and astatistically significant link between 53341\u0422 allele frequencyand several climatic geographic factors was shown .Studies on the frequencies of gene alleles associated withthe risk of DM2 and other metabolic disorders in indigenousSiberian populations have remained relevant throughout therecent decade . However,the distribution of the polymorphic variants of functionallysignificant gene TCF7L2 in Siberian populations remainsunderstudied. Polymorphism frequencies in locus rs7903146for some Siberian peoples, including Buryats and Yakuts,were presented in . Unfortunately, theauthors did not indicate the area where genetic material wascollected, which seems necessary for these large heterogeneousethnic groups populating vast territoriesThe present paper reports the results of a study into thefrequencies of polymorphisms G103894T, rs12255372, andC53341T, rs7903146 in gene TCF7L2 associated with severaldiseases in the populations of indigenous Siberian ethnicgroups, namely Buryats, Teleuts, Yakuts and Dolgans, incomparison to Russians living in Siberia.The genetic material for the present research was collectedin the field in 2000\u20132006. Blood samples were taken fromapparently healthy volunteers under their informed consentand with the approval of the local healthcare authorities andthe Ethics Committee of the Institute of Cytology and Genetics,SB RAS. Before blood sampling, all volunteers filled ina special demographic questionnaire to specify their ancestors\u2019nationalities down to 3 to 4 generations.The data obtained were used to form 7 population samplescovering Southern and Eastern Siberia. Persons of Buryatnationality with no outsider ancestors living in Alkhanay andOrlovsky settlements in the Aginsky Buryat Okrug (ABO) ofZabaykalsky Krai were included in the Eastern Buryat group (N = 132). Ethnic Buryats from settlements of Ekhirit-BulagatskyDistrict of Ust-Ordynsky Buryat Okrug (UOB) ofthe Irkutsk Region (N = 278) were included in the Westernsample. Also included in the study were Teleuts from the BelovoDistrict of the Kemerovo Region (N = 116). Two ethni-callyhomogeneous samples of Yakuts were formed as follows:the Nyurbinsky group included the residents of settlementsNyurbachan and Syultsy of the Nyurbinsky District(N = 109), and the Ust-Aldansky group \u2013 the residents of theDyupsya settlement of the Ust-Aldansky District (N = 100).The residents of the town of Dudinka and settlements Volochankaand Ust-Avam of the Taymyr Dolgan-Nenets Okrugof Krasnoyarsk Krai identifying as ethnic Dolgans were includedin the Dolgan sample (N = 180). The seventh samplecombined Russians from Zabaykalsky Krai and the IrkutskRegion (N = 133).DNA samples were isolated from the leukocyte fractionof venous blood using the BioSilica kits (Russia). Real-timeSNP genotyping in genes TCF7L2 and TCF7L2  was performed applyingcompeting TaqMan-probes complementary to polymorphicDNA segments. Primer and probe designs wereselected using the sequences available in the NCBI database(http://www.ncbi.nlm.nih.gov/) with UGENE  and Oligo Analyzer  software(Table 1).Amplification was performed in 25-\u03bcl final volume, themaster mix included 300 nM primers, 100 nM TaqMan probes,65 mM TrisHCl (\u0440\u041d 8.9), 16 mM (NH4)2SO4, 2.5 mM MgCl2,0.05 % Tween-20, 0.2 mM dNTP, 0.5\u201310 ng DNA, and 0.5 UTaq DNA polymerase . Reactionconditions were as follows: initial denaturation for 3 min at96 \u00b0\u0421 was followed by 46 cycles including denaturation at96 \u00b0\u0421 for 5 s, primer annealing, and extension at 61 \u00b0\u0421 for 30 s.Allele variant frequencies in the populations were determinedbased on observed genotype frequencies. The matchbetween empirically observed genotype frequency distributionand theoretically expected distribution at the Hardy\u2013Weinbergequilibrium was tested using Pearson\u2019s chi-squared (theequilibrium holds at p > 0.05). The statistical confidence ofallele frequency differences between the studied sampleswas evaluated using the chi-squared test with Yates continuitycorrection; the results were considered statisticallysignificant at p < 0.025 .Genotyping results for TCF7L2  and in samples of Buryats, Teleuts, Yakuts,Dolgans, and Russians from Eastern Siberia are presented inTable 2.The genotype distribution matched the Hardy\u2013Weinbergequilibrium for all polymorphic loci and samples. The frequenciesof alleles TCF7L2 (103894T ) and TCF7L2 (53341T ) inthe studied samples and some ethnic groups described in theliterature , as well as comparisonof populations , are presented in Tables 3\u20134.It was shown that TCF7L2 (103894T ) allele frequency inthe Russians sample (23.1 %) matched that in other Caucasiangroups (22\u201337 %) . The frequencyin the studied samples of indigenous Siberian peoplesvaries from 5.4 % for Western Buryats to 9.5 % for Teleuts,with no statistically significant differences observed. However,the allele frequency in all samples of indigenous populationswas significantly lower than in Russians from Eastern Siberiaand other Caucasian groups described in the literature . At the same time, it was significantlyhigher than in several East Asian populations, i. e. Chinese andVietnamese. We could also see a significant difference betweenTeleuts and Japanese not observed for other studied groups.This in-between position of indigenous Siberian populations,as exemplified by Buryats and Teleuts, had been demonstratedearlier in the polymorphism frequencies of some other metabolicprofile genes .TCF7L2 (53341T ) allele frequency in the Russians sample(25.9 %) matched that in other Caucasian groups (23\u201340 %). The studied samples of indigenouspopulations showed a significantly lower value comparedto Russians varying from 4.5 % in Yakuts from the NyurbinskyDistrict to 11.3 % in Teleuts. Statistically significantdifferences were discovered between Teleuts and the sampleswith the lowest frequency values, namely Eastern Buryats(4.9 %) and Yakuts from the Nyurbinsky District. The data onTCF7L2 (53341T) allele frequency in the samples of Buryats(6.3 %) and Yakuts (4.3 %) resembling the results obtainedin our study were presented by Trifonova et al. (2020). Unfortunately,the authors did not indicate sample sizes and theparticipants\u2019 places of residence, so confidence evaluationwas impossible to perform. The frequency values do not show significant differences between the samples of Eastern Buryatsand Yakuts from the Nyurbinsky District and the samples ofindigenous East Asian populations, namely Chinese, Japaneseand Vietnamese, available in the literature. Yakuts from theUst-Aldansky District demonstrated a significantly higherTCF7L2 (53341T ) allele frequency than Vietnamese, whileDolgans showed differences compared to some Chinesepopulations as well. Western Buryats and Teleuts showedsignificantly higher allele frequencies than all the East Asiansamples described in the literature. It was also shown thatthis polymorphism frequency in populations of indigenousSiberian peoples was significantly lower than in the Caucasiangroups described in the literature . Thus, TCF7L2 (53341T ) allele frequencies also confirmthe trend that places samples of indigenous Siberianpeoples in-between East Asian and Caucasian populationsInvestigation of the frequencies of functionally significantgene variants in the context of medical biology and genegeographyis a relevant issue for studying the population geneticstructure of indigenous Siberian peoples. In the presentpaper, we have determined the frequencies of the 103894Tand 53341T alleles in gene TCF7L2 associated with DM2 andother metabolic disorders in the populations of Buryats, Yakuts,Dolgans, and Teleuts, as well as a sample of Russians fromEastern Siberia. It was shown that these frequencies in Russiansfall within the same range as in other Caucasian populations.Meanwhile, the populations of indigenous Siberianethnic groups show significantly lower TCF7L2 (103894T )and TCF7L2 (53341T ) polymorphism frequencies, whichplaces them in-between Caucasian and East Asian populations.It was shown in several papers that indigenous Siberian andFar Eastern ethnic groups, as well as the ethnic groups from theEuropean part of Russia with a mongoloid component in theirgene pool, had lower incidence rates of metabolic syndromeand its DM2 component compared to Caucasians .It is primarily explained by the traditional lifestyle implyinga sufficient amount of physical activity and diet consistingmostly of animal source foods rich in proteins and fats withlimited carbohydrate component .Ethnic peculiarities in DM2 prevalence and manifestationsare also caused by distinctions from the European gene pool,i. e. a unique combination of frequencies of functionallysignificant genes developed as a result of adaptation to localenvironmental conditions . Differences in living conditions between indigenousand newcomer populations are alleviated due to urbanization,centuries-long traditions, and acquired dietary patterns change,and, as a result, civilizational diseases associated with metabolicdisorders, including DM2, become increasingly commonin indigenous populations . Investigation ofthe polymorphism distribution of the functionally significantgenes associated with risks of diseases in indigenous Siberianpopulations makes it possible to identify the gene pool resilience,evaluate the susceptibility of various ethnic groups tometabolic disorders under changing environmental conditions,and predict the epidemiological situation in the near future.Lower prevalence of DM2 among indigenous Siberianpopulations agrees with reduced populational frequencies ofstudied alleles TCF7L2 (103894T ) and TCF7L2 (53341T )associated with DM2 and several syntropic diseases, discoveredin the present paper. The reduced frequencies ofthese polymorphisms may affect the incidence rates of thediseases in the studied populations. With urbanization takeninto account, one might predict reduced growth in incidencerates of DM2 and other pathological states associated withstudied polymorphisms in indigenous Siberian ethnic groupscompared to newcomer Caucasians.The high frequency of TCF7L2 (53341T ) polymorphismin Teleuts from the Kemerovo Region compared to Buryatsand Yakuts may be attributed to a Caucasian component thatthis ethnic group adopted in their gene pool in the process offormation . With more comfortableliving conditions close to cities of Prokopyevsk, Kemerovo,and Novokuznetsk and a richer European-type diet, Teleutsmay face an increased risk of DM2 and associated diseases.Increased incidence of cardiovascular diseases has been observedin this ethnic group in recent decades . However, polymorphism frequencies of otherfunctionally significant genes are to be investigated to drawbetter-grounded conclusions.Ethnic peculiarities in the frequency distribution of polymorphismsin gene TCF7L2  and in the populations of Buryats, Yakuts,Dolgans, and Teleuts, as well as a sample of Russians fromEastern Siberia, have been studied in the present paper. Locusrs12255372 has been studied in various territorial groups ofBuryats and Yakuts for the first time, and the same goes forloci rs12255372 and rs7903146 in the Dolgan and Teleutpopulations. It has been shown that the samples of indigenousSiberian populations fall in-between Caucasian and East Asianpopulations with respect to studied polymorphism frequencies,following the geographic polymorphism distributiongradient.Significantly lower occurrence of TCF7L2 (103894T ) andTCF7L2 (53341T ) alleles associated with DM2 and othermetabolic disorders in the samples of indigenous Siberianpeoples compared to Russians was demonstrated, which agreeswith their lower susceptibility to metabolic disorders, includingDM2, compared to the newcomer Caucasian populationdescribed in the literature. With the transition to urbanizedlifestyle taken into account, one might predict reduced growthin incidence rates of DM2 and other pathological states associatedwith the studied polymorphisms in indigenous Siberianethnic groups, namely Buryats, Yakuts, Dolgans, and Teleuts,compared to newcomer Caucasians.To better understand the nature of ethnic differences, furtherinvestigation into population structure with respect to othermetabolic profile genes is required.The authors declare no conflict of interest.Ametov A.S., Kamynina L.L., Akhmedova Z.G. The clinical aspectsof effectiveness of incretin therapy (Wnt-pathogenic path and polymorphismof gene TCF7L2). Rossiyskiy Meditsinskiy Zhurnal =Medical Journal of the Russian Federation. 2016;22(1):47-51. DOI10.18821/0869-2106-2016-22-1-47-51. (in Russian)Anjum N., Jehangir A., Liu Y. Two TCF7L2 variants associated withtype 2 diabetes in the Han nationality residents of China. J. Coll.Physicians Surg. Pak. 2018;28(10):794-797.Asfandiyarova N.S. 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(in Russian)Kurtanov Kh.A., Sydykova L.A., Pavlova N.I., Filippova N.P., DodokhovV.V., Apsolikhova G.A., Solov\u2019eva N.A., D\u2019yakonova A.T.,Neustroeva L.M., Varlamova M.A., Borisova N.V. Polymorphismof the adiponutrin gene (PNPLA3) in the indigenous inhabitantsof the Republic of Sakha (Yakutia) with type 2 diabetes mellitus.Al\u2019manakh Klinicheskoy Meditsiny = Almanac of Clinical Medicine.2018;46(3):258-263. DOI 10.18786/2072-0505-2018-46-3-258-263.(in Russian)Mel\u2019nikova E.S., Rymar O.D., Ivanova A.A., Mustafina S.V., ShapkinaM.Ju., Bobak M., Maljutina S.K., Voevoda M.I., MaximovV.N. Association of polymorphisms of genes TCF7L2, FABP2,KCNQ1, ADIPOQ with the prognosis of the development of type 2diabetes mellitus. Terapevticheskiy arkhiv = Therapeutic Archive.2020;92(10):40-47. DOI 10.26442/00403660.2020.10.000393. (inRussian)Melzer D., Murray A., Hurst A.J., Weedon M.N., Bandinelli S., CorsiA.M., Ferrucci L., Paolisso G., Guralnik J.M., Frayling T.M.Effects of the diabetes linked TCF7L2 polymorphism in a representativeolder population. BMC Med. 2006;4:34. DOI 10.1186/1741-7015-4-34.Nobrega M.A. TCF7L2 and glucose metabolism: time to look beyondthe pancreas. Diabetes. 2013;62(3):706-708. DOI 10.2337/db12-1418.Orlov P.S., Kulikov I.V., Ustinov S.N., Gafarov V.V., Malyutina S.K.,Romashchenko A.G., Voyevoda M.I., Maksimov V.N. Associationanalysis of some single nucleotide polymorphism markers of thesecond type of diabetes with myocardial infarction. Byulleten\u2019 SORAMN = Bulletin of Siberian Branch of Russian Academy of MedicalSciences. 2011;31(5):19-24. (in Russian)Ostaptseva A.V., Shabaldin A.V., Akhmatianova V.R., Minina V.I.,Glushkov A.N., Druzhinin V.G., Zorkoltseva I.V., Shabaldin E.V.,Glushkova O.A., Makarchenko O.S., Ageyeva T.N. Molecular geneticanalysis of interleukin 4 gene polymorphism among Teleutians,Shorians, and Caucasians of Kemerovo region. MeditsinskayaImmunologiya = Medical Immunology. 2006;8(5-6):737-740. (inRussian)Ovsyannikova O.V., Podkhomutnikov V.M., Kolbasko A.V., LuzinaF.A., Gus\u2019kova E.V. Cardiovascular disease in rural Kuzbass aborigines\u2013 Teleut. Rossiyskiy Kardiologicheskiy Zhurnal = RussianJournal of Cardiology. 2007;6:59-62. (in Russian)Potapov V.A., Nosikov V.V., Shamkhalova M.N., Shestakova M.V., SmetaninaS.A., Bel\u2019chikova L.N., Suplotova L.A. TCF7L2 rs12255372and SLC30A8 rs13266634 confer susceptibility to type 2 diabetes ina Russian population. Russ. J. Genet. 2010;46(8):1001-1008. DOI10.1134/S1022795410080132.Rosales-Reynoso M.A., Arredondo-Valdez A.R., Ju\u00e1rez-V\u00e1zquez C.I.,Wence-Chavez L.I., Barros-N\u00fa\u00f1ez P., Gallegos-Arreola M.P., Flores-Mart\u00ednez S.E., Mor\u00e1n-Moguel M.C., S\u00e1nchez-Corona J. TCF7L2and CCND1 polymorphisms and its association with colorectal cancerin Mexican patients. Cell Mol. Biol. 2016;62(11):13-20. DOI10.14715/cmb/2016.62.11.3.Saxena R., Gianniny L., Burtt N.P., Lyssenko V., Giuducci C., Sj\u00f6grenM., Florez J.C., Almgren P., Isomaa B., Orho-Melander M.,Lindblad U., Daly M.J., Tuomi T., Hirschhorn J.N., Ardlie K.G.,Groop L.C., Altshuler D. Common single nucleotide polymorphismsin TCF7L2 are reproducibly associated with type 2 diabetes and reducethe insulin response to glucose in nondiabetic individuals. Diabetes.2006;55(10):2890-2895. DOI 10.2337/db06-0381.Sladek R., Rocheleau G., Rung J., Dina C., Shen L., Serre D., Boutin P.,Vincent D., Belisle A., Hadjadj S., Balkau B., Heude B., CharpentierG., Hudson T.J., Montpetit A., Pshezhetsky A.V., Prentki M.,Posner B.I., Balding D.J., Meyre D., Polychronakos C., Froguel P.A genome-wide association study identifies novel risk loci fortype 2 diabetes. Nature. 2007;445(7130):881-885. DOI 10.1038/nature05616.Smetanina S.A. Molecular-genetic and hormonal-metabolic associationsamong women of the Russian population of reproductive agewith obesity and early menarche. Meditsinskaya Nauka i ObrazovaniyeUrala = Medical Science and Education in Ural. 2015;2(1):126-129. (in Russian)Tabikhanova L.E., Osipova L.P., Voronina E.N., Bragin A.O., FilipenkoM.L. Polymorphism of lipid exchange genes in some populationsof South and East Siberia. Vavilovskii Zhurnal Genetiki i Selektsii =Vavilov Journal of Genetics and Breeding. 2019;23(8):1011-1019.DOI 10.18699/VJ19.578The 1000 Genomes Project Consortium. An integrated map of geneticvariation from 1,092 human genomes. Nature. 2012;491(7422):56-65. DOI 10.1038/nature11632.Timpson N.J., Lindgren C.M., Weedon M.N., Randall J., OuwehandW.H., Strachan D.P., Rayner N.W., Walker M., Hitman G.A.,Doney A.S., Palmer C.N., Morris A.D., Hattersley A.T., Zeggini E.,Frayling T.M., McCarthy M.I. Adiposity-related heterogeneity inpatterns of type 2 diabetes susceptibility observed in genome-wideassociation data. Diabetes. 2009;58(2):505-510. DOI 10.2337/db08-0906.Trifonova E.A., Popovich A.A., Bocharova A.V., Vagaitseva K.V.,Stepanov V.A. The role of natural selection in the formation of thegenetic structure of populations by SNP markers in association withbody mass index and obesity. Mol. Biol. 2020;54(3):349-360. DOI10.1134/S0026893320030176.Tsyretorova S.S., Bardymova T.P., Protasov K.V., Donirova O.S.,Mistyakov M.V. Ethnic features of diabetes mellitus and coronaryheart disease. Sibirskiy Meditsinskiy Zhurnal (Irkutsk) = SiberianMedical Journal (Irkutsk). 2015;136(5):15-21. (in Russian)Vikulova O.K., Zheleznyakova A.V., Lebedeva N.O., Nikitin A.G.,Nosikov V.V., Shestakova M.V. Genetic factors in the developmentof kidney chronic disease in patients with diabetes mellitus. Russ.J. Genet. 2017;53(4):420-432. DOI 10.1134/S1022795417030140.Xi B., Takeuchi F., Meirhaeghe A., Kato N., Chambers J.C., MorrisA.P., Cho Y.S., Zhang W., Mohlke K.L., Kooner J.S., Shu X.O.,Pan H., Tai E.S., Pan H., Wu J.Y., Zhou D., Chandak G.R., DIAGRAMConsortium, AGEN-T2D Consortium, SAT2D Consortium.Associations of genetic variants in/near body mass index-associatedgenes with type 2 diabetes: a systematic meta-analysis. Clin. Endocrinol.(Oxf .). 2014;81(5):702-710. DOI 10.1111/cen.12428.Zhang M., Tang M., Fang Y., Cui H., Chen S., Li J., Xiong H., Lu J.,Gu D., Zhang B. Cumulative evidence for relationships betweenmultiple variants in the VTI1A and TCF7L2 genes and cancer incidence.Int. J. Cancer. 2018;142(3):498-513. DOI 10.1002/ijc.31074."}
+{"text": "Scientific Reports 10.1038/srep11362, published online 12 June 2015Retraction of: Fig. 1e HFD/p-Src lane 1 and 3;Fig. 4c NCD/Ve-cadherin lane 1 and 3Fig. 5e HFD+\u2009APN/ICAM-1 lane 1 and 2Fig. 5f HFD/beta-catenin lane 2 and HFD+\u2009APN/beta-catenin lane 1Fig. S1d HFD/beta-catenin all lanesFig. S4c NCD/beta-catenin lane 1 and 3.The Authors have retracted this Article. After publication of this Article, concerns have been raised about irregularities in the western blot data. In particular, the following bands appear to be duplicated:Additionally, the beta-catenin subpanel in Fig. 5f was subsequently reused in another study  and descDilip Shah, Nadan Wang, Benjamin T. Suratt, Jianxin Sun, Ying Zhu, Kenneth Walsh and Ross Summer agree to this retraction. Freddy Romero, Bishnuhari Paudyal and Caleb B. Kallen have not responded to any correspondence from the editor or publisher about this retraction. The Publisher has not been able to obtain a current email address for Michelle Duong."}
+{"text": "Scientific Reports 10.1038/s41598-021-84517-x, published online 04 March 2021Correction to: The original version of this Article contained an error in Reference 32, which was incorrectly given as:et al.\u00a0Exogenic basalt on asteroid (101955) Bennu.\u00a0Nat. Astron.\u00a0https://doi.org/10.1038/s41550-020-1195-z\u00a0(2020).DellaGiustina, D. N.\u00a0The correct reference is listed below:et al. Variations in color and reflectance on the surface of asteroid (101955) Bennu. Science370, abc3660. https://doi.org/10.1126/science.abc3660 (2020).DellaGiustina, D. N. The original Article has been corrected."}
+{"text": "T and H16/1AT, of Gram-stain-positive, coagulase-negative staphylococci were isolated from separate healthy domestic dogs in Scotland. Both strains were genome sequenced and their inferred DNA\u2013DNA hybridisation indicates that H8/1T and H16/1AT represent two novel species of the genus Staphylococcus. On the basis of the results of genome sequence analysis  H8/1T is most closely related to Staphylococcus devriesei and H16/1AT most closely related to Staphylococcus felis. Also, average nucleotide identity distinguished H8/1T and H16/1AT from S. devriesei and S. felis as did minor phenotypic differences. On the basis of these results, it is proposed that H8/1T and H16/1AT represent novel species with the respective names Staphylococcus caledonicus and Staphylococcus canis. The type strain of S. caledonicus is H8/1T (=NCTC 14452T=CCUG 74789T). The type strain of S. canis is H16/1AT (=NCTC 14451T=CCUG 74790T)Two strains, H8/1 Staphylococcus  before plating onto mannitol salt agar (Oxoid) and then incubation at 37\u2009\u00b0C for 24\u2009h. Both isolates are Gram-stain-positive, catalase-positive cocci and were whole-genome sequenced using HiSeq technology (Illumina) with 2\u00d7250\u2009bp paired-end reads, read trimming and assembly . Reads were trimmed using Trimmomatic version 0.30 [de novo using SPAdes version 3.7 [https://tygs.dsmz.de/) [Staphylococcus. The TYGS phylogenetic tree generated by the Genome blast Distance Phylogeny approach (GBDP) indicated that H8/1T is most closely related to, but distinct from Staphylococcus devriesei CCUG 58238T with H16/1AT most closely related to, but distinct from Staphylococcus felis DSM 7377T  values calculated with the Type (Strain) Genome Server using the recommended settings of the Genome-to-Genome Distance Calculator (GGDC) 2.1 [blast (ANIb) [S. caledonicus H8/1T with S. devriesei CCUG 58238T. Divergence between ANI and tetranucleotide signature correlation index has been described previously with a possible explanation being that evolutionary or environmental forces may impede modifications in this genome signature despite genetic drift occurring [Staphylococcus was generated using CSI Phylogeny 1.4 [Staphylococcus aureus DSM 20231T as the reference genome, this analysis produced a single-nucleotide polymorphism tree comprising 19637 nucleotide positions  Genome Server (TYGS) (smz.de/) . The resSM 7377T . A 16S rS. felis . Althougparisons . Indeed,GDC) 2.1  and by At (ANIb) , MUMmer t (ANIb)  and the t (ANIb)  (Table 1t (ANIb) . With ont (ANIb)   according to the manufacturer\u2019s instructions alongside the type strains of S. devriesei DSM 25293T and S. felis DSM 7377T  and for DNAse activity using DNAse agar (Oxoid). In each case, both H8/1T and H16/1AT tested negative for these activities.Phenotypic characterisation of H8/1SM 7377T . H8/1T i. felis, . AdditioT and H16/1AT were susceptible to all the antimicrobials tested, which were as follows: amoxicillin/clavulanic acid, benzylpenicillin, cefovecin, cefoxitin (screen), ceftiofur, chloramphenicol, clindamycin, doxycycline, enrofloxacin, erythromycin, gentamicin, inducible clindamycin resistance, kanamycin, marbofloxacin, neomycin, nitrofurantoin, oxacillin, pradofloxacin, tetracycline and trimethoprim/sulfamethoxazole. No known antimicrobial resistance genes (perfect and strict hits) were identified in either H8/1T and H16/1AT on using The Comprehensive Antibiotic Resistance Database (CARD) (https://card.mcmaster.ca/) [Antimicrobial sensitivity testing was performed using the Vitek2 system (bioM\u00e9rieux) according to the manufacturer\u2019s instructions. Using the AST-GP80 card and applying the CLSI 2017 interpretations for coagulase-negative staphylococci, both H8/1ter.ca/) .Staphylococcus caledonicus , the country where the type strain was isolated).d-glucose, d-fructose, maltose, trehalose, d-mannitol and sucrose but not from d-mannose, lactose, xylitol, melibiose, raffinose, d-xylose, methyl \u03b1-d-glucopyranoside or N-acetylglucosamine. Has arginine dihydrolase activity and is able to reduce nitrates to nitrites. Catalase-positive and negative for clumping factor, coagulase and DNAse.Gram-stain-positive, non-spore forming, facultative anaerobe, forms non-pigmented, smooth, circular colonies about 1\u20132\u2009mm in diameter with entire margins on Columbia horse blood agar after 18\u2009h incubation at 37\u2009\u00b0C. Able to produce acid from S. caledonicus, H8/1T (=NCTC 14452 T=CCUG 74789T), was isolated from a healthy dog in Scotland during 2018. The draft genome of H8/1T is 2\u200a503\u200a367 bases in length with a DNA G+C\u2009content of 33.6\u2009mol%, it comprises 38 contigs with an average coverage of approximately 85-fold. The genome sequence data from H8/1T is available under these accession numbers: BioSample, SAMN15065541; Sequence Read Archive, SRR11909362; and assembly, JABTXV000000000.The type strain of Staphylococcus canis .d-glucose, d-fructose, d-mannose, maltose, lactose, trehalose and d-mannitol but not from xylitol, melibiose, raffinose, d-xylose, sucrose or methyl \u03b1-d-glucopyranoside. Has alkaline phosphatase and urease activity and is able to reduce nitrates to nitrites. Catalase-positive and negative for clumping factor, coagulase and DNAse.Gram-stain-positive, non-spore forming, facultative anaerobe, forms non-pigmented, smooth, circular colonies about 1\u20132\u2009mm in diameter with entire margins on Columbia horse blood agar after 18\u2009h incubation at 37\u2009\u00b0C. Able to produce acid from Staphylococcus canis, H16/1AT (=NCTC 14451T=CCUG 74790T), was isolated from a healthy dog in Scotland during 2018. The draft genome of H16/1AT is 2\u200a229\u200a149 bases in length with a DNA G+C\u2009content of 34.8\u2009mol%, it comprises 143 contigs with an average coverage of approximately 92-fold. The genome sequence data from Staphylococcus canis H16/1AT is available under these accession numbers: BioSample, SAMN14548534; Sequence Read Archive, SRR11498036; and assembly, JABANU000000000.The type strain of Click here for additional data file."}
+{"text": "None showed focal spasm that exclusively involved DCB-treated lesions. Among 27 patients with vasospastic features, DCB-treated segments showed less vasoconstriction than spastic counterparts (p < 0.001). A total of 110 DCB-treated lesions were analyzed to assess vasomotor function. Vasomotor function, defined as a combined constrictor and dilator response, was comparable between DCB-treated and angiographically normal segments (p = 0.173), while significant differences were observed against spastic counterparts (p < 0.001). In our study, DCB-treated lesions were not particularly vulnerable to vasospasm and were found to have vasomotor function similar to angiographically normal segments, supporting safety of DCB-only strategy in treating de novo native coronary lesions.Balloon-injured coronary segments are known to harbor abnormal vasomotion. We evaluated whether de novo coronary lesions treated using drug-coated balloon (DCB) are prone to vasospasm and how they respond to ergonovine and nitrate. Among 132 DCB angioplasty recipients followed, 89 patients underwent ergonovine provocation test at 6\u20139 months follow-up. Within-subject ergonovine- and nitrate-induced diameter changes were compared among three different sites: DCB-treated vs. angiographically normal vs. segment showing prominent vasoreactivity (spastic). No patient experienced clinically refractory vasospastic angina or symptom-driven revascularization during follow-up. Ergonovine induced vasospasm in seven patients; all were multifocal spasm either involving ( Drug-coated balloon (DCB) angioplasty is an emerging treatment approach for obstructive coronary artery disease (CAD). It has demonstrated its safety and efficacy in the treatment of in-stent restenosis and small vessel CAD and, currently, many on-going trials are exploring its value in a broader range of lesions ,2,3. ThePercutaneous transluminal coronary angioplasty (PTCA), including plain old balloon angioplasty (POBA) or stenting, unavoidably results in significant vascular injury, leading to a variety of deleterious consequences, such as dissection, thrombus formation, impairment of physiologic vasomotion, and restenosis . In partThis prospective all-comers study evaluated all patients with significant CAD who received percutaneous coronary intervention (PCI) with DCB-only angioplasty, concomitant DCB angioplasty and POBA, or concomitant DCB angioplasty and drug-eluting stent (DES) implantation for de novo lesions in the native coronary arteries at the Korea University Ansan Hospital between July 2019 and December 2020. The patients with a life expectancy shorter than 2 years and those who refused to provide their consent were excluded. DCB angioplasty was performed in accordance with the recent recommendations . SpecifiThe presence of significant vasospasm and coronary artery vasomotor function were assessed via intracoronary injection of ergonovine and isosorbide dinitrate . All par-Constrictor response (%):  after ergonovine administration\u2014MLD after nitrate administration)/MLD after nitrate administration \u00d7 100-Dilator response (%): (MLD after nitrate administration\u2014MLD at baseline)/MLD at baseline \u00d7 100Coronary spasm was defined as transient, total, or near-total occlusion (>90%) of a coronary artery as recommended . Spasm wThe reference vessel diameter (RVD), measured at the proximal and distal portions of each segment of interest, was used to calculate the diameter stenosis (DS) for quantitative comparative analysis. Segments with a percent change of >50% after the ergonovine provocation test were considered to have significant vasoconstriction, and patients who had at least one or more coronary segments showing significant vasoconstriction (>50%) were considered to have vasospastic features.Vasomotor response to ergonovine and isosorbide dinitrate was compared among three different segments: DCB-treated segments, angiographically normal arterial segments (control), and segments showing prominent vasoconstrictive response to ergonovine (spastic) . The cont-test. Ergonovine-induced percent diameter changes (constrictor response) between DCB-treated vs. spastic segments were compared using paired t-test or Wilcoxon signed-rank test based on normality test. Differences in combined constrictor and dilator responses across the three segments were analyzed using general linear model two-way repeated measures analysis of variance (ANOVA). Repeated measures ANOVA incorporated the Bonferroni test for pair-wise comparison. Statistical analyses were performed using the SPSS software . Statistical significance was set at p-values of <0.05.Continuous variables are expressed as means \u00b1 standard deviations and dichotomous variables as counts and percentages. Differences in MLD between baseline and after drug administration were analyzed using a paired A total of 132 patients who underwent DCB angioplasty for de novo native coronary lesions were analyzed. Clinical characteristics of the study population is described in During the median follow-up period of 18.5 months, two patients underwent repeated PCI due to recurrent ACS; one patient presented with unstable angina due to a de novo stenotic lesion in the treated vessel (2 months post-PCI); and one patient presented with non-ST elevation myocardial infarction due to post-POBA restenosis of the proximal right coronary artery but whose DCB-treated proximal left anterior descending artery remained patent. Albeit asymptomatic, three patients underwent redo DCB angioplasty owing to post-DCB angioplasty restenosis and one patient underwent DCB angioplasty owing to a newly-developed lesion at the follow-up angiography. One patient complained a sublingual nitroglycerin-responsive, alcohol-related nocturnal chest pain during the follow-up. His symptom was controlled by prescribing calcium channel blocker and oral nitrates, and he later showed ergonovine-induced multifocal vasospasm at the follow-up angiography . In summAmong 132 enrollees, patients who underwent 6\u20139 months of follow-up invasive coronary angiography and those excluded for the ergonovine provocation test are summarized in the CONSORT flow diagram in p = 0.069; DS (%): 27.64 \u00b1 10.07 vs. 31.46 \u00b1 13.64, p = 0.498]. However, ergonovine-induced constrictor response (%) was significantly lower in the DCB-treated segments than in the spastic segments : 24.19 \u00b1 0.97 vs. 24.66 \u00b1 12.53, p = 0.845).In seven patients with documented vasospasm, RVD and DS were similar between eight DCB-treated segments and their spastic counterparts [DCB-treated vs. spastic: RVD (mm): 2.98 \u00b1 0.56 vs. 2.61 \u00b1 0.37, p < 0.001; DS (%): 12.72 \u00b1 7.14 vs. 24.21 \u00b1 8.59 vs. 17.56 \u00b1 10.45, p < 0.001). DCB-treated segments showed similar RVD (p > 0.999), but greater DS compared to spastic segments (p < 0.001).A total of 274 segments  from 82 patients were analyzed after excluding those from seven vasospasm cases. Angiographic characteristics of the analyzed segments are presented in p < 0.001; DCB-treated: 1.90 \u00b1 0.42 vs. 1.58 \u00b1 0.41, p < 0.001; spastic: 1.94 \u00b1 0.46 vs. 1.37\u00b1 0.44, p < 0.001, p < 0.001; DCB-treated: 1.90 \u00b1 0.42 vs. 2.06 \u00b1 0.40, p < 0.001; spastic: 1.94 \u00b1 0.46 vs. 2.22 \u00b1 0.47, p < 0.001, After ergonovine administration, most segments showed significant constriction [MLD (mm): baseline vs. ergonovine: control: 2.54 \u00b1 0.57 vs. 2.20 \u00b1 0.57, Intriguingly, some DCB-treated segments were found to be unaffected by ergonovine. 35.5% (39 out of 110) of DCB-treated segments had a diameter of at least 20% greater than the adjacent arterial portions after ergonovine administration . These cp < 0.001, p = 0.173), while these were significantly greater in the spastic segments than in the DCB-treated and control segments (control vs. spastic: p < 0.001; DCB-treated vs. spastic: p < 0.001, Vasomotor function, as comprehensively assessed as a combined constrictor and dilator response, was analyzed using repeated measure ANOVA. Within-subject difference in the ergonovine- and nitrate-induced serial percent diameter changes was statistically significant across the three segments (ergonovine vs. nitrate: control: \u221218.78 \u00b1 10.45 vs. 7.79 \u00b1 8.48; DCB-treated: \u221222.95 \u00b1 15.57 vs. 9.36 \u00b1 10.73; spastic: \u221237.03 \u00b1 17.56 vs. 15.55 \u00b1 15.19, Coronary artery spasm is not only a functional abnormality of the coronary artery, causing transient, reversible myocardial ischemia, but it indeed plays a role in the pathogenesis of ACS . A previThe introduction of metallic stents was a major breakthrough in overcoming the limitations of PTCA, such as abrupt vessel closure, acute recoil, dissection, and vasospasm. However, permanent vessel caging by metallic implants constitutes barriers to physiological vasomotion and expansile vessel remodeling. Further, DESs implantation produces prolonged endothelial dysfunction in stented artery  and, as The mechanisms underlying the observed favorable findings may be multifactorial. Ergonovine, an ergot derivative, exerts vasoconstrictive effects by stimulating alpha-adrenergic and serotonin receptors  and inhiThis study is limited by a relatively small population. Angiographically normal segments, not evidenced by intravascular imaging, may harbor atherosclerosis. Unlike acetylcholine affecting both the endothelium and vascular smooth muscle, the ergonovine effect primarily represents endothelium-independent constriction . Thus, oAbnormal coronary vasomotion, as invasively assessed using intracoronary vasoactive substances, is an independent prognosticator for adverse cardiovascular events ,30. Thus"}
+{"text": "Muti HS, Heij LR, Keller G, et al. Development and validation of deep learning classifiers to detect Epstein-Barr virus and microsatellite instability status in gastric cancer: a retrospective multicentre cohort study. Lancet Digit Health 2021; published online Aug 17. https://doi.org/10.1016/S2589-7500(21)00133-3\u2014In figure 2A of this Article, the colours were missing. This correction has been made as of Aug 19, 2021."}
+{"text": "Streptomyces sp. HO1518. A total of ninety-eight aminooligosaccharides, including eighty potential new compounds, were detected mainly based on the characteristic fragment ions originating from quinovosidic bond cleavages in their molecules. Following an LC-MS-guided separation technique, seven new aminooligosaccharides (10\u201316) along with four known related compounds (17\u201320) were obtained directly from the crude extract of strain HO1518. Compounds 10\u201313 represent the first examples of aminooligosaccharides with a rare acarviostatin II02-type structure. In addition, all isolates displayed considerable inhibitory effects on three digestive enzymes, which revealed that the number of the pseudo-trisaccharide core(s), the feasible length of the oligosaccharides, and acyl side chain exerted a crucial influence on their bioactivities. These results demonstrated that the UPLC-QTOF-MS/MS-based metabolomics approach could be applied for the rapid identification of aminooligosaccharides and other similar structures in complex samples. Furthermore, this study highlights the potential of acylated aminooligosaccharides with conspicuous \u03b1-glucosidase and lipase inhibition for the future development of multi-target anti-diabetic drugs.A rapid and sensitive method using ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS) was applied for the analysis of the metabolic profile of acarviostatin-containing aminooligosaccharides derived from  Acarbose, a typical anti-diabetes drug functioning as an \u03b1-glucosidases inhibitor, potently inhibits the \u03b1-glucosidases in vivo to retard carbohydrate digestion and avoid blood glucose elevation +; HRESIMS: m/z 1273.4938 [M + H]+ .Acarviostatin II02 : White amorphous powder, c 1.01, H2O). UV (H2O) end absorption; IR \u03bdmax 3337, 1726, 1407, 1149, 1026 cm\u22121. 1H (500 MHz) and 13C (125 MHz) NMR spectroscopic data, see m/z 1329 [M + H]+; HRESIMS: m/z 1329.5234 [M + H]+ .D6-O-Isobutyryl-acarviostatin II02 : White amorphous powder, c 1.02, H2O). UV (H2O) end absorption; IR \u03bdmax 3329, 1721, 1365, 1147, 1014 cm\u22121. 1H (500 MHz) and 13C (125 MHz) NMR spectroscopic data, see m/z 1343 [M + H]+; HRESIMS: m/z 1343.5365 [M + H]+ .D6-O-Isovaleryl-acarviostatin II02 : White amorphous powder, c 0.99, H2O). UV (H2O) end absorption; IR \u03bdmax 3305, 1647, 1407, 1150, 1013 cm\u22121. 1H (500 MHz) and 13C (125 MHz) NMR spectroscopic data, see m/z 1357 [M + H]+; HRESIMS: m/z 1357.5520 [M + H]+ .D6-O-Propionyl-acarviostatin II03 : White amorphous powder, c 0.90, H2O). UV (H2O) end absorption; IR \u03bdmax 3304, 1734, 1402, 1148, 1023 cm\u22121. 1H (500 MHz) and 13C (125 MHz) NMR spectroscopic data, see m/z 1491 [M + H]+; HRESIMS: m/z 1491.5727 [M + H]+ .D6-O-Butyryl-acarviostatin II03 : White amorphous powder, c 0.53, H2O). UV (H2O) end absorption; IR \u03bdmax 3324, 1729, 1568, 1149, 1024 cm\u22121. 1H (500 MHz) and 13C (125 MHz) NMR spectroscopic data, see m/z 1505 [M + H]+; HRESIMS: m/z 1505.5872 [M + H]+ .D6-O-2-Methyl-butyryl-acarviostatin II03 : White amorphous powder, c 1.03, H2O). UV (H2O) end absorption; IR \u03bdmax 3303, 1645, 1406, 1149, 1014 cm\u22121. 1H (500 MHz) and 13C (125 MHz) NMR spectroscopic data, see m/z 1519 [M + H]+; HRESIMS: m/z 1519.6039 [M + H]+ .D6-9\u201320 were conducted based on a previously reported method. Commercial \u03b1-amylase inhibitor acarbose was used as the positive control [The PPA inhibitory activities of compounds 9\u201320 was evaluated according to the previously reported method. Acarbose was also used as the positive control [The sucrase inhibition assay of compounds  control .9\u201320 was performed according to the method outlined by McDougall et al. with slight modification [The lipase inhibition assay of compounds fication . Orlistax: 4.342 \u00c5; y: 24.299 \u00c5; z: 47.471 \u00c5) and the grid box dimensions (30 \u00d7 30 \u00d7 30 \u00c5) were set. The results of molecular docking were evaluated on the basis of the binding energy, criteria of binding structure, and possible interactions between ligand and the critical catalytic triad of protein 1LPB.The molecular docking simulations were performed by AutoDock Vina software, version 1.5.7 . The cryStreptomyces sp. HO1518. A total of ninety-eight aminooligosaccharides, including eighty new compounds, were detected and characterized from the extract of stain HO1518. Among them, twenty structural intriguing oligomers that ended with the 4-amino-4-deoxy-D-quinovopyranose unit at the non-reducing terminus were reported for the first time. The subsequent MS-guided fractionation method resulted in the isolation of seven new oligosaccharides (10\u201316) and four known analogs (17\u201320). Notably, compounds 10\u201313 are the first reported examples of oligosaccharides with a rarely occurring acarviostatin II02-type structure. All the compounds exhibited significant inhibitory activities against three digestive enzymes, among which compounds 9\u201316, 19 and 20 sharing two pseudo-trisaccharides were the most effective inhibitors of \u03b1-amylase and lipase. Furthermore, primary structure-activity relationships of 9\u201320 revealed that the number of the pseudo-trisaccharide core and acyl side chain play pivotal roles in their biological activity, which was evidenced by molecular docking analysis. These results of this study highlighted the advantages of UPLC-QTOF-MS/MS for the rapid structural identification of oligosaccharides, and this strategy could be extended to other investigations for high-throughput analysis of natural products with similar structures. More importantly, this study not only provided new lead compounds for further scientific research towards anti-diabetic drug discovery, but also shed light on the structural optimization of aminooligosaccharides analogs for medicinal scientists.In summary, the hyphenated system UPLC-QTOF-MS/MS that could provide qualitative retention time and reliable mass spectrometry information was utilized to reveal the metabolic profiling of aminooligosaccharides secreted by"}
+{"text": "Burkholderia spp. is an opportunistic pathogen associated with respiratory infections. Cefiderocol (CFDC), a siderophore cephalosporin approved in US and EU, is active in vitro against carbapenem-resistant Gram-negative bacteria including Burkholderia spp. This study examined in vitro and in vivo activity of CFDC against Burkholderia spp.Burkholderia spp. collected in 2014 - 2019 in 13 countries were determined by broth microdilution method according to CLSI guidelines. Only for CFDC, iron-depleted CAMHB was used. In a rat lung infection model, B. cepacia ATCC 25416  was used. Male CD  rats were infected by intrabronchial inoculation of the bacterial suspension including 1% nutrient agar. The humanized PK in plasma by administration of CFDC 2 g every 8 h (3-h infusion) and MEM 1 g every 8 h (0.5-h infusion) were recreated via the continuous intravenous infusion for 4 days, and the viable cfu in lungs were counted.MICs of CFDC and 13 marketed antibacterial drugs against 462 clinical isolates of 50/MIC90 of \u2264 0.031/1 \u00b5g/mL, which was the lowest among the tested antibiotics. Among 185 MEM non-susceptible isolates, 94% of the isolates exhibited \u2264 4 \u00b5g/mL of CFDC MIC. In a rat lung infection model, CFDC and MEM showed bactericidal activity with 2.8 and 2.4 log10 CFU/lung decrease compared with non-treated control, respectively. By recreating the humanized PK exposure in this model, 100% and ca.35% of fT >MIC of CFDC and MEM in plasma has been achieved, respectively. The bactericidal activities of both compounds vs B. cepacia ATCC 25416 would be reasonable because the fT >MIC achieved in this model exceeds the target fT >MIC  required to cause 1 log10 reduction in murine thigh infection models1,2). Against 462 strains, including 185 MEM non-susceptible isolates, CFDC showed MIC1) M. Sabet. 2019. AAC 2) R. Nakamura. 2019. AACIn vitro activity of CFDC and comparator agents against Burkholderia spp.Burkholderia spp. In critically ill patients, the recommended dosing regimen achieves 100% of fT >MIC of \u2264 4 ug/mL3).3) N. Kawaguchi. 2021. AACCFDC has potential for treating respiratory tract infections caused by Merime Oota, BSc, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Toriko Yoshitomi, -, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Rio Nakamura, BSc, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Miki Takemura, MS, SHIONOGI & CO., LTD. (Employee) Yoshinori Yamano, PhD, Shionogi (Employee) Meredith Hackel, PhD MPH, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "Hospitalized COVID-19 patients tend to be older and frequently have hypertension, diabetes or CHD, but whether these co-morbidities are more common than in the general older population is unclear. We estimated associations between pre-existing diagnoses and hospitalized COVID-19 alone or with mortality . In 269,070 UK Biobank participants aged 65+, 507 (0.2%) became COVID-19 hospital inpatients, of which 141 (27.8%) died. Common preexisting co-morbidities in hospitalized inpatients were hypertension (59.6%), history of falls/fragility fractures (29.4%), CHD (21.5%), T2 diabetes (19. 9%) and asthma (17.6%). However, in adjusted models, pre-existing diagnoses of dementia, T2 diabetes, COPD, pneumonia, depression, atrial fibrillation and hypertension emerged as independent risk factors for COVID-19 hospitalization, the first five remaining statistically significant for related mortality. There are specific high risk pre-existing co-morbidities for COVID-19 hospitalization and deaths in community based older men and women."}
+{"text": "The dependences of the crystal structure of CrAs on temperature (30\u2013400\u2005K) and on pressure (0\u20139.46\u2005GPa) are reported. The investigation shows twinning accompanying the temperature-induced isosymmetrical phase transition and the existence of a distinct Cr\u2013Cr distance within the structure showing anomalous behavior. TN = 267\u2005K can induce a change in the microstructure by twinning due to a crossing of the orthohexagonal setting of the unit-cell parameter ratio c/b. Within the crystal structure, one particular Cr\u2013Cr distance exhibits anomalous behavior in that it is nearly unaffected by temperature and pressure in the paramagnetic phase, which is stable above 267\u2005K and at high pressures. The distinction of this shortest Cr\u2013Cr distance might be of importance for the superconducting properties of CrAs.The crystal structure of CrAs was investigated using synchrotron X-ray single-crystal diffraction for separate dependences on temperature (30\u2013400\u2005K) and on pressure (0\u20139.46\u2005GPa). The isosymmetrical magnetostructural phase transition at  The iderk Fig. 1.aCmcm, with a\u2032 = ahex, b\u2032\u00a0= c\u2032 = chex. A slight displacement of the atomic positions leads to the loss of the C-centering and the symmetry reduction to space group Pmcn following the P63/mmc\u2192Cmcm\u2192Pmcn group\u2013subgroup relation  are of second order, the low-temperature transitions in MnP (from the ferro- to the helimagnetic state) and in CrAs are of first order and accompanied by abrupt changes in the unit-cell parameters, though the precise changes in MnP are controversial  at about 33\u2005GPa at very low temperatures   on the kappa diffractometer  equipped with a Pilatus CdTe 1M area detector.For the measurements in the range 35\u2013275\u2005K at ambient pressure, the temperatures were set using a Cryocool G2b-LT helium gas jet cryostat.et al., 1986The high-pressure measurements to 9.5\u2005GPa at room temperature were carried out using several diamond anvil cells, each pre-loaded with a CrAs single crystal, a ruby chip, and a 4:1 methanol\u2013ethanol mixture as pressure-transmitting medium. The pressure during all measurements in diamond anvil cells was determined using the ruby luminescence method  showed no significant difference in the overall quality of the fit. However, refinement of the parameters yCr and yAs, which are fixed to \u00bc in Pnma but free in Pn21a, show that within the errors they do not deviate from the centrosymmetric value \u00bc in the whole temperature range. This observation is similar to the findings of Selte & Kjekshus , the observed diffraction patterns exhibit extinction rules that are in accordance with space groups us 1973b regardin3.2.aTN, the structural phase transition is clearly visible from the large abrupt changes of all unit-cell parameters. The relative changes on cooling \u0394a/a \u2243 \u22120.40%, \u0394b/b \u2243 +3.51%, \u0394c/c \u2243 \u22120.83% and \u0394V/V \u2243 +2.25% confirm the previous observations from the literature. Below the transition temperature, the unit-cell parameters change smoothly; a and c decrease in a comparable way, b first increases and eventually stays nearly constant below about 200\u2005K, where due to the lack of further change in b, the volume V is dominated by the behavior of a and c \u2005\u00c5, b = 3.467\u2005(2)\u2005\u00c5, c = 6.200\u2005(3)\u2005\u00c5 and V = 121.4\u2005(1)\u2005\u00c53. The fitted unit-cell volume at zero pressure, the bulk modulus and the first derivative of the bulk modulus are V0 = 121.3\u2005(2)\u2005\u00c53, B0 = 28\u2005(4)\u2005GPa and B0\u2032 = 36\u2005(5), respectively. Compared to other MnP-type compounds [e.g. MnP: B0 = 116\u2005(12)\u2005GPa, B0\u2032 = 4.2\u2005(8) \u2005GPa, B0\u2032 = 4 \u2005GPa, B0\u2032 = 8.8\u2005(33) . In addition to the standard refinements with harmonic atomic displacement parameters (ADPs), refinements with anharmonic ADPs were carried out, as this was suggested by a visual inspection of the Fobs density maps. The resulting parameters show consistently that around the transition temperature the displacement parameters of the atoms show a significant anharmonicity, indicating that around and during the phase transition, the atoms show increased and anharmonic movement around their mean positions or that there is an increased degree of static disorder of the atoms which are slightly moved away from their highly symmetrical special positions.This formation of twin domains during the phase transition is furthermore coupled to increased displacement parameters of the atoms , as the Cr atoms carry the magnetic moment and they and their interactions are of particular importance for the magnetic and eventually superconducting properties. Above the phase transition, the distances CrI\u2013CrI and CrI\u2013CrIV follow the general trend and increase with temperature. However CrI\u2014CrII, the strongest covalent Cr\u2014Cr bond and strongest homoatomic interaction, is remarkable as it shows anomalous behavior in that it stays constant above TN. At the phase transition, the same distinction between the three Cr\u2013Cr distances is seen. Here, CrI\u2013CrI and CrI\u2013CrIV increase (upon cooling) due to the phase transition, while CrI\u2013CrII, the shortest Cr\u2013Cr distance, decreases even further. CrI\u2013CrI directly determines the length of the b axis of the unit cell, so that its increase is equivalent to the observed discontinuity in the temperature dependence of the b unit-cell parameter. CrI\u2013CrIV, lying almost in the bc plane, increases as well. The decrease in CrI\u2013CrII, which is oriented approximately along the a axis, leads to the abrupt shortening of this axis. Besides this decrease in the distance CrI\u2013CrII, the transition is also accompanied by a slight straightening of the CrI\u2013CrII\u2013CrI chains  and the \u2220CrI\u2013CrII\u2013CrI angle gets closer to the ideal value of 180\u00b0. CrI\u2013CrII thus shows two distinctions: decreasing at TN and staying constant above TN. Below the transition, CrI\u2013CrII and CrI\u2013CrIV decrease with temperature, while CrI\u2013CrI increases and is eventually constant below about 200\u2005K.The most interesting distances in the CrAs structure are the Cr\u2013Cr distances  \u2013 which decrease during the phase transition .In CrAs on compression, all Cr\u2013As and Cr\u2013Cr distances decrease with the exception of CrII Fig. 7, which ins Fig. 8 and the 4.et al.  and also in good agreement with those reported by Selte et al.  with the MnP-type structure shows that at least the behavior of the relevant interatomic T\u2013T distances on compression are similar  global, Synchrotron-275K, Synchrotron-270K, Synchrotron-260K, Synchrotron-250K, Synchrotron-240K, Synchrotron-220K, Synchrotron-200K, Synchrotron-185K, Synchrotron-170K, Synchrotron-155K, Synchrotron-140K, Synchrotron-125K, Synchrotron-110K, Synchrotron-95K, Synchrotron-80K, Synchrotron-65K, Synchrotron-50K, Synchrotron-35K, Synchrotron-1.03GPa, Synchrotron-2.53GPa, Synchrotron-3.13GPa, Synchrotron-4.45GPa, Synchrotron-6.05GPa, Synchrotron-7.32GPa, Synchrotron-8.09GPa, Synchrotron-9.46GPa, Lab-400K, Lab-390K, Lab-380K, Lab-370K, Lab-360K, Lab-350K, Lab-340K, Lab-330K, Lab-320K_02, Lab-320K_01, Lab-310K_02, Lab-310K_01, Lab-300K_02, Lab-300K_01, Lab-290K, Lab-280K_02, Lab-280K_01, Lab-275K, Lab-270K, Lab-250K, Lab-240K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-275Ksup2.hklStructure factors: contains datablock(s) Synchrotron-275K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-270Ksup3.hklStructure factors: contains datablock(s) Synchrotron-270K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-260Ksup4.hklStructure factors: contains datablock(s) Synchrotron-260K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-250Ksup5.hklStructure factors: contains datablock(s) Synchrotron-250K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-240Ksup6.hklStructure factors: contains datablock(s) Synchrotron-40K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-220Ksup7.hklStructure factors: contains datablock(s) Synchrotron-220K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-200Ksup8.hklStructure factors: contains datablock(s) Synchrotron-200K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-185Ksup9.hklStructure factors: contains datablock(s) Synchrotron-185K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-170Ksup10.hklStructure factors: contains datablock(s) Synchrotron-170K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-155Ksup11.hklStructure factors: contains datablock(s) Synchrotron-155K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-140Ksup12.hklStructure factors: contains datablock(s) Synchrotron-140K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-125Ksup13.hklStructure factors: contains datablock(s) Synchrotron-125K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-110Ksup14.hklStructure factors: contains datablock(s) Synchrotron-110K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-95Ksup15.hklStructure factors: contains datablock(s) Synchrotron-95K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-80Ksup16.hklStructure factors: contains datablock(s) Synchrotron-80K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-65Ksup17.hklStructure factors: contains datablock(s) Synchrotron-65K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-50Ksup18.hklStructure factors: contains datablock(s) Synchrotron-50K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-35Ksup19.hklStructure factors: contains datablock(s) Synchrotron-35K. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-1.03GPasup20.hklStructure factors: contains datablock(s) Synchrotron-1.03GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-2.53GPasup21.hklStructure factors: contains datablock(s) Synchrotron-2.53GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-3.13GPasup22.hklStructure factors: contains datablock(s) Synchrotron-3.13GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-4.45GPasup23.hklStructure factors: contains datablock(s) Synchrotron-4.45GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-6.05GPasup24.hklStructure factors: contains datablock(s) Synchrotron-6.05GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-7.32GPasup25.hklStructure factors: contains datablock(s) Synchrotron-7.32GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-8.09GPasup26.hklStructure factors: contains datablock(s) Synchrotron-8.09GPa. DOI: 10.1107/S2052520621005655/xk5084Synchrotron-9.46GPasup27.hklStructure factors: contains datablock(s) Synchrotron-9.46GPa. DOI: 10.1107/S2052520621005655/xk5084Lab-400Ksup28.hklStructure factors: contains datablock(s) Lab-400K. DOI: 10.1107/S2052520621005655/xk5084Lab-390Ksup29.hklStructure factors: contains datablock(s) Lab-390K. DOI: 10.1107/S2052520621005655/xk5084Lab-380Ksup30.hklStructure factors: contains datablock(s) Lab-380K. DOI: 10.1107/S2052520621005655/xk5084Lab-370Ksup31.hklStructure factors: contains datablock(s) Lab-370K. DOI: 10.1107/S2052520621005655/xk5084Lab-360Ksup32.hklStructure factors: contains datablock(s) Lab-360K. DOI: 10.1107/S2052520621005655/xk5084Lab-350Ksup33.hklStructure factors: contains datablock(s) Lab-350K. DOI: 10.1107/S2052520621005655/xk5084Lab-340Ksup34.hklStructure factors: contains datablock(s) Lab-340K. DOI: 10.1107/S2052520621005655/xk5084Lab-330Ksup35.hklStructure factors: contains datablock(s) Lab-330K. DOI: 10.1107/S2052520621005655/xk5084Lab-320K_02sup36.hklStructure factors: contains datablock(s) Lab-320K_02. DOI: 10.1107/S2052520621005655/xk5084Lab-320K_01sup37.hklStructure factors: contains datablock(s) Lab-320K_01. DOI: 10.1107/S2052520621005655/xk5084Lab-310K_02sup38.hklStructure factors: contains datablock(s) Lab-310K_02. DOI: 10.1107/S2052520621005655/xk5084Lab-310K_01sup39.hklStructure factors: contains datablock(s) Lab-310K_01. DOI: 10.1107/S2052520621005655/xk5084Lab-300K_02sup40.hklStructure factors: contains datablock(s) Lab-300K_02. DOI: 10.1107/S2052520621005655/xk5084Lab-300K_01sup41.hklStructure factors: contains datablock(s) Lab-300K_01. DOI: 10.1107/S2052520621005655/xk5084Lab-290Ksup42.hklStructure factors: contains datablock(s) Lab-290K. DOI: 10.1107/S2052520621005655/xk5084Lab-280K_02sup43.hklStructure factors: contains datablock(s) Lab-280K_02. DOI: 10.1107/S2052520621005655/xk5084Lab-280K_01sup44.hklStructure factors: contains datablock(s) Lab-280K_01. DOI: 10.1107/S2052520621005655/xk5084Lab-275Ksup45.hklStructure factors: contains datablock(s) Lab-275K. DOI: 10.1107/S2052520621005655/xk5084Lab-240Ksup46.hklStructure factors: contains datablock(s) Lab-240K. DOI: 10.1107/S2052520621005655/xk5084Lab-250Ksup47.hklStructure factors: contains datablock(s) Lab-250K. DOI: 10.1107/S2052520621005655/xk5084Lab-270Ksup48.hklStructure factors: contains datablock(s) Lab-270K. DOI: 10.1107/S2052520621005655/xk5084sup49.pdfTables S1-S12 , Appendix A and Figs. S1-S4. DOI: 2087357, 2087358, 2087359, 2087360, 2087361, 2087362, 2087363, 2087364, 2087365, 2087366, 2087367, 2087368, 2087369, 2087370, 2087371, 2087372, 2087373, 2087374, 2087375, 2087376, 2087377, 2087378, 2087379, 2087380, 2087381, 2087382, 2087383, 2087384, 2087385, 2087386, 2087387, 2087388, 2087389, 2087390, 2087391, 2087392, 2087393, 2087394, 2087395, 2087396, 2087397, 2087398, 2087399, 2087400, 2087401, 2087402, 2087403CCDC references:"}
+{"text": "The model reflects age, platelet count, hepatitis B e antigen positivity, serum albumin and total bilirubin levels, cirrhosis development, and liver stiffness values measured by transient elastography. Our new model showed better performance for predicting hepatocellular carcinoma development (Harrell\u2019s p < 0.05). Our nomogram showed acceptable performance in predicting HCC in Asian HBV-infected patients receiving potent antiviral therapy.Hepatocellular carcinoma (HCC) risk prediction is important to developing individualized surveillance approaches. We designed a novel HCC prediction model using liver stiffness on transient elastography for patients receiving antiviral therapy against hepatitis B virus (HBV) infection. We recruited 2037 patients receiving entecavir or tenofovir as first-line antivirals and used the Cox regression analysis to determine key variables for model construction. Within 58.1 months (median), HCC developed in 182 (8.9%) patients. Patients with HCC showed a higher prevalence of cirrhosis (90.7% vs. 45.9%) and higher liver stiffness values (median 13.9 vs. 7.2 kPa) than those without. A novel nomogram (score 0\u2013304) was established using age, platelet count, cirrhosis development, and liver stiffness values, which were independently associated with increased HCC risk, along with hepatitis B e antigen positivity and serum albumin and total bilirubin levels. Cumulative HCC probabilities were 0.7%, 5.0%, and 22.7% in the low- (score \u226487), intermediate- (88\u2013222), and high-risk (\u2265223) groups, respectively. The c-index value was 0.799 , higher than that of the PAGE-B (0.726), modified PAGE-B (0.756), and modified REACH-B (0.761) models (all  Globally, approximately 240 million individuals are chronically infected with hepatitis B virus (HBV), which remains a major etiology of hepatocellular carcinoma (HCC) and cirrhosis, especially in endemic areas, including the Republic of Korea ,2,3. A hSeveral efforts were made to evaluate HCC development in patients with CHB. Several models, such as the GAG-HCC, CU-HCC, and REACH-B, were designed with sufficiently good prognostic performance ,14,15. AIn the current era of potent AVT, we aimed to establish a novel prediction model for HCC development optimized for patients with CHB receiving ETV and TDF based on liver stiffness on transient elastography, one of the most reliable fibrosis markers, and validate its role in comparison with that of other prediction models.Between 2007 and 2018, patients who started AVT with ETV or TDF against chronic HBV infection at Yonsei University Severance Hospital were consecutively screened for eligibility. The inclusion criteria were as follows: (1) age \u226519 years, (2) patients who received ETV or TDF in first-line AVT, (3) reliable liver stiffness values available, (4) no previous history of HCC at enrollment, (5) no previous history of decompensated cirrhosis with Child-Pugh class C at enrollment, and (6) no history of previous organ transplant. The exclusion criteria were as follows: (1) co-infection with other hepatitis viruses, (2) HCC development within 6 months since AVT initiation, (3) death or orthotropic liver transplant within 6 months since AVT initiation, (4) uncontrolled advanced malignancy at enrollment, and (5) other significant medical illnesses.3/\u03bcL and imaging findings suggestive of cirrhosis, including a blunted, nodular liver edge accompanied by splenomegaly (>12 cm), or (2) clinical signs of portal hypertension, such as gastroesophageal varices ), intermediate- ), and high-risk  groups according to the 25th (87 points) and 75th percentiles (223 points) of the risk score distribution of the nomogram. The 2-, 3-, and 5-year cumulative probabilities of HCC development in the low- (scores: \u226487), intermediate- (88\u2013222), and high-risk (\u2265223) groups were 0%, 0.2%, and 0.7%; 1.1%, 2.9%, and 5.0%; and 7.5%, 13.3%, and 22.7%, respectively , together with categorized serum albumin levels , total bilirubin levels (<2.0 and \u22652.0 mg/dL), and HBeAg positivity, which were reported to be closely associated with the HCC risk among subjects with chronic HBV infection ,13,15. Tc-index was 0.805 (95% CI: 0.777\u20130.834), and the 2-, 3-, and 5-year TDAUCs were 0.809 (95% CI: 0.783\u20130.834), 0.808 (95% CI 0.782\u20130.834), and 0.805 (95% CI 0.779\u20130.831), respectively.Harrell\u2019s c-index value of the prediction model was 0.799 (95% CI: 0.769\u20130.829), and the 2-, 3-, and 5-year TDAUCs were 0.802 (95% CI: 0.776\u20130.827), 0.802 (95% CI: 0.776\u20130.828), and 0.799 (95% CI 0.773\u20130.826), respectively . Calibrap < 0.001), mPAGE-B , and mREACH-B  models. The 2-, 3-, and 5-year TDAUCs of the new nomogram  were also significantly higher than those of the PAGE-B , mPAGE-B , and mREACH-B  models (p < 0.001).The c-index value of the new nomogram for predicting HCC development (0.799) was significantly higher than that of the PAGE-B  models . Furthern = 324, 36.0%).For external validation, we recruited 901 patients from Gangnam Severance Hospital and Yongin Severance Hospital using the same criteria. During follow-up (median: 51.8 [IQR 34.4\u201373.1] months), HCC developed in 56 (6.2%): 10 patients without cirrhosis and 46 patients with cirrhosis in this cohort. The c-index value and 2-, 3-, and 5-year TDAUC of the nomogram were also acceptable: 0.785 (95% CI: 0.729\u2013 0.840) and 0.782 (95% CI: 0.722\u20130.841), 0.777 (95% CI: 0.719\u20130.835), and 0.771 (95% CI: 0.714\u20130.827), respectively, in overall patients. In subgroup analysis, Harrell\u2019s C-index, 2-, 3-, and 5-year TDAUC values were 0.749 (95% CI: 0.692\u20130.806), 0.649 (95% CI: 0.522\u20130.775), 0.651 (95% CI: 0.524\u20130.779), and 0.651 (95% CI: 0.525\u20130.776), respectively, in patients without cirrhosis and 0.668 (95% CI: 0.585\u20130.750), 0.653 (95% CI: 0.571\u20130.736), 0.651 (95% CI: 0.570\u20130.732), and 0.653 (95% CI: 0.573\u20130.733), respectively, in patients with cirrhosis  during the long-term follow-up period of approximately 60 months, allowed for statistical reliability and adequate power [Our study has several strengths. Firstly, our nomogram showed consistently superior c-index and TDAUC values for predicting HCC development during the long-term follow-up period, compared with that of other conventional risk prediction models, including the PAGE-B, mPAGE-B, and mREACH-B models, which showed high performance in previous literature ,16,30. Cte power . The higte power . Finallyn = 571, 28.0%), HCC surveillance can be mitigated safely, until the value calculated by our nomogram reaches more than 87. Accordingly, repeated assessment of HCC risk at every visit is required in routine practice. Contrary, because the intermediate- and high-risk groups  had significantly higher annual incidence rates of HCC , a more sensitive imaging study of detecting HCC rather than abdominal ultrasonography, for example, magnetic resonance imaging with or without contrast, is required. Further prospective studies on personalized surveillance strategies according to individual risk and cost-effectiveness are required.The 5-year cumulative incidence of HCC was <1% in the low-risk group (score: \u226487). Generally, the currently recommended bi-annual surveillance strategy is cost-effective when the annual incidence rate of HCC ranges \u22650.2% in patients with chronic HBV infection and \u22651.5% in those with cirrhosis ,34. TherThe type of AVT does not seem to influence the risk of HCC development. Our study included similar proportions of patients who first started AVT with ETV (45.0%) and TDF (55.0%), and the type of AVT was not associated with HCC development in univariate analysis. There is controversy on whether TDF is more advantageous than ETV in reducing HCC risk ; howeverUnsolved issues remain in our study. Firstly, the findings of this study were potentially subject to selection bias. Considering that our institute is the second largest tertiary hospital with approximately 2500 beds in the Republic of Korea, our study population was more likely to cover patients with advanced liver disease and a high prevalence of cirrhosis in comparison with the nationwide cohort . HoweverThis study developed a novel nomogram to predict HCC using baseline information readily available among treatment-na\u00efve patients with chronic HBV infection who started their first-line AVT with potent nucleos(t)ide analogs. The nomogram consistently showed better prognostic performance over that of conventional models. Further studies are required to validate our results among independent cohorts, including Western populations."}
+{"text": "Agastache foeniculum (Pursch) Kuntze (Lamiaceae), Gaultheria procumbens L. (Ericaceae), Heliopsis helianthoides (L.) Sweet (Asteraceae), Liatris spicata (L.) Willd. (Asteraceae), Pycnanthemum incanum (L.) Michx. (Lamiaceae), Smallanthus uvedalia (L.) Mack. ex Mack. (Asteraceae), and Verbena hastata L. (Verbenaceae) by hydrodistillation. The essential oils were analyzed by gas chromatographic techniques. The essential oil of A. foeniculum was dominated by estragole (88\u201393%), while methyl salicylate (91%) dominated the G. procumbens essential oil. Germacrene D was the major component in H. helianthoides (42%) and L. spicata (24%). 1,8-Cineole (31%) and \u03b1-terpineol (17%) were the main compounds in P. incanum essential oil. The essential oil of S. uvedalia showed \u03b1-pinene (24%), perillene (15%), and \u03b2-caryophyllene (17%) as major components. Verbena hastata essential oil was rich in 1-octen-3-ol (up to 29%) and palmitic acid (up to 22%). Four of these essential oils, H. helianthoides, L. spicata, P. incanum, and V. hastata, are reported for the first time. Additionally, the enantiomeric distributions of several terpenoid components have been determined.As part of our evaluation of essential oils derived from Native American medicinal plants, we have obtained the essential oils of  Plants have been used in traditional medicine since prehistoric times. The therapeutic properties of medicinal plants are generally attributed to secondary metabolites produced by the plants as protection against pathogens and herbivory. As with many other aboriginal peoples, Native North Americans have used plants as medicines throughout their history. Although not as extensively documented as traditional Chinese medicine or Ayurvedic medicine, there are several sources of information regarding Native American ethnopharmacology ,2,3. As Agastache foeniculum (Pursch) Kuntze (Lamiaceae) is native to north central United States and southern Canada, but has been recorded in southern Alabama [A. foeniculum as a cold medicine [A. foeniculum have been extensively studied, and the oils are typically dominated by methyl chavicol (estragole) with smaller amounts of (E)-anethole [A. foeniculum include flavonoids , polyphenolics , pentacyclic triterpenoids , and sterols  [ Alabama . The pla Alabama . Cheyennmedicine . Essentianethole . Nonvolaastanol) .Gaultheria procumbens L. (Ericaceae) naturally ranges in eastern North America from Canada, south through Alabama and Georgia [G. procumbens to treat headaches, colds  or to treat arthritis, rheumatism, and lumbago  [G. procumbens along with the roots of Epigaea repens for chronic indigestion, and they also chewed the leaves as a substitute for chewing tobacco [Gaultheria fragrantissima Wall., the essential oil of G. procumbens is dominated by methyl salicylate. Commercial G. fragrantissima essential oil  has 99.7% methyl salicylate, while methyl salicylate in G. procumbens essential oil typically ranges 96.6\u201399.8% [ Georgia . Severalawatomi) . The Che tobacco . Much li.6\u201399.8% .Heliopsis helianthoides (L.) Sweet (Asteraceae) is native to North America from Saskatchewan, Canada east to the Atlantic coast of Newfoundland and south to the Gulf of Mexico, with the western range extending as far as New Mexico [H. helianthoides, ssp. helianthoides, generally occurring east of the Mississippi River; ssp. occidentalis, found in the Great Plains region; and ssp. scabra, which is predominant in the Ozark region  [H. helianthoides as a stimulant [Scutellaria incana \u201cfor young women\u201d, presumably for menstruation-related discomforts, and sore feet were relieved by soaking in an infusion of what was called \u201cswamp sunflower\u201d [N-alkylamides [H. helianthoides.w Mexico ,11,12. Tnflower\u201d . Guaianonflower\u201d , N-alkylylamides , and ligylamides  have beeLiatris spicata (L.) Willd. (Asteraceae) is the eastern United States and Canada, east of the Mississippi River, from the south along the Gulf of Mexico including southern Alabama and northern Florida areas, north to Ontario and Quebec [L. spicata [The natural range of d Quebec ,16,17. Td Quebec . The gua spicata .Pycnanthemum incanum (L.) Michx. (Lamiaceae) ranges naturally in the eastern United States from the Mississippi River east to the Atlantic coast and from southern Ontario, Canada south to the Gulf of Mexico, though it is primarily found from the Appalachian mountain region beginning in north Georgia north into Ontario, Canada [P. incanum externally to treat headaches [P. incanum to be \u03b2-ionone, myrcene, linalool, and pulegone [, Canada ,19. The eadaches . Dein anpulegone .Smallanthus uvedalia (L.) Mack. ex Mack. (Asteraceae) is the southeastern United States, from the Ohio river basin south to the Gulf of Mexico [Smallanthus uvedalia was reportedly used internally by Native American Indians for laxative properties as well as a stimulant and also to treat swollen glands, especially mastitis [ent-kaurane diterpenoids have been isolated and characterized from S. uvedalia [S. uvedalia from several locations in north Alabama have been analyzed previously [The natural range of f Mexico . Smallanmastitis . The Chemastitis . Interesmastitis . A numbeuvedalia . The leaeviously ,24.Verbena hastata L. (Verbenaceae) ranges throughout North America [V. hastata has shown antiplasmodial [V. hastata [ America . The Che America . The ethasmodial , antinocasmodial , and antasmodial . The iri hastata .The purpose of this study was to extend our understanding of the volatile phytochemistry of Native American aromatic medicinal plants by examination of the essential oils of these seven plant species, to determine their chemical compositions as well as the enantiomeric distributions of terpenoid constituents.The essential oils of each species were obtained by hydrodistillation of dried plant material .A. foeniculum were collected from cultivated plants in Newville, Alabama. Hydrodistillation gave pale yellow essential oils in yields ranging from 1.48% to 2.30% yield. The essential oil compositions are compiled in The aerial parts of three different plant samples of A. foeniculum essential oil was dominated by the phenylpropanoid methyl chavicol (=estragole). There are apparently five different chemotypes of A. foeniculum based on essential oil chemical profiles: (1) methyl chavicol, (2) spathulenol/bornyl acetate, (3) \u03b3-cadinene/\u03b1-cadinol, (4) limonene, and (5) isomenthone [A. foeniculum essential oils belong to the methyl chavicol chemotype, however [The menthone ,34. Most however ,40,41,42A. foeniculum and has shown anti-inflammatory and anti-edematogenic [Methyl chavicol contributes to the anise-like aroma of atogenic ,44, cytoatogenic , and antatogenic ,47. Unfoatogenic ,49.G. procumbens was obtained in 4.25% yield and the major component was methyl salicylate (91.1%)  . Methyl 1.1% . Meo 99.96% ,50,51,52G. procumbens essential oil, methyl salicylate, is well-known as an anti-inflammatory, antipyretic, analgesic agent [The major component of ic agent , and accic agent ,55,56.H. helianthoides was obtained in 0.95% yield. The major component in the essential oil was germacrene D (42.4%), with a lesser amount of 4-vinylguaicol (5.5%)  . As far Centaurea hadimensis Wagenitz, K. Ertugrul & H. Dural (44.3%) [Centaurea pseudoscabiosa Boiss. & Buhse (36.0%) [Eupatorium cannabinum L. (33.5%) [Polymnia canadensis L. (63.6%) [Rudbeckia fulgida Aiton (30.1%) [Rudbeckia hirta L. (23.6%) [Solidago canadensis L. (64.1%) [Symphyotrichum novae-angliae (L.) G.L. Nesom (25.5%) [Verbesina macrophylla (Cass.) F.S. Blake (37.3%) [Verbesina turbacensis Kunth (36.9%) [Liatris spicata . Germacrene D has shown antimicrobial and cytotoxic activities [Although not necessarily a phytochemical marker of the family, germacrene D has been found to be a major component in several members of the Asteraceae. For example, germacrene D is the dominant compound in the essential oils of  (44.3%) , Centaur (36.0%) , Eupator (33.5%) , Polymni 63.6%) , Rudbeck (30.1%) , Rudbeck (23.6%) , Solidag 64.1%) , Symphyo (25.5%) , Verbesi (37.3%) , Verbesi (36.9%) , and Lia.6% , Rud.1% , SymL. spicata essential oil is presented in L. spicata.The essential oil composition of P. incanum growing wild in South Carolina. The essential oil was rich in oxygenated monoterpenoids, including 1,8-cineole (30.7%), \u03b1-terpineol (16.9%), borneol (8.2%), and cis-sabinene hydrate (5.6%). The sesquiterpene hydrocarbons (E)-\u03b2-caryophyllene (11.0%), and germacrene D (5.0%) were also relatively abundant. To our knowledge, this is the first reported analysis of P. incanum essential oil. Volatiles obtained from a diethyl ether extract have been analyzed by gas chromatography-olfactometry to determine the key odorants [P. incanum essential oil and the volatiles from the previously published diethyl ether extract [odorants . Althoug extract . ConcentS. uvedalia from South Carolina is summarized in S. uvedalia essential oil were \u03b1-pinene (23.9%), (E)-\u03b2-caryophyllene (16.9%), perillene (14.5%), germacrene D (12.2%), and limonene (6.1%). In comparison, S. uvedalia from northern Alabama (collected in September 2018) contained \u03b1-pinene (62.6%), limonene (11.4%), and \u03b2-pinene (6.0%), with lesser concentrations of (E)-\u03b2-caryophyllene (3.8%) [S. uvedalia samples collected in February, 2016, from northern Alabama were rich in (E)-\u03b2-caryophyllene (24.5% and 16.5%) and caryophyllene oxide (19.8% and 14.2%) [S. uvedalia may be attributed to geographical location and/or seasonal variation.The essential oil composition of e (3.8%) . Neither\u03b1-pinene .9%, (E)-Erechtites hieracifolia (L.) Raf. [S. uvedalia (this work). Likewise, (+)-\u03b2-pinene was the only enantiomer in Coreopsis capillacea Kunth (syn. C. triloba S.F. Blake) [Achillea ligustica All. [S. uvedalia (this work) and Solidago canadensis L. [E. hieracifolia [C. capillacea [There does not seem to be a trend in the major enantiomers for essential oils of the Asteraceae see . For exaL.) Raf. , but (\u2013). Blake) , while (ica All. . (+)-Limensis L. , whereasacifolia  and C. cpillacea .V. hastata were collected and investigated (E)-\u03b2-ionone, and (E)-nerolidol were the same for the three samples, however. As far as we know, this is the first report on the essential oil composition of V. hastata.Three different specimens of stigated . AlthougVerbena essential oil compositions to compare. However, several Verbena officinalis L. essential oils have been reported, and these samples also show wide variation in composition. The major components in V. officinalis essential oil from Morocco were spathulenol (10.8%), limonene (7.5%), 1,8-cineole (7.5%), caryophyllene oxide (7.3%), and ar-curcumene (6.0%) [V. officinalis from Italy was rich in geranial (45.5%) and isobornyl formate (41.4%) [Verbena officinalis from Algeria, on the other hand, showed limonene (17.7%), geranial (14.8%), carvone (14.2%), and caryophyllene oxide (12.4%) as major components [There are few e (6.0%) . In cont (41.4%) . Verbenamponents .V. hastata essential oils as it was in P. incanum essential oil (above). Notably, (\u2013)-1-octen-3-ol is the major enantiomer, generally greater than 97%, in mushrooms [The (\u2013)-enantiomer of 1-octen-3-ol was the only stereoisomer observed in ushrooms , and is ushrooms . Interesushrooms .V. hastata. (\u2013)-Linalool also dominated in the essential oil of Lantana camara L. (Verbenaceae) from Madagascar [Lippia alba (Mill.) N.E. Brown (Verbenaceae) from Uruguay was dominated by the (+)-enantiomer [A racemic mixture was observed for \u03b1-terpineol, but there was a higher concentration of (\u2013)-linalool over (+)-linalool in dagascar . In contantiomer .A. foeniculum, G. procumbens, and H. helianthoides were obtained from plants cultivated in at Kirkland Gardens, in Newville, Alabama , tubers , or young plants . All the plants were grown in full sun, except the G. procumbens, which was located in a partially shaded location (4 h/day average sunlight), and all were watered at least once a week. The plants were cultivated directly in the ground, which was clayey-loamy sand, which was amended with composted chicken manure, worm castings, kelp meal, and bone meal at time of planting. Pycnanthemum incanum and S. uvedalia were collected in the wild from a fully shaded forest understory roadside location near a small waterfall in northern South Carolina. The plants were located beside highway 276 near the North Carolina\u2013South Carolina border , S. uvedalia (SKL31820), and V. hastata (SKL51321) were deposited in the University of Alabama in Huntsville Herbarium (HALA); cultivated plants were not vouchered. For each species, the plant material was air-dried in the laboratory (around 23 \u00b0C) for 10 days. The dried plant materials of each species were chopped and hydrodistilled using a Likens\u2013Nickerson apparatus with continuous extraction with dichloromethane for 4 h. The dichloromethane was evaporated using a stream of dry nitrogen to give the essential oils , gas chromatography-flame ionization detection (GC-FID), and chiral GC-MS as previously described .GC-MS: Shimadzu GCMS-QP2010 Ultra, electron impact (EI) mode (electron energy = 70 eV), scan range = 40\u2013400 atomic mass units, scan rate = 3.0 scans/s, and GC-MS solution software; ZB-5ms fused silica capillary column (30 m length \u00d7 0.25 mm inner diameter) with a (5% phenyl)-polymethylsiloxane stationary phase and a film thickness of 0.25 \u03bcm; He carrier gas with a column head pressure of 552 kPa and flow rate of 1.37 mL/min; injector temperature = 250 \u00b0C, ion source temperature = 200 \u00b0C; GC oven temperature 50\u2013260 \u00b0C (2 \u00b0C/min), 1-\u03bcL injection of 5% solution of EO in dichloromethane . The essential oil components were identified by MS fragmentation, and retention indices compared with those in the databases ,30,31,322 carrier gas, and flow rate = 1.0 mL/min. The composition percentages were calculated from raw peak areas without standardization.GC-FID: Shimadzu GC 2010 equipped with flame ionization detector, a split/splitless injector, and Shimadzu autosampler AOC-20i, with a ZB-5 capillary column (30 m length \u00d7 0.25 mm inner diameter) with a (5% phenyl)-polymethylsiloxane stationary phase and a film thickness of 0.25 \u03bcm; oven temperature was programmed the same as above for GC-MS; injector temperature = 250 \u00b0C, detector temperature = 280 \u00b0C, NChiral GC-MS: Shimadzu GCMS-QP2010S, EI mode (electron energy = 70 eV) with scan range of 40\u2013400 amu and scan rate of 3.0 scans/s; Restek B-Dex 325 capillary column (30 m \u00d7 0.25 mm ID \u00d7 0.25 \u03bcm film); GC oven temperature program, 50\u2013120 \u00b0C (1.5 \u00b0C/min), 120\u2013200 \u00b0C (2 \u00b0C/min), and kept at 200 \u00b0C for 5 min; He carrier gas, flow rate = 1.8 mL/min; 0.1-\u03bcL injection of 3% solution of EO in dichloromethane . The monoterpenoid enantiomers were identified by comparison of retention times with authentic samples obtained from Sigma-Aldrich . Relative enantiomer percentages were determined based on peak areas. Chiral GC-MS chromatograms are available as Heliopsis helianthoides, Liatris spicata, Pycnanthemum incanum, and Verbena hastata, were reported for the first time. Additionally, the enantiomeric distributions of several terpenoid components have been determined. The chemical compositions presented add to our knowledge of the phytochemistry of the medicinal plants.This report presented the essential oil compositions of seven aromatic medicinal plants used by Native Americans. Four of these essential oils,"}
+{"text": "MUC1-C integrates activation of the IFN-\u03b3 pathway with suppression of the tumor immune microenvironment in triple-negative breast cancer. J Immunother Cancer. 2021;9:e002115. doi: 10.1136/jitc-2020-002115.Yamashita N, Long M, Fushimi A, This article has been corrected since it first published. The provenance and peer review statement has been added."}
+{"text": "Correction to: Microbiome 9, 241 (2021)https://doi.org/10.1186/s40168-021-01195-7Supplementary Data was missing from this article and has now been uploaded.The original article has been updated.Additional file 1: Supplementary Table S1. Details of the search terms used in the respective databases. Supplementary Table S2a. Additional summary of African Gut Microbiome studies. Supplementary Table S2b. Additional summary of African Urogenital Microbiome studies. Supplementary Table S2c. Additional summary of African Microbiome studies (Other body sites)."}
+{"text": "The DISC1 (disrupted in s\u0441hizophrenia 1) gene is associated with brain dysfunctions, which are involvedin a variety of mental disorders, such as schizophrenia, depression and bipolar disorder. This is the first study toexamine the immune parameters in Disc1-Q31L mice with a point mutation in the second exon of the DISC1 genecompared to mice of the C57BL/6NCrl strain . A flow cytometry assay has shown that intact Disc1-Q31L mice differ from the WT strain by an increase in the percentage of CD3+ T cells, CD3+CD4+ \u0422 helper cellsand CD3+CD4+CD25+ T regulatory cells and a decrease in CD3+CD8+ T cytotoxic/suppressor cells in the peripheralblood. A multiplex analysis revealed differences in the content of cytokines in the brain structures of Disc1-Q31Lmice compared to WT mice. The content of pro-inflammatory cytokines was increased in the frontal cortex  and striatum (IFN\u03b3), and decreased in the hippocampus and hypothalamus. At the same time, thelevels of IL-1\u03b2 were decreased in all structures being examined. In addition, the content of anti-inflammatory cytokinesIL-4 was increased in the frontal cortex, while IL-10 amount was decreased in the hippocampus. Immuneresponse to sheep red blood cells analyzed by the number of antibody-forming cells in the spleen was higher inDisc1-Q31L mice at the peak of the reaction than in WT mice. Thus, Disc1-Q31L mice are characterized by changes inthe pattern of cytokines in the brain structures, an amplification of the peripheral T-cell link with an increase in thecontent of the subpopulations of CD3+CD4+ T helpers and CD3+CD4+CD25+ T regulatory cells, as well as elevatedimmune reactivity to antigen in the spleen. Animal models have provided valuableopportunities to study the impact of immune dysfunctions andrelated alterations in neurotransmitter and hormonal systemsin the pathogenesis of neuropsychiatric disorders caused bymultiple risk factors, including genetic background. As shownpreviously, animals with genetic predisposition to depressiveor aggressive behavior are characterized by changes in thedistribution and ratio of the main subpopulations of T-cells inthe blood and spleen, immune responsiveness to T-dependentantigen, as well as cytokine variations in the periphery andbrain structures .It is now well established that a variety of social, environmentaland genetic factors may cause inflammatory responses that,over time, may result in development of multiple diseases,including neuropsychiatric disorders . The inflammatory processesare closely associated with alterations in the productionof cytokines , the compositionof T-cell subsets with different functional activities . Analysis of emotional,social and cognitive behaviors of this mice line showeda range of behavioral abnormalities that may be considered asa depression-like endophenotype . TheQ31L mutation in DISC1 gene is also known to be associatedwith changes in the dopaminergic (DA) activity  and other neuromediator systems, which are involvedin the neurobiological mechanisms of psychiatric disordersand in the control of immune function .Disrupted-in-Schizophrenia-1 (DISC1) gene has beenfunctionally linked to brain dysfunctions associated with impairedneurodevelopment processes and intracellular signalingpathways that predispose to schizophrenia, major depression,and bipolar disorder . Several mouse models based on DISC1 dysfunctionhave been generated to date, including a homozygousDisc1-Q31L\u2013/\u2013 mice remain to be elucidated. Given a roleof the immune system both in the development of differentpsychoemotional states and in neuroimmunomodulation was also determined.2019), the aim of this study was to analyze the basal contentof T- and B-cells in the peripheral blood and spleen, as wellas the level of pro- and anti-inflammatory cytokines in thebrain structures of Disc1-Q31LAnimals. The experiments were performed in adult (3.0\u20133.5 months old) homozygous male mice of the Disc1-Q31L\u2013/\u2013strain (n = 23) and there wild type (WT) littermates of theC57BL/6NCrl strain (n = 23) weighing 27\u201330 g. Mice werebred in the animals facility of the Scientific Research Instituteof Physiology and Basic Medicine .Mice were kept in standard cages  in groups for 5 animals per cage under standardvivarium conditions and free access to food and water.All experimental procedures were performed in accordancewith the requirements of the European Community Directive(86/609/EC) and approved by Local Ethical Committeeof the Scientific Research Institute of Physiology and BasicMedicine, protocol No. 10 (17.12.2015).Design of experiments. The levels of T- and B-lymphocytesand their subpopulations in the peripheral blood andspleen, as well as the content of proinflammatory  and anti-inflammatory (IL-4and IL-10) cytokines in the brain structures  were assessed in intactmice of the Disc1-Q31L and WT strains . The immune reactivity to sheep red blood cells (SRBC)was analyzed by measuring the number of antibody-formingcells (AFC) in the spleen of mice of both strains (n = 13 ofeach strain). SRBC were suspended in saline and injectedonce, intravenously into the tail vein at a dose of 5 \u00b7 108.Blood was immediately collected after the animals weredecapitated into tubes containing K3EDTA . Spleens were removed on ice on day 4 afterSRBC injection and placed in tubes with cooled RPMI-1640medium . Brain structures were dissectedon ice; brain samples were frozen in liquid nitrogenand stored at \u201370 \u00b0C until analysis.Determination of cell subpopulation. To analyze cellsubsets, 25 \u03bcl of blood was incubated for 30 minutes ina dark place with 1.5 \u03bcl (0.2\u20130.5 \u03bcg/\u03bcl) labeled rat anti-mousemonoclonal antibodies (MoAB) against surface markers:\u0421D3 , \u0421D4 , \u0421D8 , \u0421D25 (BrilliantViolet 421), \u0421D19  . Erythrocytes of theblood were lysed with Lysing Solution BD FASC . After a 10-minute incubation, the cells werewashed once with phosphate buffered saline (PBS), the cellpellet was resuspended in 100 \u03bcl of PBS.6/100 \u03bcl of the suspension and placed intoplates in a volume of 100 \u03bcl in each well. The cell suspensionwas incubated with the same MoAB as the blood cellsfor 20 minutes, and fixed by adding 1 % paraformaldehyde toeach tube. Isotypic antibodies were used as a control.The spleen was cut into several pieces, and then disaggregatedmechanically into single-cell suspension, which waspassed through a 50 \u03bcm cell strainer. The suspension waswashed twice with RPMI-1640 medium at 200 g for 5 minutes.The cell pellet was resuspended in RPMI-1640 medium,adjusted to 1 \u00b7 10+ T-lymphocytes, CD3+CD4+T-helpers, CD3+CD8+ cytotoxic/suppressor T-lymphocytes,CD3+CD4+CD25+ T-regulatory cells, CD19+ B-lymphocytesas a percentage of the total number of cells were determined.Immunoregulatory index was measured as a ratio of the contentof CD4+ to CD8+ \u0422-cells.The study of cell populations was performed on a FACSCANTO\u2122 II flow cytometer  usingmulti-stage gating. At least 50 000 cells were analyzed in eachsample. Data analysis was performed using the FACSDivasoftware. The contents of CD3Determination of cytokines in the brain structures.For the analysis of cytokines, detergent-soluble fractions ofbrain tissues were prepared. The samples were thawed on ice,homogenized in lysis buffer cooled to +4 \u00b0C containing PBS(pH 7.4), 0.1 % Triton X-100, 1 mM EDTA, and 1 mM PMSFusing plastic pestles. The homogenates were incubated on icefor 30\u201340 minutes. The tissue extracts were centrifuged (Centrifuge5415 R) at 4500 rpm for 20 minutes at +4 \u00b0C. Cytokineconcentrations were determined in the supernatants. Theconcentration was normalized to tissue weight (pg/g tissue)The content of cytokines in brain homogenates was determinedaccording to the manufacturer\u2019s protocol by multipleximmunoassay on a multiplex protein and nucleic acid analyzer using a kit . The resultswere analyzed using the xPONENT and Analist software.Determination of antibody-forming cells. The immuneresponse was assessed by the relative (per 106 spleen cells)and absolute  numberof IgM-AFC using the standard method .Statistical analysis. The data were analyzed using Statistica10.0 software. To verify whether data were normallydistributed, the Kolmogorov\u2013Smirnov and Shapiro\u2013Wilktests were used. Normally distributed data  were assessedby one-way ANOVA. The Mann\u2013Whitney test was used forabnormally distributed data (cytokine content and AFC number).Data are presented as mean and mean error (M \u00b1 m) withsignificance set at a level of p < 0.05.Content of T-cells, their subpopulations, and B-cells in theperipheral blood and spleen of Disc1-Q31L mice. Therewere differences in the content of all analyzed immunocompetentblood cells between nonimmunized Disc1-Q31L andWT mice. The percentage of CD3+ T-lymphocytes in mice of the Disc1-Q31L strain was significantly higher than inWT mice (F(1.18) = 45.2, p < 0.001). Analysis of T-lymphocytesubpopulations showed an increase in the content ofCD3+CD4+ \u0422-helpers (F(1.17) = 15.5, p < 0.01) in Disc1-Q31Lmice compared to WT strain, while the number of CD3+CD8+T-cytotoxic/suppressor cells was decreased (F(1.17) = 12.6,p < 0.01). As a result, the immunoregulatory index, determinedas the ratio of the content of CD4+ to CD8+ T-lymphocytes, inmutant mice was 1.3 times higher (F(1.18) = 27.5, p < 0.01)than in WT mice. The content of T-regulatory cells with theCD3+CD4+CD25+ phenotype in Disc1-Q31L mice was alsohigher than in WT mice (F(1.17) = 5.3, p < 0.05). The numberof CD19+ B-lymphocytes was decreased in the peripheralblood of mutant mice compared to WT mice (F(1.17) = 5.7,p < 0.05) (see the Table).Cytokines in the brain structures in mice of the Disc1-Q31L strain. Analysis of the cytokine profile in brain structuresof intact Disc1-Q31L mice revealed regional differencesin the content of cytokines between mutant and WT mice.In the frontal cortex, levels of the three pro-inflammatorycytokines IL-6 ( p < 0.01), IL-17 ( p < 0.01) and IFN\u03b3( p < 0.01) were higher in Disc1-Q31L mice than in WT mice,while the level of IL-1\u03b2 ( p < 0.05) decreased. IL-2 and TNF\u03b1levels were similar between the mutant and WT strains( p < 0.05). As to the content of anti-inflammatory cytokines,the level of IL-4 in Disc1-Q31L mice was higher than inWT mice ( p < 0.01), while the level of IL-10 did not change( p > 0.05) .In the striatum, the content of IFN\u03b3 ( p < 0.01) was foundto be increased in Disc1-Q31L mice compared to WT animals.The levels of other pro-inflammatory cytokines \u2013 IL-1\u03b2( p < 0.01), IL-2 ( p < 0.001) in the mutant mice were lowerthan in WT mice, while the levels of IL-6, IL-17 and TNF\u03b1remained unchanged ( p > 0.05). Similarly, there were nosignificant strain differences in the levels of anti-inflammatorycytokines IL-4 and IL-10 ( p > 0.05) .When compared to WT mice, Disc1-Q31L mice showedlower levels of IL-1\u03b2 ( p < 0.01), IL-2 (p < 0.01) and IL-17( p < 0.05) in the hypothalamus, while the levels of the restcytokines were unchanged ( p > 0.05) .The levels of proinflammatory cytokines IL-1\u03b2, IL-2,IL-17, TNF\u03b1 ( p < 0.05) were significantly lower in the hippocampusof Disc1-Q31L mice than in WT mice, with morepronounced decrease in IFN\u03b3 content ( p < 0.001). The levelsof IL-6 were equivalent between Disc1-Q31 and WT mice( p > 0.05). The content of anti-inflammatory cytokine IL-10in the hippocampus of Disc1-Q31 mice was also decreased ( p < 0.05), while the level of IL-4 did not differ from that ofWT mice ( p > 0.05) .Immune reaction of Disc1-Q31L mice to the antigen.Immunization of Disc1-Q31L mice with SRBC produced anincrease of the immune response at the peak of its developmentin the spleen of WT mice. The relative ( p < 0.001) andabsolute ( p < 0.001) numbers of AFC in Disc1-Q31L micewere significantly higher than in WT mice .+ T-helpers, and the ratio of CD4+to CD8+ T-cells . Aggressive behavior, asobserved in a variety of animal models, is also associated withan increase in T-helpers and the CD4+/CD8+ ratio, as well asa higher immune response generated by an antigen .Changes in DISC1 protein activities caused by mutations inthe DISC1 gene are known to be involved in multiple mentaldisorders, such as schizophrenia, depression, bipolar disorder. Alterations in immune variables associated withthese disorders may differentially contribute to disease development.Schizophrenia has been found to be accompanied byelevated serum numbers of B-cells, along with a decrease inthe content of T-cells, CD4+CD8+ T-suppressor/cytotoxic cells, adecrease in the immunoregulatory index and the immuneresponse suppression .On the other hand, depression is characterized by increasingnumbers of CD3+ T-lymphocytes, and their subpopulations, such asCD3+CD4+ T-helpers and CD3+CD4+CD25+ T-regulatorycells, with a consequent increase of the immunoregulatoryindex. At the same time, the mutant mice showed lower percentageof CD3+ T-lymphocytes in the spleen, that might leadto a predominance of CD19+ B-cells, thereby suggesting aredistribution of these cell subsets within the immune system.The present study demonstrates that, compared to WT mice,intact mice of the Disc1-Q31L strain have raised blood levelsof CD3Redistribution of T- and B-lymphocytes, which are knownto produce specific sets of cytokines, and the ratio of thesecells in the immunocompetent organs may significantly affectinflammatory and immune processes characteristic of geneticallydetermined behaviors and psychopathology . It seems, thus, possiblethat higher ability of Disc1-Q31L mice to respond to antigenchallenge, as measured by the numbers of AFC in the spleen,may be related to changes in immune cell distribution amongdifferent compartments of the immune system.Our results have also shown that the pattern of cytokinevariations over brain structures differ in Disc1-Q31L andWT mice and depends on the brain area, in which thesecytokines are localized. The levels of IL-6, IL-17 and IFN\u03b3were found to be simultaneously increased only in the frontalcortex of Disc1-Q31L mice compared to WT animals. Thesepro-inflammatory cytokines has long been known as verypotent signaling molecules of neuroinflammation implicatedin the pathophysiology of depression, bipolar disorder, andschizophrenia .Moreover, the frontal cortex has also been associated withthe development of various psychiatric diseases .Only the IFN\u03b3 level was increased in the striatum of Disc1-Q31L mice compared to WT mice, whereas the concentrationsof other cytokines decreased. Levels of pro-inflammatory cytokineswere also lower in the hippocampus and hypothalamusof mutant mice than in WT mice. Changes in the distributionof brain cytokines found in Disc1-Q31L mice suggest thatthis mutation may contribute to neuroinflammation, whichis an important etiological factor for affective disorders. Theobserved increase in the level of anti-inflammatory cytokineIL-4 in the frontal cortex of Disc1-Q31L mice could reflecta compensatory response to the elevations of pro-inflammatorycytokines that occurred in this brain area. These findings areconsistent with previous reports, showing that various formsof depression-like behavior or aggression are associatedwith impaired balance between pro- and anti-inflammatorycytokines in a number of brain regions including the frontalcortex and hippocampus .DISC1 has been found to form a complex with other intranucleartranscription factors, which mediate the expression ofseveral genes implicated in behavioral changes resembling human psychiatric disorders . A wide rangeof studies on behavioral phenotype of Disc1-Q31L producedconflicting results. Some data indicate that mice with Q31Lmutation in Disc1 have a depressive-like enodophenotype, while others did not show significant behavioral differencesin this strain compared with the WT control in any of thetests . Recent evidence suggests that Disc1-Q31L mice may also display aggressive behavior . In contrast to the data obtained in other models, inwhich animals developing depression-like responses showedimmunosuppression , our results revealed higher immunereactivity in Disc1-Q31L mice compared to WT control. Atthe same time, immune parameters characteristic of Disc1-Q31L mice are more relevant to those observed in aggressiveanimals. It may be due to the diverse behavioral phenotype ofthese mice displaying not only depression, but also aggressivebehavior  that is associated with increasedimmune function and specific pattern of cytokines.However, the mechanisms underlying alterations in peripheralimmune parameters and the profile of brain cytokines areunknown. There is growing evidence that immune mediatorssuch as cytokines are involved in the interactions betweenthe immune and neuroendocrine systems and can change theactivity of central neuromediator systems that contribute tocognitive, behavioral, and brain structure abnormalities seenin affective disorders . It is possible that the immune status of Disc1-Q31Lmice could be related to the neurochemical pattern of thebrain characteristic of this strain. Disc1-Q31L mice have beenshown to have decreased levels of DA combined with DOPACincrease in the nucleus accumbens , knownto implicate in neuroimmunomodulation . There is also data thatthe DOPAC/DA ratio, which may reflect the metabolic rate ofDA and synaptic activity, increases under immunostimulationobserved in animals experienced excessive aggression associatedwith elevated activity of the DA system . Taking into account changing DAergicactivity in the brain structures of Disc1-Q31 mice  and the critical role of this system in the control ofaggression and neuroimmunomodulation , it is likely that DA maycontribute to the enhancement of immune function found inthe Disc1-Q31 strain.However, it remains unclear whether variations of centralcytokines are related to brain alterations of monoamines specificfor Disc1-Q31L mice or the Q31L mutation determinestheir profile. Moreover, it has been found that not only neurotransmitterscan affect the production of cytokines , but also cytokines can modulate mediator neurotransmissionand promote changes in the neurochemicalpattern of the brain .+CD4+CD25+ T-regulatory cell subpopulations, as wellas elevated immune reactivity in the spleen induced by theantigen. Alterations in the peripheral immune variables areaccompanied with changes in the distribution of pro- andanti-inflammatory cytokines within brain structures, whichare involved both in the control of different forms of behaviorand in immune function. The Disc1-Q31L mouse strain is apromising model for further study of the relationships betweengenetic factors and neuroimmunological mechanismsand their implication in the development of psychoemotionaldisorders.Our data indicate that the Q31L point mutation in the DISC1gene leading to the substitution of glutamine to leucineat amino acid 31 has a significant influence on immunity and may result in an amplification of peripheral T-cell linkwith an increase in the content of CD3+CD4+ T-helpers andCD3The authors declare no conflict of interest.Al\u2019perina E.L. Involvement of the dopaminergic system in the mechanismsof immunomodulation. 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+{"text": "The SARS-CoV-2 B.1.617 lineage variants, Kappa (B.1.617.1) and Delta  emerged during the second wave of infections in India, but the Delta variants have become dominant worldwide and continue to evolve. Here, we compared B.1.617 variants for neutralization resistance by convalescent sera, mRNA vaccine-elicited sera, and therapeutic neutralizing antibodies using a pseudovirus neutralization assay. B.1.617.1, B.1.617.2, and AY.1 pseudoviruses showed a modest 1.5- to 4.4-fold reduction in neutralization by convalescent sera and vaccine-elicited sera. In comparison, similar modest reductions were also observed for C.37, P.1, R.1, and B.1.526 pseudoviruses, but 7- and 16-fold reductions for vaccine-elicited and convalescent sera, respectively, were seen for B.1.351 pseudoviruses. Among twenty-three therapeutic antibodies tested, four antibodies showed either complete or partial loss of neutralization against B.1.617.2 pseudoviruses and six antibodies showed either complete or partial loss of neutralization against B.1.617.1 and AY.1 pseudoviruses. Our results indicate that the current mRNA-based vaccines will likely remain effective in protecting against B.1.617 variants. Finally, the P681R substitution confers efficient cleavage of B.1.617 variants\u2019 spike proteins and the spike of Delta variants exhibited greater sensitivity to soluble ACE2 neutralization, as well as fusogenic activity, which may contribute to enhanced spread of Delta variants. SARS-CoV-2 trimeric spike (S) glycoprotein on the virion surface binds the angiotensin-converting enzyme (ACE2) to facilitate cellular entry and is the target of therapeutic neutralizing antibodies and vaccines [685|S686 into S1 and S2 subunits, which facilitates subsequent cleavage at the S2\u2032 site (R815|S816) by TMPRSS2 for viral entry into respiratory cells. The S1 subunit spans the N-terminal domain (NTD) and the receptor-binding domain (RBD) within the C-terminal domain (CTD) whereas the S2 subunit spans the fusion peptide and a linker region flanked by heptad repeat regions that drive virus-cell membrane fusion [Since its origin in December 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread globally to cause a coronavirus disease 2019 (COVID-19) pandemic that recorded more than 263 million infections and has claimed 5.2 million lives thus far  ,9,10,11.Several key substitutions in the RBD of spike were demonstrated to either enhance affinity towards ACE2 or contribute to immune escape. The E484K substitution in the RBD of B.1.351, P.1, R.1, and B.1.526 variants was previously identified among in vitro escape mutants selected against single antibody and antibody cocktails ,14. SeveThe spike protein of the B.1.617.2 variant contains nine substitutions and deletions compared to the early D614G variant used here as wild type (WT or D614G). The three substitutions  and two deletions  in NTD occur in the NTD antigenic supersite spanning between residues 14\u201320, 140\u2013158, and 245\u2013264 . In addiThe global dominance and ongoing evolution of B.1.617 lineage variants require continuing assessment of the neutralization potency of convalescent sera, vaccine-elicited sera, and therapeutic neutralizing antibodies against emerging B.1.617 variants. Here, we measured the neutralization potency of convalescent sera, vaccine-elicited sera, and therapeutic neutralizing antibodies against two independent variants each in the Kappa (B.1.617.1) and Delta  lineages and assessed the contribution of the RBD substitutions in conferring resistance. We found that resistance to convalescent and vaccine-elicited sera was predominantly conferred by RBD substitutions E484Q, T478K, and L452R. Furthermore, out of 23 therapeutic neutralizing antibodies tested, Kappa and Delta pseudoviruses displayed complete resistance to five neutralizing antibodies and partial resistance to one antibody. Finally, we showed that the P681R substitution confers enhanced furin processing in spike protein of B.1.617 lineage variants that corresponded to enhanced cell-cell fusion activity. However, only Delta spike protein exhibited greater sensitivity to soluble ACE2 inhibition, implying enhanced ACE2 affinity. This feature along with enhanced cell-cell fusion activity may contribute to the dominance of the B.1.617.2 variant.Use of de-identified sera in this study was approved by the U.S. Food and Drug Administration Research in Human Subjects Committee. Vaccine-elicited sera were collected at the U.S. Food and Drug Administration with written consent under an approved Institutional Review Board (IRB) protocol (FDA IRB Study # 2021-CBER-045).Codon-optimized full-length open reading frames of the S genes of SARS-CoV-2 variants were cloned into pcDNA3.1(+) or pVRC8400 by GenScript . The codon optimization for gene expression in human cell was performed by GenScript\u2019s OptimumGene algorithm system to optimize the following parameters: codon usage bias, GC content, CpG dinucleotides content, mRNA secondary structure, internal chi sites and ribosomal binding sites, negative CpG islands, cryptic splicing sites, premature PolyA sites, PolyT sites, RNA instability motif (ARE), and repeat sequences . The codon-optimized sequence of Wuhan-Hu-1 S gene is shown in n = 10) collected 6\u201361 days after symptom onset were purchased from Bocabiolistics . Donors were 18\u201373 years old with six males/four females. The information about the convalescent sera is shown in n = 15) or Moderna mRNA-1273 vaccinated individuals (n = 14) obtained two weeks after the second vaccination were used in this study. Vaccinated individual donors were 21\u201365 years old with six males/nine females for Pfizer BNT162b2 vaccination and eight males/six females for Moderna mRNA-1273 vaccination. All sera were tested negative for non-specific neutralization using amphotropic murine leukemia enveloped pseudovirus. Vaccinated donors were prescreened for absence of both history of SARS-CoV-2 infection and SARS-CoV-2 neutralizing antibodies prior to vaccination. Twenty-three therapeutic neutralizing antibodies against SARS-CoV-2 spike protein were donated by different pharmaceutical companies for the U.S. government COVID-19 response Therapeutics Research Team efforts to define neutralization profiles against existing and emerging SARS-CoV-2 variants [Convalescent sera from SARS-CoV-2-infected donors  as described previously .6 relative luminescence units (RLU)/mL of luciferase activity were incubated with serially diluted sera or antibodies for two h at 37 \u00b0C. Pseudovirus and serum or antibody mixtures (100 \u03bcL) were then inoculated onto the plates pre-seeded one day earlier with 3.0 \u00d7 104 cells/well. Pseudovirus infectivity was scored 48 h later for luciferase activity. The antibody concentration or inverse of the sera dilutions causing a 50% reduction of RLU compared to control was reported as the neutralization titer. Titers were calculated using a nonlinear regression curve fit . The mean titer from at least two independent experiments each with intra-assay duplicates was reported as the final titer. WT(D614G) pseudovirus was run as a control for every assay.HIV-based lentiviral pseudoviruses with spike proteins were generated as previously described . The B.16 RLU/mL) for one hour at 37 \u00b0C and 100 \u03bcL of pseudovirus and soluble ACE2 mixture was added to 293T-ACE2.TMPRSS2 cells. Luciferase activity was measured 48 h later. The soluble ACE2 concentration causing a 50% reduction of RLU compared to control was reported as the 50% inhibitory concentration or IC50.For ACE2 neutralization assay, serially diluted recombinant human soluble ACE2 was incubated with indicated pseudovirus .4 cells per well on a 96-well plate. The cells were co-cultivated for 24 h at 37 \u00b0C. The culture supernatants were then removed, and cell-cell fusion was scored by determination of the \u03b2-gal activity in co-cultured cell lysates using a Galacto-Star kit  according to the manufacturer\u2019s instructions.For measuring spike-protein-mediated cell-cell fusion, \u03b2-gal complementation assay was performed as described previously . BrieflyTo ensure equivalent amount of spike cell surface expression levels among treatments, spike-transfected \u03b2-gal \u03c9 subunit-expressing 293T cells were quantified for cell surface spike levels by flow cytometry. Spike-transfected 293T cells employed in cell-cell fusion assay were concurrently stained with SARS-CoV-2 positive human polyclonal sera at 1:20 dilution, washed twice, and then incubated with FITC-conjugated goat anti-human . The cells were washed twice and then fixed with 2% paraformaldehyde. The results were acquired using BD LSRFortessa\u2122 X-20 Cell Analyzer . The mean fluorescence intensities of spike positive cells were recorded.https://github.com/acorg/Racmacs, accessed on: 15 October 2021) [We created a geometric interpretation of neutralization titers against the tested SARS-CoV-2 pseudoviruses using Racmacs antigenic cartography software (Sam Wilks (2021), Racmacs: R Antigenic Cartography Macros. R package version 1.1.16. er 2021) ,37. The http://www.cbs.dtu.dk/services/ProP/ (accessed on: 15 July 2021) and the PiTou V3 software hosted at http://www.nuolan.net/reference.html (accessed on: 15 July 2021).The prediction of furin-specific cleavage site in spike proteins was computed using the ProP 1.0 Server hosted at p values of less than 0.05 were considered statistically significant. All neutralization titers were log2 transformed for analyses.One-way analysis of variance (ANOVA) with Dunnett\u2019s multiple comparisons tests (variants compared to WT(D614G)), Mann\u2013Whitney test for the comparison of two groups with unmatched pairs (Pfizer BNT162b2 compared to Moderna) and geometric mean titers (GMT) with 95% confidence intervals were performed using GraphPad Prism software. The We first investigated the cross-neutralization potency of convalescent sera from individuals infected with SARS-CoV-2 in the U.S. against pseudoviruses bearing spikes of B.1.617.1 and B.1.617.2 variants and their corresponding RBD mutations A. TitersThe C.37 variant also has a substitution at L452 residue (L452Q instead of L452R) along with F490S in the RBD. A modest 1.8-fold reduction in titers against C.37 pseudoviruses was observed compared to WT(D614G) pseudoviruses . A 1.4-fold reduction in titers was observed for pseudoviruses with only the L452Q and F490S substitutions, indicating that these RBD substitutions contribute to C.37 resistance. These findings are in agreement with a prior report showing a 3.3-fold reduction of convalescent sera neutralization titer for C.37 pseudoviruses compared to WT(D614G), as well as L452Q and F490S single substitutions contributing to neutralization resistance . In compBecause the convalescent sera came from individuals who were previously infected by different variants , we alsoSera from the group infected by variants that had the L452R substitution showed similar fold changes in neutralization titers as the group infected by WT(D614G) variants without the L452R except against B.1.617.1 (B) and AY.1 pseudoviruses. Fold changes in neutralization titers against B.1.617.1 (B) and AY.1 variants were 3.3 and 1.3, respectively, for the L452 group, compared to 5.2 and 2.5, respectively, for the WT(D614G) group B,C. The p = 0.1225, Mann\u2013Whitney test). The Moderna mRNA-1273 vaccine-elicited sera trended towards higher neutralization titers against most variants compared to Pfizer/BioNtech BNT162b2, possibly due to higher vaccine mRNA content and greater interval between priming and boosting for Moderna mRNA-1273 (4 weeks vs. 3 weeks for Pfizer/BioNtech BNT162b2) [We next assessed the neutralization potency of mRNA vaccine-elicited sera against WT(D614G) and B.1.617 variant pseudoviruses. Sera from fourteen individuals who received two doses of Moderna mRNA-1273 vaccine and fifteen individuals who received two doses of Pfizer/BioNtech BNT162b2 vaccine were collected approximately two weeks after the second immunization. Each vaccine-elicited serum had high neutralization titers against WT(D614G) pseudoviruses, ranging between 578 and 3935 for Pfizer/BioNtech BNT162b2 and 651 and 5853 for Moderna mRNA-1273 A,B (PfizNT162b2) . CompareNT162b2) ,39.We also investigated the contribution of individual RBD substitutions of B.1.617.1  and B.1.617.2/AY.1  on the D614G background. Titers against L452R (GMT 935 for Pfizer and 1781 for Moderna) and E484Q (GMT 798 for Pfizer and 1429 for Moderna) alone trended slightly lower. Likewise, titers against K417N (GMT 1208 for Pfizer and 2070 for Moderna) and T478K (GMT 1046 for Pfizer and 2113 for Moderna) alone or in L452R + T478K combination (GMT 964 for Pfizer and 1796 for Moderna) remained comparable to GMTs of WT(D614G) .Similarly, GMTs against B.1.1.7 (955 for Pfizer and 1917 for Moderna) and B.1.429 variant with the L452R substitution (1063 for Pfizer and 1799 for Moderna) were comparable to those against WT(D614G). Consistent with previous observations, the B.1.351 variant (GMT 169 for Pfizer and 306 for Moderna) displayed ~7-fold lower titers compared to WT(D614G), whereas C.37, P.1, R.1, and B.1.526 variants displayed modestly reduced titers that are similar to the titers against B.1.617 pseudoviruses (GMT 452\u2013707 for Pfizer and GMT 824\u20131332 for Moderna). Overall, the trends in neutralization titers for the vaccine-elicited sera against a large panel of variant pseudoviruses were similar to those for convalescent sera, though the GMTs were approximately 3- to 5-fold higher for vaccine-elicited sera.A prior study showed that convalescent sera and vaccine-elicited antibody neutralization titers against pseudoviruses bearing spikes containing L452R-E484Q-P681R substitutions displayed 2\u20135-fold reduction, compared to the neutralization titers against WT(D614G) pseudoviruses . In thisn = 10,000 antigenic maps were made by resampling sera with replacement  pseudovirus and the WT(D614G) sera and L452R sera, including K417N, L452R, T478K, L452Q + F490S, and B.1.429 (0.25 to 0.72 antigenic units (AU) from WT . B.1.617The antigenic map of the same SARS-CoV-2 variant pseudoviruses for vaccine-elicited sera showed similar patterns to the convalescent sera map but with some notable differences. Again, pseudoviruses K417N, L452R, T478K, L452Q + F490S, and B.1.429 were close to WT(D614G) (0.05\u20130.51 AU). However, E484Q and C.37 were both closer to WT(D614G) but were poorly coordinated and extended in elongated shapes around WT(D614G). B.1.617.1 was also slightly closer to WT(D614G) at 1.55 AU and was positioned adjacent to the other variants. Unexpectedly, AY.1 was between B.1.617.2 and B.1.351 and was further away from WT(D614G) (1.89 AU) than in the convalescent sera map (1.60 AU), being positioned only slightly closer to WT(D614G) than B.1.351 was from WT(D614G) (3.02 AU). These antigenic maps reinforce what was observed in terms of neutralizing antibody titers. Compared to the convalescent sera map, there was a larger antigenic difference between AY.1 and WT(D614G) on the vaccine-elicited sera map.Notably, given that the vaccine-elicited sera had much higher titers than convalescent sera across variants, the titers against AY.1 were still higher in vaccinated individuals, meaning the antigenic difference may not translate into loss of vaccine protection. Furthermore, vaccine-elicited sera saw a smaller difference between B.1.351 and WT(D614G) than the convalescent sera, which may have aligned AY.1 and B.1.351 closer together. Overall, while these antigenic maps provide meaningful information on the relative positions of antigens, they are limited by the sera being so tightly clustered. Future antigenic maps with sera against distinct variants would enable more accurate evaluation of antigenic variation among the variants.We next evaluated 23 clinical-stage therapeutic neutralizing antibodies for potency against the B.1.617 variants. These antibodies were evaluated as part of the U.S. Government COVID-19 response effort to inform the clinical testing and use of these antibodies [B.1.617.1 pseudoviruses displayed complete resistance (>50-fold) to five nAbs  and partial resistance (10\u201350-fold) to one nAb (H) . The E48B.1.617.2 pseudoviruses displayed complete resistance (>50-fold) to three nAbs  and partial resistance (10\u201350-fold) to one nAb (H) B. The L4Prior reports indicated increased infectivity of pseudoviruses containing the L452R substitution in spike in 293T-ACE2.TMPRSS2 cells due to L452R conferring enhanced RBD affinity to ACE2 ,27. We t50) of sACE2 against B.1.617.2  was 4.2-fold lower, compared to WT(D614G) (IC50: 2.88 \u00b5g/mL)  and L452R + T478K  displayed 2.4- and 2.3-fold higher sensitivity to inhibition by sACE2. However, pseudoviruses with only spike RBD substitutions , as well as the B.1.617.1 (IC50:2.03 \u00b5g/mL) and AY.1 (IC50:1.97 \u00b5g/mL) spikes, displayed comparable IC50 to WT(D614G) (0.5\u20131.5-fold change)  . In agre change) . These fSince P681H enhanced proteolytic processing of B.1.1.7 spike ,49, we nTo gain further insight into the furin processing efficiency at the S1/S2 site of the B.1.617 S, we undertook a bioinformatic approach utilizing the PiTou and ProP furin cleavage prediction tools, comparing B.1.617 to the Wuhan-Hu-1 (D614) prototype spike and B.1.1.7 spike, as well as spike proteins of several lineage-specific mammalian and animal CoVs. The PiTou algorithm combines a hidden Markov model and knowledge-based cumulative probability score functions for the functional characterization of a 20 amino acid cleavage motif from P14 to P6\u2032 for furin binding and cleavage, whereas ProP predicts furin cleavage sites based on experimental data-derived networks ,54. Both679- R685 and E661- R685 have been reported to have host neuropilin-1 attachment [While SARS-CoV-2 S1/S2 P\u2212R\u2212R\u2212A\u2212R furin cleavage site conforms to a minimal furin recognition motif, R\u2212X\u2212X\u2212R, the presence of H/R instead of P increases the total number of basic residues to four. This presence of basic residue H/R results in additional electrostatic and intramolecular hydrogen bonding to gain substrate turnover . In facttachment  and staptachment .Furin cleavage score analysis of CoV S1/S2 cleavage sites. CoV spike and influenza hemagglutinin (HA) sequences were analyzed using the ProP 1.0 and PiTou 3.0 furin prediction algorithms, generating a score with bold numbers indicating predicted furin cleavage. \u201c|\u201d denotes the position of the furin cleavage site. Sequences corresponding to the S1/S2 region of NL63 (APF29063.1), 229E (ABB90529.1), SARS-CoV-2 (QHD43416.1), SARS-CoV (AAT74874.1), MERS-CoV (AFS88936.1), HCoV-HKU1 (AAT98580.1), HCoV-OC43 (APU51936) Bat-SL-CoVZC45 (AVP78031.1) BatCoV-HKU4 (YP_001039953.1), Influenza A/Chicken/Hong Kong/822.1/01/H5N1 HA (AF509026.2), Influenza A virus HA (A/Wisconsin/67/2005(H3N2) (ACF54576.1), infectious bronchitis virus (QIV13719.1), CoV-HKU9 (YP_001039971), BatCoV-PML (AGY29650), BatCoV-HKU5 (YP_001039962.1), Bat-CoV RaTG13 (QHR63300.2), and Bat-SL-CoVZXC21 (AVP78042.1) were obtained from GenBank. Sequences corresponding to the S1/S2 region of SARS-CoV-2 B.1.1.7 (EPI_ISL_1374509), B.1.617.1 (EPI_ISL_1841346), B.1.617.2 (EPI_ISL_2229775), B.1.617.3 (EPI_ISL_2157058), Bat CoV RmYN02 (EPI_ISL_412977), as well as HA of Influenza A virus ) (EPI1859607) were obtained from GISAID.To quantify whether higher ACE2 binding and furin cleavage of B.1.617.2 spike augments fusion between virus and/or cell membranes, we performed cell-cell fusion assays by complementing \u03b2-galactosidase subunits in spike-transfected effector cells and 293T-ACE2.TMPRSS2 target cells. Compared to WT(D614G), both B.1.617.1 and B.1.617.2 spikes induced significantly higher cell-cell fusion activity when controlled for spike cell surface expression (4000 MFI of spike protein on cell surface) B. Our fiHere we show that pseudoviruses bearing B.1.617.1 spike with L452R and E484Q substitutions, and B.1.617.2 spike with K417N, L452R and T478K substitutions, have modestly reduced susceptibility to neutralization by Pfizer/BioNtech BNT162b2 or Moderna mRNA-1273 vaccine-elicited sera and convalescent sera compared to pseudoviruses bearing WT(D614G) spike. The individual L452R, T478K, E484Q, and dual L452R + T478K substitutions accounted for most but not all of the reduction in neutralization potency of the sera, suggesting contributions from substitutions in the NTD/CTD. Neutralization titers, as well as antigenic maps, indicated that the full set of RBD substitutions in combination with substitutions outside the RBD contributes to antigenic differences of B.1.617.1, B.1.617.2, and C.37 variants. Antigenic distances between the variants also tended to be more spaced apart in the map generated by the convalescent sera compared to the vaccine-elicited sera. Limitations in our study include the small number of sera samples in the convalescent and vaccine cohorts. Potential differences in COVID-19 severity in the convalescent sera cohort and time of sera collection could also affect neutralization titers. Nonetheless, most sera from convalesced and vaccinated individuals neutralized the B.1.617.1, B.1.617.2, and AY.1 variants. Furthermore, 17 of 23 therapeutic neutralizing antibodies retained complete neutralization against B.1.617 variants. Resistance to the remaining therapeutic neutralizing antibodies is due to RBD substitutions, K417N, L452R, and E484Q, but not T478K. These findings suggest that the two-dose immunization with current mRNA vaccines will likely induce protective immunity against the tested B.1.617 variants. However, as B.1.617.2 variants continue to evolve, it will be important to continue to monitor how new substitutions in spike impact their resistance to therapeutic neutralizing antibodies and vaccine efficacy."}
+{"text": "With an aging population and increasing healthcare utilization, the frequency of hospital-acquired pneumonia (HAP) is expected to increase. Since HAP is life threatening, appropriate diagnosis and treatment are required; however, large-scale Japanese data focusing on patient profiles and treatment patterns is lacking.The demographics and treatment patterns of HAP were examined using a large-scale Japanese claims database from Jan. 2016 to Apr. 2018. The HAP population included patients who received injection antibiotics \u22673 consecutive days after admission, but not within 2 days after admission, and those whose reason for hospitalization was not pneumonia but had a diagnosis of pneumonia after hospitalization (based on ICD-10 codes).2,968 HAP patients  contributing 2,979 total HAP episodes were included. The 12-month pre-index mean Charlson Comorbidity Index (CCI) score was 4.0\u00b13.1 (mean\u00b1SD), CCI scores \u22674 comprised 44.0%. Most HAP episodes (77.6%) occurred \u22675 days after hospitalization. During the 12month pre-index period including outpatients, 84.9% of patients had some type of pneumonia record, 9.1% had VAP (ventilator associated pneumonia) records, and 7.4% had anti-MRSA prescription records. For post-index HAP treatment, ampicillin/sulbactam  and piperacillin/tazobactam  were frequently prescribed as the first antibiotic prescription. Ceftriaxone (19.4%) and meropenem (9.8%) were also frequently prescribed. Examinations prescribed during HAP: 30.5% blood culture tests, 28.2% sputum examinations and 29.2% urine antigen tests. The overall mortality rate of HAP in overall hospitalization post-index was 22.0%, in which 14.4% of deaths occurred within 30 days. The mean (\u00b1SD) length of overall hospital stay was 49.9 (\u00b134.2) days (11.3 days for HAP period), with 12.4% ICU use and 17.6% ventilator use. The median total cost during hospitalization was \u00a51,924,848.18 .The data revealed patient characteristics, treatment patterns, mortality rates and healthcare costs in Japanese HAP patients. This database approach should prove useful for discussing antibiotics usage trends in highly aging Japan.Masahiro Kimata, PhD, MSD K.K., Tokyo, Japan (Employee) Yosuke Aoki, MD, PhD, MSD K.K., Tokyo, Japan SHIONOGI & Co., Ltd  Adachi Noriaki, n/a, MSD K.K., Tokyo, Japan (Employee) Takeshi Akiyama, MSc, MSD K.K., Tokyo, Japan (Independent Contractor) Akiko Harada, n/a, MSD K.K., Tokyo, Japan (Employee)"}
+{"text": "The phase IIIb ATLAS-2M study demonstrated non-inferiority of long-acting (LA) cabotegravir (CAB) + rilpivirine (RPV) dosed every 8 weeks (Q8W) compared with every 4 weeks (Q4W) for maintenance of virologic suppression. Hepatitis C virus (HCV) co-infection occurs in ~6% of people with HIV due to shared modes of transmission. We report efficacy and safety of CAB + RPV LA in participants with HIV/HCV co-infection in ATLAS-2M.Participants with HIV-1 RNA < 50 c/mL receiving CAB + RPV LA Q4W (transitioned from ATLAS [NCT02951052]) or oral comparator ART were randomized 1:1 to receive CAB + RPV LA Q4W or Q8W. Baseline HCV RNA was assessed by polymerase chain reaction. Participants with symptomatic chronic HCV infection requiring treatment within 12 months or liver enzymes not meeting entry criteria were excluded. Week 48 assessments included proportion with HIV-1 RNA \u226550 and < 50 c/mL , general and hepatic safety, and pharmacokinetics.HIV/HCV co-infection was present in 10 (1%) of 1045 participants, 60% of whom were female at birth. At Week 48, 9/10 (90%) and 972/1035 (94%) participants with HIV/HCV co-infection and HIV mono-infection, respectively, had HIV-1 RNA < 50 c/mL . No participants with HIV/HCV co-infection had HIV-1 RNA \u226550 c/mL (vs 14/1035 [1%] with HIV mono-infection) or confirmed virologic failure through Week 48 (vs 10 [1%] with HIV mono-infection); 1/10 (10%) discontinued for reasons other than adverse events (AEs). Excluding injection site reactions (ISRs), AEs and serious AEs were reported in 4 (40%) and 0 participants with HIV/HCV co-infection, respectively; the only AE reported in >1 participant was injection site pain . In participants with HIV/HCV co-infection, all ISRs were grade 1/2; none led to withdrawal. No hepatic laboratory abnormalities were reported in participants with HIV/HCV co-infection through Week 48; rates were low in those with HIV mono-infection (Table). Plasma CAB and RPV concentrations were similar between groups.CAB + RPV LA was effective and well tolerated in this small cohort of participants with HIV and asymptomatic HCV co-infection.Ronald D\u2019Amico, DO, MSc, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Paul Benn, MB ChB FRCP, ViiV Healthcare (Employee) Shanker Thiagarajah, MB ChB, GlaxoSmithKline  Susan L. Ford, PharmD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Eileen Birmingham, MD, MPH, Janssen Research and Development  Ojesh R. Upadhyay, MPH, MBA, GlaxoSmithKline (Employee) Louise Garside, PhD, GlaxoSmithKline (Employee) Rodica Van Solingen-Ristea, MD, Janssen Research and Development (Employee)ViiV Healthcare (Employee) Kati Vandermeulen, M.SC., Janssen Research and Development (Employee) William Spreen, PharmD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee)"}
+{"text": "Scientific Reports 10.1038/s41598-021-99672-4, published online 11 October 2021Correction to: The Preprint section in the original version of this Article was omitted. The Preprint section now reads:58.\u201d\u201cThis article was submitted to an online preprint archiveAs a result Reference 58 was omitted, which is listed below.et al. Host Genetics Underlying Pathological Outcomes to Mycobacterium Avium Subsp. Paratuberculosis Infection is Governed by Distinct Genetic Variants. Preprint at 10.21203/rs.3.rs-668666/v1 (2021).58. Canive, M., The original Article has been corrected."}
+{"text": "This study examined physical activity (FITNESS) and social relationships (FRIENDS) on social engagement among community older adults. Members from two Florida aging-in-village programs participated. Three five-Likert scales were used: A 5-item FITNESS , 4-item FRIEND , and a 3-item social engagement scales  . Among the 96 participants, 79% were females, 91% were whites, 56% were married, 86% had college education, and 46% living alone. Mean age was 70.7 (SD=10.10). Participants reported at least 30-min. physical activity about 4.2 days per week. Overall social engagement was high (mean=4.38), FITNESS was median (mean=3.46), and FRINED was high (mean=4.19). FITNESS was significant to more 30-min. physical activity. Yet, higher FITNESS, FRIENDS, age, and volunteers were all significant to social engagement. Results has implications on promoting social engagement among older adults participating in aging-in-community programs."}
+{"text": "Staphylococcus aureus bacteremia (pMRSAB) is unclear. Vancomycin plus ceftaroline (V/C) has demonstrated potent in vitro synergistic activity against MRSA; however, clinical data is limited. Thus, we sought to evaluate V/C salvage therapy for pMRSAB.The preferred antibiotic salvage regimen for persistent methicillin-resistant \u2265 72 hours, received anti-MRSA monotherapy initially, and subsequently received V/C \u2265 24 hours. Patients were excluded if they received other anti-MRSA antibiotics within 72 hours of V/C initiation. The primary outcome was time to BC clearance following V/C initiation. Secondary outcomes included 90-day all-cause mortality, microbiological cure, 90-day MRSAB recurrence, and length of stay (LOS). Microbiological cure was defined as BC clearance.This was a single-center, retrospective cohort study of patients with MRSAB who received V/C salvage therapy between 1/1/2016-3/10/2021. Adult patients were included if blood cultures (BC) were positive for MRSA for Of 178 patients identified, 20 were evaluated after inclusion and exclusion criteria were applied. Mean (SD) age and Pitt Bacteremia score were 38.5 (14.5) years and 4.2 (3.1), respectively. Most patients were male (70%), intravenous drug users (65%), and admitted to the intensive care unit (65%). The most common source was intravenous drug use (55%) and the majority had infective endocarditis (70%). All patients received infectious disease consultation and median (IQR) vancomycin AUC:MIC was 527 . Source control, if possible, was obtained in most patients (55%). Median (IQR) time to bloodstream clearance from first positive BC and from when ceftaroline was initiated was 9.7  and 2.4  days, respectively. 90-day all-cause mortality, microbiological cure, and 90-day MRSAB recurrence occurred in 35%, 95%, and 5% of patients, respectively. Median (IQR) LOS was 25  days.To our knowledge, this is the largest cohort to evaluate V/C for pMRSAB. Patients were medically complex; however, median time to MRSAB clearance following ceftaroline initiation was < 2.5 days and microbiological cure was obtained in nearly all patients. V/C may represent a potential salvage regimen for pMRSAB.Wesley D. Kufel, PharmD, Melinta (Research Grant or Support)Merck (Research Grant or Support)Theratechnologies, Inc. (Advisor or Review Panel member) Jeffrey Steele, Pharm.D., Paratek Pharmaceuticals (Advisor or Review Panel member)"}
+{"text": "In this study, we characterized blaNDM-carrying plasmids in Enterobacterales isolated in Vietnam, and identified several possible cases of horizontal transfer of plasmids both within and among species of bacteria. Twenty-five carbapenem-nonsusceptible isolates from a medical institution in Hanoi were sequenced on Illumina short-read sequencers, and 13 blaNDM-positive isolates, including isolates of Klebsiella pneumoniae, Escherichia coli, Citrobacter freundii, Morganella morganii, and Proteus mirabilis, were further sequenced on an Oxford Nanopore Technologies long-read sequencer to obtain complete plasmid sequences. Almost identical 73 kb IncFII(pSE11)::IncN hybrid plasmids carrying blaNDM-1 were found in a P. mirabilis isolate and an M. morganii isolate. A 112 kb IncFII(pRSB107)::IncN hybrid plasmid carrying blaNDM-1 in an E. coli isolate had partially identical sequences with a 39 kb IncR plasmid carrying blaNDM-1 and an 88 kb IncFII(pHN7A8)::IncN hybrid plasmid in a C. freundii isolate. 148\u2013149 kb IncFIA(Hl1)::IncA/C2 plasmids and 75\u201376 kb IncFII(Yp) plasmids, both carrying blaNDM-1 were shared among three sequence type 11 (ST11) isolates and three ST395 isolates of K. pneumoniae, respectively. Most of the plasmids co-carried genes conferring resistance to clinically relevant antimicrobials, including third-generation cephalosporins, aminoglycosides, and fluoroquinolones, in addition to blaNDM-1. These results provide insight into the genetic basis of CRE in Vietnam, and could help control nosocomial infections.Carbapenem-resistant Enterobacterales (CRE) represent a serious threat to public health due to the lack of treatment and high mortality. The rate of antimicrobial resistance of Enterobacterales isolates to major antimicrobials, including carbapenems, is much higher in Vietnam than in Western countries, but the reasons remain unknown due to the lack of genomic epidemiology research. A previous study suggested that carbapenem resistance genes, such as the carbapenemase gene  The same integron cassette was observed on E. coli pHPA  isolated from a human in Japan in 2002 , which had partially identical sequences with two plasmids, pMH17-012N_1  and pMH17-012N_2 , both found in C. freundii MH17-012N (isolated in 2017) (E. coli pMH13-051M_1 [IncFII(pRSB107)::IncN plasmid] and C. freundii pMH17-012N_1  commonly carried blaNDM-1 and blaTEM-1b, and pMH13-051M_1 further carried blaCTX-M-55 and rmtB. C. freundii pMH17-012N_2 [larger IncFII(pHN7A8)::IncN plasmid] carried no known AMR genes, but did have a set of conjugation-associated type IV secretion system (T4SS) genes, which shared identical sequences with E. coli pMH13-051M_1. The genetic structures surrounding blaNDM-1 in E. coli pMH13-051M_1 and C. freundii pMH17-012N_1 contained MGEs, including ISAba125, ISCR21, and two IS26, and shared identical sequences with Tn125 (in 2017) . E. colith Tn125 : 98.6% ablaNDM-1\u2013carrying plasmids identified in Vietnam with those reported in other countries. A set of conjugation-associated T4SS genes in P. mirabilis pMH13-009N_1 [72.6 kb plasmid with IncFII(pSE11)::IncN replicons] was partially identical with Salmonella enterica FDAARGOS_70 plasmid unnamed1  isolated from a human in the United States in 2013, which carried blaTEM-1b (S. enterica FDAARGOS_70 plasmid unnamed1 did not carry blaNDM-1 as P. mirabilis pMH13-009N_1 did. C. freundii pMH17-012N_2 [87.6 kb plasmid with IncFII(pHN7A8)::IncN replicons and no AMR genes] was highly identical with E. coli p103-2-4  from a goose farm in China in 2018 and with S. enterica serovar Enteritidis p12367A  isolated from a human in China in 2013  shared nearly identical blaNDM-1-carrying plasmids, pMH15-289M_1, pMH16-398D_1, and pMH15-258M_1 , and all of these plasmids carried other AMR genes, such as blaCTX-M-15, blaOXA-1, rmtC, and qnrB9 ::IncA/C2 replicons, accession: CP021952, 99.8% identity in 82% region of pMH15-289M_1] in the United States and with E. coli pK71-77-1-NDM  isolated from a human in Norway in 2010. The plasmids tig00000169_pilon and pK71-77-1-NDM had identical sequences in the T4SS region of pMH15-289M_1, and carried blaNDM-1, blaCMY-6, and rmtC, and the plasmid tig00000169_pilon further carried blaSHV-11 and qnrB58. K. pneumoniae MH16-398D had 235 sequence variants , and K. pneumoniae MH15-258M had six sequence variants (five SNVs and one replacement) relative to K. pneumoniae MH15-289M.Furthermore, three nd qnrB9 . blaNDM-K. pneumoniae isolates belonging to ST395  shared nearly identical blaNDM-1-carrying plasmids, pMH15-208H_1, pMH15-191M_1, and pMH13-055M_1 , and all plasmids carried another AMR gene, rmtB  replicon, accession: CP043383, 99.9% identity in 98% region of pMH15-208H_1] isolated from a human in China in 2018 and with K. pneumoniae pSECR18-2374C  isolated from a human in South Korea in 2018. E. hormaechei pNDM1_045001 and K. pneumoniae pSECR18-2374C had a partially identical sequence of conjugation-associated T4SS genes with pMH15-208H_1. E. hormaechei pNDM1_045001 carried blaNDM-1, blaTEM-1b, and rmtB, and K. pneumoniae pSECR18-2374C carried blaNDM-4, blaTEM-1b and rmtB. K. pneumoniae MH15-191M had three sequence variants (two SNVs and one MNV) and K. pneumoniae MH13-055M had only one SNV relative to K. pneumoniae MH15-208H.Another three ne, rmtB . blaNDM-th Tn125 : 99.5\u201310E. coli, C. freundii pMH17-012N_1 (39.2 kb plasmid with IncR replicon), pMH17-012N_2 [87.6 kb plasmid with IncFII(pHN7A8)::IncN replicons], E. coli pMH13-051M_1 [111.5 kb plasmid with IncFII(pRSB107)::IncN replicons], K. pneumoniae pMH15-258M_1 [148.8 kb plasmid with IncFIA(Hl1)::IncA/C2 replicons], and K. pneumoniae pMH15-208H_1 [75.4 kb plasmid with IncFII(Yp) replicon] were not transferred to recipient E. coli under our experimental conditions though a set of T4SS-associated genes were detected in C. freundii pMH17-012N_2, E. coli pMH13-051M_1, K. pneumoniae pMH15-258M_1, and K. pneumoniae pMH15-208H_1.In bacterial conjugation experiments with K. pneumoniae to third-generation cephalosporins, carbapenems, aminoglycosides, and fluoroquinolones were 66.4%, 17.1%, 29.5%, and 53.0%, respectively, in Vietnam  according to the following protocol. The same amount of Luria-Bertani (LB) broth cultures of each donor bacteria and the recipient azide\u2013resistant E. coli J53 , were mixed and spotted onto Mueller-Hinton agar and then incubated at 37\u00b0C overnight. The mixed cells were recovered and suspended into PBS buffer, plated onto LB agar after 10-fold serial dilution, and incubated at 37\u00b0C overnight. Transconjugants were selected on LB agar containing 2 \u03bcg/mL of meropenem and 100 \u03bcg/mL of sodium azide.Bacterial conjugation was performed using six S1 TableAlso, sequence types of multilocus sequence typing (MLST) analysis determined from genomes, carbapenemase genes detected by ResFinder in genomes, sizes and contigs of genomes, coverages in short-read sequencing, Illumina sequencing platforms, and accession numbers of genomes are shown. According to the CLSI 2020 guidelines, Breakpoints of meropenem and imipenem are as follows: \u22641 \u03bcg/mL, susceptible; 2 \u03bcg/mL, intermediate; \u22654 \u03bcg/mL, resistant (R).(TIFF)Click here for additional data file.S2 TableAlso, replicon types detected by PlasmidFinder and antimicrobial resistance genes (ARGs) detected by ResFinder in plasmids, sizes of plasmids, and coverages in long-read sequencing, and accession numbers of plasmids are shown.(TIFF)Click here for additional data file."}
+{"text": "It has come to our attention that figures 1 and 3 in this article have been reproduced by the authors without first obtaining permission from the original publishers or authors:Figure 1: Akerfelt M, Morimoto RI, Sistonen L:\u00a0https://dx.doi.org/10.1038/nrm2938?utm_medium=email&utm_source=transaction Heat shock factors: integrators of cell stress, development and lifespan. Nat Rev Mol Cell Biol. 2010, 11:545-55.\u00a0https://dx.doi.org/10.1038/nrm2938 10.1038/nrm2938\u00a0Figure 3:\u00a0Pockley AG. https://www.cambridge.org/core/journals/expert-reviews-in-molecular-medicine/article/abs/heat-shock-proteins-in-health-and-disease-therapeutic-targets-or-therapeutic-agents/21095BFDB742E7876E4D1FDBF33E971C Heat shock proteins in health and disease: therapeutic targets or therapeutic agents? Expert Rev Mol Med. 2001, 3:1-21.\u00a0https://dx.doi.org/10.1017/S1462399401003556 10.1017/S1462399401003556.In addition, Figure 3 was also not properly attributed to the original source. This was later fixed via correction. After confirming with the publisher of Figure 1 that permission to republish was not obtained, and receiving no response from the authors despite multiple attempts, we have made the decision to retract this article and completely remove Figures 1 and 3 from the online and PDF versions of the article."}
+{"text": "Coronaviruses (CoVs) belong to the subfamily Orthocoronavirinae of the family Coronaviridae. CoVs are enveloped (+) RNA viruses with unusually long genomes. Severe acute respiratory syndrome CoV (SARS-CoV), Middle East respiratory syndrome CoV (MERS-CoV), and the novel coronavirus  have been identif ied as causing global pandemics. Clinically tested vaccines are widely used to control rapidly spreading, acute, and often severe infections; however, effective drugs are still not available. The genomes of SARS-CoV-2 and SARS-CoV are approximately 80 % identical, while the genomes of SARS-CoV-2 and MERS-CoV are approximately 50 % identical. This indicates that there may be common mechanisms of coronavirus pathogenesis and, therefore, potential therapeutic targets for each virus may be the same. The enzymes and effector proteins that make up the replication-transcription complex (RTC) of coronaviruses are encoded by a large replicase gene. These enzymes and effector proteins represent promising targets for potential therapeutic drugs. The enzyme targets include papain- and 3C-like cysteine proteinases that process two large viral polyproteins, RNA-dependent RNA polymerase, RNA helicase, viral genome-modifying enzymes, and enzymes with 3\u2019\u20135\u2019 exoribonuclease or uridylate-specif ic endonuclease activity. Currently, there are many studies investigating the complex molecular mechanisms involved in the assembly andfunction of the RTC. This review will encompass current, modern studies on the properties and complexes of individualnon-structural subunits of the RTC, the structures of individual coronavirus RTC subunits, domain organizationand functions of subunits, protein-protein interactions, properties and architectures of subunit complexes, the effectof mutations, and the identif ication of mutations affecting the viability of the virus in cell culture.Key words: non-structural proteins CoVs; subunits of replicase CoVs; replication-transcription complex of CoVs; architectureof non-structural protein complexes CoVs. The 2019 coronavirus infection has spread globally, oftencausing severe respiratory, intestinal, and systemic illnesses.Coronaviruses (CoVs) belong to the Orthocoronavirinaesubfamily of the Coronaviridae family. The subfamily isfurther divided into \u03b1-, \u03b2-, \u03d2-, and \u03b4-coronaviruses. Severeacute respiratory syndrome CoV (SARS-CoV), Middle Eastrespiratory syndrome CoV (MERS-CoV), novel coronavirus, mouse hepatitis virus (MHV),and bovine coronavirus (BCoV) are all \u03b2-coronaviruses . Coronaviruses are enveloped viruses with an unusuallylong, single-stranded (+) RNA genome (26\u201332 kb).The SARS-CoV-2 genome is similar to the SARS-CoVgenome (sequence identity ~80 %), while the SARS-CoV-2and MERS-CoV genomes are less similar (sequence identity~50 %) . The structure and function of proteinsare preserved at levels as low as 30 % of amino acid sequenceidentity . This indicates that there may be commonmechanisms of pathogenesis among the CoVs and, therefore,the viruses may have the same potential therapeutic targets.The 5\u2032 proximal region of each CoV genome includes a cap,a 5\u2032 untranslated region (UTR) and a long replicase gene encoding16 non-structural proteins (comprising two-thirds ofthe genome). The 3\u2032 regions encode structural proteins, includingS (spike), E (surface), M (membrane), and N (nucleocapsid),auxiliary proteins (the number of these varies amongCoVs), a 3\u2032 UTR, and a poly(A) tract.The replication-transcription complex (RTC) of CoVs is acomplex consisting of viral and, probably, cellular proteins.The RTC produces the (+) RNA genome and a set of subgenomicCoVs RNA in infected cells. The CoV replicase genehas two overlapping open reading frames, ORF1a and ORF1b,which encode the viral components of the RTC. Expression ofthe gene leads to the formation of the pp1a polyprotein, whichis encoded by ORF1a. A ribosomal frameshift of \u20131 before theORF1a translation termination codon and ORF1b are requiredfor the formation of the pp1ab polyprotein, which is a continuationof pp1a. Polyproteins pp1a and pp1ab are processed bytwo viral cysteine proteinases, papain-like proteinase PLpro(PLP) and 3C-like proteinase 3\u0421Lpro (Mpro), which results inthe release of intermediate precursors and 16 mature highlyconserved non-structural proteins (nsps) capable of associatingwith each other and being subunits of the RTC. The pp1apolyprotein includes nsp1\u2013nsp11, while pp1ab includes allpp1a nsps, as well as nsp12\u2013nsp16 .The molecular mechanisms of the assembly and function ofthe RTC has not been studied. However, the structural andfunctional properties of conserved non-structural RTC subunitsand their complexes have been extensively researchedand are extremely important for the identification of key drugtargets against CoVs:nsp1 interacts with the 40S ribosome subunit and inhibitstranslation initiation of host proteins, including interferonresponse factors. Interaction of nsp1 with ribosomes alsoleads to the degradation of host RNA. Thus, nsp1 suppressescellular defence antiviral mechanisms ;nsp2 is not part of the RTC in cell culture. However, theabsence of nsp2 in cells infected with the MHV\u0394nsp2 orSARS-CoV\u0394nsp2 deletion mutants reduces the productionof the virus and viral RNA ;nsp3 and nsp5 are proteinases that process the pp1a andpp1ab polyproteins, resulting in the release of individual RTCcomponents. The PLP nsp3 domain(s) process the N-proximalregions of pp1a and pp1ab. The MHV nsp3 has two domains,PL1pro (PL1P) and PL2pro (PL2P). The PL1P domain cleavesthe nsp1/nsp2 and nsp2/nsp3 sites, while the PL2P domaincleaves the nsp3/nsp4 site . The SARS-CoV nsp3 has a single PL2Pdomain that cleaves all three nsp sites .The SARS-CoV PLP is an intracellular immune responseantagonist. PLP blocks the activation of transcription factorsIRF3 and NF-\u03baB, which induces the expression of IFN(I)and antiviral genes. It does this by indirectly inhibiting IKKiand TBK1 kinases that activate IRF3 and stabilizing I\u03baB\u03b1, aninhibitor of NF-\u03baB . PLP also hydrolyzeselements of ubiquitin and the product of interferon-stimulatinggene 15 of the ubiquitin-like protein, thereby blocking thecellular mechanism of post-translational ubiquitination and,in turn, enhancing viral replication .However, nsp3 stabilizes the host E3 ubiquitin ligase RCHY1through the interaction of its SUD and PLP domains withRCHY1. This activates the RCHY1-mediated degradation ofp53, a cellular inhibitor of SARS-CoV replication . SARS-CoV nsp3 interacts with nsp5, nsp6,nsp12, nsp13, nsp14, and nsp16 in the yeast two-hybrid (Y2H)system and is thought to serve as a scaffold for RTC assembly;nsp5 is a 3CLpro (Mpro). Mpro plays a key role in the processingof pp1a and pp1ab polyproteins, cleaving the centraland C-proximal regions of pp1ab at 11 highly conserved sites,which releases mature nsp4\u2013nsp16 proteins . Mpro isonly active as a dimer. Self-elimination of MERS-CoV Mproat the nsp4/nsp5 and nsp5/nsp6 sites occurs as a result of theligand-induced formation of an \u201cimmature dimer\u201d during theconvergence of Mpro III domains within two polyproteins. Structural analysis of the SARS-CoV-2Mpro complexes with known antiviral inhibitors, Boceprevir(peptidomimetic NS3/4A protease of hepatitis C virus) and GC376 (inhibitor of CoV replication), revealed atomic-levelinteractions between Mpro and these inhibitors. Such studiesare important for the optimization and design of effectivedrugs against CoVs ;nsp6 interacts with nsp2, nsp8, and nsp9 in the Y2H system. Six of the predicted hydrophobic domainsof MHV nsp6 and SARS-CoV nsp6 are transmembrane domains. MHV nsp6 and SARS-CoV nsp6are localized in the membranes of the endoplasmic reticulum(ER) and induce the formation of autophagosomes from ERmembranes and activate autophagy .Co-transfection of nsp3, nsp4, and nsp6 induces a change inthe internal membranes of the host cell through the formationof double-membrane vesicles (DMVs), similar to the DMVsinduced by SARS-CoV ;and nsp8 co-crystallize to form the nsp7/8 hexadecamericsupercomplex. The assembly of the supercomplex involvesthe formation of two different nsp7/8 heterodimers, D1 andD2, which differ in nsp8 conformation. D1 and D2 eachdimerize to form the heterotetramers T1 and T2. The interactionof two T1 with two T2, in the order T1\u2013T2\u2013T1\u2032\u2013T2\u2032 andwith ring closure through the T1\u2013T2\u2032 interaction, leads to theconstruction of the full supercomplex. The supercomplex hasa unique architecture: 16 molecules (8 nsp7 molecules and8 nsp8 molecules) interact tightly with each other, forminga hollow cylindrical structure in which two nsp8 conformationscoexist. The positive charge of the inner channel of thecylinder and its diameter (30 \u00c5) indicates the ability of thensp7/8 supercomplex to surround and interact with doublestrandedRNA (dsRNA) . The SARS-CoVnsp7/8 hexadecameric supercomplex can be formed in solutionat an equimolar nsp7: nsp8 ratio .In solution, hexadecameric SARS-CoV nsp7/8 associateswith dsRNA (Kd ~1.2 \u03bcM). The association of nsp7/8 withdsRNA is mediated by nsp8 and enhanced by nsp7 . On its own, SARS-CoV nsp8 possesses primerindependentRNA-dependent RNA polymerase (RdRp)activity and initiates, with low fidelity, the de novo synthesisof short (less than 6 nucleotides) complementary oligomers(primers) on single-stranded RNA (ssRNA) templates . SARS-CoV nsp8 and nsp7/8 complex also exhibitprimer-dependent RdRp activity . TheFCoV (feline coronavirus) nsp7/8 complex is a 2 : 1 heterotrimerformed by the association of two nsp7 molecules and onensp8 molecule. This complex does not form a hollow structure,either in crystalline form or in solution. FCoV nsp7/8has primer-independent RdRp activity ;nsp9 is an ssRNA-binding protein . Both monomeric and dimeric forms ofPDCoV (porcine \u03b4 coronavirus) nsp9 and PEDV (porcineepidemic diarrhoea virus related to \u03b1 coronaviruses) nsp9, as well as the dimeric form of SARSCoVnsp9 , have been found in solutionduring in vitro experiments. Studies of the crystal structuresof SARS-CoV nsp9 ,PDCoV nsp9, and PEDV nsp9  haverevealed dimeric forms of nsp9. The monomer SARS-CoVnsp9 is characterized by a different structure compared toother proteins involved in the replicative complexes of RNAviruses, the features of which are similar to the structures ofoligosaccharide/oligonucleotide-binding proteins . Mutations affecting the dimerization of SARS-CoVnsp9 weaken its interaction with ssRNA, which is lethal forSARS-CoV replication in cell culture ;nsp10 interacts with dsRNA, dsDNA, and ssRNA withmicromolar affinity . Crystal structurestudies of SARS-CoV nsp10 showed that the monomerstructure includes two zinc fingers, a new discovery amongzinc finger protein structures. Motifs of zinc-binding nsp10sequences have been identified. Twelve identical monomersform a unique spherical dodecameric architecture, which ishypothesized to be the functional form of nsp10 . Through two-hybrid analysis in mammaliancells, interactions of SARS-CoV nsp10 with nsp14and nsp16 have been revealed ;nsp11 is a short peptide resulting from the cleavage of thepp1a polyprotein by the 3CLpro/Mpro proteinase at the nsp10/nsp11 site. Nsp11 is encoded by the region of genomic RNAwhere the translational reading frame shift occurs (ORF1a toORF1b). This shift results in the formation of nsp12\u2013nsp16proteins from the pp1ab polyprotein. SARS-CoV-2 nsp11contains 13 amino acid residues and has a disordered conformation,the dynamics of which have been studied in thepresence of lipid-membrane mimetics. In the presence of SDSmicelles, the disordered conformation of nsp11 is transformedinto an \u03b1-helix ;nsp14 is bifunctional. The N-terminal domain of nsp14has 3\u2032\u20135\u2032 exoribonuclease activity (ExoN), and its C-terminaldomain has (guanine-N7) methyltransferase activity(N7-MTase). ExoN corrects the low fidelity of synthesis ofthe complementary RNA strand by viral RdRp nsp12 andcatalyzes the removal of 3\u2032-terminal erroneous nucleotidesin dsRNA. N7-MTase catalyzes the methylation of the viralRNA cap at the N7 guanine position in the presence of S- adenosylmethionine(methyl group donor). N7-MTase has anS- adenosylmethionine binding motif which recognizes the capof viral GpppRNA and methylates guanine N7 GpppRNA toform 7MeGpppRNA (cap-0). Cap-0 plays an important role inblocking the degradation of viral RNA by 5\u2032\u20133\u2032 exoribonucleases,translation initiation, and immune system controlescape . Mutants of the catalyticmotif MERS-CoV ExoN and SARS-CoV-2 ExoN arenot viable in cell culture . SARS- CoVnsp10 associates with the ExoN domain of SARS-CoV nsp14,increasing the ExoN activity of nsp14 by more than 35 timeswithout affecting its N7-MTase activity .Structural studies of the SARS-CoV nsp10/14 complexshowed that one nsp10 molecule associates with the ExoNdomain of nsp14, stabilizing and enhancing the activity ofthe ExoN. The architecture of the nsp10/14 complex has beenstudied and has been found to include two regions of contactbetween the nsp10 molecule and the ExoN domain of nsp14;nsp16 has (nucleoside-2\u2032O) methyltransferase activity(2\u2032O-MTase). 2\u2032O-MTase recognizes the cap-0 of viralRNA and catalyzes the transfer of the methyl group fromS-adenosylmethionine to the 2\u2032OH group of the first nucleotide\u2019sribose after N7-methylated guanine, resulting in the formation of 7MeGpppN2\u2032OMe-RNA (conversion of cap-0 intocap-1). SARS-CoV nsp10 associates with nsp16 and stimulatesthe 2\u2032O-MTase activity of nsp16 .Mutagenesis mapping of the surface amino acid residues ofSARS-CoV nsp10 involved in nsp10\u2013nsp14 interaction andstructural analysis of the nsp10/16 complex revealed overlappingsurfaces of nsp10 interacting with nsp14 and nsp16.Nsp10 can serve as a platform that recruits nsp14 or nsp16to the RTC, stimulating the ExoN activity of nsp14 or the2\u2032O- MTase activity of nsp16. Therefore, nsp10 is an importantregulator of the RTC. Mutations have been identified thatdisrupt the nsp10\u2013nsp14 and nsp10\u2013nsp16 interactions, someof which lead to a nonviable viral phenotype in cell culture;nsp12, an RdRp, catalyzes the synthesis of complementaryRNA strands on (+) and (\u2013) viral RNA templates and is a keyCoV RTC enzyme. CoVs nsp12 initiates de novo synthesisfrom the 3\u2032 end of the viral genome of the full-length (\u2013) RNAstrand, as well as for subgenomic (\u2013) RNA transcripts, whichhave differing 3\u2032 end lengths. In turn, the full-length (\u2013) RNAstrands and subgenomic (\u2013) RNA strands serve as templatesfor the synthesis of the new RNA genome and subgenomic(+) RNA transcripts. Subgenomic (+) RNA is important forthe expression of structural and accessory proteins encodedby genes in the 3\u2032 proximal region of the viral genome, whichis inaccessible to ribosomes that translate the viral genome. Full-length recombinant SARS-CoVnsp12 associates with short (20\u201330 nucleotides long) dsRNAand ssRNA , and initiatesprimer-dependent RNA synthesis on both homo- and heteropolymericRNA templates of the same length . However, the recombinant SARS-CoV nsp12 does notassociate with a primer-template that mimics the 3\u2032-terminal40 nucleotides of the SARS-CoV UTR and does not exhibitRdRp activity on this primer-template .Cryo-electron microscopy determined the structure of themonomer SARS-CoV-2 nsp12, which includes the nidovirusspecificN-terminal domain of the RdRp-associated nucleotidyltransferase (NiRAN), the interface domain, and theC- terminal RdRp domain. The C-terminal RdRp domain hasa conserved right-hand-like architecture, which includes threesubdomains: fingers, palm, and thumb. The active centre isformed by conservative motifs of amino acid residues localizedin the palm subdomain. The motifs of the channel ofentry for nucleotide triphosphates and the primer-template andthe exit of the resulting RNA strand converge in the centralcavity,where these motifs carry out matrix-dependent RNAsynthesis have been determined .The association of the nsp7/8 complex with nsp12 leadsto the formation of the nsp7/8/12 complex. This complexpossesses high RNA-binding capacity, polymerase activity,and processivity. It is also capable of initiating de novo RNAsynthesis on the 3\u2032 UTR template of the SARS-CoV genome,resulting in the elongation of the RNA product by over300 nucleotides. For nsp7/8/12 complex-mediated initiationof processive RNA synthesis, three amino acid residues fromnsp7  and one amino acid residue from nsp8(K58) are required for the interaction of nsp7/8/12 with theRNA template, while four amino acid residues from nsp8 interact with nsp12 . Moreover, nsp7/8/12 is able to associate with nsp14to form the nsp7/8/12/14 multicomplex. This ensemble ofnon-structural proteins possesses high RNA polymeraseactivity and is involved in 5\u2032 RNA capping; however, it doesnot have ExoN activity . The structure ofthe SARS-CoV-2 and SARS-CoV nsp7/8/12 complexes wasdetermined by cryo-electron microscopy. These complexes,including the nsp12 monomer, nsp8 monomer, and nsp7/nsp8heterodimer, have similar architecture: the nsp8-1 subunitinteractswith the RdRp finger subdomain, while the nsp7and nsp8-2 subunits interact with the RdRp thumb subdomain;nsp13 possesses helicase and nucleoside-triphosphatase(NTPase) activities; nsp13 interacts with nsp7, nsp8, andnsp12 in the Y2H system .The crystal structure of MERS-CoV nsp13 was determined.The structure includes an N-terminal zinc-binding domain(ZBD) rich in Cys/His residues that coordinate three zinc ions,as well as C-terminal helicase RecA1 and RecA2 domainswhich contain parallel \u03b2-chains . The nsp13helicase separates dsRNA and dsDNA strands with overhanging(5\u201320 nucleotides long) 5\u2032 ends; nsp13 interacts with thesingle-stranded 5\u2032 end of the partial nucleic acid duplex ofdsRNA and dsDNA and unwinds it in the 5\u2032\u20133\u2032 direction, usingthe hydrolysis energy of the 5\u2032-deoxy- and ribonucleotidetriphosphates .The RNA 5\u2032-triphosphatase activity of nsp13 catalyzes thecleavage of the \u03d2 phosphate from the 5\u2032-terminal nucleotideof RNA and is thought to be involved in the capping of viralRNA . Studies of SARS-CoV nsp13 haveshown that the unwinding of DNA duplexes occurs in discreteintervals of ~9.3 base pairs (bp) at a rate of 30 intervals persecond. Therefore, the unwinding speed of DNA duplexes isapproximately 280 bp/s.The helicase activity of nsp13 increases approximately2-fold when nsp13 interacts with nsp12, suggesting theinteractionof these proteins is involved in the function ofthe RTC . When SARS-CoV-2 nsp13interacts with the RTC, specifically with nsp7/2nsp8/nsp12:RNA, the stable complexes 2nsp13\u2013RTC (67 %) and nsp13\u2013RTC (20 %), as well as the dimer (2nsp13\u2013RTC)2 (13 %) areformed. Using cryo-electron microscopy, the architecture ofthe dominant 2nsp13\u2013RTC complex has been determined.This involves the interaction of the ZBD on the first nsp13molecule with the N-terminus of nsp8b and the nsp12 thumbsubdomain, as well as the interaction of the ZBD of the secondnsp13 molecule with the N-terminus of nsp8a. The catalyticRecA1 helicase domain of the first nsp13 molecule is fixedagainst nsp7 and the nsp8b head ;nsp15 is a nidovirus uridylate-specific endoribonuclease(NendoU). It cleaves RNA at the 3\u2032 uridylate in unpaired,single-stranded, and looped regions . The crystal structures of SARS-CoV-2,SARS-CoV, and MERS-CoV nsp15 are homologous, functionallyactive hexamers formed by the dimerization of trimers.Each of the hexamer protomers includes three domains: theN- terminal, middle, and C-terminal catalytic NendoU domains.During trimer assembly, the N-terminal domain ofone protomer is packed into a gap between the central andC-terminal domains of the neighbouring protomer. During assembly of the hexamer, the N-terminal domains of the protomersof the two trimers are packed back-to-back and are inthe centre of the hexamer structure. The C-terminal domainscontaining active centres are located outward at the verticesof the cloverleaf. This architecture provides nsp15 with sixfunctionally active centres .The uridylate content in RNA, Mn+2 and, to a lesser extent,Mg+2 increase the affinity of nsp15 for RNA. The hexamericstructure of SARS-CoV and MERS-CoV nsp15 is criticalfor its substrate and catalytic activity. The study of a seriesof mono-, tri- and hexameric protein mutants revealed weakinteractions with RNA (rU16\u201320) and low catalytic activity inmonomers and trimers compared to hexamers and wild-typensp15 . Nsp8 andthe nsp7/8 complex interact with MERS-CoV nsp15, enhancingthe binding ability of the nsp15 hexamer for RNA and itscatalytic activity . The NendoU activity ofnsp15 is an antagonist of the IFN-induced cellular antiviralresponse and stimulates the initiation of viral RNA translation.A large number of non-structural subunits and their respectivecomplexes within the RTC is a defining feature of CoVs.An important feature is that some subunits have domains withdifferent enzymatic activities. For example, nsp14 has bothExoN and N7-MTase activity, while nsp13 has helicase andnucleoside triphosphatase activity. Non-structural proteinshave a set of activities universal for (+) RNA viruses: proteinases, RNA-dependent RNA polymerases (nsp12),RNA helicases (nsp13). There are also unique domains involvedin mRNA capping, cap modification .Some proteins possess 3\u2032\u20135\u2032 exoribonuclease activity (nsp14),which regulate the reliability of RNA genome replication,while some possess uridylate-specific endonuclease activity(nsp15). Many proteins serve as cofactors for importantenzymes  and affect cellular processes.In particular, some proteins suppress the antiviral cellularresponse . The nsp9 and nsp10structures are unique among the protein structures of the replicativecomplexes of RNA viruses. Many CoVs subunits andtheir complexes have a complex architecture. For example,nsp7 and nsp8 form a functional and unique hexadecamericsupercomplex, the nsp10 architecture includes 12 identicalsubunits, and the nsp15 architecture is a functionally activehexamer. The complex architecture of the RTC is defined bythe 2 nsp13/2 nsp8/1 nsp7/1 nsp12 model.The lack of effective drugs against the novel coronavirus infectionis a current global challenge. The RTC of CoVs replicates(+) RNA and determines the production of the virusin infected cells; however, the molecular mechanisms of thisremain unexplored. Currently, great efforts are being undertakenaimed at creating a structural or functional network ofinteractions within the CoV proteome, as well as its interactionswith the host cell. This would identify a large-scale panelof therapeutic targets. The determination of the structures ofindividual non-structural RTC subunits and their complexesand the identification of key interacting amino acid residuesand types of bonds between them will enable the design ofselective and effective inhibitors. Investigations employingbiochemical methods and mutational analyses can identifyfactors that affect the efficiency of viral genomic RNA productionand the virus in an infected cell, the multiple effects ofviral proteins on the host cell, and potential key drug targets.The authors declare no conflict of interest.Adedeji A.O., Lazarus H. 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+{"text": "Rescue ECMO has been used worldwide in patients (pts) with ARDS caused by COVID-19. Bacterial super-infections affect 3.5-14.3% of hospitalized pts with COVID-19. Pts requiring ECMO may be at an increased risk of infection due to their severity of illness, gut translocation and ECMO impact on host immunity.This was a retrospective review of pts requiring ECMO for COVID-19 from April 2020-2021 at a single center. Strict definitions of infections  were in accordance with CDC criteria. S. aureus (25%) and P. aeruginosa (16%) . Only 2% of all organisms were multi-drug resistant. 3 pts had fungal infections . Duration of ECMO was significantly longer for infected pts  vs , p=.01. 95% of infected pts had received steroids vs. 67% of uninfected pts, p=0.09. Treatment success at 1 week was 50%, and 24% and 40% of pts had recurrent infections and persistent/recurrent organisms in clinical cultures, respectively. S. aureus (54%) and Enterobacterales (26%) were associated with persistent or recurrent clinical cultures, requiring prolonged antimicrobial therapy. Mortality rate at 30 days was 65% and was significantly higher for pts with infection than those without . 43 ECMO pts with 1065 ECMO days were evaluated. Median age was 53 yrs (range: 21-62) and median BMI was 36.2 (range: 19.4-75.8). 70% were men and 65% were white. 37 patients (86%) experienced a total of 40 infectious episodes with a median onset from ECMO cannulation to first infection of 10.5d (range: 4-50). Median SOFA and SAPSII scores at time of infection were 12 (6-20) and 63 (30-90), respectively. PNA was the most common infection  . The most common organisms isolated were Enterobacterales (37%), S. aureus. Super-infection and mortality rates of ARDS pts on ECMO for COVID-19 were worse than for ARDS pts on ECMO for influenza at our center.Super-infection (most commonly PNA) occurred in almost all COVID-19 pts requiring ECMO for >4 days, and was a significant risk factor for death. Recurrent infections among survivors were common, especially when caused by Enterbacterales or Ryan K. Shields, PharmD, MS, Shionogi  Fernanda P. Silveira, MD, MS, FIDSA, Ansun  Involved: Self): Grant/Research Support; Novartis  Involved: Self): Grant/Research Support; Qiagen  Involved: Self): Grant/Research Support; Shire  Involved: Self): Advisor or Review Panel member, Grant/Research Support; SlieaGen  Involved: Self): Grant/Research Support; Whiscon  Involved: Self): Grant/Research Support Cornelius J. Clancy, MD, Merck (Grant/Research Support)"}
+{"text": "Brachypodium distachyon are still unclear.Mitogen-activated protein kinase (MAPK) cascades are involved with signal transduction in almost every aspect of plant growth and development, as well as biotic and abiotic stress responses. The evolutionary analysis of MAPKs and MKKs in individual or entire plant species has been reported, but the evolutionary patterns in the diverse inbred lines of B. distachyon. A total of 799 MAPKs and 618 MKKs were identified from 53 B. distachyon inbred lines. Remarkably, only three inbred lines had 16 MPKs and most of those inbred lines lacked MPK7-2 members, whereas 12 MKKs existed in almost all B. distachyon inbred lines. Phylogenetic analysis indicated that MAPKs and MKKs were divided into four groups as previously reported, grouping them in the same branch as corresponding members. MPK21-2 was the exception and fell into two groups, which may be due to their exon-intron patterns, especially the untranslated regions (UTRs). We also found that differential evolution patterns of MKK10 paralogues from ancient tandem duplicates may have undergone functional divergence. Expression analyses suggested that MAPKs and MKKs likely played different roles in different genetic contexts within various tissues and with abiotic stresses.We conducted the systematical molecular evolutionary analysis of MPK21-2 genes and the differential evolution of MKK10 paralogues with ancient tandem duplication might have functional divergences. Our findings provide new insights into the functional evolution of genes in closely inbred lines.Our study revealed that UTRs affected the structure and evolution of  They are divided into three highly-conserved subfamilies that continuously act in a sequential manner in evolution and fundamental signaling transduction pathways . The MAPion loop . Activation loop .Brachypodium distachyon (2n = 10) is an annual temperate grass with a close phylogenetic relationship to other temperate cereals and an intermediate position within the Pooideae subfamily  clade, a mostly Spanish (S+) clade, and a Turkish (T+) clade, based on their flowering phenotype and geographical substructure  (BrachyPan (https://brachypan.jgi.doe.gov/) (B. distachyon inbred lines. Collected sequences were only accepted for scanning using InterPro software (5-S/T within the activation loop for MKKs. The gene identifier information of these sequences was collected and is listed in We downloaded gene information for /plaza/) . BLASTP /plaza/)  searchesoe.gov/)  to identsoftware  if they MAPKs and MKKs was performed using Gene Structure Display Server 2.0 (GSDS 2.0) software (http://gsds.gao-lab.org/). All of the full-length amino acid sequences were initially aligned using Clustal Omega with default parameters . The domains and motifs of MAPKs and MKKs were scanned using InterProScan software (http://www.ebi.ac.uk/interpro/) (http://weblogo.threeplusone.com/).The exon/intron structure of identified terpro/) . The strB. distachyon inbred lines were generated in the BrachyPan project and visualized with the CorelDRAW X3 program. Phylogenetic trees were created based on the alignment of all MAPKs or MKKs using the maximum likelihood (ML) method with the Jones\u2013Taylor\u2013Thornton (JTT) model, 2,000 bootstrap values, and partial deletion by the MEGA 6.0 software, respectively (http://pdgd.njau.edu.cn:8080/)  .B. distachyon were collected at the early flowering stages according to their different flowering times /10 h dark (18 \u00b0C) photoperiods. We harvested the root, stem, leaf blade, and leaf sheath at the eight-to-nine leaf stage. Spikelet samples from ng times . For the\u2019s instructions. The quality of total RNA was detected using Nanodrop1000 and its integrity was estimated by electrophoresis in 1.5% (w/v) agarose gel. The real-time quantitative polymerase chain reaction (RT-qPCR) was carried out in 10 \u00b5l reactions with 5\u201350 ng of first-stand cDNA products (four \u00b5l), five pmol of each primer (0.4 \u00b5l), five \u03bcl SYBR green master mix (2X), 0.2 \u00b5l ROX as a passive reference standard to normalize the SYBR fluorescent signal. The conditions for RT-qPCR were: initial activation at 95 \u00b0C for 5 min followed by 45 cycles of 95 \u00b0C for 30 s, and 60 \u00b0C for 30 s. Subsequently, the specificity of PCR products was monitored using a melting curve analysis (61\u201395 \u00b0C with fluorescence read every 0.5 \u00b0C). The B. distachyon actin (gene locus: Bradi2g24070) gene was used as an internal control for all RT-qPCR analyses; specific primers for MAPK and MKK were listed in MAPK and MKK genes was calculated using the 2\u2212\u0394\u0394Ct method.Total RNA was extracted from samples using Trizol reagent and 1\u20132 \u03bcg was reverse-transcribed into cDNA using PrimeScript RT Master Mix Perfect Real Time  according to the manufacturerBrachyPan database , Bd3-1 (7), Adi-10 (10), Gaz-8 (5), ABR5 (11), Foz1 (11), and Jer1 (11)  . The incer1 (11) . Furtherf Adi-10 . We alsof Adi-10  and S2.B. distachyon inbred lines, the phylogeny of all identified 799 MPK protein sequences were performed using ML and NJ methods, respectively. As expected, all homologues for each of the 16 Bd21 MPKs (BdMPKs) were divided into four groups  and clustered on the corresponding branch except Tek-4MPK16  had similar clustering patterns with corresponding branches and fell into four groups: A, B, C, and D  and S5. B. distachyon inbred lines consistently contained 5, 10, and 8 introns, respectively  domain , the TXYectively , suggested lines  and S9. EY motif . MAPKs hEY motif  that binEY motif . MPK7-2sed lines . Moreoveed lines . Specified lines . Furtherinal end  and 5B. inal end . This muinal end . Remarkaeudicots .5-S/T motif (2\u20133X1\u20135L/IXL/I) in the N-terminal domain  had a part mutation in the phosphorylation site which coincided with a wide range of plant species (5-S/T), which were the most crucial amino acids in B. distachyon inbred lines, were 7.2 and 3.32, respectively, while the same results were 7.2 and 3.32 in Bd21  synthase gene was observed between two MKK members  presented tandem duplication in the canonical form of the MKK-DMRL-MKK model with occasional variations (PNN (pinin) gene instead of the DMRL gene between two MKK gene members  model in Mur1, MKK-DMRL-MKK-ChaC-MKK (ChaC: ChaC-like protein) model in BdTR12c, and MKK-DMRL-PK-PK-MKK (PK: protein tyrosine kinase) model in Sig2  and abiotic stresses  of erformed  to explo0-4 gene  and 8, wresearch . Among tn Bd30-1 . MPK3s hreatment . MKK10-3d Bd30-1 . Moreove in root . MKK4s hd Bd30-1 . MPK3s hopposite . Express tissues . Almost  tissues . We furtthe root  and BdTRstresses . The strMPKs and MKKs. Our results showed that the exon-intron architecture, including lengths and numbers of intron, intron phase, and lengths of UTR, was generally conserved in corresponding orthologs  instead of the DMRL gene (MKK10s may have contributed to gene expansion and function conservation and/or divergence during the evolution process of monocots.Our analysis suggested that occurred . These roccurred , indicatMRL gene . This maMRL gene  and prefMRL gene , 2012. TMAPK and MKK genes have been characterized with corresponding functions in plant growth and development. For instance, the expression levels of AtMKK10 are high in pollen but do not appear in shoot apices, mesophyll cells, or mature leaves (CaMPK19-2 genes are highly expressed in roots and stems in pepper, while CaMPK1 is highly expressed in in leaves (B. distachyon inbred lines. The result indicated that most MPK and MKK genes had quantitative distinct expression patterns among the three different genetic contexts in different tissues and various abiotic stresses. For example, MPK17 had higher expression levels in the root, stem, leaf blade, and salt treatment in Bd30-1 compared with Bd21 and BdTR8i (MPK17s, which may result from the nonsynonymous substitutions at some pivotal amino acid sites in EF-hand CBP motif in their C-terminal extensions (MKK10-3 and MKK10-5 had similar expression patterns in the leaf blade in Bd30-1 and BdTR8i and distinct profiles in Bd21 (MKK3-2 gene had similar patterns under heat and salt condition (MAPKs and MKKs had an expression divergence which was correlated with the differential evolution in B. distachyon inbred lines.Tissue-specific expression patterns of e leaves , indicate leaves . CaMPK19n leaves , which in leaves . We inved BdTR8i  and 8. Ttensions  as descrtensions . Moreove in Bd21 . These r in Bd21  and are  in Bd21 . Furtherondition . Taken tMPK and 618 MKK genes were retrieved from 53 kinds of B. distachyon inbred lines based on their conserved TXY or S/T-X5-S/T domain, respectively, using bioinformatics approaches. Phylogenetic analyses showed that most MAPKs and MKKs clustered into same branch, with the exception of MPK21-2s, which was divided into two groups, designated as type I and II. Further analysis found that the divergence of MPK21-2 may be involved with the presence of UTRs. MKK10s expanded during the evolutionary process by ancient tandem duplications with a differential model. This may have resulted in expression differences and functional divergence. We discovered that the expression of the MPK and MKK gene members varied in different tissues and across abiotic stresses in three different genetic contexts, suggesting that these genes may have diverse biological functions. Taken together, our results revealed a more comprehensive understanding of the function and evolutionary patterns of MAPKs and MKKs in diverse B. distachyon inbred lines.A total of 799 10.7717/peerj.11238/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.11238/supp-2Supplemental Information 2Click here for additional data file.10.7717/peerj.11238/supp-3Supplemental Information 3Five MPKs including Gaz-8MPK4, Kah-1MPK20-4, Mon3MPK7-1, Tek-4MPK16, Tek-4MPK20-1, were excluded from entire sequences for reconstructing NJ tree.Click here for additional data file.10.7717/peerj.11238/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.11238/supp-5Supplemental Information 5Nine MKKs including Adi-10MKK5, Bd3-1MKK4, BdTR10cMKK10-5, Bd29-1MKK4, BdTR12cMKK3-1, BdTR5iMKK5, BdTR9kMKK10-5, Tek-4MKK3-1, Tek-4MKK3-3, were excluded from entire sequences for reconstructing NJ tree.Click here for additional data file.10.7717/peerj.11238/supp-6Supplemental Information 6Click here for additional data file.10.7717/peerj.11238/supp-7Supplemental Information 7Click here for additional data file.10.7717/peerj.11238/supp-8Supplemental Information 8Click here for additional data file.10.7717/peerj.11238/supp-9Supplemental Information 9Click here for additional data file.10.7717/peerj.11238/supp-10Supplemental Information 10Click here for additional data file.10.7717/peerj.11238/supp-11Supplemental Information 11Click here for additional data file.10.7717/peerj.11238/supp-12Supplemental Information 12Click here for additional data file.10.7717/peerj.11238/supp-13Supplemental Information 13Click here for additional data file.10.7717/peerj.11238/supp-14Supplemental Information 14Click here for additional data file.10.7717/peerj.11238/supp-15Supplemental Information 15Click here for additional data file.10.7717/peerj.11238/supp-16Supplemental Information 16Click here for additional data file.10.7717/peerj.11238/supp-17Supplemental Information 17Click here for additional data file.10.7717/peerj.11238/supp-18Supplemental Information 18Click here for additional data file.10.7717/peerj.11238/supp-19Supplemental Information 19Click here for additional data file."}
+{"text": "Dear Editor,With increased survival of patients with multiple myeloma (MM), therapy-related myelodysplastic syndrome (t-MDS) and t-acute myeloid leukemia (AML) may occur more frequently , 2. We pIn June 2015, a 64-year-old female was diagnosed with IgG kappa (\u03ba) MM. IgG levels were 46g/L, \u03ba-serum-free light chains (SFLC) 75.4mg/L and \u00df2-microglobulin 8.2mg/L Fig. . Anemia 6/L). BM assessment did not reveal increased PCs or MDS, and serological parameters indicated stable disease.First-line therapy with bortezomib, cyclophosphamide, and dexamethasone was followed by autologous stem cell transplantation and maintenance therapy with lenalidomide Fig. . After 16/L, platelets 12x106/L) and \u03ba-SFLCs increased .Due to frailty at that time, she was ineligible for intensive AML induction therapy. Therefore, treatment with decitabine/venetoclax was started in February 2019. A BM biopsy in March 2019 confirmed CR of the t-AML Fig. , right; ted Fig. . This inIn June 2019, after worsening pancytopenia re-emerged and myeloid blasts were detectable in PB smears, decitabine/venetoclax was re-initiated. The BM biopsy in August 2019 showed persisting (30%) immature myeloid blasts ."}
+{"text": "Correction to: J Exp Clin Cancer Res 41, 18 (2022)https://doi.org/10.1186/s13046-021-02203-2Fig.\u00a07i: plotting errors in the histogram; the histogram has been correctedFig. S8d: western blots (right hand side) replaced with correct blotsFig. S11: raw western blots presented for Fig.S8d have been replaced with correct blotsFollowing publication of the original article , the autThe corrected figure is given here. The correction does not have any effect on the final conclusions of the paper. The original article has been corrected.Additional file 1: Fig. S8 mTORC1\u2019s inhibitory effect on HuD. A. Comparison between mouse neuroblastoma and neurons under stress; Akt-mTOR pathway examined by Western blot assay. Full-length blots are presented in Supplementary Figure S11. B. Viability assay (control or miR375 mimic or miR375 inhibitor) in IMR-32 cells. C. Relative mRNA expression quantified by RT-qPCR (control or miR375 inhibitor and/or active mTOR via Rheb S16H construct and/or inactive mTOR via rapamycin-25 nM) in IMR-32 cells. D. Western blot analysis of HuD and pS6 in IMR-32 cells. Full-length blots are presented in Supplementary Figure S11. Data are presented as mean \u00b1 SEM; t test: *p < 0.05, **p < 0.01, ***p < 0.001. Fig. S11 Raw Western blot images for Fig. S5, S6 and S8."}
+{"text": "On January 30, 2020, the World Health Organization (WHO) declared the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic a worldwide emergency.\u00a0Worldwide there have been 170 million cases of the resulting disease coronavirus 2019 (COVID-19), of those, 3.53 million have resulted in death. The Food and Drug Administration (FDA) with Mayo Clinic as the lead institution authorized COVID-19 convalescent plasma (CCP) for treatment of SARS-CoV-2 infection. Effective therapeutic window for CCP administration had yet to be defined. We addressed this gap by characterizing longitudinal biologic response and clinical outcomes of COVID-19 patients treated with CCP. Primary outcome was discharged to home/home health. On January 30, 2020, the World Health Organization (WHO) declared the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic a worldwide emergency.\u00a0Worldwide there have been 114 million cases of the resulting disease coronavirus 2019 (COVID-19) of those, 2.54 million have resulted in death. At pandemic debut, available treatment was limited to supportive care as no approved therapy or vaccination was available. This treatment vacuum motivated the utilization of convalescent plasma infusion to complement the antibody response.Passive immunization has been successfully used to treat infectious diseases since the 1890s . ConvaleResults from small case series conducted during the prior Middle East Respiratory Syndrome (MERS) and SARS-CoV documented CP to be safe, well-tolerated and promoted faster viral clearance, particularly when given early in the disease course . The priThe Food and Drug Association (FDA) with Mayo Clinic as the lead institution authorized COVID-19 convalescent plasma (CCP) for treatment of CoV-2 infection. Effective therapeutic window for CCP administration had yet to be defined. We addressed this gap by characterizing longitudinal biologic response and clinical outcomes of COVID-19 patients treated with CCP.\u00a0\u00a0Study design and populationThis retrospective longitudinal study analyzed electronic medical record data, including but not limited to characteristics and laboratory test findings from 197 patients consecutively admitted between March 28 and August 5, 2020. Among those, 92 and 105, respectively, received CCP infusion within 48h of versus 48h after hospitalization. Primary outcome was discharged to home/home health and secondary outcome was longitudinal CRP levels post CCP infusion.\u00a0Sarasota Memorial Hospital Institutional Review Board authorized consenting COVID-19 inpatients to participate in the national CCP protocol. Written informed consent was obtained from every CCP recipient or their legal guardian. Internal medicine resident physicians identified, contacted, and facilitated logistical pathways for CCP donation in collaboration with the community blood bank.Enrolled patients were at least 18 years old with laboratory confirmed SARS-CoV-2 infection admitted for treatment of severe or life-threatening COVID-19. Severe disease was defined as the presence of at least one of the following characteristics: dyspnea, respiratory rate of 30 breaths per minute or more, oxygen saturation (SpO2) equal to or less than 93%, partial pressure of arterial oxygen to fraction of inspired oxygen less than 300 or development of lung infiltrates with more than 50% involvement within 24-48 hours (h). Life-threatening disease was defined as the development of at least one of the following: respiratory failure, septic shock or multiple organ dysfunction or failure.Data analysisAnalyses contrasted patients who underwent CCP infusion within or more than 48h after admission. Primary outcome was discharged to home/home health. Continuous data summarized as median (interquartile range\u00a0[IQR]) were compared using Kruskal-Wallis test or two-way analysis of variance (ANOVA). Discrete data were compared with Pearson\u2019s chi-square test. Two-tailed p < 0.05 was significant.2, p < 0.0001)  (32.7 (27-40)) vs. (29.4 (26-37) kg/mInitial admission vital signs and laboratory test results were not different between groups (p>.05) including temperature (98.4 (98.0-99.0) \u00b0F); SpO2  %); C-reactive protein (CRP) (2.6 (0.3-2.7) mg/dL); D-dimer (1.01 (0.69-2.03) mg/L); and ferritin (624 (281-1228) ng/mL)  vs. 81.7 (61.3-130.0) hours who respectively received CCP infusion within vs after 48h, p < 0.0001. Admission lactate dehydrogenase (LDH) was (366 (301-469) U/L) vs. (326 (274-428) U/L, p = 0.02)  vs. 29/105 (28%), p = 0.34. Days of mechanical ventilation were 8.0 (5.0-11.8) vs. 11.3 (4.5-17.9), p = 0.16. Hospital length of stay was 8.5 (4.9-15.2) vs. 13.0 (6.5-18.9) days, p = 0.03  vs. 49/105 (46%), p = 0.005.Between March 28 and August 5, 2020, Sarasota Memorial Hospital participated in the Mayo Clinic-led national FDA expanded access program providing access to convalescent plasma protocol. Throughout this period, data from 197 participants were analyzed and contrasted among those who received CCP infusion within or more than 48h after admission with a confirmed COVID-19 infection. We observed a post-CCP reduction in C-reactive protein, lower hospital length of stay and increase in discharge directly from hospital to home/home health in patients who received CCP within 48h of admission rather than later in hospitalization. Moreover, earlier CCP treatment resulted in 7% fewer patients who died or were discharged to hospice.In December 2019, a new member of Coronaviridae family, SARS-CoV-2 was detected in Wuhan, China primarily manifesting as a respiratory illness . Since tInitial evidence demonstrated CCP to be most beneficial when administered soon after SARS-CoV-2 infection. A randomized, double-blinded, placebo-controlled trial evaluated disease progression in 80 patients who received convalescent plasma within 72h after onset of mild COVID-19 symptoms vs. placebo ,11,12. ACCP transfusion is associated with a reduction in inflammatory markers, such as CRP ,10,14. IOur data should be interpreted with some caveats. We conducted a monocenter pragmatic investigation. CCP was administered before FDA required titer labeling. Therefore, we couldn\u2019t establish if antibody titers in CCP transfused across patient groups were equivalently distributed. In addition, SARS-CoV-2 serologic testing was unreliable restricting the assessment of whether a patient exhibited an impaired humoral response.Our study was conducted while no anti-viral treatment was approved by the FDA for patients hospitalized with COVID-19. We evince CCP treatment within 48h of admission was associated a reduction in hyperinflammation and hospital length of stay in patients more obese with higher LDH levels\u00a0with greater benefit for discharge to home/home health benefit and reduction in composite outcome of hospital mortality or discharge to hospice. Convalescent plasma has shown to be an effective treatment when given soon after SARS-CoV-2 infection. Unfortunately, in hospitalized patients with COVID-19, infusion of CCP late in the course of illness provides no observed benefit, as reported with other anti-viral agents."}
+{"text": "Listeria monocytogenes strains that were isolated from the invasive alien snail species Arion vulgaris in Austria in 2019.We report the draft genomes of two  Listeria monocytogenes is widespread in the environment, living as a saprophytic organism in the plant-soil compartment  DNA kit . Paired-end sequencing (2 \u00d7 300 bp) was performed with a MiSeq platform  as described . LibraryDefault parameters were used for all software unless otherwise specified. Raw reads were quality controlled using FastQC v0.11.9. Trimmomatic v0.36 . ContigsL. monocytogenes isolates, S-01 and S-02, generated 1,212,448 and 1,407,694 reads, respectively, with mean coverages of 101- to 113-fold and GC contents of 37.9% (L. monocytogenes. Strains were characterized by multilocus sequence typing (MLST) (https://www.cgmlst.org), respectively (Whole-genome sequencing (WGS) of the two of 37.9% . The NCBof 37.9% . Mash diectively .Listeria monocytogenes WGS project has been deposited in DDBJ/ENA/GenBank under BioProject no. PRJNA716154 and accession no. JAGGDY000000000 (S-01) and JAGGDX000000000 (S-02) (first versions). The raw sequence reads have been deposited in the Sequence Read Archive (SRA) under accession no. SRR14027493 (S-01) and SRR14027492 (S-02).The"}
+{"text": "Next-generation sequencing (NGS) of whole genomes has become more accessible to biomedical researchers as the sequencing price continues to drop, and more laboratories have NGS facilities or have access to a core facility. However, the rapid and robust development of practical bioinformatics pipelines partly depends on convenient access to data for the testing of algorithms. Publicly available data sets constitute a part of this strategy.Here, we provide a triplicate whole-genome paired-end sequencing data set, consisting of 1.38 billion raw sequencing reads derived from saliva DNA from a single anonymous male Caucasian donor, with the average sequencing depths aimed at 30x for two of the samples and 4x for a low-coverage sample. The raw number of single nucleotide variants were 3.3\u20134 million and the median variant read depth of GATK4-passed variants in three samples was 22, 18, and 10. 81% of all variants were found in two or three of the samples, whereas 19% were singletons. The karyotype was evaluated as 46,XY with no apparent copy-number variation.The data set is provided without restrictions for research, educational or commercial purposes. Specifications TableValue of the Data\u2022The data set provided here is relevant for the continued development and testing of bioinformatics pipelines as whole-genome sequencing become more important in biomedical research.\u2022Data access is provided by simple download and without restrictions. The triplicate sequencing of a Caucasian male may benefit bioinformaticians, biomedical researchers for testing or as control samples. The data may also be used for educational purposes.\u2022The raw sequencing data consists of biological replicates of low, medium, and higher coverage, which thus may be used for testing different workflow setups.1stats), and approximately 93.4% paired and mappable reads (GRCh37). The combined theoretical mean coverage was estimated to be 63\u201367x, depending on whether the unadjusted or mapped percentage was implemented, using the Lander and Watermann approach C\u00a0=\u00a0L*N/G . Calculations were based on an average read length of 144, 146, and 148 bp. The median GATK-passed variant read depths of the three were 22, 18, and 10 with 3.3\u20134 million variants  over the time span of approximately one year from replicate 1 to 3. For the assessment shown here, alignment implemented Burrows-Wheeler Aligner Used commandline workflowfn=${1%_L001_R1_001.fastq.gz}bwa mem -M -R ``@RG\\tID:group1\\tSM:$1\\tPL:illumina\\tLB:lib1\\tPU:unit1'' -t 23 hg38/Homo_sapiens_assembly38.fasta $1 ${1%_L001_R1_001.fastq.gz}_L001_R1_001.fastq.gz | samtools view -@23 -m 1G -Sb -> ${1%_L001_R1_001.fastq.gz}.bamgatk MarkDuplicatesSpark \\-I $fn.bam \\-O $fn.dedup.bam \\\u2013remove-sequencing-duplicatesgatk BaseRecalibrator \\-I $fn.dedup.bam \\-R hg38/Homo_sapiens_assembly38.fasta \\\u2013known-sites hg38/1000G_phase1.snps.high_confidence.hg38.vcf.gz \\\u2013known-sites hg38/Homo_sapiens_assembly38.dbsnp138.vcf \\\u2013known-sites hg38/1000G_phase1.snps.high_confidence.hg38.vcf.gz \\-O $fn.recal1.tablegatk ApplyBQSR \\-I $fn.dedup.bam \\-R hg38/Homo_sapiens_assembly38.fasta \\\u2013bqsr-recal-file $fn.recal1.table \\-O $fn.dedup.recal.bamgatk HaplotypeCaller \\\u2013native-pair-hmm-threads 23 \\-R hg38/Homo_sapiens_assembly38.fasta \\-I $fn.dedup.recal.bam \\\u2013dbsnp hg38/Homo_sapiens_assembly38.dbsnp138.vcf \\-O $fn.mnvs.vcfgatk SelectVariants \\-R hg38/Homo_sapiens_assembly38.fasta \\-V $fn.mnvs.vcf \\\u2013select-type-to-include SNP \\-O $fn.SNP.vcfgatk VariantRecalibrator \\-R hg38/Homo_sapiens_assembly38.fasta \\-V $fn.SNP.vcf \\\u2013resource:hapmap,known=false,training=true,truth=true,prior=15.0 hg38/hapmap_3.3.hg38.vcf \\\u2013resource:omni,known=false,training=true,truth=true,prior=12.0 hg38/1000G_omni2.5.hg38.vcf \\\u2013resource:1000G,known=false,training=true,truth=false,prior=10.0 hg38/1000G_phase1.snps.high_confidence.hg38.vcf \\\u2013resource:dbsnp,known=true,training=false,truth=false,prior=2.0 hg38/Homo_sapiens_assembly38.dbsnp138.vcf \\-an QD -an MQ -an MQRankSum -an ReadPosRankSum -an FS -an SOR -an DP \\-mode SNP \\-O $fn.output.recal \\\u2013tranches-file $fn.output.tranches \\\u2013rscript-file $fn.output.plots.Rgatk ApplyVQSR \\-R hg38/Homo_sapiens_assembly38.fasta \\-V $fn.SNP.vcf \\-O $fn.SNP.filtered.vcf \\\u2013truth-sensitivity-filter-level 99.5 \\\u2013tranches-file $fn.output.tranches \\\u2013recal-file $fn.output.recal \\-mode SNPInformed consent was obtained concerning the donation of biological material and genomic information. Sequencing was part of a technology assessment using anonymous donor material and does not involve any clinical evaluations or trials. Data is made freely available in order to contribute to the continued development of NGS bioinformatics and for educational purposes.Marcus H\u00f8y Hansen: Conceptualization, Methodology, Software, Validation, Formal analysis, Investigation, Resources, Data Curation, Writing, Editing, Visualization, Supervision, Project administration, Funding acquisition; Charlotte Guldborg Nyvold: Writing, Editing, Supervision.It is not a regular scientific paper.The authors declare that they have no known competing financial interests or personal relationships which have or could be perceived to have influenced the work reported in this article. Funding was provided by the first author. Disclaimer: The provided data presentation is deliberately descriptive."}
+{"text": "Klebsiella pneumoniae, especially the hypervirulent blaKPC-positive K. pneumoniae (Hv-blaKPC(+)-KP).To investigate the overall distributions of key virulence genes in K. pneumoniae from GenBank were collected and analyzed. Multilocus sequence typing, molecular serotyping, antibiotic-resistance, virulence genes and plasmid replicon typing were investigated.A total of 521 complete genomes of c-rmpA/A2) to 99.6% (entB). Totally 207 strains presented positive fimH, mrkD, entB and wzi and 190 showed positive fimH, mrkD, entB, irp2 and wzi, which were the two primary modes. A total of 94, 165 and 29 strains were denoted as hypervirulent K. pneumoniae (HvKP), blaKPC(+)-KP and Hv-blaKPC(+)-KP. ST11 accounted for 17 among the 29 Hv-blaKPC(+)-KP strains; Genes iucA, p-rmpA2 and p-rmpA were positive in 28, 26 and 18 Hv-blaKPC(+)-KP strains respectively. Among the 29 Hv-blaKPC(+)-KP strains exhibiting four super clusters from GenBank, IncHI1B plasmids carrying virulence genes and IncFII ones with blaKPC were responsible for both 23 strains respectively.Positive rates of virulence genes highly varied, ranging from 2.9 -KP. IncHI1B plasmids carrying virulence genes and IncFII ones with blaKPC constitute the primary combination responsible for Hv-blaKPC(+)-KP. The making of Hv-blaKPC(+)-KP is mostly via blaKPC(+)-KP acquiring another plasmid harboring virulence genes.Positive rates of virulence genes vary remarkably in  Klebsiella pneumoniae, a ubiquitous and an opportunistic pathogen, can induce both nosocomial and community-acquired infections (K. pneumoniae inducing such \u201cinvasive syndrome\u201d is termed as hypervirulent K. pneumoniae (HvKP), which is more virulent than \u201cclassical\u201d K. pneumoniae (cKP) typically responsible for nosocomial infections , New Delhi metallo-\u03b2-lactamase gene (blaNDM), and oxacillinases-48 gene (blaOXA-48), which are predominantly carried on the mobile genetic elements \u00a0has now become a great public health threat worldwide  gained more and more prevalence with its positive rate reaching 7.4\u201315.0% among CRKP in recent years -KP), was rarely reported. Here, we collected 521\u00a0K. pneumoniae strains from GenBank. Upon the yielded data, we could get insight into the distributions of key virulence genes in K. pneumoniae, particularly Hv-blaKPC(+)-KP.In the past decades, hypervirulence and drug-resistance advance separately in virulent . Howevervirulent . Not surortality . Due to nt years . To dateK.\u00a0pneumoniae from the GenBank Database  were analyzed in this study. Those draft genomes (contigs and scaffolds) were not included. The 521 strains included 28.4% (148 strains) from Mainland China, 4.4% (23 strains) from Taiwan of China, 1.5% (eight strains) from Hong Kong of China, 25.7% (134 strains) from USA, 9.6% (50 strains) from Australia, 6.7% (35 strains) from UK, 3.8% (20 strains) from Germany, 2.7% (14 strains) from Korea, 2.3% (12 strains) from India, 2.1% (11 strains) from France, 1.5% (eight strains) from Japan and 11.1% (58 strains) from other countries.A total of 521 complete whole genomes Table S1K. pneumoniae MLST database  and the STs were yielded.The DNA fasta sequences of the 521 genomes were compared with the database  containiK. pneumoniae from GenBank, the accession numbers were directly used to determine the capsular types via the database of Institute Pasteur (https://bigsdb.pasteur.fr/klebsiella/klebsiella.html). The potential beta-lactamase genes were determined using the Resfinder software version 3.2 (https://cge.cbs.dtu.dk/services/ResFinder/)   with thesupports Table S2peg-344), colonization , assembling channel protein for capsular polysaccharides or macromolecular exopolysaccharides , regulator of mucoid phenotype , Type 1 fimbriae (fimH), Type 3 fimbriae (mrkD), enterobactin (entB), yersiniabactin (irp2), salmochelin (iroN), and aerobactin (iucA) and capsular polysaccharide-anchor (wzi).For virulence genes in this study, they could be classified as the following categories: metabolism , EPS (by wzy-K1) and excessive siderophores , or \u22651 positive capsule-regulating genes . Non-HvKP is termed as cKP. Hv-blaKPC(+)-KP is defined as HvKP carrying blaKPC.The factors responsible for HvKP include hypercapsule . Chi-square test was used to analyze comparisons between groups; c-rmpA/A2) to 99.6% (entB) among the 521\u00a0K. pneumoniae strains. Four genes  exhibited prevalence rates of > 90.0%, 1 (irp2) > 50.0% and the others < 25.0%. For the rmpAs, the order was: p-rmpA2 (12.5%), p-rmpA (10.6%) and c-rmpA/A2 (2.9%). For the four siderophore genes, the order was: entB (99.6%), irp2 (53.4%), iucA (15.7%) and iroN (9.2%). Positive rates of iroN and iucA were both lower than that of irp2 and entB . K. pneumoniae: each \u22652 strains. Totally 207 strains presented positive fimH, mrkD, entB and wzi and 190 showed positive fimH, mrkD, entB, irp2, and wzi simultaneously, which were the two primary modes and accounted for 39.7% and 36.5% respectively.wzy-K1, p-rmpA, p-rmpA2 or c-rmpA/A2, 49 (53.8%) possessed p-rmpA and p-rmpA2, 18 (19.8%) possessing wzy-K1, p-rmpA and p-rmpA2, 15 (16.5%) possessing merely p-rmpA2. wzy-K1/p-rmpA or p-rmpA/p-rmpA2. In the 520 strains positive in entB, irp2, iroN or iucA, 278 (53.5%) harbored entB and irp2, 241 (46.3%) harboring only entB, 35 (6.7%) harboring all the four genes. iucA/iroN and irp2. Other relationships were also shown in: peg-344, allS and ST23), p-rmpA, p-rmpA2 and c-rmpA/A2), peg-344, allS and ST14), irp2, iroN and iucA) and irp2, iroN and iucA). Gene wzy-K1 was completely restricted to K1 serotype (31/31), vice versa. High prevalence of peg-344 and allS was found in K1 strains , but rarely in K2 ones . Gene allS was mainly found in K1 strains (28/33), contrary to peg-344 (22/65). K1 strains mostly belonged to ST23 (23/31) while less than a half (17/38) of K2 ones belonged to ST14. K1 strains showed higher rates of rmpAs (p-rmpA/p-rmpA2/c-rmpA/A2) and siderophore genes (iroN/iucA) than K2 ones: 23/31 vs 10/38 (p < 0.0001), 23/31 vs 9/38 (p < 0.0001), which \u201cconfirmed\u201d hypervirulence in K1 strains.Among the 91 strains harboring K. pneumoniae (HvKP), blaKPC(+)-KP and Hv-blaKPC(+)-KP, as shown in blaKPC(+)-KP shared 17.6% (29/165) among blaKPC(+)-KP. For the blaKPC(+)-KP, ST11 accounted for 34.5% (57/165) while clonal group 258, including ST11, ST258, ST340 and ST437, was positive for 65.5% (108/165), indicating the focus of blaKPC(+)-KP.According to the aforementioned criteria, 94 (18.0%), 165 (31.7%) and 29 (5.6%) strains were denoted as hypervirulent blaKPC(+)-KP strains, ranging from fimH (100.0%), mrkD (100.0%), entB (100.0%), wzi (100.0%) to c-rmpA/A2 (6.9%). Genes iucA, p-rmpA2 and p-rmpA were positive in 28 (96.6%), 26 (89.7%) and 18 (62.1%) Hv-blaKPC(+)-KP strains respectively. A sum of 28 (96.6%) strains presented \u2265 3 siderophores and 29 (100.0%) carried p-rmpA/p-rmpA2 (p\u00a0> 0.9999).blaKPC(+)-KP strains, as shown in A total of nine modes of virulence genes were found among the 29 Hv-blaKPC(+)-KP strains, ST11 accounted for the majority  although more than 10 STs were found in total (Among the 29 Hv-blaKPC(+)-KP infections revealed that the prevalence of Hv-blaKPC(+)-KP significantly increased between 2018 and 2020, mainly from China, especially Mainland China , suggesting that Hv-blaKPC(+)-KP strains were mainly induced by two different plasmids (blaKPC(+)-KP strains. ST11 accounted for 17 (58.6%) among the 29 Hv-blaKPC(+)-KP strains. Those Hv-blaKPC(+)-KP strains with ST11 typically corresponded to K47 (9/17) and K64 (8/17) serotypes and were divided into four super subgroups. Those with ST86 were all K2 serotype (4/4).Trends in virulence among Hv-K. pneumoniae, in particular Hv-blaKPC(+)-KP.This study investigated the general distributions of key virulence genes in peg-344, of which 63 were denoted as HvKP. A sensitivity of 96.9% was therefore yielded, similar as the report (p = 0.5791) , corresponding to K1 serotype, vice versa, could help K. pneumoniae yield macromolecular EPS, which confers hypervirulence  . Gene aldocument . The reairulence . Wzi is capsular . Acapsulrobactin . Intrigurobactin .p-rmpA, p-rmpA2 or c-rmpA/A2 genes. Hypercapsule played an equal role with excessive siderophores  in hypervirulence of K. pneumoniae. The reason lies in the same pLVPK-like plasmids harboring rmpAs and siderophore genes concurrently.Except for macromolecular EPS and excessive siderophores, hypercapsule could also contribute to hypervirulence , which iblaKPC was first reported from USA in 1996 (blaKPC-2(+)-KP strain was reported in mainland China in 2007  strains while blaKPC-3 was found in 30 (5.8%) strains. Our study also showed clonal group 258 but not ST11 made up the majority of blaKPC(+)-KP -KP ; The reablaKPC(+)-KP worldwide. Different prevalence of iucA, p-rmpA2 and p-rmpA in Hv-blaKPC(+)-KP strains suggested their different roles in hypervirulence. The modes of virulence genes were rather diverse in Hv-blaKPC(+)-KP. Similar prevalence of \u2265 3 siderophores and p-rmpA/p-rmpA2 (p > 0.9999) indicated their equal roles in hypervirulence of Hv-blaKPC(+)-KP strains, which also originated from the same pLVPK-like plasmids harboring rmpAs and siderophore genes simultaneously. The proportion of K64 was , lower than another report (p < 0.0001). Further, IncHI1B plasmids carrying virulence genes and IncFII ones with blaKPC were responsible for both 23 strains, suggesting IncHI1B and IncFII plasmids jointly constitute the most successful combination. Furthermore, the phylogenetic trees revealed that the 29 Hv-blaKPC(+)-KP strains belonged to four super clusters although three clusters all possessed ST11 strains.The first Hv-CRKP, belonging to K2 and ST65, was unveiled in mainland China in 2015, which was isolated from blood in Wuhan City in March 2013 . Armed wr report  (p < 0.0blaKPC(+)-KP evolution may occur through two mechanisms. The first pathway is via HvKP acquiring a plasmid carrying drug-resistance determinants (via multidrug-resistant/extreme drug-resistant cKP acquiring a pK2044- or pLVPK-like virulence plasmid or integrated virulence genes into drug-resistance plasmids (blaKPC(+)-KP mainly evolved through the second pathway, i.e. via blaKPC(+)-KP acquiring another plasmid harboring virulence genes. blaKPC entering ST11 strains; IncHI1B plasmids are different from IncFII ones: rare protospacers were found and they lacked Type IV secretion systems, e.g. traM gene. Therefore, the mechanisms behind IncHI1B plasmids entering ST11 strains would be sophisticated and intriguing.Hv-rminants  or by thrminants . The secplasmids . Our datK. pneumoniae strains are not well known. Second, some\u00a0positive virulence genes do not inevitably mean \u201cexact\u201d hypervirulence.This study has some limitations. First, the specimen types of\u00a0521\u00a0K. pneumoniae. Hypercapsule plays an equal proportion with excessive siderophores in hypervirulence of K. pneumoniae. Virulence genes iucA, p-rmpA2 and p-rmpA are primary ones inducing Hv-blaKPC(+)-KP. IncHI1B plasmids carrying virulence genes and IncFII ones with blaKPC constitute the primary combination responsible for Hv-blaKPC(+)-KP. Hv-blaKPC(+)-KP urges more insightful investigations.Taken together, positive rates of virulence genes vary overwhelmingly in https://pan.baidu.com/s/1sbsl_phsx8IRoQeeY87e-w (password: xf5l).Publicly available datasets were analyzed in this study. This data can be found here: DH, YL and PR conceived the study. DT, WC, PF, WW and XJ collected the 521 genomes. DH, YL, PR and XL did bioinformation analysis. DH and YL wrote the manuscript, which was revised by XL and XJ. All authors contributed to the article and approved the submitted version.This work was funded by research grants from the National Natural Science Foundation of China  and the Shanghai Municipal Science and Technology Commission (grant number 19JC1413002).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Previous studies have demonstrated that obesity is associated with pulmonary fibrosis. We attempted to identify whether regular aerobic exercise (AE) can protect against high-fat diet (HFD)-associated pulmonary fibrosis.Forty-eight C57BL/6 mice were randomly assigned to four groups: chow group (Ch), chow plus exercise group (CE), obesity group (Ob), and obesity plus exercise group (OE). The mice were fed either an HFD or a chow diet for 16 weeks, and low-intensity aerobic exercise (AE) was performed in the last 8 weeks. We measured the degree of pulmonary fibrosis; pulmonary inflammation; oxidative stress parameters; insulin resistance-related indicators; the number of inflammatory cells in bronchoalveolar lavage fluid (BALF); the mRNA expression levels of IL-10, IL-1\u03b2, TGF-\u03b2, TNF-\u03b1, CXCL-1, IL-17, MMP-9, MPO, NE, and sirt-1; and the BALF levels of CXCL-1, IL-17, TGF-\u03b2, IL-10, IL-1\u03b2, and TNF-\u03b1 in lung tissue.AE in obese mice protected against obesity-associated pulmonary fibrosis, chronic inflammation, pro-oxidative/antioxidative imbalance, and insulin resistance. AE ameliorated the HFD-induced inflammatory response and neutrophil infiltration in the lung. AE downregulated BALF levels of CXCL-1, IL-1\u03b2, TNF-\u03b1 IL-17, and TGF-\u03b2 but upregulated BALF levels of IL-10. AE decreased IL-1\u03b2, TGF-\u03b2, TNF-\u03b1, CXCL-1, IL-17, MMP-9, MPO, and NE mRNA expression levels but upregulated IL-10 and sirt-1 mRNA expression levels in the lung.AE protects against HFD-induced pulmonary fibrosis by improving obesity-associated insulin resistance, chronic low-grade inflammation, and pro-oxidative/antioxidative imbalance. AE improved HFD-induced pulmonary fibrosis by suppressing IL-17, TGF-\u03b2, NE, and MMP-9 expression and activating IL-10 and sirt-1 expression. The prevalence of obesity is increasing drastically and its prevalence markedly upregulates the incidence of many complications such as chronic obstructive pulmonary disease (COPD), asthma, and pulmonary fibrosis . At presvia the TGF-\u03b2 pathway .All protocols used in this study were approved the Animal Experimental Welfare of the Institute of Animal Science, Chinese Academy of Agricultural Sciences . All experiments were performed in accordance with the Animal Experimental Welfare of the Institute of Animal Science, Chinese Academy of Agricultural Sciences and the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health. The mice were anaesthetized Forty-eight male C57BL/6 mice were randomly divided into four groups: (1) chow (Ch) group, where the mice were fed a chow diet for 16 weeks; (2) chow diet plus exercise (CE) group, where the mice were fed a chow diet for 16 weeks and forced to train in the last 8 weeks; (3) the obesity (Ob) group, where the mice were fed a HFD for 16 weeks; and (4) the obesity plus exercise (OE) group, where the mice were fed a HFD for 16 weeks and forced to train in the last 8 weeks. Each group included 12 mice.\u00b0 incline, 18 m/min).OE mice were forced to train using a treadmill. Treadmill aerobic training lasted for 8 weeks and was performed once a day for 60 min per session  Biotech, CO., LTD, Hangzhou, China).Following the manufacturer\u2019s specifications, superoxide dismutase (SOD), malondialdehyde (MDA), myeloperoxidase (MPO), and glutathione (GSH) levels in the mice were detected using spectrophotometry as previously described  . We measTotal RNA was extracted from DNase I-treated cells using TRIzol as previously described . Total RAccording to the reagent manufacturer\u2019s specifications, plasma glucose levels were detected with a glucometer . The plasma insulin and adiponectin levels were detected using commercial kits .post hoc test was utilised to analyse the data in this study. The significance threshold was set at P < 0.05. GraphPad Prism 9 software was utilised to analyse the data and draw figures.All data were expressed as the mean \u00b1 SEM. Two-way ANOVA and Tukey\u2019s P < 0.001), net body weight gains (P < 0.001), abdominal fat weights (P < 0.001), and subcutaneous fat weights (P < 0.001) than did Ch mice. OE mice had lower body weights (P < 0.001), net body weight gains (P < 0.001), abdominal fat weights (P < 0.001), and subcutaneous fat weights (P < 0.001) than did Ob mice.As In the Ch group and the CE group, no inflammatory cell infiltration was observed in the lung. HFD markedly increased the number of neutrophils in the lung, while AE markedly decreased the number of neutrophils in obese mice .P < 0.001) in BALF was markedly upregulated. Regular AE markedly downregulated the number of neutrophils in obese mice (P = 0.0301).As shown in P < 0.001) and CXCL-1 (P < 0.001), profibrogenic factors TGF-\u03b2 (P < 0.001) and IL-17 (P < 0.001), and proinflammatory factor IL-1\u03b2 (P < 0.001) but significantly lower levels of the anti-inflammatory factor IL-10 (P < 0.001) in BALF than did Ch mice. AE markedly decreased BALF levels of IL-1\u03b2 (P = 0.033), IL-17 (P = 0.009), TGF-\u03b2 (P = 0.013), and TNF-\u03b1 (P = 0.025) but markedly increased BALF levels of IL-10 (P = 0.004) compared with Ob mice.As shown in Masson staining showed the degree of pulmonary fibrosis. Compared with those in the Ch group and the CE group, the magnitude of pulmonary fibrosis increased after 16 weeks of HFD administration, but AE decreased the magnitude of pulmonary fibrosis in obese mice .Sirius Red staining showed collagen fibre deposition in lung tissue. In the Ch group and the CE group, no obvious collagen fibre deposition was noted in the airway wall, while collagen fibre deposition in the airway wall increased after 16 weeks of HFD administration. AE decreased collagen fibre deposition in the airway wall in obese mice .P = 0.024) and IL-17 (P = 0.0031) in BALF. Regular AE downregulated BALF levels of TGF-\u03b2 (P = 0.013) and IL-17 (P = 0.003) in obese mice.As shown in P < 0.001), Ashcroft fibrosis (P < 0.001), lung fibrotic score (P < 0.001), and airway collagen (P < 0.001) in the lung, whereas AE markedly downregulated hydroxyproline levels (P = 0.019), Ashcroft fibrosis (P = 0.024), lung fibrotic score (P = 0.031), and airway collagen (P = 0.005) in obese mice.As P < 0.001), IL-1\u03b2 (P < 0.001), IL-17 (P = 0.0034), MMP-9 (P = 0.0089), MPO (P < 0.001), NE (P < 0.001), TGF-\u03b2 (P < 0.001), and TNF-\u03b1 (P = 0.0072) mRNA expression levels but decreased IL-10 (P = 0.013) and sirt-1 (P = 0.011) mRNA expression levels compared with the Ch group. AE for 8 weeks markedly downregulated CXCL-1 (P < 0.001), IL-1\u03b2 (P < 0.001), IL-17 (P = 0.024), MMP-9 (P = 0.0029), MPO (P = 0.013), NE (P = 0.011), TGF-\u03b2 (P = 0.0125), and TNF-\u03b1 (P < 0.001) mRNA expression levels in obese mice.The mRNA expression levels of CXCL-1 , IL-1\u03b2 , IL-10 , IL-17 , MMP-9 , MPO Fi, NE Fig, sirt-1 P = 0.0063) and SOD (P = 0.0047) and increased expression levels of MDA (P = 0.0092) and MPO (P = 0.0025) in lung tissue compared with the Ch group. Significant downregulation of MPO (P = 0.0396) and MDA (P = 0.0412) expression and significant upregulation of GSH (P = 0.0037) and SOD (P = 0.0268) expression were detected after 8 weeks of AE compared with the Ob group.The levels of GSH , MDA Fi, MPO Fi, and SODP < 0.001) and plasma insulin levels (P < 0.001) but markedly upregulated blood glucose levels (P < 0.001) compared with Ch treatment. Significant downregulation of blood glucose levels (P = 0.0164) and significant upregulation of plasma adiponectin levels (P = 0.0386) and plasma insulin levels (P = 0.0441) were detected after AE for 8 weeks. The results of the GTT (HFD feeding markedly downregulated plasma adiponectin levels ( the GTT , ITT (Fi the GTT , and IRT the GTT  demonstrPrevious studies demonstrate that obesity was closely associated with pulmonary fibrosis . HoweverMorphological analysis demonstrated that AE in obese mice reduced hydroxyproline levels and collagen fibre deposition in lung tissue. Our data demonstrated that HFD administration dramatically increased the levels of the profibrogenic factors NE, MMP-9, TGF-\u03b2, and IL-17 in BALF. Similarly, HFD administration dramatically increased NE, MMP-9, TGF-\u03b2, and IL-17 mRNA expression levels in lung tissue, which was reversed by the end of the 8-week AE programme. Thus, AE protected against pulmonary fibrosis by repressing the expression of NE, MMP-9, TGF-\u03b2, and IL-17. In addition, sirt-1 was a negative regulator of MMP-9 . TherefoNeutrophils play an important role in the pathogenesis of pulmonary fibrosis . We founHFD administration led to pro-oxidative/antioxidative imbalance, and the antioxidant effects of exercise were identified in this study. Our data identified that regular AE in obese mice significantly downregulated MDA and MPO levels but significantly upregulated SOD and GSH levels in the lung. AE in obese mice modulated the oxidative/antioxidative balance in lung tissue. Previous studies have demonstrated that sirt-1 has therapeutic effects in lung diseases because sirt-1 modulates the pro-oxidative/antioxidative balance . Our datPrevious studies identified that obesity-associated insulin resistance enhanced TGF-\u03b2 expression, which played an important role in the pathogenesis of pulmonary fibrosis . We founThe results of this study provide insight into pulmonary fibrosis and indicate that AE is a novel tool to treat pulmonary fibrosis. HFD administration leads to pulmonary fibrosis by causing insulin resistance, a chronic inflammatory response, pro-oxidative/antioxidative imbalance, and increased levels of profibrogenic factors, including TGF-\u03b2, IL-17, NE, and MMP-9. AE improves HFD-induced pulmonary fibrosis by counteracting obesity-associated insulin resistance, chronic inflammatory responses, and pro-oxidative/antioxidative imbalance and shows an antifibrogenic effect by suppressing TGF-\u03b2, IL-17, NE, and MMP-9 production in obese mice.The original contributions presented in the study are included in the article/The animal study was reviewed and approved by Animal Experimental Welfare and Ethical Inspection Form of Institute of Animal Science, Chinese Academy of Agricultural Sciences.XW: conceptualisation, methodology, software, validation, writing\u2014original draft preparation, and project administration. XY and DT: formal analysis, investigation, resources, supervision, and funding acquisition. XW, XY, and DT: writing\u2014review and editing. All authors have read and agreed to the published version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "The ts3agn mutation drastically increases the frequency ofectopic contacts (FEC) in specific regions of intercalary heterochromatin, suppresses learning/memory and affectslocomotion. As is shown in this study, the polytene X chromosome bands in reciprocal hybrids between ts3agn and thewild type strain Berlin are heterogeneous in modes of FEC regulation depending either on maternal or paternal geneorigin. Bioinformatic analysis reveals that FEC between X:11AB and the other X chromosome bands correlates with theoccurrence of short (~30 bp) identical DNA fragments partly homologous to Drosophila 372-bp satellite DNA repeat.Although learning acquisition in a conditioned courtship suppression paradigm is similar in hybrids, the middle-termmemory formation shows patroclinic inheritance. Seemingly, this depends on changes in miR-974 expression. Severalparameters of locomotion demonstrate heterosis. Our data indicate that the ts3agn locus is capable of trans-regulatinggene activity via POEs on the chromatin nuclear organization, thereby affecting behavior.Prognosis of neuropsychiatric disorders in progeny requires consideration of individual (1) parent-of-origineffects (POEs) relying on (2) the nerve cell nuclear 3D chromatin architecture and (3) impact of parent-specific miRNAs.Additionally, the shaping of cognitive phenotypes in parents depends on both learning acquisition and forgetting,or memory erasure. These processes are independent and controlled by different signal cascades: the first is cAMPdependent,the second relies on actin remodeling by small GTPase Rac1 \u2013 LIMK1 (LIM-kinase 1). Simple experimentalmodel systems such as Drosophila help probe the causes and consequences leading to human neurocognitive pathologies.Recently, we have developed a Drosophila model for Williams\u2013Beuren Syndrome (WBS): a mutant  Genome plasticity is ensured by the architecture of specificnuclear loci and nuclear localization of transcriptional machinerywhere the chromatin organization is the priority-driven factor. The outcome of recent achievements in systems biologyis the notion that the plasticity of 3D chromatin architectureof nervous cell nuclei plays the leading role in cognitionand neuropsychiatric disorders . The epigeneticcomponent is still an underestimated source of psychomotordisturbances and neuronal diversity . Therefore, a new field of human biomedical researchnamed molecular cytogenetics and cytogenomics , or chromosomics  has evolved. Themain goal of chromosomics is the study of chromosomes,their 3D architecture in the interphase nucleus, the outcomesof chromosomal sub-region plasticity and gene interactions forshaping interindividual and intercellular genomic variationsin normal behavior and disease.Recently, this topical problem has turned out to be thepursuit of understanding the concomitant role of activeforgetting, since the antithesis to learning acquisition is theforgetting or memory erasure . Bothprocesses are independent and controlled by different signalcascades: learning acquisition and memory consolidationoccurs via cAMP cascade, its components being CREB andC/EBP. Active forgetting relies on actin remodeling cascaderesponsible for structural alterations of neurons and synapses:small GTPase Rac1 \u2013 LIMK1 (the key enzyme of actin remodelingLIM-kinase 1) and its phosphorylation substrate\u0441ofilin. The absence of Rac1-dependent forgetting causesthe autistic spectrum disorders. Expression changes (activeor non-active state) of LIMK1 and cofilin lead to differentneuro pathologies. The most studied example embracing allthe aforementioned facets of manifestations is Microdeletion(Deletion) Williams\u2013Beuren Syndrome, or WBS in 7q11.23.WBS deletion leads to cardiovascular pathology, cognitivedeficit in visuospatial construction and hypersociability. This isbecause long-term synaptic plasticity and depression determinesuccessfulness of learning and memory, as well as oflocomotor behavior, and depend on epigenetic regulation of LIM-kinase 1 (LIMK1) gene, one of approximately 28 genesuncovered by WBS deletion. Epigenetic regulation of LIMK1activity involves DNA methylation, chromatin remodeling andthe noncoding RNA-mediated process .LIMK1, a member of serine/threonine (Ser/Thr) familykinases regulated by the Rho-GTPase pathway, is the keyenzyme of actin remodeling cascade. Dendritic spines areactin-rich structures, and spine dynamics is driven mainly byactin remodeling, thus sharing several molecular pathwayswith dendrite growth . Increasing sets ofevidence suggest that nuclear actin also plays a pivotal rolein transcriptional regulation and DNA repair. Interestingly,monomeric actin is a stoichiometric subunit of a variety ofchromatin remodeling complexes. Ashift between monomericand polymeric states modifies activity of histone deacetylases.agnostic locus harboringLIMK1 gene (X:11AB). This region possesses theproperties of intercalary heterochromatin. Being a hotspot ofchromosome breaks, ectopic contacts, underreplication andrecombination, the region attains strain-specific architecturemarked by single base changes and small insertion/deletions.The EMS-induced temperature-sensitive (ts) mutationts3agn carries the insertion of transposable element (TE) fromTc1/mariner superfamily ~460 bp downstream 3\u2032UTR ofDrosophila LIMK1 gene (dlimk1), as well as A/T-rich 28 bpinsertion within intron 1 of dlimk1 capable of pairing with5\u2032 TIR of the TE.Recently, we have developed a simple and appropriateDrosophila model for chromosomics  using the properties of the ts3agn shows a temperaturesensitivelethality at all stages of development except for theimaginal stage. At normal temperature, the adult flies showdrastic learning acquisition and memory retention defects, aswell as locomotor impairments and amyloid-like inclusions. Stressexposure (heat shock for 30 min at 37 \u00b0C) suppresses thesemanifestations . Also, ts3agnmutationleads to: (1) LIMK1 and p-cofilin increase in the adultbrain and salivary glands of 3rd instar larvae at 22\u201325 \u00b0C anda fall down to the level of the wild type strain Canton S at29\u201337 \u00b0C; (2) high level of ts-induced recombination withints3agn region; (3) 3-fold increase in frequency of non-allelicectopic contacts (FEC) within 2L arm of the chromosome 2 and in the 11\u0412 X chromosome region . Additionally, miRNAs expression including the biomarkersfor human neuropathologies is drastically reducedin ts3agn relative to the wild type strains .When maintained at 29 \u00b0C, ts3agn-specific nuclear organization is shaped in early embryogenesisalongside with formation of chromosomal heterochromatinregions. Intrinsic ts3agn FEC is maternally inherited. Therefore, ts3agn is a promisingmodel for studies on parent-of-origin effects (POEs) on progenyconsidered as significant causative factors of psychiatricdisorders . For instance, a 1.5 Mb WBSdeletion recurrently arises de novo and depends on POEs: maternalorigin leads to more severe developmental abnormalitiesand microcephaly . Moreover,when WBS deletion has a paternal origin, expression levels ofa number of genes within the WBS deletion decrease. Amongthese genes crucial for the brain development is a gene for generaltranscription factor II-I (GTF2I). It regulates transcriptionby binding to DNA and histone deacetylase (HDAC) . The main goal of the study is the analysisof POEs role in shaping quantitative traits, namely learningacquisition, memory retention, locomotion, and miRNAs expressionwhile using the advantages of the Drosophila modelfor POEs in progeny from reciprocal crosses between ts3agnand the wild type strain Berlin .To meet the requirements of chromosomics, FECs betweenthe region X:11AB and the other bands of the X chromosomemay be estimated as an indicator of chromosomal spatialorganization. This approach is justified by the existence oflate-replicating genomic territories including the underreplicatedregions of polytene chromosomes. These regions overlapwith late-replicating regions of mitotically dividing cells. Suppression of SUUR gene responsiblefor underreplication of intercalary heterochromatin and, as aconsequence, for the ectopic pairing leads to death in early embryogenesis. Additionally, to elucidatethe contribution of DNA sequence homology as componentsof epigenetic regulation in the ectopic chromatin pairing wehave developed the special software package Homology SegmentAnalysis.Drosophila stocks. The fly stocks used belong to Biocollectionof Pavlov Institute of Physiology of the Russian Academyof Sciences:\u2022 Berlin, a wild type strain;\u2022 Canton S, a wild type strain;\u2022 ts3agn, a temperature-sensitive mutation on Canton Sgenetic background within agnostic locus (X:11AB) affectingdlimk1 activity.ts3agn and Berlin were usedbecause Berlindlimk1 sequence is closer to FlyBase referencesequence . At the same time,the reciprocal hybrids ts3agn\u00d7Canton S (the genetic backgroundfor ts3agn) and Canton S\u00d7ts3agn demonstrate exactlythe same cognitive behavior as reciprocal hybrids with Berlin approving the usage of Berlin.The reciprocal hybrids between Berlin and ts3agn female and male parents and F1 female and male progeny from reciprocal crosses with theaccent on the putative hairpin formed in the X chromosomeby 28 bp A/T rich insertion within intron 1 of dlimk1 geneand 3\u2032 end of Tc1/mariner element.Figure 1 shows the X chromosome and autosomes architecturein Flies were maintained on standard Drosophila yeast-raisinmedium at +22 \u00b1 0.5 \u00b0\u0421 under a 12-h light/dark cycle. Forthe memory and locomotion tests, males were collected uponeclosion without narcotization and kept individually in culturevials till the behavioral experiments on the 5th dayEstimation of frequency of ectopic contacts (FECs).The aceto-orcein squash preparations were prepared fromsalivary glands of III instar D. melanogaster female larvae.20 to 30 animals were examined, therefore the number ofanalyzed chromosomes varied from 300 to 500. The examplesof ectopic contacts are presented on Fig. 2. The number ofnon-homologous contacts between the region X:11AB anddifferent bands of the X chromosome was calculated and expressedas per cent of the total number of the examined nuclei.FECs in parents and F1 reciprocal hybrids were comparedusing Student\u2019s t-test. Identification of genes localized in theX chromosome bands forming contacts with the 11AB regionwas performed using NCBI Genome Data Viewer databasehttps://www.ncbi.nlm.nih.gov/genome/gdv/ and molecularfunction of identified genes was derived from FlyBase https://flybase.org analysis of DNA segments homology.D. melanogaster genome sequence (release 6) was taken from. Special software package HomologySegment Analysis searching the matches of short singlestrandedDNA fragments within the chromosome areasinvolved in the ectopic pairing in Drosophila has been developed.The software written in Python 3 can be freely downloadedfrom . Software version from git(commit 41719cddc6283edbd79c5bf2aee237cde48d4b7d)was used. The algorithm of the program is described in brief in. The exact run parameters for 11AB region are following: segmentanalysis.py -v -s 30 dm6.nounmapped.fa.gz :X:11982050:12772070 DmelMapTable.160615c.bedHere, dm6.nounmapped.fa.gz is Drosophila genome canonicalsequence (Ensembl release 6) and DmelMapTable.160615c.bed is chromosome bands location from Ensembldatabase converted to BED format. Both files are included insoftware repository.For other tested regions, all parameters were the sameexcept for the regions location.Preparation of miRNAs libraries and bioinformaticanalysis. The detailed description of the procedure is givenin . Extract RNA reagent was used for total RNA extraction fromadult 5 days old males. To obtain the fraction of small RNA,25 \u03bcg of total RNA were separated using 15 % polyacrylamidegel electrophoresis in the presence of Urea (8 M) followingexcision of small RNA fraction corresponding to 21\u201329 nts.Illumina TruSeq Small RNA prep kit  wasused for small RNA libraries preparation. Sequencing wasperformed on an Illumina HiSeq 2000 platform.The amount of mapped miRNAs reads was counted byBEDTools (v. 2.22) and mirbase annotation (r. 19) . Analysis of differentially expressed miRNAs wasperformed using edgeR (v. 3.10.2) package in R environment(v. 3.2.2) . miRNA functions werederived from miRBase . Small RNAlibraries were deposited in NCBI SRA under the numberPRJNA633483.Locomotor activity. Computer-aided automatic device forsimultaneous registration of 20 animals is described in . The experiment lasted for 1 h. Spontaneouslocomotor activity of flies was detected in a plate witheight chambers and transparent cover using high-resolutionvideo camera. Software used for locomotor activity analysisis freely available at .The following parameters of locomotion were assessed:activity index (%); run frequency (the number of run boutsin 100 seconds); running speed (mm/s). The full record wasdivided into 1 s quanta, and the mean speed of fly movementin each quantum was calculated. If the result was less thanthe threshold value (5 mm/s), the fly was considered to beresting during this time quantum; otherwise, it was considered moving. Neighboring quanta with similar movement patternwere merged in intervals of moving and resting. Activity indexis determined as a time spent in movement. Running speedis an average fly speed, determined using only intervals ofmovement. The Kruskal\u2013Wallis analysis of variance with themultiple comparison of mean ranks was used to compare allthe experimental groups.Learning acquisition and middle-term memory formationin Drosophila males. Detailed description of learning/memory assessments in conditioned courtship suppressionparadigm (CCSP) and specially designed software for observationand statistical analysis (randomization test) of learningindices based on courtship indices is given in . CCSP employs the natural stimuli of Drosophilacourtship. Both virgin and fertilized females emit an aphrodisiacpheromone, attracting a na\u00efve male without courtshipexperience. However, a fertilized female rejects a male atthe courtship stage of attempted copulation via emitting anaversive pheromone. Repetitive rejections during 30 mintraining provoke a kind of learned helplessness when a malestops courting another female. This courtship suppressionmight last for one hour when test female is virgin and foreight hours when fertilized. Males with defective memoryformation continue to court after such a training as vigorously,as na\u00efve males.The courtship index was calculated for each male. The learning index (LI)was computed according to the formula:Analysis of spatial nuclear organizationdelimited to FECs formedby the X chromosome region 11ABBerlin, ts3agn and their hybrids are presented in Table 1. Thecolumns 1\u20134 show the pattern of FECs between the 11AB regionand other X chromosome regions pertinent to listedstrains. The absence of significant differences of FECs betweencolumns, i.e. 1 vs 2 and 3 vs 4 indicates matroclinicinheritance, 2 vs 3 indicates hybrid-specific frequencies, 2 vs 4and 1 vs 3 pinpoints the patroclinic inheritance. The polytenechromosome bands in hybrids demonstrate differences inFECs, a part of them showing either matroclinic propertiesor properties of the father strain. In certain bands, FECs aresimilar in reciprocal hybrids or depend on the direction of a cross, but significantly differfrom that of parents.Spatial nuclear organization was analyzed using microscopicimages of polytene chromosomes in larvae salivaryglands. The results of comparative analysis of FECs betweenthe 11AB region and the other X chromosome regions in Berlin, ts3agn and their hybrids are presented in Table 1. Thecolumns 1\u20134 show the pattern of FECs between the 11AB regionand other X chromosome regions pertinent to listedstrains. The absence of significant differences of FECs betweencolumns, i.e. 1 vs 2 and 3 vs 4 indicates matroclinicinheritance, 2 vs 3 indicates hybrid-specific frequencies, 2 vs 4and 1 vs 3 pinpoints the patroclinic inheritance. The polytenechromosome bands in hybrids demonstrate differences inFECs, a part of them showing either matroclinic propertiesor properties of the father strain. In certain bands, FECs aresimilar in reciprocal hybrids or depend on the direction of a cross, but significantly differfrom that of parents.NCBI Genome Data Viewer software helped to reveal thegenes located in the X chromosome bands forming the ectopiccontacts. Based on the assumption that shared location ofgenes determines their functional features ,it was worth elucidating what biological processes are underthe influence of epigenetic factors related to allelic parentof-origin. Therefore, the grouping of genes implied theirinvolvement in control of a certain biologic process .ts3agn\u00d7Berlin relative to the reciprocalhybrid are genes for motor proteins  and sensoryreceptors . Genes involved in neurodevelopment,oxidative-reduction process and proliferation do not present inBerlin\u00d7ts3agn progeny. Heterosis, when the number of genesinvolved in hybrid-specific contacts is more than 2-fold higherthan in both parents, is manifested for oxidative-reductionprocess (3), signal transduction (4), proliferation (9) and noncodingRNAs (13).Figure 3, a presents provisional definition of the biologicalprocesses controlled by genes within bands forming contactswith 11AB region either in matroclinic or in reciprocal hybridspecificmanner. Among the functional groups of genes havingincreased number in ts3agn\u00d7Berlin relative to thereciprocal hybrid), reactive oxygen species metabolic process, and metabolism . Sensory perception andsignal transduction groups do not present in Berlin\u00d7ts3agn.The biological processes and genes involved in ectopiccontacts with 11AB in a mode of father strain are shown inFig. 3, b. Among them are the genes responsible for chromatinremodeling ; metabolism ; cell proliferation ); chromatine remodeling . Reparation/recombination group does not present in Berlin\u00d7ts3agn.Data on biological processes and the number of genes involvedin contacts with 11AB region exclusively in hybridsor with FECs prevailing those of parents (FEC heterosis), areshown on Fig. 3, c. The ratio of gene numbers for Profiles of the localized fragment frequencies (LFFs)Using our specially designed software Homology SegmentAnalysis, we analyzed correlation between ectopic pairingand distribution of small identical fragments within contactingregions.The region X:11AB (~790 kb) involved in DrosophilaX \u2013 X:11AB ectopic pairing was selected as the source ofsmall 30 nt fragments, and the profile of X:11AB localizedfragment frequencies (LFFs) was constructed for the X chromosome. The region X:14B\u201315B was selected asa control region having almost equal length (~790 kb).ts3agn\u00d7Berlin FEC (X:11AB). Probably,20 nt fragments are too short to display DNA homologyspecifically associated with ectopic pairing. The increasein fragments length from 20 to 50 nt leads to decrease inLFF (X:14B\u201315B) \u2013 FEC (X \u2013 X:11AB) unspecific correlation and simultaneous increase in LFF \u2013 FEC (X:11AB) specificcorrelation. However, there is a lack of LFF (50 nt) \u2013 ECFcorrelation for ts3agn. Hence, 30 nt fragment length seems tobe optimal in search for the identical fragments within thecandidate chromosomal regions involved in ectopic pairing.Both LFF and the FEC (X \u2013 X:11AB) distributions significantlydiffer from normal. Therefore, we calculated Spearman\u2019srank-order correlation coefficients (rS) for LFF andthe strain-specific FEC (X \u2013 X:11AB) (Table 2). A positivecorrelation between LFF and FEC is observed within the region of interest X:11AB. The influence of the fragment lengthon rS value has been tested. For 30 nt fragments, rS is highlysignificant for all Drosophila strains ( p < 0.001). As to LFF(X:14B\u201315B), there is no significant correlation with FEC(X \u2013 X:11AB) ( p \u2265 0.01) pointing to specificity of analysisof DNA homology within the region of ectopic pairing. For 20 nt fragments, there is a false-positive correlation betweenLFF (X:14B\u201315B) and Berlin FEC and thelocalization frequencies for the several specific X:11AB 30 ntfragments having the highest number of occurrences (NO)in the X chromosome (Table 3). The maximal correlation isevident in the part of 372 bp middle repetitive DNA sequence). The majorityof fragments with significant positive rS values appear to bethe parts of a ~50 bp repeat (marked with #) having significantself-complementarity. This sequence also shows an almostcomplete identity to another part of 372 bp repeat. Such repeatswith slight sequence variations occur in both DNA strands ofthe X chromosome. rS is much lower for (-at-)15 repeat and isnearly absent for (-gt-)15 (-tc-)15 and (-ca-)15 repeats, althoughtheir NO can be higher. Noteworthy, all rS values for 372 bprepeat fragments are lower compared to rS for the whole setof the localized fragments (see Table 2), hence all of themseemingly impact ectopic pairing.To find out what DNA sequences impact the ectopic pairingthe most, we calculated rS between miRNAs expression profileBerlin, ts3agn and their reciprocalhybrids demonstrates significant differences betweenmales of parent strains and hybrids in content of 44 miRNAs.For the reciprocal hybrids, the heat maps of miRNAs expressionare presented in Fig. 5. Among 44 miRNAs 10 miRNAsbelong to the same cluster of testis-specific miRNAs . However, only miR-980 and miR-974are involved in memory processes. Expression of miR-980suppresses Drosophila memory ,while lowered expression of miR-974 impairs memory formationAnalysis of miRNAs expression in The heat map represents the RPM-normalized and log2-transformed counts of miRNAs reads with z-scale normalizationof the rows. Thirty percent of low-expressed miRNAswere removed from further analysis. Only the miRNAs withaltered content in a hybrid compared to at least one parentare shown. analysis of parent strainsand their reciprocal hybridsLocomotor behavior. Figure 6 presents the parameters oflocomotion. As to activity indices, they are significantly differentin ts3agn and Berlin, both hybrids differ from ts3agn andBerlin\u00d7ts3agn differs from Berlin. Running speed and runfrequency in ts3agn and Berlin are similar. Both hybrids differfrom ts3agn and Berlin\u00d7ts3agn from Berlin. However, thehybrid running speed exceeds that of parents, demonstratingheterosis. Comparatively to parents, run frequency in hybridsis intermediate. At the same time, any alterations in locomotorparameters in hybrids are similar and unidirectional.Learning acquisition and memory formation.In all strains,CIs of males decrease after training with fertilized femalescompared to na\u00efve flies . Learning/memory scoresin reciprocal hybrids show that memory formation (3 hoursafter training), but not learning acquisition (0 hours aftertraining), demonstrates patroclinic inheritance .Berlin strain in crossests3agn\u00d7Berlin, learning and 3-hour memory do not differfrom LIs of Berlin, but do differ from LIs of ts3agn. Patroclinicinheritance is evident in the reciprocal hybrid Berlin\u00d7ts3agn.In this case, 3-hour LIs in Berlin\u00d7ts3agn and ts3agn are similar.The patroclinic inheritance cannot be associated with theY chromosome, as it is likely to be similar in Berlin, ts3agnand Canton S . Also, it cannot be attributed to the X chromosomes,since they are different in Berlin and ts3agn\u00d7Berlin.Seemingly, this may be caused by some paternal epigeneticfactors, such as miRNAs in cytoplasm of male sperm.When male parent originates from Discussionts3agn gene and epigenetic factors (POEs) in spatial nucleararchitecture, learning/memory formation and spontaneouslocomotor activityStudies of genome-wide associations between DNA polymorphismsand phenotypic traits have revealed genetic variantspredisposing to different mental diseases. Findings pinpointingthe role of POEs in genetic risk for neuropathology opennew possibilities for therapy and preventive medicine . In this study, we estimated FECs in reciprocalhybrids considering an impact both of genetic variants ofdlimk1 gene.Our assumption was that FE\u0421s partly reflect the restrictedhomology of short DNA sequences in different, seemingly\u201cnon-homologous\u201d regions.To exploit the advantages given by the model, we delimitedthe analysis of spatial nuclear organization to FECs formedby the X chromosome region 11AB harboring As shown in this study for the short identic fragments withinthe contacting regions, FECs correlate with LFFs. Althoughthe correlation is rather moderate (~0.35), it is highly specificfor 30 nt fragments. Many factors affect ectopic pairing,such as DNA homology, the distance between the interactingregions and epigenetic factors causing the interstrainFECs differences . Our computationalalgorithm concerns only fragments aligned with the X chromosomewithout gaps. This reveals the partial homology ofinteracting bands. Thus, the interacting chromosomal areasmay be significantly larger than 30 or 50 nts. Although differentmechanisms are involved in ectopic pairing, includingPOEs, our data indicate the significant role of DNA sequenceitself. However, as FEC\u2013LFF correlation is mainly observedfor specific DNA fragments, their pairing mediated by someproteins or non-coding RNAs cannot be ruled out.Most of the found 30\u201350 nt fragments are similar to theD. melanogaster dispersed 372 bp A/T-rich noncoding repeat. This moderately repeated sequence islocated in the euchromatin of the X chromosome between theregions 4 and 14A in ~300\u2013400 copies per haploid genome.The 372 bp repeat is a part of 1.688 g/cm3 class of satelliteDNA (1.688X repeats) . siRNAfrom the 1.688X repeats is involved in dosage compensationin recognition of the X and autosomal chromatin, therebydelimiting activities of male-specific lethal (MSL) complexto sex chromosomes through up-regulation of the X chromosome.ts3agn, FECs in thesebands significantly decrease. In this case maternal control ofspatial localization and therefore, of gene expression is geneticallydetermined. These are genes controlling membranereceptor regulation (PPYR1) and signal transduction (gce),chromatin remodeling , axon guidance andchemosensory jumping behavior (acj6 ).The data obtained consider the common mechanisms ofectopic contacts formation and dosage compensations. Seemingly,POEs might influence the spatial chromatin organization,thereby affecting behavioral performances.This indicates that each Drosophila strain possesses itsown pattern of ectopic contacts with the region 11AB. Thepolytene chromosome bands are heterogeneous in their modesof regulation of ectopic pairing. Apart of them is regulated bygenes of either maternal, or paternal origin. A separate classis comprised of regions manifesting only hybrid properties.Similar \u0420\u041e\u0415s were observed for the pattern of methylation andnucleosome distribution within the imprinted loci in humansand plants .In both reciprocal crosses, bands 7A, 9A and 13B displaythe maternal properties. When mother is This indicates that each Drosophila strain possesses itsown pattern of ectopic contacts with the region 11AB. Thepolytene chromosome bands are heterogeneous in their modesof regulation of ectopic pairing. Apart of them is regulated bygenes of either maternal, or paternal origin. A separate classis comprised of regions manifesting only hybrid properties.Similar \u0420\u041e\u0415s were observed for the pattern of methylation andnucleosome distribution within the imprinted loci in humansand plants .ts3agn, FECs in thesebands significantly decrease. In this case maternal control ofspatial localization and therefore, of gene expression is geneticallydetermined. These are genes controlling membranereceptor regulation (PPYR1) and signal transduction (gce),chromatin remodeling , axon guidance andchemosensory jumping behavior (acj6 ).In both reciprocal crosses, bands 7A, 9A and 13B displaythe maternal properties. When mother is ts3agn\u00d7Berlin, the number of genes with knownfunctions contacting with 11AB with maternal-specific frequencyis 2-fold higher than in reciprocal cross. Possibly, thisis due to the ts3agn-specific miRNAs pattern of expression.The role of miRNAs in maternal inheritance and expressionin embryogenesis is sparsely studied. As we have shownearlier, the expression level of miR-9, miR-34 and miR-124differs in ts3agn from that in Berlin, Canton S and Oregon-R. miR-9 and miR-124 are alsoexpressed in early development (0\u201312 hrs) , miR-34 is detected in embryos till zygotic reduction. As known, the switch from maternal tozygotic development program occurs between the second andthe third hours of embryonic stage, hence miRNA found inearly development have maternal origin . ThesemiRNAs targets are Swi/Snf-like complex, neural-progenitorspecificnpBAF, repressor-element-1-silencing transcriptionfactor (REST belonging to 1 class of histone deacetylases(HDAC1/2) and silent information regulator 1 (SIRT1) \u20133 class of NAD+-dependent histone deacetylases involved inheterochromatin formation, Bourassa, Ratan, 2014). They areinvolved in neurogenesis, dendrite morphogenesis and axonguidance which depend on global chromatin remodeling.In the cross ts3agn\u00d7Berlinectopic contacts between 11AB and regions containing genesinvolved in chromosome remodeling are formed with frequencycharacteristic for the maternal genome. The products ofthese genes are: Tip60 \u2013 histone acetyltransferase, HDAC6 \u2013histone deacetylase; mxc \u2013 regulator of histone synthesis ofPolycomb group; Top1 \u2013 DNA topoisomerase. However, onlytwo regions containing genes HDAC6 and Top1 are presentin cross Berlin\u00d7ts3agn.Therefore, it is not surprising that in cross ts3agn-like matroclinicmode of inheritance is pertinent to genes responsible for actinand microtubules-binding proteins with motor function andneurodevelopment. Noteworthy, the state of actin remodelingdetermining neurologic manifestations is a diagnostic featureof ts3agn .Noteworthy, new knowledge about topologically associatingdomains (TADs) indicates that polytene, diploid, andembryonic TADs condensation along the chromosome axisis just the same everywhere . Moreover,comparison of TADs with 3D chromatin organization revealedby the Hi-C method confirms that the interphase nucleusspatial organization into TADs is directly represented by bandingpattern of polytene chromosomes .Therefore, this allows to bridge the ratio of genes forming ectopic contacts in a mode of either maternal, or paternal strainin reciprocal crosses and their physiologic manifestations.The later might result from alterations in the 11AB regionarchitecture. As shown in Fig. 3, a, the The regions with FECs similar in reciprocal hybrids, butdiffering from parents, i. e. manifesting hybrid properties,contain a large set of genes responsible for motor functions.ts3agn:meiosis, reparation, recombination; transcription factors; metabolism;proliferation; actin-binding proteins, microtubuleassociatedproteins.Figure 3, c shows genes and biological processes for chromosomalregions forming ectopic contacts with X:11AB onlyin hybrids or mainly in hybrids compared to parents. Theseprocesses are pertinent to main manifestations of ts3agn the chromosomalbands 8D, 12E, and 19D demonstrate FECs significantly exceedingthese of parents. These bands contain genes involvedin taste and odor perception and neurodevelopment, in particularof the mushroom bodies of the brain. The other examplesof father strain manifestations might result from activities oftrans-acting factors, such as miRNAs .Interestingly, in the cross Berlin\u00d7ts3agn and in progenyof Berlin\u00d7ts3agn (impaired 3-hour memory) and is similar towild type in the ts3agn\u00d7Berlin cross . Likely,miR-974 might act as trans-acting factor presumed to regulategenes in patroclinic mode. The prevailing role of the paternalgenome in memory formation is evident in Canton S and ts3agnreciprocal hybrids .miR-974 is involved in memory processes: its loweredexpression impairs memory formation .Decrease in its content in olfactory neurons and the mushroombody V2 neurons promotes 3-hour memory. Noteworthy, thecontent of miR-974 is decreased both in agnostic locusmight belong to the class of quantitative trait loci (QTL) .Taken together, our data indicate that the agnostic LIMK1 gene is a good candidate for linkingthe neuronal activity  and genetic apparatus . Additionally, quite recent andunexpected findings  reveal a new targetof intellectual disabilities: learning acquisition and memory erasure (forgetting) are governed by different signal cascades,correspondently cAMP-dependent and actin remodeling cascadesmall GTPase Rac1 \u2013 LIMK1 (the key enzyme of actinremodeling LIM-kinase 1) and its phosphorylation substrate\u0441ofilin. The absence of Rac1-dependent forgetting causesthe autistic spectrum disorders. Expression changes (activeor non-active state) of LIMK1 and cofilin lead to differentneurological disorders. Therefore, in the tradition of Russiangenetic school , the agnostic genemight be a functional link between genetic and cytogeneticprocesses within the nervous system and serve as a model forelucidating both the maternal and paternal modes of transgenerationalinheritance.One of the requirements of predictive and personalizedmedicine is consideration of POEs for prognosis of clinicalphenotype of many multifactorial neuropsychiatric disorders.These different and individual manifestations of cognitiveabilities and motor functions in patients with the same disease,i. e. behavioral plasticity, results from genome plasticity provokedby 3D chromatin architecture of the nerve cells nuclei.The evolutionary gene conservation approves the usage ofsimple low cost, fast and efficient models as Drosophila toprobe the causes, consequences and mechanisms of pathologyleading to human disease . 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How can we make sense of parent of origin effects inADHD and related behavioral phenotypes? Behav. Brain Funct.2015;11(1):33. DOI 10.1186/s12993-015-0078-4.Zerbino D.R., Achuthan P., Akanni W., Amode M.R., Barrell D.,Bhai J., Flicek P. Ensembl 2018. Nucleic Acids Res. 2018;46:D754-D761. DOI 10.1093/nar/gkx1098.Zhuravlev A. Homology Segment Analysis. Available at: https://bitbucket.org/beneor/homology-segment-analysis/src/master/ 2019a..Zhuravlev A. Homology Segment Analysis protocol. protocols.io.2019b. DOI 10.17504/protocols.io.bakyicxw.Zink F., Magnusdottir D.N., Magnusson O.T., Walker N.J., Morris T.J.,Sigurdsson A., Halldorsson G.H., Gudjonsson S.A., Melsted P., IngimundardottirH., Kristmundsdottir S., Alexandersson K.F., HelgadottirA., Gudmundsson J., Rafnar T., Jonsdottir I., Holm H.,Eyjolfsson G.I., Sigurdardottir O., Olafsson I., Masson G., GudbjartssonD.F., Thorsteinsdottir U., Halldorsson B.V., Stacey S.N., StefanssonK. Insights into imprinting from parent-of-origin phasedmethylomes and transcriptomes. Nat. Genet. 2018;50(11):1542-1552. DOI 10.1038/s41588-018-0232-7.Zykova T.Y., Levitsky V.G., Belyaeva E.S., Zhimulev I.F. Polytenechromosomes \u2013 a portrait of functional organization of the Drosophilagenome. Curr. Genomics. 2018;9(3):179-191. DOI 10.2174/1389202918666171016123830."}
+{"text": "Antenatal care is a unique opportunity to assess SARS-CoV-2 seroprevalence and antibody response in pregnant people, including those with previously unknown infection. Pregnant people were screened for SARS-CoV-2 IgG during antenatal care or delivery in Seattle, Washington with Abbott Architect chemiluminescent immunoassay which provides quantitative index (positive \u22651.4). Participants with IgG+ results or identified with RT-PCR+ results via medical records were invited to enroll in a longitudinal evaluation of antibody responses. We report preliminary results of an ongoing seroprevalence and longitudinal study with planned 18-month follow-up.\u2264280 days after first RT-PCR+ result for those with and \u2265104 days after first IgG detection for those without prior RT-PCR+ results), while 5/14 (26%) had a negative Abbott IgG test at a median of 81 days (IQR 75\u2013112) since initial testing.Between September 9, 2020\u2013May 7, 2021, we screened 1304 pregnant people; 62 (4.8%) tested SARS-CoV-2 IgG+, including 28 (45%) with known prior SARS-CoV-2 infection. Among participants testing IgG+, median age was 32 years (interquartile range [IQR] 26\u201335) and median gestational age was 21 weeks (IQR 12\u201338) at screening; median IgG index was 3.2 , including 3.9 (IQR 2.3\u20135.8) among those with vs. 2.7 (IQR 1.9\u20134.2) among those without prior RT-PCR+ results (p=0.05 by Wilcoxon rank-sum). Of 30 longitudinal study participants enrolled, 24 tested IgG+ at baseline (75% with prior RT-PCR+ result) and 6 tested IgG- on enrollment but were identified as previously RT-PCR+ via medical records; 24/30 (80%) reported previous symptoms. Of 24 participants testing IgG+ at baseline, 14 (58%) had first follow-up IgG results at median of 66 days (IQR 42\u2013104) since initial testing, with median IgG index of 2.0 (IQR 1.0\u20133.8). 9/14 (64%) participants with repeat IgG testing remained IgG+ at first follow-up  Alisa Kachikis, MD, MS, GlaxoSmithKline (Consultant)Pfizer (Consultant) Alexander L. Greninger, MD, PhD, Abbott (Grant/Research Support)Gilead (Grant/Research Support)Merck (Grant/Research Support) Janet A. Englund, MD, AstraZeneca GlaxoSmithKline (Research Grant or Support)Meissa Vaccines (Consultant)Pfizer (Research Grant or Support)Sanofi Pasteur (Consultant)Teva Pharmaceuticals (Consultant) Alison Drake, PhD, MPH, Merck (Grant/Research Support)"}
+{"text": "The cirrhosis-associated abnormalities of ACE, IL-6, VWF antigen, and antiplasmin parallel those observed in severe COVID-19.(1) Background: Cirrhotic patients have an increased risk for severe COVID-19. We investigated the renin-angiotensin-aldosterone system (RAS), parameters of endothelial dysfunction, inflammation, and coagulation/fibrinolysis in cirrhotic patients and in COVID-19 patients. (2) Methods: 127 prospectively characterized cirrhotic patients (CIRR), along with nine patients with mild COVID-19 (mild-COVID), 11 patients with COVID-19 acute respiratory distress syndrome , and 10 healthy subjects (HS) were included in the study. Portal hypertension (PH) in cirrhotic patients was characterized by hepatic venous pressure gradient (HVPG). (3) Results: With increased liver disease severity (Child\u2212Pugh stage A vs. B vs. C) and compared to HS, CIRR patients exhibited higher RAS activity (angiotensin-converting enzyme (ACE), renin, aldosterone), endothelial dysfunction (von Willebrand-factor (VWF) antigen), inflammation (C-reactive protein (CRP), interleukin-6 (IL-6)), and a disturbed coagulation/fibrinolysis profile . Increased RAS activity (renin), endothelial dysfunction (vWF), coagulation parameters  and inflammation  were significantly altered in COVID patients and followed similar trends from mild-COVID to ARDS-COVID. In CIRR patients, ACE activity was linked to IL-6 (\u03c1 = 0.26;  Severe acute respiratory distress syndrome-coronavirus-2 (SARS-CoV-2) causes substantial morbidity and mortality worldwide . CoronavPatients with ARDS due to COVID-19 exhibit severe endothelial damage, pulmonary microangiopathy, and thrombosis ,5, indicSimilarly, coagulation imbalance, inflammation, and RAS activation also represent well-known hallmarks of advanced chronic liver disease (ACLD) ,12,13. CPatients with ACLD are at particularly high risk for severe courses of COVID-19 ,17; howeACLD patients undergoing hepatic venous pressure gradient (HVPG) measurement at the Vienna General Hospital between February 2019 and December 2020 with portal hypertension (CIRR) were included in this study. Hemodynamic parameters including mean arterial pressure (MAP) were assessed at the time of HVPG measurement. Blood samples were withdrawn after the patients had rested for at least 30 min in supine position.Patients with intake of non-selective beta-blockers, antithrombotic or antiplatelet therapy, with active malignancy, portal vein thrombosis, porto-sinusoidal vascular disease or cardiac cirrhosis, as well as patients after liver transplantation were excluded from the study. If multiple HVPG measurements were performed in the same patient, only the baseline measurement  was used for this study. The ACLD patients were stratified by Child\u2212Turcotte\u2212Pugh (CTP) stage A, B, and C and by HVPG . Etiology of ACLD, comorbidities , age, and intake of concomitant medication were recorded.Moreover, inpatients at the General Hospital of Vienna with COVID-19 (mild-COVID), inpatients with ARDS due to COVID-19 (ARDS-COVID), as well as outpatients without ACLD or COVID-19  willing to participate in the study were included.For HVPG measurement, the right jugular vein was accessed via Seldinger technique using a catheter introducer set . A balloon catheter  was subsequently used for liver vein cannulation . HVPG waA colorimetric assay  was used to determine ACE activity. Plasma concentrations of renin  and aldosterone  were measured by chemiluminescence-immunoassay. VWF antigen was measured by latex agglutination assay . Prothrombin fragment F1,2 and plasminogen activator inhibitor (PAI) were assessed by ELISA , plasminogen activity, and \u03b1-2 antiplasmin (antiplasmin) activity by chromogenic assay . Routine laboratory parameters including D-dimer, C-reactive protein (CRP), and interleukin-6 (IL-6) were assessed by standard laboratory methods.Categorical variables were presented as number (n) of patients and % of these patients with the characteristic of interest, while continuous data was reported as median and interquartile range (IQR). D\u2019Agostino and Pearson and Shapiro\u2013Wilk normality tests were used to test for normal distribution. Mann-Whitney U test was implemented for comparing non-normally distributed continuous variables between two groups, and Kruskal-Wallis test for comparison of non-normally distributed continuous variables in three or more groups. Dunn\u2019s multiple comparisons test was used as post-hoc test. Group comparisons of categorical variables were computed using Pearson\u2019s Chi-squared or Fisher\u2019s exact test.p < 0.100) were included in the multivariate model. Multicollinearity was investigated via variance inflation factor (VIF). GraphPad Prism 8  and IBM SPSS 22.0 statistic software  were used for statistical analysis. A two-sided p-value of <0.05 was considered as being statistically significant.Factors associated with ACE plasma levels were assessed using linear regression models including either VWF antigen, prothrombin fragment F1,2, or antiplasmin activity. Parameters that showed a trend  (NCT03267615).The study was approved by the ethics committee (EC) of the Medical University of Vienna (study number: 1461/2020 and 1262/2017). It was performed according to the current version of the Helsinki Declaration. All patients were prospectively included in the study and gave their informed consent before the blood withdrawal. The cirrhotic cohort of this study is part of the Vienna Cirrhosis Study registry, registered under p < 0.001).In total, 127 CIRR patients with male predomination  were included in the study. Median age was 56.6 (IQR 15.5) years. Alcoholic (49.6%), viral (15.0%), and cholestatic liver disease (8.7%) were the main etiologies . There wThe mild-COVID cohort consisted of nine patients with COVID-19, who were admitted to the Vienna General Hospital due to the viral infection , while the ARDS-COVID cohort included 11 patients  and the HS cohort included 10 subjects  .p < 0.001) and VWF antigen (A: 223.4 (IQR 114.0)% vs. B: 307.0 (IQR 133.0)% vs. C: 396.0 (91.0)%; p < 0.001) significantly increased with decreasing liver function, while there was a trend for PAI levels (A: 0.89 (1.60) IU/mL vs. B: 0.92 (2.06) IU/mL vs. C: 2.68 (1.73) IU/mL; p = 0.149) (p < 0.001), plasminogen activity (A: 75.0 (21.0)% vs. B: 59.0 (12.0)% vs. C: 42.0 (9.0)%; p < 0.001), and in tendency prothrombin fragment F1,2 (A: 287.0 (292.0) pmol/L vs. B: 292.5 (241.0) pmol/L vs. C: 176.0 (194.0) pmol/L; p = 0.079) decreased with increasing CTP stage (D-dimer (A: 0.48 (IQR 0.41) \u00b5g/mL vs. B: 1.40 (IQR 2.90) \u00b5g/mL vs. C: 3.19 (3.08) \u00b5g/mL; = 0.149) . AntiplaTP stage . SimilarTP stage .p < 0.001) (p < 0.001). Similarly, more pronounced RAS activation was observed in patients with more advanced hepatic dysfunction: ACE (A: 41.3 (35.9) U/L vs. B: 54.4 (45.6) U/L vs. C: 71.3 (66.9) U/L; p = 0.006), as well as plasma renin (A: 12.4 (26.7) \u00b5IU/mL vs. B: 55.1 (115.9) \u00b5IU/mL vs. C: 594.1 (949.8) \u00b5IU/mL; p < 0.001) and aldosterone concentrations (A: 84.5 (442.0) pg/mL vs. B: 240.0 (442.0) vs. C: 397.0 (238.0) pg/mL; p < 0.001).With increasing CTP stage, there was increased inflammation, as indicated by rising CRP (A: 0.18 (0.29) mg/dL vs. B: 0.47 (0.67) mg/dL vs. C: 0.47 (1.23) mg/dL; < 0.001)  and IL-6p = 0.009), vWF antigen (mild-COVID: 247.5 (73.0) vs. ARDS-COVID: 420.0 (99.0)%; p = 0.007), D-dimer (mild-COVID: 0.54 (0.58) \u00b5g/mL vs. ARDS-COVID: 2.94 (4.75) \u00b5g/mL; p < 0.001), prothrombin fragment F1,2 (mild-COVID: 151.5 (252.0) pmol/L vs. ARDS-COVID: 429.0 (2687.0) pmol/L; p = 0.006), CRP (mild-COVID: 0.50 (1.42) mg/dL vs. ARDS-COVID: 15.10 (14.69) mg/dL; p < 0.001), and IL-6 (mild-COVID: 7.66 (14.74) pg/mL vs. ARDS-COVID: 65.70 (357.90) pg/mL; p < 0.001), indicating an increased RAS activation, endothelial dysfunction, coagulation/fibrinolysis activation, and inflammation  \u00b5IU/mL vs. ARDS-COVID: 62.7 (107.1) \u00b5IU/mL; ammation .Compared to healthy control subjects, cirrhotic patients and COVID-19 patients exhibited markedly increased VWF antigen, while the highest median VWF antigen level occurred in ARDS-COVID patients . Both ciMoreover, compared to healthy subjects, cirrhotic and COVID-19 patients had higher levels of CRP and IL-6.On the other hand, cirrhotic patients exhibited significantly lower antiplasmin activity and plasminogen activity than COVID-19 patients, while ACE activity and plasma aldosterone concentration were higher.p < 0.001), D-dimer , prothrombin fragment F1,2 , antiplasmin activity , IL-6 , and HVPG   .p = 0.001), coagulation , and of fibrinolysis , but not with parameters of inflammation   .In this study, we thoroughly characterized the state of components of the RAS, coagulation, and inflammation in cirrhotic patients of different disease severity and compared the findings to patients with mild COVID-19 and with COVID-ARDS. Importantly, we identified profound abnormalities of the coagulation system and upregulated systemic inflammation linked to cirrhosis-associated RAS activation that followed similar trends in COVID-19.By stratifying our ACLD cohort by hepatic dysfunction , as well as for portal pressure (by the diagnostic gold-standard HVPG), we observed marked endothelial dysfunction  and dysbalanced coagulation state  with increasing ACLD severity. At the same time, inflammatory markers , along with RAS components, were elevated with more severe ACLD, indicating a state of coagulation/fibrinolysis activation linked to RAS activity\u2014that reportedly is upregulated in cirrhosis ,22.COVID-19 patients exhibited very similar levels of endothelial dysfunction, coagulation/fibrinolysis, and inflammation parameters, compared to cirrhotic patients, as well as RAS activation, indicated by increased plasma renin concentration. Interestingly, these parameters were also the most important factors differing between mild-COVID and ARDS-COVID. This suggests that cirrhotic patients are in a disadvantageous position when contracting SARS-CoV-2 infection. Importantly, the pronounced RAS activation causing associated coagulation imbalance and a proinflammatory state in Child\u2212Pugh B/C patients may explain why cirrhotic patients have been reported to be particularly susceptible to severe courses of COVID-19 ,17.The RAS is intricately involved in cirrhosis development and progression . ImportaIn COVID-19, especially the non-classical RAS is subject of scientific interest, as SARS-CoV-2 uses the ACE2 receptor for cell entry  and it hCOVID-19 is strongly linked to thromboembolic complications, including microvascular thrombosis, but also macrovascular events such as stroke or pulmonary embolism ,30. EndoSimilarly, D-dimer and prothrombin fragment F1,2 were also increased both in cirrhosis and COVID-19, signifying an activation of coagulation and fibrinolysis, with the highest values in ARDS-COVID patients. Interestingly, prothrombin fragment F1,2 levels were highest in Child-A patients, indicating coagulation dysregulation already in an early stage of cirrhosis. Elevated D-dimer was identified as a marker of poor prognosis in patients with COVID-19 . The higFurthermore, high plasmin levels are observed in liver cirrhosis, likely due to decreased antiplasmin and elevated tissue-type plasminogen activator activity . ImportaFinally, both liver cirrhosis and COVID-19 were associated with increased systemic inflammation. As an infectious disease, COVID-19 triggers a complex immune response, leading to a pronounced (and sometimes excessive) inflammatory reaction . Again, Elevation of inflammatory parameters in cirrhosis may be due to gut-derived bacterial translocation ; howeverOur study has limitations: The cohorts were not age- and sex-matched. While cirrhotic patients were mainly male, the COVID-19 cohort was mainly female. Moreover, the healthy control group was significantly younger than the other groups, which may have impacted laboratory tests such as VWF, which increases with age . HoweverIn conclusion, we comprehensively investigated the role of the RAS, endothelial dysfunction, coagulation, and inflammation both in patients with cirrhosis and with COVID-19. Importantly, we found striking similarities between cirrhotic patients and COVID-19 patients, as both groups suffer from endothelial dysfunction, coagulation/fibrinolysis activation, and a systemic pro-inflammatory state. The \u201cbaseline-upregulation\u201d of the RAS in the setting of advanced cirrhosis may facilitate SARS-CoV-2 cell entry due to decreased antiplasmin activity and consecutively elevated plasmin levels. The molecular mechanisms driven by cirrhosis-associated RAS activation\u2014including the induction of an imbalanced coagulation profile, endothelial dysfunction, and systemic inflammation\u2014may mechanistically explain a susceptibility to severe COVID-19 in cirrhotic patients."}
+{"text": "Correction to: BMC Health Serv Res 21, 733 (2021)https://doi.org/10.1186/s12913-021-06757-x7,8 to D. Laplanche7.Following publication of the original article , the affThe author affiliation list has been updated above and the original article  has been"}
+{"text": "Cohort studies suggest higher rates of discontinuations (DCs) and adverse events (AEs) with integrase inhibitors (INSTIs) than is reported in clinical trials. Here, we assess DC of different INSTIs in combination with one of two tenofovir prodrugs in the first year following initiation defined as \u201cearly DC\u201d in a real-world cohort of treatment-na\u00efve patients.This analysis evaluated treatment-na\u00efve patients at a single center initiating raltegravir (RAL), elvitegravir/cobicistat (EVG/c), dolutegravir (DTG) or bictegravir (BIC) in combination with emtricitabine/tenofovir alafenamide (F/TAF) or emtricitabine/tenofovir disoproxil fumarate (F/TDF) between 10/2007-1/2020. Eligible patients had a minimum follow-up of 1 year. The primary endpoint was incidence of early INSTI DC. Secondary endpoints included AEs and risk factors for early INSTI DC and treatment-related AEs.331 patients were included. Median age was 32 years, 89% were male, 43% were non-White, 8% started RAL-based therapy, 46% started EVG/c-based therapy, 22% started DTG-based therapy and 24% started BIC/F/TAF. 36 discontinued INSTI-based therapy early yielding an incidence rate of 0.17 DCs per person-years (PPY) among RAL patients, 0.14 DCs PPY among EVG/c patients, 0.22 DCs PPY among DTG patients, and 0 DCs PPY among BIC patients, p=0.006. Treatment-related AEs occurred in 27% of RAL patients, 42% of EVG/c patients, 50% of DTG patients, and 43% of BIC patients p=0.607; and were responsible for early DC rates of 0.022 in 3 EVG/c patients and 0.075 in 5 DTG patients. No treatment-related early DCs occurred among RAL or BIC patients. No evaluated factor was significantly associated with early INSTI DC, however DTG use was significantly associated with treatment-related AEs .Table 1. Risk factors for early integrase inhibitor discontinuation and treatment-related adverse eventsIn this cohort, early DCs occurred in 11% initiating INSTI-based therapy, however of these only 2% were treatment-related. These data support use of INSTI-based regimens as preferred options for treatment-na\u00efve patients living with HIV due to their favorable safety and tolerability profiles.Charlotte-Paige M. Rolle, MD MPH, Gilead Sciences Janssen Infectious Disease ViiV Healthcare  Kiran Patel, PharmD, Gilead Sciences (Employee) Federico Hinestrosa, MD, AbbVie (Speaker\u2019s Bureau)Gilead Sciences Theratechonologies (Advisor or Review Panel member)ViiV Healthcare  Edwin DeJesus, MD, Gilead Sciences"}
+{"text": "Klebsiella pneumoniae is an increasingly important hospital pathogen. Classical K. pneumoniae (cKp) and hypervirulent K. pneumoniae (hvKp) are two distinct evolutionary genetic lines. The recently ongoing evolution of K. pneumoniae resulted in the generation of hybrid hvKP-MDR strains. K. pneumoniae distinct isolates (n = 70) belonged to 20 sequence types with the prevalence of ST395 (27.1%), ST23 (18.6%), ST147 (15.7%), and ST86 (7.1%), and 17 capsular types with the predominance of K2 (31.4%), K57 (18.6%), K64 (10.0%), K1 (5.7%) were isolated from patients of the Moscow neurosurgery ICU in 2014\u20132019. The rate of multi-drug resistant (MDR) and carbapenem-resistant phenotypes were 84.3% and 45.7%, respectively. Whole-genome sequencing of five selected strains belonging to cKp (ST395K47 and ST147K64), hvKp (ST86K2), and hvKp-MDR (ST23K1 and ST23K57) revealed blaSHV, blaTEM, blaCTX, blaOXA-48, and blaNDM beta-lactamase genes; acr, oqx, kpn, kde, and kex efflux genes; and K. pneumoniae virulence genes. Selective pressure of 100 mg/L ampicillin or 10 mg/L ceftriaxone induced changes of expression levels for named genes in the strains belonging to cKp, hvKp, and hybrid hvKp-MDR. Obtained results seem to be important for epidemiologists and clinicians for enhancing knowledge about hospital pathogens. Klebsiella pneumoniae is an increasingly important hospital pathogen causing a wide range of infections including urinary tract infections, pneumonia, bacteremia, and liver abscesses. In severe clinical cases, it can also lead to multiple organ failure, or even death [K. pneumoniae (cKp) and hypervirulent K. pneumoniae (hvKp), were described and are both global pathogens [K. pneumoniae strains belong to particular clones  producing beta-lactamases in combination with other functional classes of resistance determinants [K. pneumoniae were attributed to sequence types ST23, ST86, ST65, etc., and capsular types K1, K2, K57, K20, etc. [en death . Two difathogens . Most muK. pneumoniae resulting in the generation of hybrid hvKp-MDR strains. Mechanisms for the emergence of such strains can be a result of acquiring hypervirulent plasmids by cKp [Recent studies have shown the ongoing evolution of s by cKp , acquiris by cKp , and acqs by cKp . HoweverK. pneumoniae strains collected in Moscow neurosurgery ICU in 2014\u20132019, to identify resistance and virulence genes in their cells, and to estimate relative expression levels of such genes in selected strains belonging to epidemiology significant genetic lines cKp (ST395K47 and ST147K64), hvKp (ST86K2), and hybrid hvKp-MDR (ST23K1 and ST23K57).This study aimed to determine the genetic lines of K. pneumoniae caused about 28% among the agents of nosocomial infections in neurosurgery ICU during the period from January 2014 to May 2019. The incidence rates of K. pneumoniae infections were 8.0 per 100 patient infections of the central nervous system, 4.3/100 of bloodstream infections, 26.3/100 of respiratory infections, and 25.3/100 of urinary tract infections [K. pneumoniae clinical isolates were collected from 283 patients in this period, including those isolated from the respiratory system (n = 271), urine (n = 166), the nervous system (n = 41), blood (n = 36), surgical wounds (n = 27), and other (n = 4).n = 34), urine (n = 19), the nervous system (n = 8), blood (n = 6), and surgical wounds (n = 3). As a result, 20 sequence types were identified, the majority of them were ST395, ST23, ST147, and ST86, and a total of 17 capsular types were identified. Predominant K-types were K2, K57, K64, and K1 , ampicillin-sulbactam (90.0%), cefuroxime (86.8%), cefoxitin (68.8%), ceftriaxone (77.8%), ceftazidime (72.8%), cefoperazone-sulbactam (82.7%), cefepime (69.2%), ertapenem (48.1%), tetracycline (80.7%), ciprofloxacin (81.2%), chloramphenicol (77.0%), gentamicin (67.1%), tobramycin (83.0%), trimethoprim-sulfamethoxazole (69.2%), and nitrofurantoin (82.2%). Fewer resistant isolates were found for imipenem (33.8%), tigecycline (37.2%), and amikacin (31.8%) (It was shown that major  (31.8%) .K. pneumoniae isolates carried beta-lactamase genes blaSHV (100.0%), blaCTX-M (74.2%), blaTEM (51.4%), blaOXA-48 (40.0%), blaNDM (11.4%), class 1 (37.6%) and 2 (2.1%) integrons, and porin protein gene ompK36 (91.5%). Both blaOXA-48 and blaNDM carbapenemase genes were detected in four (5.7%) isolates. Beta-lactamase genes blaKPC, blaVIM, and blaIMP were not detected (The frequency of the multi-drug resistant (MDR) phenotype was 84.3%, with the carbapenem-resistant phenotype consitituting 45.7%. detected .K. pneumoniae isolates: rmpA (34.2%), aer (21.4%), kfu (17.1%), uge (85.7%), wabG (100.0%), fimH (97.1%), allS (94.3%), and allR (5.7%). Twelve virulence gene combinations were identified. The most prevalent combination was uge+wabG+fimH+allS (51.4% isolates), followed by rmpA+aer+uge+wabG+fimH+allS (11.4%), uge+wabG+kfu+fimH+allS (7.1%), rmpA+wabG+fimH+allS (5.7%), and rmpA+aer+ +uge+wabG+kfu+fimH+allR (5.7%). The rarest combinations were wabG+fimH+allS (4.3%), rmpA+uge+wabG+fimH+allS (4.3%), uge+wabG+allS (2.8%), rmpA+aer+wabG+fimH+allS (2.8%), rmpA+wabG+kfu+fimH+allS (1.4%), rmpA+aer+uge+wabG+kfu+fimH+allS (1.4%), and rmpA+wabG+kfu+fimH+allS (1.4%) (Eight virulence genes were detected in 70 S (1.4%) .n = 9) and urine (n = 2), belonging to genetic lines ST23K1 (n = 4), ST86K2 (n = 3), ST23K57 (n = 2), ST65K2 (n = 1), and ST218K57 (n = 1). Isolates of ST23K1 carried rmpA+aer+uge+wabG+kfu+fimH+allR; isolates of ST86K2 and ST218K57 harbored rmpA+aer+uge+wabG+fimH+allS; isolates of ST23K57 carried two combinations, rmpA+uge+wabG+fimH+allS and rmpA+wabG+kfu+fimH+allS; and isolates of ST65K2 carried rmpA+aer+uge+wabG+kfu+fimH+allS virulence gene combinations. All 11 hypermucoviscous isolates carried rmpA, wabG, and fimH genes. Major isolates of hypermucoviscousity-positive isolates were MDR (n = 6), resistant to 3\u20137 antimicrobial functional groups . Among them, two isolates were CR. The remaining isolates were resistant to ampicillin only. As a result, there was no revealed correlation between the hypermucoviscousity phenotype and genetic line or the antimicrobial phenotype or virulence gene profiles (Phenotype of hypermucoviscousity was detected for 11 of 70 isolates (15.7%) collected from the respiratory system , hvKp (ST86K2), and hvKp-MDR (ST23K1 and ST23K57), were analyzed by whole-genome sequence and quantitative analyses of antimicrobial resistance and virulence gene expression. One of these strains (B2523/18) was resistant to AMP and carried only one beta-lactamase gene of the blaSHV type. In contrast, the other four strains that belonged to the MDR category were resistant to 10\u201318 antimicrobials. The latter strains carried beta-lactamase genes of blaSHV, blaTEM, blaCTX-M, blaOXA-48, and blaNDM types and the resistance determinants for other antimicrobial groups in the genomes: aminoglycosides, fosfomycin, streptogramins, phenicols, quinolones, sulfonamide, tetracyclines, sulfonamides, macrolides, and rifamycins, as well as genes coding 4\u20135 efflux pumps and 1\u20134 genes of heavy metal resistance. The following genetic determinants of K. pneumoniae virulence were identified in the genomes: peg-344 (n = 3), rmpA (n = 3), rmpA2 (n = 1), iroB (n = 2), iroN (n = 2), iroD (n = 2), uge (n = 5), wabG (n = 5), kfu (n =1 ), fimH (n = 5), allR (n = 1), irp (n = 4), iuc (n = 3), entB (n = 3), iut (n = 4), mrk (n = 5), ybt (n = 4), fyu (n =4 ), treC (n = 5), celB (n = 5), and ureA (n = 5)  .K. pneumoniae strains during growth without selective pressure of antimicrobials were different: the chromosomal beta-lactamase genes blaSHV were transcribed significantly lower compared with those of the reference gene rpoD. In contrast, other beta-lactamase genes , as well as porin gene ompK36 were transcribed at higher levels compared with the reference gene, with the exception of blaTEM in the hvKp-MDR strain of ST23K1 which exhibited lower expression. The efflux pump genes and the virulence genes were expressed mostly at the same or lower levels compared with the rpoD gene, with the exception of two virulence genes: treC in the strains of ST395K47, ST147K64, ST23K1, and ST23K57; and celB in the strain of ST86K2  gene, and downregulation of the blaTEM (5.5-fold), acrA (2.4-fold), acrB (2.8-fold), oqxB (4.8-fold), kpnE (3.5-fold), kpnF (2.0-fold), and kdeA (3.0-fold) genes. Another MDR strain of ST147K64 was characterized by the upregulation of the blaTEM (2.2-fold), kpnE (3.2-fold), kdeA (2.7-fold), fimH (2.3-fold), and ureA (14.0-fold) genes, and downregulation of the blaOXA-48 (5.3-fold) gene. The hvKp strain of ST86K2 demonstrated upregulation of only one gene, iroN (4.1-fold), and downregulation of 13 genes: blaSHV (2.6-fold), acrA (9.0-fold), acrB (10.0-fold), oqxB (10.1-fold), kpnE (9.9-fold), kpnF (5.0-fold), kexD (4.9-fold), kdeA (4.8-fold), iroD (4.0-fold), uge (3.2-fold), wabG (6.2-fold), treC (4.8-fold), and ureA (5.3-fold). The hybrid hvKp-MDR strain of ST23K1 showed upregulation of the acrB (3.4-fold), oqxA (2.1-fold), oqxB (2.1-fold), wabG (6.2-fold), fimH (3.2-fold), celB (8.0-fold), and ureA (8.9-fold) genes. The second hvKp-MDR strain of ST23K57 expressed upregulation of the blaTEM (4.1-fold), blaCTX-M (5.3-fold), ompK36 (2.9-fold), acrB (2.3-fold), oqxA (2.9-fold), oqxB (4.6-fold), kpnE (2.3-fold), kpnF (2.0-fold), kexD (3.2-fold), kdeA (11.7-fold), uge (3.4-fold), wabG (14.5-fold), fimH (2.9-fold), celB (3.7-fold), and ureA (3.1-fold) genes  genes .K47 demonstrated downregulation of the blaTEM (3.3-fold) and acrA (2.4-fold) genes. The MDR strain of ST147K64 was characterized by upregulation of the oqxB (2.7-fold), kpnE (7.0-fold), kpnF (4.4-fold), kdeA (2.3-fold) genes, and downregulation of the blaOXA-48 (2.1-fold) gene. The hybrid hvKp-MDR strain of ST23K1 showed upregulation of the fimH (2.1-fold) gene, and downregulation of the blaTEM (2.1-fold), kdeA (5.1-fold), and ureA (2.7-fold) genes. The second hybrid hvKp-MDR strain of ST23K57 expressed upregulation of the blaSHV (2.3-fold), blaTEM (2.4-fold), blaCTX-M (6.3-fold), acrB (3.0-fold), oqxB (3.8-fold), kpnE (2.1-fold), kdeA (8.6-fold), wabG (14.8-fold), and ureA (4.2-fold) genes, and no genes were downregulated  [(16\u201320%) , and sig(16\u201320%) .K. pneumoniae isolates collected from 62 patients were attributed to specific genetic lines, virulence, and antimicrobial resistance genotypes. Single isolates were collected from 54 patients and two isolates from eight patients. Double isolates were studied from one patient in a case of their differences in ST , K-type (Patient 35), and antimicrobial resistance genes profiles (Patients 4 and 39) (K64 carried (blaTEM+blaSHV+blaCTX-M+blaOXA-48+blaNDM) and (blaSHV+blaOXA-48) beta-lactamase genes, respectively. Two isolates obtained from the trachea of Patient 39 belonged to ST23K57 (blaTEM+blaSHV+blaCTX-M) and (blaSHV+blaCTX-M+blaOXA-48) beta-lactamase genes, respectively. Different antimicrobial resistance gene profiles of K. pneumoniae named isolates possibly indicate the evolution events in the patient\u2019s body as described previously [Non-duplicate 70  and 39) . Two isoeviously .K. pneumoniae genetic lines common for classical K. pneumoniae (cKp) (ST395 and ST147) and hypervirulent K. pneumoniae (hvKp) (ST23 and ST86). ST395 was reported as prevalent for cKp-MDR strains in studies from Poland, France, Italy, and Russia [K. pneumoniae of ST23 and ST86 were described previously as hvKp causing bacteremia, sepsis, and liver abscess in India, France, Taiwan, and Russia [K. pneumoniae isolates of K64 commonly were recognized as cKp, but recently some isolates of K64 were described as hybrid hvKp-MDR due to acquiring the hvKp virulence plasmids [K20/K64, ST395K2, as well as hvKp ST23K1 and ST86K2 [K. pneumoniae isolates in our study were MDR with resistance to carbapenems for 45.7% isolates. Such high MDR rates were reported from China and Iran [blaCTX-M in our study was 74.2%, carbapenemase genes blaOXA-48 and blaNDM at 40.0% and 11.4%, respectively, similar to the prevalence of these genes published previously [n = 11) were attributed to ST23K1, ST86K2, ST23K57, ST65K2, and ST218K57 genetic lines. All of them carried the rmpA gene coding the major virulence-associated factor in hvKP isolates [rmpA-positive isolates in this study were hypermucoviscousity-positive, which is in agreement with a previously published report [K. pneumoniae among hypermucoviscousity-positive isolates in this study was 6/11, which is consistent with the modern trend for the appearance of hybrid hvKp-MDR genetic lines [Twenty STs and 17 K-types were identified. The most prevalent sequence types were ST395 27.1%), ST23 18.6%), ST147 15.7%), and ST86 (7.1%). The remaining 16 STs were each represented at \u2264 4.3%. The dominant STs were described previously as .7%, and .6%, ST14d Russia ,20,21,22plasmids . Moreoved ST86K2 ,27,28,29.1%, ST23and Iran ,31. The isolates , althougic lines ,34.K. pneumoniae strains belonging to prevalent STs and K-types  were selected for further comparative study of whole-genome sequences, antimicrobial resistance phenotypes, hypermucoviscosity, and altered expression levels of virulence and resistance genes in response to beta-lactams effect (AMP and CRO). It was shown that K. pneumoniae isolates belonging to ST86K2, ST23K1, and ST23K57 demonstrated hypermucoviscosity phenotype in contrast with isolates of ST395K47 and ST147K64, which confirmed the virulent phenotype of three isolates. All K. pneumoniae isolates carried blaSHV genes, including extended-spectrum beta-lactamase (ESBL) variant blaSHV-12, broad-spectrum variants blaSHV-28 and blaSHV-33, and narrow-spectrum variants blaSHV-67 and blaSHV-190. These alleles of blaSHV genes were previously described in Portugal, Turkey, China, Russia, and Spain [K. pneumoniae isolates  carried the blaTEM-1B, blaCTX-M-15, and blaOXA-1 genes. It was reported that these genes were horizontally transferred by the IncFIA-FIB-FII and IncHI2 plasmids [K57) carried the blaCTX-M-55 and blaOXA-1 genes; such gene combination was reported previously from China [K47 and ST147K64 additionally carried two carbapenemase genes, i.e., blaOXA-48 and blaNDM-1. Previously, it was reported that K. pneumoniae clinical isolates of ST395 and ST147 harbored blaNDM-5 and blaOXA-181/232 in Nepal and that ST11 harbored blaNDM-1 and blaOXA-48 in Greece [K. pneumoniae isolates belonging to the hvKp evolutionary branch, which acquired the resistance genes and became a hybrid hvKp-MDR. In our study, a hvKp-MDR isolate of ST23K1 carried simultaneously blaCTX-M-15 and blaOXA-48 genes, similar to a recently published study [K57 carried not only the blaCTX-M-55 and blaOXA-48 genes but additionally the blaNDM-1 gene. This is the first report describing K. pneumoniae of ST23K57 genetic-line-acquired cefalosporinase gene blaCTX-M-55, and two carbapenemase genes blaOXA-48 and blaNDM-1, which is particularly alarming. The incidence of high-risk clone ST383 carrying blaCTX-M-14b gene and two carbapenemase genes blaNDM-1 and blaOXA-48 combining both resistance and virulence elements was recently published [blaCTX-M-15, blaNDM, and blaOXA-181 genes [acrA, acrB, oqxA, oqxB, kpnE, kpnF, kdeA, and kexD) because of their clinical significance for K. pneumoniae beta-lactam resistance presented recently [K1 carried four efflux pump genes: acr and oqx of RND-type systems, kde of MATE-type, and kpn of SMF-type. The rest of the four strains carried five efflux pumps: acr, oqx, kde, kpn, and additionally the kex gene of the RND-type efflux system. The same efflux pump genes were detected recently in K. pneumoniae clinical isolates, which exhibited co-resistance to beta-lactams and aminoglycosides, glycopeptides, fluoroquinolones, and tetracyclines in India [Five nd Spain ,35,36. Tplasmids ,38. One n Greece . Of greaed study . Moreove81 genes . Additiorecently . In our in India . Interesin India .peg-344, iroB, iucA, plasmid-encoded rmpA, and rmpA2 and quantitative siderophore production (entB and ybtS) [K2, ST23K1, and ST23K57. In contrast, K. pneumoniae virulence genes , common for both hvKp and cKp, were detected in all five isolates. This is in agreement with recently published data [K1 and ST23K57 are characterized as hybrid hvKp-MDR. Thus, the data obtained in this study indicate the ongoing formation of hybrid K. pneumoniae on the base of the ST23 genetic line, which was already defined in the last decade [It is known that multiple biomarkers have been shown to predict hvKp isolates: nd ybtS) . In thist decade ,35,45.K. pneumoniae resistance and virulence genes at non-selective conditions in vitro and the fold change of expression levels in presence of AMP and CRO. It was shown that ESBL gene blaSHV-12 expressed ~4-fold higher in the MDR strain of ST395K47 than blaSHV-type genes coding broad-spectrum and narrow-spectrum beta-lactamases in the remaining K. pneumoniae strains. This is in agreement with previously reported data that ESBL variants of the blaSHV-12 gene expressed higher than non-ESBL variants [We estimated the basal expression levels of variants . These gblaTEM,blaCTX-M, blaOXA-48, and blaNDM and porin gene ompK36 were expressed at higher levels, with the exception of blaTEM in the hvKp-MDR strain of ST23K1. Notably, the expression levels of the beta-lactamase (blaTEM and blaCTX-M) and carbapenemase  genes were higher in cKp-MDR strains than those in hybrid hvKp-MDR strains. Possibly, the reason for this observation is the higher metabolic load in Klebsiella cells producing resistance and virulence factors simultaneously. Growing at AMP conditions for 90 min induced upregulation of blaTEM gene expression in cKp-MDR of ST147K64 (2.1-fold), as well as the blaCTX-M (5.3-fold) and ompK36 (2.8-fold) genes in hvKp-MDR of ST23K57, but downregulation of blaTEM gene expression in cKp-MDR of ST395K47 and the blaOXA-48 gene in cKp-MDR of ST147K64. In contrast, growing at CRO conditions induced upregulation of the blaCTX-M gene in hvKp-MDR of ST23K57 (6.5-fold) and downregulation of the blaTEM gene in cKp-MDR of ST395K47 (3.3-fold), in hvKp-MDR of ST23K1 (2.1-fold), and the blaOXA-48 gene in cKp-MDR of ST147K64 (2.1-fold).Beta-lactamase genes ropD, and 2\u20133 genes were lower than the reference. In the hvKp strain, one efflux gene expression was higher, three genes were expressed at the same level, and four genes were lower than the reference gene. In contrast, major efflux genes in hybrid hvKp-MDR strains were expressed lower than the rpoD gene, and one gene in the strain of ST23K57 was expressed on the same level as the reference. Interestingly, the previously described expression of the arcB efflux gene showed upregulation of this gene in carbapenem-resistant K. pneumoniae strains compared with non-resistant ones [Klebsiella evolutionary branches that are consistent with previous reports [K. pneumoniae genetic lines may reflect differences in bacterial surface structures in particular K-types: downregulation in the strains of K47 and K2, and upregulation in the strains of K1 and K57 in response to AMP; and upregulation in the strains of K64 and K57 in response to CRO  were present at higher levels in the cKp strains of ST395K47, ST147K64, and hvKP ST86K2 than in the hybrid hvKp-MDR strains of ST23K1 and ST23K57. In conditions containing 100 mg/L AMP, these genes were upregulated in cKp and hybrid hvKp-MDR strains (fimH 2.3-4.6-fold) and wabG in hvKp-MDR strains (6.1\u201314.9-fold), while downregulated in the hvKp strain . In conditions with 10 mg/L CRO, only the wabG gene was upregulated in hybrid hvKp-MDR strains (2.3-14.9-fold). The expression levels of the remaining virulence genes common for cKp and hvKp (celB and ureA), as well as common for only hvKp , were approximately equal in all studied strains at non-selective conditions. The celB gene expression at AMP medium was upregulated in hvKp-MDR strains (3.7\u20138.0-fold); the ureA gene expression was upregulated in the cKp strain of ST147K64 (13.9-fold) and hvKp-MDR strains of ST23K1 (9.2-fold) and ST23K57 (3.2-fold). It was detected that CRO induced upregulation of the ureA gene in a hvKp-MDR strain of ST23K57 (4.3-fold) and downregulation of this gene in a hvKp-MDR strain of ST23K1 (2.6-fold). Expression levels of virulence genes common for hvKp strains were not changed under selective pressure generated by AMP or CRO in the growth media  in a specialized Neurosurgical Hospital in Moscow, Russia. Following the requirements of the Russian Federation Bioethical Committee, each patient signed informed voluntary consent to treatment and laboratory examination. The materials used in the study did not contain the personal data of patients.K. pneumoniae isolates were collected from the respiratory system, blood, urine, cerebrospinal fluid, and wounds of 62 patients of the neuro-ICU. Bacteria identification was performed using by a Vitek-2 Compact instrument  and a MALDI-TOF Biotyper . Bacteria were grown at 37 \u00b0C with agitation on Luria-Bertani broth  and Muller-Hinton broth . Bacterial isolates were stored in 20% glycerol at minus 80 \u00b0C.Seventy n-101 and AST n-102 cards . The results were interpreted according to the European Committee on Antimicrobial Susceptibility Testing . E. coli strains ATCC 25922 and ATCC 35218 were used for quality control. Strains non-susceptible to \u22651 agent in \u22653 antimicrobial categories were identified as multi-drug resistant (MDR), according to Magiorakos et al., 2012 [Minimal inhibitory concentrations (MICs) of ampicillin (AMP), ampicillin-sulbactam (SAM), cefuroxime (CXM), cefoxitin (FOX), ceftriaxone (CRO), ceftazidime (CAZ), cefoperazone-sulbactam (CSL), cefepime (FEP), ertapenem (ETP), imipenem (IPM), tetracycline (TET), tigecycline (TGC), ciprofloxacin (CIP), chloramphenicol (CHL), gentamicin (GEN), tobramycin (TOB), amikacin (AMK), trimethoprim-sulfamethoxazole (SXT), and nitrofurantoin (NIT) were determined using a Vitek-2 instrument with VITEK-2 using AST l., 2012 .K. pneumoniae strains growing on the plates with Luria-Bertani broth  overnight at 37 \u00b0C [K. pneumoniae formed viscous strings >5 mm length using a standard bacteriological loop.The string test was used for the identification of hypermucoviscous at 37 \u00b0C . The posK. pneumoniae isolates were determined by the Multilocus Sequence Typing (MLST) scheme of Pasteur Institute  using the previously published primers [K. pneumoniae isolates was performed using specific primers for the wzy gene associated with K serotypes K1, K2, K20, and K57 [wzi gene sequencing for identification of capsular types K23, K27, K28, K31, K47, K60, K62, and K64 [Sequence types (STs) of  primers ,51. The  and K57  and by w and K64 . BacteriblaSHV, blaCTX-M, blaTEM, blaOXA-48, blaKPC, blaVIM, blaIMP, and blaNDM, class 1 and 2 integrons, and porin protein gene ompK36, as well as 8 genes associated with K. pneumoniae virulence, rmpA (hypermucoid phenotype regulator), aer (aerobactin), kfu (ferric absorption system), uge , wabG (glucosyltransferase), fimH (fimbria type I), allS and allR , were detected by PCR using specific primers as was described previously [Beta-lactamase genes eviously .http://www.generunner.net/, accessed on 1 November 2021) and in silico analysis by insilico.ehu.eus [K. pneumoniae virulence genetic determinants  and antibacterial resistance genes  were detected by RT-PCR using qPCRmix-HS SYBR  and the CFX96 Real-Time PCR system  with the following program: 95\u00b0 for the 20s, 61\u00b0 for 20s, 72\u00b0 for 30 s, repeat 40 times. Bacterial thermolysates were used as DNA templates.Oligonucleotides for RT-PCR  were des.ehu.eus  was perfhttps://github.com/ncbi/pgap, accessed on 1 November 2021). Antimicrobial resistance genes, STs, plasmids, and restriction-modification systems were identified using the web resource of the Center for Genomic Epidemiology . Virulence genes, capsular type, genes conferring resistance to heavy metals, and efflux pumps were identified by the Institut Pasteur, Paris, France, BIGS database web-resource of .WGS was carried out on the Illumina MiSeq platform using Nextera DNA Library Preparation Kit  and MiSeq Reagent Kits v3 . The obtained single reads were collected into contigs using the SPAdes 3.9.0 software . De novo assembled genomes were annotated in the GenBank database  containing/not containing antimicrobials (100 mg/L AMP or 10 mg/L CRO) and incubated at 37 \u00b0C with agitation for 90 min. Each experiment was represented by three independent repeats. All following steps for RNA isolation were performed at 4 \u00b0C to limit RNase activity. Total RNA was isolated by phenol-chloroform extraction using kit RNA-extran , following manufacturer protocol. After isolation, RNA was treated with TURBO DNase  to remove traces of genomic DNA.proC, recA, and rpoD expression. Three technical replicates per each of the three biological samples were used for statistical validity. The relative transcript levels of antimicrobial resistance and virulence genes were calculated using the 2\u2212\u0394\u0394Ct method [www.graphpad.com accessed on 1 November 2021). Gene expression levels of each gene at present of AMP and CRO were compared to those in conditions without antimicrobials.One \u00b5g of isolated total RNA was used for cDNA synthesis with RevertAid RT Reverse Transcription Kit . qPCR was performed using qPCRmix-HS SYBR  and the CFX96 Real-Time PCR system  with the following program: 40 cycles of 20 s at 95 \u00b0C for denaturation, 20 s at 61 \u00b0C for annealing, 30 s at 72 \u00b0C for extension and SYBR Green detection. The melting curve analysis in the temperature range from 60 \u00b0C to 94 \u00b0C, with a fluorescence estimation step of 0.2 \u00b0C, was performed to confirm the specificity of the reaction. Relative quantification of the target gene expression was normalized with reference genes t method . A heat"}
+{"text": "Drosophila macrophages, also known as plasmatocytes or hemocytes, utilizes enhanced cortical F-actin levels stimulated by the Drosophila member of the fos proto oncogene transcription factor family . RNA sequencing analysis and live imaging show that Dfos enhances F-actin levels around the entire macrophage surface by increasing mRNA levels of the membrane spanning molecular scaffold tetraspanin TM4SF, and the actin cross-linking filamin Cheerio, which are themselves required for invasion. Both the filamin and the tetraspanin enhance the cortical activity of Rho1 and the formin Diaphanous and thus the assembly of cortical actin, which is a critical function since expressing a dominant active form of Diaphanous can rescue the Dfos macrophage invasion defect. In vivo imaging shows that Dfos enhances the efficiency of the initial phases of macrophage tissue entry. Genetic evidence argues that this Dfos-induced program in macrophages counteracts the constraint produced by the tension of surrounding tissues and buffers the properties of the macrophage nucleus from affecting tissue entry. We thus identify strengthening the cortical actin cytoskeleton through Dfos as a key process allowing efficient forward movement of an immune cell into surrounding tissues.The infiltration of immune cells into tissues underlies the establishment of tissue-resident macrophages and responses to infections and tumors. Yet the mechanisms immune cells utilize to negotiate tissue barriers in living organisms are not well understood, and a role for cortical actin has not been examined. Here, we find that the tissue invasion of  The infiltration of immune cells into tissue underlies the establishment of tissue-resident macrophages, and responses to infections and tumors, but how do they overcome tissue barriers? This study shows that macrophages upregulate the proto-oncogene Fos, increasing the density and crosslinking of cortical actin, thereby counteracting the tension of surrounding tissues and protecting the macrophage nucleus. The classical model of cell migration on a surface postulated in the 1980s by Abercrombie has been extended . Much ofMigration in 2D and 3D environments requires actin polymerization to power forward progress. The assembly of actin at the leading edge, when coupled to Integrin adhesion to anchor points in the surrounding extracellular matrix (ECM), can allow the front of the cell to progress . This anDrosophila macrophage migration into the embryonic germband (gb) to investigate mechanisms of immune cell tissue invasion. Macrophages, also called plasmatocytes or hemocytes, are the primary phagocytic cell in Drosophila and share striking similarities with vertebrate macrophages  (control), UAS-cher RNAi KK107451, UAS-TM4SF RNAi KK102206, UAS-Lam RNAi1 GD45636, UAS-Lam RNAi2 KK107419 lines were obtained from the Vienna Drosophila Resource Center (Austria).SF, USA) . Oregon Here, we list the lines used in each figure; we state first the name from FlyBase; in parentheses, the name used in the figure panels is provided.Oregon R. srpHemo-GAL4, UAS-GFP (control). srpHemo-Gal4, srpHemo-H2A::3xmCherry/P{CaryP}attP2 (control). srpHemo-GAL4, UAS-GFP; kay1(Dfos1). srpHemo-GAL4, UAS-GFP::nls/+ (control 1). srpHemo-GAL4, UAS-GFP/+; kay1(Dfos1). srpHemo-GAL4, UAS-GFP::nls/+; kay2(Dfos2). srpHemo-GAL4, UAS-GFP::nls/(UAS-Fra)2; kay2. 10XUAS-IVS-myr::GFP/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ (control 2 and control). UAS-DfosDN/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ (mac>DfosDN). srpHemo-Gal4, srpHemo-H2A::3xmCherry/ UAS GFP::nls (ctrl). srpHemo-Gal4, srpHemo-H2A::3xmCherry/UAS-fbz (mac>DfosDN). srpHemo-Gal4, srpHemo-H2A::3xmCherry /+ (ctrl). srpHemo-Gal4, srpHemo-H2A::3xmCherry/UAS-DfosDN (mac>DfosDN). UAS-GFP; srpHemo-Gal4, srpHemo-H2A::3xmCherry (ctrl). UAS-Dfos RNAi HMS00254/srpHemo-Gal4, srpHemo-H2A::3xmCherry (mac>DfosRNAi1). UAS-Dfos RNAi JF02804/srpHemo-Gal4, srpHemo-H2A::3xmCherry (mac>DfosRNAi2). srpHemo-GAL4, UAS-GFP::nls/+ or /(UAS-Fra)2 (mac>Dfos). UAS-GFP; UAS-Dfos RNAi HMS00254/ srpHemo-Gal4, srpHemo-H2A::3xmCherry (mac>DfosRNAi1+ GFP). UAS-GFP; UAS-Dfos RNAi JF02804/srpHemo-Gal4, srpHemo-H2A::3xmCherry (mac>DfosRNAi2+ GFP).srpHemo-H2A::3xmCherry/+ (control). srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ (3 movies) and Resille::GFP/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+  and Resille::GFP/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ (3 movies) and Resille::GFP/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/UAS-DfosDN . srpHemo-Gal4, srpHemo-H2A::3xmCherry/UAS-fbz (mac>DfosDN). srpHemo-GAL4, UAS-GFP.nls/+ (control). srpHemo-GAL4, UAS-GFP.nls/+; kay2(Dfos2).UAS-Dicer2;; srpHemo-Gal4, srpHemo-H2A::3xmCherry/w1118 (control). UAS-Dicer2; UAS-TM4SF RNAi KK10220/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ (mac>TM4SF RNAi). UAS-Dicer2; UAS-cher RNAi KK107451/+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ (mac>cher RNAi). UAS-Dicer2; UAS-cher RNAi KK107451/UAS-TM4SF RNAi KK102206; srpHemo-Gal4, srpHemo-H2A::3xmCherry/+ . srpHemo-GAL4, UAS-mCherry::nls/UAS-mCD8::GFP (control). srpHemo-GAL4, UAS-mCherry::nls/UAS-mCD8::GFP; UAS-fbz/+ (mac>DfosDN). srpHemo-GAL4,UAS-mCherry::nls/UAS-cheerio::FLAG; UAS-fbz/+ . srpHemo-GAL4,UAS-mCherry.nls/UAS-TM4SF; UAS-fbz/+ . srpHemo-GAL4, UAS-mCherry::nls/ UAS-TM4SF (mac>TM4SF). srpHemo-GAL4, UAS-mCherry::nls/UAS-cher (mac>cher). srpHemo-Gal4, srpHemo-3xmCherry/+ (control). srpHemo-Gal4, srpHemo-3xmCherry/UAS-fbz (mac>DfosDN).srpHemo-3xmCherry; kay1 (Dfos1) and srpHemo-3xmCherry; +. srpHemo-Gal4, srpHemo-moe::3xmCherry/+;UAS-mCD8::GFP/+(Control). srpHemo-Gal4, srpHemo-moe::3xmCherry/UAS-fbz (mac>DfosDN). w118. srpHemo-Gal4, srpHemo-moe::3xmCherry/w118 (Control). srpHemo-Gal4, srpHemo-moe::3xmCherry/UAS-cher RNAi KK107451 (mac>cher RNAi). srpHemo-Gal4, srpHemo-moe::3xmCherry/UAS-TM4SF RNAi KK102206 (mac>TM4SF RNAi). srpHemo-GAL4, UAS-mCherry.nls/UAS-mCD8::GFP (control). srpHemo-GAL4, UAS-mCherry.nls/UAS-Dia\u0394Dad::EGFP; UAS-fbz/+ . srpHemo-GAL4, UAS-mCherry.nls/UAS-mCD8::GFP; UAS-fbz/+ (mac>DfosDN). srpHemo-GAL4, UAS-mCherry.nls/ UAS-Dia\u0394Dad::EGFP (mac>diaCA). UAS-GFPnls; srpHemo-Gal4, srpHemo-H2A::3xmCherry. #2: UAS-GFPnls/srpHemo-Gal4, srpHemo-H2A::3xmCherry; Dfos1. #3: UAS-GFPnls/ srpHemo-Gal4, srpHemo-H2A::3xmCherry; Dfos2. #4: UAS-Dia\u0394Dad::EGFP/srpHemo-Gal4, srpHemo-H2A::3xmCherry; Dfos1. #5: UAS-Dia\u0394Dad::EGFP/srpHemo-Gal4, srpHemo-H2A::3xmCherry; Dfos2. UAS-Dicer2;; srpHemo-Gal4, srpHemo-H2A::3xmCherry/P{CaryP}attP40 (control). UAS-Dicer2;+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/ UAS-dia RNAi HM05027 (mac>dia RNAi1). UAS-Dicer2;+; srpHemo-Gal4, srpHemo-H2A::3xmCherry/UAS-dia RNAi HMS00308 (mac>dia RNAi2). UAS-dia::EGFP/+; UAS-nlacz/ srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato. UAS-dia::EGFP/+; UAS-Rho1N.19)/srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato (mac>Rho1DN). UAS-dia::EGFP/+; UAS-fbz/srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato (mac>DfosDN). UAS-dia::EGFP/+; UAS-cher RNAi KK107451/srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato (mac>cher RNAi). UAS-dia::EGFP/+; UAS-TM4SF RNAi KK102206/srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato (mac>TM4SF RNAi). UAS-diaRBD::GFP/+; srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato/UAS-nlacZ (control). UAS-diaRBD::GFP/+; srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato/UAS-Rho1N.19 (mac>Rho1DN). UAS-diaRBD::GFP/UAS-fbz; srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato/+ (mac>DfosDN). UAS-diaRBD::GFP/UAS-cher RNAi KK107451; srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato/+ (mac>cher RNAi). UAS-diaRBD::GFP/UAS-TM4SF RNAi KK102206; srpHemo-Gal4, 10XUAS-IVS-myr::tdTomato/+ (mac>TM4SF RNAi).srpHemo-Gal4 UAS-LifeActGFP UAS-RedStinger (control); srpHemo-Gal4 UAS-LifeActGFP UAS-RedStinger; UAS-DfosDN (mac>DfosDN). srpHemo-Gal4, UAS-CLIP::GFP, UAS-RedStinger (control). srpHemo-Gal4, UAS-CLIP::GFP, UAS-RedStinger; UAS-fbz (mac>DfosDN). srpHemo-GAL4, UAS-mCherry.nls/UAS-mCD8::GFP (control). srpHemo-GAL4, UAS-mCherry.nls/UAS-Lamin RNAi GD45636, KK107419 . srpHemo-GAL4, UAS-mCherry.nls/UAS-LaminC RNAi TRIP JF01406 (mac>LamC RNAi). srpHemo-GAL4, UAS-mCherry.nls/UAS-mCD8::GFP; UAS-fbz/+ (mac>DfosDN). srpHemo-GAL4, UAS-mCherry.nls/UAS-Lam RNAi ; UAS-fbz/+ . srpHemo-GAL4, UAS-mCherry.nls/UAS-LaminC RNAi TRIP JF01406; UAS-fbz/+ . e22c-Gal4,srpHemo-H2A::3xmCherry/+ (control). srpHemo-QF/ srpHemo-H2A::3xmCherry; QUAS-fbz/UAS-Rho1.N12 (mac<>DfosDN). e22c-Gal4, srpHemo-H2A::3xmCherry/srpHemo-QF; +/ UAS-Rho1.N12 (ecto>Rho1DN). srpHemo-QF/ e22c-Gal4, srpHemo-H2A::3xmCherry; UAS-Rho1.N12/QUAS-fbz . +;UAS-GFP::nls, srpHemo-GAL4 (control). +;UAS-GFP::Lamin, srpHemo-GAL4 (mac>Lam).UAS-fbz (UAS-Dfos DN) line [The fragment was amplified from genomic DNA of the published DN) line  using pry1M{vas-int.Dm}ZH-2A w*; M{3xP3-RFP.attP}ZH-51D, BL 24483), to produce second chromosome inserts. All male survivors were crossed to w; Sp/CyO; PrDr/TM3Ser virgins. Transformants were recognized by eye color and crossed again to w; Sp/CyO; PrDr/TM3Ser virgins to get rid of the X chromosomal integrase.The TM4SF open reading frame was amplified from the DGRC GH07902 cDNA clone , using primers acagcgGAATTCATGGCATTGCCGAAGAAAAT and acagcgTCTAGATTAAAAGCTAATCGTCTGTCATT. The PCR product and the pUASt-aTTB vector (DGRC plasmid #1419) were digested with EcoRI and XbaI, and ligated. After sequencing, the construct was injected into the landing site line,  against the C-terminal end of g. Then 10 \u03bcg of the cleared lysate were separated by SDS-PAGE using 4% to 15% Mini-PROTEAN TGX Precast Protein gels  and blotted onto a Amersham Protran Premium western blotting nitrocellulose membrane . The nitrocellulose membrane was blocked with Pierce Clear Milk blocking buffer  and incubated in blocking buffer with anti-mCherry  at 1:1,000, and anti-Profilin  [Cages were prefed on fresh yeast plates for 2 days. Late stage 11/early stage 12 embryos were handpicked using a Leica M205 fluorescent microscope on ice-cold apple juice plates. They were transferred to RIPA buffer  with a Halt Protease/Phosphatase inhibitor cocktail  and lysed. After a 30-minute incubation on ice, they were centrifuged 15 minutes at 4\u00b0C at 15,000#chi 1J)  at 1:50 Dfos2 for which the setting was 25\u00b0C.Embryos were dechorionated in 50% bleach for 5 minutes, washed with water, and mounted in halocarbon oil 27 (Sigma) on a 24 \u00d7 50 mm high precision coverslip  between 2 bridges (approximately 0.5 cm high) of coverslips glued on top of each other or mounted in halocarbon oil 27 (Sigma) between an 18 \u00d7 18 mm coverslip  and an oxygen permeable membrane (YSI). The embryo was imaged on an upright multiphoton microscope  equipped with a W Plan-Apochromat 40X/1.4 oil immersion objective (Olympus). GFP and mCherry were imaged at 860 nm and 1,100 nm excitation wavelengths, respectively, using a Ti-Sapphire femtosecond laser system (Coherent Chameleon Ultra) combined with optical parametric oscillator technology (Coherent Chameleon Compact OPO). Excitation intensity profiles were adjusted to tissue penetration depth and Z-sectioning for imaging was set at 1 \u03bcm for tracking. For long-term imaging, movies were acquired for 60 to 150 minutes with a frame rate of 25 to 45 seconds. A temperature control unit set to 29\u00b0C was utilized for all genotypes except Dfos RNAi, embryos with 70% gb retraction (Stage 13) were used for vnc counts. The pre-gb zone was defined based on embryo and yolk autofluorescence as an area on the yolk sac underneath the amnioserosa with borders defined posteriorly by the gb ectoderm and anteriorly by the head. Macrophages were visualized using confocal microscopy with a Z-stack step size of 2 \u03bcm, and macrophage numbers within the gb or the segments of the vnc were calculated in individual slices (and then aggregated) using the Cell Counter plugin in FIJI. Total macrophage numbers were obtained using Imaris (Bitplane) by detecting all the macrophage nuclei as spots.Autofluorescence of the embryo revealed the position of the gb for staging of fixed samples. Embryos with 40% (\u00b15%) gb retraction (Stage 12) were analyzed for macrophage numbers in the pre-gb, within the gb, along the vnc, and in the whole embryo. For the srpHemo-H2A::3xmCherry and the surrounding tissues with Resille::GFP, or with only macrophages labeled by srpHemo-H2A::3xmCherry, or srpHemo>GFP::nls were imaged, and 250 \u00d7 250 \u00d7 40 \u03bcm3 3D stacks were typically acquired with approximately 0.2 \u00d7 0.2 \u00d7 1 \u03bcm3 voxel size every 39 to 41 seconds for approximately 2 hours. For imaging macrophages on vnc, frames were acquired at every 40 to 43 seconds for 30 minutes after macrophages started spreading into abdominal segment 2  are its instantaneous velocities. Only trajectories with a minimal duration of 15 time frames were used. Calculated persistence values were averaged over all trajectories to obtain a persistence index (I) for the duration of measurement (with \u22121 being the lowest and 1 the maximum). From 3 to 6 embryos were recorded and analyzed for each genotype; numbers of control and perturbed embryos are equal in each pairwise comparison.Cell speed and persistence were calculated from nuclei positions using custom Python scripts as described elsewhere . Briefly2 in each embryo. Analyses were carried out using standard Fiji software. From 4 to 5 embryos were analyzed per genotype. Macrophages in the pre-gb or gb entry zones were analyzed.The junctional intensity of F-actin  was calculated using linescan analysis as previously described  with thesrpHemo-moe::3xmCherry reporter line [DfosDN, cher RNAi, or TM4SF RNAi. Embryos were collected for 5 hours 30 minutes at 29\u00b0C, dechorionated in 50% bleach for 5 minutes, rinsed thoroughly with water, and aligned laterally side by side under a stereomicroscope using a fluorescence lamp to check for the presence of mCherry. Aligned embryos were then mounted as described in the live imaging section above. To image Moe::3xmCherry, a Zeiss LSM800 inverted microscope was used with the following settings: Plan-APOCHROMAT 40x/1.4 Oil, DIC, WD = 0.13 objective, 1.5\u00d7 zoom, 1,025 \u00d7 1,025 pixel, speed 8, heating chamber set to 29\u00b0C, z-interval 1 \u03bcm. Laser settings were kept constant in all experiments. Images were acquired during macrophage invasion into the gb (St 12). Pseudo-coloring was conducted for the mCherry red channel. Each pixel in the image has a color ascribed to it via the fire \u201cLook Up Table\u201d translating the level of intensity of the mCherry channel into a defined amount of each color. The highest intensity of the image is represented as very bright yellow, and all other gray values are depicted as colors on the scale accordingly.To quantify cortical F-actin intensity in living embryos, a ter line  was crost test with Welch\u2019s correction for DfosDN and one way-ANOVA for cher RNAi and TM4SF RNAi.For quantification of Moe::3xmCherry intensity, an ROI was drawn in Fiji software around macrophages at the gb entry site in 20 z-stacks for each embryo. The area mean intensity was measured in all ROIs, and the average/embryo was calculated. To normalize fluorescence intensities per batch, the average intensity/embryo of all ROIs in each sample was divided by the arithmetic mean of the average intensity/embryo of all ROIs in the control per batch. The normalized average intensities/embryo were then compared to each other using a https://github.com/Axmasha/Image_analysis_scripts.Methanol-fixed St 11 embryos were mounted either after staining with GFP antibody (Dia::GFP) or without staining (DiaRBD::GFP) and imaged with a Zeiss Inverted LSM800, Plain-Apochromat 63X/1.4 Oil Objective at an XY-resolution of 0.1 \u03bcm and a Z-resolution of 1 \u03bcm . All macrophages within 40 \u03bcm of the gb were analyzed. For the quantification of the levels of DiaRBD or the complete Dia protein at the plasma membrane versus the cytoplasm, confocal images were processed using Fiji and MATLAB-R2017b (MathWorks). Individual focal planes were used to segment a profile corresponding to an 8-pixel wide line drawn across the single outer membrane of individual macrophages chosen such that the extracellular portion of the line extended into surrounding tissue or space and not another macrophage. The corresponding intensity profiles of the Myr::Tomato and Dia::GFP or DiaRBD::GFP channels were extracted in Fiji using a custom macro and analyzed further using a custom MATLAB script. The membrane region was defined by finding the maximal value in the Tomato intensity profile and centering a 0.8-\u03bcm interval around it. The background was calculated for each GFP profile as the mean intensity in the 2 \u03bcm outside the cell, flanking the membrane region, and substracted from the entire profile. The integrated Dia::GFP or DiaRBD::GFP intensity at the membrane was calculated within the 0.8-\u03bcm interval defined above. The integrated cytoplasmic Dia::GFP or DiaRBD::GFP level was calculated as the mean intensity of 2 \u03bcm of the GFP profile inside the cell flanking the membrane region. Image analysis scripts are publicly available at UAS-CLIP::GFP under the control of srpHemo-Gal4. Briefly, 3D-stacks with 1 \u03bcm Z resolution were acquired every 35 to 45 seconds for approximately 1 hour. As the strength of the GAL4 expression increased over time, laser power was adjusted during acquisition to reach the best possible quality of visualization. Images acquired from mutiphoton microscopy were initially processed with ImSpector software (LaVision Bio Tec) to compile channels from the imaging data.Laterally oriented embryos were used to measure the maximal length and width of macrophages expressing We started measuring from the time the cell body of the first macrophage fully appeared at the interface between the ectoderm and mesoderm and yolk sac until it had moved 30 \u03bcm along the ectoderm mesoderm interface. At each time frame, a line was drawn in Fiji along the longest dimension of the macrophage in the direction of its front-rear polarization axis, denoted the maximal cell length, and along the orthologonal longest dimension, which was considered maximal cell width. We did not observe long CLIP::GFP protrusions, but when a small protrusion was present, it was not included in the length measurement; within this gb region, the front of the first macrophage was clearly outlined with CLIP::GFP. The border between the first and second entering macrophages was drawn based on the uninterrupted intense line of CLIP::GFP at the base of the first macrophage; only cells with a clearly visible border were measured. The length-to-width ratio was quantified for each time frame, and a probability density function was plotted: 5 embryos were recorded for each genotype.srpHemo-Gal4 UAS-LifeAct::GFP were used to image macrophage actin live with a 3D-stack resolution of 1 \u03bcm. See above description of CLIP::GFP labeled macrophage imaging for laser power and image compilation. Laser power was also increased further in the DfosDN samples to enhance actin visualization. We measured the length of the filopodia-like protrusion of the first entering macrophage with Imaris software (Bitplane) from the time when the protrusion was inserted into the ectoderm, mesoderm, and yolk sac interface until the macrophage started to translocate its cell body into that location.Laterally oriented embryos expressing w; +; srpHemo-Gal4, srpHemo-3xmCherry/+ or w; +; srpHemo-Gal4, srpHemo-3xmCherry/ UAS-DfosDN genotypes were placed into plastic cages closed with apple juice plates with applied yeast to enhance egg laying. Collections were performed at 29\u00b0C for 1 hour, then kept at 29\u00b0C for additional 5 hours 15 minutes to reach stage 11 to early stage 12. Embryos were harvested for 2 days with 6 to 7 collections per day and stored meanwhile at +4\u00b0C to slow down development. Collected embryos were dissociated and the macrophages sorted as previously described [5 macrophages were sorted within 30 minutes.Adult flies of either escribed . About 15 macrophages. RNA sequencing was performed by the CSF facility of Vienna Biocenter according to standard procedures (https://www.vbcf.ac.at/facilities/next-generation-sequencing/) on 3 replicates. Briefly, the cDNA library was synthesized using QuantSeq 3\u2032 mRNA-seq Library Prep kit and sequenced on the Illumina HiSeq 2500 platform. The reads were mapped to the Drosophila melanogaster Ensembl BDGP6 reference genome with STAR (version 2.5.1b). The read counts for each gene were detected using HTSeq (version 0.5.4p3). Flybase annotation (r6.19) was used in both mapping and read counting. Counts were normalized to arbitrary units using the TMM normalization from edgeR package in R. Prior to statistical testing, the data were voom transformed, and then the differential expression between the sample groups was calculated with limma package in R. The functional analyses were done using the topGO and gage packages in R [Total RNA was isolated from FACS-sorted macrophages using Qiagen RNeasy Mini kit (Cat No. 74104). The quality and concentration of RNA was determined using Agilent 6000 Pico kit (Cat No. 5067\u20131513) on an Agilent 2100 Bioanalyzer: on average about 100 ng of total RNA was extracted from 1.5 \u00d7 10ges in R ,109. RNARNA isolation and qPCR was performed from bones of wild-type C57BL/6 mice and from bones and OS of H2-c-fosLTR as previously described with the primers in p-value <0.05 was considered statistically significant .Mouse experiments: Data are shown as mean \u00b1 SEM. One-way ANOVA followed by Tukey multiple comparisons posttest was applied to compare experimental groups. Statistical analysis was performed using GraphPad Prism 6.0 software. A Drosophila experiments: Statistical tests as well as the number of embryos, cells, tracks, or contacts assessed are listed in the figure legends. All statistical analyses were performed using GraphPad PRISM or R Studio, and significance was determined using a 95% confidence interval. No statistical method was used to predetermine sample size.Dfos mutant analyses in Representative images of Dfos antibody staining were analyzed per replicate per genotype and in situ hybridization are from experiments that were repeated 2 times with many embryos with reproducible results. S1 TableComparison of TF mRNA expression in macrophages at stages 11\u201312 and 13\u201316, based on data in . TFs exp(TIF)Click here for additional data file.S2 TableGenes are ordered according to the adjusted p-value from the RNA sequencing. Function is based on Flybase assignments . The mur(TIF)Click here for additional data file.S1 Raw imagessrpHemo-moe::3xmCherry expressing either CD8::GFP (ctrl) or DfosDN in macrophages. Rightmost western blot also contains a w- lane. Top row shows blots probed with an mCherry antibody, bottom row the same blots probed with a profilin antibody as a loading control. Cropped versions of the blots are shown in Three original uncropped western blots of St 11 embryo extracts from (PDF)Click here for additional data file.S1 DataThis Excel file contains the raw data of the quantification of embryo macrophage counts and linescan analyses along with movie outputs. Each tab in the file names the figure panel whose graph is based on the data shown in that chart.(XLSX)Click here for additional data file.S2 DatasrpHemo-3xmCherry control embryos and those expressing DfosDN in macrophages. The mean from 3 samples is shown for each genotype, organized by Flybase IDs (Fbgn), along with statistical analyses.Compendium of the RNA sequencing data obtained from FACSed macrophages from Stages 11\u201312 (XLSB)Click here for additional data file.S1 Fig2Dfos mutant embryos compared to the control (p = 0.37) SD: 6, 7. (C) The total number of macrophages (see schematic at left) was not altered from that in the control embryos expressing DfosDN in macrophages (p = 0.12). SD: 60, 120.  The number of macrophages (green) along the vnc (outlined by black dotted line in the schematic on the left) shows no significant difference between the control and (D) macrophages that express DfosDN or (E) either of 2 RNAi lines against Dfos. (D) DfosDN p = 0.88, 0.99, >0.99. 1Dfos RNAi (TRiP HMS00254) p = 0.21, 0.06, 0.11, 0.072, 0.033, 0.30, 0.56. 2Dfos RNAi (TRiP JF02804) p = 0.34, 0.15, 0.83, 0.27, 0.47, 1.0, 0.45. (D) SD: Ctrl 3, 3, 3, 0.8; DfosDN 6, 3, 0.7. (E) SD: Ctrl 6, 3, 3, 3, 2, 0.3; 1Dfos RNAi 6, 3, 3, 3, 2, 2, 0.3; 2Dfos RNAi 6, 2, 3, 2, 3, 1, 0.4.  Macrophage numbers in the pre-gb (see schematic at left) are increased compared to the control for lines expressing (F) DfosDN or (G) one of 2 different UAS-Dfos RNAi constructs in macrophages under srpHemo-GAL4 control. (F) p = 0.04, SD: 19, 29. (G) 1Dfos RNAi p < 0.0009, 2Dfos RNAi p < 0.0001. SD: 12, 9, 14. (H) Macrophage numbers in the gb are not significantly altered compared to the control upon overexpression of Dfos in macrophages (p = 0.14). SD: 22, 14. (I) Macrophage numbers in the gb for lines expressing one of 2 different UAS-Dfos RNAi constructs in macrophages under srpHemo-GAL4 control and lines, which additionally express UAS-GFP. Control vs. 1mac>Dfos RNAi (TRiP HMS00254) or Control vs. 2mac>Dfos RNAi (TRiP JF02804), p < 0.0001. 1mac>Dfos RNAi vs. 1 + GFPmac>Dfos RNAi or 2mac>Dfos RNAi vs. 2 + GFPmac>Dfos RNAi, p > 0.99. SD: 33, 47, 34. The effect of each Dfos RNAi was eliminated upon simultaneous expression of another UAS construct. Macrophages are labeled using either srpHemo-Gal4 driving UAS-GFP or srpHemo-H2A::3xmCherry. \u201cmac>\u201d indicates srpHemo-GAL4 driver expressing UAS constructs specifically in macrophages. Histograms show mean \u00b1 SEM ***p < 0.005, **p < 0.01, *p < 0.05. Unpaired t test was used for statistics, except for G, I, which used one-way ANOVA. The number of embryos analyzed for that genotype is shown within each column in the graphs. In D, n = 6 embryos for the control and n = 9 for Dfos DN. In E, n = 9 embryos for control, 15 and 11 for Dfos RNAis. Scale bar in A: 10 \u03bcm. The data underlying the graphs can be found in (A) Dfos protein (green) is detected with an antibody in macrophages (magenta) in embryos from the stages as indicated. (B-I) Quantification in mid St 12 embryos. (B) The number of macrophages (green) in the pre-gb zone (outlined by a black dotted line in the schematic on the left) showed no significant change in (TIF)Click here for additional data file.S2 Fig2Dfos macrophages entering the gb (outlined by the dashed line). Time in minutes shown in the top right corner of each image. (C) Quantification of macrophage speed shows a significant reduction in the speed of 2Dfos macrophages in the pre-gb zone and at gb entry, but none in the head. Regions analyzed indicated in left schematic. Speed in head: control = 2.59 \u03bcm/min, 2Dfos = 2.68 \u03bcm/min, p = 0.40; speed in pre-gb = 3.38 \u03bcm/min, 2Dfos = 2.47 \u03bcm/min, p = 2.38e-06; speed in gb entry: control = 2.35 \u03bcm/min, 2Dfos = 1.62 \u03bcm/min, p = 0.0003. Macrophages are labeled using srpHemo-H2A::3xmCherry. Histograms show mean \u00b1 SEM. ****p < 0.0001, ***p < 0.005, **p < 0.01, *p < 0.05. Unpaired t test was used for statistics. The number of analyzed macrophages for each genotype shown within each graph column. Tracks were obtained from movies of 3 embryos each for control and mac>DfosDN for pre-gb entry in A, 4 each for gb entry in A, 3 each for the vnc in A, 4 each of control and 4 2Dfos embryos for head and pre-gb in C, and 3 embryos each for gb entry in C. Scale bars: 10 \u03bcm. The data underlying the graphs can be found in (A) Quantification reveals that the directional persistence of macrophages expressing DfosDN (0.58) is unchanged (0.56) in the pre-gb area (p = 0.66) but decreased during gb entry (0.65) (0.72), p = 0.038 and along the vnc (0.54) compared to the control (0.61), p = 0.00026. Left schematic shows pre-gb area in yellow, gb entry outlined in solid line. Boxed area in right schematic shows analyzed area of vnc. (B) Movie stills showing wild-type and (TIF)Click here for additional data file.S3 FigDhc36C 0.02, CG14204 0.03, CG42402 0.04, CR43767 0.046, TM4SF 0.03, CG42260 0.0011, cher 0.046, GstT4 0.018, Xrp1 0.0011, Tspo 0.046, CG31337 0.046. Frl, DAAM, dia, capu all >0.99.  Quantification of the macrophage numbers in (D) the pre-gb zone and (E) along the vnc from embryos expressing RNAi against cher (KK 107451), or TM4SF in macrophages (KK 102206) driven by srpHemo-Gal4 shows no significant alteration. The number in the column in (D) corresponds to the number of embryos analyzed. Control vs. cher RNAi p = 0.33. Control vs. TM4SF RNAi p = 0.05. Control vs. cher/TM4SF RNAi p = 0.67. (D) SD: 20, 20, 19, 13. For (E), n = 13 embryos for control and n = 15 for each cher RNAi and TM4SF RNAi. Control vs. cher RNAi p = 0.97 for T1, p = 0.33 for T2, p = 0.88 for T3. Control vs. TM4SF RNAi p = 0.52 for T1, p = 0.76 for T2, p = 0.35 for T3. SD: ctrl 6.5, 5.4, 0.6; cher RNAi 5.0, 3.3, 0.8; TM4SF RNAi 4.4, 4.9, 1.9. (F-I) q-PCR analysis of mRNA extracted from the bones of mice that are wild type, tg for MHC c-fos, viral 3\u2032 UTR, and those in which c-fos transgenesis has led to an OS. Analysis of mRNA expression shows that (F) higher Fos levels in OS correlate with higher levels of (G) the glutathione S transferase Gstt3, and (H) the slit receptor Eva1c, but not (I) Tspo. Bone and OS RNA isolated from the same transgenic mouse, n = 4 mice per group, age 5 to 6 months. p-values = 0.86, 0.0028, 0.0013 in (F), 0.79, 0.0001, 0.0003 in (G), 1.0, 0.054, 0.049 in (H), 0.37, 0.33, 0.040 in (I). SD: 0.7, 0.6, 2.6 in (F); 0.2, 0.3, 1.1 in (G); 0.4, 0.2, 1.5 in (H); 0.1, 0.2, 0.2 in (I). Histograms show mean \u00b1 SEM ***p < 0.005, **p < 0.01, *p < 0.05. Unpaired t test or one-way ANOVA with Tukey post hoc were used for statistics of quantifications. Significance is based on adjusted p-values. The data underlying the graphs can be found in (A-C) Comparative mRNA expression levels as determined from RNA sequencing analysis of FACS-sorted wild-type macrophages and those expressing DfosDN, n = 3 biological replicates.  Genes down-regulated in macrophages expressing DfosDN are shown, separated into those with (A) strong and (B) moderate expression in wild-type macrophages. (C) Expression levels of Drosophila formin family genes are unchanged. Fold enrichment is normalized. p-values: (TIF)Click here for additional data file.S4 FigsrpHemo-moe::3xmCherry expressing either CD8::GFP (ctrl) or DfosDN in macrophages. Left western blot also contains w- lane. Original uncropped western blots can be found in 1Dfos or 2Dfos embryos completely rescued the macrophage gb invasion defect. p-values: Control vs. 1Dfos or vs. 2Dfos p = 0.0004 or p = 0.0055, respectively; Control vs. 1 mac>DiaCADfos or vs. 2 mac>DiaCADfos p > 0.999; 1Dfos vs. 1 mac>DiaCADfos p = 0.0005; 2Dfos vs. 2 mac>DiaCADfos p = 0.035. SD: 20, 23, 18, 19, 7.8.  There was no significant change in the number of macrophages in (C) the pre-gb zone or (D) along the vnc in embryos expressing either of 2 different RNAi lines against dia expressed in macrophages. Pre-gb: Control vs. 1dia RNAi p = 0.54, Control vs. 2dia RNAi p = 0.77. vnc: Control vs. 1dia RNAi p = 0.99, Control vs. 2dia RNAi p = 0.95. RNAi1 = TRiP HMS05027, RNAi2 = TRiP HMS00308. (C) SD: 9, 12, 13. (D) SD: Ctrl 5.2, 6.4, 2.5, 0.4; 1dia RNAi 5.6, 6.8, 1.7, 0.2; 2dia RNAi 5.1, 4.9, 2.1, 0.6.  Two further examples of line profiles used for the determination of the membrane-to-cytoplasmic ratios in cher RNAi, or TM4SF RNAi as shown in the schematic in E. Line length approximately 8 \u03bcm. Blue lines indicate mean GFP intensity on the membrane and in cytoplasm. Histograms show mean \u00b1 SEM ***p < 0.005, **p < 0.01, *p <0.05. One-way ANOVA with Tukey post hoc was used for statistics of quantification. The number in each column corresponds to the number of analyzed embryos. \u201cmac>\u201d indicates srpHemo-GAL4 driver expressing UAS constructs in macrophages. Macrophages are labeled using srpHemo-H2A::3xmCherry. The data underlying the graphs can be found in (A) Three western blots probed with an mCherry antibody of St 11 embryo extracts from (TIF)Click here for additional data file.S5 FigsrpHemo-Gal4 (\u201cmac>\u201d) driving UAS-LifeActGFP. White stars indicate the tip of each actin protrusion. Scale bar 5 \u03bcm. (B) Microtubules are labeled with srpHemo-Gal4 driving UAS-CLIP::GFP. Spatially matched stills of the first macrophage expressing DfosDN and control extending protrusions into the gb slightly before entering with the body of the cell. As DfosDN macrophages have a delay in entry, the stills from the DfosDN movie are from a later developmental time point than the control. (C) Quantification of macrophage maximum length and maximum width shows that DfosDN expressing macrophages are 23% longer and 12% thinner than wild-type macrophages inside the gb (indicated in schematic above by dashed box). Control vs. DfosDN maximum length p = 0.0005, SD: 3.4, 5.7; control vs. DfosDN maximum width p = 0.0025, SD: 1.3, 1.0. (D) Quantification of the maximum length and maximum width of macrophages in the pre-gb zone (indicated in schematic by dashed box) shows that macrophages expressing DfosDN are 9% shorter and 9% thinner than wild-type macrophages. Control vs. DfosDN maximum length p = 0.0095, SD: 2.2, 2.0; control vs. DfosDN maximum width p = 0.005, SD: 2.3, 1.9. (E) Overexpression of UAS-Lam in macrophages through srpHemo-Gal4 (mac>) causes no change in their number in the gb compared to the control. p = 0.65, SD: 15, 18. Histograms show mean \u00b1 SEM ***p < 0.005, **p < 0.01, *p < 0.05. Unpaired t test was used for statistics of quantification. The number of measurements per genotype is shown in each columns. The data underlying the graphs can be found in (A) Representative image showing actin protrusions of the first macrophage entering the gb in the control and in lines expressing DfosDN in macrophages. Actin was visualized by (TIF)Click here for additional data file.S6 FigProposed interactions of proteins at the cell cortex in wild-type macrophages during gb infiltration as shown in (TIF)Click here for additional data file.S1 MoviesrpHemo-H2A::3xmCherry are imaged while entering the gb in control embryos (left) and embryos in which macrophages express DfosDN (right). Time in minutes is indicated in the upper right corner. Scale bar: 10 \u03bcm. DfosDN, dominant negative version of Dfos; DN, dominant negative; gb, germband.Movies corresponding to stills shown in (AVI)Click here for additional data file.S2 MoviesrpHemo-Gal4 driving UAS-GFP::nls are imaged during their migration along the segments of the vnc in control embryos (left) and embryos in which DfosDN is expressed in macrophages (right). Time in minutes is indicated in the upper right corner. Scale bar: 10 \u03bcm. DfosDN, dominant negative version of Dfos; vnc, ventral nerve cord.Movies corresponding to stills shown in (AVI)Click here for additional data file.S3 MoviesrpHemo-Gal4 driving UAS-GFP::nls are imaged while entering the gb in control embryos (left) and 2Dfos mutant embryos (right). Time is indicated in minutes. Scale bar: 10 \u03bcm. gb, germband.Movies corresponding to stills shown in (AVI)Click here for additional data file.S4 MoviesrpHemo-Gal4 driving UAS-LifeAct::GFP is imaged during gb entry in control embryos (left) and embryos with macrophages expressing DfosDN (right). Note the extended protrusion of the DfosDN expressing macrophages. Time is indicated in minutes. Scale bar: 10 \u03bcm. DfosDN, dominant negative version of Dfos; gb, germband.Movies corresponding to stills shown in (AVI)Click here for additional data file.S5 MoviesrpHemo-Gal4 driving UAS-CLIP::GFP. They are imaged during gb entry in control embryos (left) and embryos with macrophages expressing DfosDN (right). Note the extended shape of the DfosDN expressing macrophages. Time is indicated in minutes. Scale bar: 10 \u03bcm. DfosDN, dominant negative version of Dfos; gb, germband.Movies corresponding to stills shown in (AVI)Click here for additional data file."}
+{"text": "Amycolatopsis sp. CA-230715, a potentially interesting producer of natural products. The genome of CA-230715 was sequenced using PacBio, Illumina, and Nanopore technologies. It consists of a circular 10,363,158-nucleotide (nt) chromosome and a circular 12,080-nt plasmid.We report the sequencing, assembly, and annotation of the genome of  Amycolatopsis is a recognized source of secondary metabolites . The original colony was isolated from a serial dilution of a soil suspension plated onto HANOB medium  after incubation for 5\u2009weeks at 28\u00b0C/70% relative humidity. For DNA isolation, the strain was grown in liquid yeast extract-malt extract (YEME) medium . Subread generation and adapter removal were performed using SMRT Analysis v2.3 software. A KAPA HyperPlus library was sequenced on an Illumina MiSeq instrument , yielding 4,477,879 read clusters (2\u2009\u00d7\u2009150\u2009nt), totaling 1,273,625,221\u2009nt. Nanopore data were generated on a MinION device using the SQK-RBK004 kit and a FLO-MIN106D R9.4 Rev-D flow cell  . Default software parameters were used except where otherwise noted. The Illumina reads were adapter and quality trimmed using AdapterRemoval2 v2.1.7  similarity to Amycolatopsis nigrescens CSC17Ta-90, and GTDB-tk v1.5.1, R202 , SRR12367305 (PacBio), and SRR12367307 (Nanopore). The GenBank accession numbers are CP059997.1 (chromosome) and CP059998.1 (plasmid).All data are available under BioProject accession number"}
+{"text": "Legionella pneumophila secretes toxins into the host cell that induce the non-canonical processing and activation of the ER stress sensor and transcription factor ATF6 via a mechanism that is distinct from the canonical pathway activated by unfolded protein buildup. Legionella pneumophila (L.p.) secretes \u223c330 effector proteins into the host cell to sculpt an ER-derived replicative niche. We previously reported five L.p. effectors that inhibit IRE1, a key sensor of the homeostatic unfolded protein response (UPR) pathway. In this study, we discovered a subset of L.p. toxins that selectively activate the UPR sensor ATF6, resulting in its cleavage, nuclear translocation, and target gene transcription. In a deviation from the conventional model, this L.p.\u2013dependent activation of ATF6 does not require its transport to the Golgi or its cleavage by the S1P/S2P proteases. We believe that our findings highlight the unique regulatory control that L.p. exerts upon the three UPR sensors and expand the repertoire of bacterial proteins that selectively perturb host homeostatic pathways.The intracellular bacterial pathogen  Legionella pneumophila (L.p.), expertly manipulate host cell function to create their replicative niche. L.p. uses the specialized Dot/Icm Type IVB secretion system (T4SS) to translocate roughly 300 bacterial effector proteins into the host cytosol  with the host endosomal machinery. Instead these effectors facilitate the remodeling of the LCV into a compartment that supports pathogen replication  to allow for the antibody-mediated opsonization of L.p. Surprisingly, and in contrast to DTT treatment, infecting these cells with wild type L.p. (WT L.p.) resulted in the near complete processing of endogenous ATF6-FL into two distinct fragments\u2014a major fragment of \u223c75 kD that we designate as ATF6-P  did not affect ATF6-FL protein levels  is processed upon cleavage into an \u223c55 kD N-terminal fragment (ATF6-N) that translocates to the nucleus and activates transcription . Both threatment . ImportaL.p. infection has been shown to influence the IRE-1 branch of the UPR . HEK293-Fc\u03b3R cells were pre-treated for 3 h with the proteasome inhibitor MG-132 or control media. ER stress induction using DTT led to rapid ATF6 processing after 1 h, whereas prolonged exposure to DTT for 3 h resulted in recovery of ATF6 signal due to autoregulatory feedback from UPR induction . In contnfection . When cet impact . Previout impact . It was epletion . To testwith CHX . Similared cells . Howevercontrols . As L.p.ynthesis , we consynthesis . Consist WT L.p. . Taken tL.p. induced ATF6-FL processing into the ATF6-P (\u223c75 kD) and ATF6-LMW (\u223c30 kD) fragments affected its distal function as a nuclear transcription factor. To address this question, we used the N-terminal tagged GFP fusion protein of ATF6-FL (GFP-ATF6-FL)  domain, whereas GFP-ATF6 1-331, GFP-ATF6 1-343, and GFP-ATF6 1-355 possessed partial bZIP domains (WT L.p. (WT L.p. but not \u0394dotA L.p. (WT L.p. migrated on a reducing SDS\u2013PAGE gel in a manner similar to the GFP-ATF6 1-331 truncation mutant (L.p. 5 h lane at high exposure). Significantly, an in vitro transcription and translation product of the DNA encoding the ATF6 1-331 protein migrated as a sharp band at a MW of \u223c30 kD on a reducing SDS\u2013PAGE and was detected by an antibody raised against ATF6   (Chen etATF6-FL) . During ATF6-FL)  Indeed, ATF6-FL) . After 1ATF6-FL) . We thene JF-644 . Analyseme point . Strikinreatment . These r domains . We coll and GalT-RFP (red) and stained with Hoechst solution (blue). DTT was added to cells at t = 0. Still frame images were taken every 5 min over 2 h 30 min. Video 2Halo-tag WT-L.p. Infected cells were imaged directly after spin-fection with no more than 30 min elapsed time between centrifugation and image acquisition at t = 0. Still frame images were taken every 10 min over 12 h. Download videoTime-lapse wide-field microscopy movie of HeLa-Fc\u03b3RII cells expressing GFP-ATF6 (green) and stained with Hoechst solution (blue). Cells were infected with JF-646 stained L.p. infection, we monitored the mRNA levels of ATF6 target genes by quantitative real time PCR (qRT-PCR), including UPR regulator/ER chaperone BiP (HSPA5), in HEK293-Fc\u03b3R cells. As expected, UPR induction with DTT increased expression of ER quality control genes BiP and HERPUD1 by greater than fivefold in comparison to control (DMSO-treated) cells  cells . Interespression . To gainreatment . When thL.p. infection was dependent on the ATF6-LMW. As the ATF6-LMW resembled the sequence architecture of the ATF6 1-331 mutant protein .We then undertook two orthogonal approaches to determine if the transcriptional program induced during tein see , we firstein see  Fig 3D)L.p. infe3T cells . To compficiency . Indeed,L.p. demonstrate the activation of the ATF6 pathway during L.p. infection.In sum, the gene expression changes in concomitance with ATF6 processing induced by L.p. infection, ATF6-FL is processed into a ATF6-P fragment and a ATF6-LMW fragment that retains transcriptional activity  During activity  and 3A\u2013Epparatus . It is wpathways . Howeverhe Golgi . Treatmehe Golgi  and S3A,A levels  and S3B.A levels  and S3A pression  and S3B.93 cells , along wmRNA see . In macrragments . Yet sur and on both the ATF6 S1P and S2P cleavage sites   that have both overlapping and distinct specificities. Pre-treatment of cells with this protease inhibitor cocktail did not affect the loss of ATF6-FL activated by WT L.p. , Legionella wadsworthii (L. wad), and Legionella longbeachae (L. lon)\u2014were tested in addition to L. pneumophila strains\u2014Philadelphia str. (WT L.p. Phila or \u0394dotA L.p. Phila), Paris str. (L.p. Paris), Lens str. (L.p. Lens), and Serogroup 6 str. (L.p. SG6). The Legionella species and strains were used to infect HEK293-Fc\u03b3R and RAW264.7 macrophage cells and endogenous ATF6-FL levels were monitored by immunoblotting. Whereas most of the species and strains tested recapitulated the loss of ATF6-FL as seen with wild type L.p. (see above), infecting cells with either L. wadsworthii or the L. pneumophila Paris str. did not result in an efficient processing of ATF6-FL L.p. PhilL.p. effectors lpg0519 and lpg2131 for further characterization. We generated GFP tagged fusion constructs of lpg0159 and lpg2131 and transiently expressed them in U2OS and HEK293 Fc\u03b3R cells. Unfortunately, the ectopic expression of tagged Lpg2131 in cells resulted in toxicity and confounded our interpretation of the results obtained (data not shown). We thus excluded this effector from further analyses. In contrast, GFP-Lpg0519, when expressed in U2OS cells, was relatively non-toxic and localized to the ER as observed by colocalization with the ER marker mCherry-KDEL  ATF6-FL is processed into two fragments, a higher MW ATF6-P fragment (\u223c75 kD) and a lower MW ATF6-LMW fragment (\u223c30 kD)  . Analysi(\u223c30 kD)  and 2. I(\u223c30 kD)  and S2B;(\u223c30 kD) ; and (3) fashion . These fL.p. infected cells  that activates the ERSE reporter in cells  that presumably evolved from the same species of L. pneumophila process ATF6 differently in their mammalian hosts due to a small but significant divergence in their effector repertoires  Lpg0519 folds and functions as an atypical protease; or (b) Lpg0519 promotes the activity of an ER localized host protease. In either scenario, the answers to these questions using molecular tools derived from L. pneumophila, will enhance our understanding of ATF6 regulation in physiology and pathology.The primary targets of the  amoebae . It is wan hosts . Examplean hosts . In this enzymes . Homologertoires . One of o the ER . Bioinfoo the ER , did not-FL loss . This suLegionella strains were gifts from Craig Roy\u2019s laboratory at Yale University. Legionella strains used in this study were routinely cultivated on Charcoal Yeast Extract agar. The \u0394dotA, \u0394sidC-sdcA, and IPTG-inducible Halo-expressing strains were derived from the parental Lp01 strain. The \u03942,3,4,6,7, \u03942,3,6,7, and \u03942,3,6 L.p. strains (7-translation L. pneumophila strain (L. pneumophila serogroup 1 Lp02 (L. pneumophila Paris B1 strain was purchased from ATCC (ATCC 700833). Chloramphenicol (10 \u03bcg/ml), IPTG (0.1 mM), and thymidine (100 \u03bcg/ml) were added to Charcoal Yeast Extract agar plates as needed. L.p. were harvested from 2-d heavy patches and used to infect cells.All  strains , and thea strain  was a gip 1 Lp02 . L. pneulpg0519 was amplified from L.p. (Lp01) genomic DNA and cloned into the pEGFP-C2 vector.GFP-ATF6, HA-ATF6, and HA-ATF6 1\u2013373 were kind gifts from Ron Prywes . The GFP-ATF6 truncation mutants were cloned from the GFP-ATF6 backbone and generated by Genscript Inc. For in vitro transcription and translation, the ATF6 1\u2013331 fragment was sub-cloned between HindIII and SalI sites on the pGEM3Z vector carrying an SP6 promoter. GFP-ATF6 S1P mutant and GFP-ATF6 S1P/S2P mutant constructs were generated by site directed mutagenesis using the Q5 Site-Directed Mutagenesis Kit (NEB). For the effector screen, Myc-tagged effector proteins were amplified from a plasmid effector library that was a kind gift from Russell Vance, University of California, Berkeley. To generate N-terminal GFP tagged Lpg0519, 2. RAW264.7 macrophages were cultured in Roswell Park Memorial Institute media (RPMI) (Corning) supplemented with 10% FBS at 37\u00b0C and 5% CO2. Cells were placed in poly-L-lysine\u2013treated plates and grown to 90% confluency. Drug treatments were performed at final concentration, 200 nM Tg (Enzo Life Sciences), 1 mM DTT , protease inhibitor cocktail (100 \u03bcM 4-(2-aminoethyl)benzenesulfonyl fluoride [AEBSF] [Sigma-Aldrich], 10 \u03bcM Tosyl phenylalanyl chloromethyl ketone [TPCK] [Sigma-Aldrich], 10 \u03bcM Calpain Inhibitor I [Sigma-Aldrich], 10 \u03bcM E 64 Protease inhibitor [EMD Millipore], 100 \u03bcM phenylmethylsulfonyl fluoride [PMSF] [Sigma-Aldrich]), 5 nM Bafilomycin A1 (Sigma-Aldrich), 1 \u03bcM PF-429242 (Sigma-Aldrich), 5 \u03bcM/20 \u03bcM MG-132 (Enzo Life Sciences), 25 \u03bcM Cycloheximide (Sigma-Aldrich), 10 \u03bcM Ceapin A7  supplemented with 10% FBS at 37\u00b0C and 5% COeapin A7 . Ceapin-For transient transfections, cells were grown to 70% confluency and transfected with 2 \u03bcg of plasmid for 60 and 35 mm dishes, or 1 \u03bcg per well for 24-well plates using JetPRIME (Polyplus-transfection) according to the manufacturer\u2019s instructions. Cells were incubated with transfection reagent for 4 h, then media replaced with fresh DMEM supplemented with 10% FBS. For siRNA transfections, cells were grown to 30\u201350% confluency and transfected using Oligofectamine (Thermo Fisher Scientific) according to the manufactures protocol. Cells were grown for 72 h before application of treatment conditions. The following siRNA oligos used were purchased from Sigma-Aldrich: SEL1L-TTAACTTGAACTCCTCTCCCATAGA, Scramble-GCATACTCAACTACTTCGCATACTT; ATF6- GAACAGGGCTCAAATTCTC, Scramble-GCTAGTGCACAAGTACCTA.Legionella polyclonal antibody  was used at 1:2,000 and incubated for 20 min at room temperature. Immediately after infection, cells were centrifuged for 5 min at 100g. After centrifugation, cells were left at 37\u00b0C for an additional 60 min. After 1 hour, cells were washed with 1\u00d7 PBS to remove extracellular bacteria. DMEM supplemented with 10% FBS or the same media supplemented with treatment reagent was replaced after the washes in PBS. Infected cells were harvested at the designated times.Cells were infected at a MOI of 100, 50, 25 or 5. If cells required opsonization, BiP (Human) forward- CATCACGCCGTCCTATGTCG, reverse- CGTCAAAGACCGTGTTCTCG; HERPUD1 (Human) forward- AACGGCATGTTTTGCATCTG, reverse- GGGGAAGAAAGGTTCCGAAG; SEL1L (Human) forward- AAACCAGCTTTGACCGCCAT, reverse- GTCATAGGTTGTAGCACACCAC; HYOU1 (Human) forward- GAGGAGGCGAGTCTGTTGG, reverse- GCACTCCAGGTTTGACAATGG; ATF6 (Human) forward- AGAGAAGCCTGTCACTGGTC, reverse- TAATCGACTGCTGCTTTGCC; DNAJB11 (human) forward- AACCTGAGCACCTTTTGCCT, reverse- GGTTCCGGTCGGGATGAAG, BiP (mouse) forward- ACTTGGGGACCACCTATTCCT, reverse- GTTGCCCTGATCGTTGGCTA; Dnajb11 (mouse) forward- TTGGAGGAACCCCTCGTCA, reverse- CTCTTGCCGACAGTTGCATTT; Hsp90B1 (mouse) forward- GTTCGTCAGAGCTGATGATGAA, reverse- GCGTTTAACCCATCCAACTGAAT; Atf6 (mouse) forward- TCGCCTTTTAGTCCGGTTCTT, reverse- GGCTCCATAGGTCTGACTCC; Actin (mouse) forward- GGCTGTATTCCCCTCCATCG, reverse- CCAGTTGGTAACAATGCCATGT.HEK-293 FC\u03b3 cell mRNA was harvested and isolated using Direct-zol RNA Miniprep Plus (Zymo Research) according to the manufacturer\u2019s protocol. cDNA synthesis was performed using QuantiTect Reverse Transcription Kit (QIAGEN) and cDNA reactions were primed with poly dT. Relative quantitative PCR was performed using iTaq Universal SYBR Green Supermix (Bio-Rad). GAPDH or HPRT mRNA (for human cells) or Actin mRNA (for RAW264.7) was used for normalization. Uninfected and untreated HEK-293 FC\u03b3RII and RAW264.7 macrophage cells were used as the endogenous control for each qRT-PCR analysis. The following qRT-PCR primers were used: Mammalian cells were lysed in radioimmunoprecipitation assay buffer (RIPA) buffer with the addition of protease (Roche cOmplete), and phosphatase inhibitors (GB Sciences). Protein levels of lysates were determined using the Bio-Rad DC/RC assay. Equal amounts of protein lysate were boiled with SDS load buffer, and equal amounts of protein were loaded. Immunoblotting was performed with the following antibodies: GAPDH , ATF6 rabbit polyclonal , ATF6 mouse monoclonal , \u03b2-Actin , \u03b1-Tubulin , SEL1L , ABCD3 , GFP tag , BiP , ATF4 .L.p. as needed. For ubiquitin recruitment assays, HEK293 Fc\u03b3R cells were infected with L.p. at MOI = 5. 1 h after infection, cells were washed twice with PBS to remove extracellular bacteria and incubated for 2 h more. For co-localization assays, Cos7 cells were co-transfected with pcDNA-Fc\u03b3RII and GFP-ATF6\u03b1, then infected with L.p. at MOI = 10 and infected for 1, 4, or 8 h. For 4- and 8-h time points, cells were washed with PBS after 1 h to remove extracellular bacteria. Coverslips were mounted with ProLong Diamond Antifade Mountant (Thermo Fisher Scientific) incubated at 37\u00b0C for 10 min, then imaged directly. For immunofluorescence, coverslips were washed with cold 1\u00d7 PBS, fixed with 4% paraformaldehyde in PBS for 10 min at room temperature, permeabilized in 0.1% saponin in PBS, and blocked in 3% BSA in PBS, and then incubated with the appropriate primary and secondary antibodies diluted in 3% BSA. Nuclei were stained with Hoechst 33342 dye for 10 min before mounting on microscope slides. Coverslips were imaged using an inverted Nikon Eclipse Ti-E spinning disk confocal microscope equipped with a Prime 95B 25mm CMOS camera (Photometrics) camera. Antibodies used were Ubiquitin , and Secondary Antibody, Alexa Fluor 546 or Alexa Fluor 488 (Thermo Fisher Scientific).Cells were plated on 12 mm glass coverslips in 24-well plates. After 24 h, cells were fixed, treated with drugs, or infected with WT- or \u0394dotA L.p. that were previously stained with HaloTag-Janelia Fluor 646 conjugates  to facilitate HaloTag-JF646 conjugation. After 15 min incubation with ligand in the dark, L. pneumophila were washed 1\u00d7 with water. Stained bacteria were resuspended in 2 ml of DMEM lacking phenol red (Gibco) and used for infection of cells. Imaging of cells took place in a controlled chamber maintaining 37\u00b0C with 5% CO2. A random selection of cells was imaged at 60\u00d7 magnification at 5- or 10-min intervals for 12 h using a Nikon Eclipse Ti2 microscope with a Nikon DS-Qi2 camera. Cells that died or lost focus over the time course were omitted from analysis.HeLa Fc\u03b3RII cells expressing GFP-ATF6 were plated on 35 mm poly-lysine\u2013coated imaging dishes (Cellvis). Cells were infected at MOI = 5 with njugates . For staIn vitro transcription and translation of luciferase and the ATF6 1\u2013331 fragment were performed using the TnT coupled Reticulocyte Lysate Systems kit from Promega. Briefly, 1 \u03bcg of DNA encoding for luciferase or the ATF6 1\u2013331 fragment downstream of an SP6 promoter were incubated with TnT reaction buffer, RNA polymerase, amino acids, RNAsin ribonuclease inhibitor (Promega), Magnesium actetate (0.25 mM) and rabbit reticulocyte lysate as a source of ribosomes and translation machinery in a test tube at 30\u00b0C for 90 min. 20% of the total reaction mixture was then boiled with Lammeli buffer and run on a 4\u201315% reducing SDS\u2013PAGE gel before immunoblotting with an ATF6 antibody .Legionella effectors as previously described (2 with 1 s exposure. Background signal was subtracted from untreated untransfected cells. Treatment conditions were normalized to control cells and represented as fold change in relative luminiscence.ONE-Glo Luciferase assay system was purchased from Promega. HEK-293T ERSE-Luciferase cells, provided as a gift from the Peter Walter laboratory at UC San Francisco and described previously , were seescribed  for 24 ht test was performed using three biological replicates. Statistical significance: *P-value < 0.05; **P-value<0.01, ***P-value <0.001. Image analysis was performed using ImageJ.GraphPad Prism 6 software was used for statistical analysis. Where statistical analysis was performed an unpaired"}
+{"text": "Rumex madaio Makino. The bacteriostatic rate of the extract was 75% against 23 species of common pathogenic bacteria. The extract was further purified using the preparative high-performance liquid chromatography (Prep-HPLC) technique, and five separated componential complexes (CC) were obtained. Among these, the CC 1 significantly increased cell surface hydrophobicity and membrane permeability and decreased membrane fluidity, which damaged cell structure integrity of Gram-positive and -negative pathogens tested. A total of 58 different compounds in the extract were identified using ultra-HPLC and mass spectrometry (UHPLC-MS) techniques. Comparative transcriptomic analyses revealed a number of differentially expressed genes and various changed metabolic pathways mediated by the CC1 action, such as down-regulated carbohydrate transport and/or utilization and energy metabolism in four pathogenic strains tested. Overall, the results in this study demonstrated that the CC1 from R. madaio Makino are promising candidates for antibacterial medicine and human health care products.Outbreaks and prevalence of infectious diseases worldwide are some of the major contributors to morbidity and morbidity in humans. Pharmacophageous plants are the best source for searching antibacterial compounds with low toxicity to humans. In this study, we identified, for the first time, antibacterial components and action modes of methanol-phase extract from such one edible herbaceous plant  R. madaio Makino is an edible, perennial and herbaceous plant that belongs to the Dicotyledoneae class, Polygonaceae family, and Rumex genus. According to the National Compilation of Chinese Herbal Medicine (1996 Edition), leaf and root tissues of R. madaio Makino can be used as medicine such as clearing heat and detoxification, removing blood stasis, and defecating and killing insects. Nevertheless, current studies on the antibacterial activity of R. madaio Makino are rare.China is one of the richest countries in biodiversity, with very high levels of plant endemism . PharmacR. madaio Makino were for the first time identified. The objectives of this study were: (1) to extract bioactive substances from R. madaio Makino using the methanol and chloroform extraction (MCE) method, and determine their inhibition activity against 23 species of pathogenic bacteria; (2) to purify the methanol-phase extract from R. madaio Makino by preparation high-performance liquid chromatography (Prep-HPLC) analysis, and identify bioactive compounds in componential complex 1 (CC 1) using an ultra-HPLC and mass spectrometry (UHPLC-MS) technique; (3) to determine cell surface hydrophobicity, cell membrane permeability, fluidity, and the damage of four representative pathogenic strains treated with the CC 1; (4) to decipher possible molecular mechanisms underlying antibacterial activity by comparative transcriptomic analysis. The results of this study meet the increasing need for novel antibacterial agent candidates against common pathogenic bacteria.In this study, antibacterial components and action modes of methanol-phase extract from R. madaio Makino were extracted using the MCE method. The results showed that the water loss rate of the plant material was 93.32%, and extraction rates of the methanol phase and chloroform phase were 32.10% and 29.60%, respectively. Antibacterial activity of the crude extracts against 23 species of pathogenic bacteria was determined, most of which are common foodborne pathogens, and the results are presented in R. madaio Makino showed a bacteriostatic rate of 39%, inhibiting 2 species of Gram-positive and 11 species of Gram-negative pathogens  showed weak or no antibacterial activity, indicating that bioactive compounds in the methanol-phase extract from V. alginolyticus ATCC17749 and V. parahaemolyticus ATCC17802; 128 \u03bcg/mL against B. cereus A1-1; and 256 \u03bcg/mL against V. parahaemolyticus B4-10.MIC values of the CC 1 were also determined, which was 64 \u03bcg/mL against B. cereus A1-1, the 2 h treatment by the CC 1 resulted in the bacterial cell surface shrinking seriously, the flagella breaking, and some contents leaking. After being treated for 4 h, cell surface shrinkage was intensified, and more cells were ruptured. After being treated for 6 h, the cell structure was seriously damaged, a large number of contents exuded, and only a few cells still maintained rod shape  analysis. As shown in od shape A. For th for 6 h C. These p < 0.05) when compared with the control groups (V. parahaemolyticus ATCC17802 (1.47-fold), V. parahaemolyticus B4-10 (1.62-fold) and B. cereus A1-1 (1.42-fold) after being treated with the CC1 for 2 h (p < 0.05), whereas a similar change was observed in the treatment group of V. alginolyticus ATCC17749 (1.48-fold) after being treated for 4 h. Moreover, the highest increase in cell surface hydrophobicity was observed in B. cereus A1-1 (3.75-fold) after being treated with the CC1 for 6 h . However, a significant decrease in membrane fluidity of these three strains was observed after the treatment for 4 h. Additionally, cell membrane fluidity significantly declined in B. cereus A1-1 (1.20-fold) treated with the CC 1 for 2 h, and sharply lost for 6 h (8.11-fold)  B. The chd-galactopyranoside (o-nitrophenyl)-\u03b2-d-galactopyranoside (ONPG) was used as a probe to monitor the inner cell membrane permeability of the four bacterial strains, and the results were illustrated in R. madaio Makino on inner cell membrane permeability was observed among the four treatment groups. For example, V. alginolyticus ATCC17749 did not change significantly in the inner cell membrane permeability after the treatment for 2 h (p > 0.05), whereas a significant increase was observed after being treated for 4 h (1.15-fold) and 6 h (1.18-fold), respectively (p < 0.05) (The o-nitrophenyl-\u03b2- < 0.05) .N-Phenyl-1-naphthylamine (NPN) was used as a probe to monitor the bacterial outer membrane permeability. As shown in p < 0.01). The highest increase was found in B. cereus A1-1 (6.06-fold) after being treated for 6 h, whereas an opposite pattern was observed in V. parahaemolyticus ATCC17802 (1.77-fold).p < 0.05), which raised with the increase in treatment time. Significant damage was observed in B. cereus A1-1 (2.95-fold) and V. parahaemolyticus B4-10 (2.21-fold) after being treated for 2 h, whereas a similar change was found in the other two strains treated for 4 h. Moreover, cell membrane damage of B. cereus A1-1 was the most severe among the four strains after being treated for 6 h (8.54-fold).As shown in R. madaio Makino significantly increased bacterial cell surface hydrophobicity and membrane permeability and decreased membrane fluidity of V. parahaemolyticus ATCC17802, V. parahaemolyticus B4-10, V. alginolyticus ATCC17749, and B. cereus A1-1, consistent with the observed bacterial surface structure by the TEM analysis. The damaged cell surface and membrane structure integrity were beneficial for the CC1 to penetrate bacterial cell envelope to target intracellular processes.Taken together, these results demonstrated that the CC 1 from 2O was subjected to UHPLC-MS analysis. As shown in R. madaio Makino.The obtained CC 1 resolved in HR. madaio Makino, we determined transcriptomes of the four bacterial strains treated for 6 h using Illumina RNA sequencing technology. A complete list of DEGs in the four strains was available in the NCBI SRA database  under the accession number PRJNA767551. To validate the transcriptome data, we examined 32 representative DEGs , and 78 genes were down-regulated (FC \u2264 0.5). Based on the comparative transcriptomic analyses, 11 significantly changed metabolic pathways were identified, including valine, leucine and isoleucine degradation; nitrogen, histidine, tryptophan, glyoxylate and dicarboxylate metabolisms; quorum sensing (QS); lysine degradation; fatty acid degradation; amino sugar and nucleotide sugar metabolism; ABC transporters; and mitogen-activated protein kinase (MAPK) signal pathway (Approximately 6.73% (316/4698) of  pathway .V. alginolyticus ATCC17749 (2.002- to 87.807-fold) (p < 0.05) (p < 0.05); six DEGs encoding key enzymes in the histidine metabolism were also significantly up-regulated (2.001- to 3.187-fold) (p < 0.05); similarly, in the tryptophan metabolism, expression of three DEGs were significantly enhanced (2.123- to 5.154-fold) (p < 0.05); additionally, in the lysine degradation, expression of a transcriptional regulator (N646_3623) and an arginine/lysine/ornithine decarboxylase (N646_1979) were significantly up-regulated (2.972- to 3.332-fold) (p < 0.05). These four pathways are related to amino acid degradation metabolisms.Remarkably, approximately 60 DEGs involved in 10 changed metabolic pathways were significantly up-regulated in  < 0.05) . For exap < 0.05), in which, specifically, one DEG encoding a hydroxylamine reductase (N646_0236) was greatly enhanced to express (87.807-fold).Meanwhile, eight DEGs in the nitrogen metabolism were also significantly up-regulated (2.193- to 87.807-fold)  (p < 0.05)  . ABC trap < 0.05)  ( < 0.05) , which lV. parahaemolyticus ATCC17802 genes were expressed differently in the experimental group compared with the control group. Among these, 128 genes showed higher transcription levels (FC \u2265 2.0), and 789 genes were down-regulated (FC \u2264 0.5). Comparative transcriptome analyses revealed 20 significantly changed metabolic pathways, including methane, nitrogen, glycerolipid, propanoate, sulfur, starch and sucrose, taurine and hypotaurine, phosphonate and phosphinate, and biotin metabolisms; glucagon, and hypoxia inducible factor-1 (HIF-1) signaling pathway; benzoate and ethylbenzene degradation; glycolysis/gluconeogenesis; flagellar assembly; apoptosis; bacterial chemotaxis; cationic antimicrobial peptide (CAMP) resistance; necroptosis, and RNA transport  of ransport .p < 0.05) (VP_RS18295), the other seven DEGs were significantly down-regulation (0.087- to 0.433-fold) (p < 0.05); in the propanoate metabolic pathway, express of four DEGs were significantly depressed (0.051- to 0.240-fold) (p < 0.05); in the starch and sucrose metabolisms, except for a 4-alpha-glucono transfer (VP_RS22910), the other five DEGs were significantly down-regulated (0.206- to 0.499-fold) (p < 0.05). These three metabolic pathways were related to carbohydrate metabolisms. Their overall down-regulation trend indicated inactive carbon source transportation and/or utilization, which likely resulted in insufficient energy supply.Notably, approximately 77 DEGs involved in 12 changed metabolic pathways were significantly down-regulated (0.05- to 0.491-fold) ( < 0.05) . For exaV. parahaemolyticus ATCC17802 were also significantly inhibited (p < 0.05). For example, the DEG encoding a pyruvate dehydrogenase complex dihydrolipoyllysine-residue acetyltransferase (VP_RS12210) was significantly down-regulated (0.331-fold), which connects glycolysis with tricarboxylic acid cycle (TCA) and plays a key role in glucose metabolism [Approximately 44 DEGs involved in six energy metabolism pathways in tabolism . The dowtabolism , which cV. parahaemolyticus ATCC17802 (0.055- to 0.49-fold) (p < 0.05), which indicated the depressed flagellum assembly that led to inactive motility of V. parahaemolyticus ATCC17802. The 17 down-regulated DEGs in the bacterial chemotaxis [p < 0.05) provided indirect evidence for this result.The bacterial flagellum is a complex mobility machine with a diversity of roles in pathogenesis, including attachment, colonization, invasion, maintenance and post-infection dispersal in the host ,22. In temotaxis  (0.101- p < 0.05). T3SS enables pathogenic bacteria to directly inject effector proteins into host cells, facilitating bacterial colonization in the host [V. parahaemolyticus ATCC17802 was significantly reduced after being treated with the CC 1 from R. madaio Makino.Interestingly, 23 DEGs encoding type III secretory system (T3SS) components were also significantly down-regulated (0.055- to 0.490 -fold) (the host . This reVP_RS00200), a thiol: disulfide interchange protein DsbA/DsbL (VP_RS21260), an ATP-binding cassette domain-containing protein (VP_RS05670), a multidrug efflux RND transporter periplasmic adaptor subunit VmeC (VP_RS00205), and a phosphoethanolamine-lipid A transferase (VP_RS21300)  resistance system, five DEGs were significantly inhibited (0.120- to 0.489-fold), including a multidrug efflux RND transporter permease subunit VmeD (RS21300) . These rVP_RS14060) and an envelope stress sensor histidine kinase CpxA (VP_RS14065) , e.g., a response regulator (RS14065) .V. parahaemolyticus B4-10 genes were expressed differently in the experimental group when compared with the control group. Among these genes, 204 showed higher transcription levels (FC \u2265 2.0), and 579 genes were down-regulated (FC \u2264 0.5). Based on the comparative transcriptome analysis, five significantly changed metabolic pathways were identified, including styrene degradation, nitrogen metabolism, QS, folate biosynthesis, and histidine metabolism (Approximately 16.75% (783/4674) of tabolism .V. alginolyticus ATCC17749, the expression of 10 DEGs in the nitrogen metabolism were significantly up-regulated (2.129- to 107.754-fold) (p < 0.05) (VP_RS05780) was greatly up-regulated (107.754-fold). This enzyme can reduce hydroxylamine analogs such as methylhydroxylamine and hydroxyquinone as a scavenger of potentially toxic by-products of nitrate metabolism [V. parahaemolyticus B4-10 after being treated by the CC 1.Similar to  < 0.05) . Notablytabolism . MoreoveB. cereus A1-1 genes were expressed differently in the experimental group. Among these genes, 178 showed higher transcription levels (FC \u2265 2.0), and 542 genes were down-regulated (FC \u2264 0.5). The comparative transcriptome analysis revealed 17 significantly changed metabolic pathways, including flagellar assembly; bacterial chemotaxis; two-component system (TCS); thiamine and nitrogen metabolisms; ABC transporters; arginine biosynthesis; fatty acid degradation; alanine, aspartate and glutamate metabolism; riboflavin metabolism; HIF-1 signaling pathway; glycolysis/gluconeogenesis; butanoate, pyrimidine, and propanoate metabolisms; benzoate degradation; and inositol phosphate metabolism (Approximately 12.57% (720/5730) of tabolism .B. cereus A1-1 (3.325- to 150.780-fold) (p < 0.05) (BCN_RS16540) was also greatly enhanced to express in B. cereus A1-1 (150.780-fold).Similar to the other bacterial strains tested, expression of 12 DEGs involved in the nitrogen metabolism and riboflavin metabolism were significantly up-regulated in  < 0.05) . SpecifiB. cereus A1-1 (0.038- to 0.487-fold) (p < 0.05) (p < 0.05); 9 DEGs in bacterial chemotaxis were significantly down-regulated (0.063- to 0.474-fold); and expression of 33 DEGs in ABC transporters were significantly inhibited (0.051- to 0.487-fold).Conversely, 69 DEGs involved in the flagellar assembly, bacterial chemotaxis, ABC transporters, and TCS were significantly down-regulated at the transcription level in  < 0.05) , similarBCN_RS24725) was also significantly down-regulated (0.191-fold). These results indicated the inhibited signal transduction systems in B. cereus A1-1.Approximately eight DEGs in the TCSs were significantly down-regulated. TCSs are widespread regulatory systems that can help bacteria to control their cellular functions and respond to a diverse range of stimuli . In thisp < 0.05)  ( < 0.05) , which sVibrio strains were inoculated in media (pH 8.4\u20138.5) with 3.0% NaCl, while non-Vibrios in media (pH 7.0\u20137.2) with 1% NaCl [Bacterial strains and culture media used in this study are listed in  1% NaCl .R. madaio Makino was collected in Lishui City , Zhejiang Province, China in September of 2020. A 500 g of fresh leaf and stem tissues of R. madaio Makino was washed clean, dried at room temperature, and then freeze-dried using ALPHA 2-4 LD Plus Freeze Dryer  at \u221280 \u00b0C for 48 h. The freeze-dried material was crushed using FW-135 High-Speed Crusher  and passed through 300 mesh screen. Then, 10.0 g of the powder was mixed with 99-mL chloroform: methanol  at a solid to liquid ratio of 1.10 (m/v) for 5 h [2O  was then added, fully mixed, and then sonicated using Scientz IID ULtrasonic Cell Crusher  at the following parameters: power: 300 W; ultrasonic on time: 1 s; ultrasonic off time: 1 s; working time: 20 min; and probe size: 6 mm. The sonicated mixture was filtered through 20\u201325 \u03bcm membrane , and the filtration was collected for the secondary extraction. The methanol phase was separated from the chloroform phase and then individually evaporated, concentrated on pasting using Rotary Evaporator . for 5 h . A 60 mLR. madaio Makino was determined according to the method issued by Clinical and Laboratory Standards Institute (CLSI)  using Mueller-Hinton (M-H) agar (CM337) and Mueller-Hinton broth (M391) . Briefly, a 10 \u03bcL of crude extracts (500 \u03bcg/mL) was added onto each blank disc  on MH ager plates. The gentamicin disc  was used as a positive control, while the methanol-phase with water and chloroform-phase with ethanol was a negative control, respectively. The plates were incubated at 37 \u00b0C for 12 h. Bacteriostatic activity was evaluated by measuring diameters of bacteriostatic circles.Susceptibility of bacterial strains  to the e6 colony-forming unit (CFU)/mL), and then incubated at 37 \u00b0C for 12 h [Broth dilution testing (microdilution)  was used to determine MICs of the extracts. Briefly, a 100 \u03bcL/well of the extracts (1024 \u03bcg/mL) was serially diluted, mixed with 100 \u03bcL/well of Mueller-Hinton broth (CM337) and 10 \u03bcL/well of bacteria strain  were centrifuged at 12,000 rpm for 20 min. The supernatant was filtered through 0.22 \u00b5m membrane , and the filtration was collected for further analysis. Prep-HPLC was run using Waters 2707  linked with UPLC Sunfire C18 column   at the following parameters: column temperature, 40 \u00b0C; injection volume, 100 \u03bcL; and mobile phase of methanol (eluent A) and water (eluent B) at a flow rate of 4 mL/min . Photo-diode array (PDA) spectra were measured in the wavelength ranging from 200 to 600 nm.Aliquots (10 mg/mL) of freeze-dried samples resolved in H2O (v/v), and mobile phase B was acetonitrile ; column temperature: 40 \u00b0C; auto-sampler temperature: 4 \u00b0C; injection volume: 2 \u03bcL. Typical ion source parameters were: IonSpray voltage: +5500/\u22124500 V; curtain gas: 35 psi; temperature: 400 \u00b0C; ion source Gas 1:60 psi; ion source Gas 2: 60 psi; and declustering potential (DP): \u00b1100 V. The SCIEX Analyst Work Station Software (Version 1.6.3) was employed for multiple reaction monitoring (MRM) data acquisition and processing. In-house R program and database were applied for peak detection and annotation .The UHPLC\u2013MS analysis was carried out using EXIONLC System  by Shanghai Hoogen Biotech, Shanghai, China using the parameters as described previously . The mobR. madaio Makino was added in bacterial culture (5 mL) at middle logarithmic growth phase (mid-LQP), and incubated at 37 \u00b0C for 2 h, 4 h and 6 h, respectively. A 1.5 mL of the cell suspension were collected, washed, fixed, and observed using SU5000 transmission electron microscope  [Samples for TEM analysis were prepared according to the method described previously . Briefly\u00d730,000) .600 nm values of 0.55 to 0.60) and rotated for 1 min and then stood at room temperature for 30 min. The absorbance of the aqueous phase was measured at OD600 nm using BioTek Synergy 2 . To measure the membrane fluidity, a 200 \u03bcL/well of bacterial suspension was mixed with 2 \u03bcL of 10 mM 1,6-diphenyl-1,3,5-hexatriene (DPH) , and the change of fluorescence intensity of each well was measured at excitation light wavelength of 362 nm and emission light wavelength of 427 nm using BioTek Synergy 2 .Bacterial cell surface hydrophobicity and membrane fluidity were measured according to the methods by Krausova et al.  and KuhrCell membrane damage was examined according to the method described previously . BrieflyR. madaio Makino and then incubated at 37 \u00b0C for 2 h, 4 h and 6 h. Outer membrane permeability was measured according to the method described previously [Bacterial culture at the mid-LGS was mixed with 1 \u00d7 MIC concentration of the CC 1 from eviously . Brieflyeviously .415 nm using BioTek Synergy 2  every 30 min for 5 h, which was marked as OD1, while OD2 generated from the untreated bacterial suspension was used as a negative control [Inner membrane permeability was measured according to the method described previously . Briefly control .R. madaio Makino for 6 h. Total RNA was prepared using RNeasy Protect Bacteria Mini Kit  and QIAGEN RNeasy Mini Kit (QIAGEN). DNA was removed from the samples using RNase-Free DNase Set (QIAGEN). Three independently prepared RNA samples were used for each Illumina RNA-sequencing analysis. Illumina sequencing was conducted by Shanghai Majorbio Bio-pharm Technology Co. Ltd.  using Illumina HiSeq 2500 platform . High quality reads that passed the Illumina quality filters were used for sequence analyses [Bacterial culture at the mid-LGP was treated with 1 \u00d7 MIC concentration of the CC 1 from analyses .\u2212\u0394\u0394Ct method was used to calculate relative expression of genes. Oligonucleotide primers used for the RT-qPCR were synthesized by Sangon, Shanghai, China.Total RNA extraction, reverse transcription reactions, and relative quantitative PCR reactions were performed using the same kits and instrument according to the method described previously . The 16Shttp://deweylab.github.io/RSEM/, accessed on 17 October 2021). Genes with the criteria, fold-changes \u2265 2.0 or \u22640.5, and p-values < 0.05 relative to the control were defined as DEGs. These DEGs were used for gene set enrichment analysis (GSEA) against the Kyoto Encyclopedia of Genes and Genomes (KEGG) database . Significantly changed GSEA were identified when the enrichment test p-value fell below 0.05 [Expression of each gene was calculated using RNA-Seq by Expectation-Maximization . The methanol-phase extract was further purified using the Prep-HPLC technique, and five separated CCs were obtained. Among these, the CC 1 from R. madaio Makino significantly increased bacterial cell surface hydrophobicity and membrane permeability and decreased membrane fluidity of Gram-positive and Gram-negative pathogens, such as V. parahaemolyticus ATCC17802, V. parahaemolyticus B4-10, V. alginolyticus ATCC17749, and B. cereus A1-1. The damaged cell surface and membrane structure integrity facilitated the CC1 to penetrate bacterial cell envelope to target intracellular processes. A total of 58 different compounds in the extract were identified using UHPLC\u2013MS technique. Comparative transcriptomic analyses revealed a number of differentially expressed genes (DGEs) and various changed metabolic pathways mediated by the CC1 action, such as down-regulation of carbohydrate transport and/or utilization, and energy metabolism; upward regulation of amino acid and fatty acid degradation, and nitrogen metabolism; and inactive flagellar assembly and mobility in the four bacterial strains. Taken, the results in this study demonstrated that the CC1 from R. madaio Makino are promising candidates for antibacterial medicine and human health care products.In this study, we identified, for the first time, antibacterial components and action modes of methanol-phase extract from one edible herbaceous plant"}
+{"text": "Frailty, a clinical syndrome characterized by vulnerability to stressors resulting from multisystemic loss of physiological reserve, predicts future cognitive decline. However, frailty has also been proposed as a dementia risk factor, predicting future cognitive impairment. The study aim was to determine frailty in older veterans and its association with risk of dementia. Community-dwelling Veterans \u226550 years completed a mailed socio-demographic questionnaire and Self-Administered Gerocognitive Examination (SAGE), July 2019-May 2020. The information was complemented with EHR data. We calculated the CAIDE score, a validated tool predicting dementia (\u22656 points= high risk 20 years later) and the 31-item VA frailty index data . After adjusting for socio-demographic characteristics, smoking, alcohol/substance abuse, OSA and anticholinergic use, odds ratio (OR) and 95% CI were calculated using BLR to assess the cross-sectional association between frailty and dementia risk (CAIDE \u22656 points and MCI). The survey response rate was 19.75% . Participants mean age was 68.38 (SD=8.49) years, 57.50% (n=617) Caucasian, 69.34% (n=744) non-Hispanic, 95.81%  male, and 36.72% (n=394) frail. 11.84%(n=127) screened positive for MCI and 15.38% (n=165) for dementia. 689 (75.88%) veterans were at high risk for dementia of whom 426 (61.83%) were non-frail and 263 (38.17%) were frail. Frailty was cross-sectionally associated with higher risk for dementia in older Veterans, adjusted OR:1.45 (95%CI:1.016-2.070), p=.041. The mailed screening was a feasible and practical approach to screen for dementia risk. Early identification of patients with frailty can help in the implementation of interventions aimed at preventing or delaying dementia."}
+{"text": "Excessive sedentary behavior (SB) is related to deleterious health outcomes. Understanding the patterns and contexts in which SB accumulates can promote healthy aging. Daily sitting time and mean sitting bout duration (MBD) were measured by triaxial accelerometers. Participants self-reported how much time they spent sitting while: watching TV, reading, using the computer, driving, working, or taking phone calls. Data were compared across aging-related characteristics. Age-adjusted sitting time (minutes/day) for 5,838 diverse , older women (mean age 78.7\u00b16.7) were 577.2 for Hispanic women, 630.3 for Black women, and 632.0 for White women. Those in the lowest vs. highest physical function category had the longest MBD (16.1 vs. 11.7 minutes/bout). Watching television was the most common self-reported sedentary activity. The highest vs. lowest quartile of MBD spent, on average, 30.6 and 22.3 minutes/day watching television, respectively. This presentation will illuminate critical factors associated with sitting patterns in older adults."}
+{"text": "This study reports on the E. coli and 344 S. saprophyticus isolates were collected between 2019 and 2020 from 92 medical centers located in 25 countries. Most isolates (68%) tested were cultured from urine specimens collected from patients seen in ambulatory, emergency, family practice, and outpatient medical services. Bacterial identifications were confirmed by MALDI-TOF. Isolates were tested for susceptibility by CLSI methods at a central laboratory (JMI Laboratories). MIC results for oral antibiotics licensed for the treatment of uUTI and drug-resistant subsets were interpreted per CLSI guidelines. A total of 3,562 50/90, 2/2 mg/L) displayed good activity against 3,562 E. coli isolates, with 98.0% of all observed gepotidacin MICs \u22644 mg/L (Table). Susceptibility (S) rates for the other oral agents tested against these isolates were: amoxicillin-clavulanate (79.6% S), ampicillin (45.6% S), ciprofloxacin (72.5%S), fosfomycin (99.0% S), mecillinam (94.1%S), nitrofurantoin (97.3% S), and trimethoprim-sulfamethoxazole (68.2% S). When tested against the drug-resistant subsets, gepotidacin maintained similar MIC50/90 values (2/4 mg/L), except against isolates resistant to fosfomycin (2/8 mg/L). Against S. saprophyticus isolates, gepotidacin  inhibited all isolates at \u22640.25 mg/L. Most oral agents showed S results of >97% against S. saprophyticus isolates, except for penicillin (3.5%S).Gepotidacin  (Research Grant or Support)GlaxoSmithKline, LLC (Research Grant or Support)Melinta Therapeutics, LLC (Research Grant or Support)Nabriva Therapeutics (Research Grant or Support)Spero Therapeutics (Research Grant or Support) Deborah Butler, n/a, GlaxoSmithKline, LLC (Employee) Nicole Scangarella-Oman, MS, GlaxoSmithKline, LLC (Employee) Lindsey Paustian, BS (ASCP), GlaxoSmithKline, LLC (Research Grant or Support) Jennifer M. Streit, BS, GlaxoSmithKline, LLC (Research Grant or Support)Melinta Therapeutics, LLC (Research Grant or Support)Shionogi (Research Grant or Support)Spero Therapeutics (Research Grant or Support) Rodrigo E. Mendes, PhD, AbbVie (Research Grant or Support)AbbVie (formerly Allergan) (Research Grant or Support)Cipla Therapeutics (Research Grant or Support)Cipla USA Inc. (Research Grant or Support)ContraFect Corporation (Research Grant or Support)GlaxoSmithKline, LLC (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Melinta Therapeutics, LLC (Research Grant or Support)Nabriva Therapeutics (Research Grant or Support)Pfizer, Inc. (Research Grant or Support)Shionogi (Research Grant or Support)Spero Therapeutics (Research Grant or Support)"}
+{"text": "Here, we describe BIOVERA-Tree, a database on tree diversity, community composition, forest structure and functional traits collected in 120 forest plots, distributed along an extensive elevational gradient in Veracruz State, Mexico. BIOVERA-Tree includes information on forest structure from three levels of forest-use intensity, namely old-growth, degraded and secondary forest, replicated across eight elevations from sea-level to near the tree line at 3500 m and on size and location of 4549 tree individuals with a diameter at breast height \u2265 5 cm belonging to 216 species, 154 genera and 80 families. We also report measurements of eight functional traits, namely wood density for 143 species, maximum height for 216 species and leaf traits including: specific leaf area, lamina density, leaf thickness, chlorophyll content and leaf area for 148 species and leaf dry matter content for 145 species.BIOVERA-Tree is a new database comprising data collected in a rigorous sampling design along forest-use intensity and elevational gradients, contributing to our understanding of how interactive effects of forest-use intensity and elevation affect tree diversity, community composition and functional traits in tropical forests. Mountains are fascinating ecosystems and natural laboratories for evolutionary and ecological research as they encompass a wide variety of different climatic conditions over short distances . MountaiLand-use change and intensification are occurring at rapid rates and are strongly impacting mountain ecosystems . For ins> 5 cm and tree height (metres) of 4548 and 4549 individuals, respectively. BIOVERA-Tree also includes functional traits for the common tree species, with data for wood density of 143 species calculated, based on 483 individuals, maximun height for 216 species calculated, based on 4549 individuals and leaf traits for 148 species, specific leaf area (n = 3148 leaves), lamina density (n = 3194 leaves), chlorophyll content (n = 3280 leaves), leaf area (n = 3214 leaves), leaf thickness (n = 3299) and leaf dry matter content for 145 species (n = 3081 leaves).BIOVERA-Tree originated from the interdisciplinary research project BIOVERA, which aims at documenting and understanding biodiversity patterns along gradients of altitude, climate, soil and disturbance along an elevational transect at the Cofre de Perote in central Veracruz, Mexico . BIOVERAThe study area is located along an elevational gradient from sea level close to the Gulf of Mexico to near the tree line at 3545 m on the eastern slopes of the Cofre de Perote volcano, in the central part of the State of Veracruz, Mexico Fig. . This reWe selected eight sites along the elevational gradient, separated by about 500 m in elevation Fig. . At each> 5 cm and identified individual trees to the highest taxonomic resolution possible. For each individual, we measured its DBH (in cm) and tree height (in m), which was measured with a Leica laser  to close to the treeline at 3545 m elevation  along the eastern slopes of Cofre de Perote volcano (4282 m) in Veracruz State, Mexico . Individuals were identified to the species level by specialists , while some individuals could only be identified to the family or genus level or could not be identified. Vouchers of specimens were deposited at the Herbarium XAL of Instituto de Ecolog\u00eda, A.C. at Xalapa, Mexico.Tree diversity and community compositionThe database contains information of 216 tree species Data package title BIOVERA-Tree: community, functional traits and forest structure along forest-use intensity and elevational gradients in Veracruz, Mexico.6BIOVERA-Tree forest plots description13Location of the 120 plots along the elevational gradient at the eastern slopes of Cofre de Perote in Veracruz, Mexico. Available as Suppl. material BIOVERA-Tree scientific name3List of tree species along the elevational gradient and different levels of forest-use intensity. Available as Suppl. material BIOVERA-Tree community matrix2Tree community matrix composition along eight elevational sites and three different forest-use intensity levels of 216 tree species (n = 5 plots per forest-use intensity within elevation). The numbers within the matrix are the number of individuals. Available as Suppl. material BIOVERA-Tree forest structure5> 5 cm and tree height (metres) for 216 species along the elevational gradient and different levels of forest-use intensity. Available as Suppl. material Diameter at breast height (DBH) BIOVERA-Tree functional traits7Plant functional traits measured along the elevational gradient and different levels of forest-use intensity; including leaf traits, wood density and maximum height. Available as Suppl. material BIOVERA Tree metadata3metadataDefinition and categories according with Darwin Core, Functional Diversity thesaurus and this research.https://doi.org/10.5061/dryad.ngf1vhhvb.The data underpinning the analysis reported in this paper are deposited in the Dryad Data Repository at 1BCEFC74-C760-5C41-BC74-77C909F237EB10.3897/BDJ.9.e69560.suppl1Supplementary material 1BIOVERA-Tree forest plots descriptionData typeLocation DataFile: oo_549009.csvhttps://binary.pensoft.net/file/549009Mar\u00eda Leticia Monge-Gonz\u00e1lez, Patrick Weigelt, Nathaly Guerrero-Ram\u00edrez, Dylan Craven, Gonzalo Castillo-Campos, Thorsten Kr\u00f6mer, Holger Kreft.297517E1-0C9F-5503-B9D1-6B484B74C9E710.3897/BDJ.9.e69560.suppl2Supplementary material 2BIOVERA-Tree scientific nameData typeTaxonomyFile: oo_549044.csvhttps://binary.pensoft.net/file/549044Mar\u00eda Leticia Monge-Gonz\u00e1lez, Patrick Weigelt, Nathaly Guerrero-Ram\u00edrez, Dylan Craven, Gonzalo Castillo-Campos, Thorsten Kr\u00f6mer, Holger Kreft.42D688AD-8284-53B7-8D58-28F37D03F2EF10.3897/BDJ.9.e69560.suppl3Supplementary material 3BIOVERA-Tree community matrixData typeSpecies community dataBrief descriptionTree community matrix with abundancesFile: oo_549021.csvhttps://binary.pensoft.net/file/549021Mar\u00eda Leticia Monge-Gonz\u00e1lez, Patrick Weigelt, Nathaly Guerrero-Ram\u00edrez, Dylan Craven, Gonzalo Castillo-Campos, Thorsten Kr\u00f6mer, Holger Kreft.1B6E2702-97B7-5634-BDC4-A09F03A3709210.3897/BDJ.9.e69560.suppl4Supplementary material 4BIOVERA-Tree forest structureData typeForest structure, DBH, tree heightFile: oo_549014.csvhttps://binary.pensoft.net/file/549014Mar\u00eda Leticia Monge-Gonz\u00e1lez, Patrick Weigelt, Nathaly Guerrero-Ram\u00edrez, Dylan Craven, Gonzalo Castillo-Campos, Thorsten Kr\u00f6mer, Holger Kreft.8880BA0C-CC51-5419-83D8-EA722FAC054510.3897/BDJ.9.e69560.suppl5Supplementary material 5BIOVERA-Tree functional traitsData typeFunctional trait dataFile: oo_549015.csvhttps://binary.pensoft.net/file/549015Mar\u00eda Leticia Monge-Gonz\u00e1lez, Patrick Weigelt, Nathaly Guerrero-Ram\u00edrez, Dylan Craven, Gonzalo Castillo-Campos, Thorsten Kr\u00f6mer, Holger Kreft.BB3E33BB-DDFE-53F9-976F-4676697DF15D10.3897/BDJ.9.e69560.suppl6Supplementary material 6BIOVERA-Tree metadataData typemetadataBrief descriptionDefinition and categories according with Darwin Core, Functional Diversity thesaurus and this research.File: oo_549022.csvhttps://binary.pensoft.net/file/549022Mar\u00eda Leticia Monge-Gonz\u00e1lez, Patrick Weigelt, Nathaly Guerrero-Ram\u00edrez, Dylan Craven, Gonzalo Castillo-Campos, Thorsten Kr\u00f6mer, Holger Kreft."}
+{"text": "Patients (pts) with newly diagnosed acute myeloid leukemia (AML) undergoing induction chemotherapy are at increased risk for invasive fungal infections (IFI). Guidelines recommend posaconazole prophylaxis (ppx), but use is precluded by interactions and adverse effects. Micafungin (MCF) is an alternative, but data is limited by small prospective and retrospective studies. Primary objective: describe incidence of probable/proven IFI until neutrophil recovery (ANC \u2265 500 cells/\u00b5L) or 28 days after induction start date, whichever occurred first, in pts receiving MCF ppx. Secondary objective: describe incidence of clinical failure to MCF prophylaxis. Retrospective review (January 2017 to January 2020) of newly diagnosed AML adult pts undergoing 7 + 3 using idarubicin (7 + 3-ida), 7 + 3 using daunorubicin (7 + 3-dau), venetoclax/decitabine (VEN/DEC), or venetoclax/azacitadine (VEN/AZA) receiving MCF ppx for at least 7 days included. Diagnosis of IFI < 30 days prior to induction, liver function tests (LFT) 5x ULN at start of induction, or evidence of refractory disease after induction excluded. Probable/proven IFI defined by EORTC criteria. Clinical failure: changing to a different antifungal class for any reason until ANC recovery or 28 days after induction start date.Ninety-five pts included. Baseline characteristics: mean (\u00b1SD) age 57.8 (\u00b113.0) years; 53.6% males. 62% (59/95) 7 + 3-ida, 13.7% (13/95) 7 + 3-dau, 15.8% (15/95) VEN/DEC, 8.4% (8/95) VEN/AZA. Mean (\u00b1SD): 32.5% (\u00b126) blasts, WBC 13.2 (\u00b123.8), ANC 2.4 (\u00b14.6), ALC 1.9 (\u00b11.6), platelets 92.6 (\u00b1123.2). Incidence of probable IFI 2/95 (2.1%). No proven IFI cases identified. Clinical failure occurred in 37/95 (39%): 8 persistent febrile neutropenia, 29 due to suspected IFI. No MCF discontinuation due to adverse events. Our findings suggest that prophylactic MCF is safe and effective in pts with newly diagnosed AML undergoing induction chemotherapy. Outcomes were similar to those of prophylactic posaconazole studies, indicating MCF may be considered as an alternative when interactions and adverse effects preclude use of posaconazole. Our study was limited by small numbers, retrospective, single-center design. Future opportunities include prospective trials of prophylactic MCF in this setting. All Authors: No reported disclosures"}
+{"text": "Hospitalized patients with COVID-19 have created increased demands on health care infrastructure and resources. Bacterial and fungal infections have been reported and have increased the need for antimicrobial utilization. We performed a retrospective chart review to characterize bacterial infections and antibiotic utilization during the COVID-19 surge at our tertiary care center. All patients diagnosed with COVID-19 using SARS-CoV-2 PCR admitted to MedStar Georgetown University Hospital from 01Mar2020 through 31Aug2020 were included in the analysis. Data was collected on hospital-wide antimicrobial utilization [mean days of therapy per 1000-patient-days (DOT)] during the 6-month surge and was compared to antimicrobial utilization during a 6-month period that preceded the COVID-19 surge. Clinical and microbiological data and patient outcomes were also collected and analyzed.A total of 238 patients met eligibility criteria during the observation period, of which 25.6% (n = 61) developed a bacterial, fungal, or viral co-infection. Culture-positive bacterial complications were seen in 21.8% (n = 52) with 32.8% (n = 20) having a multidrug resistant organism (MDRO). There was a statistically significant difference between COVID-19 patients with co-infection and those without for intubation (p < 0.001), vasopressor use (p < 0.001), and renal replacement therapy (p = 0.001). COVID-19 patients with co-infections had a longer mean length of stay  and greater mortality  compared to those without a co-infection, respectively.Mean antimicrobial utilization for the entire hospital population was 790.6 DOT during the COVID surge compared to 928.7 DOT during a 6-month period preceding the COVID surge (p < 0.001). For all COVID-19 patients, antimicrobial utilization was 846.9 DOT; however, this increased to 1236.4 DOT for COVID-19 patients with co-infections.Table 1. DemographicsTable 2. Antimicrobial Utilization in COVID-19 PatientsAlthough hospital-wide antimicrobial utilization had decreased during the COVID surge, COVID-19 patients with co-infections demonstrated a disproportionate use of antimicrobial agents as well as ICU resources. As MDRO infections were relatively common, antimicrobial stewardship should be prioritized in the COVID-19 population.Lan Duong, Pharm.D., Astra Zeneca (Shareholder)Eli Lilly & Co. (Shareholder)Gilead Sciences, Inc. (Shareholder)Merck & Co. (Speaker\u2019s Bureau)Moderna, Inc. (Shareholder)Novavax, Inc. (Shareholder)Sarepta Therapeutics (Shareholder)Thermo Fisher Scientific (Shareholder) Princy N. Kumar, MD, AMGEN Eli Lilly (Grant/Research Support)Gilead GSK Merck & Co., Inc."}
+{"text": "The stromal elements of a malignant tumor can promote cancer progression and metastasis. The structure of the tumor stroma includes connective tissue elements, blood vessels, nerves, and extracellular matrix (ECM). Some of the cellular elements of the tumor stroma are cancer-associated f ibroblasts (CAFs). The origin and function of CAFs have been actively studied over the past thirty years. CAFs produce collagen, the main scaffold protein of the extracellular matrix. Collagen in the tumor stroma stimulates f ibrosis, enhances the rigidity of tumor tissue, and disrupts the transmission of proliferation and differentiation signaling pathways. CAFs control tumor angiogenesis, cell motility, tumor immunogenic properties, and the development of resistance to chemo- and immunotherapy. As a result of metabolic adaptation of rapidly growing tumor tissue to the nutrients and oxygen deprivation, the main type of energy production in cells changes from oxidative phosphorylation to anaerobic glycolysis. These changes lead to sequential molecular alterations, including the induction of specif ied transcriptional factors that result in the CAFs activation. The molecular phenotype of activated CAFs is similar to f ibroblasts activated during inf lammation. In activated CAFs, alpha-smooth muscle actin (\u03b1-SMA) is synthetized de novo and various proteases and f ibronectin are produced. Since CAFs are found in all types of carcinomas, these cells are potential targetsfor the development of new approaches for anticancer therapy. Some CAFs originate from resident f ibroblastsof the organs invaded by the tumor, while others originate from epithelial tumor cells, which are undergoing anepithelial-mesenchymal transition (EMT). To date, many molecular and metabolic inducers of the EMT have beendiscovered including the transforming growth factor-beta (TGF-\u03b2), hypoxia, and inf lammation. This review classifies modern concepts of molecular markers of CAFs, their functional features, and discusses the stages of epithelial-mesenchymal transition, and the potential of CAFs as a target for antitumor therapy Modern concept of tumor morphology postulates that solidtumors are formed by epithelial and stromal cells, such asfibroblasts, endothelial cells, and immune cells . Stromal cells with a fibroblast-like phenotype, theso-called cancer-associated fibroblasts (CAFs), in contrast tonormal fibroblasts, contain various chromosomal abnormalities,such as duplications, multiple rearrangements, and eventhe loss of entire chromosomes . CAFscontrol tumor angiogenesis, motility and metastasis of cancercells, tumor immunogenic properties, and the developmentof resistance to chemotherapy and immunotherapy .A meta-analysis of the clinical relevance of the tumorstroma has demonstrated the association of high CAFs contentwith the advanced stages of tumor progression, as wellas with the high risk of local recurrence after tumor resection.In 1995, a heterogeneous origin of CAFs was hypothesizedby R\u00f8nnov-Jessen and colleagues, who showed that breastcancer CAFs can originate from resident fibroblasts, vascularsmooth muscle cells, and pericytes .To date, it has been shown that precursors of mesenchymalcells from the red bone marrow, endothelial and epithelialcells, resident fibroblasts of the affected tissue, adipocytesand vascular adventitia cells can be sources of CAFs . For the initiation of theCAFs phenotype in some progenitor cells, additional stimulationwith cytokines and growth factors, such as transforminggrowth factor beta (TGF-\u03b2), fibroblast growth factor (FGF),and other signaling molecules is required (Table 1) .Tumor epithelial cells can undergo transformation intoCAFs via the epithelial-mesenchymal transition (EMT). EMT is a dynamic process of transdifferentiation ofepithelial cells into fibroblast-like cells. The EMT plays animportant role not only in cancer, but also in embryogenesisand regeneration. In particular, EMT occurs in embryonic stemcells producing mesoderm and neural crest, and in skin cellsduring wound healing . Dynamic changesin cell morphology during EMT are caused by changes inthe regulatory genes\u2019 expression with production of certainproteins. These proteins are considered as EMT markers. Among these markers, the most significant areN-cadherin and vimentin, which are responsible for the rearrangementof the cytoskeleton and the change in the shapeof the cell, as well as the change in cell-to-cell and cell-toextracellularmatrix (ECM) interactions .In addition to molecular inducers of EMT, the importantrole of hypoxic conditions has been shown. Hypoxia activatesEMT via the binding of the hypoxia-inducible factor (HIF-1)to the promoters of genes responsible for EMT activation.HIF-1 has been shown to increase the expression of the transcriptionfactors genes of the zinc finger motif family such asZEB1, Snail and SLUG. Overexpression of these factors isassociated with the mesenchymal phenotype and a decreasein the abundance of epithelial cell markers \u2013 E-cadherin andtype 1 tight junction protein (TJP1 or ZO-1) .Endothelial cells of tumor vessels can undergo an endothelial-mesenchymal transition (EndMT) and acquire thephenotype and functional features of CAFs with the loss ofendothelial cells molecular markers, such as the endothelialcell/platelet adhesion molecule (CD31), and the acquisition ofmarkers specific for mesenchymal cells, such as \u03b1-SMA andfibroblast specific protein 1 (FSP-1) .An important component of breast cancer stroma areadipose cells, which can transform into tumor-associatedadipocytes, and then into CAFs. Such changes are accompaniedby an increase in the expression of molecular markersof mesenchymal cells, including, PPARG (receptors inducedby peroxisome activators gamma), RUNX-2 (transcriptionfactor containing the Runt type 2 DNA-binding domain), andSOX9 (transcription factor of the HMG family DNA-bindingproteins) .Using the model of prostate cancer, it was shown thatmesenchymal stem cells (MSCs) can differentiate into CAFsafter the activation of the chemokine receptor type 6 (CXCR6)by its ligand CXCL16. Moreover, the activation of CXCR6results in the secretion of stromal factor-1 (CXCL12) involvedin EMT . Weber and colleagues also showedthat the extracellular structural protein osteopontin (OPN),which plays a key role in bone formation, activates TGF-\u03b2gene expression in integrin-dependent MSCs to maintain thephenotype of CAFs in breast cancer. Interestingly, even specializedcells such as Ito cells in the liver, pancreatic stellatecells, and mammary myofibroblasts can acquire the phenotypeof CAFs . These examples illustrate a widerange of cells that, responding to the molecular changes ina tumor, are able to acquire the CAFs\u2019 phenotype and, as aconsequence, be involved in tumor homeostasis.The involvement of CAFs in carcinogenesis and tumor progressionmakes them a potential target for the developmentof novel therapeutic approaches. A potentially clinicallysignificant marker of CAF is the transmembrane mucin-likeprotein podoplanin (PDPN) (Table 2); to date, PDPN has beendescribed as a marker of lymphoid capillary progenitor cellsand CAFs in lung cancers. Expression of podoplanin wasshowed in 54 (30.5 %) out of 177 CAFs\u2019 populations studied inthe work of Yurugi et al. Interestingly, all podoplanin-positiveCAFs correlated with invasiveness of adenocarcinomas, whilea podoplanin-negative phenotype was shown only in noninvasiveadenocarcinomas .Platelet-derived growth factor receptors \u03b1/\u03b2 are importantmarkers of CAFs. PDGFR\u03b1/\u03b2 belong to the 3rd class oftyrosine kinases and are activated by interaction with thePDGF ligand. PDGFR regulates the organogenesis of varioussystems during embryogenesis; however, the significanceof the PDGFR\u03b1 and -\u03b2 receptors activation in tumors is stillpoorly understood. It has been shown that the expression ofthe PDGFR\u03b2 receptor is increased in the tumor microenvironmentcells, where platelet growth factor activates CAFsand, probably, stimulates cancer progression . PDGFR\u03b1-positive CAFs have been found in thestroma of melanoma, suggesting that these CAFs originatefrom resident fibroblasts as a result of their activation . Serum amyloid A (SAA-1) protein is one of thepotential targets of CAFs; its expression and involvementin tumor progression has been shown in CAFs from gastrictumors .In the search for specific markers of the tumor stroma cells,among the CAFs of prostate adenocarcinoma, an increasedcontent of the surface protein with a single V-domain of immunoglobulin(CD90), initially found on T cells and neurons,was identified as a specific marker. The high level of CD90 onthe cell surface differentiates the tumor-associated stroma and \u201cbenign\u201d stroma. Since CD90 expression was shown only intumor associated fibroblasts, this marker is a potential targetfor therapy .Certain CAFs proteins can be prognostic markers of tumorinvasiveness. One of these markers is a protein from the familyof low molecular weight calcium-binding proteins ofthe S100 \u2013 S100A4 family . S100 familyproteins have both intracellular and extracellular activitydue to maintaining the calcium balance and Ca2+-dependentprocesses. S100A4 activates a cascade of reactions associatedmainly with the secretion of pro-inflammatory cytokines andthe expression of growth factors, extracellular matrix proteins,metalloproteinases, and others. Intracellular activity ofS100A4 is of particular interest, and is associated with theenhancement of the invasive capabilities of tumor cells, theirescape from apoptosis, and the stem phenotype of the cells. During the study of the role ofS100A4 in tumor progression, it was shown that suppressionof S100A4 decreased tumor growth . The role of stromal cells thatsecrete S100A4 was shown in the MMTV-PyVmT mousemodel with the S100A4 knocked-out gene during orthotopicco-transplantation of CSML100 mouse mammary adenocarcinomacells and MEF mouse embryonic fibroblasts. MEFcell lines were obtained by spontaneous immortalization ofprimary embryonic fibroblasts from mouse embryos with theS100A4+ and S100A4\u2013 phenotypes. Upon co-transplantationof tumor cells and fibroblasts with the S100A4\u2013 phenotypein syngeneic mice, no metastases were formed, however,upon transplantation of S100A4+ fibroblasts, the metastaticpotential of tumor cells returned. S100A4+ fibroblasts werecharacterized by increased mobility and invasiveness comparedto S100A4\u2013 fibroblasts, as well as the ability to secreteS100A4 into the tumor microenvironment .To date, it is clear that the expression of certain CAFsmarkers does not unambiguously predict the aggressivenessof the tumor. For example, the loss of caveolin-1 in breastcancer CAFs has been shown to be associated with a poorprognosis because the population of these cells stimulates thegrowth of triple negative (ER-/PR-/HER2-) breast cancer cells. In a parallel study, the expression ofcaveolin-1 in the breast cancer CAFs stimulated the remodelingof the tumor microenvironment, thereby facilitating theinvasion of malignant cells and an increased invasiveness levelcorrelated with the metastatic potential of the tumor . These contradictingresultsindicate the diverse role of caveolin-1 in histologicallydifferent tumors. More studies should be made to determinecaveolin-1 as a tumor prognostic marker.CAFs-dependent stimulation of the tumor cells proliferationand their invasion is of particular interest in the study oftumor. This interest is primarily due to the fact that even inthe precancerous phenotype of epithelial cells, some residentfibroblasts are already transformed into CAFs . Fibroblasts from intestinal tumors and polyps werea good model to confirm the contribution of stromal cells totumor growth and progression. These fibroblasts were shownto stimulate the proliferation of tumor and polyp cells .The interaction of tumor epithelial cells with CAFs wasanalyzed by comparing the histological picture of varioustypes of gastric cancer. In a study by Orimo and Weinberg,it was demonstrated that in the case of diffuse gastric cancer,CAFs and epithelial cells are more closely spaced, while inthe intestinal type, CAFs form a stroma-like matrix, due towhich tumor epithelial cells retain their glandular structure.Using the model of heterogeneous 3D spheroids, consistingof breast cancer epithelial cells and fibroblasts, Dang and colleaguesshowed that CAFs stimulated the migration of tumorcells of basal breast cancer (ER-/PR-/HER2-). Interestingly,this effect was not observed in the models of luminal breastcancer types  . These data are consistent with clinical observationsindicating a higher percentage of metastasis in triplenegativebreast cancer compared with other types of breast tumors. However, which factors make cells of basal breastcancer sensitive to CAFs stimulation remain undiscovered.In order to reach the blood and lymphatic streams, tumorcells should pass through the basement membrane (BM), separatingthem from connective tissue and vessels. Thus, CAFsare able to synthesize metalloproteinases \u2013 endopeptidasescapable of destroying proteins of all types of the BM extracellularmatrix . In 2017, Glentis andcolleagues revealed a metalloproteinase-independent CAFssupportedovercome of BM by tumor cells. They demonstratedthe ability of CAFs to stretch BM with the formation of pores,and through these pores, epithelial tumor cells and CAFs canmigrate into the bloodstream and form metastases in distantorgans. Interestingly, BM regions with low expression oflamininand type IV collagen exhibited the highest tendencyfor stretching . This alternative CAFsdependentmigration pathway explains the ineffectiveness ofthe metalloproteinase inhibitors application in patients withhead and neck tumors.The paracrine secretion of IL-1\u03b1 by CAFs in bladder cancerwith further activation of the Wnt pathway in tumor cells isthe perfect illustration of the pro-carcinogenic role of CAFs. Moreover, bladder cancer \u0421AFs secretingIL-8 are able to stimulate the secretion of neuropilin-1,which enhances the proliferation of tumor cells and is oneof the potential prognostic markers of malignancy . Interestingly, recent studies have shown thatneuropilin-1 may be a co-factor in the induction of EMT.Paracrine stimulation of the epithelial-mesenchymal transitionin tumor epithelial cells by CAFs is an important factorcontributing to tumor progression. The central mechanism ofEMT in the tumor is the TGF-\u03b2/Smad pathway activation,induced by TGF-\u03b2 from stromal fibroblasts  . The Smad factor is a transcription factor that controlsthe expression of EMT genes. Vered and colleagues showedthat cells with EMT markers are found in primary foci ofsquamous cell carcinoma of the tongue as well as in regionallymph nodes metastases. This confirms the importance of CAFs in the induction of metastasis and in the formation ofa secondary tumor node .The central mechanism of the EMT activation in ovariancancer is the induction of the CXCR4/Wnt/\u03b2-catenin pathwayin tumor epithelial cells. CAFs secreting stromal growth factor-1 (SDF-1 or CXCL12) have been shown to be the majorplayers in this process. Moreover, SDF-1 is also interlinkedwith the resistance of tumor cells to chemotherapeutic agentssuch as cisplatin .In 2020, Franz\u00e8 and colleagues demonstrated the activationof CAFs phenotype in normal fibroblasts from rectal polypsco-cultured with CAFs. They showed that CAFs derivedfrom colorectal tumors can secrete IL-34, which in normalfibroblasts activates the expression of CAFs markers suchas \u03b1-SMA, vimentin, and fibroblast activating protein (FAP).Even though most of the described functions of \u0421AFs areassociated with the stimulation of tumor progression, someauthors also describe \u0421AFs as tumor-suppressing players. Forexample, the subpopulation of CAFs expressing the melanomaadhesion molecule (CD146) in breast tumors correlated witha retarded cell proliferation in estrogen-dependent types ofbreast cancer . Since CAFs secretecytokines involved in the recruitment and maturation ofmacrophages, T-lymphocytes and natural killer cells , they increase the availabilityof the tumor to immune cells and promote antitumor immuneresponse . In the study of oral cancer,it was also shown that CAFs can suppress the proliferationof tumor cells. In particular, the population of CAFs secretingBone Morphogenetic Protein 4 (BMP4) and expressing\u03b1-SMA inhibited the proliferation of cancer stem cells (CSC). Using the mice model with a predispositionfor the development of pancreatic cancer, Rhim and colleaguesexhibited the role of CAFs in cancer progression. They excludedthe \u03b1-SMA-positive population of CAFs or CAFs withinhibited Hedgehog signaling pathway. These modificationssuppress the growth of pancreatic ductal adenocarcinoma.Histological analysis of tumors revealed abnormalities in thevessel\u2019s formation .These examples demonstrate the tumor-suppressing functionof CAFs only in high differentiated cancers with nosimilarity in undifferentiated ones. It can be assumed thatapart from origin and tissue of the affected organ, the functionof CAFs is determined by the differentiation stage of cancercells .Cell cultures obtained from patients\u2019 tumors after surgery aremost often used to study the properties of CAFs. It is necessaryto establish new CAFs cell cultures, since in vitro CAFs tendto age rapidly, and the possibilities of their use are limitedby early passages . Moreover, in commerciallyavailable cell collections American Type CultureCollection (ATCC), European Collection of AuthenticatedCell Cultures (ECACC), Russian collection of cell cultures,etc. cell lines with the CAFs phenotype are limited. For instance,in ATCC, only one CAFs cell line is available. Thiscell culture originates from the prostate adenocarcinoma and ismodified by the introduction of the telomerase transgene underthe control of the constitutive promoter of the polyoma virusSV40 hTERT PF179T CAF. This modification of fibroblastsis aimed at maintaining the proliferative properties of CAFs.To obtain cell cultures from tumor tissue, mechanical disaggregation,enzymatic dissociation, chelation and their combinationare used. Trypsin and type IV collagenase are mostoften used to destroy the stroma of tumor tissue. The choice ofwhat technique to use should consider the histological originof the tissue of interest. When tissue disaggregated, CAFs canrepresent a small population of cells and obtaining a monoculturerequires an additional stage of their separation fromthe total cell mass. In order to isolate a particular populationof CAFs, magnetic separation, or FACS of cells with immunostainingof specific CAFs markers such as FAP or \u03b1-SMAare used . The main difficulty in isolating CAFs lies in adaptingprotocols for vital staining of intracellular markers such as\u03b1-SMA, FAP, and vimentin. Therefore, it is highly desirableto include surface markers such as CD90 in the analysis.Clear understanding the tumor microenvironment role iscrucial for the development of new approaches in cancer diagnosticsand treatment. The multifaceted influence of CAFson tumor progression makes them an important object for thestudy of carcinogenesis and the development of new antitumoragents. 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+{"text": "Metabolic comorbidities including diabetes (DM) and dyslipidemia pose challenges to the long-term care of people with HIV (PWH). Incidence of cardiovascular disease and DM are reported at higher rates in PWH than the general population. Obesity is broadly prevalent in both the general population and PWH, and higher body mass index (BMI) can contribute to metabolic complications. Here we present longer-term follow up on incidence of DM, hypertension (HTN), BMI categorical shifts, and lipid changes over 144 weeks of blinded treatment from two trials of PWH initiating antiretroviral therapy.We assessed incidence of metabolic complications in adult PWH in Study 1489: bictegravir/emtricitabine/tenofovir alafenamide (B/F/TAF) vs dolutegravir/abacavir/ lamivudine (DTG/ABC/3TC) and Study 1490: B/F/TAF vs DTG+F/TAF. Treatment-emergent (TE) metabolic comorbidities were defined by standard MedDRA search lists. CDC-defined BMI categories were compared from baseline (BL) to Week 144. Analyses by sex at birth and race were performed, as well as for lipid changes.Among 1,274 total participants, median (range) age was 33 years (18-77), 90% men, 33% black. In study 1489, BL prevalence of DM and HTN was 4.5 and 12.1% with TE DM and HTN in B/F/TAF being 0.7% and 10%, and for DTG/ABC/3TC 1.3% and 6.9%, respectively. In study 1490, BL prevalence of DM and HTN was 6.8 and 18.8% with TE DM and HTN in B/F/TAF being 2.1 and 5.8%, and for DTG+F/TAF 2.3 and 6.5%, respectively. BMI shift from Normal to Obese: B/F/TAF 0%, DTG/ABC/3TC 3.2%, p=0.12 (1489) (Table 1); B/F/TAF 2.5%, DTG+F/TAF 2.9% p=1.00 (1490) (Table 2). Subgroup analyses by gender/race showed similar findings for TE DM, HTN, and BMI changes. Median changes from BL fasted lipids were small (Table 1).Table 1\u00a7. Studies 1489 and 1490: Metabolic Outcomes from Baseline to Week 144Table 2\u00b1. Shift Table of BMI Category at Week 144 by Baseline BMI Category \u2013 OverallThrough over 144 weeks of follow up, PWH randomized to initiate B/F/TAF, DTG/ABC/3TC or DTG+F/TAF had low rates of incident DM or HTN-related AEs, with no statistically significant differences by treatment group. BMI changes/categorical shifts from BL did not significantly differ by regimen, and no clinically significant change or difference by regimen in lipids were observed. While data are limited by three years of follow up, they are strengthened by randomized study design of three widely used initial ART regimens.Eric Daar, MD, Bristol-Myers Squibb (Consultant)Gilead Sciences Inc. Janssen Merck Teva ViiV Healthcare  Chloe Orkin, MD, Gilead Sciences Inc. Janssen Merck ViiV Healthcare  Paul Sax, MD, Gilead Sciences Janssen (Consultant)Merck ViiV  Jeffrey L. Stephens, MD, Gilead Sciences Inc.  Ellen Koenig, MD, Gilead Sciences Inc. (Scientific Research Study Investigator) Amanda Clarke, MD, Gilead Sciences Inc. ViiV Healthcare  Axel Baumgarten, MD, AbbVie Bristol-Myers Squibb Gilead Sciences Inc. Janssen (Speaker\u2019s Bureau)Merck (Advisor or Review Panel member) Cynthia Brinson, MD, Abbvie (Scientific Research Study Investigator)BI (Scientific Research Study Investigator)Gilead Sciences Inc. GSK (Scientific Research Study Investigator)Novo Nordisk (Scientific Research Study Investigator)ViiV Healthcare  Moti Ramgopal, MD FIDSA, Abbvie Gilead Janssen Merck ViiV  Hailin Huang, PhD, Gilead Sciences Inc.  Terry Farrow, MD, Gilead Sciences Inc.  Jared Baeten, MD, PHD, Gilead Sciences Inc.  Jason Hindman, PharmD, Gilead Sciences Inc.  Hal Martin, MD, MPH, Gilead Sciences Inc.  Kimberly Workowski, MD, Nothing to disclose"}
+{"text": "Cabotegravir (CAB) plus rilpivirine (RPV) is the first complete long-acting (LA) regimen recommended by treatment guidelines for the maintenance of HIV-1 virologic suppression. CAB+RPV LA dosed every 4 weeks (Q4W) or every 8 weeks (Q8W) demonstrated noninferior efficacy in multinational Phase 3/3b trials. This This analysis focuses on data for US/CAN participants naive to CAB+RPV (n=376) from the larger pooled population of the ATLAS, FLAIR, and ATLAS-2M Phase 3/3b studies (N=1245). Endpoints included the proportion of participants with plasma HIV-1 RNA \u2265 50 and < 50 c/mL at W48 , incidence of confirmed virologic failure , safety, and treatment preference through W48.2) previously associated with CVF. Among the US/CAN participants with a single baseline factor, none met CVF. Overall, archived RPV RAMs were observed in 3.2% (12/376), HIV subtype A6/A1 in 1.1% (4/376), and BMI \u2265 30 kg/m2 in 26.3% (99/376) of participants. Safety and injection site reaction findings were similar to the overall pooled population (Table 2). Most participants  preferred LA over oral dosing . 376 US/CAN participants received CAB+RPV LA Q4W or Q8W. Median (range) age was 39y (20\u201374); 14.9% were female, 66.0% were White. At W48, 93.1% (350/376) maintained virologic suppression (HIV-1 RNA < 50 c/mL), 1.9% (7/376) had HIV-1 RNA \u2265 50 c/mL, and 0.8% (3/376) met the CVF criterion, consistent with the overall global pooled population (Table 1). Two of the three participants with CVF had \u2265 2 of the three baseline factors Table 2. Safety summary through Week 48 following CAB+RPV LA Q4W and Q8W or comparator ART in participants naive to CAB+RPV from ATLAS, FLAIR, and ATLAS-2MIn US/CAN Phase 3/3b trial participants, CAB+RPV LA was highly effective and well tolerated, with outcomes consistent with the overall pooled population. Baseline prevalence of archived RPV RAMs and subtype A6/A1 was low and aligned with regional prevalence/surveillance data. CAB+RPV LA provides a tolerable and effective injectable LA treatment option for virologically suppressed US/CAN individuals with HIV.Babafemi O. Taiwo, MBBS, Gilead (Consultant)Merck (Consultant)ViiV Healthcare (Consultant) Darrell Tan, MD PhD, Abbvie (Grant/Research Support)Gilead (Grant/Research Support)GlaxoSmithKline (Scientific Research Study Investigator)ViiV Healthcare (Grant/Research Support) Parul Patel, PharmD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Paula Teichner, PharmD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Joseph Polli, PhD, FAAPS, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Louise Garside, PhD, GlaxoSmithKline (Employee) Ronald D\u2019Amico, DO, MSc, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Christine L. Talarico, M.S., GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Rodica Van Solingen-Ristea, MD, Janssen Research and Development (Employee)ViiV Healthcare (Employee) Bryan Baugh, MD, Janssen, Johnson & Johnson  William Spreen, PharmD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Michael Aboud, MBChB, MRCP, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Matthew Bosse, DO, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee)"}
+{"text": "Dengue fever is a mosquito-borne viral disease endemic in 128 countries. An unmet clinical need remains for an effective vaccine that can be used more broadly than the vaccine presently available. A clinical development program has evaluated the long-term safety, immunogenicity, and vaccine efficacy (VE) of TAK-003, a live attenuated tetravalent dengue vaccine with a DENV-2 backbone engineered to elicit immune responses to all 4 dengue serotypes. nd dose, and for another 25 months after a booster dose. Data up to 3 years after the second vaccination are currently available.18 clinical trials in 13 countries have involved 28,175 seropositive/seronegative participants aged from 1.5-60 years from endemic/non-endemic regions. In the ongoing pivotal phase III study, 4\u201316-year-old healthy children  were randomized 2:1 to receive two doses of TAK-003 or placebo, 3 months apart for an evaluation of VE and safety over a multi-year period stratified pre-vaccination dengue serostatus. Active surveillance throughout the trial detected symptomatic dengue. The trial will continue up to 4\u20134.5 years post 2nd dose. At 18 months, VE was 66.2% (95% CI: 49.1\u201377.5) in dengue-naive and 76.1% (95% CI: 68.5\u201381.9) in dengue pre-exposed participants, with VE of 90.4% (95% CI: 82.6\u201394.7) and 85.9% (95% CI: 31.9\u201397.1) for prevention of hospitalized VCD and dengue hemorrhagic fever, respectively. Cumulative VE against VCD from first dose to 3 years post 2nd dose was 62.0% (95% CI: 56.6\u201366.7) and 83.6% (95% CI: 76.8\u201388.4) in prevention of hospitalized VCD. Some decline in VE was observed over time mainly driven by outpatient dengue. Two doses of TAK-003 3 months apart were well-tolerated with no important safety risks identified up to 3 years after completion of the vaccination schedule. Safety and immunogenicity data from Phase I/II studies established the final formulation and dosing schedule. Overall VE in the pivotal phase III study was 80.2% [95% CI: 73.3\u201385.3] against virologically confirmed dengue (VCD) at 12 months post 2TAK-003 is immunogenic against all 4 dengue serotypes and continues to be efficacious, well-tolerated, and with no evidence of disease enhancement in seronegative population up to 3 years post-vaccination.Vianney Tricou, D Phil, Takeda Pharmaceuticals International (Employee) Shibadas Biswal, MD, Takeda Vaccines, Inc (Employee) Sanjay S. Patel, PhD, Takeda Pharmaceuticals International AG (Employee) Olaf Zent, MD, Takeda Pharmaceuticals International AG (Employee) Martina Rauscher, PhD, Takeda Pharmaceuticals International AG (Employee) Gonzalo Perez, MD, Takeda group companies (Employee) Walid Kandeil, MD, Takeda Pharmaceuticals International AG (Employee) Nicolas Folschweiller, PhD, Takeda (Employee)"}
+{"text": "KBP-7072 was active against Staphylococcus aureus , methicillin-resistant S. aureus , S. lugdunensis , and other coagulase-negative staphylococci . KBP-7072 was active against Enterococcus faecalis  and vancomycin-susceptible and -nonsusceptible E. faecium ; Streptococcus pneumoniae , including penicillin- and tetracycline-resistant strains; S. agalactiae , including macrolide-resistant strains; S. pyogenes ; and viridans group streptococci, including S. anginosus group  isolates. KBP-7072 inhibited 90.2%  of all Enterobacterales isolates, including expanded-spectrum \u03b2-lactamase-phenotype strains at \u22642\u2009mg/liter. KBP-7072 demonstrated potent activity against Acinetobacter baumannii-calcoaceticus species complex and Stenotrophomonas maltophilia isolates , Haemophilus influenzae , and Moraxella catarrhalis . Based on MIC90 values, KBP-7072 in vitro activity was generally superior to that the other tetracycline class comparators tested. The potent activity of KBP-7072, including resistant organism groups, merits further clinical investigation in infections where these organisms are likely to occur.KBP-7072 is a novel broad-spectrum tetracycline (aminomethylcycline) antibacterial in clinical development  for the treatment of acute bacterial skin and skin structure infections, community-acquired bacterial pneumonia, and complicated intra-abdominal infections. KBP-7072 is active against many of the World Health Organization priority pathogens. In this study, KBP-7072 and tetracycline class comparators were susceptibility tested against 1,057 geographically diverse surveillance isolates from 2019 according to Clinical and Laboratory Standards Institute (CLSI) guidelines. KBP-7072 demonstrated potent  The effectiveness of tetracycline antibacterials has declined since their initial discovery and introduction in the late 1940s, primarily due to the emergence of resistance caused by efflux and ribosomal protection mechanisms . This re1\u20135\u2013KBP-7072  has comp10\u2013in vitro antibacterial activity against many of the organisms listed on the World Health Organization (WHO) priority pathogen list, including methicillin-resistant Staphylococcus aureus, penicillin-nonsusceptible Streptococcus pneumoniae, vancomycin-resistant Enterococcus faecium, ampicillin-resistant Haemophilus influenzae, and carbapenem-resistant Acinetobacter baumannii , percent intermediate (I), and percent resistant (R) according to CLSI or FDA breakpoint interpretive criteria, are presented in 90 values, KBP-7072 was the most active agent tested against S. aureus, including methicillin-resistant S. aureus (MRSA) , tetracycline-resistant S. aureus , and enterococci , including vancomycin-nonsusceptible strains , S. agalactiae , S. pyogenes , and S. pneumoniae , including penicillin-resistant, tetracycline-resistant, and macrolide-resistant strains at \u22640.12\u2009mg/liter  was comparable in activity to tigecycline , 8-fold more active than omadacycline and minocycline , 16-fold more active than doxycycline , and >32-fold more active than tetracycline   was 4-fold more active than tigecycline , 8-fold more active than omadacycline and minocycline , >32-fold more active than doxycycline , and >64-fold more active than tetracycline  (H. influenzae isolates (100.0%) were inhibited by \u22640.25\u2009mg/liter KBP-7072 (Cumulative percent inhibition and MIC strains  and 2. Kmg/liter . Againstg/liter) . When teg/liter) . All H. KBP-7072 .50/90, 0.06/0.12\u2009mg/liter; 100.0% inhibited at \u22640.5\u2009mg/liter) demonstrated potent in vitro activity against 104 S. aureus isolates, including methicillin-susceptible S. aureus (MSSA)  and MRSA  organism subsets  was 2-fold more active than minocycline , omadacycline , and tigecycline , 8-fold more active than doxycycline , and 128-fold more active than tetracycline  (KBP-7072 (MIC subsets  and 2. Sas 11.5% . Based oeptible) .50/90, 0.03/0.03\u2009mg/liter; 100.0% inhibited at \u22640.03\u2009mg/liter) demonstrated potent in vitro activity against 20 Staphylococcus lugdunensis isolates where resistance to tetracycline was 5.0%  was comparable in activity to minocycline , 2-fold more active than doxycycline , omadacycline , and tigecycline  and 4-fold more active than tetracycline  (KBP-7072 (MICwas 5.0% . Based oeptible)  and 2.50/90, 0.06/0.25\u2009mg/liter, 100% inhibited at \u22640.5\u2009mg/liter) and tigecycline  were comparable in activity against 22 other coagulase-negative staphylococci (CoNS) isolates  was 2-fold more active than minocycline , 4-fold more active than omadacycline , 32-fold more active than doxycycline , and 256-fold more active than tetracycline  against other CoNS isolates  isolates  was 2-fold more active than omadacycline and tigecycline  , minocycline , and tetracycline  demonstrated limited activity against E. faecalis isolates (KBP-7072 was highly active against 51 isolates . Based oeptible) . Doxycycisolates .E. faecium  isolates, and its activity was not adversely affected by susceptibility or nonsusceptibility to vancomycin  was 2-fold more active than tigecycline  and 4-fold more active than omadacycline  , minocycline , and tetracycline  demonstrated reduced activity against E. faecium isolates  . Doxycycisolates .S. pneumoniae isolates (n\u2009=\u2009127) were highly susceptible to KBP-7072 , and its activity was not adversely affected by resistance to erythromycin , penicillin , or tetracycline   was comparable in activity to tigecycline , 2-fold more active than omadacycline , 256-fold more active than doxycycline  and minocycline , and 2,048-fold more active than tetracycline  , and 27.6%, respectively , 22.0% for clindamycin, and 18.1% for trimethoprim-sulfamethoxazole (data not shown).g/liter)  and 2. Beptible) . Resistaectively . ResistaS. agalactiae  and 51 S. pyogenes isolates , were inhibited by \u22640.06\u2009mg/liter of KBP-7072 regardless of erythromycin (macrolide) resistance  was comparable in activity to tigecycline , 4-fold more active than omadacycline , 256-fold more active than doxycycline , 512-fold more active than minocycline , and 1,024-fold more active than tetracycline   was 2-fold more active than tigecycline , 4-fold more active than omadacycline , 256-fold more active than doxycycline and minocycline , and 1,024-fold more active than tetracycline   . Similareptible) .Streptococcus anginosus group isolates (n\u2009=\u200917) were inhibited by low concentrations of KBP-7072  and tigecycline  , 256-fold more active than doxycycline (MIC90 8\u2009mg/liter), 512-fold more active than minocycline , and 1,024-fold more active than tetracycline  .All eptible) . Based oeptible) . Resista50/90, 0.25/2\u2009mg/liter; 90.2% inhibited at \u22642\u2009mg/liter) and tigecycline  were the most active tetracycline class compounds tested against 410 Enterobacterales isolates  was 8-fold more active than omadacycline  and minocycline , 16-fold more active than doxycycline , and >32-fold more active than tetracycline  against Enterobacterales isolates  was comparable in activity to tigecycline against 133 tetracycline-resistant Enterobacterales isolates  and meropenem  (data not shown).KBP-7072  and tigecycline  were the most active tetracycline class agents tested against 22 Citrobacter freundii species complex isolates  was 8-fold more active than omadacycline  and 16-fold more active than doxycycline, minocycline, and tetracycline  (KBP-7072 (MICisolates . Based oeptible) .Citrobacter koseri isolates (n\u2009=\u200921) were susceptible (100.0%) to doxycycline, minocycline, tetracycline, and tigecycline. KBP-7072 and tigecycline were the most active tetracycline class agents against C. koseri, with MIC90 values of 0.25\u2009mg/liter.All 90, 0.5\u2009mg/liter; 100.0% inhibited at \u22644\u2009mg/liter) and tigecycline  were the most active tetracycline class agents tested against 50 Enterobacter cloacae species complex isolates  was 4-fold more active than doxycycline  and 8-fold more active than minocycline , omadacycline , and tetracycline   was equally active against ceftazidime-susceptible and ceftazidime-nonsusceptible (AmpC-derepressed phenotype) E. cloacae species complex isolates (KBP-7072 (MICisolates  and 3. Beptible) . KBP-707isolates  and 3.50/90, 0.12/0.5\u2009mg/liter; 100.0% inhibited at \u22642\u2009mg/liter) was active against 77 Escherichia coli isolates, including a subset of 26 expanded-spectrum \u03b2-lactamase (ESBL)-phenotype E. coli isolates   was comparable in activity to tigecycline , 4-fold more active than omadacycline , 8-fold more active than minocycline , 32-fold more active than doxycycline , and >64-fold more active than tetracycline  (KBP-7072 (MICg/liter)  and 3. Aeptible) .90, 0.5\u2009mg/liter; 100.0% inhibited at \u22644\u2009mg/liter) and tigecycline  were the most active tetracycline class agents tested against 21 Klebsiella aerogenes isolates  was 4-fold more active than minocycline  and omadacycline , 8-fold more active than doxycycline , and 16-fold more active than tetracycline  (KBP-7072 (MICisolates . Based oeptible) .90, 0.5\u2009mg/liter; 100.0% inhibited at \u22642\u2009mg/liter) and tigecycline  were comparable in activity against 53 K. oxytoca isolates  was 4-fold more active than omadacycline , 8-fold more active than minocycline , and 16-fold more active than doxycycline and tetracycline  (KBP-7072 (MICisolates . Based oeptible) .50/90, 0.25/1\u2009mg/liter; 100.0% inhibited at \u22644\u2009mg/liter) was active against 80 K. pneumoniae isolates, including a subset of 27 ESBL-phenotype K. pneumoniae isolates   was comparable in activity to tigecycline , 8-fold more active than omadacycline , 16-fold more active than doxycycline , >16-fold more active than minocycline , and >32-fold more active than tetracycline  (KBP-7072 (MICg/liter)  and 3. Aeptible) .50/90, 1/2\u2009mg/liter; 95.0% inhibited at \u22642\u2009mg/liter) and tigecycline  were the most active tetracycline class agents tested against 20 Morganella morganii isolates , minocycline , doxycycline , and tetracycline  (KBP-7072 (MICisolates  and 3. Reptible) .Proteus mirabilis isolates, including KBP-7072 , doxycycline , minocycline , omadacycline , tetracycline , and tigecycline  (All tetracycline class agents demonstrated reduced or limited activity against 22 eptible)  and 3.90, 4\u2009mg/liter; 95.5% inhibited at \u22644\u2009mg/liter) and tigecycline  were the most active tetracycline class agents tested against 22 Providencia species isolates , minocycline , omadacycline , and tetracycline  (KBP-7072 (MICisolates  and 3. Oeptible) .90, 1\u2009mg/liter; 95.5% inhibited at \u22641\u2009mg/liter) and tigecycline  were the most active tetracycline class agents tested against 22 Serratia marcescens isolates  was 4-fold more active than minocycline , 8-fold more active than omadacycline , 16-fold more active than doxycycline , and >64-fold more active than tetracycline  (KBP-7072 (MICisolates  and 3. Beptible) .in vitro activity, KBP-7072  was the most potent tetracycline class agent tested against 22 A. baumanniicalcoaceticus species complex isolates  was 4-fold more active than tigecycline , 8-fold more active than minocycline  and omadacycline , >32-fold more active than doxycycline , and >64-fold more active than tetracycline  .Based on isolates . KBP-707eptible) . Compara50/90, 8/16\u2009mg/liter) and all tetracycline class agents was limited against 22 Pseudomonas aeruginosa isolates  and minocycline  demonstrated potent in vitro activity against 22 Stenotrophomonas maltophilia isolates  was 2-fold more active than tigecycline , 8-fold more active than omadacycline , and 64-fold more active than tetracycline .KBP-7072 , doxycycline , minocycline , omadacycline , tetracycline , and tigecycline  (All tetracycline class agents were very active against 52 eptible) .50/90, 0.25/0.5\u2009mg/liter; 100.0% inhibited at \u22640.5\u2009mg/liter) was active against 12 H. parainfluenzae isolates  was comparable in activity to tigecycline , 2-fold more active than tetracycline , and 4-fold more active than doxycycline , minocycline , and omadacycline  (KBP-7072 (MICisolates  and 3. Beptible) .50/90, 0.06/0.06\u2009mg/liter; 100.0% inhibited at \u22640.06\u2009mg/liter), were active against 21 Moraxella catarrhalis isolates  pathogens, vancomycin-resistant E. faecium and MRSA were identified as priority 2 (high) pathogens, and ampicillin-resistant H. influenzae and penicillin-nonsusceptible S. pneumoniae were identified as priority 3 (medium) pathogens  at a dose level of 200\u2009mg. The therapeutic dose for KBP-7072 is projected to be lower than the current daily oral dose for another aminomethylcycline class antibacterial (omadacycline) in community-acquired bacterial pneumonia and acute bacterial skin and skin structure infection ; tetracycline-resistant S. aureus (8-fold); E. faecium (4-fold); S. pneumoniae (2-fold), including penicillin-nonsusceptible strains; beta-hemolytic streptococci (4-fold); Enterobacterales (8-fold), including ESBL-phenotype and tetracycline-resistant strains; A. baumannii (8-fold); S. maltophilia (8-fold); and H. influenzae (4-fold).KBP-7072 has demonstrated dose-proportional pharmacokinetic/pharmacodynamic (PK/PD) properties in both animal models and phase I clinical studies supporting once-daily oral or intravenous administration 1013, 18\u20132018\u2013nfection . A lowerin vitro activity of KBP-7072 was unaffected by isolates displaying resistance to tetracycline. KBP-7072 remained active against isolates displaying resistance to other antibacterial agents, including ampicillin, ceftazidime, erythromycin, penicillin, and vancomycin.The in vitro activity of KBP-7072 against A. baumannii is also supported by a prior study using 531 isolates that included carbapenem-resistant, colistin-resistant, ESBL-positive, metallo-\u03b2-lactamase-producing, and tetracycline-resistant strains  were collected from 117 medical centers located in 35 countries, including the United States , Europe , Latin America , and the Asia-Pacific region  as part of the SENTRY Surveillance Program. The surveillance isolates utilized in this study were randomly selected from patients with skin and skin structure infections , pneumonia in hospitalized patients , and community-acquired respiratory tract infections  and included only 1 isolate/patient/infection episode. The percentage of tetracycline-resistant isolates in this study generally mimicked the 2019 worldwide SENTRY Surveillance Program distributions for A. baumannii (54.5% versus 60.3% in SENTRY), Enterobacterales (32.4% versus 34.6% in SENTRY), H. influenzae (1.9% versus 1.4% in SENTRY), S. aureus (11.5% versus 6.0% in SENTRY), S. agalactiae (80.8% versus 79.6% in SENTRY), S. pneumoniae (27.6% versus 24.7% in SENTRY), and S. pyogenes (19.6% versus 21.2% in SENTRY). Organism identifications were confirmed by matrix-assisted laser desorption ionization time-of-flight mass spectroscopy .Geographically diverse, recent (2019) bacterial clinical isolates (KBP-7072 and omadacycline (KBP-3039) powders were supplied by KBP Biosciences Co., Ltd. . Doxycycline, tetracycline, and tigecycline powders were obtained from the United States Pharmacopeial Convention . Minocycline powder was obtained from Sigma-Aldrich .E. coli and K. pneumoniae isolates, MRSA, vancomycin-susceptible and -nonsusceptible E. faecium, penicillin-susceptible, -intermediate, -resistant , and tetracycline-resistant S. pneumoniae, erythromycin (macrolide)-resistant S. agalactiae and S. pyogenes, ceftazidime-susceptible and -nonsusceptible E. cloacae, carbapenem (meropenem)-resistant A. baumannii, and ampicillin-resistant H. influenzae  phenotype for fluenzae . Most ESanalysis .Broth microdilution susceptibility testing was conducted at JMI Laboratories according to Clinical and Laboratory Standards Institute M07  and M100E. faecalis ATCC 29212, E. coli ATCC 25922, E. coli ATCC 35218, P. aeruginosa ATCC 27853, and S. aureus ATCC 29213. All (100.0%) of the doxycycline (18/18), minocycline (15/15), omadacycline (18/18), tetracycline (25/25), and tigecycline (18/18) MIC values obtained were within CLSI-approved quality control ranges (JMI Laboratories followed current CLSI quality assurance practices when performing susceptibility tests. MIC values were validated by concurrently testing the CLSI-recommended  American"}
+{"text": "Scientific Reports 10.1038/s41598-021-84500-6, published online 23 March 2021Correction to: The original version of this Article contained errors in References 27, 43 and 44, where hyperlinks to external data were omitted and as a result were incorrectly given as:27. Kramarenko, A. V. Demo of non-contac polatimetric cardiograph testing, 43. Kramarenko, A. V. Demo of an industrial application (pump work control) of a non-contact rf registration device .44. Kramarenko, A. V. Demo of a car driver monitoring system .The correct references are listed below:https://www.youtube.com/watch?v=gOGvGjJ2QnI (Accessed 23 Jul 2020a).27. Kramarenko, A. V. Demo of non-contac polatimetric cardiograph testing, https://www.youtube.com/watch?v=z-1pzf3iUyM (Accessed 3 Nov 2020b).43. Kramarenko, A. V. Demo of an industrial application (pump work control) of a non-contact rf registration device, https://www.youtube.com/watch?v=yB9uKd3MNxU (Accessed 13 Oct 2020c).44. Kramarenko, A. V. Demo of a car driver monitoring system, The original Article has been corrected."}
+{"text": "TB meningitis is the most severe form of tuberculosis (TB), associated with high morbidity and mortality. High-dose rifampin (35mg/kg/day) is safe in adults and substantially improves the bactericidal activity of standard TB regimen. However, there is conflicting data regarding its benefit in TB meningitis where outcomes may also be associated with intracerebral inflammatory responses. Mycobacterium tuberculosis infections were used for these studies . Animals received high-dose (35 mg/kg/day) or standard-dose (10 mg/kg/day) rifampin in combination with isoniazid, pyrazinamide and dexamethasone at human equipotent dosing. Bacterial burden, multi-modality positron emission tomography (PET) imaging, tissue drug concentrations, markers of neuroinflammation, and vascular leak were measured. Imaging data from a patient with TB meningitis was analyzed and correlated with the findings in animals.A novel mouse and a validated rabbit model of TB meningitis utilizing intracranial Figure 1. Mouse model of TB meningitis replicates human histopathology hallmarks. (A) Scheme of infection. (B) Histopathology hematoxylin-eosin (H&E) and acid-fast bacilli (AFB) staining in a representative M. tuberculosis-infected mouse shows regions of meningitis, ventriculitis, choroiditis, necrotizing and non-necrotizing granulomas. The bar represents 100\u00b5m. (C) Images show immunofluorescence of microglia activation in red (Iba-1) and nuclear stain in blue (DAPI). The rabbit model of TB meningitis has been described previously . Animal studies were approved by the Johns Hopkins Animal Care and Use Committee.P < 0.01) . There were no differences in intracerebral microglial activation (124I-DPA-713 PET and iDISCO) and pro-inflammatory cytokines during treatment in animals receiving high- or standard-dose rifampin regimens . Whole-brain PET and immunolabeling demonstrated spatially compartmentalized inflammation, vascular leak and rifampin exposures . Longitudinal imaging in the same animals showed a 40% decrease in vascular leak after two weeks of TB treatment. Spatially compartmentalized brain rifampin exposures and decreases in vascular edema over TB treatment were also noted in the TB meningitis patient.Administration of the high-dose rifampin regimen achieved four times higher brain concentration than the standard-dose regimen and displayed higher bactericidal activity in both mice and rabbits  Experimental scheme. R10 (rifampin 10mg/kg), R35 (rifampin 35mg/kg), H (isoniazid), Z (pyrazinamide), D (dexamethasone). (B) Bactericidal activity of high-dose rifampin  and standard-dose rifampin  regimens in mice. (C) Grouped colony forming units (CFU) and (D) rifampin brain concentration in mice after 42 days of TB treatments. (E) Bactericidal activity of high-dose rifampin  and standard-dose rifampin  regimens in rabbits. Data from untreated rabbits  is also shown. (F) Grouped CFU and (G) rifampin brain concentration in rabbits after 14 days of TB treatments. Tissues were assayed using validated ultra-high-performance liquid chromatography and tandem mass spectrometry (LC-MS/MS) for rifampin at the Infectious Diseases Pharmacokinetics Laboratory of the University of Florida. The bacterial burden is represented as CFU per gram of tissue and presented on a logarithmic scale. CFU and mass spectrometry data are represented as mean \u00b1 SD. *P < 0.05, **P < 0.01 and ***P < 0.001 calculated using a two-way ANOVA test.Figure 3. Neuroinflammatory responses during TB treatment. (A) Iba-1 and DAPI immunofluorescence in a representative untreated, high-dose and low-dose-treated mouse. (B) Quantification of Iba-1 immunofluorescence before treatment and after 6 weeks of treatment (n = 3 mice per group). Sections were imaged at 40X with Nikon A1+ confocal microscope. HALO was used for visualization and quantification. Quantification of intraparenchymal (C) INF-\u03b3, (D) TNF and (E) MCP-1 in untreated and treated mice (Milliplex Multiplex Luminex assay). (F) Coronal CT and 124I-DPA-713 PET/CT of a representative mouse with TB meningitis before treatment initiation. (G) 124I-DPA-713 PET quantification before treatment  and after 2  and 6  weeks of TB treatment. PET data is represented as median \u00b1 IQ. *P < 0.05, **P < 0.01 and ***P < 0.001 were calculated using a two-way ANOVA test.Figure 4. Changes in vascular leakage and rifampin penetration during TB treatment. (A) Experimental scheme in mice. (B) Whole-brain immunolabeling (iDISCO) of a representative M. tuberculosis-infected mouse prior to treatment initiation. Images show immunolabeling of \u03b1-smooth muscle actin in grey and microglia activation in red (Iba-1). Asterix represents the areas of vascular amputation. (C) Coronal 18F-py-Albumin PET/CT and quantification (tissue to plasma ratio) in untreated and 2 weeks-treated mice . (D) Serial imaging in a patient with TB meningitis. (E) Transverse T2 post-contrast section and maximal intensity projection (MIP) showing vasogenic edema. (F) Co-registered T2 post-contrast MIP with transverse 11C-rifampin area under the curve (AUC) heat-map, and 11C-rifampin tissue to plasma ratio quantification of the areas with and without vasogenic edema, and unaffected brain. (G) T2 post-contrast MIP at 3 and 45 months after treatment initiation. The patient with TB meningitis was recruited as part of a 11C-rifampin PET research study performed under the U.S. FDA Radioactive Drug Research Committee program for investigational drugs . Human studies were approved by the Johns Hopkins University Institutional Review Board Committee. M = moxifloxacin. PET data is represented as median \u00b1 IQ. *P < 0.05, **P < 0.01 and ***P < 0.001 calculated using a two-way ANOVA test.Our cross-species findings suggest that an intensified high-dose rifampin regimen is more efficacious than the standard treatment for TB meningitis without an increase in neuroinflammation. Vascular leak decreases during TB treatment and may account for decreases in rifampin permeability over time. These studies have important implications for antimicrobial development for TB meningitis.Charles A. Peloquin, Pharm.D., Nothing to disclose Alvaro A. Ordonez, MD, Cubresa (Consultant)Fujirebio Diagnostics (Research Grant or Support) Sanjay K. Jain, MD, Fujirebio Diagnostics, Inc., USA (Research Grant or Support)Novobiotic LLC, USA (Research Grant or Support)T3 Pharma, Switzerland (Research Grant or Support) Sanjay K. Jain, MD, Fujirebio Diagnostics, Inc., USA  Involved: Self): Research Grant or Support; Novobiotic LLC, USA  Involved: Self): Research Grant or Support; T3 Pharma, Switzerland  Involved: Self): Research Grant or Support"}
+{"text": "CLSI broth microdilution was used to determine MICs for Enterobacterales , Pseudomonas aeruginosa , Acinetobacter baumannii complex , Stenotrophomonas maltophilia , and Burkholderia cepacia complex (n\u2009=\u2009425). MICs were interpreted by CLSI-approved clinical breakpoints (February 2021). Cefiderocol inhibited 99.8, 96.7, 91.6, and 97.7% of all Enterobacterales, meropenem-nonsusceptible, ceftazidime-avibactam-nonsusceptible, and ceftolozane-tazobactam-nonsusceptible isolates, respectively, at \u22644\u2009\u03bcg/mL (susceptible breakpoint). Cefiderocol inhibited 99.9, 99.8, 100, and 99.8% of all P. aeruginosa, meropenem-nonsusceptible, ceftazidime-avibactam-nonsusceptible, and ceftolozane-tazobactam-nonsusceptible isolates, respectively, at \u22644\u2009\u03bcg/mL (susceptible breakpoint). Cefiderocol inhibited 96.0% of all A. baumannii complex isolates and 94.2% of meropenem-nonsusceptible isolates at \u22644\u2009\u03bcg/mL (susceptible breakpoint) and 98.6% of S. maltophilia isolates at \u22641\u2009\u03bcg/mL (susceptible breakpoint). B. cepacia complex isolates were tested with a MIC50 of \u22640.03\u2009\u03bcg/mL and MIC90 of 0.5\u2009\u03bcg/mL. Annual cefiderocol percent susceptible rates for Enterobacterales  and P. aeruginosa  were unchanged from 2014 to 2019. Annual percent susceptible rates for A. baumannii complex demonstrated sporadic, nondirectional differences ; the wider range for Europe (\u223c7%) was due to isolates from Russia. Annual percent susceptible rates for S. maltophilia showed minor, nondirectional differences . We conclude that clinical isolates of Enterobacterales (99.8% susceptible), P. aeruginosa (99.9%), A. baumannii (96.0%), and S. maltophilia (98.6%) collected in North America and Europe from 2014 to 2019 were highly susceptible to cefiderocol.We report  Enterobacterales, Pseudomonas aeruginosa, and Acinetobacter baumannii as pathogens of the highest  priority for development of new antibacterial agents  when limited or no other treatment options exist , P. aeruginosa, and Acinetobacter baumannii complex. Clinical development of cefiderocol continues for the treatment of serious infections attributable to resistant Gram-negative bacilli, including infections caused by carbapenem-resistant Gram-negative bacilli , and difficult-to-treat Gram-negative bacilli is increasing worldwide, and therapeutic options for infected patients are often limited \u20133. The W\u2013l agents . Cefiderns exist . In Aprins exist . In Sept bacilli .Cefiderocol possesses a unique mechanism of bacterial cell entry, making it an important addition to the antimicrobial armamentarium. The optimized chloro-catechol moiety within the C-3 side chain of cefiderocol facilitates formation of chelated complexes with ferric iron and expedites its transport across the outer membrane of Gram-negative bacilli using constitutive iron transport systems . Followi7\u2013in vitro activities for recently approved and investigational agents is critical to establishing and supporting treatment decisions and expanding the role of these agents in patient care, particularly for patients where unmet medical need exists , 8\u2009\u03bcg/mL (intermediate), and \u226516\u2009\u03bcg/mL (resistant) and for Stenotrophomonas maltophilia of \u22641\u2009\u03bcg/mL (susceptible) and >1\u2009\u03bcg/mL (nonsusceptible)  . The updeptible)  and the eptible)   collected over five consecutive annual SIDERO-WT surveillance studies (from November 2014 to December 2019) conducted in North America and Europe using the recently approved (February 2021) CLSI MIC clinical breakpoints  and 90% (MIC90) of the 31,896 isolates of Enterobacterales tested from North America and Europe from 2014 to 2019 were 0.12 and 1\u2009\u03bcg/mL, respectively  isolates of Enterobacterales; 96.7% of meropenem-nonsusceptible isolates were susceptible to cefiderocol. Cefiderocol demonstrated a higher percent susceptible rate against meropenem-nonsusceptible isolates (\u226520% higher) than ceftazidime-avibactam (77.0%), cefepime (8.7%), ceftolozane-tazobactam (7.8%), and ciprofloxacin (7.8%). A total of 91.6% of 263 isolates of ceftazidime-avibactam-nonsusceptible  Enterobacterales and 97.7% of 2,658 isolates of ceftolozane-tazobactam-nonsusceptible  Enterobacterales were susceptible to cefiderocol. In comparison, only 3.8% of ceftazidime-avibactam-nonsusceptible Enterobacterales isolates were susceptible to ceftolozane-tazobactam and 90.5% of ceftolozane-tazobactam-nonsusceptible Enterobacterales isolates were susceptible to ceftazidime-avibactam. MIC90 values for colistin  and ciprofloxacin were 1 and >8\u2009\u03bcg/mL, respectively, for all isolates of Enterobacterales tested.The minimal inhibitory concentrations of cefiderocol that inhibited 50%  demonstrated a rightward shift (of 1 to 3 doubling dilutions) to higher cefiderocol MICs compared to each respective antimicrobial-susceptible subset; however, as mentioned earlier, most meropenem (96.7%)-, ceftazidime-avibactam (91.6%)-, and ceftolozane-tazobactam (97.7%)-nonsusceptible isolates remained susceptible to cefiderocol, with MICs of \u22644\u2009\u03bcg/mL.Cefiderocol MICs for meropenem-nonsusceptible , ceftazi50 and MIC90 were 0.12 and 0.5\u2009\u03bcg/mL for 7,700 isolates of P. aeruginosa collected in North America and Europe from 2014 to 2019  P. aeruginosa were 0.25 and 1\u2009\u03bcg/mL, respectively, and 99.9% of meropenem-nonsusceptible isolates were susceptible to cefiderocol. Ceftazidime-avibactam, ceftolozane-tazobactam, and cefepime were all tested with MIC90 values of 32 or >64\u2009\u03bcg/mL against isolates of meropenem-nonsusceptible P. aeruginosa and exhibited percent susceptible rates of 76.1% (ceftolozane-tazobactam), 75.0% (ceftazidime-avibactam), and 49.0% (cefepime). MIC90 values for ciprofloxacin (31.2% susceptible) and colistin were >8\u2009\u03bcg/mL and 1\u2009\u03bcg/mL, respectively, for meropenem-nonsusceptible P. aeruginosa. A total of 100% of 477 isolates of ceftazidime-avibactam-nonsusceptible  and 99.8% of 463 isolates of ceftolozane-tazobactam-nonsusceptible  P. aeruginosa, respectively, were susceptible to cefiderocol. In comparison, only 24.3% of ceftazidime-avibactam-nonsusceptible P. aeruginosa isolates were susceptible to ceftolozane-tazobactam, and only 22.0% of ceftolozane-tazobactam-nonsusceptible P. aeruginosa isolates were susceptible to ceftazidime-avibactam.The cefiderocol MIC to 2019 . CefiderP. aeruginosa (both North American and European isolates combined) demonstrated a rightward shift (of 1 doubling dilution) to higher cefiderocol MICs compared to each respective antimicrobial-susceptible subset; however, almost every nonsusceptible isolate (99.8 to 100%) remained susceptible to cefiderocol, with a MIC of \u22644\u2009\u03bcg/mL.Cefiderocol MICs for meropenem-nonsusceptible , ceftazi50 and MIC90 of cefiderocol for isolates of A. baumannii complex from both North America and Europe were 0.12 and 1\u2009\u03bcg/mL; 96.0% of isolates demonstrated cefiderocol MICs of \u22644\u2009\u03bcg/mL  and MIC90 values (1\u2009\u03bcg/mL) for the meropenem-nonsusceptible subset of isolates  and Europe (99.3 to 99.9% susceptible) varied over very narrow ranges (0.4 to 0.6%)  than for isolates from Europe (90.4 to 97.5%). In total, there were 171 isolates of A. baumannii with cefiderocol MICs of \u22658\u2009\u03bcg/mL (nonsusceptible) collected in Europe from 2014 to 2019. Of these isolates, 74.3% (127/171) were from one country (Russia); 127/437 (29.1%) of isolates from Russia were cefiderocol nonsusceptible, with annual rates of 28.2% (11/39) in 2014, 41.2% (7/17) in 2015, 24.1% (19/79) in 2016, 42.7% (50/117) in 2017, 31.9% (36/113) in 2018, and 5.6% (4/72) in 2019. Other European countries contributing >10 isolates over the study period submitted isolates with cefiderocol-nonsusceptible MICs at rates ranging from zero (no cefiderocol-nonsusceptible isolates) to 7.3% (8/109 isolates [United Kingdom]). Annual cefiderocol MIC distributions for Enterobacterales, P. aeruginosa, A. baumannii complex, S. maltophilia, and B. cepacia complex are provided in Tables S2 to S6 in the supplemental material.Annual cefiderocol percent susceptible rates for isolates of to 0.6%) . Even leEnterobacterales from North America (99.7 to 100%) and Europe (98.2 to 98.8%) were similar , while annual percent susceptible rates for ceftolozane-tazobactam were higher in isolates from North America (93.8 to 94.9%) than in those from Europe (87.3 to 90.6%) (P. aeruginosa from North America were higher for both ceftazidime-avibactam (96.0 to 99.6%) and ceftolozane-tazobactam (96.7 to 99.6%) than for isolates from Europe .Annual percent susceptible rates for ceftazidime-avibactam for isolates of o 90.6%) . Annual Enterobacterales, P. aeruginosa, A. baumannii complex, S. maltophilia, and B. cepacia complex collected in 2019 were also tested against meropenem-vaborbactam and imipenem-relebactam . Meropenem-vaborbactam demonstrated in vitro activity similar to that of ceftazidime-avibactam against Enterobacterales (98.9% of isolates susceptible); <70% of meropenem-nonsusceptible Enterobacterales isolates were susceptible to meropenem-vaborbactam and imipenem-relebactam, compared to 93.2% susceptible for cefiderocol. Imipenem-relebactam was less active (83.9% susceptible) than ceftazidime-avibactam against P. aeruginosa, compared to 99.9% susceptible for cefiderocol. Meropenem-vaborbactam and imipenem-relebactam were largely inactive in vitro against clinical isolates of A. baumannii complex  and S. maltophilia .Isolates of Enterobacterales (99.8%), P. aeruginosa (99.9%), A. baumannii complex (96.0%), and S. maltophilia (98.6%) collected across North America and Europe from 2104 to 2019 were susceptible to cefiderocol. Data in the current study confirm and expand upon data presented in earlier studies. Cefiderocol was previously reported to demonstrate potent in vitro activity against key Gram-negative pathogens  but only limited activity against Gram-positive and anaerobic bacteria , P. aeruginosa (\u226597%), and A. baumannii complex (\u226591%) isolates, as well as MDR Enterobacterales (\u226597%), P. aeruginosa (\u226597%), and A. baumannii complex (\u226590%) isolates  and P. aeruginosa (>99%), with ceftazidime-avibactam-, ceftolozane-tazobactam-, cefepime-, ciprofloxacin- and colistin-resistant phenotypes at MICs of \u22644\u2009\u03bcg/mL , S. maltophilia , and B. cepacia complex .Data in the current study clearly demonstrate that the large majority of isolates of bacteria , 10, 15.cteria 2023 and rerales 99.%, P. aercteria 20\u201314, 24 hrales 99.%, P. aerrales 99.%, P. aerEnterobacterales, P. aeruginosa, and A. baumannii have not been identified, although the addition of avibactam, a \u03b2-lactamase inhibitor, to cefiderocol has been shown to lower the MICs for some cefiderocol-resistant isolates, primarily A. baumannii possessing various ESBLs.  or PiuA (not PirA) (P. aeruginosa) can also elevate cefiderocol MICs  or Acinetobacter spp. , and the FDA has not published breakpoints for S. maltophilia , P. aeruginosa (99.9%), A. baumannii complex (96.0%), and S. maltophilia (98.6%) in North America and Europe are susceptible to cefiderocol by the recently approved CLSI MIC breakpoints  and P. aeruginosa  were negligible. Annual percent susceptible rates for A. baumannii complex demonstrated sporadic, nondirectional differences , primarily due to isolates from Russia. Annual percent susceptible rates for S. maltophilia also showed minor, nondirectional fluctuation . In vitro susceptibility testing of cefiderocol may be of benefit when cefiderocol is being considered for treatment of patients infected with carbapenem-nonsusceptible, ceftazidime-avibactam-nonsusceptible, or ceftolozane-tazobactam-nonsusceptible isolates of Enterobacterales and P. aeruginosa, carbapenem-nonsusceptible isolates of A. baumannii complex, and MDR isolates of S. maltophilia.We conclude that most current (2014 to 2019) clinical isolates of akpoints . ImportaEnterobacterales, 7,700 isolates of P. aeruginosa, 5,225 isolates of A. baumannii complex, 2,030 isolates of S. maltophilia, and 425 isolates of B. cepacia complex were collected in North America and Europe from 2014 to 2019. All isolates were shipped to IHMA , where their identities were confirmed using matrix-assisted laser desorption ionization\u2013time of flight mass (MALDI-TOF) mass spectrometry .SIDERO-WT surveillance studies, sponsored by Shionogi & Co., Ltd., , were run annually from November 2014 to December 2019. In those studies, predefined quotas of isolates of specific Gram-negative bacilli cultured from patients with intra-abdominal, urinary tract, lower respiratory tract, skin and soft tissue, or bloodstream infections were collected from clinical laboratories in North America and Europe as previously described , 15, 20.Enterobacterales, P. aeruginosa, and Acinetobacter species of \u22644\u2009\u03bcg/mL for susceptible, 8\u2009\u03bcg/mL for intermediate, and \u226516\u2009\u03bcg/mL for resistant and MIC breakpoints for S. maltophilia of \u22641\u2009\u03bcg/mL for susceptible and >1\u2009\u03bcg/mL for nonsusceptible (CLSI-defined broth microdilution susceptibility testing was performed at IHMA using custom in-house-prepared broth microdilution panels , 16, 30.ceptible ."}
+{"text": "DRIVE-FORWARD is a phase 3 trial with a completed double-blind period comparing doravirine (DOR) 100 mg with ritonavir-boosted darunavir (DRV/r) 800/100 mg, both administered with two nucleos(t)ide reverse transcriptase inhibitors , and an ongoing open-label extension. At Week (W) 48, DOR demonstrated non-inferior efficacy to DRV/r, with a superior lipid profile. Those results were sustained at W96. Here we present efficacy and safety results through W192.Participants who completed the 96-week double-blind phase and met inclusion criteria were eligible to receive open-label DOR plus two NRTIs in a 96-week extension. Efficacy and safety at W192 were assessed in two groups: participants initially randomized to DOR and maintained on DOR (n=259) and those who switched from DRV/r to DOR at W96 (n=233).3) and those switched from DRV/r (53 cells/mm3). Protocol-defined virologic failure occurred in 3.1% and 5.6% of participants maintained on DOR and switched from DRV/r, respectively, and development of genotypic resistance was low in both groups (Table 1). Discontinuation due to adverse events was also low (Table 1). Fasting LDL-cholesterol, non-HDL-cholesterol, and triglycerides showed minimal increase in participants maintained on DOR and were reduced in those switched from DRV/r to DOR (Table 1). Participants maintained on DOR had minimal weight gain after W96 (median 1 kg), and a small increase overall ; participants who switched to DOR had a small increase after W96 (median 1.5 kg), similar to the median weight gain in the base study .HIV-1 RNA < 50 copies/mL were maintained through W192 in 81.1% of participants who continued DOR and 80.7% of those who switched from DRV/r to DOR. The mean increase in CD4 T-cell counts from W96 to W192 was similar for participants maintained on DOR ViiV Healthcare  Kathleen Squires, MD, Merck (Employee) Sushma Kumar, PhD, Merck (Employee) Hong Wan, PhD, Merck (Employee) Valerie Teal, MS, Merck (Employee) Ernest Asante-Appiah, PhD, Merck (Employee) Peter Sklar, MD, Merck (Employee) Elizabeth A. Martin, DO, MPH, MBA, Merck (Employee) Rima Lahoulou, n/a, Merck (Employee)"}
+{"text": "African Americans (AA) and Latinos, compared with Whites, experience disproportionately higher rates of morbidity and mortality in COVID-19. Exuberant inflammatory responses may explain, in part, the differences in disease severity in COVID-19 observed among different demographic groups.In a retrospective cohort study, we analyzed data from patients aged \u226518 years hospitalized for COVID-19 (confirmed by positive SARS-CoV-2 PCR) from 3/1/2020 \u2013 12/31/2020 at Emory Healthcare hospitals. Patient demographics, clinical characteristics, and peak levels of high-sensitivity C-reactive protein (hs-CRP) during hospitalization were abstracted from electronic medical record. Comorbidity burden was defined as the number of six total comorbidities assessed per patient. Multivariable logistic regression  assessed the effects of race and comorbidity burden on peak hs-CRP level.3,860 patients, median age 60 [18-108] years, 51% female, 57% AA, 28% White, 6% Latino and 9% other races were enrolled. Median comorbidity burden per patient was 2 , with prevalent comorbidities distributed as follows: 68% had hypertension, 43% renal disease, 42% diabetes, 16% cardiovascular disease, 12% lung disease, and 5% cancer. Unadjusted peak hs-CRP (mg/L) levels were highest among Latino patients (144.9) followed by other races (137), AA (130.3), and Whites (122.2). In adjusted models (including race), the mean difference in peak hs-CRP (mg/L) compared with patients who had no comorbidities was 18.7 (p=0.108), 56.7 (p< 0.001), and 78.2 mg/L (p< 0.001) for 1, 2, and \u22653 comorbidities, respectively. In adjusted models (including comorbidity burden), the mean level of peak hs-CRP, compared with Whites, was 34.2 (p< 0.001), 38.4 (p=0.003), and 36.0 mg/L (p=0.06) higher in AA, Latinos, and other races, respectively.Among patients hospitalized with COVID-19, non-White race and comorbidity burden were associated with significantly higher levels of inflammation. These findings suggest that exuberant inflammatory responses may be driving, in part, the differences in COVID-19 disease severity observed across different demographic groups. Lauren F. Collins, MD, MSc, Nothing to disclose"}
+{"text": "Ceftolozane/tazobactam (C/T) is an antipseudomonal cephalosporin combined with a \u03b2-lactamase inhibitor approved by FDA and EMA for complicated urinary tract (cUTI) and complicated intraabdominal infections (cIAI), as well as hospital-acquired/ventilator-associated bacterial pneumonia (HABP/VABP) in patients \u226518 years. Clinical trials studying the use of C/T for pediatric patients with cUTI and cIAI are completed, and HABP/VABP pediatric studies are underway. We evaluated the antimicrobial activity of C/T against gram-negative isolates collected from patients \u226417 years in the United States (US) as part of the global SMART surveillance program.P. aeruginosa were screened for genes encoding \u03b2-lactamases.In 2017-2019, 27 US clinical labs each collected up to 250 consecutive gram-negative pathogens per year. A total of 1336 isolates were collected from pediatric patients. MICs were determined using CLSI broth microdilution and breakpoints. C/T-nonsusceptible Enterobacterales (Ebact) and P. aeruginosa isolates, 40.7% and 76.4%, respectively, were collected from patients with respiratory tract infections, 37.0% and 10.0% from UTI, 13.7% and 8.2% from IAI, and 8.3% and 5.0% from bloodstream infections. The table shows antimicrobial susceptibility of Ebact, P. aeruginosa, and select phenotypes. C/T was active against 98% of Ebact, including 50 of 51 and 12 of 13 ESBL-non-CRE phenotype E. coli and K. pneumoniae, respectively. Among P. aeruginosa, C/T was active against 95% of isolates, 9-21 percentage points higher than the comparator \u03b2-lactams, and it maintained activity against 71-75% of P. aeruginosa isolates nonsusceptible to commonly used \u03b2-lactams. Among the 21 C/T-nonsusceptible Ebact, 2 isolates carried KPC and ESBL and 3 isolates carried only ESBL; in 16 isolates no \u03b2-lactamases were detected, of which 15 were species with intrinsic AmpC. Among 12 C/T-nonsusceptible P. aeruginosa, no acquired \u03b2-lactamases were detected, likely indicating chromosomally-mediated resistance mechanisms.Among the 944 collected Ebact and 220 Results TableP. aeruginosa, including resistant phenotypes.C/T could be a potential new treatment option for US pediatric patients with infections caused by Ebact and Sibylle Lob, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Meredith Hackel, PhD MPH, IHMA (Employee)Pfizer, Inc. (Independent Contractor) C. Andrew DeRyke, PharmD, Merck & Co., Inc.  Kelly Harris, PharmD, BCPS, Merck & Co. Inc (Employee) Katherine Young, MS, Merck (Employee) Mary Motyl, PhD, Merck & Co., Inc.  Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "Dear Editors,We and others have shown that kidney transplant recipients (KTR) exhibit a reduced immune response with a seroconversion (SC) rate <50% after a regular 2-dose mRNA SARS-CoV-2 vaccination regimen , 2. Very\u00ae SARS-CoV-2-TrimericS IgG chemiluminescent immunoassay, Diasorin S.p.A., Saluggia, Italy; cut-off value for seroconversion: \u226533.8\u00a0BAU/mL). After double mRNA vaccination the SC rate was 48%. Following heterologous triple vaccination an additional 54% of the initial non-responders achieved SC. Altogether, 97 out of 122 KTR (80%) achieved SC after either double mRNA vaccination or the heterologous triple vaccination. Forty-eight of the 142 KTR showed high-level SC after double mRNA vaccination. Twenty patients developed low antibody concentrations (arbitrary threshold <350\u00a0BAU/mL). After a third heterologous dose all these 20 patients significantly boosted their humoral response (1391.9 (SD 687.2) vs. 144.8 (SD 94.6) BAU/mL, p < 0.001). Non-responders after heterologous triple vaccination were significantly older , were more often treated with prednisolone or belatacept  and had a shorter median transplantation vintage . They showed a trend of lower mean eGFR  and being treated more often with mycophenolic acid . Higher mycophenolic acid doses did not correlate with inferior antibody response (p = 0.299). As a limitation, our study lacks cellular immune response and neutralizing antibody data. But anti-spike IgG antibodies are highly correlated with neutralizing antibodies, and a level >264\u00a0BAU/mL  has been found to be associated with 80% vaccine efficacy against primary symptomatic Covid-19, although limited to the B.1.177 and B.1.1.7 SARS-CoV-2 variant (vs. 4.6\u00a0years) and extended interval between second and third dose (109 vs. 80\u00a0days) in our cohort compared to the study by Reindl-Schwaighofer et al.  after double mRNA and triple heterologous vaccination . Patient"}
+{"text": "While the second wave of COVID-19 pandemic almost reached its climax, unfortunately, new viral strains are rapidly spreading, and numbers of infected young adults are rising. Consequently, chest high-resolution computed tomography (HRCT) demands are increasing, regarding patients\u2019 screening, initial evaluation and follow up. This study aims to evaluate the detection accuracy of ultra-low-dose chest CT in comparison with the routine low-dose chest CT to reduce the irradiation exposure hazards.-4.This study was prospectively conducted on 250 patients during the period from 15th December 2020 to 10th February 2021. All of the included patients were clinically suspected of COVID-19 infection. All patients were subjected to routine low-dose (45 mAs) and ultra-low-dose (22 mAs) chest CT examinations. Finally, all patients had confirmatory PCR swab tests and other dedicated laboratory tests. They included 149 males and 101 females (59.6%:40.4%). Their age ranged from 16 to 84 years (mean age 50 \u00b1 34 SD). Patients were divided according to body weight; 104 patients were less than 80 kg, and 146 patients were more than 80 kg. HRCT findings were examined by two expert consultant radiologists independently, and data analysis was performed by other two expert specialist and consultant radiologists. The inter-observer agreement (IOA) was excellent (96\u2013100%). The ultra-low-dose chest CT reached 93.53\u201396.84% sensitivity and 90.38\u201393.84% accuracy. The signal-to-noise ratio (SNR) is 12.8:16.1; CTDIvol (mGy) = 1.1 \u00b1 0.3, DLP (mGy cm) = 42.2 \u00b1 7.9, mean effective dose (mSv/mGy cm) = 0.59 and absolute cancer risk = 0.02 \u00d7 10Ultra-low-dose HRCT can be reliably used during the second wave of COVID-19 pandemic to reduce the irradiation exposure hazards. Real-time polymerase chain reaction (PCR) remains the gold standard tool for the diagnosis of the Novel coronavirus disease COVID-19) since it was first described in December 2019 and announced as a pandemic in February 2020 9 since i, 4.On the other hand, high-resolution computed tomography (HRCT) expressed more availability and more rapid results. Besides, HRCT sensitivity exceeded 90% in comparison with PCR sensitivity which could not exceed 75% \u20138. Low-dIn September 2019, the American Association of Physicists in Medicine (AAPM) suggested certain criteria for efficient low-dose CT during lung cancer screening among standard-sized persons (70\u201390 kg), including CTDIvol \u2264 3.0 mGy, DLP \u2264 75 mGy cm and effective dose \u2264 1.0 mSv . A dose While the second wave of COVID-19 pandemic almost reached its climax, unfortunately, new viral strains are rapidly spreading, and numbers of infected young adults are rapidly rising. Hence, chest HRCT demands are increasing.This study aims to evaluate the detection accuracy of ultra-low-dose chest CT in comparison with the routine low-dose chest CT during the assessment of COVID-19 patients in order to reduce the irradiation exposure hazards.This study was prospectively conducted on 250 patients who were clinically suspected of COVID-19 infection during the period from 15th December 2020 to 10th February 2021. The study was approved by the Ethics Committee of our University hospital. Patient verbal consent was accepted by the Ethics Committee respecting absolute safety of non-invasive non-therapeutic procedure without additional personal risk or burden to the public health, also assuring full respect of both patient and medical record confidentiality. The authors emphasize on the fact that the overall radiation dose in both low- and ultra-low-dose CT examinations together remains much lower than single routine chest CT examination.Patients included 149 males and 101 females (59.6%:40.4%). Their age ranged from 16 to 84 years (mean age 50 \u00b1 34 SD). By definition of the American Association of Physicists in Medicine (AAPM), the standard bodyweight for an adult ranges from 70 to 90 kg  [2) [ of air] . The staair] [2) . By defiair] [2) .Fig. 1IThe prevalence rate of each HRCT finding (isolated or mixed) was estimated as the percentage of patients with each abnormal CT finding. This was done by each observer for both groups of patients regarding both low-dose and ultra-low-dose CT examinations.Similarly, the prevalence rate of the maximum attenuation of the pathological lesions was also estimated.Cohen\u2019s Kappa test was utilized to calculate the \u201cInter-Observer Agreement\u201d (IOA) coefficient regarding each HRCT characteristic and lung attenuation.https://www.medcalc.org/calc/diagnostic_test.php) was utilized to detect the mean value and 95% confidence interval (95% CI) regarding the sensitivity, specificity, positive predictive value (PPV), negative predictive value, positive likelihood ratio, negative likelihood ratio, and accuracy of both low-dose and ultra-low-dose CT protocols.An online calculator  in the ultra-low-dose CT was lower than that in the low-dose CT (12.8\u201316.1 compared with 17.5\u201324.4 respectively). This almost did not impact the diagnostic efficacy of CT as detailed later on.Ground-glass opacities were the most common findings among our patients. They were found alone in 173 patients (69.2%). They were found also surrounding a solid nodule  among 49 patients (19.6%) and associated with interlobular septal thickening (crazy paving pattern) also among 49 patients (19.6%). Consolidative changes with or without ground-glass opacities were detected among 84 patients (33.6%). The CT attenuation of these pathological CT findings ranged from + 40 to \u2212 750 HU. The predominant CT attenuation among these CT findings ranged from \u2013 300 to \u2212 400 HU (157 patients/62.8%).In comparison with the low-dose CT, the ultra-low-dose CT protocol similarly detected all HRCT findings of COVID-19 , as well as their ranges of CT attenuation (+ 40 to \u2212 750 HU). The inter-observer agreement was excellent (100%) Figs.  and 3.FIn comparison with the low-dose CT, the ultra-low-dose CT protocol only showed lower efficacy regarding detection of small GG nodules < 1 cm and nodules with ground-glass halo sign (ranging from 77 to 94% and 91 to 96% among both observers respectively). The CT attenuation of these lesions ranged from \u2212 300 to \u2212 600 HU. Otherwise, no absolute differences were found regarding the detection of the rest of the HRCT findings and their ranges of CT attenuation. Finally, the inter-observer agreement was collectively excellent (96\u2013100%) Figs.  and 6.FA multi-compartmental flow chart Fig.  is summaCollectively the ultra-low-dose chest CT reached 93.53\u201396.84% sensitivity and 90.38\u201393.84% accuracy in comparison with the low-dose CT which showed 96.84\u201397.84% sensitivity and 90.38\u201395.21% accuracy.No differences were found between both observers at both low-dose and ultra-low-dose CT protocols regarding the final diagnosis. Ninety-two patients (88.5%) were truly positive and confirmed by PCR to have COVID-19. On the other hand, only 3 patients (3%) were false negative. Consequently, both CT protocols showed same statistical results: sensitivity (mean = 96.84% and CI 95% = 91.05 to 99.34%) and accuracy (mean = 90.38% and CI 95% = 83.03 to 95.29%).CT specificity was equally low (22.22%) for both low-dose and ultra-low-dose CT as 7/99 patients showed false-positive CT results with negative PCR swab tests and alternative diagnoses .Observer 1 truly detected 130 patients using ultra-low-dose CT compared with 136 patients using low-dose CT. Six cases were furtherly missed using the ultra-lose-dose CT. Consequently the low-dose CT showed sensitivity (mean = 97.84% and CI 95% = 93.82 to 99.55%) and accuracy (mean = 95.21% and CI 95% = 90.37 to 98.05%), while the ultra-low-dose CT showed sensitivity (mean = 93.53% and CI 95% = 88.06 to 97.00%) and accuracy (mean = 91.10% and CI 95% = 85.26 to 95.17%).Meanwhile, observer 2 truly detected 134 patients using ultra-low-dose CT compared with 136 patients using low-dose CT, while only two cases were furtherly missed using the ultra-low-dose CT. Consequently the low-dose CT showed sensitivity (mean = 97.84% and CI 95% = 93.82 to 99.55%) and accuracy (mean = 95.21% and CI 95% = 90.37 to 98.05%), while the ultra-low-dose CT showed sensitivity (mean = 96.40% and CI 95% = 91.81 to 98.82%) and accuracy (mean= 93.84% and CI 95%= 88.62 to 97.14%).CT specificity was equally low (42.86%) for both low-dose and ultra-low-dose CT as 4/146 patients showed false-positive CT results with negative PCR swab tests and alternative diagnoses .Comparison with other CT protocols is demonstrated in Table The second wave and even further crisis of COVID-19 pandemic are striking the world with increased demands for chest CT imaging .Similar to Chung et al.  and Song-4. These estimated radiation doses are 12.5:13.3 and 18.1:20 times more than that of the low-dose and the ultra-low-dose CT protocols in this study respectively. Additionally, the DLP and absolute cancer risk of the ultra-low-dose CT in this study were lower than those provided by Tabatabaei et al. [According to Kubo et al. , the DLPi et al.  and Bahri et al.   and iterative dose reduction (IDR).Ultra-low-dose HRCT can be reliably used during the second wave of COVID-19 pandemic to reduce irradiation exposure hazards."}
+{"text": "Scientific Reportshttps://doi.org/10.1038/s41598-019-51090-3, published online 10 October 2019Correction to: The original version of this Article omitted Supplementary Table S1, Supplementary Table S2, and Supplementary Table S3.These Supplementary Information files now accompany the original Article."}
+{"text": "The prognostic value of bile duct invasion (BDI) remains controversial. We aimed to investigate the prognostic value of BDI and the stage of BDI in different staging systems.Patients with hepatocellular carcinoma (HCC) from nine hepatobiliary medical centers who underwent R0 resection were included. Overall survival (OS) was assessed using the Kaplan\u2013Meier method and tested using the log-rank test. The prognostic effect of BDI was analyzed using univariate and multivariate Cox proportional hazard regression analyses. The predictive performance of these models was evaluated using the concordance index and time-dependent receiver operating characteristic curve (tdAUC).Of 1021 patients with HCC, 177 had BDI. OS was worse in the HCC with BDI group than in the HCC without BDI group (p<0.001); multivariate analysis identified BDI as an independent risk factor for OS. After adjustment for interference of confounding factors using the Cox proportional hazard regression model, HCC with BDI and without macrovascular invasion was classified as Barcelona Clinic Liver Cancer (BCLC) B, eighth edition American Joint Committee on Cancer (AJCC) IIIA, and China Liver Cancer (CNLC) IIb, respectively, whereas HCC with BDI and macrovascular was classified as BCLC C, AJCC IIIB, and CNLC IIIA, respectively. C-indexes and tdAUCs of the adjusted staging systems were superior to those of the corresponding current staging systems.We constructed adjusted staging systems with the BDI status, improved their predictive performance and facilitate clinical use. Hepatocellular carcinoma (HCC) is the sixth common cancer and fourth cancer-specific cause of death globally . HCC is th AJCC) staging system .HCC patients with microscopic or macroscopic BDI admitted to one of the nine Chinese hepatobiliary medical centers  between March 1, 2007 and March 1, 2018 were included. HCC patients without BDI were from the Mengchao Hepatobiliary Hospital of Fujian Medical University and Eastern Hepatobiliary Surgery Hospital of Second Military Medical University during the same period.The inclusion criteria were as follows: 1) both HCC and BDI were histopathologically confirmed, 2) tumors were treated with R0 resection, and 3) complete clinical data and postoperative follow-up records. The exclusion criteria were as follows: 1)\u00a0recurrent or metastatic HCC, 2) combined HCC-intrahepatic cholangiocarcinoma, 3) other accompanying cancers, and 4) clinical data or survival data are missing. R0 resection was defined as the removal of all macroscopic tumors with a microscopically negative margin. The decision of treatment depends on the discussion of the multidisciplinary team in each center, whether perform surgical resection mainly considering the liver function, residual liver volume, tumor related factors such as if complicated with portal vein main trunk tumor thrombus or vena cava tumor thrombus or distant metastasis, and whether tumor and tumor thrombus can be completely removed.All patients were regularly followed up after discharge from the hospital. Follow-up visits were scheduled once every 2\u20133 months in the first 2 years, once every 6 months from 2 to 5 years, and once every year after 5 years. Follow-up examinations were conducted using laboratory tests , abdominal ultrasonography, and/or contrast-enhanced computed tomography magnetic resonance imaging. Overall survival (OS) was defined as the time of resection to the date of either death or the latest follow-up. Data including baseline and clinical characteristics and follow-up information were extracted and censored on September 31, 2020.via clinical diagnosis. PS (performance status) score refer to ECOG  PS score. Tumor differentiation was classified according to the Edmonson\u2013Steiner grade. MVI was vascular invasion of small vessels only identifiable histologically. Major vascular invasion was defined as invasion of the branches of the main portal vein , one or more of the three hepatic veins , or the main branches of the proper hepatic artery (right or left hepatic artery) , major vascular invasion, macrovascular invasion (MaVI), preoperative serum gamma-glutamyltransferase (GGT), alkaline phosphatase (ALP), total bilirubin (TBil), AFP, Child\u2013Pugh grade, MELD (model for end-stage liver disease) score, BCLC stage, AJCC stage, and CNLC stage. Underlying liver diseases were categorized as viral liver disease (hepatitis B virus or hepatitis C virus), non-alcoholic fatty liver disease, alcoholic liver disease, and no underlying liver disease. Cirrhosis was confirmed histopathologically or  artery) . Major vhttp://www.r-project.org/); the R packages of \u201creadxl,\u201d \u201ctable1,\u201d \u201crms,\u201d \u201csurvminer,\u201d \u201csurvival,\u201d \u201cggplot2,\u201d \u201cCsChange,\u201d and \u201ctimeROC\u201d were used.Continuous variables are reported as medians with interquartile ranges and were compared using Student\u2019s t-test or the Mann\u2013Whitney test. Categorical data, presented as frequencies (%), were compared using the chi-square test or Fisher\u2019s exact test. Kaplan\u2013Meier survival curves were used to assess OS. Univariate and multivariate analyses were performed using the Cox proportional hazard regression model, and a backward stepwise selection method was used to identify independent prognostic factors and adjust for confounding factors. The Harrell\u2019s concordance index (C-index)  and timeOf 1021 patients with HCC who were included, flowchart was shown in The median follow-up time of the whole cohort was 49.03 months . The median OS times (mOST) of patients without BDI and with BDI were 59.27 months (95%CI 50.7-67.17 months) and 23.3 months (95%CI 19.67-26.23 months), respectively (P<0.001) Figure\u00a01The respective mOST of the BDI- and BDI+ groups were 76.87 months (95%CI 76.87- Not Available [NA] months) and 28.98 months (95%CI 18.53-NA months) in BCLC stage 0 (P=0.05), 75.53 months (95%CI 70.37-NA months) and 26.73 months (95%CI 20.27-35.9 months) in BCLC stage A (P<0.001), 40.33 months (95%CI 27.43-67.6 months) and 26.2 months (95%CI 23.07-NA months) in BCLC stage B (P=0.12), and 16.2 months (95%CI 12.67-20.87 months) and 16.3 months (95%CI 13.17-22.73 months) in BCLC stage C (P=0.45) D.The respective mOST of the BDI- and BDI+ groups were 76.87 months (95%CI 73.53-NA months) and 28.98 months (95%CI 23.43-NA months) in AJCC stage IA (P=0.03), 75.53 months (95%CI 70.07-NA months) and 31.8 months (95%CI 25.44-NA months) in AJCC stage IB (P<0.001), 44.97 months (95%CI 29.7-57.9 months) and 23.07 months (95%CI 19.67-26.53 months) in AJCC stage II (P<0.001), 26.93 months (95%CI 21.4-31.97 months) and 26.21 months (95%CI 12.6-NA months) in AJCC stage IIIA (P=0.35), 9.76 months (95%CI 7.57-12.63 months) and 10.23 months (95%CI 6.2-16.36 months) in AJCC stage IIIB (P=0.73) I.The mOST of the BDI- and BDI+ groups were 80.83 months (95%CI 76.87-NA months) and 28.98 months (95%CI 20.2-NA months) in CNLC stage Ia (P<0.001), 52.6 months (95%CI 48.27-67.13 months) and 24.17 months (95%CI 19.67-30.97 months) in CNLC stage Ib (P<0.001), 30.47 months (95%CI 22.93-NA months)and 26.53 months (95%CI 17.07-NA months) in CNLC stage IIa (P=0.43), 26.2 months (95%CI 19.0-40.33 months) and 24.77 months (95%CI 7.57-NA months) in AJCC stage IIIA (P=0.6), 11.77 months (95%CI 9.7-17.7 months) and 13.96 months (95%CI 7.54-16.37 months) in AJCC stage IIIB (P=0.27) N.OS curves stratified by different stages obtained from the multivariate Cox proportional hazard regression model adjusted for other covariates are shown in The BDI+ group was restaged according to the HR of each stage. BCLC stage 0/A/B BDI+ was integrated into BDI+MaVI- and classified as BCLC stage B, and BCLC stage C BDI+ was classified as BCLC stage C , Key Clinical Specialty Discipline Construction Program of Fuzhou, Fujian, P.R.C (Grant number: 201912002) and Fujian Provincial Medical Center of Hepatobiliary.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Patients with lymphoid malignancies are at high risk of severe COVID-19 disease and were not included in the phase 3 mRNA vaccine trials. Many patients with lymphoid malignancies receive immunosuppressive therapies, including B-cell depleting agents, that may negatively impact humoral response to vaccination.We recruited patients with lymphoid malignancies and healthy participants who planned to receive two doses of SARS-CoV-2 mRNA vaccine (BNT162b2 or mRNA-1273). Blood was drawn at baseline, prior to second dose of vaccine, and 28 days after last vaccination. Disease characteristics and therapies were extracted from patients\u2019 electronic medical record. An ultrasensitive, single molecule array (Simoa) assay detected anti-Spike (S), anti-S1, anti-receptor binding domain (RBD), and anti-Nucleocapsid (N) IgG from plasma at each timepoint.23 healthy participants and 37 patients with lymphoid malignancies were enrolled (Table 1). Low titers of anti-N  demonstrate no prior exposure or acquisition of COVID-19 before vaccination or during the study. 37.8% of the lymphoid malignancy cohort responded to the vaccine, using an internally validated AEB cutoff of 1.07. A significantly higher magnitude of anti-S (p< 0.0001), anti-S1 (p< 0.0001) and anti-RBD (p< 0.0001) are present in the healthy as compared to lymphoid malignancy cohort at the second dose and day 28 post-series . Anti-S IgG titers were compared between the healthy cohort, treatment na\u00efve, and treatment experienced groups . The treatment na\u00efve cohort had high titers by series completion which were not significantly different from the healthy cohort (p=0.2259), although the treatment experienced group had significantly decreased titers (p< 0.0001). Of the 20 patients who had received CD20 therapy, there was no clear correlation of anti-S IgG response with time from CD20 therapy, although most patients who received CD20 therapies within 12 months from the vaccine had no response . Table 1. DemographicsFigure 1. Anti-N, Anti-S, Anti-S1, Anti-RBD and Anti-N Ig G for healthy v. lymphoid malignancy cohortThe dotted line at 1.07 marks in an internally validated threshold to mark anti-S IgG response. The black bars denote median with 95% CI.Figure 2: Anti-S IgG for healthy v. treatment na\u00efve v. treatment experiencedThe dotted line at 1.07 marks in an internally validated threshold to mark antibody response. The black bars denote median with 95% CI.The vaccine-induced immune response was poor among treatment-experienced patients with lymphoid malignancies, especially among those who received CD20 therapies within 12 months.Figure 3. Months from CD20 therapy v. anti-S IgG titersThe dotted line at 1.07 marks in an internally validated threshold to mark antibody response.Jennifer Crombie, MD, AbbVie (Grant/Research Support)Bauer (Grant/Research Support)Karyopharm (Consultant)MorphoSys (Consultant) Philippe Armand, MD PhD, ADCT, Celgene, Morphosys, Daiichi, Miltenyi, Tessa, C4, Genmab, Enterome, Regeneron, Genentech, Epizyme, Astra Zeneca Affimed, Adaptive, BMS, Merck, Kite, IGM, Genentech  David Walt, PhD, Quanterix Corporation  Nicolas C. Issa, MD, AiCuris (Scientific Research Study Investigator)Astellas (Scientific Research Study Investigator)GSK (Scientific Research Study Investigator)Merck (Scientific Research Study Investigator)"}
+{"text": "The 2017 IDSA CDI guideline update phased out metronidazole (MTZ) and recommended vancomycin (VAN) or fidaxomicin (FDX) for first-line use. This study examined changes in CDI antibiotic use and clinical outcomes among Medicare beneficiaries with CDI pre- vs. post- the guideline update.This retrospective claims analysis used 2016-2018 national Medicare claims data. The two study samples included continuously eligible fee-for-service Medicare beneficiaries aged \u226566 years with a new CDI diagnosis followed by an antibiotic fill in the pre-period (04/01/2017-09/30/2017) and post-period (04/01/2018-09/30/2018), respectively. Outcomes included type of CDI antibiotic received; sustained response and CDI recurrence. Multivariable regressions compared pre- vs. post-period outcomes while controlling for sociodemographic and clinical factors.The pre-period  and post-period  samples had similar characteristics . Post-guideline update, absolute rates of MTZ use declined 27.7%  and VAN use increased 26.9%  . While FDX use increased 0.8% , overall use remained low (1.63%). Surprisingly, clinical outcomes did not improve between the pre- and post-period (Table 1). Even after adjustment, overall sustained response rates decreased  and overall CDI recurrence rates increased  slightly in the post- vs. pre-period. Additional analyses by type of antibiotic showed that VAN (55.0% and 35.1%) was similar in outcomes to MTZ (54.2% and 33.0%), whereas FDX (71.4% and 20.9%) had higher sustained response and lower CDI recurrence rates, respectively .Figure 1. First-line use of CDI treatments, pre- vs. post- the guideline update, among Medicare beneficiaries with CDITable 1. Clinical outcomes, pre- vs. post- the guideline update, among Medicare beneficiaries with CDIFigure 2. Clinical outcomes* by type of index CDI treatment among Medicare beneficiaries with CDIPooled rates among patients on each index CDI treatment across the pre- and post-index periods.The 2017 IDSA guideline update was associated with a substantial increase in VAN use and decrease in MTZ use. FDX use rates remained low (< 2%). Overall CDI outcomes did not improve post guideline update despite the shift to guideline-indicated VAN. This may be because VAN was not associated with meaningfully improved outcomes relative to MTZ. However, improved outcomes seen with FDX relative to VAN and MTZ suggest potential benefits from its greater use in Medicare patients.Erik R. Dubberke, MD, MSPH, Ferring (Grant/Research Support)Merck (Consultant)Pfizer Seres (Consultant)Summit (Consultant) Justin T. Puckett, BA, COVIA Health Solutions (Employee) Engels N. Obi, PhD, Merck & Co.  Sachin Kamal-Bahl, PhD, AbbVie (Consultant)Arena Pharmaceuticals, Inc. (Consultant)COVIA Health Solutions (Employee)Janssen, Inc. (Consultant)Merck Novartis (Consultant)Pfizer, Inc. PhRMA (Consultant) Kaushal Desai, PhD, AstraZeneca Pharmaceuticals (Shareholder)Merck & Co. Inc. (Employee) Bruce Stuart, PhD, COVIA Health Solutions (Consultant) Jalpa A. Doshi, PhD, Acadia Allergan (Advisor or Review Panel member)Biogen (Grant/Research Support)Boehringer Ingelheim Catabasis (Consultant)Humana (Grant/Research Support)Janssen, Inc. MeiraGTX (Consultant)Merck Novartis (Grant/Research Support)Otsuka (Advisor or Review Panel member)Regeneron (Grant/Research Support)SAGE Therapeutics (Consultant)Sanofi (Grant/Research Support)Shire (Advisor or Review Panel member)The Medicines Company (Advisor or Review Panel member)"}
+{"text": "Here, we present 36 metagenomes, 59 metatranscriptomes, and 373 metagenome-assembled genomes (MAGs) from Chesapeake and Delaware Bay water samples. This data set will be useful for studying microbial biogeochemical cycling in estuaries. Sharp. Surface (\u223c1.5 m below the seafloor [mbsf]) water samples were collected using a rosette sampler with associated conductivity-temperature-depth (CTD) profiles. The sampling scheme (\u2013\u2013https://www.bco-dmo.org/dataset/565451).Estuaries are productive aquatic environments harboring diverse flora, fauna, and microbial communities important for global carbon and nutrient cycling , 2. Omicg scheme , environg scheme \u20137 were d scheme \u2013\u20137 and arhttps://doi.org/10.6084/m9.figshare.14173664 (metatranscriptomes).Cells were separated as large- and small-cell-size fractions by passing water samples through 0.8- and 0.22-\u03bcm-pore-size filters. Nucleic acids were extracted from size-fractionated cells using the Allprep DNA/RNA minikit  . A totalPrior to assembly, Cutadapt v1.11 and Sickle v1.33 were used to remove adapters from and quality trim (Q\u2009=\u200930) Illumina-sequenced reads . Nanoporhttps://doi.org/10.6084/m9.figshare.14179448) with >80% completion and <5% contamination were taxonomically annotated using Anvi\u2019o and Microbial Genome Atlas (MiGA) v0.7.26.2 , Burkholderiales (n = 31), Flavobacteriales (n = 55), Microtrichales (n = 39), Nanopelagicales (n = 11), Pelagibacterales (n = 31), Pseudomonadales (n = 26), Rhodobacterales (n = 28), and Synechococcales (n = 13), as well as the archaeal phyla Crenarchaeota (n = 5) and Euryarchaeota (n = 2).For binning, trimmed reads from each Illumina-sequenced library were mapped to contigs \u22652,000\u2009bp in the corresponding metagenome using the default parameters (end-to-end mode) of Bowtie 2 v2.2.7 . Alignme17\u20130.7.26.2 . They bePRJNA432171.The metagenomes, metatranscriptomes, and MAGs are available on NCBI under the umbrella project"}
+{"text": "Complicated urinary tract infections (cUTI) are one of the most common bacterial infections and represent substantial burden to the health care system. Here, we examine the epidemiology and treatment patterns associated with cUTI in a large US database containing longitudinal inpatient (IP) and outpatient (OP) patient-level data.We conducted a retrospective cohort study of adult patients in the IBM MarketScan\u00ae Commercial or Medicare Supplemental Databases with at least 1 IP or non-diagnostic OP claim with a diagnosis for cUTI between January 1, 2017 and June 30, 2019. Patients meeting the following criteria were included for analysis: (1) \u226518 years of age on the index date, (2) \u22656 months of continuous enrollment (CE) with medical and pharmacy benefits prior to the index date, (3) \u226512 months of CE following the index date or evidence of death, and (4) no evidence of a prior cUTI during the 6-month baseline period. Demographics and clinical characteristics were quantified. Patients were classified as IP if they were hospitalized during 30-day post index date; remaining patients were classified as OP. Antibiotics received in the OP setting in the 12-months post index date were examined. 95,423 patients met study criteria. Most (86.4%) patients were Commercially insured, mean (SD) age was 53.6 (18.1) and 70.4% were female. Mean baseline Charlson Comorbidity Index was 0.77. During the 30-day post index date, 22.2% were treated as IP and 77.8% were strictly treated as OP. In the 12-month OP follow-up period among index IP, 78.2% required \u2265 2 antibiotics, 38.2% required \u22654 antibiotics, and 41.6% received an IV antibiotic. In the 12-month OP follow-up period among index OP, 81.8% required \u22652 antibiotics, 38.2% required \u22654 antibiotics, and 46.8% received an IV antibiotic. For both IP and OP, fluoroquinolones were the most common oral antibiotic class (57.7%), followed by cephalosporins (39.2%), penicillins (30.3%), trimethoprim-sulfamethoxazole (29.8%), and nitrofurantoin (25.2%). Cephalosporins were the most common IV antibiotic class (38.5%).Regardless of index treatment setting, approximately 40% of all cUTI patients required \u22654 antibiotic therapy and almost half with receive an IV antibiotic in the outpatient setting in the 12-months post index date.Thomas Lodise, Jr., PharmD, PhD, Astra-Zeneca (Consultant)Bayer (Consultant)DoseMe ferring (Consultant)genentech (Consultant)GSK (Consultant)Melinta (Consultant)merck nabriva (Consultant)paratek shionogi Spero (Consultant)tetraphase (Consultant)Venatrox (Consultant) Thomas Lodise, Jr., PharmD, PhD, Melinta Therapeutics  Involved: Self): Consultant; Merck  Involved: Self): Consultant, Scientific Research Study Investigator; Paratek  Involved: Self): Consultant; Shionogi  Involved: Self): Consultant, Speakers\u2019 bureau; Spero  Involved: Self): Consultant; Tetraphase Pharmaceuticals Inc.  Involved: Self): Consultant Janna Manjelievskaia, PhD, MPH, IBM Watson Health (Employee)Spero Therapeutics (Consultant) Matthew Brouillette, MPH, Spero Therapeutics, Inc.  Kate Sulham, MPH, Spero Therapeutics (Consultant)"}
+{"text": "Communications Biology 10.1038/s42003-021-02971-9, published online 10 January 2022.Correction to: The original version of this Article contained errors in Fig. 1 and Supplementary Fig.\u00a0Updated Supplementary Information"}
+{"text": "Early bacterial co-infection is rare in hospitalized COVID-19 patients, yet antibiotics are commonly prescribed. Antibiotic stewardship (AS) intervention is needed, especially in small community hospitals (SCHs), which often lack access to AS expertise.We implemented daily remote multidisciplinary tele-COVID rounds (synchronous case review between SCH providers and ID clinicians) and tele-stewardship surveillance (ID pharmacist review of COVID patients on antibiotics) on 6/24/2020 in 17 SCHs. We retrospectively included adult symptomatic COVID-19 admissions between 3/2020 and 4/2021. The primary outcome was early use of antibiotics for pneumonia (started within 48 hours of admission); mean monthly days of therapy per 1,000 patient days (DOT) were compared pre- (3/2020-6/2020) and post-intervention (7/2020-4/2021). Secondary outcomes were early use of antibiotics for any indication, estimated days of antibiotics avoided (comparing pre- and post-intervention DOT), and in-hospital mortality. Analyses were conducted using a two-tailed unpaired t-test (antibiotic use) or Fisher\u2019s exact test .Of the 1,976 patients included (124 pre- vs. 1852 post-intervention), 55.4% were male and 85.5% were white. Patients in the pre-intervention group were more likely to require hospital transfer [21.8% vs 8.8% (p< 0.001)] and ICU admission [18.5% vs. 9.7% (p=0.003)]. We observed a significant decrease in mean use of early antibiotics for pneumonia [656.9 vs. 240.1 DOT (p< 0.001)], including among non-ICU patients only [603.6 vs 240.2 DOT (p< 0.001)]. Early antibiotic use for any indication also decreased [686.2 vs. 359.3 DOT (p< 0.001)]. An estimated 3,697 days of unnecessary antibiotics for pneumonia were avoided in the 10-months post-intervention [370 days per month (95% CI 304 \u2013 435)]. Unadjusted in-hospital mortality was not different pre- vs post-intervention , but was higher among those prescribed early antibiotics .A significant, sustained reduction in antibiotic use among COVID-19 patients at 17 SCHs was observed after implementation of tele-COVID rounds and tele-stewardship surveillance without an observed difference in mortality.All Authors: No reported disclosures"}
+{"text": "In Cohort A, baseline NT-proBNP > 3000 ng/L and cTnT-HS > 50 ng/L and a relative increase of NT-proBNP > 50% during follow-up were independent prognostic factors of EFS. The slopes of logs NT-proBNP and cTnT-HS increased with time before and stabilized after tafamidis. Conclusion: ATTR-CA patients with increasing NT-proBNP had an increased risk of EFS. Tafamidis stabilize NT-proBNP and cTnT-HS increasing, even if initial NT-proBNP levels were >3000 ng/L. Thus suggesting that all patients, irrespective of baseline NT-proBNP levels, may benefit from tafamidis.Background: We assesse the evolution and prognostic value of N-terminal pro-brain natriuretic peptide (NT-proBNP) and high-sensitivity troponin T (cTnT-HS) in transthyretin amyloid cardiomyopathy (ATTR-CA) before and after tafamidis treatment. Methods and Results: 454 ATTR-CA patients without tafamidis (Cohort A) and 248 ATTR-CA with tafamidis (Cohort B) were enrolled. Event-free survival (EFS) events were death, heart transplant, or acute heart failure. In Cohort A, 27% of patients maintained NT-proBNP < 3000 ng/L and 14% cTnT-HS < 50 ng/L at 12 months relative to baseline levels. In Cohort B, the proportions were 49% and 29%, respectively. In Cohort A, among the 333 patients without an increased NT-proBNP > 50% relative to baseline EFS was extended compared to the 121 patients with an increased NT-proBNP > 50% (HR: 0.75 ;  Transthyretin amyloid cardiomyopathy (ATTR-CA) is a systemic life-threatening disease characterized by amyloid fibrils accumulating in the heart . There aIn the blood, TTR, mainly produced in the liver, transports the retinol-binding protein-vitamin A complex and to a lesser extent thyroxine. However, when TTR tetramer dissociates and the monomers misfolds TTR aggregates resulting in amyloidogenesis. Mutation of TTR (in ATTRv-CA) or aging (in ATTRwt-CA) destabilizers the tetramers and accelerate their dissociation into monomers. The TTR amyloid fibrils formed by the misfolding deposit in several tissues and organs in the body.Prior to the development of TTR stabilizers, ATTRv was treated by liver transplantation: replacing mutant TTR, in the liver, with wild-type TTR . CurrentThe phase III, double-blinded, placebo-controlled ATTR-ACT trial NCT01994889) assessed the benefit of tafamidis in ATTR patients. The all-cause mortality after 30 months was significantly lower in patients treated with tafamidis (30%) compared to those treated with placebo (43%), hazard ratio (HR): 0.70 (95% CI: 0.51\u20130.96)  (p < 0.0001)  and log (cTnT-HS) levels during follow up are shown in  0.0001) .n = 192/385) after 6 months, 30.6% (45/147) after 12 months, and 23.4% (30/128) after 18 months  and 14.5 months (95% CI: 10.9\u201319.0) in those with non-increased NT-proBNP (n = 333), HR: 0.75 (95% CI: 0.57\u20130.98); p = 0.032 (log-rank test) (p = 0.09) and 0.78 , respectively (figure not shown).At the cut-off date and after a median follow-up of 14.2 months (IQR: 3.1\u201340.4), 234 patients (52%) had reported EFS events (25 deaths and 209 acute heart failure). Among the 209 acute heart failures, 33 patients died after acute heart failure for a total number of deaths of 58 patients. 220 patients have not reported an EFS event: 176 had initiated tafamidis including 28 patients enrolled in the ATTR-ACT study (NCT01994889), and 44 (9.7%) were without an EFS event . The median EFS was 8.8 months (95% CI: 6.2\u201314.2) in patients with 50% relative increased NT-proBNP levels (nk test) A. The sen = 48) and 12.6 months (95% CI: 8.3\u201316.6) in those with non-increased levels (n = 260), HR: 0.86 (95% CI: 0.60\u20131.25); p = 0.43 (log-rank test) (p = 0.40) and 0.91 , respectively (figure not shown).We also analysed median EFS in the 308 patients with at least two cTnT-HS assessments, according to whether patients had a 50% relative increased cTnT-HS levels. The median EFS was 12.2 months (95% CI: 5.1\u201315.4) in patients with 50% relative increased levels (nk test) B. Simila2) to be significantly associated with an extended EFS at a 5% level. Of these, the multivariate analysis, identified non-50% relative increased NT-proBNP levels , as well as baseline levels of NT-proBNP \u2264 3000 ng/L and cTnT-HS \u2264 50 ng/L to be significantly and independently associated with extended EFS , cTnT-HS (\u226450 ng/L), and eGFR (>45 mL/min/mnded EFS .n = 248) was 17.5 months (IQR: 10.9\u201318.2).Among the 248 patients assessed in Cohort B, 63 (25%) had ATTRv and 185 (75%) had ATTRwt . The popn = 248), 194 patients (78.2%) were still being treated with tafamidis and 54 (21.8%) had discontinued tafamidis. In patients who discontinued tafamidis, 27 patients had died (including 25 before 18-month timepoint), 15 had aggravated disease, 8 had been enrolled in the ATTR-ACT study (NCT01994889), 1 had been enrolled in the APOLLO study (NCT01960348), 1 did not support the treatment, 1 refused to continue treatment, and 1 was non-compliant. In Cohort B (n = 248) the following proportions of patients had baseline levels of NT-proBNP maintained \u22643000 ng/L or decreased from >3000 ng/L to \u22643000 ng/L: 139/223 (62.3%) after 6 months, 89/162 (54.9%) after 12 months, and 80/145 (55.2%) after 18 months. At 18 months, 17% (25/145) of the patients were dead (In Cohort B (ere dead C.n = 248) the following proportions of patients had baseline levels of cTnT-HS maintained \u226450 ng/L or decreased from >50 ng/L to \u226450 ng/L: 90/204 (44.1%) after 6 months, 48/147 (32.7%) after 12 months, and 57/134 (42.5%) after 18 months. 19% (25/134) of the patients had died . We also generated the scatterplots for the subpopulations of patients that initiated tafamidis with NT-proBNP levels >3000 ng/L (p = 0.03). Similarly, for patients with NT-proBNP levels \u22643000 ng/L, the slope was 0.0007 (95% CI: \u22120.0070 to \u22120.0084) before tafamidis and 0.0080 (95% CI: 0.0025\u20130.0135) after tafamidis (p = 0.15). However, despite a stable positive slope of NT-proBNP levels after tafamidis, in patients with initial NT-proBNP levels \u22643000 ng/L, most of the individual NT-proBNP levels remain within a narrow logarithmic range (8 logs corresponding to 3000 ng/L).The scatterplots for log (NT-proBNP) before and after tafamidis treatment, are shown in 000 ng/L B and tho000 ng/L C. For pap < 0.01. In addition, we plotted the log (cTnT-HS) levels of patients with cTnT-HS levels >50 ng/L (p < 0.001). Similarly, for patients with cTnT-HS levels \u226450 ng/L, the slope was 0.0071 (95% CI: 0.0025\u20130.0118) before tafamidis and 0.0025 (95% CI: \u22120.0045\u20130.0094) after tafamidis (p = 0.35).The scatterplots for log (cTnT-HS) levels before and after tafamidis treatment, are shown in >50 ng/L E and tho>50 ng/L F. For paOur study is the first to describe the natural history of cardiac biomarkers in ATTR and their evolution after tafamidis treatment in a real-life cohort. Our results show that NT-proBNP and cTnT-HS levels increase in ATTR-CA natural history and that 50% relative increases in NT-proBNP levels significantly increases the risk of death, heart transplant, or acute heart failure in ATTR-CA patients. In contrast, increases in cTnT-HS levels were not associated with an increased risk of these events. Baseline levels of NT-proBNP \u2264 3000 ng/L and cTnT-HS \u2264 50 ng/L were found to be independently associated with extended EFS. Furthermore, our results show that tafamidis slows the increasing of NT-proBNP levels associated with amyloidosis evolution in ATTR-CA patients with NT-proBNP levels > 3000 ng/L when tafamidis is initiated. Similarly, tafamidis also slows the increasing of cTnT-HS levels in patients with cTnT-HS levels > 50 ng/L. The proportion of patients who maintained their baseline levels of NT-proBNP \u2264 3000 ng/L or decreased their NT-proBNP levels from >3000 ng/L to \u22643000 ng/L at 12 months was 31% for untreated patients and 55% for those treated with tafamidis.Amyloid formation is a progressive and dynamic process with cardiac biomarkers reflecting the pathophysiology of the disease. Our results show that cTnT-HS and NT-proBNP levels increase during the natural evolution of ATTR-CA.NT-proBNP is a cardiac biomarker indicative of cardiac wall stress and levels increase when heart ventricles become overloaded. NT-proBNP levels are used to diagnose congestive heart failure ,12,13,14In cardiac amyloidosis, fibrillogenesis is a dynamic process. An increase in biomarker levels may be linked to cardiomyopathy evolution, emphasizing the need to slow down their progression. Thus, increasing NT-proBNP levels may be linked with cardiac wall stress associated with amyloid infiltration. While, increasing troponin levels may be linked with the toxicity of amyloid fibrils .The prognostic value of baseline NT-proBNP and cardiac troponin levels in cardiac amyloidosis patients are well established ,21,22,23In our analysis of the cohort of patients treated with tafamidis we found that NT-proBNP levels continued to increase after initiating tafamidis but at a constant or reduced rate. We also found that tafamidis had a similar effect on cTnT-HS levels. The activity of tafamidis is reportedly due to its capacity to stabilise the TTR tetramer, reducing the build-up of amyloid fibrils in the heart . This meInteresting, our results correspond with an exploratory endpoint of the ATTR-ACT study showing a slightly higher NT-proBNP levels in patients, at 12 and 30 months  and one assessment before and another after tafamidis (Cohort B). However, a comparison of the patients included and those not included in the cohorts did not show any major imbalance. Secondly, the time intervals between NT-proBNP assessments were not control and varied. Our data were analysed using piecewise regression random model that generates slope models before and after treatment despite this variability in the time intervals. Thirdly, the cTnT-HS levels used for our analyses are different from the troponin assay usually used in cardiac amyloidosis literature. Recently, cTnT-HS assays have replaced the more traditional assays. These new assays detect the same proteins but with a much lower detection limit . It has Finally, this study was not designed to compare patients treated with tafamidis with those not treated. The patients in the two cohorts were not included during the same time period. Thus, untreated patients from Cohort A that were included in the Cohort B (tafamidis treated patients) may have less severe ATTR-CA. Indeed, a patient included in Cohort A with an EFS event would not have been included in Cohort B. Comparisons of the baseline characteristics of the cohorts support this hypothesis with lower level of NT-proBNP, cTnT-HS, Gillmore stage, and Grogan stage in treated patients. However, the sensitivity analyses that we performed gave results consistent with the results and conclusions herein reported. Nevertheless, this indirect comparison was not an objective of this study. We are currently indirectly comparing these cohorts using an adapted methodology, including the use of propensity scores and matching analysis.NT-proBNP and cTnT-HS are biomarkers for cardiac dysfunction that increase as ATTR-CA evolves. Increasing NT-proBNP levels, but not cTnT-HS levels, signal increasing cardiac dysfunction with an increased risk of death, heart transplant, or acute heart failure, in ATTR-CA patients. Our results seem to indicate that tafamidis slows disease evolution in ATTR-CA patients by stabilizing NT-proBNP and cTnT-HS levels."}
+{"text": "Achillea santolinoides and Achillea aleppica aeral parts and root were extracted with ethyl acetate, methanol, and water. Detailed phytochemical profiles were obtained using UHPLC-MS, yielding the identification of hydroxybenzoic and hydroxycinnamic acids, phenolic acid glycosides and sugar esters, acylquinic acids, O-glycosyl flavones and flavonols, and flavonoid aglycons, among others. The antioxidant properties and enzyme inhibitory activities of the extracts were assayed with in vitro tests. The phenolic content of the water extracts was significantly higher as compared to the ethyl acetate and methanol ones. A. aleppica aerial parts methanol extract possessed highest flavonoid content . Antioxidant properties assessment revealed that the methanol extract of A. santolinoides roots actively scavenged DPPH (54.11 mg TE/g) and ABTS radicals (112.53 mg TE/g) and possessed highest reducing potential . The ethyl acetate extracts of aerial parts and roots of both species showed highest inhibition against BuCHE (6.07\u20136.76 mg GALAE/g). The ethyl acetate extract of A.santolinoides aerial part showed highest inhibition against tyrosinase (73.00 mg KAE/g). These results showed that the tested Achillea species might represent novel phytotherapeutic avenues for the management of Alzheimer\u2019s disease and epidermal hyperpigmentation conditions, which are both associated with oxidative stress. This paper could shed light into future potential industrial applications using the tested Achillea species.In the current study,  Achillea genus, one of the most important genera of the Asteraceae family with ethnopharmacological significance, consists of approximately 85 species mainly distributed in Middle East regions, such as Iran, Turkey, and Serbia and Eastern regions of Europe \u2212 at m/z 357.084) were obtained (m/z 195.050 (C6H11O7\u2212) corresponding to the [gluconic acid-H]\u2212 supported by the fragment ions at m/z 177.040 [GA-H-H2O]\u2212, 87.007 [GA-H-C3H8O4]\u2212 and 59.012 [GA-H-C3H8O4-CO]\u2212  and four hydroxycinnamic acids  together with extracts . In addiobtained . They yi8O4-CO]\u2212 .O-(6-caffeoyl)-hexoside (32) was deduced from the loss of vanillic acid (168 Da) at m/z 323.077 and a subsequent transition 323.077\u2192221.046 [M-H-102]\u2212 arising from the hexose cross ring cleavage . The latter ion points out to the caffeoyl moiety at Hex C-6. Regarding 42, the prominent ion at m/z 323.077 [M-H-C7H6O3]\u2212 and a base peak at m/z 137.023 [salicylic acid-H]\u2212 together with m/z 93.033 [salicylic acid-H-CO2]\u2212 were in accordance with caffeic acid-O--hexoside (Tanacetum vulgare [Vanillic acid-4-hexoside . Both 32 vulgare .A. santolinoides. Syringic acid-hexoside (6) was presented mainly in A. allepica roots  afforded prominent ions at m/z 353.088 and 191.055 indicating the subsequent losses of a caffeoyl moiety (diCQA 33 and 37 were witnessed by the \u201cdehydrated\u201d ion of quinic acid at m/z 173.044 (100%) supported by the diagnostic ions at m/z 335.0771 [CQA-H-H2O]\u2212 and 135.044 [caffeic acid-H-CO2]\u2212 in 3,4-diCQA (33) (diCQA as suggested by the lack of ion at m/z 335 and the chromatographic behavior on the reverse phase (the most lipophilic diCQA isomer). The base peak at m/z 191.055 evidenced 1,3-diCQA (24a), 1,5-diCQA (34) and 3,5-diCQA (35) supported by the relative abundance of the ions at m/z 179.034 and m/z 135.044: 73.2% and 58.7% (24a), 6.2% and 6.6% (34), and 53.1% and 52.7% (35), respectively.Five peaks 24, 33\u201335, and 37 ([M-H]l moiety . The vicCQA (33) . The secp-coumaroyl-caffeoylquinic acids (p-CoCQA) isomers 41 and 44 at m/z 499.122 (C25H23O11) were deduced from the distinctive fragments at m/z 337.093 [M-H-caffeoyl]\u2212, m/z 163.039 [p-CoA-H]\u2212 and m/z 119.049 [p-CoA-H-CO2]\u2212 for p-coumaric acid (m/z 337.093 (83.6%) indicating a loss of caffeoyl residue before the p-coumaroyl one. This assignment was also supported by the base peak at m/z 163.039 as was registered in 3-p-CoQA \u2212 and m/z 353.270 [M-H-feruloyl]\u2212 for feruloyl-caffeolylquinic acids (FCQA). The fragment ion at m/z 335.0754 [M-H-FA]\u2212 accompanied by the \u201cdehydrated\u201d form of quinic acid suggested 3F-4CQA (40) [m/z 193.050 together with the abundant ion at m/z 134.036 (74.4%) as was registered in 3-FQA (m/z 161.023 [CA-H-H2O]\u2212 accompanied by the abundant ions at m/z 179.034 [CA-H]\u2212 (42.3%) and 367.104 (34.1%) \u2212 at m/z 327.051 (54) and 311.056 (57) supported by the relevant ions at m/z 299.056 0,2X/CO\u2212 [(M-H)-120\u201328]\u2212 and m/z 283.061, respectively. This fragmentation pathway was consistent with C-8 hexosyl luteolin/apigenin [C-6 hexosyl isomers 52 and 58 was shown by the ions at m/z 447.094 [M-H]\u2212 (100%) and 431.099, as well as 0,3X\u2212 at m/z 357.062 and 341.067, and 0,2X\u2212 at m/z 327.051 and 311.056. The aglycones luteolin  and apigenin  were discernable by the RDA ions 1,3A\u2212 (m/z 151.022), 0,4A\u2212 (m/z 107.012), 1,3B\u2212 at m/z 133.028  and 117.033 . Based on the comparison with reference standards, compounds 52, 54, 57, and 58 were identified as homoorientin, orientin, vitexin, and isovitexin, respectively.MS/MS spectra of the acquired . In the apigenin . In cont\u2212 at m/z 593.152 (C-glycosyl flavon pathway were produced at m/z 473.109 [(M-H)-120]\u2212, 383.077 [(M-H)-90\u2013120]\u2212 and 353.067 [(M-H)-2 \u00d7 120]\u2212 suggesting the presence of two C-hexosyl moieties on the flavonoid skeleton \u2212 at m/z 413.088 [(M-H)-60\u2013120]\u2212 and \u2212 at 323.057 [(M-H)-120-150]\u2212 suggesting the presence of both C-pentosyl (X0) and C-hexosyl (X1) moieties. Additionally, methylluteolin was assigned on the basis of specie at m/z 299.560 [MeLu-H]\u2212 and 298.048 Y0/0,2X1/\u2022CH3/CO [m/z 323.057 ([(M-H)-(132 + H2O)-120]\u2212 and 443.097 ([(M-H)-(132 + H2O)]\u2212 suggesting O-pentosyl unit at 2\u2033 of the primary hexose  593.152 . Concerny hexose . Diagnos\u2212 at 609.147, 47 was annotated as 6, 8-diC-hexosyl-luteolin, while 48 was assigned to O, C-dihexosyl-luteolin. The latter structure was shown by a series of diagnostic ions at m/z 447.093 [M-H-Hex]\u2212, 357.062 [M-H-Hex-90]\u2212 and 327.051 [M-H-Hex-120]\u2212. Additionally, ions at m/z 298.048 , 175.039  and 133.028 1,3A\u2212 indicated luteolin. The sugar chain of 56 was consistent with rutinose (308 Da); aglycone quercetin was witnessed by a series of fragments including RDA ions at m/z 178.998 \u2212, 163.003 \u2212, 151.002 \u2212, 121.028 \u2212, 107.012 \u2212. Based on comparison with reference standard, 56 (rutin), 60 (luteolin-7-glucoside), 65 (kaempferol-3-glucoside), 66 (isorhamnetin-3-glucoside), 67 (apigenin-7-glucoside), luteolin (50), quercetin (72), apigenin (75), kaempferol (77) and chrysoeriol (78) were unambiguously identified , 299.056  and 329.067 (71) [(M-H)-HexA]\u2212, respectively, indicating flavonoid hexuronides  was deduced from the fragment ions at m/z 243.030 [(M-H)-HexA-CH3-HCO\u2022-CO]\u2212, 227.035 [(M-H)-HexA-CH3-HCO\u2022-CO2]\u2212 as well as RDA ions at m/z 133.028 . Compound 69 was ascribed to chrysoeriol-O-hexuronide , while 70 was consistent with jaceosidin-O-hexuronide flavones . In (\u2212) llea sp. .1,3A\u2212) at m/z 165.990 , 139.039 , 136.986  and 1,2 B\u2212 at m/z 121.028. Within this group, compounds 73 (patuletin), 74 (axillarin), and quercetagetin-3,6,3\u2032(4\u2032)-trimethyl ether (80) were quercetagetin derivatives, while compounds 77 (hispidulin) and 81 (cirsimaritin) were scutellarein derivatives , 164.981 , 163.002 , 136.987 . Moreover,  at m/z 132.020 indicated 2 methoxy groups either in C-3, C-4\u2032 or C-3\u2032, C-4\u2032, as was observed in santin and eupatilin, respectively + and 243.101 [M + H-2H2O]+ and a base peak at m/z 237.111 [M + H-H2O-CH2]+. This fragmentation pathway could be associated with the presence of peroxide group and 84 was tentatively ascribed to tanaparthin-peroxide, previously isolated from Achillea nobilis (+ at m/z 307.153 gave a base peak at m/z 247.132 [M + H-CH3COOH]+ which is in accordance with the structure of achillicin/matricin. Compound 87 differs from 85 for 60 Da (CH3COOH) and revealed the same fragmentation patterns as 85. Thus, compound 87 was tentatively annotated as achillin/leucodin , 2xH2O (\u221236 Da), CO (\u221228 Da), as well as concomitant loss of H2O + CO (\u221246 Da), 2H2O + CO (\u221264 Da), 88 and 89 were ascribed to artabsin and dihydrosantamarin, respectively, and were previously isolated from Achillea collina [MS/MS spectrum of 84 [M + H] nobilis  44]. Co. Co+ at leucodin . Similarm/z 280.263 (C18H33NO) together with distinctive fragments at m/z 263.236 [M + H-NH3]+ and m/z 245.225 [M + H-NH3-H2O]+, suggesting amide of octadecadienoic acid. Additionally, the suggested structure was supported by the fragments at m/z 81.070 (C6H9), 69.070 (C5H9), 57.070 (C6H9)  . Thus, 9ectively  46]..m/z 280.A. aleppica aerial parts (55.15 mg TE/g) and A. santolinoides roots (54.11 mg TE/g) showed highest scavenging activity against DPPH. In contrast A. aleppica roots water extract (101.88 mg TE/g) and A. santolinoides roots methanol extract (112.53 mg TE/g) were most potent in scavenging ABTS. Protocatechuic acid and its derivatives identified in the A. aleppica roots water extract, A. aleppica aerial parts methanol extract, and A. santolinoides roots methanol extract, has been reported to exhibit radical scavenging activity [A. aleppica aerial parts and A. santolinoides roots showed highest reducing capabilities  and ethyl acetate and water extract of the aerial parts of A. santolinoides (27.37 and 26.06 mg EDTAE/g), respectively possessed strong chelating ability. Caffeic acid, chlorogenic acid, and protocatechuic acid were identified in aerial parts of A. aleppica water and A. santolinoides ethyl acetate extracts. Interestingly, a study conducted by Andjelkovi\u0107, et al. [A.santolinoides was previously reported to possess antioxidant effect on brain tissues in pentylenetetrazole-induced seizures Wistar rat models [A.santolinoides was also found to exhibit antioxidant potential against DPPH radical (IC50 = 129\u2013372 mg/mL) [The total antioxidant capacity of the extracts was determined using the phosphomolybdenum assay. As shown in activity ,48. Neocactivity . The redbilities . The che, et al.  has asset models . The ess2 mg/mL) .Achillea species against enzymes targeted in the management of diabetes mellitus type II, Alzheimer\u2019s disease, and skin hyperpigmentation problems was investigated. Alzheimer\u2019s disease has escalated to epidemic proportions and the need for complementary therapeutic agents to effectively manage this debilitating condition is of paramount importance. From A. aleppica aerial parts ethyl acetate extract and A. santolinoides roots methanol exhibited highest inhibition against AChE. A previous molecular docking study confirmed the interaction of orientin with AChE which showed least binding energy and highest binding affinity [A. aleppica roots and A. santolinoides aerial parts was previously reported to inhibit BuChE in an in silico study. On the other hand, the ethyl acetate extracts of A. aleppica aerial parts and roots (6.07 and 6.73 mg GALAE/g) and as well as that of A. santolinoides aerial parts (6.76 and 6.70 mg GALAE/g) were most active against BuChE. The inhibition of BuChE has been advocated in the later stage of Alzheimer\u2019s disease. During the progression of the disease, BuChE level increases, exacerbating the conditions of the patient [A. aleppica and A. santolinoides aerial parts and roots possessed weak anti-diabetic properties. Tyrosinase, a rate limiting enzyme responsible for the biosynthesis of melanin, is considered to be a key therapeutic strategy for the management of skin hyperpigmentation conditions. In the present study, methanol extracts of A. aleppica aerial parts and roots showed the highest inhibitory activity against tyrosinase. In other side, ethyl acetate and methanol extracts of both studied parts of A. santolinoides displayed strongest anti-tyrosinase activity. Hispidulin, isolated from Phyla nodiflora and identified in extracts which actively inhibited tyrosinase was previously reported to exhibit inhibitory action against tyrosinase with an IC50 value of 146 \u00b5M [The inhibitory ability of extracts prepared from the aerial parts and roots of the selected affinity . Vitexinaffinity . Acacetiaffinity . However patient . The abif 146 \u00b5M .A. aleppica roots EA and MeOH and A. santolinoides roots EA were grouped together. Similarly, in PC1 vs PC2 and PC1 vs. PC3, A. A. santolinoides roots MeOH and A. aleppica aerial parts MeOH were close together. Following PCA, a hierarchical classification was done to obtain a clearer picture of the different group. Based on the scores of samples on the three PCs, the hierarchical analysis revealed two principal clusters, each of which was divided into two sub-clusters  were characterized by higher antioxidant activity while samples of the second cluster  were marked by stronger enzyme inhibitory activity.Subsequent to comparison of the bioactivities of the samples of each species, principal component analysis (PCA) was used in order to uncover the similarities/differences among the extracts of both species, in light of assessed antioxidant and enzyme inhibitory activities. The results of PCA were displayed in clusters B. The sa2X) and biological activities (R2Y) respectively, indicating the good performance of the model.The relationship between the metabolites and biological activities Partial was assessed and result was reported in Achillea species. In respect of the genes modulation, 73, 122, 280, 122, 113, 122, 57, 57, 254, 287 and 272 mRNA were found to be up-regulated and down-regulated by artabsin, dehydroleucodin, dihydrosantamarin, leucodin, matricin, tanaparthin peroxide, neochlorogenic acid, chlorogenic acid, homoorientin, vitexin and isovitexin respectively (Artemisia capillaries, it has been demonstrated that leucodin dose dependently enhances phosphorylation of AMPK in alcohol-exposed HepG2 cells [After the phytochemical screening and in vitro evaluation of biological properties of the samples, we have been engaged in the investigation of KEGG pathway enrichment analysis of identified sesquiterpene lactones and derivatives and five of the main phenolics of ectively . As regaectively . StructuG2 cells . FurtherG2 cells . While tG2 cells . This fiA.aleppica and A. santolinoides roots and aerial part. Chlorogenic acid was the main derivative in aerial parts of both the species. 3,5-diCQA was the most important diCQA derivative in A. aleppica while 1,3diCQA was the most significant in A santolinoides. Sesquiterpene lactone and fatty acid amides have been also detected showing large chemical diversity in the constituents of the plant. The extraction with ethyl acetate, methanol and water allowed to prepare samples with different composition that were used to assess their in vitro bioactivity on several antioxidant and enzyme inhibition assays. The methanol extract of A. santolinoides roots possessed significant antioxidant activities. The ethyl acetate extracts of the aerial parts and roots of both Achillea species showed significant inhibition against butyrylcholinesterase while the ethyl acetate extract of A. santolinoides aerial part actively inhibited tyrosinase. The detailed phytochemical investigation, the evaluation of in vitro bioactivity, of the two Achillea species indicate these plants as valuable starting point for potential future studies and possible applications extracts in cosmetic, pharmaceuticals and nutraceuticals products. KEGG mapping using some of the phenolics and sesquiterpenes of the plants allowed to predict some of the possible molecular targets for significant bioactivities. This information opens new opportunities of research and application for A. aleppica and A. santolinoides extracts and isolated compounds.This study allowed obtaining a detailed phytochemical fingerprint of"}
+{"text": "Staphylococcus aureus (MRSA) SCCmec IV[2B] has become one of the most common community-associated MRSA clones in Australia. We report the complete genome sequence of one of the earliest isolated Australian S. aureus ST1-MRSA-IV strains, WBG8287, isolated from an Indigenous Australian patient living in the remote Kimberley region of Western Australia.Sequence type 1 (ST1) methicillin-resistant  Staphylococcus aureus (CA-MRSA) first emerged in Australia in the 1980s . WBG8287 was additionally sequenced using the Illumina NextSeq 500 platform, the Nextera XT DNA library preparation kit (paired-end format) , and the NextSeq 500/550 kit v2.5 (300-cycle format) (Illumina). The Illumina sequencing produced 2,083,742 reads, with a maximum read length of 151\u2009bp and a mean read length of 147\u2009bp. The Nesoni clip tool (github.com/Victorian-Bioinformatics-Consortium/nesoni) was used to remove adaptor sequences and quality filter reads. The assembly was polished with the Illumina reads 5\u00d7 using Minimap2 v2.17-r941 , using MinKNOW v20.10.3 and MinKNOW Core v4.1.2 software. Base calling was performed using Guppy v4.4.1\u2009+\u20091c81d62 (ONT) with the dna_r9.4.1_450bps_hac.cfg model. Sequencing produced 470,277 reads and a read-length CP070986.1 (chromosome), CP070987.1 (pWBG750), and CP070988.1 (pWBG751). The sequencing reads have been deposited in the Sequence Read Archive under accession numbers SRX11246054 and SRX11246053.The WBG8287 sequences are deposited in NCBI GenBank under accession numbers"}
+{"text": "Macrolides are a significant family of natural products with diverse structures and bioactivities. Considerable effort has been made in recent decades to isolate additional macrolides and characterize their chemical and bioactive properties. The majority of macrolides are obtained from marine organisms, including sponges, marine microorganisms and zooplankton, cnidarians, mollusks, red algae, bryozoans, and tunicates. Sponges, fungi and dinoflagellates are the main producers of macrolides. Marine macrolides possess a wide range of bioactive properties including cytotoxic, antibacterial, antifungal, antimitotic, antiviral, and other activities. Cytotoxicity is their most significant property, highlighting that marine macrolides still encompass many potential antitumor drug leads. This extensive review details the chemical and biological diversity of 505 macrolides derived from marine organisms which have been reported from 1990 to 2020. Lamellomorpha strongylata (La. strongylata) [Symbiodinium sp. [The term \u201cmacrolide\u201d was coined by Woodward in 1957 ,3,4. Thengylata) , and thenium sp. . MacroliThis literature review from 1990 to 2020 highlights 505 new macrolides derived from marine organisms  . CompareTheonella sp. (T. sp.) sponges produced a series of dimeric macrolides called swinholides A\u2013G (1\u20137) and isoswinholide A (8) [9), isobistheonellide A (10), and bistheonellic acids A (11) and B (12)\u2014are also produced by Okinawan T. sp. sponges [13), which is weakly cytotoxic and obtained from Japanese sponge Polyfibrospongia sp., was elucidated by X-ray single crystal diffraction [14) was isolated from Mycale adhaerens (M. adhaerens) and identified by spectroscopic analysis [The Okinawan de A (8) ,17,18,19 sponges . The strfraction . 13-Deoxanalysis . 15) and halistatin 2 (16) were isolated from Phakellia carteri from the Comoros Islands and Axinella cf. carteri (Dendy) from the Western Indian Ocean [17) was produced in extremely small quantities by Phakellia sponges collected at Chuuk [The antimitotic macrolides halistatin 1 , 2 (19), and 3 (20), which were isolated from Spongia sp. in the Republic of Maldives and identified via spectral data without stereochemistry [21), 5 (22), 6 (23), 7 (24), 8 (25), and 9 (26) from Spirastrella spinispirulfera (S. spinispirulfera) on the southeast coast of Africa [Independent groups have reported the potent antitumor macrolides spongiastatins 1 , C (28), and D (29) have been isolated from the Caledonian sponge Neosiphoniu superstes [Three macrolides sphinxolides B (uperstes . 30), B (31), and C (32), were reported without stereochemical data in an Okinawan Juspis sponge [Three new trisoxazole macrolides, jaspisamides A (30), B , and C (33) was isolated from a Republic of Maldives Spongia sponge and exhibited significant cytotoxicity towards murine P388 lymphocytic leukemia [34) and B (35), have been produced by the marine sponge Reidispongia coerulea (R. coerulea) [A new 22-membered macrocyclic lactone named dictyostatin 1 (leukemia . The rel4) and B , have beoerulea) . The reloerulea) . 36) and superstolide B (37) have been isolated from the deep-water marine sponge Neosiphonia superstes (N. superstes) [38), was produced by the shallow-water Caribbean sponge Forcepia sp. [39), belonging to the halichondrin family, was isolated from the New Zealand deep-water sponge Lissodendoryx sp. (Li. sp.) [40) and B (41) have an unprecedented scaffold and were isolated from the Indian Ocean sponge Phorbas sp. (P. sp.), with complete stereochemistry and absolute configuration determined by spectroscopy and partial synthesis [42) and laulimalide B (43) isolated from Okinawan sponge Fasciospongia rimosa were determined by X-ray analysis [44), neolaulimalide (45) and zampanolide (46), have been produced by the F. rimosa genus [47), isolated from the marine sponge Halichodria okadai, exhibited significant inhibition of vascular cell adhesion molecule 1 (VCAM-1) [Cytotoxic superstolide A (perstes) ,39. Anotepia sp. . IsohomoLi. sp.) . Phorboxynthesis ,43. The analysis . Other climalide  and zamppanolide , have be48), exhibiting antifungal and cytotoxic activities, was obtained from the sponge Leucascandra caveolata (Le. caveolata) [Callipelta sp.  contains the first member of a new class of marine-derived macrolides, callipeltoside A (49), which incorporates an unusual chlorocyclopropyl group and an amino sugar [50\u201352) and 5-desacetylaltohyrtin A (53) were isolated from the sponge Hyrtios altum and their absolute stereochemistries were determined by spectroscopy [La. strongylata for cytotoxicity towards the P388 cell line yielded swinholide H (54) [Leucascandrolide A , exhibittoside A , which ino sugar ,51,52. Chyrtin A  were isotroscopy . Screenie H (54) .Li. produced the antitumor macrolides neonorhalichondrin B (55), neohomohalichondrin B (56), 55-methoxyisohomohalichondrin (57), 53-methoxyneoisohomohalichondrin B (58a) and 53-epi-53-methoxyneoisohomohalichondrin B (58b) [Another deep-water (> 100 m) sponge of the genus ondrin B , neohomo59) and B (60) were isolated from the Haliclona sponge, representing a potentially important new class of antitumor leads [61) and C (62), two members of a novel class of marine glycoside macrolides, were isolated from the sponge Cal. sp. [Macrolide salicylihalamides A  and B  were iso63\u201366), were isolated from an Okinawan marine sponge, Halichondria sp. [Four new oxazole-containing compounds, halishigamides A\u2013D , showing modest cytotoxicity [68), 32-hydroxymycalolide A (69), and 38-hydroxymycalolide B (70), were isolated from the marine sponge M. magellanica and showed cytotoxicity towards L1210 cells [A Palau toxicity . Three m10 cells . 71), a thiazole-containing macrolide with an unique dilactone functionality, was isolated from M. sp. sponge [72\u201375), were produced by the marine sponge Amphimedon spp. collected in the Great Australian Bight [Pateamine 1 (. sponge . Four nean Bight . 76), NB (77), NC (78), ND (79) and NE (80) were isolated from the New Caledonian Litbistida sponges N. superstes and R. Coerulea [81\u201383) and reidispongiolide C (84) are new cytotoxic macrolides from Okinawan species of Ircinia [85) is a 16-membered macrolide from the calcareous sponge Le. caveolata from the northeastern waters of New Caledonia [M. sp. contained the polyoxygenated, pyranose ring-containing, 16-membered macrolide peloruside A (86) [Cytotoxic macrolides haterumalides NA  , which we A (86) . 87) has been isolated from the Vanuatu marine sponge Spongia sp. [88), was isolated from a marine sponge of the genus Dactylospongia. This has been synthesized and the relative stereochemistry of the acyloxymethine and the absolute configuration of the whole molecule have been determined [Ha. sp. was found to contain the cyclic metabolite haliclamide (89) [Cytotoxic spongidepsin (ngia sp. . A new ctermined . The Vanide (89) .90\u201393) have been found in sponge Myriastra clavosa [94\u2013100) are antimitotic macrolides isolated from the Caribbean marine sponge S. coccinea [Clavosolides A\u2013D  and (19Z)-halichondramide (102), and the open ringed secohalichondramide. Neohalichondramide and (19Z)-halichondramide exhibited significant cytotoxicity and antifungal activity toward the human leukemia cell-line K562 and Candida albicans  [The sponge lbicans) .103), 32-hydroxymycalolide A (104) and 38-hydroxymycalolide B (105), have been isolated from a Japanese M. magellanica [106\u2013110) were isolated from Forcepia sponge collected in the U.S. Gulf of Mexico [111), isolated from the marine sponge Geodia exigua, was reported to inhibit fertilization of sea urchin (Hemicentrotus pulcherrimus) gametes but not embryogenesis [Three cytotoxic mycalolides, 30-hydroxymycalolide A (ellanica . The fivf Mexico . Exiguologenesis . The absogenesis .112) and B (113) were obtained from a deep-water (>200 m) Leiodermatium sponge [114), isolated from Ircinia sp. (Papua New Guinea), was found to be potently cytotoxic, causing S-phase arrest, suggestive of protein synthesis inhibition [115\u2013118) were produced by Pachastrissa nux (P. nux) (Gulf of Thailand) [Cytotoxic macrolides leiodolides A  . 119), was isolated from P. nux sponge [120) and the related hurghadolide A (121), with cytotoxicity towards human colon cancer cells, were produced by T. swinhoei  [An antiplasmodial macrolide, kabiramide L  .122) was isolated from Red Sea sponge Negombata corticata and showed significant antifungal and anticancer activities, suggesting it as a potential member of the bioactive latrunculin family [123) with potential cytotoxic and antifungal activities. This compound was synthesized to determine its absolute configuration and the relative stereochemistry of C-13 [124), a complex mixture of acyl esters of a macrolide related to tedanolide, was isolated from Candidaspongia sp. (Can. sp.) (Papua New Guinea) and Can. flabellata  [125\u2013130) were produced by sponge Cacospongia mycofijiensis (Cac. mycofijiensis)  [131\u2013135) are chlorocyclopropane macrolides isolated from marine sponge P. sp. [Oxalatrunculin B  . FijianoVanuatu) . Phorbase P. sp. ,97. 136\u2013138), 18-epi-latrunculol (139) and latrunculones A (140) and B (141), were obtained from Cac. mycofijiensis [142), salarin B (143) and tulearin A (144) were obtained from repeated collections of the Madagascan sponge Fascaplysinopsis sp. (F. sp.) [Latrunculin analogs, latrunculol A\u2013C  . 145), which was considered to be the precursor of salarins A and B [Siliquariaspongia mirabilis contained an antitumor macrolide lactam named mirabilin (146) [147) was isolated from the Madagascar sponge F. sp. and was found to inhibit proliferation of K562 leukemia cells [148), containing a rare hexahydro-1H-isoindolone and trichlorocarbinol ester, was isolated from marine sponge of the genus Phorbas [A further collection led to the isolation of salarin C ( A and B . Marine in (146) . The nitia cells . Muirono Phorbas . 149), B-1092 (150), B-1020 (151) and B-1076 (152), were extracted from the Poecilosclerid sponge Li. sp. in microgram quantities and their structures were elucidated by capillary NMR spectroscopy [Four variants of halichondrin B, B-1140 (troscopy . 153) and two analogs, 15-O-methylenigmazole A (154) and 13-hydroxy-15-O-methylenigmazole A (155), were extracted from the marine sponge Cinachyrella enigmatica collected in Papua New Guinea [Cytotoxic phosphate-containing macrolide enigmazole A (w Guinea  and theiw Guinea .156\u2013162) were obtained from the Madagascan F. sp. sponge. Scalarins D, E, H, and J inhibited cell proliferation in a dose- and time-dependent manner [163\u2013165) were obtained from the Okinawan marine sponge T. sp. and absolute configurations were determined by combining a JBCA method, a universal NMR database, and a 13C-acetonide method [Seven scalarin analogs D\u2013J  and swinholide K (167) [168), with strong cytotoxicity towards human Jurkat J16 T and Ramos B lymphocytes, was isolated from marine sponge Cal. sp. [169) and C (170) were isolated from Petrosiidae sponge with stereochemical assignment via enantioselective synthesis of the macrocyclic core [171), F (172), and G (173) were isolated from the marine sponge Poecillastra sp. [The Indonesian sponge  K (167) . An unusCal. sp. . Cytotoxlic core . Cytotoxstra sp. .Periconia byssoides (Per. byssoides), obtained from the sea hare Aplysia kurodai (Ap. sp.), was reported to produce the cytotoxic triols pericosides A and B, and four new macrolides, macrosphelides E\u2013H (174\u2013177) [178) and macrosphelides E\u2013H from Per. byssoides isolated from Ap. kurodai were also reported elsewhere [179) and H produced by Per. byssoides from Ap. kurodai, and the cytotoxic macrosphelide M (180) [Penicillum verruculosum (IMI352119) was reported to produce three macrolides with antifungal activity: BK223-A (181), BK223-B (182) and BK223-C (183) [Varicosporina ramulosa has been reported to produce -colletodiol (184), -colletodiol (185) and colletoketol (186) [187) and pandangolide 2 (188) were extracted from an unidentified fungus isolated from marine sponge collected in Indonesia [The fungus 174\u2013177) . Macrosplsewhere . Macrosplsewhere . The synlsewhere ,118. Abs M (180) ,119,120.-C (183) . The mitol (186) ,123. Thendonesia . 189), macrolide dimer pandangolide 4 (190), and a new acetyl derivative of 5-hydroxymethylfuran-2-carboxylic acid were produced by the fungus Cladosporium herbarum (Cla. herbarum), associated with the sponge Callyspongia aerizusa  and collected in Bali [191) was obtained from an extract of the marine fungus Myrothecium roridum (M. roridum) [192\u2013196) were isolated from the mangrove fungus Aigialus parvus BCC 5311 [197) and 15G256\u0461 (198) were obtained from the marine fungus Hypoxylon oceanicum LL-15G256 [199) and B (200), were produced by the fungus Cladosporium isolated from the brown alga Actinotrichia fragilis  [Pandangolide 3  . The 14-BCC 5311 . PotentiL-15G256 . Two cyt, Japan) . Sargassum sp.  was the source of two 12-membered ring lactones (201\u2013202) [203), roridin Q (204) and 2,3-deoxyroritoxin D (205) were obtained from M. roridum on submerged wood in Palau [Gliocladium sp. isolated from the alga Durvillaea antarctica  yielded 4-ketoclonostachydiol (206) [An unidentified endophytic fungus from the brown alga 201\u2013202) . 12-Hydrin Palau . Glioclaol (206) . 207) and 2\u2032-hydroxyzearalanol (208) were isolated from the marine-derived fungus Penicillium sp. (Pen. sp.) [209) was obtained from the culture broth of the fungus Fusarium sp. 05ABR26 [210\u2013212) were isolated from the culture broth of the marine sponge-derived fungus Aspergillus ostianus (As. ostianus)  [The 14-membered resorcylic acid lactone derivatives 8\u2032-hydroxyzearalanone (en. sp.) . \u03b2-resor 05ABR26 . The cytronesia) .As. sp. SCSGAF 0076 was reported to produce the 16-membered macrolide aspergillide D (213) [214) and enantiomers of curvularin (215\u2013220) were isolated from Curvularia sp. (Cur. sp.) [221) was produced by Cur. sp. isolated from the marine alga Gracilaria folifera and inhibited the growth of B. subtilis, Microbotryum violaceum, Septoria tritici, and Chlorella fusca [The marine-derived fungus  D (213) . Apralacur. sp.) ,138. Thela fusca . 222\u2013224), were purified from an ethyl acetate extract of Corynespora cassiicola isolated from leaf tissues of the Chinese mangrove medicinal plant Laguncularia racemose [225) and B (226) (as the diacetate) were isolated from the fungus Pestalotiopsis spp., which is associated with mangrove twigs of Rhizophora mucronata [227\u2013229), 15G256\u03b1 (230), and 15G256\u03b2 (231) were obtained from crude extracts of the fungus Calcarisporium sp. KF525 isolated from German Wadden Sea water samples [Three decalactones, xestodecalactones D\u2013F , were obtained from the fungus Dendrodochium sp. derived from sea cucumber Holothuria nobilis Selenka in the South China Sea [Thirteen new 12-membered macrolides, dendrodolides A\u2013M  was produced by the gorgonian-derived fungus Cochliobolus lunatus (Coc. lunatus) [Cochliomycin C (lunatus) , its abslunatus) .Pen. sumatrense MA-92, associated with the mangrove Lumnitzera racemose, yielded the sulfur-containing curvularin derivatives sumalarins A\u2212C (246\u2013248) [Coc. lunatus (TA26-46) with histone deacetylase inhibitors led to the elucidation of two 14-membered resorcylic acid lactones: 5-bromozeaenol (249) and 3,5-dibromozeaenol (250) [251\u2013255) were obtained from a sponge-derived fungus Gliomastix sp. ZSDS1-F7-2, their structures being determined by spectroscopy and single crystal X-ray diffraction [256\u2013257) were isolated from the marine-derived fungus Pen. meleagrinum var. viridiflavum [Penicillium sp. DRF2 led to the isolation of cyclothiocurvularins (258\u2013260) and cyclosulfoxicurvularins (261\u2013262) [263) was produced by the mangrove endophytic fungus Cladosporium sp. (Cla. sp.) SCNU-F0001 and its absolute configuration was determined by X-ray diffraction [264\u2013268) were isolated from another mangrove-derived endophytic fungus species in the same Cla. genus [269) was isolated from endophytic fungus Aplosporella javeedii [270\u2013271), were obtained from the endophytic fungus Paramyrothecium roridum isolated from the medicinal plant Morinda officinalis [The fungus 246\u2013248) . Chemicaol (250) . Gliomasfraction . Two 13-diflavum . Applica261\u2013262) . Thioclafraction . Thioclaa. genus . The macjaveedii . Two triicinalis .272) was isolated from a marine bacterium in the order Actinomycetales [273) was produced by Streptomyces hygroscopicus (S. hygroscopicus) isolated from the marine fish Halichoeres bleekeri [O-Succinyl macrolactin F (274) and 7-O-succinyl macrolactin A (275) were isolated from a culture of marine Bacillus sp. (B. sp.) Sc026 [The 24-membered macrolide maduralide  Sc026 .276) was obtained from marine actinomycete L-25-ES25-008 [277) was isolated from marine Streptomycete isolate B7064 and was bioactive in both microorganisms and microalgae [278\u2013279) have been extracted from culture broths of bacteria isolated from the surface of the Caribbean brown alga Lobophora variegata  [280\u2013282) were produced by Micromonospora sp. (M. sp.) and demonstrated inhibition of gastrulation in starfish embryos [Cytotoxic macrolide IB-96212  . Micromo embryos ,164. 283\u2013286) were isolated from actinomycete \u201cMarinispora\u201d. These marinomycins showed antibacterial activity towards methicillin-resistant S. aureus (MRSA), while marinomycin A inhibited vancomycin-resistant S. faecium (VREF) and C. albicans (weakly). Marinomycins A\u2013C demonstrated cytotoxic activity against a panel of 60 tumor cell lines, including six of the eight melanoma cell lines [Marinomycins A\u2013D , with arenicolides A showing moderate cytotoxicity [290) has been reported in a culture of marine Bacillus sp. [Marinispora\u201d yielded polyene macrolides marinisporolides A (291) and B (292), which photoisomerized to the geometric isomers marinisporolides C\u2013E, suggesting that they may be artefacts [S. hygroscopicus  produced halichoblelides B (293) and C (294), which are cytotoxic to tumor cells [Marine actinomycete toxicity . Macrolallus sp. . The actrtefacts . S. hygror cells .295\u2013296), were obtained from the culture of a marine actinomycete S. sp. isolated from a sediment sample collected at North Cat Cay in the Bahamas [Two 36-membered macrolides, bahamaolides A and B ( Bahamas . B. subtilis isolated from marine sediment collected at Gageocho (Republic of Korea) was a source of three new glycosylated methoxy-macrolactins (297\u2013299) [300\u2013302), featuring an oxetane, an epoxide, and a tetrahydropyran ring, were isolated from an ethyl acetate extract of a marine B. sp. [303) was produced by a marine-derived actinomycete strain (CNJ-878) [M. strain FIM07-0019 isolated from shallow coastal waters near the island of Chiloe (Chile) produced a 20-membered macrolide, levantilide C (304) [297\u2013299) . Three ne B. sp. . CytotoxCNJ-878) . The M.  C (304) . S. sp. in sediment from Heishijiao Bay  yielded 11\u2032,12\u2032-dehydroelaiophylin (305) and 11,11\u2032-O-dimethyl-14\u2032-deethyl-14\u2032-methylelaiophylin (306)\u2014both 6-deoxyhexose-containing antibiotics\u2014with the former exhibiting inhibition of MRSA and vancomycin-resistant Enterococci pathogens [307), was produced by a marine-derived filamentous sulfur bacteria, Thioploca sp. [308), was isolated from marine sediment-derived actinomycete S. sp.  [309) and B (310) were identified in marine-derived bacteria of the genus Nocardiopsis and demonstrated inhibition towards TNF-\u03b1-induced NF\u03baB activation (fijiolide A to a greater extent than fijiolide B) [311) and B (312) were obtained from S. hygroscopicus in the alkaline soil of the Saratov region of Russia. They exhibited significant cytotoxicity towards doxorubicin-resistant human leukemia cells [313) and B (314), were isolated from the actinobacterium Catenulispora sp. KCB13F192 [Investigation of a athogens . A rare loca sp. . A poten U.S.A.) . Fijioliolide B) . Astolidia cells . Two hygCB13F192 . Scytonema mirabile BY-8-1, S. burmanicum DO-4-1, and S. ocellatum DD-8-1, FF-65-1 and FF-66-3 have been reported to produce tolytoxin (315). S. burmanicum DO-4-1 also yielded scytophycin B (316), 6-hydroxyscytophycin B (317), 19-O-demethylscytophycin C (318), 6-hydroxy-7-O-methylscytophycin E (319), and scytophycin E (320) [321), was isolated from Japanese Oscillatoria sp. and demonstrated inhibition towards fertilized echinoderm eggs [Lyngbya bouillonii (L. bouillonii) collected on Laing Island (Papua New Guinea) produced lyngbyaloside (322) [323), madangolide (324), and laingolide A (325) [326), for which the configuration of C-11 was later revised [Cyanobacteria  E (320) . A macroerm eggs . The marde (322)  in addit A (325) ,185, and revised ,187.327) and B (328), together with swinholide A previously obtained from the marine sponge T. swinhoei [Geitlerinema sp. collected in Madagascar [329), demonstrating significant molluscicidal activity towards the snail vector Biomphalaria glabrata, was also isolated from L. bouillonii from Papua New Guinea [330) was isolated from L. sp. and showed strong apoptosis-inducing activity in HeLa S3 and HL60 cells [331\u2013333), were produced by another L. cyanobacterium sampled on Tokunoshima Island (Japan) [2+ concentration in HeLa S3 cells [Two glycosylated swinholides, ankaraholides A  . BiselynS3 cells . 334) [335\u2013338), have been isolated from Leptolyngbya sp. collected in Okinawa [The Caribbean Okeania cyanobacterium VQR28MAR11-2 has been reported to produce polycavernoside D (334) , while f Okinawa .339) was isolated from the Okinawan flatworm Amphiscolops sp. (Amphis. sp.) and exhibited cytotoxicity towards murine leukemia cells L1210 and L5178Y [340), G (341) and H (342) were produced by dinoflagellate Amphidinium sp. (Amphid. sp.) associated with the Okinawan flatworm Amphis. breviviridis [Amphidinolide E  and K (344) were isolated from symbiotic dinoflagellate Amphid. sp. and later synthesized [345), B2 (346) and B3 (347) were also isolated from Amphid. sp. [348), M (349) and N (350) [Amphidinolides G and H were elucidated by X-ray diffraction analysis and interconversion . Amphidithesized ,200. Amphid. sp. ,202,203, N (350) ,205,206.351) and P (352), were also isolated from Amphis. sp. [353), showing moderate cytotoxicity towards murine lymphoma L1210 cells in vitro (IC50 6.4 \u03bcg/mL), was obtained from the symbiotic flatworm Amphis. sp. of dinoflagellate Amphid. sp. [R, 7R, 9S, 11R, and 13R on the basis of NMR analysis and a modified Mosher\u2019s method [354) and S (355) were also isolated from Amphid. sp. [356) was obtained from a cultured Amphid. sp. Y-56 isolated from the flatworm Amphis. sp. in Okinawa [357), was also obtained from the Y-56 dinoflagellate strain and exhibited cytotoxicity towards P388, L1210 and KB cells [358) [A. sp. Y-5 produced the 14-membered polyene amphidinolide V (359) [360), T3 (361), T4 (362), and T5 (363) were produced by Amphid. sp. [364), H3 (365), H4 (366), H5 (367), G2 (368), and G3 (369) were produced by Amphid. sp. strain Y-42 isolated from marine acoel flatworms Amphis. sp. The absolute configurations of these compounds were determined by coupling constant data, distance geometry calculations, and chemical means [S)-lactate via a 16-step linear sequence [370) was isolated from an Amphid. sp. and the absolute stereochemistry determined by a combination of J-based configuration analysis and modified Mosher\u2019s method [371) and Y (372) were produced by symbiotic dinoflagellate Amphid. sp. strain Y-42 from Okinawan Amphis. species. Amphidinolide Y exists as a 9:1 equilibrium mixture of the 6-keto- and 6(9)-hemiacetal forms (373). Both amphidinolides X and Y showed significant cytotoxicity against murine lymphoma L1210 and human epidermoid carcinoma KB cells in vitro [374) and B7 (375), were isolated from a culture of a symbiotic dinoflagellate Amphid. sp. from Amphis. sp. and demonstrated cytotoxicity against human B lymphocyte DG-75 cells [376) was isolated from dinoflagellate Amphid. sp. (Y-71 strain) [The structure of amphidinolide N was later revised and stereochemistry assigned . Cytotoxhis. sp. . The abshis. sp. . The 12-hid. sp. . The abss method . Cytotoxhid. sp. . The 20- Okinawa . A 25-meKB cells . Y-56 hals [358) , while t V (359) . Total s V (359) . Analogshid. sp. ,220. Ampal means . Amphidisequence . Amphidis method . Total ss method . Amphidiin vitro ,226. Two75 cells . Amphidi strain) . Amphid. strain S1-36-5 yielded the highly cytotoxic 26-membered caribenolide I (377) [The  I (377) .378) and a 26-membered macrolide amphidinolactone B (379) have been isolated from cultures of Amphid. sp. Amphidinolactone A was synthesized totally via a ring-closing metathesis reaction and the absolute configuration was elucidated [380) and B (381) were isolated from a symbiotic dinoflagellate Symbiodinium sp. (Y-6 strain), which was associated with Amphis. sp. [Prorocentrum maculosum Faust yielded the fast-acting toxin prorocentrolide B (382) [383) was identified in the marine dinoflagellate P. hoffmannianum [384), -1b (385) and -1c (386) were isolated from a marine benthic dinoflagellate Amphid. sp. (strain HYA024) [The 13-membered macrolide amphidinolactone A 37 and a 26ucidated ,231. Thehis. sp. ,233. Bio B (382) . Hoffmanannianum . The 20- HYA024) ,237. 387) was also obtained from Amphid. sp. [388) containing an allyl epoxide was obtained from Amphid. sp. strain HYA024 and was potently cytotoxic to human B lymphocyte DG-75 cells and Epstein\u2013Barr virus (EBV)-infected Raji cells [389) and -5a (390) were isolated from a benthic dinoflagellate Amphid. sp. (strain HYA024) and showed moderate cytotoxicity towards human B lymphocytes DG-75 [391) and -11a (392) were cytotoxic towards human cervix adenocarcinoma HeLa and murine hepatocellular carcinoma MH134 cells [The cytotoxic 23-membered iriomoteolide-2a (hid. sp. . The 15-ji cells . Iriomotes DG-75 . The 15-34 cells .393) and -12a (394) were isolated from a marine dinoflagellate Amphid. sp. (KCA09053 strain) with iriomoteolide-10a being cytotoxic to human cervix adenocarcinoma HeLa and murine hepatocellular carcinoma MH134 cells [395) was isolated from the symbiotic dinoflagellate Symbiodinium sp. (S. sp.) and showed significant voltage-dependent N-type Ca2+ channel-opening activity at 7 nM and immediately ruptured the surface tissue of the acoel flatworm Amphis. sp. at 2.5 mM [S,18R,21R) configurations were determined by synthesis [S,40S) and (C-1\u2032\u2013C-25\u2032) [396) caused potent stimulation of actomyosin ATPase activity [397) was isolated from cultures of the marine dinoflagellate Prorocentrum belizeanum [398) was extracted and purified from a culture of dinoflagellate Alexandrium ostenfeldii from the Baltic Sea [399) was isolated from a culture of the symbiotic marine dinoflagellate S. sp. [Iriomoteolide-10a  ,244,245.activity . The 25-lizeanum . Gymnodiltic Sea . Symbiode S. sp. .Polycavernosa tsudai (Gracilaria edulis) contained the macrolide polycavernoside A (400), which led to human illness and death in Guam [401), A3 (402), B (403) and B2 (404) were also obtained from Polycavernosa red algae [ in Guam . The rel in Guam ,252. Its in Guam . Polycaved algae . 405) and C2 (406), were isolated from the red alga Gracilaria edulis (G. edulis) [407\u2013409) were isolated from extracts of red alga G. coronopifolia [410) and manauealide C were extracted from Hawaiian G. coronopifolia [Callophycus serratus (C. serratus) led to the isolation of three diterpene-benzoate natural products: bromophycolides A (411) and B (412), and a nonhalogenated compound (413). Bromophycolides A and B exhibited moderate antibacterial and antifungal properties while bromophycolides A demonstrated potent anti-HIV and moderate cytotoxic activities [414\u2013420) were also isolated from extracts of C. serratus. All the bromophycolides exhibited modest antineoplastic activity towards a range of human tumor cell lines while bromophycolides F and I showed weak antifungal activity [Two analogs of polycavernosolide A, polycavernosides C ( edulis) . Manaueaopifolia . Anhydroopifolia . Investitivities . Bromophactivity . C. serratus extract yielded a series of unusual antimalarial diterpene-benzoate macrolides, bromophycolides J\u2013Q (421\u2013428), with a range of moderate to strong antimicrobial and anticancer properties [C. serratus was also a source of the diterpene-benzoate macrolides bromophycolides R\u2013U (429\u2013432). These demonstrated modest cytotoxicity toward selected human cancer cell lines while bromophycolide S was active (at submicromolar concentrations) against the human malaria parasite Plasmodium falciparum  [Further investigation of the operties . C. serrciparum) . 433) and B (434) were obtained from the Fijian red alga Neurymenia fraxinifolia [Ecklonia stolonifera produced ecklonialactones C (435) and D (436) containing a 14-membered lactone moiety, and ecklonialactones E (437) and F (438), with a 16-membered moiety [439\u2013446) with a macrolide scaffold and one cymathere-type oxylipin with an open ring were isolated from the brown alga Eisenia bicyclis. The absolute configurations of compounds 439\u2013443 and 446 were determined by NMR spectroscopy with the relative stereochemistry at C-9 in 446 remaining unassigned [447) was isolated from the crustose coralline alga Hydrolithon reinboldii and its absolute relative configuration was determined by NMR spectroscopy with the relationships of the two side chains to the macrolide ring remaining unassigned [The \u03b1-pyrone macrolides neurymenolides A  and B1e (449), exhibiting moderate antifouling activity were obtained from Anthrogorgia caerulea collected in the South China Sea [Two avermectin congeners, avermectins B1c (hina Sea .450) from the marine bryozoan Bugula neritina (L.) was carried out to provide material for clinical study [451) has been converted to bryostatin 1 and bryostatin 12 (452) by selective protection and deprotection involving the C-26 hydroxyl group [p-bromobenzoate (453) [1H- and 13C-NMR were later revised [B. neritina and reinvestigation of 2D NMR spectroscopic data revised the structure of bryostatin 3 to structure 454 [Large-scale isolation of bryostatin 1  , while t revised . Bryostature 454 .B. neritina led to the identification of bryostatins 14 (455) and 15 (456) [457) have been elucidated by NMR spectroscopy [458) was determined to be the major cytotoxic component of B. neritina [459), 17 (460), and 18 (461), were isolated in trace amounts from B. neritina from the Gulf of Mexico [462) was isolated from B. neritina collected from the South China Sea [463), was produced by the larvae of B. neritina and its structure determined by spectral comparison with previously described bryostatins [464), which was cytotoxic towards the P388 lymphocytic leukemia cell line [Further investigation of 15 (456) . The strtroscopy ,273,275.troscopy . Bryostaneritina . Three af Mexico . Antineohina Sea . A furthostatins . Bioassaell line .465) was isolated from an extract of the sea hare Stylocheilus longicauda and synthesized [Aplysia kurodai Baba contained the novel and potently cytotoxic macrolides aplyronines A (466), B (467) and C (468). The absolute configuration of aplyronine A was assigned following enantioselective synthesis of its degradation products and total synthesis was also reported [469\u2013473), were also isolated from the Japanese sea hare Aplysia kurodai [474) and the diacetyl derivative dolabelide B (475), both cytotoxins, were obtained from the Japanese sea hare Dolobella auricularia [476) and D (477) were also isolated from Dolabella auricularia, the originally assigned structure of dolabelide D being confirmed by total synthesis [478\u2013482) were found in the skin of the marine mollusk Aplysia depilans, and were ichthyotoxic to the mosquito fish Gambusia affinis [483) and 7 (484) were isolated from Patinopecten yessoensis scallops [485) as a pectenotoxin accumulating in Norwegian blue mussels (Mytilus edulis) and cockles (Cerastoderma edule) [486), containing a 14-membered macrocyclic lactone linked to a 2,4-di-O-methyl-L-R-rhamnopyranoside, was found in the Gulf of California in the shell-less mollusk Dolabella auricularia [Aplysiatoxin  . Dolastaicularia . The steicularia .487) and D (488), were isolated from the Okinawan tunicate Eudistoma cf. rigida [489) and B (490), were reported in the tunicate Aplidium lobatum [A. lobatum from shallow waters in Australia, A. sp. from deep water, and an unidentified Philippine ascidian have been reported as sources of a series of macrolides, lobatamides C\u2013F (491\u2013494), demonstrating cytotoxicity towards human tumor cell lines [495) was obtained from an Okinawan ascidian L. sp. by bioassay-guided isolation and was shown to inhibit the first cleavage of fertilized sea urchin eggs at 0.01 \u03bcg/mL [Didemnidae sp. was the source of the macrolides biselides A (496) and B (497) [D. sp. led to the isolation of biselides C (498), D (499) and E (500) which exhibited cytotoxicity against human cancer cells NCI-H460 and MDA-MB-231 [501) was obtained from the Antarctic tunicate Synoicum adareanum [Two 24-membered macrolide sulfates showing antineoplastic activity, iejimalides C  . FurtherA-MB-231 . Cytotoxdareanum  and its dareanum ,307. 502\u2013505) were isolated from Lissoclinum ascidian collected in Algoa Bay near Port Elizabeth and the surrounding Nelson Mandela Metropole in South Africa [Glycosylated macrolides mandelalides A\u2212D (h Africa .The biological activities of marine-derived macrolides have been studied extensively. As listed in This review presents a summary of 505 marine-derived macrolides reported from 1990 to 2020 and highlights their chemical and biological diversity. As shown in For macrolides with larger macrocyclic rings, such as reidispongiolides A and B , symbiod"}
+{"text": "ADG20 is a fully human IgG1 monoclonal antibody engineered to have potent and broad neutralization against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other SARS-like CoVs with pandemic potential as well as an extended-half-life. ADG20 is administered intramuscularly (IM). A QSP/PBPK model was constructed to support dose selection for a COVID-19 Phase 2/3 prevention trial (EVADE: NCT04859517).90]) for authentic SARS-CoV-2 were also evaluated.A QSP/PBPK model and a CDC reference adult body weight distribution (45\u2013150 kg) were used to simulate 1000 concentration-time profiles for candidate single-dose regimens of ADG20 (150\u2013450 mg IM). As serum virus neutralizing antibody (sVNA) titers are reportedly a key correlate of protection from COVID-19, a regression equation between time-matched serum ADG20 concentrations (following a 300 mg IM dose) and sVNA titers was developed using measured titers against authentic SARS-CoV-2 determined by a plaque reduction neutralization assay. Projected ADG20 serum concentrations relative to neutralization potency in vitro .The measured 50% neutralization titer  was 1382 (32.7%) 13 days after a single 300 mg IM dose of ADG20. This was within the range of peak sVNA titers reported for COVID-19 vaccine recipients. Using the linear equation relating serum ADG20 concentration to time matched individual MN50 titers and the QSP/PBPK median PK prediction, the anticipated median MN50 exceeded the threshold for protection from SARS-CoV-2 infection established in a non-human primate adoptive transfer model for up to 52 weeks. Based on the QSP/PBPK median PK prediction, median ADG20 serum concentrations are projected to remain >100-fold above the ADG20 ICFollowing administration of a single 300 mg IM dose, sVNA titers and concentrations of ADG20 are projected to remain above thresholds anticipated to be required for protection against COVID-19 for up to 52 weeks. These data support the evaluation of a single ADG20 300 mg IM dose for the prevention of COVID-19.Figure. QSP/PBPK model forecast of ADG20 300 mg IM in adults.Predicted median serum ADG20 concentration is shown with the dotted line representing 100\u00d7 in vitro IC90 of 0.011 mg/L or 1.1 mg/L; the solid black line represents the simulated median; the shaded area represents the 90% prediction interval. The predicted median half-life of ADG20 300 mg IM exceeded 74 days. PBPK model inputs include Ln-normal Kd,FcRn of 9.55 nM (10% IIV); IM bioavailability of 100%; 15% IIV on muscle lymph RC; and Centers for Disease Control and Prevention weight distribution of 45\u2013150 kg. FcRn, neonatal Fc receptor; IIV, inter-individual variability; Kd, dissociation constant; Ln, log-normal; RC, reflection coefficient.Scott A. Van Wart, PhD, Adagio Therapeutics, Inc. (Independent Contractor) Evan D. Tarbell, PhD, Adagio Therapeutics, Inc. (Independent Contractor) Kristin Narayan, PhD, Adagio Therapeutics, Inc. (Employee) Laura M. Walker, PhD, Adagio Therapeutics, Inc.  Lynn E. Connolly, MD, PhD, Adagio Therapeutics, Inc. (Employee) Paul G. Ambrose, PharmD, Adagio Therapeutics, Inc. (Employee)"}
+{"text": "Integrase inhibitor (INSTI)-based antiretroviral therapies (ART) are first-line for HIV infection. Recent studies have identified metabolic complications of INSTI regimens, especially when TAF is included. The objective of this study was to assess lipid changes associated with INSTI-based ART.This was a retrospective, observational, single-center study. Patients age \u2265 18 with HIV infection, receiving care Jan 2004 to Jul 2019, and prescribed the same ART for \u2265 6 consecutive months were randomly selected from a computer-generated list. Unique ART regimens prescribed per patient were considered \u201cexposures\u201d. Patients were followed up to 2 years per exposure. INSTI-based exposures included an INSTI plus one or more NRTI. Non-INSTI-based exposures included a PI or NNRTI plus one or more NRTI. Lipid panel results were recorded before and during exposures. Lipid changes with INSTI and non-INSTI exposures were compared. Other metabolic outcomes were recorded.2, respectively. 72 (37.5%) were ART-na\u00efve. 53.5% INSTI recipients received BIC or DTG. Hypertriglyceridemia (HTG) occurred more often during INSTI exposures . Mean change in TG concentration was +10.1 and -15.0 mg/dL for INSTI and non-INSTI regimens, respectively (p = 0.105). Patients receiving INSTIs were more commonly prescribed a new lipid-lowering therapy (LLT) or an increase in dose of preexisting LLT . Of those that received an INSTI-regimen and either TAF, TDF, or no tenofovir, 17.4%, 10.3%, and 8.3% developed HTG, respectively (p = 0.49). There was no difference between groups in HDL or LDL changes. Among INSTI and non-INSTI groups, weight increased by +3.8 and +2.1 kg, respectively (p = 0.129). Mean weight change was less in INSTI recipients whose regimen included TDF .192 exposures were identified  among 132 patients. The majority were Black (69.3%) and male (61.9%). Mean age and BMI were 42.0 \u00b1 11.3 and 26.2 \u00b1 5.5 kg/mTreatment with INSTI-based ART was associated with HTG. LLTs were utilized more often among INSTI recipients, and this limited our ability to fully characterize lipid changes in this real-world study. Weight gain among INSTI recipients may be attenuated if TDF is included in the regimen.James Johnson, PharmD, FLGT (Shareholder) Caryn Morse, MD, Chimerix (Scientific Research Study Investigator)Covis Pharma (Scientific Research Study Investigator)Gilead Sciences Inc. (Scientific Research Study Investigator)Ridgeback Biotherapeutics (Scientific Research Study Investigator)Roche (Scientific Research Study Investigator)SCYNEXIS, Inc. (Scientific Research Study Investigator)Theratechnologies (Advisor or Review Panel member)Viiv (Advisor or Review Panel member)"}
+{"text": "Staphylococcus lugdunensis and determine its association with sequence types (STs) determined by multilocus sequence typing (MLST) and oxacillin susceptibility. Primers were designed to detect and sequence types IIIA and IIC CRISPR-Cas in 199 S. lugdunensis isolates. MLST and oxacillin susceptibility tests were also performed on the isolates. We found that 84 S. lugdunensis isolates had type IIIA CRISPR-Cas, while 46 had type IIC. The results showed a strong association between STs and CRISPR-Cas types. The ST1, ST6, ST12, and ST15 isolates had type IIIA CRISPR-Cas systems, and the ST4, ST27, and ST29 isolates had type IIC CRISPR-Cas. Interestingly, of 83 isolates containing type IIIA CRISPR-Cas, 17 (20.5%) were oxacillin-resistant S. lugdunensis (ORSL), and all of these ORSL isolates belonged to ST6 cluster 1. Moreover, spacers 23 and 21 were found in 16 and 17 ORSL isolates, respectively. In contrast, all 46 isolates with type IIC CRISPR-Cas were susceptible to oxacillin. Our results showed that 41.3% of CRISPR-Cas IIIA spacers were homologous to plasmids and 20.2% were homologous to phages. However, in type IIC CRISPR-Cas, 11.8% and 39.9% of spacers showed sequence homology with plasmids and phages, respectively. In conclusion, we found that the distribution and composition of the CRISPR-Cas system in S. lugdunensis was associated with STs and oxacillin susceptibility.Clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) genes (CRISPR-Cas) are present in many bacterial genomes with functions beyond adaptive immunity. We aimed to characterize the CRISPR-Cas system in the pathogenic Gram-positive bacterium IMPORTANCE CRISPR-Cas systems have been characterized as playing several biological roles in many bacterial genomes. Moreover, CRISPR-Cas systems are useful for epidemiological, diagnostic, and evolutionary studies of pathogenic bacteria. However, the characteristics of CRISPR-Cas systems in Staphylococcus lugdunensis have been rarely reported. In this study, we revealed that type IIIA CRISPR-Cas was dominant in S. lugdunensis isolates, followed by type IIC CRISPR-Cas. Moreover, the composition of CRISPR-Cas spacers was strongly associated with multilocus sequence typing and oxacillin susceptibility of S. lugdunensis. These results advance our understanding of the evolution of CRISPR-Cas systems; however, the biological functions of CRISPR-Cas systems in S. lugdunensis remain to be further characterized. Staphylococcus lugdunensis is a Gram-positive, catalase-positive, and coagulase-negative Staphylococcus (CoNS) bacterium. Staphylococcus lugdunensis has emerged as an important pathogen, implicated in clinically invasive infections such as endocarditis and bacteremia in recent years  has been frequently reported over the last decade (Staphylococcus lugdunensis expressing penicillin-binding protein 2a (PBP2a), encoded by the mecA gene, shows a lower affinity for \u03b2-lactams and is responsible for \u03b2-lactam resistance   , 5.S. lugdunensis isolates from different clinical and geographic sources, using DNA sequence analysis of seven housekeeping genes: aroE, dat, ddl, gmk, ldh, recA, and yqiL (S. lugdunensis isolates could be defined into 20 sequence types (STs) and 5 clonal complexes  and CRISPR-associated (Cas) genes (CRISPR-Cas) are present in many bacterial and archaeal genomes . CRISPR-lococcus \u201313. Rossococcus \u2013, 15. HowS. lugdunensis  isolates and determined the distribution of CRISPR-Cas in the isolates. The PCR results showed that 130/199 (65.3%) of the S. lugdunensis isolates had CRISPR-Cas systems, including 84 (42.2%) with type IIIA and 46 (23.1%) with type IIC . Most ST1 and ST6 clusters (clusters 1 to 5) had spacers 1 and 2, which showed homology with Campylobacter phage CP220 and Clostridium botulinum strain CDC_67071 plasmid pNPD7, respectively. All ST15 cluster 7 isolates had spacers 94, 95, 96, and 97, of which 94 and 96 showed homology with phages in the S. lugdunensis genome . We were unable to determine the homologs of spacers 12 and 81.To further validate the association between CRISPR-Cas types and STs, we performed a phylogenetic analysis based on the sequences of seven housekeeping genes for MLST. The results revealed a strong association between ST1, ST6, ST12, and ST15 , which had type IIIA CRISPR-Cas systems . Among tisolates . Spacer s genome . Spacer S. lugdunensis isolates containing type IIIA CRISPR-Cas systems was 10.2 (818 spacers/80 isolates) (S. lugdunensis chromosome (The mean number of spacers in solates) . We founsolates) . Moreoveromosome . Overallromosome .S. lugdunensis isolates with type IIC CRISPR-Cas comprised 8 clusters and 33 CTs (S. lugdunensis genome (Table\u00a0S5). All ST29 isolates had spacers 55 and 56 (Secundilactobacillus paracollinoides strain TMW 1.1979 plasmid pL11979-2 (Table\u00a0S5). We were unable to determine the homologs of spacer 56. Spacers 20, 21, 29, 31, 32 to 36, 44 to 46, and 53 were highly conserved in ST4 isolates, but ST4 cluster 1 lacked spacer 53 (Staphylococcus genomes (Table\u00a0S5).Forty-three d 33 CTs . Three id 33 CTs  and 3B. d 33 CTs . Howeverd 33 CTs . Spacers5 and 56 . Spacer pacer 53 . Among tS. lugdunensis containing type IIC CRISPR-Cas systems was 13.7 (591 spacers/43 isolates) (S. lugdunensis containing type IIIA CRISPR-Cas systems (mean 10.2 spacers/strain) (S. lugdunensis chromosome (All isolates with type IIC CRISPR-Cas were susceptible to oxacillin. The mean number of spacers in solates) , higher /strain) . Our res/strain) . Moreoveromosome . Overallromosome . Of spacromosome .S. lugdunensis. We found 84 S. lugdunensis isolates with type IIIA CRISPR-Cas and 46 isolates with type IIC  and related IncF epidemic plasmids . Surprisingly, spacer 23 showed homology with a S. aureus strain 16405 plasmid that expressed PBP2A encoded by mecA. Therefore, the activity of CRISPR-Cas systems in plasmid acquisition and their association with oxacillin susceptibility of S. lugdunensis merits further investigation.Our results demonstrated that CRISPR-Cas spacer sequences were strongly associated with MLST and oxacillin susceptibility. Previous studies have shown that pandemics caused by multidrug-resistant plasmids . The absplasmids , 16. In isolates . In addiisolates . The homS. lugdunensis typing, including a classical length-based multiple loci VNTR analysis (MLVA) method and a sequence-based MLVA method known as the tandem repeat sequence typing (TRST) method (fbl-typing) has also been used for S. lugdunensis typing  method . Sequenc) method . DNA seqs typing . Among tdunensis , 9. CompAcinetobacter baumannii global clone 1 (GC1) isolates  were included in the study. All information associated with the 199 isolates  was collected. Staphylococcus lugdunensis isolates were initially identified by Gram staining, biochemical methods , and rapid PCR detection . The isolates were stored in tryptic soy broth containing 20% glycerol at \u221280\u00b0C until use.A total of 199 etection . All thecas1 gene for CRISPR-Cas types IIIA and IIC, respectively  database, including sequences for type IIIA-positive strain VISLISI_33 (accession no. CP020769.1) and type IIC-positive strain C_33 (accession no. CP020768.1). The PCRs were carried out in a total volume of 20\u2009\u03bcl containing 2\u00d7 master mix , 10\u2009pmol of each primer, and 1\u2009\u03bcl DNA template. The PCR cycling conditions for CRISPR-Cas system detection were 95\u00b0C for 3\u2009min; 30 cycles of 30 s at 95\u00b0C, 30 s of annealing at 50\u00b0C, and 1\u2009min of extension at 72\u00b0C; and a final extension for 3\u2009min at 72\u00b0C.The Crispr-IIIA-F/Crispr-IIIA-R and Crispr-IIC-F/Crispr-IIC-R primers were designed to specifically detect the The primers Crispr-IIIA-F/Crispr-IIIA-R and Crispr-IIC-F/Crispr-IIC-R were designed for also determining type IIIA and type IIC CRISPR-Cas spacer sequences, respectively  for ST type determination. Sequence types were identified based on the allele profiles. Further analysis was performed using eBURST (http://eburst.mlst.net) to identify clonal complexes (CCs) and founders, as well as to determine the overall population structures. A minimum-evolution (ME) tree of the concatenated sequences (aroE-dat-ddl-gmk-ldh-recA-yqiL) for each ST shared by the S. lugdunensis isolates was generated using Mega X and the Kimura two-parameter model to estimate genetic distances. Statistical support of the nodes in the ME tree was assessed by performing 1,000 bootstrap resamplings.MLST was performed for all us study . Seven hS. lugdunensis isolates to oxacillin was determined using an agar dilution assay and interpreted according to the Clinical and Laboratory Standards Institute guidelines (Staphylococcus lugdunensis isolates with a MIC\u2009\u2265\u20094\u2009\u03bcg/mL were defined as resistant. Staphylococcus aureus ATCC 29213 was used as a control strain. Antimicrobial susceptibility testing was performed in duplicate to ensure reproducibility.Susceptibility of the idelines . StaphylS. lugdunensis isolates were subjected to SCCmec typing and mecA detection using a multiplex PCR assay to amplify the ccr and mec complexes according to a previous study (All us study ."}
+{"text": "Arabidopsis thaliana, Gossypium herbaceum (A1), Gossypium arboreum (A2), Gossypium raimondii (D5), and Gossypium hirsutum (AD1) genomes demonstrated that four HSF genes were inherited from a common ancestor, A0, of all existing cotton A genomes. Members of the HSF gene family in G. herbaceum (A1) genome indicated a significant loss compared with those in G. arboretum (A2) and G. hirsutum (AD1) A genomes. However, HSF genes in G. raimondii (D5) showed relative loss compared with those in G. hirsutum (AD1) D genome. Analysis of tandem duplication (TD) events of HSF genes revealed that protein-coding genes among different cotton genomes have experienced TD events, but only the two-gene tandem array was detected in Gossypium thurberi (D1) genome. The expression analysis of HSF genes in G. hirsutum (AD1) and Gossypium barbadense (AD2) genomes indicated that the expressed HSF genes were divided into two different groups, respectively, and the expressed HSF orthologous genes between the two genomes showed totally different expression patterns despite the implementation of the same abiotic stresses. This work will provide novel insights for the study of evolutionary history and expression characterization of HSF genes in different cotton genomes and a widespread application model for the study of HSF gene families in plants.Heat shock transcription factors (HSFs) are involved in environmental stress response and plant development, such as heat stress and flowering development. According to the structural characteristics of the HSF gene family, HSF genes were classified into three major types  in plants. Using conserved domains of HSF genes, we identified 621 HSF genes among 13 cotton genomes, consisting of eight diploid and five tetraploid genomes. Phylogenetic analysis indicated that HSF genes among 13 cotton genomes were grouped into two different clusters: one cluster contained all HSF genes of HSFA and HSFC, and the other cluster contained all HSF genes of HSFB. Comparative analysis of HSF genes in  Gossypium) is one of the most important economic crops worldwide, which provides the major resource of natural fiber for human beings over the past decades. The genus Gossypium contains over 45 diploid (designated as A to G and K) and 7 tetraploid (designated as AD1 to AD7) species (Gossypium herbaceum (A1) (Gossypium arboreum (A2) (Gossypium thurberi (D1) (Gossypium raimondii (D5) (Gossypium turneri (D10)   (Gossypioides kirkii (K) (Gossypium hirsutum (AD1) (Gossypium barbadense (AD2) (Gossypium tomentosum (AD3) (Gossypium mustelinum (AD4) (Gossypium darwinii (AD5) (Bombax ceiba at approximately 24 million years ago (Mya). Subsequently, cotton A0 and D5 genome species diverged from their ancestor at approximately 4.8 Mya. After the split of their ancestor, cotton A0 and D5 genome species crossed and generated cotton allotetraploid AD genome species at approximately 1.6 Mya (G. hirsutum (AD1) and G. barbadense (AD2) (G. herbaceum (A1) and G. arboreum (A2) (Cotton (genus  species . To facieum (A1) , Gossypieum (A2) , Gossypieri (D1) , Gossypidii (D5) , Gossypiri (D10) , Gossypilyx (F1) , Gossypiale (G2) , and Gosrkii (K) , and fivum (AD1) , Gossypise (AD2) , Gossypium (AD3) , Gossypium (AD4) , and Gosii (AD5) . Previou 1.6 Mya . Later, se (AD2) . Less theum (A2) . A cleareum (A2) .Solanum lycopersicum (24)  (Oryza sativa (25) (Zea mays (25) (Glycine max (26) (Brassica oleracea (35) (Brassica rapa (36) (Brassica napus (64) (Sesamum indicum (30) (Prunus mume (18) (Fagopyrum tataricum (29) (Cicer arietinum (20) (Vitis vinifera (19) (Capsicum annuum (25) (Brassica juncea (60) (G. hirsutum (AD1) D genome using EST assembly and genome-wide analyses (Heat shock transcription factors (HSFs) serve as the major activators of heat shock proteins (HSPs) with the help of binding to the promoter regions of HSP genes in plants. This enables HSP genes to regulate transcription in response to heat stress for adapting to various environmental dynamic changes . HSF procum (24) , Arabidoana (21) , Oryza siva (25) , Zea mayays (25) , Glycinemax (26) , Brassiccea (35) , Brassicapa (36) , Brassicpus (64) , Sesamumcum (30) , Prunus ume (18) , Fagopyrcum (29) , Cicer anum (20) , Vitis vera (19) , Capsicuuum (25) , and Bracea (60) . For theanalyses .G. herbaceum (A1), G. arboreum (A2), G. raimondii (D5), and G. hirsutum (AD1), we detected the formation of the HSF gene family and traced the evolution of HSF genes in G. hirsutum (AD1) genome. According to the genome-wide analysis of TD events, we investigated the influence of TD events on the generation of HSF genes among 13 cotton genomes. However, we used the expression analysis to reveal the functional differences of HSF genes under the same abiotic stresses in G. hirsutum (AD1) acc. TM-1 and G. barbadense (AD2) acc. Hai7124. This project will provide novel insights for understanding the evolutionary history and expression characterization of the HSF gene family, paving the way for genetic breeding and molecular improvement of cotton diploid and tetraploid species.In this project, we performed genome-wide comparative genomics analysis for the HSF gene family among 13 cotton genomes. Based on the evolutionary relationship among G. herbaceum , G. arboreum , G. thurberi , G. raimondii , G. turneri , G. longicalyx , G. australe , and G. kirkii , and tetraploid genomes, G. hirsutum , G. barbadense , G. tomentosum , G. mustelinum , and G. darwinii , are downloaded from CottonGen1  acc. TM-1 and G. barbadense (AD2) acc. Hai7124 are retrieved from the Sequence Read Archive (SRA) database with accession number PRJNA490626  program with \u201ctrusted cutoff\u201d as threshold . All proA. thaliana and Gossypium species followed the same procedures.ClustalW2 was employed to perform MSA with protein sequences of target HSFs from 13 different cotton species. Then, the MSA file with \u201cmeg\u201d format were used to construct the phylogenetic tree through MEGA 7 with maximum likelihood (ML) statistical method and 1,000 bootstrap replications . The phyFollowing the identification of tandemly duplicated genes in PTGBase, all-against-all BLAST of protein sequences in 13 cotton species was employed to identify the paralogous gene pairs within single cotton species with E-value cut-off \u22641e-20 . After cG. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes, as well as between G. raimondii (D5) and G. hirsutum (AD1) D genomes, were detected by the MCscanX software and validated by phylogenetic analysis (G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes and between G. raimondii (D5) and G. hirsutum (AD1) D genomes. So, phylogenetic analysis of these four cottons and A. thaliana genomes was used to validate collinear relationships among cotton genomes. The collinear analysis of G. hirsutum (AD1) and G. barbadense (AD2) A genomes, as well as G. hirsutum (AD1) and G. barbadense (AD2) D genomes, follows the above procedures.Orthologous gene pairs between analysis . First, G. hirsutum (AD1) and G. barbadense (AD2) reference genome sequences through HISAT (version 2.2.1)  , and alln 2.2.1) . The Strn 2.2.1) . The hien 2.2.1) .G. herbaceum , with 17 in G. arboreum , 29 in G. thurberi , 33 in G. raimondii , 36 in G. turneri , 38 in G. longicalyx , 36 in G. australe , 29 in G. kirkii , 78 in G. hirsutum , 78 in G. barbadense , 84 in G. tomentosum , 79 in G. mustelinum , and 80 in G. darwinii  genomes (G. herbaceum (A1) has the least number of HSF genes and G. australe (G2) has the greatest number of HSF genes in cotton diploid genomes. Out of five cotton tetraploid genomes, G. hirsutum (AD1) and G. barbadense (AD2) have the least number of HSF genes, and G. tomentosum (AD3) has the greatest number of HSF genes.With the development of genome sequencing technology, more and more cotton genome sequences were released for the community, which facilitate the analysis of key gene families within plant whole genomes. Totally, 13 latest cotton genomics data, consisting of eight diploid and five tetraploid genomes, were downloaded from CottonGen , G. tomentosum (AD3), G. mustelinum (AD4), and G. darwinii (AD5) genomes, respectively. Finally, we obtained 78 representative HSF genes in G. barbadense (AD2) genome, with 84, 79, and 80 HSF genes in G. tomentosum (AD3), G. mustelinum (AD4), and G. darwinii (AD5) genomes, respectively.To identify accurately the members of the HSF gene family, we used the entire available protein sequences in different cotton genomes. After curation, we found that several cotton tetraploid genomes released multiple protein sequences for one gene due to an alternative splice in genomes. Using the profile HMMs of HSF_DNA-bind, we identified 120, 135, 122, and 106 HSF protein sequences in Based on the characteristics of conserved domains of HSF genes, we obtained 621 target HSF genes among 13 cotton genomes. With protein sequences of the identified HSF genes, we constructed a phylogenetic tree to investigate the evolutionary relationship of all HSF genes among 13 cotton genomes. Following the phylogenetic tree, all HSF genes were grouped into two different clusters, including I and II clusters, among 13 cotton genomes . A previG. hirsutum (AD1), was generated from hybridization between G. raimondii (D5) and the common progenitor, A0 genome, of G. herbaceum (A1) and G. arboreum (A2). However, out of four HSF genes in G. herbaceum (A1) genome, two HSF genes belonged to HSFA, with one HSF gene belonging to HSFB and one HSFC gene belonging to HSFC. For HSF genes in G. arboreum (A2) genome, six HSF genes were classified into HSFA, with eight HSF genes belonging to HSFB and three HSF genes belonging to HSFC. These HSF genes in G. herbaceum (A1) and G. arboreum (A2) originated from a common progenitor of all existing A genomes, including A genome in G. hirsutum (AD1). So, different types of HSF genes in G. herbaceum (A1) genome indicated a significant loss compared with G. arboreum (A2) and G. hirsutum (AD1) A genomes. However, different types of HSF genes in G. raimondii (D5) experienced relative loss compared with those in G. hirsutum (AD1) D genome.Upland cotton, Arabidopsis thaliana is an important model plant, and its genomic data were released more than 20 years, which brings opportunity for studying HSF genes in A. thaliana , G. arboreum (A2), G. raimondii (D5), and G. hirsutum (AD1) genomes to perform comparative genomics analysis of HSF genes in A. thaliana and cotton genomes. Through phylogenetics analysis of HSF genes in A. thaliana and cotton genomes, all HSF genes among five species were clustered into two different groups, namely the I and II groups  has two orthologous genes (Ghe05G03320 and Ghe08G31530) in G. herbaceum (A1) genome, two orthologous genes (Gar05G03090 and Gar08G31350) in G. arboreum (A2) genome, two orthologous genes (D5.v1.pred_00000361-RA and D5.v1.pred_00033482-RA) in G. raimondii (D5) genome, and three  and two (Ghi_D05G01481 and Ghi_D07G00601) orthologous genes in G. hirsutum (AD1) A and D genomes, respectively (G. arboreum (A2) genome, three orthologous genes  in G. raimondii (D5) genome, and three  and three  orthologous genes in G. hirsutum (AD1) A and D genomes, respectively , two genes (Gar02G18780 and Gar10G10520) in G. raimondii (D5) genome, three genes  in G. raimondii (D5) genome, and three  and three  genes in G. hirsutum (AD1) A and D genomes were clustered together and divided into two groups following the divergence from the common ancestor of A. thaliana and Gossypium lineage  was identified to belong to HSFC. Through the analysis of phylogeny, AT3G24520.1 has one orthologous gene (Ghe06G07640) in G. herbaceum (A1) genome, three orthologous genes  in G. arboreum (A2) genome, three orthologous genes  in G. raimondii (D5) genome, and three  and three  in G. hirsutum (AD1) A and D genomes, respectively (In ectively . AT3G029ectively . This re lineage . In A. tectively .G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes and between G. raimondii (D5) and G. hirsutum (AD1) D genomes. This result will help to trace the evolution of HSF genes in G. hirsutum genome clearly. In G. hirsutum (AD1) genome, 78 HSF genes were identified, namely 40 and 38 HSF genes distributed into A and D genomes, respectively. From the comparisons between G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes, we got 11 and 64 all-against-all HSF gene pairs, respectively , G. arboreum (A2), G. raimondii (D5), and G. hirsutum (AD1), four HSF genes in G. herbaceum (A1) genome were investigated to get four HSF orthologous genes in G. hirsutum (AD1) A genome, and 17 HSF genes in G. arboreum (A2) genome and 17 HSF orthologous genes in G. hirsutum (AD1) A genome (G. hirsutum (AD1) A genome were detected to have orthologous genes in G. herbaceum (A1) and G. arboreum (A2) genomes, which represented 42.5% of the total HSF genes in G. hirsutum (AD1) A genome. A previous study indicated that all existing A genomes in Gossypium lineage originated from a common progenitor, A0 genome, and G. herbaceum (A1) and G. arboreum (A2) genome experienced independent evolution after the split of the A0 genome (G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genome existed in ancestral genome A0 and remained in G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes although they experienced independently long-term evolutionary history, but HSF genes in G. herbaceum (A1) genome missed more according to orthologous analysis between G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes.To trace the evolutionary history of HSF genes in the Upland cotton genome, we combined the MCscanX software and validation of phylogeny to confirm HSF orthologous gene pairs between ectively . CombineA genome  were obt0 genome . FollowiG. hirsutum (AD1) D genome, 33 of 38 HSF genes were detected to have orthologous genes in G. raimondii (D5) genome, representing 86.8% of the total HSF genes in G. hirsutum (AD1) D genome. For G. raimondii (D5) genome, all HSF genes were detected to have orthologous genes in G. hirsutum (AD1) D genome. These results meant that members of the HSF gene family in G. raimondii (D5) genome and G. hirsutum (AD1) D genome kept excellent collinear relationships after the formation of G. hirsutum (AD1), but HSF genes in G. hirsutum (AD1) D genome showed significant expansion compared with those in G. raimondii (D5) genome.In G. herbaceum, G. arboreum, G. thurberi, G. raimondii, G. turneri, G. longicalyx, G. australe, G. kirkii, G. hirsutum, G. barbadense, G. tomentosum, G. mustelinum, and G. darwinii, representing 12.9, 11.83, 19.79, 14.76, 12.49, 21.03, 14.71, 13.44, 10.27, 34.66, 33.94, 35.23, and 27.75% of total protein-coding genes in G. herbaceum, G. arboreum, G. thurberi, G. raimondii, G. turneri, G. longicalyx, G. australe, G. kirkii, G. hirsutum, G. barbadense, G. tomentosum, G. mustelinum, and G. darwinii genomes (G. thurberi (D1) genome were identified following the identification of tandemly duplicated genes in plants. However, for the remaining 12 cotton genomes, we did not get any tandemly duplicated genes of the HSF gene family in these cotton genomes. So, the TD events brought an increase of the members of the HSF gene family in G. thurberi (D1) genome, but the HSF gene family in 12 cotton species did not experience TD event, which meant that members of the HSF gene family were discretely distributed into 12 cotton genomes.In this part, we investigated this important mechanism that occurred in different cotton genomes to get more evidence for the expansion of the HSF gene family. With the method implemented in PTGBase, we obtained 5,672, 5,118, 6,237, 6,013, 4,956, 8,070, 5,630, 4,930, 7,635, 25,877, 26,570, 26,306, and 21,726 tandemly duplicated genes in  genomes . ThroughG. hirsutum (AD1) acc. TM-1 and G. barbadense (AD2) acc. Hai7124. These treatments of different periods (hours) include control , heat , and cold  in G. hirsutum (AD1) and G. barbadense (AD2), respectively. After curation, we got 32 treatments in total. Using all available RNA-seq short-reads data of different samples of G. hirsutum (AD1) and G. barbadense (AD2), we implemented an expression analysis of total protein-coding genes in G. hirsutum (AD1) and G. barbadense (AD2) genome and further retrieved expression values of 78 and 78 HSF genes from different G. hirsutum (AD1) and G. barbadense (AD2) samples, respectively (G. hirsutum (AD1) genome, 76 HSF genes were detected to have expression in 16 samples, with 2 HSF genes (Ghi_A01G06131 and Ghi_D01G05506) having no expression in any of the samples. In G. barbadense (AD2) genome, 76 HSF genes have expression among 16 samples and two HSF genes (Gobar.A01G125400.1 and Gobar.D01G132500.1) were not detected to have expression in any of the samples. In G. hirsutum (AD1) genome, the 76 expressed HSF genes were divided into two different groups, I and II, and the I group contained 35 HSF genes, with the II group containing 41 HSF genes (G. barbadense (AD2) genome, 76 expressed HSF genes also were divided into two different groups, I and II, but the I group contained 46 HSF genes and the II group contained 30 HSF genes (G. hirsutum (AD1) and G. barbadense (AD2) indicated different expression patterns despite the implementation of identical experimental materials with identical heat and cold stresses.To investigate the expression differences of cotton HSF genes for heat stress tolerance, we analyzed the expression profiling of HSF genes in leaves under different treatments  of ectively . Out of SF genes . In G. bSF genes . These rG. hirsutum (AD1) and G. barbadense (AD2), we obtained 61 HSF orthologous gene pairs between the two genomes. In G. hirsutum (AD1) genome, 31 and 30 HSF genes from 61 orthologous gene pairs were located on G. hirsutum (AD1) A and D genomes, respectively, with one HSF gene locating on unknown pseudomolecular in G. hirsutum (AD1) D genome. In G. barbadense (AD2) genome, there were 61 HSF genes from 61 orthologous gene pairs and one HSF gene was not detected on any chromosome, with 30 HSF genes locating on A genome and 30 HSF genes locating on D genome. The no expression HSF genes, Ghi_A01G06131 versus Gobar.A01G125400.1 and Ghi_D01G05506 versus Gobar.D01G132500.1, were orthologous gene pairs between G. hirsutum (AD1) and G. barbadense (AD2) genomes. After removing the no expression HSF genes, 59 HSF orthologous gene pairs between G. hirsutum (AD1) and G. barbadense (AD2) genomes were selected to investigate the difference of expression patterns (G. herbaceum (A1), G. hirsutum (AD1), and G. barbadense (AD2), four HSF genes in G. herbaceum (A1) were detected to have four and four orthologous HSF genes in G. hirsutum (AD1) and G. barbadense (AD2), respectively. Only three of four HSF orthologous gene pairs between G. hirsutum (AD1)  and G. barbadense (AD2)  were expressed in different abiotic stress treatments and demonstrated different downregulated expression patterns regardless of control, cold, or heat stresses (According to the collinear relationship between patterns . Throughstresses .G. hirsutum (AD1) genome, we collected five genome-sequenced plant species, including the model plant A. thaliana, three cotton diploid species  and one tetraploid species [G. hirsutum (AD1)]. The allotetraploid cotton, G. hirsutum (AD1), named Upland cotton, is one of the most economically important crops which dominates over 98% of cotton production worldwide. Previous reports illustrated that G. hirsutum (AD1) was generated from hybridization between cotton A-genome progenitor and D-genome ancestor by chromosome doubling. Upland cotton D genome originated from G. raimondii (D5), and the A genome originated from the common progenitor, A0 genome, of all existing A genomes in Gossypium lineage. Furthermore, the formation of G. hirsutum (AD1) has preceded the split of G. herbaceum (A1) and G. arboreum (A2). These two cotton A genomes were shown to evolve independently from their common ancestor (G. hirsutum (AD1) genome, we got 40 and 38 HSF genes in G. hirsutum (AD1) A and D genomes, which meant that the D genome in G. hirsutum (AD1) missed two HSF genes compared with G. hirsutum (AD1) A genome. Through comparative genomic analysis between G. raimondii (D5) and G. hirsutum (AD1) D genome, we got 33 HSF orthologous gene pairs between the two genomes, which meant that G. raimondii (D5) missed five HSF genes after independent evolution. For the comparisons of G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genomes, we got 17 HSF orthologous gene pairs between G. arboreum (A2) and G. hirsutum (AD1) A genomes, containing four HSF orthologous gene pairs, which meant that G. herbaceum (A1) and G. arboreum (A2) missed more HSF genes after the split of their common ancestor, but they kept four HSF orthologous genes inherited from their common ancestor, A0 genome. These four common conserved HSF genes in G. herbaceum (A1), G. arboreum (A2), and G. hirsutum (AD1) A genome might experience stronger natural selection pressures and performed a key biological function for resisting heat stress tolerance for adapting to the changing environment. From the evolutionary history of cotton diploid and tetraploid genomes, we knew that ancestral cotton genomes experienced loss of the members of the HSF gene family, but the current cotton tetraploid genomes kept more members of the HSF gene family.To trace the evolutionary history of HSF genes in ancestor . ThroughHeat shock transcription factor gene family analysis among different cotton diploid and tetraploid genomes showed significant differences in number variation of the members of the HSF gene family. Apparently, members of the HSF gene family in cotton tetraploid genomes indicated a larger family size of the HSF gene family than those in cotton diploid genomes. Members of the HSF gene family in subgenomes of cotton tetraploid genomes showed expansion compared with those in cotton diploid genomes. These results demonstrated that HSF genes in cotton diploid genomes experienced weaker selection pressures leading to fast loss of HSF genes during the evolutionary history, but the HSF genes in subgenomes of cotton tetraploid genomes experienced stronger selection pressures after hybridization. The analysis of number variation of the members of the HSF gene family illustrated that the cotton tetraploid species may indicate stronger ability for the resistance of heat stress tolerance than diploid species.G. hirsutum (AD1) and G. barbadense (AD2) diverged after the formation of cotton allotetraploid AD genome at approximately 0.96 Mya, suggesting that these two genomes kept many orthologous gene pairs (G. hirsutum (AD1) and G. barbadense (AD2) genomes, respectively. From collinear analysis, 122 HSF genes from 61 orthologous gene pairs in G. hirsutum (AD1) and G. barbadense (AD2) were used to detect the expression pattern of HSF orthologous genes in cotton tetraploid species. After the analysis of difference expression, 61 and 61 HSF genes in G. hirsutum (AD1) and G. barbadense (AD2), respectively, indicated totally different expression pattern, which meant the functional divergence of HSF orthologous genes between the two genomes. These results provided novel insights for functional studies and molecular breeding for the cotton community.A previous study illustrated that ne pairs . Based oG. thurberi (D1), G. raimondii (D5), G. turneri (D10), G. longicalyx (F1), G. australe (G2), and G. kirkii (K) compared with those in G. herbaceum (A1) and G. arboreum (A2). As expected, cotton tetraploid genomes have more members of the HSF gene family than diploid genomes. Phylogenetic analysis revealed that 621 HSF genes in 13 cotton genomes were clustered into two different groups, and the I group contained all HSF genes of HSFA and HSFC, with the II group containing all HSF genes of HSFB according to the conserved domains and phylogeny of HSF genes. Comparing A. thaliana, G. herbaceum (A1), G. arboreum (A2), G. raimondii (D5), G. herbaceum (A1), and G. arboreum (A2), four HSF genes were inherited from the common progenitor, A0, of all existing cotton A genomes, and members of the HSF gene family indicated a significant expansion in G. arboreum (A2) and G. hirsutum (AD1) A genome compared with G. herbaceum (A1). However, HSF genes in G. hirsutum (AD1) D genome experienced a significant expansion compared with those in G. raimondii (D5). Analysis of TD events of HSF genes revealed that the HSF gene family in G. thurberi (D1) genome has experienced TD event, but not in the remaining 12 cotton genomes. Expression analysis revealed that HSF orthologous gene pairs between G. thurberi (D1) and G. barbadense (AD2) showed a totally different expression trend under identical heat and cold stresses, which is beneficial for cotton molecular breeding and directed screening. This study is the first to trace the evolutionary history and expression features of the HSF gene family in 13 cotton genomes and provides more biological evidence to study the HSF gene family in response to abiotic stress tolerance in cotton genomes, particularly expression diversity of HSF genes in different treatments of leaves among different G. hirsutum (AD1) and G. barbadense (AD2) accessions. We hope this work will provide novel insights for the study of HSF genes under environmental stress in different cotton genomes and a widespread application model for the study of HSF gene families in plants.According to the conserved domains of HSF gene family in plants, we identified 621 HSF genes among 13 genome-sequenced cotton species distributed into eight diploid and five tetraploid species. Comparative genomics analysis demonstrated that members of the HSF gene family in cotton diploid genome showed a significant expansion in The original contributions presented in the study are included in the article/YL and QC conceived and designed the experiments. YL analyzed the data and prepared the manuscript. JW and JZ performed the experiments and analyzed the data. ZG performed phylogenetic analysis and revised the manuscript draft. ZL collected genomic data, performed comparative analysis of HSF genes, and revised the manuscript draft. XA and XL performed the tandem duplication analysis of HSF genes and revised the manuscript draft. QC revised the manuscript. All authors read and approved the final manuscript.ZL was employed by company Adsen Biotechnology Corporation. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Rapid antigen tests  are cheaper and faster than nucleic acid amplification tests  for SARS-CoV-2 infection, with variable reported sensitivity. A horse racetrack in California experienced a COVID-19 outbreak among staff and used BinaxNOW to supplement RT-PCR. Utility of BinaxNOW in detecting SARS-CoV-2 infection in a workplace outbreak was assessed.t) < 30.Between November 25\u2013December 22, 2020, anterior nasal swabs were collected from racetrack staff for six rounds of paired BinaxNOW and RT-PCR tests. BinaxNOW tests were interpreted according to manufacturer instructions. RT-PCR was performed at the state public health lab using the ThermoFisher TaqPath COVID-19 Combo Kit. Staff with positive results on either test were isolated and removed from subsequent testing. Viral cultures were attempted on specimens with cycle threshold  (17.8 vs. 28.5) (p < 0.001). In dual positive pairs, median time from specimen collected to RT-PCR result reported was 4 days (range 1-6), compared to the 15-minute BinaxNOW reporting time. Of 100 Ct < 30 specimens, 51 resulted in positive virus isolation, 45 (88.2%) of which were BinaxNOW-positive.Overall, 769 paired results from 342 staff were analyzed. Most were of Hispanic ethnicity (62.0%) and ages ranged from 18 to 92 years (median 52). BinaxNOW performance compared to RT-PCR (95% CI) was as follows: positive percent agreement (PPA) 43.3% (34.6%\u201352.4%); negative percent agreement (NPA) 100% (99.4%\u2013100%); positive predictive value (PPV) 100% (93.5%\u2013100%); negative predictive value 89.9% (87.5%\u201392.0%). Among 127 RT-PCR-positive specimens, those with paired BinaxNOW-positive results (n = 55) had a lower mean Ct value and positive viral cultures, which could suggest that among RT-PCR-positive specimens, those that are BinaxNOW-negative may be less likely to contain infectious virus than those that are BinaxNOW-positive.High NPA and PPV support immediate isolation of BinaxNOW-positive individuals, while low PPA supports confirmatory testing following BinaxNOW-negative results. BinaxNOW performed better in paired specimens with lower CDavid Seftel, M.D., M.D., M.B.A., Enable Biosciences, Inc"}
+{"text": "Authors would like to correct the errors in their publication.The original article has been corrected.Incorrect Supplementary Information (ESM2) deleted now.The line \u201cA color version of this figure is available online\u201d to be deleted from all figure captions since the color figures in print version is not charged now.Missing references for the text citations were updated hereAlongi, 2002Alongi DM (2002) Present State and Future of the World\u2019s Mangrove Forests. Environ Conserv 29:331\u2013349. 10.1017/S0376892902000231Alongi and Dixon, 2000Alongi DM, Dixon P (2000) Mangrove primary production and above-and below-ground biomass in Sawi Bay, southern Thailand. Phuket Mar Biol Cent Spec Publ 22:31-38.Arfi et al. 2012Avicennia marina and Rhizophora stylosa. FEMS Microbiol Ecol 79:433\u2013444. 10.1111/j.1574-6941.2011.01236.xArfi Y, Bu\u00e9e M, Marchand C, et al (2012) Multiple markers pyrosequencing reveals highly diverse and host-specific fungal communities on the mangrove trees Krishna and Mohan, 2017Krishna MP, Mohan M (2017) Litter decomposition in forest ecosystems: a review. Energy, Ecol Environ 2:236\u2013249. https://doi.org/10.1007/s40974-017-0064-9Murdiyarso et al. 2015Murdiyarso D, Purbopuspito J, Kauffman JB, et al (2015) The potential of Indonesian mangrove forests for global climate change mitigation. Nat Clim Chang 5:1089\u20131092. 10.1038/nclimate2734Quadros et al. 2019Quadros AF, Nordhaus I, Reuter H, Zimmer M (2019) Modelling of mangrove annual leaf litterfall with emphasis on the role of vegetation structure. Estuar Coast Shelf Sci 218:292\u2013299. 10.1016/j.ecss.2018.12.012"}
+{"text": "We observed altered gene expression in several DNA repair pathways including homologous recombination repair, non-homologous end-joining and mismatch repair, with hypoxia primarily resulting in downregulation of gene expression. The extent of gene expression changes was dependent on hypoxic severity. Some, but not all, of these downregulations were directly under the control of HIF activity. For example, the downregulation of LIG4, a key component of non-homologous end-joining, was reversed upon inhibition of the hypoxia-inducible factor (HIF). In contrast, the downregulation of the mismatch repair gene, PMS2, was not affected by HIF inhibition. This suggests that numerous molecular mechanisms lead to hypoxia-induced reprogramming of the transcriptional landscape of DNA repair. Whilst the global impact of hypoxia on DNA repair gene expression is likely to lead to genomic instability, tumorigenesis and reduced sensitivity to anti-cancer treatment, treatment re-sensitising might require additional approaches to a simple HIF inhibition.Glioblastoma, a grade IV astrocytoma, has a poor survival rate in part due to ineffective treatment options available. These tumours are heterogeneous with areas of low oxygen levels, termed hypoxic regions. Many intra-cellular signalling pathways, including DNA repair, can be altered by hypoxia. Since DNA damage induction and subsequent activation of DNA repair mechanisms is the cornerstone of glioblastoma treatment, alterations to DNA repair mechanisms could have a direct influence on treatment success. Our aim was to elucidate the impact of chronic hypoxia on DNA repair gene expression in a range of glioblastoma cell lines. We adopted a NanoString transcriptomic approach to examine the expression of 180 DNA repair-related genes in four classical glioblastoma cell lines  exposed to 5 days of normoxia (21% O Surviva options . Alterna options . Despite options .2  components leading to reduced HRR capacity . Also, cWe adopted a NanoString transcriptomic approach to assess the impact of chronic hypoxia on DNA repair genes in glioblastoma cell lines. We found that hypoxia specifically affects mismatch repair (MMR), non-homologous end-joining (NHEJ) and homologous recombination repair (HRR). Additionally, we provide evidence that the hypoxia inducible factor (HIF) play a role in downregulation of some, but not all, key DNA repair genes. Downregulation of DNA repair genes by hypoxia will have significant clinical impact for cancer management and should be considered when designing new treatment methods and protocols.Cell culture reagents were from Gibco Life Technologies and Foetal Calf Serum from Harlam Seralab (UK). Acriflavin was purchased from Sigma Aldrich. \u03b2-Actin (Ab8226), PMS2 (Ab110638) and anti-mouse HRP (Ab6808) antibodies were from abcam. HIF-1\u03b1 (20960-1-AP) and HIF-2\u03b1 (A700-003) were from Bethyl. Anti-rabbit HRP (7074S) antibody was from Cell Signalling.2. For hypoxic experiments, cells were incubated in a Don Whitley H35 Hypoxystation for 1% O2 or a New Brunswick Galaxy 48R incubator for 0.1% O2. Cells were routinely tested for mycoplasma infection.U87-MG  and T98G  were purchased from ATCC. U251-MG  were purchased from CLS. D566-MG cells  were a kind gift from Prof. DD Bigner . U87-MG and T98G were cultured in modified Eagle\u2019s Medium (EMEM) with 10% FCS, 1% Sodium pyruvate. D566-MG and U251-MG were cultured using EMEM, 10% FCS, 1% sodium pyruvate and 1% non-essential amino acids. Cells were maintained at 37 \u00b0C in 5% CO2. RNA was extracted using High Pure RNA Extraction kit . A total of 100 ng of total RNA was used in the NanoString assay. The expression of 180 DNA repair genes, plus 12 housekeeping genes, were assessed using the NanoString nCounter Vantage\u2122 RNA Panel for DNA Damage and Repair (LBL-10250-03) following the manufacturer\u2019s protocol. Analysis of the NanoString data was conducted using nSolver\u2122 Analysis Software (3.0) with nCounter\u00ae Advanced Analysis plug-in (2.0.115). Data are available at https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE139250 and inCells were cultured in 6 cm dishes for 5 days at 21%, 1% or 0.1% O\u00ae VILO  following the manufacturer\u2019s guidelines. RT-PCR experiments were performed and analysed as described in LIG4 Forward: TCCCGTTTTTGACTCCCTGG Reverse: GGCAAGCTCCGTTACCTCTG, ABL Forward: TGGGGCATGTCCTTTCCATC Reverse: GATGTCGGCAGTGACAGTGA, ERCC4 Forward: CTCCCTCGCCGTGTAACAAA Reverse: ACACCAAGATGCCAGTAATTAAATC, FEN1 Forward: GTTCCTGATTGCTGTTCGCC Reverse: ATGCGAATGGTGCGGTAGAA, MSH5 Forward: GTTTGCGAAGGTGTTGCGAA Reverse: GTCTGAGACCTCCTTGCCAC, PARP1 Forward: GCCCTAAAGGCTCAGAACGA Reverse: CTACTCGGTCCAAGATCGCC, UBE2T Forward: ATGTTAGCCACAGAGCCACC Reverse: ACCTAATATTTGAGCTCGCAGGT, WRN Forward: TCACGCTCATTGCTGTGGAT Reverse: CAACGATTGGAACCATTGGCA, PMS2 Forward: AGCACTGCGGTAAAGGAGTT Reverse: CAACCTGAGTTAGGTCGGCA, CYCLOA Forward: GCTTTGGGTCCAGGAATGG Reverse: GTTGTCCACAGTCAGCAATGGT. Cyclophillin A was used as a housekeeping gene.RNA was extracted using High Pure RNA Extraction kit . Reverse transcription was conducted using SuperScriptLIG4 (NM_001352604.1) and PMS2 (NM_000535.7) was obtained through the UCSC Genome Browser using human genome assembly h38 , and GLUT1 or VEGFA levels assessed by RT-PCR to confirm hypoxia had indeed resulted in hypoxia-induced gene expression changes , D566-MG (0.1% = 2.6-fold) and U87-MG (0.1% = 1.6-fold)  at 0.1% O2  and MutL\u03b1  actively translocates along the DNA in search of discontinuity . EXO1 is 0.1% O2 , yet litsistance , was als.6-fold)  and 3B, .6-fold) . However studies . Other c 0.1% O2 . MSH5 is 0.1% O2 , therefo 0.1% O2 , potentiXRCC6, Ku80-XRCC5, DNA-PK-PRKDC) remain largely unaffected by hypoxia. However, in all cell lines tested, LIG4 (DNA Ligase IV) was downregulated at least 1.5-fold in hypoxia (1% and 0.1% O2.), although, this was only statistically significant in U87-MG  complex recognises the DSB . The endn U87-MG  and 3D.  hypoxia  and 3D. in XRCC4 .PMS2 and LIG4, essential components of their respective repair pathways, are involved in chemoresistance   , resulti2 (PHD2) . To testactivity . Treatmeget gene . In bothnificant . In contreatment , suggestPMS2 (MMR) and LIG4 (NHEJ), essential components of their respective pathways, were downregulated across multiple cell lines after chronic and acute hypoxic exposure, providing potential targets for re-sensitisation of hypoxic tumour cells to DNA damaging therapies.Tumour hypoxia plays an essential role in tumour progression, metastasis and drug resistance, leading to poor patient outcome, particularly for GBM patients. Hypoxia has been shown to impact numerous signalling pathways including the cell cycle, metabolism and apoptosis, yet additionally, hypoxia can have a significant impact on DNA repair mechanisms. We have shown here that several key DNA repair pathways were downregulated by hypoxia, including MMR and NHEJ, both essential pathways for the repair of modified/mismatch bases and double strand breaks respectively. Further analysis determined that PMS2 , and siRNA of LIG4 enhanced cell lethality of both chemotherapeutics U87-MG, (B) U251-MG and (C) D566-MG was utilised in RT-PCR experiments to assess the expression levels of selected genes. Data are expressed as logClick here for additional data file.10.7717/peerj.11275/supp-3Supplemental Information 32 and treated with 0\u201310 \u00b5M Acriflavin for 24 h. Control cells remained in 21% O2. RT-PCR was performed to assess GLUT1 expression levels. Data represent the fold change in GLUT1 expression relative to 21% O2. Data are from a single experiment. (B) HIF inhibition does not restore PMS2 mRNA levels in hypoxia. D566\u2011MG and U87-MG cells were incubated in 21% and 1% O2 for 24 h with and without 5 \u00b5M Acriflavin, the plots show the log2 fold change in mRNA levels with respect to levels for the 21% O2 samples. Data are the mean of at least two independent experiment.(A) D566-MG cells were incubated in 0.1% OClick here for additional data file.10.7717/peerj.11275/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.11275/supp-5Supplemental Information 5The data for each figures are on separate tabs.Click here for additional data file.10.7717/peerj.11275/supp-6Supplemental Information 62 for 5 days before examination of gene expression using the NanoString nCounter Vantage\u2122 RNA Panel for DNA Damage and Repair. Differential gene expression analysis was performed using nSolver\u2122.U87-MG, D566-MG, U251-MG and T98G were exposed to 21%, 1% or 0.1% OClick here for additional data file."}
+{"text": "P. aeruginosa is a common infection among hospitalized patients, with increased levels of morbidity and mortality. This pathogen exhibits multiple resistance mechanisms to antibiotics. We analyzed the molecular epidemiology and activity of the main therapeutic options against P. aeruginosa isolated from RTI in Latin America (LATAM).Respiratory Tract Infection (RTI) caused by Isolates were collected from 36 sites in 10 countries during 2017-2019. Non-duplicate samples were consecutively collected. MICs were determined by broth microdilution and interpreted by CLSI criteria. A subset of imipenem non-susceptible isolates was selected for characterization of carbapenemase encoding genes via multiplex PCR and DNA sequencing. \u03b2-lactamase genes encoding ESBLs, carbapenemases, and plasmid-mediated AmpCs were investigated.P. aeruginosa were collected from RTI. Overall C/T [87.8% susceptible (S)] was the most active antimicrobial tested against P. aeruginosa isolates followed by amikacin (85.8% S) and imipenem/relebactam . Other antimicrobials had less than 80% susceptibility. C/T remained the most active agent including activity against imipenem and piperacillin/tazobactam non-susceptible isolates . 583 imipenem non-susceptible P. aeruginosa were selected for molecular analysis (Table 1). Thirty  isolates were confirmed to be producers of serine-carbapenemases , while 83 (14.2%) were MBL producers. KPC-2 was found in Colombia (9), Chile (6), Puerto Rico (4), Guatemala (3), and Brazil (2). GES-5 was identified in Mexico (3), Argentina (2) and Brazil (1). VIM-2 was the most common MBL encoding gene identified. IMP variants were observed in Brazil , Ecuador (IMP-13), Mexico (IMP-18), Panama (IMP-18) and Puerto Rico (IMP-18). SPM-1 was only encountered in Brazil. The production of ESBLs was low in most LATAM countries, except for Guatemala (80%) . A total of 2,044 Figure 1. Activities of selected antimicrobial agents against 2,044 P. aeruginosa isolated from respiratory tract infections in Latin America (2017 to 2019).TABLE 1. Molecular analysis of imipenem non-susceptible P. aeruginosa isolates in Latin America (LATAM) from Respiratory Tract Infection (N=583).Figure 2. Carbapenemases identified in 583 imipenem non-susceptible P. aeruginosa isolated from patients with respiratory tract Infections in Latin America (LATAM).P. aeruginosa infections. The prevalence of carbapenemases-encoding genes varied geographically in LATAM.CT, amikacin and IMI/REL showed good activity against RTI isolates and could represent effective treatment options for Leandro Cardinal, PharmD, PhD, MSD (Employee) Cicera P. Marcelino, n/a, MSD (Employee) Aline Okuma, n/a, MSD (Employee)MSD Brazil (Employee) Gustavo Mizuno, PharmD, Merck Sharp Dohme (Employee) Felipe Tuon, PhD, Merck Sharp Dohme Brazil (Scientific Research Study Investigator) Ana C. Gales, MD, MSD Pfizer  Marina Della Negra, Medical Doctor, MSD Brazil (Employee) Thales Polis, Medical Doctor, MSD Brazil (Employee)"}
+{"text": "OBJECTIVES/GOALS: We compared the validity of an International Classification of Diseases, Clinical Modification (ICD) algorithm for identifying high-grade cervical intraepithelial neoplasia and adenocarcinoma in situ (together referred to as CIN2+) from ICD 9th revision (ICD-9) and 10th revision (ICD-10) codes. METHODS/STUDY POPULATION: Using Tennessee Medicaid data, we identified cervical diagnostic procedures in 2008-2017 among females aged 18-39 years in Davidson County, TN. Gold-standard cases were pathology-confirmed CIN2+ diagnoses validated by HPV-IMPACT, a population-based surveillance project in catchment areas of five US states. Procedures in the ICD transition year (2015) were excluded to account for implementation lag. We pre-grouped diagnosis and procedure codes by theme. We performed feature selection using least absolute shrinkage and selection operator (LASSO) logistic regression with 10-fold cross validation and validated models by ICD-9 era  and ICD-10 era . RESULTS/ANTICIPATED RESULTS: Of 7864 cervical diagnostic procedures, 880 (11%) were true CIN2+ cases. LASSO logistic regression selected the strongest features of case status: Having codes for a CIN2+ tissue diagnosis, non-specific CIN tissue diagnosis, high-grade squamous intraepithelial lesion, receiving a cervical treatment procedure, and receiving a cervical/vaginal biopsy. Features of non-case status were codes for a CIN1 tissue diagnosis, Pap test, and HPV DNA test. The ICD-9 vs ICD-10 algorithms predicted case status with 68% vs 63% sensitivity, 95% vs 94% specificity, 63% vs 64% positive predictive value, 96% vs 94% negative predictive value, 92% vs 89% accuracy, and C-indices of 0.95 vs 0.92, respectively. DISCUSSION/SIGNIFICANCE OF IMPACT: Overall, the algorithm\u2019s validity for identifying CIN2+ case status was similar between coding versions. ICD-9 had slightly better discriminative ability. Results support a prior study concluding that ICD-10 implementation has not substantially improved the quality of administrative data from ICD-9."}
+{"text": "The article presents the results of studying the biodiversity and biotechnological potential of halophilicmicroorganisms from the thermal highly mineralized Berikey Lake, the salty Lake Tarumovskoye and saline soils ofthe Peri-Caspian Lowland (Republic of Daghestan). Denitrifying halophilic bacteria of the genus Halomonas andVirgibacillus were identified using microbiological methods and 16S rRNA gene analysis. A new species Halomonas sp. G2 (MW386470) with a similarity of the nucleotide sequences of the 16S rRNA genes is 95 %. Strain G2 isan extreme halophile capable of growing in the range of 5\u201325 % NaCl (optimum 25 %) and forming a carotenoidpigment. Mesophil, 30\u201337 \u00b0\u0421 (optimum 30 \u00b0\u0421); neutrophil, pH 6\u20138 (optimum 7.2\u20137.4). Strain G2 chemolithotroph;reduces nitrate or nitrite as electron donors; catalase-, amylase-, protease- and \u03b2-galactosidase-positive; lipase-,oxidase- and urease-negative. Not able to hydrolyze inositol, indole; produces lysine, gelatin, ectoine; uses citrateand sodium malate as a source of carbon and energy; does not produce ornitin, H2S or acid from d-mannose, sucrose, glycerol, cellobiose, except for lactose and d-glucose. Susceptible to trimethoprim, ciprofloxacin, ofloxacin,kanamycin, vancomycin, rifampicin, cefuroxime, ampicillin, ceftazidime, fosfomycin, clarithromycin, cefepime, cefaclor. The G+C content in DNA is 67.3 %. A distinctive characteristic of the isolate was the production of industrially significant hydrolytic enzymes such as amylase, protease, \u03b2-galactosidase, and oxidoreductase  at aNaCl concentration of 25 % in the medium. Habitat: saline soils on the territory of the Tersko-Kumskaya lowland. The rest of the halophilic isolates of H. ventosae G1 (MW386469), H. elongata G3(MW386471), V. salinarius B2 (MW386472), and V. salinarius B3 (MW386473) had a high degree of similarity (100 %)with the type strains of H. elongata DSM 2581\u0422and V. salarius DSM 18441\u0422; the content of G+C in DNA was 65.8,66.5, 42.8 and 37.3 %, respectively. The strains had a high biotechnological potential at NaCl concentrations of 5 and25 % in the medium. The data obtained expanded the understanding of the diversity and ecological significanceof denitrifying bacteria in the functioning of arid ecosystems and make it possible to identify strains producingenzymes of industrial importance. The interest in extremophilic microorganisms is relativelyhigh due to their biological uniqueness. Agreat contributionto the study of natural microbial communities was made bythe school of Russian scientists . Archaeaand highly-specialized bacteria of the genera Alcaligenes,Bacillus, Halobacillus, Virgibacillus, Micrococcus, andPseudomonas  occupy a dominant place in ecologicalniches with a high salt content  and anthropogenic ecosystems with an increasedlevel of mineralizationThe largest ecosystems on the planet are saline and hypersaline environments . Daghestan isa unique natural province of Russia that has a variety ofnatural landscapes due to the influence of tectonic processes,the erosive activity of flowing waters, transgressive andregressive dynamics of the Caspian Sea, and arid climate.There are a number of works on the study of microbial communities of various ecological niches of the region: lithotrophic sulfur-oxidizing representatives of sulfide sources,hydrocarbon-oxidizing bacteria of the geothermal sourceof the Kizlyar field .The highly mineralized lakes of the Tersko-Kumskayalowland with high salinity form conditions for the existence of halophilic bacteria. Microorganisms from extremehabitats are the producers of valuable industrially importantenzymes, antibiotics; they can participate in soil biodegradation, and are highly resistant to contamination by foreignmicroflora .The study considers the spatial distribution of halophilicmicrobial communities of halophyte plants, saline soils, and highly mineralized lakes in arid regions of the Peri-CaspianLowland . Chemoorganoheterotrophic bacteria of the genera Virgibacillus, Bacillus,Halomonas and Salimicrobium from the phylum Firmicutesand Proteobacteria have proved to be the main componentsof the microbial flora of the Tersko-Kumskaya and TerskoSulakskaya provinces. A major correlation was revealedbetween isolated microbial communities and concentrationsof chemicals Na, K, Ca, Mg, Cl, Cu, Sr, SO4, Cl, and HCO3,as one of the main regulators of microbiological activity insoils and lakes.The aim of the paper is a molecular taxonomic studyof isolated halophilic bacteria and their biotechnologicalpotential.The objects of research are natural microbial communitiesof saline reservoirs and soils in the territory of the PeriCaspian Lowland of the Republic of Daghestan  (Table 1). Samples were taken in July\u2013September 2014.Cultivation. For the cultivation of halophilic bacteria,a modified medium of the following composition (g/l) wasused: bacto yeast extract \u2013 10.00, Na3C6H5O7 \u22195.5H2O \u20133.0, NaCl \u2013 50, 100, 250, KCl \u2013 2.0, MgSO4 \u2219 7H2O \u201320.0, glycerin \u2013 4.0 . Bacto-peptone  \u2013 5 g/lwas used as a substrate; the pH of the medium was adjustedto 7.2\u20137.4 by 1N HCl or 4M KOH (Russia) using a HannaInstrumentals pH 211 pH meter (Germany). The cultureswere incubated in a Binder-115 microbiological incubator(USA) at an operating temperature of (30\u201337)\u00b11 \u00b0C for3\u201320 days.Morphology of bacterial cells  was studied using a light microscope CX21 FS1  and thePowerShot A640 digital camera  at a working magnification of \u00d7600.Ecological and physiological characteristics of growth. Effect of NaCl concentration in an amount of 2 % ofthe medium volume for cell growth in liquid and solid mediawas determined at 30\u201337 \u00b0C in the Binder-115 incubator(USA). The growth was monitored at 24-hour intervals for7 days by measuring turbidity using a Genesys-20 spectrophotometer . The effect of temperature (30 and 37 \u00b0C) on the growth rate was determinedby cultivation under the same conditions.The use of growth substrates  was studied using standard methods.The use of electron acceptors. The ability to use nitrateas an electron acceptor was determined using the BD BBLTaxo Differentiation Discs Nitrate , in compliance with the manufacturer\u2019sinstructions. The discs were impregnated with a solutioncontaining 40 % potassium nitrate and 0.1 % sodium molybdate. The reduction of nitrate to nitrite was detected by theaddition of sulfanilic acid and N,N-dimethyl-\u03b1-naphthylamine, which reacts with nitrite to form a red-colored substance-n-sulfobenzenazo-\u03b1-naphthylamine (positive result).In the absence of a change in color after the addition ofreagents (negative result), zinc dust was added to determinethe presence of unrecoverable nitrate or products other thannitriteDetermination of enzymatic activity. Hydrolase-producing bacteria were screened for amylase, proteinase,\u03b2-galactosidase, lactase, lipase, urease, and oxidoreductases on dishes of starch, tributyrin, andgelatin agar, depending on the concentrations of NaCl.Amylase activity \u2013 on the elective medium . The isolateswere incubated at 45 \u00b0C for 24\u201336 hours, tested with Lugolsolution . Potential amylase producers were selected based onthe ratio of the clearance zone diameter to the colony diameter. Protease was determined on media with agar and 10 %skimmed milk; \u03b2-galactosidase (lactase) \u2013 using indicatordisks impregnated with a special reagent-ortho-nitrophenyl\u03b2-d-galactopyranoside ; urease \u2013 using CLOtest ; lipase \u2013 tested on dishes with1 % tributyrin. Isolates showing clear zones of tributyrinhydrolysis were identified as lipase-producing bacteria.Determination of oxidoreductases: catalase \u2013 using 3 %H2O2 as a substrate in the medium for 24\u201348 hours, oxidase \u2013 by Kovac\u2019s method . All screening testsfor enzymatic activity were performed in three repetitions.The bacteria were incubated at 37 \u00b0C for seven days.Antibiotic resistance  was determined by the intensity of bacterialgrowth on the basic agarized medium \u201cB\u201d in Petri dishesusing standard disks \u201cIndicator paper systems for identifyingmicroorganisms\u201d of NPO Microgen of Natsimbio Holding(Russia) with 10\u201330 \u03bcg of antimicrobial agent .G+C content and phylogenetic analysis. Genomic DNAwas isolated according to Marmur (1961) and Thomas  methods. The DNA nucleotide compositionwas determined by thermal denaturation (0.5 \u00b0C\u2219min\u20131)using a Cary-100 Bio UV-VIS spectrophotometer . The GC content in the composition of DNA \u2013according to the method . Escherichiacoli K-12 DNA (51.7 %) was used as the standard.For phylogenetic analysis, the DNA was isolated fromsamples using the modified Birnboim\u2013Doly alkaline DNA method  and the Wizard technologyof Promega (USA) . The concentration of the resulting DNA sample when using this methodwas 30\u201350 \u03bcg /ml. RNA in the resulting preparation is present in trace amounts .For polymerase chain reaction (PCR) and further sequencing of PCR fragments of the 16S rRNA gene for each ofthe studied samples, universal primer systems were usedto detect both eubacteria (11f-1492r)  and archaea (8fa-A915R) . The volumeof the amplification mixture was 50 \u03bcl with the followingcomposition: 1\u00d7 buffer of BioTaq DNA polymerase (17 mM(NH4)2SO4, 67 mM Tris-HCl, pH 8.8, 2 mM MgCl2);12.5 nmol of each dNTP, 50 ng of the DNA matrix; 5 pmolof the corresponding primers and 3 units of BioTaq DNApolymerase . The temperature-timeprofile of the PCR was as follows: the first cycle \u2013 94 \u00b0C\u00d79 min, 55 \u00b0C\u00d71 min, 72 \u00b0C\u00d72 min; the next 30 cycles \u201394 \u00b0C\u00d71 min, 55 \u00b0C\u00d71 min, 72 \u00b0C\u00d72 min; the final cycle \u201372 \u00b0C\u00d77 min. The PCR products were analyzed by electrophoresis in 2 % agarose gel at an electric field strengthof 6 V/cm. Isolation and purification of PCR products werecarried out from low-melting agarose using a set of reagentsby WizardPCRPreps , according to themanufacturer\u2019s recommendations.Sequencing of PCR products was carried out at the Centerof \u201cBioengineering\u201d of the Russian Academy of Sciences,Moscow, by the method of  using a setof BigDyeTerminatorv.3.1 reagents on the ABIPRIZM 3730genetic analyzer . Standardprimers were used for sequencing .Analysis of 16S rRNA sequences. The primary analysisof the similarity of the nucleotide sequences of the 16S rRNAgenes of the studied strains was performed using the BLASTprogram on the following web-site: https://blast.ncbi.nlm.nih.gov .The 16S rRNA gene sequences of all studied strains aredeposited in GenBank: G1 \u2013 MW386469, G2 \u2013 MW386470,G3 \u2013 MW386471, B2 \u2013 MW386472, B3 \u2013 MW386473.Strains of halophilic bacteria G1, G2, G3, B2 and B3,isolated from salt lakes and salt marshes of the TerskoKumskaya and Tersko-Sulakskaya lowlands, grew at atemperature of 30\u201337 \u00b0C and pH 6.4\u20137.4. The culturesshowed steady growth in the agarized elective mediumin the presence of 5\u201325 % NaCl with an optimum of 5,10, 25 %, which indicated that they belonged to moderateand extreme halophiles following the known classification.The analysis of the 16S rRNA gene sequence allowed usto determine their phylogenetic position. The 16S rRNAgene sequences of the new halophilic strains were analyzedand compared with the 16S rRNA sequences of the validlydescribed bacterial species. The analysis has shown that thenew isolates belong to two genera of bacteria containing halophilic microorganisms Halomonas and Virgibacillus. Table 2 and Figure 1 demonstrate that theG2 strain represents a new species in the genus Halomonas.The H. ventosae G1 (MW386469) and H. elongata G3(MW386471) strains seem to belong to the species H. ventosae and H. elongata, respectively, while the V. salinariusB2 (MW386472) and V.salinarius B3 (MW386473) strainsbelong to the group of species related to V. salinarius.Characteristics of the Halomonas sp. G2 strainThe main object of further research is the strain Halomonas sp. G2. The GC content in the G2 strain DNA was67.3 %.Morphology of cells and colonies. Rod-shaped, gramnegative, mobile bacillus of the size of 0.8\u20131.0\u00d71.5\u20133.0 \u00b5m.The cells were observed singly, in pairs, or in short chains. Cell mobility was ensured by one or two lateralflagella located on one side of the cell, forming endospores.On an elective solid medium, the strain formed an activegrowth of colonies of a round shape with a wavy edge, yellow and pale-yellow color with a gloss. With an increase inthe concentration of NaCl in the culture medium, a brightcarotenoid pigment appeared in the beige-colored colonies.On meat-peptone agar (MPA) \u2013 small colonies located closeto each other in a chain, rounded shape with a wavy edge,turning into a solid growth; smooth, shiny, light beige witha pinkish tinge. In all variants, a smearing consistency wasobserved .Physiology of strain growth . When determining the optimal growth parameters, theG2 strain was assigned to mesophiles  and moderate alkalophiles . As a representative of the genus Halomonas, it cangrow in a wide range of NaCl concentrations from 10 to25 % with an optimum of 25 %; an extreme halophileThe substrates and electron acceptors used. Oxygenratio. The G2 strain is capable of denitrification by usingnitrates as an electron acceptor, reducing them to nitrites.The differentiating characteristics of the G2 strain arepresented in Table 3. The strain had a positive reaction tolysine, gelatin, ectoine, lactose, and d-glucose; utilized citrate and Na malonate. Tests for \u03b2-galactosidase, amylase,protease, and catalase are positive; for oxidase, lipase, andurease \u2013 negative. Growth does not occur under anaerobicconditions.Antibiotic sensitivity. The G2 culture differed in sensitivity to trimethoprim of the sulfanilamide group; fluoroquinolones of the 1st and 2nd generations ; vancomycin of the macrolide group;rifamycins of the rifampin group; cefuroxime and ampicillin of the penicillin group; antibiotics of the 3rd generationcephalosporins of the macrolides ; antibiotics of the 4th generation cephalosporins .Characteristics of isolates G1, G3, B2, B3Rod-shaped, mobile cells of strains G1, G3 have the following sizes: 0.6\u20130.8 \u00d7 1.6\u20131.9 \u00b5m (G1), 0.7\u20131.0 \u00d7 1.5\u20132.5 \u00b5m (G3) . Single cells and chains ofthem were observed. Mobility was provided by flagellalocated on one side of the cell. The cells of the B2 andB3 strains were mobile, in the form of rods, with sizes:0.5\u20130.7\u00d71.0\u20132.5 \u00b5m (B2) and 0.2\u20130.7\u00d71.0\u20135.0 \u00b5m (B3);formed endospores. The biomass of isolated strains on theMPA medium is represented by a chain of colonies locatedone after another, differing in shape, color, size, pigment andmorphology . On an elective agar mediumwith 5\u201325 % NaCl  and 5\u201310 % NaCl , thecultures formed colonies with lipochromic pigment.The results of phylogenetic analysis of 16S rRNA genesequences indicated that the closest type strains (100 %) forG1 and G3 are the type strain of H. elongata DSM 2581T,and for B2 and B3 \u2013 V. salarius DSM 18441T. On thedendrogram, the cultures formed a common cluster withthe typical strains, making it possible to classify isolated cultures as these species. Related cultures for G1 and G3 areH. ventosae GQ903443, H. elongata NR 074782; for B2 andB3 \u2013 V. salarius MK785132, B. brevis JF802177, V. olivaeNR 043572, H. arcis MK063873, V. salarius NR 041270,which combined the typical features of moderate and extreme halophiles.The distinctive characteristics differentiating the culturesof H. ventosae G1 and H. elongata G3 were: optimumgrowth at 5\u201325 % NaCl vs. 32 %; pH 7.2\u20137.4 vs. 7\u20139; noutilization of sucrose, glycerol, d-mannose, cellobiose,lactose, and production of urease, oxidase, and protease(except G3) at a concentration of 5\u201325 % NaCl in themedium (see Table 3). At the same time, such traits forV.salarius B2 and B3 strains in comparison with the typicalV. salarius DSM 18441T were: no need for d-mannose, theability to produce amylase, protease, and \u03b2-galactosidaseenzymes at a concentration of 5\u201310 % NaCl in the medium(Table 4).Characteristics of genera Halomonas and VirgibacillusThus, based on phenotypic and genetic studies, we providea brief description of the strain of the new species Halomonas sp. G2 and halophilic strains of H. ventosa G1, H. elongata G3, V. salinarius B2, V. salinarius B3.Currently, the genus Halomonas includes 91 species,where H. elongata acts as the type species . The Halomonadaceaefamily was first described in 1988 by combining moderatelyhalophilic and marine bacteria of the genera Deleya andHalomonas . Over the past threedecades, many species have been assigned to the genusHalomonas, the domain Bacteria, the phylum Proteobacteria, the class Gammaproteobacteria, the order Oceanospirillales, and the family Halomonadaceae; however, at the timeof writing, 7 species have been reclassified. Representativesof the genus \u2013 gram-negative, facultative anaerobes, aerobes,prototrophs, mesophiles, denitrifying, produce exopolysaccharides; they mainly utilize oxygen, nitrate or nitrite as anelectron acceptor; under conditions of saline stress, theysynthesize ectoine, which protects cells from adverse environmental influences .The genus Virgibacillus was created as a result of thereclassification of Bacillus pantothenticus, Virgibacilluspantothenticus and, subsequently, established as a speciesof Virgibacillus . At the moment, the genus consists of 27 species,representatives of which are gram-positive, obligate aerobesor facultative anaerobes, moderate halophiles, chemotaxonomic; the main fatty acid is C15:0 .Description of the strain of the new species Halomonas sp. G2. The G2 strain cells are encapsulated, motile,aerobic, gram-negative bacillus 0.8\u20131.0 \u00d7 1.5\u20133.0 \u00b5m;observed singly or in a chain of 2 to 4 interlocked cells.The G2 strain is an extreme halophile, capable of growingin the range of 10\u201325 % NaCl (25 % optimum) and forming a carotenoid pigment. On an elective solid mediumwith 25 % NaCl, it forms colonies of a round shape with awavy edge, beige color with a gloss; forms areas of brightcarotenoid pigment. The strain grows on meat-peptonebroth. The species is mesophile, the temperature range is30\u201337 \u00b0C (30 \u00b0C optimum). Neutrophil, pH 6\u20138 (7.2\u20137.4optimum). Denitrifying strain; chemolithotrophic; reducesnitrate or nitrite as electron donors; catalase-, amylase-,protease-, and \u03b2-galactosidase-positive; lipase-, oxidase-,and urease-negative. Unable to hydrolyze inositol, indole;produces lysine, gelatin, ectoine; uses sodium citrate andmalate as a source of carbon and energy; does not produceornithine; H2S and acid from d-mannose, sucrose, glycerol,cellobiose, except lactose and d-glucose. Susceptible to trimethoprim, ciprofloxacin, ofloxacin, kanamycin, vancomycin, rifampicin, cefuroxime, ampicillin, ceftazidime, fosfomycin, clarithromycin, cefepime, cefaclor. The GC contentin DNA is 67.3 %.Based on its physiological, biochemical, and phylogeneticproperties, the G2 strain represents a new species, calledHalomonassp. G2. Adistinctive characteristic of the isolateis the production of hydrolytic enzymes protease, amylase,\u03b2-galactosidase, and oxy-reductase-catalase at a concentration of 25 % NaCl in the medium.Habitat: soil  on the territory of theTersko-Kumskaya lowland .Tersko-Kumskaya lowland .Description of the strains Halomonas ventosae G1(MW386469), and Halomonas elongata G3 (MW386471).Halomonas G1 and G3 strains are aerobes, gram-negative,denitrifying; mesophiles, prototrophs, chemolithotrophs,and extreme halophiles (5 to 25 % NaCl). Unable to hydrolyze inositol; produce lysine, ornithine, gelatin, ectoine;reduce nitrate or nitrite as electron donors; utilize citrate(excluding G3) and sodium malonate as a source of carbonand energy; do not produce H2S and acid from d-mannose,sucrose, glycerol, cellobiose, except for d-glucose. TheGC content in the DNA for G1 and G3 are 65.8 and 66.5 %,respectively. On the basis of phenotypic and genotypic characteristics, isolated bacteria are classified as H. ventosae G1(MW386469), and H. elongata G3 (MW386471).Habitat: saline soils  and Lake Tarumovskoe on the territoryof the Tersko-Kumskaya lowland . The type strain of H. elongata DSM 2581\u0422 wasisolated from equipment for extraction of salt from brine.Description of V. salinarius B2 (MW386472) andB3 (MW386473) strains. The strains are gram-positive;mesophiles, neutrophils, chemolithotrophs, moderate halophiles . The cultures are unable tohydrolyze inositol, sodium malonate; they do not producelysine (except for B3), indole, H2S and acid from d-mannose,sucrose, except for d-glucose, reduce nitrate to nitrite andare capable of utilizing the polypeptide substrate gelatin andsodium citrate as a carbon source. The GC content in theDNA of strains B2 andB3 was 42.8 and 37.3 %, respectively.Based on the phenotypic and genotypic characteristics, theisolated cultures have been classified as V. salinarius B2(MW386472) and V. salinarius B3 (MW386473) strains.Habitat: waters of technogenic, highly mineralizedBerikey Lake . The type strain Virgibacillus salarius DSM 18441\u0422isolated from the salt crust of the Gharsah Salt Lake in Shattal Gharsah (Sahara) in Tunisia .Biotechnological potentialof halophilic microorganismsHalophilic bacteria are increasingly being studied for theirbiotechnological potential for the production of biochemically active and resistant enzymes to alkaline pH, hightemperature and salt concentration . These multifaceted properties are useful fora variety of industries , suchas fermented food, textiles, pharmaceuticals, cosmetics,and leather production .Most producers of extracellular hydrolytic enzymes lipase,amylase, protease, inulinase, xylanase, cellulase, DNase,and pectinase are halophilic bacteria, including strains ofthe genera Halomonas and Virgibacillus .Isolation of natural strains in these studies made it possibleto discover a new species Halomonas sp. G2 (MW386470)and new strains Halomonas G1 (MW386469) and G3(MW386471), Virgibacillus B2 (MW386472) and B3(MW386473), capable of producing hydrolytic enzymesand oxidoreductase .The study confirms the biotechnological and scientific importance of halophilic denitrifying bacteria inhabiting theextremophilic ecological niches of the Peri-Caspian Lowland in the Republic of Daghestan. The strains of bacteriaof the genera Halomonas and Virgibacillus isolated provedto be not strictly confined to the salt lakes and soils of thePeri-Caspian Lowland ,having a wide distribution area, including the ecologicalniches of Bonaire Island (Netherlands Antilles) and Tunisia.Isolation and study of natural strains have revealed a newspecies Halomonassp. G2 that complement the collection ofthe already known strains \u2013 producers of industrially usefulenzymes such as amylase, protease, lactase, lipase, urease,\u03b2-galactosidase, catalase, and oxidase.The authors declare no conflict of interest.Banciu H.L., Enach M., Rodriguez R.M., Oren A., Ventosa A. Ecology and physiology of halophilic microorganisms \u2013 Thematic issuebased on papers presented at Halophiles 2019 \u2013 12th InternationalConference on Halophilic Microorganisms, Cluj-Napoca, Romania,24\u201328 June, 2019. FEMS Microbiol. Lett. 2020;366(23):1-4. DOI10.1093/femsle/fnz250.\u0412aumann P., Baumann L. 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+{"text": "Neisseria gonorrhoeae is a global health concern with worrying trends of decreasing susceptibility to also the last-line extended-spectrum cephalosporin (ESC) ceftriaxone. A dramatic increase of reported gonorrhea cases has been observed in Sweden from 2016 and onward. The aim of the present study was to comprehensively investigate the genomic epidemiology of all cultured N. gonorrhoeae isolates in Sweden during 2016, in conjunction with phenotypic AMR and clinical and epidemiological data of patients. In total, 1279 isolates were examined. Etest and whole-genome sequencing (WGS) were performed, and epidemiological data obtained from the Public Health Agency of Sweden. Overall, 51.1%, 1.7%, and 1.3% resistance to ciprofloxacin, cefixime, and azithromycin, respectively, was found. No isolates were resistant to ceftriaxone, however, 9.3% of isolates showed a decreased susceptibility to ceftriaxone and 10.5% to cefixime. In total, 44 penA alleles were found of which six were mosaic (n = 92). Using the typing schemes of MLST, NG-MAST, and NG-STAR; 133, 422, and 280 sequence types, respectively, and 93 NG-STAR clonal complexes were found. The phylogenomic analysis revealed two main lineages (A and B) with lineage A divided into two main sublineages (A1 and A2). Resistance and decreased susceptibility to ESCs and azithromycin and associated AMR determinants, such as mosaic penA and mosaic mtrD, were predominantly found in sublineage A2. Resistance to cefixime and azithromycin was more prevalent among heterosexuals and MSM, respectively, and both were predominantly spread through domestic transmission. Continuous surveillance of the spread and evolution of N. gonorrhoeae, including phenotypic AMR testing and WGS, is essential for enhanced knowledge regarding the dynamic evolution of N. gonorrhoeae and gonorrhea epidemiology.The increasing transmission and antimicrobial resistance (AMR) in  Neisseria gonorrhoeae, the causative agent of gonorrhea, is a global public health concern with increasing transmission and development of antimicrobial resistance (AMR) (ce (AMR) . The incce (AMR) . This trce (AMR) . In 2020ce (AMR) . Nonethece (AMR) . Furtherce (AMR)  and the ce (AMR) .N. gonorrhoeae, there are many different molecular AMR determinants, for example, target mutations in: penA, encoding penicillin-binding protein 2 (PBP2), associated with resistance and decreased susceptibility to ESCs and penicillins; 23S rRNA associated with macrolide resistance; gyrA and parC, encoding subunits of DNA gyrase and DNA topoisomerase IV, respectively, associated with fluoroquinolone resistance; and 16S rRNA associated with spectinomycin resistance  result in decreased influx and increased efflux of antimicrobials, respectively. These mutations decrease the antimicrobial susceptibility further  is, due to its high resolution and accuracy, an ideal method for molecular epidemiology and enables collection of comprehensive data on gonococcal population characteristics and AMR determinants internationally  in Sweden in 2016, representing 72.0% of all reported gonorrhea cases (n = 1777) in the year, were included. When multiple isolates from the same gonorrhea episode were available, isolates from extragenital sites were prioritized. Accordingly, the examined gonococcal isolates were cultured from the urethra (n = 521), rectum (n = 291), pharynx (n = 150), or other/not reported (n = 65) in men and from cervix (n = 119), pharynx (n = 44), urethra (n = 34), rectum (n = 21), vaginal swabs (n = 14), or other/not reported (n = 20) in women. Multiple gonococcal infections of the same patient were considered separate episodes if the infections were > 3 weeks apart. AMR testing was conducted using Etest, according to the instructions of the manufacturer . Minimum inhibitory concentrations  of ceftriaxone, cefixime, ciprofloxacin and spectinomycin were interpreted according to the clinical breakpoints recommended by the European Committee on Antimicrobial Susceptibility Testing. For azithromycin, no clinical breakpoints are recommended and the epidemiological cut-off value of 1 mg/L (1 v11.0) was applied (isolates with azithromycin MIC > 1 mg/L are referred to as resistant hereafter). Additionally, isolates with MIC values of 0.064 to 0.125 mg/L for ceftriaxone and cefixime were interpreted to have a decreased susceptibility , and country of infection were extracted.2 was used to calculate significance and significance was set at p < 0.05.Statistical analysis was performed using logistic regression in IBM SPSS Statistics (version 25) with calculation of a 95% confidence interval and odds ratio (OR). Pearson\u2019s \u03c7Ethical approval for the study was obtained from the Swedish Ethical Review Authority .Genomic DNA was extracted using the QIASymphony DSP virus/pathogen kit  according to the manufacturer\u2019s instructions, including RNase treatment and elution with Tris-HCl (pH 8), on the QIASymphony instrument (Qiagen). All DNA extractions were paired-end sequenced using Illumina HiSeq X platform . DNA extractions and sequencing libraries were quality controlled using Qubit BR/HS assays  and appropriate assays on the TapeStation according to manufacturer\u2019s instructions .N. gonorrhoeae genomes (n = 1279) were sequenced with 50-100-fold coverage and initially quality controlled for contamination using Kraken (v1.1.1)  (N. gonorrhoeae multi-antigen sequence typing (NG-MAST)  (All (v1.1.1) . All rea(v1.1.1) ], geneti(v1.1.1) , N. gonoNG-STAR)  and N. gNG-MAST)  sequenceNG-MAST) . The NG-NG-MAST)  based onv3.14.1)  with theN. gonorrhoeae reference strain FA1090 (accession number LT591897) to obtain a multiple sequence alignment using multiple_mappings_to_bam pipeline3 with BWA-MEM  visualized using Microreact .All WGS sequence reads are available from the ENA (PRJEB47922).n = 21) except one (V\u00e4sterbotten) were represented. Regarding origin of infection, 68.4% and 29.6% were reported as domestic (n = 875) and foreign (n = 379), respectively (n = 70), Germany (n = 32), Spain (n = 31), Denmark (n = 25), and Turkey (n = 18) (The study population included 252 women and 1027 men with median (mean) age of 24 (27.4) years and 30 (32.1) years, respectively . In totaectively . The mos(n = 18) . A highe4 and The phenotypic antimicrobial susceptibility of all gonococcal isolates by regional council is summarized in Figure 1n = 119) of all isolates (n = 1279) showed a decreased susceptibility. Cefixime resistance was found in 1.7% of isolates , and the highest resistance levels were in \u00d6sterg\u00f6tland (4.0%), Stockholm (2.4%), and Sk\u00e5ne (2.3%) (n = 134) of isolates had a decreased susceptibility to cefixime. Eighty (6.3%) isolates had a decreased susceptibility to both ceftriaxone and cefixime. One of these 80 isolates was also resistant to azithromycin, i.e., a high-level azithromycin-resistant (MIC > 256 mg/L) strain cultured from a heterosexual male with a sexual contact in Thailand. Based on the WGS sequences, 20 (90.9%) of the cefixime-resistant isolates (n = 22) had mosaic penA alleles [penA-10.001 (n = 18) and penA-34.001 (n = 2)]  cefixime-resistant isolates did not contain any mosaic penA, however, these isolates harbored PBP2 A501V and P551S alterations (penA-13.001) as well as mtrR and porB1b mutations. Overall, 44 penA alleles were found among the 1279 isolates, of which the most common were penA-2.001 (n = 325), penA-2.002 (n = 232), penA-19.001 (n = 101), penA-44.001 (n = 98), and penA-14.001 (n = 94). Ninety-two isolates (7.2%) had mosaic penA alleles: penA-34.001 (n = 60), penA-10.001 (n = 23), penA-34.008 (n = 6), penA-72.001, penA-105.001, and penA-180.001 (n = 1 each) resulting in ceftriaxone MICs < 0.002-0.125 mg/L and cefixime MICs < 0.016-0.5 mg/L. The combination of mosaic penA with mtrR and/or porB1b mutations, which further decrease the ESC susceptibility, was found in 81 isolates, of which 49 (60.5%) isolates had decreased susceptibility to ceftriaxone and 17 (21.0%) and 60 (74.1%) isolates had resistance and decreased susceptibility to cefixime, respectively. PBP2 A501V/T, P551S and G542S alterations were found in 162 (12.7%), 56 (4.4%), and 45 (3.5%) isolates, respectively. The ceftriaxone and cefixime MIC range (mean) was 0.004-0.125 (0.035) mg/L and < 0.016-0.125 (0.040) mg/L for A501V/T, 0.004-0.064 (0.034) mg/L and < 0.016-0.064 (0.029) mg/L for P551S, and 0.004-0.032 (0.016) mg/L and < 0.016-0.032 (0.020) mg/L for G542S, respectively. The combination of PBP2 A501V/T and P551S alterations was present in 48 isolates (3.8%) with ceftriaxone and cefixime MIC range (mean) of 0.004-0.125 (0.07) mg/L and < 0.016-0.25 (0.07) mg/L, respectively. The combination of PBP2 A501T and G542S alterations was present in six isolates (0.5%) with ceftriaxone and cefixime MIC range (mean) of 0.016-0.064 (0.051) mg/L and 0.016-0.064 (0.045) mg/L, respectively.No resistance to ceftriaxone was found, however, 9.3% (e (2.3%) . FurthermtrR -35 A-deletion was found in all of them, two isolates had additionally a mosaic mtrD and the remaining three isolates had porB1b mutations. Mosaic mtrR was found in 1.1% (n = 14) of isolates, mosaic mtrC in 1.1% (n = 14), mosaic mtrD in 8.4% (n = 107), and mosaic mtrE in 1.0% (n = 13). The combination of mosaic alleles in all genes of the mtrRCDE operon was found in 1.0% (n = 13) of isolates with azithromycin MICs of 0.032-1.0 mg/L. One isolate harbored mosaic alleles in all genes except mtrE (azithromycin MIC = 1.0 mg/L). Notably, one isolate, which lacked known azithromycin resistance determinants, contained the previously described GC deletion in mtrC (Azithromycin resistance was found in 16 (1.3%) isolates, and the highest resistance levels were found in Dalarna (11.1%) and Stockholm (1.8%) . High-le in mtrC  (azithroparC was found in 451 (35.3%) isolates, all except one had the GyrA S91F mutation, consisting of the amino acid substitutions D86N (n = 137), S87I/N/R (n = 311), S88P (n = 27), and E91K (n = 10).Ciprofloxacin resistance was found in 653 (51.1%) isolates . All isorpsE genes.No resistance was found to spectinomycin, including no spectinomycin resistance determinants in the 16S rRNA or mtrR  and porB1b mutations  were found in 540 (42.2%) and 356 (27.8%) isolates, respectively.In total, n = 133), ST7363 (n = 125), ST1901 (n = 93), ST1588 (n = 89), and ST7359 (n = 67). Fifty-five MLST STs represented by single isolates were found. Using NG-MAST, 422 STs were identified and the five most prevalent STs were ST5441 (n = 89), ST5793 (n = 56), ST2992 (n = 41), ST11461 (n = 39), and ST387 (n = 35). In total, 260 NG-MAST STs were found in only one isolate. With NG-STAR, 280 STs were observed and the five most prevalent ones were ST442 (n = 133), ST55 (n = 63), ST158 (n = 63), ST231 (n = 61), and ST520 (n = 52). One hundred and forty-eight NG-STAR STs were represented by single isolates. Lastly, 92 NG-STAR CCs were found and 19 isolates were ungroupable. The five most common CCs were CC442 (n = 134), CC158 (n = 100), CC63 (n = 91), CC42 (n = 70), and CC390 (n = 69). Twenty-six NG-STAR CCs were represented by single isolates.Based on the WGS sequences, the most common MLST, NG-MAST, NG-STAR STs and NG-STAR CCs by sex and sexual orientation is summarized in penA types in sublineages A1 and A2 and linage B are summarized in n = 133) and ST1901 (n = 90); NG-MAST ST5441 (n = 89) and ST2992 (n = 41); NG-STAR ST442 (n = 133) and ST90 (n = 51); and NG-STAR CCs 442 (n = 134) and 63 (n = 91). The sexual orientation of the patients in A2 was homosexual male (57.9%), bisexual male (2.6%), heterosexual male (23.4%), women (14.5%), and not reported (1.7%) (penA (49/81) predominantly belonged to A2. The isolates with decreased susceptibility to ceftriaxone (n = 89) were cultured from MSM , heterosexual men , women  and not reported . The majority  of these infections were domestic and 73.0% (n = 65) were diagnosed in Stockholm. The most common molecular types among the isolates with decreased susceptibility to ceftriaxone were MLST ST1901 (n = 34), NG-MAST ST1407 (n = 25), NG-STAR ST90 (n = 33), and NG-STAR CC90 (n = 35). The 21 (95.5%) cefixime-resistant isolates in A2 were cultured from infections acquired heterosexually  or MSM (3/21). Furthermore, 14 isolates were from domestic and seven from foreign infections, in six different countries. The majority  were diagnosed in Stockholm, however, all cefixime-resistant isolates were from patients in larger metropolitan areas. The most common molecular types among the cefixime-resistant isolates were MLST ST7363 (n = 16), NG-MAST ST13876 (n = 9), NG-STAR ST232 (n = 10), and NG-STAR CC348 (n = 15).Phylogenomic analysis revealed two main lineages (A and B), of which one (A) was subdivided into two main sublineages (A1 and A2) (d (1.7%) . Isolaten = 11), or mtrR -35 A-deletion plus mosaic mtrD (n = 2). Sexual orientation of these patients was homosexual male (46.2%), heterosexual male (15.4%) and women (38.5%). These azithromycin-resistant isolates did not belong to any predominant ST nor did they cluster together in the phylogenomic tree  also belonged to A2, and contained 23S rRNA gene mutations (n = 90), NG-MAST ST1407 (n = 33), NG-STAR ST90 (n = 51), and NG-STAR CC90 (n = 61).Resistance to ciprofloxacin was also prevalent (53.2%) in A2. The most common molecular types among the ciprofloxacin-resistant isolates in A2 were MLST ST1901 (n = 101) and ST1588 (n = 89); NG-MAST ST12001 (n = 21), ST9184 (n = 19), and ST18710 (n = 19); NG-STAR ST158 (n = 63) and ST426 (n = 22); and NG-STAR CC158 (n = 100) and CC309 (n = 48). The sexual orientation of the patients in A1 was homosexual male (32.8%), bisexual male (1.9%), heterosexual male (40.0%), women (22.5%), and not reported (2.8%) (penA). The remaining three  azithromycin-resistant isolates also belonged to A1 and appeared to represent the same gonococcal strain . All these isolates were also lacking 23S rRNA mutations and mosaic mtrD, but had mtrR -35 A-deletion plus porB1b mutations (azithromycin MICs of 2 mg/L). Furthermore, ciprofloxacin resistance was very common in A1 (90%). The most common molecular types among the ciprofloxacin-resistant isolates in A1 were MLST ST7363 (n = 101), NG-MAST ST12001 (n = 21), NG-STAR ST158 (n = 63), and NG-STAR CC158 (n = 100) (In lineage A1 (360 isolates), the most common molecular types were MLST ST7363 (d (2.8%) . A1 incln = 100) .n = 67) and ST1599 (n = 53), NG-MAST ST5793 (n = 56) and ST11461 (n = 39), NG-STAR ST55 (n = 63) and ST231 (n = 61), and NG-STAR CC42 (n = 69) and CC390 (n = 64). The sexual orientation of the patients was homosexual male (52.4%), bisexual male (0.8%), heterosexual male (22.8%), women (22.9%), and not reported (1.4%) (penA but no isolates comprised mosaic mtrD. The majority (93.3%) of isolates were ciprofloxacin susceptible.In lineage B (359 isolates), the most frequent molecular types were MLST ST7359 (d (1.4%) . Lineagen = 5) and epidemiological data  were diagnosed in metropolitan areas, especially in the capital city Stockholm. In 2016, resistance to ceftriaxone and cefixime remained low nationally; i.e., no resistance to ceftriaxone and 1.7% to cefixime. Resistance to azithromycin was also low (1.3%). The cefixime resistance was caused by penA-10.001 (n = 18), penA-34.001 (n = 2), or PBP2 A501V and P551S alterations in combination with mtrR and porB1b mutations (n = 2). The azithromycin resistance was caused by 23S rRNA A2059G (n = 1), 23S rRNA C2611T (n = 10) or mtrR -35 A-deletion plus mosaic mtrD (n = 2) or mtrR -35 A-deletion plus porB1b mutations (n = 3). None of the cefixime-resistant isolates was resistant to azithromycin. Prior to 2019, the Swedish gonorrhea guideline recommended ceftriaxone 500 mg, plus azithromycin 2 g if pharyngeal infection, for treatment of uncomplicated gonorrhea. However, from 2019, ceftriaxone 1 g monotherapy is the recommended treatment  and A1 contained the lowest proportion of MSM. Lineage B harbored most of the antimicrobial-susceptible isolates.Phylogenomic analysis showed that the isolates grouped into two main lineages as previously described , of whicpenA alleles in MLST ST1901 and ST7363 was originally documented in Japan  of the 21 Swedish regions, making preventions targeted on single sexual networks or risk group and in general gonorrhea control very challenging.The most prevalent MLST STs found in the present study have been described in previous studies. A national genomic Norwegian study described 21 STs with \u2265 10 isolates , all of in Japan  and thesin Japan , 2021. Iectively . The majd Europe . In SwedN. gonorrhoeae, NG-STAR CC  due to the lack of phylogenomic similarity of the isolates. NG-STAR with CC designation, which clusters based on AMR determinants and the closest alleles, appeared to correspond relatively well with the genome-based phylogeny and can also inform regarding resistance or decreased susceptibility to cefixime and ceftriaxone and presence of associated AMR determinants.The newly proposed classification method for -STAR CC , assigne-STAR CC , where iN. gonorrhoeae population in 2016 had a low prevalence of resistance to cefixime and no resistance to ceftriaxone. However, the limited cefixime resistance and especially the high level of decreased susceptibility to ESCs, predominantly domestically transmitted, remain worrisome and is a risk for development of resistance to ceftriaxone. The prevalence of azithromycin resistance was also low. Resistance and decreased susceptibility to ESCs and azithromycin and associated AMR determinants, such as mosaic penA and mosaic mtrD, were mainly found in the phylogenomic sublineage A2. Resistance to cefixime and azithromycin was more prevalent among heterosexuals and MSM, respectively, and both were predominantly spread through domestic transmission. Continuous surveillance of the spread and evolution of N. gonorrhoeae, including phenotypic AMR testing and WGS, is essential for enhanced knowledge regarding the dynamic evolution of N. gonorrhoeae and gonorrhea epidemiology.In conclusion, the Swedish https://www.ebi.ac.uk/ena, PRJEB47922.The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: The studies involving human participants were reviewed and approved by the Swedish Ethical Review Authority . Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.MU and RH designed, initiated, and coordinated the study. RH, DG, and LE were involved in the laboratory work, WGS, and/or bioinformatic analysis. A-KO, E-LE, and YL provided gonococcal isolates. IV and HF contributed with epidemiological data. RH, DG, and MU analyzed and interpreted all the data. RH and MU wrote the first draft of the manuscript. All authors read, commented, and approved the submitted manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Inflammatory osteolysis after total joint replacement (TJR) may cause implant failure, periprosthetic fractures, and be a severe threat to global public health. Our previous studies demonstrated that melatonin had a therapeutic effect on wear-particles induced osteolysis. Gut microbiota is closely related to bone homeostasis, and has been proven to be affected by melatonin. However, whether melatonin could play its anti-osteolysis effects through reprogramming gut microbiota remains elusive.Here, we demonstrated that melatonin could alleviate Ti-particles induced osteolysis, while this therapeutic effect was blocked by antibiotic cocktail treatment. Interestingly, transplantation of fecal microbiota from mice treated with melatonin reappeared the same beneficial effect. Analysis of the 16S rRNA revealed that melatonin could reverse dysbacteriosis triggered by osteolysis, and elevate the relative abundance of some short chain fatty acid (SCFA) producing bacteria. Moreover, butyrate was enriched by exogenous melatonin administration, while acetate and propionate did not show an evident difference. This was consistent with the results of the metagenomic approach (PICRUSt2) analysis, which revealed a general increase in the synthetic enzymes of butyrate. More importantly, direct supplementation of butyrate could also recapitulate the anti-osteolysis effect of melatonin. Further analysis identified that butyrate alleviated osteolysis via activating its receptor GPR109A, and thus to suppress the activation of NLRP3 inflammasome triggered by Ti-particles.Taken together, our results suggested that the benefits of melatonin mainly depend on the ability of modulating gut microbiota and regulating butyrate production.The online version contains supplementary material available at 10.1186/s12951-021-00915-3. As the essential treatment for end-stage arthritis, total joint replacement (TJR) could effectively relieve joint pain, stiffness, and improve joint function . However.  ** p < .01). Figure S3. Principle coordinate analysis (PCoA) plot between Ti-trans and MT-trans groups based on the Bray Curits distance. n = 6. Figure S4. Ti-particles induced osteolysis could be blocked by the treatment of either NLRP3 or caspase-1 inhibitor. (A) Representative view of calvarium from sham, Ti, MCC950, and Ac-YVAD-CMK groups via Micro-CT 3D reconstruction. (B-D) Quantification of bone erosion parameters (BV/TV) bone volume to tissue volume ratio, (BMD) bone mineral density, and total porosity. n=6. (E) Percentage of osteoclasts surface per bone surface . n = 5.(F) H&E and (G) TRAP staining of calvarial slices from each group. (H) H&E and (I) TRAP staining of calvarial slices from Sham, Ti and Butyrate group. Results are expressed as mean \u00b1 SEM (One-way ANOVA[post hoc:SNK] *** p < .001). Figure S5. Quantification of NLRP3, Caspase-1, IL-1\u03b2 and GPR109A staining. (A) Quantification of NLRP3 immunohistochemical staining using integrated optical density/specimen area (IOD/Area). n=5. (B) Quantification of Caspase-1 immunofluorescence staining using mean density (integrated density/specimen area). n=5. (C) Quantification of IL-1\u03b2 immunohistochemical staining using integrated optical density/specimen area (IOD/Area). n=5. (D) Quantification of GPR109A immunohistochemical staining. n=5. Results are expressed as mean \u00b1 SEM ."}
+{"text": "Anxiety Disorder (AD) is common in inpatient pediatric burn patients and likely related to pain/stress associated with acute care. This study ascertained if burn survivors reported higher anxiety levels based on sex, visibility of scars, or TBSA \u2265 50%.Burn-injured youth completed the Screen for Child Anxiety Related Disorders (SCARED) with parental consent. This 41 item self-report measures DSM-IV pediatric anxiety disorder symptoms: panic disorder (PD), separation anxiety (SA), generalized anxiety disorder (GAD), social phobia (SP) school phobia (SCP) and total anxiety (TA). The percentage of respondents above threshold for each disorder was calculated.112 survivors, mean age of 13, included boys (51%) & girls (49%). 83 reported visible scars. Females had higher percentages for TA (53%) vs. males (21%) (p < 0.001), PD (47%) vs. (7%) (p< 0.001), GAD (40%) vs. (16%) (p < 0.005), & SA (51%) vs. (21%) (p < 0.001). Youth with TBSA \u2265 50% (n=22) had higher precents for GAD (46%) vs. < 50% (24%) (p < 0.01). The visibly scarred had higher percent for GAD (38%) vs. hidden (7%) (p< .01).Female, visibly scarred, and patients with burns > 50% revealed increased AD symptoms. AD may be chronic, interfere with a child\u2019s home & school function and lead to chronic distress, substance abuse, and isolation. Screening for anxiety in burn-injured youth is recommended."}
+{"text": "The primary analysis of the STAT study demonstrated the feasibility, efficacy, and safety of using DTG/3TC as a first-line regimen in a test-and-treat setting through 24 weeks, with therapy adjustments for baseline resistance or hepatitis B virus (HBV) co-infection. Here we present secondary analyses through Week 48 of virologic outcomes in participants by baseline viral load (VL). 2, then antiretroviral therapy (ART) was potentially adjusted and participants remained on study. Efficacy analyses included proportion of participants with HIV-1 RNA < 50 c/mL regardless of ART regimen at Week 48, among all participants  and among participants with available HIV-1 RNA data at Week 48 . STAT is a single-arm study of treatment-naive adults with HIV-1 infection who initiated DTG/3TC \u2264 14 days after HIV-1 diagnosis without availability of screening/baseline laboratory results. If baseline testing indicated DTG or 3TC resistance, HBV co-infection, or creatinine clearance < 30 mL/min/1.73 mOf 131 enrolled, DTG/3TC treatment was adjusted in 10 participants, and of those with available data (n=7), all (100%) achieved HIV-1 RNA < 50 c/mL at Week 48. At Week 48, 82% (107/131) of all participants  and 97% (107/110) of those with available data  achieved HIV-1 RNA < 50 c/mL. Of participants with baseline VL \u2265 500,000 c/mL, 89% (17/19) achieved HIV-1 RNA < 50 c/mL at Week 48; the remaining 2 withdrew from study. Of participants with baseline VL \u2265 1,000,000 c/mL, 90% (9/10) achieved HIV-1 RNA < 50 c/mL at Week 48 (Table); the remaining participant withdrew consent. Of the 17 participants with baseline VL \u2265 500,000 c/mL with available data through Week 48, 76% (13/17) achieved virologic suppression by Week 24. One participant with baseline VL \u2265 500,000 c/mL switched from DTG/3TC before the Week 48 assessment. Of the 9 participants with baseline VL \u2265 1,000,000 c/mL with available data through Week 48, most participants  were suppressed by Week 24.Figure 1. Virologic outcomes at Week 48, overall and by baseline VL and CD4+ cell count: ITT-E missing = failure analysis.Figure 2. Virologic outcomes at Week 48, overall and by baseline VL and CD4+ cell count: observed analysis.Table. Viral Load by Study Visit Among Participants with Baseline HIV-1 RNA \u22651,000,000 c/mLThese data provide evidence for the efficacy and feasibility of using DTG/3TC as a first-line regimen in a test-and-treat setting, including among participants with very high baseline VL.Charlotte-Paige M. Rolle, MD MPH, Gilead Sciences Janssen Infectious Disease ViiV Healthcare  Tulika Singh, MD MS AAHIVS, Gilead ViiV  Moti Ramgopal, MD FIDSA, Abbvie Gilead Janssen Merck ViiV  Dushyantha Jayaweera, MD, mrcog(uk), face, Gilead (Research Grant or Support)Janssen (Research Grant or Support)viiv (Research Grant or Support) Peter Leone, MD, viiv healthcare (Employee) Jessica Matthews, BS, ViiV Healthcare (Employee) Michael Cupo, Ph.D., GlaxoSmithKline (Employee) Mark Underwood, PhD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Kostas Angelis, PhD, GSK  Brian Wynne, MD, ViiV Healthcare  Deanna Merrill, PharmD, MBA, AAHIVP, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Christopher T. Nguyen, MD, ViiV Healthcare (Employee) Jean A. van Wyk, MB,ChB, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee) Andrew Zolopa, MD, GlaxoSmithKline (Shareholder)ViiV Healthcare (Employee)"}
+{"text": "Due to antiretroviral treatment success, individuals with HIV are living longer. People aging with HIV  may be more likely to experience nutritional risk compared to their HIV-negative counterparts due to biopsychosocial factors. The DETERMINE checklist measure accounts for social and economic factors as well as aspects of the aging process that are not typically considered when examining nutritional risk and are important for PAWH. The current study examined nutritional risk and health-related quality of life (HRQoL) in PAWH using the DETERMINE checklist and PROMIS t-scores  through secondary analyses of 158 participants in the Strengthening Therapeutic Resources in Older patients agiNG with HIV (STRONG) study. DETERMINE nutritional risk scores (0-21) were separated into 4 groups . The sample was 55% male, 94% Black/African American and had a mean age=59 (SD=5.5). Most of the sample (74%) were at high or very high nutritional risk and low HRQoL t-score: physical M=43.7 (SD=9.5), and mental M=45.7 (SD=10.1). Mental and physical HRQoL were significantly (p<.001) associated with nutritional risk group as tested through linear regressions. Means were as follows: physical HRQoL low-risk M=53.4 (SD=10.6), moderate-risk M=47.4 (SD=8.9), high-risk M=43.5 (SD=8.1), very high-risk M=38.4 (SD=8.9); mental HRQoL low-risk M=54.0 (SD=8.9), moderate-risk M=49.1(SD=7.9), high-risk M=46.1(SD=9.5), and very high-risk M=39.5 (SD=9.7). These associations remained significant after controlling for age and sex. Higher nutritional risk as measured by the DETERMINE checklist in PAWH was associated with poorer physical and mental HRQoL."}
+{"text": "In a Phase 3 trial, the Janssen COVID-19 vaccine, Ad26.COV2.S, showed robust efficacy against severe\u2013critical COVID-19 in countries where different SARS-CoV-2 variants were circulating. We evaluated Ad26.COV2.S-elicited antibody neutralizing activity against variants of concern (VOC) B.1.1.7 (Alpha), B.1.351 (Beta), and B.1.617.2 (Delta) in sera from participants in clinical trials following a single dose of Ad26.COV2.S.10 viral particles [vp]) against VOC were assessed by wild-type virus neutralizing (wtVNA) and pseudovirion neutralization (psVNA) assays in sera from participants in Phase 1/2a and Phase 3 clinical trials, respectively. Geometric mean titers (GMTs) were determined at Days 29 and 71 after vaccination.Neutralizing activities of Ad26.COV2.S . Day 71 titers against the Delta variant were maintained for at least 8 months following a single dose of Ad26.COV2.S (5 x 1010 vp). In serum samples from Phase 3 participants (n = 8), psVNA titers against VOC were lower than the original strain at Day 71 after vaccination, with the lowest titers observed for the Beta variant . Smaller reductions in Nab titers for VOC were observed in the psVNA assay compared to wtVNA.In serum samples from Phase 1/2a participants (n = 6), at Day 29 after 1 dose of Ad26.COV2.S, wtVNA titers against VOC were lower than for the original strain (GMT = 573), with GMT = 65, 14, and 15 for Alpha, Beta, and Delta, respectively, representing 8.8-, 40.9-, and 37.7-fold decreases. By Day 71 after vaccination (n = 14), fold differences between the original strain (GMT = 375) and VOC  were smaller  than at Day 29, suggestive of B-cell maturation , B.1.351 (Beta), and B.1.617.2 (Delta) lineages in serum samples from participants who received Ad26.COV2.S. n = 6 samples at Day 29 and n = 14  samples at Day 71 after vaccination with a single dose of Ad26.COV2.S (5 x 10^10 vp dose level) were analyzed in wild-type virus neutralization assays against the SARS-CoV-2 Victoria strain , the B.1.1.7  the B.1.351 , and the B.1.617.2  lineages. Dots represent the IC50 (inhibitory concentration) titers per participant. Geometric mean titers (GMTs) and fold decrease in neutralizing activity between the original Victoria strain and each lineage are shown.Ad26.COV2.S-elicited serum neutralizing activity against VOC showed an overall decrease in titers relative to the original strain that was largest for the Beta variant, even though vaccine efficacy against severe\u2013critical COVID-19 was maintained in countries where these variants were circulating versus in countries where they were not circulating. Over time, titers against variants increased, suggesting B-cell affinity maturation leading to increasing coverage of VOC.Mathieu Le Gars, n/a, Johnson & Johnson  Jerald Sadoff, MD, Johnson & Johnson  Mandy Jongeneelen, n/a, Johnson & Johnson  Dirk Heerwegh, n/a, Janssen Research and Development (Employee) Georgi Shukarev, MD, Janssen (Employee) Carla Truyers, n/a, Janssen Research and Development (Employee) Anne Marit de Groot, n/a, Johnson & Johnson (Employee) Gert Scheper, n/a, Johnson & Johnson  Jenny Hendriks, n/a, Johnson & Johnson  Boerries Brandenburg, n/a, Johnson & Johnson  Frank Struyf, n/a, Johnson & Johnson  Johan Van Hoof, n/a, Johnson & Johnson  Macaya Douoguih, MD, MPH, Janssen (Employee) Hanneke Schuitemaker, PhD, Johnson & Johnson"}
+{"text": "BRCA. In this study, we investigated the somatic mutation of DNA damage response genes in epithelial ovarian cancer patients using a multiple-gene panel with next-generation sequencing. In all, 69 serous, 39 endometrioid and 64 clear cell carcinoma patients were enrolled. Serous carcinoma patients (69.6%) had higher percentages of DDR gene mutations compared with patients with endometrioid (33.3%) and clear cell carcinoma (26.6%) . The percentages of DDR gene mutations in patients with recurrence  or cancer-related death  were higher than those without recurrence or living patients. In endometrioid carcinoma, patients with \u22652 DDR gene mutations had shorter PFS  and OS  than those with one mutation or none. In clear cell carcinoma, patients with \u22652 DDR gene mutations had significantly shorter PFS  and OS  than those with 1 DDR mutation or none. In the EOC patients, somatic DDR gene mutations were associated with advanced-stage tumor recurrence and tumor-related death. Type I EOC patients with DDR mutations had an unfavorable prognosis, especially for clear cell carcinoma.DNA damage response (DDR) is important for maintaining genomic integrity of the cell. Aberrant DDR pathways lead to accumulation of DNA damage, genomic instability and malignant transformations. Gene mutations have been proven to be associated with epithelial ovarian cancer, and the majority of the literature has focused on  Epithelial ovarian carcinoma (EOC) is a major cause of death in women worldwide, and patients are usually diagnosed at an advanced stage with a 5-year survival of less than 50% ,2,3,4. CBRCA mutations or homologous recombination deficiency (HRD) status is critical for selecting potential patients, but both positive and negative patients as defined by current HRD assays benefited from PARPi [Precision medicine is the current direction for cancer management depending on the specific genetic or molecular features of cancer. There are several subtypes of EOC\u2014high-grade serous, clear cell, endometrioid, mucinous and low-grade serous\u2014that could be viewed as distinct diseases for their differences in clinical course and pathological features ,7. To daom PARPi ,13,14,15BRCA 1/2 genes participating in HR and maintaining PARPi therapy for BRCA-mutated EOC is a good example of synthetic lethality [DNA damage response (DDR) is important for maintaining a cell\u2019s genomic integrity, and the DDR pathway is composed of various molecules that detect DNA damage, activate cell-cycle checkpoints, trigger apoptosis, and coordinate DNA repair ,17,18. Sethality . Severalethality ,17. UndeThe study protocol was approved by the National Taiwan University Hospital Research Ethics Committee . Informed consent from all participants was obtained and the methods were performed in accordance with the guidelines and regulations. From December 2015 to October 2018, 172 women diagnosed with epithelial ovarian cancer who had received debulking surgery and adjuvant chemotherapy were enrolled. The cancerous tissue specimens collected during debulking surgery were immediately frozen in liquid nitrogen and stored at \u221270 \u00b0C. A portion of the tissue specimens were sent for pathological examinations to confirm the diagnosis and ensure tumorous tissue sufficient for the following experiments. Clinical data were obtained from medical records, including age, cancer stage, the findings during debulking surgery, treatment course and recurrence. Optimal debulking surgery was defined as a maximal residual tumor size <1 cm following surgery. The tumor grade based on International Union Against Cancer criteria, and cancer stage was based on International Federation of Gynecology and Obstetrics (FIGO) criteria . All patWe selected 60 genes involved in DNA damage response (DDR) for the gene panel , includi260/280 ratio.Genomic DNA was isolated using a QIAGEN Genomic DNA extraction kit according to the manufacturer\u2019s instructions . The purity and concentration of the genomic DNA were checked by agarose gel electrophoresis and the ODThe genomic DNA was fragmented with Covaris fragmentation protocol . The size of the fragmented genomic DNA was checked by Agilent Bioanalyzer 2100  and NanoDrop spectrophotometer . The target gene library was generated with NimblGen capture kits . The samples were sequenced by Illumina MiSeq with paired-end reads of 300 nucleotides.The analysis algorithm was conducted according to our previous protocol . BrieflyThe sequence variants were classified according to the IARC variant classification . The patp value of less than 0.05.All statistical analyses were performed using the Statistical Package for Social Sciences software package  and R . One-way ANOVA was used to compare continuous variables and a chi-squared test was used for categorical variables. Survival curves were generated using the Kaplan\u2013Meier method, and differences were calculated using the log-rank test. A multivariate Cox\u2019s regression model was used to evaluate the prognostic factors for progression-free survival (PFS) and overall survival (OS). Statistical significance was set as a There were 172 EOC patients enrolled: 69 serous, 39 endometrioid and 64 clear cell carcinomas . There whttp://www.ncbi.nlm.nih.gov/snp, accessed on 28 September 2021) and bioinformatic analyses , MUTYH (6.4%) and BRCA2 (5.8%) for all patients. Serous carcinoma\u2014TP53 (56.5%), BRCA2 (5.8%) and RAD51C (5.8%); endometrioid carcinoma\u2014TP53 (15.4%), ATM (12.8%) and MSH2 (7.7%); clear cell carcinoma\u2014MUTYH (9.4%), TP53 (4.7%), BRCA2 (3.1%) and ERCC8 (3.1%). The top three prevalent mutated subgroups of DDR genes were CCR (30.8%), HR (10.5%) and BER (7.0%) for all patients. Serous carcinoma\u2014CCR (58.05%), HR (15.9%) and BER (5.8%); endometrioid carcinoma\u2014CCR (23.1%), MMR (15.4%) and HR (7.7%); clear cell carcinoma\u2014BER (9.4%), CCR (6.3%) and HR (6.3%). For detailed information, please refer to The pattern of prevalent mutated DDR genes was different among the histological subtypes . The prop = 0.048, chi-squared test). Endometrioid carcinoma (15.38%) had a higher percentage of MMR mutations than those of serous carcinoma (2.90%) and clear cell carcinoma (4.69%) . Low-grade tumors had a higher percentage of MMR mutations compared with high-grade tumors . Type II tumors had a higher percentage of DSBR mutations than type I tumors . Serous carcinoma (57.97%) had a higher percentage of CCR mutations than those of endometrioid carcinoma (23.08%) and clear cell carcinoma (6.25%) . Type II tumors had higher percentage of CCR mutations than those of type I tumors . The advanced-stage patients had a higher percentage of CCR mutations than the early-stage patients . The recurrent patients had a higher percentage of CCR mutations than those without recurrence . Patients who died of EOC had higher percentages of CCR mutations than living patients . Serous carcinoma (69.57%) had higher percentage of DDR mutations than those of endometrioid carcinoma (33.33%) and clear cell carcinoma (26.56%) . Type II tumors had a higher percentage of DDR mutations than type I tumors . The advanced stage patients had higher percentage of DDR mutations than the early-stage patients . Recurring patients had a higher percentage of DDR mutations than those without recurrence . Patients who died of EOC had a higher percentage of DDR mutations than living patients .We evaluated the correlations between the mutation of DDR genes, the clinicopathologic parameters and outcome of the EOC patients. As shown in p = 0.0072, log-rank test, p = 0.022, log-rank test, p = 0.56, log-rank test, p = 0.47, log-rank test, p = 0.0035, log-rank test, p = 0.015, log-rank test, p = 0.0056, log-rank test, p = 0.0046, log-rank test, EOC patients without DDR gene mutation had longer progression-free survival (PFS) (p = 0.011), 1 DDR gene mutation (HR: 1.71 (1.12\u20132.60), p = 0.013), endometrioid carcinoma (HR: 0.17 (0.08\u20130.37), p < 0.001), type II tumor (HR: 2.69 (1.81\u20134.00), p < 0.001), advanced-stage carcinoma (HR: 5.29 (3.16\u20138.85), p < 0.001), high-grade tumor (HR: 5.57 (2.26\u201313.70), p < 0.001) and optimal debulking surgery (HR: 0.28 (0.18\u20130.41), p < 0.001) were significant in the univariate Cox regression model (p = 0.001) and optimal debulking surgery (HR: 0.51 (0.32\u20130.80), p = 0.004) were important prognostic factors in the multivariate analysis. Cancer-related death with TLS gene mutation (HR: 33.76 (3.95\u2013289.00), p = 0.001), 1 DDR gene mutation (HR: 1.96 (1.20\u20133.20), p = 0.007), endometrioid carcinoma (HR: 0.12 (0.04\u20130.38), p < 0.001), type II tumor (HR: 1.88 (1.19\u20132.96), p = 0.007), advanced-stage carcinoma (HR: 6.84 (3.28\u201314.25), p < 0.001), high-grade tumor (HR: 17.97 (2.50\u2013129.29), p = 0.004) and optimal debulking surgery (HR: 0.26 (0.16\u20130.41), p < 0.001) were significant in the univariate Cox regression model. Type II tumor (HR: 0.35 (0.20\u20130.60), p < 0.001), TLS gene mutation (HR: 9.57 (1.08\u201384.83), p = 0.042), advanced-stage carcinoma (HR: 4.82 (2.09\u201311.09), p < 0.001) and optimal debulking surgery (HR: 0.38 (0.22\u20130.64), p < 0.001) were important prognostic factors in the multivariate analysis.Tumor recurrence with CCR gene mutation (HR: 1.68 (1.12\u20132.50), on model . AdvanceATM, BRCA1/2, BRIP1, MLH1, MSH2, MSH6, PALB2, RAD51C and RAD51D [BRCA test alone. The percentage of BRCA 1/2 somatic mutation in serous carcinoma was 7.2, which was compatible with the 6\u20137% in previous studies [BRCA HR somatic mutation of our study was more than 10% in serous and endometrioid carcinomas, and the MMR somatic mutation was around 15% in endometrioid carcinomas, which was compatible with the previous study [Our study showed that nearly half of the epithelial ovarian cancer (EOC) patients had DNA damage response (DDR) gene mutations with varied proportions of histological subtypes. Two-thirds of serous adenocarcinoma patients, one-third of endometrioid adenocarcinoma patients and one-fourth of clear cell carcinoma patients had DDR gene mutations. Our DDR gene panel consisted of the genes involved in single-strand break repair, double-strand break repair and cell cycle regulation, including the genes recommended by National Comprehensive Cancer Network (NCCN) guidelines as cost-effective tools for assessing the lifetime risk of EOC, such as d RAD51D . The maj studies ,35,36,37us study .ARID1A, PIK3CA, KRAS and PPP2R1A) might be related to chromatin remodeling, cell proliferation, cell cycle checkpointing and cytoskeletal organization [ARID1A, PIK3CA, PPP2R1A or TP53 in ovarian clear cell carcinoma did not correlate well with the prognosis [ARID1B, ARID3A, CREBBP, CSMD3, CTNNB1, LPHN3, LRP1B, MAGEE1, MLH1, MLL3, MUC4, PIK3R1, PTEN and TP53 [Our study showed that ovarian clear cell carcinoma patients with DDR gene mutations had an unfavorable survival prognosis. Those who had somatic DDR mutations were significantly associated with advanced-stage carcinomas, tumor recurrence and tumor-related death. The trend was different in histological subtypes as serous carcinomas or type II tumors with DDR mutation showed a better survival trend. Non-serous or type I EOC patients with DDR mutations had a poor prognosis, especially in clear cell carcinoma. Ovarian clear cell carcinoma is an aggressive drug-resistant subtype of EOC in association with endometriosis and glycogen accumulation. It accounts for about 5\u201313% of all EOCs in Western populations, but up to 20\u201325% in East Asia, including Taiwan [nization ,47,48,49rognosis . Other iand TP53 ,46,48,49BRCA gene tests or companion HRD assays are currently suggested for PARPi, but there are unmet problems that need to be resolved [BRCA (tBRCA) mutations, including germline (gBRCA) or somatic (sBRCA), derived the greatest benefit from PARPi maintenance therapy [BRCA mutation, and another 6\u20137% patients had an sBRCA mutation with a negative gBRCA test [BRCA mutated patients tested negative for tBRCA [BRCA HR gene mutations were usually pooled together to interpret the association with clinical outcomes in previous studies because of their relatively low prevalence [BRCA somatic mutations  derived benefit from olaparib in study 19 [BRCA HR gene mutations , but the sensitivity in discriminating a rucaparib response was only 11% [BRCA wild type EOC patients still benefitted from PARPi, which indicated that a BRCA test by itself was inadequate for selecting EOC patients for PARPi [BRCA HR genes could be used to predict a PARPi response, especially in non-serous EOC patients.Our DDR gene panel could provide a scientific rationale for patient selection in future clinical trials that target DNA damage repair response pathways, especially in clear cell carcinoma. resolved ,14,15,20resolved ,50,51. G therapy ,13,14,15RCA test ,35,36,37or tBRCA ,53,54. Tevalence ,56,57. Tstudy 19 . In ARIEonly 11% . HoweverATM, BRCA1/2, BRIP1, MLH1, MSH2, MSH6, PALB2, RAD51C, RAD51D and STK11 to assess the lifetime risk of EOC [BRCA gene, there is a difference among laboratories in the VUS reporting rate (3\u201350%), detection protocols and management strategies [There were limitations to our study. First, germline gene mutations were not investigated. These not only inform the patients but also identify family members of the possible risk of malignancy ,53,54. Tk of EOC , but howk of EOC ,61,62. Tk of EOC . Even inrategies . FurtherOur study found that nearly half of the EOC patients had DDR gene mutations of varying proportions in the histological subtypes. Patients with somatic DDR mutations were significantly associated with advanced-stage carcinoma, tumor recurrence and tumor-related death. Type I EOC patients with DDR mutations had an unfavorable prognosis, especially for clear cell carcinoma. A broad multiple-gene DDR panel would provide not only comprehensive information of gene mutations but also a rationale for a future study of a novel therapy target for DNA damage response pathways."}
+{"text": "Correction to: BMC Pharmacol Toxicol 20, 5 (2019)https://doi.org/10.1186/s40360-018-0280-8After publication of the original article , an erroThe incorrect sentence is:P < 0.0001).SIB peak plasma concentrations were 207.1 mg/L for SPC and 12.6 mg/L for SM tablets. All pharmacokinetic parameters differed significantly between formulations (The correct sentence is:P < 0.0001).SIB peak plasma concentrations were 207.1 ng/L for SPC and 12.6 ng/L for SM tablets. All pharmacokinetic parameters differed significantly between formulations (The original article has been corrected."}
+{"text": "Breast cancer is the most common cancer in women . Triple-First, we generated a cisplatin (DDP)-resistant TNBC cell line, MDA-MB-231/DDP, which is 5-fold more resistant to cisplatin than its parental line MDA-MB-231  Fig.\u00a0a. IntereP\u2009=\u20090.005). Consistently, in both validation cohort of 128 samples . Likewise, low-SNORD33 group showed significantly reduced overall survival (OS) compared with high-SNORD33 group , validation group  and combined cohorts   patients who received first-line platinum-based chemotherapy Fig. f. In theles Fig. a, low-SN38) Fig.  revealedROC test was used to evaluate the value of SNORD33 in predicting PFS in mTNBC patients treated with platinum-based regimens in the combined cohort. SNORD33 signature showed the strongest predictive value (AUC\u2009=\u20090.652) for PFS, compared with other single clinicopathological risk factors , suggesting better performance of SNORD33 signature as a surrogate predictor for platinum-based chemotherapy outcome Fig. m. Based P\u2009=\u20090.023) than high-SNORD33 group  [RPL13A (host gene), DICER enzyme (an endoribonuclease), or SNORD32a (homologue to SNORD33 modifying 18S rRNA during cell proliferation) in MDA-MB-231 cells  . It part(RPL13A) . Does SNlls Fig.\u00a0a. The insis Fig. b. We obssis Fig. b. We thenes Fig. c. Down-rnes Fig. . ConsistTogether, we identified plasma SNORD33 signature as a predictor for the sensitivity of platinum-based chemotherapy in mTNBC patients. Furthermore, we showed that MeCP2 could convey SNORD33 associated platinum resistance. Our findings may have immediate translational relevance for precision platinum-based chemotherapy.Additional file 1. Methods.Additional file 2: Supplementary Figure 1. Aberrantly expressing RNA in 231/DDP cells. Supplementary Figure 2. SNORD33 knockdown increases proliferation and decreases apoptosis of TNBC cells. Supplementary Figure 3. Reduced SNORD33 level is correlated with poor prognosis of mTNBC patients who received first-line platinum-based chemotherapy. Supplementary Figure 4. Increased cell viability was observed in cisplatin treated cell lines with SNORD33 knockdown. Supplementary Figure 5. Reduced SNORD33 level correlates with poor prognosis of non-small cell lung cancer (NSCLC) patients who received first-line platinum-based chemotherapy. Supplementary Figure 6. MeCP2 is a candidate protein binding with SNORD33. Supplementary Figure 7. Down-regulation of MeCP2 partially rescues SNORD33 knockdown increased cell colony formation. Supplementary Figure 8. Down-regulation of MeCP2 rescues SNORD33 knockdown decreased cell apoptosis and induced alteration of apoptotic markers.Additional file 3: Supplementary Table 1. Baseline characteristics in mTNBC patients received first-line platinum-containing regimens. Supplementary Table 2. Univariate and multivariate analysis of prognostic factors associated with progression-free survival and overall survival in the combined cohort. Supplementary Table 3. Baseline characteristics in mTNBC patients received first-line non-platinum-containing regimens. Supplementary Table 4. Baseline characteristics in lung adenocarcinoma patients received first-line platinum-containing regimens. Supplementary Table 5. Univariate and multivariate analysis of prognostic factors associated with progression-free survival in lung adenocarcinoma patients.Additional file 4."}
+{"text": "CMV infection is common post-kidney transplant (KT). Valganciclovir (VGC) prophylaxis (Px) has lessened CMV infection among high-risk (CMV D+/R-) KT recipients (KTRs), but VGC can induce neutropenia. We quantified the burden of CMV infection among CMV D+/R- KTRs and healthcare resources required to manage these patients (pts).Retrospective study of pts undergoing KT between Jan 2014-Dec 2018. Study and control groups (gps) were CMV D+/R- and R+ KTRs, respectively. Standard post-KT immunosuppression was tacrolimus and mycophenolate mofetil (MMF). D+/R- and R+ KTRs received VGC Px (900 mg/day) for 6 and 3 months (mos), respectively.Clinical characteristics did not differ between D+/R- (n=131) and R+ (n=140) pts. Median VGC Px duration was longer for D+/R- . Within the first 6 mos post KT, a higher proportion of D+/R- KTRs received \u22651-course of granulocyte-stimulating factor (G-CSF) . VGC Px was stopped prematurely/intermittently in 20% and 10% of D+/R- and R+, respectively, due to neutropenia (p=0.02); corresponding data for stopping MMF for \u22651 mos were 32% and 21% (p=.05). 50% of D+/R- pts received < 3 mos Px. Leukopenia prompted hospitalization in 3% of D+/R- vs 0% of R+ pts (p=.05). CMV infections did not differ between gps ; however, VGC-resistant CMV was higher in D+/R- gp . Between 6-12 mos post-KT, D+/R- KTRs had higher rates of CMV infection , VGC resistance , hospitalization due to CMV , MD intervention , and infectious disease (ID) referral . 57% of CMV resistance was observed in pts who prematurely stopped VGC. Hospitalizations were longer for CMV infections in D+/R- KTRs . There was a trend toward higher rejection for D+/R- KTRs .Universal VGC Px in D+/R- KTR remains challenging and requires significant resources for monitoring and intervention for neutropenia, including MD involvement and ID referral. Intermittent/premature stop of VGC may have led to VGC-resistant CMV,and stop of MMF may have led to a trend of higher cellular rejection at 1 yr. There is critical need for new CMV agents with a better safety profile.Amit D. Raval, PhD, Merck and Co., Inc. (Employee) Yuexin Tang, PhD, JnJ Merck & Co., Inc.  Cornelius J. Clancy, MD, Merck (Grant/Research Support) Minh-Hong Nguyen, MD, Merck (Grant/Research Support)"}
+{"text": "Carbapenem (Carb) minimum inhibitory concentration (MIC) breakpoints were lowered by CLSI in 2010 and recognized by FDA in 2012. Adoption of revised breakpoints is often slow, which may lead to under-reporting of Carb non-susceptibility (NS) by facilities. We compare facility-reported rates of Carb-NS ENT to the CLSI MIC breakpoints for a large nationwide collection of isolates in the United States (US) from 2016-2019.All adults with a positive non-contaminant ENT culture  in ambulatory/inpatient settings from up to 300 US hospitals from 2016-2019 were evaluated (BD Insights Research Database). Facility-reported Carb-NS was defined as: susceptible (S), intermediate (I) or R to ertapenem (ETP), imipenem (IPM), meropenem (MEM) and/or doripenem (DOR) per commercial panels. Where available, MICs were interpreted using CLSI 2010 MIC breakpoints (\u00b5g/ml): \u2264 0.5 (S), 1 (I), \u2265 2 (R) for ETP and \u22641 (S), 2 (I), and \u2265 4 (R) for IPM/MEM/DOR. For evaluable ENT isolates we compared susceptibility results as reported by the facility to CLSI MIC breakpoints.Overall, 77.4%  and 90.6%  non-duplicate ENT isolates with facility-reported susceptibility results also had interpretable MIC results for ETP and IPM/MEM/DOR, respectively (Tables). ETP S rates were 99.3% and 99.1% as reported by facilities and using CLSI criteria, respectively. S rates of other Carbs were 98.9% and 98.4% by facility reporting and CLSI criteria, respectively. Systematic application of CLSI breakpoints under-reported EPT-I and \u2013R isolates by 24.2% and 16.4%, respectively, and identification of IPM/MEM/DOR-I and \u2013R isolates by 31.3% and 22.7%, respectively.Systematic application of CLSI breakpoints in 2016-19 would have had minimal impact on ENT S rates in the US. However, facility reporting failed to identify 18.8% of ETP I or R and 26.5% of IPM/MEM/DOR I or R isolates. The clinical implications of this observation are unknown. Facilities should know their local epidemiology, decide if under-reporting might be an issue, and assess if there is any impact on their patients.Vikas Gupta, PharmD, BCPS, Becton, Dickinson and Company  Kalvin Yu, MD, BD (Employee) Jason M Pogue, PharmD, BCPS, BCIDP, Merck (Consultant)QPex (Consultant)Shionogi (Consultant)Utility Therapeutics (Consultant)VenatoRX (Consultant) Janet Weeks, PhD, Becton, Dickinson and Company (Employee) Cornelius J. Clancy, MD, Merck (Grant/Research Support)"}
+{"text": "Scientific Reports 10.1038/s41598-021-98868-y, published online 30 September 2021Correction to: The original version of this Article contained an error in the order of the References 16 and 17, which was incorrectly given as:et al. Vitamin D increases glucocorticoid efficacy via inhibition of mTORC1 in experimental models of multiple sclerosis. Acta Neuropathol. 10.1007/s00401-019-02018-8 (2019).16. Hoepner, R. FASEB J23, 3649\u20133658 (2009).17. Ayroldi E, Riccardi C. Glucocorticoid\u2010induced leucine zipper (GILZ): a new important mediator of glucocorticoid action. The correct order of the References is listed below:FASEB J23, 3649\u20133658 (2009).16. Ayroldi E, Riccardi C. Glucocorticoid\u2010induced leucine zipper (GILZ): a new important mediator of glucocorticoid action. et al. Vitamin D increases glucocorticoid efficacy via inhibition of mTORC1 in experimental models of multiple sclerosis. Acta Neuropathol. 10.1007/s00401-019-02018-8 (2019).17. Hoepner, R. The original Article has been corrected."}
+{"text": "Effective treatment of glioma requires a nanocarrier that can cross the blood\u2013brain barrier (BBB) to target the tumor lesion. In the current study, elemene (ELE) and cabazitaxel (CTX) liposomes were prepared by conjugating liposomes with transferrin (Tf) and embedding the cell membrane proteins of RG2 glioma cells into liposomes , which exhibited effective BBB infiltration to target glioma.The findings showed that Tf-ELE/CTX@BLIP was highly stable. The liposomes exhibited highly significant homologous targeting and immune evasion in vitro and a 5.83-fold intake rate compared with classical liposome (ELE/CTX@LIP). Bioluminescence imaging showed increased drug accumulation in the brain and increased tumor penetration of Tf-ELE/CTX@BLIP in orthotopic glioma model nude mice. Findings from in vivo studies indicated that the antitumor effect of the Tf-ELE/CTX@BLIP led to increased survival time and decreased tumor volume in mice. The average tumor fluorescence intensity after intravenous administration of Tf-ELE/CTX@BLIP was 65.2, 12.5, 22.1, 6.6, 2.6, 1.5 times less compared with that of the control, CTX solution, ELE solution, ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP groups, respectively. Histopathological analysis showed that Tf-ELE/CTX@BLIP were less toxic compared with administration of the CTX solution.These findings indicate that the active-targeting biomimetic liposome, Tf-ELE/CTX@BLIP, is a promising nanoplatform for delivery of drugs to gliomas.The online version contains supplementary material available at 10.1186/s12951-021-01048-3. P-gp) occurs on the BBB, which further increases clearance of chemotherapy drugs \u2009=\u200920\u00a0\u03bcg/mL) and incubated for 2\u00a0h. In addition, RG2 cells were seeded into a 6-well plate (3\u2009\u00d7\u2009105 cells/well) and incubated with ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP and Tf-ELE/CTX@BLIP (Rho B\u2009=\u200920\u00a0\u03bcg/mL) for 2\u00a0h. The medium was removed and cells were fixed with 4% paraformaldehyde, stained with DAPI and washed thrice with PBS. The cellular uptake was determined by confocal laser scanning microscope imaging  and flow cytometry [Tf-ELE/CTX@BLIP was incubated with SPC-A-1 cells, A549 cells, MDA-MB-231 cells, LM-3 cells, U251 cells, C6 cells, and RG2 cells, separately. Cells were inoculated into 6-well plates  for 2\u00a0h. Effect on immune evasion was then determined by CLSM and flow cytometry .Cytotoxicity effects of ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP and Tf-ELE/CTX@BLIP on RG2 cells were determined using a CCK-8 kit. RG2 cells (3000\u00a0cells/well) were inoculated in 96-well plates and incubated for 24\u00a0h. After incubation, cells were exposed to ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP and Tf-ELE/CTX@BLIP (CTX concentrations ranging from 0.4 to 200\u00a0ng/mL) for 48\u00a0h. Non-treated cells were used as negative controls. Medium was evaluated as the blank control. Cytotoxicity was quantitatively determined by measuring the absorbance at 450\u00a0nm using a Spark multi-functional microporous plate testing platform .Furthermore, apoptotic cells were determined by FACS analysis . RG2 cells were incubated with ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP and Tf-ELE/CTX@BLIP and 50\u00a0ng/mL CTX for 48\u00a0h then treated with the apoptosis detection kit  for 10\u00a0min. Percentage of apoptotic cells was determined using a FACS Calibur System. Non-treated cells were used as the negative control .5\u00a0cells/well) and incubated for 24\u00a0h. Rhodamine 123  was preconditioned with ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP, Tf-ELE/CTX@BLIP (50\u00a0ng/mL) and verapamil (0.625\u00a0\u03bcg/mL) separately for 30\u00a0min then it was exposed to bEnd.3 for 2\u00a0h. The medium was removed and bEnd.3 cells were fixed with 4% paraformaldehyde for 30\u00a0min and washed thrice with PBS. Cells were then analyzed using fluorescence microscope and flow cytometry. Relative expression level of P-gp in bEnd.3 cells was determined by WB assay. bEnd3 cells in 6-well plates were conditioned with medium and verapamil, ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP, Tf-ELE/CTX@BLIP were added [bEnd.3 cells were cultured as described in the \u201cre added .Female nude mice  were obtained from Shanghai Slack Laboratory Animal Co. LTD. Mice were allowed to acclimatize at room temperature for 7\u00a0days prior to the performing animal studies. Animals were housed under the animal care facility and allowed free access to food/water. All animal experiments were approved by the animal ethics committee of Hangzhou Normal University .7 glioma-luc cells in medium were injected slowly into the brain right striatum of nude mice . The needle was maintained at the injection point for 1\u00a0min after injection and then pulled out slowly. Alcohol cotton swabs were used to disinfect the skin, the skin was sewn with a needle and thread. The mice were then placed back to the cage to wake up naturally. Rate of growth of intracranial tumor was monitored by magnetic resonance imaging (MRI) and fluorescence imaging.Glioma-luc cells were injected into the right striatum of nude mice to develop a orthotopic glioma-bearing model. Nude mice were anesthetized through administration of 10% chloral hydrate and the head was immobilized on a stoelting . Approximately 2.5\u2009\u00d7\u200910Cypate\u2009=\u20090.1\u00a0mg/mL) were injected into normal mice and glioma-bearing mice through the tail vein. Bioluminescence imaging was performed at fixed times  using a small animals in vivo 3D bioluminescence imaging system .The nude mice were then sacrificed, and brains, liver, heart, spleen, lung, and kidney were collected for quantitative biodistribution analysis and ex vivo bioluminescence imaging [Orthotopic glioma-bearing model was established by injecting RG2 cells into the brain striatum as described above. Cypate, Cypate@LIP, Cypate@BLIP, Tf-Cypate@LIP and Tf-Cypate@BLIP (CP-gp.Orthotopic glioma-bearing model nude mice were randomly assigned into seven groups (6 mice/group) as follows: (1) control ; (2) CTX solution; (3) ELE solution (25\u00a0mg/kg ELE); (4) ELE/CTX@LIP; (5) ELE/CTX@BLIP; (6) Tf-ELE/CTX@LIP; (7) Tf-ELE/CTX@BLIP, to explore the anti-tumor effect in vivo. Treatments were administered by intravenous injection through the tail at day 1, 3, 5, 7, 9 and 11. Group 2, 4, 5, 6, 7 was administered with 2.5\u00a0mg/kg (CTX) on the first time and 0.625\u00a0mg/kg for the subsequent 5 injections. Bioluminescence imaging was performed at day 1, 5, 10, and 15 to explore tumor growth. Body weight and survival time were recorded every 3\u00a0days. Nude mice were euthanized 2\u00a0h post-treatment and their brains were collected for H&E staining, TUNEL immunofluorescence staining and detection of glioma Glioma-bearing nude mice were assigned into seven groups and were administered with (1) control ; (2) CTX solution; (3) ELE solution; (4) ELE/CTX@LIP; (5) ELE/CTX@BLIP; (6) Tf-ELE/CTX@LIP; (7) Tf-ELE/CTX@BLIP, 6 times and every other day. Whole blood samples were collected from retro-orbital sinus of glioma-bearing nude mice 2\u00a0h post-treatment. A portion of the complete blood samples was used for complete blood count analysis. The supernatant (serum) of the portion of blood obtained after centrifugation  was used for determination of biochemical indexes of liver and kidney, including total bilirubin, blood urea nitrogen, uric acid, alanine transaminase, creatinine and aspartate transaminase level. Liver, heart, spleen, lung and kidney sections were stained with H&E to evaluate effects of free ELE, CTX solution and liposomes toxicity .t test for comparison between two groups and ANOVA for comparison among multiple groups. Statistical difference was defined as significant for *p\u2009<\u20090.05 and highly significant for **p\u2009<\u20090.01.GraphPad Prism 8.0.2.263 software  was used for statistical analysis. Data were expressed as mean\u2009\u00b1\u2009SD. Experimental data were analyzed using two-tailed Student Additional file 1: Table S1. Encapsulation efficiency stability of the drug in 4 liposomes at 3 months\u00a0(n = 3). Fig. S1. Diameter and \u03b6-potential of Tf-ELE/CTX@BLIP, Tf-ELE/CTX@LIP, ELE/CTX@BLIP and ELE/CTX@LIP. Fig. S2. 7-days stability of TF-ELE/CTX@BLIP diameter in different medium. Fig. S3. Flow cytometry analysis of RG2 glioma cells after incubation with Tf-ELE/CTX@BLIP, Tf-ELE/CTX@LIP, ELE/CTX@BLIP and ELE/CTX@LIP for 2 h. Rho B = 20 \u03bcg/mL. Fig. S4. CLSM images of RG2, U251 and C6 glioma cells treated with Tf-ELE/CTX@BLIP for 2 h. Scale bar = 50 \u03bcm. Fig. S5. Flow cytometry analysis of RAW264.7 cells treated with Tf-ELE/CTX@BLIP, Tf-ELE/CTX@LIP, ELE/CTX@BLIP and ELE/CTX@LIP for 2 h. Rho B = 20 \u03bcg/mL. Fig. S6. WB analysis of P-gp in bEnd.3 cells. Relative protein expression was calculated. Cells preconditioned with (1) control; (2) verapamil; (3) ELE/CTX@LIP; (4) ELE/CTX@BLIP; (5) Tf-ELE/CTX@LIP; (6) Tf-ELE/CTX@BLIP. . Fig. S7. Corresponding quantitative fluorescent analysis of brain, liver, heart, spleen, lung and kidney at 48 h post-injection in glioma-beard mice. *p < 0.05. Fig. S8. In vivo fluorescence imaging of Tf-Cypate@BLIP, Tf-Cypate@LIP, Cypate@BLIP and Cypate@LIP in normal mice. Cypate = 0.5 mg/kg. Fig. S9. Averaged fluorescent intensity of saline, free CTX, free ELE, ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP and Tf-ELE/CTX@BLIP in nude mice bearing orthotopic glioma brain within 15 days of treatments. Fig. S10. H&E staining of brain sections of orthotopic glioma-bearing mice in different formulation groups. Fig. S11. Biochemical parameter analysis after treated with saline, free CTX, free ELE, ELE/CTX@LIP, ELE/CTX@BLIP, Tf-ELE/CTX@LIP and Tf-ELE/CTX@BLIP. (A) BLI-T: total bilirubin, (B) BUN: blood urea nitrogen, (C) URIC: uric acid, (D) ALT: alanine transaminase, (E) CRE: creatinine, (F) AST: aspartate transaminase."}
+{"text": "Compared with WT mice, S\u2212/\u2212tat1 had reduced neutrophils, Th1, and Th17 cell infiltration. To evaluate corticosteroid sensitivity, mice were treated with either vehicle, 1 or 3 mg/kg fluticasone propionate (FP). Corticosteroids significantly reduced eosinophil infiltration and cytokine levels in both c-di-GMP + MA-challenged WT and \u2212/\u2212Stat1 mice. However, histological and functional analyses show that corticosteroids did not reduce airway inflammation, epithelial mucous cell abundance, airway smooth muscle mass, and AHR in c-di-GMP + MA-challenged WT or \u2212/\u2212Stat1 mice. Collectively, our data suggest that increased Th1 inflammation is associated with a decrease in corticosteroid sensitivity. However, increased airway pathology and AHR persist in the absence of STAT1 indicate corticosteroid insensitivity in structural airway cells is a STAT1 independent process.Corticosteroid insensitivity in asthma limits the ability to effectively manage severe asthma, which is characterized by persistent airway inflammation, airway hyperresponsiveness (AHR), and airflow obstruction despite corticosteroid treatment. Recent reports indicate that corticosteroid insensitivity is associated with increased interferon-\u03b3 (IFN-\u03b3) levels and T-helper (Th) 1 lymphocyte infiltration in severe asthma. Signal transducer and activator of transcription 1 (STAT1) activation by IFN-\u03b3 is a key signaling pathway in Th1 inflammation; however, its role in the context of severe allergic airway inflammation and corticosteroid sensitivity remains unclear. In this study, we challenged wild-type (WT) and  Corticosteroids have broad anti-inflammatory effects that contribute to the effective management of asthma symptoms and exacerbations. In contrast to the corticosteroid-sensitive mild asthma, people with severe asthma exhibit greater airway inflammation, remodeling, airway hyperresponsiveness (AHR), and more frequent exacerbations despite treatment with higher doses of corticosteroids , 2. Alth+ T lymphocytes in bronchoalveolar lavage samples  levels . Studies samples , 9. Alth samples . These o samples , 11\u201313.  samples , 11, imp\u2212Stat1/\u2212 mice exhibit reduced Th1 responses and IFN-\u03b3 production  is a transcription factor that is activated by IFN-\u03b3 and mediates proinflammatory responses in immune and airway structural cells . Given toduction , 16. Preoduction . Given t\u2212Stat1/\u2212 mice using a chronic mixed allergen (MA) model augmented with c-di-GMP. We hypothesized that ablation of STAT1 would disrupt Th1 inflammation and increase corticosteroid sensitivity in chronic severe allergic airway inflammation. However, our data demonstrate reduced corticosteroid sensitivity in both WT and \u2212Stat1/\u2212 mice. Examination of immunological, functional, and structural aspects of chronic allergic airway inflammation shows that persistent airway remodeling remains increased in \u2212Stat1/\u2212 mice despite treatment with corticosteroids. Our studies demonstrate that corticosteroid insensitivity remains in the absence of STAT1 signaling during severe allergic airway inflammation.In this study, we examined corticosteroid sensitivity in wild-type (WT) and \u2212Stat1/\u2212 (Stock No. 012606) mice were procured from Jackson Laboratory . Mice were maintained in Research Building III at the Abigail Wexner Research Institute and provided food and water ad libitum. Adult male and female mice were mated to generate pup litters that were randomly assigned a treatment group. Newborn male and female mice were intranasally (in) sensitized and challenged with PBS or mixed allergens (MA), which consists of 10 \u00b5g Alternaria alternata , 10 \u00b5g Aspergillus fumigatus , 10 \u00b5g Dermatophagoides pteronyssinus , and 10 \u00b5g ovalbumin (OVA) plus 0.5 \u00b5g c-di-GMP , three times per week for 7 wk. During the last 2 wk, mice were intraperitoneally (ip) injected with vehicle (PBS containing 0.015% DMSO) or 1 mg/kg or 3 mg/kg FP  on the same day as MA challenge. Necropsy was performed \u223c24 h after last allergen challenge  and hallenge A. Mice wg for 5 min. Supernatant was collected and stored at \u221280\u00b0C for further analyses. Cellular pellets were resuspended in PBS for total and differential cell count analysis. Total counts were performed using a hemocytometer. Cytospins were generated on microscope slides and differentially stained using a modified Wright-Giemsa Stain . Cells were imaged on a bright-field microscope and counted by a blinded investigator until a total of 200 cells had been counted.Bronchoalveolar lavage fluid (BAL) was harvested as previously described . BrieflyOnce anesthetized, the trachea was exposed and cannulated with a 19-gauge blunt tip cannula. The mouse was placed on a 37\u00b0C heating pad and attached to Y-tubing on the Flexivent . A series of measurements including Snapshot  maneuvers were performed following nebulization with PBS, 6.25, 12.5, 25, and 50 mg/mL methacholine . Airway hyperresponsiveness is reported as total resistance in response to methacholine.Lung tissue was homogenized in RIPA buffer, processed, and protein concentration measured using Bradford assay. Each well in custom Meso scale U-Plex Multiplex ELISA plates  was coated with antibodies against IL-4, IL-17A, and IFN-\u03b3. Analytes were measured in whole lung homogenates following manufacturer instructions. IL-13 levels in BAL were measured using ELISA  according to the manufacturer\u2019s instructions.Samples were quantified by BCA assay (Pierce) and 25 \u00b5g of protein was analyzed per sample. Samples were separated on a 14% SDS-PAGE gel and transferred to nitrocellulose membrane (Bio-Rad). Membranes were blocked for 1 h at room temperature in 1% milk/TBS-T and then incubated in primary antibody overnight at 4\u00b0C. Primary antibodies were purchased from Cell Signaling : monoclonal rabbit anti-Stat1 antibody at 1:1,000 dilution, Cat. No. 14994S; polyclonal rabbit antiphospho-Stat1 (Ser727) antibody at 1:500 dilution, Cat. No. 9177S; monoclonal rabbit anti-Stat6 at 1:500, Cat. No. 5397S; and monoclonal rabbit anti-phospho-Stat6 (Tyr641) at 1:500, Cat. No. 56554S. The following day, samples were washed in Tris-buffered saline-Tween 20 (TBST), incubated with secondary antibodies  for 1 h at room temperature, washed with TBST, and imaged on an Odyssey Clx (LI-COR). Protein quantification was performed using ImageStudio Software (LI-COR).2O. Lungs were processed, paraffin embedded, cut into 6 \u00b5m sections, and stained with hematoxylin and eosin . H&E slides were analyzed and scored for inflammation  and bronchial-associated lymphoid tissues (BALTs) were quantified by a blinded investigator. Inflammation scores are based on the degree of immune cell infiltration/aggregation around peribronchiolar and perivascular spaces. To quantify BALTs, compact lymphoid nodules were counted in proximal and distal regions. To quantify abundance of mucous cells in the airway epithelium, left lung lobe sections were stained with Alcian Blue-Periodic Acid Schiff (AB-PAS) and photomicrographs of four different airways were taken at \u00d7100 using an Olympus BX-40 light microscope and digital camera . PAS-positive and -negative cells were quantified by a blinded investigator using ImageJ (NIH) and reported as a percentage of total airway epithelial cells counted.Left lung lobes were inflated with 10% neutral-buffered formalin at 25 cm HFormalin-fixed, paraffin embedded left lung lobe sections (6 \u00b5m) were used for immunohistochemical staining for \u03b1-smooth muscle actin (\u03b1-SMA). Sections were deparaffinized with xylene (2 \u00d7 5 min each) and rehydrated with graded ethanol . Antigen retrieval was performed with 10 mM sodium citrate at 100\u00b0C for 1 h. The slides were blocked for 1 h in TBS containing 4% goat serum and 0.04% Triton X-100. After washing, slides were incubated overnight in monoclonal mouse anti-\u03b1-SMA at 1:100 dilution . After incubation, slides were washed and incubated with anti-mouse-FITC secondary antibody  at 1:1,000 dilution. Negative control slides followed the same protocol but without addition of a primary antibody. Slides were counterstained with DAPI, and images taken at \u00d7100 using Lionheart . Smooth muscle actin area was analyzed using ImageJ (NIH). Data were normalized to length of airway basement membrane to report as airway smooth muscle (ASM) mass per \u00b5m basement membrane.12\u201318 primers and Superscript IV (Thermo Fisher Scientific Applied Biosystems). Quantitative real-time PCR with TaqMan using Muc5ac (Mm0127618_m1), Muc5b (Mm00466391_m1), and GAPDH (Mm99999915_g1) primer sets was performed (Life Technologies) according to the manufacturer\u2019s instructions. Samples were run on a QuantStudio 3 96-well Real-Time PCR system (Thermo Fisher Scientific Applied Biosystems). Results were analyzed using the comparative Ct method for TaqMan assays.RNA (300\u20131000 ng) from profiled samples was cDNA transcribed using Oligo dTg for 10 min, and resuspended in 10 mL of DMEM medium. Cells were counted using a hemocytometer. For Th stimulation, two million cells were incubated in DMEM medium containing leukocyte activation cocktail with Golgi plug , protein transported inhibitor containing Monensin , and 10% FBS for 37\u00b0C for 4 h. Cells were collected and permeabilized/fixed using fixation/permeabilization buffer and 1\u00d7 permeabilization/wash buffer  and resuspended in 200 \u03bcL of PBS. Single cell suspensions were used in flow cytometry analysis.Lungs were perfused with 5 mL of PBS through right ventricle of the heart, and then placed in C-tubes  and digested for 30 min in 432 U/mL Collagenase Type IV  and 64 U/mL DNAase I  in DMEM medium. Lung tissue was homogenized using Gentlemacs octo dissociator (Miltenyi Biotec), strained through 70 \u03bcm nylon cell strainer, centrifuged at 300 1 isotypes were used. All antibodies were purchased from BioLegend. Data were acquired using a BD LSRII flow cytometer  and analyzed using FlowJo software (v. 10.7.1).Single cell suspensions were stained with various fluorochrome conjugated anti-mouse monoclonal antibodies for cellular phenotyping as follows: FITC-conjugated anti-CD45 (30-F11), BV421-conjugated anti-CD3 (17A2), PE-conjugated anti-CD4 (A161A1), Alexa 700-conjugated anti-IFN\u03b3 (XMG1.2), APC-conjugated anti-IL-4 (11B11), and APC-conjugated anti-IL-17A (TC11-18H10.1). To control for intracellular cytokine staining, APC (RTK2071) and Alexa fluor 700 (RTK2071) IgGP < 0.05.Data were analyzed by performing two-way ANOVA with Bonferroni post hoc analysis for multiple comparisons. Data were analyzed and graphed using GraphPad Prism 9 Software . Values are presented a means \u00b1 standard error and significant differences indicated by \u2212Stat1/\u2212 mice challenged with c-di-GMP + MA exhibited increased total BAL immune cells compared with PBS-challenged WT mice  and  WT mice B. BAL to WT mice C. Differ WT mice D. In con\u2212/\u2212 mice E. Treatm WT mice B, wherea\u2212/\u2212 mice D and E. \u2212/\u2212 mice C.+CD4+ T cell numbers were significantly increased in c-di-GMP + MA-challenged WT and \u2212Stat1/\u2212  and IL-4 and IL-13 expression. Th2 lymphocytes were increased in c-di-GMP + MA-challenged WT and \u2212Stat1/\u2212 mice and were not reduced with 1 or 3 mg/kg FP treatment  cells and IL-17A levels were significantly higher in c-di-GMP + MA-challenged WT and \u2212Stat1/\u2212 mice  nodules quantified. Compared with PBS challenge, c-di-GMP + MA challenge significantly increased airway inflammation in WT and \u2212/\u2212 mice A and B. \u2212/\u2212 mice B. To anagenotype C. In regcontrols D. A signmg/kg FP C.\u2212Stat1/\u2212mice exhibited significant increases in mucous cells compared with PBS-challenged mice  mass in lung sections stained for \u03b1-smooth muscle actin. C-di-GMP + MA-challenged WT and mg/kg FP .\u2212Stat1/\u2212 mice showed significantly increased AHR compared with PBS-challenged mice. Treatment with 1 mg or 3 mg/kg fluticasone propionate did not significantly reduce AHR in WT and \u2212Stat1/\u2212 mice  was measured using forced oscillation maneuvers following administration of methacholine (0\u201350 mg/mL). C-di-GMP + MA-challenged WT and \u2212/\u2212 mice . To analcontrols .\u2212Stat1/\u2212 mice that exhibit dysfunctional Th1 signaling  and c-di-GMP exhibited reduced AHR even in the absence of corticosteroids, implicating Th1 inflammation in increased AHR . Although we observed that corticosteroids significantly reduced IL-4 and IL-13 expression levels in \u2212Stat1/\u2212 mice, their levels did not return to baseline. IL-4 and IL-13 at relatively low levels have been shown to induce mucus production, AHR, and remodeling  expression and mucus production are associated with increased Th2-associated inflammation  model, Ifn\u03b3\u2212/\u2212 and \u2212Irf5/\u2212 mice exhibited evidence of increased Th2 inflammation with greater eosinophil infiltration than allergen-challenged WT mice , and persistent airway remodeling could be contributing factors in the lack of reduced airway inflammation and corticosteroid sensitivity in \u2212Stat1/\u2212 mice.Our findings in  WT mice . However\u2212Stat1/\u2212 mice, persistent airway inflammation and corticosteroid insensitivity may involve enhanced Th2 inflammation. Negative regulation of Th2 inflammatory responses by Th1 inflammation is an established and important immune regulatory mechanism , R01 HL155095 (to R. D. Britt), R01 AI121405 (to M. Guerau-de-Arellano), R03 AI151769 (to M. Guerau-de-Arellano), and startup funds from the Abigail Wexner Research Institute at Nationwide Children\u2019s Hospital.No conflicts of interest, financial or otherwise, are declared by the authors.B.W.L., M.H.G., and R.D.B.J. conceived and designed research; B.W.L., D.J., S.A.A., J.W., M.G., S.G., E.C., A.V.B., and R.D.B.J. performed experiments; B.W.L., D.J., S.A.A., J.W., M.G., S.G., E.C., A.V.B., M.G.-d.-A., and R.D.B.J. analyzed data; B.W.L., M.G.-d.-A., M.H.G., and R.D.B.J. interpreted results of experiments; B.W.L. and R.D.B.J. prepared figures; B.W.L., M.G.-d.-A., M.H.G., and R.D.B.J. drafted manuscript; B.W.L., D.J., S.A.A., J.W., M.G., S.G., E.C., A.V.B., M.G.-d.-A., M.H.G., and R.D.B.J. edited and revised manuscript; B.W.L., D.J., S. A.A., J.W., M.G., S.G., E.C., A.V.B., M.G.-d.-A., M.H.G., and R.D.B.J. approved final version of manuscript."}
+{"text": "Chalara sp. is able to biotransform the epigenetic modifier vorinostat to form unique, aniline-containing polyketides named chalanilines. Here, we sought to expand the chemical diversity of chalaniline A-type molecules by changing the aniline moiety in the precursor vorinostat. In total, twenty-three different vorinostat analogs were prepared via two-step synthesis, and nineteen were incorporated by the fungus into polyketides. The highest yielding substrates were selected for large-scale precursor-directed biosynthesis and five novel compounds, including two fluorinated chalanilines, were isolated, purified, and structurally characterized. Structure elucidation relied on 1D and 2D NMR techniques and was supported by low- and high-resolution mass spectrometry. All compounds were tested for their bioactivity but were not active in antimicrobial or cell viability assays. Aminofulvene-containing natural products are rare, and this high-yielding, precursor-directed process allows for the diversification of this class of compounds.The plant endophyte  Tolypocladium sp. which results from highly reactive intermediates that can detoxify various synthetic and naturally derived antifungals via nucleophilic substitutions +  and an m/z value of 416.0904 [M + Na]+ , resulting in a molecular formula of C22H16FNO5. The UV spectrum showed maxima at 380, 308, and 244 nm, representing the chalaniline A-type backbone. The 1H NMR spectrum exhibited a broad hydroxyl peak (\u03b4H 13.75), an N-H resonance (\u03b4H 11.84) with a large coupling constant (14.5 Hz) to one methine (\u03b4H 8.74), seven aromatic/olefinic hydrogens, one methoxy, and one methyl group , 2J (26.2 and 21.4 Hz), and 3J (9.4 Hz) C-F coupling constants in the carbon spectrum, and the proton spectrum revealed two 3J (10.7 and 8.0 Hz) H-F coupling constants , as well as the correlation between H-11 and C-2\u2032 supported the structural assignment. The placement of the fluorine at C-3\u2032 was confirmed by C-F coupling constant analysis  was isolated as a yellow amorphous solid and the HRESIMS provided an m/z value of 394.1088 [M + H] +  and m/z value of 416.0909 [M + Na]+ for the sodium adduct . The 1H NMR spectrum of 2 was very similar to the spectrum of 1, the only differences can be found in the number of aromatic signals and the coupling and integration pattern for the fluoro-benzene moiety. Two sets of aromatic signals, with each an integration of ~2H, supported the symmetric, para-substituted fluoro-aniline incorporation , 2J C-F coupling of 23.0 Hz for C-3\u2032/5\u2032, 3J C-F coupling of 8.3 Hz for C 2\u2032/6\u2032, and 4J C-F coupling (2.6 Hz) for C-1\u2032  and the 4-fluoro anilino ring C-2\u2032/6\u2032, and also from the H-11 to C-2\u2032  was isolated as an amorphous yellow solid. The HRESIMS gave an m/z value of 406.1284 [M + H] +  and m/z value of 428.1102 [M + Na]+ for the sodium adduct . The 1H NMR spectrum of 3 showed a similar pattern of signals as previously reported for chalaniline A  + , indicative of a regioisomer of 3. The para-substitution in the 4-methoxy aniline moiety became evident in the proton NMR with two doublets each with an integration of ~2H , as well as the correlation between bridging methine (H-11) to the fulvene  assisted in the structure assembly  was isolated as a yellow amorphous solid and the HRESIMS gave an m/z value of 426.1341 [M + H]+ , and an m/z value of 448.1160 [M + Na]+ for the sodium adduct  supporting a molecular formula of C26H19NO5. The 1H and 13C NMR spectra were similar to chalaniline A with additional signals for the naphthyl moiety instead of the phenyl in the aromatic region  and melanoma (SK-MEL-5) cancer cell models by measuring the reduction of the tetrazolium salt MTT -2,5-diphenyltetrazolium bromide) by metabolically active cells + , m/z 416.0904 [M + Na]+ 3-Fluoro chalaniline A (2): yellow amorphous solid; IR (ATR): \u03bdmax = 3430, 2925, 2853, 1712, 1651, 1589, 1508, 1466, 1365, 1209, 1098 cm\u22121; UV (MeCN) \u03bbmax: 380, 308, 244 nm; 13C NMR  and 1H NMR  see m/z 394.1088 [M + H]+ , m/z 416.0909 [M + Na]+, 4-Fluoro chalaniline A (3): yellow amorphous solid; IR (ATR): \u03bdmax = 3290, 2924, 2850, 1703, 1647, 1601, 1510, 1470, 1252, 1190, 1050, 840, 735 cm\u22121; UV (MeCN) \u03bbmax = 383, 310, 246 nm; 13C NMR  and 1H NMR  see m/z 406.1284 [M + H]+ , m/z 428.1102 [M + Na]+  3-Methoxy chalaniline A (4): yellow amorphous solid; IR (ATR): \u03bdmax = 3290, 2924, 1703, 1647, 1510, 1490, 1252, 1050, 820, 745 cm\u22121; UV (MeCN) \u03bbmax: 382, 308, 244 nm; 13C NMR  and 1H NMR  see m/z 406.1286 [M + H]+, 4-Methoxy chalaniline A (5): yellow amorphous solid; IR (ATR): \u03bdmax = 3410, 2926, 2852, 1737, 1647, 1614, 1465,1372, 1307, 1207, 1098, 830, 767 cm\u22121; UV (MeCN) \u03bbmax: 374, 305, 242, 214 nm; 13C NMR  and 1H NMR  see m/z 426.1341 [M + H]+ , m/z 448.1160 [M + Na]+ Naphthyl chalaniline A (Staphylococcus aureus (ATCC 25923), methicillin-resistant Staphylococcus aureus (ATCC BAA-41), multidrug-resistant Staphylococcus aureus (ATCC BAA-44), Pseudomonas aeruginosa (ATCC 15442), Candida albicans (ATCC 90027), Candida krusei (ATCC 34135), and Mycobacterium smegmatis (ATCC 14468) in microbroth assays performed following an established protocol [Antimicrobial Assays. Extracts and fractions were tested for inhibitory activity against protocol ,31. FracCell proliferation assay. Cytotoxic activities of extracts and pure compounds were evaluated against colon (HCT-116) and melanoma (SK-MEL-5) cancer models by measuring the reduction of the tetrazolium salt MTT -2, 5-diphenyltetrazolium bromide) by metabolically active cells following standard procedures ,33."}
+{"text": "P. aeruginosa from bloodstream infections (BSI) to those from other infection types.Ceftolozane/tazobactam (C/T), an antipseudomonal cephalosporin combined with a \u03b2-lactamase inhibitor, was approved for treatment of complicated urinary tract (cUTI) and intraabdominal infections (cIAI), and hospital-acquired/ventilator-associated bacterial pneumonia (HAP/VAP). Imipenem/relebactam (IMI/REL) is a combination of imipenem/cilastatin with relebactam, an inhibitor of class A and C \u03b2-lactamases. IMI/REL was approved for HAP/VAP and for infections due to aerobic gram-negative organisms in adults with limited treatment options . We compared the activity of C/T and IMI/REL against Pa isolates were collected. MICs were determined using CLSI broth microdilution and breakpoints.As part of the SMART program, 24 hospitals in the US and 8 in Canada each collected up to 250 consecutive gram-negative isolates per year in 2018-2019 from patients with BSI, lower respiratory tract infections (LRTI), IAI, and UTI. A total of 2351 Pa isolates from BSI tended to show higher susceptibility than IAI, UTI, and especially LRTI isolates (Table). Susceptibility to the tested comparator \u03b2-lactams was 11-12 percentage points lower among LRTI than BSI isolates, while C/T and IMI/REL susceptibility was only 2-5% lower. Even among BSI isolates, the comparator \u03b2-lactams were active against only 75-88% of isolates, while C/T and IMI/REL were active against >95%. Only amikacin showed higher activity. Analyzing coverage by either C/T or IMI/REL, 98.7% of Pa isolates from BSI were susceptible to one or both agents. C/T and IMI/REL maintained activity against 89% and 69% of meropenem-nonsusceptible (MEM-NS) Pa isolates from BSI (n=36), respectively, and 87% and 76% of piperacillin/tazobactam (P/T)-NS Pa (n=38).Results TablePa susceptibility to commonly used \u03b2-lactams like MEM and P/T was < 90%, 7-23% lower than C/T and IMI/REL. Given the desirability of \u03b2-lactams among clinicians and the >98% coverage by either C/T or IMI/REL of Pa isolates from BSI, both agents represent important options in the treatment of patients with BSI.Even among BSI isolates, which were generally more susceptible than those from other infection types, Sibylle Lob, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Meredith Hackel, PhD MPH, IHMA (Employee)Pfizer, Inc. (Independent Contractor) C. Andrew DeRyke, PharmD, Merck & Co., Inc.  Kelly Harris, PharmD, BCPS, Merck & Co. Inc (Employee) Katherine Young, MS, Merck (Employee) Mary Motyl, PhD, Merck & Co., Inc.  Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "Lactococcus lactis may represent a potential approach for IBD therapy.Inflammatory bowel diseases (IBDs) are generally characterized by persistent abdominal pain and diarrhea caused by chronic inflammation in the intestine. Cathelicidins are antimicrobial peptides with pleiotropic roles in anti-infection, wound healing, and immune modulation. However, the sensitivity to the acidic environment and short half-life of cathelicidins limit their application in IBD treatment. Recombinant cathelicidin-related antimicrobial peptide (CRAMP)-producing L. lactis NZ9000 and explore the role and mechanism of recombinant L. lactis NZ9000 expressing CRAMP in colitis.The aim of this study was to develop recombinant CRAMP-producing L. lactis NZ9000 with different plasmids pMG36e (L.L-pMU45CR) or pNZ8148 (L.L-pNU45CR), which use a Usp45 secretion signal to drive the secretion of CRAMP. Bacterial suspensions were orally supplemented to mice with a syringe for 4 days after dextran sodium sulfate (DSS) treatment. Body weight change, disease active score, colon length, and colonic histology were determined. The expression of tight junction  and cytokines  in colon was performed by qPCR. The expression of p-ERK, p-p38, and p-p65 was determined by Western blot analysis.We constructed two strains of CRAMP-producing L. lactis NZ9000 strains protected against colitis, as shown by reduced weight loss and disease activity score, improved colon shortening, and histopathological injury. In addition, CRAMP-producing L. lactis NZ9000 restored gut barrier by upregulating ZO-1, ZO-2, and occludin. Moreover, CRAMP-producing L. lactis NZ9000 regulated the colonic cytokines profile with reduced IL-6, IL-1\u03b2, and TNF-\u03b1 production, and increased IL-10 production. By further analysis, we found that CRAMP-producing L. lactis NZ9000 reduced the expression of p-p38 and p-p65.Both CRAMP-producing L. lactis NZ9000 attenuated dextran sulfate sodium-induced colitis by colonic colonization and inhibiting p38/NF-\u03baB signaling. Orally administered recombinant CRAMP-secreting L. lactis NZ9000 represents a potential strategy for colitis therapy.Together, our data suggested that CRAMP-secreting  Lactococcus lactis NZ9000 were constructed with different plasmids pMG36e (L.L-pMU45CR) or pNZ8148 (L.L-pNU45CR), which use a Usp45 secretion signal to drive the secretion of CRAMP.Two strains of recombinant CRAMP-producing L. lactis NZ9000 strains protected mice from colitis via suppressed activation of p-p38/NF-\u03baB signaling, thus resulting in a restored cytokines profile and an improved gut barrier integrity.CRAMP-producing L. lactis NZ9000 represents a novel intervention strategy for colitis treatment.CRAMP-producing Inflammatory bowel diseases (IBDs) are inflammatory conditions in the intestine, which generally include ulcerative colitis (UC) and Crohn\u2019s disease (CD) . Both UCCathelicidins are antimicrobial peptides found in humans and mice, which includes LL-37 and cathelicidin-related antimicrobial peptide (CRAMP), that are homologous in nature, . These hLactococcus lactis (L. Lactis) is widely used in food fermentation and oral delivery of therapeutic proteins (Lactococcus lactis NZ9000 (L. Lactis NZ9000) plays a key role in industrial fermentation  or pNZ8148 (L.L pNU45CR) plasmids, respectively. We also evaluated the role and mechanism of CRAMP-secreting L. lactis NZ9000 in experimental colitis.In this study, we constructed two recombinant Lactococcus lactis NZ9000  was grown in M17 medium  containing 0.5% (w/v) glucose  at 30\u00b0C. CRAMP gene (Lactococcus lactis NZ9000 was transformed with pMG36e-Usp45-CRAMP (L.L-pMU45CR) and pNZ8148-Usp45-CRAMP (L.L-\u200bpNU45CR) by electroporation (Lactococcus lactis NZ9000 was transformed with empty pMG36e (L.L-pMVectro) and empty pNZ8148 (L.L-pNVector) as controls. Nisin  was used for 4 h to induce gene expression in L.L-pNU45CR and L.L-pNVector as controls (AMP gene  containiAMP gene  was syntAMP gene  (pMG36e-AMP gene  (pNZ8148poration . Lactococontrols . Bacterin = 5) as follows: [1] control (without DSS), [2] 3% DSS + sterilized water, [3] 3% DSS + L.L-pMVector, [4] 3% DSS + L.L-pNVector, [5] 3% DSS + L.L-pMU45CR, and [6] 3% DSS + L.L- pNU45CR. After 7 days of DSS treatment, bacterial suspensions (1010 CFU) were orally administrated with syringe once daily for 4 days. Bacterial suspensions were prepared as described below. The bacterial cells were centrifuged at 3,000 g at 4\u00b0C for 20 min and washed twice with PBS. These bacterial cells were resuspended at concentrations of 5 \u00d7 1010 CFU/mL based on optical density (OD600). Mice were gavaged with 200 \u00b5L of bacterial suspension (1010 CFU) with syringe. To check the number of CFUs, plate counts were performed on M17 agar supplemented with 0.5% glucose at 30\u00b0C without agitation. After 24 h, the plates were observed.Male C57BL/6 mice (7\u20138 weeks old) were purchased from Su Pu Si Biotechnology Co. Ltd  and were randomly divided into six groups  according to the manufacturer\u2019s instructions.Colon tissues were fixed and embedded in paraffin via standard methods , and sec-\u2206\u2206ct method was used for calculation and normalized to \u03b2-actin. All primers were synthesized by Thermo Fisher Scientific . Primers were designed with Primer 5 software . To check the specificity of primers, blast program  was used for colonic RNA isolation. SuperRT cDNA synthesis kit  was used for reverse transcription. SYBR Green  was used for quantitation. The 2 program  was usedL. Lactis cultures at a given optical density of 600 nm (OD600) were harvested by centrifugation at 3,000\u00b4g for 20 min at 4\u00b0C. The equivalent of 1 mL of 1 OD600 unit of culture (cell or supernatant) was concentrated in a 100 \u03bcL final volume as described below, and 10 \u03bcL was loaded for SDS-PAGE. Supernatants were precipitated by the addition of 10% trichloroacetic acid, harvesting by centrifugation at 10,000\u2005\u00d7\u2005g at 4\u00b0C. The resulting pellet was dissolved in a 1:20 volume of 50 mM NaOH. Cell pellets were resuspended in 70 \u03bcL of TES containing lysozyme (1 mg/mL). After 30 min of incubation at 37\u00b0C, cells were lysed with 30 \u03bcL of 20% SDS. Equal volumes of 2\u00d7 loading buffer were added to all samples. RIPA buffer with phosphatase and protease inhibitors was purchased from Songon Biotech  and used in the preparation of colon samples. Electrophoresis and transfer were performed as described before  and statistically analyzed by GraphPad Prism software  using one-way analysis of variance followed by Tukey\u2019s post-hoc test. L.L-pMU45CR or L.L-pNU45CR. As shown in L.L-pMU45CR and L.L-pNU45CR expressed CRAMP, and the levels of CRAMP were higher in supernatants than in bacterial cells  to confirm whether CRAMP with secretion signal peptides (Usp45 + CRAMP) were expressed in al cells . CollectL. lactis NZ9000 on colitis, we treated mice with recombinant L. lactis NZ9000 for 4 days after dextran sodium sulfate (DSS)-induced colitis. We found that both L.L-pMU45CR and L.L-pNU45CR attenuated colitis in DSS-treated mice, as shown by reduced weight change  L. lactis, which transit with diet, could survival through acidic conditions in gastric juice (L. lactis NZ9000 is able to colonize the colon and has been used to express bioactive molecules in the treatment of colitis, such as theme oxygenase-1 and insulin-like growth factor I (L. lactis are varied due to the characteristics of heterologous proteins. Usp45 is a signal peptide widely applied in driving protein secretion (L. lactis NZ9000 with Usp45 signal peptides.ic juice . Recombifactor I , 19. Theecretion . We achiL. Lactis-treated mice. However, the recombinant CRAMP producing L. lactis may not have a direct effect on serum CRAMP, because bioactive peptides were not able to cross the gut wall intact, except dipeptides and tripeptides (Cathelicidins have multifunctional roles, such as anti-microbial, anti-inflammation, anti-apoptosis activities, and wound healing. Serum LL-37 levels are positively correlated with recovery in IBD patients . We obsepeptides , 39. Conpeptides , 41.L. lactis increased the expression of IL-10. Similarly, cathelicidins increased the expression of IL-10 in human mononuclear cells (L.L-pMU45CR and L.L-pNU45CR reduced IL-6, TNF-\u03b1, and IL-1\u03b2 expression by suppressing the expression of p-p38 and p-p65. These results revealed that recombinant CRAMP-producing L. lactis regulates proinflammatory cytokine expression by p-p38/NF-\u03baB p-p65 signaling.Cytokines exert major impacts on intestinal inflammation and related clinical symptoms in IBD. The unbalanced cytokines profile between proinflammatory and regulatory cytokines promotes mucosal inflammation. IL-10 regulates intestinal homeostasis, and its deficiency leads to spontaneous colitis in mice . We founar cells , 44. Bloar cells , 45, 46.Escherichia coli is increased in IBD patients and can disrupt tight junctions (L. lactis restored the expression of tight junctions by regulating the cytokines profile in colitis, although it is unclear whether recombinant CRAMP-producing L. lactis regulates adherent-invasive E. coli.Epithelial tight junctions are crucial in regulating intestinal barrier and permeability. Disruption of intestinal barrier results in the transfer of intestinal bacteria and antigens into submucosa, thus subsequently leading to inflammatory response, such as transcription factor activation and immune cell infiltration. Tight junctions are regulated by intestinal microbes and cytokines in IBD. Adherent-invasive unctions . Severalunctions . Previouunctions  and epidunctions . TogetheL. lactis NZ9000 strains protected mice from colitis via suppressed activation of p-p38/NF-\u03baB p-p65 signaling, thus resulting in a restored cytokines profile and improved gut barrier integrity (L. lactis NZ9000 may represent a potential strategy for IBD therapy.Our data revealed that two CRAMP-secreting ntegrity . Our dat"}
+{"text": "Correction to: BMC Public Health 22, 354 (2022)https://doi.org/10.1186/s12889-022-12776-yThe original publication of this article contained an incorrect Fig. 1. The incorrect and correct Fig. 1 are included in this correction article .The original article has been updated.Fig. 1 Incorrect version of figure 1 as originally publishedFig. 2 Correct version of figure 1 as corrected"}
+{"text": "We investigated the infectivity of 128 severe acute respiratory disease coronavirus 2\u2013associated deaths and evaluated predictive values of standard diagnostic procedures. Maintained infectivity (20%) did not correlate with viral RNA loads but correlated well with anti-S antibody levels. Sensitivity >90% for antigen-detecting rapid diagnostic tests supports their usefulness for assessment. Deaths associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have raised concerns that contact with the corpses of deceased persons might pose a risk for transmitting infection   , panel Apies/mL) , panel B7 copies/mL, IQR 3.7 \u00d7 104\u20133.3 \u00d7 108) among culture-positive corpses did not differ significantly from PMI  and RNA loads  among culture-negative corpses (4 copies/mL), in contrast with previous findings among living patients , indicative of potentially decreased infectivity.Virus isolation proved infectivity was maintained in 26/128 (20%) corpses . PMI  (We confirmed seroconversion in 18/44 (41%) blood samples, 15/43 (35%) anti-nucleocapsid positive and 17/44 (39%) anti-spike positive  , panel C = 0.23) , panel Chttps://www.abbott.com), sensitivity was 80.3% (95% CI 72.3%\u201386.4%) and specificity 100.0% (95% CI 95.0%\u2013100.0%); for the SARS-CoV-2 Rapid Antigen Test (Roche https://www.roche.com), sensitivity was 86.4% (95% CI 79.1%\u201391.9%) and specificity 98.6% (95% CI 93.0%\u2013100.0%); and for the SARS-CoV-2 Antigen Rapid Test (MEDsan https://www.medsan.eu), sensitivity was 84.1% (95% CI 76.6%\u201390.0%) and specificity 95.8% (95% CI 88.0%\u201399.0%) .Antigen-detecting rapid diagnostic tests (Ag-RDTs) are considered adequate alternative swift diagnostic tools in living patients (6 RNA copies/mL (n = 74) revealed 100% (95% CI 95.1%\u2013100.0%) sensitivity in Abbott (n = 74) and Roche and MEDsan (n = 73 each) assays. In contrast, neither PMI (p = 0.34) nor putrefactive changes (p = 0.90) were predictive for testing positive in Ag-RDTs . Ag-RDT sensitivity in infectious corpses was 92.3% (95% CI 74.9%\u201399.1%) for Abbott, 96.2% (95% CI 80.4%\u201399.9%) for Roche, and 96.2% (95% CI 80.4%\u201399.9%) for MEDsan. We detected 2 SARS-CoV-2 variants of concern despite relatively low viral RNA loads (4.83 log10); the 2 samples tested positive by Abbott and Roche but were missed by MEDsan.We found SARS-CoV-2 RNA load correlated with Ag-RDT positivity in univariate and multivariate analyses (p<0.001), thereby confirming their predictive value (The first limitation of our study is that blood was not available from all corpses, and the serologic assays and Ag-RDTs used are not approved for cadaveric samples. Furthermore, because of a shortage of reagents and supplies, we had to use different tests to quantify RNA, and slight deviations cannot be ruled out.In summary, we show that cadavers from SARS-CoV-2\u2013associated deaths remain infectious long after death in a considerable proportion of cases. Postmortem infectivity does not correlate with PMI or viral RNA load but correlates with the absence of virus-specific antibodies. Ag-RDTs performed well, enabling rapid on-site detection. Because previous studies among living patients indicate that Ag-RDTs reliably detect all SARS-CoV-2 variants (Additional information about study of infectivity of cadavers from SARS-COV-2 deaths"}
+{"text": "Klebsiella pneumoniae (hvKp), unlike classical K. pneumoniae (cKp), are often responsible for community-acquired infections in otherwise healthy individuals. The acquisition of hypervirulence genes by sequence type 11 (ST11) carbapenem-resistant (CR) Kp endemic in Asia is a grave threat. Aztreonam-avibactam (ATM-AVI) is a monobactam combined with a \u03b2-lactamase inhibitor for the treatment of infections caused by Enterobacterales isolates that carry Class A, B, C and some Class D \u03b2-lactamases.Hypervirulent K. pneumoniae isolates were collected from 17 sites in China in 2019 as a part of the ATLAS global surveillance study. 220 isolates with MICs >1 \u00b5g/ml to meropenem (MEM), ceftazidime or ATM were selected for whole genome sequencing (Illumina Hiseq 2x150 bp reads). Analyses were carried out using the CLC Genomics Workbench (Qiagen). Presence of the aerobactin synthesis locus differentiated hvKp and cKp. Antimicrobial susceptibility was determined by CLSI broth microdilution.487 90 values for ATM-AVI  were lower than those for any comparator tested, with only two isolates testing with MIC >4 \u00b5g/ml. Of the isolates sequenced, 82/220 (37.3%) were ST11. 53/82 (64.6%) of these ST11 isolates were hvKp  and showed percentages of susceptibility < 90% to three last-line agents (0% MEM-susceptible (S); 18.9% amikacin (AMK)-S; 88.7% tigecycline (TGC)-S). Isolates of other STs (Non-ST11) were less frequently identified as hvKp  and more Non-ST-11 hvKp and cKp alike were S to MEM and AMK relative to isolates of ST11 . Likewise, the ATM-AVI MIC90 value (0.25 \u00b5g/ml) was 4-fold lower for Non-ST11 isolates.Of the 487 isolates, MICResults Tablein vitro activity against these isolates which displayed resistance to a range of last-line agents. CST and TGC also displayed some activity but are limited in utility due to nephrotoxicity and poor accumulation in blood, respectively. The spread of virulence factors leading to the complicated clinical presentation of hvKp infection into multidrug-resistant lineages warrants continued surveillance.CR ST11 hvKp represented at least 10.9% of the collected Kp isolates. ATM-AVI retained potent Mark Estabrook, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Krystyna Kazmierczak, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Francis Arhin, PhD, Pfizer, Inc. (Employee) Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "During the COVID-19 pandemic, a task force was assembled to collect data on patient characteristics and treatment exposures to assess what factors may contribute to patient outcomes, and to help develop institutional treatment guidelines. apriori. Covariates of interest included baseline comorbidities, admission level-of-care, vital signs, mortality outcomes, need for intubation, and specific pharmacological treatment exposures. Logistic regression was performed on our final model and reported as OR +/- 95% CI. A retrospective study was performed on COVID-19 inpatient admissions within a four-hospital community health system over a six-month period from April-October 2020. Positive COVID-19 immunology results and/in conjunction with an inpatient admission was criteria for inclusion. Covariates for age, gender, race were added A total of 349 patients met inclusion criteria. Pharmacotherapies were not associated with a difference in mortality in a four-hospital system. Corticosteroids (p = 0.99); Remdesivir (p = 0.79); hyrdroxychloroquine (p = 0.32); tocilizumab (p = 0.91); were not associated with mortality. ACE-inhibitor or angiotensin II receptor blockers OR 0.29 (0.09-0.93) (p = 0.03); convalescent plasma OR 7.85 (1.47-42.1) (p = 0.02); neuromuscular blocking agents (NMBA) OR 5.51 (1.28-23.8) (p = 0.02); vasopressors OR 17.6 (5.62-54.9) (p = 0.00) were associated with in-hospital mortality. Covariates that were associated with a difference in mortality were: age > 60 years OR 2.73 (1.04-7.14) (p = 0.04); structural lung disease OR 3.02 (1.28-7.10) (p = 0.01). Covariates not associated with mortality included African American race (p = 0.30); critical care admission (p = 0.19); obesity (p = 0.06); cardiovascular disease (p = 0.89); diabetes (p = 0.28).The use of corticosteroids, remdesivir, tocilizumab, and hydroxychloroquine, and admission to a critical care bed was not associated with a difference of in-hospital mortality. Patients who required vasopressors or NMBA were associated with in-hospital mortality. Despite national trends reporting increased mortality in patients with obesity, diabetes, cardiovascular disease, and of African American race, this was not observed in our health system safety net hospitals. All Authors: No reported disclosures"}
+{"text": "Achromobacter spp. is intrinsically resistant to multiple antibiotics, and the treatment options are limited. Cefiderocol (CFDC), a siderophore cephalosporin approved in US and EU, is active against a wide variety of aerobic Gram-negative bacteria, including carbapenem-resistant strains. In this study, in vitro and in vivo antibacterial activity of CFDC against Achromobacter spp. was evaluated.In vivo efficacy of CFDC was compared with meropenem (MEM), piperacillin-tazobactam (PIP/TAZ), ceftazidime (CAZ), and ciprofloxacin (CIP) in a neutropenic murine lung infection model (n=5), and compared with MEM in a immunocompetent rat lung infection model (n=3-7) caused by 2 A. xylosoxydans. In the murine model, treatment was given 2, 5, and 8 hours post-infection, and the numbers of viable cfu in lungs were determined 24 hours post-infection. In the rat model, the humanized PK in plasma resulting from CFDC 2 g every 8 h (3-h infusion) or meropenem 1 g every 8 h (0.5-h infusion) were recreated via continuous intravenous infusion for 4 days, following which cfu in lungs were determined.A total of 334 global isolates collected by IHMA from 39 countries in 2015-2019 were used. Minimum inhibitory concentrations (MICs) of CFDC and comparators were determined by broth microdilution method using iron-depleted CAMHB or CAMHB, respectively, as recommended by CLSI guidelines. in vitro activity with MIC50/90 of 0.06/0.5 \u00b5g/mL against 334 Achromobacter spp. Only 7 isolates (2.1%) had MICs > 4 \u00b5g/mL. These were the lowest values among all compound tested (Table). In the murine model, CFDC caused > 1.5 log10 decrease of viable cfu in lungs at 100 mg/kg dose (%fT >MIC: < 50%) from baseline control against both of strains (CFDC MIC: 0.5 and 2 \u00b5g/mL) (P< 0.05). No decrease of cfu in lungs was observed for the comparators at 100 mg/kg . In the rat model, humanized CFDC dosing reduced the viable cfu by >1 log10 CFU/lung compared with baseline controls (P< 0.05). MEM showed no significant activity.CFDC showed In vitro activity of CFDC and comparator agents against Achromobacter spp.334 Achromobacter spp. isolates collected from 2015 and 2019. The majority of isolates tested were A. xylosoxidans , followed by A. insolitus , Achromobacter sp. , A. denitrificans , and A. piechaudii .in vivo efficacy reflecting in vitro activity against A. xylosoxidans. The results suggested that CFDC has the potential to be an effective therapeutic option for Achromobacter spp. infections.CFDC showed potent Ryuichiro Nakai, MSc, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Ayaka makino, BSc, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Toriko Yoshitomi, -, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Rio Nakamura, BSc, Shionogi TechnoAdvance Research & Co., Ltd. (Employee) Meredith Hackel, PhD MPH, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Miki Takemura, MS, SHIONOGI & CO., LTD. (Employee) Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Yoshinori Yamano, PhD, Shionogi (Employee)"}
+{"text": "Pseudomonas aeruginosa isolated from cystic fibrosis sputum. Short reads were de novo assembled into 190 contigs and scaffold assembled to a length of 6.26\u2009Mbp. PhiSpy predicts that PA291 is free of prophages.Here, we report the genome sequence of PA291, a nonmucoid, multidrug-resistant strain of  Pseudomonas aeruginosa causes difficult-to-treat and often multidrug-resistant (MDR) infections, which severely impacts individuals with cystic fibrosis (CF) (sis (CF) , 2. CF psis (CF) . Phage tsis (CF) , 5; in pP. aeruginosa. The strain was named PA291 and found to have a nonmucoid phenotype, and a broth microdilution assay  revealed strong antibiotic resistance to ceftazidime (MIC\u2009>\u200916\u2009\u03bcg/ml) and intermediate resistance to aztreonam , gentamicin , and meropenem . Moreover, PA291 was lysed by phages PAK_P1, E215, and E217, which are under investigation to treat P. aeruginosa infections  was cultured overnight on blood agar at 37\u00b0C, and matrix-assisted laser desorption ionization\u2013time of flight (MALDI-TOF) mass spectrometry identified it as belonging to the species de novo assembled using SPAdes v3.14.1 (AE004091.2), W16407 (CP008869.2), VA-134 (CP013245.1), DVT779 (CP050330.1), SE5357 (CP054844.1), H47921 (CP008861.1), SE5443 (CP046405.1), PAO1161 (CP032126.1), PABL048 (CP039293.1), and PAC6 (CP053705.1). The quality and metrics were analyzed using QUAST v5.02   and alginate biosynthesis genes  , a polymyxin resistance gene (arnC), and a fosfomycin resistance gene (fosA). PhiSpy v4.2.6 (PA291 DNA extraction and purification was performed after overnight growth in Lennox broth (LB) at 37\u00b0C using the phenol-chloroform method . The DNA v3.14.1  into 190 v3.14.1  with the161 CP0326.1, PABLg, kinB)  were useP. aeruginosa strain isolated from CF sputum. The size of PA291 is consistent with that of other P. aeruginosa genomes, between 5.5 and 7\u2009Mbp  under accession number SRR14436913. The RAST annotation of the genome is available on Zenodo (https://zenodo.org/record/5114517#.YPXHVehKguV). The genome sequence of P. aeruginosa PA291 was deposited in GenBank under accession number JAGYWJ000000000, where the Prokaryotic Genome Annotation Pipeline (PGAP) annotation is also available.Data for"}
+{"text": "Nosocomial pneumonia (NP) remains associated with excess morbidity and mortality. The effect of NP on other measures of outcome and quality, such as re-admission at 30 days, remains unclear. Moreover, differing types of NP may have varying impacts on re-admissions.We conducted a multicenter retrospective cohort study within the Premier Research database, a source containing administrative, pharmacy, and microbiology data. The rate of rehospitalization at 30 days following the index discharge served as our primary endpoint. We compared NP patients readmitted with pneumonia (RaP) as the principal diagnosis to those readmitted for other reasons (RaO). We also compared readmission rates as function of the type of NP: ventilator-associated bacterial pneumonia (VABP), ventilated hospital-acquired bacterial pneumonia (vHABP), and non-ventilated HABP (nvHABP).Among 17,819 patients with NP, 14,123 (79.3%) survived to discharge, of whom 2,151 (15.2%) required an acute readmission within 30 days of index discharge. Of these, 106 (4.9%) were RaP, and the remainder were RaO. At index hospitalization, RaP patients were older (mean age (SD) 67.4 (13.9] vs. 63.0 (15.2) years), more likely medical (44.3% vs. 36.7%), and less chronically ill (median [IQR] Charlson scores (3 [2-5] vs. 4 [2-5]) than persons with RaO. Bacteremia (10.4% vs. 17.5%), need for vasopressors (15.1% vs. 20.0%), dialysis (9.4% vs. 16.5%), and/or sepsis (9.4% vs. 16.5%) or septic shock 14.2% vs. 17.1%) occurred less frequently in the RaP group. With respect to NP type, nvHABP was most common in RaP (47.2%) and VABP in RaO (38.1%). One in seven survivors of a hospitalization complicated by NP requires an acute rehospitalization within 30 days. However, few of these readmissions had a principal diagnosis of pneumonia, irrespective of NP type. This suggests that short-term readmission does not capture the quality of care initially delivered to patients for their NP. Of the 5% of NP subjects with RaP, the plurality initially suffered from nvHABP. Marya Zilberberg, MD, MPH, Cleveland Clinic (Consultant)J&J (Shareholder)Lungpacer Merck (Grant/Research Support)scPharma (Consultant)Sedana Spero (Grant/Research Support) Brian Nathanson, PhD, Lungpacer (Grant/Research Support)Merck (Grant/Research Support)Spero (Grant/Research Support) Laura A. Puzniak, PhD, Merck & Co., Inc. (Employee) Andrew F. Shorr, MD, MPH, MBA, Merck (Consultant)"}
+{"text": "Children infected with SARS-CoV-2 often have mild or no symptoms, making symptom screening an ineffective tool for determining isolation precautions. As an infection control measure, universal pre-procedural and admission SARS-CoV-2 testing for pediatric patients was implemented in April and August 2020, respectively. Limited data exist on the utility screening programs in the pediatric population. We performed a retrospective cohort study of pediatric patients (birth to 18 years) admitted to a tertiary care academic medical center from April 2020 to May 2021 that had one or more SARS-CoV-2 point-of-care or polymerase chain reaction tests performed. We describe demographic data, positivity rates and repeat testing trends observed in our cohort.A total of 2,579 SARS-CoV-2 tests were performed among 1,027 pediatric inpatients. Of these, 51 tests (2%) from 45 patients (4.3%) resulted positive. Community infection rates ranged from 4.5-60 cases/100,000 persons/day during the study period. Hispanic patients comprised 16% of the total children tested, but were disproportionately overrepresented (40%) among those testing positive . Of 654 children with repeated tests, 7 (0.1%) converted to positive from a prior negative result. Median days between repeat tests was 12 (IQR 6-45), not necessarily performed during the same hospital stay. Five of these 7 patients had tests repeated < 3 days from a negative result, of which only 2 had no history of recent infection by testing performed at an outside facility. Pre-procedural tests accounted for 35% of repeat testing, of which 0.9% were positive. Repeated tests were most frequently ordered for patients in hematology/oncology (35%) and solid organ transplant/surgical (33%) wards, each with < 3% positive conversion rate. Notably, no hematopoietic stem cell transplant patients tested positive for SARS-CoV-2 during the study period.Pediatric SARS-CoV-2 Testing Distributed by Race/EthnicityThe positivity rate of universal pre-procedural and admission SARS-CoV-2 testing in pediatric patients was low in our inpatient cohort. Tests repeated < 3 days from a negative result were especially low yield, suggesting limited utility of this practice. Diagnostic testing stewardship in certain populations may be useful, especially as community infection rates decline.Michael J. Smith, MD, M.S.C.E, Merck (Grant/Research Support)Pfizer (Grant/Research Support) Rebekah W. Moehring, MD, MPH, UpToDate, Inc."}
+{"text": "Staphylococcus aureus (MRSA) is a prominent colonizer in cystic fibrosis (CF) patients that causes acute pulmonary exacerbation (APE). Vancomycin is the first line treatment for APE of CF; however, optimal alternatives remain poorly defined. The goal of this study was to determine the safety and efficacy of ceftaroline in CF patients presenting with an APE caused by MRSA.Methicillin-resistant > 10% lower than the patient\u2019s baseline. A positive MRSA culture within 90 days before or 21 days after hospital admission and receipt of > 7 days of either vancomycin or ceftaroline was required for inclusion. Patients were excluded for receipt of a lung transplant, > 48 hours of alternative MRSA therapy, renal replacement therapy, or an APE secondary to fungal or mycobacterium infection. The primary outcome was the return to > 90% of baseline lung function measured by discharge %FEV1 in comparison to baseline %FEV1.This study was a single-center, retrospective cohort study from January 1, 2011 to January 1, 2020. The study included adult CF patients admitted for APE with %FEV1 Clostridioides difficile infection 0 (0%) vs. 1 (3%), P = >0.99; or acute kidney injury 2 (9%) vs. 5 (15%), P = 0.69. Fifty-six patients were included in the analysis . There were no differences in baseline characteristics (Table 1). Eleven (50%) patients in the ceftaroline group and 19 (56%) in the vancomycin group met the primary outcome (P = 0.79) . FEV1 measurements at baseline, admission, and discharge were not different between treatments . Patients treated with ceftaroline had a longer length of stay during hospital admission, 14 days (IQR 13-14) vs.10 days (IQR 7-14), P = 0.01. Other secondary outcomes were similar between the ceftaroline and vancomycin groups, respectfully, including 30-day readmission rate, 6 (27%) vs. 12 (35%), P = 0.57; 30-day mortality, 0 (0%) vs. 2 (6%), P = 0.51; neutropenia 3 (12%) vs. 1 (3%), P = 0.29; Table 1. Baseline characteristics for ceftaroline and vancomycin treated patients1Lumacaftor/ivacaftor, tezacaftor/ivacaftor; 2Piperacillin/tazobactam, aminoglycoside, furosemide, contrast dye, lisinopril, NSAIDs, colistin, phenylephrine; 3Methimazole, sulfasalazine, trimethoprim/sulfamethoxazole; 4Albuterol, hypertonic saline, dornase alpha, azithromycin, ibuprofen, inhaled aminoglycoside, inhaled colistin, corticosteroid; 5Azithromycin, aminoglycoside, fluroquinolone, cephalosporin, carbapenem, piperacillin/tazobactam. Data represents n (%) unless noted. CFTR=cystic fibrosis transmembrane conductance regulator.Figure 1. %FEV1 trend from baseline to discharge in patients treated with ceftaroline or vancomycin(A) Percentage (%) of patients who met the primary outcome in each group; (B) Mean %FEV1 change between ceftaroline (square) and vancomycin (circle) with error bars representing standard deviationsThis study found no difference in safety and efficacy outcomes between vancomycin and ceftaroline. Our small cohort supports ceftaroline as an alternative agent for the treatment of MRSA mediated APE of CF. All Authors: No reported disclosures"}
+{"text": "Bambusa oldhamii Munro, known as \u201cgreen bamboo\u201d, is famous for its edible bamboo shoots and fast-growing timber. The green and yellow striped-culm B. oldhamii variety, named B. oldhamii f. revoluta W.T. Lin & J. Y. Lin, is an attractive system for researching the culm color variation of B. oldhamii.The clumping bamboo 1, GA3, GA4, and GA7 was performed using HPLC\u2013MS/MS platforms.Millions of clean reads were generated and assembled into 604,900 transcripts, and 383,278 unigenes were acquired with RNA-seq technology. The quantification of ABA, IAA, JA, GAB. oldhamii f. revoluta. Phytohormone contents, especially GA1 and GA7, were higher in B. oldhamii. Approximately 21 transcription factors (TFs) were differentially expressed between the two groups: the bZIP, MYB, and NF-YA transcription factor families had the most DEGs, indicating that those TFs play important roles in B. oldhamii culm color variation. RNA-seq data were confirmed by quantitative RT-PCR analysis of the selected genes; moreover, phytohormone contents, especially those of ABA, GA1 and GA7, were differentially accumulated between the groups. Our study provides a basal gene expression and phytohormone analysis of B. oldhamii culm color variation, which could provide a solid fundamental theory for investigating bamboo culm color variation.Differential expression analysis showed that 449 unigenes were differentially expressed genes (DEGs), among which 190 DEGs were downregulated and 259 DEGs were upregulated in  Dendrocalamus latiflorus, Bambusa oldhamii, and Bambusa chungii; and running bamboos , such as Phyllostachys edulis  and fast-growing culm timber  (Paeonia suffruticosa (Rosa chinensis (m timber . B.\u00a0oldholorata) , Paeoniaruticosa , and Roshinensis .Basidiomycota phylum; they comprise red to red-violet betacyanins and yellow-orange betaxanthins  and jasmonic acid (JA) could promote anthocyanin biosynthesis, while auxin and gibberellin (GA) could inhibit anthocyanin biosynthesis . Abscisike genes . JA treake genes . JA treake genes . The accke genes . The plake genes . GA3 incing time . Treatmeotenoids .MdDFR and MdUFGT and influence fruit coloration  were labeled LZ_1, LZ_2, and LZ_3, and those from B.\u00a0oldhamii f. revoluta W.T. Lin & J. Y. Lin (HLZ) were labeled HLZ_1, HLZ_2, and HLZ_3, representing three biological replicates of each type of bamboo. The culm skin samples were frozen in liquid nitrogen immediately for further phytohormone detection, RNA-seq, and relative gene expression. Total RNA was isolated using plant RNA isolation kits .The middle and lower culm internode epidermis samples that were removed from https://www.illumina.com) at Wuhan Metware Biotechnology Co., Ltd. . The output data contained raw reads in fastq format, that were then processed for quality control, including filtering and trimming of low confidence bases, biased nucleotide composition, adapters, duplicates and low-quality reads to acquire clean reads. Trinity (2.6.6) (Library construction and sequencing steps were performed based on the Illumina HiSeq platform for RNA-seq protocols ( (2.6.6)  and Cors (2.6.6)  were usee-value = 1e\u22125, and the Pfam annotation was performed using the hmmscan command of the HMMER 3.2 package with e-value = 0.01. Transcription factor annotation was performed with iTAK (1.7a) (Nr (NCBI nonredundant protein sequences), Pfam (Protein family), KOG (Protein family), Swiss-Prot, Trembl, KEGG (Kyoto Encyclopedia of Genes and Genomes), and GO (Gene Ontology) were used for gene annotation. The Nr, KOG, Swiss-Prot, Trembl, KEGG, and GO annotations were performed using BLAST (v2.7.1) with an K (1.7a)  with defWe used the assembled transcriptome of Trinity as a reference and then mapped the clean reads of each sample to the reference with RSEM software. FPKM (Fragments Per Kilobase of transcript per Million fragments mapped) values were calculated to estimate gene expression and abundance after normalization of the mapped reads and transcript lengths. The R package Pheatmap was used to draw a heatmap with normalized log2(FPKM+1) data and clusters of expression patterns with kmeans_k = 10. The color from red to blue indicates gene expression from high to low.After acquiring the abundance information and performing normalization, gene expression between the groups was compared. DESeq2 (1.22.2) was used to calculate the differentially expressed genes between the LZ and HLZ groups, which were corrected with FDR  by Benjamini\u2013Hochberg methods. Differentially expressed genes were filtered with the condition of \u2014log2(Fold Change)\u2014 \u2265 1 and FDR <\u00a00.05.\u00ae II Q RT SuperMix for qPCR . Quantitative Real-Time PCR (qRT-PCR) was performed on a LightCycler\u00ae 480 II Real-Time PCR system  using the Unique Aptamer\u2122\u00a0qPCR SYBR\u00ae Green Master Mix. The components of the qRT-PCR were as follows: SYBR Premix Ex Taq (2x) (10 \u00b5l), forward primers (0.5 \u00b5M), reverse primers (0.5 \u00b5M), cDNA template (2 \u00b5l), and ddH2O to 20\u00a0\u00b5l. Then, qRT-PCR was performed as follows: initial denaturation at 95\u00a0\u00b0C for 5 min; 40 cycles of denaturation at 95\u00a0\u00b0C for 10 s and annealing at 72\u00a0\u00b0C; and finally, steps for melt-curve analysis . Actin was used as the internal control  was performed by Genepioneer Biotechnologies Co., Ltd.  using an HPLC\u2013MS/MS platform.The quantification of endogenous abscisic acid (ABA), auxin , jasmonic acid (JA), and gibberellic acid (GAn\u00a0=\u00a09) with a significant difference (p\u00a0<\u00a00.05) according to unpaired t-tests.The enrichment of up- and downregulated genes was determined using GOseq and KOBAS . The corB.\u00a0oldhamii (LZ) is a species of clumping bamboo (B.\u00a0oldhamii variety referred to as B.\u00a0oldhamii f. reboluta W.T. Lin & J. Y. Lin (HLZ) has green culms with yellow stripes of random widths. The culm skin was removed from LZ and HLZ to research the correlation of culm color variation on the phytohormone contents and gene expression levels.g bamboo  and it hB.\u00a0oldhamii with three biological replicates marked LZ_1 - LZ_3 and from B.\u00a0oldhamii f. revoluta with three biological replicates marked HLZ_1 - HLZ_3. After the cDNA library was constructed and sequenced, approximately 282 million raw sequence reads were obtained from the RNA-seq experiment, and 267 million clean sequence reads remained after filtering with a Q20 above 98% after quality control was performed. The error correction and GC content are shown as follows . The annotation results produced 38.64%, 70.18%, 44.92%, 69.23%, 40.03%, 57.24%, and 47.26% unigenes annotated in the KEGG, NR, SwissProt, Trembl, KOG, GO, and Pfam databases, respectively. Approximately 274,681 unigenes were annotated in at least one database.All of the unigenes were annotated using seven databases  (https:/The unigenes annotated with GO functions were assigned to three main ontologies: molecular function (MF), cellular component (CC), and biological process (BP). The terms cellular process (GO:0009987), metabolic process (GO:0008152), biological regulation (GO:0065007), and response to stimulus (GO:0050896) were the most common BP ontologies; the terms cell (GO:0005623), cell part (GO:0044464), and organelle (GO:0043226) were the most common CC ontologies; and binding (GO:0005488), catalytic activity (GO: 0003824), transporter activity (GO:0005215), and transcription regulator activity (GO:0140110) were the most common MF ontologies . The uniunigene-21666.123476) annotated with metallothionein that was highly expressed in all six samples, and cluster 1 included 8 genes that were more highly expressed in LZ samples. unigene-21666.69211 (mitogen-activated protein kinase kinase kinase ANP1), unigene-21666.134631 (SAUR family protein), unigene-21666.128797 (EREBF-like factor), and others without a specific annotation were included. The upregulated genes in HLZ were classified into 10 clusters , unigene-21666.90795 ; unigene-21666.169778 (serine/threonine-protein kinase PBS1), unigene-21666.167280 (granule-bound starch synthase), unigene-21666.149531 (acyl-[acyl-carrier-protein] desaturase), unigene-21666.125526 (anthranilate O-methyltransferase), unigene-21666.107961 (serine/threonine-protein kinase PBS1), and others without specific annotations , metabolic process (GO:0008152), response to stimulus (GO:0050896), and biological regulation (GO:0065007) terms were the most common BP ontologies; cell (GO:0005623), cell part (GO:0044464), organelle (GO:0043226), and membrane (GO:0016020) were the most common CC ontologies; and binding (GO:0005488) and catalytic activity (GO:0003824) were the most common MF ontologies . For theDifferential expression analysis between the LZ and HLZ groups showed that 190 and 259 DEGs were down- and upregulated in the HLZ culm skin samples. The GO functional enrichment revealed that the downregulated DEGs were more enriched in monocarboxylic acid transport (GO:0008028), transporter complex (GO:1990351), transmembrane transporter complex (GO:1902495), replication fork (GO:0005657), organelle envelope lumen (GO:0031970), lipid transporter activity (GO:0005319), and monocarboxylic acid transmembrane transporter activity (GO:0008028) .The upregulated DEGs were more enriched in biological processes and molecular functions. In particular, the top three terms were L-phenylalanine metabolic process (GO:0006558), L-phenylalanine catabolic process (GO:0006559), and entrainment of the circadian clock (GO:0009649), followed by photoreceptor activity (GO:0009881), phenylalanine ammonia-lyase activity (GO:0045548), NAD(P)H dehydrogenase (quinone) activity (GO:0003955), and carbon-nitrogen lyase activity (GO:0016840) .The downregulated DEGs were enriched in the alpha-linolenic acid metabolism pathway (ko00592) , and theB. oldhamii, the content of endogenous abscisic acid (ABA), auxin , jasmonic acid (JA), and gibberellic acid  was detected  and gene expression was analyzed with corresponding samples. There were 18 genes whose expression was significantly related to ABA accumulation patterns (1 (7 (1 and GA7 were more enriched in the GO terms of chloride channel complex (GO:0034707), ion channel complex (GO:0034702), protein kinase complex (GO:1902911), transporter complex (GO:1990351), anion channel activity (GO:0005253), chloride channel activity (GO:0005254), phosphatidylinositol bisphosphate binding (GO:1902936), and phosphatidylinositol-3,5-bisphosphate binding (GO:0080025)  pathway (ko04712) (unigene-21666.126192 and unigene-21666.239597) and protein FLOWERING LOCUS T (FT) (unigene-54902.0) were up-regulated. Moreover, the expression of genes significantly related to ABA, GA1 and GA7 showed that unigene-21666.90795  was relatively highly expressed in HLZ  . All theko04712) . In the d in HLZ . The genunigene-21666.57044 (bHLH), unigene-21666.218036 (MYB-related), and unigene-21666.41135 (SBP) were not detected in the HLZ samples and were only detected in the LZ samples. bHLH transcription factors can interact with MYB transcription factors to regulate anthocyanin biosynthesis (unigene-21666.194872 (FAR1), unigene-21666.98709 (MADS-M-type), unigene-21666.139384 (MYB), and unigene-21666.212035 (TRAF) were not detected in the LZ samples and were only detected in the HLZ samples. MADS-box transcription factors are involved in flower promotion and development, and simultaneous death usually follows after mass production of bamboo flowers  were annotated as the transcription factor HY5. The target genes of HY5 participate in many biological signaling processes, such as light signaling, circadian clock, anthocyanin biosynthesis, and chlorophyll biosynthesis , CHI , and FLS (flavonol synthase) to regulate anthocyanin biosynthesis  that were members of the MYB transcription factor family. MYB transcription factors could regulate anthocyanin biosynthesis . The psbP protein is required for the photosystem II complex and normal thylakoid architecture in Arabidopsis thaliana (unigene-54902.0 (FT) gene is upregulated in the circadian rhythm (plant) pathway. The FT gene (FLOWERING LOCUS T), a mobile stimulus expressed in leaves and then translocated to the shoot apex, is essential for floral induction in Arabidopsis (unigene-21666.98709 (MADS-M-type) is relatively highly expressed in HLZ. MADS-box genes are essential for flower induction, promotion, and maturation . These proteins are usually divided into four classes: (1) R2R3-MYB, (2) 1R-MYB, MYB-related, and others, (3) 3R-MYB, and (4) 4R-MYB . Among tis genes .unigene-21666.57044, was differentially expressed. These results demonstrate that MYB and bHLH transcription factors could regulate the color variation of bamboos. However, how the crosstalk among the transcription factors and hormone regulation influences color variation still needs further investigation.The family of Basic Helix-Loop-Helix (bHLH) transcription factors contains approximately 60 conserved amino acid domains that bind to the promoter of E-box cis-elements to regulate downstream genes , and theB. oldhamii via combined RNA-seq and endogenous phytohormone content variation analyses. The results showed that bZIP, MYB, HY5, and other differentially expressed transcription factors play a role in B. oldhamii culm color variation. Moreover, phytohormone contents, especially GA1 and GA 7, were more highly accumulated in LZ, but many flower-regulated genes were more highly expressed in HLZ, which indicates that HLZ may flower more rapidly than LZ and that the senescence pathways may be involved in bamboo culm color variation. The transcription factors HY5, MYB, and bHLH participate in culm color variation by regulating pigment biosynthesis pathways to cause bamboo culm color variation, but how the regulatory pathways between transcription factors and phytohormones influence culm color variation still needs to be deeply investigated.This paper focused on investigating the culm color variation of the clumping bamboo 10.7717/peerj.12796/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.12796/supp-2Supplemental Information 2Click here for additional data file.10.7717/peerj.12796/supp-3Supplemental Information 3Click here for additional data file.10.7717/peerj.12796/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.12796/supp-5Supplemental Information 5Click here for additional data file.10.7717/peerj.12796/supp-6Supplemental Information 6Click here for additional data file.10.7717/peerj.12796/supp-7Supplemental Information 7Click here for additional data file.10.7717/peerj.12796/supp-8Supplemental Information 8Click here for additional data file.10.7717/peerj.12796/supp-9Supplemental Information 9Click here for additional data file.10.7717/peerj.12796/supp-10Supplemental Information 10Click here for additional data file.10.7717/peerj.12796/supp-11Supplemental Information 11Click here for additional data file.10.7717/peerj.12796/supp-12Supplemental Information 12Click here for additional data file.10.7717/peerj.12796/supp-13Supplemental Information 13Click here for additional data file.10.7717/peerj.12796/supp-14Supplemental Information 14Click here for additional data file.10.7717/peerj.12796/supp-15Supplemental Information 15Click here for additional data file.10.7717/peerj.12796/supp-16Supplemental Information 16Click here for additional data file.10.7717/peerj.12796/supp-17Supplemental Information 17Click here for additional data file.10.7717/peerj.12796/supp-18Supplemental Information 18Click here for additional data file.10.7717/peerj.12796/supp-19Supplemental Information 19Click here for additional data file.10.7717/peerj.12796/supp-20Supplemental Information 20Click here for additional data file.10.7717/peerj.12796/supp-21Supplemental Information 21Click here for additional data file.10.7717/peerj.12796/supp-22Supplemental Information 22Click here for additional data file."}
+{"text": "Helicobacter pylori (H.pylori) may cause dyspepsia and/or unexplained functional nonspecific, gastrointestinal complaints of the irritable bowel syndrome (IBS) spectrum. Hitherto, in H. pylori infected patients with symptoms of the IBS spectrum the occurrence of additional food intolerance/malabsorption is not evaluated. We used a retrospective analysis of charts from 548 patients who presented with gastrointestinal complaints of the irritable bowel syndrome spectrum. An enzyme-linked IgA immunosorbent assay or histologic evaluation of gastric mucosa were used to detect H. pylori infection. A hydrogen breath (H2) test was performed to evaluate fructose malabsorption (FM) and lactose intolerance (LIT). Serum diamine oxidase value of <10 U/ml and a response to a histamine-reduced diet was used to identify histamine intolerance (HIT). We found 293 patients infected with H. pylori, within these were 58 H. pylori patients with LIT, 23 H. pylori LIT patients with FM and 46 H. pylori LIT patients with HIT. Additionally, 13 H. pylori, lactose- and histamine intolerance patients also had FM. The Kruskal Wallis test and pairwise comparison were used to analyze differences of the area under the curve of expiratory hydrogen. In lactose H2 breath tests compared with LIT-only patients, LIT with H. pylori, LIT and H. pylori with HIT, LIT and H. pylori with FM showed significantly higher exhaled H2 levels (p=0.022). Pairwise comparison demonstrated H. pylori infected patients with LIT exhaled more H2 compared to LIT-only (p=0.029). H. pylori with lactose- and histamine intolerance, and H. pylori with lactose-, histamine intolerance and FM compared to H. pylori-only patients indicated a significantly higher occurrence of stomach pain during lactose H2 breath tests . We demonstrate that LIT patients with high expiratory H2 levels in lactose breath tests may have H. pylori infection and possibly additional food intolerance/malabsorption. Subsequently, besides H. pylori eradication, a dietician is necessary for an individually tailored reduction- or exclusion diet of symptom triggering food components.Infection with  AUC, Area under the curve; DAO, diamine oxidase; FM, fructose malabsorption; GI, gastrointestinal; HIT, histamine intolerance; IBS, irritable bowel syndrome; LIT, lactose intolerance.Helicobacter pylori (H. pylori) infection is the most prevalent human pathogen and is present in more than 50 % of worldwide populations. If H. pylori is detected, then an eradication therapy is mandatory. Some association of H. pylori infection and dyspepsia and/or unexplained functional, nonspecific, non-allergic gastrointestinal (GI) complaints of the irritable bowel syndrome (IBS) spectrum is documented. Reduction of these symptoms due to H. pylori eradication has been shown . Based al., 2019).Unexplained functional, nonspecific, non-allergic GI complaints and dyspepsia, including IBS and IBS-like syndromes, are widespread and costly, and a main reason for consultations in primary care settings . GeneraH. pylori-infected patients for additional food intolerance/malabsorption, including CD, FM, HIT and LIT. All included patients presented with dyspepsia/unexplained functional, nonspecific, non-allergic GI complaints of the IBS spectrum. Compared to LIT-only, patients with LIT and H. pylori combined with or without additional food intolerance/malabsorption demonstrated significantly higher expiratory hydrogen (H2) values during lactose tolerance breath tests. In this study we have evaluated H. pylori infection. 293 were infected with H. pylori and in 255 H. pylori was not found. Eradication therapy was started after performance of H2 breath tests and included in this evaluation were only patients who took part in all tests. Patients with alarming symptoms, such as vomiting, rectal bleeding or unintentional weight loss were excluded.During the retrospective evaluation, we found 548 patients who presented with dyspepsia/unexplained functional, nonspecific, non-allergic GI complaints of the IBS spectrum. Main symptoms included abdominal pain, diarrhea, loose stools, bloating, constipation and postprandial fullness. They were examined for additional food intolerance/malabsorption, including CD, FM, HIT, and LIT. In evaluated patients, either an enzyme-linked IgA immunosorbent assay  or a histologic evaluation of gastric mucosa was used to detect 2 breath tests  were used for LIT and FM testing as described earlier .Hal., 2020). After A thorough anamnesis, concerning abdominal complaints, and a timely relation to the ingestion of food or drinks, including pharmaceutical treatments that might influence dyspepsia/unexplained functional, nonspecific, non-allergic GI complaints of the IBS spectrum was performed. A registered dietitian was consulted to implement and monitor an individualized diet to reduce and/or eliminate symptom-triggering foods. For screening of celiac disease, antibodies against tissue transglutaminase were measured with the anti-tTG IgA ELISA  method. The study follows the ethical guidelines of the Declaration of Helsinki and was approved by the Ethical Committee of the Johannes Kepler University in Linz, Austria (No. K-107-16).2 breath tests were calculated and compared with the Kruskal-Wallis test, and pairwise comparison. The Chi-square and the Fisher's exact test were applied for evaluation of symptoms. The level of significance was set to 5 %. The Bonferroni correction was used for multiple testing.Descriptive statistics are presented as medians with interquartile ranges (IQR). The data distribution was assessed with a Shapiro Wilk test. For not normally distributed data non-parametric tests were applied. The areas under the curve (AUC) of exhaled hydrogen during HStatistical analyses were performed with IBM SPSS statistics version 25.0 , and GraphPad Prism version 9.0.0.  was used for the generation of figures. H. pylori, male/female 93/200, median age 50 years (IQR 21), age range 17-93 years. We identified 68 patients with H. pylori infection only (12.4 %) - male/female 25/43, median age 56 years (IQR 22), age range 25-93 years. Hydrogen breath tests neither showed LIT nor FM in these patients, and serum DAO values were within normal >10 IU/mL (median 17.9 IU/mL (IQR 12.1), range 10.1-63.4 IU/mL). 58 patients with H. pylori had additional LIT (10.6 %) - male/female 20/38, median age 50 years (IQR 17), age range 22-84 years. Hydrogen breath tests did not show FM in these patients, and DAO values were >10 IU/mL (median 17 IU/mL (IQR 9.5), range 10.5-80 IU/mL). 17 H. pylori patients had only additional FM (3.1 %), male/female 3/14, median age 57 years (IQR 15), age range 23-78 years. Hydrogen breath tests did not show LIT in these patients, and DAO values were >10 IU/mL (median 14.7 IU/mL (IQR 5.6), range 10.7-30.6 IU/mL). However, 47 H. pylori patients had additional HIT (8.6 %), male/female 12/35, median age 50 years (IQR 22), age range 24-91 years. Their DAO values were <10 IU/mL (median 5 IU/mL (IQR 5.2), range 1.5-9.7 IU/mL). Moreover, hydrogen breath tests did show neither LIT nor FM in this group. For this retrospective evaluation, we used 548 patients with IBS spectrum symptoms. We found 293 infected withH. pylori combined with LIT and FM (4.2 %) - male/female 6/17, median age 45 years (IQR 21), and age range 17-82 years. Their DAO values were >10 IU/mL (median 16.6 IU/mL (IQR 9.5), range 12.4-71 IU/mL). In 46 patients we found H. pylori combined with LIT and HIT (8.4 %) - male/female 17/29, median age 43 years (IQR 21), age range 22-83 years. Accordingly, their DAO values were <10 IU/mL (median 6.6 IU/mL (IQR 3.0), range 1.5-9.9 IU/mL) and hydrogen breath tests did not show FM. In 17 H. pylori patients we found additional FM combined with HIT (3.1 %) - male/female 6/11, median age 54 years (IQR 22), age range 28-79 years. Their DAO values were <10 IU/mL (median 4.1 IU/mL (IQR 6.7), range 1.5-10 IU/mL). Hydrogen breath tests did not show LIT in these patients. 13 patients with H. pylori had LIT and FM combined with HIT (2.4 %). Their DAO values were <10 IU/mL (median 7.8 IU/mL (IQR 3.5), range 2.1-9.9 IU/mL). Antibodies against tissue transglutaminase were not detectable in 289 H. pylori patients. Four patients with H. pylori infection demonstrated celiac disease (0.8 %) with elevated tissue transglutaminase antibodies and following histologic diagnosis of the duodenal mucosa as shown in Table 1Additionally, 23 patients showed 2 breath tests. 102 patients (18.6 %) were positive for FM , range 19-91 years). Their DAO values were > 10 IU/mL, with median 16.9 IU/mL (IQR 7.9) and range 10.2-62.5 IU/mL and they did not show LIT in lactose H2 breath tests. In all of these 255 patients neither antibodies against H. pylori nor tissue transglutaminase were detected.In 153 (27.9 %) patients , age range 16-84 years, LIT only was diagnosed. Their DAO values were >10 IU/mL (median 16.6U/ml (IQR 10.2), range 10.2-80 IU/mL) and they did not show FM in fructose HH. pylori, to H. pylori, LIT with HIT and, to H. pylori, LIT with FM demonstrated significantly higher exhaled hydrogen values (p=0.022) , corrected for multiple testing (test statistic 35.206 and standard error 12.48). Due to the low number of H. pylori patients with LIT, FM and HIT these were not included in Figure 1The Kruskal Wallis test for the comparison of the AUCs of LIT-only, to patients with LIT and  Figure 1. PairwisH. pylori and LIT patients were abdominal pain, bloating, diarrhea, and increased bowel movements. Other GI symptoms included nausea, heartburn, belching, fullness, and excessive mucus in the throat. Extra-intestinal symptoms were headache, fatigue, vertigo and one eczema. Various double and few triple combinations of these symptoms were indicated. GI-symptoms were indicated by 160 of 279 LIT patients (57 %) during H2 breath tests. As shown in Figure 2H. pylori with LIT and FM, and H. pylori with LIT, FM and HIT compared to H. pylori-only patients demonstrate significantly higher occurrence of stomach pain . Patients with H. pylori-only infection did specify symptoms, although neither bloating nor diarrhea were mentioned during lactose breath tests. All patients >50 years old were screened by colonoscopy and no pathology was present.During performed lactose breath tests patients specified their GI and extra-intestinal symptoms on paper. Indicated GI symptoms by H. pylori in 262 patients, male/female 84/178, median age 51 years (IQR 22), age range 17-93 years, with mean 77.2 IU/mL (range 21-200 IU/mL) IgA antibodies. 63 patients had H. pylori only, male/female 25/38, median age 53 years (IQR 24), age range 25-93 years, and they had mean 73.9 IU/mL (range 21-200 IU/mL) IgA antibodies. 45 patients had H. pylori combined with LIT, male/female 17/28, median age 49 years (IQR 16), age range 26-86 years, with mean 63.8 IU/mL (range 21-200 IU/mL) IgA antibodies. 46 were H. pylori and HIT patients male/female 9/37, median age 49.5 years (IQR 24), age range 24-91 years, they had IgA antibodies mean 65.3 IU/mL (range 22-200 IU/mL). 19 were H. pylori and FM patients, male/female 3/16, and median age 56 years (IQR 17), age range 23-78 years, with mean 74.1 IU/mL (range 21-200 IU/mL) IgA antibodies. 44 were H. pylori and LIT with HIT patients, male/female 17/27, median age 43 years (IQR 18), age range 22-83 years, IgA antibodies with mean 84.9 IU/mL (range 21-200 IU/mL). 19 were H. pylori patients with LIT and FM male/female 6/13, median age 51 years (IQR 21), age range 17-79 years, with mean 111 IU/mL (range 34-200 IU/mL) IgA antibodies. 15 were H. pylori, FM and HIT patients male/female 5/10, median age 54 years (IQR 28), age range 28-58 years, IgA antibodies with mean 95.3 IU/mL (range 24-200 IU/mL). 11 H. pylori, LIT, HIT with FM patients, male/female 2/9, median age 41 years (IQR 32), age range 24-83 years, with mean 92.8 IU/mL (range 25-200 IU/mL) IgA antibodies.The enzyme-linked IgA immunosorbent assay  detected IgA antibodies against H. pylori significant different among all groups (p=0.044). Patients with H. pylori, LIT and FM had the highest quantity of IgA antibodies. Pairwise comparisons showed significantly lower antibody values comparing H. pylori, LIT and FM to H. pylori and LIT (p=0.005), to H. pylori and HIT (p=0.005), to H. pylori with FM (p=0.034) and, to H. pylori only (p=0.016). However, correction for multiple testing showed no significance.As shown in Figure 32 (p=0.39) comparing all groups (data not shown). Fructose malabsorption breath tests (n=159), using the Kruskall-Wallis test for evaluation of AUCs, showed no differences of exhaled HSee also the Supplementary data.From a clinical perspective, considerable overlap exists between food intolerance/ malabsorption and unexplained dyspepsia/ functional, nonspecific, non-allergic GI complaints and disorders, including IBS and IBS-like disorders. Currently, IBS is classified as a functional gastrointestinal disorder, according to Rome IV criteria, but new disease models are being proposed continuously Talley, 2020). Food i020. FoodH. pylori, the hormone gastrin and the biogenic amine histamine are main stimulants of gastric acid secretion . In patal., 2018). Due toal., 2018), we useal., 2020).H. pylori infection and its association with deficiencies of micronutrients, minerals and vitamins was reported . A relaley, 2020). Succesal., 2000). Additial., 2020). Severakas, 2020). FinallH. pylori patients with dyspepsia/functional, nonspecific, non-allergic GI complaints of the IBS spectrum, besides a thorough anamnesis, it seems useful to include examinations for food intolerance/malabsorption with FM- and lactose intolerance H2 breath tests, serum DAO determination and screening for celiac disease . 2 production. Therefore, the clinical diagnosis of LIT and FM is primarily performed with H2 breath tests. Impaired degradation of ingested histamine due to an anticipated gastrointestinal DAO deficiency, promotes HIT. So far, serum DAO is not established to reflect GI histamine-degrading DAO activity , fermen al. 2021). Noneth al. 2021; Mu\u0161i\u010d e al. 2021). Althou al. 2021).Generally, only few studies have reported on combined occurrence of food intolerance/malabsorption. Nonetheless, more than 30 % of patients with dyspepsia/ unexplained functional, nonspecific, non-allergic GI complaints and carbohydrate intolerance/malabsorption may have combinations of these . Next tH. pylori, the presence of additional food intolerance/ malabsorption induced significantly increased expiratory H2 values . In thikas, 2020).H. pylori patients when this infection is combined with LIT and FM or with LIT, FM and HIT . HoweveH. pylori infection may have additional food intolerance/malabsorption. Moreover, we show that in LIT the presence of H. pylori infection causes significantly higher expiratory H2 values in lactose tolerance breath tests. Subsequently, besides eradication, a registered and experienced dietician is essential for the development of an individually tailored reduction- or exclusion diet of symptom triggering food components.In conclusion, we describe, that patients with dyspepsia/unexplained functional, nonspecific, non-allergic GI complaints of the IBS spectrum having LIT and We are indebted to Mrs. Katharina Schnedl, Cardiff University, Cardiff, UK, who performed English language corrections.Wolfgang J. Schnedl received speaking honoraria from Sciotec. The other authors have no conflict of interest."}
+{"text": "Mycobacterium smegmatis mc2 155 host. The genome of JeTaime is 75,099 bp (142 predicted genes), and that of Luna22 is 53,730 bp (87 predicted genes). Both phages exhibit Siphoviridae morphology.The mycobacteriophages JeTaime (E cluster) and Luna22 (Q cluster) were isolated from soil in Providence, Rhode Island, and Charleston, South Carolina, respectively, using a  Mycobacterium smegmatis mc2 155, as described in the SEA-PHAGES Phage Discovery Guide  program through the Howard Hughes Medical Institute (HHMI) . The phale tails .de novo into a single contig using Newbler v2.9 (https://cpt.tamu.edu/computer-resources/pause), genome termini with 3\u2032 cohesive overhangs were identified, as follows: JeTaime, 5\u2032-CGCTTGTCA-3\u2032; Luna22, 5\u2032-TAAGCCGCGCGGTA-3\u2032. The Promega Wizard DNA cleanup system was used to extract DNA from phage lysates, and sequencing libraries were prepared with a NEBNext Ultra II DNA library prep kit v3). Libraries for each phage were independently generated, and genomes were sequenced separately. Sequencing was performed at the Pittsburgh Bacteriophage Institute on an Illumina MiSeq system (MiSeq reagent kit v3) , TOPCONS v2 (https://phagesdb.org/DNAMaster). All programs used default settings.Each genome was annotated on PECAAN  using GLPCONS v2 , and thePCONS v2 . tRNAs aPCONS v2  and tRNAPCONS v2 , and comKT222941) genomes. The other 103 published E-cluster members typically contain endonuclease VII and/or DNA methylase in that genome region. Also, the JeTaime gp89 (unidentified function) is rare, occurring in only one other E-cluster bacteriophage, 244 (GenBank accession number DQ398041). Whole-genome BLASTn alignment (NC_004681.1) (99.24% identity and 96% coverage), Phrux (GenBank accession number NC_021309.1) (99.20% identity and 95% coverage), and ShereKhan (GenBank accession number MH513983.1) (99.10% identity and 95% coverage).JeTaime is an E-cluster phage with 142 predicted protein-coding genes (54 assigned functions), while Luna22 is a Q-cluster phage with 87 predicted protein-coding genes (42 assigned functions) . Intereslignment  revealedMT114159) and New Zealand  (GenBank accession number MK494095) retain >98% nucleotide sequence identity to the Luna22 genome. Also, the genome of Giles (GenBank accession number EU203571) is nearly identical to that of Luna22 (99.96% identity and 100% coverage)  \u201321; the Individual GenBank and SRA numbers are listed in"}
+{"text": "Defining the metabolic syndrome (MetS) in children remains challenging. Furthermore, a dichotomous MetS diagnosis can limit the power to study associations. We sought to characterize the serum metabolite signature of the MetS in early childhood using high-throughput metabolomic technologies that allow comprehensive profiling of metabolic status from a biospecimen.In the Family Atherosclerosis Monitoring In earLY life (FAMILY) prospective birth cohort study, we selected 228 cases of MetS and 228 matched controls among children age 5 years. In addition, a continuous MetS risk score was calculated for all 456 participants. Comprehensive metabolite profiling was performed on fasting serum samples using multisegment injection-capillary electrophoresis-mass spectrometry. Multivariable regression models were applied to test metabolite associations with MetS adjusting for covariates of screen time, diet quality, physical activity, night sleep, socioeconomic status, age, and sex.Compared to controls, thirteen serum metabolites were identified in MetS cases when using multivariable regression models, and using the quantitative MetS score, an additional eight metabolites were identified. These included metabolites associated with gluconeogenesis (glucose (odds ratio (OR) 1.55 [95% CI 1.25\u20131.93]) and glutamine/glutamate ratio (OR 0.82 [95% CI 0.67\u20131.00])) and the alanine-glucose cycle ), amino acids metabolism (tyrosine (OR 1.33 [95% CI 1.10\u20131.63]), threonine (OR 1.24 [95% CI 1.02\u20131.51]), monomethylarginine (OR 1.33 [95% CI 1.09\u20131.64]) and lysine (OR 1.23 [95% CI 1.01\u20131.50])), tryptophan metabolism (tryptophan (OR 0.78 [95% CI 0.64\u20130.95])), and fatty acids metabolism (carnitine (OR 1.24 [95% CI 1.02\u20131.51])). The quantitative MetS risk score was more powerful than the dichotomous outcome in consistently detecting this metabolite signature.A distinct metabolite signature of pediatric MetS is detectable in children as young as 5\u2009years old and may improve risk assessment at early stages of development.The online version contains supplementary material available at 10.1186/s12916-021-02162-7. Obesity and related metabolic sequelae such as type 2 diabetes are increasingly prevalent among children in high-income countries , 2. The N-oxide or TMAO) Additional file 2. Supplemental Methods [IDEFICS MetS definitions and MSI-CE-MS data processing]Additional file 3: Table S1. [Serum metabolites characteristics]Additional file 4: Table S2. [Unadjusted and intermediate adjusted models]Additional file 5: Table S3. [Pathway analysis]Additional file 6: Fig. S2. [Clustering of MetS cases]"}
+{"text": "Orgyia postica. The complete mitochondrial genome was a circular molecule with 15,258\u2009bp in full-length, including 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs) and 2 ribosomal RNA genes (rRNAs). The nucleotide composition of O. postica mitogenome was A: 37.6%, C: 7.9%, G: 14.8%, T: 39.7%. In addition, the results of the phylogenetic analysis indicate that O. postica had the closest relationship with Laelia suffusa. Our study will contribute to further research on the evolution of Lymantriidae and the identification of larva species.The high-throughput sequencing method was used for the first time to determined complete mitochondrial genome of  Orgyia postica Walker, 1855 (Lepidoptera: Lymantriidae) is a crop pest that mainly damages stamens and leaves, It mainly damages more than 70 crops such as tea, cotton, strawberries, loofah, asparagus, radishes, peaches, grapes, pears, citrus, mangoes, causing poor growth, reduced production and even death . The specimen was deposited at the Guangxi Key Laboratory of Agric-Environment and Agric-Products Safety  (Voucher specimen number: FDSW202318382-1r) and identified by the head of herbarium Mr Li (Email: lijunlijun1981@163.com). Genomic DNA was extracted using Genomic DNA Kit  and constructed the libraries with an average length of 350\u2009bp using the NexteraXT DNA Library Preparation Kit . Then the libraries were sequenced on Illumina Novaseq 6000 platform, 9.23\u2009Gb clean data was assembled by SPAdes (version 3.11.0) . The original sequencing data were uploaded to the NCBI database (https://www.ncbi.nlm.nih.gov/sra/) with the accession number SRR15041082. The complete mitochondrial genome was assigned Genbank accession number MW355619.The sample of O. postica is a closed circular molecule of 15,258\u2009bp, consisting of complex I (NADH dehydrogenase), complex IV (cytochrome C oxidase), ATP synthase, transfer RNAs, ribosomal RNAs and other genes, with the base content of A 37.6%, C 7.9%, G 14.8%, and T 39.7%. The A\u2009+\u2009T content is 77.3%, showing a strong AT skew. The genome encodes 37 genes, including 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), and 2 ribosomal RNA genes (rRNAs). With the gene rrnS as a starting point, the gene order of complete mitochondrial is rrnS, trnV-TAC, rrnL, trnL-TAG, ND1, trnS-TGA, CYTB, ND6, trnP-TGG, trnT-TGT, ND4L, ND4, trnH-GTG, ND5, trnF-GAA, trnE-TTC, trnS-GCT, trnN-GTT, trnR-TCG, trnA-TGC, ND3, trnG-TCC, COX3, ATP6, ATP8, trnD-GTC, trnK-CTT, COX2, trnL-TAA, COX1, trnY-GTA, trnC-GCA, trnW-TCA, ND2, trnQ-TTG, trnI-GAT, trnM-CAT. Among these genes, the shortest and longest were 62 (trnR-TCG) and 1692\u2009bp (ND5), and the average gene length was 390.78\u2009bp.The mitochondrial genome of O. postica within lepidoptera. 21 published complete mitochondrial genome sequences from lepidoptera including Euproctis pseudoconspersa (KJ716847.1), Euproctis cryptosticta (KY996558.1), Anarta trifolii (MN715147.1), Sesamia inferens (JN039362.1), Spodoptera littoralis (MT816470.1), Spodoptera litura (JQ647918.1), Spodoptera frugiperda (KM362176.1), Spodoptera exigua (JX316220.1), Spodoptera exempta (MT906792.1), Parasa consocia (KX108765.1), Aglaomorpha histrio (KY800518.1), Spilarctia subcarnea (KT258909.1), Lemyra melli (KP307017.1), Hyphantria cunea (GU592049.1), Amata formosae (KC513737.1), Euproctis similis (KT258910.1), Euproctis seitzi (MN916588.1), Lymantria dispar (FJ617240.1), Laelia suffusa (MN908152.1), Lymantria umbrosa (KY923066.1) and Lymantria sugii (MT265380.1) species were collected and aligned with O. postica by MAFFT7.037 (Katoh and Standley, L. suffusa has the closest relationship with O. postica.To confirm the phylogenetic position and understand the relationship of"}
+{"text": "Women aged 65 and older experience nearly three-fourths of the 2 million osteoporotic fractures annually in the US, yet whether accelerometer-measured volumes and intensities of physical activity and sedentary behavior (SB) are associated with reduced fracture risk is understudied. We investigated associations of accelerometer-measured light physical activity (LPA), moderate-to-vigorous physical activity (MVPA), sedentary time (ST), and mean sedentary bout duration (MBD) with incident clinical fractures  in the WHI OPACH cohort. Participants  without prior hip fracture wore the ActiGraph GT3X+ for 7 days between May 2012-April 2014 and were followed through March 2020 for incident clinical fracture (N=711). Cox models estimated hazard ratios (HR) and 95% confidence intervals (CI), adjusting for age, race-ethnicity, education, alcohol, smoking, height, weight, falls history, RAND-36 physical function, diabetes, thiazide use, prescription osteoporotic therapy, and age at menopause. The HR(95% CI) across MVPA quartiles was 1.00(reference), 1.15(0.93-1.41), 0.90(0.72-1.13), and 0.79(0.61-1.02); p-trend=0.01. The HR(95% CI) for a one-interquartile range increment in MVPA (42 minutes/day) was 0.86(0.76-0.97). Associations were modified by prescription osteoporotic therapy  and varied in magnitude by age, BMI , and race-ethnicity . LPA, ST, or MBD were not associated with incident fractures. These data suggest that MVPA may reduce and not increase fracture risk and that LPA and SB do not increase fracture risk."}
+{"text": "Scientific Reports 10.1038/s41598-021-81257-w, published online 18 January 2021Correction to: This Article contains an error, where Supplementary Data files Supplementary Data 1.Supplementary Data 2.Supplementary Data 3."}
+{"text": "Monarda (family Lamiaceae) contains 22 species of which three are native to southern Alabama, M. citriodora, M. fistulosa, and M. punctata. Several species of Monarda have been used in traditional medicines of Native Americans, and this present study is part of an ongoing project to add to our understanding of Native American pharmacopeia. Plant material from M. citriodora, M. fistulosa, and M. punctata was collected in south Alabama and the essential oils obtained by hydrodistillation. The essential oils were analyzed by gas chromatographic techniques to determine the chemical compositions as well as enantiomeric distributions. The compounds thymol, carvacrol, p-cymene, and their derivatives were the primary terpenoid components found in the essential oils. The known biological activities of these compounds are consistent with the traditional uses of Monarda species to treat wounds, skin infections, colds, and fevers.The genus  Monarda L. (Lamiaceae) species, 18 of which occur in the United States [Monarda species native to south Alabama, namely Monarda citriodora Cerv. ex Lag., Monarda fistulosa L., and Monarda punctata L.  and carvacrol (RIdb = 1296). The other major components were p-cymene (RIdb = 1024) and thymol methyl ether (RIdb = 1239).The M. citriodora essential oils revealed the (+)-enantiomers to be the major stereoisomers for \u03b1-thujene, \u03b1-pinene, \u03b2-pinene, \u03b1-phellandrene, \u03b4-3-carene, \u03b1-terpinene, cis-sabinene hydrate, trans-sabinene hydrate, \u03b1-terpineol, \u03b1-copaene, (E)-\u03b2-caryophyllene, and germacrene D. On the other hand, the (\u2212)-enantiomer was dominant for \u03b2-phellandrene, borneol, carvone, and \u03b4-cadinene. Limonene showed variation in the enantiomeric distributions with (+)-limonene in 26.7%, 63.3%, and 57.3% for aerial parts #1, #2, and roots essential oils, respectively. Likewise, linalool also showed variation with (+)-linalool of 71.9%, 49.7%, and 50.1%. (+)-Terpinen-4-ol was the predominant enantiomer in the aerial parts essential oils (60.2% and 58.5%), but (\u2212)-terpinen-4-ol (79.1%) was dominant in the root essential oil.Chiral gas chromatography\u2013mass spectrometry (GC-MS) analysis of the Monarda fistulosa essential oils were obtained in 2.66\u20134.83% yields as bright orange oils. The chemical compositions of the essential oils from the aerial parts of M. fistulosa are summarized in db = 1289) dominated the compositions  with lesser quantities of p-cymene , limonene , carvacrol , and thymoquinone . Curiously, sample #3, although qualitatively similar, had a very different quantitative composition with thymoquinone as the most abundant constituent (41.3%) followed by p-cymene (21.9%), but with lower concentrations of thymol (8.9%) and carvacrol (1.6%).M. citriodora essential oils, in M. fistulosa essential oils, the (+)-enantiomer was the major for \u03b1-thujene, \u03b1-pinene, \u03b2-pinene, \u03b1-phellandrene, \u03b4-3-carene, \u03b1-terpinene, cis-sabinene hydrate, trans-sabinene hydrate, \u03b1-terpineol, \u03b1-copaene, (E)-\u03b2-caryophyllene, and germacrene D, while the (\u2212)-enantiomer was predominant for \u03b2-phellandrene and borneol. (\u2212)-Limonene (97.4\u201399.5%) and (\u2212)-linalool (62.1\u201362.5%) dominated in all three M. fistulosa samples. (+)-Camphene (100%), (+)-sabinene (58.4\u201359.0%), and (+)-terpinen-4-ol (63.2\u201363.3%) were also dominant.As was observed in M. punctata aerial parts gave bright orange essential oils in 0.781% and 0.658% yield. The most abundant components in the essential oils were thymol , p-cymene , \u03b3-terpinene , and carvacrol  (see Hydrodistillation of two samples of wild-growing .1%) see .M. punctata essential oils were analogous to those observed for M. citriodora and M. fistulosa oils with the exception of limonene, which was virtually racemic in sample #1, but 100% (\u2212)-limonene in sample #2.The enantiomeric distributions of terpenoids in Monarda citriodora and M. fistulosa have been introduced throughout temperate regions of the world as popular herbal medicines as well as ornamentals [M. citriodora in the present study were rich in both thymol and carvacrol, whereas essential oils from Europe and Asia were dominated by thymol with much lower concentrations of carvacrol. Monarda fistulosa, in particular, showed wide variation with at least three different chemotypes  in this study fit into the thymol-rich chemotype. Interestingly, there was a high concentration of thymoquinone in M. fistulosa sample #3, with concomitant lower concentrations of thymol and carvacrol. Thymol was reported as the major component of M. punctata in two old reports [M. punctata from China was rich in thymol (75.2%), which is in agreement with the aerial parts essential oils from Alabama.amentals ,5,6. The reports ,12. Consp-cymene are consistent with the traditional uses of Monarda spp. to treat skin infections, wounds, fevers, and respiratory problems. Thymol [p-cymene [The high concentrations of thymol, carvacrol, and . Thymol , carvacr. Thymol , and p-cp-cymene  have demp-cymene ,35, as wp-cymene . Thymol p-cymene  and carvp-cymene , in addip-cymene , have shp-cymene . Furtherp-cymene  and carvp-cymene  have shop-cymene .Monarda species. Several investigations on the enantiomeric distributions in other members of the Lamiaceae have been reported in the literature, however. There seems to be much variation in the enantiomeric distribution of monoterpenoids across the family. Consistent with what was observed in Monarda essential oils, (+)-\u03b1-pinene was the major enantiomer found in Coridothymus capitatus [Rosmarinus officinalis [Lepechinia heteromorpha [Ocimum canum, and Ocimum kilimandscharicum [C. capitatus [Monarda essential oils. On the other hand, (\u2212)-\u03b2-pinene dominates in R. officinalis [Lepechinia mutica [Mentha \u00d7 piperita) and spearmint (Mentha spicata) have shown nearly racemic mixtures of \u03b1- and \u03b2-pinenes [Monarda essential oils. In marked contrast, however, (\u2212)-\u03b1-phellandrene and (+)-\u03b2-phellandrene predominated in L. mutica essential oil [M. fistulosa essential oil, peppermint (M. piperita) and spearmint (M. spicata) essential oils [C. capitatus [O. canum, and O. kilimandscharicum [R. officinalis) essential oil [C. capitatus [Salvia schimperi [Pycnanthemum incanum [O. canum, and O. kilimandscharicum [Lavandula angustifolia [R. officinalis [As far as we are aware, this work presents the first chiral analysis of terpenoid constituents of apitatus , Rosmariicinalis , Lepechiromorpha , Ocimum charicum . Likewisapitatus  as well icinalis  and Lepea mutica . The ess-pinenes . (+)-\u03b1-Ptial oil . (\u2212)-Limial oils  whereas apitatus , O. canucharicum , and a ntial oil . (+)-Linapitatus , Salvia chimperi , Pycnant incanum , O. canucharicum , whereasstifolia  and R. oicinalis .Monarda citriodora was cultivated in Kirkland Gardens, Newville, AL, USA  from seeds . The cultivated Monarda spp. were grown in loamy clayey-sand and fertilized with chicken manure, kelp meal, and bone meal at planting in full sun. The aerial parts of M. citriodora were collected from separate plants on separate occasions . The roots of M. citriodora were obtained from plant #2.Monarda fistulosa was cultivated in Kirkland Gardens, Newville, AL, USA  from seedlings  as above. The aerial parts of three different plant samples were collected on 25 June 2020.Monarda punctata was collected from wild-growing plants near Newville, AL, USA ; the edge of a planted pine forest, disturbed grassland, full/partial sun, sandy-clay soil that had been intentionally burned (prescribed burn) 1.5 years before collection. The aerial parts of two different plants were collected on 1 June 2020.M. citriodora (SKL61820), M. fistulosa (SKL72020), and M. punctata (SKL9620). The Monarda plant materials were allowed to dry in the shade for several days, the air-dried plant materials were pulverized and subjected to hydrodistillation using a Likens-Nickerson apparatus with continuous extraction with dichloromethane (Plants were identified by S.K. Lawson and a voucher specimen of each plant was deposited in the University of Alabama in Huntsville Herbarium (HALA); voucher numbers for omethane .The essential oils were analyzed by gas chromatography\u2013mass spectrometry (GC-MS), gas chromatography with flame ionization detection (GC-FID), and chiral GC-MS as previously reported .n-alkanes. Compounds identified by comparison of the MS fragmentation and retention indices with those in the databases [Shimadzu GCMS-QP2010 Ultra, ZB-5ms GC column, GC oven temperature 50 \u00b0C\u2013260 \u00b0C (2 \u00b0C/min), 1-\u03bcL injection of 5% solution of EO in dichloromethane . Retention indices (RIs) were determined with reference to a homologous series of atabases ,8,9,10.Shimadzu GC 2010, FID detector, ZB-5 GC column, GC oven temperature 50 \u00b0C\u2013260 \u00b0C (2.0 \u00b0C/min). The percent compositions were determined from raw peak areas without standardization.Shimadzu GCMS-QP2010S, Restek B-Dex 325 column, GC oven temperature 50 \u00b0C\u2013120 \u00b0C (1.5 \u00b0C/min) then 120 \u00b0C\u2013200 \u00b0C (2.0 \u00b0C/min), 0.1 \u03bcL injection of 5% solution of EO in dichloromethane . The enantiomeric distributions were determined by comparison of retention times with authentic samples obtained from Sigma-Aldrich . Relative enantiomer percentages were calculated from peak areas.Monarda growing in south Alabama. In addition, the enantiomeric distribution of terpenoids was also carried out. This work illustrates the wide variation in essential oil compositions based on geographical location as well as variations in enantiomeric distribution. It would be interesting to compare enantiomeric distributions for Monarda essential oils from other geographical locations and for other Monarda species. Nevertheless, the phenolic monoterpenoids thymol and/or carvacrol were found to dominate the compositions of M. citriodora, M. fistulosa, and M. punctata and support the traditional medicinal uses of these plants.This study presents, for the first time, analyses of the essential oils of three species of"}
+{"text": "Ferrigenium kumadai An22T (= JCM 30584T = NBRC 112974T = ATCC TSD-51T) is a microaerophilic iron oxidizer isolated from paddy field soil and belongs to the family Gallionellaceae. Here, we report the complete genome sequence of F. kumadai An22T, which was obtained from the hybrid data of Oxford Nanopore long-read and Illumina short-read sequencing. Ferrigenium kumadai, isolated from paddy soil has been described (F. kumadai An22T (= JCM 30584T = NBRC 112974T = ATCC TSD-51T).Microaerophilic iron-oxidizing bacteria, which are capable of oxidizing ferrous iron (Fe) under circumneutral pH and microoxic conditions, are a key player in the Fe redox cycle in environments . Howeverescribed . Here, wT and DNA preparation were described previously  were removed with Sickle v1.33 (https://github.com/najoshi/sickle). Short ONT reads  were filtered out with Filtlong v0.2.0 (https://github.com/rrwick/Filtlong), and then error correction of the reads was carried out on FMLRC v0.1.2 (Betaproteobacteria. Genes were annotated with DFAST v10 (Sideroxydans lithotrophicus ES-1 (GenBank accession number CP001965.1), Rhodopseudomonas palustris TIE-1 (CP058907.1), Mariprofundus ferrooxydans PV-1T (DS022294.1), and Acidithiobacillus ferrooxidans ATCC 23270T (CP001219.1). An orthologous gene cluster table among the genome sequences used was created  was found as a sole candidate gene of Fe(II) oxidation.The genome size of strain An22F. kumadai An22T was deposited to the DDBJ database under the accession number AP019536.1. The raw sequencing data were deposited under BioProject accession number PRJDB7995 and SRA accession numbers DRR168795 and DRR168796.The genome sequence of"}
+{"text": "Elevated resistance rates have been reported in ICUs. Aztreonam (ATM) combined with avibactam (AVI) is being developed for use against drug-resistant Enterobacterales (Ebact), including metallo-\u03b2-lactamase (MBL)-positive isolates. We examined the activity of ATM-AVI and comparators against Ebact isolates collected from geriatric patients in ICU and non-ICU wards as part of the ATLAS surveillance program.Escherichia coli, Klebsiella spp. and Proteus mirabilis with ATM or ceftazidime MIC >1 \u00b5g/mL.23754 non-duplicate Ebact isolates were collected in 53 countries in Asia/Pacific (excluding mainland China and India), Europe, Latin America, and Middle East/Africa from patients \u226565 years with lower respiratory tract (LRTI), urinary tract (UTI), skin and soft tissue (SSTI), intra-abdominal (IAI), and bloodstream (BSI) infections. Susceptibility testing was performed by CLSI broth microdilution and values interpreted using CLSI 2021 breakpoints. PCR and sequencing were used to determine the \u03b2-lactamase genes present in isolates with meropenem MIC >1 \u00b5g/mL, and 90 values were up to 32-fold higher. ATM-AVI MIC90 values were within one doubling-dilution across all studied strata (0.12-0.25 \u00b5g/mL), were comparable to or lower than for meropenem in all strata, and were 2 to \u22659 dilutions lower than all other tested comparators. MBL-positive Ebact were found in 1.5% of LRTI (n=91), 1.2% of UTI (n=70), 1.1% of SSTI (n=52), 1.3% of BSI (n=49), and 0.7% of IAI isolates (n=22). MBL-positive rates were higher among ICU  than non-ICU isolates . ATM-AVI MIC90 values were 0.5 \u00b5g/mL against MBL-positive isolates from all ward and infection types except SSTI (MIC90 0.25 \u00b5g/mL) and BSI (MIC90 1 \u00b5g/mL), 2-4 dilutions lower than tigecycline and at least 5-10 dilutions lower than the other comparators.Susceptibility of the studied comparator agents was generally slightly lower among Ebact from BSI than other infection types (Table). Susceptibility was also generally lower among Ebact from ICU than non-ICU wards by up to 10 percentage points, and MICResults TableATM-AVI could provide a valuable therapeutic option for treatment of infections caused by Ebact in patients \u226565 years old in both ICU and non-ICU wards.Sibylle Lob, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Krystyna Kazmierczak, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Francis Arhin, PhD, Pfizer, Inc. (Employee) Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "Rhizopus species is the primary fungi responsible for 70% of mucormycosis cases.During the second wave of the COVID-19 pandemic, an unusual increase in cases of mucormycosis was observed in India, owing to immunological dysregulation caused by the SARS-CoV-2 and the use of broad-spectrum antibiotics, particularly in patients with poorly controlled diabetes with ketoacidosis to have contributed to the rise, and it has been declared an epidemic in several states of India. Because of the black colouring of dead and dying tissue caused by the fungus, it was dubbed \"black fungus\" by several Indian media outlets. In this study, attempts were taken to unmask novel therapeutic options to treat mucormycosis disease. Rhizopus delemar such as CotH3, Lanosterol 14 alpha-demethylase and Mucoricin which plays a crucial role in the virulence of Mucorales. Initially, we explored the physiochemical, structural and functional insights of proteins and later using AutoDock Vina, we applied computational protein\u2013ligand binding modelling to perform a virtual screening around 300 selected compounds against these three proteins, including FDA-approved drugs, FDA-unapproved drugs, investigational-only drugs and natural bioactive compounds. ADME parameters, toxicity risk and biological activity of those compounds were approximated via in silico methods. Our computational studies identified six ligands as potential inhibitors against Rhizopus delemar, including 12,28-Oxamanzamine A, vialinin B and deoxytopsentin for CotH3; pramiconazole and saperconazole for Lanosterol 14 alpha-demethylase; and Hesperidin for Mucoricin. Interestingly, 12,28-Oxamanzamine A showed a maximum binding affinity with all three proteins .We chose three important proteins from the In summary, our investigation identified 12,28-Oxamanzamine A, vialinin B, deoxytopsentin, pramiconazole, saperconazole and hesperidin as potent bioactive compounds for treating mucormycosis that may be considered for further optimisation techniques and in vitro and in vivo studies.The online version contains supplementary material available at 10.1186/s42269-022-00704-4. Rhizopus species are the most common fungi in the order of Mucorales responsible for over 70% of mucormycosis cases  and diabetic ketoacidosis. SARS-Cov-2, in addition to mucormycosis, is a fatal combination that has caused a considerable number of deaths, particularly in India  acts as a fungal ligand for host cell GRP78 and mediates pathogenic host-cell interactions. The presence of CotH3 in Mucorales also explained why DKA patients with high GRP78 levels are more susceptible to mucormycosis  require a thorough examination of their structure and function, which will bring unique insights into the development of an effective, low-cost medicine with minimal side effects. Therefore, the current study aims to collect 300 compounds  that exhibit antiviral, antifungal, antibacterial and antimicrobial properties have been identified through different literature reviews, and it was screened against CotH3, Lanosterol 14 alpha-demethylase and Mucoricin by applying several in silico tools, viz., protein modelling, binding pocket prediction, molecular docking, ADME and drug-likeness screening, bioactivity prediction and toxicity prediction , cytochrome P450 enzyme Lanosterol 14 alpha-demethylase (ACCESSION: EIE87079) and ricin-like toxin Mucoricin (ACCESSION: EIE81863) . Various physicochemical properties of the CotH3, Lanosterol 14 alpha-demethylase and Mucoricin were calculated using ExPasy's ProtParam tool   NPS@ .All three proteins  were subjected to 3D modelling. CotH3 was modelled via SWISS-MODEL  3.0 was used to predict probable binding pockets of the proteins , part of MGLTools  program  alpha helix, 13.95% (36 residues) extended strand, 2.71% (7 residues) beta turn and 43.02% (111 residues) random coil, while Lanosterol 14 alpha-demethylase showed to have 49.80% (254 residues) alpha helix, 10.78% (55 residues) extended strand, 3.14% (16 residues) beta turn and 36.27% (185 residues) random coil. Similarly, Mucoricin exhibited 5.44% (8 residues) alpha helix, 40.14% (59 residues) extended strand, 14.97% (22 residues) beta turns and 39.46% (58 residues) random coil.Bacillus cereus CotH kinase (PDB ID: 5JD9) \u2009=\u20090.02\u00a0nM) and acts as a promising anti-allergic agent . The compound displayed potent low nanomolar inhibitory activity against MRSA PK with significant concomitant selectivity over human PK orthologues -dien-3, \u03b2-6\u03b1-diol and 14\u03b1-methylergosta-5,7,22,24(28)-tetraenol , Parsiguine (B), Haliclonacyclamine B (C), Vialinin B (D), 6-Deoxymanzamine X (E), Natamycin (F), Olorofim (G), Deoxytopsentin (H), Manzamine E (I), Fascioquinol A (J). Figure S2. 2D visualisation of docking analysis of Lanosterol 14 alpha-demethylase binding with Pramiconazole (A), 12,28-Oxamanzamine A (B), Fascioquinol D (C), Saperconazole (D), Nakadomarin A (E), Plakinamine A (F), Fascioquinol C (G), Parsiguine (H), Hesperidin (I), Epoxyazadiradione (J). Figure S3. 2D visualisation of docking analysis of Mucoricin binding with 12,28-Oxamanzamine A (A), Manzamine A (B), Parsiguine (C), Halicyclamine A (D), Tetrahydrohaliclonacyclamine A (E), Phaeosphenone (F), 6-Deoxymanzamine X (G), Goniodomin A (H), Hesperidin (I), Stelletin A (J).Additional file 2: Table S1. The variants of mucormycosis and the symptoms associated with the disease .Additional file 3: Data 1. Binding affinities of all bioactive compunds with our three target proteins."}
+{"text": "Literature on SARS-CoV-2 infection in cancer patients is scarce in Latin America. This population seems to have a higher risk for adverse outcomes. This study aims to correlate clinical characteristics with outcomes in patients with cancer in a referral center in Mexico.We included patients with cancer and confirmed SARS-CoV-2 infection, from April, 19 to December 30, 2020, at the Instituto Nacional de Cancerolog\u00eda, Mexico. Clinical information was obtained from medical and epidemiological records. We conducted a descriptive analysis. For the association between variables with hospitalization, invasive mechanical ventilation (IMV), and mortality; univariate and multivariate logistic regression was performed; odds ratios and 95% confidence intervals were calculated.Four hundred thirty-three patients were included; 268 (62%) were female, the median age was 55 years. One hundred thirty-five (31%), 130 (30%), and 93 (21%) patients had obesity, hypertension, and diabetes mellitus (DM), respectively. Three hundred forty-one (79%) had solid cancer; 82 (19%) hematological malignancy (HM), and 10 (2%) were under evaluation for cancer diagnosis. One hundred seventy (39%) had advanced or metastatic cancer. One hundred ninety-eight (46%) patients were hospitalized. Risk factors were: age (p= 0.001); woman (p=0.019); HM (p=0.050) and advanced or metastatic cancer (p= 0.041). Fourty-five (10%) patients required IMV. Age (p=0.018); DM (p=0.041); C-Reactive Protein (p= 0.002), and LDH (p= 0.033) were associated with invasive mechanical ventilation. Mortality within 30-days after diagnosis was 19% (82 cases). Associated characteristics were: age (p=0.041); lymphocytes (p=0.049); creatinine (p=0.005) and albumin (p=0.001).In this study, patients with cancer showed higher mortality, need of hospitalization, and invasive mechanical ventilation compared with groups of patients without cancer. We did not find an increased risk in mortality for hematological malignancies. Although our cohort was younger than others previously reported, age was a strong predictor of adverse outcomes. Variables associated with IMV and death were similar to those previously described in cancer patients with COVID-19.All Authors: No reported disclosures"}
+{"text": "Despite well-established guidelines for urinary tract infection (UTI) treatment, prescribing practices vary. We examined the association between inappropriate (IA) or suboptimal (SO) antibiotic (AB) prescribing (RX) and hospitalization, healthcare resource use (HRU), and costs among patients with uncomplicated UTI (uUTI) in the US.Table 1) were assessed from Optum claims data during index episode (within 28 days of index) and 12-month follow-up.This retrospective cohort study used linked Premier Healthcare/Optum claims data from female outpatients (\u2265 12 years old) with a uUTI diagnosis . Patients with complicated UTIs  were excluded. HRU and costs between patients with IA/SO and appropriate and optimal (AP&OP) AB RX (defined in Table 1. Definitions of appropriateness of AB RXTable 2). During index episode, mean ambulatory care and pharmacy claims were significantly higher for IA/SO versus AP&OP AB RX , and total HRU cost per patient was higher for IA/SO (&2616) versus AP&OP AB RX . During follow-up, 267 (9.7%) patients with IA/SO AB RX had a UTI-related emergency department (ED) visit versus 202 (6.5%) patients with AP&OP AB RX (p < 0.001). Mean UTI-related HRU costs were significantly higher for IA/SO (&5048) versus AP&OP AB RX . After adjusting for patient characteristics, patients with IA/SO AB RX were 40% more likely than those with AP&OP AB RX to have a UTI-related ED visit  during follow-up (Table 3). Adjusted HRU costs for IA/SO AB RX (vs AP&OP) were numerically higher for index uUTI episode (by &1772), and UTI-related (by &1102) and all-cause (by &1528) charges during follow-up .Of 5870 patients, 1856 (31.6%) had IA and 1255 (21.4%) had SO AB RX. Patients with IA/SO AB RX (47.1%) were older and more likely to have a Charlson Comorbidity Index score > 0 than those with AP&OP AB RX , stratified by appropriateness of AB RX at index and during follow-upIA/SO AB RX was associated with higher overall and UTI-related HRU and costs during index episode and 12-month follow-up, highlighting a need for education on applying prescription guidelines and the use of culture-based RX.Rena Moon, MD, Premier Applied Sciences, Premier Inc. (Employee) Alen Marijam, MSc, GlaxoSmithKline plc.  Fanny S. Mitrani-Gold, MPH, GlaxoSmithKline plc.  Daniel C. Gibbons, PhD, GlaxoSmithKline plc.  Alex Kartashov, PhD, Premier Applied Sciences, Premier Inc. (Employee) Ning Rosenthal, MD, Premier Applied Sciences, Premier Inc.  Ashish V. Joshi, PhD, GlaxoSmithKline plc."}
+{"text": "Oncogene (2020) 39:5616\u20135632, 10.1038/s41388-020-01388-8, published online 13 July 2020Correction to: Following the publication of the above article, the authors noted two errors in Fig. p\u2009<\u20090.05, ** p\u2009<\u20090.01.Supplemental Fig. 2: Overexpression of LINC01503 promotes NPC cell growth, migration, and invasion in vitro. (a) Relative expression of LINC01503 in 5-8F and HONE1 cells transfected with LINC01503-expressing plasmid and empty vector. (b) LINC01503 overexpression promoted cell growth of 5-8F and HONE1 cells as shown by CCK-8 assays. (c) LINC01503 overexpression facilitated cellular survival effects as evaluated by colony formation assays. (d) LINC01503 overexpression accelerated the movement of 5-8F and HONE1 cells as assessed by wound healing assays. Scale bar, 100 \u03bcm. (e) LINC01503 overexpression promoted the migration and invasion ability of 5-8F and HONE1 cells as determined by transwell assays. Scale bar, 100\u2009\u03bcm. * The original article has been corrected.Supplementary Fig. 2"}
+{"text": "Hemophagocytic lymphohistiocytosis (HLH) is a disease that can affect both children and adults. HLH can be categorized as primary or secondary. Secondary HLH (sHLH) may be secondary to various viral infections. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus infection is a pandemic with multi-system involvement. HLH in COVID-19 positive patients is a recognized entity. However, in post-COVID-19 patients who have recovered and are negative by serological tests and reverse transcription-polymerase chain reaction test may present with sHLH due to dysregulation of the immune system. We highlight this unusual finding of post-COVID-19 sHLH in two cases, who were diagnosed by the new revised H-score. Hemophagocytic lymphohistiocytosis (HLH) is a lethal disorder of varying etiology and encompasses a wide range of diseases. Familial HLH (FHLH) and immune-related HLH constitute the primary HLH spectrum whereas infections, drug-related and transplantation, malignancies, and macrophage activation syndrome (MAS) constitute the secondary HLH (sHLH) spectrum . LaboratCase 12 was 88%, and the temperature was 40\u030aC. On examination, pallor was present. Bilateral crepitations and rhonchi were present in respiratory system auscultation. The cardiovascular system was unremarkable and central nervous system examination showed no focal neurological deficit. Ultrasonography of the whole abdomen revealed hepatosplenomegaly. High-resolution computed tomography (HRCT) thorax showed CO-RADS category 6 and CT severity of 15/25. Bone marrow aspirate showed evidence of hemophagocytosis  and was hospitalized for 1.5 months. At presentation, the patient was conscious, alert, and oriented.\u00a0Her blood pressure was 96/58 mmHg, pulse rate was 118 beats per minute, respiratory rate was 33/min and SPO3 /\u00b5L with differential leukocyte count (DLC) of N87L10M03E0, and platelet count of 20x103 /\u00b5L. Biochemical investigations revealed serum urea of 61 mg/dL (reference range [RR]: 17-45 mg/dL), serum creatinine of 0.9 mg/dL (RR: 0.6-1.4 mg/dL), D-dimer of 1.58 mg/L (RR: 0-0.5 mg/L), aspartate aminotransferase (AST) of 76 IU/L (RR: 5-40 IU/L), alanine aminotransferase (ALT) of 98 IU/L (RR: 5-55 IU/L), alkaline phosphatase (ALP) of 137 IU/L (RR: 20-140 IU/L), total bilirubin of 2.5 mg/dL (RR: up to 1.2 mg/dL), direct bilirubin of 1.3 mg/dL (RR: 0-0.2 mg/dL), IL-6 of 11.56 pg/mL (RR: 5-15 pg/mL), fibrinogen of 2.3 g/L (RR: 2-4 g/L), vitamin B12 of 489.5 pg/mL (RR: 160-950 pg/mL), ferritin of 170.2 ng/L (RR:12-250 ng/L), and C-reactive protein of 77mg/L (RR: 0-10 mg/L). Direct Coombs test was negative and indirect cold antibodies titer was more than 256 (RR: less than 164). All viral markers, anti-nuclear antibody (ANA), and glucose-6-phosphate dehydrogenase (G6PD) were negative. The H-score estimated as per the H-score of 2014 was 213 points with a probability of sHLH to be 93% to 96%  of 3.3 g%, total leukocyte count (TLC) of 18.3x10According to the protocols of our institute, the patient was isolated and started on intravenous steroids (dexamethasone acetate 10 mg/day), low molecular weight heparin (LMWH), antibiotics, and oxygen support. Considering the deranged kidney function, strict monitoring and adequate hydration were provided along with four units of whole blood transfusion. The patient developed congestive cardiac failure during the hospital stay and oral diuretics were initiated. The patient responded to treatment without the addition of etoposide and was hemodynamically stable. She was discharged after four weeks of hospital stay. The patient was reviewed after 14 days and was stable.Case 23 /\u00b5L with DLC N86L10M02E02, and platelet count of 60x103 /\u00b5L. Biochemical investigations showed serum ferritin of 188.0 ng/mL (RR: 7-140 ng/mL), IL-6 of 1,057 pg/mL (RR: 1-5 pg/mL), D-dimer of 1.76 mg/L (RR: 0-0.5 mg/L), AST of 182 IU/L (RR:10-40 IU/L), ALT of 77 IU/L (RR:10-40 IU/L), ALP of 77 IU/L (RR: <350 IU/L), C-reactive protein of 62.1 mg/L (RR: 0.02-14.5 mg/L), and fibrinogen of 2.13 g/L (RR: 2-4 g/L). Malaria, scrub typhus, HBsAg, anti-HCV were negative. Bone marrow aspirate revealed features of hemophagocytosis with increased iron , diarrhea, and vomiting for two days. The patient had no previous episodes in the past and developmental milestones were achieved at his age normally. On examination of muscle tone, neck control was decreased. A possible diagnosis of post-viral encephalitis was considered. The patient was positive for SARS-CoV-2 virus by RT-PCR and was admitted to COVID-19 designated hospital. He was discharged after testing negative for the SARS-CoV-2 virus.\u00a0After two weeks of discharge, he presented with the above symptoms but was negative for COVID-19 by RAT and RT-PCR. MRI brain showed features suggestive of viral encephalitis with atrophy of cerebral hemispheres and peritrigonal white matter hyperintensities. Laboratory findings revealed Hb of 7.1 g%, TLC of 7.3x10The H-score estimated as per the H-score of 2014 was 239 points with a probability of sHLH to be 98% to 99%\u00a0, laboratory , and histological parameters , which was modified in 2004 . The HLHThe optimal cut-off for a diagnosis of HLH was considered to be 169 by Fardet et al. with 93% sensitivity and 86% specificity . In our HLH in COVID-19 positive patients is reported in various case reports ,15. HoweSARS-CoV-2 infection is a recent pandemic with important complications inherent to its multisystemic affections.\u00a0Recent information\u00a0about etiopathogenesis, clinical signs\u00a0and symptoms, and treatment protocols are getting highlighted. sHLH is a condition primarily occurring in COVID-19 positive patients; however, sHLH in post-COVID-19\u00a0patients is rare. Immune dysregulation following infection by the SARS-CoV-2 virus may be the prime reason for sHLH in such patients. We highlighted two such post-COVID-19\u00a0patients who were negative for COVID-19 at the time of presentation but were diagnosed as cases of sHLH based on the H-score."}
+{"text": "Otostigmus (O.) lewisiO. (O.) beroni Lewis, 2001. The latter was also recorded from Jilong County  by O. (O.) beroni from Jilong County with new materials of O. (O.) lewisi from Jiacha County, we reaffirm that O. (O.) lewisi is a valid species. Otostigmus Porat, 1876 currently comprises about 56 species (O. (O.) aculeatus Hasse, 1887, O. (O.) astenus , O. (O.) beroni Lewis, 2001, O. (O.) lewisi Song, Gai, Song & Zhu, 2005, O. (O.) martensi Lewis, 1992, O. (O.) politus Karsch, 1881, O. (O.) scaber Porat, 1876, and O. (O.) xizangensis Niu, Li & Di, 2021.The subgenus  species . Among t species : O. (O.)Otostigmus (O.) lewisi was identified as a synonym of O. (O.) beroni (O. (O.) lewisi has complete paramedian sutures on sternites 4\u201319, sternite 21 has a shallow medial longitudinal depression, and the coxopleural process is short. However, the sternite paramedian sutures in O. (O.) beroni are incomplete, and the coxopleural process is long. These differences were not shown in the corresponding figures of O.lewisi is a junior subjective synonym of O.beroni\u201d. We would also like to highlight that all type materials used in the O. (O.) lewisi were sub-adults.) beroni . AccordiMHBU .Studied materials were collected by hand and preserved in 75% ethanol. All studied materials in this paper were examined under a stereomicroscope (Motic K700). Photographs and measurements were taken using a Leica Stereomicroscope (M205 A). The standard terminology followed Otostigminae Kraepelin, 1903Subfamily Otostigmus Porat, 1876Genus Taxon classificationAnimalia\ufeffLewis, 200160ED176E-3416-5BCE-9AFD-79035CE69070Otostigmusberoni Lewis, 2001: 22; MHBU-SoJL1608050101\u2013Ar.-MHBU-SoJL1608050120: Jilong Town, Jilong (Gyirong) County, Xizang (Tibet), China, 28.4370\u00b0N, 85.2568\u00b0E, 5/8/2016, leg. Zhiyong Di. Ar.-MHBU-SoJL21080101: Jilong Town, Jilong (Gyirong) County, Xizang (Tibet), China, 28.4350\u00b0N, 85.2570\u00b0E, 1/8/2021, leg. Zhiyong Di. Housed in MHBU.Ar.-Maximum length 57 mm. Antennae 18 articles, the basal 2.25 to 2.40 glabrous dorsally Fig. . With 4 MHBU-SoJL1608050101). Length.Pigmentation : body color yellow-brown; legs and antennae yellow; cephalic plate, tergites 1\u20132, tergites 20\u201321 and penultimate and ultimate legs blue-green : cephalic plate blue-green, tergites blue-brown, penultimate legs and ultimate legs yellow with blue middle part in each segment, the other legs yellow : with complete paramedian sutures from 3 to 20, marginate from 7 to 21 : smooth, with incomplete paramedian sutures from 5 to 19 : L1\u201316 and L18\u201319 with 2 tarsal spurs, L1\u20134, left L5 and right L6 with 1 tibial spur and L1 with 1 femoral spur.Ultimate prefemur with 4 ventro-lateral, 2 ventro-medial, 3 medial, 2 dorso-medial spines and 1 corner spine Fig. .There are multiple differences among individuals as addressed below. The number of teeth of forcipular tooth plates 4+4 (11 specimens), 5+5 (6 specimens), 3+3 (2 specimen) or 3+4 (2 specimens). Coxopleural process with 4\u20137 spines . One tibial spur on L1\u20134 (8 specimens), L1\u20135 (5 specimens), L1\u20132 (2 specimens), L1\u20133 (3 specimens), or L1\u20136 (2 specimens). Two tarsal spurs on 1 to 18 or 19 pairs of legs; 1 tarsal spur on subsequent to penultimate legs. Ultimate legs without tarsal spur. Ultimate legs prefemur with 11\u201314 spines   and Nepal Fig. .Taxon classificationAnimalia\ufeffSong, Gai, Song & Zhu, 2005 80F267B0-9D5B-5FC6-A29B-D11CC060ACBEOtostigmuslewisiMHBU-SoJC1908060301\u2013 Ar.-MHBU-SoJC1908060307: Jiacha County (Gyaca County), Xizang (Tibet), China, 29.0857\u00b0N, 92.3430\u00b0E, 6/8/2019, leg. Zhiyong Di. Ar.-MHBU-SoJC1608DX01: Jiacha County, Xizang, China, 29.1188\u00b0N, 92.6969\u00b0E, 12/8/2016, leg. Zhiyong Di. Ar.-MHBU-SoJC1608120401: Jiacha County, Xizang, China, 29.1387\u00b0N, 92.6880\u00b0E, 12/8/2016, leg. Zhiyong Di. Housed in MHBU.Ar.-Maximum length 77 mm. Antennae 17\u201320 articles, basal 2.2\u20133 glabrous dorsally Fig. . With 3 O. (O.) lewisi holotype described in The O. (O.) lewisi and O. (O.) beroni; however, the description and figures of O. (O.) lewisi were not consistent in O. (O.) martensi (O. (O.) lewisi (O. (O.) lewisi (O. (O.) martensi (Following O. (O.) lewisi is similar to O. (O.) beroni, and distinguished O. (O.) lewisi from the latter by the length of coxopleural process and the shape of the ultimate sternite. Because O. (O.) lewisi in O. (O.) lewisi is a junior subjective synonym of O. (O.) beroni  beroni .O. (O.) lewisi reported by O. (O.) lewisi described previously were sub-adults.The characteristics of the holotype of MHBU-SoJC1908060301). Length.Pigmentation : cephalic plate and tergites yellow with light green; antennae and legs yellow; penultimate legs and ultimate legs green : antennae light blue mainly, cephalic plate and tergites brownish, penultimate legs and ultimate legs yellow with blue middle part in each segment, the rest of the legs yellow : with complete paramedian sutures from 4 to 20; marginate from 6 to 21 : smooth, with incomplete paramedian sutures from 3 to 4, complete paramedian sutures from 5 to 19 : L2\u20139, right L11 and left L12 with 2 tarsal spurs; 1 tarsal spur on subsequent to penultimate legs; L1 and left L2 with 1 tibial spur and right L1 with 1 femoral spur.The left prefemur with 1 corner spine, 3 ventro-lateral spines, 1 ventro-medial spines, 2 medial spines, 1 dorso-medial spine; the right prefemur with 2 corner spines, 5 ventro-lateral spines, 2 ventro-medial spines, 7 medial spines, 3 dorso-medial spines Fig. .Adult and juvenile individuals differ primarily in body length and pigmentation Figs , 3A, B. Found under stones in arid mountain bush Fig. .China (Xizang) Fig. .Otostigmus, namely, O. (O.) beroni, O. (O.) lewisi, O. (O.) martensi and O. (O.) xizangensis, were reported in Xizang, China  lewisi can be easily distinguished from O. (O.) beroni using the following characteristics: (1) larger average body length: 73 mm in O. (O.) lewisi (average of 2 adults), versus 48 mm in O. (O.) beroni (average of 5 adults); (2) body color (live): primarily brownish cephalic plate and tergites in O. (O.) lewisi, and blue-green cephalic plate and blue-brown tergites in O. (O.) beroni  lewisi 3+3 in 5 specimens or 4+4 in 4 specimens versus O. (O.) beroni 4+4 in 11 specimens or 5+5 in 6 specimens; (4) the number of tibial spurs on legs: O. (O.) lewisi with one tibial spur on legs 1\u20132 in 6 specimens or on legs 1 in 3 specimens. In contrast, O. (O.) beroni with one tibial spur on legs 1\u20134 in 8 specimens or 1\u20135 in 5 specimens; (5) the number of tarsal spurs on legs: O. (O.) lewisi with 2 tarsal spurs on legs 1(2)\u20139(13), with no regularity. In O. (O.) beroni, however, 2 tarsal spurs on legs 1\u201318(19); (6) the length of coxopleural process: coxopleural process of O. (O.) lewisi is shorter than that of O. (O.) beroni. The specimens described by Only four species of the genus ina Fig. , geograponi Fig. ; (3) theO. (O.) beroni and O. (O.) martensi. The main difference between O. (O.) beroni and O. (O.) martensi are as follows: (1) the number of teeth of tooth plates: O. (O.) martensi 3+3, whereas, O. (O.) beroni 4+4 or 5+5; (2) the number of spines on the coxopleural process: O. (O.) martensi coxopleural process with 1 apical spine, 1 lateral spine and 1 dorsal spine. In contrast, O. (O.) beroni coxopleural process with 2\u20133 apical spines, 1\u20132 lateral spines, and 1\u20132 dorsal spines; (3) spurs on legs: O. (O.) martensi L1\u20134 or 5 with 2 tarsal spurs, whereas O. (O.) beroni L1\u201319 with 2 tarsal spurs, L20 with 1 tarsal spur (O. (O.) lewisi and O. (O.) martensi are obviously different species.The most geographically close species of this genus in Xizang, China are sal spur . Given tO. (O.) beroni in China) and Zhangmu Town  martensi in China)) is about 78 km, and the distance between Jilong Town and Jiacha County  lewisi in China) is about 714 km. No species of the genus Otostigmus was found in the area between Jilong Town (or Zhangmu Town) and Jiacha County. The habitat of O. (O.) beroni in Jilong Town is more humid than the habitat of O. (O.) lewisi in Jiacha County  lewisi is a valid species.The distance between Jilong Town (the locality of nty Fig. . Based o"}
+{"text": "Oniscidea) are the most diverse group of troglobionts in caves of continental Portugal. They occur in all karst regions of Portugal, play a major role in decomposition of organic matter in caves and may act as umbrella species for the conservation of all other cave-adapted invertebrates.Terrestrial isopods (We present the IUCN Red List profiles for the cave-adapted terrestrial isopods from continental Portugal, based on recent distribution data from caves. Cave-adapted fauna have a high and global conservationist interest . Cave-adOniscidea) evolved to be the only truly terrestrial crustaceans . The maps were generated in the open source software QGIS 3.14.16, with the layer of the natural protected areas of Portugal (Extent of occurence (EOO) and area of occupancy (AOO) were calculated using the Geospatial Conservation Assessment Tool (GeoCAT) with an approximation to the standard IUCN 2 km \u00d7 2 km cells : Suppl. materials 2Basis of EOO and AOO: Known habitat extent2.Basis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 kmMin Elevation/Depth (m): 380Trichoniscoidesbellesi is known from Algar do Javali Cave in the Montejunto karst massif (Range description: t massif .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Trichoniscoidesbellesi occurs in a single cave : A total of three specimens have been collected in the type locality .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesTrichoniscoidesbellesi was found in the deepest and most thermally insulated parts of the cave, at 10 m depth (Habitat (narrative):  m depth .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 1.7 mm , 1.8 mm Generation length (yr): 1Dependency of single sp?: UnknownTrichoniscoidesbellesi is a blind and depigmented troglobiont species  :  species . It shara, 2012)  and Roncra, 2013 ; an unde\u00f1o, 2010  and a neEucalyptus intensive plantations, which substituted the original native vegetation and is located 50 m from a road, 1.6 km from a quarry and 2.9 km from the closest village.Justification for threats: The cave entrance is surrounded by Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.2. Agriculture & aquaculture - Wood & pulp plantations3.2. Energy production & mining - Mining & quarrying4. Transportation & service corridorsThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.2. Agriculture & aquaculture - Wood & pulp plantations3.2. Energy production & mining - Mining & quarrying4. Transportation & service corridorsJustification for conservation actions: Even though this cave is protected under legislation by the \u201cRede Natura 2000\u201d , this raConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTrichoniscoidesbroteroiScientific name: Species authority: Vandel, 1946AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 3Basis of EOO and AOO: Known habitat extent2.Basis (narrative): The extent of occurrence (EOO) and area of occupancy (AOO) are both 4 kmMin Elevation/Depth (m): 100Trichoniscoidesbroteroi is a troglobiont isopod known from Alqueves Cave, located in the Sic\u00f3 karst area (Range description: rst area . This carst area .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Trichoniscoidesbroteroi is known from a single cave : Alqueves Cave is considered a pre- and proto-historic site. It is a natural funerary cave as bone fragments and other human artifacts were there found, testifying to its human occupation at the end of the Neolithic period, beginning of the Chalcolithic . This caTrend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 4 mm , 4.5 mm Generation length (yr): 1Dependency of single sp?: UnknownTrichoniscoidesbroteroi is a blind and depigmented troglobiont species and a single cave endemic (Porcelliocavernicolus (Ecology and traits (narrative):  endemic . It sharrnicolus .Justification for threats: The entrance to the cave is located within a fully urbanised area, more specifically in the middle of a roundabout . This caThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.3. Residential & commercial development - Tourism & recreation areas4. Transportation & service corridors9.1. Pollution - Domestic & urban waste water9.5. Pollution - Air-borne pollutantsThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.3. Residential & commercial development - Tourism & recreation areas4. Transportation & service corridors9.1. Pollution - Domestic & urban waste water9.5. Pollution - Air-borne pollutantsTrichoniscoidesbroteroi : Suppl. materials 4Basis of EOO and AOO: Known habitat extent2 and the area of occupancy (AOO) is 40 km2.Basis (narrative): The extent of occurrence (EOO) is 217.7 kmMin Elevation/Depth (m): 95Max Elevation/Depth (m): 485Trichoniscoidesmeridionalis is recorded from 10 caves distributed along the Estremenho karst massif: Algar do Vale da Pena, Algar do Z\u00e9 de Braga, Alcobertas, Lapa da Ch\u00e3 de Cima, Moinhos Velhos, Almonda, Papagaio, Algar do Burro, Algar do Pena and Algar do Ladoeiro (Range description: Ladoeiro .EOO (km2): 217.7Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 40Number of locations: 10Trichoniscoidesmeridionalis occurs in ten caves : A total of 43 specimens have been collected: six in Moinhos Velhos cave, five in Algar do Vale do Pena, five in Algar do Burro, one in Algar do Z\u00e9 de Braga, fifteen in Algar do Ladoeiro, ten in Almonda and one in Papagaio Caves .Number of subpopulations: 10Trend: Decline (inferred)Extreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): The caves are located at an altitude ranging from 95 to 485 m a.s.l. . Amonda Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 2 mm , 2.5 mm Generation length (yr): 1Dependency of single sp?: UnknownTrichoniscoidesmeridionalis is a blind, depigmented troglobiont that is adapted to life in the underground (Ecology and traits (narrative): erground . This sperground .Justification for threats: Almonda Cave is located 50 m from a factory that extracts water from a subterranean river and 420 m from a village, which has many agricultural fields. Algar do Ladoeiro's Cave entrance is 840 m from the closest urbanisation. The Moinhos Velhos Cave is the largest show cave of Portugal with around 140,000 visitors per year. Alcobertas Cave has also been subject to structural alterations with the intention to transform it into a show cave during the last century and it is exploited for tourism by a local association . The subThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.1. Agriculture & aquaculture - Annual & perennial non-timber crops3.2. Energy production & mining - Mining & quarrying3.3. Energy production & mining - Renewable energy6.1. Human intrusions & disturbance - Recreational activities9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.1. Agriculture & aquaculture - Annual & perennial non-timber crops3.2. Energy production & mining - Mining & quarrying3.3. Energy production & mining - Renewable energy6.1. Human intrusions & disturbance - Recreational activities9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Almonda Cave is classified, since 1993, as a Property of Public Interest (IIP) and protected due to its archaeological heritage . The arcConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTrichoniscoidesouremensisScientific name: Species authority: Vandel, 1946AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 5Basis of EOO and AOO: Known habitat extent2.Basis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 kmTrichoniscoidesouremensis is only recorded from Lapa da Salgada Cave, located in the F\u00e1tima Plateau, in the eastern part of the Estremenho karst massif (Range description: t massif .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Trichoniscoidesouremensis is recorded from a single cave : Lapa da Salgada Cave is composed of a main underground gallery and the floor is mostly covered by flowstone and clay, with a few bat guano deposits.Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3.5 mm Generation length (yr): 1Dependency of single sp?: UnknownTrichoniscoidesouremensis is classified as a troglobiont species, being blind and depigmented. It is a single cave endemic (Ecology and traits (narrative):  endemic .Justification for threats: The cave entrance is located 270 m from a road in which trucks transport goods from warehouses 600 m away. The cave is also located 1 km away from the closest town.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas4. Transportation & service corridors9.1. Pollution - Domestic & urban waste water9.2. Pollution - Industrial & military effluentsThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas4. Transportation & service corridors9.1. Pollution - Domestic & urban waste water9.2. Pollution - Industrial & military effluentsJustification for conservation actions: Measures need to be put in place in order to protect the habitat and species from the disturbances caused by truck movements and proximity to the urban areas.Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTrichoniscoidesserraiScientific name: Species authority: Cruz, 1993AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 6Basis of EOO and AOO: Known habitat extent2.Basis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 kmMin Elevation/Depth (m): 577Trichoniscoidesserrai is only recorded from Santo Adri\u00e3o Cave, located in the palaeokarst of Vimioso in north-eastern Portugal : 4Trend: UnknownCauses ceased?: UnknownCauses understood?: YesCauses reversible?: YesExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Trichoniscoidesserrai occurs in a single cave : Santo Adri\u00e3o mines are located at 580 m a.s.l. in the Municipality of Miranda do Douro, District of Bragan\u00e7a . The cavTrend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3.6 mm ; 4.2 mm Generation length (yr): 1Dependency of single sp?: UnknownTrichoniscoidesserrai is classified as a blind, depigmented troglobiont, adapted to life in the underground (Ecology and traits (narrative): erground .Justification for threats: The cave entrance is in the border of an active quarry, very close to a road .Threat type: OngoingThreats: 3.2. Energy production & mining - Mining & quarrying4.1. Transportation & service corridors - Roads & railroads9.2. Pollution - Industrial & military effluentsThreat type: OngoingThreats: 3.2. Energy production & mining - Mining & quarrying4.1. Transportation & service corridors - Roads & railroads9.2. Pollution - Industrial & military effluentsJustification for conservation actions: Even though this cave is protected under legislation by the \u201cRede Natura 2000\u201d , this spConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTrichoniscoidessicoensisScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: Class: MalacostraIsopodaOrder: TrichoniscidaeFamily: Trichoniscoidessicoensis is blind and depigmented and it can be distinguished from all species of the genus because the male pereopod 7 merus has a lobe on the mid-sternal margin, the male pleopod 1 exopod has a broadly rounded outer margin and two unequal setae, the endopod has a fusiform distal article with a distinct circular suture in the middle and the male pleopod 2 endopod has a thickset distal article bearing two short triangular lobes and two setae at the apex  or Photo(s): Fig. 1Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 7Basis of EOO and AOO: Known habitat extentBasis (narrative): The extent of occurrence (EOO) is 129.2 km\u00b2 and the maximum estimated area of occupancy (AOO) is 20 km\u00b2.Min Elevation/Depth (m): 20Max Elevation/Depth (m): 380Trichoniscoidessicoensis is recorded from six caves located in the Sic\u00f3 karst area: Cer\u00e2mica, Santa Maria da Estrela, Soprador do Carvalho, Algarinho, Arrifana and S\u00e3o Sim\u00e3o (Range description: \u00e3o Sim\u00e3o .EOO (km2): 129.2Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 20Number of locations: 6Trichoniscoidessicoensis is known from six caves : The largest population was found in the type locality Cer\u00e2mica Cave, where a higher number of specimens has been collected, followed by Santa Maria da Estrela Cave and then Soprador do Carvalho, Algarinho, Arrifana and S\u00e3o Sim\u00e3o Caves .Number of subpopulations: 6Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesTrichoniscoidessicoensis inhabits the deepest and isolated parts of caves of the Sic\u00f3 karst area, found up to a maximum altitude of 380 m a.s.l. The easternmost locality is Soprador do Carvalho Cave and the westernmost is Santa Maria da Estrela Cave, its distribution being limited at the north by the Arrifana Cave and at the south by the Cer\u00e2mica Cave. Cer\u00e2mica Cave opens at 355 m a.s.l., it has a horizontal development of 120 m, a depth of 21 m :  of 21 m  and is t of 21 m . Santa M of 21 m . Sopradoalleries .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3.5 mm , 3.9 mm Generation length (yr): 1Dependency of single sp?: UnknownTrichoniscoidessicoensis is classified as a troglobiont species, blind and depigmented. It is endemic to the Sic\u00f3 karst area (Ecology and traits (narrative): rst area . Most ofrst area .Eucalyptus intensive plantations. It is located 270 m from a road, 550 m from an animal farm, 1.6 km from the closest village and 3.6 km from a quarry. Santa Maria da Estrela Cave is located 86 m from a touristic site called Monstro das Bolachas, 230 m from the Nossa Senhora da Estrela viewpoint, 250 m from the closest urbanised area, 80 m from the road, 220 m from agricultural fields and 2.6 km from two quarries. Soprador do Carvalho Cave is surrounded by agricultural lands distances and is located 67 m from the closest house and 1.4 km from a quarry. This cave is also affected by touristic activities, as people who visit the cave walk all over the habitat : Suppl. materials 8Basis of EOO and AOO: Known habitat extent2.Basis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 kmMin Elevation/Depth (m): 122Trichoniscoidessubterraneus is only recorded from Alta do Cabe\u00e7o dos Mosqueiros Cave, located in Carvalhal de Aljubarrota, the western part of the Estremenho karst massif (Range description: t massif .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Trichoniscoidessubterraneus occurs in a single cave : This cave is currently part of the Geocaching network; therefore, it is subject to human disturbance without regulation.Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3 mmGeneration length (yr): 1Dependency of single sp?: UnknownTrichoniscoidessubterraneus is classified as a blind and depigmented troglobiont species (Ecology and traits (narrative):  species .Justification for threats: This cave is frequently visited by geocachers looking for the geocache installed inside the cave.Threat type: OngoingThreats: 6.1. Human intrusions & disturbance - Recreational activitiesThreat type: OngoingThreats: 6.1. Human intrusions & disturbance - Recreational activitiesJustification for conservation actions: Measures to limit the number of visits to the cave should be implemented in order to ensure the integrity of the habitat and the survival of this species.Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelMetatrichoniscoidessalirensisScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Metatrichoniscoidessalirensis can be distinguished from all species of the genus because the pleopod 1 exopod of the male has two long distal setae of subequal length and its pleopod 2 endopod has a thickset distal article, ending in a thinner sinuous part with a beak-like small lobe medially directed : Suppl. materials 9Basis of EOO and AOO: Known habitat extentBasis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 km\u00b2Min Elevation/Depth (m): 60Metatrichoniscoidessalirensis is only known from Salir Cave, a single isolated cave located in the western border of Caldas da Rainha Typhonic Valley : 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Metatrichoniscoidessalirensis is only recorded from one cave : A total of seven specimens have been collected in the type locality .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): Salir Cave was discovered in the 1960s by the labour force of a quarry. It is located near the sea, at an altitude of 60 m a.s.l. The substrate is mostly composed of flowstone, clay and marine sand that can be found in the deepest parts of the cave .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 2.2 mm Generation length (yr): 1Dependency of single sp?: UnknownMetatrichoniscoidessalirensis is classified as a troglobiont species, blind and depigmented and is hitherto the only troglobiont species known from this cave and from this karst area (Ecology and traits (narrative): rst area . The meaJustification for threats: The cave entrance is located in a former quarry, at 400 m from the closest house and 1 km from the village centre. Salir Cave has an easy access and mostly horizontal development, which attracts casual visitors. It has suffered recurrent episodes of vandalism, which include paintings inside the first chamber and breakage of many lithological formations. This cave is currently listed in the Geocaching network and subject to human disturbance without regulation.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: The cave should be a protected site, as it is the only locality known for this species; therefore, a site of maximum priority for biodiversity conservation. The entrance should be closed and the access regulated in order to prevent the recurrent vandalism and human disturbance to the ecosystem.Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTroglonethesolissipoensisScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Taxonomic notes: Antennae have five flagellar articles, the male pleopod 1 exopod is triangular, as wide as long and the male pleopod 2 endopod distal article has a basal and a distal hook-like process .Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 10Basis of EOO and AOO: Known habitat extentBasis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 km\u00b2.Min Elevation/Depth (m): 42Troglonethesolissipoensis is only recorded from Alvide Cave, located in Cascais Municipality in the Lisbon metropolitan area (Range description: tan area .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Troglonethesolissipoensis occurs in a single cave : A total of 25 specimens have been collected in the type locality .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): The entrance to Alvide Cave is through a house currently serving as headquarters of Den\u00edvel Association. Part of the cave's ceiling has concrete that serves as the base of a residential building. This cave is in the margin of a small canyon and it is composed of three levels of horizontal galleries connected by pits, it has a horizontal development of 708 m and total depth of 28 m.Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3.5 mm , 4.2 mm Generation length (yr): 1Dependency of single sp?: UnknownTroglonethesolissipoensis is a troglobiont species, being blind and depigmented. It is endemic and the only cave-adapted species known from this cave (T.olissipoensis were collected in the deepest parts of the cave (Ecology and traits (narrative): his cave . The avethe cave .Justification for threats: Alvide Cave is located below an over-urbanised area .Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Measures should be taken to minimise the pernicious effects due to the cave's close proximity to the urbanisation on the habitat.Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTroglonethesarrabidaensisScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Taxonomic notes: Antennae have three flagellar articles, the male pereopod 7 carpus is enlarged in the basal part, the male pleopod 1 exopod is triangular and as wide as long and the male pleopod 2 endopod has the distal article with an apical hook-like process .Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 11Basis of EOO and AOO: Known habitat extentBasis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 km\u00b2.Min Elevation/Depth (m): 0Troglonethesarrabidaensis is only recorded from Frade Cave, located in the Arr\u00e1bida karst massif (Range description: t massif .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Troglonethesarrabidaensis occurs in a single cave : A total of 17 specimens have been collected in Frade Cave .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): Frade Cave is located near the seashore, with several anchialine lakes inside, which are influenced by the sea tides with a slight delay period . The cavTrend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 2.7 mm Generation length (yr): 1Dependency of single sp?: UnknownTroglonethesarrabidaensis is classified as a troglobiont, depigmented, blind and with an elongated body (Ecology and traits (narrative): ted body . It is aJustification for threats: The cave entrance is located 600 m from a very touristic beach, mainly accessed by boat; therefore, fuel residues brought in by the tidal movement might be a concerning pollutant inside the cave. Contamination of groundwater should be evaluated.Threat type: OngoingThreats: 9. Pollution6.1. Human intrusions & disturbance - Recreational activitiesThreat type: OngoingThreats: 9. Pollution6.1. Human intrusions & disturbance - Recreational activitiesJustification for conservation actions: Even though this cave is protected under legislation by the \u201cRede Natura 2000\u201d , this siConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelMiktoniscuslongispinaScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Taxonomic notes: The male pereopod 7 has a long and stout seta on the distal corner of the ischium and a triangular male pleopod 1 exopod . The speFigure(s) or Photo(s): Fig. 2Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 12Basis of EOO and AOO: Known habitat extentBasis (narrative): The extent of occurrence (EOO) is 137.96 km\u00b2 and the area of occupancy (AOO) is 12 km\u00b2.Min Elevation/Depth (m): 145Max Elevation/Depth (m): 355Miktoniscuslongispina is recorded from three caves with disjunt distribution. Bolhos Cave, also known as Casal da Lebre Cave, located in the Cesaredas Plateau and Ervilha and Cer\u00e2mica Caves located in the centre of the Sic\u00f3 karst area (Range description: rst area .EOO (km2): 137.96Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 12Number of locations: 3Miktoniscuslongispina can be found in three caves : A total of 12 specimens were collected in the three localities: two in Bolhos Cave, eight in Cer\u00e2mica Cave and two in Algar da Ervilha Cave .Number of subpopulations: 3Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: YesSystem: TerrestrialHabitat specialist: YesHabitat (narrative): Bolhos Cave has a horizontal development of 130 m and is located in Cesaredas Plateau, 93 km away from the other localities in the Sic\u00f3 karst area. Cer\u00e2mica Cave has a horizontal development of 120 m and is the richest cave in troglobiont species in central Portugal. Algar da Ervilha Cave has a depth of 52 m and a horizontal development of 150 m . The cavTrend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3.5 mm Generation length (yr): 1Dependency of single sp?: UnknownMiktoniscuslongispina is a troglobiont, with a depigmented and elongated body. All specimens, except for the ones collected in Bolhos Cave, are blind.Ecology and traits (narrative): Eucalyptus intensive plantations. It is located 270 m from a road, 550 m from an animal farm, 1.6 km from the closest village and 3.6 km from a quarry. Algar da Ervilha Cave is located in the border of a road in the westernmost access to Ereiras Village, at 90 m from the closest house and 3.5 km from the same quarry as Cer\u00e2mica Cave.Justification for threats: Bolhos Cave is located 130 m from an energy windmill and 700 m from the closest village, in an area of extensive agricultural fields. Cer\u00e2mica Cave is surrounded by agricultural fields and Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.2. Agriculture & aquaculture - Wood & pulp plantations2.3. Agriculture & aquaculture - Livestock farming & ranching3.2. Energy production & mining - Mining & quarrying3.3. Energy production & mining - Renewable energy4. Transportation & service corridors9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas2.2. Agriculture & aquaculture - Wood & pulp plantations2.3. Agriculture & aquaculture - Livestock farming & ranching3.2. Energy production & mining - Mining & quarrying3.3. Energy production & mining - Renewable energy4. Transportation & service corridors9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: The caves located in the Sic\u00f3 karst area are protected under the \u201cRede Natura 2000\u201d . Bolhos Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelMoseriusinexpectatusScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: TrichoniscidaeFamily: Moserius species due to the peculiar shape of the male pleopod 1 exopod, with a truncate and sinuous, rather than triangular, distal point : Suppl. materials 13Basis of EOO and AOO: Known habitat extent2.Basis (narrative): Both the extent of occurrence (EOO) and area of occupancy (AOO) are 4 kmMin Elevation/Depth (m): 95Moseriusinexpectatus is only recorded from Almonda Cave, also known as Olho do Moinho da Fonte, located in the easternmost subunit of the Estremenho karst massif (Range description: t massif .EOO (km2): 4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 4Number of locations: 1Moseriusinexpectatus is endemic to a single cave : Only one specimen was collected from the type locality .Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesHabitat (narrative): Almonda Cave, the type locality for this species, is the largest cave in Portugal .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 1.5 mm Generation length (yr): 1Dependency of single sp?: UnknownMoseriusinexpectatus is a troglobiont blind and depigmented isopod. This is the third species described from the genus Moserius, previously known from Slovenia and Italy (Ecology and traits (narrative): nd Italy .Justification for threats: Almonda Cave is located 50 m from a factory that extracts and uses water from a subterranean stream and 420 m from a village, which has many agricultural fields.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.1. Agriculture & aquaculture - Annual & perennial non-timber crops9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.1. Agriculture & aquaculture - Annual & perennial non-timber crops9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: In 1993, Almonda Cave was classified as a Property of Public Interest (IIP) and protected due to its archaeological heritage . The arcConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelCordioniscuslusitanicusScientific name: Species authority: Reboleira & Taiti 2015AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: StyloniscidaeFamily: Taxonomic notes: The male pereopod 7 ischium has a rounded hyaline basal lobe, the triangular male pleopod 1 exopod is as long as the endopod and the male pleopod 2 endopod has a complex apical part .Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 14Basis of EOO and AOO: Known habitat extent2 and the area of occupancy (AOO) is 16 km2.Basis (narrative): The extent of occurrence (EOO) is 5,893.93 kmMin Elevation/Depth (m): 10Max Elevation/Depth (m): 370Cordioniscuslusitanicus is recorded from five caves, located in two isolated karst areas: Algar de Santo Ant\u00f3nio from the Estremoz-Cano karst massif and Ibne Ammar, Algar\u00e3o do Remexido, Vale Telheiro and Senhora Caves from the Algarve karst massif (Range description: t massif .EOO (km2): 5,893.93Trend: UnknownJustification for trend: The four caves, located in the Algarve karst massif, are at 200 km distance from the cave in the Estremoz-Cano karst massif.Causes ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 16Number of locations: 5Cordioniscuslusitanicus occurs in five caves : A total of 26 specimens have been collected: nine from Algar de Santo Ant\u00f3nio, six from Ibne Ammar, six from Algar\u00e3o do Remexido and four from Senhora  and one Number of subpopulations: 5Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: UnknownCordioniscuslusitanicus was collected in two karst areas, Estremoz-Cano and Algarve, located more than 200 km apart, which are isolated from each other by the dry and flat areas of the Alentejo Province. In Algar de Santo Ant\u00f3nio, the specimens were collected in deep layers of soil at the bottom of an entrance pit of the cave (Habitat (narrative): the cave .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 3 mm , 5 mm Generation length (yr): 1Dependency of single sp?: UnknownCordioniscuslusitanicus is classified as a troglobiont and endogean species. It is blind, depigmented and has an elongated body (Ecology and traits (narrative): ted body .Justification for threats: Algar de Santo Ant\u00f3nio is located in the middle of an urbanised area of a village. Algar\u00e3o do Remexido is located under agricultural lands, 370 m from the closest house and 1.7 km from the closest village. Senhora Cave is located 168 m from the closest house and 900 m from an industrial complex. Ibne Ammar Cave is located right in the flooding zone of the Arade river, 380 m from the national road IC4 and 1.4 km from the nearest town. Vale Telheiro Cave is located 50 m from a road and 150 m from the closest house.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.2. Agriculture & aquaculture - Wood & pulp plantations4. Transportation & service corridors9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.2. Agriculture & aquaculture - Wood & pulp plantations4. Transportation & service corridors9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Of the five caves, only Ibne Ammar Cave is protected under legislation by the \u201cRede Natura 2000\u201d  and thisConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelPorcelliocavernicolusScientific name: Species authority: Vandel, 1946AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: PorcellionidaeFamily: Figure(s) or Photo(s): Fig. 3Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 15Basis of EOO and AOO: Known habitat extent2 and the area of occupancy (AOO) is 24 km2.Basis (narrative): The extent of occurrence (EOO) is 713 kmMin Elevation/Depth (m): 20Max Elevation/Depth (m): 380Porcelliocavernicolus is recorded from seven caves located in two isolated massifs, Gruta d\u2019el Rey in the Cantanhede-Outil karst massif and Santa Maria da Estrela, Soprador do Carvalho, Algarinho, Cer\u00e2mica, Abrigo Tomar I and Furjaca caves, located in the Sic\u00f3 massif (Range description: \u00f3 massif .EOO (km2): 713Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 24Number of locations: 7Porcelliocavernicolus occurs in seven caves : More than 63 specimens have been collected: 28 from Gruta d\u2019el Rey, 16 from Santa Maria da Estrela, seven from Soprador do Carvalho, three from Cer\u00e2mica, four from Abrigo Tomar I and five from Furjaca. The specimens, collected from Algarinho Cave, are simply described as \u201cmany\u201d .Number of subpopulations: 7Trend: UnknownExtreme fluctuations?: UnknownSevere fragmentation?: UnknownSystem: TerrestrialHabitat specialist: YesPorcelliocavernicolus inhabits the most superficial parts of caves and it occurs on roots that hang from the ceiling. Specimens are easily distinguishable due to their whitish colouration (Habitat (narrative): ouration .Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 10 mm Generation length (yr): 1Dependency of single sp?: UnknownPorcelliocavernicolus is classifid as a troglobiont, endemic to seven caves from central Portugal, distributed in two isolated karst massifs (Ecology and traits (narrative):  massifs .Eucalyptus intensive plantations. It is located 270 m from a road, 550 m from an animal farm, 1.6 km from the closest village and 3.6 km from a quarry. Abrigo Tomar I Cave is located in a quite pristine location and it is a local protected area by the ONG Quercus. Furjaca Cave is located in an abandoned quarry which, after closure, was used as a trash dumping site.Justification for threats: Gruta d\u2018el Rey is located in the middle of an urbanised area, 1 km from a quarry and 1.2 km from highway A14. Santa Maria da Estrela Cave is located 86 m from a touristic site called Monstro das Bolachas, 230 m from the Nossa Senhora da Estrela viewpoint, 250 m from the closest house, 80 m from the closest road, 220 m from agricultural fields and 2.6 km from two quarries. Soprador do Carvalho is surrounded by agricultural lands and is located 67 m from the closest house and 1.4 km from a quarry. This cave is also affected by touristic activities, as people who visit the cave walk all over the habitat . The subThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.2. Agriculture & aquaculture - Wood & pulp plantations3.2. Energy production & mining - Mining & quarrying4. Transportation & service corridors6.1. Human intrusions & disturbance - Recreational activities9.1. Pollution - Domestic & urban waste water9.4. Pollution - Garbage & solid wasteThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.2. Agriculture & aquaculture - Wood & pulp plantations3.2. Energy production & mining - Mining & quarrying4. Transportation & service corridors6.1. Human intrusions & disturbance - Recreational activities9.1. Pollution - Domestic & urban waste water9.4. Pollution - Garbage & solid wasteJustification for conservation actions: Of the seven locations, only three are protected under legislation by the \u201cRede Natura 2000\u201d . Almost Conservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelTroglelumamachadoiScientific name: Species authority: AnimaliaKingdom: ArthropodaPhylum: MalacostracaClass: IsopodaOrder: ArmadillidiidaeFamily: Figure(s) or Photo(s): Fig. 4Region for assessment: EuropeReviewers: ReviewersEditor: EditorReviewers: ReviewersEditor: EditorBiogeographic realm: PalearcticCountries: PortugalMap of records (Google Earth): Suppl. materials 16Basis of EOO and AOO: Known habitat extent2 and the area of occupancy (AOO) is 16 km2.Basis (narrative): The extent of occurrence (EOO) is 356.4 kmMin Elevation/Depth (m): 10Max Elevation/Depth (m): 260Troglelumamachadoi is recorded from six caves, located in the Algarve karst massif: Ibne Ammar, Algar\u00e3o do Remexido, Vale Telheiro, Senhora, Algar\u00e3o do Paulino and Abismo Novo (Range description: smo Novo .EOO (km2): 356.4Trend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownTrend: UnknownCauses ceased?: UnknownCauses understood?: UnknownCauses reversible?: UnknownExtreme fluctuations?: UnknownAOO (km2): 16Number of locations: 6Troglelumamachadoi occurs in six caves : Trend in extent, area or quality?: Decline (inferred)Habitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesHabitat importance: Major ImportanceHabitats: 7.1. Caves and Subterranean Habitats (non-aquatic) - CavesSize: 8 mmGeneration length (yr): 1Dependency of single sp?: UnknownTroglelumamachadoi is a blind and depigmented species, classified as a troglobiont. Specimens are found in the most deep and well isolated parts of of caves, they have the integument covered with clay, while some others were found walking on cave walls and completely clean of clay, probably recently moulted :  moulted . The temicamagna , the relgmaticus , the dyslitoides , the milocalense , the dipamendesi , the giagharbica  and the arvensis .Justification for threats: Ibne Ammar Cave is located right in the flooding zone of the Arade River, 380 m from the national road IC4 and 1.4 km from the nearest town. Algar\u00e3o do Remexido is located under agricultural lands, 370 m from the closest house and 1.7 km from the closest village. Senhora Cave is located 168 m from the closest house and 900 m from an industrial complex. Abismo Novo Cave is located 100 m from the closest house and 500 m from the village centre and is also located 1 km from Senhora Cave. Algar\u00e3o do Paulino is located near a road, 90 m from the closest house and 800 m from the closest village. Vale Telheiro Cave is located 50 m from a road and 150 m from the closest house.Threat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.1. Agriculture & aquaculture - Annual & perennial non-timber crops4. Transportation & service corridors9.1.2. Pollution - Domestic & urban waste water - Run-offThreat type: OngoingThreats: 1.1. Residential & commercial development - Housing & urban areas1.2. Residential & commercial development - Commercial & industrial areas2.1. Agriculture & aquaculture - Annual & perennial non-timber crops4. Transportation & service corridors9.1.2. Pollution - Domestic & urban waste water - Run-offJustification for conservation actions: Of the five locations, only three are protected under legislation by the \u201cRede Natura 2000\u201d . MeasureConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelConservation action type: NeededConservation actions: 1.1. Land/water protection - Site/area protection1.2. Land/water protection - Resource & habitat protection2.1. Land/water management - Site/area management4. Education & awareness5.1.3. Law & policy - Legislation - Sub-national levelCave-adapted terrestrial isopods are key species for cave ecosystem conservation: i) they are the most diverse group of cave-adapted species in continental Portugal, ii) they have several single cave endemics that are under threat and require specific protection measures, iii) they are basal in the trophic chains in caves and serve as a food source for several other zoological groups; iv) they play a vital role on the decomposition of organic matter in caves; and v) they are very sensitive to contaminants and climate change .Almost all cave-adapted terrestrial isopod species face direct anthropogenic threats, such as point or diffuse pollution, direct habitat destruction by mining and quarry activities or excess cave visitation . With thOniscidea  . So far,niscidea . Therefod groups .It is a priority to establish concrete protection strategies for cave-adapted species in continental Portugal. We need to improve the knowledge about population size and dynamics, real extent of subterranean distribution, improve our knowledge on the functional ecology, understand species life cycle and evaluate their sensitivity to disturbance. This contribution may be used as a support for decision-making for territory management and to define conservation measures for cave endemic species. Cave-adapted terrestrial isopods have the potential to be used as umbrella species for the conservation of other cave-adapted species sharing the same subterranean habitats.Discussion2F1C3046-2169-5E36-AB53-F8D02E3A9E1410.3897/BDJ.10.e78796.suppl1Supplementary material 1Distribution of cave-adapted terrestrial isopods in continental PortugalData typeSpecies distribution mapBrief description(A) Overview of the distribution of cave-adapted terrestrial isopods in continental Portugal; (B) Sic\u00f3 karst area distribution detail; (C) Estremenho and Montejunto karst massifs, Caldas da Rainha Typhonic Valley and Cesaredas Plateau distribution detail; (D) Algarve karst massif distribution detail; (E) Lisbon Peninsula and Arr\u00e1bida karst massif and Estremoz-Cano karst massif distribution detail; and (F) Vimioso paleokarst distribution detail.Species:Trichoniscoidesbroteroi (orange circle), T.ouremensis (pink circle), T.serrai (dark blue circle), T.subterraneus (yellow circle), T.meridionalis (purple circle), T.bellesi (forest green circle), T.sicoensis (light blue circle), Metatrichoniscoidessalirensis (dark blue diamond), Troglonethesolissipoensis (dark blue star), T.arrabidaensis (pink star), Miktoniscuslongispina (yellow cross), Moseriusinexpectatus (light blue hexagon), Cordioniscuslusitanicus (dark blue triangle outline), Porcelliocavernicolus (pink triangle) and Troglelumamachadoi (pink circle outline).File: oo_607434.pnghttps://binary.pensoft.net/file/607434A.S.P.S. Reboleira, R.P. Eus\u00e9bioDD91E314-7C4E-5774-8FF0-6086CE34BF8A10.3897/BDJ.10.e78796.suppl2Supplementary material 2TrichoniscoidesbellesiDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidesbellesi distribution: Algar do Javali Cave, Montejunto karst massif.File: oo_607707.tifhttps://binary.pensoft.net/file/607707A.S.P.S. Reboleira, R.P.Eus\u00e9bio5914DE8F-D8BC-5B98-9A9F-9D4D7A6E1EE910.3897/BDJ.10.e78796.suppl3Supplementary material 3TrichoniscoidesbroteroiDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidesbroteroi distribution: Alqueves Cave, Sic\u00f3 karst area.File: oo_607708.tifhttps://binary.pensoft.net/file/607708A.S.P.S. Reboleira, R.P. Eus\u00e9bio5FB36EF8-919B-5348-86EF-333B2BE58A3510.3897/BDJ.10.e78796.suppl4Supplementary material 4TrichoniscoidesmeridionalisDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidesmeridionalis distribution. (A) Detail of distribution: Algar do Vale do Pena; Alcobertas Cave; Algar do Z\u00e9 de Braga Cave; Lapa da Ch\u00e3 de Cima Cave; Algar do Ladoeiro Cave; Algar do Pena Cave; Moinhos Velhos Cave; Papagaio Cave; Algar do Burro Cave; and Almonda Cave.File: oo_607711.tifhttps://binary.pensoft.net/file/607711A.S.P.S. Reboleira, R.P. Eus\u00e9bio597F809C-3090-5016-9767-C3E76CCD0A1610.3897/BDJ.10.e78796.suppl5Supplementary material 5TrichoniscoidesouremensisDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidesouremensis distribution: Lapa da Salgada Cave, F\u00e1tima Plateau.File: oo_607712.tifhttps://binary.pensoft.net/file/607712A.S.P.S. Reboleira, R.P. Eus\u00e9bioF92DCDAA-FA35-5326-85D4-F63245509A6C10.3897/BDJ.10.e78796.suppl6Supplementary material 6TrichoniscoidesserraiDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidesserrai distribution: Santo Adri\u00e3o Cave, Vimioso karst area.File: oo_607713.tifhttps://binary.pensoft.net/file/607713A.S.P.S. Reboleira, R.P. Eus\u00e9bio48CB91A0-5194-5E31-94A2-A512AA8133E510.3897/BDJ.10.e78796.suppl7Supplementary material 7TrichoniscoidessicoensisDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidessicoensis distribution. (A) Detail of the distribution: Arrifana Cave; Santa Maria da Estrela Cave; Cer\u00e2mica Cave; S\u00e3o Sim\u00e3o Cave; Algarinho Cave; and Soprador do Carvalho Cave.File: oo_607714.tifhttps://binary.pensoft.net/file/607714A.S.P.S. Reboleira, R.P.Eus\u00e9bio774C7847-3F7C-544E-81E9-48B9C9A29AFE10.3897/BDJ.10.e78796.suppl8Supplementary material 8TrichoniscoidessubterraneusDistribution of Data typeSpecies distribution mapBrief descriptionTrichoniscoidessubterraneus distribution: Alta do Cabe\u00e7o dos Mosqueiros Cave, Aljubarrota Plateau.File: oo_607720.tifhttps://binary.pensoft.net/file/607720A.S.P.S. Reboleira, R.P. Eus\u00e9bio71845157-2E1F-5082-AAA4-DDC0924AFDBD10.3897/BDJ.10.e78796.suppl9Supplementary material 9MetatrichoniscoidessalirensisDistribution of Data typeSpecies distribution mapBrief descriptionMetatrichoniscoidessalirensis distribution: Salir Cave, Caldas da Rainha.File: oo_607721.tifhttps://binary.pensoft.net/file/607721A.S.P.S. Reboleira, R.P. Eus\u00e9bio425A22BF-FE18-5FDB-B23D-8937DEED57B610.3897/BDJ.10.e78796.suppl10Supplementary material 10TroglonethesolissipoensisDistribution of Data typeSpecies distribution mapBrief descriptionTroglonethesolissipoensis distribution: Alvide Cave, Lisbon.File: oo_607722.tifhttps://binary.pensoft.net/file/607722A.S.P.S. Reboleira, R.P. Eus\u00e9bio85C91C9B-5B83-533D-AB14-9F6ABE3F8A6B10.3897/BDJ.10.e78796.suppl11Supplementary material 11TroglonethesarrabidaensisDistribution of Data typeSpecies distribution mapBrief descriptionTroglonethesarrabidaensis distribution: Frade Cave, Arr\u00e1bida karst massif.File: oo_607723.tifhttps://binary.pensoft.net/file/607723A.S.P.S. Reboleira, R.P.Eus\u00e9bioBC5A499B-29AA-5FBE-B1CE-77CC7E4752A510.3897/BDJ.10.e78796.suppl12Supplementary material 12MiktoniscuslongispinaDistribution of Data typeSpecies distribution mapBrief descriptionMiktoniscuslongispina distribution. (A) Detail of distribution: Casal da Lebre Cave; Algar da Ervilha Cave; and Cer\u00e2mica Cave.File: oo_607725.tifhttps://binary.pensoft.net/file/607725A.S.P.S. Reboleira, R.P. Eus\u00e9bioDB2A85A3-79E9-53CC-AC9E-E2C21922CF6F10.3897/BDJ.10.e78796.suppl13Supplementary material 13MoseriusinexpectatusDistribution of Data typeSpecies distribution mapBrief descriptionMoseriusinexpectatus distribution: Almonda Cave, Estremenho karst massif.File: oo_607726.tifhttps://binary.pensoft.net/file/607726A.S.P.S. Reboleira, R.P. Eus\u00e9bioC9060AAB-01F8-5426-86EE-22A80F48CE2110.3897/BDJ.10.e78796.suppl14Supplementary material 14CordioniscuslusitanicusDistribution of Data typeSpecies distribution mapBrief descriptionCordioniscuslusitanicus distribution. (A) Detail of distribution: Ibne Ammar Cave; Algar\u00e3o do Remexido Cave; Vale Telheiro Cave; Senhora Cave; and Algar de Santo Ant\u00f3nio Cave.File: oo_607729.tifhttps://binary.pensoft.net/file/607729A.S.P.S. Reboleira, R.P. Eus\u00e9bio9A58FB28-25DF-5F94-842E-3D0690437F8C10.3897/BDJ.10.e78796.suppl15Supplementary material 15PorcelliocavernicolusDistribution of Data typeSpecies distribution mapBrief descriptionPorcelliocavernicolus distribution. (A) Detail of distribution: Gruta d\u2019el Rey; Furjaca Cave; Santa Maria da Estrela Cave; Cer\u00e2mica Cave; Algarinho Cave; Soprador do Carvalho Cave; and Abrigo Tomar I Cave.File: oo_607732.tifhttps://binary.pensoft.net/file/607732A.S.P.S. Reboleira, R.P. Eus\u00e9bioF77EA613-7319-514F-BCF6-CF2EFDC4F8E210.3897/BDJ.10.e78796.suppl16Supplementary material 16TroglelumamachadoiDistribution of Data typeSpecies distribution mapBrief descriptionTroglelumamachadoi distribution. (A) Detail of distribution: Ibne Ammar Cave; Algar\u00e3o do Remexido Cave; Vale Telheiro Cave; Algar\u00e3o do Paulino Cave; Abismo Novo Cave; and Senhora Cave.File: oo_607736.tifhttps://binary.pensoft.net/file/607736A.S.P.S. Reboleira, R.P. Eus\u00e9bio"}
+{"text": "Monocyte derived dendritic cells were generated from these peripheral blood mononuclear cells (PBMC), loaded with SARS-CoV-2 peptides, and used to induce autologous CD4+ and CD8+ T cell activation. The SARS-CoV-2 peptides demonstrated antigenicity against the T-cells from individuals with previous SARS-CoV-2 infection indicating that this approach holds promise as a method to activate anti-SAR-CoV-2 T-cell responses from conserved regions of the virus which are not included in vaccines utilising the Spike protein.The pandemic caused by SARS-CoV-2 has led to the successful development of effective vaccines however the prospect of variants of SARS-CoV-2 and future coronavirus outbreaks necessitates the investigation of other vaccine strategies capable of broadening vaccine mediated T-cell responses and potentially providing cross-immunity. In this study the SARS-CoV-2 proteome was assessed for clusters of immunogenic epitopes restricted to diverse human leucocyte antigen. These regions were then assessed for their conservation amongst other coronaviruses representative of different alpha and beta coronavirus genera. Sixteen highly conserved peptides containing numerous HLA class I and II restricted epitopes were synthesized from these regions and assessed  NC_045512.2, MN996532.2, MT072864.1, MT040333.1, KC881005.1, KY417144.1, KT444582.1, KF367457.1, KC881006.1, KY417152.1, MK211376.1, AY572034.1, AY572035.1, AY572038.1, AY686864.1, AY686863.1, NC_004718.3, AY390556.1, KY352407.1, NC_014470.1, GU190215.1, MG772933.1, MG772934.1, MK211374.1, JX993987.1, MK211377.1, DQ084199.1, GQ153539.1, DQ084200.1, DQ022305.2, KJ473815.1, KJ473813.1, KJ473812.1, KP886808.1, DQ648857.1, DQ412043.1, KY417147.1, DQ648856.1, JX993988.1, FJ588686.1, MK211378.1, KJ473816.1, KJ473811.1, MG596802.1, MG596803.1, NC_009020.1, MG021452.1, MK129253.1, JX869059.2, MT576585.1, NC_038294.1, DQ648794.1, KJ473821.1, KJ473822.1, KX442564.1, EF065505.1, NC_009019.1, KC869678.4, NC_039207.1, KC545386.1, KJ473806.1, NC_026011.1, NC_017083.1, FJ647223.1, KF686343.1, AY391777.1, NC_030886.1, NC_009021.1, HM211098.1, MK211375.1, KY770860.1, KU182964.1, MT430884.1, LC469308.1, NC_048212.1, MF593268.1, NC_034440.1, NC_028814.1, JQ989273.1, NC_018871.1, AF304460.1, JQ410000.1, NC_048216.1, AY567487.2, NC_046964.1, NC_028833.1, KJ473820.1, NC_028811.1, NC_022103.1, EF203064.1, NC_028824.1, DQ648858.1, EU420138.1, KJ473795.1, EU420137.1.The pandemic caused by the novel coronavirus, SARS-CoV-2, has prompted a global response to produce effective vaccines. A number of vaccines have been approved for use having demonstrated varying levels of efficacy in clinical trials . These vin vitro.T-cell responses to SARS-CoV-1 demonstrate greater durability than those of neutralising antibody  and are https://www.ncbi.nlm.nih.gov/labs/virus/vssi/), alignment using Clustal Omega with default settings  and analysis of conservation using Microsoft Excel. Accession numbers of coronaviruses strains can be found in the supplementary material file.Analysis of SARS-CoV-2 and other coronaviruses protein sequences from proteins harbouring epitope rich regions was performed using FASTA sequences deposited in the NCBI database (https://www.iedb.org/).Identification of SARS-CoV-2 HLA restricted epitopes using prediction algorithms and experimentally validated epitopes deposited in the Immune epitope database and by using the NetMHCpan EL 4.1 (2020.09) prediction algorithm against an HLA allele reference set (Selection of conserved immunogenic regions between 15\u201330 amino acids in length (synthetic long peptides) was determined by assessing their suitability for synthesis based upon the physiochemical properties of the amino acids in the sequence and potential as CPP (cell penetrating peptides) defined by a net positive charge. Selected peptides were synthesized .Peripheral blood mononuclear cells (PBMC) were purchased from the national blood service  prior to the distribution of SARS-CoV-2 vaccines. PBMC demonstrating responses to a SARS-CoV-2 consensus peptide pool and serum antibody responses to the SARS-CoV-2 spike protein were defined as having been previously infected with SARS-CoV-2 whilst PBMC and sera lacking detectable responses were defined as SARS-CoV-2 na\u00efve.\u22121 IL-4 and 50 ng ml\u22121 GM-CSF . These mDDC were subsequently co-cultured with the T-cells enriched from the CD14- fraction of PBMC using anti-CD3 magnetic beads  in the presence of SARS-CoV-2 peptides individual and in groups, and control peptides including CEF , CEFT  SARS-CoV-2 reference group  (1 \u00b5g ml\u22121 for each peptide) for antigenicity assays.The generation of mDDC (monocyte-derived dendritic cells) was performed using established protocols. CD14+ cells were isolated by positive selection using anti-CD14 conjugated magnetic beads . The CD14+ cells were cultured for 6 days in RPMI  10% foetal calf serum  and 5% streptomycin/penicillin  of 10 ng ml4 mDDC: 2\u00d7105 T-cells) were incubated in IFN-\u03b3 ELISPOT plates  for 48 h in order to assess antigen specific T-cell activation. After 48 h ELISPOT plates were processed according to the manufacturers protocol. Briefly, plates were washed in PBS \u00d73 prior to the incubation with detection anti-IFN-\u03b3 antibody for 2 h. Plates were washed \u00d73 in PBS wash buffer and Strep-Biotin reagent was added for 1 h. Plates were washed \u00d73 prior to the addition of substrate solution. Spot formation was observed and the plates were washed once in distilled H2O and left to dry before enumeration using a CTL Immunospot entry ELISPOT plate reader. Positive responses compared to the no peptide control were defined as >1.5-fold change and statistical significance using Student\u2019s t-test.IFN-\u03b3 ELISPOT (enzyme-linked immunosorbent spot) assays using peptide pulsed mDDC-T-cell co-cultures  since the physiochemical properties of the peptides may make them unsuitable for synthesis or for targeting toward antigen presenting cells or homology between the peptides and human proteins may make them unsuitable for vaccination. Amino acid modifications were made outside of epitope containing regions in order to improve synthesis, stability and internalisation. Each of the peptides was differently conserved between other coronaviruses and contained a different number of epitopes restricted to different HLA. A total of 16 of these candidate SLP were selected as an immunogenic pool for The selected peptides were assessed for their ability to activate T-cell responses. PBMC from seven individuals with previous SARS-CoV-2 infection and four individuals seronegative for SARS-CoV-2, without selection for specific HLA expression, were used to generate monocyte derived dendritic cells , loaded 4895-915 which demonstrated a CD4+ T cell response from the PBMC of one individual but no CD8+ T cell responses. This pattern of responses was consistent with other reports studying anti-SARS T-cell responses [Next a flow cytometry-based activation induced marker (AIM) assay was used to gain a greater insight into the T-cell responses raised by the peptides  and CD8+esponses . Interesesponses .in vitro T-cell antigenicity of SARS-CoV-2 derived SLP containing epitopes restricted to multiple HLA and conserved between SARS-CoV-2 variants and other coronaviruses.Taken together these data demonstrate the SARS-CoV-2 vaccines based upon the Spike protein have demonstrated between 60\u201395% efficacy in phase three trials and are now in widespread use. Although these vaccines are highly efficacious numerous issues remain unresolved. These include supply, cost and the requirement of some vaccines for a cold chain. From an immunological perspective there remains concern that variation in the Spike protein may evolve against which antibodies raised by vaccination are less effective, demonstrated to some degree by the Gamma  and DeltThe broad therapeutic activation of SARS specific T-cell immune responses may resolve or ameliorate a number of these issues. T-cell responses are more stable than humoral responses  whilst pIn this study, immunogenic regions from SARS-CoV-2 proteins other than the spike were identified and their conservation amongst selected alpha and beta coronaviruses was assessed. The selected peptides contain multiple epitopes restricted to the most common HLA class I molecules and which have previously demonstrated induction of T-cell activation in response to SARS-CoV-2 . ImportaThe peptides are derived from the immunodominant viral proteins other than the spike  so couldin vitro experiments DC-T-cell co-cultures generated from the PBMC of individuals with previous SARS-CoV-2 infection responded with IFN-\u03b3 expression to an average of five peptides derived from an average of four proteins. Previous studies have demonstrated that broad T-cell responses against multiple epitopes are more effective than narrow responses targeting fewer epitopes [A recent study showed that CD8+ T cell responses against SARS-CoV-2 were raised against approximately 17 epitopes derived from between 1\u20136 viral proteins (average 2.7) [epitopes  and lessepitopes  indicatiAnalysis of T-cell responses to the ChAdOx1 spike-based vaccine showed that nearly 30% of unique TCRs raised by the vaccine mapped to a single region of the spike protein which is mutated in the Beta variant of SARS-CoV-2 . This maSynthetic long peptides of the kind studied here have been used in numerous therapeutic vaccines for both infectious disease and cancer and demonstrated an ability to induce efficacious T-cell responses . PeptideClick here for additional data file."}
+{"text": "People admitted to a skilled nursing facility (SNF) for post-acute care undergo comprehensive evaluation and rehabilitation, potentially enabling prediction of future functional recovery. We identified the first SNF admission per beneficiary  between 07/01/2014 \u2013 06/30/2016 in a 5% Medicare sample, using the Minimum Data Set (MDS) and the Outcome and Assessment Information Set (OASIS). Episodes were excluded for non-community discharge  or no OASIS admission assessment within 14 days of SNF discharge . A deficit accumulation Frailty Index (FI) was measured on admission MDS assessment and categorized into robust (MDS-FI<0.15), pre-frailty (MDS-FI0.15-0.24), mild frailty (MDS-FI0.25-0.34), and moderate or worse frailty (MDS-FI\u22650.35). Outcomes were functional decline obtained from OASIS, readmission, or death after initiation of home care. Functional status was measured by activities of daily living from OASIS assessments. A total of 135,310 SNF episodes were matched to OASIS episodes. Of these, there were 6,472 (4.8%) robust patients, 38,923 (28.8%) pre-frail, 63,727 (47.1%) mildly frail and 26,053 (19.3%) moderately frail or worse. In a logistic regression after adjustment for OASIS admission function, compared to robust status, frailty was associated with hospital readmission or death within 30 days of OASIS admission, (mild frailty OR1.33 [95%CI 1.23-1.45] and moderate or worse OR1.81 [95%CI 1.66-1.97]). Frailty was also associated with functional decline at OASIS discharge, after adjustment for OASIS admission function (mild frailty OR1.50 [95%CI 1.38-1.63] and moderate or worse OR2.30 [95%CI 2.11-2.50]). Among those discharged from SNF with home services, a SNF-based MDS-FI is associated with increased likelihood of poor community outcomes."}
+{"text": "Tracheal, bronchus, and lung (TBL) cancer is the leading cause of cancer death globally, but trends in TBL mortality attributable to tobacco, ambient particulate matter pollution (APMP), and household air pollution (HAP) were unequally distributed within global population subgroups over the last three decades. We used data from the Global Burden of Disease 2019 study to quantify the impact of sex, time, sociodemographic development index (SDI), and age for each exposure from 1990\u20132019. During that interval, tobacco dominated the TBL cancer mortality landscape, with its minimum global age-adjusted death rate of 16.71 deaths/100,000 : 15.27\u201318.13) outstripping maximums of 3.85 deaths/100,000 (UI: 2.82\u20134.83) and 2.54 deaths/100,000 (UI: 1.69\u20133.54) for APMP and HAP, respectively. In 2019, tobacco male TBL death rates exceeded female rates by a factor of 4.4:1. Ratios of 1.9:1 for APMP and 2.1:1 for HAP were seen. Our analysis indicates that both-sex middle SDI and female low, low-middle, and high-middle SDI populations are suffering increasing tobacco TBL burden. Efforts producing successful global reductions in HAP-associated TBL mortality should continue, with attention to low SDI female death rate increases. Finally, except for high SDI populations, global APMP-attributable TBL cancer burden is increasing and represents a major health concern. Lung cancer is the leading cause of cancer death globally , with trRecent studies have shed light on TBL incidence, DALYs, and mortality trends in the GBD 2017 and GBD 2019 data ,11,12. WAnnual TBL cancer deaths and death rate estimates attributable to environmental exposures were extracted from the GBD 2019 dataset . Death rt-tests of the 95% confidence interval of the difference between a given subgroup\u2019s 1990 and 2019 death rates, at a significance level of p < 0.05 . Statistically significant findings are termed \u201csignificant\u201d in the Results section; non-significant findings are not preceded by a word modifier. Lastly, using Microsoft Excel software, we generated heat maps for each exposure using death rate stratification by sex, SDI, and age between 1990 and 2019 to highlight granular trends among subpopulations. Other visualizations were generated using the GBD Compare tool [Specific considerations for each stratifying variable were as follows: (1) Time: TBL deaths and age-standardized death rate estimates were available for all years between 1990\u20132019; (2) SDI: The GBD 2019 authors created SDI as a country\u2019s \u201ccomposite indicator of socio-demographic development status\u201d utilizing \u201ctotal fertility rate in those under 25 years old, mean education for those 15 years or older, and lag-distributed income per capita\u201d to give each country an index value from 0 to 100 [are tool .From 1990\u20132019, tobacco dominated the TBL cancer mortality landscape. Tobacco-attributed global age-adjusted death rates ranged from 16.71 deaths/100,000 (UI: 15.27\u201318.13) to 19.99 deaths/100,000 (UI: 19.10\u201320.86), which outstripped interval maximum rates of 3.85 deaths/100,000 (UI: 2.82\u20134.83) and 2.54 deaths/100,000 (UI: 1.69\u20133.54) for APMP and HAP, respectively c. Globalp < 0.01). Males in the global population experienced a large, significant decrease (p < 0.01), while the female trend was a non-significant increase (p = 0.80). Exploring these trends by SDI highlighted significant decreases of 30% in HSDI (p < 0.01) and 14% in HMSDI (p = 0.01) both-sex populations (p < 0.01) (p = 0.06) (p = 0.10); MSDI females saw a 31% increase (p = 0.07) (p = 0.02) and LMSDI females a non-significant increase, while male subgroups saw non-significant decreases . The global male rate increased 12% to 5.78 deaths/100,000 (UI: 4.19\u20137.18), while the female rate increased 60% (p = 0.06) to 2.08 deaths/100,000 (UI: 1.49\u20132.71) (p = 0.11), with similar decreases in the male and female rates during this period (p = 0.01) and 133% (p < 0.01) for MSDI populations (p = 0.01), 263% in LMSDI (p < 0.01), 178% in MSDI (p < 0.01), and 81% in HMSDI (p = 0.02). LMSDI and MSDI males additionally saw a significant increase of 157% (p = 0.01) and 119% (p = 0.01), respectively, while LSDI and LMSDI males also saw non-significant increases  to 3.78 deaths/100,00 (UI: 2.79\u20134.86) between 1990\u20132019 (49\u20132.71) a\u2013c. HSDIp = 0.01) from 2.54 deaths/100,000 (UI: 1.69\u20133.54) to 0.97 deaths/100,000 (UI: 0.55\u20131.93). The decrease in males was larger than that in females, with significant reductions of 66% (p = 0.01) in the former and a 54% reduction (p = 0.03) in the latter  and, more recently, China (HMSDI) do offer some indication that implementing regulations aimed at improving APMP levels can reduce related TBL mortality . HMSDI fHAP trends offered the most universally positive outlook among the three exposures studied. HAP-attributable TBL mortality reductions across almost every global subpopulation can be taken as a sign that international efforts to reduce the burden of disease associated with these exposures, such as indoor stove interventions ,23, haveOur findings further characterize the complex, dynamic, exposure-related trends in TBL cancer mortality from 1990\u20132019. Our analysis indicates that the most urgent areas for tobacco-related intervention are both-sex MSDI populations and female LSDI, LMSDI, and HMSDI populations suffering from increasing tobacco-attributable TBL burden. Successful efforts that have brought about global reductions in HAP-associated TBL burden should continue, with special attention to LSDI female populations, where death rate increases were seen. Finally, except for both-sex HSDI and younger male HMSDI populations, the TBL cancer burden attributed to APMP is increasing and represents an urgent threat to the health of many global populations. With these targeted insights, global leaders, policy makers, and health care professionals will be better able to address the contribution of TBL cancer to the global burden of disease."}
+{"text": "We externally validated the recently suggested FSAC prediction model for hepatocellular carcinoma (HCC) in treatment-na\u00efve Asian chronic hepatitis B patients starting potent antiviral therapy (AVT). The model reflects age, sex, presence of cirrhosis, and on-therapy changes in non-invasive fibrosis markers (NFMs) after 12 months of antiviral therapy, such as APRI and FIB-4. Our results highlighted better predictive performance for the FSAC model for HCC (Harrell\u2019s c-index: 0.770) than the PAGE-B, modified PAGE-B, modified REACH-B, LSM-HCC, and CAMD models, which only use baseline parameters. A simplified version of FSAC score ), including only NFMs at 12 months, also showed a high c-index value (0.763). Our retrospective study suggests that the accurate measurement of intra-hepatic fibrotic burden during adequate AVT is necessary for predicting HCC development.n = 3026; median age, 50.0 years; male predominant (61.3%); cirrhosis in 1391 (46.0%) patients) receiving potent AVTs for >18 months between 2007 and 2018. During follow-up (median 64.0 months), HCC developed in 303 (10.0%) patients. Patients with low FIB-4 or APRI levels at 12 months showed significantly lower HCC risk than those with high NFM levels at 12 months . Cumulative 3-, 5-, and 8-year HCC probabilities were 0.0%, 0.3% and 1.2% in the low-risk group (FSAC \u2264 2); 2.1%, 5.2%, and 11.1% in the intermediate-risk group (FSAC 3\u22128); and 5.2%, 15.5%, and 29.8% in the high-risk group (FSAC \u2265 9) (both p < 0.001 between each adjacent pair). Harrell\u2019s c-index value for FSAC score (0.770) was higher than those for PAGE-B (0.725), modified PAGE-B (0.738), modified REACH-B (0.737), LSM-HCC (0.734), and CAMD (0.742). Our study showed that the FSAC model, which incorporates on-therapy changes in NFMs, had better predictive performance than other models using only baseline parameters.Antiviral therapy (AVT) induces the regression of non-invasive fibrosis markers (NFMs) and reduces hepatocellular carcinoma (HCC) risk among chronic hepatitis B (CHB) patients. We externally validated the predictive performance of the FSAC prediction model for HCC using on-therapy NFM responses. Our multicenter study consecutively recruited treatment-na\u00efve CHB patients ( Chronic hepatitis B virus (HBV) infection is a major public health problem affecting approximately more than 250 million people worldwide; it remains a leading cause of hepatocellular carcinoma (HCC), especially in endemic areas such as Korea ,2,3,4,5.Many models have been developed to help with predicting the risk of HCC development among HBV patients with or without AVT. Since the prognostic role of baseline serum HBV-DNA levels has been substantially attenuated in the present era of potent nucleos(t)ide analogues, models established within one decade  have adopted the presence of baseline cirrhosis rather than virological factors ,14,15. MHere, we aimed to externally validate the predictive performance of the newly developed FSAC model in comparison with other risk prediction models assessed at one time-point in an independent HBV cohort treated with ETV, TDF, or TAF.\u00ae  at the time of AVT initiation, as described in previous reports. Cirrhosis was histologically or clinically diagnosed as follows: (1) platelet count of <150/\u00d7103/\u03bcL and imaging findings suggestive of cirrhosis, including a blunted, nodular liver edge accompanied by splenomegaly (>12 cm) or (2) clinical signs of portal hypertension, such as gastroesophageal varices ) and were compared using Student\u2019s The predictive performances of the risk scoring models for HCC development were assessed using Harrell\u2019s C-index, time-dependent area under the receiver operating characteristic curve (TDAUC) at 3-, 5-, and 8-years from the index date and integrated area under the receiver operating characteristic curve (iAUC). Furthermore, lower values for the Akaike information criterion (AIC) were considered reflective of a better discriminatory ability for each model. Model performance was presented graphically using calibration plots, which compared the model prediction probability with the actual probability of HCC development. Discrimination and calibration were evaluated using the bootstrap method with re-sampling 1000 times.Statistical differences in parameters of predictive performance between FSAC and the other HCC-risk prediction models were evaluated using the bootstrap method with re-sampling 1000 times. If 95% CIs contained zero, there was deemed to be no significant difference in the parameters of predictive performance between the two models.http://cran.r-project.org/, accessed on 10 August 2021). Two-sided p-values < 0.05 were considered to indicate statistical significance.All statistical analyses were conducted using R package ; and higher median values of liver stiffness (14.3 vs. 7.7 kPa), FIB-4 (3.38 vs. 2.11), and APRI , compared to those without HCC   months, HCC developed in 303 (10.0%) patients (1.82 per 100 patient-years), and the cumulative probabilities of HCC development at 3-, 5-, and 8-years were 2.5%, 7.5%, and 15.3%, respectively. Patients who developed HCC were predominantly male (73.3% vs. 60.0%) and showed a significantly older age (median: 55.0 vs. 49.0 year); higher prevalence of cirrhosis (79.5% vs. 42.2%), diabetes mellitus (31.0% vs. 16.6%), hypertension (42.5% vs. 22.1%), and HBeAg negativity  . Likewisp = 0.001) and D , whose FIB-4 values at 12 months were \u22651.45, compared to those without , 952 (31.5%), 87 (2.9%), and 852 (28.2%) were classified into groups A, B, C, and D, respectively. Patients who developed HCC showed a higher prevalence of being classified into groups B  and D , whose APRI values at 12 months were \u22650.5, compared to those without , 733 (24.2%), 402 (13.3%), and 490 (16.2%) were classified into groups A, B, C, and D, respectively. Patients who developed HCC showed a higher prevalence of being classified into groups B . Subsequent multivariable analysis revealed that older age , male sex , presence of cirrhosis , higher liver stiffness values , hypertension , and lower platelet counts  were independently associated with an increased risk of HCC development , 0.769 (95% CI 0.745\u20130.791), 0.768 (95% CI 0.745\u20130.790), 0.768 (95% CI 0.743\u20130.789), and 0.769 (95% CI 0.744\u20130.791), respectively. The calibration plots for predicting 3-, 5-, and 8-year HCC development showed that the predicted probabilities were very close to the observed incidences .Harrell\u2019s c-index values for PAGE-B, modified PAGE-B, modified REACH-B, LSM-HCC, and CAMD were 0.725 (95% CI 0.699\u20130.750), 0.738 (95% CI 0.712\u20130.764), 0.737 (95% CI 0.710\u20130.763), 0.734 (95% CI 0.706\u20130.762), and 0.742 (95% CI 0.715\u20130.768), respectively, whereas their iAUC values were 0.718 (95% CI 0.693\u20130.741), 0.722 (95% CI 0.701\u20130.744), 0.724 (95% CI 0.699\u20130.748), 0.731 (95% CI 0.705\u20130.755), and 0.743 (95% CI 0.717\u2013 0.768), respectively . The FSAHarrell\u2019s c-index and iAUC values of FSAC (2) model were 0.763 (95% CI 0.737\u20130.787) and 0.763 (95% CI 0.739\u20130.784), respectively .n = 1391, 46.0%) showed that the Harrell\u2019s c-index and the iAUC value of the FSAC model were 0.668 (95% CI 0.633\u20130.701) and 0.661 (95% CI 0.627\u20130.694), respectively : scores 0\u20132); intermediate- (n = 1172 (38.2%): scores 3\u20138); and high-risk groups (n = 1009 (33.3%): scores 9\u201312), according to a previous study [p < 0.001, n = 0), 0.3% (n = 2), and 1.2% (n = 5) in the low-risk group; 2.1% (n = 23), 5.2% (n = 48), and 11.1% (n = 75) in the intermediate-risk group; and 5.2% (n = 49), 15.5% (n = 126), and 29.8% (n = 196) in the high-risk group, respectively (both p < 0.001 between each adjacent pair).Using FSAC score, we stratified patients into the three risk groups: low-  occurred during the median follow-up period of 64.0 months, allowing for highly acceptable statistical power. Second, among the low-risk group defined according to FSAC score (n = 845), only 6 patients developed HCC, with an annual incidence of 0.15%. Considering that current surveillance strategies to detect early-stage HCC may be cost-effective when annual risk exceeds at least 0.2% in patients without cirrhosis and 1.5% in patients with cirrhosis, the 27.9% of individuals in the low-risk group could likely avoid biannual abdominal ultrasonography-based HCC surveillance safely. Conversely, the intermediate- and high-risk groups, accounting for the remaining 72.1% of this study population, may require more delicate surveillance, given the suboptimal diagnostic sensitivity of abdomen ultrasonography and the overall poor prognosis of HCC detected at advanced stages. Thus, in order to achieve higher detection rates of HCC at early stages and greater cost-effectiveness, further studies on how to implement individualized surveillance strategies based on optimal visit intervals and the adoption of novel diagnostic modalities using radiology and/or serum biomarkers are warranted.Our study had several clinical implications. First of all, the prognostic performance of the FSAC model over other HCC-risk prediction models was reproduced. The large sample of >3000 patients with long-term follow-up enhanced the statistical reliability of the results. Moreover, a sufficient number of HCC cases ) which included only NFMs assessed at 12 months, also showed a Harrell\u2019s c-index value similar to the FSAC model . The observations in this study are consistent with reports of excellent predictive performance for the CAGE-B and SAGE-B models that incorporate liver stiffness values on transient elastography after 5 years of AVT.n = 1102, 36.4%) [p < 0.001). Hence, further large-scale study with the serial follow-up of transient elastography during long-term AVT should be required.Although the predictive performance of FSAC model among our study population was acceptable, further research using novel biomarkers that can exclude the overestimation caused by active necro-inflammation before starting AVT  should be required in the near future , in orde, 36.4%) , we confOur study has several limitations. First, although liver stiffness values by transient elastography were available in all patients at the baseline, approximately two thirds of patients did not undergo transient elastography during long-term AVT, primarily because it is not reimbursed by the National Health Insurance Service in Korea. Since the predictive performances might vary depending on the sample size and HCC incidence, further studies are required in the setting of paired transient elastography tests. Second, when the HCC prediction models were assessed among a subgroup with cirrhosis, their prognostic performances were generally attenuated. This is most likely because the discriminatory power of the variables constituting the models  might become considerably attenuated in the relatively \u201chomogenous\u201d cirrhotic subgroup. Given that the majority of HCC rises in the setting of cirrhosis, especially among patients in Western countries, further studies concerning development of novel biomarkers should be required, allowing the optimized prognostication in a subgroup with cirrhosis.In this external validation study of a large-scale cohort, the FSAC model exhibited more robust predictive performance for HCC development, compared to other HCC-risk prediction models that use only baseline characteristics. For predicting HCC development, accurate measurement of fibrotic burden during long-term AVT is necessary."}
+{"text": "Pseudomonas aeruginosa. The activity of the investigational combination cefepime-taniborbactam (FTB) and comparator agents was evaluated against clinical isolates of Enterobacterales from a 2018-2020 global surveillance study.Taniborbactam (formerly VNRX-5133) is a novel cyclic boronate-based broad-spectrum \u03b2-lactamase inhibitor with potent and selective direct inhibitory activity against both serine- and metallo-\u03b2-lactamases . Taniborbactam restores the activity of cefepime against many difficult to treat organisms, including cephalosporin- and carbapenem-resistant Enterobacterales and MICs of cefepime with taniborbactam fixed at 4 \u00b5g/mL and comparators were determined following CLSI M07-A11 guidelines against 10,543 Enterobacterales. Isolates were from community and hospital infections collected from 259 sites in 56 countries in 2018-2020. Resistant phenotypes were based on 2021 CLSI breakpoints. A set of 827 isolates with meropenem MIC \u22654 \u00b5g/mL (n=421) or with cefepime and/or ceftazidime MIC \u22652 \u00b5g/mL (n=406) was evaluated for the presence of MBLs, KPC, ESBLs, and OXA-48 group genes via PCR and sequencing. Forty-eight isolates with FTB MIC values of 16 \u00b5g/mL or greater were interrogated by WGS.50/90 values of 0.06/0.25 \u00b5g/mL and 99.5% inhibited at \u22648 \u00b5g/mL. FTB maintained activity against MBL-, KPC-, OXA-48 group, and ESBL-positive isolates . Isolates with elevated FTB MICs had IMP-type enzymes, variation in the cefepime target (penicillin binding protein 3), permeability defects in combination with acquired \u03b2-lactamases, and/or possible up-regulated efflux.Overall, 23.0% and 15.9% of isolates were nonsusceptible (NS) to cefepime and piperacillin-tazobactam (TZP), respectively (Table). FTB had potent activity against all Enterobacterales, with MICResults Tablein vitro activity of cefepime against Enterobacterales, including isolates nonsusceptible to recently-approved BL/BLI combinations and expressing serine and metallo-\u03b2-lactamases. This support the continued development of FTB as a potential new treatment option for challenging infections due to resistant Gram-negative pathogens.Taniborbactam significantly restored the Meredith Hackel, PhD MPH, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Mark G G. Wise, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor) Daniel F. Sahm, PhD, IHMA (Employee)Pfizer, Inc. (Independent Contractor)"}
+{"text": "Mycobacterium kansasii\u2014termed rBCG-Mkan85B\u2014which was used together with a booster immunization with plasmid DNA expressing the same M. kansasii Ag85B gene (DNA-Mkan85B). We reported that rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost immunization elicited various NTM strain-specific CD4+ and CD8+ T cells and induced Mycobacterium\u00a0tuberculosis-specific immunity. In this study, to investigate the protective effect against M. kansasii infection, we challenged mice vaccinated with a rBCG-Mkan85B or rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost strategy with virulent M. kansasii. Although BCG and rBCG-Mkan85B immunization each suppressed the growth of M. kansasii in the mouse lungs, the rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost vaccination reduced the bacterial burden more significantly. Moreover, the rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost vaccination induced antigen-specific CD4+ and CD8+ T cells. Our data suggest that rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost vaccination effectively enhances antigen-specific T cells. Our novel rBCG could be a potential alternative to clinical BCG for preventing various NTM infections.The incidence of infections with nontuberculous mycobacteria (NTM) has been increasing worldwide. The emergence of multidrug-resistant NTM is a serious clinical concern, and a vaccine for NTM has not yet been developed. We previously developed a new recombinant Bacillus Calmette\u2013Gu\u00e9rin (rBCG) vaccine encoding the antigen 85B (Ag85B) protein of  Mycobacterium tuberculosis complex and Mycobacterium leprae, such as Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium kansasii. Recently, the incidence of NTM infection has been increasing worldwide [Nontuberculous mycobacteria (NTM) are mycobacterium species other than the orldwide ,2,3. Altorldwide ,5,6,7,8.Since NTM pathogens are intracellular, cell-mediated immunity plays a major role in protecting against NTM. Previous reports have indicated that protective immunity against tuberculosis (TB) and NTM infection might overlap . EpidemiM. kansasii\u2014termed rBCG-Mkan85B\u2014which we used with a booster vaccination with plasmid DNA expressing the same M. kansasii Ag85B gene (DNA-Mkan85B) [M. tuberculosis and M. leprae [+ T cells and CD8+ T cells [d)-restricted epitopes that induced cross-reactive responses to M. tuberculosis and other related mycobacteria, including M. kansasii, in both BALB/c (H2d) mice and CB6F1 (H2b/d) mice [d-restricted peptide epitopes elicited polyfunctional CD8+ T cell responses that were also highly cross-reactive with those of other proteins in the Ag85 complex [M. kansasii, we challenged rBCG-Mkan85B/DNA-Mkan85B-prime\u2013boost-vaccinated CB6F1 mice with M.kansasii and evaluated the protective effect and cellular immunity in this study.Previously, we developed a new recombinant BCG (rBCG) vaccine encoding the antigen 85B (Ag85B) protein of Mkan85B) . The Ag8Mkan85B) . The Ag8. leprae ,16. We r. leprae . We also T cells . Moreove/d) mice . The H2- complex . TherefoSpecific pathogen-free female CB6F1 mice, aged between 6 and 8 weeks, were purchased from Japan SLC Inc. . All animal studies were carried out under institutional guidelines approved by the Nihon University Animal Care and Use Committee (AP19MED029-3), the institutional committee for gene recombination experiments (2018MED21), and biorisk management and control (30-10-4). The institutional animal experimental guidelines are in accordance with the ILAR Guide. Mice were allowed free access to sterile water and standard mouse food, and their physiological conditions were assessed every few days.We used the previously prepared rBCG-Mkan85B strain and DNA-Mkan85B plasmid  in this BCG was cultured in Middlebrook 7H9 broth (Difco) supplemented with albumin dextrose complex (ADC) enrichment (Difco) and 0.05% Tween 80 at 37 \u00b0C. rBCG-Mkan85B was cultured in Middlebrook 7H9 broth (Difco) supplemented with ADC enrichment (Difco), 0.05% Tween 80, and 10 mg/mL kanamycin at 37 \u00b0C. Bacterial culture density was monitored by measuring the absorbance at 470 nm and 600 nm.6 CFU or 0.1 mg of bacilli intradermally (i.d.), and 100 \u00b5g of plasmid DNA in saline intramuscularly (i.m.) three times [Mice were immunized with the BCG vaccine or rBCG-Mkan85B at a concentration of 4 \u00d7 10ee times  . The bacteriological experiment was carried out under institutional guidelines approved by Nihon University biorisk management and control (30-10-4). M. kansasii was grown in Middlebrook 7H9 broth (Difco) supplemented with ADC enrichment (Difco) with 0.05% Tween 80 at 37 \u00b0C and harvested during the exponential growth stage. Infection of mice with M. kansasii was conducted intratracheally. Mice were anaesthetized with isoflurane and inoculated with 50 \u00b5L of a bacterial suspension containing 1 to 10 \u00d7 104 CFU of M. kansasii using a yellow pipet tip.At 6 weeks after infection, the mice were sacrificed, and the numbers of viable bacilli in their lungs were estimated. Lung tissues were homogenized with sterilized water. Tenfold serial dilutions of homogenate were inoculated onto duplicate 7H10-OADC agar plates (Difco) to determine bacterial loads. Colonies were counted after incubation for 3 weeks at 37 \u00b0C.Fractioned lung samples from each mouse were fixed with 10% neutral-buffered formalin and embedded in paraffin wax. The sections from these tissues were 4 mm thick and stained with hematoxylin and eosin (H&E) or with the Ziehl-Neelsen stain for acid-fast bacilli.+ epitope peptide (peptide-25: FQDAYNAAGGHNAVF) [M. kansasii. A seven-color flow cytometry panel was used to simultaneously analyze multiple cytokines at the single-cell level. The gating strategy used to identify cytokine-producing CD8+ and CD4+ T cells in mouse splenocytes is shown in Splenocytes and immune cells isolated from the lung were stimulated with a 9-mer Ag85B CD8 epitope peptide (YYQSGLSIV) (Pep8) , 15-mer GGHNAVF) , or tubeGGHNAVF) ,17. PPD All comparisons among recombinant and control groups and among immunization groups were conducted using one-way ANOVA tests with the Tukey-Kramer test using the JMP program (SAS Institute) and R version 4.0.3 (R core team 2020). Data are expressed as the mean \u00b1 SD.M. avium and M. abscessus [M.kansasii infection.According to previous epidemiological reports, BCG vaccination can reduce the risk of NTM infection in humans ,20,21. Ibscessus . TherefoM.kansasii strain for infection for another 6 weeks. The immunization schedule is shown in 4 (\u00b11.5 \u00d7 104) pulmonary CFU per animal. Although BCG and rBCG-Mkan85B vaccination both reduced the pulmonary CFU  (p < 0.01) (The CB6F1 (H2b/d) mice were either left unimmunized or immunized with BCG or rBCG-Mkan85B for 6 weeks, followed by a nasal exposure to a virulent  < 0.01) , the eff < 0.01) A,D. Acid < 0.01) G,J. Alth < 0.01) B,C, the  < 0.01) H,I.M.kansasii infection. The CB6F1 mice were either left unimmunized or immunized with BCG or rBCG-Mkan85B/DNA-Mkan85B and then challenged with a virulent M.kansasii strain for another 6 weeks. The immunization schedule is shown in Next, we evaluated the efficacy of the rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost vaccination against To analyze the adaptive immune responses induced by BCG, rBCG-Mkan85B, and rBCG-Mkan85B/DNA-Mkan85B vaccination, we examined the induction of antigen-specific polyfunctional T cells using an intracellular cytokine staining (ICS) method.M. kansasii infection  [M. malmoense, M. marinum, or M. kansasii [M. avium and M. abscessus cross-reactive T cells in humans [+ T cell epitopes of the mycobacterial Ag85 complex elicited similar levels of cytokine production by polyfunctional T cells in mice vaccinated with an rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost strategy [Generally, NTM patients are considered to have been infected by environmental NTM, not by NTM from other patients. In other words, NTM do not cause airborne or droplet infections, unlike TB and SARS-CoV-2, the latter of which is responsible for coronavirus disease 2019 (COVID-19). However, increases in NTM incidence rates have been reported recently [ex (MAC) , M. malmn humans . Previoustrategy .M. kansasii. We showed that both BCG and rBCG-Mkan85B could protect the mice from virulent M. kansasii infection. However, contrary to our expectation, the efficacy of rBCG-Mkan85B vaccination was not superior to that of BCG vaccination. An ICS analysis revealed that M. kansasii antigen-specific polyfunctional CD4+ T cells were equally induced by BCG and rBCG-Mkan85B vaccination. However, similar to previous reports [M. kansasii peptides could not induce antigen-specific polyfunctional CD8+ T cells. Moreover, the present study demonstrated that rBCG-Mkan85B vaccination alone was not sufficient to induce M. kansasii Ag85B-specific polyfunctional CD8+ T cells. Therefore, we immunized mice with rBCG-Mkan85B and boosted them with DNA-Mkan85B three times. BCG and recombinant BCG prime\u2013boost immunization were reported to be beneficial strategies for inducing antigen-specific CD8+ T cells [+ and CD4+ T cells. Regarding antigen-specific CD4+ T cells, several critical roles for eliminating mycobacteria have been reported [+ T cell counts have a high risk of developing NTM infection. In addition to that of CD4+ T cells, the importance of CD8+ T cells to protecting against mycobacterial infection was revealed by previous studies by other researchers. For instance, the airborne infection of mice [M. avium induced the activation of both CD4+ T cells and CD8+ T cells. However, the significance of CD8+ T cells in protection against NTM infection, especially M. avium infection, has been controversial. Although the depletion or genetic elimination of CD8+ T cells significantly decreases immunological responses to TB and increases susceptibility to M. tuberculosis [+ T cells was reported to be unrelated to M. avium infection [+ T cells occurred during experimental M. avium infection [+ T cells in M. kansasii infection has not been investigated thoroughly. In the present study, although BCG or rBCG-Mkan85B immunization could activate only M. kansasii-specific polyfunctional CD4+ T cells, rBCG-Mkan85B/DNA-Mkan85B prime-boost vaccination was able to elicit M. kansasii-specific polyfunctional CD8+ T cells. Nevertheless, the BCG- or rBCG-Mkan85B-immunized mice were shown to be protected against M. kansasii infection, and the rBCG-Mkan85B/DNA-Mkan85B prime-boost strategy drastically reduced the bacterial burden in the lungs of the mice. The results suggest a role for antigen-specific polyfunctional CD8+ T cells in protecting against M. kansasii infection.In the present study, we first challenged unvaccinated and BCG- or rBCG-Mkan85B-vaccinated mice with virulent  reports ,22,23,25 T cells ,17,24. Ireported ,26,27. I of mice , calves  of mice , and rherculosis ,31, the nfection . Gilbertnfection . On the + and CD8+ T cells [Since the end of 2019, COVID-19 has been a serious public health concern worldwide. Looking back on history, human beings have suffered from many kinds of infections. We have to continue to fight various pathogenic microorganisms and infectious diseases. Vaccines are the gold standard for protecting against infectious diseases. One of the aims of a vaccine is to induce effective neutralizing antibodies against the target pathogen. To protect against several pathogenic microbes, such as mycobacteria, the activation of cellular immunity is required in addition to the induction of humoral immunity. BCG has been used globally as a live attenuated vaccine against TB for a century. Various recombinant BCG strains have been designed, developed, and evaluated as novel vaccines against various infectious diseases. For instance, several rBCG strains expressing HIV antigens, such as HIV gag and env, have been reported to induce anti-HIV neutralizing antibodies and antigen-specific CD4 T cells ,33,34. W T cells ,36,37. MM. kansasii infection by inducing M. kansasii-specific polyfunctional CD4+ T cells. Furthermore, rBCG-Mkan85B/DNA-Mkan85B prime\u2013boost vaccination induced antigen-specific CD4+ and CD8+ T cells and drastically reduced the CFU of M. kansasii in the lungs of vaccinated mice. Our data suggest that the combination of rBCG expressing Ag85B derived from M. kansasii with DNA that also expresses Ag85B derived from M. kansasii may be effective in enhancing antigen-specific T cells, resulting in more efficient control of M. kansasii infection.In conclusion, we demonstrated that BCG and rBCG-Mkan85B immunization protected CB6F1 mice from"}
+{"text": "The Centers for Disease Control and Prevention (CDC) estimated direct healthcare costs of &271 million in 2018. CDC 2015 guidelines  recommended cephalosporin plus azithromycin for GC. We used real-world data to assess patterns of inappropriate Table 1) during the first uUGG episode . Generalized linear models were used for multivariate analysis.A retrospective cohort study of IBM MarketScan data  in patients \u2265 12 years old with uUGG. Eligible patients had an AB prescription \u00b15 days of uUGG diagnosis (index date) and continuous health-plan enrollment with \u2265 6 months\u2019 baseline/\u2265 12 months\u2019 follow-up data. Patients with complicated urogenital GC were excluded. Patients were stratified by AB prescription , 77.1% had an IA/SO prescription (mostly due to IA AB class [~82.0%] and duration [24.0%]), while only 22.9% had an AP&OP prescription; uUGG episodes were more frequent with IA/SO (n=2386) than AP&OP (n=714) prescriptions during follow-up. Patients with IA/SO prescriptions had higher GC-related total adjusted costs per patient (PP) per index episode (&196) vs those with AP&OP prescriptions . Patients with IA/SO prescriptions also had higher GCrelated total adjusted costs PP during follow-up (&220) vs those with AP&OP prescriptions , mostly driven by higher outpatient ambulatory and emergency room (ER) adjusted costs with IA/SO  vs AP&OP prescriptions . ER visits PP at index and during follow-up were higher with IA/SO vs AP&OP prescriptions .Figure. GC-related costs per patient with uUGG, stratified by appropriateness of AB prescription*Table 2. GC-related HRU per patient with uUGG, stratified by AB prescriptionMost patients with uUGG were not prescribed treatments in accordance with CDC 2015 guidelines. High IA/SO AB prescriptions and associated healthcare costs suggest an unmet need for improved prescribing practices for uUGG in the US.Madison T. Preib, MPH, STATinMED Research  Fanny S. Mitrani-Gold, MPH, GlaxoSmithKline plc.  Ziyu Lan, MSc, STATinMED Research  Xiaoxi Sun, MA, STATinMED Research  Ashish V. Joshi, PhD, GlaxoSmithKline plc."}
+{"text": "The predictive value of silver stained nucleolar organiser regions (AgNORs) was assessed in 229 patients with transitional cell bladder cancer followed up for over 10 years. The AgNORs were enumerated in pretreatment biopsy specimens. The AgNORs were related to clinical stage (T) (P = 0.0111), papillarity (P less than 0.0001), WHO grade (P less than 0.0001), DNA ploidy (P = 0.0010) and S-phase fraction (P less than 0.0001). Tumours presenting with pelvic lymph node involvement (P = 0.0085) or metastasis (P = 0.0780) at the time of diagnosis had more AgNORs than tumours confined to the bladder wall. Progression in T-, N- and M-categories (P = 0.0010-0.0030) was related to AgNORs and consequently they predicted bladder cancer related survival (P = 0.0005). The diploid tumours could be regrouped according to survival by AgNORs (P = 0.0001). In papillary tumours AgNORs predicted progression (P = 0.0110) and survival (P = 0.0038). In Ta-T1 tumours AgNORs predicted progression (P = 0.11) and survival (P = 0.0751) and also in T2-T3 tumours AgNORs contributed to survival significantly (P = 0.0039). The AgNORs subdivided WHO grade III tumours according to their ability to progress during the follow-up time (P = 0.0711). In a multivariate analysis AgNORs predicted progression independently in Ta-T1 category (P = 0.0165). AgNORs predicted recurrence free period like SPF (P = 0.0010). In conclusion, AgNORs are inferior to classic prognostic factors or DNA flow cytometric variables in muscle invasive bladder cancers whereas they have independent predictive value in superficial cancers."}
+{"text": "The role of cholecystokinin (CCK) has been explored in pancreatic carcinogenesis following pancreatobiliary diversion (PBD), using the specific CCK receptor antagonist CR-1409. Male Wistar rats (n = 80) weighing 70-100 g were given weekly i.p. injections of azaserine (30 mg kg-1 week-1) for 3 consecutive weeks. One week later animals were randomised to receive either PBD or sham PBD and thereafter to receive s.c. injections of either saline or CR-1409 . Six months after operation surviving rats were killed as follows: sham + saline 20, PBD + saline 19, sham + CR-1409 14, PBD + CR-1409 11. Cardiac blood was taken for CCK assay and the pancreas was excised for wet weight measurement and quantitative estimation of atypical acinar cell foci (AACF), the precursor of carcinoma. PBD reduced median body weight (3-20% less than shams) but trebled the absolute and relative pancreatic weights (P < 0.001). CR-1409 blunted this adaptive response to PBD, reducing absolute pancreatic weight by 35% (P < 0.005). PBD quadrupled circulating CCK concentrations, regardless of the antagonist treatment. Acidophilic AACF occurred only in rats with PBD. CR-1409 markedly reduced the number of observed acidophilic AACF by 90% (P < 0.001) and the number of foci per pancreas by 93% (P < 0.001). Moreover, CR-1409 reduced the mean focal diameter of each lesion by 18% (P < 0.005), the mean focal volume by 58% (P < 0.05) and the percentage of pancreas occupied by acidophilic foci by 95% (P < 0.001). PBD enhances pancreatic carcinogenesis by causing hypercholecystokininaemia, and CR-1409 largely inhibits this enhancement."}
+{"text": "Cytokines are small proteins that regulate immunity in vertebrate species. Marsupial and eutherian mammals last shared a common ancestor more than 180 million years ago, so it is not surprising that attempts to isolate many key marsupial cytokines using traditional laboratory techniques have been unsuccessful. This paucity of molecular data has led some authors to suggest that the marsupial immune system is 'primitive' and not on par with the sophisticated immune system of eutherian  mammals.IFN-\u03b3, IL-2, IL-4, IL-6, IL-12 and IL-13, in the genome of the grey short-tailed opossum (Monodelphis domestica). Many of these genes were not predicted in the publicly available automated annotations.The sequencing of the first marsupial genome has allowed us to identify highly divergent immune genes. We used gene prediction methods that incorporate the identification of gene location using BLAST, SYNTENY + BLAST and HMMER to identify 23 key marsupial immune genes, including The power of this approach was demonstrated by the identification of orthologous cytokines between marsupials and eutherians that share only 30% identity at the amino acid level. Furthermore, the presence of key immunological genes suggests that marsupials do indeed possess a sophisticated immune system, whose function may parallel that of eutherian mammals. The marsupial and eutherian  lineages diverged approximately 180 million years ago. Marsupials are chiefly distinguished from other mammals by their unique reproductive strategies, with young born in an immature state with only the most rudimentary neurological and immunological systems , T Cell However, conventional experimental strategies using degenerate primers for reverse-transcriptase polymerase chain reaction (RT-PCR) and heterologous probes for screening genetic libraries have only identified the most phylogenetically conserved immune molecules, with cytokines proving particularly difficult to isolate . To datein vitro Mixed Lymphocyte Response -x(3)-L) is conserved . Inverted triangles indicate cysteine residues that are conserved across species. Dots represent identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (Q9HBE4), Mus musculus (NP_068554.1), Gallus gallus (NP_001020006.1), Canis familiaris (NP_001003347.1), Sus scofa (Q76LU6), Bos taurus (Q76LU5).Click here for fileSyntenic region between human chromosome 4q27 and opossum chromosome 5, illustrating the gene cluster of interleukin 2 and 21. Transcriptional directions are indicated by arrows.Click here for fileIL-2R\u03b3 amino acid sequences. Conserved cysteine residues are marked with an inverted triangle. Dots represent identity to Monodelphis domestica sequence. Completely conserved residues are shaded. Sequences used for alignment: Homo sapiens (NP_000197.1), Mus musculus (NP_038591.1), Gallus gallus (NP_989858.1), Rattus norvegicus (NP_543165.1), Canis familiaris (NP_001003201.1), Sus scrofa (NP_999248.1), Bos taurus (NP_776784.1).Click here for fileAlignment of IL-5 amino acid sequences. Dots represent identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_000870.1), Macaca mulatta (NP_001040598.1), Bos taurus (NP_776347.1), Canis familiaris (NP_001006951.1), Mus musculus (NP_034688.1), Macropus eugenii (AAD37462.1), Gallus gallus (NP_001007085.1).Click here for fileSyntenic region between human chromosome 5q23.3 and opossum chromosome 1, illustrating the gene cluster of interleukin 5, 4 and 13. Transcriptional directions are indicated by arrows.Click here for fileAlignment of IL-6 amino acid sequences. Residues involved in receptor binding in human IL-6 are denoted with diamonds. Cysteine residues conserved among all species are marked with an inverted triangle. PROSITE family motif is boxed. Dots represent identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_000591.1), Mus musculus (NP_112445.1), Oryctolagus cuniculus (Q9MZR1).Click here for fileAlignment of IL-12\u03b1 amino acid sequences. Cysteine residues conserved among all species are marked with an inverted triangle. Dots represent identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_000873.2), Mus musculus (NP_032377.1), Gallus gallus (NP_998753.1), Rattus norvegicus (NP_445842.1), Ovis aries (NP_001009736.1) Canis familiaris (NP_001003293.1).Click here for fileAlignment of IL-10 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_000563.1), Mus musculus (NP_034678.1), Gallus gallus (NP_001004414.1), Trichosurus vulpecular (AAD01799), Canis familiaris (NP_001003077.1), Sus scofa (Q29055), Cervus elaphus(P51746).Click here for fileAlignment of IL-19 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_037503.2), Mus musculus (NP_001009940.1).Click here for fileAlignment of IL-20 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_061194.2), Mus musculus (NP_067355.1), Tetraodon nigroviridis (AAP57416.1).Click here for fileAlignment of IL-24 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_006841.1), Mus musculus (NP_444325.1), Rattus norvegicus (NP_579845.1), Tetraodon nigroviridis (AAP57418.1).Click here for fileAlignment of IL-26 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_060872.1), Danio rerio (NP_001018635.1).Click here for fileAlignment of IL-22 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_065386.1), Mus musculus (NP_058667.1), Sus scofa (AAX33671.1), Rattus norvegicus (ABF82262.1), Danio rerio (NP_001018628.1).Click here for fileNeighbour-Joining tree of IL-10 family ligand protein sequences rooted by midpoint. JTT amino acid substitution matrix was used and 500 bootstrap replicates performed. Branches supported by bootstrap values over 70 are in bold. Opossum sequences are marked by triangles. Sequences used for this analysis were Homo sapiens IL-10 (NP_000563.1), IL-19 (NP_715639.1), IL-20 (NP_061194.2), IL-22 (NP_065386.1), IL-24 (NP_006841.1), IL-26 (NP_060872.1); Mus musculus IL-10 (NP_034678.1), IL-19 (NP_001009940.1), IL-20 (NP_067355.1), IL-22 (NP_058667.1), IL-24 (NP_444325.1); Rattus norvegicus IL-24 (NP_579845.1); Sus scofa IL-10 (Q29055); Bos taurus IL-10 (P43480); Trichosurus vulpecular IL-10 (AAD01799); Gallus gallus IL-10 (NP_001004414.1); Cyprinus carpio IL-10 (BAC76885.1); Tetraodon nigroviridis IL-10 (CAD67786.1); Takifugu rubripes IL-10 (CAD62446.1) Danio rerio IL-26 (NP_001018635.1) and Monodelphis domestica. Sequences labels in the tree are abbreviated by the first letter from the genus with the first two letters from the specific name followed by the gene name.Click here for fileAlignment of CD4 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_000607.1), Mus musculus (NP_038516.1), Macaca mulatta (BAA09671.1) Felis cattus (NP_001009250.1), Rattus norvegicus (NP_036837.1) Oncorhynchus mykiss (AAY42068.1).Click here for fileAlignment of CD8 amino acid sequences. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_001759.3), Mus musculus (Q60965), Gallus gallus (NP_990566.1), Canis familiaris (NP_001002935.1), Sus scofa (NP_001001907.1), Rattus norvegicus (AAH88126.1).Click here for fileAlignment of IFNGR-2 amino acid sequences. Cysteine residues conserved among all species are marked with an inverted triangle. Dots indicate identity to Monodelphis domestica sequence. Sequences used for alignment: Homo sapiens (NP_005525.2), Mus musculus (NP_032364.1), Gallus gallus (NP_001008676.1).Click here for file"}
+{"text": "Animals were inoculated either with 10(6) or 10(7) MAT-LyLu cells, or with saline to serve as controls. Carcass weight in tumour-bearing (TB) animals decreased despite similar food and water intake in both groups. Absence of metastatic tumour cells from HL of all TB animals was confirmed by histological examination. Twenty-one days after inoculation, 31P MRS showed a 2.5-fold increase in [Pi]/[ATP] ratios in HL in vivo (P < 0.001) which was confirmed by 31P MRS of liver extracts in vitro (P < 0.005). Phosphodiester to ATP ratios were significantly increased (P < 0.05) in HL in vivo, but absolute PDE levels were similar in both groups. Phosphomonoester to ATP ratios did not change, although absolute phosphomonoester levels in HL were reduced by -41% (not significant). In HL extracts in vitro, sharp reductions in the levels of glucose-6-phosphate (P < 0.05), fructose-6-phosphate (P = 0.05), phosphocholine (P < 0.001), glycerophosphocholine (P < 0.001), and glycerophosphoethanolamine (P < 0.001) were observed. Electron microscopy revealed increased amounts and altered distribution of rough endoplasmic reticulum in HL. These findings show that experimental prostate cancer significantly affects hepatic phosphorylation status, phospholipid metabolism, and gluconeogenesis in the host animal, and demonstrate the value of combined MRS in vivo and in vitro in monitoring HL metabolism in cancer."}
+{"text": "A longitudinal study of iron status markers (haemoglobin (Hb), serum (S-) iron, S-transferrin, transferrin saturation, S-ferritin) was performed in 31 chemotherapy treated patients with small cell lung cancer. At discovery, eight patients were anaemic (Hb less than 121 g l-1). Hb, S-iron and transferrin saturation were lower (P less than 0.01), and S-ferritin higher (P less than 0.01) than in healthy subjects. Chemotherapy induced an immediate fall in Hb (P less than 0.003), increase in S-iron (P less than 0.003) and transferrin saturation (P less than 0.001). Later in the disease a fall in S-transferrin (P less than 0.006) and an increase in S-ferritin (P less than 0.02) occurred. Thirty patients died during the 2 years observation. S-ferritin at discovery was correlated to performance status score  and to survival . Patients with S-ferritin less than or equal to 400 micrograms l-1 (n = 13) had longer survival than those with S-ferritin greater than 400 micrograms l-1 (n = 18) (P = 0.004)."}
+{"text": "Sequential exposure to these factors , however, resembling the order of secretion during liver embryogenesis, induced both glycogen-storage and cytokeratin (CK)18 expression. Additional exposure of the cells to trichostatin A (TSA) considerably improved endodermal differentiation, as evidenced by acquisition of an epithelial morphology, chronological expression of hepatic proteins, including hepatocyte-nuclear factor (HNF)-3\u03b2, alpha-fetoprotein (AFP), CK18, albumin (ALB), HNF1\u03b1, multidrug resistance-associated protein (MRP)2 and CCAAT-enhancer binding protein (C/EBP)\u03b1, and functional maturation, i.e. upregulated ALB secretion, urea production and inducible cytochrome P450 (CYP)-dependent activity.hMSC are able to undergo mesenchymal-to-epithelial transition. TSA is hereby essential to promote differentiation of hMSC towards functional hepatocyte-like cells. By utilizing in vitro models, pharmaceutical companies attempt to reduce clinical failure rates by accurately evaluating efficacy and safety much earlier in the drug discovery process , CD45-PE (DAKO), CD105-FITC , CD73-PE, CD90-PE, CD166-PE  and analyzed using a flow cytometer . Mesodermal differentiation was verified by means of different stainings (described below). To exclude spontaneous differentiation, hMSC were cultivated in regular MSCGM medium (Cambrex) and analyzed in parallel.3 cells/cm2 on 1 mg/ml collagen gel type I (BD) in basal medium supplemented with 2% (v/v) foetal bovine serum . Basal medium consisted of 60% (v/v) DMEM and 40% (v/v) MCDB-201  supplemented with 100 IE/ml penicillin , 100 \u03bcg/ml streptomycin, 1 mg/ml linoleic-acid bovine serum albumin, 0.1 mM L-ascorbic acid, 0.03 mM nicotinamide, 0.25 mM sodium pyruvate and 1.623 mM glutamine . At 100% confluence, cells were exposed to hepatogenic cytokines and growth factors, added either simultaneous as a cocktail ('referred to as 'cocktail-condition/-set-up/-exposure' or a derivative thereof)  or sequentially  . Differentiation media were changed every 3 days. From day 6 onwards, 1 \u03bcM TSA (Sigma) was added. The cocktail- and sequential-conditions, supplemented with 1 \u03bcM TSA will be further referred to as 'cocktail TSA-condition/-set-up/-exposure', and 'sequential TSA condition/-set-up/-exposure, respectively, or a derivative hereof.hMSC were plated at 21.5 10Cells were fixed with 10% (w/v) formalin  for 10 min at room temperature  or with methanol  for 2 min at -20\u00b0C (osteogenic differentiation). After fixation, adipocytes were identified as red-colored lipid droplets upon staining with Oil-red O (from Sigma) . MineralChondrocytic differentiation was determined using rabbit polyclonal anti-collagen II antibody .Cells were fixed either with ethanol (Merck) for 10 min at -20\u00b0C  or with 4% (w/v) paraformaldehyde  for 10 min at 4\u00b0C (nuclear and cytoplasmic markers). After fixation, liver-specific protein expression was analysed using primary antibodies against AFP (goat), (HNF3\u03b2 (goat), HNF1\u03b1 (rabbit), C/EBP\u03b1 (rabbit), MRP2 , CK18  and ALB . HDAC inhibition was assessed using anti-acetyl histone H4 antibody . Respective secondary antibodies came from Jackson Immunoresearch, Cambridgeshire, UK. As a negative control, cells were incubated with appropriate gamma immunoglobulines (Jackson Immunoresearch) and immunostained under the same conditions. Cells were analysed using fluorescence microscopy with a Zeiss Axiovert scope.Cell morphology was analysed using phase-contrast light-microscopy (Nikon).ALB concentrations, secreted into the culture media, were analysed by ELISA .4Cl (Sigma), were measured colometrically in the culture media according to the manufacturer's instructions . Fresh culture media supplemented with 6 mM NH4Cl (Sigma) and 4 hours-cultured adult hepatocytes were used as negative and positive controls, respectively.Urea concentrations produced, after 24 hours-exposure of the cells to 6 mM NHEROD- and PROD-activities were assessed as previously described  with somTo evaluate the inducibility of CYP2B6 and CYP1A1/2, cells were, after 18 days of differentiation, exposed to PB  and MC , respectively. Media, supplemented with either PB or MC, were daily renewed. Fresh culture media and 4 hours-cultured adult hepatocytes were used as negative and positive controls, respectively.Results are expressed as mean \u00b1 SD. Statistical analyses were performed using Oneway Anova and Student's t-test. The significance level was set at 0.05.Albumin (ALB); alpha-fetoprotein (AFP); CCAAT-enhancer binding protein (C/EBP); cytochrome P450 (CYP); cytokeratin (CK); dimethylsulfoxide (DMSO); ethoxyresorufin-O-deethylase (EROD); fibroblast growth factor-4 (FGF-4); hepatocyte growth factor (HGF); hepatocyte nuclear factor (HNF); histone deacetylase inhibitor (HDAC-I); human adipose tissue-derived stromal cells (hADSC); human mesenchymal stem cells (hMSC); insulin-transferrin-sodium-selenite (ITS); lactate dehydrogenase (LDH); multidrug resistance-associated protein (MRP); multipotent adult progenitor cells (MAPC); methylcholantrene (MC); new chemical entities (NCEs); oncostatin M (OSM); pentoxyresorufin-O-dealkylase (PROD); Periodic-acid-Schiff (PAS) staining; phenobarbital (PB); trichostatin A (TSA).SS: Differentiation of expanded hMSC under hepatocyte-specific conditions. Analysis of multilineage  differentiation by means of staining, functionality assays, microscopic analysis. Preparation of the manuscript.TV, PP and MV: Optimization of the concentration of TSA and onset of exposure to TSA. Critical revision of the manuscript. ADB and IVR: Isolation, expansion and routine characterization of hMSC . Critical revision of the manuscript. VR: Head of department of Toxicology and promotor of the hereby associated PhD thesis. Microscopic analysis. Critical revision of the manuscript.All authors read and approved the final manuscript.3H] thymidine  incorporation into trichloroacetic acid precipitated-DNA  was measured using a scintillation counter . Graphs shown are representative for 3 independent experiments.DNA synthesis of sequentially (+/-1 \u03bcM TSA) and cocktail (+/-1 \u03bcM TSA)-exposed hMSC. hMSC, plated on 1 mg/ml collagen gel type I, were at 100% confluence treated with either the cocktail- or sequential-condition. From day 6 onwards, 1 \u03bcM TSA was added . Differentiation media were changed every 3 days. Samples for DNA synthesis were taken 2, 4, 6, 12 and 18 hours upon media change. [Methyl-Click here for fileCell viability analysis of sequentially (+/-1 \u03bcM TSA) and cocktail (+/-1 \u03bcM TSA)-exposed hMSC. hMSC, plated on 1 mg/ml collagen gel type I, were at 100% confluence treated with either the cocktail- or sequential-condition. From day 6 onwards, 1 \u03bcM TSA was added . LDH leakage into culture media was measured throughout culture time according to the Bergmeyer procedure  using a commercial kit . Graphs shown are representative for at least 3 independent experiments.Click here for fileCell death analysis of sequentially (+/-1 \u03bcM TSA) and cocktail (+/-1 \u03bcM TSA)-exposed hMSC. hMSC, plated on 1 mg/ml collagen gel type I, were at 100% confluence treated with either the cocktail- or sequential-condition. From day 6 onwards, 1 \u03bcM TSA was added . Differentiation media were changed every 3 days. Cells were, 12 hours upon media change, incubated with Alexa Fluor 488 annexin V (green fluorescent), propidiumiodide (red fluorescent) and the nuclear counterstain DAPI (blue fluorescent). The red, green, and both red and green-stained cells represent necrotic, apoptotic and death cells, respectively. 20 \u00d7 10 original magnification, phase contrast. Stainings shown are representative for at least 3 separate experiments.Click here for file"}
+{"text": "PLoS Genetics, vol 2, issue 4, DOI:  10.1371/journal.pgen.0020053In http://www.ncbi.nlm.nih.gov) accession numbers for the sequence discussed in this paper are HVS1 (AY72095-AY721592)\u201d should be \u201cThe GenBank (http://www.ncbi.nlm.nih.gov) accession numbers for the sequence discussed in this paper are HVS1 (AY720957-AY721592).\u201dIn the Supporting Information section, the sentence \u201cThe GenBank ("}
+{"text": "The expression of oestrogen receptor protein (ER) was examined in 151 cases of symptomatic or screening detected pure ductal carcinoma in situ (DCIS) of the breast by immunocytochemical assay (ERICA), in formalin-fixed paraffin-embedded tissue, with the monoclonal antibody H 222 (Abbott). Forty-eight tumours (31.8%) of cases were ER positive. Twenty-seven (17.9%) of cases showed high level ER expression and 21 (13.9%) of cases showed low level ER immunoreactivity. Significant associations of positive tumour ER immunoreactivity and non-comedo architecture chi 2 = 6.76; (d.f. = 1): P < 0.001, small cell size chi 2 = 4.49; (d.f. = 1): P = 0.034, higher S-phase fraction chi 2 = 4.71; (d.f. = 1): P = 0.03 and lack of c-erbB-2 protein overexpression chi 2 = 7.96; (d.f. = 1): P < 0.01 were identified. No significant associations of ER expression and patient age, histological grade of necrosis in DCIS, or DNA ploidy were found. ER expression is detectable in less than one third of symptomatic and screening detected cases of DCIS, implying that endocrine therapy of DCIS may be a more appropriate form of management for morphological subtypes of DCIS which show higher rates of oestrogen receptor expression, particularly those of non-comedo and small cell type."}
+{"text": "Scientific Reports 10.1038/s41598-018-20137-2, published online 29 January 2018Correction to: This Article contains errors in References 8 and 9 which are incorrectly given as:et al. Colistin hetero-resistance in multidrug-resistant Acinetobacter baumannii clinical isolates from the Western Pacific region in the SENTRY antimicrobial surveillance programme. J Infection58, 138\u2013144, 10.1016/j.jinf.2008.11.002 (2009).\u2019\u2018Yau, W. et al. Synergistic killing of NDM-producing MDR Klebsiella pneumoniae by two \u2018old\u2019 antibiotics-polymyxin B and chloramphenicol. J Antimicrob Chemoth70, 2589\u20132597, 10.1093/jac/dkv135 (2015).\u2019\u2018Rahim, N. A. 2.The correct references are listed below as refs"}
+{"text": "Microbacterium paludicola CC3 exhibits the capability to produce polysaccharide bioflocculants. Here, we report the whole-genome sequence of M.\u00a0paludicola CC3, which may be helpful in understanding the genetic basis of the biosynthesis of polysaccharide bioflocculants as well as in promoting its production and application in industrial fields. Paenibacillus shenyangensis, Agrobacterium tumefaciens F2, and Paenibacillus wulumuqiensis  RSII platform using P6-C4 chemistry. The resulting sequencing reads with 320.4-fold coverage were then de novo assembled using Hierarchical Genome Assembly Process (HGAP) (http://www.ncbi.nlm.nih.gov/COG/), and the Gene Ontology (GO) Consortium (http://www.geneontology.org/). The metabolic pathways were predicted using the KEGG Automatic Annotation Server (KAAS) (http://www.genome.jp/tools/kaas/). rRNAs and tRNAs were identified using Barrnap 0.4.2 (http://www.vicbioinformatics.com/software.barrnap.shtml) and tRNAscan-SE version 2.0 (http://lowelab.ucsc.edu/tRNAscan-SE/), respectively. The clustered regularly interspaced short palindromic repeat (CRISPR) elements were detected using PILER-CR  , 9. Genes (HGAP) . FunctioPILER-CR .M.\u00a0paludicola CC3. No plasmid sequences were detected. The chromosome was composed of 3,410,829\u00a0bp, with an average G+C content of 70.10%, which comprised 3,390 predicted genes, of which 3,209 were protein coding genes (CDSs), 32 were tRNA genes, 146 were rRNA genes, and 3 were microRNA genes. Pseudogenes and prophage genes were not identified, whereas 14 CRISPR candidates were detected in the genome of strain CC3. A series of genes encoding polysaccharide biosynthesis/modification proteins, such as mannose-1-phosphate guanylyltransferase .The sequence data for the genome of"}
+{"text": "AbstractPhlaeothripinae, Mystrothripslevissp. n. and Urothripslancangensissp. n., are described from China. Pentagonothripsantennalis Haga & Okajima and Plectrothripsbicolor Okajima are newly recorded in China.Two new species of fungivorous  Phlaeothripinae belong to Phlaeothrips lineage, which are usually taken from dead branches or leaf-litter and feed on fungal hyphae .All thrips specimens were extracted by using Tullgren funnels from leaf litter, and then sorted and preserved in 90% alcohol. Examined specimens were mounted into Canada balsam using the method outlined by Taxon classificationAnimaliaThysanopteraPhlaeothripidaehttp://zoobank.org/5399B364-0B5C-4B37-A2F5-896C751EF8A6. Holotype. Female aptera: CHINA, Guangdong: Guangzhou City, South China Botanical Garden (23Guangdong: Guan), in leaf litter of bamboo, 9.viii.2014 (Chao Zhao).Paratypes. 8 females 2 males, collected with holotype; 5 females 1 male, the same locality but collected on 20.xi.2015 (Chao Zhao).Female aptera . Distended body length 2030. Head length 190, width 185; eyes length 50; postocular setae length 75. Antennae length 470, segments I\u2013VIII length (width) as follows: 49(43); 53(41); 68(32); 67(34); 65(34); 61(30); 54(24); 52(12). Pronotum median length 140, width across median part 310; length of major setae: pronotum anteromarginal setae 48, anteroangular setae 68, midlateral setae 90, posteroangular setae 85, epimeral setae 80. Metanotum median setae 40. Pelta length 100, width at base 170. Abdominal tergite IX S1 setae length 190, intermediate setae length 85, S2 length 190. Tube length 155, width at base 93, at apex 42; anal setae length 135.Male aptera . Distended body length 1620. Head length 160, width 155; eyes length 35; postocular setae length 70. Antennae length 375, segments I\u2013VIII length (width) as follows: 34(36); 44(29); 50(32); 50(31); 55(28); 50(25); 45(20); 45(13). Pronotum median length 125, width across median part 260; length of major setae: pronotum anteromarginal setae 60, anteroangular setae 45, midlateral setae 73, posteroangular setae 66, epimeral setae 63. Metanotum median setae 30. Pelta length 65, width at base 110. Abdominal tergite IX setae S1 length 145, intermediate setae length 60, S2 length 45. Tube length 130, width at base 80, at apex 32; anal setae length 110.China (Guangdong).levis, is from the Latin adjective, meaning \u201csmooth\u201d, and refers to the dorsal surface of head and pronotum which are largely smooth. In contrast, most species of this genus are sculptured with distinct polygonal reticulation on head and pronotum.The specific epithet, Mystrothrips  to Japan  head and pronotum largely smooth ; (2) antennae uniformly brown (versus antennae segments I and II distinctly lighter than remaining segments in M.flavidus); (3) pelta semicircular with short lateral lobes ; (4) S1 and S2 setae on abdominal tergite IX much longer than tube (vs shorter than tube in M.flavidus).Of the seven species worldwide listed in the genus rothrips , two areto Japan . The newPageBreakTaxon classificationAnimaliaThysanopteraPhlaeothripidaeHaga & OkajimaPentagonothripsantennalis Haga & Okajima, 1979: 147.CHINA, Hunan: 2 females and 1 male, Zhuzhou City, Yanling County, Shennong Valley , in leaf litter, 16. ix. 2014 (Chao Zhao). Hubei: 1 male, Huanggang City, Yingshan County, Taohuachong , in leaf litter, 23.iv.2014 (Chao Zhao).Dorsal surface of body entirely reticulate; head longer than width, cheeks distinctly incut behind eyes; postocular setae well-developed with expanded at apex. Antennae 7-segmented, morphological segments VII and VIII fused with an incomplete suture, segments III and IV with two and three sense cones, respectively. Maxillary stylets short, wide apart. Pronotum with five pairs of well-developed major setae, strongly expanded at apex. Basantra absent. Fore tarsal tooth present in both sexes. Pelta transverse, shaped as a squashed ellipse, with distinctly polygonal reticulation. Sternite VIII with pair of stout or leaf-like posteromarginal setae submedially. Tube short than head. Pore plate of male absent.China ; Japan.Pentagonothrips, was originally established from Japan ; mesothoracic furcae closely fused together medially as well as metathoracic furcae joined together medially (but metathoracic furcae separated in Mystrothrips).The monobasic genus, om Japan . P.anteTaxon classificationAnimaliaThysanopteraPhlaeothripidaeOkajimaPlectrothripsbicolor Okajima, 1981: 313.CHINA, Guangdong: 1 female, Guangzhou City, Arboretum of South China Agricultural University , in leaf litter, 20.xi.2004 (Jun Wang); 1 male, Guangzhou City, Dafushan Forest Park , in leaf litter of Litchichinensis,17.iv.2016 (Chao Zhao).Body bicolored, yellow and brown. Head, thorax and tube brown, abdomen yellowish brown; all legs yellow; antennal segments II and III yellow, remaining segments brown. Head longer than broad, dorsal surface smooth except weakly sculptured posterolaterally. Antennal segments III and IV with two and three sense cones, respectively, segment VI with two sense cones. Maxillary stylets short, maxillary bridge developed and arched. Pronotum smooth, surrounded by stippled membrane with a distinct median longitudinal line. Metanotum with longitudinal striae medially. Mid tibia and hind tibia with one and two apical spur-like stout setae, respectively. Forewing parallel-sided with seven duplicated cilia. Pelta irregularly triangular with slender lateral lobes and a pair of campaniform sensilla. Abdominal tergites II\u2013VII each with a pair of wing retaining setae; sternites V\u2013VII with a pair of worm-like reticulate areas in both sexes; tergite IX S1 and S2 setae pointed, S1 setae longer than S2 but shorter than tube; tergite IX in male with a small median projection on posterior margin.China (Guangdong); Japan; Indonesia.P.bicolor, originally described from Japan and Indonesia . Holotype. Female aptera, CHINA, Yunnan province, Pu\u2019er City, Lancang County, Nuozhadu Nature Reserve , 5.xi.2016 (Chao Zhao).Paratypes. 6 females, 3 males, collected with holotype.Female aptera . Body length 1400. Head length 180; maximum width 190. Pronotum length 110; median width 250; epimeral setae 20. Metathoracic epimeral setae 20. Abdominal tergite IX length 120, basal width 75, distal width 40. Tube length 130, basal width 22, apical width 25; anal setae 430. Antennal segments I\u2013VIII length (width) as follows: 20(36), 28 (31), 37 (23), 39 (24), 45 (20), 40 (15), 47 (12).Male aptera. . Body length 1050. Head length 160; maximum width 160. Pronotum length 90; median width 185; epimeral setae 13. Metathoracic epimeral setae 13. Abdominal tergite IX length 105, basal width 55, distal width 40. Tube length 115, basal width 20, apical width 22; anal setae 370. Antennal segments I\u2013VIII length (width) as follows: 22(33), 23 (31), 32 (19), 29 (22), 33 (20), 31(17), 39(13).China (Yunnan).The specific epithet is named after the type locality, Lancang County, Yunnan Province, China.Urothripstarai , particularly in the shape of antennae, but it can be differentiated from the latter by the following diagnostic characters: (1) head broadly rounded in front (vs slightly produced in U.tarai); (2) dorsal surfaces of head and pronotum largely sculptured with polygonal reticulation (vs head and pronotum distinctly tuberculate and without reticulation in U.tarai); (3) major body setae on head, pronotum, especially on abdominal tergites are stout, dilated and fan-shaped at apex ; (4) fore femora brown (while fore femora yellow in U.tarai).There are ten species recognized in this genus , of whicPageBreak"}
+{"text": "Negative, cognitive and depressive symptoms, as well as physical comorbidities, have a great impact on the real-world functioning in patients with schizophrenia (SZ) . However, not all the studies have employed accurate psychometric instruments to assess these symptoms, nor have all these factors been studied simultaneously.The aim of the current study is to analyze the determinants of functionality in SZ measured by the Personal and Social Performance (PSP) scale, and considering not exclusively psychopathological and cognitive variables, but also aspects related to physical health and inflammation.Sample: 73 outpatients with SZ, duration of illness \u226410 years, under stable maintenance treatment .Clinical variables: PANSS, CGI-Severity, Clinical Assessment Interview of Negative Symptoms (CAINS) -Motivation/Pleasure (MAP) & Expression (EXP) domains-, Brief Negative Symptom Scale (BNSS), Calgary Depression Scale (CDS), MATRICS Consensus Cognitive Battery (MCCB), PSP.Biological variables: Glucose, cholesterol, LDL, HDL, triglycerides, TSH, prolactin, insulin, uric acid, alkaline phosphatase (APh), C-reactive protein (CRP), TNF-\u03b1, interleukin(IL)-6, IL-2, IL-1\u03b2, IL-1RA, homocysteine, HT (% hemolysis), lipid peroxidation (LPO), catalase.Pearson correlations were performed to select variables significantly related to PSP scores which were later included in stepwise multiple linear regression analyses. Age, sex, education, smoking, alcohol use, BMI, antipsychotic equivalent doses and other confounding factors were considered.Final model for PSP total score  identified that CGI-Severity (\u03b2= -0.279), PANSS-NM (negative Marder Factor) (\u03b2 = -0.218), Asociality subscale of BNSS (\u03b2= -0.383) and IL-2 (\u03b2= -0.269) were significant predictors.Predicting variables included in regression models for specific PSP domains:- Self-care : PANSS-NM (\u03b2=0.458), Avolition subscale of BNSS (\u03b2=0.248), IL-2 (\u03b2=0.221), APh (\u03b2=0.201).- Useful activities : CGI-S (\u03b2=0.245), Avolition (\u03b2=0.557).- Social relationships : Asociality (\u03b2=0.578), PANSS-GP (\u03b2=0.276), CAINS-EXP (\u03b2=0.178).- Aggresive behaviour : PANSS-P (\u03b2=0.408), CDS (\u03b2=0.273).1.Negative symptoms are the most important determinants of a deficit in the real-world functioning in SZ, especially \u201casociality.\u201d2.\u201cApathy\u201d has a negative impact on self-care and useful activities domains.3.Proinflammatory cytokine IL-2 marks poor functionality.1. Harvey (2014). Assessing disability in schizophrenia: tools and contributors. J Clin Psychiatry.2. Strassnig et al. (2015). Determinants of different aspects of everyday outcome in schizophrenia: The roles of negative symptoms, cognition, and functional capacity. Schizophr Res.3. Menendez-Miranda et al. (2015). Predictive factors of functional capacity and real-world functioning in patients with schizophrenia. Eur Psychiatry."}
+{"text": "Marinomonas fungiae strain AN44T was isolated from mucus of the coral Fungia echinata. Optimum growth occurs at 3 to 5% NaCl. The draft genome is 4.2\u2009Mb, with 3,776 protein-coding genes. It harbors genes for the degradation of aromatic compounds, such as quinate, ferulate, p-coumarate, protocatechuate, and p-hydroxyphenylacetate. Marinomonas fungiae strain AN44T (JCM 18476T) is an aerobic, Gram-negative, motile, and rod-shaped bacterium belonging to the class Gammaproteobacteria and was isolated from mucus of the coral Fungia echinata from the Andaman Sea (N44T JCM 8476T is aman Sea .M. fungiae strain AN44T was generated at the DOE Joint Genome Institute (JGI) using the Illumina HiSeq 2000 platform (https://github.com/lh3/wgsim), and AllPaths-LG , respectively. The draft genome contains a total of 3,776 CDSs, 78 pseudogenes, 53 tRNAs, 16 rRNAs , 5 noncoding RNAs (ncRNAs), and 1 clustered regularly interspaced short palindromic repeat (CRISPR). Based on COG functional categories, the CDSs of the M. fungiae genome were distributed into categories of amino acid transport and metabolism (9.84%), carbohydrate transport and metabolism (5.92%), cell cycle control, cell division, and chromosome partitioning (1.08%), cell motility (3.51%), cell wall/membrane/envelope biogenesis (5.06%), chromatin structure and dynamics (0.09%), coenzyme transport and metabolism (5.47%), defense mechanisms (2.18%), energy production and conversion (6.61%), extracellular structures (0.44%), function unknown (5.41%), general function prediction only (7.31%), inorganic ion transport and metabolism (5.19%), intracellular trafficking, secretion, and vesicular transport (1.55%), lipid transport and metabolism (3.35%), mobilome prophages and transposons (1.08%), nucleotide transport and metabolism (2.47%), posttranslational modification, protein turnover, and chaperones (4.15%), RNA processing and modification (0.03%), replication, recombination, and repair (3.83%), secondary metabolite biosynthesis, transport, and catabolism (2.12%), signal transduction mechanisms (8.13%), transcription (7.97%), translation, ribosomal structure, and biogenesis (7.18%), and unassigned (28.88%).The genome was annotated using the JGI Microbial Genome Annotation Pipeline . Genes wp-coumarate, along with a single operonic gene cluster for central aromatic compound degradation via the protocatechuate meta-cleavage pathway, were found in the genome. Additionally, a single operonic gene cluster for the degradation of p-hydroxyphenylacetate was found.Further, putative genes or gene clusters for peripheral aromatic compound degradation pathways of quinate, ferulate, and LIQF00000000. The version described in this paper is the first version, LIQF01000000, and consists of sequences LIQF01000001 to LIQF01000036.This whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession number"}
+{"text": "Nucl. Acids Res. (04 January 2016) 44 (D1): D447\u2013D456. doi: 10.1093/nar/gkv1145.3Medizinisches Proteom Center (MPC), Ruhr-Universit\u00e4t Bochum, D-44801 Bochum, Germany.The authors wish to correct the affiliation of one of the authors. Gerhard Mayer's only affiliation should be 1European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SD, UK.He does NOT have a second affiliation at: The authors apologise to the readers for this error."}
+{"text": "With limited and low-genetic barrier drugs used for the prevention of mother-to-child transmission (PMTCT) of HIV in sub-Saharan Africa, vertically transmitted HIV-1 drug-resistance (HIVDR) is concerning and might prompt optimal pediatric strategies.The aim of this study was to ascertain HIVDR and viral-tropism in majority and minority populations among Cameroonian vertically infected children.A comparative analysis among 18 HIV-infected children (7 from PMTCT-exposed mothers and 11 from mothers without PMTCT-exposure) was performed. HIVDR and HIV-1 co-receptor usage was evaluated by analyzing sequences obtained by both Sanger sequencing and ultra-deep 454-pyrosequencing (UDPS), set at 1% threshold.10 copies/mL, and 526 (282\u2013645) cells/mm3, respectively. All children had wild-type viruses through both Sanger sequencing and UDPS, except for 1 PMTCT-exposed infant harboring minority K103N (8.31%), born to a mother exposed to AZT+3TC+NVP. X4-tropic viruses were found in 5 of 15 (33.3%) children (including 2 cases detected only by UDPS). Rate of X4-tropic viruses was 0% (0/6) below 5 years , and became relatively high above 5 years  versus CD4 >15% ; similarly for CD4 \u2264200 (3/4 [75%]) versus CD4 >200  age, viremia, and CD4 count were 6 (4\u201310) years, 5.5 (4.9\u20136.0) logNGS has the ability of excluding NRTI- and NNRTI-mutations as minority species in all but 1 children, thus supporting the safe use of these drug-classes in those without such mutations, henceforth sparing ritonavir-boosted protease inhibitors or integrase inhibitors for the few remaining cases. In children under five years, X4-tropic variants would be rare, suggesting vertical-transmission with CCR5-tropic viruses and possible maraviroc usage at younger ages. From these observations, we postulated that minority DRMs in HAART-na\u00efve children might grow-up through selective drug-pressure and populate plasma in a short-frame, herein justifying the rapidly emerging DRMs we observed at failure. Although not yet clinically endorsed, pediatric minority DRMs might be more concerning in the context of PMTCT, henceforth underscoring an unmet clinical need.,11 Coupled to previous knowledge on the detection of DRMs by next-generation sequencing (NGS),\u201314 we thus hypothesized that using NGS to assess DRMs in vertically infected HAART-na\u00efve children would contribute in designing long-term HAART strategies for SSA-children.As the footprint of long-term HAART depends largely on the effectiveness of first-line drugs in sustaining viral suppression, establishing adequacy between pediatric HAART and DR-mutations (DRMs) would be clinically relevant. As HAART would be reaching 1.5 million children by 2020, as high as 20% virological failure (VF) is expected, favored by high-viremia and poor adherence in children.,16 Without optimal strategies, VF would quickly overcome HAART success, maintaining children vulnerable.Current pediatric HAART-regimens consist of lamivudine (3TC), abacavir (ABC), or zidovudine (AZT), associated to ritonavir-boosted lopinavir (LPV/r) or NVP. LPV/r is recommended to overcome PMTCT-resulting non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance, whereas NVP matches with postnatal prophylaxis.,8 Although not yet approved for under 16 years, the CCR5 antagonist\u2014maraviroc\u2014might represent a suitable antiretroviral alternative for children, pending proof-of-concept towards relevant pediatric clinical trials. Particularly, there are limited evidence on the potential effectiveness of maraviroc for SSA-children in PMTCT, initial-HAART and/or following treatment-failure.\u201321 With rising concerns of minority variants on response to several classes of antiretrovirals, a genuine delineation of HIV-1 tropism, considering both minority and majority quasi-species,,23 could rationalize maraviroc suitability for pediatric HAART-policies in SSA.Moreover, pediatric HAART options are limited in SSA, urging the quest for a wider therapeutic portfolio.Based on these assumptions, we aimed to ascertain DRMs and HIV-1 co-receptor usage, in majority and minority viral populations, from children according to maternal PMTCT-exposure in a resource-limited setting (RLS).22.1A comparative study was conducted in 2015 among 18 HIV-1 vertically infected Cameroonian children, all HAART-na\u00efve, stratified according to maternal antiretroviral exposure during pregnancy: control-group  versus case-group (7 children from mothers exposed to reverse transcriptase inhibitors [RTIs]). For each child, a plasma sample was collected to perform both Sanger- and 454 ultra-deep pyrosequencing (UDPS).2.2 Briefly, viral RNA was extracted from plasma using QIAamp Viral RNA minikit , following manufacturer's instructions. PR/RT-containing region was then reverse-transcribed and amplified using SuperScript One-Step for long templates reverse transcriptase polymerase chain reaction (RT-PCR) of Invitrogen kit , with an eventual second-round seminested PCR. Direct sequencing was then performed using 7 overlapping primers.Protease (PR)/RT Sanger sequencing was performed as previously described. Briefly, viral RNA containing the V3-loop region was reverse-transcribed and amplified using an RT/Taq mix, with an eventual second-round seminested PCR. Direct sequencing was then performed using 4 overlapping primers.V3 loop Sanger sequencing was performed as previously described.2.32O DNase RNase free, 1\u200a\u03bcL forward primer (10\u200a\u03bcmol/L), 1\u200a\u03bcL reverse primer (10\u200a\u03bcmol/L), 1\u200a\u03bcL RNase Out (40\u200aU/\u03bcL Invitrogen) and 1.2\u200a\u03bcL RT/TAQ, for a final volume of 50\u200a\u03bcL. RT-PCR conditions were the following: 1 cycle 50\u00b0C, 30 minutes; 1 cycle 94\u00b0C, 2 minutes; 40 cycles ; a final extension 68\u00b0C, 10 minutes. Forward and reverse primers were respectively 5\u2019GACAGGCTAATTTTTTAGGG3\u2019  and 5\u2019GATAAATTTGATATGTCCATTG3\u2019 . Nested-mid PCR was then performed with the Fast Start HiFi PCR system  using 5 pairs of barcoded-modified forward and reverse primers for each amplicon , 8\u200a\u03bcL MgSO4 (5\u200ammol/L), 2.8\u200a\u03bcL H2.4Ten microliters viral RNA were reverse transcribed with 1-step RT-PCR system using forward  and reverse  primers, under the following conditions: 1 cycle 50\u00b0C, 30 minutes; 1 cycle 94\u00b0C, 2 minutes; 35cycles ; a final extension 68\u00b0C, 10 minutes. A nested mid-PCR was then performed with the Fast Start HiFi PCR system  as previously described.2.5PR/RT PCR products  and V3 loop (one fragment of 367 bps) were purified using Agencourt AMPure PCR purification beads  and quantified with Quant-iT PicoGreen double-stranded DNA assay kit  on a GloMax multidetection system . Phylogenetic analyses excluded any possible sample contamination (data not shown).Pooled purified PCR products were clonally amplified by emulsion PCR and pyro-sequenced on the 454 GS junior platform  as previously described.2.6The entire PR (amino acid position: 1\u201399), RT (1\u2013251) and the entire V3 loop (1\u201335) sequences obtained after 454-pyrosequencing were de-multiplexed and then quantified using the SFF tool Roche. Using a home-made Perl script and SHORAH package 0.5.1, sequences were filtered and corrected for homopolymeric region-associated errors and aligned against HIV-1 consensus B. Final alignments were manually checked for insertion or deletion in homopolymeric regions that could result in a frame shift. Nucleotidic/aminoacidic variants were evaluated and quantified by a home-made pearl script, and sequences were considered reliable when showed an intra-patient frequency \u22651% in both forward and reverse strands.2.7http://hivdb.stanford.edu/pages/download/resistanceMutations_handout.pdf) and geno2pheno.v2.5 (http://coreceptor.geno2pheno.org/), respectively. Using a quantitative interpretation, viruses were considered CXCR4-tropic (X4-variants) by UDPS when \u22652% viral species had a false-positive rate (FPR) \u22643.5%, or by Sanger sequencing when FPR was \u226410%, describing the probability of classifying an R5-virus falsely as an X4-variant.PR/RT DRMs and HIV-1 co-receptor usage were interpreted using Stanford HIVdb list  available at 2.9HIV-1 DRMs and coreceptor usage were compared between the two PMTCT-groups. Coreceptor results by Sanger sequencing and UDPS were considered concordant if viral-tropism was identical from both sequencing technologies. Viral-tropism was explored according to age and CD4 count.P values <.05 were considered statistically significant.All statistical analyses were performed using the statistical open source environment R.v.3.1.1. 2.10Ref.\u200aN\u00b0034/NEC/SE), proxy-informed consent was provided, unique identifiers were used for privacy and confidentiality, and a material transfer agreement was established.Ethical clearance was obtained from the Cameroon National Ethics Committee  cells/mm3, respectively, without any significant difference between the 2 groups (data not shown). In the control, neither children nor their mothers had any antiretroviral exposure. Antiretroviral history of children belonging to the case-group, considered at higher risk of HIVDR, is described in Table Overall, median (interquartile range [IQR]) age, viremia, and CD4 count were 6 (4\u201310) years, 5.5 (4.9\u20136.0) log3.2HIV-1 subtyping revealed 50% CRF02_AG (9/18), 33.3% F (6/18), 11.1% CRF01_AE (2/18), and 5.6% CRF11.cpx (1/18).3.3PR/RT sequences were successfully obtained both through Sanger sequencing and UDPS for 17/18 children. The median UDPS coverage was of 1642 (IQR: 1269\u20135193) reads. In the entire covered PR/RT regions, the 2 sequencing technologies showed total concordance in variants detection, and all UDPS variants with frequencies <20% were not detected by Sanger sequencing , an accessory polymorphism weakly selected under etravirine (ETR) and rilpivirine (RPV), was found in a child aged 8 years from the control group.By using UDPS, 1 (aged 1 year) of 7 children 14.3%) from the case-group harbored viruses with K103N , a nonpolymorphic mutation causing high-level resistance to NVP and efavirenz (EFV). This infant was born from an RTI-treated mother (AZT\u200a+\u200a3TC\u200a+\u200aNVP). Thus, Sanger sequencing and UDPS were performed also for the mother (ID-18613). UDPS revealed a virus harboring 2 major DRMs: L74\u200aV at minority-level (2.5%), causing high- and intermediate-level resistance respectively to didanosine and to ABC; Y181C at population-level (96.7%), causing high- and intermediate-level resistance respectively to NVP and to EFV, ETR, and RPV , a polymorphic accessory mutation selected under EFV, in a child aged 6 years Table .Other variants, found even at RTI-associated drug resistance positions, were with minimal or no effect on drug susceptibility or virological response. Of note, in either group, no major DRMs to ritonavir-boosted protease inhibitors (PI/r) were found by both Sanger sequencing and UDPS.3.4V3 loop sequencing was successful by both Sanger sequencing and UDPS for 15 of 18 children and the mother ID-18613, with an overall viral-tropism concordance of 87.5% 14/16) between Sanger sequencing and UDPS (Table /16 betweX4-tropic viruses were found in 5 of 15 (33.3%) children (including 2 cases detected only by UDPS), all aged above 5 years. Specifically, in 1 child (ID-11621) UDPS provided an added value in tropism-determination compared to Sanger sequencing. Indeed, a clinically relevant quantity of minority X4-tropic variants (frequency: 3.9%) was detected by UDPS in this child . In another child (ID-10196), despite an R5-tropism (FPR\u200a=\u200a79.7%) determined by Sanger sequencing, a discordant tropism was observed through UDPS with a high percentage of X4-tropic variants , because of insertions detected only at minority levels.P\u200a=\u200a.040). As expected, X4-tropic viruses were higher with CD4 \u226415% (4/9 [44.4%]) versus CD4 >15% ; similarly for CD4 \u2264200 (3/4 [75%]) versus CD4 >200 . No statistical difference was found in X4-variants between the 2 PMTCT-groups: 2 of 7 (28.6%) case group versus 3 of 8 (37.5%) control group, P\u200a=\u200a1.000.Of relevance, the rate of X4-tropic viruses was 0% (0/6) among children under 5 years , and became significantly higher as from 5 years and above , known to be associated with resistance to NNRTIs used both for PMTCT and first-line HAART in SSA, was found in a PMTCT-exposed infant, thus suggesting NNRTI-sparing regimens for such children.,30,34 Discrepancy in DRMs between mother and infant would be due to sample collection later after delivery (at the moment of infant HIV diagnosis), with possible selection following prophylaxis/breastfeeding; as previously reported in similar RLS . This infant (aged 1 year), compared to the median age of the study population (6 years), suggests that circulating DRMs might have fade-up with increasing age.,33 NNRTI mutations (E138A and V179D), found in children without PMTCT-exposure, are known as polymorphisms with little or no effect on drug susceptibility or virological response. The ability of NGS in excluding minority RTI-mutations  supports the safe use of NNRTIs/NRTIs in those without such mutations, thus sparing from inappropriate switch to PI/r- or integrase inhibitor-containing regimens.,17,33\u201335In this high CRF02_AG-infected population,,37 as well as a baseline FPR <60 as previously demonstrated.,39 Further investigations might help in establishing novel public health strategies for an eventual usage of maraviroc in children.,40 As current PMTCT-practice might not be an independent factor for viral-tropism , CCR5-antagonist (maraviroc) could be a useful therapeutic weapon for pediatric HAART.,18,40Coreceptor usage in these children provides a clue for clinical application. Indeed, X4-variants appeared to be associated with older ages and lower CD4 cells, suggesting limited vertical transmission by CXCR4-tropic viruses, and later appearance of X4-variants with chronicity, immunological impairment,,39 Interestingly, by detecting minority insertions associated with a complete discrepant result on Sanger sequencing, UDPS appears very useful in validating tropism determination for non-B subtypes.Of the two children showing discordant results between the two sequencing techniques, the added value of UDPS in detecting X4-tropic minority variants is in accordance with previous reports.,22,41Therefore, UDPS might provide additional information in detecting DRMs and viral-tropism, confirming the added value of this technology for both clinical diagnostics and management of non-B HIV-infected children.,43In spite of this added value of UDPS, implementing NGS is more challenging in RLS , suggesting the need for simpler and affordable approaches integrating minority variants (point-of-care or pragmatic sequencing).,13,44\u201346 This study therefore provides relevant data to be used as base for further/enlarged studies.A potential study limitation could be the relatively small sample size, which makes the study probability relatively large. Also, in the PMTCT-exposed group, only 3 of 7 were exposed to triple ART, calling for subsequent investigations with scale-up of option B+. Moreover, HIV-1 variants were investigated only in plasma compartment, suggesting the need for exploring HIV variability in several compartments  and the impact on treatment and monitoring strategies in SSA.In a nutshell, NGS could help in identifying PMTCT-exposed children harboring minority NNRTI-DRMs, therefore serving for a timely switch of treatment and limiting failure rate. NGS also reveals a possible absence of X4-variants among children below 5 years, thus suggesting possible public health approaches using maraviroc. These preliminary evidences, generated on a small sample of mainly CRF02_AG-infected individuals, merit further investigations for improved pediatric-HAART strategies in RLS.Conceptualization: A. Nanfack, C-F. Perno, C. Fokunang, C. Tangimpundu, D. Armenia, D. Takou, E. Temgoua, F. Ceccherini-Silberstein, G. Cappelli, J. Fokam, M-M. Santoro, P. Koki, V. Colizzi.Data curation: D. Armenia, F. Ceccherini-Silberstein, J. Fokam, L. Carioti, M. Bellocchi, M-M. Santoro.Formal analysis: D. Armenia, F. Ceccherini-Silberstein, J. Fokam, L. Carioti, M. Bellocchi, M-M. Santoro.Funding acquisition: C-F. Perno, J. Fokam, V. Colizzi.Investigation: A. Ndjolo, A. Nanfack, C-F. Perno, C. Fokunang, C. Tangimpundu, D. Takou, E. Temgoua, F. Ceccherini-Silberstein, G. Cappelli, J. Fokam, J. Torimiro, M-M. Santoro, P. Koki, V. Colizzi.Methodology: D. Armenia, D. Takou, F. Continenza, F. Ceccherini-Silberstein, J. Fokam, L. Carioti, M. Bellocchi, M-M. Santoro.Project administration: A. Ndjolo, C-F. Perno, C. Fokunang, F. Ceccherini-Silberstein, G. Cappelli, J. Fokam, J. Torimiro, P. Koki, V. Colizzi.Resources: A. Ndjolo, C-F. Perno, J. Fokam, V. Colizzi.Software: D. Armenia, D. Takou, F. Continenza.Supervision: A. Ndjolo, C-F. Perno, C. Fokunang, F. Ceccherini-Silberstein, G. Cappelli, M-M. Santoro, P. Koki, V. Colizzi.Validation: A. Ndjolo, C-F. Perno, C. Fokunang, C. Tangimpundu, D. Armenia, E. Temgoua, F. Continenza, F. Ceccherini-Silberstein, G. Cappelli, J. Fokam, J. Torimiro, L. Carioti, M. Bellocchi, M-M. Santoro, P. Koki, V. Colizzi.Visualization: A. Nanfack, C-F. Perno, C. Tangimpundu, D. Takou, E. Temgoua, F. Continenza, J. Fokam, M-M. Santoro.Writing \u2013 original draft: C-F. Perno, F. Ceccherini-Silberstein, J. Fokam, M-M. Santoro.Writing \u2013 review & editing: A. Ndjolo, A. Nanfack, C. Fokunang, C. Tangimpundu, D. Armenia, D. Takou, E. Temgoua, F. Continenza, G. Cappelli, J. Torimiro, L. Carioti, M. Bellocchi, P. Koki, V. Colizzi.We are appreciative to our institutional staff that participated locally in the enrolment and in sample processing. We thank Domenico Di Carlo for statistical analyses."}
+{"text": "Halopteris filicina (Grateloup) K\u00fctzing, Dictyota dichotoma (Hudson) J. V. Lamouroux, Posidonia oceanica (L.) Delile and Flabellia petiolata (Turra) Nizamuddin from the Adriatic Sea (single point collection). VOCs were investigated by headspace solid-phase microextraction (HS-SPME) and analysed by gas chromatography and mass spectrometry (GC-MS/FID). H. filicina headspace contained dimethyl sulfide , C8-compounds ), benzaldehyde , alkane C17, dictyopterene D and C , tribromomethane (V), 1-iodopentane, others. F. petiolata headspace was characterized by DMS (22.2%), 6-methylhept-5-en-2-one (9.5%), C17 (9.1%), II (6.5%), compounds I-V. DMS (59.3%), C15 (14.5%), C17 (7.2%) and C19 (6.3%) dominated in P. oceanica headspace. Sesquiterpenes were found in D. dichotoma, predominantly germacrene D (28.3%) followed by other cadinenyl (abundant), muurolenyl and amorphenyl structures. Determined VOCs may be significant for chemosystematics and chemical communications in marine ecosystem.Performed phytochemical study contributes to the knowledge of volatile organic compounds (VOCs) of  The research on the algae VOCs has been continued by different teams cyclohepta-1,4-diene). Proposed biogenesis of dictyopterenes starts from (3S)-1,cis-5-undecadien-3-ol and (3S)-1,cis-5,cis-8-undecatrien-3-ol by dehydration and cyclization.Carotenoid cleavage dioxygenases (CCDs) catalyze oxidative cleavage of carotenoids, resulting in the production of norisoprenoids\u2014apocarotenoids  via glycso occur . The bre2-unit, which can potentially occur via either the \u03b2-oxidative pathway or non-oxidatively [Benzenoid and phenylpropanoid volatile compounds, primarily derived from phenylalanine require shortening of the carbon skeleton side chain by a Cdatively .P. oceanica followed by F. petiolata and H. filicina indicating those plants as source of sulfur compounds in marine ecosystem. Their headspace contained individually variety of C8-compounds (e.g. fucoserratene), benzaldehyde, alkanes C15, C17 and C19, dictyopterene D and C, others. Sesquiterpenes were found in D. dichotoma, predominantly germacrene D indicating similarity to terrestrial aromatic plants. Identified VOCs contain different types of organic compounds that may be significant for chemosystematics and ecology .Considering limited data available on the chemical composition of marine plants from the Adriatic Sea, the present research is contribution toward their better chemical characterization. Significant differences were found among the headspace VOCs from 3 seaweeds and 1 seagrass. High abundance of DMS was found in"}
+{"text": "In t test)  .S1 Fign = 8 biological replicates per genotype. (B) Percent of male only Spen KD larvae floating. (C) Percent of female only Spen KD larvae floating. (D) FB-specific Spen KD  with different insertion site as Spen KD in Fig 1A compared to KD control (dcg>iw). (E) Genetic background controls (iSpen/+ and iw/+) for (D). (F) As the Spen hairpin insertion site appears to result in a lean phenotype, KD animals were normalized to their genetic background. (G) As in (A), three additional independent Spen hairpin constructs (dcg>iSpen) tested in different density solutions and compared to KD control (dcg>iw). (H) Genetic background controls (iSpen/+\u2019s and iw/+) for (G). P value obtained by ANOVA. *P < 0.05, ** P < 0.01, ***P < 0.001, **** P < 0.0001. Error bars represent SEM.(A) Percent of floating larvae in different density solutions. FB-specific Spen KD  as in Fig 1A with additional dcg/+ background control. Fifty larvae per genotype per experimental replicate, (TIF)Click here for additional data file."}
+{"text": "Klebsiella\u00a0pneumoniae infections. Herein, we report the draft genome sequences of two colistin-resistant K.\u00a0pneumoniae isolates (BA41763 and B6753). The sequence data indicate that BA41763 and B6753 contain genomes of ~5.9 and 5.7\u00a0Mb in size with several plasmids.Resistance to colistin is a major threat that limits therapeutic choices for treating carbapenem-resistant  Klebsiella pneumoniae (CRKP) infections are difficult to manage, with very few remaining treatment options, such as colistin and tigecycline  with 49\u00d7 and 23\u00d7 coverage, respectively. Assembled genome sequences were annotated in PATRIC, the bacterial bioinformatics database and analysis resource (http://www.patricbrc.org) (http://rast.nmpdr.org/) (http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html).To further understand the molecular mechanism behind colistin resistance, we determined their whole-genome shotgun sequences in Ion Torrent PGM using 400-bp chemistry. https://www.patricbrc.org). Similarly for B6753, 5,754,328\u00a0bp with 6,248 CDS were identified. Also identified were 13 rRNAs, 72 tRNAs with 34 and 82 ARDB and CARD antimicrobial resistance genes and 72 and 181 virulence factors from the VFDB and Victors database, respectively.The genome of BA41763 had 5,944,266\u00a0bp with 6,678 coding sequences (CDS), 10 rRNAs, and 66 tRNAs. The annotation revealed 32 and 85 ARDB and CARD antimicrobial resistance genes, respectively. In addition, 74 and 183 virulence factors from VFDB and the Victors database, respectively, were identified (https://cge.cbs.dtu.dk//services/MLST/) (https://cge.cbs.dtu.dk//services/ResFinder/) (rmtf and aacA4), beta-lactam , fluoroquinolones lb-cr, oqxA, qnrB66), fosfomycin (fosA), macrolide (mph(A)), rifampin (ARR-2), and sulfonamide (sul2) resistance genes. Similarly, for B6753, we found aminoglycoside -Vla), beta-lactam , fluoroquinolone (aac(6\u2032)lb-cr), fosfomycin (fosA), macrolide (mph(E), msr(E)), phenicol (catB3), sulfonamide (sul1), tetracycline (tet(D), and trimethoprim . However, plasmid-mediated colistin resistance genes mcr-1 and mcr-2 were absent. B6753 had a mutation in the mgrB gene with a premature stop codon resulting in a truncated protein of 27\u00a0amino acids, but there were no mutations in the mgrB gene of BA41763.Next-generation sequencing (NGS) also revealed that BA41763 and B6753 were of sequence types (STs) ST147 and ST14, respectively, as analyzed by the MLST 1.8 tool (https://cge.cbs.dtu.dk//services/PlasmidFinder/) (PlasmidFinder 1.3 (Finder/)  revealedLZYN00000000 and MEBR00000000, respectively. The versions described in this paper are the first versions, LZYN01000000 and MEBR01000000.This whole-genome shotgun project of both the isolates BA41763 and B6753 has been deposited at DDBJ/ENA/GenBank under the accession numbers"}
+{"text": "EditorThe following manuscript has been retracted from our November \u2013 December, 2016 issue. It has been retracted on a request from the authors as they forgot to properly acknowledge the source of their data. The authors are grateful to those who pointed out this mistake. - Retraction in: Pak J Med Sci 2016;32(6):1500-1505. doi: https://doi.org/10.12669/pjms.326.11460Link: http://pjms.com.pk/index.php/pjms/article/view/11460/4800Effective role of lady health workers in immunization of children in Pakistan1, Azka Naeem2, Unaiza Shahid3, Wajiha Noor Syed4, Urva Khan5, Nayyar Misal Zaidi6Saira AfzalRetracted on February 7, 2017"}
+{"text": "A 24-year-old male from Wangdue Phodrang district, Bhutan, presented with a history of unexplained intermittent low-grade fever and cough for 8 months. He also complained of weight loss, abdominal discomfort, and one episode of hemoptysis and convulsion. He was a cow herder by profession. A computed tomography scan showed hepatosplenomegaly  with mesenteric lymphadenopathy. Biochemical evaluation showed elevated alkaline phosphatase 1096 IU/L), lactate dehydrogenase 330 IU/L), and C-reactive protein (40.8 mg/dL) and reverse albumin/globulin ratio. Complete blood count showed pancytopenia confirmed by peripheral smear. In addition, left shift of neutrophils and giant platelets were observed. Erythrocyte sedimentation rate (26 mm/h) was increased. Prothrombin time (20 s) and activated partial thromboplastin time (51 s) were prolonged. The bone marrow aspirate smear revealed intracellular and extracellular Leishman-Donovan bodies . In view096 IU/L,30 IU/L,"}
+{"text": "C. elegans cell divisions that produce an apoptotic daughter cell exhibit Daughter Cell Size Asymmetry (DCSA), producing a larger surviving daughter cell and a smaller daughter cell fated to die. Genetic screens for mutants with defects in apoptosis identified several genes that are also required for the ability of these divisions to produce daughter cells that differ in size. One of these genes, ham-1, encodes a putative transcription factor that regulates a subset of the asymmetric cell divisions that produce an apoptotic daughter cell. In a survey of C. elegans divisions, we found that ham-1 mutations affect primarily anterior/posterior divisions that produce a small anterior daughter cell. The affected divisions include those that generate an apoptotic cell as well as those that generate two surviving cells. Our findings suggest that HAM-1 primarily promotes DCSA in a certain class of asymmetric divisions. Caenorhabditis elegans somatic development is essentially invariant. Almost all of the somatic divisions are asymmetric, generating two daughter cells that differ in fate  [4p::gfp] .LG II. ynIs25 [flp-12p::gfp] [2p::gfp] , 19.LG III. nIs107[tbh-1p::gfp + lin-15(+)] [rdvIs1 [egl-17p::mCherry:his-24 + egl-17p::myristolated mCherry + pRF4] [n-15(+)] , rdvIs1  + pRF4] .LG IV. ham-1(gm279) [ ham-1gm29 [7].LG V. zuIs178 [-119(+)] .LGX. gmIs81 [7p::gfp] .ltIs44[pie-1p-mCherry::PH(PLC1delta1) + unc-119(+)]  [lqIs80[scmp::gfp::CaaX] [Unmapped: -119(+)] , leIs270-119(+)] , lqIs80[p::CaaX] .kyEx581[ocr-4p::gfp + lin-15(+)] , Ex  [Extra-chromosomal arrays: ::dsRed] .C. elegans promoter. The A/PVM, SDQR/L, A/PQR and URXR/L neurons were detected using the gmIs81 reporter. The SMB, OLQ, ASK, MC and RIC neurons were detected using the reporters flp-12p::gfp, ocr-4p::gfp, srbc-66p::gfp, ceh-19p::gfp and tbh-1p::gfp, respectively. When an extra neuron was detected in a ham-1 mutant, its position was in close proximity to the normal position of the single neuron found in wild-type animals. Missing neurons were only scored when using integrated transgenes, since extra-chromosomal arrays can be lost during cell divisions. Statistical analysis was performed using the two-sample Z-test for proportions.All neurons were detected with transcriptional reporters that express fluorescent proteins under control of the indicated lqIs80[scmp::gfp::CaaX]. V5.pa lineage analysis was performed by compiling and reordering images of rdvIs1[egl-17p::myr mcherry + egl-17p::H2B::mcherry] L2 larvae. The mcherry markers are upregulated in all cells of the V5.pa lineage. V5.paa daughter cells size measurements were performed at the 3- and early 4-cell stages, before V5.paap and V5.paaa migrations occurred. T.pp and V5.pa neuroblast daughter cell sizes measurements were performed as previously described for the Q neuroblasts' daughters , which labels the cells of the P3-8 lineage. Ratios are indicated in the text \u00b1 standard deviation.T.p lineage analysis was performed in early L2 larvae using aughters , 12. TheFor embryonic neuroblasts, automated lineaging was performed as described . After nFor each genotype and ACD, the measured daughter cells size ratio was plotted on graphs where horizontal dotted lines indicate 1:1 ratios, and horizontal grey bars indicate the median of each ratio distribution. Statistical analysis was performed using the Mann-Whitney U-test.ham-1 mutants produce abnormal numbers of neurons in specific lineages . The cells boundaries are indicated by dotted lines and the name of each cell is indicated. Right panels are schematic representations of the corresponding cells. Black arrows describe cell displacements. Colored arrows indicate the direction of neurite extensions. Upon V5.pa division, the anterior daughter V5.paa moves to a more dorsal position above its sister V5.pap. The V5.paa neuroblast divides asymmetrically before V5.pap to produce the smaller anterior daughter V5.paaa, which will become the PDE neuron, and the larger posterior daughter V5.paap. The V5.pap glioblast's division produces daughter cells of equal size, the anterior daughter V5.papa, which will become the PDE socket cell, and the posterior daughter V5.papp, which will become the PDE sheath cell. Before the V5.paap division, both V5.paap and V5.paaa migrate anteriorly and ventrally, respectively. V5.paap then divides to generate the larger anterior daughter V5.paapa which will become the PVD neuron, and the smaller posterior daughter V5.paapp, which dies. After completion of all cell divisions, PDE and PVD neurons extend their neurites. Abbreviations: so: socket cell; sh: sheath cell. Scale bars: 10 \u03bcm.(A) Schematic diagram of the V5.pa lineage, which produces two neurons (PDE and PVD), two neuron-associated support cells (PDE socket cell and PDE sheath cell) and one apoptotic cell (V5.paapp). V5.paa and V5.paap daughters are unequal in size, whereas V5.pap division is symmetric in size. (B) Description of the cell division sequence and cell movements that occur during posterior deirid development. Left panels are representative fluorescent labeling of the left V5.pa lineage at all stages of development by (TIF)Click here for additional data file."}
+{"text": "Objectives. In this study, we assessed the extra-articular symptoms in constellation with selected serum cytokines and disease activity in spondyloarthritis (SpA). Patients and Methods. We studied 287 SpA patients: 131 had AS, 110 had PsA, and 46 had SAPHO. We assessed extra-articular symptoms in all cases. In 191 SpA patients, we measured serum interleukin-6 (IL-6), interleukin-18 (IL-18), interleukin-23 (IL-23), endothelin-1 (ET-1), vascular endothelial growth factor (VEGF), and epidermal growth factor (EGF). Results. Patients with acute anterior uveitis (AAU) had higher VAS (P = 0.0008), BADSDAI (P = 0.0001), ASDAS-ESR (P = 0.04), CRP (P = 0.006), IL-6 (P = 0.02), and IL-18 (P = 0.03) levels. Patients with inflammatory bowel disease (IBD) had higher VAS (P = 0.03), CRP (P = 0.0009), and IL-6 (P = 0.0003) levels. Patients with skin psoriasis had lower VAS (P = 0.001) and BASDAI (P = 0.00007) levels. Patients with psoriatic onycholysis had lower VAS (P = 0.006), BASDAI (P = 0.00001), and CRP (P = 0.02) and higher IL-23 (P = 0.04) levels. Patients with PPP had lower BASDAI (P = 0.04) and higher ET-1 (P = 0.001) levels. Conclusions. SpA patients with increased serum IL-18 and decreased serum ET-1 had an increased risk of extra-articular symptoms. In SpA patients, increased disease activity was associated with an increased risk of AAU and IBD and a decreased risk of skin psoriasis, psoriatic onycholysis, and PPP. Ankylosing spondylitis (AS), psoriatic arthritis (PsA), and SAPHO syndrome (SAPHO) are seronegative spondyloarthropathies (SpA) which are connected with the presence of HLA-B27 antigen \u20133. In thSeveral proinflammatory cytokines such as interleukin-6 (IL-6), interleukin-18 (IL-18), interleukin-23 (IL-23) are considered to play a role in pathogenesis of SpA \u201314. AlsoThere are data that development of different extra-articular symptoms is connected with elevated levels of different markers of inflammatory process , 6, 13. The aim of our study was to assess extra-articular symptoms in constellation with selected serum cytokines and disease activity in SpA. We selected cytokines that play a role in the pathogenesis of SpA and are involved in angiogenesis and endothelial function.This study was approved by the Ethics Committee of the Pomeranian Medical University in Szczecin . Informed consent was obtained from all patients. We studied 287 SpA patients: 131 had AS, 110 had PsA, and 46 had SAPHO. All patients were Caucasian. The following data were recorded: age, sex, disease duration, and extra-articular symptoms: acute anterior uveitis (AAU), inflammatory bowel disease (IBD), skin psoriasis, psoriatic onychopathy, and positivity for HLA B27. The diagnosis of AS was made according to modified New York criteria . The patWe also assessed the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI). This index has a possible score of 0\u201310, with a higher score indicating greater disease activity. We regarded patients as active if the BASDAI score was > 4 .The Ankylosing Spondylitis Disease Activity Score (ASDAS) was assessed using ESR. The ASDAS-ESR was calculated, in AS patients and PsA patients with axial joint involvement, using online calculator available at the Assessment of Spondyloarthritis International Society website.Disease activity score calculation, in PsA patients, were made use of free online Disease Activity Score 28 (DAS28) calculator.In the first 191 SpA patients, 81 had AS, 76 had PsA, and 34 had SAPHO; we studied serum levels of selected cytokines: IL-6, IL-23, IL-18, endothelin-1(ET-1), VEGF, and EGF. Additionally, we studied CRP and ESR. The controls were 30 healthy volunteers. The methods of assessment of these cytokines were presented in our previous paper .R values of correlations were determined and corresponding P values <0.05 were considered significant. The groups were compared using Student's t-test, Mann\u2013Whitney U test, and Kruskal-Wallis test. To assess parameters Pearson's chi-squared test (\u03c72), logistic regression analysis, and step-wise analysis were performed. The level of significance was set at P < 0.05. The statistical analysis was performed using STATISTICA version 8.0, StatSoft, Inc., Tulsa, United States.Data distributions were assessed using the Kolmogorov\u2013Smirnov test. Data are described as mean \u00b1 standard deviation and median (Q1 and Q3). The The clinical characteristics of SpA patients and the prevalence of extra-articular symptoms are presented in P < 0.000001) and in AS than in SAPHO (P = 0.0001). We compared SpA patients with AAU to those without AAU. SpA patients with AAU had longer disease duration (P = 0.0002) and higher prevalence of HLA-B27 antigen (P = 0.0001). SpA patients with AAU had higher disease activity as assessed using the VAS (P = 0.0008), BADSDAI (P = 0.0001), ASDAS-ESR (P = 0.04), and CRP (P = 0.006). Additionally, they had lower disease activity as assessed using the DAS28 (P < 0.0001)  .P = 0.01) and BASDAI (P = 0.005) were associated with increased risk of AAU (P = 0.02) and IL-18 (P = 0.03) levels (P = 0.02). SpA patients with AAU compared to healthy controls had higher serum IL-18 levels (P = 0.00009). In SpA patients as compared to healthy controls, increased serum levels of IL-6 (P = 0.02), IL-23 (P = 0.03), and Il-18 (P = 0.0006) were associated with increased risk of AAU (P = 0.0007) were associated with increased risk of AAU (In SpA patients increased CRP (k of AAU . SpA pat) levels . SpA patk of AAU . In SpA k of AAU .P = 0.004). We compared SpA patients with IBD to those without IBD. SpA patients with IBD had higher prevalence of HLA-B27 antigen (P = 0.04) and higher disease activity as assessed using the VAS (P = 0.03), CRP (P = 0.0009), and IL-6 (P = 0.0003). Additionally, they had lower disease activity as assessed using the DAS28 (P < 0.0001) (P = 0.007). When SpA patients were compared to controls, increased serum levels IL-18 (P = 0.03) were associated with an increased risk of IBD (IBD was present in 3.5% SpA patients and in 7.6% AS patients. No one patient with PsA or SAPHO had IBD . The pre 0.0001) . SpA patk of IBD .P = 0.004), lower prevalence of HLA-B27 antigen (P = 0.00004), and lower disease activity as assessed using the VAS (P = 0.001), and BASDAI (P = 0.00007) (P = 0.003) and BASDAI (P = 0.002) were associated with decreased risk of skin psoriasis  . In SpA soriasis .P > 0.05). In SpA patients as compared to healthy controls, increased serum levels of IL-18 (P = 0.0002) and decreased serum levels of ET-1 (P = 0.006) were associated with increased risk of skin psoriasis , lower prevalence of HLA-B27 antigen (P = 0.0004), and lower disease activity as assessed using the VAS (P = 0.006), BASDAI (P = 0.00001), CRP (P = 0.02), and IL-23 (P = 0.04) (We compared SpA patients with psoriatic onychopathy to those without psoriatic onychopathy. SpA patients with psoriatic onychopathy had shorter disease duration ( = 0.04) .P = 0.02).SpA patients with psoriatic onychopathy compared to healthy controls had higher IL-23 level (P = 0.002) had lower risk of psoriatic onychopathy (SpA patients with increased BASDAI (chopathy .P = 0.0002) and decreased serum levels of ET-1 (P = 0.008) were associated with increased risk of psoriatic onychopathy , lower prevalence of HLA-B27 antigen (P = 0.0008), and lower disease activity as assessed using the BASDAI (P = 0.04). They had higher ESR (P = 0.004) and higher serum ET-1 (P = 0.001) (P = 0.4). In SpA patients compared to healthy controls increased serum levels of IL-18 (P = 0.01) were associated with increased risk of PPP (PPP was present in 14.6% SpA patients and in 91.3% SAPHO patients . We comp= 0.001) . There wk of PPP .We presented extra-articular symptoms in constellation with disease activity and selected serum cytokines which are involved in disease activity, angiogenesis, and endothelial function in SpA patients.The percentage of AAU in SpA patients and AS patients in our study was similar to those presented by other authors \u20136, 9.The most important genetic factor associated with AAU is HLA-B27 . We confIn our study AAU was the most frequent in AS patients. Additionally, SpA patients with AAU had higher levels of ASDAS-ESR and lower DAS28. These confirm observations of other authors that AAU is more frequent in axial SpA , 8, 9. TWe also confirmed as other authors that patients with AAU had longer disease duration and higher disease activity , 8.IL-6 plays role in arthritis but its role in SpA pathogenesis is controversial. Kramer et al.  presenteIL-18 is considered as one of proinflammatory cytokine which also activate and deregulate endothelial function. In our previous study we confirmed higher levels of IL-18 in SpA patients and AS patients . In currThe percentage of IBD in SpA patients in our study was similar to those presented by other authors , 5. PatiThe percentage of SpA patients with skin psoriasis in our study was similar to the results presented by others , 9. PeluWe also found that SpA patients with skin psoriasis, compared to SpA group without that symptom, had lower levels of proinflammatory cytokines such as IL-6, IL-23, IL-18, VEGF, and EGF. Nevertheless, we confirmed that increased serum IL-18 levels were associated with increased risk of skin psoriasis in SpA. These foundlings suggest that some markers of disease activity did not influence on prevalence of psoriatic skin changes. On the other hand it can confirm the role of IL-18 in pathogenesis of skin psoriasis by influence on disease activity and endothelial function.IL-23 is produced by keratinocytes and is considered to play a role in SpA pathogenesis. In our previous study by Przepiera-B\u0119dzak et al.  we showePsoriatic onychopathy was observed only in PsA group. Peluso et al. have presented psoriatic onychopathy in 89.2% PsA patients . In SpA The axis IL-17/ILK-23 is considered to play the crucial role in SpA pathogenesis. In our study we presented association between psoriatic onychopathy and serum IL-23. SpA patients with psoriatic onychopathy had higher serum IL-23 than healthy controls. In our previous study by Przepiera-B\u0119dzak et al.  we confiThere were data which confirmed increased serum IL-18 in SAPHO. These suggested role of IL-18 in pathogenesis of SAHO , 13. In ET-1 plays a role in inflammation and vasculopathy. Kuryliszyn-Moskal et al.  presentepatients with AAU had higher serum IL-6 and IL-18;patients with IBD had higher levels of IL-6;patients with nail psoriasis had higher levels of IL-23;patients with PPP had higher levels of ET-1.In SpA patients, increased serum IL-18 and decreased serum ET-1 were associated with an increased risk of extra-articular symptoms. Additionally, increased serum IL-6 and IL-23 increased the risk of AAU. Among SpA patients, increased disease activity was associated with an increased risk of AAU and IBD and a decreased risk of skin psoriasis, psoriatic onycholysis, and PPP.The analysis of serum levels of selected cytokines in SpA patients with and without extra-articular symptoms and healthy controls confirmed the following:"}
+{"text": "AbstractSciaroidea) are a globally species rich group of lower Diptera. In Europe, Fennoscandian peninsula in particular holds a notable diversity, ca. 1000 species, of which 10 % are still unnamed. Fungus gnats are predominantly terrestrial insects, but some species dwell in wetland habitats.Fungus gnats  and Mycetophilidae , are described. Four of the species are known from Fennoscandia only whilst two are supposed to have boreo-alpine disjunct ranges, i.e. having populations in Fennoscandia and the Central European Alps. One of the species probably has a boreal range . Type material of Boletinacurta Sasakawa & Kimura from Japan was found to consist of two species, and a further species close to these taxa is described from Finland. Phroniaelegantula Hackman is redescribed and reported for the first time from Norway. DNA barcodes are provided for the first time for five species.Eight new fungus gnat species, belonging to the families  Sciaroidea are lower Diptera traditionally classified to the infraorder Nematocera, thread-horned flies , black-winged fungus gnats (Sciaridae) and gall midges (Cecidomyiidae) (ed flies . Nematocfamilies . Sciaroimyiidae) . In thismyiidae) .Fungus gnats are a highly diverse group of flies, having over 5000 known species globally , and butDNA barcoding has become a standard procedure in biodiversity surveys and taxonomic studies e.g. . The metOrfelia Costa, Sciophila Meigen, Boletina Staeger and Phronia Winnertz. Orfelia is a keroplatid genus with 36 Holarctic species, of which 25 are known from the Palaearctic region .Images of male hypopygia were taken using an Olympus SZ61 stereomicroscope equipped with a Canon 650D camera and a LM Digital SLR Adapter. Habitus photos of the new Sciaroidea specimens. Legs or 2\u20133 abdominal segments of the specimens were placed in 96% ethanol in a 96-well lysis microplate and dispatched to the Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario where DNA was extracted and sequenced using standard protocols and primers  was sequenced from a total of 10  primers . The frahttp://v4.boldsystems.org/index.php/IDS_OpenIdEngine) in order to search for conspecific taxa and to assess the COI divergence between the new species and the taxa available in the BOLD database. We used \u201ccurrent database\u201d and \u201cAll Barcode Records on BOLD\u201d as options in the queries. The queries were made during October 2016 and at that time the BOLD database held 4,708,558 sequences (with a minimum sequence length of 500 bp), of which 46206 belonged to the family Mycetophilidae and 3665 to the Keroplatidae. Genetic similarities presented here are based on K2P distances and were calculated by the BOLD identification engine.Barcodes of the Finnish specimens  were submitted to the BOLD  identifiSalmelasp. n.urn:lsid:zoobank.org:act:B5A81F63-3F71-4A8F-B23B-ADF5CA4211D3Type status:Holotype. Occurrence: catalogNumber: DIPT-JS-2014-0233; recordedBy: M. M\u00e4kil\u00e4; individualCount: 1; sex: M; lifeStage: adult; Taxon: phylum: Arthropoda; class: Insecta; order: Diptera; Location: country: Finland; stateProvince: Lapponia kemensis pars orientalis; municipality: Savukoski; locality: T\u00f6rm\u00e4oja Conservation Area; decimalLatitude: 67.823; decimalLongitude: 29.439; Identification: identifiedBy: Jukka E. Salmela; Event: eventDate: 2014-08-07; Record Level: institutionCode: ZMUTType status:Paratype. Occurrence: catalogNumber: BIOUG08366-D12; recordNumber: bayw.17; recordedBy: G. Sellmayer; individualCount: 1; sex: female; Location: country: Germany; stateProvince: Bavaria; locality: Nationalpark Bayerischer Wald, 11.3 km N of Grafenau; decimalLatitude: 48.9509; decimalLongitude: 13.422; Event: eventDate: 2012-09-13/22; Record Level: institutionCode: ZSMMale. Head bicolored, vertex with a triangular dark area, laterally yellowish brown Fig. a, b, c. 1Scutum yellowish with three longitudinal brown stripes; median stripe consisting of two stripes that are largely merged, a narrow anterior gap between the stripes is present Fig. b, c. DarWings yellowish, with a faint subapical dark band extending from C to M2. Veins dark brown except bm\u2013cu and bRs that are lighter. Veins R1 and bCuA with dorsal setae, R5 setose both ventrally and dorsally. Sc ending in C before bRs. R4 very short, about 0.12 times longer than apical portion of R5. Wing length 4.1 mm.Coxae yellowish brown - brown, bearing short dark setae, legs yellowish. Ratio of femur to tibia for fore, mid and hind legs: 0.79, 0.68, 0.63. Ratio of tibia to basitarsus for fore, mid and hind legs: 1.67, 1.0, 1.0. Anterior spur of mid-tarsus about 0.5 times longer than posterior spur.Abdominal tergites and sternites brown, bearing dark setae. Hypopygium brown. 9th tergite widest medially, apex rounded. Gonocoxites dorsally with an outgrowth, bearing a few long apical setae and having a mesial protrusion Fig. a. Cerci 22222Female.The paratype female is lacking all legs except right fore leg and right hind femur. The specimen is slightly paler than the holotype male. The specimen may be somewhat teneral or it has bleached in the Malaise trap or later in the ethanol. Otherwise the specimen is very similar to the holotype. Antennal flagellomeres, except first and last, are wider than long (length:width ratio of 4th segment is 0.78). Cerci short, apically truncated, gonocoxite 8 short and rounded. Wing length 3.9 mm.O.nigricornis (Fabricius) and O.subnigricornis Zaitzev & Menzel have a bunch of setae.The new species is characterised by the short and dark antennae, a yellow scutum with contrasting scutellar stripes, brown pleural sclerites of the thorax, brown, unicolorous abdomen and a short R4 vein. The dorsal lobe of gonostylus is strongly curved. The ventral lobe of gonostylus has only two black apical setae, while The name of the new species refers to its putative boreo-alpine, disjunct range in Europe. The name is a noun in apposition.O.boreoalpina sp.n. has a disjunct European range, having populations in the northern Fennoscandia and the Central European mountains  dominated boreal forest. Bavarian site is a conifer-dominated mountain forest . If using the key provided by O.boreoalpina sp.n. has an orange scutum and brown pleura, thus dropping out already in the first couplet. In the key provided by O.nigricornis, that has elongated palpal segments and one pointed outgrowth on the dorsal side of gonocoxites. Orfelianigricornis, however, has a tuft of setae on the apex of dorsal lobe of the gonostylus while O.boreoalpina has only two dark setae. Other more or less similar species are 1) O.subnigricornis, that is characterized by the yellow scape and pedicel and median flagellomeres that are 1.4 times longer than wide , 2) O.sachalinensis (Matsumura) has a yellow abdomen and indistinct scutal stripes   and 3) O.minima (Giglio-Tos) that has a yellowish scape, pedicel and a yellowish abdomen .The new species is rather distant to all other Holarctic species, but it may be closest to han wide   shared by no other members. The nearest specimens are rather distant: 97 closest sequences have similarity values between 88.25 and 86.35, being assigned to O.nemoralis (Meigen) (54 specimens), O.nigricornis (2), Keroplatidae (40) and Mycetophilidae (1). DNA barcode and associated data of the paratype is available from the BOLD Public data portal.The DNA barcode of the paratype specimen is almost identical to the holotype, their similarity is 99.54 %. The type specimens belong to the same BIN old-growth boreal forest, dominated by birch , black. Scape:pedicel length ratio 1.30; scape with a rounded, a bit depressed sensory field in its lateral base, having 7 minute setae. Flagellomeres cylindrical, length:width ratio of 1st flagellomere 1.51, 4th flagellomere 1.76 and apical flagellomere 3.13. Flagellomeres covered by dense light setosity, setae slightly curved, their length shorter than width of respective flagellomere; polygon-like (reticulate) pattern present, especially so in apical flagellomeres.Thorax black. Scutum covered by pale setae. Anepimeron bare, other sclerites setose. Scutellum with eight setae in a curved row. Halteres light brown with pale setae; apical part of stem and base of knob infuscated.Wings hyaline, lamina covered by both macro and microtrichia. Base of Rs, R4 and r-m bare, other veins setose, veins light brown to dark brown. C exceeding tip of R5 25 % of the distance between R5 and M1. Sc2 situated between base of Rs and R4. Furcation point of median fork at the level of bRs. M1+M2 very short. Length ratio of M1+2:r-m = 0.53. Wing length 3.2 mm.Fore coxae light brown, mid and hind coxae dark brown, with pale setae, trochanters dark-brown. Legs yellowish brown, femora basoventrally darkened; apices of mid and hind coxae infuscated, the latter more clearly so. Setae on femora mostly dark, tibial and tarsal setae dark. Length ratio of femur to tibia for fore, mid and hind legs: 0.93, 1.03, 0.89. Length ratio of tibia to basitarsus for fore, mid and hind legs: 1.36, 1.57, 1.89. Anteroapical depressed area of the fore tibia with two rows of pale setae, proximal row curved with ca. 17 setae and distal row almost straight with ca. 20 setae. Ratio of apical width of tibia:length of longest tibial spur for fore, mid and hind legs: 0.52, 0.33, 0.33.Abdominal tergites and sternites dark brown - almost black, covered by dark setae. Distal margin of 9th tergite rounded Fig. a. Gonoco44444This is a very dark species with the head, antennae, thorax, and abdomen black or dark brown. The 9th tergite is apically rounded. The ventral lobe of the gonostylus has a prominent apical outgrowth. The aedeagus is about as long as the parameres with the apex truncated. The parameres are rather thin with their apices contorted.The new species is named after Mr. Tuomas Holopainen, the founder, songwriter and keyboardist of a Finnish metal band, \"Nightwish\". The name is a genitive.The new species is so far known only from eastern Finnish Lapland, the north boreal ecoregion Fig. .The type locality in T\u00f6rm\u00e4oja Conservation Area was a sloping birch forest in a river canyon, close to a spring brook.Sciophila. The number of large setae on the small median appendage of gonostylus is varying, it may be two or three, thus making the use of Zaitzev's , but is otherwise very different, having e.g. a high number of comb-like megasetae on the large median lobe of the gonostylus (63\u201365 vs. 19 in S.holopaineni sp.n.). If three setae, the species runs to the couplet 79 and thereafter to 95, 99, 109 and finally to 114, but the new species does not fit either S.kashmirensis Zaitzev or S.stackelbergi Zaitzev. Holotype male had two and three setae, paratype male had two in both gonostyli.The new species seems to be rather distant from the known Holarctic species of aitzev's  key probSciophila. JS checked a few specimens in his collection (JES), and the character was present in S.buxtoni Freeman, S.curvata sp.n., Leptomorphusforcipatus Landrock, Polyleptaborealis Lundstr\u00f6m and Allocotocerapulchella (Curtis), but it was absent among Anaclileiadziedzickii (Landrock). In Acnemiatrifida Zaitzev there was an ventroapical sensory field at the scape, with hyaline cover. It is possible, that this trait is symplesiomorphic  amongst Sciophilinae and is lost in some genera.We were not able to find any notes in the literature on the presence of a sensory field at the base of scape among Salmelasp. n.urn:lsid:zoobank.org:act:1B97183D-C325-46CA-9E12-AA1E98678CFEType status:Holotype. Occurrence: catalogNumber: DIPT-JS-2015-0252; recordedBy: J. Salmela; individualCount: 1; sex: male; Location: country: Finland; stateProvince: borealis pars borealisOstrobothnia ; verbatimLocality: Kemij\u00e4rvi, Pyh\u00e4-Luosto National Park, Karhunotko; verbatimLatitude: 67.001; verbatimLongitude: 27.133; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2014-6-10/7-11; habitat: old-growth boreal forest with an intermittent brook; Record Level: institutionCode: ZMUTType status:Other material. Occurrence: recordedBy: A. Polevoi; individualCount: 1; sex: male; Location: country: Russia; stateProvince: Karelia; verbatimLocality: Kivach Nature Reserve; verbatimLatitude: 62.272; verbatimLongitude: 33.986; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 1990-8-13/9-11; habitat: Myrtillus pine forest; Record Level: institutionCode: FRIPMale. Head dark, almost black. Ocelli arranged in a shallow triangle, approximately on the median part of vertex; ratio of distance of lateral ocellus from median ocellus: distance of lateral ocelli from eye = 0.59. Vertex covered by dark setae, face covered by small setae and clypeus by longer setae. Eyes pubescent. Palpi pale, covered by pale setae. Length ratio of palpal segments 3\u20135: 3:4=0.81, 4:5=0.52. Penultimate segment 4.3 times as long as wide, last segment 11.0 times as long as wide. Antennae 16-segmented , brown, first flagellomere light brown. Scape:pedicel length ratio 1.38. Scape with a slightly depressed sensory field in its base, having 5-6 minute setae. Flagellomeres cylindrical, length:width ratio of 1st flagellomere 1.54, 4th flagellomere 1.3 and apical flagellomere 2.90. Flagellomeres covered by dense light setosity, setae slightly curved, their length shorter than width of respective flagellomere. Polygonal (reticulate) pattern not present in basal and median flagellomeres, and either unclearly present or absent on the apical flagellomeres; apical flagellomeres of the holotype are slightly wrinkled.Thorax dark brown. Scutum covered by pale setae. Anepimeron bare, other sclerites setose. Scutellum with eight setae in a curved row. Halteres light brown with pale setae.Wings hyaline, both macro and microtrichia present on lamina. Base of Rs and R4 bare, other veins setose, veins brown to dark brown. C exceeding tip of R5 22 % of the distance between R5 and M1. Sc2 situated above R4. Furcation point of median fork slightly before the level of R4. Length ratio of M1+2:r-m = 0.71. Wing length 2.6 mmCoxae yellow, with pale setae, trochanters infuscated. Legs yellow, femora ventrobasally darkened, setae on femora pale, tibial and tarsal setae darker. Length ratio of femur to tibia for fore, mid and hind legs: 0.98, 0.92, 0.83. Length ratio of tibia to basitarsus for fore, mid and hind legs: 1.81, 1.64, 2.20. Anteroapical depressed area of the fore tibia with ca. 16 pale setae in a row. Ratio of apical width of tibia:length of longest tibial spur for fore, mid and hind legs: 0.65, 0.27, 0.26.Abdominal tergites and sternites dark brown, covered by pale setae. 9th tergite triangular, apex pointed Fig. a. Gonoco66666S.californiensis Zaitzev; the 9th tergite of the latter species is medially constricted, in the former the outline of the 9th tergite is triangular.The new species is characterised by the presence of three setae on the small median lobe of the gonostylus, very narrow dorsal lobe of the gonostylus and strongly curved parameres. The new species is closest to curvata Latin, curved, an adjective) refers to the curved parameres of the male hypopygium.The name of the new species  and having 18 comb-like megasetae . Although not mentioned in the description and improperly figured Boletinacurta Zaitzev 1994: 209 Type status:Paratype. Occurrence: recordedBy: M. Sasakawa; individualCount: 1; sex: male; Location: country: Japan; stateProvince: Honsu; verbatimLocality: Otsu, Mt. Hiei; verbatimLatitude: 35.06; verbatimLongitude: 135.83; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: eventDate: 1974-5-3; Record Level: institutionCode: OSAKABoletinacurta is a poorly known East Palaearctic species, hitherto recorded from Japan, Honshu  range between April 27 and May 3. However, Russian specimens reported by , Honshu  and Russ, Honshu . BoletinBoletinacurta was described from Honshu, the main island of Japan .of Japan . The typcription . The secBoletinacurta can be separated from the closely related B.sasakawai sp.n. and B.norokorpii sp.n. based on the following characters: 1) two stout setae present on the ventral lobe of the gonostylus  and 3) the proximal row of stout setae on cerci (comb-like rows) is wider than apical row  and the first flagellomere of B.curta is yellowish ; pedicel:first flagellomere length ratio of B.curta is 0.32 (0.24 in B.sasakawai sp.n. and 0.36 in B.norokorpii sp.n.). For the details of the male hypopygium of B.curta, please see Fig. ow Fig. 7b, 9b. FuBoletina species, such as B.trivittata Staeger, occur in both early and late season . Hence, it might be possible in theory that B.sasakawai sp.n. is just a late summer/autumn morph of B.curta, likewise the butterfly species Araschnialevana (Linnaeus), that has two distinct colour morphs within a season , elongated . Scape yellowish and brownish, pedicel yellow and first flagellomere basally yellowish. Length ratio of pedicel:first flagellomere 0.24. Flagellomeres dark, palpus yellow.Thorax dark-brown with pale setosity. Scutum shining, pleural sclerites with weak microtrichosity. Antepronotum yellow. Halter yellow. Femora yellow, bearing pale setae. Trochanters infuscated. Femora yellow, but mid and hind femora ventrobasally infuscated. Legs gradually darkening toward tarsi. Tibial spurs brownish. Length ratio of femur to tibia for fore, mid and hind legs: 0.77, 0.66, 0.66. Length ratio of tibia to basitarsus for fore, mid and hind legs: 1.06, 1.68, 1.68.Apex of wing slightly infuscated. Bases of M1 and M2, M1+2, r-m, bM1+2, Rs, A1 and Sc bare, other veins setose. C exceeding tip of R5 36 % of the distance between R5 and M1. Sc ending in C at the level of Rs. Length ratio of M1+2:r-m = 1.19. Cu forking slightly beyond M end of r-m. Wing length 5.0 mm.Abdomen dark-brown, tergites 2\u20134 laterodistally yellowish. 9th tergite elongated; cerci bearing two rows of combs, that are about equally wide, having ca. 45 stout setae Fig. b. Ventra8888B.curta it is curved and bearing two stout setae. The apices of parameres with a conspicuous pair of horn-like outgrowths.A large species with a vaguely infuscated wing apex, abdominal tergites 2\u20134 laterally yellowish and relatively long first flagellar segment (about 4-times the length of the pedicel). The ventral lobe of gonostylus bare, sinuous; in the closely related The new species is named after Dr. Mitsuhiro Sasakawa, Japanese entomologist and the collector of the holotype. The name is a genitive.Known only from the type locality (Yoshino in Japan). The holotype male was collected at the end of October.Boletinacurta.See above Salmela & Kolcs\u00e1rsp. n.urn:lsid:zoobank.org:act:D906B0FD-0C14-4FDC-9FF7-EADF472C4476Type status:Holotype. Occurrence: catalogNumber: DIPT-JS-2016-0044; recordedBy: J. Salmela; individualCount: 1; sex: male; Location: country: Finland; stateProvince: borealis pars borealisOstrobothnia ; verbatimLocality: Ylitornio, Tuorerommas Mire Conservation Area; verbatimLatitude: 66.479; verbatimLongitude: 24.757; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2012-7-2/8-6; habitat: old-growth boreal forest with a spring brook; Record Level: institutionCode: ZMUTMale. Head black, vertex covered by pale setae, frons glabrous and face with scattered setae. Ocelli in a shallow triangle, median ocellus smallest. Clypeus not much longer than wide . Scape and pedicel brownish, first and second flagellomeres yellowish, base of third flagellomere yellowish. Length ratio of pedicel:first flagellomere 0.36. Flagellomeres dark, palpus yellow.Thorax dark-brown with pale setosity. Antepronotum yellow. Halter yellow. Femora yellow, bearing pale setae. Trochanters infuscated. Femora yellow. Legs gradually darkening toward tarsi. Tibial spurs brownish. Length ratio of femur to tibia for fore and hind legs: 0.93, 0.76. Length ratio of tibia to basitarsus of hind leg: 1.68.Apex of wing slightly infuscated. Bases of M1 and M2, M1+2, r-m, bM1+2, Rs, A1 and Sc bare, other veins setose. C exceeding tip of R5 16 % of the distance between R5 and M1. Sc ending in C at the level of Rs. Sc2 present. Length ratio of M1+2:r-m = 1.14. Cu forking slightly beyond M end of r-m. Wing length 4.1 mm.Abdomen dark-brown, tergites 2-4 laterodistally yellowish. 9th tergite elongated; cerci bearing two rows of combs, that are about equally wide, having 18 stout setae . The caudal and proximal combs of the cerci are equally wide, having relatively a small number (18) of stout setae (over 40 in both B.curta and B.sasakawai sp.n.)A species very close to The new species is named after Dr. Yrj\u00f6 Norokorpi, Finnish forest researcher and former area manager at Parks & Wildlife Finland. The name is a genitive.So far known from SW Finnish Lapland only Fig. .Parisquadrifolia and Calypso bulbosa.The Finnish trapping site was an old-growth boreal forest characterised by vascular plants typical for base-rich soils, such as B.curta and B.sasakawai sp.n. It is likely, however, that the eastern species are more related to each other than to B.norokorpii sp.n. For example, presence of Sc2 (absent in other species), shorter basal flagellar segments  and the small number  of stout setae of combs in the cerci separate the new species from the eastern Palaearctic taxa. We assume that both eastern Palaearctic species have restricted ranges in Japan, Far East Russia and neighbouring areas, whereas B.norokorpii sp.n. might have a widespread boreal range.The new species is very close to the eastern Palaearctic species SCFI744-16. GenBank accession number: KY062991.Holotype: BOLD Sample ID: DIPT-JS-2016-0044. BOLD Process ID: AATATTATATTTTATTTTTGGAGCTTGATCAGGAATAATTGGTACATCATTAAGAATTCTTATTCGTGCTGAATTAGGACACCCTGGAGCATTAATTGGAGATGATCAAATTTATAATGTTATTGTAACAGCTCATGCATTTGTAATAATTTTTTTTATAGTAATACCTATTATAATTGGAGGATTTGGTAATTGATTAATCCCTTTAATATTAGGAGCTCCTGATATAGCATTCCCTCGAATAAATAATATAAGATTTTGACTACTTCCTCCTTCATTAATATTACTTTTATCCAGAAGTTTAGTTGAAACAGGGGCTGGTACAGGTTGAACAGTGTACCCACCATTATCCTCAACAATTGCTCATGCAGGAGCATCTGTTGATTTAGCAATTTTTTCATTACATTTAGCAGGAATTTCTTCTATTTTAGGAGCTGTAAATTTTATTACTACAATTATTAATATACGAGCTCCTGGAATTACTTTTGAACGAATACCTCTTTTTGTATGATCAGTTTTAATTACAGCTATTTTATTATTATTATCTCTCCCAGTTTTAGCTGGAGCTATTACTATACTTTTAACAGACCGTAATTTAAATACATCATTTTTTGATCCTGCTGGAGGAGGAGATCCTATTTTATATCAACACTTATTCBOLD:ADD1952, shared by no other specimens. In BOLD database the closest matches to this specimen are three Boletinalundbecki Lundstr\u00f6m and four Boletina unassigned to taxonomic species .The new species is assigned to the BIN Salmelasp. n.urn:lsid:zoobank.org:act:B3FBE538-6CFA-451B-AB4F-F62698CC3E27Type status:Holotype. Occurrence: catalogNumber: DIPT-JS-2014-0011; recordedBy: J. Salmela; individualCount: 1; sex: male; Location: country: Finland; stateProvince: Regio kuusamoensis; verbatimLocality: Salla, V\u00e4rri\u00f6 Strict Nature Reserve, Kuntasjoki; verbatimLatitude: 67.749; verbatimLongitude: 29.616; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2013-6-4/29; habitat: old-growth boreal riparian forest with seepages; Record Level: institutionCode: ZMUTType status:Paratype. Occurrence: catalogNumber: DIPT-JS-2015-0101; recordedBy: J. Salmela; individualCount: 1; sex: male; Location: country: Finland; stateProvince: Regio kuusamoensis; verbatimLocality: Salla, V\u00e4rri\u00f6 Strict Nature Reserve, Kuntasjoki; verbatimLatitude: 67.750; verbatimLongitude: 29.620; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2013-6-29/7-29; habitat: riparian forest with lush vegetation and large amount of decaying trees,; Record Level: institutionCode: JESType status:Paratype. Occurrence: catalogNumber: DIPT-JS- 2014-0404; recordedBy: J. Salmela; individualCount: 1; sex: male; Location: country: Finland; stateProvince: Lapponia kemensis pars orientalis; verbatimLocality: Savukoski, Urho Kekkonen National Park, Tyyroja; verbatimLatitude: 68.143; verbatimLongitude: 28.574; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2014-7-1/8-5; habitat: riparian meadow, spring brook with abundant Palustriella mosses; Record Level: institutionCode: JESMale. Head brown, vertex covered by pale setae, frons glabrous. Ocelli in a line, central ocellus smallest, lateral ocelli close to eyes, their distance from eye less than their own width. Eyes pubescent. Palpi brown, bearing light setae. Length ratio of palpal segments 3-5: 3:4=0.88, 4:5=0.61. Penultimate segment 2.62 times as long as wide, last segment 4.67 times as long as wide. Third palpomere with a sensory pit in its base. Antennae brown, 16-segmented , pedicel and base of first flagellomere yellowish brown. Scape:pedicel length ratio 1.47. Flagellomeres cylindrical, length:width ratio of 1st flagellomere 2.27, 4th flagellomere 1.67 and apical flagellomere 1.90. Flagellomeres covered by dense light setosity, setae slightly curved, their length shorter than width of respective flagellomere.Thorax generally brown, except scutum that has yellowish brown anterior corners. Scutum with mainly pale setosity, two stout and long posterodorsal setae are present just above scutellum. Mediotergite bare, other sclerites bearing setae. Scutellum with four stout marginal setae. Halteres pale, bearing weak light setae and setulae.Wings hyaline, veins light brown. Bases of M1 and M2, M1+2, base of r-m, bM1+2, bRs and Sc bare, other veins setose. C slightly exceeding tip of R5. Sc ending free. Length ratio of M1+2:r-m = 1.03. Wing length 1.8 mm.Coxae yellow, bearing dark setae, legs yellowish. Length ratio of femur to tibia for fore, mid and hind legs: 1.02, 1.0, 0.84. Length ratio of tibia to basitarsus for fore, mid and hind legs: 1.28, 1.6, 1.63. Anteroapical depressed area of the fore tibia ovate, having ca. 20 light setae arranged in a curved row. Ratio of apical width of tibia:length of longest tibial spur for fore, mid and hind legs: 0.37, 0.30, 0.29.Abdominal tergites and sternites brown, bearing light setae. 9th tergite and cerci without peculiar characteristics. Ventroapical margin of gonocoxite with a marked median emargination Fig. b. Ventra10101010A small species that is different from the known member of the genus. The male hypopygium has the following diagnostic characters: the ventroapical margin of the gonocoxites has a deep notch; the mesial projection of the gonostylus lacks comb-like structures but bears a prominent finger-like projection and a rounded, hyaline protrusion; the aedeagal complex is about as long as broad and is apically notched.Lapponia kemensis pars orientalis, abbreviated as Lkor, is in Finnish \"Sompion Lappi\". The name is a noun in apposition.The name of the new species refers to the old Forest Sami name of the region, Sompio, meaning large area bordered by aapamires. The biogeographical province of The species has been collected so far from three localities, all of these from eastern Finnish Lapland close to the Russian border. In fact, all of the collecting sites belong to the River Tuuloma catchment area east of the Maanselk\u00e4 divide, so the waters finally flow to the Barents Sea in Russia.Collecting sites are small waterbodies  surrounded by old-growth boreal forests.The new species stands apart from all other Holarctic members of the genus.SCFI001-15.BOLD Sample ID: DIPT-JS-2014-0011. BOLD Process ID: SCFI164-15.BOLD Sample ID: DIPT-JS-2015-0101. BOLD Process ID: SCFI102-15.BOLD Sample ID: DIPT-JS-2014-0404. BOLD Process ID: Barcoding of the holotype and paratypes failed.Salmelasp. n.urn:lsid:zoobank.org:act:082AAF64-6B68-438B-991A-9C42736838A3Type status:Holotype. Occurrence: catalogNumber: DIPT-JS-2015-0272; recordedBy: J. Salmela; individualCount: 1; sex: male; Location: country: Finland; stateProvince: Regio kuusamoensis; verbatimLocality: Salla, Iso Pyh\u00e4tunturi; verbatimLatitude: 66.776; verbatimLongitude: 28.810; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2013-7-19/8-8; habitat: poor - intermediate rich sloping fen; Record Level: institutionCode: ZMUTType status:Paratype. Occurrence: catalogNumber: 1386 (3); recordedBy: G.P. Ostroverkhova; individualCount: 1; sex: male; preparations: slide mounted; Location: country: Russia; stateProvince: Krasnoyarsk region; verbatimLocality: Tungussko-Chunsky District, village Vanavary; verbatimLatitude: 60.33; verbatimLongitude: 102.30; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: sweep net; eventDate: 1972-7-26; habitat: swampy forest; Record Level: institutionCode: TSUMale. Head dark-brown, vertex covered by pale setae, frons glabrous. Ocelli in a line, central ocellus slightly smaller than laterals; lateral ocelli close to eyes, their distance from eye less than their own width. Eyes pubescent. Palpi brown, bearing light setae. Length ratio of palpal segments 3\u20135: 3:4=0.83, 4:5=0.69. Penultimate segment 3.6 times as long as wide, last segment 5.3 times as long as wide. Third palpomere with a sensory pit in its base. Antennae brown, 16-segmented , base of pedicel and base of first flagellomere yellowish brown. Scape:pedicel length ratio 1.60. Flagellomeres cylindrical, length:width ratio of 1st flagellomere 2.86, 4th flagellomere 1.75 and apical flagellomere 3.0. Flagellomeres covered by dense light setosity, setae slightly curved, their length shorter than width of respective flagellomere.Thorax generally dark-brown, except scutum that has yellowish anterior corners. Scutum with mainly pale setosity. Mediotergite bare, other sclerites bearing setae. Scutellum with four stout setae. Halteres pale, bearing weak light setae and setulae.Wings hyaline, veins brown. Bases of M1 and M2, M1+2, base of r-m, bM1+2, base of Rs and Sc bare, other veins setose. C exceeds tip of R5 very slightly. Sc ending free. Length ratio of M1+2:r-m = 1.18. Wing length 3.1 mm.Coxae yellow, bearing pale setae, legs yellowish, except femora ventrobasally and apices of hind femora infuscated. Length ratio of femur to tibia for fore, mid and hind legs: 0.95, 0.99, 0.82. Length ratio of tibia to basitarsus for fore, mid and hind legs: 1.03, 1.34, 1.60. Anteroapical depressed area of the fore tibia ovate, having ca. 19 light setae arranged in a slightly curved row. Ratio of apical width of tibia:length of longest tibial spur for fore, mid and hind legs: 0.39, 0.27, 0.24.Abdominal tergites and sternites brown, bearing light setae. 9th tergite and cerci normal for the genus Fig. a. Ventro11111111111111P.braueri Dziedzicki but differs in the following features; the apices of the parameres are non-setose (the setae here are long in P.braueri), the internal outgrowth of the ventral lobe of the gonostylus is curved and apically notched (not curved or notched in P.braueri), and the ventral lobe of the gonostylus also has a narrow club-like basal projection .The new species is close of reducta, reduced, an adjective) is referring to the non-setose apices of the male parameres.The name of the new species  was close to a pine and spruce dominated pristine boreal forest.P.annulata, (= P.braueri). These two taxa are indeed closely related, but due to differences in the male hypopygia and DNA barcodes are considered as distinct species .The holotype male is the only member of the BIN Salmelasp. n.urn:lsid:zoobank.org:act:3A162039-F1B7-458E-BEBC-0317C31C25B7Type status:Holotype. Occurrence: catalogNumber: DIPT-JS-2015-0215; recordedBy: E. Rundgren; individualCount: 1; sex: male; Location: country: Finland; stateProvince: Lapponia inarensis; verbatimLocality: Inari, Muotkatunturi Wilderness Area, Kielajoki; verbatimLatitude: 69.146; verbatimLongitude: 26.292; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2014-6-26/8-5; habitat: lush and swampy riparian birch forest; Record Level: institutionCode: ZMUTType status:Paratype. Occurrence: catalogNumber: MYCFI183-11; recordedBy: Finnmarksprosjektet; individualCount: 1; sex: male; Location: country: Norway; stateProvince: Finnmark; verbatimLocality: Alta, Goppaelva; verbatimLatitude: 70.027; verbatimLongitude: 23.394; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela, G. S\u00f6li; Event: samplingProtocol: sweep net; eventDate: 2010-6-13; Record Level: institutionCode: NHMOType status:Paratype. Occurrence: catalogNumber: MYCFI184-11; recordedBy: Finnmarksprosjektet; individualCount: 1; sex: male; Location: country: Norway; stateProvince: Finnmark; verbatimLocality: Alta, Goppaelva; verbatimLatitude: 70.027; verbatimLongitude: 23.394; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela, G. S\u00f6li; Event: samplingProtocol: sweep net; eventDate: 2010-6-13; Record Level: institutionCode: NHMOType status:Paratype. Occurrence: catalogNumber: BC-ZSM-DIP-22552-E10; recordedBy: D. Doczkal, S. Schmidt & J. Voith; individualCount: 1; sex: male; Location: country: Germany; stateProvince: Bavaria; verbatimLocality: Allg\u00e4u, Oberstdorf, Schochen; verbatimElevation: 2032 m; verbatimLatitude: 47.3936; verbatimLongitude: 10.3692; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: J. Salmela; Event: samplingProtocol: Malaise trap; eventDate: 2014-6-6/21; habitat: Blaugras-Horstseggenrasen; Record Level: institutionCode: ZSMType status:Other material. Occurrence: catalogNumber: BIOUG21868-H06; recordedBy: BIObus; individualCount: 1; sex: female; Location: country: Canada; stateProvince: British Columbia; verbatimLocality: Vancouver Island; verbatimLatitude: 49.044; verbatimLongitude: -125.684; verbatimCoordinateSystem: decimal degrees; verbatimSRS: WGS84; Identification: identifiedBy: BOLD ID engine; Event: samplingProtocol: sweep net; eventDate: 2014-6-30; habitat: old growth temperate rain forest; Record Level: institutionCode: BIOUGMale. Head brown, vertex covered by pale setae, frons glabrous. Ocelli arranged in a very shallow triangle, central ocellus slightly smaller than laterals; lateral ocelli close to eyes, their distance from eye less than their own width. Eyes pubescent. Palpi brown, bearing light setae. Length ratio of palpal segments 3\u20135: 3:4=0.92, 4:5=0.62. Penultimate segment 3.51 times as long as wide, last segment 10.0 times as long as wide. Third palpomere with a sensory pit in its base. Antennae brown, 16-segmented . Scape:pedicel length ratio 1.28. Flagellomeres cylindrical, length:width ratio of 1st flagellomere 3.0, 4th flagellomere 2.58 and apical flagellomere 3.2. Flagellomeres covered by dense light setosity, setae slightly curved, their length shorter than width of respective flagellomere.Thorax generally brown, scutum dorsally dark-brown. Scutum with mainly pale setosity, including the two stout and long dorso-posterior setae above scutellum. Mediotergite bare, other sclerites bearing setae. Scutellum with four stout marginal setae. Halteres pale, bearing weak light setae and setulae.Wings hyaline, veins light brown. Bases of M1 and M2, M1+2, r-m, bM1+2, bRs and apex of Sc bare, other veins setose. C slightly exceeding tip of R5. Sc ending free. Length ratio of M1+2:r-m = 1.29. Wing length 3.2-3.5 mm.Coxae and legs yellowish brown, bearing dark setae. Length ratio of femur to tibia for fore and mid legs : 0.93, 0.9, 0.76. Length ratio of tibia to basitarsus for fore and mid legs: 0.96, 1.21, 1.5. Anteroapical depressed area of the fore tibia ovate, having numerous light setae over the area. Ratio of apical width of tibia:length of longest tibial spur for fore and mid legs: 0.67, 0.33, 0.23.Abdomen. 9th tergite and cerci as in Fig. 141414141414Female. In general, similar to male. Antennae dark except scape, pedicel and base of 1st flagellomere yellowish brown. Scape:pedicel length ratio 1.54. Length:width ratio of 1st flagellomere 3.1, 4th flagellomere 2.14 . Length ratio of M1+2:r-m = 1.58. Wing length 3.5 mm.Phroniaprolongata sp.n. is a closely related species of P.exigua , the aedeagus  is evenly curved along its length (less curved in P.exigua) and the parameres are very long, about as long as the aedeagus .prolongata, an adjective) of the new species refers to the elongated, prolonged parameres of the male hypopygium.The name , Norway, Finland and Germany Fig. . FennoscThe Finnish collecting site was a swampy riparian birch forest and in Germany the species was collected from an alpine meadow.P.exigua, all sharing a beaked, setose ventral lobe of the gonostylus  and a row of ventral setae on the hind tibia .The new species belongs to a group of species clastered with nd tibia . The newsee e.g. c. The clSCFI251-15. GenBank accession number: KY062993.BOLD Sample ID: DIPT-JS-2015-0215. BOLD Process ID: AATTTTATACTTTATTTTTGGTGCTTGATCTGGAATAGTAGGAACTTCCCTAAGAATTATTATTCGTGCTGAACTTGGTCATCCAGGAGCATTGATTGGAAATGATCAAATTTATAATGTAATTGTTACTGCTCATGCTTTCATTATAATTTTTTTTATAGTTATGCCCATTATAATTGGTGGGTTTGGTAACTGACTTGTCCCATTGATATTGGGGGCCCCTGATATAGCTTTTCCTCGAATAAATAATATAAGTTTCTGATTATTGCCTCCCTCATTAACACTTCTTCTTTCAAGAAGTTTAGTCGAAGCTGGGGCTGGTACAGGTTGAACTGTTTATCCCCCTCTTTCTTCTACTATTGCTCACGCAGGATCTTCTGTTGATCTAGCTATTTTTTCTCTTCATTTAGCTGGTATTTCTTCAATTTTAGGGGCGGTAAATTTTATCACAACTATTATTAATATACGAGCTCCAGGAATTTCCTTTGATCGTTTACCTTTATTTGTTTGATCTGTTCTTATTACTGCTGTATTGCTTCTTTTATCGCTACCAGTTTTAGCTGGGGCTATTACTATACTTTTAACTGATCGAAATTTAAACACATCTTTCTTTGACCCTGCCGGAGGGGGGGACCCTATTCTTTATCAACATTTATTTP.exigua range between 95.57 and 94.8, and between the new species and P.egregia 89.98-87.86. The new species displays a notable intraspecific variation: the Canadian non-type female has 97.06 similarity compared to the holotype and the two Norwegian specimens have 96.6 similarity. Interestingly the similarity of the holotype and German paratype is 98.94, and these two are classified to the same Barcode index number (BIN) by the BOLD (BOLD:ACW2188), shared by no other specimens. The new species is, however, monophyletic  Brachypogonsociabilis (Goetghebuer) and Bezziarhynchostylata Remm in Finnmark, Norway, were characterised by relatively high intraspecific distances (4.0-3.8 %) and were classified to four and three BINs, respectively ; scape, pedicel and basal half of first flagellomere yellowish. Scape with a prominent dorsal seta, about as long as first flagellomere. Scape:pedicel length ratio 1.33. Flagellomeres cylindrical, length:width ratio of 1st flagellomere 2.98, 4th flagellomere 1.75 and apical flagellomere 2.95. Flagellomeres covered by dense light setosity, setae slightly curved, their length shorter than width of respective flagellomere.Thorax generally brown. Scutum dorsally with three dark stripes, that are almost confluent; the stripes are separated by very narrow yellowish gaps; anterolateral corners yellowish. Scutum with mainly pale setosity. Mediotergite bare, other sclerites bearing setae. Scutellum with four stout setae. Halteres pale, bearing weak light setae and setulae.Wings hyaline, veins light brown. Bases of M1 and M2, M1+2, r-m, bM1+2, base of Rs and apex of Sc bare, other veins setose. C very slightly exceeding tip of R5. Sc ending free. Length ratio of M1+2:r-m = 1.29. Wing length 2.2 mm.Coxae and legs yellow, apices of mid and hind femora sligthly infuscated, bearing dark setae. Length ratio of femur to tibia for fore, mid and hind legs: 0.99, 0.97, 0.79. Length ratio of tibia to basitarsus for fore, mid and hind legs: 1.08, 0.97, 0.8. Anteroapical depressed area of the fore tibia ovate, having ca. 20 light in a row. Ratio of apical width of tibia:length of longest tibial spur for fore, mid and hind legs: 0.35, 0.28, 0.25.Abdomen mostly dark brown, but first, second and third tergites caudolaterally yellowish; these yellow areas are most extensive in second and third tergite. Sternum of second and third segments yellowish. Hypopygium dark brown. Ventroapical margin of gonocoxite with a wide and shallow median emargination, with a moderate median peak Fig. b. Ventra17171717Female. Similar to male. Antennae dark except scape, pedicel and base of 1st flagellomere yellowish brown. Scape:pedicel length ratio 1.32. Length:width ratio of 1st flagellomere 3.9, 4th flagellomere 2.80, apical flagellomere 2.5. Length ratio of M1+2:r-m = 1.84. Wing length 2.2 mm.Phronia species with a yellowish pattern on the abdominal tergites 1\u20133. The ventral lobe of the gonostylus is rounded and at its widest basally. The mesial projections are finger-like and the inner lamella of the ventral lobe of the gonostylus bears a tuft of setae. The species is somewhat close to P.elegans Dziedzicki and P.signata Winnertz, that have similarly shaped ventral lobe of the gonostylus; P.elegantula can be distinguished from these due to differences in the structure of the aedeagus, the ventral lobe of gonostylus and the mesial portion of the gonostylus.A A European species. The species was described from eastern Finland (Ok: Sotkamo and Ks: Kuusamo) and has been later recorded from southern and northern parts of the country . The species has been found from Russian Karelia  and MurmSampling sites are coniferous forests, mixed forests and wetlands.Phroniaelegantula is somewhat similar to P.signata and P.elegans, and has the same yellowish anterolateral corners to the scutum as well as a rotund ventral lobe of the gonostylus. However the abdomen of P.elegans is dark brown as opposed to some yellowish colouration on abdominal tergites 1\u20133 of P.elegantula. Phroniasignata have only moderately emarginated ventroapical margins of the gonocoxites, whereas this character is much more conspicuous in P.elegantula. Phroniasignata has ca. 14 setae on the ventral edge of the ventral lobe of gonostylus . The nearest specimens in BOLD database belong to P.disgrega Dziedzicki, being 90.98 % similar to P.elegantula. DNA barcode and associated data of the German paratypes and female specimens is available from the BOLD Public data portal.All studied specimens belong to the BIN"}
+{"text": "Edwardsiella ictaluri is a Gram-negative bacillus that has recently been implicated in disease outbreaks in tilapia and zebrafish. We report here the complete and annotated genome sequence of an isolate from a Nile tilapia (Oreochromis niloticus), which contains a chromosome of 3,630,639\u00a0bp and two plasmids. Ictalurus punctatus)  was first detected in the late 1970s, predominantly in pond-reared fingerling channel catfish . Raw data 754\u00a0Mb) were error-corrected and trimmed to a total of 129\u00a0Mb in 8,795 sequences  and assembled using Canu version 1.3 .The circularized and completed genome was submitted to the National Center for Biotechnology Information (NCBI) Prokaryotic Genome Annotation Pipeline (PGAP) for annotation and submission to GenBank. The genome was also submitted for Rapid Annotations using Subsystems Technology (RAST) analysis , 11, empE.\u00a0ictaluri RUSVM-1 genome consists of one circular chromosome with 3,630,639\u00a0bp (57.4% G+C content). PGAP annotation predicted 3,377 genes encoding 3,254 proteins and 93 tRNAs. The RUSVM-1 genome is 5% smaller than that of E.\u00a0ictaluri 93-146, isolated from catfish, which contains 3,783 predicted genes. RNAmmer ( E.\u00a0ictaluri isolate 93-146 (E. anguillarum isolate LADL05-105 (E. piscicida isolate S11-285 (E. tarda isolate FL95-01 (E.\u00a0hoshinae isolate ATCC 35051 (E. ictaluri isolated from tilapia will be useful for future investigations into host-pathogen interactions and comparative Edwardsiella analyses.The e 93-146 , 92.4% (DL05-105 , 92.1% ( S11-285 , 83.0% ( FL95-01 , and 82.CC 35051 . Two plaCC 35051 . This coEdwardsiella ictaluri isolate RUSVM-1 has been deposited in GenBank under the accession no. CP020466.The complete genome sequence for"}
+{"text": "D. Adams) A. Rich., Erythrina sigmoidea Hua (Fabaceae), Imperata cylindrical Beauv. var. koenigii Durand et Schinz, Nauclea pobeguinii (Pob\u00e9g. ex Pellegr.) Merr. ex E.M.A., Piper capense L.f., Polyscias fulva (Hiern) Harms., Uapaca togoensis Pax., Vepris soyauxii Engl. and Xylopia aethiopica  A. Rich. Prominent antiproliferative compounds include: isoquinoline alkaloid isotetrandrine (51), two benzophenones: guttiferone E (26) and isoxanthochymol (30), the isoflavonoid 6\u03b1-hydroxyphaseollidin (9), the naphthyl butenone guieranone A (25), two naphthoquinones: 2-acetylfuro-1,4-naphthoquinone (4) and plumbagin (37) and xanthone V1 (46). However, only few research activities in the African continent focus on cytotoxic drug discovery from botanicals. The present review is expected to stimulate further scientific efforts to better valorize the African flora.Cancer remains a major health hurdle worldwide and has moved from the third leading cause of death in the year 1990 to second place after cardiovascular disease since 2013. Chemotherapy is one of the most widely used treatment modes; however, its efficiency is limited due to the resistance of cancer cells to cytotoxic agents. The present overview deals with the potential of the flora of Central, Eastern and Western African (CEWA) regions as resource for anticancer drug discovery. It also reviews the molecular targets of phytochemicals of these plants such as ABC transporters, namely P-glycoprotein (P-gp), multi drug-resistance-related proteins (MRPs), breast cancer resistance protein  as well as the epidermal growth factor receptor (EGFR/ErbB-1/HER1), human tumor suppressor protein p53, caspases, mitochondria, angiogenesis, and components of MAP kinase signaling pathways. Plants with the ability to preferentially kills resistant cancer cells were also reported. Data compiled in the present document were retrieved from scientific websites such as PubMed, Scopus, Sciencedirect, Web-of-Science, and Scholar Google. In summary, plant extracts from CEWA and isolated compounds thereof exert cytotoxic effects by several modes of action including caspases activation, alteration of mitochondrial membrane potential (MMP), induction of reactive oxygen species (ROS) in cancer cells and inhibition of angiogenesis. Ten strongest cytotoxic plants from CEWA recorded following  Cancer is a term for a series of malign diseases characterized by abnormal cell proliferation, leading to invasion and metastasis, the ultimate causes of deaths by cancer. The burden of neoplastic diseases affects the entire world population. Over the past two decades, there has been a slight improvement in cancer statistics due to diagnostic and therapeutic progresses and a better understanding of tumor biology -binding cassette (ABC) proteins are amongst the largest protein families found in all living organisms from microbes to humans . Additionally, other ABC transporters such as ABCC1/MRP1 , a signal transducer for cell growth and differentiation, is the cell-surface receptor belonging to the ErbB family of receptors. This family consists of four closely related receptor tyrosine kinases, namely EGFR/HER1/ErbB-1, HER2/c-neu/ErbB-2, HER3/ErbB-3, and HER4/ErbB-4. Mutations affecting the activity or expression of EGFR can contribute to carcinogenesis . They can be classified into three main types, that are initiator caspase , executioner or effector caspases  and inflammatory caspases   are generated. In many cancer cells, mitochondria appear to be dysfunctional , with a shift in energy metabolism from oxidative phosphorylation to active glycolysis and an increase in the generation of ROS  pathway or Ras-Raf-MEK-ERK pathway is one of the most important signal transduction pathways. The MAPK/ERK pathway regulates growth, proliferation, differentiation and survival of the cells. Its deregulation is observed in various diseases such as cancer, degenerative syndromes, immunological and inflammatory diseases, making it an important drug target  will be discussed. According to the criteria of the American National Cancer Institute, 20 \u03bcg/mL is the upper IC50 limit to be considered as promising for cytotoxic crude extracts   from Garcinia punctata Oliv.  and isobavachalcone (27) isolated from Dorstenia barteri Bureau  isolated from Dorstenia mannii Hook.f.  from Morus mesozygia Stapf.  from Dorstenia angusticornis Engl.  from Erythrina sigmoidea Hua  from Erythrina excelsa Baker  from Milicia excelsa Welw C.C. Berg.  from Vismia laurentii De Wild.  Engl. et Diels.  A. Rich.  Harms.  K.Schumex.Engl.  A. Rich.  M\u00fcll.-Arg.   and damnacanthol (20) from Pentas schimperi (Hook f.) Verde  from Maesa lanceolata Forssk., Myrsine africana L., Embelia keniensis R.E.Fr., Embelia schimperi Vatke and Rapanea pulchra Gilg & Schellenb. and the naphthoquinone, plumbagin (37) from Plumbago and Diospyros species  isolated Xylopia aethiopica  A.Rich.  from Polygonum limbatum Meisn  from Erythrina sigmoidea Hua  from Ficus chlamydocarpa Mildbr. & Burret  from Dorstenia mannii Hook.f.  from Dorstenia poinsettifolia Engl.    and sophorapterocarpan A (45)  from Pycnanthus angolensis (Welw.) Ward  from Milicia excelsa Welw C.C. Berg.  M\u00fcll.-Arg.  Engl. et Diels.  Harms.  Hochst (Anacardiaceae)  and isoquinoline alkaloid isotetrandrine (51), anthraquinones 19 and 20    and the xanthone 46 . Plants and compounds inducing hypersensitivity in these cell lines are summarized in Tables Some botanicals and phytochemicals from CEWA were screened against ABC transporters-expressing cell lines. The most investigated cell lines included the P-gp-overexpressing CEM/ADR5000 leukemia cell line, the MRP1-expressing HL60/AR leukemia cell line and BCRP-expressing MDA-MB-231/Aframomum arundinaceum (Oliver & Hanbury) K. Schum (Zingiberaceae)   Merr. ex E.M.A. (Rubiaceae)     from Polygonum limbatum Meisn  from Dorstenia barteri  from Erythrina sigmoidea Hua  from Aframomum arundinaceum (Oliver & Hanbury) K. Schum  from Uapaca togoensis Pax.    Merr. ex E.M.A.   isolated from Aframomum arundinaceum (Oliver & Hanbury) K. Schum  from Hypericum lanceolatum Lam.  from Aframomum arundinaceum (Oliver & Hanbury) K. Schum  from Erythrina sigmoidea Hua  from Garcinia nobilis Engl.  . They included: Albizia adianthifolia (Schum.) and Alchornea cordifolia (Schum. & Thonn.) M\u00fcll.-Arg.   Stapf. (Euphorbiaceae)   Merr. ex E.M.A.   A.Rich. (Annonaceae)  from Hypericum lanceolatum Lam.   compared to its sensitive counterpart HCT116 (p53+/+) cell line included: Beilschmiedia acuta Kosterm (Lauraceae)  A. Rich. (Compositae)   Merr. ex E.M.A.  Harms.    from Erythrina senegalensis DC  Robyns & Wilczek or Tylostemon acutifolius Engl. & K. Krause] belongs to the family Lauraceae. The plant is mainly found in Cameroon and Central African Republic, where it is traditionally used to treat cancer and gastrointestinal infections , CEM/ADR5000 cells (IC50: 7.96 \u03bcg/mL)  and HL60AR cells (IC50: 9.24 \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50: 8.61 \u03bcg/mL), MDA-MB-231-BCRP cells (IC50: 6.52 \u03bcg/mL), colon carcinoma HCT116 (p53+/+) cells (IC50: 3.58 \u03bcg/mL), HCT116 (p53\u2212/\u2212) cells (IC50: 3.29 \u03bcg/mL), gliobastoma U87MG cells (IC50: 13.55 \u03bcg/mL) and U87MG.\u0394EGFR cells (IC50: 11.15 \u03bcg/mL), hepatocarcinoma HepG2 cells (IC50: 14.32 \u03bcg/mL)   is a tree of 3\u20136 m, or 10\u201320 m belonging to the Fabaceae family. The plant is mainly found in Cameroon and Chad, where it is used as antidote , diuretic, febrifuge and to treat arthritis, rheumatism, pulmonary troubles, stomach troubles, infectious diseases and kidney diseases , atalantoflavone (15), bidwillon A (16), neobavaisoflavone (35), neocyclomorusin (36), and sigmoidin I (44) , CEM/ADR5000 cells (IC50: 20.06 \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50: 22.37 \u03bcg/mL), MDA-MB-231-BCRP cells (IC50: 27.42 \u03bcg/mL), HCT116 (p53+/+) cells (IC50: 19.63 \u03bcg/mL), HCT116 (p53\u2212/\u2212) cells (IC50: 16.22 \u03bcg/mL), U87MG cells (IC50: 45 \u03bcg/mL), U87MG.\u0394EGFR cells (IC50: 29.80 \u03bcg/mL), and HepG2 cells (IC50:22.34 \u03bcg/mL)  and CEM/ADR5000 cells (IC50: 7.18 \u03bcg/mL), pancreatic MiaPaca-2cells (IC50: 12.11 \u03bcg/mL) , HL60AR cells (IC50: 30.60 \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50: 5.19 \u03bcg/mL), MDA-MB-231-BCRP cells IC50: 10.04 \u03bcg/mL), HCT116 (p53+/+) cells (IC50: 4.37 \u03bcg/mL), HCT116 (p53\u2212/\u2212) cells (IC50: 4.60 \u03bcg/mL), U87MG cells (IC50: 19.99 \u03bcg/mL), U87MG.\u0394EGFR cells (IC50: 10.68 \u03bcg/mL), and HepG2 cells (IC50: 18.28 \u03bcg/mL)  is a deciduous, small to medium-sized tree growing up to 30 m tall, sometimes a shrub. In CEWA, the plant is distributed in South Tropical Africa especially in Angola, Zambia, West Tropical Africa: Burkina, Ghana, Guinea, Guinea-Bissau, Ivory Coast, Nigeria, Senegal, Sierra Leone, West-Central Tropical Africa: Cameroon, Central African Republic, Congo, DR Congo, Gabon. The plant is used in traditional medicine as abortive and to treat stomach ache and infectious diseases , CEM/ADR5000 cells , HCT116 (p53+/+) cells  and HCT116 (p53\u2212/\u2212) cells  , CEM/ADR5000 (IC50: 6.56 \u03bcg/mL) and MiaPaca-2 cells (IC50: 8.92 \u03bcg/mL) , HL60AR cells (IC50: \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50:4.17 \u03bcg/mL), MDA-MB-231-BCRP cells (IC50: 19.45 \u03bcg/mL), HCT116 (p53+/+) cells (IC50: 4.64 \u03bcg/mL), HCT116 (p53\u2212/\u2212) cells (IC50: 4.62 \u03bcg/mL), U87MG cells (IC50: 13.48 \u03bcg/mL), U87MG.\u0394EGFR cells (IC50: 7.44 \u03bcg/mL), HepG2 cells (IC50: 16.07 \u03bcg/mL) , 22.63 \u03bcg/mL (CEM/ADR5000 cells), 3.27 \u03bcg/mL (MDA-MB-231 cells), 16.67 \u03bcg/mL (MDA-MB-231/BCRP cells), 14.66 \u03bcg/mL (HCT116 p53+/+cells), 5.98 \u03bcg/mL (HCT116 p53\u2212/\u2212cells), 4.15 \u03bcg/mL (U87MG cells), 16.35 \u03bcg/mL (U87MG.\u0394EGFR cells), and 12.99 \u03bcg/mL (HepG2 cells)  against the above cancer cell lines  Hutch., Uapaca perrotii Beille Uapaca somon Aubr\u00e9v. & Leandri) is an evergreen tree. The plant grows in tropical Africa, from Senegal to southern Chad and Central African Republic and from Gabon to DR Congo and northern Angola. In traditional medicine, the plant is used as antiemetic, lotion for skin disorders , CEM/ADR5000 cells (IC50: 4.44 \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50: 25.85 \u03bcg/mL), MDA-MB-231-BCRP cells (IC50: 4.17 \u03bcg/mL), HCT116 (p53+/+) cells , HCT116 (p53\u2212/\u2212) cells (IC50: 3.09 \u03bcg/mL), U87MG cells (IC50: 8.01 \u03bcg/mL), U87MG.\u0394EGFR cells (IC50: 8.68 \u03bcg/mL), and HepG2 cells (IC50: 19.90 \u03bcg/mL)  and an alkaloid,arborinin (49)  is a plant of the family Rutaceae mostly found throughout West Africa, from Sierra Leone, Liberia, Ivory Cost, Mali, Ghana to Nigeria, and Cameroon. In traditional medicine, the plant is used as anti-fibriomyoma and to treat stomachache, malaria , CEM/ADR5000 cells (IC50: 11.72 \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50: 7.52 \u03bcg/mL), MDA-MB-231-BCRP cells (IC50: 12.93 \u03bcg/mL), HCT116 (p53+/+) cells (IC50: 8.59 \u03bcg/mL), HCT116 (p53\u2212/\u2212) cells (IC50:9.70 \u03bcg/mL), U87MG cells (IC50: 8.75 \u03bcg/mL), U87MG.\u0394EGFR cells (IC50: 4.09 \u03bcg/mL) and HepG2 cells (IC50: 13.60 \u03bcg/mL) , CEM/ADR5000 cells (IC50:7.4 \u03bcg/mL)and Mia PaCa-2 cells (IC50: 6.86 \u03bcg/mL) , breast adonocarcinoma MCF7 cells (IC50: 60.2 \u03bcg/mL), human oral squamous carcinoma KB cells (IC50: 62.5 \u03bcg/mL) , HL60AR cells (IC50: 30.60 \u03bcg/mL), MDA-MB-231-pcDNA cells (IC50: 5.19 \u03bcg/mL), MDA-MB-231-BCRP cells (IC50: 10.04 \u03bcg/mL), HCT116 (p53+/+) cells (IC50: 4.37 \u03bcg/mL), HCT116 (p53\u2212/\u2212) cells (IC50: 4.60 \u03bcg/mL), U87MG cells (IC50: 19.99 \u03bcg/mL), U87MG.\u0394EGFR cells (IC50: 10.68 \u03bcg/mL) and HepG2 cells (IC50: 18.28 \u03bcg/mL)  and trans-tiliroside  isolated from Xylopia aethiopica was amongst the most active alkaloids reported in CEWA plants. This compound displayed interesting cytotoxic effects with IC50 values below 10 \u03bcM toward a panel of sensitive and MDR cancer cell lines. These cell lines included: CCRF-CEM cells (IC50: 1.53 \u03bcM), CEM/ADR5000 cells (IC50: 2.36 \u03bcM), MDA-MB-231-pcDNA cells (IC50: 7.28 \u03bcM), MDA-MB-231-BCRP cells IC50:6.70 \u03bcM), HCT116 (p53+/+) cells (IC50: 2.39 \u03bcM), HCT116 (p53\u2212/\u2212) cells(IC50: 4.55 \u03bcM), U87MG cells (IC50: 3.89 \u03bcM), U87MG.\u0394EGFR cells (IC50: 1.45 \u03bcM) and HepG2 cells (IC50: 3.28 \u03bcM)  and isoxanthochymol (30) isolated from Garcinia punctata Oliv. (Guttiferae)  , CEM/ADR5000 cells, HL60 cells , HL60AR cells , MDA-MB-231-pcDNA cells , MDA-MB-231-BCRP cells , HCT116 (p53+/+) cells , HCT116 (p53\u2212/\u2212) cells , U87MG cells , U87MG.\u0394EGFR cells  and HepG2 cells  , isobavachalcone (27), neocyclomorusin (36), poinsettifolin B (38), 6\u03b1-hydroxyphaseollidin (9), isoneorautenol (29), neobavaisoflavone (35), sigmoidin I (44) and sophorapterocarpan A (45) , CEM/ADR5000 cells (IC50: 5.51 \u03bcM), MDA-MB-231-pcDNA cells (IC50: 5.70 \u03bcM), MDA-MB-231-BCRP cells (IC50: 5.87 \u03bcM), HCT116 (p53+/+) cells (IC50: 5.68 \u03bcM), HCT116 (p53\u2212/\u2212) cells (IC50: 4.60 \u03bcM), U87MG cells (IC50: 4.91 \u03bcM), U87MG.\u0394EGFR cells (IC50: 4.91 \u03bcM) and HepG2 cells (IC50: 6.44 \u03bcM), a major component of the leaves of Guiera senegalensis displayed good cytotoxic effects on a panel of human cancer cell lines. The cytotoxicity of 25 was documented toward CCRF-CEM cells (IC50: 2.31 \u03bcM), CEM/ADR5000 cells (IC50: 3.19 \u03bcM), MiaPaCa-2 cells (IC50: 12.39 \u03bcM), Capan-1 cells (IC50: 29.08 \u03bcM), MCF-7 cells (IC50: 3.42\u03bcM), 786-0 cells (IC50: 11.32 \u03bcM), U87MG cells (IC50: 7.78 \u03bcM), lung carcinoma A549 cells (IC50: 2.28 \u03bcM), skin melanoma Colo-38 cells (IC50: 7.69 \u03bcM), cervical carcinoma HeLa cells (IC50: 1.61 \u03bcM), and Caski cells (IC50: 3.73 \u03bcM)   and plumbagin (37), showed remarkable cytotoxic effects (Table 4 showed good cytotoxicity toward PF-382 cells (IC50: 2.36 \u03bcM), MiaPaCa-2 cells (IC50: 7.48 \u03bcM), MCF-7 cells (IC50: 6.68 \u03bcM), U87MG cells (IC50: 8.02 \u03bcM), Colo-38 cells (IC50: 2.77 \u03bcM), HeLa cells (IC50: 1.65 \u03bcM), and Caski cells (IC50: 0.70 \u03bcM)  , 786-0 cells (IC50: 9.62 \u03bcM), U87MG cells (IC50: 9.64 \u03bcM), A549 cells (IC50:10.13 \u03bcM), Colo-38 cells (IC50: 3.02 \u03bcM), HeLa cells (IC50: 0.58 \u03bcM), and Caski cells (IC50: 0.61 \u03bcM)  Table  is one oThe present review paper aimed at compiling and summarizing relevant data on the potential of medicinal plant and isolated natural products from Central, Eastern and Western Africa to combat cancer with emphasis on their possible cellular targets. This report could not deliver medical results on the therapeutic capacities of the flora of these three African Regions as anticancer drugs. Nonetheless, in phytochemical and pharmacological basic sciences it clearly shows that efforts are being made by African scientists and their international collaborators to achieve this goal in the future. However, few research teams in the continent are already involved in the cytotoxic drug discovery from botanicals and it is expected that this review will stimulate other laboratories to undertake similar research projects to better valorize the African flora.AM and VK wrote the manuscript; VK and TE designed and corrected the work. All authors read and approved the final version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "In this review, a comprehensive overview about the antifouling compounds from marine invertebrates is described. In total, more than 198 antifouling compounds have been obtained from marine invertebrates, specifically, sponges, gorgonian and soft corals. Biofouling includes microfouling (mainly by bacteria and diatoms) and macrofouling  in the marine environment . BiofoulMarine invertebrates have developed prominent chemical defense systems against biofouling in the course of evolution. Lots of AF compounds have been isolated from marine invertebrates. Several books ,7 and reMarine invertebrates, specifically, sponges, gorgonian and soft corals, are rich sources of novel and bioactive secondary metabolites. Studies of the natural chemistry of these interesting groups of marine invertebrates began in the late 1950s. They are recognized to mainly produce novel diterpenoids, sesquiterpenoids, prostanoids, alkaloids, and highly functionalized steroids that are largely unknown from terrestrial sources. Most of these compounds showed AF activity.Terpenoids, especially isocyanoterpenoids, were the typical AF metabolites of marine sponges.l\u20133) dipodazine (162) and other four dipodazine analogs (163\u2013166) with a dipodazine group significantly inhibited the settlement of B. improvisus larvae with EC50 values of 0.034, 5.8, 1.5, 2.4 and 6.7 \u03bcM [167) from the sponge G. barrette inhibited the settlement of B. improvisus larvae with EC50 value of 15 nM [AF indole alkaloids : Alkaloitoxicity . Barettiectively . In 2006d 6.7 \u03bcM , respectof 15 nM .168), isoaaptamine (169), and demethylated aaptamine (170) isolated from the sponge Aaptos aaptos showed AF activity against zebra mussel attachment [Haliclona exigua was rich in bis-1-oxaquinolizidine alkaloid (171), exhibiting significant AF activity against the growths of seven fouling bacterial strains and against the settlement of B.amphitrite larvae [Other AF alkaloids : Aaptamiptamine 1 isolatedlkaloid 1, exhibitBesides the above characteristic terpenoids, alkaloids and steroids, there were many other kinds of AF compounds isolated from marine invertebrates, such as polyacetylenes, butenolides, phenol derivatives, and peptides.172), callypentayne (173), callytriols A-E (174\u2013178) and callyspongins A-B (179\u2013180) from the sponge Callyspongia truncate showed potent metamorphosis-inducing activity towards the ascidian Halocynthia roretzi larvae with ED100 values of 0.13\u20131.3 \u00b5g/mL, and 174\u2013180 also showed AF activity against B. amphitrite larvae with ED50 values of 0.24\u20134.5 \u00b5g/mL [AF polyacetylene derivatives : Callyte181\u2013183) from a soft coral Sinularia sp. showed moderate AF activity against the barnacle B. amphitrite [R)-5-(1-ethoxypropyl)-5-hydroxy-3,4-dimethylfuran-2(5H)-one (184) as a pair of inseparable epimers, along with (S)-5-hydroxy-3,4-dimethyl-5-propylfuran-2(5H)-one (185) and (S)-5-hydroxy-3,4-dimethyl-5-pentylfuran-2(5H)-one (186) were obtained from the gorgonian S. suberosa. Compounds 184\u2013186 exhibited moderate AF activity against the settlement of B. amphitrite larvae [AF butenolides : Sinularphitrite . Butenole larvae . The strDyside. It was believed that this type of compound was biosynthesised by the symbiotic cyanobacteria of the sponge. Five polybrominated diphenyl ethers including 187 from a sponge Callyspongia sp., 188 from Dysidea granulosa, and 189\u2013191 from D. herbacea were investigated against several taxa of prominent fouling organisms including marine bacteria, the diatom A. coffeaeformis, the barnacle B. amphitrite and the mussel Mytilus edulis. All of these compounds exhibited significant antibacterial and AF activity. Compound 187 was the strongest in all the bioassays with non-toxicity. It inhibited the growth of all of the tested bacterial strains with MIC \u00bc 0.02\u20131.52 \u00b5M, and inhibited the larval settlement of A. coffeaeformis, B. amphitrite and M. edulis larvae with EC50 values of 0.24, 0.66 and 1.26 \u00b5M, respectively [AF brominated phenol derivatives : Brominaectively .1c and B1e (192 and 193), avermectin B2a (194) and ivermectin A1a (195) from the gorgonian Anthogorgia caerulea exhibited potent antilarval activity towards B. amphitrite larvae with low-toxicity [O-palmityl-sn-glycero-3-phosphocholine (196) from the sponge Crella incrustans showed strong inhibition against the settlement of B. amphitrite larvae [197) and B (198) from the sponge Geodia barrette showed significant antilarval activity against the settlement of B. improvises larvae at concentrations of 0.6 and 6 \u03bcM, respectively [Other AF compounds : Four avtoxicity . 1-O-pale larvae . Two novectively .50 < 15 \u03bcg/mL, and AF compounds having high LC50/EC50 ratios (>15) are potentially good candidate antifoulants [Totally, over 198 AF compounds have been obtained from marine invertebrates, especially, sponges, gorgonian and soft corals. These compounds covered isocyanoterpenoids, sesquiterpenes, diterpenes, sesterterpenes, triterpenoids, alkaloids , steroids, polyacetylenes, butenolides, peptides, and phenol derivatives, which played important chemical defense roles in the marine invertebrates. In here, the AF activities of 198 compounds towards microfouling and macrofouling were summarized in foulants . From Ta"}
+{"text": "AbstractKSA) was published in 2013 and contained a total of 582 species; an addendum to this list was published in 2015 adding 142 species and bringing the total number recorded from the province to 724 insect species representing 17 orders. The previous two studies excluded Jabal Shada al-A\u2019la Nature Reserve (SANR), so the present study in SANR, as belonging to Al-Baha Province, are complementary to the previous two. The present study presents a preliminary list of Diptera (Insecta) in SANR, with remarks on their zoogeography, and is the first of a series of planned ecological and systematic studies on different insect orders as one of the outputs of a project proposed to study the entire insect fauna of SANR.The first list of insects of Al-Baha Province, Kingdom of Saudi Arabia ; Saropogon sp. [Asilidae]; Spogostylumtripunctatum  [Bombyliidae]; Phycus sp. [Therevidae]; Hemeromyia sp.; Meoneurapalaestinensis Hennig, 1937 [Carnidae]; Desmometopainaurata Lamb, 1914 [Milichiidae]; Stomoxysniger Macquart, 1851 [Muscidae]; and Sarcophagapalestinensis  [Sarcophagidae].A total number of 119 PageBreakZoogeographic affinities of recorded fly species suggest a closer affiliation to the Afrotropical region (46%) than to the Palearctic region (23.5%) or the Oriental region (2.5%). This supports the previous studies\u2019 conclusions and emphasizes the fact that parts of the Arabian Peninsula, including Al-Baha Province, ought to be a part of the Afrotropical Region rather than of the Palaearctic Region or the Eremic Zone. KSA) between the Holy Makkah and Asir provinces. It is the smallest province in KSA , situated at 41\u201342 \u00b0E and 19\u201320 \u00b0N. It is characterized by natural tree cover (SANR) lies between latitudes 19.8149N\u201319.8763N and longitudes 41.2855E\u201341.3501E ], striped hyaena [Hyaenahyaenasultana ], Arabian wolf , sand cat , and reportedly the Arabian leopard , makes this small protected area a unique PageBreaktreasure of biological diversity. Small communities on the mountain grow a distinctive variety of coffee and other crops in terraced fields (d fields .Diptera (Insecta) in SANR, Al-Baha Province, KSA, with remarks on their zoogeography. This is not the final list of Diptera that occur at SANR with the study serving as a basis for further investigations as many additional collected species are still unidentified and further studies are planned to be carried out at SANR. Also, this is the first of a series of planned ecological and systematic studies on different insect orders as one of the outputs of a project proposed to study the entire insect fauna of SANR.The purpose of this paper is to present a preliminary list of SANR, so the present study and other future studies in SANR are complementary to the previous two studies. Studies on the fauna of SANR are of particular interest as this area lies in a part of the Arabian Peninsula which is thought by many authors to touch three of the main zoogeographical regions: the Palaearctic, the Afrotropical, and the Oriental  Vahl, Blepharisciliaris (L.) B.L.Burtt and Hypoestesforskaolii (Vahl) R.Br. (Acanthaceae); Aloeofficinalis Forssk. (Aloeaceae), Aervajavanica (Burm.f.) Juss. ex Schult., Aervalanata (L.) A. L. Juss. ex Schultes and Celosia spp. (Amaranthaceae); Adeniumobesum (Forssk.) Roem. & Schlt. and Carissaedulis (Forssk.) Vahl (Apocynaceae); Commiphoraquadricinta Schweinf. and Cappariscartilaginea Decne. (Burseraceae); Commelinaforskaolii Vahl (Commelinaceae); Conyzastricta Willd., Echinops sp., Psiadiapunctulata (DC.), Pulicariaundulata (DC.), Rhamnusdispermus (L.), Tagetesminuta L. and Vernoniaschimperi DC. (Compositae); Sansevieriaehrenbergii Schweinf. ex Baker (Dracaenaceae); succulent Euphorbia spp. (Euphorbiaceae); Acaciaasak (Forssk.), Acaciaetbaica Schweinf and Indigoferaspinosa Forssk. (Fabaceae); Asparagusafricanus Lam. (Liliaceae); Hibiscusmicranthus L. and Hibscusdeflersii Schweinf. ex Cufod. ; Ficusingens (Miq.) (Moraceae); Commicarpus spp. (Nyctaginaceae); Aristidaadscensionis L., Cenchrusciliaris L., Eragrostistenella (L.) P. Beauv. ex Roemer & Schultes and Pennisetumdivisum (Gmel.) Henr. (Poaceae); Solanumincanum L. (Solanaceae); Grewiatembensis Fresen and Grewiatenax (Forssk.) (Tiliaceae); Cissusrotundifolius (Forssk.) Vahl (Vitaceae); in addition to semi-evergreen PageBreaksclerophyllous woodlands of the Afromontane vegetation .SANR over two successive years, 2014 and 2015 by the authors. Twelve collecting trips were made, six in 2014 in February, April, June, August, October and December, and six in 2015 in January, March, May, July, September and November. Collections were made in 6 different localities representing different altitudinal levels and habitats in SANR .Images of newly recorded species were made using a Leica MZ 125 stereo-binocular microscope  fitted with a digital camera  at Many studies and keys have been consulted in order to identify, classify and estimate the zoogeographical affiliation of collected specimens and such studies are indicated after each taxon in the list, in addition to the following: Unidentified specimens (or photos of specimens)were sent to experts for identification, as indicated after each of these taxa in the list.Nematocera were examined and preserved in alcohol, while other flies were glued to pinned stiff paper points, and all are deposited at the King Saud University Museum of Arthropods, Riyadh, Saudi Arabia (KSMA).Flies of suborder Abbreviations used: Afrotropical AF Bait trap BT Hand-collecting HP King Saud University Museum of Arthropods, Riyadh, Saudi Arabia KSMAPageBreak Light trap LT Malaise trap MT Nearctic NE Oriental OR Palaearctic PA Pitfall trap PT Jabal Shada al-A\u2019la Nature Reserve SANR Sweeping and areal nets SW Vacuuming VCSANR through the present study. Some species have been identified only to genus and listed herein as the genera were not previously recorded from KSA.A total of 119 fly species belonging to 87 genera, 31 tribes, 42 subfamilies, and representing 30 families was recorded from Most of the recorded fly species are characteristic of the Afrotropical region. Table (2) indicates the zoogeographic affinities of recorded species suggesting a closer affiliation to the Afrotropical region (46%) than to the Palearctic region (23.5%) or the Oriental region (2.5%).PageBreakDipteraOrder NematoceraSuborder BibionidaeFamily Dilophustridentatus Walker, 184815 February 2014 (MT1), 5 May 2015 (SW1).Identification: AF.Known distribution: CeratopogonidaeFamily CeratopogoninaeSubfamily CulicoidiniTribe Culicoideskingi 23 August 2014 .Identification: AF.Known distribution: ForcipomyiinaeSubfamily Forcipomyiasahariensis Kieffer, 192323 August 2014 (LT1).Identification: AF. First record from KSA.Known distribution: CulicidaeFamily AnophelinaeSubfamily Anophelesmulticolor Cambouliu, 190223 August 2014 (LT2), 15 February 2014 (LT3).Identification: PA.Known distribution: CulicinaeSubfamily Aedescaspius 15 February 2014 .Identification: PA.Known distribution: Culexpipiens Linnaeus, 175823 August 2014 (PT4).Identification: Known distribution: Cosmopolitan.SciaridaeFamily Chaetosciara sp. Fig. 715 February 2014 (MT1), 23 August 2014 (LT2).Remark: This seems to be the first record of Sciaridae from KSA..Identification: Known distribution: Undetermined.PageBreakPageBreakBrachyceraSuborder AsilomorphaInfraorder AsiloideaSuperfamily AsilidaeFamily AsilinaeSubfamily AsiliniTribe Neolophonotus sp1. Fig. 414-15 February 2014 , 21 April 2014 (LT3), 27 January 2015 , 5 May 2015 (SW1), 27 July 2015 (LT2).Remark: This seems to be the first record of this genus from KSA.Identification: Dr. Jason G.H. Londt, from photos .Known distribution: Undetermined.Neolophonotus sp2. Fig. 615 February 2014 (MT3), 15 November 2015 (MT3).Remark: This seems to be the first record of this genus from KSA.Identification: Dr. Jason G.H. Londt, from photos .Known distribution: Undetermined.ApocleinaeSubfamily Promachussinaiticus Efflatoun, 1934 Fig. 320 April 2014 (LT6), 3 June 2014 , 3-5 June 2014 (SW2), 15 November 2015 (MT6).Identification: PA. First record of the species from the KSA.Known distribution: DasypogoninaeSubfamily DasypogoniniTribe Saropogonlongicornis  Fig. 53 June 2014 (MT3).Identification: PA. First record from KSA.Known distribution: Saropogon sp.15 November 2015 (MT6).Remark: This seems to be the first record of this genus from KSA.Identification: Known distribution: Undetermined.LaphystiinaeSubfamily Trichardisleucocomus 3 June 2014 (MT5), 5 May 2015 (MT5).Identification: Dr Torsten Dikow, from photos .PA.Known distribution: PageBreakBombyliidaeFamily BombyliinaeSubfamily BombyliiniTribe Bombylelladelicata Wiedemann, 18305 June 2014 (SW6), 28 July 2015 (SW3).Identification: Magdi El-Hawagry using AF.Known distribution: Bombyliuspallidipilus Greathead, 196715 February 2014 (MT1), 23 August 2014 (LT2).Identification: Magdi El-Hawagry using AF.Known distribution: Systoechushorridus Greathead, 198021 April 2014 (LT2), 3 May 2015 (LT5), 14 November 2015 (LT6).Identification: Magdi El-Hawagry using PA.Known distribution: AnthracinaeSubfamily AnthraciniTribe Anthraxsticticus Klug, 1832LT).22 April 2015 4 June 2014 (SW6).Identification: Magdi El-Hawagry using OR, PA.Known distribution: Spogostylumisis 29 July 2015 (PT5).Identification: Magdi El-Hawagry using PA.Known distribution: Spogostylumtripunctatum 4-5 June 2014 (SW2), 2 September 2015 (LT6).Identification: Magdi El-Hawagry using PA. First record from KSA.Known distribution: ExoprosopiniTribe Defilippianigrifimbriata 17 October 2014 (MT5).Identification: Magdi El-Hawagry using AF.Known distribution: Exoprosopadisruptatihamae Greathead, 1980 Fig. 93 June 2014 (SW1).Identification: Magdi El-Hawagry using PageBreakPageBreakAF.Known distribution: Heteraloniabisecta Greathead, 198829 July 2015 (PT5).Identification: Magdi El-Hawagry using AF.Known distribution: Pterobateschalybaeus 3 November 2014 (HP6).Identification: Magdi El-Hawagry using PA.Known distribution: VilliniTribe Exhyalanthraxtriangularis Bezzi, 192427 January 2015 (MT5), 5 May 2015 , 15 November 2015 (MT4).Identification: Magdi El-Hawagry using AF.Known distribution: Pachyanthraxcirce 5 May 2015 (MT4).Identification: Magdi El-Hawagry using AF.Known distribution: Villa bivirgata Austen, 19373 June 2014 (SW4), 5 May 2015 (SW4).Identification: Magdi El-Hawagry using PA.Known distribution: Villa paniscoides Bezzi, 19123 June 2014 (SW4), 27-28 July 2015 (SW1), 15 November 2015 (MT4).Identification: Magdi El-Hawagry using AF.Known distribution: XeramoebiniTribe Desmatoneura sp.4 June 2014 (SW4).Identification: Magdi El-Hawagry using Known distribution: Undetermined.Petrorossiaalbula Zaitzev, 19625 June 2014 (SW2), 27 July 2015 (SW1).Identification: Magdi El-Hawagry using PA.Known distribution: Petrorossialetho 5 June 2014 (SW4), 27 July 2015 (SW1).PageBreakIdentification: Magdi El-Hawagry using PA.Known distribution: Petrorossiatropicalis Bezzi, 19213-5 June 2014 , 5 May 2015 (MT3), 27 July 2015 (SW4).Identification: Magdi El-Hawagry using AF.Known distribution: TherevidaeFamily Phycus sp.1 June 2014 (LT5), 24 August 2014 (LT6).Remark: This seems to be the first record of the genus from KSA.Identification: Dr Martin Hauser .AF.Known distribution: EmpidoideaSuperfamily DolichopodidaeFamily DiaphorinaeSubfamily Asyndetusalbifacies Parent, 1929SW).27 July 2015 (Identification: AF.Known distribution: DolichopodinaeSubfamily Dolichopus sp.23 August 2014 (LT4), 10 December 2014 (LT6), 26 January 2015 (PT4), 27 July 2015 (LT6).Identification: Known distribution: Undetermined.Tachytrechusplanitarsis Becker, 190723 August 2014 (LT2).Identification: PA.Known distribution: NemestrinoideaSuperfamily NemestrinidaeFamily Trichopsideacostata Loew, 185810 December 2014 (LT6).Identification: AF.Known distribution: PageBreakTabanoideaSuperfamily TabanidaeFamily Haematopotapluvialis 15 November 2015 (LT6).Identification: PA.Known distribution: MuscomorphaInfraorder AschizaSection PlatypezoideaSuperfamily PhoridaeFamily Megaseliascalaris 23 April 2014 , 5 June 2014 (PT4), 2 March 2015 (PT4), 29 July 2015 (PT5), 23 August 2015 (LT3).Identification: Magdi El-Hawagry.Known distribution: Cosmopolitan.SchizophoraSection AcalyptrataeSubsection CarnidaeFamily Hemeromyia sp. 23 August 2014 (LT1).Remark: This seems to be the first record of the genus from KSA.Identification: Known distribution: Undetermined.Meoneurapalaestinensis Hennig, 193723 August 2014 .Identification: PA.Known distribution: ChloropidaeFamily ChloropinaeSubfamily Pachylophuspellucidus Becker, 191024 August 2014 (MT6).Identification: AF.Known distribution: Thaumatomyianotata 27 January 2015 (LT1).Identification: AF, PA.Known distribution: OscinellinaeSubfamily Anatrichuspygmaeus Lamb, 191827 July 2015 (VC5).Identification: AF.Known distribution: PageBreakAphanotrigonumsubfasciellum Collin, 19494 June 2014(SW4), 24 August 2014 (LT6).Identification: PA.Known distribution: Lasiochaetavulgaris 15 February 2014 (MT1), 8 December 2014 , 5 May 2015 (MT4).Identification: AF.Known distribution: Polyodaspisrobusta 15 February 2014 , 17 October 2014 (LT1), 27 July 2015 (VC2).Identification: AF.Known distribution: Scoliophthalmusmicantipennis Duda, 19355 May 2015 (MT6).Identification: Identification: AF.Known distribution: Scoliophthalmustrapezoides Becker, 19035 May 2015 (MT6).Identification: Identification: AF.Known distribution: SiphonellopsinaeSubfamily Apotropinagregalis 23 August 2014 , 17 October 2014 (LT5), 8 December 2014 (VC4), 2-3 March 2015 , 17 July 2015 , 15 November 2015 (LT6).Identification: Identification: AF.Known distribution: ChyromyidaeFamily ChyromyinaeSubfamily Somatiosomaeremicolum Ebejer, 200815 February 2014 (MT4).Identification: AF.Known distribution: ConopidaeFamily MyopinaeSubfamily ZodioniniTribe Zodioncinereum 5 May 2015 (MT6).Mei & Stuke J-H (2008) has been consulted to identify this species.Identification: PA.Known distribution: PageBreakDiopsidaeFamily Diopsisapicalis Dalman, 18175 May 2015 .Identification: AF.Known distribution: Sphyracephalabeccarii 2 June 2014 (LT6), 3 June 2014 , 3 June 2014 (MT2), 27 January 2015 (LT4), 5 May 2015 , 15 November 2015 (LT6).Identification: AF.Known distribution: DrosophilidaeFamily DrosophilinaeSubfamily DrosophiliniTribe Drosophilamelanogaster Meigen, 183017-18 October 2014 , 8 December 2014 (PT2), 26-27 January 2015 , 2 March 2015 .Identification: Magdi El-Hawagry.Known distribution: Cosmopolitan.Zaprionusindianus Gupta, 19702 March 2014 (PT5), 23 August 2014 (LT2), 18 October 2014 .Identification: OR.Known distribution: EphydridaeFamily DiscomyzinaeSubfamily DiscomyziniTribe Actocetorindicus , 17 October 2014 (LT4).Identification: AF.Known distribution: Actocetormargaritatus Wiedemann, 1830 Fig. 828 February 2014 (PT3), 23 August 2014 , 10 December (2014 (LT6), 5 May 2015 .Identification: AF.Known distribution: PsilopiniTribe Psilopanilotica 15 February 2014 , 4 June 2014 (SW4), 29 July 2015 .Identification: AF, PA.Known distribution: PageBreakHydrelliinaeSubfamily Notiphilaignobilis Loew, 186229 July 2015 (MT6).Identification: AF.Known distribution: LonchaeidaeFamily LonchaeinaeSubfamily LonchaeiniTribe Silbavirescens Macquart, 185115 February 2014 (SW6).Identification: AF.Known distribution: MilichiidaeFamily MadizinaeSubfamily Desmometopainaurata Lamb, 1914 Fig. 1027 January 2015 (LT2), 29 July 2015 (PT4).Identification: AF. First record from KSA.Known distribution: Desmometopavaripalpis Malloch, 19275 May 2015 (PT5), 29 July 2015 (PT6).Identification: Identification: AF.Known distribution: PhyllomyzinaeSubfamily Phyllomyza sp.15 February 2014 (LT2), 27 July 2015 (LT2).Identification: Known distribution: Undetermined.PyrgotidaeFamily Campyloceraferruginea Macquart, 184315 November 2015 (LT6).Identification: Dr Valery Korneyev, from photos .AF.Known distribution: Eupyrgotalatipennis 3 June 2014 (LT2), 14 November 2015 (LT2).Identification: Dr Valery Korneyev, from photos .AF.Known distribution: PageBreakSphaeroceridaeFamily Rachispodafuscipennis 15 February 2014 , 23 August 2014 (PT6), 18 October 2014 , 8-11 December 2014 .Identification: Magdi El-Hawagry, compared with museum specimens.PA.Known distribution: TephritidaeFamily DacinaeSubfamily DaciniTribe Bactrocerazonata 23 August 2014 (LT2), 5 May 2015 (SW1), 27 July 2015 (SW1).Identification: OR.Known distribution: TephritinaeSubfamily TephritiniTribe Acanthiophilushelianthi 23 August 2014 (LT2).Identification: AF, OR, PA.Known distribution: Dioxynasororcula 15 February 2014 (MT4), 3 June 2014 (MT4), 8 December 2014 .Identification: AF.Known distribution: Goniurelliatridens 23 August 2014 (LT2).Identification: PA.Known distribution: Trupaneastellata 3 June 2014 (LT2).Identification: PA.Known distribution: UlidiidaeFamily UlidiinaeSubfamily UlidiiniTribe Physiphoraalceae 15 February 2014 , 21 April 2014 (LT1), 6 June 2014 (LT1), 23 August 2014 (LT1), 17-18 October 2014 , 27 January 2015 , 5 May 2015 (LT1), 27 July 2015 , 15 November 2015 .Identification: Known distribution: Cosmopolitan.PageBreakCalyptrataeSubsection AnthomyiidaeFamily AnthomyiinaeSubfamily AnthomyiiniTribe Anthomyiapluvialis 15 February 2014 (MT1), 27 January 2015 (MT3), 4-5 May 2015 , 15 November 2015 (LT5).Identification: PA.Known distribution: HydrophoriiniTribe Deliaplatura 15 February 2014 , 23 August 2014 (LT2), 17 October 2014 , 27 January 2015 .Identification: Known distribution: Cosmopolitan.CalliphoridaeFamily CalliphorinaeSubfamily Calliphoracroceipalpis Jaennicke, 186715 February 2014 (MT4).Identification: AF.Known distribution: Calliphoravicina 3 June 2014 (SW6).Identification: Known distribution: Cosmopolitan.ChrysomyinaeSubfamily Chrysomyaalbiceps 4 June 2014 (SW1), 2 September 2015 (LT6), 15 November (LT3).Identification: AF.Known distribution: Chrysomyaputoria 3 June 2014 (SW4).Identification: AF.Known distribution: Chrysomyaregalis Robineau-Desvoidy, 183015 February 2014 (MT3), 4 June 2014 (MT6), 10 December 2014 (LT6).Identification: AF.Known distribution: PageBreakLuciliinaeSubfamily Luciliasericata 16 February 2014 (HP6), 21 February 2014 (LT3), 10 December 2014 (LT6).Identification:Known distribution: Cosmopolitan.PolleniinaeSubfamily Polleniahungarica Rognes, 198717 October 2014 (LT6).Identification: PA.Known distribution: Polleniarudis 17 October 2014 (LT5).Identification: PA.Known distribution: MuscidaeFamily AtherigoninaeSubfamily AtherigoniniTribe Atherigonahumeralis Wiedemann, 183015 November 2015 (SW5).Identification: AF.Known distribution: Atherigonalaevigata 15 February 2014 (MT1), 8 December 2014 (VC4).Identification: AF.Known distribution: Atherigonareversura Villeneuve, 193615 February 2014 (MT3), 23 August 2014 , 17 October 2014 , 5 May 2015 (MT2), 15 November 2015 (MT4), 2 September 2015 (LT6).Identification: OR.Known distribution: CoenosiinaeSubfamily CoenosiiniTribe Coenosiaattenuata Stein, 190315 February 2014 , 23 April 2014 (PT1), 23 August 2014 (LT2), 17 October 2014 , 18 October 2014 (PT5), 5 May 2015 (MT4), 15 November 2015 (MT4).Identification: Known distribution: Cosmopolitan.Coenosiahumilis Meigen, 1826PageBreak5 May 2015 (MT6).Identification: Known distribution: Cosmopolitan.LimnophoriniTribe Lispenivalis Wiedemann, 183015 February 2014 (LT6).Identification: AF.Known distribution: Lispepectinipes Becker, 190323 August 2014 , 17 October 2014 (LT5), 5 May 2015 , 14-15 November 2015 .Identification: PA.Known distribution: MuscinaeSubfamily MusciniTribe Muscaalbina Wiedemann, 18305 May 2015 (MT6).Identification: AF, OR, PA.Known distribution: Muscaautumnalis De Geer, 177623 August 2014 (LT2), 5 May 2015 (MT2).Identification: Known distribution: Cosmopolitan.Muscacalleva Walker, 184914 November 2015 (LT4).Identification: AF.Known distribution: Muscadomestica Linnaeus, 175815 February 2014 , 3 June 2014 , 23 August 2014 , 5 May 2015 (MT6), 2 September 2015 (LT5), 15 November 2015 (LT6).Identification: Known distribution: Cosmopolitan.Muscalucidula 3 June 2014 (MT6).Identification: AF, PA.Known distribution: Muscasorbens Wiedemann, 18305 May 2015 (MT1), 15 November 2015 (LT5).Identification: AF.Known distribution: PageBreakStomoxyiniTribe Stomoxysniger Macquart, 1851 Fig. 1115 February 2014 (MT4), 17 October 2014 (LT5).Identification: AF. First record from KSA.Known distribution: PhaoniinaeSubfamily DichaetomyiiniTribe Dichaetomyialuteiventris 2 March 2015 (PT5).Identification: AF.Known distribution: PhaoniiniTribe Helinaconiformis 15 February 2014 , 21 April 2014 (LT2), 17 October 2014 , 27 January 2015 , 14-15 November 2015 .Identification: AF.Known distribution: Helinalucida 21 April 2014 (LT5).Identification: AF.Known distribution: RhiniidaeFamily Cosminaviridis Townsend, 191715-16 February 2014 , 17 October 2014 (LT5), 27 January 2015 , 4-5 May 2015 .Identification: AF.Known distribution: Isomyiaterminata 15 February 2014 .Identification: AF.Known distribution: Rhiniaapicalis 15 February 2014 (MT5), 3 June 2014 (SW4), 17 October 2014 , 14-15 November 2015 .Identification: AF.Known distribution: PageBreakSarcophagidaeFamily MiltogramminaeSubfamily Taxigrammaheteroneura 15 February 2014 (MT5), 3 June 2014 (SW4), 27 January 2015 (MT4), 5 May 2015 , 27-29 July 2015 (PT5).Identification: Thomas Pape  and the first author.NE, PA.Known distribution: ParamacronychiinaeSubfamily Wohlfahrtiaerythrocera Villeneuve, 191028 July 2015 (PT6).Identification: Thomas Pape  and the first author.AF.Known distribution: Wohlfahrtianuba Wiedemann, 18303 May 2015 (PT5).Identification: Thomas Pape  and the first author.AF.Known distribution: SarcophaginaeSubfamily Blaesoxiphaalgeriensis 23 August 2014 (LT5).Identification: Thomas Pape  and the first author.PA.Known distribution: Blaesoxipharufipes 3 June 2014 (SW4).Identification: Thomas Pape  and the first author.Known distribution: Cosmopolitan.Sarcophagaadhamae 21 April 2014 (BT6).Identification: AF.Known distribution: Sarcophagaafrica 5 May 2015 (SW4).Identification: Thomas Pape  and the first author.Known distribution: Cosmopolitan.Sarcophagababiyari 3 June 2014 (LT6).Identification: Thomas Pape  and the first author.AF.Known distribution: Sarcophagadux Thompson, 186915 February 2014 (MT1).Identification: Thomas Pape  and the first author.Known distribution: Cosmopolitan.PageBreakSarcophagapalestinensis  Fig. 1221 February 2014 (LT1).Identification: Thomas Pape  and the first author.PA.Known distribution: TachinidaeFamily ExoristinaeSubfamily EryciiniTribe Drinolota 15-16 February 2014 , 17 October 2014 , 14-15 November 2015 .Identification: AF, PA.Known distribution: ExoristiniTribe Exoristalarvarum 3 June 2014 .Identification: NE, PA.Known distribution: GoniiniTribe Goniacapitata 5 May 2015 (MT1).Identification: PA.Known distribution: Sturmiabella 15 February 2014 (MT1), 21 April 2014 (LT1), 3 June 2014 (SW4), 27-30 January 2015 , 27 July 2015 (LT5).Identification: OR, PA.Known distribution: PhasiinaeSubfamily CylindromyiiniTribe Cylindromyiabicolor 7 June 2014 (SW4).Identification: PA.Known distribution: TachininaeSubfamily TachininiTribe Dejeaniabombylans 10 December 2014 (LT6).Identification: AF.Known distribution: PageBreakKSA, including Al-Baha Province, is considered to be the most important part of the country and the Arabian Peninsula in general. Floristically and ecologically, this area is similar to the high altitude mountains of north-eastern and eastern parts of Africa, and like other areas in the south-western part of the Arabian Peninsula, contains montane woodlands and evergreen shrub lands, with strong Afromontane affinities  as shown in the present results which obviously emphasize the fact that Al-Baha Province, as lying in the south-western part of the Arabian Peninsula, should be included in the Afrotropical Region rather than in the Palaearctic Region or the Eremic Zone.The authors would like to extend their sincere appreciation to the Deanship of Scientific Research at King Saud University for its funding this research group NO (RGP-1437-009)."}
+{"text": "Enterobacter sp. strain EA-1 is an electrochemically active bacterium isolated from tropical sediment in Singapore. Here, the annotated draft genome assembly of the bacterium is reported. Whole-genome comparison indicates that Enterobacter sp. EA-1, along with a previously sequenced Enterobacter isolate from East Asia, forms a distinct clade within the Enterobacter genus. Enterobacter belong to the class Gammaproteobacteria and are rod-shaped, Gram-negative, motile, facultative anaerobes. They are commonly found in soil, water, sewage, and the intestines of animals and humans and have been associated with nosocomial infections  broth at room temperature and with shaking at 150\u2009rpm. The Wizard genomic DNA purification kit  was used according to the protocol for Gram-negative bacteria supplied by the manufacturer.The draft genome sequence was determined by shotgun sequencing on a MinION Mk1B sequencer  using R9 chemistry and library protocol SQK-LSK108. Raw sequencing data were assembled using Canu version 1.3 , manuallEnterobacter cloacae ATCC 13047, Enterobacter cloacae SBP-8, and Enterobacter sp. strain FY-07. However, a whole-genome comparison indicated an average nucleotide identity (E. cloacae ATCC 13047 (NCBI accession number CP001918), 78.09% compared to E. cloacae SBP-8 (NCBI accession number CP016906), and 93.38% compared to FY-07 (NCBI accession number CP012487), suggesting that EA-1 forms, together with FY-07, a distinct clade within the Enterobacter genus related to but separate from E. cloacae.The draft genome of EA-1 is composed of 5,145,523 bp, with an average G+C content of 54.56%, and it contains 81 tRNAs, 22 rRNAs organized in 7 operons, and 1 transfer-messenger RNA (tmRNA). A comparison of the 16S rRNA gene sequence revealed >98% identity to the 16S rRNA genes of http://www.ebi.ac.uk/Tools/psa/) using the EMBOSS Needle tool . In FY-07, these genes are arranged in three BC-producing operons: bcsI, bcsII, and bcsIII , AKI40_0197 (CWS02_01215), AKI40_0198 (CWS02_01220), AKI40_0199 (CWS02_01225), AKI40_0200 (CWS02_01230), AKI40_0201 (CWS02_01235), and AKI40_0202 (CWS02_01240). For bcsII, the FY-07 and corresponding EA-1 locus tags (in parentheses) are AKI40_0206 (CWS02_01260), AKI40_0207 (CWS02_01265), AKI40_0208 , AKI40_0209 (CWS02_01285), AKI40_0210 (CWS02_01290), and AKI40_0211 (CWS02_01295). In the case of the EA-1 locus tag corresponding to FY-07\u2019s AKI40_0208, multiple ORFs were stitched together during nucleotide alignment in order to identify the complete homologous gene sequence which was interrupted due to frameshifting errors associated with the sequencing technology used. For bcsIII, the FY-07 and corresponding EA-1 locus tags (in parentheses) are AKI40_0894 (CWS02_05060), AKI40_0893 (CWS02_05055), AKI40_0892 (CWS02_05050), and AKI40_0891 (CWS02_05045). For other BC-related genes (not operon-associated), the FY-07 and corresponding EA-1 locus tags are AKI40_0203 (CWS02_01245), AKI40_0204 (CWS02_01250), and AKI40_0205 (CWS02_01255).In-detail genome alignments between EA-1 and FY-07 using the Artemis Comparison Tool , are alsd bcsIII . In bcsICP025776.The whole-genome shotgun project was deposited in NCBI under the accession number"}
+{"text": "Salmonella enterica subsp. diarizonae serovar 61:k:1,5,(7) strain CRJJGF_00165 , isolated from ground beef in 2007.Here, we report a 4.78-Mb draft genome sequence of the  Salmonella enterica subsp. diarizonae serovar 61:k:1,5,(7) strain CRJJGF_00165  distinguishes it from the phylogenetically distinct monophasic S. enterica subsp. IIIa serovar 61:k:1,5,(7)  is mostly associated with reptiles  strain CRJJGF_00165 was isolated from ground beef using standard microbiology techniques. SMART-PCR was used to serotype this isolate  and tRNAs were predicted with Prodigal . DNA libraries were constructed using the Nextera XT DNA preparation kit, and paired-end sequencing was performed on the Illumina HiSeq2500 platform  using a 500-cycle MiSeq reagent kit. A total of 3,848,000 reads were generated. Reads were g Velvet  into 108Prodigal  and ARAGProdigal , respectProdigal , CRISPRFProdigal , and PHAProdigal , respectRG-ANNOT . The genS. enterica subsp. IIIb serovar 61:k:1,5,(7) strain CRJJGF_00165 has been deposited in GenBank (NCBI) under the accession number JQYQ00000000. The version described here is the first version.The genome sequence of"}
+{"text": "Addendum to: Translational Psychiatry (2017) 7, e1001;doi:10.1038/tp.2016.263; published online 17 January 2017post hoc tests for clarifying the direction of the three MANOVA results thatfound significant associations between functional connectivity changes and long-termclinical outcomes. In each of the three regression tests, greater increases infunctional connectivity were associated with improved long-term clinical outcomes, asascertained by positive beta weights for the following regressors: longitudinal positivepsychotic symptoms =2.79;P=0.015), longitudinal affective symptoms =2.36; P=0.035) and subjective long-termrecovery =2.56;P=0.024). These tests replace the correlation tests originally reportedin the article.After publication, the authors determined that regression tests are the appropriate"}
+{"text": "In sub-Saharan Africa, 20%\u201325% of people starting antiretroviral therapy (ART) have severe immunosuppression; approximately 10% die within 3 months. In the Reduction of EArly mortaLITY (REALITY) randomized trial, a broad enhanced anti-infection prophylaxis bundle reduced mortality vs cotrimoxazole. We investigate the contribution and timing of different causes of mortality/morbidity.Participants started ART with a CD4 count <100 cells/\u00b5L; enhanced prophylaxis comprised cotrimoxazole plus 12 weeks of isoniazid + fluconazole, single-dose albendazole, and 5 days of azithromycin. A blinded committee adjudicated events and causes of death as  tuberculosis, cryptococcosis, severe bacterial infection (SBI), other potentially azithromycin-responsive infections, other events, and unknown.P < .05) but not tuberculosis, SBI, potentially azithromycin-responsive infections, or other causes (P > .3); and reduced nonfatal/fatal tuberculosis and cryptococcosis (P < .05), but not SBI, other potentially azithromycin-responsive infections, or other events (P > .2).Median pre-ART CD4 count was 37 cells/\u00b5L. Among 1805 participants, 225 (12.7%) died by week 48. Fatal/nonfatal events occurred early (median 4 weeks); rates then declined exponentially. One hundred fifty-four deaths had single and 71 had multiple causes, including tuberculosis in 4.5% participants, cryptococcosis in 1.1%, SBI in 1.9%, other potentially azithromycin-responsive infections in 1.3%, other events in 3.6%, and unknown in 5.0%. Enhanced prophylaxis reduced deaths from cryptococcosis and unknown causes (Enhanced prophylaxis reduced mortality from cryptococcosis and unknown causes and nonfatal tuberculosis and cryptococcosis. High early incidence of fatal/nonfatal events highlights the need for starting enhanced-prophylaxis with ART in advanced disease.ISRCTN43622374. In sub-Saharan Africa, 20%\u201325% of human immunodeficiency virus (HIV)\u2013infected individuals starting antiretroviral therapy (ART) have a CD4 count <100 cells/\u00b5L . In the The REALITY trial ISRCTN43622374) recruited 1805 HIV-infected ART-naive adults and children \u22655 years (98% aged \u226513 years) from Zimbabwe, Uganda, Malawi, and Kenya with a CD4 count <100 cells/\u00b5L. In brief, participants were randomized 1:1 to enhanced prophylaxis vs standard prophylaxis (cotrimoxazole) ), other events (n = 63 [3.6%]), SBI (n = 33 [1.9%]), other potentially azithromycin-responsive infections (n = 23 [1.3%]), and cryptococcosis (n = 20 [1.1%]). Sixty-seven (29.8%) deaths were adjudicated as compatible with IRIS. Enhanced prophylaxis reduced all-cause mortality by 24 weeks by 27%  log-rank 8.4% wereP = .06) . There w4 and 48 .P = .007), cryptococcosis , and IRIS-compatible events  (P = .26), potentially azithromycin-responsive infections (P = .34), or other events (P = .81). Although occurring at much higher rates  events of each category occurred in 674 (37.3%) participants. The most common were other events (n = 479 participants [27.0%]) , followe = .001) , but noter rates , similarer rates . Rates fAt 48 weeks post\u2013ART initiation, rates of fatal/nonfatal events were lowest  and participants with lower baseline CD4 count (P = .02) or self-reported vomiting (P = .005) (P = .03); those with lower baseline CD4 count (P = .05), creatinine clearance (P < .001), or albumin (P = .01); higher bilirubin (P = .05); without previous healthcare contact (P = .04); or with wasting/severe weight loss (P = .02), participant-reported fever (P = .08), or problems with mobility (P = .05) or self-care (P = .05). There was no effect of water source or household toilet type on either cause of death (P > .3).Given the impact of enhanced prophylaxis on deaths from cryptococcosis and unknown causes, we considered whether predictors of these 2 categories of death were similar, suggesting unascertained cryptococcal disease could be driving these effects. Deaths from cryptococcosis were more common in standard prophylaxis ( = .005) . Deaths Considering the 1716 participants alive and in follow-up at week 4, median last postbaseline VL before death was 95  copies/mL , with reIn the REALITY trial of patients starting ART with severe immunosuppression in sub-Saharan Africa, morbidity/mortality rates were high in the first month after starting ART, regardless of type of event, and dropped exponentially thereafter. However, even within a randomized trial with intensive follow-up, structured clinical narratives, and independent endpoint review, causes of death were difficult to ascertain and the commonest category was unknown. Furthermore, multiple causes were identified in almost one-third of deaths, highlighting the complexity of clinical presentations in patients starting ART with advanced immunosuppression.REALITY showed that enhanced prophylaxis reduces mortality by 27% , due to Of note, 62.7% of participants had no serious morbidity/mortality during their first 48 weeks on ART, despite very low pre-ART CD4 and high VL. Nevertheless, their low CD4 put them at substantially increased risk of impaired short- and long-term immunological response, reinforcing the importance of undertaking baseline CD4 to identify these patients. Where CD4 is becoming less available, our data reinforce the need to further develop CD4 point-of-care tests which at least qualitatively identify the most immunosuppressed individuals. Alternatively, total lymphocyte counts could be used to identify at least some of those at greatest risk , 21.Given current WHO guidance focusing on diagnostic tools to rule out opportunistic infections before ART initiation , our praClinical Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at Supplementary Material 1Click here for additional data file."}
+{"text": "Juniperus phoenicea (Arar), Anastatica hierochuntica (Kaff Maryam), and Citrullus colocynthis  with a set of reference compounds with established modes of action. Cluster analyses of the cytological profiles of the tested compounds suggested that these plants contain possible topoisomerase inhibitors that could be effective in cancer treatment. Using histone H2AX phosphorylation as a marker for DNA damage, we discovered that some of the compounds induced double-strand DNA breaks. Furthermore, chemical analysis of the active fraction isolated from Juniperus phoenicea revealed possible anti-cancer compounds. Our results demonstrate the usefulness of cell-based phenotypic screening of natural products to reveal their biological activities.Natural products have been used for medical applications since ancient times. Commonly, natural products are structurally complex chemical compounds that efficiently interact with their biological targets, making them useful drug candidates in cancer therapy. Here, we used cell-based phenotypic profiling and image-based high-content screening to study the mode of action and potential cellular targets of plants historically used in Saudi Arabia\u2019s traditional medicine. We compared the cytological profiles of fractions taken from  Cancer is a leading cause of morbidity and mortality worldwide, exceeding the number of cases of illness and death due to HIV/AIDS, malaria, and tuberculosis . In 2012High-content screening (HCS) optimizes the discovery of biologically active small molecules and their subsequent development into therapeutic compounds by the use of automated microscopy in conjunction with image analysis. A high-throughput platform based on image-processing software that reviews images from automated fluorescence microscopy \u201310, HCS Anastatica hierochuntica (Kaff Maryam), Juniperus phoenicea (Arar), and Citrullus colocynthis , that have been used in traditional medicine in Saudi Arabia. Previous reports reported anti-cancer activity in Juniperus phoenicea and Citrullus colocynthis .\u201331.1H NM 4 and 5 , 33. TheThis study demonstrates the capability of HCS for cytological profiling for drug discovery in natural products. In recent years, HCS has developed from a promising concept into an efficient methodology and indispensable tool. HCS now can be implemented during the early drug discovery process as a result of its recent technological advances , 34.C. colocynthis, J. phoenicea, and A.hierochuntica. We used a library of known small molecules with assigned modes of action as reference compounds. We compared their cytological profiles with profiles retrieved from the fractionated plant extracts. We were able to predict the mode of action of pure compounds as well as fractionated extracts -\u03b2Dgalactopyranoside triacetate and Estra-1,3,5(10)-triene-3,6beta,17beta-triol triacetate. Estra-1,3,5(10)-triene-3,6beta,17beta-triol triacetate (PubChem CID: 5756) is used as anti-inflammatory drug but no anti-inflammatory activity has been reported for Methyl6-O--\u03b2-Dgalactopyranoside triacetate compound.The 2,2-dimethoxybutane (PubChem CID: 137941) found in the JUN_C2_60% fraction was found to be toxic to microbial membranes \u201361, but polymers . This mae needed . 3-Trifl effects , 67. It  effects , in agre Tunisia . Anti-in Tunisia . Ethyl 4 Tunisia . Of the  Tunisia \u201375. SphiOur results demonstrate the power of HCS for drug discovery. HCS allows traditionally used medicinal plants to be assessed for the presence of bioactive compounds. Cytological profiles allow us to identify topoisomerase II inhibitor activity in the plant fractions that we tested to verify the presence of anti-cancer compounds in the plants. Further studies are needed to evaluate and better characterize the active compounds identified in our study. This can be done by guided fractionation using HPLC to isolate and eliminate the active from the non-active compounds and to retest the active compounds with HCS. Furthermore, active compounds can be synthesized and tested in vivo.S1 Table(DOCX)Click here for additional data file.S2 TableThis figure were adapted from the cited publication.(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.S5 Table(DOCX)Click here for additional data file.S6 Table(DOCX)Click here for additional data file.S1 FigEight peaks were identified from the LC-MS chromatogram: (1) 185.98322 m/z, (2) 144.98225 m/z, (3) 144.98225 m/z, (4) 288.28995 m/z, (5) 256.26364 m/z, (6) 387.18086 m/z, (7) 415.21219 m/z, and (8) 637.30585 m/z.(TIFF)Click here for additional data file.S2 Fig4H5NO4S.Acesulfame-K, Chemical Formula: C(TIFF)Click here for additional data file.S3 Fig2H6ClO3P.Ethephon, Chemical Formula: C(TIFF)Click here for additional data file.S4 FigSphinganine, Chemical Formula: C17H37NO2.(TIFF)Click here for additional data file.S5 Fig16H33NO.Palmitic amide, Chemical Formula: C(TIFF)Click here for additional data file.S6 Fig22H26O6.Burseran or (+)Eudesmin, Chemical Formula: C(TIFF)Click here for additional data file.S7 Fig24H30O6.Estra-1,3,5(10)-triene-3,6beta,17beta-triol triacetate. Chemical Formula: C(TIFF)Click here for additional data file.S8 Fig29H48O15.Methyl 6-O--\u03b2-D-galactopyranoside. Chemical Formula: C(TIFF)Click here for additional data file.S9 FigThe spectrum was recorded at room temperature using a 600-MHz NMR spectrometer.(TIFF)Click here for additional data file.S10 FigThe spectrum was recorded at room temperature using a 600-MHz NMR spectrometer. This figure verifies the MS finding of the -6-methyl-\u03b2-D-glucopyranuronosyl]-\u03b2-D-galactopyranoside) molecules with signals of several CH3 groups around 1 ppm in (TIFF)Click here for additional data file.S11 FigA) Estra-1,3,5(10)-triene-3,6beta,17beta-triol triacetate B) Burseran C) (+)Eudesmin D) Extended aromatic region of the NMR spectrum.(TIFF)Click here for additional data file.S12 Fig(TIFF)Click here for additional data file.S13 Fig(TIFF)Click here for additional data file.S14 Fig(TIFF)Click here for additional data file."}
+{"text": "Magnetospirillum sp. 15-1. This strain was isolated from a planted fixed-bed reactor based on its ability to degrade toluene under anaerobic conditions. The genome assembly consists of 5.4\u00a0Mb in 28 contigs and 5,095 coding sequences containing the genes involved in anaerobic toluene degradation.Here, we report the draft genome sequence of  Betaproteobacteria strains, represented by Thauera aromatica and Azoarcus sp.  continuously fed with toluene (De novo assembly was conducted with the A5-miseq pipeline (http://www.geneious.com)  (coding percentage 98.82%). The assembled sequences were functionally annotated using the RAST online service  , generat (M.\u00a0magnetotacticum MS-1 (86%) (Magnetospirillum sp. XM-1 (Magnetospirillum sp. SO-1 (89%)  (M. bellicus VDY (77%) (Magnetospirillum sp. WD (77%)  , M.\u00a0magn-1 (86%) , Magnetosp. XM-1  (88%), M-1 (89%) , and M.\u00a0-1 (76%)  than to DY (77%)  and MagnWD (77%) . This fibss, bbs, and bam), as observed for Thauera aromatica K172 (bss operon of strain 15-1 against other anaerobic toluene degraders were performed through BLAST (https://blast.ncbi.nlm.nih.gov/Blast.cgi). Results showed similarities of 99% to Magnetospirillum sp. TS-6 and 80% to both T.\u00a0aromatica K172 (Azoarcus sp. (Magnetospirillum strains (excluding strain 15-1) showed that none of these strains contain genes related to aromatic compound degradation pathways.The genes for degradation of toluene under anaerobic conditions were found to be organized in three operons on the chromosome  under the accession no."}
+{"text": "Globally, endometrial cancer is the sixth leading cause of female cancer-related deaths. Non-atypical endometrial hyperplasia (EH), has a lifetime progression rate to endometrial cancer ranging from less than 5%, if simple without atypia, to 40%, if complex with atypia. Site specific, long-acting intrauterine devices (IUDs) provide fertility sparing, progestin-based EH medical management. It is unclear which IUD is most beneficial, or if progesterone sensitizing metformin offers improved outcomes. For resolution, PubMed searches for \u201cMirena\u201d or \u201cMetformin,\u201d \u201ctreatment,\u201d \u201cendometrial hyperplasia,\u201d or \u201cstage 1 endometrial cancer,\u201d were performed, yielding 33 articles. Of these, 19 articles were included. The 60 mg high-dose frameless IUD/20 mcg levonorgestrel has achieved sustained regression of Grade 3 endometrial intraepithelial neoplasia for 14 years. Case series on early stage endometrial cancer (EC) treatment with IUDs have 75% or greater regression rates. For simple through complex EH with atypia, the 52 mg-IUD/10\u201320 mcg-LNG-14t has achieved 100% complete regression in 6-months. Clearly, IUDs have an outcome advantage over oral progestins. However, studies on metformin for EH, and of progestins or metformin for early stage EC management are underpowered, with inadequate dose ranges to achieve significant differences in, or optimal outcomes for, the treatment modalities. Therefore, outcomes from the feMMe trial for the 52 mg-IUD/10\u201320 mcg-LNG-14t and metformin will fill a gap in the literature. Endometincrease . Therefoincrease . Insulinincrease . Based on unopposed estrogen driving malignant endometrial proliferation, progestins have been the mainstay for fertility sparing medical management of EH and Stage I, grade I EC ,8,12,13.p < 0.04, after adjusting for age, clinical stage, grade, chemotherapy treatment, radiation treatment and presence of hyperlipidemia [Metformin has been successfully used in a few cases of progestin nonresponsive atypical EH . Cell liipidemia . With inipidemia . This waipidemia . Progestipidemia . As metfIt will be shown that oral continuous or intermittent progestin has lower regression rates (higher progression rates), than do progestin containing IUDs. In other words, the locally- long-acting intrauterine progestin delivery provided by the 52 mg-IUD/10\u201320 mcg-LNG-14t, the 60 mg-IUD/14 mcg-LNG, the 60 mcg-HD-frameless-IUD/20 mcg-LNG and the 60 mcg-LD-frameless-IUD/14 mcg-LNG more effectively medically manages EH than does continuous or intermittent oral progestin. PubMed searches on June 3, 2017, search terms \u201cMirena treatment endometrial hyperplasia\u201d and \u201cmetformin treatment endometrial hyperplasia\u201d with the parameters English language, free full text, published from 2012 onwards yielded 25 articles. Of these, 1 was on contraception, 1 was redundant, 1 was concerned with diagnosis, 1 focused on PCOS, 1 focused on hysterectomy in obese patients, 1 included hysteroscopic resection, 1 explored progestin resistance, and 18 were included. PubMed searches on February 11, 2017, search terms \u201cMirena treatment stage 1 endometrial cancer\u201d and \u201cmetformin treatment stage 1 endometrial cancer\u201d with the parameters English language, free full text, published from 2012 onwards, yielded 8 articles. Of these, 3 were excluded as a duplicate, 2 were concerned with hysteroscopic resection as treatment or pretreatment followed by progestin treatment, 1 was concerned with ovarian cancer and 1 was concerned with advanced EC, leaving 1 inclusion. From the four PubMed searches, 19 articles were included, 14 articles were excluded. Hand search yielded seven additional included articles as shown in Progestins modulate endometrial glands\u2019 secretory differentiation, inhibit estrogen receptor function and endometrial cell mitosis, and are pro-apoptotic . ProgestGrade 2 endometrial endometrioid adenocarcinoma in an 18-year old nullipara with Class III obesity, NIDDM and PCOS was treated with a 52 mg-IUD/10\u201320 mcg-LNG-14t . CompletA multi-center randomized controlled trial (RCT) with 170 participants compared the 52 mg-IUD/10\u201320 mcg-LNG-14t to 10 mg MPA for 10 days monthly or continuously for EH treatment . Patholop = 0.001 [p = 0.001 [p = 0.30 [For further surveillance, 153 participants from the above trial participated in a national, multicenter RCT of the 52 mg-IUD/10-20 mcg-LNG-14t compared to cyclic MPA 10 mg for 10 days per cycle, and continuous MPA 10 mg daily for 6-month\u2019s treatment followed by reevaluation at 6, 12, 18, and 24 months . Of the  = 0.001 . Consist = 0.001 . Howeverp = 0.30 . It is ip = 0.30 .p = 0.013, this study, which did not use intent to treat analysis, was underpowered for the 18.9 percentage-point regression rate difference to achieve statistically significance [A RCT of 60 participants in total, compared EH treatment with the 52 mg-IUD/10\u201320 mcg-LNG-14t to MPA 10 mg orally daily for 12 days monthly for 3-months [p < 0.01) and 55.7% cyclic NET for simple hyperplasia respectively at 12 months [At 1-year follow up, a randomized controlled trial (RCT) with 59 patients assigned to the 52 mg-IUD/10\u201320 mcg-LNG-14t versus 61 patients assigned to cyclic norethindrone (NET) 15 mg daily for 3-week cycles, indicated that the 52 mg-IUD/10\u201320 mcg-LNG-14t was more effective than cyclic NET medical management of non-atypical EH ,21. InteA single case series of 20 women with non-atypical and atypical EH, achieving sustained, complete regression for an average of 32-months with the 60mg-IUD/14 mcg-LNG or 60 mg-HD-frameless-IUD/20 mcg-LNG has been reported . A 44-yep < 0.0001 [p < 0.00001), however, this was predominantly spotting which resolved in 1-year [p = 0.08) [p = 0.28), or cancer-induced death,  [Meta-analysis of three RCT with a total of 359 participants found that the 52 mg-IUD/10\u201320 mcg-LNG-14t prevented tamoxifen-induced endometrial polyp formation, odds ratio (OR) 0.18, 95% confidence interval (CI): 0.13 to 1.02, < 0.0001 . Interesn 1-year . The 52  = 0.08) . The 52  = 0.71) .Clearly the 52 mg-IUD/10\u201320 mcg-LNG-14t\u2019s 1 in 1000 uterine perforation risk and invasive placement in the uterus are limitations that oral medications lack. Studies with IUDs cannot be completely blinded, and cannot have an innocuous a placebo control as other RCT can have. Normally an IUD string is left palpable in the vagina for IUD removal. The presence of the IUD string alerts the participant to the in utero IUD. Furthermore, any object placed in utero can exert a mass effect on the endometrium and uterus, therefore, a plastic IUD lacking hormonal or metallic active component is not inert. Incomplete participant and trial personnel blinding contributes to a performance bias . The 52 Clearly, the 52 mg-, 60 mg-, and frameless-IUD have long acting reservoir adherence advantages over daily oral progesterone for endometrial protection. While depo progestin precludes immediate treatment discontinuation, IUDs can be removed and progestin diffusion immediately halted. Oral progestin must be continuous to assure EH regression . NeverthLike progestins, metformin is anti-proliferative on the endometrium ,13,17. MPhosphoinositide 3-kinase (P13K) proto-oncogene serine/threonine protein kinase B (Akt) mammalian target of rapamycin  signaling pathway inhibition, mitogen-activated protein kinase (MAPK) inhibition, and glucose metabolism changes form part of metformin\u2019s anti-proliferative effect ,11. MetfMetformin upregulates GLUT4 mRNA and protein which are normally decreased in PCOS in comparison to non-PCOS . Like prp = 0.002 [A case-controlled trial of 28 participants taking 850 mg twice daily for 7 to 30 days versus 12 participants not taking metformin found an average daily dose dependent 17.2% reduction in atypical EH and endometrioid EC Ki-67, 95% confidence interval (CI) \u22127.0%, \u221227.4%,  = 0.002 . This st = 0.002 . 2, p = 0.004 [p = 0.25 [A case series of five insulin resistant PCOS patients diagnosed by the Rotterdam criteria, who had Stage 1A Grade 1 EC treated with metformin 1000 mg daily and Cyproterone/Ethinyl Estradiol 2 mg/35 mcg 21 days per month for 6 months achieved 100% complete regression . This tr = 0.004 . HOMA-IRp = 0.25 . Cyprotep = 0.25 . Cyprotep = 0.25 . TherefoA pilot study compared megestrol acetate 160 mg daily to metformin 500 mg trice daily with megestrol acetate 160 mg daily for treatment of atypical EH . Only 16Metformin use is limited by its gastrointestinal adverse effect profile. Metformin is associated with abdominal pain, diarrhea, lactic acidosis, nausea and vomiting, and taste changes . MetformConsideration should be given to head-to-head trials of the 52 mg-IUD/10\u201320 mcg-LNG-14t and metformin combined with oral progesterone for EH and EC medical management. In vivo dose-escalation studies on metformin are needed to establish the optimal dose for EH treatment . MetformAromatase inhibitors that would reduce estrogen concentrations, gonadotrophin-releasing hormone agonists, and selective estrogen receptor modulators could be investigated for efficacy in preventing and/or treating EH and early stage EC . GLUT1 aProgestins have least three anti-proliferative mechanisms: anti-angiogenesis, estrogen receptor inhibition and IGF-1 inhibition . HoweverCyclic progestin has a 55.7 to 72% regression rate for EH, whereas continuous progestin has an 88 to 100% regression rate ,21. The p = 0.001 [p = 0.30 [Of note, with oral and intrauterine progestin therapy, premenopausal status and higher mean estrogen level are associated with EH recurrence,  = 0.001 . Conversp = 0.30 . While bHopefully, the feMMe trial, an international, phase II, RCT of the 52 mg-IUD/10\u201320 mcg-LNG-14t, metformin with the 52 mg-IUD/10\u201320 mcg-LNG-14t, and weight loss with metformin and the 52 mg-IUD/10\u201320 mcg-LNG-14t, for atypical EH and early stage EC patients will provide clarity on the effective of metformin concurrent with a progestin IUD . It is bIf the feMMe trial supports metformin use with progestin IUDs for medical management of EH and early stage EC, a combined metformin and LNG IUD may become a reasonable treatment modality. A combined metformin\u2013LNG IUD may also have contraceptive and antiglycemic benefits for female, reproductive age diabetics. However, as the feMMe trial excludes oral progestins, unanswered questions will remain after the feMMe trial. The comparative effectiveness of oral progestins combined with metformin versus that of the IUDs combined with metformin for EH or early stage EC medical management remains to be answered."}
+{"text": "Brucella species are the etiological agent of brucellosis in humans and animals. Here, we report the whole-genome sequence of Brucella melitensis strain CIIMS-BH-2, belonging to biovar 2. The draft assembly of CIIMS-BH-2 is 3.31\u2009Mb in size, with 57.2% G+C content. Brucella species. Brucella spp. are Gram-negative, nonmotile, and facultative intracellular coccobacilli. Brucellosis in humans is characterized by undulant fever, arthritis, and gastrointestinal complications. It also causes a negative impact on livestock and public health , B. abortus (cattle), B. suis (swine), B. canis (dogs), B. ovis (sheep), B. neotomae (wood rats), B. ceti , B. pinnipedialis , B. microti (voles and foxes), B. inopinata (humans), B. papionism (baboons), and B. vulpis (foxes)  .Brucella spp. lack classical virulence factors, they have the ability to adapt to various environmental conditions  using version 2 chemistry. The sequencing of the isolate was performed on an Illumina HiSeq 2500 platform. The de novo assembly was carried out using SPAdes (version 3.10.1)  . The seqBrucella melitensis 16M. As predicted by RAST, the CIIMS-BH-2 genome consists of 3,216 open reading frames (ORFs), including 3,113 protein-coding sequences with 67 RNAs (The 16S rRNA gene sequence of CIIMS-BH-2 revealed 100% similarity to that of  67 RNAs . FurtherBrucella melitensis CIIMS-BH-2 genome sequence was deposited in the GenBank database under accession numbers CP025680 and CP025681.The"}
+{"text": "Volume 29, no. 3, Page 326\u2013328, 2014Page 327, Legend for Fig. 1IncorrectFig. 1. Changes in viability (black line) and ATP levels (gray line) during carbon-starvation conditions.CorrectFig. 1. Changes in viability (gray line) and ATP levels (black line) during carbon-starvation conditions."}
+{"text": "Bevacizumab addiction to triplet chemotherapy, according to FIr-B/FOx schedule, as first-line treatment in young-elderly metastatic colorectal CANCER (MCRC) patients may be more effective. Tailored treatments show worse clinical outcome in unfit patients.Elderly patients were clinically evaluated according to age and comorbidity  to select FIr-B/FOx regimen in fit or tailored treatments in unfit elderly. Limiting toxicity syndromes (LTS) were evaluated.KRAS genotype. G3-4 toxicities were diarrhea 21%, mucositis 11%, neutropenia 11%. LTS were 46%, significantly more multiple than single site. At 8 months follow-up, in 37 unfit patients: ORR 37%, PFS 7 months, OS 13 months. PFS was significantly different in KRAS wild-type compared to mutant patients, while not OS. PFS and OS were significantly worse in KRAS c.35 G > A compared to wild-type and/or other mutant.At 17 months follow-up, in 28 young-elderly patients treated with first line FIr-B/FOx: objective response rate (ORR) 79%, progression-free survival (PFS) 11 months, overall survival (OS) 21 months. Clinical outcome was not significantly different according to KRAS, and specifically c.35 G > A mutant genotype, may significantly affect clinical outcome in patients unfit for FIr-B/FOx.Careful decision-making process including evaluation of patient's fitness, and individual safety should be included to select FIr-B/FOx intensive first line regimen in young-elderly MCRC patients.  RAS genotype [KRAS wild-type patients [KRAS genotype) represents a major challenge in clinical management of MCRC patients.Different treatment options and lines of medical treatment in metastatic colorectal cancer (MCRC) patients are currently tailored according to fitness , comorbidities), metastatic extension (liver-limited (L-L) or other/multiple metastatic (O/MM)), and genotype \u20137. Firstgenotype , 7, 8, 1genotype , 10, 6. patients , 6. The MCRC patients are prevalently elderly but often under-treated in clinical practice, and usually underrepresented in clinical trials. They require a decision-making process combining the evaluation of fitness, according to co-morbidity, functional, and nutritional status , and selNo impact on PFS and OS was observed by age and/or comorbidities in patients treated with FOLFOX or FOLFIRI added or not to cetuximab . AdditioRecent retrospective analyses reported by our group, evaluating clinical outcome and safety of first-line FIr-B/FOx intensive regimen in fit young-elderly MCRC patients , 33, or et al. [2/w), OXP (32.5 mg/m2/w), associated to bolus plus continuous infusion 5-FU  880 mg/m2/w (88%)) failed to confirm significantly improved clinical efficacy compared to FOLFIRI in unresectable MCRC patients [HORG-FOLFOXIRI schedule proposed by Soug-lakos et al. , charact, OXP 32. mg/m2/w,patients , 5.In an unplanned subgroup evaluation of the phase III study, prognostic relevance of elderly status in both FOLFIRI and FOLFOXIRI treated patients was evaluated , and cliHORG/FOLFOXIRI intensive regimen showed a worse safety profile compared to FOLFIRI, with significantly more dose reductions and treatment delays; in elderly compared with younger patients, significantly more dose reductions , treatment delays , particularly due to toxicity were reported . In the FOLFIRI arm, dose reductions  and treatment delays were not different. However, there was no significantly different rDI of drugs in elderly compared with younger patients, in both FOLFOXIRI and FOLFIRI arms.Grade 3-4 diarrhea was significantly more prevalent in elderly, compared to younger patients, in both chemotherapy regimens  . Moreove2, 12h-timed-flat-infusion, 2 days weekly; CPT-11 160 mg/m2/BEV 5 mg/kg or OXP 80 mg/m2, weekly alternating. Cumulative Index Rating Scale (CIRS) was used to evaluate the comorbidity status, and only patients with primary and intermediate CIRS stage were enrolled [We retrospectively evaluated consecutive young-elderly patients 65 to 75 years enrolled in the previously reported phase II trial  and in tenrolled . Primaryenrolled . LTS werKRAS wild-type patients, ORR was 92%, liver metastasectomies 23% (50% in L-L patients), median PFS 14 months (4-78+ months), median OS 38 months (8-78+ months). Among 13 KRAS mutant patients, ORR was 77%, liver metastasectomies 15%, (20% in L-L patients), median PFS 7 months (3-69+ months), median OS 19 months (6-69+ months). Neither PFS nor OS were significantly different in KRAS wild-type compared with mutant patients, according to log-rank test. No BRAF mutations were detected.Young-elderly patients were 28 (42%) among overall MCRC patients enrolled fitting for FIr-B/FOx intensive treatment, according to inclusion criteria, WHO PS 0 89%, CIRS primary/intermediate. At a median follow-up of 17 months, ORR was 79%, liver metastasectomies 18% (37.5% in L-L patients), median PFS 11 months (3-78+), median OS 21 months (6-78+) . Patients\u2019 distribution according to age and comorbidities was: young-elderly 22%, old-elderly 54%; CIRS stage primary 11%, intermediate 40%, secondary 42%. Medical treatment regimens were tailored according to age and comorbidity status in the individual patients. Eighteen patients (49%) were treated with triplet regimens; 15 patients (40%) with doublet regimens; 4 patients (11%) with mono regimens; 3 underwent up-front surgery.At a median follow-up of 8 months, ORR was 37%, median PFS was 7 months (1-13+), median OS 13 months (1+-23+). Among patients treated with triplet regimens, ORR was 37.5%, median PFS 8 months (3-12), median OS 12 months (3-23+ months) , median OS 13 months (1+-23+ months). Among 12 KRAS mutant patients evaluable for activity, ORR was 25%, median PFS 6 months (1-11 months), median OS 8 months (3-18 months). A significantly different PFS (p = 0.043), but not OS, was reported in KRAS wild-type compared with mutant patients. Significantly worse PFS and OS were reported in c.35 G > A KRAS mutant compared to wild-type , and to other mutant patients .Among 14 KRAS wild-type patients, demonstrated clinical outcome and safety profile equivalent to younger patients [Fit young elderly patients, PS<2, treated with triplet regimens consisting of chemotherapeutic drugs, or BEV addiction to doublets, or doublets plus EGFR-inhibitors in patients , 5, 8, 1patients , 34 obtaRetrospective analysis of randomized clinical trials showed that doublets CPT-11, or OXP, added to fluoropyrimidin in older patients eligible for clinical study reported ORR 18-59.4%, PFS 4.9-10.0 months and OS 8.5-20.7 months , 32, 30.Published studies showed that limiting toxicities were not significantly different in elderly patients treated with 5-FU or CPT-11 \u201316, sligKRAS wild-type compared to mutant patients. More, significantly worse clinical outcome (PFS and OS) may be influenced by KRAS c.35 G > A mutant genotype, compared to wild-type and/or other mutant, confirming that KRAS genotype, and specifically c.35 G > A mutant, confers different biological aggressiveness [KRAS genotype may significantly explain different PFS, and c.35 G > A KRAS mutant a significantly worse PFS and OS, compared to wild-type and other mutant.In clinical practice, selection of patients eligible for intensive medical treatment and to achieve optimal activity and clinical outcome could be performed by the evaluation of age, PS, CIRS and careful monitoring of individual safety using LTS Table . Patientsiveness \u201341, lessKRAS genotype, and specifically the prevalent KRAS c.35 G > A mutant status, that can discriminate significantly different clinical outcome particularly in unfit MCRC patients.We revised intensive medical treatment consisting of triplet chemotherapy regimens or more intensive triplet chemotherapy plus anti-angiogenic drug, bevacizumab, in elderly MCRC patients, that we previously developed, discussed compared with tailored medical regimens selected for unfit MCRC in clinical practice , 7, 32,"}
+{"text": "The correct name is: Victor W. M. van Hinsbergh. The correct citation is: Nauta TD, Duyndam MCA, Weijers EM, van Hinsbergh VWM, Koolwijk P (2016) HIF-2\u03b1 Expression Regulates Sprout Formation into 3D Fibrin Matrices in Prolonged Hypoxia in Human Microvascular Endothelial Cells. PLoS ONE 11(8): e0160700. doi:"}
+{"text": "Primulina cardaminifolia Yan Liu & W.B. Xu (Gesneriaceae), a distinct new species with imparipinnate leaves, is described and illustrated from a limestone valley in Guangxi Zhuangzu Autonomous Region, China. To assure its generic placement and phylogenetic affinity, phylogenetic analyses were performed using DNA sequences of nuclear ITS and chloroplast trnL-F intron spacer region. Additionally, somatic chromosome number was counted and pollen stainability was tested.Primulina; however, two phylogenetically distinct ITS sequence types were detected, suggesting a probable hybrid origin. Its pollen stainability is 100% and its chromosome number, 2n = 36, is congruent with all known counts of diploid species of the genus.Phylogenetic analyses support its placement in Primulina cardaminifolia and suggest that it could have derived from homoploid hybrid speciation. Color plates, line drawings and a distribution map are provided to aid in identification.All available data support the recognition of the new species The online version of this article (doi:10.1186/1999-3110-54-19) contains supplementary material, which is available to authorized users. Chirita D. Don  Mich. M\u00f6ller & A. Weber] made without support from molecular data by Weber et al. .Although the new generic circumscription reflects better the evolutionary relationships and ecological preferences of the genus, diagnostic characters of  rosette  and the r et al.  was recePrimulina, which is described and illustrated here. Chromosome count of the new species is in agreement with those reported for Primulina in Christie et al.  has been deposited in HAST. Root tips were gathered and pretreated in 2 mM 8-hydroxyquinoline at 15\u201318\u00b0C for about 6 h and fixed overnight in an ethanol-acetic acid solution (3:1) below 4\u00b0C. The chromosomes were stained and macerated in 2% acetic orcein with 1 N hydrochloric acid (10:1). Classification of chromosome complement based on centromere position at mitotic metaphase followed Levan et al.  and cycle-sequenced using the T7 and SP6 promoter primer pairs.DNA sequences of the nuclear ribosomal internal transcribed spacers (ITS) and the chloroplast u et al. . BecausetrnL-F regions as separated matrices. Primulina pinnata (W.T. Wang) YinZ. Wang, which was not included in Xu et al.  YinZ. Wang, and Didymocarpus podocarpus C.B. Clarke chosen as outgroups based on recent phylogenetic analyses  and maximum likelihood (ML) criteria implemented in MEGA5. MP trees were searched using the Tree-Bisection-Reconnection (TBR) search option with the initial trees setting at 50, MP search level setting at 5, and maximum number of trees setting at 2000. Clade supports were calculated based on 100 bootstrap resamplings . ML trees were reconstructed using the nearest-neighbor-interchange (NNI) method with all site used and the initial tree automatically selected under the model(s) selected by MEGA5. Clade supports of ML analysis were evaluated based on 100 bootstrap resamplings .For phylogenetic analyses, matrices of Xu et al.  were adou et al.  because Wei-Bin Xu & Yan Liu 08050 . \u788e\u7c73\u85ba\u8449\u5831\u6625\u82e3\u82d4 Figures\u00a0Yan Liu & W.B. Xu, sp. nov.\u2014TYPE: CHINA. Guangxi Zhuangzu Autonomous Region, Laibin Shi (City), Fenghuang Zhen (Township), alt. 280\u00a0m, on moist limestone rock face in a valley, 14 July 2008, Primulina cardaminifolia Yan Liu & W.B. Xu resembles Primulina pinnata (W.T. Wang) YinZ. Wang in having imparipinnate leaves, but is clearly distinct from this species by the ovate-cordate terminal leaflet of 3\u20137 \u00d7 3\u20136.5 cm, 1 or 2 pairs of broadly ovate to sub-ovate lateral leaflets, 1\u20133-branched cymes with 3 to 10-flowers, and the entire-margined calyx lobes with acuminate apex.Herbs perennial. Rhizome subterete, 3\u20135 mm across. Leaves 5\u20137, in basal rosette, imparipinnate, 10\u201320 cm long, papery when dry; petiole subterete, 6\u201312.5 cm long, densely pubescent; terminal leaflet ovate-cordate, 3\u20137 \u00d7 3\u20136.5 cm, apex obtuse, base cordate, margin repand to irregularly pinnately lobed, densely pubescent on both surfaces; lateral leaflets 1 or 2 pairs, opposite or alternate, broadly ovate to rotund, 1\u20133 \u00d7 1\u20133 cm, margin repand to irregularly pinnately lobed, densely pubescent on both surfaces, petiolules short, 2\u201310 mm long. Cymes 2\u20134, axillary, 1\u20133-branched, 3\u201310-flowered; peduncle 4\u20138 cm long, 1\u20132 mm in diam., densely pubescent; bracts 2, opposite, linear-lanceolate, 7\u20138 \u00d7 2\u20133 mm, margin entire, pubescent; pedicel 4\u20137 mm long, densely pubescent. Calyx 5-parted to base, lobes lanceolate-linear, 8\u201312 \u00d7 2\u20133 mm, apex acuminate, outside pubescent, inside glabrous, margins entire. Corolla white to pale purple, 3.2\u20133.5 cm long, outside glandular-pubescent, inside sparsely puberulent, with 2 pale-yellow stripes; corolla tube 2.1\u20132.3 cm long, 8\u201312 mm in diam. at the mouth, 2.5\u20133 mm in diam. at the base; limb pale purple, distinctly 2-lipped; adaxial lip 2-parted to over the middle, lobes oblong, 8\u201310 \u00d7 6\u20137 mm; abaxial lip 3-lobed to over the middle, lobes oblong, 12\u201313 \u00d7 4\u20135 mm; stamens 2, adnate to 1.5 cm above the corolla tube base; filaments linear, ca. 1.2 cm long, geniculate above the base, sparsely glandular-puberulent; anthers ca. 3 mm long, ca. 1.5 mm wide, dorsifixed, glabrous; staminodes 2, ca. 5 mm long, apex capitate, glabrous, adnate to ca. 6 mm above the base of corolla tube. Disc ring-like, ca. 1 mm in height, margin repand, glabrous. Pistil 2.5\u20132.8 cm long, ovary 7\u20138 mm long, ca. 1.5 mm across, puberulent; style 1.5\u20131.8 cm long, ca. 0.6 mm across, puberulent; stigma obtrapeziform, ca. 2 mm long, apex 2-lobed. Capsules not seen.CHINA. Guangxi Zhuangzu Autonomous Region, Laibin Shi, Fenghuang Zhen, 3 July 2008, Wei-Bin Xu & Yan Liu 08040 (IBK); same locality, 8 Sep 2008, Wei-Bin Xu & Kuo-Fang Chung 08472 (IBK), Shin-Ming Ku et al. 2035 (HAST); same locality, 28 June 2007, Hong-Jin Wei & Wei-Bin Xu 07244 (IBK).Primulina cardaminifolia is extremely rare, currently known only from the type locality in Laibin Shi, Guangxi Zhuangzu Autonomous Region, China .The specific epithet is derived from the leaves resembling those of the genus Primulina cardaminifolia resembles Primulina pinnata (W.T. Wang) YinZ. Wang  YinZ. Wang  and 634 (P. cardaminifolia-B) bp, respectively, while only one cpDNA sequence type was detected in the species. With the addition of these two ITS sequences, the ITS matrix contained 28 accessions of 675 aligned positions, of which 182 (26.96%) were parsimoniously informative. Based on the Kimura 2-parameter model using a discrete Gamma distribution (K2\u2009+\u2009G) with 5 rate categories selected by the corrected Akaike Information Criterion (AICc) implemented in MEGA5, a single ML tree (log likelihood\u2009=\u2009\u22124114.9869) was recovered  and low  supports in ITS and cpDNA datasets, respectively . Primulina cardaminifolia was collected from the type locality [S.M. Ku 2035(HAST)].GenBank accession numbers: Species: (ITS/Didymocarpus podocarpus C.B. Clarke: (DQ912688/FJ501514); Petrocodon dealbatus Hance: (FJ501358/FJ501537); Petrocodon scopulorum (Chun) YinZ. Wang [=Tengia scopulorum Chun]: (GU350637/GU350669); Primulina bipinnatifida (W.T. Wang) Yin Z. Wang [=Chiritopsis bipinnatifida W.T. Wang]: (DQ872842/DQ872806); Primulina cardaminifolia Yan Liu & W.B. Xu: ; Primulina cordifolia (D. Fang & W.T. Wang) YinZ. Wang [=Chiritopsis cordifolia D. Fang & W.T. Wang]: (DQ872845/DQ872803); Primulina dryas (Dunn) Mich. M\u00f6ller & A. Weber [=Chirita sinensis Lindl.]: (FJ501348/FJ501524); Primulina gemella (D. Wood) YinZ. Wang [=Chirita gemella D. Wood]: (FJ501345/FJ501523); Primulina glandulosa  Yin Z. Wang : (DQ872841/DQ872804); Primulina heterotricha (Merr.) YinZ. Wang [=Chirita heterotricha Merr.]: (DQ872826/DQ872816); Primulina linearifolia (W.T. Wang) YinZ. Wang [=Chirita linearifolia W.T. Wang]: (DQ872834/DQ872810); Primulina longgangensis (W.T. Wang) YinZ. Wang [=Chirita longgangensis W.T. Wang]: (FJ501347/AJ492290); Primulina luochengensis (Yan Liu & W.B. Xu) Mich. M\u00f6ller & A. Weber [=Wentsaiboea luochengensis Yan Liu & W.B. Xu]: (HQ633046/HQ632949); Primulina minutimaculata (D. Fang & W.T. Wang) YinZ. Wang [=Chirita minutimaculata D. Fang & W.T. Wang]: (DQ872828/DQ872815); Primulina multifida B. Pan & K.F. Chung: (JX507031/JX506756); Primulina ophiopogoides (D. Fang & W.T. Wang) YinZ. Wang [=Chirita ophiopogoides D. Fang & W.T. Wang]: (DQ872829/DQ872814); Primulina pinnata (W.T. Wang) YinZ. Wang [Chirita pinnata W.T. Wang]: (FJ501349/FJ501526); Primulina pinnatifida (Hand.-Mazz.) Yin Z. Wang [=Chirita pinnatifida (Hand.-Mazz.) B.L. Burtt]: (FJ501350/FJ501527); Primulina pseudomollifolia W.B. Xu & Yan Liu: (JX506869/JX506759); Primulina pteropoda (W. T. Wang) Yan Liu [=Chirita pteropoda W.T. Wang]: (DQ872827/DQ872817); Primulina repanda var. guilinensis (W.T. Wang) Mich. M\u00f6ller & A. Weber [=Chiritopsis repanda var. guilinensis W.T. Wang]: (DQ872846/DQ872808); Primulina spadiciformis (W.T. Wang) Mich. M\u00f6ller & A. Weber [=Chirita spadiciformis W.T. Wang]: (FJ501346/AJ492291); Primulina spinulosa (D. Fang & W.T. Wang) YinZ. Wang [=Chirita spinulosa D. Fang & W.T. Wang]: (DQ872830/DQ872813); Primulina tabacum Hance: (FJ501352/AJ492300); Primulina weii Mich. M\u00f6ller & A. Weber : (DQ872832/DQ872811); Primulina wentsaii (D. Fang & L. Zeng) YinZ. Wang [=Chirita wentsaii D. Fang & L. Zeng]: (DQ872831/DQ872812)."}
+{"text": "Listeria monocytogenes is a Gram-positive, rod-shaped, non-spore-forming bacterium which is an important foodborne bacterial pathogen for humans worldwide with high mortality rates. Here, we report a 2,964,284-bp draft genome sequence of Listeria monocytogenes strain ATCC 7644 (American Type Culture Collection). Listeria monocytogenes is a Gram-positive, rod-shaped, non-spore-forming bacterium and an important foodborne bacterial pathogen for humans worldwide with high mortality rates  isolated from a human was obtained from ATCC  and stored at \u221280\u00b0C. Genomic DNA was isolated from overnight cultures grown at 37\u00b0C on brain and heart infusion agar using the Qiagen DNeasy blood and tissue DNA purification kit . Three sequencing libraries were constructed from three independent cultures/DNA extractions using the Nextera XT DNA sample preparation kit , and library quality was analyzed using a BioAnalyzer . Libraries were sequenced with the Illumina MiSeq platform  using v3 reagent kits. A total of 2,889,385 300-bp paired-end reads were generated.http://jgi.doe.gov/data-and-tools/bbtools/). Preprocessed reads were assembled using SPAdes v3.10.1 , and overlapping pairs were merged (bbmerge) using the bbtools software suite ( v3.10.1  and poli v3.10.1 . Assemblh v1.1.1 , which ph v1.1.1 . Two of NGZM00000000. The version described in this paper is the first version, NGZM01000000.This whole-genome shotgun project has been deposited in DDBJ/ENA/GenBank under the accession no."}
+{"text": "P = 0.03, I2 = 0%; CC vs. GG: OR = 0.59, 95% CI = 0.36\u20130.97, P = 0.04, I2 = 0%; GC+CC vs. GG: OR = 0.61, 95% CI = 0.37\u20130.99, P = 0.05, I2 = 0%). In summary, current evidence indicates that the microRNA-608 rs4919510 G>C polymorphism maybe an important factor of DSCs susceptibility, especially in Caucasian population.Previous epidemiologic studies have revealed a possible association between microRNA-608 rs4919510 G>C polymorphism and digestive system cancers (DSCs) risk, but the results were not consistent. We therefore performed an updated meta-analysis to explore the association between microRNA-608 rs4919510 G>C polymorphism and DSCs risk. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to assess the relationship between the microRNA-608 rs4919510 G>C polymorphism and DSCs risk. Heterogeneity, cumulative analyses, sensitivity analyses, and publication bias were also conducted to examine the statistical power. Eight published articles with nine independent case-control studies involving 10,836 individuals were included in this meta-analysis. Overall, no significant association was found between microRNA-608 rs4919510 G>C polymorphism and DSCs risk in general populations. But some significant protective effects were observed in the subgroup of Caucasian population group in three genetic models (C vs. G: OR = 0.82, 95% CI, 0.68\u20130.99,  Digestive system cancers (DSCs), which comprise esophageal cancer (EC), gastric cancer (GC), hepatocellular cancer (HC), pancreatic cancer (PC) colorectal cancer (CRC) and other solid carcinoma, is one of the most common malignancies with increasing incidence and mortality worldwide  are the most common type of genetic variation in people. SNPs alter gene function and/or expression, consequently affecting downstream biologic pathways and increasing cancer risk  statement  were searched for relevant studies that focused on the association between microRNA-608 rs4919510 G>C polymorphism and DSCs risk from inception up to January 1, 2018. Only studies that were written in English and Chinese were included. Moreover, the bibliographies of the collected studies and relevant reviews were retrospected to identify additional articles. The following search terms and strategy was used :#1 microRNA 608#2 microRNA-608#3 mir 608#4 mir-608#5 rs4919510#6 #1 OR #2 OR #3 OR #4 OR #5#7 polymorphism#8 variant#9 mutation#10 #7 OR #8 OR #9#11 cancer#12 tumor#13 neoplasm#14 #11 OR #12 OR #13#15 #6 AND #10 AND #14Studies were included based on the following criteria: (1) only case-control studies that focusing on the association between the microRNA-608 rs4919510 G>C polymorphism and DSCs risk; and (2) studies had to provided sufficient frequency data on the genotype distribution to evaluate the crude odds ratios (ORs) and 95% confidence intervals (CIs); and; (3) studies had to be published only in English and Chinese; and (4) only publications with the largest or most recently updated sample data were included when there were some overlapping or duplicate publications on the same theme.Two researchers (Li and Song) independently reviewed and extracted relevant information from all included studies, including name of the first author, publishing date, country or region where the study was conducted, race, control design, sample sizes of the cases and controls, frequency data of genotype distribution, genotyping method, Hardy-Weinberg equilibrium (HWE) assessment in controls, and cancer type. Quality evaluation of the included studies was performed by the two researchers using the modified Newcastle-Ottawa scale (NOS). The scores ranged from 0 points (worst) to 11 points (best) Table ; studiesQ-test and I2 statistical method , co-dominant models (GC vs. GG and CC vs. GG), dominant model (GC+CC vs. GG), and recessive model (CC vs. GG+GC). Heterogeneity between the included studies was calculated using Cochran's A total of 164 articles were collected. The study selection process was presented in Figure P = 0.98, I2 = 53.4%; GC vs. GG: OR = 1.07, 95% CI = 0.93\u20131.24, P = 0.35, I2 = 43.0%; CC vs. GG: OR = 1.01, 95% CI = 0.83\u20131.24, P = 0.90, I2 = 56.4%; GC+CC vs. GG: OR = 1.05, 95% CI = 0.89\u20131.23, P = 0.58, I2 = 56.4%,  .Publication bias was examined with Begg's test and no apparent asymmetry of the funnel plot was found Figure . These rMicroRNAs is a family of small, noncoding, evolutionarily conserved endogenous RNAs consisting of 22 nucleotides that can regulate gene expression through a post-transcriptional pathway binding to the 3\u2032-UTR of target mRNAs , and the results were calculated without gene-gene and gene-environment risk factors, leading to a failure to interpret the potential interaction mechanisms. Third, almost all of the included studies focused on Asian and Caucasian populations, which would restrict the application of our results to other populations.In summary, our meta-analysis demonstrated that the microRNA-608 rs4919510 G>C polymorphism might play an important role in DSCs susceptibility. Further studies in different races and regions with larger population sizes are needed to confirm our findings.X-FL, J-KS, and Y-MN conceived the study. X-FL, J-KS, and J-WC searched the databases and extracted the data. J-KS, Y-QZ, and ML analyzed the data. X-FL, J-KS, and J-WC wrote the draft of the paper. JZ and Y-MN reviewed the manuscript. All the authors approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Molecular classifications of breast cancer have resulted in improved treatments. However, treatments for triple negative breast cancer (TNBC) are lacking. Analysis of molecular targets for TNBC is a priority. One potential candidate is androgen receptor (AR) phosphorylation. This study assessed the role of AR phosphorylation at ser81/ser515 and their two upstream effectors, cyclin-dependent kinase 1 (pCDK1) and extracellular-regulated kinase 1/2 (pERK1/2) in 332 ductal breast cancer patients by immunohistochemistry.p = 0.038), decreased size (p = 0.001), invasive grade (p < 0.001), necrosis (p = 0.003), b-lymphocytes (p = 0.020), molecular subtype (p < 0.001) and estrogen receptor (ER)/progesterone receptor (PR)-status (p < 0.001). The cohort was therefore stratified into ER+ve and ER-ve patients. In ER+ve tumours, pERK1/2 combined with AR-515 associated with improved CSS (p = 0.038), smaller size (p = 0.004), invasive grade (p = 0.001), decreased b-lymphocytes (p = 0.013) and increased plasma cells (p = 0.048). In contrast, in TNBC patients, phosphorylation of AR-515 associated with poorer CSS (p = 0.007). pERK1/2 combined with AR-515 associated with decreased inflammation (p = 0.003), increased tumour stroma (p = 0.003) and tumour budding (p = 0.011), with trends towards decrease CSS (p = 0.065) and macrophage levels (p = 0.093).pERK1/2 combined with AR-515 associated with improved cancer-specific survival (CSS, In Conclusions, AR-515 may be an important regulator of inflammation in breast cancer potential via ERK1/2 phosphorylation. AR-515 is a potential prognostic marker and therapeutic target for TNBC. Approximately 1.7 million cases of breast cancer are diagnosed every year. About 522,000 die of breast cancer yearly making it the 5th most common cause of cancer death globally . In the The majority of breast cancers comprise of luminal A and luminal B subtypes. These subtypes display oestrogen (ER) and/or progesterone receptors (PR) rendering these cancer cells susceptible to endocrine therapies e.g. anti-oestrogen therapies (Tamoxifen) , 8. In HIt has been shown that TNBC patients displaying pathological complete response (pCR) had better outcomes in terms of progression-free survival than patients who had not achieved pCR . pCR is Although numerous studies on AR have been published, the exact role of AR as tumour-promoting or anti-tumorigenic remains undecided. It is known that AR signalling plays a role in regulating cancer cell proliferation and apoptosis \u201319. AR aIt was also reported that when Tamoxifen-resistant breast cancer cells that expressed high levels of AR were exposed to bicalutamide (anti-androgen), cell growth was shown to decrease as well as reversing Tamoxifen resistance , indicatThe MAPK/ERK signalling cascade is heavily involved in the regulation of cancer cell proliferation, growth and differentiation. The stimulation of this pathway leads to the activation of a cascade of kinases , which ultimately leads to the activation of ERK1/2, which is associated with poor prognosis in breast cancer . One funin vitro kinase assays demonstrated an increase in ser-308 phosphorylation by CDK1 is responsible for Serine\u201381 phosphorylation [In contrast, AR phosphorylation at ser-81 is the prototypical site for dihydrotestosterone (DHT) activation, which is thought to be facilitated by activated Cdk1, as observed by Chen et al where Cdk1 inhibitors decreased ser-81 phosphorylation of AR in LNCaP cells and similarly decreased AR protein expression and transcriptional activity. Whereas transfected Cdk1 stimulated AR phosphorylation at ser-81 and increased AR protein expression . This surylation . Cdk1 phrylation .This study aims to further understand the role of androgens and AR activation in breast cancer by determining its effect on patient survival. This is followed by the determination of the role of upstream pathways (i.e. Cdk1 and ERK1/2 pathways) on AR activation and their effects on cancer specific survival (CSS). We aim to confirm that the phosphorylation of AR at either ser-515 or ser-81 is a surrogate for AR activation and assess if the pathways responsible for this phosphorylation are potential targets for therapeutic intervention.n = 332, Figure Only patients with staining for AR, AR-81 and AR-515 were included in the analysis (p = 0.842) nor AR-515 (p = 0.563) were associated with CSS alone. Therefore, upstream effectors of AR phosphorylation were investigated. AR-81 significantly correlated with nuclear CDK1 (p < 0.001) and AR-515 correlated with nuclear pERK1/2 (p < 0.001) as expected. However, AR-81 still showed no associations even when combined with CDK1 in the nucleus (p = 0.344). In contrast, AR-515 was significantly associated with improved CSS when combined with ERK1/2 phosphorylation in the nucleus . When nuclear pERK1/2 was assessed alone it was also significantly associated with improved CSS (p = 0.004). Therefore, further analysis was confined to nuclear pERK1/2 and AR-515.The relationship between nuclear AR activation and CSS is shown in Table p = 0.001), lower grade (p < 0.001), ER/PR+ve tumours (p < 0.001), molecular subtype (p < 0.001), lower necrosis (p = 0.003), weak Klintrup-Makinen (KM) grade (p = 0.004) and lower b-lymphocyte levels (p = 0.020). There was also a trend towards decreased HER2 (p = 0.096) and lower lymphovascular invasion (p = 0.099). When pERK1/2 was combined with AR-515 significant associations were still seen with smaller size (p = 0.017), lower grade (p < 0.001), ER+ve tumours (p = 0.013), PR+ve tumours (p = 0.008), decreased HER2 (p = 0.041), molecular subtype (p < 0.001), lower necrosis (p = 0.006), weak KM grade (p = 0.005) and lower b-lymphocytes (p = 0.023). Significant associations were now also seen with decreased proliferation (p = 0.009), lymphovascular invasion (p = 0.017) and increased tumour budding (p = 0.047).The relationship between pERK1/2, AR-515, clinicopathological characteristics, and local inflammatory response is presented in Table p = 0.008). As pERK1/2 and AR-515 strongly correlated in ER+ve patients (p < 0.001) they were combined. However, when combined the survival effect was weakened (p = 0.038), suggesting that pERK can also act independently of AR-515 in ER+ve breast cancers. When ER+ve patients where stratified further, there was no difference in CSS between luminal A and luminal B subtypes for pERK1/2 alone or combined with AR-515 therefore further analysis was carried out in all ER+ve tumours (n = 228).As pERK1/2 plus AR-515 associated with molecular subtype, we firstly stratified patients into ER+ve and ER-ve tumours . pERK1/2p = 0.004), lower grade (p = 0.001), molecular subtype (p = 0.024), decreased proliferation (p = 0.006), lower b-lymphocytes (p = 0.013) and increased plasma cells (p = 0.048). There was also a trend towards decreased necrosis (p = 0.096). When pERK1/2 was combined with AR-515 significant associations where still seen with lower grade (p = 0.002), molecular subtype (p = 0.001), decreased proliferation (p = 0.001), lower b-lymphocytes (p = 0.006) and increased plasma cells (p = 0.023). Significant associations were now also seen with decreased necrosis (p = 0.009) and lymphovascular invasion (p = 0.029).The relationship between pERK1/2, AR-515, clinicopathological characteristics, and local inflammatory response in ER+ve tumours is presented in Table p = 0.023) may be due to differences between the two ER-ve subtypes. Therefore, ER-ve patients were stratified into HER2-type and TNBC to further investigate these effects (p = 0.010), however no significant effect on CSS was observed for pERK1/2 alone (p = 0.534) and only a trend towards poorer outcome was seen when combined with AR-515 (p = 0.065). However, high levels of AR-515 combined with total AR, representing full AR activation, associated with a significantly poorer outcome  and no significant associations were seen in these patients. However, there was a trend towards improved patient survival with AR-515 phosphorylation (p = 0.056) alone and when combined with pERK1/2 (p = 0.076), confirming our hypothesis that these two ER-ve subtypes respond differently to AR-515 phosphorylation.It was observed in patients with ER-ve breast cancer that high pERK1/2 plus AR-515 had a trend towards an effect on survival rate. It was hypothesised that the loss of significance and weaker correlation between pERK1/2 and AR-515 in ER-ve patients ( effects . In patip = 0.033), lower KM grade (p = 0.004), increased tumour-stroma percentage (TSP) (p = 0.033) and increased tumour budding (p = 0.007). When combined with pERK1/2 associations were still seen with lower KM grade (p = 0.003), increased TSP (p = 0.003) and increased tumour budding (p = 0.011). Trends towards association were also seen for increased age (p = 0.075) and decreased macrophages (p = 0.093).Due to the detrimental effect of AR-515 phosphorylation in TNBC, associations with clinicopathological characteristics were assessed Table . Full acp = 0.002), invasive grade (p = 0.013), lymph node involvement (p = 0.009), necrosis (p < 0.001), proliferation index (p < 0.001), lymphatic invasion (p < 0.001), cytotoxic t-lymphocytes (p = 0.005), TSP (p = 0.009), tumour budding (p = 0.001) and angiogenesis (p = 0.043) were significantly associated with poorer CSS. Lymphovascular invasion (p = 0.054) and b-lymphocytes (p = 0.064) trended towards significance.For ER+ve tumours, since pERK1/2 plus AR-515 had significant associations with improved patient survival, these factors were taken into univariate and multivariate analysis (Table p = 0.036), proliferation index (p = 0.014), cytotoxic t-lymphocytes (p = 0.001), and TSP (p = 0.033) were independent prognostic factors, with lymphatic invasion (p = 0.060), tumour budding (p = 0.084), angiogenesis (p = 0.051) and pERK1/2 (p = 0.073) showing trends towards independence. However, pERK1/2 plus AR-515 does not appear to be independently prognostic in these patients (p = 0.682).On multivariate survival analysis including pERK1/2 plus AR-515; necrosis (p = 0.022), lymph node involvement (p < 0.001), necrosis (p = 0.030), lymphatic invasion (p = 0.001), lymphvascular invasion (p < 0.001), macrophages (p = 0.011) and cytotoxic t-lymphocytes (p = 0.047) were significantly associated with poorer CSS. Age (p = 0.055) and plasma cells (p = 0.055) trended towards significance.For TNBC, since AR-51 plus total AR had significant associations with poorer patient survival, these factors were taken into univariate and multivariate analysis (Table 7). On univariate analysis, size (p = 0.006), lymphovascular invasion (p = 0.002), and macrophages (p = 0.002) were independent prognostic factors. However, AR-515 plus total AR (p = 0.957) nor AR alone (p = 0.801) appear to be independently prognostic in these patients.On multivariate survival analysis including AR-515 plus total AR; lymph node involvement . The study was approved by the Research Ethics Committee of the West Glasgow University Hospitals NHS Trust.Patients presenting with invasive ductal breast cancer at Glasgow Royal Infirmary, Western Infirmary and Stobhill Hospital between 1995 and 1998 with formalin-fixed paraffin embedded blocks of the primary tumour available for evaluation were studied  were retrieved from the routine reports. Tumour grade was assigned according to the Nottingham Grading System. ER and PR status were assessed on tissue microarrays (TMAs) using immunohistochemistry (IHC) with Dako  ER antibody and Leica  PR antibody and scored according to the American Society of Clinical Oncology and College of American Pathologists guidelines with a cut-off value of 1% positive tumour nuclei . Her2 stFull-section haematoxylin and eosin (H&E) slides for the 474 patients were used to score local inflammatory infiltrate according to Klintrup-Makinen (KM) criteria. KM scoring of slides was carried out as previously described. Briefly, tumours were scored on four-point scores based on appearances at the tumour invasive margin. A score of 0 signified that there were no inflammatory cells at tumour's invasive margin; score 1 indicated a mild and patchy inflammatory cells; score 2 denoted a prominent band-like inflammatory reaction at the invasive margin; and score 3 revealed a florid cup-like inflammatory infiltrate at the invasive edge , 44. IndFull-section H&E slides were also used to score the tumour stroma percentage (TSP) as previously reported . BrieflyLymphatic and blood vessel (lymphovascular) invasion were assessed, on 2.5-\u03bcm thick sections, using IHC staining with the lymphatic endothelial marker D2-40  diluted 1:100 and vascular endothelial marker Factor VIII  diluted 1:100 as previously described . Ki67 prThe molecular subtypes were defined as follows: Luminal A: oestrogen (ER) and/or progesterone receptor (PR) positive, Her-2 negative or low proliferative index (< 15%); Luminal B: hormone receptor positive, Her-2 positive or high proliferative index (> 15%); Her-2 subtype: Her-2 positive and hormone receptor negative, any proliferative index; and triple negative: Her-2 negative, hormone receptor negative, any proliferative index.2O2. Sections were further blocked using 5% horse serum (10% casein for pERK1/2) in Tris-buffered saline (TBS). Antibodies for pCdk1 (Abcam), pERK1/2 , AR-81 , AR-515  were incubated overnight at 4\u00b0C diluted in Dako antibody diluent at 1:150 (Dako UK Ltd.) Total AR antibody (Dako UK Ltd.) was incubated overnight at 4C at 1:4000. Bound antibody complex was visualised using EnVision plus kit (Dako UK Ltd.) followed by 3,3-diaminobenzidine tetrahydrochloride . Nuclei were counterstained with haematoxylin and blued with Scots Tap Water Substitute, dehydrated through graded alcohol and xylene and mounted with Di-N-Butyl Phthalate in xylene (DPX).Antibody validation for AR-81, AR-515, pERK1/2 and pCDK1 was carried out as previously described , 26. ImmTMAs were scanned using the Hamamatsu NanoZoomer  at \u00d7 20 magnification, and visualization was carried out using Slidepath Digital Image Hub, version 4.0.1 .Only nuclear expression was scored as this represents the active form of each protein. Tissue staining intensity was scored by two blinded independent observers using a weighted histo-score (H-score) method , 49. Then = 332). Cut-off values for high or low protein expression were determined by the median of the average values of the intensity of staining. The relationships between variables were assessed using contingency table analysis with the X2 test for linear trend. Correlations coefficients were analysed using a Spearman's rho. Kaplan\u2013 Meier curves with log rank analysis was used to examine the effect of protein expression on CSS. Univariate survival analysis was performed using Cox proportional hazards regression. Variables with P-value of < 0.05 were entered into a multivariable model using a backwards conditional method for all patients and triple negative patients. All statistical analyses were two-sided and significance defined as P-value < 0.05. All statistical analysis was performed using the SPSS software version 22 .Only patients with a score for AR, AR-81 and AR-515 were included in the analysis ("}
+{"text": "The absolute configurations of four terpenes were determined based on their optical rotary powers. Incubation experiments with  R)-(+)-limonene is found in citrus fruits. Cineol is one of the main constitutents of eucalyptus oil and (+)-carvone is present in caraway. Some famous sesquiterpenes are \u03b1-humulene from hops, \u03b1-patchoulene from patchouli oil, and \u03b2-cedrene from juniper D = +120) , wh, whStrepades ago , the fultructure . The sig carbons . The HMB = +120) , while i2+ cofactor to which in turn the substrate\u2019s diphosphate portion is bound D24 = +7.1 . This points to the same enantiomer as reported from the heartwood of Cryptomeria japonica D22 = \u221251.3 . The opposite enantiomer was reported from Eucalyptus macarthuri ([\u03b1]D = +30.5) + calcd for C15H24+, 204.1873; found, 204.1872; IR (diamond ATR) : 2964 (m), 2925 (m), 2873 (m), 1670 (w), 1447 (w), 1412 (w), 1377 (w), 1259 (s), 1090 (s), 1016 (s), 866 (w), 797 (s), 700 (w), 662 (w) cm\u22121.I = 1483 ); MS (EI(+)-T-Muurolol (2): GC (HP 5): I = 1640 (literature (HP 5): I = 1640 + calcd for C15H26O+, 222.1978; found, 222.1982; IR (diamond ATR) : 3353 , 2956 (m), 2925 (m), 2869 (m), 1722 (m), 1639 (w), 1446 (m), 1376 (s), 1319 (m), 1258 (w), 1180 (m), 1081 (s), 955 (m), 910 (w), 837 (m), 802 (w) 614 (w), 539 (w) cm\u22121.I = 1493 ); MS (EI(\u2212)-7-epi-\u03b1-Eudesmol (4): GC (HP 5): I = 1661 (literature (HP 5): I = 1662 + calcd for C15H26O+, 204.1873, found. 204.1870; IR (diamond ATR) : 3388 , 2966 (s), 2910 (s), 2850 (s), 1662 (w), 1440 (s), 1376 (w), 1281 (m), 1260 (m), 1220 (s), 1139 (m), 1101 (m), 1021 (s), 937 (w), 873 (m), 841 (s) 798 (w), 755 (w), 707 (w), 649 (w), 568 (w) cm\u22121.(\u2212)-\u03b3-Cadinene (5): GC (HP 5): I = 1514 (literature (HP 5): I = 1513 + calcd for C15H24+, 204.1873; found, 204.1865; IR (diamond ATR) : 2925 (s), 2850 (s), 1525 (m), 1458 (s), 1366 (s), 1302 (s), 1192 (w), 1087 (s), 1030 (s), 963 (m), 921 (w), 823 (s), 691 (m) 608 (s) cm\u22121.I = 1513 ); MS (EI(\u2212)-(E)-\u03b2-Caryophyllene (6): GC (HP 5): I = 1429 (literature (HP 5): I = 1428 + calcd for C15H24+, 204.1873; found, 204.1868; IR (diamond ATR) : 3066 (w), 2924 (s), 2856 (s), 1670 (m), 1631 (m), 1449 (s), 1382 (s), 1367 (s), 1276 (m), 1257 (w), 1227 (w), 1182 (m), 1106 (w), 1067 (w); 1019 (w), 936 (m), 885 (s), 842 (m), 813 (m), 764 (w) 742 (s), 641 (w), 544 (m) cm\u22121.I = 1428 ); MS (EIE. coli BL 21 transformants was inoculated from an overnight preculture. The cells were grown to an OD600 = 0.4 before IPTG (0.4 mM) was added. The cultures were incubated at 18 \u00b0C and 160 rpm overnight. E. coli cells were harvested by centrifugation at 4 \u00b0C and 8000 rpm for 10 min. Protein purification was performed by Ni2+-NTA affinity chromatography with Ni2+-NTA superflow (Novagen) as reported above. For the incubation experiment in 2H2O with 7-epi-\u03b1-eudesmol synthase and (6-13C)FPP, the soluble enzyme fraction was washed with binding buffer  and then eluted twice with 10 mL elution buffer . Each pure protein fraction from 0.5 L 2YT liquid culture was concentrated with a Vivaspin20 concentration tube  for 0.5 to 1.5 h at 6000 rpm to 2 mL enzyme fraction. Incubation experiments were performed with the pure protein (2 mL) and the 13C-labeled substrate (0.8 mg to 3.0 mg in 2 mL 2H2O or H2O) at 28 \u00b0C overnight. The reaction mixture was extracted with 2 \u00d7 0.4 mL (2H6)benzene and directly measured by NMR.For each incubation experiment a 0.5 L 2YT liquid culture (containing kanamycin (50 mg/L)) of File 11\u20135.Gas chromatograms of extracts from enzyme reactions and NMR spectra of compounds"}
+{"text": "GNB3) gene has been implicated in obesity risk; however, the molecular mechanism of GNB3-related disease is unknown. GNB3 duplication is responsible for a syndromic form of childhood obesity, and an activating DNA sequence variant (C825T) in GNB3 is also associated with obesity. To test the hypothesis that GNB3 overexpression causes obesity, we created bacterial artificial chromosome (BAC) transgenic mice that carry an extra copy of the human GNB3 risk allele. Here we show that GNB3-T/+ mice have increased adiposity, but not greater food intake or a defect in satiety. GNB3-T/+ mice have elevated fasting plasma glucose, insulin, and C-peptide, as well as glucose intolerance, indicating type 2 diabetes. Fasting plasma leptin, triglycerides, cholesterol and phospholipids are elevated, suggesting metabolic syndrome. Based on a battery of behavioral tests, GNB3-T/+ mice did not exhibit anxiety- or depressive-like phenotypes. GNB3-T/+ and wild-type animals have similar activity levels and heat production; however, GNB3-T/+ mice exhibit dysregulation of acute thermogenesis. Finally, Ucp1 expression is significantly lower in white adipose tissue (WAT) in GNB3-T/+ mice, suggestive of WAT remodeling that could lead to impaired cellular thermogenesis. Taken together, our study provides the first functional link between GNB3 and obesity, and presents insight into novel pathways that could be applied to combat obesity and type 2 diabetes.The G-protein beta subunit 3 ( LEP) as the first obesity gene [delta]Ct method using Inguinal WAT (iWAT) and BAT were dissected, weighed and immediately homogenized on ice in CelLytic MT Mammalian Tissue Lysis/Extraction Reagent  with protease inhibitor cocktail. Total protein concentration was determined using Pierce BCA Protein Assay Kit .Citrate synthase activity for iWAT and BAT extracts were measured using the Citrate Synthase Assay Kit  following the manufacturer\u2019s instructions.t-test was used to compare two groups, and one-way ANOVA was used to compare more than two groups. Comparisons with p-values <0.05 were considered significant.Statistical analyses were performed using GraphPad Prism version 6 for Mac . Data are presented as mean \u00b1 SD (or SEM where indicated). Unpaired Student\u2019s GNB3-T/+. To determine the expression levels of human GNB3 and endogenous Gnb3 we performed quantitative RT-PCR in RNA from whole brain, olfactory bulb, hypothalamus, and cerebellum of 5-week-old GNB3-T/+ and WT mice. As expected, we did not detect human GNB3 in WT mice. Notably, human GNB3 expression was much greater than endogenous Gnb3 in whole brain, olfactory bulb, hypothalamus and cerebellum of GNB3-T/+ mice as calculated by delta cycle threshold values , and 50-fold greater than endogenous Gnb3 in iWAT and BAT  were not significantly different in  WT mice . At 20 wther sex . Inflamm obesity . Growth ale mice . It is iGNB3-T/+ mice could be due to greater food intake, lack of activity, or a metabolic defect in energy expenditure. To investigate these possibilities, we measured food consumption at ages 5, 10, 15, 20 and 25 weeks  in male mice. At 5 weeks old, VO2, heat and RER measurements are similar in GNB3-T/+ and WT mice , this difference was not significant.In our transgenic model, GNB3-T/+ mice, indicated by elevated fasting plasma glucose levels and GTT response. Though GNB3-T/+ mice did not have glucose intolerance prior to obesity, 5-week old female GNB3-T/+ mice had elevated fasting blood glucose, and both female and male GNB3-T/+ mice had elevated fasting plasma insulin. This could indicate the beginning stage of impaired glucose metabolism prior to obesity. However, GNB3-T/+ mice do not respond to the ITT like other mouse models of type 2 diabetes, indicating a milder phenotype.Glucose intolerance and type 2 diabetes are apparent at 20 weeks in GNB3-T/+ mice, consistent with lower circulating GH in obese humans [GNB3-T/+ mice at 20 weeks. Thyroid hormones control multiple physiological systems and have an important role in regulating basal metabolic rate, lipolysis, as well as the differentiation process in the adipose tissue [GNB3-T/+ mice at 20 weeks, which could indicate hypogonadotropic hypogonadism [GH was reduced in obese e humans . Anothere tissue . FSH, LHgonadism .GNB3-T/+ mice could be due to increased calorie intake, reduced activity, or a defect in metabolism that results in lower energy expenditure. Our results from food intake measurements and levels of fasting ghrelin, PYY, and amylin hormones revealed that hyperphagia or a satiety defect are not responsible. Novelty suppressed feeding tests also show no significant difference in the amount of sucrose eaten, latency to feed, or total feeding time between the GNB3-T/+ and WT mice, indicating that anxiety-like feeding behaviors are not involved in GNB3-related obesity. GNB3-T/+ and WT mice do not have a statistically significant difference in locomotor activity or oxygen consumption. However, it is possible that subtle differences in locomotion and/or oxygen consumption could contribute to increased adiposity. Future energy expenditure experiments conducted at thermoneutrality and/or brown fat induction experiments could shed light on the effects of GNB3 overexpression. Further, behavioral assessments indicate that there are no substantial anxiety- or depressive-like phenotypes in GNB3-T/+ mice, and that these affective phenotypes are unlikely to add to the relationship between GNB3 overexpression and obesity.Obesity is caused by an energy imbalance between calories consumed and calories expended. The increased adiposity in GNB3-T/+ mice. Ucp1 in mitochondria dissipates chemical energy in the form of heat, mainly in BAT, through a process called nonshivering thermogenesis [+ cells [+ adipose tissue which has a gene expression pattern more similar to beige cells than to classic brown cells in the mouse [GNB3 overexpression alters the gene expression profiles of both BAT and WAT, GNB3 appears to have the greatest effect in beige-cell containing iWAT.Ucp1 expression in adipose tissues provides a clue to the underlying defect in ogenesis . Recentlogenesis . In addi[+ cells \u201348 known[+ cells  or brown[+ cells . Beige c[+ cells  and deve[+ cells . Adult hhe mouse . Though GNB3-T/+ mice has lower expression of beige and brown adipocyte markers. Additionally, GNB3-T/+ mice showed markedly worsened beige adipocyte function in subcutaneous fat pads as indicated by lower levels of Ucp1 in iWAT. Subcutaneous adipose tissue in GNB3-T/+ mice acquired properties of visceral fat indicated by elevated expression of adipogenic markers, Pparg and adiponectin. Overall, GNB3 overexpression stimulates a conversion of subcutaneous WAT, particularly in the inguinal depot, into a less UCP1+ and a less beige but whiter tissue. We show that this white-like remodeling of iWAT and loss of brown and beige properties in BAT is accompanied by increased adiposity in mice fed normal chow. Together, these data implicate GNB3 overexpression in impaired WAT and BAT, and for the first time provide a functional link between GNB3 and obesity pathogenesis. However, the specific causes of GNB3-related obesity remain to be determined. Future studies of GNB3 overexpression are needed to dissect the molecular mechanisms by which GNB3 alters adipose tissue metabolism, signaling, and energy expenditure.BAT in S1 Fign = 8\u201311 mice per group in A, C, and D; n = 1\u20138 mice per group in B; n = 14\u201322 mice per group in E, F. Data are \u00b1 SD. No significant difference between GNB3-T/+ and WT of same sex by unpaired Student\u2019s t-test.A. gWAT weight/body weight (gWAT%), B. iWAT weight/body weight (iWAT%), C. BAT weight/body weight (BAT%), D. liver weight/body weight (liver%), E. DXA lean mass and F. DXA fat mass of 5-week-old mice. (TIF)Click here for additional data file.S2 Fign = 8\u201311 mice per group in A, n = 7 mice per group in B-D, n = 9\u201313 mice per group in E-H, n = 10\u201314 mice per group in I, n = 11\u201315 mice per group in J. Data are \u00b1 SD. *P < 0.05, **P < 0.01 vs. WT of same sex by unpaired Student\u2019s t-test.A. Fasting blood glucose, B. fasting plasma insulin, C. C-peptide, D. leptin, E. triglycerides, F. cholesterol, G. phospholipids and H. non-esterified fatty acids. I. GTT and AUC of female and J. male mice. All mice are 5 weeks old. (TIF)Click here for additional data file.S3 FigGNB3-T/+ mice have elevated fasting plasma resistin during obesity.Male  A,B. Fasting plasma glucagon; C,D. resistin; E,F. GIP; G,H. IL-6; I.J., TNF-a; and K,L. MCP-1 in 5-week and 20-week-old mice, respectively. n = 7 mice per group in A, C, E, G, I, K; n = 10\u201313 mice per group in B, n = 11\u201320 mice per group in D, F, H, J, L. Data are \u00b1 SD. *P < 0.05 vs. WT of same sex by unpaired Student\u2019s t-test.(TIF)Click here for additional data file.S4 FigGNB3-T/+ male mice have lower GH, TSH, FSH, LH and prolactin during obesity. A,B. Fasting plasma growth hormone (GH); C,D. thyroid-stimulating hormone (TSH); E,F. follicle-stimulating hormone (FSH); G,H. luteinizing hormone (LH); I,J. prolactin; K,L. adrenocorticotropic hormone (ACTH) levels of 5 week and 20 week old mice, respectively. n = 7 mice per group in A, C, E, G, I, K; n = 11\u201320 mice per group in B, D, F, H, J, L. Data are \u00b1 SD. *P < 0.05, **P < 0.01, ***P < 0.001 vs. WT of same sex by unpaired Student\u2019s t-test.(TIF)Click here for additional data file.S5 Fign = 7 mice per group in A-C, n = 19\u201321 mice per group in D-F. Data are \u00b1 SD in A-C, and \u00b1 SEM in D-F. *P < 0.05, **P < 0.01, ***P < 0.001 vs. WT of same sex by unpaired Student\u2019s t-test.A. Fasting plasma ghrelin, B. PYY and C. amylin. D. Total movement in X-plane, E. horizontal ambulatory activity, and F. vertical activity averaged over a 72-hour period. All mice are 5 weeks old. (TIF)Click here for additional data file.S6 Fig2, B. heat produced normalized over body weight, and C. RER (VCO2/VO2) averaged over a 72-hour period. D. Acute cold stress at 4\u00b0C. All mice are 5 weeks old. n = 19\u201321 mice per group in A-C, n = 5\u20138 mice per group in D. Data are \u00b1 SEM in A-C; and \u00b1 SD in D-F. ****P < 0.0001 vs. WT of same sex by unpaired Student\u2019s t-test.A. VO(TIF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 TableGNB3-T/+ mice and WT littermates were subjected to behavioral tests in order to evaluate anxiety/depressive-like behaviors and learning and memory.(DOCX)Click here for additional data file."}
+{"text": "Nature Communications7: Article number: 11929; DOI: 10.1038/ncomms11929 (2016); Published 06222016; Updated 03172017In this Article, residues D128 and D132 of ASC are consistently referred to incorrectly as D130 and D134, respectively. These errors appear in the Results, Methods, Fig. 2, Fig. 3 and Supplementary Fig. 1."}
+{"text": "Enterobacter cloacae is a major nosocomial pathogen causing bloodstream infections. We retrospectively conducted a study to assess antimicrobial susceptibility and phylogenetic relationships of E. cloacae bloodstream isolates in two tertiary university-affiliated hospitals in Shanghai, in order to facilitate managements of E. cloacae bloodstream infections and highlight some unknowns for future prevention.E. cloacae bloodstream isolates were consecutively collected from 2013 to 2016. Antimicrobial susceptibility was determined by disk diffusion. PCR was performed to detect extended-spectrum \u03b2-lactamase (ESBL), carbapenemase and colistin resistance (MCR-1) gene. Plasmid-mediated AmpC \u03b2-lactamase (pAmpC) genes were detected using a multiplex PCR assay targeting MIR/ACT gene  and other plasmid-mediated genes, including DHA, MOX, CMY, ACC, and FOX. eBURST was applied to analyze multi-locus sequence typing (MLST).Fifty-three non-duplicate E. cloacae isolates, 8(15.1%) were ESBL producers, 3(5.7%) were carbapenemase producers and 18(34.0%) were pAmpC producers. ESBL producers bear significantly higher resistance to cefotaxime (100.0%), ceftazidime (100.0%), aztreonam (100.0%), piperacillin (87.5%), tetracycline (75.0%), and trimethoprim-sulfamethoxazole (62.5%) than non-producers (p<0.05). PAmpC- and non-producers both presented low resistance rates (<40%) to all antibiotics (p>0.05). SHV  and MIR/ACT  predominated in ESBL and pAmpC producers respectively. Moreover, 2 isolates co-carried TEM-1, SHV-12, IMP-26 and DHA-1. MLST analysis distinguished the 53 isolates into 51 STs and only ST414 and ST520 were assigned two isolates of each (2/53).The rates of resistance to all tested antibiotics were <40%. Among 53 E. cloacae bloodstream isolates in the two hospitals. Multiclonality disclosed no evidence on spread of these isolates in Shanghai. The simultaneous presence of ESBL, carbapenemase and pAmpC detected in 2 isolates was firstly reported in Shanghai, which necessitated active ongoing surveillances and consistent prevention and control of E. cloacae.The antimicrobial resistance rates were low among 53  Enterobacter cloacae is an important emerging pathogen, causing various nosocomial infections, including respiratory infections, bloodstream infections (BSIs) and surgical site infections ). Escherichia coli ATCC 25922 was used for quality control.Antimicrobial susceptibility of the  by CLSI . The ant Edition). For tig Edition); for polblaTEM, blaSHV, blaCTX-M , blaOXA , blaVEB, blaGES, and blaPER), carbapenemase genes  and plasmid-mediated colistin resistance gene (mcr-1), using primers previously described [http://blast.ncbi.nlm.nih.gov/BLAST).Polymerase chain reaction (PCR) was performed to confirm the existence of ESBL genes  as described by Miyoshi-Akiyama et al.[http://pubmlst.org/ecloacae/). STs not found in the database were submitted. The eBURST version 3.0 was used to analyze the clustering of related STs. The eBURST algorithm can group strains according to their allelic profiles by employing a user-specified group definition, as well as drawing a rough sketch to show the genetic relationship[A multilocus sequence typing (MLST) scheme was used to assign ma et al.. The comationship. In thisP value of <0.05.SAS 8.2  was used for statistical analysis. Continuous variables were presented as median and interquartile range. The chi-square or Fisher\u2019s exact test was used to compare the disparity between different groups for categorical variables. Differences were considered statistically significant at a two-tailed The age of the 53 patients ranged from 23 to 86 years . Male patients (37/53) were more than females (16/53). Near half of the episodes were derived from the surgery (22/53), and where most patients suffered from malignant tumor (10/22) .E. cloacae isolates, 8(15.1%) were ESBL producers, 3(5.7%) were carbapenemase producers and 18(34.0%) were pAmpC producers (P<0.05). Of note, pAmpC- and non-producers both presented low resistance rates (<40%) to all antibiotics, which did not differ significantly in the two groups (p>0.05) (As summarized in roducers . The hig(p>0.05) .E. cloacae isolates were positive for pAmpC genes, of which 15 (83.3%) isolates were detected with MIR/ACT gene and the other 3 (16.7%) were detected with DHA-1 gene , blaVEB, blaGES, blaPER, blaVIM, blaKPC, blaOXA-48 and blamcr-1 were detected.Of the 8 ESBL producers, SHV enzymes  predominated with all identified as SHV-12, and CTX-M enzymes  followed, including CTX-M-15 and CTX-M-65  . A totalA-1 gene . Of the A-1 gene . Besideshttps://pubmlst.org/bigsdb?db=pubmlst_ecloacae_seqdef). It was indicated in MLST analysis distinguished 51 different STs, clustered into 3 non-overlapping clonal complexes (CCs) and 43 singletons . Only STE. cloacae was frequently implicated in nosocomial infections[E. cloacae isolates[E. cloacae worsening clinical outcome and prolonging hospitalization duration have been alarmingly increasing since 21st century[E. cloacae bloodstream isolates harboring resistant genes were mostly associated with elderly patients from the surgery or Intensive Care Unit  produced ESBL, which was higher than that in north-eastern USA (10.1%)[Enterobacter spp. collected from all sites[E. cloacae [E. cloacae isolates due to various modes of transmission.Of all 15.1% 8/5 producedall sites. Unlike all sites, SHV wasall sites, and simall sites. SHV-12, cloacae . What\u2019s  cloacae , 26, whiE. cloacae was once reported in Taiwan[PAmpC can preferentially hydrolyze all \u03b2-lactams except fourth-generation cephalosporins and carbapenems and may serve as the reservoir for the emergence of antibiotic resistance . The higin Taiwan, but firPseudomonas aeruginosa isolate in Singapore [E. cloacae isolates, but interestingly, the two IMP-producers in our study were both detected as IMP-26. It was still noted that IMP-producing E. cloacae had caused several outbreaks or possible spread in other countries [E. cloacae. NDM-1 was the dominant carbapenemase of carbapenems-resistant E. cloacae in Henan China, and in contrast to VIM-1 the most prevalent in Spain and other southern Europe [Carbapenemase spread has been increasingly reported worldwide over the last decade. We founingapore  and few ountries , 33, 34,n Europe , 36; whiE. cloacae has once denoted the association of distinct clonal groups with genetic lineages of higher prevalence and/or wider geographical spread [E. cloacae bloodstream isolates did not evolve from a unique ancestral background with 51 STs distinguished from 53 isolates and 8 isolates (15.1%) assigned to 3 CCs Click here for additional data file."}
+{"text": "VOLUME 104104(3) July, page 242http://dx.doi.org/10.3163/1536-5050.104.3.014.Bramer WM, Giustini D, de Jonge GB, Holland L, Bekhuis T. De-duplication of database search results for systematic reviews in EndNote. J Med Libr Assoc. 2016 Jul;104(3):240\u20133. DOI: Page 242: Table 1, row C3, should state:Review the top references without page numbers and those with page numbers starting with the number 1 for equivalent author names.Only articles without page numbers or page numbers starting with 1 should be reviewed."}
+{"text": "Correction to:The ISME Journal (2017) 11, 932\u2013944; doi:10.1038/ismej.2016.172Updated online 28 February 2017: This article was originally published under a CC BY-NC-SA 4.0 license, but has now been made available under a CC-BY 4.0 license. The PDF and HTML versions of the paper have been modified accordingly."}
+{"text": "In the original article, there was a mistake in the published supplementary material. The literature search string as mentioned in the section \u201cLiterature Search and Selection Strategy\u201d never appeared in the published material. The search string as applied in Medline appears below. The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way.Ovid Technologies, Inc. Email ServiceDatabase: Epub Ahead of Print, In-Process & Other Non-Indexed Citations, Ovid MEDLINE(R) Daily and Ovid MEDLINE(R)<1946 to Present>Search Strategy:1 Hyperparathyroidism, Primary/dt [Drug Therapy] (99)2 Hyperparathyroidism, Primary/(2211)3 Parathyroid adenom*.mp. (4188)4 (Primary adj2 Hyperparathyr*).mp. (9043)5 2 or 3 or 4 (11256)6 exp Diphosphonates/(22931)7 bisphosphonate*.mp. (14211)8 diphosphonat*.mp. (17321)9 bisphosphonic*.mp. (152)10 clodron*.mp. (2326)11 alendron*.mp. (4714)12 etidron*.mp. (3123)13 ibandron*.mp. (1063)14 incadron*.mp. (78)15 medron*.mp. (4405)16 minodron*.mp. (104)17 neridron*.mp. (94)18 olpadron*.mp. (77)19 pamidron*.mp. (2920)20 risedron*.mp. (1775)21 tiludron*.mp. (156)22 zoledron*.mp. (4145)23 cinacalcet.mp. (1071)24 Cinacalcet Hydrochloride/(735)25 \"amg 073\".mp. (24)26 amg073.mp. (0)27 krn 1493.mp. (3)28 krn1493.mp. (4)29 mimpara.mp. (20)30 parareg.mp. (0)31 regpara.mp. (1)32 sensipar.mp. (30)33 exp Isoflavones/(15928)34 ipriflavone.mp. (268)35 Denosumab/(925)36 denosumab.mp. (1712)37 amg 162.mp. (33)38 amg162.mp. (3)39 amgiva.mp. (0)40 prolia.mp. (32)41 xgeva.mp. (16)42 blosozumab.mp. (14)43 ly 2541546.mp. (0)44 ly2541546.mp. (1)45 romosozumab.mp. (45)46 amg 785.mp. (27)47 amg785.mp. (0)48 cdp 7851.mp. (0)49 cdp7851.mp. (3)50 odanacatib.mp. (156)51 (\"mk 0822\" or mk0822 or mk822 or mk 822).mp. (5)52 etelcalcetide.mp. (5)53 (amg 416 or amg416).mp. (12)54 (kai 4169 or kai4169).mp. (1)55 (ono 5163 or ono5163).mp. (0)56 telcalcetide.mp. (0)57 velcalcetide.mp. (4)58 or/6-57 (49081)59 5 and 58 (459)60 1 or 59 (491)The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "We report here the first complete genome sequences of genotype GI.3, GI.4, GI.6, GI.7, and GII.7 sapovirus strains, detected from fecal samples of acute gastroenteritis patients. Complete or nearly complete genome sequences of all 18 genotypes of human sapoviruses are now available for phylogenetic analysis and primer design. Genetically diverse sapovirus (SaV) strains have been detected in fecal specimens from patients with acute gastroenteritis \u20134. We ha\u20136\u2013de novo assembly of consensus SaV genome sequence were performed first, as described ((AY237420), GII.3 (AY603425), GII.5 (LC190463), and GII.6 (AY646855) SaV genomes in combination with two primers designed in the previously determined VP1-encoding-region sequence  or QIAamp viral RNA minikit (Qiagen). Library preparation from the purified RNA for sequencing on an Illumina MiSeq platform (Illumina) and escribed , 10. Forescribed  using thsequence . FurtherGII.5 LC10463, andsequence 11. The 5GII.5 LC10463, andThe genomes of GI.3 Hu/OH08021/2008/JP, GI.4 Hu/SV/Chiba/000496/2000, GI.6 Hu/SV/Chiba/000764/2000, GI.7 Hu/D1714-B/2008/JPN, and GII.7 Hu/20072248/2008/JP SaV strains consist of 7,442, 7,436, 7,443, 7,452, and 7,462\u2009nt, respectively, excluding the poly(A) tail. All of these SaV genomes were predicted to encode two ORFs, a short 5\u2032 untranslated region (UTR) (12 or 13\u2009nt long) and a 3\u2032-UTR 78 to 112\u2009nt long). The 5\u2032 terminal sequence was conserved as GTG, similarly to those of other SaVs , 11.2\u2009nt longThe new sequence data determined in this study will be useful in designing more broadly reactive primers and probes for human SaV detection PCR, as well as establishment of a nonstructural protein coding region typing scheme like that recently established for norovirus .AB623037, AJ606693, AJ606694, AB522390, and AB630067, respectively.The genome sequences of GI.3 Gu/OH08021/2008/JP, GI.4 Hu/SV/Chiba/000496/2000, GI.6 Hu/SV/Chiba/000764/2000, GI.7 Hu/D1714-B/2008/JPN, and GII.7 Hu/20072248/2008/JP SaV strains have been deposited in GenBank under the accession no."}
+{"text": "Nature Communications8: Article number: 15110; DOI: 10.1038/ncomms15110 (2017); Published: 04272017; Updated: 0613201752\u2019 and \u2018pSANG10-3F-BG4 (Addgene 55756)52 was transformed into\u2026.\u2019 The correct citation is given below.In this Article, the antibody \u2018BG4\u2019 is consistently referred to incorrectly as \u2018hf2\u2019. These errors appear in the Results and Methods. Additionally, the Article incorrectly cites reference 52 in the sentences \u2018A recent study described hf2, an engineered single-chain antibody specific for G4 DNANat. Chem.5, 182\u2013186 (2013).Biffi, G., Tannahill, D., McCafferty, J. & Balasubramanian, S. Quantitative visualization of DNA G-quadruplex structures in human cells."}
+{"text": "The impact of MMP-1 , MMP-3 Lys45Glu (A/G), MMP-7 -181A/G, and MMP-12 -82A/G variants and plasma MMP levels on obesity and microvascular reactivity in Tunisians.  Our population included 202 nonobese and 168 obese subjects. Anthropometric, biochemical, and microvascular parameters were determined according to standard protocols. PCR-RFLP and ELISA were used to determine the genetic variants and levels of MMPs, respectively. 2): 30\u2009\u00b1\u20090.51 versus 27.33\u2009\u00b1\u20090.8, P = 0.004; MMP-3 levels: 7.45 (4.77\u201311.91) versus 5.21 (3.60\u201310.21) ng/ml, P = 0.006). The MMP-12 -82G allele was also associated with higher BMI values when compared to subjects carrying the AA genotype . Individuals carrying the MMP-3 45G or MMP-12 -82G variants were also associated with a higher risk for severe forms of obesity . Similarly, the MMP-7 -181G allele was associated with a higher MMP-7 level and an increased risk for morbid obesity when compared to AA genotype carriers (0.32 (0.31\u20130.60) versus 0.18 (0.17\u20130.24) ng/ml, P = 0.01; OR\u2009=\u20091.67, P = 0.02, resp.).  The MMP-3 45Glu (G) allele associates with higher anthropometric values and MMP-3 levels compared to AA genotype carriers (BMI (kg/m MMP-3, MMP-7, and MMP-12 polymorphisms associate with obesity risk and its severity. Obesity is a pathological condition that is closely related to genes with environmental modulators, including sedentary life and positive energy balance . It is cUntil now, SNPs in MMP genes have been described to alter their expression . As an eUp to now, only a few studies have determined the impact of MMP variants and plasma MMP levels on obesity-related phenotypes and microvascular reactivity. The goals of the current report were to explore the impact of MMP-1 , MMP-3 Lys45Glu (A/G), MMP-7 -181A/G, and MMP-12 -82A/G SNPs on the development of obesity and its severity among Tunisians. We also determined whether these SNPs affect biochemical and microvascular reactivity, as well as plasma levels of the corresponding proteins.2) while 168 subjects were obese (BMI\u2009\u2265\u200930\u2009kg/m2). Obese individuals were categorized according to the WHO classification as class I with 30\u2009\u2264\u2009BMI\u2009<\u200935\u2009kg/m2, class II with 35\u2009\u2264\u2009BMI\u2009<\u200940\u2009kg/m2, and class III (morbidly obese) with 40\u2009kg/m2\u2009\u2264\u2009BMI [2 were excluded from the study. Diabetic individuals and subjects with a history of cardiovascular disease, smoking habits, malignancy, and renal, thyroid, and liver diseases were also excluded from the study. Other exclusion criteria were subjects using medications that might alter the endothelial or smooth muscle-dependent responses. All participants signed a written informed consent, and the study was approved by the Farhat Hached Hospital Ethics Committee.370 randomly selected Tunisian subjects (181 men and 189 women) with a mean age of 38 years were selected for the current study. 202 individuals were classified as nonobese subjects (18.5\u2009\u2264\u2009BMI\u2009<\u200925\u2009kg/mm2\u2009\u2264\u2009BMI . Overwei2), waist circumference (WC) (cm), and hip circumference (HC) (cm) were determined according to standard protocols. Waist-to-height ratio (WHtR) and waist-to-hip ratio (WHR) were also measured by WC/height ratio and WC/HC ratio, respectively. The fasting glucose (FG), total cholesterol (TC), and triglycerides (TG) were determined by the corresponding oxidase methods . The immune-inhibition method  was used to measure high-density lipoprotein cholesterol concentration (HDL-C) while LDL-C was determined with the Friedwald formula [For all participants, weight (kg), height (m), BMI (kg/m formula . The senmax) represented the endothelium-dependent response, registered after three cumulative doses of acetylcholine (ACh) (arbitrary unit). The forearm flow response to heating the skin (FSBFRHS) represented the endothelium-independent response registered after the increase in skin temperature (44\u00b0C) without ACh infusion (arbitrary unit). Data were also presented as cutaneous vascular conductance (CVC) that represents the cutaneous blood flow (CBF)/mean arterial pressure (MAP) ratio. The MAP was determined as 1/3 SBP\u2009+\u20092/3 DBP. The difference between the peak CVC upon ACh stimulation (after three doses of ACh) and the baseline CVC was considered as the endothelium-dependent response (\u0394ACh-CVC). The difference between the peak CVC following RHS-induced vasodilation and the baseline was considered as the endothelium-independent response (\u0394RHS-CVC) [Endothelial function was determined in all subjects by assessing the forearm microvascular cutaneous vasoreactivity using laser Doppler flowmetry coupled with iontophoresis  . The maxRHS-CVC) .\u03bcl) contained 100\u2009ng of genomic DNA, 25\u00a0\u03bcM of each deoxynucleotide triphosphate , 50\u2009pmol of each primer, 2.5\u2009mM of MgCl2, and 0.5 units of Taq polymerase . Details of primers and PCR-RFLP conditions are reported in DNA was isolated as previously described . AllelicMMP-1 and MMP-3 levels were determined by enzyme-linked immunosorbent assays (ELISA) using commercially available kits . MMP-7 and MMP-12 plasma levels were evaluated by Magnetic Luminex .t-test or the Mann\u2013Whitney U test as appropriate. Comparison of categorical variables including calculation for deviation from the Hardy\u2013Weinberg equilibrium (HWE) was performed using the chi-square (\u03c72) test. The evaluation of odds ratios (ORs) and 95% confidence intervals (CIs) was carried out using an unconditional logistic regression model adjusted for age and sex. Haplotypes and linkage disequilibrium (LD) were analyzed with SNPstats (http://bioinfo.iconcologia.net/SNPstats). Results were considered statistically significant if the P value <0.05. The Bonferroni correction (Pc) was applied to adjust for multiple comparisons.SPSS\u00ae 17.0 software  and GraphPad Prism (version 6.04) were used for statistical analyses. Comparisons of quantitative variables were carried out using a 2\u2009\u00b1\u20090.44. In this study, we used a BMI cutoff point of 30\u2009kg/m2 for obesity status. As expected, higher BMI and abdominal obesity values  were associated with obesity status. SBP, DBP, MAP, and hs-CRP were increased in obese subjects. As indicated in max as well as the FSBF dose-response to ACh expressed as area under the curve (AUC) was decreased in the obese group (P < 0.001). Basal CVC, as well as peak ACh-CVC and \u0394ACh-CVC values, was also decreased in obese subjects (P < 0.001). BMI and central adiposity (WC and WHR) were inversely correlated with the FSBFmax response to ACh . Additionally, basal CVC and peak ACh-CVC were inversely associated with BMI  and WC values . The abdominal obesity marker (WHtR) was inversely associated with peak ACh-CVC .Anthropometric, biochemical, and microvascular values are indicated in 2\u2009\u00b1\u20090.61 versus 28.5\u2009kg/m2\u2009\u00b1\u20090.63, For the MMP-1 (-519A/G) polymorphism, no significant variation in BMI values was observed in individuals carrying the dominant model (AG+GG) genotype in comparison to AA genotype carriers ; WC ; WHtR , 2\u2009\u00b1\u20090.49 versus 27.19\u2009kg/m2\u2009\u00b1\u20090.49, P = 0.04) ; WC ; WHtR , Regarding the MMP-3 Lys45Glu (A/G) SNP, individuals carrying the (AG/GG) genotype showed increased BMI, WC, and WHtR values when compared to AA genotype carriers (BMI (30\u2009kg/m = 0.04) ). HigherP = 0.06). No significant correlation was observed between MMP-1, MMP-3, MMP-7, and MMP-12 polymorphisms, FG, and hs-CRP levels. No significant association was also found between MMP-1 and MMP-7 polymorphisms and TG, TC, LDL-C, and HDL-C levels. Regarding the MMP-3 polymorphism, a higher amount of LDL-C was found in subjects carrying the AG or GG genotypes compared to AA genotype carriers . A significant increase in LDL-C was also found among obese subjects carrying the AG or GG genotypes when compared to AA genotype carriers . Investigation of the MMP-12 (-82A/G) SNP also revealed that individuals carrying the AG or GG genotypes present higher LDL-C levels when compared to AA genotype carriers . Analysis of endothelial function in the sampled population revealed, however, no significant relationships between the MMP-1, MMP-7, MMP-3, and MMP-12 variants and microvascular reactivity parameters (data not shown).The SBP, DBP, and MAP revealed no significant association with MMP-1, MMP-3, and MMP-7 polymorphisms. Regarding the MMP-12 (-82A/G) SNP, there was an increase in DBP among subjects carrying the AG or GG genotypes compared to AA genotype carriers . When subjects were categorized according to severity of obesity, a higher level of MMP-7 was observed in morbidly obese subjects carrying the -181G variant (AG+GG: 0.32 (0.31\u20130.6), AA: 0.18 (0.17\u20130.24), P = 0.01, Pc = 0.04).We also determined whether the MMP polymorphisms affect plasma levels of the corresponding proteins. For the MMP-1  SNPs, there was no significant impact on MMP-1 levels in the sampled population  ng/ml, AG+GG: 2 (0.75\u20134.16) ng/ml; \u20131607 1G/2G), 1G1G: 2.53 (0.75\u20134.17) ng/ml, 1G2G+2G2G: 1.66 (0.66\u20133.81) ng/ml). For the MMP-3 Lys45Glu (A/G) polymorphism, subjects carrying the AG or GG genotypes present higher MMP-3 levels compared to individuals having the AA genotype (7.45 (4.77\u201311.91) ng/ml versus 5.21 (3.6\u201310.21) ng/ml, P = 0.03, Pc = 0.24; OR\u2009=\u20091.61, P = 0.02, Pc = 0.08, resp., P = 0.01; OR\u2009=\u20092.4, P = 0.006; OR\u2009=\u20092.04, P = 0.004, resp., P = 0.004, Pc = 0.016). When subjects were categorized according to severity of obesity, the frequency of the Lys45Glu (G) allele was more frequent in nonmorbidly obese (48.58%) compared to nonobese subjects  . For the MMP-12 (-82A/G) SNP, the frequencies of AA, AG, and GG genotypes were 76.4%, 23%, and 0.6% in obese participants and 88.6%, 11.4%, and 0% in nonobese participants. We found a significant association between the AG genotype, the combined genotypes AG+GG, and the development of obesity . The frequencies of A and G alleles were 94.28% and 5.72% in nonobese subjects and 87.88% and 12.12% in obese individuals. The G allele was associated with enhanced obesity when compared to A allele carriers . The G allele was also associated with nonmorbid obesity when compared to the A allele . Similarly, the AG genotype alone or in combination with the GG variant was associated with a higher risk for severe forms of obesity when compared to AA genotype carriers . The G allele was also significantly associated with morbid obesity when compared to the A allele .Analyses were carried out to study the distribution of the different SNPs according to obesity status and severity based on BMI Tables  and 4. F, resp.) . BesidesD\u2032 = 0.42, r = 0.13). Haplotypes follow this order: MMP-1 (-1607 1G/2G), MMP-1 (-519A/G), MMP-7 (-181A/G), MMP-3 Lys45Glu (A/G), MMP-12 (-82A/G). Comparison of the haplotype frequencies between obese and nonobese groups revealed an increased risk for obesity for the haplotype s1G\u22121607:G\u2212519:G\u2212181:GLys45Glu:A\u221282 (Supplementary File 1 available online at https://doi.org/10.1155/2017/6198526).We further determined the haplotype frequencies of the different polymorphisms in obese and nonobese subjects. Linkage disequilibrium analysis revealed that MMP-3 and MMP-12 SNPs were in mild linkage disequilibrium . Several studies reported an enhanced expression of MMP-1 in preadipocytes from obese subjects suggesting a role in ECM remodeling in obesity . MMP-1 hMMP-3 (stromelysin I) is a proteoglycanase and a key player in ECM degradation and remodeling . MMP-3 wMMP-7 (matrilysin-1) is another metalloproteinase with an inflammatory regulatory role that displays a proteolytic activity against various kinds of ECM components . Previouin vitro adipogenesis decreases adipocyte expansion by increasing endostatin levels and inhibiting adipose tissue vascularity. Up to now, several SNPs were reported in the MMP-12 locus. Among these, the -82A/G SNP was found to affect both AP-1 binding affinity and MMP-12 promoter activity [MMP-12 is a macrophage metalloelastase that has a role in ECM remodeling processes . A previactivity . A reporactivity  revealedactivity  revealedactivity , 54. WitIn the current report, when individuals were stratified by BMI, the MMP-3 45Glu (G) and MMP7 (-181G) variants were more frequent in individuals with severe forms of obesity. Although further studies are required, these results may highlight that the contribution of MMP-3 and MMP-7 SNPs in the development of the morbid phenotype can be substantially higher than in lower classes of BMI and could, therefore, be valuable markers to predict the risk for the development of extreme classes of obesity. Consistently, previous studies provided evidence of a weak population-related risk for common obesity for several genetic variants. As an example, Villalobos-Compar\u00e1n et al.  found thThe limitations of this study should be considered. A first limitation is the small sample size of the screened population. Our findings need, therefore, to be replicated in a larger population or an independent severe obesity cohort. A second limitation is the lack of measures of MMP activities in serum, as well as in adipose tissue.MMP-3 Lys45Glu (A/G), MMP-7 (-181A/G), and MMP-12 (-82A/G) variants were associated with obesity and its anthropometric indicators. The current findings also highlight a significant correlation between the MMP-3Lys45Glu (A/G) and MMP-7 (-181A/G) variants and severe forms of obesity among Tunisians. Our results also revealed significant alterations in microvascular reactivity in obese subjects. MMP variants may not, however, be determinant factors of endothelial dysfunction in the studied population.Supplementary file 1. Estimated haplotype frequencies of MMP-1, MMP-3, MMP-7 and MMP-12 polymorphisms in obese and non-obese subjects."}
+{"text": "Scientific Reports7:1110; doi:10.1038/s41598-017-01016-8; Article published online 24 April 2017The original version of this Article contained errors. In the Abstract,\u201cThe use of SC79, an Akt1/2 inhibitor\u201dnow reads:\u201cThe use of SC79, an Akt1/2 activator\u201dIn the Introduction,\u201cwildlife and animal exposure including fetal growth\u201dnow reads:\u201cwildlife and animal exposure including reduced fetal growth\u201dIn addition,\u201cinducing Sertoli cell injury and Leydig cell steroidogenic function\u201dnow reads:\u201cinducing Sertoli cell injury and disrupting Leydig cell steroidogenic function\u201dFinally,\u201cThis mechanistic study thus provides additional insight on the molecular mechanism by which PFOS mediates reproductive\u201dnow reads:\u201cThis mechanistic study thus provides additional insight on the molecular mechanism by which PFOS causes reproductive\u201dThe original version of this Article omitted Xiang Xiao and Wing-yee Lui as corresponding authors. Correspondence and request for materials should also be addressed to X.X. (xxiao@zjams.com) or W.Y.L. (wylui@hku.hk)In the Results section, under the sub-heading \u2018PFOS perturbs Sertoli cell TJ-barrier function through changes in the spatial expression of actin binding and regulatory proteins\u2019,\u201cPalladin also no longer stretched across the Sertoli cell cytosol to organize actin filaments into bundles, instead, paladin retracted from cell cytosol and found closer to the cell nuclei .\u201dnow reads:\u201cPalladin also no longer stretched across the Sertoli cell cytosol to organize actin filaments into bundles as seen in controls, instead, palladin retracted from cell cytosol and found closer to the cell nuclei following PFOS treatment \u201dThe Results section contains an error in a sub-heading, where:\u201cRescue of PFOS-induced Sertoli cell TJ-barrier disruption by SC79, an activator of Akt through changes in the re-distribution of BTB-associated proteins.\u201dnow reads:\u201cRescue of PFOS-induced Sertoli cell TJ-barrier disruption by SC79, an activator of Akts through changes in the re-distribution of BTB-associated proteins.\u201dThere were also errors in the Figure legends. In Figure 2 legend,\u201cInstead, palladin was retraced from the cell peripheries but concentrated to cell nuclei after PFOS treatment.\u201dnow reads:\u201cInstead, palladin was retracted from the cell peripheries but concentrated to cell nuclei after PFOS treatment.\u201dIn Figure 3(c) legend6 cells/cm2 for 3 days were pre-treated with 2\u2009\u03bcg/ml SC79 for 30\u2009min, and then cells were rinsed and treated with 20\u2009\u03bcM PFOS for 24 hr. Thereafter, cells were then fixed and examined by immunofluorescence microscopy. SC79 treatment was found to block PFOSinduced mis-localization of TJ proteins CAR and ZO-1, as well as basal ES proteins N-cadherin and \u03b2-catenin, apparently through enhanced endocytosis. BTB-associated proteins at the Sertoli cell-cell interface in control and treated cells were annotated by corresponding white and yellow brackets, respectively\u201d\u201c(C) Sertoli cells cultured at 0.04\u2009\u00d7\u200910now reads:6 cells/cm2 for 3 days were pre-treated with 2\u2009\u03bcg/ml SC79 for 30\u2009min, and then cells were rinsed and treated with 20\u2009\u03bcM PFOS for 24 hr. Thereafter, cells were then fixed and examined by immunofluorescence microscopy. SC79 treatment was found to block PFOS-induced mis-localization of TJ proteins CAR and ZO-1, as well as basal ES proteins N-cadherin and \u03b2-catenin. BTB-associated proteins at the Sertoli cell-cell interface in control and PFOS-treated cells were annotated by corresponding white and yellow brackets, respectively\u201d\u201c(C) Sertoli cells cultured at 0.04\u2009\u00d7\u200910In Figure 4 legend\u201cyellow rectangles in FOS-treated cells\u201dnow reads:\u201cyellow rectangles in PFOS-treated cells\u201dIn Figure 5 legend, Figure 1A now reads Figure 1BIn Figure 6 legend,\u201cThereafter, cells were fixed with ice-cold methanol and visualized byimmunofluorescence microscopy.\u201dnow reads:\u201cThereafter, cells were fixed with ice-cold methanol and visualized by immunofluorescence microscopy using corresponding antibodies (Table 1)\u201dIn addition, the Acknowledgments section now reads:We wish to thank Dr. Ben Margolis and Dr. Shuling Fan who kindly provided us with some anti-CRB3 antibody to validate some experiments in this report. This work was supported by grants from the National Institutes of Health ; National Natural Science Foundation of China (NSFC) (31371176 to X.X.), Qianjiang Talents Program (QJD1502029 to X.X.), Zhejiang Province Funding ; Hong Kong Research Grants Council (RGC) General Research Fund (GRF) , NSFC/RGC Joint Research Scheme (N_HKU 717/12 to W.M.L.), and University of Hong Kong CRCG Seed Funding (to W.M.L.)The Author Contributions statement now reads:C.Y.C., Y.G. and H.C. designed research; Y.G., H.C. and C.Y.C. performed research; X.X., W.Y.L., W.M.L., D.D.M. and C.Y.C. contributed new reagents/analytic tools; Y.G., H.C. and C.Y.C. analyzed data; Y.G. and C.Y.C. wrote the paper; Y.G., H.C. and C.Y.C. prepared all figures. All authors reviewed the manuscript.There were also errors in References 43 and 76, which now read:in vitro a useful model to study molecular mechanisms in spermatogenesis? Semin Cell Dev Biol59, 141\u2013156, doi:10.1016/j.semcdb.2016.01.003 (2016).43. Li, N., Mruk, D. D., Lee, W. M., Wong, C. K. C. & Cheng, C. Y. Is toxicant-induced Sertoli cell injury et al. SC79 protects retinal pigment epithelium cells from UV radiation via activating Akt-Nrf2 signaling. Oncotarget7, 60123\u201360132, doi:10.18632/oncotarget.11164 (2016).76. Gong, Y. These errors have now been corrected in the HTML and PDF versions of this Article."}
+{"text": "Bariatric surgery has emerged as a competitive strategy for obese patients. However, its comparative efficacy against non-surgical treatments remains ill-defined, especially among nonseverely obese crowds. Therefore, we implemented a systematic review and meta-analysis in order for an academic addition to current literatures.Literatures were retrieved from databases of PubMed, Web of Science, EMBASE and Cochrane Library. Randomized trials comparing surgical with non-surgical therapies for obesity were included. A Revised Jadad's Scale and Risk of Bias Summary were employed for methodological assessment. Subgroups analysis, sensitivity analysis and publication bias assessment were respectively performed in order to find out the source of heterogeneity, detect the outcome stability and potential publication bias.P > 0.05). The pooled results of primary endpoints (weight loss and diabetic remission) revealed a significant advantage among surgical patients rather than those receiving non-surgical treatments (P < 0.05). Furthermore, except for certain cardiovascular indicators, bariatric surgery was superior to conventional arms in terms of metabolic secondary parameters (P < 0.05). Additionally, the pooled outcomes were confirmed to be stable by sensitivity analysis. Although Egger's test (P < 0.01) and Begg's test (P<0.05) had reported the presence of publication bias among included studies, \u201cTrim-and-Fill\u201d method verified that the pooled outcomes remained stable.25 randomized trials were eligibly included, totally comprising of 1194 participants. Both groups displayed well comparability concerning baseline parameters (Bariatric surgery is a better therapeutic option for weight loss, irrespective of follow-up duration, surgical techniques and obesity levels. Emerging as a costly burden of global healthcare system, obesity has currently attracted worldwide attentions due to its uncontrollably rising incidence, especially in industrialized countries . EconomiAccording to SAGES  and NICEBariatric surgery, initially reported in 1995 , has beeversus non-surgical interventions is rarely described, especially lacking of a well summarized evidence. Additionally, the clinical value of bariatric surgery for nonseverely obese patients requires further analysis. Hence we performed this systematic review and meta-analysis in order to comprehensively make comparisons between both strategies, aiming to provide novel evidences for future guidelines.Nevertheless, long-term (3-year or more) efficacy of surgical The preliminary 1076 entries were rigorously screened to generate 25 eligible studies for pooling analysis, with a total amount of 1194 participants and individually ranging from 16 to 150 Figure . Merely According to Revised Jadad's Scale, 13 trials were appraised as high-quality in methodology, while DSS, Heindorff 1997 and Mingrone 2002 were identified as low-quality investigations  , 2-year (P < 0.00001) or long-term (3 years or more) follow-up duration (P < 0.00001) , Roux-en-Y gastric bypass (P < 0.00001), laparoscopic adjustable gastric banding (P < 0.00001) and biliopancreatic diversion (P < 0.00001)  and severe obesity , metabolic surgery was a more effective tool for losing weight against non-surgical treatments   .P < 0.0001), 2-year (P = 0.004) and long-term follow-up (P < 0.0001)  .P < 0.00001), Roux-en-Y gastric bypass (P < 0.00001), laparoscopic adjustable gastric banding (P = 0.0008) and biliopancreatic diversion (P = 0.001)  .P = 0.0004) and severely obese (P < 0.0001) sufferers significantly benefited from surgical remedies, compared with non-surgical patients   .P < 0.00001), 2-year (P < 0.00001) or long-range follow-up (P = 0.006), bariatric surgery was far more efficient to eliminate excessive weight than non-surgical interventions (Regardless of 1-year (ventions .P < 0.00001), Roux-en-Y gastric bypass (P < 0.00001), laparoscopic adjustable gastric banding (P = 0.0008) and biliopancreatic diversion (P < 0.00001) was performed  and severe obesity (P < 0.00001)  .P < 0.00001)  .P = 0.0005), 2-year (P = 0.0003) or long-term follow-up (P = 0.02), surgical management was significantly effective in decreasing fasting glucose compared to non-surgical interventions (Regardless of 1-year (ventions .P = 0.0005), Roux-en-Y gastric bypass (P < 0.00001) and laparoscopic adjustable gastric banding (P < 0.0001) fell behind in fasting glucose reduction against conserved therapeutics, except for biliopancreatic diversion, which was statistically equivalent to contrastive group (P = 0.05) (None of sleeve gastrectomy ( = 0.05) .P = 0.001) and severely obese (P < 0.00001) patients obtained greater downregulation on fasting glucose following the surgical interventions   .P < 0.00001), 2-year (P = 0.02) and long-term follow-up (P < 0.00001), rather than those with conservative interventions , Roux-en-Y gastric bypass (P < 0.00001), laparoscopic adjustable gastric banding (P = 0.002) and biliopancreatic diversion (P = 0.03), bariatric surgery was significantly superior to non-surgical approaches in terms of reducing elevated glycated hemoglobin (Irrespective of sleeve gastrectomy (moglobin .P < 0.00001) and severely obese patients (P < 0.00001)  .P < 0.00001)  .P < 0.00001), 2-year (P < 0.00001) and long-term follow-up (P < 0.00001), patients that were surgically treated obtained larger decline on waist circumference than those were conventionally cured (Among the three subgroups of 1-year (ly cured .P < 0.0001), Roux-en-Y gastric bypass (P < 0.00001), laparoscopic adjustable gastric banding (P < 0.00001) and biliopancreatic diversion (P < 0.00001)  .P < 0.00001) and severely obese sufferers (P < 0.00001), more reduction on waist circumference was observed following surgical management in contrast to traditional treatments (Based on the pooled outcomes featuring nonseverely (eatments .P < 0.00001)  .P = 0.30), while bariatric surgery was a superior option among sufferers with 1-year (P = 0.001) and long-range follow-up (P = 0.02)  .P = 0.36), laparoscopic adjustable gastric banding (P = 0.36) and biliopancreatic diversion (P = 0.90) was statistically comparable with non-surgical strategies in terms of reduction on systolic pressure, except for the dominant efficacy of Roux-en-Y gastric bypass (P = 0.007) (Bariatric surgery featuring sleeve gastrectomy (= 0.007) .P < 0.00001) and severely obese patients (P = 0.41) respectively, in terms of reduction on systolic pressure   .P = 0.26) and 2-year (P = 0.55). However, surgical interventions were inferior to non-invasive remedies amid patients with long-term follow-up (P = 0.03)  .P = 0.03) were in a significantly superior status, the remaining techniques were statistically comparable to conservative regimens concerning the decrease on diastolic pressure, inclusive of sleeve gastrectomy (P = 0.67), laparoscopic adjustable gastric banding (P = 0.57) and biliopancreatic diversion (P = 0.52) (Although Roux-en-Y gastric bypass ( = 0.52) .P = 0.23) and severe obesity (P = 0.73) (Patients undergoing both interventions exhibited comparable efficacy on diastolic pressure reduction regardless of nonsevere obesity ( = 0.73) .P < 0.00001)  .P < 0.0001) and 2-year (P < 0.0001) follow-up had a greater decline on triglycerides level than those being conventionally treated, except for the comparable efficacy among patients with long-term follow-up (P = 0.06)  .P = 0.0005) and laparoscopic adjustable gastric banding (P = 0.004) exerted a more evident influence among enrolled patients, while sleeve gastrectomy (P = 0.08) and biliopancreatic diversion (P = 0.14) was therapeutically comparable with conventional remedies  and severe obesity (P < 0.0001)  .P = 0.13)  .P = 0.18) and 2-year (P = 0.07), there was no therapeutic difference between both interventions. However, patients with long-term follow-up achieved more reduction on total cholesterol following non-surgical managements, in contrast to bariatric surgery (P = 0.03) (Among obese sufferers being followed up for 1-year ( = 0.03) .P = 0.004), patients receiving bariatric surgery gained comparable efficacy of total cholesterol reduction against non-surgical enrollees, including sleeve gastrectomy (P = 0.67), Roux-en-Y gastric bypass (P = 0.78) and laparoscopic adjustable gastric banding (P = 0.62) (Besides the preponderant role of biliopancreatic diversion ( = 0.62) .P = 0.01), while nonseverely obese counterparts reported no significant difference between surgical and non-surgical interventions (P = 0.76)  .P < 0.00001)  .P < 0.00001), 2-year (P = 0.0005) and long-term follow-up (P = 0.001), metabolic surgery led to more increase on high density lipoprotein among included patients, compared to non-surgical recipients (Despite of 1-year (cipients .P = 0.003) and Roux-en-Y gastric bypass (P < 0.00001) displayed a higher increase on high density lipoprotein. However, the increase on high density lipoprotein was identically obtained between non-surgical treatments and bariatric surgery of laparoscopic adjustable gastric banding (P = 0.19) and biliopancreatic diversion (P = 0.27) (The recipients of sleeve gastrectomy ( = 0.27) .P < 0.00001) or severe obesity (P < 0.00001), bariatric surgery was a more capable pattern of elevating high density lipoprotein than traditional modes   .P = 0.24) and 2-year follow-up (P = 0.14), recipients of conventional strategies exceled those with long-term postoperative follow-up duration, with regard to the reduction on low density lipoprotein (P < 0.00001)  .P = 0.34), Roux-en-Y gastric bypass (P = 0.71) and laparoscopic adjustable gastric banding (P = 0.46), it was mathematically comparable between surgical and non-surgical patients regarding the decrease on low density lipoprotein. Nevertheless, patients undergoing biliopancreatic diversion had an advantage over those were conventionally cured (P < 0.00001) (Based on subgroup statistics of sleeve gastrectomy (0.00001) .P = 0.76) and severely obese patients (P = 0.09)  .Firstly, by interchanging random-effects and fixed-effects models, the overall as well as subgroup outcomes of primary endpoints (weight loss and diabetic remission) were confirmed to be statistically stable Table .Secondly, by eliminating four low-quality trials of DSS, Heindorff 1997, Mingrone 2002 along with O'Brien 2013, the stability of primary endpoints were numerically verified, irrespective of overall or subgroup analysis  and 6 added for diabetic remission   and diabetic remission , Egger's test and Begg's test consistently confirmed the presence of publication bias among the included studies . However 0.0001) .Although Golomb et al had retrospectively doubted the dominant role of bariatric surgery at 5-year follow-up , its conIt is empirically regarded that compared to severe obesity, patients suffering from nonsevere obesity manifest with better general conditions which makes it more economical to be conventionally cured. However, deriving from our pooled evidence, nonseverely obese patients should rather receive bariatric surgery than conservative managements, which is similar to those featuring morbid obesity and is a vital addition to current literatures.Adolescent obesity and overweight crowds have been targeted as further research highlights of bariatric surgery. Uncontrollable weight increase has severer impacts on pubescents than adults, simultaneously damaging their sexual development and intelligence growth . O'BrienNotwithstanding the statistical outcomes are of great strength in both methodology and statistics, there are still some limitations within. First of all, the internal heterogeneity is still in a considerable level despite the subgroup analyses have been additionally performed. Difference from chemical examination methods, surgical manipulation techniques and diversified non-surgical interventions (life-style and medications) may contribute to inerasably internal heterogeneity. More accurate grouping on different confounding factors may benefit the source seeking of heterogeneity. Secondly, raw data of life-quality and survival prognostication are inadequate for meta-analyzing, making it less valuable in terms of comparisons on life expectancy. Moreover, mutual comparison of financial burdens is still lacking, blockading a more comprehensive appraisal of both strategies.Taken together, despite of the shortcomings of surgical therapeutics  , bariatrIn line with the PRISMA Checklist  and CochIn order to guarantee the integrity of literature retrieval, databases of PubMed, Web of Science, EMBASE and Cochrane Library were electronically searched with a search term \u201cbariatric randomized OR bariatric randomised OR obesity surgery\u201d. Both abstracts and full-texts were elaborately examined for fear of omission or ineligible inclusion. Additionally, citation lists of formerly published meta-analysis were screened as well.versus non-surgical interventions among obese patients (body mass index > 30); 3. Adequate raw data of interested endpoints including metabolic and cardiovascular indicators;Studies were eligibly included because of the following criteria: 1. English-written and officially published trials until December 2015; 2. Randomized controlled trials evaluating the comparative efficacy of surgical Studies were eventually eliminated due to the following reasons: 1. Duplicated publications; 2. Inadequate sample-size (< 10); 3. Lack of follow-up materials;With regard to baseline, primary  and secondary parameters , a well-prepared electronic form was designed to facilitate date extraction from tables, figures, text contents and supplementary information within the eligible trials. Overlapped data deriving from a single registered trial was mathematically combined to prevent repetitive counting. All continuous variables were rounded to one decimal place.Firstly, a Revised Jadad's Scale  was emplSecondly, as an addition to Revised Jadad's Scale, Review Manager 5.3 assisted us to summarize the risk of bias amid the qualified literatures. The symbol of green, yellow and red represented low risk of bias, unclear risk of bias and high risk of bias respectively, in terms of seven constituted categories . The higher proportion that green occupies, the lower risk of bias there is .2) [2 < 25%. Otherwise a random-effects model was preferred under the remaining circumstances [via eliminating low-quality trials, exclusion of controversial studies and comparing the outcome variation between fixed-effects and random-effects models. Moreover, the incorporated outcomes were additionally classified into various subgroups for purpose of more specific and instructive discoveries. With aid of STATA 12.0, publication bias was numerically examined by Begg's test and Egger's test [P < 0.05.Review Manager 5.3 was employed as a statistical platform for our quantitative analysis. The effect-sizes of dichotomous and continuous variable were calculated by models of odds ratio and weighted mean difference respectively, along with 95% confidence interval. If the source data on endpoints were not offered explicitly, median was statistically regarded as mean while standard deviation was derived from range, interquartile range or 95% confidence interval as appropriate , 49. A d2) . Revealimstances , in order's test . A \u201cTrim"}
+{"text": "SmC5-MTase genes were identified from the genome of Salvia miltiorrhiza, a well-known traditional Chinese medicine material and an emerging model medicinal plant. Based on conserved domains and phylogenetic analysis, eight SmC5-MTase genes were divided into four subfamilies, including MET, CMT, DRM and DNMT2. Genome-wide comparative analysis of the C5-MTase gene family in S. miltiorrhiza and Arabidopsis thaliana, including gene structure, sequence features, sequence alignment and conserved motifs, was carried out. The results showed conservation and divergence of the members of each subfamily in plants. The length of SmC5-MTase open reading frames ranges widely from 1,152 (SmDNMT2) to 5,034\u00a0bp (SmMET1). The intron number of SmC5-MTases varies between 7 (SmDRM1) and 20 (SmCMT1 and SmCMT2b). These features were similar to their counterparts from Arabidopsis. Sequence alignment and conserved motif analysis showed the existence of highly conserved and subfamily-specific motifs in the C5-MTases analyzed. Differential transcript abundance was detected for SmC5-MTases, implying genome-wide variance of DNA methylation in different organs and tissues. Transcriptome-wide analysis showed that the transcript levels of all SmC5-MTase genes was slightly changed under yeast extract and methyl jasmonate treatments. Six SmC5-MTases, including SmMET1, SmCMT1, SmCMT2a, SmCMT2b, SmCMT3 and SmDRM1, were salicylic acid-responsive, suggesting the involvement of SmC5-MTases in salicylic acid-dependent immunity. These results provide useful information for demonstrating the role of DNA methylation in bioactive compound biosynthesis and Dao-di herb formation in medicinal plants.Cytosine DNA methylation is highly conserved epigenetic modification involved in a wide range of biological processes in eukaryotes. It was established and maintained by cytosine-5 DNA methyltransferases (C5-MTases) in plants. Through genome-wide identification, eight putative  Cytosine DNA methylation, a dominating epigenetic modification mechanism, plays vital roles in gene expression regulation, transposon silencing, gene imprinting, and X chromosome inactivation . It is wde novo through distinct mechanisms (De novo methylation is mainly established by RNA-directed DNA methylation (RdDM), in which domains-rearranged methyltransferases (DRMs) are guided to target locus to direct methylation of all three sequence contexts via 24-nucleotide short interfering RNAs , another CMT subfamily member . Arabidopsis AtC5-MTase proteins were blast-analyzed against the S.\u00a0miltiorrhiza 99-3 whole genome sequence using tBLASTn algorithm  using the BLASTx and BLASTn program, respectively (http://www.ncbi.nlm.nih.gov/blast/). Obtained raw genomic sequences, open reading frame (ORF) sequences and deduced protein sequences were listed in Sequences of 11 lgorithm . S.\u00a0milthttp://gsds.cbi.pku.edu.cn/index.php) using the coding sequences and the corresponding genomic sequences as inputs. Protein amino acid number, molecular weight (Mw) and theoretical isoelectric point (pI) were analyzed using the EXPASY PROTOPARAM tool (http://www.expasy.org/tools/protparam.html). Multiple sequence alignment was carried out using ClustalW  plants grown in a field nursery at the Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences, were collected in August and stored in liquid nitrogen until use. Total RNA was extracted from three biological replicates for each tissue using the Quick RNA isolation kit . Each biological replicate represents independent single plant. RNA integrity and quantity were analyzed using agarose gel and NanoDrop 2000C Spectrophotometer , respectively. Total RNA was reverse-transcribed using the PrimeScript\u2122\u00a0 RT reagent kit . qRT-PCR primers listed in http://frodo.wi.mit.edu/primer3/). qRT-PCR analysis was conducted in triplicate on the CFX96\u2122 real-time PCR detection system  for each biological replicate using SYBR premix Ex Taq\u2122\u00a0kit  as described  (S. miltiorrhiza hairy roots treated with yeast extract (YE) and methyl jasmonate (MeJA) were downloaded from SRA database under the accession number SRP111399  were downloaded from SRA database under the accession number SRX1423774  . RNA-seqRP111399 . RNA-seqX1423774 . DiffereX1423774 . Heat maX1423774 .Arabidopsis cytosine DNA methyltransferase proteins against the whole S.\u00a0miltiorrhiza 99-3 genome sequence resulted in the identification of eight SmC5-MTase genes. The number of SmC5-MTase genes in S.\u00a0miltiorrhiza is comparable with that in other plants, such as Arabidopsis  varies from 4.89 (SmCMT3) to 8.03 (SmCMT2a) and the molecular weight (Mw) varies from 43.7 kDa (SmDNMT2) to 189.4 kDa (SmMET1)  . These sterparts . Gene stse genes . The undS. miltiorrhiza and Arabidopsis. Four highly conserved motifs, including I, IV, VI and X  and methyl jasmonate (MeJA) are effective elicitors for tanshinone and phenolic acid production in iorrhiza . The resry roots . The trary roots . CompareSmC5-MTases in plant defense, RNA-seq data from SA-treated S.\u00a0miltiorrhiza cell cultures was analyzed -dependent immunity . In ordeanalyzed . The trareatment , suggestS. miltiorrhiza. Understanding the regulatory mechanism of their biosynthesis and metabolism is important for S. miltiorrhiza quality improvement. Most of the studies about S. miltiorrhiza gene function concentrate on key enzyme genes and transcription factors associated with secondary metabolism  through regulating SA-dependent defense genes in trans or cis (S. miltiorrhiza.Recent studies suggest that DNA methylation is involved in regulation of secondary metabolism . Consistnd xylem . Althougreatment . Dynamiced by SA . Analysis or cis . SimilarSmC5-MTase genes were identified using whole genome sequence and RNA-seq data from S. miltiorrhiza. Based on phylogenetic tree and conserved domain distribution, SmC5-MTase genes were divided into four subfamilies, including SmMET, SmCMT, SmDRM and SmDNMT2. Comparative analysis of SmC5-MTases and AtC5-MTases revealed the conservation and divergence of plant C5-MTases. The transcript abundance analysis of SmC5-MTase genes suggests functional importance of SmC5-MTases in secondary metabolism and stress response in S. miltiorrhiza. The results provide useful information for understanding the role of DNA methylation in medicinal plant development and bioactive compound biosynthesis.Eight 10.7717/peerj.4461/supp-1Table S1Click here for additional data file.10.7717/peerj.4461/supp-2Table S2Click here for additional data file.10.7717/peerj.4461/supp-3Table S3Click here for additional data file.10.7717/peerj.4461/supp-4Figure S1Click here for additional data file.10.7717/peerj.4461/supp-5Figure S2Click here for additional data file.10.7717/peerj.4461/supp-6Figure S3Click here for additional data file.10.7717/peerj.4461/supp-7Figure S4Click here for additional data file.10.7717/peerj.4461/supp-8Figure S5Click here for additional data file.10.7717/peerj.4461/supp-9Data S1Raw data.Click here for additional data file."}
+{"text": "Protective effects of boswellic acid (BA) against acetaminophen (APAP)-induced hepatotoxicity in Balb/ cA mice were examined. BA, at 0.05 or 0.1%, was supplied for 4 weeks. Acute liver injury was induced by APAP treatment. Results showed that BA intake increased hepatic BA bioavailability. APAP treatment decreased glutathione (GSH) level, increased reactive oxygen species (ROS) and oxidized glutathione (GSSG) production; and lowered activity and protein expression of glutathione reductase (GR) and heme oxygenase (HO)-1 in liver. BA intake at both doses alleviated subsequent APAP-induced oxidative stress by retaining GSH content, decreasing ROS and GSSG formations, reserving activity and expression of GR and HO-1 in liver, and lowering hepatic cytochrome P450 2E1 activity and expression. APAP treatment enhanced hepatic levels of interleukin-6, tumor necrosis factor-alpha and monocyte chemoattractant protein-1. BA pre-intake diminished APAP-induced release of those inflammatory cytokines and chemokines. APAP up-regulated hepatic protein expression of toll-like receptor (TLR)-3, TLR-4, MyD88, nuclear factor kappa B (NF-\u03baB) p50, NF-\u03baB p65 and JNK. BA pre-intake at both doses suppressed the expression of NF-\u03baB p65 and p-JNK, and only at 0.1% down-regulated hepatic TLR-3, TLR-4 and MyD88 expression. APAP led to obvious foci of inflammatory cell infiltration in liver, determined by H&E stain. BA intake at both doses attenuated hepatic inflammatory infiltration. These findings support that boswellic acid is a potent hepato-protective agent. It is metabolized by hepatic cytochrome P450 system, especially CYP2E1, which leads to the overproduction of reactive free radicals and n-acetyl-p-benzoqui- noneimine (NAPQI) , 4. In aet al. [et al. [Toll-like receptors (TLRs) are pattern recognition receptors mainly responsible for immunomodulation. The activation of TLR-3 and TLR-4 has been implicated in APAP-induced hepatic damage because both TLRs could stimulate the production of adaptor proteins including MYD88 and nuclear factor kappa B (NF-\u03baB), which in turn promote the generation of oxidants and inflammatory cytokines and chemokines , 8. Furtet al.  reportedBoswellia species including Boswellia carteri, Boswellia serrata and Boswellia sacra [via its anti-oxidative and anti-inflammatory activities [et al. [Boswellic acid BA, , a pentaia sacra . It is rtivities . The stu [et al.  revealed [et al. . Thus, i\u03baB, TLR-3 and TLR-4 was evaluated. Furthermore, histological analysis was processed in order to provide more solid evidence to support the benefit of using BA as a hepatic protective agent.The purpose of this animal study was to examine the protective effects and action modes of BA at various doses upon liver of acetaminophen treated mice. The influence of this compound upon GSH, ROS and cytokines variation, CYP2E1 activity and protein expression of associated factors such as NF-2.2.1.Boswellic acid  was synthesized and provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, China. APAP (98%) was purchased from Sigma Chemical Co. .2.2.Five-to six-week-old male Balb/cA mice were obtained from National Laboratory Animal Center . Mice were housed on a 12-h light-12-h dark schedule, and fed with water and mouse standard diet . Use of the mice was reviewed and approved by the China Medical University animal care committee (104-305).2.3.BA at 0.05 or 0.1 g was mixed with 99.95 or 99.9 g powder diet to prepare 0.05 and 0.1% BA diets. Mice were divided into five groups: normal group ; BA group (0.1% BA diet), APAP group ; BA-low-APAP group (0.05% BA diet plus APAP treatment); BA-high-APAP group (0.1% BA diet plus APAP treatment). After 4 weeks supplement, normal and BA groups were sacrificed, and the other three groups were treated by a single intraperitoneal injection of APAP (400 mg/kg body weight). APAP was dissolved in phosphate-buffered saline (PBS). Mice were killed with carbon dioxide after 24 h. The mortality of mice due to APAP injection was zero. After sacrificed, liver from each mouse was collected and weighted. There was no mice die before sacrificed. Blood was also collected, and serum was separated from erythrocyte immediately. Liver tissue, 0.2 g, was homogenized on ice in 2 ml phosphate buffer (pH 7.2), and the filtrate was collected. The protein concentration of serum and liver filtrate was determined by a commercial assay kit  with bovine serum albumin as standard.2.4.et al. [g for 5 min at 4\u00b0C. Agilent 1100 series HPLC system  equipped with a C18 reversed-phase column  was used. BA quantification was performed using the external standard method. The limit of detection was 0.1 mg/g tissue. The relative standard deviations of precision and accuracy were less than 5.2%.The HPLC method of Lozano-Mena et al.  was used2.5.Serum activities of ALT and AST were determined by using commercial assay kits . CRP level (mg/l) was measured by an ELISA kit .2.6.Liver tissue was homogenized with cold PBS containing 0.05% Tween 20 and 1 mM EDTA. After centrifuging, supernatants were used for measurements. GSH and GSSG concentrations (nmol/mg protein) in liver were determined by commercial colorimetric GSH and GSSG assay kits . ROS level was quantified by using 2\u2019, 7\u2019-dichlorofluores- cein diacetate. Fluorescence value was measured by using a fluorescence microplate reader at excitation and emission wavelengths of 485 and 530 nm, respectively. Relative fluorescence unit (RFU) was the difference in fluorescence values obtained at time 0 and 5 min. Results are expressed as RFU/mg protein. The activity (U/mg protein) of GPX and GR was assayed by using commercial kits purchased from EMD Biosciences Co. .2.7.Hepatic levels of IL-6, TNF-alpha and MCP-1 were measured by using cytoscreen immunoassay kits . The sensitivity of assay with the detection limit was 5 pg/ml for IL-6, and 10 pg/ml for TNF-alpha and MCP-1.2.8.The activity of CYP2E1 in freshly prepared liver microsome was estimated by colorimetrically measuring the formation of 4-nitrocatechol, a product from p-nitrophenol hydroxylation catalyzed specifically by CYP2E1. The protein concentration of CYP2E1 was measured by ELISA, and a rabbit anti-CYP2E1 antibody  was used for detection. The formed 4-nitrocatechol was expressed as nmol/mg protein.2.9.Hepatic tissue, 40 mg, was homogenized in buffer containing protease-inhibitor cocktail purchased from Sigma-Aldrich Chemical Co.  and 0.5% Triton X-100. Homogenate was then mixed with buffer , and followed by boiling for 5 min. Protein sample at 40 was electrophoresed on 10% SDS-polyacrylamide gel, and further transferred onto nitrocellulose membranes  for 1 h. After blocking with a protein solution containing 5% skim milk for 1 h, membranes were treated with monoclonal antibody  against heme oxygenase (HO)-1, CYP2E1 (1:1000), NF-\u03baB p50, NF-\u03baB p65, JNK (1:500), TLR-3, TLR-4, MyD88 (1:2000) at 4\u00b0C overnight, and followed by incubating with horseradish peroxidase-conjugated antibody at room temperature for 3.5 h. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as a loading control, and the bands were quantified by an ATTO image analyzer .2.10.Partial liver tissue from each mouse was fixed in 10% phosphate-buffered formalin, and embedded in paraffin. Paraffin section at 5 mm thickness was cut and processed with hematoxylin-eosin (H&E) stain, and followed by examining under a light microscope for histological analysis. The severity of hepatic inflammatory injury was assayed according to the Ishak scoring system . The inj2.11.P < 0.05.The effect of each treatment was analyzed from 10 mice (n = 10) in each group. Data were reported as means \u00b1 standard deviation (SD), and subjected to analysis of variance. Differences among means were determined by the Least Significance Difference Test with significance defined at 3.3.1.. The intake of BA at both doses alleviated subsequent APAP-induced elevation of ALT, AST and CRP levels in serum (P < 0.05).BA intake at 0.1% increased hepatic BA content, and did not affect body weight, feed intake, water intake and liver weight 3.2.P < 0.05); but BA pre-intake attenuated APAP-induced GSH depletion and decreased hepatic GSSG and ROS production (P < 0.05). APAP raised GPX activity and reduced GR activity in liver (P < 0.05). BA intake failed to change hepatic GPX activity (P > 0.05); but significantly maintained hepatic GR activity (P < 0.05). As shown in P < 0.05). BA intake did not affect GPX protein expression (P > 0.05); but significantly retained protein expression of GR and HO-1 in liver (P < 0.05). APAP treatment enhanced CYP2E1 activity and expression (P < 0.05); however, BA intake significantly suppressed subsequent APAP-induced elevation of CYP2E1 activity and protein expression (P < 0.05). APAP treatment also significantly increased hepatic release of TNF-alpha, IL-6 and MCP-1 (P < 0.05). BA intake lowered APAP-induced hepatic production of these cytokines (P < 0.05).As shown in 3.3.P < 0.05). BA pre-intake at both doses down-regulated the expression of NF-\u03baB p65 and p-JNK (P < 0.05), and only at high dose suppressed hepatic TLR-3, TLR-4 and MyD88 expression (P < 0.05).APAP up-regulated hepatic protein expression of TLR-3, TLR-4, MyD88, NF-\u03baB p50, NF-\u03baB p615, JNK and p-JNK (\u03baB p50 expression (P > 0.05).BA did not alter NF-3.4.P < 0.05). BA intake at 0.05 and 0.1% decreased hepatic infiltration by inflammatory cells and lowered Ishak inflammation scores (P < 0.05), in which 0.1% BA treatment exhibited greater anti-inflammatory effects than 0.05% BA (P < 0.05).Histological data revealed that BA intake at 0.1% only (without APAP) did not cause inflammatory stress and showed similar Ishak inflammation score as normal groups . APAP tr4.Boswellia species, increased its deposit in liver and protected liver against subsequent APAP induced oxidative and inflammatory injury. Besides increasing GSH retention, we found that BA effectively decreased CYP2E1 activity and expression, lowered ROS, TNF-alpha and MCP-1 production, as well as suppressed HO-1, TLR-3, TLR-4, NF-\u03baB p65 and p-JNK expression. In addition, our histological results indicated that BA improved diffuse ballooning degeneration and lobular inflammation caused by APAP These findings support that BA is a potent hepatic protective agent.Our present study revealed that the pre-intake of BA, an active compound of APAP at high dose led to GSH depletion and GSSG accumulation . Our datvia its anti-oxidative activities, which definitely led to less stimulation for hepatic NF-\u03baB activation and JNK phosphorylation. Our western blot data regarding hepatic protein expression of NF-\u03baB p65 and p-JNK agreed that BA intake at both doses limited the activation of these two signal pathways. Consequently, it is reasonable to observe lower hepatic levels of IL-6, TNF-alpha and MCP-1 in BA treated mice. The decrease in serum levels of ALT, AST and CRP in BA treated mice also agreed that BA intake declined APAP caused liver inflammatory stress. In addition, our histological data further supported that BA pre-intake effectively improved hepatic inflammatory injury.APAP overdose activated NF-\u03baB and JNK pathways through stimulating ROS generation , 23. Theet al. [et al. [in vivo anti-inflammatory data of BA, and extended BA\u2019s mediating activity to TLRs and MyD88.TLRs play crucial roles in the pathological progression of inflammatory liver diseases because TLRs recognize endogenous damage-associated molecular patterns during inflammatory reactions . TLR-3 iet al.  reported [et al.  reportedBA is naturally present in many edible plants including vegetables and herbs , 30. The5.via maintaining hepatic GSH content, retaining activity and expression of GR and HO-1, decreasing inflammatory cytokines and suppressing protein expression of CYP2E1, NF-\u03baB p65, JNK, TLR-3 and TLR-4. These findings support that boswellic acid was a potent hepatic protective agent.Boswellic acid pre-intake protected mice liver against subsequent acetaminophen-induced oxidative and inflammatory injury All authors claimed that no Conflict of Interest."}
+{"text": "Pochazia shantungensis adults and nymphs, newly recorded pests, were evaluated. The LC50 values of Thymus vulgaris, Ruta graveolens, Citrus aurantium, Leptospermum petersonii and Achillea millefolium oils were recorded as 57.48, 84.44, 92.58, 113.26 and 125.78\u2009mg/L, respectively, against P. shantungensis nymphs using the leaf dipping bioassay, and 75.80, 109.86, 113.26, 145.06 and 153.74\u2009mg/L, respectively, against P. shantungensis adults using the spray bioassay method. Regarding volatile components identified in T. vulgaris oil, the LC50 values of carvacrol and thymol using the leaf dipping bioassay against P. shantungensis nymphs were 56.74 and 28.52\u2009mg/L, respectively. The insecticidal action of T. vulgaris oil against P. shantungensis could be attributed to carvacrol and thymol. Based on the structure-toxicity relationship between thymol analogs and insecticidal toxicities against P. shantungensis nymphs similar to the LC50 values against P. shantungensis adults, the LC50 values of thymol, carvacrol, citral, 2-isopropylphenol, 3-isopropylphenol, and 4-isopropylphenol were 28.52, 56.74 and 89.12, 71.41, 82.49, and 111.28\u2009mg/L, respectively. These results indicate that the insecticidal mode of action of thymol analogs may be largely attributed to the methyl functional group. Thymol analogues have promising potential as first-choice insecticides against P. shantungensis adults and nymphs.The insecticidal toxicities of five essential oils against  Pochazia shantungensis Chou & Lu. is a hemipterous insect that belongs to the Ricaniidae insect family, which includes about 400 described species in over 40 generaP. shantungensis is a newly recorded pest that is economically devastating for various trees including apple, blueberry, peach and persimmon, mainly in Wanjugun, KoreaP. shantungensis was first recorded in Chungcheongnamdo in 2010P. shantungensis has subsequently occurred sporadically in several other provinces across Korea2P. shantungensis adults and nymphs, despite the urgent demandP. shantungensis.Cynanchum paniculatum oilMentha piperita oilMentha spicata oil3789Plant oil is a very complex mixture that can contain approximately 30\u201365 constituents at various concentrations35Thymus vulgaris oil-derived constituents against P. shantungensis. Therefore, the aims of the present study were first to investigate the insecticidal properties of T. vulgaris oil-derived components against P. shantungensis adults and nymphs, and then to determine the structure-activity relationship between thymol analogs and insecticidal toxicities.So for no report has been received about the insecticidal toxicities of P. shantungensis adults and nymphs. Essential oils of Achillea millefolium flowers, Citrus aurantium fruits, Leptospermum petersonii leaves, Ruta graveolens leaves and T. vulgaris leaves were analyzed  of P. shantungensis adults and nymphs. Differences in the insecticidal toxicities of plant-derived oils may be explained on the basis of species-specific responses to plant species, phytochemicals, and the weight and size of P. shantungensis adults and nymphsThis study was undertaken within the framework of a more general study involving the natural products for insecticidal toxicities against analyzed . The yieexposure . From thP. shantungensis adults and nymphs, the components of A. millefolium, C. aurantium, L. petersonii, R. graveolens, and T. vulgaris oils were investigated by GC-MS analysis. The identified components, together with the percentages present in the essential oils are displayed in A. millefolium oil; limonene (87.75%), citral (3.21%), limonene oxide (2.29%), and (\u2212)-carveol (1.75%) in C. aurantium oil; citral (48.12%), \u03b2-citronellal (19.50%), isopulegol (8.14%), geraniol (4.64%), and linalool (3.33%) in L. petersonii oil, and thymol (23.34%), undecyl trichloroacetate (18.52%), methyltridecyl pentanoate (16.18%), and 2,2-dimethylpropanoic acid (7.03%), 2-acetoxytetradecane (6.98%), palmitic acid (5.07%), and methyl linolenate (4.78%) in R. graveolens oil. The major components in T. vulgaris oil were thymol (40.04%), \u03c1-cymene (29.97%), \u03b3-terpinene (8.17%), linalool (4.99%), terpinolene (3.26%), \u03b1-pinene (2.84%), \u03b2-caryophyllene (2.50%), carvacrol (2.45%), limonene (1.25%), \u03b1-phellandrene (1.20%), myrcene (0.91%), camphene (0.89%), and caryophyllene oxide (0.21%). Together, thymol, \u03c1-cymene and \u03b3-terpinene made up 78.18% of T. vulgaris oil. The volatile components consisted of 8 monoterpene hydrocarbons , 1 monoterpene alcohol , 2 monoterpene phenols (carvacrol and thymol) and 2 sesquiterpene hydrocarbons (\u03b2T. vulgaris oil were borneol (0.98%), camphene (0.60%), carvacrol (2.81%), \u03b2-caryophyllene (2.39%), 1,8-cineol (0.96%), p-cymene (25.2%), myrcene (1.93%), linalool (2.86%), \u03b1-pinene (1.16%), 1-octen-3-ol (1.19%), \u03b1-terpinene (1.02%), \u03b3-terpinene (6.37%), \u03b1-thujene (1.50%) and thymol (42.27%). The fact that the essential oil of T. vulgaris leaves dried more at 45\u2009\u00b0C than at 30\u2009\u00b0C can be explained mainly by the increase of thymol by 8% and carvacrol by 12%, while the quantity of \u03b3-terpinene was decreased by 4.9%T. vulgaris were affected by the environmental conditions, including harvest time, genotype, storage period, handling method and intraspecific variability, and the experimental conditions, which included the extraction method, extracted plant parts, and plant tissue drying temperatureTo further explore the insecticidal toxicities of the five essential oils against oxide 2.2%, and  derived from the five essential oils were evaluated using spray and leaf dipping bioassays against P. shantungensis adults and nymphs -carveol, geraniol, and bornyl acetate) did not exhibit any insecticidal toxicity against P. shantungensis adults and nymphs (data not shown). The insecticidal toxicities of the essential oils appear to be connected to their chemical composition. The insecticidal toxicities of T. vulgaris and R. graveolens oils could be due to the existence of thymol and carvacrol, which exhibited the greatest insecticidal toxicities. The essential oils of C. aurantium and L. petersonii contain citral, which showed insecticidal toxicities against P. shantungensis adults and nymphs, however its toxicity was weaker than thymol and carvacrol. Furthermore, P. shantungensis nymphs were more susceptible to T. vulgaris oil, carvacrol and thymol, when compared to P. shantungensis adults -carveol, (+)-carvone, \u03b2-caryophyllene, caryophyllene oxide, \u03b2-citronellal, citral, 1,8-cineole, d nymphs . Based os adults . In a prP. shantungensis adults and nymphs, thymol, carvacrol, 2-isopropylphenol, 3-isopropylphenol, and 4-isopropylphenol were selected as thymol analogs for testing , decyl chloroformate (97%), dodecanoic acid (98%), \u03b2-farnesene (90%), geranyl acetate (97%), geraniol (98%), isopulegol (98%), linalool (97%), limonene (97%), limonene oxide (97%), methyl linolenate (99%), myrcene (90%), myristic acid (99%), palmitic acid (99%), \u03b1-phellandrene (85%), \u03b1-pinene (98%), pivalic acid (99%), sabinene (75%), \u03b1-terpineol (90%), \u03b3-terpinene (97%), 4-terpineol (95%), terpinolene (90%), and tymol (99%) were obtained from Sigma-Aldrich . Achillea millefolium L. flowers, Citrus aurantium L. fruits, Leptospermum petersonii F. M. Bailey leaves, Ruta graveolens L. leaves, and Thymus vulgaris L. leaves were collected from a local store in Chonju, Korea. Sample specimens were authenticated by Jeongmoon Kim at Chonbuk National University, Korea. Essential oils of the five plants were obtained by steam distillation extraction, and finally dried over Na2SO4 to extract the pure essential oils (Bornyl acetate (99%), camphene (95%), camphor (96%), carvarol (98%), (\u2212)-carveol (95%), (+)-carvone (96%), \u03b2-caryophyllene (98.5%), caryophyllene oxide (95%), \u03b2-citronellal (95%), citral (95%), 1,8-cineole (99%), ial oils .P. shantungensis adults and nymphs were collected from persimmon trees in Wanjugun, Korea and classified the fourth instar stages of P. shantungensis nymphs and adults as detailed elsewhere2P. shantungensis adults and nymphs were assessed  and were separated with HP-Innowax capillary column and DB-5 column . The conditions of the column were as follows: Helium at 0.75\u2009mL/min; column temperature (51 to 201\u2009\u00b0C) at 2\u2009\u00b0C/min; injector temperature (211\u2009\u00b0C); split ration (48:1); ion source temperature (231\u2009\u00b0C); ionization potential (70e\u2009V); and mass spectra range (50\u2013800\u2009amu). The components of T. vulgaris oils were evaluated according to retention times, retention indices, and mass spectra and were identified by comparison with a spectrum library (T. vulgaris oil constituent (%) was measured by comparison with internal standards.The components of the essential oil extracted from  library . The rel50) value and the slope of the regression lines were calculated using the statistical package SPSS, version 12.0 for Windows.Data obtained for each dose response bioassay were subjected to probit analysis. The median lethal concentration .Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations."}
+{"text": "MiR-486 was found to be associated with cancer\u2019s diagnosis and prognosis. This meta-analysis aimed to investigate the potential effect of miR-486 on cancer detection and prognosis.We searched PubMed, Cochrane library, Embase, Chinese National Knowledge Infrastructure (CNKI) and Wanfang databases to find all correlated articles. The STATA 11.0 was applied to estimate the pooled effects, heterogeneity and publication bias.The pooled sensitivity (SEN), specificity (SPE) and Area under the curve (AUC) were 82% (95% CI: 78\u201385%), 88% (95% CI: 83\u201392%) and 0.91 (95% CI: 0.88\u20130.93). Subgroup analysis indicated miR-486 from circulating samples exhibited higher diagnostic accuracy with the AUC was 0.90 (95% CI: 0.87\u20130.92) than miR-486 from other specimen with the AUC of 0.78 (95% CI: 0.75\u20130.82) and miR-486 obtained a better diagnostic value in the Asian population with the AUC of 0.94 (95% CI: 0.91\u20130.95) than the Caucasian and Caucasian/African population with the AUC of 0.80 (95% CI: 0.76\u20130.83) and 0.89 (95% CI: 0.86\u20130.91) respectively. MiR-486 obtained high value for the diagnosis of non-small cell lung cancer with SEN, SPE and AUC were 0.82 (95% CI: 0.0.77\u20130.87), 0.90 (95% CI: 0.84\u20130.94) as well as 0.92 (95% CI: 0.89\u20130.94) respectively. For the 7 prognostic tests, the pooled hazard ratio (HR) was 0.48 (95% CI: \u20130.13\u20131.08) for low versus high miR-486 expression.This meta-analysis indicated that miR-486 can be used as ideal biomarkers in the cancer\u2019s diagnosis. However, Low miR-486 expression did not increase the risk of poor outcome. MicroRNA is a group of 19\u201322 nucleotide, small, single-stranded and conserved non-coding RNA that acts as a regulator of gene expression at both the post-transcriptional and the translational levels through acting on the 3\u2019-untranslated region (UTR) of messenger RNA (mRNA) , SPE [TN/ (FP+TN)], the positive likelihood ratio (PLR) [(SEN/ (1-SEN)], the negative likelihood ratio (NLR) [(1-SPE)/SPE)], the diagnostic odds ratio (DOR) [PLR/ NLR] and the pooled HR with its 95% Confidence Interval (95% CI) respectively. We also constructed the SROC curve and calculated the AUC value. Simultaneously, we assessed the heterogeneity among the selected studies through the I2 value . The P vnificant . As for This was the first meta-analysis to confirm the potential value of miR-486 on cancer diagnosis and prognosis. The expression of miR-486 might be an effective biomarker for detection of human cancer."}
+{"text": "A new recombinant norovirus GII.P16-GII.2 outnumbered pandemic GII.4 as the predominant GII genotype in the winter of 2016\u20132017 in Hong Kong, China. Half of hospitalized case-patients were older children and adults, including 13 young adults. This emergent norovirus targets a wider age population compared with circulating pandemic GII.4 strains. Noroviruses are leading causes of acute gastroenteritis (http://www.rivm.nl/mpf/norovirus/typingtool).Since August 2012, we have conducted an ongoing molecular surveillance of norovirus genotype distribution in all hospitalized gastroenteritis patients in our teaching hospital in Hong Kong . The female-to-male ratio was 1.04:1. We successfully genotyped 357 (90.8%) samples. The top 3 circulating VP1 genotypes during the study period were GII.4 (n = 214 [54.5%]), GII.2 (n = 86 [21.9%]), and GII.3 (n = 16 [4.1%]). Before this season, GII.2 had been a rare genotype, accounting for <1% of total strains circulating locally , panel Ahttp://www.megasoftware.net)  from the samples of 20 case-patients  as previously described (are.net) , panel AThe GII.2 strains we identified clustered most closely with the recombinant GII.P16-GII.2 strains from Germany during the same period . SubsequU-test)  , panel CU-test) , panel CU-test)  Figure. We noted a gradual narrowing in the age distribution of GII.4 infections to young children <5 years of age in the past 5 seasons , presumaWe report the emergence of a recombinant norovirus GII.P16-GII.2 variant that surpassed the previously predominant genotype GII.4 in hospitalized acute gastroenteritis case-patients in the winter of 2016\u20132017 in Hong Kong. However, unlike the recently emerged epidemic GII.17 Kawasaki variant that predominated only in part of Asia (China and Japan) during 2014\u20132016  apart from causing outbreaks. Collectively, these findings might have important implications for norovirus vaccine formulation and vaccination strategy. Close monitoring of the global spread of GII.P16-GII.2 is warranted.Incidence of hospitalization in case-patients with GII.4 and GII.2 infections, Hong Kong, China, July 2016\u2013February 2017."}
+{"text": "Mycobacterium porcinum is a rapidly growing environmental mycobacterium responsible for opportunistic infections. The 7,025,616-bp draft genome of M. porcinum strain CSURP1564 exhibits a 66.71% G+C content, 6,687 protein-coding genes, and 65 predicted RNA genes. In silico DNA-DNA hybridization confirms its assignment to the Mycobacterium fortuitum complex. Mycobacterium porcinum, a member of the Mycobacterium fortuitum third biovariant complex, is a nontuberculous mycobacterium primarily recovered from swine with lymphadenitis  oleic acid-albumin-dextrose-catalase  under a 5% CO2 atmosphere. Colonies were confirmed by matrix-assisted laser desorption ionization\u2013time of flight mass spectrometry (M. porcinum CSURP1564 genome was incorporated into in silico DNA-DNA hybridization (DDH) (M. porcinum IP141460001 (GenBank accession number MVIG00000000), 49.5% with M. boenickei CIP 107829 (FUWC00000000), 34.5% with M. conceptionense D16 (CTEF00000000), 34.4% with M. farcinogenes DSM 43637 (CCAY000000000) and M. neworleansense ATCC 49404 (CWKH00000000), 31.40% with M. fortuitum CT6 (NCBI reference sequence number NZ_CP011269), and 20.5% with M. avium 104 (NCBI reference sequence NC_008595).trometry . Reads itrometry , and contrometry , GapFilltrometry , and mantrometry  yielded on (DDH)  using GGon (DDH) , with reM. porcinum strain CSURP1564 differs from M. conceptionense in the M. fortuitum complex. This is the first report of MinION technology applied to the genome sequencing of a nontuberculous Mycobacterium species , European Nucleotide Archive (ENA), under the accession number OLMG00000000 (OLMG01000001 to OLMG01000005).The draft genome sequence of"}
+{"text": "Lactobacillus plantarum Oregon-R-modENCODE strain BDGP2 was isolated from the gut of Drosophila melanogaster for functional host microbial interaction studies. The complete genome comprised a single circular genome of 3,407,160\u00a0bp, with a G+C content of 44%, and four plasmids. Lactobacillus plantarum increases amino acid metabolism and promotes larval growth in Drosophila melanogaster under nutrient-scarce conditions (L.\u00a0plantarum strains include one derived from a laboratory fly strain (KP) and one from wild-caught flies (DF)  . Here, wL. plantarum Oregon-R-modENCODE strain BDGP2 was isolated from a fecal swab. Bacteria were streaked onto Difco de Man, Rogosa, and Sharpe (MRS) agar  agar  agar (BDRS) agar  and sequRS) agar . DNA wasRS) agar , and whoRS) agar . A singlsion 2.0 , 9. The sion 2.0  and the sion 2.0 .L.\u00a0plantarum strains, ours contains integrated likely prophages . There are seven copies, six ranging in size from 36,225 to 46,235\u00a0bp and in sequence similarity from 45 to 70% and a partial copy of only 13,544 kb. Together they constitute 7.5% of the genome. Two cornerstone proteins highly conserved in prophages are the large terminase subunit and the portal protein (L. plantarum strain DF plasmid unnamed1 (GenBank accession number CP013754) and L. plantarum strain TMW 1.277  plasmid pL1277-4 (accession number CP017367), respectively.The chromosomal genome annotation predicts 3,148 protein-coding genes, 131 pseudogenes, 5 rRNA operons, a single 5S rRNA, and 85 tRNAs . Of the 3,148 protein-coding genes, 328 are contained within candidate prophages. Like other  protein . These cL. plantarum Oregon-R-modENCODE strain BDGP2 have been deposited in GenBank under the accession numbers CP023174 (chromosome) and CP023175 (pLtBDGP2A), CP023176 (pLtBDGP2B), CP023177 (pLtBDGP2C), and CP023178 (pLtBDGP2D) (plasmids).The complete genome sequences of the chromosome and four plasmids of"}
+{"text": "Both spontaneous hepatitis C virus (HCV) clearance and the achievement of sustained virological response (SVR) by anti-viral therapy greatly reduce the incidence of hepatocellular carcinoma (HCC). The current study aimed to compare the risk of HCC between the two patient groupsA total of 313 subjects with spontaneous HCV clearance (SC) and 564 age- and sex-matched patients in the treatment-induced SVR group were enrolled for analysis.Nineteen (2.2%) of the 877 patients developed HCC during 6,963 person-years of follow-up. Fourteen (2.5%) SVR patients and 5 (1.6%) SC patients developed HCC (P=0.004). Cox regression analysis of factors predictive of HCC included SVR , diabetes (HR/CI:3.41/1.21-9.58), and age (HR/CI: 1.07/1.01-1.14). Of the 564 SVR patients, eleven (5.9%) of the 187 patients with fibrosis stage 2-4 (F2-4) and 2 (0.9%) of the 226 patients with F01 developed HCC (P=0.01). Compared to SC subjects, only SVR patients with F2-4 (P<0.001) but not F0-1(P=0.60) had a higher risk of HCC development. Cox-regression analysis using liver fibrosis as a variable demonstrated that factors associated with HCC included SVR with F2-4 (versus SC: HR/CI: 10.06/2.20-45.98), diabetes (HR/CI:3.23/1.14-9.19), and age (HR/CI: 1.08 1.02-1.15).Compared to subjects with spontaneous viral clearance, subjects with antiviral treatment-induced HCV viral clearance remain at high risk for HCC development, especially if they have significant hepatic fibrosis. These results may provide important information for decision-making regarding the prioritization of current direct antiviral agents in resource-limited countries. Spontaneous hepatitis C virus (HCV) seroclearance occurs in 20-30% of patients after acute infection \u20133. SubjeA total of 313 subjects in the SC group (participants who had anti-HCV seropositivity and undetectable HCV RNA at baseline (n = 291) and those who had anti-HCV seropositivity and detectable HCV RNA levels at baseline, but HCV RNA became undetectable after more than one-year follow-up (n = 22)) and 564 age- and sex-matched patients in the SVR group were enrolled for analysis with a follow-up period of 4,224and 2,739 person-years, respectively. The baseline characteristics are shown in Table Nineteen 2.2%) of the 877 patients developed HCC over 6,963 person-years of follow-up, including fourteen (2.5%) SVR patients and 5 1.6%) SC patients with an incidence of 0.51% and 0.12% per person-year, respectively (P=0.004)  of the 564 SVR patients had liver histology available within six months of initiating the antiviral therapy. Of them, eleven (5.9%) of the 187 patients with F2-4 and two (0.9%) of the 226 patients with F0-1 developed HCC (Log-Rank test P=0.01). Compared to SC subjects, only SVR patients with F2-4 (Log-Rank test P<0.001), but not F0-1(Log-Rank test P=0.60), had significantly higher risk of HCC development  HCV cohort. The R.E.V.E.A.L-HCV study is a community-based cohort and enrolled participants from seven townships in Taiwan during 1991-1992. The details of the study participants, interview, blood collection and laboratory examinations have been described previously in detail. , 23 Sex-Interleukin 28B (IL-28B) rs8099917 was selected as the single-nucleotide polymorphism (SNP) to be tested for its association with HCC. The genetic testing was determined using methods that have been described previously , 25\u201327.The baseline characteristics of the clinical cohort and R.E.V.E.A.L.-HCV cohort were compared by chi-squared tests. The cumulative risk of the incidence of HCC among patients who occurred spontaneous HCV RNA clearance (R.E.V.A.E.L.-HCV cohort) and treatment-induced RNA clearance was estimated by Kaplan-Meier method, and the statistical significance of the difference was examined by log-rank tests. Cox proportional hazards models were used to estimate multivariate-adjusted hazard ratios (HR) with 95 percent confidence intervals (95% CI) for HCC risk for various groups with RNA clearance after adjustment for other risk factors. Statistical significance levels were determined by a 2-sided P-value of 0.05. The proportionality assumption (non-changing HRs over time) of Cox models was examined, and the assumption was not violated. All analyses were performed using the SAS statistical software package ."}
+{"text": "ATZD00000000) is an aerobic, motile, Gram-negative, non-spore-forming rod that was isolated from an effective nitrogen-fixing nodule of Medicago laciniata (L.) Miller from a soil sample collected near the town of Guatiza on the island of Lanzarote, the Canary Islands, Spain. This strain nodulates and forms an effective symbiosis with the highly specific host M. laciniata. This rhizobial genome was sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) sequencing project. Here the features of 10.1601/nm.1335 Mlalz-1 are described, together with high-quality permanent draft genome sequence information and annotation. The 6,664,116\u00a0bp high-quality draft genome is arranged in 99 scaffolds of 100 contigs, containing 6314 protein-coding genes and 74 RNA-only encoding genes. Strain Mlalz-1 is closely related to 10.1601/nm.133510.1601/strainfinder?urlappend=%3Fid%3DIAM+12611T, 10.1601/nm.1334 A 321T and 10.1601/nm.1783110.1601/strainfinder?urlappend=%3Fid%3DORS+1407T, based on 16S rRNA gene sequences. gANI values of \u226598.1% support the classification of strain Mlalz-1 as 10.1601/nm.1335. Nodulation of M. laciniata requires a specific nodC allele, and the nodC gene of strain Mlalz-1 shares \u226598% sequence identity with nodC of M. laciniata-nodulating 10.1601/nm.1328 strains, but \u226493% with nodC of 10.1601/nm.1328 strains that nodulate other Medicago species. Strain Mlalz-1 is unique among sequenced 10.1601/nm.1335 strains in possessing genes encoding components of a T2SS and in having two versions of the adaptive acid tolerance response lpiA-acvB operon. In 10.1601/nm.1334 strain 10.1601/strainfinder?urlappend=%3Fid%3DWSM+419, lpiA is essential for enhancing survival in lethal acid conditions. The second copy of the lpiA-acvB operon of strain Mlalz-1 has highest sequence identity (> 96%) with that of 10.1601/nm.1334 strains, which suggests genetic recombination between strain Mlalz-1 and 10.1601/nm.1334 and the horizontal gene transfer of lpiA-acvB.10.1601/nm.1335 Mlalz-1 (INSDC = The online version of this article (10.1186/s40793-017-0270-2) contains supplementary material, which is available to authorized users. Medicago spp.) are some of the most important and extensively grown pasture legumes and their specific symbiosis with strains of rhizobia belonging to either 10.1601/nm.1328 (synonym 10.1601/nm.1339) meliloti or the closely related species 10.1601/nm.1334 [Symbiotic nitrogen fixation by pasture legumes and their associated root nodule bacteria provides a critical contribution to sustainable animal and plant production, and the maintenance of soil fertility in agricultural systems \u20133. As su/nm.1334 , 6 has b/nm.1334 .Medicago laciniata (L.) Miller (cut leaf medic), an annual native of southern and eastern Mediterranean and Saharo-Sindian countries, is of importance because of its ability to grow in comparatively arid habitats and marginal cropping areas [Medicago sativa L.  or Medicago truncatula Gaertn. [nod genes, in particular a specific nodC allele [M. truncatula and M. laciniata shared chromosomal identity, but differed in their nodC alleles [M. laciniata and bv. meliloti for the classical 10.1601/nm.1335 group that efficiently nodulates M. sativa. However, in subsequent studies the diversity observed within bv. medicaginis strains indicate that this group is certainly heterogeneous [ng areas \u201311. It i Gaertn. , 13. ThiC allele . For exa alleles . Based oM. laciniata is native to the Canary Islands and is present on all of the islands of this archipelago, growing in environments that range from arid to subhumid. 10.1601/nm.1335 strain Mlalz-1 was isolated from a N2-fixing nodule of M. laciniata grown in alkaline soil (pH\u00a09.0) collected in Guatiza, in the arid Northeast of Lanzarote Island, in 2007. This strain was one of the rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 GEBA-RNB project proposal [proposal , 18. HerLeft and Center). It is fast growing, forming colonies after 3\u20135\u00a0days when grown on \u00bdLA, TY, or a modified yeast-mannitol agar [Right). Minimum Information about the Genome Sequence (MIGS) for strain Mlalz-1 is provided in Table\u00a010.1601/nm.1335 Mlalz-1 is a motile, non-sporulating, non-encapsulated, Gram-negative strain in the class 10.1601/nm.809. The rod shaped form has dimensions of approximately 0.5\u00a0\u03bcm in width and 1.0\u20132.0\u00a0\u03bcm in length  [T (= 10.1601/strainfinder?urlappend=%3Fid%3DLMG+19920T) [T [T, 99.7% to 10.1601/nm.1334 A 321T and 99.5% to 10.1601/nm.1783110.1601/strainfinder?urlappend=%3Fid%3DORS+1407T. In contrast, 10.1601/nm.133510.1601/strainfinder?urlappend=%3Fid%3DIAM+12611T and 10.1601/nm.133710.1601/strainfinder?urlappend=%3Fid%3DLMG+7834T [Previous studies using multilocus sequence typing showed that al group . Phyloge+6133T) , 10.160120T) [T . The avaG+7834T  were onlM. laciniata, which is suited for cultivation in arid environments [10.1601/nm.1335 Mlalz-1 was selected for sequencing at the U.S. Department of Energy funded Joint Genome Institute as part of the GEBA-RNB project , 18. Theronments . The 10.ronments  and a hironments . Sequenc600nm of 0.6 was reached. DNA was isolated from 50\u00a0ml of cells by Peter van Berkum according to the method described by van Berkum [\u22121.10.1601/nm.1335 Mlalz-1 (= 10.1601/strainfinder?urlappend=%3Fid%3DUSDA+1984) was cultured on MAG solid media  for thren Berkum . The finThe draft genome of 10.1601/nm.1335 Mlalz-1 was generated at the DOE Joint genome Institute (JGI) using Illumina technology . An IlluUniProt, TIGRFam, Pfam, KEGG, COG, and InterPro databases. The tRNAScanSE tool [Walnut Creek, CA, USA.Genes were identified using Prodigal , as partnSE tool  was usednSE tool . Other nnSE tool . AdditionSE tool  developeThe genome is 6,664,116\u00a0bp with 62.16% GC content Table\u00a0 and compT (= 10.1601/strainfinder?urlappend=%3Fid%3DLMG+6133T), 10.1601/nm.1334 A 321T (= 10.1601/strainfinder?urlappend=%3Fid%3DLMG+19920T) and 10.1601/nm.1783110.1601/strainfinder?urlappend=%3Fid%3DORS+1407T. As the genomes of these type strains have not been sequenced or are not publically available, gANI values [repC, which is present as a single copy on 10.1601/nm.1335 pSymA and pSymB. The 10.1601/nm.1335 Mlalz-1 genome carried 2 repC loci (A3CADRAFT_00120 and A3CADRAFT_01676) with highest encoded protein identity to RepC proteins of 10.1601/nm.1335 strains. Mlalz-1 A3CADRAFT_00120 RepC1 had highest identity (98.10%) to the RepC1 protein encoded by SMb20044 on pSymB of 10.1601/nm.1335 1021. 10.1601/nm.1335 Mlalz-1 A3CADRAFT_01676 RepC2 had highest identity (99.00%) to the RepC2 protein encoded by SMa2391 on pSymA of 10.1601/nm.1335 1021. This indicated the presence of two megaplasmids in strain Mlalz-1, and revealed that strain Mlalz-1 has a similar genome architecture to that of 10.1601/nm.1335 1021.10.1601/nm.1335 Mlalz-1 is one of seven strains of 10.1601/nm.1335 that have been sequenced from the GEBA-RNB genome sequencing projects . On the I values  had to bI values . The totgspDOGLMCKEFHIJ necessary for a functional T2SS, but lacks the gspS gene found only in certain genera [.All 29 10.1601/nm.1335 strains within the gANI clique share a core set of 4948 orthologous genes, using cut off values of 1e-5 and 30% minimum protein identity. 10.1601/nm.1335 Mlalz-1 contains 176 unique genes, 96 (54.5%) of which encode hypothetical proteins. The unique genes include those encoding the components of a T2SS, located on scaffold A3CADRAFT_scaffold_5.6 Fig.\u00a0, as welln genera  , actP, actR, actS, phrR, exoR, exoH, lpiA, acvB, degP1, mdh3, fbaB, groS, kdpB, kdpC, fixN2 and fixO2 [lpiA-acvB operon. One operon (A3CADRAFT_01189-A3CADRAFT_01190) is found on scaffold A3CADRAFT_scaffold_3.4, in a gene region that is conserved in other 10.1601/nm.1335 (sequence similarity >98%) and is located on the chromosome of the fully sequenced 10.1601/nm.1335 1021. The second version of the lpiA-acvB operon (A3CADRAFT_05694-A3CADRAFT_05695) is located on A3CADRAFT_scaffold_47.48, in a gene region that is conserved in 10.1601/nm.1334 genomes (sequence similarity >96%) and is located on the pSMED02 symbiotic plasmid of the fully sequenced 10.1601/nm.133410.1601/strainfinder?urlappend=%3Fid%3DWSM+419. The regulatory gene fsrR, required for the acid activated expression of lpiA in 10.1601/nm.133410.1601/strainfinder?urlappend=%3Fid%3DWSM+419 [lpiA-acvB gene regions of all other 10.1601/nm.1335 sequenced genomes. These findings suggest that 10.1601/nm.1335 Mlalz-1 acquired the plasmid-borne lpiA-acvB operon and associated fsrR regulatory gene by lateral transfer from an 10.1601/nm.1334 strain.The Mlaz-1 genome also contains acid-tolerance or acid-responsive genes that are orthologous to the genes identified in the comparatively acid tolerant strain 10.1601/nm.133410.1601/strainfinder?urlappend=%3Fid%3DWSM+419. Acid-tolerance or acid-responsive genes identified in Mlaz-1 include nd fixO2 \u201352  genes identified in the 10.1601/nm.1335 Mlalz-1 genome  with nodC of other M. laciniata-nodulating 10.1601/nm.1328 strains in the NCBI database, whereas there is a lower sequence identity (\u2264 93%) with nodC of 10.1601/nm.1328 strains that nodulate other Medicago species. Nodulation of Medicago hosts requires Nod factors that are sulfated at the reducing terminus and acylated at the non-reducing terminus, with a polyunsaturated fatty acyl tail [Medicago species since the nodEF, nodL and nodHPQ genes that are required for these specific decorations of the Nod factor are present in the genome. 10.1601/nm.1335 Mlalz-1 also possesses the three nodD genes that mediate host-specific activation of nodABC in the symbiotic interactions of 10.1601/nm.1335 with Medicago [Essential symbiotic  with that of sequenced 10.1601/nm.1334 strains, which infers horizontal gene transfer of this region from 10.1601/nm.1334.10.1601/nm.1335 Mlalz-1 is a rhizobial strain that is able to nodulate and fix nitrogen with the highly specific host  strains . HoweverAdditional file 1: Table S1.Ensifer meliloti Mlalz-1. (DOCX 52\u00a0kb)Associated MIGS record for Additional file 2: Table S2-S4.Table S2. Acid responsive gene orthologs present in Ensifer strains. Table S3. The nodulation genes of Ensifer meliloti Mlalz-1. Table S4. The nitrogen fixation genes of Ensifer meliloti Mlalz-1. (DOCX 65\u00a0kb)"}
+{"text": "In the Local score development  subsection of the Results section, there is an error in the model equation following the second paragraph. The correct equation is:Log odds of in-hospital mortality = -4.67 + (moderate hypoxemia x 0.43) + (severe hypoxemia x 1.62) +  +  +  + (wheeze x -0.35) + (unconsciousness x 1.74)"}
+{"text": "Since 2005, anti-hepatitis B virus (anti-HBV) vaccine is part of the Expanded Program on Immunization (EPI) for infants born in Cameroon, with 99% anti-HBV coverage. In a context of generalized HIV epidemiology, we assessed paediatric anti-HBV vaccine response according to HIV status, feeding option and age in a tropical context.Prospective, observational and cross-sectional study conducted among 82 children , after complete anti-HBV vaccination (Zilbrix Hepta: 10\u03bcg AgHBs) at the Essos Health Centre in Yaounde, Cameroon, classified as group-A: HIV unexposed (28), group-B: HIV-exposed/uninfected (29), group-C: HIV-infected (25). Quantitative anti-HBs ELISA was interpreted as \u201cno\u201d, \u201clow-\u201d or \u201cprotective-response\u201d with <1, 1\u201310, or \u226510 IU/L respectively; with p-value<0.05 considered significant.95% 0.933\u20137.500], p = 0.041. According to feeding option during first six months of life, 47.67% (21/45) developed a protective-response on exclusive breastfeeding vs. 43.24% (16/37) on mixed or formula feeding, OR: 1.148 [CI95% 0.437\u20133.026], p = 0.757. According to age, protective-response decreased significantly as children grow older: 58.33% (28/48) <24 months vs. 26.47% (9/34) \u226524 months, OR: 3.889 [CI95% 1.362\u201311.356], p = 0.004; and specifically 67.65% (23/34) \u22646 months vs. 0%, (0/5) 33\u201341 months, p = 0.008.Children were all HBV-unexposed (AcHBc-negative) and uninfected (HBsAg-negative). Response to anti-HBV vaccine was 80.49% (66/82), with only 45.12% (37/82) developed a protective-response (\u226510IU/L). According to HIV status, 60.71% (17/28) developed a protective-response in group-A, vs. 51.72% (15/29) and 20% (5/25) in group-B and group-C respectively, Odds Ratio (OR): 2.627 [CIAnti-HBV vaccine provides low rate of protection (<50%) among children in general, and particularly if HIV-exposed, infected and/or older children. Implementing policies for early vaccination, specific immunization algorithm for HIV-exposed/infected children, and monitoring vaccine response would ensure effective protection in tropical settings, pending extensive/confirmatory investigations. Viral hepatitis B (HBV) and C are globally known as prevailing agents in hepatocellular carcinoma (HCC) and associated mortality . As the Out of 36.7 million of HIV-infected individuals (including 2.6 million children) worldwide, ~70% are living in SSA, with consistent risks of mother-to-child transmission ,5. FurthIn HBV-endemic settings like SSA, anti-HBV vaccination is strongly recommended to every newborn ,11. Of nIn contrast to the high vaccine-induced HBV-protection rate (90\u201395%) observed among children living in western countries , childreIn a tropical context with highly prevalent HIV/HBV coinfection and consistent MTCT like Cameroon ,7,14, inWe herein sought to ascertain the overall post-vaccination response against HBV in children, compare the degree of HBV protection according to HIV vertical exposure and infection, evaluate the impact of infant feeding option and age on vaccine response.th May\u2013 04th August in 2014 among HBV-vaccinated children at the Essos Health Centre (EHC) in Yaounde, Cameroon; stratified according to HIV profile, feeding option and age range.A prospective, observational and cross-sectional study was conducted from 052 featuring a tropical wet and dry climate with constant temperature (22.9\u201325.7\u00b0C mean daily temperature); the population is over 2.5 million inhabitants, for a density of 14,000/km2. EHC currently has a large accommodation capacity of ~250 beds, innovative technical equipment and a very high-performance within a wide range of specialised services: general medicine, emergency, anaesthesiology, reanimation, surgery and sub-specialities, gynaecology, ophthalmology, dentistry, medical imaging, laboratories (including molecular biology), pharmacy, neonatology, paediatrics, etc. Specifically, EHC is a reference antiretroviral treatment (ART) centre for both adults and children since 2005, with ~450 HIV-infected children on ART to date. This performance ranked the EHC as the four national reference center for paediatric ART in Cameroon.EHC is a reference health facility based in the capital city of Cameroon, a township of 180 kmAt the EHC, PMTCT of HBV at the study site follows national guidelines, and starts with antenatal screening for HBV infection for all pregnant women during antenatal care. For positive-HBsAg mothers, early neonatal administration of anti-HBV immunoglobulin is strongly recommended within 48 hours after birth. Both for HBV vertically exposed and unexposed infants, anti-HBV vaccine (Zilbrix Hepta) is administered  at a series of three monthly doses. Assessing antibody response consists of anti-HBs serological testing recommended at least 1\u20134 months after the final (third) vaccine dose.Based on a convenient sampling, the sample size was calculated using the following formula: (42.9%) , and \u201cD\u201d3) for HIV-infected children. Four children had received the third vaccine dose \u22654 months ago prior to enrolment, while two others had insufficient samples for laboratory analysis. Final sampling therefore had 82 study participants. The HIV status of the 82 participants, as well as the CD4 count of HIV-infected children, was confirmed from their medical records at EHC.At the vaccination unit of the EHC paediatric service, mothers and their children attending the routine immunization program were provided with information on the study objectives. Then, 200 mothers of potentially eligible children received the written information sheet and the study informed consent form. Of these 200 women, 34% (88) mothers returned with a signed informed consent form, and their children were enrolled. Vaccination charts of the 88 children were then checked by retrieving information to confirm eligibility to study criteria, mainly based on age (6\u201359 months), on complete anti-HBV vaccination , and on the absence of advanced stage of immunodeficiency , HIV-exposed but uninfected (group-B), and HIV-exposed and infected (group-C).As per the study secondary outcomes, participants were furthered stratified according to feeding option and age in the frame of anti-HBV vaccine response.Evaluation of post-vaccination response was based on a component of three different HBV-specific biomarkers, among which anti-HBs, anti-HBc and HBsAg tests.0.10 for the mean negative control and of \u22650.80 for the mean positive control. Cut-off value was calculated using the mean negative control x 2.1. Samples were considered Anti HBS positive if D.O \u22651.00, Anti HBS negative or \u201cno vaccine response\u201d if D.O <1.00. \u201cProtective response\u201d was defined as Anti HBS titer >10 UI/ml; \u201clow response\u201d as Anti HBS titer: 1\u20139 UI/ml; and \u201cno response\u201d Anti HBS titer <1 UI/ml.Levels of anti-HBs antibody titer was measured to determine the threshold of effective protection in HBV vaccinated children, using the quantitative anti-HBs Sandwich enzyme-linked immunosorbent assay (ELISA) of CTK Biotech, USA (lot: E0604K3), as per the manufacturer\u2019s instructions. Briefly, reagents were brought at room temperature (18\u201325\u00b0c), wash solutions were prepared with phosphate buffer saline (1/30), and 50 \u03bcl of controls (negative and positive) and samples were added in respective wells. 50 \u03bcl of conjugate was added in each well (except for the blank), followed by incubation at 37\u00b0c for 60 min. Wells were washed and 50 \u03bcl of substrates A and B added in every well (including the blank), then incubated for 30 min at room temperature in a back room. 100\u03bcl of stop solution was then added to each well and reading of optical densities performed at 450 nm wave length, with reference to 630\u2013690 nm. Assay validation was performed with optical density of \u2264 This anti-HBs ELISA kit is coupled to the systematic detection of anti-HBc marker, irrespective of anti-HBs present or absent in the analysed serum .\u00ae, USA), with 50\u03bcl of serum. Validated results were reported as Anti-HBc \u201cresponse\u201d or \u201cno response\u201d. Both Anti-HBc results were all concordant, confirming HBV unexposed status.Anti-HBc marker was used to qualitatively detect any child with past/on-going exposure with HBV. In addition to aforementioned testing of anti-HBc, anti-HBc was also tested using the lateral flow chromatographic immunoassay HBV-5 Rapid Test  of BioSino Biotechnology & Science Inc , with 100\u03bcl of serum. Validated results were reported as HBsAg reactive (HBV-positive) or non-reactive (HBV-negative).S) \u201clow-response\u201d (1\u20139 UI/ml anti-HBS), or \u201cno response\u201d (<1 UI/ml anti-HBS).Vaccine response was defined by a profile of anti-HBs positive, anti-HBc negative and HBsAg negative; further stratified as \u201cprotective-response\u201d  were performed using Chi-square and Fisher Exact test where appropriate, with corresponding 95% CI. P-values, generated by SAS, SPSS, R software, were considered statistically significant if p<0.05.Administrative authorisation (Reference: N\u00b0005/14/DCHE/DA/CE/CHE/CNPS) was issued and ethical clearance for the study obtained from the Institutional Review Board (IRB) of the Essos Health Centre (Reference: N\u00b02014/001/EC-CHE). Mothers of potential participating children received detailed information on the study (with provision of an information sheet). Compliant mothers then provided each a signed written proxy-informed consent; this consent procedure was approved by the IRB. Laboratory results were freely offered for participant clinical benefits, data management was under strict confidentiality by using specific identifiers and restricted access.Study dataset is provided as supporting information .In total, 82 eligible children were enrolled in the study, all being anti-HBc and HBsAg negative. Sex ratio was similar (F/M: 5/4), median  age was 27 [IQR: 9\u201347] months, min-max: 6\u201359 months. According to maternal HBV status, 22 children were born to HBV-negative mothers (of whom 7 were HBV-vaccinated), four from HBV-positive mothers, and 56 from mothers with unknown HBV-status.All HIV-infected children were normal clients  attending the routine immunization program; all were receiving antiretroviral therapy and the majority were asymptomatic without any event of advanced immunodeficiency .According to feeding option of the 82 children during the first six months of life, 54.87% experienced exclusive-breastfeeding, 26.83% formula feeding, and 18.30% mixed feeding. All these children were not exposed to HBV, reported by a negative result both to anti-HBc and HBsAg testing.Overall assessment of anti-HBV post-vaccine response showed that: 80.49% (66/82) of children responded to anti-HBs testing (positive anti-HBsAb), indicating an immune response following anti-HBs vaccination, while 19.51% (16/82) did not responded to vaccination (negative anti-HBsAb).Analysis on the level of protective vaccine response revealed that only 45.12% (37/82) developed a protective immune response against HBV (anti-HBsAb \u226510IU/L) against 35.37% (29/82) who developed a low post-vaccine response.95% 0.933\u20137.5], p = 0.041), as shown in In group-A (HIV unexposed children), 82.14% (23/28) responded to HBV-vaccination and 60.71% (17/28) had a protective response. In the group of HIV-vertically exposed children, 79.63% (43/54) responded to HBV-vaccination and only 37.03% (20/54) had a protective response. Compared to HIV-unexposed children, the level of anti-HBV protection dropped significantly (~two fold) in HIV-exposed children as compared to : 2.627 [CI95%1.099\u201317.602], p = 0.065).Among HIV-exposed/uninfected children, 82.76% (24/29) responded to HBV-vaccination and 51.72% (15/29) had a protective response. Among HIV-exposed/infected children, 76.00% (19/25) responded to HBV-vaccination and only 20.00% (5/25) developed a protective response. Compared to HIV-exposed/uninfected children, the level of anti-HBV protection dropped in HIV-exposed/infected children , as shown in Among HIV-infected children, 76.00% (19/25) responded to HBV-vaccination and only 20.00% (5/25) had a protective response. Among all HIV-uninfected children (exposed or unexposed), 82.46% (47/57) responded to HBV-vaccination and 56.14% (32/57) had a protective response. Compared to HIV-uninfected children, the level of anti-HBV protection dropped significantly (~three fold) in HIV-infected children  on exclusive breastfeeding, against 43.24% (16/37) on mixed (15) or formula feeding (22), OR: 1.079 [CI = 0.757 .95% 1.362\u201311.356], p = 0.004; with specifically 67.65% (23/34) \u22646 months vs. 0%, (0/5) 33\u201341 months, p = 0.008.Evaluation of vaccine efficacy according to the age ranges of children revealed that protective-response decreased from 67.65% (23/34) for age 6\u201314 months, 35.71% (5/14) for age 15\u201323 months, 20% (1/5) for age 24\u201332 months, 0% (0/5) for age 33\u201341 months . Most imFurthermore, among all HIV-exposed children (n = 54), age \u226412 vs. >12 months, respectively, confirmed the decreasing protection with higher age and HIV-infection: uninfected  and infected .In highly endemic HBV and HIV settings like Cameroon, considering paediatric HIV-exposure when monitoring paediatric anti-HBV vaccine response might be of great clinical and public health relevance, and supports optimal performance of the EPI ,7,10\u201312.Though the majority (80.49%) of children were showed a response to anti-HBs testing, a non-negligible rate 19.51%) failed completely, highlighting the necessity for routine evaluation of post-vaccination to either confirm efficacy or determine eligibility for re-vaccination .51% fail. This inHaemophilus influenzae type b, etc) as earlier addressed [et al. in Kenya [Levels of anti-HBV protection dropped significantly for HIV-vertically exposed and/or infected children. This reflects the potential impact of maternal immunodeficiency on the child protectiveness ,9, and cddressed ,20,24. Tddressed ,22,25. Tin Kenya ,22,26.Though feeding options, especially exclusive breastfeeding, had no significant impact on anti-HBV vaccine response (p = 0.757) in our dataset, further investigations with higher sample size are needed on the impact of feeding option on anti-HBV vaccine response. This is because breast-feeding is generally known to positively modulate response to vaccines  through significant developmental changes in Th-1 pipeline response , as compared to formula-fed infants . This isThe quality of response appears to be decreasing, as children grow older, thus supporting the fact that anti-HBV immunization stimulates higher production of Th2-type cytokines and higher levels of antigen-specific type 2 immunoglobulins in newborns than in adults . Though The high rate of non-consenting mothers is the major limitations to our study, as this is directly related to the statistical power in the three arms. Non-consenting were due to (a) believe that vaccination automatically provide the required protection, (b) fear that the child sample could be used for other purpose beside informed research goals, (c) fear of phlebotomy, (d) undisclosed or unknown motivations. These indicate needs for counselling parents on the importance of controlling vaccine efficacy, as well as sensitizing health staff on implementing such measures in routine care, for programmatic effectiveness .Higher sampling would have allowed closer age-match, gender-match, and wider immune status analysis  with respect to anti-HBV vaccine response . Lack ofDespite high response to anti-HBV vaccination among Cameroonian children, protectiveness is poor in general (<50%), worse with HIV-exposure, and even worst with HIV-infection; and a decreasing response as children grow older. Strategies supporting early-age vaccination as well as implementing policies to monitoring post-vaccination response and cost-effective analysis of specific immunization algorithms for HIV vertically exposed- or infected-children would be of high public health benefit in resource-limited settings. These observations therefore merit wider sampling for confirmation towards a stronger policy-implementation.S1 File(ZIP)Click here for additional data file."}
+{"text": "Correction to:Leukemia (2017) 31, 2717\u20132725; doi:10.1038/leu.2017.143; published online 23 May 2017This article was originally published under a CC BY-NC-ND 4.0 license, but has now been made available under a CC BY 4.0 license. The PDF and HTML versions of the paper have been modified accordingly."}
+{"text": "SALL4 and ZNF217 have been widely acknowledged as pivotal effectors stimulating embryonic immortalization as well as oncogenicity. Nevertheless, their prognostic worthiness towards solid tumors remains obscure. Hence we performed this comprehensive meta-analysis aiming to unveil the survival significance of both aberrantly expressed proteins.Overall we included 22 eligible entries comprising of 3093 participants. Over-expression of SALL4 and ZNF217 were negatively correlated with clinical prognosis of 3-year, 5-year, 10-year and disease-free survival in solid malignancies, irrespective of cancer types, source regions, mean-age and sex predominance. Results of sensitivity analysis additionally verified the stability of the pooled outcomes. No publication bias was observed on the basis of Egger's test and Begg's test.Studies were eventually included via database searching and rigorous eligibility appraisal. Data extraction and methodological assessment were implemented under a standard manner. Review Manager 5.3 and STATA 12.0 were utilized as statistical platforms following the recommendations by Cochrane Collaboration protocols.Aberrant amplification of SALL4 and ZNF217 serve as unfavorable predictors of survival expectancy among cancer sufferers, revealing great potential as targeted spots in future therapeutics. Over the past decade, the global healthcare system has been heavily burdened by soaring amount of solid malignancies . Includiin vivo and in vitro, SALL4 was first described as an oncoprotein in leukemia carcinogenesis [SALL4, expressively silenced in mature entities, is constitutively enriched in embryonic tissue and serves to maintain self-renewal capability , 4. Aberogenesis . Similarogenesis . NeverthAs a novel zinc finger transcription factor, ZNF217 was initially identified as a tumorigenic stimulator in breast carcinoma and was frequently amplified in diverse malignancies . HoweverTherefore, as representatives of zinc-finger transcriptional factors with independent performances, we performed this comprehensive meta-analysis aiming to clarify the prognostic roles of SALL4 and ZNF217 in solid malignancies and provide promising targeted spots.Deriving from 742 preliminarily retrieved entries, 22 observational cohorts were eventually selected (13 for SALL4 and 9 for ZNF217), with a total sample-size of 3093 participants (2111 for SALL4 and 982 for ZNF217). The selection flow chart was depicted in Figure n = 5) and liver cancer (n = 9) respectively. The wide spectrum of sample-size ranged from 38 to 337, with a median value of 144. Baseline parameters were statistically comparable between both contrastive groups. Additional details were demonstrated in Table Among the studies featuring SALL4 expression, the chief source region and cancer type were Japan (n = 4). Meanwhile, China (n = 3) became the major source region of literatures, followed by USA (n = 2) and France (n = 2). The median sample-size was 84, with a wide range from 44 to 319    . However, a poorer prognosis of cancer patients was observed concerning 5-year (P = 0.008) and 10-year disease free survival (P = 0.003)  (Other types: P = 0.01). Meanwhile, its unfavorable impact on overall survival was similarly observed among patients with 5-year follow-up duration (Liver cancer: P < 0.00001) (Other types: P = 0.0003)  .P = 0.004) (Mean-age < 60: P < 0.0001) and 5-year overall survival (Mean-age > 60: P = 0.04) (Mean-age < 60: P < 0.00001)  disclosed a prognostic disadvantage among solid cancer patients, irrespective of 3-year (Mean-age > 60: 0.00001) .P < 0.00001) and 5-year survival expectancy (P < 0.00001). Similar outcomes were obtained in non-Asian subgroup (3-year: P < 0.00001) (5-year: P < 0.00001)  .P < 0.0001)  and 5-year overall prognosis    .P = 0.0001)   , 5-year (P = 0.003) and 10-year disease frees survival (P = 0.006)  (Others types: P = 0.02) and 5-year prognosis respectively (Breast cancer: P = 0.01) (Others types: P = 0.0004)  .P = 0.04) and 5-year follow-up duration (P < 0.00001). Nevertheless, there was no significant correlation between ZNF217 overexpression and long-term prognosis among patients with mean-age < 50 (3-year:P = 0.06) (5-year: P = 0.06)  .P = 0.02) (Non-Asian: P = 0.005) and 5-year overall survival in solid malignancies (Asian: P = 0.002) (Non-Asian: P = 0.002)  .P = 0.03)  and 5-year overall survival in solid malignancies    .P < 0.0001) and 5-year (P < 0.00001) overall survival rate, despite that Han et al. [P = 0.0002) and 5-year (P < 0.00001) overall survival in solid malignancies.Firstly, we performed sensitivity analysis by elimination of low-quality trials (NOS = 6). Outcomes remained stable in terms of 3-year  or 5-year overall survival (P < 0.00001).Secondly, we implemented another sensitivity analysis by excluding studies with cytoplasmic staining. Nguyen et al.  and Li eP = 0.125; ZNF217: P = 0.790) and Begg's test  jointly confirmed that there was no publication bias among the included studies in both groups  expression and clinical prognosis in solid malignancies; 3. The expression of SALL4 or ZNF217 was detected by immunohistochemistry (IHC).Studies were ruled out due to the following reasons: 1. Inadequate survival data for further quantification or the follow-up duration was shorter than 3 years; 2. Overlapped or duplicated studies; 3. Inappropriate article types such as reviews and case-reports; 4. Studies involving less than 10 participants as its sample-size.Two investigators independently implemented the selection process and any disagreement was settled by mutual discussion.Standardized extraction forms were applied for data extraction. Concerning the classification standards of SALL4 and ZNF217 expression, we generally recognized low-expression as < 25% of cell positivity while high-expression as > 25% of cell positivity. However, this criterion was adaptively adjusted if necessary, mainly based on original subgroups within individual trials. Two authors collaboratively extracted prognostic materials from main texts or Kaplan-Meier curves.Owing to the observational properties of included studies, a Newcastle-Ottawa Scale (NOS) was employed for methodological evaluation. The scale consisted of three categories including selection, comparability and outcome, with a maximum score of nine. Studies graded with more than six scores were identified as high quality trials in methodology. Each study was appraised by two evaluators in an independent way. Any disapproval was resolved by mutual discussion.I2 as a heterogeneity indicator with its value < 25%, 25%\u201350% and > 50% defined as low, moderate and significant heterogeneity respectively. A moderate or significant heterogeneity was adjusted by a random-effects model, otherwise a fix-effects model was preferred. The statistical significance within all comparisons was mathematically signified as P < 0.05. Moreover, sensitivity analysis was applied to examine the stability of pooled outcomes. Publication bias was statistically analyzed via Egger's test and Begg's test, based on the results from STATA 12.0.Our quantitative calculation was performed on Review Manager 5.3 under Cochrane Collaboration protocols. Odds ratio (OR) along with Mantel-Haenszel model were adopted for dichotomous variables. We acknowledged"}
+{"text": "Visceral adiposity is associated with higher productions of C-reactive protein (CRP) and interleukin-6 (IL-6). Inflammation of obese adipose tissues could contribute to systemic metabolic dysregulation, especially thermogenic activity of white adipose tissues, namely, beige adipogenesis, characterized by altered irisin expression. Thus, we investigated the roles of inflammation and adipocyte beiging in Chinese centrally obese (CO) adults with metabolic syndrome (MetS).  This cross-sectional study was conducted on 54 CO and 58 non-CO subjects drawn from 1492 Chinese people with age and sex matched during November 2010 and August 2013. Twenty (37.0%) of the CO subjects fulfilled the IDF worldwide definition of MetS. Serum CRP, IL-6, and irisin levels were examined.  Higher CRP and IL-6, but lower irisin, levels were manifested in MetS versus non-MetS subjects with or without CO. Multiple linear regression identified high-density lipoprotein cholesterol level as the only independent risk factor for irisin level. Categorized by median of CRP and IL-6 levels, a lower irisin level was only observed in high CRP group.  Under the condition of central obesity, chronic inflammation and impaired beige adipogenesis are associated with MetS in Chinese adults. People with metabolic syndrome (MetS) are at a heightened risk for overt diabetes and cardiovascular events . RelativC-reactive protein (CRP), an established marker of inflammation, is an acute phase protein secreted from liver, playing a key role in the underlying causes of endothelial dysfunction as indicated by decreased nitric oxide and increased endothelin-1, as well as reduced plasminogen activator inhibitor-1 (PAI-1) levels . Moreove\u03b3 coactivator-1 \u03b1/uncoupling protein-1 signaling cascades to elicit beige adipogenesis, muscular mitochondrial biogenesis, and fibre-type switching, resulting in loss of body weight Low level of HDL-C Raised blood glucose This case-matched, cross-sectional study was conducted on 112 subjects drawn from 1492 Hong Kong Chinese people aged from 24 to 86 years with age and sex matched during November 2010 and August 2013 , 28. AllWaist circumference and blood pressure were measured by a trained researcher (Yu AP) . Waist ct- or Mann\u2013Whitney U tests. Univariate and stepwise multiple linear regressions were employed to estimate the correlations of each clinical characteristic with serum CRP, IL-6, and irisin levels. A two-tailed test with P \u2264 0.05 was considered significantly different. All statistical analyses were conducted using SPSS Statistics 24.0.Data were presented as mean\u2009\u00b1\u2009SD (standard deviation) or median (25th and 75th percentiles) for continuous variables and assessed for normality by Shapiro-Wilk test. Either one-way analysis of variance or Kruskal-Wallis test was used for comparing differences across MetS, CO, and non-CO groups. Group differences were compared using either Student's P < 0.01 and P < 0.001), systolic BP (P < 0.001), diastolic BP (P < 0.001), plasma triglyceride (P < 0.001), and fasting blood glucose (P < 0.001), but lower HDL-C (P < 0.001) in MetS compared with non-MetS subjects with or without CO, respectively, whereas there was no significant difference between CO and non-CO, non-MetS subjects, except waist circumference (P < 0.001).The clinical characteristics of our subjects were categorized by the presence of CO in P < 0.01) and non-CO, non-MetS subjects (P < 0.01) (P < 0.05) and non-CO, non-MetS subjects (P < 0.01).There were significantly higher serum levels of CRP and IL-6 in MetS versus CO ( < 0.01) . On the P < 0.001), diastolic BP (P = 0.04), and triglycerides (P < 0.001), but negatively correlated with HDL-C levels (P < 0.001). Serum IL-6 levels were positively associated with waist circumference (P = 0.03), systolic BP (P = 0.008), diastolic BP (P = 0.04), triglycerides (P < 0.001), and fasting plasma glucose (P = 0.002) and also negatively correlated with HDL-C levels (P = 0.002). More importantly, serum irisin levels were negatively correlated with systolic BP (P = 0.04) and fasting plasma glucose (P = 0.03), but positively associated with HDL-C levels (P = 0.001) (P < 0.001) and irisin (P = 0.005) levels, respectively, whereas systolic BP was found to be the only independent risk factor for serum IL-6 levels (P = 0.006).The correlations of clinical characteristics with serum levels of CRP, IL-6, and irisin were analyzed using univariate and stepwise multiple linear regression models. Serum CRP levels were shown to be positively correlated with systolic BP (= 0.001) . StepwisP < 0.001] and IL-6  levels (Tables P < 0.001) and triglycerides (P = 0.01), but lower HDL-C (P = 0.001) and irisin (P = 0.05) levels in high versus low CRP groups (P = 0.003) and fasting plasma glucose (P = 0.009), but lower HDL-C (P = 0.02), were observed , serum irisin levels were also negatively correlated with waist circumference (R = \u22120.15). In addition, stepwise multiple linear regression determined HDL-C as the only independent predictor for serum irisin levels, substantiating circulating irisin as a surrogate marker of improved lipid profile as previously described [Adipose tissue is metabolically active in nature and a highly secretory endocrine organ that is capable of modulating signals in appetite, energy expenditure, immune responses, inflammation, insulin sensitivity, and so forth . In enerescribed \u201333.\u03b2 levels [P = 0.28). This phenomenon could be attributed to dual functions of IL-6 in both pro- and anti-inflammation, and its conflicting roles are largely dependent on (1) its expression levels of which lower extent exerts anti-inflammatory actions and vice versa, (2) different cell contexts as exemplified by its varying roles in hepatocytes, endothelial cells, skeletal muscle, and macrophages, and (3) its selective involvement in either classic or alternative transsignaling, where components of the transsignaling were positively associated with MetS, endothelial dysfunction, and arterial stiffness in male subjects at high risk of cardiovascular disease, and effective blockade of this signaling has been experimentally proven to attenuate innate immunity-triggered inflammatory responses in phenotypically IL-6-transsignaling knockout-like mice [It is known that in obesity, inflamed adipose tissues could cause impaired beige adipogenesis and hence less efficient energy metabolism through complex interactions between macrophages and adipocytes , yet the\u03b2 levels . In contike mice . HoweverThe present study provides a novel insight on how chronic inflammation affects the thermogenic activity of adipose tissue possibly through altered circulating levels of irisin in Chinese adults. Future studies are warranted to further delineate the exact mechanism(s) underlying the interplay among a spectrum of inflammatory mediators and irisin."}
+{"text": "Scientific Reports5: Article number: 16768; 10.1038/srep16768 published online: 11252015; updated: 10202017.This Article contains errors in the legend of Figure 5.A) Systemic zinc chloride  significantly inhibited NaHS-induced scratching. (B) Local application of ZnCl2 (i.d. 5\u2009nmol) significantly inhibited NaHS-induced scratching in both RTX- and vehicle-treated mice. (C) ZnCl2 (i.d. 5\u2009nmol) significantly inhibited NaHS-induced both forelimb wiping and hindpaw scratching in cheek model. (D) ZnCl2(i.pl. 5\u2009nmol) significantly inhibited NaHS-induced flinching. (E) Systemic ascorbic acid  significantly inhibited NaHS-induced scratching. (F) Asc (i.d. 1\u2009nmol) significantly inhibited NaHS-induced scratching in both RTX- and vehicle-treated mice. (G) Asc (i.d. 1\u2009nmol) significantly inhibited NaHS-induced both forelimb wiping and hindpaw scratching in cheek model. (H) Asc (i.pl. 1\u2009nmol) significantly inhibited NaHS-induced flinching. (I) Local application of mibefradil (Mib) (i.d. 5\u201325\u2009nmol) dose-dependently inhibited NaHS-induced scratching in mice. (J) Mib (i.d. 10\u2009nmol) significantly inhibited NaHS-induced scratching in RTX-treated mice. (K) Mib (i.d. 10\u2009nmol) significantly inhibited NaHS-induced both forelimb wiping and hindpaw scratching in cheek model. (L) Mib (i.d. 10\u2009nmol) significantly inhibited NaHS-induced flinching. All data are expressed by means\u2009\u00b1\u2009SEM. n\u2009=\u20096\u20138 mice per group. *P\u2009<\u20090.05; **P\u2009<\u20090.01, ***P\u2009<\u20090.001 vs. vehicle control, Student\u2019s t test\u201d.\u201c(should read:A) Local application of mibefradil (Mib) (i.d. 5\u201325 nmol) dose-dependently inhibited NaHS-induced scratching in mice. (B) Mib (i.d. 10 nmol) significantly inhibited NaHS-induced scratching in RTX-treated mice. (C) Mib (i.d. 10 nmol) significantly inhibited NaHS-induced both forelimb wiping and hindpaw scratching in cheek model. (D) Mib (i.d. 10 nmol) significantly inhibited NaHS-induced flinching. (E) Systemic zinc chloride  significantly inhibited NaHS-induced scratching. (F) Local application of ZnCl2 (i.d. 5 nmol) significantly inhibited NaHS-induced scratching in both RTX- and vehicle-treated mice. (G) ZnCl2 (i.d. 5 nmol) significantly inhibited NaHS-induced both forelimb wiping and hindpaw scratching in cheek model. (H) ZnCl2 (i.pl. 5 nmol) significantly inhibited NaHS-induced flinching. (I) Systemic ascorbic acid  significantly inhibited NaHS-induced scratching. (J) Asc (i.d. 1 nmol) significantly inhibited NaHS-induced scratching in both RTX- and vehicle-treated mice. (K) Asc (i.d. 1 nmol) significantly inhibited NaHS-induced both forelimb wiping and hindpaw scratching in cheek model. (L) Asc (i.pl. 1 nmol) significantly inhibited NaHS-induced flinching. All data are expressed by means\u2009\u00b1\u2009SEM. n\u2009=\u20096\u20138 mice per group. *P\u2009<\u20090.05; **P\u2009<\u20090.01, ***P\u2009<\u20090.001 vs. vehicle control, Student\u2019s t test\u201d.\u201c("}
+{"text": "This E.\u00a0coli is serotype O25b and sequence type 131, a pandemic clonal group, causing worldwide antimicrobial-resistant infections.The draft genome sequence has been determined for an extended-spectrum-\u03b2-lactamase (ESBL)-producing ( Escherichia coli sequence type 131 (ST131) has emerged as a globally dispersed pathogen, the predominant lineage causing extraintestinal infections, and is often resistant to antibiotic treatment . The ESBL-producing E.\u00a0coli strain E10394 (= CCUG 62462) was isolated from a urinary tract infection (UTI) sample as part of an outbreak of ESBL-producing Enterobacteriaceae in a neonatal postsurgery ward at Sahlgrenska University Hospital, Gothenburg, Sweden, in 2008  with command line arguments \u201c\u2014stringency 3\u2014retain_unpaired.\u201d After quality filtering, 1,424,760 paired reads and 2,356/75 unpaired forward/reverse reads were retained for analysis. The high-quality reads were assembled de novo with SPAdes version 3.7.0  -IIa on contig LZET01000076, and aadA5, mph(A), sul1, and dfrA17 on contig LZET01000047. PlasmidFinder (IncFIA (nucleotide accession no. AP001918) on contig LZET01000040, IncFIB (accession no. AP001918) on contig LZET01000042, and IncFII (accession no. AY458016) on contigs LZET01000050, LZET01000068, and LZET01000072. In addition, genes involved in type IV secretion systems (T4SS) were identified, using CONJscan (F), relaxases , and type IV coupling proteins (T4CP). Three small plasmids were completely assembled. First, a 5,627-bp plasmid (contig LZET01000060), which was 99% identical to the pJJ1886_3 plasmid previously described in E.\u00a0coli strain JJ1886 (accession no. CP006787.1), was found. Second, a 5,166-bp plasmid (contig LZET01000063), and last, a 1,718-bp plasmid (contig LZET01000087), which was 99% identical to plasmid pEC648 previously described in E.\u00a0coli strain ST648 (accession no. CP008716.1), were found.The  = CCUG 6462 was iLZET00000000. The version described in this paper is version LZET00000000.1.This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession no."}
+{"text": "Avena L. (Poaceae) and the outgroups were used for maximum likelihood and Bayesian analyses. The evolution of cultivated oat (Avena sativa L.) and its close relatives was inferred to have involved ancient allotetraploidy and subsequent recent allohexaploidy events. The crown ages of two infrageneric lineages (Avena sect. Ventricosa Baum ex Romero-Zarco and Avena sect. Avena) were estimated to be in the early to middle Miocene, and the A. sativa lineages were dated to the late Miocene to Pliocene. These periods coincided with the mild seasonal climatic contrasts and the Mediterranean climate established in the Mediterranean Basin. Our results suggest that polyploidy, lineage divergence, and complex reticulate evolution have occurred in Avena, exemplifying the long-term persistence of tetraploids and the multiple origins of hexaploids related to paleoclimatic oscillations during the Miocene-Pliocene interval in the circum-Mediterranean region. This newly-resolved infrageneric phylogenetic framework represents a major step forward in understanding the origin of the cultivated oat.Understanding the diversification of polyploid crops in the circum-Mediterranean region is a challenging issue in evolutionary biology. Sequence data of three nuclear genes and three plastid DNA fragments from 109 accessions of  Genome duplication following hybridization  is common among flowering plants, and is found in nearly a quarter of Poaceae that provide crops and fuel worldwideAvena L. (Poaceae) contains ca. 29 species exhibiting considerable morphological and ecological diversity in the Mediterranean Basin, Eastern Africa, Europe, Asia, and the Americas4Avena: Avenotrichon (Holub) Baum, Ventricosa Baum ex Romero-Zarco, Agraria Baum, Tenuicarpa Baum, Ethiopica Baum, Pachycarpa Baum, and Avenax\u2009=\u200914), AB- and A\u2019C (DC)-genome tetraploids (4x\u2009=\u200928), to ACD-genome hexaploids (6x\u2009=\u200942)9The genus ds 4x\u2009=\u20092, to ACD-A. clauda DurA. canariensis Baum & Raj. & SampA. insularis Ladiz., A. maroccana Grand., and A. murphyi Ladiz. may contain the D genome found in hexaploid oat11Avena.Molecular data support a close relationship between D and A genomes12A. strigosa Schreb. DNA was homologous to the A-genome sequences of the cultivated oatA. canariensisA. longiglumis DurA. weistii SteudA. damascena Rajah & Baum, A. hirtula Lag., and A. wiestii Steud. rather than from one particular species9812Cultivated oat offers a model for unraveling the dynamic evolutionary process of polyploid crops in the Mediterranean BasinA. sativa21A. clauda)Given chromosome structural differentiation, the C-genome origin of cultivated oat has been under intense scientific scrutiny. Eighteen chromosomes were involved in intergenomic translocations between C and A genomes of Avena with 28 (96.55%) species 3 polyploid cool-season grasses, e.g., fodder ryegrassesThe Mediterranean Basin, encompassing an area between 28\u00b0\u201348\u00b0N and 10\u00b0\u201339\u00b0E, is one of the 34 global biodiversity hotspots with c. 24,000  plant speciesth 28 96.5% specieAvena species  parsimony-informative characters. The maximum likelihood (ML) analyses and the Bayesian inference (BI) showed an identical topology for Avena . Three clades and two nodes were observed in the ppcB1 phylogram: A\u2019C-PPI  , and A- and A\u2019-type sequences hexaploids ]  and hexaploids ] (A. wiestii and A\u2019(D)-type sequences of hexaploids (without A. fatua and A. nuda)] (PP\u2009=\u20090.80) , and A\u2019 and C-type sequences of tetraploids  and hexaploids (without A. nuda)] (PP\u2009=\u20090.54) (Avena (The monophyly of \u2009=\u20090.98) ; node A\u2019erilis)] ; node ABntalis)] ; A-PPI -types of \u2009=\u20091.00) . C-type \u2019C-PPIII . As for in Avena .GBSSI matrix had 1352 characters, including exons 9, 10, 11, 12, 13, and 14, and introns 8, 9, 10, 11, 12, 13, and 14, with the lengths of 53\u2009bp, 81\u2009bp, 194\u2009bp, 88\u2009bp, 129\u2009bp, 22\u2009bp, 47\u2009bp, 148\u2009bp, 153\u2009bp, 127\u2009bp, 154\u2009bp, 152\u2009bp, and 4\u2009bp, respectively (GBSSI data provided 434 (32.1%) parsimony-informative characters. The ML and BI analyses generated different topologies for Avena  ] were observed in the GBSSI tree: A\u2019C-GBI    and hexaploid A. fatua]  , A\u2019-type sequences of A. maroccana and A. murphyi and hexaploids] (Avena (The monophyly of \u2009=\u20091.00)  and 4. I\u2009=\u20091.00) ; AB-GBI \u2009=\u20090.93) ; and AB&aploids] . The cla] ] were observed in the GBSS1 tree: A\u2019C-GBI   , and A\u2019-type sequences of A. maroccana and A. murphyi)] (PP\u2009=\u20090.50)  and hexaploids (without A. nuda and A. sterilis)] (PP\u2009=\u20090.66) (A. sterilis). Clade AB-GBI was sister to clade AB-GBII, and this group (PP\u2009=\u20090.66) plus clade A\u2019C-GBII (PP\u2009=\u20090.50) was assigned to a single monophyletic lineage (PP\u2009=\u20090.98), which was sister to clade A\u2019C-GBI. This large clade received strong support  in Avena -type sequences of tetraploids \u2009=\u20091.00) ; A\u2019C-GBI\u2009=\u20090.50) ; AB-GBI \u2009=\u20090.50) ; and AB-\u2009=\u20090.66) . Clades in Avena .GBSSI sequences were identified in four accessions of A. sativa , consistent with its hexaploid origin. These sequences fell into three distinct groups. In clade A\u2019C-GBI, C-type sequences of A. sativa clustered with C-genome diploids, C-type sequences of tetraploids  and hexaploids (without A. nuda)  , and C]-types of \u2009=\u20091.00) . A-type  AB-GBII , respectanalysis , and conanalysis .gpa1 matrix had 1034 characters, including exons 10, 11, 12, introns 10, 11, and 12; with the lengths of 22\u2009bp, 94\u2009bp, 60\u2009bp, 681\u2009bp, 92\u2009bp, and 85\u2009bp, of which 137 (13.25%) were parsimony-informative. ML and BI analyses had an identical topology for Avena . Seven clades were observed for the gpa1 tree  and five hexaploids (without A. nuda)] ; A\u2019C-GPII  ; A-GPI (A. canariensis and A-type sequence of A. hybrida) ; AB-GPI  ; A\u2019C-GPIII (A. hirtula and A\u2019-type sequences of A. maroccana) ; and AB-GPII  . Clades A-GPII, A-GPI, AB-GPI, A\u2019C-GPIII, and A\u2019C-GPII formed one monophyletic lineage , and this lineage in turn was sister to clade A\u2019C-GPI with strong support , then the large group was sister to clade C-GPI with strong support  (The monophyly of pa1 tree : C-GPI (\u2009=\u20091.00) .gpa1 sequences were identified in a single accession of A. sativa (Liu 310). These sequences fell into two distinct groups, with A\u2019(D)-type sequence of A. sativa nested within clade AB-GPII, and C-type sequences of A. sativa nested within clade A\u2019C-GPI (Two [A\u2019(D)- and C-] types of  A\u2019C-GPI .Avena ; A\u2019C-NRR \u2009+\u2009AB-NRR . Clade A\u2019C-NRR\u2009+\u2009AB-NRR  was sister to clade C-NRR in Avena  were parsimony-informative. The BEAST analysis generated a well-supported tree, which was identical to the topologies obtained from ML and BI analyses of Avena . Two clain Avena . Here weAvena was 20.04 [95% highest posterior density (HPD) 3.56\u201335.06] mya (node 1). This was also the stem ages for clades C-NRR and A\u2019C-NRR\u2009+\u2009AB-NRR, whose crown ages were 10.71 (HPD: 1.62\u201320.25) and 14.54 (HPD: 2.68\u201325.02) mya, respectively (nodes 2 and 3). The crown age of clade A\u2019C-NRR\u2009+\u2009AB-NRR was also the divergence time for nodes A\u2019C-NRR and AB-NRR (nodes 4 and 8). The crown ages of the A. sativa lineages were 2.43, 2.46, and 2.97 mya , respectively (The uncorrelated-rate relaxed molecular clock suggests that the crown age of ectively .Avena were identified by the plastid data: the C-genome diploid lineage (Avena sect. Ventricosa) containing A. clauda, A. eriantha, and A. ventricosa in clade C-NRR; and the A-genome diploid-polyploid lineage (Avena sect. Avena) containing other congeneric species in clade AB-NRR\u2009+\u2009A\u2019C-NRR -genome tetraploids and hexaploids.Two strongly supported infrageneric lineages within \u2009A\u2019C-NRR . Members\u2019C-PPIII  and A\u2019C-\u2019C-PPIII . In the Avena sect. Avena was proposed for the A-genome diploid-polyploid lineage including nodes with low support in the plastid tree , A. damascena (Ad-genome), and A. longiglumis (Al-genome) clustered with three A\u2019C(DC)-genome tetraploids  in node A\u2019C-PPII , A. canariensis (Ac-genome), A. damascena (Ad-genome), and A. prostrata (Ap-genome) clustered with AB-genome tetraploids in node AB-PPI , A. damascena (Ad-genome), and A. longiglumis (Al-genome) clustered with AB-genome tetraploids in clade AB-GBII -genome tetraploids originated from different A-genome diploid ancestors 5gpa1 tree  showed specific genetic divergence from A. hirtula and A. hispanica (group 3) in high-density GBS analysis6) and the genome size (9.08\u2009\u00b1\u20090.1112), A. hirtula can be easily differentiated from the two As-genome diploids, that have a similar genome size to the smallest Ad-genome diploid A. damasceneAvena lusitanica and A. hispanica might represent ecotypes of A. hirtula found in the circum-Mediterranean, western Asia, and EuropeWithin the As-genome diploids, tid tree . Howeverca group  showed sAvena sect. Ventricosa and the C-copy sequences of A\u2019C-genome tetraploids plus hexaploids was a novel discovery which suggested their C-genome donor to be the ancestor of Avena sect. Ventricosa. This was consistent with the hypothesis that the paleotetraploidy events pre-dated and potentially triggered divergence of the extant A\u2019C(DC)-genome tetraploids in narrow ranges of the Mediterranean Basingpa1 tree, A\u2019C(DC)-genome tetraploids together with hexaploids comprised the clade A\u2019C-GPI - and C-genome diploid ancestors to form A\u2019C -genome tetraploids. The second step includes subsequent recent allohexaploidy events involving hybridization between DC-genome tetraploids and the more recent A-genome diploid progenitors to form the extant ACD-genome hexaploids A\u2019C-GPI . TherefoppcB1 tree, i.e., A. atlantica, A. longiglumis, and A. wiestii were embedded within the A\u2019C-PPII-A1, A\u2019C-PPII-A2, and A-PPI subclades  located in node A\u2019C-PPII and clade A-PPI (FL intron2 clones (CIav 9053)A. wiestii is certainly different from other As-genome diploids. Although the different genome forms of A. wiestii were close in genome sizeA wiestii deserves further investigation. Two plausible explanations can be proposed for the ploidy level of allelic variation found in A. wiestii. First, the three diploids may have arisen by allopolyploidy and subsequent unequal diploidization led to heterozygotes. Second, introgression may have brought about very subtle morphological and genetic changes in Avena  and 10.71 (HPD: 1.62\u201320.25) mya, respectively  seem to correlate with highly seasonal climatic oscillation. Geographic isolation might have contributed to genetic differentiation in the progenitor-derivative species pair, the presumed D(or A\u2019)-genome progenitors having disjunct distributions in the Mediterranean region A. wiestii (eastern-most) from A. atlantica (western-most). Therefore, the independent hexaploidy events of cultivated oat were modulated by harsh climatic oscillation, thus A. sativa was able to adapt to new habitats.Cultivated oat may have arisen multiple times in response to selection pressure such as geographic isolation. The long-term aridity of the Mediterranean Basin summer became more severe along a south-eastern to north-western gradient during the late Miocene to PlioceneAvena represents a remarkable model to study because its history of polyploidy, lineage divergence, and complex reticulate evolution. The complex evolution of cultivated oat and its close relatives involved paleotetraploidy events between the ancient A(or A\u2019)- and C-genome diploid ancestors and subsequent recent allohexaploidy events between A\u2019C(DC)-genome tetraploids and the more recent A-genome diploid progenitors. The pattern of recurrent polyploidizations in Avena and their temporal relationships with paleoclimatic oscillations is unparalleled among polyploid crops occurring in the circum-Mediterranean region4AvenaSupplementary Table S1Eighty-nine accessions of 27 species were sampled to represent the morphological diversity and geographic range of six sections in phosphoenolpyruvate carboxylase B1 (ppcB1), granule-bound starch synthase I (GBSSI) and G protein alpha subunit 1 (gpa1), were used. The ppcB1 gene encodes PEPC enzyme for the oxaloacetate replenishment of the tricarboxylic acid cycle in C3 plantsGBSSI gene encodes GBSSI enzyme for the amylose synthesis in plantsgpa1 gene encodes a G-protein \u03b1 subunit for signal transduction in flowering plants247448Three low-copy nuclear genes, et alppcB1, GBSSI, and gpa1) and plastid  fragments, which were amplified using designed or published primers and protocols listed in Table S2Escherichia coli TOP10 competent cells following the protocol of TOPO TA Cloning Kit . The resulting sequences were edited using Sequencher v.5.2.3  and aligned with MUSCLE v.3.8.31http://tree.bio.ed.ac.uk/software/seal/). All sequences were deposited in GenBank .Genomic DNA was extracted following Liu Phylogenetic analyses were performed using maximum likelihoodhttp://tree.bio.ed.ac.uk/software/tracer). All resulting trees were then combined with LogCombiner v.1.6.1 (http://beast.bio.ed.ac.uk/) with 25% burn-ins. The remaining trees  were used to calculate the Bayesian posterior probabilities (PP) for internal nodes. Data sets and phylogenetic trees are available at TreeBase   based on a likelihood ratio testAvena. Three plastid markers were partitioned using BEAUTI v.1.8.2 (within BEAST) with the best fit model determined by Modeltest v.3.7  of three runs was obtained using TreeAnnotator v.1.8.2 (within BEAST) with a PP limit of 0.5 and mean lineage heights. The convergence between two runs was checked using the \u201ccumulative\u201d and \u201ccompare\u201d functions in AWTYHow to cite this article: Liu, Q. et al. Unraveling the evolutionary dynamics of ancient and recent polypoidization events in Avena (Poaceae). Sci. Rep.7, 41944; doi: 10.1038/srep41944 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations."}
+{"text": "The first author is listed incorrectly in the citation. Please view the correct citation here:https://doi.org/10.1371/journal.pone.0186602Guardo AC, G\u00f3mez CE, D\u00edaz-Brito V, Pich J, Arnaiz JA, Perdiguero B, et al. (2017) Safety and vaccine-induced HIV-1 immune responses in healthy volunteers following a late MVA-B boost 4 years after the last immunization. PLoS ONE 12(10): e0186602."}
+{"text": "Leishmania but not found in mammals, is considered a potentially useful target for chemotherapy against leishmaniasis. Leishmania (Viannia) braziliensis is endemic in Latin America and causes American tegumentary leishmaniasis. We demonstrated that IPCs are localized internally in parasites, using a specific monoclonal antibody. Treatment with 5 \u03bcM myriocin  rendered promastigotes 8-fold less infective than controls in experimental hamster infection, as determined by number of parasites per inguinal lymph node after 8 weeks infection, suggesting the importance of parasite IPC or sphingolipid derivatives in parasite infectivity or survival in the host. IPC was isolated from promastigotes of three L. (V.) braziliensis strains and analyzed by positive- and negative-ion ESI-MS. The major IPC ions were characterized as eicosasphinganine and eicosasphingosine. Negative-ion ESI-MS revealed IPC ion species at m/z 778.6 (d20:1/14:0), 780.6 (d20:0/14:0), 796.6 (t20:0/14:0), 806.6 (d20:1/16:0), and 808.6 (d20:0/16:0). IPCs isolated from L. (V.) braziliensis and L. (L.) major showed significant differences in IPC ceramide composition. The major IPC ion from L. (L.) major, detected in negative-ion ESI-MS at m/z 780.6, was composed of ceramide d16:1/18:0. Our results suggest that sphingosine synthase  in L. (V.) braziliensis is responsible for synthesis of a long-chain base of 20 carbons (d20), whereas SPT in L. (L.) major synthesizes a 16-carbon long-chain base (d16). A phylogenetic tree based on SPT proteins was constructed by analysis of sequence homologies in species of the Leishmania and Viannia subgenera. Results indicate that SPT gene position in L. (V.) braziliensis is completely separated from that of members of subgenus Leishmania, including L. (L.) major, L. (L.) infantum, and L. (L.) mexicana. Our findings clearly demonstrate sphingoid base differences between L. (V.) braziliensis and members of subgenus Leishmania, and are relevant to future development of more effective targeted anti-leishmaniasis drugs.Inositol phosphorylceramide (IPC), the major sphingolipid in the genus  Leishmania. The prevalence of leishmaniasis worldwide was recently estimated as 12 million cases, with ~1.5\u20132 million new cases per year  and Leishmania (Viannia), species of which are responsible for various clinical pathologies  and related biological, molecular, and biochemical features  has received considerable research attention during the past three decades because of their biological relevance as structural cell membrane components, and as bioactive compounds involved in cell-cell recognition, cell adhesion, cell growth, and signal transduction , with condensation by serine palmitoyltransferase (SPT) of L-serine and palmitoyl-CoA (16:0) to form 3-ketosphinganine, followed by reduction of this intermediate to produce sphinganine  in a reaction that involves NADPH  that contain two hydroxyl groups, and are denoted by the letter \u201cd\u201d (di) followed by the number of carbons in the chain. SLs in plants and fungi are derived from dihydroxylated or trihydroxylated sphingoid bases, and are denoted respectively by the letter \u201cd\u201d or \u201ct.\u201d Trihydroxylated sphingoid bases in plants and fungi are later acylated to form phytoceramide, a precursor of inositol phosphorylceramides (IPCs) and glycosyl inositol phosphorylceramides (GIPCs)  donovani promastigotes , sphingomyelin (SM), IPCs, and GIPCs  braziliensis promastigotes are present in fractions enriched in membrane microdomains resistant to non-ionic detergent at 4\u00b0C  mexicana  major  braziliensis contain mainly C14:0 fatty acid  braziliensis, one of the major etiologic agents of cutaneous and mucocutaneous leishmaniasis in South America. In cutaneous leishmaniasis, parasites spread from the skin to the naso-oropharyngeal mucosa. The causative factors for such mucosal dissemination and resulting mucosal disease are poorly understood. Effective systemic treatment of cutaneous leishmaniasis caused by L. (V.) braziliensis will presumably reduce the risk of mucosal disease development. We focused on characterization of IPCs from L. (V.) braziliensis mainly because these molecules, and enzymes involved in their specific biosynthetic pathways, are promising targets for anti-Leishmania drug development  tandem MS with electrospray ionization (ESI) in both negative-ion mode [M-H]Stock solutions of 2 mM myriocin (a SL synthesis inhibitor)  were prepared in dimethyl sulfoxide and stored at \u221270\u00b0C for a maximum of 1 month.L. (V.) braziliensis strains MHOM/BR/1987/M11272, MHOM/BR/2001/BA778  and MHOM/BR/1975/M2903 were cultured at 23\u00b0C by several passages of log-phase parasites in Medium 199 supplemented with 10% heat-inactivated fetal calf serum , 2 mM L-glutamine (Sigma-Aldrich), 0.02 mg ml\u22121 bovine hemin, 100 U ml\u22121 penicillin, 100 \u03bcg ml\u22121 streptomycin, and 2% sterile male human urine (complete medium). The starting inoculum (0.4 \u00d7 107 cells ml\u22121) consisted of parasites isolated from early stationary phase. Parasites were collected either in log-phase or after 48 h (stationary phase). L. (V.) major strain MRHO/SU/59/P (LV39) was also cultured at 23\u00b0C in complete Medium 199.Promastigotes of Leishmania (Viannia) braziliensis MHOM/BR/2001/BA778 promastigotes were cultured (starting inoculum 0.4 \u00d7 107 cells ml\u22121) in complete medium in the presence of myriocin (5 \u03bcM) or equivalent vehicle concentration for 6 days. Cell growth was estimated by counting cells with a hemocytometer (Improved Double Neubauer). Parasite viability was determined by SYTOX\u00ae Blue staining  as per the manufacturer's protocol, and shown to be >85% in control and myriocin-treated cultures. Parasite cultures were washed with PBS, cells were resuspended in PBS, and 1 \u00d7 106 parasites (0.05 ml) were inoculated subcutaneously in footpads of female golden hamsters (Mesocricetus auratus) (groups of four). Eight weeks after inoculation (infection), inguinal lymph nodes were removed and homogenized in 3 ml culture medium. Parasite suspension from each node was plated in complete Medium 199, and parasites were quantified by limiting dilution in 96-well plates containing 120 \u03bcl well\u22121. Plates were kept at 23\u00b0C for 7 days. Parasite number (PN) per lymph node was estimated based on the highest dilution at which parasites were detected after 7 days  \u00d7 3 . Experiments were performed in triplicate. All animal procedures were conducted in accordance with the recommendations of the Guide for the Care and Use of Laboratory Animals of the Brazilian National Council of Animal Experimentation (www.cobea.org.br). The protocol was approved by the Research Ethical Committee of Federal University of S\u00e3o Paulo .Leishmania IPC; Godoy et al., MS in prep) and SST-1  .Parasites were fixed with 2% formaldehyde in 10 mM phosphate buffer, pH 7.2, containing 150 mM NaCl (PBS), for 10 min. Some fixed cells were permeabilized with 0.1% saponin. Cells were washed, resuspended in 1 ml PBS, and 100 \u03bcl of the solution was added to coverslips pretreated with 0.1% poly-L-lysine. Coverslips were blocked for 3 h with 0.1% gelatin in PBS and for 1 h with 10% skimmed milk and 1% bovine serum albumin (BSA) in PBS. Parasites were incubated sequentially with primary antibody LST-1   and with chloroform/methanol (CM)   20 psi, and nebulizing gas (N2) 40 psi. Each MS2 individual ion fragmentation was optimized with regard to capillary and collision energy to minimize variations in relative ion abundance resulting from differences in dissociation rates. The CID gas was argon at 2 mTorr. The inlet system consisted of a direct infusion pump (Harvard Apparatus), and methanol/ water  with 5 mM ammonium formate as mobile phase, flow rate 30 \u03bcl min\u22121. Precursor ion scan (PREIS) for detection of ion containing phosphoinositol derivative m/z 259 (inositol monophosphate anion) and m/z 241  was performed in negative ion mode with capillary energy -110 V and collision energy 40 V. Typically, 20 scans were used for accumulation of non-targeted scan range, and 30 scans for analysis of precursor ions. Ion characterization was performed by CID in negative and positive ion mode with collision energy 15, 30, or 45 V  with ESI source. Data acquisition was performed using the Varian MS Workstation program, V. 6.9. Sample analysis was conducted in positive-ion and negative-ion ESI modes with respective needle voltages 5.8 and 5 kV  infantum, L. (L.) donovani, L. (L.) mexicana, L. (L.) major, L. (V.) braziliensis, L. (V.) guyanensis, L. (V.) panamensis, T. brucei, and T. cruzi CL Brener. Sequences were chosen by homology search using BLAST software with annotated yeast and/or human proteins as baits. Sequences were retrieved from GenBank or GeneDB (TriTrypDB) .Phylogenetic analysis was performed using gene data from SPT enzymes from many distinct species of the family Trypanosomatidae: B) Table . ID sequt-test, using GraphPad Prism software program V. 5.0 . Differences between means were considered statistically significant for p < 0.01. All experiments were performed in triplicate.Quantitative data were analyzed by Student's Leishmania surface, parasites were double-labeled with monoclonal antibodies (mAbs) directed to L. (V.) braziliensis glycolipids (mAb SST-1), and to Leishmania IPC (mAb LST-1). Representative images from a z-stack confocal series are shown in Figure To determine whether IPCs are expressed on the  mAb SST-, and to  mAb SST-, and to L. (V.) braziliensis IPC expression, growth rate, and cytokinesis , and at m/z 241, corresponding to inositol-1,2-cyclic phosphate anion (InsP-H2O). ESI-MS of [M-H]\u2212 ions of IPC fractions purified from three L. (V.) braziliensis strains  were performed, and similar spectral profiles were observed for all strains. In full scan of IPC fraction from negative-ion ESI-MS of L. (V.) braziliensis (M11272), major ions were detected at m/z 778.6 and 780.6, and minor components were detected at m/z 796.6, 806.6, and 808.6  major IPC fraction showed a distinctive profile: the major IPC ion was at m/z 778.6, relative abundance of m/z 780.6 was significantly reduced, and minor IPC ions were detected at m/z 796.6 and 806.6  braziliensis, and at m/z 780.6, 782.6, 798.6, and 808.6 for L. (L.) major  braziliensis and L. (L.) major  braziliensis and L. (L.) major  braziliensis  and 241 (InsP-H2O).CID was performed for major IPC ions detected in  Figures  and at ms Figure , phosphoL. (V.) braziliensis [M+H]+ ions at m/z 780.6, using collision energy of 15V, yielded expected fragments at m/z 762 (M-H2O), 520 (M-InsP), and 502 (M-InsP-H2O) . Application of higher collision energy (45 V) for the ion at m/z 780.6 , suggesting that this compound was d20:1/14:0-IPC. Figure L. (V.) braziliensis [M+H]+ ion at m/z 780.6.Ceramide moieties were characterized by CID in positive-ion mode using various collision energies. MS/MS product ion spectra of ) Figure . We also6 Figure , in addiL. (L.) major [M+H]+ ion at m/z 780.6, using collision energies 15 and 30V, gave rise as expected to ions at m/z 762 (M-H2O), 520 (M-InsP), and 502 (M-InsP-H2O)  braziliensis. The ion profile, in terms of LCB and fatty acid composition, was distinct from that of L. (V.) braziliensis. Ions were detected at m/z 236 (indicating the presence of d16:1 sphingoid base), and fragments at m/z 308 and 284 , identifying the 18:0-fatty acyl substituent, as described by Hsu et al.  braziliensis [M+H]+ ion at m/z 782.6 using collision energies 15 and 45 V , 522 (M-InsP), 504 (M-InsP-H2O), 294 , 252, and 228  , indicating that this compound is d20:0/14:0-IPC.ESI-MS/MS spectra of  Figures  generateL. (V.) braziliensis. ESI-MS/MS spectra of [M-H]\u2212 ions at m/z 796.6, 806.6, and 808.6 are shown in Figures m/z 259 (InsP) and 241 (InsP-H2O) were detected in all IPC ions.We also characterized minor IPC ions of + ion at m/z 798.6 allowed us to characterize the ceramide moiety. Using collision energy 45V , 520 (M-InsP-H2O), 328, 310, 292 (characteristic products of phytosphingosine t20:0), 252, and 228  . These findings suggest that this ion (m/z 798.6 in positive-ion mode) corresponds to t20:0/14:0-IPC.Analysis of ESI-MS/MS spectra of the [M+H]V Figure , the spe+ ion at m/z 808.6 using collision energy 45 V , 530 (M-InsP-H2O), 292 (characteristic of sphingoid base d20:1), 280, and 256 (corresponding to fatty acid 16:0 acyl substituent) , suggesting that this compound is d20:1/16:0-IPC.Analysis of ESI-MS/MS spectra of the [M+H]V Figure  generate+ ion at m/z 810.6 using collision energy 45 V , 532 (M-InsP-H2O), 294 (characteristic of d20:0), 280, and 256 (corresponding to fatty acid 16:0 acyl substituent) , suggesting that this compound is d20:0/16:0-IPC.Analysis of ESI-MS/MS spectra of the [M+H]V Figure  generateL. (V.) braziliensis are summarized in Table L. major  braziliensis display predominantly 20-carbon LCBs , and C14:0 and C16:0 fatty acids. Positive-ion ESI-CID-MS/MS of IPC ions from L. (V.) braziliensis did not detect product ions related to d16:0 (m/z 238) or d16:1(m/z 236). Positive-ion ESI-CID-MS/MS of L. (L.) major IPC ion at m/z 780.6 did not detect sphingoid base product related to d20:1 (m/z 292)  braziliensis strains M11272, BA778, and M2903.IPC proposed structures and positive- and negative-ion ESI-CID-MS/MS fragments of IPC ions from ) Figure , Table 2L. (V.) braziliensis is responsible for synthesis of 20-carbon LCB (d20), whereas L. (L.) major presents mainly 16-carbon LCB (d16), and T. brucei and T. cruzi SLs present mainly 18-carbon sphingoid bases (d18)  braziliensis, L. (V.) guyanensis and L. (V.) panamensis) were completely separated from those of the Leishmania subgenus (L. (L.) infantum, L. (L.) mexicana, and L. (L.) major), and comprised two distinct clades. SPTs of T. brucei and T. cruzi also represented different clades.SPT in Leishmania SPTs, we also analyzed N-terminal and catalytic portions of SPTs of L. (V.) braziliensis and L. (L.) major to determine whether sequence homologies refer specifically to the catalytic portion. Phylogenetic results were similar for sequences of SPT whole proteins, N-terminal regions, and catalytic regions  braziliensis  showed that IPCs and GIPLs are not co-localized in L. (V.) braziliensis promastigotes; GIPLs are found on the parasite surface whereas IPCs are localized internally. Labeling of IPCs was observed only when parasites were permeabilized with 0.1% saponin, confirming the internal localization of IPCs. Previous studies have demonstrated the involvement of Leishmania SLs in a variety of biological processes, including differentiation, replication, trafficking, and virulence  braziliensis promastigotes treated with the SPT inhibitor myriocin displayed reduced IPC accumulation and incomplete cytokinesis  braziliensis infectivity. Golden hamsters were infected by subcutaneous footpad injection of promastigotes treated (or not) with 5 \u03bcM myriocin for 6 days. After 8 weeks of infection, PN was counted in inguinal lymph nodes, and found to be ~8-fold lower for myriocin-treated parasites in comparison with controls. Our findings suggest that IPCs (and/or their intermediate SLs) are important for L. (V.) braziliensis infectivity, for their survival during the first few hours inside macrophages, for differentiation to amastigotes, and for amastigote proliferation. We obtained similar results in a previous study of L. (L.) amazonensis mouse experimental infection, in which parasite burden was reduced by infection of mouse footpads with promastigotes pretreated with a different SL inhibitor: the IPC synthase inhibitor aureobasidin A  major displayed defective membrane trafficking events in extracellular promastigotes, thus delaying promastigote infection in BALB/c mice  braziliensis contain mainly C14:0 fatty acid  major and L. (L.) donovani contain mainly C18:0 fatty acids  braziliensis strains, isolated from patients living in different regions of Brazil: reference strain MHOM/BR/1975/M2903 from a subject in Par\u00e1 (Northern Brazil), strain MHOM/BR/01/BA788 from a cutaneous leishmaniasis patient in Bahia State (Northeastern Brazil), and strain MHOM/BR/1987/M11272 from a cutaneous leishmaniasis patient in Paran\u00e1 State (Southern Brazil). IPC molecules were detected by negative-ion ESI-MS, as described by Hsu et al.  braziliensis, and to identify novel IPC structures. The major IPC ions detected by negative-ion ESI-MS were at m/z 778 and 780 for all three strains, regardless of whether ions were isolated at logarithmic or stationary growth phase. These two ions accounted for >70% of IPCs expressed in the parasites. In negative-ion mode, the full scan profile of IPC fractions isolated from the three strains showed ions at m/z 778.6, 780.6, 796.6, 806.6, and 808.6  major and L. (L.) donovani  major  donovani  braziliensis IPC [M+H]+ ions at m/z 780.6 and 782.6 displayed fragments at m/z 292 (20:1-LCB), 294 (20:0-LCB), and 228 (C14:0-fatty acyl substituent). IPC ions with the same mass in positive-ion mode (m/z 780 and 782) from L. (L.) major presented d16:1 and d16:0 LCBs, respectively, with C18:0-fatty acid. IPCs are the major SLs in Leishmania; these findings therefore suggest strongly that L. (V.) braziliensis preferentially synthesizes sphingoid bases through condensation of L-serine and stearoyl-CoA (18:0) to produce d20:0 in LCBs and ceramide, whereas L. (L.) major and L. (L.) donovani (subgenus Leishmania) preferentially synthesize d16:0 sphingoid base through condensation of L-serine and myristoyl-CoA (14:0)  braziliensis showed that the SPT protein sequence of L. (V.) braziliensis is completely separated from those of species in the Leishmania subgenus, suggesting that L. (V.) braziliensis and the Leishmania subgenus species represent different clades. The constructed trees suggest that genetic factors are involved in SPT separation. More precise studies are needed to clarify this issue. The two Leishmania subgenera may have differentiated at different times, as evidenced by the SPT protein data, and this separation may be responsible for observed differences in enzyme functions and specificities. The overall conclusion from the SPT phylogenetic tree is that neither L. major nor L. braziliensis is closer to T. cruzi or T. brucei. It appears that the common SPT ancestor of L. braziliensis and L. major separated from the Trypanosomatidae prior to the separation of Leishmania.In the present study, we investigated the role of SLs in u et al. , who dem6 Figure . PredomiL. (V.) braziliensis vs. L. major, in addition to differences in LCBs, show distinctive fatty acid compositions. In L. (V.) braziliensis IPCs, the major fatty acid is C14:0, and there are only trace amounts of fatty acid 16:0, as detected in positive-ion mode ESI-MS/MS at m/z 808.6 and 810.6. In contrast, the major fatty acid in L. (L.) major and L. (L.) donovani IPCs is stearic acid (C18:0)  braziliensis, preferentially use d20:0 and myristoyl-CoA (14:0) or palmitoyl-CoA (16:0), whereas members of the Leishmania subgenus, e.g., L. (L.) major and L. (L.) donovani, preferentially use d16:0 and stearoyl-CoA (18:0) amino)hexanoyl)sphingosine C6-ceramide.The existence of o et al.  reporteda et al.  showed tLeishmania enzyme specificities in LCB synthesis and concurrent ceramide synthesis will contribute to the development of more effective drugs having high ligand efficiency indices against L. (V.) braziliensis, L. (L.) major, or L. (L.) donovani, but low (or zero) ligand efficiency to mammalian host cells.Future studies on ED and AS conceived and designed the experiments, and wrote the manuscript. ED performed the experiments. ED and MT performed mass spectrometric analysis. RW and DB performed phylogenetic studies. ED and RM performed confocal microscopy and image analysis. HT critically reviewed and revised the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Human leukocyte antigen- (HLA-) A, HLA-B, HLA-C, HLA-DRB1, and HLA-DQB1 allele and haplotype frequencies were studied in a subset of 237 volunteer bone marrow donors registered at the South African Bone Marrow Registry (SABMR). Hapl-o-Mat software was used to compute allele and haplotype frequencies from individuals typed at various resolutions, with some alleles in multiple allele code (MAC) format. Four hundred and thirty-eight HLA-A, 235 HLA-B, 234 HLA-DRB1, 41 HLA-DQB1, and 29 HLA-C alleles are reported. The most frequent alleles were A\u221702:02g (0.096), B\u221707:02g (0.082), C\u221707:02g (0.180), DQB1\u221706:02 (0.157), and DRB1\u221715:01 (0.072). The most common haplotype was A\u221703:01g~B\u221707:02g~C\u221707:02g~DQB1\u221706:02~DRB1\u221715:01 (0.067), which has also been reported in other populations. Deviations from Hardy-Weinberg equilibrium were observed in A, B, and DRB1 loci, with C~DQB1 being the only locus pair in linkage disequilibrium. This study describes allele and haplotype frequencies from a subset of donors registered at SABMR, the only active bone marrow donor registry in Africa. Although the sample size was small, our results form a key resource for future population studies, disease association studies, and donor recruitment strategies. The ~4\u2009Mb human leukocyte antigen (HLA) complex on chromosome 6 in humans is amongst the most polymorphic gene regions in the genome . SeventeBone Marrow Donors Worldwide (BMDW) is a centralized databank of HLA phenotypes and other relevant data of unrelated stem cell donors which aims to support HSCT programmes . The SouDonor registries continuously try to improve their recruitment strategies through increasing donor numbers , recruitIn this study, we describe HLA allele and haplotype frequency data from 237 donors registered with the SABMR, which serves as the source of unrelated marrow donors in South Africa. Frequencies of HLA-A, HLA-B, HLA-C, HLA-DRB1, and HLA-DQB1 alleles and haplotypes were analyzed with the aim of developing a resource for disease association, anthropology, and evolutionary studies. Furthermore, these data will support models for population-specific vaccine development  and willhttps://hml.nmdp.org/MacUI/.Two hundred and thirty-seven (237) SABMR-registered-consenting volunteer bone marrow donors HLA typed at varying resolutions were included in this study. This subset was accessed following an extensive reconsenting procedure of donors in the SAMBR. The self-reported ethnic grouping of the study population was Asian, Black, Chinese, Coloured, White, and some unknown. High-resolution typing has recently been adopted by SABMR, with most donors having low-resolution typing (two digit) , 21 whichttp://www.ebi.ac.uk/ipd/imgt/hla/) [Allele and haplotype  frequencies were estimated by resolving phase and allelic ambiguities using the expectation-maximization (EM) algorithm , 37 in Hgt/hla/) , 3 and rgt/hla/) . Global gt/hla/) . MAC codSelf-reported ethnicity was not considered for analysis in this study owing to redundancy and simplicity of this classification as previously discussed , 42. Onep < 0.05), with HLA-C and HLA-DQB1 having insignificant (p > 0.05) differences between expected and observed heterozygosity (p < 0.001), as summarized in In this donor subset, HLA-A, HLA-B, and HLA-DRB1 genotypes deviated from the expected HWE proportions (zygosity . No signzygosity . In addi\u221702:01g (0.096), A\u221703:01g (0.093), and A\u221701:01g (0.057); B\u221707:02g (0.082), B\u221708:01g (0.049), and B\u221758:02 (0.048); C\u221707:02g (0.180), C\u221707:01g (0.104), and C\u221704:01g (0.091); DRB1\u221715:01 (0.072), DRB1\u221715:03 (0.065), and DRB1\u221707:01 (0.057); and DQB1\u221706:02 (0.157), DQB1\u221703:01 (0.139), and DQB1\u221705:01 (0.118).The full list of alleles including those derived from MACs and their frequencies are listed in \u221733:95~B\u221707:231N (1.08725E-06), A\u221703:01g~C\u221707:02g~DQB1\u221703:02 (1.03519E-06), and A\u221711:01g~C\u221701:02g~DRB1\u221701:01~DQB1\u221705:01 (2.8507E-06) were less frequent two, three, and four locus haplotypes, respectively (\u221703:01g~B\u221707:02g~C\u221707:02g~DRB1\u221715:01~DQB1\u221706:02 being the most frequent (0.067).All two, three, four, and five (extended) haplotype frequencies are detailed in Supplementary ectively . The twehttps://hml.nmdp.org/MacUI) were analyzed using a robust Hapl-o-Mat [Although this study had a limited sample size of 237, we provide an in-depth analysis of HLA diversity in a subset of donors in the SABMR. Mixed resolution HLA typing data with multiple allele codes (pl-o-Mat  package pl-o-Mat . Althougp < 0.001 in p > 0.05), which contrasts to the significant deviation (p < 0.05) observed in the current study  encoded alleles [The main thrust of our study has been the ability to estimate, with high confidence, haplotype frequencies from mixed resolution typings including MAC , despitAlthough results reported here are from a small subset of SABMR registered donors, allele and haplotype frequencies generated by Hapl-o-Mat tool  could be"}
+{"text": "Bacillus velezensis YJ11-1-4 is a strain that exhibits broad-spectrum antimicrobial activity against various pathogens. It was isolated from doenjang, a traditional Korean fermented soybean paste. The genome comprises a single circular chromosome of 4,006,637\u00a0bp with 46.42% G+C content without plasmids. Bacillus velezensis was first proposed by Ruiz-Garc\u00eda et al.  which has revealed good antimicrobial activities against various foodborne pathogens, including Bacillus\u00a0cereus, Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, and Aspergillus flavus subsp. Flavus, was isolated from homemade doenjang, and its complete genome was sequenced.a et al.  as a surdization . Recentldization  suggesteB. velezensis YJ11-1-4 was sequenced using a Pacific Biosciences RS II platform with a 20-kb SMRTbell template library at Macrogen . It generated a total of 969,934,944\u00a0bp (about 198-fold coverage) with 112,837 reads. The sequencing reads were assembled using the Hierarchical Genome Assembly Process version 2.0 (HGAP2.0). The assembled genome of strain YJ11-1-4 comprises a single circular chromosome of 4,006,637\u00a0bp with a 46.42% G+C content without plasmids. The complete genome sequence was annotated using the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html). The tRNA and rRNA genes were detected using tRNAscan-SE  encoded by bacABCDE (AAV30_01545 to AAV30_01565) (acnABCDEF (AAV30_04380 to AAV30_04405) (fenEDCBA (AAV30_10135 to AAV30_10160), bacillomycin D (targeting yeast) encoded by bmyDABC (AAV30_10275 to AAV30_10290), surfactin (targeting virus) encoded by srfABCD (AAV30_17390 to AAV30_17405), and an LCI gene (targeting some Gram-negative bacteria) (AAV30_17550)  . Strain B.\u00a0velezensis YJ11-1-4 was deposited in NCBI under GenBank accession number CP011347.The genome information for"}
+{"text": "The genetic basis for biofilm formation among nontyphoidal salmonellae (NTS) remains poorly understood. This draft genome submission provides initial insights on the genetic differences between biofilm-forming and non-biofilm-forming clinical and environmental NTS serovars. Nontyphoidal salmonellae (NTS) are pathogens of global importance  with varSalmonella enterica serovars , which possess diametrically opposed biofilm phenotypes (biofilm formers and non-biofilm formers) as assessed by the crystal violet binding assay , and S.\u00a0Dublin MN-12 (D15-051537) were isolated from a poultry barn, bovine lymph node, and porcine lung, respectively. The three non-biofilm-forming strains S.\u00a0Heidelberg MN-618, S.\u00a0Typhimurium variant 5 MN-62 (D15-043619), and S.\u00a0Dublin MN-69 (D15-009047) were isolated from a poultry barn, bovine brain, and porcine lung, respectively.The NTS isolates sequenced in this study include pairs of strains from each of three ng assay . The thrde novo assembled by the A5-miseq pipeline (http://rast.nmpdr.org), and PGAP, of the NCBI Prokaryotic Genome Annotation Pipeline (http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html). The occurrence of antibiotic resistance genes, clustered regularly interspaced short palindromic repeat (CRISPR) regions, prophages, Salmonella pathogenicity islands (SPIs), and number of plasmids on assembled sequences were predicted using ResFinder version 2.1 (http://crispr.i2bc.paris-saclay.fr/Server/), PHAST (https://cge.cbs.dtu.dk/services/SPIFinder), and PlasmidFinder version 1.3  and processed on the MiSeq platform . Paired-end reads were pipeline . The asssion 2.1 , CRISPRF), PHAST , SPI Finsion 1.3 , respectsion 1.3 .S.\u00a0Heidelberg MN-714 , S. Typhimurium variant 5 MN-74 (D15-052810) , and S.\u00a0Dublin MN-D12 (D15-051537)  possessed a higher number of plasmids than the non-biofilm-forming strains S.\u00a0Heidelberg MN-618 (ColpVC and IncX1), S.\u00a0Typhimurium variant 5 MN-62 (D15-043619) (no plasmid), and S.\u00a0Dublin D MN-69 (D15-009047) (IncA/C2). Biofilms provide an ideal niche for plasmid exchange and also promotes plasmid stability  relative to non-biofilm formers. A larger genome size would favor increased functional gene content and could enhance the ability of organisms to form biofilms under stressful conditions. It is also noteworthy that the biofilm-forming strains D15-04361 , S.\u00a0Dublin MN-12 (D15-051537), S.\u00a0Heidelberg MN-618, S.\u00a0Typhimurium variant 5 MN-62 (D15-043619), and S.\u00a0Dublin MN-69 (D15-009047) have been deposited in DDBJ/ENA/GenBank under the accession numbers listed in Whole-genome sequences of the six NTS isolates"}
+{"text": "In the article titled \u201cChitosan Prevents Gentamicin-Induced Nephrotoxicity via a Carbonyl Stress-Dependent Pathway\u201d , the namhttps://www.doi.org/10.1016/j.toxlet.2013.01.024. Additionally, the same picture of Figure 2(e) was presented as Figure 2(c). The corrected Also, there was an error in Figure 2. Figures 2(b) and 2(e) were inadvertently reused from Yi-Chieh Li, Yi-Min Shih, and Jen-Ai Lee, \u201cGentamicin caused renal injury deeply related to methylglyoxal and N\u025b-(carboxyethyl)lysine (CEL),\u201d Toxicology Letters, Volume 219, Issue 1,"}
+{"text": "Arthrobacter alpinus R3.8 is a psychrotolerant bacterial strain isolated from a soil sample obtained at Rothera Point, Adelaide Island, close to the Antarctic Peninsula. Strain R3.8 was sequenced in order to help discover potential cold active enzymes with biotechnological applications. Genome analysis identified various cold adaptation genes including some coding for anti-freeze proteins and cold-shock proteins, genes involved in bioremediation of xenobiotic compounds including naphthalene, and genes with chitinolytic and N-acetylglucosamine utilization properties and also plant-growth-influencing properties. In this genome report, we present a complete genome sequence of A. alpinus strain R3.8 and its annotation data, which will facilitate exploitation of potential novel cold-active enzymes.The online version of this article (doi:10.1186/s40793-017-0264-0) contains supplementary material, which is available to authorized users. Antarctica. The optimum growth temperature range of this bacterium is 10\u201316 \u00b0C, which rendered it a promising source for discovery of novel cold-adapted enzymes. The complete genome sequence of 10.1601/nm.20084 strain R3.8 was generated using Single Molecule Real Time sequencing technology to provide a rapid and complete insight into its biotechnological potential. Here, we highlight various genome features that indicate the potential biotechnological value of 10.1601/nm.20084 strain R3.8 in the context of xenobiotic biodegradation and metabolism, chitin utilization, and as a potential component in bio-fertilizers.The production of cold-adapted enzymes by psychrotolerant bacteria has important scientific and industrial interest due to their highly specific activity and catalytic efficiency at low and moderate temperatures . The useArea No.129 . Strain R3.8 was isolated using basal medium supplied with C6-HSL as sole carbon source. An isolation temperature of 4\u00a0\u00b0C was used to select for psychrophilic or psychrotolerant bacteria maintained on Luria Bertani (LB) agar  and urease gamma subunit [AOC05_18490]). Urease is important in catalyzing one of the metabolic pathways involved in microbial-induced calcite precipitation. MICP is a promising approach in the containment of heavy metals such as lead and cadmium in contaminated soils , 24.Various other xenobiotic biodegradation genes and pathways of 10.1601/nm.20084 strain R3.8 are available from the PATRIC server.Chitinase is a biotechnologically-important enzyme widely used in waste management industries for the degradation of chitinous waste into simpler depolymerized substances , in agriN-acetylglucosamine utilization being identified, including beta-hexosaminidase (EC 3.2.1.52) , eukaryotic type N-acetylglucosamine kinase (EC 2.7.1.59) [AOC05_12050], PTS system, N-acetylglucosamine (EC 2.7.1.69) [AOC05_12345], N-acetylglucosamine-6-phosphate deacetylase (EC 3.5.1.25) [AOC05_15120], eukaryotic type N-acetylglucosamine kinase (EC 2.7.1.59) [AOC05_15120], chitinase (EC 3.2.1.14) [AOC05_02965], N-acetyl-glucosamine kinase 2, ROK family (EC 2.7.1.59) [AOC05_03295], transcriptional regulator of N-acetylglucosamine utilization, GntR family [AOC05_07215] and glucosamine-6-phosphate deaminase (EC 3.5.99.6) [AOC05_10045].The full chitinolytic potential of 10.1601/nm.20084 strain R3.8 was also identified here, with various genes involved in chitin and N-(5\u2032-phosphoribosyl) anthranilate isomerase [AOC05_12350] were identified. Production of bacterial IAA is important to assist plants to overcome abiotic stresses and inhibitory compounds, and thus contributes to plant growth stimulation [Application of psychrotrophic PGP bacteria to vegetation can promote growth and improve cold tolerance of crops . From thmulation . SeveralAntarctica. The strain was sequenced to explore its biotechnological potential. By analyzing the complete genome of 10.1601/nm.20084 strain R3, we identified genes involved in xenobiotic biodegradation and metabolism, and chitin utilization, as well as genes that potentially promote plant growth. Further comparative genomic studies with related isolates together with functional studies will provide better understanding of the potential biotechnological value of this strain.We report the complete genome sequence of 10.1601/nm.20084 strain R3.8 that was originally isolated from the soil collected from Rothera Point, Adelaide Island, maritime"}
+{"text": "Delineation of high-risk LTBI can, however, allow for chemoprophylaxis and curtail majority cases of active tuberculosis (ATB). There is epidemiological evidence to support the view that LTBI in context of HIV-1 co-infection is high-risk for progression to ATB relative to LTBI among HIV-ve persons. We recently showed that assays of M.tb thymidylate kinase (TMKmt) antigen and host specific IgG can differentiate ATB from LTBI and or no TB . In this study, we aimed to expose the differential levels of TMKmt Ag among HIV+ve co-infected LTBI relative to HIV-ve LTBI as a strategy to advance these assays for designating incipient LTBI.Precise designation of high risk forms of latent Mycobacterium TMKmt host specific IgM and IgG detection Enzyme Immuno-Assays (EIA) were conducted on 40 TB exposed house-hold contacts (22 LTBI vs. 18 no TB (NTB) by QunatiFERON-TB GOLD\u00ae); and TMKmt Ag detection EIA done on 82 LTBI (46 HIV+ve vs 36 HIV-ve) and 9 NTB (American donors). Purified recombinant TMKmt protein was used as positive control for the Ag assays.IgM levels were found to be equally low across QuantiFERON-TB GOLD\u00ae prequalified NTB and TB exposed house-hold contacts. Higher TMKmt host specific IgG trends were found among TB house-hold contacts relative to NTB controls. TMKmt Ag levels among HIV+ve LTBI were 0.2676 \u00b1 0.0197 (95% CI: 0.2279 to 0.3073) relative to 0.1069 \u00b1 0.01628 (95% CI: 0.07385 to 0.14) for HIV-ve LTBI (supporting incipient nature of LTBI in context of HIV-1 co-infection). NTB had TMKmt Ag levels of 0.1013 \u00b1 0.02505 (5% CI: 0.0421 to 0.1606) (intimating that some were indeed LTBI).TMKmt Ag levels represent a novel surrogate biomarker for high-risk LTBI, while host-specific IgG can be used to designate NTB from LTBI.The online version of this article (10.1186/s12879-018-3007-y) contains supplementary material, which is available to authorized users. Mycobacterium tuberculosis (M. tb)  as per our prior work ) relative to HIV-ve LTBI (<0.14: 0.1069 \u00b1 0.01628[95% CI: 0.07385 to 0.14]) (see Table On one hand:(a) for n=128 and prevalence of 80.0 , the sensitivity, specificity, PPV and NPV of pre-set TMKmt Ag capture-EIA-OD cut-off for differentiating ATB from NTB alone at 95% CI were respectively: 99.0 , 68.0 , 92.7 , and 94.4  [yielding a ROC-area of 83.5 ] compared to 96.6 , 21.1 , 78.9  and 66.7  [ROC-area 58.8 ] obtained using pre-set TMKmt host-specific IgG-EIA-OD cut-offs on n=154 participants and prevalence of 74.0 . On the other hand, (b) for n=220 and prevalence of 63.0 , the sensitivity, specificity, PPV and NPV of pre-set TMKmt Ag capture-EIA-OD cut-off for differentiating ATB from both LTBI & NTB at 95% CI were respectively: 73.9 , 90.2 , 92.7  and 67.3  [ ROC-area of 82.1 ] compared to 92.6 , 34.8 , 78.9  and 64.0  [ROC-area 63.7 ] obtained using pre-set TMKmt host-specific IgG-EIA-OD cut-offs for n=167 participants and prevalence of 72.0   as a surrogate for disease progression. Although several M.tb targets inclusive of lip-arabinomannose (LAM), IP-10, early secretory antigen 6(ESAT-6) and colony filtrate protein 10 (CFP-10) have previously been developed for the purpose of detecting TB, none meets the criteria for designating delineating high-risk LTBI [tb targets [A key limitation of our work\u2014as is the case for all projects that aim to advance novel biomarkers for LTBI, is the absence of a gold standard for precisely designating LTBI and NTB \u201319. As nisk LTBI \u20137. On pa targets , 51. It  targets \u201319.In conclusion, TMKmt Ag and host specific IgG antibodies offer us novel surrogate biomarkers for LTBI in-context of HIV-1 co-infection. Precisely, Levels of TMKmt\u00a0Ag represent a novel surrogate biomarker for high-risk LTBI, while host-specific IgG can be used to designate NTB from LTBI. TMKmt host specific IgM levels might be relevant towards evaluating TB exposure among residents of low TB endemic areas (NTB) who recently travelled to a high TB endemic area.Additional file 1:This file shows processing of the duplicate EIA readings for TMKmt Ag capture among the 46 HIV+ve LTBI and 36 HIV-ve LTBI relative to the 9 NTB controls and 2 rTMKmt positive controls. Overall, readings of blanks were subtracted from the average ODs of the test wells to obtain a corrected average ODs used to draw the graphs. (XLSX 29 kb)"}
+{"text": "Drosophila wing imaginal discs. We used genetic tools to ablate the wing primordium to induce regeneration, and carried out transcriptional profiling of the regeneration blastema by fluorescently labeling and sorting the blastema cells, thus identifying differentially expressed genes. Importantly, by using genetic mutants of several of these differentially expressed genes we have confirmed that they have roles in regeneration. Using this approach, we show that high expression of the gene moladietz (mol), which encodes the Duox-maturation factor NIP, is required during regeneration to produce reactive oxygen species (ROS), which in turn sustain JNK signaling during regeneration. We also show that JNK signaling upregulates mol expression, thereby activating a positive feedback signal that ensures the prolonged JNK activation required for regenerative growth. Thus, by whole-genome transcriptional profiling of regenerating tissue we have identified a positive feedback loop that regulates the extent of regenerative growth.Regenerating tissue must initiate the signaling that drives regenerative growth, and sustain that signaling long enough for regeneration to complete. How these key signals are sustained is unclear. To gain a comprehensive view of the changes in gene expression that occur during regeneration, we performed whole-genome mRNAseq of actively regenerating tissue from damaged  Drosophila imaginal discs, the epithelial structures in the larva that will form the adult animal during metamorphosis, have been an important model system for tissue repair and regeneration for over 60 years. Here we show that damage-induced JNK signaling leads to the upregulation of a gene called moladietz, which encodes a co-factor for an enzyme, NADPH dual oxidase (Duox), that generates reactive oxygen species (ROS), a key tissue-damage signal. High expression of moladietz induces continuous production of ROS in the regenerating tissue. The sustained production of ROS then continues to activate JNK signaling throughout the course of regeneration, ensuring maximal tissue regrowth.Regenerating tissue must initiate the signaling that drives regenerative growth, and then sustain that signaling long enough for regeneration to complete.  The capacity to regenerate damaged or lost organs or limbs is significantly greater in some animals than others. The use of model organisms with varying degrees of regenerative capacity, from whole-body regeneration in planaria and hydra, to limb regeneration in amphibians, organ and fin regeneration in zebrafish, and the limited tissue regeneration that occurs in mammalian models, has advanced our understanding of this process , antz (mol)  , AdoR (F0039747) , and kay0001297)   , Nlaz (FE FBgn00117 activi0003900) , and zfh0004606)  showed u (corto) . Some offeration ,73, and % were dipuckered, can be observed . Vari. VariDroresponse , during response ,85 and aresponse . After acontrols . A seconneration . Additio (Alp-4) , the 4E-4) , an89) /TM6B (gift from M. Uhlirova)[nlaZ:GFP[R2] (gift from M. Ganfornina)[w1118; P{10xStat92E-GFP}1 (BL26197)[cn1P{PZ}dve01738/CyO; ry506 (BL11073)[cn1P{PZ}sm05338/CyO; ry506 (BL11403)[y1w*; P{PTT-GB}LamCCB04957ttvCB04957/SM6a (BL51528)[y1w*; Mi{PT-GFSTF.1}AdoRMI01202-GFSTF.1/TM6C, Sb1Tb1 (BL60165)[y1w*; P{lacW}Thork13517 (BL9558)[y1w*; Mi{PT-GFSTF.0}kayMI05333-GFSTF.0 (BL63175)[w1118; PBac{corto-GFP.FPTB}VK00037 (BL42268), w1118; PBac{Hr78-GFP.FLAG}VK00037 (BL38653), w1118;PBac{NC2\u03b2-GFP.FPTB}VK00033 (BL56157), y1 w*; Mi{PT-GFSTF.0}CatMIO4522-GFSTF.0 (BL60212)[Hml\u0394RFP (gift from K. Bruckner)[TRE-red and gstD-GFP (gifts from D. Bohmann)[Ets21Cf0369 (BL18678)[y1w*; Mi{MIC}CG9336MI03849 (BL36397)[y1w67c23; P{lacW}Col4a1K00405/CyO (BL10479)[y1w67c23; P{lacW}vkgk00236 (BL10473)[P{PZ}Thor06270cn1/CyO; ry506 (BL11481)[w1; P{UAS-Sod1.A}B36 (BL24754), w1; P{UAS-CatA}2 (BL24621), w1; P{UAS-Sod2.M}UM83 (BL24494), w1118; PBac{RB}mole02670/CyO (BL18073)[y1sc*v1; P{TRiP.HMS02560}attP40 (UAS-molRNAi) (BL42867), y1v1; P{TRiP.GL00678}attP40 (UAS-DuoxRNAi) (BL38907), y1v1; P{TRIP.GL00678}attP40 (UAS-NoxRNAi) (BL32902), y1w*; Mi{MIC}NoxMI15634/SM6a (BL61114)[pucE69 [UAS-JNKDN [w* hepr75/FM7C (BL6761)[w*; P{neoFRT}82B p38a1 (BL8822)[Flies were reared on standard molasses medium at 25\u00b0 C except during regeneration experiments. The following (BL37920), y1,w67c(BL10440), y1, w* (BL56274), P{PZ}Al(BL12285), w1118; BL11515), y1w67c(BL12173), ry506PS (BL684), w;;pBACUhlirova), nlaZ:GFnfornina), w1118; (BL26197), cn1P{P(BL11073), cn1P{PBL11403), y1w*; (BL51528), y1w*; (BL60165), y1w*; (BL63175), w1118; (BL60212), Hml\u0394RFP15, y1w603, y1w*(BL36397), y1w67c(BL11481), w1; P{U(BL61114), pucE69 AS-JNKDN , w* hepr (BL6761), w*; P{n (BL8822).Immunostaining was carried out as previously described .Anitbodies and dilutions used were Anti-Nubbin (1:500) (gift of S. Cohen) , mouse aAlexa Fluor (AF) secondary antibodies from Molecular Probes were AF488, AF555 and AF633 (used at 1:500). Nuclei were labeled with DAPI (Sigma)(1:5000).EdU incorporation was carried out using the click-it EdU Alexa Fluor 594 Imaging kit (Molecular Probes) as previously described . SamplesImmunostained samples were imaged on a Zeiss LSM 700 confocal microscope and images were processed using ZenLite, Adobe Photoshop, and Image J software. Bright-field imaging of adult wings was done on an Olympus SZX10 microscope using the CellSens Dimension software, and images were processed using Image J.ROS were detected in imaginal discs using Dihydroethidium (DHE)  using the protocol described in Owusu-Ansah et al. , with slgstD-GFP, whose expression was not uniform and thus was quantified by measuring GFP intensity in the entire pouch area. Average intensities of multiple wing discs were combined to calculate the final average intensity plotted in the graphs. For measuring the pouch area, a maximum projection of all the confocal slices was taken and the Nubbin-expressing area measured in Image J. Graphs were plotted in Excel, R, and GraphPad Prism 7.0.Fluorescence intensity analysis was performed using single confocal slices. Average intensity was calculated by measuring intensity values in three equal-sized boxes in the pouch region of the wing disc in Image J, except for the For imaginal disc measurements and immunofluorescence quantifications, the Welch\u2019s t-test was performed using R and GraphPad Prism 7.0. For the adult wing size assay, chi-squared tests were performed using GraphPad online tools. Statistical analyses for adult wing measurements were performed using Welch\u2019s t-test.nub-GFP/+; rn-Gal4, GAL80ts, UAS-rpr /+, while the mock-ablated controls had the genotype nub-GFP/+; rn-Gal4, GAL80ts/+. Cells were isolated for the transcriptional profile as previously described [The ablated regenerating discs had the genotype escribed . Brieflyescribed . RNA quaDrosophila melanogaster genome  with a maximum of 2 mismatches permitted. Intron length was set between 20 and 150,000, and a gene model was provided as GTF . FPKM estimation was done using Cufflinks (v.0.0.7) [https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE101797).Fastq reads were trimmed using FASTQ Quality Trimmer (v.1.0.0) and adaptor sequences were removed using Clip (v.1.0.1) in Galaxy . Reads wv.0.0.7) , and botv.0.0.7) , geometrv.0.0.7) . All biov.0.0.7) . The datv.0.0.7)  and are S1 Fig(A) Variation in the size of the wing pouch as marked by anti-Nub immunostaining in undamaged and ablated wing discs. Note the minimal variation in pouch size at R0, indicating consistency of ablation. Undamaged n = 8, regenerating n = 28. Error bars are SEM. (B) Quantification of percentage of regenerating animals that had pupariated by each day, showing the asynchronous progression to pupariation that occurred after tissue damage. Undamaged animals were w1118;; rnGAL4, Gal80TS/+, regenerating animals were w1118;; rnGAL4, UAS-reaper, Gal80TS/+, and both experienced the 24-hour temperature shift. Three independent experiments, total undamaged n = 144 pupae, regenerating n = 176 pupae. (C) Percentage of wings of different sizes on w1118;; rnGAL4, UAS-reaper, Gal80TS/+ animals that eclosed on different days after egg laying due the asynchronous development induced by tissue damage. Note that the animals that eclosed first (day 18) had smaller wings than those that eclosed on day 19, and those that eclosed on day 20 had the largest wings. Thus, variation in wing size after disc regeneration is partly determined by length of time for regeneration. Three independent experiments, total n = 371 wings.(TIF)Click here for additional data file.S2 Fig(A) Venn diagrams showing genes at least 1.3-fold upregulated or downregulated in three transcription profiles  of r(TIF)Click here for additional data file.S3 FigAdoR-GFP MiMIC enhancer trap. (B-B\u2019) Kayak-GFP protein trap. (C-C\u2019) Thor-lacZ enhancer trap. (D-D\u2019) Corto-GFP protein trap. (E-E\u2019) NLaz-GFP MiMIC enhancer trap. (F-F\u2019) anti-Twist. (G-G\u2019) zfh1-lacZ enhancer trap. Blue dashed line outlines the wing primordium. Scale bars are 100 \u03bcm. (H) Quantification of upregulation of mol, Nox, and Ets21C expression using qPCR. Four biological replicates each. Error bars are SEM, *p<0.05.Undamaged (A-G) and regenerating (R24) (A\u2019-G\u2019) wing discs. (A-A\u2019) (TIF)Click here for additional data file.S4 Figfru3/+ males had larger wings after regeneration than controls. Three independent experiments, w1118 n = 112 wings, fru3/+ n = 95 wings, p<0.001 by a chi-squared test. (B) Adult htlAB42/+ animals had larger wings after regeneration than controls. Three independent experiments, w1118 n = 316 wings, htlAB42/+ n = 223 wings, p<0.001 by a chi-squared test. Error bars are SEM.(A) Adult (TIF)Click here for additional data file.S5 Fig(A-D) Hemolectin-RFP (Hml-RFP) (green) showing hemocytes near undamaged  and R24  wing discs.  Anti-Nimrod (green) also showing hemocytes near an R24 wing disc, confirming the Hml-RFP results. (B), (D), and (F) are orthogonal slices with the columnar epithelium or the disc proper (DP) toward the bottom and the peripodial epithelium (PE) toward the top of the images. (G) Schematic of an undamaged and a regenerating epithelium showing the location of a hemocyte outside the PE.(TIF)Click here for additional data file.S6 FigmolRNAi caused smaller adult wings after regeneration of the imaginal discs than controls. Three independent experiments, w1118 n = 402 wings, UAS-molRNAi n = 221 wings, p<0.0001 by a chi-squared test. (B-C) Cellular debris is visually distinct from the regenerating epithelium. Regenerating w1118 (B) and mole02670/+ (C) discs at R24, expressing the nub-GFP enhancer trap and stained with DHE. The speckled, grainy GFP tissue is cellular debris, outlined with yellow. The smooth GFP tissue is the intact epithelium, outlined with blue. The side panels are zoomed-in views of debris and epithelium, to show that they are easily distinguished.  Undamaged and R24 w1118 discs stained with DAPI and the ROS detector H2DCFDA. (F-J) gstD-GFP expression in undamaged w1118 (F), and regenerating w1118  and mole02670/+  wing discs at R24  and R48 . (K-O) Anti-Myc immunostaining in w1118  and mole02670/+  regenerating wing discs at R24  and R48 . (O) Quantification of immunofluorescence in Myc staining. R24 w1118 n = 14 discs, mole02670/+ n = 12 discs. R48 w1118 n = 11 discs, mole02670/+ n = 14 discs. Scale bars are 100 \u03bcm. Error bars are SEM except where noted. *p<0.02.(A) Expression of (TIF)Click here for additional data file.S7 Figw1118 and UAS-DuoxRNAi animals. Three independent experiments. w1118 n = 390 wings, UAS-DuoxRNAi n = 200 wings, p = 0.0005 using a chi-squared test. (B) Sizes of adult wings after disc regeneration in w1118 and UAS-NoxRNAi animals. Two independent experiments, thus error bars are SD. w1118 n = 299 wings, UAS-NoxRNAi n = 257 wings, p<0.0001 using a chi-squared test. (C) Sizes of adult wings after disc regeneration in w1118 and NoxMI15634/+ animals. Three independent experiments. w1118 n = 349 wings, NoxMI15634/+ n = 180 wings, p<0.0001 using a chi-squared test.  qPCR showing effectiveness of Duox (D) and Nox (E) RNAi. The RNAi was expressed under rn-GAL4 control in the pouch of normally developing wing discs for 24 hours before collecting for qPCR. Three biological replicates each, *p<0.05. (F) Total number of mitotic cells as identified by anti-phospho-Histone H3 staining in the wing pouch as identified by anti-Nub staining in the indicated genotypes. R0 w1118 n = 14 discs, NoxMI15634/+ n = 15 discs, UAS-NoxRNAi n = 15 discs, R24 w1118 n = 12 discs, NoxMI15634/+ n = 17 discs, UAS-NoxRNAi n = 15 discs, R48 w1118 n = 11 discs, NoxMI15634/+ n = 18 discs, UAS-NoxRNAi n = 19 discs. (G) Area of the wing pouch as marked by anti-Nub staining was measured at R0, R24 and R48 for the indicated genotypes. R0 w1118 n = 14 discs, NoxMI15634/+ n = 15 discs, UAS-NoxRNAi n = 15 discs, R24 w1118 n = 12 discs, NoxMI15634/+ n = 17 discs, UAS-NoxRNAi n = 15 discs, R48 w1118 n = 11 discs, NoxMI15634/+ n = 18 discs, UAS-NoxRNAi n = 19 discs. (H) Pupariation timing for regenerating animals of the indicated gentoypes. Three independent experiments. w1118 n = 204, NoxMI15634/+ n = 113, UAS-NoxRNAi n = 206 (I) Pupariation timing for normally developing animals that did not experience the thermal shift and so did not ablate and regenerate the wing primordia. Note that because these animals did not experience the thermal shift, the pupariation timing cannot be compared directly to the pupariation timing in panel H, in which the animals did experience the thermal shift. w1118 n = 97, NoxMI15634/+ n = 49, UAS-NoxRNAi n = 122. Error bars are SEM.Regeneration assays using adult wing size to assess extent of regenerative growth in the imaginal discs. (A) Sizes of adult wings after disc regeneration in (TIF)Click here for additional data file.S8 Figmol-lacZ reporter (green) in w1118 undamaged discs (A), and in regenerating wing discs at R0 (B), R24 (C) and R48 (D). (E) Quantification of mol-lacZ expression changes. Undamaged n = 3, R0 n = 7, R24 n = 10, R48 n = 10.  Anti-\u03b2-galacosidase immunostaining showing expression of the mol-lacZ reporter (green) in w1118 (F) and p38a1/+ (G) R24 discs. (H) Quantification of the fluorescence from the immunostaining. Two independent experiments, for a total w1118 n = 10 discs, p38a1/+ n = 12 discs. Scale bars are 100 \u03bcm. Error bars are SEM.(A-E) Anti-\u03b2-galacosidase immunostaining showing expression of the (TIF)Click here for additional data file.S1 Table(PDF)Click here for additional data file.S2 Table(PDF)Click here for additional data file.S1 Text(DOCX)Click here for additional data file."}
+{"text": "Staphylococcus aureus , including methicillin-resistant S. aureus (MRSA); streptococci , including Streptococcus pneumoniae, viridans group streptococci, and beta-hemolytic streptococci; Enterobacteriaceae, including Escherichia coli ; Haemophilus influenzae ; and Moraxella catarrhalis . Omadacycline merits further study in serious infections where resistant pathogens may be encountered.Omadacycline was tested against 21,000 bacterial isolates collected prospectively from medical centers in Europe and the United States during 2016. Omadacycline was active against  Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE), Enterobacteriaceae that produce extended-spectrum \u03b2-lactamases (ESBLs) and carbapenemases, and multidrug-resistant (resistant to \u22653 classes of agents) strains of Acinetobacter spp. and Stenotrophomonas maltophilia  were collected prospectively from hospitalized patients in 68 medical centers in the United States and Europe for the 2016 SENTRY Antimicrobial Surveillance Program. Isolate identifications were established by the participating medical centers and confirmed at JMI Laboratories , when necessary. Omadacycline MIC values were determined using the reference Clinical and Laboratory Standards Institute (CLSI) broth microdilution method (Enterobacteriaceae isolates were classified as susceptible (S) to ceftazidime , nonsusceptible (NS) to ceftazidime , R to imipenem , R to tetracycline , and NS to tigecycline . Other resistant phenotypes included MRSA (oxacillin MIC \u2265 4 mg/liter or cefoxitin MIC > 8 mg/liter), vancomycin-NS enterococci (MIC \u2265 8 mg/liter), tetracycline-R A. baumannii, staphylococci, enterococci , and Streptococcus pneumoniae (MIC \u2265 4 mg/liter), macrolide-R S. pneumoniae (erythromycin MIC \u2265 1 mg/liter and azithromycin MIC \u2265 2 mg/liter), and macrolide-R \u03b2-hemolytic streptococci (BHS) (erythromycin MIC \u2265 1 mg/liter). Haemophilus influenzae isolates were divided into \u03b2-lactamase-positive and -negative groups. QC strains were tested concurrently and included Escherichia coli ATCC 25922 and ATCC 35218, S. aureus ATCC 29213, Pseudomonas aeruginosa ATCC 27853, Enterococcus faecalis ATCC 29212, and S. pneumoniae ATCC 49619. All QC results, including all omadacycline MIC values, were within published ranges.Bacterial isolates  isolates were methicillin R, 40.0% of Enterococcus faecium isolates were vancomycin NS, 11.6% of S. pneumoniae isolates were penicillin R, 31.4% of S. pneumoniae isolates were macrolide R, 14.4% of E. coli isolates were ceftazidime NS, 28.6% of Klebsiella pneumoniae isolates were ceftazidime NS, 27.9% of Enterobacter cloacae species complex (SC) isolates were ceftazidime NS, and 8.1% of K. pneumoniae isolates were imipenem R , 43.6% of BHS, 32.8% of Enterobacteriaceae, 66.4% of Acinetobacter baumannii, 0.2% (data not shown) of H. influenzae, and 11.1% (data not shown) of Haemophilus parainfluenzae  , omadacycline inhibited 99.9% of isolates at \u22642 mg/liter, including 100.0% of methicillin-S S. aureus (MSSA) and 99.7% of MRSA isolates . Tetracycline resistance had little effect on omadacycline MIC values against S. aureus  or CoNS  isolates  than against E. faecalis  and was not adversely affected by vancomycin or tetracycline resistance , VGS , and BHS  isolates, regardless of species and susceptibility to penicillin, macrolides, or tetracycline  , indole-positive Proteus spp. , and tigecycline-NS K. pneumoniae    and K. pneumoniae  (50/MIC90 2/4 mg/liter) against ceftazidime-NS  E. cloacae isolates as it was against ceftazidime-S  E. cloacae isolates , K. pneumoniae , Klebsiella oxytoca , E. cloacae SC , and Citrobacter spp.   inhibited 71.2% of isolates at \u22644 mg/liter  at \u22644 mg/liter (data not shown). Omadacycline  demonstrated good in vitro activity against S. maltophilia (Table 2). Omadacycline was equally active against \u03b2-lactamase-negative and -positive isolates of H. influenzae  and was also very active against Moraxella catarrhalis isolates  (Omadacycline MIC distributions are shown in isolates . All CoNsistance  and S1. acycline  and S1. g/liter)  except Pg/liter)  and S2. g/liter)  and S2. ctively)  and S2. isolates  and S2. olates])  and S2. mg/liter . Omadacyg/liter)  and S2.Compared to older tetracyclines, omadacycline has advantages that include a low propensity for selection of resistance, enhanced binding to the 30S ribosomal subunit, the ability to overcome common tetracycline resistance mechanisms, a lack of effect of other resistance mechanisms, availability as intravenous or oral formulations, a prolonged half-life, and once-daily administration . Omadacyin vitro activity of omadacycline against bacterial isolates (United States and Europe) from the 2016 SENTRY survey. Overall, omadacycline provided broad coverage against Gram-positive and fastidious Gram-negative bacteria (S. pneumoniae isolates (E. coli and somewhat less active against ESBL-producing K. pneumoniae and ceftazidime-NS E. cloacae strains. Tetracycline-R Enterobacteriaceae were slightly less susceptible to omadacycline than tetracycline-S strains. Imipenem and amikacin were the most active agents against Enterobacteriaceae, including the resistant subsets. Omadacycline was the only agent with useful activity against A. baumannii, and omadacycline and trimethoprim-sulfamethoxazole were the only agents with useful activity against S. maltophilia isolates.This study documents the isolates  and S1. These data build on previous SENTRY surveillance surveys  and indi"}
+{"text": "However, IDH mutations make classification of gliomas according to the WHO2007 edition controversial. Here, we characterized IDH-1R132H mut status in a cohort of 670 adult patients with different WHO2007 grades of diffuse glioma. Patient characteristics, clinical data and prognoses were obtained from medical records. Patients with IDH-1R132H mut were younger and had better clinical outcomes than those without mutations. Differences in age among patients with astrocytomas of different WHO2007 grades were eliminated after patients were grouped based on IDH-1R132H status. IDH-1R132H mut was present more often in patients with lower Ki-67 and MGMT protein levels and higher mutant p53 levels. Ki-67 was also strongly associated with WHO2007 grade independently of IDH-1R132H mut status. Moreover, patients with Ki-67<30 survived longer than those with Ki-67\u226530, regardless of IDH-1R132H mut status. Patients in the IDH-1R132H mut group with lower MGMT protein levels also had better clinical outcomes than those in other groups. Our results indicate that to better treat gliomas, IDH mutation status should be included when determining WHO2007 grade in glioma patients.WHO The median OS of 36.230 months (95% CI 31.103\u201341.357) in patients with Ki-67<30 was higher than the OS of 15.370 months (95% CI 12.842\u201317.898) in patients with Ki-67\u226530 . We subdivided IDH-1R132H mut or IDH-1R132H wt gliomas based on Ki-67<30, and found differences in mOS among the four groups  vs. 19.000 months (95% CI 13.754\u201324.246); p<0.001, Breslow test, Figure R132H wt [median 17.400 months (95% CI 13.627\u201321.173) vs. 13.270 months (95% CI 11.141\u201315.399); p=0.039, Breslow test, Figure R132H mut/Ki-67\u226530 and IDH-1R132H wt /Ki-67<30 patients .IDH-1s Figure . Patientp<0.001, Breslow test). Furthermore, mOS differed among IDH-1R132H-mut-MGMTneg , IDH-1R132H mut-MGMTpos , IDH-1R132H wt-MGMTneg  and IDH-1R132H-WT-MGMTpos  patients .The mOS of 15.670 months (95% CI 13.273\u201318.067) in MGMT-positive patients was shorter than the mOS of 32.500 months (95% CI 22.439\u201342.561) observed in MGMT-negative patients  and IDH wt (median OS=355 days) groups. The median OS in our study was closer to that found by Zeng et al. These results indicate that Ki-67 index is a reliable candidate for determining prognosis in glioma patients in addition to IDH-1 status. Although the prognostic value of MGMT protein levels is controversial [Recent data suggests that IDH-1 mutations, Ki-67 index, and MGMT protein levels are prognostic factors for diffuse gliomas , 30\u201332. oversial , we founR132H mut in a large cohort of glioma patients. IDH-1R132H mut was associated with specific WHO2007 histological grades and younger age. Age differences between different WHO2007 grades of astrocytoma were strongly influenced by IDH-1R132H mutation status. Low Ki-67 index values occurred much more often in patients with lower WHO2007 grades and IDH-1R132H mutation. Finally, our study indicated that Ki-67 index and MGMT protein levels, together with IDH mutation status, were predictive of prognosis in different glioma subtypes.In summary, we characterized the expression of IDH-1Tumor samples were obtained from Sanbo Brain Hospital. Informed consent was obtained from all patients prior to the study. All experiments using human tissues were approved by the Institutional Review Board of Sanbo Brain Hospital. 670 adult patients with diffuse supratentorial gliomas were involved in the study. WHO classification of all specimens was performed by two independent neuropathologists . In the R132H (Dianova 1:100), p53 (1:100 Invitrogen), MGMT (1:150 Invitrogen), and Ki-67 (1:200 Invitrogen) were used. The cutoff values were 10% for IDH-1R132H mut, 10% for mutant p53, 10% for MGMT, and 30% for Ki-67. Representative images of high and low IDH-1R132H mut  was considered statistically significant.SPSS 22.0 was used for all statistical analyses. The \u03c7"}
+{"text": "Nature Communications8:312 doi:10.1038/s41467-017-00390-1; Article pubilshed online: 22 Aug 2017Nat. Commun.7, 12374 (2016).\u201dWe regretfully omitted to give credit to a previous figure upon which the surface-tension scheme in Fig. 1b is based. The caption to Fig. 1 should have included the following: \u201cThe surface-tension scheme in Fig. 1b is adapted from Fig. 1a in Noh, J., Jeong, S. & Lee, J.-Y. Ultrafast formation of air-processable and high-quality polymer films on an aqueous substrate."}
+{"text": "Xiphinema includes a remarkable group of invertebrates of the phylum Nematoda comprising ectoparasitic animals of many wild and cultivated plants. Damage is caused by direct feeding on root cells and by vectoring nepoviruses that cause diseases on several crops. Precise identification of Xiphinema species is critical for launching appropriate control measures. We make available the first detailed information on the diversity and distribution of Xiphinema species infesting wild and cultivated olive in a wide-region in southern Spain that included 211 locations from which 453 sampling sites were analyzed. The present study identified thirty-two Xiphinema spp. in the rhizosphere of olive trees, ten species belonging to Xiphinema americanum-group, whereas twenty-two were attributed to Xiphinema non-americanum-group. These results increase our current knowledge on the biodiversity of Xiphinema species identified in olives and include the description of four new species , and two new records for cultivate olives . We also found evidence of remarkable prevalence of Xiphinema spp. in olive trees, viz. 85.0% (385 out of 453 sampling sites), and they were widely distributed in both wild and cultivated olives, with 26 and 17 Xiphinema spp., respectively. Diversity indexes  were significantly affected by olive type. We also developed a comparative morphological and morphometrical study together with molecular data from three nuclear ribosomal RNA genes . Molecular characterization and phylogenetic analyses allowed the delimitation and discrimination of four new species of the genus described herein and three known species. Phylogenetic analyses of Xiphinema spp. resulted in a general consensus of these species groups. This study is the most complete phylogenetic analysis for Xiphinema non-americanum-group species to date.The genus  Soil is most likely one of the more species-rich habitats of terrestrial ecosystems because over one quarter of all living species on Earth are inhabiting the soil , 2. One Meloidogyne spp.) and cyst nematodes (Heterodera spp. and Globodera spp.), likewise the ectoparasitic nematodes belonging to the family Longidoridae Thorne, 1935 [Xiphinema Cobb, 1913 [Nepovirus, family Comoviridae) that cause significant damage to a wide range of crops [Xiphinema was divided into two different species groups [Xiphinema americanum-group comprising a complex of about 60 species [Xiphinema non-americanum-group which comprises a complex of more than 215 species [Xiphinema is based mainly on classical diagnostic features; however, due to a high degree of intraspecific morphometric variability can lead to overlapping among Xiphinema species and increase the risk of species miss-identification [The most important nematodes economically include endoparasitic species such as the root-knot  species , 17, 18.fication .Xiphinema species  have been characterized molecularly by ribosomal genes , constituting a useful tool for molecular-based species identification [Xiphinema species identification becomes difficult when dealing with morphological closely species that co-occur in a sample or region, as often detected in the Iberian Peninsula [Xiphinema spp. detected in the Iberian Peninsula [Xiphinema have been reported in Spain, mainly associated with woody, ornamental and vegetable plant species [Recently, 96 fication \u201333. Xipheninsula , 28. Seveninsula \u201336. In p species , 38.Xiphinema spp.) [Xiphinema spp. associated with olive trees, except for the recent contributions of Archidona-Yuste et al. [Xiphinema macrodora Archidona-Yuste et al., 2016, Xiphinema oleae Archidona-Yuste et al., 2016, Xiphinema plesiopachtaicum Archidona-Yuste et al. 2016, and Xiphinema vallense Archidona-Yuste et al. 2016 [Xiphinema spp. infecting wild and cultivated olives in southern Spain, we surveyed a total of 211 localities at the eight provinces of Andalusia where both olive forms were present. This survey raised 385 populations of Xiphinema species, apparently morphologically related to other known Xiphinema spp. This prompted us to carry out an integrative taxonomic study to identify the species within this complex genus.Olive, in wild and cultivated forms, is widely distributed in the Mediterranean Basin, and particularly in southern Spain , 39\u201341. ma spp.) , 42. Howe et al. , 18, 28 al. 2016 , 18, 28.Xiphinema species and to test the resemblance between morphological and molecular data within Xiphinema species, and the specific objectives were: i) to identify the 385 Spanish populations of Xiphinema spp. detected in wild and cultivate olives; ii) to carry out a molecular characterisation of these Xiphinema populations based on sequences of the D2-D3 expansion segments of the 28S nuclear ribosomal RNA gene, the ITS1 of rRNA, and partial 18S rRNA sequences; and iii) to study the phylogenetic relationships of Xiphinema spp.The general objectives of this research was to study the occurrence and abundance of No specific permits were required for the described fieldwork studies. Permission for sampling the olive orchards was granted by the landowner. The samples from wild olives were obtained in public areas, forests, and other natural areas studied and do not involve any species endangered or protected in Spain. The sites are not protected in any way.et al. [Nematodes were surveyed from 2012 to 2015 during the spring season in wild and cultivate olives growing in Andalusia, southern Spain , Fig 1. et al. .3 sub-sample of soil by a modification of Cobb\u00b4s decanting and sieving method [et al. [Xiphinema.Nematodes were extracted from a 500-cmg method . Since rg method , 44, theg method . The nem [et al. . PPN froXiphinema spp. populations detected infesting soils from olives in Andalusia, conventional ecological and diversity indexes were performed in order to evaluate the distribution and changes in the diversity in wild and cultivated olives. In this regard, abundance and prevalence of each Xiphinema species identified were estimated. For each sampling site, abundance was calculated as the mean number of Xiphinema nematodes per 500 cm3 of soil for all samples. The prevalence was computed by dividing the number of samples in which a Xiphinema species was detected by the total number of samples and expressed as a percentage.Based on the P \u2264 0.05 using the general model procedure of SAS .Several diversity indexes including Hill\u00b4s diversity, Hill\u00b4s reciprocal of D (Simpson\u00b4s dominance index) and Hill\u00b4s evenness indexes  were calXiphinema specimens for light microscopy were killed by gentle heat, fixed and examined Xiphinema specimens as described by Archidona-Yuste et al. and Seinhorst [viz. Xiphinema cadavalense Bravo & Roca 1995 [viz. Xiphinema conurum Siddiqi, 1964 [Xiphinema species already described were analysed morphologically and molecularly in this study and proposed as standard and reference populations for each species given until topotype material becomes available and molecularly characterized. Voucher specimens of these described species have been deposited in the nematode collection of Institute for Sustainable Agriculture, IAS-CSIC, C\u00f3rdoba, Spain.einhorst , 50. Theeinhorst , 27, 51.oca 1995 , and Dr.qi, 1964 . NematodXiphinema. Following morphological confirmation, the specimens were removed from the slides and DNA extracted.For molecular analyses, in order to avoid mistakes in the case of mixed populations, two live nematodes from each sample were temporary mounted in a drop of 1M NaCl containing glass beads (to avoid nematode crushing/damaging specimens) to ensure specimens conformed to the unidentified populations of et al. [et al. [Detailed protocols for nematode DNA extraction, PCR and sequencing were applied as described by Castillo et al. . The D2-et al. , 55\u201358.  [et al. . The newXiphinema spp. from GenBank were used for phylogenetic reconstruction. Outgroup taxa for each dataset were chosen according to previous published data [http://molevol.cmima.csic.es/castresana/Gblocks_server.html) using the less stringent option . Percentage similarity between sequences was calculated using a sequence identity matrix in BioEdit. For that, the score for each pair of sequences was compared directly and all gap or place-holding characters were treated as a gap. When position of both sequences has a gap they do not contribute as a difference. Phylogenetic analyses of the sequence data sets were performed based on Bayesian inference (BI) using MrBayes 3.1.2 [6 generations, respectively. The Markov chains were sampled at intervals of 100 generations. Two runs were performed for each analysis. After discarding burn-in samples and evaluating convergence, the remaining samples were retained for further analyses. The topologies were used to generate a 50% majority rule consensus tree. Posterior probabilities (PP) are given on appropriate clades. Trees were visualised using TreeView [D2-D3 expansion segments of 28S rRNA, ITS1, and partial 18S rRNA sequences of different hed data , 17, 18.hed data , strateghed data  and edithed data  in Castres 3.1.2 . The beses 3.1.2  with theTreeView .http://zoobank.org/. The LSID for this publication is: urn:lsid:zoobank.org:pub:CE945C7D-7B14-46DD-8A17-A93A05750590. The electronic edition of this work was published in a journal with an ISSN, and has been archived and is available from the following digital repositories: PubMed Central, LOCKSS.The electronic edition of this article conforms to the requirements of the amended International Code of Zoological Nomenclature (ICZN), and hence the new names contained herein are available under that Code from the electronic edition. This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the ICZN. The ZooBank LSIDs (Life Science Identifiers) can be resolved and the associated information viewed through any standard web browser by appending the LSID to the prefix Xiphinema spp. found in this study including specimens of sampling sites used in morphological and/or molecular analyses, are shown in Xiphinema spp. and sampling sites are presented in Xiphinema spp. were detected in the rhizosphere of olive trees, ten species belonging to X. americanum-group, whereas 22 were attributed to X. non-americanum-group  Kirjanova 1951 [X. parachydermum Guti\u00e9rrez-Guti\u00e9rrez et al., 2012 [X. turcicum Luc, 1963 [X. vallense), while the remaining 21 identified species where present either in wild or cultivated olives only.All um-group . From aled olive ; Fig 1. l., 1992 , X. coxian, 1964 , X. duril., 1993 , X. incel., 1983 , X. italyl, 1953 , X. macrl., 1992 , X. pachova 1951 , 71, X. l., 2012 , X. turcuc, 1963  and X. vXiphinema spp. were present in low to high densities  in both wild and cultivated olives, being observed in cultivated olives in higher densities than in wild olives (X. americanum-group species was significantly higher (P < 0.01) in cultivated than wild olives  in both olive types in the Xiphinema non-americanum-group  (P < 0.001) for X. americanum group than X. non-americanum-group species (Xiphinema species with the highest nematode density was X. pachtaicum (414 nematodes per 500 cm3 of soil), which showed a higher average density in cultivated than wild olives , both belonging to X. non-americanum-group, showing lower average density in cultivated than in wild olives  , which r species . On the d olives . Howeverd olives .Xiphinema spp. in olive was 85.0% (385 out of 453 sample sites) in Andalusia  than cultivated olives    that was detected in both olive types in the most of the Andalusia provinces, at exception of Almer\u00eda , that was found in both olive types in Almer\u00eda, C\u00e1diz, Huelva and M\u00e1laga provinces, but only in wild olive in C\u00f3rdoba, Granada, Ja\u00e9n and Seville provinces  . In addive types . As indie sites) . The sub Almer\u00eda . Anotherrovinces .Xiphinema spp. in wild and cultivated olive (Xiphinema spp. detected in each sampling site (Richness index) was significantly affected (P < 0.05) by olive type (P < 0.001) in wild than cultivated olives (X. non-americanum-group species were significantly higher (P < 0.05) in wild than in cultivated olive (X. americanum-group species (P < 0.001) in X. americanum-group than in X. non-americanum-group (P < 0.05) for both diversity indexes were observed when Xiphinema species groups were considered separately  than X. americanum-group (Tables P < 0.01) among them when it was considered both olive types (P < 0.01) than that of wild olives (P < 0.05) detected in cultivated than wild olives (X. americanum-group was significantly higher (P < 0.001) than that of X. non-americanum-group species , and Hill\u00b4s evenness ), and teed olive . Overallive type , showingd olives . Similared olive , but the species . Overallum-group . Diversium-group . Thus, sparately . On the  P < 0.00 in wild p Tables  resultedve types . Evennesd olives  accordind olives . On the  species .Nematoda Linnaeus, 1758 Dorylaimida Pearse, 1942 Longidoridae Thorne, 1935 [Longidorinae Thorne, 1935 [Xiphinema Cobb, 1913 [bb, 1913 Xiphinema andalusiense Archidona-Yuste, Navas-Cort\u00e9s, Cantalapiedra-Navarrete, Palomares-Rius & Castillo, sp. nov.urn:lsid:zoobank.org:act:95E9BE47-B822-4AAF-A11C-50EF7A016137Figs 5Olea europaea subsp. silvestris (Miller) Lehr) , at Belmez, C\u00f3rdoba province, Spain; collected by G. Leon Ropero, March 14, 2015; mounted in pure glycerine and deposited in the nematode collection at Institute for Sustainable Agriculture (IAS) of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection number AR093-2).Adult female, collected from the rhizosphere of wild olive  of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection numbers AR093-5-AR093-7); two females at Istituto per la Protezione Sostenibile delle Piante (IPSP), Consiglio Nazionale delle Ricerche (CNR), Bari, Italy (AR093-8); and one female at USDA Nematode Collection, Beltsville, MD, USA (T-6774p); collected by G. Leon Ropero, March 14, 2015.Xiphinema andalusiense sp. nov. is an apparently parthenogenetic species belonging to morphospecies Group 5 from the Xiphinema non-americanum-group species [pars dilatata uteri, and small spiniform structures and crystalloid bodies in low number; female tail short, convex-conoid to conical shape with distinctly digitate terminus, and bearing three pairs of caudal pores; c\u00b4 ratio (1.0\u20131.3); and specific D2-D3, ITS1 rRNA and partial 18S rRNA sequences . According to the polytomous key of Loof & Luc [ species . It is cof & Luc , the newThe species epithet refers to the autonomous community from Spain, Andalusia, where the species was detected and moderately distributed.ca 76\u201388% of lip region diam. and located at ca two-thirds of lip region height. Odontostyle long, 1.6\u20131.9 times longer than odontophore, and the latter with moderate-developed flanges 9.5\u201312.5 \u03bcm wide. Guiding ring with average guiding sheath length of 16.0 \u03bcm. Pharynx occupying about 8\u201315% of body length, consisting of an anterior slender narrow part 346\u2013541 \u03bcm long and extending to terminal pharyngeal bulb occupying ca 19\u201327% of total pharyngeal length, 112\u2013139 \u03bcm long and 22.5\u201329.5 \u03bcm wide. Glandularium 99.5\u2013119.0 \u03bcm long. Nucleus of dorsal pharyngeal gland (DN) located at beginning of basal bulb (10.4\u201314.3%), ventrosublateral nuclei (SVN) situated ca halfway along bulb (46.9\u201359.4%)  similar to that of female, including almost conoid tail shape ending in a digitate terminus  were identified using morphological characters such as body length, length of replacement and functional odontostyle , Fig 5 , 76. Speterminus , becominXiphinema andalusiense sp. nov. was collected from the rhizosphere of wild olive (Olea europaea subsp. silvestris (Miller) Lehr) of two localities belonging to C\u00f3rdoba and Ja\u00e9n provinces, being one of the new species described in this work which has a broader distribution in Andalusia, concretely on North of Andalusia , slightly lower c\u00b4 ratio (1.0\u20131.3 vs 1.1\u20131.8), the presence of spiniform structures or crystalloid bodies along tubular portion of uterus vs absent, and the absence vs presence of males [X. andalusiense sp. nov. mainly differs from X. cadavalense in having a shorter odontostyle and odontophore length  resulting in a shorter stylet length (215.5\u2013239.5 vs 244.5\u2013278.5 \u03bcm), a narrower lip region (12.0\u201315.5 vs 14.0\u201319.5 \u03bcm), and higher a and c\u00b4 ratios  [X. andalusiense sp. nov. differs from X. turdetanense in having a slightly longer odontostyle length (137.0\u2013151.0 vs 121.0\u2013142.0 \u03bcm), a slightly narrower lip region (11.5\u201315.5 vs 14.0\u201316.0 \u03bcm), higher number of globular bodies present in the Z-differentiation (11\u201316 vs 6\u20138), size and number of spiniform structures presents along tubular part of uterus , presence of crystalloid bodies along uterus vs absence, and the absence vs presence of males [According to the polytomous key by Loof & Luc  and sortl., 2013 , X. cadal., 2013 . Xiphineof males . On the ctively) . Finallyof males .X. andalusiense sp. nov. is molecularly related to X. macrodora, but it can be clearly differentiated in having a smaller nematode body and odontostyle length  [In addition, ctively) .X. andalusiense sp. nov. (KX244884-KX244888) was 97% similar to X. baetica , X. macrodora  and X. cadavalense (KX244900); sequence variation among these species was from 24 to 34 nucleotides and from 3 to 8 indels  in relation to the ITS1 region were also X. baetica , X. cadavalense , and X. macrodora . Intraspecific variation for this marker was 44 nucleotides and 23 gaps amongst the five studied populations , with several Xiphinema spp. such as X. baetica (KC567148-KC567149), X. cadavalense (KX244932), X. macrodora (KU171050) and X. coxi europaeum (KC567153).D2-D3 region of 8 indels . Xiphineulations . FinallyXiphinema celtiense Archidona-Yuste, Navas-Cort\u00e9s, Cantalapiedra-Navarrete, Palomares-Rius & Castillo, sp. nov.urn:lsid:zoobank.org:act:17E565E4-18E8-4D60-AA57-55253F3E257EFigs 7Olea europaea subsp. silvestris (Miller) Lehr) , at Pe\u00f1aflor, Seville province, Spain; collected by A. Archidona-Yuste, April 22, 2014; mounted in pure glycerine and deposited in the nematode collection at Institute for Sustainable Agriculture (IAS) of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection number AR083-01).Adult female, collected from the rhizosphere of wild olive  of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection numbers AR083-03-AR083-06); two females and one juvenile at Istituto per la Protezione Sostenibile delle Piante (IPSP), Consiglio Nazionale delle Ricerche (CNR), Bari, Italy (AR083-22); two females and two juveniles at Royal Belgian Institute of Natural Sciences, Brussels, Belgium (RIT 852); and two females and two juveniles at USDA Nematode Collection, Beltsville, MD, USA (T-6775p); collected by A. Archidona-Yuste, April 22, 2014.Xiphinema celtiense sp. nov. is a Xiphinema non-americanum-group species belonging to morphospecies Group 5 sensu Loof & Luc [of & Luc . It is aof & Luc , the newCelti\u2019, where the type specimens were collected.The species name is derived from originating Roman city of Pe\u00f1aflor, \u2018ca 58\u201378% of lip region diam. Guiding ring with average guiding sheath length of 15.5 \u03bcm. Odontostyle long, 1.4\u20131.8 times longer than odontophore, and the latter with well-developed flanges 13.0\u201316.5 \u03bcm wide. Pharynx very long occupying about 10\u201314% of body length, consisting of an anterior slender narrow part 379\u2013510 \u03bcm long and extending to pharyngeal bulb, 126.0\u2013168.0 \u03bcm long and 22.5\u201336.0 \u03bcm wide. Glandularium 110\u2013155 \u03bcm long. Nucleus of dorsal pharyngeal gland (DN) located at beginning of basal bulb (11.5\u201316.1%), ventrosublateral nuclei (SVN) situated ca halfway along bulb (50.5\u201362.3%)  \u03bcm wide and 7.2 \u00b1 1.4 (4.5\u20139.5) \u03bcm high. Amphidial fovea aperture extending for  Coomans . Vestigiize Figs . Small cExtremely rare, only one male specimen was found in type locality. Male genital tract diorchic with testes containing multiple rows of different stages of spermatogonia. Tail short, convex-conoid with a broadly rounded terminus and thickened outer cuticular layer. Spicules moderately long and slightly curved ventrally; lateral guiding pieces more or less straight or with curved proximal end. One pair of adanal and 4 mid-ventral supplements.ca 75 \u03bcm. Tail morphology of second-stage juvenile similar to J1 expect to absence of knob-like expansion, and tail conoid and subdigitate with rounded terminus for third-stage juvenile. In J4 tail conoid with a short bulge rounded terminus  were identified using morphological characters such as body length, length of replacement and functional odontostyle , Fig 5 , 76. Speterminus . All juvterminus , Fig 7.Xiphinema celtiense sp. nov. was found in the rhizosphere soil of wild olive (Olea europaea subsp. silvestris (Miller) Lehr) in one additional locality belonging to C\u00f3rdoba province. , posterior vulva position (50.0\u201355.0 vs 46.0\u201351.0%), a longer odontostyle and odontophore  resulting in a longer stylet length (241.0\u2013263.05 vs 213.0\u2013234.0 \u03bcm), a narrower lip region (13.5\u201316.0 vs 15.5\u201317.0 \u03bcm), frequency of males (extremely rare vs frequent), and the female and J1 tail shape , posterior vulva position (50.0\u201355.0 vs 47.1\u201351.8%), and presence vs absence of crystalloid bodies along uterus [X. turcicum by slightly higher a and c ratios , presence vs absence of crystalloid bodies in the tubular portion of uterus, and different shape of J1 tail  although in both species the tail ends in a knob-like expansion more or less separated from the anterior part of tail , and female tail shape  [X. cohni it mainly differs by the presence vs absence of Z-differentiation containing numerous granular bodies, and female tail shape  , 65. AndX. celtiense sp. nov. (KX244889-KX244890) differed with the closest related species, X. hispanum (GU725074) by 24 nucleotides and 3 gaps (97% similarity) and from X. cohni  from 27 nucleotides and 1 indel (97% similarity). Intraspecific variation of D2-D3 segments detected between the two studied population of X. celtiense sp. nov. consisted of 7 nucleotides (99% similarity), and no indels (X. hispanum (GU725061) and 86% (141 nucleotides and 34 indels) with X. cohni (KX244933). Intraspecific variation of the ITS1 for these sequences (KX244926-KX244927) was 44 nucleotides and 18 gaps, 95% similarity (X. celtiense sp. nov. (KX244943) showed a high level of similarity (99%) with several sequences deposited in GenBank such as X. hispanum (GU725083), X. adenohystherum (GY725084), and X. nuragicum (GU725080).D2-D3 sequences from o indels . ITS1  , at Izn\u00e1jar, C\u00f3rdoba province, Spain; collected by J.E. Palomares-Rius, December 3, 2014; mounted in pure glycerine and deposited in the nematode collection at Institute for Sustainable Agriculture (IAS) of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection number JAO-25-1).Adult female, collected from the rhizosphere of cultivated olive  of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection numbers JAO-25-2-JAO-25-7); one female and one male at Istituto per la Protezione Sostenibile delle Piante (IPSP), Consiglio Nazionale delle Ricerche (CNR), Bari, Italy (JAO-25-12); two females and one juvenile at Royal Belgian Institute of Natural Sciences, Brussels, Belgium (RIT 853); and two females, one male, and one juvenile at USDA Nematode Collection, Beltsville, MD, USA (T-6777p); collected by J.E. Palomares-Rius, December 3, 2014.Xiphinema iznajarense sp. nov. is an amphimictic species belonging to morphospecies Group 5 from X. non-americanum-group species sensu Loof & Luc [ca 71 \u03bcm), and one pair of adanal supplement plus 4\u20135 pairs of ventromedian supplements; and specific D2-D3, ITS1 rRNA and partial 18S rRNA sequences . According to the polytomous key of Loof & Luc [of & Luc . It is cof & Luc , the newThe species epithet refers to the type locality, Izn\u00e1jar, where the species was detected.ca 63\u201374% of lip region diam. and located at ca two-thirds of lip region height. Guiding ring with average guiding sheath length of 12.0 \u03bcm. Odontostyle moderately long, 1.5\u20131.8 times longer than odontophore, and the latter with well-developed flanges in the most of specimens studied, 11.5\u201322.0 \u03bcm wide. Pharynx consisting of an anterior slender narrow part 265\u2013414 \u03bcm long, extending to a cylindrical, terminal pharyngeal bulb occupying ca 23\u201336% of total pharyngeal length, cylindrical, 117\u2013153 \u03bcm long and 20\u201329 \u03bcm wide. Glandularium 101\u2013135 \u03bcm long. Nucleus of dorsal pharyngeal gland (DN) located at beginning of basal bulb (11.6\u201312.6%), ventrosublateral nuclei (SVN) situated ca halfway along bulb (50.5\u201357.8%) , a reflexed oviduct with well-developed pars dilatata oviductus separated from uterus by a well-developed sphincter. Uteri tripartite, comprising a developed pars dilatata uteri continuing into a narrower, muscular tube-like portion, and a well-developed Z-differentiation with weakly muscularised wall and containing numerous small granular bodies. Uterine wall wrinkles present along uterus, being more numerous in the proximal part of pars dilatata uteri and ovejector . Ovejector well-developed 35.5\u201356.0 \u03bcm wide, vagina perpendicular to body axis, 18.0\u201324.0 \u03bcm long in lateral view. Vulva slit-like, pre-equatorial. Tail conoid and short, dorso-ventrally convex, ending in a rounded and broadly terminus, bearing in four to five pairs of caudal pores on each side.Habitus in specimens killed by gentle heat usually almost straight anterior to the vulva, more curved behind the vulva, occasionally open C-shaped. Cuticle 2.0\u20134.0 \u03bcm thick at mid-body, more thickened in the lip region (4.0\u20136.0 0 \u03bcm wide) and tail tip region (5.5\u201310.0 \u03bcm wide). Lateral hypodermical chords occupying about 26\u201346% of the corresponding maximum body diameter. Lip region hemispherical, broadly rounded frontally, usually low and offset from body contour by a shallow constriction; 15.5\u201317.0 \u03bcm wide and 5.5\u20137.5 \u03bcm high. Amphidial fovea aperture extending for vejector . Small svejector . In someFrequent but less abundant than female (ratio = 1: 2). Morphologically similar to female except for genital system and more curved posterior part of body. Male genital tract diorchic with testes containing multiple rows of different stages of spermatogonia. Tail short, convex-conoid with short bulge rounded terminus and thickened outer cuticular layer Figs . Spiculeca 63 \u03bcm. Tail morphology in second-stage juvenile similar to J1 expect for the presence a slightly depression at the level of the hyaline region in both sides. On the other hand, the tail was conoid and subdigitate with a rounded terminus for J3, while for fourth-stage juvenile was conoid with rounded terminus and short bulge  were identified using morphological characters such as body length, length of replacement and functional odontostyle , Fig 5 , 76. In rt bulge . All juvrt bulge .Xiphinema iznajarense sp. nov. was only found in type locality, Izn\u00e1jar (C\u00f3rdoba province), being extracted from the rhizosphere of cultivated olive (Olea europaea subsp. europaea L.) (Morphometric variability is described in paea L.) , Fig 1.X. iznajarense sp. nov. closely resembles with X. celtiense sp. nov., X. coronatum and X. turcicum. Xiphinema iznajarense sp. nov. can be differentiated from X. celtiense sp. nov. by the characters discussed above. From X. coronatum it differs in having a longer body (4.5\u20135.8 vs 3.8\u20134.6 mm), higher a ratio (75.2\u2013106.3 vs 65.5\u201375.5), a shorter odontophore and lower oral aperture-guiding ring distance , frequency of males (frequent vs extremely rare), presence vs absence of crystalloid bodies in the tubular portion of uterus, female tail shape , and shape of J1 tail (conoid elongate with rounded terminus vs a long clavate peg) , a shorter odontostyle length (132.0\u2013151.0 vs 152.0\u2013182.0 \u03bcm), the presence vs absence of crystalloid bodies along uterus, the frequency of males (frequent vs rare), the female tail shape , and shape of J1 tail  , lower c\u00b4 ratio (0.7\u20131.1 vs 1.4\u20132.2), a longer odontostyle (132.0\u2013151.0 vs 107.0\u2013131.0 \u03bcm), and the presence vs absence of crystalloid bodies along uterus  was 97% similar (26 nucleotides and 1 indel) to X. adenohystherum (GU725075), X. hispidum (KC567181) and 95% similar (36 nucleotides and 2 indels) to X. hispanum (GU725074). No intraspecific variation of D2-D3 segments was detected amongst the studied individuals (100% similarity) (X. hispanum (GU725061), X. adenohystherum (GU725063) and X. hispidum (HM921367) with similarity values of 88% (131 nucleotides and 31 indels), 87% (145 nucleotides and 29 indels) and 84% (175 nucleotides and 52 indels), respectively (X. iznajarense sp. nov. (KX244944) closely matched with several species of Xiphinema, some of them were X. adenohystherum (GU725084), X. hispanum (GU725083), X. gersoni Roca & Bravo, 1993 [X. sphaerocephalum Lamberti, Castillo, G\u00f3mez Barcina & Agostinelli, 1992 [D2-D3 region of ilarity) . Similarectively . ITS1 alvo, 1993   , at Meng\u00edbar, Ja\u00e9n province, Spain; collected by J. Mart\u00edn-Barbarroja, March 25, 2012; mounted in pure glycerine and deposited in the nematode collection at Institute for Sustainable Agriculture (IAS) of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection number O3C4-01).Adult female, collected from the rhizosphere of cultivated olive  of Spanish National Research Council (CSIC), C\u00f3rdoba, Spain (collection numbers O3C4-02-O3C4-08); one female, one male and one juvenile at Istituto per la Protezione Sostenibile delle Piante (IPSP), Consiglio Nazionale delle Ricerche (CNR), Bari, Italy (O3C4-19); one female and one male at Royal Belgian Institute of Natural Sciences, Brussels, Belgium (RIT 854); and one female and one male at USDA Nematode Collection, Beltsville, MD, USA (T-6776p); collected by J. Mart\u00edn-Barbarroja, March 25, 2012.Xiphinema mengibarense sp. nov. belongs to the X. non-americanum Group 5 in Loof and Luc [Xiphinema non-americanum Group 5 and has the following specific alphanumeric codes (codes in parentheses are exceptions): A4-B2+3-C5a-D6(5)-E6(5)-F45-G32-H2-I2-J6-K2-I2. and Luc ; and it  and Luc , the speThe species epithet refers to the type locality, Meng\u00edbar, where the species was detected.ca 64\u201378% of lip region diam. and located at ca two-thirds of lip region height. Guiding ring and guiding sheath variable in length depending on degree of protraction/retraction of stylet. Odontostyle moderately long, 1.5\u20131.7 times longer than odontophore, in the most specimens the latter with moderate-developed flanges, but in some specimens it was observed weaker, 8.5\u201314.0 \u03bcm wide. Pharynx composed by an anterior slender narrow flexible part 317\u2013417 \u03bcm long, and a posterior muscular expanded part with three nuclei. Terminal pharyngeal bulb variable in length, 120\u2013173 \u03bcm long and 19.5\u201329.5 \u03bcm wide. Glandularium 104\u2013148 \u03bcm long. Nucleus of dorsal pharyngeal gland (DN) located at beginning of basal bulb (9.2\u201315.0%), ventrosublateral nuclei (SVN) situated ca halfway along bulb (45.7\u201358.0%) , with slightly bulging rounded terminus with a distinct terminal blind canal. Three to four caudal pores present on each side.Body cylindrical in an open C-shape when heat relaxed. Cuticle 3.1 \u00b1 0.3 (2.0\u20134.5) \u03bcm thick at post-lip region, 2.8 \u00b1 0.5 (2.0\u20134.0) \u03bcm wide at mid-body, but thicker just posterior to anus, 6.4 \u00b1 1.8 (5.0\u201310.0) \u03bcm thick. Lateral chord 13.0 \u00b1 4.8 (8.0\u201320.0) \u03bcm wide, occupying 17\u201342% of corresponding body diam. Lip region flatly rounded, slightly offset from body contour by a depression, 13.9 \u00b1 0.7 (12.5\u201315.5) \u03bcm wide and 6.9 \u00b1 0.8 (5.5\u20138.5) \u03bcm high. Amphidial fovea aperture extending for  Coomans ). CardiaFunctional, less abundant than females (ratio = 1: 2). Reproductive system diorchic with testes occupying 45\u201357% of body length, and spindle-shaped sperm. Spicules dorylaimoid, massive, well sclerotised, 57.5\u201366.0 \u03bcm long, ventrally curved with tubular lateral guiding pieces 13.5\u201318.0 \u03bcm long. One pair of adanal supplements located at 16.6 \u00b1 1.2 (15.5\u201319.0) \u03bcm from cloacal opening and a series of four to five ventromedian supplements. Tail similar to that of female, dorsally more convex than female, and ending in a rounded terminus with short bulge.ca 53 \u03bcm, and shorter distance from anterior end to stylet guiding-ring than that in adult stages. Tail morphology in second and third juvenile stages similar to J1, becoming progressively shorter and stouter in each progressively moult. However, tail shape in fourth-stage similar into that of female, broadly dorsally convex-conoid with slightly bulging rounded terminus , being extracted from the rhizosphere of cultivated olive (Olea europaea subsp. europaea L.) (Morphometric variability is described in paea L.) , Fig 1.X. mengibarense sp. nov. groups with X. herakliense Tzortzakakis et al., 2015 [X. hispanum, and X. lanceolatum Roca & Bravo, 1993 [Xiphinema mengibarense sp. nov. can be differentiated from X. herakliense by higher a and c ratios , a shorter odontostyle (120.0\u2013131.5 vs 127.0\u2013157.0 \u03bcm), shorter spicules (57.5\u201366.0 vs 70.0\u201381.0 \u03bcm) [X. mengibarense sp. nov. mainly differs from X. hispanum in having higher a ratio (80.0\u201398.2 vs 73.1\u201383.9), a shorter odontostyle (120.0\u2013131.5 vs 131.2\u2013142.3 \u03bcm), the number of spiniform structures present in the Z-differentiation (lower vs abundant), the presence vs absence of crystalloid bodies in the tubular portion of uterus, and the frequency of males (frequent vs rare) , a shorter odontostyle and odontophore  resulting in a shorter stylet (194.5\u2013215.0 vs 255.5\u2013283.0 \u03bcm), a slightly narrower lip region (12.5\u201315.5 vs 14.5\u201318.0 \u03bcm), posterior vulva position (48.5\u201357.0 vs 43.5\u201350.0%), the presence vs absence of males, and the number of spiniform structures and crystalloid bodies (lower vs very abundant)  was 94% similar to several Xiphinema species such as X. italiae , X. pyrenaicum Dalmasso, 1969 [X. sphaerocephalum . Xiphinema mengibarense sp. nov. showed a high homogeneity for the D2-D3 region  in the three sampled populations (X. hispanum (GU725061) and X. cohni (KC567159), with a similarity of 84%  (X. mengibarense sp. nov. (KX244945) closely matched (99% similarity) those for X. italiae (FJ713154), X. pyrenaicum (GU725085) and X. gersoni (KC567154).D2-D3 region of so, 1969  (GU72507ulations . The cloctively) . Low intX. adenohystherum, X. baetica, X. cohni, X. coxi europaeum, X. duriense, X. hispanum, X. hispidum, X. incertum, X. index Thorne & Allen, 1950 [X. italiae, X. lupini Roca & Pereira, 1993 [X. macrodora, X. madeirense Brown, Faria, Lamberti, Halbrendt, Agostinelli & Jones, 1993 [X. nuragicum, X. oleae, X. opisthohysterum Siddiqi, 1961 [X. pachtaicum, X. parapachydermum, X. plesiopachtaicum, X. rivesi Dalmasso, 1969 [X. santos Lamberti, Lemos, Agostinelli & D\u2019Addabbo, 1993 [X. sphaerocephalum, X. turcicum, X. turdetanense, and X. vallense have been previously recorded within studies of dagger and needle nematodes infesting olives and vineyards in southern Spain [viz. X. cadavalense, X. conurum and X. pseudocoxi Sturhan, 1985 [Morphological and morphometrical data, and molecular delineation (rDNA) of en, 1950 , X. italra, 1993 , X. macres, 1993 , X. nuraqi, 1961 , X. pachso, 1969 , X. santbo, 1993 , X. spharn Spain , 18, 28.an, 1985 ), a brieXiphinema collected from the rhizosphere of cultivated olive (Olea europaea subsp. europaea L.) at Espiel (C\u00f3rdoba province) corresponds fairly well with studied paratypes and original description of X. cadavalense. This population was characterised by a long body; lip region hemispherical, rounded both anteriorly and laterally and set off from body contour by slightly depression; long odontostyle and odontophore; reproductive system didelphic-amphidelphic with both branches about equally developed with a well-developed Z-differentiation with weakly muscularised wall and comprising 9\u201315 sclerotized bodies of variable size and petal shape, each one consisting of a large portion, irregularly spherical surrounded by a variable number of refractive pieces; spiniform structures and crystalloid bodies in very small size and low number present along the narrower and muscular tube-like of uterus; tail dorsally convex-conoid  ending in a terminal peg with blind canal , posterior guiding ring position from oral aperture (149.5\u2013167.0 vs 126.5\u2013148.5 \u03bcm) [X. cadavalense has the following specific alphanumeric codes (codes in parentheses are exceptions): A4-B2+3-C5a-D65-E56-F5-G4-H2-I3-J-K-L1.The amphimictic population of nd canal . The obs and Luc , which aavalense . In addi48.5 \u03bcm) . These d48.5 \u03bcm) , this SpX. cadavalense (KX244900) was 98% similar (14 nucleotides and no indels) to X. baetica (KC567168), 97% similar (24 nucleotides and 3 indels) to X. andalusiense sp. nov. (KX244884-KX244888) and 96% similar (30 nucleotides and 10 indels) to X. macrodora . ITS1 sequence (KX244932) region also agrees with results obtained from D2-D3, this sequence was 90% similar (105 nucleotides and 28 indels) to X. baetica (KC567157), 89% (121 nucleotides and 35 indels) to X. andalusiense sp. nov. (KX244921-KX244925) and 86% (157 nucleotides and 70 indels) to X. macrodora (KU171048). The partial 18S region of X. cadavalense (KX244946), was very similar to several sequences of Xiphinema spp., including X. diversicaudatum  Thorne, 1939 [X. baetica (KC567149) and X. bakeri Williams, 1961 [D2-D3 segments of nse KX2440 was 98%et al. [The Spanish population of this species from the rhizosphere of olive was characterised by a lip region rounded offset from the rest of the body by a conspicuous depression, two equally developed female genital branches, vulva slightly anterior to mid-body, uterus with uterine differentiation, presence of Z-differentiation with small granular bodies plus small spines (in low number), female tail conical, ventral profile nearly straight, dorsal profile regularly curved with rounded terminus . The moret al. . Up to oX. conurum (KX244902) matched well, 99% similar with former sequences from Tunisia deposited in GenBank (KX062671-KX062673); and ITS1 (KX244934) was 95\u201396% similar with former sequences from Tunisia deposited in GenBank (KX062696-KX062697). And partial 18S (KX244947) was provided for the first time in this research, being 99% similar to several Xiphinema spp. such as X. nuragicum (GU725081) or X. israeliae Luc, Brown & Cohn, 1982 [D2-D3 sequence for TS1 KX2444 was 95\u2013Xiphinema collected from the rhizosphere of wild olive (Olea europaea subsp. silvestris (Miller) Lehr) at Alcaracejos (C\u00f3rdoba province) agrees fairly well with original description of X. pseudocoxi. This population was characterised by a moderately long body in an open C-shaped after fixation; lip region distinct from the body contour by a depression, frontally rounded; female reproductive system didelphic-amphidelphic having both branches about equally developed; Z-differentiation with weakly muscularised wall formed by 6\u201310 globular bodies similar in size, and irregularly spherical surrounded by a variable number of refractive pieces; no spiniform structures and, crystalloid bodies nor sperm cells observed along uterus; female tail convex-conoid, varying slightly in shape, and ending in a terminal peg with a blind canal : A4-B2-C5a-D45-E4(5)-F4(5)-G2-H2-I3-J-K-L1.The amphimictic population of nd canal , Table 9 and Luc , which ieudocoxi . Additioeudocoxi , 94, 95.eudocoxi , the newX. pseudocoxi (KX244915-KX244916) were obtained for the first time in this study. The closet species regarding D2-D3 segments of X. pseudocoxi (KX244915-KX244916) were X. globosum Sturhan, 1978 [X. diversicaudatum  and X. coxi europaeum . Similarly, ITS1 region (KX244939-KX244940) also showed some similarity with X. globosum , X. diversicaudatum  and X. coxi europaeum . Finally, the partial 18S of X. pseudocoxi (KX244948) matched closely (99%) with several Xiphinema spp., such as X. globosum (GU549476), X. diversicaudatum (EF538761), X. bakeri (AY283173), X. vuittenezi Luc, Lima, Weischer & Flegg, 1964 [X. index (AY687997).Sequences for an, 1978   were used for further phylogenetic studies. Sequences for X. andalusiense sp. nov., X. cadavalense, X. celtiense sp. nov., X. duriense, X. iznajarense sp. nov., X. mengibarense sp. nov., X. opisthohysterum and X. pseudocoxi were obtained for these species in this study. On the other hand, sequences from X. adenohystherum, X. cohni, X. conurum, X. hispanum, X. hispidum, X. incertum, X. index, X. italiae, X. nuragicum, X. parapachydermum, X. turcicum and X. turdetanense matched well with former sequences deposited in GenBank, and spread out the molecular diversity of these species to the newly studied areas.The amplification of D2-D3 expansion segments of 28S rRNA, ITS1 rRNA, and partial 18S rRNA yielded a single fragment of approximately 800 bp, 1000 bp, and 1800 bp, respectively, based on gel electrophoresis. Sequences from other species of Xiphinema non-americanum-group species inferred from analyses of D2-D3 expansion segments of 28S, ITS1, and the partial 18S rDNA gene sequences using BI are given in Figs X. non-americanum-group spp. based in a multiple edited alignment including 103 sequences and 753 total characters showed two clearly separated (PP = 1.00) major clades  with X. cohni  and X. hispanum , this clade was related (PP = 1.00) with another subclade which was formed by X. iznajarense sp. nov. (KX244891-KX244892), X. adenohystherum , X. hispidum  and X. gersoni (KC567180). Finally, X. mengibarense sp. nov. formed a low-supported subclade (PP = 0.76) with X. italiae , X. pyrenaicum (GU725073), and X. meridianum Heyns, 1971 [X. bakeri and X. index which belong to Groups 7 and 8, respectively. This clade grouped sequences from the new species X. andalusiense sp. nov. (KX244884-KX244888) and the new accessions from X. cadavalense (KX244900), X. conurum (KX244902), and X. pseudocoxi (KX244915-KX244916). Xiphinema andalusiense sp. nov. (KX244884-KX244888) from wild olive occupied a superior position within the clade II forming a well-supported subclade (PP = 1.00) with X. cadavalense (KX244900) from cultivated olive, X. baetica  and X. macrodora . Finally, X. pseudocoxi (KX244915-KX244916) was phylogenetically related to X. globosum (GU549474) forming a well-supported clade (PP = 0.99).Phylogenetic relationships among r clades . Clade IX. non-americanum-group species, was 1113 characters after discarding ambiguously aligned regions from the alignment. Two new accessions were used as outgroup, X. duriense (KX244935) and X. opisthohysterum (KX244938). The 50% majority rule consensus BI tree of X. non-americanum-group spp. showed two major clades (PP = 1.00) similar to those obtained for D2-D3 region , X. pseudocoxi (KX244939-KX244940) and X. cadavalense (KX244932). Xiphinema andalusiense sp. nov. (KX244921-KX244925) and X. cadavalense (KX244932) clustered with X. baetica (KC567156-KC567157) and X. macrodora (KU171048) in a well-supported subclade (PP = 1.00), these results agree with the results obtained with D2-D3 region. Xiphinema pseudocoxi and X. globosum were also phylogenetically related to this marker and they were placed in a well-supported subclade (PP = 1.00) which was related (PP = 0.96) at the same time with X. turdetanense (KC567163). Clade II grouped thirteen species from different morphospecies Groups 1, 4, 5 and 7, including X. celtiense sp. nov., X. iznajarense sp. nov. and X. mengibarense sp. nov. Xiphinema iznajarense sp. nov. (KX244928-KX244929), and X. celtiense sp. nov. (KX244926-KX244927) clustered together with X. cohni (KX244933), X. adenohystherum (GU725063), X. hispanum (GU725061) and X. hispidum (HM921367) as occurred in the D2-D3 tree. Finally, X. mengibarense sp. nov. (KX244930-KX244931) formed a low-supported subclade with X. barense Lamberti, Roca, Agostinelli & Bleve-Zacheo, 1986 [X. pyrenaicum (GU725060) although this relation also was poorly supported. The new accessions for X. duriense (KX244935) and X. opisthohysterum (KX244938) clustered together with X. pachtaicum (AY430178) as an outgroup, all of them from the X. americanum-group  clustered with X. cadavalense, X. macrodora and X. baetica within the same well-supported subclade (PP = 1.00). Phylogenetic inferences based on 18S also suggest that X. pseudocoxi and X. globosum are related species, although this relation was poorly supported (X. iznajarense sp. nov. (KX244944), X. celtiense sp. nov. (KX244943) and X. mengibarense sp. nov. (KX244945) clustered in this case with X. cohni (KC567151), X. hispanum (GU725083), X. adenohystherum (GU725084), X. italiae , X. barense (KM199695), X. gersoni (KC567154), X. sphaerocephalum (GU725082), and X. pyrenaicum (GU725085) within a well-supported subclade (PP = 1.00).The 50% majority rule BI tree of a multiple alignment including 60 18S sequences and 1647 bp long showed several major clades . Additioupported . FinallyXiphinema associated with wild and cultivated olives in southern Spain, as well as their distribution and molecular phylogeny. This was conducted in an extensive and systematic nematological survey that included 211 locations and 453 sampling sites. We found 385 Spanish populations of Xiphinema spp. infesting olive soils. We described four new Xiphinema species, enlarging the diversity of Xiphinema species in the Iberian Peninsula which is in agreement with previous data obtained for the phylogeny and biogeography of the genus Xiphinema and Longidorus in the Euro-Mediterranean region [Xiphinema based on nuclear rDNA markers.This study aimed to get knowledge and a better understanding on the occurrence, abundance and biodiversity of dagger nematodes of the genus n region , 101. ToXiphinema is one of the most diverse PPN associated with olive, with twenty species  reported in various countries of the Mediterranean Basin [Xiphinema species detected in olive worldwide, including four new species from the X. non-americanum-group. All these species were new records for olive with the exception of X. pachtaicum, X. index, X. italiae, X. nuragicum and X. turcicum [Xiphinema spp. observed in both olive types, our study showed a great species diversity, that was mainly associated with the X. non-americanum-group species  [X. non-americanum-group, i.e. X. italiae found in all provinces, X. nuragicum in 7 out of 8 provinces, and X. coxi europaeum in 5 out of 8 provinces. In this sense, the presence of a high number of frequent species belonging to X. non-americanum-group  explains the higher value observed in Hill\u00b4s 2 (Dominance diversity) index with respect to X. americanum-group  . Other sXiphinema cause damage to olive by feeding on unmodified plant root cells and causing cell necrosis and galling in root apex [X. diversicaudatum and X. vuittenezi [Xiphinema spp. were responsible for 5 to 10% of loss production resulting in an estimated $39 million loss [Xiphinema spp. in both olive types studied  because of their sensitivity to fast desiccation, large body size, and the absence of survival-resistance forms\u201d. Unfortunately, little is known about the ecological requirements of Xiphinema nematodes and further research is needed [Longidorus spp. showed evidence of some geographic species associations in Andalusia [Xiphinema spp. in southern Spain and other wider areas.Overall, nematode diversity decreases rapidly to agricultural management including plant-parasitic nematodes . Our resin areas . Althoug species , the geoe et al.  for Longs needed . Some prndalusia . ConsequX. cadavalense, X. pseudocoxi, and X. conurum) and for ITS1 . D2-D3 expansion region was more useful for establishing phylogenetic relationships among Xiphinema species than ITS1 or 18S. Phylogenetic analyses based on D2-D3, ITS1, and partial 18S using BI resulted in a consistent position for the newly described species of X. non-americanum-group species from Spain, which grouped in two separated clades, and mostly agree with the clustering obtained by other authors [X. andalusiense sp. nov., X. baetica, and X. cadavalense. In the case of X. andalusiense sp. nov. vs X. baetica, only lower a and c\u2019 ratios, the absence of spines in the uterus, the absence of males and different ribosomal genes could separate X. baetica from X. andalusiense sp. nov. These species probably evolved in the Iberian Peninsula as they occur only there. The Iberian Peninsula has been suggested as a possible center of recent speciation for PPN nematode genera such as Longidorus, Trichodorus or Rotylenchus species [Xiphinema celtiense sp. nov., X. iznajarense sp. nov. and X. mengibarense sp. nov. could be clearly separated morphologically and molecularly from the other Xiphinema species. The majority of the species showed congruence in the phylogenetic relationships within D2-D3, ITS1, and partial 18S using the DNA from the same individual and these markers matched very well with the sequences deposited in the GenBank. This result is in contrast with the close related genus Longidorus found in a similar sampling scheme and localities in which the diversity of species was lower and all the species occupies two major positions in the phylogenetic clade [Sequences of nuclear ribosomal RNA genes, particularly D2-D3 and ITS1, are useful molecular markers for providing accurate species identification of Longidoridae , 30, 127 authors , 18. The species . Xiphineic clade .X. americanum-group and the non-americanum group species. This research provides molecular markers for precise and unequivocal diagnosis of some species of Xiphinema in order to differentiate virus vector or quarantine species. Furthermore, it reflects that similar intensive and extensive integrative studies on Xiphinema species based on widest areas may help to elucidate the evolutionary origin of Xiphinema species. In this sense, further studies based on widespread species (i.e. X. pachtaicum) could also help to clarify if the main speciation occurred in Africa leading to many apomictic species in tropical and subtropical environments as hypothesised by Coomans [In summary, this study provides new insights into the diversity of this genus associated with the olive in Mediterranean conditions with important differences related to the species within the  Coomans , or in SS1 Table(DOCX)Click here for additional data file."}
+{"text": "Alchornea cordifolia and Canthium subcordatum were obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS). The essential oils were subjected to in vitro antibacterial, antifungal and cytotoxic activity screening. Thirty-eight compounds comprising 97.7% of A. cordifolia oil and forty-six constituents representing 98.2% of C. subcordatum oil were identified. The major components in A. cordifolia oil were methyl salicylate (25.3%), citronellol (21.4%), \u03b1-phellandrene (7.4%), terpinolene (5.7%) and 1,8-cineole (5.5%). Benzaldehyde (28.0%), \u03b2-caryophyllene (15.5%), -\u03b1-farnesene (5.3%) and methyl salicylate (4.5%) were the quantitatively significant constituents in C. subcordatum fruit essential oil. A. cordifolia essential oil demonstrated potent in vitro antibacterial activity against Staphylococcus aureus (MIC = 78 \u03bcg/mL) and marginal antifungal activity against Aspergillus niger (MIC = 156 \u03bcg/mL). C. subcordatum showed antibacterial activity against Bacillus cereus and S. aureus (MIC = 156 \u03bcg/mL) and notable antifungal activity against A. niger (MIC = 39 \u03bcg/mL). However, no appreciable cytotoxic effects on human breast carcinoma cells (Hs 578T) and human prostate carcinoma cells (PC-3) were observed for either essential oil. The antimicrobial activities of A. cordifolia and C. subcordatum fruit essential oils are a function of their distinct chemical profiles; their volatiles and biological activities are reported for the first time.Bacterial resistance has been increasingly reported worldwide and is one of the major causes of failure in the treatment of infectious diseases. Natural-based products, including plant secondary metabolites , can be exploited to ameliorate the problem of microbial resistance. The fruit essential oils of  Antimicrobial resistance is one of the most serious public health threats that results mostly from the selective pressure exerted by antibiotic use and abuse ,2. DurinAlchornea cordifolia (Schumach. and Thonn.) M\u00fcll. Arg. (Euphorbiaceae) is a shrub found along the coastal regions of West Africa. It has multipurpose utilization as fodder, food and medicine. The leaves, roots and stem bark extracts are used extensively in traditional medicine in the preparation of drugs for urinary, respiratory and gastro intestinal disorders [A. cordifolia has been a subject of scientific studies: anti-microbial, antioxidant and anticancer activities, etc. [A. cordifolia has been reported [A. cordifolia has demonstrated antibacterial activity against 21 bacterial strains tested and showed the highest levels of antibacterial activity with MICs against methicillin-resistant Staphylococcus aureus (MRSA) in the range of 1.6\u20133.1 mg/mL and MBCs in the range of 6.3\u201312.5 mg/mL among 24 other plant species studied [A. cordifolia leaf [isorders . A slurrisorders . The leaisorders . Hithertes, etc. ,11,12,13reported . The aqu studied . Phyto-clia leaf ,17,18,19Canthium subcordatum DC.  is a tree that grows in central and western Africa and reaches a height of more than 10 m [C. subcordatum [S,9R)-roseoside [C. subcordatum and their structures deduced. GC-MS analysis and anticancer activity of ethanol leaf extract of C. parviflorum have also been reported [A. cordifolia and C. subcordatum.han 10 m . Its roohan 10 m  and the han 10 m . Recentlcordatum . A numbeoseoside  and shanoseoside  have beereported ,27. As pA. cordifolia and C. subcordatum were collected in the month of July 2004, from the campus of the University of Ibadan, Nigeria. Plant samples were authenticated by F. Usang of the Herbarium Headquarters, Forest Research Institute of Nigeria (FRIN), Ibadan, Nigeria, where voucher specimens  were deposited.The mature fresh fruits of The fruit essential oils were obtained by hydrodistillation (4 h) of the pulverized air-dried plant samples (500 g) in an all glass Clevenger-type apparatus following the British Pharmacopoeia specifications . The fruhttp://www.agilent.com/en-us/products/software-informatics/massspec-workstations/gc-msd-chemstation-software). The GC column was an HP-5ms fused silica capillary with a (5% phenyl)-methyl polysiloxane stationary phase (30 m \u00d7 0.25 \u03bcm film thickness). The carrier gas was helium with a column head pressure of 7.07 psi and flow rate of 1.0 mL/min. Inlet temperature was 200 \u00b0C and MSD detector temperature was 280 \u00b0C. The GC oven temperature program was used as follows: 40 \u00b0C initial temperature, held for 10 min; increased at 3 \u00b0C/min to 200 \u00b0C; increased 2 \u00b0C/min to 220 \u00b0C. The sample was dissolved in CH2Cl2, and 1 \u03bcL was injected using a splitless injection technique.The essential oils were subjected to GC-MS analysis on an Agilent system consisting of a model 6890 gas chromatograph, a model 5973 mass selective detector (MSD) , and an Agilent ChemStation data system  and by comparison of their mass spectral fragmentation patterns  and with the literature .A. cordifolia and C. subcordatum essential oils were screened for antibacterial activity against Bacillus cereus (ATCC No. 14579), Staphylococcus aureus (ATCC No. 29213), Pseudomonas aeruginosa (ATCC No. 27853), and Escherichia coli (ATCC No. 10798). Minimum inhibitory concentrations (MICs) were determined using the micro broth dilution technique [w/w solutions of samples in DMSO plus 50 \u03bcL CAMHB. The sample solutions were serially diluted (1:1) in CAMHB in 96-well plates to give concentrations of 2500, 1250, 625, 313, 156, 78, 39, and 19.5 \u03bcg/mL. Organisms at a concentration of approximately 1.5 \u00d7 108 colony-forming units (CFU)/mL were added to each well. Plates were incubated at 37 \u00b0C for 24 h; the minimum inhibitory concentration (MIC) was determined as the lowest concentration without turbidity. Gentamicin was used as a positive antibiotic control; DMSO was used as a negative control .echnique . Dilutioi.e., serial dilution, concentrations of 2500, 1250, 625, 313, 156, 78, 39, and 19.5 \u03bcg/mL) using Candida albicans (ATCC No. 10231) in a yeast-nitrogen base growth medium with approximately 7.5 \u00d7 107 CFU/mL. Amphotericin B was used as the positive control. An additional test for antifungal activity against Aspergillus niger (ATCC No. 16888) was determined as above using yeast mold (YM) broth inoculated with A. niger hyphal culture diluted to a McFarland turbidity of 1.0. Amphotericin B was the positive control.Antifungal activity was determined, as described above for bacteria (2 environment at 37 \u00b0C in Dulbecco\u2019s modified Eagle medium (DMEM) with 4500 mg glucose per liter of medium, supplemented with 10% fetal bovine serum, 10 \u03bcg bovine insulin, 100,000 units penicillin and 10.0 mg streptomycin per liter of medium, and buffered with 44 mM NaHCO3, pH 7.35.Human Hs578T breast ductal carcinoma cells (ATCC No. HTB-129)  were gro2 environment at 37 \u00b0C in RPMI-1640 medium with l-glutamine, supplemented with 10% fetal bovine serum, 100,000 units penicillin and 10.0 mg streptomycin per liter of medium and buffered with 15 mM Hepes and 23.6 mM NaHCO3, pH 7.30.Human PC-3 prostatic carcinoma cells (ATCC No. CRL-1435)  were gro5 cells per well and PC-3 cells at 1.9 \u00d7 104 cells per well. The volume in each well was 100 \u03bcL for both cell types. After 48 h, supernatant fluid was removed by suction and replaced with 100 \u03bcL growth medium containing either 2.5 or 1.0 \u03bcL of dimethylsulfoxide (DMSO) solution of oils (1% w/w in DMSO), giving a final concentration of 250 or 100 \u03bcg/mL, respectively, for each oil. Hs 578T cells were tested with final concentrations at 250 \u03bcg/mL and PC-3 at final concentration of 100 \u03bcg/mL. Solutions were added to wells in four replicates. Medium controls and DMSO controls (25 or 10 \u03bcL DMSO/mL) were used. Tingenone (250 or 100 \u03bcg/mL) was used as a positive control [\u00aeAQueous Non-Radioactive Cell Proliferation assay was performed [oil/% killDMSO) were calculated.Hs 578T cells were plated into 96-well cell culture plates at 1.0 \u00d7 10 control . After terformed . After cA. cordifolia and C. subcordatum fruits, according to their elution order on HP-5ms capillary column are presented in w/w) yield. Thirty-eight compounds comprising 97.7% of A. cordifolia oil and forty-six constituents representing 98.2% of C. subcordatum oil were identified. The fruit essential oil of A. cordifolia consisted of oxygenated monoterpenoids and aromatic esters (52.9% and 26.5%), monoterpene and sesquiterpene hydrocarbons (17.2% and 14.6%) and low amounts of aliphatic alcohol and aldehyde (3.7%). The major components identified in this sample include methyl salicylate (25.3%), citronellol (21.4%), \u03b1-phellandrene (7.4%), terpinolene (5.7%) and 1,8-cineole (5.5%). Other minor constituents detected in considerable quantities were p-cymene (3.6%), \u03b1-humulene (3.0%), \u03b2-caryophyllene (2.8%) (E)-\u03b2-damascenone (2.0%) and 1-octen-3-ol (2.0%). Two uncommon essential oil constituents were identified as (Z)-rose oxide and geosmin in A. cordifolia essential oil. (Z)-Rose-oxide (isobutenyl-4-methyl tetrahydropyran) is a perfumery ingredient and a constituent  of Pelargonium essential oils and secretions of Aromia moschata [Actinomycetes, Streptomyces riseus and Streptomyces odourifer) are reported to be present on some fruits, grapes for example, and are known for their ability to produce geosmin during metabolism [et al. [A. cordifolia reported by Okoye et al. [E)-\u03b1-bergamotene (4.54%). Twenty-five constituents consisting 90.3% of the composition of the leaf essential oil were identified in 0.13% w/w yield.The relative concentrations of the volatile components in moschata . This cymoschata . Geosmintabolism . Accordi [et al. , geosmin,EE)-\u03b1-farnesene (5.3%) and methyl salicylate (4.5%) were the quantitatively significant constituents in C. subcordatum fruit essential oil. C. subcordatum fruit oil is comprised of 41.3% hydrocarbons and 56.9% oxygen-containing compounds. The five classes of organic compounds identified and reported are twenty-four hydrocarbons (41.3%), twelve alcohols (18.6%), three aldehydes (28.7%), four aromatic esters (5.8%), one ketone and one ether (3.8%). The fruit volatile oil is dominated by sesquiterpenoid compounds (50.8%), followed by aromatic compounds (33.8%), monoterpenoid compounds (7.3%), simple aldehydes and alcohols (6.3%). The monoterpenoid profile consisted of two monoterpene hydrocarbons (2.8%) and six oxygenated monoterpenes (4.5%). Twenty-two sesquiterpene hydrocarbons and seven oxygenated sesquiterpenes made up the sesquiterpenoid profile of the oil. Two unusual sesquiterpenoids, \u03b1-calacorene and longiborneol were identified in C. subcordatum fruit oil. The sesquiterpene alcohol, longiborneol is a constituent of Juniperus, Pinus, Cupressus, Dacrydium species and Cedrus deodara. It is reported to be a plant growth regulator (inhibits cress root growth and promotes wheat germination) [Canthium species. The high concentration of benzaldehyde in essential oil samples has been reported by other workers. Lei et al. [Cerasus subhirtella and C. serrulata contain 31.2% and 42.1% benzaldehyde, respectively as major constituents. Benzaldehyde (96.96%) was also indicated as a major component in the leaf essential oil of Prunus myrtifolia [Benzaldehyde (28.0%), \u03b2-caryophyllene (15.5%), (ination) . A literi et al.  showed trtifolia .A. cordifolia and C. subcordatum showed promising antimicrobial activity  and moderate antifungal activity against A. niger (MIC = 156 \u03bcg/mL); C. subcordatum oil exhibited moderate antibacterial activity against B. cereus and S. aureus (MIC = 156 \u03bcg/mL) and good antifungal activity (MIC = 39 \u03bcg/mL) against A. niger. It has been documented that Gram-positive bacteria are more sensitive to chemical compounds than Gram-negative bacteria due to differences in the structures of their cell walls; Gram-negative bacteria are less susceptible to hydrophobic small molecules such as essential oil components due to hydrophilic lipopolysaccharides in their outer membrane [A. cordifolia essential oil, Laportea aestuans essential oil, rich in methyl salicylate (54.50%), has been shown to exhibit antimicrobial activity against S. aureus, B. subtilis, P. aeruginosa, E. coli and C. albicans at 200 mg/mL compared to the standard drug; however, it was more active against the fungi Rhizopus stolon and A. niger at 25 mg/mL [Pelargonium graveolens essential oil, dominant in citronellol (26.7%), inter alia, and citronellol, also demonstrated strong inhibitory activity against S. aureus and E. coli [P. myrtifolia and apricot, Prunus armeniaca, seed essential oils) exhibited antimicrobial activity against S. aureus, S. epidermidis, B. subtilis, P. aeruginosa, E. coli and C. albicans [Stachys cretica essential oil  and \u03b2-caryophyllene are reported to exhibit strong antimicrobial activity, particularly against P. aeruginosa and B. subtilis [The antibacterial and antifungal activities of the fruit volatile oils of activity . Alchornmembrane . This is25 mg/mL ; Pelargo E. coli ,44. Simity MIC =  \u03bcg/mL agalbicans . Additiosubtilis .A. cordifolia and C. subcordatum fruit essential oils were screened for in vitro cytotoxic activity against Hs 578T human breast adenocarcinoma and PC-3 human prostatic carcinoma cells. Neither oil showed activity, however, with 0% kill at the concentrations tested.Both A. cordifolia and C. subcordatum essential oils are a function of their distinct chemical profiles. The fruit essential oil of A. cordifolia, rich in methyl salicylate and citronellol, showed antibacterial activity against S. aureus and antifungal activity against A. niger. The antimicrobial activity can be attributed to these two major components, which have shown antimicrobial activities [C. subcordatum fruit oil was active against B. cereus, S. aureus, and A. niger, but the activity is not likely due to the major component benzaldehyde, which is generally not antimicrobial [A. cordifolia and C. subcordatum essential oils are consistent with traditional uses of these medicinal plants.The antimicrobial activities of tivities ,47,48. Cicrobial , but rat"}
+{"text": "Triticosecale Wittmack) is an important feed crop which suffers severe yield, grade and end-use quality losses due to Fusarium head blight (FHB). Development of resistant triticale cultivars is hindered by lack of effective genetic resistance sources. To dissect FHB resistance, a doubled haploid spring triticale population produced from the cross TMP16315/AC Ultima using a microspore culture method, was phenotyped for FHB incidence, severity, visual rating index (VRI), deoxynivalenol (DON) and some associated traits  followed by single nucleotide polymorphism (SNP) genotyping. A high-density map consisting of 5274 SNPs, mapped on all 21 chromosomes with a map density of 0.48 cM/SNP, was constructed. Together, 17 major quantitative trait loci were identified for FHB on chromosomes 1A, 2B, 3A, 4A, 4R, 5A, 5R and 6B; two of incidence loci (on 2B and 5R) also co-located with loci for severity and VRI, and two other loci of VRI (on 1A and 4R) with DON accumulation. Major and minor loci were also identified for all other traits in addition to many epistasis loci. This study provides new insight into the genetic basis of FHB resistance and their association with other traits in triticale.Triticale (x Triticosecale Wittmack; 2n = 6 \u00d7 = 42) crop suffers severe losses in yield, grade and end-use quality due to Fusarium head blight (FHB) caused by Fusarium graminearum, Fusarium avenaceum, Fusarium poae and other spp. of genus Fusarium , five main effect QTL for Type-II resistance , seven main effect QTL for VRI  and seven main effect QTL for Type-III resistance  (R2) and were considered major QTL  ; Table 4ajor QTL . The totIn addition to the above main effect QTL, epistasis QTL were also identified for DS and VRI on chrs 1A, 2A, 3A and 3B ; Table 5QErg.lrdc-4A), 5R (QErg.lrdc-5R) and 7A (QErg.lrdc-7A)) (R2) and were considered major QTL ) ; Table 6ajor QTL ; Table 6In addition to above main effect QTL, epistasis QTL were also identified for reduced ERG incidence on chrs 6B and 6R ; Table 7QGpc.lrdc-2A), 2B (QGpc.lrdc-2B), 4R (QGpc.lrdc-4R), 5R (QGpc.lrdc-5R), 6R (QGpc.lrdc-6R) and 7B (QGpc.lrdc-7B)) (R2) and were considered major QTL ) ; Table 6QTwt.lrdc.1R), 4R (QTwt.lrdc.4R), 5A  and 5R (QTwt.lrdc.5R)) (R2) and were considered major QTL ) ; Table 6ajor QTL ; Table 7QYld.lrdc-4A), 4R (QYld.lrdc-4R.2), 6A (QYld.lrdc-6A) and 6R (QYld.lrdc-6R)) (R2) and were considered major QTL ) ; Table 6ajor QTL ; Table 7QPht.lrdc-5A), 5R (QPht.lrdc-5R), 6R (QPht.lrdc-6R) and 7R (QPht.lrdc-7R)) (QLdg.lrdc-2R), 4R , 6R (QLdg.lrdc-6R), 7B (QLdg.lrdc-7B) and 7R (QLdg.lrdc-7R)) (R2) explained by these QTL ranged from 6.0 to 38.0%. One of the above identified QTL on chr 5R (QPht.lrdc-5R) for reduced PHT was commonly shared with reduced Type-II and -III resistance and interestingly favorable allele for these, reduced PHT, DS and DON content, were contributed by TMP16315 ) ; Table 6rdc-7R)) ; Table 6Additionally, epistasis QTL were identified for PHT  but not for LDG ; Table 7QFhb.lrdc-1A) and DON (QDon.lrdc-1A), also harbored QTL for TWT (QTwt.lrdc-1A) (QFhs.lrdc-2A) harbored a QTL for PHT (QPht.lrdc-2A), a 2B region for DI (QFhi.lrdc-2B), DS (QFhs.lrdc-2B) and VRI (QFhb.lrdc-2B) harbored QTL for GPC (QGpc.lrdc-2B), a 4R region for VRI (QFhb.lrdc-4R) and DON  harbored QTL for TWT (QTwt.lrdc.4R) and LDG (QLdg.lrdc-4R), another 4R region for GPC (QGpc.lrdc-4R) harbored QTL for YLD (QYld.lrdc-4R.2), a 5A region for DON (QDon.lrdc-5A) harbored QTL for PHT (QPht.lrdc-5A), a 5R region for DI (QFhi.lrdc-5R), DS (QFhs.lrdc-5R.1) and VRI (QFhb.lrdc-5R.1) harbored QTL for TWT (QTwt.lrdc-5R), another 5R region for DON (QDon.lrdc-5R) and PHT (QPht.lrdc-5R) also harbored QTL for DS (QFhs.lrdc-5R.2) and a 6R region for GPC (QGpc.lrdc-6R) and YLD (QYld.lrdc-6R) harbored QTL for PHT (QPht.lrdc-6R) and LDG (QLdg.lrdc-6R)  QTL listed above, fifteen more  QTL were also co-localized  with other QTL. One region on chr 1A, which harbored QTL for VRI (lrdc-1A) . Similarlrdc-6R) .Fusarium head blight (then scab) in England in 1884, many scientific studies have been conducted in wheat, barley and other cereals. The genetic factors, their chromosomal locations and sources  for FHB resistance have been identified and well-studied in wheat [After the first report of in wheat . Howeverin wheat . Previouin wheat , predomiin wheat  and possin wheat ,12. In ain wheat ,55, diffin wheat  and theiin wheat  was subjParents to make this DH population were selected for commercial breeding program and neither of the parents was highly susceptible for FHB, ERG and both parents shared many other favorable characters . This inSecale cereale L. genome, which is a mosaic of various progenitor genomes [Gene associated SNPs, high-throughput genotype calling and low cost per data point of recently developed SNP assays for wheat (Infinium iSelect 90K SNP assay)  and rye  genomes . Around  genomes  also obs genomes  to 5.2 c genomes , however genomes  to 4907. genomes  cM. On t genomes  also obs genomes  construc genomes . We also genomes  and can  genomes  or previ genomes  and use R2) (R2) another FHB/DON resistance QTL. Interestingly 1A QTL region  was never identified in any of the previous triticale studies. This region also harbored an important QTL for TWT  and was derived from AC Ultima, where most of the other important QTL were derived from TMP16315. This 5A QTL seems to be common with one of the three  QTL of widely used Chinese cv. Sumai-3. However, phenotypic data indicate that AC Ultima possess low level of resistance in comparison to Sumai-3. This is obvious since AC Ultima may not have 3B and 6B QTL. However, effect of 5A QTL on FHB resistance can be validated in multiple environments for its further use in breeding programs. These observations suggest that while TMP16315 possess quantitative inheritance of minor additive genes/QTLs, AC Ultima carries a major QTL for FHB resistance along with some minor QTL. Since both parents, AC Ultima and TMP16315, contribute favorable QTL alleles, they can be utilized to pyramid and obtain a high level of FHB resistance in triticale. In this study, a few of the DH lines showed significantly better phenotype for FHB resistance than both parents. Phenotypic data largely showed continuous variation for FHB-DI, -DS, -VRI and DON traits . These oR2)  which inR2)  though QQPht.lrdc-5R); most likely this region was flanked by markers Xiac130 and Xiac128 in Kalih et al. [rPt509721 and wPtm8731 in Kalih et al. [QErg.lrdc-7A which had an additive effect of 34.0 and LOD score 8.1 (QPht.lrdc-5R) on chr 5R, which have also been most likely identified in earlier studies, Kalih et al. [Previous studies showed negative relationship of FHB resistance with plant height ,19,20,22h et al. . Howevercore 8.1 . Though,core 8.1 . Similarh et al. . Since, The understanding of genetic architecture of FHB resistance and some related traits like ERG, GPC, TWT, YLD, PHT and LDG in spring triticale is important for developing superior cultivars. Therefore, for the first time, a high-density spring triticale SNP genetic map was generated and several new major and minor effects and epistasis QTL were mapped for FHB resistance and other related traits. These QTL includes both specific (such as a 3A QTL for reduced DON content) and common/pleotropic QTL . On the other hand, both parents contributed favorable QTL alleles  and identified QTL expressed either across environments  or specifically in an environment (such as 2B QTL for VRI and 1A QTL for DON). Most of the identified FHB resistance QTL were also co-localized with one or more agronomic traits such as a QTL on interstitial region of chr 5R for reduced DS, VRI and DON was found pleotropic for reduced PHT. At many pleotropic QTL regions, such as those mentioned above, the favorable alleles were contributed by same parent and these QTL can efficiently be used for marker-assisted selection (MAS) without any disadvantageous results to improve FHB resistance and other agronomic traits simultaneously. The high-density spring triticale SNP map also promises a starting platform for the fine mapping and map-based cloning of major stable QTL identified during the present study."}
+{"text": "Among the variants, the homozygote variant CC genotype of mTOR rs17036508 C>T were associated with higher PCa risk than the wild TT genotypes (adjusted OR = 3.73 (95% CI = 1.75-7.94), P = 0.001). The GT genotype of mTOR rs2295080 G>T was more protective than the TT genotypes (adjusted OR=0.54 (95% CI=0.32-0.91), P=0.020). The distributions of Raptor rs1468033 A>G genotypes differed between cases and controls, especially in subgroups defined by age, BMI, smoking status, and ethnicity. The CT/CC genotypes of AKT2 rs7250897 C>T were associated with an increased risk of PCa, particularly in subgroups of age >71 and BMI >24 kg/m2. These findings suggest that SNPs in the PI3K/AKT/mTOR pathway may contribute to the risk of PCa in Chinese men.In this hospital-based case-control study of 413 prostate cancer (PCa) cases and 807 cancer-free controls, we investigated the role of functional single nucleotide polymorphisms (SNPs) of pivotal genes in the PI3K/AKT/mTOR pathway. We genotyped 17 SNPs in  For Raptor variants, heterozygote genotypes AG of polymorphism rs1468033 A>G were associated with PCa risk compared to genotypes GG (adjusted OR=1.61 (95% CI =1.25-2.06), P <0.001). We also found that rs1468033 A>G was associated with PCa risk . Further analysis of the combined genotypes of these three SNPs and AKT2 rs7250897 C>T showed enhanced PCa susceptibility with increasing numbers of putative high-risk genotypes (Ptrend<0.001) , P = 0.006). Further, the mTOR rs17036508 CC genotypes were associated with increased PCa risk by recessive genetic model for subgroups of age \u226471(adjusted OR=4.75 (95%CI =1.78-12.71), P = 0.002), age >71 (adjusted OR=4.88 (95%CI =1.65-14.38), P = 0.004), BMI \u226424 kg/m2 (adjusted OR=3.20 (95%CI =1.2-8.53), P = 0.02), BMI >24 kg/m2 (adjusted OR=8.11 (95%CI =2.62-25.11), P< 0.001), ever smokers (adjusted OR=6.39 (95%CI =2.46-16.6), P< 0.001), and Uygur population (adjusted OR=5.09 (95%CI =2.2-11.78), P< 0.001). On the contrary, the mTOR rs2295080 GT/GG genotypes were associated with decreased PCa risk by a dominant genetic model in BMI >24 kg/m2 and ever smokers subgroups. However, the mTOR rs2295080 GG genotypes were associated with PCa risk among age subgroups according to the recessive genetic model. For Raptor rs1468033 A>G, AG/AA genotypes were associated with increased PCa risk by the dominant genetic model, particularly in subgroups of age >71(adjusted OR=1.81 (95%CI =1.31-2.48), P = 0.003), BMI >24kg/m2 (adjusted OR=2.02 (95%CI =1.42-2.87), P< 0.001), never smokers (adjusted OR=1.61 (95%CI =1.21-2.30), P = 0.009), ever smokers (adjusted OR=1.58 (95%CI =1.13-2.19), P = 0.068), and Uygur population (adjusted OR=1.66 (95%CI =1.26-2.20), P< 0.001). Furthermore, increased PCa risk was observed by recessive genetic model for the BMI >24 kg/m2 subgroup (adjusted OR=5.13 (95% CI=1.94-13.59), P = 0.001). Increased PCa risk was observed for AKT2 rs7250897 C>T by the dominant genetic model, particularly in subgroups of age >71(adjusted OR=1.83 (95%CI =1.33-2.52), P = 0.002) and BMI >24kg/m2 (adjusted OR=1.58 (95%CI =1.12-2.24), P = 0.010). However, further homogeneity tests showed no difference in risk estimates between subgroups for most strata except age group by mTOR rs17036508 CT/CC genotypes (P=0.032), mTOR rs2295080 GG genotypes (P=0.002), AKT2 rs7250897 CT/CC genotypes (P<0.001) and BMI group by Raptor rs1468033 AA genotypes (P<0.001). The details are shown in Table The mTOR rs17036508 CC and rs2295080 TT, Raptor rs1468033 AG/AA, and AKT2 rs7250897 TT/CT) and four environmental risk factors . Among all 8 factors, the Raptor rs1468033 A>G variant was the best one-factor model. Likewise, interactions between mTOR rs17036508 CC, rs2295080 TT and Raptor rs1468033 AG/AA represented the best three-factor model. Age was the most promising environmental risk factor associated with the four genetic factors. The details are presented in Table We performed the multifactor dimensionality reduction (MDR) analysis by including the genotypes of four significant genetic factors  values at variant prior probability levels for all positive findings Table . Some himTOR rs17036508 CC, mTOR rs2295080 GG/GT, Raptor rs1468033 AG/AA, and AKT rs7250897 TT/CT genotypes were associated with PCa risk, especially in age, BMI, smoker-status, and ethnic subgroups. To the best of our knowledge, this is the first post-GWAS study analyzing associations of these six pivotal genes of PI3K/AKT/mTOR pathway with PCa risk.In the current single institution based case-control study, functional SNPs of six pivotal genes of PI3K/AKT/mTOR pathway were associated with PCa risk. Briefly, the mTOR gene is a critical cellular protein with more than 2651 SNPs reported across the whole region [mTOR rs2295080 GT/GG genotypes protected against PCa risk in Han Chinese populations [mTOR rs2295080 GT/GG genotypes were also observed in esophageal squamous cell carcinoma, gastric carcinoma, and renal cell carcinoma [In vitro and in vivo studies suggested that mTOR rs2295080 T allele probably increased the affinity of special transcription factors to this promoter region and contributed to enhanced mTOR activity [mTOR rs2295080 was linked to eight potential functional variants of mTOR with higher linkage disequilibrium (LD) coefficient >0.8, including rs1064261, rs1074078, rs1135172, rs1883965, rs4845860, rs6540965, and rs6671083. Among these, the variant rs1064261 probably interrupted the exonic splicing enhancer or silencer motif and correlated with neuroendocrine tumors [mTOR, variant rs1883965 was associated with esophageal carcinoma, gastric cancer, and hepatocellular carcinoma [mTOR was located within a miRNA binding site and an exonic splicing enhancer or silencer motif, thereby affecting pre-RNA splicing. The association between rs17036508 and PCa was also observed previously [ANGPTL7), resulting in upregulation of ANGPTL7 by hypoxia in cancer cells, which exerts a pro-angiogenesis effect [mTOR and ANGPTL7 simultaneously.The e region . Howeverulations , 19. Thearcinoma \u201322. In vactivity . Variante tumors , gastrice tumors , and esoe tumors . Similars effect . Taken tRaptor gene, located in 17q25.3 with 34 exons, regulates responses to nutrient and insulin levels [in vitro and in vivo studies have shown that Raptor acts as a scaffold protein that regulates mTOR-dependent signaling [Raptor gene polymorphisms were associated with increased risk for bladder cancer; physical activity, energy balance and genetic variants in the mTOR pathway co-coordinately influenced bladder cancer risk [Raptor rs1468033 was associated with PCa risk, particularly in subgroups of age, BMI, and smoking status, similar to previous findings. Silico analysis indicated that rs1468033 was found with several potential functional polymorphisms of Raptor gene, including rs2292639, rs499609, rs6565500, and rs9899178. Among these, rs2292639, rs4999609, and rs6565500 were predicted as transfactor binding sites, whereas rs9899178 was predicted as splicing site. It is plausible that these variants modulate the expression of Raptor gene, and affect mTOR activity.The n levels . The Rapignaling . One of cer risk . In thisAKT2 was not known. The different AKT isoforms  play contradictory regulatory roles; for example, AKT1 activation inhibits cell migration whereas AKT2 promotes it [et al. reported that AKT2 rs7254617 increased prostate cancer risk [AKT2 rs7250897 was associated with increased PCa risk by a dominant genetic model in subgroups of age >71and BMI >24kg/m2, indicating that rs7250897 altered AKT2 expression, which subsequently affected synthesis of adipose-related proteins. Since the three variants were far from each other, we postulated that they were associated with carcinogenesis by different mechanisms. These findings need to be further validated.Although mTOR activity was previously implicated in promoting PCa cell invasion, the role of motes it , 32. Manmotes it . Additiomotes it . Recentlcer risk . Also, ain vitro and in vivo experiments are necessary to unravel molecular mechanisms for the genetic associations that we have postulated in this study.There were few limitations in the present study that need to be addressed. First, the limited sample size in our study may have decreased detection of weaker genetic effects in carcinogenesis. Second, information regarding predisposition to PCa was not collected for the analysis and might confuse the stratified positive associations. Third, the case-control study may have inherent selection and information biases. Because of insufficient medical records, we could not perform correlative analyses between stages of PCa and variants of AKT pathway that may have provided more information in PCa carcinogenesis. Moreover, further mTOR leading to PCa susceptibility.In summary, we showed that variants of PI3K/AKT/mTOR signal pathway genes may associate with PCa risk. The combined genotypes of these variants enhanced PCa risk with increasing numbers of putative high-risk genotypes. Moreover, the three-factor model  was the best model to predict PCa risk. In conclusion, our study postulated that the genetic variants may alter the expression and activity of We recruited 413 newly diagnosed PCa cases and 807 frequency-matched cancer-free controls from genetically unrelated Chinese Han and Uygur participants in Xinjiang province between January 2003 and January 2015. The cases were histopathologically confirmed as primary prostate adenocarcinoma at the First Affiliated Hospital of Xinjiang medical University (XJMU). Pathological grades of the PCa were determined by Gleason scores from the radical prostatectomy specimens according to the latest WHO criteria . The recAll subjects were interviewed with a written informed consent. Response rate was 92% and 90% for cases and controls, respectively. The experimental and research protocols were approved by the Institutional Review Board of XJMU.http://www.ncbi.nlm.nih.gov/projects/SNP) and SNPinfo web server (http://snpinfo.niehs.nih.gov/snpfunc.htm). The criteria were: 1) minor allele frequency (MAF) was at least 5% in Chinese populations; 2) SNPs were potentially functional according to SNPinfo prediction platform. Ultimately, 17 potentially functional variants were selected involving mTOR , Raptor , AKT1 , AKT2 , PTEN (rs701848 C>T), and K-ras (rs7312175 A>G) based on the bioinformatics analysis performed with HaploView software 4.2. All these SNPs were genotyped by the TaqMan real-time PCR method as described previously [For the six pivotal genes in PI3K/AKT/mTOR pathway, selection strategy for the potentially functional SNPs was based on the NCBI dbSNP database  to identify the best n-factor interaction model [The multifactor dimensionality reduction (MDR) analysis was conducted by the MDR V2.0 beta 8.2 software (on model . We furt"}
+{"text": "Pseudomonas resinovorans strain MO-1, which possesses a high ability to oxidize Mn(II), has been isolated from oligotrophic pond sediment. The draft genome sequence consists of 6,252,942 bp and has a G+C content of 63.4%. Strain MO-1 has 5,694 coding sequences, including 13 putative Mn(II) oxidation genes. The extracted DNA was sequenced using a 101-bp paired-end sequencing method with an Illumina HiSeq 2500 platform at Hokkaido System Science Co., Ltd. , obtaining 22,074,708 reads, with approximately 350-fold genome coverage. After the adaptors were trimmed using the Trimmomatic program version 0.36 (mnxG (locus tag PputGB1_2447 under GenBank accession no. CP000926), mcoA (locus tag PputGB1_2665 under GenBank accession no. CP000926) and mopA (locus tag PputGB1_3353 under GenBank accession no. CP000926) of Pseudomonas putida GB-1 (\u221250)  oxidation genes, much more than the 3 annotated Mn(II) oxidation genes in P. putida GB-1.Two gene families encoding multicopper oxidase (MCO) and heme peroxidase oxidase domains play an important role in Mn(II) oxidation in several MnOB. Of our 5,694 CDSs, 1, 8, and 4 CDSs were identified as being related to Mn(II) oxidation; these were homologous to ida GB-1 , respect-1 (\u221250) . The higlgorithm  are 82.2P. resinovorans strain MO-1 genome sequence has been deposited in DDBJ/EMBL/GenBank under the accession no. BDMA00000000. The version described in this paper is the first version, BDMA01000000.The"}
+{"text": "Escherichia coli J53, which is used as a recipient in conjugation experiments and is a laboratory strain derived from E. coli K-12. This genome sequence will help in the development of a comprehensive genetic analysis of conjugative elements.We report here the complete genome sequence of  Escherichia coli J53 is a laboratory mutant of E. coli K-12  and Oxford Nanopore Technologies MinION (R9.4 flow cell) systems. A circular chromosome was obtained by a http://mlst.warwick.ac.uk/mlst/dbs/Ecoli), similar to other E. coli K-12 strains. A genomic comparison against the K-12 reference genome for substrain MG1655 (GenBank accession no. U00096) using progressiveMauve (1A (2 copies), IS5, proB, proA, four tRNAs, a part of prophage CP4-6, and a part of a repetitive extragenic palindromic sequence (REP321) in 7 segments. A total of 57,947 bp of insertions included IS10R (6 copies), rtT small RNA (sRNA), and the lambda prophage (GenBank accession no. J02459), in 8 segments. J53 contained an 825,667-bp inversion block flanked by 2 copies of IS5 between nucleotide positions 3652638 and 4476957 of the MG1655 genome.The complete genome contains a 4,682,574-bp chromosome with a GC content of 50.8%, 4,530 coding sequences, 82 tRNA-coding genes, and 22 rRNA-coding operons. The strain belongs to sequence type 10, as per the Achtman multilocus sequence typing scheme (E. coli K-12 genomes available on the NCBI Assembly resource (https://www.ncbi.nlm.nih.gov/assembly/) as of 9 April 2018 and the J53_AICK draft genome sequence, we made a phylogenetic tree based on core single nucleotide polymorphisms (SNPs) with kSNP3.0 (secA gene, which is associated with azide resistance (Using the 39  kSNP3.0 . The J53E. coli J53 has been deposited in GenBank under accession no. CP028702.The complete sequence of the chromosome of"}
+{"text": "AbstractGammarus are described and illustrated from Tibetan Plateau. Gammarusaltussp. n. and G.limosussp. n. are characterized by pereopods III\u2013IV with a few short setae and uropod III with marginal spines accompanied by short setae. Gammaruskangdingensissp. n. and G.gonggaensissp. n. are characterized by pereopods III\u2013IV with long straight setae on posterior margins and inner ramus of uropod III 0.4 times as long as outer ramus. Detailed morphological comparisons with related species are discussed. A key to 15 Gammarus species from the Tibetan Plateau and a map of their distributions are provided.Four new species of the genus  PageBreakunderstood, except fishes , Beijing.The specimens were collected along the bank of streams flowing from high mountains with a fine-meshed hand net. Samples were preserved in 95% ethanol in the field and deposited in a -20\u00b0C refrigerator for long-term preservation. The body length of each amphipod was recorded by holding the specimen straight and measuring the distance along the dorsal side of the body from the base of the first antenna to the base of the telson. All dissected appendages were mounted in glycerol on slides. Appendages were drawn using a Leica DM2500 compound microscope equipped with a drawing tube. The specimens are lodged in the Taxon classificationAnimaliaORDOFAMILIAGenusFabricius, 1775Gammaruspulex .PageBreakTaxon classificationAnimaliaORDOFAMILIAhttp://zoobank.org/B32F6C92-AEAB-4A62-BD6F-4916FCC73377IZCAS-I-A0061-1), 11.6 mm, Maniganggo Town , Dege County, Sichuan Province, altitude 4000 m, August 12, 2001, collected by Xianjin Peng. Paratype: female (IZCAS-I-A0061-2), 10.1 mm; paratypes: five males and five females (IZCAS-I-A0061-3), same data as holotype.Holotype: male , 11.6 mm., 10.1mm.: second article of palp with two groups of B-setae (a group of B-setae); bases of pereopods V\u2013VII elongated (broad), narrowing distally (posterior margin of pereopods V and VI nearly straight); uropod III with spines accompanied by simple setae on inner margin (uropod III with spines accompanied by plumose setae); and telson with no medial spines on surface .G.sichuanensis Hou, Li & Zheng, 2002 in peduncle of antenna I and II with two or three groups of setae along anterior and posterior margins, and pereopod IV with a few setae on posterior margin. It differs from G.sichuanensis (G.sichuanensis in parentheses) by pereopod III with short setae on posterior margins PageBreakPageBreakof merus and carpus (with long setae on posterior margins of merus and carpus); bases of pereopods V\u2013VII elongated (broad in G.sichuanensis); uropod III with no plumose setae (both rami with plumose setae on inner and outer margins).This species is similar to Taxon classificationAnimaliaORDOFAMILIAhttp://zoobank.org/2883210F-54D8-4945-95C0-9C9DEE5D0DA2IZCAS-I-A0059-1), 11.8 mm, Erdaohe, Kangding County , altitude 2470 m, August 19, 2001, collected by Xianjin Peng. Paratype: female (IZCAS-I-A0059-2), 8.5 mm; paratypes: two males and three females (IZCAS-I-A0059-3), same data as holotype.Holotype: male , 11.8 mm, slender., 8.5 mm. in antenna II peduncle having short setae along anterior and posterior margins, and calceoli ; uropod III inner ramus 0.4 times the length of outer ramus (inner ramus 0.7 times the length of outer ramus); uropod III terminal article of outer ramus longer than adjacent spines .Gamamruskangdingensis sp. n. is similar to G.altus sp. n. in the shape of gnathopods I and II. It differs from G.altus sp. n.  in having pereopods III and IV with long setae on posterior margins (with a few short setae on posterior margins); and inner ramus of uropod III 0.4 times the length of outer ramus (0.3), inner margins of inner and outer rami with a row of plumose setae (with no plumose setae).Taxon classificationAnimaliaORDOFAMILIAhttp://zoobank.org/ED25CC10-321D-44B1-98F3-E2E30CC99286IZCAS-I-A0065-1), 10.0 mm, Hepinggou, Baoxing County , Gongga Mountains, altitude 2110 m, June 15, 2001, collected by Jinzhong Fu and Yuezhao Wang. Paratype: female (IZCAS-I-A0065-2) 8.6 mm; paratypes: two males (IZCAS-I-A0065-3), same data as holotype.Holotype: male , 10.0 mm., 8.6 mm. by antenna II peduncle having long setae along anterior and posterior margins, calceoli absent ; outer ramus of uropod III with a few plumose setae on inner margin, inner ramus with no plumose marginal setae (with a row of plumose setae on inner margins of inner and outer rami); terminal article of uropod III shorter than adjacent spines (longer than adjacent spines).The new species of Gammarusgonggaensis sp. n. is similar to G.emeiensis Hou, Li & Koenemann, 2002 in antenna II peduncle with long setae along anterior and posterior margins, calceoli absent, and pereopods III and IV with long setae on posterior margins. It differs from G.emeiensisG.emeiensis.Gammarusgonggaensis sp. n. differs from G.altus sp. n.  by anternna II peduncle with long setae, calceoli absent ; pereopods III and IV with long setae on posterior margin (with a few short setae); uropod III with plumose setae on inner margin of inner ramus (with no plumose setae), terminal article shorter than adjacent spines (longer than adjacent spines).Taxon classificationAnimaliaORDOFAMILIAhttp://zoobank.org/9ACD4C39-16EC-47AF-A3B0-C32C3BA8EAC3IZCAS-I-A0063-1), 8.6 mm, Baxoi County , Tibet, altitude 4400 m, August 21, 2001, collected by Xianjin Peng. Paratype: female (IZCAS-I-A0063-2), 5.5 mm; paratypes: five males and five females (IZCAS-I-A0063-3), same data as holotype.Holotype: male , 8.6 mm., 5.5 mm. by pereopod III with few setae on posterior margin (merus with four groups of setae on posterior margin); epimeral plate III acute on posterodistal corner (blunt); and uropod III inner ramus longer than half of outer ramus length (inner ramus 0.3 times the length of outer ramus).Gammaruslimosus sp. n. is very similar to G.balcanicus Sch\u00e4ferna, 1922 (widespread in Europe). It differs from the latter by pereopods III\u2013IV and uropod III having few setae; bases of pereopods VI and VII slender.Gammarus recorded from the Tibetan Plateau can be classified into four groups based on morphological comparison: (1) G.lacustris group, is characterized by uropod III inner ramus longer than half of outer ramus length, both rami fringed with plumose setae, and includes five species: G.lacustris Sars, 1863, G.lasaensis Barnard & Dai, 1988, G.hongyuanensis Barnard & Dai, 1988, G.frigidus Hou & Li, 2004, and G.jaspidus Hou & Li, 2004; (2) cave species with no eyes, including G.abstrusus Hou, Platvoet & Li, 2006, and G.praecipuus Li, Hou & An, 2013; (3) G.sinuolatus Hou & Li, 2004 with long simple setae on uropod III; and (4) G.kangdingensis group is characterized by uropod III having a few simple or plumose setae, and includes G.emeiensis Hou, Li & Koenemann, 2002, G.sichuanensis Hou, Li & Zheng, 2002, G.glaber Hou, 2017, Gammarusaltus sp. n., G.kangdingensis sp. n., G.gonggaensis sp. n., and G.limosus sp. n. A key to these species is presented as follows.The species of the genus"}
+{"text": "MLL) gene produce fusion proteins such as MLL-AF4 that disrupt epigenetic pathways and cause poor-prognosis leukemias. Here, we find that at a subset of gene targets, MLL-AF4 binding spreads into the gene body and\u00a0is associated with the spreading of Menin binding, increased transcription, increased H3K79 methylation (H3K79me2/3), a disruption of normal H3K36me3 patterns, and unmethylated CpG regions in the gene body. Compared to other H3K79me2/3 marked genes, MLL-AF4 spreading gene expression is downregulated by inhibitors of the H3K79 methyltransferase DOT1L. This sensitivity mediates synergistic interactions with additional targeted drug treatments. Therefore, epigenetic spreading and enhanced susceptibility to epidrugs provides a potential marker for better understanding combination therapies in humans.Understanding the underlying molecular mechanisms of defined cancers is crucial for effective personalized therapies. Translocations of the mixed-lineage leukemia (  \u2022MLL-AF4 binding requires an unmethylated CpG (uCpG) island and Menin\u2022MLL-AF4 and Menin can spread into the gene body of some targets\u2022Spreading targets are highly transcribed and have an aberrant chromatin signature\u2022Spreading of MLL-AF4 is a predictor of sensitivity to DOT1L inhibitors Translocations of the MLL gene produce fusion proteins such as MLL-AF4 that cause poor-prognosis leukemias. Kerry et\u00a0al. show that MLL-AF4 can spread into the gene body of some target genes. Spreading targets have an aberrant chromatin signature and are sensitive to DOT1L inhibitors. MLL) gene produce over 120 different MLL fusion proteins (MLL-FPs) that cause aggressive acute leukemias, the most common one being the MLL-AF4 fusion  at target genes, an epigenetic mark associated with gene activation . H3K79MeHere, we reveal a strong co-dependent relationship between MLL-AF4 and Menin binding at a small number of target genes containing uCpGs. At a subset of these gene targets, we observe MLL-AF4 and Menin spreading that is bookended by uCpGs. These spreading targets are distinct from super-enhancers, are associated with high levels of gene transcription, have an aberrant H3K79me2/H3K36me3 signature, and are predictive of a poor overall survival in patients with acute lymphoblastic leukemia (ALL). These gene targets also display a remarkable dependence on H3K79me2 and the fusion protein for their sustained expression in leukemia. Together, this work shows that MLL-FP spreading occurs at genes important in MLL leukemogenesis and has the potential to act as a biomarker for therapeutic response.Using MLL(N) and AF4(C) chromatin immunoprecipitation sequencing (ChIP-seq) in the human MLL-AF4 SEM cell line C, we ideIf the CXXC domain is essential for MLL-AF4 recruitment, we would expect all MLL-AF4 binding sites to occur at regions of uCpGs. To test this, we used a biotinylated CXXC affinity purification (Bio-CAP) assay  for highTo determine whether other CXXC domain-containing proteins (\u201cCXXC proteins\u201d from now on) are also restricted to only a proportion of uCpG sites, we performed ChIP-seq for CFP1 (a CXXC protein member of the SET1 complex associated with gene activation), and KDM2B (a CXXC protein involved in the recruitment of the polycomb group repressive complex [PRC] ), in SEM2\u00a0= 0.96)  I, wherea\u00a0= 0.96) J and S1HChIP-seq on two members of PAFc, PAF1 and LEO1 C, overlaLedgf siRNA  system see A legend gf siRNA F and S2GNA level F.HOXA9 locus  gene target isoforms  ChIP-seq in the MLL-AF6 cell line ML-2 detects the fusion protein unambiguously due to a deletion of the wild-type isoforms , and simisoforms B. The hiisoforms . ER-taggisoforms , biotin-isoforms , MLL-AF4isoforms , MLL-AF4, at wild-type MLL binding sites in SEM cells C and 4D,ARID2, JMJD1C, MBNL1, MEF2C, or RUNX2 alone is associated with at least one indicator of poor prognosis in ALL patients .Cell lines, culture methods, and drug treatment protocols used in this study are listed in Western blot analysis was performed as previously described . AntibodChIP and ChIP-seq experiments were performed as described in MLL-AF4, Menin, and PAF1 cDNA were inserted downstream of the FS2-TetR coding sequence in the original pCAGFS2TetR vector, by ligation-independent cloning (LIC). Plasmids were transfected into TetO mESCs using Lipofectamine 2000, and clones stably expressing TetR fusions were selected using puromycin (1\u00a0\u03bcg/mL), or MLL-AF4 cDNA was transiently transfected into mESCs at 60%\u201370% confluency using Lipofectamine 2000. Cells were collected 24\u00a0hr after transfection.For the TetR recruitment assay, we used the previously engineered Tet-operon (TetO) mESC line . MLL-AF4ChIP-seq peaks were called as described in Nascent RNA-seq and gene expression analysis was performed as described in Clinical datasets and survival analyses are detailed in GSE13313, GSE28460, GSE29130, GSE34861, GSE73528, GSE74812, and GSE84116; and ArrayExpress: E-MTAB-3593 (for a detailed list of datasets, see Data were analyzed using Fisher\u2019s exact test, Wilcoxon test, and Mann-Whitney U test, where appropriate. Results were deemed significant if p\u00a0< 0.05. Unless otherwise indicated, data are shown as mean \u00b1 SD. This paper analyzed datasets from GEO: Conceptualization, T.A.M. and J.K.; Formal analysis, J.K., E.R., H.G., and H.M.; Investigation, J.K., L.G., M.T., N.P.B., H.M., E.B., T.A.M.; Resources, S.O., F.P., O.H., A.R., I.R.; Writing \u2013 Original draft, T.A.M. and J.K.; Writing \u2013 Review & Editing, T.A.M., J.K., O.H., A.R., I.R., M.K., R.J.K., H.G.; Visualization, T.A.M. and J.K.; Supervision, T.A.M., M.K., and R.J.K.; Funding Acquisition, T.A.M., O.H., A.R., I.R., R.J.K., M.K."}
+{"text": "Riemerella anatipestifer, the tetracycline susceptibility of 212 R. anatipestifer isolates from China between 2011 and 2017 was tested. The results showed that 192 of 212 (90.6%) R. anatipestifer isolates exhibited resistance to tetracycline (the MICs ranged from 4 to 256 \u03bcg/ml). The results of PCR detection showed that, 170 of 212 (80.2%) R. anatipestifer isolates possessed the tet(X) gene. Other genes, including tet(A), tet(M), tet(Q), tet(O), tet(B), and tet(O/W/32/O), were found at frequencies of 20.8, 4.7, 1.4, 0.9, 0.9, and 0.5%, respectively. However, tet(C), tet(E), tet(G), tet(K), and tet(W) were not detected in any isolate. In these tet gene positive strains, 31 (14.6%), 2 (0.9%), 5 (2.4%), 1 (0.5%), 3 (1.4%) were detected containing tet(A)/tet(X), tet(M)/tet(O), tet(M)/tet(X), tet(O)/tet(X), and tet(Q)/tet(X) simultaneously, respectively. One isolates, R131, unexpectedly contained three tet genes, i.e., tet(M), tet(O), and tet(X). Sequence analysis of the tet gene ORFs cloned from R. anatipestifer isolates confirmed that tet(A), tet(B), tet(M), tet(O), tet(Q) and an unusual mosaic tet gene tet(O/W/32/O) were present in R. anatipestifer. The MIC results of R. anatipestifer ATCC 11845 transconjugants carrying tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), tet(Q), and tet(X) genes exhibited tetracycline resistance with MIC values ranging from 4 to 64 \u03bcg/ml. Additionally, the tet(X) gene could transfer into susceptible strain via natural transformation (transformation frequencies of ~10\u22126). In conclusion, the tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), tet(Q), and tet(X) genes were found and conferred tetracycline resistance in R. anatipestifer isolates. Moreover, the tet(X) is the main mechanism of tetracycline resistance in R. anatipestifer isolates. To our knowledge, this is the first report of tet(A), tet(B), tet(M), tet(O), tet(Q), and mosaic gene tet(O/W/32/O) in R. anatipestifer.To investigate tetracycline resistance and resistant genotype in  In addition, an unknown gene tet(U) was reported to be located on the pKq10 plasmid in Enterococcus faecium  gene was reported to be located on the pRA0511 plasmid in R. anatipestifer  was reported to be the main mechanism of tetracycline resistance in R. anatipestifer isolates , tet(B), tet(O), tet(O/W/32/O), tet(Q) in R. anatipestifer and their resistant function were confirmed by transferring into tetracycline-susceptible R. anatipestifer ATCC 11845.To determine the tetracycline resistance mechanisms of R. anatipestifer field isolates were isolated from 58 large-scale duck farms in different regions of China between 2011 and 2017, including Sichuan, Jiangsu, Guizhou, Anhui, Guangdong, Chongqing, Guangxi, Hainan, Jiling, Henan and Beijing provinces. Under sterile conditions, the brains, hearts or livers were collected from infected or died ducks, and samples were streak-inoculated on tryptic soybean agar plates  supplementary with 10% sheep-blood. A single colony from one duck was purified and cultured repeatedly. A total of 212 isolates were identified as R. anatipestifer by PCR amplifying 16S rRNA and sequencing and biochemical analyses. R. anatipestifer strains were cultured at 37\u00b0C in GC broth (GCB) or on GC agar (GCA) plates  strains DH5\u03b1 and S17-1 were grown at 37\u00b0C in Luria-Bertani  broth or on LB agar. When required, antibiotics were added as follows: 5 \u03bcg/ml tetracycline (TET); 2 \u03bcg/ml cefoxitin (FOX); 40 \u03bcg/ml kanamycin (KAN) or 100 \u03bcg/ml ampicillin (AMP). All antibiotics were obtained from Dalian Meilun Biotech Co., Ltd. .The bacteria and plasmids used in this study are listed in Table 6 CFU/ml (100 \u03bcl/well). The inoculated micro-plates were incubated at 37\u00b0C for 24 h. E. coli ATCC 25922 was used as a quality control strain. The experiments were repeated at least three times. Due to the lack of CLSI-approved tetracycline breakpoints applicable to R. anatipestifer  in 96-well microtiter-plates  according to Clinical and Laboratory Standards Institute (CLSI) guideline specific for bacteria isolated from animals CLSI, . The finp. CLSI, , i.e., stet) characterized previously in other bacteria were determined in all R. anatipestifer field isolates by PCR , tet(B), tet(C), tet(E), tet(G), tet(K)], 4 ribosomal protection genes , enzymatic inactivation gene tet(X), and a mosaic tetracycline resistance gene tet(O/W/32/O). The primers and protocols described previously  was used as positive control. Since the absence of available positive controls for other tet genes, all PCR products were size-confirmed by electrophoresis on a 1.5% agarose gel. All amplicons of tet(B), tet(M), tet(O), tet(O//W/32/O), and tet(Q) from R. anatipestifer isolates were verified by sequencing , while only 12 amplicons of tet(A) were sequenced.The PCR templates were prepared by treating the bacteria with lysis buffer  and boiled for 10 min. The presence of the tetracycline resistance genes (tet(X) gene was amplified by PCR from R. anatipestifer CH-2 using the primer sets tet(X)-F2/tet(X)-R2, listed in Table tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), and tet(Q) gene were also amplified from tet-positive isolates confirmed above using the additional primers that were designed based on the previously described tet sequences in other bacteria (Table tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), and tet(Q) genes, respectively. The purified amplicons were sequenced and digested with restriction enzymes, and then ligated to the same digested shuttle vector pLMF03  and KAN (40 \u03bcg/ml) and identified by PCR. At the same time, the negative control empty vector pLMF03 was transferred into ATCC 11845, resulting in the transconjugant ATCC 11845 (pLMF03). The MICs of tetracycline for the wild-type and transconjugants were measured as described above.The correct recombinant plasmids were transferred into tet(X) gene was verified by natural transformation as described previously  were respectively used as donor DNA and transferred into the recipient strain R. anatipestifer ATCC 11845. The transformants, designated ATCC 11845 [tet(X)], were screened using GCB plates supplemented with TET (5 \u03bcg/ml). The insertion of the transferred tet(X) genes was verified by PCR and sequencing. The tetracycline resistance phenotypes of transformant was determined as described above.The transferability of the tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), and tet(Q) genes in R. anatipestifer isolates R100, R98, R131, R96, and R159 in this study have been deposited in GenBank under the accession numbers from MF969099 to MF969104, respectively.The sequences of All animals studies were conducted in strict accordance with the recommendations of the Guide for the Care and Use of Laboratory Animals, National Research Council. The animal-use procedures were approved by the Animal Ethics Committee of the Sichuan Agricultural University .R. anatipestifer field isolates were resistance to tetracycline by susceptibility test. The MICs of tetracycline in 212 R. anatipestifer field isolates ranged from 0.25 to 256 \u03bcg/ml  was detected in 170 of 212 isolates, exhibiting the highest rate of occurrence among tet genes (80.2%). The detected frequency of other tet genes was as followed: tet(A) (20.8%), tet(M) (4.7%), tet(Q) (1.4%), tet(O) (0.9%), tet(B) (0.9%), and tet(O/W/32/O) (0.5%). However, tet(C), tet(E), tet(G), tet(K), and tet(W) were not detected in any of 212 tested R. anatipestifer field isolates (Table tet genes simultaneously (Table tet(X) was detected simultaneously along with tet(A), tet(M), tet(O), and tet(Q) in 31(14.6%), 5 (2.4%), 1 (0.5%), and 3 (1.4%) isolates, respectively. Two isolates (0.9%) carried tet(M) and tet(O) genes simultaneously. Moreover, one isolate (0.5%) had three commensal tet genes, tet(M), tet(O), and tet(X), respectively. The R. anatipestifer field isolates containing one or more tet genes exhibited tetracycline resistance with MICs ranging from 4 to 256 \u03bcg/ml, while no tet genes were detected in 19 tetracycline-susceptible isolates and 1 intermediately resistant isolate. Although several isolates carried more than one tet genes in this study, they did not display higher MIC values. Thus, the positive rate of genotype was in line with that of tetracycline resistance phenotype.To assess the prevalence of the R. anatipestifer isolates selected for ORFs sequencing were listed in Table tet(A) gene cloned from six different R. anatipestifer isolates shared 99\u2013100% identity (Table MF969099) showed 99% sequence identity with the tet(A) gene from transposon Tn1721  gene cloned from R. anatipestifer isolates showed 99% identity each other. The one in R98 isolate (GenBank accession no. MF969100) showed 100% identity with tet(B) gene from transposon Tn10  genes from five different R. anatipestifer isolates shared 99\u2013100% sequence homology. The one in R133 isolate (GenBank accession no. MF969101) exhibited 96 and 95% sequence identity with tet(M) genes from Streptococcus pneumoniae  genes cloned from R. anatipestifer isolates were 100% identity. The tet(O) gene in R131 isolate (GenBank accession no. MF969102) exhibited 99% sequence identity with the tet(O) genes from Streptococcus mutans DL5  from R. anatipestifer isolate R96 (GenBank accession no. MF969103) was a mosaic tetracycline resistance gene derived from tet(O) and tet(W) genes. Sequence alignment result showed that it had 97 and 99% sequence identity with the tet(W/32/O) gene from Bifidobacterium thermophilum (GenBank accession no. AM710601)  gene from Streptococcus suis integrative conjugative element ICESsu32457 (GenBank accession no. FR823304)  gene from three different R. anatipestifer isolates shared 100% identity. The tet(Q) gene in R159 isolate (GenBank accession no. MF969104) exhibited 97% sequence identity with the tet(Q) gene from Bacteroides thetaiotaomicron  gene in R. anatipestifer CH-2  gene previously reported on the pRA0511 plasmid in R. anatipestifer , tet(B), tet(M), tet(O), tet(O/W/32/O), tet(Q), and tet(X) genes were responsible for tetracycline resistance in R. anatipestifer, recombinant plasmids carrying these genes amplified from different R. anatipestifer isolates were transferred into the tetracycline-susceptible R. anatipestifer ATCC 11845 by conjugative transfer. The tetracycline MICs for all transconjugants ranged from 4 to 64 \u03bcg/ml (Table tet(A), tet(B), or tet(O) genes showed low-level tetracycline resistance, and their MIC values were 4\u20138 \u03bcg/ml. The transconjugants containing tet(M), tet(O/W/32/O), or tet(X) genes exhibited tetracycline resistance of 32 \u03bcg/ml. The tetracycline resistance of ATCC 11845 [pLMF03-R159-tet(Q)] was the highest among all transconjugants .To further verify whether the identified tet(X) gene, we transferred the tet(X) gene from R. anatipestifer CH-2 to ATCC 11845 by natural transformation. The results showed that the tet(X) gene could be successfully transferred into tetracycline-susceptible ATCC 11845. The maximum transformants were obtained with transformation frequency about ~10\u22126 for 5 \u03bcg R. anatipestifer CH-2 genome. Compared with wild-type strain ATCC11845, the transformant ATCC 11845 [tet(X)] exhibited 128-fold increased tetracycline resistance  gene could confer tetracycline resistance and be easily transferred by natural transformation in R. anatipestifer.To study the transferability of the R. anatipestifer isolates in China. The results of antimicrobial susceptibility showed that tetracycline resistance in R. anatipestifer was widespread in China between 2011 and 2017, although the usage of this antibiotic treatment was decreased in the avian industry. By PCR detection, the genotypes of tetracycline resistance were abundant in our investigated R. anatipestifer isolates, including efflux genes [tet(A) and tet(B)], ribosomal protection genes , enzymatic gene tet(X), and mosaic tetracycline resistance gene tet(O/W/32/O). The total rate of positive resistance genes was as high as 90.6%. The tet(X) gene had the highest occurrence frequency and was the dominant mechanism conferring tetracycline resistance in R. anatipestifer isolates. Unexpectedly, this conclusion was in contrast to previous reports that ribosomal protection and efflux pumps were the classical tetracycline resistance mechanisms in other bacteria  detected in this study was also in contrast to the previous conclusion that tet(C) was the main mechanism of tetracycline resistance in R. anatipestifer isolates , tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), and tet(Q) genes were detected in R. anatipestifer.Tetracyclines have been widely used for disease treatment and growth promotion in livestock Roberts, . In thistet genes or an isolate may contain different tet genes on different plasmids or some tet genes on plasmid and other tet genes in the chromosome  or tet(S) gene was reported to accompany with tet(M) gene /tet(B), tet(A)/tet(C), and tet(A)/tet(D)] /tet(C), and tet(C)/tet(M)]. In this study, multiple tet genes were also found in one R. anatipestifer isolate, such as tet(A)/tet(X), tet(M)/tet(O), tet(M)/tet(X), tet(O)/tet(X), tet(Q)/tet(X), and tet(M)/tet(O)/tet(X). This phenomenon might be due to strong selective pressure and horizontal gene transfer among the various bacteria , the tet(X), tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), and tet(Q) genes were most likely located in the chromosome of R. anatipestifer. We confirmed that the tet(X) gene could be transferred by natural transformation, and this transferability might contribute to its wide dissemination in R. anatipestifer.The Roberts, . In genetet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), tet(Q), and tet(X) genes to susceptible strain, we verified that all tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), tet(Q), and tet(X) genes were functional and conferred tetracycline resistance in R. anatipestifer. Interestingly, tet genes detected in this study have high identities with others reported previously, while they exhibited comparatively lower tetracycline resistance in R. anatipestifer , tet(B), and tet(O) conferred the tetracycline resistance with the MIC values from 4 to 8 \u03bcg/ml. We speculated that the low-level of resistance in R. anatipestifer might result from the low-level of tet gene expression, which needed to be further illustrated.Finally, through transferring the tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), and tet(Q) tetracycline resistance genes in R. anatipestifer. We confirmed that all the tet(A), tet(B), tet(M), tet(O), tet(O/W/32/O), tet(Q), and tet(X) confer the tetracycline resistance in R. anatipestifer. The predominant tetracycline resistance mechanism in R. anatipestifer is conferred by tet(X) gene.To the best of our knowledge, this is the first report of the presence of tet genes in R. anatipestifer isolates. D-KZ, X-XZ, and R-YJ cloned and sequenced the tet genes. SC, K-FS, and QY analyzed the sequences of tet genes. H-YL, YW, and X-YC transferred the shuttle vectors. D-KZ, H-YL, and M-SW performed the natural transformation of tet(X) gene. HL, D-KZ, and A-CC drafted and revised the manuscript. All authors have read and approved the final version of the manuscript.D-KZ, M-SW, and A-CC conceived and designed the project. H-YL, M-FL, and M-SW performed the tetracycline susceptibility test and prevalence of The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Case Reports in Psychiatry has retracted the article titled \u201cCombined Case of Blood-Injury-Injection Phobia and Social Phobia: Behavior Therapy Management and Effectiveness through Tilt Test\u201d . The artHeimberg RG et al.: Psychometric properties of the Liebowitz Social Anxiety Scale. Psychological Medicine, 1999, 29, 199\u2013212 (not cited);Ayala ES, Meuret AE, Ritz T: Treatments for blood-injury-injection phobia: a critical review of current evidence. J Psychiatr Res. 2009 Oct; 43(15):1235-42. doi: 10.1016/j.jpsychires.2009.04.008;Lars-G\u00f6ran \u00d6st, Ulf Sterner: A specific behavioral method for treatment of blood phobia. Behaviour Research and Therapy Volume 25, Issue 1, 1987, Pages 25\u201329 doi: 10.1016/0005-7967(87)90111-2;Lars-G\u00f6ran \u00d6st, Anita Jerremalm, Jan Johansson: Individual response patterns and the effects of different behavioral methods in the treatment of social phobia. Behaviour Research and Therapy. Volume 19, Issue 1, 1981, Pages 1\u201316 doi: 10.1016/0005-7967(81)90107-8;Ryan C. Shorey and Gregory L. Stuart: Manualized Cognitive-Behavioral Treatment of Social Anxiety Disorder: A Case Study. Clin Case Stud. 2012 Feb 1; 11(1): 35\u201347. doi: 10.1177/1534650112438462 (not cited)."}
+{"text": "Spiroplasma citri causes stubborn disease in Citrus spp. and diseases in other plants. Here, we report the nucleotide sequence of the 1,599,709-bp circular chromosome and two plasmids of S. citri strain R8-A2T. This information will facilitate analyses to understand spiroplasmal pathogenicity and evolutionary adaptations to lifestyles in plants and arthropod hosts. Sympt\u2013coplasma \u201313. Onlyoplasma \u2013, 15, fors coined , was thema citri , 17.Spiroplasma kunkelii CR2-3X (S.\u00a0citri strain R8-A2T (Morocco-R8-A2 [ATCC 27556T]), which was originally isolated from stubborn diseased citrus [Citrus sinensis (L.) Osbeck var. Washington navel] , CP013198 (plasmid pSCI26), and CP013199 (plasmid pSCI15). The sequence versions described in this paper are CP013197.1, CP013198.1, and CP013199.1, respectively.This genome project has been deposited in GenBank under BioProject ID PRJNA296877, BioSample accession number SAMN04110376, and GenBank accession numbers"}
+{"text": "Psammomys obesus as an animal model of diet-induced type 2 diabetes when subjected to a hypercaloric regimen.Type 2 diabetic retinopathy is the main cause of acquired blindness in adults. The aim of this work was to examine the retinal function of the sand rat Psammomys obesus by high caloric diet . The visual function of control (n = 7) and diabetic (n = 7) adult rodents were followed up during 28 consecutive weeks with full-field electroretinogram(ERG) recordings evoked to flashes of white light according to the standard protocol of the International Society for Clinical Electrophysiology of Vision protocol (ISCEV).Hyperglycemia was induced in Twenty-eight weeks following the induction of diabetes, results revealed significantly reduced and delayed photopic and scotopic ERG responses in diabetic rats compared to control rats. More specifically, we noted a significant decrease in the amplitude of the dark-adapted 0.01ERG (62%), a- and b-wave amplitudes of the dark-adapted 3.0 ERG  and the four major oscillatory potentials components (OP1-OP4) . In photopic conditions, diabetic rats showed a significant decrease in a- and b-wave , photopic negative response (55.3%), 30 Hz flicker (63.7%), OP1-OP4 and S-cone (34.7%). Significantly delayed implicit times were observed for all ERG components in the diabetic animals. Results obtained are comparable to those characterizing the retinal function of patients affected with advanced stage of diabetic retinopathy.Psammomys obesus is a useful translational model to study the pathophysiology of diabetic retinopathy in order to explore new therapeutic avenues in human patients. Diabetic retinopathy (DR), which can lead to blindness in severe cases, is reported to affect more than 90% of diabetic patients . The patPsammomys obesus (P.obesus) fed with a high caloric diet will spontaneously develop type 2 diabetes, with accompanying retinopathy, thus making them a valid translational model of this retinal disorder . Of interest, in the present study, all P.obesus which developed DR after 28 weeks of high-fat treatment also developed a retinal dysfunction as measured with the ERG.Electrophysiological evaluation after the morphological assessment in diabetic P.obesus  was needinopathy , 32, 33.inopathy  on the msubjects , making P.obesus. These functional changes suggest that photoreceptor function, as well as synaptic transmission from the photoreceptors to bipolar cells, were affected in vivo by the high-fat induced hyperglycemia [P.obesus showed a decreased expression of both PKC \u03b1 and \u03b6 isoforms that are Ca2+ independent and co-localized in rod bipolar cells[P.obesus retina. Another alteration detected in diabetic retina in our previous study affecting the synaptic terminals by an increase in synaptic proteins such as synaptophysin [P.obesus viewed as an indicator of retinal cell stress [in vitro by Baccouche et al. [P.obesus were present for both scotopic (rod-mediated) and photopic (cone-mediated) conditions, indicating that both retinal systems are affected in our animal model of diabetic retinopathy. Based on ERG abnormalities and histologic findings in advanced human DR, a significant decrease in amplitudes of scotopic a- and b- waves from pre-proliferative DR [P.obesus at 28 weeks after diabetes induction. A calculation of b-/a-wave amplitude ratios in the present study revealed a declined in scotopic condition, suggesting that bipolar cells are more affected than photoreceptors. However, in photopic condition the unchanged ratio showed that the decrease of b-wave amplitude was correlated to the photoreceptoral response.Our results demonstrated that the amplitude of a- and b- waves ERG of diabetic rodents at 28weeks were significantly reduced, and the implicit times were considerably delayed compared to control glycemia , 34. Thelar cells. This detophysin . The uprl stress . These ce et al. . ERG abnative DR  indicateative DR . The darative DR  is reporative DR \u201341. Thusative DR . Chung eative DR  reportedative DR \u201346 and iP.obesus may be explained by the loss of amacrine cells numbers at different cell retinal layers including retinal ganglion cells [P.obesus ERG were comparable to what is reported in human patients at the proliferative stage in diabetic retinopathy [P.obesus.Our results also provide evidence of retinal impairments beyond the level of the photoreceptors in rodents with diabetes, given that OPs were shown to signal inner retinal activity, particularly the amacrine cells , 48. Alton cells . These con cells , 49\u201351, inopathy \u201354. Thisinopathy , 55\u201357, P.obesus model mimics several of the DR features observed in human diabetes and should therefore be considered as a valid alternative to test therapeutic pharmacological molecules on type 2 DR functions.Currently available diabetic rodent models can be used to study the initial (acute) or latent phase of diabetic retinopathy and several studies have reported a concurrent change in OPs , 58, 59.P.obesus after 7 months of diabetic induction.The i-wave of the human full-field photopic ERG response is a relatively small positive deflection following the b-wave and is thought to originate in an off-circuitry in the inner retina . AlthougP.obesus were previously reported by our laboratory showing a reduction by 44% of RGCs in diabetic P.obesus retina after 7 months of diabetes progression [Degeneration of the retinal ganglion cells (RGC) and deterioration of the inner nuclear layer in the retinas of diabetic gression . In the gression , 69. In gression , 70, 71.P.obesus was reduced in amplitude and delayed in timing, similar to what was previously reported in human diabetes [P.obesus. Recognition of these defects will enhance our understanding of the pathophysiology of diabetic retinopathy.Our results also showed that the photopic flicker ERG of our diabetic diabetes , 72, 73.diabetes . The timdiabetes  describeP.obesus was significantly reduced in amplitude and delayed in timing. The cone short-wavelength opsin staining was detectably reduced in P.obesus diabetic retinas and this was confirmed by Western blot analysis in our previous study [Finally, as previously shown in diabetic human subjects , 76, theus study .P.obesus and the corresponding changes in human DR and support the notion that it represents a valid translational model to study the retinal pathophysiological processes involved in the onset and progression of type 2 diabetic retinopathy.\u201dIn summary, the totally of our findings regarding ERG presented in this report further confirms the similarity between the characteristics of retina changes in diabetic P.obesus. Such studies should include fundus photography, optical coherence tomography and fluorescein angiography in order to clarify to what extent the ERG findings correlate (precede or follow) with morphological changes in the retina.Clearly, more studies are needed to better correlate structural and functional changes in the diabetic retina of The present study clearly demonstrated for the first time that long-lasting and significant alterations in visual function detected by full-field ERG take place after 28 weeks of diet-induced type 2 diabetes in the retina of the sand rat. Thus, the diabetic sand rat appears to be an animal model that mimics several important features of the human form of diabetic retinopathy.P.obesus as a useful translational model to study diet-induced type 2 diabetic retinopathy . We strongly believe that adding this new model to the researchers\u2019 armamentarium will not only be instrumental in increasing our understanding of the pathophysiology of human diabetic retinopathy but also help in the development of new therapeutic strategies.Our results confirm the validity of the S1 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S2 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S3 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S4 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S5 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S6 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S7 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S8 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S9 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S10 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S11 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file.S12 Fig2 flash. (B) Mixed response using 3 cd.s/m2 flash. (C) Photopic responses using 3 cd.s/m2 flash. (D) Photopic 30 Hz flicker response. (E) Photopic S-cone response using 0.0045cd.s/m2 blue flash on an orange background.Representative traces indicating: (A) Rod responses using 0.01 cd.s/m(TIF)Click here for additional data file."}
+{"text": "Microbacterium hominis 84-0209T and M.\u00a0laevaniformans 91-0039 were studied. Genome sizes  were 3,506,522 (70.96%) and 2,999,965 (69.51%), respectively. Annotation revealed: (M.\u00a0hominis) three rRNA sequences, 45 tRNA genes, and 3,218 coding sequences; (M.\u00a0laevaniformans) three rRNA sequences, 49 tRNA genes, and 2,874 coding sequences.Draft genomes for  Microbacterium are environmentally derived, Gram stain-positive bacilli which infrequently cause opportunistic infections in humans (T (CDC group A-4) and LCDC 91-0039 (CDC group A-5) were shared by Health Canada\u2019s Laboratory Centre for Disease Control  with European and Japanese colleagues as part of an effort to assign provisionally named bacteria to validly published genera. Many CDC group A-4 and A-5 isolates were assigned to the genus Microbacterium in 1995 after 16S rRNA gene sequencing analysis  as well as M.\u00a0laevaniformans LCDC 91-0039  (T and LCDC 91-0039.Members of the genus n humans . Some ofn humans . LCDC 84DNA was extracted using the TruseqR DNA HT sample preparation kit. Paired-end whole-genome shotgun libraries were constructed using TruseqR Nano DNA HT library preparation kit and mate-pair libraries for each strain were prepared using the Nextera mate-pair sample preparation kit (Illumina). Sequencing was performed using a MiSeq sequencer (Illumina).M.\u00a0hominis and M.\u00a0laevaniformans generated 2,296,026 and 1,583,940 reads; mate-pair libraries gave rise to 2,272,076 reads and 2,768,080 reads, respectively. Overlapping paired-end reads were merged using FLASH (fast length adjustment of short reads)  predicted three rRNA sequences, 45 tRNA genes, and 3,218 coding sequences. Data was highly consistent (>99% symmetric identity [SI]) with an unpublished genome of M.\u00a0hominis NBRC 15708T (Genbank [GB] accession no. NZ_BCW100000000.1); both genomes were not closely related (63% SI) to GB NZ_JWSZ00000000.1. Annotation of M.\u00a0laevaniformans using Prokka  and repeat sequences; mate-pair reads were added to contigs using Sspace (v2.0.0) to produce scaffolds . This reM.\u00a0hominis LCDC 84-0209T and M.\u00a0laevaniformis LCDC 91-0039 have been deposited at DDBJ/EMBL/GenBank under accession no. LRYC00000000 and LRAD00000000.Draft whole-genome projects for"}
+{"text": "Escherichia coli (ETEC) is a major cause of diarrhoea in children below 5 years of age in endemic areas, and is a primary cause of diarrhoea in travellers visiting developing countries. Epidemiological analysis of E. coli pathovars is traditionally carried out based on the results of serotyping. However, genomic analysis of a global ETEC collection of 362 isolates taken from patients revealed nine novel O-antigen biosynthesis gene clusters that were previously unrecognized, and have collectively been called unclassified. When put in the context of all isolates sequenced, one of the novel O-genotypes, OgN5, was found to be the second most common ETEC O-genotype causing disease, after O6, in a globally representative ETEC collection. It\u2019s also clear that ETEC OgN5 isolates have spread globally. These novel O-genotypes have now been included in our comprehensive O-genotyping scheme, and can be detected using a PCR-based and an in silico typing method. This will assist in epidemiological studies, as well as in ETEC vaccine development.Enterotoxigenic  CF, colonization factor; ETEC, enterotoxigenic Escherichia coli; LT, heat-labile toxin; O-AGC, O-antigen biosynthesis gene cluster; OgUT, O-genotype untypeable; ST, heat-stable toxin; STEC, Shiga toxin-producing Escherichia coli.Escherichia coli isolates were deposited in GenBank/EMBL/DDBJ under the accession numbers shown in Table S1 .1. Assembled draft genomes for 55 enterotoxigenic wzy genes from positive control strains  were deposited in GenBank/EMBL/DDBJ under the accession numbers LC177546\u2013LC177554 and LC223608\u2013LC223610, respectively.2. Sequences of nine annotated O-antigen biosynthesis gene clusters and three Escherichia coli (ETEC) isolates. The novel O-genotype OgN5 was found to be the second most common ETEC O-genotype globally, with no prior information regarding the contribution to the burden of disease. To gain more information about trends in ETEC OgN epidemiology, further studies of global OgN isolates are needed. The PCR method described in this study and an in silico typing method may help the surveillance and monitoring of the OgN groups.We identified nine novel O-genotypes (OgN) in a global collection of enterotoxigenic Escherichia coli (ETEC) [The first diarrhoeal illness that infants often experience in endemic areas is caused by enterotoxigenic i (ETEC) . In 2010i (ETEC) . The majE. coli isolates, especially pathogenic E. coli, for taxonomical and epidemiological studies. Most of what we know about E. coli prevalence, outbreaks and surveillance is described in terms of the O-serogroup. Recently, sequence analyses show that phenotypic O-serogroup diversification can be correlated with differences in the gene content and genetic diversity of the O-antigen biosynthesis gene cluster (O-AGC) located on the chromosome [wzx (encoding the O-antigen flippase), wzy (encoding the O-antigen polymerase), and the wzm and wzt genes (encoding components of the ABC transporter pathway) located on the O-AGCs are highly variable in sequence, and can be used as gene markers for the identification of O-serogroups via molecular approaches [in silicoblast analysis using a wzx/wzy and wzm/wzt sequence set extracted from O1 to O187 O-AGCs, we subtyped our 362 global ETEC isolates [n=38), O25 (n=24), O27 (n=18), O114 (n=17), O115 and O159 . In addition to the ETEC isolates classified into 48 known O-genotypes, 55 isolates carried wzx/wzy genes that showed <50\u200a%\u2009or no sequence identity to the previously defined sequences. These ETEC isolates were categorized as O-genotype untypeable (OgUT), but carried novel O-AGCs.O-serogrouping remains the gold standard for the subtyping of romosome . In partproaches . By applIn this study, we focused on characterizing the OgUT ETEC isolates detected in our previous study . Such ETwzx and wzy sequences were extracted from each O-AGC. The known wzx/wzy sequences from typical E. coli O-serogroups  [Shigella O-serogroups  [E. coli OX serogroups  [E. coli  [wzy genes of OX25 and S. dysenteriae type 6, which are not found in the O-AGCs). A complete set of fliC sequences [ETEC O-AGC sequences were obtained from draft genomes used in a previous study . The wzx groups) , Shigell sonnei) , E. coliwzx and wzy were constructed by using the neighbour-joining algorithm using mega5 software [clustal W program [Phylogenetic trees of software , followi program .wzy sequences. PCR was performed as follows: the 30\u2009\u00b5l reaction mixture contained 2\u2009\u00b5l genomic DNA, 6\u2009\u00b5l 5\u00d7 Kapa Taq buffer, dNTP mix , MgCl2 , primers  and 0.8 U Kapa Taq DNA polymerase (Kapa Biosystems). The thermocycling conditions were: 25 cycles of 94\u2009\u00b0C for 30\u2009s, 58\u2009\u00b0C for 30\u2009s and 72\u2009\u00b0C for 1 min. The PCR products were visualized following agarose gel (1.5\u200a%) electrophoresis in 0.5x TBE  and staining with ethidium bromide (1 mg/ml). Three strains  were used as positive-control strains for PCR.OgN-specific PCR primers targeting OgN3, OgN5 and OSB16 were designed based on each alignment of wca) (upstream) and the histidine biosynthesis (his) (downstream) extracted from OgUT ETEC genomes contained nine novel O-AGCs, OgN2, OgN3, OgN4, OgN5, OgN13, OgN14, OgN15, OgN16 and OgN17  (wzx and wzy were highly conserved within each O-genotype (99.8\u2013100\u200a%\u2009sequence identity). ETEC OgN5 strains were isolated in five countries, Guatemala (n=12), Argentina (n=5), Egypt (n=4), Mexico (n=4) and Indonesia (n=4), between 1989 and 2003. All ETEC OgN5 isolates carried the ST-encoding gene, except for a single isolate (E1542) that carried the LT-encoding gene, and all OgN5 isolates were confirmed negative for all known CFs by dot-blot and PCR-based analyses in our previous study [fliC-based H-typing showed that the OgN5 isolates were classified into eight H-types: H5, H9, H10, H16, H18, H19, H32 and H39 (see Table S1). Phylogenetic analysis showed OgN5 isolates were spread across phylogenetically distinct lineages where most/some isolates grouped based on their H-types . The major groups were OgN5\u200a:\u200aH10 and OgN5\u200a:\u200aH16  belonging to a phylogenetic lineage L14 [E. coli phylogroup A and a lineage L22 (designed in this study) of phylogroup B, respectively . Additionally, OgN5\u200a:\u200aH18 , OgN5\u200a:\u200aH9 , OgN5\u200a:\u200aH32 , OgN5\u200a:\u200aH19 , OgN5\u200a:\u200aH34 , OgN5\u200a:\u200aH16  and OgN5\u200a:\u200aH5  were observed. No significant association between lineages and geographical origins was observed (see Table S1).Comparative analyses revealed that chromosomal O-AGC sequences flanked by gene clusters of the colanic acid biosynthesis  . Among t (OSB16) . A summafliC of H45, and were confirmed positive for STh and two CFs, CFA/I (a fimbria) and CS21 (a type IV pilus) (see Table S1). All OgN3\u2009:\u2009H45 isolates were phylogenetically grouped into L6 of phylogroup A, and originated from Central and South America, including Mexico, Guatemala and Argentina . Six of eight OSB16 strains carried the fliC of H32, and were confirmed positive for LT, LT+STh or LT+STp, and negative for all known CFs (see Table S1). All OSB16\u200a:\u200aH32 and OSB16\u200a:\u200aH2 isolates were phylogenetically grouped into L13 of phylogroup A, and originated from Guatemala and Argentina . Additionally, three OgN13, two OgN15 and single OgN2, OgN4, OgN14, OgN16 and OgN17 strains were observed in our ETEC collection (see Table S1).Six of eight OgN3 strains carried the wzy genes  associated with diarrhoeal patients in Egypt , Boliviawzx/wzy from O-AGCs were highly conserved within each O-genotype, indicating that these O-AGCs have been spread across this species by horizontal gene transfer. Phylogenetic analysis in our previous study [Novel O-genotype ETEC isolates were found in several distinct phylogenetic lineages, and sequences of us study  showed tin silico-based serotyping web tool, SerotypeFinder, of the Center for Genomic Epidemiology (https://cge.cbs.dtu.dk/services/SerotypeFinder) [E. coli O-antigen associated genes  from O1 to O187 and H-antigen associated genes (fliC and its homologues) from H1 to H56 can be used to estimate E. coli O\u2009:\u2009H serotypes of sequenced isolates. If it is possible to organize and update the database with new wzx and wzy sequences, it becomes possible to meet the requirements for broader O-genotypes, including OgN5, OgN3 and OSB16 in the in silico typing.Recently, we have been able to access whole-genome sequencing data due to the appearance of next-generation sequence technologies. A publicly available eFinder) , which hin silico typing method.In conclusion, by using detailed whole genome sequence-based analyses, it is clear that there is a hidden diversity of ETEC O-genotypes for which there is no prior information regarding their contribution to the burden of disease. It should be noted that even from our global collection, ETEC isolates from Africa are underrepresented. Hence, it is possible and perhaps likely that additional novel ETEC O-types and their associated lineages will continue to be discovered. These novel O-genotypes identified in this study have now been included in our comprehensive O-genotyping scheme and can be detected using the PCR-based method described in this study and by the rfb) cluster of Escherichia coli O111. Gene 1995;16:17\u201323. GenBank/EMBL/DDBJ accession no.: AF078736.Bastin DA, Reeves PR. Sequence and analysis of the O antigen gene . Appl Environ Microbiol 2005;71:7995\u20138001. 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Structural and genetic characterization of Shigella boydii type 17 O antigen and confirmation of two new genes involved in the synthesis of glucolactilic acid. Biochem Biophys Res Commun 2006;349:289\u2013295. GenBank/EMBL/DDBJ accession no.: DQ875941.Senchenkova SN, Feng L, Wang Q, Perepelov AV, Qin D Escherichia coli O128 antigen biosynthesis cluster and functional identification of an alpha-1,2-fucosyltransferase. FEBS Lett 2003;553:99\u2013103. GenBank/EMBL/DDBJ accession no.: AY217096.Shao J, Li M, Jia Q, Lu Y, Wang PG. Sequence of Shigella boydii O11 and functional identification of its wzy gene. FEMS Microbiol Lett 2004;234:125\u2013132. GenBank/EMBL/DDBJ accession no.: AY529126.Tao J, Feng L, Guo H, Li Y, Wang L. The O-antigen gene cluster of et al. Molecular analysis of Shigella boydii O1 O-antigen gene cluster and its PCR typing. Can J Microbiol 2005;51:387\u2013392. GenBank/EMBL/DDBJ accession no.: AY630255.Tao J, Wang L, Liu D, Li Y, Bastin DA Escherichia coli O157 O antigen gene cluster and identification of its specific genes. Infect Immun 1998;66:3545\u20133551. GenBank/EMBL/DDBJ accession no.: AF061251.Wang L, Reeves PR. Organization of Shigella boydii O-antigen loci: implication for Escherichia coli and Shigella relationships. Infect Immun 2001;69:6923\u20136930. GenBank/EMBL/DDBJ accession no.: AF402312\u2013AF402315.Wang L, Qu W, Reeves PR. Sequence analysis of four et al. Sequence of the E. coli O104 antigen gene cluster and identification of O104 specific genes. Gene 2001;30:231\u2013236. GenBank/EMBL/DDBJ accession no.: AF361371.Wang L, Briggs CE, Rothemund D, Fratamico P, Luchansky JB Escherichia coli O55:H7 and identification of a new UDP-GlcNAc C4 epimerase gene. J Bacteriol 2002;184:2620\u20132625. GenBank/EMBL/DDBJ accession no.: AF461121.Wang L, Huskic S, Cisterne A, Rothemund D, Reeves PR. The O-antigen gene cluster of et al. Molecular markers for detection of pathogenic Escherichia coli strains belonging to serogroups O 138 and O 139. Vet Microbiol 2005;111:181\u201390. GenBank/EMBL/DDBJ accession nos: DQ109551, DQ109552.Wang L, Liu B, Kong Q, Steinr\u00fcck H, Krause G et al. Genetic and structural analyses of Escherichia coli O107 and O117 O-antigens. FEMS Immunol Med Microbiol 2009;55:47\u201354. GenBank/EMBL/DDBJ accession no.: EU694095.Wang Q, Perepelov AV, Feng L, Knirel YA, Li Y et al. Development of a DNA microarray for detection and serotyping of enterotoxigenic Escherichia coli. J Clin Microbiol 2010;48:2066\u20132074. GenBank/EMBL/DDBJ accession nos: GU014554, GU014555.Wang Q, Wang S, Beutin L, Cao B, Feng L et al. Development of a serogroup-specific multiplex PCR assay to detect a set of Escherichia coli serogroups based on the identification of their O-antigen gene clusters. Mol Cell Probes 2010;24:286\u2013290. GenBank/EMBL/DDBJ accession nos: GU068041, GU068044\u2013GU068046.Wang Q, Ruan X, Wei D, Hu Z, Wu L et al. Molecular cloning and characterization of genes for Shigella sonnei form I O polysaccharide: proposed biosynthetic pathway and stable expression in a live salmonella vaccine vector. Infect Immun 2002;70:4414\u20134423. GenBank/EMBL/DDBJ accession no.: AF294823.Xu DQ, Cisar JO, Ambulos Jr N, Burr DH Shigella dysenteriae serotype 1 O-antigen in live Salmonella Typhi vaccine vector Ty21a: preclinical evidence of immunogenicity and protection. Vaccine 2007;25:6167\u20136175. GenBank/EMBL/DDBJ accession no.: AY585348.Xu DQ, Cisar JO, Osorio M, Wai TT, Kopecko DJ. Core-linked LPS expression of"}
+{"text": "Corynebacterium CDC group F-1 were assembled and studied. Genome sizes ranged between 2.3 and 2.44\u00a0Mb, with G+C content between 60.4% and 60.7%.Three draft and one complete genome sequence from strains isolated from urine and consistent with  Corynebacterium based on biochemical results and other phenotypic traits  published an identification scheme for isolates consistent with the genus roup F-1 . Here, fDNA was extracted using the TruSeq DNA high-throughput (HT) sample preparation kit. Paired-end whole-genome shotgun libraries were constructed using TruSeq Nano DNA HT library preparation kit. A mate-pair library for strain NML 98-0116 was prepared using the Nextera mate-sample prep kit (Illumina). Sequencing was performed using a MiSeq sequencer (Illumina), and assembly was performed with SPAdes version 3.1.1) . Contigs.1.1 . Cohttps://www.ncbi.nlm.nih.gov/genome/annotation_prok). Assembled whole genomes demonstrated essentially identical G+C contents, genome sizes, and number of coding regions , with a G+C content of 56.72% (Corynebacterium species (ermX gene (associated with resistance to erythromycin and clindamycin) homologous to that found in Corynebacterium urealyticum, Corynebacterium\u00a0jeikeium, and Corynebacterium\u00a0striatum. This was consistent with published antibiograms for these strains  Prokaryotic Genome Annotation Pipeline (PGAP) ( regions . The gen regions . Using t regions . NML 140 strains , consist strains .Corynebacterium CDC group F-1 isolates have been deposited in GenBank under accession numbers shown in Complete or draft genome sequences of four"}
+{"text": "The publisher apologizes for the error. The correct citation is: Zou W, Yadav S, DeVault L, Jan YN, Sherwood DR (2015) RAB-10-Dependent Membrane Transport Is Required for Dendrite Arborization. PLoS Genet 11(9): e1005484. doi:10.1371/journal.pgen.1005484This article was republished on October 21, 2015, to correct an XML error causing the 4S1 File(PDF)Click here for additional data file.S2 File(PDF)Click here for additional data file."}
+{"text": "Plasmodium parasites to bind and invade target cells. Hence, characterising molecules that participate in reticulocyte interaction is key to understanding the molecular basis of Plasmodium vivax invasion. This study focused on predicting functionally restricted regions of the P. vivax GPI-anchored micronemal antigen (PvGAMA) and characterising their reticulocyte binding activity.Adhesin proteins are used by pvgama gene was initially found in P. vivax VCG-I strain schizonts. According to the genetic diversity analysis, PvGAMA displayed a size polymorphism very common for antigenic P. vivax proteins. Two regions along the antigen sequence were highly conserved among species, having a negative natural selection signal. Interestingly, these regions revealed a functional role regarding preferential target cell adhesion.The PvGAMA reticulocyte binding properties for the first time. Conserved functional regions were predicted according to natural selection analysis and their binding ability was confirmed. These findings support the notion that PvGAMA may have an important role in P. vivax merozoite adhesion to its target cells.To our knowledge, this study describes The online version of this article (doi:10.1186/s13071-017-2183-8) contains supplementary material, which is available to authorized users. Plasmodium vivax is a human malaria-causing parasite whose eradication is a priority on the international health agenda C[G/C]C[A/T]AA[C/T][C/G][A/G/C][G/A]AC[G/C/A] repeat which was not present in P. cynomolgi, P. inui, P. fragile, P. knowlesi or P. coatneyi , P\u2009>\u20090.1). However, synonymous divergence was greater than non-synonymous divergence (P\u2009<\u20090.0001) when comparing pvgama sequences to each related species: KN-KSP. vivax/P. cynomolgi\u2009=\u2009-0.041 (0.006); KN-KSP. vivax/P. inui\u2009=\u2009-0.062 (0.008); KN-KSP. vivax/P. fragile\u2009=\u2009-0.030 (0.006); KN-KSP. vivax/P. knowlesi\u2009=\u2009-0.072 (0.009); KN-KSP. vivax/P. coatneyi\u2009=\u2009-0.049 (0.007). The evolutionary rate \u03c9 (dN/dS and KN/KS) sliding window showed that two highly conserved regions amongst species (codons 80\u2013320 and 514\u2013624) might be under negative selection (\u03c9\u2009<\u20090.5). Furthermore, 308 negatively-selected codons were observed amongst species  whilst 67.5% of them recognised rPvGAMA-Ct (0.47 cut-off point). These data agreed with a study of the profile of the humoral immune response for P. vivax in which rPvGAMA was recognised by 54.5% of the sera used in the array AN[A/G][N/D] was predicted. This RR was common in different ce Pv12 ) or in tce . DN. DNP. vi  or in t regions , 53. Thimsp4  showed higher rosetting activity, unlike the F1 region (aa 22 to 344) (amino fragment) : percentage of sites evolving under positive selection. P-value corrected for multiple tests using the Holm-Bonferroni method. (TIF 470\u00a0kb)Lineage-specific positive selection. Branches under positive episodic selection were identified by using the REL-site branch method. Episodic selection acts very quickly and involves a switch from negative to positive natural selection and back to negative and might enable adaptation to a new host. Phylogeny was inferred in MEGA v6 by the maximum likelihood method using the GTR\u2009+\u2009G evolutionary model. \u03c9Additional file 3: Figure S3.PvGAMA-Nt and -Ct stained with Coomassie blue or analysed by western blot using anti-polyhistidine antibodies, respectively. c Purifying conserved (CR1 and CR2) and variable (VR1 and VR2) PvGAMA regions. Lanes 2, 4, 6 and 8 show purified recombinant proteins and lanes 3, 5, 7 and 9 show western blot detection. The proteins\u2019 molecular markers are indicated in Lane 1 on all figures. (TIF 5327\u00a0kb)Obtaining recombinant proteins. a, b Recombinant GAMA protein expression and purification. Lanes 2\u20133 show non-induced and induced cell lysate, respectively. Lanes 4\u20135 show purified rAdditional file 4: Figure S4.Selection strategy for immature and mature erythrocyte populations. The doublets were excluded by plotting FSC-H against FSC-A. Cells were then selected by their granularity, using an SSC-A vs FSC-A cytogram. The CD45 vs CD71 signal was plotted for selecting reticulocyte (CD71\u2009+\u2009CD45-) and mature erythrocyte (CD71-CD45-) populations and omitting activated lymphocytes (CD71\u2009+\u2009CD45+). The percentage of cells having bound protein was calculated using the PE signal (CD71\u2009+\u2009CD45-PE+). A representative histogram from three independent experiments analysing the PE signal for the CR2 binding assay compared to control is also shown. (TIF 10448\u00a0kb)"}
+{"text": "During 2000\u20132013, among adults aged 50\u201375 years, the use of colorectal cancer tests or procedures increased for all racial and ethnic groups shown. Non-Hispanic Asian adults had the largest increase; the percentage more than doubled from 20.6% in 2000 to 51.2% in 2013. Although increases were observed among all groups, in 2013 the prevalence of colorectal cancer screening remained higher among non-Hispanic white (60.4%) and non-Hispanic black (58.2%) adults and lower among non-Hispanic Asian (51.2%) and Hispanic (41.5%) adults.Source: Health, United States, 2014 . Available at http://www.cdc.gov/nchs/data/hus/hus14.pdf.Reported by: Hashini Khajuria, MPA, hwq6@cdc.gov, 301-458-4253."}
+{"text": "Acute myocarditis (AM) is an inflammatory disease of the heart muscle. Clinical presentation and outcome are variable, ranging from asymptomatic to fulminant myocarditis (FM), presenting with decompensated heart failure or cardiogenic shock needing inotropic or mechanical circulatory support. Cardiac magnetic resonance (CMR) is increasingly employed in the diagnosis and follow-up (f.u.) of AM. We investigated the differences in functional parameters evaluated with CMR between patients (pts) presenting with FM versus non fulminant AM (NFM), at baseline and f.u.We retrospectively analyzed consecutive pts with AM who underwent CMR  within 30 days from clinical presentation. Cine, STIR T2-weighted and late-enhancement images were acquired on matching planes. CMR diagnosis of AM was based on Lake Louise Criteria. Indexed LV end-diastolic (LVEDVi) and end-systolic (LVESVi) volumes, LV ejection fraction (LVEF), indexed LV mass (LVmi), indexed RV end-diastolic (RVEDVi) and end-systolic (RVESVi) volumes and RV ejection fraction (RVEF) were calculated for each patient. F.u. scans were similarly analyzed, if available.2, p=0.028) and LVEF improved from 56% to 66% (p=0.028); in NFM LVEDVi and LVEF were unchanged at f.u. . F.u. LVEF and RVEF did not differ significantly between FM and NFM, 66% (60-69) vs 65% (61-69), p=0.79 and 62% (57-68) vs 63% (59-68),p=0.69 respectively. In both FM and NFM LVmi significantly decreased from baseline . There was a trend towards a more pronounced reduction of LVmi in FM, but the difference fell short of statistical significance .Four pts with FM died and 3 were heart transplanted before performing CMR and thus were excluded from the analysis; none of the pts with NFM died or were transplanted before CMR. In total, 68 pts  were included: 11 (16%) with FM and 57 (84%) with NFM underwent CMR. Baseline scans were obtained earlier in the NFM group, 5 (3-9) vs 12 (7-21) days after presentation, p = 0.0035. Baseline LVEF was significantly lower in FM compared to NFM, 56% (43-65) vs 64% (59-68),p=0.026. RVEF was lower in FM than in NFM pts, 56% (52-58) vs 63% (58-66),p=0.0024. F.u. scans were available in 47 pts  and were obtained after a median of 169 (110-203) days from baseline scan in FM and 148 (93-242) days in NFM (p=0.78). In FM, LVEDVi was increased at f.u. (from 75 to 77 mL/mPts with FM achieved normal LVEF at f.u., despite a lower baseline LVEF, even if a mild LV remodeling was observed at f.u. This might in part be due to a more pronounced decrease in LVmi, likely secondary to more severe myocardial edema in FM, but this observation warrants further investigation and longer f.u. to better understand long term consequences of FM on ventricular remodeling and prognosis."}
+{"text": "Talaromyces produce useful biomass-degrading enzymes and secondary metabolites. However, these enzymes and secondary metabolites are still poorly understood and have not been explored in depth because of a lack of comprehensive genetic information. Here, we report a 36.51-megabase genome assembly of Talaromyces pinophilus strain 1\u201395, with coverage of nine scaffolds of eight chromosomes with telomeric repeats at their ends and circular mitochondrial DNA. In total, 13,472 protein-coding genes were predicted. Of these, 803 were annotated to encode enzymes that act on carbohydrates, including 39 cellulose-degrading and 24 starch-degrading enzymes. In addition, 68 secondary metabolism gene clusters were identified, mainly including T1 polyketide synthase genes and nonribosomal peptide synthase genes. Comparative genomic analyses revealed that T. pinophilus 1\u201395 harbors more biomass-degrading enzymes and secondary metabolites than other related filamentous fungi. The prediction of the T. pinophilus 1\u201395 secretome indicated that approximately 50% of the biomass-degrading enzymes are secreted into the extracellular environment. These results expanded our genetic knowledge of the biomass-degrading enzyme system of T. pinophilus and its biosynthesis of secondary metabolites, facilitating the cultivation of T. pinophilus for high production of useful products.Species from the genus  Talaromyces pinophilus, formerly designated Penicillium pinophilum, is a fungus that produces biomass-degrading enzymes such as \u03b1-amylase1, cellulase2, endoglucanase3, xylanase2, laccase4 and \u03b1-galactosidase2. In one study, a blended enzyme cocktail produced by T. pinophilus and Chrysoporthe cubensis improved the efficiency of hydrolysis of glucan and xylan in sugarcane bagasse for glucose and xylose production, compared with enzymes from a single strain2. A relatively high level of \u03b2-glucosidase activity is observed in T. pinophilus under solid state fermentation5. Therefore, T. pinophilus is considered a potential alternative to Trichoderma reesei for cellulase production and efficient biomass hydrolysis. T. pinophilus produces a variety of medically useful metabolites such as 3-O-methylfunicone, which is used to inhibit mesothelioma cell motility6, and talaromycolides 1\u20133, 5 and 11, which inhibit the growth of the human pathogen methicillin-resistant Staphylococcus aureus7.T. pinophilus 1\u201395 was isolated from the soil of a dried, ploughed field in Wuzhou, China. This strain produces a highly efficient, calcium-independent \u03b1-amylase1. Application of calcium-independent \u03b1-amylase in starch conversion avoids problems caused by addition of calcium ions8. Additionally, we found that T. pinophilus 1\u201395 produces 1.21\u2009\u00b1\u20090.30\u2009U/mL of filter-paper cellulase, 10.72\u2009\u00b1\u20090.74\u2009U/mL of carboxymethylcellulose cellulase, 0.71\u2009\u00b1\u20090.02\u2009U/mL of p-nitrophenyl-\u03b2-cellobioside cellulase, 0.27\u2009\u00b1\u20090.01\u2009U/mL of p-nitrophenyl-\u03b2-glucopyranoside cellulase and 41.93\u2009\u00b1\u20092.84\u2009U/mL of xylanase activities in submerged flask cultivation (data not shown). However, a comprehensive understanding of the biomass-degrading enzyme system in this fungus is still lacking.The fungal strain de novo whole-genome assembly of T. pinophilus strain 1\u201395, a nearly complete genome sequence of a high biomass-degrading enzyme-producing species in the genus Talaromyces. Carbohydrate-active enzyme (CAZyme) genes and secondary metabolism gene clusters were observed in the sequenced genome. Comparative genomic analysis suggested that T. pinophilus harbors more biomass-degrading enzymes and secondary metabolites than other related filamentous fungi. In addition, the predicted secretory protein patterns of T. pinophilus 1\u201395 were analyzed.We describe the T. pinophilus 1\u201395 (CGMCC No. 2645), isolated from soil in a dried, ploughed field in Wuzhou, China, was performed using a combination of single molecule real-time (SMRT) DNA sequencing and next generation sequencing technology. A high-quality genome sequence of T. pinophilus 1\u201395 was generated on the PacBio RS II platform. Approximately 1.94\u2009Gb of clean subreads, with sequences from a single pass of a polymerase on a single strand of an insert within a SMRTbell template and no adapter sequences with an N50 size of 10,045\u2009bp and average length 8,102\u2009bp were generated. Additionally, a paired-end (PE) library with a 500-bp average insert size was constructed using the Illumina HiSeq 4000 platform, and 3.88\u2009Gb clean, short-sequence PE reads were generated with a length of 125\u2009bp. Reads were used to correct wrong bases in the assembled genome sequence on the PacBio RS II platform. Finally, a 36.51-Mb genome of T. pinophilus 1\u201395 was generated with 159-fold coverage. This size was in accordance with the estimated genome size of 28\u201336\u2009Mb for three Talaromyces species11. The genome was covered by nine scaffolds, including eight large scaffolds (accession number CP017344-CP017351 in GenBank) without gaps , GeneMark-ES (http://exon.gatech.edu/GeneMark/), Genewise (http://www.ebi.ac.uk/Tools/psa/genewise/), SNAP14 and an unsupervised learning system program Glean version 1 (https://sourceforge.net/projects/glean-gene/). The number of coding genes was significantly higher than other filamentous fungi that produce biomass-degrading enzymes , were annotated in the Gene Ontology (GO) database (http://geneontology.org/), 12,828 (95.21%) in the UniProt database (http://www.uniprot.org/), 12,946 (96.09%) in the NCBI non-redundant (NR) protein sequences database (ftp://ftp.ncbi.nlm.nih.gov/blast/db/) and 4437 (32.93%) in the Clusters of Orthologous Groups of proteins database (http://www.ncbi.nlm.nih.gov/COG/). A total of 6817 (50.6%) genes belonging to 331 pathways were also annotated in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database (http://www.kegg.jp/)  search of all-vs.-all findings showed that T. pinophilus 1\u201395 and T. cellulolyticus Y-94 shared 10,260 orthologous proteins, accounting for 76.16% of the total proteome, with an average amino acid sequence identity of 98.28%. In contrast, low identity was observed to other sequenced Talaromyces species17, Trichoderma sp.18, Penicillium sp.20 and Aspergillus sp.22, ranging from 70% to 88% , 35 families of glycosyl transferases (GTs), 13 families of carbohydrate esterases (CEs), 10 families of auxiliary activities (AAs), 5 families of polysaccharide lyases (PLs) genes and 19 families of carbohydrate-binding modules (CBMs)  Fig.\u00a0. FurtherMs) Fig.\u00a0.Figure 3T. cellulolyticus Y-94 and P. oxalicum HP7-1, larger numbers of BGLs were classified into GH families 1 and 3 in the T. pinophilus 1\u201395 genome , 8 \u03b2-1,4-endoglucanases  and 29 \u03b2-glucosidases  were included as cellulases. Of these, the known cellulases included Cel7A-2 (TP09412), Cel5A (TP03457), Cel5B (TP07499), Cel7B (TP08514) and Bgl3A (TP09042). Compared with known filamentous fungi used for cellulase production, i.e., T. pinophilus 1\u201395 genome was rich in hemicellulose-degrading enzymes (97 genes) assigned into 29 predicted CAZyme families; this finding was compared to 77 genes in T. cellulolyticus Y-94 and 80 genes in P. oxalicum HP7\u20131, which includes Xyn11A (TP00436) and Xyn10A (TP06900), encoding the important \u03b2-1,4-endoxylanases. This result supports the high xylanase activity of T. pinophilus 1\u201395. The predicted hemicellulases in T. pinophilus 1\u201395 were divided into 19 types by substrate specificities. The large differences among T. pinophilus 1\u201395, T. cellulolyticus Y-94 and P. oxalicum HP7\u20131 broadly covered most of the listed hemicellulase types (see Supplementary Table\u00a0T. pinophilus 1\u201395 possessed more \u03b2-D-xylosidases (EC 3.2.1.37), acetyl xylan esterases (EC 3.1.1.72) and feruloyl esterases (EC 3.1.1.73) than the two others. \u03b2-D-xylosidases hydrolyze xylobiose or linear xylooligosaccharides to the monomer xylose. Acetyl xylan esterases liberate acetic acid esterifying position 2 and 3 on mono- and di-O-acetylated \u03b2-1,4-linked D-xylopyranosyl residues in xylan chains. Feruloyl esterases liberate trans-ferulic acid from 5-O-feruloylated L-arabinofuranosyl residues. These enzymes facilitate the hydrolysis of hemicellulose26. In contrast, the T. pinophilus 1\u201395 genome possessed a similar number of pectin-degrading enzymes as T. cellulolyticus Y-94 and P. oxalicum HP7\u20131 , which mainly hydrolyze \u03b1-1,4-glucosidic linkages from the nonreducing ends of starch chains with the release of \u03b2-D-glucose28; and a 1,4-\u03b1-glucan branching enzyme (EC 2.4.1.18) that cleave \u03b1-1,4 glucosidic linkages of glucan chain, and then transfer the cut end to 6-position of glucose residue within the cleaved or another glucan chain, resulting the generation of an \u03b1-1,6 glucosidic linkage29. Comparative analysis indicated that T. pinophilus 1\u201395 had more \u03b1-glucosidases and glucoamylases than the other investigated fungi  containing zinc-finger structures, such as the Zn2Cys6 type, the C2H2 type and CCHC type, followed by the winged helix repressor DNA-binding family (97 members) , the cellulase transcription activator CLR-2 (TP10486), the starch degradation regulator AmyR (TP09286) and the xylan degradation regulator XlnR (TP02627) (Table\u00a0T. pinophilus 1\u201395.Transcription factors (TFs) are essential for modulating diverse biological processes by regulating gene expression. In total, 943 TFs were found in the predicted proteome of T. pinophilus 1\u201395 had a wealth of secondary metabolites using the AntiSMASH web service30. A total of 68 secondary metabolism gene clusters harboring 401 putative genes were identified. The predicted products of 52 secondary metabolism gene clusters were classified into 8 different types: 28 T1 polyketide synthase (T1PKS) gene clusters, 9 non-ribosomal peptide synthase (NRPS) gene clusters, 9 terpene gene clusters, 2 Nrps-T1pks gene clusters, 1 phosphonate gene cluster, 1 T1pks-Indole gene cluster, 1 T1pks-Nrps gene cluster and 1 Terpene-T1pks gene cluster; the remaining 16 gene clusters synthesized other unknown secondary metabolites , comprising 831 classical and 372 nonclassical secretory proteins , Cel7B (TP08415), Cel5A (TP13457), Cel5B (TP07499), Cel5C (TP08784), Cel45A (TP06957) and Bgl3A (TP09042). We also identified 62 hemicellulose-degrading enzymes and 6 pectin-degrading enzymes, including eight \u03b2-1,4-endoxylanases, nine acetyl xylan esterases, four \u03b1-galactosidases, 10 \u03b1-L-arabinofuranosidases, five \u03b1-L-fucosidases, one endo-1,4-\u03b2-mannanase and four feruloyl esterases, as well as two pectin esterases, two tannases, one pectate lyase and one pectin lyase , glucoamylases (5 coding genes) and \u03b1-glucosidases (13 coding genes) among species we compared. BGLs are important for releasing inhibition of cellulase activity36. Furthermore, the predicted secretome of T. pinophilus 1\u201395 showed that approximately 50% of CWDEs and starch-degrading enzymes were secreted into the extracellular region, including major cellulases, hemicellulases and amylases. This result indicated a promising application of T. pinophilus in biorefining. These results also demonstrated that T. pinophilus 1\u201395 is more excellent cellular machinery for biomass-degrading enzymes than that of P. oxalicum HP7\u20131 and T. cellulolyticus Y-94 as previously observed35, meriting further study.Comparative genome analysis indicated that the most closely related species to Talaromyces, Trichoderma, Penicillium and Aspergillus, indicated that T. pinophilus 1\u201395 possessed the most secondary metabolism gene clusters except for A. niger and A. oryzae. T. pinophilus 1\u201395 had more T1PKs than Trichoderma sp., Penicillium sp. and Aspergillus sp., and fewer NRPS than Aspergillus species. These results suggested that T. pinophilus 1\u201395 has potential for producing bioactive secondary metabolites. Although thus far, several bioactive secondary metabolites have been extracted and characterized from T. pinophilus7, according to the genomic data, additional secondary metabolites could be generated using this species.Comparative analysis to 10 filamentous fungi from four genera, Talaromyces. The result provided new insights for a comprehensive understanding of the biomass-degrading enzyme system of Talaromyces at the genome level. Detailed comparative genomic analysis revealed a complex biomass-degrading enzyme system in T. pinophilus, indicating its promising application in biomass biorefineries. This study provides a genome-sequence basis for developing strategies that use T. pinophilus as a microbial cell factory for production of high-value enzymes and secondary metabolites.In summary, this study provided a nearly complete genome sequence for the genus T. pinophilus 1\u201395 was isolated from soil in a dried, ploughed paddy field in Wuzhou, China1 and was deposited at the China General Microbiological Culture Collection Center (CGMCC) under accession number CGMCC No. 2645. Total DNA extraction from mycelia was performed using a phenol-chloroform method with some modifications37. Mycelia were ground in liquid nitrogen and put in 1\u2009mL lysate reagent  per 100\u2009mg mycelia powder. Genomic DNA was collected by centrifugation at 11,300\u2009\u00d7\u2009g for 10\u2009min.T. pinophilus strain 1\u201395 genome was sequenced using a PacBio RS II platform and Illumina HiSeq 4000 platform at the Beijing Genomics Institute . Four SMRT cells zero-mode waveguide arrays of sequencing, were used by the PacBio platform to generate the subreads set. PacBio subreads (length\u2009<\u20091\u2009kb) were removed. The program Pbdagcon (https://github.com/PacificBiosciences/pbdagcon) was used for self-correction. Draft genomic unitigs, which are uncontested groups of fragments, were assembled using the Celera Assembler38 against a high-quality corrected circular consensus sequence subreads set. Order, distance and orientation of unitigs and combined scaffolds were generated using software SSPACE39. An upgraded draft genome was obtained after filling or reducing as many captured gaps as possible using software PBJelly40. To improve the accuracy of the genome sequences, GATK (https://www.broadinstitute.org/gatk/) and SOAP tool packages 42 were used to make single-base corrections.The 43 used reasonable PE sequence data from Illumina libraries to perform comprehensive variant detection and improve genome assembly.A DNA library of 500\u2009bp inserts was constructed and PE sequenced. For generated HiSeq reads, Q20, representing the probability of the incorrectness of the corresponding base call, was detected. If Q20 reads accounted for less than 60%, they were discarded. Software PilonT. pinophilus 1\u201395 genome were predicted independently with the gene prediction programs Augustus , GeneMark-ES (http://exon.gatech.edu/GeneMark/), Genewise (http://www.ebi.ac.uk/Tools/psa/genewise/), SNAP14, and the unsupervised learning system program Glean (https://sourceforge.net/projects/glean-gene/) version 1. Augustus and SNAP, using default parameters, were trained on gene models for A. oryzae, P. oxalicum, T. marneffei and T. stipitatus  and Swiss-Prot and TrEMBL databases (http://www.mrc-lmb.cam.ac.uk/genomes/madanm/pres/swiss2.htm)  and KEGG database (http://www.kegg.jp/) version 76, were used to assign general protein function profiles. We used cut-off E-value\u2009\u2264\u20091e-5, overlap 0.4 and identity 30. InterProScan5 (http://www.ebi.ac.uk/interpro/), stand-alone version 55, and GO (http://geneontology.org/) were also used to annotate the predicted proteome. TFs were predicted based on InterPro IDs in the Fungal Transcription Factor Database (http://ftfd.snu.ac.kr/). The hmmsearch program in the HMMER 3.1b2 package (http://hmmer.org/), was used to search all predicted proteomes with the family-specific hidden Markov model profiles of CAZymes from the dbCAN database26. Primary results were processed with an E-value threshold of 1E-7. Protein kinases and phosphatases were detected using hmmsearch based on the Eukaryotic Kinase and Phosphatase Database (http://ekpd.biocuckoo.org/). Membrane transport proteins were classified and identified by a BLASTp search against the transport classification database44, with E-value threshold 1E-10, overlap 0.4 and identity of 30. AntiSMASH30 was used to annotate secondary metabolism gene clusters.For functional annotation of translated proteins in the 45. We selected 2,082 single-copy genes from 118,099 genes in 11 fungal genomes. MUSCLE (http://www.ebi.ac.uk/Tools/msa/muscle/) version 3.7 with default parameters was used to perform multiple sequence alignment of single-copy genes. A neighbor-joining tree was calculated using TreeBeST46 with bootstrapping set to 100. The phylogenetic tree was visualized using SVGKit (http://svgkit.sourceforge.net/) and PERL scripts.An all-against-all pairwise BLASTp similarity search was performed using proteomes from 11 filamentous fungi  and for glycosylphosphatidyl inositol anchors use web server PredGPI (http://gpcr.biocomp.unibo.it/predgpi/). Software tRNAscan-SE (http://lowelab.ucsc.edu/tRNAscan-SE/) version 1.3 was used for transfer RNA prediction using the T. pinophilus 1\u201395 genome with option C and other default parameters.The total set of 13,472 proteins of Supplementary Infromation for SREP-16-39329Dataset 1Dataset 2Dataset 3"}
+{"text": "Bradyrhizobium canariense, isolated from root nodules of Lupinus microanthus and Lupinus angustifolius, and 1 strain of Bradyrhizobium japonicum isolated from root nodules from Lupinus angustifolius in Algeria. These sequences add to the known diversity of this agronomically important genus.We report here the whole-genome sequences of 14 strains of Bradyrhizobium canariense , mungbean (Vigna radiata), cowpea (Vigna unguiculata), and siratro (Macroptilium atropurpureum)  is rather recent (Lupinus angustifolius and Lupinus micranthus (Papilionoideae: Genisteae), collected at 2 sites in the National Park El-Kala . For whole-genome sequencing, DNA libraries were generated with a Nextera XT kit . Sequencing was performed on a MiSeq sequencer (Illumina) in three different runs generating 2 \u00d7 250-bp paired-end reads (version 2 chemistry) and 2 \u00d7 250-bp and 2 \u00d7 300-bp paired-end reads (version 3 chemistry). Quality control of the reads was assessed with FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc). Genome assemblies were computed with SPAdes genome assembler 3.10  and 8,379,024\u00a0bp (UBMAN05), with G+C contents from 62.94%  to 63.06% (UBMA122), and Bradyrhizobium japonicum UBMA197 had a larger genome, at 10,442,239\u00a0bp, with 63.3% G+C content. Genome coverage varied from 46-fold (UBMA510) to 255-fold (UBMA060). PlasmidFinder  , 4. Bactrpureum) , 2, 5. Brpureum) , Bradyrhs Poland , 7, Itals Poland , and Mors Poland . While dr recent . The 15 ler 3.10  and analler 3.10 . For BraidFinder  and PlasidFinder  detectedidFinder  identifiAll genome sequences have been deposited at GenBank under the accession numbers reported in"}
+{"text": "The effect of degradable starch microspheres (DSM) on the intrahepatic distribution of a low molecular weight marker, 99Tcm-labelled methylene diphosphonate (MDP), was studied in rats with hypovascular HSN liver tumours. MDP was injected regionally, via the hepatic artery, alone or co-administered with DSM, with or without subsequent occlusion of either the hepatic artery or the portal vein. Tumour vascularity was measured with 57Co-labelled microspheres. Co-injection with DSM immediately significantly increased hepatic retention of marker in both tumour (T) (median 22.40 (range 16.82-39.58)% injected dose) and normal liver (N) (9.08 (4.85-12.59) %ID) the greater effect seen in T (P < 0.01). After DSM degradation, very little MDP remained in N (0.61 (0.28-1.40) %ID) but there was significant retention in T (10.01 (6.73-20.28) %ID, P < 0.01). Clamping the hepatic artery had minimal effect on the retention of MDP when administered alone. Regional injection of 16.5 microM 57Co microspheres resulted in a N:T ratio of 2.25:1. Concomitant injection of the 40 microM DSM was 57Co microspheres reversed this ratio to 1:2. The results indicate that DSM selectively enhances the retention of MDP to a hypovascular hepatic tumour, not by causing intra-tumour stasis, but by directing a greater arterial flow to hypovascular areas in the liver."}
+{"text": "Primary chemotherapy in operable breast invasive carcinoma enables tumour reduction and conservative surgery. In order to search for one or more biological factors capable of predicting tumour behaviour under primary chemotherapy, and subsequent patient survival, an immunohistochemical study was performed with specific antibodies to p53, c-erbB-2 (Her-2/neu), Mib1 (antiKi-67), pS2, GST pi, oestrogen receptors (ERs) and progesterone receptors (PRs). Core biopsies, obtained before primary chemotherapy, were available from a series of 128 breast invasive carcinomas treated between January 1985 and April 1989, with a median follow-up of 93.3 months. Univariate statistical analysis showed that negative ER detection by immunohistochemistry (IHC) was highly correlated with chemosensitivity (P = 0.001). A high percentage of Mib1-positive tumour cells (> 40%), as well as initial tumour size less than 4 cm, were also correlated with tumour responsiveness to chemotherapy (P = 0.009 and P = 0.03). By multivariate analysis IHC-ER, Mib1 and initial tumour size were independent predictors, the last parameter being the most important. Concerning subsequent patient survival, c-erbB-2 overexpression, as detected by IHC, was significant with respect to overall survival (OS) (P = 0.0006), disease-free interval (DFI) (P = 0.03) and metastasis-free interval (MFI) (P = 0.008) by univariate analysis. Furthermore, c-erbB-2 was the major independent prognostic factor for OS and MFI by multivariate analysis."}
+{"text": "A sentence was omitted from Section 2.2 of the Methods . The corrected paragraph should read: DMEM, bovine serum and penicillin/streptomycin were purchased from Hyclone . Nilotinib (TasignaR)  was obtained as a gift from Novartis pharmaceuticals . Imatinib  and dasatinib  were purchased from ChemieTeck Inc. . Paclitaxel, vincristine, doxorubicin, colchicine, p-aminophenylmethylsulfonyl fluoride, bovine serum albumin, dimethyl sulfoxide (DMSO) and 1--3,5-diphenylformazan (MTT), the monoclonal mouse antibody against P-gp (P7965), the secondary horseradish peroxidase-labeled anti-goat or anti-mouse IgG were purchased from Sigma-Aldrich Chemical Co. . A polyclonal goat antibody against human MRP7 (D-19) was purchased from Santa Cruz Biotech Inc.<br/>. A polyclonal antibody against human MRP1/ABCC1 was kindly provided by Dr. Akiyama  [21]. A monoclonal antibody BXP-34 against BCRP was acquired from Signet Laboratories Inc. . [3H]-paclitaxel  was purchased from Moravek Biochemicals ."}
+{"text": "Two tris-chelated water-coordinated units are bridged by a 2,4-dihydroxy\u00adpyrimidine-5-carboxyl\u00adate dianion, which is disordered about a center of inversion. The metal center has a monocapped square-anti\u00adprismatic coordination.The water-coordinated neodymium(III) atom in the centrosymmetric title compound, [Nd DOI: 10.1107/S1600536808001499/ci2552Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:"}
+{"text": "B. microti infection (7).\u201dIn \u201cTransfusion-Associated Babesiosis after Heart Transplant\u201d by Joseph Z. Lux et al., errors occurred in the text. On page 118, left column, lines 20\u201322, the sentence should read \u201che became symptomatic during the typical 2- to 8-week incubation period for transfusion-transmitted http://www.cdc.gov/ncidod/eid/vol8no12/02-0149.htm.The corrected text appears online at"}
+{"text": "The 4,4\u2032-bipyridine ligands bridge the metal ions, forming one-dimensional chains along different directions, which are further connected by formate ligands into a topologically (1010.124.14)(10)3 three-dimensional network.In the title compound, [Cd(HCOO) DOI: 10.1107/S1600536809009969/jh2074Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:"}
+{"text": "From January 1983 to December 1986 seventy-six previously untreated children with non-Hodgkin's lymphoma (NHL) were treated by combination chemotherapy. Burkitt's lymphoma patients were ineligible. The treatment regimens include intermittent chemotherapy and for non-localized patients, prophylactic central nervous system chemotherapy. Intrathoracic non-Hodgkin's lymphoma patients also had cranial prophylactic radiotherapy. Sixty-six patients (86.8%) achieved complete remission. Two year failure-free survival rate was 82.1% for localized (stage I and II) NHL and 53.3% for non-localized (stage III and IV) NHL patients. Failure-free survival did not differ significantly for the two major histologic diagnoses, but two year survival rate was lower in diffuse poorly differentiated lymphoblastic than undifferentiated non-Burkitt's lymphoma (50% versus 66.8% respectively). Failure-free survival rate was 53.7% in mediastinal disease and, 73.2% in abdominal disease at 24 months. Relapse rate was higher in mediastinal cases (46.1%) than primary abdominal cases (24.3%) at 24 months. Eleven (13.5%) died of treatment related sepsis. Although the overall survival rate was 72.4% at 2 years we need novel or more intensive programmes for mediastinal and non-localized disease."}
+{"text": "Correction for:10.1371/journal.pmed.0030184Wagstaff SC, Laing GD, Theakston RDG, Papaspyridis C, Harrison RA (2006) Bioinformatics and multiepitope DNA immunization to design rational snake antivenom. PLoS Med 3(6): e184. doi:In the Supporting Information section of this paper, Figure S1 and Figure S2 were incorrectly linked. Figure S1 and Figure S2 labeled and in the order originally intended are provided here.Figure S1(48 KB PDF).Click here for additional data file.Figure S210.1371/journal.pmed.0050209.sg002 (32 KB PDF).Found at doi:Click here for additional data file."}
+{"text": "Drosophila wing growth by fueling a wave front of Fat-Dachsous signaling that recruits new cells into the wing primordium.The secreted morphogen Wingless promotes  Drosophila wing primordium undergoes a dramatic increase in cell number and mass under the control of the long-range morphogens Wingless  and Decapentaplegic . This process depends in part on the capacity of wing cells to recruit neighboring, non-wing cells into the wing primordium. Wing cells are defined by activity of the selector gene vestigial (vg) and recruitment entails the production of a vg-dependent \u201cfeed-forward signal\u201d that acts together with morphogen to induce vg expression in neighboring non-wing cells. Here, we identify the protocadherins Fat (Ft) and Dachsous (Ds), the Warts-Hippo tumor suppressor pathway, and the transcriptional co-activator Yorkie  as components of the feed-forward signaling mechanism, and we show how this mechanism promotes wing growth in response to Wg. We find that vg generates the feed-forward signal by creating a steep differential in Ft-Ds signaling between wing and non-wing cells. This differential down-regulates Warts-Hippo pathway activity in non-wing cells, leading to a burst of Yki activity and the induction of vg in response to Wg. We posit that Wg propels wing growth at least in part by fueling a wave front of Ft-Ds signaling that propagates vg expression from one cell to the next.During development, the  Drosophila, wings develop at the larval stage from wing primordia. Recently, we discovered that the morphogen Wingless promotes growth of the Drosophila wing by inducing the recruitment of neighboring cells into the wing primordium. Wing cells are defined by the expression of the \u201cselector\u201d gene vestigial. Recruitment depends on the capacity of wing cells to send a short-range, feed-forward signal that allows Wingless to activate vestigial in adjacent non-wing cells. Here, we identify the molecular components and circuitry of the recruitment process. We define the protocadherins Fat and Dachsous as a bidirectional ligand-receptor system that is controlled by vestigial to generate the feed-forward signal. Further, we show that the signal is transduced by the conserved Warts-Hippo tumor suppressor pathway via activation of its transcriptional effector Yorkie. Finally, we propose that Wingless propels wing growth by fueling a wave front of Fat-Dachsous signaling and Yorkie activity that propagates vestigial expression from one cell to the next.Under normal conditions, animals and their various body parts grow until they achieve a genetically predetermined size and shape\u2014a process governed by secreted organizer proteins called morphogens. How morphogens control growth remains unknown. In  Growth is a fundamental property of animal development. Under normal conditions, animals of a given species, as well as their various body parts, achieve a characteristic size, shape, and pattern under tight genetic control. However, the basis of this control is poorly understood.Drosophila wing, the morphogens Wingless  and Decapentaplegic  drive a rapid \u223c200-fold increase in cell number and mass that occurs during larval life Morphogens, such as secreted factors of the Wingless/Int (Wnt), Bone Morphogenetic Protein (BMP), and Hedgehog (Hh) families, control growth. For example, in the classic paradigm of the Another system involved in growth is the evolutionarily conserved Warts-Hippo tumor suppressor pathway Drosophila, two protocadherins, Dachsous (Ds) and Fat (Ft), have been implicated as a ligand-receptor pair that acts, via the atypical myosin Dachs (D), to regulate Wts kinase activity In Ft and Ds are also important for planar cell polarity (PCP), in which cells within epithelial sheets adopt a common orientation, e.g. as manifest by their secreting hairs that point in the same direction Drosophila wing growth by morphogen vestigial (vg), the selector gene that defines the wing state vg expressing cells to send a feed-forward (FF) signal that induces neighboring cells to activate vg in response to Wg vg expression from cell to cell in response to Wg spreading from the border cells.Recently, we defined a new mechanism for the control of vg expression and initiate a new cycle of FF signaling. Based on these findings, we posit that Wg (and likely Dpp) promote wing growth by fueling the propagation of a wave front of Ft-Ds signaling that transiently suppresses the Wts-Hpo pathway and elevates Yki activity to recruit new cells into the wing primordium.In our initial analysis of the recruitment process, we speculated that Ft and Ds might be involved in the FF mechanism vg Boundary Enhancer (BE) to generate a stripe of vg expressing \u201cborder cells\u201d vg expression in surrounding cells via the vg Quadrant Enhancer (QE)   [4],[5],apterous (ap) in D, but not V, cells ap null discs (henceforth oap discs), the D-V segregation fails, vg and wg expressing border cells are not specified, and the nascent wing primordium is subsequently lost  and ds, itself, are expressed at peak levels in complementary domains that abut at the wing periphery, fj in the vgON domain . Such clones express moderate levels of exogenous Vg, a few fold lower than the peak endogenous level observed in wild type discs, and rescue wing development cell-autonomously fj-lacZ and repress ds-lacZ . Hence, prospective wing cells in these discs behave as if they have constitutively activated the FF signal transduction pathway but can mount only a weak QE response owing to the low levels of cryptic Wg available As described above, imordium  and 2B [o apoft discs is Wg dependent. First, the QE response is abolished in clones of o Dfz2ofz cells, which are unable to transduce Wg (UAS.Nrt-wg clones) drive peak levels of Vg and 5XQE.DsRed expression in o apoft discs, both within the clones and in abutting cells . These results confirm that ods clones serve as ectopic sources of FF signal, capable of inducing QE-dependent vg expression in neighboring cells, provided that the responding cells also receive Wg.In the presence of Nrt-Wg, ame disc . In the F signal : they inoft clones elicited a strictly cell autonomous response, both in Nrt-Wg expressing oap discs  and hence should be potent sources of FF signal; nevertheless they behave as if devoid of the capacity to signal. Note that this failure cannot be attributed to a generic inability of oft cells to send intercellular signals. First, oft clones repolarize their neighbors, whereas o ftods clones do not, indicating that they have the capacity to send the Ds PCP signal oft clones in the wing primordium can also send DSL-Notch, Wg, and Dpp signals  and generate ectopic FF signal in oap discs, as monitored by the induction of 5XQE.DsRed expression in adjacent wild type cells  that wing cells require Ft to generate FF signal and (ii) that non-wing cells require both Ft and Ds to receive the signal.Although wing cells require Ft, but not Ds, to send the FF signal, cells undergoing recruitment are also in position to receive an opposing Ds signal coming from non-wing cells on the other side, raising the possibility that this Ds input may also contribute to activating the QE and recruiting cells into the wing primordium.oap discs and asked if the resulting disparity in Ds signaling across the clone border is sufficient to induce the QE response in surrounding cells.To assess this, we generated Ds over-expressing clones in UAS.ds cells in oap discs generate levels of Ds that are several fold higher than endogenous Ds . In the absence of exogenous Wg, such UAS.ds clones had little effect on surrounding cells, only occasionally inducing 5XQE.DsRed expression just outside the clone (unpublished data). However, when supplemented with exogenous Wg (using co-expression of a UAS.wg transgene), most UAS.ds clones induced 5XQE.DsRed expression both within the clone and in surrounding cells . Hence, if the FF signal is transduced by the Wts-Hpo pathway, manipulations that promote Yki action , as expected if D is not available to block Wts activity in the absence of Ds and/or Ft.First, we examined the consequences of generating ods and od clones in oUAS.wg ap discs. Under these conditions, the ods clones both expressed Vg and induced Vg expression in neighboring wild type cells but failed to induce detectable expression in abutting cells belonging to the od clone, resulting in their exclusion from the rescued wing pouch . In this context, Ds and Ft are thought to function as a ligand-receptor pair, with tissue-wide gradients of Ds signal serving to activate Ft to appropriate levels within each cell To date, Ft-Ds signaling has been studied in two contexts: the control of Yki target genes in tissue growth and the orientation of cell structures in PCP. Most work on tissue growth has focused on Yki target genes that control basic cell parameters, such as survival, mass increase, and proliferation  and non-wing (vgOFF) cells. Further, we posit that these differentials are transduced in cells undergoing recruitment  that work in conjunction with FF propagation see also . And thiDrosophila wing, Wts-Hpo activity and YAP appear to function in these contexts in a manner that is distinct from their generic roles in the regulation of cell survival, growth, and proliferation, namely as part of an intercellular signaling mechanism that specifies cell type. We suggest that this novel employment of the pathway constitutes a new, and potentially general, mechanism for regulating tissue and organ size.Our identification of Ft-Ds signaling, the Wts-Hpo pathway, and Yki as key components of the FF recruitment process provides a striking parallel with the recently discovered involvement of the Wts-Hpo pathway and Yki/YAP in regulating primordial cell populations in vertebrates, notably the segregation of trophectoderm and inner cell mass in early mammalian embryos (i) Flp/FRT mediated mitotic recombination Hsp70.flp transgene . Wing discs from mature third instar larvae were dissected, fixed, and processed for immuno-fluorescence by standard methods, using anti-Vg, anti-Wg, anti-HA, and anti-\u03b2Gal antisera  by heat shock induced expression of an vg QE activity was monitored by expression of 1XQE.lacZ and 5XQE.DSRed reporter transgenes as well as by the expression of Vg protein in the absence of DSL-Notch signaling fj-lacZ enhancer trap allele P1fjvg expression. All four assays gave similar results, with the 5XQE.DSRed and fj-lacZ reporters showing the greatest sensitivity.http://flybase.bio.indiana.edu/) The following amorphic mutant alleles and transgenes were employed Exel6006, UA071ds, 2D60bds, E1ex, P1fj, 15ft, P21fz, C1Dfz2 , 83b27Rvg, and X1wts.Mutant alleles: apintraUAS.N, UAS.Nrt-wg, UAS.wg, Tub\u03b11>GFP,y+>vg, C765.Gal4, nub.Gal4, Tub\u03b11>Gal80>Gal4, GSUAS.ds, UAS.ft, UAS.d, UAS.yki, Hsp70.GFP.Transgenes: Exact genotypes, by Figure panel:56f fjP1/+.y w 5XQE.DsRed/y w Hsp70.flp; FRT39 ap(2A) 56f fjP1/FRT39 ap56f.y w 5XQE.DsRed/y w Hsp70.flp; FRT39 ap(2B) 2D60b FRT39 ap56f vg83b27R/+.y w 5XQE.DsRed/y w Hsp70.flp; ds(2C) 2D60b FRT39 ap56f vg83b27R/FRT39 ap56f.y w 5XQE.DsRed/y w Hsp70.flp; ds(2D) 56f vg83b27R 5XQE.DsRed/FRT39 ap56f vg83b27R fjP1; Tub\u03b11>flu-GFP,y+>vg/+.y w Hsp70.flp/y w Hsp70.flp; ap(2E) 56f vg83b27R 5XQE.DsRed/ds2D60b FRT39 ap56f vg83b27R; Tub\u03b11>flu-GFP,y+>vg/+.y w Hsp70.flp/y w Hsp70.flp; ap(2F) 56f/Hsp70.flu-GFP FRT39 ap56f fjP1; Tub\u03b11>flu-GFP,y+>vg UAS.Nrt-flu-wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; FRT39 ap(2G) 56f/ds2D60b FRT39 ap56f vg83b27R; Tub\u03b11>flu-GFP,y+>vg UAS.Nrt-flu-wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; FRT39 ap(2H) UA071 FRT39 ap56f/dsUA071 FRT39 ap56f; UAS.wg/+.y w 5XQE.DsRed/y w Hsp70.flp; ds(3A) 15 FRT39 ap56f/dsUA071 ft15 FRT39 ap56f fjP1.y w 5XQE.DsRed/y w 5XQE.DsRed; ft(3B) UA071 ft15 FRT39 ap56f/dsUA071 ft15 FRT39 ap56f fjP1; Tub\u03b11>CD2,y+>Gal4/+.y w 5XQE.DsRed/y w 5XQE.DsRed; ds(3C) 15 FRT39 ap56f/ft15 FRT39 ap56f; fzP21 Dfz2C1 FRT2A/Hsp70.CD2 Hsp70.flu-GFP FRT2A.y w 5XQE.DsRed/y w Hsp70.flp; ft(3D) 15 FRT39 ap56f/ft15 FRT39 ap56f; UAS>CD2,y+>Nrt-flu-wg C765.Gal4/+.y w 5XQE.DsRed/y w Hsp70.flp; ft(3E) UA071 FRT39 ap56f/Hsp70.flu-GFP Tub\u03b11.Gal80 FRT39 ap56f fjP1.y w 5XQE.DsRed/y w Hsp70.flp Tuba1.Gal4 UAS.GFPnls; ds(4A) 15 FRT39 ap56f/Hsp70.flu-GFP Tub\u03b11.Gal80 FRT39 ap56f fjP1; UAS.wg/+.y w 5XQE.DsRed/y w Hsp70.flp; ft(4B) UA071 ft15 FRT39 ap56f fjP1/Hsp70.flu-GFP Tub\u03b11.Gal80 FRT39 ap56f; C765.Gal4/+.y w 5XQE.DsRed/y w Hsp70.flp; ds(4C) UA071 FRT39 ap56f/Hsp70.flu-GFP FRT39 ap56f; UAS.Nrt-flu-wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; ds(4D) 15 FRT39 ap56f/Hsp70.flu-GFP FRT39 ap56f; UAS.Nrt-flu-wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; ft(4E) UA071 FRT39 ap56f/Hsp70.flu-GFP FRT39 ap56f; UAS>CD2,y+>Nrt-flu-wg C765.Gal4/1XQE.lacZ.y w Hsp70.flp/y w Hsp70.flp; ds(4F) UA071 Tub\u03b11.Gal80 FRT39 ap56f vg83b27R/Hsp70.flu-GFP FRT39 ap56f fjP1; UAS.ft/Tuba1.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; ds(5A) 15 Tub\u03b11.Gal80 FRT39 ap56f/Hsp70.flu-GFP FRT39 ap56f fjP1; UAS.ft/Tub\u03b11.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; ft(5B) 56f fjP1/FRT39 ap56f UAS.flu-wg; UAS.ft/Tub\u03b11>Gal80,y+>Gal4.y w 5XQE.DsRed/y w Hsp70.flp UAS.GFPnls; FRT39 ap(5C) 56f 1XQE.lacZ/dsUA071 FRT39 ap56f; UAS.ds/Tub\u03b11>Gal80,y+>Gal4 UAS.wg.y w 5XQE.DsRed/y w Hsp70.flp UAS.GFPnls; ap(5D) 56f/Hsp70.flu-GFP Tub\u03b11.Gal80 FRT39 ap56f fjP1; UAS.d/UAS.wg.y w 5XQE.DsRed/y w Hsp70.flp Tuba1.Gal4 UAS.GFPnls; FRT39 ap(6A) 56f UAS.flu-wg/FRT39 ap56f fjP1; Tub\u03b11>Gal80,y+>Gal4 UAS.yki/+.y w 5XQE.DsRed/y w Hsp70.flp UAS.GFPnls; FRT39 ap(6B) 56f/ap56f UAS.flu-Nrt-wg; FRT82 wtsx1/FRT82 Hsp70.flu-GFP.y w Hsp70.flp/y w Hsp70.flp; nub-Gal4 FRT39 ap(6C) e1 FRT39 ap56f/Hsp70.flu-GFP FRT39 ap56f fjP1; UAS.wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; ex(6D) UA071 Hsp70.flu-GFP FRT39 ap56f/dGC13 FRT39 ap56f fjP1; UAS.wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; ds(7A) GC13 FRT39 ap56f fjP1/dGC13 FRT39 ap56f; Tub\u03b11>flu-GFP,y+>vg UAS.Nrt-flu-wg/C765.Gal4.y w 5XQE.DsRed/y w Hsp70.flp; d(7B) UA071 dGC13 FRT39 ap56f/Hsp70.flu-GFP Tuba1.Gal80 FRT39 ap56f fjP1; UAS.wg/+.y w 5XQE.DsRed/y w Hsp70.flp Tuba1.Gal4 UAS.GFPnls; ds(7C) UA071 FRT39 ap56f/dsUA071 FRT39 ap56f; UAS.Nrt-flu-wg/Tub\u03b11>Gal80,y+>Gal4.y w 5XQE-DsRed/y w Hsp70.flp; ds(S1A) 56f UAS>CD2,y+>Nrt-flu-wg/nub-Gal4 FRT39 ap56f; FRT82 wtsx1/FRT82 Hsp70.flu-GFP.y w Hsp70.flp/y w Hsp70.flp; ap(S1B) 56f 1XQE.lacZ/FRT39 ap56f; UAS>CD2,y+>Nrt-flu-wg C765.Gal4/UAS.yki.y w Hsp70.flp/y w Hsp70.flp; ap(S1C) (S2A) as (4E).15 FRT39 ap56f/Df(2L)Exel6006 Hsp70.flu-GFP FRT39 ap56f; UAS.Nrt-flu-wg/C765.Gal4.y w 5XQE-DsRed/y w Hsp70.flp; ft(S2B) 15 Tuba1.Gal80 FRT39 ap56f/Hsp70.flu-GFP FRT39 ap56f; UAS>CD2,y+>Nrt-flu-wg C765.Gal4/1XQE.lacZ.y w Hsp70.flp/y w Hsp70.flp; ft(S2C) cx4 FRT39 ap56f/Hsp70.flu-GFP Tub\u03b11.Gal80 FRT39 ap56f; UAS.Nintra/1XQE.lacZ.y w Hsp70.flp Tub\u03b11.Gal4 UAS-GFPnls/y w Hsp70.flp; wg(S3A) 15 wgcx4 FRT39 ap56f/Hsp70.flu-GFP Tub\u03b11.Gal80 FRT39 ap56f; lqf1227 Hsp70-CD2 FRT2A UAS.Nintra/+.y w 5XQE-DsRed/y w Hsp70.flp Tub\u03b11.Gal4 UAS-GFPnls; ft(S3B) 15 FRT39 ap56f/Tub\u03b11.Gal80 FRT39; UAS.wg/+.y w Hsp70.flp Tub\u03b11.Gal4 UAS-GFPnls/y w Hsp70.flp; ft(S3C) 15 FRT39 ap56f/Tub\u03b11.Gal80 FRT39; UAS.dpp/+.y w omb-lacZ/y w Hsp70.flp Tub\u03b11.Gal4 UAS-GFPnls; ft(S3D) Figure S1Quadrant enhancer activity is Wingless dependent in oap discs that lack either Dachsous or Warts or that over-express Yorkie. (A) A UAS.Nrt-wg clone in a o apods disc. Both Vg and 5XQE.DsRed are expressed at peak levels in the clone and in surrounding cells that abut the clone, as observed for UAS.Nrt-wg clones in o apoft discs , Vg is strongly up-regulated in the UAS.Nrt-wg clones and abutting cells. (C) UAS.Nrt-wg clones generated in an oUAS.yki ap disc; same outcome as in (A), except a 1XQE.lacZ transgene was used instead of the 5XQE.DsRed transgene.po discs . (B) UAS(3.99 MB TIF)Click here for additional data file.Figure S2Exceptional cases of local non-autonomous Quadrant enhancer activity associated with oft clones can be attributed to induction by their sibling 2\u00d7+ft clones. (A) A oft clone (marked by the absence of GFP) associated with local, non-autonomous activity of the 5XQE.DsRed transgene (appears yellow in A') in an o UAS.Nrt-wgap disc. Note that this non-autonomous expression is associated with a sibling 2\u00d7+ft clone . In this experiment, 26/43 oft clones were associated with strictly cell-autonomous QE activity , and in the remaining 10/17 cases, 9/10 had no detectable twin, and 1/10 had a twin clone located elsewhere. Thus, the majority of oft clones analyzed in this experiment showed a strictly cell-autonomous response, and in 7/8 cases in which local, non-autonomous 5XQE.DsRed expression was observed and a 2\u00d7+ft twin survived, the twin spot was associated with the 5XQE.DsRed expression. Based on these results, we attribute the exceptional cases of non-autonomous 5XQE.DsRed expression associated with oft clones to signaling by their 2\u00d7+ft sibling clones, a conclusion further supported by experiments in panels (B) and (C). (B) A oft clone generated and marked as in (A), except under conditions in which its sibling 2\u00d7+ft clone died, owing to homozygosity for Df(2L)Exel6006. Note the strictly cell-autonomous expression of the 5XQE.DsRed transgene. 39/45 clones generated in this experiment behaved in this way; 6/45 showed local non-autonomy. We have not determined how quickly the sibling 2\u00d7+ Df(2L)Exel6006ft clones die after being generated in this experiment; it is possible that rare 2\u00d7+ Df(2L)Exel6006ft clones survive long enough to induce self-sustaining vg and 5XQE.DsRed expression in neighboring cells prior to their loss. (C) A UAS.Nrt-wg 2\u00d7+ft clone  and its oft sibling clone (marked by the absence of GFP) in an oap disc. Note the association of the 2\u00d7+ft clone with ectopic Vg expression as well as the local induction of Vg expression by Nrt-Wg in neighboring cells . This result corroborates the evidence shown in (A) and (B), that 2\u00d7+ft clones generated in 1\u00d7+ apoft discs have the capacity to induce 5XQE-DsRed and vg expression.ty as in : of thesft+ twin , and the(2.19 MB TIF)Click here for additional data file.Figure S3oft clones can send Delta/Serrate/Lag2 (DSL), Wingless, and Decapentaplegic signals. (A) A intra wgoUAS.N clone generated in an oap disc. intraN encodes a constitutively active form of Notch; clones of intra wgoUAS.N cells in oap discs up-regulate the expression of the Notch ligands Delta and Serrate and activate Notch in adjacent cells, as visualized by the induction of a ring of ectopic, Wg-expressing D-V border cells encircling the clone . These ectopic border cells suffice to initiate the long-range propagation of QE-dependent vg expression in surrounding cells, as indicated by the broad halo of 1XQE-lacZ expression. (B) A intra wgo ftoUAS.N clone generated an oap disc. Essentially the same experiment shown in (A), except that the clones are also oft. The result is the same (except that a 5XQE-DsRed reporter was used in place of the 1XQE-lacZ reporter), indicating that cells in the clone can send DSL signals to the surround, even though they are devoid of Ft. (C) A oUAS.wg ft clone generated in a wild type disc. QE-dependent vg expression depends on the level of Wg input. As a consequence UAS.wg clones up-regulate Vg expression in surrounding cells within the wing pouch, as seen in this example, even though the clone is also oft. (D) A oUAS.dpp ft clone generated in a wild type disc. Ectopic Dpp expressed by the clone has induced ectopic omb-lacZ expression in the surround, even though the clone is oft.(3.03 MB TIF)Click here for additional data file."}
+{"text": "The expression of retinoblastoma (Rb), c-Myc and Bcl-2 proteins was studied by immunohistochemical methods in 104 cases of renal adenocarcinoma. One tumour was completely negative for Rb protein and altered expression pattern was detected in 36% of cases. A low fraction of Rb-positive nuclei was related to high grade (P = 0.016) and high mitotic index (P = 0.012). Twenty-eight per cent of the tumours expressed c-Myc in cancer cell nuclei and 87% showed cytoplasmic positivity. Cytoplasmic expression of c-Myc was related to high grade (P = 0.002), while nuclear expression of c-Myc was related to small tumour diameter (P = 0.034), low T category (P = 0.04), low mitotic index (P = 0.019) and expression of c-ErbB-2 (P = 0.0007). Overexpression of c-myc predicted favourable outcome in M0 tumours (P = 0.0157). Bcl-2 was expressed in 20% of tumours and it was related to small tumour size (P < 0.0001), low T category (P < 0.0001), lack of venous invasion (P = 0.008), node negativity (P = 0.015) and absence of metastasis (P = 0.017). In multivariate analysis the expression of Rb, Bcl-2 and c-Myc had no independent prognostic value over T category (P < 0.001), mitotic index (P = 0.008) and combined nuclear grade (P = 0.056)."}
+{"text": "P= 0.08). A negative association was found between ARs and cell proliferative activity: MIB-1 scores were higher (25.4%) in AR-negative than in AR-positive cases . A strong positive association (P= 0.0001) was found between ARs and oestrogen receptors (ERs). In univariate analysis, ARs (as well as ERs and PGRs) were not correlated with overall survival; tumour histological grade (P= 0.02), size (P= 0.01), p53 expression (P= 0.0008) and MIB-1 scores (P= 0.0003) had strong prognostic value. In multivariate survival analysis, only p53 expression (P= 0.002) and histological grade (P= 0.02) retained independent prognostic significance. In conclusion, the lack of association between AR and most clinicopathological features and survival, together with the absence of prognostic value for ER/PGR status, suggest that MBCs are biologically different from female breast carcinomas and make it questionable to use antihormonal therapy for patients with MBC. \u00a9 1999 Cancer Research CampaignAndrogen receptor (AR) expression was retrospectively analysed in 47 primary male breast carcinomas (MBCs) using a monoclonal antibody on formalin-fixed, paraffin-embedded tissues. AR immunopositivity was detected in 16 out of 47 (34%) cases. No association was found with patient age, tumour stage, progesterone receptor (PGR) or p53 protein expression. Well-differentiated MBCs tended to be AR positive more often than poorly differentiated ones ("}
+{"text": "P for trend = 0.022, adjusted odds ratio 1.20, 95% confidence interval 1.07\u20131.33 respectively. No association was found between HHV-6 and ALL, CML or RAEB. \u00a9 1999 Cancer Research CampaignThe relationships between acute myeloid leukaemia (AML), acute lymphocytic leukaemia (ALL), chronic myeloid leukaemia (CML) and refractory anaemia with excess of blasts (RAEB) and human herpes virus (HHV)-6 antibody level were investigated in a multicentre case-control study. An association between increased HHV-6 seropositivity and geometric mean titre ratio with AML was shown:"}
+{"text": "We have studied the cytotoxicity of bleomycin (4--10 u/kg/day for 6 days) given by continuous i.p. infusion (using an osmotic minipump) compared to daily i.p. bolus administration, against P388 leukaemic spleen colony-forming-units(LCFU-S). Continuous i.p. bleomycin at 8 u/kg/day caused a 0.5 log greater reduction of LCFU-S than did an identical dose given by intermittent bolus administration. The infusion minipump provided constant bleomycin plasma levels of 0.62 +/- 0.03 mu/ml and a total plasma AUC (area under the plasma decay curve) of 89.0 mu.h/ml for 6 days at 8 u/kg/day. Intermittent bolus bleomycin at 8 u.kg/day had a terminal-phase plasma t1/2 of 15 min and a total 6-day plasma AUC of 90.8mu.h/ml. These pharmacokinetic data validate the osmotic minipump as a constant drug-delivery system, and suggest that the two administration schedules resulted in equal total bleomycin dosages. Although high peak bleomycin plasma levels (i.e. 32 mu/ml) were achieved with the intermittent bolus administration, continuous-infusion bleomycin's greater inhibition of LCFU-S was probably related to the drug's schedule-dependent cell-killing characteristics. The results of this study provide further rationale for the continuing use of infusion bleomycin schedules in cancer patients."}
+{"text": "P < 0.0001), 100 nM NPY (P < 0.0001), 100 nM VIP (P < 0.001), 100 nM CGRP (P < 0.001), 100 nM SP (P < 0.01), and 0.1 mM histamine (P < 0.01), whereas 0.1 mM 5-HT, mM acetylcholine, and 1 \u03bcM isoproterenol             (\u03b2-adrenergic agonist) had no effect. Proliferation(incorporation of tritiated thymidine) was increased by CGRP (P = 0.004) and histamine (P < 0.02), but decreased by isoproterenol (P = 0.002), 5-HT (P = 0.003), and acetylcholine (P < 0.05). The percentage of multinucleate cells was decreased after isoproterenol (2.5%), and increased after 5-HT (21.3%), GABA (15%), and histamine (15.1%). Compared to controls, thymulin in the supernatant was decreased after challenge with acetylcholine (52%), isoproterenol (71%), 5-HT (73%), and histamine (84%). This study demonstrates direct effectsof neuropeptides and neurotransmitters on functional aspects of cultured thymic epithelial cells.To determine if major thymic neuropeptides and neurotransmitters can directly influence the functional activity of cultured rat thymic epithelium, neuropeptides and neurotransmitters were applied, and intercellular communication, proliferation, and thymulin secretion assessed. After injections of a mixture of lucifer yellow dextran (too large to pass gap junctions) and cascade blue (which does) into single cells, some neuropeptides decrease dye coupling: 0.1 mM GABA ("}
+{"text": "The atomic arrangement consists of an infinite long-chain polyphosphate organization. Chains, with a period of four PO4 tetra\u00adhedra, run along the a-axis direction. Two other polymorphs of this phosphate are known, in space groups P21/n and C2/c.The title compound, potassium yttrium polyphosphate, KY(PO DOI: 10.1107/S1600536808013925/br2073Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:"}
+{"text": "The 4,6-dihydroxy\u00adbenzene-1,3-disulfonate anion, acting as a counter-ion, is also located on the mirror plane. The crystal packing is stabilized by O\u2014H\u22efO hydrogen bonds, forming a three-dimensional supra\u00admolecular network.In the title compound, [Zn(CH DOI: 10.1107/S1600536810006525/hy2283Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:"}
+{"text": "Since 1997, the largest epidemic of highly pathogenic avian influenza (H5N1) ever recorded has caused 172 human and several billion bird deaths. Recently administered questionnaires determined that live poultry exposures have declined by \u224863% in Hong Kong since 2004 and that, in Vietnam, domestic backyard exposures to poultry are likely more important than retail exposures. MappingIn Hong Kong, random household telephone interviewing of 1 adult >17 years of age selected by Kisch grid (which randomizes selection of persons within households) was conducted from December 2005 through 2006 from a list of 5,000 numbers. Simultaneously in Vietnam, stratified cluster sampling was carried out throughout 2 districts in each of 5 northern provinces. Within 3 of these provinces, 1 district with and 1 without an HPAI epidemic history were selected. Within each district, 1 urban and 1 rural commune each provided 100 households randomly selected from electoral rolls. Kisch grid selected 1 adult from each household for face-to-face interviews.Respondents estimated their live poultry purchase frequency and touching at purchase ; 64% of respondents were women and 36% were men; their median age, 44 years . VietnamIn Hong Kong in 2006, 18,586 standardized purchases averaged 10.56 chickens/household/year . This is a territory-wide gender-adjusted rate of 11.05 chickens/household/year, which indicated that 22,673,000 live chickens were purchased during the preceding year, 41% fewer than in 2004. Households buying poultry bought an average of 15.6 chickens/household/year. Among respondents personally buying, 7.5% touched the poultry during purchasing (compared with 11% in 2004), giving \u22481,700,500 exposures/year. Adjustment for touching frequency and gender differences in reported touching  gave \u22481,110,900 contacts  for Hong Kong in 2006, or 0.76 exposures/buying household/year . Applied retrospectively to Hong Kong 2004 data, this gave an adjusted exposure rate of 8.6% (95% CL 6.8\u201310.3), \u22483,311,300 contacts, and exposure frequency of 2.07 exposures/buying household/year (0.62/person/year) in 2004. These adjusted estimates indicate an absolute exposure decline of 3.7% (95% CL 2.25%\u20134.91%), a relative decline of 43% between 2004 and 2006. Less purchasing and touching reduced annualized buying exposures by 63% overall.2 = 45.57, df = 4, p<0.001), after adjustment for gender proportion and reported touching, was 63% (62%\u201364%). Estimated exposures in the surveyed provinces from buying were \u224813,097,000 \u00d7 0.63 = \u22488,251,000 exposures/year. When these rates were used, national per capita exposure estimates (assuming 4.49 persons/household) from touching when buying are \u224862,479,000 exposures/year, 2.24 exposures/person/year in buying households, 0.76 exposures/person/year overall.In Vietnam, respondents reported 10,659 standardized purchases, averaging 5.36 chickens/household/year, giving a gender-adjusted  rate of 5.43 chickens/household/year. Estimated number of live birds purchased in the sampled provinces  was 13,097,000 chickens per year. Buying households  buy on average 15.97 chickens per year, comparable to the Hong Kong 2006 purchase rate. Touching frequency during purchasing  that raised poultry, 92 (5%) ceased keeping poultry from February 2005 through February 2006 . HousehoWhile 34% (32%\u201336%) of households buy live chickens, 53% (52%\u201354%)  raise live poultry at home, and 12% (10%\u201313%) do both. Assume a 53% national average and, conservatively, that all persons within households rearing backyard poultry have at least weekly physical contact with their birds, bird eggs, or feces. Household size in the surveyed districts averages 3.38 persons . Thus, 224,685,500 exposures/year would occur from backyard poultry in surveyed districts, an average exposure within backyard poultry raising households of \u2248175 exposures/person/year. Households buying live poultry have 8,251,000 /820,080 = 10.1 exposures/household/year (2.99 exposures/person/year) from these purchases. Total purchase-related plus backyard exposure events then equal  + 224,685,500 = 232,968,300 exposures/year. Average household exposure is therefore \u224896 exposures/household/year (28 exposures/person/year) in sampled districts. If daily backyard exposure occurs, then there are \u22481,581,081,300 total exposures, \u2248655 exposures/household/year (194 exposures/person/year). Nationally, average household size is 4.49 persons. Hence, between \u22482,322,546,000 (weekly contact) and \u224815,882,953,000 (daily contact) exposures/year, \u2248127\u2013869 exposures/household/year occur nationally. If multiple contacts occur daily, these figures would be much higher.Epidemic and nonepidemic district-buying frequency CL overlapped (exposure 3.4 [1.9\u20134.8] chickens/household/year vs. nonexposure 5.8 [4.5\u20137.0] chickens/household/year). Dual adjusted touching frequencies were 69% (62%\u201376%) in epidemic and 60% (57%\u201363%) in nonepidemic districts, respectively. Backyard poultry were more common in epidemic districts (71% [67%\u201375%] vs. 45% [42%\u201348%]), where keeping poultry declined 17% (14%\u201320%) compared with 8% (6%\u201310%) in nonepidemic districts. Epidemic and nonepidemic districts had comparable average incomes .In Hong Kong, government import restrictions have reduced poultry availability by 41% from 2004 to 2006. Purchase and touching declines prompted by health education messages have together reduced exposure by \u224860%.www.worldbank.org/data/quickreference/quickref.html), live chickens costs $16.6\u2013$18.0 and $21.8\u2013$31.0  each in Hong Kong and in Vietnam, respectively. Hence, temptation to use sick, dying, or dead poultry is high, increasing the risk for human influenza (H5N1) infection (Fewer Vietnamese households bought live chickens, but those that did so bought at comparable frequencies to Hong Kong 2006 households. Chickens are relatively more expensive in Vietnam. Adjusted for purchasing power parity (Limitations include generalizing from 5 northern Vietnamese provinces to the country as a whole and using arbitrary estimates for backyard exposure frequency. Nonetheless, valuable data are presented on differential exposure patterns."}
+{"text": "Objective: Octreotide, a somatostatin analogue, hasbeen shown to prevent angiogenesis in diverse invitro models. We evaluated its effect on retinal neovascularizationin vivo, using a neonatal rat retinopathymodel.                    Methods: We used, on alternating days, hypoxia(10% O2) and hyperoxia (50% O2) during the first 14days of neonatal rats, to induce retinal neovascularization.Half of the rats were injected subcutaneouslywith octreotide 0.7 \u03bcg/g BW twice daily. Atday 18 the eyes were evaluated for the presence ofepiretinal and vitreal hemorrhage, neovascularizationand epiretinal proliferation. Octreotide pharmacokineticsand its effect on serum growth hormone(GH) and insulin-like growth factor I (IGF-I) wereexamined in 28 rats.                    Results: Serum octreotide levels were 667 \u03bcg/1 twohours after injection, 26.4 \u03bcg/1 after nine hours and3.2 \u03bcg/1 after 14 hours. GH levels were decreasedby 40% (p = 0.002) two hours after injection butthereafter returned to baseline. IGF-I levels were unchangedtwo hours after injection and were elevatedby 26% 14 hours after injection (p = 0.02). Epiretinalmembranes were highly associated with epiretinalhemorrhages (p < 0.001), while retinal neovascularizationwas notably associated with vitreal hemorrhages (p <  0.001).                    Conclusions: Twice-daily injections of octreotidefailed to produce sustained decrease in serum GH,but produced rebound elevation of serum IGF-I.Accordingly, no statistically significant effect of injectionson retinal pathology was noted. This finding,however, does not contradict our assumptionthat GH suppression may decrease the severity ofretinopathy."}
+{"text": "The crystal structure contains six sites in the asymmetric unit: two sites are statistically occupied by rare-earth atoms with Gd:Sc ratios of 0.967\u2005(4):0.033\u2005(4) and 0.031\u2005(3):0.969\u2005(3), one site (.m. symmetry) is occupied by Sc atoms, and three distinct sites (two of which with .m. symmetry) are occupied by Ge atoms. The rare-earth atoms form two-dimensional slabs with Ge atoms occupying the trigonal-prismatic voids.Gd DOI: 10.1107/S1600536809007211/mg2062Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:"}
+{"text": "We looked for associations between spontaneous apoptosis, p53 gene mutation, p53 protein accumulation, growth fraction, bcl-2 expression and histological parameters in 64 ovarian, four tubal and three peritoneal carcinomas. Apoptotic cells were detected with the TUNEL method. p53 gene variants were detected by the single-strand conformation polymorphism and were sequenced directly. P53, Ki-67 and bcl-2 protein expressions were detected immunohistochemically. A weighed multiple logistic regression model was applied. Apoptotic index (Al) ranged 0.02\u20130.18 (mean 0.11); proliferation index (PI) ranged 3\u201390% (mean 54%). p53 gene mutations were present in 51, p53 protein accumulation in 46, and diffuse bcl-2 expression in 29 of 71 tumours. The AI was positively associated with the presence of p53 gene mutation (P = 0.011). However, the PI included into the analysis did positively influence the AI (P = 0.02) and diminished the association with p53 gene mutation (P = 0.082). The AI was negatively associated with good histological differentiation (P = 0.0006), the serous tumour type (P = 0.002), and diffuse bcl-2 expression (P = 0.025). Strong bcl-2 expression was associated with endometrioid tumour type (P = 0.002). FIGO stage and p53 protein accumulation were the only parameters that influenced overall survival time. Thus, our results suggest that histological tumour type and grade are major determinants of spontaneous apoptosis in ovarian carcinomas;p53 alterations do not adversely but rather positively affect spontaneous apoptosis by increasing growth fraction. This, in turn, suggests p53-independency of spontaneous apoptosis in ovarian carcinomas. \u00a9 2000 Cancer Research CampaignChanges in cell survival contribute to tumour development, influence tumour biology and its response to chemotherapy."}
+{"text": "Lactobacillus plantarum LQ80 is a strain isolated from liquid feed for pigs. We determined the complete genome sequence of this strain using the PacBio RS II platform. LQ80 contained a single circular chromosome of 3,230,192 bp, with 44.66% G+C content and seven plasmids. Lactobacillus plantarum is found in various environmental niches, e.g., vegetable pickles  (https://www.qiagen.com/us/resources/resourcedetail?id=8b1b2b4f-3dca-4447-8d21-f6fd64c3a729), and a 5-kb library was constructed with shearing. One SMRT cell was used for sequencing on the PacBio RS II platform with a 360-min movie time. The result of 362,945 reads with a mean length of 1,519 bp was assembled with HGAP3, and five circular contigs were obtained, representing five plasmids . From this analysis, another four smaller plasmids were constructed.Strain LQ80 was cultured with de Man-Rogosa-Sharpe medium at 37\u00b0C. For the genome sequencing of LQ80, DNA was isolated at the early log phase. DNA was purified using a PowerClean DNA cleanup kit , followed by 20-kb library construction for P6-C4 chemistry with shearing. Five single-molecule real-time (SMRT) cells were used for sequencing on the PacBio RS II platform  with a 180-min movie time. We obtained 320,152 reads with a mean length of 5,268 bp.  (HGAP3) . Four ciThe LQ80 chromosome was 3,230,192 bp in length, with a G+C content of 44.66%. In addition, LQ80 had seven plasmids: pLQ801 , pLQ802 , pLQ803 , pLQ804 , pLQ805 10,218 bp), pLQ806 , and pLQ807 . The complete genome was annotated using NCBI Prokaryotic Genome Annotation Pipeline (PGAP)  and Micr bp, pLQ8L. plantarum LQ80 have been deposited in GenBank under accession numbers CP028977 (chromosome), CP028978 (pLQ801), CP028979 (pLQ802), CP028980 (pLQ803), CP028981 (pLQ804), CP028982 (pLQ805), CP028983 (pLQ806), and CP028984 (pLQ807).The complete genome sequences of the chromosome and seven plasmids of"}
+{"text": "Salmonella typhimurium (S. typhimurium) A1-R, a facultative anaerobe that is an auxotroph of leucine and arginine. The tumor-targeting efficacy of S. typhimurium A1-R was demonstrated in vivo and vitro using several malignant cell lines including melanoma, sarcoma, glioma, breast, pancreatic, colon, cervical, prostate, and ovarian cancers. Our laboratory also developed a patient-derived orthotopic xenograft (PDOX) model by implanting patient-derived malignant tumor fragments into orthotopic sites in mice. We reviewed studies of S. typhimurium A1-R against recalcitrant cancers. S. typhimurium A1-R was effective against all PDOX tumor models tested and showed stronger efficacies than chemotherapy or molecular-targeting therapy against some tumors. Furthermore, the synergistic efficacy of S. typhimurium A1-R when combined with chemotherapeutic agents, molecular-targeting agents, or recombinant methioninase was also demonstrated. We suggest potential clinical uses of this S. typhimurium A1-R treatment.We developed tumor-targeting  Streptococcus pyogenes (S. pyogenes). He then developed Coley\u2019s toxin, a mixture of killed S. pyogenes and Serratia marcescens, achieving clinical responses for many malignant tumors [Dr. William B. Coley began bacterial therapy of cancer using t tumors .Live bacteria can actively penetrate tumors to reach lesions distant from blood vessels, where chemotherapeutic drugs cannot be delivered, and damage malignant cells by several cytotoxic mechanisms . ObligatClostridium. butyricum (C. butyricum) M-55 resulted in oncolysis and accumulation in treated tumors [C. novyi-NT induced intratumor infection and necrosis [Salmonella typhimurium (S. typhimurium) VNP20009 attenuated by msbB and purI mutations, showed bacterial colonization of treated melanomas [S. typhimurium showed bacterial colonization in treated tumors after local or systemic administration [In clinical studies, limited antitumor efficacy has been shown thus far. d tumors ,7. Intraelanomas ,9. ObjecS. typhimurium is a facultative anaerobe. Green fluorescence protein (GFP)-labeled S. typhimurium A1-R developed by our laboratory has high tumor-targeting efficacy, due to the leucine\u2013arginine auxotroph, resulting in broad antitumor efficacy and limited adverse effects [S. typhimurium A1-R was quickly eliminated from normal organs including the liver and spleen seven days after intravenous administration [S. typhimurium A1-R remained at high density in CT26 tumors. In addition, tumors treated with S. typhimurium A1-R were significantly smaller than those treated with S. typhimurium VNP20009. The efficacy of S. typhimurium A1-R was demonstrated in orthotopic nude mouse models of prostate [S. typhimurium A1-R was also effective in metastatic cancer models [ effects . In a CTstration . In contprostate , breast prostate ,14, pancprostate ,16, and prostate , as wellprostate  and glioprostate ,20. S. tr models ,22.S. typhimurium A1-R against malignancies in patient-derived orthotopic xenograft (PDOX) nude mouse models, in which human tumors are orthotopically implanted in mice, to examine future clinical applicability.We review in the present report the therapeutic efficacy of We established PDOX tumors as follows: when the original tumors derived from primary sites, the PDOX tumors were implanted into the same primary sites in mice; when the original tumors derived from recurrent or metastatic sites, the PDOX tumors were implanted into original, recurrent, or metastatic sites in mice.S. typhimurium A1-R in PDOX models were identified. All of the 17 studies of human cancer of different histological types were evaluated and the efficacy of S. typhimurium A1-R was compared with the efficacy of chemotherapy  [S. typhimurium A1-R ranged from 5 \u00d7 105 colony-forming units (CFUs) to 1.5 \u00d7 108 CFUs. Intravenous injection twice weekly with a dose of 5 \u00d7 107 CFUs was used in most experiments. S. typhimurium A1-R treatment was performed either as a monotherapy or a polytherapy in combination with chemotherapeutic or molecular-targeting agents, or recombinant methioninase (rMETase), which reduces the plasma methionine on which cancer cells are addicted [s (i.v.) ,37,38,39s (i.v.) ,25, intrs (i.v.) ,28, or is (i.v.) . A singladdicted . PolytheS. typhimurium A1-R was evaluated via culture of resected specimens. A fluorescent microscope was used to detect GFP-expressing S. typhimurium A1-R in tumors grown in PDOX models treated with S. typhimurium A1-R i.t., i.p., i.v., or i.a. injection [S. typhimurium A1-R grown from a Ewing\u2019s sarcoma PDOX tumor [S. typhimurium A1-R were present in STS PDOX tumors, including undifferentiated STS [S. typhimurium A1-R were not detectable in adjacent muscles, suggesting selective tumor-targeting efficacy [The tumor-targeting efficacy of njection ,34,37,38OX tumor . Abundanated STS , pleomorated STS , follicuated STS , melanomated STS ,29,30,34ated STS , a GIST ated STS , and a Cefficacy ,37. The treatment efficacy of S. typhimurium A1-R was confirmed in all PDOX models . SignifiS. typhimurium A1-R showed stronger antitumor efficacy than gemcitabine, cisplatinum, or fluorouracil treatment in the pancreatic PDOX model [S. typhimurium A1-R was more effective than cisplatinum [S. typhimurium A1-R resulted in greater tumor growth inhibition compared to doxorubicin treatment [S. typhimurium A1-R showed stronger efficacy than imatinib in the GIST PDOX model [OX model . In the platinum ,32. In treatment ,35,39. MOX model .S. typhimurium A1-R with chemotherapy [S. typhimurium A1-R had additional efficacy when combined with gemcitabine or gemcitabine plus bevacizumab [S. typhimurium A1-R with temozolomide or vemurafenib significantly reduced tumor growth compared to monotherapy with these agents [S. typhimurium A1-R, rMETase, and cisplatinum was more effective than double therapy using S. typhimurium A1-R with rMETase, or monotherapy of these agents [Synergistic treatment efficacy was observed with combination of otherapy ,36, moleotherapy ,26,29,30otherapy ,35. In pacizumab ,36. Addie agents .S. typhimurium A1-R-treated tumors showed extended necrosis compared to untreated tumors. As an example, S. typhimurium A1-R caused central tumor necrosis to a large extent in the Ewing\u2019s sarcoma PDOX model, while the untreated tumors grew without necrosis (S. typhimurium A1-R treatment resulted in changes in sarcoma cell shape but not necrosis [S. typhimurium A1-R treatment in combination with cisplatinum and rMETase resulted in tumor necrosis. Moreover, S. typhimurium A1-R showed more extensive necrosis when combined with chemotherapy or molecular-targeting agents than S. typhimurium A1-R monotherapy on undifferentiated STS and melanoma PDOX models [S. typhimurium A1-R induced a higher degree of necrosis in several PDOX models including pancreatic cancer, STS, osteosarcoma, and GIST [S. typhimurium A1-R led to more extensive necrosis than intravenous administration in the osteosarcoma PDOX model [Established tumors in the PDOX model had a similar morphologic appearance to the original patient tumor A,B. S. tnecrosis C\u2013G. In anecrosis . S. typhX models ,29,30. Wand GIST ,32,33,37OX model . These rS. typhimurium A1-R compared to the untreated control.None of the PDOX experiments showed adverse effects, in terms of significant weight loss, in mice treated with S. typhimurium A1-R against recalcitrant-caner PDOX models, indicating advantages that S. typhimurium A1-R may have over chemotherapy. Therefore, for rare malignancies or cancers of unknown primary origin, for which effective treatments have not been established, S. typhimurium A1-R treatment can be a good candidate. Moreover, for highly aggressive malignancies such as pancreatic cancer or melanomas, S. typhimurium A1-R was highly effective when combined with chemotherapy or molecular-targeting therapy. In addition, adverse effects were shown to be limited. Therefore, S. typhimurium A1-R treatment has clinical potential. PDOX models are theoretically better at mimicking the human disease than heterotopic tumors, increasing the robustness of drug discovery studies. The present review demonstrates the strong antitumor efficacy of S. typhimurium penetrated the cancer cells in vitro by being attracted to small molecules such as ribose and serine [S. typhimurium A1-R targeted tumors in several PDOX mouse models. The antitumor efficacy of S. typhimurium A1-R against many kinds of cancer cell lines was demonstrated and suggested that S. typhimurium A1-R kills cancer cells directly [S. typhimurium A1-R expanded and burst, resulting in loss of viability [S. typhimurium A1-R, a facultative anaerobe, can grow under anaerobic condition [S. typhimurium A1-R induces central tumor necrosis (Salmonella plays the role of inducing antitumor immune responses in an immunocompetent model [Salmonella enhances both innate and adaptive immunity. Salmonella induces cytokine production, including interferon-\u03b3, via Toll-like receptor 4 signaling [S. typhimurium resulted in recruitment of CD8+ lymphocytes, CD4+ lymphocytes and B lymphocytes as well as macrophages and granulocytes in the tumor [S. typhimurium A1-R was correlated with CD8+ lymphocyte infiltration into treated tumors in a pancreatic cancer syngeneic immunocompetent mouse model [S. typhimurium A1-R acts as a decoy. It induces the cancer cells to leave the chemo-sensitive state of the cell cycle, making the cancer cells highly sensitive to chemotherapy [S. typhimurium A1-R. d serine ,42. The directly ,43,44. Uiability . Importaondition ,44. As anecrosis . In addint model . Salmoneignaling . Upregulignaling ,47. Avogse model . Moreoveotherapy . These fS. typhimurium A1-R against recalcitrant cancer PDOX models. Pre-clinical efficacy studies of S. typhimurium A1-R were completed and only a toxicity test needs to be performed to enable S. typhimurium A1-R to begin phase I clinical studies.The present review discusses the antitumor efficacy of"}
+{"text": "Siphoviridae bacteriophage, characterized by an unusual prolate capsid, containing a 72,658-base-pair double-stranded DNA genome with 132 predicted protein-coding genes. Conserved among cluster O bacteriophages, the Ryadel genome contains 31 copies of a unique 17-bp sequence with dyad symmetry.Mycobacteriophage Ryadel is a newly isolated cluster O  Siphoviridae bacteriophage, characterized by an unusual prolate capsid, containing a 72,658-base-pair double-stranded DNA genome with 132 predicted protein-coding genes. Conserved among cluster O bacteriophages, the Ryadel genome contains 31 copies of a unique 17-bp sequence with dyad symmetry.Mycobacteriophage Ryadel is a newly isolated cluster O  Mycobacterium smegmatis mc2155 at 37\u00b0C for 48 hours and resulted in small-sized lytic plaques. Ryadel was purified by collecting virus from well-isolated plaques from the direct isolation and two successive rounds of serial dilutions. Negative-staining transmission electron microscopy . Soil samples were placed in 7H9 liquid medium, and the supernatant was passed through a 0.22-\u00b5m filter for direct isolation of bacteriophages. Filtered supernatant was incubated with croscopy  of isolahttps://blast.ncbi.nlm.nih.gov/) whole-genome alignment (MG099943) and Catdawg (GenBank accession number KF017002)  (http://phagesdb.org/DNAMaster/), and PECAAN (https://pecaan.kbrinsgd.org/). Mycobacteriophage Ryadel was predicted to contain 132 protein-coding genes, and no tRNA genes were identified by ARAGORN v1.2.38  encode DNA polymerase III sliding clamp beta, Ku-like double-stranded DNA (dsDNA) break-binding protein, and ParB-like dsDNA partitioning protein.Glimmer v3.02 , 7 and Gor.org/) , DNA Mas v1.2.38  or tRNAs v1.2.38 . Putativ v1.2.38  and NCBI v1.2.38 . SimilarMH590592. Raw reads are available in the SRA under accession number SRX4721442.The mycobacteriophage Ryadel genome is available at GenBank under accession number"}
+{"text": "Few studies have compared the effectiveness of brief training courses on point-of-care ultrasound (POCUS) skill acquisition of novice attending physicians vs. trainees. The purpose of this study was to evaluate the change in POCUS image interpretation skills and confidence of novice attending physicians vs. trainees after a 1-day POCUS training course.A 1-day POCUS training course was held in March 2017 in Japan. A standardized training curriculum was developed that included online education, live lectures, and hands-on training. The pre-course assessment tools included a written examination to evaluate baseline knowledge and image interpretation skills, and a physician survey to assess confidence in performing specific ultrasound applications. The same assessment tools were administered post-course, along with a course evaluation. All learners were novices and were categorized as trainees or attending physicians. Data were analyzed using two-way analysis of variance.p\u2009<\u20090.001). The post-course physician confidence scores in using ultrasound significantly increased in all skill categories for both groups. Both trainees and attending physicians demonstrated similar improvement in their post-course test scores and confidence with no statistically significant differences between the groups. The course evaluation scores for overall satisfaction and satisfaction with faculty members\u2019 teaching skills were 4.5 and 4.6 on a 5-point scale, respectively.In total, 60 learners attended the course, and 51 learners (85%) completed all tests and surveys. The 51 novice learners included 29 trainees  and 22 attending physicians . The mean pre- and post-course test scores of novice trainees improved from 65.5 to 83.9% while novice attending physicians improved from 66.7 to 81.5%  contains supplementary material, which is available to authorized users. Point-of-care ultrasound (POCUS) is defined as ultrasonography at the patient\u2019s bedside that is performed in real-time by a physician caring for the patient . In contMultiple studies have demonstrated the effectiveness of POCUS training courses on knowledge and skills acquisition by different learner groups. These courses have varied in duration, content, delivery, and target audience , 8, 12. Even though POCUS ultrasound training courses have been proven to be effective, , 8, 12 tAs a preparatory step, a 2-day train-the-trainer course was conducted in November 2016 to standardize the educational curriculum for a future POCUS training course in March 2017. The train-the-trainer course curriculum was based on the principles shown to be effective for POCUS skills training \u201315. The The same twelve POCUS faculty that participated in the train-the-trainer course in November 2016 served as the faculty for a 1-day POCUS training course in March 2017 at the 14th Japanese Society of Hospital General Medicine Semi-Annual Meeting. The 1-day POCUS course was a separate session from the main conference and the enrollment cap was 60 participants. The 1-day POCUS course was geared toward novices learners and included pre-course internet-based modules, live lectures 3.5\u00a0h), and hands-on skills training with live models (1.5\u00a0h)  and included three focused cardiac ultrasound (FOCUS) sessions, and one session each for deep vein thrombosis, lung/diaphragm, and abdominal ultrasound. The faculty-to-learner ratio was 1:3 during these hands-on sessions. Learning objectives were displayed, and faculty used standardized printed materials as teaching aids at each station . We analyzed differences in examination and survey scores between the trainees and attending physicians. Data analyses were performed using R statistical software (R version 3.1.3.)In total, 60 learners attended the course, and 51 learners (85%) completed all tests and surveys. The 51 novice learners included 29 trainees  and 22 attending physicians  (Table\u00a0p\u2009<\u20090.001). However, there was no statistically significant difference between the pre-course examination scores (p\u2009=\u20090.80) nor the post-course examination scores (p\u2009=\u20090.43) between trainees vs. attending physicians. Descriptive statistics and an ANOVA table of the examination scores for each application are shown in Additional\u00a0file\u00a0The mean pre- and post-course written examination scores for all learners were 66.0% (standard deviation [SD] 12.9) and 82.8% (SD 9.0), respectively. The mean pre-course examination scores of trainees vs. attending physicians were 65.5% (SD 13.0) and 66.7 (SD 13.0), respectively. The mean post-course examinations scores of trainees vs. attending physicians were 83.9% (SD 9.0) and 81.5% (SD 9.0), respectively. Post-course examination scores were significantly improved compared with pre-course examination scores in both groups (es p\u2009=\u20090.3 betweenp\u2009<\u20090.001). However, there were no significant differences in improvement of post-course confidence scores between the trainees vs. attending physicians in all categories . The pre- and post-course self-evaluation survey results are shown in Additional\u00a0files\u00a0The mean confidence scores for all learners in the pre-course vs. post-course self-evaluation surveys were: general ultrasound skills, 2.37 (SD 0.90) vs. 3.32 (SD 0.71); focused cardiac ultrasound, 2.56 (SD 0.84) vs. 3.60 (SD 0.71); vascular diagnostics, 1.94 (SD 0.90) vs. 3.55 (SD 0.70); lung/diaphragm ultrasound, 1.77 (SD 0.76) vs. 3.30 (SD 0.70); and abdominal ultrasound, 2.95 (SD 0.97) vs 3.81 (SD 0.75). Post-course confidence scores were significantly higher compared with pre-course scores in all categories for both groups Additional file 2:Pre- and post-course physicians survey. (DOCX 17 kb)Additional file 3:Post-course satisfaction surveys. (DOCX 15 kb)Additional file 4:Descriptive statistics and an ANOVA table of the test scores for pre- and post-course written examinations. (DOCX 19 kb)Additional file 5:Descriptive statistics for the pre- and post-course self-evaluation survey. (DOCX 19 kb)Additional file 6:Analysis of variance table for pre- and post-course self-evaluation survey. (DOCX 22 kb)"}
+{"text": "Using the Meikirch model as a theoretical underpinning, the present study aimed to examine population health disparities by discerning age variations within and across rural urban areas. Secondary data analysis was conducted using the CDC\u2019s 2017 Behavioral Risk Factor Surveillance System. The study\u2019s outcome variables included physical health and mental health burden . A total sample of 96,568 adults were included with a mean age of 66.05 years (SD = 9.91). Individuals were classified in the following age groups: 43% middle-aged (45-64 years), 33% young-old (65-74 years), and 25% old-old adults (75+ years). The sample was largely female (61%), Non-Hispanic White (86%), and urban (67%). A series of chi-square tests of independence \u2013 post hoc tests when applicable \u2013 were completed. Overall, rural residents reported a higher prevalence of severe physical and mental health symptom burden. Regarding physical health burden, a significant difference was found within urban settings (X2(4) = 50.74, p < .001), where, unexpectedly, young-old adults reported the best physical health. Regarding mental health burden, a significant difference was found for both urban (X2(4) = 1661.72, p < .001)) and rural settings (X2(4) = 820.65, p < .001), with middle-aged adults reporting greater mental illness and the old-old adults reporting greater mental health resiliency. Findings suggest that a multidimensional framework of health usefully informs public health and clinical service interventions, identifying populations and locations in need ."}
+{"text": "Bacillus sp. strain YSP-3 was isolated from a salt lake.The halophilic, alkaliphilic bacterium  Bacillus sp. strain YSP-3 was isolated from a salt lake. It grows optimally at 8% (wt/vol) NaCl (pH 9.0). The draft genome is composed of 4,006 predicted genes. Genomic analysis showed that various genes are potentially involved in the adaptation mechanisms for osmotic stress and pH homeostasis.The halophilic, alkaliphilic bacterium  Bacillus sp. strain YSP-3 was isolated from a salt lake in Jilin, China. In sodium bicarbonate-buffered medium (Bacillus aurantiacus K1-5T (97.9 \u200a%) and Bacillus populi FJAT-45347T (97.01%). A phylogenetic tree was reconstructed by a neighbor-joining (B. aurantiacus K1-5T and B. populi FJAT-45347T (data not shown), which revealed that YSP-3 belonged to the genus Bacillus. To understand the adaptive strategies for survival under hypersaline conditions, draft genome sequencing of Bacillus sp. YSP-3 was performed using an Illumina platform.The halophilic and alkaliphilic bacterium d medium , growth d medium . The resde novo assembled using MicrobeTrakr plus v. 0.9.1 (http://microbetracker.org/). Genome annotation was processed using NCBI PGAP (www.ncbi.nlm.nih.gov/genome/annotation_prok). The total length of the draft genome sequence was 4,047,843\u2009bp and yielded 12 contigs, with a G+C content of 48.26%. Among the predicted 4,006 genes, 3,910 putative protein-coding genes (CDSs) were identified. Furthermore, 87 complete RNAs, including 10 rRNAs , 72 tRNAs, and 5 noncoding RNAs (ncRNAs) were found.Total genomic DNA (2\u2009\u03bcg) was isolated using a DN12 microbial DNA isolation kit  according to the manufacturer\u2019s instructions. A library for genome sequencing was constructed using the NEBNext Ultra DNA library prep kit for Illumina . The genBacillus sp. YSP-3 contains 1 gene cluster  for ectoine biosynthesis from aspartate semialdehyde, 1 glnA gene for l-glutamine biosynthesis from l-glutamate, 9 genes of glycine/betaine ABC transporter, 3 genes of Na+/solute symporter, 3 genes of Na+/alanine symporter, and 2 genes of Na+/proline symporter, indicating that Bacillus sp. YSP-3 may resist osmotic stress when salinity increases by taking up compatible solute  into the cell from the extracellular environment (+ uptake system also implies that strain YSP-3 possibly gains isosmotic cytoplasm though K+ as an osmolyte via a \"salt-in strategy\" when coping with a rapid osmotic shock (Bacillus sp. YSP-3 contains nine Na+/H+ antiporter genes (+ antiporter gene (+/H+ antiporter gene (Bacillus sp. YSP-3 survival in alkaline environments (+ homeostasis under salinity stress.Analysis of the genome sequence revealed that ironment , 7. The ic shock . Alkalipter gene , and a Kter gene , which mronments . These pBacillus sp. YSP-3 has been deposited at DDBJ/ENA/GenBank under the accession number PDOF00000000.The draft genome assembly of"}
+{"text": "Verticillium nonalfalfae, a soilborne vascular fungus, shows promise for biocontrol of highly invasive Ailanthus altissima strains. This announcement provides draft genome sequences of the aggressive isolate G1/5 (wild-type strain), the highly aggressive isolate Vert56 (improved strain), and the mildly aggressive isolate I3/2, all obtained from symptomatic A. altissima trees in Austria. Verticillium nonalfalfae, a soilborne vascular fungus, shows promise for biocontrol of highly invasive Ailanthus altissima strains. This announcement provides draft genome sequences of the aggressive isolate G1/5 (wild-type strain), the highly aggressive isolate Vert56 (improved strain), and the mildly aggressive isolate I3/2, all obtained from symptomatic A. altissima trees in Austria. Verticillium nonalfalfae is an important vascular wilt pathogen on several dicotyledonous crops as well as on tree of heaven   \u20133. This issima) \u2013\u20137. For telection  and resuKT223526 and WGLJ00000000) (WGLK00000000), and another less pathogenic isolate, I3/2 (GenBank accession numbers KT223527 and SPUV00000000)  with Milli-Q water for elution. Whole-genome sequencing was performed by Vienna BioCenter Core Facilities GmbH  using an Illumina HiSeq 2500 system with a single-end 100-bp setup. Sequencing libraries were prepared according to the Westburg next-generation sequencing (NGS) DNA library prep kit protocol v3.1 . A total of 60.09 million (Vert56), 54.24 million (I3/2), and 57.87 million (G1/5) reads were received, representing in total 6,008.9 million, 5,423.5 million, and 5,786.7 million bases, respectively. Default parameters were used for all software, unless otherwise noted. A read quality check was performed using FastQC v0.11.4) (.4 (11) we strain , and theV. nonalfalfae strains Vert56, I3/2, and G1/5, respectively.Sequencing completeness was estimated using BUSCO (v3.0.2)  based onWGLJ00000000 , SPUV00000000 , and WGLK00000000 . The versions described in this paper are the first versions, WGLJ01000000, SPUV0100000000, and WGLK01000000, respectively. Sequence reads were deposited under SRA project accession numbers SRR8271219, SRR8271218, and SRR8271217, respectively, and BioProject accession number PRJNA507541.These whole-genome shotgun projects have been deposited at DDBJ/EMBL/GenBank under the accession numbers"}
+{"text": "Dictyostelium discoideum forms a fruiting body consisting of spores and a multicellular stalk. Originally, the chlorinated alkylphenone differentiation-inducing factors (DIFs) -1 and -3 were isolated as stalk cell inducers in D. discoideum. Later, DIFs and their derivatives were shown to possess several biologic activities including antitumor and anti-Trypanosoma properties. In this study, we examined the antibacterial activities of approximately 30 DIF derivatives by using several bacterial species. Several of the DIF derivatives strongly suppressed the growth of the Gram-positive bacteria Staphylococcus aureus, Bacillus subtilis, and Enterococcus faecalis and Enterococcus faecium, at minimum inhibitory concentrations (MICs) in the sub-micromolar to low-micromolar range. In contrast, none of the DIF derivatives evaluated had any noteworthy effect on the growth of the Gram-negative bacterium Escherichia coli . Most importantly, several of the DIF derivatives strongly inhibited the growth of methicillin-resistant S. aureus and vancomycin-resistant E. faecalis and E. faecium. Transmission electron microscopy revealed that treatment with DIF derivatives led to the formation of distinct multilayered structures consisting of cell wall or plasma membrane in S. aureus. The present results suggest that DIF derivatives are good lead compounds for developing novel antimicrobials.At the end of its life cycle, the cellular slime mold  Dictyostelium discoideum has long been studied as an excellent model system in the fields of cell and developmental biology; at the end of its developmental process, this organism forms fruiting bodies, each consisting of spores and a multicellular stalk [D. discoideum [D. discoideum [The cellular slime mold ar stalk ,2,3. Difar stalk ,5 and moscoideum . DIF-3  [Trypanosoma drugs [In addition to these developmental roles, DIF-1, DIF-3, and several of their derivatives B,C exertomiasis) . Importaomiasis) ,26,27,28ma drugs . MoreoveStaphylococcus aureus, Bacillus subtilis, Enterococcus faecalis and, Enterococcus faecium. In addition, several of the DIF derivatives evaluated strongly inhibited the growth of methicillin-resistant S. aureus and vancomycin-resistant enterococci . Our results support the investigation of such DIF derivatives as candidate lead compounds for developing novel antimicrobials.In this study, we investigated the antibacterial activities of DIF derivatives in vitro and show that several of them exert strong antibacterial effects against Gram-positive bacteria, including S. aureus , methicillin-resistant S. aureus , vancomycin-susceptible E. faecalis , vancomycin-resistant E. faecalis , vancomycin-resistant E. faecium , and B. subtilis (ATCC6633) and the Gram-negative bacteria Escherichia coli (NIHJ) and Mycobacterium bovis  were used in this study. DIF derivatives (The Gram-positive bacteria methicillin-susceptible ivatives  were synS. aureus suspension (109 CFU/mL) in Mueller\u2013Hinton broth (MHB) (0.2% (w/v) beef extract, 1.75% (w/v) casein hydrolysate, 0.15% (w/v) starch) into 10 mL of MHB agar (MHB containing 1.7% (w/v) agar; final bacterial density, 107 CFU/mL) and spread this suspension in a plastic culture dish . Paper discs  impregnated with 2 \u00b5L of EtOH, DMSO, a 10 mM solution of the DIF or DIF derivative of interest in EtOH or DMSO (approx. 5.5\u20138 \u00b5g DIF/disc), or the purchased penicillin\u2013streptomycin solution (10 units penicillin and 10 \u00b5g streptomycin/disc) were placed on the bacterial agar plate. In addition, paper discs  containing minomycin (30 \u00b5g/disc), imipenem (10 \u00b5g/disc), gentamicin (10 \u00b5g/disc), levofloxacin (5 \u00b5g/disc), or clindamycin (2 \u00b5g/disc) were placed on a MHB bacterial agar plate as controls. The plates were incubated for 20 h at 37 \u00b0C, after which the bacterial growth-inhibition zones around each disc was noted.We mixed 0.1 mL of S. aureus in MHB (107 CFU/mL) was spread on an MHB agar plate , after which 2 \u00b5L of each 10 mM DIF solution was dropped directly on the agar surface. The plates were incubated for 20 h at 37 \u00b0C, after which the bacterial growth-inhibition zones around the discs were noted.In addition, a diffusion assay without paper discs was performed for comparison. Briefly, 10 mL of 5 CFU/mL; 0.1 mL/well) were incubated for 20 to 24 h at 37 \u00b0C in 96-well plates  in the presence of vehicle, various concentrations of serially diluted DIF derivatives, or known antibiotics; MIC was defined as the lowest concentration of the additives that inhibited visible bacterial growth.Gram-positive and Gram-negative bacteria in MHB (5 \u00d7 10M. bovis was grown in Hayflick modified PPLO (pleuropneumonia-like organisms) broth (0.5% (w/v) glucose, 5% (w/v) beef heart infusion, 2.5% (w/v) yeast extract, 1% (w/v) peptone, 0.5% (w/v) NaCl, 15% (v/v) heat-inactivated horse serum, 50 \u00b5g/mL of ampicillin) and used at concentrations of 104 to 107 CFU/mL for the minimum inhibitory concentration (MIC) assay as described; tiamulin  and enrofloxacin (Sigma-Aldrich) were used as positive-control antibiotics.S. aureus (strain ATCC29213) in MHB  was incubated for 1.5 h at 37 \u00b0C in 6-well plates (Corning) in the presence of 0.2% DMSO (vehicle) or 4 \u00b5M DIF derivative. After incubation, the bacteria were transferred to centrifuge tubes, collected by centrifugation , fixed overnight at 4 \u00b0C in 0.5 mL of 2.5% (v/v) glutaraldehyde in 100 mM phosphate buffer (pH 7.4) , and postfixed in 1% (w/v) OsO4 in 100 mM phosphate buffer. Fixed specimens were dehydrated through a graded series of ethanol and embedded in Epon812 . Ultrathin sections were cut by using an ultramicrotome  and stained with uranyl acetate and lead citrate. These sections were examined under a transmission electron microscope .Specimens for transmission electron microscopy were prepared according to standard procedures . BrieflyS. aureus (MSSA: strain 209P) (S. aureus (MRSA: strain MS29202)  from those of the known antibiotics that we used.We first examined the effects of DIF-1, DIF-3 A, and thin 209P) A and metMS29202) B by usinMS29202) A, a largMS29202) B, peniciMS29202) C. These B. subtilis (ATCC6633) and on the Gram-negative bacterium E. coli (NIHJ), and MIC values of DIF derivatives were determined   and enrofloxacin (0.7 \u00b5M).We next examined the effects of DIF derivatives on the growth of the Gram-positive bacteria MSSA (strain 209P), MRSA (MS29202), and termined . Several>100 \u00b5M) ; M. boviNote that the MIC values of DIF-3 against MSSA and MRSA were comparable to those of DIF-1 , even thvanB and vanA, respectively . Transmission electron microscopy revealed distinct multilayered structures consisting of cell wall and/or plasma membrane in the DIF-treated MSSA cells . This fiD. purpureum [S. aureus (strain IID671), MRSA (IID1677), E. faecalis (ATCC29212), and B. subtilis (ATCC6633) are 1.56 \u00b5g/mL (3.4 \u00b5M), 3.13 \u00b5g/mL (6.8 \u00b5M), 50 \u00b5g/mL (109 \u00b5M), and 0.78 \u00b5g/mL (1.7 \u00b5M), respectively [Polysphondylium filamentosum [The antibacterial substance AB0022A, a chlorinated dibenzofuran A, was isurpureum  and is sectively , and thumentosum , but theWhen we assessed the MIC values of these dichlorinated dibenzofurans in MRSA (strain MS29202), neither AB0022A nor Pf-2 had any noteworthy inhibitory effect  on the growth of this strain B. In conPenicillium notatum have been isolated from fungi and Actinobacteria [Most of the many antibiotics  identified since the discovery of penicillin in the fungus bacteria ,34,35,36bacteria ,35,36,37D. discoideum has long been used as a model organism in the study of cell and developmental biology. However, slime molds have recently been identified as excellent sources of potential lead compounds for drug discovery and the development of novel medicines [D. discoideum-derived DIFs and their derivative compounds in various types of eukaryotic cells [The cellular slime mold edicines ,38. In tic cells ,16,29.D. discoideum DIF derivatives possess antibacterial activity against Gram-positive bacteria, including S. aureus, E. faecalis, E. faecium, and B. subtilis. In contrast, these DIF derivatives did not inhibit the growth of Mycoplasma or the Gram-negative bacterium E. coli  and E. faecium (containing vanA) were resistant to both vancomycin and ampicillin (E. faecalis and E. faecium cause the great majority of enterococcal infections, and isolates that carry drug-resistance genes such as vanA or vanB can cause serious infections [In the present study, we showed that several  E. coli . Most im E. coli . Note th E. coli B but als E. coli C, wherea E. coli . In addipicillin . Althougfections ,40. OverAlthough the antibacterial activities of the DIF derivatives varied among the Gram-positive strains of bacteria examined in this study , the strS. aureus, Ph-DIF-1, Ph-DIF-3, and Bu-DIF-3 induce the formation of distinct multilayered structures composed of cell wall or membrane (Regarding the mechanism through which DIF derivatives exert their antibacterial effects, DIF derivatives have mitochondrial uncoupling activity in mammalian cells ,18. Givemembrane . AlthougD. discoideum possess strong antimicrobial activities against Gram-positive bacteria including MRSAs and VREs. Our results suggest that the DIF derivatives are good lead compounds for developing novel antimicrobials.In this study, we showed that several derivatives of DIF-1 and DIF-3, chlorinated polyketides found in The following authors hold a patent related to this article:Kubohara, Y.; Kikuchi, H.; Oshima, Y. Antibacterial drugs. Japanese Patent No. 6478378, 15 February 2019."}
+{"text": "Haematobia irritans irritans , a hematophagous external parasite of cattle, causes considerable economic losses to the livestock industry worldwide. This pest is mainly controlled with insecticides; however, horn fly populations from several countries have developed resistance to many of the products available for their control. In an attempt to better understand the adult horn fly and the development of resistance in natural populations, we used an Illumina paired-end read HiSeq and GAII approach to determine the transcriptomes of untreated control adult females, untreated control adult males, permethrin-treated surviving adult males and permethrin + piperonyl butoxide-treated killed adult males from a Louisiana population of horn flies with a moderate level of pyrethroid resistance. A total of 128,769,829, 127,276,458, 67,653,920, and 64,270,124 quality-filtered Illumina reads were obtained for untreated control adult females, untreated control adult males, permethrin-treated surviving adult males and permethrin + piperonyl butoxide-treated killed adult males, respectively. The de novo assemblies using CLC Genomics Workbench 8.0.1 yielded 15,699, 11,961, 2672, 7278 contigs (\u2265 200\u202fnt) for untreated control adult females, untreated control adult males, permethrin-treated surviving adult males and permethrin + piperonyl butoxide-treated killed adult males, respectively. More than 56% of the assembled contigs of each data set had significant hits in the BlastX (UniProtKB/Swiss-Prot database) (E <0.001). The number of contigs in each data set with InterProScan, GO mapping, Enzyme codes and KEGG pathway annotations were: Untreated Control Adult Females \u2013 10,331, 8770, 2963, 2183; Untreated control adult males \u2013 8392, 7056, 2449, 1765; Permethrin-treated surviving adult males \u2013 1992, 1609, 641, 495; Permethrin + PBO-treated killed adult males \u2013 5561, 4463, 1628, 1211.The horn fly,  Specifications TableValue of the data\u2022Haematobia irritans irritans.Resource for investigations of the molecular basis of insecticide resistance in the horn fly, \u2022Provides candidate protein coding regions for the development of control strategies targeting adult flies.1https://www.ncbi.nlm.nih.gov/sra/SRP131897 or through SRA accession number SRP131897. The adult horn fly transcriptome Shotgun Assembly project has been deposited at DDBJ/EMBL/GenBank under the accession GGLM00000000. The version described in this paper is the first version, GGLM01000000. The overall BioProject ID is PRJNA429442 and the BioSample accessions are SAMN08355023, SAMN08355024, SAMN08355025, and SAMN08355026.RNA was isolated from unfed, newly emerged adult horn flies, including untreated control adult females, untreated control adult males, permethrin-treated surviving adult males and permethrin + piperonyl butoxide-treated killed adult males. Subsequently, a single lane of 2 \u00d7 54\u202fbp paired end RNASeq reads were obtained, de novo assembled and annotated. The raw reads are accessible at NCBI\u05f3s SRA through the direct link 22.12, ~LD25) or permethrin  + 1% piperonyl butoxide (PBO) by the impregnated filter paper assay Adult flies were collected with aerial sweep hand nets from pastured cattle at the St. Gabriel Research Station, Saint Gabriel, Louisiana, and incubated in large inverted Erlenmeyer flasks to collect eggs that were immediately seeded into manure to allow adult fly emergence. The unfed, newly emerged flies were sexed and either immediately frozen at \u221280\u202f\u00b0C for sequencing  or exposed to low doses of permethrin  and the FastRNA Pro Green Kit .2.3https://www.qiagenbioinformatics.com/) or Trimmomatic programmable-0.33 https://de.cyverse.org/de/?type=apps&app-id=8cb5c088-6b3e-11e7-a22d-008cfa5ae621&system-id=de)  followed by Sickle-quality-based-trimming_version_1.0\u00a0\u00a0https://de.cyverse.org/de/?type=apps&app-id=9f5710c6-3424-11e7-9a58-008cfa5ae621&system-id=de)  and Illumina adaptor sequences and low quality bases were removed. Trimmomatic and Sickle were both run on CyVerse/Discovery Environment https://de.cyverse.org/de/?type=apps&app-id=trinity-wrangler-2.5.1u2&system-id=agave) or version 11.10.13 (https://de.cyverse.org/de/?type=apps&app-id=trinity-stmpde-11.10.13u2&system-id=agave) and SoapdenovoTrans version 1.0.3 https://de.cyverse.org/de/?type=apps&app-id=Soaptrans-1.0.3u1&system-id=agave). Both Trinity versions and SoapdenovoTrans were run on CyVerse/Discovery Environment Sequencing was performed at the National Center for Genome Research  using the standard Illumina RNAseq library preparation protocol and a single lane of the RNAseq. 2 \u00d7 54 paired-end approach. A total of 134,671,818, 132,374,494, 68,856,572, 65,427,160 paired-end Illumina raw reads were produced for untreated control adult females, untreated control adult males, permethrin-treated surviving adult males and permethrin + PBO-treated killed adult males, respectively . The rawhttps://pods.iplantcollaborative.org/wiki/display/DEapps/Compute+Contig+Statistics), rnaQUAST_1.2.0 (de novo based) https://de.cyverse.org/de/?type=apps&app-id=980dd11a-1666-11e6-9122-930ba8f23352&system-id=de) and BUSCO-v3.0\u00a0\u00a0https://de.cyverse.org/de/?type=apps&app-id=7f948668-7a53-11e7-a680-008cfa5ae621&system-id=de) (The assembled transcriptomes were then compared using three tools on CyVerse/Discovery Environment m-id=de) . Assemblm-id=de)  and were-3) using Blast2GO PRO version 5.0.21 The transcriptomes were BlastX aligned against the UniProtKB/SwissProt database (E-value = 1.0 e"}
+{"text": "AbstractSinocentrus Yuan, S.brevicornis Li & Chen, sp. nov. from China, is described and illustrated. A checklist and key to species of the Sinocentrus are provided.A new species of the treehopper genus  Sinocentrus was established by Yuan , 1 r-m and 1 m-cu crossveins present, apical limbus broad, with wrinkles. Metathoracic trochanter without spines, tibia with 3 rows of cucullate setae.Large sized. Frontoclypeus distinct. Suprahumeral horns narrow, acuminate, horizontally extended laterally, width between their apex ca 0.5 to 1.0 times body length. Pronotum highly developed, with anterior part strongly inflated and evenly rounded in profile, metopidium vertical, glabrous and minutely punctate without obvious pubescence. Posterior pronotal process strongly elevated above scutellum at base, slender, elongate, evenly tapered toward apex, straight or slightly sinuate, lateral and dorsal carina well developed, apex extended beyond forewing clavus. Scutellum entirely exposed, posterior margin emarginate. Basal one-fifth of forewing with opaque sclerotization, veins M and Cu fused basally to approximately one-fifth to one-fourth of wing length then strongly divergent, veins M+Cu and R fused basally, with 1 m-cu, 2 r-m, and 1 s crossveins. Hindwing vein R branched into RCentrotinae genera by the following characters: pronotum highly developed, strongly inflated with anterior part evenly rounded, glabrous with minute punctures and no obvious pubescence, suprahumeral horns extended laterad, posterior pronotal process elevated far above scutellum, scutellum emarginate.This genus can be distinguished from other oriental China  Fig. .S.brevicornis Li & Chen, sp. nov.; China (Hainan)S.sinensis Yuan, 2002; China (Yunnan); elevation: 1600 m.Taxon classificationAnimaliaHemipteraMembracidaeLi & Chensp. nov.0F416316-4D26-546E-8578-BDD13EA0550Dhttp://zoobank.org/73BE02FB-C98E-4D45-AC88-D597817F3597Holotype: \u2642, CHINA: Hainan, Bawangling, 29 April 2017, Hong-Xing Li. Paratypes: 2\u2640\u2640, same data as holotype.N = 1), female 8.9\u20139.3 mm (N = 2); forewing length: male 7.3 mm (N = 1), female 7.3\u20137.9 mm (N = 2); width between humeral angles apices: male 3.3 mm (N = 1), female 3.5\u20133.8 mm (N = 2); width between suprahumeral horns apices: male 4.6 mm (N = 1), female 4.6\u20135.2 mm (N = 2).Body length: male 8.1 mm . Humeral angles triangular with apices somewhat blunt. Suprahumeral horns short, width between horns apices nearly half length of body. Scutellum humped basally, large punctures present, longer than wide, apex extended antero-dorsally male, curved ventrally in female .brevicornis\u201d is derived from the Latins words \u201cbrevi-\u201d and \u201ccornu\u201d, referring to having the short suprahumeral horns.The word \u201cS.sinensis Yuan, 2002, but differs from the latter in: (1) forewing veins black and apical limbus black  (2) suprahumeral horns short, width between suprahumeral horns apices shorter than body length (as long as body length in S.sinensis); (3) posterior pronotal process nearly straightly ; (4) scutellum longer than wide (wider than long in S.sinensis); (5) apex of posterior pronotal process not reaching M3+4 veins (exceeding M3+4 veins in S.sinensis).This species is similar to Taxon classificationAnimaliaHemipteraMembracidaeYuan, 2002074481DF-471C-5038-857A-48AFE38F7BD3Sinocentrussinensis Yuan, 2002: 170, by original designation.Coloration. General color reddish-brown with golden setae .While holotype was not examined, an online image of the holotype Fig.  and detaCentrotinae, Sinocentrus and treated the genus as Centrotinae, incertae sedis.In their phylogeny and genus-level revision of Sinocentrus: (1) frontoclypeus distinct (indistinct in Centrotypini); (2) posterior pronotal process elevated far above the scutellum, entirely exposed ; (3) male lateral plate with short dorsoapical lobe extending dorsally, style clasp angled ventrally; style shank with arch at central section ; (4) mesothoracic femur without ablateral and adlateral cucullate setae; metathoracic leg cucullate setae row II irregular. Although the above characteristics can suggest that the genus is related to Leptocentrini, the shape of the female second valvulae closely align S.brevicornis with the tribe Centrotypini. Given these mixed affinities, we follow Wallace and Deitz, in treating Sinocentrus as Centrotinae, incertae sedis. Proper tribal placement may be affirmed by future phylogenetic analyses of combined morphological and molecular data.We provide the following additional details on"}
+{"text": "Lower physical activity is cross-sectionally associated with greater fatigability; whether such a relationship holds for longitudinal changes in fatigability is under-studied. We examined this question in offspring  enrolled in the Long Life Family Study, a two-generation cohort enriched for exceptional longevity and their spousal controls. At Visit 2 (2014-2017), we measured self-reported physical activity (PA) with the Framingham Physical Activity Index . Perceived physical fatigability was assessed using the Pittsburgh Fatigability Scale  at Visit 2 and repeated during a follow-up contact 2.7\u00b10.92 years later. We constructed a repeated-measures linear mixed-effect model to examine the effect of PA on longitudinal change in PFS by median age  adjusted for family structure, field center, follow-up time, sex, and self-rated health. We found a strong dose-response relationship of PFS scores across the four age/PA groups (ptrend<0.001). Specifically, older/less active (N=310) participants had the highest annual PFS increases of 0.37 points/yr (p<0.001) while those older/more active (N=340) had annual increases of 0.17 points/yr (p=0.03). Younger/less active (N=371) participants had annual PFS increases of 0.09 points/yr (p=0.008); those younger/more active (N=341) had annual decreases (improvement) of 0.18 points/yr (p<0.001). Although annual PFS changes were modest, our findings indicate physical activity attenuated age differences in these trajectories. Physical activity is emerging as a potential target for intervention aimed at reducing fatigability - an important risk factor in the disability pathway."}
+{"text": "Their structures were elucidated on the basis of extensive 1D and 2D NMR spectroscopic data analyses. Significant antifeedant activity of 1, 3 and 10 against the beet armyworm (Spodoptera exigua) was found, indicating that these diterpenoids might also be involved in the plant defense against insect herbivores.Three new grayanane diterpenoids, pierisoids C\u2012E ( Pieris, Rhododendron, Kalmia, Craibiodendron and Leucothoe +; HR-ESI-MS: m/z 507.2567 [M + Na]+ ; 1H- and 13C-NMR data: see Pierisoid D (2): Colorless oils;c = 0.1, MeOH); UV (MeOH) \u03bbmax (log \u03b5): 202 (2.48) nm; IR (KBr) \u03bdmax: 3441, 3432, 2971, 2939, 1735, 1730, 1630, 1382, 1179, 1049, 1025 cm\u22121; ESI-MS: m/z 463 (88) [M + Na]+; HR-EI-MS: m/z 440.2420 ; 1H- and 13C-NMR data: see Pierisoid E (3): Colorless crystals; mp 271\u2013272 \u00b0C;c = 0.1, MeOH); UV (MeOH) \u03bbmax (log \u03b5): 202 (2.29) nm; IR (KBr) \u03bdmax: 3441, 2942, 1787, 1734, 1631, 1372, 1264, 1237, 1053 cm-1; ESI-MS: m/z 661 (68) [M + Na]+; HR-EI-MS: m/z 638.2560 ; 1H- and 13C-NMR data: see Spodoptera exigua) were purchased from the Pilot-Scale Base of Bio-Pesticides, Institute of Zoology, Chinese Academy of Sciences. A modified dual-choice bioassay was performed for an antifeedant test as previously described  \u00d7 100, where C and T represent the control and treated leaf areas consumed by the insect. The insect antifeedant potency of the test compound was evaluated in terms of the EC50 value, which was determined by probit analysis for each insect species.Beet armyworms , as well as 10 known ones (4\u201213), were identified from the flowers of P. formosa via their extensive 1D and 2D NMR spectroscopic data analyses. Notably, compounds 1, 3 and 10, especially 10, exhibited obvious antifeedant activity against the beet armyworm (S. exigua), suggesting that these diterpenoids were important defensive substances in P. formosa against natural enemies.Terpenoids play an important role in natural product chemistry and biology, such as antifungal and insecticidal activities . Seconda"}
+{"text": "Drosophila Hippo pathway maintains NSC quiescence, but its regulation during brain development remains unknown. Here, we show that CRL4Mahj, an evolutionarily conserved E3 ubiquitin ligase, is essential for NSC reactivation (exit from quiescence). We demonstrate that damaged DNA-binding protein 1 (DDB1) and Cullin4, two core components of Cullin4-RING ligase (CRL4), are intrinsically required for NSC reactivation. We have identified a substrate receptor of CRL4, Mahjong (Mahj), which is necessary and sufficient for NSC reactivation. Moreover, we show that CRL4Mahj forms a protein complex with Warts (Wts/large tumor suppressor [Lats]), a kinase of the Hippo signaling pathway, and Mahj promotes the ubiquitination of Wts. Our genetic analyses further support the conclusion that CRL4Mahj triggers NSC reactivation by inhibition of Wts. Given that Cullin4B mutations cause mental retardation and cerebral malformation, similar regulatory mechanisms may be applied to the human brain.The ability of neural stem cells (NSCs) to transit between quiescence and proliferation is crucial for brain development and homeostasis.  During the transition from quiescence to reactivation of neural stem cells, the E3 ubiquitin ligase CRL4Mahj promotes their reactivation by inhibiting Wts, a core kinase of Hippo signalling pathway. The ability of stem cells to switch between quiescence and proliferation is crucial for tissue homeostasis and regeneration. Most adult neural stem cells (NSCs) exist in quiescence, a reversible dormant state, without proliferation or differentiation . In respDrosophila NSCs, called neuroblasts, have emerged as a powerful model to study mechanisms underlying NSC reactivation in vivo. Most Drosophila NSCs enter quiescence at the end of embryogenesis and resume proliferation shortly after larval hatching (ALH) #44974) driven by insc-Gal4 at 24 h ALH . Similarly, 31.2%  and 26.3%  of NSCs extended a cellular process in cul4G1-3 and cul4JJ11 mutants, compared with only 6.1% of NSCs  of WT brain lobes . Moreover, at 24 h ALH upon knockdown of cul4 by two different RNAi lines driven by NSC-driver insc-Gal4, significantly more quiescent NSCs, as judged by the lack of EdU incorporation and by the presence of cellular processes, were observed  versus 12.4%  and 10.3%  PH3+ NSCs were observed in cul4RNAi1/VDRC#105668 and cul4RNAi2/VDRC#44829  and ddb1HK-2-3 , ddb15-1 , and cul4G1-3 mutant brains  were proliferating at 24 h ALH on food depleted of amino acids, as indicated by their ability to incorporate EdU , overexpression of Cul4KR resulted in a marked decrease of EdU+ NSCs to 57.2%   in control MARCM clones were EdU+, only 7.5% (n = 53) and 19.5% (n = 41) of Mira+ NSCs in cul4G1-3 and cul4JJ11 MARCM clones were EdU+ (cul4G1-3 and cul4JJ11 was completely rescued by overexpression of UAS-cul4 transgene (n = 105 and n = 69). By contrast, overexpression of UAS-Cul4KR failed to rescue the quiescent NSC phenotypes of cul4G1-3 and cul4JJ11, as indicated by 3.7% (n = 27) and 3.2% (n = 31) EdU+ NSCs, respectively  . Further control . These p mutants . At 96 here EdU+ . The quiectively . These r+ NSCs , whereas overexpression of an active form of InR (InRCA) in NSCs triggered NSC reactivation . By contrast, ddb1 knockdown dramatically suppressed this effect induced by InRCA overexpression . This result suggested that DDB1 likely functions downstream of the InR pathway during NSC reactivation.We next tested whether DDB1 is required for premature reactivation induced by overactivation of InR signaling pathway. Similar to previous reports ,11, in tDrosophila ortholog of human DCAF1/Vpr (HIV-1) Binding Protein (VprBP) #34912 and mahjRNAi2: VDRC#110669) driven by insc-Gal4 led to an increase in proportion of NSCs with a cellular process to 23.1%  and 27.1%   and 75.1%  upon mahj RNAi knockdown, whereas there were 96.6%  EdU+ NSCs in the control  in mahjRNAi1 and 9.1%  in mahjRNAi2, compared with 20.6%  in control  , as the t = 575) . Consist control . In addi control .mahj1, that was previously generated by imprecise P-element excision , depending on the combinations of constructs) displayed no PLA foci, and the rest of the cells displayed weak PLA foci (1\u201310 foci); on average, there were 0\u20130.4 \u00b1 0.2\u20130.8 PLA foci per cell (n = 281) coexpressing Flag-Wts and Myc-Mahj showed PLA signals: 26.0%, 30.6%, and 41.3% of cell with strong (>20 foci), moderate (11\u201320 foci), and weak (1\u201310 foci) signals, respectively; they resulted in 16.2 \u00b1 14.9 foci per cell (n = 176) expressing Flag-Wts and Myc-DDB1 (n = 176) showed PLA signals: 10.1%, 27.0%, and 60.7% of the cells with strong, moderate, and weak PLA signals; and on average, these cells displayed 10.7 \u00b1 9.94 PLA foci per cell (n = 157) also indicated positive physical interaction: 11.9 \u00b1 10.5 PLA foci per cell and 97.5% cells showed strong, moderate, and weak PLA signals of 56.7%, 26.1%, and 14.6%, respectively   . By contn of Wts . These rS2 cells .mahj depletion could aggravate NSC reactivation defects observed upon ddb1 depletion. Whereas 97.2% of control NSCs  were positive for EdU, 84.7%  of ddb1RNAi and 81.5%  of mahjRNAi controls incorporated EdU  of NSCs were EdU+  of NSCs were positive for EdU, comparable to that of the control brains   of NSCs in mahj knockdown were EdU+ at 24 h ALH, upon simultaneous knockdown of both mahj and wts by insc-Gal4, 97.1%  of NSCs incorporated EdU . These data suggest that wts knockdown significantly suppresses NSC reactivation defects observed in mahj RNAi brains. To maintain NSC quiescence, Wts phosphorylates and inactivates the transcriptional coactivator Yki to turn off expressions from Yki target genes  XE-2900) transheterozygous mutants were labeled with EdU, Dpn, and Mira. (M-N) Quantification of NSCs that are EdU+ or with process of various genotypes in (L). (O) MB NSC lineages in larval brains from WT and mahj1 at 24 h ALH were labeled with EdU, Dpn, and Dac. The enlarged views of white dotted boxes are shown to the right. Yellow arrowheads, MB NSCs surrounded by Dac-positive MB neurons. (P) At 0 h ALH, larval brains of grh-gal4>UAS-CD8-GFP were labeled with Mahj and a NSC marker (Dpn). Blue arrows, NSCs. (Q) Quantification of number of NSCs in ddb1\u2212, cul4\u2212, and mahj\u2212 mutant brains at 24 h ALH. Data are presented as mean \u00b1 SD. **** for P \u2264 0.0001, *** for P \u2264 0.001, ** for P \u2264 0.01, * for P \u2264 0.05, and ns for P > 0.05. Scale bars: 10 \u03bcm. The data underlying this figure can be found in Drosophila Stock Center; cul4, Cullin 4; Dac, dachshund; Dcr2, Dicer 2; ddb1, damaged DNA-binding protein 1; Dpn, Deadpan; EdU, 5-ethynyl-2\u2032-deoxyuridine; GFP, green fluorescent protein; Mahj, Mahjong; MB, mushroom body; Mira, Miranda; ns, statistically nonsignificant; NSC, neural stem cell; PH3, phospho-Histone H3; ROD, relative of density; UAS, upstream activating sequence; VDRC, Vienna Drosophila Resource Center; WT, wild type.(A-F) At 24 h ALH, larval NSCs in control ((TIF)Click here for additional data file.S4 FigDmMahj and DmDDB1. (A) View of Mahj 1110\u20131441 aa  and DDB1  heterodimers. (B) Detail of predicted interaction between residues R1120 and R1123 (red stick) side chains in the HLH motif of Mahj (yellow ribbon) and DDB1 . Relevant residues are depicted in stick models: R1220 and R1123 , D1134 , and D788 . The predicted H-bonds are depicted by black dashed lines. Mahj R1120 and R1123 are corresponding to HsDCAF1 R1053 and R1056, respectively. (C) Detail of predicted interaction between R1307 and R1343 (red stick) side chains in the WD40 domain of Mahj (yellow ribbon) and DDB1 . Relevant residues are depicted in stick models: R1307 and R1342 , E1295 , and E201 . The predicted H-bonds are depicted by black dashed lines. Mahj R1307 and R1343 are corresponding to HsDCAF1 R1247, R1283, respectively. (D) A schematic diagram illustrating different Wts domains and truncated Wts proteins. (E) Wts C-terminal fragment containing kinase domain interacts with Mahj. Co-IP between Myc-Mahj and Flag-Wts or indicated truncated constructs. Anti-Myc were used for IP, followed by western blotting probed with anti-Myc, anti-Flag, or anti-Actin antibodies. Actin served as a loading control. aa, amino acid; DDB1, damaged DNA-binding protein 1; Dm, D. melanogaster; HLH, helix-loop-helix; HsDCAF1, H. sapiens DDB1-Cul4 associated factor 1; IP, immunoprecipitation; Mahj, Mahjong; Wts, Warts.(A-C) A homology model illustrating the interaction between (TIF)Click here for additional data file.S5 Fign = 272), Flag-Wts + Myc-control (n = 247), Flag-control + Myc-Mahj (n = 889), Flag-control + Myc-DDB1 (n = 284), Flag-control + Myc-Cul4 (n = 263), Flag-Wts + Myc-Mahj (n = 281), Flag-Wts + Myc-DDB1 (n = 176), and Flag-Wts + Myc-Cul4 (n = 157). The data underlying this figure can be found in (A) Co-IP between Flag-DDB1 and HA-Wts. S2 cells were cotransfected with Flag-DDB1 and HA-Wts or respective controls. Immunoprecipitation was performed using anti-Flag antibodies, and western blot was performed using anti-Flag or anti-HA antibodies. (B) A schematic illustration for PLA. (C) In situ PLA assay between Flag-Wts and either Myc-Mahj, Myc-DDB1, or Myc-Cul4. S2 cells transfected with the indicated plasmids were stained with Flag, Myc, and DAPI and detected for PLA signal (red). Cell outline was shown by DIC images. Scale bar, 10 \u03bcm. (D) Quantification graph showing the percentage of cells with PLA foci in (B). (E) Quantification for the average number of PLA foci per cell in (C). The number of cells used for quantification in  are coexpression of Flag-control + Myc-control ((TIF)Click here for additional data file.S6 FigctrlRNAi: \u03b2-galRNAi), wtsRNAi +\u03b2-galRNAi, ddb1RNAi (VDRC#44974) +\u03b2-galRNAi, and ddb1RNAi + wtsRNAi were labeled with Dpn, Mira, and EdU. (B-C) Quantification of Dpn+ Mira+ NSCs that are EdU+ (B) or with cellular process (C). In (B), control : brain lobes n = 8, total NSCs t = 517; wtsRNAi+\u03b2-galRNAi: n = 11, t = 705; ddb1RNAi+\u03b2-galRNAi: n = 10, t = 647; ddb1RNAi + wtsRNAi: n = 15, t = 955. In (C), control : n = 4, t = 320; wtsRNAi+\u03b2-galRNAi: n = 6, t = 485; ddb1RNAi+\u03b2-galRNAi: n = 6, t = 598; ddb1RNAi + wtsRNAi: n = 16, t = 1,403. (D) Larval NSCs in ctrlRNAi , YkiS168A+\u03b2-galRNAi, ddb1RNAi+ \u03b2-galRNAi, and ddb1RNAi + YkiS168A were labeled with Dpn, Mira, and EdU. (E-F) Quantification of Dpn+ Mira+ NSCs that are EdU+ (E) or with cellular process (F). In (E), control : n = 7, t = 388; YkiS168A+\u03b2-galRNAi: n = 10, t = 645; ddb1RNAi+\u03b2-galRNAi: n = 9, t = 551; ddb1RNAi + YkiS168A: n = 11, t = 809. In (F), control : n = 5, t = 429; YkiS168A+\u03b2-galRNAi: n = 4, t = 255; ddb1RNAi+\u03b2-galRNAi: n = 5, t = 357; ddb1RNAi + YkiS168A: n = 9, t = 755. (G) Larval NSCs in ctrlRNAi , wtsRNAi+\u03b2-galRNAi, cul4RNAi (VDRC#105668)+\u03b2-galRNAi, and cul4RNAi + wtsRNAi were labeled with Dpn and EdU. (H-I) Quantification of Dpn+ NSCs that are EdU+ or with Mira+ cellular process. In (H), control : n = 13, t = 1,179; wtsRNAi+\u03b2-galRNAi: n = 19, t = 1,665; cul4RNAi+\u03b2-galRNAi: n = 14, t = 1,073; cul4RNAi + wtsRNAi: n = 19, t = 1,404. In (I), control : n = 8, t = 702; wtsRNAi+\u03b2-galRNAi: n = 8, t = 696; cul4RNAi+\u03b2-galRNAi: n = 9, t = 772; cul4RNAi + wtsRNAi: n = 10, t = 847. (J) Larval NSCs in ctrlRNAi+\u03b2-galRNAi, YkiS168A+\u03b2-galRNAi, cul4RNAi+\u03b2-galRNAi, and cul4RNAi + YkiS168A were labeled with Dpn and EdU. (K-L) Quantification of Dpn+ NSCs that are EdU+ or with Mira+ cellular process. In (K), control : n = 12, t = 990; YkiS168A+\u03b2-galRNAi: n = 11, t = 917; cul4RNAi+\u03b2-galRNAi: n = 13, t = 1,056; cul4RNAi + YkiS168A: n = 15, t = 950. In (L), control : n = 7, t = 701; YkiS168A+\u03b2-galRNAi: n = 4, t = 344; cul4RNAi+\u03b2-galRNAi: n = 11, t = 974; cul4RNAi + YkiS168A: n = 10, t = 895. Driver is insc-Gal4, tub-Gal80ts (A-F), and insc-Gal4; UAS-Dcr2 (G-L). Note that to balance the number of UAS elements across different genotypes, additional control UAS line, \u03b2-galRNAi for (A-F) or CD8-GFP/BDSC#32186 for (G-L), was added to various RNAi lines, resulting in weaker phenotype in RNAi lines compared with those without additional UAS control shown earlier in this study. Yellow arrows, EdU+ NSCs. White arrows, EdU-negative NSCs. Data are presented as mean \u00b1 SD. **** for P \u2264 0.0001, *** for P \u2264 0.001, ** for P \u2264 0.01, * for P \u2264 0.05, and ns for P > 0.05. Scale bars, 10 \u03bcm. The data underlying this figure can be found in Drosophila Stock Center; CRL4, Cullin4-RING ligase; ddb1, damaged DNA-binding protein 1; Dpn, Deadpan; EdU, 5-ethynyl-2\u2032-deoxyuridine; Mira, Miranda; ns, statistically nonsignificant; NSC, neural stem cell; RNAi, RNA interference; UAS, upstream activating sequence; VDRC, Vienna Drosophila Resource Center; Wts, Warts; Yki, Yorkie.(A) Larval NSCs in control ((TIF)Click here for additional data file.S1 TableC-term, C-terminal fragment; F, forward; FL, full length; HLH+WD40, helix-loop-helix motif and WD40 domain; N-term, N-terminal fragment; R, reverse; sc, with stop codon; wo sc, without stop codon.(XLSX)Click here for additional data file.S2 TableF, forward; R, reverse.(XLSX)Click here for additional data file.S3 TableDDB1, damaged DNA-binding protein 1; F, forward; R, reverse; RNAi, RNA interference; UAS, upstream activating sequence.(XLSX)Click here for additional data file.S1 Text(DOCX)Click here for additional data file.S1 Data(XLSX)Click here for additional data file."}
+{"text": "To investigate the diagnostic value of the interferon-\u03b3 release assay (IGRA) for detecting tuberculosis (TB) infection in patients with Beh\u00e7et\u2019s disease (BD).We retrospective analyzed the data collected from 173 BD patients hospitalized between 2010 and 2015. Ninety-nine healthy volunteers were enrolled as a control group. IGRA was performed using T-SPOT.TB. The diagnosis of active TB (ATB) was based on clinical, radiological, microbiological, histopathological information and the response to anti-TB therapy. Latent TB (LTB) infection was defined as asymptomatic patients with positive T-SPOT.TB.6 PBMC was the optimal cutoff for diagnosing ATB, with an area under the curve of 0.891. Furthermore, the median SFCs in ATB group was significantly higher than those in LTB infection  or previous TB infection . A significant discrepancy between T-SPOT.TB and tuberculin skin test was noted .TB infection was documented in 59 BD patients (34.1%). The sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio and negative likelihood ratio of T-SPOT.TB for the diagnosis of ATB were 88.9%, 74.8%, 29.1%, 98.3%, 3.53 and 0.15, respectively. The receiver-operating-characteristic curve demonstrated that spot-forming cells (SFCs) of 70/10T-SPOT.TB, an IGRA, may assist in the diagnosis of ATB in BD patients, and the higher SFCs suggest ATB in BD patients. M.tuberculosis may act as a trigger of BD through molecular mimicking. On the other hand, defective cell-mediated immunity in BD patients may increase the susceptibility of TB [Behcet\u2019s disease (BD) is a multi-system inflammatory disorder, characterized by recurrent oral ulcers, genital ulcers, skin lesions, and ocular involvement of unknown etiology. Notably, the clinical manifestation of orogenital ulcers and skin lesions including erythema nodosum could be mimicked by tuberculosis (TB). Additionally, active TB (ATB) is frequently observed in patients with BD, and those symptoms were relieved after anti-TB therapy , 2. BD aty of TB , and thety of TB . Therefo. Given IGRAs do not cross-react with BCG vaccination or nontuberculous mycobacteria infection, IGRAs show higher specificity than TST [Tuberculin skin test (TST), a routine screening test for latent TB (LTB) infection, has limited diagnostic value in BD patients due to cross-reactivity with the bacillus Calmette-Gu\u00e9rin (BCG) vaccine and nontuberculous mycobacteria as well as pathergy reaction . In addithan TST . IGRAs, than TST  and rheuthan TST . HoweverM.tuberculosis. Highly-probable TB was defined as highly suggestive clinical and radiological features for TB in combination with the appropriate response to anti-TB treatment [Medical records of the hospitalized BD patients from Peking Union Medical College Hospital between January 2010 and March 2015 were retrospectively reviewed. All patients fulfilled the International Study Group BD criteria or the new International Criteria for BD . T-SPOT.reatment . LTB infreatment .M.tuberculosis. Briefly, peripheral blood mononuclear cells were isolated from whole blood samples by density gradient separation, then stimulated with two TB specific antigens, early secreted antigenic target 6 (ESAT-6) and culture filtrate protein 10 (CFP-10), phytohaemagglutinin (PHA) (positive control) or AIM-V  (negative control) in microplate wells (2.5\u2009\u00d7\u2009105 per well) precoated with interferon-\u03b3 (IFN-\u03b3) capture antibodies. The number of spot-forming cells (SFCs) was calculated after 16 to 20\u2009h of incubation. The result was considered positive if the number of SFCs was six or more after subtracting the spot count of negative control [IGRAs were performed using T-SPOT.TB, a simplified enzyme-linked immunospot (ELISPOT) for detection of effector T cells that respond to stimulation by antigens specific for  control .TST was performed with an intradermal injection using standardized 0.1\u2009ml (5\u2009U)-purified protein derivative into the ventral surface of the forearm according to the Mantoux method . On the p values <\u20090.05 is considered statistically significant. Statistical analysis was performed by using SPSS  and the GraphPad Prism .Numerical data were expressed as mean\u2009\u00b1\u2009standard deviation or median , and categorical data were expressed as frequencies/ percentages. The Mann-Whitney U test and Pearson chi-square test were used to compare differences between two groups. The concordance of TST and T-SPOT.TB results were assessed using \u03baappa coefficient. . The diagnostic performance of T-SPOT.TB on ATB was evaluated by calculation of its sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), positive likelihood ratio (PLR), and negative likelihood ratio (NLR). The area under the receiver operating characteristic curve of the T-SPOT.TB on peripheral blood mononuclear cell (PBMC) diagnostic cutoff for ATB was calculated. A two-sided with Among the 173 BD patients performed with TB-SPOT.TB test, 114 (65.9%) were men. The mean age was 37.07\u2009\u00b1\u200914.74\u2009years, and median disease duration of BD was 84\u2009months (range 1\u2013608). The most common clinical manifestation was recurrent oral ulceration (98.3%). Other common findings included fever (81.5%), genital ulcers (58.4%), skin lesions (58.4%), gastrointestinal involvement (39.3%), vascular involvement (28.3%), positive pathergy test (22.5%), ocular involvement (19%), neurologic involvement (15%), cardiac involvement (9.8%) and hematological involvement (5.2%). The median BDCAF2006 score for disease activity was 2 (range 0\u20135). Elevated erythrocyte sedimentation rate (ESR) and high-sensitivity C-reactive protein (hs-CRP) levels were detected in 98 (56.6%) and 118 patients (68.2%), respectively. The median ESR was 21 (range 1\u2013140) mm/first hour and the median hs-CRP was 11.54 (range 0.12\u2013262.77) mg/L. In the past 2 years, 113 patients (65.3%) were treated with glucocorticoids, including 13 patients (7.5%) received glucocorticoid pulse therapy. Ninety-seven patients (56.1%) were under glucocorticoids treatment when T-SPOT.TB assay was performed, and the median daily dose of corticosteroids was 40 (range 5\u2013100) mg. Ninety-four patients received immunosuppressants, including cyclophosphamide (30.1%), leflunomide (5.2%), cyclosporin A (4.6%), azathioprine (4.6%), methotrexate (4.6%) and salazosulfapyridine (2.9%). Sixteen patients (9.2%) received two or more immunosuppressants. Biological agents were used in 12 patients (6.9%), including infliximab injection in six patients, etanercept injection in five patients, etanercept followed by infliximab in one patient , especially erythema nodosa  compare to BD-LTB infection patients.TB infection was documented in 59 BD patients 34.1%) in our study, including 18 (10.4%) ATB (BD-ATB), 29 (16.8%) LTB (BD-LTB) infection, and 12 (6.9%) patients with evidence of previous TB. Of the 18 BD-ATB cases, culture-confirmed TB and highly probable TB were diagnosed in 4 and 14 patients, respectively. Of the four culture-confirmed TB patients, pulmonary TB, tuberculous lymphadenitis, tuberculous pleuritis, tuberculous meningitis were diagnosed in one patient, respectively. Among 14 highly-probable TB, highly-probable pulmonary TB was diagnosed in 7 cases based on fever, cough, expectoration or chest pain, and CT-confirmed pulmonary cavity, infiltrating or nodules as well as % in our 6 PBMC (IQR: 108\u2013925). Among patients with evidence of previous TB, ten patients (83.3%) had positive T-SPOT.TB with the median number of SFCs being 96/106 PBMC (IQR: 31.75\u2013187). The median number of SFCs in BD-LTB infection patients was 68/106 PBMC (IQR: 38\u2013208). The median number of SFCs in the BD-ATB group was higher than those in the BD-LTB infection group (p\u2009=\u20090.007) or those in previous TB patients (p\u2009=\u20090.018). The median numbers of SFCs in the BD-ATB group, BD-LTB infection group and previous TB patients were higher than those in the non-TB infection group (p\u2009=\u20090.000) , and the median number of spot-forming cells (SFCs) was 466/10The sensitivity, specificity, PPV and NPV of the T-SPOT.TB test for the diagnosis of ATB were 88.9% (95%CI 63.9\u201398.1%), 74.8% (95%CI 67.1\u201381.3%), 29.1% (95%CI 18\u201343.1%), and 98.3% (95%CI 93.4\u201399.7%), respectively. PLR was 3.53(95%CI 2.57\u20134.85) and NLR was 0.15 (95%CI 0.04\u20130.55).6 PBMC was the optimal cutoff for diagnosing BD-ATB, with an area under the curve of 0.891 (95%CI 0.796\u20130.987)  Fig.\u00a0. Accordi6 PBMC (IQR:80\u2013308). There was no significant difference in the proportion of LTB infection  and the number of SFCs detected by T-SPOT.TB  between BD patients and healthy controls.LTB infection was documented in 27(27.3%) healthy controls, and the median number of SFCs was 192/10p\u2009=\u20090.002).Both T-SPOT.TB and TST results were simultaneously tested in 63 patients. Among these patients, positive results of TST and T-SPOT.TB were presented in 44.4% 28/63) and 46% (29/63) patients, respectively. 30.2% (19/63) of the 63 patients had both positive TST and T-SPOT.TB, and 60.3% (38/63) had positive results on either TST or T-SPOT.TB (Table\u00a08/63 and The sensitivity of TST for detecting ATB was 78.6% (95%CI 48.8\u201394.3%), which was lower than that of the T-SPOT.TB test (92.9%), but there was no significant difference between the two groups. The specificity of TST was similar with T-SPOT.TB test (Table\u00a0p\u2009=\u20090.06). However, there was no difference in the sensitivity between parallel testing and T-SPOT.TB assay alone.We also evaluated the diagnostic values of the combination of T-SPOT.TB and TST for ATB diagnosis in these 63 patients  TB report . Patientnfection , but thenfection .M.tuberculosis-specific antigens [In recent years, IGRAs emerges as accurate diagnostic assays for screening of TB infection. IGRAs specifically detect the presence of specific effector T cells which response to antigens . Dai Y. antigens . And Theantigens . Consistantigens .6 PBMC as the cutoff, the specificity and PLR are improved for diagnosing ATB in BD patients. Although, neither T-SPOT.TB nor TST can clearly distinguish between LTB infection, ATB or history of TB infection [In this study, we evaluated T-SPOT.TB and TST for the diagnosis of ATB in BD patients. To our knowledge, this is the first study that reports the diagnostic value of T-SPOT.TB for TB in BD patients. We reported the high sensitivity (88.9%) and specificity (74.8%) of T-SPOT.TB for the diagnosis of ATB in BD patients. Compared to Dai Y\u2019s study , our stunfection , 26, we nfection . Meanwhinfection .It has been reported that the pooled sensitivity of T-SPOT.TB was higher than that of TST for the diagnosis of ATB in culture-confirmed or non-confirmed TB . A discrMycobacterium tuberculosis antigens with no evidence of clinically ATB. Given the risk of LTB infection reactivation increases in patients receiving immunosuppressive therapy or with immune dysfunction [LTB infection is defined as the sustained immune response to function , it is ifunction \u201333 suggefunction \u201336. Alsofunction , 37, andfunction  . In ourfunction , 38. A nfunction . SecondlM.tuberculosis bacteriologically confirmed, which might introduce a high risk of bias. Fourth, given no gold standard of LTB infection is available, the precision of the diagnosis of LTB infection by T-SPOT.TB could not be assessed in this study. Nevertheless, this is the first study on the diagnostic value of T-SPOT.TB for TB in BD patients. A large-scale, multi-center study enrolled bacteriologically confirmed TB patients are warranted to confirm our findings.Our study has several limitations. First, our study is a retrospective analysis of a case series, which may have incomplete data of TST. Second, since all BD patients were enrolled from a single-center with relatively small sample sizes, and our center is a national referred center for complicated rheumatic diseases, potential selection bias of BD patients is possible. Third, it is challenging to identify active TB in BD patients based on clinical features since systemic inflammation is a shared feature of both BD and TB. The majority of our ATB patients were diagnosed according to clinical criteria rather than 6 PBMC) strongly suggested ATB. Further study is needed to investigate the diagnostic value of T-SPOT.TB for LTB infection in BD patients.T-SPOT.TB may assist in the diagnosis of ATB in BD patients, and the higher number of SFCs (>\u200970/10"}
+{"text": "In this study, resistance spectrum of TA7733 was assayed by using 15 Blumeria graminis f. sp. tritici (Bgt) isolates prevalent in different regions of China. The result indicated that TA7733 was highly resistant to all tested Bgt isolates and the gene(s) on chromosome 2Mb conferred broad-spectrum resistance to powdery mildew. In order to characterize mechanism of powdery mildew resistance by identifying candidates R-genes derived from Ae. biuncialis chromosome 2Mb and develop 2Mb-specific molecular markers, we performed RNA-seq analysis on TA7733 and CS. In total we identified 7,278 unigenes that showed specific expression in TA7733 pre and post Bgt-infection when compared to CS. Of these 7,278 unigenes, 295 were annotated as putative resistance (R) genes. Comparatively analysis of R-gene sequences from TA7733 and CS and integration CS Ref Seq v1.0 were used to develop R-gene specific primers. Of 295 R-genes we identified 53 R-genes were specific to 2Mb and could be involved in powdery mildew resistance. Functional annotation of majority of the 53 R-genes encoded nucleotide binding leucine rich repeat (NLR) protein. The broad-spectrum resistance to powdery mildew in TA7733 and availability of 2Mb-derived putative candidate R-gene specific molecular markers identified in this study will lay foundations for transferring powdery mildew resistance from 2Mb to common wheat by inducing CS-Ae. biuncialis homoeologous recombination. Our study also provides useful candidates for further isolation and cloning of powdery mildew resistance gene(s) from Ae. biuncialis chromosome 2Mb.Powdery mildew is one of the most widespread diseases of wheat. The development and deployment of resistant varieties are one of the most economical and effective methods to manage this disease. Our previous study showed that the gene(s) at 2M Triticum aestivum L., 2n = 6x = 42, AABBDD), one of the most widely planted crops in the world provides 20% of the calories and 25% of its protein consumed by human [Fusarium head blight (FHB) and powdery mildew. Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most destructive diseases all over the world, with severe yield losses ranging from 13% to 50% [Common wheat  was more than 80 at 54 loci [Pm genes were derived from wild relatives of wheat. However, some Pm genes had been defeated by new virulent Bgt races or by races that were previously present at very low frequencies in the pathogen population [Wild relatives of common wheat contain a large number of  54 loci , of whicpulation ,11, and pulation ,13. TherAegilops biuncialis  is a tetraploid wild relative of wheat, belonging to the section Polyeides of the genus Aegilops. The Ub genome of it was derived from Ae. umbellulata , and Mb genome from the diploid Ae. comosa  [Aegilops biuncialis owns many desired agronomic traits for wheat improvement, such as resistance to yellow rust [Ae. biuncialis with wheat, develop a series of wheat-Ae. biuncialis addition lines, and transfer desired genes from Ae. biuncialis into wheat [Ae. biuncialis 2Mb disomic addition line TA7733 conferred high resistance to powdery mildew compared with its recipient parent CS [ 14, MM) . Aegiloplow rust , brown rlow rust , powderylow rust , toleranlow rust \u201321, highlow rust , and spelow rust . Successto wheat ,24,25. Iarent CS .Ph) genes in hexaploid wheat backgrounds [Thinopyrum intermedium by RNA-seq analysis [Pm60 from T. urartu by combining genetic mapping and RNA-seq analysis [The isolation and cloning of plant disease resistance genes had great significance for both plant disease resistance breeding and the study on molecular mechanisms of disease resistance. Map-based cloning is currently an important method to isolate novel genes. However, it is very challenging to perform fine mapping and map-based cloning of alien genes derived from wild relatives of wheat due to the strict control of homoeologous recombination by pairing homoeologous  from chromosome 2Mb, as well as further understanding of the molecular and genetic mechanisms of disease resistance conferred by Ae. biuncialis chromosome 2Mb.In this study, we report the assays of a broad-spectrum resistance gene(s) on chromosome 2Mn = 6x = 42, AABBDD), Ae. comosa TA2102 , and CS-Ae. biuncialis 2Mb disomic addition line TA7733 (2n = 44) where a pair of 2Mb chromosomes derived from Ae. biuncialis were added into CS genetic background were used in this study. All the materials were kindly provided by the Wheat Genetics Resource Center at Kansas State University, USA and maintained at the experimental station of Henan Agricultural University, China.Common wheat landrace CS  . The cytGenomic DNA (gDNA) was extracted from fresh leaves using a modified hexadecyl trimethyl ammonium bromide (CTAB) method . The conin situ hybridization (GISH) was applied to analyze the chromosomal composition of TA7733. Genomic DNA of Ae. comosa accession TA2102  and wheat CS were respectively used for probe labeling with fluorescein-12-dUTP and blocking at a ratio of 1:130 to distinguish Ae. biuncialis 2Mb chromosome. GISH was carried out as described by Liu et al. (2017) [Genomic . (2017) . Hybridiin situ hybridization (ND-FISH) [10. The first six were labeled with 6-carboxytetramethylrhodamine (TAMRA) generating red signals, and the last two being labeled with 6-carboxyfuorescein (FAM) generating green signals. All the oligonucleotides were synthesized at Sangon Biological Technology, Shanghai, China.After GISH, the hybridization signals were washed off with phosphate-buffered saline (PBS). Eight single-strand oligonucleotides were then used as probes for dual-color nondenaturing fluorescence ND-FISH) ,39. The Bgt isolates collected in Henan Province were used to evaluate the resistance of CS and CS-Ae. biuncialis 2Mb disomic addition line TA7733. Fifteen prevalent Bgt isolates collected from different regions of China were chosen to evaluate the resistance spectrum of TA7733 at the seedling stage by using CS as a susceptible control. The 15 Bgt isolates were provided by Prof. Pengtao Ma, Yantai University, China. They were all single-pustule-derived powdery mildew virulent isolates by separate artificial inoculation. The infection type (IT) were scored 7\u201310 days post-inoculation using a 0 to 4 rating scale [A mixture of prevailing ng scale , with 0 Bgt inoculation, the first leaves of TA7733 and CS were cut into 2 cm segments and stained with coomassie brilliant blue following Li et al. (2016) [Bgt development on the leaves.At 10 days post-. (2016)  for furtBgt isolates. Leaves at 0, 12, 24, 48 and 72 hours post-inoculation (hpi) were respectively collected, rapidly frozen in liquid nitrogen and stored at -80\u00b0C for RNA extraction.Seeds of CS and TA7733 soaking in water for 24 h at 23\u00b0C were transferred into a mixture of nutrient soil and vermiculite (1:1). Seedlings with full extended first leaf were dusted using fresh conidiophores of Total RNA of ten samples  were extracted for transcriptome sequencing. Then equal amounts of RNA samples 12\u201372 hpi from TA7733 and CS were mixed to generate RNA-seq sample RI and SI, respectively. RNA at 0 hpi from TA7733 and CS were accordingly represented as RNA-seq sample RC and SC. Two biological replicates were performed in this study, forming a total of eight RNA samples . The designations 1 and 2 are used to represent replicates 1 and 2, respectively. Libraries with an average insert size of 200 bp constructed from these eight samples were then sequenced using the Illumina HiSeqTM 2500 by the Beijing Genomics Institute.\u22125) against the NCBI non-redundant (NR) protein, SWISSPROT, gene ontology (GO), eukaryotic orthologous groups (KOG), kyoto encyclopedia of genes and genomes (KEGG) and plant resistance gene (PRG) databases.Prior to assembly, sequencing raw reads were pre-processed using a Perl script dynamic-Trim.pl to remove the adaptor sequences, low-quality sequences, low complexity sequences, short reads and empty reads. Reads data with a quality score (Qphred) \u2265 50 (Q50: ratio of an error rate of 0.01%) were then merged and input into the data assembly software Trinity for assembling into transcripts. The generated unigenes were annotated by a Blastx alignment search , 0.25 \u03bcl forward primer (10 \u03bcmol/l), 0.25 \u03bcl reverse primer (10 \u03bcmol/l), 7.5 \u03bcl Taq MasterMix  and 5 \u03bcl ddH2O. PCR cycling conditions were as follows: 94\u00b0C for 5 min followed by 35 cycles of 94\u00b0C for 30 s, 50\u201366\u00b0C for 30 s, and 72\u00b0C for 1 min, followed by a final 10-min extension at 72\u00b0C. The PCR products were digested with four base-restriction enzymes. Five microliters of a restriction enzyme mixture containing 2.8 \u03bcl of ddH2O, 2.0 \u03bcl of CutSmart buffer, and 0.2 \u03bcl of an enzyme stock solution was added to 15 \u03bcl of PCR products and incubated for 3.5 h at 65\u00b0C. The PCR or restricted PCR products were separated on a 2.0% agarose gel-electrophoresis stained with ethidium bromide and visualized by UV light.R gene-specific primer sets were designed based on their transcriptome sequences to perform PCR amplification using gDNA from TA7733 and CS as templates to verify 2MAe. biuncialis chromosome 2Mb. Comparative maps of 2Mb-specific R genes were made using MapDraw software referring homoeologous chromosome locations of CS Ref Seq v1.0.Genome sequences of wheat landrace CS (CS Ref Seq v1.0) were used as references in Blastn searches to obtain position information for R genes from Ae. biuncialis 2Mb disomic addition line TA7733 by using fluorescein-labeled gDNA from M genome donor Ae. comosa as a probe and wheat CS DNA as blocker. As shown in Ae. biuncialis 2Mb chromosomes in TA7733, confirming the disomic addition of chromosome 2Mb.GISH and ND-FISH were respectively performed to confirm the chromosome composition of CS-Bgt isolates collected in Henan Province was used to inoculate seedlings with fully-extended first leaves of TA7733 and its recipient parent CS in the greenhouse. Ten days post-inoculation, the leaves of CS were covered with a large number of Bgt hyphae, with ITs of 3\u20134, whereas TA7733 showed only stunted spores, with ITs 0 to 1  to all the 15 Bgt isolates tested, whereas its recipient parent CS was highly susceptible (IT = 4 or 3) to all tested Bgt isolates. These results indicated that the chromosome 2Mb in TA7733 conferred broad-spectrum resistance to powdery mildew of wheat.The resistance spectrum of TA7733 was further assayed at the seedling stage by inoculation of 15 prevalent Ae. biuncialis 2Mb disomic addition line TA7733 and its recipient parent CS were respectively conducted pre and post Bgt-infection. A total of 158,953 unigenes were assembled with a total length of 198,364,757 bp. The average unigene size was 1247.95 bp ranging from 301 to 19,496 bp , 48,724 (30.65%), 40,543 (25.51%), 37,008 (23.28%), 13,414 (8.44%) and 10,969 (6.92%) annotated unigenes, respectively  unigenes were assigned to one or more GO term annotations , of whicThe KEGG database was used to systematically describe the pathway where the unigenes involved. Out of a total 158,953 annotated unigenes, 26,589 unigenes were assigned to 23 KEGG pathways . The mosAe. biuncialis chromosome 2Mb, which should be only expressed in TA7733 other than in CS. Based on pairwise comparison of unigenes of TA7733 vs CS, a total of 7,278 genes were uniquely expressed in TA7733, of which 4,382 unigenes were significantly differentially expressed post vs before Bgt-inoculation, and the remaining 2,896 unigenes had insignificantly different expression levels. In consideration of the fact that expression levels of some cloned resistance genes did show no significant difference before and after pathogen infection [b-derived R-genes involved in powdery mildew resistance.One of the objectives of this study was to explore putative R genes specific to nfection , these 2Ae. biuncialis 2Mb chromosome.To analyze the biological pathways of these 4,382 unigenes, the statistical enrichment of differentially expressed genes (DEGs) in KEGG pathways were tested using the KOBAS software. In consequence, 399 out of 4,382 DEGs were allocated to 162 KEGG pathways . The mosBased on transcriptome data analysis, 7,278 unigenes were uniquely expressed in TA7733. Only 295 of 7,278 unigenes were annotated as putative R-genes by Blastx alignment against the PRG and NCBI databases. However, of 295 R-genes sequences when blastn searched against CS Ref Seq v1.0, only 61 (20.68%) R-genes mapped to wheat homoeologous group 2, and the remaining 234 (79.32%) R-genes to none of the homoeologous group 2 chromosome of wheat.Ae. biuncialis chromosome 2Mb, a total of 61 sets of PCR primer pairs were designed based on transcriptome sequences of these R genes. PCR amplification of gDNA of CS and TA7733 confirmed 40 R genes to be specific to chromosome 2Mb, which producing unique amplification in CS-Ae. biuncialis 2Mb disomic addition line TA7733  structures, a central nucleotide-binding (NB) subdomain and a leucine-rich repeat (LRR) domain, 12 in NL class containing NBS and LRR domains, but lack of CC domain, five in class RLP which contains leucine-rich receptor-like repeat, a transmembrane region of 25AA, and a short cytoplasmic region, each one for TNL class which contains a central NB subdomain, a LRR domain, a interleukin-1 receptor (1L-1R) domain, and N class only containing NBS domain . The remAe. biuncialis chromosome 2Mb specificity with those in CS Ref Seq v1.0  on Ae. biuncialis 2Mb chromosome should be a new Pm gene(s).Development of resistant wheat varieties is the most important and environment-friendly way to control programs . Wild re group 2 , 42\u201344. Pm60, a map-based cloned powdery mildew gene, showed no significant differences at various time points after Bgt E09 infection based on qRT-PCR analysis [bsr-d1 in rice were not significantly up-regulated after blast infection [Bgt-resistance related candidate genes from all specifically expressed R genes in TA7733 regardless of significance of their expression level changes post vs before Bgt-infection. After PCR verification by using R gene sequence-based primer sets and integrating transcriptome sequences blastn against CS Ref Seq v1.0, we finally verified 53 R genes candidates of chromosome 2Mb specificity, which included 47 unregulated, four up-regulated and two down-regulated genes.Previous studies have generally focused on the significantly differentially expressed genes in interactions between plant and pathogens to explore disease resistance-related genes by transcriptome sequencing \u201347. Howenfection . So, thePm genes including Pm2 [Pm3 [Pm8 [Pm21 [Pm60 [b-specific R genes were predicted to encode CC-NBS-LRR protein. These 14 R genes should be considered as the most promising candidate genes for further isolating and cloning Pm genes carried by Ae. biuncialis chromosome 2Mb.Isolation of plant resistance (R) gene is greatly helpful to breed resistant varieties and elucidates resistance molecular mechanisms. Conventional map-based cloning proved to be a effective method to clone R genes , howeverding Pm2 , Pm3 [50Pm3 [Pm8 , Pm21 [2m8 [Pm21 ,52 and P21 [Pm60 , have beD. villosum 6V#4S-specifc markers using transcriptome data [Ae. longissima chromosome-specific markers by RNA-seq [D. villosum#4 based on transcriptome data [Ae. biuncialis chromosome 2Mb. These markers will be useful to assist the transfer resistance gene(s) from 2Mb into common wheat by inducing CS-Ae. biuncialis 2Mb homoeologous recombination for wheat disease breeding in the future.GISH is a popular visual method to identify alien chromosome or chromatin in wheat background. Whereas GISH is expensive and time-consuming especially used to screen a large population derived from distant crossing between wheat and its wild relatives . In contome data . Wang et RNA-seq . Li et aome data . Furtherome data . Therefoome data ,58. In tAe. biuncialis chromosome 2Mb was verified to be board-spectrum in this study. It could be a valuable disease-resistance resource for wheat breeding programs. Fifty-three disease resistance gene candidates of 2Mb specificity, which were selected based on transcriptome sequencing analyses, will be greatly helpful to further isolate and clone Pm gene(s) derived from chromosome 2Mb and provide the insights into molecular mechanism of 2Mb-conferred powdery mildew resistance. Furthermore, 53 R gene sequence-based functional molecular markers of 2Mb specificity in this study will facilitate the transfer of resistance gene(s) from 2Mb to common wheat by inducing CS-Ae. biuncialis homoeologous recombination.In summary, powdery mildew resistance gene(s) on S1 Fig(M) 100 bp DNA Ladder.  CS.  TA7733. (A) CL119404Contig1. (B) CL88277Contig1. (C) CL82670Contig1. (D) 82789Contig1. (E) CL82700Contig1. (F) CL85355Contig1. (G) CL66003Contig1. (H) CL89405Contig1. (I) CL106750Contig1. (J) CL119216Contig1. (K) CL19981Contig2. (L) CL93721Contig1. (M) CL84424Contig1. (N) CL88613Contig1. (O) CL91022Contig1. (P) 96221Contig1. (Q) comp19533_c0_seq1_6. (R) CL85258Contig1. (S) CL113949Contig1. (T) CL86521Contig1. (U) CL105879Contig1. (V) CL90029Contig1. (W) CL84846Contig2. (X) CL87530Contig1. (Y) CL29910Contig1. (Z) CL92547Contig1. (AA) CL75219Contig1. (AB) CL108886Contig1. (AC) CL113652Contig1. (AD) CL80063Contig1. (AE) CL89447Contig1. (AF) CL93169Contig1. (AG) CL114224Contig1. (AH) CL116612Contig1. (AI) CL67241Contig1. (AJ) CL119539Contig1. (AK) CL90483Contig1. (AL) CL91742Contig1. (AM) comp84147_c0_seq1_6. (AN) CL124Contig7. (AO) CL100654Contig1. (AP) CL104996Contig1. (AQ) CL107524Contig1. (AR) CL107607Contig1. (AS) CL465Contig5. (AT) CL66266Contig1. (AU) CL72629Contig1. (AV) CL75868Contig1. (AW) CL86319Contig1. (AX) CL79458Contig1. (AY) comp121700_c0_seq1_5. (AZ) comp80277_c0_seq1_7. (BA) comp93868_c0_seq1_7.(TIF)Click here for additional data file.S1 Raw_imageAegilops biuncialis 2Mb disomic addition line TA77333; 3, common wheat CS; 4, CS-Ae. biuncialis 2Mb disomic addition line TA77333. (A) CL119404Contig1. (B) CL88277Contig1. (C) CL82670Contig1. (D) 82789Contig1. (E) CL82700Contig1. (F) CL85355Contig1. (G) CL66003Contig1. (H) CL89405Contig1. (I) CL106750Contig1. (J) CL119216Contig1. (K) CL19981Contig2. (L) CL93721Contig1. (M) CL84424Contig1. (N) CL88613Contig1. (O) CL91022Contig1. (P) 96221Contig1. (Q) comp19533_c0_seq1_6. (R) CL85258Contig1. (S) CL113949Contig1. (T) CL86521Contig1. (U) CL105879Contig1. (V) CL90029Contig1. (W) CL84846Contig2. (X) CL87530Contig1. (Y) CL29910Contig1. (Z) CL92547Contig1. (AA) CL75219Contig1. (AB) CL108886Contig1. (AC) CL113652Contig1. (AD) CL80063Contig1. (AE) CL89447Contig1. (AF) CL93169Contig1. (AG) CL114224Contig1. (AH) CL116612Contig1. (AI) CL67241Contig1. (AJ) CL119539Contig1. (AK) CL90483Contig1. (AL) CL91742Contig1. (AM) comp84147_c0_seq1_6. (AN) CL124Contig7. (AO) CL100654Contig1. (AP) CL104996Contig1. (AQ) CL107524Contig1. (AR) CL107607Contig1. (AS) CL465Contig5. (AT) CL66266Contig1. (AU) CL72629Contig1. (AV) CL75868Contig1. (AW) CL86319Contig1. (AX) CL79458Contig1. (AY) comp121700_c0_seq1_5. (AZ) comp80277_c0_seq1_7. (BA) comp93868_c0_seq1_7.Lanes: M, 100 bp Ladder DNA Marker; 1, common wheat CS; 2, CS-(PDF)Click here for additional data file.S1 Table(XLS)Click here for additional data file.S2 Table(XLS)Click here for additional data file.S3 Table(XLS)Click here for additional data file.S4 Table(XLS)Click here for additional data file.S5 Table(XLS)Click here for additional data file.S6 Table(XLS)Click here for additional data file."}
+{"text": "Scyphiphora hydrophyllacea Gaertn. collected in Hainan Province of China resulted in the isolation of a new iridoid, scyphiphin C (1) and a known iridoid glycoside, shanzhiside methyl ester (2). Their structures were elucidated by a study of their physical and spectral data.Chemical investigation of the ethanol extract of the aerial parts of  Scyphiphora hydrophyllacea Gaertn. F. (Rubiaceae), one of the mangrove plants, is distributed from south to southeast Asia, Caroline Islands, Australia, and New Caledonia  to give 26 fractions. Fraction 16 (5.02 g) was subjected to column chromatography over silica gel eluted with light petroleum-EtOAc (4:6) to afford nine further fractions. Sub-fraction 4 (624.2 mg) was fractionated by column chromatography (Sephadex LH-20) eluted with 95 % EtOH and then rechromatographed on a silica-gel column with light petroleum-EtOAc (3:7) to afford compound 1 (18.7 mg). Fraction 18 (10.06 g) was subjected to vacuum liquid column chromatography over RP-18 eluted with MeOH-H2O  gradually to afford eight further fractions. Sub-fraction 1 (2.18 g) was purified by silica gel CC eluted with CHCl3-MeOH (9:1) to afford compound 2 (290.2 mg).The dried, milled aerial parts of Scyphiphin C (1): White amorphous powder,  = - 13.4 . IR (KBr): n = 3875, 3858, 3605, 3308, 2941, 2247, 1482, 865 cm-1. HR-ESI-MS (positive): m/z = 269.0998 . 1H and 13C-NMR: see Shanzhigenin methyl ester (2): White amorphous powder,  = - 380.0 . IR (KBr): n = 3922, 3886, 3779, 3654, 3700, 3543, 3112, 3081, 2326, 1583, 891 cm-1. HR-ESI-MS (positive): m/z = 429.1380 .Scyphiphora hydrophyllaceaGaertn., a new iridoid scyphiphin C (1) and a known iridoid glucoside shanzhigenin methyl ester (2) were isolated. Their structures were established on the basis of spectroscopic evidence.As a part of our chemical investigation on"}
+{"text": "Open Biol.7, 170165. (Published online 13 September 2017). (doi:10.1098/rsob.170165)A correction is required to The correct version of table 2 is as follows:"}
+{"text": "Flammulina rossica (wood-rotting basidiomycete) genome was performed to identify its carbohydrate-active enzymes (CAZymes). De novo genome assembly (31 kmer) revealed a total length of 35,646,506 bp (49.79% GC content). In total, 12,588 gene models of F. rossica were predicted using an ab initio gene prediction tool (AUGUSTUS). Orthologous analysis with other fungal species revealed that 7433 groups contained at least one F. rossica gene. Additionally, 12,033 (95.6%) of 12,588 genes for F. rossica proteins had orthologs among the Dikarya, and F. rossica contained 12 species-specific genes. CAZyme annotation in the F. rossica genome revealed 511 genes predicted to encode CAZymes including 102 auxiliary activities, 236 glycoside hydrolases, 94 glycosyltransferases, 19 polysaccharide lyases, 56 carbohydrate esterases, and 21 carbohydrate binding-modules. Among the 511 genes, several genes were predicted to simultaneously encode two different CAZymes such as glycoside hydrolases (GH) as well as carbohydrate-binding module (CBM). The genome information of F. rossica offers opportunities to understand the wood-degrading machinery of this fungus and will be useful for biotechnological and industrial applications.Next-generation sequencing (NGS) of the  Flammulina rossica  was first identified in 1999 by Redhead and Petersen E) , suggesting that these GHs can be secreted (The GH16 family consists of agarase (EC 3.2.1.81), lichenase (EC 3.2.1.73), \u03ba-carrageenase (EC 3.2.1.83), xyloglucan xyloglucosyltransferase (EC 2.4.1.207), endo-\u03b2-1,3-glucanase (EC 3.2.1.39), endo-\u03b2-1,3-1,4-glucanase (EC 3.2.1.6), and endo-\u03b2-galactosidase (EC 3.2.1.103), and most of these enzymes contain conserved motifs such as Glu-Xaa-Asp-Xaa-(Xaa)-Glu (EXDX[X]E). The first and last glutamic acid (E) residues of the conserved motif are characterized as a nucleophile and Br\u00f8nsted acid/base, respectively, and play an important role in the catalytic activity of GH16 family enzymes ,52,53. AXDX[X]E) . Althougsecreted . F. rossica also contains a series of genes related to cellulase (27 genes), xylanase (6 genes), and chitinase (23 genes) in its genome sequence  . Cellulasequence . Cellobis genome .F. rossica with more than 200 genes encoding various GHs showed strong potential for diverse applications, such as biotechnology and industry.Simultaneous actions of several GHs are necessary to effectively degrade plant cell wall complexes composed of cellulose and xylan. Recently, genome sequencing of various bacterial and fungal species has reviewed the various activities of GHs on cellulose, chitin, and xylan degradation and their potential for biotechnological applications and industrial degradation of biopolymers ,59,60,61F. rossica genome based on three different databases  searches (PLs EC 4.2.2.-) cleave polymer chains of polysaccharides, essential cellular components of all living organisms, through a \u03b2-elimination mechanism to produce unsaturated polysaccharides ,63. PLs .2.2.- clsearches . Among tsearches . In addisearches . PL20 wasearches  64]..F. rossid-galacturonate) or pectin  is an acidic polysaccharide in plant cell walls [F. rossica and other fungal species. However, according to this study, genes encoding PL families 2 and 10 were not found in F. rossica and in other fungal species genomes  but not in other fungal species  are classified into AA families, which are mainly involved in the depolymerization of non-carbohydrate structural components (lignin) of plants . These Asequence . Among tectively . Interes species . In addiomycetes .3-His-Xaa4-Met/Leu/Phe [18-Glu that interacts with the FAD cofactor [F. rossica was also found to contain 24 genes predicted to encode AA3, and 17 genes contained the \u03b2-\u03b1-\u03b2 dinucleotide binding motif in their amino acid sequences  has conserved motifs (copper binding motifs) within its amino acid sequence such as His-Xaa-His, His-Xaa-His-Gly, His-Xaa-Xaa-His-Xaa-His, and His-Cys-His-Xaa/Leu/Phe . Our rescofactor ,71,72. Fequences . These rAmino acid sequence within a carbohydrate-active enzyme with carbohydrate-binding activity is known as CBM ,76. CBM F. rossica genome based on three different database  searches. CBM family 13 was prominent, and six families, including CBM18, -20, -22, -35, -48, -63, and -67, were represented by only one CBM in the F. rossica genome  genes that simultaneously encode CBM suggest that CBM is required for efficient substrate degradation , trisaccharide (cellotriose), and oligosaccharide (cellotetraose) to ethanol at similar recovery rates to glucose and to convert glucose to ethanol at a similar level as S. cerevisiae [F. velutipes is suitable for consolidated bioprocessing (CBP), which is considered an effective process for bioethanol production from lignocellulosic biomass [F. velutipes, the closest white rot fungus to F. rossica, is a very attractive model for bioethanol production due to the numerous genes associated with lignocellulolytic enzymes such as CAZymes [F. rossica genome to identify genes involved in lignocellulose degradation. As described above, various CAZyme genes were identified in the F. rossica genome including 102 auxiliary activities, 236 glycoside hydrolases, 94 glycosyltransferases, 19 polysaccharide lyases, 56 carbohydrate esterases, and 21 carbohydrate binding-modules. Although further studies on CAZyme genes are needed, this study suggests that F. rossica has great potential for future production of biomaterials such as bioenergy.This study aimed to improve the understanding of lignocellulolytic machinery in revisiae ,80. This biomass ,82,83. P"}
+{"text": "Multiple medication use within one year is associated with increased fall injury risk in older adults. However, chronically using multiple medications and treated fall injury have rarely been explored, particularly in cohort studies linked with claims data. We examined using >5 medications in 2 or more consecutive years (chronic medication use) as a risk factor for treated fall injury in 1,898 community-dwelling adults  with linked Medicare Fee-For-Service (FFS) claims from the Health, Aging and Body Composition Study since 1997/98 clinic visit. Incident fall injury (N=546) was the first claim from 1998/99 clinic visit to 12/31/08 with an ICD-9 fall code and non-fracture injury code, or fracture code with/without a fall code. Stepwise Cox models with a time-varying predictor of chronic medication use before fall injury or censoring (N=414) vs. not using >5 medications at the same time (N=1008) were adjusted for baseline demographics, lifestyle factors, fall history, quadriceps strength, cardiovascular disease (CVD), diabetes, sensory nerve impairment, and kidney function. Fall injury risk increased for chronic medication users (37%) vs. non-users (29%) (HR=1.25[1.00-1.57]), though was attenuated after adjustment for CVD and diabetes (HR=1.18[0.93-1.51]). Sensitivity analyses excluding fall-risk-increasing drugs (FRIDs) from medication counts (HR=1.32[0.54-3.20]), or including those using >5 medications non-chronically (N=365) in referent groups (HR=1.22[0.96-1.55]) had consistent findings. Unmeasured comorbidity differences may confound associations of chronic medication use and treated fall injury risk in older adults with Medicare FFS. Considering both chronic diseases and medication use in fall risk assessments is needed."}
+{"text": "Stenotrophomonas maltophilia is a prevalent nosocomial pathogen with multidrug resistance. Here, we describe the complete genome of S. maltophilia myophage Moby, which shares characteristics with Enterobacteria phage T4 and is closely related to Stenotrophomonas phage IME-SM1. Moby has a 159,365-bp genome with 271 predicted protein-coding genes and 24 predicted tRNAs. Stenotrophomonas maltophilia is a prevalent nosocomial pathogen with multidrug resistance. Here, we describe the complete genome of S. maltophilia myophage Moby, which shares characteristics with Enterobacteria phage T4 and is closely related to Stenotrophomonas phage IME-SM1. Moby has a 159,365-bp genome with 271 predicted protein-coding genes and 24 predicted tRNAs. Stenotrophomonas maltophilia is a multidrug-resistant Gram-negative bacterium with rising prevalence as a nosocomial pathogen  was grown aerobically at 30\u00b0C in nutrient broth or agar (BD), and the phage was propagated by the soft-agar overlay method (www.bioinformatics.babraham.ac.uk/projects/fastqc), and trimming was performed using FastX Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/). Assembly into a single contig with 673.7-fold coverage was performed using SPAdes v3.5.0 , and, unless otherwise stated, were executed using default parameters (Moby was isolated from filtered (0.2-\u03bcm pore size) wastewater collected in Bryan, TX. The y method . Phage Dy method . An Illust ATCC 1807 was gs v3.5.0 . Rho-inds v3.5.0 . Gene fus v3.5.0 \u201315. Stru v3.5.0 \u2013. progresost ATCC 807 was grameters . Phage mrameters .pac-type headful DNA packaging mechanism used by T4-like phages (Stenotrophomonas phage IME-SM1 (GenBank accession no. KR560069), an unclassified myovirus. No introns were identified in Moby, including in the predicted thymidylate synthase and nrdB genes, which contain introns in phage T4 (GenBank accession no. NC_000866)  .MN095772, BioProject accession no. PRJNA222858, SRA accession no. SRR8893605, and BioSample accession no. SAMN11414490.The genome sequence and associated data for phage Moby were deposited under GenBank accession no."}
+{"text": "Triticum aestivum L.) through development and cultivation of superior genotypes incorporating yield-related agronomic and physiological traits derived from genetically diverse and complementary genetic pool. Despite significant breeding progress, yield levels in wheat have remained relatively low and stagnant under marginal growing environments. There is a need for genetic improvement of wheat using yield-promoting morpho-physiological attributes and desired genotypes under the target production environments to meet the demand for food and feed. This review presents breeding progress in wheat for yield gains using agronomic and physiological traits. Further, the paper discusses globally available wheat genetic resources to identify and select promising genotypes possessing useful agronomic and physiological traits to enhance water, nutrient-, and radiation-use efficiency to improve grain yield potential and tolerance to abiotic stresses . Finally, the paper highlights quantitative trait loci (QTL) linked to agronomic and physiological traits to aid breeding of high-performing wheat genotypes.Enhanced grain yield has been achieved in bread wheat ( Triticum aestivum L., 2n = 6x = 42) is the world\u2019s third important staple food crop after maize (Zea mays) and rice (Oryza sativa). It is the most widely grown crop globally  are discussed. Finally, the paper highlights QTL linked to agronomic and physiological traits to aid breeding of high-performing wheat genotypes.Grain yield response in wheat is influenced by several agronomic and physiological traits . AgronomIndia, Russia, China, and Kazakhstan are currently the leading wheat producers with approximately 30, 27, 24, and 12 million hectares devoted to wheat production, respectively. In terms of total production, China is the world\u2019s leading wheat producer with approximately 131 million tons per year . Other c-1 yr-1), Chile (246 kg ha-1 yr-1), France (123 kg ha-1 yr-1), and Mexico (41.77 kg ha-1 yr-1), whereas relatively lower genetic progress were reported in Spain (24 kg ha-1 yr-1), Australia (25\u2005kg ha-1 yr-1), and Siberia (15.3 kg ha-1 yr-1). Annual yield gains in Egypt, India, and Pakistan were estimated at 27.4 (0.55%), 21.4 (0.62%), 111.6 (1.13%), 32.5 (0.83%), and 18.5 kg ha-1 yr-1 (0.5%), respectively (-1 (0.65%) (Wheat yield gains across the major wheat producing countries are presented in ectively . Genetic (0.65%) . Countri (0.65%) .Genetic progress is relatively lower under low-yielding environments compared to high-yielding environments . TherefoGenetic gains in grain yield have been attributed to development and deployment of high-yielding wheat genotypes with improved agronomic and physiological traits related with high yield potential . For exaGrain yield in wheat is influenced by several agronomic traits  which haVrn-A1, Vrn-B1, and Vrn-D1 regulate flowering and maturity in wheat . The effect of Vrn loci on heading and maturity, and grain yield potential are ranked as follows: Vrn-A1 < Vrn-B1 < Vrn- D1  among Chinese wheat cultivars followed by Vrn-A1 , grain yield (23 and 40%), and HI (31 and 50%) from tall and dwarf genotypes, respectively. Canadian spring wheat carrying dominant allele of Vrn-B1, photo-period insensitive allele of Ppd-D1, and height reducing allele Rht-1 produced shorter plants and higher grain yield , Rht2 (Rht-D1b), Rht-D1c, and Rht8  but resulted in reduced grain number per spike and increased 1,000\u2013kernel weight, above-ground biomass and grain yield in wheat  have negligible effects on biomass production, whereas some  can increase above-ground biomass, kernel weight, and grain yield  and 0.3% yr-1 for TKW among Chinese and American wheat genotypes, respectively. Similarly, -1 among Brazilian wheat genotypes. TKW is reportedly linear with moderate to high correlation with grain yield (Grain yield improvement has been significantly associated with increased thousand kernel weight (TKW) . On the in yield  suggestiin yield , rather -1 y-1 (0.44% yr-1) between 1980 and 2003 attributed to high number of kernels spike-1 (0.24 kernels spike-1 yr-1) in Spain. Similarly, -2 yr-1 through breeding and selection spanning over 50 years in Iran. Grain number among Brazilian wheat genotypes was increased by 77.89 grains yr-1. In China grain number per spike varied between 26 for landraces developed in 1941 to 38 for improved wheat genotypes released between 2007 and 2011 (The number of grains per spike has been identified as an important trait for improving grain yield . Yield gand 2011 . Among wand 2011 . Althougand 2011 . Therefoand 2011 . The reland 2011 . On the and 2011 .-1 was derived from \u2018Zhoumai 16\u2019 x \u2018Liken 4\u2019. Zhoumai 16 was developed from \u2018Yumai 21\u2019 x \u2018Zhou 8425B\u2019 whereby Zhou 8425B is characterized by large spikes and high TKW (Spike fertility (SF) is a grain yield component that influences the increase in the number of kernels per spike . For inshigh TKW , demonstOther spike characteristics useful for breeding include SL, SPS, and SC . The numTin1 tiller inhibition gene can increase grain number per spike  and/or anthesis date genes (e.g. Ppd1) resulted in improvement of WSC. In addition, wheat genotypes with high WSC produce more fertile tillers, reduced days to anthesis, increased biomass, grain number, and grain yield than genotypes with low WSC  possess useful source of alleles for enhancing drought tolerance and improving yield and its component traits (Aegilops tauschii) with durum wheat (T. durum Desf.) (T. turgidum L.) is also identified as a useful genetic resource for wheat breeding possessing functional genes that surpass the early maturity effect caused by the early flowering allele Ppd-A1a found in T. turgidum L.   . Tetraplamidale) . Wild emamidale) . Wheat gamidale) . Furtheramidale) .QTLs QTn.ipk-5D, QTn.ipk-2D, QTn.ipk-3B, and QTn.ipk-1B which are associated with productive tiller number (QFlt.dms-2D, QFlt.dms-5B, QFlt.dms-2D, QFlt.dms-7A, and QFlt.dms-6B.2 are linked to days to flowering; whereas, QTLs QMat.dms-2D, QMat.dms-2D, QMat.dms-7A.2, and QMat.dms-4A.1 are associated with days to maturity (QTL mapping of agronomic and physiological traits is important for marker-assisted breeding in wheat . Agronomr number . QTL QFlmaturity . About 4maturity . New QTLmaturity . QTLs fomaturity Table 3 maturity . QTL\u2019s fmaturity . The idematurity .2 concentration were co-located with QTL for grain yield and/or yield components (mponents . QTL whimponents . Such plmponents . Marker-mponents . Generalmponents , their imponents . TherefoIn conclusion, genetic improvement can be achieved by either direct selection for primary traits such as grain yield or indirectly through selection of secondary traits related to higher grain yield potential. Breeding high-yielding genotypes incorporating yield-promoting agronomic and physiological traits has accelerated yield gains in wheat. As a result, further grain yield improvement will likely be achieved through in/direct selection targeting yield-related agronomic and physiological attributes. Furthermore, QTL associated with agronomic and physiological traits linked to grain yield are useful for marker-assisted selection of high-performing wheat genotypes.All authors listed have made substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Brassica rapa, an important vegetable cultivated worldwide.Genome-wide maps of histone modifications have been obtained for several plant species. However, most studies focus on model systems and do not enforce FAIR data management principles. Here we study the H3K27me3 epigenome and associated transcriptome of B. rapa leaves and inflorescences. To analyze these data we developed a Reproducible Epigenomic Analysis pipeline using Galaxy and Jupyter, packaged into Docker images to facilitate transparency and reuse. We found that H3K27me3 covers roughly one-third of all B. rapa protein-coding genes and its presence correlates with low transcript levels. The comparative analysis between leaves and inflorescences suggested that the expression of various floral regulatory genes during development depends on H3K27me3. To demonstrate the importance of H3K27me3 for B. rapa development, we characterized a mutant line deficient in the H3K27 methyltransferase activity. We found that braA.clf mutant plants presented pleiotropic alterations, e.g., curly leaves due to increased expression and reduced H3K27me3 levels at AGAMOUS-like loci.We performed H3K27me3 chromatin immunoprecipitation followed by high-throughput sequencing and transcriptomic analysis by 3\u2032-end RNA sequencing from B. rapa development. Our work reveals the epigenomic landscape of H3K27me3 in B. rapa and provides novel genomics datasets and bioinformatics analytical resources. We anticipate that this work will lead the way to further epigenomic studies in the complex genome of Brassica crops.We characterized the epigenetic mark H3K27me3 at genome-wide levels and provide genetic evidence for its relevance in  The epigenome comprises alternative chromatin states that can affect gene activity . These iArabidopsis thaliana (hereinafter referred to as Arabidopsis) the core PRC2 subunits are well conserved and H3K27me3 is crucial for plant development . Ca. CaB. raRRID:SCR_006989) [P <\u00a00.05; and Plant GO slim ontology type); data were visualized reduced in complexity and redundant GO terms using REViGO  [Gene Ontology analysis was performed using agriGO v2.0   (Fisher _005825)  with defB. rapa v3.0 gene models was obtained from Zhang et al. [B. rapa genome v3.0 was obtained by blasting coding sequences against B. rapa genome v1.5  and Arabidopsis .Functional annotation of g et al. . Custom B. rapa AGAMOUS genes, we found that BraA.AG.a (BraA01g010430.3C) gene structure annotation was not correct in the recent B. rapa genome v3.0. We curated BraA.AG.a gene structure using AUGUSTUS  [B. rapa genome V1.5) gene information at the B. rapa database [During our analyses of H3K27me3 levels on _008417)  and Bra0database .Project name: Epigenomics Workflow on Galaxy and Jupyterhttps://github.com/wilkinsonlab/epigenomics_pipelineProject home page: Operating systems: Platform independentProgramming languages: Python, R, BashOther requirements: noneLicense: MITRRID:SCR_017544https://github.com/wilkinsonlab/epigenomics_pipeline; this is associated with a Zenodo release to match the configuration used in this publication [RRID:SCR_017544 and biotools: Epigenomics_Workflow_on_Galaxy_and_Jupyter at SciCrunch and bio.tools databases, respectively. Compiled Docker images are available at https://hub.docker.com/u/mpaya. All supporting data are available in the GigaScience GigaDB database [ChIP-seq and RNA-seq data sets supporting the results of this article are available at NCBI SRA under the accession number PRJNA542357. Latest versions of the components of the REA pipeline, and instructions to deploy the Galaxy/Jupyter containers and run the analysis, can be found in the GitHub repository lication . The REAdatabase .Figure S1: Differences between epic2 and MACS2 peak-calling algorithmsB. rapa inflorescencesFigure S2: ChIP-seq analysis of H3K27me3 regions in B. rapa inflorescencesFigure S3: Comparisons of H3K27me3 ChIP-seq signal in B. rapa H3K27 methyltransferase-coding genesFigure S4: Transcript levels of BraA.clf-1 mutant phenotypesFigure S5: B. rapa plant sampling materialsFigure S6: Pictures of Figure S7. ChIP-seq sample-sample correlation testsFigure S8. RNA-seq sample-sample correlation testsTable S1. Alignment statisticsTable S2. Comparison of Bowtie2 and BWA performanceB. rapaTable S3. H3K27 trimethylated regions identified in B. rapa leavesTable S4. List of H3K27me3 peaks in B. rapa inflorescencesTable S5. List of H3K27me3 peaks in Table S6. 3\u2032RNA-seq resultsTable S7. List of H3K27me3 differentially marked regionsTable S8. H3K27me3-regulated genesTable S9. Primer listgiz147_GIGA-D-19-00256_Original_SubmissionClick here for additional data file.giz147_GIGA-D-19-00256_Revision_1Click here for additional data file.giz147_Response_to_Reviewer_Comments_Original_SubmissionClick here for additional data file.giz147_Reviewer_1_Report_Original_SubmissionDiep Ganguly -- 8/28/2019 ReviewedClick here for additional data file.giz147_Reviewer_1_Report_Revision_1Diep Ganguly -- 11/7/2019 ReviewedClick here for additional data file.giz147_Reviewer_2_Report_Original_SubmissionJingyu Zhang -- 9/1/2019 ReviewedClick here for additional data file.giz147_Supplemental_FilesClick here for additional data file.M: log2 fold-change of the normalized H3K27me3 read densities in leaves relative to inflorescences calculated by MAnorm; MACS: Model-based Analysis of ChIP-Seq; mRNA: messenger RNA; NCBI: National Center for Biotechnology Information; PRC2: Polycomb repressive complex 2; REA: Reproducible Epigenomic Analysis; RNA-seq: RNA sequencing; RT-qPCR: real-time quantitative reverse transcription PCR; s.d: standard deviation; SEA: singular enrichment analysis; SICER: Spatial Clustering for Identification of ChIP-Enriched Regions; SRA: Sequence Read Archive.3\u2032RNA-seq: 3\u2032-end mRNA high-throughput sequencing; AG: AGAMOUS; bp: base pairs; cDNA: complementary DNA; ChIP: chromatin immunoprecipitation; ChIP-seq: ChIP followed by high-throughput sequencing; ChIP-qPCR: ChIP followed by real-time quantitative PCR; CLF: CURLY LEAF; DAG: days after germination; DEG: differentially expressed genes; FAIR: findability, accessibility, interoperability, and reusability; FC: fold change; FDR: false discovery rate; GO: gene ontology; H3K27me3: histone H3 lysine 27 trimethylation; IGV: integrative genomics viewer; kb: kilobase pairs; LOWESS: Locally Weighted Scatterplot Smoothing; The authors declare that they have no competing interests.This work was supported by grants RTI2018-097749-B-100, BIO2015\u201368031-R and RYC-2013\u201314689 to P.C., and BES-2016\u2013078939 fellowship to L.P.V. from the Spanish Ministerio de Economia y Competitividad ; and by the \"Severo Ochoa Program for Centres of Excellence in R&D\u201d from the Agencia Estatal de Investigaci\u00f3n of Spain, grant SEV-2016-0672 (2017\u20132021) to the CBGP. M.P.M. was supported by a Postdoctoral contract associated with the Severo Ochoa Program.P.C. conceived the work; M.P.M. performed computational biology analyses; L.P.V. performed initial bioinformatic analysis and all experimental research. P.M.U. and D.L.A. performed 3\u2032RNA-seq library preparation and sequencing. M.W. contributed analytical tools and metadata generation. P.C. and M.P.M. wrote the first draft of the manuscript, which was completed with the assistance of L.P.V. and M.W. All authors approved the final version of the article."}
+{"text": "Scientific Reports 10.1038/s41598-019-50208-x, published online 26 September 2019Correction to: The Acknowledgements section in this Article is incomplete.\u201cWe thank Michael Miller at W.S.I. for discussion.\u201dshould read:\u201cWe thank Michael Miller at W.S.I. for discussion. We gratefully acknowledge Dr. Tyler Maxwell for his contribution to Figure S11c.\u201d"}
+{"text": "The publisher apologizes for the error.The fifth author is listed incorrectly in the citation. The correct citation is: Ljones K, Ness HO, Solvang-Garten K, Gaustad SE, H\u00f8ydal MA (2017) Acute exhaustive aerobic exercise training impair cardiomyocyte function and calcium handling in Sprague-Dawley rats. PLoS ONE 12(3): e0173449."}
+{"text": "Scientific Reports 10.1038/s41598-019-49575-2, published online 17 October 2019Correction to: This Article contains errors in Reference 46 which was incorrectly given as:Odorico, J. S. et al. Extracellular matrix scaffold and hydrogel derived from decellularized and delipidized human pancreas. Sci. Rep. 8, 1\u201316 (2018)The correct reference is listed below as Reference"}
+{"text": "Luminal B cancers show much worse outcomes compared to luminal A. This present study aims to screen key lncRNAs and mRNAs correlated with luminal-B breast cancer.cis nearby-targeted DEmRNAs of DElncRNAs. DElncRNA-DEmRNA co-expression networks were performed. The mRNA and lncRNA expression profiles were downloaded from The Cancer Genome Atlas (TCGA) database to validate the expression patterns of selected DEmRNAs and DElncRNAs.Luminal-B breast cancer tissue samples and adjacent tissue samples were obtained from 4 patients with luminal-B breast cancer. To obtain differentially expressed mRNAs (DEmRNAs) and lncRNAs (DElncRNAs) between luminal-B breast cancer tumor tissues and adjacent tissues, RNA-sequencing and bioinformatics analysis were performed. Functional annotation of DEmRNAs and protein-protein interaction networks (PPI) construction were performed. DEmRNAs transcribed within a 100\u2009kb window up- or down-stream of DElncRNAs were searched, which were defined as A total of 1178 DEmRNAs and 273 DElncRNAs between luminal-B breast cancer tumor tissues and adjacent tissues were obtained. Hematopoietic cell lineage, Cytokine-cytokine receptor interaction, Cell adhesion molecules (CAMs) and Primary immunodeficiency were significantly enriched KEGG pathways in luminal-B breast cancer. FN1, EGFR, JAK3, TUBB3 and PTPRC were five hub proteins of the PPI networks. A total of 99 DElncRNAs-nearby-targeted DEmRNA pairs and 1878 DElncRNA-DEmRNA co-expression pairs were obtained. Gene expression results validated in TCGA database were consistent with our RNA-sequencing results, generally.This study determined key genes and lncRNAs involved in luminal-B breast cancer, which expected to present a new avenue for the diagnosis and treatment of luminal-B breast cancer. Breast cancer is the leading cause of cancer-related death in women, both overall and in less developed countries . It is aWith advances in high-throughput technology, it is discovered that human transcriptome mainly consists of non-coding RNAs (ncRNAs) with limited or no protein-coding capacity , 10. Loncis nearby-targeted DEmRNAs of DElncRNAs and construction of DElncRNA-DEmRNA co-expression networks were performed. In this light, we expect this study could represent a new avenue to improve the understanding of the pathogenesis and be helpful for treatment of luminal B breast cancer.In this study, differentially expressed lncRNAs (DElncRNA) and mRNAs (DEmRNAs) in tumor tissues of patients with luminal-B breast cancer were identified by RNA-sequencing. Subsequently, protein-protein interaction (PPI) networks of DEmRNAs were conducted. Identification of Luminal-B breast cancer tissue samples and adjacent tissue samples were obtained from 4 patients with luminal-B breast cancer in People\u2019s Hospital of Deyang City, which were free of treatment. The detailed characteristics of patients are displayed in Table\u00a0http://www.bioinformatics.babraham.ac.uk/projects/fastqc/), the FASTQ sequence data were acquired from the RNA-sequencing data. Read QC tool in FastQC version 0.11.4 (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/) was used for the quality control of FASTQ data with Q\u2009>\u200930. Trimming of raw data was performed with cutadapt version 1.16 (http://cutadapt.readthedocs.io). Reads with low quality  were removed to obtain the clean reads.According to the manufacturer\u2019s protocol, RNA was extracted with PAXgene blood RNA kit . With Agilent 2100 Bioanalyzer (Agilent RNA 6000 Nano Kit), the concentration, integrity and RNA integrity number (RIN) values of RNA were assessed. Sequencing was performed based on the Illumina Hiseq X-ten platform  with PE150 bp sequencing mode. The sequencing was done with paired-ends and 10G depth. With Base Calling version 0.11.4 (ftp://ftp.ncbi.nlm.nih.gov/genomes/Homo_sapiens), HISAT2 version 2.1.0 (https://ccb.jhu.edu/software/hisat2/index.shtml) was applied. Expression of mRNAs and lncRNAs was normalized and outputted with StringTie version 1.3.3b (http://ccb.jhu.edu/software/stringtie/). Fragments per Kilobase of exon per million fragments mapped (FPKM) of lncRNAs and mRNAs were calculated with StringTie. With edgeR version 3.24 (http://www.bioconductor.org/packages/release/bioc/html/edgeR.html), both DEmRNAs and DElncRNAs were obtained with |log2FC|\u2009>\u20091 and p-value <\u20090.05. By using R package \u201cpheatmap\u201d, hierarchical clustering analysis of DElncRNAs and DEmRNAs were conducted.In order to align the clean reads with the human reference genome, Ensemble GRCh38.p7 (http://metascape.org/gp/index.html). A value of p\u2009<\u20090.05 was set as the cut-off for significance.Functional annotation, including Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses was performed with Metascape (http://www.uniprot.org/database/DB-0184). Then, PPI networks were visualized with Cytoscape software .Top 100 up- and down-regulated DEmRNAs were scanned with the BioGrid .A total of 1178 DEmRNAs (666 up-regulated and 512 down-regulated DEmRNAs) and 273 DElncRNAs (181 up-regulated and 92 down-regulated DElncRNAs) were obtained. The top 10 up- and down-regulated DEmRNAs and DElncRNAs were exhibited in Table\u00a0p\u2009=\u20091.14E-50), cytokine-mediated signaling pathway (p\u2009=\u20095.74E-44) and cytokine production (p\u2009=\u20098.78E-38) were significantly enriched GO terms in luminal-B breast cancer , Cytokine-cytokine receptor interaction (p\u2009=\u20098.19E-21), Cell adhesion molecules (CAMs) (p\u2009=\u20093.61E-18) and Primary immunodeficiency (p\u2009=\u20091.17E-14) were significantly enriched KEGG pathways in luminal-B breast cancer , EGFR (degree\u2009=\u200914), JAK3 (degree\u2009=\u200911), TUBB3 (degree\u2009=\u200911) and PTPRC (degree\u2009=\u200910) were five hub proteins of the PPI networks , regulation of cell-cell adhesion (p\u2009=\u20097.86E-12), cytokine binding (p\u2009=\u20093.41E-07), T cell selection (p\u2009=\u20097.75E-07) and negative regulation of secretion (p\u2009=\u20091.38E-06) were significantly enriched GO terms , Cytokine-cytokine receptor interaction (p\u2009=\u20092.81E-03), Primary immunodeficiency (p\u2009=\u20097.71E-03), AMPK signaling pathway (p\u2009=\u20099.30E-03) and Wnt signaling pathway (p\u2009=\u20091.46E-02) were significantly enriched KEGG pathways , cytokine-mediated signaling pathway (p\u2009=\u20099.35E-28), cytokine production (p\u2009=\u20091.68E-27), leukocyte migration (p\u2009=\u20093.20E-22) and alpha-beta T cell activation (p\u2009=\u20094.89E-20) were significantly enriched GO terms , Hematopoietic cell lineage (p\u2009=\u20099.66E-16), Primary immunodeficiency (p\u2009=\u20092.85E-15), Th1 and Th2 cell differentiation (p\u2009=\u20093.38E-13) and Cell adhesion molecules (CAMs) (p\u2009=\u20091.66E-12) were significantly enriched KEGG pathways  . During Metastasis-associated lung adenocarcinoma transcript\u00a01 (MALAT1) is a highly conserved lncRNA, and its over-expression in multiple cancerous tissues has been linked to the proliferation and metastasis of tumor cells. It was first identified as being up-regulated in lung tumors, and a prognostic marker for metastasis and patient survival in non-small cell lung cancer (NSCLC), specifically in early stages of lung adenocarcinoma . Subsequ+ T cells promotes metastasis in luminal breast cancer  is primarily expressed in heart and fetal brain tissue . DysreguThe Wilms\u2019 tumor 1 (WT1) was first cloned in 1990 as a suppressor in Wilms\u2019 tumor, which was located at chromosome 11p13 . WT1 genWilms tumor 1 Antisense RNA (WT1-AS) is located upstream of the WT1 gene, and these two genes are bi-directionally transcribed from the same promoter region. Down-regulation of WT1-AS was related to a poorer prognosis in ovarian clear cell adenocarcinoma . Lv et aIn conclusion, a total of 1178 DEmRNAs and 273 DElncRNAs between luminal-B breast cancer tumor tissues and adjacent tissues were obtained. We discussed and emphasized the importance role of three DElncRNA-DEmRNA pairs, including MALAT1-S100A7, MIAT-CCL5 and WT1-AS-WT1, involved in luminal B breast cancer, which expected to provide new insight into understanding the mechanism underlying pathogenesis of luminal B breast cancer. The small sample size was a limitation of our study. Although the validation results in TCGA database indicated that our RNA-sequencing results were generally reliable, larger cohorts of patients and further experimental validation studies are needed to conduct to verify this conclusion."}
+{"text": "Oriens lacunosum-moleculare (O-LM) interneurons constitute 40% of hippocampal interneurons expressing Somatostatin (SST). Recent evidence has indicated a dual origin for these cells in the medial and caudal ganglionic eminences (MGE and CGE), with expression of Htr3a as a distinguishing factor. This is strikingly different from cortical SST interneurons that have a single origin within the MGE/preoptic area (POA). We reassessed the origin of hippocampal SST interneurons using a range of genetic lineage-tracing mice combined with single-cell transcriptomic analysis. We find a common origin for all hippocampal SST interneurons in NKX2-1-expressing progenitors of the telencephalic neuroepithelium and an MGE/POA-like transcriptomic signature for all SST clusters. This suggests that functional heterogeneity within the SST CA1 population cannot be attributed to a differential MGE/CGE genetic origin.  \u2022+ progenitors generate all hippocampal CA1 somatostatin (SST) interneuronsNKX2-1\u2022+ cells generated in the absence of NKX2-1 do not migrate to the hippocampusSST\u2022LHX6 expression maintained in MGE-derived cortical and hippocampal CA1 interneurons\u2022Htr3a is detected in MGE-derived hippocampal CA1 interneuronsLow expression of  In this article, Kessaris and colleagues combine genetic lineage tracing and single-cell transcriptomic analysis to identify the embryonic origin of hippocampal CA1 Somatostatin interneurons. They report a common origin in the medial ganglionic eminence/preoptic area from neuroepithelial precursors expressing NKX2-1. The pri the CGE . The pre the CGE .oriens lacunosum-moleculare (O-LM) interneurons, which have their cell bodies within stratum oriens (s.o.) and represent 40% of all SST interneurons in the hippocampus  mapping of Sst interneurons in the developing mouse brain suggested that most, if not all, telencephalic Sst cells originate in the diagonal area (or anterior entopeduncular area [AEP]) and not the pallidal neuroepithelium  (Recent ithelium . This su (E13.5) .Lhx6 to label MGE-derived interneurons in the cortex and hippocampus   B\u20131F. Aro30 (P30) B and 1F.30 (P30)  or from 30 (P30) A (Fogart30 (P30) . Analysi30 (P30) C and 1F,30 (P30) D and 1F.with GFP E and 1F.rneurons .Figure\u00a01Sst interneurons in the developing mouse brain suggested that most, if not all, telencephalic Sst cells originate in the AEP  and that failure to label 30% of cortical interneurons in Nkx2-1-Cre mice may have been caused by lack of NKX2-1 expression in the AEP at E13.5  embryos for the presence of SST-positive cells. At E13.5, most Sst cells were missing from the telencephalon in Nkx2-1 KO embryos compared with controls  \u223c30% of SST cells with typical O-LM characteristics were labeled with GFP in Htr3a-GFP mice; (2) a proportion of s.o. SST O-LM cells were labeled with GFP in Mash1-CreER;RCE transgenic mice following tamoxifen administration; and (3) there was only \u223c10% overlap in expression of tdTomato and GFP in Nkx2-1-Cre;tdTOM;Htr3a-GFP mice 1Wdr (MGI:3761164) ((Nkx6-2-icre)1Kess (MGI:4355562) (JAX 027798), Tg(Lhx6-icre)1Kess (MGI:4355717) (JAX 026555) (tm1(EGFP/cre)Cjt (MGI:3053959) (JAX 005622) (tm2.1(cre)Zjh (MGI:4838416) (JAX 013044) (KI (JAX 006148) (KI reporter mice (JAX 004077)  , Tg(Nkx6 026555) , Shhtm1( 005622) , Ssttm2. 013044) , R26R-YF 006148) , and R26 004077)  have beed allele . All aniLhx6- and Sst DIG-labeled probes were generated from a plasmid (kind gift from V. Pachnis) and IMAGE clone 4218815 (Source Biosciences), respectively.Tissue processing, immunohistochemistry, and ISH were carried out as previously described . Primary+ cells per animal were counted in order to generate the data shown in Images were captured using a Hamamatsu C4742-95 camera on\u00a0a\u00a0Zeiss Axioplan fluorescence microscope and Digital Pixel software. Confocal images were captured on a Leica CTR6500 confocal microscope or a Zeiss LSM880 with Airyscan. ISH images were captured on a Zeiss Axio Scan.Z1 scanner. Image composites were assembled using Microsoft ICE software  and processed with Adobe Photoshop CS6 . Figures were generated in Adobe Illustrator CS6 (Adobe Systems). Quantification was performed as described previously . Three ahttps://github.com/cortexlab/Transcriptomics. To visualize different cell subtypes, we used the negative binomial t-stochastic neighbor-embedding (nbtSNE) algorithm as previously described (The single-cell data presented were obtained by reanalyzing our previously published datasets . The codescribed .Conceptualization, Z.A. and N. Kessaris; Methodology, N. Kessaris and K.D.H.; Investigation, Z.A., L.M., N. Ktena, K.D.H., and N. Kessaris; Writing \u2013 Original Draft, Z.A. and N. Kessaris; Writing \u2013 Review\u00a0& Editing, Z.A., L.M., K.D.H., and N. Kessaris; Funding Acquisition, K.D.H. and N. Kessaris."}
+{"text": "We observed that both EBV- and HCMV-specific T cell responses were significantly lower in SLE patients compared with healthy subjects. We reported decreased EBV- and HCMV-specific T cell responses among medium-high immunosuppressed patients compared to low immunosuppressed patients. Immunosuppressive level could exert a role in the control of herpesviruses reactivation, even if the immunosuppressive condition of SLE remains the driving cause of skewed virus-specific T cell response.Systemic lupus erythematosus (SLE) is an autoimmune disease with a complex etiology. Opportunistic viral pathogens, such as human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV), are particularly relevant. The role of the T cell response in SLE has not been deeply studied; we investigated the role of HCMV- and EBV-specific T cell responses in SLE patients also in relation to their pharmacological immunosuppressive status. PBMCs from 70 SLE patients and 50 healthy controls were stimulated with EBV- and HCMV-specific antigens, and IFN- Herpesviridae family, and they may complicate the disease course [\u03b3) is also suggested to play a crucial role in this context [Systemic lupus erythematosus (SLE) is a complex pathological condition  which mae course \u201315 or mie course . Furthere course . It has e course , 23, whie course . In this context .\u03b3 ELISPOT assay. By using a novel approach we provided a good estimation of both CD4+ and CD8+ antigen-specific T cell responses, avoiding predepletion assay [+ and CD8+ antigen-specific T cell response, avoiding the intracellular cytokine staining approach that is labor intensive and requires a larger number of cells. However, this approach cannot be considered as precise as flow cytometry strategy, but could represent an easier way for the estimation of antigen-specific T cell response. For comparison, T cell response to the nonspecific mitogen (PHA) was also investigated.The aim of this study was to evaluate and characterize T cell responses to HCMV and EBV in SLE patients using IFN-on assay . In thisSeventy patients  38.0-57.8) fulfilling the 1997 ACR classification criteria for SLE  and refeSLE patients had a median age at disease onset of 30 (IQR 23-46) years and a median disease duration of 121.5 (IQR 42.3-228.5) months. In all cases, disease activity was evaluated according to SLEDAI 2k score [For practical purposes, we divided the patients into two groups, according to the degree of pharmacological immunosuppression: patient treatment with hydroxychloroquine and/or with prednisone\u2009\u2264\u20095\u2009mg/day was considered low pharmacological immunosuppression . Patient treatment with mycophenolate mofetil, methotrexate, cyclosporin A, rituximab, belimumab, and/or prednisone\u2009>\u20095\u2009mg/day was considered medium-high pharmacological immunosuppression .\u03bcg/ml streptomycin, 25% human albumin , and 10% DMSO ), and stored in liquid nitrogen (10\u2009\u00d7\u2009106 cells/ml) until analysis. After thawing, about 50-60% of cells were still viable and could be used in the ELISPOT assay.Peripheral blood was collected into vacutainer tubes (BD) containing heparin. Whole blood was used for viral genome quantification and determination of T cell subsets; plasma was separated for serological analyses. Peripheral blood mononuclear cells (PBMCs) were isolated by density gradient centrifugation , cryopreserved in freezing medium , IgG anti-VCA, and anti-EBNA were quantified using an ELISA kit , according to the manufacturer's instructions. Healthy subjects and SLE patients were considered EBV-seropositive when IgG anti-VCA and anti-EBNA were positive.IgM anti-HCMV and IgG anti-HCMV were quantified using an ELISA kit , according to the manufacturer's instructions. Healthy subjects and SLE patients were considered HCMV-seropositive when IgG anti-HCMV was positive.DNA was purified using NucliSENS\u00aeeasyMAG\u00ae . EBV DNA and HCMV DNA were quantified using real-time PCR (lower limit detection HCMV and EBV DNA 90 copies/ml), as previously described  were determined by flow cytometry , using Flow-Count Fluorospheres. Gating strategy was set up on CD45+ and side scatter (SSC).Fresh whole blood was stained with anti-CD3-PC5, anti-CD45-FITC, anti-CD4-RD1, and anti-CD8-ECD monoclonal antibodies . After lysis of red blood cells, absolute CD3\u03bcg/ml streptomycin with 8% of DMSO. Resuspended peptide pools were used after dilution (1\u2009:\u2009100) in RPMI 1640 supplemented with 2\u2009mM L-glutamine, 100\u2009U/ml penicillin and 100\u2009\u03bcg/ml streptomycin, and 10% FBS and then used as antigens. A lytic pool, containing peptides spanning the full length of BZLF-1 (59 peptides) and BMRF-1 (99 peptides) EBV proteins; an Epstein-Barr nuclear antigen (EBNA) pool, containing peptides spanning the full length of EBNA 1 (158 peptides), EBNA3a (234 peptides), EBNA 3b (279 peptides), and EBNA 3c (265 peptides) EBV proteins; and a latent membrane protein (LMP) pool, containing peptides spanning the full length of LMP1 (94 peptides) and LMP2 (122 peptides) EBV proteins, were used as EBV-specific antigens  at a final concentration of 0.25\u2009\u03bcg/ml for each individual peptide in the corresponding pool. Peptide pools representative of whole HCMV proteins IE-1 (120 peptides), IE-2 (143 peptides), and pp65 (138 peptides) (JPT Peptide Technologies) were used at a final concentration of 0.25\u2009\u03bcg/ml for each individual peptide in the corresponding pool.Lyophilized peptide pools, 15 amino acids in length with an 11 amino acid overlap, were resuspended in RPMI 1640 supplemented with 2\u2009mM L-glutamine, 100\u2009U/ml penicillin, and 100\u2009\u03b3 ELISPOT kits  and Multiscreen-IP membrane-bottomed 96-well plates  were used as described [\u03b3 and stored at 4\u00b0C. After washing with PBS, plates were blocked with culture medium  (Euroclone)) for 2 hours at room temperature. Cells were plated in duplicate (1 \u00d7 105/100\u2009\u03bcl per well) and stimulated with the corresponding antigens or with phytohemagglutinin  or with medium alone (negative control) and incubated at 37\u00b0C in a 5% CO2 humidified atmosphere for 24 hours. After washing, plates were incubated overnight at 4\u00b0C with biotinylated IFN-\u03b3 detection antibody. Plates were washed, streptavidin-alkaline phosphatase conjugate was added, and plates were incubated at 37\u00b0C in a 5% CO2 atmosphere for 1 hour. Plates were then washed, and 5-bromo-4-chloro-3-indolyl phosphate/nitro blue tetrazolium (BCIP/NBT) was added for 20 minutes at room temperature. Wells were then washed several times under running water and air-dried overnight. Spots were counted by using an automated AID ELISPOT reader system . The mean number of spots from duplicate wells was adjusted to 1 \u00d7 106 PBMCs. The net spots per million PBMCs was calculated by subtracting the number of spots responding to the negative control from the number of spots responding to the corresponding antigenand results were given as net spots/million PBMCs. Furthermore, results were normalized to absolute CD4+ and CD8+ T cell counts, as previously described [Human IFN-escribed \u201331. Briet-test or Mann-Whitney test was used for quantitative variables in order to perform comparison between groups. Spearman's test was used for the correlation analysis. All tests were two-tailed. A P value\u2009<\u20090.05 was considered statistically significant. Analyses were performed using the GraphPad Prism 5 .Descriptive data were reported or considered as absolute and relative frequencies, mean and standard deviation, median, and interquartile range (IQR) based on the type of the variable distribution. For qualitative variables, Fisher's test was used, while + and CD8+ T cells were significantly lower in SLE patients than in healthy controls (P < 0.0001). In SLE patients, the median CD4+ and CD8+ T cell counts were 434.5 (IQR 270.3-620.3) cells/\u03bcl and 287.0 (IQR 196.0-397.3) cells/\u03bcl, respectively, while in healthy subjects, the median CD4+ and CD8+ T cell counts were 1054.0 (IQR 760.5-1361.0) cells/\u03bcl and 532.0 (IQR 376.8-748.8) cells/\u03bcl, respectively. EBV- and HCMV-specific antibodies were analyzed in all enrolled subjects, as well as HCMV and EBV DNAemia. Demographic data are summarized in T cell subsets were analyzed in 70 SLE patients and 50 healthy subjects with homogeneous characteristics. The two groups were not substantially different in terms of age and sex distribution. Both CD4+ and CD8+ T cell responses to all EBNA, LMP, and lytic overlapping 15-mer peptide pools were evaluated in 68 EBV-seropositive SLE patients, 25 EBV-seropositive healthy subjects, and four EBV-seronegative individuals .EBV-specific CD4P = 0.0002)  vs 1.140 (IQR 0.5235-2.6180) EBV-specific CD4+ T cells/\u03bcl EBV-specific CD4+ T cells/\u03bcl; P < 0.0001) (+ T cell response was significantly different in the two groups of subjects (0.0905 (IQR 0.0215-0.2148) and 0.5620 (IQR 0.2335-1.231) EBV-specific CD8+ T cells/\u03bcl, respectively, P < 0.0001) (+ and CD8+ T cell responses with the SLEDAI 2k score in SLE patients (data not shown).Median EBV-specific T cell response was significantly lower in EBV-seropositive SLE patients than in EBV-seropositive healthy subjects (340.5 (IQR 75-738.8) vs. 890 (IQR 360-1983) net spots/million PBMCs;  0.0002) . Normali 0.0001) . Similar 0.0001) . There wn = 25, 36.8%) or mhp-IS . The latter had significantly lower EBV-specific T cell response than the other group (170 (IQR 70-530) and 500 (IQR 237.5-905.0) net spots/million PBMCs, respectively; P = 0.0224) (+ and CD8+ T cells. The EBV-specific CD4+ T cell response was lower in mhp-IS than in lp-IS (0.0620 (IQR 0.0170-0.2680) and 0.2420 (IQR 0.1065-0.4515) EBV-specific CD4+ T cells/\u03bcl; P = 0.0124) (+ T cell response in SLE patients receiving mhp-IS was lower than that observed in patients receiving lp-IS (0.1430 (IQR 0.0800-0.3335) and 0.0450 (IQR 0.0180-0.1640) EBV-specific CD8+ T cells/\u03bcl; P = 0.0174)  . Results 0.0124) . Similar 0.0174) .+ and CD8+ T cell EBV-specific ELISPOT response in SLE patients with undetectable EBV DNA  and detectable EBV DNA . A trend toward statistical significance was observed when we compared the EBV-specific T cell response measured as net spots/million PBMCs in SLE patients with undetectable (median 387.5 (IQR 119.8-953.8)) and detectable EBV DNA (220 (IQR 56.3-538.8); P = 0.0753) (+ T cells/\u03bcl between the two groups (0.202 IQR (0.038-0.450) and 0.072 IQR (0.016-0.236) EBV-specific CD4+ T cells/\u03bcl; P = 0.0773)  EBV-specific CD8+ T cells/\u03bcl and 0.0710 IQR (0.0157-0.1565) EBV-specific CD8+ T cells/\u03bcl; P = 0.1070)  . Similar 0.0773) , althoug 0.1070) .P = 0.0373) (+ T cell response in mhp-IS patients (median 0.040 IQR (0.013-0.160) EBV-specific CD4+ T cells/\u03bcl) was compared with lp-IS patients (median 0.223 IQR (0.097-0.398) EBV-specific CD4+ T cells/\u03bcl; P = 0.0573) (+ T cell response in mhp-IS (0.034 IQR (0.011-0.120) EBV-specific CD8+T cells/\u03bcl and lp-IS patients (0.097 IQR (0.080-0.210) EBV-specific CD8+ T cells/\u03bcl; P = 0.0394)  were receiving mhp-IS and nine (30%) lp-IS. In a comparison of the two groups, EBV-specific T cell response measured as net spots/million PBMCs was higher in lp-IS group (median 110 IQR (37.5-460)) than in mhp-IS (median 380 IQR (313-750);  0.0373) . A trend 0.0573) . A signi 0.0394) . No diffP = 0.3679)  vs 0.6065 (IQR 0.1368-1.250) HCMV-specific CD4+ T cells/\u03bcl; P = 0.0004) (+ T cell response was significantly different in the two groups of subjects (0.6490 (IQR 0.1480-1.280) vs 2.279 (IQR 0.7108-3.570) HCMV-specific CD8+T cells/\u03bcl; P = 0.0059). Both groups of HCMV-seropositive subjects  had HCMV-specific responses higher than HCMV-seronegative controls (P < 0.0001) . Seventeen out of 55 (30.9%) SLE patients were receiving lp-IS, while the remaining 38 (69.1%) were receiving mhp-IS. The HCMV-specific ELISPOT response measured as net spots/million PBMCs was significantly higher in lp-IS patients (median 2695 (IQR 1003-6525)) than in mhp-IS patients (median 1055 (IQR 271-1810)) (P = 0.0282) (+ T cell response was significantly reduced in this mhp-IS patients (0.271 IQR (0.125-1.083) vs 0.927 IQR (0.611-2.315) HCMV-specific CD4+ T cells/\u03bcl; P = 0.0179) (+ T cell response between mhp- and lp-IS (0.359 IQR (0.071-0.738) vs 0.910 IQR (0.383-2.090) HCMV-specific CD8+ T cells/\u03bcl; P = 0.0111) . HCMV-specific T cell response was lower in HCMV-seropositive SLE patients than in HCMV-seropositive healthy subjects (median 1155 (IQR 415-3925) and 1850 (770-3103) net spots/million PBMCs, respectively), but the difference was not statistically significant ( 0.3679) . Results 0.0004) . Similar 0.0001) . There w 0.0282) . Similar 0.0179) . Finally 0.0111) .We observed that ELISPOT T cell response in the 52 SLE patients with undetectable HCMV DNA was higher than the response observed in the three patients with detectable HCMV DNAemia (data not shown). However, due to the low number of patients in the latter group, the difference was not considered statistically valid.All patients with detectable HCMV DNA were medium-high immunosuppressed. Two of them were also positive for EBV DNA.+ and CD8+ T cells producing IFN-\u03b3 in response to PHA, a nonspecific antigen, were significantly lower in 70 SLE patients than in 41 healthy controls. PHA-specific T cell responses measured as net spots/million PBMCs were 4795 (IQR 2956-7193) and 6915 (5449-8165) (P = 0.0140), respectively,  and 6.890 (IQR 4.542-10.89) PHA-specific CD4+ T cells/\u03bcl (P < 0.0001), respectively,  and 3.859 (IQR 2.199-5.700) PHA-specific CD8+T cells/\u03bcl (P < 0.0001)  . There w+ and CD8+ T cells producing IFN-\u03b3, as previously described [\u03b3 ELISPOT is one of the most widespread assays used to evaluate antigen-specific T cell response. It allows the evaluation of total CD4+ and CD8+ antigen-specific T cell response by detection of IFN-\u03b3-producing T cells. As known, IFN-\u03b3 is a cytokine mainly produced by activated T helper 1 (Th1) and T cytotoxic cells in response to specific antigens, having a crucial role inactivating lymphocytes to enhance antimicrobial and antitumor effects. The traditional ELISPOT assay does not easily distinguish between CD4+ and CD8+ T cell responses unless separate antigens to detect CD4+ and CD8+ T cell responses or lymphocyte subset depletion are used [+ and CD8+ T cells [+ and CD8+ T cell counts, in order to estimate both CD4+ and CD8+ T cells producing IFN-\u03b3, in response to different viral antigens. Even if it does not allow a precise quantification of CD4+ and CD8+ antigen-specific T cell response, our previous results evidenced a good correlation between intracellular cytokine staining and normalized ELISPOT approach [The aim of this study was to evaluate EBV- and HCMV-specific T cell responses by ELISPOT assay. In this setting, we used the normalization on CD4 and CD8 T cells in order to provide an estimation of CD4escribed . As our are used . EBV- an T cells , 34. Resapproach .Among viral pathogens, EBV infection is the most common in patients with SLE, and more importantly, it has been hypothesized that EBV may play a role in SLE disease induction . In a st+ and CD8+ T cell responses to EBV-specific peptide pools were significantly lower in SLE patients compared with controls. Berner et al. demonstrated that the frequency of EBV-specific CD8+ T cells was similar between SLE and healthy subjects when analyzed by using MHC I tetramers with a lytic cycle EBV antigen peptide. However, CD8+ T cell EBV-specific IFN-\u03b3 production was significantly lower in SLE patients than in healthy controls, when assayed by ELISPOT [+ T cell EBV-specific response was provided. In contrast with our results, Kang et al. showed an increased frequency of EBV-specific CD69+ CD4+ T cells producing IFN-\u03b3 in SLE patients compared with controls. However, the frequency of EBV-specific CD69+ CD8+ T cells producing IFN-\u03b3 tended to be lower in SLE patients [Both CD4 ELISPOT . Neverthpatients . These d\u03b3 production upon stimulation with EBNA 1 or EBV early antigen diffuse (EBV-EA/D) in SLE patients, suggesting the decreased control of EBV infection in these subjects [Similarly, Draborg et al. showed a significantly reduced number of activated T cells and IFN-subjects . InteresA few studies \u201339 have + CD4+ T cells producing IFN-\u03b3 and TNF-\u03b1 was similar in SLE patients and healthy controls, while the frequency of HCMV-specific CD69+ CD8+ T cells producing IFN-\u03b3 and TNF-\u03b1 was lower in SLE patients, although the difference was not statistically significant [+ and CD8+ T cell IFN-\u03b3-producing cells.Larsen et al. did not find immune alterations in HCMV-specific T cell responses in SLE patients compared with controls . Anothernificant . On the Analyzing HCMV-specific T cell response in SLE patients according to immunosuppression level, we do not have the evidence of the difference between lp-IS SLE patients and healthy HCMV-seropositive subjects, while mhp-IS patients showed a markedly reduced HCMV-specific T cell response if compared to lp-IS patients as well as to healthy HCMV-seropositive subjects. This could support the role of immunosuppressive treatment in the control of HCMV reactivation.Interestingly, in our study, a significantly lower T cell response to the specific PHA antigen in SLE patients compared with healthy subjects was demonstrated, while other authors have not shown any differences , 38, sug+ and CD8+ T cell responses in SLE patients treated with medium-high immunosuppression. Similarly, HCMV-specific CD4+ and CD8+ T cells had reduced IFN-\u03b3 production in medium-high immunosuppressed SLE patients. On the contrary, there was no difference in terms of PHA-specific T cell response between low and medium-high immunosuppressed SLE patients.In contrast with other studies , 39, we The role of immunosuppressive therapy appears to be crucial mainly to the response to EBV but also to HCMV-specific T cell response. Furthermore, a generally deficient immune response in SLE patients with respect to healthy controls was observed, supporting the hypothesis that SLE disease exerts a general immunosuppressive action regardless of the therapy. To corroborate this hypothesis, virus-specific T cell responses need to be analyzed in a larger number of patients, in order to stratify patients according to immunosuppressive status and iatrogenic risk factors."}
+{"text": "The near-complete genome sequence of foot-and-mouth disease virus (FMDV) serotype A potential vaccine strain BAN/CH/Sa-304/2016 is reported here. Its genome revealed antigenic heterogeneity with the current Indian vaccine strain IND40/00, with four amino acid substitutions in antigenically critical sites of the VP1 protein. The near-complete genome sequence of foot-and-mouth disease virus (FMDV) serotype A potential vaccine strain BAN/CH/Sa-304/2016 is reported here. Its genome revealed antigenic heterogeneity with the current Indian vaccine strain IND40/00, with four amino acid substitutions in antigenically critical sites of the VP1 protein. Aphthovirus in the family Picornaviridae. FMDV exists as seven immunologically distinct serotypes  with multiple topotypes, lineages, and sublineages (Foot-and-mouth disease (FMD) is a highly contagious disease of domestic and wild cloven-hoofed animals worldwide . Foot-anlineages . Among alineages . All thelineages , 5.r > 0.3)  . The serr > 0.3) . The ampr > 0.3) .The near-complete genome of the vaccine candidate BAN/CH/Sa-304/2016 is 8,221 nucleotides (nt) in length, consisting of 53.49% GC content, with a 5\u2032 untranslated region (5\u2032 UTR), coding sequence (CDS), and 3\u2032-UTR region of 1,101, 6,999, and 121 nt, respectively. The CDS codes for a polyprotein 2,332 amino acids (aa) long, containing four structural proteins, VP1 to -4, and eight nonstructural proteins , while the 5\u2032-UTR poly(C) tract and 3\u2032-UTR poly(A) tail are 16 nt and 25 nt long, respectively. Phylogenetically, the isolates BAN/CH/Sa-304/2016 and BAN/CH/Sa-302/2016  clustered in a separate clade closely related to other isolates from Bangladesh and India within genotype VII of the Asia topotype .KJ754939) and 91% with Indian vaccine strain IND40/00 (GenBank accession no. HM854025). The VP1, VP2, and VP3 protein sequences of BAN/CH/Sa-304/2016 had 8%, 6%, and 5% variability, with 16, 12, and 9 aa substitutions compared to the respective regions of vaccine strain IND40/00. Only 4 aa substitutions  of VP1 were in antigenically critical sites (59 position of BAN/CH/Sa-304/2016.The homology search in GenBank using BLAST showed that BAN/CH/Sa-304/2016 shares 96% nucleotide similarity with previous serotype A isolate BAN/GA/Sa-197/2013 (GenBank accession no. al sites . One amiWe report here the near-complete genome sequence of FMDV serotype A potential vaccine strain BAN/CH/Sa-304/2016 for FMD control in Bangladesh.MK088171.The near-complete genome sequence of FMDV serotype A strain BAN/CH/Sa-304/2016 has been deposited in the NCBI GenBank database under the accession no."}
+{"text": "Trypanosoma equiperdum primarily parasitizes the genital organs and causes dourine in equidae. We isolated a new T. equiperdum strain, T. equiperdum IVM-t1, from the urogenital tract of a horse definitively diagnosed as having dourine in Mongolia. Trypanosoma equiperdum primarily parasitizes the genital organs and causes dourine in equidae. We isolated a new T. equiperdum strain, T. equiperdum IVM-t1, from the urogenital tract of a horse definitively diagnosed as having dourine in Mongolia. Here, we report the whole-genome sequence, the predicted gene models, and their annotations. Trypanosoma equiperdum belongs to the kingdom Protista, phylum Sarcomastigophora, class Zoomastigophorea, order Kinetoplastida, family Trypanosomatidae, genus Trypanosoma, subgenus Trypanozoon together with T. brucei and T. evansi   , EthiopiICT2011) , and Ven in 2005 , 2015 (7T. equiperdum strain IVM-t1 isolate was cultivated using Hirumi's modified Isocove's medium-9 (HMI-9) soft-agarose medium  at 37\u00b0C in 5% CO2 (T. equiperdum IVM-t1 was extracted and purified using Tris-EDTA (TE)-saturated phenol (Sigma-Aldrich Japan) and phenol-chloroform isoamyl-alcohol solution (Sigma-Aldrich Japan) , a DNA/polymerase binding kit P6 v2, and a DNA sequencing kit 4.0 v2 for PacBio RS II sequencing . The MiSeq sequencer produced 28,337,050 paired-end reads with an average read length of 300\u2009bp, and the PacBio RS II sequencer produced 7,508 reads with an average read length of 8,523\u2009bp . The low-quality reads were trimmed using FastQC v0.11.5 and FASTX-Toolkit v0.0.13.The h Japan) . PurifieT. equiperdum by Metassembler v1.5 (T. equiperdum IVM-t1 were performed using the published genome of T. brucei brucei (T. b. brucei) TREU927 as a reference via the Companion pipeline (https://companion.sanger.ac.uk/) (Whole-genome assembly was performed with ABySS-2.0.2  with parler v1.5  with the.ac.uk/)  with defN50 value of 859,849\u2009bp and a cumulative length of 26,988,997\u2009bp (\u224827 Mbp). The T. equiperdum IVM-t1 draft genome contains 7,718 protein-coding genes, 102 noncoding genes, and 2,473 pseudogenes. Following comparison of the predicted genes of T. equiperdum IVM-t1 with the reference gene sets of T. b. brucei TREU927 using OrthoMCL v2.0.7 (T. equiperdum IVM-t1-specific protein-coding genes were identified. Seven short genes that have less than 25 amino acids were not included in the comparison (https://doi.org/10.6084/m9.figshare.7552262.v1).The integrated draft genome consisted of 45 contigs with an L v2.0.7 , 6,831 pT. equiperdum-specific genes.In conclusion, the whole-genome draft assembly produced in the present study provides a resource for future trypanosome genetic studies and identifies some QSBY00000000. The version described in this paper is the first version, QSBY01000000. Raw sequence reads have been deposited in the NCBI SRA database under the accession no. PRJNA477427.This whole-genome project has been deposited in DDBJ/ENA/GenBank under the accession no."}
+{"text": "N-methyl-d-aspartate/glutamate receptor (NMDAR) is one of the major voltage-sensitive ligand-gated cation channel. Several noncompetitive NMDAR antagonists contribute to pathophysiology of schizophrenia and mood disorders; however, the effects of inhibition of NMDAR on several transmitter system have not been well clarified. Thus, this study determined the selective NMDAR antagonist, MK801 (dizocilpine), on thalamocortical, mesothalamic, and mesocortical transmissions associated with l-glutamate, GABA, serotonin, norepinephrine, and dopamine using multiprobe microdialysis. Perfusion with MK801 into the medial prefrontal cortex (mPFC) increased and decreased respective regional releases of monoamine and GABA without affecting l-glutamate. The mPFC MK801-induced monoamine release is generated by the regional GABAergic disinhibition. Perfusion with MK801 into the reticular thalamic nucleus (RTN) decreased GABA release in the mediodorsal thalamic nucleus (MDTN) but increased releases of l-glutamate and catecholamine without affecting serotonin in the mPFC. The RTN MK801-induced l-glutamate release in the mPFC was generated by GABAergic disinhibition in the MDTN, but RTN MK801-induced catecholamine release in the mPFC was generated by activation of \u03b1-amino-3-hydroxy-5-methyl-4-isoxazolepropionate/glutamate receptor (AMPAR) which received l-glutamate release from thalamocortical glutamatergic terminals in the mPFC. Perfusion with MK801 into the dorsal raphe nucleus (DRN) decreased GABA release in the DRN but selectively increased serotonin release in the MDTN and mPFC. These DRN MK801-induced serotonin releases in the both mPFC and MDTN were also generated by GABAergic disinhibition in the DRN. These results indicate that the GABAergic disinhibition induced by NMDAR inhibition plays important roles in the MK801-induced releases of l-glutamate and monoamine in thalamic nuclei and cortex. N-methyl-d-aspartate/glutamate receptor (NMDAR) modulation has been a source of scientific discussion in psychiatry and psychopharmacology, since various clinical studies have demonstrated that a noncompetitive NMDAR antagonist, ketamine contributes to pathophysiology of schizophrenia and mood disorders   = 20.9 (p < 0.05), FTime = 43.7 (p < 0.05), FMK801*Time = 36.4 (p < 0.05)]  = 18.8 (p < 0.05), FTime = 32.0 (p < 0.05), FMK801*Time = 25.9 (p < 0.05)]  = 7.4 (p < 0.05), FTime = 56.2 (p < 0.05), FMK801*Time = 22.2 (p < 0.05)] (l-glutamate level in the mPFC (Perfusions with MK801 (1 and 5 \u03bcM) into the mPFC increased regional extracellular levels (mPFC MK801-induced release) of serotonin [F< 0.05)] C, norepi< 0.05)] D and dop< 0.05)] E, but de< 0.05)] B withoutthe mPFC A. Extracthe mPFC C\u2013E; howethe mPFC B. Therefl-glutamate, GABA, serotonin, norepinephrine, or dopamine  = 11.4 (p < 0.05), FTime = 74.3 (p < 0.05), FMK801*Time = 17.0 (p < 0.05)])  = 9.1 (p < 0.05), FTime = 65.2 (p < 0.05), FMK801*Time = 12.1 (p < 0.05)]  = 7.3 (p < 0.05), FTime = 47.1(p < 0.05), FMK801*Time = 5.3(p < 0.05)] ]) C, norepi< 0.05)] D and dop< 0.05)] E without< 0.05)] B. These l-glutamate release in the mPFC    = 16.4 (p < 0.05), FTime = 46.7 (p < 0.05), FMK801*Time  = 18.5 (p < 0.05)]  = 9.0 (p < 0.05), FTime = 49.2 (p < 0.05), FMK801*Time = 21.3 (p <0.05)]  into the RTN increased extracellular levels of < 0.05)] A, norepi< 0.05)] D, and do <0.05)] E without <0.05)] B,C. Extr <0.05)] D,E; howe\u03bcM MK801 A. Therefl-glutamate   = 7.2 (p < 0.05), FTime = 21.3 (p < 0.05), FMK801*Time = 7.5 (p < 0.05)]  = 9.0 (p < 0.05), FTime = 73.7 (p < 0.05), FMK801*Time = 12.6 (p < 0.05)]  into the RTN increased extracellular levels of < 0.05)] A and ser< 0.05)] C, and de< 0.05)] B without< 0.05)] D. The exl-glutamate level was increased by 5 \u03bcM MK801, but not by 1 \u03bcM MK801  and perampanel (AMPAR antagonist) into the MDTN and mPFC on transmitter releases in the mPFC and MDTN induced by 50 \u03bcM MK801 perfusion into the RTN .The specific noradrenergic, dopaminergic and serotonergic terminals from respective LC (locus coeruleus), VTA  and DRN  project to deeper layers of frontal cortex ,26,33,37A-R agonist)  and 1 \u03bcM perampanel (AMPAR antagonist)  into MDTN reduced 50 \u03bcM RTN MK801-induced l-glutamate release in the mPFC  = 7.5 (p < 0.05), FTime = 65.2 (p < 0.05), FPER*Time = 8.5 (p < 0.05)] and mPFC   = 11.6 (p <0.05), FTime = 48.1 (p <0.05), FMus*Time = 17.8 (p <0.05)], but perfusion with 1 \u03bcM muscimol into the mPFC did not affect  = 5.0 (p < 0.05), FTime = 107.2 (p < 0.05), FPER*Time = 9.8 (p < 0.05)] and mPFC   = 5.3 (p < 0.05), FTime = 87.2 (p < 0.05), FMus*Time = 22.7 (p < 0.05)], but perfusion with muscimol into the mPFC did not affect (Contrary to < 0.05)] D. Perfust affect D. Simila< 0.05)] E. Perfust affect E.l-glutamate release in the mPFC is generated by the GABAergic disinhibition and relatively activation of AMPAR in the MDTN, but is not modulated by these receptors in the mPFC. Contrary to l-glutamate, RTN MK801-induced catecholamine release in the mPFC is regulated by the GABAergic disinhibition in the MDTN, and activation of AMPAR in the MDTN and mPFC, but is not modulated by GABAA-R in the mPFC. Therefore, RTN MK801-induced catecholamine release in the mPFC is activated by AMPAR in the mPFC via activation of thalamocortical glutamatergic transmission.These results suggest that RTN MK801-induced A-R agonist) into MDTN reduced 50 \u03bcM RTN MK801-induced l-glutamate release in the MDTN, whereas perfusion with 1 \u03bcM perampanel (AMPAR antagonist) into the MDTN did not affect  was regulated by independent system compared with other catecholamine, since serotonin release in the mPFC is not affected by thalamocortical glutamatergic transmission, but conversely mesothalamic serotonergic transmission possibly affects thalamocortical glutamatergic pathway in the MDTN  . To clarMK801 = 16.8 (p < 0.05), FTime = 42.9 (p < 0.05), FMK801*Time = 16.1 (p < 0.05)]  into the DRN concentration-dependently increased extracellular serotonin level in the mPFC [F< 0.05)] C withoutdopamine A,B,D,E. dopamine C. TherefMK801 = 41.9 (p < 0.05), FTime = 63.8 (p < 0.05), FMK801*Time = 24.0 (p < 0.05)] ] C withoutdopamine A,B,D,E. MK801 = 11.6 (p < 0.05), FTime = 26.0 (p < 0.05), FMK801*Time = 6.3 (p < 0.05)]  into the DRN concentration-dependently increased extracellular serotonin level in the MDTN [F< 0.05)] C withoutdopamine A,B,D. Exdopamine C. Our prMK801 = 21.5 (p < 0.05), FTime = 36.3 (p < 0.05), FMK801*Time = 6.1 (p < 0.05)] ] C. TherefMK801 = 13.6 (p < 0.05), FTime = 53.3 (p < 0.05), FMK801*Time = 15.8 (p < 0.05)]  = 9.9 (p < 0.05), FTime = 42.9 (p < 0.05), FMK801*Time = 12.1 (p < 0.05)]  into the DRN concentration-dependently increased and decreased extracellular levels of serotonin [F< 0.05)] C and GAB< 0.05)] B in the  the DRN A,D,E. Ex the DRN B,C. TherMK801 = 22.0 (p < 0.05), FTime = 42.0 (p < 0.05), FMK801*Time = 6.1 (p < 0.05)] ] C, wherea< 0.05)] B. These The threshold concentration of local administration (perfusion) of MK801 into the mPFC, MDTN, RTN, and DRN on several transmission systems demonstrated by this study and previous reports ,16,17,20l-glutamate, GABA, serotonin, norepinephrine and dopamine in the mPFC, MDTN and DRN. According to the results in this study and published neural circuits [The present study also demonstrated the presence of several regulatory systems in the thalamocortical (RTN-MDTN-mPFC) glutamatergic, mesothalamic (DRN-MDTN), and mesocortical (DRN-mPFC) serotonergic pathways controlling releases of circuits ,38,39,40l-glutamate release [A-R in the mPFC (perfusion with 1 \u03bcM muscimol into the mPFC). Therefore, inhibition of NMDAR in the mPFC increases monoamine release induced by presynaptic GABAergic disinhibition in the mPFC. Indeed, the threshold concentrations of local administration of MK801 into the mPFC on GABA release (1 \u03bcM) is more sensitive compared with that of monoamine (5 \u03bcM)  .l-glutamate, norepinephrine and dopamine in the mPFC without affecting those of GABA or serotonin. The threshold concentration of local administration of MK801 into the RTN on releases of l-glutamate and catecholamine (norepinephrine and dopamine) in the mPFC were 1 \u03bcM and 5 \u03bcM, respectively. The RTN MK801-induced l-glutamate release in the mPFC was inhibited by the activation of GABAA-R and inhibition of AMPAR in the MDTN, but was not affected by the activation of GABAA-R or inhibition of AMPAR in the mPFC. These results suggest that an activation of glutamatergic neuronal activity induced by GABAergic disinhibition in the MDTN contributes to RTN MK801-induced l-glutamate release in the mPFC.Contrary to intra mPFC regulation system, local administration of MK801 into the RTN increased releases of l-glutamate release in the mPFC is thalamocortical (from MDTN to mPFC) glutamatergic pathway, but generating mechanisms is GABAergic disinhibition from RTN to MDTN through NMDAR inhibition in the RTN.Various thalamic nuclei, which receive GABAergic inhibition from RTN , projectl-glutamate release, the RTN MK801-induced catecholamine release in the mPFC was inhibited by the activation of GABAA-R in the MDTN, inhibition of AMPAR in the MDTN and mPFC, but was not affected by the activation of GABAA-R in the mPFC. Electrophysiological study demonstrated that electrical stimulation of the MDTN increased the releases of glutamate and catecholamine in the mPFC without affecting those of serotonin [Contrary to erotonin . Other lerotonin ,24,40. Terotonin ,24. TakeThis study indicates the several specific regulation systems of serotonergic transmission in the mPFC and MDTN which are independent upon catecholamine release regulation system. The first, intra mPFC regulation of serotonergic transmission associated with NMDAR is resembling to the regulation systems of catecholaminergic transmission, since the selective noradrenergic, dopaminergic and serotonergic terminals in the deeper layers of mPFC receive GABAergic inhibition. However, the serotonergic terminal in the mPFC from DRN does not contact with thalamocortical glutamatergic afferents, whereas mesothalamic serotonergic transmission activates thalamocortical glutamatergic transmission, since an activation of serotonergic neuronal activities in the DRN increases serotonin release in the MDTN. Recent multiprobe microdialysis studies demonstrated that an activation of serotonergic neuronal activity enhances MDTN glutamatergic neurons through activation of excitatory 5-HT7 receptor in the MDTN ,16. In tA-R predominantly, as indicated by decreased regional serotonin release following local administration of muscimol (GABAA-R agonist) into the DRN; however, GABA release is regulated by excitatory 5-HT7 receptor, reflecting decreased regional GABA release following local perfusions with SB269970 (5-HT7R antagonist) into the DRN [A-R and 5-HT7R predominantly inhibit serotonergic neurons and enhance GABAergic neurons, respectively. In the present study, local administration of MK801 into the DRN, decreased and increased releases of GABA and serotonin in the DRN, respectively. The opposite effects of MK801 between releases of GABA and serotonin were inhibited by local administration of muscimol in the DRN. Therefore, NMDAR in the DRN regulates predominantly GABAergic neurons rather than serotonergic neuronal activity in the DRN.In the DRN, serotonin release is regulated by inhibitory GABA the DRN . Previou the DRN . In cont the DRN . Taken wThe present study demonstrated that inhibition of NMDAR directly inhibited GABAergic transmission, but enhanced indirectly glutamatergic and monoaminergic transmissions induced by GABAergic disinhibition. In other words, during resting stage, MK801 selectively inhibits NMDAR on GABAergic neurons, but cannot affect NMDAR on the glutamatergic or monoaminergic neurons. In generally, GABAergic interneuron is more sensitive to NMDAR antagonist compared with other types of neurons, since the resting membrane potential of GABAergic interneurons are more positive (\u221250 ~\u221260 mV) rather than those of monoaminergic and glutamatergic neurons ,44. NMDAIn this study, the threshold concentrations of local administrations of MK801 into the mPFC, RTN and DRN on regional GABAergic transmission were almost equal to be 1 \u03bcM. Contrary to GABA, the threshold concentrations of local administration of MK801 into the mPFC and RTN on catecholamine release in the mPFC were 5 \u03bcM, whereas those into mPFC and DRN on serotonin release in the mPFC were 5 \u03bcM and 1 \u03bcM, respectively. Therefore, serotonergic transmission is more sensitive to NMDAR antagonist rather than other catecholaminergic transmissions in the mPFC.Subanesthetic doses of ketamine produces transient dissociative and psychotomimetic effects that resemble the positive and negative symptoms of schizophrenia ; howeverR)-ketamine into the infralimbic mPFC and hippocampus also produced antidepressant-like action [R)-ketamine into the prelimbic mPFC nucleus accumbens could not produce [R)-ketamine. Moreover, the binding affinity  of (S)-ketamine and (R)-ketamine to NMDAR are 0.7 and 2.6 \u03bcM, respectively [S)-ketamine is potent than that of (R)-ketamine [Neither clinical mechanisms between psychotomimetic and antidepressant effects of ketamine have remained to be clarified. Local administration of ketamine into the infralimbic mPFC reproduced the antidepressant-like actions of systemic ketamine administration . Local ae action , whereas produce . These fectively . Indeed,ketamine . The MK8ketamine . Clinicaketamine ,57. Thesketamine .l-glutamate and secondary catecholamine release in the mPFC, suggests that lower concentration of MK801 affects emotional rather than cognitive disturbances. In the present study, inhibition of AMPAR in the MDTN and mPFC attenuated the MK801-induced glutamatergic and monoaminergic transmissions without affecting GABAergic disinhibition. Several preclinical studies reported the possibility that NMDAR inhibition conversely activates glutamatergic transmission associated with AMPAR through GABAergic disinhibition [The thalamocortical glutamatergic transmission plays important roles in the function of neuro-cognition, including learning, memory, and perceptual integration ,59,60. Thibition ,65. In shibition , whereashibition . The detl-glutamate release in the mPFC activates AMPAR on the regional co-releasing terminals resulting in an increase in releases of norepinephrine and dopamine, but does not affect serotonin release. Inhibition of NMDAR in the DRN enhances serotonin release in the DRN, MDTN and mPFC via GABAergic disinhibition in the DRN. The sensitivities of serotonin release in the mPFC to local administration of MK801 into the DRN is more predominant rather than that of catecholamine release in the mPFC induced by local MK801 administration into the RTN.The present study determined the effects of MK801 on the thalamocortical (RTN-MDTN-mPFC) glutamatergic, mesothalamic (DRN-MDTN), and mesocortical (DRN-mPFC) serotonergic transmissions using multiprobe microdialysis, to clarify the NMDAR associated regulation systems in these three pathways. Inhibition of NMDAR in the RTN activates thalamocortical glutamatergic transmission induced by GABAergic inhibition and secondarily activated AMPAR in the MDTN. The enhanced"}
+{"text": "Of note, abdominal fat tissue of obese pre-DM patients treated with metformin therapy presented higher SIRT6 expression and lower NF-\u03baB, PPAR-\u03b3, and SREBP-1 expression levels compared to pre-DM control group. Collectively, results show that SIRT6 is involved in the inflammatory pathway of subcutaneous abdominal fat of obese pre-DM patients and its expression responds to metformin therapy.The role of sirtuin 6 (SIRT6) in adipose abdominal tissue of pre-diabetic (pre-DM) patients is poorly known. Here, we evaluated SIRT6 expression in visceral abdominal fat of obese pre-diabetic patients and the potential effects of metformin therapy. Results indicated that obese pre-DM subjects showed low SIRT6 protein expression and high expression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-\u03baB), peroxisome proliferator-activated receptor gamma (PPAR-\u03b3), and sterol regulatory element-binding transcription factor 1 (SREBP-1). Obese pre-DM patients showed high values of glucose, insulin resistance (HOMA-IR), C reactive protein (CRP), nitrotyrosine, tumor necrosis factor-\u03b1 (TNF-\u03b1) and interleukin 6 (IL-6), and low values of insulin ( Visceral fat and superficial adipose tissue of obese patients express cytokines cross-talking with the cardiovascular system . It has p < 0.05) (p < 0.05), lower values of insulin (p < 0.05), and higher values of HOMA-IR (p < 0.05) than NG obese patients (group 1) (p < 0.05), inflammatory markers, such as CRP (p < 0.05), IL-6 (p < 0.05), TNF-\u03b1 (p < 0.05), and nitrotyrosine (p < 0.05), than NG obese patients (group 1) (p < 0.05)  . Obese pgroup 1) . Moreovegroup 1) . The sam < 0.05) .p < 0.01). Obese pre-DM not-metformin users presented lower values of SIRT6 (p < 0.01 vs. NG) and higher values of NF-\u03baB, PPAR-\u03b3 and SREBP-1 (p < 0.01 vs. NG) (p < 0.05) and lower values of NF-\u03baB, PPAR-\u03b3, and SREBP-1 protein levels (p < 0.05) compared to obese pre-DM not-metformin users  . Moreovein users . p < 0.05). However, a regulatory effect on SIRT6 expression might be the main determinant of oxidative stress and nitrotyrosine reduction in pre-DM receiving an anti-oxidative drug therapy with metformin  buffer, 30 mM Tris-HCl, pH 8.8. Tissues were homogenized with a Precellys-24 system  and centrifuged at 800\u00d7 p < 0.05 was considered significant. Statistical analysis was performed using the SPSS software package for Windows 17.0 .Data were presented as group mean \u00b1 SD. One-way analysis of variance (ANOVA) was used to compare baseline data, followed by Scheffe\u2019s test for pairwise comparisons. Simple and partial correlation were used to evaluate relationships between variables."}
+{"text": "Eggerthellaceae within the class Coriobacteriia (phylum Actinobacteria), Adlercreutzia muris WCA-131-CoC-2 (= DSM 29508 = KCTC 15543) and Ellagibacter urolithinifaciens CEBAS 4A (= CCUG 70284 = DSM 104140).Here, we report the annotated draft genome sequences of two type strains belonging to the family  Eggerthellaceae within the class Coriobacteriia (phylum Actinobacteria), Adlercreutzia muris WCA-131-CoC-2 (= DSM 29508 = KCTC 15543) and Ellagibacter urolithinifaciens CEBAS 4A (= CCUG 70284 = DSM 104140).Here, we report the annotated draft genome sequences of two type strains belonging to the family  Eggerthellaceae are typical members of the mammalian gut and have been isolated from, e.g., humans  under anaerobic conditions of N2-CO2 (80:20) in flushed brain heart infusion medium (Merck) supplemented with 0.5% yeast extract, 0.05% l-cysteine monohydrochloride (Roth), 1\u2009mg ml\u22121 resazurin sodium salt, 2.5\u2009mg liter\u22121 heme solution, and 2\u2009\u03bcg ml\u22121 vitamin K1 solution (Sigma-Aldrich). Ellagibacter urolithinifaciens DSM 104140T was cultured (37\u00b0C) under anaerobic conditions of N2-CO2 (80:20) in flushed anaerobe basal broth (Oxoid). DNA extraction was done using a Qiagen blood and tissue kit. DNA was quantified with a double-stranded DNA (dsDNA) high-sensitivity (HS) assay on a Qubit version 2.0 fluorometer (Thermo Fisher Scientific), according to the manufacturer\u2019s instructions, and adjusted to a concentration of 0.2\u2009ng \u03bcl\u22121.The following type strains were obtained from the German Collection of Microorganisms and Cell Cultures (DSMZ): escribed , 13. AdlA. muris DSM 29508T, while 1,067,866 reads were generated for E. urolithinifaciens DSM 104140T. Data processing was done as previously described  using a BLASTn search . Whole-genome shotgun sequencing was performed on an Illumina MiSeq benchtop sequencer using a 500-cycle v2 kit . In total, 1,469,853 reads were generated for escribed , 13. Seqescribed , 16. The 2.3.3.1 . To elimn search . All conn search . To calcPipeline . The assPipeline  was usedA. muris DSM 29508T and E. urolithinifaciens DSM 104140T, has been deposited at DDBJ/ENA/GenBank under BioProject number PRJNA574580. The versions described in this publication are the first versions and are listed in A. muris DSM 29508T have been deposited under SRA accession number SRX6974936, while the raw reads of E. urolithinifaciens DSM 104140T have been deposited under accession number SRX6974937.This whole-genome shotgun project, including raw reads of"}
+{"text": "This study explores associations between frailty and oral health in cross-sectional data of 1,202 community-dwelling older people. Two dichotomous outcomes were used: 1. Potential frailty, using routine primary care data; 2. Self-reported frailty, using a questionnaire. Oral health concerned dental record data and self-reported oral problems. Following exploration of univariate associations, age and sex adjusted multivariate logistic regressions were performed. For potential frailty and self-reported frailty associations were found with dental emergency visit (odds ratio (OR)= 2.0, 95% confidence Interval (CI)=1.33;3.02 respectively OR=1.58, 95% CI=1.00;2.49), experiencing oral problems , making dietary adjustments . Additional associations were found for self-reported frailty with wearing dental prosthesis  and missing periodontal information . The cross-sectional data of this study show that in community dwelling older people oral health is associated with frailty."}
+{"text": "Scientific Reports 10.1038/s41598-019-38633-4, published online 14 February 2019Correction to: This Article contains an error in the Figure legends of Figures 2 and 3 which have been inadvertently reversed.The correct legend for Figure 2 is:A) Map of specimens CUE JJ_M01-3 showing four-track Minisauripus trackway, and additional isolated fifth track on small unconnected slab. Map based on counterpart cast of track-bearing surface. Red outline shows part of surface preserved as natural impressions. Pterosaur manus tracks, desiccation cracks (stippled areas), raindrop impressions and invertebrate traces also shown. Compare with Fig. 2 and text for details. (B) Shows microstratigraphy of part and counterpart of track-bearing slab. (C) Shows four track-trackway with dashed line to highlight steps and pace angulations. See Table 1 for measurements. Map made by M. G. Lockley and K-S Kim with layout created in Canvas X .\u201d\u201c Counterpart slab CUE JJ_M01 showing trackway with four consecutive Minisauripus track casts TL1-TR2. (B) Natural impression slab (CUE JJ_M02) showing tracks TL2 and TR2. (C) Isolated track specimen (CUE JJ_M03). Compare with Fig. 3. Photographs by K-S Kim and layout created in Canvas X .\u201d\u201c("}
+{"text": "Rhodomyrtus tomentosa, which belongs to the Myrtaceae family. In the current study, we investigated the properties of rhodomyrtone as a potential drug candidate for the treatment of stress-caused depression.Rhodomyrtone is one of the main active compounds derived from We assessed the function of rhodomyrtone in chronic unpredictable mild stress, a well-validated depression model in mice. Depression-like behavior tests, including a sucrose performance test, social interaction test, and forced swimming test, were used to validate the antidepressant effects of rhodomyrtone. The Morris water maze was used to evaluate the mice\u2019s learning and memory ability. Spine density, glycogen synthase kinase-3\u03b2, brain-derived neurotrophic factor, postsynaptic density protein 95, and apoptosis-associated protein were detected to reveal the underlying mechanism.Rhodomyrtone was found to prevent source consumption decrease, decreased social behaviors, and increase immobility in the forced swimming test, suggesting a protective effect of rhodomyrtone against depression-like behaviors. Additionally, rhodomyrtone prevented the impairment of spatial memory in mice exposed to chronic unpredictable mild stress. Rhodomyrtone administration also reversed dendritic spine density defects in chronic unpredictable mild stress. Furthermore, rhodomyrtone inhibited the increase of glycogen synthase kinase-3\u03b2 activity and reversed the decrease of brain-derived neurotrophic factor and postsynaptic density protein 95 in chronic unpredictable mild stress mice. Elevated expression of apoptosis-associated protein Bax and cleaved-caspase 3 was also reversed by rhodomyrtone treatment.These results suggested that the antidepressant effect of rhodomyrtone involves the regulation of neurogenesis, neuronal survival, and synaptic plasticity in the hippocampus. We identified a protection effect of rhodomyrtone against depression-like behaviors in a well-validated depression model in mice. Rhodomyrtone administration can prevent increase of depression-like behaviors, impairment of cognitive abilities, and morphological changes in dendritic spines in CUMS model mice. Rhodomyrtone also prevents CUMS-induced changes in depression-related proteins, including GSK3 activity, BDNF expression, Bax activity, and caspase-3 cleavage. These observations indicate that rhodomyrtone exhibits antidepressant activity involving the promotion of neurogenesis and neuronal survival in the hippocampus.As one of the common mood disorders, depression is correlated with stressful events in life . The cliRhodomyrtus tomentosa, belongs to the acylphloroglucinol derivative family. R. tomentosa has been used in traditional medicine for the treatment of many diseases  and used in the current study. Standard chow and water were supplied ad libitum. Mice were housed in a temperature-controlled (20\u00b0C\u00b11\u00b0C) room with a normal 12-h-light/-dark cycle, with the lights on at 7:00 Mice in CUMS+vehicle and CUMS+rhodomyrtone groups were exposed to CUMS as described previously . InitialThe SPT wAn adult The mouseSpatial learning and memory were assessed by the Morris water maze (MWM) . The MWMGolgi staining was performed using a GolgiStain kit according to the protocol provided by the manufacturer   on ice. Each sample of lysates (20 \u03bcL) was added into SDS-PAGE and electroblotted onto PVDF membranes, blocked by 5% bovine serum albumin, and incubated with primary antibodies overnight at 4\u00b0C, including postsynaptic density protein 95 (PSD-95) , cleaved caspase-3 , caspase3 , brain-derived neurotrophic factor (BDNF) , glycogen synthase kinase-3\u03b2 (GSK3\u03b2) , p-GSK3\u03b2 , Bax (BD), and \u03b2-actin (Sigma). Secondary antibodies were purchased from Sigma.Rhodomyrtone  was i.p. injected at 15 mg/kg. The dosages were chosen based on the behavioral results from the 5-mg/kg (low dosage), 15-mg/kg (high dosage 1), and 45-mg/kg (high dosage 2) groups . RhodomyTwo-way ANOVA was used to analyze the effects of exposure to the CUMS protocol and rhodomyrtone treatment, and a posthoc Tukey\u2019s test was conducted. All values were expressed as mean\u00b1SEM.F=10.38, CUMS: F=38, 72, Rho: F=17.21, all P<.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; F=19.72, CUMS: F=14.23, Rho: F=26.02, all P<.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; F=12.65, P<.01; F=8.41, P<.01; Rho: F=2.66, P=.11; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; To investigate the protective effects of rhodomyrtone on CUMS-induced depression, mice were exposed to CUMS protocol for 35 days, and rhodomyrtone or vehicle saline was i.p. injected daily during the last 3 weeks. Meanwhile, mice from vehicle control and rhodomyrtone control group were group-housed with no stress and were i.p. injected with vehicle or rhodomyrtone . DepressF=10.27, P<.01; day 3 interaction: F=15.47, P<.01; day 4 interaction: F=47.60, P<.001; F=0.49, P=.48; F=2.20, P=.15; CUMS: F=13.36, P<.01; rhodomyrtone: F=7.38, P<.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.05; 2-way ANOVA; F=8.45, P<.01; CUMS: F=11.25, P<.01; rhodomyrtone: F=7.2, P<.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; To investigate whether rhodomyrtone could protect against impaired spatial cognition performance in the CUMS group, the MWM task was performed. The escape latency improved in all 4 groups with trial training. However, mice in the CUMS group spent more time to locate the hidden platform during the training. Interestingly, rhodomyrtone treatment significantly improved the latency time of CUMS mice =4.39, P<.05; CUMS: F=12.93, P<.01; rhodomyrtone: F=4.39, P<.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.05; Alterations in synaptic and dendritic structure and function are associated with impaired learning and memory in depressive disorders . We deteF=8.01, P<.01; CUMS: F=25.83, P<.01; rhodomyrtone: F=10.59, P<.01; posthoc Tukey\u2019s tests found CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; F=2.12, P=.155; CUMS: F=58.38, P<.01; rhodomyrtone: F=7.08, P<.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.05; F=0.48, P=.49; CUMS: F=24.26, P<.01; rhodomyrtone: F=9.02, P<.01; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; Numerous studies have shown that abnormal activity of GSK3\u03b2 has multiple effects and is correlated with the severity of depressive symptoms, including neurogenesis defects . Thus, wF=6.21, P<.05; CUMS: F=10.05, P<.01; rhodomyrtone: F=2.23, P>.05; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.05; F=10.75, P<.01; CUMS: F =13.90, P<.01; rhodomyrtone: F=7.59, P<.01; posthoc Tukey\u2019s test CUMS+Veh. vs Control+Veh., P<.01; CUMS+Rho. vs CUMS+Control, P<.01; Volume decrease of the hippocampus in depression is associated with a reduction of neuronal cell body size and increased neural apoptosis in depression patients . AntidepIn the current study, we found that rhodomyrtone exhibited antidepressant activity in a mouse CUMS model. We observed that rhodomyrtone administration protected against CUMS-caused, depression-like behaviors and cognitive defects. Further, CUMS-decreased spine density was reversed by rhodomyrtone treatment. Impairment of neurogenesis and increase of apoptosis are 2 potential mechanisms of depression. We found that rhodomyrtone reversed the CUMS-induced changes in GSK3\u03b2 activity and BDNF expression level, 2 essential regulators of hippocampal neurogenesis. The activation of apoptosis-related protein Bax and caspase-3 was also reversed by rhodomyrtone treatment. Our findings demonstrated the antidepressant-like effects of rhodomyrtone through regulation of neurogenesis and apoptosis in a classic depressive mouse model induced by CUMS. Studies also showed that rhodomyrtone exhibits antibacterial, anticancer, antiinflammatory, and antioxidant activities . Here, tIt has been reported that defects in emotional memory and spatial learning are key characteristics of depression besides depression-like behaviors . The hipHippocampus shrinkage has been observed in both depressive patients and animal models . InteresAnother important explanation for hippocampus shrinkage is increased apoptosis . ChronicIn summary, for the first time to our knowledge, we showed that rhodomyrtone exhibits antidepressant-like activity in a mouse depression model. Rhodomyrtone can prevent CUMS-induced increase of depression-like behaviors, impairment of cognitive abilities, and morphological changes in dendritic spines. Rhodomyrtone also prevents CUMS-induced changes in GSK3 activation, BDNF expression, Bax activation, and caspase-3 cleavage. These observations indicate that rhodomyrtone has antidepressant effects involving the promotion of neurogenesis and neuronal survival in the hippocampus.None.Supplementary Figure 1Click here for additional data file.Supplementary MaterialClick here for additional data file."}
+{"text": "Arabidopsis NBS genes revealed a clade-specific nesting pattern in CNLs, with RNLs nested in the CNL-A clade, and species-specific nesting pattern for TNLs. Surprisingly, we found a moderate bootstrap support (BS = 50%) for CNL-A clade being nested within TNL clade making both the CNL and TNL clades paraphyletic. Arabidopsis and sunflower showed 87 syntenic blocks with 1049 high synteny hits between chromosome 5 of Arabidopsis and chromosome 6 of sunflower. Expression data revealed functional divergence of the NBS genes with basal level tissue-specific expression. This study represents the first genome-wide identification of NBS genes in sunflower paving avenues for functional characterization and potential crop improvement.Nucleotide Binding Site\u2014Leucine-Rich Repeat (NBS-LRR) genes encode disease resistance proteins involved in plants\u2019 defense against their pathogens. Although sunflower is affected by many diseases, only a few molecular details have been uncovered regarding pathogenesis and resistance mechanisms. Recent availability of sunflower whole genome sequences in publicly accessible databases allowed us to accomplish a genome-wide identification of Toll-interleukin-1 receptor-like Nucleotide-binding site Leucine-rich repeat (TNL), Coiled Coil (CC)-NBS-LRR (CNL), Resistance to powdery mildew8 (RPW8)-NBS-LRR (RNL) and NBS-LRR (NL) protein encoding genes. Hidden Markov Model (HMM) profiling of 52,243 putative protein sequences from sunflower resulted in 352 NBS-encoding genes, among which 100 genes belong to CNL group including 64 genes with RX_CC like domain, 77 to TNL, 13 to RNL, and 162 belong to NL group. We also identified signal peptides and nuclear localization signals present in the identified genes and their homologs. We found that NBS genes were located on all chromosomes and formed 75 gene clusters, one-third of which were located on chromosome 13. Phylogenetic analyses between sunflower and  CNL-C (I) clade, sister clade to CNL-C (II) and CNL-D constituted 79 genes. CNL-B clade constituted three genes . The remaining 12 genes did not belong to any clade of Arabidopsis CNL genes. The TNL group formed a species-specific clade, except ten genes that formed a small clade with AT5G36930, named TNL-D clade with strong bootstrap support of 100%. We found a moderate bootstrap support (BS = 50%) for CNL-A clade being nested within TNL clade making both the CNL and TNL clades paraphyletic. Another tree constructed using RNL genes of A. thaliana and H. annuus showed two distinct clades for two lineages: activated disease resistance gene 1 (ADR1) and N-required gene 1 (NRG1)  was used in phylogenetic analyses. Phylogenetic relationships among the sunflower NBS sequences are shown in NL group . The nes1 (NRG1) . The Newhomologs . The synbidopsis .H. annuus clone Ha-NTIR11g CC-NBS-LRR gene (Pl8). HanXRQChr13g0425411, HanXRQChr13g0425361, and HanXRQChr13g0425431 showed more than an 80% identity to the Pl8 gene suggesting the probable homologs to that gene. HanXRQChr04g0123041, belonging to the NL group has shown homology to Lycopersicon esculentum EIX receptor 1 (LeEIX1), a gene that encodes receptor-like proteins (RLPs). Similarly, HanXRQChr17g0552491 showed homology to MLA10, HanXRQChr13g0420141 to N, HanXRQChr17g0552491 to both MLA12 and MLA13 and HanXRQChr17g0552491 to Sr33 protein with greater than 60% identity. Sunflower Genome Database with H. annuus r1 annotations was employed to obtain expression data for predicted NBS genes. We compared accessions of H. annuus r1.2 annotations to H. annuus r1 to obtain the expression data for NBS proteins. Since there were many duplicates for H. annuus r1.2 annotations, we used only the sequences with the unique names. The raw Read Per Kilobase Million) (RPKM) values of gene expression were downloaded separately. The expression values were from bract, corolla, leaves, ligule, ovary, pollen, seed, stamen and stem. Only expression data for 9 CNL type, 33 TNL type, 23 NL type and 6 RNL type genes were retrieved from the database and employed to generate heatmap after deseq normalization of the data using MeV package  [Arabidopsis (~0.43%) [C. sativus (~0.21%) [Carica papaya (~0.21%) [P. vulgaris (~1.19%) [Manihot esculenta (~0.9%) [V. vinifera (~1.3%) [G. max (~0.73%) [Our findings on the NBS-encoding genes in this study is based on recently sequenced sunflower genome . Previoual. 2004  identifial. 2008  identifi(~0.66%) . This nu(~0.43%) , C. sati(~0.21%) , Carica (~0.21%)  and lowe(~1.19%) , Manihot (~0.9%) , V. vini (~1.3%) , and G. (~0.73%) ,24. We p(~0.73%) , Plocik (~0.73%) , and Rad(~0.73%)  showed 7A. thaliana (1:2), A. lyrata (1:2), B. rapa (1:2), Eucalyptus grandis (1:1.25), and Thellungiella salsuginea (1:1.5) as numbers of TNLs were higher than CNLs in these species [M. truncatula, A. thaliana, and B. rapa [A. thaliana and B. rapa [Following the classification of NBS genes by Shao et al. 2016  and Yu e species ,65,66,67 species ,57. The  B. rapa ,27. HanX B. rapa .S. tuberosum [A. thaliana, P. vulgaris, G. max, and P. trichocarpa [P. trichocarpa [B. rapa [A. thaliana, P. trichocarpa, and O. sativa [NBS-encoding genes also called NBS-LRR genes encode proteins having TIR/CC at the N-terminal, NBS domain in the center and LRR at the C-terminal . Among tuberosum . All TIRchocarpa ,24,63,69chocarpa . All of [B. rapa . Kelch m. sativa .Arabidopsis CLAVATA2  [Cladosporium fulvum in tomato [We further compared our pipeline with another pipeline, RGAugury , for theAtRLP10)  is involn tomato .M. esculenta, 143 NBS genes positioned in 39 clusters [C. sativus, 33 NBS genes were located in nine clusters [B. oleracea (3.04), B. rapa (2.7), A. thaliana (2.8) [G. max (4), V. vinifera (6), M. truncatula (5) [Gossypium species such as G. arboretum (3.4), G. raimondii (5.5), G. hirsutum (5.3), and G. barbadense (3.5) [Arabidopsis [aK/sK values of less than one, indicating that these genes are under the influence of purifying selection.A variety of clustering patterns of NBS-encoding genes, frequently observed in almost all plant species, is one of the major reasons for rapid evolution of the NBS genes ,79. The clusters . In C. sclusters , Brassicna (2.8) , Fabaceatula (5) , Gossypise (3.5) . Both sese (3.5) . HoweverC. sativus CNL genes while compared to their respective TNL genes [Arabidopsis, as TNL clades constitute larger numbers of genes than CNL clade [HanXRQChr02g0057361, HanXRQChr02g0057351, and HanXRQChr13g0425771 in the subclade CC (VI) possessed in the range of five to seven. Similarly, subclade TIR (II) possessed gene members with introns in the lowest range (three to five). TIR (I), TIR (III), TIR (IV), TIR (V) and TIR (VI) gene members possessed introns in range of 3 to 17, 2 to 7, 1 to six, 1 to 15, and 1 to 13, respectively. Similar patterns were also observed in the phylogenetic tree of CNL and TNL in C. sativus [Sunflower CNL genes were similar to NL genes . HoweverNL clade . Subclad sativus . The dif sativus . In addi sativus .H. annuus in the analysis as the genome was not available by then. Our results indicate a surprising position of RNLs within TNLs in sunflower making the clades of TNL and CNL potentially paraphyletic. Upon reconstruction of the phylogenetic tree with Arabidopsis NBS genes, RNL genes of sunflower were observed in a CNL-A clade  [ADR1 and NRG1, and two ancient lineages separated before the Angiosperms diversified. The RNL genes, ADR1 and NRG1, have been characterized in Arabidopsis and Nicotiana, respectively. A separate tree, constructed to observe the relationships among sunflower RNLs and Arabidopsis RNLs, formed two clades. The sunflower RNL genes HanXRQChr02g0046611 and HanXRQChr05g0129181 were nested with AT4G3330 (ADR1-L1), AT1G33560 (ADR1) and AT5G04720 (ADR1-L2 or PHX21), with bootstrap support of 90%. On the other hand, HanXRQChr02g0048181, HanXRQChr11g0331571, HanXRQChr03g0067681, HanXRQChr0073241, and HanXRQChr04g0095241 were nested with AT5G66630 (RNL) and AT5G66910 (homologous to NRG1), with bootstrap support of 63%. This suggests that the sunflower RNLs mentioned above are orthologous to the ADR1 and NRG1 homologs of Arabidopsis. ADR1 proteins play a role as helper genes for receiving signals from the R genes in downstream signaling of effector-triggered immunity [A. lyrata (2.5%), A. thaliana (4.2%), B. rapa (4.4%), Capsella rubella (4.7%) and T. salsuginea (5.7%) [a/KsK ratios values for the clade. This supports the hypothesis of high conservation and slow evolutionary rates among the RNL genes [We found that RNLs were nested within the clade of TNLs in sunflower (a member of the Asterids lineage) although RNLs in the families Brassicaceae and Fabaceae (Rosids lineage) were found to be related to CNLs ,30. The s study) . The CNLs study)  suggesteimmunity . Similarimmunity . Since timmunity . Only 5.NL genes .S. tuberosum nematode resistance protein (Gro1.4) [S. tuberosum subsp. andigena RY-1 (conferring resistance to potato virus Y) [N. glutinosa Tobacco Mosaic Virus resistance (N) gene [A. thaliana RPS2 (Resistant to P. syringae 2) [Cucumis melo VAT (resistance to Aphis gossypii) [H. annuus Pl8 [O. sativa Pi36 (conferring resistance to Blast fungus) [H. vulgare subsp. vulgare RPG1 (conferring resistance to stem rust fungus) [http://www.prgdb.org has shown some of them to be the possible homologs of the reference proteins . The Pl8 gene is involved in conferring resistance to P. halstedii, a causative agent to downy mildew [L. esculentum EIX receptor 2 (Eix2), a gene that encodes receptor-like proteins (RLPs) involved in detecting ethylene-inducing xylanase, a fungus elicitor [MLA locus is highly polymorphic and encode allelic CNL type resistance proteins such as MLA1, MLA2, and MLA3 that confer resistance to barley powdery mildew fungus  [Puccinia graminis f. sp. tritici [H. annuus r1.2 annotations compared to H. annuus r1 annotations. From the available expression data, it can be deduced that NBS genes can be expressed at a basal level with tissue specificity in unchallenged conditions [Sunflower NBS proteins identified in this study formed clades with reference proteins such as Pi36, Pl8, Rps2, VAT, RPG1, Gro1.4, RY-1, and N proteins, suggesting their homologous relationships . The sun(Gro1.4) , S. tubevirus Y) , and N. (N) gene . Similaringae 2) , Cucumisossypii) , H. annunuus Pl8 , O. sati fungus) , and H.  fungus) . The BLAproteins . Sunflowy mildew . HanXRQCelicitor . Other iei, Bgh) . Another tritici . We werenditions . In the nditions . Thus, dPl8, LeEIX1, MLA10-13, Sr33 resistance genes. Further characterization of the NBS genes will help us to understand resistance pathways and to develop durable resistance necessary for crop improvement in sunflower, one of the major oilseed crops in the world.We identified 352 NBS genes in sunflower and studied their clustering, phylogenetic relationships, gene homology and functional divergence. These genes formed clusters and showed structural conservation in signature domains and exon/intron architecture in CNL, TNL and RNL types of NBS genes. The RNLs belonged to the CNL-A clade, which in turn was found nested within the TNL clade, making both CNL and TNL clades paraphyletic. This warrants further rigorous analysis. All of the NBS-encoding genes have undergone purifying selection and available expression data have revealed their functional divergence. We confirmed homology of sunflower NBS genes to multiple previously characterized"}
+{"text": "Histopathological and functional outcomes were evaluated on P10, P18, and P90. Baseline outcome parameters were established in sham-treated and healthy control animals. Gross brain injury did not significantly differ between treatment groups at any time point. On P10, caspase-3 activation and caspase-independent apoptosis were similar between treatment groups; cell proliferation and the number of BrdU-positive vessels did not differ on P18 or P90. Neurobehavioral assessment did not reveal significant differences between treatment groups in accelerod performance, open field behavior, or novel object recognition capacity on P90. Turning behavior was more frequently observed in G-CSF/SCF- and FL-treated animals. No sex-specific differences were detected in any outcome parameter evaluated. In hypoxic-hyperoxic ischemic neonatal brain injury, G-CSF/SCF and FL treatment does not convey neuroprotection. Prior to potential clinical use, meticulous assessment of these hematopoietic growth factors is mandated.Hematopoietic growth factors are considered to bear neuroprotective potential. We have previously shown that delayed treatment with granulocyte colony-stimulating factor (G-CSF)/stem cell factor (SCF) and Fms-related tyrosine kinase 3 ligand (FL) ameliorates excitotoxic neonatal brain injury. The effect of these substances in combined-stressor neonatal brain injury models more closely mimicking clinical conditions has not been investigated. The aim of this study was to assess the short-, mid-, and long-term neuroprotective potential of G-CSF/SCF and FL in a neonatal model of hypoxic-hyperoxic ischemic brain injury. Five-day-old (P5) CD-1 mice were subjected to unilateral common carotid artery ligation and subsequent alternating periods of hypoxia and hyperoxia for 65 minutes. Sixty hours after injury, pups were randomly assigned to intraperitoneal treatment with (i) G-CSF (200\u2009 Neonatal brain injury is a problem of great global concern , 2. AdvaOur research group has recently been able to show that systemic treatment with the hematopoietic growth factors/cytokines granulocyte colony-stimulating factor (G-CSF)/stem cell factor (SCF) and Fms-related tyrosine kinase 3 ligand (FL) starting 60 hours after insult protects against N-methyl-D-aspartate receptor-mediated developmental excitotoxic brain damage by reducing injury extent and apoptotic cell death . WhetherFurthermore, any neuromodulatory intervention taking place during potentially critical or sensitive periods of brain development possibly has not only short-, but also long-term impact . The assThus, the aim of the current study was to investigate short-, mid-, and long-term effects of delayed treatment with G-CSF/SCF and FL in a mouse model of combined hypoxic-hyperoxic ischemic neonatal brain injury. From our previous findings we hypothesised that G-CSF/SCF and FL reduce injury extent and apoptotic cell death, stimulate neuroproliferation, and improve neurobehavioral outcome following hypoxia-hyperoxia ischemia.Heparin (Heparin Immuno 1000\u2009IU/ml) was obtained from Ebewe Pharma , and phenobarbital  was obtained from Desitin Arzneimittel GmbH ; murine G-CSF, SCF, and FL were obtained from Peprotech , and 5-bromo-2\u2032-deoxyuridine (BrdU) was obtained from Sigma-Aldrich .\u03bcg/kg bw)/SCF (50\u2009\u03bcg/kg bw) in vehicle , (ii) FL (100\u2009\u03bcg/kg bw) in vehicle, or (iii) vehicle only and received an intraperitoneal (i.p.) injection every 24 hours for three or five consecutive days, depending on timing of endpoint determination. Endpoints were assessed on P10, P18, and P90. Sex was visually determined by assessment of the anogenital distance and confirmed by polymerase chain reaction  or was determined by visual assessment alone  depending on mouse age as described previously ; 1x PBS: 37.1 (11.1), G-CSF/SCF: 37.3 (5.8), FL: 34.6 (6.3); n=11-12 per group, F=0.377, p=0.689, One-Way ANOVA, two degrees of freedom). Female mice were significantly lighter in 1x PBS and G-CSF/SCF treatment groups, whereas body weight did not differ between male and female experimental animals in FL-treated mice (mean bodyweight (SD) [g]; 1x PBS: male 44.6 (9.6), female 28.0 (2.7), n=5-6, t=3.727, p= 0.005, Student's t-test; G-CSF/SCF: male 41.1 (3.8), female 31.9 (2.8), n=5-7, t=4.528, p=0.001, Student's t-test; FL: male 35.5 (7.2), female 33.0 (5.0), n=4-7, t=0.618, p=0.552, Student's t-test; Bonferroni-adjusted level of significance p<0.025).\u03c72=1.263, p=0.532, Pearson Chi-Square, two degrees of freedom; Turning behavior with counter-clockwise movements and hyperactivity was not observed in healthy control animals, but it occurred in six animals subjected to HHI, indicating severe brain damage (1x PBS: n=1 (9.1%) , G-SCF/SCF: n=2 (16.7%) , FL: n=3 (27.3%) ; With regard to accelerod performance, baseline time until task disruption in healthy control animals was 97.2 \u00b1 46\u2009sec (mean \u00b1 SD).In HHI, accelerod performance did not significantly differ between treatment groups (mean time until task disruption (SD) [sec]; 1x PBS: 107.7 (46.7), G-CSF/SCF: 116.3 (35.6), FL: 146.7 (74.4); n=11-12 per group, F=1.624, p=0.213, One-Way ANOVA, two degrees of freedom). No statistically significant sex-specific differences were detected .With regard to open field assessments, baseline anxiety index in sham-treated animals was 80.6%  (median (IQR)), baseline travel distance as an indicator of horizontal activity was 3000.3\u2009cm  (median (IQR)), and baseline time spent rearing as an indicator of vertical activity was 68.1\u2009sec (12.9) (mean (SD)).In HHI, anxiety (median anxiety index (IQR) [%]; 1x PBS: 85.0 , G-CSF/SCF: 78.8 , FL: 85.4 ; n=34, H=2.291, p=0.318, Kruskal-Wallis test, two degrees of freedom), horizontal activity (median travel distance (IQR) [cm]; 1x PBS: 2871.6 ; G-CSF/SCF: 3132.2 , FL: 3420.1 ; n=34, H=4.191, p=0.123, Kruskal-Wallis test, two degrees of freedom), and vertical activity (mean time spent rearing (SD) [sec]; 1x PBS: 71.6 (29.4), G-SCF/SCF: 85.0 (33.4), FL: 68.7 (49.3); n=11-12 per group, F=0.603, p=0.554, One-Way ANOVA, two degrees of freedom) did not significantly differ between treatment groups. No statistically significant sex-specific differences were detected .With regard to novel object recognition, baseline discrimination index (DI) in sham-treated animals was 0.16  (median (IQR)) after one hour, 0.29  (median (IQR)) after three hours, and 0.35  (median (IQR)) after five hours.In HHI, no overall differences in DI between treatment groups were observed after one (median DI (IQR); 1x PBS: 0.41 , G-CSF/SCF: 0.47 , FL: 0.22 ; n=34, H=2.269, p=0.322, Kruskal-Wallis test, two degrees of freedom), three (median DI (IQR); 1x PBS: 0.63 , G-CSF/SCF: 0.61 , FL: 0.46 ; n=34, H=1.942, p=0.379, Kruskal-Wallis test, two degrees of freedom), or five hours (median DI (IQR); 1x PBS: 0.40 , G-CSF/SCF: 0.26 , FL: 0.43 ; n=34, H=1.250, p=0.535, Kruskal-Wallis test, two degrees of freedom). Details can be seen in The identification of novel treatment options for brain injury is one of the biggest challenges encountered in neonatal neuroscience , 40, 41.In the study at hand, we used a combined hypoxic-hyperoxic ischemic brain injury model, thought to more adequately reflect the clinical situation in neonatal intensive care units , 34. AlsOn a histomorphological basis, we did not observe statistically significant differences between treatment groups at any time point, either in gross brain injury, or in caspase-dependent or -independent apoptosis as indicated by AIF-positivity. Another mechanism of action described for G-CSF/SCF is its trophic effect on neurons . An incrThis is in discordance with previous neonatal rodent brain injury studies, reporting reduced damage extent, apoptotic cell death, and increased cell proliferation following both single and repetitive G-CSF treatments , 24, 28.\u03bcg/g body weight was chosen, based on previous publications and our own studies, showing a dose response curve [For the present study, a higher dose of G-CSF was selected. The dose of 200\u2009se curve , 30. These curve  The diffse curve  had no ese curve  A directInterestingly, turning behavior as an indicator of severe brain damage was more frequently observed in G-CSF/SCF- and FL-treated animals than in vehicle-treated controls. Even though the finding was not statistically significant, this might still be of relevance as turning behavior has been shown to correlate with histological damage and sensory motor deficit in rodents exposed to cerebral ischemia . CautiouThe importance of evaluating sex-specific differences has been proven by several studies, especially when considering the variable responses of male and female immature rodents to brain injury , 47, 48.To conclude, in this multistressor mouse model of hypoxia-hyperoxia ischemia we cannot confirm the protective effect of G-CSF/SCF and FL on neonatal brain injury previously reported. As adverse effects cannot be ruled out, meticulous assessment of these hematopoietic growth factors in extensive preclinical studies is mandated prior to potential clinical use."}
+{"text": "This article has been corrected: Due to a labeling error, the legends for Figures 2 and 3 were accidentally switched. The proper figure legends are given below. The authors declare that these corrections do not change the results or conclusions of this paper.Figure 2: Induction of fibrosis in the mammary gland of NeuT/ATTAC mice following AP21087 treatment for four weeks. Six-week-old NeuT/ATTAC mice were injected i.p. with vehicle (NeuT/ATTAC) or 0.4 mg/kg AP21087 (NeuT/ATTAC+AP) for four weeks. FFPE sections were stained as described in Figure 2. Magnification 400\u00d7.Figure 3: Induction of fibrosis in NeuT/ATTAC mice. Mice at 6 weeks-of-age were injected i.p. with vehicle (NeuT/ATTAC) three times weekly for 5.5 months (NeuT/ATTAC) or with 0.4 mg/kg AP21087 (NeuT/ATTAC+AP) for 4 months. FFPE sections were stained with H&E, for collagen (PicroSirius Red) and with antibodies against Ki-67, CD31, \u03b1-smooth muscle actin (SMA), F4/80, Cxcl1, Foxp3, CD8 and PD-L1. Magnification 400\u00d7.8042-8053. https://doi.org/10.18632/oncotarget.24233Original article: Oncotarget. 2018; 9:"}
+{"text": "A post-conflict vaccination campaign, Central African Republic. Bull World Health Organ. 2018 Aug 1;96(8):540\u201347. on page 544,"}
+{"text": "Scientific Reports 10.1038/s41598-018-34229-6, published online 31 October 2018Correction to: In this Article the Competing Interests section was omitted. This should read:\u201cT.S. received research funding from Sony IP&S Inc.\u201d"}
+{"text": "This article has been corrected: The correct order of authors' names is given below:1,2, Milind Shivajirao Patole1, Punam Vasudeo Nagvenkar1, Sonali Shankar Kamble2 and Rajesh Nivarti Gacche2,3Bhimashankar Gurushidhappa Utage30304-30323. https://doi.org/10.18632/oncotarget.25717Original article: Oncotarget. 2018; 9:"}
+{"text": "The correct name is: Ashwani K. Singal. The correct citation is: Axley P, Mudumbi S, Sarker S, Kuo Y-F, Singal AK (2018) Patients with stage 3 compared to stage 4 liver fibrosis have lower frequency of and longer time to liver disease complications. PLoS ONE 13(5): e0197117."}
+{"text": "Correction to: J Immuno Therapy Cancer. (2018) 6 (Suppl 1)https://doi.org/10.1186/s40425-018-0422-yhttps://doi.org/10.1186/s40425-018-0423-xAfter publication of this supplement , 2, it wP128Multiplexed immunofluorescent assay development for study of the PD-1/PD-L1 checkpoint in the tumor immune microenvironment (TIME)Sneha Berry, MS, Nicolas Giraldo, MD PhD, Peter Nguyen, MS, Benjamin Green, BS, Haiying Xu, Aleksandra Ogurtsova, Abha Soni, DO, Farah Succaria, MD, Daphne Wang, MS, Charles Roberts, Julie Stein, MD, Elizabeth Engle, MSc, Drew Pardoll, MD, PhD, Robert Anders, MD, PhD, Tricia Cottrell, MD, PhD, Janis M. Taube, MDJohns Hopkins University School of Medicine, Baltimore, MD, USACorrespondence: Sneha Berry (jtaube1@jhmi.edu)Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P128Janis M. Taube, MD is a contributing author and has therefore been added to the author list in this correction article. Contributing author Sneha Berry, MS is listed three times in the original article; this is no longer the case in this correction article.BackgroundMultispectral immunofluorescent (mIF) staining of formalin-fixed paraffin-embedded (FFPE) tissue allows spatially-resolved quantitative analysis of cell position and protein expression. The design and validation of mIF panels is a challenge. Our goal was to develop a 7-plex assay for characterizing PD-1 and PD-L1 expression, with high sensitivity for multiple markers and minimal bleed-through between fluorescent channels, while avoiding steric hindrance among markers occupying the same cellular compartment.MethodsSingle IF slides were stained for PD-1, PD-L1, CD8, FoxP3, CD163, and a tumor marker (e.g. Sox10/S100 for melanoma) using primary antibodies at manufacturer\u2019s recommended concentrations and visualized with an Opal kit (PerkinElmer). Positive signal was compared to chromogenic IHC (n=3 tonsil specimens). In some instances, the kit\u2019s HRP-polymer was substituted for one that provided greater amplification. Primary antibody titrations were performed, and the concentration with comparable signal to chromogenic IHC that showed the highest IF signal to noise ratio was selected. Using the selected primary antibody concentration, TSA dilution series were performed on n=5 tumor specimens to minimize bleed-through. Finally, the optimized single IF stains were combined into multiplex format, which was again validated to ensure no positivity loss. Images were scanned with the Vectra 3.0 and processed using inForm (Ver 2.3).Resultshigh vs. PD-1low lymphocytes.The percent positive pixels for CD163, CD8, and tumor marker expression by IF were comparable to chromogenic IHC with manufacturer\u2019s recommended protocols (p>0.05). However, PD-1, PD-L1, and FoxP3 showed ~50% loss of signal (p<0.05), which was recovered by replacing the Opal kit's secondary HRP polymer with PowerVision (Leica). Unbalanced fluorescence intensities between 540 to 570 Opal dyes resulted in significant bleed-through and led to false positive pixels. This error was minimized >2 fold (2.5% to 1.1%) by concentrating the 570 dye and ensuring that this dye pair was used to study markers in different cellular compartments (nuclear FoxP3 vs. membrane CD8), so any residual bleed-through could be discounted during image analysis. Using the optimized panel, we are able to reliably identify cell types contributing PD-L1 and PD-1 to the TIME, and even resolve populations of PD-1ConclusionsWe demonstrate successful optimization of a 7-color multiplex panel characterizing the PD-1/PD-L1 axis to provide high quality data sets for whole slide or regional analysis of the TIME. With the use of multiparametric assays such as this, we hope to guide improved approaches to patient selection and potentially identify additional tumor types likely to respond to anti-PD-(L)1 immunotherapy.Ethics ApprovalThe study was approved by Johns Hopkins University Institutional Review Board.P308A Phase 1 study of MEDI5752, a bispecific antibody that preferentially targets PD-1 and CTLA-4 expressing T cells, in patients with advanced solid tumors2, Mark Voskoboynik3, James Kuo4, Yung-Lue Bang5, Hyun-Cheo Chung6, Myung-Ju Ahn7, Sang-We Kim8, Ayesh Perera1, Daniel Freeman1, Ikbel Achour1, Raffaella Faggioni1, Feng Xiao1, Charles Ferte1, Charlotte Lemech4Ben Tran1MedImmune, Gaithersburg, MD, USA; 2Peter MacCallum Cancer Center, Melbourne, Australia; 3Nucleus Network, Melbourne, Australia; 4Scientia Clinical Research, Sydney, Australia; 5Seoul National University Hospital, Seoul, Korea, Republic of; 6Yonsei Cancer Center, Yonsei University, Seoul, Korea, Republic of; 7Samsung Medical Center, Seoul, Korea, Republic of; 8Asan Medical Center, Songpa-Gu, Korea, Republic ofCorrespondence: Charles Ferte (fertec@MedImmune.com) Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P308Jeff Brubaker was not a contributing author and has therefore been removed from the author list in this correction article. The credentials as shown on the original article are no longer listed on this correction article.BackgroundBased on demonstrated clinical activity and manageable safety profiles, checkpoint inhibiting antibodies blocking PD 1, PD-L1, or CTLA-4 have received regulatory approvals for the treatment of various malignancies [1-5]. The combination therapy with anti-PD-1 and anti-CTLA-4 agents is approved by FDA for metastatic melanoma, renal cell carcinoma and microsatellite instability-high (MSI-H) or mismatch repair deficient (dMMR) metastatic colorectal cancer, based on improved overall survival versus either agent alone [6-10]. Numerous clinical studies of combination immunotherapy are currently investigating the same combination across a range of solid tumors [11- 15]. Although the efficacy of these drug combinations is dose dependent, the toxicity associated with anti-CTLA-4 agents, in particular, is dose limiting, thereby potentially affecting treatment outcomes with combination therapy.- MEDI5752 is a bispecific humanized IgG1 monoclonal antibody that binds PD-1 and CTLA-4. In contrast to the combination therapy, MEDI5752 exhibits a novel T cell targeting mechanism that could provide a favorable toxicity profile. In addition, we have shown that MEDI5752 can impact cell surface expression of PD-1. Based on these novel mechanisms of action, MEDI5752 may show improved efficacy and safety in comparison to co- administration of conventional anti-PD1/anti-PD-L1 and anti-CTLA-4 antibodies.MethodsThis is a Phase 1, first-time-in-human, multicenter, open-label study in patients with advanced solid tumors. The dose-escalation phase will evaluate approximately six MEDI5752 dose levels to identify a maximum tolerated dose. Dose escalation will be followed by two dose-expansion cohorts in defined setting with patients with advanced or metastatic solid tumor and tested against a control arm. Subjects will remain on treatment until confirmed progressive disease, initiation of alternative cancer therapy, unacceptable toxicity, or other reason for discontinuation. The primary endpoints are safety and efficacy (objective response in the dose-expansion phase). Secondary endpoints include additional efficacy assessment across both phases, pharmacokinetics, and immunogenicity.Trial RegistrationNCT03530397References1. Opdivo Prescribing Information. Princeton, NJ: Bristol-Myers Squibb Company, 2018. [revised 2018 Apr].2. Keytruda Prescribing Information. Whitehouse Station, NJ: Merck Sharp & Dohme Corp, 2018 [revised 2018 Jun].3. Tecentriq Prescribing Information. San Francisco (CA): Genentech, Inc, 2018. [revised 2019 Jul].4. Imfinzi Prescribing Information. Wilmington (DE): AstraZeneca Pharmaceuticals LP, 2018. [revised 2018 Feb].5. Yervoy Prescribing Information. Princeton (NJ): Bristol-Myers Squibb Company, 2018. [revised 2018 Apr].6. Larkin J, Chiarion-Sileni V, Gonzalez R, Grob JJ, Cowey CL, Lao CD, et al. Combined nivolumab and ipilimumab or monotherapy in untreated melanoma. N Engl J Med. 2015;373:23-34.7. Postow MA, Chesney J, Pavlick AC, Robert C, Grossmann K, McDermott D, et al. Nivolumab and ipilimumab versus ipilimumab in untreated melanoma. N Engl J Med. 2015;372:2006-2017.8. Motzer RJ, Tannir NM, McDermott DF, Frontera OA, Melichar B, Choueiri TK, et al. Nivolumab plus ipilimumab versus sunitinib in advanced renalcell carcinoma. N Engl J Med. 2018;378:1277-1290.9. Overman MJ, Lonardi S, Wong KYM, Lenz H-J, Gelsomino F, Aglietta M, et al. Durable clinical benefit with nivolumab plus ipilimumab in DNA mismatch repair\u2013deficient/microsatellite instability\u2013high metastatic colorectal cancer. J Clin Oncol. 2018;36:773-779.https://www.pharmacytimes.com/news/nivolumab-ipilimumabcombo-approved-by-fda-for-msihdmmr-colorectal-cancer. Accessed 18 July, 2018.10. Broderick JM. Nivolumab, ipilimumab combo approved by FDA for MSI-H/dMMR colorectal cancer. Pharmacy Times, 11 July, 2018. Available at: 11. Antonia S, Goldberg SB, Balmanoukian A, Chaft JE, Sanborn RE, Gupta A, et al. Safety and antitumour activity of durvalumab plus tremelimumab in non-small cell lung cancer: a multicentre, phase 1b study. Lancet Oncol. 2016a;17(3):299-308.12. Haddad R, Gillison M, Ferris RL, Harrington K, Monga M, Baudelet C, et al. Double-blind, two-arm, phase 2 study of nivolumab (nivo) in combination with ipilimumab (ipi) versus nivo and ipi-placebo (PBO) as first-line (1L) therapy in patients (patients) with recurrent or metastatic squamous cell carcinoma of the head and neck (R/M SCCHN)\u2014CheckMate 714. Ann Oncol. 2016;27(suppl_6):1017TiP-TiP.13. Kelley RK, Abou-Alfa GK, Bendell JC, Kim T-Y, Borad MJ, Yong W-P, et al. Phase I/II study of durvalumab and tremelimumab in patients with unresectable hepatocellular carcinoma (HCC): Phase I safety and efficacy analyses. J Clin Oncol. 2017;35(15_suppl):4073.14. Janjigian YY, Ott PA, Calvo E, Kim JW, Ascierto PA, Sharma P, et al. Nivolumab \u00b1 ipilimumab in patients with advanced (adv)/metastatic chemotherapy-refractory (CTx-R) gastric (G), esophageal (E), or gastroesophageal junction (GEJ) cancer: CheckMate 032 study. J Clin Oncol. 2017;35(15_suppl):4014.15. Escudier B, Tannir NM, McDermott DF, Frontera OA, Melichar B, Plimack ER, et al. LBA5CheckMate 214: Efficacy and safety of nivolumab + ipilimumab (N+I) v sunitinib (S) for treatment-na\u00efve advanced or metastatic renal cell carcinoma (mRCC), including IMDC risk and PD-L1 expression subgroups. Ann Oncol. 2017;28(suppl_5):mdx440.029- mdx440.029.P309Phase I dose-finding study of MIW815 (ADU-S100), an intratumoral STING agonist, in patients with advanced solid tumors or lymphomas2, Theresa Werner3, Stephen Hodi4, Wells Messersmith, MD5, Nancy Lewis6, Craig Talluto7, Mirek Dostalek6, Aiyang Tao6, Sarah McWhirter8, Damian Trujillo8, Jason Luke, MD, FACP9Funda Meric-Bernstam, MD2MD Anderson Cancer Center, Houston, TX, USA; 3University of Utah, Salt Lake City, UT, USA; 4Dana-Faber Cancer Institute, Boston, MA, USA; 5University of Colorado Cancer Center, Aurora, CO, USA; 6Novartis Pharmaceuticals Corporation, East Hanover, NJ, USA; 7Novartis Institutes for BioMedical Resea, Cambridge, MA, USA; 8Aduro Biotech Inc, Berkeley, CA, USA; 9The University of Chicago Medicine, Chicago, IL, USACorrespondence: Funda Meric-Bernstam (fmeric@mdanderson.org)Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P309Janis Callister was not a contributing author and has therefore been removed from the author list in this correction article. The redundant affiliation Articulate Science as shown on the original article is no longer listed on this correction article.BackgroundMIW815 (ADU-S100) is a novel synthetic cyclic dinucleotide that can activate human STING (STimulator of INterferon Genes) in antigenpresenting cells. In preclinical models, STING pathway activation can induce tumor antigen-specific T-cell priming within the tumor microenvironment, leading to antitumor immunity and tumor destruction.MethodsEligible patients  include those with advanced/metastatic solid tumors or lymphomas with progressive disease despite standard of care or for whom there is no standard treatment.MIW815 (ADU-S100) is administered by weekly intratumoral injections (3 weeks on/1 week off) at escalating doses (starting dose: 50\u03bcg) in 28-day cycles. Primary objectives are to characterize safety and tolerability and to identify a recommended dose for future studies.Secondary objectives include preliminary efficacy, pharmacokinetics (PK), and pharmacodynamics (PD). The study is currently in dose escalation.ResultsAs of June 15, 2018, 41 heavily pretreated patients  with various solid tumors or lymphomas were enrolled. Thirty-five patients have discontinued from the study for the following reasons: disease progression (n=26), physician/patient decision (n=8), and death (n=1); 6 patients continue to receive treatment. No dose-limiting toxicities (DLTs) were reported during the first cycle at any dose level. The most common (\u226510% of patients) treatment-related AEs (TRAEs) were pyrexia , injection site pain , and headache . Grade 3/4 TRAEs included increased lipase  and elevated amylase, tumor pain, dyspnea, respiratory failure, and injection site reaction . Systemic MIW815 (ADU- S100) exposure increased with dose. On-treatment tumor biopsies showed increases in CD8 T cells infiltrating the injected tumors in a subset of patients. Preliminary antitumor activity, PK analysis, and PD data from injected lesions, noninjection lesions, and peripheral blood, will be presented.ConclusionsIntratumoral injection of MIW815 (ADU-S100) was well tolerated in doses tested thus far in patients with advanced solid tumors and lymphoma, with no DLTs reported to date. Trials evaluating combinations of MIW815 (ADU-S100) with anti-PD1 or anti-CTLA4 antibodies are ongoing.Ethics ApprovalThis study was approved by an independent ethics committee or institutional review board at each site.P383Combination therapy with M7824 (MSB0011359C) and NHSmuIL12 enhances antitumor efficacy in preclinical cancer models2, Bo Marelli2, Jin Qi2, Guozhong Qin2, Huakui Yu2, Molly Jenkins2, Kin-Ming Lo2, Joern-Peter Halle2, Yan Lan, MD2Chunxiao Xu, PhD2EMD Serono Research and Development, Belmont,MA, USACorrespondence: Yan Lan (yan.lan@emdserono.com)Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P383Colleen Stanton was not a contributing author and has therefore been removed from the author list in this correction article. The redundant affiliation Nucleus Global as shown on the original article is no longer listed on this correction article.BackgroundPD-1/PD-L1 pathway inhibition is a clinically validated approach in cancer therapy. However, most patients do not respond to the monotherapy due to multiple immunosuppressive mechanisms. Combining anti-PD-1/ PD-L1 with other immunotherapeutic agents targeting additional immunomodulatory pathways in the tumor microenvironment (TME) is one strategy to overcome resistance and improve response rates. M7824 is an innovative first-in-class bifunctional fusion protein composed of two extracellular domains of TGF-\u03b2 receptor II (a TGF-\u03b2 \u201ctrap\u201d) fused to a human anti-PD-L1 IgG1 monoclonal antibody. Through simultaneous blockade of the PD-L1 and TGF-\u03b2 pathways, M7824 demonstrated enhanced anti-tumor activity in preclinical models [1]. NHS-IL12, and the surrogate NHS- muIL12, are immunocytokines designed to target tumor necrotic regions to deliver IL-12 into the TME, where they can activate NK cells and CD8+ T cells to increase their cytotoxic functions. The surrogate NHSmuIL12 has demonstrated antitumor efficacy in preclinical models [2].This study is designed to investigate whether M7824 treatment may further benefit from combination therapy with NHS-muIL12.MethodsMice bearing MC38, EMT-6, or 4T1 tumors were treated with M7824, NHS-muIL12, or combination therapy. Tumor growth and survival were assessed in each model, and tumor recurrence following remission and rechallenge was evaluated in the EMT-6 model. Immune cell populations in the spleens and tumors were evaluated by flow cytometry and the frequency of tumor antigen-reactive IFN\u03b3-producing CD8+ T cells was evaluated by an ELISpot assay in the MC38 model.ResultsCombination of M7824 and NHS-muIL12 enhanced antitumor activity and extended the survival relative to either monotherapy in preclinical tumor models. Combination therapy also enhanced the proliferation, infiltration, and cytotoxicity of CD8+ T cells relative to monotherapies. In addition, the combination therapy increased the frequency of tumor antigenreactive T cells and induced the generation of tumor-specific immune memory, as demonstrated by protection against tumor rechallenge.ConclusionsThese data demonstrate that combination therapy with M7824 and NHS-muIL12 improved anti-tumor efficacy in multiple preclinical tumor models and suggest that combining these therapies may be a promising therapeutic strategy for patients with solid tumors.References1. Lan Y, et al. Enhanced preclinical antitumor activity of M7824, a bifunctional fusion protein simultaneously targeting PD-L1 and TGF-\u03b2. Sci Trans Med. 2018;10(424).2. Fallon J, et al. The immunocytokine NHS-IL12 as a potential cancer therapeutic. Oncotarget 2014;5(7):1869-84.P391A phase 1b/2 trial of lenvatinib in combination with pembrolizumab in patients with advanced melanoma2, Nicholas Vogelzang, MD3, Allen Cohn3, Daniel Stepan4, Robert Shumaker, PhD,4, Corina Dutcus4, Matthew Guo4, Emmett Schmidt, MD PhD5, Drew Rasco6Matthew Taylor, MD2Oregon Health and Science University, Portland, OR, USA; 3McKesson Specialty Health, Las Vegas, NV, USA; 4Eisai Inc., Woodcliff Lake, NJ, USA; 5Merck & Co., Inc., Kenilworth, NJ, USA; 6South Texas Accelerated Research Therape, San Antonio, TX, USACorrespondence: Matthew Taylor (taylmatt@ohsu.edu) Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P391The redundant affiliation 1. Oxford PharmaGenesis, Oxford, UK as shown on the original article is no longer listed on this correction article.BackgroundLenvatinib is a multikinase inhibitor of VEGFR 1\u22123, FGFR 1\u22124, PDGFR\u03b1, RET, and KIT. Pembrolizumab, an anti-PD-1 antibody, is approved for the first-line treatment of patients with advanced melanoma, with objective response rates (ORR) of 21\u201334% . Preclinical studies indicate that lenvatinib decreases the population of tumorassociated macrophages, increases CD8+ T cell infiltration, and augments the activity of PD-1 inhibitors; therefore, lenvatinib is a rational combination partner for pembrolizumab . We report interim results of an ongoing phase 1b/2 trial evaluating lenvatinib in combination with pembrolizumab in patients with solid tumors, focusing on the advanced melanoma cohort.MethodsIn this multicenter, open-label study (NCT02501096), patients with measurable, confirmed, metastatic melanoma and ECOG performance status \u22641 received lenvatinib  + pembrolizumab . Patients were not preselected based on PDL1 status. Tumor assessments were performed by study investigators using immune-related RECIST (irRECIST). The phase 2 primary end point was ORR at 24 weeks (ORRWK24). Secondary end points included ORR, progression-free survival (PFS), and duration of response (DOR).ResultsAt the data cutoff of March 1, 2018, 21 patients were enrolled: 14 (67%) patients were PD-L1(+), 4 (19%) were PD-L1(-), 3 (14%) were not tested; and 38% of patients had \u22651 prior anticancer therapy. The primary end point of ORRWK24 was 47.6% . Additional efficacy outcomes are summarized in the table (Table 1). All patients experienced \u22651 treatment-related adverse event (TRAE). Grade 3 and 4 TRAEs occurred in 13 (62%) and 1  patients respectively. There were no fatal TRAEs. The most common any-grade TRAEs were fatigue (52%), decreased appetite (48%), diarrhea (48%), hypertension (48%), dysphonia (43%), and nausea (43%). Dose reduction and interruption due to TRAEs occurred in 13 (62%) and 10 (48%) patients, respectively.ConclusionsThe lenvatinib and pembrolizumab combination regimen was welltolerated and demonstrated encouraging clinical activity. The combination may potentially improve upon the antitumor activity of antiPD-1 monotherapies, supporting further evaluation of this regimen in patients with advanced melanoma.Trial RegistrationNCT02501096References1. Ribas A, et al. Pembrolizumab versus investigator-choice chemotherapy for ipilimumab-refractory melanoma (KEYNOTE-002): a randomised, controlled, phase 2 trial. Lancet Oncology. 2015;16(8):908-18.2. Robert C, et al. Pembrolizumab versus ipilimumab in advanced melanoma. N Engl J Med. 2015;372(26):2521- 2532.3. Kato Y et al. Effects of lenvatinib on tumor-associated macrophages enhance antitumor activity of PD-1 signal inhibitors. Mol Cancer Ther. 2015;14 (12Suppl2):A92.4. Kato Y. Upregulation of memory T cell population and enhancement of Th1 response by lenvatinib potentiate antitumor activity of PD-1 signaling blockade. Cancer Res. 2017;77 (13 Suppl):4614. Ethics Approval This study was approved by all relevant institutional review boards.Table 1 (abstract P391). See text for description.P392A phase 1b/2 trial of lenvatinib in combination with pembrolizumab in patients with non-small cell lung cancer2, Nicholas Vogelzang, MD3, Christopher Di Simone3, Sharad Jain, MD3, Donald Richards3, Carlos Encarnacion3, Drew Rasco4, Robert Shumaker, PhD5, Corina Dutcus5, Daniel Stepan5, Matthew Guo5, Emmett Schmidt, MD PhD6, Matthew Taylor, MD7Marcia Brose2Abramson Cancer Center of the University, Philadelphia, PA, USA; 3McKesson Specialty Health, Las Vegas, NV,USA; 4South Texas Accelerated Research Therape, San Antonio, TX, USA; 5Eisai Inc., Woodcliff Lake, NJ, USA; 6Merck & Co., Inc., Kenilworth, NJ, USA; 7Oregon Health and Science University, Portland, OR, USACorrespondence: Marcia Brose (Brosem@mail.med.upenn.edu) Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P392The redundant affiliation 1. Oxford PharmaGenesis, Oxford, UK as shown on the original article is no longer listed on this correction article.BackgroundLenvatinib is a multikinase inhibitor of VEGFR 1\u22123, FGFR 1\u22124, PDGFR\u03b1, RET, and KIT. Pembrolizumab, an anti-PD-1 antibody, is approved as a monotherapy for previously treated patients with metastatic PD-L1\u2013positive (tumor proportion score [TPS] \u22651%) non-small cell lung cancer (NSCLC), with an objective response rate (ORR) of 18% [1]. We report interim results of an ongoing phase 1b/2 trial evaluating lenvatinib in combination with pembrolizumab in patient with solid tumors, focusing on the metastatic NSCLC cohort.MethodsIn this multicenter, open-label study (NCT02501096), patients with measurable, confirmed metastatic NSCLC and ECOG performance status \u22641 received lenvatinib  and pembrolizumab . In the phase 2 portion, patients must have had \u22642 prior lines of systemic therapy; there was no limit for phase 1b. Patients were not preselected based on PD-L1 status. Tumor assessments were performed by study investigators using immune-related RECIST (irRECIST). The phase 2 primary end point was ORR at 24 weeks (ORRWK24). Secondary end points included ORR, progressionfree survival (PFS), and duration of response (DOR).ResultsAt the data cutoff of March 1, 2018, 21 patients were enrolled. 9 (43%) Patients were PD-L1(+) (TPS \u22651%); 5 (24%) were PD-L1(-); 7 (33%) were not tested. 3 (14%) Patients were treatment-na\u00efve; 7 (33%), 10 (48%), and 1 (5%) patients had 1, 2, and \u22653 prior lines of systemic therapy, respectively. The primary end point of ORRWK24 was 33.3% . Additional efficacy outcomes are summarized in the table (Table 1). Grade 3 and 4 treatment-related adverse events (TRAEs) occurred in 10 (48%) and 1  patients, respectively. There was 1 fatal TRAE . The most common grade 3 TRAEs were hypertension (24%), fatigue (14%), diarrhea (14%), proteinuria (10%), and arthralgia (10%).ConclusionsThe combination of lenvatinib and pembrolizumab showed promising clinical activity with a manageable safety profile in previously treated patients with metastatic NSCLC who were not preselected for PD-L1 status. Further study is warranted.Trial RegistrationNCT02501096ReferencesHerbst RS et al. Pembrolizumab versus docetaxel for previously treated, PD-L1-positive, advanced non-small-cell lung cancer (KEYNOTE-010): a randomised controlled trial. Lancet. 2016;387(10027):1540-50.Ethics ApprovalThis study was approved by all relevant institutional review boards.Table 1 (abstract P392). See text for description.P393A phase 1b/2 trial of lenvatinib in combination with pembrolizumab in patients with urothelial cancer2, Carlos Encarnacion2, Allen Cohn2, Christopher Di Simone2, Drew Rasco3, Donald Richards2, Matthew Taylor, MD4, Corina Dutcus5, Daniel Stepan5, Robert Shumaker, PhD5, Matthew Guo5, Emmett Schmidt, MD PhD6, James Mier, MD7Nicholas Vogelzang, MD2McKesson Specialty Health, Las Vegas, NV, USA; 3South Texas Accelerated Research Therape, San Antonio, TX, USA; 4Oregon Health and Science University, Portland, OR, USA; 5Eisai Inc., Woodcliff Lake, NJ, USA; 6Merck & Co., Inc., Kenilworth, NJ, USA; 7Beth Israel Deaconess Medical Center, Boston, MA, USACorrespondence: Nicholas Vogelzang (nicholas.vogelzang@usoncology.com) Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P393The redundant affiliation 1. Oxford PharmaGenesis, Oxford, UK as shown on the original article is no longer listed on this correction article.BackgroundPembrolizumab, an anti-PD-1 antibody, is approved in the secondline setting for patients (objective response rate [ORR] 21%) with advanced/metastatic urothelial cancer and in the first-line setting for patients who are ineligible for cisplatin with combined positive score \u226510 or ineligible for platinum-based chemotherapy, with ORR  [1\u20133]. However, there is still an unmet need for effective therapeutic options for advanced urothelial cancer. Lenvatinib is a multikinase inhibitor of VEGFR 1-3, FGFR 1-3, PDGFR\u03b1, RET and KIT. Tyrosine kinase inhibitors, such as lenvatinib, have demonstrated activity in urothelial cancer and may reverse the immunosuppressive environment that leads to immuno-oncology (IO) therapy failure. Here we present a phase 1b/2 trial to determine the safety and efficacy of lenvatinib in combination with pembrolizumab in patients with advanced urothelial cancer.MethodsIn this multicenter, open-label study (NCT02501096), patients with confirmed metastatic urothelial cancer and an ECOG PS of 0 or 1 received lenvatinib 20 mg orally once daily and 200 mg pembrolizumab intravenously every 3 weeks. Patients were not preselected based on PD-L1 status. The phase 2 primary end point was ORR at week 24 (ORRwk24), as assessed by study investigators using immune-related RECIST (irRECIST). Secondary end points included ORR, duration of response (DOR), and progression-free survival (PFS).ResultsAt the time of data cutoff , 20 patients were enrolled. 9 (45%) Patients were PD-L1(+); 5 (25%) were PD-L1(-); 6 (30%) were not tested. 4 Patients (20%) were treatment-na\u00efve, whereas 11 (55%) and 5 (25%) patients had had 1 and 2 lines of prior anticancer therapies, respectively. No patient had received prior IO therapy. The primary end point of ORRwk24 was 25% (95% CI: 8.7\u201349.1). Additional efficacy outcomes are summarized in the table (Table 1). 18 (90%)Patients experienced treatment-related adverse events (TRAEs).Grade 3 and 4 TRAEs occurred in 5 (25%) and 5 (25%) patients, respectively. There was 1 fatal TRAE . The most common any-grade TRAEs were proteinuria (45%), diarrhea (40%), fatigue (30%), hypertension (30%), and hypothyroidism (30%).ConclusionsThe tyrosine kinase inhibitor (lenvatinib) and immunotherapy (pembrolizumab) regimen demonstrated activity in this study, which included patients receiving later-line treatment. The combination of lenvatinib and pembrolizumab deserves further investigation in patients with metastatic urothelial cancer.Trial RegistrationNCT02501096References1. Bellmunt J, et al. Pembrolizumab as second-line therapy for advanced urothelial carcinoma. N Engl J Med. 2017;376(11):1015-26.2. Vuky J et al. Updated efficacy and safety of KEYNOTE-052: A single arm phase 2 study investigating first-line pembrolizumab (pembro) in cisplatin-ineligible advanced urothelial cancer (UC). J Clin Oncol. 2018;36(15 Suppl):4524.3. Keytruda\u00ae (pembrolizumab) [package insert]. Whitehouse Station, NJ: Merck & Co., Inc.; 2017.Ethics ApprovalThis study was approved by all relevant institutional review boards.Table 1 (abstract P393). See text for description.P569Novel Pharmacobiotic approach to enhance the tamoxifen efficacy using bacterial extracellular vesicles as the immunotherapy in breast cancer1, Yeun-yeoul Yang1, Won-Hee Lee2, Jinho Yang2, Jong-kyu Kim1, HyunGoo Kim1, Se Hyun Paek1, Jun Woo Lee1, Joohyun Woo1, Jong Bin Kim1, Hyungju Kwon1, Woosung Lim1, Nam Sun Paik1, Yoon-Keun Kim2, Byung-In Moon1Jeongshin An, MD,PhD1Ewha Womans University, Seoul, Korea, Republic of; 2MD healthcare company, Seoul, Korea, Republic ofCorrespondence: Jeongshin An (rulru81@hanmail.net) Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P569Byung-In Moon is a contributing author and has therefore been added to the author list in this correction article.BackgroundThe anti-cancer effect of bacteria has a long history. According to Bierman et al., spontaneous remission of cancer has been observed in patients with severe bacteremia [1]. The reason was not revealed at that time, but we studied that in breast cancer. There are four main ways in which microbiota affects cancer: probiotics, prebiotics, drugs that target microbial enzymes and microbial products that have anticancer properties [2]. Among them, bacterial extracellular vesicles(EVs) are one of microbial products. In this study, we investigated the effects of bacterial EVs on the growth of breast cancer cells and tamoxifen efficacy.MethodsHere, we analized microbiota of urine samples by NGS to select the target EVs that were expected to affect the growth of breast cancer cells. A total of 347 female urine samples \u2013 from 127 breast cancer patients (cancer group) and 220 normal individuals (control group) \u2013 were collected and analyzed by NGS using a universal bacterial primer of 16S rDNA. Human breast cancer cells were cultured, and the cells were treated with EVs of S. aureus and K.pneumoniae for 72 h. Real-time polymerase chain reaction (PCR) and Western blotting for signalling molecule analysis were performed after treatment of EVs in each breast cancer cell.ResultsThere was a significant difference in the distribution of bacterial EVs between the urine samples from breast cancer patients and from normal controls. Especially, S.aureus EVs were predominant in the normal group, and K.pneumoniae was abundant in the breast cancer group. Therefore, we selected these two bacterial EVs that may have an effect on breast cancer cell growth. We found that S.aureus and K.pneumoniae EVs down-regulated cell growth in MDA-MB-231 cells. We also found that S.aureus or K.pneumoniae EVs had a synergic effect on growth inhibition of while co-treated with tamoxifen. S.aureus EVs down-regulated mRNA expression of cyclin E2 and up- regulated that of TNF-alpha which was related ERK pathway while co-treated with tamoxifen.ConclusionsThe anti-cancer effect of S.aureus and K.pneumoniae was initiated by its bacterial EVs and consequently inhibited the growth of breast cancer cells in triple negative breast cancer cells and improved the efficacy of tamoxifen in ER-positive cells. In the near future, we plan to conduct animal studies which are expected to further clarify the effect of bacterial EV on breast cancer. Ethics ApprovalThe study was approved by Ewha Womans University Medical Center\u2018s Ethics Board.ConsentWritten informed consent was obtained from the patient for publication of this abstract and any accompanying images. A copy of the written consent is available for review by the Editor of this journal.References1. Bierman, H. R. et al. Remissions in leukemia of childhood following acute infectious disease. Staphylococcus and streptococcus, varicella, and feline panleukopenias. Cancer 6, 591-605 (1953).2. Zitvogel, L., Daill\u00e8re, R., Roberti, M. P., Routy, B. & Kroemer, G. Anticancer effects of the microbiome and its products. Nature Reviews Microbiology 15, 465 (2017).Fig. 1 (abstract P569). See text for descriptionFig. 2 (abstract P569). See text for descriptionFig. 3 (abstract P569). See text for descriptionFig. 4 (abstract P569). See text for descriptionFig. 5 (abstract P569). See text for descriptionFig. 6 (abstract P569). See text for descriptionFig. 7 (abstract P569). See text for descriptionP651First-in-human study of FAZ053, an anti-PD-L1 mAb, alone and in combination with spartalizumab, an anti- PD-1 mAb, in patients with advanced malignancies2, David Tan3, Juan Martin-Liberal4, Shunji Takahashi5, Ravit Geva, MD6, Ayca Gucalp7, Xueying Chen8, Kulandayan Subramanian9, Jennifer Mataraza9, Jennifer Wheler9, Philippe Bedard, MD10Filip Janku, MD, PhD2MD Anderson Cancer Center, Houston, TX, USA; 3National University Cancer Institute, Singapore, Singapore 4Vall d\u2019Hebron Institute of Oncology, Barcelona, Spain; 5The Cancer Institute Hospital of JFCR, Tokyo, Japan; 6Tel Aviv Sourasky Medical Center, Tel-Aviv, Israel; 7Memorial Sloan Kettering Cancer Center, New York, NY, USA 8Novartis Pharmaceuticals Corporation, East Hanover, NJ, USA 9Novartis Institutes for BioMedical Resea, Cambridge, MA, USA; 10Princess Margaret Cancer Centre, Toronto, ON, CanadaCorrespondence: Filip Janku (fjanku@mdanderson.org)Journal for ImmunoTherapy of Cancer 2018, 6(Suppl 1):P651Janis Callister was not a contributing author and has therefore been removed from the author list in this correction article. The redundant affiliation Articulate Science as shown on the original article is no longer listed on this correction article.BackgroundFAZ053 and spartalizumab are humanized immunoglobulin G4 monoclonal antibodies (mAbs) that bind anti-programmed death ligand-1 (PD-L1) and programmed death-1 (PD-1), respectively. We report the dose-escalation results from an ongoing Phase I study of FAZ053 \u00b1 spartalizumab in patients with advanced malignancies, enriched for patients with chordoma, a rare subtype of sarcoma.MethodsPatients received escalating doses of single-agent (SA) FAZ053 intravenously once every 3 weeks (Q3W) or 6 weeks (Q6W), or FAZ053 + spartalizumab Q3W. The primary objective was to assess the safety and tolerability of FAZ053 \u00b1 spartalizumab, and determine recommended doses for expansion (RDEs). Dose escalation was guided by an adaptive Bayesian logistic regression model following the escalation with overdose control principle.ResultsAs of the data cutoff of March 30, 2018, 61 patients received SA FAZ053 at doses 80\u20131600 mg Q3W or 800\u20131600 mg Q6W. Most patients  received prior treatment; 1 (2%) received prior anti-PD-1. FAZ053 exposure was generally dose proportional, with terminal half-life of ~16\u201318 days. A dose-limiting toxicity occurred in 1 patient . RDE was determined to be 1200 mg Q3W or 1600 mg Q4W. Adverse events (AEs) of all grades assessed as possibly related to treatment were reported for 33 patients (54%); most commonly (\u226510%) fatigue  and pruritus ; 4 patients (7%) had Grade 3/4 treatment-related AEs, including elevated amylase (3%), renal failure, elevated lipase, elevated AST, and elevated blood CPK (each 2%). For these patients treated with SA FAZ053, partial responses (PRs) were demonstrated in 4 patients (7%) with chordoma, alveolar soft part sarcoma (ASPS), poorly differentiated carcinoma of scalp, and penile squamous cell carcinoma . Among 5 patients with chordoma treated with SA FAZ053, 1 patient has a PR, treatment ongoing >12 months, and 4 patients have stable disease ongoing (+4% to \u201329%). Data for 57 patients treated with combination FAZ053 (20\u20131200 mg) + spartalizumab 300 mg Q3W are preliminary. Updated results and biomarker data for patients receiving SA and combination treatment, including additional patients with chordoma, will be presented.ConclusionsSA FAZ053 was well tolerated and the RDE was determined to be 1200 mg Q3W. Clinical activity was observed in a range of indications including chordoma, a rare tumor without standard therapy options.Trial Registrationwww.clinicaltrials.gov; NCT02936102"}
+{"text": "Scientific Reports 10.1038/s41598-018-25836-4, published online 10 May 2018Correction to: In this Article, the legend of Figure 3 is incorrect:\u201cA flowchart of the rapid (RAP) system for bone staining (RAP-B). The new procedure for bone staining which is rapid, nondestructive, and allowed to obtain high-definition, fine bone-stained specimens, was based on our RAP system. Cartilage staining (RAP-C) can be applied as single staining or optionally added before the bone staining for double staining of bone and cartilage . FIXATIVE: 5% formalin, 5% Triton X-100, 1% potassium hydroxide (KOH); B-STAINING SOLUTION: 0.05% alizarin red S, 20% ethylene glycol, 1% KOH; C-STAINING SOLUTION: 50\u201370% ethanol, 20% acetate, 0.015\u20130.02% alcian blue; CLEARING SOLUTION: 20% Tween 20, 1% KOH; ENHANCEMENT SOLUTION: 20% ethylene glycol, 5% Triton X-100, 1% KOH.\u201dshould read:\u201cA flowchart of the rapid (RAP) system for bone staining (RAP-B). The new procedure for bone staining which is rapid, nondestructive, and allowed to obtain high-definition, fine bone-stained specimens, was based on our RAP system. Cartilage staining (RAP-C) can be applied as single staining or optionally added before the bone staining for double staining of bone and cartilage . FIXATIVE: 5% formalin, 5% Triton X-100, 1% potassium hydroxide (KOH); B-STAINING SOLUTION: 0.05% alizarin red S, 20% ethylene glycol, 1% KOH; C-STAINING SOLUTION: 50\u201370% ethanol, 20% acetic acid, 0.015\u20130.02% alcian blue; CLEARING SOLUTION: 20% Tween 20, 1% KOH; ENHANCEMENT SOLUTION: 20% ethylene glycol, 5% Triton X-100, 1% KOH.\u201d"}
+{"text": "Plasmodium vivax infections. Knowledge of the metabolism of PQ is critical for understanding the therapeutic efficacy and hemolytic toxicity of this drug. Recent in vitro studies with primary human hepatocytes have been useful for developing the ultra\u00a0high-performance liquid chromatography coupled with high-resolution mass spectrometric (UHPLC-QToF-MS) methods for simultaneous determination of PQ and its metabolites generated through phase I and phase II pathways for drug metabolism.Primaquine (PQ), an 8-aminoquinoline, is the only drug approved by the United States Food and Drug Administration for radical cure and prevention of relapse in These methods were further optimized and applied for phenotyping PQ metabolites from plasma and urine from healthy human volunteers treated with single 45\u00a0mg dose of PQ. Identity of the metabolites was predicted by MetaboLynx using LC\u2013MS/MS fragmentation patterns. Selected metabolites were confirmed with appropriate standards.Besides PQ and carboxy PQ (cPQ), the major plasma metabolite, thirty-four additional metabolites were identified in human plasma and urine. Based on these metabolites, PQ is viewed as metabolized in humans via three pathways. Pathway 1 involves direct glucuronide/glucose/carbamate/acetate conjugation of PQ. Pathway 2 involves hydroxylation (likely cytochrome P450-mediated) at different positions on the quinoline ring, with mono-, di-, or even tri-hydroxylations possible, and subsequent glucuronide conjugation of the hydroxylated metabolites. Pathway 3 involves the monoamine oxidase catalyzed oxidative deamination of PQ resulting in formation of PQ-aldehyde, PQ alcohol and cPQ, which are further metabolized through additional phase I hydroxylations and/or phase II glucuronide conjugations.This approach and these findings augment our understanding and provide comprehensive view of pathways for PQ metabolism in humans. These will advance the clinical studies of PQ metabolism in different populations for different therapeutic regimens and an understanding of the role these play in PQ efficacy and safety outcomes, and their possible relation to metabolizing enzyme polymorphisms.The online version of this article (10.1186/s12936-018-2433-z) contains supplementary material, which is available to authorized users. Plasmodium vivax malaria + (intense peak), 175.0842 [M\u2009+\u2009H-C5H8O2]+, 101.0593 [M\u2009+\u2009H-C10H10N2O]+, 83.0493 [M\u2009+\u2009H-C10H12N2O2]+ , 260.177 , 243.1523 , 175.0895 , 86.0959  , 160.0619 [M\u2009+\u2009H-C8H16NO2]+, 88.0384 [M\u2009+\u2009H-C14H18N2O]+ , 147.0553 [M\u2009+\u2009H-C6H11NO]+, 86.0970 [M\u2009+\u2009H-C9H6N2O2]+ , 2-Hydroxy PQ (18) and 4-hydroxy PQ (19) with the same retention time (Table\u00a017) could not be established due to non-availability of corresponding synthetic standard.ESI-HRMS of these four metabolites gave identical molecular ion peak at m/z 453.1881 [M\u2009+\u2009H]+ which corresponds to the molecular formula C21H29N2O9. , 161.0751 [M\u2009+\u2009H-C5H11N]+, 86.0966 [M\u2009+\u2009H-C9H8N2O]+, 69.0721 [M\u2009+\u2009H-C9H11N3O]+ , 86.0966 [M\u2009+\u2009H-C12H12N2O2]+ , 175.0884 [M\u2009+\u2009H-C11H16O8]+, 101.0593 [M\u2009+\u2009H-C16H18N2O7]+  (2) and cPQ (m/z 275.1385) (5), only seven additional metabolites of PQ were detected in plasma. These include four phase I metabolites namely, hydroxy-primaquine quinone-imine (m/z 274.1549) (1), carboxyprimaquine lactam (m/z 257.1273) (3), Dihydroxy-carboxyprimaquine methylester (m/z 321.1454) (8) and carboxyprimaquine methyl ester (m/z 289.1560) (6) and three phase II metabolites, namely, glycosylated PQ (m/z 422.2315) (7), PQ N-carbamoyl glucuronide (m/z 480.1992) (4) and PQ methyl carbamate (m/z 318.1789) (9). No hydroxylated metabolites of PQ were detected in plasma.Very limited numbers of PQ metabolites were detected in plasma from the individuals treated with single dose of PQ. Besides PQ (m/z 260.1769) (2) and the major plasma metabolite cPQ (m/z 275.1385) (5) were detected in urine also. Besides these, the carboxy PQ lactam (m/z 257.1273) (3), predictably formed by spontaneous cyclization of carboxy side chain of cPQ and PQ carbamoyl glucuronide (m/z 480.1992) (4), a unique phase II metabolite formed through direct N-carbamoylation of PQ followed by glucuronidation, which were present in plasma, were also detected in urine. Glycosylated PQ (m/z 422.2315) (7), a prominent metabolite detected in plasma, was not detected in urine. Three acetylated metabolites were present in urine namely, PQ acetate (m/z 302.1875) (29), acetylated hydroxyl-PQ glucuronide conjugate (m/z 494.2127) (20) and N-Acetyl-PQ quinone-imine (m/z 316.1676) (35). Two metabolites with different retention times but identical accurate mass and MS/MS fragmentation profile corresponding to PQ glucuronide conjugate (m/z 436.2100)  were also detected in urine.The urine from the individuals treated with PQ contained large numbers of PQ metabolites. PQ (m/z 276.1694) correspond to 2-OH (18), 3-OH (21), and 4-OHPQ (14), and the fourth (17) could be 7-OH, 8-N\u2013OH, 1-N-oxide, or 8-N-oxide of PQ. Desmethy-PQ (m/z 246.1606) (19), and desmethyl-PQ glucuronide (m/z 422.1908) (12) were also detected in urine. Two metabolites predicted as PQ-quinone-imine (m/z 274.1549)  and PQ-5,6-ortho-quinone (m/z 260.1403) (10), which is spontaneously generated from 5-OH-PQ, were also detected in urine. None of these phase I metabolites were detected in plasma. Further metabolism of hydroxyl-PQ was evident from presence of two dihydroxy-PQ (m/z 292.1647) metabolites  and a dihydroxyl PQ quinone-imine (m/z 290.15) (24). Phase II conjugation metabolites, generated from further metabolism of hydroxyl PQ metabolites, detected in urine were two monohydroxy-PQ glucuronides (m/z 452.2011)  trihydroxy-PQ glucuronide (m/z 484.1932) (23) and desmethyl-PQ glucuronide (m/z 422.1908) (12).Surprisingly, four mono hydroxylated metabolites of PQ, likely to be formed through CYP mediated pathways, were detected in urine. Three of these metabolites (m/z 451.1722) (32), hydroxyl cPQ glucuronide (m/z 467.1649) (27), trihydroxy-cPQ glucuronide (m/z 499.1545) (26) and dihydroxy cPQ methylester (m/z 321.1454) (8). Hydroxy-PQ alcohol (m/z 277.1559) (33), PQ alcohol glucuronide (m/z 437.1911) (31), hydroxyl-PQ alcohol glucuronide (m/z 453.1881) (15) detected in urine may be generated through further metabolism of PQ alcohol, the metabolite from oxidative deamination of PQ side chain alkyl amine.Further metabolism of cPQ, a major plasma metabolite, through phase I and phase II pathways of drug metabolism was evident from presence of several cPQ metabolites namely, cPQ glucuronide  [N-carbamoylation and glucuronidation [2 tension yield N-carbamoyl glucuronide conjugates [N-carbamoyl glucuronide conjugate of PQ as a predominant metabolite, when PQ was incubated in vitro with primary human hepatocytes [S-PQ generated more than two-fold higher levels of the conjugate than (\u2212)-R-PQ [R-PQ to the oxidative deamination pathway [S-PQ than from (\u2212)-R-PQ. In the human hepatocyte study, glycosylated PQ was detected at time zero and thought to be a simple adduct of the primary amine group of PQ and the aldehyde form of glucose via non-enzymatic means. This metabolite could also be formed in plasma by the same mechanism. Predominant metabolism of (+)-S-PQ through these pathways may explain the somewhat lower plasma concentrations of the (+)-S-PQ, despite its negligible metabolism through oxidative deamination to cPQ [Based on the comprehensive LC-ESI-HRMS and MS/MS fragmentation profiles of PQ metabolites identified in plasma and urine of healthy human volunteers treated with single dose of 45\u00a0mg, PQ may be regarded as metabolized broadly through three pathways. Pathway 1 involves direct glucuronide/glucose/carbamate/acetate conjugation of PQ. The presence of s (UGTs) . Althougnidation , 36. Incatocytes . Formati(\u2212)-R-PQ . This ma pathway . Glycosy pathway  and was n to cPQ , 20.orthoquinone was only detected in urine but not in plasma. Recent PK and tissue distribution studies in mice showed rapid appearance of PQ-5,6-orthoquinone in high concentrations in the liver and in low concentration in plasma and its fast disappearance from the liver and circulation [orthoquinone PQ. Earlier in vitro studies have indicated the role of multiple CYP isoforms in hemolytic toxicity of PQ with predominant involvement of CYP2D6 and CYP3A4 [orthoquinone metabolite, has been linked to therapeutic efficacy of PQ [orthoquinone through CYP2D6 mediated pathway [The pathway 2  mediated pathways, generates monohydroxylated PQ metabolites. These metabolites have been implicated in therapeutic efficacy and hemolytic toxicity of PQ. Further, extension of these studies to quantitative profiling of key metabolites in malaria infected individuals treated with PQ and their correlation with therapeutic outcomes vis-\u00e0-vis CYP2D6 genotypes may be useful in determining the rational approach for application of PQ in malaria radical cure, the primary application of this drug, as well as malaria control programs.This is the most exhaustive study on identification and characterization of human metabolites of primaquine, the only approved drug for malaria radical cure. Based on overall metabolites\u2019 profile, three pathways were predicted for metabolism by humans, (a) direct glucuronide/glucose/carbamate/acetate conjugation of PQ (b) hydroxylation of PQ at different positions on the quinoline ring and (c) oxidative deamination of PQ at the terminal amine of the aminoalkyl side chain. Metabolism of PQ through direct conjugation and oxidative deamination pathways may determine pharmacokinetics and pharmacodynamics of this important antimalarial drug. Metabolism of PQ through hydroxylations on the quinolone ring, predictably catalyzed by cytochrome PAdditional file 1: Fig. 1S. Key fragments of PQ and other metabolites using LC/ESI-QToF"}
+{"text": "This article has been corrected: The authors made a mistake in size of scale bar in captions of Figures 2-5. The authors declare that this correction does not change the results or conclusions of this paper. The authors sincerely apologize for this error. The correct Figure captions are provided below. The correct scale bar size is marked in bold font.Ink4a+/CD34+) in human skeletal muscle after resistance exercise.Figure 2. Senescent endothelial progenitor cells (p16 (A) Representative immunohistochemical co-staining of muscle cross-sections (senescent cells indicated by arrows). Scale bar 55 \u03bcm. (B) Senescent endothelial progenitor cells decreased in human skeletal muscle after a single bout of resistance exercise, and to a greater extends under low protein supplemented condition. * Significant difference against Baseline, P < 0.05; \u2020 Significant difference against Low Protein, P < 0.05. Low protein: 14% protein; High protein: 44% protein in weight.+) in human skeletal muscle after resistance exercise.Figure 3. Phagocytic macrophage (CD68 (A) Representative immuno-histochemical staining of a muscle cross-section (CD68+ macrophage indicated by an arrow). Scale bar 100 \u03bcm. (B) Low protein supplementation before and after resistance exercise enhanced CD68+ macrophage infiltration in skeletal muscle above High protein trial. * Significant difference against Baseline, P < 0.05; \u2020 Significant difference against Low Protein, P < 0.05. Low protein: 14% protein; High protein: 44% protein in weight.+) in human skeletal muscle after resistance exercise.Figure 4. Regenerative macrophage (CD163 (A) Representative immunohistochemical staining of a muscle cross-section (CD163+ macrophage indicated by an arrow). Scale bar 100 \u03bcm. (B) High protein supplementation before and after resistance exercise increased CD163+ macrophage presence in human skeletal muscle 48 h after exercise. * Significant difference against Baseline, P < 0.05; \u2020 Significant difference against Low Protein, P < 0.05. Low protein: 14% protein; High protein: 44% protein in weight.Figure 5. Centrally nucleated fibers in human skeletal muscle after resistance exercise. (A) Representative hematoxylin and eosin staining of a muscle cross-section . Scale bar 100 \u03bcm. (B) (\u2020\u2020) No difference between Low and High protein trials was found. *Significant difference against Baseline, P < 0.05. Low protein: 14% protein; High protein: 44% protein in weight.1356-1365 . https://doi.org/10.18632/aging.101472Original article: Aging. 2018; 10:1356\u20131365."}
+{"text": "Streptococcus pneumoniae Serotype 15A-ST63 Clone in Japan, 2012\u20132014 . The article has been corrected online (https://wwwnc.cdc.gov/eid/article/24/2/17-1276_article).Some data were inaccurate in the text and figures in Spread of Meropenem-Resistant"}
+{"text": "The Supporting Information files were published with tracked changes. Please view the correct files below.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.S5 Table(DOCX)Click here for additional data file.S6 Table(DOCX)Click here for additional data file.S7 Table(DOCX)Click here for additional data file.S1 Fig(DOCX)Click here for additional data file.S2 Fig(DOCX)Click here for additional data file.S3 Figvs. progressors using eGFR , Gd-IgA1 biomarkers, and Oxford classification . Area under the curve, AUC = 0.936. b- Receiver operating characteristic (ROC) curve for non-progressors vs. progressors using eGFR  and Oxford classification . Area under the curve, AUC = 0.836.a- ROC curve for non-progressors (DOCX)Click here for additional data file.S4 Fig2 at the time of renal-biopsy; group 2 (n = 42), eGFR <60 mL/min/1.73 m2 at the time of renal-biopsy. Prediction equation from logistic regression (predicts probability to choose group 1): Pred(group 1) = 1 / (1 + exp(-(1517.5\u20131.2E-02*AB-IgA-24.9*eGFR))).Area under the curve = 1.00. Accuracy of the discrimination is 100%. Group 1 (n = 35), eGFR \u226560 mL/min/1.73 m(DOCX)Click here for additional data file.S5 FigS-creat, serum creatinine (\u03bcmol/L); eGFR ; serum IgG autoantibody specific for Gd-IgA1 (U/mL). Group 1 (n = 35), eGFR \u226560 mL/min/1.73 m2 at the time of renal biopsy; group 2 (n = 42), eGFR <60 mL/min/1.73 m2 at the time of renal biopsy.(DOCX)Click here for additional data file."}
+{"text": "Scientific Reports 10.1038/s41598-018-34127-x, published online 23 October 2018Correction to: This Article contains errors in Figure 1, where 4 microbial diversity patterns were omitted. The correct Fig.\u00a0As a result, the Figure legend,A. mali gut microbiotas. Alpha-diversity in bacterial  and fungal  gut communities of A. mali gut. Beta-diversity in bacterial (E) and fungal (F) gut communities estimated via principal coordinate analysis (PCoA) based on Bray-Curtis distance.\u201d\u201cMicrobial diversity patterns for should read:A. mali gut microbiotas. Alpha-diversity in bacterial  and fungal  gut communities of A. mali gut. Beta-diversity in bacterial (E) and fungal (F) gut communities estimated via principal coordinate analysis (PCoA) based on Bray-Curtis distance.\u201d\u201cMicrobial diversity patterns for"}
+{"text": "Scientific Reports 10.1038/s41598-019-54905-5, published online 05 December 2019Correction to: This Article contains a typographical error in the Acknowledgements section.\u201cMinistry of Education and Science of Russian Federation (Grant \u2116 12.1223.2017/AP to S.S. and E.R.).\u201dshould read:\u201cMinistry of Education and Science of Russian Federation (Grant No. 18-15-00172 to S.S.and E.R.)\u201d"}
+{"text": "Acinetobacter radioresistens strain DD78 (= CCUG 69565) is a soil hydrocarbon-degrading and biosurfactant-producing bacterium isolated from chronically crude oil-polluted soil of the Aconcagua River mouth in Chile. The 3.25-Mb A. radioresistens DD78 genome (41.8% GC content) was completely sequenced, with 4 replicons, 2,970 coding sequences, and 77 tRNAs. Acinetobacter radioresistens strain DD78 (= CCUG 69565) is a soil hydrocarbon-degrading and biosurfactant-producing bacterium isolated from chronically crude oil-polluted soil of the Aconcagua River mouth in Chile. The 3.25-Mb A. radioresistens DD78 genome (41.8% GC content) was completely sequenced, with 4 replicons, 2,970 coding sequences, and 77 tRNAs. Acinetobacter species are aerobic Gram-negative coccobacilli present in soil, freshwater, sediment, polluted environments, and clinical samples , improving hydrocarbon bioavailability (A. radioresistens DD78 (CCUG_69565), a soil hydrocarbon-degrading and biosurfactant-producing bacterium from crude oil-polluted soil of the Aconcagua River mouth in Central Chile, which was isolated using enrichment in diesel (0.1% [vol/vol]) in Bushnell Hass mineral medium . The PacBio reads were trimmed and assembled into 5 contigs using SMRTLink v5 and the Hierarchical Genome Assembly Process v4.0 (HGAP4.0) with default parameters , proteins for osmoprotectant glycine betaine synthesis , and the osmoprotectant transporters osmo-dependent choline transporter, sodium/proline symporter, and aspartate/alanine antiporter  were identified (>30% identity) with BLASTP using the Uniprot-KB/Swiss-Prot database. Arsenic and heavy metal  resistance genes are in pAr1. Catabolic genes encoding an alkane monooxygenase , a rubredoxin NAD(H) reductase/rubredoxin system (E3H47_10395 and E3H47_10400), and the catechol  and benzoate catabolic pathways  are chromosomal. Comparative complete 16S rRNA gene sequence analysis showed a close relationship (99.7% identity) with A. radioresistens FO-1T  and <97.5% identity with other Acinetobacter species (A. radioresistens CIP 103788T, A. lwoffii ATCC 9957T, and A. tandoii CIP 107469T showed ANIb values of 98.9%, 74.1%, and 73.6%, respectively, indicating that strain DD78 most likely belongs to the species A. radioresistens. The genome sequence of A. radioresistens DD78 provides essential data on alkane degradation, salt resistance, and biosurfactant production. samples , 2. A. b alkanes , 4. A. rlability , 6. Here78 CCUG_6565, a some sizes . The 5 cme sizes . The DD7me sizes  identifime sizes  identifi isolate . Genes e species  with A. Acinetobacter radioresistens DD78 genome sequences have been deposited in DDBJ/ENA/GenBank under the accession number GCA_005519305 and the SRA accession number SRX5548971. The version discussed here is the first version. The BioProject number for the publicly available raw data is PRJNA528312.The"}
+{"text": "Blood cultures were negative. Laparoscopic nodal biopsy showed sheets of medium-sized lymphocytes diffusely expressing CD30, TIA-1, granzyme B, and ALK, but not T-cell markers including CD2, CD3, CD4, CD5, CD7, CD8, and \u03b2F1, indicating ALK+ ALCL of null cell phenotype. Bone marrow biopsy showed two small aggregates of tumor cells in a background of normal tri-lineage hematopoiesis. ALK immunostaining revealed singly scattered positive cells (Anaplastic large cell lymphoma (ALCL) frequently involves both nodal and extranodal sites and is rarely leukemic. A 21-year-old male presented with abdominal pain. His complete blood count, which had been normal four months ago, showed increasing white cell counts from 14.9x10ve cells  in additve cells . The dis"}
+{"text": "Correction: J Intensive Carehttps://doi.org/10.1186/s40560-018-0352-2In the original publication of this article , an AuthSome data concerning specifically ECMO-treatment in patients with septic shock have been published previously in a German journal with respect to a completely different research question: Friedrichson B, Fichte J, Banjas N, Sch\u00fctz M, Hopf H-B. Extrakorporale Membranoxygenierung bei erwachsenen Patienten mit septischem Schock. An\u00e4sthesie und Intensivmed. 2018;59:698\u2013704."}
+{"text": "Haemophilus influenzae and Streptococcus pneumoniae has been implicated in the pathogenesis of otitis media, mainly in chronic and recurrent cases. We studied the \u201cin vitro\u201d biofilm production by these 2 species isolated alone or together from the nasopharynx of children with acute otitis media.Biofilm production by H. influenzae and 191\u2009S. pneumoniae strains.The studied strains were from 3 pneumococcal conjugate vaccine (PCV) periods: pre-PCV7, post-PCV7/pre-PCV13 and post-PCV13. A modified microtiter plate assay with crystal violet stain was used to study the biofilm production of 182 H. influenzae and 128/191 (66.8%) S. pneumoniae strains produced biofilm. The proportion of biofilm-producing H. influenzae strains was greater with than without the isolation of S. pneumoniae in the same sample . Conversely, the proportion of biofilm-producing S. pneumoniae strains was not affected by the presence or not of H. influenzae (66.3% vs 67.4%). S. pneumoniae serotypes 6B, 15B/C, 19A, 35F and 35B were the better biofilm producers (80%). Serotypes 11A, 14, 15A, 19F and 19A were more associated with H. influenzae biofilm-producing strains. Overall, 89/94 (94.6%) of cases with combined isolation showed biofilm production by S. pneumoniae or H. influenzae.Overall, 117/181 (64.6%) H. influenzae and S. pneumoniae strains isolated from the nasopharynx of children with acute otitis media, which reinforces the results of studies suggesting the importance of biofilm in the pathogenesis of acute otitis media.This study emphasizes the high proportion of biofilm production by The online version of this article (10.1186/s12879-018-3657-9) contains supplementary material, which is available to authorized users. Acute otitis media (AOM) is the leading cause of bacterial infections in childhood, about 700 million cases each year, and the leading cause of antibiotic prescription . More thStreptococcus pneumoniae and non-typable Haemophilus influenzae (NT-Hi)  as well as Moraxella catarrhalis : strong production >\u20091.10, moderate production 0.35\u20131.10, negative <\u20090.35.The cut-offs used to determine the intensity of biofilm production in isolates of S. pneumoniae  were classified as strong biofilm producers, 55 (30.2%) moderate producers and 64 (35.1%) non-producers. One strain could not be evaluated. Overall, 117/181 (64.6%) NT-Hi strains produced biofilm  were classified as strong biofilm producers, 64 (33.5%) moderate producers and 63 (33%) non-producers. Overall, 128/191 (66.8%) S. pneumoniae strains produced biofilm , whether isolated alone or with S. pneumoniae strains  or with S. pneumoniae strains . Overall, the proportion of NT-Hi biofilm-producing strains was greater when isolated with S. pneumoniae   or with NT-Hi strains (71.9 and 62.5% in pre-PCV7 and post-PCV13 periods). Overall, the proportion of S. pneumoniae biofilm-producing strains did not differ whether isolated alone or with NT-Hi strains   that produced biofilm was similar to that for other serotypes: 57.4% (31/54) versus 70.6% (96/137), p\u00a0=\u20090.09 (data not shown). Moreover, the proportion did not differ between the 6 additional S. pneumoniae serotypes included in PCV13 and other serotypes . Strains of serotypes 6B, 15B/C, 19A, 35F, and 35B produced biofilm in more than 80% of the cases. In contrast, strains of serotypes 23B, 23F,19F were the lowest producers (40% of strains)  of the combination. H. influenzae biofilm-producing strains were isolated more often with serotypes 11A, 14, 15A, 19F and 19A. This association did not affect biofilm production by S. pneumoniae.The p\u00a0=\u20090.44.Susceptibility or resistance to penicillin did not differ with and without biofilm production: 64.2% (61/96) versus 69.5% (66/95), -Hi strains produced biofilm when isolated with S. pneumoniae strains. Serotypes of S. pneumoniae were not associated with biofilm production by NT-Hi strains.Overall, 75.5% (72/96) of NTS. pneumoniae or H. influenzae or both produced biofilm in 94.6% (89/94) of cases  or fever + otalgia . These relations were not observed for S. pneumoniae .Table -Hi and S. pneumoniae. Among NT-Hi strains, the proportion of \u03b2-lactamase\u2013producing strains decreased from 51.5% (33/64) in the pre-PCV7 period to 17.8% (10/56) and 17.7% (11/62) in the post-PCV7/pre-PCV13 and post-PCV13 periods, respectively (p\u2009<\u20090.001). As well, the proportion of S. pneumoniae strains with decreased susceptibility to penicillin decreased from 65.7% in the pre-PCV7 period to 45.4% (29/64) and 39.7% (25/63) in the post-PCV7/pre-PCV13 and post-PCV13 periods (p\u2009=\u20090.003). Among S. pneumoniae strains, the proportion of those with erythromycin susceptibility increased from 46.8% (30/64) to 81% (51/63) between the post-PCV7/pre-PCV13 and post-PCV13 periods, respectively (p\u2009<\u20090.001).The proportion of resistant strains significantly decreased from the pre-PCV7 to post-PCV13 periods for both NT. pneumoniae serotypes changed according to the study period  with no variation in proportion for S. pneumoniae over PCV implementation periods. Of note, the proportion of NT-Hi biofilm-producing strains was greater when H. influenzae strains were isolated with S. pneumoniae, which agreed with the results of Hong et al. : strong production >\u20091.10, moderate production 0.35\u20131.10, negative <\u20090.35. Strains were classified according to agreement of at least 2 of the 3 methods used to calculate biofilm production. (XLS 195 kb)Supplementary Data S1: Different cut-offs used to determine the biofilm production by Additional file 2:H. influenzae: the different cut-offs used were as followed where AB represented the stained wells containing attached bacteria, CW the stained control wells containing bacteria-free medium only and G the bacterial growth control i) AB \u2013 CW: strong production >\u20090.30, moderate production 0.10\u20130.30, negative <\u20090.10 ii) AB / CW: strong production >\u20096, moderate production 2\u20136, negative <\u20092 iii)[(AB \u2013 CW) / G]: strong production >\u20091.10, moderate production 0.35\u20131.10, negative <\u20090.35. Strains were classified according to agreement of at least 2 of the 3 methods used to calculate biofilm production. (XLSX 76 kb)Supplementary Data S2: Different cut-offs used to determine the biofilm production by Additional file 3:. pneumoniae serotypes by study period . (DOCX 15 kb)Supplementary Data S3: Distribution of S"}
+{"text": "Scientific Reports 10.1038/s41598-017-01512-x, published online 02 May 2017Correction to: This Article contains an error in Reference 29:PLoS One12(4), e0175267 (2017)\u2019\u2018Jose Carlos Mancera Gracia, Silvie Van den Hoecke, Xavier Saelens, Kristien Van Reeth & Mirco Schmolke. Effect of serial pig passages on the adaptation of an avian H9N2 influenza virus to swine. should read:PLoS One12(4), e0175267 (2017)\u2019\u2018Mancera Gracia, J. C., Van den Hoecke, S., Saelens, X. & Van Reeth, K. Effect of serial pig passages on the adaptation of an avian H9N2 influenza virus to swine."}
+{"text": "Ralstonia solanacearum is a major phytopathogenic bacterium that attacks many crops and other plants around the world. In this study, a novel actinomycete, designated strain NEAU-SSA 1T, which exhibited antibacterial activity against Ralstonia solanacearum, was isolated from soil collected from Mount Song and characterized using a polyphasic approach. Morphological and chemotaxonomic characteristics of the strain coincided with those of the genus Streptomyces. The 16S rRNA gene sequence analysis showed that the isolate was most closely related to Streptomyces aureoverticillatus JCM 4347T (97.9%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain formed a cluster with Streptomyces vastus JCM4524T (97.4%), S. cinereus DSM43033T (97.2%), S. xiangluensis NEAU-LA29T (97.1%) and S. flaveus JCM3035T (97.1%). The cell wall contained LL-diaminopimelic acid and the whole-cell hydrolysates were ribose, mannose and galactose. The polar lipids were diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), hydroxy-phosphatidylethanolamine (OH-PE), phosphatidylinositol (PI), two phosphatidylinositol mannosides (PIMs) and an unidentified phospholipid (PL). The menaquinones were MK-9(H4), MK-9(H6), and MK-9(H8). The major fatty acids were iso-C17:0, C16:0 and C17:1 \u03c99c. The DNA G+C content was 69.9 mol %. However, multilocus sequence analysis (MLSA) based on five other house-keeping genes , DNA\u2013DNA relatedness, and physiological and biochemical data showed that the strain could be distinguished from its closest relatives. Therefore, it is proposed that strain NEAU-SSA 1T should be classified as representatives of a novel species of the genus Streptomyces, for which the name Streptomyces sporangiiformans sp. nov. is proposed. The type strain is NEAU-SSA 1T (=CCTCC AA 2017028T = DSM 105692T). Ralstonia solanacearum is the causal agent of bacterial wilt, one of the most devastating plant pathogenic bacteria around the world [he world , which hhe world , includihe world . TherefoStreptomyces, within the family Streptomycetaceae, is the largest genus of the phylum Actinobacteria, first proposed by Waksman and Henrici (1943) [http://www.bacterio.net/streptomyces.html), which are widely distributed in soils throughout the world. Therefore, members of novel Streptomyces species are in demand as sources of novel, environmentally friendly, commercially significant, naturally bioactive compounds [T with inhibitory activity against phytopathogenic bacterium Ralstonia solanacearum was isolated and subjected to the polyphasic taxonomy analysis. Results demonstrated that the strain represents a novel species of the genus Streptomyces, for which the name Streptomyces sporangiiformans sp. nov. is proposed.The actinobacteria are known to produce biologically active secondary metabolites, including antibiotics, enzymes, enzyme inhibitors, antitumour agents and antibacterial compounds ,5,6. Thei 1943)  and currompounds ,9. Durin43  and cT was isolated from soil collected from Mount Song , Dengfeng, Henan Province, China. The soil sample was air-dried at room temperature for 14 days before isolation for actinomycetes. After drying, the soil sample was ground into powder and then suspended in sterile distilled water, followed by a standard serial dilution technique. The diluted soil suspension was spread on humic acid-vitamin agar (HV) [\u22121) and nalidixic acid (20 mg L\u22121). After 28 days of aerobic incubation at 28 \u00b0C, colonies were transferred and purified on the International Streptomyces Project (ISP) medium 3 [v/v) at \u221280 \u00b0C for long-term preservation.Strain NEAU-SSA 1gar (HV)  supplemew/v) were tested in GY (Glucose-Yeast extract) medium [2PO4/0.1 M NaOH; pH 9.0\u201310.0, 0.1 M NaHCO3/0.1 M Na2CO3; and pH 11.0\u201312.0, 0.2 M KH2PO4/0.1 M NaOH. Hydrolysis of Tweens  and production of urease were tested as described by Smibert and Krieg [2S were examined as described previously [Gram staining was carried out by using the standard Gram stain, and morphological characteristics were observed using light microscopy  and scanning electron microscopy  using cultures grown on ISP 3 agar at 28 \u00b0C for 6 weeks. Samples for scanning electron microscopy were prepared as described by Jin et al. . Cultura) medium  at 28 \u00b0Cnd Krieg . The utieviously ,20.18 Column  that had a mobile phase consisting of acetonitrile: 0.05 mol L\u22121 phosphate buffer pH 7.2  at a flow rate of 0.5 mL min\u22121. The peak detection used an Agilent G1321A fluorescence detector  with a 365 nm excitation and 455 nm longpass emission filters. The whole-cell sugars were analyzed according to the procedures developed by Lechevalier and Lechevalier [18 Column  at 270 nm. The mobile phase was acetonitrile-iso-propyl alcohol . To determine cellular fatty acid compositions, the strain NEAU-SSA 1T was cultivated in GY medium in shake flasks at 28 \u00b0C for 4 days. Fatty acid methyl esters were extracted from the biomass as described by Gao et al. [Biomass for chemotaxonomic studies was prepared by growing the organisms in GY medium in shake flasks at 28 \u00b0C for 5 days. Cells were harvested using centrifugation, washed with distilled water, and freeze-dried. The isomer of diaminopimelic acid (DPA) in the cell wall hydrolysates was derivatized and analyzed using an HPLC (High Performance Liquid Chromatography) method  with an hevalier . The polhevalier . Menaquihevalier . Extracto et al.  and analT was cultured in GY medium for 3 days to the early stationary phase and harvested using centrifugation. The chromosomal DNA was extracted according to the method of sodium dodecyl sulfate (SDS)-based DNA extraction [T (1412bp) was obtained and compared with type strains available at the EzBioCloud server (https://www.ezbiocloud.net/), retrieved using NCBI BLAST  and then submitted to the GenBank database. Phylogenetic trees were constructed based on the 16S rRNA gene sequences of strain NEAU-SSA 1T and related reference species. Sequences were multiply aligned in Molecular Evolutionary Genetics Analysis (MEGA) software version 7.0 using the Clustal W algorithm and trimmed manually where necessary. Phylogenetic trees were constructed with neighbor-joining [T to its closely related strains, phylogenetic relationships of the strain NEAU-SSA 1T were also confirmed using sequences of five individual housekeeping genes  for core-genome analysis. The sequences of NEAU-SSA 1T and its related strains were obtained from the genomes or GenBank/EMBL/DDBJ (European Molecular Biology Laboratory/DNA Data Bank of Japan). GenBank accession numbers of the sequences used are given in atpD-gyrB-recA-rpoB-trpB. Phylogenetic analysis was performed as described above. Genome mining for bioactive secondary metabolites was performed using \u201cantibiotics and secondary metabolite analysis shell\u201d (antiSMASH) version 4.0 [For DNA extraction, strain NEAU-SSA 1traction . PCR amptraction ,30. The -joining  and maxi-joining  algorith-joining . The sta-joining . A dista-joining . All pos-joining . To furtsion 4.0 .T was extracted using the method of SDS-based DNA extraction [\u00ae 2.0 Fluorometer (Thermo Scientific). Whole-genome sequencing was performed on the Illumina HiSeq PE150  platform. A-tailed, ligated to paired-end adaptors, and PCR amplified samples with a 350 bp insert were used for the library construction at the Beijing Novogene Bioinformatics Technology Co., Ltd. Illumina PCR adapter reads and low-quality reads from the paired-end were filtered using a quality control step using our own compling pipeline. All good-quality paired reads were assembled using the SOAP (Short Oligonucleotide Alignment Program) denovo [https://github.com/aquaskyline) into a number of scaffolds. Then, the filter reads were handled by the next step of the gap-closing.For draft genome sequencing and assembly, the genomic DNA of strain NEAU-SSA 1traction . The har) denovo ,39 , then sheared using a JY92-II ultrasonic cell disruptor . The DNA renaturation rates were determined in 2 \u00d7 SSC at 70 \u00b0C. The experiments were performed with three replications and the DNA\u2013DNA relatedness value was expressed as a mean of the three values. Several genomic metrics are now available to distinguish between orthologous genes of closely related prokaryotes, including the calculation of average nucleotide identity (ANI) and digital DNA\u2013DNA hybridization (dDDH) values [T and S. flaveus JCM3035T (JOCU00000000) using the ortho-ANIu algorithm from Ezbiotaxon and the genome-to-genome distance calculator (GGDC 2.0) at http://ggdc.dsmz.de.Because of a lacking number of genome sequences of y et al.  under coy et al. , using atraction . The con) values ,43. In tT against two pathogenic bacteria  was evaluated using the agar well diffusion method [T, the strain was cultured in tryptone-glucose-soluble starch-yeast extract medium , and the inhibitory activity was tested. Briefly, strain NEAU-SSA 1T was inoculated into MB medium and incubated at 28 \u00b0C for seven days in a rotary shaker. The supernatant (100 mL for this study) was obtained via centrifugation at 8000 rpm and 4 \u00b0C for 10 min and subsequently extracted by using an equal volume of ethyl acetate. Then, the extract was dried in a rotary evaporator at 40 \u00b0C and eluted with proper volume methanol (1 mL used in this study). The cell precipitate was extracted with an equal volume of methanol and also condensed as above. After that, the antibacterial activity was evaluated using the agar well diffusion method, and each well contained 200 \u00b5L of the methanol extract. To examine the effect of temperature on antibacterial activity, the ten-fold dilution methanol extract was placed in a water bath at 40, 60, 80, and 100 \u00b0C for 30 min, and then cooled to room temperature. The antibacterial activity was evaluated using the agar well diffusion method.The antibacterial activity of strain NEAU-SSA 1n method  with theT showed that the strain had the typical characteristics of the genus Streptomyces. Observation of 6-week cultures of strain NEAU-SSA 1T grown on ISP 3 medium revealed that it formed well-developed, branched substrate hyphae and aerial mycelia. Sporangia consisted of cylindrical, and rough-surfaced spores (0.6\u20130.8 \u03bcm \u00d7 0.9\u20131.6 \u03bcm) were produced on aerial mycelia, but spore chains were not observed  with an optimal level of 0\u20131% (w/v). Detailed physiological characteristics are presented in the species description  (29.5%), MK-9(H6) (41.2%), and MK-9(H8) (29.4%). The cellular fatty acid profile of strain NEAU-SSA 1T was composed of iso-C17:0 (30.9%), C16:0 (26.4%), C17:1\u03c99c (19.9%), C15:0 (7.8%), C17:0 (4.4%), C14:0 (3.3%), iso-C16:0 (1.7%), anteiso-C15:0 (1.7%), C18:1\u03c99c (1.7%), C16:0 1-OH (1.2%), and iso-C18:0 (1.1%). The polar lipids of the strain consisted of diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), hydroxy-phosphatidylethanolamine (OH-PE), phosphatidylinositol (PI), two phosphatidylinositol mannosides (PIMs), and an unidentified phospholipid (PL)  . All theT were affiliated with the genus Streptomyces and most closely related to S. aureoverticillatus JCM 4347T (97.9%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain formed a cluster with S. vastus JCM4524T (97.4%), S. cinereus DSM43033T (97.2%), S. xiangluensis NEAU-LA29T (97.1%), and S. flaveus JCM3035T (97.1%) in the neighbor-joining tree  and S. flaveus JCM 3035T (7%); and the strain could grow at pH 11.0, while S. vastus JCM4524T, S. cinereus DSM 43033T, and S. xiangluensis NEAU-LA29T could not. Other phenotypic differences included the production of H2S; decomposition of cellulose; liquefaction of gelatin; growth temperature; hydrolysis of Tweens ; and utilization of L-arabinose, D-galactose, D-fructose, D-maltose, lactose, L-rhamnose, D-ribose, D-sorbitol, D-mannose, raffinose, D-xylose, myo-inositol, L-glutamine, glycine, L-threonine, L-tyrosine, L-serine, L-proline, L-asparagine, and L-arginine.Comparison of phenotypic characteristics between strain NEAU-SSA 1 strains . DiffereT is considered to represent a novel species within the genus Streptomyces, for which the name Streptomyces sporangiiformans is proposed.On the basis of morphological, physiological, chemotaxonomic, and phylogenetic results, strain NEAU-SSA 1Streptomyces sporangiiformans .w/v) NaCl. Growth was observed at temperatures between 15 and 45 \u00b0C, with an optimum temperature of 28 \u00b0C. Positive for decomposition of Tweens (40 and 80) and cellulose, hydrolysis of aesculin and starch, liquefaction of gelatin and production of urease; and negative for coagulation and peptonization of milk, hydrolysis of Tween 20, production of H2S, and reduction of nitrate. D-fructose, D-galactose, D-glucose, inositol, lactose, D-maltose, D-mannose, D-raffinose, L-rhamnose, and D-sucrose were utilized as sole carbon sources, but not L-arabinose, dulcitol, D-ribose, D-sorbitol, or D-xylose. L-alanine, L-arginine, L-asparagine, L-aspartic acid, creatine, L-glutamic acid, L-glutamine, L-proline, L-serine, and L-threonine were utilized as sole nitrogen sources, but not glycine or L-tyrosine. Cell wall contained LL-diaminopimelic acid and the whole-cell hydrolysates were ribose, mannose, and galactose. The polar lipids contained diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), hydroxy-phosphatidylethanolamine (OH-PE), phosphatidylinositol (PI), two phosphatidylinositol mannosides (PIMs), and an unidentified phospholipid (PL). The menaquinones were MK-9(H4), MK-9(H6), and MK-9(H8). Major fatty acids were iso-C17:0, C16:0, and C17:1\u03c99c.Gram-stain-positive, aerobic actinomycete that formed well-developed, branched substrate hyphae and aerial mycelia. Sporangia consisted of cylindrical and rough surfaced spores (0.6\u20130.8 \u03bcm \u00d7 0.9\u20131.6 \u03bcm) were produced on aerial mycelia, but spore chains were not observed. Good growth on ISP 3, ISP 4, ISP 7, and Nutrient agar media; moderate growth on ISP 1, ISP 2, ISP 5, ISP 6, and Czapek\u2019s agar media; and poor growth on Bennett\u2019s agar medium. Growth occurred at pH values between 6.0 and 11.0, the optimum being pH 7.0. Tolerates up to 6.0% NaCl and grows optimally in 0\u20131% (T (=CCTCC AA 2017028T = DSM 105692T), isolated from soil collected from Mount Song, Dengfeng, Henan Province, China. The DNA G+C content of the type strain was 69.9 mol %, calculated from the assembly for the draft genome sequence. The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain NEAU-SSA 1T is MH842151. This Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under the accession VCHX00000000. The version described in this paper is version VCHX00000000.2.The type strain was NEAU-SSA 1T exhibited antibacterial activity against Ralstonia solanacearum with inhibitory zone diameters of 23 mm  suggested that strain NEAU-SSA 1T belonged to the genus Streptomyces. Physiology and biochemical characteristics, together with DDH relatedness values and ANI values, clearly indicated that strain NEAU-SSA 1T could be differentiated from the closely related strains S. aureoverticillatus JCM 4347T, S. vastus JCM 4524T, S. cinereus DSM 43033T, S. xiangluensis NEAU-LA29T, and S. flaveus JCM 3035T. Based on the polyphasic analysis, it is proposed that strain NEAU-SSA 1T should be classified as representatives of a novel species of the genus Streptomyces, for which the name Streptomyces sporangiiformans sp. nov. is proposed. The type strain is NEAU-SSA 1T (=CCTCC AA 2017028T = DSM 105692T).A novel strain NEAU-SSA 1"}
+{"text": "These articles have been corrected: The Figure 2G in \u2018Upregulation of long non-coding RNA PlncRNA-1 promotes proliferation and induces epithelial-mesenchymal transition in prostate cancer,\u2019 (https://doi.org/10.18632/oncotarget.15318), is a duplication of Figure 3F in the published article \u2018PlncRNA-1 induces apoptosis through the Her-2 pathway in prostate cancer cells,\u2019 (https://doi.org/10.4103/1008-682X.178849). The proper Figure 2G is shown below. The authors declare that these corrections do not change the results or conclusions of this paper.26090-26099.https://doi.org/10.18632/oncotarget.15318Original article: Oncotarget. 2017; 8:26090\u201326099."}
+{"text": "Lysinibacillus sp. OL1 and Enterococcus sp. OL5, were isolated from boron fertilizer-amended cauliflower plantation field soils in India. Here, we report the draft genome sequences of OL1 (4.87\u2009Mb) and OL5 (3.93\u2009Mb) to explore the intricacies of boron tolerance in bacteria.Two novel boron-tolerant, arsenic-resistant, Gram-positive bacterial strains,  Lysinibacillus sp. OL1 and Enterococcus sp. OL5, were isolated from boron fertilizer-amended cauliflower plantation field soils in India. Here, we report the draft genome sequences of OL1 (4.87\u2009Mb) and OL5 (3.93\u2009Mb) to explore the intricacies of boron tolerance in bacteria.Two novel boron-tolerant, arsenic-resistant, Gram-positive bacterial strains,  The enriched culture was dilution plated on boric acid-supplemented tryptone soya agar (TSA) for isolation and screening of boron-tolerant strains, as described previously (3BO3 and sodium arsenite (NaAsO2), respectively, was tested in TSA supplemented with different concentrations of boron or arsenic , following methods described previously (Boron (B) occurs in soil solution as a nonionized molecule (Ht growth . Boron-t arsenic  with minhttp://www.usadellab.org/cms/?page=trimmomatic) high-quality data, which were subsequently assembled into 179 scaffolds  using SPAdes v3.11.1 with default parameters (http://hannonlab.cshl.edu/fastx_toolkit/download.html). For OL5, a genomic DNA library was constructed using an Ion Xpress Plus fragment library kit  and sequenced on a 530 chip of the Ion S5 system (Thermo Fisher Scientific). Before sequence retrieval from Ion S5, low-quality and polyclonal reads were filtered out and the adaptor sequences trimmed using the inbuilt PGM software. A total of 3,993,359\u2009bp of adapter-free, quality-filtered reads  were obtained and subsequently assembled into 46 contigs  using SPAdes v3.13.0 with default parameters  under the accession numbers PRJNA521315 and PRJNA549579.These whole-genome shotgun projects have been deposited at DDBJ/ENA/GenBank under the accession numbers"}
+{"text": "HERC family emerged >595 million years ago and has undergone gene duplication and gene loss events throughout its evolution. The structural topology of the RCC1-like domain and HECT domains from all HERC paralogs is highly conserved among evolutionarily diverse vertebrates despite low sequence homology. Functional analyses showed that the human small HERCs exhibit different degrees of antiviral activity toward HIV-1 and that HERC5 provides the strongest inhibition. Notably, coelacanth HERC5 inhibited simian immunodeficiency virus (SIV), but not HIV-1, particle production, suggesting that the antiviral activity of HERC5 emerged over 413 million years ago and exhibits species- and virus-specific restriction. In addition, we showed that both HERC5 and HERC6 are evolving under strong positive selection, particularly blade 1 of the RCC1-like domain, which we showed is a key determinant of antiviral activity. These studies provide insight into the origin, evolution, and biological importance of the human restriction factor HERC5 and the other HERC family members.In humans, homologous to the E6-AP carboxyl terminus (HECT) and regulator of chromosome condensation 1 (RCC1)-like domain-containing protein 5 (HERC5) is an interferon-induced protein that inhibits replication of evolutionarily diverse viruses, including human immunodeficiency virus type 1 (HIV-1). To better understand the origin, evolution, and function of HERC5, we performed phylogenetic, structural, and functional analyses of the entire human small-HERC family, which includes HERC3, HERC4, HERC5, and HERC6. We demonstrated that the IMPORTANCE Intrinsic immunity plays an important role as the first line of defense against viruses. Studying the origins, evolution, and functions of proteins responsible for effecting this defense will provide key information about virus-host relationships that can be exploited for future drug development. We showed that HERC5 is one such antiviral protein that belongs to an evolutionarily conserved family of HERCs with an ancient marine origin. Not all vertebrates possess all HERC members, suggesting that different HERCs emerged at different times during evolution to provide the host with a survival advantage. Consistent with this, two of the more recently emerged HERC members, HERC5 and HERC6, displayed strong signatures of having been involved in an ancient evolutionary battle with viruses. Our findings provide new insights into the evolutionary origin and function of the HERC family in vertebrate evolution, identifying HERC5 and possibly HERC6 as important effectors of intrinsic immunity in vertebrates. Vertebrates possess multiple defense mechanisms to inhibit the replication of viruses. This defense system is largely composed of specialized hematopoietic cells that react nonspecifically to pathogens (innate immunity), an antibody-dependent and cell-mediated response (adaptive immunity), and core cellular effector proteins called restriction factors (intrinsic immunity). Restriction factors are considered to be the front line of defense against viral infection, since their activity typically does not require virus-triggered signaling or intercellular communication . The imp3\u2013Retroviridae, Orthomyxoviridae, Flaviviridae, Togaviridae, Herpesviridae, Poxviridae, Arteriviridae, and Pneumoviridae (\u201328\u201332\u201332\u2013Interferon-stimulated gene 15 (ISG15) and/or its conjugation to newly translated proteins (referred to as ISGylation) exhibits broad antiviral activity toward evolutionarily diverse viruses, including those belonging to the families oviridae \u201331. The idae \u201328\u2013, 32\u201336. By virtue of their RCC1-like domains, HERCs also belong to the phylogenetically widespread RCC1 superfamily of proteins , 39. The32\u2013https://genome.ucsc.edu]) and NCBI gene and protein sequence databases for the presence of small HERC gene members. The oldest small-HERC member is HERC4, which is present in one of the only surviving lineages of jawless fish, sea lampreys (originating \u223c595 million years ago [mya]) , just after the small-HERC family expanded with the duplication and divergence of HERC4 , giving rise to two different chromosomal HERC loci in most vertebrates, where the HERC3-HERC5-HERC6 locus is flanked by FAM13A and PIGY-PYURF and HERC4 by MYPN and SIRT1  . To bettngements . This alngements . A singlof HERC4 . HERC3 and SIRT1 .HERC family expanded further after the divergence of cartilaginous fish (\u223c430 mya), with the emergence of HERC6, which is present in most jawed vertebrates with an apparent absence in platypus and some fish  and bird  species (HERC family occurred after the divergence of ray-finned fish (\u223c413 mya), with the likely duplication of HERC6, giving rise to HERC5 , metatherian  mammals , and rodents    43, 45), 45HERC HERC sequences showed segregation of the small HERC genes into four major clusters consisting of HERC3, HERC4, HERC5, and HERC6 . Notably, coelacanth and lizard HERC5 sequences clustered on their own, showing more sequence similarity to HERC6 genes than to other HERC5 genes, possibly indicating that these genes are actually HERC6 or a hybrid of HERC5 and HERC6. Similar tree topologies regarding the main branching were predicted using several tree-generating algorithms . Consistent with the approximate emergence times of the small-HERC family members shown in HERC4 is the oldest of the HERC paralogs, followed by HERC3, HERC6, and then HERC5.Phylogenetic analysis of nd HERC6 . Most HEH structural measure derived from alignment of the predicted tertiary structures of RCC1-like domains (https://bmm.crick.ac.uk/~populus/) and showed that each of the RCC1-like domains of HERC3 to -6 adopted the characteristic \u03b2-propeller structure of the superfamily, despite their low sequence homology  constructs for the ability to knock down endogenous HERC mRNA and protein levels. As shown in HOS-CD4-CXCR4 cells were cotransfected with plasmids carrying HERC shRNA and HIV-1 R9 (a full-length replication-competent NL4-3 derivative). After 72 h of replication (\u223c2 or 3 rounds), quantitative Western blot analysis of cell lysates or virions produced from cells showed that cells knocked down for HERC5 expression exhibited a significant increase in Gag particle production (\u223c8-fold), whereas HERC3, -4, and -6 released modestly more virions into the supernatant than the control cells (\u223c2-fold) . The amo53\u2013To test the effect of increased HERC expression on single-round HIV-1 particle production, human 293T cells, which do not support multiround replication, were cotransfected with plasmids carrying HIV-1 (R9) and either empty vector, HERC3, HERC4, HERC5 or HERC6. As expected, HERC5 potently inhibited HIV-1 particle production . HERC3 oWe previously showed that human HERC5 blocked nuclear export of Rev-dependent HIV-1 RNA . To detecis-acting element called the Rev-response element (RRE) located within an HIV-1 intron  and Rev-independent  constructs, as previously described , 58. HIV1 intron . HIV-1 m1 intron . Success1 intron  and F. THERC5, we tested the ability of coelacanth HERC5 to inhibit HIV-1 virus production. To assess potential virus-specific effects, we also tested the antiviral activity toward another, related nonhuman retrovirus, simian immunodeficiency virus (SIV) , which is thought to be at least 32,000 years older than HIV-1  or blade 1 (H6:H5blade1) from human HERC5 with the corresponding region in human HERC6. We then measured the abilities of these HERC6 mutants to inhibit HIV-1 particle production. As shown in HERC family has an ancient marine origin, where HERC4 emerged at least 595 mya and expansion of the family occurred sometime after the divergence of jawed vertebrates from jawless vertebrates (\u223c476 to 595 mya). Elephant sharks are among the oldest and most slowly evolving jawed vertebrates and have accumulated a small number of chromosomal rearrangements  comprise a substantial portion of vertebrate genomes and appear to have an ancient marine origin, with evidence of ERV sequences found in the genomes of elephant shark, coelacanth, and possibly lamprey , resulting in a smaller repertoire of immune genes than in humans . However, HERC3 and HERC4 do exhibit differential tissue-specific expression  supplemented with 10% heat-inactivated fetal bovine serum (FBS), 100 U/ml penicillin, and 100 \u03bcg/ml streptomycin at 37\u00b0C with 5% COHomo sapiens (human) (NP_055421.1), Gorilla gorilla gorilla (gorilla) (XP_004039158.1), Pan troglodytes (chimpanzee) (XP_517337.3), Nomascus leucogenys (gibbon) (XP_003265945.1), Papio anubis (baboon) (XP_003898995.1), Saimiri boliviensis boliviensis (squirrel monkey) (XP_003924058.1), Callithrix jacchus (marmoset) (XP_002745644.1), Bos taurus (cow) (NP_001077132.1), Ovis aries (sheep) (XP_004009758.1), Felis catus (cat) (XP_003985228.1), Canis lupus familiaris (dog) (XP_535653.3), Ailuropoda melanoleuca (giant panda) (XP_002913643.1), Equus caballus (horse) (XP_001496703.3), Callorhinchus_milii (elephant shark) (XM_007902532.1), Danio rerio (zebrafish) (NM_001145624.1), Takifugu rubripes (pufferfish) (XM_011612933.1), Oreochromis niloticus (tilapia) (XM_005456948.2), Latimeria chalumnae (coelacanth) (XM_006005071.2), Xenopus tropicalis (frog) (XM_002938630.3), Anolis carolinensis (lizard) (XM_008111037.2), Geospiza fortis (finch) (XM_005418124.1), Meleagris gallopavo (turkey) (XM_003205471.2), Gallus gallus (chicken) (XM_015276202.1), Monodelphis domestica (opossum) (XM_007495699.2), Mus musculus (mouse) (NM_028705.3), and Dasypus novemcinctus (armadillo) (XM_004480548.2); for HERC4, human (NP_071362.1), gorilla (XP_004049535.1), chimpanzee (XP_001167753.1), gibbon (XP_003258260.1), baboon (XP_003903924.1), squirrel (XP_003928728.1), marmoset (XP_002756356.1), cow (NP_001070362.2), sheep (XP_004021455.1), dog (XP_849808.1), panda (XP_002913788.1), horse (XP_001503636.1), elephant shark (XM_007897346.1), zebrafish (XM_005173035.3), pufferfish (XM_011602912.1), tilapia (XM_005473848.2), coelacanth (XM_005992449.2), frog (NM_001128650.1), lizard (XM_008115175.2), finch (XM_005428501.2), turkey (XM_010714265.1), chicken (XM_015278711.1), Ornithorhynchus anatinus (platypus) (XM_007667149.1), opossum (XM_016421882.1), mouse (NM_030114.2), armadillo (XM_012529817.1), and Petromyzon marinus (lamprey) ENSPMAG00000001626; for HERC5, human (NP_057407.2), chimpanzee (XP_003310459.1), gorilla (XP_004039179.1), marmoset (XP_002745648.1), baboon (XP_003898997.1), squirrel monkey (XP_003924055.1), gibbon (XP_003265940.1), horse (XP_001915115.2), giant panda (XP_002913645.1), sheep (XP_004009762.1), cow (NP_001095465.1), dog (XP_535652.3), cat (XP_003985249.1), coelacanth (XM_014498805.1), lizard (XM_008111035.2), and armadillo (XM_012520464.1); for HERC6, human (NP_060382.3), gorilla (XP_004039178.1), chimpanzee (XP_001160851.1), gibbon (XP_003265938.1), baboon (XP_003899001.1), squirrel (XP_003924053.1), marmoset (XP_002745681.1), cow (NP_001179573.1), sheep (XP_004010096.1), cat (XP_003985250.1), dog (XP_851549.1), horse (XP_001494887.1), pufferfish (XM_011618146.1), tilapia (XM_005474674.1), coelacanth (XM_014498807.1), frog (XM_002938624.2), lizard (XM_008111034.2), opossum (XM_007495704.1), mouse (NM_025992.2), and armadillo (XM_012520459.1).The MEGA 7.0 package was used for phylogenetic analysis . The amiCallorhinchus milii 6.1.3 GCF_000165045.1 (elephant shark), Danio rerio GRCz10 GCF_000002035.5 (zebrafish), Takifugu rubripes FUGU5 GCF_000180615.1 (pufferfish), Oreochromis niloticus Orenil1.1 GCF_000188235.2 (tilapia), Latimeria chalumnae LatCha1 GCF_000225785.1 (coelacanth), Xenopus tropicalis Xtropicalis_v7 GCF_000004195.2 (frog), Anolis carolinensis AnoCar2.0 GCF_0000090745.1 (lizard), Geospiza fortis GeoFor_1.0 GCF_000277835.1 (finch), Meleagris gallopavo Turkey_5.0 GCF_000146605.2 (Turkey), Gallus_gallus-5.0 GCF_000002315.4 (chicken), Ornithorhynchus_anatinus-5.0.1 GCF_000002275.2 (platypus), Monodelphis domestica MonDom5 GCF_000002295.2 (opossum), Dasypus novemcinctus Dasnovv3.0 GCF_000208655.1 (armadillo), Loxodonta africana Loxafr3.0 GCF_000001905.1 (elephant), Mus musculus GRCm38.p3 GCF_000001635.23 (mouse), Rattus norvegivcus Rnor_6.0 GCF_000001895.5 (rat), Elephantulus edwardii EleEdw1.0 GCF_000299155.1 (shrew), Chrysochloris asiatica ChrAsi1.0 GCF_000296735.1 (cape golden mole), Canis lupus familiaris CanFam3.1 GCF_000002285.3 (dog), and Homo sapiens GRCh38.p2 GCF_000001405.28 (human).Synteny maps were derived using the following reference assemblies: http://www.ncbi.nlm.nih.gov/tools/cobalt/) (Homo sapiens (human) (NP_055421.1), Gorilla gorilla gorilla (gorilla) (XP_004039158.1), Pan troglodytes (chimpanzee) (XP_517337.3), Nomascus leucogenys (gibbon) (XP_003265945.1), Papio anubis (baboon) (XP_003898995.1), Saimiri boliviensis boliviensis (squirrel monkey) (XP_003924058.1), Callithrix jacchus (marmoset) (XP_002745644.1), Bos taurus (cow) (NP_001077132.1), Ovis aries (sheep) (XP_004009758.1), Felis catus (cat) (XP_003985228.1), Canis lupus familiaris (dog) (XP_535653.3), Ailuropoda melanoleuca (giant panda) (XP_002913643.1), and Equus caballus (horse) (XP_001496703.3); for HERC4, human (NP_071362.1), gorilla (XP_004049535.1), chimpanzee (XP_001167753.1), gibbon (XP_003258260.1), baboon (XP_003903924.1), squirrel (XP_003928728.1), marmoset (XP_002756356.1), cow (NP_001070362.2), sheep (XP_004021455.1), dog (XP_849808.1), panda (XP_002913788.1), and horse (XP_001503636.1); for HERC5, human (NP_057407.2), chimpanzee (XP_003310459.1), gorilla (XP_004039179.1), marmoset (XP_002745648.1), baboon (XP_003898997.1), squirrel monkey (XP_003924055.1), gibbon (XP_003265940.1), horse (XP_001915115.2), giant panda (XP_002913645.1), sheep (XP_004009762.1), cow (NP_001095465.1), dog (XP_535652.3), and cat (XP_003985249.1); for HERC6, human (NP_060382.3), gorilla (XP_004039178.1), chimpanzee (XP_001160851.1), gibbon (XP_003265938.1), baboon (XP_003899001.1), squirrel (XP_003924053.1), marmoset (XP_002745681.1), cow (NP_001179573.1), sheep (XP_004010096.1), cat (XP_003985250.1), dog (XP_851549.1), and horse (XP_001494887.1). At least 2 independent sequences were available for human, sheep, baboon, marmoset, gibbon, and squirrel monkey. The following sequences were not independently validated: cat, dog, cow, horse, sheep, and giant panda. The identification of site-specific positive selection and purifying selection was generated using the Selecton server (http://selecton.tau.ac.il/index.html). The HERC5 phylogenetic tree was used in the Selecton analysis. Nested pairs of models  and a nonnested pair (M8a and MEC) were compared using the likelihood ratio test implemented in the Selecton program.Positive-selection analysis was performed as previously described . HERC secobalt/) . The folPlasmids encoding Flag-tagged HERC3, HERC4, HERC5, and HERC6 were created by first PCR amplifying the various HERC coding regions from their respective templates (as described above) using the following primers: for HERC3, forward, 5\u2032 ACG TGA ATT CCA TGT TAT GTT GGG GAT ATT GG 3\u2032, and reverse, 5\u2032 ACG TGG TAC CTC AGG CCA AAC TAA ACC CTT CAT AGT TGT C 3\u2032; for HERC4, forward, 5\u2032ACG TGA ATT CTA TGT TGT GCT GGG GAA ATG C 3\u2032, and reverse, 5\u2032 ACG TTC TAG ATT ATA TTA AAC TGA AGC CTT CAT TGT G 3\u2032; for HERC5, forward, 5\u2032 AAT CGA GAT CTT ATG GAG CGC CGC AGC 3\u2032, and reverse, 5\u2032 TAT GCG GAT CCT CAG CCA AAT CCT CTG 3\u2032; and for HERC6, forward, 5\u2032 AGA TAA GAT CTT ATG TAC TTC TGT TGG GGC 3\u2032, and reverse, 5\u2032 TAG GAG ATA TCT TAT GAC TGT GTG AGC ATG 3\u2032. The amplified products were then cloned into p3xFLAG-CMV10 using the following restriction enzymes: for HERC3, EcoRI and KpnI; for HERC4, EcoRI and XbaI; for HERC5, BglII and BamHI; and for HERC6, BglIII and EcoRV. The resulting plasmids were named pHERC3, pHERC4, pHERC5, and pHERC6.To generate pH 6:H5RLD, the HERC5 RCC1-like domain was PCR amplified from pHERC5 using the following primers: forward, 5\u2032 GGA TGA CGA TGA CAA GAT GGA GCG CCG CAG CC 3\u2032, and reverse, 5\u2032 TAT GTT CCA GCA AAA ATT ATT AAC TCC TTT TCT GAG GTA TGG CTT TCA AG 3\u2032. The backbone of pHERC6 was PCR amplified using the following primers: forward, 5\u2032 TTT TTG CTG GAA CAT ATG CCA ACT TTG 3\u2032, and reverse, 5\u2032 CTT GTC ATC GTC ATC CTT GTA ATC GAT G 3\u2032. The two amplified fragments were cloned using the fast cloning technique .To generate pH6:H5blade1, blade 1 of HERC5 (amino acids 1 to 100) was PCR amplified from pHERC5 using the following primers: forward, 5\u2032 GGA TGA CGA TGA CAA GAT GGA GCG CCG CAG CCG CCC CAA CAG AAG TAC ATC TTG TCA TCG TCA TCC TTG TAA TCG ATG 3\u2032, and reverse, 5\u2032 GCT CCT TCC CGC AGC TCA CGT GGA TCT TCA TGT TCT TGC CCA GC 3\u2032. The backbone of pHERC6 was PCR amplified using the following primers: forward, 5\u2032 GCT GGG CAA GAA CAT GAA GAT CCA CAG CTG CGG GAA GGA GCA C 3\u2032, and reverse, 5\u2032 GGC TGC GGC GCT CCA TCT TGT CAT CGT CAT CCT TGT AAT CGA TG 3\u2032. The two amplified fragments were cloned using the Gibson cloning technique per the manufacturer's instructions (New England Biolabs).To generate pH6:R10A, site-directed mutagenesis was performed on pHERC6 using the following primers: forward, 5\u2032 TTC TGT TGG GGC GCC GAC TCC GCG GAG CTG CAG CGC CGG AGG 3\u2032, and reverse, 5\u2032 CCT CCG GCG CTG CAG CTC CGC GGA GTC GGC GCC CCA ACA GAA 3\u2032. pH6:E67A was generated similarly using the following primers: forward, 5\u2032 GCA GCG CGG GGA GCT GCC AGC ACC AAT TCA GGC ATT GGA AAC C 3\u2032, and reverse, 5\u2032 GGT TTC CAA TGC CTG AAT TGG TGC TGG CAG CTC CCC GCG CTG C 3\u2032. pH6:R10A/E67A was generated similarly, except pH6:R10A was used as the template with the following primers: forward, 5\u2032 GCA GCG CGG GGA GCT GCC AGC ACC AAT TCA GGC ATT GGA AAC C 3\u2032, and reverse, 5\u2032 GGT TTC CAA TGC CTG AAT TGG TGC TGG CAG CTC CCC GCG CTG C 3\u2032. pR10G was generated similarly, using the following primers: forward, 5\u2032 CGC CGA CTC CGG GGA GCT GCA 3\u2032, and reverse, 5\u2032 TGC AGC TCC CCG GAG TCG GCG 3\u2032. pHERC3-\u0394RLD was generated similarly, using the following primers: forward, 5\u2032 GAC GAT GAC AAG ATG AGC TCA CCA CCA GAT GTT GAA G 3\u2032, and reverse, 5\u2032 CAT CTG GTG GTG AGC TCA TCT TGT CAT CGT CAT CCT TGT AAT CG 3\u2032. pHERC4-\u0394RLD was generated similarly, using the following primers: forward, 5\u2032 ACG ATG ACA AGA TGA ATT GGT ACC CCT ATA ATG GGC AGT G 3\u2032, and reverse, 5\u2032 TAG GGG TAC CAA TTC ATC TTG TCA TCG TCA TCC TTG TAA TCG 3\u2032. pHERC5-\u0394RLD was generated previously . pHERC6-HERC3, HERC4, HERC5, and HERC6 were obtained from the following sources: HERC3 (NM_014606.2), HERC6 (NM_017912.3) (HERC5 (NP_057407.2) (HERC4 (NM_015601.3) was isolated from HeLa cells by first reverse transcribing total RNA and then PCR amplification of cDNA using the following primers: forward, 5\u2032ACG TGA ATT CTA TGT TGT GCT GGG GAA ATG C 3\u2032, and reverse, 5\u2032 ACG TTC TAG ATT ATA TTA AAC TGA AGC CTT CAT TGT G 3\u2032. All the constructs were verified by sequencing. Transfections were performed using Lipofectamine 2000 (Invitrogen) according to the manufacturer's instructions unless otherwise indicated. Cotransfections of HERC plasmids with pR9 were performed at a ratio of 10:1 unless otherwise noted. Standard Western blot analyses were performed as previously described  , and HER57407.2) . HERC4 (escribed . DensitoSaccharomyces cerevisiae-based cloning of diverse HIV-1 strains (S. cerevisiae Hanson (MYA-906) (MAT\u03b1 ade6 can1 his3 leu2 trp1 URA3) was obtained from the American Type Culture Collection (ATCC). The yeast was grown at 30\u00b0C in appropriate medium , depending on the cloning step. Transformations/recombinations were performed using the lithium acetate (LiAc) method. Briefly, the linearized vector DNA (\u223c1 \u03bcg) and PCR product (\u223c3 \u03bcg) were added to competent cells at a 1:3 ratio, along with 50 \u03bcg of single-stranded salmon sperm carrier DNA  and sterile polyethylene glycol (50%)-TE -LiAc (100 mM). Following agitation for 30 min at 30\u00b0C, the yeast was heat shocked at 42\u00b0C for 15 min and plated on C-leu agar plates containing the appropriate selection.For the construction of pSIVmac239 (pREC_nfl_SIV239), the SIVmac239 Spx vector was obtained from the NIH AIDS Reagent Program. We previously constructed pREC_nfl_HIV and pCMV_cplt vectors for  strains . We deve strains . For yeaThe following reagents were obtained through the NIH AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH: HIV-1 p24 monoclonal antibody (183-H12-5C) from Bruce Chesebro and Kathy Wehrly and anti-SIVmac p17 monoclonal antibody (KK59) from Karen Kent and Caroline Powell. Anti-FLAG and anti-hemagglutinin (HA) (clone 3F10) were purchased from Sigma, anti-myc and anti-\u03b2-actin were purchased from Rockland, anti-eGFP was purchased from Clontech, and anti-GAPDH  (clone 6C5) was purchased from EMD/Millipore. Anti-HERC3 (H00008916-B01P), anti-HERC4 (H00026091-A01), anti-HERC5 (H00051191-A01), and anti-HERC6 (H00055008-A01) were purchased from Abnova.TC) for each PCR. The gene-specific forward and reverse primer sets used were as follows: Gag- ; Rev . Quantification of endogenous HERC mRNA was run on the QuantStudio5 qPCR machine (Applied Biosystems) under the following cycling conditions: 2 min at 95\u00b0C and 40 cycles of 5 s at 95\u00b0C, 10 s at 60\u00b0C, and 20 s at 72\u00b0C. QuantStudio design and analysis desktop software (version 1.4) was used to determine the TC for each PCR. The primer pairs were as follows: HERC3 , HERC4 , HERC5 , HERC6 , GAPDH , and eGFP . To ensure no carryover of DNA into each total purified RNA sample, 3 \u03bcg of the purified RNA was used directly as the template without reverse transcription for qPCR, using the primer sets described above.Total RNA was extracted using the PureLink RNA minikit . Three micrograms of RNA was reverse transcribed to cDNA using Moloney murine leukemia virus (MMLV) reverse transcriptase and oligo(dT) primers (Life Technologies). Prior to qPCR, the cDNA samples were diluted 1:5 with water. Each PCR mixture consisted of 10 \u03bcl of SYBR green master mix, 2 \u03bcl of gene-specific primers (1 \u03bcl of 10 \u03bcM forward primer and 1 \u03bcl of 10 \u03bcM reverse primer), 5 \u03bcl of diluted cDNA, and water to a total volume of 20 \u03bcl. For quantification of incompletely and fully spliced HIV RNAs, qPCR was run on the Rotor-Gene 6000 qPCR machine (Corbett Life Science) under the following cycling conditions: 10 min at 95\u00b0C and 40 cycles of 10 s at 95\u00b0C, 15 s at 60\u00b0C, and 20 s at 72\u00b0C. The Rotor-Gene 6000 series software (version 1.7) was used to determine the cycle threshold (P values and statistical tests used are mentioned where appropriate. P values of less than 0.05 were deemed significant.GraphPad Prism v6 was used for all statistical analyses mentioned in the text. The"}
+{"text": "Xueshuan Xinmaining Tablet (XXT) is a widely used traditional Chinese medicine for the treatment of stroke, chest pain, coronary heart disease, and angina pectoris caused by blood stasis. Having a multiple-component preparation, it is still far from meeting the requirements of modernization and standardization because its detailed chemical basis and action mechanism have not been clarified. In this work, the different batches of XXT samples were analyzed by HPLC and the typical sample was analyzed by UPLC-ESI-Q-TOF/MS to understand its chemical profiling. As a result, 77 chromatographic peaks were detected, among which 63 constituents were identified or tentatively characterized based on the comparison of retention time and UV spectra with authentic compounds as well as by summarized MS fragmentation rules and matching of empirical molecular formula with those of published components. This is the first systematic report on the chemical profiling of the commercial XXT products, which provides the sufficiently chemical evidence for the global quality evaluation of XXT products. Chuanxiong rhizoma), Huaihua (Sophorae flos), Danshen , Shuizhi (Hirudo), Maodongqing (Hairy holly root), Rengong Niuhuang , Rengong Shexiang (Moschus artifactus), Renshen Jingye Zongzaogan , Bingpian (Borneolum syntheticum), and Chansu (Bufonis venenum) + (ESI+) or [M+HCOO]\u2212 (ESI-) were detected in MS spectrum. In MS/MS spectrum, a series of fragmentation ions were observed due to the neutral loss of glucose, rhamnose, arabinose, or xylose  were unambiguously identified as ginsenoside Rg1, ginsenoside Re, ginsenoside Rb2, ginsenoside Rb3, 20S-ginsenoside F1, ginsenoside Rd, 20S-ginsenoside F2, and 20S-ginsenoside Rh2. Unknown ginsenoside saponins followed the similar dissociation pathways. For the ginsenosides of protopanaxadiol, the ions at m/z 407, 425, and 443 could be found in the positive ion mode. Meanwhile the ginsenosides of protopanaxatriol showed the ions at m/z 405, 423, and 441 in the positive ion mode.lose see . These f1 showed [M+Na]+ ion at m/z 823.4824 (C42H72O14Na) in the positive ion mode and [M+HCOO]\u2212 ion at m/z 845.4895 (C43H73O16) in the negative ion mode. The fragmentation ions at m/z 621.4368 and 603.4259 were produced from the protonated ion [M+H]+ by losing glucose and the further loss of H2O. In the negative MS/MS analysis, [M-H]\u2212 at m/z 799.4838 gave the characteristic ion at m/z 637.4307 [M-H-glc]\u2212and 475.3773[M-H-2glc]\u2212. Compounds 23 and 40 showed the quasimolecular ions at m/z 823 [M+Na]+ and 845 [M+HCOO]\u2212 in the full mass spectrum. Compound 23 has the same retention time and MS data as those of reference ginsenoside Rg1 and it was identified as ginsenoside Rg1. As for compound 40, although the fragmentation ions at m/z 405, 423, or 441 could not be detected in the MS/MS spectrum, it was temporarily identified as ginsenoside Rf.Ginsenoside Rg+ ion at m/z 969.5427 (C48H82O18Na) in the positive ion mode and [M+HCOO]\u2212 ion at m/z 991.5482 (C49H83O20) in the negative ion mode. The fragmentation ions at m/z 945.5428 [M-H]\u2212, 799.4845 [M-H-rha]\u2212, 783.4893 [M-H-glc]\u2212, and 637.1430 [M-H-rha-glc]\u2212 were found in MS/MS spectrum in the negative ion mode. Compound 24 showed similar retention time and MS data as those of reference ginsenoside Re and it was identified as ginsenoside Re.Ginsenoside Re showed [M+Na]2, [M+Na]+ ion at m/z 1101.5868 (C53H90O22Na) in the positive ion mode and [M+HCOO]\u2212 ion at m/z 1123.5908 (C54H91O24) in the negative ion mode are shown in the full mass spectrum. The fragmentation ions at m/z 945.5425 [M-H-ara]\u2212 and 783.4889 [M-H-ara-glc]\u2212 were detected in MS2. Reference ginsenoside Rb3  and 683.4375 [M+HCOO]\u2212 (C37H63O11Na). In the negative MS/MS experiment, the fragmentation ion at m/z 475.3785 [M-H-glc]\u2212 was detected. Compound 49 showed similar retention time and MS data as those of reference 20S-ginsenoside F1.20 m/z 969.5379 [M+Na]+ and 991.5495 [M+HCOO]\u2212 in the full mass spectrum. The fragmentation ions at m/z 783.4892 [M-H-glc]\u2212, 621.4368 [M-H-2glc]\u2212, and 459.3842 [M-H-3glc]\u2212 were produced by losing glucose. Compound 53 was identified as ginsenoside Rd by displaying the same retention time and MS information.Reference ginsenoside Rd showed the adduct ions atS-ginsenoside F2, it showed the quasimolecular ions at m/z 807.4890 [M+Na]+ and 829.4949 [M+HCOO]\u2212. The deprotonated ion peak at m/z 783.4881 [M-H]\u2212 produced the fragmentation ion at m/z 621.4355 [M-H-glc]\u2212 in MS2. Compounds 45, 61, 64, and 65 gave the same adduct ions [M+Na]+ in positive MS and [M+HCOO]\u2212 in negative MS, indicating the molecular formula as C42H72O13. Compound 45 was deduced as protopanaxatriol type and the latter three compounds were deduced as protopanaxadiol type on the base of the fragmentation rules described above. Among them, compound 61 has similar ions in full mass spectrum and MS/MS experiment as those of reference 20S-ginsenoside F2 and was unambiguously identified as 20S-ginsenoside F2. The deprotonated ion of 45 at m/z 783.4910 [M-H]\u2212 indicated the ion at m/z 475.3791 [M-H-rha-glc]\u2212, supporting that 45 was ginsenoside Rg2. Compounds 64 and 65 displayed same MS data; however, unfortunately, MS2 information could not be detected and they were tentatively identified as 20R-ginsenoside Rg3 and 20S-ginsenoside Rg3.As for reference 202 showed the ions at m/z 1245.8995 [2M+H]+, 645.4348 [M+Na]+, and 667.4437[M+HCOO]\u2212. Compound 74 was identified as 20S-ginsenoside Rh2 by comparing the retention time and MS data.Reference ginsenoside Rh 35, 36, and 37 have the same molecular formula of C48H82O19 and were tentatively characterized as 20S-glc-ginsenoside Rf, beads ginseng saponin F1, and notoginsenoside R3. They gave [M+Na]+ ions at m/z 985.5332, 985.5319, and 985.5339 in the positive ion mode and [M+HCOO]\u2212 ions at m/z 1007.5457, 1007.5457, and 1007.5459 in the negative ion mode. In their MS/MS experiments, the fragmentation ions of 35 at m/z 799.4843 and 637.4273 were produced from the ion at m/z 961.5391 [M-H]\u2212 by loss of glucose unit. Similarly, the fragmentation ions of 36 and 37 at m/z 799 and 637 were yielded. The retention order of three chromatographic peaks is determined according to the description in the literature [Compounds 42 and 43 are calculated as C41H70O13 based on the ions at m/z 793.4705 [M+Na]+ in the positive ion mode and 815.4818 [M+HCOO]\u2212 in the negative ion mode. Both compounds showed the fragmentation ions at m/z 405, 423, or 441 indicating that they are due to protopanaxatriol type. In MS/MS analysis, the ions at m/z 637.4326 [M-H-xyl]\u2212 and 475.3794 [M-H-xyl-glc]\u2212 for compound 42, as well as 637.4319 [M-H-ara]\u2212 and 475.3788 [M-H-ara-glc]\u2212 for compound 43, were detected from the deprotonated ions at m/z 769.4753 [M-H]\u2212 and 769.4758 [M-H]\u2212, respectively. Compounds 42 and 43 were identified as notoginsenoside R2 and ginsenoside F3.The chemical formulas of compounds 54 showed the quasimolecular ions at m/z 939.5284 [M+Na]+ and 961.5390 [M+HCOO]\u2212, indicating the molecular formula as C47H80O17. The fragmentation ions 783.4901 [M-H-xyl]\u2212 and 621.4363 [M-H-xyl-glc]\u2212 were detected, supporting that 54 was gypenoside IX.Compound 60 showed the quasimolecular ions at m/z 789.4742 [M+Na]+ and 811.4871 [M+HCOO]\u2212 and the molecular formula was calculated as C42H70O12. In the MS/MS analysis, the deprotonated ion at m/z 765.4792[M-H]\u2212 gave the ions at m/z 603.4261 [M-H-glc]\u2212 and 441.3730 [M-H-2glc]\u2212. Further loss of H2O yielded the ions at m/z 423 and 405. It was identified as ginsenoside Rk1.Compound 34 showed [M+Na]+ ion at m/z 1117.5740 and [M+HCOO]\u2212 ion at m/z 1139.5891, suggesting the molecule formula of C53H90O23. Besides, it showed [M-H]\u2212 ion at m/z 1093.5826 and fragment ions at m/z 961 [M-H- xyl/ara]\u2212, 799 [M-H-xyl/ara-glc]\u2212, and 637 [M-H-xyl/ara-2glc]\u2212, concerning consecutive loss of 132, 294, and 456. These data were in accordance with those of floral ginsenoside P, floranotoginsenoside D, notoginsenoside FT3, or yesanchinoside H [Compound 72 showed [M+Na]+ ion at m/z 789.4742 and [M+HCOO]\u2212 ion at m/z 811.4847, indicating the molecule formula as C42H70O12. Besides, the ions at m/z 603.4251 [M-H-162]\u2212 and 471.3466 [M-H-162-132]\u2212 in the MS/MS analysis suggested that the glycoside chain of 72 consisted of a molecule of glucose and a molecule of arabinose or xylose. It could be one of ilexoside A or ilexoside D described in the literature + and deprotonated ion at m/z 609.1461 [M-H]\u2212 as well as the ion at m/z 1219.3008 [2M-H]\u2212. Quercetin, with UV \u03bbmax absorption at 253 and 356 nm, displayed the deprotonated ion at m/z 301.0376 [M-H]\u2212. Four compounds  were determined as flavonoid derivatives due to the typical UV absorption. Especially, compound 9 was one of main constituents of XXT. Compounds 9 and 26 were unambiguously identified as rutin and quercetin based on the direct comparison of their UV spectra, and mass spectra with those of the authentic compounds. Compound 10 has UV \u03bbmax absorption at 264 and 343 nm. The molecular formula was calculated as C27H30O15 by the quasimolecular ion at m/z 595.1652 [M+H]+, 617.1472 [M+Na]+, and 593.1505 [M-H]\u2212. Other fragment ions at m/z 449.1083 [M+H-rha]+ and 287.1055 [M+H-rha-glc]+ were detected. Therefore, compound 10 was identified as kaempferol-3-O-rutinoside. Compound 13 has UV \u03bbmax absorption at 252 and 339 nm. The molecular formula was calculated as C28H32O16 by the quasimolecular ion at m/z 625.1754 [M+H]+, 647.1578 [M+Na]+, and 623.1616 [M-H]\u2212, suggesting it was isorhamnetin 3-O-rutinoside \u2212 ion at m/z 395.07949 was found. The MS/MS spectrum of [M-H]\u2212 exhibited an obvious fragment ion, [M-H-H2O]\u2212 at m/z 179.0338, and further loss of COOH obtained ion at m/z 135.0438. Compound 1 was identified as danshensu.Danshensu has UV \u03bbmax absorption at 323 nm. In the full mass spectrum, the deprotonated ion at m/z 353.0869 [M-H]\u2212 was found. The MS/MS spectrum of [M-H]\u2212 showed ions at m/z 191.0548, 179.0336, and 135.0438. Compound 2 was identified as neochlorogenic acid.Neochlorogenic acid has UV \u03bbmax absorption at 325 nm and showed the quasimolecular ion at m/z 353.0872 [M-H]\u2212 and 707.1482 [2M-H]\u2212. The fragment ion at m/z 191.0553 was yielded by losing C9H6O3. Compound 4 was identified as chlorogenic acid.Chlorogenic acid has UV \u03bbmax absorption at 229, 278, and 310 nm. The [M-H]\u2212 ion at m/z 137.0236 was detected. In the MS/MS analysis, the ion at m/z 108.0204 was yielded by losing CHO and further losing oxygen produced from the ion at m/z 92.0256. Compound 5 was identified as protocatechuic aldehyde.Protocatechuic aldehyde has UV \u03bbmax absorption at 238 and 322 nm. It showed [M-H]\u2212 ion at m/z 179.0339 in the full mass spectrum and the ion at m/z 135.0438 by losing CO2 in the MS/MS experiment. Compound 7 was identified as caffeic acid.Caffeic acid has UV 11, 14, and 15) at m/z 515 [M-H]\u2212 and 517 [M+H]+ were easily located in the chromatogram of XXT, suggesting the molecular formula of C25H24O12. They were assigned as dicaffeoylquinic acids by comparison with retention times. Besides, they gave the same fragment ions with those of chlorogenic acid, such as m/z 191, 179, 173, and 135. in MS/MS experiments. The fragmentation pathways are in accordance with those described in the literature terature . Based o 12 has UV \u03bbmax absorption at 244 and 327 nm. The molecular formula was calculated as C27H22O12 on the basis of the quasimolecular ions at m/z 539.1149 [M+H]+ and 537.1031 [M-H]\u2212. In the MS/MS experiment, it indicated the characteristic ion at m/z 339.0500 [M-H-C9H10O5]\u2212, supporting that compound 12 was salvianolic acid H.Compound 17 has UV \u03bbmax absorption at 250 and 306 nm. It showed the ion at m/z 341.0666 [M+H]+. In the MS/MS analysis, the ions at m/z 295.0617 and 279.0659 were yielded by losing CH2O2 and CH2O3. Compound 17 was identified as salvianolic acid G.Compound 18 has UV \u03bbmax absorption at 251 and 317 nm. It showed [M-H]\u2212 ion at m/z 493.1134 and the molecular formula was calculated as C26H22O10. In the MS/MS analysis, the fragment ion at m/z 295.0604 [M-H-C9H10O5]\u2212 was detected, suggesting compound 18 was salvianolic acid A.Compound 32 showed the ions at m/z 567.1498 [M+H]+, 589.1320 [M+Na]+, and 565.1342 [M-H]\u2212 in MS spectrum, revealing molecular formula of C29H26O12. Besides, fragment ion at m/z 369.0971 [M-H-C9H10O5]\u2212 was observed. Compound 32 was identified as ethyl lithospermate.CompoundBufadienolides. The MS/MS behaviors of bufadienolides have been extensively described + ions was triggered by initial loss of 60 Da (HOAc). The elimination of CO was significant for bufadienolides with a 19-formyl group, and the 19-hydroxyl group could be characterized by the loss of 30 Da (HCHO). These fragmentation rules were applied to the identification of bufadienolides in XXT sample. As shown in 19), arenobufagin (25), bufotalin (41), bufalin (50), resibufogenin (55), and cinobufagin (56) by comparison with reference substances isolated from toad venom. The other two bufadienolides were tentatively identified as bufarenogin (20) and hellebrigenin (29) + ion at m/z 403.2486 was found and the fragment ions at m/z 385.2366 [M+H-H2O]+, 367.2262 [M+H-2H2O]+, and 349.2169 [M+H-3H2O]+ were detected in MS2.escribed , 18. BriBile acids. Bile acid derivatives are in lack of conjugated system and their UV absorption is not obvious. However, they always present high sensitivity in the negative ion mode. The deprotonated ion [M-H]\u2212 and adduct ion [M+HCOO]\u2212 were obviously detected in MS spectrum. The loss of side chain was commonly observed in MS2. Four bile acid derivatives were detected from XXT sample, two of which were unambiguously identified as ursodeoxycholic acid  and chenodeoxycholic acid  based on the direct comparison of reference substances. The other two compounds were tentatively identified as cholanic acid (57) and hyodeoxycholic acid (67). They are derived from the raw material, Rengong Niuhuang  [ifactus) . m/z 391.2851 [M-H]\u2212, 437.2916 [M+HCOO]\u2212, and 783.5778 [2M-H]\u2212 in the negative ion mode. In the positive ion mode, the ion at m/z 357.2807 [M+H-2H2O]+ was assigned as the loss of two molecules of H2O. The ion at m/z 321.2587 represented its side chain loss in MS2. Compound 58 was identified as UDCA.Ursodeoxycholic acid showed the ions at m/z 391.2872 [M-H]\u2212, 437.2907[M+HCOO]\u2212, and 783.5798 [2M-H]\u2212 in the negative ion mode. In the positive ion mode, the loss of H2O unit yielded [M+H-2H2O]+ ion at m/z 357.2809. The occurrence of the ion at m/z 321.2586 in MS2 was due to side chain loss. Compound 70 was identified as CDCA.CDCA showed the ions at 57 showed [M+Na]+ ion at m/z 431.2778 in positive mode, and [M-H]\u2212 ion at m/z 407.2803, [M+HCOO]\u2212 ion at m/z 453.2857, and [2M-H]\u2212 ion at m/z 815.5698 in the negative mode. The ions representing a series of H2O loss was observed, such as 373.2749 [M+H-2H2O]+ and 355.2649 [M+H-3H2O]+. It was tentatively characterized as cholanic acid.Compound 67 showed [M-H]\u2212 ion at m/z 391.2852, [M+HCOO]\u2212 ion at m/z 437.2903, and [2M-H]\u2212 at m/z 783.5792 in the full mass spectrum. The molecular formula was calculated as C24H40O4. It was tentatively deduced as hyodeoxycholic acid.CompoundQuinones derivatives. Quinones derivatives are another kind of active constituents from Danshen and they were easily detected in XXT sample. Compounds 63, 71, and 76 were identified as dihydrotanshinone, cryptotanshinone, and tanshinone II A by comparison with reference substances. They displayed similar fragmentation pathways concerning successive eliminations of H2O and CO molecules. Tanshinone IIA was used as an example to illustrate the fragmentation pathway of quinones constituents as shown in  31, 1-oxo tanshinone IIA 59, neotanshinone D 62, tetrahydrotanshinone I 66, methyltanshinonate 69, methylenetanshinquinone 73, and miltirone 77. + ion at m/z 207.1024. The above data were in accordance with those of 4-hydroxyl-3-butylphthalide in the literature [ 21 was tentatively assigned as 4-hydroxyl-3-butylphthalide. It could be derived from individual herb Chuanxiong rhizome [terature . Compoun rhizome .Among the identified compounds, most constituents were derived from the raw materials, Danshen and Renshen Jingye Zongzaogan, and a small proportion of compounds were considered from Huaihua, Maodongqing, Chuanxiong, Chansu, and Rengong Niuhuang. The other ingredients, Rengong Shexiang, Bingpian, and Shuizhi, were not characterized in the present HPLC and UPLC-QTOF/MS condition. This could be related to the prescription of raw materials and the manufacturing process employed. Usually, muscone, one of active constituents in Rengong Shexiang and borneol are detected by GC or GC-MS . They arIn this work, HPLC analysis was employed to find out the common chromatographic peak in various batches of XXT samples and UPLC-Q-TOF/MS was used for the identification of main constituents in the typical XXT sample. As a result, a total of 63 constituents including twenty saponins, four flavonoids, fifteen phenolic acids, eight steroids, four bile acids, ten quinones, and other two compounds were identified or tentatively characterized based on the comparison of retention time and UV spectra with authentic compounds as well as by summarized MS fragmentation rules and matching empirical molecular formula with those of published components. The present investigation clearly understood the nonvolatile constituents in XXT and provided good basis for further study on the active substances and quality control of this preparation."}
+{"text": "Scientific Reports 10.1038/s41598-019-42959-4, published online 29 April 2019Correction to: In Figure 5 there is a missing axis label. The y-axis of the bottom graph should be labelled \u201cIntensity (arb. units)\u201d The correct Figure 5 appears below as Figure\u00a0Additionally, this Article contains errors in Reference 44 which is incorrectly given as:Jpn. J. Appl. Phys. 56, 01AC06 (2016)Ito, T., Uchida, G., Nakajima, A., Takenaka, K. & Setsuhara, Y. Selective production of reactive oxygen and nitrogen species in the plasma-treated water by using a nonthermal high-frequency plasma jet. The correct reference is listed below as ref."}
+{"text": "Decline in driving skills begins in preclinical AD, when an older adult remains cognitively normal, but the underlying disease process has begun. Preclinical AD is detectable among cognitively normal individuals using molecular biomarkers: positron emission tomography (PET) imaging and cerebrospinal fluid (CSF). The aim of this prospective, longitudinal study is to determine whether naturalistic driving behavior using in-vehicle dataloggers can distinguish older adults with (n=36) and without preclinical AD (n=134). Driving data was calculated as mean/month for several variables  for participants followed between one to 46 months. Using stepwise logistic regression, the area under the receiver operating curve (AUC) and 95% confidence interval for these five variables was 0.73 (0.63-0.79) in distinguishing those with and without preclinical AD via amyloid imaging. When age, gender, race, and education were added, the model improved: 0.80 (0.72-0.88). Finally, when apolipoprotein \u03b54 allele (APO\u03b54), obtained via blood or saliva, was added to the model, accuracy improved: 0.84 (0.77-0.89). Similar results were found using CSF biomarker tau/A\u03b242: AUCs (95% CI) were 0.68 (0.58-0.79) for driving variables alone, 0.77 (0.69-0.86) for driving variables and demographics, and 0.87 (0.80-0.94) driving variables, demographics, and apolipoprotein \u03b54 allele. These promising findings suggest that naturalistic driving behavior can predict those with and without preclinical AD. The AUC is further improved with demographics and APO\u03b54, an easily obtainable genetic biomarker. This model may be used in clinical/research settings as a screen or adjunct for diagnostics and prognostics purposes."}
+{"text": "Rhodococcus erythropolis X5 is a psychrotrophic (cold-adapted) hydrocarbon-degrading bacterium, as it showed effective n-alkane destruction at low positive temperatures. Here, the genome of strain X5 was completely sequenced; it consists of a 6,472,161-bp circular chromosome (62.25% GC content) and a 526,979-bp linear plasmid, pRhX5-526k (62.37% GC content). Rhodococcus erythropolis X5 is a psychrotrophic (cold-adapted) hydrocarbon-degrading bacterium, as it showed effective n-alkane destruction at low positive temperatures. Here, the genome of strain X5 was completely sequenced; it consists of a 6,472,161-bp circular chromosome (62.25% GC content) and a 526,979-bp linear plasmid, pRhX5-526k (62.37% GC content). Rhodococcus erythropolis X5 (VKM Ac-2532D) was isolated from oil-polluted soil (R. erythropolis X5 is a psychrotrophic (cold-adapted) hydrocarbon-degrading bacterium, as it showed effective n-alkane destruction at low positive temperatures . Sequencing was performed using a MinIon sequencer (Oxford Nanopore Technologies [ONT]) at the Center of Analytical and Genetic Engineering Research . A library was prepared with the ONT ligation sequencing kit . Guppy v3.2.4 software was used for base calling, which yielded a total of 301 Mbp, distributed in 33,708 reads. Reads with a Q score of >10 were used for further analysis. Additionally, the same DNA sample was sequenced with an Illumina MiSeq platform using a MiSeq reagent kit v3 (2\u2009\u00d7\u2009300\u2009bp). A paired-end library for sequencing was prepared with the MuSeek library preparation kit . The Nanopore reads were assembled into 2 contigs using Canu assembler v1.8 (Genomic DNA was isolated from a fresh culture biomass (a colony) of R. erythropolis. For the average nucleotide identity (ANI) analysis, we used all available GenBank data on the R. erythropolis representatives and on Rhodococcus sp. strains. Average nucleotide identity parameter analysis revealed that the closest phylogenetic relatives of strain X5 are Rhodococcus sp. strain 008 (GenBank accession number CP012749), Rhodococcus sp. strain NJ-530 (CP034152), R. erythropolis CCM2595 (CP003761), and Rhodococcus sp. strain YL-1 (CP017299).The X5 genome consists of a 6,472,161-bp circular chromosome (62.25% GC content) and a 526,979-bp linear plasmid, pRhX5-526k (62.37% GC content). Chromosome circularization and plasmid linearity were specified by Canu. Also, there were no reads overlapping with plasmid ends. The strain was previously identified as R. erythropolis X5 bears 5 copies of alkane hydroxylase-encoding gene alkB. These genes are essential for the degradation process of alkanes with a chain length of C12 to C20  v4.6 , which i2 to C20 , 10. In 2 to C20 , 12. ApaCP044283 and CP044284, BioSample number SAMN12818508, BioProject number PRJNA573614, and SRA accession number PRJNA573614.This genome project has been deposited in the NCBI database under GenBank accession numbers"}
+{"text": "Elaeagnus macrophylla were analyzed via conserved DNA-derived polymorphism molecular markers. A total of 289 discernible loci were obtained from 102 individuals via fifteen primers, and 100% of the loci were polymorphic. The observed number of alleles was 1.9654, and the effective number of alleles was 1.2604. Nei\u2019s genetic diversity index was 0.1724 on average, and Shannon\u2019s information index was 0.2869, indicating that Elaeagnus macrophylla had lower levels of genetic diversity than those reported for its continental relatives and other continental species. The average percentage of polymorphic loci was 42.1%, and the maximum and minimum were 80.97% and 14.88%, respectively, which were associated with the Nanji Island and Liugong Island populations, respectively. The populations of Elaeagnus macrophylla were highly differentiated. Cluster analysis revealed that the similarity between the tested samples was related to their geographical location, that the samples from the same island tended to cluster together, and that there was no cross-clustering between samples. The Nanji Island and Da Rushan populations differentiated into two subpopulations. Last, we detected no correlation between genetic distance and geographic distance between populations .The genetic diversity and genetic structure of five natural populations of the island and coastal endangered plant species  Elaeagnus\u00a0macrophylla is an endangered evergreen shrub species of East Asian coastal areas and islands. It is distributed in the Shandong, Zhejiang, and Jiangsu Provinces of China, mainly on offshore islands and in coastal lowlands  molecular markers (Neolitsea sericea based on random amplified polymorphic DNA (RAPD) molecular markers  have shown that CDDP markers have many advantages, including convenience, low cost, and rich polymorphism, and can effectively mark sequences of target traits (Solanum dulcamara (Chrysanthemum (Paeonia suffruticosa (Vaccinium\u00a0vitis-idaea (Rosa rugosa (E.\u00a0macrophylla.The conserved DNA-derived polymorphism (CDDP) method is based on a single primer amplification reaction, with primers designed to target conserved sequences of plant functional genes, mostly transcription factors such as WRKYs, MYBs, MADs, ERFs, KNOXs, and ABP1. Because of the strong conservation of some sequences of plant DNA, CDDP molecular marker technology can be used across different species. Studies of rice . Only one individual each was found on Lingshan Island and Putuo Island. After the samples were collected, silica gel was used to quickly dry the specimens, after which they were stored at 20\u00a0\u00b0C.A total of 102 individual leaf samples were collected from 7 islands and offshore sites  from AprE. macrophylla via the modified cetyltrimethylammonium bromide (CTAB) method  method . The qua2O, 1 \u00b5l of 30 ng/\u00b5l DNA template, and 1.0 \u00b5l of 10 pmol/\u00b5l primers . A standard PCR thermocycler  was used and the PCR program was as follows: an initial denaturation step at 94\u00a0\u00b0C for 3 min; 35 annealing cycles of 94 \u00a0\u00b0C for 1 min, 50\u00a0\u00b0C for 1 min and 72\u00a0\u00b0C for 2 min; and a final extension of 72 \u00a0\u00b0C for 5 min. The PCR products were subsequently stored at 4\u00a0\u00b0C. The products were then electrophoresed on a 2% agarose gel (110 V and 110 mA) for 1.5\u20132 h; a DL2000 marker was used as a size marker. The electrophoresis results were imaged and recorded by a gel imaging system. All amplification procedures were repeated at least twice to ensure the repeatability of the experiment.The DNA from one sample per population was selected to screen 21 CDDP primers  synthes. PCR wasWe used POPGEN v.1.32  to compuA dendrogram was generated by the unweighted pair group method with arithmetic mean (UPGMA) clustering procedure in NTSYS-pc 2.10e software . The relK values) was set from 1 to 10, with 10 independent runs for each K. The contribution of the accessions to the genotypes was calculated on the basis of a 105 iteration burn-in period and 105 iteration sampling period. The optimal number of K clusters was then identified according to the methods of The genetic structure of populations was further assessed via the Bayesian clustering approach implemented in STRUCTURE v.2.3.4 . The numE. macrophylla was high.The DNA of 102 samples was amplified with 15 primers, yielding 289 bands, and the fragment length was between 500 and 2,000 bp . The numAt the population level, the PPL ranged from 14.88% to 80.28%, with an average of 48.928%, whereas it was 96.54% at the species level. The Na ranged from 1.1488 to 1.8028, while the Ne varied from 1.0739 to 1.2410. H varied from 0.0446 to 0.1580, with an average of 0.1149, and I ranged from 0.0690 to 0.2613, with an average of 0.1848. At the species level, H and I were 0.1724 and 0.2869, respectively . The Na,The Ht and Hs were 0.1706 and 0.1149, respectively, as calculated by POPGEN v.1.32 software. The Gst was 0.3263, indicating that 67.37% of the variation was within the populations and that 32.63% of the variation occurred between the populations. A certain degree of genetic differentiation was observed between the populations. The Nm was 1.0325, indicating that there was some  genetic exchange between populations.E. macrophylla, the genetic distance was between 0.0490 and 0.1443 .The applied measure of genetic similarity was used to construct UPGMA dendrograms . The cluThe UPGMA clustering map provided a clear division of the 102 samples . NotablyE. macrophylla best fit three genetic groups, and when K\u00a0=\u00a05, the delta K value was also large (K\u00a0=\u00a03 (K\u00a0=\u00a05 (K\u00a0=\u00a05: a northwestern group (indicated in yellow) and a southeastern group (indicated in blue). The DRS population differentiated into two subpopulations when K\u00a0=\u00a03 and K\u00a0=\u00a05: Nos. 9\u201318 and Nos. 19\u201328, respectively.The results of the Bayesian clustering analysis of the genetic structure showed that the populations of so large . When K\u00a0e   (Elaeagnus mollis)   and Paeo\u00a00.2682)  estimate mollis) , the geniversity . E. macrlination . The Ht is sweet . The frutinction . Island tinction .E. macrophylla were 0.1706 and 0.1149, respectively. Compared with endangered and Chinese secondary protected plants  studies   . This ge >\u00a00.15) . Furtherulations .E. macrophylla displayed little gene flow (Nm = 1.0325), and the UPGMA clustering analysis of samples revealed no hybridization among individuals from different localities. Structure analysis (K\u00a0=\u00a05) revealed that most of the populations had a simple pedigree, and the genetic exchange between each pair of populations was low. These results were mainly due to the geographical isolation of the islands (mainly barriers posed by seawater), which limited the range of dispersal by pollen- and seed-dispersing birds ; it is difficult for small pollinators to spread pollen across islands separated by vast seas. In addition to pollen, seeds play an important role in the spread of gene flow. One important aspect of seed movement is its role in the initial founding of a population  of N. sericea were dispersed within a range of approximately 480 m (Hakdongri on Keojae Island) and 680 m (Naechori on Oenaro Island), respectively, from their maternal plants . After possible causes such as the prevention of gene flow were excluded, the reason may be related to differences in habitats within the population. The NJD nature reserve has numerous islands and reefs with meandering shorelines, headlands, and numerous bays. There are many types of coastal beaches, such as mudflats, gravel beaches, and rocky reefs, and NJD is at the intersection of the Taiwan Warm Current and the Jiangsu and Zhejiang Coastal Currents. The flow system is complex, so the habitat is complex  was significantly greater than that of island species with relatively small populations   NJD (0.1580)>DRS (0.1290)>DGD (0.1222)>LS (0.1208)>LGD (0.0446), with an average value of 0.1149. The average genetic diversity index of the NJD population is much greater than that of the LGD population. The values of the other three populations are relatively similar. The NJD population is the largest, the DRS, DGD, and LS populations are similar in size, and the LGD population is declining. Human disturbances such as excessive logging, habitat destruction, and the introduction of exotic species are considered to be the main causes endangering island species  . The ave species , which a species . NJD is E. macrophylla.Am in situ conservation method was proposed because the conservation of sufficient natural population numbers and sizes to prevent a reduction in genetic diversity is urgently needed. The best strategy for in situ conservation of genetic diversity during an endemic is the preservation of natural habitat . In thisElaeagnus macrophylla using conserved DNA-derived polymorphism markers to investigate the distribution and genetic variation. The results showed that conserved DNA-derived polymorphism markers can be effectively used to study the genetic diversity of Elaeagnus macrophylla populations and revealed that Elaeagnus macrophylla populations have low genetic diversity and high genetic differentiation. The low levels of gene flow between populations are the main cause of the high levels of genetic differentiation. On the basis of these findings, some conservation measures for Elaeagnus macrophylla are proposed.The present study is the first genetic investigation of 10.7717/peerj.8498/supp-1Supplemental Information 10-1 matrix obtained from the results of electropherogram.Click here for additional data file.10.7717/peerj.8498/supp-2Supplemental Information 2It can be seen that the three primer amplification bands are clear and specific and can be used for experiments.Click here for additional data file.10.7717/peerj.8498/supp-3Supplemental Information 3It can be seen that the MYB1, MYB2, ERF1 amplified bands are clear and specific and can be used for experiments.Click here for additional data file.10.7717/peerj.8498/supp-4Supplemental Information 4It can be seen that the WRKY-F1, WRKY-R1 amplified bands are clear and specific and can be used for experiments. Although the WRKY-R2 has a strip, the strip is weak and therefore not used.Click here for additional data file.10.7717/peerj.8498/supp-5Supplemental Information 5It can be seen that the WRKY-R3 amplified band is clear and specific, and can be used for experiments. WRKY-R2B, WRKY-R3B is not effective, so it is not used.Click here for additional data file.10.7717/peerj.8498/supp-6Supplemental Information 6It can be seen that KNOX-2, KNOX-3, MADS-1 amplified bands are clear and specific and can be used for experiments.Click here for additional data file.10.7717/peerj.8498/supp-7Supplemental Information 7It can be seen that the ABP1-1, ABP1-3 amplified bands are clear and specific and can be used for experiments. ABP1-2 did not amplify the band and therefore could not be used.Click here for additional data file.10.7717/peerj.8498/supp-8Supplemental Information 8It can be seen that the MADS-4 amplification band is clear and specific and can be used for experiments. Although the MADS-2 has a band, it is not effective in use, and the MADS-3 does not amplify the band, so it is not used.Click here for additional data file.10.7717/peerj.8498/supp-9Supplemental Information 9Amplification results of WRKY-F1 on LGD1-8 and DRS9-20.Click here for additional data file.10.7717/peerj.8498/supp-10Supplemental Information 10Amplification results of WRKY-F1 on DRS21-28, NJD1-12.Click here for additional data file.10.7717/peerj.8498/supp-11Supplemental Information 11Amplification results of WRKY-F1 on NJD13-NJD32.Click here for additional data file.10.7717/peerj.8498/supp-12Supplemental Information 12Amplification results of WRKY-F1 on NJD 33, LS 1-13, PTD1, LSD1.Click here for additional data file.10.7717/peerj.8498/supp-13Supplemental Information 13Amplification results of WRKY-F1 on DGD1-13.Click here for additional data file.10.7717/peerj.8498/supp-14Supplemental Information 14Amplification results of WRKY-F1 on DGD14-26.Click here for additional data file.10.7717/peerj.8498/supp-15Supplemental Information 15Amplification results of WRKY-R1 on LGD1-8 and DRS9-20.Click here for additional data file.10.7717/peerj.8498/supp-16Supplemental Information 16Amplification results of WRKY-R1 on DRS21-28, NJD1-12.Click here for additional data file.10.7717/peerj.8498/supp-17Supplemental Information 17Amplification results of WRKY-R1 on NJD13-32.Click here for additional data file.10.7717/peerj.8498/supp-18Supplemental Information 18Amplification results of WRKY-R1 on NJD 33, LS1-13, PTD1, LSD1.Click here for additional data file.10.7717/peerj.8498/supp-19Supplemental Information 19Amplification results of WRKY-R1 on DGD1-20.Click here for additional data file.10.7717/peerj.8498/supp-20Supplemental Information 20Amplification results of WRKY-R1 on DGD21-26.Click here for additional data file.10.7717/peerj.8498/supp-21Supplemental Information 21Amplification results of WRKY-R3 on LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-22Supplemental Information 22Amplification results of WRKY-R3 on DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-23Supplemental Information 23Amplification results of WRKY-R3 on DRS21-28, NJD13-32 samples.Click here for additional data file.10.7717/peerj.8498/supp-24Supplemental Information 24Amplification results of WRKY-R3 on NJD33, LS1-13, PTD1, LSD1 samples.Click here for additional data file.10.7717/peerj.8498/supp-25Supplemental Information 25Amplification results of WRKY-R3 on DGD1-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-26Supplemental Information 26Amplification results of WRKY-R3 on DGD21-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-27Supplemental Information 27Amplification results of MYB1 on LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-28Supplemental Information 28Amplification results of MYB1 on DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-29Supplemental Information 29Amplification results of MYB1 on NJD13-33 samples.Click here for additional data file.10.7717/peerj.8498/supp-30Supplemental Information 30Amplification results of MYB1 on NJD33, LS1-13, PTD1, LSD1 samples.Click here for additional data file.10.7717/peerj.8498/supp-31Supplemental Information 31Amplification results of MYB1 on DGD1-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-32Supplemental Information 32Amplification results of MYB1 on DGD21-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-33Supplemental Information 33Amplification results of MYB2 on LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-34Supplemental Information 34Amplification results of MYB2 on DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-35Supplemental Information 35Amplification results of MYB2 on NJD13-32 samples.Click here for additional data file.10.7717/peerj.8498/supp-36Supplemental Information 36Amplification results of MYB2 on NJD33, LS1-13, PUD1, LSD1 samples.Click here for additional data file.10.7717/peerj.8498/supp-37Supplemental Information 37Amplification results of MYB2 on DGD1-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-38Supplemental Information 38Amplification results of MYB2 on DGD21-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-39Supplemental Information 39Amplification results of ERF1 on LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-40Supplemental Information 40Amplification results of ERF1 on DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-41Supplemental Information 41Amplification results of ERF1 on NJD13-32 samples.Click here for additional data file.10.7717/peerj.8498/supp-42Supplemental Information 42Amplification results of ERF1 on NJD33, LS1-13, PTD1, LSD1 samples.Click here for additional data file.10.7717/peerj.8498/supp-43Supplemental Information 43Amplification results of ERF1 on DGD1-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-44Supplemental Information 44Amplification results of ERF1 on LS1-6, DGD21-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-45Supplemental Information 45Amplification results of ERF2 on LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-46Supplemental Information 46Amplification results of ERF2 on DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-47Supplemental Information 47Amplification results of ERF2 on NJD13-32 samples.Click here for additional data file.10.7717/peerj.8498/supp-48Supplemental Information 48Amplification results of ERF2 on NJD33, LS1-13, PTD1, and LSD1 samples.Click here for additional data file.10.7717/peerj.8498/supp-49Supplemental Information 49Amplification results of ERF2 on DGD1-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-50Supplemental Information 50Amplification results of ERF2 on DGD21-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-51Supplemental Information 51Amplification results of ERF3 on LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-52Supplemental Information 52Amplification results of ERF3 on DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-53Supplemental Information 53Amplification results of ERF3 on NJD13-32 samples.Click here for additional data file.10.7717/peerj.8498/supp-54Supplemental Information 54Amplification results of ERF3 on NJD33, LS1-13, PTU1, LSD1, and DGD24-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-55Supplemental Information 55Amplification results of ERF3 on DGD1-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-56Supplemental Information 56Amplification results of ERF3 on DGD24-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-57Supplemental Information 57KNOX-1 amplification results for LGD1-8, DRS9-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-58Supplemental Information 58KNOX-1 amplification results for DRS21-28, NJD1-12 samples.Click here for additional data file.10.7717/peerj.8498/supp-59Supplemental Information 59KNOX-1 amplification results for NJD13-32 samples.Click here for additional data file.10.7717/peerj.8498/supp-60Supplemental Information 60KNOX-1 amplification results for NJD33, LS1-13, PUD1, LSD1 samples.Click here for additional data file.10.7717/peerj.8498/supp-61Supplemental Information 61KNOX-1 amplification results for DGD1-20 samples.Click here for additional data file.10.7717/peerj.8498/supp-62Supplemental Information 62KNOX-1 amplification results for DGD21-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-63Supplemental Information 63KNOX-2 amplification results for LGD1-8, DRS9-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-64Supplemental Information 64KNOX-2 amplification results for DRS24-28, NJD1-18 samples.Click here for additional data file.10.7717/peerj.8498/supp-65Supplemental Information 65KNOX-2 amplification results for NJD19-33, LS1-6 samples.Click here for additional data file.10.7717/peerj.8498/supp-66Supplemental Information 66KNOX-2 amplification results for LS7-13, PTD1, LSD1and DGD1-13 samples.Click here for additional data file.10.7717/peerj.8498/supp-67Supplemental Information 67KNOX-2 amplification results for DGD14-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-68Supplemental Information 68KNOX-3 amplification results for LGD1-8, DRS9-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-69Supplemental Information 69KNOX-3 amplification results for DRS24-28, NJD1-18 samples.Click here for additional data file.10.7717/peerj.8498/supp-70Supplemental Information 70KNOX-3 amplification results for NJD19-33, PTD1, LSD1, LS1-4 samples.Click here for additional data file.10.7717/peerj.8498/supp-71Supplemental Information 71KNOX-3 amplification results for LS5-13, DGD1-13 samples.Click here for additional data file.10.7717/peerj.8498/supp-72Supplemental Information 72KNOX-3 amplification results for DGD14-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-73Supplemental Information 73Amplification results of MADS-1 on LGD1-8, DRS9-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-74Supplemental Information 74Amplification results of MADS-1 on DRS24-28, NJD1-18 samples.Click here for additional data file.10.7717/peerj.8498/supp-75Supplemental Information 75Amplification results of MADS-1 on NJD19-NJD33, PTD1, LSD1, LS1-4 samples.Click here for additional data file.10.7717/peerj.8498/supp-76Supplemental Information 76Amplification results of MADS-1 on LS5-13, DGD1-13 samples.Click here for additional data file.10.7717/peerj.8498/supp-77Supplemental Information 77Amplification results of MADS-1 on DGD14-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-78Supplemental Information 78Amplification results of MADS-4 on LGD1-8 and DRS9-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-79Supplemental Information 79Amplification results of MADS-4 on DRS24-28, NJD1-18 samples.Click here for additional data file.10.7717/peerj.8498/supp-80Supplemental Information 80Amplification results of MADS-4 on NJD19-33, PUD1, LSD1, LS1-4 samples.Click here for additional data file.10.7717/peerj.8498/supp-81Supplemental Information 81Amplification results of MADS-4 on LS5-13, DGD1-13 samples.Click here for additional data file.10.7717/peerj.8498/supp-82Supplemental Information 82Amplification results of MADS-4 on DGD14-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-83Supplemental Information 83ABP1-1 amplification results for LGD1-8, DRS9-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-84Supplemental Information 84ABP1-1 amplification results for DRS24-28, NJD1-18 samples.Click here for additional data file.10.7717/peerj.8498/supp-85Supplemental Information 85ABP1-1 amplification results for NJD19-33, LS1-4 samples.Click here for additional data file.10.7717/peerj.8498/supp-86Supplemental Information 86ABP1-1 amplification results for LS5-13, DGD1-13 samples.Click here for additional data file.10.7717/peerj.8498/supp-87Supplemental Information 87ABP1-1 amplification results for DGD14-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-88Supplemental Information 88ABP1-3 amplification results for LGD1-8, DRS9-23 samples.Click here for additional data file.10.7717/peerj.8498/supp-89Supplemental Information 89ABP1-3 amplification results of DRS24-28, NJD1-18 samples.Click here for additional data file.10.7717/peerj.8498/supp-90Supplemental Information 90ABP1-3 amplification results for NJD19-33, PTD1, LSD1, LS1-4 samples.Click here for additional data file.10.7717/peerj.8498/supp-91Supplemental Information 91ABP1-3 amplification results for LS5-13, DGD1-13 samples.Click here for additional data file.10.7717/peerj.8498/supp-92Supplemental Information 92ABP1-3 amplification results for DGD14-26 samples.Click here for additional data file.10.7717/peerj.8498/supp-93Supplemental Information 93When K=3, the value of delat K is the largest and best fit three genetic groups. When K=5, the value of delat K is also large, so we analyze the cases of K=3 and K=5.Click here for additional data file.10.7717/peerj.8498/supp-94Supplemental Information 94Nm represents the gene flow, and Gst represents the genetic differentiation coefficient.Click here for additional data file.10.7717/peerj.8498/supp-95Supplemental Information 95Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-96Supplemental Information 96Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-97Supplemental Information 97Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-98Supplemental Information 98Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-99Supplemental Information 99Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-100Supplemental Information 100Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-101Supplemental Information 101Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-102Supplemental Information 102Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-103Supplemental Information 103Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-104Supplemental Information 104Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-105Supplemental Information 105Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-106Supplemental Information 106Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-107Supplemental Information 107Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-108Supplemental Information 108Two repetitions per sample.Click here for additional data file.10.7717/peerj.8498/supp-109Supplemental Information 109Raw data format when generating a sample cluster map.Click here for additional data file.10.7717/peerj.8498/supp-110Supplemental Information 110Generate raw data for population clustering analysis.Click here for additional data file."}
+{"text": "Xanthomonas sp. strain ATCC PTA-13101, which was isolated from rice. The 44-kbp Pagan genome contains direct terminal repeats and contains 59 genes, 27 of which have a predicted function. Pagan is most closely related to Xanthomonas phage phi Xc10 and Xylella phage Prado.The T7-like podophage Pagan infects  Xanthomonas sp. strain ATCC PTA-13101, which was isolated from rice. The 44-kbp Pagan genome contains direct terminal repeats and contains 59 genes, 27 of which have a predicted function. Pagan is most closely related to Xanthomonas phage phi Xc10 and Xylella phage Prado.The T7-like podophage Pagan infects  Xanthomonas consists of many diverse species of Gram-negative, rod-shaped phytopathogenic gammaproteobacteria , and phages were propagated via the soft agar overlay method described previously (www.bioinformatics.babraham.ac.uk/projects/fastqc). Sequence reads were then trimmed using the FASTX-Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/). The genome was assembled through SPAdes v3.5.0 (using default parameters) with a sequencing coverage of 6.3-fold  , mixed freshwater collected in Vidor, TX, and infects the rice-associated eviously , 7. Full6.3-fold . Next, txy-pub/) . Structuxy-pub/) \u201314. Geney-pub/) \u2013\u201317. Like-pub/) \u2013\u2013, 19. Rho-pub/) \u2013\u2013. progres-pub/) \u2013\u2013. Phage m-pub/) \u2013\u2013.Xanthomonas genome average of 64% G+C content (Xanthomonas phage phi Xc10 (GenBank accession number MF375456) and Xylella phage Prado (accession number KF626667), with 93.46% and 69.55% nucleotide sequence identity and 53 and 48 shared proteins, respectively (The 44,448-bp podophage Pagan genome has a coding density of 96.95% and contains 59 predicted genes but no tRNAs. The genome of Pagan displayed a G+C content of 62.3%, near the host  content . Pagan iectively . These sMK903278, BioProject accession number PRJNA222858, SRA accession number SRR8892144, and BioSample accession number SAMN11408680.The genome sequence and associated data for phage Pagan were deposited under GenBank accession number"}
+{"text": "Klebsiella pneumoniae, a Gram-negative pathogen whose multidrug-resistant strains are a public health issue. Here, we describe the annotation of the 157,741-bp Magnus genome and its similarity to other myophages.Bacteriophage Magnus infects  Klebsiella pneumoniae, a Gram-negative pathogen whose multidrug-resistant strains are a public health issue. Here, we describe the annotation of the 157,741-bp Magnus genome and its similarity to other myophages.Bacteriophage Magnus infects  Klebsiella pneumoniae is an opportunistic, Gram-negative enteric bacterium. Carbapenemase-producing K. pneumoniae (KPC) strains are resistant to carbapenem antibiotics and are an emerging cause of nosocomial and systemic human infections (fections . Phage tK. pneumoniae strain 39427 (BioSample number SAMN06218024) (www.bioinformatics.babraham.ac.uk/projects/fastqc). Quality-controlled reads were trimmed using the FASTX-Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/) and then assembled into a single contig of circular assembly at 146-fold coverage using SPAdes v3.5.0   collected at a wastewater treatment facility in Austin, TX, against  v3.5.0 68. Termin v3.5.0 6 and BLAS v3.5.0 6, 12. Bioxy-pub/) , 14. Unlxy-pub/) .Magnus is a myophage that has a 157,741-bp genome and a coding density of 92%, with 212 predicted protein-coding genes, 71 of which have a predicted function. The Magnus genome also contains 5 tRNA genes . Its G+C content was determined to be 46.3%. It was predicted by PhageTerm that Magnus uses a headful packaging system .Klebsiella phage 0507KN21 (GenBank accession number AB797215) (Serratia phage vB_Sru_IME250 (GenBank accession number KY073123) and Shigella phage Ag3 (GenBank accession number FJ373894), all classified within the Ackermannviridae. Additionally, Magnus contains at least 27 genes with significant amino acid similarity to type phages T4 (GenBank accession number NC_000866) and K (GenBank accession number KF766114), including core structural and replication proteins. The large terminase gene (NCBI accession number QEG07939) harbors an intein element.By BLASTp and progressiveMauve analyses, Magnus has 75.15% nucleotide identity to, and shares 146 similar proteins with, B797215) . Magnus MN045230, BioProject accession number PRJNA222858, SRA accession number SRR8869229, and BioSample accession number SAMN11360383.The genome sequence and associated data for phage Magnus were deposited under GenBank accession number"}
+{"text": "Psychrobacter spp. Analysis of Arctic psychrophilic Psychrobacter sp. DAB_AL32B genome content provided an insight into its overall stress response, and genes conferring protection against various life-limiting factors  were recognized and described. Moreover, it was revealed that the strain carries a large plasmid pP32BP2. Its replication system was used for the construction of two novel shuttle vectors (pPS-NR\u2014Psychrobacter-Escherichia coli-specific plasmid and pPS-BR\u2014Psychrobacter-various Proteobacteria-specific plasmid) of an increased carrying capacity, which may be used for genetic engineering of Psychrobacter spp.Cold-active bacteria are currently of great interest in biotechnology, and their genomic and physiological features have been extensively studied. One of the model psychrotolerant bacteria are The online version of this article (10.1007/s00203-018-1595-y) contains supplementary material, which is available to authorized users. Psychrobacter spp. belong to the Moraxellaceae family (Gammaproteobacteria). Bacteria of this genus are frequently isolated from various cold environments, including seawater, ice, permafrost and Arctic and Antarctic ornithogenic soils. Some strains  are considered to be opportunistic pathogens, as they are occasionally isolated from human patients, as well as from infected animals , 77 genomes (including 65 drafts) and an additional 65 plasmid sequences of Psychrobacter spp. are available (22nd June 2018). Analysis of the Psychrobacter genomes revealed the presence of various adaptation mechanisms allowing their survival in extremely cold environments. Hence, psychrophilic Psychrobacter strains are model, cold-active bacteria useful for studying bacterial adaptation to extreme conditions  a pABW1 vector containing a ColE1-type replication system. Therefore, the broad host range vector pBBR1 MCS-2 with pBBR1-type replication system was tested. Surprisingly, this replication system was also found to be inactive in tested Psychrobacter spp. These negative results encouraged us to construct novel, shuttle Psychrobacter\u2013Escherichia coli-specific vectors.It was previously reported that plasmids carrying ColE1- and p15a-type replication systems were stably maintained in E. coli-specific replication system originating from the pMB1 plasmid, i.e., high copy number replicon) or pWSK29  plasmids, and carrying replication systems of the pP43BP3 and pP43BP4 plasmids originating from psychrophilic Psychrobacter sp. DAB_AL43B, were reported , based on the pBGS18  was obtained. The genome analyses provided brief insights into the stress adaptation mechanisms of the strain. Moreover, we found that the strain carries a large, approximately 60-kb, plasmid. Although this plasmid was not fully assembled during genome drafting, its replication system was used for the construction of two novel Psychrobacter-specific vectors that are suitable for cloning of large DNA fragments.In this study, the draft genome sequence of Psychrobacter spp.) or 37\u00a0\u00b0C (Escherichia coli DH5\u03b1). The medium was solidified by the addition of 1.5% (w/v) agar. Where necessary, the media were supplemented with X-gal, IPTG and antibiotics: kanamycin (20\u00a0\u00b5g/ml for Psychrobacter spp. or 50\u00a0\u00b5g/ml for E. coli) and rifampin (50\u00a0\u00b5g/ml).The bacterial strains and plasmids used in this study are listed in Table\u00a0The CTAB/lysozyme method was used for isolation of genomic DNA .ACCGGTGCGAACCACTGTGAGTATTG-3\u2032 and 32B_REPR\u20145\u2032-ATACCGGTTTAATTCTATCGCCCGCCTG-3\u2032. To obtain the PCR product, the following conditions were applied: 35 cycles  preceded by 3-min denaturation at 95\u00a0\u00b0C and followed by 2-min extension at 72\u00a0\u00b0C.Plasmid DNA was isolated using a GeneMATRIX Plasmid Miniprep DNA Purification Kit  and alkaline lysis method  was specified as taxon separating from other bacterial taxa.Genome completeness was assessed by the presence/absence of bacterial orthologs according to the OrthoDB database using BUSCO  available at the Ribosomal Database Project (RDP) website (Nawrocki and Eddy P. adeliensis DSM 15333T (HE654007.1), P. aestuarii SC35 (EU939718.1), P. alimentarius JG-100 (AY513645.1), P. allis E2 (JX122558.1), P. aquaticus CMS 56 (NR_042206.1), P. aquimaris SW-210 (AY722804.1), P. arcticus 273-4 (NR_075054.1), P. arenosus R7T (AJ609273.1), P. celer SW-238 (NR_043225.1), P. ciconiae 176/10 (KM486054.1), P. cryohalolentis K5 (NR_075055.1), P. faecalis DSM 14664 (NR_118025.1), P. fjordensis BSw21516B (GQ358940.1), P. fozii NF23 (NR_025531.1), P. frigidicola DSM 12411 (NR_042222.1), P. fulvigenes KC 40 (NR_041688.1), P. glacialis DD43 (AJ539102.1), P. glaciei BIc20019 (NR_148850.1), P. glacincola DSM 12194 (NR_042076.1), P. halophilus DD2 (AJ539103.1), P. immobilis ATCC 43116 (NR_118808.1), P. jeotgali YKJ-103 (NR_025205.1), P. luti NF11 (NR_025532.1), P. lutiphocae IMMIB L-1110 (NR_044602.1), P. marincola KMM 277 (NR_025458.1), P. maritimus Pi2-20 (NR_027225.1), P. meningitidis SBA4 (KR091838.1), P. muriicola 2pS (NR_114669.1), P. namhaensis SW-242 (AY722805.1), P. nivimaris 88/2\u20137 (AJ313425.1), P. oceani 4k5 (AB910522.1), P. okhotskensis MD17 (NR_024806.1), P. pacificensis NBRC 103191 (NR_114238.1), P. pasteuri CIP110853 (KY292376.1), P. phenylpyruvicus ATCC 23333 (NR_118815.1), P. piechaudii CIP110854 (KY292375.1), P. piscatorii T-3-2 (NR_112807.1), P. piscidermidis 45 (FJ613616.1), P. pocilloporae S6-60 (KT444699.2), P. proteolyticus HAMBI 2948 (LT899990.1), P. psychrophilus BBDP29 (DQ337513.1), P. pulmonis CCUG 46240 (NR_118026.1), P. salsus DD 48 (NR_042166.1), P. sanguinis 13983 (HM212668.1), P. submarinus KMM 225 (AJ309940.1), P. urativorans DSM 14009T (AJ609555.1), P. vallis CMS 39 (AJ584832.1), P. cibarius JG-219 (AY639871.1), Psychrobacter sp. DAB_AL32B (JF714884.1) and Psychrobacter sp. DAB_AL43B (JF714885.1).Phylogenetic analysis was performed based on the comparison of partial 16S rRNA gene sequences of the DAB_AL32B strain, DAB_AL43B strain and strains representing 48 Psychrobacter sp. DAB_AL32B draft genome sequence has been deposited in the GenBank (NCBI) database under accession numbers NEXU01000001-NEXU01000218 and the sequences of vectors constructed in this study under accession numbers MH539767 (pPS-BR) and MH539768 (pPS-NR).The Psychrobacter spp. genomes available in the NCBI database (the average length of the Psychrobacter genome is 3.07\u00a0Mb). An assembly of the DAB_AL32B draft genome resulted in 37 scaffolds composed of 218 contigs. The average GC content is 41.9%, which is typical for Psychrobacter spp. . An automatic annotation of the DAB_AL32B genome performed applying the NCBI Prokaryotic Genome Annotation Pipeline resulted in 2,799 predicted genes with an average length of 973.43\u00a0bp, which covers about 80.08% of the genome. Additionally, 41 tRNA genes were identified .The estimated size of the DAB_AL32B genome is 3,211,529\u00a0bp, which is similar to the size of other Gammaproteobacteria BUSCO set (containing 452 BUSCO groups), was performed. The gene set predicted within the DAB_AL32B genome contains 94.3% of genes present in an applied BUSCO set (92.5% complete and 1.8% fragmented genes), which suggests that the genome assembly is complete or very close to completeness.To estimate the completeness of the obtained draft genome, a BUSCO analysis, using Psychrobacter genomes, with only short fragments homologous to genomes of phylogenetically closely related Moraxella and Acinetobacter. Other two contigs (contig00048 and contig00081) exhibit similarity to Psychrobacter genomes in majority of their fragments, while the remaining fragments are either homologous to Acinetobacter, Moraxella, and other unrelated bacteria, or not homologous to any sequence in the nr database. However, BLASTx search revealed significant similarity to proteins encoded within most of these fragments to Psychrobacter and Acinetobacter proteins. For three other contigs  the best BLAST hits were sequences from either Acinetobacter or Moraxella; however, they showed comparable similarity to Psychrobacter genomes. One contig (contig00082) was mistakenly automatically classified as a contaminant, because of a non-significant hit to zebrafish genome in low-complexity region. As a result of above analysis, these 15 contigs were recategorized as non-contaminant sequences. Seven other contigs  exhibited no significant homology to sequences available in the GenBank database , therefore it is unclear if these are contaminants or not. Finally, only nine contigs   may be considered as potential contaminants, as they do not show similarities to neither Psychrobacter sequences nor closely related bacteria.To examine if obtained draft genome is contaminated with other genomic sequences, an analysis using Taxoblast was performed . This analysis revealed that 30 out of 218 contigs were classified as possible contaminants or did not exhibit significant homology to sequences available in the GenBank database. These contigs were manually examined. Putative prophage was found within contig00038. Seven contigs  mostly exhibited homology to Psychrobacter frigidicola DSM 12411. As several new Psychrobacter species have been described since the last analysis, we performed another analysis with dataset extended on 14 new Psychrobacter-type strains  in NCBI database . Unfortunately, the genome of P. frigidicola DSM 12411 or some other representative of this species, is not publicly available. The phylogenomic analysis revealed that the DAB_AL32B strain is most related with Psychrobacter sp. DAB_AL43B . However, to predict if these strains may be considered as representatives of novel Psychrobacter species, the calculation of the ANI value with P. frigidicola is needed.For more precise classification of DAB_AL32B at the species level, the ANI analysis, based on whole-genome comparison between various Psychrobacter sp. DAB_AL32B had to cope not only with low temperatures, but also increased ultraviolet (UV) radiation, osmotic pressure and oxidative stress. The genes involved in the response to oxidative, osmotic, and cold shock encoded within the DAB_AL32B genome are listed in Table\u00a0Various adaptive mechanisms enable survival of psychrophilic bacteria in Arctic. Identification and analysis of genes encoding various stress response proteins of psychrophiles is important for studying their biology and adaptation, as well as for proper recognition of their biotechnological potential , lipids cross-linking with proteins and disturbances to membrane structure affecting its fluidity are observed  formed in the cell. Increased ROS formation in cells may occur as a result of (i) depletion of the UV-protective ozone layer in the Arctic region , peroxidase (1), superoxide dismutase (1), peroxiredoxin (2), rubredoxin (1) and glutaredoxins (3) Table\u00a0.2+). Hydroxyl radicals are then produced via Fenton reaction (Touati 2+ ions) and formation of very stable complexes with DNA , decreasing gene expression and membrane fluidity  from the environment or synthesise them in elevated quantities. Compatible solutes increase the stability of proteins and cell membranes without interfering with cellular function , cryptic plasmid pP32BP1  within the N-terminal part of this protein and winged helix DNA-binding domain (residues 1\u2013136 and 140\u2013233). Upstream to the repA gene, the putative origin of replication (oriV) was found . It is composed of five 20-bp-long direct repeats DR1.1-1.5 (5\u2032-ATAACACTAAATGATGTGGG-3\u2032) followed by a pair of 19-bp long inverted repeats, IR1 and IR2   of pP32BP2 shows 76% identity with RepB protein (GenBank: AFD62164) of pP62BP1 of parS, were found. ParA contains the ParA domain (residues 4-182) and shows 93% identity to ParA-like protein (GenBank: ABE76262) encoded within the plasmid 1 of Psychrobacter cryohalolentis K5. Within the ParB protein, the ParG domain, specific to the ParB-like proteins of, e.g., Salmonella enterica plasmid TP228 and Pseudomonas alcaligenes plasmid pRA2, was found (Hayes Psychrobacter urativorans R10.10B. The putative parS centomer-like sequence is located upstream to the parA gene. It consists of eight direct repeats DR2.1-2.8 (5\u2032-(A/T)ATACTCA-3\u2032) , and centromer-like sequence nd Hayes . The prePsychrobacter-specific cloning vectors. Currently, all available Psychrobacter-specific vectors are based on replication systems of small (not exceeding 6.4\u00a0kb) plasmids, which may exclude their application as molecular tools for cloning of large genetic modules. Since Psychrobacter spp. recently became a model bacterium representing cold-active microorganisms  plasmid pP32BP2, encouraged us to use this system for the construction of novel Psychrobacter spp., were constructed . Its functionality was confirmed by introducing it  into two Psychrobacter spp. strains, DAB_AL12R and DAB_AL43BR. Vector stability was tested in both these strains, and after approximately 30 generations of growth without antibiotic selection pressure, plasmid pPS-NR was found in 21% and 61% of the host cells, respectively . The vector was successfully introduced via triparental mating into Psychrobacter sp. DAB_AL12R and DAB_AL43BR. To analyse the carrying capacity of the pPS-BR vector, we cloned and successfully introduced to Psychrobacter spp. two relatively large restriction fragments of the DAB_AL32B genome, of a length of 5.8\u00a0kb (approx. the size of the vector) and 12.7\u00a0kb (approx. two times bigger than the vector), respectively.For the construction of this vector, the PCR-amplified (1.8-kb) DNA fragment containing the replication system of the pP32BP2 plasmid was cloned within PfoI site of the pBBR1 MCS-2 vector. This resulted in construction of a unique Psychrobacter spp. and E. coli (pPS-NR) or various Proteobacteria (pPS-BR), (iii) the system enabling mobilization to conjugal transfer, (iv) aphA gene-conferring resistance to kanamycin, (v) multiple cloning site (MCS) and (vi) a selection marker (lacZ\u2019 gene) enabling blue-white screening of clones in E. coli  two replication systems, which enable their replication in oli Fig.\u00a0. The obtBelow is the link to the electronic supplementary material.Supplementary material 1 (DOCX 308 KB)Supplementary material 2 (DOCX 50 KB)Supplementary material 3 (DOCX 31 KB)"}
+{"text": "In this work, we investigate the self-assembly between Ag(I) and Au(I) centers and pyridyl donors to form hexagonal metallacycles and related linear complexes. The precipitation of hexagonal metallacycles upon assembly in chloroform/methanol mixtures results in high solid-state photo-stability. Whereas, the Ag(I) species have fast kinetics and high formation constants in acetone, this solvent interferes in the formation of the analogous Au(I) complexes. The photophysical properties of this suite of metallacycles was investigated including steady-state absorption, emission, and time-resolved lifetime measurements. All ligands and hexagons exhibited ligand-centered singlet emissions with ground-state absorption and emission perturbed upon coordination. The ligand-based fluorescent photoluminescence was affected by the heavy-atom effect when halide or metals are present, attenuating quantum yields as evidenced by increases in the experimentally measured non-radiative rate constants. The formation of group 11 metallacycles is motivated by their potential applications in mixed-matrix materials wherein metal ions can interact with substrate to facilitate separations chemistry with reduced energy requirements, in particular the isolation of ethylene and light olefins. Existing processes involve cryogenic distillation, an energy intensive and inefficient method. Coordination-driven self-assembly  precursors  , acetone, dichloromethane (DCM), ethyl acetate (EtOAc), n-hexanes, toluene, diethyl ether, petroleum ether, potassium hydroxide (KOH), sodium sulfate (Na2SO4) and potassium carbonate (K2CO3) were purchased from Fisher Scientific. N,N-dimethylformamide (DMF) was purchased from Macron Fine Chemicals. Tetrahydrofuran (THF), triethylamine (NEt3), and chloroform (CHCl3) were purchased from EMD Millipore. Ethanol  was purchased from Decon Labs, Inc. Tetrakis(triphenylphosphine)palladium(0) (Pd(PPh3)4), and bis(triphenylphosphine)palladium(II) chloride (Pd(PPh3)2Cl2), and 1,3-dibromobenzene were purchased from Matrix Scientific. 4-pyridinylboronic acid (pyB(OH)2) was purchased from AK Scientific. Copper (I) iodide (CuI) was purchased from Strem Chemicals. Tetrahydrothiophene (tht) was purchased from Aldrich. Silver(I) hexafluorophosphate (AgPF6) was purchased from Oakwood Chemical. Hydrogen tetrachloroaurate(III) trihydrate (HAuCl4\u00b73H2O) was purchased from J&J Materials Inc. 4-ethynylpyridine hydrochloride and 1,3,5-tribromobenzene were purchased from Ark Pharm Inc. Deionized water was used whenever water was needed. THF was purified and dried through a Pure Process Technology free standing solvent purification system. NEt3 was purified and dried by distillation in KOH. DMF was filtered and dried using activated molecular sieves under N2 and freeze-pump-thawed with water before use for synthesis.All reagents and solvents were reagent grade and used as received without further purification unless noted. Methanol (CH1H Nuclear Magnetic Resonance (NMR) spectra were recorded on either Varian 300 or 400 MHz spectrometer. Chemical shifts (\u03b4) are reported in parts per million (ppm) referenced using the residual protio-solvent peaks as internal standards. Coupling constants (J) are quoted in Hertz (Hz), and the following abbreviations are used to describe the signal multiplicities: s (singlet); d (doublet); t (triplet); q (quartet); m (multiplet). Fourier Transform-Ion Cyclotron Resonance (FT-ICR) mass spectra were acquired using a Bruker Daltonics SolariX 12T FT-ICR Mass Spectrometer that was calibrated with > 90% Angiotensin I purchased from Sigma Aldrich under either Electrospray Ionization (ESI) or Laser Desorption Ionization (LDI) modes. Single Crystal X-ray Diffraction (SC-XRD) Crystallography was performed using a Bruker D8 Venture diffractometer in a fixed-chi geometry, equipped with a Photon-100 CMOS area detector, Oxford Cryosystems cryostat, a molybdenum source , and a graphite monochromator. Fourier Transform Infrared Resonance (FT-IR) spectra were collected from a Perkin Elmer 1760 FT-IR spectrometer equipped with horizontal attenuated total reflectance (HATR) from 4,000 to 500 cm\u22121 wavenumbers (\u03bd). The following abbreviations are used to describe signal intensities: w (weak); m (medium); s (strong). All UV-Vis absorption spectra were acquired from an Agilent Cary 8454 UV-Vis Diode Array system. Blank spectra with pure solvents were acquired before each run. All emission spectra were collected using a Horiba Scientific FluoroMax-4 Spectrofluorometer equipped with Quanta-\u03c6 Integrating light sphere with Spectralon coating for quantum yield (\u03a6) measurements and Delta-Hub DH-HT High Throughput Time-correlated Single Photon Counting (TCSPC) Controller and Nano-LED NL-C2 pulsed-diode controller with N-350 NanoLED Source for lifetime measurements. All solution-state absorbance and emission spectra were collected using a 10-mm rectangular quartz cuvette from Starna Cells Inc.L12 complex was self-assembled using a modified literature procedure  AgPF6 was dissolved in 80.0 mL MeOH then layered over the L1 solution. After 48 h, white needle-like crystals form. These crystals were filtered and washed with 10-mL portions of CHCl3 (3x) to afford 1.19 g (100% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.81 , 7.82 , 7.77 , 7.71 , 7.65 , 7.46 . FT-IR : 3069 (w), 2235 (w), 2187 (w), 1609 (m), 1558 (w), 1506 (w), 1470 (w), 1433 (w), 1405 (w), 1223 (w), 1152 (w), 1070 (w), 1031 (w), 991 (w), 915 (w), 881 (m), 815 (m), 775 (m), 757 (m), 743 (m), 678 (m), 663 (m), 553 (m), 531 (m).AgL2}6 hexagon, was self-assembled using a modified literature procedure  AgPF6 was dissolved in 5.0 mL MeOH then layered over the L2 solution. After 48 h, white powdered product formed which was filtered and washed with 10-mL portions of CHCl3 (3x) to afford 0.062 g (73% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.72 , 7.90 , 7.83 , 7.62 . FT-IR : 3076 (w), 2219 (w), 1613 (m), 1557 (w), 1507 (w), 1435 (m), 1338 (w), 1291 (w), 1221 (m), 1174 (w), 1162 (w), 1136 (w), 1107 (w), 1066 (w), 1028 (w), 993 (w), 964 (w), 864 (m), 821 (s), 748 (w), 736 (w), 663 (m), 573 (w), 552 (s).{AgL3}6 hexagon, was self-assembled using a modified literature procedure  AgPF6 was dissolved in 5.0 mL MeOH then layered over the L3 solution. After 48 h, white powdered product formed which was filtered and washed with 10-mL portions of CHCl3 (3x) to afford 0.096 g (100% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.74 , 7.86 , 7.74 , 7.66 , 7.63 \u2013 7.56 . FT-IR : 3107 (w), 2203 (w), 1615 (m), 1543 (w), 1501 (w), 1431 (w), 1227 (w), 1167 (w), 1064 (w), 1025 (w), 878 (w), 821 (m), 752 (w), 733 (w), 682 (w), 556 (m), 541 (m).{AgL4}6 hexagon, was self-assembled using a modified literature procedure  AgPF6 was dissolved in 2.0 mL MeOH then layered over the L4 solution. After 48 h, white powdered product formed which was filtered and washed with 10-mL portions of CHCl3 (3x) to afford 0.052 g (92% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.76 , 8.25 , 8.00\u20137.87 , 7.80\u20137.71 . FT-IR : 3092 (w), 1618 (m), 1545 (w), 1513 (w), 1478 (w), 1431 (w), 1411 (w), 1314 (w), 1230 (w), 1069 (w), 1031 (w), 1019 (w), 873 (m), 818 (s), 791 (s), 736 (m), 695 (m), 663 (w), 637 (m), 613 (w), 555 (s).{AgL12 complex was self-assembled using a modified literature procedure  Au(tht)Cl were dissolved in 6.0 mL acetone. After 6 h, the solution was concentrated in vacuo and the collected solid was washed with 6-mL portions of CHCl3 (3x) to afford 0.007 g (20% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.84 , 7.89 , 7.80 , 7.68 , 7.50 \u2013 7.40 . FT-IR : 3044 (w), 2918 (w), 2856 (w), 2218 (m), 2187 (w), 1943 (w), 1801 (w), 1614 (m), 1588 (m), 1558 (w), 1496 (w), 1463 (w), 1429 (w), 1404 (w), 1261 (w), 1234 (w), 1217 (w), 1158 (w), 1105 (w), 1088 (w), 1060 (w), 1038 (w), 987 (w), 906 (w), 876 (w), 836 (m), 814 (w), 789 (m), 747 (w), 713 (w), 691 (w), 680 (m), 655 (m), 565 (w), 535 (m).AuL2}6 hexagon was self-assembled using a modified literature procedure  Au(tht)Cl were dissolved in 6.0 mL CHCl3. After 24 h, the solution was concentrated in vacuo and the collected yellow solid was washed with 6-mL portions of CHCl3 (3x) to afford 0.025 g (78% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.84\u20138.71 , 8.09 \u2013 7.82 . FT-IR : 3100 (w), 3044 (w), 2210 (m), 1615 (s), 1553 (w), 1496 (w), 1430 (m), 1336 (w), 1288 (w), 1211 (m), 1171 (w), 1134 (w), 1105 (w), 1063 (m), 1046 (w), 992 (w), 966 (w), 863 (m), 826 (m), 761 (w), 721 (w), 672 (m), 587 (w), 574 (m), 527 (w).{AuL3}6 hexagon was self-assembled using a modified literature procedure  Au(tht)Cl were dissolved in 6.0 mL CHCl3. After 24 h, the solution was concentrated in vacuo and the collected yellow solid was washed with 6-mL portions of CHCl3 (3x) to afford 0.026 g (89% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.76 , 8.08 \u2013 7.70 , 7.66\u20137.57 . FT-IR : 3092 (w), 3029 (w), 2203 (m), 1943 (w), 1609 (s), 1529 (w), 1492 (w), 1427 (w), 1324 (w), 1213 (w), 1162 (w), 1150 (w), 1125 (w), 1093 (w), 1065 (w), 1042 (w), 991 (w), 942 (w), 902 (w), 831 (m), 792 (w), 754 (w), 729 (w), 666 (w), 569 (w), 544 (w).{AuL4}6 hexagon was self-assembled using a modified literature procedure  Au(tht)Cl were dissolved in 6.0 mL CHCl3. After 24 h, the solution was concentrated in vacuo and the collected white solid was washed with 6-mL portions of CHCl3 (3x) to afford 0.027 g (100% yield) isolated product. 1H NMR : \u03b4 (ppm) = 8.82 , 8.35 , 8.22 , 8.11 , 7.86 . FT-IR : 3060 (w), 1611 (s), 1546 (w), 1508 (w), 1472 (w), 1443 (w), 1427 (w), 1398 (m), 1340 (w), 1318 (w), 1266 (w), 1231 (m), 1186 (w), 1109 (w), 1076 (m), 1034 (w), 970 (w), 935 (w), 915 (w), 857 (w), 832 (m), 786 (s), 761 (w), 732 (m), 687 (w), 667 (w), 645 (m), 632 (w), 606 (w), 535 (s).{AuL1), 5-bromo-1,3-bis(4-ethynylpyridyl)benzene (L2), 1,3-bis(4-ethynylpyridyl)benzene (L3), and 1,3-bis(4-pyridyl)benzene (L4), were prepared as donors for self-assembly reactions. As illustrated in L1, L2, and L3 were prepared following a typical Sonogashira coupling reaction while L4 was prepared following a typical Suzuki coupling reaction. The 1H NMR spectra of L1, L2, L3, and L4 in acetone-d6 are summarized in L11H spectrum shows the pyridyl-H doublets at 8.63 and 7.49 ppm and the benzyl-H singlet at 7.78 ppm, two doublets at 7.65 and 7.60 ppm, and triplet at 7.42 ppm. The L2 spectrum shows the pyridyl-H doublets at 8.66 and 7.53 ppm and the benzyl-H singlets at 7.87 and 7.82 ppm. The L3 spectrum shows the pyridyl-H doublets at 8.65 and 7.51 ppm and the benzyl-H singlet at 7.83 ppm, doublet at 7.69 ppm, and triplet at 7.56 ppm. The L4 spectrum shows the pyridyl-H doublets at 8.68 and 7.78 ppm and the benzyl-H singlet at 8.17 ppm, doublet at 7.89 ppm, and triplet at 7.70 ppm.Four pyridyl ligands, 1-bromo-3(4-ethynylpyridyl)benzene (L1 was used to synthesize linear Ag(I) and Au(I) complexes, while L2, L3, and L4 were used to self-assemble Ag(I) and Au(I) hexagonal metallacycles. The general method used to self-assemble the complexes and hexagons is shown in L12, was prepared by layering one equivalent of AgPF6 in MeOH over two equivalents of L1 in CHCl3. This resulted in the crystallization of AgL12 after 48 h. Ag(I) hexagons, {AgL}6 where L = L2, L3, and L4, were prepared by layering one equivalent of AgPF6 in MeOH over one equivalent of L in CHCl3. The hexagons precipitated at almost quantitative yields as white powders after 48 h. The gold analogs, Au(I) hexagons, {AuL}6 where L = L2, L3, and L4, were prepared by mixing 1:1 equivalents of Au(tht)Cl and L in CHCl3. After 24 h, powdered metallacycles were isolated upon solvent removal and CHCl3 washing. In the case of the monomeric Au(I) complex, AuL12, previous attempts using pure CHCl3 or 1:1 (v/v) CHCl3/MeOH did not result in any complex formation. However, the complex was isolated at a relatively low yield by mixing 1:1 equivalents of Au(tht)Cl and L1 in acetone.N,N'-dimethylsulfoxide (DMSO), which are typically used in solution-state structural analyses of Ag(I) and Ag(I) complexes proved to be unsuitable for our complexes and hexagons  and Au(I) complex and hexagons is their limited solubilities. Coordinating solvents such as acetonitrile (MeCN) and ns Laye, . In all 1H NMR peaks as well as peak integration agreements  complexes and hexagons  as well reements . In addi1H NMR spectra of {AuL2}6, {AuL3}6, and {AuL4}6. The full spectra of each Au(I) hexagon are shown in 1H NMR peaks, the peak integration agreements, and the similarity in peak splitting patterns indicate the formation of pure Au(I) hexagonal metallacycles. Although the solubility of these species is limited, our identification of NMR-suitable solvents rules out the formation of polymeric species as such coordination polymers are fully insoluble. ESI-MS (see below) further confirms the stoichiometries of self-assembly.For the Au(I) hexagons, ligand dissociation occurs when MeCN, DMSO, and acetone were used as solvents. These hexagons are insoluble in all other solvents except for nitromethane, in which we were able to observe intact Au(I) hexagons. 3 or in CHCl3 mixtures with MeOH. MeCN and DMSO competitively coordinate with the Ag(I) and Au(I) resulting in ligand dissociation. Acetone was a suitable solvent for the building blocks, as well as the final assemblies, enabling the study of complex formation. Using 1H NMR methods, we initially acquired a series of spectra for a solution containing 1:1 equivalents of Ag:L in acetone upon mixing and after 24 h (All of these complexes were synthesized in pure CHClter 24 h . We obseL in acetone to ensure complete ligand coordination. The acquired 1H NMR spectra at 0 (upon mixing), 6, 24, and 48 h were summarized in L12 complex self-assembles within 6 h then partially dissociates thereafter. In the case of both {AuL2}6 and {AuL3}6, an equilibrium was established immediately upon mixing that stays up to at least 48 h. Products of {AuL4}6 hexagon assembly crash out of solution immediately upon mixing such that we only observe uncoordinated L4 and partially dissolved products. In all cases, the initial coordination is fast; however, quantitative analyses of the NMR peaks is not straightforward using a one-step metal-ligand binding model. There are reports on the mechanism and reducing ability of acetone to form Au nanoparticles from Au(III) and Au(I)  complex and hexagons. In this case, we prepared solutions containing 1:1, 2:1, and 3:1 equivalents of Au:High-resolution mass spectrometry (HRMS) was used to probe the stoichiometry of our self-assemblies. We used Fourier transform-ion cyclotron mass spectrometry (FT-ICR MS) with electrospray ionization (ESI) for soluble complexes and hexagons, while laser desorption ionization (LDI) was used for hexagons that are unstable in acetone. ESI, as a soft ionization technique, is advantageous and popularly used in detection of intact coordination complexes and cages with high molar masses  complex and hexagons and for Au(I) complex and hexagons are shown in  complex  as well  hexagon  and + complex (\u2212 counter-ion by ESI-FT-ICR MS. The Au(I) hexagons, which are unstable is acetone were detected by LDI-FT-ICR MS. We were able to detect singly-charged intact [{AuL2}6+AuCl+K]+ hexagon (L3}6+Na]+ hexagon (L4}6+Na]+ hexagon (L}6 fragments (For the Au(I) complex and hexagons, we were able to detect the intact  cores that ionized by the loss of counterions.Silver(I) and gold(I) mononuclear complexes and hexagonal rings were self-assembled from AgPFLonger-term photostability of all complexes and hexagons in the solid-state were confirmed by FT-IR. Significant peak shifts to higher wavenumbers were observed upon coordination of the ligands to either Ag(I) or Au(I). No changes to the spectra were observed after storing the compounds at ambient conditions under room light for one week.L3 and L4 were the most and least emissive ligands, respectively. In the solid-state, all complexes and hexagons have diminished emission except for {AgL2}6, {AgL4}6, and {AuL4}6 wherein aggregation-induced emission was observed. In the solution-state, all ligands and hexagons exhibited ligand-centered singlet emissions while AgL12 and AuL12 both exhibited metal-perturbed ligand-centered singlet emissions.Ligand-centered fluorescence emission was observed for many of these complexes. In both the solid- and solution-state, This work establishes that Ag(I) and Au(I) centers are effective linear nodes for self-assembly reactions with dipyridyl ligands and that the resultant materials stabilize these ions with respect to oxidation and photodecomposition. Due to the modular nature of coordination-driven self-assembly, efforts are ongoing to exploit ligand-tuning to incorporate functional groups that will enhance metallacycle solubility to improve their processability for incorporation into mixed-matrix materials.L12 (CCDC No. 1879931) for this study can be found in the Cambridge Crystallographic Data Centre https://www.ccdc.cam.ac.uk/solutions/csd-system/components/csd/.The dataset AgCF designed and carried out the experiments and wrote the manuscript. SK assisted in ligand synthesis. AF carried out the mass spec characterization. TC designed the project and wrote the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Alcaligenes aquatilis strain QD168 (= CCUG 69566) is a marine hydrocarbon-degrading bacterium isolated from crude oil-polluted sediment from Quintero Bay, Central Chile. Here, we present the 4.32-Mb complete genome sequence of strain QD168, with 3,892 coding sequences, 58 tRNAs, and a 56.3% G+C content. Alcaligenes aquatilis strain QD168 (= CCUG 69566) is a marine hydrocarbon-degrading bacterium isolated from crude oil-polluted sediment from Quintero Bay, Central Chile. Here, we present the 4.32-Mb complete genome sequence of strain QD168, with 3,892 coding sequences, 58 tRNAs, and a 56.3% G+C content. Alcaligenes are Gram-negative, aerobic, and motile bacteria that are frequently isolated from water, soil, and clinical samples, as well as from industrial and contaminated environments , a marine hydrocarbon-degrading bacterium capable of growing on n-hexadecane and diesel in M9 minimal medium agar  in Bushnell Hass mineral medium and artificial seawater.Members of the genus ronments \u20136. Alcalg phenol , atrazing phenol , endosulg phenol , phenantg phenol , pyrene,a]pyrene . Here, wium agar  at 30\u00b0C.Genomic DNA of QD168 cells cultivated in Luria-Bertani broth was extracted using a Qiagen Genomic-tip 100/G kit . Next-generation sequencing was performed using a PacBio RS II platform  with one single-molecule real-time (SMRT) cell and an average 20-kb insert library, obtaining 87,836 reads with an average length of 12,761\u2009bp. PacBio reads were trimmed and assembled using the Hierarchical Genome Annotation Pipeline (HGAP) v3.0 , obtainiectABCD; D3M96_13270, D3M96_13265, D3M96_13260, and D3M96_13255) and the bioplastic polyhydroxyalkanoates  were identified. Aromatic catabolic genes encoding catechol 1,2-dioxygenase , catechol 2,3-dioxygenase , homogentisate 1,2-dioxygenase , protocatechuate 4,5-dioxygenase  and gentisate 1,2-dioxygenase  were identified.Gene functional annotation was performed using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v4.6 , identifhttps://www.ebi.ac.uk/Tools/psa/emboss_needle/) showed a close relationship (99.9% identity) with A. aquatilis LMG 22996T, while identities with those of the type strains of other Alcaligenes species were <98.5% (species threshold)   and A. faecalis DSM 30030T, resulted in ANIb values of 96.8% and 90.5% , respectively.Comparative 16S rRNA gene sequence analyses using EMBOSS Needle (reshold) . MultiloW v.1.81  and manuT (ANIb) , using JT (ANIb)  with A. A. aquatilis strain.This is the first complete genome sequence of an Alcaligenes aquatilis strain QD168 has been deposited in DDBJ/ENA/GenBank under the accession number CP032153. The version described in this paper is the first version, CP032153.1. The accession number for the publicly available raw data at NCBI is PRJNA489687.The genome sequence of"}
+{"text": "Klebsiella pneumoniae represents an emerging public health issue. Here, we present the draft whole-genome sequences of K. pneumoniae clinical strains KPL0.1 (OXA-48 carbapenemase) and KPL0.2 (NDM-1 carbapenemase).Carbapenemase-producing  Klebsiella pneumoniae represents an emerging public health issue. Here, we present the draft whole-genome sequences of K. pneumoniae clinical strains KPL0.1 (OXA-48 carbapenemase) and KPL0.2 (NDM-1 carbapenemase). These genome sequences should help in investigating pathophysiological mechanisms of digestive colonization or infection with these highly resistant bacteria.Carbapenemase-producing  Enterobacteriaceae (CPE) are resistant to most beta-lactams, including last-line options such as carbapenems, and are an emerging public health issue , KPL0.1 (producing OXA-48 carbapenemase) and KPL0.2 (producing NDM-1 carbapenemase). Both strains were isolated from blood cultures using the Virtuo automated system  and identified by matrix-assisted laser desorption ionization\u2013time of flight (MALDI-TOF) mass spectrometry . KPL0.1 was isolated from a 77-year-old patient following digestive surgery, and KPL0.2 came from an 84-year-old patient with a urinary tract infection.Carbapenemase-producing th issue . Klebsieescribed . In Franescribed . Two strde novo assembled with Unicycler. Genomes were annotated using the Prokaryotic Genome Annotation Pipeline (PGAP) v4.5 , and quality control was performed using a Qubit v3.0 fluorometer . Libraries were prepared using the Nextera XT DNA Library prep kit , followed by paired-end 2 \u00d7 150-bp) sequencing on a HiSeq platform (Illumina). Genome coverage was about 650\u00d7 for each isolate. Paired reads were filtered, and Nextera adapters were removed using Trimmomatic on a Galaxy server using default settings . Process0-bp sequblaOXA-48, blaSHV-28, blaTEM-1B, blaCTX-M-15, blaOXA-1), four aminoglycoside resistance genes , five fluoroquinolone resistance genes , and individual antimicrobial resistance genes .The draft genome of KPL0.1 consists of 5,825,863\u2009bp and has a mean G+C content of 56.70%. A total of 5,628 protein-coding genes were annotated, including 92 RNA-coding genes and 75 tRNAs, and the remaining genes were annotated as hypothetical proteins. KPL0.1 belongs to sequence type 307 (ST-307) and possesses the KL102 capsule locus , four aminoglycoside resistance genes , and individual antimicrobial resistance genes .The draft genome of KPL0.2 consists of 5,741,089\u2009bp with a mean G+C content of 56.94%. A total of 5,570 protein-coding genes were annotated, including 93 RNA-coding genes and 79 tRNAs, and the remaining genes were annotated as hypothetical proteins. KPL0.2 belongs to ST-147 and possesses the KL64 capsule locus. The following resistance genes were predicted: five beta-lactam resistance genes  and PYBH00000000 (KPL0.2). The versions described in this paper are QHMA01000000 (KPL0.1) and PYBH01000000 (KPL0.2). The SRA accession number is SRP155589.These two draft whole-genome shotgun projects have been deposited at DDBJ/ENA/GenBank under the accession numbers"}
+{"text": "Spodoptera litura, and significantly influences the development and fecundity of S. litura at either lethal or sublethal doses. Herein, Illumina HiSeq Xten (IHX) platform was used to explore the transcriptome of S. litura and to identify genes responding to fluralaner exposure.Fluralaner is a novel isoxazoline insecticide with a unique action site on the \u03b3-aminobutyric acid receptor (GABAR), shows excellent activity on agricultural pests including the common cutworm S. litura transcriptome and annotated according to the COG, GO, KEGG and NR databases. These genes included 156 detoxification enzyme genes  and 24 insecticide-targeted genes . There were 3275 and 2491 differentially expressed genes (DEGs) in S. litura treated with LC30 or LC50 concentrations of fluralaner, respectively. Among the DEGs, 20 related to detoxification  and 5 were growth-related genes (1 chitin and 4 juvenile hormone synthesis genes). For 26 randomly selected DEGs, real-time quantitative PCR (RT-qPCR) results showed that the relative expression levels of genes encoding several P450s, GSTs, heat shock protein (HSP) 68, vacuolar protein sorting-associated protein 13 (VPSAP13), sodium-coupled monocarboxylate transporter 1 (SCMT1), pupal cuticle protein (PCP), protein takeout (PT) and low density lipoprotein receptor adapter protein 1-B (LDLRAP1-B) were significantly up-regulated. Conversely, genes encoding esterase, sulfotransferase 1C4, proton-coupled folate transporter, chitinase 10, gelsolin-related protein of 125\u2009kDa (GRP), fibroin heavy chain (FHC), fatty acid synthase and some P450s were significantly down-regulated in response to fluralaner.A total of 16,572 genes, including 451 newly identified genes, were observed in the S. litura whilst the DEGs identified sheds further light on the molecular response to fluralaner.The transcriptome in this study provides more effective resources for the further study of  Spodoptera litura Fabricius (Lepidoptera: Noctuidae), is a destructive polyphagous pest with a broad host plant range of more than 100 species of crops and vegetables  of fluralaner dissolved in acetone. After 24\u2009h, 15 alive and normal S. litura from each treatment were randomly collected, equally divided into three 1.5\u2009mL Eppendorf tubes, respectively, immediately frozen in liquid nitrogen and stored at \u2212\u200980\u2009\u00b0C until use.A laboratory strain of s report . Flurala. (2019) . The 3rdS. litura from each group  by TRIzol\u2122 Reagent . The concentration and integrity of total RNA were measured using NanoDrop 2000  and with the RNA Nano 6000 Assay Kit by Agilent Bioanalyzer 2100 system , respectively.Total RNA was isolated from whole body of RNA sequencing libraries were generated from each sample and the sequencing was carried out by BioMarker  following the manual of the NEB Next\u00ae Ultra\u2122 RNA Library Prep Kit  in Illumina. Briefly, 1\u2009\u03bcg of total RNA from each sample was enriched using magnetic beads with Oligo (dT) and broken into short fragments, which was carried out using divalent cations under elevated temperature by adding NEB Next First Strand Synthesis Reaction Buffer (5x) (NEB). These short fragments of mRNA served as templates for synthesis of the first-strand complementary DNA (cDNA) with random hexamer primers and Moloney Murine Leukemia Virus (M-MuLV) Reverse Transcriptase (NEB). Using buffer, dNTPs, RNase H and DNA Polymerase I, the second-strand cDNA was subsequently synthesized. Remaining overhangs were converted into blunt ends via exonuclease/ polymerase activities. After adenylation of 3\u2032 ends of DNA fragments, NEB Next Adaptor with hairpin loop were ligated for hybridization. The purified cDNAs were subjected to end repair by adding an adenosine triphosphate (A) base to the 3\u2032 end and connected with sequencing adaptor. Suitable fragments of cDNAs were extracted by the AMPure XP system . Three microliter of USER Enzyme (NEB) was used for size selection, and adaptor-ligated cDNAs at 37\u2009\u00b0C for 15\u2009min followed by 5\u2009min at 95\u2009\u00b0C before PCR. Then PCR was performed with Phusion High Fidelity DNA polymerase (NEB), Universal PCR primers, and Index (X) Primer (NEB). PCR products were purified with the AMPure XP system (Beckman Coulter) and library quality was assessed on the Agilent Bioanalyzer 2100 system (Agilent).The clustering of the index coded samples was carried out on a cBot Cluster Generation System (Illumina) using TruSeq PE Cluster Kit v4-cBot-HS (Illumina) according to the manufacturer\u2019s instruction. After cluster generation, the prepared libraries were sequenced on an Illumina platform and paired end reads were generated.S. litura . The clean reads with a perfect match or with only one mismatch were mapped to the reference genome of . litura  by the H. litura . The map. litura  and all . litura  accordin. litura , Gene On. litura , Kyoto E. litura  and Non-. litura . In addiS. litura transcriptome. Referring to the previous studies [S. litura, B. mori, D. melanogaster and A. mellifera, then the phylogenetic trees were mapped with 1000 bootstrap replications using the neighbor-joining method to evaluate the branch strength of each tree using MEGA 7 [Genes encoding detoxification enzymes including P450s, GSTs, CarEs, and insecticide-targeted genes including ionotropic GABARs, GluCl, VGSCs, nAChRs, RyR and AChEs, were identified from the  studies , 21, 62,g MEGA 7 . The phyg MEGA 7 . Referrig MEGA 7 , 48, 65,g MEGA 7  and Treeg MEGA 7 .30- and LC50-treated groups were analyzed using DEGseq method [>\u00a02) and false discovery rate (FDR <\u20090.01) from corrected P-value were used as a judgment standard for the significant differences in gene expression between two samples [Premix Ex Taq\u2122 II (Tli RNaseH Plus) \u2002Co.,\u2002Ltd., Liaoning, China) on a Quant Studio\u2122 6 and 7 Flex Real-Time PCR System  according to the procedures from Liu et al. (2018) [-\u25b3\u25b3Ct method [The relative expression levels of genes among the control, LCq method , which p samples . The DEG. (2018) . The hou. (2018) . Three tt method . The priAdditional file 1: Text S1. Gene annotation with COG and GO databases.Additional file 2. All pathways annotated by KEGG database.Additional file 3. Cytochrome P450 nucleotide sequences of the S. litura transcriptome.Additional file 4. Neighbor-joining phylogenetic analysis of the P450s genes from D. melanogaster, H. armigera, P. xylostella,S. litura and B. mori.Additional file 5. Glutathione S-transferase nucleotide sequences of the S. litura transcriptome.Additional file 6. Neighbor-joining phylogenetic analysis of the GST genes from D. melanogaster, A. mellifera,S. litura and B. mori.Additional file 7. Carboxylesterase nucleotide sequence of the S. litura transcriptome.Additional file 8. Neighbor-joining phylogenetic analysis of the CarE genes from D. melanogaster, H. armigera, P. xylostella,S. litura and B. mori.Additional file 9. Nucleotide sequences of annotated insecticide-targeted genes of the S. litura transcriptome.Additional file 10. Neighbor-joining phylogenetic analysis of the cys-loop ligand-gated ion channel superfamily genes from S. litura and other species. The subunits sequence used are as follow: Am-alpha1 (AAY87890.1), Am-alpha2 (AAS48080.1), Am-alpha3 (AAY87891.1), Am-alpha4 (AAY87892.1), Am-alpha5 (AJE70263.1), Am-alpha6 (AAY87894.1), Am-alpha7 (AAR92109.1), Am-alpha8 (NP_001011575.1), Am-alpha9 (AAY87896.1), Am-beta1 (AAY87897.1), Am-beta2 (AAY87898.1), Am-GluCl (ABG75737.1), Am-RDL (AJE68941.1), Am-GRD (AJE68942.1), Am-LCCH3 (AJE68943.1), Am-CG8916 (NP_001071290.1), Am-HisCl (ABG75739.1), Am-pHCl (ABG75741.1), Am-CG6927 (ABG75747.1), Am-CG12344 (ABG75746.1), Dm-alpha1 (AGB96296.1), Dm-alpha2 (NP_524482.1), Dm-alpha3 (NP_525079.3), Dm-alpha4 (NP_001097669.2), Dm-alpha5 (NP_995708.1), Dm-alpha6 (NP_995674.1), Dm-alpha7 (NP_996514.1), Dm-beta1 (NP_523927.2), Dm-beta2 (NP_524483.1), Dm-beta3 (AAF51485.1), Dm-RDL (NP_523991.2), Dm-LCCH3 (NP_996469.1), Dm-GRD (CAA55144.1), Dm-CG8916 (AAF48539.4), Dm-HisCl1 (AAL74413.1), Dm-HisCl2 (AAL74414.1), Dm-NtR (NP_651958.2), Dm-pHCl1 (NP_001034025.2), Dm-pHCl2 (NP_651861.1), Dm-CG6927 (AAF45992.1), Dm-CG7589 (AAF49337.2), Dm-CG12344 (NP_610619.2), Bm-alpha1 (XP_021203289.1), Bm-alpha2 (NP_001103397.1), Bm-alpha3 (NP_001103387.2), Bm-alpha4 (NP_001166816.1), Bm-alpha5 (NP_001166811.1), Bm-alpha6 (NP_001166813.1), Bm-alpha7 (NP_001166818.1), Bm-alpha8 (NP_001166817.1), Bm-alpha9 (ABV72691.1), Bm-beta1 (ABV72692.1), Bm-beta2 (ABV72693.1), Bm-beta3 (ABV45510.1), Bm-RDL1 (ADM88014.1), Bm-RDL2 (NP_001182629.1), Bm-RDL3 (NP_001182630.1), Bm-LCCH3 (BAT57341.1), Bm-GluCl (BAO58781.1), Bm-pHCl (BAX77827.1).Additional file 11. Phylogenetic analysis of the AChE genes.Additional file 12. Functional classification of DEGs according to COG database. Note: A represented the DEGs and all genes after the exposure of LC30 fluralaner, B represented the DEGs and all genes after the exposure of LC50 fluralaner, respectively.Additional file 13. Functional classification of DEGs according to KEGG database.Additional file 14. Changes of DEGs related to detoxification and development of S. litura after exposure of fluralaner.Additional file 15. Primers for RT-qPCR validation."}
+{"text": "Pneumonia and influenza (P&I) increase morbidity and mortality among older adults, especially those residing in long-term care facilities (LTCFs). Facility-level characteristics may affect P&I risk beyond resident-level determinants. However, the relationship between facility characteristics and P&I is poorly understood. We therefore identified potentially modifiable facility-level characteristics that might influence the incidence of P&I across LTCFs. We conducted a retrospective cohort study using 100% of 2013-2015 Medicare claims linked to Minimum Data Set 3.0 and LTCF-level data. Short-stay (<100 days) and long-stay (\u2265100 days) LTCF residents aged \u226565 were followed for the first occurrence of hospitalization, LTCF discharge, Medicare disenrollment, or death. We calculated LTCF risk-standardized incidence rates (RSIRs) per 100 person-years for P&I hospitalizations by adjusting for over 30 resident-level demographic and clinical covariates using hierarchical logistic regression. The final study cohorts included 1,767,241 short-stay  and 922,863 long-stay residents . LTCFs with lower RSIRs had more Physician Extenders (Nurse Practitioners or Physician\u2019s Assistants) among short-stay  and long-stay residents , higher Registered Nurse hours/resident/day among short-stay and long-stay residents (Mean (SD): 0.5 (0.7) vs. 0.4 (0.4), p<0.001), and fewer residents prescribed antipsychotics among short-stay (21.4% (11.6) vs. 23.6% (13.2), p<0.001) and long-stay residents (22.2% (14.3) vs. 25.5% (15.0), p<0.001). LTCF characteristics may play an important role in preventing P&I hospitalizations. Hiring more Registered Nurses and Physician Extenders, increasing staffing hours, and reducing antipsychotic use may be modifiable means of reducing P&I in LTCFs. Funding provided by Sanofi Pasteur."}
+{"text": "LncRNA MIR4435-2HG is observed in a variety of cancers, while its role in colorectal cancer is unknown. We aimed to demonstrate the relationship between MIR4435-2HG and colorectal cancer based on The Cancer Genome Atlas (TCGA) database.Patients with colorectal cancer were collected from TCGA. We compared the expression of MIR4435-2HG in colorectal cancer and normal tissues with Wilcoxon rank sum test, and logistic regression was used to evaluate the relationship between MIR4435-2HG and clinicopathological characters. Moreover, Kaplan\u2013Meier and Cox regression was performed to evaluate the correlation between MIR4435-2HG and survival rate. Gene set enrichment analysis (GSEA) was also conducted to annotate biological function of MIR4435-2HG.P-value <0.05). High MIR4435-2HG expression had a poorer progression-free survival (p = 0.048), and overall survival (OS) (P = 0.028), which were validated in the GSE92921 and GSE29621 datasets. MIR4435-2HG expression ) was independently correlated with OS. GSEA demonstrated that the P38/MAPK pathway, the VEGF pathway, the cell adhesion molecules cams, the NOD-like receptor signaling pathway, the cell surface interactions at the vascular wall, and integrin cell surface interactions were differentially enriched in MIR4435-2HG high expression phenotype.MIR4435-2HG level was elevated in colorectal cancer tissues. Increased level of MIR4435-2HG was significantly correlated with TNM stage , stage (OR = 1.66 for stage 1/2 vs. stage 3/4), and carcinoembryonic antigen level before treatment (OR = 1.70 for <5 vs. \u22655) (all Increased MIR4435-2HG might be a potential biomarker for the diagnosis and prognosis of colorectal cancer. Moreover, MIR4435-2HG might participate in the development of colorectal cancer via the P38/MAPK and VEGF pathway. The prevalence of colorectal cancer is increasing worldwide . SurgeryMIR4435-2HG, also known as AGD2, MORRBID, LINC00978, MIR4435-1HG, lncRNA-AWPPH, has been regarded as a new oncogenic lncRNA in many types of cancer, like breast cancer and bladder cancer . A recenTherefore, we aimed to demonstrate the correlation between MIR4435-2HG and colorectal cancer, and analyze the prognostic role of MIR4435-2HG in colorectal cancer based on The Cancer Genome Atlas (TCGA). To achieve this goal, we analyzed the expression level of LncRNA MIR4435-2HG in colorectal cancer and normal tissues based on TCGA. And the correlation between LncRNA MIR4435-2HG and prognostic values was performed. Furthermore, MIR4435-2HG related biological pathways involved in colorectal cancer were detected using Gene set enrichment analysis (GSEA).The present study showed increased MIR4435-2HG was related to poor prognosis of colorectal cancer, and P38/MAPK pathway, VEGF pathway, cell adhesion molecules cams, Nod like receptor signaling pathway, cell surface interactions at the vascular wall, and integrin cell surface interactions were associated with LncRNA MIR4435-2HG expression phenotype using GSEA.n = 9) and with overall survival (OS) less than 30 days (n = 42) were excluded. The related RNA-seq data and clinicopathological data were extracted. The expression of lncRNA MIR4435-2HG in colorectal adenocarcinoma were analyzed and compared with that in adjacent normal tissues. Considering the fact that the relationship between LncRNA MIR4435-2HG expression and the development of a tumor is independent of the follow-up days, we used 622 RNA-seq data for further association analysis, and 580 data for survival analysis. The characteristics of patients including gender, race, differentiation, venous or perineural invasion, preoperative pretreatment, TNM stage, and tumor location were recorded. Some of them were not available, which were treated as missing value. In addition, the Gene Expression Omnibus (GEO) database was used to validate the association between MIR4435-2HG expression and colorectal cancer outcome. Two raw gene expression datasets  were downloaded from GEO for further study.In the TCGA database, 459 patients with colonic adenocarcinoma and 172 cases with rectal adenocarcinoma were collected. Finally, 580 cases were included in the further analysis, while cases without RNA-seq data .In this study, GSEA was used to elucidate the significant survival difference between high- and low- MIR4435-2HG groups. The number of gene set permutations were 1,000 times for each analysis. The expression level of lncRNA MIR4435-2HG was used as a phenotype label. The pathways enrichment was analyzed based on nominal Statistical analysis was performed using R (v.3.5.1) . Comparin = 240), and 156 cases (31.52%) had history of colon polyps. Stage I disease was found in 102 patients (18.21%), stage II in 206 (36.79%), stage III in 168 (30.00%), and stage IV in 84 (15.00%). Most tumors  were of colonic adenocarcinoma, and 27.07% (n = 157) were rectal adenocarcinoma. The topography distribution included 3.29% T1 (n = 19), 17.82% T2 (n = 103), 68.17% T3 (n = 394), and 10.73% T4 (n = 62). A total of 123 cases (24.50%) had venous invasion, and 57 (26.03%) perineural invasion. A total of 83 cases (16.12%) had distant metastases.The characteristics of patients including gender, race, differentiation, venous or perineural invasion, preoperative pretreatment, TNM stage, and tumor location were collected, as shown in P < 0.001) (P < 0.001)  . In addi< 0.001) , indicatP < 0.001), lymph node metastasis (P < 0.001), stage (P < 0.001), and CEA level before treatment (P = 0.013), as shown in A total of 580 colorectal cancer samples with LncRNA MIR4435-2HG expression data were analyzed from TCGA. Increased expression of MIR4435-2HG was correlated significantly with the grade of topography distribution , stage , and CEA level before treatment   . Univari= 0.015) . These rP = 0.048) (P = 0.028)  was significantly poorer in patients with high LncRNA MIR4435-2HG expression than those with low LncRNA MIR4435-2HG expression (P = 0.044) based on GSE92921 (P = 0.021) based on GSE29621 ), older patients ), higher CEA level ), higher TNM stage ; N: P < 0.001, HR = 2.991 (95% CI [2.027\u20134.414]); M: P < 0.001, HR = 4.737 (95% CI [3.156\u20137.11])), higher disease stage ), and venous invasion )  was significantly poorer in patients with high LncRNA MIR4435-2HG expression than those with low LncRNA MIR4435-2HG expression (= 0.048) \u20133C, simi= 0.028) \u20133F. In aGSE92921 \u2013S1C, andGSE29621 \u2013S1F. We GSE29621 . High Ln3.728])) .P = 0.040, HR = 1.955 (95% CI [1.031\u20133.710])), age ), status ), lymph node metastasis ), and disease stage ) were independently correlated with OS in multivariate analysis  in enrichment of MSigDB Collection (c2.cp.v6.2.symbols.gmt).Gene set enrichment analysis was used to identify signaling pathways involved in colorectal cancer between low and high LncRNA MIR4435-2HG expression data sets, and demonstrated significant differences  The Kaplan\u2013Meier curve, (B) Number at risk, and (C) Number of censoring of DFS in colorectal cancer based on GSE92921 dataset. (D) The Kaplan\u2013Meier curve, (E) Number at risk, and (F) Number of censoring OS in colorectal cancer based on GSE29621 dataset. DFS: disease-free survival; OS: over survival.Click here for additional data file.10.7717/peerj.6683/supp-3Supplemental Information 3Click here for additional data file.10.7717/peerj.6683/supp-4Supplemental Information 4Click here for additional data file.10.7717/peerj.6683/supp-5Supplemental Information 5Click here for additional data file."}
+{"text": "Healthy People 2020 included objectives to improve health of LGBT persons.In recent decades, public health awareness of health disparities among lesbian, gay, bisexual, and transgender (LGBT) populations has increased ( BRFSS is an annual state-based, random-digit\u2013dialed telephone survey of noninstitutionalized U.S. adults aged \u226518 years, which collects information on health-related topics.The prevalence and 95% confidence intervals of demographic characteristics  and of engaging in the five health-related behaviors was estimated by sexual orientation status for men and women separately, and by transgender status. The health-related behaviorsIn 2016, among 86,185 men who answered the sexual orientation question, 92.7% reported being heterosexual, 2.2% reported being gay, and 1.5% reported being bisexual; among 114,842 women, 91.7% reported being heterosexual, 1.3% reported being lesbian, and 2.3% reported being bisexual . OverallCompared with heterosexual men, gay men had a lower prevalence of not currently smoking cigarettes (77.0% versus 81.4%) and moderate or no drinking (51.8% versus 60.8%), but had a higher prevalence of performing any leisure-time exercise (82.0% versus 77.9%); gay men also had a higher prevalence of having a normal body weight (40.3%) than did bisexual (29.8%) and heterosexual men (25.0%). The prevalence of not currently smoking cigarettes, moderate or no alcohol consumption, and getting \u22657 hours\u2019 sleep during a 24-hour period was higher among heterosexual women  than among lesbian  and bisexual women . Engaging in any leisure-time exercise was more prevalent among lesbian (80.6%) and bisexual women (77.6%) than among heterosexual women (73.8%); however, having a normal body weight was less prevalent among lesbian women (30.4%) than among heterosexual women (37.0%); the difference in prevalence between heterosexual women and bisexual women (35.8%) was not statistically significant. In addition, the prevalence of reporting zero or one health-related behavior was higher among lesbian (10.0%) and bisexual (10.7%) women than among heterosexual women (4.9%), and the prevalence of reporting all five health-related behaviors was lower among lesbian (5.4%) and bisexual (6.9%) women than among heterosexual women (10.6%) .Among 200,874 adults from the 25 states and Guam who answered the gender identity question, 98.3% reported being cisgender, 0.2% reported being male-to-female transgender, and 0.1% each reported being female-to-male transgender and transgender nonconforming . Being aThe prevalence of performing any leisure-time exercise was higher among cisgender adults (75.5%) than among male-to-female transgender adults (56.7%). More than three quarters (77.4%) of male-to-female transgender adults reported sleeping \u22657 hours during a 24-hour period compared with cisgender adults (65.0%), female-to-male transgender adults (58.9%), and transgender nonconforming adults (52.9%). In addition, male-to-female transgender adults had a lower prevalence of engaging in any two of five health-related behaviors (12.3%) than did cisgender adults (18.6%), but had a higher prevalence of engaging in any three of five health-related behaviors (47.2%) than did female-to-male transgender adults (28.2%) .The findings from this study support those of other studies showing that disparities in sociodemographic characteristics and health-related behaviors exist among the LGBT populations , the prevalence of not currently smoking cigarettes, moderate or no drinking, maintaining a normal body weight, performing any leisure-time physical activity, and sleeping \u22657 hours per day was lower among lesbian (5.4%) and bisexual women (6.9%). Male-to-female transgender adults had a lower prevalence of engaging in any two of five health-related behaviors (12.3%) than did cisgender adults (18.6%), but had a higher prevalence of engaging in any three of five health-related behaviors (47.2%) than did female-to-male transgender adults (28.2%).Implementation of targeted strategies to increase community-based health intervention programs and mass media campaigns to improve health-related behaviors of lesbian, gay, bisexual, and transgender adults are needed."}
+{"text": "Lavandula species, which continues to increase in a variety of industries. Lavandula pubescens might be a good alternative, as it exhibits strong antibacterial activity. In this study, the chemical composition of the essential oils from different organs  of L. pubescens was identified using gas chromatography-mass spectrometry. Furthermore, the antimicrobial activities of different solvent extracts  and different organ  extracts of L. pubescens were evaluated. Only the ethyl acetate extracts of L. pubescens exhibited antibacterial activity against all bacterial strains tested, including Staphylococcus haemolyticus, Escherichia coli (KF 918342), Aeromonas hydrophila (KCTC 12487), E. coli (ATCC 35150), Cronobacter sakazakii (ATCC 29544), and Aeromonas salmonicida (KACC 15136). In particular, the extracts exhibited significant activity against S. haemolyticus. Ethyl acetate extract of the leaf exhibited the best activity against all bacterial strains. This study provides valuable information on the chemical compositions in essential oils and antimicrobial properties of L. pubescens.The discovery of a new species exhibiting more effective antibacterial properties is necessary because of the demand on  Lavandula pubescens, also known as downy lavender, is an aromatic flowering plant belonging to the Lamiaceae family. Lavandula, consisting of approximately 39 known species, has been largely cultivated as ornamental plants for gardens and scenery use. This perennial plant is widely distributed in the Mediterranean, North Africa, Southwest Asia, western Iran, and eastern India [Lavandula angustifolia, Lavandula officinalis, Lavandula latifolia, and Lavandula vera. Lavandula pubescens is a newly discovered species of lavender, which occurs in the Mediterranean. In recent years, the plants have been extensively studied as resources for medicine and aromatic products and used largely for their medicinal potentials, because these plants contain a number of bioactive compounds that act against human and plant pathogens [rn India . The widathogens , as wellLavandula is widely studied because of its commercial use in the fragrance industry [Lavandula species is presently used in food manufacturing industries for flavoring. Lavandula essential oil contains monoterpenes and is used in soaps, shampoos, mouthwashes, and household cleaners [Lavandula essential oil has been used in the food industry [Presently, industry ,5. The pindustry . The esscleaners . Previoucleaners  as well cleaners ,10. Furtindustry ,12.Essential oils, also called volatile or ethereal oils, are defined as an aromatic hydrophobic liquid, including a broad diversity of volatile compounds, and are derived from plant materials, such as the roots, stem, flowers, leaves, buds, seeds, and fruits . LavendeLavandula has been widely studied [Lavandula essential oil have been reported [L. pubescens using gas chromatography-mass spectrometry (GC-MS) and the antibacterial activities of different plant parts of L. pubescens.The antimicrobial activity of  studied ,19,20. L studied ,22. Antireported ,24,25. TEthanol (EtOH), methanol (MeOH), hexane, and ethyl acetate (EA) were purchased from Samchun Pure Chemical, Pyeongtaek, Korea. Diethyl ether (DE) and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich, Yongin, Korea. Luria-Bertani (LB) broth and streptomycin sulfate was purchased from MB cell, Seoul, Korea.L. pubescens were collected from the green house of Chungnam National University , Staphylococcus haemolyticus (KCTC 3341), Aeromonas hydrophila (KCTC 12487), Escherichia coli (ATCC 35150), Cronobacter sakazakii (ATCC 29544), and Aeromonas salmonicida (KACC 15136). These strains were collected from the medicine department of Chungnam National University. The bacterial strains (Escherichia coli (KF 918342), Staphylococcus haemolyticus (KCTC 3341), Escherichia coli (ATCC 35150), Cronobacter sakazakii (ATCC 29544), and Aeromonas salmonicida (KACC 15136)) were cultured at 37 \u00b0C and the strain (Aeromonas hydrophila (KCTC 12487)) was cultured at 30 \u00b0C for 18\u201324 h in a 50 mL sample of LB broth prepared in a 250 mL conical flask medium placed on an orbital shaker. Optical density (OD) measurement was performed at 600 nm using a UV-1800 spectrophotometer . The culture flask was inoculated at 0.1 OD600  with freshly prepared LB medium under the same culture conditions. The mid log phase bacterial cultures were used for the antibacterial studies. Staphylococcus haemolyticus (KCTC 3341) was the only gram-positive bacteria used in this study; all others were gram-negative bacteria.Six bacterial strains were used in this study: Volatile compounds were extracted and analyzed using a previously reported GC-MS method . GC-MS a600 of the different overnight bacterial cultures was swabbed on a 25 mL LB agar plate. Whatman disk was then placed on the plates. A 30 \u00b5L sample of different solvent extracts of L. pubescens was added to the sterilized disk  and incubated overnight at 37 \u00b0C. For screening of the different plant organs, the powdered samples were dissolved in ethyl acetate, and the resulting extract was added to the Whatman disk. Streptomycin (250 \u00b5g/mL) was used as the standard antibacterial agent. The experiment was performed in triplicate.A 0.1 ODL. pubescens extracts was established according to the method of Abdullah Al-Dhabi et al. [8 CFU/mL of each of the six bacterial strains was added to the 96-well plate and incubated at 37 \u00b0C for 17 h. After the incubation time, the minimum inhibitor concentration was determined by the lowest visible growth in LB broth. The experiment was performed in triplicate.The minimum inhibitory concentration (MIC) of the i et al.  by usingL. pubescens, such as the roots, stems, leaves, and flowers, were identified by GC-MS analysis. Most of the essential oils from different organs were characterized by the dominant presence of terpenes, including monoterpenes, diterpenes, and sesquiterpenes (p-cymenene (18.15%), terpinolene (3.31%), isothymol methyl ether (2.8%), m-cymene (2.1%), \u03b3-terpinene (1.64%), carvacrol (0.78%), 2,5,5-trimethyl-1,3,6-heptatriene (0.76%), and trans-3-caren-2-ol (0.76%); nine sesquiterpenes\u2014alloaromadendrene (2.45%), caryophyllene (2.45%), \u03b2-bisabolene (0.96%), \u03b1-muurolene (0.17%), (\u2212)-\u03b2-elemene (0.16%), \u03b2-ylangene (0.13%), \u03b2-guaiene (0.11%), \u03b1-calacorene (0.11%), and acoradien (0.07%); and one diterpene\u2014jolkinol D (0.16%), comprising 94.62% of the total leaf-derived essential oil.Chemical composition of the essential oils of different organs of terpenes . The oilp-cymenene (7.66%), m-cymene (2.89%), carvacrol (2.54%), trans-3-caren-2-ol (2.51%), 2,5,5-trimethyl-1,3,6-heptatriene (1.83%), and 4-terpinyl acetate (1.34%); and 10 sesquiterpenes\u2014caryophyllene (9.97%), \u03b2-bisabolene (3.52%), \u03b2-ylangene (1.46%), longifolene (1.04%), \u03b1-cubebene (0.65%), \u03b2-eudesmene (0.58%), \u03b1-ylangene (0.38%), caryophyllene oxide (0.33%), aromandendrene (0.11%), and acoradien (0.08%), comprising 99.47% of the total stem-derived essential oil.The essential oil from the stems contained 10 monoterpenes\u2014neo-allo-ocimene (15.68%), isothymol methyl ether (13.35%), \u03b3-terpinene (11.09%), terpinolene (10.17%), m-cymene (49%), \u03b1-terpinene (4.22%), 4-terpinyl acetate (3.73%), (\u2212)-4-terpineol (3.05%), p-xylene (2.46%), \u03b3-terpinene (2.28%), d-limonene (2.0%), p-cymenene (1.99%), terpinolene (1.90%), \u03b1-terpinene (1.57%), carvacrol (0.41%) neo-allo-ocimene (0.36%), trans-3-caren-2-ol (0.33%), and isothymol methyl ether (0.35%); and 21 sesquiterpenes\u2014(\u2212)-calamenene (6.33%), \u03b1-gurjunene (3.30%), \u03b1-copaene (1.54%), \u03b2-eudesmene (1.21%), \u03b1-bergamotene (1.00%), caryophyllene (0.91%), \u03b4-cadinene (0.88%), \u03b1-muurolene (0.86%), cyclosativene (0.81%), (\u2212)-\u03b2-elemene (0.66%), \u03b2-copaene (0.61%), \u03b3-cadinene (0.48%), \u03b1-calacorene (0.48%), \u03b1-cubebene (0.43%), \u03b2-guaiene (0.42%), \u03b1-ylangene (0.41%), acoradien (0.38%), caryophyllene oxide (0.19%), alloaromadendrene (0.16%), (+)-ledene (0.14%), and aromandendrene (0.07%), comprising 99.03% of the total root-derived essential oil.The essential oil from the roots contained 14 monoterpenes\u2014p-cymenene (11.66%), neo-allo-ocimene (8.55%), 1,3,5,5-tetramethyl-1,3-cyclohexadiene (2.95%), 2,5,5-trimethyl-1,3,6-heptatriene (2.75%), \u03b3-terpinene (2.39%), m-cymene (1.31%), trans-3-caren-2-ol (1.12%), \u03b1-terpinene (0.87%), carvacrol (0.27%), p-xylene (0.15%), and d-limonene (0.08%); and 5 sesquiterpenes\u2014longifolene (2.2%), \u03b2-bisabolene (0.72%), \u03b1-bergamotene (0.36%), caryophyllene (0.23%), and acoradien (0.07%), comprising 99.44% of the total flower-derived oil.The essential oil from flowers contained 13 monoterpenes\u20144-terpinyl acetate (16.99%), L. pubescens was performed using five different organic solvents. The ethyl acetate extract showed more antimicrobial activity against the six pathogenic bacteria because the disk diffusion method revealed that the ethyl acetate extract produced higher activity than that by the other organic solvents. In particular, L. pubescens possessed significant activity against S. haemolyticus, followed by E. coli (KF 918342), A. hydrophila (KCTC 12487), E. coli (ATCC 35150), C. sakazakii (ATCC 29544), and A. salmonicida (KACC 15136)  . TherefoLavandula extract was studied by the micro broth dilution method and the results are shown in Lavandula ethyl acetate extract at different concentrations inhibited all the growth of all bacterial strain broths. The MIC for E. coli was 6.25 \u00b5L and for S. haemolyticus, A. hydrophila, and A. salmonicida was 12.5 \u00b5L. Cronobacter sakazakii inhibition was observed at 25 \u00b5L. Streptomycin showed better MIC values in comparison with those of the Lavandula ethyl acetate extracts. MIC values are shown in The MIC of crude L. pubescens through GC-MS analysis. The most abundant constituents were terpenes, including monoterpenes, diterpenes, and sesquiterpenes. Similarly, previous studies have shown that terpene metabolites were the major components of essential oils from a variety of Lavandula species: Barocelli et al. reported a total of 23 constituents in Lavandula hybrida Reverchon \u201cGrosso\u201d essential oil, with major constituents such as linalool (33.4%), linalyl acetate (36.2%), camphor (7.6%), and 1,8-cineole (5.8%) [Lavandula stoechas L. (Spanish lavender) [L. angustifolia essential oil were 1,8-cineole (44.9%), camphor (14.3%), \u03b2-phellandrene (5.0%), and \u03b1-pinene (4.7%) [Lavandula bipinnata essential oil had transcarveol (18.93%), pulegone (8.45%), camphor (7.09%), and menthol (5.89%) [L. latifolia Med.) and lavandin (Lavandula \u00d7 intermedia) essential oils contained camphor (32.70%), 1,8-cineole (26.9%), and caryophyllene (4.88%) and 1,8-cineol (40.5%), linalool (33.1%), and camphor (8.17%), respectively [L. pubescens, with main components including carvacrol (72.7%), carvacrol methyl ether (7.0%), and caryophyllene oxide (5.9%) [In this study, 71 volatile compounds were identified in the essential oils derived from the roots, stems, leaves, and flowers of e (5.8%) . Fenchonavender) . The maje (4.7%) . Shirugu (5.89%) . Spike llinalool .4%, linae (5.9%) .L. pubescens tested in the present study showed strong antibacterial activity against the six bacterial strains tested. However, antibacterial activities of the extracts dissolved in the other solvents were not observed. This could be caused by the difference in the chemical composition of these extracts. Often, variations in chemical composition may result from differences in the extraction solvents, season, and presence of secondary metabolites [Ethyl acetate extract of abolites . Previouabolites ,37,38.L. pubescens possessed strong antibacterial activity against the six pathogenic bacteria, and L. pubescens leaves had the most powerful antimicrobial capacity. Our results are supported by those of previous studies showing that the essential oil from Lavandula species had antibacterial properties [lavender essential oil showed anti-bacterial activities against E. coli and Staphylococcus aureus [L. angustifolia was effective against all tested turtle-borne pathogenic bacteria: A. hydrophila, Aeromonas caviae, and Aeromonas dhakensis [C. sakazakii [L. angustifolia showed anti-bacterial activity against S. aureus, E. coli, Citrobacter freundii, Enterobacter aerogenes, Propionibacterium acnes, Proteus vulgaris, Pseudomonas aeruginosa, Shigella sonnei, and Streptococcus pyogenes [Lavandula plants are expected to possess important antibacterial properties against various bacterial species.Ethyl acetate extracts of operties . Hui et s aureus . Hossainhakensis . Furtherakazakii . The esspyogenes . TherefoL. pubescens and the different antibacterial activity of ethyl acetate extracts of these plant parts. These properties could be successfully exploited to treat several diseases caused by bacterial infections. This may indicate that lavender species used in traditional remedies may possess beneficial biological activity. Thus, this study suggests the potential use of L. pubescens roots, stems, leaves, and flowers in traditional herbal medicine applications.According to the World Health Organization (WHO), approximately 80% of the world\u2019s population depends on herbal remedies for their primary healthcare ,45. ThisL. pubescens, and its extract possessed strong antibacterial activity against Escherichia coli (KF 918342), Staphylococcus haemolyticus (KCTC 3341), Aeromonas hydrophila (KCTC 12487), Escherichia coli (ATCC 35150), Cronobacter sakazakii (ATCC 29544), and Aeromonas salmonicida (KACC 15136). In particular, the leaves exhibited the strongest antimicrobial activity against these bacteria among the different plant parts tested. Therefore, this study suggests that L. pubescens could be considered a good source for human health.In this study, ethyl acetate was the most effective solvent for extracting a broad variety of compounds from"}
+{"text": "Their structures were elucidated by extensive NMR spectroscopy studies. Compound 2 represents the first example of an R-configuration in the prenylated moiety. All these isolated compounds were examined for antimicrobial and cytotoxic activities. Compounds 1\u20139 exhibited antimicrobial activities against some human- and plant-pathogenic microbes with MIC values ranging from 2 to 64 \u03bcg/mL. Moreover, compound 9 displayed considerable inhibitory activity against the THP-1 cell line in vitro, with an IC50 value of 7.0 \u03bcg/mL.Considerable attention has been paid to marine derived endophytic fungi, owing to their capacity to produce novel secondary metabolites with potent bioactivities. In this study, two new compounds with a prenylated diphenyl ether structure\u2014diorcinol L ( Aspergillus sp. + and 351.12 [M + Na]+; HRESIMS at m/z 329.1377 [M + H]+  and 351.1199 [M + Na]+ .(R)-Diorcinol B (2): yellowish powder; c 0.1, MeOH); UV (MeOH) \u03bbmax (log \u03b5) 205 (4.44), 281 (3.30) nm; 1H and 13C-NMR data, see m/z 333.16 [M + H]+; HRESIMS m/z 333.1689 [M + H]+ .Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, Ralstonia solanacearum and plant pathogenic fungi Alternaria alternata, Cochliobolus heterostrophus, Gaeumannomyces graminis, Glomerella cingulata, Mucor hiemalis, and Thielaviopsis basicola were carried out using the well diffusion method [Antimicrobial assays against the human- and plant-pathogenic bacteria n method . ChloramThe cytotoxic activity of the isolated compounds against six tumor cell lines including A549 , Du145 (human prostate cancer cell line), HeLa (human cervix carcinoma cell line), MCF-7 (human breast adenocarcinoma cell line), MDA-MB-231 (human breast cancer cell line), and THP-1 (human monocytic cell line) were determined according to the previously reported the Cell Counting Kit-8 (CCK-8) colorimetric method .Aspergillus tennesseensis. Among them, compound 2 represents the first example of an R-configuration in the prenylated moiety. The antimicrobial and cytotoxic activities of the isolated compounds were evaluated, and some of the compounds showed promising activities. These results indicated that the algal-derived endophytic fungi are a prolific resource of novel bioactive natural products.In conclusion, nine prenylated diphenyl ethers, including two new ones, were isolated and identified from the marine algal-derived endophytic fungus"}
+{"text": "We performed a meta-analysis to obtain the diagnostic and grading accuracy of 18F-FDOPA PET and PET/CT in patients with gliomas.Positron emission tomography (PET) and PET/computed tomography (PET/CT) imaging with 3,4-dihydroxy-6-[18F-FDOPA PET or PET/CT in glioma patients. Pooled sensitivity, specificity, and area under the summary receiver operating characteristic (SROC) curve were calculated from eligible studies on a per-lesion basis.PubMed, Embase, Cochrane Library and Web of Science were searched through 13 May 2019. We included studies reporting the diagnostic performance of 18F-FDOPA PET and PET/CT were 0.90 (95%CI: 0.86\u20130.93) and 0.75 (95%CI: 0.65\u20130.83). Across 7 studies (219 patients) for glioma grading, 18F-FDOPA PET and PET/CT showed a pooled sensitivity of 0.88 (95%CI: 0.81\u20130.93) and a pooled specificity of 0.73 (95%CI: 0.64\u20130.81).Eventually, 19 studies were included. Across 13 studies (370 patients) for glioma diagnosis, the pooled sensitivity and specificity of 18F-FDOPA PET and PET/CT demonstrated good performance for diagnosing gliomas and differentiating high-grade gliomas (HGGs) from low-grade gliomas (LGGs). Further studies implementing standardized PET protocols and investigating the grading parameters are needed. Glioma is the most common primary brain tumor, accounting for 81% of all malignant brain tumors with an annual incidence of 5.26 per 100,000 individuals , 4. Thus18F-FDG) has been the classic PET tracer used in tumor imaging, but it is not ideal in detecting gliomas due to the high physiologic uptake in normal brain tissue. Given the diagnostic limitations of 18F-FDG, amino acid tracers have been extensively investigated, such as 3,4-dihydroxy-6-[18F] fluoro-L-phenylalanine (18F-FDOPA). Unlike gadolinium, 18F-FDOPA is transported across the intact blood brain barrier (BBB) , indicating a moderately high overall diagnostic value of als Fig. c.p-value\u2009=\u20090.0006, I-square\u2009=\u200965.1%) through Chi-square, Cochran-Q, and I-squared tests , specificity of 0.76 (95% CI: 0.66\u20130.85), LR+ of 2.87 (95% CI: 2.01\u20134.11), LR- of 0.13 (95% CI: 0.07\u20130.23), DOR of 29.65 (95% CI: 13.09\u201367.15), and AUC of 0.90 (SE\u2009=\u20090.06). In the newly-diagnosed subgroup, only two studies were eligible for meta-analysis (in another three studies only sensitivity can be computed after stratification). In comparison with recurrent group, pooled data of newly-diagnosed group revealed a lower sensitivity of 0.71 (95% CI: 0.54\u20130.85) and a higher specificity of 0.86 (95% CI: 0.42\u20131.00). The LR+, LR- and DOR were 3.71 (0.87\u201315.81), 0.36 (0.19\u20130.68) and 10.88 (1.57\u201375.31) respectively.No threshold effect was detected in two subgroups. Pooled data and SROC curve manifested a high diagnostic value of 18F-FDOPA PET and PET/CT for Grading Gliomas.Aim 2: Investigating the Performance of 18F-FDOPA PET and PET/CT presented a pooled sensitivity of 0.88 (95% CI: 0.81\u20130.93), specificity of 0.73 (95% CI: 0.64\u20130.81), LR+ of 2.90 (95% CI: 2.19\u20133.85), LR- of 0.16 (95% CI: 0.08\u20130.36), DOR of 25.87 (95% CI: 10.53\u201363.54), and AUC of 0.89 (SE\u2009=\u20090.03) for differentiating HGGs from LGGs.As illustrated in the Fig. p-value\u2009=\u20090.0045), no significant heterogeneity was found among studies for specificity , LR+ , LR-  and DOR .Other than sensitivity , and for HGGs  and LGGs  respectively. Besides the evidence presented above, our systematical analysis has also shown that 18F-FDOPA PET and PET/CT provide more reliable results in discriminating recurrent lesions from treatment related changes, with a pooled sensitivity and specificity of 0.92 and 0.76.Of increased interest is the value of patients . Howevera et al.  reportedi et al.  proved s18F-FDOPA PET and PET/CT is the utility in the detection of HGGs. Due to the differences between LGGs and HGGs in therapeutic regimen and prognosis evaluation, initial grading for gliomas through imaging approaches is of great significance. This meta-analysis proved the high overall value of 18F-FDOPA PET and PET/CT in evaluating tumor grade, with the AUC of 0.89. However, studies with respect to grading of recurrent gliomas demonstrated controversial results. In Fueger et al. -methyl-L-methionine (11C-MET). In the only study with direct comparison between 18F-FDOPA PET and 11C-MET, 18F-FDOPA performed equally well as 11C-MET in the imaging of brain tumors [11C-MET is being replaced by tracers labelled with fluorine-18  due to its short half-life (20\u2009min). The use of another amino acid tracer O-(2-[18F]-fluorethyl)-1-tyrosine (18F-FET) has rapidly grown in recent years. A previous meta-analysis of 5 studies including 180 patients reported a sensitivity and specificity of 0.82 and 0.76 of 18F-FET PET for the differential diagnosis of primary brain tumor [18F-DOPA and 18F-FET shared similar uptake pattern in primary and recurrent HGGs and had similar diagnostic accuracy in recurrent HGGs [18F-FET, the elevated physiological 18F-FDOPA uptake in striatum has been a concern which may limits its use to detect brain tumor with striatum involvement [18F-FET to examine patients with possible involvement of basal ganglia irrespective of tumor grade. Similarly, Morana et al. [18F-FDOPA PET/CT in dorsal striatum was concordant with its overall accuracy, while not in the ventral striatum, which required fused MRI to improve its performance.Evidence for comparisons of c tissue . 18F-FDO 18F-FDG , 10, 23.27\u20130.50) . The lon27\u20130.50) C-methyl-in tumor . In lateent HGGs , 41. Howolvement , 43. Thea et al.  reported18F-FDOPA uptake with glioma molecular characteristics cannot be analyzed.Several study limitations also need to be addressed. Although 19 studies were included in this meta-analysis, the sample size of each study tended to be small, and the number of true negative cases was particularly limited, which might possibly yield less replicable results, especially the pooled specificity. Limited sample size also led to data loss in stratification of subgroups when true negative case was less than one . Besides, multiple specimens were obtained from one patient in three studies , 27, 30,18F-FDOPA PET and PET/CT have high diagnostic accuracy for the detection of gliomas, especially the discrimination between tumor recurrence and radiation-induced changes. 18F-FDOPA PET and PET/CT also demonstrate good grading performance for the distinction between HGGs and LGGs. However, the grading parameters needs further investigation with standardized imaging protocols in prospective studies.In conclusion, this meta-analysis provides evidence that"}
+{"text": "Brassica napus) is preferred over black-seeded rapeseed for the desirable properties of the former. This study evaluated the metabolites and nutritive values of black-seeded rapeseed meal and yellow-seeded meal from the progeny of a B. napus\u2013Sinapis alba hybrid. Yellow-seed meal presented higher protein (35.46% vs. 30.29%), higher sucrose (7.85% vs. 7.29%), less dietary fiber (26.19% vs. 34.63%) and crude fiber (4.56% vs. 8.86%), and less glucosinolates (22.18 vs. 28.19 \u03bcmol/g) than black-seeded one. Amounts of ash (3.65% vs. 4.55%), phytic acid (4.98% vs. 5.60%), and total polyphenols (2.67% vs. 2.82%) were decreased slightly in yellow-seeded meal compared with black-seeded meal. Yellow-seeded meal contained more essential amino acids than black-seeded meal. Levels of the mineral elements Fe, Mn, and Zn in yellow-seeded meal were higher than black-seeded meal. By contrast, levels of P, Ca, and Mg were lower in yellow-seeded meal. Moreover, yellow-seeded meal showed lower flavonol  content than black-seeded meal. Comparison of metabolites between yellow and black rapeseed confirmed the improved nutritional value of meal from yellow-seeded B. napus, and this would be helpful to the breeding and improvement of rapeseed for animal feeding.Breeding of yellow-seeded rapeseed ( Brassica napus) is the second most abundant oil crop worldwide next to soybean. Soybean meal is broadly used as an animal fodder for its abundant nutrients, but the supply of soybean meal is insufficient to meet current demands [B. napus [B. napus is an optimal perspective in breeding of rapeseed, and cultivars/lines with yellow seed coat have been selected from hybridization among Brassica species with yellow seed character  [Sinapis alba) have been reported [B. napus has revealed different nutrient contents in the seed meals [et al., found higher protein , higher sucrose (10.2% vs. 8.8% DM), and less total dietary fiber (24.1% vs. 30.1% DM) in meal derived from yellow-seeded B. napus than black seed [et al., identified 31 polyphenols in canola meal, of which sinapine is the most abundant chemical, comprising 80% of the total phenolics [et al., reported that the unpleasant taste of meal with higher sinapine content affected the intake of pigs, which directly reduced their average daily feed intake and average daily weight gain [et al., reported significant differences in the PA contents of yellow- and black-seeded B. napus [Rapeseed  . Introdureported . Comparied meals ,9,10. Beed meals . Slominsack seed . Antinuthenolics . Sinapinght gain ,15. Flavght gain . Jiang eB. napus .B. napus\u2013S. alba hybrids, and the oil content of this yellow-seeded rapeseed line (W82) was 6% higher than that of black-seeded rapeseed [B. napus cv. Yangyou 6) and yellow-seeded rapeseed . This comprehensive comparison will facilitate understanding of the differences in nutritional values between the two rapeseed lines and might contribute to the utilization and molecular dissection of yellow-seeded rapeseed.Rapeseed with yellow-seeded germplasm does not naturally exist. We obtained a novel yellow-seeded rapeseed from the progeny of rapeseed . In the B. napus were produced using a high-temperature press, degreasing with petroleum ether. The chemical compositions of seed meals from yellow- and black-seeded rapeseeds are listed in B. napus presented higher protein (35.46% vs. 30.29% DM) and higher sucrose (7.85% vs. 7.29% DM). Significant differences in protein and sucrose levels were identified between yellow- and black-seeded B. napus seed meals (p < 0.05). Proteins are the main nutrients in rapeseed meal, supplying energy for metabolism after animal absorption. Improved protein content in yellow-seeded rapeseed meal indicates higher nutritional value for animal feeding. Theodoridou et al., reported easier animal assimilation of yellow-seeded rapeseed meal from B. napus than that of black-seeded rapeseed meal [B. juncea and B. napus exhibited higher absorption efficiency of proteins in the rumen and intestines than when they were fed with meal from black-seeded B. napus. Higher intestinal digestibility of total metabolizable protein and lower degraded protein balance were reported when yellow-seeded rapeseed meal was consumed compared with black-seeded rapeseed meal. Thus, the increase in protein content of the novel yellow-seeded B. napus derived from somatic hybrids of B. napus\u2013S. alba indicates improved nutritional value for animal feeding, especially in the actual absorbed protein. High sucrose levels would improve the digestibility of rapeseed meal because sucrose is a carbohydrate that can be easily digested. Theander et al., reported the sucrose content in seed meal of Brassica to be approximately 6.2%\u20138.0% [Rapeseed meals from yellow- and black-seeded .2%\u20138.0% ,17.vs. 34.63% DM), crude fiber (4.56% vs. 8.86% DM), and glucosinolates (22.18 \u03bcmol/g DM vs. 28.19 \u03bcmol/g DM) were significantly higher in black-seeded rapeseed meal than in yellow-seeded rapeseed meal  and total polyphenol (2.67% vs. 2.82% DM) contents were lower in yellow-seeded rapeseed meal than in black-seeded rapeseed meal , methionine and cysteine (1.36 mg/g vs. 0.21 mg/g), isoleucine (0.33 mg/g vs. 0.20 mg/g), glycine (0.62 mg/g vs. 0.39 mg/g), alanine (2.95 mg/g vs. 1.06 mg/g), phenylalanine (2.08 mg/g vs. 1.26 mg/g), histidine (0.98 mg/g vs. 0.50 mg/g), lysine (1.65 mg/g vs. 0.80 mg/g), and arginine (1.59 mg/g vs. 1.17 mg/g) (p < 0.05) (vs. 7.06 mg/g), lysine (9.52 mg/g vs. 9.02 mg/g), threonine (9.37 mg/g vs. 8.29 mg/g), isoleucine (13.39 mg/g vs. 11.97 mg/g), asparagine (12.91 mg/g vs. 11.71 mg/g), glutamic acid (23.75 mg/g vs. 21.18 mg/g), methionine and cysteine (32.15 mg/g vs. 29.01 mg/g), tyrosine (15.44 mg/g vs. 13.80 mg/g), glycine (2.28 mg/g vs. 1.83 mg/g), alanine (6.99 mg/g vs. 6.04 mg/g), histidine (6.73 mg/g vs. 5.88 mg/g), and arginine (12.13 mg/g vs. 10.44 mg/g) were significantly higher in yellow-seeded rapeseed meal than in black-seeded rapeseed meal  directly affects the nutritional value of seed meal [et al., showed that the digestibility of total amino acids was higher in chickens fed with yellow-seeded rapeseed meal than in those fed with black-seeded rapeseed meal [Profiles of essential amino acids and total amino acids are listed in  < 0.05) . In termeed meal . Methioneed meal .vs. 87.48 mg/kg), Mn (48.12 mg/kg vs. 37.63 mg/kg), Zn (74.89 mg/kg vs. 54.82 mg/kg), Cu (4.96 mg/kg vs. 4.67 mg/kg), and K , and the differences of Fe, Mn, and Zn contents between the two types of rapeseed meal were significant (p < 0.05). By contrast, the amounts of P , Ca (3990.40 mg/kg vs. 5645.36 mg/kg), and Mg (3767.20 vs. 3860.59 mg/kg) were lower in yellow-seeded rapeseed meal than in black-seeded rapeseed meal  derived from the progeny of B. napus\u2013S. alba hybrids and black-seeded rapeseed (B. napus cv. \u201cYangyou 6\u201d) were used in this study [et al. [Seeds of yellow-seeded is study ,36. \u201cYan [et al. , seed meet al. [2SO4 (5 mL) in 30 g/L trichloroacetic acid (5 mL). After shaking for 2 h at room temperature, the suspension was collected after centrifugation at 13,000 g for 10 min. The pellet was re-extracted and all the suspension was combined. The phytic acid was precipitated with 0.3 g/L FeCl3 (2 mL), and its amount was calculated according to the difference of phosphorus value between the initial supernatant and the supernatant after precipitated with FeCl3. A conversion factor of 3.55 from phosphorus and phytic acid was used. Glucosinolates were determined via near-infrared spectroscopy, which developed a new calibration to measure major fraction, and no external standards were needed [et al. [et al. [For determination of protein contents, a nitrogen analyzer was used to obtain the content of N, and the content of crude protein was calculated by N \u00d7 6.25. Seed meal (0.5 g) was treated with 6 N HCl (20 mL) at 110 \u00b0C for 24 h, and total amino acid content was then determined using ninhydrin colorimetry and an Biochrom 30 amino acid analyzer . For free amino acid analysis, the seed meal (0.5 g) was extracted with picric acid (20 mL) then determined on the amino acid analyzer . Total det al. . Seed me [et al. . Inducti [et al. . Calibraet al. with some modifications, and the extracts were filtered through 0.45 \u03bcm Teflon membranes  and then used for high-performance liquid chromatography (HPLC)-electrospray ionization (ESI)/tandem mass spectrometry (MS2) analysis [et al. [2CO3 (50 \u03bcL) was added. The total phenol content was reported as (\u2212)-epicatechin equivalents based on a calibration curve. Using (\u2212)-epicatechin as standard, the absorbance was measured at 725 nm after the incubation for 45 min to acknowledge the content of phenol content. The total phenol content was reported as (\u2212)-epicatechin equivalents based on a calibration curve. HPLC analysis of 10 \u03bcL extracts was performed on an Agilent 6460 instrument  equipped with automatic injector, quaternary pump, thermostatted column compartment, and diode array detector (DAD). The Agilent Masshunter Qualitative Analysis software was used for data collection and analysis. LC/MS data of tentatively identified polyphenolics in meal of black- and yellow-seeded B. napus and their quantitative analysis using DAD at 280 and 330 nm. Samples were separated using XB-C18 column , with a flow rate of 0.3 mL/min, oven temperature of 30 \u00b0C, and a detection range of 190\u2013800 nm. The mobile phase consisted of a combination of solvent A  and solvent B . The linear gradient was as follows: 10% B (v/v) for 5 min, 10%\u201390% B for 40 min, 90% B for 10 min, 90%\u201310% B for 1 min, and hold at 10% B for 10 min.Phenolics were extracted using the method previously described by Shao analysis . Seed me [et al. : extract2) pressure = 15 psi, sheath gas temperature 250 \u00b0C, sheath gas flow = 7 L/min, capillary voltage = 4000 V, and spraying voltage = 500 V. Full scan spectra of 500 ms and an m/z range of 90\u20132000 were selected. MS2 fragmentation patterns of phenolics were determined under 15\u201340 V, using the Agilent MassHunter Workstation Data Acquisition and Agilent MassHunter Qualitative Analysis software. Procyanidin B2, (\u2212)-epicatechin, sinapine, and quercetin-3-O-\u03b2-d-glucoside, which were purchased from Sigma , and isorhamnetin-3-O-glucoside, which was purchased from ChromaDex , were used as standards for phenolic quantification. Since not all the standards of the chemicals identified in the O-\u03b2-d-glucoside were adequate for quantification of all the chemicals, of which quercetin-3-O-\u03b2-d-glucoside was used as standard for quercetin, isorhamnetin and kaempferol. Statistical analyses of the contents were expressed as mean \u00b1 standard error of samples from three independent extractions. A significance level of p < 0.05 was considered.Tandem mass spectrometry was operated in positive and negative ESI modes. The mass spectrometer used in phenolic analysis was MS QQQ coupled in LC/MC system . The parameters were as follows: drying gas temperature = 300 \u00b0C, drying gas flow = 10 L/min, nebulizer gas (NB. napus improves the quality of rapeseed meal. In this study, a novel yellow-seeded line of B. napus derived from a somatic B. napus\u2013S. alba hybrid was shown to be of greater nutritional value compared with black-seeded rapeseed, that is, yellow-seeded rapeseed meal exhibited more nutrient substances and less antinutrients than black-seeded rapeseed meal. The chemical composition of rapeseed meal, in which proteins are the resources for animal feeding, but the antinutrients influence the actual digestibility and assimilation of the nutrients in rapeseed meal is rather complicated. Thus, further animal feeding experiments using seed meals from both yellow- and black-seeded rapeseeds are necessary to evaluate the actual value of yellow-seeded rapeseed meal.Rapeseed meal is abundant in protein compared with other seed meals, making it optimal for animal feeding. However, antinutrients in rapeseed meal limit its application. Breeding of yellow-seeded"}
+{"text": "Correction to: BMC Cancer (2018) 18:325https://doi.org/10.1186/s12885-018-4261-5Following publication of the original article , the autAdditional file 1:Table S1. Sequences of the qPCR primers used in this study. Table S2. Cell numbers per well used in this study for cell proliferation, cell cycle and immunofluorescent assays. Table S3. KPNA7 silencing efficiencies of the cell lines used in Fig. 1 24\u2009h after siRNA treatment. (DOCX 17 kb)"}
+{"text": "Correction to: BMC Genomics (2017) 18:693https://doi.org/10.1186/s12864-017-3951-8Following the publication of this article , the autFurthermore, in the \u2018Availability of data and materials\u2019 declaration the sentence \u201cSupplementary material can be found online at http://www.rna.uni-jena.de/en/supplements/nothobranchius-furzeri-mirnome/\u201c\u00a0should now read \u201cSupplementary material can be found online at https://osf.io/25mxb/ (DOI 10.17605/OSF.IO/25MXB).\u201d"}
+{"text": "AbstractScolopsislacrimasp. nov. is described from a single specimen (213.6 mm standard length) collected from Grande-Terre Island, New Caledonia. The new species closely resembles S.meridiana, both species having the upper part of the pectoral-fin base with reddish blotch when fresh, two bands across the top of the snout, a dorsal scaled area on the head reaching anteriorly to between the anterior margin of the eye and anterior nostril, a similar number of lateral-line scales, and absence of a small antrorse spine below the eye. However, S.lacrimasp. nov. is distinguished from S.meridiana by having diagonal lines on the body absent , a dark longitudinal band below the lateral line , the caudal fin central area not patterned , a narrower body and shallower caudal peduncle.The new monocle bream  Scolopsis Cuvier, 1814 is widespread throughout shallow Indo-West Pacific tropical and subtropical waters, some species being marketed in Southeast Asia  recognised as a valid species by The monocle bream genus ast Asia , 2001. TScolopsis, a single specimen from New Caledonia, having a distinctively elongate body and unique colouration, was examined. It is described herein as a new species of Scolopsis.During a taxonomic study of DASMN). Examined specimens of S.meridiana and Scolopsistaenioptera  are listed in Counts and proportional measurements followed Taxon classificationAnimaliaSpariformesNemipteridaehttp://zoobank.org/5AF65765-E484-41A9-ABB2-E88A93034AE5Scolopsistaeniopterus (non Cuvier): Scolopsistaenioptera (non Cuvier): MNHN 2002\u20132930, 213.6 mm SL, Noum\u00e9a, Grande-Terre Island, New Caledonia, 1 Aug 2002, purchased at market by P. B\u00e9arez.Scolopsis with the following combination of characters: pectoral-fin rays 17; lateral-line scales 47; no antrorse spine below eye; dorsal scaled area on head reaching anteriorly to between anterior margin of eye and anterior nostril; bony opercular ridge and lower limb of preopercle without scales; 3rd anal-fin spine longer than 2nd anal-fin spine; narrow body, its depth at dorsal, pelvic, and anal fin origins 29.2, 29.5 and 26.6% of SL, respectively; caudal-peduncle depth 10.4% of SL; head length 29.9% of SL; upper part of pectoral-fin base with reddish blotch when fresh; two dark bands across dorsum of snout; body below lateral line with a dark longitudinal band, without diagonal lines; no blotches or lines on central area of caudal fin.A species of st dorsal-fin spine length 6.2; 2nd dorsal-fin spine length 8.5; 3rd dorsal-fin spine length 10.4; 4th dorsal-fin spine length 10.8; 5th dorsal-fin spine length 11.0; 6th dorsal-fin spine length 11.0; 7th dorsal-fin spine length 10.9; 8th dorsal-fin spine length 11.0; 9th dorsal-fin spine length 10.7; 10th dorsal-fin spine length 10.4; longest dorsal-fin soft ray length 16.0; 1st anal-fin spine length 3.9; 2nd anal-fin spine length 7.4; 3rd anal-fin spine length 8.0; anal-fin base length 15.0; pectoral-fin length 21.3; pelvic-fin spine length (measured on right side because the left side damaged) 13.8; longest pelvic-fin soft ray length 24.0.Dorsal-fin rays X, 9; anal-fin rays III, 7; pectoral-fin rays (left / right) 17 / 17; pored lateral-line scales 47; pelvic-fin rays I, 5; scale rows above lateral line 5; scale rows below lateral line 10; gill rakers (upper / lower) 5 / 8; preopercle scale rows (behind eye) 3; preopercle scale rows (below eye) 4. The following morphometrics are expressed as percentages of SL: body depth at dorsal-fin origin 29.2; body depth at pelvic-fin origin 29.5; body depth at anal-fin origin 26.6; body depth at posterior margin of orbit 23.7; body depth at anterior margin of orbit 17.5; pre-dorsal-fin length 32.7; pre-pelvic-fin length 37.9; pectoral-pelvic length 15.4; pre-anus length 61.4; head length 29.9; snout length 11.4; posterior nostril  0.8; posterior nostril  0.8; upper-jaw length 10.7; orbit diameter 7.8; interorbital width 10.0; suborbital depth 5.7; caudal-peduncle length 22.8; caudal-peduncle depth 10.4; dorsal-fin base length 54.1; 1st to 10th dorsal-fin spines, thereafter more steeply to caudal peduncle. Ventral profile of body lowering from lower-jaw tip to anus, thereafter rising to caudal peduncle. Dorsal-fin origin just above posteriormost point of opercle, base extending posterior to posteriormost point of anal-fin base. First to 5th dorsal-fin spines gradually lengthening, 5th to 8th spine lengths similar, 8th to 10th spines gradually shortening. Seventh dorsal-fin soft ray longest. All dorsal-fin soft rays non-filamentous. Uppermost point of pectoral-fin base slightly posterior to posteriormost point of opercle. Lowermost point of pectoral-fin base anterior to pelvic-fin origin. Posterior tip of pectoral fin pointed, reaching to vertical through 7th dorsal-fin spine origin. Pelvic-fin origin posterior to dorsal-fin origin. Posterior tip of depressed pelvic fin reaching anus, not reaching anal-fin origin. Anal-fin origin below 1st dorsal-fin ray origin, ending below 6th dorsal-fin ray origin. First anal-fin spine shortest, 3rd spine longest. Caudal-fin forked, upper lobe longer than lower lobe. Posterior tip of both lobes of caudal fin pointed, non-filamentous. Anus oblong, anterior to anal-fin origin. Eye and pupil round. Lower margin of eye above a line from snout tip to uppermost part of pectoral-fin base. Nostrils round, paired, positioned close together anterior to orbit, anterior nostril with small dermal flap. Snout pointed. Posterior tip of maxilla not reaching to vertical through anterior margin of eye. Distinct suborbital spine posteriorly directed. Small antrorse spine below eye absent. Posterior margins of suborbital and preopercle serrated. Scales ctenoid; both lips, snout, area around eye, and bony opercular ridge and lower limb of preopercle scaleless. Lateral line complete, originating above opercle, extending to central part of caudal-fin base. Both jaws with small conical teeth, forming dense bands. Canine teeth absent. Gill rakers long, slender.Body oblong, rather compressed, deepest at pelvic-fin origin. Dorsal profile rising from snout tip to dorsal-fin origin, lowering slightly between origins of 1Based on colour photograph of holotype  from New Caledonia. The latter two specimens have been reduced to bones and otoliths only. However, fresh colour photographs of both specimens prior to dissection (Fig. S.lacrima.ion Fig.  support Scolopsismeridiana and S.taenioptera are restricted to northern Australia and Southeast Asia, respectively (Fig. Scolopsis with a reddish blotch on the upper part of the pectoral-fin base are allopatrically distributed in the Indo-West Pacific.ely Fig. , probabl"}
+{"text": "Lactobacillus reuteri used prior to or concurrently with Staphylococcus aureus infection can increase epidermal keratinocyte survival, p < 0.01. Phenolics may not have been extensively studied for atopic dermatitis or skin burns. However, phenolics do have a role in photoprotection. The phenolic rutin increases ultraviolet B radiation filter reactive oxygen species scavenging at 75%, p < 0.002, and peak wavelength absorption, p < 0.001. While oral and topical probiotics have untapped potential for atopic dermatitis amelioration and skin infection prevention, phenolics will be increasingly used for photoprotection. With optimized bioavailability, dosage, and formulation, nutraceuticals will become crucial for healthy skin maintenance.Nutraceuticals are important for healthy skin maintenance. Probiotics, phenolics, and vitamins are just a few of the nutraceuticals meant to potentially prevent and assist medical management of dermatologic conditions. Among these, probiotics, vitamin E, and green tea catechins may offer the broadest array of skin protective mechanisms with probiotics having the greatest clinical range. Probiotics\u2019 amelioration of atopic dermatitis and opportunistic infections of skin burns has been targeted in recent research efforts. This includes the improvement of Scoring Atopic Dermatitis index scores,  Nonmelanoma skin cancers (NMSC), which are comprised primarily of basal cell carcinomas (BCC) and squamous cell carcinomas (SCC), are one of the most common human cancers with an 18-fold to 20-fold higher incidence than melanoma . MeanwhiUltraviolet A radiation (UVA), which has a 320 nm to 400 nm spectrum range, is the deepest penetrating UVR at 1000 \u03bcm into the epidermis and dermis and comprises 90% to 95% of UVR . UVA actKeratinocytes are the major cells of the epidermis, which is the outermost skin layer . The mosStaphylococcus aureus in 42% of AD patients [Bifidiobacterium (Acintobacteria)-based intestinal microbiome [Increased outer horny skin layer thickness is associated with atopic dermatitis  . About 1patients . Consistpatients ,13. In ccrobiome ,13.Although Hippocrates publicized foods\u2019 medicinal value, nutraceutical is a twentieth century term . Based oPropionibacterium and Corynebacterium, Firmicutes Staphylococcus, Proteobacteria and Bacteroidetes is achieved in childhood [S. aureus colonization with less than 106 cfu/cm2 of skin occurs in 20% of people and intermittent carriage occurs in 60% of people [S. aureus at greater than or equal to 106 cfu/cm2 of skin is considered an infection [S. aureus causes dose dependently severe opportunistic infections from 105 cfu/mL or greater [S. aureus infections with any skin barrier disruption, which occurs with burns, AD, and acne vulgaris [Long-term skin commensal microbiome stability dominated by the Actinobacteria hildhood . Skin cohildhood . Skin cohildhood . Stable f people ,18. NormLactobacillus plantarum have shown nonspecific immune system boosting and regulation [Probiotics are live, active, microbial cultures that are generally regarded as safe (GRAS) when consumed in quantities that achieve health benefits to the host ,9. Heat-gulation ,20. LactLactobacillus paracasei and Bifidobacterium longum reuter reportedly decreased leg dryness and facial roughness in 33 adult females over an eight-week trial [B. longum reuter lysate (disrupted bacteria) topically applied to reactive skin twice daily for two months achieved increased skin barrier resistance measured by reduced trans-epidermal water loss (TEWL) and reduced skin sensitivity and dryness in response to thermal and chemical physical changes [Historically, with oral formulations, probiotics had to survive gastric transport, adhere to, and colonize the gastrointestinal tract . These cek trial . Subsequ changes .2 = 61%, Absolute Risk Reduction 44 cases per 1000 children; 95% CI 20 to 64 [Lactobacillus rhamnosus Goldin and Gorbach (LGG), when not heat inactivated, significantly improves the Scoring Atopic Dermatitis index (SCORAD) scores in comparison to heat inactivated LGG or the placebo, p = 0.02 [p = 0.036 [9 cfu, oral Lactobacillus fermentum VRI-033 PCC, twice daily for 8-weeks achieved SCORAD improvements with sustained T-helper 1 IFN-\u03b3 responses, p = 0.046 for 2-months post-treatment, which were directly proportional to the SCORAD score improvement [L. fermentum VRI-003 use was not associated with reduced topical corticosteroid use [Based on a meta-analysis of 19 trials totaling 4076 intervention and 3700 control events, respectively, maternal probiotic consumption in late pregnancy and, during lactation, could reduce AD incidence in children younger than five years old, Risk Ratio 0.78, 95% Confidence Interval (CI) 0.68 to 0.90, I20 to 64 . A systeLactobacillus salivarius synbiotic (probiotic with prebiotics to increase probiotic delivery) was trialed against prebiotic only in sixty 2- to 14-year-old children with moderate to severe AD [L. salivarius synbiotic recipients had significantly lower SCORAD scores (27.4 \u00b1 12.7) than prebiotic recipients  [L. salivarius LS01 has been shown to preserve Th1 cytokine production and Th1:Th2 ratios when compared to maltodextrin controls after four months of treatment [L. salivarius\u2019 reduction of AD severity via normalization of T-cell function is consistent with the hygiene hypothesis [A evere AD . At eigh= 0.022) ,24. In areatment . L. salipothesis .Bifidobacterium longum infantis and B. breve combination in fermented infant formula achieved greater IgA responses to the poliovirus vaccine challenge than standard formula [L. salivarius and B. breve BR03 combination led to improved SCORAD indices in adult AD patients [B. breve Bb99 synbiotic with three additional probiotics given to 459 women during the last two to four weeks of pregnancy and given to their newborns for six months after delivery reduced AD in comparison to 463 controls (Odds Ratio (OR) = 0.74; 95% CI, 0.55\u20130.98, p = 0.04) [L. rhamnosus LCS-742 and B. longum infantis M63 synbiotic administered to 39 term newborns for six months reduced AD in comparison with 45 formula-fed control newborns  [p = 0.03, based on a SCORAD-weighted mean difference improvement of 7.32 for 80 treated participants from three studies [B. breve M-16V synbiotic versus placebo control in 90 infants less than seven months of age did not find differences in cytokine production or circulating regulator T-cell proportions [L. rhamnosus LC705, B. breve Bb99, and Propionibacterium freudenreichii spp. Shermanii JS, with 1223 subjects found that in cesarean delivered infants, IgE-associated allergic diseases at five years of age were significantly reduced  [A  formula . A L. sapatients . B. brev = 0.04) . Similar < 0.05) . Overall studies . Consistportions . However= 0.035) . It has L. plantarum K8 (KCTC 10887BP) lysates. The 2.1% L. plantarum K8 lysates increased forearm skin hydration by the fourth week (p = 0.03), but not after eight weeks of use (p = 0.068) [p = 0.00) and eight weeks (p = 0.007) of use [L. plantarum K8 lysates were associated with significantly decreased horny layer thickness of the forearm and face at four weeks and eight weeks with p = 0.002 and p = 0.000 for the face and p = 0.007 and p = 0.000 for the forearm [L. plantarum K8 lysates use was associated with statistically significant decreased TEWL of the forearm and face at eight weeks with p = 0.002 and p = 0.008, respectively [L. plantarum K8 lysates may be regarded as orally consumed skin moisturizers that reduce the need for topical corticosteroids and emollients for AD treatment [Lysates offer cost-savings over the use of selected single components. Lysates can be included in prepared foods. A randomized controlled trial (RCT) with 41 dry, dark-skinned 25- to 60-year-old patients tested orally consumed 2.1% Pseudomonas aeruginosa is known for opportunistic infections in burn patients through the use of a toll-like receptor-4 (TLR-4) associated lipopolysaccharide receptor complex to induce an inflammatory and adaptive immune response [L. plantarum, 105 cfu/mL injected into infected areas on days three, four, five, seven, and nine post infection inhibited P. aeruginosa PA100 colonization of an in vivo burned-murine model, which was measured by the lack of viable P. aeruginosa PA 100 cells, p < 0.001 [L. plantarum was tried against silver sulphadiazine in 80 patients with delayed, infected second-degree and third-degree burns, or early, non-infected third-degree burns [L. plantarum achieved a 2.72% lower bacterial load in second-degree burns and 1.07% less infection in early third-degree burns, but had 16.90% more infection in delayed, infected third-degree burns [response . L. plan < 0.001 . L. planee burns . Within ee burns . These ree burns . Lactobacillus reuteri ATCC 55730 at 105 to 108 cfu/mL and L. rhamnosus AC413 coinfections increase normal primary human epidermal keratinocytes (NHEK) survival in the presence of concurrent S. aureus infection from 8.8% in the absence of probiotics to 53.1% (p = 0.0012) and 42.7% (p < 0.0001), respectively [L. reuteri ATCC 55730 lysate also protects NHEK from S. aureus, p = 0.01. Heat-killed L. reuteri ATCC 55730 is ineffective against S. aureus infection [L. reuteri ATCC 55730 exposure after S. aureus infection is ineffective. L. reuteri ATCC 55730 introduction before or concurrently with S. aureus allows L. reuteri ATCC 55730 to competitively exclude S. aureus from NHEK binding sites, p = 0.026 and p = 0.0078, respectively [L. reuteri ATCC 55730, L. reuteri ATCC 55730 lysates are less effective staphylococcal adhesion inhibitors, p = 0.0002 versus p = 0.032 [L. salivarius UCC118 cannot be substituted for L. reuteri ATCC 55730 since L. salivarius UCC118 adheres less effectively to NHEK than does L. reuteri ATCC 55730, 5.6 \u00b1 0.1 log cfu versus 6.7 \u00b1 0.1 log cfu respectively, p = 0.005 [ectively . L. reutnfection . L. reutectively . Compare = 0.032 . L. saliS. aureus exposure with p = 0.006 and p = 0.01 after a 24-h incubation [S. aureus infection, p = 0.005 and p = 0.01, respectively [S. aureus from receptors [S. aureus from keratinocytes\u2019 receptors [S. aureus growth. However, spent culture did not inhibit S. aureus growth [In vitro studies show that LGG lysates and spent culture fluid can protect human keratinocytes from subsequent cubation . LGG andectively . Live LGeceptors . Live LGeceptors . Live LGs growth .Camellia sinensis derived tea is 20% green tea and is the second most common drink [Pigmented polyphenols absorb UVR primarily in the UVB spectrum with some UVA and UVC absorption, which facilitates sunscreen function . Globallon drink ,31. Topion drink . In murion drink .2) preceding UVR exposure also reduced photodamage [p = 0.02 [Anti-inflammatory, antioxidant, polyphenolic green tea catechins (GTC) inhibit cyclooxygenase-2 and lipoxygenases . Conjugatodamage . A 34-dap = 0.02 .2 to 28 mJ/cm2 in both intervention and placebo groups [+ T-lymphocyte counts were similar in each group, p = 0.85 and p = 0.62, respectively [Oral GTC metabolites are incorporated into the skin . Oral GTo groups . Post-UVectively .p < 0.05 versus 20.4 \u00b1 2.7%) [p < 0.05 [Topical green tea EGCG effects IL-12-dependent DNA repair . However \u00b1 2.7%) . \u03b1-Lipoi \u00b1 2.7%) . Interesp < 0.05 .p < 0.05, p < 0.01, and p < 0.001, respectively. Ferulic acid is least active with IC30 > 30 against melanin production and tyrosinase activity [p < 0.002) and peak wavelength absorption is increased (p < 0.001) [p \u2264 0.05) [p \u2264 0.05) [Aspalathus linearis (rooibos) tea. Caffeic acid and ferulic acid are constituents of Polypodium leucotomas, a tropical fern [P. leucotomas extract is commercially available in several products [P. leucotomas extract significantly reduced tumorigenesis and immunosuppression associated cyclobutane pyrimidine dimers formed by UVB action on DNA, p < 0.001 [P. leucotomas [Dietary phenolic acids are primarily UVA absorbing antioxidants with indirect regulatory effect on the nuclear factor E2-related factor 2-antioxidant responsive element (Nrf2-ARE) pathway . In primactivity . Nonetheactivity . Althoug< 0.001) . Rutin i \u2264 0.05) . UVA pho \u2264 0.05) . Of notecal fern . P. leucproducts . In a sm < 0.001 . This fiucotomas .Photoprotection and anti-photocarcinogenesis murine trials of resveratrol abound . In vivoSilymarin is up to 80% silibinin is GRAS . In addiVitamins C and E are normal skin components . UVR-indElaies guineensis , which was transported globally by the Portuguese in the 15th Century and the Dutch in the 19th Century [Elaeis oleifera  has more oleic and linoleic acid and less palmitic and saturated acids than E. guineensis [E. guineensis tocotrienols are unusual since rice bran oil is the only other tocotrienol containing vegetable oil [p < 0.03, when compared to control cells [Vitamin E comprised 30% of mixed tocopherols and 70% of mixed tocotrienols and a water-soluble phenolic-flavonoid-rich antioxidant complex are minor components of  Century . Elaeis ineensis . E. guinable oil . Palm oiable oil . Howeverable oil . Tocotriable oil . Consistable oil ,45. Nanoable oil . In vitrable oil . Cell viol cells .RRR-\u03b1-tocopherol daily for 12-weeks reduced dorsal skin erythema (p < 0.01) by week nine in comparison to 25 mg total carotenoids daily [Lactobacillus johnsonii 5.10 \u00d7 108 cfu was the intervention in a 12-week randomized, double-blinded, placebo-controlled trial with 60 patients who had polymorphic light eruptions [L. johnsonii 5.10 \u00d7 108 cfu was able to reduce the effect of a single exposure to longwave UV radiation (UVA1), p < 0.001, but that of multiple exposures [p < 0.001, and p = 0.022 in the placebo group [p = 0.03), increased procollagen deposition (p = 0.05), reduced UVR-induced matrix metalloproteinase-1 (p = 0.04), and reduced mitochondrial DNA 3895-base pair deletion after 3\u00d7 minimal erythema dose UV exposure (p = 0.01) [Based on 20 healthy adults, an oral combination of 25 mg total carotenoids and 335 mg  ds daily . Combinads daily . This isds daily . A commeruptions . The comxposures . UVA1-inxposures ,49. Intr = 0.01) . Based o = 0.01) . The dos = 0.01) .2 \u00b1 0.04, 50 mg coenzyme Q10 MED was 0.72 J/cm2 \u00b1 0.06 and 150 mg coenzyme Q10 MED was 0.70 J/cm2 \u00b1 0.06, p = 0.49 [The antioxidant, coenzyme Q10 is essential for mitochondrial energy metabolism . In contp = 0.49 .2, p < 0.01, and reduced p53 UV-induced skin expression by 50%, p < 0.01 [A 3-month, 42-person, double-blind randomized trial of omega-3 polyunsaturated fatty acids 4 g daily, eicosapentaenoic acid (EPA), or monounsaturated oleic acid found that EPA was the most promising for photo-protection . EPA incp < 0.01 . This waL. salivarius symbiotic should be considered for AD treatment given reduced AD severity measured by SCORAD, p = 0.022 [B. breve may need additional trials as a combination probiotic in fermented foods before significant effects on AD was seen [L. rhamnosus LC705, B. breve Bb99, and Propionibacterium freudenreichii spp. Shermanii JS, reduced IgE-associated allergic diseases in cesarean-delivered infants (p = 0.035), but B. breve as a single administered probiotic did not affect cytokine production or circulating regulator T-cell proportion [The preceding sections and the synopsis of selected references in  = 0.022 . Probiotwas seen . The comoportion ,27. Succoportion ,27.L. plantarum K8 lysates are skin moisturizers, which increase facial skin hydration through eight weeks of use (p = 0.007), decrease the horny layer thickness of the forearm and face through eight weeks of use (p = 0.000 for the face and forearm), and decrease TEWL of the forearm and face at eight weeks  [L. plantarum K8 lysates should reduce the need for topical corticosteroids and emollients for AD treatment [L. plantarum is a candidate for further larger scale in vivo study as a topical burn treatment [Orally consumed 2.1% ctively) . Thereforeatment . Decreasreatment ,8. Givenreatment ,29. Ideareatment .L. reuteri ATCC 55730 for S. aureus opportunistic infection ensures the effectiveness of the non-denatured probiotic [Live or lysate probiotic preparations are biologically plausibly efficacious where heat-killed preparations are ineffective ,18. Liverobiotic ,18. Therrobiotic .Oral and topical polyphenols hold the photoprotection and anti-photocarginogenesis promise. Additional human trials on GTC and EGCG are needed. Formulations with improved GTC and EGCG bioavailability such as combinations with \u03b1-lipoic acid may allow anti-inflammatory photo protective effectiveness at doses equivalent to less than 10 cups of green tea daily ,30,32. H"}
+{"text": "Escherichia coli (UPEC) are one of the main bacteria causing urinary tract infections (UTIs). The rates of UPEC with high resistance towards antibiotics and multidrug-resistant bacteria have increased dramatically in recent years and could difficult the treatment.Uropathogenic E. coli isolated from community (79 isolates) and hospital-acquired (31 isolates) urinary tract infections. The plasmid-mediated quinolone resistance genes presence was also investigated. A subset of 18 isolates with a quinolone-resistance phenotype was examined for common virulence genes encoded in diarrheagenic and extra-intestinal pathogenic E. coli by a specific E. coli microarray.The aim of the study was to determine multidrug-resistant bacteria, antibiotic resistance profile, virulence traits, and genetic background of 110 kpsMTII gene. Overall, fimH (71.8%) and fyuA (68.2%), had the highest prevalence as virulence genes among isolates. However, the profile of virulence genes displayed a great diversity, which included the presence of genes related to diarrheagenic E. coli. Out of 110 isolates, 25 isolates (22.7%) were positive to qnrA, 23 (20.9%) to qnrB, 7 (6.4%) to qnrS1, 7 (6.4%) to aac(6\u2032)lb-cr, 5 (4.5%) to qnrD, and 1 (0.9%) to qnrC genes. A total of 12.7% of the isolates harbored blaCTX-M genes, with blaCTX-M-15 being the most prevalent.Female children were the group most affected by UTIs, which were mainly community-acquired. Resistance to trimethoprim\u2013sulfamethoxazole, ampicillin, and ampicillin\u2013sulbactam was most prevalent. A frequent occurrence of resistance toward ciprofloxacin (47.3%), levofloxacin (43.6%) and cephalosporins (27.6%) was observed. In addition, 63% of the strains were multidrug-resistant (MDR). Almost all the fluoroquinolone (FQ)-resistant strains showed MDR-phenotype. Isolates from male patients were associated to FQ-resistant and MDR-phenotype. Moreover, hospital-acquired infections were correlated to third generation cephalosporin and nitrofurantoin resistance and the presence of E. coli may be difficult to treat empirically due to high resistance to commonly used antibiotics. Continuous surveillance of multidrug resistant organisms and patterns of drug resistance are needed in order to prevent treatment failure and reduce selective pressure. These findings may help choosing more suitable treatments of UTI patients in this region of Mexico.Urinary tract infection due to  Escherichia coli is an important cause of extra-intestinal infections, enteric disease, and systemic infections in humans and animals. Uropathogenic Escherichia coli (UPEC), one of the members of the extra-intestinal pathogenic E. coli (ExPEC) is a predominant pathogen causing urinary tract infections (UTIs) . Trimethoprim\u2013sulfamethoxazole and cephalosporins [cefotaxime (p\u2009=\u20090.0232) and ceftazidime (p\u2009=\u20090.0001)] also showed a co-resistance.Co-resistance phenotype between cephalosporins and beta-lactams towards fluoroquinolones was frequently observed [(amoxicillin\u2013clavulanic acid (p\u2009=\u20090.0306), ceftriaxone (p\u2009=\u20090.0305), and nitrofurantoin (p\u2009=\u20090.0002) resistance phenotype (Table\u00a0p\u2009=\u20090.02329). MDR-phenotype (69.6%) as well as the FQ-resistant phenotype  was also prevalent among older adults (>\u200960\u00a0years old).Overall, hospital-acquired isolates were slightly more resistant than community-acquired isolates . Both hospital and community-acquired isolates showed higher resistance towards ampicillin (77.4% vs 68.4%) and trimethoprim\u2013sulfamethoxazole (77.4% vs 70.9%). UPEC isolates from hospital-acquired infections were associated to ceftazidime (E. coli isolates. The phylo-groups D (23.6%) and A (19.1%) were the most commonly found, followed by B1 (15.5%), C (13.6%), B2 (11.8%), F (10%), cryptic clades (5.5%) and phylo-group E (0.9%). The different phylo-groups and cryptic clades were distributed in both hospital and community settings, except the phylo-group E which was only distributed in the hospital-acquired infections. MDR strains were distributed into all phylogenetic-groups: 26% (16/70) to A, 17.1% (12/70) to D, 15.7% (11/70) to B1, 12.9% (9/70) to C, 11.4% (8/70) to B2 and F, and 6.7% (5/70) to cryptic clades. Phylo-group D was significantly associated to MDR-phenotype (p\u2009=\u20090.0339) as well as FQ-resistant phenotype . Phylo-groups B2 and D were more common among community-acquired isolates  than hospital-acquired isolates , as well as in females  than in males  but not significantly associations were found.All seven phylogenetic lineages  and cryptic clades were found on the 110 urinary fimH (71.8%), fyuA (68.2%), and agn43 (54.5%) were the virulence genes with the highest distributions among isolates, while afa/dra (8.2%) and cnf1 (2.7%) had the lowest. Others virulence genes tested had the following distribution: chuA, 49.1%; papC, 42.7%; kpsMTII, 37.3%; vat, 20%; yfcV, 20%; sfaS 10%, and hlyA, 9.1% (Table\u00a0fyuA (71.0% vs 67.1%), papC (48.4% vs 40.5%), yfcV (25.8% vs 55.7%), hlyA (12.9% vs 7.6%), cnf1 (3.2% vs 2.5%), kpsMTII (58.1% vs 29.1%) and chuA (51.6% vs 48.1%) were higher among UPEC isolated from hospital-acquired than community-acquired infections. However, only the kpsMTII was significantly associated with hospital-acquired isolates . The presence of the chuA gene was associated to UPEC isolated from children (p\u2009=\u20090.0483). Furthermore, the presence of vat was associated to UPEC isolates from adults (p\u2009=\u20090.0153).Overall, nd agn43 4.5% werefimH (72.9%), fyuA (65.7%), agn43 (55.7%) and chuA , fyuA (67.3%) and chuA (57.7%) were highly distributed among FQ-resistant bacteria.Virulence genes ), fyuA 6.7%, agn4E. coli isolates belonging to the phylo-groups B2, D and F. Overall, 145 virulence and 40 antimicrobial resistance genes among the 315 and 82 genes and variants investigated were detected at least once in one or more of the isolates. The total number of virulence genes per isolate ranged from 18 to 63 genes. The median number of the virulence genes per isolate was 35. Microarray hybridizations demonstrated that all the E. coli isolates tested possessed virulence genes related to a pathotype including ExPEC and diarrheagenic E. coli (DEC). The total number of antimicrobial resistance genes per isolate ranged from 0 to 21 genes with a median number of 6.5 genes. Since the microarray carries a large set of virulence factors, numerous incomplete ExPEC that would normally be missed in a PCR-based assay were found. Thus, various unusual gene combinations were discovered, such as ExPEC pathogenic profiles with assorted EPEC genes coding for the type III secretion system 2 proteins ErpJ and SpaS . Moreover, detection of virulence (mean 39.0\u2009\u00b1\u200916.9) and antimicrobial resistance (mean 9.0\u2009\u00b1\u20091.9) genes were higher on hospital-acquired UTIs isolates compared to community-acquired infections , 20.9% (23 isolates) to qnrB, 6.4% (7 isolates) to qnrS1, 0.9% (1 isolates) to qnrC, 4.5% (5 isolates) to qnrD, and 6.4% (7 isolates) to aac(6\u2032)-Ib-cr , levofloxacin (LEV), and norfloxacin (NOR). The double mutations in Ser-83\u2009\u2192\u2009Leu and Asp-87\u2009\u2192\u2009Tyr or Asp-87\u2009\u2192\u2009Asn substitution in gyrA were present in 92% of the strain tested. Ser-80\u2009\u2192\u2009Ile substitution in parC were found in 92% of the isolates tested, which also were resistant to both ciprofloxacin and levofloxacin. The substitution Thr-66\u2009\u2192\u2009Ser , Thr-66\u2009\u2192\u2009Asn , and Thr-66\u2009\u2192\u2009Tyr  were also found in parC. Other substitutions in parC as Glu-84\u2009\u2192\u2009Val  and Glu-84\u2009\u2192\u2009Ala were also detected. Interestingly, one strain (isolated UEc 76) which was resistant to ciprofloxacin, levofloxacin and norfloxacin, belonging to B2 phylo-group and blaCTX-M-containing, also carried the double mutations in Ser-83\u2009\u2192\u2009Leu and Asp-87\u2009\u2192\u2009Asn substitution in gyrA and the triple mutation in parC Ser-80\u2009\u2192\u2009Ile, Thr-66\u2009\u2192\u2009Tyr and Glu-84\u2009\u2192\u2009Val.Overall, 22% of the isolates were positive for rB, 6.4%  isolatesblaCTX-M (12.7%/14 isolates), blaTEM , blaPSE , and blaOXA (9.1%/10 isolates). Sequencing of the blaCTX-M amplicons identified the presence of blaCTX-M-15 in 7.2% of the strains (8 isolates), blaCTX-M-12 in 4.5% (5 isolates), blaCTX-M-3 in 1.8% (2 isolates), and blaCTX-M-14 in 1.8% (2 isolates). Furthermore, blaROB-1,blaSHV-37, and blaCMY-2 genes, were identified in one isolate (0.9%) through the microarray assay.ESBL genes were investigated through microarray and PCR assay. A total of 27 isolates (24.5%) suspected as cephalosporinase producers showed positive PCRs for blaCTX-M genes and PMQR genes as well as mutation in QRDR were found in both community- and hospital-acquired infections as well as in male and female genders. The PMQR qnrA (21.4%), and qnrB (18.6%), were highly distributed on MDR-phenotype (Table\u00a0aac(6\u2032)-Ib-cr gene compare to female isolates . Quinolone-resistance isolates also presented higher prevalence to blaCTX-M compared to quinolone-sensitive phenotype isolates .Overall pe Table\u00a0. IsolateIn accordance with global trends, our results reveal higher prevalence of urinary tract infections in female patients than in males , 34\u201336. Multidrug-resistant strain prevalence (63.3%) was higher in this work than in other study from Mexico, which reported only 16.4% of MDR strains . While wE. coli isolates from hospital-acquired UTIs are correlated to the presence of kpsMTII gene that has been associated to pyelonephritis, a more severe infection of the upper urinary tract [fyuA-encoding yersiniabactin receptor and, fimH encoding type 1 fimbrial adhesion.As detected by Ochoa et al. , phylo-gry tract . In agrery tract , 41, ourE. coli (EAEC and DAEC), including capU , deoK (deoxyribokinase), shf , virK  and astA gene (EAEC heat-stable enterotoxin 1). This was also reported in previous works [E. coli reported in ExPEC strains. The deoK operon is frequently associated with strains isolated from infected urine and blood and is part of a large genetic island carrying genes contributing to the strain intrinsic virulence and/or adaptive properties [Detailed analysis of a strains subset using microarrays, revealed that some of them from both, hospital- and community-acquired infections carried virulence related genes of enteroaggregative and diffusely adherent us works \u201345, whicoperties . These soperties .E. coli (EPEC) related genes on five UPEC isolates. These genes included the eae , as well as virulence related genes such as espG , eprJ (E. coli type III secretion system 2 protein ErpJ), epaS (E. coli type III secretion system 2 protein EpaS), and eivG (E. coli type III secretion system 2 protein EivG) [Another finding was the presence of enteropathogenic in EivG) , 46. T3Sin EivG) , 48. Thein EivG) , 49, 50.qnrA, qnrB, qnrS, qnrC and qnrD as well as aac(6\u2032)-Ib-cr were detected. The most common PMQR-genes identified were qnrA and qnrB, which is in contrast to previous studies were aac(6\u2032)-Ib-cr gene detection was more frequent on ExPEC isolates [qnrA and qnrB were frequently found as a part of MDR-phenotype. qnrD gene was found in 4.5% of the isolates. Previous papers reported qnrD in bacteria isolated from rooks [Salmonella enterica serovar Kentucky, serovar Bovismorbificans, Proteus mirabilis and Morganella morganii [E. coli from dogs [qepA was not identified. Although PMQR genes provide a low level of FQ-resistance, they have been reported to favor the selection of additional chromosome-encoded resistance mechanism [gyrA and parC were prevalent in community-acquired infections (89% vs 11%). Overall, mutations in QRDR were equally present among isolates from males and females patients.In this study, UPEC strains carrying PMQR genes associated to quinolones resistance including isolates , 51. Morom rooks  and humamorganii , 53, as rom dogs . In accorom dogs , 55, 56,echanism . In our blaCTX-M , blaTEM, blaPSE, and blaOXA were detected. For the isolates carrying resistance genes, ESBL genes such as TEM, SHV and CTX-M, are the most widespread and frequently detected in E. coli [In our study, ESBL genes  E. coli . In addi E. coli , 59.E. coli UTI isolates from hospital and community in Mexico. Diverse genotypes and phenotypes including multidrug-resistance, fluoroquinolone resistance and carriage of virulence genes related to several enteropathogenic E. coli were found among UPEC isolates. Thus, continuous surveillance for antimicrobial resistance of UPEC is needed in order to prevent treatment failure and to improve the strategies to mitigate the occurrence of antibiotic resistance organisms and ensure the best treatment to UTI patients. This study highlights the importance of antimicrobial resistance of virulent E. coli from urinary tract infection in Mexico.Our study describes significant antimicrobial resistance in"}
+{"text": "The impact of STAR-VA on psychotropic drug use among residents with behavioral symptoms of dementia was evaluated through a difference-in-differences framework. STAR-VA residents enrolled 2013-2017 were evaluated longitudinally pre-post intervention. The primary outcome was the number of as needed administrations with an indication of \u2018anxiety\u2019 or \u2018agitation\u2019. The analytical cohort included 214 training cases and 1,870 controls from untrained sites meeting eligibility criteria. STAR-VA cases were less white (48% vs. 54%), less black (11% vs. 14%), and had significantly longer median length of stay (830 vs. 261 days), respectively. STAR-VA cases had on average 3.5 as needed doses/month of psychotropic medication before the intervention and 1.7 after, controls averaged 1.8 doses/month. After adjustment for person-time-fixed effects, enrollment was associated with 55%  reduction or an average 0.8 as needed psychotropic doses/month. Findings demonstrate effectiveness in decreasing as-needed psychotropic drug use among CLC residents, supporting continued implementation of STAR-VA."}
+{"text": "Nature Communications 10.1038/s41467-017-02626-6; published online 15 Jan 2018Addendum\u00a0to: Nat. Commun. 7, 11369 (2016) and Phys. Rev. Mat. 1, 035604 (2017)), which report the spontaneous helix formation in a polar smectic liquid crystal phase made of achiral bent-core mesogens.We would like to make our readers aware of the related publications by S.P. Sreenilayam et al. ("}
+{"text": "Scientific Reports 10.1038/s41598-019-46738-z, published online 12 August 2019Correction to: This Article contains an error in the Acknowledgements section.\u201cThis study also partly supported by Incheon National University (INU) Research Grant (2016) to H.W.K. and the INU Postdoctoral Fellowships to U.Y. and R.I.\u201dshould read:\u201cThis study also partly supported by Incheon National University (INU) Research Grant (2016) to H.W.K.\u201d"}
+{"text": "Correction to: J ImmunoTher Cancer (2019) 7:288https://doi.org/10.1186/s40425-019-0778-7Following publication of the original article , the aut3)\u201d should change to black font;A: Red font \u201cMHC-I c reduces macrophage frequency and activates macrophages, cDC, B and NK cells. Figure S3. Poly(I:C)c triggers transcriptional reprogramming of MDSC. Figure S4. Significantly regulated genes in PMN- and M-MDSC upon poly(I:C)c therapy."}
+{"text": "Scientific Reports 10.1038/s41598-017-10484-x, published online 31 August 2017Correction to: The Acknowledgements section in this Article is incomplete.\u201cWe thank D.M. Buss for his comments.\u201dshould read:\u201cWe thank D.M. Buss for his comments. Study was funded by National Science Centre-PL-2014/13/B/HS6/02644\u201d"}
+{"text": "AbstractKSU). Included in this material, three species of Labiobaetis Novikova & Kluge, 1987 are recorded, two of them being new to science. Labiobaetispotamoticus Gattolliat & Al Dhafer, sp. n. is described from both larvae and adults, whereas Labiobaetisalahmadii Gattolliat & Al Dhafer, sp. n. is only known from the larval stage. The two species are compared morphologically with Palearctic and Afrotropical species of Labiobaetis. A third species, Labiobaetisglaucus  is reported for the first time from the Arabian Peninsula. The species was originally described from South Africa and subsequently reported from the east and northeast of Africa. A molecular reconstruction including 18 Afrotropical and Palearctic species of Labiobaetis was performed using 658 bp of the mitochondrial gene CO1. The reconstruction highly supported the validity of the two new species and confirmed the occurrence of L.glaucus in KSU.Mayfly larvae and imagos were collected at approximately 50 localities of the Kingdom of Saudi Arabia ( Labiobaetis Novikova & Kluge, 1987 is a species-rich genus with an almost worldwide distribution ; it is mainly diversified in Afrotropical (28 species) and Oriental realms (23 species)  . The staspecies) . Moleculphyletic . Larvae KSU) and Yemen despite potential suitable habitats present in Oman, Jordan and the United Arab Emirates  were mentioned from KSU ; other paratypes are housed in the Museum of Zoology, Lausanne, Switzerland (MZL). Each GBIFCH code refers to a tube with group of specimens in or a slide with a single specimen (sequenced or not).The majority of the material was collected in November 2012 during a scientific expedition organised by King Saud University Museum of Arthropods, College of Food and Agriculture Sciences, Department of Plant Protection, CO1) gene. Specimens belonging to the different \u201cmorphospecies\u201d and collected in the same localities were selected for genetic analysis. The CO1 gene was sequenced using LCO1490 and HCO2198 primers ) as the maximal value for intraspecific divergence : Saudi Arabia (AR44); Wadi Shahadan; 17\u00b028'36\"/ 42\u00b042'50\"; Alt. 190m; 13.XI.2012; Coll. J-L Gattolliat.Paratypes: 4 larvae (GBIFCH00235716 + GBIFCH00235735 (Genetics)), 1 male imago (GBIFCH00235732 (Genetics)) + 3 larvae (KSU: GBIFCH00526192); same data as holotype.19\u00b055'32\"/ 41\u00b026'17\"; Alt. 752m; 13.X.2010; Coll. B. Kondratieff.42 larvae : Saudi Arabia (AR01); Al Jiwah, Thee Aine; 18\u00b047'29\"/ 41\u00b056'19\"; Alt. 490m; 13.III.2012; Coll. Al Dhafer, H. & Kondratieff, B.14 larvae (GBIFCH00235721): Saudi Arabia (AR20); Wadi Baqrah; 19\u00b055'46\"/ 41\u00b026'34\"; Alt. 760m; 3.VI.2012; Coll. Al Dhafer, H. & Kondratieff, B.5 larvae (GBIFCH00235722): Saudi Arabia (AR28); Thee Ain, Al-Baha; 19\u00b055'46\"/ 41\u00b026'34\"; Alt. 760m; 8.XI.2012; Coll. J-L Gattolliat.2 larvae (GBIFCH00235714): Saudi Arabia (AR31); Thee Ain, Al-Baha; KSU: GBIFCH00526173): Saudi Arabia (AR32); Wadi Elarj, near Adam; 20\u00b027'11\"/ 40\u00b048'56\"; Alt. 440m; 9.XI.2012; Coll. J-L Gattolliat.58 larvae , GBIFCH00517521 (Genetics)) + 11 larvae (KSU: GBIFCH00526224): Saudi Arabia (AR43a); Wadi Shahadan; 17\u00b028'36\"/ 42\u00b051'25\"; Alt. 460m; 12.XI.2012; Coll. J-L Gattolliat.23 larvae , GBIFCH00465155 (Genetics)) + 4 larvae (KSU: GBIFCH00526237): Saudi Arabia (AR43b); Wadi Shahadan; 17\u00b028'17\"/ 42\u00b051'14\"; Alt. 455m; 12.XI.2012; Coll. J-L Gattolliat.3 larvae .1 larva (on slide), Saudi Arabia, Wadi Buwah, 1340m, Larva: Tergites I-X medium brown with peculiar pattern formed of six ecru dots Fig. . Scape oLarva.Length: fully grown female: Body 5.1\u20137.7 mm, cerci 3.6\u20134.0 mm, terminal filament 2.5\u20132.6 mm. Fully grown male: Body 5.0\u20135.3 mm, cerci 3.4\u20133.6 mm, terminal filament 2.5 mm.Colouration : Saudi Arabia (AR43a); Wadi Shahadan; 17\u00b028'36\"/ 42\u00b051'25\"; Alt. 460m; 12.XI.2012; Coll. J-L Gattolliat.Paratypes: 151 larvae , GBIFCH00235737) + 8 larvae (KSU: GBIFCH00526208): same data as holotype.KSU: GBIFCH00526227): Saudi Arabia (AR43b); Wadi Shahadan; 17\u00b028'17\"/ 42\u00b051'14\"; Alt. 455m; 12.XI.2012; Coll. J-L Gattolliat.47 larvae (GBIFCH00235710) + 10 larvae (KSU: GBIFCH00526223): Saudi Arabia (AR39); Wadi Damad; 17\u00b012'21\"/ 43\u00b001'35\"; Alt. 260m; 11.XI.2012; Coll. J-L Gattolliat.29 larvae (GBIFCH00235727) + 6 larvae (KSU: GBIFCH00526179): Saudi Arabia (AR44); Wadi Shahadan; 17\u00b028'36\"/ 42\u00b042'50\"; Alt. 190m; 13.XI.2012; Coll. J-L Gattolliat.15 larvae , GBIFCH00465155 (Genetics) + GBIFCH00517526 (Genetics), GBIFCH00517527 (Genetics)) + 4 larvae Baetisglaucus Agnew, 1961: 14.Pseudocloeonglaucum , Labiobaetisglaucus , 19\u00b055'32\"/ 41\u00b026'17\"; Alt. 752m; 13.X.2010; Coll. B. Kondratieff.18 larvae (GBIFCH00235711 + 4 slides GBIFCH00235741 (Genetics), GBIFCH00235746, GBIFCH00235750 (Genetics), GBIFCH00235756: Saudi Arabia (AR01); Al Jiwah, Thee Aine; 19\u00b005'22\"/ 41\u00b058'16\"; Alt. 490m; 13.III.2012; Coll. Al Dhafer, H.3 larvae (GBIFCH00235708): Saudi Arabia (AR19); Wadi Khat; 19\u00b055'43\"/ 41\u00b026'34\"; Alt. 760m; 3.VI.2012; Coll. Al Dhafer, H. & Kondratieff, B.1 larva (GBIFCH00235712): Saudi Arabia (AR28); Thee Ain, Al-Baha; 19\u00b055'43\"/ 41\u00b026'34\"; Alt. 760m; 8.XI.2012; Coll. J-L Gattolliat.3 larvae (GBIFCH00235713): Saudi Arabia (AR31); Thee Ain, Al-Baha; 17\u00b028'36\"/ 42\u00b051'25\"; Alt. 460m; 12.XI.2012; Coll. J-L Gattolliat.1 larva (GBIFCH00235707): Saudi Arabia (AR43a); Wadi Shahadan; 17\u00b028'17\"/ 42\u00b051'14\"; Alt. 440m; 12.XI.2012; Coll. J-L Gattolliat.2 larvae GBIFCH00465151 (Genetics): Saudi Arabia (AR43b); Wadi Shahadan; 17\u00b028'36\"/ 42\u00b042'50\"; Alt. 190m; 13.XI.2012; Coll. J-L Gattolliat.7 larvae (GBIFCH00235723 + 1 slide GBIFCH00235738), 3 male imagos (GBIFCH00235724 + 1 slide GBIFCH00235731 (Genetics)): Saudi Arabia (AR44); Wadi Shahadan; Larva: abdominal pattern  with moderate current. The substrate was a mix of sand, cobbles and rocks Figs , 52. ThiL.potamoticus and L.alahmadii as monophyletic clades (BS (Bootstrap support) of 83% and 100% respectively), with intraspecific K2P distances below 1% , with the three populations  supported as monophyletic sister-clades . The sister group of Labiobaetispotamoticus is an undescribed species from South Africa; the distance between the two taxa is slightly higher than intraspecific distance (between 4.2 and 5.1%). Labiobaetispotamoticus possesses high distances to all the other species included in the study (16.2 to 25.5%). The relationships of L.alahmadii and L.glaucus with other species of Afrotropical and Palaearctic origins also are unclear and have no molecular support  and L.balcanicus and the Afrotropical species L.boussoulius . The shape of the distolateral process of the second segment of the labial palp is of taxonomic importance to separate the different species: more elongated and curved in L.potamoticus , the shape of the apex of the segment II of the maxillary palp , distal margin of tergites , tarsal claw less curved and less stout in L.glaucus  and the setation of the ventral margin of femora . Labiobaetispotamoticus and L.piscis show a peculiar pattern on the abdominal tergites . Labiobaetispotamoticus was initially identified as L.balcanicus  and the shape of the distomedial projection of the segment II of the labial palp .Labiobaetis are generally similar. The presence or absence of the hindwings and the shape of the genital plates are the main characters to separate the species (Labiobaetispotamoticus cannot be separated from most other species of Labiobaetis with hindwings and broad, apically flat genital plates. Labiobaetisglaucus and L.boussoulius differ from most other species of the genus by the presence of a well-marked triangular expansion on the inner margin of the gonopods (Fig. L.tricolor Tshernova, 1928 (fig. 111a in The imagos of the different species of  species . The malods Fig. . A similL.glaucus in KSU is rather unexpected despite that this species is widely distributed in Afrotropical Region (South Africa, Angola, Kenya, Lesotho, Namibia, and Zimbabwe (KSU. We found no clear morphological differences between them and they fully correspond to the original description and subsequent redescriptions (KSU form a highly supported monophyletic clade. Genetic distances between the three populations are clearly of intraspecific range especially if we consider that intermediate populations from East and North-East Africa are not included in the analysis.The discovery of Zimbabwe ; Comorosriptions . As alreriptions , segment"}
+{"text": "Salmonella enterica serovar Typhimurium is a Gram-negative pathogen and a primary cause of foodborne illnesses worldwide. Here, we present the complete 47,393-bp genome sequence of the siphophage Skate, which was isolated against S. Typhimurium strain LT2. Salmonella enterica serovar Typhimurium is a Gram-negative pathogen and a primary cause of foodborne illnesses worldwide. Here, we present the complete 47,393-bp genome sequence of the siphophage Skate, which was isolated against S. Typhimurium strain LT2. Salmonella enterica serovar Typhimurium is a Gram-negative pathogen and a primary cause of foodborne illnesses worldwide  was used for quality control of the reads. The reads were trimmed with FASTX-Toolkit 0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/download.html) before being assembled into a single contig at 160.9-fold coverage using SPAdes 3.5.0 (http://transterm.cbcb.umd.edu/). Sequence similarity searches by BLASTp 2.2.28 (https://cpt.tamu.edu/).The phage Skate was isolated from soil in the cattle holding pen of a cattle harvesting facility in Michigan in August 2016 using rain LT2 . The pharain LT2 . Phage grain LT2 . Pooled es 3.5.0 . Contig es 3.5.0  and Metaes 3.5.0  were usees 3.5.0 . Rho-indp 2.2.28  were usep 2.2.28  interfacSalmonella phages, like phage IME207 (GenBank accession number KX523699) and E1 (GenBank accession number AM491472)  , 19, shaMH321493. The associated BioProject, SRA, and BioSample accession numbers are PRJNA222858, SRR8787571, and SAMN11259649, respectively.The genome sequence of phage Skate was submitted to GenBank as accession number"}
+{"text": "The generated UHPLC-MS data were analysed and dereplicated using the scientific databases Dictionary of Natural Products and SciFinder Scholar in order to potentially identify new dihydro-\u03b2-agarofurans from local Celastraceae plants. These investigations led to the large-scale extraction and isolation work on a prioritised fruit sample that belonged to the rainforest plant Denhamia celastroides. Chemical investigations resulted in the purification of four new natural products, denhaminols O\u2013R (1\u20134), along with the related and known compound, denhaminol G (5). The structures of all the new compounds were determined via detailed analysis of NMR and MS data.An analytical method using UHPLC-MS was developed and applied to 16 crude CH To confirm the presence of the new compounds and to unambiguously identify the structures, we conducted a large-scale extraction of the plant material and MS-directed purification to isolate the targeted compounds with the distinct molecular weights before NMR experiments were conducted.The scientific databases search  indicateD. celastroides (10 g) were sequentially extracted with CH2Cl2. Subsequent purifications using silica gel column chromatography and RP-HPLC afforded five dihydro-\u03b2-agarofurans , respectively. During the isolation of the three targeted compounds, another new dihydro-\u03b2-agarofuran  and a known congener, denhaminol G (5) +; (+)-HRESIMS m/z 637.2595 [M + Na]+ .Colourless gum; c 0.075, MeOH); ECD \u03bbext (MeOH) 217 (\u22124.79), 268 (\u22123.81) nm; UV (MeOH) \u03bbmax (log \u03b5) 282 (4.38) nm; IR (UATR) \u03bdmax 2952, 1739, 1705, 1373, 1197, 1162, 1076, 975, 769 cm\u22121; 1H-NMR  see 13C-NMR  see m/z 657 [M + H]+; (+)-HRESIMS m/z 679.2693 [M + Na]+ .Colourless gum; c 0.510, MeOH); ECD \u03bbext (MeOH) 216 (\u22124.60), 253 (\u22122.01), 301 (0.29) nm; UV (MeOH) \u03bbmax (log \u03b5) 630 (4.441) nm; IR (UATR) \u03bdmax 3419, 2971, 1747, 1710, 1391, 1278, 1120, 1163, 1078, 974, 713 cm\u22121; 1H-NMR  see 13C-NMR  see m/z 631 [M + H]+, 653 [M + Na]+; (+)-HRESIMS m/z 653.2531 [M + Na]+ .Colourless gum; c 0.110, MeOH); ECD \u03bbext (MeOH) 216 (\u22125.37), 235 (0.92), 279 (\u22121.31) nm; UV (MeOH) \u03bbmax (log \u03b5) 224 (4.42), 281 (4.39) nm; IR (UATR) \u03bdmax 3414, 2981, 1746, 1709, 1635, 1277, 1199, 1163, 1077, 973, 712 cm\u22121; 1H-NMR  see 13C-NMR  see m/z 653 [M + H]+; (+)-HRESIMS m/z 675.2400 [M + Na]+ .Colourless gum; 1\u20134) along with a known compound denhaminol G (5) were successfully isolated and characterised from one prioritised plant sample, namely the fruits of D. celastroides. Other Celastraceae samples that were prioritised for large-scale extraction and isolation studies will be investigated in the future. The pure compounds reported in this paper will be added to the Davis Open-Access Compound Library which is housed at Compounds Australia, Griffith University [A UHPLC-MS method and dereplication process was developed for the rapid identification of new dihydro-\u03b2-agarofurans from Celastraceae plants. This study further exemplifies how UHPLC-MS data and database mining can be used to extract molecular features related to characteristic secondary metabolites of a plant family and thus expedite new discoveries in natural products research. This approach was successfully applied to 16 Australian Celastraceae plant extracts. Consequently, four previously undescribed dihydro-\u03b2-agarofurans (denhaminols O\u2013R, iversity ,23,24,25"}
+{"text": "Scientific Reports 10.1038/s41598-018-21753-8, published online 23 February 2018Correction to: In the original version of this Article, Jakub Wlodarczyk was incorrectly affiliated with \u201cCenter of New Technologies, University of Warsaw, Banacha 2c Street, Warsaw, 02-097, Poland\u201d. The correct affiliation is listed below.Nencki Institute of Experimental Biology, Polish Academy of Sciences, Pasteura 3, Warsaw, 02-093, Poland.Additionally, the Acknowledgements section was incomplete.\u201cThis work was supported by the National Science Centre Grant - UMO-2015/17/B/NZ3/00557  and by an EMMA-West grant from the EU (S. B.). SB was also supported by a FASTTRACK grant (SR/FTP/ETA-04/2012) from DST and a UGC Research Award (F.30-31/2016 SA-II) from the Government of India. D.P. and S.B. were supported by the Polish National Science Centre (2014/15/B/ST6/05082) and D.P. by COST BM1405 and BM1408 EU actions.\u201dnow reads:\u201cThis work was supported by the National Science Centre Grant - UMO-2015/17/B/NZ3/00557  and by an EMMA-West grant from the EU (S.B.). S.B. was also supported by a FASTTRACK grant (SR/FTP/ETA-04/2012) from DST and a UGC Research Award (F.30-31/2016 SA-II) from the Government of India. M.M was also supported by the National Science Center Grant 2016/21/N/NZ3/03273. N.D. was supported by the PURSE-II project . D.P. and S.B. were supported by the Polish National Science Centre (2014/15/B/ST6/05082), and by Foundation for Polish Science (TEAM to D.P.). D.P. was also supported by grant 1U54DK107967-01 \u2018Nucleome Positioning System for Spatiotemporal Genome Organization and Regulation\u2019 within 4DNucleome NIH program (TEAM grant to D.P.) and COST BM1405 and BM1408 EU actions.\u201dThese errors have now been corrected in the PDF and HTML versions of the Article."}
+{"text": "Salmonella enterica serovar Heidelberg phage Matapan. A myophage with a 157,408-kb genome, Matapan is most closely related to Vi01-like phages.Here, we describe the  Salmonella enterica serovar Heidelberg phage Matapan. A myophage with a 157,408-kb genome, Matapan is most closely related to Vi01-like phages.Here, we describe the  Salmonella enterica serovar Heidelberg is a member of Enterobacteriaceae and a zoonotic pathogen  at 37\u00b0C with aeration, and phage isolation and propagation were done using the soft agar overlay method (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/) and trimmed with the FASTX-Toolkit 0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/). The genome was assembled to 26.9-fold coverage with SPAdes v3.5.0, with default parameters, and closed by PCR  and Sanger sequencing  (\u201314\u2013Matapan was collected from filtered (0.2-\u03bcm pore size) municipal wastewater in College Station, TX, via enrichment on y method . Phage my method . The geny method . Librarixy-pub/) , 9. Genexy-pub/) \u201312. The y-pub/) \u2013. Predictub/) \u201314\u2013. Whole-gub/) \u201314\u2013.Salmonella phage Vi01 (GenBank accession number FQ312032), Escherichia phage ECML-4 (GenBank accession number JX128257), Salmonella phage STML-13-1 (GenBank accession number JX181828), and Salmonella phage Marshall (GenBank accession number KF669653), respectively, and 196 similar proteins with phage Vi01. Similar to other Vi01-like phages, Matapan is predicted to use headful or pac-type packaging using PhageTerm (rIIA-like gene.Matapan is a Vi01-like myophage with a 157,408-kb genome with a G+C content of 44.9%, a 93.0% coding density, 214 protein-coding genes, and 3 tRNAs. As determined by progressiveMauve, Matapan shares nucleotide similarities of 91.3%, 89.5%, 88.2%, and 87.9% with MN066127, BioProject accession number PRJNA222858, SRA accession number SRR8869240, and BioSample accession number SAMN11360418.The genome sequence and associated data for phage Matapan were deposited under GenBank accession number"}
+{"text": "T was isolated from the gut microbiota of a 28-year-old woman and exhibits a 2.23-Mb  draft genome sequence containing 1,902 protein-coding genes, 49 tRNAs, and 3 rRNAs. The 16S rRNA sequencing and in silico DNA-DNA hybridization indicated that strain Marseille-P9525T represents a new species to be described.Strain Marseille-P9525 T was isolated from the gut microbiota of a 28-year-old woman and exhibits a 2.23-Mb  draft genome sequence containing 1,902 protein-coding genes, 49 tRNAs, and 3 rRNAs. The 16S rRNA sequencing and in silico DNA-DNA hybridization indicated that strain Marseille-P9525T represents a new species to be described.Strain Marseille-P9525 Enorma massiliensis was the first species of the genus Enorma recently described within the family Coriobacteriaceae . Here, we report the genome sequence of strain Marseille-P9525T and describe its genomic content.eriaceae . The genmosphere \u20133. A culosphere \u2013 of the sT was subcultured on Colombia agar incorporating 5% sheep blood  at 37\u00b0C under anaerobic conditions . DNA was extracted in 50 \u03bcl using the EZ1 BioRobot and EZ1 DNA tissue kit  and was quantified using a Qubit assay with a high-sensitivity kit  to 0.2\u2009ng/liter. DNA was then sequenced using MiSeq technology  with paired-end applications. The DNA was fragmented and amplified by limited PCR (12 cycles), introducing dual-index barcodes and sequencing adapters. After purification on AMPure XP beads , libraries were normalized and pooled for MiSeq paired-end sequencing. An automated cluster generation of 12 runs with dual-indexed 2\u2009\u00d7\u2009251-bp reads was performed for 40 hours. A total of 7,810,996 paired-end reads with a 35 to 251-bp sequence length were quality checked using FastQC, trimmed using Trimmomatic version 0.36.6 , 96.76% identity with E. timonensis (NR_144707), 95.30% with E. massiliensis (NR_125606), 94.03% with Collinsella tanakaei (NR_112899), and 93.96% with Collinsella ihuae (LN881598).Strain Marseille-P9525T is smaller than those of E. phocaeensis, E. massiliensis, E. timonensis, C. tanakaei, and C. ihuae . Based on the 16S rRNA gene sequence proximity, genomes were selected and incorporated into an in silico genomic comparison. Genomic similarities estimated using OrthoANI version 0.93.1 (https://www.ezbiocloud.net/tools/orthoani) and in silico DNA-DNA hybridization (http://ggdc.dsmz.de/ggdc.php#) estimated using the GGDC version 2.0 online tool yielded 84.20% and 28.4% sequence similarity with E. phocaeensis, 80.77% and 25.3% with E. timonensis, 78.68% and 24.9% with E. massiliensis, 77.35% and 23.7% with C. ihuae, and 74.57% and 22.2% with C. tanakaei, respectively.The 2.23-Mb draft genome sequence of strain Marseille-P9525CABFVT000000000 and ERR3427549, respectively.The genome sequence reported here and the raw reads have been deposited at EMBL/GenBank under the accession numbers"}
+{"text": "Vibrio parahaemolyticus is the lead causative agent for seafood-borne human gastroenteritis. While its occurrence has traditionally been uncommon in Europe and the United Kingdom, rising sea surface temperatures have resulted in an increased prevalence. Here, we present the complete genome sequences of four novel V. parahaemolyticus strains isolated in the United Kingdom. Vibrio parahaemolyticus is the lead causative agent for seafood-borne human gastroenteritis. While its occurrence has traditionally been uncommon in Europe and the United Kingdom, rising sea surface temperatures have resulted in an increased prevalence. Here, we present the complete genome sequences of four novel V. parahaemolyticus strains isolated in the United Kingdom. Vibrio parahaemolyticus is a ubiquitous marine bacterium and an important causative agent for human gastroenteritis  EXE V18/004 from Ostrea edulis at Chichester Harbor (2018), (ii) V12/024 from Crassostrea gigas at Weymouth (2012), (iii) V05/313 from Eriocheir sinensis in the River Thames (2005), and (iv) V05/027 from an unknown shellfish source in Southampton (2005). Strains EXE V18/004 and EXE V13/004 were isolated at the University of Exeter, while strains V12/024 and V05/027 were isolated at Cefas Weymouth Laboratories. All four of these strains were isolated directly from shellfish following previously described methods V. parahaemolyticus was initially identified based on colony morphology on selective agar (marine agar and thiosulphate citrate bile sucrose agar) and by PCR targeting the toxR region (9 cells\u2009\u00b7\u2009ml\u22121) was added directly to a Qiagen DNeasy PowerWater kit (Germany) for DNA extraction, followed by library preparation for short-read  and long-read  sequencing. For short-read libraries, DNA was quantified in triplicate with the Quant-iT double-stranded DNA (dsDNA) high-sensitivity (HS) assay in an Ependorff AF2200 plate reader. Genomic DNA libraries were prepared using a Nextera XT library prep kit (Illumina) following the manufacturer\u2019s protocol with the following modifications: 2 ng of DNA instead of 1 ng was used as the input, and PCR elongation time was increased from 30\u2009s to 1\u2009min. DNA quantification and library preparation were carried out on a Hamilton Microlab Star automated liquid handling system. Pooled libraries were quantified using the Kapa Biosystems library quantification kit for Illumina on a Roche LightCycler 96 quantitative PCR (qPCR) machine. Libraries were sequenced on an Illumina HiSeq instrument. Long-read sequencing was performed in-house using a multiplexed SQK-LSK108 library preparation and sequenced on a FLO-MIN106 flow cell, following Oxford Nanopore Technologies protocol (https://github.com/rrwick/Porechop) with the following settings: -require_two_barcodes -discard_unassigned -discard_middle. Hybrid genome assembly was performed using Unicycler v0.4.7 (https://sourceforge.net/projects/bbmap/) with the following settings: idfilter=0.95 covstats=covstats.txt. Long-read coverage was calculated by first using minimap2 v.2.17 (https://github.com/lh3/minimap2) and SAMtools v.1.9 (http://samtools.sourceforge.net/) to create a bam file (minimap2 -t 16 -ax map-ont <assembly> <long_reads> | samtools view -F 4 -buS | samtools sort -o long.sorted.bam) and then using BBMap\u2019s pileup.sh to calculate coverage (pileup.sh in=long.sorted.bam ref=ref.fa out=long.covstats.txt). Default parameters for all software were used unless otherwise noted.R region . IsolateR region  at 37\u00b0C protocol . Short rprotocol  with a sr v0.4.7 , and eacr v0.4.7 , developEach strain was found to have two chromosomes , one withttps://img.jgi.doe.gov/) using the following taxon identifiers (IDs): 2816332655 (V. parahaemolyticus EXE V18/004), 2816332656 (V. parahaemolyticus V12/024), 2816332657 (V. parahaemolyticus V05/313), and 2816332658 (V. parahaemolyticus V05/027). Read data are available from the European Nucleotide Archive under the following strain names and accession numbers: V. parahaemolyticus EXE V18/004, ERS3342146; V. parahaemolyticus V12/024, ERS3342147; V. parahaemolyticus V05/313, ERS3342148; and V. parahaemolyticus V05/027, ERS3342149.Assembled and annotated genomes are publicly available from JGI IMG/M ("}
+{"text": "In sub-Saharan Africa, individuals infected with HIV who are severely immunocompromised have high mortality (about 10%) shortly after starting antiretroviral therapy (ART). This group also has the greatest risk of morbidity and mortality associated with immune reconstitution inflammatory syndrome (IRIS), a paradoxical response to successful ART. Integrase inhibitors lead to significantly more rapid declines in HIV viral load (VL) than all other ART classes. We hypothesised that intensifying standard triple-drug ART with the integrase inhibitor, raltegravir, would reduce HIV VL faster and hence reduce early mortality, although this strategy could also risk more IRIS events.3, from eight urban/peri-urban HIV clinics at regional hospitals in Kenya, Malawi, Uganda, and Zimbabwe were randomised 1:1 to initiate standard triple-drug ART, with or without 12-week raltegravir intensification, and followed for 48 weeks. The primary outcome was 24-week mortality, analysed by intention to treat. Of 2,356 individuals screened for eligibility, 1,805 were randomised between 18 June 2013 and 10 April 2015. Of the 1,805 participants, 961 (53.2%) were male, 72 (4.0%) were children/adolescents, median age was 36 years, CD4 count was 37 cells/mm3, and plasma viraemia was 249,770 copies/mL. Fifty-six participants (3.1%) were lost to follow-up at 48 weeks. By 24 weeks, 97/902 (10.9%) raltegravir-intensified ART versus 91/903 (10.2%) standard ART participants had died , with no evidence of interaction with other randomisations (pheterogeneity > 0.7) and despite significantly greater VL suppression with raltegravir-intensified ART at 4 weeks  and 12 weeks . Through 48 weeks, there was no evidence of differences in mortality ; in serious , grade-4 , or ART-modifying  adverse events ; in events judged compatible with IRIS  or in hospitalisations . At 12 weeks, one and two raltegravir-intensified participants had predicted intermediate-level and high-level raltegravir resistance, respectively. At 48 weeks, the nucleoside reverse transcriptase inhibitor (NRTI) mutation K219E/Q (p = 0.004) and the non-nucleoside reverse transcriptase inhibitor (NNRTI) mutations K101E/P (p = 0.03) and P225H (p = 0.007) were less common in virus from participants with raltegravir-intensified ART, with weak evidence of less intermediate- or high-level resistance to tenofovir (p = 0.06), abacavir (p = 0.08), and rilpivirine (p = 0.07). Limitations of the study include limited clinical, radiological, and/or microbiological information for some participants, reflecting available services at the centres, and lack of baseline genotypes.In a 2\u00d72\u00d72 factorial open-label parallel-group trial, treatment-naive adults, adolescents, and children >5 years old infected with HIV, with cluster of differentiation 4 (CD4) <100 cells/mmAlthough 12 weeks of raltegravir intensification was well tolerated and reduced HIV viraemia significantly faster than standard triple-drug ART during the time of greatest risk for early death, this strategy did not reduce mortality or clinical events in this group and is not warranted. There was no excess of IRIS-compatible events, suggesting that integrase inhibitors can be used safely as part of standard triple-drug first-line therapy in severely immunocompromised individuals.NCT01825031.ClinicalTrials.gov ISRCTN 43622374.International Standard Randomised Controlled Trials Number  In the factorial randomised REALITY trial, Sarah Walker and colleagues document outcomes of intensified initial antiretroviral treatment for people with advanced HIV disease. 3) have high risks of dying shortly after starting treatment.Individuals in Africa who are HIV positive and initiating treatment with severe immunosuppression  to standard triple-drug antiretroviral therapy for 12 weeks would reduce HIV viral load faster: we wanted to find out whether this would reduce high early mortality.We randomly allocated 1,805 adults, adolescents, and older children to receive standard antiretroviral therapy with or without 12 weeks of adjunctive raltegravir.We found that the group receiving 12 weeks of adjunctive raltegravir had significantly faster declines in HIV viral load.However, we found no significant difference in deaths within 24 weeks of starting treatment between those receiving adjunctive raltegravir (97/902 [10.9%]) or not (91/903 [10.2%]); nor were there differences in clinical disease progression, immune reconstitution, inflammatory syndrome, or adverse events.In this study, we found that intensifying the potency of initial antiretroviral therapy did not reduce early mortality on treatment, providing no support for its widespread use in individuals initiating treatment with severe immunosuppression.However, it also did not increase the rates of clinically important immune reconstitution inflammatory syndrome, suggesting that integrase inhibitors could replace other components of standard first-line antiretroviral therapy safely. Absolute 24-week mortality difference was +0.6% . By 48 weeks, 110 (12.4%) versus 115 (13.0%) participants, respectively, had died  with no evidence of differences by cause (Table 2). There was no difference in longer-term (after week 48) mortality . Estimated mortality rates decreased sharply from day 19 through week 12 , with 71 (31.6%) of the 225 deaths occurring by 4 weeks, and 147 (65.3%) by 12 weeks. There was no evidence of early differences in mortality rates  and no evidence of heterogeneity in the impact of raltegravir intensification over time on ART . Of nine preplanned and five exploratory subgroup analyses . Specifically, there was no evidence that mortality differences depended on pre-ART VL or pre-ART CD4 . No subgroup analyses suggested heterogeneity in the impact of raltegravir intensification on suppression <50 copies/mL at week 4 (pheterogeneity > 0.05) .By 24 weeks (primary endpoint), 97 (10.9%) raltegravir-intensified ART versus 91 (10.2%) standard ART participants died .There was no evidence of an impact of raltegravir intensification on any disease progression clinical outcome (S2 Text). One participant had predicted intermediate-level (mutations T97A+R263K) and two predicted high-level  raltegravir resistance mutations . No patient had predicted intermediate- or high-level dolutegravir resistance.Integrase genotypes were obtained in 33 raltegravir-intensified ART participants with VL >1,000  copies/mL at 12 weeks (see p = 0.004) and the NNRTI mutations K101E/P (p = 0.03) and P225H (p = 0.007) were less common in raltegravir-intensified ART, with no evidence of differences for other mutations . There was marginal evidence suggesting less intermediate- or high-level resistance with raltegravir-intensified ART to tenofovir , abacavir , and rilpivirine    copies/mL at 48 weeks. The NRTI mutation K219E/Q ; however, there was marginal evidence of a small difference at 48 weeks . There was similar weak evidence of slightly (2%\u20135%) greater percentages with CD4 \u2265 200 cells/mm3 at later time points . Changes in weight were also similar through week 24 , with small but significant differences appearing from 36\u201348 weeks. Similar late differences were observed for BMI in adolescents/adults (data not shown), fat mass , and muscle mass . There was no evidence of heterogeneity in these differences by age . Absolute CD8 count increases were similar in both groups through 48 weeks .Absolute CD4 count increases were similar in both groups through 24 weeks , grade-4 AEs (log-rank p = 0.29), grade-3/4 AEs (log-rank p = 0.72), grade-4 AEs definitely/probably/possibly related to ART (log-rank p = 0.52), grade-4 AEs definitely/probably related to ART (log-rank p = 0.07), AEs leading to ART modification (log-rank p = 0.51) or new hospitalisations (log-rank p = 0.59) (Table 2). Raltegravir was modified for AEs in 19 (2.1%) participants (S2 Table), including renal (n = 6) events, hepatic (n = 6) events, hypersensitivity reactions , and Stevens-Johnson syndrome in one participant . Only one participant experienced a grade-4 AE adjudicated as definitely/probably related to raltegravir (Table 2). There was no evidence of any difference in total hospitalisation-days  or total hospitalisations .There was no evidence for differences in time to first SAEs (log-rank p = 0.79) (Table 2); 36 (4.0%) versus 31 (3.4%), respectively, experienced fatal IRIS events (p = 0.54). Tuberculosis IRIS events occurred in 53 (5.9%) participants with raltegravir-intensified ART versus 54 (6.0%) with standard ART (exact p = 1.00) and cryptococcal IRIS events in 15 (1.7%) versus 16 (1.8%) participants, respectively (exact p = 1.00) (S3 Table). IRIS events occurred a median 3.4 (IQR 2.0\u20136.3) weeks from randomisation, with rates declining from the third week on ART . IRIS was more common in participants initiating ART at older ages (p = 0.005), with lower CD4 counts (p < 0.001) or with pre-existing TB (p = 0.007); IRIS was less common in those initiating ART with enhanced prophylaxis against opportunistic infections (p = 0.001) (S4 Table). There was no evidence that raltegravir intensification was associated with IRIS after adjusting for these factors (p = 0.63).Fatal or nonfatal events judged compatible with IRIS occurred in 89 (9.9%) raltegravir-intensified ART versus 86 (9.5%) standard ART participants . However, the differential suppression occurred between weeks 4 and 12, with overall only 15.8% less time spent with VL <50 c/mL in the standard ART group through week 24 . Therefore, benefits at a population level would likely be modest at best. However, such a strategy might have particular value in swiftly reducing VL in women identified as HIV-infected during pregnancy to reduce mother-to-child transmission .S2B Fig). Association with twice-daily dosing, rather than pill burden, is supported by the lack of difference in self-reported ART adherence between participants randomised in this trial to enhanced-prophylaxis versus standard-of-care co-trimoxazole  .The full trial protocol can be accessed at (PDF)Click here for additional data file.S1 Text(DOC)Click here for additional data file.S2 Text(DOC)Click here for additional data file.S1 TableSAE, serious adverse event.(DOC)Click here for additional data file.S2 TableAE, adverse event.(DOC)Click here for additional data file.S3 TableIRIS, immune reconstitution inflammatory syndrome.(DOC)Click here for additional data file.S4 TableIRIS, immune reconstitution inflammatory syndrome.(DOC)Click here for additional data file.S5 TableCD4, cluster of differentiation 4.(DOC)Click here for additional data file.S1 Fig(TIF)Click here for additional data file.S2 FigSelf-reported percentage reporting missing doses of any ART in the last 4 weeks (a) overall and (b) by backbone NRTI and NNRTI frequency. ART, antiretroviral therapy; NNRTI, non-nucleoside reverse transcriptase inhibitor; NRTI, nucleoside reverse transcriptase inhibitor.(TIF)Click here for additional data file.S3 FigVL suppression (a) <400 copies/mL and (b) <1,000 copies/mL. VL, viral load.(TIF)Click here for additional data file.S4 Fig(TIF)Click here for additional data file.S5 FigART, antiretroviral therapy.(TIF)Click here for additional data file.S6 Fig(TIF)Click here for additional data file.S7 FigVL, viral load.(TIF)Click here for additional data file.S8 FigNNRTI, non-nucleoside reverse transcriptase inhibitor; NRTI, nucleoside reverse transcriptase inhibitor; VL, viral load.(TIF)Click here for additional data file.S9 FigVL, viral load.(TIF)Click here for additional data file.S10 FigCD4, cluster of differentiation 4.(TIF)Click here for additional data file.S11 FigChanges in body composition, (a) fat mass and (b) muscle mass.(TIF)Click here for additional data file.S12 FigChanges in (a) CD4 cell count and (b) weight in children/adolescents (5\u201317 years) versus adults (18 years or older) at ART initiation. ART, antiretroviral therapy; CD4, cluster of differentiation 4.(TIF)Click here for additional data file.S13 FigCD8, cluster of differentiation 8.(TIF)Click here for additional data file.S14 FigIRIS, immune reconstitution inflammatory syndrome.(TIF)Click here for additional data file."}
+{"text": "Diet plays a crucial role in sculpting microbial communities. Similar diets appear to drive convergence of gut microbial communities between host species. Captivity usually provides an identical diet and environment to different animal species that normally have similar diets. Whether different species\u2019 microbial gut communities can be homogenized by a uniform diet in captivity remains unclear.Rhinolophus ferrumequinum, Vespertilio sinensis, and Hipposideros armiger) in captivity and in the wild using 16S rDNA sequencing. In captivity, R. ferrumequinum and V. sinensis were fed yellow mealworms, while H. armiger was fed giant mealworms to rule out the impact of an identical environment on the species\u2019 gut microbial communities.In this study, we compared gut microbial communities of three insectivorous bat species (R. ferrumequinum and V. sinensis in captivity clustered together. All microbial functions found in captive V. sinensis were shared by R. ferrumequinum. Moreover, the relative abundances of all metabolism related KEGG pathways did not significantly differ between captive R. ferrumequinum and V. sinensis; however, the relative abundance of \u201cGlycan Biosynthesis and Metabolism\u201d differed significantly between wild R. ferrumequinum and V. sinensis.We found that the microbial communities of the bat species we studied clustered by species in the wild, while the microbial communities of Our results suggest that consuming identical diets while in captivity tends to homogenize the gut microbial communities among bat species. This study further highlights the importance of diet in shaping animal gut microbiotas. Trillions of microorganisms reside in animal guts, and these microorganisms constitute the animal\u2019s gut microbiota, which is important for animal health . Ley et Wild animals in captivity are usually housed under uniform conditions that include identical diets and environments . This reRhinolophus ferrumequinum), the Asian parti-colored bat (Vespertilio sinensis) and the great Himalayan leaf-nosed bat (Hipposideros armiger) in the wild and in captivity. We then compared the bacterial communities in both the wild and captive samples between these three species. We captured bats in the wild, brought them back to the laboratory and housed them in identical environments but provided different food. Rhinolophus ferrumequinum and V.\u00a0sinensis were fed the same food , while H.\u00a0armiger were provided giant mealworms, thus forming a comparison to eliminate the impact of environment on the gut microbiome. Given that diet strongly influences microbiome composition and similar diets appear to drive convergence of gut microbial communities between host species  are the second largest mammalian group . Most ba species , we predosystems . Microbiosystems . Thus, mRhinolophus ferrumequinum feeds preferentially on lepidopterans, particularly the noctuid species, which constitute approximately 41% of the bat\u2019s diet (V.\u00a0sinensis mainly comprises Lepidoptera (mean relative percentage: 32.8%), Diptera (27.5%) and Coleoptera (22.6%), but the proportion of each order varies seasonally , while the great leaf-nosed bats were fed giant mealworms (Zophobas morio) for comparison. We kept the bats for 4\u20136 months, and collected their fecal pellets less than 15\u00a0min after defecationin the laboratory. Each bat\u2019s sex, age, weight, forearm length, and reproductive condition was recorded  and the Forestry Bureau of Jilin Province, China  approved all study protocols.\u00aeStool DNA Kit  per the manufacturer\u2019s instructions and stored at \u221220\u00a0\u00b0C for further analysis. Extracted DNA was measured using a NanoDrop NC2000 spectrophotometer  and agarose gel electrophoresis to estimate DNA quantity and quality, respectively.Fifty-three fecal samples were used, including 23 from the wild bats and 30 from the captive bats. DNA was extracted from all fecal samples using the E.Z.N.A.2O. The PCR conditions consisted of initial denaturation at 98\u00a0\u00b0C for 2 min, followed by 25 denaturation cycles at 98\u00a0\u00b0C for 15 s, annealing at 55 \u00a0\u00b0C for 30 s, extension at 72\u00a0\u00b0C for 30 s, and a final extension at 72\u00a0\u00b0C for 5 min. PCR products were purified with Agencourt AMPure Beads  and quantified using the PicoGreen dsDNA Assay Kit . The individual PCR products were then pooled in equal amount, and sequenced using the paired-end 2\u00a0\u00d7300 bp method on the Illumina MiSeq platform with MiSeq Reagent Kit v3 at Shanghai Personal Biotechnology Co., Ltd. . All raw sequences were deposited into the NCBI Sequence Read Archive under accession numbers SRR8238420\u2013SRR8238472.The V3-V4 region of the bacterial 16S rRNA genes were amplified via PCR using the forward primer, 338F (5\u2032-ACTCCTACGGGAGGCAGCA-3\u2032), and the reverse primer, 806R (5\u2032-GGACTACHVGGGTWTCTAAT-3\u2032) . Sample-Sequencing data were processed using the Quantitative Insights Into Microbial Ecology  . Brieflyt-test and the Monte Carlo permutation test with 1,000 permutations, then visualized using box-and-whiskers plots. For UniFrac distance-based pairwise comparisons among groups, we used a very conservative Bonferroni post-hoc correction method to perform the multiple corrections and evaluate the significance of the comparison. Permutational multivariate analysis of variance (PERMANOVA)   and analRMANOVA)  were conRMANOVA)  to assesRMANOVA)  based onRMANOVA) . MicrobiRMANOVA)  in the KRMANOVA)  based onRMANOVA) . ResultsA total of 768,990 and 1,466,150 16S rDNA sequences were obtained from the microbiomes of the 23 wild and 30 captive bats, respectively, and the average sequence numbers per sample were 33,434 and 48,872, respectively. Rarefaction analysis demonstrated that the sequencing depth was sufficient for each sample . A totalV. sinensis and R. ferrumequinum were shared, but only 18% were shared by these two species in the wild. The proportions of OTUs shared by V. sinensis and H. armiger were approximately 39% and 12% in captivity and the wild, respectively. Minimal difference was noted between the proportions of shared OTUs in the captive and wild R. ferrumequinum and H. armiger, of which, the proportions were nearly 36% and 29%, respectively.Venn diagrams were plotted to visualize the shared and unique OTUs  among three species of wild and captive bats. The captive bats we sampled shared more OTUs than did the wild bats . A totalV.\u00a0sinensis and R. ferrumequinum fed yellow mealworms) clustered together, while the gut microbial communities of H. armiger, fed giant mealworms, clustered alone. A PCoA based on unweighted UniFrac distances also demonstrated similar clustering results using NMDS based on unweighted UniFrac distances. In the wild bats, PC1, PC2 and PC3 accounted for nearly 42% of the variation, and samples were separated roughly by bat species   , Table 2 samples , Table 2H. armiger were shared by the other two species, and all microbial functions in captive V. sinensis were shared by R. ferrumequinum. Thus, in terms of presence/absence, the microbial functions appeared converged in the captive bats.Finally, we predicted the microbial functions of wild and captive bats using PICRUSt, which yielded 5,971 and 4,771 KEGG pathways, respectively. Venn diagrams showed that in total 5,495 KEGG pathways were shared among the wild bat samples, and 3,964 were shared among the captive bat samples . Unlike R. ferrumequinum and V. sinensis, while the relative abundance of \u201cGlycan Biosynthesis and Metabolism\u201d differed significantly between the wild R. ferrumequinum and V. sinensis  fed the same food converged markedly, while they differed from those of H. armiger fed different food. This result highlighted the importance of diet on gut microbial communities. Diet shapes the gut microbiota by providing substrates that differentially support or enhance specific microbial growth  than were R. ferrumequinum and V. sinensis, and the gut microbial communities of captive H. armiger did not converge with the other two bat species. This result eliminated the impact of environment on the gut microbial communities because the three bat species were housed in identical environments in the laboratory. Further, this suggested that identical diets contribute to microbial community convergence in various bat species. However, fecal samples usually include bacteria that are ingested with the food , and distinguishing these bacteria from the host-derived bacteria in the fecal microbiome is difficult. Thus, the gut microbiota in this study did not specifically refer to the host-derived bacteria. The microbial community compositions between species fed uniform diets may have converged due to changes in the compositions of the host-associated gut microbiome or a much larger shared component of the fecal microbiome based on a completely shared diet comprising mealworms and their commensal bacteria or both. Our results highlight the need for future studies to address this issue, for example, incorporating dietary classifications via metabarcoding and classifying the microbiomes of the invertebrate prey. In addition, our results differed from those obtained by Comparing the microbial communities in fecal samples from three captive bats species showed that the microbial compositions of two bat species  Wild bats (B) Captive bats. Points are means \u00b1 SE, with the numbers of bats per group shown in Click here for additional data file.10.7717/peerj.6844/supp-2Figure S2Wild (A) and captive (B) bats\u2019 fecal bacterial communities clustered using principal coordinates analysis. Each point corresponds to a fecal sample colored according to bat species with different symbols corresponding to host family .Click here for additional data file.10.7717/peerj.6844/supp-3Figure S3X-axis, pairwise comparisons among groups. Y-axis, UniFrac distances. Box borders represent upper and lower interquartile ranges. Red lines, whiskers, and \u201c+\u201d represent the median values, 1.5 times the interquartile range beyond upper and lower quartiles, and outliers respectively. Significant differences in the UniFrac distances for pairwise comparisons among groups are shown in Click here for additional data file.10.7717/peerj.6844/supp-4Figure S4V. sinensis, R. ferrumequinum and H. armiger collected from the wild respectively.WFVs, WFRf and WFHa represent fecal samples from Click here for additional data file.10.7717/peerj.6844/supp-5Figure S4V. sinensis, R. ferrumequinum and H. armiger respectively.FVs, FRf and FHa represent fecal samples from captive Click here for additional data file.10.7717/peerj.6844/supp-6Data S1Sex, age, weight, forearm length, and sample site information of each bat.Click here for additional data file.10.7717/peerj.6844/supp-7Data S2raw data of FHa1.Click here for additional data file.10.7717/peerj.6844/supp-8Data S3raw data of FHa1.Click here for additional data file.10.7717/peerj.6844/supp-9Data S4raw data of FHa2.Click here for additional data file.10.7717/peerj.6844/supp-10Data S5raw data of FHa2.Click here for additional data file.10.7717/peerj.6844/supp-11Data S6raw data of FHa3.Click here for additional data file.10.7717/peerj.6844/supp-12Data S7raw data of FHa3.Click here for additional data file.10.7717/peerj.6844/supp-13Data S8raw data of FHa4.Click here for additional data file.10.7717/peerj.6844/supp-14Data S9raw data of FHa4.Click here for additional data file.10.7717/peerj.6844/supp-15Data S10raw data of FHa5.Click here for additional data file.10.7717/peerj.6844/supp-16Data S11raw data of FHa5.Click here for additional data file.10.7717/peerj.6844/supp-17Data S12raw data of FHa6.Click here for additional data file.10.7717/peerj.6844/supp-18Data S13raw data of FHa6.Click here for additional data file.10.7717/peerj.6844/supp-19Data S14raw data of FHa7.Click here for additional data file.10.7717/peerj.6844/supp-20Data S15raw data of FHa7.Click here for additional data file.10.7717/peerj.6844/supp-21Data S16raw data of FHa8.Click here for additional data file.10.7717/peerj.6844/supp-22Data S17raw data of FHa8.Click here for additional data file.10.7717/peerj.6844/supp-23Data 18raw data of FHa9.Click here for additional data file.10.7717/peerj.6844/supp-24Data S19raw data of FHa9.Click here for additional data file.10.7717/peerj.6844/supp-25Data S20Raw data of FHa10.Click here for additional data file.10.7717/peerj.6844/supp-26Data S21Raw data of FHa10.Click here for additional data file.10.7717/peerj.6844/supp-27Data S22Raw data of FRf1.Click here for additional data file.10.7717/peerj.6844/supp-28Data S23Raw data of FRf1.Click here for additional data file.10.7717/peerj.6844/supp-29Data S24Raw data of FRf2.Click here for additional data file.10.7717/peerj.6844/supp-30Data S25Raw data of FRf2.Click here for additional data file.10.7717/peerj.6844/supp-31Data S26Raw data of FRf3.Click here for additional data file.10.7717/peerj.6844/supp-32Data S27Raw data of FRf3.Click here for additional data file.10.7717/peerj.6844/supp-33Data S28Raw data of FRf4.Click here for additional data file.10.7717/peerj.6844/supp-34Data S29Raw data of FRf4.Click here for additional data file.10.7717/peerj.6844/supp-35Data S30Raw data of FRf5.Click here for additional data file.10.7717/peerj.6844/supp-36Data S31Raw data of FRf5.Click here for additional data file.10.7717/peerj.6844/supp-37Data S32Raw data of FRf6.Click here for additional data file.10.7717/peerj.6844/supp-38Data S33Raw data of FRf6.Click here for additional data file.10.7717/peerj.6844/supp-39Data S34Raw data of FRf7.Click here for additional data file.10.7717/peerj.6844/supp-40Data S35Raw data of FRf7.Click here for additional data file.10.7717/peerj.6844/supp-41Data S36Raw data of FRf8.Click here for additional data file.10.7717/peerj.6844/supp-42Data S37Raw data of FRf8.Click here for additional data file.10.7717/peerj.6844/supp-43Data S38Raw data of FRf9.Click here for additional data file.10.7717/peerj.6844/supp-44Data S39Raw data of FRf9.Click here for additional data file.10.7717/peerj.6844/supp-45Data S40Raw data of FRf10.Click here for additional data file.10.7717/peerj.6844/supp-46Data S41Raw data of FRf10.Click here for additional data file.10.7717/peerj.6844/supp-47Data S42Raw data of FVs1.Click here for additional data file.10.7717/peerj.6844/supp-48Data S43Raw data of FVs1.Click here for additional data file.10.7717/peerj.6844/supp-49Data S44Raw data of FVs2.Click here for additional data file.10.7717/peerj.6844/supp-50Data S45Raw data of FVs2.Click here for additional data file.10.7717/peerj.6844/supp-51Data S46Raw data of FVs3.Click here for additional data file.10.7717/peerj.6844/supp-52Data S47Raw data of FVs3.Click here for additional data file.10.7717/peerj.6844/supp-53Data S48Raw data of FVs4.Click here for additional data file.10.7717/peerj.6844/supp-54Data S49Raw data of FVs4.Click here for additional data file.10.7717/peerj.6844/supp-55Data S50Raw data of FVs5.Click here for additional data file.10.7717/peerj.6844/supp-56Data S51Raw data of FVs5.Click here for additional data file.10.7717/peerj.6844/supp-57Data S52Raw data of FVs6.Click here for additional data file.10.7717/peerj.6844/supp-58Data S53Raw data of FVs6.Click here for additional data file.10.7717/peerj.6844/supp-59Data S54Raw data of FVs7.Click here for additional data file.10.7717/peerj.6844/supp-60Data S55Raw data of FVs7.Click here for additional data file.10.7717/peerj.6844/supp-61Data S56Raw data of FVs8.Click here for additional data file.10.7717/peerj.6844/supp-62Data S57Raw data of FVs8.Click here for additional data file.10.7717/peerj.6844/supp-63Data S58Raw data of FVs9.Click here for additional data file.10.7717/peerj.6844/supp-64Data S59Raw data of FVs9.Click here for additional data file.10.7717/peerj.6844/supp-65Data S60Raw data of FVs10.Click here for additional data file.10.7717/peerj.6844/supp-66Data S61Raw data of FVs10.Click here for additional data file.10.7717/peerj.6844/supp-67Data S62Raw data of WFHa1.Click here for additional data file.10.7717/peerj.6844/supp-68Data S63Raw data of WFHa1.Click here for additional data file.10.7717/peerj.6844/supp-69Data S64Raw data of WFHa2.Click here for additional data file.10.7717/peerj.6844/supp-70Data S65Raw data of WFHa2.Click here for additional data file.10.7717/peerj.6844/supp-71Data S66Raw data of WFHa3.Click here for additional data file.10.7717/peerj.6844/supp-72Data S67Raw data of WFHa3.Click here for additional data file.10.7717/peerj.6844/supp-73Data S68Raw data of WFHa4.Click here for additional data file.10.7717/peerj.6844/supp-74Data S69Raw data of WFHa4.Click here for additional data file.10.7717/peerj.6844/supp-75Data S70Raw data of WFHa7.Click here for additional data file.10.7717/peerj.6844/supp-76Data S71Raw data of WFHa7.Click here for additional data file.10.7717/peerj.6844/supp-77Data S72Raw data of WFHa8.Click here for additional data file.10.7717/peerj.6844/supp-78Data S73Raw data of WFHa8.Click here for additional data file.10.7717/peerj.6844/supp-79Data S74Raw data of WFHa9.Click here for additional data file.10.7717/peerj.6844/supp-80Data S75Raw data of WFHa9.Click here for additional data file.10.7717/peerj.6844/supp-81Data S76Raw data of WFHa10.Click here for additional data file.10.7717/peerj.6844/supp-82Data S77Raw data of WFHa10.Click here for additional data file.10.7717/peerj.6844/supp-83Data S78Raw data of WFRf1.Click here for additional data file.10.7717/peerj.6844/supp-84Data S79Raw data of WFRf1.Click here for additional data file.10.7717/peerj.6844/supp-85Data S80Raw data of WFRf14.Click here for additional data file.10.7717/peerj.6844/supp-86Data S81Raw data of WFRf14.Click here for additional data file.10.7717/peerj.6844/supp-87Data S82Raw data of WFRf24.Click here for additional data file.10.7717/peerj.6844/supp-88Data S83Raw data of WFRf24.Click here for additional data file.10.7717/peerj.6844/supp-89Data S84Raw data of WFRf25.Click here for additional data file.10.7717/peerj.6844/supp-90Data S85Raw data of WFRf25.Click here for additional data file.10.7717/peerj.6844/supp-91Data S86Raw data of WFRf26.Click here for additional data file.10.7717/peerj.6844/supp-92Data S87Raw data of WFRf26.Click here for additional data file.10.7717/peerj.6844/supp-93Data S88Raw data of WFRf27.Click here for additional data file.10.7717/peerj.6844/supp-94Data S89Raw data of WFRf27.Click here for additional data file.10.7717/peerj.6844/supp-95Data S90Raw data of WFRf28.Click here for additional data file.10.7717/peerj.6844/supp-96Data S91Raw data of WFRf28.Click here for additional data file.10.7717/peerj.6844/supp-97Data S92Raw data of WFRf29.Click here for additional data file.10.7717/peerj.6844/supp-98Data S93Raw data of WFRf29.Click here for additional data file.10.7717/peerj.6844/supp-99Data S94Raw data of WFVs1.Click here for additional data file.10.7717/peerj.6844/supp-100Data S95Raw data of WFVs1.Click here for additional data file.10.7717/peerj.6844/supp-101Data S96Raw data of WFVs3.Click here for additional data file.10.7717/peerj.6844/supp-102Data S97Raw data of WFVs3.Click here for additional data file.10.7717/peerj.6844/supp-103Data S98Raw data of WFVs4.Click here for additional data file.10.7717/peerj.6844/supp-104Data S99Raw data of WFVs4.Click here for additional data file.10.7717/peerj.6844/supp-105Data S100Raw data of WFVs6.Click here for additional data file.10.7717/peerj.6844/supp-106Data S101Raw data of WFVs6.Click here for additional data file.10.7717/peerj.6844/supp-107Data S102Raw data of WFVs7.Click here for additional data file.10.7717/peerj.6844/supp-108Data S103Raw data of WFVs7.Click here for additional data file.10.7717/peerj.6844/supp-109Data S104Raw data of WFVs8.Click here for additional data file.10.7717/peerj.6844/supp-110Data S105Raw data of WFVs8.Click here for additional data file.10.7717/peerj.6844/supp-111Data S106Raw data of WFVs10.Click here for additional data file.10.7717/peerj.6844/supp-112Data 107Raw data of WFVs10.Click here for additional data file."}
+{"text": "Scientific Reports6: Article number: 2291110.1038/srep22911; published online: 03182016; updated: 08092018This Article contains typographical errors in Table 2 in the Multivariate logistic regression analysis section. The OR values for Platelet count, Glucose level, WBC count and C-reactive protein level are duplicated from their partial regression coefficients (\u03b2).The correct values should read 2.85, 1.14, 1.13, and 1.23 respectively."}
+{"text": "Correction to: Arthritis Res Therhttps://doi.org/10.1186/s13075-018-1790-xFollowing publication of the original article , the autn\u00a0=\u200925) and/or MRA .The sentence should read: Angiography was performed in all c-TA patients, including conventional angiography, CTA, and/or magnet resonance angiography (MRA) as the initial diagnostic modality in 36 (35.6%), 57 (56.4%), and 9 (8.9%), respectively, while 28 (27.7%) patients experienced additional catheter-based angiography for interventional therapy after TA diagnosis by CTA (24.7%,"}
+{"text": "N-arylation of azole compounds utilizing selective aryl-transfer TMP-iodonium (III) reagents . For the title compound, phenyliodonium(III) acetate (Ph(TMP)IOAc), the single-crystal X-ray diffraction measurement together with NMR analysis, like also the method of synthesis and crystallization are presented. The X-ray structure analysis has revealed that the two types of geometries regarding the acetate anion attached to phenyl (TMP)iodonium (III) cation are found in the crystal states.The data in this article are related to research article \u2018\u2018Efficient  The str22.1The solvents, starting materials, and reagents were purchased from Nacalai tesque and Tokyo Chemical Industry CO. Ltd.2.2Ph(TMP)IOAc was prepared according to our reported procedure 2.31H and 13C NMR spectra were recorded on an ECS 400 NMR spectrometer  at 400 MHz and 100 MHz, respectively, using CDCl3 as the solvent. The chemical shifts (\u03b4) are expressed in ppm relative to tetramethylsilane (TMS) as an internal standard. Coupling constants (J) are expressed in Hz. Signal multiplicities are represented as singlet (s), doublet (d), and triplet (t). Assignments of the proton and carbon positions in the compound were performed by PFG-HMQC and PFG-HMBC analyses.The 2.43, TMS standard. Concentration: 13 mg in 0.75 mL : \u03b4 1.95 , 3.83 , 6.12 , 7.29 , 7.41 , 7.92 . 13C NMR : \u03b4 24.6 (CH3COO), 55.7 (p-OMe), 56.5 (o-OMe), 90.9 (m-TMP), 91.0 (ipso-TMP), 119.4 (ipso-Ph), 130.3 (p-Ph), 130.8 (m-Ph), 133.8 (o-Ph), 160.6 (o-TMP), 165.7 (p-TMP), 178.8 (CH3COO).JEOL ECS 400 NMR spectrometer, solvent CDCl 0.75 mL , Fig.\u00a04.2.5The crystals were obtained at room temperature from chloroform/hexane mixture under a shading condition. Ph(TMP)IOAc was dissolved in chloroform and the insoluble material was removed by filtration. Hexane was added to the filtrate in sample bottle to reach the chloroform/hexane ratio 2/5. After standing for 1 day, the several crystals suitable for the X-ray structural analysis were obtained.2.6The single-crystal X-ray diffraction experiment was performed on the HPC diffractometer (Rigaku XtaLAB P200)). The two types of geometries for Ph(TMP)IOAc in a crystal state are shown in"}
+{"text": "Metarhizium anisopliae, chitindegradation is also used to breach the host cuticle during infection. In view ofthe putative role of NAGases as virulence factors, this study explored thetranscriptional profile and evolution of putative GH20 NAGases(MaNAG1 and MaNAG2) and GH3 NAGases(MaNAG3 and MaNAG4) identified inM. anisopliae. While MaNAG2 orthologs are conserved inseveral ascomycetes, MaNAG1 clusters only with Aspergilllus sp.and entomopathogenic fungal species. By contrast, MaNAG3 and MaNAG4 werephylogenetically related with bacterial GH3 NAGases. The transcriptionalprofiles of M. anisopliae NAGase genes were evaluated in sevenculture conditions showing no common regulatory patterns, suggesting that theseenzymes may have specific roles during the Metarhizium lifecycle. Moreover, the expression of MaNAG3 andMaNAG4 regulated by chitinous substrates is the firstevidence of the involvement of putative GH3 NAGases in physiological cellprocesses in entomopathogens, indicating their potential influence on celldifferentiation during the M. anisopliae life cycle.Cell walls are involved in manifold aspects of fungi maintenance. For severalfungi, chitin synthesis, degradation and recycling are essential processesrequired for cell wall biogenesis; notably, the activity of\u03b2-N-acetylglucosaminidases (NAGases) must be present for chitin utilization. Forentomopathogenic fungi, such as  Chitin is the second most abundant polymer on Earth and its recycling from carapaces,cuticles and fungal cell walls impacts on carbon and nitrogen cycles. The chitinpolymer is composed of \u03b2-1,4-linked N-acetyl-D-glucosamine (GlcNAc) subunits  and its NAGases are classified into three glycoside hydrolase (GH) families, 3, 20, and 84,on the basis of their amino acid sequence similarities 2-3 substrates. This report furthersupports the existence of GH3 NAGases in other fungal species, especially inascomycetes, considering their expansion of chitinolytic machinery genes . In these species, thechitinolytic system has, probably, two main biological functions: Firstly, as chitinis the major component of fungal cell walls, chitin-degrading enzymes act on thecell wall remodeling, which is necessary for hyphal vegetative growth , are processes that require chitin degradation  (A.nidulans (XP_659106) (T.atroviride (EHK40646 and EHK46127)  was used as a query in the tBLASTn search (R. miehei CAU-432(AGC24356), the only fungal GH3 family member with NAGase activity to date were cultured under seven different growth conditions prior to RNAextraction: i) Cove\u2019s Complete medium (MCc) containing(w/v) 1% glucose, 0.6% NaNO3, 0.15% casein hydrolisate, 0.05% yeastextract, 0.2% peptone, pH 7.0 plus 2% (v/v) salts solution  and 0.04% (v/v) Trace Elements Solution  (ii) 0.25% GlcNAc in minimum mediumcomposed of 0.6% NaNO3 (w/v) plus 0.25% GlcNAc) (w/v) as carbohydratesource, with salts and trace element solutions (iii) 1%Chitin in minimum medium composed of 0.6% NaNO3 (w/v)plus 1% crystalline chitin from crab shells as a carbohydrate source, with saltsand trace element solutions  for 72 h at 28 \u00b0C, then washedwith sterile distilled water and filtered through Miracloth andfrozen in liquid nitrogen for total RNA extraction; iv)Autolysis: medium for mycelium autolysis induction (1% glucose(w/v) and 0.6% NaNO3 (w/v), sustained for 9 days) (v) Sporulation: on MCc agar plates for conidia RNAextraction; vi) Blastospores: Inoculation of5104 conidia/mL on ADAMEK medium for blastospore production , shaking for 64 h at28 \u00baC (vii)Appressoriuminduction medium: 5105 conidia/mL wasinoculated in 0.004% yeast extract solution on 500 glass coverslips for 16 h at28 \u00baC .at28 \u00baC ; vii)ApM. anisopliae cells harvested underall seven different growth conditions was performed in triplicate. Samples wereground using a mortar and pestle in liquid nitrogen, prior to standard RNAextraction using Trizol Reagent .Residual DNA was removed with DNase . Thereafter,extracted RNAs were passed through RNeasy Cleanup columns . RNA samples were quantified using a Qubit fluorometer , and stored at -80 \u00b0C. One microgram oftotal RNA was used for cDNA synthesis using MMLV-RT enzyme . All procedures were performed according to themanufacturer\u2019s instructions.Total RNA extraction from Polymerase chain reactions were carried out on ABI-7500 Real-Time PCR System. Platinum SYBR Green qPCRSuperMix-UDG  was used to monitordsDNA synthesis. Each biological sample was analyzed in technical triplicates;no-template and no-reverse transcriptase controls were included.TableS1). Five housekeeping genes were evaluated:act (\u03b3-actin), gapdh , tef1-\u03b1 (translation elongationfactor 1-\u03b1), trp1 (tryptophan biosynthesis enzyme), andtub (\u03b1-tubulin). The efficiency of each reference geneacross samples was analyzed using geNorm version 3.5 .Primers for qPCR assays were designed using VECTOR NTI software  (-DDCt method (p < 0.05) were performed to determine statisticaldifferences among 2-DDCt values of the seven experimentalconditions.Melting curves from each qPCR reaction were analyzed to confirm specificity ofthe synthesized products and absence of primer dimers. Relative transcriptexpressions were analyzed by Cq (quantification cycle) values, applying the2t method . ResultsM. anisopliae genome, usingNagA from A. nidulans and NAG1 and NAG2 from T.atroviride as queries, resulted in the identification of twoputative GH20 NAGases, named MaNAG1  and MaNAG2 . All otherfungal GH20 NAGase sequences and GH20 conserved domain sequences used as queriesresulted in alignments with the same two previously detected contigs. Therefore,MaNAG1 and MaNAG2 are most probably the only M. anisopliaeputative GH20 NAGases. The GH20 family domain (IPR015883) and the conservedmotif of GH20 proteins (H/N-x-G-A/C/G/M-D-E-A/I/L/V) were found in both MaNAG1and MaNAG2 sequences . Additionally, the putative GH20 NAGasesalso exhibited a chitobiase/beta-hexosaminidase N-terminal domain (IPR029018)(The survey of NAGase genes of the R029018).M. anisopliae genome allowed theidentification of seven positive matches. However, phylogenetic analysis clearlyrevealed that only two sequences, named MaNAG3  and MaNAG4  could be putative GH3 NAGases.Furthermore, these sequences exhibit higher similarity with bacterial GH3NAGases and the RmNag GH3 . MaNAG3 and MaNAG4 share a conserveddomain with GH3 family members (IPR001764) and exhibit the conserved sequencemotif of GH3 proteins (K-H-F/I-P-G-H/L-G-x-x-x-x-D-S/T-H). Furthermore, MaNAG3 and MaNAG4sequences present a conserved GH3 C-terminal domain (IPR002772) , raising the possibility of functionalequivalence.The GH3 domain screening of the R002772) . The othM. anisopliae putative NAGasesare listed in A.nidulans NagA (68 kDa) (T. atrovirideNag1 (73 kDa) (T. harzianum P1 Nag1 (72 kDa) , containingthe highest expected number of introns (4) and theoretical molecular mass (98.71kDa), with N-glycosylation translational modification signals on six sites. Incontrast, MaNAG4 ORF size is 2,057 bp, the theoretical molecular mass is 60.67kDa and the pI of predicted mature protein is 5.6. The predicted molecular massof MaNAG4 is similar to most known bacterial GH3 NAGases, as S.thermoviolaceus NagA (60 kDa) , were recognizedin all of GH20 orthologs .Twenty-six MaNAG1 and MaNAG2 orthologs were identified in 15 filamentous fungigenomes. Most of them are single copy of each putative GH20 NAGase of M. anisopliae and the other fifteen ascomycetes revealed anearly duplication event in GH20 NAGases, resulting in two distinct main clades. This cluster also formed a statistically supported cladewith species from the Aspergillus genus. In contrast, MaNAG2exhibits a more diverse evolutionary history, with orthologs present inTrichoderma sp., Fusariumsp, Neurospora sp., and Magnaporthe sp.Interestingly, the present evolutionary analysis revealed that both NAG1 andNAG2 from the mycoparasite T. atroviride, used in theM. anisopliae genome screening, are evolutionarily morerelated to MaNAG2 . All 15 filamentous fungi have MaNAG3orthologs. However, the M. acridum gene ortholog was notincluded in the phylogenetic analysis, since it was not properly annotated inthe M. acridum genome. In turn, onlyTrichoderma sp., Aspergillus sp., and theentomopathogens C. militaris and B. bassianahave MaNAG4 orthologs.Twenty-three MaNAG3 and MaNAG4 orthologs were identified on the filamentous fungigenomes examined. Conserved sequence motifs of GH3 proteins (K-H-F / I-P-G-H /L-G-x-x-x-x-D-S / T-H) were found in all of them, however, few amino acidresidues substitutions were observed . Additionally, geneduplication of MaNAG3 and MaNAG4 orthologs was not observed.The phylogenetic tree revealed that MaNAG3 and MaNAG4 orthologs formed twodistinct clusters . Both MaS. thermoviolaceus NagA, Clostridiumparaputrificum NagZ, Alteronomonas sp. HexA,V. furnissii NagZ, Thermotoga maritma NagAand T. neapolitana CbsA) grouped into distinct clades fromfungal NAGases. This is probably due to the fact that some bacterial NAGases donot necessarily have GlcNAc hydrolysis specificity over chitooligosaccharides.For example, E. coli NagZ cleaves GlcNAc from muropeptidespresent in the bacterial cell wall  and GH3 family (MaNAG3 andMaNAG4) of M. anisopliae genome wereanalyzed.Virulence determinants are the main focus in the study of entomopathogenic fungi. As chitM. anisopliae by gel-filtration,with a pI of 6.4 and molecular mass of 110-120 kDa. We hypothesize that thisM. anisopliae purified enzyme could be the MaNAG1 presentedhere, based on the predicted pI (6.07) and molecular mass (66.98 kDa) of MaNAG1,likely forming a homodimer. In fact, some fungal GH20 NAGases  clustered together with characterized \u03b2-glucosidases, again suggesting thepossibility of similar function  would repressexpression of chitinolytic enzymes by their own activity products in M.anisopliae (catabolic repression)   in T.reesei, the product of which, an MaNAG3 ortholog, holds suggestedNAGase activity. The phylogenetic analyses indicate that MaNAG3 and T.reesei NAG3 are phylogenetically related  exhibits 63%identity with characterized GH84 from Penicillium chrysogenum(XP_002557703.1). The P. chrysogenum GH84 NAGase not only exhibitactivity against GlcNAc substrates, but also hydrolyzes substrates withgalacto-configuration and exhibits transglycosylation activity(Our results are in agreement to previous suggestions of the presence of GH3 NAGasesin fungi (RmNag) ( related . The exiM. anisopliae and entomopathogenic fungi. Thisanalysis will allow the selection of genes for further functional characterizationto elucidate the process and to identify redundancies and specificities. The viewthat chitinase diversity is merely redundant may not correct (M. anispoliae putative GH20 NAGase genesrevealed induced transcript production in the presence of chitin, potentially in theextracellular milieu. The detection of MaNAG3 andMaNAG4 putative genes is the first evidence for the presence ofa possible GH3 family of NAGases in entomopathogenic fungi. MaNAG3and MaNAG4 expression is responsive to chitinous substrates,suggesting their potential influence on cell differentiation during the M.anisopliae life cycle.In conclusion, this study explored relevant evolutionary aspects of putative GH3 andGH20 NAGase genes and the expression analysis highlighted possible functions forthese genes in"}
+{"text": "Scientific Reports 10.1038/s41598-018-26198-7, published online 18 May 2018Correction to: The Acknowledgements section in this Article is incomplete.\u201cWe thank Sara Olenich, Kayley Abell-Hart and Thanh Von for technical assistance. This study was funded by the Osher Foundation, and in part by NCCIH RO1 AT009267.\u201dshould read:\u201cWe thank Sara Olenich, Kayley Abell-Hart and Thanh Von for technical assistance. This study was supported by the Osher Foundation (H.M.L.), NCCIH RO1 AT009267 (H.M.L.), Susan G. Komen Foundation PDF16376814 (J.B.), National Institutes of Health (NIH) R35 CA210057 (J.J.Z.) and Breast Cancer Research Foundation (J.J.Z.).\u201d"}
+{"text": "Serratia marcescens is a ubiquitous Gram-negative opportunistic pathogen. This announcement describes the isolation and genome annotation of S. marcescens T5-like siphophage Slocum. Terminal repeats, 170 protein-coding genes, and 23 tRNAs were predicted in the 112,436-bp Slocum genome. Serratia marcescens is a ubiquitous Gram-negative opportunistic pathogen. This announcement describes the isolation and genome annotation of S. marcescens T5-like siphophage Slocum. Terminal repeats, 170 protein-coding genes, and 23 tRNAs were predicted in the 112,436-bp Slocum genome. Serratia marcescens is a Gram-negative bacillus that is ubiquitous throughout nature and is a member of the family Enterobacteriaceae , was cultured aerobically at 30\u00b0C and 37\u00b0C in LB (BD), and phages were propagated via the soft-agar overlay method (http://www.bioinformatics.babraham.ac.uk/projects/fastqc). After trimming was performed using the FastX Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit), the raw Slocum contig was assembled using SPAdes v3.5.0, with default parameters, to 409.3-fold coverage  (Bacteriophage Slocum originated from filtered (0.2-\u03bcm filter) municipal wastewater in Bryan, Texas. The host, y method . After py method . Phage gy method , and theoverage 810. Rho-ioverage 8. Functiooverage 8\u201315. Addiverage 8\u2013. Structuverage 8\u2013. The genverage 8\u2013. All lisaxy-pub) , 21. UnlEscherichia phage T5 (GenBank accession no. NC_005859) and T5-like Escherichia phage slur09 (GenBank accession no. LN887948), with 27 to 29% nucleotide identity to slur09 and several Salmonella phages, fragmented across the genome.The phage Slocum genome consists of 112,436 bp of double-stranded DNA, with a G+C content of 44.8%. With 170 predicted protein-coding genes and 23 predicted tRNA genes, the genome protein coding density is 88%. Additionally, 12,521-bp predicted direct terminal repeats were used as the boundary for genome reopening. At the amino acid level, phage Slocum shares 81 similar proteins with MN095770, BioProject accession no. PRJNA222858, SRA accession no. SRR8892204, and BioSample accession no. SAMN11411459.The genome sequence and associated data for phage Slocum were deposited under GenBank accession no."}
+{"text": "Gemcitabine/nab-paclitaxel and FOLFIRINOX demonstrated significantly increased survival compared with gemcitabine in metastatic pancreatic ductal adenocarcinoma (PDAC): objective response rate (ORR) 23 and 31.6%, progression-free survival (PFS) 5.5 and 6.4 months, overall survival (OS) 8.7 and 11.1 months. Present phase II study evaluated recommended first-line triplet FIr/FOx schedule.010%, p130%, power 80%, \u03b15%, \u03b220%. Projected ORR: I step, 1/10; II 5/29. Schedule: 12h-timed-flat-infusion/5-fluorouracil 750-800-900 mg/m2 d1-2,8-9,15-16,22-23; irinotecan 120-140-160 mg/m2 d1,15; oxaliplatin 70-80 mg/m2 d8,22; every 4 weeks, according to clinical parameters , liver function). Activity and efficacy were evaluated, and compared using log-rank; limiting toxicity syndromes (LTS), using chi-square.Simon two-step design: pP 0.022).Twenty-nine consecutive patients were enrolled, according to primary/intermediate/secondary Cumulative Illness Rating Scale (CIRS). Median age 62; elderly 13 (44.7%); PS2 3 (10.4%), secondary CIRS 5 (17.2%). Primary endpoint was met: ORR 53% (7/13 patients) as-treated, 50% intent-to-treat. Cumulative G3-4 toxicities: diarrhea 17%, asthenia 14%, hypertransaminasemy 7%, mucositis 7%, vomiting 3%, anemia 3%, thrombocytopenia 3%. LTS were 27.5% overall, 38.4% in elderly. At 3 months median follow-up, PFS 4 months, OS 11 months. PS2 patients showed significantly worse OS (Intensive first-line triplet FIr/FOx is tolerable at modulated doses, and confirms high activity/efficacy in metastatic PDAC. Patients\u2019 careful selection, and exclusion of PS2, can maintain safety profile and efficient dose intensity. Fixed-dose rate gemcitabine (10 mg/m2/minute) maximized intracellular concentrations of the phosphorylated active forms of gemcitabine, and may substitute standard infusion over 30 minutes [Metastatic pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with approximately 6 months median overall survival (OS) \u20133. In th minutes , 5.BRCA 2 or PALB mutations).Even if phase III studies proposing gemcitabine-based associations failed to improve OS \u201316, exceThe association of nab-paclitaxel to gemcitabine reported highest clinical benefit in patients with PDAC, and significantly raised up OS 8.7 months, PFS 5.5 months, and ORR 23%, compared with gemcitabine alone , 23. IntThe most relevant issue limiting the feasibility of addition of more drugs in a chemotherapy combination is the proper design of the schedule assuring the balance between tolerability for individual patients and effective received dose intensity (DI) of each drug in order to obtain the expected efficacy of the combination. Intensive regimens based on triplet chemotherapy in MCRC, and FOLFIRINOX regimen in metastatic PDAC, frequently required proper clinical management of toxicity and treatment modulations due to moderate/severe toxicities , 25.PM to 10AM) timed-flat-infusion (TFI) 5-FU (TFI/5-FU), without leucovorin, associated to irinotecan (CPT-11) and OXP, according to a weekly alternating schedule, also added to bevacizumab [Over the last 10 years, we developed triplet chemotherapy regimen according to FIr/FOx schedule, characterized by 12-hour , lung 2 (6.8%), lymph nodes 22 (75.8%), local recurrence 2 (6.8%), cutaneous/subcutaneous tissue 1 (3.4%), peritoneal carcinomatosis 12 (41.3%), bone 2 (6.8%). Metastatic site was single in 2 patients (6.8%), multiple in 27 (93.1%). Single metastatic sites: lymph nodes 2 patients (6.8%). Liver metastases were single in 1 patient (3.4%), multiple in 17 (58.6%).Five patients underwent surgery of primary pancreatic tumor. One patient received adjuvant gemcitabine, 1 patient underwent radiotherapy associated with capecitabine.Baseline CA19.9 measurement was normal in 3 patients, elevated in 26.In the first step, according to two-steps Simon\u2019s design , assuminThe phase II study was performed among the projected 29 patients. In the intent-to-treat analysis, 14 patients were evaluable, 13 patients did not received at least 3 cycles of treatments, and 2 patients were on-treatment at data cut-off. OR were 7 out of 14 patients, ORR 50%    ; PFS was not significantly different (P = 0.078) , even if OS was trendy worse in patients with other than head pancreatic tumor location , even if OS was trendy worse in patients with liver metastases , median OS 11 months (0-33); among 12 patients with other than head pancreatic tumor locations, median PFS was 2 months (0-21), median OS 3 months (0-25). PFS and OS were not significantly different (Ten patients (34.4%) received, at least, a second line treatment: FIr/FOx re-challenge in 1 patient (3.4%); gemcitabine/nab-paclitaxel association in 8 patients (27.5%); intra-arterial chemotherapy in 1 patient (3.4%). Two patients (6.8%) received a third line treatment: gemcitabine in 1 patient (3.4%); capecitabine in 1 patient (3.4%).All evaluable patients with an increased Ca19.9 baseline value had a >20% decrease: 58% \u226550% decrease, 42% \u226570% and 90% decrease. CA19-9 levels were not significantly correlated with PFS and OS.Median number of administered cycles was 2 (range 1-11).2/w, 70.4% of projected-DI (pDI); CPT-11 56 (30-80) mg/m2/w, 70%; OXP 29 (11-40) mg/m2/w, 72.5%  mg/m2/w, 79%; OXP 33.5 (0-40) mg/m2/w, 83.75%. In yE patients, median rDIs per cycle were: 5-FU 1500 (375-1800) mg/m2/w, 83.3% of pDI; CPT-11 64 (30-80) mg/m2/w, 80%; OXP 34 (11-40) mg/m2/w, 85%.Median received dose intensities (rDI) per cycle were: 5-FU 1268.5 375-1800) mg/m75-1800 mTable Overall, LTS were observed in 8 patients 27.5%) , and OS 11 months , in the preliminary analysis of efficacy. Activity and clinical outcome of FIr/FOx exceeded that reported with gemcitabine: median OS 5.7 months, and 1-year OS rate 20%, median PFS 2.08 months, ORR 5.4% .P < 0.001) [Recently, phase III trials evaluating more intensive first-line chemotherapy regimen, such as gemcitabine plus nab-paclitaxel and triplet FOLFIRINOX, demonstrated to be much more, and equivalently effective, with significantly increased survival benefit over standard gemcitabine alone in metastatic PDAC patients : ORR 23%< 0.001) . OS rate< 0.001) , 27.Our present real life study on consecutive, unselected patients showed that 13 patients (44.8%) were not evaluable in the ITT analysis because they did not received at least 3 cycles of treatment, thus confiming that, even today, the primary challenge of clinical management of metastatic PDAC patients is to start and safely perform at least 3 cycles of intensive chemotherapy treatment, to evaluate activity contributing to increase clinical outcome.P = 0.022), thus confirming overall benefit in clinical outcome achieved by metastatic PDAC patients with PS 0-1, treated with gemcitabine-based combinations [Among patients treated with intensive triplet FIr/FOx regimen, PFS and OS were not significantly different among patients treated with modulated and standard drug dosage, due to clinical parameters requiring treatment modulations. PS 2 significantly affected worse OS , the safety profile was less favourable than that reported in the gemcitabine arm [Median rDIs were >80% of the projected dose for each drug, also in elderly patients. Cumulative G3-5 toxicities were prevalently represented by: diarrhea (17%), stomatitis/mucositis (6%), asthenia (14%), nausea (3%), vomiting (3%), hypoalbuminemia (3%), hypokalaemia (7%), hypertransaminasemia (7%), hyperbilirubinemia (3%), hypercreatininemia (3%), anemia (3%), neutropenia (17%), thrombocytopenia (3%). In the previously reported phase II trial proposing intensive first-line triplet chemotherapy plus bevacizumab according to FIr-B/FOx regimen, as first-line treatment in MCRC patients, cumulative G3-4 toxicities were similar: diarrhea (28%), stomatitis/mucositis (6%), asthenia (6%), hypertension (2%), hypertransaminasemy (4%), neutropenia (10%). Triplet FIr/FOx, even associated to bevacizumab, determined only 10% G3-4 neutropenia, while FOLFOXIRI schedule, added or not to bevacizumab , 27, prebine arm . More ad2 plus nab-paclitaxel 125 mg/m2 association, 3 out of 4 weeks [In the phase III trial, gemcitabine 1000 mg/m 4 weeks , prevale 4 weeks .In the present study, LTS were observed in 27.5% of individual patients and in 38.4% of elderly patients treated with FIr/FOx regimen. The innovative clinical evaluation of LTS, consisting of at least the LT associated or not to other G2 or LT, introduced to better evaluate, in the individual patient, the presence of LT alone, LTS-ss, or the association of major toxicities in different sites, LTS-ms, showed that: overall, they were 3.4% and 24.1%, respectively; among elderly patients, they were all LTS-ms 38.4%. LTS were not significantly represented by LTS-ms compared to LTS-ss, even if the small number of enrolled patients requires further analyses. LTS-ms were mostly represented by diarrhea, mucositis, asthenia, neurotoxicity, thrombocytopenia and/or anemia, associated to nausea, vomiting, anorexia and/or neutropenia, hypokaliemia, hypoalbuminemia, hypertransaminasemia. In the individual patient, limiting and moderate toxicities often characterized LTS, previously observed in 44% MCRC patients treated with FIr-B/FOx, and equally involving single or multiple sites .DPYD, UGT1A1, ABCB1, CYP3A4, specifically influencing fluoropyrimidines and CPT-11-related adverse events, justifying inter-patients variability in safety profile, may help selection of patients fit for triplet chemotherapy, and may predict the occurrence of individual LTS, prevalently gastrointestinal [Pharmacogenomic analysis evaluating 5-FU degradation rate (FUDR) and/or detection of a panel of DNA Single Nucleotide Polimorphisms (SNPs) involving different genes, such as testinal , 41, 42.Reported data confirmed that intensive regimens, such as FIr/FOx, frequently required proper clinical management of toxicity and treatment modulations due to moderate/severe toxicities. Careful selection of eligible patients, based on age, PS, comorbidity index, and monitoring of individual safety, also according to LTS in individual patients, are major parameters to optimize clinical management of metastatic PDAC. More, close monitoring of patients, expertise with a particular regimen, and toxicity management, remained the physician-related factors, that can guide personalized selection of first-line regimens in individual PDAC patients.Patients were eligible if they had clinical and/or histological/cytological confirmed diagnosis of measurable metastatic PDAC; age \u226518 years, specifically <65 years (non-elderly), \u226565 <75 years (yE), and \u226575 years (oE); WHO PS \u2264 2; adequate hematological, renal and hepatic functions; life expectancy more than 3 months.Ineligibility criteria: pregnancy and breast-feeding; uncontrolled severe diseases; cardiovascular disease ; thromboembolic disease, coagulopathy, pre-existing bleeding diatheses; sensory and/or motor polineuropathy; surgery within the previous 28 days; previous adjuvant chemotherapy or radiotherapy completed less than 6 months before. CIRS was used to evaluate the comorbidity status . Primaryin label for metastatic PDAC treatment in Italian public hospitals, and published in Gazzetta Ufficiale Repubblica Italiana . The study was approved by the Regional Review Board , and conducted in accordance with the Declaration of Helsinki. All patients provided written, informed consent.Treatment was approved by Agenzia Italiana del Farmaco (AIFA) for administration This was a single-arm phase II study evaluating safety and activity of weekly alternating 5-FU, CPT-11, and OXP (FIr/FOx) as first-line treatment of metastatic PDAC.2/die, over 12-hour (from 10:00 p.m to 10:00 a.m.), days 1-2, 8-9, 15-16 and 22-23; CPT-11 , 120-140-160 mg/m2, administered over 90 minutes as an intravenous infusion in 250 ml of NaCl 0.9%, days 1 and 15; OXP  over 2-hours as an intravenous infusion in 250 ml of dextrose 5%, at the dose of 70-80 mg/m2, days 8 and 22. Cycles were repeated every 4 weeks. 5-FU was administered by a portable pump  using a venous access device. According to patients\u2019 fitness, and particularly in patients with PS 2, and/or \u226575 years, secondary CIRS stage, and/or abnormal liver functional laboratory tests, such as \u2265 G2 hypertransaminasemia at baseline, drugs\u2019 doses were modulated in individual patients as reported, providing a received DI >75% for each drug, reported as active and efficacious in previous studies of triplet chemotherapy regimens [FIr/FOx association consisted of 5-FU associated to alternating CPT-11 or OXP according to the following weekly schedule: TFI/5-FU , 750-800-900 mg/mregimens , 26\u201331.Physical examination and routine laboratory studies were performed at baseline and every week on-treatment, including complete blood cell count, electrolytes, liver and renal function tests, urine examination and coagulation function; tumor markers every cycle; electrocardiogram every cycle, and echocardiogram at baseline, and every 3 cycles.Primary end-point was ORR; secondary end-points were toxicity, PFS, OS. ORR was evaluated according to RECIST criteria ; PFS andLTS, consisting of at least a LT associated or not to other limiting or G2 toxicities, were evaluated as previously reported , 32. TheCorrelations between maximum decrease from baseline in CA19.9 level and PFS and OS were analyzed, to assess possible relationships between CA19.9 and clinical outcomes.0 was considered as the estimated activity reported with gemcitabine alone (median ORR 10%), and confirmed with the association of gemcitabine plus erlotinib (ORR 8.6%) [1 as the projected ORR using the present intensive triplet combination, according to FIr/FOx schedule, increasing the activity \u2265 20% in metastatic PDAC patients, as reported with FOLFIRINOX (ORR 31.6%), and with the association of gemcitabine and nab-paclitaxel in the phase I/II trial  [This phase II study was planned according to two-steps Simon\u2019s design : assuminRR 8.6%) , 16; p1 I trial) \u201324.FIr/FOx intensive triplet chemotherapy in metastatic PDAC patients preliminary showed high activity. Present schedule was feasible in non-elderly and elderly patients, with PS 0-1, with manageable toxicities, at proper CPT-11, OXP, and 5-FU doses. LTS were prevalently characterized by diarrhea, mucositis, asthenia, anemia, neurotoxicity and/or thrombocytopenia, associated to nausea, vomiting, anorexia and/or neutropenia. Elderly patients preliminary showed trendy, but not significantly worse OS. Adequate selection of suitable patients, based on clinical parameters, aimed to maintain safety profile at efficacious DIs, will verify if more intensive approaches, such as triplet chemotherapy regimen could increase OS, compared to historical gemcitabine control, in the clinical practice management of metastatic PDAC patients, and if it could be a treatment option in locally advanced/borderline resectable pancreatic cancer patients."}
+{"text": "Also, the data include the bond lengths and angles of [Ln2(IMBA)6(dmp)2] .In this data article, we present the FT-IR and PXRD data of the lanthanide complexes constructed by 4-iodo-3-methylbenzoic acid (IMBA) and 4,7-dimethyl-1,10-phenanthroline (dmp). Detailed structure analysis, luminescence and sensing properties were discussed in our previous study, \u201cHighly Luminescent Lanthanide Complexes as Bifunctional Sensor for Et Specifications tableValue of the data\u2022This structure information would be valuable for FT-IR analysis of lanthanide complexes.\u2022This data would be worthy for further investigation of the PXRD properties.\u2022This data provide a new process to synthesize two ligands coordinated lanthanide complexes.1[Ln2(IMBA)6(dmp)2] ,  are isostructural. They crystallize in triclinic space group P-1 (no. 2). These complexes are dinuclear cluster structures which contain two lanthanide ions , six deprotonated IMBA and two dmp, forming an electroneutral unit (1a that incubation in aqueous solution for as long as the sensing time (1\u202fh) was in line with the as-synthesized sample and calculated data, confirming that the sensor 1a is a highly stable 6(dmp)2]\u00b7(1a). Yield: 37% based on Eu3+. Anal. Calcd (%): C, 39.92; H, 2.645. Found (%): C, 40.13; H, 2.633. FT-IR (\u22121): 3450 (m), 2947 (w), 1620 (s), 1584 (w), 1438 (s), 1403 (m), 1305 (m), 1193 (w), 1033 (m), 934 (w), 872 (m), 789 (s), 747 (w), 550 (w), 482 (w).[Eu3. FT-IR  (KBr pel2(IMBA)6(dmp)2] (1b). Yield: 34% based on Gd3+. Anal. Calcd (%): C, 39.74; H, 2.633. Found (%): C, 39.61; H, 2.618. FT-IR (\u22121): 3435 (m), 2995 (w), 1648 (w), 1592 (s), 1542 (m), 1495 (w), 1438 (s), 1305 (m), 1152 (w), 1033 (s), 943 (m), 872 (s), 789 (s), 739 (m), 600 (w), 550 (w), 488 (m).[Gd8. FT-IR  (KBr pel2(IMBA)6(dmp)2] (1c). Yield: 35% based on Tb3+. Anal. Calcd (%): C, 39.68; H, 2.629. Found (%): C, 39.79; H, 2.637. FT-IR (\u22121): 3443 (m), 2980 (w), 1627 (w), 1592 (s), 1542 (m), 1494 (w), 1430 (s), 1312 (w), 1152 (w), 1033 (s), 934 (w), 872 (m), 824 (m), 789 (s), 739 (w), 550 (w), 488(w).[Tb7. FT-IR  3\u00b76H2O , dmp and IMBA at a molar ratio of 1:1:1.5 at 333\u2009K for 120\u2009h. The colorless block single crystals of 1a\u20131c were collected by filtration, and mounted on a glass fiber Lanthanide complexes Single crystal X-ray diffraction data were obtained on an instrument of Bruker SMART 1000 CCD, at wavelength of 0.71073\u2009\u00c5 (Mo-Ka radiation) at 25\u2009\u00b0C. The structures were refined by full-matrix least-squares methods with SHELXL-97 module. Phase purity of bulk samples were tested by PXRD, on a DMAX2200VPC diffractometer"}
+{"text": "Oribacterium sp. strain C9, isolated from the rumen of a steer grazing on Rhodes grass in Rockhampton, Queensland, Australia.We report the 3.7-Mb genome sequence of  Oribacterium sp. strain C9, isolated from the rumen of a steer grazing on Rhodes grass in Rockhampton, Queensland, Australia. This draft genome consists of 3,720,024\u2009bp with a 42.8% G+C content, 3,130 predicted coding DNA sequences (CDSs), and 67 RNAs.We report the 3.7-Mb genome sequence of  Oribacterium was first proposed by Jean-Philippe Carlier  in Australia , using an anaerobic in vitro culture with 0.8% yeast extract. DNA sequencing was performed at Macrogen (South Korea) on a HiSeq 2500 instrument  (2 \u00d7 100-bp paired-end sequencing) to produce 58,649,712 total reads with 15\u00d7 coverage. Raw reads were trimmed with Trimmomatic version 0.32   version 4 (https://www.ncbi.nlm.nih.gov/genome/annotation_prok) . The average nucleotide identity (ANI) was calculated using the algorithm described by Yoon et al. (Oribacterium sp. strains P6A1 (GenBank accession number JNKO00000000), WCC10 (FOPH00000000), NK2B42 (AUJX00000000), FC2011 (JNJM00000000), and KHPX15 (FNRG00000000) (from the Hungate1000 project [O. parvum strain ACB1T (AFZC00000000), and O. sinus F0268 (ACKX00000000), respectively.The 16S rRNA gene sequence was found to be 95% identical to that of in ACB1T  and Oribn et al.  with theOribacterium sp. strains P6A1, WCC10, NK2B42, FC2011, and KHPX15, O. sinus F0268, and O. parvum strain ACB1T are 28.4%, 24.9%, 24.4%, 24%, 24.1%, 21.1%, and 26.1%, respectively. Having both ANI values lower than 95% and dDDH values lower than 70% indicated that isolated strain C9 is a species distinct from the other isolates.The digital DNA-DNA hybridization (dDDH) was determined using the Genome-to-Genome Distance Calculator (GGDC) version 2.1 with default settings . The esthttp://www.cazy.org/) (http://csbl.bmb.uga.edu/dbCAN/)  (zy.org/)  and dbCA/dbCAN/) , we idenOribacterium sp. strain C9 has been deposited in DDBJ/EMBL/GenBank under the accession number MUHW00000000, BioProject number PRJNA362832, and BioSample number SAMN06249574. The genomic raw sequencing reads are available in the Sequence Read Archive (SRA) database under accession number SRR8449972.The draft genome sequence for"}
+{"text": "Peptide nucleic acids (PNAs) are very useful tools for gene regulation at different levels, but in particular in the last years their use for targeting microRNA (anti-miR PNAs) has provided impressive advancements. In this respect, microRNAs related to the repression of cystic fibrosis transmembrane conductance regulator (CFTR) gene, which is defective in cystic fibrosis, are of great importance in the development of new type of treatments. In this paper we propose the use of an anti-miR PNA for targeting miR-145, a microRNA reported to suppress CFTR expression. Octaarginine-anti-miR PNA conjugates were delivered to Calu-3 cells, exerting sequence dependent targeting of miR-145-5p. This allowed to enhance expression of the miR-145 regulated CFTR gene, analyzed at mRNA  and CFTR protein (Western blotting) level. MicroRNAs  are from 19 to 25 nucleotide-long noncoding RNAs that regulate gene expression by targeting mRNAs, leading to a translational repression or mRNA degradation ,4. SinceEpigenetic regulation of expression of cystic fibrosis transmembrane conductance regulator (CFTR) gene by miRNAs has been recently explored by different groups in cystic fibrosis (CF) primary bronchial epithelial cells in vitro or from bronchial brushings ex vivo. Expression of miR-145 and miR-494 was found anti-regulated with that of CFTR . The effPeptide nucleic acids (PNAs) are DNA analogues of outstanding properties ,13,14, sThe objective of this study was to design a PNA targeting miR-145, determine its activity in inhibiting miR-145, and verify whether it induces an increase of CFTR production. The PNA was conjugated to a poly-arginine tail, since this type of constructs was previously used by our group for the delivery of PNA into cell lines ,25,26. AIn Homo sapiens (H. sapiens), Pan troglodytes (Chimp), Macaca Mulatta (Rhesus), Saimiri sciureus (Squirrel), Mus musculus (Mouse), Rattus norvegicus (Rat), Oryctolagus cuniculus (Rabbit), Sus scrofa (Pig), Bos taurus (Cow), Felis catus (Cat), Canis lupus familiaris (Dog), Myotis lucifugus (Brown bat) and Loxodonta africana (Elephant). In red the differences with respect to the Homo sapiens sequence are underlined. The CFTR sequence complementary to miR-145-5p seed region is boxed. The region showing the highest level of homology (reaching 100% homology in some positions) belongs to the seed-complementary stretch, as also shown in 8) peptide was conjugated at N-terminus of the PNA chain since it induces an efficiency in the delivery which approaches 100% , as elsewhere published 4+, 1255.97 (1256.07) [MH5]5+, 1046.81 (1046.9) [MH6]6+, 897.41 (897.48) [MH7]7+, 785.36 (785.42) [MH8]8+, 698.21 (698.26) [MH9]9+.R8-PNA-a145-MUT: sequence H-R8-AGAGATGCCTTGGAGAAC-Gly-NH2; Rt = 10.23 min, HRMS: calculated MW: 6272.4698 g/mol; m/z found : 1569.70 (1569.84) [MH4]4+, 1255.97 (1256.07) [MH5]5+, 1046.81 (1046.9) [MH6]6+, 897.41 (897.48) [MH7]7+, 785.36 (785.42) [MH8]8+, 698.21 (698.26) [MH9]9+.R8-PNA-a509: sequence H-R8-CTACCCACAGACGTACCA-Gly-NH2; Rt = 11.17 min, HRMS: calculated MW: 6097.4456 g/mol; m/z found : 1525.95 (1526.07) [MH4]4+, 1220.96 (1221.06) [MH5]5+, 1017.64 (1017.71) [MH6]6+, 872.40 (872.47) [MH7]7+, 763.48 (763.54) [MH8]8+, 678.76 (678.81) [MH9]9+, 610.98 (611.03) [MH10]10+R8-PNA-a494: sequence H-R8-TTTCCCGTGTATGTTTCA-Gly-NH2; Rt = 12.30 min, HRMS: calculated MW: 6146.4044 g/mol; m/z found : 1538.19 (1538.32) [MH4]4+, 1230.75 (1230.86) [MH5]5+, 1025.80 (1025.88) [MH6]6+, 879.40 (879.47) [MH7]7+, 769.60 (769.66) [MH8]8+, 684.20 (684.25) [MH9]9+, 615.88 (615.93)[MH10]10+R8-PNA-a433: sequence H-R8-CGGGGAACCCTTCAAGAT-Gly-NH2; Rt = 10.47 min, HRMS: calculated MW: 6208.4525 g/mol; m/z found : 1036.14 (1036.22) [MH6]6+, 888.26 (888.33) [MH7]7+, 777.35 (777.41) [MH8]8+, 691.09 (691.15) [MH9]9+.2/air in DMEM/F12 medium  supplemented with 10% fetal bovine serum , 100 units/mL penicillin and 100 \u03bcg/mL streptomycin  and 1% NEEA (100\u00d7) . To determine the effect on proliferation, cell growth was monitored by determining the cell number/mL using a Z2 Coulter Counter .Calu-3 ,30 and NCultured cells were trypsinized and collected by centrifugation at 1500 rpm for 10 min at 4 \u00b0C, washed with PBS, lysed with Tri-Reagent , according to manufacturer\u2019s instructions. The isolated RNA was washed once with cold 75% ethanol, dried and dissolved in nuclease free pure water before use. RNA from human tissue sections was extracted and purified with miRNeasy FFPE Kit  according to the manufacture's procedures. For miRNA quantification using real-time RT-qPCR reagents, the primers and probes  were obtained from Applied Biosystems . Reverse transcriptase (RT) reactions were performed using the TaqMan MicroRNA Reverse Transcription Kit (Applied Biosystems); real-time PCR was performed according to the manufacturer\u2019s protocols . The quaGene expression analysis was performed by RT-qPCR. 300 ng of the total RNA was reverse transcribed by using random hexamers. Quantitative real-time PCR (qPCR) assays were carried out using gene-specific double fluorescently labeled probes. Primers and probes used to assay CFTR (Assay ID: Hs00357011_m1) gene expression were purchased from were from Applied Biosystems, (Applied Biosystems). The relative expression was calculated using the comparative cycle threshold method and, as reference genes, the human RPL13A (Assay ID: Hs03043885_g1) ,53. g for 10 min at 4 \u00b0C. Protein concentration was determined by the method of Lowry after precipitation with 5% Trichloroacetic acid (TCA), utilizing bovine serum albumin as a standard. For CFTR analysis 20 \u03bcg of total protein was heated in Laemmli buffer (Bio-Rad) at 37 \u00b0C for 10 min and loaded onto a 3 to 8% Tris-acetate gel (Bio-Rad). The gel proteins were transferred to PVDF membrane (Bio-Rad) by using Trans Blot Turbo (Bio-Rad) and processed for Western blotting by using mouse monoclonal antibody, clone 596, against NBD2 domain of CFTR  at a dilution of 1:2500 by an overnight incubation at 4 \u00b0C. After washes, membranes were incubated with horseradish peroxidase-coupled anti-mouse immunoglobulin  at room temperature for 1 h and after washes the signal was developed by enhanced chemiluminescence . After membranes stripping, \u03b2-Actin monoclonal antibody (Sigma-Aldrich) was used to confirm the equal loading of samples [Cell pellets were lysed in 1% Nonidet P40 (IGEPAL), 0.5% sodium deoxycholate, 200 mM NaCl, 10 mM Trizma base, pH 7.8, 1 mM EDTA plus protease inhibitor mixture and 1 mM PMSF for 30 min in ice. Lysates were cleared by centrifugation at 10,000\u00d7  samples . Annexin V and Dead Cell assay on Calu-3 cells, untreated and treated for 72 h with 2 \u03bcM PNA-a145, 5 \u03bcM Stattic together with 10% DMSO, were performed with \u201cMuse\u201d  method, according to the instructions supplied by the manufacturer. This procedure utilizes Annexin V to detect PS (PhosphatidylSerine) on the external membrane of apoptotic cells. A dead cell marker is also used as an indicator of cell membrane structural integrity. It is excluded from live, healthy cells, as well as early apoptotic cells. Four populations of cells can be distinguished in this assay. Cells were washed with sterile PBS 1X, trypsinized, suspended and diluted (1:2) with the one step addition of the Muse Annexin V & Dead Cell reagent. After incubation of 10 min at room temperature in the dark, samples were put on ice, vortexed and analyzed. Data from prepared samples are acquired and recorded utilizing the Annexin V and Dead Cell Software Module (Millipore) .t test. Statistical significance was defined with p < 0.05  and p < 0.01 .Results are expressed as mean \u00b1 standard deviation (S.D.) Comparisons between groups were made by using paired Student\u2019s"}
+{"text": "Brassica campestris L. ssp. chinensis var. rosularis Tsen). However, there is little information on the global gene expression of BRs under LT stress in wucai. In this study, the molecular roles of 24-epibrassinolide (EBR) after exogenously application, were explored by RNA sequencing under LT conditions.Brassinosteroids (BRs) have a positive effect on many processes during plant growth and development, and in response to various abiotic stressors. Low-temperature (LT) stress constricts the geographic distribution, growth, and development of wucai (According to the Gene Ontology (GO) term and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, photosynthesis was significantly enriched after spraying EBR under LT. The transcripts encoding the photosystem II (PSII) oxygen-evolving enhancer protein, photosystem I (PSI) subunit, light-harvesting chlorophyll protein complexes I and II, and ferredoxin were up-regulated after the application of EBR. Transcripts encoding several key enzymes involved in chlorophyll biosynthesis were also up-regulated, accompanied by significant differences in the contents of 5-aminolevulinic acid (ALA), porphobilinogen (PBG), protoporphyrin IX (Proto IX), Mg-protoporphyrin IX (Mg-proto IX), protochlorophyllide (Pchl), and photosynthetic pigments. Notably, transcriptional and physiological analyses revealed that under LT stress, plant responses to EBR involved a major reorientation of photosynthesis, as well as porphyrin and chlorophyll metabolism.This study explored the role of EBR as an LT stress tolerance mechanism in wucai. At the transcription level, LT tolerance manifests as an enhancement of photosynthesis, and the amelioration of porphyrin and chlorophyll metabolism. Brassica campestris L. ssp. chinensis var. rosularis Tsen) is a variant of non-heading Chinese cabbage (Brassica campestris L.), a crucial species in the Brassicaceae family  [\u03c8o / (1\u2013 \u03c8o)]. Maximum quantum yield of primary photochemistry was calculated as: \u03c6Po\u2009=\u2009Fv / Fm\u2009=\u2009(Fm - Fo) / Fm [In vivo chlorophyll formulae . The flumeasured . The folFo) / Fm . Other pp\u2009<\u20090.05. Analyses were conducted using SPSS v19.0 for Windows . Figures were plotted using GraphPad Prism v7.0 and Origin Pro v9.1 software .All of the data from the 4 treatments were subjected to an analysis of variance (ANOVA). The mean separation was performed using the Fisher\u2019s protected least significant difference (LSD) test with a significance level of Additional file 1: Figure S1. The quality control of six samples. A-F represent LT-1, LT-2, LT-3, LT\u2009+\u2009EBR-1, LT\u2009+\u2009EBR-2, and LT\u2009+\u2009EBR-3, respectively.Additional file 2: Figure S2. The reads mapping of six samples. A-F represent LT-1, LT-2, LT-3, LT\u2009+\u2009EBR-1, LT\u2009+\u2009EBR-2, and LT\u2009+\u2009EBR-3, respectively.Additional file 3: Figure S3. The 20 DEGs we randomly selected for qRT-PCR assay in EBR-mediated LT stress.Additional file 4: Figure S4. The results of identifying EBR concentration.Additional file 5: Table S1. Summary of sequence assembly after illumine sequencing.Additional file 6: Table S2. Number of reads sequenced and mapped to the Brassica rapa genome.Additional file 7: Table S3. The expression patterns of the cold acclimation and cold-induced genes.Additional file 8: Table S4. The expression patterns of the BR-responsive genes.Additional file 9: Table S5. Primers of verifying genes.Additional file 10: Table S6. Primers of photosynthesis genes.Additional file 11: Table S7. DEGs of photosynthesis\u2013antenna proteins and photosynthesis.Additional file 12: Table S8. Parameters derived from the OJIP transient for use in the current study."}
+{"text": "An update of xanthones encountered in lichens is proposed as more than 20 new xanthones have been described since the publication of the compendium of lichen metabolites by Huneck and Yoshimura in 1996. The last decades witnessed major advances regarding the elucidation of biosynthetic schemes leading to these fascinating compounds, accounting for the unique substitution patterns of a very vast majority of lichen xanthones. Besides a comprehensive analysis of the structures of xanthones described in lichens, their bioactivities and the emerging analytical strategies used to pinpoint them within lichens are presented here together with physico-chemical properties (including NMR data) as reported since 1996. H-xanthen-9-one scaffold [Xanthones are ubiquitous polyphenolic compounds displaying a common 9scaffold . Bioactiscaffold  likely tscaffold . Thus, mscaffold . Even thscaffold , variousAs with most lichen metabolites , the bioet al. [Most lichen xanthones arise through the folding of a polyketide intermediate as described by Cacho et al. , resultiPyrenochaeta terrestris [ortho carboxybenzophenone that might follow several metabolic fates. A first possibility is the 1,4-addition of a B-ring phenol to the A-ring dienone followed by dehydration and decarboxylation to access ravenelin-like xanthones after a final oxidation [Aspergillus variecolor, a skeleton thus far unknown from lichens.In contrast, a limited number of structures arise via a distinct biosynthetic pathway that leads to the ravenelin skeleton, with the methyl group in position 3. This biosynthetic scheme begins with the widespread anthraquinone emodin as a precursor  Figure . To begirrestris ,10. The rrestris . Deeper rrestris , who proxidation . This reHypericum androsaemum) or 1,3,5-trihydroxyxanthone (Centaurium erythraea) [It is noteworthy that the xanthone nucleus of plants is of mixed biosynthetic origin with the A-ring being acetate-derived whereas shikimic acid pathway\u2013derived 3-hydroxybenzoic acid gives rise to the C-ring . Aromatiythraea) .Anthocleista djalonensis [Croton cuneatus [, Cupania cinerea [, Feroniella lucida [, Minquartia guianensis [Zanthoxylum microcarpum [Z. valens [Hypericum ascyron [Penicillium [Penicillium patulum [Ulocladium [Penicillium patulum [Given these different biosynthetic pathways, only few lichen xanthones are known from non-lichenized organisms. However, some can be produced by higher plants, such as lichexanthone , vineto ascyron ). Likewiicillium , norlich patulum  and the ocladium ; 1,3,6-t patulum . Secalon patulum .Vismia parviflora [Weddelina squamulosa [Penicillium strain [Cassia obtusifolia [Astrocystis sp. BCC 22166 [Cassia occidentalis [Phomopsis sp. [Talaromyces bacillosporus [Engyodontium album [O-methyl-2-deprenylrheediaxanthone B ). It can therefore be stated that xanthones are highly unique to each realm, legitimating joint efforts on higher plants, non-lichenized fungi and lichens to widen the chemical diversity of these privileged structures.Likewise, very few xanthones are common to higher plants and fungi, with some such examples being 1,7-dihydroxyxanthone  1,8-dihusifolia  and fungCC 22166 ), pinseldentalis  and sevepsis sp. , Talaromlosporus  and the um album ), 6-O-meillardii  and fungpsis sp. ), as welca. 2000 occurrences of xanthones sensu lato . These considerable efforts might be explained by the discovery of promising leads such as the anti-angiogenic molecule Vadimezan (AS 404) which is currently undergoing phase III clinical trials as a tumor vascular-disrupting agent [Recent data regarding the number of naturally occurring xanthones are scarce, with the last numbered record of 278 xanthones listed by Vieira and Kijjoa more than 10 years ago . By Janung agent ,36, as wng agent ,38. ObviStructural diversity of lichen xanthones mostly stems from variations in the orientation and degree of chlorination of the norlichexanthone or ravenelin core, as well as from the position and extent of methylation of the phenolic groups. Elix and Crook tremendously advanced the understanding of chlorination and methylation processes affecting the xanthone core. Through the example of 40 different species of lichens, theoretical biosynthetic schemes mirroring the sequence of biogenetic events leading to the joint occurrence of xanthones among lichens were constructed . This reet al. in 2002 [Aspergillus nidulans, through the identification of prenyltransferase genes belonging to the fungal indole prenyltransferases, so far known for their involvement in the prenylation of amino acids [Further structural modifications might be taken into account to extend the chemodiversity of lichen xanthones. Important contributors to the chemical diversity of xanthones are prenyl groups which are most often incorporated as dimethylallyl moieties . General in 2002 . Underlyno acids . Glycosyno acids  and hirtno acids . Blennoria sp. [ortho and para C positions [The diversity of lichen xanthones is also extended through some dimeric xanthones. Although the identification of key dimerization processes still warrants further investigation, xanthone dimers are most likely obtained after the biaryl linkage of monomers. This hypothesis is preferred to that of a tandem biosynthetic pathway which would involve a side-by-side cyclization of a double-length polyketide, especially since the discovery of the long-sought-after monomeric units of secalonic acids  within the fungus oria sp. . Enzymatositions .Reductive dearomatizations are sometimes observed on xanthones to yield dihydro-, tetrahydro- or hexahydroxanthones. To date, in lichens, such reduced species were only observed from dimeric xanthones: secalonic acids, hirtusneanoside and eumitrin A1 (bis tetrahydroxanthones) and eumitrin A2, B and T . Overall, a vast majority of xanthones reported from lichens have a monomeric and fully aromatized structure.Lecanora dispersa [Lecanora rupicola. This lichen is known to produce metabolites of various polyketide classes: lecanoric acid (depsides), hematommic and orsellinic acids (monocyclic phenols), eugenitol and sordidone (chromones) and arthothelin (xanthone). The whole chemosyndrome could be produced from axenically grown mycobionts, with the notable exception of arthothelin [Physconia distorta but not by its isolated mycobiont [Pyrenula japonica and P. pseudobufonia revealed the biosynthesis of xanthones that cannot be evidenced from the lichen as a whole (see further), suggesting their significance in the pre-lichenized condition [Even though xanthones from free-living fungi are well known, a plant-fungus collaboration has been suggested for several lichen xanthones. As an example, the typical lichen xanthone 2,7-dichlorolichexanthone could be isolated from the lichen dispersa . Howeverhothelin , suggestycobiont . Adverseondition ,52.Dehalococcoides mccartyi that has gathered attention because of its ability to dechlorinate a large array of anthropogenic pollutants [p-dioxins (dioxins), such bacterial strains might be of paramount interest for bioremediation purposes [A total of 72 xanthones containing one to four chlorine atoms are currently known from lichens. Even though recent studies have shed light on chlorinated xanthone biosynthesis, no study has focused on their degradation. Dechlorination of organochlorines can be achieved by some bacterial strains, referred to as organohalide respirers. Such bacteria comprise the famous llutants ,54,55. Lllutants ,57,58. Tllutants , which illutants ,61,62. Hpurposes . Acting purposes ,63.Lecidella asema, without the need for matrix assistance [in situ [Lecidella asema shows an exhaustive chemical profile  to perform separation and therefore to collect retention time and UV/Vis data as obtained with more traditional dereplication protocols [However, regarding the specific example of lichen xanthones, the lack of mutually supportive data such as chromatographic retention times and/or UV/Vis spectra precludes the differentiation of isomers and is a severe limitation of such methods. Possible ways to overcome such limitations while keeping an rotocols . Even throtocols . Such inrotocols ,83. Thisrotocols .\u00ae was dedicated to the identification of lichen metabolites from an original database of 550 compounds, including HPLC retention indices, TLC-Rf values, UV/visible colors of TLC spots and thalline reactions to guide their identification [Lichenized and non-lichenized Ascomycetes) [Together with analytical achievements, informatics tools were also developed to alleviate the dereplication holdup. A computer software named Wintabolitesfication . A most mycetes) , a databmycetes) .Lecanora schofieldii [O-methylnorlichexanthone , 2,4-dicalderae ), 2,5-diia fusca  among otulescens  and Phyldatinica ), 4,5,7-leprosum ) and 2,4rpallida ). Chlorirpallida . One migBesides these fully rationalized structures, a limited number of metabolites exhibit structural variations that are more or less difficult to explain. As an example, demethylchodatin displays a methoxy group in position 4 that might be introduced by a xanthone oxidase on this trichlorinated norlichexanthone derivative. Another unusual modification of the lichexanthone series is the occurrence of two acetyl groups in erythrommone. The chemical structures of xanthones with trivial names discussed in this manuscript are enlisted in Rinodina thiomela.However, 22 lichen xanthones display unusual structural features that make their biosynthetic intermediates tricky to unravel  [Plasmodiumfalciparum and Trypanosoma brucei [Aedes aegypti [Very few lichen xanthones have been investigated to date for their bioactivities. Lichexanthone exerts a weak activity against rculosis  and M. aM. aurum . DespiteM. aurum . Lichexa: 21 \u03bcM) . No antia brucei . Lichexaa brucei  and indua brucei . Lichexaa brucei . At last aegypti .lck tyrosine kinase at 200 \u03bcg/mL [50 values ranging from 0.3 to 12 \u03bcM [Norlichexanthone was shown to display promising cytotoxic activities . Indeed,00 \u03bcg/mL  and inhito 12 \u03bcM . Norlichto 12 \u03bcM . Dayan ato 12 \u03bcM . Alongsito 12 \u03bcM .Haematomma fluorescens as a response to a 365 nm UV light exposure, emphasizing its protective role as a light filter [As most structural classes of lichen metabolites, xanthones display strong UV-absorbing properties, predominantly in the wavelength range of UVA . Indeed,t filter . Depositt filter ,110. Suct filter . A Time-t filter .The past 20 years witnessed the isolation of 23 further lichen xanthones. It is noteworthy that those new xanthones displayed original substitution patterns and revealed original moieties compared to the previous state of the art.Pyrenula japonica and P. pseudobufonia afforded a series of five new ravenelin-type lichen xanthones, the first reports of such xanthones from aposymbiotically grown lichen mycobionts. A first report elucidated the structures of 1,5,8-trihydroxy-3-methylxanthone, 1,8-dihydroxy-5-methoxy-3-methylxanthone and 1,7-dihydroxy-3-methylxanthone [Cassia occidentalis [Cultures of the spore-derived mycobionts of xanthone  while a xanthone . Notablydentalis ) and alsdentalis . Surprisdentalis . Besidesdentalis .Umbilicaria proboscidea [d-glucopyranose unit anchored at position O-7, whereas umbilicaxanthoside B stands for a C2,C8-diprenylxanthone with a disaccharide \u03b2-d-glucopyranose-(1\u21924)-\u03b2-d-glucopyranose moiety at O-7. These compounds were the first prenylated xanthones to be isolated from a lichen source and glycosides are not frequently encountered in lichens [O-glycosides corresponding to linolenoyl, lineoyl, palmitoleoyl, oleoyl, palmitoyl, eicosenoyl and stearoyl esters of both umbilicaxanthosides A and B [Two highly unusual glycosylated prenylxanthones were subsequently identified from the Ural lichen boscidea . Umbilic lichens . Besides A and B . FragmenCladonia incrassata [Sporormiella minimoides [Staphylococcus aureus even though the paucity of the compound precluded the determination of a minimal inhibitory concentration [The last monomeric xanthone isolated so far from a lichen source is cladoxanthone A , obtained from crassata . The occcrassata . It is nnimoides   as well as antifungal (Microbotryum violaceum) and antialgal agent (Chlorella fusca) [Gliocladium sp. T 31 streamlined the isolation of secalonic acid D as the cytotoxic metabolite against four cell lines in a highly variable range of 0.03\u201315 \u03bcM, suggesting selective pharmacodynamic properties [50 of secalonic acid D on the carcinoma KB cells and an inhibition of human topoisomerase 1 with a promising IC50 of 0.16 \u03bcg/mL [++-dependent enzymes through competitive inhibition [Regarding dimeric xanthones, known metabolites of the secalonic acid series were first isolated from lichen sources in 2009, anescens . The joianescens . Secalonanescens . Secalonc acid D ,119, wara oryzae . Likewisa fusca) . Severala fusca) . Bioassaoperties . Further16 \u03bcg/mL . More re16 \u03bcg/mL . As a la16 \u03bcg/mL . Anotherhibition . Subsequhibition . As a cohibition ,128.Usnea hirta corresponding to a rhamnoside of an unsymmetrical dimeric tetrahydroxanthone [Staphylococcus aureus and Bacillus subtilis [l-rhamnose and the aglycone named hirtusneanin, the physico-chemical data of which were also reported by Rezanka and Sigler [Hirtusneanoside is a new xanthone dimer isolated from xanthone . If the subtilis . The enzd Sigler .Available physico-chemical properties regarding the newly described lichen compounds are listed below.Cladoxanthone A14H7O4Cl3 (343.94099)CYellow powdermax (MeOH)(log \u03b5): 381 (3.24), 323 (3.50), 259 (4.11) nmUV \u03bbmax (CHCl3): 3353, 3056, 1642, 1597, 1450, 800 cm\u22121IR \u03bdCladonia incrassata [Sources: crassata .1,8-Dihydroxy-3-hydroxymethyl-5-methoxyxanthone15H12O6 (288.06339)CYellow needles, mp 221\u2013222 \u00b0Cmax (MeOH)(log \u03b5): 387.5 (3.27), 341 (3.84), 271 (4.16), 263 (4.24), 255 (4.29), 236 (4.17)UV \u03bbmax (KBr): 3533, 1659, 1634, 1609, 1589, 1493 cm\u22121IR \u03bdPyrenula japonica [Sources: Mycobiont of japonica .1,2,8-Trihydroxy-5-methoxy-3-methylxanthone15H12O6 (288.06339)CYellow needles, mp 214\u2013215 \u00b0Cmax (MeOH) (log \u03b5): 421 (3.42), 352 (3.58), 280 (4.27), 263 (4.18), 243 (4.17), 206 (4.23)UV \u03bbmax (KBr): 3487, 1663, 1636, 1607, 1576, 1489 cm\u22121IR \u03bdPyrenula japonica [Sources: Mycobiont of japonica .1,7-Dihydroxy-3-methylxanthone14H10O4 (242.05791)CYellow needles, mp 259\u2013260 \u00b0Cmax (MeOH) (log \u03b5): 384 (3.84), 290 (3.98), 261 (4.58), 235 (4.45)UV \u03bbmax (KBr): 3285, 1653, 1607, 1585, 1483 cm\u22121IR \u03bdPyrenula japonica [Sources: Mycobiont of japonica .1,5,8-Trihydroxy-3-methylxanthone14H10O4 (258.05282)CYellow needles, mp 277\u2013278 \u00b0Cmax (MeOH) (log \u03b5): 403 (3.54), 341 (4.01), 272 (4.58), 263 (4.40), 255 (4.47), 236 (4.32)UV \u03bbmax (KBr): 3461, 1661, 1633, 1606, 1591, 1497 cm\u22121IR \u03bdPyrenula japonica and Pyrenula pseudobufonia [Sources: Mycobiont of obufonia .1,8-Dihydroxy-5-methoxy-3-methylxanthone15H12O5 (272.06847)CYellow needles, mp 214\u2013215 \u00b0Cmax (MeOH) (log \u03b5): 393 (3.54), 340 (4.00), 272 (4.30), 254 (4.49), 235 (4.36)UV \u03bbmax (KBr): 3445, 1661, 1631, 1609, 1585, 1489 cm\u22121IR \u03bdPyrenula japonica and Pyrenula pseudobufonia [Sources: Mycobiont of obufonia .Hirtusneanoside40H46O17 (798.27350)CFaint yellow crystals, mp 231\u2013232 \u00b0Cc = 0.02 in MeOH)max (MeOH)(log \u03b5): 340 (3.24), 275 (4.01), 230 (4.52) nmUV \u03bbmax (CHCl3): 3290, 1735, 1620, 1590, 870 cm\u22121IR \u03bdUsnea hirta [Sources: ea hirta .Hirtusneanine34H36O13 (652.21559)CPale yellow crystals, mp 251\u2013253 \u00b0Cc = 0.01 in MeOH)max (MeOH)(log \u03b5): 338 (3.97), 275 (4.04), 231 (4.43) nmUV \u03bbmax (CHCl3): 3290, 1734, 1608, 1590, 870 cm\u22121IR \u03bdUsnea hirta [Sources: ea hirta .Umbilicaxanthoside A25H28O11 (504.16316)CYellow needles, mp 114 \u00b0Cmax (MeOH) (log \u03b5): 345 (3.90), 310 (4.15), 270 (4.05), 245 (4.50).UV \u03bbmax (KBr): 3320, 2945, 2905, 1643 cm\u22121IR \u03bdUmbilicaria proboscidea [Sources: boscidea .Umbilicaxanthone A19H18O6 (342.11034)CUmbilicaria proboscidea [Sources: boscidea .Umbilicaxanthoside B36H46O16 (734.27859)CPale yellow needles, mp 133 \u00b0Cmax (MeOH) (log \u03b5): 355 (4.06), 319 (4.01), 271 (3.96), 244 (4.08)UV \u03bbmax (KBr): 3350, 2950, 2900, 1640 cm\u22121IR \u03bdUmbilicaria proboscidea [Sources: boscidea .Umbilicaxanthone B24H26O6 (410.17294)CUmbilicaria proboscidea [Sources: boscidea .Linolenoylumbilicaxanthoside B54H74O17 (994.49260)CPale yellow needles, mp 133 \u00b0Cmax (MeOH) (log \u03b5): 355 (4.06), 319 (4.01), 271 (3.96), 244 (4.08)UV \u03bbmax (KBr): 3350, 2950, 2900, 1640 cm\u22121IR \u03bdUmbilicaria proboscidea [Sources: boscidea .Secalonic acid A32H30O14 (638.16356)CFaint yellow crystals, mp 231\u2013232 \u00b0Cc = 0.1 in CHCl3)max (MeOH)(log \u03b5): 338 (4.52), 260 (4.12), 228 (4.32) nmUV \u03bbmax (CHCl3): 3464, 1726, 1610, 1585, 1566, 1436, 1233, 1067 cm\u22121IR \u03bdDiploicia canescens [Sources: anescens .Spectral data: .c = 0.14 in CHCl3)Enantiomeric secalonic acid D: similar physicochemical data except Secalonic acid B32H30O14 (638.16356)CFaint yellow crystals, mp 231\u2013232 \u00b0Cc = 0.38 in CHCl3)max (MeOH)(log \u0190): 338 (4.52), 260 (4.12), 228 (4.32) nmUV \u03bbmax (CHCl3): 3582, 3014, 1747, 1611, 1589, 1214, 1058, 796, 726 cm\u22121IR \u03bdDiploicia canescens [Sources: anescens .Spectral data: Even though lichens offer the widest diversity of compounds in the fungal realm, the bioactivities of their secondary metabolites remain under-investigated in regards to any other fungi , especia"}
+{"text": "Cancer still is one of the leading causes of death and its death toll is predicted to rise further. We identified earlier the potential tumour suppressor zygote arrest 1 (ZAR1) to play a role in lung carcinogenesis through its epigenetic inactivation.ZAR1 is epigenetically inactivated not only in lung cancer but also across cancer types, and ZAR1 methylation occurs across its complete CpG island. ZAR1 hypermethylation significantly correlates with its expression reduction in cancers. We are also the first to report that ZAR1 methylation and expression reduction are of clinical importance as a prognostic marker for lung cancer and kidney cancer. We further established that the carboxy (C)-terminally present zinc-finger of ZAR1 is relevant for its tumour suppression function and its protein partner binding associated with the mRNA/ribosomal network. Global gene expression profiling supported ZAR1's role in cell cycle arrest and p53 signalling pathway, and we could show that ZAR1 growth suppression was in part p53 dependent. Using the CRISPR-dCas9 tools, we were able to prove that epigenetic editing and reactivation of ZAR1 is possible in cancer cell lines.We are the first to report that ZAR1 tumour suppressor in cancer.ZAR1 is a novel cancer biomarker for lung and kidney, which is epigenetically silenced in various cancers by DNA hypermethylation. ZAR1 exerts its tumour suppressive function in part through p53 and through its zinc-finger domain. Epigenetic therapy can reactivate the  ZAR1 was reported to be expressed in porcine and bovine brain and testis -dCas9 (100087), DNMT3A-dCas9 (100090). Epigenetic editing of endogenous ZAR1 was performed in the ZAR1 partially methylated Hela, if not mentioned otherwise. ZAR1 RNA guides are #1 ACTTTCGCTCACTTAGCCAG, #2 TGGTTCCCTTACGGATCAGC, #3 GTAGGGAGAAGGACGAAGAG, #4 GTCGCCTATTTAGGGTGCGG, #5 CGCGGCCACCAAGGGCAAGG, and #6 CCGCGGTACAGTGCTCGCTG and are positioned relative to TSS at \u2212402 #1, \u2212230 #2, \u2212133 #3, \u22123 #4, +120 #5, and +386 #6.CRISPR-Cas9 vector px549 was obtained from Lienhard Schmitz  and adapted for epigenetic editing by inactivation of Cas9 (dCas9 site-directed mutation). 2. I brief, beads were resuspended in two volumes urea buffer  and incubated shaking for 30 min at room temperature. Cysteins were alcylated at 55 mM final concentration of iodoacetamide, shaking at room temperature and in the dark for another 30 min. Peptidolysis was then initiated with 0.5 \u03bcg Lys-C  for 3 h shaking at room temperature, followed by dilution to 2 M urea/thiourea, addition of 0.5 \u03bcg trypsin (Serva) and an overnight shaking incubation at room temperature. Peptide-containing supernatants were brought to 1% NH3 and loaded onto three-layer SAX tips equilibrated previously with 30 \u03bcl of 0.1% NH3. After sequential washes with 30 \u03bcl 0.1% NH3 and 30 \u03bcl NH3 in 2-propanol, respectively, columns were syringe dried, peptides eluted using 30 \u03bcl 80% acetonitrile, 0.1% formic acid and vacuum dried. In-solution chemical labelling was performed as described [2 analysis used an in-house packed 70 \u03bcm ID, 15 cm reverse phase column emitter  with a buffer system comprising solvent A  and solvent B . Relevant instrumentation parameters are extracted using MARMoSET [ZAR1-EYFP, ZAR1delZF-EYFP vs. EYFP-empty were overexpressed in HEK293T cells (24 h), and pulldown was performed according to manufacturer's protocol by GFP Trap (ChromoTek). Triplicate sample pairs were processed by off-bead digest, strong anion exchange (SAX) extraction, and dimethyl-labelling, followed by LC-MSescribed , 72. PepMARMoSET  and inclMARMoSET , 75  APS = 16.2 (407) Avg = 12.8, Source: GEO ID: gse36133 Dataset Date: 2012-03-20. Additional file ZAR1 expression in cancer cell lines vs. normal tissues, 1555775_a_at, log2, data Roth vs. Broad, Anova one way. ZAR1 methylation in normal to tumour tissues and cancer cell lines, cg22773661/cg1753764, data Lokk vs. Heyn vs. Esteller, Anova one way. ZAR1 methylation in normal to tumour tissues and cancer cell lines relative to CpG island/shores and for all ZAR1 (cg) reporters from array. T-SNE analysis on Broad and Roth, Transform: zscore, no gene filter, no sample filter, perplexity = 50, Colour mode: Colour by Gene (ZAR1), Transform log 2. Overview on R2 used datasets : Normal Various - Roth - 504 \u2013 MAS5.0 - u133p2, Source: GEO ID: gse7307 Dataset Date: 2007-04-09; Cellline CCLE Cancer Cell Line Encyclopedia - Broad - 917 - MAS5.0 - u133p2, Source: GEO ID: gse36133 Dataset Date: 2012-03-20; Normal Tissues - Lokk - 70 - custom - ilmnhm450, Source: GEO ID: gse50192 Dataset Date: 2014-02-26; Tumor Types (landscape) - Heyn - 493 - custom - ilmnhm450, Source: GEO ID: gse76269 Dataset Date: 2017-06-07; Cellline Cancer Pharmacogenomic - Esteller - 1028 - custom - ilmnhm450, Source: GEO ID: gse68379 Dataset Date: 2016-07-05. Figure Wanderer [ZAR1, dataset project: TCGA, data type: 450k Methylation Array, for LUAD lung adenocarcinoma and KIRC Kidney renal clear cell carcinoma. Pan-cancer mRNA RNA-seq using KM Plotter [MethSurv [PhyloP; UCSC genome browser [BioEdit [NCBI RefSeq [Swiss-Model [PhosphoSitePlus [cBioPortal [String v11 [Gene expression, promoter methylation correlation, and Kaplan\u2013Meier calculations were performed using Platform , WandereWanderer , KM Plot Plotter \u201381, and MethSurv . Gene serv [GSEA . The fol37, data . ZAR1 exZAR1 15555_a_at AP Plotter , Tumor tMethSurv  on TCGA  browser  and BioE[BioEdit  matrix: I RefSeq . Swiss-Mss-Model  predictiioPortal , 90. FigZAR1 promoter (186 bp) were upper primer GGAGAAGGAYGAAGAGGGGTTTTT and lower primer TCCCCCAAAACCRCCATAAAC, and pyrosequencing primer was TGGTAGGAAGGGYGTGGAGG. Primers for RT-PCR were ZAR1 AGCTGGGCAAGGAGCGGCTG and GGTGGGGCCGTTTAGGGTCCA (264 bp), GAPDH TGGAGAAGGCTGGGGCTCAT and GACCTTGGCCAGGGGTGCTA (176 bp), ACTB CCTTCCTTCCTGGGCATGGAGTC and CGGAGTACTTGCGCTCAGGAGGA (226 bp), p21 CCTTGTGCCTCGGTCAGGGGAG and GGCCCTCGCGCTTCCAGGAC (183 bp), p27 GTGCGAGAGAGGCGGTCGTG and TCCACCGGGCCGAAGAGGTT (146 bp), WEE1 CACACGCCCAAGAGTTTGC and CACTTGAGGAGTCTGTCGCA (135 bp) and WEE1 3\u2032UTR primers are: pair 1 CTCCCCCTGAACACTGTGAC and ACTGACACCAATCGAGAAAGT (87 bp), pair 2 CACCAGCCTTTCCAGGGTTA and GGTCACTACAGGGAAAGACACC (92 bp), pair 3 AGCCTTCAATGTACCTGTGTGT and TGCCTACAAAGTGCTCCCAG (93 bp), pair 4 CTGGGAGCACTTTGTAGGCA and AGCAGCAAATTCACAAGGCA (77 bp), pair 5 AGTTTTGTCTTTGCTGTAAACTTGT and CATCAAAAGCAGCTATACATTTCAC (100 bp), pair 6 TGCACCCTTTCCCTCCTTTG and GTCCGGGAAGGACATTACCA (89 bp), pair 7 TGTTTTGCCCGGTTTTTCTCT and GTCAGAAGTCATTCTGGCATTTCA (95 bp), pair 8 TTTGCACTTGTCTTTGACTTGTGT and AGGTAAGCTCAGAGTGACTTTT (70 bp), pair 9 GCCATTTGACTAATAATACTGGCT and ACACAAGTCAAAGACAAGTGC (106 bp).Primers for CoBRA analysis of the Additional file 1: Figure S1. Overview ZAR1 genomic structure, expression pattern and GO-term correlation, Figure S2.ZAR1 promoter is hypermethylated across cancer cell lines. Figure S3.ZAR1 methylation in ovarian carcinoma. Figure S4. Epigenetic inactivation of ZAR1 across human cancers. Figure S5. ZAR1 is conserved across human, mouse, and xenopus with high C-terminal homology and structure prediction. Figure S6. Confirming human ZAR1 RNA binding ability. Figure S7. ZAR1 is cytosolic. Figure S8. Reexpression of ZAR1 alters the transcriptome and reveals association with mRNA 3\u2032end processing. Figure S9. ZAR1 function depends on its zinc-finger. Figure S10. ZAR1 binding partner GO-term association overview. Figure S11. ZAR1 binding partner GO-term association overview. Figure S12. Targeting the ZAR1 genomic region with CRISPR ZAR1 guide RNA oligos. Figure S13. Effective epigenetic editing of ZAR1 with distinct RNA guide combinations upstream the TSS."}
+{"text": "Escherichia coli 4s is a Gram-negative bacterium found in the equine intestinal ecosystem alongside diverse other coliform bacteria and bacteriophages. This announcement describes the complete genome of the T7-like E. coli 4s podophage Penshu1. From its 39,263-bp genome, 54 protein-encoding genes and a 179-bp terminal repeat were predicted. Escherichia coli 4s is a Gram-negative bacterium found in the equine intestinal ecosystem alongside diverse other coliform bacteria and bacteriophages. This announcement describes the complete genome of the T7-like E. coli 4s podophage Penshu1. From its 39,263-bp genome, 54 protein-encoding genes and a 179-bp terminal repeat were predicted. Escherichia coli is a Gram-negative bacterium found living among the intestinal microbiome of all mammals. Due to multiple protective abilities, including extreme acid resistance, E. coli colonizes the intestine as a commensal . After trimming using FastX-Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit/), they were assembled into a single contig at 376.4-fold coverage using SPAdes v3.5.0   wastewater treatment sample from Bryan, TX, using the host . Followithe host . The phathe host . The DNAs v3.5.0 . Contig xy-pub/) , 10. Proxy-pub/) , 12. No xy-pub/) . Protein-pub/) 1417. Rho-i-pub/) 14. Genome--pub/) 14.Escherichia phage ST31 (GenBank accession number KY962008) and 80.1% nucleotide sequence identity with Escherichia phage YZ1 (GenBank accession number MG845865). As for phage T7, Penshu1 has a slippery sequence in the major capsid protein (NCBI accession number QEG09806) that can lead to translation by frameshift of the minor capsid protein (NCBI accession number QEG09807).Penshu1 has a 39,263-bp genome, with a 93.4% coding density and 50.6% G+C content. Analysis predicted 54 protein-coding genes, with 30 being assigned a putative function. The Penshu1 genome was reopened at T7-like direct terminal repeats of 179 bp predicted by PhageTerm . Penshu1MK903281, BioProject accession number PRJNA222858, SRA accession number SRR8893626, and BioSample accession number SAMN11414580.The genome sequence and associated data for phage Penshu1 were deposited under GenBank accession number"}
+{"text": "HERC1 and HERC2, are present in most metazoan taxa. They encode very large proteins  that contain more than one RCC1-like domain as a structural characteristic. Accumulating evidences show that these unusually large proteins play key roles in a wide range of cellular functions which include neurodevelopment, DNA damage repair, and cell proliferation. To better understand the origin, evolution, and function of the Large HERC family, this minireview provides with an integrated overview of their structure and function and details their physiological implications. This study also highlights and discusses how dysregulation of these proteins is associated with severe human diseases such as neurological disorders and cancer.Homologous to the E6AP carboxyl terminus (HECT) and regulator of chromosome condensation 1 (RCC1)-like domain-containing proteins (HERCs) belong to the superfamily of ubiquitin ligases. HERC proteins are divided into two subfamilies, Large and Small HERCs. Despite their similarities in terms of both structure and domains, these subfamilies are evolutionarily very distant and result from a convergence phenomenon rather than from a common origin. Large HERC genes,  Proteins containing a HECT domain are ubiquitin ligases (E3). These enzymes participate in the ubiquitylation process accepting ubiquitin from a ubiquitin-conjugating enzymes (E2) and catalysing its transfer to the protein to be ubiquitylated . In animAlthough traditionally classified together with the Small HERC proteins, Large and Small HERCs form two distant protein families . Large HMonosiga brevicollis and Salpingoeca rosetta, the emergence of HERC1 occurred in Metazoa. Both proteins are already present in the placozoan Trichoplax adhaerens and in most metazoan phyla, with the absence of HERC1 in certain insect clades . HERC2 induces BRCA1 degradation in breast cancer. This is inhibited either by binding of TUSC4 to HERC2  or by BAHERC1 deletions affect MSH2 protein levels  and certain HERC2 SNPs can interfere OCA2\u2019s expression thus affecting eye, skin, and hair pigmentation , for HERC1: T. adhaerens (XP_002116356.1), Anoplophora glabripennis (Asian long-horned beetle) (XP_018567016.1), Callorhinchus milii (Elephant shark) (XP_007906088.1), Latimeria chalumnae (Coelacanth) (XP_005987767.1), Danio rerio (Zebrafish) (XP_021333511.1), Oreochromis niloticus (Tilapia) (XP_013121789.1), Xenopus tropicalis (Frog) (XP_004916026.1), Gekko japonicus (Gecko) (XP_015275623.1), Gallus gallus (Chicken) (XP_004943789.1), Ornithorhynchus anatinus (Platypus) (XP_028921613), Monodelphis domestica (Opossum) (XP_016284147.1), Sarcophilus harrisii (Tasmanian devil) (XP_023353554.1), Equus caballus (Horse) (XP_023471806.1), Rattus norvegicus (Rat) (XP_017451567.1), Mus musculus (Mouse) (NP_663592.3), Homo sapiens (Human) (NP_003913.3), Pan troglodytes (Chimpanzee) (XP_001174017.1), Canis lupus familiaris (Dog) (XP_544717.3), Felis catus (Cat) (XP_023110993.1), and Lepidosiren paradoxa (Lungfish) . For HERC2: M. brevicollis (XP_001743304.1), S. rosetta (XP_004996155.1), T. adhaerens (XP_002112421), Asian long-horned beetle (XP_018562376.1), Drosophila melanogaster (Fruit fly) (NP_608388.2), Apis mellifera (Honey bee) (XP_01677273), Coelacanth (XP_014343905.1), Zebrafish (XP_021332870.1), Tilapia (XP_013130527.1), Frog (XP_012813156.1), Gecko (XP_015266462.1), Chicken (XP_015133197.1), Platypus (XP_016083014.1), Opossum (XP_007501467.1), Tasmanian devil (XP_023356163.1), Horse (XP_023507879.1), Mouse (NP_001347009.1), Rat (XP_006229362.1), Human (NP_004658.3), Chimpanzee (XP_024204823.1), Dog (XP_022272508.1), Cat (XP_023110801.1), Elephant shark (XP_007889299.1), and Lungfish .Amino acid sequences were aligned using the Mafft FFT-NS-i algorithm . The phyHuman RCC1 (NCBI accession number P18754) and HECT domain from E6AP  three-dimensional structures were modelled using Swiss-Model online software . RLDs anJG-C and JR conceived and designed the manuscript. JG-C, AM-M, and JR analysed the data and performed figures and tables. JG-C, AM-M, JS-G, LP, and JR wrote the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Staphylococcus aureus clones and genes encoding antimicrobial resistance and toxins were examined among 120 S. aureus strains from nosocomial infections in tehran, Iran.In the current research, the prevalence of mec, agr, spa, and MLST, the isolates were typed. Antimicrobial susceptibility was examined, based on disk diffusion and PCR method to identify resistance and toxin-encoding genes. Based on the polymorphisms in SCCS. aureus isolates, 85 (70.8%) were methicilin resistant S.aureus (MRSA), and 35 (29.2%) were methicilin sensetive S. aureus (MSSA). The tested isolates contained resistance genes, including ant(4\u0384)-Ia (90%), aac(6\u0384)-Ie/aph(2\u02dd) (80%), aph(3\u0384)-IIIa (30%), erm(A) (26.7%), erm(B) (10.8%), erm(C) (11.7%), msr(A) (40.8%), msr(B) (14.2%), tet(M) (45.8%), and mupA (8.3%). The MRSA strains were clustered into six different clones. The most common genotypes included ST239-SCCmec III/t037 (23.3%), ST239-SCCmec III/t388 (22.5%), ST22-SCCmec IV/t790 (8.3%), ST15-SCCmec IV/t084 (7.5%), ST585-SCCmec III/t713 (5%), and ST239-SCCmec III/t924 (4.2%), respectively. ST182/t196 (8.3%) and ST123/t171 (5%) belonged exclusively to MSSA strains. Overall, 10 (66.7%) and 5 (33.3%) out of 15 isolates with pvl genes were attributed to clones ST22-SCCmec IV/t790 and ST15-SCCmec IV/t084, respectively. ST22-SCCmec IV/t790, ST239-SCCmec III/t037, and ST15-SCCmec IV/t084, were related to high-level mupirocin-resistant phenotypes. Among 120 S. aureus was confirmed in our hospitals, and ST239-SCCmec III/t037 showed a relatively high prevalence in our study. It seems that assessment of resistance and virulence genes in different S. aureus molecular types is necessary for proper antibiotic consumption.The genetic diversity of  Staphylococcus aureus, which is described as a common nosocomial pathogen, is responsible for various diseases, such as food poisoning, osteomyelitis, wound infections, and even fatal conditions, such as endocarditis (S. aureus (MRSA) occurred in the UK  is recognized as a staphylococcal cassette chromosome mec (SCCmec). Generally, SCCmec is categorized into 11 types with respect to mec genes and ccr gene complexes  infections. HA- and CA-MRSA strains can be distinguished with respect to some genotypic, phenotypic, and epidemiological characteristics, as well as virulence factors  genes have significantly limited the availability of antibiotics over the past decades. In addition, the growing emergence of MDR-MRSA strains poses a major global health concern . Wide remupA and mupB genes are responsible for resistance to mupirocin which is used to treat various types of skin diseases caused by S. aureus  including aminoglycoside nucleotidyltransferases, aminoglycoside phosphotransferases and aminoglycoside acetyltransferases . mupA an. aureus . Resistaing site . Thus, i spa, SCCmec and agr techniques in clinical samples taken from patients in Tehran, Iran.This study was conducted to identify antibiotic resistance patterns and the carriage of resistance and virulence genes as well as major MRSA clones by MLST,Sampling, MRSA isolation and antibacterial susceptibility testingThis cross-sectional study included 368 clinical samples from wound, blood, and urine specimens during April-December 2016. Ethics Committee of Shahid Beheshti University of Medical Sciences approved the implementation of this study (IR.SBMU.SM.REC.1395.157). All patients signed written informed consent forms.S. aureus identification was performed, based on the conventional biochemical tests. S. aureus identification was confirmed based on the PCR assay for nucA gene , clindamycin (CD 2 \u00b5g), ciprofloxacin (CIP 5 \u00b5g), trimethoprim- sulfamethoxazole (TS 2.5 \u00b5g), kanamycin (K 30 \u00b5g), ceftriaxon (CRO 30 \u00b5g), quinupristin-dalfopristin (SYN 15 \u00b5g), erythromycin (E 15 \u00b5g), amikacin (AK 30 \u00b5g), gentamicin (GM 10 \u00b5g), tobramycin (TN 10 \u00b5g), teicoplanin (TEC 30 \u00b5g), penicillin (PG 10 \u00b5g), and linezolid (LZD 30 \u00b5g)  were determined, based on the CLSI criteria  were considered as the reference strains for the quality control purposes.Using E-test strips (bioMe\u00b4rieux), the minimum inhibitory concentrations (MICs) were measured for mupirocin and vancomycin. Resistance to three antibiotic groups or more, besides beta-lactams, was defined as MDR. High mupirocin resistance was defined as antibiotic use \u2265 256 mg/l. Growth in a well containing clindamycin and erythromycin  indicated inducible macrolide-lincosamide-streptogramin B and/or clindamycin resistance phenotypes; otherwise, constitutive MLSB and/or clindamycin resistance phenotype was confirmed . ATCC292Extraction of genomic DNAS. aureus cultures were used on 5% sheep blood agar , based on the protocols of InstaGene Matrix kit . For extracting genomic DNA, pure overnight Resistance and toxin genes profilingetb, tst, pvl, eta) and resistance (tet(M), aac (6\u0384)-Ie/aph (2\u02dd), mupA, erm(A), msr(A), msr(B), erm(B), erm(C), ant (4\u0384)-Ia, aph (3\u0384)-IIIa) genes, PCR assay was carried out. The details of the degenerated primers in this study are described in To identify toxin  and reverse (agr1 to agr4) primers for the agr groups as previously recommended by Gilot et al . Chromas 1.45  was used to edit the sequences. To assign the sequences to specific spa types, the Ridom SpaServer database was searched.On the other hand, lleagues . After tMLST techniqueS. aureus isolates. The internal fragments of housekeeping genes were used to identify the allelic profiles; these genes included gmk, arcC, aroE, glpF, pta,yqiL, and tpi. The isolate was assigned a sequence type (ST) after comparing the sequences with the S. aureus MLST database.Via amplification and sequencing, MLST was carried out on Sampling and antibiotic susceptibility S. aureus. These isolates originated from wound (60%), blood (20.8%) and urinary tract infections (19.2%). Of the 120 S. aureus clinical isolates obtained from the hospitalized patients, 85 (70.8%) were MRSA and 35 (29.2%) were methicillin susceptible S. aureus (MSSA). In this study, out of 368 samples obtained from various clinical specimens, 120 isolates (83 (69.2%) obtained from men and 37 (30.8%) from women) were identified as  in vitro antimicrobial susceptibility tests. All isolates were susceptible to vancomycin, among which 55 (45.8%), 48 (40%) and 17 (14.2%) isolates had a MIC of 0.5, 1 and 2 \u00b5g/ml, respectively. None of the isolates were susceptible to all of the antimicrobial agents tested regardingP= 0.145 and 0.128, respectively). The frequency of resistance for MRSA and MSSA isolates to different antibacterial agents are presented in Among the 35 MSSA isolates, no mupirocin resistance was detected, whereas 30 MRSA isolates (35.3%) were mupirocin resistant. Of these mupirocin resistant isolates, 14 (46.7%) and 16 (53.3%) had high and low resistance levels, respectively. All the high-level mupirocin-resistant (HLMUPR) isolates were collected from wound samples. Patients diagnosed with mupirocin-susceptible and -resistant MRSA infections were not significantly different in terms of age and gender  were defined as MDR. The predominant multiple drug resistance profile among the MDR isolates were resistance to 9 and 7 antibiotics found in 75 (62.5%) and 12 (10%) isolates, respectively. Distribution of resistance profile and different clinical sample in S. aureus isolated from nosocomial infections are presented in Of the 120 The distribution of resistance genesant(4\u0384)-Ia (90%), aac(6\u0384)-Ie/aph(2\u02dd) (80%), aph(3\u0384)-IIIa (30%), erm(A) (26.7%), erm(B) (10.8%), erm(C) (11.7%), msr(A) (40.8%), msr(B) (14.2%), tet(M) (45.8%), and mupA (8.3%) (ant(4\u0384)-Ia, aac(6\u0384)-Ie/aph(2\u02dd), and mecA genes. Other detected antibiotic resistance genes included tet(M) (64.7%), msr(A) (57.6%), aph(3\u0384)-IIIa (42.3%), erm(A) (37.6%), msr(B) (20%), erm(C) (16.5%), erm(B) (15.3%), and mupA (11.8%), while 31.4% and 56.7% of MSSA strains were found to respectively carry aac(6\u0384)-Ie/aph(2\u02dd) and ant(4\u0384)-Ia. Particularly, the MRSA strains contained more resistance genes, compared to the MSSA isolates.In addition, the frequency of antibiotic resistance genes was measured. The genes included A (8.3%) . The MRSS. aureus isolates were identified as HLMUPR-MRSA, while mupA gene was confirmed in 10 isolates (71.4%). In addition, ant(4\u0384)-Ia , followed by aac(6\u0384)-Ie/aph(2\u02dd) , was recognized as the most common aminoglycoside resistance gene. A total of 75 (62.5%) isolates contained ant(4\u0384)-Ia, as well as aac(6\u0384)-Ie/aph(2\u02dd). On the other hand, 15 (12.5%) isolates harbored aph(3\u0384)-IIIa, besides ant(4\u0384)-Ia. Also, eleven (9.2%) isolates harbored ant(4\u0384)-Ia, aph(3\u0384)-IIIa, and aac(6\u0384)-Ie/aph(2\u02dd) genes, while 10 (8.3%) contained aac(6\u0384)-Ie/aph(2\u02dd), as well as aph(3\u0384)-IIIa; however, ant(4\u0384)-Ia alone was confirmed in 7 (5.8%) strains. Resistance to tetracycline was observed among 84 (70%) S. aureus isolates, 55 (45.8%) of which harbored tet(M) gene. Antibiotic resistance genes showed the highest prevalence among MRSA strains from wound infections.Fourteen (11.7%) out of 30 mupirocin-resistance Virulence gene profilingtst  and etb  genes, respectively . S. aureus isolates harbouring tst gene were isolated from wound , blood , and urinary tract infections  while pvl positive isolates were detected in wound  and blood  samples. Eight isolates carrying pvl and tst genes, simultaneously, had mupA gene.Among toxin-encoding genes, the highest and lowest frequencies were attributed to ectively . In presDistribution of SCCmec typesAccording to SCCmec typing, 66 (77.6%) and 19 (22.4%) MRSA isolates contained SCCmec types III and IV, respectively. No isolate harbored SCCmec type V, II, or I. Based on the multiplex PCR, isolates positive for PVL were attributed to SCCmec type IV, while tst gene was found in MRSA isolates from SCCmec III and IV. The HA-MRSA origin was emphasized by the presence of SCCmec type III.Frequency of agr typesagr typing indicated that type I was the predominant agr type present in 91 isolates (75.8%), followed by type III which was present in 20 isolates (16.7%). agr type II was detected in 9 isolates (7.5%). Among 35 MSSA isolates, 20 and 15 harboured agr types I and III respectively .spa typingS. aureus isolates. spa typing discriminated eight different types: t037 (23.3%), t388 (22.5%), t713 (16.8%), t924 (8.3%), t790 (8.3%), t196 (8.3%) t084 (7.5%) and t171 (5%)  . All theMLSTmec III/t037 was found to be the most prominent MRSA clone identified in this study. Distribution of MRSA clones isolated from nosocomial infections are presented in Apparently, 120 isolates belonged to six different STs including ST239 (65 strains), ST22 (10 strains), ST182 (10 strains), ST15 (9 strains), ST585 (20 strains) and ST123 (6 strains). ST239, ST585, ST182 and ST123 were found in MSSA strains. It should be noted that of these STs, ST182 and ST123 belonged exclusively to MSSA strains. In conclusion, MRSA strains are clustered into six different groups. ST239-SCCmec IV/t790  and ST15-SCCmec IV/t084  clones. Among the isolates under study, 58 (48.3%) isolates harboring tst-1 were distributed in ST239-SCCmec III/t037 , ST239-SCCmec III/t388 , ST15-SCCmec IV/t084 , ST585-SCCmec III/t713 , ST239-SCCmec III/t924 , and ST22-SCCmec IV/t790  clones. Among examined isolates, nine isolates (7.53%) were found to carry the eta gene. The eta positive isolates were distributed in ST15-SCCmec IV/t084 , ST22-SCCmec IV/t790  and ST239-SCCmec III/t037  clones. The etb gene was detected in ST585-SCCmec III/t713  and ST22-SCCmec IV/t790  clones. MRSA clones resistance profile varied. Resistance to mupirocin was detected in all the MRSA clones with the exception of ST585-SCCmec III/t713 clone. Interestingly, mupirocin resistant MSSA isolates belonged to ST182/spa type t196. HLMUPR-MRSA strains were detected in ST22-SCCmec IV/t790 , ST15-SCCmec IV/t084  and ST239-SCCmec III/t037  clones. Fifteen PVL-carrying strains in our study belonged to ST22-SCCB phenotype was detected in ST239-SCCmec III/t037 , ST239-SCCmec III/t388 , ST585-SCCmec III/t713 , ST15-SCCmec IV/t084 , ST239-SCCmec III/t924  and ST22-SCCmec IV/t790  while iMLSB phenotype distributed among 3 major clones ST239-SCCmec III/t388 , ST22-SCCmec IV/t790  and ST15-SCCmec IV/t084 . Of 12 MSSA isolates with cMLSB phenotype, 5 isolates belonged to ST182/t196, 4 isolates belonged to ST239/t924, 2 isolates belonged to ST585/t713 and 1 isolate belonged to ST123/t171. All the iMLSB phenotype among MSSA strains belonged to ST585/t713. Characteristics of MRSA clones are presented in cMLSet al. /aph(2\u2032\u2032) gene (40.3% ) in comparison with other AME genes, in the present study, ant(4\u0384)-Ia (90%) was the most frequent AME gene in S. aureus isolates. However, in this study, aph(3\u0384)-IIIa gene frequency rate was higher than the study reported by Ida et al. (8.9%)  and Margtudy (aac\u2032/aph(2\u2032\u2032S. aureus strains (S. aureus isolates were found to be lower. We detected the resistance gene mupA in 10 isolates (8.3%) which is lower than 12.6%  gene that may cause tetracycline resistance was detected in 55 (45.8%) isolates. In accordance with the results of our previous study  a high tet al. (7%) reported low frequency of iMLSB phenotype among S. aureus isolates as well  presented cMLSB phenotype while the frequency of iMLSB phenotype was 10% (12/120). In consistent with our findings, Schreckenberger  as well . Accordiospitals . They fo. in USA  reportedn survey , the fre ) or transposons (erm(A) and erm(B)) and msr genes expressing active efflux pumps mainly msr(A)) (msr(A), erm(A), msr(B),erm(C), and erm(B) genes to be 40.8%, 26.7%, 14.2%, 11.7%, and 10.8% respectively. In contrast to Rashidi Nezhad\u2019s study (erm(A) gene as the predominant gene among the isolates with inducible phenotype and erm(C) among the isolates with the constitutive phenotype, our finding revealed that the msr(A) gene was the most common gene among strains with the constitutive phenotype , followed by erm(A) , erm(C) , erm(B) , msr(B)  while erm(A) , erm(B) , erm(C) , msr(A)  and msr(B)  were much more common among the isolates with inducible phenotype. It is worth mentioning that the frequency of erm and msr genes depends on the bacterial species as well as the geographic region in which the study is carried on. As previously mentioned, resistance to macrolides is encoded by genes often carried on plasmids ( msr(A)) . Our res\u2019s study  who repomec (77.6%), according to the results of SCCmec typing, associated with an MDR pattern among MRSA isolates. This is in consistence with the results reported by Japoni and colleagues , type III (16.7%), and type II (7.5%), respectively. Goudarzi et al. . This spa type was reported from Saudi Arabia, China, Iran as well as among HA-MRSA isolates found in Europe, America and other regions of Asia . The review of the literature reveals that the multiresistant ST239 clone is responsible for at least 90% of HA-MRSA infections in Europe, United States, and some Asian countries, including Kuwait and Malaysia  was the second most commonly detected MRSA clone. A similar result was reported by Ohadian Moghadam and ST30 .mec IV/t790 (8.3%). This clone was associated with high resistance to mupirocin, carrying resistance genes, including mecA, msr(A), ant(4\u0384)-Ia, msr(B), tet(M), and mupA. In our study, all ST22-SCCmec IV/t790 strains contained pvl genes. There are reports of S. aureus ST22 harboring pvl gene from Iran  was the fourth most commonly detected clone. The low frequency of this clone has been previously reported in 16 European countries . The presence of eight different spa types, i.e., t037, t388, t713, t924, t790, t196, t084, and t171, was also confirmed. For the first time in Iran, STs 182 and 123, as well as spa types t196 and t171, were detected, which might be indicative of the emergence of new clones. Further studies on other neighboring regions, focusing on the emergence of new circulating clones, are necessary to reach an overall understanding of dynamic MRSA clones in Iran and the Middle East.To sum up, the present findings showed that MRSA isolates have various genetic backgrounds in our hospitals and involve six major clones. Certain molecular types were associated with some resistance and virulence genes (e.g.,"}
+{"text": "Macrophages exposed to the Th2 cytokines interleukin (IL) IL-4 and IL-13 exhibit a distinct transcriptional response, commonly referred to as M2 polarization. Recently, IL-4-induced polarization of murine bone marrow-derived macrophages (BMDMs) has been linked to acetyl-CoA levels through the activity of the cytosolic acetyl-CoA-generating enzyme ATP-citrate lyase (ACLY). Here, we studied how ACLY regulated IL-4-stimulated gene expression in human monocyte-derived macrophages (MDMs). Although multiple ACLY inhibitors attenuated IL-4-induced target gene expression, this effect could not be recapitulated by silencing ACLY expression. Furthermore, ACLY inhibition failed to alter cellular acetyl-CoA levels and histone acetylation. We generated ACLY knockout human THP-1 macrophages using CRISPR/Cas9 technology. While these cells exhibited reduced histone acetylation levels, IL-4-induced gene expression remained intact. Strikingly, ACLY inhibitors still suppressed induction of target genes by IL-4 in ACLY knockout cells, suggesting off-target effects of these drugs. Our findings suggest that ACLY may not be the major regulator of nucleocytoplasmic acetyl-CoA and IL-4-induced polarization in human macrophages. Furthermore, caution should be warranted in interpreting the impact of pharmacological inhibition of ACLY on gene expression. Macrophages respond to changes in their environment, such as bacterial or viral infection, hormones, cytokines, or nutrients, with remodeling their transcriptome. Consequently, they alter their phenotype, a response known as macrophage polarization . Historide novo lipogenesis and thus, cell proliferation  . IL-4 trSince we previously noticed considerable differences in metabolic requirements of human vs. murine macrophages toward IL-4-induced polarization , we quesHuman peripheral blood mononuclear cells were isolated from commercially obtained buffy coats  using Ficoll  density centrifugation. Monocytes were separated from lymphocytes by adherence to plastic after 1 h-incubation in serum-free medium, and differentiated into macrophages by culture in RPMI-1640 medium  containing 3% heat-inactivated AB-positive human serum for 7\u201310 days. THP-1 cells were purchased from ATCC (cat. no. TIB-202) and maintained in RPMI-1640 medium containing 10% fetal bovine serum, penicillin and streptomycin. THP-1 cells were differentiated to macrophages by 24 h-treatment with 50 nM phorbol myristate acetate  followed by overnight culture in serum-free medium. Unless indicated otherwise, cells were treated with 5 \u03bcM BMS-303141 (cat. no. 4609), 25 \u03bcM SB 204990 (cat. no. 4962) (both Tocris), 100 \u03bcM MEDICA 16 (cat. no. M5693), 20 mM hydroxycitrate (cat. no. 59847), 5 mM sodium acetate (cat. no. S2889), 5 mM sodium octanoate (cat. no. C5038), 1,2,3-benzene-tricarboxylic acid (cat. no. 51520), CTP inhibitor (cat. no. SML0068) , 5 \u03bcg/mL TOFA (cat. no. 10005263) (Cayman), 20 ng/mL IL-4 (cat. no. 200-04), or IL-13 (cat. no. 200-13) (Peprotech).\u2212(Ct(target)\u2212Ct(reference))] using expression of \u03b22-microglobulin or GAPDH as reference genes.Total RNA from macrophages was isolated using PeqGold RNAPure kit  followed by reverse transcription using cDNA Synthesis kit . Quantitative real time PCR (Q-PCR) was carried out on a CFX96 system from Bio-Rad using iQ SYBR green . Primer sequences are listed in Supplementary Table Total cell lysates were prepared by scraping the cells into Laemmli buffer  containing protease inhibitors  followed by sonification. For histone extraction, cell pellets were lyzed in PBS containing 0.5% Triton X-100 , 5 mM sodium butyrate , and protease inhibitors for 10 min on ice. Lysates were centrifuged 0.5 min at 16,000 g, 4\u00b0C, and pellets containing nuclei were incubated with 0.2 M HCl  overnight. After centrifugation of insoluble material for 10 min at 500 g, 4\u00b0C, supernatants were neutralized with NaOH , mixed with 5x Laemmli buffer and heated 5 min at 95\u00b0C.Protein lysates from macrophages were run on 7.5\u201315% polyacrylamide gels and blotted on nitrocellulose membranes. Following primary antibodies were used: pSTAT6 (Y641) (cat. no. #9361), STAT6 (clone D3H4) (cat. no. #5397), pSTAT3 (Y705) (cat. no. #9131), STAT3 (clone 124H6) (cat. no. #9139), pACLY (S454) (cat. no. #4331), histone H3 acK14 (clone D4B9) (cat. no. #7627), histone H3 acK23 (clone D6Y7M) (cat. no. #14932), DDK tag (clone 9A3) (cat. no. #8146) , ACLY , histone H3 acK9 (clone Y28) , histone H3 acK27 , histone H3 , nucleolin . Membranes were incubated with IRDye 700/800-coupled secondary antibodies, scanned and quantified using Odyssey imaging system (Licor).Silencing of ACLY was performed using siGENOME SMARTpool  at 50 nM and Hyperfect transfection reagent . Cells were treated 96 h post-transfection.Human MDMs were transfected with 1 \u03bcg DDK-tagged ACLY  or GFP-encoding plasmids  using Viromer Red transfection reagent  according to manufacturer's instructions. Twenty-four hours after transfection, cells were stimulated with 20 ng/mL IL-4 for 24 h.13C2-acetyl-CoA . Acetyl-CoA concentration was determined by reference to the standard. Analyst 1.6.2 and MultiQuant 3.0 (both Sciex), were used for data acquisition and analysis, respectively. Acetyl-CoA amounts in the sample were normalized to sample DNA concentration measured following incubation with H\u00f6chst 33342 fluorescent DNA dye  on a Tecan fluorescence plate reader.Cells were rapidly washed with saline, and metabolism was quenched by putting the dishes into liquid nitrogen. Cells were scraped into methanol:water (5:3) mix on ice/dry ice followed by addition of cold chloroform , and vortexing for 10 min at 4\u00b0C. Aqueous phase was separated, evaporated, and re-suspended in 10% methanol . Acetyl-CoA concentration was determined with a Sciex QTrap5500 mass spectrometer operating in multiple reaction monitoring mode in positive electrospray ionization mode. Chromatographic separation was performed on an Agilent 1290 Infinity LC system (Agilent) using an Acquity HSS T3 column. The mobile phase consisted of (A) water, 10 mM ammonium formate , 0.01% ammonia, and (B) methanol, 10 mM ammonium formate, 0.01% ammonia. Elution of analytes was carried out under gradient conditions at a flow rate of 0.3 mL/min going from 2% B to 70% B in 5 min, increasing to 95% B in 1 min, hold 95% B for 0.5 min, and equilibrate at 2% B for 2.5 min. Calibration curve was performed with an authentic standard. All samples and dilutions of the standards were spiked with heavy isotope labeled internal standard containing pLentiCRISPRv2 (Addgene: cat. no. #52961) vectors harboring different sgRNAs designed using the benchling software package Table . Cell-frpost-hoc means comparison using GraphPad Prism. Differences were considered statistically significant at p < 0.05.Data are presented as means \u00b1 S.E. of at least three independent experiments. Data were analyzed by one-way analysis of variance (ANOVA) with Bonferroni Investigations were conducted in accordance with the ethical standards and according to the Declaration of Helsinki and to the national and international guidelines and have been approved by the authors' institutional review board. The ethics committee of Goethe-University waived the necessity of written informed consent when using the buffy coats from anonymized blood donors.To investigate the role of ACLY in IL-4-stimulated human macrophage polarization we initially analyzed mRNA expression of arachidonate 15-lipoxygenase (ALOX15) in MDMs in the presence of pharmacological ACLY inhibitors BMS 303141 , SB 2049A previous study showed that ACLY was necessary for IL-4-induced expression of a subset of IL-4 target genes in murine macrophages . To asseNext, we assessed the effect of silencing ACLY mRNA expression on IL-4-induced MDM polarization. For this, we treated MDMs for 96 h with control or ACLY siRNAs prior to 24 h-treatment with IL-4. Surprisingly, a knockdown of ACLY failed to reproduce the effect of ACLY inhibitors on IL-4-stimulated gene expression Figures , despiteConsidering these discrepancies, we decided to investigate the impact of ACLY inhibitors on human macrophage metabolism in more detail. Surprisingly, we did not find any differences in the levels of acetyl-CoA in cells treated with BMS 303141 or SB 204990 for 24 h Figure . AccordiWe then investigated the influence of ACLY inhibitors on initial steps of IL-4-induced signal transduction by pre-incubating MDMs with inhibitors for 1 h followed by 0.5 h-treatment with IL-4. ACLY inhibitors did not affect IL-4-triggered tyrosine phosphorylation of STAT6 with the exception of BMS 303141 and hydroxycitrate Figure . We notiMurine BMDMs responded to IL-4-stimulation with an Akt-dependent increase of ACLY phosphorylation at Ser454, which exhibited a delayed kinetics as compared with IL-4-induced Akt phosphorylation . We follde novo lipogenesis . Thus, the role of ACLY in metabolism and epigenetic regulation of terminally differentiated human macrophages remains unclear and warrants further research. Of note, ACLY may have greater impact during human macrophage differentiation, since this process is characterized by a temporary rise of de novo lipogenesis . ACSS2 contributes to histone acetylation in cancer cells, especially under hypoxia or glucose deprivation , 38, andOur results also highlight the notorious proneness of pharmacological inhibitors to off-target effects, which is in our case particularly remarkable, since we used structurally dissimilar substances . Off-target effects of ACLY inhibitors on IL-4-stimulated gene transcription could only be revealed using ACLY knockout cell line. Our findings also point to the limitation of studying human primary cells, since ACLY knockdown leaves substantial residual activity left whereas pharmacological inhibitors are unsuitable due to off-target activities. Whether induced pluripotent stem cell\u2014derived macrophages, where creation of knockout models is possible , will beDN designed the study, performed the experiments, and wrote the manuscript. SZ and IF contributed to acetyl-CoA measurements. FS, NK, and DF contributed to CRISPR/Cas9 THP-1 knockout cell line creation. BB contributed to study design and edited the final manuscript. All authors contributed to manuscript revision, read, and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Coastal regions experiencing declining dissolved oxygen are increasing in number and severity around the world. However, despite the importance of microbial metabolism in coastal hypoxia, few metagenomic surveys exist. Coastal regions experiencing declining dissolved oxygen are increasing in number and severity around the world. However, despite the importance of microbial metabolism in coastal hypoxia, few metagenomic surveys exist. Our data set from within the second largest human-caused hypoxic region provides opportunities to more deeply explore the microbiology of these systems. The northern Gulf of Mexico experiences seasonal bottom water hypoxia that can exceed 22,000\u2009km2 (http://www.noaa.gov/media-release/gulf-of-mexico-dead-zone-is-largest-ever-measured), impacting fisheries and other coastal industries (Marine systems suffering from declines in dissolved oxygen (DO) are becoming more numerous across the globe . Many codustries ; howeverActinobacteria (n = 12), Alphaproteobacteria (n = 5), Bacteroidetes (n = 6), Gammaproteobacteria (n = 3), Gemmatimonidetes (n = 1), Ignavibacteriae (n = 2), Nitrospina (n = 5), Planctomycetes (n = 7), Proteobacteria (n = 1), Synechococcus (n = 3), Verrucomicrobia (n = 3), and unclassified (n = 2). Twenty-four of the MAGs were estimated at >50% complete, with 16 estimated at >75% complete based on CheckM (https://doi.org/10.6084/m9.figshare.6911729.v1). All but 2 MAGs had estimated contamination of <9%, with 40 having estimated contamination of <3%. While several of our Actinobacteria MAGs grouped with sequences near the important OM1 clade of marine Actinobacteria (8https://doi.org/10.6084/m9.figshare.6911729.v1), but make useful references for future studies of the group.Site selection, marine chemistry metadata, and extraction, sequencing, and assembly methods were previously described , 4. Brieetes n = , Proteobacteria 810, theseeria n = , Gemmatihttps://doi.org/10.6084/m9.figshare.6911729.v1). Nitrospina MAGs were the only ones with predicted nitrite-oxidizing metabolism, matching observations from 2012 , including dissimilatory nitrate reduction to ammonium, and a few had predicted capacity for sulfur lithotrophy and possible autotrophy  (see Table S1 at https://doi.org/10.6084/m9.figshare.6911729.v1). Future comparisons of these data with those from other low-DO systems will illuminate common functional features associated with hypoxia and also provide information about biogeographic distinctions among taxa associated with these regimes.Metabolic reconstruction and carbohydrate-active enzyme (CAZyme) prediction were also completed as described  database under the accession numbers tandards . The ann"}
+{"text": "Serratia marcescens strains cause serious nosocomial infections in humans. Here, we present the annotated genome sequence of S. marcescens podophage Pila. Similar to its closest relative, Enterobacteria phage T7, Pila has a 38,678-bp genome, predicted to encode 51 protein-coding genes, and contains 148-bp direct terminal repeats.Multidrug-resistant  Serratia marcescens strains cause serious nosocomial infections in humans. Here, we present the annotated genome sequence of S. marcescens podophage Pila. Similar to its closest relative, Enterobacteria phage T7, Pila has a 38,678-bp genome, predicted to encode 51 protein-coding genes, and contains 148-bp direct terminal repeats.Multidrug-resistant  Serratia marcescens, a facultative, Gram-negative bacterium of the family Enterobacteriaceae, causes nosocomial infections in severely immunocompromised or critically ill patients  aerobically at 30\u00b0C and 37\u00b0C in LB broth and agar (BD). This phage was plaque purified, via the soft-agar overlay method (www.bioinformatics.babraham.ac.uk/projects/fastqc) was used for quality control of the 4,413 total reads in the phage-containing index, and reads were trimmed with the FastX Toolkit v0.0.14 (http://hannonlab.cshl.edu/fastx_toolkit). SPAdes v3.5.0 was used with default parameters to assemble a single contig at 14.9-fold coverage , using default parameters unless otherwise stated . PhageTerm analysis predicted the genomic termini to be 148-bp direct terminal repeats, typical for T7-like phages .The 38,678-bp double-stranded DNA genome of podophage Pila has a G+C content of 48.8%. The 51 predicted protein-coding genes correspond to a coding density of 92.5%. Notably, Pila shares 83.7% nucleotide identity with e phages , which iMN098329, BioProject accession no. PRJNA222858, SRA accession no. SRR8892142, and BioSample accession no. SAMN11408657.The genome sequence and associated data for phage Pila were deposited under GenBank accession no."}
+{"text": "Its\u2019 LOD is more than 74 times lower than the traditional Cu@Ni CSNPs modified working electrode. It also has higher sensitivity and wider linear range. This indicates the great potential of applying this kind of nano composites in electrode modification.In present work, a highly sensitive biosensor with high selectivity for glucose monitoring is developed based on novel nano-composites of nitrogen doped graphene quantum dots (N-GQDs) and a novel bimetallic Cu/Ni core-shell nanoparticles (CSNPs) (Cu@Ni CSNPs/N-GQDs NCs). With the tuned electronic properties, N-GQDs helped bimetallic core-shell structure nanomaterials from aggregation, and separate the charges generated at the interface. This novel nano-composites also have the good electrical conductivity of N-GQDs, catalyst property of Cu/Ni bimetallic nano composite, Cu@Ni core-shell structure and the synergistic effect of the interaction between bimetallic nano composite and N-GQDs. While modified the electrode with this novel nano-composites, the sensor\u2019 linear range is 0.09 ~ 1 mM, and the limit of detection (LOD) is 1.5 \u03bcM (S/N = 3) with a high sensitivity of 660 \u03bcA mM With the rapid development of nanotechnology, a lot of carbon materials have emerged in recent years, such as fullerenes, multi-wall carbon nanotubes, carbon nanofibers, graphene, graphenOwing to these properties, GQDs have gained wide attention for their enormous potential in varies applications. For instance they are widely applied in photovoltaics, organic light emitting diodes, fuel cells, photocatalysis, bioimaging, biosensing, biomedicine, environmental monitoring, thermal interface materials etc\u20138. Biose2 NTs[core-shellnanostructure of Ni/Cu bimetallic have attracted researchers\u2019 attention for glucose sensing[On the other hand, various nano-structured metals, alloys and metal oxides have been extensively applied in non-enzymatic electrode modification, due to their increased specific surface area, rapid mass transport, significant catalytic activity and so on\u201322. Amon2 NTs, Ni/Cu/M2 NTs are all 2 NTs\u201339. On te sensing.-1 cm-2. Comparing with the sensor based on Cu@Ni CSNPs modified working electrode, the electrode modified with Cu@Ni CSNPs/ N-GQDs NCs makes the sensor\u2019s LOD more than 74 times lower, higher sensitivity and wider linear range. These indicate that well designed N-GQDs\u2019 composites have great application prospects in improving electron migration rate and electrocatalytic performance of electrode surface.In this work, the novel bimetallic Cu/Ni and N-GQDs nano-composites (Cu@Ni CSNPs/N-GQDs NCs) have been synthesized by hydrothermal method and a one-pot solvothermal method. With our gentle synthesis method, the size of N-GQDs can be uniform with the nitrogen content about 12.93%. The size of Cu @ni CSNPs/N-GQDs NCs is 30\u201380 nm, and N-GQDs is uniformly coated on its surface, with bimetallic Cu/Ni co-catalytic performance and good electrical conductivity. A series of non-enzymatic glucose sensors are constructed with Cu@Ni CSNPs/N-GQDs NCs modified glassy carbon electrode (Cu@Ni CSNPs/N-GQDs/GCE), Cu@Ni CSNPs/GCE and Cu@Ni CSNPs/N-GQDs/GCE. Their electrochemical properties and electrocatalytic activities are compared. It is excited that Cu@Ni CSNPs/N-GQDs/GCE shows the best electrocatalytic performance for glucose oxidation, and displays the lowest detection limit(LOD) 1.5 \u03bcM (S/N = 3), a wider linear range from 0.09 mM to 1 mM and high sensitivity 660 \u03bcA mM2\u00b72H2O, 99.0%), Nickel (II) chloride hexahydrate , NaOH (96.0%), AA (\u226599.7%), sucrose, glucose and NaCl (\u226599.5%) were purchased from XiLong Scientific. Citric acid (99.5%) and DA (98%) were purchased from Aladdin. 5wt% of Nafion was purchased from Sigma Aldrich. Ethylene glycol , maltose, fructose, D-galactose and UA (99%) were purchased from Sinopharm Chemical Reagent Co., Ltd. Ultrapure water  was used throughout the experiments, and all reagents which were not mentioned above were of analytical grade.Urea (\u226599.0%), N, N-dimethyl formamide , Copper (II) chloride dihydrate (CuCl2 20), ultraviolet-visible spectra , Fourier transform infrared spectra  and X-ray diffraction measurement . All electrochemical experiments were performed on the electrochemical workstation . The working electrodes were modified glassy carbon electrode , the reference electrode was saturated calomel electrode (SCE), and a platinum wire electrode was usedas the auxiliary electrode.The samples were characterized by X-ray photoelectron spectra , transmission electron microscopy (TEM) and high resolution transmission electron microscope , Ipc = 6.1 \u00d710\u22123 v + 0.45125 (R2 = 0.99109), as shown in Ipa = -5.12 \u00d710\u22123 v\u20140.0932 (R2 = 0.99914), Ipc = 2.56 \u00d710\u22123 v + 0.06168 (R2 = 0.99952), as shown in 3+ and Ni2+ species[The modified Cu@Ni CSNPs/Nafion/GCE and Cu@Ni CSNPs/N-GQDs/Nafion/GCE electrodes were placed in cell containing 0.1 M NaOH solution to be tested at different scan rates. As shown in + species,42.-1. In Ipa = -0.00127 c\u20142.12802 (R2 = 0.95372), Ipc = 2.34641 \u00d710\u22125 c + 0.59356 (R2 = 0.63127). In Ipa = - 8.9 \u00d710\u22124 c\u20140.9307 (R2 = 0.99045), Ipc = -1.8 \u00d710\u22124 c + 0.3803 (R2 = 0.99435). Compared with the above two sets of experiments, Cu@Ni CSNPs/N-GQDs/Nafion/GCE has better linearity and better determination of glucose concentration.CVs was used to study the relationship between current and glucose concentration. The CVs was obtained by placing modified electrodes in solutions containing different glucose concentrations at scan rate of 100 mV si-t curve test.The CVs of modified Cu@Ni CSNPs/N-GQDs/Nafion/GCE electrode at different scan rates in 0.1 M NaOH containing 10 \u03bcM glucose is shown in i-t curve of Cu@Ni CSNPs/Nafion/GCE at different glucose concentrations. The linear relationship between current and glucose concentration is shown in i = 0.0853 c\u20140.00018, R = 0.99973). The detection limit of glucose using Cu@Ni CSNPs/Nafion/GCE is found to be 111.4 \u03bcM (S/N = 3) with the sensitivity of 85.3 \u03bcA mM-1 cm-2. As shown in i-t curve of Cu@Ni CSNPs/N-GQDs/Nafion/GCE at different glucose concentrations. The linear relationship between current and glucose concentration is shown in i = 0.66 c + 0.125, R = 0.99952). The detection limit of glucose using Cu@Ni CSNPs/N-GQDs/Nafion/GCE is found to be 1.5 \u03bcM (S/N = 3) with the sensitivity of 660 \u03bcA mM-1 cm-2. From this, it can be clearly concluded that Cu@Ni CSNPs/N-GQDs/Nafion/GCE is more sensitive to glucose determination. This is because GQDs itself is conductive, and N-GQDs has more hole electron pairs due to nitrogen atoms have been successfully doped, which greatly improve the electrical conductivity. Glucose is catalyzed by Cu and Ni in the composites, and N-GQDs are coated on the surface of Cu@Ni CSNPs, which improves the electron mobility between the electrode and the electrolyte, so that the working electrode to detect glucose within a very short time, which greatly improves the sensitivity of the electrode and reduces the detection limit, providing conditions for real-time, rapid and accurate determination of glucose concentration.Under the optimal conditions, +0.6 V is selected as the constant potential in the range of anodic potential of +0.5 ~ 0.7 V. In The anti-interference property and selectivity for glucose determination are crucial in the development of glucose biosensors. Chemical species such as uric acid (UA), dopamine (DA), ascorbic acid (AA), and NaCl that easily oxidize are always present with glucose in human blood. In this study, interference experiments were detected by adding 0.1 mM interference component to a 0.1 M NaOH solution containing 0.5 mM glucose. As shown in -1 cm-2, rapid response time (3 s). Comparing with the sensor based on Cu@Ni CSNPs modified working electrode, this novel nano-composite of Cu@Ni CSNPs/ N-GQDs NCs makes the sensor\u2019s LOD more than 74 times lower, also has higher sensitivity and wider linear range. Furthermore, the interference components show insignificant interference in determination of glucose, and Cu@Ni CSNPs/N-GQDs/Nafion/GCE has high selectivity for glucose determination. The results indicate that the biosensor based on Cu@Ni CSNPs/N-GQDs/Nafion/GCE has potential application prospect in the determination of glucose, the application of GQDs in biosensor has a great prospect. It also indicates the great potential to apply this kind of nano composites in electrode modification and high sensitivity biomolecule detection.In summary, Cu@Ni CSNPs and Cu@Ni CSNPs/N-GQDs NCs were successfully synthesized by one-pot solvothermal method. Our experiments show that the electrochemical response of Cu@Ni CSNPs/N-GQDs/Nafion/GCE to glucose determination was the highest. Cu@Ni CSNPs/N-GQDs/Nafion/GCE has the advantages of low cost, high sensitivity and good selectivity. One more advantage of Cu@Ni CSNPs/N-GQDs/Nafion/GCE is its\u2019 wide linear range is 0.09 ~ 1 mM. And the detection limit is 1.5 \u03bcM (S/N = 3), high sensitivity of 660 \u03bcA mMS1 Table(TIF)Click here for additional data file.S1 FigA) full spectrum, (B) C1s spectrum, (C) N1s spectrum, (D) O1s spectrum.((TIF)Click here for additional data file.S2 FigA) C1s spectrum, (B) Cu2p spectrum, (C) Ni2p spectrum.((TIF)Click here for additional data file.S3 FigMapping of Cu@Ni CSNPs/N-GQDs NCs (a), mapping-Cu (b), mapping-Ni (c).(TIF)Click here for additional data file.S4 FigA) and the histogram (B) of the Cu@Ni CSNPs/N-GQDs/GCE with successive addition of 0.5 mM glucose, 0.1 mM DA, 0.1 mM AA, 0.1 mM UA, 0.1 mM NaCl and 0.5 mM glucose in 0.1 M NaOH solution at +0.6 V, respectively.Amperometric response ((TIF)Click here for additional data file.S5 FigA) and the histogram (B) of the Cu@Ni CSNPs/N-GQDs/GCE with successive addition of 0.5 mM glucose, 0.1 mM maltose, 0.1 mM sucrose, 0.1 mM fructose, 0.1 mM D-galactose, 0.5 mM glucose in 0.1 M NaOH solution at +0.6 V, respectively.Amperometric response ((TIF)Click here for additional data file."}
+{"text": "Testing sampling effort and relative abundance descriptors of belowground ectomycorrhizal fungi in a UK planted scots pine woodland. Mycology. Agerer R, Ammirati J, Blanz P, Courtecuisse R, Desjardin DE, Gams W, Hallenberg N, Halling R, Hawksworth DL, Horak E, Korf RP, Mueller GM, Oberwinkler F, Rambold, G, Summerbell RC, Tribel D and Watling R. 2000. Open letter to the scientific community of mycologists. Canadian Journal of Botany. 78:981\u2013983Agerer R. 1991. Characterization of ectomycorrhiza. In: Norris JR, Read DJ, Varma AK, editors. Techniques for the study of mycorrhiza. Methods in microbiology. 23:25\u201373.Palmer MW. 1991. Estimating species richness: the second order jackknife reconsidered. Ecology. 72:1512\u20131513.Erland S and Taylor AFS. 2002. Diversity of ecto-mycorrhizal fungal communities in relation to the abiotic environment. In: van der Heiden MGA and Sanders IR, editors. Mycorrhizal ecology. Ecological Studies. Berlin: Springer; p. 163\u2013200.When the above article was first published online, the following references were omitted by mistake:The caption for Figure 2 incorrectly read:\u201c(a) Illustration of transect line (blue dashed line) and replicated sampling points (red dots). (b) Field sampling procedure in stand 8. (1) transect line, (2) soil samples in a bucket, (3) soil corer. (c) Detail of soil sample (1) in the soil corer (2) (3) Ruler 15 cm.\u2019\u2019This has now been corrected to:\u201c(a) Illustration of transect line (dashed line) and replicated sampling points (dots). (b) Field sampling procedure in stand 8. (1) transect line, (2) soil samples in a bucket, (3) split soil core sampler. (c) Detail of soil sample (1) in the split soil core sampler (2) (3) Ruler 15 cm.\u2019\u2019The caption for figure 5 incorrectly read:\u201cResult of regression analysis: r2 = 0.676**, p < 0.01**.\u2019\u2019This has now been corrected to:\u201cResult of regression analysis: r2 = 0.676, p < 0.01.\u2019\u2019In Table 1 the reference to Agerer, mistakenly listed the year as 2006 in three places, this has now been corrected to Agerer 1987\u20132002.These items have now been corrected in both the print and online versions of the journal.Taylor and Francis apologises for these errors."}
+{"text": "This article has been corrected: During the assembly of Figure 2B, the same image was inadvertently used for both H1299 (pEGFP-C3) and H1299 (pEGFP-4.1N) groups at 0h time point. The proper Figure 2 B specific for H1299 (pEGFP-C3) and H1299 (pEGFP-4.1N) groups are shown below. The authors declare that these corrections do not change the results or conclusions of this paper.509-523. https://doi.org/10.18632/oncotarget.6312Original article: Oncotarget. 2016; 7:509\u2013523."}
+{"text": "Butia ordorata) and fishtail palm (Caryota uren), using liquid chromatography electrospray ionization quadrupole time-of-flight mass spectrometry (LC-ESI-QTOF/MS) and assess their antioxidant potential. The total phenolic content (TPC), total tannins content (TTC), 2,2-diphenyl-1-picrylhydrazyl (DPPH) antioxidant assay and 2,2\u2032-azinobis-(3-ethylbenzo-thiazoline-6-sulfonic acid) (ABTS) scavenging abilities and ferric reducing antioxidant power (FRAP) were higher in the jelly palm fruit while total flavonoid contents (TFC) were higher in the fishtail palm. The LC-ESI-QTOF/MS tentatively identified a total of 86 phenolic compounds in both jelly and fishtail palm fruits. Although both palm fruits exhibited different phenolic profiles, hydroxycinnamic acids and flavonols were the most common in both. In high performance liquid chromatography photodiode array (HPLC-PDA) quantification, 4-hydroxybenzoic acid (317.46 \u00b1 4.68 \u00b5g/g) and catechin (4724.00 \u00b1 32.39 \u00b5g/g) were the most abundant phenolic acid and flavonoid quantified in the jelly palm fruit, respectively. Quercetin (557.28 \u00b1 7.81 \u00b5g/g) and kaempferol 3-O-glucoside (220.99 \u00b1 2.06 \u00b5g/g) were the most abundant flavonoids quantified in the fishtail palm. Our study indicates that palm fruit is a good source of polyphenols and has strong antioxidant potential for health promotion. Furthermore, this study provides the scientific basis for an exploitation of jelly and fishtail palm fruits in the food, pharmaceutical and nutraceutical industries. Palm fruits have gained growing attention for their nutrition values and health promotion perspectives. They have a diverse range of bioactive compounds including carotenoids, vitamins, dietary fibres and especially polyphenolic compounds. These polyphenolic compounds contribute to the putative health benefits of palm fruits. Nevertheless, the detailed information about these polyphenols in palm fruits is limited. The present work was conducted to comprehensively characterize polyphenols in two palm fruits, jelly palm ( Palm fruits are mostly grown in tropical regions and are rich sources of bioactive compounds such as polyphenols, vitamin E, carotenoids and unsaturated fatty acids . BioactiButia ordorata) is a tropical palm species whose fruits and leaves can be processed into different food products +m/z 339.1081) and 3-O-methylrosmarinic acid ([M + H]+m/z 375.1069) were tentatively characterized in fishtail palm. Hoffman et al. [p-coumaroylquinic acid (p-coumaric acid) in the methanolic extract of jelly palm pulp and nectar.Compound 2 and 5 (retention time (RT) = 19.734 and 37.691 min) with [M \u2212 H]ectively , while c355.1017 . There wion mode , while flm fruit . Sinapinly palms . The ferly palms . The prely palms ,29,30. C+ at m/z 139.0394 and 4-hydroxybenzoic acid 4-O-glucoside with RT = 7.456 min and [M \u2212 H]\u2212 at m/z 299.0793 in extracts of jelly and fishtail palm, respectively (m/z 137.0242) and hydroxybenzoic hexose (m/z 299.0070) have already been characterized in jelly palm by Hoffmann, Carvalho, Barbieri, Rombaldi and Chaves + at m/z 167.0687 and 227.0916 were assigned as 3-hydroxyphenylpropionic acid and dihydrosinapic acid, while 3-hydroxy-3-(3-hydroxyphenyl) propionic acid was tentatively identified by a molecular ion at m/z 181.0513 in negative ionization mode. The hydroxyphenylpentanoic acid compounds conjugated with valerolactone were proposed for both palm fruits  and 5--\u03b3-valerolactone ([M \u2212 H]\u2212m/z 207.0667) were detected  and 23  detected in ethanolic extract of fishtail palm fruits were tentatively identified as kaempferol 3-O-glucosyl-rhamnosyl-galactoside and kaempferol 3-O-glucoside, respectively, in negative ionization mode (O-glucoside (14) tentatively identified by the parent ion at m/z 449.1072 in ESI+ mode , quercetin 3-O-rutinoside (16), and quercetin (17) exhibiting precursor ion at m/z 465.1008, 611.1579 and 303.0486 in positive ionization mode were proposed for jelly palm fruit (O-glucoside (RT = 37.133 min) giving precursor ion at m/z 463.0878 in ESI\u2212 mode . The detection of aglycone quercetin in the jelly palm was reported previously d Chaves  also cha studies . Compounm/z 595.1634 and 595.163, respectively, was tentatively characterized in extracts of jelly and fishtail palm fruits. Compound 21  in the jelly palm and compound 32  in the fishtail palm were tentatively identified as apigenin 7-O-apiosyl-glucoside and apigenin 6-C-glucoside, respectively. The aglycone apigenin has been identified in different solvent extracts of jelly palm [\u2212 ion at m/z 461.1093 and [M + H]+ ion at m/z 549.1220, were proposed as chrysoeriol 7-O-glucoside and chrysoeriol 7-O-, respectively  and fishtail palm fruit , respectively. The aglycone hesperetin displaying [M \u2212 H]\u2212 ion at m/z 301.0707 has been proposed for jelly palm fruits in the study of Hoffmann, Carvalho, Barbieri, Rombaldi and Chaves + ion at m/z 481.1340 was proposed as 3\u2019-O-methyl-(-)-epicatechin 7-O-glucuronide. Catechin has also been detected in ethanolic extracts of jelly palm in a study reported by Beskow, Hoffmann, Teixeira, Fachinello, Chaves and Rombaldi Rombaldi . ProcyanO-sambubioside-5-O-glucoside (RT = 25.648 min) and peonidin 3-O-diglucoside-5-O-glucoside (RT = 26.857 min) exhibiting precursor ions at m/z 756.2092 and 786.2195 were detected in jelly palm (O-sophoroside (RT = 44.148 min) with parent ions at m/z 624.1664 was tentatively characterized in fishtail palm (m/z 772.2033 and 564.1457 were assigned as cyanidin 3-O-diglucoside-5-O-glucoside (29) and pelargonidin 3-O-sambubioside (32), respectively. In fishtail palm fruit, compound 40 exhibiting precursor ion at m/z 520.1225 was proposed as petunidin 3-O-(6\u2019\u2019-acetyl-glucoside). Previously, two cyanidin derivative anthocyanins were characterized in jelly palm fruit + ion at m/z 151.1108 was detected in fishtail palm. Pyrogallol was the only compound belonging to other polyphenols subclass detected in both palm fruits in the positive mode of ionization (Compound 54 (RT = 33.745 min) and 55 RT = 79.779 min) tentatively characterized in the extract of fishtail palm fruit with [M \u2212 H]opoletin . 3-Methynization . Previou.779 min The screening and characterization of polyphenolic compounds showed that some of the polyphenols presented in two palm fruits have strong antioxidant potential. Hydroxycinnamic acid derivatives, hydroxybenzoic acids and their derivatives, protocatechuic acid, chlorogenic acid, catechin, matairesinol, hydroxytyrosol, quercetin and kaempferol derivatives are regarded as potential compounds showing considerable free radical scavenging capacity ,38,39. TO-glucoside, catechin, quercetin and kaempferol 3-O-glucoside) based on the LC-ESI-QTOF/MS characterization and previously reported antioxidant activities. The quantification of individual polyphenols was computed considering the UV\u2013Vis absorption and calibration curves of external standards.Seven polyphenols were targeted to quantify through HPLC-PDA including three phenolic acids  and four flavonoids (quercetin 3-fw) was higher than fishtail palm fruit (140.23 \u00b1 1.78 \u00b5g/g fw). Boeing et al. [fw) was only detected in jelly palm fruit, while protocatechuic acid  was only found in fishtail palm. Beskow, Hoffmann, Teixeira, Fachinello, Chaves and Rombaldi [The polyphenols quantified through HPLC-PDA in the jelly palm were higher than in fishtail palm. These HPLC results support our TPC values determined using the Folin\u2013Ciocalteu method. Among the three selected phenolic acids, chlorogenic acid was the only phenolic acid detected in both palm fruits . The cong et al.  already fw in jelly palm and 557.28 \u00b1 7.81 \u00b5g/g fw in fishtail palm fruit. Beskow, Hoffmann, Teixeira, Fachinello, Chaves and Rombaldi [fw), which might be involved in higher antioxidant activity.Flavonols were found to be higher in fishtail palm fruit as compared to jelly palm fruits. Quercetin was the major compound in this group, with a concentration of 360.19 \u00b1 4.53 \u00b5g/g It has been established in this work that LC-ESI-QTOF/MS is an effective and powerful analytical tool to characterize most of the polyphenols present in jelly and fishtail palm fruits. The TPC, TTC, DPPH, FRAP and ABTS scavenging activity was higher in jelly palm compared to fishtail palm. LC-ESI-QTOF/MS characterizes a total of 42 and 60 phenolic compounds in jelly and fishtail palm fruits, respectively. Hydroxycinnamic acids and flavonols were the most common polyphenols reported in both palm fruits. The HPLC-PDA enabled the quantification of some targeted phenolic compounds and found that catechin and quercetin were the most abundant polyphenols in jelly and fishtail palm, respectively. In short, both palm fruits are a good source of polyphenols and could be utilized in food, feed and pharmaceutical industries."}
+{"text": "P < 0.001, RFS; P < 0.001), cytoplasmic expression of FAM83H , nuclear expression of PANX2 , cytoplasmic expression of PANX2 , co-expression pattern of nuclear FAM83H and nuclear PANX2 . In multivariate analysis, nuclear expression of FAM83H  and the co-expression pattern of nuclear FAM83H and PANX2  were independent indicators of shorter survival of CCRCC patients. Cytoplasmic expression of FAM83H was associated with shorter RFS (P = 0.030) in multivariate analysis. In Caki-1 and Caki-2 CCRCC cells, knock-down of FAM83H decreased PANX2 expression and cell proliferation, and overexpression of FAM83H increased PANX2 expression and cell proliferation. These results suggest that FAM83H and PANX2 might be involved in the progression of CCRCC in a co-operative manner, and their expression might be used as novel prognostic indicators for CCRCC patients.FAM83H is primarily known for its role in amelogenesis; however, recent reports suggest FAM83H might be involved in tumorigenesis. Although the studies of FAM83H in kidney cancer are limited, a search of the public database shows a significant association between FAM83H and pannexin-2 (PANX2) in clear cell renal cell carcinomas (CCRCCs). Therefore, we evaluated the clinicopathological significance of the immunohistochemical expression of FAM83H and PANX2 in 199 CCRCC patients. The expression of FAM83H and PANX2 were significantly associated with each other. In univariate analysis, individual, and co-expression pattern of FAM83H and PANX2 was significantly associated with shorter overall survival (OS) and relapse-free survival (RFS) of CCRCC patients: nuclear expression of FAM83H (OS;  Family with sequence similarity 83 H (FAM83H) is one of the eight FAM83 family members, from FAM83A to FAM83H , 2. Accohttp://www.oncolnc.org). In addition, the cBioPortal database  indicated pannexin-2 (PANX2) as the gene most significantly associated with FAM83H expression in CCRCCs   is the most common type of kidney cancer . Most CCn CCRCCs , 12. Altst 2018) , 12. PANst 2018) . Howeverst 2018) . TherefoThis study evaluated CCRCC patients who underwent surgical resection between July 1998 and August 2011. Among them, 199 CCRCCs with complete medical records were available with original H&E slides, and paraffin-embedded tissue blocks. Clinical information was obtained by reviewing the medical records. The histopathologic information was retrospectively reviewed according to the 2016 World Health Organization classification of the renal tumors , and staIn this study, tissue microarrays were used for immunohistochemical staining. The tissue microarrays have 3.0 mm cores, and one core was arrayed per case from the areas composed mainly of tumor cells with no degenerative change or necrosis. The histologic sections from the tissue microarray tissue blocks were deparaffinized and boiled for 20 min in pH 6.0 antigen retrieval solution  in a microwave oven. Primary antibodies for FAM83H  and PANX2  was used. The slides were visualized with the enzyme substrate 3-amino-9-ethylcarbazole and counterstained with hematoxylin. The scoring for the immunohistochemical staining slides was performed without clinical information by two pathologists (KYJ and KMK) with consensus under a multi-viewing microscope. The immunohistochemical positivity for FAM83H and PANX2 was separately scored according to their expression patterns in the cytoplasm and nuclei. The immunohistochemical scores were obtained by adding their staining intensity score ranging from zero to three  and staining area score ranging from zero to five  , 19. The2 and 37\u00b0C.In this study, we used two human CCRCC cell lines. Caki-1 and Caki-2 cells were purchased from the Korean Cell Line Bank . The Caki-1 and Caki-2 cells were cultured in DMEM medium with 10% fetal bovine serum  and streptomycin and penicillin (100 U/ml) in 5% COThe vector of shRNA for FAM83H was purchased from GenePharma . The sense and antisense sequences of the FAM83H shRNA were the 5\u2032-CAC CGC TCA TCT TCA GCA CGT CAC ATT CAA GAG ATG TGA CGT GCT GAA GAT GAG CTT TTT TG-3\u2032 and 5\u2032-GAT CCA AAA AAG CTC ATC TTC AGC ACG TCA CAT CTC TTG AAT GTG ACG TGC TGA AGA TGA GC-3\u2032, respectively. The vector for FAM83H overexpression  was purchased from GeneCopoeia . JetPRIME transfection reagent  was used for transfection.To isolate protein, the cells were lysed with PRO-PREP Protein Extraction Solution  with 1x phosphatase inhibitor cocktails 2, 3 (Sigma-Aldrich). The primary antibodies for FAM83H , PANX2 , and actin (Sigma-Aldrich) were used for western blots.RNA was isolated with an RNeasy Mini Kit . The isolated RNA was reverse transcribed by probing 1.5 \u03bcg RNA with Reverse Transcription kits  to generate cDNA that was used for quantitative polymerase chain reaction using Applied Biosystems Prism 7900HT sequence Detection System and SYBR Green Master PCR Mix (Applied Biosystems). The values were normalized to the expression of the glyceraldehyde-3-phosphate dehydrogenase reference housekeeping gene. All experiments were performed in triplicate. The sequences of the primers used in quantitative reverse-transcription polymerase chain reaction are listed in Table 5) and Caki-2 (2 \u00d7 105) cells were seeded in 24-well plates and the number of viable cells was counted by using hemocytometer. For the MTT assay, Caki-1 (3 \u00d7 103) and Caki-2 (3 \u00d7 103) cells were seeded in 96-well culture plates, and absorbance was measured at 560 nm using a microtiter plate reader .The proliferation of cells was evaluated by counting the number of cells and performing a 3--2,5-diphenyltetrazonium bromide (MTT)  assay and a colony-forming assay. To compare the number of cells in control, FAM83H knock-down, and FAM83H overexpression groups, Caki-1  was used throughout, and P < 0.05 were considered statistically significant.The cut-off points of immunohistochemical staining scores for FAM83H and PANX2 were determined by receiver operating characteristic curve analysis , 19. SurP < 0.001), Cy-FAM83H (P < 0.001), Nu-PANX2 (P < 0.001), and Cy-PANX2 (P = 0.002) was significantly associated with death of patients from CCRCC  and higher tumor stage (P = 0.001) (Table P = 0.004), larger tumor size (P < 0.001), higher tumor stage (P < 0.001), and higher histologic grade (P = 0.037) (Table P = 0.009), tumor size (P = 0.001), tumor stage (P < 0.001), and tumor necrosis (P = 0.003) (Table P = 0.032), tumor size (P < 0.001), tumor stage (P < 0.001), lymph node metastasis (P = 0.018), nuclear grade (P = 0.006), and tumor necrosis (P = 0.011) , tumor size (P < 0.001), tumor stage (P < 0.001), tumor necrosis (P = 0.006), Nu-FAM83H (P < 0.001), Cy-FAM83H (P < 0.001), Nu-PANX2 (P < 0.001), and Cy-PANX2 (P < 0.001) (Table P = 0.033), age (P = 0.022), tumor size (P < 0.001), tumor stage (P < 0.001), lymph node metastasis (P = 0.010), nuclear grade (P = 0.006), Nu-FAM83H (P < 0.001), Cy-FAM83H (P < 0.001), Nu-PANX2 (P < 0.001), and Cy-PANX2 (P < 0.001) , Nu-FAM83H , Cy-FAM83H , Nu-PANX2 , and Cy-PANX2 positivity  are presented in Figure Nuclear positivity of FAM83H expression had a 6.205-fold  greater risk of death and a 4.949-fold  greater risk of relapse or death of CCRCC patients. Cy-FAM83H positivity showed a 4.617-fold  greater risk of death and a 5.369-fold  greater risk of relapse or death of CCRCC patients (Table P = 0.047), tumor stage (P = 0.018), tumor necrosis (P = 0.031), and Nu-FAM83H positivity (P < 0.001) (Table P < 0.001), Nu-FAM83H positivity (P = 0.003), and Cy-FAM83H positivity (P = 0.030) were the factors significantly associated with RFS of CCRCC patients in multivariate analysis , intermediate (Nu-FAM83H+/Nu-PANX2\u2212), and poor prognostic (Nu-FAM83H+/Nu-PANX2+) groups that were significantly associated with OS  and RFS  in univariate analysis  greater risk of OS and 4.903-fold  greater risk of RFS compared with the Nu-FAM83H\u2212/Nu-PANX2\u2212 and Nu-FAM83H\u2212/Nu-PANX2+ subgroups  greater risk of lower OS and a 160%  greater risk of lower RFS compared with the Nu-FAM83H\u2212/Nu-PANX2\u2212 and Nu-FAM83H\u2212/Nu-PANX2+ subgroups , higher expression of FAM83H RNA was associated with shorter OS of CCRCC   predicted shorter survival of hepatocellular carcinoma patients . In a reConcerning the mechanism how FAM83H is involved in tumorigenesis, the role of FAM83H in the proliferation of cancer cells has been suggested in prostatic cancer cells . In addihttps://www.proteinatlas.org) also reported FAM83H expression to be in the cytoplasmic membrane and cytoplasm. In colorectal cancer cells, FAM83H expression was localized in the cytoplasm in association with keratin cytoskeleton structures   , 12, theP = 0.004) and RFS (P = 0.015). The patients with Nu-PANX2-positive CCRCC had a 3.186-fold  greater risk of cancer-related death and a 2.302-fold  greater risk of relapse or cancer-related death compared with the patients with Nu-PANX2-negative CCRCC. Therefore, despite limited reports on the role of PANX2 in human cancers, our results suggest PANX2 as a potential biologic marker of CCRCC. In addition, when we searched the OncoLnc database (http://www.oncolnc.org), higher expression of PANX2 RNA was associated with shorter OS of CCRCC patients  greater risk of death compared to CCRCC patients with low expression of PANX2 RNA. However, in contrast, PANX2 inhibited proliferation and tumor formation of C6 glioma cells in in vitro and in vivo   , 12. Morn; 0.36) . In addiIn conclusion, this study presents the roles for FAM83H and PANX2 in CCRCCs and suggests that FAM83H and PANX2 are closely associated and involved in the progression of CCRCCs. Especially, the expression patterns of FAM83H and PANX2 were significantly associated with shorter survival of CCRCC patients. In addition, the co-expression patterns of FAM83H and PANX2 were also significantly associated with the survival of CCRCC patients. Therefore, individual and co-expression patterns of FAM83H and PANX2 might be useful prognostic indicators for CCRCC patients. Furthermore, understanding for the role of FAM83H in conjunction with PANX2 might be helpful in establishing a new therapeutic strategy for CCRCCs.KMK, UH, JB, S-HP, KSK, SH, HP, HL, MC, WM, MK, and KJ participated in the study design. KMK, UH, and JB performed the experiment. KMK, UH, JB, S-HP, KSK, SH, HP, HL, MC, WM, MK, and KJ were involved in data collection and data interpretation. KMK, UH, JB, S-HP, KSK, SH, HP, MC, WM, and KJ participated in the statistical analyses. KMK, UH, JB, S-HP, KSK, SH, HP, HL, MC, WM, MK, and KJ wrote the manuscript. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "The correct citation is: Bispo PC, Balzter H, Malhi Y, Slik JWF, Santos JR, Renn\u00f3 CD, et al. (2017) Drivers of metacommunity structure diverge for common and rare Amazonian tree species. PLoS ONE 12(11): e0188300. In the Funding section, the author Fernando D. Esp\u00edrito-Santo\u2019s initials are listed incorrectly. The sentence should read: FDES was supported by Natural Environment Research Council (NERC) grants (BIO-RED NE/N012542/1 and AFIRE NE/P004512/1) and Newton Fund (The UK Academies/FAPESP Proc. N\u00b0: 2015/50392-8 Fellowship and Research Mobility)."}
+{"text": "The family environment is often overlooked in caregiver research and assessment, despite having implications for caregiver health and well-being . The purpose of the present study was to examine differences on two types of family conflict (beliefs and support) among a diverse sample of caregivers. The present sample consisted of help-seeking (n = 375) and non-help-seeking (n = 415) caregivers . Caregivers filled out the Caregiver Reaction Scale , a multidimensional assessment of the caregiver experience. Results of a 2  x 2  ANOVA indicated that help-seeking caregivers reported significantly more conflict over family beliefs than did non-help-seeking caregivers , F = 21.10 p < .001. Adult children caregivers reported significantly greater conflict over family beliefs (M = 1.91) than did spouse caregivers (M = 1.60), F = 10.66, p < .001. Adult children caregivers also reported significantly greater conflict over family support (M = 1.87) than did spouse caregivers (M = 1.57), F = 16.23, p < .001. Results highlight that certain caregiving contexts  potentially increase family conflict, which has implications for caregiver burden. Family conflict over beliefs is also related to help-seeking in caregivers. Findings inform appropriate assessment and intervention regarding the family environment in caregiving."}